HAL Id: hal-01264545https://hal.archives-ouvertes.fr/hal-01264545

Submitted on 29 Jan 2016

HAL is a multi-disciplinary open accessarchive for the deposit and dissemination of sci-entific research documents, whether they are pub-lished or not. The documents may come fromteaching and research institutions in France orabroad, or from public or private research centers.

L’archive ouverte pluridisciplinaire HAL, estdestinée au dépôt et à la diffusion de documentsscientifiques de niveau recherche, publiés ou non,émanant des établissements d’enseignement et derecherche français ou étrangers, des laboratoirespublics ou privés.

Sensory evaluation and storage stability of UHT milkfortified with iron, magnesium and zinc

Atallah H. Abdulghani, Sangeeta Prakash, Mahmood Y. Ali, Hilton C. Deeth

To cite this version:Atallah H. Abdulghani, Sangeeta Prakash, Mahmood Y. Ali, Hilton C. Deeth. Sensory evaluation andstorage stability of UHT milk fortified with iron, magnesium and zinc. Dairy Science & Technology,EDP sciences/Springer, 2015, 95 (1), pp.33-46. �10.1007/s13594-014-0188-z�. �hal-01264545�

ORIGINAL PAPER

Sensory evaluation and storage stability of UHT milkfortified with iron, magnesium and zinc

Atallah H. Abdulghani & Sangeeta Prakash &

Mahmood Y. Ali & Hilton C. Deeth

Received: 12 April 2014 /Revised: 25 July 2014 /Accepted: 25 August 2014 /Published online: 12 September 2014# INRA and Springer-Verlag France 2014

Abstract Fortification of staple foods is an effective means of addressing dietarydeficiencies of iron, magnesium and zinc, which are common throughout the world.In this work, homogenized whole milk was fortified with iron, magnesium and zinc,individually and collectively, shortly before ultra-high-temperature (UHT) processing(145 °C, 6 s). Sulphates of iron, magnesium and zinc were added to give totalconcentrations (natural plus added) of approximately 25, 50, 75 and 100% of therecommended daily intakes (RDI) per litre of the UHT milk. The highest concentra-tions (100% RDI) were 8, 320 and 16 mg.L−1 for iron, magnesium and zinc,respectively. The fortified milks were analysed to determine the distribution of theminerals, their sensory characteristics and their stability during storage. Zinc and ironpreferentially became associated with the casein fraction, while most of the magnesiumremained in the serum phase. Some sensory panellists could perceive a different taste inUHT milk supplemented to 75% of the RDI per litre, but not in milks supplemented to50%. During storage at 30 °C of milks supplemented to 50%, changes in flavour weredetected in milks supplemented with zinc and iron after 30 and 60 days, respectively;this generally corresponded with increases in oxidation. Viscosity and proteinhydrolysis also increased but not to detrimental levels. It was concluded thatfortification of UHT milk with magnesium and possibly zinc has considerablepotential, but fortification with iron is more challenging due its ability to catalyseoxidation.

Dairy Sci. & Technol. (2015) 95:33–46DOI 10.1007/s13594-014-0188-z

This paper is part of the special issue dedicated to the 2nd Internationl Symposium on Minerals & DairyProducts (MADP2014) held on 26–28th February in Auckland, New Zealand.

A. H. AbdulghaniDepartment of Food Science, College of Agriculture, Tikrit University, Tikrit, Iraq

S. Prakash : H. C. Deeth (*)School of Agriculture and Food Sciences, The University of Queensland, Brisbane 4072, Australiae-mail: h.deeth@uq.edu.au

M. Y. AliDepartment of Food Science, College of Agriculture and Forestry, University of Mosul, Mosul, Iraq

Keywords UHTmilk . Fortification . Iron .Magnesium . Zinc . Sensory

1 Introduction

Inadequate intake of the essential micronutrients, iron (Fe), magnesium (Mg) and zinc(Zn), is common in many countries. This is particularly important for women (Yangand Huffman 2011), and infants and young children (Krebs et al. 2006). The inadequateintake is exacerbated in many cases by the presence in the diet of mineral-bindingsubstances such as phytate and oxalate.

Iron is essential for many metabolic processes in the human body, and two thirds of itis associatedwith haemoglobin in the red cells of the blood. Low levels in the blood causeiron deficiency anaemia which is one of the most widespread disorders in both developedand developing countries (Stolzfus 2003; Toxqui et al. 2013). WHO has estimated thatanaemia, whose primary cause is iron deficiency, affects 52% of preschool-age children,34% of pregnant women and 44% of non-pregnant women in the world (de Benoist et al.2008). A high incidence occurs in African and Asian populations. In the USA, Killipet al. (2007) reported that 2% of adult men were iron deficient but the prevalence inwomen was much higher, up to 20% in black and Mexican-American women. USDA(2009) estimated about 10% of Americans consumed less than the recommended dailyintake (RDI) of iron. Low iron status has been linked to several diseases in women in thepost-menopausal period (Bendich and Zilberboim 2010).

Magnesium is necessary for healthy bone development and maintenance (Abramset al. 2014). About half of the approximately 25 g of magnesium contained in a humanbody is in bones (Hunt and Nielsen 2009). It has been suggested that magnesium maybe even more significant than calcium in bone development (Abrams et al. 2014) and inthe bone disorder, osteoporosis (Abraham and Grewal 1990). USDA (2009) estimatedthat 57% of the US population may have an inadequate intake of magnesium.

Zinc is involved in many body activities such as protein and nucleic acid biosyn-thesis, carbohydrate and lipid metabolism, immunity, growth and fertility (Samman2007; Prasad 2009). The adult human body contains 1.5–2.5 g of zinc, 95% of which isintracellular. Wessells and Brown (2012) estimated that around 17% of the world’spopulation may have inadequate zinc intake while USDA (2009) estimated that 29% ofAmericans may be zinc deficient.

Therefore, there is a need for fortification of foods with Fe, Mg and Zn to alleviatedeficiencies in these elements. The increasing worldwide demand for ultra-high-temperature (UHT)-processed milk (Chavan et al. 2011) makes it an ideal product for suchfortification, particularly as it naturally contains relatively low levels of these elements.UHT milk, which is processed at 135–145 °C for 1–10 s, has a shelf life at roomtemperature of 6 months or more and hence can be distributed widely without refrigeration.

Milk naturally contains low levels of Fe, 0.20–0.5 mg.L−1 (Fransson and Lonnerdal1983; Holt 1997; Hunt and Meacham 2001), and Zn, 3–5 mg.L−1 (Hunt and Meacham2001), and moderate amounts of Mg, 110 mg.L−1 (Holt 1997). Most of the Fe and Zn isin the colloidal phase with the Zn being mostly associated with the colloidal calciumphosphate and the Fe being bound to the polypeptide chains of the caseins (Silva et al.2001). By contrast, approximately two thirds of the Mg resides in the milk serum(Gaucheron 2011; Tsioulpas et al. 2007). The Mg in the colloidal phase is associated

34 A.H. Abdulghani et al.

with the colloidal calcium phosphate. The RDIs for adults for Fe, Mg and Zn accordingto various sources are around 8–15 mg, 280–420 mg and 11–16 mg, respectively.

UHT milk undergoes several changes during storage (Burton 1988). These includechanges in viscosity which can lead to gelation (Kocak and Zadow 1985), proteolysisdue to milk plasmin and bacterial proteinases which results in increases in non-caseinnitrogen (NCN) (López-Fandiño et al. 1993; Valero et al. 2001) and changes in flavourdue, in part, to lipid oxidation (Perkins et al. 2005) which can be accelerated by metalssuch as copper and iron.

The aim of this work was to fortify UHT milk with Fe, Mg and Zn so that each litrecontributed significantly to the RDI of each element. The sensory characteristics andstorage stability of UHT-processed fortified milk was investigated. The distribution ofthe added minerals among the serum, colloidal (micellar casein) and lipid phases of theUHT milk was also investigated to determine whether the added minerals distribute inthe same manner as the natural minerals.

2 Materials and methods

2.1 Experimental plan

The work was carried out in three parts:

1. Sensory evaluation of UHT milk fortified with iron, magnesium and zinc to 75%RDI per litre. Three separate batches of milk were processed and evaluated.

2. Distribution of the elements in UHTmilk fortified with iron, magnesium, zinc and allthree elements to 25, 50, 75 and 100%RDI per litre. The distributions were comparedwith that in unfortified UHT milk. Three separate batches of milk were processed.

3. Storage trial of UHT milk fortified with iron, magnesium and zinc to 50% RDI perlitre at 30 °C for 60 days. Sensory evaluation and physico-chemical analyses(viscosity, lipid oxidation, proteolysis) were performed. Three separate batches ofmilk were processed.

2.2 Milk

Nine different batches of homogenized, pasteurized whole bovine milk were collectedfrom a local dairy processor immediately after production. The concentrations (%, mean±SD, n=9) of the milk components, determined by infrared analysis, were the follow-ing: total solids, 12.6±0.5; fat, 3.89±0.18; protein, 3.27±0.12 (casein, 2.68±0.11; wheyproteins, 0.58±0.02); lactose, 4.67±0.17 and ash, 0.75±0.03 (by difference).

2.3 Minerals

Analytical grade FeSO4·7H2O, MgSO4 and ZnSO4.H2O were obtained from Sigma-Aldrich (Sydney, NSW, Australia). Depending on the recommended daily intake (RDI)for adults, fortified milk samples with different levels were prepared as shown inTable 1. The RDI values of iron, magnesium and zinc adopted were 8, 320 and

Mineral fortification of UHT milk 35

16 mg, respectively. In preparing the magnesium-fortified milk samples, the amount ofmagnesium in normal bovine milk (~120 mg.L−1) was taken into consideration.Trisodium citrate (dihydrate) was added to the milk samples fortified with magnesiumto prevent fouling in the UHT plant (Boumpa et al. 2008; Datta and Deeth 2007;Prakash 2007). All other reagents used were of the highest purity available and at leastanalytical grade.

2.4 Mineral fortification of milk

Accurately weighed amounts of the salts were thoroughly dissolved in 25 mL of high-purity Milli-Q water per each litre of milk. Fortified whole milk samples were preparedby adding the dissolved salts into the milk and mixing gently shortly before heattreatment. Only 25 mL of distilled water was added to the control milk sample.

2.5 UHT processing

UHT processing was conducted in an indirect (tubular) bench-top UHT plant at theUniversity of Queensland as described by Prakash et al. (2010). The temperature–timeconditions in the preheating and high-temperature sections were 95 °C for 8 s and145 °C for 6 s, respectively. The UHT-processed milks were filled aseptically intogamma ray pre-sterilized polyethylene terephthalate (PET) containers in a laminar flowcabinet sterilized by ultra-violet light (UV) shortly before processing and filling. For thekeeping quality trials, all samples were placed in incubators at 30 °C for 60 days.

2.6 Sensory evaluation

Panellists Sensory evaluation of UHT milk samples was conducted by ten trainedpanellists (UQ students and staff, seven males, three females, age 20–50 years, healthysubjects, lactose tolerant) selected after completing scaling exercise and basictaste and aroma exercises as per ISO 22935 and trained for 48 h for QDA® ofpasteurized and UHT milk. The evaluation was conducted in individual booths inthe Food Sensory Laboratory. This study is covered by the Human EthicsApproval #2010000300.

Table 1 Mineral salts added (mg.L−1/mM) to homogenized whole milk

Fortification (% RDI) FeSO4·7H2O MgSO4 ZnSO4·H2O Trisodium citratea

25 9.9/0.04 0 10.9/0.06 0

50 19.9/0.07 198/1.64 21.9/0.12 322/1.1

75 29.8/0.11 594/4.94 32.9/0.18 968/3.3

100 39.8/0.14b 990/8.22c 43.9/0.24d 1,612/5.5

a Added to samples fortified with magnesiumbCorresponds to ~8 mg.L−1 Fec Corresponds to ~320 mg.L−1 Mgd Corresponds to ~16 mg.L−1 Zn

36 A.H. Abdulghani et al.

Sensory evaluation test Triangle tests were performed to determine if addition ofminerals contributed to perceivable differences in colour, odour and taste of the UHTmilk samples. The fortified samples were compared with unfortified UHT milk. Threesamples were presented simultaneously to panellists, two from one formulation (eitherfortified or unfortified UHT milk) and one from a different formulation (either fortifiedor unfortified UHT milk) in a balanced random order. Panellists were asked to identifythe odd sample. To improve the power of the analysis and to be able to detect truediscriminators, duplicate triangle tests were conducted for each batch. The data wasanalysed using chi-square distribution.

The sensory evaluation was carried out in two stages:

Stage 1: In the first stage, UHT milk samples with 75% of the RDI of iron, magnesiumand zinc stored refrigerated for 10 days after processing were evaluated for colour,aroma and taste. The panellists evaluated two different batches of UHT fortified milksover two sessions. In each session, duplicate sets of samples were evaluated for colour,aroma and taste.

Stage 2 (storage trial): In the second stage, UHT milk samples with 50% of the RDI ofiron, magnesium and zinc were evaluated by the trained panel for taste. Soon afterprocessing, the samples were stored refrigerated for 7 days to allow the sulphuryflavour to dissipate. Sensory evaluation was performed on these samples after 7, 30and 60 days of storage at 30 °C.

2.7 Physico-chemical analyses

2.7.1 Distribution of minerals

The skim milk and milk fat fractions were obtained from whole homogenized milk bycentrifuging at 7,000×g and 20 °C for 12 min, using a Beckman Coulter centrifuge (JA-12 rotor, Beckman, USA). The skimmed milk was carefully removed. Sodium azidewas added as preservative (0.1 g.L−1) before storage at 5 °C until analysis.

Ultracentrifugation was used to study the minerals distribution (de la Fuente et al. 1996)between the soluble andmicellar casein phases. The skimmilk samples were centrifuged at100,000×g and 20 °C for 2 h, using a Beckman Coulter, Optima L-100 XPUltracentrifuge(Ti 50.2 rotor, Beckman, USA). The supernatant was carefully removed from the pelletedcasein and filtered through Whatman 40 paper and stored at 5 °C until analysis.

Samples of whole milk, skim milk and soluble phase of the milks were prepared forinitial digestion for 20 min by addition of 6 and 2 mL of nitric and hydrochloric acidsper gramme of sample, respectively. After cooling, 10 mL of triplicate deionized water(TDI) was added to the each sample before digestion in a microwave digester (CEMMSP 1000) for 20 min under pressure of 120 lb per square inch. Hydrogen peroxide(30%, 10 mL) was added to the cooled samples and digested again. Finally, the sampleswere placed into 25-mL volumetric flasks and the volume was made up with TDI. Theconcentrations of the mineral elements were determined by inductively coupled plasmaoptical emission spectrometry (ICP-OES). All analyses were performed in duplicate.

Mineral fortification of UHT milk 37

2.7.2 Viscosity

The viscosity of control UHT milk and fortified milks were measured at 20 °C using aBrookfield DV-II+ viscometer. A UL adapter (ULA) was used for the determination ofviscosity of milk, and the spindle speed was set at 60 rpm. Readings were taken afterthe spindle had been rotating for 30 s, and the mean of three readings was recorded. Theviscosity was expressed in millipascal-second. The tests were done at 0, 15, 30, 45 and60 days of storage period.

2.7.3 Lipid oxidation

The concentration of thiobarbituric acid reactive substances (TBARS) was determinedaccording to King (1962). The tests were done at 0, 10, 20, 30, 40, 50 and 60 days ofstorage, and the average of three readings was recorded. A standard curve was preparedwith 1,1,3,3-tetraethoxypropane (TEP) as the reactive substance as described by King(1962). The standard curve equation obtained was the following: y=0.119x−0.0007.

2.7.4 Proteolysis

The trinitrobenzene sulphonate (TNBS) method was used for measuring the level ofproteolysis (McKellar 1981). The tests were done at 0, 12, 24, 36, 48 and 60 days ofstorage, and the average of three readings of each sample was recorded. A standardcurve was prepared with glycine as the reactive substance as described by McKellar(1981). The standard curve equation obtained was: y=0.208x−0.0029.

2.8 Statistical analysis

All statistical analyses on the physio-chemical data were performed using StatisticalAnalysis Systems (SAS) software (2002, v.9). The general linear model analysiswas performed with the GLM ANOVA tests procedure. When the main ANOVAtests were significant, multiple comparisons of treatments were conducted with theDuncan test.

For the sensory evaluation, duplicate sets of triangle tests were presented in arandomized balanced order and assessed by the trained sensory panel in the samesession. The data were analysed using chi-square distribution. A P value <0.05indicates a significant difference.

3 Results and discussion

3.1 Distribution of iron, magnesium and zinc in milk components

3.1.1 Iron

The average concentration of natural iron in the milk samples was 0.26 mg.L−1. Thedistribution of iron among the serum, casein and fat phases in the control and iron-fortified milk samples is shown in Table 2. The levels were highest in the casein

38 A.H. Abdulghani et al.

fraction, and these increased to a plateau level of 82–89% with fortification up to ~50%RDI per litre, much higher than in the control, 60.9%. Concomitantly, there was amarked decrease in the percentage in the fat phase compared with the control. Thedistributions in the milk samples fortified with all three minerals were similar to thosein the samples fortified with iron only. Silva et al. (2001) reported that iron binds to thepolypeptide backbone of caseins in the micelle; this contrasts with calcium, magnesiumand zinc which are associated with the colloidal calcium phosphate. It is thereforeapparent that added iron also shows a high affinity for the proteins in the casein micelle.This is supported by the work of Raouche et al. (2009) which showed the caseinmicelle to be a good carrier of iron. They reported 84–91% of added iron (up to20 mM) became associated with the casein micelle. The iron-fortified micelles with upto 15 mM added iron were stable to heat.

3.1.2 Magnesium

The average concentration of natural magnesium in the milk samples was 114.9 mg.L−1

(4.59 mM). Since the RDI for magnesium is 320 mg, no magnesium was added for the25% RDI per litre samples but was added for the 50% and higher samples. Sampleswith added magnesium caused severe fouling of the UHT plant. This was overcome byaddition of low levels of trisodium citrate to chelate the magnesium ions.

Processing of the fortified milk samples did not change the general distribution ofmagnesium in the milks. It preferentially resides in the milk serum (62.8%) ofunfortified milk which contrasts with the other major bivalent ion in milk, calcium(Gaucheron 2011). The proportion of magnesium in the serum phase increased withincreasing level of fortification (Table 3). In milk fortified with all three elements, themagnesium distribution was similar with 66.3 to 77.1% being present in the serum.Most likely, most of magnesium was present as a citrate salt since trisodium citrate wasadded as a stabilizer (Flynn and Cashman 1997).

These results agree with that of de la Fuente et al. (2004) who found 75% ofmagnesium in milk fortified with 450 mg.L−1. They reported that two thirds of themagnesium is usually present in the serum; this is close to the figure obtained for thecontrol in this work (62.8%).

Table 2 Iron distribution in con-trol and mineral-fortified UHTwhole milks (mean values, n=3)

UHT milk sample Fortification(% RDI)

Distribution (%)

Serum Casein Fat

Control 0 11.4 60.9 26.6

Fortified with iron only 25 16.6 76.4 7.1

50 5.9 89.0 5.1

75 7.7 87.8 4.3

Fortified with iron,magnesium and zinc

25 9.8 78.3 12.0

50 7.5 85.3 7.20

75 7.3 87.0 5.70

100 7.3 82.7 10.1

Mineral fortification of UHT milk 39

3.1.3 Zinc

The average concentration of zinc in the control milk (4.59 mg.L−1) was within therange (0.2–19 mg.L−1) cited by Walstra et al. (2006). In the control milk, the majority(>80%) of the zinc was associated with the casein, the remainder being distributedapproximately equally between the serum and fat phases (Table 4). In the zinc-fortified milks, an even higher proportion was associated with the micellarcasein fraction, and this generally increased with increasing level of fortifica-tion, up to 92.7%. The distribution is similar to that reported by Flynn et al.(1989). They reported 94% associated with the casein micelle of which 63%was closely associated with the colloidal calcium phosphate and 31% wasloosely associated with the caseins. Silva et al. (2001) reported that in themicelle, zinc, along with magnesium and calcium, is mostly associated with thecalcium phosphate. Since most added zinc became associated with the caseinmicelle, it can reasonably be assumed that it associated with the calcium phosphate also(Table 4).

Table 3 Magnesium distributionin control and mineral-fortifiedUHT whole milks(mean values, n=3)

UHT milk sample Fortification(% RDI)

Distribution (%)

Serum Casein Fat

Control 0 62.8 34.2 3.1

Fortified with magnesium only 25 65.7 31.6 2.7

50 69.8 27.2 3.1

75 66.3 29.2 4.5

Fortified with iron, magnesiumand zinc

25 66.7 30.1 3.1

50 72.4 24.2 3.4

75 77.1 18.2 4.8

100 62.8 34.2 3.1

Table 4 Zinc distribution in con-trol and mineral-fortified UHTwhole milks (mean values, n=3)

UHT milk sample Fortification(% RDI)

Distribution (%)

Serum Casein Fat

Control 0 10.2 80.6 9.3

Fortified with zinc only 25 5.0 87.7 7.3

50 4.3 89.3 6.4

75 3.6 92.7 3.7

Fortified with iron,magnesium and zinc

25 4.4 87.0 8. 7

50 5.7 88.3 6.0

75 5.4 88.0 6.6

100 5.6 77.3 17.1

40 A.H. Abdulghani et al.

3.2 Sensory evaluation

3.2.1 Stage 1: UHT milks fortified to 75% RDI per litre

Triangle tests based on overall sensory differences were used to determine whetherfortified UHT milk samples could be differentiated from control UHT milk samples bya trained sensory panel. The results for colour and aroma (not presented) indicated nosignificant difference in the responses for the fortified and control samples (P>0.05) inall the three sessions.

The responses for taste indicate that panellists significantly perceived differences intaste of the iron-fortified samples (P>0.05) but not of the zinc-fortified samples(P>0.05) compared with the control samples in all three sessions. In two of the threesessions, the panellists significantly (P>0.05) identified differences in taste of themagnesium-fortified samples from the control UHT milk samples (Table 5).

Since the panel could perceive differences between the control UHT milk and the UHTmilk fortified with Fe andMg to 75%RDI, it was decided to reduce the level of fortificationto 50% RDI for the storage trial in which sensory evaluation was also conducted.

No sensory evaluation was performed on samples fortified to less that 50% RDI.

3.2.2 Stage 2: UHT milks fortified to 50% RDI per litre and stored at 30 °Cfor 60 days

The storage trial contained an additional sample, one to which all three minerals (ofiron, magnesium and zinc, each to 50% RDI) were added. The results of the duplicatetriangle tests with fortified and control UHT milk samples after 7, 30 and 60 days ofstorage are presented in Table 5. For samples with P values <0.05, the panellists wereable to differentiate between fortified and control samples. The results suggest that after7 days of storage, the panellists could not perceive any difference in taste betweencontrol and fortified UHT milk samples. However, they could differentiate the iron-

Table 5 Panellists’ ability to differentiate between control UHT whole milk and UHT whole milk fortifiedwith Mg, Zn, Fe and all (Mg, Zn and Fe) to 75 and 50% of recommended daily intake per litre

P value

Mg Zn Fe All

75% of RDI per litre

Storage (day 10) 0.00 0.07 0.01 NA

Storage (day 10) 0.09 0.09 0.01 NA

Storage (day 10) 0.00 0.09 0.01 NA

50% of RDI per litre

Storage (day 71 ) 0.22 0.43 0.22 0.06

Storage (day 30) 0.09 0.59 0.02 0.61

Storage (day 60) 0.67 0.00 0.00 0.49

NA not applicable

Mineral fortification of UHT milk 41

fortified UHT milk from the control sample after 30 days of storage and after 60 days ofstorage, they could differentiate both iron- and zinc-fortified UHT milk samples fromthe control (P>0.05, Table 5). The panellists could not perceive any sensory differencebetween the control and the samples fortified with magnesium and all three minerals.

The sensory results show the potential for as well as the limitations in fortifying UHTmilk with Fe, Mg and Zn. The Fe results show that addition of ferrous sulphate causesinstability in the product during storage which would limit its application. Althoughferrous sulphate is soluble in milk, imparts little colour and contains iron in a bioavail-able form, Fe fortification using ferric iron may be preferable for product stability.Recently, Mittal et al. (2012, 2013) reported the preparation of a soluble ferric iron–milkprotein complex from calcium-depleted milk which has potential for fortification offoods including UHT milk products. The use of iron–milk protein complexes forfortifying foods has recently been reviewed by Ellis et al. (2012). The results for Mgindicate considerable potential for fortification of UHTmilk, provided it is accompaniedby addition of citrate to prevent fouling during processing, while the zinc results suggestonly low levels could be incorporated without induction of off-flavours during storage.

3.3 Physico-chemical characteristics of UHT milk fortified to 50% RDI and storedat 30 °C for 60 days

3.3.1 Viscosity

The viscosity data are shown in Fig. 1. The viscosity was similar for all milk samples atzero time. After 60 days, there were no significant differences between the iron-,magnesium- and zinc-fortified milks, but the results showed significant differenceswith the control (P>0.05). The viscosity of all milk samples gradually increased duringstorage (P>0.05) with that of the milk samples fortified with all three mineralsincreased significantly (P>0.05) more than the viscosities of the control and samplesfortified with one mineral only. However, all remained below 10 mPa.s, the level atabout which gelation occurs (Kocak and Zadow 1985). There were no signs of clottingor gelation (Datta and Deeth 2007).

0 15 30 45 60

Vis

cosi

ty (

mP

a.s)

Storage time (days)Fig. 1 Viscosity during storage of UHTwhole milks fortified with Fe, Mg and Zn to 50% recommended dailyintake per litre (mean values, n=3). Control. Fe. Mg. Zn. All

42 A.H. Abdulghani et al.

3.3.2 Lipid oxidation

All values of TBARS were below the threshold levels at which an oxidized flavourcould be perceived (King 1962). They showed moderate increase during the first40 days of storage and then a more rapid increase up to 60 days of storage. Therewas no significant difference between the control and zinc-fortified samples whichshowed the smallest increase during the storage time (Fig. 2). The greatest increaseoccurred in the samples containing iron. Similar results were obtained with the iron-fortified sample and the sample fortified with all three minerals. According to King(1962), the description of the flavour of milk with the highest levels of oxidationobserved here would be “questionable to very slight”. Although whole milk (3.89% fat)was fortified, processed and bottled in PET transparent containers, the low levels ofoxidation in the milk samples can be attributed to their being stored in the dark andhaving minimal headspace (Smet et al. 2009). In addition, sulfhydryl compounds

0.050

0.075

0.100

0.125

0.150

0.175

0.200

0.225

0 10 20 30 40 50 60

TB

AR

S (n

mo

l TE

P/m

L)

Storage time (days)

Fig. 2 Thiobarbituric acid reactive substances (TBARS) during storage of UHTwhole milks fortified with Fe,Mg and Zn to 50% recommended daily intake per litre (mean values, n=3). Control. Fe.

Mg. Zn. All

1.10

1.15

1.20

1.25

1.30

1.35

1.40

1.45

0 12 24 36 48 60Pro

tein

hyd

roly

sis

(G

lyci

ne,

µm

ol/m

l)

Storage time (days)Fig. 3 Proteolysis during storage of UHT whole milks fortified with Fe, Mg and Zn to 50% recommendeddaily intake per litre (mean values, n=3). Control. Fe. Mg. Zn. All

Mineral fortification of UHT milk 43

produced during UHT treatment would have acted as antioxidants (Shipe et al. 1978).Greater oxidation would be expected with a longer storage period, especially in thesamples containing iron which is a well-known pro-oxidant.

3.3.3 Proteolysis

The level of proteolysis in all milks, including the control, increased slowly during thefirst 36 days and then increased at a higher rate (Fig. 3). Overall, the increases weresmall and there was no obvious effect of the minerals added on the proteolysis level.

4 Conclusion

Sensory evaluation of UHT milks fortified with various levels of iron, magnesium andzinc revealed that the trained panellists could perceive the taste of milk samplesfortified to 75% of RDI per litre but not those fortified to 50%. During storage for60 days at 30 °C, milk fortified to 50% of RDI per litre with iron and zinc exhibited off-flavours mostly due to oxidation, which was generally consistent with the TBARSresults. Viscosity and proteolysis levels increased slightly during the storage period butnot to a level which would cause instability in the product.

The distributions of iron, magnesium and zinc among the casein, serum and fatphases of the fortified milk samples were similar to those in unfortified milks.However, increasing fortification generally increased the proportion in the preferredphase, that is, casein micelle for iron and zinc and serum for magnesium. Neither UHTprocessing nor fortification changed the distribution of the minerals in milk.

Acknowledgements The authors extend sincere appreciation to Honest Madziva, Lesleigh Force and Shiou(Aaron) Pang for assistance with the practical lab work. They also thank Olena Kravchuk for her help instatistics. The support of Iraqi Ministry of Higher Education and Dairy Innovation Australia Ltd is gratefullyacknowledged.

References

Abraham GE, Grewal H (1990) A total dietary program emphasizing magnesium instead of calcium: effect onthe mineral density of calcareous bone in postmenopausal women on hormonal therapy. J Reprod Med35:503–507

Abrams SA, Chen Z, Hawthorne KM (2014) Magnesium metabolism in 4-year old to 8-year old children. JBone Miner Res 29(1):118–122

Bendich A, Zilberboim R (2010) Iron and women’s health. In: Yehuda S, Mostofsky DI (eds) Iron deficiencyand overload: from basic biology to clinical medicine (nutrition and health). Humana Press, NewYork, pp327–350

Boumpa T, Tsioulpas A, Grandison AS, Lewis MJ (2008) Effects of phosphates and citrates on sedimentformation in UHT goats’ milk. J Dairy Res 75:160–166

Burton H (1988) UHT processing of milk and milk products. Elsevier Applied Science, LondonChavan RS, Chavan SR, Khedkar CD, Jana AH (2011) UHT milk processing and effect of plasmin activity on

shelf life: a review. Comp Rev Food Sci Food Safety 10:251–268Datta N, Deeth HC (2007) UHT and aseptic processing of milk and milk products. In: Advances in thermal

and non-thermal food preservation. Blackwell Publishing, Ames, pp 63–69

44 A.H. Abdulghani et al.

de Benoist B, McLean E, Egli I, Cogswell M (eds) (2008) Worldwide prevalence of anaemia 1993–2005: WHO global database on anaemia. WHO Press, World Health Organization, Geneva,Switzerland

De la Fuente MA, Fontecha J, Juárez M (1996) Partition of main and trace minerals in milk: effect ofultracentrifugation, rennet coagulation, and dialysis on soluble phase separation. J Agric Food Chem 44:1988–1992

De la Fuente MA, Belloque J, Juarez M (2004) Mineral contents and distribution between the soluble and themicellar phases in calcium-enriched UHT milks. J Sci Food Agric 84:1708–1714

Ellis A, Mittal V, Sugiarto M (2012) Innovation in iron fortification. Is the future in iron-binding milk proteins.In: Ghosh D, Das S, Bagshi D, Smarta RB (eds) Innovation in healthy and functional foods. CRC Press,Boca Raton, pp 249–268

Flynn A, Cashman K (1997) Nutritional aspects of minerals in bovine. In: Fox PF (ed) Advanced dairychemistry. Lactose, water, salts and vitamins. Chapman and Hall, London, pp 251–302

Flynn A, Singh H, Kiely J, Fox PF (1989). Effect of colloidal calcium phosphate on distribution andbioavailability of zinc in cow's milk and infant formulae. J Dairy Res 56:547–547

Fransson G-B, Lonnerdal B (1983) Distribution of trace elements and minerals in human and cow’s milk.Pediatr Res 17:912–915

Gaucheron F (2011) Milk salts| distribution and analysis. In: Fuquay JW, Fox PF, McSweeney PLH (eds)Encyclopedia of dairy sciences (second edition), vol 3. Academic, San Diego, pp 908–916

Holt C (1997) The milk salts and their interaction with casein. In: Fox PF (ed) Advanced dairy chemistry 3:lactose, water, salts and vitamins, 2nd edn. Chapman & Hall, London, pp 233–256

Hunt CD, Meacham SL (2001) Aluminum, boron, calcium, copper, iron, magnesium, manganese, molybde-num, phosphorus, potassium, sodium, and zinc: concentrations in common Western foods and estimateddaily intakes by infants; toddlers; and male and female adolescents, adults, and seniors in the UnitedStates. J Am Diet Assoc 101:1058–1060

Hunt CD, Nielsen FH (2009) Nutritional aspects of minerals in bovine and human milks. In: McSweeneyPLH, Fox PF (eds) Advanced dairy chemistry vol 3 lactose, water, salts and minor constituents. Springer,New York, pp 391–456

Killip S, Bennett JM, Chambers MD (2007) Iron deficiency anemia. Am Fam Physician 75(5):671–678King RL (1962) Oxidation of milk fat globule membrane material. 1. Thiobarbituric acid reaction as a measure

of oxidized flavor in milk and model systems. J Dairy Sci 45:1165–1171Kocak HR, Zadow JG (1985) Age gelation of UHT whole milk as influenced by storage temperature. Aust J

Dairy Technol 40:14–18Krebs NF, Westcott JE, Butler N, Robinson C, Bell M, Hambidge KM (2006) Meat as a first complementary

food for breastfed infants: feasibility and impact on zinc intake and status. J Pediatr Gastroenterol Nutr42(2):207–214

López-Fandiño R, Olano A, Corzo N, Ramos M (1993) Proteolysis during storage of UHT milk: differencesbetween whole and skim milk. J Dairy Res 60:339–347

McKellar RC (1981) Development of off-flavors in ultra-high temperature and pasteurized milk as a functionof proteolysis. J Dairy Sci 64:2138–2145

Mittal V, Ellis A, Das S, Ye A, Singh H (2012) Micronutrient fortification process and its usage (ironfortification for dairy products). New Zealand patent application 600756

Mittal V, Ye A, Ellis A, Das S, Singh H (2013) Formation of soluble protein-iron complexes in calcium-depleted milk. Abstract of poster presented at Int Dairy Fed World Dairy Summit, Rediscovering milk, 28October - 1 November, Yokohama, Japan

Perkins ML, Elliott AJ, D’Arcy BR, Deeth HC (2005) Stale flavour volatiles in Australian commercial UHTmilk during storage. Aust J Dairy Technol 60:231–237

Prakash S (2007) Burn-on in ultra-high-temperature processing of milk. PhD thesis, University of Queensland,Queensland

Prakash S, Huppertz T, Kravchuk O, Deeth HC (2010) Ultra-high temperature processing of chocolate-flavoured milk. J Food Eng 96:174–184

Prasad AS (2009) Impact of the discovery of human zinc deficiency on health. J Am Coll Nutr 28:257–265Raouche S, Dobenesque M, Bot A, Lagaude A, Marchesseau S (2009) Casein micelles as a vehicle for iron

fortification of foods. Eur Food Res Technol 229:929–935Samman S (2007) Zinc. Nutr Diet 64(Suppl 4):S131–S134Shipe WF, Bassette R, Deane DD, Dunkley WL, Hammond EG, Harper WJ, Kleyn DH, Morgan ME, Nelson

JH, Scanlan RA (1978) Off flavors of milk: nomenclature, standards, and bibliography. J Dairy Sci 61:855–869

Mineral fortification of UHT milk 45

Silva FV, Lopes GS, Nóbrega JA, Souza GB, Nogueira ARA (2001) Study of the protein-boundfraction of calcium, iron, magnesium and zinc in bovine milk. Spectrochim Acta B At Spectrosc56:1909–1916

Smet K, de Block J, de Campeneere S, de Brabander D, Herman L, Raes K, Dewettinck K, Coudijzer K(2009) Oxidative stability of UHTmilk as influenced by fatty acid composition and packaging. Int Dairy J19:372–379

Stolzfus RJ (2003) Iron deficiency: global prevalence and consequences. Food Nutr Bull 24(4Suppl):S99–S103

Toxqui L, Pérez-Granados AMP, Blanco-Rojo R,Wright I, González-Vizcayno C, Vaquero MP (2013) Effectsof an iron or iron and vitamin D-fortified flavored skim milk on iron metabolism: a randomized controlleddouble-blind trial in iron-deficient women. J Am Coll Nutr 32(5):312–320

Tsioulpas A, Lewis MJ, Grandison AS (2007) Effect of minerals on casein micelle stability of cows’ milk. JDairy Res 74:167–173

USDA (United States Department of Agriculture, Agricultural Research Service) (2009) Nutrient intakes.Percent of population 2 years old and over with adequate intakes based on average requirement (http://www.ars.usda.gov/Services/docs.htm?docid=15672, accessed 23 February 2014)

Valero E, Villamiel M, Miralles B, Sanz J, Martínez-Castro I (2001) Changes in flavour and volatilecomponents during storage of whole and skimmed UHT milk. Food Chem 72:51–58

Walstra P, Wouters J, Geurts T (2006) Milk components. In: Dairy science and technology. CRC Press, Taylor&Francis Group, Boca Raton

Wessells KR, Brown KH (2012) Estimating the global prevalence of zinc deficiency: results based on zincavailability in national food supplies and the prevalence of stunting. PLoS ONE 7(11):e50568

Yang Z, Huffman SL (2011) Review of fortified food and beverage products for pregnant and lactating womenand their impact on nutritional status. Matern Child Nutr 7(Suppl 3):19–43

46 A.H. Abdulghani et al.