nutrient presentation

STAKEHOLDER PANEL ON INFANT FORMULA & ADULT NUTRITIONALS (SPIFAN)& ADULT NUTRITIONALS (SPIFAN)

MINERALS

Eric Poitevin – Nestlé Research Center (Switzerland)[email protected]

August 24th , 2013Chicago, IL ‐ USA

Background and HistoryInorganicelements

CAS number Atomic Mass (Number)

Classification (Group)

Year of discovery

1st IP (eV)

Calcium (Ca) 7440‐70‐2 40.078 (20) Alkali earth metal (2) 1808 6.113

Magnesium (Mg) 7439‐95‐4 24.305 (12) Alkali earth metal (2) 1755 7.646

Potassium (K) 7440‐09‐7 39.098 (19) Alkali metal (1) 1807 4.34

Sodium (Na) 7440 23 5 22 990 (11) Alkali metal (1) 1807 5 139Sodium (Na) 7440‐23‐5 22.990 (11) Alkali metal (1) 1807 5.139

Phosphorus (P) 7723‐14‐0 30.974 (15) Nonmetal (15) 1669 10.487

Copper (Cu) 7440‐50‐8 63.546 (29) Transition metal (11) 9000 BC 7.726

Manganese (Mn) 7439‐96‐5 54.938 (25) Transition metal (7) 1774 7.432

Zinc (Zn) 7440‐66‐6 65.390 (30) Transition metal (12) 1746 9.394

Iron (Fe) 7439‐89‐6 55.845 (26) Transition metal (8) 3500 BC 7.90

Background and History1800’ Di f j ti l i l• 1800’s Discovery of major essential minerals

• 1867 Liebig‘s first marketed “ideal IF”• 1915 1st SMA IF with minerals added• 1929 Spectrometer with nebulizer and spray chamber• 1929 1st soya IF with minerals added• 1937 1st commercial grating spectrograph (ARL)1937 1 commercial grating spectrograph (ARL)• 1941 1st FDA label declaration (Ca, P, Fe) • 1959 Iron fortification in IF (microcytic anemia)

1960’ H i d IF ( l l t l d C P ti )• 1960’s Humanized IF (renal solute load, Ca:P ratio)• 1974 1st commercial ICP-AES spectrometer• 1981 Codex standard for IF (all minerals regulated)( g )• 2000’s Axial and dual view ICP-AES spectrometers• 2010’s rc/cc-ICP-MS and MIP-AES spectrometers

Background and HistoryMilk Product Water

(%)Fat(%)

Protein(%)

Carbohydrate(%)

Ash/Minerals(%)

Whole liquid 87 3.7 3.3 4.8 0.7

Whole liquid (human) 87.5 3.8 0.9 7.0 0.2

Skimmed liquid 90 <0.1 3.4 4.9 0.75

Whole powder 2‐4 25‐28 25‐27 37‐38 6‐7

Skimmed powder 3‐5 0.7‐1.3 35‐37 49‐52 7.5‐8

Whey powder 3‐6 0 8‐1 5 9‐13 65‐73 7 5‐12Whey powder 3 6 0.8 1.5 9 13 65 73 7.5 12

Infant formula 2‐3 26‐39 10‐18 40‐60 8

Reference:Dairy Powders and Concentrated products A Y Tamine 2009 Wiley‐Blackwell ISBN 978‐1‐4051‐5764‐3Dairy Powders and Concentrated products. A.Y. Tamine.2009.Wiley‐Blackwell. ISBN 978‐1‐4051‐5764‐3.J.H. Kastle. Am J Physiol.,22 ,1908.

Background and HistoryElement Whole milk

(mg/100g)Skimmed milk(mg/100g)

Whey protein powder(mg/100g)

Casein powder(mg/100g)

Ca 114‐123 117‐123 350‐480 1000‐1500

P 103 103 330‐500 800

Na 41‐46 44‐49 280‐460 1200‐1300

K 152‐155 150‐164 650‐1190 1650

Mg 7‐13 70‐150 5‐7 30‐52

Element Cow milk(mg/L)

Human milk(mg/L)(mg/L) (mg/L)

Fe 0.5 0.4

Zn 4 3

C 0 9 0 4Cu 0.9 0.4

Mn 0.03 0.01

Background and HistoryElement Sources Benefits Risks

Ca Dairy products,Legumes, green

Bones and teeth formationNerve stimulation

Stunted growthOsteoporosis, osteomalaciag g

vegetables Cell permeabilityEnzyme activation

pTetanyRickets

P All foods Bones and teeth formationEnergy metabolism enzyme formation

Stunted growthRicketsEnergy metabolism, enzyme formation Rickets

Na All foods Osmotic pressure, Acid‐base balanceCell permeability, nerve stimulation and muscle contraction

Nausea, vomiting, muscularweakness, heart failure

K All foods Same as Na. Protein synthesis and glycogen formation

Rare: nausea, vomiting, cramps

Mg Green vegetables

Structural (bones and teeth) Nerve stimulation

Renal or alcoholism diseaseTremors leading to seizurevegetables

Nuts, grains, milk seafood

Nerve stimulationMuscle contractionEnzyme activation

Tremors leading to seizure

Background and HistoryElement Sources Benefits Risks

Fe Organ meats, fish eggs dairy

Hemoglobin and myoglobin formationEssential component of many enzymes

Anemia, decrease in oxygentransport and cellularfish, eggs, dairy

productsEssential component of many enzymes transport and cellular

immunity, muscle weakness

Zn Oysters, fish, meat, milk, nuts,

Constituent of many enzymesCarbon dioxide transport

Retard growth and sexualdevelopment, poor

grains, legumes Vitamin A utilization immunity and skin lesions, dwarfism, taste sensitivity

Cu Organ meats, oyters, nuts,

Necessary for Fe utilization and hemoglobin formation, constituent of

Anemia, neutropenia, leucopenia, skeletaloyters, nuts,

fruits, driedlegumes

hemoglobin formation, constituent of cytochrome oxidase, involved in boneand elastic tissue development

leucopenia, skeletaldemineralization

Mn Tea, grains, nuts Cofactor of large number of enzymes, anti oxidant involved in normal brain

Reduced growth, abnormalreproduction and boneanti‐oxidant, involved in normal brain

function, reproduction and bonestructure

reproduction and bonedevelopment

Background and History

• Iron deficiency (Anemia) • Zinc deficiency (Retard growth)

Minerals Chemistry• Ca, P: Ca phosphates, Ca caseinate, Ca citrate, ionic form (50% casein bound)

N K f i i f (N i d i h Chl id ) i i f i• Na, K: free ionic forms (Na associated with Chloride) in casein fraction

• Mg: 40% Mg citrate, 16% as free ion and 7% as Mg phosphate

• Fe: 32% Fe citrate, 26% (whey proteins), 24% (caseins) and 14% (fat)

• Zn: 32% bound to casein and the rest bound to colloidal Ca phosphateZn: 32% bound to casein and the rest bound to colloidal Ca phosphate

• Cu: 47% in low Mw fraction, 44% (caseins), 8% (whey proteins) and 2% (fat)

• Mn: 67% bound to caseins, 14% (whey proteins), 18% (low Mw) and 1% (fat)

Regulations

Legislation Infant formulae Follow‐on FormulaeFormulae for special medical

purpose

C d Ali i FAO/WHO (2007) FAO/WHO (1987) C d FAO/WHO (2007)Codex AlimentariusCommission

FAO/WHO (2007)Codex Stan 72‐1981 –Revised Section A

FAO/WHO (1987) Codex Stan 156

FAO/WHO (2007)Codex Stan 72‐1981 –Revised Section B

USA FDA (1985, 2007a,b)Infant Formula Act 21 CFR

Not Applicable FDA (1985, 2007a,b)Infant Formula Act 21 CFRInfant Formula Act 21 CFR

107Infant Formula Act 21 CFR 107

EU EU (2006b) Commission Directive 2006/141/EC

EU (1999) Commission Directive 1999/21/EC

EU (1989) Council Directive 89/398/EC

Australia & New Zealand

Food Standards Australia and New Zealand Standard 2.9.1a Infant Formula Products)

China GB 10765‐2010 (Infant Formula and Follow‐on Formula)China ( )

Recommended Daily Allowances (RDA/AI)Life stage Age Ca (mg/day) P (mg/day) Na (mg/day) K (mg/day) Mg (mg/day)

Infants 0‐6 months 210 (AI) 100 (AI) 120 (AI) 400 (AI) 30 (AI)

Infants 7‐12 months 270 (AI) 275 (AI) 370 (AI) 700 (AI) 75 (AI)

Children 1‐3 years 500 (AI) 460 1000 (AI) 3000 (AI) 80



Children 14‐18 years 1300 (AI) 1250 1500 (AI) 4700 (AI) 410 (360F)

Adolescents‐adults 19‐30 years31‐50 years

1000 (AI) 700 1500 (AI) 4700 (AI) 420 (320 ‐F)400 (310 ‐F)

Adults 51 ‐70 years 1200 (AI) 700 1300 (AI) 4700 (AI) 420 (320 ‐F)

Adults 70 years+ 1200 (AI) 700 1200 (AI) 4700 (AI) 420 (320 ‐F)

Pregnancy 19‐30 years 1000 (AI) 700 1000 (AI) 4700 (AI) 350Pregnancy 19 30 years31‐50 years

1000 (AI) 700 1000 (AI) 4700 (AI) 350360

Breast feeding 19‐30 years31‐50 years

1000 (AI) 700 1000 (AI) 5100 (AI) 310320

Recommended Daily Allowances (RDA/AI)Life stage Age Fe (mg/day) Mn (mg/day) Zn (mg/day) Cu (µg/day)

Infants 0‐6 months 0.27 (AI) 0.003 (AI) 2 (AI) 200 (AI)

Infants 7‐12 months 11 0.6 (AI) 3 220 (AI)

Children 1‐3 years 7 1.2 (AI) 3 340

Children 4‐8 years 10 1.5 (AI) 5 440

Children 9‐13 years 8 1.9 (1.6‐F) (AI) 8 700

Children 14‐18 years 11 (15‐F) 2.2 (1.6‐F) (AI) 11 (9‐F) 890

Adolescents‐adults 19‐30 years31‐50 years

8 (18‐F) 2.3 (1.8‐F) (AI) 11 (8‐F) 900

Adults 51 ‐70 years 8 2.3 (1.8‐F) (AI) 11 (8‐F) 900

Adults 70 years+ 8 2.3 (1.8‐F) (AI) 11 (8‐F) 900

Pregnancy 19‐30 years 220 2.0 (AI) 11 1000Pregnancy 19 30 years31‐50 years

220220

2.0 (AI) 11 1000

Breast feeding 19‐30 years31‐50 years

290290

2.6 (AI) 12 1300

Tolerable Uptake level (UL)Life stage Age Ca P Na K Mg Fe Mn Zn CuLife stage Age Ca P Na K Mg Fe Mn Zn Cu

Infants 0‐6 months ND* ND ND ND ND 40 ND 4 ND

Infants 7‐12 months ND ND ND ND ND 40 ND 5 ND

Children 1 3 years 2500 3000 1500 ND 65 40 2 7 1Children 1‐3 years 2500 3000 1500 ND 65 40 2 7 1

Children 4‐8 years 2500 3000 1900 ND 110 40 3 12 3



Adolescents‐adults 19‐70 years 2500 4000 2300 ND 350 45 11 40 10

Adults 70 years+ 2500 3000 2300 ND 350 45 11 40 10y

Pregnancy 14‐18 years19‐50 years

2500 3500 2300 ND 350 45 9 3440

810

Breast feeding 14‐18 years19 50

2500 4000 2300 ND 350 45 11 3440

81019‐50 years 40 10

* ND: Not Determined

Regulations – Codex (Minerals content)Element Product Target population Min Max GUL UnitElement Product Target population Min Max GUL Unit

Calcium Infant formulaFollow‐up

0 – 6 months6 – 12 months

50 90

‐ 140 mg/100 kcal

Phosphorus Infant formulaF ll

0 – 6 months6 12 th

25 60 ‐ 100 mg/100 kcalFollow‐up 6 – 12 months 60 g/

Ca:P ratio Infant formulaFollow‐up


1:1 2:1 ‐ mg/100 kcal

Sodium Infant formulaF ll


20 20

60 85 ‐ mg/100 kcalFollow‐up 6 – 12 months 20 85 g/

Potassium Infant formulaFollow‐up


60 80

180 ‐ ‐ mg/100 kcal

Magnesium Infant formulaF ll


5 6 ‐ 15 mg/100 kcalFollow‐up 6 – 12 months 6 g/

Iron Infant formulaFollow‐up


0.45 1 2 ‐ mg/100 kcal

Zinc Infant formulaF ll


0.50 5

1.5 mg/100 kcalFollow‐up 6 – 12 months 0.5 ‐ g/

CopperManganese

Infant formulaInfant formula


35 1 ‐

120100 µg/100 kcal

Regulations – Entity (Minerals content)Element Product Target population Minimum Maximum UnitElement Product Target population Minimum Maximum Unit

Ca Infant formulaFollow‐up


60 (USA)50 (EU,NZA) 140 (EU) mg/100 kcal

P Infant formulaF ll


30 (USA)/25‐30*(EU)25 (EU NZA) 30*(EU) 90‐100*(EU)/100(NZA) mg/100 kcalFollow‐up 6 – 12 months 25 (EU,NZA) ‐ 30*(EU) ( )/ ( ) g/

Ca:Pratio

Infant formulaFollow‐up


1.2:1 (NZA) mg/100 kcal

Na Infant formulaF ll

0 – 6 months6 12 th 60 (EU,NZA) mg/100 kcalFollow‐up 6 – 12 months ( , ) g/

K Infant formulaFollow‐up


80 (USA,NZA)60 (EU)

200(USA,NZA)/160(EU)160 (EU)/200 (NZA) mg/100 kcal

Mg Infant formulaF ll


6 (USA)5 (EU NZA) 15 (EU,NZA) mg/100 kcalFollow‐up 6 – 12 months 5 (EU,NZA) ( , ) g/

Fe Infant formulaFollow‐up


0.15(USA)/0.3‐0.45*(EU)/0.8(NZA)0.6‐0.9*(EU)/0.8(NZA)

3 (USA)/1.3‐1.9*(EU)1.7‐2.5* (EU)/1.5(CHN) mg/100 kcal

Zn Infant formulaF ll


0.75*(EU)0 75*(EU)

2.4* (EU)/Zn:Cu15:1(NZA)2 4*(EU) /Z C 20 1(NZA) mg/100 kcalFollow‐up 6 – 12 months 0.75*(EU) 2.4*(EU) /Zn:Cu20:1(NZA) g/

CuMn

Infant formulaInfant formula


60 (USA,NZA)/35 (EU)5 (USA), 4.8 (CHN)

100 (EU)/80(NZA)100(NZA) µg/100 kcal

(* Soy protein)

Minerals calculated range for IFElement K

(mg/100g)Ca

(mg/100g)P

(mg/100g)Na

(mg/100g)Mg

(mg/100g)Regulations limits 300‐1000 250‐ 700 75‐500 100‐425 25‐75

Minimum 460 350 300 110 30Minimum 460 350 300 110 30

Maximum 750 640 400 330 55

Concentrate/premixes 1100‐1650 800‐8000 600‐800 500‐1200 75‐4000

Element Fe(mg/100g)

Zn(mg/100g)

Cu(mg/100g)

Mn(mg/100g)

Regulations limits 0.75‐20 2‐9 0.18‐ 0.9 0.005‐0.5

Minimum 1 2 0.2 0.01

Maximum 10 7.1 0.6 0.4

Concentrate/Premixes 20‐10000 50‐9000 5‐4000 0.1‐9000

Minerals ‐ Sample digestion

Digestion techniqueDry sample

size (g)

Digestion time

(hour)

Degree of digestion (way of losses)

Sample throughput (number of

samples per Basic Equipment

Dry Ashing

losses)run)

1-5Muff le furnace

Crucibles 1-10Incomplete

(volatilization) 10-20

Microwave Open vessel

High Pressure Ashing

Wet ashing

< 0.3 High Pressure asher Quartz tubes

1 Very High (retention)

15-30

0.5-2 Microw ave Glass vessel

< 1 Incomplete (volatilization)

5-10

Microwave Closed vessel

< 0.5 Microw ave Teflon vessel

gashing ( )

< 1 High (retention)

20-40

Minerals ‐ AnalysisAnalyte Macro‐and Micronutrients a yteCriterion FAAS XRF ICP‐OES

Typical IDL (µg/L) XRF (in mg/kg)

1‐1000 1‐1000 1‐100

Precision (%) Short term (< 4 h)

0.1‐1 1‐3 0.3‐2

ac o a d c o ut e ts

Dynamic range (orders of magnitude)

1‐2 4‐5 4‐5

Speed for element (excluding sample preparation time)

Fast 5‐30 sec

(nebulization sample uptake: 5‐6 mL/min

Slow 5 ‐ 10 Minutes

Intermediate 1‐2 minutes

(nebulization sample uptake: ~1 mL/minpreparation time) (nebulization sample uptake: 5 6 mL/min (nebulization sample uptake: ~1 mL/minOverall speed

(including multi‐element capability)

Slow limited multielement capability

6 at most

Fast / Non destructive Multielement capability

Fast Multielement capability

Interferences FewHigh matrix depedency match standard needed

Numerous but well characterized Spectral and non spectral Interferences Few match standard needed

Calibration vs ICP‐OES neededLine selection, internal standardization and

ion buffer needed Multi‐element capability

commonly None Up to 6

Yes Yes

Safety Fire hazard, flammable gasRadiation hazards

Precautions for X‐Ray protectionSafe

inert gas ArPrecautions for X Ray protection inert gas Ar

• ICP‐MS with reaction/collision cells (costly, interferences) • MP‐AES (promising technology, only supplier application notes)

Minerals ‐ Official Methods• AOAC 985.35 Codex Type II method: Ca,Mg,Fe,Zn,Cu,Mn,Na,K‐ Flame atomic absorption spectrophotometry after dry ashingValidated: Infant Formula (enteral soy based RTF whey powder)‐ Validated: Infant Formula (enteral, soy‐based , RTF, whey powder)

• AOAC 986.24 Codex Type II method: P‐Spectrophotometry after dry ashing‐ Validated: Infant Formula (enteral, soy‐based , RTF, whey powder)• AOAC 984.27 Codex Type III method: Ca,Cu,Fe,Mg,Mn,P,K,Na,Zn‐ Validated: Infant Formula (RTF, soy‐based, whey powder)( , y , y p )‐ ICP‐AES after acid digestion• ISO 8070:2007 (IDF 119) Codex Type II method: Ca,Mg,Na,KFlame atomic absorption spectrophotometry after acid digestion‐ Flame atomic absorption spectrophotometry after acid digestion

‐ Validated: Skim and whey milk powders, protein concentate, casein

Minerals ‐ Official Methods• Existing Official Methods suffer from:

‐ use of old and non safe digestion systems (ashing, acid digestion with HCLO4)‐ use of old technology (FAAS, Radial ICP‐AES)l k f ifi i ( ilk b d f l i h‐ lack of specificity (non milk‐based formulas with

added salts)lack of sensitivity (high analytical range of‐ lack of sensitivity (high analytical range of

minerals from µg to g/100g)‐ Relative poor multi‐elemental capacity and high p p y gsample throughput

Minerals ‐ Official Methods• New AOAC 2011.14 First Action: Ca,Mg,Fe,Zn,Cu,Mn,Na,K‐ ICP‐AES after microwave digestionValidated: Fortified Food products including IF‐ Validated: Fortified Food products including IF

Minerals ‐ Official Methods• AOAC 2011.14 First Action method: Ca,Mg,Fe,Zn,Cu,Mn,Na,K‐ Validated with 2 IF, NIST Non fat milk powder, chocolate milk powder with complete digestion covering AOAC sectors 5 6 7powder with complete digestion covering AOAC sectors 5, 6,7 ‐ Ring trial with 9 independent Nestlé laboratories‐ Additional data from 2 proficiency‐tests including 4 IF and 4 Adult nutritionals (2012) for final Action

100 % fat ICP-AES analysisMicrowave digestion

2 4

3

1

0 %

pro

tein

0 % carbohydrate

IN 1

OUT

T- connector

Peristaltic pump

To ICP-AES nebuliser

Internal standards

SampleFrom autosampler

From internal standard solution

75 9

86

100 % protein

100 % carbohydrate

0 % fat

IN 2

Dilute to 50 mL, HNO3 10% v/vDF = 1000.5 g + 5 mL HNO3

Minerals ‐ Official Methods• AOAC 2011.14 First Action method : data for Final Action (end 2013)

‐ Data collected from 2 p‐tests on 2012 (29 independant labs)Satisfactory results for 8 IF and adult nutritionals matrices‐ Satisfactory results for 8 IF and adult nutritionals matrices

Test Matrices (4 samples per p‐test) Mineral Number of labs Unit Consensus Value Range

RSDR (%)

Recovery Min‐Max range for labs

(%)Horrat value

Calcium 9 ‐ 21 mg/100g 78 ‐ 603 < 5 84 ‐ 118 0.64 ‐ 1.09

1 Starter Infant Formula, Milk Based

2 Follow‐Up Infant Formula, Whey Enriched

1 Growing Up Milk Powder

Copper 7 ‐ 21 mg/100g 0.14 ‐ 0.44 < 10 86 ‐ 157 0.66 ‐ 0.79

Iron 12 ‐ 22 mg/100g 1.46 ‐ 8.63 < 8 90 ‐ 124 0.57 ‐ 0.80

Magnesium 7 ‐ 21 mg/100g 26.0 ‐ 60.4 < 4 93 ‐ 107 0.51‐ 0.58

M 7 21 /100 0 055 0 313 < 10 91 117 0 37 1 10Growing Up Milk Powder

1 Adult Complete Formula, Hydrolysed Proteins

2 Adult Complete Formula

1 Adult Formula, Blenderized Natural Foods

Manganese 7 ‐ 21 mg/100g 0.055 ‐ 0.313 < 10 91 ‐ 117 0.37 ‐ 1.10

Phosphorus 8 ‐ 21 mg/100g 67 ‐ 425 < 6 71 ‐ 110 0.60 ‐ 1.10

Potassium 8 ‐ 20 mg/100g 124 ‐ 622 < 6 97 ‐ 110 0.66 ‐ 1.19

Sodium 7 ‐ 22 mg/100g 86 ‐ 231 < 5 94 ‐ 108 0.87 ‐ 1.01

Zinc 7 ‐ 20 mg/100g 1.42 ‐ 6.30 < 6 67 ‐ 110 0.54 ‐ 0.64

Fitness for Purpose

Minerals Working Group

Fitness for purpose statement• A multi‐elemental analytical method to determine level of

nine minerals in infant formula and adult nutritionals asd fi d b AOAC SPIFAN i l d idefined by AOAC SPIFAN, in regulated concentration ranges:

Mineral K(mg/100g)

Ca(mg/100g)

P(mg/100g)

Na(mg/100g)

Mg(mg/100g)

R l i li i (IF) 300 1000 250 700 75 500 100 425 25 75Regulations limits (IF) 300‐1000 250‐ 700 75‐500 100‐425 25‐75

Minimum 200 150 25 30 10

Maximum (IF/Adult) 3200 2400 1600 1600 750

Mineral Fe(mg/100g)

Zn(mg/100g)

Cu(mg/100g)

Mn(mg/100g)

Regulations limits (IF) 0.75‐20 2‐9 0.18‐ 0.9 0.005‐0.5

Minimum 0.5 1 0.2 0.005

Maximum (IF/Adult) 85 32 21 0.5

Fitness for purpose statement

• An analytical method to determine level of nine minerals in i f f l d d l i i l i l finfant formula and adult nutritionals using complete safe digestion systems and updated ICP‐AES technology

• An analytical method having the same performances than AOAC 2011.14 validated for IF and adult nutritionals

• The method needs to be capable of dispute resolution

Questions?Questions?

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