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IS 2557 (1994): Annatto Colour for Food Products [FAD 8:Food Additives]

18 2667 : 1994. ( RNfflrmed 1999 )

~~ ~ ~ 3t ..;=fllT~-~

( q~ffi' ~if"€te11lT )

Indian Standard

ANNATTO COLOUR FOR FOOD PRODUCTS­SPECIFICATION

( First Revision)

Second Reprint SEPTEMBER 200 )

UDC 633-863·92: 637-2/-3

OBIS 1994

BUREAU OF INDIAN STANDARDSMANAK BHAVAN, 9 BAHADUR SHAH ZAFAR MARG

NBW DELHI 110002

Octob" 1994 Price Gro.,1

xxxx2009

AMENDMENT NO. 1 FEBRUARY 2006TO

IS 2557: 1994 ANNATTO COLOUR FOR FOODPRODUCTS - SPECIFICATION

( Fuwt RevLdo,,)

( Page 1,clause4.2.1 ) - Substitutethe following for the existing clause:

,Annatto extract in oil, as solution or suspension, is preparedby extractionof theouter coatingof seeds with vegetable oils. In the preparation of the solution ofannatto colour in oil, only the vegetable oils included under the Prevention ofFood Adulteration Rules, 1955 shall be used, either singly or in a mixture andshall conform to relevant Indian Standards where it exists.'

(FAD 8)

Reprography Unit, 818, New Delhi, India

AMENDMENT NO.2 APRIL 2011TO

IS 2557: 1994 ANNATTO COLOUR FOR FOODPRODUCTS - SPECIFICAnON

(First Revision)

[Page 3. clouse 6.2.l(c») - Substitute 'net fluid quantity' for 'net fluidcontents'.

(Page 4, clause 8-1.1) - Substitute the following for the existing formula:

Percent total carotenoidA 1000000

(expressed as bixin) = -- x -------2.826 Sample mass (mg) x 100

(Page 4, clause B-2.1) - Substitute the following for the existing formula:

Percent total carotenoid

(expressed as norbixin) = _A_ x 1000 0003.473 Sample mass (mg) x 100

(FAD 8)Reprography Unit. SIS, New Delhi, India

Food Additives Sectional Committee, FAD 8

FOREWORD

This Indian Standard ( First Revision) was adopted by the Bureau of Indian Standards, after the draftfinalized by the Food Additives Sectional Committee had been approved by the Food and AgricultureDivision Council.

Annatto colour is obtained from the aril <coating of the seed) commonly known in commerce as theseed of the plant Bixa orellana L. Its solution in oil is used for colouring butter and other food productswhile its solution in water with a little alkali is used for colouring cheese and other similar products.

This standard is one of a series of Indian Standards on natural food colours permitted under thePrevention of Food Adulteration Rsles, 1955 with the latest amendments, issued by the Ministry ofHealth, Government of India. In the preparation of this standard due consideration has been given tothese Rules. Due consideration has also been given to the Standards 0/ Weights and Measures ( PackagedCommodities) Rules, 1977. However, this standard is subject to the restriction imposed under theseRules wherever applicable.

This standard was first published In 1963. It has been revised taking into account the latest technolo­gical developments in the country and also to bring it in line with the International specifications. In thisrevision minimum limits for carotenoids and maximum limits for heavy metals have been included aswell as identification tests for annatto.

In the preparation of this standard, assistance has been derived from the following publications.Specification for Identity and Purity of Food Additives, Food and Nutrition Paper 19, 'publishedby Food and Agriculture Organization of the United Nations and the World Health Organization.Food Chemical Codex published by the National Academy of Sciences and National ResearchCouncil, Washington DC, USA. Second supplement o~ 1981 Edition.

DescrlptloDClass - CarotenoidsCode Number - CI ( 1975 ) No. 75120

CI ( 1975 ) Natural Orange 4EEe No. £.160 b

Chemical Name

Annatto extract in oil contains several coloured components, the major single one being bfxin, whichmay be present in both Cis and trans forms. Thermal degradation products of bixin may also bepresent.

Water soluble annatto contains norbixin, the hydrolysis product of bixin, in the form of sodium orpotassium salt, as the major colouring principle. Both Cu and trans forms may be present.

Chemical Formula - Bixin C.H,.O,Norbixin c..H1sO.

Structural Formula

CIS AND TRANS BIXINSHAVE THE STRUCTURES

CIS AND TRANS· NORBIXINS(Na 00 K SALIS) HAVE THESTRUCTURES

where R - Na or K( Co",,,,,,,d 011 t"',d cover)

( ContinuedIron! second cover)

Molecular Weight - BixinNorbixin

394·50380-48

For the purpose of deciding whether a particular requirement of this standard is complied with,final value, observed or calculated, expressing the result of a test or analysis, shall be rounded oftaccordance with IS 2: 1960 'Rules for rounding off numerical values (revised)'. The numbersignificant places retained in the rounded off value should be the same as that of the specified valuethis standard.

IS '1.557 : 1994

Indian Standard

ANNATfOCOLOURFORFOODPRODUCTS-­SPECIFICATION( First Revision)

0.320 g2.02g

2011To make solutionto one litre

These ~agenlssball be oftbe analytical reagent grade(see IS 2SO : 1964 and IS 266 : 1993 ). Althougb tbesolution retains its tinctorial value for a considerabletime, after prolonged storage, its optical clarity sha!l heexamined before use, to ensure that no alteration hastaken place.

NO'rESI Diluted solution ofannatto colour in amyl acetate is notstable in colour quality particularly if exposed to light,and measurement should be carried out on the dilutedsolution without unduedelay.2 In using the lintometer, the maker's instrudions shallbe rigidly followed.

4.3 Solution of Annatto Colour In Water for Use inCheese and Other Food Products

4.3.1 Water soluble annatto is prepared by extraction oftbe outer coating of tile seeds with aqueous alkali( sodium or potassium bydroxide ). In tbe preparationof tbe solution, potable water shall be used. A littlequantity (0.5 to 3 percent) of alkali may be added(see IS 376 : 1986 and IS 296 : 1986 ).

4.J-Z The solution shall be clear and shall remain so onstorage in suitable containers at a temperature of 15°C.

4.2.2 The solution of annatto colour in oils shill beclear and shall remain so on storage in suitableconlainel5at 1SOC except fora sligbldepositofstearineor sball be in tbe form of. suspension. The suspe_ionon dilution witb bot oil to bring tbe bixin content to 0.24percent sbaII bea clear solution.

4.1.3 ColourThe colour of solution in amyl acetate ( see IS 231 :1957) at a dilution of 1 : 1000 (wlv), when measuredin a Lovibond TinlODtelerwitb I l-COI cell, sball be notless than tbe following:

Yellow units 5.0Red units 0.4

or be not less than the rolour of tbe following ioorganicsolution at a liquid deptb ofone centimetre whicb D18Y

be employed for matching tbe stated dilution in aplunger type colorimeter using incident ligbt closelyapproxhuating the north day light

Potassium bicbromate

Cobalt ammoniumsulphate(CoS04, (NH4}2 S04, 6H20 )Sulphuric acid sp-gr 1.84

Disti lied water

1 SCOPE

1.1 This slandlrd prescribes the requirements and themethods of test for annatto colour derived from tbeplant Biro orellana L. and supplied in tbe form of<a> solution in oil, and (b) solution in water, for use inbutter, cheese and other food products.

Z REFERENCES

The Indian Standards listed at Annex A are necessaryadjuncts to this standard.

3 TYPES

3.1 The n18 teria I shall be of tbe following two types:

a) Solution in oil for use in butter and otber foodproducts, and

b) Solution in water for usc in cheese and otherfood products.

4 REQIJlREMENTS

4.1 General

The material shall be derived only from tbe plant Bixaorellana L. and shall not contain any extraneous colour­ing matter, It shall be processed, packed, stored anddistributed under hygienic conditions in licencedpremises ( see IS 2491 : 1972).

4.2 Solution of Annatto Colour In Oil for Use inButter and Other Food Products

4.%.1 Annatto extract in oil, as solution or suspension,is prepared by extraction of tbe outer coating of seedswitb vegetable oils. In tbe preparation of the solutionofannatto colour in oil, only the vegetable oils includedunder the Prevention cfFoodAduiteration Rilles, 1955shall be used, either singly or in a mixture and shallconform to tbe following specifications:

8) Coconut oil - conforming to IS 542: 1968b) Groundnut oil - conforming to IS 544 : 1968c) Mustard oil - conformieg to IS S46 : 1975d) Sesame oil - conforming to IS 547 : 1968e) Linseed oil - confonning to IS 75 : 1973t) Nigerseed oil -l.'"Onfomting to IS 3490: 1965g) Samower seed oil - confooning to IS 3491 :

lQ(i5b) Olive oil - conforming to specifications given

under Indian Pharn18copoeiaj) Poppy seed oil - ronfonniag to specifiC'l lions

given under Prevention of Food AdulterationRilles, 1955.

182557: 19M

4.3.3 Colour the column. 1be Ol'lnge-ral zone (bixin) It the top ofTIle colour or tbe solution in D.l N sodium hydroxide the colUnlB immediltely becomes blue-green.or polUSium bydroxide II I dilution 1 : 1 000 (",'v) 4A.2.3 Woter $01,""e annattomeuuR:d iD I l-an cell sull be lbe SiDle Is""l Tl'IlISfer2 ml or 2 g to I SD-mlscpll'ltory funnellndspecified iD4.2.3. . ldel sufficient 2 N sulpburic lcid to make tbe solution4.4 IcIeIItilkatioa Icidic to pH test paper (pH 1-2). Dissolve the ReI

precipitate of norbixin by mixiog tile solution with SO4.4.1 Spectropholometry mI of benzene. Discard tbe water layer and Wlsb lbeAaaauo extrKll in oil, diluted with chloroformexibits benzene pbase with Wlter until it no longer gives ID.bIorbince xi.. II 439, 470 .nd SOl RID. Water acid IUction. Remove Iny undissolved norbixin bysoluble tlO extlllda, diluted with water exibit ab- centrifugation or filtmlion and dry tbe solution overlO....ncc maxiDII at 453 aDd482 1lDl. anhydrous sodium sulphate. Transfer 3-5 011 of tbe dry4A.2 Col"." CluomtJlOgraphy antiCa"-Prke solution to tbe top of ID IlunU'" columa pre..re ISRetJClitM descnbed above. FJute thecolumn witb beozeae,three4.4.1.1 Preflllml;OIf 0/01,"";l1licolUIIUI ID-m) volumes ofdry cblorofoml, followed by Sml of

r" Carr-Price ragent added to the top ortbe column. TbePrep.re I smlll cbmlllltoppby COIuIlUl by tilting I omoge-red bind of norbixin immedialely becomesgillS tube (7 X 200 Ram mlerMl diameter), stoppered blue-green.willi gillS wool, with llumi. ( 800 - 200 mesh) .... ~ ..... L ell ,nIslurried iabeDZeDesolblttllesettledlJulllina fiUsabout ~ I."'" ayer "romalDgrtlyr'Ytwo-tbircls ortbe tube. Fill tile column witb benzene I lid 4.4.3.1 A",",,,o extract in oilIdjusl tile flow rite to lboul 30 drops/min using a Use silica gel with 12 percent calcium sulphite IS

rubber outlet tube ~nd c"mp until the bellZene llyer is binding .genl,.nd a mixture ofacetic I,'-id,chloroform2-3 mmlbove tbe .Iuminl SUrfltt. and acetone (1 : 50 : SO, by volume) IS developing

NOTE- It i» advisable to place a glass-wool plug at the solvent. 1be sample in oil produces .t Ie.st 3 or 4 redtop ortbe IlumiDl to prnent cbumio. when solvent is or yellow spots. 1\vo red spots Ire clearly olO~ inte.-eIdded,and to applya reservoir orsolvent at the top of the tbln tbe otller spots. Dissolve the two spols in benzenecolumn, which will exert a mild pralle on the and identify the spot lhat is bixin by tbe method givenchromatoppbic ooIumn. The sudioa m..t be c.efully in 4.4.2.2.oonb'olled 10 prnent channelling in the alumina. Avoid 4.4.3.% Water soillble annattoelutiog tbe benzene or otber solvent completely. It isneaaaarylbatthesurfaceofthealuminaisbptcovered Prepare a cbroRlIlogram as indicaled under 4.4.3.1.with liquid layer. Three or four spots Ire obtained,!Jfwbicb 2 spots bave

4A.2.% Annattoextract in oil an ol'lnge colour and tbe others an orange-yellowcolour. Dissolvetbe omngespots in benzene aDd ideo­

Add to tbe top oftbe aluOOIll coJulnn3 mJ ofa solution tifyone oftbe spots that is norbixin by tbe Dletbod givencontlining suffICient saRlple, in benzene, to impart I in 4.4.1.3.colour equivalent to a O.I% potlssiuDI dichromate solu-lion; elute with benzene until. pale yellow fmeliOn is 5 PIJRITY TESTSwashed frolll the coluam, W.sb the column witb three S.t TIle Illalerill shall conform to tbe requirements10-ml ~olumes of dry chlorof~nn; tben add S ml of given in Table 1.Carr-Pnce ~geDllnd allow It to nan onto the top of

Table 1 Requirelllents for An.aUo

( Clause5.1 )

SINo. Requirement Method or -rest. Ker tn

(1)i)

ii)iii)iv)

v)

(2)(~rotenoids

a) Annatto extract in oil,expresaed as bixin, percentby mau.Mill

b) Water-soluhle Innal~o.

expreued 8. norbixin,perceot by mui, M;"

Arsenic, milk., MaxLead, m~g, MtlXC.oppermglkg, MuHeavy metals, mllkg, MfIX

(3)

0.24

0.24

310

30._

40

2

Annex of ThisSiandard

(4)

8

B

Ref 10 nause ofIS 16" : lQQ4

(S)

ISISIS16

, PACKING, MARKING AND STORAGt;

'.1 Packing

The mlerial shall be tilled in suitable containers witbIS little air space as pos.~iblc so IS 10 preclude con­'anunalion of tbe contents with metals or otber im­puriljes~ and sbould preferably exclude ligbt.

6.z Marldlll

6.2.1 Eacb contlincr shall be marked legibly andindelibly to give tbe following infomlltion:

a) NIDle and type of the material;b) Sourceof manufacture;c) Mininlunl net fluid rootents;d) Ball-hor code number;e) Dale of Inanufacture;t) Declaration of bixin and Ilorbixin conteut;g) A s,alclnenl to the effect lhaltbc n1BleriB) should

he kept in I (,"001 ptace, and in tbe case of glasscontainers, away fro In light; and

b) Any otber de~aik as specifled under tbe Stand­ards o/Weights and Measures( PllckagedCom­modities ) Rilles, 1977/Preleention of FoodAdulterotion Rilles, 1955.

6.z.z SIS Certification M(,rkin~

The product may also be marked witb tbe StandardMark.

IS 1557 : 1"4

6.1.1.1 The use of tbe Standard Mark is governed bythe provisions of the Bureau of Indian Standards Act,1986 and the Rules and Regulations made thereunder.The details of conditions under whi("b the licence fortbe use of Standard Mark loay be glinted to manufac­turers or produttl'S .nay be obtained from the Bureauof Indian Standards.

6.3 Storage

The nllterial shall be stored in a cool place.

7 SAMPLING

7.1 Representative samples oflhe Inaterial for tbe tesL~

shall be drawn according to tbe oletbod prescribed in 4of IS 1699 : 1994.

8 'rlt:Sl"S

8.1 Tests shall be carried out as prescribed in col 4and 5 of Table 1.

1.2 Quality of Reagents

Unless specified otherwise, pure ("bcn,icals anddistilled waler ( see IS 1070 : 1992 ) shall be empjoyedin tests.

NOTE - •Pure chemicals" shall mean chemicals that donot contain impurities which affed the experimentalresults.

ANNEXA(Clause 2)

IJIST OF REFERRED INDIAN STANDARDS

IS No.

7S: 1973

231 : 1957

250: 1964

266: 1993

296: 1986

376: 1986

542: 1968

S44: 1968

Tille

Linseed oil, raw and refined(second revision)Amyl acetate

Potassium bichromate, lechnicaland analyti("al reagent (revised)Sulpburic 8~id ('!lird revision)Sodium carbonate. anbydrous( third revision )Sodiunl bydroxide, analyticalreagent (tllird revision )Coconut oil ( second revision)

Groundnut oil (second revision)

3

ISNo.

546: 1975

547: 1968

1070: 1992

1699: 1994

2491 : 1972

3490: 1965

3491 : 1965

ntle

Muslard oil (second revision )

SeasaOle lli I ( second revision )

Reagent grade water ( thirdrevision )

Methods of 58lupling and lest forfood colours ( second revision )Code for hygienic conditions forfood processing units ( firstrevision )Nigerseed oil

SlOlower oil

IS ZSS7 : 1994

ANNEXB"[ Table 1, Item (i) ]

DETERMINAnON OF CAROTENOIDS

Percent totalcarotenoid

B-1 ANNAITO EXTRACT IN OIL

B-I.l Transfer 0.1 to 1 g of tbe SImple, acculltelyweighed, into a lOO-ml, volametric Dask. Dissolve inchloroform, dilute to volume witb chloroform. and mix.Transfer a J-ml portion of the solution into anotberl00-lul volunlelric nask, and dilute to 100 ml,Measure the Ibsorhanc.<te A of this solution at the wave­lengtb of 470 nm,

A 100 ()()().---x--------2.826 Sanlple miss (Dig) X tOO

(expressed as hixin)

B-Z WATER·S()I~IJ8LE ANNATIO

solution (1 in 10) and wlter to SO ml, and add 2 ml ofdiluled sulphuric acid. Continue '0 exillcilbe solulionwith each 10 011ofbenzene until tbe benzene extract isnot coloured. Combine benzene extract, Wlsb threethues witb each S nJI of water, and allow to stand toseparate the Wiler layer. Transfer the benzene extractinto another separating runnel containing tbe Wilerlayer three times with 2 ml of benzene, 100 combieetbe washings 10 tbe benzene extract. To the benzeneextract, add an equal volume of petroleum benzenemix, and continue to extract with each S 011 of 0.01 Nsodium hydroxide until sodium hydroxide solution isnot coloured. Combine extracts, and add 0.01 sodiumhydroxide to 100 ml. Measure tbe absorbace A at thewavelength of 453 nm,

8-%.1 Weigb accurately 0.1 to 1 g of tbe sample, add0.01 N sodium bydroxide to 100 ml, and shakethorougbly. Transfer I ml portion of the solutioninto a separating funnel, add 10 nil of sodium chloride

4

Percent totalcarotenoid

(expressed asnorbixin)

A 100000.--x---------3.473 Sample 1118SS (mg) X 100

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This Indian Standard has been developed from Doc: No. FAD X ( 256 ).

Amendments Issued Since Punhcatinn

Amend No. Date of Issue

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