IS 9967-1: Milk and milk products - Determination of nitrate and nitrate contents, Part 1: Method using cadmium reduction and spectrometry

2008·6.9 MB·English

by  Bureau of Indian Standards

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timely dissemination of this information in an accurate manner to the public.
“जान1 का अ+धकार, जी1 का अ+धकार” “प0रा1 को छोड न’ 5 तरफ”
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“The Right to Information, The Right to Live” “Step Out From the Old to the New”
IS 9967-1 (2008): Milk and milk products - Determination of
nitrate and nitrate contents, Part 1: Method using cadmium
reduction and spectrometry [FAD 19: Dairy Products and
Equipment]
“!ान $ एक न’ भारत का +नम-ण”
Satyanarayan Gangaram Pitroda
““IInnvveenntt aa NNeeww IInnddiiaa UUssiinngg KKnnoowwlleeddggee””
“!ान एक ऐसा खजाना > जो कभी च0राया नहB जा सकता हहहहै””ै”
Bhartṛhari—Nītiśatakam
“Knowledge is such a treasure which cannot be stolen”

IS 9967 (Part 1) : 2008
ISO 14673-1 : 2004

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Indian Standard
MILK AND MILK PRODUCTS
DETERMINATION OF NITRATE AND
NITRITE CONTENTS
PART 1 METHOD USING CADMIUM REDUCTION AND
SPECTROMETRY
( Second Revision )
ICS 67.100.01
© SIS2008
BUREAU OF INDIAN STANDARDS
MANAK SHAVAN. 9 SAHADUR SHAH lAFAR MARG
NEW DELH I ' 10002
Aprif 2008 Price Group 7
Dairy Products and Equipment SectionalCommittee.FAD 19
NATIONAL FOREWORD
This Indian Standard (Part 1) (Second Revision) which is identical with ISO 14673-1 :2004 'Milk and
milk products - Determinationof nitrate and nitrite contents:Part 1Method using cadmium reduction
and spectrometry'issued by the International Organization for Standardization (ISO) was adopted by
the Bureau of IndianStandardson therecommendationofthe Dairy Productsand Equipment Sectional
Committee and approval ofthe Food and Agriculture Division Council.
This standard was first published in 1981 based on the earlier version of International Standard.
ISO4099 :1978andsubsequently revisedin1997 toalign with the revised ISO4099 :1984 underdual
numbering and its scope covered determination of nitrate and nitrite contents in cheese using
cadmium reduction and spectrometry. Subsequently ISO 4099 was withdrawn and revised into three
parts of ISO 14673 to include in its scope a variety of milk and milk products and also including two
more methods fordetermination of nitrate and nitrite contents.
Subsequently toalign with ISO 14673 on thesubject, IS9967 is now being revised into following three
parts underthe general title:'Milk and milk products - Determination of nitrate and nitrite contents'
Part 1 Method using cadmium reduction and spectrometry and is identical to
ISO 14673-1 :2004;
Part 2 Method using segmented flow analysis (routine method) and is identical to
ISO 14673-2 :2004; and
Part 3 Method using cadmium reduction and flow injection analysis with in-line dialysis
(routine method) and isidentical to ISO 14673-3 :2004.
The text of ISO Standard has been approved as suitable for publication as an Indian Standard
without deviations.Certain conventions are,however, not identical to those used in Indian Standards.
Attention IS particularlydrawn to the following:
a) Wherever the words 'International Standard'appear refernng to this standard, they should
be read as'Indian Standard'.
b) Comma (,) has been used as a decimal marker while in Indian Standards, the current
practice isto useapoint (.)asthedecimal marker.
Inthisadoptedstandard,referenceappearstocertain InternationalStandardsforwhich IndianStandards
also exist. The corresponding Indian Standards which are to be substituted in their places are given
below along withtheir degree of equivalence forthe editions indicated:
tnternetionet Standard Corresponding Indian Standard Degree of
Equivalence
IS 460 Specification for test sieves: }
ISO 565 : 1990 Test sieves
Metalwirecloth,perforatedmetal (Part 1) : 1985 Wire cloth test sieves Technically
plate and electroformecl sheet (thirdrevision) Equivalent
Nominal sizes ofopenings (Part 2) : 1985 Perforated plate test
sieves(third revision)
ISO 648 : 1977 Laboratory IS 1117 :1975 Specification for one-mark do
glassware - One-mark pipettes pipettes (first revision)
ISO 835-1 : 1981 laboratory IS 4162 (Part 1) : 1985 Specification for
do
glassware Graduated graduated pipettes: Part 1 General
pipettes - Part 1: General requirements (first revision)
requirements
(Continued on thirdcover)
IS9967 (Part 1) : 2008
ISO 14673·1 :2004
Indian Standard
MILK AND MILK PRODUCTS
DETERMINATION OF NITRATE AND
NITRITE CONTENTS
PART 1 METHOD USING CADMIUM REDUCTION AND
SPECTROMETRY
( Second Revision )
WARNING- The use ofthis International Standard may involvehazwous materials, operations and
equipmentThisstandarddoes notpurportto addressaUthesafetyproblemsassociatedwith its use.
It is the responsibility of the user of this standard to establish Hfety and health practices and
determinetheapplicabilityofregulatorylimitationspriorto use.
1 Scope
This part of ISO 1467311DF189specifies 8 method for the determinatIon ofthe nitrate and nrtntecontents of
milkandmilkproductsbycadmium reductionandspectrometry The method isapplicableto
whole and partly skimmedandskimmeddriedmilk;
hard, semi-hardandsoft cheeses;
processedcheese;
whey cheese, caseinsandcaseinates,anddriedwhey.
The methodmay be performedusingautomaticequipment, inparticularby segmentedflowanalysIS(SFA) or
ftCINInjection analysis(FIA), thus reducingcadmium contaminationinlaboratoryworkplacesandwastewater
NOTE TheM rne«hod.aredeacribedinISO14673-2iIOF189-2andISO1~73-3110F 189-3. respectrwly
2 Normative references
The foIIa'Mng referenced documents are indape,..able for the application of this document For dated
references, only the edition cited applies. For undated references. the latest eCXtion of the referenced
document(inducinglInylImendments)applies.
ISO565, restsilMts- Met'"wint doth. perforetedmetalpI«eende/edroformedsheet- Nominalsilesof
openings
ISO648,LaboratoryQlassWIJlW- One-miDpipettes
ISO 835-1,l.IIborIItoryglassware- Greduetedpipettes- Part 1:Generalrequirements
ISO 1042,LetJoretoryglassware- On&-merl<voIum«ric flaSKS

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IS 9967(Part 1):2008
ISO14673-1 :2004
3 Termsand definitions
Forthe purposesofttusdocument, the followingterms and definttionsapply.
3.1
nitratecontent
mass fractIonofnitratedeterminedbythe procedurespecifiedinthis partofISO 1467311DF 189
NOTE ThenltratecontertISexpt'essed asthemassinmiMigramsofnitrateions(N03- )perkilogram ofproduct
3.2
nitritecontent
massfractionatnitrite determinedbytheprocedurespecifiedinthispartofISO 1467311DF 189
NOTE Thenitritecontent isexpresaed8Sthemassinmiligramsofnitriteions (002- )perkilogramofproduct.
4 Principle
Atest portion is dispersedin warm water. with precipitation ofthe fat and proteins.then filtration. The nitrate
Ionsare reducedtonatriteionsinaportionofthe filtrateby meansofcopperizedcadmium.
A red colour is developed in poI'tions of both unreduced filtrate and the reduced solution, by addition of
sulfanilamideand N-1-naphthylethylenediaminedhydrochloride.Spectrometricmeasurementsare carriedout
atawavelen~hof 538 nm.
The nwlte content of the sample and the total nitrite content after reduction of nitrate ions are calculated by
companng the measured abeorbances with those ~ a setof sodum nitrite calibration solutions. The nitrate
content IScalculatedfrom the ci1ferencebetweenthesetwocontents.
5 Reagents
Useonly reagentsofrecognizedanalyticalgradeunlesaothefwtsespecified.
5.1 W••r.distilletJ0(deioniZed,orwaterofeqUIValentpurity. freefrom nitrateand nitriteions.
ToaVOId the poulble ,"cl~onof small gas bubbles in the copperized cadmium column (9.16). freshly boil
the distilledor deioniZedwaterand cool toroom temperature.Usethe thus-preperedwaterfor thepreparation
ofthe column(91),to checkthe reducingc.peatyofthecolumn(9.2). and toregeneratethecolumn(9.3).
5.2 cadmium granu....ofdameter0.3mmto0.8 mm.
Preparecadmium l1anulett,Ifnot BV8Ilabiecommercially, _ follows.
PlaceaSUItablenumberofzincrods in abeaker.Covertheroda with cadmium sulfatesolution (5.3). SCrape
the cadmium s\X>ngefrom lt1erods from time totime CN8f8 period~24 h.Removethezincrods and decant
the liqUIduntil only suftlaentremainsto cover thecadmium sponge.W_h the spongetwo orthreetimeswith
water. Transfer the cadmium sponge to a laboratory blender together with 400ml ~ the hydrochloric acid
workingsolution(5 7) 8lldblendfor afewsecondsto obtaingranules ofthe requiredsize.Returnthe contents
ofthe blendertothe beakerand leave10standforseveralhours,whilestirringoccasionallyto remCNebubbles.
Decantmostoftheliquid and immedatelycoppenzethe granules88describedin9.1.
WARNING - BecauseofitlI toxicity, the usedcadmium should bedelivered aschemical wasteto the
relevantauthorttJes.

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IS9967 (Part 1): 2008
ISO14673·1 : 2004
5.3 Cadmiumsulfatesolution, c(CdSO..8H20) =401)'1
DIssolve409of cadmium sulfate solutIon Inwater In a , 000ml volumetrIcflask (64) DIlute tothe mark WIth
water andmix
5.4 Copper(lI)sulfatesolution, c(CuSO..5H =20gil
20)
Dissolve 20g of copper(ll) sulfate In water In a , 000 ml volumetnc nask (64) Dilute to the mark WIth water
andmix
5.5 Hydrochloricacid (HCI),(P20 = 1,19glml)
5.6 Dilutehydrochloricacid, c(HCI) '"2molJl.
Carefully add 160ml of hydrochloric acid (5.5) to about 700 ml of water In a , 000 ml volumetnc flask (64)
while regularly sli'>1rlingthe contents. Cool the contentstoroomtemperature DIlute tothe mark WIthwater and
mixcarefully.
5.7 Hydrochloricacid working solution, c(HCI)..0,1 molll
Add 50ml of dilute hydrochloric acid (56) to a 1000 ml volumetric nask (64) Dilute to the mark WIthwater
andmix.
5.8 Zincsulfatesolution,c(ZnSO..·7H20)=535gil
DIssolve53,5 0ofzinc sulfateinwater ina ,00mlvolumetncnask (64) DIlutetothe markWIthwater and mIx
5.9 Potassium hexacyanoferrllte(1l)solution, c(K.rFe(CN)s1·3~O)" 172gil
Dissolve 17,20of potassium hexacyanoferrate(lI)trihydratein waterIn a 100ml volumetncnask (64) Dilute
tothemarkwithwater and mix
Dissolve 33,Sg atEDTA in about 900ml water in a 1000ml volumetric flask (64) DIlute to the mark WIth
water andmix.
5.11 SolutionI.
Carefully add 450 ml 01 hydrochloric acid (5.5) to a 1000 ml volumetric nask (64) Dilute to the mark 'Mth
water andmix.
5.12 SolutionII.
Dissolveby heatingon awater beth (615)0,50ofs~anllamide (NH~eH..S02N~) in a mixture of 75ml of
water and 5mlof hydrochloricaCId(5.5) in a100mlvolumetricflask (64).Cool toroom temperature DIluteto
themarkwithwaterand mix. Filter the obtainedsolution, ifnecessary
5.13 SolutionIn
Dissolve 0,19 of N-1-naphthyt ethylenediamine dhydrochloride (C1oH.,NHC~CH2NH22HCI)in water in a
100mlvolumetricflask (6.4). Dilutetothe markwith water andmix.Filterthe obtainedsolution, Ifnecessary
Thesolution maybestoredfor upto 1weekinawetl-stopperedbrown banle in arefrigerator
5.14 Sodiumnitritestocksolution(NaN02)·
Dry a few grams atsodium nitrite in an oven (6.16) at 110'C to 12O·C to constant man (Ie until the
differencebetween twosuccessive'N8ighings doesnot exceed 1mo) DIssolve0,1500ofthesodium nrttlteto
water ina 1000 mlvolumetricflask(6.4). Dilute tothe mark withwaterand mix.

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IS9967 (Part 1) :2008
ISO14673-1 :2004
5.15 Sodium nitriteworkingsolution.
Preparethe sodlum nitrite working solution ontheday ofuse.Transferwith apipette (6.5), 10ml ofthestock
sotuuon(514) and 20ml of the buffer solution (5.19) to a 1000ml volumetric flask (6.4). Diluteto the mark
withwater andrnrx Thenitrite contentofthesodlum nitriteworking solutionis 1IJgJml.
5.18 Potassium nitratestocksolution(KN03)·
Dry afew grams of potassium nitrate in an oven (6.16) at 110·C to 120·C to constant mass (ie. until the
difference between two successive weighings does not exceed 1mg). Dissolve 1,4689 of the potassium
nitrateinwater ina1000 mlvolumetricflask (6.4).Dilutetothemarkwith water and mix.
5.17 Potassium nitrateworkingsolution.
Prepare the potassium nitrate working solution on the day of use. Transferwith a pipette (6.5), 5ml of the
pctassuan nitrate stock solution (5.16) and 20ml of the buffer solution (5.19) to a 1000 ml volumetric flask
(64) Dilute to the mark .,.,;th water and mix. The nitrate content of the potassium nitrate working solution is
4,50IJglml.
5.18 Ammoniasolution(NH3),(P20=0,91g/ml).
If an ammonia solution of above-mentioned concentration is not available, an equivalent amount of a more
concentrated ammonia solution may be usedin5.19[e.g. 103ml of a35% (massfraction) ammoniasolution
(P20=1,19glml»)
5.19 Buffersolution,pH 9,6to9,7.
Dilute SOml ofhydrochloric acid (5.5) ......th600 ml ofwater in a conicalflask(6.3)and mix.Add 135ml of the
ammonia solution (5.18) and etlute to 1000 ml......th 215ml of water and mix.Adjust the pH, if necessary, to
between 9,6and9.7
6 Apparatus
Clean allg1auwarethoroughlyand rinse withdistilled water to ensurethatitisfree from nitrateandnitrite ions.
Usua/laboratory eqUIpmentand,Inparticular,thefollo......ng
8.1 Analyticalbalance,capable ofweighingtothe nearest1mg,with areadabilityof0,1mg.
8,2 Samplecontainer,providedwith anairtightlid
8.3 Conical"asks,ofcapacity 2SOml,SODmland 1000mi.
8.4 Volumetric flasks,of nominal capacity 100ml,500 ml and 1000 ml, complyingwith the requirements
ofISO 1042.dass B
8.5 Pipettes, capable of delivering 2ml, 4ml, 5ml,6ml, 8ml, 10ml, 12ml, 20 ml and 25ml, complying
with the requirements 01ISO648,dassA, orISO 835-1.Whereappropriate.burettesmaybe usedinsteadd
pipettes
8.8 Measuringcytinders,ofcapecities5ml,10ml,25ml, 100ml,250ml,500mland 1000ml.
8.1 Glasafunnefs,ofetameter7em.'Mthshortstem.
1.8 Filter papef.mediumgrade.ofdiameterabout 15em,free from nitrateandnitrite ions.
8.9 Reductioncolumn,made of~...anexampleofwhidlis!tieninFlOUre 1.

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IS9967 (Part 1):2008
ISO 14673-1 :2004
8.10 Spectrometer,suitabletor measuringabsorbance at awavelength ot538 nm,WIthcells ofopllcal path
length 1cmto2em
8.11 Grinding device,appropnate tor grindingthe test sample,Itnecessary Toavord loss of mcssture the
deviceshould notproduceundue heat Ahammer shall notbeused '
8.12 Laboratory mixer or homogenizer, with glass containers r:A capaoty 250 ml or 400 rnt sUitable for
suspendingtest portionsotcheese and whey cheese,
6.13 Test sieve,ofwoven wire doth.r:A diameter200rnm, nominal SIZeotopenings 500 ~m, and 8 recerver
complymg WIththe requirements ofISO 565.
6.14 Magneticstirrer,
6.15 Water bath. capableotboiling water,
6.16 Oven, capable ofmaintainingatemperatureofbetween 110·C and 120·C
7 Sampling
Sampling is not part of the method specified in thiS part of ISO 1467311DF 189 Arecommended samphng
methodisgiven inISO707,
Itisimportantthat the laboratory receive asample which IStruly represent8tlve and has notbeen damagedor
changed duringtransportorstorage.
Storetheteatsampleinsuch away thatdetenoration andchange incomposmon areprevented
8 Prep.ratlon oftest sample
8.1 Dried milkand driedwhey
Transfer the test sample to a sample container (6.2) ot capacity about IWlcethe volume of the test sample
Close the container immediately, Mix the teat sample thoroughly by repeatedly shakIng and inverting the
containeruntilahomogeneoussampleisobtained
8.2 caseinsandcaseinates
8.2.1 Thoroughlymix the teat sample. if necessaryafter transferring all ofit to a.amplecontalner (62) of
suitableC8p&city. byrepeatedyshakingandinvertingthecontainer
8.2.2 Transfer50gofthe test sampletothetest sieve (613) Itthe 50gportlon passes drect1ythroughthe
sieve,oralmostcompletely, passthe wholemIXedtest sample(821)through the &leve,
8.2.3 If the teat sample does not pass completely through the sieve. uae the gnndmg device (6 i1) to
aenevethat condition.Immediatelytransfer aU the sievedtest sample to the sample contalner (6 2}and mil'
thoroughlyinthe closedcontainer.Duringthese operations,take precautionsto avOIdany change10thewater
content ofthe product.
8.2.4 After the test sample has been prepared. proceed with the preparation r:A the test portion (94) as
soonaspossible.

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