JECFA Monograph 22

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COMPENDIUM
OF FOOD ADDITIVE FAO JECFA Monographs 22
SPECIFICATIONS
84th Meeting 2017
(JECFA), which was held in
M
COMPENDIUM
Y
CM
OF FOOD ADDITIVE
MY
CY
SPECIFICATIONS
CMY
Joint FAO/WHO Expert Committee on Food Additives
86th Meeting 2018
FAO / WHO
FAO JECFA Monographs 22
COMPENDIUM
OF FOOD ADDITIVE
SPECIFICATIONS
86th Meeting
Geneva, 12 – 21 June 2018
Food and Agriculture Organization of the United Nations

World Health Organization
Geneva, 2018
II
Required citation:
FAO and WHO. 2018. Compendium of Food Additive Specifications. Joint FAO/WHO Expert Committee on Food
Additives (JECFA), 86th Meeting June 2018. FAO JECFA Monographs 22. Rome. 167 pp.
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FAO JECFA Monographs 22 III
SPECIAL NOTE
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IV FAO JECFA Monographs 22
FAO JECFA Monographs 22 V
Contents
LIST OF PARTICIPANTS ..........................................................................................................VII

INTRODUCTION ........................................................................................................................IX
SPECIFICATIONS FOR CERTAIN FOOD ADDITIVES ............................................................... 1
ANIONIC METHACRYLATE COPOLYMER............................................................................. 3
BASIC METHACRYLATE COPOLYMER ................................................................................. 9
CASSIA GUM ........................................................................................................................ 15
CITRIC AND FATTY ACID ESTERS OF GLYCEROL (TENTATIVE) ..................................... 23
ERYTHROSINE ..................................................................................................................... 27
GLYCEROL ESTER OF WOOD ROSIN ................................................................................ 35
INDIGOTINE .......................................................................................................................... 41
LUTEIN FROM TAGETES ERECTA ...................................................................................... 47
MODIFIED STARCHES ......................................................................................................... 53
NEUTRAL METHACRYLATE COPOLYMER (TENTATIVE) .................................................. 87
SPIRULINA EXTRACT (TENTATIVE).................................................................................... 93
SPECIFICATIONS FOR CERTAIN FLAVOURING AGENTS .................................................... 99
GROUP 1: MISCELLANEOUS NITROGEN-CONTAINING SUBSTANCES ......................... 100
GROUP 2: SATURATED ALIPHATIC ACYCLIC BRANCHED-CHAIN ................................. 102
PRIMARY ALCOHOLS, ALDEHYDES, AND ACIDS ............................................................ 102
GROUP 3: LINEAR AND BRANCHED-CHAIN ALIPHATIC, UNSATURATED,
UNCONJUGATED ALCOHOLS, ALDEHYDES, ACIDS, AND RELATED ESTERS ............. 103
GROUP 4: CARVONE AND STRUCTURALLY RELATED SUBSTANCES ......................... 104
GROUP 5: MENTHOL AND STRUCTURALLY RELATED SUBSTANCES .......................... 106
GROUP 6: MALTOL AND RELATED SUBSTANCES .......................................................... 110
GROUP 7: ALICYCLIC PRIMARY ALCOHOLS, ALDEHYDES, ACIDS ............................... 111
AND RELATED ESTERS..................................................................................................... 111
GROUP 8: FURAN SUBSTITUTED ALIPHATIC HYDROCARBONS, ALCOHOLS,
ALDEHYDES, KETONES, CARBOXYLIC ACIDS AND RELATED ESTERS, ...................... 113
SULFIDES, DISULFIDES AND ETHERS ............................................................................. 113
REVISIONS TO EXISTING FLAVOUR SPECIFICATIONS .................................................. 126
SPECTRA OF CERTAIN FLAVOURING AGENTS .............................................................. 127
CORRIGENDUM ..................................................................................................................... 137
ANNEX I: SUMMARY OF RECOMMENDATIONS FROM THE 86th JECFA ............................ 141
ANNEX 2. GENERAL INFORMATION .................................................................................... 153
ANNEX 3. FUTURE WORK AND RECOMMENDATIONS....................................................... 155
SPECIFIC FOOD ADDITIVES (OTHER THAN FLAVOURING AGENTS) ............................ 155
FLAVOURING AGENTS ...................................................................................................... 157
VI FAO JECFA Monographs 22
FAO JECFA Monographs 22 VII
LIST OF PARTICIPANTS
JOINT FAO/WHO EXPERT COMMITTEE ON FOOD ADDITIVES, 86th MEETING

Geneva, 12 – 21 June 2018
Members
Dr S. Barlow, Brighton, East Sussex, England, United Kingdom
Dr J. Bend, Department of Pathology and Laboratory Medicine, Schulich Medicine & Dentistry,
Western University, London, Ontario, Canada
Dr D. Benford, Cheddington, London, England, United Kingdom
Dr R. Cantrill, Halifax, Nova Scotia, Canada (Vice-Chairperson)
Dr E. Dessipri, General Chemical State Laboratory, Athens, Greece, currently European
Directorate for the Quality of Medicines and Health Care, Strasbourg, France
Dr D. Folmer, Office of Food Additive Safety, Center for Food Safety and Applied Nutrition, United
States Food and Drug Administration, College Park, Maryland, USA (Joint Rapporteur)
Ms T. Hambridge, Food Standards Australia New Zealand (FSANZ), Kingston, ACT, Australia
Dr A. Mattia, Office of Food Additive Safety, Center for Food Safety and Applied Nutrition, United
States Food and Drug Administration, College Park, Maryland, USA (Chairperson)
Dr U. Mueller, Australian Pesticides and Veterinary Medicines Authority (APVMA), Symonston,
Australian Capital Territory (ACT), Australia (Joint Rapporteur)
Dr O.E. Orisakwe, University of Port Harcourt, Choba, Port Harcourt, Rivers State, Nigeria
Dr J. Schlatter, Zurich, Switzerland
Dr J. Smith, Bio|Food|Tech, Charlottetown, Prince Edward Island, Canada
Dr J.R. Srinivasan, Division of Biotech and GRAS Notice Review, Office of Food Additive Safety,
Center for Food Safety and Applied Nutrition, United States Food and Drug Administration,
College Park, Maryland, USA
Dr M. Veerabhadra Rao, Hyderabad, India
Secretariat
Dr J.H. Andersen, National Food Institute, Technical University of Denmark, Lyngby, Denmark
(FAO Expert)
Dr J.N. Barrows, Office of Cosmetics and Colors, Center for Food Safety and Applied Nutrition,
United States Food and Drug Administration, College Park, Maryland, USA (FAO Expert)
Dr C. Béchaux, ANSES, Maisons-Alfort, France (WHO Temporary Adviser)
Dr M. DiNovi, Office of Food Additive Safety, Center for Food Safety and Applied Nutrition, United
States Food and Drug Administration, College Park, Maryland, USA (WHO Temporary
Adviser)
Dr B. Fallico, Food Science and Technology Unit, University of Catania, Catania, Italy (FAO
Expert)
VIII FAO JECFA Monographs 22
Mr Y. Fan, China National Center for Food Safety Risk Assessment, Beijing, China (CCFA
Chairperson)
Dr M.J.F. Fernandez, Universidad Miguel Hernández, Alicante, Spain (FAO Expert)
Dr B. Fields, Food Standards Australia New Zealand, Barton, Australian Capital Territory (ACT),
Australia (WHO Temporary Adviser)
Ms F. Hill, Food Standards Agency, London, United Kingdom (WHO Temporary Adviser)
Dr S.M.F. Jeurissen, Centre for Nutrition, Prevention and Health Services, National Institute for
Public Health and the Environment (RIVM), Bilthoven, the Netherlands (WHO Temporary
Adviser)
Dr X. Jia, Laboratory of Toxicology, China National Center for Food Safety Risk Assessment,
Beijing, China (WHO Temporary Adviser)
Dr S. Kim, Department of Food Safety and Zoonoses, World Health Organization, Geneva,
Switzerland (WHO Secretariat)
Ms K. Laurvick, United States Pharmacopeial Convention, Rockville, Maryland, USA (FAO
Expert)
Dr M. Lipp, Agriculture and Consumer Protection Department, Food and Agriculture Organization
of the United Nations, Rome, Italy (FAO Joint Secretary)
Dr P. Mosesso, Department of Ecological and Biological Sciences, Università degli Studi della
Tuscia, Viterbo, Italy (WHO Temporary Adviser)
Ms J. Odrowaz, Toronto, Ontario, Canada (WHO Technical Editor)
Mr K. Petersen, Department of Food Safety and Zoonoses, World Health Organization, Geneva,
Switzerland (WHO Secretariat)
Dr L. Rosenfeld, Division of Petition Review, Office of Food Additive Safety, Center for Food
Safety and Applied Nutrition, United States Food and Drug Administration, College Park,
Maryland, USA (WHO Temporary Adviser)
Dr J. Rotstein, Pre-Market Toxicology Assessment Section, Chemical Health Hazard
Assessment Division, Bureau Chemical Safety, Food Directorate, Health Products and Food
Branch, Health Canada, Ottawa, Ontario, Canada (WHO Temporary Adviser)
Dr N. Sugimoto, Division of Food Additives, National Institute of Health Sciences (NIHS),
Kanagawa, Japan (FAO Expert)
Dr S. Takasu, Division of Pathology, Biological Safety Research Center, National Institute of
Health Sciences, Kawasaki-shi, Kanagawa, Japan (WHO Temporary Adviser)
Dr A. Tritscher, Department of Food Safety and Zoonoses, World Health Organization, Geneva,
Switzerland (WHO Joint Secretary)
Dr T. Umemura, Faculty of Animal Science Technology, Yamazaki Gakuen University, Tokyo,
Japan (WHO Temporary Adviser)
Dr X. Yang, Food Safety and Health Research Center, Southern Medical University, Guangzhou,
Guangdong Province, China (WHO Temporary Adviser)
Ms L. Zhang, Joint FAO/WHO Food Standards Programme, Food and Agriculture Organization
of the United Nations, Rome, Italy (Codex Secretariat)
FAO JECFA Monographs 22 IX
INTRODUCTION
This volume of FAO JECFA Monographs contains specifications of identity and purity prepared at
the 86th meeting of the Joint FAO/WHO Expert Committee on Food Additives (JECFA), held in
Geneva, 12 – 21 June 2018. The specifications monographs are one of the outputs of JECFA’s risk
assessment of food additives, and should be read in conjunction with the safety evaluation, reference to
which is made in the section at the head of each specifications monograph. Further information on the
meeting discussions can be found in the summary report of the meeting (see Annex 1), and in the full
report which will be published in the WHO Technical Report series. Toxicological monographs of the
substances considered at the meeting will be published in the WHO Food Additive Series.
Specifications monographs prepared by JECFA up to the 65th meeting, other than specifications for
flavouring agents, have been published in consolidated form in the Combined Compendium of Food
Additive Specifications which is the first publication in the series FAO JECFA Monographs. This
publication consists of four volumes, the first three of which contain the specifications monographs on
the identity and purity of the food additives and the fourth volume contains the analytical methods, test
procedures and laboratory solutions required and referenced in the specifications monographs. FAO
maintains an on-line searchable database of all JECFA specifications monographs from the FAO
JECFA Monographs, which is available at: http://www.fao.org/food/food-safety-quality/scientific-
advice/jecfa/jecfa-additives/en/. The specifications for flavourings evaluated by JECFA, and previously
published in FAO Food and Nutrition Paper 52 and subsequent Addenda, are included in a database for
flavourings specifications. All specifications for flavourings that have been evaluated by JECFA since
its 44th meeting, including the 79th meeting, are available in the online searchable database at the JECFA
website at FAO: http://www.fao.org/food/food-safety-quality/scientific-advice/jecfa/jecfa-flav/en/.
The databases have query pages and background information in English, French, Spanish, Arabic and
Chinese. Information about analytical methods referred to in the specifications is available in the
Combined Compendium of Food Additive Specifications (Volume 4), which can be accessed from the
query pages.
An account of the purpose and function of specifications of identity and purity, the role of JECFA
specifications in the Codex system, the link between specifications and methods of analysis, and the
format of specifications, are set out in the Introduction to the Combined Compendium, which is
available in shortened format online on the query page, which could be consulted for further information
on the role of specifications in the risk assessment of additives.
Chemical and Technical Assessments (CTAs) for some of the food additives have been prepared as
background documentation for the meeting. These documents are available online at:
http://www.fao.org/food/food-safety-quality/scientific-advice/jecfa/technical-assessments/en/.
Contact and Feedback
More information on the work of the Committee is available from the FAO homepage of JECFA at:
http://www.fao.org/food/food-safety-quality/scientific-advice/jecfa/en/. Readers are invited to address
comments and questions on this publication and other topics related to the work of JECFA to:
jecfa@fao.org
X FAO JECFA Monographs 22
FAO JECFA Monographs 22 1
SPECIFICATIONS FOR CERTAIN FOOD ADDITIVES
New and revised specifications
New (N) or revised (R) specifications monographs were prepared for 11 food additives and
these are presented in this publication.
Anionic methacrylate copolymer (AMC) (N, T)

Basic methacrylate copolymer (BMC) (N)
Cassia gum (R)
Citric and fatty acid esters of glycerol (R, T)
Erythrosine (R)
Glycerol ester of wood rosin (R)
Indigotine (R)
Lutein (R)
Modified starches 1 (R, T)
Neutral methacrylate copolymer (NMC) (N, T)
Spirulina extract (N)
In the specifications monographs that have been assigned a tentative status (T), there is
information on the outstanding data and a timeline by which this information should be
submitted to the FAO JECFA Secretariat.
1Applying to all 16 modified starches: INS 1400, 1401, 1402, 1403, 1404, 1405, 1410, 1412, 1413,
1414, 1420, 1422, 1440, 1442, 1450, 1451
2 FAO JECFA Monographs 22
ANIONIC METHACRYLATE COPOLYMER
New specifications prepared at the 86th JECFA (2018) and

published in FAO JECFA Monographs 22 (2018). No ADI was
established at the 86th JECFA (2018).
SYNONYMS E 1207, INS No. 1207, acrylates copolymers, Methyl acrylate,

methyl methacrylate, methacrylic acid polymer; methacrylic acid,
polymer with methyl acrylate and methyl methacrylate
DEFINITION Anionic methacrylate copolymer is a copolymer comprised of

monomers, methyl acrylate, methyl methacrylate, and
methacrylic acid in the molar ratio of 7:3:1. The copolymer is
manufactured by emulsion polymerization of the monomers with
water soluble radical initiators. The product is purified by water
vapour distillation and filtration to remove residual monomers,
excess water, other volatile low-molecular weight substances
and coagulum. The copolymer is standardized as a 30%
aqueous dispersion. The copolymer dispersion may contain
residual monomers (methyl acrylate, methyl methacrylate, and
methacrylic acid). Anionic methacrylate copolymer is used as a
coating and glazing agent
for food supplements and products for special medical purposes.
Chemical name Poly (methyl acrylate-co-methylmethacrylate-co-methacrylic

acid) 7:3:1
C.A.S. number 26936-24-3
Chemical formula Poly[(CH2:CHCO2CH3)-co-(CH2:C(CH3)CO2CH3)-co-

(CH2:C(CH3)COOH)]
Structural formula
••• •••
The above formula is provided for illustrative purposes; in this

copolymer no definitive structural unit can be defined.
Formula weight 280,000 (weight-average ), 77,000 (number-average)
Assay 9.2 – 12.3 % methacrylic acid units on the dried basis

See description under Tests
DESCRIPTION Commercial form (30% aqueous dispersion) is a low viscosity,

milky-white liquid.
FUNCTIONAL USES Coating agent, glazing agent.
CHARACTERISTICS
IDENTIFICATION
Viscosity (Vol. 4) Not more than 20 mPa•s
Determine viscosity using Brookfield viscometer at 20° and 30 rpm

using UL adapter.
pH (Vol 4) 2.0 – 3.5
Infrared absorption The infrared absorption spectrum of a dry film of sample

(Vol. 4) corresponds to the infrared spectrum in the Appendix.
Apply one drop of sample to a glass plate, cover with a water-

resistant crystal disc (AgCl, KRS 5), press lightly, remove the
crystal disc and dry for about 15 minutes at 60°.
PURITY
Loss on drying 68.5 – 71.5% (110°, 5 h)

(Vol. 4)
Sulfated ash (Vol. 4) Not more than 0.2%
Test 5 g of the sample (Method I)
Methanol (Vol. 4) Not more than 1,000 mg/kg
Residual monomers Methyl acrylate: Not more than 1 mg/kg
Methyl methacrylate: Not more than 3 mg/kg
Methacrylic acid: Not more than 1 mg/kg
See description under TESTS
Lead (Vol. 4) Not more than 1.0 mg/kg in the dispersion

Determine using a method appropriate to the specified level.

The selection of sample size and method of sample
preparation may be based on principles of methods described
in Volume 4 (under “General Methods, Metallic Impurities”).
Microbiological criteria Total plate count: Less than 1,000 cfu/g

(Vol. 4)
Yeast and moulds: Less than 100 cfu/g
Coliforms: Negative in 10 g
TESTS
IDENTIFICATION
TESTS
Residual monomers Determined by liquid chromatography (Vol. 4)
Standards and Reagents:

• Acetonitrile: HPLC grade with UV absorption: Amax of 1%
at 190 nm
• Acetone, methanol, isobutanol and deionized water:
HPLC grade
• Phosphoric acid solution (pH 2): Adjust phosphoric acid
(85 %) with an appropriate volume of deionized water to
pH 2.
• Standards: methyl acrylate, methyl methacrylate and
methacrylic acid (>99%)
Preparation of mixed standard solutions:

Stock mixed standard solution:
Pipette 5 ml of isobutanol into a 50 ml volumetric flask.
Accurately weigh approximately 10 mg of methyl acrylate,
12 mg of methyl methacrylate and 11 mg of methacrylic
acid, add to isobutanol and dilute to volume with acetone.
Intermediate mixed standard solution-1:

Dilute 5.0 ml of stock mixed standard solution to 50 ml
with acetone in a volumetric flask.
Intermediate mixed standard solution-2: Dilute 20.0 ml of

intermediate mixed standard solution-1 to 50 ml with
acetone in a volumetric flask.
Working mixed standard solution:

Dilute 5 ml of Intermediate mixed standard solution-2 to
25 ml with methanol:phosphoric acid-pH 2 (70:30) in a
volumetric flask.
Preparation of sample solution:

Accurately weigh approximately 11 g of sample, dissolve in
acetone and dilute to 50 ml in a volumetric flask. Add 5.0 ml of
the solution dropwise (precipitation of the polymer should be

slow to avoid entrapment of monomer in the precipitate) to 20 ml
methanol and phosphoric acid- pH 2 (70:30 v/v). Centrifuge until
the supernatant is clear and use the supernatant as the sample
solution.
Procedure:
• Use an HPLC with diode array or UV detector at 200 nm
• Column: Octadecylsilane chemically bonded to porous
silica (125 cm x 4.6 mm i.d.x 7 µm)
• Injection volume: 20 μl
• Mobile phase: Acetonitrile:Phosphoric acid-pH 2 (10:90
v/v)
• Flow rate: 2 ml/min
Inject separately 20 µl each of working mixed standard solution

and sample solution. Calculate the amount of each monomer in
the sample from the peak areas obtained in the chromatograms
of working mixed standard solution (rR) and sample solution (rS);
amount of standard (R, mg), weight of sample (W, g) and dilution
factor (40).
𝑟𝑟𝑟𝑟 × 𝑅𝑅 × 40
𝐶𝐶𝐶𝐶𝐶𝐶𝐶𝐶. 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 [𝜇𝜇𝜇𝜇/𝑔𝑔] =
𝑟𝑟𝑟𝑟 × 𝑊𝑊
Total monomers in the sample (µg/g) = Sum of monomers in the

sample and correct the results for recovery.
METHOD OF ASSAY Accurately weigh about 5 g sample and dissolve completely in

90 ml isopropyl alcohol and 10 ml water. Titrate with 0.5 N
sodium hydroxide standard solution to a potentiometric endpoint.
Perform a blank titration under the same conditions. One ml
0.5 N NaOH corresponds to 43.045 mg methacrylic acid units.
𝑀𝑀𝑀𝑀𝑀𝑀ℎ𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢 (%𝑤𝑤/𝑤𝑤, 𝑜𝑜𝑜𝑜 𝑡𝑡ℎ𝑒𝑒 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏) =

𝑚𝑚𝑚𝑚 𝑜𝑜𝑜𝑜 0.5 𝑁𝑁 𝑁𝑁𝑁𝑁𝑁𝑁𝑁𝑁 × 430.45
𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤ℎ𝑡𝑡 (𝑔𝑔)× % 𝑑𝑑𝑑𝑑𝑑𝑑 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 𝑖𝑖𝑖𝑖 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠
Appendix: Infrared spectrum of anionic methacrylate copolymer

BASIC METHACRYLATE COPOLYMER
New specifications prepared at the 86th JECFA (2018) and

published in FAO JECFA Monographs 22 (2018). An ADI of “not
specified” was established at 86th JECFA (2018).
SYNONYMS E 1205; INS No. 1205; basic butylated methacrylate copolymer;

amino methacrylate copolymer; aminoalkyl methacrylate
copolymer E; butyl methacrylate, dimethylaminoethyl
methacrylate, methyl methacrylate polymer;
butyl methacrylate, methyl methacrylate, dimethylaminoethyl
methacrylate copolymer
DEFINITION Basic Methacrylate Copolymer is a cationic copolymer comprised

of the monomers dimethylaminoethyl methacrylate, butyl
methacrylate and methyl methacrylate in the molar ratio of 1:2:1.
The copolymer is manufactured by a controlled polymerization
process using a free radical donor initiation system. After
completion of polymerization, the viscous copolymer solution is fed
into an extruder to remove solvents and volatile substances, by
actively degassing through vacuum and heating. The solid granu-
les of basic methacrylate copolymer formed in the extruder can be
milled to a powder. The copolymer may contain residual
monomers (dimethylaminoethyl methacrylate, butyl methacrylate,
methyl methacrylate,). Basic methacrylate copolymer is used as a
coating and glazing agent for food supplements and foods for
special medical purposes.
Chemical name Poly(butyl methacrylate-co-(2-dimethylaminoethyl)methacrylate-

co-methyl methacrylate)
C.A.S. number 24938-16-7
Chemical formula Poly[(CH2:C(CH3)CO2(CH2)2N(CH3)2)-co-(CH2:C(CH3)CO2CH3)-co-

(CH2 :C(CH3)CO2(CH2)3CH3)]
Structural formula
••• •••
The above formula is provided for illustrative purposes; specific

repeat units cannot be defined.
Formula weight 47,000 (weight-average), 22,000 (number-average)

Assay 20.8 – 25.5 % dimethylaminoethyl (DMAE) groups on the dried

basis
DESCRIPTION White powder
FUNCTIONAL USES Coating agent, glazing agent
CHARACTERISTICS
IDENTIFICATION
Viscosity 3 - 6 mPa∙s
Determine viscosity using Brookfield viscometer at 20° and 30 rpm

using UL adapter.
Refractive index n20

D : 1.380 - 1.385
(Vol. 4)
Solubility (Vol. 4) Freely soluble in methanol, ethanol, and 1 N aqueous hydrochloric

acid

corresponds to the infrared spectrum in the Appendix.

PURITY
Loss on drying (Vol. 4) Not more than 2.0% (110°, 3 h)
Particle size < 50 μm: at least 95 %

< 20 μm: at least 50 %
< 3 μm: not more than 10 %
Residual solvents Methanol: Not more than 50 mg/kg;

(Vol. 4)
Butanol: not more than 50 mg/kg and
Propan-2-ol: not more than 100 mg/kg.
(See General Methods, Organic Components, Residual Solvents,

Method 1)
Residual monomers Dimethylaminoethyl methacrylate: Not more than 500 mg/kg
Butyl methacrylate: Not more than 100 mg/kg
Lead (Vol. 4) Not more than 1.0 mg/kg
Determine using a method appropriate to the specified level. The

selection of sample size and method of sample preparation may
be based on principles of methods described in Volume 4 (under
“General Methods, Metallic Impurities”).
Microbiological criteria Total plate count: Not more than 1,000 cfu/g
(Vol. 4)
Yeast and moulds: Not more than 100 cfu/g
TESTS
PURITY TESTS
Particle size Determine using the light diffraction measurement method

according to Ph. Eur. 2.9.31
(European Pharmacopeia; Particle Size Analysis by Laser Light

Diffraction. 8 01/2010:0333)
Residual monomers Method for the determination of methyl methacrylate and

butyl methacrylate:

Phosphate buffer (0.0625 M, pH 2.0):
Prepare an aqueous solution containing 8.9 g of anhydrous
dibasic sodium phosphate and 8.5 g of monobasic
potassium phosphate in 1 L deionized water. Adjust with
phosphoric acid to pH 2.0.
Mobile phase:
Prepare a mixture of methanol and pH 2.0 phosphate
buffer (55:45).
Diluent:
Acetonitrile:Buffer (40:60)
Preparation of standard solution

Stock mixed standard solution:
Accurately weigh 20 mg of butyl methacrylate and 10 mg of
methyl methacrylate, dissolve in 3 ml of n-butanol and
dilute to volume to 10 ml with diluent in a volumetric flask.
Intermediate mixed standard solution:

Pipette 1.0 ml of stock mixed standard solution into a 10 ml
volumetric flask and dilute to 10 ml with diluent.
Working mixed standard solution:

Pipette 1.0 ml of intermediate standard solution into a 25
ml volumetric flask and dilute to volume with diluent. This
solution contains about 8 µg/ml of butylmethacrylate and 4
µg/ml of methyl methacrylate.
Preparation of Sample solution

Accurately weigh about 1.0 g of sample, dissolve in diluent and
make up to 50 ml with diluent and mix.
Procedure
Chromatographic system:
The liquid chromatograph is equipped with a UV/diode
array detector capable of working at 205 nm and a column
(4.6 mm × 12 cm, packing material: octadecylsilane
chemically bonded to porous silica or ceramic
microparticles, 1.5-10 µm)). Flow rate: 2 ml/min.
Chromatograph the working standard solution, and record the

peak responses. The resolution, R, between butyl methacrylate
and methyl methacrylate is not less than 10; and the relative
standard deviation for replicate injections is not more than 3.0%.
Separately inject 50 µl each of the working standard

solution and sample solution and record the peak areas of the
monomers.
Calculate the quantity of each monomer in the sample using the

formula:
Monomer concentration (µg/g) = (rU/rS) × (CS/CU) × F

rU = Peak area for the monomer in the sample
chromatogram
rS = Peak area for the monomer in the working standard
chromatogram
CS = Concentration of monomer in the working standard
solution (µg/ml)
CU = Concentration of polymer in the sample solution
(mg/ml)
F = Conversion factor (103 mg/g)
Method for the determination of 2-Dimethylaminoethyl

methacrylate

Monobasic potassium phosphate buffer solution (0.025M):
Prepare an aqueous solution containing 3.4 g of
monobasic potassium phosphate per litre.
Mobile phase:
Tetrahydrofuran:monobasic potassium phosphate buffer
solution (75:25).
Preparation of standard solution

Stock standard solution (200 µg/ml):
Accurately weigh about 20 mg of (2-dimethylaminoethyl)
methacrylate, dissolve in tetrahydrofuran, make up to
volume in a 10 ml volumetric flask with tetrahydrofuran and
mix.
Working standard solution (8 µg/ml):

Dilute 2.0 ml of the stock standard solution to 50 ml in a
volumetric flask with tetrahydrofuran and mix.
Preparation of Sample solution

Accurately weigh about 1.0 g of sample, dissolve in
tetrahydrofuran, dilute to 50 ml with tetrahydrofuran in a volumetric
flask and mix.
Procedure
Chromatographic system:
The liquid chromatograph is equipped with a UV/diode
array detector capable of working at 215 nm and a column
(4.6 mm × 12 cm, packing material: an essentially
monomolecular layer of aminopropylsilane chemically
bonded to totally porous silica gel support, 1.5-10 µm) .
Flow rate: 2 ml/min.
Chromatograph the working standard solution, and record the

peak area. The relative standard deviation for replicate injections
is not more than 2.0%.
Separately inject 50 µl of the working standard solution and

the sample solution and record the peak areas.
Calculate the quantity of each monomer in the sample using the

formula”
2-Dimethylaminoethyl methacrylate, (µg/g) = (rU/rS) × (CS/CU) × F
rU = Peak area for the monomer in the sample

chromatogram
rS = Peak area for the monomer in the standard
chromatogram
CS = Concentration of monomer in the working standard

solution (µg/ml)
CU = Concentration of polymer in the sample solution
(mg/ml)
F = Conversion factor (103 mg/g)
METHOD OF ASSAY Determine the percentage of Dimethylaminoethyl (DMAE) groups

using a potentiometric titration.
Dissolve 200 mg of dried sample in 4 ml water and 96 ml of glacial

acetic acid. Titrate with 0.1 N standard perchloric acid solution to a
potentiometric end point. Perform a blank determination.
𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷 𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔 (%𝑤𝑤/𝑤𝑤, 𝑜𝑜𝑜𝑜 𝑡𝑡ℎ𝑒𝑒 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏)

(𝑉𝑉𝑉𝑉 − 𝑉𝑉𝑉𝑉) × 𝑁𝑁 × 𝐹𝐹
= × 100
𝑊𝑊
VS = titrant volume consumed by the sample (ml)

VB = titrant volume consumed by the blank (ml)
N = actual normality of the titrant (mEq/ml)
F = equivalency factor, 72.1 mg/mEq
W = dried sample weight (mg)
Appendix: Infrared spectrum of basic methacrylate copolymer
90
%
T 80
r 70
a
n 60
s
m 50
i
t 40
t 30
a
n 20
c
e 10
4000 3000 2000 1000

Wavenumbers (cm-1)
CASSIA GUM
Prepared at the 86th JECFA (2018) and published in FAO

JECFA Monographs 22 (2018), superseding tentative
specifications prepared at the 82nd JECFA (2016) and
published in FAO JECFA Monographs 19 (2016). An ADI “not
specified” was established at the 71st JECFA (2009)
SYNONYMS INS 427
DEFINITION Cassia gum is obtained from the ground purified endosperm of

the seeds of Cassia tora and Cassia obtusifolia (Fam.
Leguminosae) containing less than 0.05% of Cassia
occidentalis. It consists mainly of high molecular weight
(approximately 200,000-300,000) polysaccharides composed
of galactomannans with a mannose:galactose ratio of about
5:1. The seeds are dehusked and degermed by thermal and
mechanical treatment followed by milling and screening of the
endosperm. The ground endosperm is purified by extraction
with isopropanol.
Structural formula
The structure above is provided for illustrative purposes. A

specific repeat unit for Cassia gum cannot be defined.
Assay Not less than 75% of galactomannans
DESCRIPTION Pale yellow to off-white, odourless free-flowing powder. Forms

colloidal solutions in cold water.
FUNCTIONAL USES Thickener, emulsifier, foam stabilizer, moisture retention agent,

and texturizing agent.
CHARACTERISTICS
IDENTIFICATION
Solubility Insoluble in ethanol
Gel formation with Add sodium borate TS to an aqueous dispersion of the sample
borate to raise the pH above 9; a gel is formed.
Gel formation with Passes test

xanthan gum
Gum constituents Proceed as directed under ‘Gum Constituents Identification’

(Vol. 4) using 100 mg of sample (instead of 200 mg) and 1-10 µl of the
hydrolysate (instead of 1-5 µl). Use galactose and mannose
as reference standards. These constituents should be present.
Viscosity Less than 500 mPa × s
pH (Vol. 4) 5.5-8.0 (1% solution)
PURITY
Loss on drying Not more than 12% (105º , 5 h)

(Vol. 4)
Total ash (Vol. 4) Not more than 1.2%
Acid-insoluble matter Not more than 2.0%

(Vol. 4)
Protein (Vol. 4) Not more than 7.0%
Multiply percent nitrogen by 6.25.
Crude fat Not more than 1%
Starch To a 1 in 10 dispersion of the sample add a few drops of iodine

TS; no blue colour is produced.
Anthraquinones Not more than 0.5 mg/kg
Residual solvents Isopropanol: Not more than 1.0%

(Vol. 4)
Lead (Vol. 4) Not more than 1 mg/kg

preparation may be based on the principles of the method

described in Volume 4 (under "General Methods, Metallic
Impurities").
Microbiological Total plate count: Not more than 5,000 cfu/g

criteria
(Vol. 4) Yeast and mould: Not more than 100 cfu/g
E. coli: Negative in 1 g
Salmonella: Negative in 25 g
TESTS
IDENTIFICATION
TESTS
Gel formation with Weigh 1.5 g of sample, 1.5 g of xanthan gum and blend them.
xanthan gum Add this blend (with rapid stirring) to 300 ml water at 80º in a
400 ml beaker. Stir until the mixture is dissolved and continue
stirring for an extra 30 min after dissolution (maintain the
temperature above 60º during the stirring period). Discontinue
stirring and allow the mixture to cool to room temperature for
at least 2 h.
A firm, viscoelastic gel forms after the temperature drops

below 40º, but no such gel forms in a 1% control solution of
cassia gum or xanthan gum alone prepared in a similar
manner.
Viscosity Add 495 ml of deionized water into a 1L beaker, insert a

magnetic stir bar and place the beaker on a magnetic stirrer
equipped with a heater. Adjust the stirrer speed to about
750 rpm. Weigh 5 g of sample and quickly add to the beaker.
Switch on the heater and heat the beaker to reach 90º and
keep it at 90o for 15 min. Cool the solution to room
temperature (25º ±1.5º) in a water bath. Measure the viscosity
at 25º, after 2 h, using a RVT Brookfield Spindle 1 and 20 rpm
speed. Repeat the procedure with a sample of 5 g of carob
(locust) bean gum. The viscosity of the cassia gum (150 –
500 mPa × s) must be less than 50% that of carob bean gum
(2000 - 3500 mPa × s)
PURITY TESTS
Residual solvents Determine residual solvents using headspace gas

chromatography (Vol. 4; Method I) under the following
conditions.
Internal standard solution

Add 50.0 ml water to a 50 ml vial and seal. Accurately weigh

and inject 15 µl of 3-methyl-2-pentanone through the septum
and reweigh the vial to within 0.01 mg.
Standard solution
Add 50.0 ml water to a 50 ml vial and seal weigh accurately.
Inject 15 µl isopropanol and reweigh the vial.
Blank solution:
Add 5.0 ml of water and pipette 1.0 ml of the internal
standard solution into a headspace vial. Seal the vial
and mix the contents using a vortex mixer.
Calibration solution:
Add 4.0 ml of water into the headspace vial. Pipette
1.0 ml each of the internal standard solution and the
standard solution. Seal the vial and mix the contents
using a vortex mixer.
Preparation of sample:
Pipette 5 ml of water and 1 ml internal standard
solution into a headspace vial. Accurately weigh 0.500
± 0.001 g of sample in a small weighing boat and add
the sample carefully to prevent clumping of sample at
the bottom of the vial. Seal the vial and mix the
contents using a vortex mixer. Do not shake the
sample vial.
Follow the procedure described in Vol. 4 for the determination

of residual solvents.
Crude fat Apparatus

The apparatus consisting of a Butt-type extractor, as shown
below, having a standard-taper 34/45 female joint at the upper
end, to which is attached a Friedrichs- or Hopkins-type
condenser, and a 24/40 male joint at the lower end, to which is
attached a 125-ml Erlenmeyer flask.
Procedure
Transfer about 10 g of the sample, previously ground to
20-mesh or finer and accurately weighed, to a cellulose
thimble or a 15-cm filter paper (roll the paper tightly around the
sample), and place it in a suitable extraction shell). Plug the
top of the thimble or the extraction shell with cotton previously
extracted with hexane, and place it in the extractor. Attach the
extractor to a dry 125-ml Erlenmeyer flask containing about 50
ml of hexane and to a water-cooled condenser, apply heat to
the flask to produce 150 to 200 drops of condensed solvent
per min, and extract for 16 h. Disconnect the flask, and filter
the extract to remove any insoluble residue. Rinse the flask
and filter with a few ml of hexane, combine the washings and
filtrate in a tared flask, and evaporate on a steam bath until no
odour of solvent remains. Dry in a vacuum for 1 h at 100°,
cool in a desiccator, and weigh.
Butt-Type Extractor for fat determination.
NOTE: The method for crude fat is referenced from Appendix

X: Crude fat in the Food Chemicals Codex, 11th Edition, 2018.
Reproduced from USP-NF with permission from The U.S.
Pharmacopeial Convention (USP)
Anthraquinones Principle
Anthraquinones are extracted with chloroform and
determined by High Performance Liquid
Chromatography (Vol.4) using the conditions below.
NOTE: Anthraquinones are photosensitive. Samples and

standards shall be protected from light and all manipulations
shall be carried out under the subdued light.

Emodin, Aloe-emodin, Physcion (1,8-dihydroxy-3-
methoxy-6-methyl-anthraquinone), Rhein and
Chrysophanic acid (>99%).
Internal standard: Danthrone (1,8-dihydroxy-
anthraquinone, >99%)
Methanol, acetonitrile, deionized water, chloroform,

trifluoroacetic acid, sulfuric acid and sodium hydrogen
carbonate
Individual stock standard and internal standard solutions (100

µg/ml)
Accurately weigh about 10 mg of the standards and
internal standard, transfer to 100 ml volumetric flasks
with about 5 ml of methanol, sonicate for 15 min and
dilute to volume with methanol.
Store these solutions in amber coloured bottles at 4º
(the solutions are stable for 2 weeks under these
conditions).
Internal spike standard solution (20 µg/ml)

Dilute 2 ml of internal standard stock solution to 10 ml
with methanol.
Mixed standard solution (10 µg/ml)

Transfer 1 ml of each of the anthraquinones stock
standard solution into a 10 ml volumetric flask and
dilute to volume with methanol.
Working standard solutions

To each of five 10 ml volumetric flasks transfer 0, 0.5,
1, 2 and 5 ml respectively of the mixed standard
solution, and 1 ml of the internal spike standard
solution (20 µg/ml), dilute to volume with methanol and
mix.
Sample preparation:
Accurately weigh about 4.0 g of the sample into a
250 ml Erlenmeyer flask. Add 80 µl of internal standard
solution (100 µg/ml), and 100 ml 2N H2SO4 to the flask.
Stopper the flask using a PTFE stopper and heat at
103º for 3.5 hours in an oven. After cooling to room
temperature, add 100 ml of chloroform and shake well.
Allow phase separation. Evaporate 50 ml of the
chloroform layer to dryness in a rotary evaporator at
68º. Dissolve the residue in 2 ml of methanol. Filter the
solution through a PTFE membrane syringe filter.
Chromatographic conditions:
Column: Hypersil ODS C18 (250 mm x 4.6 mm ID,
5 µm)
Mobile phase:
(A) 0.1 % trifluoroacetic acid in water
(B) Acetonitrile
Injection volume: 20 µl
Gradient:
Time, min % (A) % (B)

0 86 14
10 86 14
15 80 20
25 80 20
55 20 80
60 0 100
66 86 14
Flow rate: 1 ml/min

Detector: Photodiode Array/UV Detector operated at 435 nm.
Procedure
Inject individual standard solutions and internal
standard solution (dilute, if required) and record
retention times.
Construction of standard curves

Inject 20 µl of each working standard solution.
Construct the standard curves by plotting the ratios of
the peak areas of each of the specific anthraquinone /
internal standard against the concentrations of each
working standard solution (µg/ml).
Inject 20 µl of the Sample solution. Calculate the ratios

of the peak areas of each anthraquinone / internal
standard, and obtain the amount (A) of each
anthraquinone from the respective standard curve.
Concentration of
anthraquinone in the sample (µg/g) = (A × 4)/ W
Where
A = the amount of each anthraquinone (µg) obtained from the
standard curve
W = Mass of sample (g)
4 = Dilution factor for sample
METHOD OF ASSAY % Galactomannans = 100 – (L + A + I + P + F)
L % Loss on Drying
A %Total Ash
I %Acid-Insoluble Matter
P %Protein
F %Crude Fat
CITRIC AND FATTY ACID ESTERS OF GLYCEROL (TENTATIVE)
Prepared at the 86th JECFA (2018) and published in FAO JECFA

Monographs 22 (2018), superseding specifications prepared at the
82nd JECFA (2016), and published in FAO JECFA Monographs 19
(2016). An ADI 'not limited' was established at the 17th JECFA
(1973)
Information required:
• A validated method for the determination of total citric acid
content
• Performance characteristics (method validation data) of the
citric acid determination method
• Data on the total citric acid content, in at least five batches
of products currently available in commerce, determined
using the above method.
SYNONYMS Citric acid esters of mono- and di-glycerides, citroglycerides,

CITREM; INS No. 472c
DEFINITION Citric and fatty acid esters of glycerol (CITREM) consists of mixed
esters of citric acid and edible fatty acids with glycerol. It may
contain free fatty acids, glycerol, citric acid and mono- and
diglycerides, in minor quantities. The mono- and diglycerides may
include either one or two edible fatty acids from C12:0 to C18:0,
mainly the saturated palmitic (C16:0) and stearic (C18:0) acids. It
may also contain minor amounts of other fatty acids such as
myristic (C14:0), oleic (C18:1), linoleic (C18:2) and arachidic acid
(C20:0). CITREM is obtained by esterification of glycerol with citric
acid and edible fatty acids, or by reaction of a mixture of mono- and
diglycerides of edible fatty acids, with citric acid. CITREM may be
partially or wholly neutralized with sodium hydroxide or potassium
hydroxide.
Structural formula
Where at least one of R1, R2 or R3 represents a citric acid moiety,

one represents a fatty acid moiety and the remainder may
represent citric acid, fatty acid or hydrogen.
DESCRIPTION White to ivory coloured, oily to waxy material.
FUNCTIONAL USES Stabilizer, emulsifier, dough conditioner, antioxidant synergist

CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Insoluble in water; soluble in oils and fats; insoluble in ethanol
Test for fatty acids Passes test

(Vol. 4)
Test for citric acid Information required
Test for glycerol Passes test

(Vol. 4)
PURITY
Sulfated ash (Vol. 4) Non-neutralized products: not more than 0.5%

Partially or wholly neutralized products: not more than 10%; test 2 g
of the sample (Method I)
Free glycerol (Vol. 4) Not more than 4%
Total glycerol 8-33%
Total citric acid 13-50%
(Information required)
Total fatty acid 37-81%

Lead (Vol. 4) Not more than 2 mg/kg.

(Not more than 0.1 mg/kg for use in infant formula and formula for
special medical purposes intended for infants)

selection of sample size and method of sample preparation may be
based on the principles of the methods described in Volume 4
(under “General Methods, Metallic Impurities”).
TESTS
PURITY TESTS
Total glycerol CITREM is hydrolyzed, glycerol in the aqueous phase is oxidized

using known excess of sodium periodate in a strongly acid medium
and the unreacted periodate is back titrated using standard sodium
thiosulfate solution.
Procedure:
Accurately weigh about 2 g of the sample into a saponification
flask, add 50 ml of 0.5 M ethanolic potassium hydroxide, and reflux
for 30 min.
To a 1-L volumetric flask add 99 ml + 0.2 ml of chloroform using a

burette and add 25 ml of glacial acetic acid using a graduated
cylinder. Quantitatively transfer the content of the saponification
flask to the volumetric flask, using three 25 ml portions of water.
Add about 500 ml of water further, and shake vigorously for about
1 min. Dilute to volume with water, stopper, mix thoroughly and set
aside for separation of layers.
Pipet 50 ml of acetic periodic acid TS into a series of 400 ml

beakers. Prepare two blanks by adding 50 ml of water to each.
Pipet 50 ml of the aqueous layer into one of the 400 ml beakers
containing 50 ml of acetic periodic acid TS; shake gently to mix;
cover with watch glass, and allow to stand 30 min but not longer
than 1.5 h. Add 20 ml of 15% potassium iodide solution, shake
gently to mix, and allow to stand at least 1 min. but not more than
5 min. Do not allow to stand in bright or direct sunlight. Add 200 ml
of water and titrate with 0.1 N sodium thiosulfate. Use a variable
speed electric stirrer to keep the solution thoroughly mixed.
Continue the titration to the disappearance of the brown iodine
colour from the aqueous layer. Add 2 ml of starch TS and continue
the titration to the disappearance of iodine from the tiny chloroform
layer separated during titration and the disappearance of the blue
iodine-starch complex colour from the aqueous layer. Read the
burette to the nearest 0.01 ml. Treat the blanks in the same way as
the sample.
Calculation
% total glycerol = [(B − S) × N × 2.302 × 900]/(W × 50)
where
B volume of 0.1 N sodium thiosulfate used for the blank, ml
S volume of 0.1 N sodium thiosulfate used for the sample, ml
N exact normality of 0.1 N sodium thiosulfate
W mass of sample, g
Total citric acid Information required

Total fatty acid Principle: This method measures total fatty acids by extracting with
diethyl ether.
Procedure
Weigh accurately 5 g of the sample into a 250-ml round-bottomed
flask, add 50 ml of potassium hydroxide, ethanolic, TS, and reflux for
1 h on a boiling water bath.
Quantitatively transfer the contents of the saponification flask to a

1,000 ml separating funnel, using three 25 ml portions of water, and
add 5 drops of methyl orange indicator solution.
Cautiously add 50% hydrochloric acid until the colour of solution

changes to orange red.t. Add 1 ml of excess acid. Shake well to
mix the contents and separate the fatty acids.
Cool to room temperature and extract the separated fatty acids with
three 100 ml portions of diethyl ether. Combine the extracts, and
wash with 50 ml portions of 10% sodium chloride solution until the
washed sodium chloride solution becomes neutral.
Dry the ether solution with anhydrous sodium sulfate. Then

evaporate off ether on a steam bath, leave additional 10 min on the
steam bath, and weigh the residue. This is the weight of the total
fatty acids.
Calculation:
mass of fatty acids g × 100

Total Fatty acids % =
mass of sample g
ERYTHROSINE
Prepared at the 86th JECFA and published in FAO JECFA Monograph 22 (2018)
superseding specifications prepared at the 41st JECFA (1993), published in FNP 52
Add 2 (1993). Metals and arsenic specifications revised at the 59th JECFA (2002). An
ADI of 0-0.1 mg/kg bw was established at the 37th JECFA (1991) and confirmed at
the 86th JECFA (2018).
SYNONYMS INS No. 127, CI Food Red 14, CI (1975) No. 45430,
Food Red No. 3, FD&C Red No. 3
DEFINITION Erythrosine consists of the disodium salt of 2-(2,4,5,7-

tetraiodo-6-oxido-3-oxoxanthen-9-yl)benzoate
monohydrate and subsidiary colouring matters. Sodium
chloride and/or sodium sulfate are the principal
uncoloured components. Erythrosine is manufactured by
iodination of fluorescein, the condensation product of
resorcinol and phthalic anhydride.
Erythrosine may be converted to the corresponding

aluminium lake in which case only the requirements in
the General Specifications for Aluminium Lakes of
Colouring Matters apply.
Chemical names Disodium 2-(2,4,5,7-tetraiodo-6-oxido-3-oxoxanthen-9-

yl)benzoate monohydrate;
Disodium;2',4',5',7'-tetraiodo-3-oxospiro[2-benzofuran-
1,9'-xanthene]-3',6'-diolate;
Disodium 2′,4′,5’,7′-tetraiodofluorescein monohydrate
C.A.S. number 16423-68-0
Chemical formula C20H6I4Na2O5 · H2O
Structural formula
Formula weight 879.86
Assay Not less than 87% total colouring matters

DESCRIPTION Red powder or granules
FUNCTIONAL USES Colour
CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Soluble in water, slightly soluble in ethanol
Spectrophotometry Maximum wavelength approximately 527 nm

(Vol. 4)
Determine the UV-visible absorption spectrum of the
sample dissolved in water.
PURITY
Loss on drying, Not more than 13%

chloride and sulfate as
sodium salts Determine chloride as sodium chloride, sulfate as sodium
(Vol. 4) sulfate, and loss on drying (135°, 6 h) as described in
Volume 4 (under “Specific Methods, Food Colours”).
Inorganic iodides Not more than 0.1% calculated as sodium iodide
Water insoluble matter Not more than 0.2%

(Vol. 4)
Zinc (Vol. 4) Not more than 50 mg/kg
Determine using a method appropriate to the specified

level. The selection of sample size and method of
sample preparation may be based on the principles of
the method described in Volume 4 (under “General
Methods, Metallic Impurities”).

level. The selection of sample size and method of
sample preparation may be based on the principles of
the method described in Volume 4 (under “General
Methods, Metallic Impurities”).
Subsidiary colouring Not more than 4% (except fluorescein)

matters
Note: Do not allow the sample and standard solutions to

be exposed to direct sunlight.
Fluorescein Not more than 20 mg/kg
Organic compounds Triiodoresorcinol: Not more than 0.2%

other than colouring
matters 2-(2,4-dihydroxy-3,5-diiodobenzoyl)benzoic acid: Not
more than 0.2%
Ether extractable From a solution of pH not less than 7, not more than
matter (Vol. 4) 0.2%
Hydrochloric acid- Not more than 0.5%

insoluble matter in
Erythrosine Lake See description under TESTS
TESTS
PURITY TESTS
Inorganic iodides Weigh 1.0 g of the sample into a 100-ml beaker. Add
75 ml distilled water and a magnetic stirrer. Stir to
dissolve. Immerse an iodide specific electrode and a
reference electrode in the solution and use a suitable
millivoltmeter to read the potential of the system in
millivolts.
Add 0.001 M silver nitrate solution from a burette initially

in 0.5 ml aliquots, reducing these to 0.1 ml as the end-
point approaches as indicated by an increasing change
in potential for each addition. After allowing time for the
reading to stabilize, record the millivolt readings after
each addition. Continue the titration until further additions
make little change in the potential.
Plot the millivolt readings against the volume of silver

nitrate solution added. The equivalence point is the
volume corresponding to the maximum slope of the
curve.
The percentage of sodium iodide in the sample = Titre ×

0.015%
where
Titre ml-equivalent of silver nitrate solution
0.015% 0.001 mol/l x 10-3 l/ml x 149.89 g sodium

iodide/mol x 1 mol/equivalent x 1/1.0 g
(sample weight) x 100.
Subsidiary colouring Determine subsidiary colouring matters content by

matters reversed-phase HPLC (Vol. 4) using the following
conditions:
- Column: C8 (250 mm x 4.6 mm i.d., 5 µm particle

size)
- Eluent A: 0.1 M ammonium acetate in water
- Eluent B: methanol
- Injection volume: 20 μl
- Column temperature: ambient
- Detector: UV-visible/diode array at 514 nm
- Flow rate: 1.0 ml/min
Gradient:
Elution time Eluent A (%) Eluent B (%)

(min)
0 55 45
20 34 66
21.1 0 100
25.5 0 100
26.0 55 45
40.0 55 45
Reagents: HPLC grade
Standards:
- 2′,4′,5′-Triiodofluorescein (C.A.S. 56254-06-9) –
synthesized material (see Appendix)
- 2′,4′,7′-Triiodofluorescein (C.A.S. 83498-90-2) –
synthesized material (see Appendix)
- 4′,5′-Diiodofluorescein, disodium salt (C.A.S. 33239-
19-9) – Alfa Aesar, Cat. No. A15626 or equivalent
- 2′-Monoiodofluorescein, disodium salt
(C.A.S. 52010-85-2) – synthesized material (see
Appendix)
- 4′-Monoiodofluorescein, disodium salt
(C.A.S. 52010-86-3) – synthesized material (see
Appendix)
- Erythrosine (C.A.S. 16423-68-0) – TCI, >95.0%
disodium 2′,4′,5’,7′-tetraiodofluorescein, Cat.
No. F0139 or equivalent (use if subsidiary colouring
matter standards are not available)
Prepare standard solutions as required.
Sample preparation:
Weigh accurately 200±2 mg sample and dissolve
in 100 ml of water. Dilute the solution, if required,
to separate subsidiary colours from the primary
colour component in order to improve their
resolution.
Calculations:
Construct the relevant standard curves. Integrate
all peaks of the chromatogram obtained at
514 nm. If Erythrosine is used as a standard,
calculate the ratio of the sum of all peaks not
corresponding to Erythrosine to the sum of all
peaks.
Fluorescein Determine fluorescein by the test for subsidiary colouring

matters content except use the following conditions:
Standard: Fluorescein, disodium salt (C.A.S. 518-47-8) –

TCI, Cat. No. F0096 or equivalent
Sample preparation:
Weigh accurately 2.00±0.05 g sample and
dissolve in 10 ml of water.
Organic compounds Determine organic compounds other than colouring

other than colouring matters by reversed-phase HPLC (Vol. 4) using the
matters following conditions:
- Column: C18 (150 mm x 2.1 mm i.d., 5 µm particle
size)
- Eluent A: 0.05 M sodium dihydrogen phosphate in
95/5 water/methanol, pH 4.0
- Eluent B: methanol
- Column temperature: 27°
Gradient:

(min)
0 95 5
3 95 5
5 80 20
13 35 65
15 0 100
25 0 100
27 95 5
37 95 5
Standards:
- 2,4,6-Triiodoresorcinol (C.A.S. 19403-92-0) – Alfa
Chemistry, Cat. No. ACM19403920 or equivalent
- 2-(2,4-Dihydroxy-3,5-diiodobenzoyl)benzoic acid
(C.A.S. 3480-21-5) – Wako, Cat. No. 043-32981 or
equivalent
Prepare standard solutions as required. Dissolve the

standards in methanol. Use an amber glass volumetric
flask for 2,4,6-triiodoresorcinol and prepare the standard
and calibration solutions immediately before use.
Sample preparation:
in 10 ml of methanol.
Calculations:
all peaks of the chromatogram obtained at
223 nm.
Hydrochloric acid- Reagents

insoluble matter in - Concentrated hydrochloric acid
Erythrosine Lake - Hydrochloric acid, 0.5% v/v
- Dilute ammonium hydroxide solution (dilute 10 ml of
14.5 M ammonium hydroxide to 100 ml with water).
Procedure
Accurately weigh approximately 5 g of the lake into a
500-ml beaker. Add 250 ml water and 60 ml
concentrated hydrochloric acid. Boil to dissolve the
alumina while the Erythrosine converts to its "free acid"
form, which is insoluble in acid. Filter through a tared
No. 4 sintered glass crucible. Wash the crucible with a
small amount of hot 0.5% hydrochloric acid and then with
some hot distilled water. Remove the acid filtrate from
the filter flask, replace the crucible, and wash with hot
dilute ammonium hydroxide solution until the washings
are colourless. Dry the crucible to constant weight at
135º. Express the residue as a percentage of the weight
taken.
METHOD OF ASSAY Determine total colouring matters content by

spectrophotometry using Procedure 1 in Volume 4
(under “Specific Methods, Food Colours”) and an
appropriate solvent.
Using water as the solvent:

absorptivity (a) = 110 l/(g × cm)
wavelength of maximum absorbance = 527 nm.
GLYCEROL ESTER OF WOOD ROSIN
Prepared at the 86th JECFA and published in FAO JECFA Monographs 22 (2018),
superseding specifications prepared at the 77th JECFA (2013) and published in FAO
JECFA Monographs 14 (2013). An ADI of 0-25 mg/kg bw for glycerol ester of wood
rosin was established at the 77th JECFA (2013).
SYNONYMS INS No. 445(iii)
DEFINITION Glycerol ester of wood rosin (GEWR) is a complex mixture of

glycerol di- and tri- esters of resin acids from wood rosin, with a
residual fraction of glycerol monoesters. In addition, neutrals
(non-acidic saponifiable and unsaponifiable substances) and
residual free resin acids are present. Wood rosin is obtained by
the solvent extraction of aged pine stumps, followed by a
liquid-liquid solvent refining process. Refined wood rosin is
composed of approximately 90% resin acids and
approximately 10% neutrals. The resin acid fraction is a
complex mixture of isomeric diterpenoid monocarboxylic acids
having the typical empirical formula C20H30O2, of which the
main components are dehydroabietic and abietic acids. GEWR
is produced by esterifying the resin acids with food grade
glycerol. The product is then purified by steam stripping or by
direct countercurrent steam distillation.
These specifications do not cover substances derived from

gum rosin, an exudate of living pine trees, and substances
derived from tall oil rosin, a by-product of kraft (paper) pulp
processing.
C.A.S. number 8050-30-4
DESCRIPTION Hard, yellow to pale amber-coloured solid
FUNCTIONAL USES Emulsifier, density adjustment agent (flavouring oils in

beverages), stabilizer, plasticizer (in chewing gum bases).
CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Insoluble in water, soluble in acetone
Infrared absorption The infrared spectrum of a thin film of the sample (potassium
(Vol. 4) bromide disc) corresponds with the typical infrared spectrum
below
Sulfur test Negative
Weigh 40-50 mg of sample into a test tube and add 1- 2 drops

of a 20% (w/v) solution of sodium formate. Place a strip of lead
acetate test paper over the mouth of the test tube. Heat the
tube until fumes are formed that contact the test paper.
Continue heating for 2-5 min. The formation of a black spot of
lead sulfide indicates the presence of sulfur-containing
compounds. (Detection Limit: 50 mg/kg sulfur)
Gas chromatography Passes test

of resin acids and
glycerol See description under TESTS
PURITY
Specific gravity d (20, 25): Not less than 0.935 (50% solution in d-limonene)
(Vol. 4)
Ring and ball Not less than 82° (see “Specific Methods, Glycerol Esters of
softening point (Vol. 4) Rosins”)
Acid value (Vol. 4) Between 3 and 9 (see “Specific Methods, Fats, Oils, and
Hydrocarbons”)
Determine using method appropriate to the specified level. The

selection of sample size and method of sample preparation
may be based on the principles of the method described in
Volume 4 (under “General Methods, Metallic Impurities”).
TESTS
IDENTIFICATION
TESTS
Gas chromatography The ester groups in the glycerol esters of wood rosin are
of resin acids reduced with a metal hydride to form a mixture of
corresponding resin alcohols and glycerol which are analyzed
by gas chromatography (Vol. 4). The characteristic
chromatogram shows predominant peaks for abietic and
dehydroabietic alcohols.
Apparatus
- Gas Chromatograph equipped with a flame ionization
detector.
- Centrifuge: table top, capable of achieving 3200 rpm
Standards and reagents

- Internal Standard (1,4-Butanediol: >99%
- Toluene
- Sodium Vitride Reagent [(Sodium bis(2-methoxyethoxy)
aluminium dihydride], 70% in toluene:(~ 3.5 mol/l)
Sodium Vitride solution:

Pipet 10.0 ml of sodium vitride reagent into a 100 ml
volumetric flask dilute to volume with toluene and mix
thoroughly.
Hydrolysis solution:
Slowly add 50 ml of concentrated sulfuric acid, reagent
grade, to 200 ml distilled water while stirring in an ice
bath. Cool to room temperature.
Procedure
Sample preparation
Weigh 250-300 mg sample into a 25 ml Erlenmeyer
flask containing a Teflon coated stirrer bar. Pipet 5.0 ml
toluene into the flask and stir until sample is dissolved.
Pipet 5.0 ml of sodium vitiride solution into the flask,
stopper the flask and stir for 30 min. While stirring, pipet
3.0 ml of hydrolysis solution into the flask. Continue
stirring for 3 min. Transfer contents of flask to
centrifuge tube (15 ml), stopper, and shake vigorously.
Vent and centrifuge at 2800-3200 rpm for 5 min. Inject
0.5 µl of the toluene layer into the gas chromatograph
operating under the following conditions and record the
chromatogram. Compare with the chromatogram
shown below to verify the approximate retention order
of the resin alcohols.
Chromatographic conditions
- Column: DB-1 methyl silicone (bonded and crosslinked)
wide-bore capillary (15 m x 0.53 mm i.d.,1.5 µm).
- Injector: Flash vaporization injector
- Flow rates: Carrier Gas (He): 30 ml/min at 63 psi,
Hydrogen: 30 ml/min and
- Air: 240 ml/min
- Temperatures: Column: Isothermal, 190º; Injector:
250º , and Detector: 250º
Gas chromatography Standards and reagents
of glycerol - Glycerol: >99%
- 1,4-Butanediol (Internal standard): >99%
Internal Standard Solution:

Weigh 0.1 g of 1,4-butanediol into a 100 ml volumetric
flask. Dilute to volume with distilled water and mix
thoroughly.
Glycerol solution:
Weigh 0.1 g of 1,4-butanediol and 0.1 g glycerol into a
100 ml volumetric flask. Dilute to volume with distilled
water and mix thoroughly
Phenolphthalein Solution: 1% in ethanol.
Sodium Hydroxide Solution:

Dissolve 16 g of reagent grade NaOH in 70-80 ml of
distilled water and cool to room temperature. Dilute to
100 ml with distilled water and mix thoroughly. Store in
a polyethylene bottle.
Procedure
Sample preparation
Proceed as in the sample preparation for the analysis
of resin acids until the centrifugation step. Using a pipet
or syringe, remove the toluene layer and part of the
aqueous layer leaving approximately 2 ml of the
aqueous layer in the centrifuge tube. Add 1 drop of
phenolphthalein solution to the remaining aqueous
layer in the centrifuge, and neutralize with the sodium
hydroxide solution (aluminium salts will precipitate).
Pipet 5 ml of the internal standard solution into the
tube, dilute to 15 ml with distilled water, stopper, shake,
and then centrifuge at 2800-3200 rpm for 5 min. Inject
1 µl of the clear supernatant liquid into the gas
chromatograph operating under the following conditions
and record the chromatogram. Inject 1 µl of the glycerol
solution and record the chromatogram. Measure the
retention times of any observed peaks relative to
1,4-butanediol. Compare retention times to that of
glycerol standard.
Chromatographic conditions
- Column: DB-WAX polyethyleneglycol (bonded and
cross-linked), wide bore capillary (15 m x 0.53 mm i.d.,
1.0 µm)
- Flow rates: Carrier Gas (He): 30 ml/min at 60 psi,
Hydrogen: 30 ml/min and
- Air: 240 ml/min
- Temperatures: Column: Programmed, 120 to 200º at 6º
/min; Injector: 250º,
- and Detector: 250º
Gas chromatography of resin acids in GEWR (determined as

alcohols)
Typical GC-FID chromatogram of a GEWR sample.
Retention times correspond to pimaric (18.8 min),
isopimaric (22.4 min), palustric (23.1 min),
dehydroabietic (25.9 min), abietic (29.6 min), and
neoabietic (35.2 min) alcohols. This is a product
derived from a plant-based source which can

demonstrate significant variability and relative
intensities.
Gas chromatogram
FTIR Spectrum of Glycerol esters of wood rosin

INDIGOTINE
Prepared at the 86th JECFA and published in JECFA Monograph 22 (2018)

superseding specifications prepared at the 28th JECFA (1984), published in FNP 31/1
(1984) and in FNP 52 (1992). Metals and arsenic specifications revised at the 59th
JECFA (2002). An ADI of 0-5 mg/kg bw was established at the 18th JECFA (1974) and
confirmed at the 86th JECFA (2018).
SYNONYMS INS No. 132, CI Food Blue 1, CI (1975) No. 73015, Indigo
Carmine, Food Blue No. 2, FD&C Blue No. 2
DEFINITION Indigotine consists of a mixture of disodium 3,3'-dioxo-

[delta2,2'-biindoline]-5,5'-disulfonate and disodium 3,3'-
dioxo-[delta2,2'-biindoline]-5,7'-disulfonate and subsidiary
colouring matters. Sodium chloride and/or sodium sulfate
are the principal uncoloured components. Indigotine is
manufactured by heating indigo in the presence of sulfuric
acid. The indigo (or indigo paste) is manufactured by the
fusion of N-phenylglycine (prepared from aniline and
formaldehyde) in a molten mixture of sodamide and
sodium and potassium hydroxides under ammonia
pressure. It is isolated and subjected to purification
procedures prior to sulfonation.
Indigotine may be converted to the corresponding

aluminium lake in which case only the requirements in the
General Specifications for Aluminium Lakes of Colouring
Matters apply.
Chemical names Disodium 3,3'-dioxo-[delta2,2'-biindoline]-5,5'-disulfonate
Disodium (2E)-3-oxo-2-(3-oxo-5-sulfonato-2,3-dihydro-
1H-indol-2-ylidene)-2,3-dihydro-1H-indole-5-sulfonate
Disodium;(2E)-3-oxo-2-(3-oxo-5-sulfonato-1H-indol-2-
ylidene)-1H-indole-5-sulfonate
C.A.S. number 860-22-0 (5,5' isomer)
Chemical formula C16H8N2Na2O8S2
Structural formula

Assay Not less than 85% total colouring matters
Not more than 18% of disodium 3,3'-dioxo-[delta2,2'-

biindoline]-5,7'-disulfonate
DESCRIPTION Blue powder or granules
CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Soluble in water, sparingly soluble in ethanol
Spectrophotometry Maximum wavelength approximately 610 nm

(Vol. 4)
Determine the UV-visible absorption spectrum of the
sample dissolved in water.
PURITY
Loss on drying, Not more than 15%

chloride and sulfate
as sodium salts (Vol. 4) Determine chloride as sodium chloride, sulfate as sodium
sulfate, and loss on drying (135°, 6 h) as described in
Volume 4 (under “Specific Methods, Food Colours”).
Water insoluble matter Not more than 0.2%

(Vol. 4)

level. The selection of sample size and method of sample
preparation may be based on the principles of the method
described in Volume 4 (under “General Methods, Metallic
Impurities”).
Subsidiary colouring Not more than 18% disodium 3,3'-dioxo-[delta2,2'-

matters biindoline]-5,7'-disulfonate (isomeric subsidiary colouring
matter)
Not more than 1% other subsidiary colouring matters
Organic compounds other Not more than 0.5% of sum of isatin-5-sulfonic acid, 5-
than colouring matters sulfoanthranilic acid, and anthranilic acid
Unsulfonated primary Not more than 0.01% calculated as aniline

aromatic amines
(Vol. 4)
Ether extractable matter Not more than 0.2%

(Vol. 4)
Weigh accurately about 2 g sample instead of the 5 g
stated in the general methods
TESTS
PURITY TESTS
Subsidiary colouring Determine subsidiary colouring matters content by

matters reversed-phase HPLC (Vol. 4) using the following
conditions:
- Column: C18 (250 mm x 4 mm i.d., 5 µm particle
size)
- Eluent A: 0.2 M ammonium acetate in water
- Eluent B: acetonitrile
- Column temperature: ambient
Gradient:

(min)
0 100 0
20.0 40 60
30.0 40 60
32.0 100 0
40.0 100 0
Standards:
- Indigotine disodium salt (5,7′ isomer) (isomeric
subsidiary colour) (C.A.S. 27414-68-2) – Angene
Chemical, Cat. No. AGN-PC-0R372R or
equivalent
- Sodium indigo sulfonate monosodium salt
(monosulfonated subsidiary colour) (C.A.S. 27414-
69-3) – Atomax Chemicals Co., Ltd., Cat No.
AM27414693 or equivalent
- Trisodium indigo-5,5′,7′-trisulfonate
- (trisulfonated subsidiary colour), potassium salt
(C.A.S. 67627-18-3) – Sigma-Aldrich, Cat
No. 234087 or equivalent
- Indigotine (C.A.S. No. 860-22-0) – TCI, Cat.
No. F0148 or equivalent (use if subsidiary
colouring matter standards are not available)
Sample preparation:
in 100 ml of water. Dilute the solution, if required,
to separate subsidiary colours from the primary
colour component in order to improve their
resolution. Analyze immediately after preparation.
Calculations:
all peaks of the chromatogram obtained at 610
nm. If Indigotine is used as a standard, calculate
the ratio of the sum of all peaks not corresponding
to Indigotine to the sum of all peaks.
Organic compounds other Determine organic compounds other than colouring

than colouring matters matters content by reversed-phase HPLC (Vol. 4) using
the following conditions:
- Column: Luna C18 (250 mm x 4.6 mm i.d., 5 µm
particle size) or equivalent
- Eluent A: 0.1% trifluoroacetic acid in water
- Eluent B: acetonitrile
Gradient:

(min)
0 100 0
15 80 20
20 0 100
22 0 100
22.1 100 0
32 100 0

Standards:
- Isatin-5-sulfonic acid sodium salt dihydrate
(C.A.S. 207399-16-4) – Sigma-Aldrich Cat.
No. 58245 or equivalent
- 5-Sulfoanthranilic acid (2-amino-5-sulfobenzoic
acid) (C.A.S. 3577-63-7) – TCI Cat No. S0802 or
equivalent
- Anthranilic acid (C.A.S. 118-92-3) – Sigma-Aldrich
Cat No. A89855 or equivalent
Sample preparation:
in 100 ml of water. Analyse immediately after
preparation.
Calculations:
the chromatogram peaks obtained at 244 nm.
METHOD OF ASSAY Determine total colouring matters content by

spectrophotometry using Procedure 1 in Volume 4 (under
“Specific Methods, Food Colours”) and an appropriate
solvent. Analyse immediately after preparation.
Using water as the solvent:

absorptivity (𝑎𝑎) = 48.0 𝑙𝑙/(𝑔𝑔 × 𝑐𝑐𝑐𝑐)
wavelength of maximum absorbance = 610 nm.
Determine isomer content by HPLC using the test for

subsidiary colouring matters.
LUTEIN FROM TAGETES ERECTA
Prepared at the 86th JECFA (2018) and published in FAO

JECFA Monograph 22 (2018), superseding specifications
prepared at the 63rd JECFA (2004) and published in FNP52
Add 12 (2004). A group ADI of “not specified” was established
for Tagetes extract, Lutein from Tagetes erecta, Lutein esters
from Tagetes erecta, Zeaxanthin (synthetic), and meso-
zeaxanthin at the 86th JECFA (2018) superseding the group
ADI of 0 - 2 mg/kg bw for lutein from T. erecta L. and synthetic
zeaxanthin established at the 63rd JECFA (2004).
SYNONYMS INS No. 161b(i), Vegetable lutein; vegetable luteol; Bo-Xan,

luteine
DEFINITION Lutein from Tagetes erecta is a purified extract of xanthophylls

obtained from oleoresin in marigold. The oleoresin is prepared
from hexane extracts of Tagetes erecta L. flowers, saponified
with potassium hydroxide in either methanol or propylene
glycol. The resulting reaction mixture is diluted with water and
dried. The crystalline product contains lutein along with minor
components that include other carotenoids and waxes.
Chemical names 3R,3’R,6’R-β,ε-carotene-3,3'-diol; all-trans-lutein; 4’,5’-

didehydro-5’,6’-dihydro-beta,beta-carotene-3,3’-diol
C.A.S. number 127-40-2
Chemical formula C40H56O2
Structural formula
Assay Not less than 80% total carotenoids, not less than 70% lutein
DESCRIPTION A free-flowing, orange-red powder
FUNCTIONAL USES Colour, nutrient supplement

CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Insoluble in water, soluble in hexane
Spectrophotometry A 2 mg/l solution in acetone shows maximum absorbance at

(Vol. 4) approximately 446 nm.
Test for carotenoids The colour of 2 ml of a 2 – 4 mg/l solution of the sample in

(Vol. 4) acetone immediately disappears after successive addition of
about 0.5 ml of 5% sodium nitrite and about 0.5 ml of 0.5 M
sulfuric acid.
PURITY
Moisture (Vol. 4) Not more than 1.0%
Ash (Vol. 4) Not more than 1.0%
Zeaxanthin Not more than 9.0%
See description under METHOD OF ASSAY
Lead (Vol. 4) Not more than 3 mg/kg.

preparation may be based on the principles of the methods
described in Vol. 4 (under “General Methods, Metallic
Impurities”).
Hexane (Vol. 4) Not more than 50 mg/kg
Methanol (Vol. 4) Not more than 10 mg/kg
Propylene glycol Not more than 1000 mg/kg
Waxes Not more than 14.0%

TESTS
PURITY TESTS
Propylene glycol Determine by gas chromatography (Vol. 4) under the following

conditions.
Internal standard solution

Prepare a 500 μg/ml solution of ethylene glycol in
tetrahydrofuran.
Standard solutions
Prepare a range of standard solutions containing 1, 5,
10, 25 and 50 μg/ml of propylene glycol and 5 μg/ml of
ethylene glycol in tetrahydrofuran.
Chromatography conditions
- Column: Polydimethylsiloxane (30 m x 0.32 mm i.d.
with 0.25 μm film)
- Carrier gas: Helium
- Flow rate: 1.5 ml/min (Constant flow)
- Detector: FID
- Temperatures: injection port: 230°
- Column Temperature: Hold for 3 min at 40°, then 40-
250° at 20°/min, hold for 5 min at 250°
- Detector Temperature: 270°
The retention times of ethylene glycol and propylene glycol

derivatives under the above conditions are approx. 7.6 min
and
7.8 min, respectively.
Procedure
Weigh accurately 1 g of the sample into a 10-ml
volumetric flask, and add 100 μl of the internal
standard solution. Dissolve and make up to volume
with tetrahydrofuran. Take 0.5 ml of the sample
solution in a centrifugation tube, and add 0.25 ml of
1,1,1,3,3,3-hexamethyldisilazane (HMDS) and 0.1 ml
of trimethylchlorosilane (TMCS). After sealing the tube,
shake it vigorously, let stand for 30 min at room
temperature, then centrifuge. Inject 1.0 μl of this
centrifugal supernatant into the chromatograph.
Standard curve
Prepare following the same procedure using 0.5 ml of
the standard solutions in place of the sample solution.
Calculate the concentration of propylene glycol in mg/kg (CPG)

from:
CPG (mg/kg) = C × 10 / W
where
C is polyethylene glycol concentration determined (µg/ml); and
W is weight of sample (g)
Waxes Determine by gas chromatography (Vol. 4) using the following

conditions:
- GC column DB-5 (30 m x 0.25 mm ID with a 0.25 μm
film thickness) or equivalent.
- GC injector temperature: 280°
- FID temperature: 350°
- GC temperature program: 50° (2 min) 13°/min to 340°
and hold for 8 min
- Carrier gas (Helium) flow rate: 1.0 ml/min
- Injection mode: splitless
- Injection volume: 1.0 μl
Standards:
- Hydrocarbons mixed standard: C25 to C46
- Internal standard: Hexatriancontane (C36)
Standard solutions:
Prepare standard solutions by addition of hydrocarbon
standards to methylene chloride to get hydrocarbon
concentrations of 2.0, 5.0, 10, 25, 50, mg/l respectively.
Add required quantity of hexatriancontane internal
standard to get a final concentration 50 mg/l in all
standard solutions.
Sample Preparation
Accurately weigh 100 mg of sample into a centrifuge
tube and dissolve in exactly 20 ml of methylene
chloride. Sonication or vortex mixing may be required
to completely dissolve the product. Centrifuge sample
at 2500 rpm for 5 min, if the sample appears turbid.
Add 1.6 ml of methylene chloride and 20 μl of
(5000 mg/l) hexatriancontane solution (to a final
concentration of 50 mg/l) into 2 ml volumetric flask.
Transfer 40 μl of sample solution and dilute with
methylene chloride to the 2 ml. Transfer the solution
into a 2 ml autosampler vial.
Analysis
Inject 1.0 μl of each of the standards solutions. Record
the peak areas. Construct standard curves using the
peak ratios of each hydrocarbon to the internal
standard against the concentration of the hydrocarbon.
Inject 1.0 μl of the sample solution and determine
individual wax in the sample (mg/l) from the respective
standard curve.
Add the concentration of individual waxes to get the
total wax concentration in the sample solution (mg/l)
Calculation:
C (mg/l) × 2 ml × 20 ml × 100
Waxes % w/w =
1000 (ml/l) × W (mg) × 0.04 ml
= (100 × C)/W
Where:
C is the total concentration of waxes, mg/l in the sample
W is the weight of sample, mg
METHOD OF ASSAY Determine the total carotenoid content and the content of lutein
and zeaxanthin by UV spectrophotometry and HPLC using the
following conditions:
Reagents:
- Hexane (HPLC grade)
- Ethyl acetate (HPLC grade)
- Acetone
- Dehydrated ethyl alcohol (absolute alcohol)
- Toluene
- Solvent Mixture: (10:6:7:7 hexane:dehydrated ethyl
alcohol:acetone:toluene, v/v/v/v)
System Suitability Solution for HPLC:

150 µg/ml of lutein standard in solvent mixture (use
USP Lutein RS available from U. S. Pharmacopeia, or
equivalent standard)
Apparatus
UV/Vis spectrophotometer; 1-cm cuvettes
HPLC system with suitable diode array detector,
autosampler, column oven, signal processor and
degasser.
Analytical column: 3 µm silica, 4.6 mm x 250 mm
Instrument Conditions
- Temperature: ambient
- Mobile Phase: 70:30 (v:v) hexane/ethyl acetate
(isocratic elution)
- Flow Rate: 1.5 ml/min
- Injection: 10 µl
- Detection: UV/Vis 446 nm
- Run Time: approximately 40 min
Concentrated Sample Preparation

For the UV/Vis spectrophotometry weigh sample
(30 mg) into a glass weighing funnel. Using the solvent
mixture, wash crystals into a 100 ml volumetric flask,
dilute to the mark with the solvent mixture and stir for
10 min.
Sample Preparation
Pipette 1 ml of concentrated sample preparation into a
100 ml volumetric flask. Dilute up to the mark with

dehydrated ethyl alcohol, mix by inversion for
20 seconds. Read samples in a spectrophotometer at
446 nm using dehydrated ethyl alcohol as the blank.
For HPLC, evaporate 1 ml of the concentrated sample

preparation to dryness using a stream of nitrogen,
dissolve solids in 1 ml 70:30 hexane:ethyl acetate, and
add 0.5 ml to HPLC vials. Analyze this sample and the
system suitability solution for HPLC using the HPLC
conditions above.
Results
Compare the results of the chromatogram from the
system suitability solution for HPLC to identify the lutein
and zeaxanthin peaks at a resolution of not less than 3.
Calculation
Using the results obtained from the UV/Vis
spectrophotometry calculate the % Total carotenoids
Absorbance at 446 nm × 10000 × 100

% Total carotenoids =
sample weight in g × 2550
Note: The factors 10000 and 2550 are the dilution factor and
extinction value for a 1% solution, respectively.
Using the chromatogram of the sample, calculate the

concentration of lutein and zeaxanthin.
𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿

𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿 (%) = % 𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 ×
𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇
𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍ℎ𝑖𝑖𝑖𝑖 (%) = % 𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 ×

𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍𝑍
𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇
MODIFIED STARCHES

Monograph 22 (2018), superseding specifications included in 16
individual specification monographs prepared at the 82nd JECFA
(2016), published in FAO JECFA Monographs 19 (2016).
MODULAR MONOGRAPH consisting of "GENERAL

SPECIFICATIONS"(a) that contains common specifications to all
modified starches(INS 1400, 1401, 1402, 1403, 1404, 1405, 1410,
1412, 1413, 1414, 1420, 1422, 1440, 1442, 1450, 1451), and 8
ANNEXES that contain specifications related to the chemical treatments
of native starches:
- ANNEX 1(a) – Fragmentation.

- ANNEX 2(a) – Bleaching.
- ANNEX 3(a) – Esterification and/or crosslinking with
phosphorous containing compounds.
- ANNEX 4(a) – Acetylation.
- ANNEX 5(a) – Oxidation.
- ANNEX 6(a) – Esterification with octenyl succinic anhydride.
- ANNEX 7(a) – Etherification with propylene epoxide.
- ANNEX 8(a) – Esterification and crosslinking with adipic
anhydride.
The General specifications are applicable to the following modified

starches, each of which should additionally fulfil the specifications of
the ANNEXES as follows:
Modified INS Annex ADI STATUS

Starch
Dextrin roasted 1400 1 N.S.(1) T(3)
starch
Acid treated 1401 1 N.S.(1) T(3)
starch
Alkaline treated 1402 1 N.S.(1) T(3)
starch
Bleached starch 1403 2 N.S.(1) T(3)
Oxidized starch 1404 5 N.S.(1) T(3)
Enzyme-treated 1405 1 N.S.(1) T(3)
starch
Monostarch 1410 3 N.S.(1) T(3)
phosphate
Distarch 1412 3 N.S.(1) T(3)
phosphate
Phosphated 1413 3 N.S.(1) T(3)
distarch
phosphate
Acetylated 1414 3, 4 N.S.(1) T(3)
distarch
phosphate
Starch acetate 1420 4 N.S.(1) T(3)

Acetylated 1422 4, 8 N.S.(1) T(3)
distarch adipate
Hydroxypropyl 1440 7 N.S.(1) T(3)
starch
Hydroxypropyl 1442 3, 7 N.S.(1) T(3)
distarch
phosphate
Starch sodium 1450 6 N.S.(1) T(3)
octenylsuccinat
e
Acetylated 1451 4, 5 N.S.(2) T(3)
oxidized starch
Should any of the modified starches be subjected to additional chemical
treatment, the appropriate specifications outlined in the respective
ANNEX should be met. Consequently, for all fragmented and/or
bleached starches the specifications of ANNEXES 1 and/or 2
respectively should be met.
(a)
Monograph 22 (2018), superseding specifications included in
specification monographs prepared at the 82nd JECFA (2016),
published in FAO JECFA Monographs 19(2016).
(1)
An ADI “not specified” was established at the 26th JECFA (1982).
(2)
An ADI “not specified” was established at the 57th JECFA (2001).
(3)
T: TENTATIVE
Summary Table
GENERAL REQUIREMENTS
IDENTIFICATION PURITY
Solubility Microscopy Iodine Stain Copper Reduction pH Loss on Drying Lead Microbiological Criteria Sulfur dioxide
≤0.2mg/k ≤50 mg/kg d.w.
Granular Red precipitate after g d.w. Aerobic Plate Count: ≤1000 for modified
structure Colour from dark blue addition of hot alkaline Cereal starch ≤15.0%; Pb (≤0.1 CFU/g; Yeasts and molds: cereal
Insoluble in cold
typical of to red after addition of cupric tartrate to a test 3.0 -9.0 Potato starch: ≤21.0%; mg/kg) for ≤1000 CFU/g; Total Coliforms: starches; ≤10
water
the starch tri-iodide sample refluxed under Other starches: ≤18.0% OSA for ≤10 cfu/g; Information mg/kg d.w. for
source acidic condition infant required. other modified
formula starches
SPECIFIC REQUIREMENTS
Modified Starch Annex IDENTIFICATION PURITY
Dextrin roasted Dispersion index (Information Required); Reducing sugars
1 No additional
(INS 1400) (Information Required)
Acid treated (INS Dispersion index (Information Required); Reducing sugars
1 No additional
1401) (Information Required)

Alkaline treated Dispersion index (Information Required); Reducing sugars
1 No additional
starch (INS 1402) (Information Required)
Bleached (INS
2 No additional Carboxyl groups (≤0.1% d.w.);Residual oxidising substances (Information Required)
1403)
Oxidized (INS
5 hypochlorite oxidized starch Carboxyl groups (≤1.3% d.w.);Residual hypochlorite (Information Required)
1404)
Enzyme-treated Dispersion index (Information Required); Reducing sugars
1 No additional
(INS1405) (Information Required)
Monostarch
phosphate (INS 3 Information required Phosphate (≤0.5% d.w. for potato or wheat starches; ≤0.4% d.wfor other starches)
1410)
Distarch
phosphate (INS 3 Information required Phosphate (≤0.5% d.w. for potato or wheat starches; ≤0.4% d.w. for other starches)
1412)
Phosphated
distarch phosphate 3 Information required Phosphate (≤0.5% d.w. for potato or wheat starches; ≤0.4% d.w. for other starches)
(INS 1413)
Acetylated distarch
Phosphate (≤0.14% d.w. for potato or wheat starches; ≤0.04% d.w. for other starches)
phosphate (INS 3, 4 Acetyl group; Ester group; Information required
Acetyl groups (≤2.5%d.w.); Ester groups (≤0.5%d.w.)
1414)
Starch acetate
4 Acetyl group; Ester group Acetyl groups (≤2.5%d.w.); Ester groups (≤0.5%d.w.)
(INS 1420)
Acetylated distarch Acetyl groups (≤2.5%d.w.); Vinyl acetate (≤0.1 mg/kg); Ester groups (≤0.5%d.w.)
4, 8 Acetyl group; Ester group; Information required
adipate (INS 1422) Adipate groups (≤0.135%d.w.); Residual free adipic acid (Information Required)
Hydroxypropyl
7 Hydroxypropyl ether groups Hydroxypropyl groups (≤7.0% d.w.); Propylene chlorohydrins (≤1 mg/kg d.w.)
starch (INS 1440)
Hydroxypropyl
Phosphate (≤0.14% d.w. for potato or wheat starches; ≤0.04% d.w. for other starches)
distarch phosphate 3, 7 Hydroxypropyl ether groups; Information required
Hydroxypropyl groups (≤7.0% d.w.); Propylene chlorohydrins (≤1 mg/kg d.w.)
(INS 1442)
Starch sodium
octenylsuccinate 6 No additional Octenylsuccinyl groups (≤3% d.w.); Residual free octenylsuccinic acid (≤0.3%d.w.);
55
(INS 1450)
Acetylated
Acetyl groups (≤2.5%d.w.); Vinyl acetate (≤0.1 mg/kg); Ester groups (≤0.5%d.w.)
oxidized starch 4, 5 Acetyl group
Carboxyl groups (≤1.3% d.w.);Residual hypochlorite (Information Required)
(INS 1451)
GENERAL SPECIFICATIONS FOR MODIFIED STARCHES
(VERSION 2018 - TENTATIVE)
Information required:
• Suitable microbiological acceptance criteria and supporting
data
DEFINITION Starch consists mainly of amylose and amylopectin. Amylose is a

linear molecule of α-D-glucopyranosyl units linked by (1-4)-α-
linkages. Amylopectin is a highly-branched polymer of α-D-
glucopyranosyl units linked by (1-4)-α-linkages and by(1-6)-α-
linkages that constitute the branch points. Each glucose unit
possesses a maximum of three hydroxyls that can undergo
chemical substitution.
Native starches can be physically (pre-gelatinized starches) and/or

chemically modified for improved functionality. The most common
sources of native starch used in these modifications are various
roots, tubers, cereals and legumes. Modified starches are used in
applications requiring special properties not attainable by native
starches.
Chemical modifications of native starches are often performed, in

an aqueous suspension under controlled conditions of pH, time
and temperature, unless otherwise indicated in the description of
the respective annex. After sufficient reaction time, the modified
starch is recovered by filtration or centrifugation, washed with
water, dried and packaged. The relevant modification reactions
can be, separately or in combination, fragmentations (hydrolysis,
oxidation, enzymatic), bleaching, oxidation, esterification,
etherification or phosphorylation of one or more of the hydroxyl
groups of the α-D-glucopyranosyl units or crosslinking using
polyfunctional agents.
See the appropriate Annex or Annexes for the treatment that is

applicable to individual modified starch products.
C.A.S numbers See ANNEXES
DESCRIPTION White or nearly white powder or granules or (if pre-gelatinized)

flakes, or amorphous powder or coarse particles.
FUNCTIONAL USES Thickener, stabilizer, binder, emulsifier

CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Insoluble in cold water (if not pre-gelatinised); forming typical
colloidal solutions with viscous properties in hot water; insoluble in
ethanol.
Microscopy Passes test
Iodine stain Passes test
Copper reduction Passes test
PURITY
General Requirements:
pH 3.0 – 9.0
Loss on drying (Vol 4) Cereal starch: not more than 15.0%
Potato starch: not more than 21.0%
Other starches: not more than 18.0%
Conditions: 120°, 4 h, vacuum not exceeding 100 mm Hg
Lead (Vol. 4) Not more than 0.2 mg/kg on the dried basis
Not more than 0.1 mg/kg on the dried basis for Starch sodium
octenylsuccinate (INS 1450) for use in infant formula and formula for
special medical purposes intended for infants(see Annex 6)

selection of sample size and method of sample preparation may be
based on principles of methods described in Volume 4 (under
Microbiological Criteria Aerobic plate count: Not more than 1000 CFU/g
(Vol 4)
Yeasts and moulds: Not more than 1000 CFU/g
Total coliforms: Not more than 10 CFU/g
Information required
Sulphur dioxide (Vol. 4) Not more than 50 mg/kg on the dried basis for modified cereal
starches
Not more than 10 mg/kg on the dried basis for other modified
starches
TESTS
IDENTIFICATION
TESTS
Microscopy Each modified starch, which has not been pre-gelatinized, retains its
granular structure and can be identified as a starch by microscopic
observation. The typical polarization cross is observed when sample
is examined with a polarizing microscope, in polarized light under
crossed Nicol prisms.
Corn starch: Polygonal, rounded or spherical granules up to 35 µm

diameter having a circular or several-rayed central cleft.
Potato starch: Irregular shaped, ovoid, pear-shaped granules (30-

100 µm diameter, occasionally >100 µm); both, the ovoid, the pear-
shaped granules and the rounded granules have an eccentric hilum.
All granules show clearly visible concentric striations.
Tapioca starch: Spherical granules with one truncated side (5-35 µm

diameter) usually having a circular or several-rayed central cleft.
Wheat starch: large and small granules (10-60 µm diameter). The

central hilum and striations are visible and barely visible.
Iodine stain Add a few drops of 0.1 N potassium triiodide to an aqueous

suspension of the sample. The modified starch stains with iodine in
the same way as native starches. The colour can range from dark
blue to red.
Copper reduction Place about 2.5 g of the sample previously washed with water, in a
boiling flask; add 10 ml of dilute hydrochloric acid (3%) and 70 ml of
water; mix, reflux for about three hours and cool. Add 0.5 ml of the
resulting solution to 5 ml of hot alkaline cupric tartrate TS. A copious
red precipitate is produced.
PURITY TESTS
pH (Vol. 4) Suspend 20 g of the sample with 80 ml of water, and agitate

continuously at a moderate rate for 5 min (In the case of pre-
gelatinised starches, 3 g should be suspended in 97 ml of water).
ANNEX 1: ADDITIONAL SPECIFICATIONS FOR STARCHES MODIFIED BY

FRAGMENTATION
Information is required on:
• A suitable method for dispersion and a method for

reducing sugars and data on at least 5 representative
batches using the method(s) from each of the
fragmentation processes
APPLIES TO Dextrin roasted starch (INS No. 1400)
Acid treated starch (INS No. 1401)
Alkaline treated starch (INS No. 1402)
Enzyme-treated starch (INS No. 1405)
All modified starches that are fragmented
SYNONYMS Modified starch by fragmentation, converted starch,

hydrolysed starch.
TREATMENT The fragmentation of native starch results in products

containing polymers with a lower average molecular weight
and reduced viscosity. The manufacturing details for the
various modified starches by fragmentation in this
monograph are described as below:
- Dextrin roasted starch, INS. 1400: is manufactured by
dry heating or roasting of native starch with
hydrochloric acid or ortho-phosphoric acid in heated
and/or agitated vessels. The final dextrin roasted
starch is obtained by drying.
- Acid treated starch, INS. 1401 is obtained by treating
a slurry or a suspension of native food starch with
dilute hydrochloric acid, ortho-phosphoric acid, or
sulphuric acid.
- Alkaline treated starch, INS. 1402 is obtained by
treating a suspended solution of native food starches
with sodium hydroxide or potassium hydroxide.
- Enzyme-treated starch, INS 1405 is obtained by
treating a suspension of native food starch with one
or more food-grade amyolytic-enzymes (e.g., α-
amylase (E.C. 3.2.1.1), β-amylase (3.2.1.2),
glucoamylase (3.2.1.3), isoamylase (3.2.1.68),
pullulanase (E.C. 3.2.1.41)).
The properties of the modified starches by fragmentation

vary depending on the source of native starch, reaction
conditions (pH, reaction time, reaction temperature,
fragmenting reagent etc.) The alteration of native starch
allows for applications that require reduced viscosity in hot
solutions and/or typically utilise high levels of modified
starches.
C.A.S number 9004-53-9 (Dextrins)
65996-63-6 (Acid-hydrolysed starch)
68909-37-5 (Acid-hydrolysed amylopectin)
9005-84-9 (Starch soluble)
65996-64-7 (Enzyme-hydrolysed starch)
1001439-91-3 (Enzyme-treated amylopectin).
CHARACTERISTICS
IDENTITY
Dispersion identity Information required.
Reducing sugars Information required
TESTS
IDENTIFICATION
TESTS
Dispersion test Information required
Reducing sugars Information required

ANNEX 2: ADDITIONAL SPECIFICATIONS FOR BLEACHED STARCHES

• Suitable method(s) for the determination of residual
reagents and data on at least 5 representative
batches using the method(s).
APPLIES TO Bleached starch INS No. 1403
All modified starches that are bleached
TREATMENT Peracetic acid and/or hydrogen peroxide, or sodium

hypochlorite, sodium chlorite, sulfur dioxide, alternative
permitted forms of sulphites, potassium permanganate, or
ammonium persulfate
Bleaching is performed to improve physical attributes such as

colour due to oxidation of traces of pigments such as
carotenoids and xanthophylls. The change is essentially in the
colour only. Residual reagents are either removed or limited to
technically unavoidable levels.
C.A.S number 977075-42-5
and all other modified starches submitted to bleaching
CHARACTERISTICS
PURITY
Manganese (Vol. 4) Not more than 50 mg/kg on the dried basis

The selection of sample size and method of sample preparation
may be based on principles of methods described in Volume 4
(under “General Methods, Metallic Impurities”).
Residual oxidising Information required

substances
Carboxyl groups (Vol. Not more than 0.1% on the dried basis applying the correction
4) for phosphate content as outlined in Note 6 of the method for
starches esterified with phosphorus containing compounds.
ANNEX 3: ADDITIONAL SPECIFICATIONS FOR STARCHES ESTERIFIED

AND/OR CROSSLINKED WITH PHOSPHORUS CONTAINING COMPOUNDS
Information required on: A suitable method for identification of

crosslinking and data on at least 5 representative batches of
crosslinked and non-crosslinked starches.
APPLIES TO Monostarch phosphate (INS No. 1410)
Distarch phosphate (INS No. 1412)
Phosphated distarch phosphate (INS No. 1413)
Acetylated distarch phosphate (INS No. 1414)
Hydroxypropyl distarch phosphate (INS No. 1442)
TREATMENT The phosphorus containing compounds ortho-phosphoric

acid, sodium or potassium ortho-phosphate and sodium
tripolyphosphate, can be used for esterification and the
sodium trimetaphosphate or phosphorus oxychloride for
crosslinking.
- Monostarch phosphate (INS 1410) is obtained by
esterification/crosslinking of unmodified food starch
with ortho-phosphoric acid, or sodium or potassium
ortho-phosphate, or sodium tripolyphosphate
- Distarch phosphate (INS 1412) is obtained by
crosslinking of unmodified food starch with sodium
trimetaphosphate or phosphorus oxychloride
- Phosphated distarch phosphate (INS 1413) is
obtained by esterification/crosslinking of unmodified
food starch with sodium trimetaphosphate or
phosphorus oxychloride combined with esterification
with ortho-phosphoric acid, or sodium or potassium
ortho-phosphate, or sodium tripolyphosphate
- Acetylated distarch phosphate (NS 1414) is obtained
by esterification/crosslinking of unmodified food
starch with sodium trimetaphosphate or phosphorus
oxychloride combined with esterification with acetic
anhydride or vinyl acetate
- Hydroxypropyl distarch phosphate (INS1442) is
obtained by esterification of unmodified food starch
with sodium trimetaphosphate or phosphorus
oxychloride combined with etherification by
propylene oxide
Phosphorylation results in partial substitution of the 2, 3- or

6- position of the anhydro glucose unit unless the 6-position
is occupied for branching. In the case of cross-linking, where
a polyfunctional substituting agent, such as phosphorus
oxychloride, connects two chains, the structure can be
represented by: Starch-O-R-O-Starch, where R = cross-

linking group and Starch refers to the linear and/or branched
structure.
C.A.S numbers Monostarch phosphate (INS 1410)

11120-02-8(Modified starch)
63055-37-8 (Modified amylopectin)
Distarch phosphate (INS No. 1412)

Phosphated distarch phosphate (INS No. 1413)

Acetylated distarch phosphate (INS No. 1414)


CHARACTERISTICS
PURITY
Phosphate For monostarch phosphate (INS No. 1410), distarch

(calculated as phosphate (INS No. 1412), and phosphate distarch
phosphorus) (Vol. 4) phosphate (INS No. 1413)
Not more than 0.5% on the dried basis for potato or

wheat starches
Not more than 0.4% on the dried basis for other

starches
For acetylated distarch phosphate (INS No. 1414) and

hydroxypropyl distarch phosphate (INS No. 1442)
Not more than 0.14% on the dried basis for potato

and wheat starch
Not more than 0.04% on the dried basis for other

starches
IDENTITY
Crosslinking Information Required

ANNEX 4: ADDITIONAL SPECIFICATIONS FOR ACETYLATED STARCHES
Version 2018
APPLIES TO Acetylated distarch phosphate (INS No. 1414)
Starch acetate (INS No. 1420)
Acetylated distarch adipate (INS No. 1422)
Acetylated oxidized starch (INS No. 1451)
TREATMENT This type of modified starch is obtained by esterification

with acetic anhydride or vinyl acetate. Acetylation results in
substitution of hydroxyl groups with acetyl esters.
- Acetylated distarch phosphate (INS 1414) is

obtained by esterification/cross-linking of
unmodified food starch with sodium
trimetaphosphate or phosphorus oxychloride
combined with esterification with acetic anhydride
or vinyl acetate.
- Starch acetate (INS 1420) is obtained by
esterification of food starches with acetic anhydride
or vinyl acetate
- Acetylated distarch adipate (INS 1422) is obtained
by esterification of unmodified food starch with
acetic anhydride and esterification/cross-linking
with adipic anhydride
- Acetylated oxidized starch (INS 1451) is obtained
by treatment of food starch with sodium
hypochlorite followed by esterification with acetic
anhydride
C.A.S numbers Acetylated distarch phosphate (INS No. 1414)

Starch acetate (INS No. 1420)

9045-28-7 (Modified starch)
Acetylated distarch adipate (INS No. 1422)


CHARACTERISTICS
IDENTIFICATION
Specific reaction for Passes TEST

Acetyl groups
Ester groups Passes TEST
PURITY
Acetyl groups Not more than 2.5% on the dried basis
Vinyl acetate Not more than 0.1 mg/kg
TEST
IDENTIFICATION
TESTS
Specific reaction for Principle

acetyl groups Acetate is liberated upon saponification of acetylated
starch and converted to acetone by heating with
calcium hydroxide. The acetone thus produced stains
blue with o-nitrobenzaldehyde.
Procedure
Suspend about 10 g of the sample in 25 ml water. Add
20 ml of 0.4 M NaOH. After shaking for 1 h filter the
starch off and evaporate the filtrate in an oven at 110°.
Dissolve the residue in a few drops of water and
transfer to a test tube. Add calcium hydroxide and
heat the tube. If the sample is acetylated starch,
acetone vapours are produced. These produce a blue
colour on a paper strip soaked in a fresh saturated
solution of o-nitrobenzaldehyde in 2 M NaOH. The
blue colour is more distinct when the original yellow
colour of the reagents is removed with 1 drop of a 1 in
10 solution of hydrochloric acid.
Ester groups The infrared spectrum of a thin film gives a typical

absorption band at about 1720 cm-1 which is an indication
for ester groups. The limit of detection is about 0.5% acetyl

groups in the product.
PURITY TESTS
Acetyl groups Accurately weigh about 5 g of the sample and transfer into a
250 ml conical flask. Suspend in 50 ml of water, add a few
drops of phenolphthalein TS, and titrate with 0.1 M sodium
hydroxide to a permanent pink end-point. Add 25.0 ml of
0.45 M sodium hydroxide, stopper the flask, and shake
vigorously for 30 min, preferably with a mechanical shaker.
(NOTE: the temperature should not exceed 30o as some
starches may gelatinise). Remove the stopper, wash the
stopper and sides of the flask with a few ml of water, and
titrate the excess alkali with 0.2 M hydrochloric acid to the
disappearance of the pink colour. Record the volume, in ml of
0.2 M hydrochloric acid required as S.
Perform a blank titration on 25.0 ml of 0.45 M sodium

hydroxide, and record the volume, in ml, of 0.2 M hydrochloric
acid required as B.
(B − S) × M × 0.043 × 100
Acetyl groups % =
W
where
M is the molarity of hydrochloric acid solution; and
W is the weight of sample, in grams.
ANNEX 5: ADDITIONAL SPECIFICATIONS FOR STARCHES SUBJECTED TO

OXIDATION

• A suitable method for determination of residual
hypochlorite and data on at least 5 representative
batches using the method.
APPLIES TO Oxidized starch (INS No. 1404)
TREATMENT Sodium hypochlorite is used for oxidation.
• Oxidized starch (INS 1404) is obtained by treatment of

food starch with sodium hypochlorite.
• Acetylated oxidized starch (INS 1451) is obtained by

treatment of food starch with sodium hypochlorite
followed by esterification with acetic anhydride.
Oxidation involves the deliberate production of carboxyl

groups.
C.A.S number Oxidised starch (INS No. 1404)

65996-62-5 (modified starch)
113894-86-3 (modified amylopectin)
Acetylated oxidised starch (INS No. 1451)

68187-08-6
CHARACTERISTICS
IDENTIFICATION
Test for hypochlorite Passes test

oxidized starch
PURITY
Carboxyl groups Not more than 1.3% on the dried basis

(Vol. 4)
Residual hypochlorite Information required

TESTS
IDENTIFICATION
TESTS
Test for hypochlorite Principle

oxidized starch Because of the carboxyl group content, hypochlorite-
oxidized starch has anionic properties. It can be dyed
with positively charged dyes such as methylene blue.
The test is not suitable for slightly oxidized potato starch
due to the presence of phosphate groups.
Procedure
50 mg of the sample are kept in suspension for 5-
10 min in 25 ml of a 1% aqueous dye solution and
stirred occasionally. After decantation of the excess
solution, the starch is washed with distilled water.
Microscopic inspection clearly shows colouring, if the
sample is hypochlorite-oxidized starch. By this test
hypochlorite-oxidized starch is distinguished from native
and acid modified starch of the same botanical origin.
ANNEX 6: ADDITIONAL SPECIFICATIONS FOR STARCHES ESTERIFIED WITH

OCTENYLSUCCINIC ANHYDRIDE
Version 2018
APPLIES TO Starch sodium octenylsuccinate (INS No. 1450)
TREATMENT Octenylsuccinic anhydride can be used for the esterification

and either sodium hydroxide or sodium carbonate as a pH
buffer for neutralisation.
C.A.S numbers Starch sodium octenylsuccinate

CHARACTERISTICS
PURITY
Octenylsuccinyl Not more than 3% on the dried basis

groups
Residual free Not more than 0.3% on the dried basis

octenylsuccinic acid
PURITY TEST
Octenylsuccinate Principle
groups and residual Residual free octenylsuccinic acid in the sample is extracted
free octenylsuccinic and determined by HPLC/UV. Total octenylsuccinic content is
acid in Starch sodium determined using the same method after hydrolysis of the
octenyl succinate sample. Octenylsuccinate ester groups on the modified starch
are calculated by subtraction of the residual free
octenylsuccinic acid from the total.
Standard and Reagents
Octenylsuccinic anhydride:
2-Octen-1-ylsuccinic anhydride, mixture of cis and trans
(>97%) (CAS: 42482-06-4)
0.1 N potassium hydroxide:

Weigh1.4 g of potassium hydroxide, dissolve in water
and dilute to 250 ml
0.073 mol/l phosphoric acid:

Dilute 1 ml of phosphoric acid (85%, density
1.686g/cm3) to 200 ml with water.
Preparation of standard solutions

Accurately weigh about 20 mg of octenylsuccinic
anhydride, add 10 ml of 0.1N potassium hydroxide,
stopper and heat at 80° for 3 hours. After cooling, add
8 ml of 0.073mol/l phosphoric acid and dilute with water
to 20 ml. Pipette 2 ml of this solution into a 20 ml
volumetric flask and dilute with water. Pipette 1 ml,
2 ml, 5 ml, and 10 ml of the resulting solution into four
separate 20-ml volumetric flasks, and dilute each to
volume with water to prepare standards of 5 μg/ml,
10 μg/ml, 25 μg/ml and 50 μg/ml respectively.
Preparation of test solution A (for residual octenylsuccinic

acid):
Accurately weigh about 0.1 g of sample, add 20 ml of
methanol, and shake for 18 hours or more. Centrifuge
the mixture at about 3000 rpm for 5 minutes, pipette
10 ml of the supernatant, and evaporate to dryness
under vacuum at 40°. Dissolve the residue and dilute
with water in a 5 ml volumetric flask.
Preparation of test solution B (for total octenylsuccinic acid):

Accurately weigh about 20 mg of sample, dissolve in
10 mL of 0.1N potassium hydroxide, stopper and heat
at 80°for 3 hours. After cooling, add 8 ml of 0.05mol/l
phosphoric acid, dilute with water to 20 ml.
Procedure
HPLC operating conditions
- Column: A octadecylsilanized silica gel column
(250 mm x 4.6 mm, 5µm) (L-Column ODS-V CERI or
equivalent)
- Detector: UV at 205 nm
- Mobile phase: A 1:1 mixture of 0.1% (v/v) phosphoric
acid solution / acetonitrile
- Injection volume: 20µl
- Flow rate: Adjust the retention time of the main peak to
about 9 minutes.
Inject the test solution A and B and the standard solutions into
an HPLC under the same conditions.
Measure the sum of the peak areas of two main peaks of cis-
and trans-2-octenylsuccinic acid for each standard solution,
and prepare a standard curve for octenylsuccinic anhydride
from the sums obtained and the concentrations of
octenylsuccinic anhydride in the standard solutions. Measure
the sum of the peak areas of two main peaks for the test
solutions A and B. Determine the concentration of
octenylsuccinic acid (µg/ml) in the test solutions A and B from

the standard curve, and calculate residual and total
octenylsuccinic acid, respectively. The value of
octenylsuccinate groups in the sample is calculated by the
following formula:
Calculation:
𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅𝑅 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 %

= 𝐶𝐶𝑂𝑂𝑂𝑂 𝑥𝑥 1.086 / 𝑊𝑊𝑟𝑟 𝑥𝑥 100
𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 % = 𝐶𝐶𝑜𝑜𝑜𝑜 𝑥𝑥 1.086 / 𝑊𝑊𝑠𝑠 𝑥𝑥 500
Where
- 1.086 is the molecular weight of octenylsuccinic acid
divided by the molecular weight of octenylsuccinic
anhydride
- Cos is the octenylsuccinic anhydride concentration
(µg/ml);
- Wr or W s is the dry-basis weight of the sample (g).
Content (%) of octenylsuccinyl groups =

Content of total octenyl succinic acid – Content of
residual octenylsuccinic acid.
ANNEX 7: ADDITIONAL SPECIFICATIONS FOR STARCHES ETHERIFIED WITH

PROPYLENE OXIDE
(Version 2018 - TENTATIVE)

• A suitable method for the determination of propylene
chlorohydrin with detection limit lower than 0.1 mg/kg
and data on at least 5 representative batches of
Hydroxypropyl starch using the method.
APPLIES TO Hydroxypropyl starch (INS No. 1440)
TREATMENT Propylene oxide is used for etherification.
• Hydroxypropyl starch (INS No. 1440) is obtained by

etherification of unmodified food starch with propylene
oxide.
• Hydroxypropyl distarch phosphate (INS No. 1442) is
obtained by esterification of unmodified food starch with
sodium trimetaphosphate or phosphorus oxychloride
combined with etherification by propylene oxide.
Hydroxypropylation results in substitution of hydroxyl groups

with 2-hydroxypropyl ether.
C.A.S numbers Hydroxypropyl starch(INS No. 1440)


CHARACTERISTICS
IDENTIFICATION
Hydroxypropyl ether Passes test

groups
PURITY
Hydroxypropyl groups Not more than 7.0% on the dried basis

Propylene Not more than 1 mg/kg

chlorohydrins
TESTS
IDENTIFICATION
TESTS
Hydroxypropyl ether Ninhydrin reagent

groups 2 A 3% solution of 1,2,3-triketohydrindene crystals in 5%
aqueous sodium bisulfite solution.
Procedure
Weigh 100 mg of the sample into a 100-ml volumetric
flask and add 12.5 ml of 2 N sulfuric acid. Prepare a
sample of unmodified starch of the same source (i.e.
corn or potato) in the same manner. Place the flasks in
a boiling water bath and heat until the samples are in
solution. Cool and dilute the contents to 100 ml with
water. Pipet 1 ml of the solutions into 25-ml graduated
test tubes with glass stoppers and, with the tubes
immersed in cold water, add dropwise 8 ml of
concentrated sulfuric acid to each. Mix well and place
the tubes in a boiling water bath for exactly 3 min.
Immediately transfer the tubes to an ice bath until the
solution is chilled. Add 0.6 ml of ninhydrin reagent,
carefully allowing the reagent to run down the walls of
the test tubes. Immediately shake well, and place the
tubes in a 25° water bath for 100 min. Adjust the
volume in each tube to 25 ml with concentrated sulfuric
acid and mix by inverting the tubes several times. (Do
not shake).
A violet colour develops only in the modified sample

within 5 min due to the presence of hydroxypropyl
groups (starch ether).For all other non-hydroxypropyl
treated starches a light pink colour is observed.
PURITY TEST
Hydroxypropyl groups Ninhydrin reagent

A 3% solution of 1,2,3-triketohydrindene crystals in 5%
aqueous sodium bisulfite solution.
Procedure
Accurately weigh 50 - 100 mg of the sample into a
100-ml volumetric flask and add 25 ml of 1 N sulfuric
acid. Prepare a sample of unmodified starch of the
2 USP29-NF34: U.S. Pharmacopeial Convention, Hydroxylpropyl corn starch monograph, 2015.
Reproduced from the USP-NF with permission from The U.S. Pharmacopeial Convention (USP)
same source (i.e. corn or potato) in the same manner.

Place the flasks in a boiling water bath and heat until
the samples are in solution. Cool and dilute the
contents to 100 ml with water. If necessary, dilute the
sample further to assure the presence of no more than
4 mg of hydroxypropyl group per 100 ml, and then dilute
the blank starch in the same proportion. Pipet 1 ml of
the solutions into 25-ml graduated test tubes with glass
stoppers and, with the tubes immersed in cold water,
add dropwise 8 ml of concentrated sulfuric acid to each.
Mix well and place the tubes in a boiling water bath for
exactly 3 min. Immediately transfer the tubes to an ice
bath until the solution is chilled. Add 0.6 ml of ninhydrin
reagent, carefully allowing the reagent to run down the
walls of the test tubes. Immediately shake well, and
place the tubes in a 25° water bath for 100 min. Adjust
the volume in each tube to 25 ml with concentrated
sulfuric acid and mix by inverting the tubes several
times. (Do not shake). Immediately transfer portions of
the solutions to 1-cm cells and after exactly 5 min,
measure the absorption (A) at 590 nm, using the starch
blank as the reference. Prepare a calibration curve with
1-ml aliquots of standard aqueous solutions, containing
10, 20, 30, 40 and 50 µg of propylene glycol per ml.
Calculation
𝐶𝐶 × 0.7763 × 10 × 𝐹𝐹
𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔 (%) =
𝑊𝑊
where
- C is the amount of propylene glycol in the sample
solution read from the calibration curve (µg/ml);
- F is the dilution factor (if a further dilution has been
necessary); and
- W is the weight of sample (mg).
Propylene Principle
chlorohydrins Propylene chlorohydrins (1-chloro-2-propanol and 2-
chloro-1-propanol) in sample are determined by
capillary gas chromatography.
Standards and Reagents

propylene chlorohydrins
Preparation of standard addition calibration curve

Accurately weigh about 50 mg propylene
chlorohydrins, and dilute with water to 100 ml. Dilute
10 ml of this solution to 100 ml with water to make a
standard stock solution (50 μg/ml). Take four
Erlenmeyer flasks and weigh 50.0 g of sample in
each one. Add 125 ml of 1 M sulfuric acid in each
one. Add 0.5 ml, 1 ml, 2 ml, or 5 ml of standard stock
solution, to the 1st, 2nd, etc flasks respectively.
Proceed as directed for the test solution, beginning
with “and swirl the flask to disperse the contents,” to

prepare the standard solutions of added
concentration 5, 10, 20 and 50 µg/ml respectively.
Preparation of test solution

Accurately weigh 50 g of sample into an Erlenmeyer
flask, add 125 ml of 1 M sulfuric acid, and swirl the
flask to disperse the contents. Stopper loosely, heat
in a water bath at 100°for 10 min, mix the contents
well, and heat for an additional 30 min. For starches
that are not easy to hydrolyze, such as wheat starch,
heating time should be longer (90 min). Cool to room
temperature, adjust the pH to 7 with 25% sodium
hydroxide solution, and filter with suction through a
glass-fiber filter paper. Wash the flask and the
residue on the filter paper with 25 ml of water, and
combine the washings with the filtrate. Add 30 g of
anhydrous sodium sulfate, stir for 5–10 min to
dissolve, and transfer the solution into a separating
funnel. Wash the flask with 25ml of water, and add
the washings to the funnel. If precipitate remains, stir
well with a small amount of water to dissolve it
completely, and add the solution to the funnel.
Extract five times with five 50ml portions of diethyl
ether. Combine the diethyl ether extracts, add 3 g of
anhydrous sodium sulfate, let it stand for a few
minutes and filter through a filter paper. Wash the
flask and the filter paper with 25 ml of diethyl ether,
and combine the washings with the filtrate.
Evaporate to 4 ml in a water bath at about 40° under
atmospheric pressure, cool, transfer to a 5 ml
volumetric flask and add diethyl ether to the exact
volume.
Procedure
GC operating conditions
- GC equipped with a flame ionization detector (FID).
- Column: A fused silica column coated with
polyethylene glycol (30 m x 0.25 mm i.d., 0.25 µm)
(Inert Cap WAXGL Sciencesor equivalent）
- Carrier gas: N2 or He
- Flow rate: Adjust the retention time of 1-chloro-2-
propanol to about 15 min
- Column temperature: 40°-for 2 min; heat at 5°/min to
80°, keep for 8 min, heat at 25°/min to 230°, keep for
5 min
- Injector temperature: 150°
- Detector temperature: 230°
- Split-less（purge start: 1 min after injection）
Analyse 1-µl portions of the test solution and the

standard solutions by gas chromatography, using the
operating conditions given above. Prepare a standard
addition curve: Plot in the y axis the sum of the peak
areas corresponding to 1-chloro-2-propanol and 2-
chloro-1-propanol in the chromatographs and in the x-
axis the added concentration of propylene chlorohydrins
in the standard solution. For the test solution the added
concentration is equal to 0. A linear calibration curve
should be obtained. Extrapolate to 0 in the y axis. The
concentration (µg/ml) of propylene chlorohydrins in the
test solution is equal to the absolute value of the
concentration at the point where the curve intercepts the
x axis (Ct). Determine the content of propylene
chlorohydrins in the sample using the following formula:
Calculation
Content (mg/kg) of Propylene chlorohydrins = C𝑡𝑡 × 5 / W
where
- Ct: amount of propylene chlorohydrins in test solution
(µg/mL);
- W: mass of sample (g, on the dried weight basis)
ANNEX 8: ADDITIONAL SPECIFICATIONS FOR STARCHES CROSSLINKED

WITH ADIPIC ANHYDRIDE
(Version 2018 – Tentative)

• A suitable method for identification of crosslinking
and data on at least 5 representative batches of
crosslinked and non-crosslinked starches
• Levels of free adipic acid in at least 5 representative
batches
APPLIES TO Acetylated distarch adipate (INS No. 1422)
TREATMENT Adipic anhydride can used for esterification and crosslinking.

In cases of cross-linking, where adipic anhydride connects
two chains, the structure can be represented by: Starch-O-
R-O-Starch, where R = CO-(CH2)4-CO and starch refers to
the linear and/or branched structure.
C.A.S numbers Acetylated distarch adipate

63055-36-7 (modified amylopectin)
CHARACTERISTICS
PURITY
Adipate groups Not more than 0.135% on the dried basis
Free adipic acid Information Required
PURITY TEST
Adipate groups and Determine by gas chromatography after derivatization
free adipic acid
Principle
Free adipic acid in the sample is extracted and determined by
capillary gas chromatography after trimethylsilyl-derivatization.
Total adipic acid is determined using the same method after
hydrolysis of the sample and adipate groups are calculated by
subtraction of the free adipic acid from the total.
Standards and Reagents

Adipic acid (>99%)
Glutaric acid (>99%)
Starch, unmodified (of the same botanical origin as the sample)
Sodium hydroxide solution (4N): weigh 40g of NaOH, dissolve in
water and dilute to 250 ml.
Concentrated HCl (36%)
Ethyl acetate
Sodium sulphate, anhydrous
N,O-Bis(trimethylsilyl)trifluoroacetamide
Pyridine
Internal standard solution (1 mg/ml)

Accurately weigh 0.1 g of glutaric acid, dissolve in water and
dilute to 100 ml.
Standard stock solution (1 mg/ml)

Accurately weigh 0.1 g of adipic acid, dissolve in 90 ml of warm
water, cool to room temperature, dilute to 100 ml and mix.
Working standard solutions (0.02, 0.1, 0.2 and 0.4 mg/ml)

Pipette 1, 5, 10, and 20 ml of the standard stock solution in four
separate 50 ml volumetric flasks, and dilute with water.
Procedure
Preparation standard curve solutions
Weigh 1.0 g of starch into each of four Erlenmeyer flasks, add
50 ml of water and 1 ml of internal standard solution. Add 5 ml
each of the four working standard solutions, respectively.
Stopper the flask and shake them well to disperse the starch,
add 50 ml of 4N sodium hydroxide solution, and shake for
5 min. Place the flasks in a water bath, at room temperature,
and add cautiously 20 ml of conc. hydrochloric acid. Cool, and
quantitatively separately transfer the contents of the flasks into
four separation funnels with a little amount of water. Extract
three times with 100 ml of ethyl acetate each time. Collect the
ethyl acetate layers separately in four dry Erlenmeyer flasks,
add 20 g of anhydrous sodium sulphate, allow to stand for
10 min with occasional shaking, and filter into a rotary
evaporator flask. Wash the Erlenmeyer flask and the residue
on the filter paper twice with a small quantity of ethyl acetate,
and combine the washings with the filtrate. Evaporate the ethyl
acetate under a reduced pressure of 6.7 kPa at a temperature
below 40°. Remove the remaining ethyl acetate completely by
nitrogen stream. The evaporation of ethyl acetate should be
effected as quickly as possible. Successively add 2 ml of
pyridine and 1 ml of N,O-bis(trimethylsilyl)trifluoroacetamide to
the residue and stopper the flask. Allow the solution to stand
for 1 hour, transfer 2 ml of it into a GC vial, and immediately
stopper tightly. Use these solutions to construct standard curve
(Internal standard 1 mg/g starch, standards 0.1, 0.5, 1 and
2 mg/g starch respectively)
Preparation of test solution A (for residual free adipic acid)

Weigh accurately about 5 g of sample into an Erlenmeyer flask,
add 100 ml of water and 1 ml of the internal standard solution.
Shake well for 1 hour, and filter through a 0.45 µm membrane
filter. To the filtrate, add exactly 1 ml of hydrochloric acid (in the
case of pre-gelatinized starch or water-soluble starch, directly
add 1 ml of hydrochloric acid to the resulting suspension
without filtering), and transfer into a separation funnel. Proceed
as directed for the preparation of standard solutions, beginning
with “…and wash the inside of the flask with a little amount of
water into the funnel.” Use this solution for the determination of
residual free adipic acid (Internal standard 1 mg/ 5 g starch).
Preparation of test solution B (for total adipic acid)

Weigh accurately about 1 g of sample into an Erlenmeyer flask,
add 50 ml of water and exactly 1 ml of the internal standard
solution. Shake the mixture well to disperse the starch, add
50 ml of 4N sodium hydroxide solution and shake well for
5 minutes. Place the flask in a water bath at room temperature,
and add cautiously 20 ml of concentrated hydrochloric acid.
After cooling, transfer the contents in the flask into a separation
funnel. Proceed as directed for the preparation of standard
solution, beginning with “…and wash the inside of the flask with
a little amount of water into the funnel.” Use this solution for the
determination of total adipic acid (Internal standard 1 mg/g
starch).
Procedure
GC operating conditions
• GC equipped with a flame ionization detector (FID)
Column:A fused silica column coated with a mixture of
50% diphenyl and 50% dimethylpolysiloxane (15 m x
0.25 mm i.d., 0.25 µm)
• Carrier gas: He
• Column flow 1.0 ml/min.
• Column temperature: 120°-5 min-5°/min-150°
(Glutaric and adipic acids elute at about 5 min and
8 min respectively)
• Injector temperature: 250°
• Detector temperature: 250°
• Injection volume 1µl
• Split ratio: 30:1
Inject standard curve solutions into the capillary GC under the

conditions indicated and construct a standard curve using the
peak area ratios of adipic acid and glutaric acid against the
amounts of adipic acid in the standard solutions (in g). Inject
the test solution A and Band obtain the peak area ratio of
adipic acid to glutaric acid for each of the test solutions A and
B.
Determine the amount of adipic acid in each test solution from

the standard curve and calculate the percent of adipate groups
using the following formula:
Free adipic acid, %w/w = [CF/MF] x 100
Adipate groups. %w/w = [CT/MT–CF/MF] x 100
where
CT = amount of the total adipic acid in the test solution
B (g)
CF = amount of the free adipic acid in the test solution
A (g)
MT =mass of sample in the test solution for the
determination of total adipic acid (g, on the dried
weight basis)
MF =mass of the sample in the test solution for the
determination of free adipic acid (g, on the dried
weight basis)
NEUTRAL METHACRYLATE COPOLYMER (TENTATIVE)
New tentative specifications prepared at the 86th JECFA (2018)

and published in FAO JECFA Monographs 22 (2018). An ADI of
“not specified” was established at 86th JECFA (2018).
Information required on:

• A validated method for the assay of neutral methacrylate
copolymer (e.g., quantitative IR)
• Performance characteristics (method validation data) of
the assay method
• Assay and monomers data on at least five batches of
products currently available in commerce
SYNONYMS E 1206, INS No. 1206, Ethyl acrylate methyl methacrylate

polymer, Ethyl acrylate methyl methacrylate polymer; Ethyl
acrylate polymer with methyl methacrylate, Methyl methacrylate
ethyl acrylate polymer, Methyl methacrylate polymer with ethyl
acrylate.
DEFINITION Neutral Methacrylate Copolymer is a copolymer comprised of the

monomers ethyl acrylate and methyl methacrylate in the molar
ratio of 2:1. The copolymer is manufactured by emulsion
polymerization of the monomers with water-soluble radical
initiators. The product is purified by water vapour distillation and
filtration to remove residual monomers, excess water, other
volatile low-molecular weight substances and coagulum. The
copolymer is standardized as a 30% aqueous dispersion with
polyethylene glycol monostearyl ether. The copolymer dispersion
may contain the residual monomers (methyl methacrylate and
ethyl acrylate). Neutral methacrylate copolymer is used as a
coating and glazing agent for food supplements and foods for
special medical purposes
Chemical name Poly(ethyl acrylate-co-methylmethacrylate)
C.A.S. number 9010-88-2
Chemical formula Poly[(CH2:CHCO2CH2CH3)-co-(CH2:C(CH3)CO2CH3)]

Structural formula
••• •••
The above formula is provided for illustrative purposes; in this

copolymer no definitive structural unit can be defined.
Formula weight 600,000 (weight-average), 220,000 (number-average)
Assay Information required
DESCRIPTION Commercial form (30% aqueous dispersion) is a low viscosity

milky-white liquid
FUNCTIONAL USES Coating agent, binding agent, glazing agent
CHARACTERISTICS
IDENTIFICATION
Viscosity (Vol. 4) Not more than 50 mPa.s
Determine viscosity using Brookfield viscometer at 20° and

300 pm using UL adapter.
pH (Vol 4) 5.5 – 8.6

corresponds to the infrared spectrum in the Appendix.

PURITY
Loss on drying (Vol 4) 68.5 – 71.5% (110°, 3 h)

Residual solvents Methanol: Not more than 100 mg/kg

(Vol. 4)
Residual monomers Ethanol: Not more than 1,000 mg/kg
Ethyl acrylate: Not more than 20 mg/kg
Lead (Vol. 4) Not more than 1.0 mg/kg

selection of sample size and method of sample preparation may
be based on principles of methods described in Volume 4 (under
Microbiological criteria Total plate count: Not more than 1,000 cfu/g
(Vol. 4)
Yeast and moulds: Not more than 100 cfu/g
TESTS
PURITY TESTS
Residual monomers Determined by liquid chromatography (Vol. 4)

• Acetonitrile: UV absorption: Amax of 1% at 190 nm
• Tetrahydrofuran and deionized water:
• Sodium perchlorate (35g/l)
• Standards: Ethyl acrylate and methyl methacrylate (>99%)
Preparation of mixed standard solutions

Stock mixed standard solution (200 µg/ml):
Accurately weigh about 10 mg each of ethyl acrylate and methyl
methacrylate, dissolve in tetrahydrofuran and make up to 50 ml
with tetrahydrofuran in a volumetric flask.
Intermediate mixed standard solution-1 (20 µg/ml):

Dilute 1.0 ml of stock mixed standard solution to 10 ml with
tetrahydrofuran in a volumetric flask.
Intermediate mixed standard solution-2 (2 µg/ml):

Dilute 1.0 ml of intermediate mixed standard solution-1 to 10 ml
with tetrahydrofuan in a volumetric flask.
Working mixed standard solution (0.67 µg/ml):

To 10 ml of Intermediate mixed standard solution-2 add 5 ml of
sodium perchlorate and mix. Dilute 5 ml of this mixture to 10 ml
with deionized water.
Preparation Sample Solution

Accurately weigh approximately1.0 g of sample, dissolve in
tetrahydrofuran and dilute to 50.0 ml in a volumetric flask. To 5 ml
of sodium perchlorate solution, add 10 ml of sample solution drop
wise, whilst stirring continuously. Centrifuge and filter the clear
supernatant. Dilute 5 ml of this mixture to 10 ml with deionized
water.
Procedure
Use a HPLC with diode array/UV detector at 205 nm
Column: Octadecylsilyl silica gel (12 cm x 4.6 mm i.d., 5-10 µm.)
Injection volume: 50 μl
Mobile phase: Acetonitrile:Water (15:85)
Flow rate: 2 ml/min
Inject separately 50 µl each of working mixed standard solution

and sample solution. Calculate the amount of each monomer in
the sample from the peak areas obtained in the chromatograms
of working mixed standard solution (rR) and sample solution (rS);
amount of standard in the injected solution (R, µg) and weight of
sample in injected sample solution (W, g)
rS × R
Amount of each monomer (µg/g) =
rR × W
Total monomers in the sample (µg/g) = Sum of monomers in the

sample
METHOD OF ASSAY Information Required

Appendix: Infrared spectrum of neutral methacrylate copolymer

SPIRULINA EXTRACT (TENTATIVE)
New specifications prepared at the 86th JECFA (2018),

published in FAO JECFA Monograph 22 (2018). A temporary
ADI “not specified” was established at the 86th JECFA (2018).
Information Required on:

• Full compositional characterization of commercial
products in both liquid and powder forms.
• Full compositional characterization of the aqueous
extract before formulation/standardization.
• Validated analytical methods for identification of the
substance with a suitable specificity (including
validation data and representative batch data).
• Validated analytical methods for the determination of
the purity of the substance with a suitable specificity
(including validation data and representative batch
data).
SYNONYMS INS 134; Spirulina colour
DEFINITION Spirulina extract is obtained by aqueous extraction of the

biomass of Arthrospira platensis, an edible cyanobacterium.
The organism is cultivated and harvested under conditions that
prevent the growth of other cyanobacteria and the production
of microcystins. The material extracted from the biomass is
further treated by steps that may include pH adjustment,
centrifugation, filtration, concentration, sterilization, drying, and
dilution to the desired degree of pigment concentration. The
main colouring principles are two phycobiliproteins, C-
phycocyanin and allophycocyanin, which are water-soluble
pigment-protein complexes where the chromophore is
covalently bonded to the protein. Extracts may also contain
trace amounts of chlorophyll, beta-carotene, and other
carotenoids. Spirulina extract may contain peptides, other
proteins, carbohydrates and minerals. Commercial products
are formulated in liquid and powder forms.
C.A.S. number 20298-86-6
(Phycocyanobilin; 3-[(2Z,5E)-2-[[3-(2-carboxyethyl)-5-[(Z)-
[(3E,4R)-3-ethylidene-4-methyl-5-oxopyrrolidin-2-
ylidene]methyl]-4-methyl-1H-pyrrol-2-yl]methylidene]-5-[(4-
ethyl-3-methyl-5-oxopyrrol-2-yl)methylidene]-4-methylpyrrol-3-
yl]propanoic acid)
Chemical formula C33H38N4O6 (Phycocyanobilin)

Structural formula
Phycocyanobilin
Formula weight 586.68 (Phycocyanobilin)
Assay Total phycocyanins as the sum of C-phycocyanin and

allophycocyanin not less than declared.
DESCRIPTION Clear blue liquid or blue powder
CHARACTERISTICS
IDENTIFICATION
Solubility (Vol. 4) Freely soluble in water. Insoluble in ethanol.
Colour Value Not less than declared (15 to 300 for powdered products on
the dried basis and 10 to 70 for liquid products).
PURITY
Loss on drying (Vol. Not more than 6% for the powdered product (105°, 4h)
4)
Arsenic (Vol. 4) Not more than 1 mg/kg

Impurities”).
Cadmium (Vol. 4) Not more than 1 mg/kg

Impurities”).

Impurities”).
Mercury (Vol. 4) Not more than 1 mg/kg

Impurities”).
Microbiological Total (aerobic) plate count: less than 1000 CFU/g

criteria (Vol. 4)
Yeast and moulds: less than 100 CFU/g
Coliforms: absent in 10 g
Salmonella spp.: absent in 25 g
S. aureus: absent in 10 g
Microcystins Less than 0.5 µg/g as microcystin-LR (dried basis)
TESTS
PURITY TESTS
Microcystins Principle
Determine microcystins by enzyme linked immunoassay
(ELISA) under the following conditions:
Reagent
Methanol/water (75:25, v/v)
Equipment
Use a commercially available ELISA kit with cross reactivity for
microcystin-LR and other microcystins.
Sample preparation
In the absence of other instructions provided by the ELISA kit
manufacturer, follow the procedure presented here.
Dry an appropriate amount of spirulina extract. Homogenize

3.0 g of the dried material in 20.0 ml of the methanol/water
reagent for 20 minutes. Centrifuge the resulting suspension at
4500 rpm for 10 minutes. Transfer the supernatant into a glass
flask. Add 10.0 ml of the methanol/water reagent to the
homogenizer and homogenize the residue for 30 seconds.
Centrifuge the resulting suspension at 4500 rpm for 10
minutes. Combine the supernatants and dilute with water to a
concentration within the range indicated by the ELISA kit
manufacturer.
Procedure
Follow the instructions provided by the ELISA kit
manufacturer.
Colour Value For the purpose of this specification, Colour Value is based on
the absorbance of a buffered solution at 618 nm.
Reagent
Sodium phosphate buffer (100 mM, pH 6.0): Transfer 14.04 g
of sodium phosphate monobasic dihydrate and 1.75 g of
sodium phosphate dibasic anhydrous into a 1000 ml
volumetric flask and dilute to volume with water containing
0.05% sodium azide. Adjust the pH to 6.0 with a few drops of
phosphoric acid or 1 M NaOH if needed.
Procedure
Transfer 330 mg of spirulina extract into a 100 ml volumetric
flask and dilute to volume with water. Transfer 10 ml of the
solution into a second 100 ml volumetric flask and dilute to
volume with the sodium phosphate buffer (100 mM, pH 6.0).
Determine the absorbance (A618) of the solution in a 1-cm cell
at 618 nm with a suitable spectrophotometer using sodium
phosphate buffer (100 mM, pH 6.0) as the reference.
Calculate the Colour Value of the spirulina extract as follows:
Colour Value = A618 x 100 / W 1
Where
W1 is the weight of spirulina extract taken, in g
METHOD OF ASSAY Principle
Determine total phycocyanins as the sum of C-phycocyanin

and allophycocyanin under the following conditions:
Reagent
Sodium phosphate buffer (100 mM, pH 6.0): Transfer 14.04 g
of sodium phosphate monobasic dihydrate and 1.75 g of
sodium phosphate dibasic anhydrous into a 1000 ml
volumetric flask and dilute to volume with water containing
0.05% sodium azide. Adjust the pH to 6.0 with a few drops of
phosphoric acid or 1 M NaOH if needed.
Procedure
Transfer 100 mg of spirulina extract into a 25 ml volumetric
flask and dilute to volume with sodium phosphate buffer
(100 mM, pH 6.0). Sonicate the mixture for 30 minutes
maintaining the temperature at 8°. Incubate at 30° for 8 h,
shaking manually every hour. Mix the contents of the flask and
transfer to a centrifugation tube; centrifuge at 3500 rpm for
4 minutes. Determine the absorbance of the supernatant in a
1-cm cell at 620 nm (A620) and 650 nm (A650) with a suitable
spectrophotometer using sodium phosphate buffer (100 mM,
pH 6.0) as the reference. The dilution should be adjusted with
additional buffer, if needed, to obtain absorbance values of
0.2 to 0.6 at 620 nm.
Calculate the C-phycocyanin content of the spirulina extract

(%, w/w) as follows:
TcPC = (0.162 x A620) – (0.098 x A650) x V1 x 100 / W1
Where
W1 is the weight of spirulina extract taken, in mg
V1 is the volume of the volumetric flask used to prepare the
sample solution, in mL
Calculate the allophycocyanin content of the spirulina extract

(%, w/w) as follows:
TaPC = (0.180 x A620) – (0.042 x A650) x V1 x 100 / W1
Where
W1 is the weight of spirulina extract taken, in mg
V1 is the volume of the volumetric flask used to prepare the
sample solution, in mL
Calculate the total phycocyanin content of the spirulina extract

as follows:
TPC = TcPC + T aPC

SPECIFICATIONS FOR CERTAIN FLAVOURING AGENTS
At the 86th meeting, the Committee prepared specifications of identity and purity of 69
flavourings in 8 sub-categories for the following numbers: 380.1, 380.2, 427, 433, 619, 973-
975, 980-982, 1480, 1491-1526, 2103-2105, 2123, 2235-2244, 2246-2255.
Information on specifications for flavouring agents is given in the tables, most of which are
self-explanatory: Name; Chemical name (Systematic name, normally IUPAC name);
Synonyms; Flavour and Extract Manufacturers' Association of the United States (FEMA) No;
FLAVIS (FL) No; Council of Europe (COE) No; Chemical Abstract Service Registry (CAS) No;
Chemical formula (Formula); Molecular weight (MW); Physical form/Odour; Solubility;
Solubility in ethanol, Boiling point (B.P. ºC – for information only); Identification test (ID)
referring to type of test (NMR: Nuclear Magnetic Resonance spectrometry; IR: Infrared
spectrometry; MS: Mass spectrometry); Assay min % (Gas chromatographic (GC) assay of
flavouring agents); Acid value max; Refractive index (R.I.) (at 20º, if not otherwise stated);
Specific gravity (S.G) (at 25º, if not otherwise stated). The field called "Other requirements"
contains four types of entry:
1. Items that are additional requirements, such as further purity criteria or other tests.
2. Items provided for information, for example the typical isomer composition of the
flavouring agent. These are not considered to be requirements.
3. Substances which are listed as Secondary Constituents (SC) which have been taken
into account in the safety evaluation of the named flavouring agent. If the commercial
product contains less than 95% of the named compound, it is a requirement that the
major part of the product (i.e. not less than 95% is accounted for by the sum of the
named compound and one or more of the secondary constituents.
4. Information on the status of the safety evaluation.
The fields named Session/Status contain the number of the meeting at which the
specifications were prepared and the status of the specification. All specifications prepared at
the 86th meeting were assigned full status.
The flavouring agents were evaluated using the Procedure for the Safety Evaluation of
Flavouring Agents and a list for conclusions in alphabetical order is given in Annex I.
GROUP 1: MISCELLANEOUS NITROGEN-CONTAINING SUBSTANCES
JECFA Other
No. Name FEMA Chemical Formula Solubility ID test R.I. (20°) requirements
Solubility in
Status Chemical Name FLAVIS M.W ethanol Assay min % S.G. (25°)
Physical form; Information
Synonyms CE Odour B.P. ° Acid value required
Sessio
n CAS
2-(((3-(2,3-Dimethoxyphenyl)-1H-1,2,4-triazol-5- Phosphate buffer, pH MS, 1H-NMR,
2235 yl)thio)methyl)pyridine 4798 C16H16N4O2S 7.1 = 0.30 mM 13C-NMR, IR NA mp: 114.0 - 116.0 º
Pyridine, 2-(((3-(2,3-dimethoxyphenyl)-1H-1,2,4-
Draft triazol-5-yl)thio)methyl) 328.39 Soluble >98 % NA
2-((5-(2,3-dimethoxyphenyl)-2H-1,2,4-triazol-3-
ylthio)methyl)pyridine
White to off-white solid
86 902136-79-2
(S)-1-(3-(((4-Amino-2,2-dioxido-1H-
benzo[c][1,2,6]thiadiazin-5-
yl)oxy)methyl)piperidin-1-yl)-3-methylbutan-1- MS, 1H-NMR, mp: 236-238 º; <5%
2236 one 4802 C18H26N4O4S Soluble 13C-NMR, IR NA R-enantiomer
(S)-1-(3-(((4-amino-2,2-dioxido-1H-
benzo[c][1,2,6]thiadiazin-5-yl)oxy)methyl)piperidin-1-
Draft yl)-3-methylbutan-1-one 394.49 Soluble >95 % NA
Off-white solid
100
86 1469426-64-9
2-(4-Methylphenoxy)-N-(1H-pyrazol-3-yl)-N- Slightly soluble at MS, 1H-NMR, mp: 115-
2237 (thiophen-2-ylmethyl)acetamide 4809 C17H17N3O2S pH 2.8 13C-NMR, IR NA 116.5 °C
N-(1H-pyrazol-3-yl)-N-(thiophen-2-ylmethyl)-2-(4-
Draft tolyloxy)acetamide 16.133 327.40 Soluble 99% NA
White to off-white solid

86 1374760-95-8
101
GROUP 2: SATURATED ALIPHATIC ACYCLIC BRANCHED-CHAIN
PRIMARY ALCOHOLS, ALDEHYDES, AND ACIDS
JECFA Other
Solubility in
Information
Synonyms CE Physical form; Odour B.P. ° Acid value required
Session CAS
MS, 1H-NMR, 1.4224-
2238 8-Methyldecanal 4795 C11H22O very slightly soluble 13C-NMR 1.4421
0.879-0.919
Draft 8-methyldecanal 170.29 Soluble 95% (20 °)
colorless liquid/ citrus/green odor 222-223 º 10
86 127793-88-8
MS, 1H-NMR, 1.4256 -
2239 8-Methylnonanal 4803 C10H20O very slightly soluble 13C-NMR 1.4260
0.8227 -
Draft 8-Methylnonanal 156.27 Soluble >95% 0.8231 (20 °)
Colorless, transparent liquid/Sweet
Isodecanal, Isodecaldehyde aroma of fruit with green notes 196-197 º
86 3085-26-5
102
GROUP 3: LINEAR AND BRANCHED-CHAIN ALIPHATIC, UNSATURATED, UNCONJUGATED ALCOHOLS,
ALDEHYDES, ACIDS, AND RELATED ESTERS
JECFA Other
Solubility in
Information
Session CAS
1H-NMR, 13C-
2240 trans-6-Octenal 4787 C8H14O Slightly soluble NMR 1.4377
Draft (E)-Oct-6-enal 126.20 Soluble 95% 0.8536 (20 º)
(E)-6-Octenal; (6E)-Octenal Colorless oil / Green, melon-like

aroma 171-172º
86 63196-63-4
Secondary
component: 1-6%
2,6-dimethyl-5-
heptenal (No. 349)
The sum of 2,6-
dimethyl-5-heptenol
MS, 1H-NMR, and 2,6-dimethyl-5-
2241 2,6-Dimethyl-5-heptenol 4789 C9H18O Slightly soluble 13C-NMR 1.428-1.459 heptenal is ≥ 95%
Draft 2,6-Dimethylhept-5-en-1-ol 142.24 Soluble >90% 0.794-0.904
Colorless to light yellow,
2,6-dimethyl-5-hepten-1-ol, Melonol transparent liquid/Floral fruity
aroma reminiscent of melon 204-206 °
86 4234-93-9
103
GROUP 4: CARVONE AND STRUCTURALLY RELATED SUBSTANCES
JECFA Other
Solubility in
Information
Sessio
n CAS
Practically insoluble
2242 Pinocarvyl isobutyrate 4525 C14H22O2 to insoluble MS 1.445-1.450
6,6-dimethyl-2-
methylidenebicyclo[3.1.1]heptan-3-yl 2-
Draft methylpropanoate 222.32 Soluble >95% 0.912-0.915
Propanoic acid, 2-methyl-, 6,6-dimethyl-
2-methylenebicyclo[3.1.1]hept-3-yl ester Viscous colourless to slight/pale
yellow liquid / Warm woody
balsamic aroma with fruity notes 264-266 º
86 929116-08-5
mp: 144-145 º;
Mixture of
(2R,4S)-carvyl
palmitate and
Practically insoluble NA (2S,4S)-carvyl
2243 Carvyl palmitate 4515 C26H46O2 to insoluble MS, 1H-NMR palmitate
5-Isopropenyl-2-methyl-2-cyclohexen-1-yl NA
Draft palmitate 390.64 Soluble >95%
Carvyl hexadecanoate Waxy solid/Rich fatty spearmint
to caraway aroma
86 929222-96-8
104
NA
2244 6-Hydroxycarvone 4523 C10H14O2 Slightly soluble MS, 1H-NMR, IR mp: 185 º
3-hydroxy-2-methyl-5-(prop-1-en-2- NA
Draft yl)cyclohex-2-en-1-one 166.22 Soluble >95%
Solid/cool mint-like aroma

86 51200-86-3
105
GROUP 5: MENTHOL AND STRUCTURALLY RELATED SUBSTANCES
JECFA Chemical
No. Name FEMA Formula Solubility ID test R.I. (20°) Other requirements
Solubility in
Physical form;
Synonyms CE Odour B.P. ° Acid value Information required
Sessio
n CAS
2246 Menthyl formate 4509 C11H20O2 to insoluble MS, IR 1.446-1.452 racemic mixture
Draft 2-isopropyl-5-methylcyclohexyl formate 184.28 Sparingly soluble >95% 0.933-0.939
Clear colorless
liquid/Sweet minty
aroma 219-220°
86 2230-90-2
2247 Menthyl propionate 4510 C13H24O2 to insoluble MS 1.444-1.449
2-isopropyl-5-methylcyclohexyl
Draft propionate 212.34 Sparingly soluble >95% 0.918-0.923
Clear colorless
liquid/fruity cool
aroma 246-247 º
86 86014-82-6
106
2248 l-Menthyl butyrate 4524 C14H26O2 to insoluble MS 1.445-1.450
Draft 2-isopropyl-5-methylcyclohexyl butyrate 226.36 Sparingly soluble >95% 0.912-0.915

clear colorless
liquid/tropical cool
aroma 262-263 º
86 68366-64-3
2249 dl-Isomenthol 4729 C10H20O to insoluble MS, 1H-NMR, IR NA mp: 82 º; racemic mixture
5-methyl-2-(propan-2-yl)cyclohexan-1-
ol
Draft 156.27 Sparingly soluble >95% NA
Cyclohexanol, 5-methyl-2-(1-
methylethyl)-, (1-alpha,2-beta,5-beta)-
(±)-; Menthol, trans-1,3,cis-1,4-(±)-; (±)-
Isomenthol; Cyclohexanol, 5-methyl-2-
(1-methylethyl)-, (1-alpha,2-beta,5-
beta)-; Isomenthol Solid/cool minty
aroma
(1S, 2R, 5R) (+)-Isomenthol
(1R, 2S, 5S) (-)-Isomenthol
86 3623-52-7
107
Practically insoluble NA
2250 Dimenthyl glutarate 4604 C25H44O4 to Insoluble 1H-NMR mp: Approx. 40 °
bis(2-isopropyl-5-methylcyclohexyl) NA
Draft glutarate 09.935 408.61 Very slightly soluble >95%
Pentanedioic acid, bis[5-methyl-2-(1-
methylethyl)cyclohexyl] ester Amber amorphous or
(9CI);Glutaric acid, di-(p-menth-3-yl) crystalline solid/Fresh
Ester minty aroma < 1.0
86 406179-71-3
Melting Range: 41-44° ((-)-
very soluble in alcohol menthol);
and volatile oils; Nonvolatile Residue: =< 0.05%;
slightly soluble in Angular Rotation: -52° to -40° ((-)-
427 Menthol 2665 C10H20O water IR 1.461 menthol): -2° to +2° (dl-menthol)
3-p-Menthanol 95%; sum of (+/-) 0.901 (20°);
Full 156.27 isomers 0.891 (30°)
colourless, hexagonal
crystals, usually
needle-like; fused
masses or crystalline
powder with a
pleasant, peppermint- 212° ((-)-isomer
63 like odour 216.5)
51 89-78-1
108
2251 (±)2-[(2-p-Menthoxy)ethoxy]ethanol 4718 C14H28O3 to insoluble MS, 1H-NMR, IR 1.444 -1.484 (1R, 2S, 5R) (1S, 2R, 5S)
>95% (as a sum 0.947 -

2-(2-((2-isopropyl-5- of the + and – 0.987 (20°);
methylcyclohexyl)oxy)ethoxy)ethan-1- isomers; racemic 0.945 -0.985
Draft ol 244.37 Soluble mixture) (25°)
2-[2-(2-Isopropyl-5-methyl- Colorless viscous

cyclohexyloxy)-ethoxy]-ethanol liquid/Minty herbal
aroma with fruity
notes 97° (0.2 mmHg) NA
86 28804-53-7
109
GROUP 6: MALTOL AND RELATED SUBSTANCES
JECFA No. Name FEMA Chemical Formula Solubility ID test R.I. (20°) Other requirements
Status Chemical Name FLAVIS M.W Solubility in ethanol Assay min % S.G. (25°)
Synonyms CE Physical form; Odour B.P. ° Acid value Information required
Session CAS
Ethyl maltol
2252 isobutyrate 4534 C11H14O4 Practically insoluble to insoluble MS, IR 1.480-1.496 Secondary component: 2-3% Ethyl maltol (No. 148
2-ethyl-4-oxo-4H-
Draft pyran-3-yl isobutyrate 210.23 Soluble 93-94% 1.132-1.138
Clear light yellow liquid/sweet fruity aroma 58-65 º (2 mm Hg)
86 852997-28-5
110
GROUP 7: ALICYCLIC PRIMARY ALCOHOLS, ALDEHYDES, ACIDS
AND RELATED ESTERS
JECFA
No. Name FEMA Chemical Formula Solubility ID test R.I. (20°) Other requirements
Solubility in
Synonyms CE Physical form; Odour B.P. ° Acid value Information required
Session CAS
Mixture of 1-Vinyl-3- 60%-70% 1-vinyl-3-
cyclohexenecarbaldehyde and cyclohexenecarbaldehyde
4-Vinyl-1- Very slightly MS, 1H-NMR, 1.4870 - and 25-35% 4-vinyl-1-

2253 cyclohexenecarbaldehyde 4783 C9H12O soluble 13C-NMR 1.4930 cyclohexenecarbaldehyde
1-vinylcyclohex-3-ene-1-
carbaldehyde and 4-
vinylcyclohex-1-ene-1- >95% (sum of 0.9565 -
Draft carbaldehyde 136.19 Soluble mixture) 09665
Liquid; Spicy herbal fruit
aroma 205 °
1049017-63-
1049017-63-1 1049017- 1; 1049017-
86 68-6 68-6
Insoluble in
water; soluble in 1.504- Safety evaluation not
973 p-Mentha-1,8-dien-7-al 3557 C10H14O alcohols and oils NMR 1.513 completed
p-Mentha-1,8-dien-7-al Miscible at room 0.948-
Full 150.22 temperature 97% 0.956
Perilla aldehyde;Dihydrocuminic
aldehyde;Perillaldehyde;4-
Isopropenyl-1-cyclohexene-1- Pale, yellowish oily liquid;
carboxaldehyde powerful, fatty-spicy, oily-
11788 herbaceous odour 104° (10 mm Hg) 3
111
86 2111-75-3
(1-Methyl-2-(1,2,2-
trimethylbicyclo[3.1.0]hex-3- very slightly MS, 1H-NMR, 1.4820 -
2254 ylmethyl)cyclopropyl)methanol 4776 C15H26O soluble 13C-NMR 1.4880 1S,2S 55% 1R,2R 40%
1-methyl-2-(1,2,2-
trimethylbicyclo[3.1.0]hex-3- 0.941 -
Draft ylmethyl)cyclopropyl)methanol 222.37 Soluble >95% 0.951 (20°)
Liquid; Strong musky, sweet,
balsamic woody aroma
Javanol reminiscent of sandalwood 268 °
86 198404-98-7
(±)-Bicyclo[2.2.1]hept-5-ene-2- very slightly 1.448- Racemic mixture of R-
2255 carboxylic acid, ethyl ester 4790 C10H14O2 soluble MS, 1H-NMR 1.488 and S-
ethyl bicyclo[2.2.1]hept-5-ene-2- 1.007-
Draft carboxylate 166.09 Soluble >95% 1.047 (20°)
Ethylbicyclo[2.2.1]hept-5-ene-2-
carboxylate; 5-Norbornene-2-carboxylic Clear liquid; Floral aroma
acid, ethyl ester; 2,5-Endomethylene-3- with earthy fermented
cyclohexene carboxylic acid, ethyl ester undertones 215-217 °
86 10138-32-6
112
GROUP 8: FURAN SUBSTITUTED ALIPHATIC HYDROCARBONS, ALCOHOLS, ALDEHYDES,
KETONES, CARBOXYLIC ACIDS AND RELATED ESTERS,
SULFIDES, DISULFIDES AND ETHERS
JECFA Other
Solubility in
Information
Session CAS
Slightly soluble in
1491 2-Pentylfuran 3317 C9H14O water NMR 1.443-1.449
Full 2-Pentylfuran 13.059 138.21 Soluble 99% 0.886-0.893
58-60° (10 mm
2-Amylfuran Colourless liquid; Fruity aroma Hg) 1
86 3777-69-3
1492 2-Heptylfuran 3401 C11H18O Insoluble in water NMR 1.446-1.452
Full 2-Heptylfuran 13.069 166.26 Soluble 99% 0.860-0.866
Colourless to yellowish liquid;
Nutty, coffee-like aroma 209-210° 1
86 3777-71-7
113
1493 2-Decylfuran 4090 C14H24O Insoluble in water NMR (13C) mp: 30°
Full 2-Decylfuran 13.106 208.34 Soluble 95%

Colourless solid; Spicy, fatty
aroma
86 83469-85-6
Slightly soluble in
1494 3-Methyl-2-(3-methylbut-2-enyl)-furan 4174 C10H14O water MS 1.473-1.479
Full 3-Methyl-2-(3-methylbut-2-en-1-yl)furan 13.148 150.22 Soluble 98% 0.998-1.004
Rosefuran;2-(3-Methyl-2-butenyl)-3- Colourless liquid; Caramel
methylfuran aroma 70° (11 mm Hg) 1
86 15186-51-3
1497 3-(2-Furyl)acrolein 2494 C7H6O2 Insoluble in water NMR mp: 49-52°
Full 3-(2-Furyl)prop-2-enal 13.034 122.12 Soluble 97%
Furylacrolein;3-(2-Furyl)acrylaldehyde;3- White or yellow needles;
(2-Furyl)-2-propenal;2-Furanacrolein Cooked spicy-herb aroma 3
86 623-30-3
114
Slightly soluble in
1499 3-(5-Methyl-2-furyl)prop-2-enal 4175 C8H8O2 water NMR (13C) 1.006-1.012
Full 3-(5-Methyl-2-furyl)prop-2-enal 13.150 136.15 Soluble 95% 0.998-1.004
(5-Methylfuryl)acrolein;3-(5-Methyl-2-
furanyl)-2-propenal;1-(5-Methyl-2-
furanyl)-1-propen-3-al;5-Methyl-2- Colourless liquid; Sweet spicy
furanacrolein aroma 101° (5 mm Hg) 3
86 5555-90-8
Very slightly
soluble in water;
Slightly soluble in
propylene glycol,
1503 2-Furyl methyl ketone 3163 C6H6O2 vegetable oils IR 1.505-1.510
Full 2-Acetylfuran 13.054 110.11 Soluble 97% 1.102-1.107

Yellow to brown liquid; Coffee-
Acetylfuran;Methyl 2-furyl ketone like aroma 67° (10 mm Hg) 1
86 1192-62-7
Slightly soluble in
water; Soluble in
1504 2-Acetyl-5-methylfuran 3609 C7H8O2 corn oil HNMR IR 1.511-1.517
1.066-1.072
Full 2-Acetyl-5-methylfuran 13.083 124.14 Soluble 99% (20°)
Methyl 5-methyl-2-furyl ketone;1-(5- Colourless liquid; Strong,
Methyl-2-furyl) ethanone nutty aroma 71-72° (8 mm Hg) 2
86 1193-79-9
115
1505 2-Acetyl-3,5-dimethylfuran 4071 C8H10O2 Insoluble in water MS 1.494-1.500 mp: 18°
1522 Difurfuryl ether 3337 C10H10O3 Insoluble in water NMR 1.138-1.144
Full Difurfuryl ether

2-Acetyl-3,5-dimethylfuran 13.061 178.19
138.17 Soluble
Soluble 97%
95% 1.506-1.512
1.041-1.047
Colourlessliquid
Colourless to yellow liquid;
to solid;
Furfuryl ether
3,5-Dimethyl-2-furyl methyl ketone Coffee-like,
Sweet balsamicnutty aroma
aroma 88-89° (1 mm Hg)
195° 11
86 22940-86-9
1507 2-Butyrylfuran 4083 C8H10O2 Insoluble in water MS 1.489-1.495
86 4437-22-3
Full 2-Butyrylfuran 13.105 138.17 Soluble
Practically 95% 1.050-1.056
Furyl n-propyl ketone;1-(2-Furyl)-1- Colourless liquid; Balsamic insoluble in water
butanone;2-Furyl propyl ketone aroma and hexane;
195° 1
1523 2,5-Dimethyl-3-furanthiol acetate 4034 C8H10O2S Soluble in ether HNMR IR MS 1.527-1.533
Full 2,5-Dimethyl-3-thioacetoxyfuran 13.116 170.23 Soluble 98% 1.137-1.143
S-(2,5-Dimethyl-3-furyl)
86 4208-57-5
ethanethioate;Thioacetic acid S-(2,5- Colourless liquid; Fruity floral
dimethyl-furan-3-yl) ester aroma Soluble230°
in ether, 5
1508 (2-Furyl)-2-propanone 2496 C7H8O2 triacetin NMR 1.499-1.505
Full 1-(2-Furyl)-propan-2-one 13.045 124.14 Soluble 97% 1.074-1.080
Furyl acetone;Methyl furfuryl Liquid; Aroma suggestive of
ketone;Furfuryl methyl ketone raddish 179-180° 1
86 55764-22-2
86 6975-60-6
Slightly soluble in
1509 2-Pentanoylfuran 4192 C9H12O2 water MS 1.486-1.492
Full 2-Pentanoylfuran 13.163 152.19 Soluble 95% 1.009-1.015
1-(2-Furanyl)-1-pentanone;Butyl 2-furyl Colourless liquid; Sweet
ketone caramel aroma 101° (10 mm Hg) 1
116
86 3194-17-0
Slightly soluble in
1510 1-(2-Furyl)butan-3-one 4120 C8H10O2 water MS mp: 37°
Full 1-(2-Furyl)butan-3-one 13.138 138.17 Soluble 95%
1-(2-Furyl)-3-butanone;4-(2-Furyl)-2- Colourless solid; Spicy
butanone;Furfurylacetone caramel aroma
86 699-17-2
1511 4-(2-Furyl)-3-buten-2-one 2495 C8H8O2 Insoluble in water NMR mp: 37-40°
Full 4-(2-Furyl)but-3-en-2-one 13.044 136.15 Soluble 98%

Colourless needle crystals;
Furylidene acetone;Furfural acetone Spicy aroma 1
86 623-15-4
Very slightly
1513 Ethyl 3-(2-furyl)propanoate 2435 C9H12O3 soluble in water NMR mp: 24-25°
Full Ethyl 3(2-furyl)propionate 13.022 168.19 Soluble 95%
Low melting solid, turning
Ethyl furfurylacetate;Ethyl yellow on exposure to air;
furylpropionate;Ethyl 2-furanpropionate Fruity aroma 5
86 10031-90-0
117
Very slightly
1514 Isobutyl 3-(2-furan)propionate 2198 C11H16O3 soluble in water NMR 1.531-1.537
Full 2-Methylpropyl 3-(2-furyl)propanoate 13.024 196.25 Soluble 96% 1.007-1.013

Isobutyl 2-furanpropionate;Isobutyl Colourless to pale, straw-
furylpropionate;Isobutyl 3-(2- yellow liquid; Fruity, winey,
furyl)propanoate brandy-like aroma 105° (3 mm Hg) 5
86 105-01-1
1515 Isoamyl 3-(2-furan)propionate 2071 C12H18O3 Insoluble in water NMR 1.549-1.557
Full 3-Methylbutyl 3-(2-furan)propanoate 13.023 210.27 Soluble 96% 0.987-0.993
Colourless to pale yellow
2-Isoamyl furfurylacetate;3-Methylbutyl 3(2- liquid; Sweet, green, slightly
furyl)propionate;Isoamyl 2-furanpropionate floral aroma 258° 5
86 7779-67-1
1516 Isoamyl 4-(2-furan)butyrate 2070 C13H20O3 Insoluble in water NMR 1.551-1.555
Full 3-Methylbutyl 4-(2-furan)butanoate 13.021 224.3 Soluble 95% 0.975-0.981
Isopentyl 2-furanbutyrate;alpha-Isoamyl Pale yellowish liquid; Sweet-
furfurylpropionate;3-Methylbutyl 2- buttery, fruity and caramel-like
furanbutyrate aroma 263-265° 5
86 7779-66-0
118
Insoluble in water;
1517 Phenethyl 2-furoate 2865 C13H12O3 Soluble in oils NMR 1.585-1.593
Full Phenethyl 2-furoate 13.006 216.24 Soluble 96% 1.136-1.142
Colourless liquid; Warm,
2-Phenylethyl 2-furoate;Phenylethyl 2- fruity-caramel, slightly earthy,
furoate oily aroma 275° 5
86 7149-32-8
Insoluble in water;
1520 Furfuryl methyl ether 3159 C6H8O2 Soluble in ether NMR 1.454-1.460
Full Furfuryl methyl ether 13.052 112.13 Soluble 99% 1.013-1.019
Clear to yellow liquid; Airy,
Methyl furfuryl ether roasted coffee aroma 134-135° 1
86 13679-46-4
Slightly soluble in
1521 Ethyl furfuryl ether 4114 C7H10O2 water MS 1.449-1.455
Full Ethyl furfuryl ether 13.123 126.15 Soluble 95% 0.982-0.988
Colourless liquid; Sweet, spicy
Furfuryl ethyl ether aroma 150° 1
86 6270-56-0
119
120
Slightly soluble in
water; Soluble in SC: 6-7% Di-(2-
pentane, diethyl methyl-3-furyl)
1524 Furfuryl 2-methyl-3-furyl disulfide 4119 C10H10O2S2 ether HNMR IR 1.581-1.587 disulfide
Full Furfuryl 2-methyl-3-furyl disulfide 226.32 Soluble 90% 1.277-1.283
3-[2-Furanylmethyl)dithio]-2-
methylfuran;(2-Methyl-3-furyl)furfuryl
disulfide;2-Methyl-3-[(2-furanylmethyl)- Colourless liquid; Strong,
dithio]furan;3-(Furfuryldithio)-2-ethylfuran sulfurous aroma 294° 3
86 109537-55-5
Soluble in ethyl
acetate, triacetin,
Practically
1525 3-[(2-Methyl-3-furyl)thio]-2-butanone 4056 C9H12O2S insoluble in water HNMR MS 1.510-1.516
Full 3-((2-Methyl-3-furyl)thio)-2-butanone 13.190 184.25 Soluble 99% 1.104-1.110
3-[(2-Methyl-3-furanyl)sulfanyl]-2-
butanone;3-(2-Methyl-3-furylthio)-2-
butanone;3-[(2-Methyl-3-furyl)sulfanyl]-2- Colourless liquid; Spicy, floral
butanone aroma 70° (0.75 mm Hg) 1
86 61295-44-1
121
Practically
insoluble in water;
Soluble in diether
O-Ethyl S-(2- ether, ethyl
1526 furylmethyl)thiocarbonate 4043 C8H10O3S acetate HNMR IR MS 1.504-1.510
Full O-Ethyl S-(2-furylmethyl)thiocarbonate 186.23 Soluble 99% 1.167-1.173
O-Ethyl S-(2-
furanylmethyl)thiocarbonate;Ethoxy
carbonyl furfurylthiol;O-Ethyl S-(furan-2- Colourless liquid; Spicy, floral
ylmethyl) thiocarbonate aroma 130-135° 5
86 376595-42-5
Insoluble in water,
1495 2,3-Dimethylbenzofuran 3535 C10H10O Soluble in fats NMR 1.554-1.563
Full 2,3-Dimethylbenzofuran 13.074 146.19 Soluble 97% 1.031-1.037
Clear to yellow liquid; Nutty 96-98° (15 mm
spicy aroma Hg) 1
86 3782-00-1
1496 2,4-Difurfurylfuran 4095 C14H12O3 Insoluble in water NMR (13C) mp: 153°
Full 2,4-Difurfurylfuran 13.107 228.24 Soluble 95%
Colourless solid; Floral, fruity
aroma
86 64280-32-6
Insoluble in water;
122
1498 2-Methyl-3(2-furyl)acrolein 2704 C8H8O2 Soluble in oils NMR 1.567-1.573

Full 3-(2-Furyl)-2-methylprop-2-enal 13.046 136.15 Solube 96% 1.097-1.103
2-Methyl-3-(2-furyl)propenal;alpha-
Methylfurylacroleine;Furfurylidene-2- Pale yellowish liquid; Mild,
propanal warm, cinnamon-like aroma 225° 3
86 874-66-8
Insoluble in water;
1500 3-(5-Methyl-2-furyl)-butanal 3307 C9H12O2 Soluble in oils NMR 1.575-1.581
Full 3-(5-Methyl-2-furyl) butanal 13.058 152.19 Soluble 98% 1.006-1.012
Colourless liquid; Vegetable, 88-91° (12 mm
3-(5-Methyl-2-furyl)-butyraldehyde fruity aroma Hg) 3
86 31704-80-0
Insoluble in water;
1501 2-Furfurylidenebutyraldehyde 2492 C9H10O2 Soluble in oils NMR 1.570-1.576
Full Furfurylidene-2-butanal 13.043 150.18 Soluble 98% 1.057-1.063
3(2-Furyl)-2-ethylacrolein;2-Ethyl-3(2-
furyl)-2-propenal;2-Ethyl-3(2- Pale yellowish liquid; Mild,
furyl)acrolein warm, vegetable-like aroma 240° 3
86 770-27-4
1502 2-Phenyl-3-(2-furyl)prop-2-enal 3586 C13H10O2 Insoluble in water NMR mp: 56-57°
123
Full 3-(2-Furyl)-2-phenylprop-2-enal 13.137 198.22 Soluble 99%

2-Furfurylidenephenylacetaldehyde White solid; Berry aroma
86 65545-81-5
Slightly soluble in
water; Soluble in
propylene glycol,
1506 3-Acetyl-2,5-dimethylfuran 3391 C8H10O2 most fixed oils NMR 1.488-1.490
Full 3-Acetyl-2,5-dimethylfuran 13.066 138.17 Soluble 99% 1.037-1.039
Clear to yellow liquid;
Powerful, slightly roasted,
2,5-Dimethyl-3-acetylfuran nutty aroma 83° (11 mm Hg) 1
86 10599-70-9
Slightly soluble in
1512 Pentyl 2-furyl ketone 3418 C10H14O2 water NMR 1.490-1.496
Full 2-Hexanoylfuran 13.070 166.22 Soluble 99% 0.992-0.998

Colourless to yellow liquid; 65-67° (0.5 mm
2-Furyl pentyl ketone Apricot, peach-like aroma Hg) 1
86 14360-50-0
124
1518 Propyl 2-furanacrylate 2945 C10H12O3 Insoluble in water NMR

1.071-1.077
Full Propyl 3(2-furyl)prop-2-enoate 13.047 180.2 Soluble 97% (20°)
2-Propenoic acid, 3-(2-furanyl)-, liquid; Light strawberry, pear-
propyl ester;Propyl 3-(2-furyl)acrylate like aroma 119° (7 mm Hg) 5
86 623-22-3
SC: 1-3% 4-
Hydroxy-2,5-
dimethyl-3(2H)-
2,5-Dimethyl-3-oxo-(2H)-fur-4-yl furanone and 1-
1519 butyrate 3970 C10H14O4 Insoluble in water NMR 1.467-1.473 3% Butyric acid
4-Butyroxy-2,5-dimethyl-3(2H)-
Full furanone 198.22 Soluble 93% 1.095-1.103
liquid; Spicy, sweet aroma 287° 5
86 114099-96-6
Practically
insoluble to
2103 (E)-Ethyl 3-(2-furyl)acrylate 4541 C9H10O3 insoluble in water MS 1.542-1.548
Full Ethyl (2E)-3-(furan-2-yl)prop-2-enoate 166.17 Soluble 95% 1.090-1.096
Ethyl(E)-3-(2-furyl)-2-propenoate Viscous liquid; Sweet aroma 230-233°
86 53282-12-5
Practically
125
insoluble to
2104 di-2-Furylmethane 4540 C9H8O2 insoluble in water MS 1.501-1.507
1.097-1.103
Full 2,2'-Methanediyldifuran 148.16 Soluble 95% (20°)
Colourless clear liquid; Rich
di-2-Furyl methane roasted aroma 194-195°
86 1197-40-6
Practically
insoluble to
2105 2-Methylbenzofuran 4543 C9H8O insoluble in water MS 1.548-1.560
Full 2-Methyl-1-benzofuran 132.16 Soluble 95% 1.052-1.057
Colourless liquid; Burnt
2-Methyl benzo(b)furan phenolic aroma 197-198°
86 4265-25-2
REVISIONS TO EXISTING FLAVOUR SPECIFICATIONS
126
JECFA
No. Name FEMA Chemical Formula Solubility ID test R.I. (20°) Other requirements
Solubility in
Physical form;
Synonyms COE Odour B.P. ° Acid value Information required
Sessio
n CAS
433 l-menthyl l-lactate 3748 C13H24O3 IR Melting point c.25°
(1R,2S,5R)-2-Isopropyl-5-methylcyclohexyl
Full (2S)-2-hydroxypropanoate 228.33 97%
colourless liquid or white
crystalline solid with a
weak chamomile or
tobacco odour 142° (5 mmHg) 2
86 61597-98-6
619 l-malic acid Melting point: 100°;
Fumaric Acid: max. 1.0%;
Heavy Metals: max 10
ppm; Maleic acid: max.
0.05%; Residue on
soluble in water and Ignition: max 0.1 %; Water
alcohol; 1 gm in 0.8 Insoluble Matter: max.
2655 C4H6O5 ml water IR 0.1%
Full 2-Hydroxybutanedioic acid 134.09 1 g in 1.4 ml alcohol 99%

White crystalline powder,
granules, or needles;
Acid taste; odourless or
having a very faint
caramellic acrid odour
Hydroxysuccinic acid;2-Hydroxybutanedioic acid 17 and a tart aciduous taste
86
m.p.: 200-204º
2123 Glutamyl-valyl-glycine 4709 C12H21N3O6 Soluble in water MS, HNMR (decomposition)
(2S)-2-Amino-5-({(2S)-1-[(carboxymethyl)amino]-3-
Full methyl-1-oxobutan-2-yl}amino)-5-oxopentanoic acid 303.31 Practically insoluble >95%
Solid off-white powder;
Light savoury almost
L-gamma-glutamyl-L-valyl-glycine 38837-70-6 yeastlike aroma
86
SPECTRA OF CERTAIN FLAVOURING AGENTS
433
619
974
975
980
981
2123 HNMR 2123 MS
2235 HNMR
2235 C13 NMR
2235 IR 2235 MS
2236 HNMR
2236 C13NMR
2236 IR 2236 MS
2237 HNMR
2237 C13NMR
2237 IR 2237 MS
2238 C13NMR 2238 HNMR
2238 MS 2239 C13NMR
2239 HNMR 2239 MS

2240 C13NMR 2240 HNMR
2241 C13NMR 2241 HNMR
2241 MS
2242 MS
2243 HNMR 2243 MS
2244 IR
2244 HNMR
2244 MS 2246 IR
2246 MS 2247 MS
2248 MS 2249 HNMR
2249 MS
2249 IR
2251 HNMR
2250 HNMR
2251 MS
2251 IR
2252 MS
2252 IR
2253 HNMR
2253 C13NMR
2253 MS 2254 C13NMR
2254 MS
2254 HNMR
2255 HNMR 2255 MS

CORRIGENDUM
The following requests for corrections, reported to the JECFA secretariats, were evaluated
by the 86th JECFA meeting and found to be necessary. These corrections, however, will
only be made in the electronic versions and in the on-line database.
Food additive Original text New text Additional

information
Calcium disodium CAS No. 662-33-9 CAS No. 62-33-9 Transcription error
ethylenediaminetetr
aacetate (INS 385)
Monograph 1 (2006)
Chlorophyllins, Accurately weigh Accurately weigh Correction
copper complexes about 1 g of the about 1 g of the
sodium and sample and dissolve sample and mix in 20
potassium salts (INS in 20 ml of arachid ml of arachid oil….
141(ii)) oil….
Monograph 5 (2008)
Test for ”Free
ionisable copper”
Curcumin (INS: The criteria for several Acetone: Not more Improves readability
100(i)) residual solvents are than 30 mg/kg
Monograph 1 (2006) listed under the Hexane: Not more It was unclear
heading “Residual than 25 mg/kg whether the criterion
solvents” (see Fig. 1). Methanol: Not more “Not more than
than 50 mg/kg 50 mg/kg” extended to
Ethanol: Not more methanol, ethanol,
than 50 mg/kg isopropanol and ethyl
Isopropanol: Not more acetate.
than 50 mg/kg
Ethyl acetate: Not
more than 50 mg/kg
Ethyl acetoacetate CAS No. 1648615 CAS No. 6413-10-1 Transcription error
ethyleneglycol ketal
JECFA No: 1969
JECFA 73 (2010)
Ethyl 2-methyl CAS No. 28959-02-6 CAS No. 39255-32-8 Wrong CAS number
pentanoate
JECFA No: 214
JECFA 55 (2000)
cis-3-Hexen-1-ol 98.0% (sum of (Z) and 98.0% (sum of (Z) and Transcription error
JECFA No.: 315 (E) isomers, =<92.0% (E) isomers, =>92.0%
JECFA 51 (1998) (Z)) (Z))
Monosodium L- CAS No. 142-47-2 CAS No. 6106-04-3 Wrong CAS number
glutamate (INS: 621)
Monograph 1 (2006)
Myrcene Specific gravity: Specific gravity: Transcription error
JECFA No.: 1327 0.789–1.793 0.789–0.793
JECFA 63 (2004)
Food additive Original text New text Additional

information
Polyoxyethylene (20) CAS No. 9005-07-6 CAS No. 9005-67-8 Wrong CAS number
sorbitan
monostearat
(Polysorbate 60)
(INS 435)
Monograph 16
(2014)
Sodium aluminium Within the assay, the Within the assay, the Transcription error
silicate (INS 554) limits for silicon limits for silicon
Monograph 20 dioxide, aluminium dioxide, aluminium
(2017) oxide and sodium oxide and sodium
oxide are expressed oxide are expressed
“on dried basis”. “on ignited basis”.
Silicon dioxide, CAS No. 112696-00-8 CAS No. 112926-00-8 Transcription error
amorphous (INS (hydrated silica) (hydrated silica)
551) Pyrogenic silica is Pyrogenic silica is Transcription error
Monograph 20 produced in an produced in an
(2017) essentially anhydrous essentially anhydrous
state, whereas the state, whereas the
wet process products wet process products
are obtained as are obtained as
hydrates or contain hydrates or contain
surface absorbed surface adsorbed
water. water.
Sodium thiosulfate CAS No. 7772-98-7 CAS No. 10102-17-7 CAS No. 7772-98-7
(INS 539) refers to the
Monograph 1 (2006) anhydrous form. The
specifications in the
monograph refer to
the pentahydrate
form.
Brown HT and its Text in the Table 1 See Table 1, below
aluminium lake “Values for synthetic
(FAO JECFA colours for use in
Monographs 19, 82nd performing tests for
meeting, 2016) colouring matters
content by
spectrophotometry”
Fast Green FCF Chemical structure in
(FAO JECFA Table 1 “Values for
Monographs 19, synthetic colours for
82nd meeting, 2016) use in performing
tests for colouring
matters content by
spectrophotometry”
CAS: Chemical Abstracts Service; INS: International Numbering System for Food Additives; No.: number
Bolding and underlining for clarity only. This formatting will not be shown in the online database.
The criteria for several residual solvents are listed under the heading “Residual solvents” (see Fig. 1)
Figure1: Residual solvent criteria for curcumin as displayed in Monograph 1, 2006
Table 1
Replacement of the text for the spectrophotometric data for Brown HT and its aluminium lake
originally published in “Table 1. Values for synthetic colours for use in performing tests for
Colouring Matters Content by Spectrophotometry” (FAO JECFA Monographs 19, 82nd meeting, 2016)
JECFA Sample
name weight Structure Spectral data Visible absorption spectrum
Brown 245.6 Water, pH 7
HT mg λmax = 464 1.2
A = 0.9957
1
Spec abs = 403
0.8
a = 40.3 Water pH 7
0.6
0.4
Water
λmax = 464 0.2
A = 0.9804 0
350 450 550 650 750
Spec abs = 397
a = 39.7 Wavelength (nm)
0.04 N AmAc
λmax = 461
A = 0.9206
Spec abs = 373
a = 37.3
Brown 53.3 mg Straight colour
HT (blue)
1.2
Aluminiu 0.04 N AmAc 0.04 N AmAc
m Lake 1
λmax = 461
Absorbance
0.8
A = 0.9206
0.6
Lake (red) 0.4
0.04 N AmAc 0.2

λmax = 458 0
A = 1.0451 350 450 550 650 750
Wavelength (nm)
ANNEX I: SUMMARY OF RECOMMENDATIONS FROM THE 86th JECFA
JOINT FAO/WHO EXPERT COMMITTEE ON FOOD ADDITIVES

Eighty-sixth meeting
Geneva, 12–21 June 2018
SUMMARY AND CONCLUSIONS

Issued 3 July 2018
A meeting of the Joint FAO/WHO Expert Committee on Food Additives (JECFA) was held in
Geneva, Switzerland, from 12 to 21 June 2018. The purpose of the meeting was to evaluate
certain food additives (including flavouring agents).
Dr A. Mattia, Center for Food Safety and Applied Nutrition, United States Food and Drug
Administration, served as Chairperson, and Dr. Richard Cantrill, Canada, served as Vice-
Chairperson.
Dr M. Lipp, Office for Food Safety, Food and Agriculture Organization of the United Nations, and
Dr A. Tritscher, Department of Food Safety and Zoonoses, World Health Organization, served as
Joint Secretaries.
The present meeting was the eighty-sixth in a series of similar meetings. The tasks before the
Committee were (a) to undertake safety evaluations of certain food additives (including flavouring
agents); and (b) to review and prepare specifications for certain food additives (including flavouring
agents).
The Committee evaluated the safety of eight food additives, revised the specifications for 19 other
food additives (including 16 modified starches), evaluated 69 flavouring agents according to the
revised Procedure for the Safety Evaluation of Flavouring Agents and revised the specifications for
three flavouring agents.
The report of the meeting will be published in the WHO Technical Report Series. Its presentation will
be similar to that of previous reports – namely, general considerations, comments on specific
substances and recommendations for future work. An annex will include detailed tables (similar to
the tables in this report) summarizing the main conclusions of the Committee in terms of acceptable
daily intakes and other toxicological, dietary exposure and safety recommendations. Information on
the specifications for the identity and purity of certain food additives (including flavouring agents)
examined by the Committee will also be included.
The participants in the meeting are listed in Annex 1. Items of a general nature that the Committee
would like to disseminate quickly are included in Annex 2. Future work and recommendations are
listed in Annex 3.
Toxicological and dietary exposure monographs on most of the substances that were considered
will be published in WHO Food Additives Series No. 77. New and revised specifications for the
identity and purity of the compounds will be published in FAO JECFA Monographs 22.
More information on the work of JECFA is available at:
http://www.fao.org/food/food-safety-quality/scientific-advice/jecfa/en/
and
http://www.who.int/foodsafety/areas_work/chemical-risks/jecfa/en/
The issuance of this document does not constitute formal publication. The document may, however,
be freely reviewed, abstracted, reproduced or translated, in whole or in part, but not for sale or use
in conjunction with commercial purposes.
Food additives evaluated toxicologically and assessed for dietary exposure

Acceptable daily intakes (ADIs) and other
Food additive Specifications toxicological and dietary exposure conclusions
Anionic N, Ta The Committee was unable to complete the
methacrylate evaluation of AMC. While the copolymer itself is not
copolymer (AMC) of health concern, genotoxicity concerns remains for
the residual monomer methacrylic acid. The
specifications were made tentative pending the
completion of the safety evaluation of AMC.
Basic methacrylate N The Committee established an ADI “not specified”
copolymer (BMC) for basic methacrylate copolymer.
The Committee concluded that the use of BMC that
complies with the specifications established at the
current meeting is not of safety concern when the food
additive is used as a coating or glazing agent for solid
food supplements and for foods for special medical
purposes and micronutrient encapsulation for food
fortification. The NOAELs for BMC ranged from 750-
2000 mg/kg bw per day which were the highest doses
tested.
The Committee evaluated exposure to BMC for the
copolymer and its monomers (n-butyl methacrylate, 2-
(dimethylamino)ethyl methacrylate and methyl
methacrylate). Estimated exposures to BMC range
from 3.0 to 135 mg/kg bw per day. The total
monomeric content of BMC is less than 0.3%. The
Committee concluded that the toxicological data on
the residual monomers do not give rise to concerns
when taking into account the low dietary exposures.
Erythrosine Rb The Committee concluded that the new data that
have become available since the previous
evaluation of erythrosine do not give reason to
revise the ADI and confirmed the previous ADI of
0–0.1 mg/kg bw.
The Committee noted that the dietary exposure
estimate for erythrosine of 0.09 mg/kg bw per day
(95th percentile for children) was close to the upper
bound of the ADI. Given that this estimate of exposure
is for children and it is a high percentile for consumers
only, such a level is unlikely to occur every day over a
lifetime. Therefore, the Committee concluded that
dietary exposures to erythrosine for all age groups do
not present a health concern.

Indigotine Rb The Committee considered the new data that had
become available since the previous evaluation as
well as previously evaluated studies and concluded
that there are no reasons to revise the ADI and
confirmed the previous ADI of 0–5 mg/kg bw.
The Committee noted that the conservative dietary
exposure estimate of 0.8 mg/kg bw per day (95th
percentile for children and toddlers) is less than the
upper bound of the ADI of 0–5 mg/kg bw. The
Committee concluded that dietary exposure to
indigotine for all age groups does not present a health
concern.
Lutein Rc,d Free lutein, lutein esters and free zeaxanthin including
meso-zeaxanthin are biochemically and
toxicologically equivalent. At the present meeting the
Committee concluded that there were sufficient
toxicological data to complete a safety assessment of
lutein and lutein esters from Tagetes erecta, synthetic
zeaxanthin and meso-zeaxanthin. Free lutein, lutein
esters and free zeaxanthin and meso-zeaxanthin are
substances of low toxicity for which no adverse effects
have been observed in a broad range of toxicological
studies in laboratory animals and clinical studies in
humans.
Based on the absence of toxicity in a wide range of
studies, the Committee established a group ADI
"not specified" for lutein from Tagetes erecta,
lutein esters from Tagetes erecta and zeaxanthin
(synthetic).
Meso-zeaxanthin was not included in this group ADI,
as specifications are not currently available.
The group ADI of 0-2 mg/kg bw for lutein from
Tagetes erecta and zeaxanthin (synthetic) was
withdrawn.
Neutral N, T The Committee established an ADI “not specified”
methacrylate for NMC. The ADI “not specified” was made
copolymer (NMC) temporary because the specifications are
tentative.
The Committee concluded that the use of NMC that
complies with the specifications established at the
current meeting is not of safety concern when the food
additive is used as a coating or glazing agent for solid
food supplements and for foods for special medical
purposes. The NOAELs for NMC ranged from 454–
2000 mg/kg bw per day, and these were the highest
doses tested.
The Committee evaluated exposure to NMC for the
copolymer and its monomers (methyl methacrylate
and ethyl acrylate). Estimated exposures to NMC
range from 5.8 to 86 mg/kg bw per day. The total
monomeric content of NMC is less than 0.01%.
Toxicological data on the residual monomers do not

give rise to concerns when taking into account the low
dietary exposures.
Sorbitol syrup - Sorbitol syrup (INS 420(ii)) is currently included in the
Codex General Standard for Food Additives (GSFA)
although it has not been assigned an ADI or
determined, on the basis of other criteria, to be safe.
The Committee was therefore requested to consider
the previous evaluations of sorbitol, hydrogenated
glucose syrups and other relevant substances, and
advise on the need for a separate evaluation of
sorbitol syrup or if the ADI “not specified” for sorbitol
is also applicable for sorbitol syrup.
Based on the similarity of the chemical constituents of
sorbitol syrup to the previously evaluated sorbitol,
maltitol syrup and polyglycitol syrup, the Committee
concluded that there is no need for a separate
evaluation of sorbitol syrup and established an
ADI “not specified” for sorbitol syrup.
Spirulina extract N, T The Committee established a temporary ADI “not
specified” for spirulina extract. The ADI was based
on the absence of toxicity in repeated-dose animal
studies with spirulina extract and dried spirulina. The
ADI “not specified” was made temporary due to
the tentative nature of the specifications.
Expressed as phycocyanins, estimated dietary
exposure from the use of spirulina extract as a food
colour based on the Budget method and exposure to
spirulina extract and dried spirulina from other dietary
sources, including food ingredients, dietary
supplements, and coatings of food supplements was
190 mg/kg bw for adults (60 kg/person) and 650
mg/kg bw for a child (15 kg/person). The Committee
concluded that this dietary exposure does not present
a health concern.
- : no specifications prepared; N: new specifications; R: existing specifications revised; T: tentative
specifications
a
The specifications were made tentative pending the completion of the safety evaluation of AMC.
b
At the current meeting, high-performance liquid chromatographic (HPLC) methods were added for
determining subsidiary colouring matters and organic compounds other than colouring matters.
The method of assay was changed to visible spectrophotometry, and spectrophotometric data
were provided for the colour dissolved in water.
c
The specifications for lutein esters from Tagetes erecta and zeaxanthin (synthetic) were
maintained.
d
At the current meeting, the identity test for melting range was deleted, the identity tests for
carotenoids and spectrophotometry were updated, the test for propylene glycol was incorporated
verbatim and the previous reference removed, and the method of assay was updated.
Food additives considered for specifications only

Food additive Specifications
Cassia gum Ra
Citric and fatty acid esters of glycerol R, Tb
Glycerol ester of wood rosin Rc
Modified starches Rd, T
R: existing specifications revised; T: tentative specifications
a
The Committee, at its current meeting, received analytical methods and included the most
suitable validated method in the specifications monograph. However, this method uses
chloroform for the extraction of anthraquinones. Extraction with n-hexane and diethyl ether
resulted in poor recovery of anthraquinones. The Committee recommends that the JECFA
Secretariat be notified if an alternative extraction solvent is identified. The specifications were
revised and the tentative status was removed.
b
The Committee did not receive a replacement method for the obsolete packed column gas
chromatographic method for the determination of total citric acid, in its specifications
monograph. The Committee noted further that the method for total glycerol still uses chloroform.
The Committee encouraged the submission of a method for total glycerol that eliminates the
use of chloroform. Specifications were revised and made tentative pending the availability of
data. Specifications will be withdrawn if suitable information is not provided by December 2019.
c
The Committee received information on the manufacture of GEWR from the rosin obtained from
the stumps of two additional species namely Pinus halepensis and Pinus brutia as source
materials. Recognizing the natural variability of the composition of wood rosin, the Committee
removed the restriction to certain pine species within the specifications. Since the specifications
monograph for GEWR does not contain an assay, the Committee recommended that the JECFA
Secretariat be notified upon the development and validation of an appropriate assay. The
existing specifications were revised.
d
The Committee reviewed data on the method of manufacture, identity, and purity of all 16
modified starches. Based on the information received, and available information the Committee
noted that:
• All processes are performed under similar manufacturing conditions and result in minor
chemical modifications. Given the chemical and physical similarities of modified starches, the
Committee at previous meetings considered the application of a read-across approach to be
appropriate for the toxicological evaluation of these substances.
• All 16 modified starches had been assigned an ADI of “not specified”.
• All modified starches can be additionally bleached or fragmented; therefore revision in the
specifications of bleached or fragmented starches would imply the revision of all 16
monographs;
• Microbiological specifications were not present in the existing specifications for all modified
starches.
• Several specifications were common to all modified starches (such as for heavy metals
impurities content and microbiological considerations). Revision of those common
specifications would affect all 16 monographs;
• As a result of the wide range of products manufactured, the identification tests required to
unambiguously chemically characterize each modified starch in individual specifications may
be cumbersome, potentially unavailable, and unlikely to reflect market requirements.
• It may not be possible to publish identification tests based on market requirements without
unduly revealing proprietary information.
• Based on the points noted above, individual specifications for several modified starches may
remain tentative for an indefinite period or may need to be withdrawn.
The Committee therefore recommended that a new approach to the specifications monographs
should be introduced to account for the chemical similarity between all modified starches, their
functional diversity, the variety of chemicals used in their manufacture, and the corresponding
diversity of impurities. The Committee recommended that all modified starches be included in a
modular monograph titled ‘Modified Starches’ that contains common requirements [General
specifications for modified starches] consisting of specifications that apply to all 16 modified
starches (INS 1400, 1401, 1402, 1403, 1404, 1405, 1410, 1412, 1413, 1414, 1420, 1422, 1440,
1442, 1450, 1451), and annexes with specifications applicable to each individual modified starch
based on the treatment(s) received. The Committee drafted a new modular specifications
monograph titled “Modified starches” consisting of an explanatory introduction, “General
specifications for modified starches,” and eight annexes. The new modular specifications
monograph for modified starches is printed in FAO Monograph 22, and will replace the 16
existing individual specifications for modified starches (INS 1400, 1401, 1402, 1403, 1404, 1405,
1410, 1412, 1413, 1414, 1420, 1422, 1440, 1442, 1450, 1451).
The specification for lead included in the General specifications be decreased from 2 mg/kg to
0.2 mg/kg. The limit of lead for starch sodium octenylsuccinate for use in infant formula and
formula for special medical purposes intended for infants was set to 0.1 mg/kg in the General
specifications.
The methods for the determination of free adipic acid and adipate groups, residual vinyl acetate,
free octenyl succinic acid and octenyl succinate esters were revised and a method for the
determination of propylene chlorohydrins was added.
Flavouring agents evaluated by the revised Procedure for the Safety Evaluation of
Flavouring Agents
A. Alicyclic primary alcohols, aldehydes, acids and related esters

Conclusion based on
current estimated
Flavouring agent No. Specifications dietary exposure
Structural class I
Mixture of 1-Vinyl-3- 2253 N No safety concern
cyclohexenecarbaldehyde and 4-Vinyl-
1-cyclohexenecarbaldehyde
p-Mentha-1,8-dien-7-ol 974 N No safety concern
p-Mentha-1,8-dien-7-yl acetate 975 N No safety concern
Formyl-6,6-dimethylbicyclo[3.1.1]hept- 980 N No safety concern
2-ene
Myrtenol 981 N No safety concern
Myrtenyl acetate 982 M No safety concern
Structural class II
(1-Methyl-2-(1,2,2- 2254 N No safety concern
trimethylbicyclo[3.1.0]hex-3-
ylmethyl)cyclopropyl)methanol
Structural class III
(±)-Bicyclo[2.2.1]hept-5-ene-2- 2255 N No safety concern
carboxylic acid, ethyl ester
Flavouring agent excluded at Step 1
of the Procedure
p-Mentha-1,8-dien-7-al 973 M Genotoxicity data for
(Perillaldehyde) p-mentha-1,8-dien-7-
al raise concerns for
potential genotoxicity
N: new specifications
M: existing specifications maintained;
B. Carvone and structurally related substances
Conclusion based on
current estimated
Flavouring agent No. Specifications dietary exposure
Structural class I
Pinocarvyl isobutyrate 2242 N No safety concern
Carvyl palmitate 2243 N No safety concern
6-Hydroxycarvone 2244 N No safety concern
Flavouring agents not evaluated according to the revised Procedure
(+)-Carvone 380.1 M The Committee did not re-
evaluate (+)-carvone (No.
380.1) according to the
revised Procedure given
the lack of information on
the oral exposure from all
sources and the need to
review the ADI.
A review of the ADI is
recommended based on
the evaluation of all
biochemical and
toxicological data. Also,
data are needed for an
exposure assessment for
oral exposure to (+)-
carvone from all sources
to complete the evaluation
for (+)-carvone.
(-)-Carvone 380.2 M The Committee did not re-
evaluate (-)-carvone (No.
380.2) according to the
revised Procedure given
the lack of information on
the oral exposure from all
sources and the lack of
toxicological data.
M: existing specifications maintained; N: new specifications
C. Furan-substituted aliphatic hydrocarbons, alcohols, aldehydes, ketones, carboxylic acids

and related esters, sulfides, disulfides and ethers
Conclusion based on
current estimated dietary
Flavouring agent No. Specifications exposure
2-Pentylfuran 1491 Ma No safety concern
a
2-Heptylfuran 1492 M No safety concern
a
2-Decylfuran 1493 M No safety concern
a
3-Methyl-2-(3-methylbut-2-enyl)- 1494 M No safety concern
furan
Conclusion based on
2,3-Dimethylbenzofuran 1495 Ma No safety concern
2,4-Difurfurylfuran 1496 Ma No safety concern
a
3-(2-Furyl)acrolein 1497 M No safety concern
2-Methyl-3(2-furyl)acrolein 1498 Ma No safety concern
a
3-(5-Methyl-2-furyl)prop-2-enal 1499 M No safety concern
a
3-(5-Methyl-2-furyl)butanal 1500 M No safety concern
a
2-Furfurylidene-butyraldehyde 1501 M No safety concern
a
2-Phenyl-3-(2-furyl)prop-2-enal 1502 M No safety concern
2-Furyl methyl ketone 1503 Ma No safety concern
2-Acetyl-5-methylfuran 1504 Ma No safety concern
a
2-Acetyl-3,5-dimethylfuran 1505 M No safety concern
3-Acetyl-2,5-dimethylfuran 1506 Ma No safety concern
2-Butyrylfuran 1507 Ma No safety concern
a
(2-Furyl)-2-propanone 1508 M No safety concern
2-Pentanoylfuran 1509 Ma No safety concern
1-(2-Furyl)butan-3-one 1510 Ma No safety concern
a
4-(2-Furyl)-3-buten-2-one 1511 M No safety concern
Pentyl 2-furyl ketone 1512 Ma No safety concern
Ethyl 3-(2-furyl)propanoate 1513 Ma No safety concern
a
Isobutyl 3-(2-furan)propionate 1514 M No safety concern
Isoamyl 3-(2-furan)propionate 1515 Ma No safety concern
Isoamyl 3-(2-furan)butyrate 1516 Ma No safety concern
a
Phenethyl 2-furoate 1517 M No safety concern
Propyl 2-furanacrylate 1518 Ma No safety concern
2,5-Dimethyl-3-oxo-(2H)-fur-4-yl 1519 Ma No safety concern
butyrate
Furfuryl methyl ether 1520 Ma No safety concern
a
Ethyl furfuryl ether 1521 M No safety concern
Difurfuryl ether 1522 Ma No safety concern
2,5-Dimethyl-3-furanthiol acetate 1523 Ma No safety concern
a
Furfuryl 2-methyl-3-furyl disulfide 1524 M No safety concern
3-[(2-Methyl-3-furyl)thio]-2-butanone 1525 Ma No safety concern
O-Ethyl S-(2- 1526 Ma No safety concern
furylmethyl)thiocarbonate
(E)-Ethyl 3-(2-furyl)acrylate 2103 Ma No safety concern
di-2-Furylmethane 2104 Ma No safety concern
2-Methylbenzofuran 2105 Ma No safety concern
M: existing specifications maintained
a
The text indicating that the safety evaluation for these flavouring agents had not been completed
was removed from the specifications and the specifications were maintained as full
D. Linear and branched-chain aliphatic, unsaturated, unconjugated alcohols, aldehydes,

acids and related esters
Conclusion based on
Structural class I
trans-6-Octenal 2240 N No safety concern
2,6-Dimethyl-5-heptenol 2241 N No safety concern
E. Maltol and related substances

Conclusion based on
Structural class II
Maltol 1480 M No safety concerna
Ethyl maltol isobutyrate 2252 N No safety concern
a
The previously established ADI for maltol was withdrawn by the Committee.
F. Menthol and structurally related substances

Conclusion based on
Structural class I
Menthyl formate 2246 N No safety concern
Menthyl propionate 2247 N No safety concern
l-Menthyl butyrate 2248 N No safety concern
dl-Isomenthol 2249 N No safety concern
Dimenthyl glutarate 2250 N No safety concern
Menthol 427 M No safety concerna
(±)-2-[(2-p- 2251 N No safety concern
Menthoxy)ethoxy]ethanol
a
The ADI of menthol of 0–4 mg/kg bw established at the fifty-first meeting was maintained.
G. Miscellaneous nitrogen-containing substances
Conclusion based on
2-(((3-(2,3-Dimethoxyphenyl)-1H- 2235 N No safety concern
1,2,4-triazol-5-
yl)thio)methyl)pyridine
S)-1-(3-(((4-Amino-2,2-dioxido-1H- 2236 N No safety concern
benzo[c][1,2,6]thiadiazin-5-
yl)oxy)methyl)piperidin-1-yl)-3-
methylbutan-1-one
2-(4-Methylphenoxy)-N-(1H- 2237 N No safety concern
pyrazol-3-yl)-N-(thiophen-2-
ylmethyl)acetamide
H. Saturated aliphatic acyclic branched-chain primary alcohols, aldehydes, and acids

Conclusion based on
Structural class I
8-Methyldecanal 2238 N No safety concern
8-Methylnonanal 2239 N No safety concern
Flavouring agents considered for specifications only

Flavouring agent No. Specifications
L-menthyl lactate 433 Ra
L-malic acid 619 Rb
Glutamyl-valyl-glycine 2123 Rc
a
The CAS number was changed from 59259-38-0 to 61597-98-6 and the name to L-menthyl L-
lactate.
b
The specification for specific rotation were removed
c
The melting point range was revised
ANNEX 2. GENERAL INFORMATION
This section is deliberately blank

ANNEX 3. FUTURE WORK AND RECOMMENDATIONS
SPECIFIC FOOD ADDITIVES (OTHER THAN FLAVOURING AGENTS)
Anionic Methacrylate Copolymer
The Committee noted that there were insufficient data to reach a conclusion on the genotoxic
potential of methacrylic acid. Further studies to clarify the in vivo carcinogenic potential are required.
Citric and fatty acid esters of glycerol
The specifications of CITREM were made tentative, requiring a suitable validated method for the
determination of total citric acid content, along with performance characteristics of the method and
data on the total citric acid content in at least five batches of products currently available in
commerce, determined using that method.
The Committee noted that the method for total glycerol still uses chloroform. The Committee
encouraged the submission of a method for total glycerol that eliminates the use of chloroform.
Specifications were revised and made tentative. Specifications will be withdrawn if suitable
information is not provided by December 2019.
Neutral Methacrylate Copolymer
The Committee noted that there was no data submitted for a suitable method of assay. Tentative
specifications for NMC were prepared and made tentative requiring a suitable validated method of
assay.
Spirulina extract
The Committee received limited analytical data on spirulina extract. In order to remove the tentative
designation from the specifications, the following information on the products of commerce is
requested by December 2019:
• Full compositional characterization of commercial products in both liquid and powder forms.
• Full compositional characterization of the aqueous extract before
formulation/standardization.
• Validated analytical methods for identification of the substance with a suitable specificity
(including validation data and representative batch data).
• Validated analytical methods for the determination of the purity of the substance with a
suitable specificity (including validation data and representative batch data).
Modified starches
The Committee requested additional data and a suitable method for the determination of propylene
chlorohydrins in Hydroxypropyl starch (INS 1440) and Hydroxypropyl distarch phosphate (INS 1442)
in order to consider lowering this limit.
The Committee requests suitable microbiological acceptance criteria and supporting data for all
modified starches.
Table 1. The annexes and the modified starches to which they apply along with required information:
ANNEX Modification Starches to which it applies Information required

1 Minor INS 1400: Dextrin roasted A suitable method for dispersion
fragmentation starch; and a method for reducing sugars
INS 1401: Acid treated and data on at least 5
starch; representative batches using the
INS 1402: Alkaline treated method(s) from each of the
starch; fragmentation processes.
INS 1405: Enzyme-treated
starch
All modified starches that are
additionally fragmented.
2 Bleaching INS 1403: Bleached starch Suitable method(s) for the
All modified starches if determination of residual reagents
additionally bleached. and data on at least 5
representative batches using the
method(s).
3 Esterification INS 1410: Monostarch A suitable method for identification
and/or phosphate; of crosslinking and data on at least
crosslinking INS 1412: Distarch 5 representative batches of
with phosphate; crosslinked and non-crosslinked
phosphorus INS 1413: Phosphated starches.
containing distarch phosphate;
compounds INS 1414: Acetylated distarch
phosphate;
INS 1442: Hydroxypropyl
distarch phosphate
4 Acetylation INS 1420: Starch acetate; Currently no additional information
INS 1414: Acetylated distarch required.
phosphate;
INS 1422: Acetylated distarch
adipate; INS 1451:
Acetylated oxidized starch
5 Oxidation INS 1404: Oxidized starch; A suitable method for determination
INS 1451: Acetylated of residual hypochlorite and data on
oxidized starch at least 5 representative batches
using the method.
6 Esterification INS 1450: Starch sodium Currently no additional information
with octenyl octenyl succinate required.
succinic
anhydride
7 Etherification INS 1440: Hydroxypropyl A suitable method for the
with propylene starch; determination of propylene
epoxide INS 1442: Hydroxypropyl chlorohydrin with detection limit
distarch phosphate lower than 0.1 mg/kg and data on at
least 5 representative batches of
Hydroxypropyl starch using the
method
8 Crosslinking INS 1422: Acetylated distarch A suitable method for identification
with adipic adipate of crosslinking and data on at least
anhydride 5 representative batches of
crosslinked and non-crosslinked
starches.
Levels of free adipic acid in at least

5 representative batches
FLAVOURING AGENTS
Carvone and structurally related substances
For (+)-carvone (No. 380.1), the Committee concluded that a review of the ADI is recommended
based on the evaluation of all biochemical and toxicological data. Also, data are needed for an
exposure assessment for the oral exposure to (+)-carvone from all sources.
The ADI for (+)-carvone is maintained pending review of the ADI at a future meeting. The Committee
recommends that the re-evaluation is completed within three years.
For (-)-carvone (No. 380.2), the Committee concluded that toxicological data on (-)-carvone are
necessary. Also, data are needed for an exposure assessment for the oral exposure to (-)-carvone
from all sources
Maltol and related substances
The Committee could not verify the NOEL of 100 mg/kg bw in rats that was used to derive the ADI
of 0–1 mg/kg bw for maltol (No. 1480) during its twenty-fifth meeting because of uncertainties in the
administered dose levels and the effects observed in several studies described in the monograph of
that meeting.
The Committee withdrew the ADI for maltol. The Committee concluded that access to either the
original studies or submission of new data would be needed to reaffirm or amend the ADI.
The ADI for ethyl maltol was maintained.

New_Cover_2016.pdf 1 26/04/2016 15:05:26
20
ISSN 1817-7077
COMPENDIUM
OF FOOD ADDITIVE FAO JECFA Monographs 20
SPECIFICATIONS
86th Meeting 2018
This document contains food additive specification monographs,

analytical methods, and other information prepared at the eighty-fourth
meeting of the Joint FAO/WHO Expert Committee on Food Additives
COMPENDIUM OF FOOD ADDITIVE SPECIFICATIONS

(JECFA), which was held in Geneva, 12–21 June 2018. The specification
monographs provide information on the identity and purity of food
additives used directly in foods or in food production. The main three
objectives of these specifications are to identify the food additive that
has been subjected to testing for safety, to ensure that the additives are
of the quality required for use in food or in processing and to reflect and
encourage good manufacturing practice. This publication and other
C
M
documents produced by JECFA contain information that is useful to all
those who work with or are interested in food additives and their safe use
COMPENDIUM
Y
CM
in food.
OF FOOD ADDITIVE
MY
CY
SPECIFICATIONS
CMY
84th Meeting 2017
FAO / WHO

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84th Meeting 2017

(JECFA), which was held in

Joint FAO/WHO Expert Committee on Food Additives

86th Meeting 2018

Food and Agriculture Organization of the United Nations

ISBN 978-92-5-131100-4 (FAO)

LIST OF PARTICIPANTS ..........................................................................................................VII

JOINT FAO/WHO EXPERT COMMITTEE ON FOOD ADDITIVES, 86th MEETING

Contact and Feedback

SPECIFICATIONS FOR CERTAIN FOOD ADDITIVES

New and revised specifications

Anionic methacrylate copolymer (AMC) (N, T)

ANIONIC METHACRYLATE COPOLYMER

New specifications prepared at the 86th JECFA (2018) and

SYNONYMS E 1207, INS No. 1207, acrylates copolymers, Methyl acrylate,

DEFINITION Anionic methacrylate copolymer is a copolymer comprised of

Chemical name Poly (methyl acrylate-co-methylmethacrylate-co-methacrylic

C.A.S. number 26936-24-3

Chemical formula Poly[(CH2:CHCO2CH3)-co-(CH2:C(CH3)CO2CH3)-co-

The above formula is provided for illustrative purposes; in this

Formula weight 280,000 (weight-average ), 77,000 (number-average)

Assay 9.2 – 12.3 % methacrylic acid units on the dried basis

See description under Tests

DESCRIPTION Commercial form (30% aqueous dispersion) is a low viscosity,

FUNCTIONAL USES Coating agent, glazing agent.

Viscosity (Vol. 4) Not more than 20 mPa•s

Determine viscosity using Brookfield viscometer at 20° and 30 rpm

pH (Vol 4) 2.0 – 3.5

Infrared absorption The infrared absorption spectrum of a dry film of sample

Apply one drop of sample to a glass plate, cover with a water-

Loss on drying 68.5 – 71.5% (110°, 5 h)

Sulfated ash (Vol. 4) Not more than 0.2%

Test 5 g of the sample (Method I)

Methanol (Vol. 4) Not more than 1,000 mg/kg

Residual monomers Methyl acrylate: Not more than 1 mg/kg

Methyl methacrylate: Not more than 3 mg/kg

Methacrylic acid: Not more than 1 mg/kg

See description under TESTS

Lead (Vol. 4) Not more than 1.0 mg/kg in the dispersion

Determine using a method appropriate to the specified level.

Microbiological criteria Total plate count: Less than 1,000 cfu/g

Residual monomers Determined by liquid chromatography (Vol. 4)

Standards and Reagents:

Preparation of mixed standard solutions:

Intermediate mixed standard solution-1:

Intermediate mixed standard solution-2: Dilute 20.0 ml of

Working mixed standard solution:

Preparation of sample solution:

the solution dropwise (precipitation of the polymer should be

Inject separately 20 µl each of working mixed standard solution

Total monomers in the sample (µg/g) = Sum of monomers in the

METHOD OF ASSAY Accurately weigh about 5 g sample and dissolve completely in

Appendix: Infrared spectrum of anionic methacrylate copolymer

BASIC METHACRYLATE COPOLYMER

New specifications prepared at the 86th JECFA (2018) and

SYNONYMS E 1205; INS No. 1205; basic butylated methacrylate copolymer;

DEFINITION Basic Methacrylate Copolymer is a cationic copolymer comprised