determination of iodine content...

DETERMINATION OF IODINE CONTENT IN LOCALLY AVAILABLE FOODS USING

SPECTROPHOTOMETRIC KINETIC METHOD

by

Ajenesh Chandra

A thesis submitted in fulfilment of the requirements for the

Degree of Master of Science in Chemistry

Copyright © 2019 by Ajenesh Chandra

School of Biological and Chemical Sciences

Faculty of Science, Technology and Environment

The University of the South Pacific

January, 2019

STATEMENT BY AUTHOR AND SUPERVISORS

Statement by author

I, Ajenesh Chandra, declare that this thesis is my own work and that, to the best of my

knowledge, it contains no material previously published, or substantially overlapping

with material submitted for the award of any other degree at any institution, except

where due acknowledgment is made in the text.

Signature Date 2 / 1/201

Name: Ajenesh Chandra

Student ID No. S11022698

Statement by supervisor

The research in this thesis was performed under my supervision and to my

knowledge is the sole work of Ajenesh Chandra.

Signature Date 2 / /201

Principal Supervisor: Professor Surendra Prasad

Professor of Chemistry

SBCS, FSTE, USP, Suva, Fiji

Signature Date 2 / /201

Co-supervisor: Dr. Matakite Maata

Senior Lecturer in Chemistry

SBCS, FSTE, USP, Suva, Fiji

i

DEDICATION

I would like to dedicate my work to my parents especially my dad the late Mr.

Suresh Chandra who passed away during the course of this research, all my family

and my wife Bijeta Chandra.

ii

ACKNOWLEDGEMENT

This Master of Science thesis has been carried out at the School of Biological and

Chemical Sciences, the University of the South Pacific, Suva, Fiji. I would like to

acknowledge the following people for their support and help and without whom this

project would not be successful. Therefore, I express my gratitude to them all.

This MSc thesis work was carried out under the able guidance of my Principal

Supervisor, Professor Surendra Prasad. A special thanks to him for all his help,

support, advice, encouragement, criticism, comments and time for this entire project.

My sincere thanks to Dr. Matakite Maata for being Co-supervisor.

A special thanks to the FSTE Research and Graduate Affairs team for approving the

funding for this research from the FSTE research funds.

My appreciation also goes to the Chief Technician, Mr. Steve Sutcliffe and Senior

Technician, Mr. Shelvin Prasad for helping and guiding me on the procurement of the

required chemicals and the support and help throughout the project. In particular, I

wish to thank the academic and technical staff of the Chemistry discipline, who have

assisted me in various ways. Special thanks to the technicians Timaima Waqainabete,

Joslin Lal and Thomas Tunidau for going out of their ways to assist me in obtaining

necessary glassware, chemicals and apparatus on time.

I would also like to acknowledge my parents, the late Mr. Suresh Chandra and Mrs.

Suneela Chandra, my brother Atish Chandra, and his wife Jyotika, my two sisters

Ashika Maharaj and Ashmeeta Singh for their love and support throughout my life and

this study. I would also like to thank my late uncle, Mr. Ravin for his words of

encouragement for this project.

Finally, a special thanks goes to my wife Bijeta for the love, motivation, help and

encouragement throughout this research.

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ABSTRACT

Iodine is one of the essential trace elements that is important to human health and has

been of great interest in nutritional research studies. It is vital for the generation of

hormones in the thyroid. These hormones triiodothyronine (T3) and thyroxine (T4) are

needed for the proper functioning, growth and development of the human body. A

deficiency of iodine in the human body for prolonged periods will result in the thyroid

working harder to maintain the right amount of hormones in the blood and eventually

leads to goitre which is the enlarged thyroid or swelling of the thyroid gland. Iodine

deficiency in the human body is also linked to other health problems such as endemic

cretinism, infant mortality, infertility, miscarriage, mental retardation, neuromuscular

defects, and dwarfism. All these are known as Iodine Deficiency Disorders (IDDs).

IDDs are major health problems throughout the world, especially for young children

and pregnant women. Many studies have reported that, IDDs pose a threat to the social

and economic development of countries as well. Therefore, a knowledge of the daily

iodine intake (DII) as recommended by the World Health Organisation (WHO) is

important. Food being the major iodine source for the human body needs to be

carefully analysed for iodine contents so people can understand, know and plan their

DII. Thus there is every need for all citizens around the world to be knowledgeable of

iodine and especially for Fijians because of limited research and public awareness on

the topic.

The spectrophotometric kinetic method for iodine determination in food samples was

validated in this research based on the iodide catalysed reaction which involves the

reduction of Ce4+ to Ce3+ by As3+. The incineration of the organic matter was achieved

sulphate (ZnSO4) for 3 hours. The absorbance of the kinetic reaction was measured at

370 nm f

against the different iodine concentrations. A linear relationship was seen with the R2

value of 0.9998, which indicated a good reproducibility. Trace levels of iodine (ng)

were determined successfully using the above mentioned spectrophotometric kinetic

method for the 22 food samples (4 samples of each category consisting of 88 sub-

samples).

iv

The Fiji seaweeds, lumiwawa (brown seaweed) and sea grapes (green seaweed) gave

the highest levels of average iodine content of 6373.30 ± 0.39 ng/g and 1162.81 ± 0.61

ng/g, respectively followed by fresh seawater fish with an average iodine content of

1043.24 ± 0.75 ng/g. The decreasing trend of iodine content followed the order where

egg had 730.10 ± 0.47 ng/g, canned sardine 586.66 ± 0.40 ng/g, processed powdered

milk 580.04 ± 0.45 ng/g, canned tuna 536.92 ± 0.49 ng/g, clam 499.98 ± 0.48 ng/g,

cheese 377.57 ± 0.27 ng/g, dalo/taro 311.93 ± 0.28 ng/g, cassava 262.76 ± 0.19 ng/g,

potato 255.87 ± 0.27 ng/g, fresh liquid milk 237.70 ± 0.24 ng/g, butter/margarine

218.52 ± 0.20 ng/g, lettuce 114.81 ± 0.08 ng/g, English cabbage 108.40 ± 0.06 ng/g,

Chinese cabbage 104.01 ± 0.06 ng/g, pumpkin 101.24 ± 0.08 ng/g, rice 99.92 ± 0.11

ng/g, long bean 97.61 ± 0.10 ng/g, banana 76.18 ± 0.10 ng/g and tomato 40.32 ± 0.04

ng/g. Fresh food samples also showed higher iodine concentrations than the factory

processed foods. In addition, brown seaweeds (lumiwawa) had higher iodine content

than sea grapes (green seaweed).

The coefficient of variation for the sample analysis was less than 5.92 % with a mean

and standard deviation of 2.57 ± 0.28% for the 22 food samples (88 sub-samples), each

analysed four times. The limit of detection (LOD) was 1.54 ng/mL and the limit of

quantification (LOQ) was 4.90 ng/mL. The recovery of iodine added to different food

samples ranged from 97.42 ± 3.41% to 103.13 ± 4.76% with an average recovery of

100.18 ± 3.02% (mean ± standard deviation). The analytical coefficient of variation

was calculated to be 0.54% for the 22 food samples analysed. This shows exceptional

system analytical stability of the method used in this study.

v

LIST OF ABBREVIATIONS AND ACRONYMS

% - Percentage

~ - Approximately

× - Multiplied by

AAS - Atomic Absorption Spectrometer

ACS - American Chemical Society

ADHD - Attention deficit hyperactivity disorders

AOAC - Association of Official Analytical Chemists

AR - Analytical reagent

As - Arsenic

As3+ - Arsenic trioxide

b - Blank

Ce3+ - Reduced cerium oxide

Ce4+ - Ammonium cerium sulphate

Conc. - Concentration

d - Dilution

DII - Daily Iodine Intake

EU - European Union

FAO - Food and Agricultural Organization

FT-IR - Fourier Transform Infrared

GC - Gas Chromatography

GC-ECD - Gas Chromatography- Electron Capture Detector

H2SO4 - Sulfuric acid

HCl - Hydrochloric acid

Hg - Mercury

HNO3 - Nitric acid

HPLC - High Pressure Liquid Chromatography

hr - hour

I- - Iodide

I - Iodine

IAS - Institute of Applied Science

vi

ICCIDD - International Council for Control of Iodine Deficiency

Disorders

ICP-MS - Inductively Coupled Plasma Mass Spectrophotometry

IDDs - Iodine Deficiency Disorders

IEC - Ion Exchange Chromatography

IQ - Intelligence quotient

ISE - Ion selective electrodes

KI - Potassium iodide

KIO3 - Potassium iodate

KOH - Potassium hydroxide

LOD - Limit of detection

LOQ - Limit of quantification

m - Slope

Max - Maximum

Min - Minimum

n - Number

NaCl - Sodium chloride

R2 - Value of linear regression

RDI - Recommended daily intake

rpm - Revolutions per minute

RSD - Relative standard deviation

s - Sample

SAE - Standard analytical error

SD - Standard deviation

SRM - Standard reference material

t - Student’s t-value

T3 - Triiodothyronine

T4 - Thyroxine

TG - Thyroglobulin

TMAH - Tetramethylammonium hydroxide

TSH - Thyroid stimulating hormone

TXRF - Total Reflection X- Ray Fluorescence

UHT - Ultra heat treated

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UI - Urinary iodine

UNICEF - United Nations Children’s Fund

USA - United States of America

USFDA - United States Food and Drug Administration

USP - University of the South Pacific

UV - Ultraviolet

vs - Versus

WHO - World Health Organisation

ZnSO4 - Zinc Sulphate

- Change

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UNITS OF MEASUREMENTS

°C - Degrees Celsius

μg - Microgram

μg/d - Micrograms per day

μL - Microliter

μm - Micron

A - Absorbance

g - Grams

hr - Hour

L - Liter

mg - Milligram

min - Minute

mL - Milliliter

ng - Nanogram

ng/g - Nanogram per gram

nm - Nanometer

sec - Second

t - Time

W - Watts

w/v - Weight over volume ratio

/min - Change in absorbance per minute

- Wavelength

- Micro

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TABLE OF CONTENTS PAGES

DEDICATION .......................................................................................................... i

ACKNOWLEDGEMENT ........................................................................................ ii

ABSTRACT ............................................................................................................ iii

LIST OF ABBREVIATIONS AND ACRONYMS ................................................. v

UNITS OF MEASUREMENTS ........................................................................... viii

TABLE OF CONTENTS ........................................................................................ ix

LIST OF FIGURES .......................................................................................... xv

LIST OF TABLES ....................................................................................... xviii

CHAPTER 1 ........................................................................................................... 1

INTRODUCTION .................................................................................................. 1

1.1. General background .......................................................................................... 1

1.1.1. Iodine in water................................................................................................. 2

1.1.2. Iodine in air .................................................................................................... 2

1.1.3. Iodine in soil ................................................................................................... 2

1.1.4. Iodine in food and plants ................................................................................ 3

1.1.5. Iodine in human body ..................................................................................... 3

1.1.6. Iodine in thyroid ............................................................................................. 3

1.1.7. Iodine in plasma ............................................................................................. 4

1.1.8. Iodine in brain ................................................................................................ 5

1.1.9. Iodine in hair .................................................................................................. 5

x

1.1.10. Iodine in human milk.................................................................................... 5

1.1.11. Iodine in urine .............................................................................................. 6

1.2. Problems associated with low and excess iodine intake ................................... 6

1.3. Justification of the study and significance of research ...................................... 9

1.4. Aim. ................................................................................................................. 10

1.5. Objectives ........................................................................................................ 10

CHAPTER 2 ......................................................................................................... 11

LITERATURE REVIEW .................................................................................... 11

2.1. Introduction ..................................................................................................... 11

2.2. Global estimates .............................................................................................. 12

2.3. Recommended iodine intake ........................................................................... 14

2.4. Urinary iodine .................................................................................................. 15

2.5. Iodine in foods ................................................................................................. 15

2.5.1. Food fortification to prevent iodine deficiency ............................................ 18

2.5.2. Salt iodisation to prevent IDDs. ................................................................... 19

2.6. Analytical methods for iodine quantification .................................................. 20

2.6.1. Inductively coupled plasma-mass spectrophotometry (ICP-MS)................. 22

2.6.2. Inductively coupled plasma-optical emission spectrophotometry (ICP-

OES) ....................................................................................................................... 24

2.6.3. Neutron activation analysis (NAA) ............................................................... 24

2.6.4. Atomic absorption spectrometry (AAS) ....................................................... 25

2.6.5. Electrochemical and potentiometric probes ................................................. 26

xi

2.6.6. Gas, liquid and ion chromatographic methods ............................................. 27

2.6.7. UV-Visible spectrophotometry .................................................................... 28

2.7. Sample digestion ............................................................................................. 38

2.8. Sandell-Kolthoff (S-K) reaction for iodine determination .............................. 39

2.9. Drawbacks of the Sandell-Kolthoff reaction method in iodine determination 40

2.10. Conclusion ..................................................................................................... 41

CHAPTER 3 ......................................................................................................... 43

RESEARCH METHODOLOGY ........................................................................ 43

3.1. Chemical and reagents ..................................................................................... 43

3.2. Instrumentation ................................................................................................ 44

3.3. Standard calibration curves ............................................................................. 45

3.4. Food samples and sampling ............................................................................ 45

3.5. Sample storage and preparation....................................................................... 46

3.6. Ashing procedure ............................................................................................ 46

3.7. Sample analysis ............................................................................................... 48

3.8. Precision .......................................................................................................... 48

3.9. Limit of detection ............................................................................................ 49

3.10. Limit of quantification ................................................................................... 49

3.11. Quality control ............................................................................................... 49

3.11.1. Chemicals ................................................................................................... 50

3.11.2. Preparation of Millipore water ................................................................... 50

xii

3.11.3. Glassware ................................................................................................... 50

3.11.4. Data recording ............................................................................................ 51

3.11.5. Standard operating procedure of analysis................................................... 51

3.11.6. Analysis of duplicate samples .................................................................... 51

3.11.7. Analysis of blanks ...................................................................................... 51

3.11.8. Analysis of standard samples ..................................................................... 52

3.11.9. Standard calibration and linear equation .................................................... 52

3.11.10. Spike recoveries ....................................................................................... 52

3.11.11. Analysis of Standard Reference Materials (SRM) ................................... 52

3.11.12. Statistical analysis of data ........................................................................ 53

CHAPTER 4 ......................................................................................................... 54

RESULTS .............................................................................................................. 54

4.1. Analysis of blanks ........................................................................................... 54

4.2. Time-absorbance curves .................................................................................. 55

4.3. Calibration curves ............................................................................................ 59

4.4. Food samples ashing ....................................................................................... 61

4.5. Sample analysis ............................................................................................... 62

4.6. Precision .......................................................................................................... 74

4.7. Limit of detection ............................................................................................ 75

4.8. Limit of quantification ..................................................................................... 76

4.9. Quality control ................................................................................................. 76

xiii

4.9.1. Analysis of duplicate samples for food sample analysis .............................. 76

4.9.2. Recovery analysis from standard samples .................................................... 76

4.9.3. Spike recoveries from real samples .............................................................. 77

4.9.4. Analysis of standard reference materials (SRM).......................................... 79

CHAPTER 5 ......................................................................................................... 81

DISCUSSION ........................................................................................................ 81

5.1. Discussion of results obtained ......................................................................... 81

5.1.1. Cluster analysis ............................................................................................. 92

5.2. Comparison of iodine content with previous published data .......................... 93

CHAPTER 6 ....................................................................................................... 101

CONCLUSION AND RECOMMENDATIONS ............................................. 101

6.1. Conclusion ..................................................................................................... 101

6.2. Recommendations ......................................................................................... 103

6.2.1. Recommendations to the general public..................................................... 103

6.2.2. Recommendations for future study ............................................................ 104

REFERENCES ....................................................................................................... 105

APPENDICES ........................................................................................................ 117

Appendix 1: Average absorbance at different iodine concentrations for t = 0 min. 117

Appendix 2: Average absorbance at different iodine concentrations for t = 0.5

min. .......................................................................................................................... 118

Appendix 3: Average absorbance at different iodine concentrations for t = 1 min. 120

xiv

Appendix 4: Absorbance for the determination of iodine in standard iodine solutions

at 4, 12, and 18 ng/mL and their recovery. .............................................................. 122

Appendix 5: Absorbance for the determination of iodine in different food samples

analysed along with average iodine contents and coefficient of variation............... 123

Appendix 6: Absorbance for the recovery study along with the determined iodine

contents in some selected food samples by adding 4, 12 and 18 ng/mL iodine. ..... 145

Appendix 7: Determination (recovery) of iodine in NIST Standard Reference Material

(SRM No. 3530). ...................................................................................................... 155

xv

LIST OF FIGURES PAGES

Figure 1. Perkin Elmer Lambda 365 UV visible spectrophotometer equipped with 10

mm quartz cells (3) connected to a computer with the UV Express software (2), a

printer (4) together with a thermostatic water bath to control the temperature of the

reagents and reaction system ......................................................... 45

Figure 2. Muffle furnace with digital temperature controller (1) and temperature

ramping setting (1 and 2). .......................................................................................... 47

Figure 3. The Simplicity brand Millipore Milli-Q system used to obtain Millipore

water. .......................................................................................................................... 50

Figure 4. Typical absorbance time curve for the blank analysis up to 1 min............ 54

Figure 5. Typical absorbance – time recording of the catalysed reaction up to 1 min at

different iodine concentrations of 0, 2.5, 5, 10, 15, 20 and 25 ng/mL at 370 nm at 37

............................................................................................................................... 56

Figure 6. Plot of average absorbance at 370 nm for the reduction of Ce4+ by As3+

against time in the presence of different iodine concentrations of 0, 2.5, 5, 10, 15, 20

and 25 ng/mL at analysis time of ...................................................... 59

Figure 7. Calibration curve i.e. plot of average change in absorbance per minute

...................................................................... 60

Figure 8. Typical UV-visible recording of absorbance against time

at 370 nm for different food samples analysed. ......................................................... 63

Figure 9. Graphical representation of iodine contents in different brands of rice

analysed on a fresh weight basis. ............................................................................... 64

Figure 10. Graphical representation of iodine contents in different root crops analysed

on a fresh weight basis. .............................................................................................. 66

Figure 11. Graphical representation of iodine contents in different fish/meat products


xvi

Figure 12. Graphical representation of iodine contents in different dairy products


Figure 13. Graphical representation of iodine contents in commonly consumed leafy

vegetables analysed on a fresh weight basis. ............................................................. 71

Figure 14. Graphical representation of iodine contents in commonly consumed fruits

and vegetables analysed on a fresh weight basis. ...................................................... 73

Figure 15. Graphical representation of iodine contents in commonly consumed

seaweeds analysed on a fresh weight basis. ............................................................... 74

Figure 16. Typical UV-visible spectra of blank and NIST Standard Reference Material

(SRM No. 3530 - Iodised S .................... 79

Figure 17. Graphical representation of mean iodine contents in commonly consumed

rice and root crops analysed on a fresh weight basis. ................................................ 82


fish/meat analysed on a fresh weight basis. ............................................................... 84


dairy products analysed on a fresh weight basis. ....................................................... 85


leafy vegetables analysed on a fresh weight basis. .................................................... 87


fruits and vegetable analysed on a fresh weight basis................................................ 88


seaweeds analysed on a fresh weight basis. ............................................................... 89

Figure 23. Graphical representation of determined average iodine contents (ng/g) of

the analysed food samples .......................................................................................... 93

xvii

Figure 24. Dendrogram of cluster analysis (Ward’s method) of determined average

iodine contents (ng/g) of the analysed food samples……………………………………….94

xviii

LIST OF TABLES PAGES

Table 1. Daily iodine intake recommended by World Health Organisation. .............. 7

Table 2. Tolerable daily iodine intake by population in different countries of different

age groups. ................................................................................................................... 7

Table 3. Spectrum of iodine deficiency disorders (IDDs) for different age groups. ... 8

Table 4. Recommended daily intake of iodine in Australia and New Zealand in

different age groups.................................................................................................... 15

Table 5. Criteria for assessing iodine nutrition in groups based on median UI

concentrations. ........................................................................................................... 16

Table 6. Analytical methods for the determination of iodine in different samples. .. 30

Table 7. Change in absorbance in blank analysis (0 ng/mL- iodine). ....................... 55

Table 8. Absorbance analysis at different iodine concentrations and different

times. .......................................................................................................................... 57

Table 9. The average absorbances at different iodine concentrations from 0 to 25

ng/mL at three different times (n = 7). ....................................................................... 58

Table 10. ............................................ 60

Table 11. Iodine contents in different brands of rice (Oryza sativa) analysed on a fresh

weight basis. ............................................................................................................... 64

Table 12. Iodine contents in different root crops analysed on a fresh weight basis. . 65

Table 13. Iodine contents in different fish/meat products analysed on a fresh weight

basis. ........................................................................................................................... 66

Table 14. Iodine contents in different dairy products analysed on a fresh weight basis.

.................................................................................................................................... 69

xix

Table 15. Iodine contents in commonly consumed leafy vegetables analysed on a fresh

weight basis. ............................................................................................................... 70

Table 16. Iodine contents in commonly consumed fruits and vegetables analysed on a

fresh weight basis. .................................................................................................... 712

Table 17. Iodine contents in commonly consumed seaweeds analysed on a fresh

weight basis. ............................................................................................................... 73

Table 18. Change in the absorbance per minute for 7 runs (n = 7) for blank analysis (0

ng/mL –iodine), with standard deviation for LOD and LOQ determination... .......... 75

Table 19. Analysis of standard iodine solutions at 4, 12, and 18 ng/mL and their

recovery. ..................................................................................................................... 77

Table 20. Recovery of iodine from different food samples after spiking with 4, 12 and

18 ng/mL iodine. ........................................................................................................ 78

Table 21. Summary of the recovery results obtained from NIST SRM No. 3530 –

Iodised Salt analysis. .................................................................................................. 80

Table 22. NIST SRM No. 3530 iodine recovery using the spectrophotometric kinetic

method. ....................................................................................................................... 80

Table 23. Mean iodine contents in commonly consumed rice and root crops analysed


Table 24. Mean iodine contents in commonly consumed fish/meat products analysed


Table 25. Mean iodine contents in commonly consumed dairy products analysed on a

fresh weight basis. ...................................................................................................... 85

Table 26. Mean iodine contents in commonly consumed leafy vegetables analysed on

a fresh weight basis. ................................................................................................... 86

Table 27. Mean iodine contents in commonly consumed fruits and vegetables analysed


xx

Table 28. Mean iodine contents in commonly consumed seaweeds analysed on a fresh

weight basis. ............................................................................................................... 89

Table 29. Mean iodine contents in commonly consumed food samples analysed on a

fresh weight basis. ...................................................................................................... 90

Table 30. Comparison of iodine contents in selected food samples from the present

research with previous published data analysed on a fresh weight basis. .................. 98

1

CHAPTER 1

INTRODUCTION

1.1. General background

Iodine was discovered in the year 1811 by Courtois and is an important biogenic

element (Patzeltová, 1993). Iodine having a symbol I, and molar mass 126.9, is the

heaviest member of the halogens and occurs naturally. There are many forms of iodine

in the environment. Elemental iodine does not exist in a stable form in nature. The two

most common forms of naturally occurring iodine are iodide (I-) and iodate (IO3-) ions.

The iodate ion (IO3-) is a very good oxidising agent while iodide is a reducing agent

(Winger et al., 2005).

Plants and animals including human beings have very little contact with iodine due to

its low concentration in the environment. The iodine content in the earth’s crust

including the sea and the atmosphere accounts for 6 × 10-6 % of the total earth’s mass

according available geochemical data (Patzeltová, 1993). The Chilean alum which is

considered to be the richest source of iodine has 0.2% iodine in the form of sodium

iodide (Patzeltová, 1993). Iodine species exists in a variety of forms in the environment

and in different biological functions. It exists mainly in water, air, soil and food

(Blazewicz, 2012) and thus is found in the human body as well. Some research have

been carried out to identify the concentrations of iodine in the different organs or body

parts (Hou et al., 1997; Okerlund, 1997; Tadros et al., 1981; Zabala et al., 2009;

Andrási et al., 2004; Levine et al., 2007; Braetter et al., 1998).

The concentration of iodine that transfers from the soil to plants is generally low and

as a result there is only a small iodine intake through the plant root system. It is likely

that the uptake of atmospheric iodine by the aerial parts of plants is a vital process

being a major source for grazing animals. However, human intake of iodine is mainly

from food sources. The other source of iodine is from drinking water (Ronald and

Christopher, 1986).

2

1.1.1. Iodine in water

The literature review shows that the iodine content of rainwater is mostly within the

range 0.5 - 2.5 mg/L (Ronald and Christopher, 1986). Earlier literature compilations

of data for surface waters suggest that they generally contain < 20 mg/L iodine with a

range of 0.5 - 5 mg/L (Ronald and Christopher, 1986). It has been also found that

ground waters are more enriched in iodine than surface waters (Ronald and

Christopher, 1986). In water, iodine is mostly found as iodate (IO3-) and iodide (I-)

ions. The other forms of iodine in water exist as periodate (IO4-), hypoiodite (IO-),

methyl iodide (CH3I), methyl diiodide (CH2I2), ethyl iodide (C2H5I), propyl iodide

(C3H7I), butyl iodide (C4H9I) and methyl bromide iodide (CH2BrI). It has also been

reported that the organic iodine concentrations are higher in fresh water sources such

as water from lakes, rivers and rain (Blazewicz, 2012).

1.1.2. Iodine in air

Iodine in air is mostly in particulate form. Inorganic gaseous form exists as I2 and

hypoiodous acid (HIO) whereas the organic gaseous iodine mostly exists as CH3I and

CH2I2. It has also been reported that high concentrations of iodine are found in urban

areas due to the combustion of oil and coal (Blazewicz, 2012). Gaseous iodine (I2) is

high in concentrations in coastal areas due to the emissions from seawater, sea spray

and algae and is dependent on the location, season and climate (Blazewicz, 2012).

1.1.3. Iodine in soil

The weathering of rock materials containing iodine leads to the enrichment of soil with

iodine (Ashworth, 2009). The average reported iodine contents in igneous rocks was

0.24 mg/kg, 5 – 200 mg/kg for sediments, 2.7 mg/kg for carbonates, 2.3 mg/kg for

shales and 0.8 mg/kg for sandstones (Ronald and Christopher, 1986). It has been found

that sedimentary rocks have higher iodine content than other rock types (Ronald and

Christopher, 1986). Iodine present in soil is mostly due to its atmospheric transport

from the ocean and deposition in soil. The weathered rocks and soil exhibit higher

iodine concentrations. Soil samples close to the coast, where there was high rainfall

and areas with high organic matter exhibited high iodine values. Iodine retention in

3

soils is influenced by many factors such as soil pH, moisture, porosity and the

composition of organic and inorganic matters (Blazewicz, 2012 9).

1.1.4. Iodine in food and plants

As iodine is present in air, water and soil, the iodine levels in food and plants may vary

due to geographical locations leading to the differences in topsoil irrigation ways and

other factors. Iodine is not categorised as an essential element for plant growth.

However, the plant roots uptake iodine through the liquid phase of soils. This becomes

an important phase as iodine entering plants is transferred to the atmosphere and may

enter the food chain (Ashworth, 2009). Investigations have shown that iodised salt,

fish, eggs, meat, poultry, shellfish and milk are the main sources of iodine in diets

where iodine in humans is very similar to the levels in fish (Anke et al., 1995; Centre

for Food Safety - Hong Kong, 2011; Cressey, 2003; Eckhoff and Maage, 1997;

Haldimann et al., 2005; Jooste and Strydom, 2010; Leufroy et al., 2015). However, the

widest encountered iodine is present in seaweeds where concentrations vary in

different species of seaweed. For example, brown seaweed have mostly I- however

the green seaweeds are home for many organic molecules to which the iodine is bound

(Blazewicz, 2012 ).

1.1.5. Iodine in human body

Iodine is also present in the human body as an essential element. In clinical practice,

iodine is commonly analysed in urine, serum, blood and a variety of tissues. The

bioavailability of organic iodine, especially associated with macromolecules, is low

compared to I- and IO3- which have high bioavailability (Hou, 2009). A study revealed

that 96.4% of the potassium iodide (KI) is absorbed in humans (USFDA, 2009). Iodine

analysis in humans has been mainly carried out in the thyroid, plasma, brain, hair,

human milk and urine (Blazewicz, 2012; Nitschke and Stengel, 2015; WHO, 2007).

1.1.6. Iodine in thyroid

Iodine exists in the form of triiodothyronine (T3) and thyroxine (T4) in the thyroid

hormones in humans and mammals and plays a key structural role. In thyroid samples,

iodine is measured as precursor forms such as monoiodotyrosine (MIT) and

4

diiodotyrosine (DIT) or isomeric forms such as reverse triodothyronine (rT3) may also

be measured. It has been shown that iodine accounts for 65% of the molecular weight

of T4 and 59% of T3 (Blazewicz, 2012). In thyroid and hormones, 15 – 20 mg iodine

is concentrated and the other 70% is distributed in other tissues (Blazewicz, 2012).

The T3 and T4 thyroid hormones regulate many biochemical reactions, such as protein

synthesis and enzymatic activity where the major target organs are the brain, heart,

muscle, pituitary and kidney (Muhammad et al., 2014). If the body is unable to

synthesize the thyroid hormones because of insufficient iodine, the hormones cannot

regulate metabolism in every cell of the body which plays a role in all physiological

functions, thus having a devastating impact on human health (Muhammad et al., 2014).

In a study reported by Hou et al. (1997), the average iodine contents determined in six

tissues have been as follows heart 46.6 ± 14.9 ng/g, liver 170 ± 34 ng/g, spleen 26 ±

8.6 ng/g, lungs 33.3 ± 10.6 ng/g, muscle 23.5 ± 14.3 ng/g and finally in hair as 927 ±

528 ng/g. In the USA, the mean value of iodine in the thyroid was found as 10 mg per

thyroid while patients with autoimmune thyroiditis and hypothyroidism had a 2.3

mg/thyroid for 13 patients tested (Okerlund, 1997).

Tadros et al. (1981) determined iodine concentrations which ranged from 0.02 to 3.12

mg/g of thyroid tissues obtained from 48 thyroids at an autopsy while the mean

concentration was 1.03 ± 0.67 mg/g. Another interesting research was conducted by

Zabala et al. (2009) to determine the iodine content in thyroids of the male population

in Caracas, Venezuela. The median thyroidal iodine concentration was 1443 ± 677

μg/g (wet weight) ranging from 419 to 3430 μg/g which corresponded to a median

total iodine content of 15 ± 8 mg. There was also no relation seen between the iodine

concentration compared to the age and the weight of the thyroid gland.

1.1.7. Iodine in plasma

Iodine exists in plasma and represents about 0.5% of the total plasma iodine. The other

portion in plasma occurs as specific plasma protein which is also known as protein-

bound iodine (Allain et al., 1993). The determination of iodine in plasma is an

alternative to iodine determination in urine as urine is difficult to collect (Aumont and

Tressol, 1987). The total plasma iodine concentrations have been reported to be 40 –

5

80 μg/L (Aumont and Tressol, 1987). In the study, the authors have also concluded

that anyone having less than 40 μg/L total plasma iodine concentrations is more likely

to develop hypothyroidism and for individuals with a total plasma iodine concentration

of 80 – 250 μg/L will probably develop hyperthyroidism and Graves’ disease (Allain

et al., 1993).

1.1.8. Iodine in brain

The quantitative data on the iodine concentrations in the human brain is scarce. The

location and nature of the forms of iodine binding in the human brain is still unknown.

In an investigation, on the iodine distribution in the lipid fraction and brain tissue

without lipid showed that the mean iodine content was 910 ± 147 and 281 ± 68 ng/g

dry weight, respectively depending on the brain region. The highest iodine

concentration was found in susbstantia nigra and the lowest concentration in the vermis

crebelli (Andrási et al., 2004).

1.1.9. Iodine in hair

A study was conducted by Levine et al. (2007) to determine the iodine content in hair

samples. It was observed that the iodine content in hair samples ranged from 0.483 to

15.9 μg/g. In another study, iodine in hair was determined in autistic children. It was

confirmed that iodine content was lower in this group compared to the iodine in the

hair of the control group children. Low levels of iodine in the hair of autistic children

indicated that iodine could be important in the aetiology of autism due to its effect on

the thyroid function (Adams et al., 2006).

1.1.10. Iodine in human milk

Iodine present in human milk is mainly in the form of I- and comprises of about 80%

of total iodine. The other 20% iodine is comprised of higher molecular weight

molecules (Braetter et al., 1998). Iodine was analysed in human milk and infant milk

formulae from different manufacturers in several European countries by Fernández-

Sánchez and Szpunar (1999). Another method was developed by using inductively

coupled plasma mass spectrometry (ICP-MS) to determine the iodine in human milk

and infant formulae and it was seen that human milk had 144.0 ± 93.2 μg/kg iodine

6

whereas infant formulae had around 53.3 ± 19.5 μg/kg iodine (Fernández-Sánchez et

al., 2007).

1.1.11. Iodine in urine

Iodine in urine occurs as I- but other forms can also be found. The bioavailability of

iodine is high and about 90% of iodine consumed is excreted in urine (Jooste and

Strydom, 2010). Therefore, urinary iodine (UI) serves as a good indicator of the dietary

iodine intake and thus the overall iodine status (Jooste and Strydom, 2010). Iodine

content in a urine sample is a direct indicator of daily iodine consumption by humans

(Patzeltová, 1993). Thus, the measurement of UI concentration is the primary tool in

the assessment of the nutritional iodine status and to evaluate iodine supplementation.

According to WHO (2004), the recommended median UI concentration of 100 - 199

μg/L is considered optimal. Populations having iodine concentrations between 50 - 99

μg/L are classed as having mild iodine deficiency. Populations with 20 - 49 μg/L

iodine concentrations are classified as having moderate iodine deficiency whereas

populations with < 20 μg/L iodine are classed as severe iodine deficiency. Populations

having iodine concentrations between 200 - 299 μg/L are said to be at risk of iodine

induced hyperthyroidism while those having > 300 μg/L UI are at risk of adverse

health consequences (WHO, 2004).

1.2. Problems associated with low and excess iodine intake

The bulk of iodine entering the human body is via the food chain. It is therefore

becomes essential to have knowledge of iodine levels in food stuffs and diets to assess

the amount of iodine intake by humans to see if the recommended daily iodine intakes

are met with. The lack of iodine in the body leads to iodine deficiency disorders

(IDDs), while excessive iodine intake can result in pathological problems

2009).

Iodine deficiency also causes cognitive impairment (Shelor and Dasgupta, 2011). It is

said that the brain is extremely sensitive to the effects of low iodine intake but is

dependent on the timing and severity of the deficit. Even a mild iodine deficiency is

seen to affect the intelligence and functions of children. Infants relying on their

mothers’ milk are vulnerable if the mothers’ iodine intake is low. This shortage of

7

iodine in infants can lead to congenital hypothyroidism which has drastic effects on

the neurological functions. It has been reported that even mild iodine deficiency is

linked to behavioural and cognitive dysfunction (Shelor and Dasgupta, 2011). Studies

have shown that children with less than 100 μg/L UI have lower intelligence quotient

(IQ) and higher chances of behavioural disorders than their peers (WHO, 2004;

FAO/WHO, 2006; WHO, 2007). In case of women, mild iodine deficiency correlates

with attention deficit hyperactivity disorders (ADHD) in their children (Shelor and

Dasgupta, 2011; Marjan et al., 2013; WHO, 2007). Thus, iodine deficiency is a global

problem and studies show that 30% of the world’s population live in areas with iodine

deficient soils (Marjan et al., 2013). The WHO (2007), has recommended a daily

iodine intake as shown in Table 1.

Table 1. Daily iodine intake recommended by World Health Organisation.

Age group (month/years) Daily iodine intake (μg)

Preschool children (up to 59 months) 90

School children (6-12 years) 120

Adolescents/ adults 150

Pregnant and lactating women 250

Iodine intake of more than the recommended levels or excess iodine intake can lead to

health problems such as hyperthyroidism and thyroid autoimmune diseases (Marjan et

al., 2013). Thus, the European Union (EU) and major countries, including USA and

Canada have set the upper level of iodine intake as shown in Table 2.

Table 2. Tolerable daily iodine intake by population in different countries of different

age groups.

Age (years) EU (μg) Age (years) USA/Canada

(μg)Children 1-3 200 Children 1-3 200

Children 4-6 250 Children 4-8 300

Children 7-10 300 Children 9-13 600

Children 11-14 450

Adolescents 15-17 500 Adolescents 14-18 900

Adults 18 and above 600 Adults 19 and above 1100

8

Source: European Commission/ Scientific Committee on Food, 2002 and U.S.

Institute of Medicine (IOM), 2001.

Nitschke and Stengel (2015) have reported that an inadequate iodine intake may lead

to major health conditions such as the dysfunction of the thyroid gland. It can also lead

to subclinical enlargement of the thyroid gland, which is a condition called goitre.

Iodine deficiency can also adversely affect reproduction (Nitschke and Stengel, 2015).

Thus, the WHO (2007) has summarised the health consequences of iodine deficiency

for different age groups as shown in Table 3.

Table 3. Spectrum of iodine deficiency disorders (IDDs) for different age groups.

Physiological groups Iodine deficiency health consequences

All ages Goitre

Hypothyroidism

Increased susceptibility to nuclear radiation

Fetus Spontaneous abortion

Stillbirths

Congenital anomalies

Perinatal mortality

Neonate Endemic cretinism plus mental deficiency with

mixture of mutism, spastic diplegia, squint,

hypothyroidism and short stature

Infant mortality

Child/adolescent Impaired mental function

Delayed physical development

Iodine-induced hyperthyroidism

Adults Impaired mental function

Iodine-induced hyperthyroidism

9

It has also been reported by Nitschke and Stengel (2015) that excess iodine intake can

cause hyperthyroidism. Most iodine enters the human body through ingestion.

Therefore, it is important to understand the amounts of iodine content in foods and

natural products intended for consumption to estimate the iodine intake by humans

through these sources.

1.3. Justification of the study and significance of research

Based on the above discussions, it is very clear that iodine is one of the essential trace

elements and is of much interest in nutritional research. It is needed for the generation

of T3 and T4 hormones for the proper growth and development of the human body. The

major part of the essential iodine enters the body through food intake (Bhagat et al.,

2009). However, iodine deficiency is still a global public health concern because it

leads to a number of functional and development abnormalities known as IDDs, goitre

being one of the well known (Blazewicz, 2012; Shelor and Dasgupta, 2011). Other

disorders associated with iodine deficiency include cretinism and mental retardation.

Many countries have introduced supplementation programmes, promoting the use of

iodised salt or iodised vegetable oil to prevent iodine deficiency (Leufroy et al., 2015).

It has also been reported that Pacific Island populations have one of the world’s highest

thyroid cancer rates (Leufroy et al., 2015).

The analysis and compilation of nutritional levels of various commonly consumed Fiji

foods have been done by the Institute of Applied Sciences (IAS) together with the

Food and Agricultural Organization (FAO) and presented as Pacific Food

Composition Table (Dignan et al., 2004). However, iodine levels in those foods have

neither been determined nor reported in the Pacific Foods Composition Table (Dignan

et al., 2004). Hence, considering public health issue, it was essential to determine

iodine levels in commonly consumed foods in Fiji. Since reliable information was

lacking on the iodine content of food products in Fiji, publications from this project

will provide a valuable source of data for public health purposes. Thus, the project was

initiated with the following aim and objectives.

10

1.4. Aim: To determine the iodine content in locally available foods in Fiji using the

spectrophotometric kinetic method.

1.5. Objectives:

The objectives of the proposed project were to:

Validate the kinetic method for the determination of iodine.

Determine the iodine contents in commonly consumed foods in Fiji.

Compare the iodine contents in some fresh and factory-processed selected food

products.

11

CHAPTER 2

LITERATURE REVIEW

This chapter in general explains the importance of iodine in the human body and the

issues associated with iodine deficiency. Iodine deficiency is further described as a

global concern and comparison is made between countries and populations studied.

The chapter also explains the iodine deficiency studies carried out in Fiji. The

recommended iodine intake for people based on their age groups is further discussed.

This chapter further explains the iodine contents in different types of foods using

different analytical methods. The Sandell-Kolthoff reaction, which is one of the most

common methods for iodine determination in different foods and biological samples

is explained in detail in this chapter.

2.1. Introduction

Iodine is an essential trace element and of much interest in nutritional research. In the

human body, it is essential for the production of T3 and T4 hormones which are

responsible for the proper functioning and the development of the human body (Bhagat

et al., 2009). These hormones regulate body temperature and metabolic rate in adults

and children. They also play an important role in the normal development of the brain

and nervous system, before birth in babies, and young children. It is therefore

particularly important that pregnant women, breast feeding mothers and young

children have an adequate dietary iodine intake (Nitschke and Stengel, 2015).

Iodine also helps in the maturation of the central nervous system, and the development

of foetal and early postnatal life (Gónzalez-Iglesias et al., 2012). The most known

adverse effect of iodine deficiency is goitre. Goitre is an enlargement of the thyroid

which lies in the front of the neck where the thyroid gland lies. Goitre is however just

one effect of iodine deficiency, the others include endemic cretinism, infant mortality,

infertility, miscarriage, mental retardation, neuromuscular defects, and dwarfism. All

these are commonly known as IDDs (Eckhoff and Maage, 1997).

12

Assessment of iodine nutrition in the populations is mostly carried out by measuring

UI concentration, thyroid stimulating hormone (TSH), serum thyroglobulin (TG), and

the goitre rate. Those recommended indicators are complementary where UI is a

sensitive indicator of recent iodine intake (days) while TG shows an intermediate

response (weeks to months). On the other hand, changes in the goitre rate reflect long-

term iodine nutrition status (months to years) and the TSH is a valuable indicator of

iodine deficiency in neonates (Gónzalez-Iglesias et al., 2012).

2.2. Global estimates

Low iodine resulting in IDDs is a global concern, while excessive iodine intake is not

that common . IDDs occur when the iodine intake falls below the

required intake levels Stengel, 2015). This is a natural

ecological phenomenon which occurs globally. The erosion of soil due to loss of

vegetation for agricultural production, overgrazing and deforestation result in iodine

losses from the soil and, thus, it is a global issue. Therefore, foods grown and water in

such areas will be iodine deficient. It has been reported by WHO that IDDs mostly

affect the brain. Populations/ communities with low iodine intake resulting in IDDs

can sustain brain damage and reduced cognitive capacity. In this way, the potential of

the whole community is reduced by iodine deficiency. In cases of severe deficiency,

there is little chance of achievement among populations and communities and under

development is seen (WHO, 2007).

On a global basis, iodine deficiency is the single most preventable cause of brain

damage (WHO, 2007). It has been highlighted that people with severe iodine

deficiency may have an IQ of up to 13.5 points lower than of those having no iodine

deficiency (WHO, 2007). This mental deficiency has an immediate effect on child

learning capacity, women’s health, the quality of life in communities and economic

productivity. Surprisingly, IDDs are among the easiest and the least expensive of all

nutrient disorders to prevent (WHO, 2007). In addition, there is also a risk of taking

excessive amounts of iodine which can cause hyperthyroidism and thyroid immune

diseases (Shelor and Dasgupta, 2011).

13

It has been estimated that more than two billion people have insufficient iodine intake

and are at a risk of developing IDDs (Shelor and Dasgupta, 2011). In their review of

analytical methods for the quantification of iodine in complex matrices, Shelor and

Dasgupta (2011) have referred mostly to the US population and concluded that the

status for most countries regarding iodine nutrition is not better than that of the USA.

It has been reported that 91% of the global population totalling some 130 countries are

regularly checked for their iodine nutrition status (Shelor and Dasgupta, 2011). Iodine

deficiency became a recognised problem in 1993 in 123 countries but by the year 2006,

this number has decreased where 63 countries still do not have regular iodine screening

procedures. Thus the present scenario on iodine nutrition is complicated, therefore this

makes it a global challenge to try and simplify iodine nutrition status (Shelor and

Dasgupta, 2011). Furthermore, Leufroy et al. (2015) highlighted that Pacific Island

populations have the world’s highest thyroid cancer incidences rates. They have also

highlighted that data on iodine in foods is limited in the Pacific and has not even been

reported in the FAO’s Pacific Island Food Composition Table (Leufroy et al., 2015;

Dignan et al., 2004).

In addition, Judprasong et al. (2016) have reported that Thailand was classified as a

country with the optimum iodine nutrition but not so for all regions in Thailand. North

and North-East regions of Thailand still have mild iodine deficiency. It was seen that

in Thailand, to overcome IDDs, food fortification with iodine was applied in table salt,

fish sauce and soy sauce. Kulkarni et al. (2013) highlighted that in India, about 71

million people suffer from IDDs. In areas like Uttar Pradesh, Bihar, Madhya Pradesh,

Maharashtra and Gujarat states, contribute to almost 70% population, having IDDs

shown in a statistics by the Ministry of Health and Family Welfare, Government of

India (Kulkarni et al., 2013).

In 1993, the WHO published the first version of the Global Database on Iodine

Deficiency with global estimates on the prevalence of iodine deficiency (WHO, 1993).

The international community and authorities in most countries where iodine deficiency

was a public health problem decided to take measures to control this. The WHO

recommended a strategy to prevent and control IDDs through salt iodisation

programmes (WHO, 2007).

14

The problem of IDDs remains a global problem due to the decreases or losses of iodine

content during processing or cooking as seen in several parts of the world especially

in developing countries (Salau et al., 2010). In Fiji, IDDs were recognised as a public

health problem in the year 1996. In the same year, the Government of Fiji passed a

Gazette in Cabinet that prohibited the import of non–iodised salt into Fiji. This Gazette

was passed because of a survey of iodine status by the United Nations Children’s Fund

(UNICEF), WHO, Ministry of Health and Ministry of Education in 1994. Later, this

law was incorporated under the Fiji Islands Food Safety Regulations 2009, under the

Fiji Islands Food and Safety Act 2003 (Food Safety Regulations – Fiji Islands, 2009).

This survey was carried out in Ba, Sigatoka and Suva. The prevalence of goitre was

studied in school children and pregnant women by ultrasound and was found to be

around 45% of goitre cases. Mean UI studies were also conducted in 15 schools in the

same areas and the UI ranged from 2 - 94 μg/L with an average of 26 μg/L indicating

moderate to severe IDDs (International Council for Control of Iodine Deficiency

Disorders, 2009). Thus, it is important that Fiji population should consume iodine rich

foods as iodine supplementation.

2.3. Recommended iodine intake

The International Council for Control of Iodine Deficiency Disorders (ICCIDD),

UNICEF and WHO recommended that the daily intake of iodine should be 90 μg for

preschool children (0 to 59 months), 120 μg for schoolchildren (6 to 12 years), 150 μg

for adolescents (above 12 years) and adults, 250 μg for pregnant and lactating women

(WHO, 2007). The U.S. Institute of Medicine (IOM) daily recommended adequate

intake of iodine for the different population groups is as follows: 0 - 6 months – 110

μg, 7 - 12 months – 130 μg, 1 - 8 years – 90 μg, 9 - 13 years – –

150 μg, pregnant women – 220 μg, and lactating women – 290 μg (Shelor and

Dasgupta, 2011). For comparison, the recommended daily intake (RDI) of iodine for

Australia and New Zealand population based on their age and gender is shown in Table

4 (Nutrition Australia, 2010).

15

Table 4. Recommended daily intake of iodine in Australia and New Zealand in

different age groups.

Age (years) Gender RDI (μg/day)

1 – 8 Boys and girls 90



19 – >70 Men 150

19 – >70 Women 150

Pregnancy Women 220

Lactation Women 270

2.4. Urinary iodine

In the implementation of IDDs control programs, the measure of UI is the principal

indicator of iodine determination rather than the thyroid size, TSH or TG. Thyroid size

is however a more useful assessment of the severity of the IDDs and has a role in

assessing the long term impact of control programs. Over 90% of dietary iodine

appears in the urine. The urine samples are easy to collect and available for analysis

thus making it an ideal candidate for analysis to measure the dietary iodine intake. The

UI excretion can vary in individuals from day to day. Table 5, therefore, shows the

criteria for assessing iodine nutrition in groups based on median urinary iodine

concentrations (n = 30) as well

as lactating women and children < 2 years old (Marjan et al., 2013).

2.5. Iodine in foods

The natural dietary sources of iodine in foods include; milk, cereals, fruits, vegetables,

eggs, meat, spinach and sea foods. These natural sources may not satisfy the

requirements of iodine intake in humans as these iodine sources may not be

bioavailable in a form as needed by the body and also that the iodine concentrations

are low (Kulkarni et al., 2013). Iodine occurs in food mainly as inorganic iodide which

is readily and almost completely absorbed in the gastro intestinal tract (Longvah et al.,

2013). Marine macro algae (seaweeds) represent an important source of food,

supplements, fertilisers and medicine in many parts of the world. They are considered

16

to have high nutritional value and their metabolites and associated biological activities

have a lot of significance for multiple nutrceutical, cosmetical and pharmaceutical

applications (Nitschke and Stengel, 2015). Seaweeds are generally divided into three

main categories: (1) brown algae (phaeophyceae), (2) red algae (rhodophyta), and (3)

green algae (chlorophyta). It is said that the brown algae accumulates high levels of

iodine (Nitschke and Stengel, 2015). In all categories of algae, iodine can be retained

both in an inorganic as well as organic form.

Table 5. Criteria for assessing iodine nutrition in groups based on median UI

concentrations.

Median UI (mg/L) Iodine intake Iodine nutrition

< 20 Insufficient Severe deficiency

20 – 49 Insufficient Moderate deficiency

50 – 59 Insufficient Mild deficiency

100 – 199 Adequate Optimal

200 – 299 More than adequate Risk of iodine-induced

hyperthyroidism

> 300 Excessive Risk of adverse health

consequences- iodine induced

hyperthyroidism, autoimmune

thyroid disease

Pregnant women

< 150 Insufficient

150 – 249 Adequate

250 – 499 More than adequate

Excessive

Lactating women and children < 2 years old

< 100 Insufficient

Adequate

Source: Marjan et al. (2013).

17

A study on the iodine contents in fresh mass foodstuffs by Koutras et al. (1985)

revealed that iodine content was low in the range of 10 - 200 μg/kg, but it was observed

that iodine content (fresh mass) in seafoods was higher. In another study Moreda-

Pin˜eiro et al. (2007) determined the total iodine concentrations in edible seaweed

samples in Northwest Spain. They concluded that most of the iodine is present as

iodide (I-) in brown seaweed samples which is 90% of the total iodine whereas the red

seaweed has iodide lower than 30% and green seaweeds lower than 80%.

According to Judprasong et al. (2016), the top 10 commonly consumed food items

identified from the Thai National Food Consumption Survey were: jasmine rice, kale,

boiled banana, prawn, steamed short-bodied mackerel, iodine enriched hen egg, yard

long-bean, chicken thigh, milk powder, fermented fish and shrimp paste. For

comparison of the methods, iodine determination was carried out using alkali dry-

ashing, dissolved in water and analysed by spectrophotometric and ICP-MS methods.

The iodine contents in these foods were found to be in the range of 3 – 1304 μg/100g.

An analysis by Leufroy et al. (2015) on the determination of iodine in French

Polynesian foods, showed a lot of variation in iodine content. Fruits tested had a 0.014

– 0.032 mg/kg iodine content, starchy samples showed 0.014 – 0.081 mg/kg, green

vegetables 0.027 – 1.85 mg/kg, fish 0.222 – 5.19 mg/kg, shellfish 6.51 – 85.6 mg/kg

and 0.004 – 1.39 mg/kg iodine in beverages. The database on the iodine content in

foods and diets showed that iodine content was highest in marine fish 1456 μg/kg

followed by fresh water fish 106 μg/kg, leafy vegetables 89 μg/kg, dairy 84 μg/kg,

other vegetables 80 μg/kg, meat 68 μg/kg, cereals 56 μg/kg, fresh fruit 31 μg/kg and

bread 17 μg/kg (Leufroy et al., 2015). The results show that grain crops are poor

sources of iodine when compared to leafy vegetables. There are also some evidences

that indicate leafy vegetables have high iodine content than some other vegetables

(Leufroy et al., 2015). The iodine content of foods varies with the different geographic

locations therefore, iodine content from one country cannot be universally used to

estimate the iodine intake for another population (Longvah et al., 2013).

18

2.5.1. Food fortification to prevent iodine deficiency

Food processing is one of the earliest technologies that humans have been using due

to inherent advantages of ensuring food supply, increase stability, improving

flavouring and decreasing the possibility of toxicity. The effect of processing of foods

was studied by Salau et al. (2010). The results indicated that a significant reduction of

iodine is seen in processed foods compared with raw forms. The authors concluded

that consideration must be given to the different food processing methods when

assessing iodine intake from different processed foods. Some of the food processing

methods include fermentation, frying and cooking which introduce chemicals that

affect the food nutritional value adversely (Salau et al., 2010). However, iodine

deficiency can be controlled through fortification of food and food products. Adding

iodised salt in cooked food is an example.

Many countries have regulations that control the levels of iodine intake through the

diet (Bhagat et al., 2009). Many countries have introduced supplementation

programmes, promoting the use of iodised salt or iodised vegetable oil to prevent

iodine deficiency (Nitschke and Stengel, 2015). Many countries have a significant

portion of daily iodine intake achieved by supplementation such as iodised salts

. However, there is a declining use of salt at homes because of possible

adverse health concerns for excessive sodium consumption and high blood pressure.

Generally, the iodine content in most foods is low. Despite this, about 90 % of dietary

iodine is derived from food and the remaining 10 % from drinking water. The iodine

content of grains, fruits and vegetables is generally determined by the environmental

factors in which they grow in (soil, water, geographical location and use of fertilizers).

With marine foods, both of animal and plant origin, the levels, can still be variable

based on the amount of iodine which has been accumulated from the sea water

(Haldimann et al., 2005). Iodine has also been reported to be commercially added in

bread making through iodine based dough conditioners which increase the iodine

content in breads (Salau et al., 2010).

Food fortification was a solution implemented by most countries to eliminate the

chances of iodine deficiency. However, recent studies in the USA have shown that

iodine intake has decreased over time and thus more people are at risk due to iodine

19

deficiency (Shelor and Dasgupta, 2011). The decrease in iodine intake may be due to

changes in practices among dairy and cereal manufacturers, removal of iodate based

conditioners from breads, an increased reliance on pre-packed, pre-packaged and fast

foods which may have a lot of salt but not iodised salt. It has been highlighted that

most of the salt consumed outside the home in the US is not iodised salt (Shelor and

Dasgupta, 2011). The authors have further reported that pregnant and lactating women

who make efforts to “eat healthy” during pregnancy or in the early motherhood may

be at a risk of iodine deficiency when they try and restrict the consumption of salt at

home.

2.5.2. Salt iodisation to prevent IDDs

Salt iodisation has been considered the preferred strategy that is economical,

convenient and an effective means to control and to prevent IDDs. However, iodine in

iodised salt may be volatile and lost through the cooking process of foods. One study,

showed that iodine losses occur from iodised salt through the cooking processes where

a mean iodine retention of 60 ± 21% was observed in 139 commonly consumed Indian

foods (Longvah et al., 2012). Other studies of different foods using iodide salt showed

a variation of 14 – 94% in the retention of iodine during food preparation

(Szymandera-Buszka and Waszkowiak, 2004; Amr and Jabay, 2004; Azanza et al.,

1998). The retention of iodine from iodised salt has been seen to be affected by many

factors such as food variety, type of cooking utensil used, recipe used and the time of

addition of iodised salt to the food during preparation (Longvah et al., 2013).

Kulkarni et al. (2013) have also confirmed that adequate iodine intake can be achieved

by consumption of iodised salt. This iodisation is done by adding iodate to salt samples

due to its good stability and bioavailability. Salt fortification is normally done with

potassium iodide (KI) and potassium iodate (KIO3) due to their low cost and good

iodine availability. However, international organisations like WHO and UNICEF have

recommended KIO3 or iodate (IO3-) ion over iodide due to their stability. It has also

been recommended that after iodisation the salt should be stored in warm and humid

conditions (WHO, 2007). This is due to the fact that 20% of the iodine in salt is lost

from production to a house hold supply. Another 20% is lost during cooking before

consumption (WHO, 2007).

20

The investigation of iodine levels in foods and drinking water has shown that the

highest iodine concentrations are present in seafood (Bhagat et al., 2009; Chilean

Iodine Educational Bureau, 1952; Haldimann et al., 2005; Leufroy et al., 2015; Varo

et al., 1982; Mahesh et al., 1992). However, this source from seafood is not usually

sufficient to supply daily requirements especially in pregnant women. To meet daily

human iodine requirements, iodine can be provided through iodine supplementation

through iodised salt. Many countries have adapted to this approach of food fortification

to eliminate the chances of iodine deficiency as well as the chances of excess iodine

intake by controlling the limits of iodine in salts. The demand and knowledge of iodine

contents in foods including salt is therefore needed to combat the issues associated

with IDDs globally. Thus presently, salt iodisation is the most common method used

to control and eliminate IDDs (Marjan et al., 2013).

2.6. Analytical methods for iodine quantification

The main sources of iodine in a normal, balanced diet are fish, shellfish, milk and

iodised salt. Other sources of iodine in the normal diet are food supplements containing

iodine. As most iodine enters the human body through food intake, the knowledge of

iodine contents in foods and natural products is essential for estimating the daily iodine

intake (DII). The food supplements have a complex composition of vitamins and

minerals thus making the determination of iodine very difficult (Osterc and Stibilj,

2006).

Various analytical methods have been used for the determination of trace amounts of

iodine in different types of samples. These include inductively coupled plasma mass

spectrometry (ICP-MS) (Leufroy et al., 2015; Romarís–Hortas et al., 2011; Gónzalez-

Iglesias et al., 2012; Eckhoff and Maage, 1997; Haldimann et al., 2005; Dyke et al.,

2009; Pieter, 2010) radiochemical neutron activation analysis (RNAA) (Adotey et al.,

2011; Osterc and Stibilj, 2006), ion chromatography (IC) (Rebary et al., 2010; Rong

et al., 2007; Malongo et al., 2008; Bruggink et al., 2007; Hu et al., 2009), high

performance liquid chromatography (HPLC) (Melichercik et al., 2006), HPLC with

UV detection (Nitschke and Stengel, 2015), HPLC-diode array detection (Gupta et al.,

2011), inductively coupled plasma–atomic emission spectrophotometry (ICP-AES)

21

(Varga, 2007) and atomic absorption spectrophotometry (AAS) (Haase and Broekaert,

2002). However, all these methods require expensive instrumentation and complex

sample preparation. As an inexpensive alternative, spectrophotometric kinetic

methods to determine iodine content in food and dairy products has continuously been

used (Mahesh et al., 1992; Pieter, 2010; Moreda-Pin˜eiro et al., 2007; Longvah and

Deosthale, 1998; Cressey, 2003). It is an attractive procedure because of its high

sensitivity and accuracy without using expensive equipment (Shelor and Dasgupta,

2011).

Iodine nutrition assessment methods require affordable and accurate quantification

methods in different samples of soil, plants, foods, serum, urine, etc. Iodine

measurements in samples are presently carried out by two common methods: One is

the kinetic spectrophotometric method known as Sandell-Kolthoff reaction (Sandell

and Kolthoff, 1934, 1937). In this indicator reaction, yellow Ce4+ is reduced to

colourless Ce3+ by As3+. Usually this reaction is very slow. Iodide catalyses this

reaction making it faster and can be used in the kinetic spectrophotometric method

(Shelor and Dasgupta, 2011). The reaction involved is shown below.

2Ce4+ + 2I- 3+ + I2

As3+ + I2 5+ + 2I-

This reaction is also catalysed by iodate but to a much smaller extent and because of

the presence of arsenite in an acidic medium, iodate gets converted to iodide. The rate

of the disappearance of the yellow colour is a measure of the iodine concentration.

Ce3+ is fluorescent with ex 254 nm and em 350 nm. Thus, this reaction can also be

fluorometrically monitored. However, the draw back for this method is that there can

potentially be organic species that may be blocking/interfering with the Ce4+ and the

Ce3+ which may affect the rate of the indicator reaction. In order to eliminate any

organic compounds, complete mineralisation of the sample is thus required in the

digestion step.

The other method that is most widely used for iodine quantification and analysis in

recent years is ICP-MS. This ICP-MS method permits superb sensitivity and in some

cases allows the direct sample determination of iodine such as in urine after dilution.

22

However, the use of internal standards is required in ICP-MS method to account for

any matrix effects (Shelor and Dasgupta, 2011).

Most of the techniques mentioned above, except NAA, are not selective, suffer from

interferences and need pre-concentration or separation procedures which may lead to

the loss of iodine (Bhagat et al., 2009). Another reason is that iodine concentrations in

food matrices are low and losses due to its high volatility make it challenging to

analyse (Leufroy et al., 2015). Thus, the majority of methods use the initial step for

the determination of iodine in biological materials. This requires the conversion of

iodine into a form such as iodide and iodate which may be reliably analysed.

Conversions are mostly carried out in the form of dry alkaline in an alkaline medium

or wet ashing involving digestion using a strong acid

medium. Reproducible results are obtained only when the losses of iodine are avoided

at the incineration stage (Patzeltová, 1993).

2.6.1. Inductively coupled plasma-mass spectrophotometry (ICP-MS)

In this quantification technique, microwave or radio frequency power is applied

through an induction coil to generate high temperature argon plasmas and electron

temperatures. The sample gets atomised by the plasma and then strips the atoms of one

or more valence electrons. As a result the positive ion enters a quadrupole mass

analyser for sorting out ions of different m/z and then detected. The first ionization

potential of iodine is reasonably high to form I+ of 10eV. The drawback for this

technique is that during the ionization step iodine present is partially ionised

(approximately 25%). Despite this partial ionisation, the sensitivity of iodine in ICP-

MS is considered superior in terms of other techniques for iodine determination

(Shelor and Dasgupta, 2011).

A recent study for the determination of total iodine in French Polynesian foods used

this highly expensive ICP-MS technique (Leufroy et al., 2015). Fresh food samples

were freeze dried and ground to obtain <300 μm powder prior to analysis. 0.1 – 0.5 g

dry samples or 1 g liquid samples were weighed in 50 mL polypropylene flasks in

duplicates and extracted with 5 mL of ultrapure water and 1 mL of

tetramethylammonium hydroxide (TMAH – rs in a heating block.

23

The extracts were cooled, diluted to 25 mL and centrifuged at 4000 revolutions per

minute (rpm) and filtered through a 5 μm filter. A final filtration was done through

0.45 μm filter before sample dilution and determination by ICP-MS. The limit of

quantification (LOQ) was 0.027 mg/kg and the limit of detection (LOD) was 0.014

mg/kg. A total of 124 food samples were successfully analyzed (Leufroy et al., 2015).

Haldimann et al. (2005) also analysed iodine contents in different food groups

available in the Swiss market by isotope dilution ICP-MS using the enriched long lived

nuclide 129I. Iodine contents in food were generally low, therefore accurate

determination was required with high sensitivity which was achieved by the use of

miniaturised sample introduction system and the application of isotope dilution

analysis.

A study was conducted to measure iodine concentrations in conventional, organic and

ultra-heat treated (UHT) semi-skimmed milk at retail in the UK by ICP-MS after

alkaline extraction (Stevenson et al., 2018). Milk samples were collected from

supermarkets and stored at - carried out by

diluting 100 μL milk sample with 10 mL of 0.22 M TMAH in ultrapure water and

containing 5 μg/L Rh as an internal standard. The samples were filtered using a 0.45

μm filter to remove fats which could block the nebuliser in the ICP-MS. Analysis of

results showed that conventional milk had higher iodine content followed by UHT

milk and then organic milk. It was also seen that milk produced in summer had lower

iodine contents to those in winter (Stevenson et al., 2018).

In addition, in an analysis for iodine contents in foods, a comparison of methods was

done between the spectrophotometric and the ICP-MS methods (Judprasong et al.,

2016). Food samples were treated by alkaline ashing where 30% w/v potassium

carbonate (K2CO3) and 10% w/v zinc sulphate (ZnSO4) were added and evaporated on

a steam bath until dry. Then the samples were dry-as

for 2 hrs to remove organic species. If ashing was not complete 1 mL of 10% ZnSO4

solution was added and the charred residue was broken with a glass rod to disperse it

in the solution. The samples were again heated on a steam bath until dry. Ashing was

repeated until a white ash was obtained. The samples were then analysed

spectrophotometrically using the Sandell-Kolthoff reaction at 410 nm and also by ICP-

24

MS. The recovery results obtained by spectrophotometry were in the range of 80 –

108% whereas 92 – 103% was achieved by ICP–MS. This study made analysis by

ICP–MS as the recommended method for iodine analysis in foods (Judprasong et al.,

2016; Stevenson et al., 2018; Leufroy et al., 2015).

2.6.2. Inductively coupled plasma-optical emission spectrophotometry (ICP-OES)

ICP-OES is also known as inductively coupled plasma-atomic emission

spectrophotometry (ICP-AES) and both use plasmas for atomization and excitation or

ionization. The light emitted from the excited atom is measured at a particular

wavelength which is specific for the analyte. This technique for iodine determination

is not frequently used because the sensitivities are not good (Shelor and Dasgupta,

2011).

In a study by Varga (2007) using ICP-AES to determine iodine in dietary products, the

operating conditions were a Plasma 27.1 MHz crystal driven radio frequency (RF)

generator with air flow rates cooling at 12 dm3 min 1, plasma 0.6 dm3 min 1, aerosol

0.8 dm3 min 1, a nebulizer GMK type with 2.4 mL min 1 sample uptake rate. It was

found that ICP-AES measurement was seriously affected by spectral line coincidence

between the prominent line of iodine and the adjacent phosphorus line thus was not

used as a suitable indicator for iodine in dietary supplements. Moreover, this technique

was also quite expensive for use in common laboratories.

2.6.3. Neutron activation analysis (NAA)

The neutron activation analysis (NAA) is not a readily available instrumentation to all.

In NAA, the sample is irradiated with neutrons and the emission is monitored in the

form of a radioactive isotope. The advantage of this method is that sample preparation

is not needed. However, self-protection is required from the radioactive emissions.

Iodine is a monoisotopic element and all its isotopes are radioactive except for 127I.

Thermal neutron energy of 0.025 eV is commonly used for iodine determination.

Interference from sodium, potassium, bromine and chlorine is common (Shelor and

Dasgupta, 2011). In normal chemical analysis of iodine, the sample is usually

decomposed by ashing or acid digestion in which high temperatures and strong

oxidising and reducing reactions are required to dissolve the sample completely or

25

convert iodine into a suitable form for determination. NAA which is considered a non-

destructive analytical method, where the sample and standard solution of iodine

existing as inorganic iodine compounds must be dried and irradiated with neutrons

(Xiaolin et al., 1998).

In a study by Muhammad et al. (2014), the NAA was compared to the traditional

Sandell-Kolthoff reaction. In the NAA method, 0.5 g of dried and homogenised food

sample was weighed and digested with 7 mL ultrapure HNO3 in a microwave oven at

1000W for 2 min. The system was cooled to room temperature and further cooled in

ice bath for one hr. The clear solution of the digested sample was mixed with 1 g of

hydrazine sulphate. The sample cup and lid were washed with 3 portions of 5 mL of

5% hydrazine sulphate solution and deionised water. The sample and washings were

combined and diluted to 100 mL. The pH was then adjusted to 2 - 3 using 10%

ammonia solution. Iodine was co-precipitated with 1 mL of 0.05 M bismuth sulphate

and 0.25 M thioacetamide. The precipitate was filtered, dried, weighed and packed for

irradiation. The samples were individually irradiated and after a decay of about 2 min,

the nuclide spectra were obtained by counting the samples for 600 sec using 441keV

gamma ray of 128I. The spectra acquired were processed and analysis was done via a

software (Muhammad et al., 2014).

Most analysis for iodine quantification are not selective, suffer interferences or need

pre-concentration or separation procedures which usually leads to iodine losses. The

NNA technique using reactor neutrons is one of the best techniques for iodine

determination due to its favourable nuclear properties that lead to high sensitivity and

thus applicability to measure trace amounts of iodine in samples like food and food

products (Bhagat et al., 2009).

2.6.4. Atomic absorption spectrometry (AAS)

Direct determination of iodine using AAS is difficult due to the fact that there is no

commercial lamp available and in addition, the best iodine absorption band lies in the

vacuum UV region in which the optical path needs to be purged with an inert gas.

Good detection limits have been achieved by indirect iodine analysis. Iodide solutions

from alkaline ashing of samples were precipitated with Ag+ as AgI. The precipitate

26

was then washed with dilute ammonia for the removal of silver salts and then dissolved

in dilute thiosulfate followed by the determination of silver which was correlated to

iodine (Yebra and Bollain, 2010).

Haase and Broekaert (2002) have reported the indirect determination of iodide with

AAS where the sample solutions containing iodide were mixed mostly offline with

mercury (Hg) prior to analysis. The authors developed an online procedure for the

indirect determination of iodide based on its interference in the determination of Hg

with cold vapour AAS. The procedure made use of a commercially obtained Hg

analyser where the interference of NaCl on the determination of Hg was used. It was

noticed that the interference caused by halogenides with the system resulted in an

indirect determination of iodide. Subsequently, an online procedure, based on mixing

iodide and Hg in a flow injection system, was developed for the indirect determination

of iodide by flow injection cold-vapor AAS.

2.6.5. Electrochemical and potentiometric probes

Ion selective electrodes (ISEs) for iodine determination are commercially available

and have been applied for iodine determination for a long time. Commercial iodide

ISEs are based on insoluble silver salt membranes and also react to other high level

anions forming insoluble silver salts. Recent studies have focused on fabricating iodide

selective ionophores having higher selectivity than silver salt based ISEs.

Electrochemical detections have widely been used because of their fast response time,

sensitivity and selectivity for the analyte iodine. The silver working electrode is the

most commonly used electrode for iodide analysis using the electrochemical technique

(Zhang et al., 2005).

In an assessment to rapidly measure the iodide content in milk samples, an ISE method

was used (Melichercik et al., 2006). An Orion ion selective electrode and a general

purpose electrode were used on a Radiometer set to potentiometric mode. Analysis of

milk samples for iodide confirmed that ISE method, because of its low cost and

simplicity, is well suited method for rapid screening of iodide in raw and processed

milk (Melichercik et al., 2006).

27

2.6.6. Gas, liquid and ion chromatographic methods

Chromatography is the most commonly used separation technique in modern

analytical laboratories. Though the catalytic spectrophotometric method is associated

with low cost equipment but chromatographic systems with different detectors are the

techniques for fast, simple, reliable and sensitive methods (Blazewicz, 2012).

Recently, a high pressure liquid chromatography (HPLC) method for the

determination of iodine in natural samples of edible seaweeds and commercial

seaweed food products was developed as a simple and reliable method for an accurate

determination of total iodine contents (Nitschke and Stengel, 2015). Potential

interfering compounds were removed from the process by incinerating the organic

compounds and the inorganic compounds were removed by a chromatographic

separation process. The cost effectiveness was maintained by using a standard HPLC

equipped with a diode array detector. The analysis time was around 20 min (Nitschke

and Stengel, 2015).

Analytical methods for the determination of iodine by HPLC are generally based on

ion chromatography. In milk, iodine is almost exclusively found as iodide. Thus, the

quantification of iodide in milk by HPLC is a recognised official method by the

Association of Official Analytical Chemists (AOAC). The results for the analysis of

iodide in milk samples using HPLC gave very reliable results. Thus, the authors

strongly recommended the use of HPLC for the determination of iodide in milk

samples (Melichercik et al., 2006).

The estimation of iodine intake and the analysis of iodine content in seaweed using

gas chromatography with electron capture detector (GC-ECD) was used by (Yeh et al.,

2014). The iodine in seaweeds was derivatized with 3-pentanone and detected by GC-

ECD with a detection limit of 0.5 mg/kg. The method developed was compared with

ICP-OES and GC-ECD was said to be the low cost alternative to ICP-OES for iodine

detection in seaweeds.

Another GC method for the determination of total iodine in foods was based on the

reaction of 3-pentanone with iodine (Mitsuhashi and Kaneda, 1990). An alkaline

28

ashing techniques was used for organic matter destruction and the ash residue was

diluted in water. This water extract was then oxidised in the presence of H2SO4 by

adding Cr2O72- to liberate the iodine. This liberated iodine then reacted with 3-

pentanone to form 2-iodo-3-pentanone, which was extracted into n-hexane and then

determined by GC-ECD. The detection limit for iodine was 0.05 μg/g and the recovery

from spiked food samples was in the range of 91.4 - 99.6% (Mitsuhashi and Kaneda,

1990). Another GC-ECD technique was applied to the determination of elemental

iodine in toluene and cyclohexane solvents where the retention index was based on the

alkyl iodide and n-paraffin series. Resolutions were obtained to resolve I2 from alkyl

iodides. A detection limit of 39 ng I2 was obtained (Fernandez et al., 1984).

Furthermore, silver-based solid carbon paste electrode was developed for use as a

detector in ion chromatography (IC) for the sensitive determination of iodide in real

samples (Malongo et al., 2008). This method was successfully applied to the

determination of iodide in complex samples such as table salts, sea products and iodide

bound drug compounds. Determination of iodide and iodate in edible salt by IC with

integrated amperometric detection was found feasible method for idodate

(Rebary et al., 2010). A rapid method for the direct

determination of inorganic iodine in plasma using ion exchange chromatography (IEC)

and the Sandell-Kolthoff reaction was developed by Aumont and Tressol (1987). The

separation of plasma inorganic iodine from other organic iodine was carried out after

the precipitation of plasma protein with ethanol. Iodide was then then determined by

alkaline ashing and via Sandell-Kolthoff

a spectrophotometer at 420 nm (Aumont and Tressol, 1987). Inorganic plasma iodine

as low as 3 μg L-1 concentration could be determined with only 0.5 mL sample.

However, only a few of these methods are currently used in routinely for analysis of

iodine (Rong et al., 2007; Malongo et al., 2008; Rebary et al., 2010; Bruggink et al.,

2007; Hu et al., 2009). This is due to the fact that these chromatographic methods

require expensive instrumentation.

2.6.7. UV-Visible spectrophotometry

Iodine is an oxidant and many chromogenic substrates can be oxidised to coloured

compounds. The trick is to retain the iodine in a form which the iodine derived species

29

is the only oxidant (Shelor and Dasgupta, 2011). A rapid and feasible method to

determine total iodine and iodide in edible seaweeds by an inexpensive analytical

technique, catalytic spectrophotometry, was developed by Moreda-Pin˜eiro et al.

(2007). Seaweed samples were treated by applying a microwave-assisted alkaline

digestion with TMAH. The percentage of iodide was measured directly in the alkaline

digests by observing the catalytic effect of iodide on the oxidation of As3+ by Ce4+ in

H2SO4/HCl medium (Moreda-Pin˜eiro et al., 2007). Thus, a microwave-assisted

sodium tetrahydroborate (NaBH4) reduction was optimized to determine total iodine

using a UV spectrophotometer based on its catalytic effect on the said oxidation

reaction (Moreda-Pin˜eiro et al., 2007).

In a rapid assessment method to determine iodate in table salt samples, Kulkarni et al.

(2013) used an iodometric reaction between iodate, excess iodine and an acid. The

iodine emitted is allowed to react with variamine blue dye in the presence of sodium

acetate to yield a violet coloured species which showed absorbance maxima at 550 nm.

This method allowed the analysis of iodate in the range of 10 - 25 ppm. The kinetics

of the reaction was very fast and thus a large number of samples could be easily

screened for their iodate content in a short period of time.

Furthermore, another sensitive spectrophotometric kinetic determination of iodine in

foodstuffs was presented by Mahesh et al. (1992) where food samples were alkaline

ashed using KOH and ZnSO4 water, centrifuged

and analysed. The reaction used was the reduction of Ce4+ to Ce3+ in the presence of

As3+ catalysed by iodide ion (I-) and the absorbance was measured at 370 nm. The

recovery presented was in the range of 94 – 102% with a mean of 97% and the

detection limit and sensitivity were 0.4 ng and 40 pg, respectively. Various analytical

methods for the determination of iodine in different types of samples are summarised

in Table 6 along with their LOD, LOQ and some other relevant comments.

30

Tab

le 6

. Ana

lytic

al m

etho

ds fo

r the

det

erm

inat

ion

of io

dine

in d

iffer

ent s

ampl

es.

Met

hod

Sam

ples

LO

D

(LO

Q)

RSD

(%)

Spik

e

reco

very

(%)

Com

men

tsR

efer

ence

HPL

C w

ith U

V d

etec

tion

Nat

ural

seaw

eed

and

com

mer

cial

seaw

eed

food

prod

ucts

~ 0.

2 ng

/μL

(~ 1

ng/

μL)

1.34

-5.6

992

.6–

108.

9D

ry a

lkal

ine

inci

nera

tion

and

UV

iodi

de d

etec

tion

at 2

23

nm.

Nits

chke

and

Sten

gel,

2015

Sand

ell-K

olth

off r

eact

ion

pris

m sp

ectro

phot

omet

er

Seru

mN

/AN

/AM

ean

-92.

9A

lkal

ine

inci

nera

tion

of

sam

ples

.

Acl

and,

195

7

Sand

ell-K

olth

off r

eact

ion

UV

Vis

ible

spec

troph

otom

eter

Seaw

eeds

9.2

μg/g

(30.

7 μg

/g)

10.4

97–

100

Mic

row

ave

assi

sted

dig

estio

n

and

redu

ctio

n.

Mor

eda-

Pin˜

eiro

et

al.,

2007

Sand

ell-K

olth

off r

eact

ion

Ion

exch

ange

chro

mat

ogra

phy-

spec

troph

otom

eter

Plas

ma

3 μg

/L10

.39

98.0

7A

lkal

ine

inci

nera

tion.

Det

ectio

n at

420

nm

.

Aum

ont a

nd

Tres

sol,

1987

31

Sand

ell-K

olth

off r

eact

ion

UV

vis

ible

spec

troph

otom

eter

Urin

e an

d

milk

2 ng

/g<

893

.08

-95.

41A

lkal

ine

inci

nera

tion.

Det

ectio

n at

420

nm

.

Aum

ont a

nd

Tres

sol,

1986

UV

spec

troph

otom

etry

Sand

ell-K

olth

off r

eact

ion

Food

0.4

μg/

100g

(1.3

4 μg

/100

g)

8.2

85-1

08Sp

ectro

phot

omet

ry a

chie

ved

low

er a

ccur

acy.

Judp

raso

ng

et a

l., 2

016

ICP-

MS

Food

0.03

μg/

100g

(0.1

0 μg

/100

g)

4.9

92-1

03IC

P-M

S pr

ovid

ed g

reat

er

accu

racy

.

Judp

raso

ng

et a

l., 2

016

Sand

ell-K

olth

off r

eact

ion

phot

o-co

lorim

eter

Bio

logi

cal

mat

eria

ls

N/A

(10

μg/

dm)

N/A

80.2

Wet

ash

ing

-aci

dic

med

ium

.

Abs

orba

nce

at 4

05 n

m.

Ave

rage

iodi

ne lo

ss o

f 19.

8%.

Patz

elto

vá,

1993

Sand

ell-K

olth

off r

eact

ion

UV

vis

ible

spec

troph

otom

etry

Edib

le

seaw

eeds

9.2

μg/

g –

iodi

de

28.5

μg/

g –

iodi

ne

2.6

%fo

r

iodi

de

5.8%

for

tota

l iod

ine

9.7

-94.

9M

icro

wav

e as

sist

ed d

iges

tion/

dist

illat

ion.

Die

go e

t al.,

2005

ICP-

MS

Food

sam

ples

0.01

4 m

g/kg

(0.0

27 m

g/kg

)

5.8

-22.

480

-120

Alk

alin

e di

gest

ion

usin

g

TMA

H.

Leuf

roy

et

al.,

2015

32

Epith

erm

al n

eutro

n

activ

atio

n an

alys

is

(EN

NA

)

Food

pro

duct

s

and

salt

0.02

–0.

6

mg/

kg

1-1

3N

/AB

oron

car

bon

filte

r.B

haga

t et a

l.,

2009

Iodo

met

ric re

actio

n

spec

troph

otom

etry

Tabl

e sa

lt0.

25 μ

gN

/AN

/AA

sim

ple

rapi

d m

etho

d fo

r

iodi

de d

eter

min

atio

n 2

- 30

μg.

Kul

karn

i et

al.,

2013

Pre-

conc

entra

tion

neut

ron

activ

atio

n

anal

ysis

Sand

ell-K

olth

off r

eact

ion

Food

sN

/A11

.34

N/A

Rea

ctor

use

d fo

r iod

ine

anal

ysis

.

Muh

amm

ad

et a

l., 2

014

Inst

rum

enta

l neu

tron

activ

atio

n an

alys

is

Bio

logi

cal

mat

eria

ls

N/A

N/A

90–

99Io

dine

loss

es d

urin

g sa

mpl

e

prep

arat

ion

and

anal

ysis

was

mon

itore

d.

Xia

olin

et a

l.,

1998

Sand

ell-K

olth

off r

eact

ion

UV

vis

ible

spec

troph

otom

etry

Food

stuf

fs;

wat

er; t

issu

es

and

body

fluid

s

0.4

ng<

6 94

-102

Rec

over

y re

sults

wer

e go

od.

UV

det

ectio

n at

370

nm

.

Mah

esh

et

al.,

1992

33

Qua

rtz c

ryst

al

mic

roba

lanc

e

Food

stuf

fs0.

0005

mg/

L4.

193

.2-1

01.1

Met

hod

base

d on

sens

itive

resp

onse

to m

ass c

hang

e at

elec

trode

of p

iezo

elec

tric

quar

tz c

ryst

al. I

nter

fere

nce

foun

d fr

om b

rom

ine.

Yao

et a

l.,

1999

Ion

chro

mat

ogra

phy

with

UV

det

ectio

n

Salt

45.5

3 μg

/L2.

198

.4-1

01.6

Pum

p cy

clin

g-co

lum

n-

tech

niqu

e co

uple

d w

ith h

igh

exch

ange

cap

acity

col

umns

.

Zhon

gpin

g et

al.,

2013

ICP-

MS

Edib

le

seaw

eed

24.6

ng/

g

(82.

0 ng

/g)

N/A

N/A

Mic

row

ave

assi

sted

alk

alin

e

dige

stio

n pr

oces

s and

in-v

itro

dige

stio

n pr

oced

ure

used

.

Rom

arís

–

Hor

tas e

t al.,

2011

Cat

alyt

ic

spec

troph

otom

etric

(UV

)

Iodi

ne in

coa

l0.

09μg

/g

(0.2

9 μg

/g)

5.87

102.

58A

bsor

banc

e m

easu

red

at 4

20

nm.

Wu

et a

l.,

2007

Ion-

spec

ific

elec

trode

vs.

HPL

C m

etho

d

Milk

HPL

C –

6

μg/L

(HPL

C –

20

μg/g

)

N/A

87-1

14%

for

ISE

91 -

100%

for

HPL

C

HPL

C m

etho

d id

eal w

ith lo

w

dete

ctio

n lim

it.

Mel

iche

rcik

et a

l., 2

006

34

Ion

chro

mat

ogra

phy

with

inte

grat

ed a

mpe

rom

etric

dete

ctio

n

Edib

le sa

ltN

/A5.

23N

/AR

educ

tion

of io

date

to io

dide

with

sodi

um b

isul

phite

.

Reb

ary

et a

l.,

2010

Gas

diff

usio

n flo

w

inje

ctio

n an

d

chem

ilum

ines

cenc

e

Phar

mac

eutic

al p

rodu

cts

0.1

mg/

L4.

8N

/AIo

dide

oxi

dise

d to

iodi

ne g

as

and

mea

sure

d.

Rat

anaw

imar

nwon

g et

al.,

2005

Ion

chro

mat

ogra

phy

UV

det

ectio

n

Seaw

ater

19 μ

g/L

(66

μg/L

)

4.19

99.6

-101

.2U

V d

etec

tion

at 2

20nm

.R

ong

et a

l.,

2007

Ion

chro

mat

ogra

phy

usin

g a

silv

er-b

ased

carb

on p

aste

ele

ctro

de

Rea

l iod

ine

sam

ples

0.47

μg/

L3

72.9

-104

.9R

edox

beh

avio

ur o

f iod

ide

ions

was

stud

ied

at e

ach

elec

trode

by

cycl

ic

volta

mm

etry

.

Mal

ongo

et

al.,

2008

Sand

ell-K

olth

off r

eact

ion

UV

det

ectio

n

Food

sam

ples

8 μg

/kg

12.5

93-1

01Io

dine

rete

ntio

n fr

om

unco

oked

to c

ooke

d fo

od w

as

mea

sure

d.

Long

vah

et

al.,

2013

ICP-

MS

Fish

and

food

prod

ucts

0.08

1 μg

/LN

/AN

/AN

itric

aci

d sa

mpl

e di

gest

ion.

Eckh

off a

nd

Maa

ge, 1

997

35

Sand

ell-K

olth

off r

eact

ion

UV

vis

ible

spec

troph

otom

eter

Food

s and

wat

er

N/A

N/A

N/A

Stud

y do

ne in

Nor

th E

ast

Indi

a.

Long

vah

and

Deo

stha

le,

1998

Isot

ope

dilu

tion-

ICP-

MS

Food

N/A

N/A

N/A

Sam

ple

dige

stio

n in

HN

O3

acid

.

Hal

dim

ann

et

al.,

2005

Kin

etic

col

orim

etric

met

hod

Dai

ry

prod

ucts

0.04

mg/

kg –

dry

solid

sam

ples

(0.0

2 m

g/kg

liqui

d or

sem

i-

solid

sam

ples

)

14.8

77-1

10A

lkal

ine

dry

ashi

ng o

f

sam

ples

.

Cre

ssey

,

2003

Tota

l ref

lect

ion

X-r

ay

fluor

esce

nce

(TX

RF)

and

ICP-

atom

ic e

mis

sion

spec

trom

etry

(IC

P-A

ES)

Die

tary

supp

lem

ent

prod

ucts

0.37

mg/

LN

/AN

/ASa

mpl

e ex

tract

ion

with

amm

onia

solu

tion.

Var

ga, 2

007

Chr

omat

ogra

phic

tech

niqu

es w

ith U

V a

nd

ICP-

MS

dete

ctio

n

Seaw

eeds

0.12

μg/

L-

Iodi

de

0.2

μg/L

-

Ioda

te

Iodi

de -1

.0

Ioda

te -

1.2

N/A

Alk

alin

e di

gest

ion

of sa

mpl

es.

Shah

et a

l.,

2005

36

Four

ier t

rans

form

infr

ared

(FT-

IR)

Palm

oil

N/A

N/A

N/A

Iodi

ne sp

ectru

m o

f pal

m o

il at

a ra

nge

of 3

025-

2992

cm

-1.

Man

and

Setio

wat

y,

1999

Per-

vapo

ratio

n-flo

w

inje

ctio

n w

ith

chem

ilum

ines

cenc

e

dete

ctio

n

Mul

tivita

min

tabl

ets

0.5

mg/

L5.

281

.3-1

17Io

dide

oxi

dise

d to

iodi

ne.

Che

milu

min

esce

nt e

mis

sion

at

425

nm.

Nac

apric

ha

et a

l., 2

007

HPL

C-d

iode

arr

ay

dete

ctio

n

Tabl

e sa

lt3.

7 μg

/L7.

998

.42-

Iodo

sobe

nzoa

te a

nd N

,N-

dim

ethy

lani

line

have

bee

n

used

at p

H 6

.4 fo

r sel

ectiv

e

conv

ersi

on o

f iod

ide

to

4-io

do-N

, N-d

imet

hyla

nilin

e

whi

ch w

as e

xtra

cted

with

etha

nol,

whe

n th

e ph

ase

sepa

ratio

n oc

curr

ed b

y

addi

tion

of a

mm

oniu

m

sulp

hate

.

Gup

ta e

t al.,

2011

37

Ion

chro

mat

ogra

phy

Urin

e20

μg/

L1.

586

-98

Sam

ple

oxid

ised

to io

date

and

the

alka

line

dige

stio

n so

lutio

n

neut

ralis

ed.

Elec

troch

emic

al p

re-tr

eatm

ent.

Hu

et a

l.,

2009

Spec

troph

otom

etric

dete

rmin

atio

n- fl

ow

inje

ctio

n an

alys

is

Iodi

sed

salt

0.02

mg/

L1.

297

.3-1

00.1

Met

hod

base

d on

reac

tion

of

ioda

te w

ith h

ydro

xyla

min

e in

acid

ic so

lutio

n.

Shab

ani e

t

al.,

2011

Sand

ell-K

olth

off r

eact

ion

colo

rimet

ry

Geo

chem

ical

sam

ples

0.05

μg/L

3.87

75-9

0V

anad

ium

pen

toxi

de u

sed

a

flux

for p

yroh

ydro

lysi

s.

Rae

and

Mal

ik, 1

996

NA

: Not

ava

ilabl

e

38

2.7. Sample digestion

Most analytical techniques require sample decomposition, which is a basic problem in

iodine determination because of the highly volatile iodine compounds such as HI and

CH3I. This problem can be resolved in two ways: 1) the conversion of all iodine species

into its elementary form (I2) either by distillation or by combustion with the trapping

of analyte for further processing. 2) The conversion of all volatile iodine species to a

non-volatile state such as iodide or iodate . Hence, the sample

digestion/preparation is extremely important in iodine determination.

The first described method of sample decomposition involves distillation in an oxygen

atmosphere in a closed flask and combustion in a stream of oxygen flowing through a

heated tube . The second described method for decomposition of iodine

in foodstuffs comprises of dry ashing (fusion) with alkaline ashing aids such as NaOH,

NaOH plus NaNO3, Na2CO3, and oxidative fusion with Na2O2 in a Parr bomb

2009). Wet ashing involving oxidising acids can also be used provided the oxidation

potential is high to oxidise iodine to the non-volatile iodate using mixtures such as

H2SO4-chromate, H2SO4-HNO3-HClO4 or HClO3-HNO3. It has been reported that no

losses of iodine were found in wet digestion with HNO3 at different pressures and

controlled temperatures in closed devices

TMAH also did not show

measurable losses of iodine . The analysis of serum protein bound

iodine has been carried out by alkaline incineration (Acland, 1957) and this method is

still in use.

Among many other methods, alkaline ashing prior to iodine determination by the

Sandell-Kolthoff reaction is one of the commonly used methods and is an official

AOAC method for the analysis of iodine (Aumont and Tressol, 1986). This method is

recommended by the authors because of its high precision, operational simplicity,

great recovery and is a rapid method for routine iodine analysis. It has also been

concluded that the proposed method has a low detection limit of 2 ng/g. Another ashing

procedure with slight modification was used by Aumont and Tressol (1987) to

determine the iodine in plasma using ion chromatography and the Sandell-Kolthoff

reaction.

39

Wet ashing technique with the Sandell-Kolthoff reaction to determine micro quantities

of iodine was used by Patzeltová (1993). This digestion procedure showed that the

recovery was unsatisfactory when K2CO3 was used instead of Na2CO3 which

confirmed that digestion is the most critical step in the iodine estimation (Patzeltová,

1993). Longvah et al. (2013) used the principle of alkaline incineration of the samples

mination of iodine by

measuring the rate of catalytic activity of iron thiocyanate by nitrite in the presence of

iodine.

In an analysis for the total iodine in edible seaweed by the Sandell-Kolthoff reaction,

Moreda-Pin˜eiro et al. (2007), used a microwave assisted TMAH digestion technique

for iodide determination where alkaline digests collected were analysed for iodide

content. For comparison of ICP-MS and spectrophotometry methods for iodine

determination in foods common alkali-ashing digestion procedure was used. Then

iodine was determined by kinetic spectrophotometric method at 410 nm as well as by

ICP-MS (Judprasong et al., 2016). All this discussion confirmed that the sample

preparation is a critical step for iodine quantification. The extraction using

concentrated acids should be avoided due to the formation of volatile species such as

hydrogen iodide (HI) and I2 which generally results in low recoveries (Leufroy et al.,

2015).

2.8. Sandell-Kolthoff (S-K) reaction for iodine determination

The most widely used technique for iodine determination is the Sandell-Kolthoff

reaction (Sandell and Kolthoff, 1934, 1937). Iodine is determined by the reduction of

Ce4+ coupled with the oxidation of As3+ to As5+. The reaction involved is shown as

follows (Patzeltová, 1993):

2Ce4+ + 2I- 3+ + I2

2Ce4+ + 2IO3- 3+ + I2 + 3O2

As3+ + I2 5+ + 2I-

The reduction of Ce4+ ions to Ce3+ changes the absorbance and the progress of the

reaction is monitored by the change in the yellow colour due to Ce4+ (Patzeltová,

40

1993). The role of the iodide ions is highly specific in this reaction. The reaction is

also much dependent on the temperature, concentration of H2SO4 and chloride. The

role of H2SO4 is to increases the reaction rate whereas the chloride stabilises it by

preventing the oxidation of iodine to iodate. The reaction mixture is kept fairly acidic

to prevent the precipitation of cerium(IV) arsenate (Patzeltová, 1993).

The reduction of Ce4+ to Ce3+ is typically measured at 405 – 420 nm. This

determination using the Sandell-Kolthoff reaction can be determined in two ways: 1)

the complete absorbance profile measured with time, and 2) measuring the absorbance

of the sample at a fixed time. The most commonly used method is measuring the

absorbance of the sample at a fixed time. Iodine concentration can be measured from

the difference in absorbance between the blank and sample (Shelor and Dasgupta,

2011). Aumont and Tressol (1986, 1987) have also used the Sandell-Kolthoff reaction

and the absorbance was measured at 420 nm on a spectrophotometer for the

determination of iodine in different samples. They concluded that the method is

simple, rapid, cheap and uses a small amount of sample.

2.9. Drawbacks of the Sandell-Kolthoff reaction method in iodine determination

The most commonly used Sandell-Kolthoff reaction has some drawbacks which have

been discussed by Shelor and Dasgupta (2011). This method is interfered by

uncharacterised organics. The presence of high concentrations of thiocyanate also

interferes with the analysis for iodine determination. Traces of metal ions such as silver

and mercury have also been found to interfere with the iodine analysis. Items that

readily undergo oxidation such as ferrous ion (Fe2+), nitrite (NO2-) and ascorbic acid

also interfere with the iodine analysis (Shelor and Dasgupta, 2011). Zinc was also said

to be an inhibitor for the catalytic action on the Ce4+ Ce3+ reaction and thus it

interferes at higher iodine concentrations (Acland, 1957). The analysis of iodine in

urine and milk samples by alkaline ashing procedure and determination by the Sandell-

Kolthoff reaction reported a low recovery of iodine as 93.08 - 94.41% (Aumont and

Tressol, 1986).

Mahesh et al. (1992) have discussed the use of dry ashing incineration methods for the

destruction of organic matters present in serum and biological materials. An

41

incomplete ashing was observed in food samples even after 12 – 36 hrs of ashing at

and this was common in cereals and oil seeds. To overcome this problem of

incomplete ashing, Moxon and Dixon (1980) had recommended the use of K2CO3 and

ZnSO4 for ashing. This method was successfully used for analysing iodine in milk and

food stuffs but poor recoveries and iodine losses were observed in the range of 10 –

30% (Mahesh et al., 1992). Moreda-Pin˜eiro et al. (2007) used microwave assisted

digestion and reduction for iodide and iodine determination in edible seaweeds

respectively and found that the analysis of the certified reference material was about

16% higher than the certified iodine concentrations.

According to Diego et al. (2005), the main drawback of the Sandell-Kolthoff reaction

is attributed to the fact that all iodine species must be present as iodide for the reaction

to occur. Therefore, sample pre-treatment which guarantees that all iodine species are

converted to iodide is needed. Acid digestion procedures are mostly reported as

common procedures for total element determination, however the risks of iodine losses

as iodine vapour happens as iodide reacts with protons. The use of alkaline digestion

procedures using TMAH or ammonia was therefore recommended (Diego et al., 2005).

2.10. Conclusion

Iodine deficiency is a major obstacle to the human and social development of

communities living in iodine deficient environments. The correction of iodine

deficiency would thus be the major contribution to the iodine deficient environment

(Longvah and Deosthale, 1998). Most iodine enters the human body through ingestion

and thus, the knowledge of iodine contents of food and natural products intended for

consumption is extremely important to estimate intake levels (Nitschke and Stengel,

2015). The quantification of iodine in foods is directly linked with the iodine uptake.

However, difficulties in the extraction and quantification explain why data/literature

on the iodine levels in foods are limited (Mahesh et al., 1992).

The inadequate accuracy of the analytical methods used for iodine analysis may also

contribute to the problem (Patzeltová, 1993). Thus, several analytical methods for

iodine quantification have been discussed above. Iodine intake are obtained from

iodised salt, however this does not always fulfil the requirements for recommended

42

iodine intake levels. Since food is being the major contributor of the total iodine

exposure for humans, ascertaining the iodine nutritional status of Fiji foods and dairy

products is of great importance as part of public health programs. Therefore, in the

proposed project, the determination of iodine content in selected Fiji foods has been

undertaken using inexpensive spectrophotometric catalytic kinetic method.

43

CHAPTER 3

RESEARCH METHODOLOGY

Iodine contents of food vary with different geographical locations due to a large

variation of iodine contents for different environmental areas. Thus, reliable data on

the iodine contents in foods can only be obtained by careful investigation using

appropriate and accurate analytical techniques. The choice of analytical methods with

strict quality control measures make an analysis and the data obtained of high quality.

The other prerequisite for obtaining valid and meaningful results is the availability of

a sufficiently homogenous and representative sample. Iodine determination in

foodstuffs has been a difficult analytical problem for many years and inconsistent

results are common in inter-laboratory studies (Shelor and Dasgupta, 2011; Haldimann

et al., 2005; ; Leufroy et al., 2015). There are a number of analytical

methods developed for iodine determinations in food stuffs, but the main difficulty in

these methods is the volatility of iodine when present in the elementary or in its volatile

form or state . Literature survey has indicated that the procedures of

iodine determination vary in terms of digestion techniques, analytical principles,

detection limits, specificity, sensitivity, accuracy, precision, recovery, robustness, time

involved, costs, equipment and the ease of performance. Therefore, this chapter

outlines the selected research methodologies that were used for iodine determination

and help to achieve the objectives of the research.

3.1. Chemical and reagents

All chemicals used were of high purity grade meeting the American Chemical Society

(ACS) reagent requirements except for arsenic trioxide, potassium hydroxide and zinc

sulphate which were analytical reagents (AR). All high purity grade chemicals were

purchased from Sigma-Aldrich, Australia. Thus, the ACS reagents potassium iodide

( ), diammonium ceric nitrate ((NH4)2Ce(NO3)6, ), nitric acid (HNO3,

70%), hydrochloric acid (HCl, 37%), sulphuric acid (H2SO4, 95 - 98%), potassium

hydroxide (KOH, AR grade), arsenic trioxide (As2O3, and zinc sulphate

(ZnSO4, AR grade) were used in this study.

44

Iodine stock standard was prepared from KI, by mixing 130.8 mg in 1 L of Milli-Q-

water. Iodine working standard solutions were then prepared by diluting the KI stock

solution in the range of 2.5 – 25 ng/mL. H2SO4 and HCl combined reagent was

prepared by adding 19.6 mL concentrated H2SO4 (specific gravity 1.840 g/cm3) to 500

mL water. This was mixed well and allowed to cool to room temperature. Then 5.4

mL concentrated HCl (specific gravity 1.20 g/cm3) was added, mixed and diluted to 1

L with Milli-Q-water (Mahesh et al., 1992).

0.05 M Ce4+ solution was prepared from diammonium ceric nitrate ((NH4)2Ce(NO3)6)

by dissolving 0.274 g in 10 mL Milli-Q-water. To this, 50 mL concentrated HNO3

(specific gravity 1.41 g/cm3) was added followed by addition of 5 mL of H2SO4. The

solution was allowed to cool to room temperature and made up to 100 mL with Milli-

Q-water. As3+ 0.030 M was prepared by dissolving 0.593 g of As2O3 and 0.6 g of KOH

in 100 mL of Milli-Q-water KOH (6 M) solution was prepared by dissolving

168.33 g KOH in 500 mL of Milli-Q-water. 0.52 M ZnSO4 solution was also prepared

by dissolving 74.76 g ZnSO4 in 500 mL Milli-Q-water.

3.2. Instrumentation

An oven with an automatic temperature control was used to maintain the oven

temperature at 120 ± 0.1 A high temperature Muffle furnace with a programmable

temperature setup (Model: YC-1400S)

Perkin Elmer Lambda 365 UV visible spectrophotometer equipped with 10 mm quartz

cells with a thermostatic water bath (Thermoline, Australia) was used to control the

temperature of the reagents and reaction . The absorbance of the

reduction of Ce4+ in the presence of As3+ within a fixed time (1 min) was measured at

370 nm. Figure 1 shows the instrumental set-up for UV visible spectrophotometer

connected to a computer, thermostatic water bath and printer.

45

3.3. Standard calibration curves

Iodine working standard solutions were prepared from KI stock solution in the range

of 2.5 – 25.0 ng/mL. The concentrations prepared were: 2.5, 5, 10, 15, 20 and 25

ng/mL. These standards were analysed using the Sandell-Kolthoff reaction and the

absorbance monitored for 1 min using a UV visible spectrophotometer to obtain the

calibration curve.

3.4. Food samples and sampling

The following 22 commonly consumed foods namely: rice (Oryza sativa), potato

(Solanum tuberosum), cassava (Manihot esculenta), dalo (Colocasia esculenta), fresh

fish, clam (Margaritifera), canned tuna, canned sardine, chicken egg, cheese, liquid

fresh milk, processed milk, butter, leafy (lettuce (Lactuca sativa), English cabbage

(Brassica oleracea), Chinese cabbage (Brassica chinensis)), fruit (tomato (Solanum

lycopersicum), banana (Musa), long green bean (Vigna unguiculata ssp.

Sesquipedalis) and pumpkin (Cucurbita moschata)) and sea grapes/green seaweeds

(Caulerpalentillifera) and lumiwawa/brown seaweeds (Gracilaria maramae) were

selected to analyse for their iodine contents.

Figure 1. Perkin Elmer Lambda 365 UV visible spectrophotometer equipped with 10

mm quartz cells (3) connected to a computer with the UV Express software (2), a printer

(4) together with a thermostatic water bath to control the temperature of the reagents and

reaction system at (1).

46

The selected food samples were purchased randomly from the Suva municipal market

and supermarkets. For each food item, 4 samples of different brands or varieties, or

from 4 different vendors were purchased. Thus, a total of 22 food samples (4 samples

of each category consisting of 88 sub-samples) were analysed on a fresh weight basis.

3.5. Sample storage and preparation

Fresh food samples were collected into clean polythene bags and kept in an ice box.

The samples were immediately transported to the laboratory. The edible part of each

sample was individually prepared and pulverized to obtain particles of required size

and analysed. All samples followed the same procedure except for liquid milk,

powdered milk and butter or margarine which were not pulverized. All other prepared

samples which could not be analysed on the same day were kept at -

washed screw capped plastic bottles until analysis.

3.6. Ashing procedure

The alkaline ashing procedure reported by Mahesh et al. (1992) as well as Nitschke

and Stengel (2015) was modified and used for the determination of iodine in food

samples. The fresh food samples (1 g) were transferred into clean dry test tubes in

duplicates. For recovery studies, 1 mL standard iodine solutions containing iodine

concentrations of 4, 12 and 18 ng/mL were added and mixed with the samples in the

test tube for analysis. To the food samples and recovery test tubes, 500 μL of 6M KOH

was added and mixed well. The test tubes with samples were then placed in an oven at

120 ± 0.1 for 24 hrs. The test tubes

containing the samples were then transferred to a Muffle furnace, shown in Figure 2,

which was operating at 120 for 30 min. The temperature of the Muffle furnace was

over 30 min and incineration was continued for exactly

1 hr.

47

Air was renewed in the Muffle furnace chamber every 15 min via a timer by opening

the furnace door for 10 - 15 sec. After 1 hr the test tubes with incinerated samples were

transferred to a desiccator and allowed to cool. Then 500 μL of 0.52M ZnSO4 was

added to the test tubes and the contents dried at 120 for 3 hrs. The test tubes were

then transferred to the Muffle furnace and another ashing procedure was performed

for 2 hrs renewing the air in the chamber every 15 min. The resultant ashed

samples, which were in white powder form (free from any carbon) were dissolved in

Milli-Q-water by placing them in an ultrasonic bath for 10 min then transferred to

centrifuge tubes and centrifuged at 4500 rpm for 10 min. The supernatant was filtered

using a 0.45 μm filter and stored at room temperature for analysis of total iodine.

Figure 2. Muffle furnace with digital temperature controller (1) and temperature

ramping setting (1 and 2).

48

3.7. Sample analysis

The kinetic spectrophotometric method using the Sandell-Kolthoff reaction was used

to determine the iodine concentration in foods. The catalytic effect of iodide on the

redox Ce4+ – As3+ reaction is a widely used technique to assess iodine by kinetic assay.

To initiate this reaction, 0.25 mL of Milli-Q-water, 0.25 mL of H2SO4- HCl mixture,

0.25 mL Ce4+ reagent and 0.25 mL As3+ reagent were transferred to a 10 mm path

length cuvette. The contents of the cuvette were mixed for 10 sec and the cuvette was

pre- min. The 0.25 mL catalyst iodine (blank/standard/sample)

was always added at the end to initiate the reaction. The decrease in the absorbance

due to the reduction of Ce4+ to Ce3+ was monitored for 1 min at 370 nm. The rate of

disappearance of the yellow colour in Ce4+ – As3+ indicator reaction system is analysed

as a measure of iodine content using the Perkin Elmer Lambda 365 UV visible

spectrophotometer equipped with 10 mm quartz cells (Mahesh et al., 1992).

The initial velocities i.e. A/min were calculated from the time-absorbance (A) curves

obtained for different iodine concentrations. Each standard was analysed 7 times and

the average change in absorbance was calculated. A standard calibration curve of

change in absorbance per min versus iodine concentrations was plotted as shown in

Figure 7. The iodine content of samples was calculated using the following formula

blank, m is slope of calibration curve and d is sample dilution (mL).

As Ab m × 4 × d = iodine (ng/g)3.8. Precision

Precision is a measure of errors associated with a number of repeated measurements

of the same parameter within a sample. Precision generally measures the closeness of

results and is normally expressed as absolute standard deviation, relative standard

deviation, variance, coefficient of variation or relative percent difference (Wisconsin

Department of Natural Resources Laboratory Certification Program, 1996). Thus, the

49

coefficient of variation was calculated as a measure of precision for all samples

analysed. A 6% coefficient of variation was observed by Mahesh et al. (1992) and a

mean ± standard deviation of 3.4 ± 1.77% for more than 20 foodstuff analysed

(Mahesh et al., 1992). The aim in the present research was also to see whether

precision of the method can be maintained or improved further.

3.9. Limit of detection

LOD or detection limit is the lowest concentration that can be determined to be

statistically different from a blank (Wisconsin Department of Natural Resources

Laboratory Certification Program, 1996). The LOD was calculated using the following

formula.

LOD ngmL = 3.143 (students t value) × standard deviation of analysis (n = 7)3.10. Limit of quantification

The LOQ is a level above which quantitative results can be achieved with a specified

degree of confidence. Generally, the LOQ is worked out as equal to 10 times the

standard deviation of a set of results for a series of replicates used to determine a

justifiable LOD (Wisconsin Department of Natural Resources Laboratory

Certification Program, 1996). The LOQ for this research was worked out using the

following formula.

LOQ ngmL = 10 × standard deviation of analysis (n = 7)3.11. Quality control

Quality control involves those steps taken to ensure that the analysis is under statistical

control. In this research, the following quality measures were carefully looked at to

ensure that the research was under good quality control.

50

3.11.1. Chemicals

All chemicals used in this study were of high purity grade meeting the American

Chemical Society (ACS) reagent requirements, except for AS2O3, KOH and ZnSO4

which were analytical reagents (AR) purchased from Sigma-Aldrich, Australia.

3.11.2. Preparation of Millipore water

Water used throughout the research was -1). This was

achieved by filtering deionised distilled water through the Millipore Milli-Q system

shown in Figure 3.

3.11.3. Glassware

All glassware used in this research were of certified A grade, unless specified

otherwise. All glassware used were soaked in 5 M HNO3 for a minimum of 12 hrs and

washed with distilled water, rinsed with deionised distilled water and dried before use.

Figure 3. The Simplicity brand Millipore Milli-Q system used to obtain Millipore water.

51

3.11.4. Data recording

Data was recorded manually in a lab note book with all details such as dates of work

and experiments conducted, quantity of reagents used, recording all details relating to

the research and all data or results collected. A softcopy of analytical data was kept in

the hard drive of the computer which had the Perkin Elmer Lambda 365 UV-visible

spectrophotometer connected. Another set of analytical data was stored on a flash drive

as a backup.

3.11.5. Standard operating procedure of analysis

A fixed procedure was agreed to after all trials and ensured that this fixed procedure

was followed throughout the analysis. This fixed procedure for analysis was time

based. This ensured the reagent mixing in the cuvette was for exactly 30 sec and the

reagents were for exactly 2 min. Sample/blank or

standard was always added at the end of the 2 min and the absorbance was measured

for exactly 1 min. This fixed procedure was used throughout this research.

3.11.6. Analysis of duplicate samples

Duplicate samples were always analysed as an efficient method of determining the

precision of an analysis. As far as the calibration curves were concerned, each sample

was analysed 7 times and then the averages were calculated. Finally, time versus

absorbance graphs was plotted. From this time versus absorbance graph, the change in

absorbance for the 7 sets data (n = 7) was used to work out the average change in

absorbance per minute at different iodine concentrations. Each food sample was also

analysed four times (n = 4) to provide accuracy and precision to the analysis.

3.11.7. Analysis of blanks

Analysis of blanks was carried out every time before starting the analysis of samples

on the Perkin Elmer Lambda 365 UV visible spectrophotometer to enable correction

of the measured signal for contributions from sources other than the analyte. Method

blanks were used to identify and correct systematic errors due to impurities in the

reagent, contamination in the glasswares and the instrumentation.

52

3.11.8. Analysis of standard samples

The analysis of standards containing a known concentration of iodine was used to

monitor the systems state of statistical control. New standards at concentrations of 4,

12 and 18 ng/mL were used to test the performance of the standard calibration curve

and the stability of the test method.

3.11.9. Standard calibration and linear equation

The standard calibration curve of the average absorbance versus time was plotted. A

very good linear relationship was obtained with a high R2 value (0.9998) indicating

the reliability of the determination of iodine in food samples.

3.11.10. Spike recoveries

The recovery analysis was carried out to confirm the precision and repeatability of the

data. The recovery was determined by dividing the obtained analyte iodine

concentration with the known concentration of the prepared standard. To get the

percentage recovery, the result was multiplied with 100 as shown below.

ecovery (%) = Concentration of analyte found by analysisConcentration of analyte added in sample × 100The iodine recovery was also carried out at 3 concentrations by adding 4, 12 and 18

ng/mL iodine to different samples. The standard deviation was also calculated for all

recovery values.

3.11.11. Analysis of Standard Reference Materials (SRM)

Analysis of Standard Reference Materials (SRM) was also performed alongside

samples to monitor the overall analytical performance. The SRM material chosen for

this research was Iodised Table Salt (Iodine as Iodide) SRM # 3530 from National

Institute of Standards and Technology (NIST).

53

3.11.12. Statistical analysis of data

Significance of differences between iodine concentrations in foods from different

sources and varieties was examined using the t-test and ANOVA. Measurements of

each sample were carried out four times and the average was calculated. The mean,

standard deviation and relative standard deviation were evaluated for

interpretation/comparison of data.

54

CHAPTER 4

RESULTS

In this chapter the results obtained from this research are presented. In the analysis of

iodine by the spectrophotometric kinetic method, the absorbance of the blanks,

standards and samples were automatically monitored. As discussed, the kinetic

reaction is initiated as soon as As3+ is added to the reaction mixture. In this reaction,

yellow Ce4+ is reduced to colourless Ce3+ by As3+. Usually this reaction is very slow.

Iodide helps in catalysing this reaction making the reaction faster and has been used

for the determination of iodine in different food samples as presented in this chapter.

4.1. Analysis of blanks

The analysis of blanks was carried out by adding the following in a cuvette, 0.25 mL

of Milli-Q-water, 0.25 mL of H2SO4- HCl mixture, 0.25 mL Ce4+ reagent and 0.25 mL

As3+ reagent. The contents were mixed for 10 sec and the cuvette was pre-incubated

-Q-water) was added at the end to initiate

the reaction. The decrease in the absorbance of the blank was monitored for 1 min at

370 nm as shown in Figure 4 below.

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

1.1

1.2

0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0

Abs

orba

nce(

AU

)

Time (Min)

BlankCurve o

Figure 4. Typical absorbance time curve for the blank analysis up to 1 min.

55

The allowable change in the absorbance per min for the blanks was in the range of

0.185 - 0.192. This was derived from Table 7 below in which the A/min for the blank

was analysed 7 times (n = 7). Method blanks as indicated in Table 7 below were used

to identify and correct systematic errors due to impurities in the reagent and

contamination in the glasswares and the instrumentation. The standard deviation of the

blank measurements was closely monitored.

Table 7. Change in absorbance in blank analysis (0 ng/mL - iodine).

Number of analysis (n) A/min

1 0.187

2 0.191

3 0.192

4 0.189

5 0.191

6 0.185

7 0.191

Average 0.189

Standard deviation (SD) 0.003

Relative standard deviation (RSD) (%) 1.36

The RSD for the blank analysis which was 1.36% is a measure of the sources of error

in the analysis and thus was used to measure the analytical coefficient of variation.

The data recorded with SD of 0.003 and a RDS of 1.36% show an exceptional system

stability.

4.2. Time-absorbance curves

The kinetic assay involved the following procedure: 0.25 mL of Milli-Q-water, 0.25

mL of H2SO4-HCl mixture, 0.25 mL Ce4+ reagent and 0.25 mL As3+ reagent were

separately transferred to a 10 mm path length cuvette. The contents in the cuvette were

mixed for 10 sec and the cuvette was pre- min. A 0.25 mL

volume of the catalyst iodine (0 - 25 ng/mL) was added to initiate the reaction. The

absorbance of the standards was recorded at 370 nm exactly up to 1 min. A total of 7

runs (n = 7) was carried out for each standard and then the average absorbance was

56

determined which was then plotted against the time to get the calibration curve. A

typical absorbance versus time curve which was recorded at 370 nm for 1 min at

different iodine concentrations from 0 to 25 ng/mL is shown in Figure 5.

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

1.1

1.2

0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0

Absorb

ance(A

U)

Time (Min)

BlankIodine 2.5 ng/mLIodine 5 ng/mLIodine 10 ng/mLIodine 15 ng/mLIodine 20 ng/mLIodine 25 ng/mLCurve of BlankCurve of Iodine 2.5Curve of Iodine 5 nCurve of Iodine 10Curve of Iodine 15Curve of Iodine 20Curve of Iodine 25

Figure 5. Typical absorbance – time recording of the catalysed reaction up to 1 min

at different iodine concentrations of 0, 2.5, 5, 10, 15, 20 and 25 ng/mL at 370 nm at

57

Table 7. Absorbance analysis at different iodine concentrations and different times.

Time (min)

Number of analysis (n)

Absorbance values (A) at different iodine concentrations (ng/mL)

Blank 2.5 5.0 10.0 15.0 20.0 25.0

0.01

1.002 0.998 1.003 1.003 0.997 1.003 0.9990.5 0.905 0.892 0.887 0.875 0.851 0.855 0.8181.0 0.815 0.792 0.783 0.753 0.729 0.719 0.667

0.02

1.001 0.999 0.999 0.996 0.996 0.996 1.0050.5 0.904 0.899 0.894 0.872 0.858 0.831 0.8331.0 0.810 0.797 0.791 0.762 0.728 0.699 0.688

0.03

1.001 0.999 0.998 0.994 0.990 0.997 0.9960.5 0.900 0.898 0.892 0.871 0.849 0.837 0.8221.0 0.810 0.800 0.792 0.760 0.723 0.697 0.663

0.04

1.007 1.000 0.996 0.997 1.000 1.000 1.0040.5 0.905 0.897 0.876 0.872 0.861 0.845 0.8361.0 0.817 0.799 0.771 0.756 0.732 0.709 0.686

0.05

0.997 1.002 0.998 1.000 1.000 0.997 1.0000.5 0.901 0.896 0.883 0.868 0.858 0.843 0.8281.0 0.806 0.794 0.776 0.759 0.730 0.707 0.680

0.06

1.003 1.005 0.999 0.999 0.996 0.999 0.9990.5 0.903 0.904 0.884 0.865 0.856 0.835 0.8271.0 0.818 0.806 0.775 0.747 0.730 0.692 0.676

0.07

1.006 0.995 0.998 1.001 0.997 1.007 1.0070.5 0.905 0.891 0.883 0.870 0.857 0.846 0.8341.0 0.814 0.791 0.778 0.757 0.726 0.704 0.683

From the time versus absorbance data shown in Table 8 above, the Tables in

Appendices 1, 2 and 3 were generated at different times 0, 0.5 and 1 min, respectively

which show the statistical control of the data obtained.

The statistical data analysis (Appendices 1, 2 and 3) from the absorbance at different

iodine concentrations shows that the relative standard deviation (RSD) for all the

standards analysed at the three time intervals (0, 0.5 and 1 min) were in the range of

58

0.2 - 1.4 %. This relative standard deviation (0.2 - 1.4%) shows exceptional system

stability. Based on the absorbance data above, the average absorbance at the 3 time

intervals (0, 0.5 and 1 min) were compiled together and tabulated in Table 9.

Table 8. The average absorbances at different iodine concentrations from 0 to 25

ng/mL at three different times (n = 7).

Time

(Min)

Average absorbance (A) at different iodine concentrations (ng/mL)

Blank 2.5 5.0 10.0 15.0 20.0 25.0

0.0 1.000 1.000 1.000 1.000 1.000 1.000 1.000

0.5 0.903 0.897 0.886 0.870 0.856 0.842 0.828

1.0 0.813 0.797 0.781 0.756 0.728 0.704 0.678

The absorbance versus time graph (Figure 6) was plotted using the average absorbance

data shown in Table 9. Figure 6 clearly shows that all absorbances for the blank and

other standards start at absorbance 1.000 and finish at their respective absorbances

exactly after 1 min. The absorbances were all measured at a wavelength of 370 nm and

59

4.3. Calibration curves

the different iodine concentrations were calculated from Figure

6 and computed in Table 10 from seven different measurements. The calibration curve

was then plotted from the change in the average i.e. fixed

time procedure was followed and is shown in Figure 7. The change in absorbance per

min) is also known as the initial rate.

R² = 0.9996

R² = 0.9999

R² = 0.9993

R² = 0.9987

R² = 0.9987

R² = 0.9984

R² = 0.9986

0.600

0.700

0.800

0.900

1.000

1.100

0 0.5 1

Abs

orba

nce

at 3

70 n

m

Time (Min)

Blank

Iodine 2.5 ng/mL

Iodine 5 ng/mL

Iodine 10 ng/mL

Iodine 15 ng/mL

Iodine 20 ng/mL

Iodine 25 ng/mL

Figure 6. Plot of average absorbance at 370 nm for the reduction of Ce4+ by As3+

against time in the presence of different iodine concentrations of 0, 2.5, 5, 10, 15, 20

and 25 ng/mL at analysis time of

60

Table 9. Chang min).

Iodine

conc.

(ng/mL)

Average

n = 1 n = 2 n = 3 n = 4 n =5 n = 6 n = 7

Blank 0.187 0.191 0.192 0.189 0.191 0.185 0.191 0.189 ± 0.003

2.5 0.205 0.202 0.199 0.201 0.207 0.199 0.204 0.202 ± 0.003

5.0 0.220 0.208 0.205 0.224 0.221 0.224 0.220 0.217 ± 0.008

10.0 0.250 0.234 0.234 0.240 0.241 0.251 0.244 0.242 ± 0.007

15.0 0.268 0.268 0.267 0.269 0.270 0.266 0.271 0.268 ± 0.002

20.0 0.284 0.297 0.299 0.291 0.290 0.306 0.303 0.296 ± 0.008

25.0 0.332 0.317 0.333 0.318 0.320 0.323 0.321 0.323 ± 0.007

R² = 0.9998

0.150

0.170

0.190

0.210

0.230

0.250

0.270

0.290

0.310

0.330

0.350

0 5 10 15 20 25Iodine Concentration (ng/mL)

Figure 7. Calibration curve i.e. plot of average change in absorbance per minute

n.

61

4.4. Food samples ashing

Decomposition of organic matter is generally based on acid or alkaline digestion. Acid

digestion methods as discussed in the literature are generally effective in the

destruction of organic matter but are not recommended for routine analytical purposes

due to the fact that they require large amounts of concentrated acids for each sample

and they are hazardous to humans from the fumes they emit (Nitschke and Stengel,

2015; Acland, 1957; Aumont and Tressol, 1987; Aumont and Tressol, 1986;

Patzeltová, 1993; Leufroy et al., 2015; Romarís–Hortas et al., 2011; Eckhoff and

Maage, 1997; Haldimann et al., 2005; Cressey, 2003; Shah et al., 2005). The use of

acid digestion becomes impractical when analysing a large number of samples

manually. A number of papers as discussed in the literature uses alkaline dry ashing

for the analysis of iodine in biological materials and food stuffs (Nitschke and Stengel,

2015; Acland, 1957; Aumont and Tressol, 1987; Aumont and Tressol, 1986; Mahesh

et al., 1992). The use of 30% K2CO3 and 10% ZnSO4 for alkaline dry ashing resulted

in incomplete ashing and low recoveries of iodine from plants and foodstuffs (Mahesh

et al., 1992).

As reported by Mahesh et al. (1992), even the use of sodium carbonate (Na2CO3) as

an ashing agent gave poor recoveries. Incomplete ashing and poor recoveries were also

observed when KOH was used as an ashing agent (Mahesh et al., 1992). Thus, to

overcome all these problems, Mahesh et al. (1992) used 0.1 mL 6 M KOH and 0.1 mL

0.5 M ZnSO4 and ashing was carried out in two steps: 1 hr with KOH and 2 hrs with

ZnSO4. All their samples were therefore ashed for exactly 3 hrs.

In the current study, initially poor recoveries were obtained when the procedure

specified by Mahesh et al. (1992) was followed. The poor recoveries were attributed

to incomplete ashing and the presence of organic matter. Thus, to overcome this

problem, a modified alkaline ashing procedure reported by Nitschke and Stengel

(2015) was referred to in which the authors used < 200 mg of sample and 400 μL of

17 M KOH s. This was followed by 4 hrs decomposition

62

Finally, to overcome the poor recoveries, all the food samples in the present study were

analysed by a modified ashing procedure as discussed in the methodology section (cf.

section 3.6).

4.5. Sample analysis

Sample analysis was carried out in a similar manner as the blanks and standard

solutions were analysed. A typical absorbance versus time curve of some food samples

analysed at 370 nm for 1 min is shown in Figure 8. Of the 22 samples analysed, each

sample was analysed 4 times (n = 4) and the average iodine content calculated from

those determinations. The standard deviation, coefficient of variation and the

confidence interval were also calculated to evaluate the overall performance of the

method as indicated in Tables 11 -17.

63

0.00.20.40.60.81.01.21.4

0.00.1

0.20.3

0.40.5

0.60.7

0.80.9

1.0

Absorbance(AU)

Time (

Min)

Blan

k Ana

lysis

Dalo

Rew

a butt

er P

owde

red m

ilkLu

miw

awa

Lettu

ce

Seag

rape

s Ch

iken e

gg

Chee

se

Clam

Cu

rve of

Blan

k Ana

lysis

Curve

of D

alo

Curve

of R

ewa b

utter

Curve

of P

owde

red m

ilkCu

rve of

Lum

iwaw

a Cu

rve of

Lettu

ce

Curve

of S

eagr

apes

Cu

rve of

Chik

en eg

g Cu

rve of

Che

ese

Curve

of C

lam

Figu

re 8

.Typ

ical

UV

-vis

ible

reco

rdin

g of

abs

orba

nce

agai

nst t

i.

64

The iodine contents in different brands of rice are reported in Table 11. The iodine

contents in different brands of rice are also been presented in the form of bar diagram

as shown in Figure 9. The highest iodine content was present in Sunwhite Calrose rice

with the mean value of 195.47 ± 0.19 ng/g.

Table 10. Iodine contents in different brands of rice (Oryza sativa) analysed on a fresh

weight basis.

Sample Iodine content Mean ± SDa

(ng/g)

Coefficient of variation (%)

Confidence intervalb

Punjas Jasmine rice 65.11 ± 0.09 3.83 0.14

FMF Sungrown rice 53.89 ± 0.06 3.10 0.09

Punjas Long Grain rice 85.19 ± 0.11 3.77 0.18

Sunwhite Calrose rice 195.47 ± 0.19 2.74 0.30a Mean for four determinations (n = 4)b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

Figure 9. Graphical representation of iodine contents in different brands of rice

analysed on a fresh weight basis.

65.1

1

53.8

9 85.1

9

195.

47

0

50

100

150

200

250

Punjas Jasmine rice FMF Sungrown rice Punjas Long Grainrice

Sunwhite Calroserice

Iodi

ne c

onte

nt (n

g/g)

65

The iodine contents in different root crops are reported in Table 12 and graphically

presented in form of bar diagram as shown in Figure 10. The variation in the results

for the analysed different food samples (potato, cassava and dalo/taro) can be clearly

seen.

Table 11. Iodine contents in different root crops analysed on a fresh weight basis.

Sample (Scientific name)Iodine content Mean ± SDa

(ng/g)



Potato (Solanum tuberosum)

MH supermarket 265.60± 0.18 1.88 0.28

Shop and Save supermarket 260.45 ± 0.30 3.19 0.47

New World supermarket 262.17 ± 0.37 3.96 0.59

Potato Market 235.23 ± 0.21 2.51 0.34

Cassava (Manihot esculenta)

Cassava vendor 1 345.91 ± 0.22 1.78 0.35

Cassava vendor 2 219.64 ± 0.12 1.50 0.19

Cassava vendor 3 235.75 ± 0.16 1.84 0.25

Cassava vendor 4 249.75 ± 0.26 2.92 0.41

Dalo/Taro (Colocasia esculenta)

Taro vendor 1 305.09 ± 0.30 2.74 0.47

Taro vendor 2 335.34 ± 0.28 2.33 0.44

Taro vendor 3 379.85 ± 0.32 2.38 0.51

Taro vendor 4 227.43 ± 0.20 2.48 0.32a Mean for four determinations (n = 4) b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

66

Figure 10. Graphical representation of iodine contents in different root crops analysed

on a fresh weight basis.

The iodine contents in different fish/meat products are reported in Table 13 and

graphically presented in form of bar diagram in Figure 11. The variation in results for

the investigated different brands of food samples (fresh marine fish, clam, canned tuna,

canned sardine and chicken eggs) is quite evident.

Table 12. Iodine contents in different fish/meat products analysed on a fresh weight

basis.


(ng/g)



Daisy parrot fish (Chlorurus sordidus) 966.55 ± 0.42 1.90 0.66

Malabar grouper (Soisoi) (Epinephelus Malabarcius) 1048.32 ± 0.93 3.88 1.47

Russell's snapper fish (Kwake) (Lujanus russelli) 1069.91 ± 0.74 3.04 1.18

Pacific yellow tail emperor fish (Sabutu) (Lethrinus atkinsoni) 1088.17 ± 0.89 3.59 1.41

265.

6

260.

45

262.

17

235.

25

345.

91

219.

64

235.

75

249.

75 305.

09

335.

34

379.

85

227.

43

050

100150200250300350400

Iodi

ne c

onte

nt (n

g/g)

67

Clam (Margaritifera)

Clam vendor 1 449.72 ± 0.52 3.21 0.82

Clam vendor 2 483.40 ± 0.25 1.45 0.40

Clam vendor 3 587.74 ± 0.53 2.52 0.84

Clam vendor 4 479.04 ± 0.63 3.70 1.01

Canned tuna

Sunbell Tuna in vegetable oil 660.77 ± 0.68 2.89 1.09

Burnswick Tuna in vegetable oil 313.55 ± 0.34 2.99 0.53

Sunbell Ovalau Blue (Light Tuna flakes) 514.04 ± 0.68 3.68 1.07

Skipper Tuna in vegetable oil 659.32 ± 0.25 1.07 0.40

Canned sardine

Burnswick Sardine in vegetable oil 534.11 ± 0.54 2.83 0.86

Burnswick Sardine in Spring Water 956.15 ± 0.42 2.10 0.67

Burnswick Sardine in Tomato Sauce 504.92 ± 0.21 1.15 0.33

Burnswick Sardine in Lemon Sauce 351.45 ± 0.42 3.37 0.67

Chicken EggRam Sami & Sons egg 612.96 ± 0.83 3.78 1.32

Egg vendor 2 591.79 ± 0.34 1.93 0.53

Egg vendor 3 603.58 ± 0.69 3.21 1.10

Egg vendor 4 1112.08 ± 0.03 0.16 0.05a Mean for four determinations (n = 4)b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

68

Figure 11. Graphical representation of iodine contents in different fish/meat products


The iodine contents in different dairy products are reported in Table 14 and also

presented in form of bar diagram in Figure 12. Table 14 and Figure 12 clearly show

the variation in results for the different food samples (cheese, fresh liquid milk,

processed powdered milk and butter/margarine).

966.

55 1048

.32

1069

.91

1088

.17

449.

72

483.

4

587.

74

479.

04

660.

77

313.

55

514.

04

659.

32

534.

1195

6.15

504.

92

351.

45

612.

96

591.

79

603.

5811

12.0

8

0

200

400

600

800

1000

1200

Iodi

ne c

onte

nt (n

g/g)

69

Table 13. Iodine contents in different dairy products analysed on a fresh weight basis.

Sample Iodine content Mean ± SDa

(ng/g)



Cheese

Rewa Tasty (Fiji Dairy cheese) 191.64 ± 0.27 3.91 0.43

Lemnos cheese 490.79 ±0.29 1.67 0.47

Chesdale cheese 406.92 ± 0.26 1.82 0.42

Devondale cheese 420.92 ± 0.24 1.60 0.38

Fresh liquid milk

Rewa Life full cream milk 210.53 ± 0.25 3.27 0.39

Anchor Regular milk 158.36 ± 0.08 1.48 0.13

Meadow Fresh milk 242.09 ± 0.29 3.31 0.45

Fresh cow milk - unprocessed 339.83 ± 0.33 2.74 0.53

Processed milk

Rewa full cream milk powder 471.77 ± 0.41 2.42 0.65

Redcow full cream milk powder 397.02 ± 0.29 2.03 0.46

Rewa Skim milk powder 655.89 ± 0.03 0.14 0.05

Dairy Fresh full cream milk powder 795.49 ± 1.07 3.76 1.70

Butter/Margarine

Rewa butter 124.02 ± 0.15 3.32 0.23

Anchor butter 313.94 ± 0.39 3.49 0.62

Flora margarine 263.49 ± 0.11 1.15 0.17

Meadowlea margarine 172.62 ± 0.15 2.48 0.24a Mean for four determinations (n = 4)b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

70

Figure 12. Graphical representation of iodine contents in different dairy products


The determined iodine contents in different leafy vegetables are reported in Table 15.

The graphical representation of these data is shown in Figure 13 which clearly depicts

the variation in the results for the tested food samples that include lettuce, English

cabbage and Chinese cabbage.

Table 14. Iodine contents in commonly consumed leafy vegetables analysed on a fresh

weight basis.


(ng/g)



Lettuce (Lactuca sativa)

Lettuce vendor 1 178.17 ± 0.14 2.16 0.22

Lettuce vendor 2 40.15 ± 0.03 2.15 0.05

Lettuce vendor 3 81.75 ± 0.05 1.86 0.09

Lettuce vendor 4 159.15 ± 0.10 1.85 0.17

191.

64

490.

79

406.

92

420.

92

210.

53

158.

36 242.

09 339.

83 471.

77

397.

02

655.

89 795.

4912

4.02

313.

94

263.

49

172.

62

0100200300400500600700800900

Iodi

ne c

onte

nt (n

g/g)

71

English cabbage (Brassica oleracea)

English cabbage vendor 1 56.92 ± 0.03 1.39 0.05




Chinese cabbage (Brassica chinensis)

Chinese cabbage vendor 1 44.64 ± 0.04 2.27 0.06




a Mean for four determinations (n = 4)b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

Figure 13. Graphical representation of iodine contents in commonly consumed leafy

vegetables analysed on a fresh weight basis.

The iodine contents determined in different fruits and vegetables are reported in Table

16. The graphical representation of the determined iodine contents is shown in Figure

14. The data obtained clearly depicts the variation in the iodine contents for the

investigated food samples such as tomato, banana, long bean and pumpkin.

178.

1740

.15

81.7

5

159.

15

56.9

2

143.

7

87.1

7

145.

81

44.6

4

126 132.

87

112.

530

20406080

100120140160180200

Iodi

ne c

onte

nt (n

g/g)

72

Table 15. Iodine contents in commonly consumed fruits and vegetables analysed on a

fresh weight basis.


(ng/g)



Tomato (Solanum lycopersicum)

Tomato vendor 1 24.17 ± 0.02 2.09 0.03

Tomato vendor 2 48.34 ± 0.04 2.60 0.04

Tomato vendor 3 35.13 ± 0.06 4.51 0.09

Tomato vendor 4 53.62 ± 0.04 1.89 0.06

Banana (Musa)

Banana vendor 1 29.85 ± 0.06 5.31 0.61

Banana vendor 2 21.40 ± 0.04 4.73 0.13

Banana vendor 3 183.06 ± 0.23 3.59 0.37

Banana vendor 4 70.40 ± 0.07 2.83 0.33

Long bean (Vigna unguiculata ssp. Sesquipedalis)

Long bean vendor 1 37.64 ± 0.04 3.11 0.07

Long bean vendor 2 183.98 ± 0.19 2.87 0.30

Long bean vendor 3 105.40 ± 0.06 1.56 0.09

Long bean vendor 4 63.40 ± 0.12 5.14 0.19

Pumpkin (Cucurbita moschata)

Pumpkin vendor 1 48.87 ± 0.03 1.87 0.05

Pumpkin vendor 2 63.92 ± 0.03 1.51 0.05

Pumpkin vendor 3 183.58 ± 0.20 3.03 0.32

Pumpkin vendor 4 108.57 ± 0.05 1.22 0.08


73

Figure 14. Graphical representation of iodine contents in commonly consumed fruits

and vegetables analysed on a fresh weight basis.

The iodine contents in different seaweeds are reported in Table 17 and graphically

presented in Figure 15. The variation in the results for the studied food samples such

as sea grapes and lumiwawa is clearly evident. The iodine contents in lumiwawa are

relatively much higher than sea grapes.

Table 16. Iodine contents in commonly consumed seaweeds analysed on a fresh

weight basis.


(ng/g)



Sea grapes (Caulerpa lentillifera)

Sea grapes vendor 1 1359.06 ± 0.63 2.77 1.00




24.1

7 48.3

4

35.1

3 53.6

2

29.8

5

21.4

183.

0670

.4

37.6

4

183.

9810

5.4

63.4

48.8

7 63.9

2

183.

5810

8.57

0

20

40

60

80

100

120

140

160

180

200

Iodi

ne c

onte

nt (n

g/g)

74

Lumiwawa (Gracilaria maramae)

Lumiwawa vendor 1 9185.72 ± 0.89 3.78 1.42

Lumiwawa vendor 2 11000.00 ± 0.32 1.28 0.51

Lumiwawa vendor 3 2868.81 ± 0.25 3.72 0.40

Lumiwawa vendor 4 2438.68 ± 0.10 1.72 0.15a Mean for four determinations (n = 4)b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

Figure 15. Graphical representation of iodine contents in commonly consumed

seaweeds analysed on a fresh weight basis.

The iodine contents presented in Tables 11 to 17 have clearly demonstrated that there

has been great variability in the iodine contents in different food samples of different

origin.

4.6. Precision

For all the samples analysed, the coefficient of variation was calculated. It has been

reported that a coefficient of variation less than 6.0% with a mean and standard

deviation of 3.4 ± 1.77% for more than 20 foodstuff analysed on three or more

occasions was sufficient to judge the precision. (Mahesh et al., 1992). The coefficient

of variation for the sample analysis in our case was also less than 6.0 % with a mean

and standard deviation of 2.57 ± 0.28% for the 22 samples analysed each four times.

Thus this method was quite precise. The analytical coefficient of variation in the

1359

.06

1525

.83

914.

42

851.

92

9185

.72

1100

0

2868

.81

2438

.68

0

2000

4000

6000

8000

10000

12000

Iodi

ne c

onte

nt (n

g/g)

75

present study was worked out to be 0.54% for the 22 food samples analysed which

showed exceptional system analytical stability.

4.7. Limit of detection

The LOD was calculated by multiplying the sample standard deviation by the student’s

t value. Thus, for seven replicates and six degrees of freedom, the student’s t was taken

as 3.143 at 98% confidence level (Wisconsin Department of Natural Resources

Laboratory Certification Program, 1996). The data from the blank analysis were used

for the LOD calculation. The change in the absorbance per min ( i.e. initial

rate for the blank analysis is shown in Table 18. The standard deviation was calculated

to be 0.49 which was used for the LOD calculation.

Table 17. Change in the absorbance per minute for 7 runs (n = 7) for blank analysis (0

ng/mL –iodine), with standard deviation for LOD and LOQ determination.

Number of analysis (n)

Iodine concentration (ng/mL)

Iodine recovery (ng/mL)

1 0 0.187 -0.382 0 0.191 0.383 0 0.192 0.574 0 0.189 0.005 0 0.191 0.386 0 0.185 -0.757 0 0.191 0.38

Standard deviation (SD) 0.49

Using the standard deviation obtained in Table 18, the LOD was calculated as follows:

LOD = 3.143 × standard deviation of analysis (n = 7)

LOD = 3.143 × 0.49 ng/mLLOD = 1.54 ng/mL

76

4.8. Limit of quantification

The LOQ was determined following the method reported by Wisconsin Department of

Natural Resources Laboratory Certification Program (1996) as shown below:

LOQ = 10 × standard deviation of analysis (n = 7)LOQ ngmL = 10 × 0.49 ng/mL

LOQ = 4.9 ng/mL4.9. Quality control

Under quality control, the following aspects were accounted for:

4.9.1. Analysis of duplicate samples for food sample analysis

As indicated in Table 11 – 17, each food sample was analysed 4 times (n= 4) from

which the average iodine contents were calculated. The coefficient of variation and the

confidence interval were also calculated alongside each sample to measure the sources

of errors and the degree of variation.

4.9.2. Recovery analysis from standard samples

The analysis of known concentrations of iodine was used to monitor the statistical

control. Thus, iodine concentrations of 4, 12 and 18 ng/mL were used for the recovery

study. Iodine concentrations were calculated using the calibration equation shown

below which was obtained from the calibration curve shown in Figure 7. The recovery

ranged from 99.84 – 100.24% showing excellent quality control which is presented in

Table 19.

A = 0.0053[I] + 0.189[I] = A 0.1890.0053

77

Where: [I] = iodine concentration

0.189 = intercept on Y- axis and

0.0053 = slope of the calibration curve i.e. sensitivity.

Table 18. Analysis of standard iodine solutions at 4, 12, and 18 ng/mL and their

recovery.

Iodine

concentration

(ng/mL)

Nominal ± SDaRecovery ± RSD

(%)

Standard

analytical error

(%)

Confidence

intervalb

4.0 4.01 ± 0.24 100.24 ± 5.92 0.60 0.38

12.0 11.98 ± 0.11 99.84 ± 0.91 0.23 0.17

18.0 18.02 ± 0.47 100.10 ± 2.64 0.88 0.76

4.9.3. Spike recoveries from real samples

Recovery study was further carried out using real samples. The samples that were

previously analysed were randomly selected from different categories. This included

Punjas Jasmine rice, Ram Sami and Sons egg, Rewa Full Cream powdered milk,

Burnswick sardine and Anchor butter. For the recovery analysis, iodine in the form of

KI at concentrations 4, 12 and 18 ng/mL were added to each sample and the ashing

procedure was applied followed by the kinetic assay. The percentage recovery was

calculated using the following equation:

Recovery (%) = Concentration of iodine found by analysisConcentration of total in sample × 100The recovery results from the spiked real samples are shown in Table 20. The recovery

of iodine added to different food samples ranged from 97.42 ± 3.41% to 103.13 ±

4.76% with an average recovery of 100.18 ± 3.02% (mean ± standard deviation). The

results presented in Table 20 clearly demonstrated the method used produced quality

data where average recovery was excellent as 100.18%.


78

Table 19. Recovery of iodine from different food samples after spiking with 4, 12 and

18 ng/mL iodine.

SampleSpiked concentration (ng/mL)

Nominal ± SDa Recovery ± RSD (%)

Standard analytical error (%)


Punjas Jasmine rice

Unspiked 65.11 ± 0.09 - 0.23 0.14

4.0 67.87 ± 0.08 98.20 ± 4.95 0.21 0.12

12.0 77.92 ± 0.15 101.05 ± 5.68 0.40 0.2518.0 85.72 ± 0.14 103.13 ± 4.76 0.37 0.23

Chicken Egg Unspiked 612.96 ± 0.83 - 1.44 1.324.0 646.04 ± 0.37 101.81 ± 1.24 0.38 0.3112.0 625.96 ± 0.56 100.16 ± 3.87 1.15 0.9118.0 621.40 ± 0.45 98.48 ± 3.15 0.91 0.72

Rewa powdered milk

Unspiked 471.77 ± 0.41 - 0.78 0.654.0 464.75 ± 1.88 97.68 ± 1.88 0.55 0.4412.0 489.40 ± 0.20 101.16 ± 1.77 0.42 0.3118.0 492.64 ± 0.29 100.59 ± 2.30 0.60 0.46

Burnswick sardine in vegetable oil

Unspiked 534.11 ± 0.54 - 0.99 0.864.0 552.64 ± 0.21 102.70 ± 1.52 0.43 0.3312.0 558.51 ± 0.54 102.27 ± 4.26 1.12 0.86

18.0 545.02 ± 0.37 98.72 ± 3.01 0.78 0.59

Anchor butter Unspiked 313.94 ± 0.39 - 0.83 0.624.0 309.74 ± 0.29 97.42 ± 3.41 0.64 0.4512.0 319.49 ± 0.08 98.02 ± 0.72 0.17 0.1318.0 336.40 ± 0.33 101.34 ± 2.83 0.70 0.53

Average 100.18 ± 3.02 0.66 0.51a Mean for four determinations (n = 4) b Confidence level at 95% with 3 degrees of freedom (t = 3.182)

79

4.9.4. Analysis of standard reference materials (SRM)

Standard Reference Material Iodised Table Salt (SRM No. 3530) was purchased from

the National Institute of Standards and Technology (NIST), USA and analysed to

evaluate the accuracy of the kinetic spectrophotometric method used for iodine

determination. The adequacy of applied methodology was verified by usual measures

of accuracy, reproducibility, and recovery in which 300 mg of the SRM (Iodised Salt)

was dissolved in 1L of Milli-pore water. The sample was then placed in an ultrasonic

bath for 10 min to facilitate complete dissolution of the salt material prior to kinetic

spectrophotometric analysis. The typical absorbance against time recording for the

blank and NIST SRM No. 3530 is shown in Figure 16. The recovery results obtained

as per calibration graph in terms of ng/mL which was further converted to mg/kg as

reported in NIST SRM No. 3530 certificate is shown in Table 21 and Table 22.

Figure 16. Typical UV-visible spectra of blank and NIST Standard Reference Material

(SRM No. 3530 - Iodised Salt) anal .

80

Table 20. Summary of the recovery results obtained from NIST SRM No. 3530 –

Iodised Salt analysis.

Iodine concentration

(ng/mL)

Iodine calculated ± SDa

(ng/mL)

Iodine recovery ± RSD

(%)

Standard analytical error (%)


15.66 15.61 ± 0.37 99.68 ± 2.38 0.72 0.44

a Mean for seven determinations (n = 7)b Confidence level at 95% with 6 degrees of freedom (t = 3.143)

Table 22 compares the results obtained from the SRM analysis by the Sandell-Kolthoff

kinetic spectrophotometric method to the certified value of the iodine content of the

Iodised Table Salt - 52.2 ± 4.2 mg/kg analysed by ICP-MS at NIST, USA according

to the certificate for SRM No. 3530. The recovery data presented in Table 22 shows

great accuracy and are well within the error range which is 99.68 ± 2.38 %. Thus, the

method applied in the present study is very reliable for the determination of iodine

contents in food samples.

Table 21. NIST SRM No. 3530 iodine recovery using the spectrophotometric kinetic

method.

NIST SRM No. 3530

Certified value ± SD(mg/kg)

Found value Mean ± SDa

(mg/kg)

Recovery ± RSD (%)

Iodised Table Salt (Iodide) 52.2 ± 4.2 52.03 ± 0.37 99.68 ± 2.38

a Mean for seven determinations (n = 7)

81

CHAPTER 5

DISCUSSION

This chapter explains the results obtained in the analysis of the food samples. The

average, minimum and the maximum iodine contents in different commonly consumed

food groups are also discussed. The results are expressed in graphical format for easier

comparison\representation. Iodine contents in foods have been further compared to

previously published data and comparison is made with the similarities or the

differences seen in the results obtained.

5.1. Discussion of results obtained

The twenty two commonly consumed food samples were successfully analysed.

Average values for each type of food sample together with the mean, minimum and

maximum iodine contents were calculated with the average standard deviation

associated with each mean value and shown category wise in Tables 23 – 28 and

collectively in Table 29. These data were used to compare the results from previous

studies to see if the present determined iodine contents correspond to the previously

reported values on the determination of iodine contents in foods analysed on a fresh

basis.

The mean iodine contents in commonly consumed rice and root crops are reported in

Table 23. Rice had a mean iodine content 99.92 ± 0.11 ng/g. The lowest or minimum

iodine content in rice was 53.89 ± 0.06 ng/g whereas the maximum iodine content was

found to be 195.47 ± 0.19 ng/g. Potato had a determined mean iodine content of 255.87

± 0.27 ng/g with the minimum of 235.25 ± 0.21 ng/g and the maximum of 265.60 ±

0.18 ng/g. Cassava, had a mean iodine content of 262.76 ± 0.19 ng/g for the 4 different

samples analysed. The minimum iodine content was found to be 219.64 ± 0.12 ng/g

while the highest was 345.91 ± 0.22 ng/g. Dalo/taro showed the highest mean analysed

iodine content of 311.93 ± 0.28 ng/g with a minimum iodine content of 227.43 ± 0.20

ng/g and the highest analysed iodine content of 379.85 ± 0.32 ng/g. The mean iodine

contents in commonly consumed rice and root crops are also presented in the form of

a bar diagram as shown in Figure 17. Figure 17 clearly shows that dalo/taro had the

highest mean iodine content of 311.93 ± 0.28 ng/g and rice had the lowest mean iodine

82

content of 99.92 ± 0.11 ng/g. Thus, this gives an indication that dalo, cassava and

potato are preferred over rice to increase the daily dietary iodine intake.

Table 22. Mean iodine contents in commonly consumed rice and root crops analysed


Fresh samples (Scientific name)

No. of samples analysed (n)

Iodine content (ng/g)

Mean ± SDa Minimum (Mean ± SDa)

Maximum (Mean ± SDa)

Rice(Oryza sativa) 4 99.92 ± 0.11 53.89 ± 0.06 195.47 ± 0.19

Potato (Solanum tuberosum) 4 255.87 ± 0.27 235.25 ± 0.21 265.60 ± 0.18

Cassava(Manihot esculenta) 4 262.76 ± 0.19 219.64 ± 0.12 345.91 ± 0.22

Dalo/Taro (Colocasia esculenta) 4 311.93 ± 0.28 227.43 ± 0.20 379.85 ± 0.32


rice and root crops analysed on a fresh weight basis.

99.9

2

255.

87

262.

76

311.

93

0.00

50.00

100.00

150.00

200.00

250.00

300.00

350.00

Rice Potato Cassava Dalo/Taro

Iodi

ne c

onte

nt (n

g/g)

a Mean of four samples of each food analysed four times (n = 4)


83

The mean iodine contents in commonly consumed fish/meat are reported in Table 24.

Fresh marine fish had the highest mean iodine content with a reported value of 1043.24

± 0.75 ng/g. The highest iodine content analysed was 1088.17 ± 0.89 ng/g and the

lowest iodine content was found to be 966.55 ± 0.42 ng/g. Clam had a mean iodine

content of 499.98 ± 0.48 ng/g with a minimum of 449.72 ± 0.52 ng/g and a maximum

of 587.44 ± 0.53 ng/g. The mean iodine content in canned sardine was 586.66 ± 0.40

ng/g with the minimum mean of 351.45 ± 0.42 ng/g and the highest mean of 956.15 ±

0.42 ng/g. Chicken egg had the analysed mean iodine content of 730.10 ± 0.47 ng/g

with a minimum mean of 591.79 ± 0.34 ng/g and the highest mean iodine content of

1112.08 ± 0.03 ng/g. The mean iodine contents in commonly consumed fish/ meat are

also presented in the form of a bar diagram as shown in Figure 18. The bar diagram

clearly depicts that fresh marine fish shows the highest mean iodine with a reported

value of 1043.24 ± 0.75 ng/g. The clam with the mean iodine content 499.98 ± 0.48

ng/g shows the least. Thus, it can be concluded that fresh marine fish and chicken eggs

present a good source of iodine, followed by others from this group of commonly

consumed foods analysed.

Table 23. Mean iodine contents in commonly consumed fish/meat products analysed



No. of samples analysed

(n)




Fresh marine fish 4 1043.24 ± 0.75 966.55 ± 0.42 1088.17 ± 0.89 Clam(Margaritifera) 4 499.98 ± 0.48 449.72 ± 0.52 587.44 ± 0.53

Canned tuna 4 536.92 ± 0.49 313.55 ± 0.34 660.77 ± 0.68

Canned sardine 4 586.66 ± 0.40 351.45 ± 0.42 956.15 ± 0.42

Chicken egg 4 730.10 ± 0.47 591.79 ± 0.34 1112.08 ± 0.03



84


fish/meat analysed on a fresh weight basis.

The mean iodine contents in commonly consumed dairy products are shown in Table

25. In this group of foods analysed, processed powdered milk showed the highest mean

iodine content of 580.04 ± 0.45 ng/g. The lowest mean iodine content was found to be

397.02 ± 0.29 ng/g and the highest iodine content was found to be 795.49 ± 1.07 ng/g

in the same food product. Cheese had the second highest iodine content in this group

which had the mean iodine content of 377.57 ± 0.27 ng/g with a minimum mean of

191.64 ± 0.27 ng/g and a maximum mean iodine content of 490.79 ± 0.29 ng/g. Next

was fresh liquid milk with a mean iodine content of 237.70 ± 0.24 ng/g and a minimum

mean value of 158.36 ± 0.08 ng/g and a maximum value of 339.83 ± 0.33 ng/g.

Butter/margarine were reported to have 218.52 ± 0.20 ng/g mean iodine content and

the reported minimum and maximum values were 124.02 ± 0.15 ng/g and 313.94 ±

0.39 ng/g, respectively. The mean iodine contents in commonly consumed dairy

products are also presented in the form of a bar diagram as shown in Figure 19, which

1043

.24

499.

98

536.

92 586.

66

730.

10

0.00

200.00

400.00

600.00

800.00

1000.00

1200.00

Fresh fish Clam Canned tuna Canned sardine Chicken egg

Iodi

ne c

onte

nt (n

g/g)

85

makes it easy to identify foods with high and low iodine contents. The processed

powdered milk shows the highest mean iodine content while butter/margarine shows

the lowest iodine content from the in this group of dairy products analysed.

Table 24. Mean iodine contents in commonly consumed dairy products analysed on

a fresh weight basis.

Fresh samples


(n)




Cheese 4 377.57 ± 0.27 191.64 ± 0.27 490.79 ± 0.29

Fresh liquid milk 4 237.70 ± 0.24 158.36 ± 0.08 339.83 ± 0.33Processed powdered milk 4 580.04 ± 0.45 397.02 ± 0.29 795.49 ± 1.07

Butter/margarine 4 218.52 ± 0.20 124.02 ± 0.15 313.94 ± 0.39


dairy products analysed on a fresh weight basis.

377.

57

237.

70

580.

04

218.

52

0.00

100.00

200.00

300.00

400.00

500.00

600.00

700.00

Cheese Fresh liquid milk Processedpowdered milk

Butter/Margarine

Iodi

ne c

onte

nt (n

g/g)



86

Three types of commonly consumed leafy vegetables (lettuce, English cabbage and

Chinese cabbage) have also been investigated for their iodine contents. The mean

iodine contents of commonly consumed leafy vegetables are shown in Table 26. The

highest mean iodine content in this group of leafy vegetables was found in lettuce with

a value of 114.81 ± 0.08 ng/g. The highest reported mean value was 178.17 ± 0.14

ng/g and the minimum value of 40.15 ± 0.03 ng/g. English cabbage showed the mean

iodine content of 108.40 ± 0.06 ng/g with a minimum and maximum of 56.92 ± 0.03

ng/g and 145.81 ± 0.07 ng/g respectively. Chinese cabbage had the lowest iodine

content in this group of leafy vegetables. The reported mean value was 104.01 ± 0.06

ng/g with the minimum mean of 44.64 ± 0.04 ng/g and the maximum mean of 132.87

± 0.12 ng/g. The mean iodine contents in commonly consumed leafy vegetables are

also presented in form of bar diagram (Figure 20) which depicts that lettuce with the

highest mean iodine content while Chinese cabbage showed the lowest mean iodine

content.

Table 25. Mean iodine contents in commonly consumed leafy vegetables analysed




(n)




Lettuce(Lactuca sativa) 4 114.81 ± 0.08 40.15 ± 0.03 178.17 ± 0.14

English cabbage (Brassica oleracea) 4 108.40 ± 0.06 56.92 ± 0.03 145.81 ± 0.07

Chinese cabbage(Brassica chinensis) 4 104.01 ± 0.06 44.64 ± 0.04 132.87 ± 0.12



87


leafy vegetables analysed on a fresh weight basis.

Table 27 shows the mean iodine contents in commonly consumed fruits and

vegetables. From this table, pumpkin had the mean iodine content of 101.24 ± 0.08

ng/g with the maximum mean of 183.58 ± 0.20 ng/g and the minimum mean iodine

content of 48.87 ± 0.03 ng/g. Following this was long bean which had 97.61 ± 0.10

ng/g mean iodine content. The reported minimum value was 37.64 ± 0.04 ng/g with a

maximum mean value of 183.98 ± 0.19 ng/g. Banana had 76.18 ± 0.10 ng/g mean

iodine content with 21.40 ± 0.04 ng/g being the minimum mean and 183.06 ± 0.23

ng/g was the maximum value. Tomato showed the mean iodine content of 40.32 ± 0.04

ng/g with the minimum of 24.17 ± 0.02 ng/g and maximum of 53.62 ± 0.04 ng/g mean

iodine content.

The mean iodine contents in commonly consumed fruits and vegetables are also

presented in the form of a bar diagram as shown in Figure 21. The bar diagram shows

that the studied fruits and vegetables follow a decreasing trend of iodine contents as:

tomato < banana < long bean < pumpkin.

114.

81

108.

40

104.

01

98.00

100.00

102.00

104.00

106.00

108.00

110.00

112.00

114.00

116.00

Lettuce English cabbage Chinese cabbage

Iodi

ne c

onte

nt (n

g/g)

88

Table 26. Mean iodine contents in commonly consumed fruits and vegetables




(n)




Tomato(Solanum lycopersicum)

4 40.32 ± 0.04 24.17 ± 0.02 53.62 ± 0.04

Banana(Musa) 4 76.18 ± 0.10 21.40 ± 0.04 183.06 ± 0.23

Long bean(Vigna unguiculata ssp. Sesquipedalis)

4 97.61 ± 0.10 37.64 ± 0.04 183.98 ± 0.19

Pumpkin (Cucurbita moschata)

4 101.24 ± 0.08 48.87 ± 0.03 183.58 ± 0.20


fruits and vegetable analysed on a fresh weight basis.

40.3

2

76.1

8

97.6

1

101.

24

0.00

20.00

40.00

60.00

80.00

100.00

120.00

Tomato Banana Long bean Pumpkin

Iodi

ne c

onte

nt (n

g/g)



89

Two types of commonly consumed seaweeds (lumiwawa and sea grapes) have also

been studied for their iodine contents. As shown in Table 28, lumiwawa showed the

highest value of the mean iodine content as 6373.30 ± 0.39 ng/g and is commonly

consumed seaweeds. This brown seaweed had the maximum mean of 11000.00 ± 0.32

ng/g and the minimum mean of 2438.68 ± 0.10 ng/g. This was followed by sea grapes

with a mean of 1162.81 ± 0.61 ng/g. The maximum mean determined was 1525.83 ±

0.83 ng/g with a minimum mean iodine of 851.92 ± 0.34 ng/g. These sea grapes are

referred to as the green seaweeds sold commonly in the markets. The mean iodine

contents in commonly consumed seaweeds are also presented in self-explanatory bar

diagram shown in Figure 22.

Table 27. Mean iodine contents in commonly consumed seaweeds analysed on a

fresh weight basis.



(n)




Sea grapes(Caulerpa lentillifera) 4 1162.81 ± 0.61 851.92 ± 0.34 1525.83 ± 0.83

Lumiwawa (Gracilaria maramae) 4 6373.30 ± 0.39 2438.68 ± 0.10 11000.00 ± 0.32


seaweeds analysed on a fresh weight basis.

1162

.81

6373

.30

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

Sea grapes (green seaweed) Lumiwawa (brown seaweed)

Iodi

ne c

onte

nt (n

g/g)



90

Table 28. Mean iodine contents in commonly consumed food samples analysed on a

fresh weight basis.



(n)




Rice(Oryza sativa) 4 99.92 ± 0.11 53.89 ± 0.06 195.47 ± 0.19

Potato (Solanum tuberosum) 4 255.87 ± 0.27 235.25 ± 0.21 265.60 ± 0.18

Cassava(Manihot esculenta) 4 262.76 ± 0.19 219.64 ± 0.12 345.91 ± 0.22

Dalo/Taro (Colocasia esculenta) 4 311.93 ± 0.28 227.43 ± 0.20 379.85 ± 0.32

Fresh marine fish 4 1043.24 ± 0.75 966.55 ± 0.42 1088.17 ± 0.89 Clam(Margaritifera) 4 499.98 ± 0.48 449.72 ± 0.52 587.44 ± 0.53

Canned tuna 4 536.92 ± 0.49 313.55 ± 0.34 660.77 ± 0.68

Canned sardine 4 586.66 ± 0.40 351.45 ± 0.42 956.15 ± 0.42

Chicken egg 4 730.10 ± 0.47 591.79 ± 0.34 1112.08 ± 0.03

Cheese 4 377.57 ± 0.27 191.64 ± 0.27 490.79 ± 0.29

Fresh liquid milk 4 237.70 ± 0.24 158.36 ± 0.08 339.83 ± 0.33

Processed powdered milk 4 580.04 ± 0.45 397.02 ± 0.29 795.49 ± 1.07

Butter/Margarine 4 218.52 ± 0.20 124.02 ± 0.15 313.94 ± 0.39

Vegetables

Lettuce(Lactuca sativa) 4 114.81 ± 0.08 40.15 ± 0.03 178.17 ± 0.14

English cabbage (Brassica oleracea) 4 108.40 ± 0.06 56.92 ± 0.03 145.81 ± 0.07

Chinese cabbage(Brassica chinensis) 4 104.01 ± 0.06 44.64 ± 0.04 132.87 ± 0.12

91

Fruits

Tomato(Solanum lycopersicum)

4 40.32 ± 0.04 24.17 ± 0.02 53.62 ± 0.04

Banana(Musa) 4 76.18 ± 0.10 21.40 ± 0.04 183.06 ± 0.23

Long bean (Vigna unguiculata ssp. Sesquipedalis)

4 97.61 ± 0.10 37.64 ± 0.04 183.98 ± 0.19

Pumpkin (Cucurbita moschata) 4 101.24 ± 0.08 48.87 ± 0.03 183.58 ± 0.20

Seaweeds

Sea grapes (Caulerpa lentillifera) 4 1162.81 ± 0.61 851.92 ± 0.34 1525.83 ± 0.83

Lumiwawa (Gracilaria maramae) 4 6373.30 ± 0.39 2438.68 ± 0.10 11000.00 ± 0.32

The summary of the determined iodine contents in 22 commonly consumed food

samples shown in Table 29 clearly shows that seaweeds lumiwawa and sea grapes had

the highest levels of average iodine content being 6373.30 ± 0.39 ng/g and 1162.81 ±

0.61 ng/g, respectively followed by fresh seawater fish with the mean iodine content

of 1043.24 ± 0.75 ng/g. Egg had 730.10 ± 0.47 ng/g, canned sardine 586.66 ± 0.40

ng/g, processed powdered milk 580.04 ± 0.45 ng/g, canned tuna 536.92 ± 0.49 ng/g,

clam 499.98 ± 0.48 ng/g, cheese 377.57 ± 0.27 ng/g, dalo/taro 311.93 ± 0.28 ng/g,

cassava 262.76 ± 0.19 ng/g, potato 255.87 ± 0.27 ng/g, fresh liquid milk 237.70 ± 0.24

ng/g, butter/margarine 218.52 ± 0.20 ng/g, lettuce 114.81 ± 0.08 ng/g, English cabbage

108.40 ± 0.06 ng/g, Chinese cabbage 104.01 ± 0.06 ng/g, pumpkin 101.24 ± 0.08 ng/g,

rice 99.92 ± 0.11 ng/g, long bean 97.61 ± 0.10 ng/g, banana 76.18 ± 0.10 ng/g and

tomato 40.32 ± 0.04 ng/g. Comparison of iodine contents for all the 22 food samples

is shown in Figure 23.

Lumiwawa is brown seaweed which has the highest iodine content. Other studies have

confirmed that brown seaweeds generally have the highest iodine content (Nitschke

and Stengel, 2015; Moreda-Pin˜eiro et al., 2007; Diego et al., 2005; Romarís–Hortas

et al., 2011; Shah et al., 2005). The lowest iodine values were observed in fruits and



92

vegetables. Similar results have been previously reported for food samples of different

origins (Longvah et al., 2013; Centre for Food Safety - Hong Kong, 2011; Wenlock et

al., 1982; Pennington et al., 1995; Haldimann et al., 2005)

93

Figu

re 2

3.G

raph

ical

repr

esen

tatio

n of

det

erm

ined

aver

age

iodi

ne c

onte

nts(

ng/g

)of t

he a

naly

sed

food

sam

ples

.

99.92

255.87

262.76

311.93

1043.24

499.98

536.92

586.66

730.10

377.57

237.70

580.04

218.52

114.81

108.40

104.01

40.32

76.18

97.61

101.24

1162.81

6373.30

0.00

1000

.00

2000

.00

3000

.00

4000

.00

5000

.00

6000

.00

7000

.00


94

5.1.1. Cluster analysis

Figure 24. Dendrogram of cluster analysis (Ward’s method) of determined average

iodine contents (ng/g) of the analysed food samples.

95

The Hierarchical method for the cluster analysis of the food samples was used

(Kelepertzis, 2014; Davis et al., 2009; Argyraki et al., 2014). To create dendrograms,

Ward’s method and Euclidian distance was used. The height of the cluster clades

shows the distance between data points. This height can also be used to determine the

similarity or the difference between them (Singh et al., 2016). Figure 24 shows two

clusters. A cluster for single food is seen for Lumiwawa (brown seaweed) which shows

the highest level of average iodine content of 6373.30 ± 0.39 ng/g among the 22

commonly consumed food samples analysed. The next main cluster is for all other

food groups from rice to chicken egg. The second main cluster is further grouped in

two groups: one is from rice to cheese and while the other cluster is for food groups

from fresh fish to chicken eggs. Food samples with lowest average iodine contents are

in one cluster i.e. rice to banana while food samples with higher average iodine

contents fall in other clusters.

The food samples in the lowest clade with similar average iodine contents consisted of

rice having 99.92 ± 0.11 ng/g, pumpkin 101.24 ± 0.08 ng/g, long bean 97.61 ± 0.10

ng/g, English cabbage 108.40 ± 0.06 ng/g, Chinese cabbage 104.01 ± 0.06 ng/g, lettuce

114.81 ± 0.08 ng/g, tomato 40.32 ± 0.04 ng/g and banana 76.18 ± 0.10 ng/g. The other

clade consists of potato 255.87 ± 0.27 ng/g, cassava 262.76 ± 0.19 ng/g, fresh liquid

milk 237.70 ± 0.24 ng/g, butter/margarine 218.52 ± 0.20 ng/g, dalo/taro 311.93 ± 0.28

ng/g and cheese 377.57 ± 0.27 ng/g which had similar average iodine contents. Fresh

fish and sea grapes (green seaweed) made one clade because their average iodine

contents ranged between 1043.24 ± 0.75 ng/g and 1162.81 ± 0.61 ng/g. Canned

sardine, processed powdered milk, clam, canned tuna and chicken egg have made one

clade with the average iodine content 586.66 ± 0.40 ng/g, 580.04 ± 0.45 ng/g, 499.98

± 0.48 ng/g, 536.92 ± 0.49 ng/g, 730.10 ± 0.47 ng/g, respectively (cf. Figure 24).

5.2. Comparison of iodine content with previous published data

There are very few studies on iodine content in different foods in ng/g on a fresh weight

basis (Haldimann et al., 2005; Wenlock et al., 1982; Pennington et al., 1995; Catalan

Food Safety Agency, 2016; Centre for Food Safety - Hong Kong, 2011). The results

obtained in the present study were compared to other published data as reported in

Table 30.

96

The comparison of results from this present study of iodine contents in foods analysed

on a fresh weight basis shows some similarities to the results published by other

researchers. There were similarities in the results for fish (marine) with the present

study which had the mean iodine content of 1043.24 ± 0.75 ng/g. Other studies of

marine fish on a fresh weight basis reported by Pennington et al. (1995) had 1160 ±

880 ng/g fresh weight while one reported by Wenlock et al. (1982) showed a mean of

750 ng/g with the minimum of 320 ng/g and the maximum of 1440 ng/g iodine

concentration.

The mean value for the iodine content in chicken egg was 730.10 ± 0.47 ng/g. This

data was compared to that of Wenlock et al. (1982) which had an average iodine

content of 525 ng/g. The next comparison was done for the milk samples. Liquid milk

samples showed a mean iodine content of 237.70 ± 0.24 ng/g. Results from previous

published data showed almost similar results as 230 ng/g with the minimum 50 ng/g

and the maximum 550 ng/g (Wenlock et al., 1982) and 230.2 - 702.7 ng/mL as reported

by Travnicek et al. (2006) for raw milk samples.

Processed powdered milk samples in this study had 580.04 ± 0.45 ng/g iodine. This

was compared to the value reported by Judprasong et al. (2016) which was 544 ng/g

with the minimum of 413 ng/g and the maximum of 675 ng/g. Almost similar results

were also obtained by the Centre for Food Safety - Hong Kong (2011) with an average

of 430 ng/g showing 300 ng/g minimum and 580 ng/g maximum iodine contents. The

values for cheese analysed in the present study having an average iodine content of

377.57 ± 0.27 ng/g was again compared to that of the Centre for Food Safety - Hong

Kong (2011) which gave a mean concentration of 420 ng/g, while the Catalan Food

Safety Agency (2016) reported an average iodine content of 577.6 ng/g.

The average mean obtained for rice of 99.92 ng/g was compared to the average

achieved by Longvah and Deosthale (1998) of 104 ng/g (minimum 88 ng/g and

maximum 129 ng/g). In comparison of vegetables, the iodine content of long bean of

97.61 ng/g (37.64 ng/g minimum – 183.98 ng/g maximum) was compared to that

reported by Judprasong et al. (2016) who reported an average value of 109.33 ng/g

with a range of 63 - 171 ng/g. These compared data are presented in Table 30.

97

These comparisons of iodine contents in the food samples from the present study

clearly show that there were variability of iodine contents in different foods from

different regions/ countries. Iodine contents reported from commonly consumed foods

and water from the Northeast region in India showed that iodine contents in foods and

water were lower than other non-endemic areas indicating that environmental iodine

deficiency was evident in the Northeast region (Longvah and Deosthale, 1998). This

also indicates that iodine contents may vary in different regions or geographic

locations. Haldimann et al. (2005) reported that iodine content of plant foods varied

from species to species. It was noted that the nutrient components of plants such as

fruits and vegetables had lower iodine contents.

Iodine contents of food vary with different geographical locations due to a large

variation of iodine contents for different environmental areas . For

plants, iodine content in them is dependent on the iodine content of the soil where they

grow. This factor determines the iodine contents for food chains. Seafoods were found

to have high iodine concentration in this research. This is due to the fact that a large

amount of iodine in the upper crust of the earth is leached and carried out to the sea

9). The ability of the individual species in the sea to accumulate iodine

determines the iodine levels in them. Thus, for this reason, a significant variation is

observed in foods analysed from the sea ; Koutras et al., 1985). A

correlation of the geographic location and the atmospheric transport of iodine from the

sea and the deposition in soil have been highlighted by Blazewicz (2012). In general,

area closer to the coast having high rainfall exhibited higher iodine values in organic

matter and, thus, suggests that the atmospheric transport could be the reason for higher

iodine levels.

98

Tab

le 2

9. C

ompa

rison

of i

odin

e co

nten

ts in

sele

cted

food

sam

ples

from

the

pres

ent r

esea

rch

with

pre

viou

s pub

lishe

d da

ta a

naly

sed

on

a fr

esh

wei

ght b

asis

.

Food

sam

ple

(type

)

Ave

rage

iodi

ne c

onte

nt in

fres

h sa

mpl

es w

ith ra

nge

(min

imum

–m

axim

um) o

r ± S

D (n

g/g)

Pres

ent s

tudy

Mea

n(M

in –

Max

)

Cen

tre fo

r Foo

d Sa

fety

- H

ong

Kon

g, 2

011

Wen

lock

et

al.,

1982

Cat

alan

Foo

d Sa

fety

Age

ncy,

20

16

Penn

ingt

on e

t al.,

19

95H

aldi

man

n et

al

., 20

05

Fres

h fis

h (m

arin

e)10

43.2

4(9

66.5

5 –

1088

.17)

17

0(5

0 - 6

00)

750

(320

- 14

40)

259.

7(7

0 - 6

24)

1160

± 8

80

486

(89

- 159

3)

Cla

m49

9.98

(449

.72

– 58

7.44

) 11

00(5

90 -

2100

) -

667

- -

Can

ned

tuna

53

6.92

(313

.55

– 66

0.77

) 98 (7

4 –

120)

-

153

- -

Can

ned

sard

ine

586.

66(3

51.4

5 –

956.

15)

190

(100

- 34

0)

- 23

0-

-

Chi

cken

egg

73

0.10

(591

.79

– 11

12.0

8)

290

(82

- 430

) 52

546

148

0 ±

390

324

(247

– 4

28)

Che

ese

377.

57(1

91.6

4 –

490.

79)

420

(160

- 14

00)

577.

6-

-

Fres

h liq

uid

milk

237.

70(1

58.3

6 –

339.

83)

91 (56

- 130

) 23

0(5

0 - 5

50)

146

(45

- 204

) 20

0 ±

80

124

(59

– 19

9)

Proc

esse

d po

wde

red

milk

58

0.04

(397

.02

– 79

5.49

) 43

0(3

00 -

580)

-

- -

-

But

ter/

Mar

garin

e21

8.52

(124

.02

– 31

3.94

) -

- 2.

8 - 8

.8

- -

99

a- R

epre

sent

atio

n of

ave

rage

iodi

ne c

onte

nts i

n al

l veg

etab

les.

b - R

epre

sent

atio

n of

ave

rage

iodi

ne c

onte

nts i

n al

l fru

its.

Min

- M

inim

umM

ax -

Max

imum

Veg

etab

les

(< 2

0 –

280)

a <

10a

5a (1 –

22)

Lettu

ce11

4.81

(40.

15 –

178

.17)

1

- 21

- -

Engl

ish

cabb

age

108.

40(5

6.92

– 1

45.8

1)

- -

- -

-

Chi

nese

ca

bbag

e 10

4.01

(44.

64 –

132

.87)

25 (2

1 –

28)

- -

- -

Frui

ts(<

20 –

80)

b <

30b

3b (0.3

– 1

3)

Tom

ato

40.3

2(2

4.17

– 5

3.62

) -

- 5.

7-

-

Ban

ana

76.1

8(2

1.40

– 1

83.0

6)

- -

<4-

-

Long

bea

n 97

.61

(37.

64 –

183

.98)

6 (4

– 7

) -

6.5

- -

Pum

pkin

10

1.24

(48.

87 –

183

.58)

-

- -

- -

Sea

grap

es

1162

.81

(851

.92

– 15

25.8

3)

(840

– 2

9000

00)

- -

- -

Lum

iwaw

a 63

73.3

0(2

438.

68 –

110

00.0

0)

- -

- -

-

100

A very good correlation has been seen in the iodine contents determined in the present

study and with the previously published data. Only the fruits and vegetable samples

showed higher iodine values than reported in the literature. Studies have indicated that

green leafy vegetables have higher iodine contents when compared to other vegetable

types (Fordyce, 2003; Haldimann et al., 2005). In the present study, fruits exhibited

high iodine contents compared to the data presented in earlier studies. This could be

due to the difference in geographic location and the atmospheric transport of iodine

from the ocean and deposition in soil. Soil samples close to the coast, where there was

high rainfall and areas with high organic matter exhibited high iodine values

(Blazewicz, 2012).

In another analysis on the iodine content in foods and diets, iodine content was highest

in marine fish (1456 μg/kg) followed by fresh water fish (106 μg/kg), and then leafy

vegetables (89 μg/kg), dairy (84 μg/kg), other vegetables (80 μg/kg), meat (68 μg/kg),

cereals (56 μg/kg), fresh fruit (31 μg/kg) and bread (17 μg/kg) (Fordyce, 2003). The

results show that grain crops are poor sources of iodine than leafy vegetables. There

are also some evidences that indicate that leafy vegetables have higher iodine content

than some other vegetables (Haldimann et al., 2005; Leufroy et al., 2015). The iodine

content of foods varies with different geographic locations therefore, iodine content

from one country cannot be universally used to estimate the iodine intake for another

population (Longvah et al., 2013). Furthermore, a research in Central Europe indicated

that iodine concentration of drinking water decreases with increasing distance from

the ocean indicating that iodine contents in water and food are likely to be higher in

locations that are closer to the ocean (Anke et al., 1995).

With this justification, the iodine concentrations of fruits and vegetables obtained in

the present research are acceptable. Fiji being a small island nation, surrounded by the

ocean can have higher soil iodine concentrations compared to other countries

2009; Koutras et al., 1985). This was evident in the higher iodine contents in the

vegetable and fruits analysed. Further research needs to be carried out to fully justify

this assumption.

101

CHAPTER 6

CONCLUSION AND RECOMMENDATIONS

6.1. Conclusion

Iodine is a mineral that is important for human health. As seen in the literature, iodine

is needed in the body to make hormones in the thyroid. These hormones are needed by

the human body for the proper functioning of the body which includes growth,

metabolism and also for the development of a baby’s brain during pregnancy. A low

intake of iodine for a long period of time will result in the thyroid to work harder to

maintain the right amount of thyroid hormone in the blood. This will directly lead to

an increase in the thyroid size in order to trap more iodine from the body (The British

Dietetic Association , 2016). This swelling or increase in the thyroid size is called

goiter and could be easily visible in the neck (The British Dietetic Association , 2016).

Iodine deficiency in the human body is also linked to other problems such as endemic

cretinism, infant mortality, infertility, miscarriage, mental retardation, neuromuscular

defects, and dwarfism. All these are known as IDDs. Therefore, a knowledge of the

daily iodine intake as recommended by WHO is important. Food being the major

iodine source for the human body needed to be critically analysed for iodine contents

so people can understand, know and plan their daily iodine intake in Fiji and the region

from different foods.

In this research, the objectives of the proposed project were met by firstly validating

the spectrophotometric kinetic method for iodine determination (Mahesh et al., 1992).

It has now been confirmed that very trace levels of iodine (ng/g) can be analysed

successfully using modified ashing procedure and the spectrophotometric kinetic

method. A number of studies have been conducted to identify iodine contents in foods

using the kinetic method for concentrations in μg and mg levels, but there was limited

data that studied the iodine levels in very trace levels at ng/g. This was successfully

achieved in this research.

Secondly, as the main objective, some commonly consumed Fiji foods were

successfully analysed for their iodine contents using the spectrophotometric kinetic

102

method. It was observed in this study that the following commonly consumed foods in

Fiji had the following iodine contents as reported below.

To sum up, seaweeds lumiwawa and sea grapes showed the highest levels of average

iodine content being 6373.30 ± 0.39 ng/g and 1162.81 ± 0.61 ng/g, respectively

followed by fresh seawater fish with an average iodine content of 1043.24 ± 0.75 ng/g.

Chicken egg had 730.10 ± 0.47 ng/g, canned sardine 586.66 ± 0.40 ng/g, processed

powdered milk 580.04 ± 0.45 ng/g, canned tuna 536.92 ± 0.49 ng/g, clam 499.98 ±

0.48 ng/g, cheese 377.57 ± 0.27 ng/g, dalo/taro 311.93 ± 0.28 ng/g, cassava 262.76 ±

0.19 ng/g, potato 255.87 ± 0.27 ng/g, fresh liquid milk 237.70 ± 0.24 ng/g,

butter/margarine 218.52 ± 0.20 ng/g, lettuce 114.81 ± 0.08 ng/g, English cabbage

108.40 ± 0.06 ng/g, Chinese cabbage 104.01 ± 0.06 ng/g, pumpkin 101.24 ± 0.08 ng/g,

rice 99.92 ± 0.11 ng/g, long bean 97.61 ± 0.10 ng/g, banana 76.18 ± 0.10 ng/g and

tomato 40.32 ± 0.04 ng/g.

Thirdly, based on the comparison of the iodine contents of fresh foods versus factory

processed foods, it can be concluded that fresh foods showed higher iodine

concentrations than those of the factory processed foods. Fresh fish had the mean

iodine concentration of 1043.24 ± 0.75 ng/g for the 4 different species of fish ranging

from 966.55 ± 0.42 to 1088.17 ± 0.89 ng/g. It was compared with the iodine

concentration in different brands of canned tuna ranging from 313.55 ± 0.34 to 660.77

± 0.68 ng/g. Furthermore, 4 different canned sardine samples were also analysed and

the iodine concentration ranged from 351.45 ± 0.42 to 956.15 ± 0.42 ng/g with an

average of 586.66 ± 0.40 ng/g. Fresh liquid milk samples were also analysed to verify

the assumption that iodine content is low in factory processed foods than fresh foods.

Three factory processed liquid milk samples were analysed for iodine content which

ranged from 158.36 ± 0.08 to 242.09 ± 0.29 ng/g. Fresh cow milk (unprocessed) had

an iodine concentration of 339.83 ± 0.33 ng/g. Thus, there is clear evidence that fresh

foods have higher iodine content than processed foods unless the processed food items

are fortified.

It was also observed that for the seaweeds studied, lumiwawa (brown seaweed) had an

average iodine concentration of 6373.30 ± 0.39 ng/g which was higher than the sea

grapes (green seaweed) which had an average iodine content of 1162.81 ± 0.61 ng/g.

103

It can be concluded that brown seaweed has a higher iodine content than the green

seaweed.

In a report by WHO, based on the country data on the urinary iodine and national

estimate of iodine nutrition has classed Fiji in the category of insufficient iodine intake

(WHO, 2004). The classification of iodine nutrition for Fiji was moderate iodine

deficiency when the study was done at a district level with a 479 sample size (WHO,

2004). Food being the major contributor of iodine in the body, makes the knowledge

of iodine contents in foods very important for Fiji. The data presented from this

research will make people aware of the daily dietary iodine intakes or will be able to

inform people about foods rich in iodine.

The spectrophotometric kinetic method for iodine determination at trace levels (ng/g)

in food samples has been validated through this study. The data presented give clear

indication of the iodine content of some commonly consumed foods in Fiji and forms

a basis of a basic database on iodine levels from foods in the country and in the region.

The spectrophotometric kinetic method used in this research has definite advantage of

being very sensitive, versatile and can be adapted easily with minimum equipment and

chemicals as an inexpensive method for the determination of iodine in foods.

6.2. Recommendations

Iodine contents in commonly consumed foods analysed in this study form a foundation

of determined iodine contents in food available in Fiji. Iodine is an essential nutrient

in the human body. Despite this fact, there had not been any studies conducted in Fiji

for community awareness. The data presented in this study on the iodine contents in

foods form a basis of an upgrade to the Pacific Islands Food Composition Tables and

thus would now serve as a basis for educating the general public. Recommendations

for further research on iodine studies in foods are also explained below.

6.2.1. Recommendations to the general public

- More awareness be made by the Health authorities on the role of iodine to the

human body.

104

- The data presented in this study be made available to the general public. The

general public can also use the data presented in this study to regulate their

daily iodine intake.

6.2.2. Recommendations for future study

The data collected in this study is hoped to be significant for public health purposes,

however more studies need to be carried out. The following are recommendations for

future iodine related research:

- More foods need to be analysed to update the current database on iodine

contents in foods.

- A total diet study of iodine in Fiji foods needs to be carried out using validated

kinetic spectrophotometric methods similar to what is used in this research.

- The Pacific Island Food Composition Table needs to be updated. If possible

iodine contents in foods need to be incorporated in the Tables.

- The effect of cooking and the losses of iodine during food preparation need to

be studied to determine the correct iodine intake by people in Fiji.

105

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APPENDICES

Appendix 1: Average absorbance at different iodine concentrations for t = 0 min.


Time 0 min Run (n = 7)

Average absorbance (A)

Standard deviation (SD)

RSD (%)

Blank 1.002 1.00 0.00 0.31.0011.0011.0070.9971.0031.006

2.5 0.998 1.00 0.00 0.30.9990.9991.0001.0021.0050.995

5.0 1.003 1.00 0.00 0.20.9990.9980.9960.9980.9990.998

10.0 1.003 1.00 0.00 0.30.9960.9940.9971.0000.9991.001

15.0 0.997 1.00 0.00 0.30.9960.9901.0001.0000.9960.997

118

20.0 1.003 1.00 0.00 0.40.9960.9971.0000.9970.9991.007

25.0 0.999 1.00 0.00 0.41.0050.9961.0041.0000.9991.007

Appendix 2: Average absorbance at different iodine concentrations for t = 0.5 min.


Time 0.5 min Run (n = 7)



RSD (%)

Blank 0.905 0.903 0.00 0.20.9040.9000.9050.9010.9030.905

2.5 0.892 0.897 0.00 0.50.8990.8980.8970.8960.904

0.891

5.0 0.887 0.886 0.01 0.70.8940.8920.8760.8830.8840.883

119

10.0 0.875 0.870 0.00 0.40.8720.8710.8720.8680.8650.870

15.0 0.851 0.856 0.00 0.50.8580.8490.8610.8580.8560.857

20.0 0.855 0.842 0.01 1.00.8310.8370.8450.8430.8350.846

25.0 0.818 0.828 0.01 0.80.8330.8220.8360.8280.8270.834

120

Appendix 3: Average absorbance at different iodine concentrations for t = 1 min.


Time 1 min Run (n = 7)



RSD (%)

Blank 0.815 0.813 0.00 0.50.8100.8100.8170.8060.8180.814

2.5 0.792 0.797 0.01 0.70.7970.8000.7990.7940.8060.791

5.0 0.783 0.781 0.01 1.00.7910.7920.7710.7760.7750.778

10.0 0.753 0.756 0.01 0.70.7620.7600.7560.7590.7470.757

15.0 0.729 0.728 0.00 0.40.7280.7230.7320.7300.7300.726

20.0 0.719 0.704 0.01 1.30.6990.6970.709

121

0.7070.6920.704

25.0 0.667 0.678 0.01 1.40.6880.6630.6860.6800.6760.683

122

Appendix 4: Absorbance for the determination of iodine in standard iodine solutions

at 4, 12, and 18 ng/mL and their recovery.


(ng/mL)in


Iodine recovery(%)

4.0 0.210 3.96 99.064.0 0.209 3.77 94.344.0 0.210 3.96 99.064.0 0.212 4.34 108.49

Average 4.01 ± 0.24 100.24 ± 5.92Standard analytical error (%) 0.60


(ng/mL)in


Iodine recovery (%)

12.0 0.253 12.08 100.6312.0 0.252 11.89 99.0612.0 0.252 11.89 99.0612.0 0.253 12.08 100.63



(ng/mL)n


Iodine recovery (%)

18.0 0.284 17.92 99.5818.0 0.284 17.92 99.5818.0 0.288 18.68 103.7718.0 0.282 17.55 97.48


123

Appendix 5: Absorbance for the determination of iodine in different food samples analysed

along with average iodine contents and coefficient of variation.

1 Rice

Sample No. of

analysis (n)

Sample dilution (mL)

Analysis Iodide content (ng/g)

Coefficient of variation

(%)

Punjas Jasmine rice

1

7

0.2009 2.25

3.832 0.2020 2.453 0.2012 2.304 0.2012 2.30

Average iodine (ng/g) = 65.11 ± 0.09Analytical coefficient of variation (%) = 0.23

Sample No. of

analysis (n)




(%)

FMF Sun Grown rice

1

7

0.1996 2.00

3.102 0.1989 1.873 0.1990 1.894 0.1993 1.94


Sample No. of

analysis (n)




(%)

Punjas Long Grain rice

1

7

0.2054 3.09

3.772 0.2049 3.003 0.2058 3.174 0.2044 2.91


Sample No. of

analysis (n)




(%)

Sunwhite Calrose rice

1

7

0.2253 6.85

2.742 0.2257 6.923 0.2275 7.264 0.2255 6.89


124

2 Potato

Sample No. of

analysis (n)




(%)

Potato (MH)

1

7

0.2381 9.26

1.882 0.2392 9.473 0.2404 9.704 0.2394 9.51


Sample No. of

analysis (n)




(%)

Potato (Shop &

Save)

1

7

0.2401 9.64

3.192 0.2381 9.263 0.2387 9.384 0.2363 8.92


Sample No. of

analysis (n)




(%)

Potato (New World)

1

7

0.2407 9.75

3.962 0.2368 9.023 0.2399 9.604 0.2371 9.08


Sample No. of

analysis (n)




(%)

Potato (Market)

1

7

0.2342 8.53

2.512 0.2332 8.343 0.2346 8.604 0.2321 8.13


125

3 Cassava

Sample No. of

analysis(n)




(%)

Cassava (Vendor 1)

1

7

0.2547 12.40

1.782 0.2539 12.253 0.2560 12.644 0.2533 12.13


Sample No. of

analysis (n)




(%)

Cassava (vendor 2)

1

7

0.2315 8.02

1.502 0.2302 7.773 0.2304 7.814 0.2302 7.77


Sample No. of

analysis (n)




(%)

Cassava (Vendor 3)

1

7

0.2327 8.25

1.842 0.2345 8.583 0.2332 8.344 0.2341 8.51


Sample No. of

analysis (n)




(%)

Cassava (Vendor 4)

1

7

0.2383 9.30

2.922 0.2354 8.753 0.2360 8.874 0.2354 8.75


126

4 Dalo

Sample No. of

analysis (n)




(%)

Dalo (Vendor 1)

1

7

0.2450 10.57

2.742 0.2469 10.923 0.2488 11.284 0.2463 10.81


Sample No. of

analysis (n)




(%)

Dalo (Vendor 2)

1

7

0.2528 12.04

2.332 0.2533 12.133 0.2535 12.174 0.2503 11.57


Sample No. of

analysis (n)




(%)

Dalo (Vendor 3)

1

7

0.2585 13.11

2.382 0.2625 13.873 0.2616 13.704 0.2610 13.58


Sample No. of

analysis (n)




(%)

Dalo (Vendor 4)

1

7

0.2305 7.83

2.482 0.2322 8.153 0.2327 8.254 0.2328 8.26


127

5 Fresh Fish

Sample No. of

analysis (n)




(%)

Parrot fish(Chlorurus sordidus)

1

11

0.3046 21.81

1.902 0.3081 22.473 0.3029 21.494 0.3061 22.09


Sample No. of

analysis (n)




(%)Malabar Grouper

(Soisoi) fish (Epinephelus Malabarcius)

1

11

0.3128 23.36

3.882 0.3106 22.943 0.3219 25.084 0.3158 23.92


Sample No. of

analysis (n)




(%)Russell's

Snapper fish (Kwake) (Lujanus russelli)

1

11

0.3128 23.36

3.042 0.3193 24.583 0.3173 24.214 0.3221 25.11


Sample No. of

analysis (n)




(%)

Pacific yellowtail

emperor fish (Sabutu)

(Lethrinus atkinsoni)

1

11

0.3190 24.53

3.592 0.3270 26.04

3 0.3174 24.23

4 0.3169 24.13Average iodine (ng/g) = 1088.17 ± 0.89

Analytical coefficient of variation (%) = 1.47

128

6 Clam

Sample No. of

analysis (n)




(%)

Clam Vendor 1

1

7

0.2751 16.25

3.212 0.2771 16.623 0.2737 15.984 0.2706 15.40


Sample No. of

analysis (n)




(%)

Clam Vendor 2

1

7

0.2819 17.53

1.452 0.2807 17.303 0.2807 17.304 0.2787 16.92


Sample No. of

analysis (n)




(%)

Clam Vendor 3

1

7

0.2963 20.25

2.522 0.3029 21.493 0.3008 21.094 0.3010 21.13


Sample No. of

analysis (n)




(%)

Clam Vendor 4

1

7

0.2787 16.92

3.702 0.2756 16.343 0.2809 17.344 0.2835 17.83


129

7 Canned Tuna

Sample No. of

analysis (n)




(%)

Sunbell Tuna

1

7

0.3119 23.19

2.892 0.3112 23.063 0.3192 24.574 0.3140 23.58


Sample No. of

analysis (n)




(%)

Burnswick Tuna

1

7

0.2505 11.60

2.992 0.2491 11.343 0.2467 10.894 0.2471 10.96


Sample No. of

analysis(n)




(%)Sunbell

Ovalau Blue (Light Tuna

Flakes)

1

7

0.2826 17.66

3.682 0.2912 19.283 0.2857 18.254 0.2857 18.25


Sample No. of

analysis(n)




(%)

Skipper Tuna

1

7

0.3156 23.89

1.072 0.3126 23.323 0.3140 23.584 0.3130 23.40


130

8 Canned Sardine

Sample No. of

analysis (n)




(%)

Burnswick sardine in

vegetable oil

1

7

0.2929 19.60

2.832 0.2861 18.323 0.2907 19.194 0.2907 19.19


Sample No. of

analysis (n)




(%)


spring water

1

12

0.2977 20.51

2.102 0.2929 19.603 0.2931 19.644 0.2946 19.92


Sample No. of

analysis (n)




(%)


tomato sauce

1

7

0.2840 17.92

1.152 0.2836 17.853 0.2846 18.044 0.2861 18.32


Sample No. of

analysis(n)




(%)


lemon sauce

1

7

0.2584 13.09

3.372 0.2539 12.253 0.2536 12.194 0.2562 12.68


131

9 Chicken Egg

Sample No. of

analysis(n)




(%)

Ram Sami & Sons egg

1

7

0.3031 21.53

3.782 0.3114 23.093 0.3041 21.724 0.3015 21.23


Sample No. of

analysis (n)




(%)

Egg Market Vendor 2

1

8.5

0.2829 17.72

1.932 0.2789 16.963 0.2809 17.344 0.2823 17.60


Sample No. of

analysis(n)




(%)

Egg Market Vendor 3

1

7

0.3077 22.40

3.212 0.3013 21.193 0.3046 21.814 0.2994 20.83


Sample No. of

analysis (n)




(%)

Egg Market Vendor 4

1

14

0.2943 19.87

0.162 0.2940 19.813 0.2943 19.874 0.2944 19.89


132

10 Cheese

Sample No. of

analysis (n)




(%)Rewa

Tasty (Fiji Dairy

cheese)

1

7

0.2257 6.92

3.912 0.2268 7.133 0.2234 6.494 0.2252 6.83


Sample No. of

analysis(n)




(%)

Lemnos cheese

1

7

0.2825 17.64

1.672 0.2825 17.643 0.2830 17.744 0.2796 17.09


Sample No. of

analysis(n)




(%)

Chesdale cheese

1

7

0.2663 14.58

1.822 0.2660 14.533 0.2642 14.194 0.2676 14.83


Sample No. of

analysis (n)




(%)

Devondale cheese

1

7

0.2694 15.17

1.602 0.2701 15.303 0.2676 14.834 0.2676 14.83


133

11 Fresh Liquid Milk

Sample No. of

analysis (n)




(%)

Rewa Life full cream

milk

1

7

0.2303 7.79

3.272 0.2272 7.213 0.2293 7.604 0.2286 7.47


Sample No. of

analysis(n)




(%)

Anchor regular milk

1

7

0.2194 5.74

1.482 0.2185 5.573 0.2193 5.724 0.2187 5.60


Sample No. of

analysis (n)




(%)

Meadow fresh milk

1

7

0.2350 8.68

3.312 0.2336 8.423 0.2338 8.454 0.2369 9.04


Sample No. of

analysis (n)




(%)

Fresh cow milk un-processed

1

7

0.2550 12.45

2.742 0.2509 11.683 0.2533 12.134 0.2541 12.28


134

12 Processed Milk

Sample No. of

analysis (n)




(%)Rewa full

cream powdered

milk

1

7

0.2767 16.55

2.422 0.2762 16.453 0.2805 17.264 0.2798 17.13


Sample No. of

analysis (n)




(%)Redcow full

cream powdered

milk

1

7

0.2632 14.00

2.032 0.2628 13.923 0.2662 14.574 0.2644 14.23


Sample No. of

analysis (n)




(%)

Rewa Skim milk powder

1

7

0.3131 23.42

0.142 0.3130 23.403 0.3134 23.474 0.3131 23.42


Sample No. of

analysis (n)




(%)

Dairy Fresh full cream

milk powder

1

7

0.3368 27.89

3.762 0.3434 29.133 0.3451 29.454 0.3330 27.17


135

13 Butter

Sample No. of

analysis (n)




(%)

Rewa butter

1

7

0.2135 4.62

3.322 0.2116 4.263 0.2124 4.424 0.2124 4.42


Sample No. of

analysis (n)




(%)

Anchor butter

1

7

0.2454 10.64

3.492 0.2491 11.343 0.2501 11.534 0.2491 11.34


Sample No. of

analysis (n)




(%)

Flora margarine

1

7

0.2386 9.36

1.152 0.2397 9.573 0.2388 9.404 0.2384 9.32


Sample No. of

analysis (n)




(%)

Meadowlea margarine

1

7

0.2221 6.25

2.482 0.2218 6.193 0.2223 6.284 0.2205 5.94


136

Vegetables14 Lettuce

Sample No. of

analysis (n)




(%)

Lettuce - Vendor 1

1

7

0.2225 6.32

2.162 0.2218 6.193 0.2234 6.494 0.2232 6.45


Sample No. of

analysis (n)




(%)

Lettuce - Vendor 2

1

7

0.1968 1.47

2.152 0.1966 1.433 0.1966 1.434 0.1964 1.40


Sample No. of

analysis (n)




(%)

Lettuce- Vendor 3

1

7

0.2043 2.89

1.862 0.2044 2.913 0.2043 2.894 0.2049 3.00


Sample No. of

analysis (n)




(%)

Lettuce-Vendor 4

1

7

0.2197 5.79

1.852 0.2187 5.603 0.2186 5.584 0.2195 5.75


137

15 English cabbage

Sample No. of

analysis (n)




(%)

English cabbage

(Vendor 1)

1

7

0.1999 2.06

1.392 0.1996 2.003 0.1999 2.064 0.1997 2.02


Sample No. of

analysis (n)




(%)

English cabbage

(Vendor 2)

1

7

0.2165 5.19

0.792 0.2161 5.113 0.2162 5.134 0.2160 5.09


Sample No. of

analysis (n)




(%)

English cabbage

(Vendor 3)

1

7

0.2058 3.17

2.472 0.2059 3.193 0.2052 3.064 0.2051 3.04


Sample No. of

analysis (n)




(%)

English cabbage

(Vendor 4)

1

7

0.2165 5.19

1.362 0.2162 5.133 0.2166 5.214 0.2171 5.30


138

16 Chinese cabbage

Sample No. of

analysis (n)




(%)

Chinese cabbage

(Vendor 1)

1

7

0.1975 1.60

2.272 0.1977 1.643 0.1973 1.574 0.1973 1.57


Sample No. of

analysis (n)




(%)

Chinese cabbage

(Vendor 2)

1

7

0.2126 4.45

1.112 0.2129 4.513 0.2127 4.474 0.2132 4.57


Sample No. of

analysis (n)




(%)

Chinese cabbage

(Vendor 3)

1

7

0.2148 4.87

2.482 0.2139 4.703 0.2134 4.604 0.2145 4.81


Sample No. of

analysis (n)




(%)

Chinese cabbage

(Vendor 4)

1

7

0.2101 3.98

0.772 0.2103 4.023 0.2105 4.064 0.2103 4.02


139

Fruits17 Tomato

Sample No. of

analysis (n)




(%)

Tomato market

Vendor 1

1

7

0.1936 0.87

2.092 0.1935 0.853 0.1935 0.854 0.1937 0.89


Sample No. of

analysis (n)




(%)

Tomato (New World)

1

7

0.1983 1.75

2.602 0.1980 1.703 0.1979 1.684 0.1984 1.77


Sample No. of

analysis (n)




(%)

Tomato market

Vendor 3

1

7

0.1959 1.30

4.512 0.1955 1.233 0.1959 1.304 0.1953 1.19


Sample No. of

analysis (n)




(%)

Tomato market

Vendor 4

1

7

0.1990 1.89

1.892 0.1992 1.923 0.1994 1.964 0.1990 1.89


140

18 Banana

SampleNo. of

analysis (n)




(%)

Banana market

Vendor 1

1

7

0.1949 1.11

5.312 0.1949 1.113 0.1945 1.044 0.1943 1.00


Sample No. of

analysis (n)




(%)

Banana market

Vendor 2

1

7

0.1929 0.74

4.732 0.1929 0.743 0.1933 0.814 0.1931 0.77


SampleNo. of

analysis (n)




(%)

Banana market

Vendor 3

1

7

0.2228 6.38

3.592 0.2255 6.893 0.2231 6.434 0.2232 6.45


SampleNo. of

analysis (n)




(%)

Banana market

Vendor 4

1

7

0.2020 2.45

2.832 0.2026 2.573 0.2020 2.454 0.2027 2.58


141

19 Long Bean

SampleNo. of

analysis (n)




(%)

Bean Vendor 1

1

7

0.1960 1.32

3.112 0.1959 1.303 0.1964 1.404 0.1962 1.36


Sample No. of

analysis (n)




(%)

Bean Vendor 2

1

7

0.2234 6.49

2.872 0.2235 6.513 0.2231 6.434 0.2253 6.85


SampleNo. of

analysis (n)




(%)

Bean Vendor 3

1

7

0.2091 3.79

1.562 0.2088 3.743 0.2086 3.704 0.2093 3.83

Average iodine (ng/g) = 105.40± 0.06 Analytical coefficient of variation (%) = 0.15

Sample No. of

analysis (n)




(%)

Bean Vendor 4

1

7

0.2012 2.30

5.142 0.2015 2.363 0.2001 2.094 0.2012 2.30


142

20 Pumpkin

SampleNo. of

analysis (n)




(%)

Pumpkin- Vendor 1

1

7

0.1983 1.75

1.872 0.1984 1.773 0.1983 1.754 0.1980 1.70


SampleNo. of

analysis (n)




(%)

Pumpkin- Vendor 2

1

7

0.2010 2.26

1.512 0.2012 2.303 0.2013 2.324 0.2009 2.25


SampleNo. of

analysis (n)




(%)

Pumpkin- Vendor 3

1

7

0.2232 6.45

3.032 0.2230 6.423 0.2253 6.854 0.2235 6.51


SampleNo. of

analysis(n)




(%)

Pumpkin Vendor 4

1

7

0.2095 3.87

1.222 0.2095 3.873 0.2099 3.944 0.2093 3.83


143

Seaweeds21 Sea Grapes (green seaweeds)

Sample No. of

analysis(n)




(%)

Sea grapes Vendor 1

1

15

0.3106 22.94

2.772 0.3103 22.893 0.3041 21.724 0.3112 23.06


Sample No. of

analysis (n)




(%)

Sea grapesVendor 2

1

16

0.3139 23.57

3.842 0.3174 24.233 0.3207 24.854 0.3094 22.72


SampleNo. of

analysis(n)




(%)

Sea grapesVendor 3

1

16

0.2606 13.51

3.712 0.2667 14.663 0.2663 14.584 0.2653 14.40


Sample No. of

analysis (n)




(%)

Sea grapesVendor 4

1

16

0.2617 13.72

2.572 0.2573 12.893 0.2593 13.264 0.2599 13.38


144

22 Lumiwawa (brown seaweeds)

Sample No. of

analysis (n)




(%)

Lumiwawa Vendor 1

1

97

0.3109 23.00

3.782 0.3114 23.093 0.3212 24.944 0.3144 23.66


Sample No. of

analysis (n)




(%)

Lumiwawa Vendor 2

1

110

0.3197 24.66

1.282 0.3238 25.433 0.3213 24.964 0.3212 24.94


Sample No. of

analysis(n)




(%)

Lumiwawa Vendor 3

1

107

0.2248 6.75

3.722 0.2245 6.703 0.2228 6.384 0.2260 6.98


Sample No. of

analysis (n)




(%)

Lumiwawa Vendor 4

1

110

0.2181 5.49

1.722 0.2178 5.433 0.2188 5.624 0.2188 5.62


145

Appendix 6: Absorbance for the recovery study along with the determined iodine contents

in some selected food samples by adding 4, 12 and 18 ng/mL of iodine.

Rice

Sample No. of analysis (n)

Sample dilution

(mL)


Iodine content

in sample (ng/g)

SAE (%)

Punjas Jasmine

rice

1

7

0.2009 2.25 62.87 0.232 0.2020 2.45 68.683 0.2012 2.30 64.454 0.2012 2.30 64.45

Average iodine (ng/g) = 65.11 ± 0.09

Recovery Rice + 4 ng/mL


Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Punjas Jasmine

rice

1

11

0.1973 1.57 68.91 100.30

0.212 0.1969 1.49 65.58 105.383 0.1968 1.47 64.75 106.734 0.1977 1.64 72.23 95.69

RSD (%) = 4.95Recovery (%) = 98.20


Sample Iodine present (ng/g)

Iodine added

(ng/mL)

Total iodine found (ng/g)

Recovery ± RSD (%)

Punjas Jasmine

rice65.11 ± 0.09 4.0 67.87 ± 0.08 98.20 ± 4.95

146



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Punjas Jasmine

rice

1

7

0.2035 2.74 76.60 100.67

0.402 0.2044 2.91 81.36 94.78

3 0.2044 2.91 81.36 94.78

4 0.2027 2.58 72.38 106.54RSD (%) = 5.68

Recovery (%) = 101.05



Iodine added

(ng/mL)


Recovery ± RSD (%)

Punjas Jasmine

rice65.11 ± 0.09 12.0 77.92 ± 0.15 101.05 ± 5.68



Sample dilution

(mL)

Analysis Iodide content

(ng)

Iodine content

in sample

(ng)

%Recovery

SAE (%)

Punjas Jasmine

rice

1

7

0.2049 3.00 84.00 98.94

0.372 0.2059 3.19 89.28 93.09

3 0.2043 2.89 80.83 102.82

4 0.2058 3.17 88.75 93.64RSD (%) = 4.76

Recovery (%) = 103.13

Average iodine (ng) = 85.72 ± 0.14

Sample Iodine present (ng)

Iodine added

(ng/mL)


Recovery ± RSD (%)

Punjas Jasmine

rice65.11 ± 0.09 18.0 85.72 ± 0.14 103.13 ± 4.76

147

Chicken egg


Sample dilution

(mL)


Iodine content

in sample (ng/g)

SAE (%)

Ram Sami & Sons

egg

1

7

0.3031 21.53 602.79 1.442 0.3114 23.09 646.643 0.3041 21.72 608.084 0.3015 21.23 594.34


Recovery Chicken egg + 4 ng/mL


Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Ram Sami & Sons

egg

1

10

0.2726 15.77 630.94 97.78

0.382 0.2732 15.89 635.47 97.093 0.2723 15.72 628.68 98.144 0.2708 15.43 617.36 99.94

RSD (%) = 1.24Recovery (%) = 101.81



Iodine added

(ng/mL)


Recovery ± RSD (%)

Ram Sami & Sons

egg612.96 ± 0.83 4.0 628.11 ± 0.19 101.81 ± 1.24

148



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Ram Sami & Sons

egg

1

11

0.2635 14.06 618.49 101.05

1.152 0.2689 15.08 663.32 94.22

3 0.2626 13.89 611.02 102.28

4 0.2626 13.89 611.02 102.28RSD (%) = 3.87

Recovery (%) = 100.16Average iodine (ng/g) = 625.96 ± 0.57


Iodine added

(ng/mL)


Recovery ± RSD (%)

Ram Sami & Sons

egg612.96 ± 0.83 12.0 625.96 ± 0.56 100.16 ± 3.87



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Ram Sami & Sons

egg

1

11

0.2640 14.15 622.64 101.34

0.912 0.2672 14.75 649.21 97.19

3 0.2622 13.81 607.70 103.83

4 0.2620 13.77 606.04 104.11RSD (%) = 3.15

Recovery (%) = 98.48Average iodine (ng/g) = 621.40 ± 0.45


Iodine added

(ng/mL)


Recovery ± RSD (%)

Ram Sami & Sons

egg612.96 ± 0.83 18.0 621.40 ± 0.45 98.48 ± 3.15

149

Rewa powdered milk


Sample dilution

(mL)


Iodine content

in sample (ng/g)

SAE (%)

Rewa powdered

milk

1

7

0.2767 16.55 463.32 0.782 0.2762 16.45 460.683 0.2805 17.26 483.404 0.2798 17.13 479.70

Average iodine (ng/g)= 471.77 ± 0.41

Recovery Milk + 4 ng/mL


Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Rewa powdered

milk

1

8

0.2649 14.32 458.26 103.82

0.552 0.2649 14.32 458.26 103.823 0.2680 14.91 476.98 99.754 0.2661 14.55 465.51 102.20

RSD (%) = 1.88Recovery (%) = 97.68



Iodine added

(ng/mL)


Recovery ± RSD (%)

Rewa powdered

milk471.77 ± 0.41 4.0 464.75 ± 0.28 97.68 ± 1.88

150



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Rewa powdered

milk

1

11

0.2466 10.87 478.19 101.17

0.422 0.2477 11.08 487.32 99.273 0.2486 11.25 494.79 97.774 0.2489 11.30 497.28 97.28

RSD (%) = 1.77Recovery (%) = 101.16



Iodine added

(ng/mL)


Recovery ± RSD (%)

Rewa powdered

milk471.77 ± 0.41 12.0 489.40 ± 0.20 101.16 ± 1.77



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Rewa powdered

milk

1

10

0.2537 12.21 488.30 100.30

0.602 0.2565 12.74 509.43 96.143 0.2539 12.25 489.81 99.994 0.2530 12.08 483.02 101.40

RSD (%) = 2.30Recovery (%) = 100.59



Iodine added

(ng/mL)


Recovery ± RSD (%)

Rewa powdered

milk471.77 ± 0.41 18.0 492.64 ± 0.29 100.59 ± 2.30

151

Burnswick Sardine


Sample dilution

(mL)


Iodine content

in sample (ng/g)

SAE (%)

Burnswick sardine in vegetable

oil

1

7

0.2929 19.60 548.91 0.992 0.2861 18.32 512.983 0.2907 19.19 537.284 0.2907 19.19 537.28


Recovery Sardine + 4 ng/mL


Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)


oil

1

10

0.2620 13.77 550.94 97.67

0.432 0.2622 13.81 552.45 97.403 0.2637 14.09 563.77 95.454 0.2610 13.58 543.40 99.03

RSD (%) = 1.52Recovery (%) = 102.70



Iodine added

(ng/mL)


Recovery ± RSD (%)


oil

534.11 ± 0.54 4.0 552.64 ± 0.21 102.70 ± 1.52

152



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)


oil

1

11

0.2544 12.34 542.94 100.58

1.122 0.2596 13.32 586.11 93.183 0.2577 12.96 570.34 95.754 0.2534 12.15 534.64 102.15

RSD (%) = 4.26Recovery (%) = 102.27



Iodine added

(ng/mL)


Recovery ± RSD (%)


oil

534.11 ± 0.54 12.0 558.51 ± 0.54 102.27 ± 4.26



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)


oil

1

11

0.2527 12.02 528.83 104.40

0.782 0.2532 12.11 532.98 103.593 0.2565 12.74 560.38 98.534 0.2562 12.68 557.89 98.97

RSD (%) = 3.01Recovery (%) = 98.72



Iodine added

(ng/mL)


Recovery ± RSD (%)


oil

534.11 ± 0.54 18.0 545.02 ± 0.37 98.72 ± 3.01

153

Anchor Butter


Sample dilution

(mL)


Iodine content

in sample (ng/g)

SAE (%)

Anchor butter

1

7

0.2454 10.64 297.96

0.832 0.2491 11.34 317.513 0.2501 11.53 322.794 0.2491 11.34 317.51


Recovery Anchor butter + 4 ng/mL


Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Anchor butter

1

9

0.2353 8.74 314.49 101.10

0.642 0.2349 8.66 311.77 101.983 0.2358 8.83 317.89 100.024 0.2324 8.19 294.79 107.85

RSD (%) = 3.41Recovery (%) = 97.42



Iodine added

(ng/mL)


Recovery ± RSD (%)

Anchor butter 313.94 ± 0.39 4.0 309.74 ± 0.29 97.42 ± 3.41

154



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Anchor butter

1

7

0.2494 11.40 319.09 102.15

0.172 0.2499 11.49 321.74 101.313 0.2489 11.30 316.45 103.004 0.2497 11.45 320.68 101.64

RSD (%) = 0.72Recovery (%) = 98.02



Iodine added

(ng/mL)


Recovery ± RSD (%)

Anchor butter 313.94 ± 0.39 12.0 319.49 ± 0.08 98.02 ± 0.72



Sample dilution

(mL)


Iodine content

in sample (ng/g)

%Recovery

SAE (%)

Anchor butter

1

7

0.2501 11.53 322.79 102.83

0.702 0.2530 12.08 338.11 98.183 0.2539 12.25 342.87 96.814 0.2537 12.21 341.81 97.11

RSD (%) = 2.83Recovery (%) = 101.34



Iodine added

(ng/mL)


Recovery ± RSD (%)

Anchor butter 313.94 ± 0.39 18.0 336.40 ± 0.33 101.34 ± 2.83

155

Appendix 7: Determination (recovery) of iodine in NIST Standard Reference

Material (SRM No. 3530).

NIST SRM No.3530 (Iodised Salt) conc. (ng/mL)

A/min Iodine recovery (ng/mL)

Iodine recovery (%)

15.66

0.2723 15.72 100.360.2754 16.30 104.100.2713 15.53 99.160.2708 15.43 98.560.2727 15.79 100.850.2697 15.23 97.230.2699 15.26 97.47Average 15.61 99.68SAE (%) 0.72

SAE – Standard analytical error.

determination of iodine content...

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