ANNEX 5

A PROCEDURE FOR THE SAFETY EVALUATION OF FLAVOURING SUBSTANCES

I.C. Munro, Ph.D., FRCPath

CanTox, Inc.

Mississauga, Ontario

Canada

This paper was considered at the forty-fourth meeting of the

Joint FAO/WHO Expert Committee on Food Additives. The conclusions of

the Committee relating to its consideration of the paper are provided

in the report, which has been published in the WHO Technical Report

Series. This paper was not endorsed by the Committee, and the opinions

expressed therein are those of the author. Comments on the approach

outlined in the paper are invited, which should be addressed to the

WHO Joint Secretary of JECFA, International Programme on Chemical

Safety, 20, Avenue Appia, World Health Organization CH-1211 Geneva 27,

Switzerland.

ANNEX 5

A PROCEDURE FOR THE SAFETY EVALUATION OF FLAVOURING SUBSTANCES

Table of Contents

1. INTRODUCTION

2. CONCEPTS EMPLOYED IN THE SAFETY EVALUATION OF FLAVOURING

SUBSTANCES

2.1 Exposure to flavouring substances through food

2.2 Structure-activity relationships

2.3 Use of toxicity data

3. INTEGRATED METHOD FOR THE SAFETY EVALUATION OF FLAVOURING

SUBSTANCES

3.1 Structure-activity relationships and metabolic fate

3.2 Integrating information on exposure and toxicity

3.3 Evaluation criteria

3.4 Integrating data on the consumption ratio

4. CONCLUSIONS

5. REFERENCES

APPENDICES

Appendix A Estimating distribution of intakes of food ingredients

across the population

Appendix B Substances in reference database by order of structural

class

1. INTRODUCTION

The objective of this paper is to provide a procedure that can be

used by the Joint FAO/WHO Expert Committee on Food Additives (JECFA)

for the safety evaluation of flavouring substances. The procedure

extends principles and procedures recommended by IECFA in the past and

is consistent with international concepts in safety evaluation. These

principles have been discussed by JECFA (JECFA, 1972; 1974b; 1976a;

1978; 1980a,b; 1982a; 1983a; 1984a; 1986; 1987a,b; 1989; 1990a; 1991a;

1992) and have been recapitulated in a report, which outlines criteria

for the safety evaluation of flavouring substances (WHO, 1987).

This paper takes this previous background into account and

provides a scientific basis for assessing the safety of flavouring

substances used in food by presenting a procedure for evaluating this

large group of compounds in a consistent, timely, and appropriate

manner.

2. CONCEPTS EMPLOYED IN THE SAFETY EVALUATION OF FLAVOURING

SUBSTANCES

Criteria for the safety evaluation of flavouring substances have

been put forward by several national and international authorities.

Included among these are JECFA, a special Task Group convened by the

World Health Organization (WHO, 1987), the Committee of Experts on

Flavoring Substances of the Council of Europe (CE), the Commission of

the European Communities' Scientific Committee for Food (SCF, 1991),

the British Industrial Biological Research Association (BIBRA), and

the Flavor and Extract Manufacturers' Association of the United States

Expert Panel (FEXPAN).
In 1987, WHO published a health criteria document entitled

Principles of the Safety Evaluation of Food Additives and

Contaminants in Food, which contains a discussion of principles

related to the safety evaluation of flavouring substances. This report

recapitulated principles previously stated by JECFA on numerous

occasions (JECFA, 1968; 1972; 1974b; 1976a; 1978; 1980a,b; 1982a;

1983a; 1984a; 1986; 1987a,b; 1989; 1990a; 1991a; 1992). Early on, it

was recognized by JECFA that the safety evaluation of flavouring

substances warranted special consideration in light of use patterns

and typically low levels of human exposure (JECFA, 1972). This view

also has been recognized by the (SCF) and has been recorded in their

document entitled Guidelines for the Evaluation of Flavorings for Use

in Foodstuffs. 1. Chemically, Defined Flavoring Substances

(SCF, 1991).

In addition, several organizations including JECFA (WHO, 1987),

FEXPAN (Woods & Doull, 1991), SCF (1991), and the Council of Europe

(CE, 1974, 1981, 1992) have noted that knowledge of structure-activity

relationships and metabolism plays a key role along with exposure in

the safety evaluation of flavouring substances. In this regard, JECFA

(WHO, 1987) has used structure-activity relationships in evaluating

groups of structurally related flavouring substances in a homologous

series where toxicology studies exist on only one or a few members of

the series. Structure-activity relationships can provide a useful

means of assessing substances that lack toxicity data based on

structurally related substances that have adequate toxicity data.

2.1 Exposure to flavouring substances through food

Flavouring substances are used in processed foods and beverages

to impart desirable organoleptic qualities and to provide the specific

flavour profile traditionally associated with certain food products.

Unlike many substances which are added to food to achieve a

technological purpose, the use of flavouring substances is generally

self-limiting and governed by the flavour intensity required to

provide the necessary organoleptic appeal. Thus, flavouring substances

are used generally in low concentrations resulting in human exposures

that are very low.

Estimates of the intake of flavouring substances by the

population typically have involved the acquisition of data on poundage

used in foods. Between 1970 and 1987, the U.S. National Academy of

Sciences/National Research Council (NAS/NRC) conducted, under contract

with the Food and Drug Administration (FDA), a series of poundage

surveys of substances intentionally added to food (NAS, 1978, 1979,

1984, 1989). These surveys obtained information, both from ingredient

manufacturers and from food processors, on the poundage of each

substance committed to the food supply and on the usual and maximum

levels at which each substance was added to foods in each of a number

of broad food categories.

Estimates of flavouring substance intake in the U.S. have been

performed in two ways. One is to assume that the total poundage

reported in food annually is completely consumed by the total

population. Numerous checks using data from independent sources, such

as imports, show that, in general, the reported poundage in the survey

accounted for only 60% of the total used. Much more detailed and

elaborate analyses (see Appendix A) led to the conclusion that it was

conservative but reasonable to assume that each flavouring substance

is consumed by only 10% of the population. A second method of

calculating intake is to combine data on the level of use in specific

food groups with data on food consumption to calculate the intake of

each flavouring substance. Both methods tend to overestimate human

intakes of flavouring substances because they deal with disappearance,

that is the amount presumed to be used in food, and take no account of

losses and waste during food manufacture, storage, preparation and

consumption.

Estimating intake of flavouring substances based on level of use

data, coupled with data on the consumption of foods, typically leads

to substantial overestimates of intake. This is because intake data

for food commodities are stratified into broad categories of food

products (e.g., baked products). Thus, cardamom, ordinarily used only

in certain types of coffee cakes, would be assumed to be present in

all breads, rolls, cakes and pastries In extreme cases, this may lead

to overestimates of intake which are exaggerated several hundred-fold.

Through a series of detailed studies conducted between 1970 and

1987 (see Appendix A) it has become clear that, while there is at

present no perfect way to estimate intake of flavouring substances,

poundage used provides a reasonable basis for calculating intakes. The

intake data reported in this paper rely on annual poundage used in

food and the estimates of intake reflect the assumptions that: i) the

available survey data accounted for only 60% of production; and ii)

the total amount produced is consumed by 10% of the population. Table

1 presents the intake data for 1323 chemically defined flavouring

substances permitted for use in the U.S. calculated in this fashion.

As can be seen from Table 1, most flavouring substances are consumed

in amounts of less than 1 mg/person/day. The data are taken from the

most recent U.S. NAS/NRC survey (NAS, 1989) of poundage used in food.
For reasons previously stated, it can be assumed that these intake

estimates are overestimated.

Another important factor to consider in the evaluation of human

exposure to flavouring substances is the extent to which flavouring

substances intentionally added to foods also occur naturally in the

food supply. The natural presence of flavouring substances in food is

of course not necessarily indicative of safety. For many flavouring

substances that occur naturally in foods, such natural occurrence,

rather than intentionally added use, is the principal component of

human exposure. The comparison of natural occurrence to intentional

addition has been expressed as the consumption ratio (CR) (Stofberg &

Kirschman, 1985). A CR of greater than one indicates food predominance

(i.e. the flavouring substance is consumed at a higher level from

foods than as an added substance). A CR greater than 10 indicates an

almost insignificant contribution (-10%) of the flavouring substance

as a food additive to the total intake (Stofberg & Grundschober,

1987). Stofberg and Grundschober (1987) calculated that out of 499

flavouring substances, 415 (83%) were food predominant (i.e. CR > 1)

and 309 (62%) made an insignificant contribution to the food supply

(i.e. CR > 10). As can be seen from this analysis, the use of natural

occurrence as part of the safety evaluation provides an important

perspective on the impact of intentional addition of flavouring

substances to foods.

If a flavouring substance is one of the few that for any reason,

including high intake, is of relatively high safety concern, then a

high consumption ratio likely enhances that concern because it

indicates a much larger and uncontrolled exposure from natural

occurrence than from intentional addition. If, on the other hand, a

flavouring substance is one of the vast majority that have few, if

any, safety issues and consequently low inherent concern, then a high

consumption ratio reduces any concern still further because it

indicates that intentionally added use is trivial. Stated in another

way, if the added use of a flavouring substance amounts to less than

10% of its natural occurrence, this would indicate a minimal safety

concern about added use. If added use is < 1% of natural use (i.e. CR

> 100) then the added use can at most be of trivial safety concern.

2.2 Structure-activity relationships

Toxicity is dependent on the chemical structure of a substance,

its pharmacokinetics, and its metabolic reaction pathways. Available

metabolic pathways are usually dose-dependent and, to a large extent,

govern the magnitude of the toxic effect. Therefore, chemical

structure, pharmacokinetics, metabolic fate and dose are key

determinants of toxicity and play a critical role in safety evaluation

of flavouring ingredients.

Table 1. Number of flavouring substancesa within various intake

categories

Intake categoryb Cumulative

(μg/day) No. of flavours frequency

(% of total)

< 0.01 349 26

0.01-0.1 93 33

0.1-1 274 54

1-10 224 71

10-100 204 86

100-1000 111 95

1000-10 000 45 98

10 000-100 000 16 99

100 000+ 7 100

TOTAL 1323

a Chemically defined flavouring substances permitted for use in the

U.S. excluding botanicals.

b Intake data calculated assuming: survey poundage reflects 60% of

actual usage, 10% of population exposed, U.S. population in 1987

was 240 million. Formula: Intake (μg/person/day) = [(annual

flavour usage in μg)‰0.6]‰(24×106 persons × 365 days). Poundage

data from 1987 NAS/NRC survey data.

Refinements to the initial concepts of structure-activity came as

a result of increasing knowledge and confidence in predicting

structure-activity relationships for flavouring substances. This

formed the basis of a paper by Cramer et al. (1978) which, through

the use of a "decision tree" approach, permitted the classification of

flavouring substances into "classes of concern" based on structure and

other considerations, similar in many respects to, but predating

the "Concern Level" concept outlined by the U.S. Food and Drug

Administration in its "Redbook" (FDA, 1982, 1993).
The concept of establishing concern levels also has been

investigated further by BIBRA to evaluate its applicability to food

chemicals more generally (Phillips et al., 1987). This group

evaluated and established concern levels for several food additives,

plastic monomers, as well as flavouring substances. Although they

reported that, in their opinion, the Cramer et al. (1978) decision

tree misclassified a few substances, the decision tree was likely to

be a more realistic approach for predicting toxicity than any other

reported quantitative structure-activity relationship (QSAR)

technique. Thus, there is general consensus, based on the work of

Cramer et al. (1978), the subsequent work by BIBRA (Phillips

et al., 1987), and the fact that the FDA (1982; 1993) uses

structure-activity relationships in defining Concern Levels for food

substances, that structure-activity has a solid basis in science when

applied to substances of simple and closely related structures

encountered at low exposures, a number of which are of known low

toxicity and safe metabolic disposition. This is particularly the case

for all but a very few flavouring substances. As will be discussed

later in this paper, structure-activity relationships play an

important role in the evaluation of flavouring substances.

2.3 Use of toxicity data

Traditional safety evaluation procedures are only partially

applicable to flavouring substances. Traditional approaches to the

safety assessment of food additives typically involve the evaluation

of considerable toxicological data, usually in an amount sufficient

to establish a no-observed-effect-level (NOEL), permitting the

establishment of an acceptable daily intake (ADI). Approximately half

of the flavouring substances currently in use are naturally occurring

simple acids, aldehydes, alcohols and esters. With few exceptions,

these are rapidly metabolized to innocuous end products, the safety of

which is well established or can be assumed from metabolic and

toxicity data on the substance in question or on structurally related

substances. Moreover, the acquisition of extensive toxicity data is

unnecessary for the majority of flavouring substances because

structure-activity relationships can be used as a means of assessing

substances in a homologous series, in which only a few substances have

toxicology data, to determine safety in use. This concept has been

used by JECFA in the evaluation of structurally related flavouring

substances, including the allyl esters, amyl acetate and isoamyl

butyrate, benzyl compounds, citral compounds, alpha- and ß-ionones,

and nonanal and octanal (JECFA, 1967: 1968; 1980a; 1984a,b; 1990a,b;

1991a,b; 1993a,b). In addition, as previously indicated in Table 1,

95% of flavouring substances are consumed at exposure levels less than

1 mg/person/day and in keeping with the safety evaluation procedure

outlined in this paper, only limited toxicological data are required

in such circumstances.

When exposure is extremely low and there are organoleptic

limitations on use levels, a primary consideration is whether there is

a need to establish a numerical ADI for flavouring substances. There

are several reasons why it is not appropriate or necessary to

establish ADIs in the case of the majority of flavouring substances.

ADIs are based on toxicological data and the establishment of a NOEL,

an approach that differs from the concept that a safety evaluation

can be performed in many instances on the basis of exposure and

structure-activity relationships. Moreover, the organoleptic and

gustatory properties of flavouring substances typically limit their

use and, consequently, exposure to them. In addition, because a

majority of flavouring substances occur in nature, there is a long

history of human experience in flavouring substance consumption from

traditional foods. Nearly fifty percent of flavouring substances given

full ADIs by IECFA have consumption ratios greater than 1, indicating

their predominant natural occurrence in food (Stofberg & Grundschober,

1987). The above factors have been noted by WHO (1987) as important in

the evaluation of flavouring substances. It is also evident that very

large safety margins exist for flavouring substances, as evidenced by

the fact that the margin between the NOEL and the per capita intake of

flavouring substances ranges from more than 50 to more than 1 000 000

times for the flavouring substances given full ADIs by JECFA (Table 2).

Table 2. Safety margins between NOELs and per capita daily

exposure for various flavouring substances given full ADIs

by JECFAa

Safety Margin Number of Flavours

<100 1

100 - 1000 4

1000 - 10 000 13

10 000 - 100 000 9

100 000 + 7

TOTAL 34
a JECFA, 1967; 1968; 1970; 1971; 1972; 1974a,b; 1976a.b; 1978;

1980a,b,c; 1981a,b; 1982a,b; 1983a,b; 1984a,b; 1986; 1987a,b;

1989; 1990a,b; 1991a,b; 1992; 1993a.b,c.

The factors discussed above lend support to the argument that it

is unnecessary to establish ADIs for the majority of flavouring

substances. Not establishing a numerical ADI for the great majority of

flavouring substances is also consistent with the concept that

presumed or known metabolism can be used as a basis for safety

evaluation.

3. INTEGRATED METHOD FOR THE SAFETY EVALUATION OF FLAVOURING

SUBSTANCES

In a continuing effort to improve the basis for the safety

evaluation of flavouring substances, this paper presents a procedure

which integrates information on exposure, structure-activity

relationships, metabolic fate and toxicity. It presents a safety

evaluation procedure which allows a determination of the safety of

flavouring substances under conditions of intended use. The key

elements of the safety evaluation procedure are discussed below.

3.1 Structure-activity relationships and metabolic fate

Without exception, flavouring substances are volatile organic

chemicals. The vast majority of flavourings ingredients have simple,

well characterized structures with a single functional group and low

molecular weight (< 300). More than 700 of the 1323 chemically

defined flavouring substances used in food in the U.S. are simple

aliphatic acyclic and alicyclic alcohols, aldehydes, ketones,

carboxylic acids, and related esters, lactones, ketals, and acetals.

Other structural categories include aromatic (e.g., cinnamaldehydes

and amhranilates), heteroaromatic (e.g., pyrazines and pyrroles) and

heterocyclic (e.g., furanones and thiofurans) substances with

characteristic organoleptic properties (e.g., furanones providing a

strawberry note). For most flavouring substances, the structural

differences are small. Incremental changes in carbon chain length and

the position of a functional group or hydrocarbon chain typically

describe the structural variation in groups of related flavouring

substances. These systematic changes in structure provide the basis

for understanding the effect of structure on the chemical and

biological properties of a substance.

Toxicity is dependent on the chemical structure and metabolism of

a substance. The "decision tree" procedure (Cramer et al., 1978)

relies primarily on chemical structure and estimates of total human

intake to assess toxic hazard and to establish priorities for

appropriate testing. The procedure utilizes recognized pathways of

metabolic deactivation and activation, data on toxicity, and the

presence of the substance as a component of traditional foods and as

an endogenous metabolite. Substances are classified according to three

categories:

Class I - Substances of simple chemical structure and efficient

modes of metabolism which would suggest a low order

of oral toxicity (e.g., butyl alcohol or isoamyl

butyrate).

Class II - Contains structures that are intermediate. They

possess structures that are less innocuous than

substances in Class I, but do not contain structural

features suggestive of toxicity like those substances

in Class III. Members of Class II may contain

reactive functional groups (e.g., furfuryl alcohol,

methyl 2-octynoate, and allyl propionate).

Class III - Substances of a chemical structure that permit no

strong initial presumption of safety, or may even

suggest significant toxicity (e.g., 2-phenyl3-carbethoxy furan and benzoin).

The decision tree is a tool for classifying flavour ingredients

according to levels of concern. The majority of flavouring substances

fall into Class I because they are simple alcohols, aldehydes,

ketones, acids or their corresponding esters, acetals and ketals that

occur naturally in food and, in many cases, are endogenous substances.

They are rapidly metabolized to innocuous products (e.g., carbon

dioxide, hippuric acid, and acetic acid) by well recognized reactions

catalyzed by cellular enzymes that exhibit high specificity and high

catalytic efficiency (e.g., alcohol dehydrogenase and isovaleryl

coenzyme A dehydrogenase). Substances that do not undergo detoxication

via these highly efficient pathways (e.g., fatty acid pathway and

citric acid cycle) are metabolized by reactions catalyzed by enzymes

of low specificity and relatively low efficiency (e.g., cytochrome

P-450 and glutathione transferase). For some groups of substances

(e.g., branched-chain carboxylic acids, allyl esters, and linear

aliphatic acyclic ketones), metabolic thresholds for intoxication have

been identified (Krasavage et al., 1980; Deisinger et al., 1994;

Jaeschke et al., 1987). The dose range, over which a well-defined

change in metabolic pathway occurs, generally correlates with the dose
range over which a transition occurs from a no-observed-adverse-effect

level to an adverse-effect level. For such groups of substances the

dose range at which this transition occurs is orders of magnitude

greater than the level of exposure from use as flavour ingredients.

Most substances in Class II belong to either of two categories;

one includes substances with functional groups which are similar to,

but somewhat more reactive than functional groups in Class I

(e.g., allyl and alkyne); the other includes substances with more

complex structures than substances in Class I, but that are common

components of food. This category includes heterocyclic substances

(e.g., 4-methylthiazole) and terpene ketones (e.g., carvone).

The majority of the flavouring substances within Class III

include heterocyclic and heteroaromatic substances and cyclic ethers.

Many of the heterocyclic and heteroaromatic substances have sidechains

with reactive functional groups. In a few cases, metabolism may

destroy the heteroaromaticity of the ring system (e.g., furan).

Although metabolism studies have been performed for Class III

flavouring substances with elevated levels of exposure, the metabolic

fate of many substances in this structural class cannot be confidently

predicted. Review of the group of substances in each of the structural

classes indicates that as structural complexity increases (Class I -

III), the number of flavouring substances and the levels of exposure

decrease significantly (Table 3). In all structural classes,

one-quarter or more of the flavouring substances are consumed at

levels below 0.01 μg/day or 0.2 μg/kg bw/day.

Table 3. Number of flavouring substancesa divided by structural class

within various intake categories

Intake Categoryb No. of Flavours (% of Total)

(μg/day)

Class I Class II Class III

<0.01 212 (24) 68 (28) 69 (34)

0.01-0.1 55 (6) 20 (8) 18 (9)

0.1-1 169 (19) 48 (20) 57 (28)

1-10 145 (17) 45 (19) 34 (17)

10-100 147 (17) 39 (16) 18 (9)

100-1000 95 (11) 12 (5) 4 (2)

1000-10 000 34 (4) 9 (4) 2 (1)

10 000-100 000 16 (2) 0 0

100 000+ 5 (0.6) 2 (0.8) 0

TOTAL 878 243 202

a Chemically defined flavouring substances permitted for use in the

U.S. excluding botanicals.

b Intake data calculated assuming: survey poundage reflects 60% of

actual usage, 10% of population exposed, U.S. population in 1987

was 240 million. Formula: Intake (μg/person/day) = [(annual

flavour usage in μg)‰0.6]‰(24×106persons × 365 days). Poundage

data from 1987 NAS/NRC survey data.

3.2 Integrating information on exposure and toxicity

One of the key elements of the safety evaluation procedure is

based on the premise that intake levels can be specified for

flavouring substances that would not present a safety concern. This

paper presents a procedure which provides the scientific underpinnings

for defining toxicologically inconsequential exposures for flavouring

substances and expressing these as human exposure thresholds. The

concept of specifying human exposure thresholds relies on principles

that permit specifying the daily intake of a substance which can be

considered, for practical purposes, as presenting no toxicological

risks (and thus of no health or safety risk to consumers) even in the

absence of specific toxicological data on the substance (Federal

Register, 1993; Munro, 1990; Rulis, 1986; Frawley, 1967). The

concept relies on knowledge of the range of toxicological risks for

structurally related substances and on knowledge regarding the

toxicological potency of relevant classes of chemicals for which good

toxicity data exist. The principles underpinning the establishment of

human exposure thresholds have been embodied in a recent United States

government Federal Register (1993)1 notice emanating from the FDA,

which provides the scientific basis for the conclusion that an

exposure level to indirect food additives can be specified, below

which no risk to public health would likely accrue. This exposure

level has, in turn, been used by FDA to establish a proposed

"threshold of regulation" for indirect food additives which precludes

the need for toxicological evaluation of substances migrating into

food from food-contact articles provided the amount that migrates does

not lead to a dietary level in excess of 500 ppt (equivalent to

1.5 μg/person/day assuming a daily food intake of 3000 g). The FDA has

noted that such a level would result in negligible risk to consumers

even if the substance was shown later to be a carcinogen. This concept
is in keeping with the well-established principle that resources

should be directed to the safety evaluation of substances having high

exposure and therefore greater potential for adverse effects and not

toward substances with trivial exposure. The concept is particularly

applicable to substances of low toxicity and with known or predictable

metabolic fate. The scientific basis for the establishment of human

exposure thresholds and the proposed FDA regulation are discussed

below.

Frawley (1967) initially suggested the concept of establishing

human exposure thresholds. He showed, on the basis of studies

conducted on several well-tested substances, including food additives,

industrial and consumer chemicals, and pesticides that a generic

"no-effect" level could be established that could preclude the need

for toxicity studies and safety evaluation for a majority of

substances intended for use as food packaging materials. Frawley

constructed a reference database of non-tumorigenic endpoints using

220 two-year rodent studies. He presented the NOELs for all 220

compounds. Frawley (1967) reported that if he excluded heavy metals

and pesticides from the analysis, there was no compound in the

remaining database (except for acrylamide) which showed evidence of

chronic toxicity at dietary concentrations of less than 100 ppm.

Application of a typical 100-fold safety factor to the 100 ppm

generalized NOEL would mean that humans could safely consume any of

the materials provided the dietary concentration did not exceed 1 ppm.

Frawley (1967), noting that his database was incomplete, proposed

adding an additional safety factor of 10 which would translate to a

toxicologically insignificant human exposure level of 0.1 ppm in the

diet. Assuming an individual consumes 1500 grams of food per day, an

exposure of 150 μg/person/day (approximately 2.5 μg/kg/day) or less to

a chemical of unknown toxicity would be considered toxicologically

insignificant. According to Frawley, such exposures could be

considered of no safety concern.

More recently, Rulis (1986) conducted a similar analysis of the

FDA's Priority-Based Assessment of Food Additives (PAFA) database

containing 159 compounds with subchronic or chronic toxicity data. He

came to the same conclusion as Frawley (1967). Essentially there is no

risk of toxicity in rodents exposed to certain food additives at

dietary levels of less than 1 mg/kg bw/day or in human terms,

approximately 1 to 10 μg/kg bw/day depending on the safety factor

applied. Even twenty years apart, using different databases, the

toxicologically inconsequential levels proposed by Frawley (1967) and

Rulis (1986) were nearly identical.

Munro (1990) used a database of approximately 350 substances

compiled by Gold et al. (1984, 1989) to develop a human exposure

threshold value to be applied to substances for which no presumption

of safety can be made because of a complete lack of data on metabolism

and potential toxicity. Munro (1990) proposed a threshold of

regulation of up to 1000 ppt for indirect additives which would

translate to a daily intake of 1.5 to 3.0 μg/person/day depending upon

assumptions regarding food intake. The acceptable exposure of

1.5 μg/day is equivalent to that considered by FDA (Federal Register,

1993)1 to present no regulatory concern for a food packaging

material even if later it was determined to be a carcinogen.

1 FDA has proposed a dietary concentration of 500 ppt as the

threshold of regulation for substances used in food-contact

articles. Assuming that an individual consumes 1500 g of solid

food and 1500 g of liquid food per day, this threshold would

equate to a toxicologically inconsequential level of 1.5 μg/day

(Federal Register, 1993).

The work conducted by the FDA (Federal Register, 1993), Frawley

(1967), Rulis (1986) and Munro (1990) is expanded upon in this paper

through the compilation of a large database of reference substances

(Appendix B) from which a distribution of NOELs could be derived for

chemicals of various structural types. The reference database

describes the relationships between exposure, structure and toxicity

for a wide variety of chemicals of divergent structure and it can be

used as a reference point from which to judge the safety of flavouring

substances.

In compiling the database, strict criteria were applied in the

selection of data sets. The objective of the exercise was to identify

as many high quality toxicological studies as possible representing a

variety of toxic endpoints and chemical structures. To accomplish

this, the study types included those typically conducted in

toxicology, such as subchronic, chronic, reproductive and teratology

studies. Short term and acute studies were not included since these

were considered not to be relevant for establishing chronic NOELs. The

database consisted mainly of studies in rodents and rabbits. Very few

studies in dogs and other species were found that met the established

criteria. An evaluation of randomly selected dog and primate studies

indicated that many had too few animals per group to derive a

statistically valid NOEL. Moreover, for many dog studies, a common

endpoint was reduced body weight and/or food consumption which was
due, in many cases, either to palatability problems with the diet or

vomiting. In addition, most studies in dogs and other non-rodent

species were simply too short in duration to be classified as chronic

studies. Only oral studies were included in the database. A further

criterion for inclusion in the database was that a study had to have a

demonstrated lowest-observed-effect level (LOEL) as well as a NOEL,

thus ensuring that the study was rigorous enough to detect toxic

effects. In some instances NOELs were included for studies not

demonstrating a LOEL, and these were substances such as major food

ingredients that were without toxicity at the highest dose tested in

well-conducted studies. It should be noted that the inclusion of such

substances in the database would not bias the database in favour of

higher NOELs since the true NOEL for such substances probably would

exceed the NOEL established from the available studies.

In order to group NOELs for substances with only subchronic

studies with those with chronic studies to derive the cumulative

distribution of NOELs, subchronic NOELs were divided by a factor of

three to approximate the most likely NOEL that would be derived from a

chronic study. This conversion factor is based on research defining

the relationship between subchronic and chronic NOELs. Weil and

McCollister (1963) compared 3 month NOELs with 2 year NOELS for 33

different substances (including pharmaceuticals, pesticides, and food

additives) fed to rats. They found that for most of the compounds

(30), the ratio of the NOELs between subchronic and chronic studies

was 5 or less and more than half of the compounds had a ratio equal to

2 or less. More recently, it has been discovered through further

analysis of more chemical substances, that a more accurate adjustment

factor for extrapolating NOELs derived from subchronic studies to

lifetime was between 2 and 3 (Dourson, personal communication;

Lewis et al., 1990; Beck et al., 1993).

In compiling the database, emphasis was placed on retrieving data

from certain databases known to contain well-validated toxicological

endpoints for a series of well-defined chemical structures. An

exhaustive search was made of compounds evaluated by JECFA. Other

sources included the U.S. Environmental Protection Agency's (EPA)

Integrated Risk Information System (IRIS) on-line database, the

National Toxicology Program (NTP) studies, the Developmental and

Reproductive Toxicity (DART) on-line database from EPA and the U.S.

National Institute of Environmental Health Sciences and the published

literature in general. The data entered into the database included the

name of the chemical, Chemical Abstracts Service Registry Numbers (CAS

No.), structural classification as assessed using the Cramer et al.

(1978) decision tree and the FDA "Redbook", species, sex, route of

administration, dose levels tested, study type, duration, endpoints

reported, LOEL, NOEL and references. In an effort to be conservative

in the construction of the reference database, NOELs selected by the

author(s) of each study were used even though in some cases authors

tended to over-interpret their data. In some instances, it was found

that the stated NOEL may have been based on a misjudgment of an

adverse effect by the author (e.g., physiological versus toxicological

effects) or on artifactual effects (e.g., fetal toxicity as a result

of maternal toxicity). An example of this is isopropyl alcohol, which

has been reported to produce teratogenic effects at very low doses

(0.018 mg/kg) in one study; however, its structure, known metabolism

and other toxicological data provide no evidence for concluding

teratogenicity. Even though scientifically, some of these

author-derived NOELs were not thoroughly substantiated, they were

included in the reference database, thereby increasing the degree of

its conservative nature. NOELs selected by EPA for the IRIS database

were entered without further review. In all, the database consists of

612 substances representing a range of industrial chemicals,

pharmaceuticals, food substances and environmental and consumer

chemicals likely to be encountered in commerce. Since the database was

developed as a reference database for the evaluation of flavouring

substances, all of which are organic chemicals, no organometallic or

inorganic compounds were included in the database. For many of the

substances, more than one NOEL was identified from the literature

resulting from the fact that some substances were tested in more than

one species and sex and/or demonstrated a range of endpoints suitable

for establishing a NOEL. This led, in some cases, to multiple NOELs

for individual substances. In all, the database contains 2944 entries.

In order to correlate chemical structure with toxicity, the

substances in the database were classified into three groups

corresponding to the three structural classes outlined in the

Cramer et al. (1978) decision tree. For each substance, the most

conservative NOEL was selected from the reference database based on

the most sensitive species, sex and endpoint. The cumulative

distribution of the NOELs within each class is shown in Figure 1,

along with the lognormal distributions fitted to these data. These

results clearly delineate the effects of structural class on toxicity,

with the median (50th percentile) NOEL decreasing from Class I through

III. Similar differences among structural classes exist in the range

between the 5th and 95th percentile.

The human exposure threshold for each of the structural classes

was calculated from the 5th percentile NOEL. The 5th percentile NOEL

was chosen because this value would provide 95% confidence that any
other substance of unknown toxicity but of the same structural class

as those comprising the reference database would not have a NOEL less

than the 5th percentile for that particular structural class within

the reference database.

The 5th percentile NOELs for each structural class are shown in

Table 4. In converting the 5th percentile NOELs to human exposure

thresholds (Table 4) for the various structural classes, a 100-fold

safety factor was used since such a factor would inherently be applied

in establishing safe intake levels for the substances comprising the

database. The use of such a factor provides a substantive margin of

safety since the human exposure thresholds are based on a large

database of approximately 612 compounds with good supporting

toxicological data. Furthermore, 5th percentile NOELs were used to

calculate the thresholds, providing a more conservative figure than

the arithmetic mean. Moreover, the estimated daily intakes of

flavouring substances to which the human exposure threshold are

compared are greatly overestimated as they represent the "eaters only"

(10%) population. Thus, it is believed that a 100-fold safety factor

provides a wide margin of safety in relating the results of the

analysis of the reference database to flavouring substance exposure.

It is evident from Table 4 that there are substantial differences

in the 5th percentile NOELs for the various structural classes,

indicating an obvious effect of structure on toxic potency.

It is enlightening to compare the human exposure thresholds with

present intakes of chemically defined flavouring substances in the

U.S. As shown in Table 5, it is clear that for nearly all (93 to

97%) flavouring substances used in the U.S., intakes are below the

human exposure threshold for their respective structural class.

Because most flavouring substances possess simple structures and their

metabolism is known or reasonably predictable, it can be concluded

that it is highly improbable that they would present a toxicological

risk at exposure levels below the human exposure threshold for their

respective structural class. However, even if information on

structural class, metabolic fate and existing toxicity studies were

not available, 743/1323 (56%) of flavouring substances used in the

U.S. are consumed in amounts less than the 1.5 μg/person/day standard

proposed by the FDA (Federal Register, 1993) and Munro (1990). This

indicates that for approximately half of the existing list of 1323

chemically defined flavouring substances permitted for use in the

U.S., exposure can be considered to be trivial.

Table 4. Fifth percentile NOELs and human exposure thresholds for

Cramer et al. (1978) structural classes in the reference

database

5th Percentile NOELs, Human Exposure

(μg/kg bw/day) Threshold,(μg/day)a,b

I 137 2993 1800

II 28 906 540

III 447 147 88

a The human exposure threshold was calculated by multiplying the

5th percentile NOEL by 60 (assuming an individual weighs 60 kg)

and dividing by a safety factor of 100, as discussed in the text.

b Numbers rounded to two (2) significant figures.
Table 5. Flavouring substances within each Cramer et al. (1978)

structural class consumed in amounts below human exposure

thresholds

No. of No. of Flavours

Human Exposure Flavours Under Human

Structural Class Threshold Within Exposure

(μg/day) Structural Threshold (%)

Classa

I 1,800 878 835 (95)

II 540 243 227 (93)

III 88 202 195 (97)

a Adapted from the FEMA flavouring substance database of flavouring

substances permitted for use in the U.S.

3.3 Evaluation criteria

The evaluation criteria proposed for application to flavouring

substances in this paper involve the integration of information on

exposure, structure-activity relationships, metabolism and, as

required, toxicity data. It should be noted that the safety evaluation

criteria outlined below are not intended to be applied to any

flavouring substances with unresolved toxicity problems or to

substances that are presumed or known carcinogens. Such substances

warrant special consideration and must be evaluated using more

traditional methods of safety evaluation. While toxicity data exist on

numerous flavouring substances and can be used as the basis for

evaluation, there are many flavouring substances that lack toxicity

data. The evaluation criteria are intended to provide a means of

evaluating such substances. The criteria incorporate, where the data

permit, the concept that metabolic fate can be predicted on the basis

of presumed structure-activity relationships. The criteria also rely,

in part, on the NOEL reference database which provides a human

exposure threshold for each of the three structural classes of

flavouring substances, as shown in Table 4. The evaluation criteria

also incorporate, where available, toxicity data on flavouring

substances and closely related structural analogues as a basis for

safety evaluation. One of the criteria (number 5 below) incorporates

the concept of a minimum human exposure threshold based on the

1.5 μg/person/ day standard proposed by the FDA (Federal Register,

1993) and Munro (1990). This standard can be applied to those

flavouring substances for which metabolic fate is unknown and cannot

be confidently predicted and for which there are no toxicity data on

the flavouring substance or on a structurally related material from

which to conclude any inference of safety in use.

Flavouring substances that meet one of the five numbered criteria

outlined below can be declared safe for their intended use without

further evaluation:

1. a) The flavouring substance has a simple structure and will be

metabolized and excreted through known detoxication pathways

to innocuous end-products; and

b) the conditions of intended use do not result in an exposure

greater than the human exposure threshold for the relevant

structural class, indicating a low probability of potential

for adverse effects.

2. a) The conditions of intended use result in an exposure that

exceeds the human exposure threshold for the relevant

structural class; however

b) the flavouring substance has a simple structure and will be

metabolized and excreted through known detoxication pathways

to innocuous end-products and it or its metabolites are

endogenous human metabolites with no known biochemical

regulating function.

3. a) The flavouring substance has a simple structure and will be

metabolized and excreted through known detoxication pathways

to innocuous end-products; and

b) the conditions of intended use result in an exposure that

exceeds the human exposure threshold for the relevant

structural class; however

c) toxicity data exist on the flavouring substance which

provide assurance of safety under conditions of intended

use, or there are toxicity data on one or more close

structural relatives which provide a NOEL high enough to

accommodate any perceived difference2 in toxicity between

the flavouring substance and the structurally related

substance having toxicity data.
4. a) The metabolic fate of the flavouring substance cannot be

confidently predicted on the basis of structure; however

b) the conditions of intended use result in an exposure below

the human exposure threshold for the relevant structural

class indicating a low probability of potential for adverse

effects; and

c) there are toxicity data on the substance, or on one or more

structurally related substances with a NOEL at least 10

times greater than the 5th percentile NOEL for the relevant

structural class.

5. a) The metabolic fate of the flavouring substance cannot be

confidently predicted on the basis of structure; however

b) the conditions of intended use result in an exposure below

the human exposure threshold of 1.5 μg/day2, providing

assurance that the substance will be safe under conditions

of intended use.

[NOTE: This criterion accommodates flavouring substances for

which there are limited data on structure-activity and toxicity,

but they would not be considered to present a safety concern

below the human exposure threshold of 1.5 μg/day (Federal

Register, 1993)].

The evaluation criteria listed above identify substances which

are considered safe under conditions of intended use or those that may

require additional data and further evaluation. Figure 2 presents the

same criteria in the form of an evaluation sequence. The sequence

contains a number of questions on structure, metabolism, exposure data

and toxicity and provides an integrated mechanism to evaluate the

safety of a flavour ingredient. Although the procedure incorporates

relevant toxicity data on a substance or related substances where

available, it does not require them. The procedure identifies those

substances for which additional safety data may be required in order

to perform an adequate safety evaluation.

The effective application of this safety evaluation procedure

depends on a substantial knowledge of toxicology, chemistry,

metabolism, and exposure to flavouring substances. It can be applied

most effectively when groups of structurally related flavouring

substances are evaluated together. In a group evaluation, conclusions

reached on the safety of individual substances are supported by

similar conclusions for structurally related substances. For example,

the results of the evaluation for butyl butyrate should be consistent

with results for other esters formed from aliphatic acyclic linear

saturated alcohols and acids having similar levels of exposure. These

similar substances will pass through the same branch of the safety

evaluation procedure because they fall into the same structural class,

possess similar metabolic fate, and exhibit similar patterns of

exposure from use as flavour ingredients and as components of food.

2 In most instances, groups of structurally related materials have

toxicology data on at least one member of the group, usually the

flavoring substance with the highest poundage. In most cases a

large margin of safety ( i.e., 100- to 1000-fold) exists

between the NOEL and the calculated exposure to the substance

having toxicological data. Such margins of safety would be

expected to accommodate any perceived difference between the

toxicity of a flavouring substance having no toxicological data

and its close structural relative for which a NOEL has been

established.

In the first step of the safety evaluation procedure (Figure 2)

the user must assign a decision tree structure class (Cramer et al.,

1978) to the substance. Following assignment of structure class, a

question on metabolic fate appears. This question identifies those

substances which are anticipated to be efficiently metabolized

to innocuous products (e.g., 1-butanol) versus those which are

transformed to more toxic metabolites (e.g., estragole) or have

limited information on which to predict confidently the metabolic fate

(e.g., 2-phenyl-3-carbethoxy furan).

Once a substance has been sorted according to structure class

and knowledge of metabolic fate, the next question compares the

substance's daily per capita intake ("eaters only") from use as a

flavour ingredient to the human exposure threshold (Table 4) for the

same structure class.

If the substance is metabolized to innocuous products (Step

No. 2) and has an intake less than the human exposure threshold for

the structure class (Step No. 3), the substance is considered safe

(e.g., 1-octanol). If the intake is greater than the human exposure

threshold (Step No. 3) and the substance or its metabolites are

endogenous (Step No. 4), the substance is also considered safe, even

though the intake is greater than the human exposure threshold (e.g.,
butyric acid). If the substance is not endogenous, then the substance

or related substances must have a NOEL (Step No. 5) significantly

greater than the intake of the substance in order to be considered

safe (e.g., citral). If no such data exist or the NOEL is not

significantly greater than the intake for the substance, then

additional data are required in order to complete the safety

evaluation.

If metabolic fate cannot be confidently predicted and the intake

(Step No. 2) is greater than the human exposure threshold (Step

No. 3), additional data on metabolic fate or toxicity on the substance

or structurally related substances are required to complete the safety

evaluation (e.g., dihydro-coumarin). If the intake is less than the

human exposure threshold, the substance or structurally related

substances must have a NOEL significantly greater than the NOEL for

the structure class (Step No. 6) in order for the substance to be

considered safe (e.g., 2-ethyl-4-hydroxy-3(2H)-furanone). If an

adequate toxicity study is not available and the substance has an

intake less than 1.5 μg/day (Step No. 7), the substance is considered

safe (e.g., 3-acetyl-2,5-dimethylthiophene). Otherwise additional data

are required in order to complete the safety evaluation (e.g.,

2-ethylfuran).

The principal objective of the safety evaluation procedure is to

identify two groups of flavourings substances: i) those substances

whose structure, metabolism, and relevant toxicity data clearly

indicate that the substance would be expected not to be a safety

concern under current conditions of intended use; and ii) those

substances which may require additional data in order to perform an

adequate safety evaluation.

3.4 Integrating data on the consumption ratio

As pointed out by WHO (1987) and SCF (1991), natural occurrence

is no guarantee of safety, but it is important to recognize that the

safety evaluation of added use of flavouring substances needs to be

conducted with an appreciation of the consumption ratio. Clearly, if

added use of flavouring substances results in an exposure that exceeds

that from natural sources, this will increase awareness of the need to

consider carefully overall exposure in the light of existing data on

toxicity and structure-activity relationships. On the other hand, if

the added use is trivial with a consumption ratio of 10 to 100, that

is, it increases total exposure by only 1 to 10%, then this fact needs

to be taken into consideration when applying the criteria outlined

above.

The substances of primary concern are those which, in Figure 2,

receive a "No" answer to Step No. 5, or a "Yes" answer to Step No. 7,

indicating a possible need for additional data and evaluation beyond

that included in the evaluation procedure outlined in this paper. In

such an evaluation, as stated immediately above, the extent of natural

occurrence should be given appropriate weight.

Of much less concern with respect to consumption ratio are those

substances that drop out of further consideration as a result of "No"

answers to Step Nos. 3 or 7, or "Yes" answers to Step Nos. 4 or 6. The

derivation of the thresholds, the estimation of intakes (see Appendix

A), and the special factors applicable to the use of flavours

(volatility, self-limiting use, etc.) build in multiple conservative

assumptions more than adequate to cover additional exposure from

natural occurrence to substances that in any case are of low inherent

concern.

The advances in analytical chemistry in the past 50 years provide

virtual assurance that no flavouring substances of extensive natural

occurrence remain unknown. Those of potential value yet to be
discovered (e.g., as yet unknown components of roast beef, coffee, or

chocolate flavour) are being sought at the ppb and ppt level. This

does not suggest exposures above any of the thresholds discussed in

this paper.

4. CONCLUSIONS

It is neither possible nor necessary to conduct toxicological

studies on all individual flavouring substances used in food. The vast

majority of flavouring substances are members of groups of substances

with common metabolic pathways, and typically, individual members of

such a group display a similar toxicity profile. This is not

surprising in light of the close structural similarity of the various

flavouring substances comprising a chemical group. As demonstrated in

this paper, exposure to flavouring substances is usually low and, in

the majority of cases, below the human exposure threshold values

presented in this paper. This knowledge, coupled with knowledge

regarding structure-activity relationships, metabolic disposition, and

toxicology data, permits the establishment of the proposed paradigm

for safety assessment which provides a solid foundation to assure the

safety of flavouring substances under conditions of intended use.

5. REFERENCES

BECK, B.D., CONOLLY, R.B., DOURSON, M.L., GUTH, D., HATTIS, D.,

KIMMEL, C. & LEWIS, C. (1993). Symposium overview. Improvements in

quantitative noncancer risk assessment. Fund Appl Toxicol.,

20: 1-14.

COUNCIL OF EUROPE (1974). Natural Flavouring Substances, Their

Sources, And Added Artificial Flavouring Substances. Council of

Europe, Strasbourg, France.

COUNCIL OF EUROPE ( 1981 ). Flavouring Substances And Natural Sources

Of Flavourings, Third Edition. Council of Europe, Strasbourg, France.

COUNCIL OF EUROPE (1992). Flavouring Substances and Natural Sources

of Flavourings. Volume I. Chemically-Defined Flavouring Substances,

Fourth Edition. Council of Europe, Maissoneuve, France.

CRAMER, G.M., FORD, R.A. & HALL, R.L. (1978). Estimation of toxic

hazard-A decision tree approach. Food Cosmet Toxicol., 16: 255-276.

DEISINGER, P.J., BOATMAN, R.J. & GUEST, D. (1994). Metabolism of

2-ethylhexanol administered orally and dermally to the female Fischer

344 rat. Xenobiotica, 24: 429-440.

FOOD AND DRUG ADMINISTRATION (1982). Toxicological Principles for the

Safety Assessment of Direct Food Additives and Color Additives Used in

Food. Redbook. U.S. Food and Drug Administration, Bureau of Foods,

Washington, DC.

FOOD AND DRUG ADMINISTRATION (1993). Toxicological Principles for the

Safety Assessment of Direct Food Additives and Color Additives Used in

Food. Redbook II (Draft). U.S. Food and Drug Administration, Center

for Food Safety and Applied Nutrition, Washington, DC.

FEDERAL REGISTER (1993). Food additives; threshold of regulation for

substances used in food-contact articles. 58(195): 52719-52729.

FRAWLEY, J.P. (1967). Scientific evidence and common sense as a basis

for food-packaging regulations. Food Cosmet Toxicol., 5: 293-308.

GOLD, L.S., SAWYER, C.B., MAGAW, R., BACKMAN, G.M., DE VECIANA, M.,

LEVINSON, R., HOOPER, N.K., HAVENDER, W.R., BERNSTEIN, L., PETO, R.,

PIKE, M. & AMES, B.M. (1984). A carcinogenic potency database of the

standardized results of animal bioassays. Environ Health Perspect.,

58: 9-319.

GOLD, L.S., SLONE, T.H. & BERNSTEIN, L. (1989). Summary of

carcinogenic potency and positivity for 492 rodent carcinogens in the

carcinogenic potency database. Environ. Health Perspect., 79: 259-272.

JAESCHKE, H., KLEINWAECHTER, C. & WENDEL, A. (1987). The role of

acrolein in allyl alcohol-induced lipid peroxidation and liver cell

damage in mice. Biochem. Pharmacol., 36: 51-57.

JECFA (1967). Toxicological Evaluation of Some Flavouring Substances

and Non-Nutritive Sweetening Agents. Eleventh Meeting of the Joint

FAO/WHO Expert Committee on Food Additives.

JECFA (1968). Specifications for the Identity and Purity of Food

Additives and their Toxicological Evaluation: Some Flavouring

Substances and Non-nutritive Sweetening Agents. Eleventh Report of the

Joint FAO/WHO Expert Committee on Food Additives, FAO Nutrition

Meetings Report Series No. 44, WHO Technical Report Series No. 383.

JECFA (1970). Toxicological Evaluation of Some Extraction Solvents and

Certain Other Substances. Fourteenth Meeting of the Joint FAO/WHO

Expert Committee on Food Additives.
JECFA (1971). Evaluation of Food Additives. Fourteenth Report of the

Joint FAO/WHO Expert Committee on Food Additives, WHO Technical Report

Series No. 462.

JECFA (1972). Evaluation of Food Additives. Some Enzymes, Modified

Starches, and Certain Other Substances: Toxicological Evaluations and

Specifications and a Review of the Technological Efficacy of Some

Antioxidants. Fifteenth Report of the Joint FAO/WHO Expert Committee

on Food Additives, WHO Technical Report Series No. 488.

JECFA (1974a). Toxicological Evaluation of Certain Food Additives

Including Anticaking Agents, Antimicrobials, Antioxidants, Emulsifiers

and Thickening Agents. Seventeenth Meeting of Joint FAO/WHO Expert

Committee on Food Additives, WHO Food Additive Series No. 5.

JECFA (1974b). Toxicological Evaluation of Certain Food Additives with

a Review of General Principles and of Specifications. Seventeenth

Report of the Joint FAO/WHO Expert Committee on Food Additives, WHO

Technical Report Series No. 539.

JECFA (1976a). Evaluation of Certain Food Additives. Twentieth Report

of the Joint FAO/WHO Expert Committee on Food Additives, FAO Nutrition

Meetings Report Series No. 1, WHO Technical Report Series No. 599.

JECFA (1976b). Toxicological Evaluation of Certain Food Additives.

Twentieth Meeting of the Joint FAO/WHO Expert Committee on Food

Additives, WHO Food Additive Series No. 10.

JECFA (1978). Evaluation of Certain Food Additives and Contaminants.

Twenty-second Report of the Joint FAO/WHO Expert Committee on Food

Additives, WHO Technical Report Series No. 631.

JECFA (1980a). Evaluation of Certain Food Additives. Twenty-third

Report of the Joint FAO/WHO Expert Committee on Food Additives. World

Health Organization, Geneva. WHO Technical Report Series No. 648.

JECFA (1980b). Evaluation of Certain Food Additives. Twenty-fourth

Report of the Joint FAO/WHO Expert Committee on Food Additives. World

Health Organization, Geneva. WHO Technical Report Series No. 653.

JECFA (1980c). Toxicological Evaluation of Certain Food Additives.

Twenty-third Meeting of the Joint FAO/WHO Expert Committee on Food

Additives, WHO Food Additive Series No. 14.

JECFA (1981a). Evaluation of Certain Food Additives. Twenty-fifth

Report of the Joint FAO/WHO Expert Committee on Food Additives,

Technical Report Series No. 669.

JECFA (1981b). Toxicological Evaluation of Certain Food Additives.

Twenty-fifth Meeting of the Joint FAO/WHO Expert Committee on Food

Additives, WHO Food Additive Series No. 16.

JECFA (1982a). Evaluation of Certain Food Additives and Contaminants.

Twenty-sixth Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 683.

JECFA (1982b). Toxicological Evaluation of Certain Food Additives.

Twenty-sixth Meeting of the Joint FAO/WHO Expert Committee on Food

Additives, WHO Food Additive Series No. 17.

JECFA (1983a). Evaluation of Certain Food Additives and Contaminants.

Twenty-seventh Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 696.

JECFA (1983b). Toxicological Evaluation of Certain Food Additives and

Contaminants. Twenty-seventh Meeting of the Joint FAO/WHO Expert

Committee on Food Additives, WHO Food Additive Series No. 18.

JECFA (1984a). Evaluation of Certain Food Additives and Contaminants.

Twenty-eighth Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 710

JECFA (1984b). Toxicological Evaluation of Certain Food Additives and

Contaminants. Twenty-eighth Meeting of the Joint FAO/WHO Expert

Committee on Food Additives, WHO Food Additive Series No. 19.

JECFA (1986). Evaluation of Certain Food Additives and Contaminants.

Twenty-ninth Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 733.

JECFA (1987a). Evaluation of Certain Food Additives and Contaminants.

Thirtieth Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization. Geneva. WHO Technical Report

Series No. 751.

JECFA (1987b). Evaluation of Certain Food Additives and Contaminants.
Thirty-first Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 759.

JECFA (1989). Evaluation of Certain Food Additives and Contaminants.

Thirty-third Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 776.

JECFA (1990a). Evaluation of Certain Food Additives and Contaminants.

Thirty-fifth Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 789.

JECFA (1990b). Toxicological Evaluation of Certain Food Additives and

Contaminants. Thirty-fifth Meeting of the Joint FAO/WHO Expert

Committee on Food Additives, WHO Food Additive Series No. 26.

JECFA (1991a). Evaluation of Certain Food Additives and Contaminants.

Thirty-seventh Report of the Joint FAO/WHO Expert Committee on Food

Additives. World Health Organization, Geneva. WHO Technical Report

Series No. 806.

JECFA (1991b). Toxicological Evaluation of Certain Food Additives and

Contaminants. Thirty-seventh Meeting of the Joint FAO/WHO Expert

Committee on Food Additives, WHO Food Additive Series No. 28.

JECFA (1992). Evaluation of Certain Food Additives and Naturally

Occurring Contaminants. Thirty-ninth Report of the Joint FAO/WHO

Expert Committee on Food Additives. World Health Organization, Geneva.

WHO Technical Report Series No. 828.

JECFA (1993a). Evaluation of Certain Food Additives and Contaminants.

Forty-first Report of the Joint FAO/WHO Expert Committee on Food

Additives, Technical Report Series No. 837.

JECFA (1993b). Toxicological Evaluation of Certain Food Additives and

Contaminants. Forty-first Meeting of the Joint FAO/WHO Expert

Committee on Food Additives, WHO Food Additive Series No. 32.

JECFA (1993c). Toxicological Evaluation of Certain Food Additives and

Naturally Occurring Toxicants. Thirty-Ninth Meeting of the Joint

FAO/WHO Expert Committee on Food Additives, WHO Food Additive Series

No. 30.

KRASAVAGE, W.J., O'DONOGHUE, J.L., DIVINCENZO, G.D. & TERHAAR, C.J.

(1980). The relative neurotoxicity of methyl-n-butyl ketone, n-hexane

and their metabolites. Toxicol. Appl. Pharmacol., 52: 433-441.

LEWIS, S.C., LYNCH, J.R. & NIKIFOROV, A.I. (1990). A new approach to

deriving community exposure guidelines from "no-observed-adverseeffect levels". Regul. Toxicol. Pharmacol., 11: 314-330.

MUNRO, I.C. (1990). Safety assessment procedures for indirect food

additives: An overview. Regul. Toxicol. Pharmacol., 12: 001-0011.

NAS (1978). Resurvey of the Annual Poundage of Food Chemicals

Generally Recognized as Safe, National Technical Information Service

(NTIS) PB288-081.

NAS (1979). Comprehensive Survey of Industry on the Use of Food

Additives, National Technical Information Service (NTIS) PB80-113418.

NAS (1984). Poundage Update of Food Chemicals, National Technical

Information Service (NTIS) PB84-162148.

NAS (1989). 1987 Poundage and Technical Effects Update of Substances

Added to Food. National Research Council, Washington, DC. Prepared

for: Food and Drug Administration, Washington, D.C. NTIS Report

No. PB91-127266.

PHILLIPS, J.C., PURCHASE, R., WATTS, P. & GANGOLLI, S.D. (1987). An

evaluation of the decision tree approach for assessing priorities for

safety testing of food additives. Food Addit. Contam., 4(2): 109-123.

RULIS, A.M. (1986). De Minimis and the Threshold of Regulation.

In: Felix, C.W. (Ed.) Food Protection Technology. Lewis Publishers

Inc., Chelsea, MI. pp. 29-37.

SCF (1991). Guidelines for the Evaluation of Flavourings for Use in

Foodstuffs: I. Chemically Defined Flavouring Substances. Commission of

the European Communities, Scientific Committee for Food, Brussels,

Belgium.

STOFBERG, J. & KIRSCHMAN, J. (1985). The consumption ratio of

flavouring materials: A mechanism for setting priorities for safety

evaluation. Food Chem. Toxicol., 23: 857-860.

STOFBERG, J. & GRUNDSCHOBER, F. (1987). Consumption ratio and food

predominance of flavouring materials. Perfumer Flavourist,

12: 27-68.
WEIL, C.S. & McCOLLISTER, D.D. 1963. Safety evaluation of chemicals.

Relationship between short- and long-term feeding studies in designing

an effective toxicity test. Agr. Food Chem., 11(6): 486-491.

WHO (1987). Principles For The Safety Assessment Of Food Additives

And Contaminants In Food. WHO Environmental Health Criteria. World

Health Organization (WHO), International Program on Chemical Safety in

Cooperation with the Joint FAO (Food & Agriculture Organization of the

United Nations), WHO Expert Committee on Food Additives (JECFA),

Geneva, Switzerland, Environmental Health Criteria 70.

WOODS, L.A. & DOULL, J. (1991). GRAS evaluation of flavouring

substances by the Expert Panel of FEMA. Regul. Toxicol. Pharmacol.,

14: 48-58.

ANNEX 5, Appendix A

ESTIMATING DISTRIBUTION OF INTAKES OF FOOD INGREDIENTS ACROSS

THE POPULATION

In response to the Food Additives Amendment of 1958 to the U.S.

Food, Drug and Cosmetic Act, the Flavor and Extract Manufacturers'

Association (FEMA) in 1960 conducted a comprehensive survey of all

flavoring materials then thought or claimed to be in use. Information

included: identity; poundage used annually in food; year of first use;

levels (in ppm) used in major food categories; toxicological data; and

other information related to safety. After evaluation by a panel of

external experts, this and subsequent information was published in a

series of "FEMA GRAS Lists" (Hall and Oser, 1965, 1970; Oser and Hall,

1972; Oser and Ford, 1973ab, 1974, 1975, 1977, 1978, 1979; Oser et al.,

1984, 1985; Burdock et al., 1990; Smith and Ford, 1993).

FEMA determined in 1968 that another survey was needed to monitor

trends and to improve on the previous effort. In addition, the U.S.

Food and Drug Administration (FDA) began to plan to review its

Generally Recognized as Safe (GRAS) lists, and in so doing, made use

of the prior experience of FEMA. FEMA and FDA efforts merged into the

first comprehensive survey of ingredients used in food, conducted by

the National Academy of Sciences/National Research Council (NAS) in

1970 (NAS, 1973) with several subsequent surveys (NAS, 1978, 1979,

1984, 1989).

Emerging from the 1960 and 1970 surveys was a clear indication

that per capita intake of an ingredient (from annual poundage),

although useful for comparison with other ingredients, has a

significant limitation. As an average of intakes by those who consume

foods containing the ingredient ("eaters") and by those who do not

(non-eaters"), it does not represent the intake by the individuals of

concern ( i.e., the "eaters"). This is not the case for ingredients

commonly consumed by almost everyone ( e.g., sugar), but the

difference between the average and "eaters" is significant for other

ingredients consumed only by few people ( e.g. raspberry flavor).

Because of these issues, an alternative method of calculating

intakes was used in the first (1973) NAS report. This method involved

the average level of use of an ingredient in a broad food category

(e.g., baked goods) multiplied by the average daily consumption of

that food category per person. The daily consumption for each category

was obtained by multiplying the average portion size for the category,

obtained from the U.S. Department of Agriculture (USDA) Food

Consumption Survey by the average daily frequency of consumption of

foods in that category over a 14-day survey period. The frequency of

consumption data came from a 12,000+ member panel operated by the

Market Research Corporation of America (MRCA; see note b to Table 1).

Thus the average intake of an ingredient for all relevant food

categories provided the "possible average daily intake" (PADI). These

intakes were often highly exaggerated, as much as a hundred-fold,

because an ingredient used in only a few foods in a food category was

assumed to be at the average level in all foods in the category. Thus,

for example, cardamom, used only in a few coffee cakes, was assumed to

be used in each food in the entire category of baked goods ( i.e.,

all breads, rolls, crackers, and pastries).

To address this issue, another method was developed to give the

probable distribution of intakes for the entire population without

exaggerating intake. This required an informed estimate of the

probability that each food eaten would contain the ingredient of

interest. The first series of such intake estimates used calculation

"A", as shown in Table 1, and the results are summarized in Section

"A" of Table 2.

In Table 2, Column 1 ("Substance") lists the substances selected

to test and improve the more refined method. In Section A, nutmeg and

mace were chosen, among other reasons because they are not grown

domestically in the U.S. and import data, therefore, provide a firm

external check on pounds used. Furthermore, more than half of all

nutmeg is consumed in four foods: bologna, pork sausage, frankfurters,

and doughnuts. It thus presented an opportunity to see how heavy

consumers of those foods would skew total intake distribution. The
flavor ingredient 4-(p-hydroxyphenyl)butan-2-one was chosen because it

is used in, and occurs naturally, almost entirely in only one flavor,

raspberry flavor, and is far less widely consumed than chocolate,

vanilla, or citrus flavors. Thus, it is a good example of an uncommon

ingredient consumed by those who particularly like the few foods in

which it is used (Hall, 1975).

Column 2 [" per capita (External Check)"] lists per capita

intake derived from import data, product manufacturer surveys, or

tariff reports, all independent of the NAS survey to serve as an

external check. Column 3 (" per capita [NAS(lbs)/0.6]") lists the

per capita intake from the poundage reported in the NAS survey

assumed to be 60% complete. Further research (see references to NRC

surveys) showed this to be generally a valid assumption, but with

certain qualifications. Ingredients such as sugar, corn syrup, and

salt, could not be fully or reliably reported by bakers and other

small food processors, whereas the flavor industry reported more

fully, and the 60% assumption, while used, usually resulted in a

poundage per capita overestimate.

For Section A, Column 4 ["50th Percentile (Total Panel)"]

presents the total panel 50th percentile for each substance, and

Column 4a ("50th Percentile 3 S.D. Total Panel"), the 50th percentile

plus 3 standard deviations. Column 4b ["very high (Total Panel)"]

shows the so-called "very high intake". This is not a realistic

figure, because no person could consume all foods using an ingredient

at the highest level for every food in every major category. Note

particularly, also, the earlier discussion of the exaggeration that

arises from assuming that an ingredient used only in a few foods is

used in all foods in a major food category. The "very high intake"

serves simply as a wholly unreachable and therefore unrealistic value.

Given the remaining and inevitable uncertainties due to possible

errors in estimations, and possible non-food uses, the agreement

between the total panel 50th percentile (Section A, Column 4), the NRC

per capita, (Column 3), and the per capita from external sources

(Column 2), is entirely satisfactory. However, calculation "A" did

not provide data on only those who actually consumed the substance

("eaters only"), and the statistical treatment was not as solidly

based as was desirable. This led to an improved calculation, "B", as

shown in Table 3. Table 2, Section B, presents the results of these

calculations.

The substances shown in Column 1 of Section B were selected, as

discussed in the referenced reports themselves, by the FDA or the NAS

committee (NAS, 1976). An improved method of calculation was used, as

described below, which provides the total sample mean (Column 4 ["Mean

(Total Panel)"]) as a check against the per capita data in Columns 2

and 3.

The external per capita figure for calcium propionate is twice

the total panel mean, probably due primarily to usage by many small

bakeries not included in the survey, and also to the high wastage of

bread. The slightly smaller discrepancy for MSG may well reflect

several added sources of error. Even so, comparison of the total panel

mean with the per capita data from the NAS survey and the external

checks shows satisfactory agreement. Food consumption surveys have

significant and well-recognized sources of error in addition to those

affecting the NRC surveys, and it seems unlikely that it is practical

to expect intake estimates derived by different means to correspond

more closely than within a factor of two.

For Section B, Column 4 ["Mean (Total Panel)"] provides the total

panel mean, Column 4a ["Mean (Eaters Only)"] the "eaters only" mean

age, 2 and over and Column 4b ["S.D. (Eaters Only)"], the standard

deviation for "eaters only".

The procedure in calculation B does not eliminate all sources of

error. Intake is still calculated from "disappearance figures"

( i.e, the levels introduced into food during processing, not the

lower levels actually consumed). Thus, ignoring the often large losses

due, for example, to volatilization or leaching in processing,

preparation and storage result in exaggerated estimates of intake. In

the other direction, heavy (frequent) eaters will often eat larger

portion sizes, resulting in a tendency to underestimate higher

intakes. The result overall, however, remains highly satisfactory.

Table 1 Calculation "A"

Step Operation Result

For each specific food (SF)a e.g. bologna,

within a broad food category, eg., meats,

obtain from MRCAb the

1 total number of eatings of each SF by the total
panel, in two weeks

14 days × 12,337 (persons in panel) = mean frequency of eating, p/d

×

2 mean portion size in gc = mean consumption of SF, g/p/d

×

3 probability of SF containing ingredient "I"d = probable mean consumption of SF

containing I g/p/d

×

4 usual use level of "I" in ppm of major food = 50th percentile of probable intake of I

category containing SF ‰ 1,000e from SF in mg/p/d

×

5 90th percentile MRCA frequency of eating = 90th percentile of probable intake of I

50th percentile MRCA frequency of eating from SF in mg/p/d

6 & 7 sum 4 and 5 separately across all SFs in that = 50th and 90th percentiles of probable

major food category intake of I from that major food category

Table 1 (cont'd) Calculation "A"

Step Operation Result

8 fit a curve, assuming normal distribution, to = 50th percentile and standard deviation

the points determined in 6 and 7 and calculate (SD) for each major food category

9 repeat steps 1 through 8 and sum across all = 50th percentile and SD in mg/p/d

food categories probable total intake

10 repeat steps 4 and following, using "average = "very high intake"

maximum use level" from the survey for each

major food category

a Specific food (SF), in this context, denotes a narrow category of very similar foods serving

the same dietary and market niches (e.g. all bologna, all cola drinks). The eatings of each SF

include in one total for each SF all eatings of similar brands or home-prepared versions of the

same SF. There were approximately 10,000 such SFs.

b The Market Research Corporation of America (MRCA) is a private market research company that

surveys usage of food products and components It uses a nationally representative panel and

records for each panel member the number of servings of each specific loud for each day of a

two-week period. The panelists' reporting periods are distributed evenly throughout the year. The

1973 NAS report used data from the third Menu Census (1967-1968) involving 12,871 individuals.

The dataused in calculations "A" and "B" came from the Fourth Menu Census (1972-1973) involving

12,337 individuals.

c The USDA Food Consumption Survey (USDA, 1965)

d Obtained from a consensus panel of experts knowledgeable about the ingredient and the food.

e From NAS, 1973

Table 2 Distribution of MRCA 14-Day Average Daily Intakes (in mg/person/day)a of Selected Food Ingredients

(1) (2) (3) (4) (4a) (4b) (5)

"High Eaters

Section Substance per capita per capita Mean Mean plus "Very High" Only"

(External [NAS (lbs.)/0.6] (Total Panel) 3 S.D.b (Total Panel) [NAS

Check) Total Panel (lbs.)/0.6]×10

A Nutmeg 23.6 10.3 15.7 51.3 71.1 103

Nutmeg Oil 1.2 1.0 2.0 5.7 7.1 10

A Mace 3.3 2.8 3.3 8.6 14.4 28

Mace, Oil NAc 0.26 0.19 0.67 0.46 2.6

Mace, Oleoresin NA 0.070 0.13 0.39 0.41 0.70

A 4-p-Hydroxyphenyl 0.048 0.086 0.041 0.16 0.37 0.86

Butan-2-one

Table 2 (cont'd).

(1) (2) (3) (4) (4a) (4b) (5)

"High Eaters
Section Substance per capita per capita Mean Mean plus "Very High" Only"

(External [NAS (lbs.)/0.6] (Total Panel) 3 S.D.b (Total Panel) [NAS

Check) Total Panel (lbs.)/0.6]×10

Mean Mean S.D.

(Total Panel) (Eaters Only) (Eaters Only)

B Calcium Propionate 92 33 44.8 45.1 20.7 330

Sodium Propionate NA 16 33,2 33.3 14.7 160

B Carob Bean Gum NA 26 35.6 36.1 35.0 260

Chondrus Extract 19.3 13 10.6 11.1 18.5 130

B Mono- and diglycerides NA 727 884 885 467 7,270

Monosodium Glutamate 186 194 98.8 99.0 88.5 1,940

(MSG)

a See text for explanation of columns.

b S.D. = Standard Deviation.

Table 3 Calculation "B"

1 From the MRCA survey, obtain the total

number of eatings of each SF, by each panel

member, each day over the 14 - day period

×

2 USDA mean portion size, for that person's = quantity of SF in g for that

age group and the relevant major food person, that day

category

×

3 weighted mean of usual use level of I in = quantity of I in mg/d, that

ppm ‰ 1,000 person, that day, if all of

the SF contained I

×

4 probability that particular SF contains I = expected intake of I for

that person for that day,

mg/d

5 Repeat steps 1 through 4 for that person for = expected intake of I for

each of the 14 days that person for 14 days,

mg

6 ‰ 14 = average daily intake over

14 days for that person,

g/p/d

7 tabulate all intakes and compute = total panel mean intake of I

mg/p/d

8 discard all persons with zero intake for the = "eaters only" mean and SD

14 day period, tabulate all "eaters only" of for I, mg/p/d

I, age 2 and over, and compute

Unfortunately, this more refined approach, while successful, is

extremely elaborate, tedious, and expensive. The MRCA panel is

expensive to maintain and operate, and the data are correspondingly

costly. The approach requires extensive input from a broadly-based,

necessarily large and well-informed panel of expert technologists.

Such an effort can only be justified in those few cases where it is

necessary to obtain the distribution of intakes with maximum accuracy.

Given the wide safety margins food ingredients typically enjoy, and

the uncertainties and assumptions inherent in interpreting

toxicological data, such effort is rarely necessary. A much more

appropriate, but quick and easy, method of estimating intake was

needed, so long as it was conservative, but not excessively so. This

led Rulis and others in FDA (Rulis et al., 1984; Rulis, 1987) to

test and adopt the assumption that, except for an unusually narrow

margin of safety or specific concern, the per capita "eaters only"

intake could safely and conservatively be assumed - even for heavy

eaters - simply by multiplying the best available estimate of per

capita intake by ten (Column 5 ["High Eaters Only"]). Comparison of

Column 5 with Column 4b, for Section B, makes clear that in every case

the: "High Eaters Only" ( per capita × 10) estimate far exceeds the

"Eaters Only mean plus 3 standard deviations", yet in no case are the

estimates so high as to be unusable.

With the special factors surrounding the use of flavoring
ingredients (i.e., completeness of the surveys, typical volatility

with consequent loss in processing, and self-limiting nature) the

procedure in Column 5 is practical but highly conservative. It has

been widely used in other publications (FDA, 1982, 1993; Easterday

et al., 1993). It can be used safely in all instances except when a

very narrow margin of safety requires maximally precise data for both

toxicological effects and exposure.

Recently, still more sophisticated methods for analyzing USDA

Food Consumption Survey data have become available under license for

those applications in which the need for relatively high precision of

estimates justifies the cost (Helmbach, 1994). In nearly all instances

where broadly- based ingredient disappearance data are available,

per capita × 10, in actuality, a very high "eaters only" estimate,

remains the approach that is by far the simplest and least expensive,

while being conservative but realistic.

REFERENCES

Burdock, G.A., Wagner, B.M., Smith, R.L., Munro, I.C. and Newberne,

P.M. 1990. Recent progress in the consideration of flavoring

ingredients under the food additives amendment. 15. GRAS substances.

A list of flavoring ingredient substances considered generally

recognized as safe by the Flavor & Extract Manufacturers' Association

Expert Panel. Food Technol 44(2):78, 80, 82, 84, & 86.

Easterday, O.D., Ford, R.A., Hall, R.L., Stofberg, J., Cadby, P. and

Grundschober, F. 1992. A Flavor Priority Ranking System, Acceptance

and Internationalization. In: Finley, J.W., Robinson, S.F. and

Armstrong, D.J. (Eds.) Food Safety Assessment. American Chemical

Society, ACS Symposium Series No. 484.

FDA. 1982. Toxicological Principles for the Safety Assessment of

Direct Food Additives and Color Additives Used in Food. Redbook. U.S.

Food and Drug Administration, Bureau of Foods, Washington, DC.

FDA. 1993. Toxicological Principles for the Safety Assessment of

Direct Food Additives and Color Additives Used in Food. Redbook II

(Draft). U.S. Food and Drug Administration, Center for Food Safety and

Applied Nutrition, Washington, DC.

Hall, R.L. and Oser, B.L. 1965. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment. III. GRAS

substances. Food Technol 19(2):151-197.

Hall, R.L. and Oser, B.L. 1970. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment. IV. GRAS

substances. Food Technol 24(5):25-28, 30-32, 34.

Hall, R.L. 1975. Unpublished data from personal communication to the

NAS/NRC Committee on GRAS List Survey - Phase III, and to the FEMA.

Heimbach, J.T. 1994. Personal communication re: TAS-DIET from J.T.

Helmbach, Technical Assessment Systems, Inc., Washington, D.C.

NAS. 1973. A comprehensive Survey on the Use of Food Chemicals

Generally Recognized as Safe, National Technical Information Service

(NTIS) PB-221949.

NAS. 1976. Estimating Distribution of Daily Intakes of Certain GRAS

Substances. National Research Council, Washington, DC. Prepared for:

Food and Drug Administration, Washington, DC. December 1976. National

Technical Information Service (NTIS) PB-299-381.

NAS. 1978. Resurvey of the Annual Poundage of Food Chemicals Generally

Recognized as Safe, National Technical Information Service (NTIS)

PB288-081.

NAS. 1979. Comprehensive Survey of Industry on the Use of Food

Additives, National Technical Information Service (NTIS) PB80-113418.

NAS. 1984. Poundage Update of Food Chemicals, National Technical

Information Service (NTIS) PB84-162148.

NAS. 1989. 1987 Poundage and Technical Effects Update of Substances

Added to Food. National Research Council, Washington, DC. Prepared

for: Food and Drug Administration, Washington, D.C. NTIS Report

No. PB91-127266.

Oser, B.L. and Ford, R.A. 1973a. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 6. GRAS

substances. Food Technol 27(1):64-67.

Oser, B.L. and Ford, R.A. 1973b. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 7. GRAS

substances. Food Technol 27(11):56-57.

Oser, B.L. and Ford, R.A. 1974. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 8. GRAS

substances. Food Technol 28(9):76-80.
Oser, B.L. and Ford, R.A. 1975. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 9. GRAS

substances. Food Technol 29(8):70-72.

Oser, B.L. and Ford, R.A. 1977. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 10. GRAS

substances. Food Technol 31(1):65-74.

Oser, B.L. and Ford, R.A. 1978. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 11. GRAS

substances. Food Technol 32(2):60-70.

Oser, B.L. and Ford, R.A. 1979. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 12. GRAS

substances. Food Technol 33(7):65-73.

Oser, B.L. and Hall, R.L. 1972. Recent progress in the consideration

of flavoring Ingredients under the Food Additives Amendment 5. GRAS

substances. Food Technol 26(5):35-42.

Oser, B.L. and Hall, R.L. 1977. Criteria employed by the Expert Panel

of FEMA for the GRAS evaluation of flavoring substances. Food Cosmet

Toxicol 15:457-466.

Oser, B.L., Ford, R.A. and Bernard, B.K. 1984. Recent Progress in the

consideration of flavoring ingredients under the Food Additives

Amendment 13. GRAS substances. Food Technol 38(10):66-89.

Oser, B.L., Well, C.S., Woods, L.A. and Bernard, B.K. 1985. Recent

progress in the consideration of flavoring ingredients under the Food

Additives Amendment 14. GRAS substances. Food Technol 39(11):108-117.

Rulis, A.M., Hattan, D.G., and Morgenroth, V.H.,III. 1984. FDA's

priority-based assessment of food additives. I. Preliminary results.

Reg Toxicol Pharm 4:37-56.

Rulis, A.M. 1987. Safety assurance margins for food additives

currently in use. Reg Toxicol Pharm 7:160-168.

Smith, R.L. and Ford, R.A. 1993. Recent progress in the consideration

of flavoring ingredients under the Food Additives Amendment 16. GRAS

substances. Food Technol-June 1993, pp. 104-117.

ANNEX 5 Appendix B

SUBSTANCES IN REFERENCE DATABASE BY ORDER OF STRUCTURAL CLASS

Cramer et al. (1978) Structural Class I

1 acetic acid

2 acetoin

3 acetone

4 adipic acid

5 allura red AC

6 aminoundecanoic acid, 11-

7 ascorbic acid

8 ascorbic acid, 1-

9 benzaldehyde

10 benzoic acid

11 benzyl acetate

12 benzyl alcohol

13 bis(2-ethylhexyl)phthalate

14 brilliant black BN

15 butanediol, 1,3-

16 butanol, 2-

17 butanol, n18 butyl benzyl phthalate

19 butylated hydroxyanisole

20 butyrolactone, gamma21 calcium cyclamate

22 calcium formate

23 calcium stearoyl lactylate

24 carotenoic acid, beta-apo-8'-, methyl ester

25 cinnamaldehyde

26 citral

27 citranaxanthin

28 cumene

29 di(2-ethylhexyl)adipate

30 dibutyl phthalate

31 diethyl phthalate

32 diethylene glycol

33 diethylene glycol monoethyl ether

34 dimethoxane

35 dimethyl terephthalate

36 dimethylcarbonate

37 dimethyldicarbonate

38 dimethylphenol, 2,4-

39 dimethylphenol, 2,6-

40 dimethylphenol, 3,4-

41 dioctyl sodium sulfosuccinate

42 disodium 5'-guanylate
43 disodium 5'-inosinate

44 dodecyl gallate

45 ethanol

46 ethyl acetate

47 ethyl acrylate

48 ethyl butyrate

49 ethyl ether

50 ethyl formate

51 ethyl glycol monomethyl ether

52 ethyl heptanoate

53 ethyl nonanoate

54 ethyl-1-hexanol, 2-

55 ethylbenzene

56 ethylbutyric acid, 2-

57 ethylene glycol

58 ethylhexanoic acid, 2-

59 ethylphthalyl ethylglycolate

60 eugenol

61 FD & C blue No. 1

62 FD & C blue no. 2

63 formaldehyde

64 fumaric acid

65 geranyl acetate

66 glutamate, monosodium

67 glutamic acid hydrochloride

68 glutamic acid, 1-

69 glycerol

70 glyceryl tribenzoate

71 hexenal, 2- (trans)

72 hexylresorcinol, 4-

73 hydroquinone

74 hydroxybenzoic acid butyl ester, p75 hydroxybenzoic acid ethyl ester, p76 hydroxybenzoic acid methyl ester, p77 hydroxybenzoic acid propyl ester, p78 hydroxybenzyl acetone, p79 inosine monophosphate

80 ionone

81 isoamyl butyrate

82 isoamyl salicylate

83 isobutyl alcohol

84 isomaltitol

85 isopropyl alcohol

86 isovaleric acid

87 lactitol

88 limonene, d89 lithocholic acid

90 malonaldehyde, sodium salt

91 mannitol

92 mannitol, d93 menthol

94 methanol

95 methyl ethyl carbonate

96 methyl methacrylate

97 methyl salicylate

98 methyl-1-phenylpentan-2-ol, 4-

99 methylenebis, 2,2'-

100 methylphenol, 3-

101 methylphenylcarbinyl acetate

102 myrcene, beta103 nonalactone, gamma104 octyl acetate

105 octyl gallate

106 oleylamine

107 oxalic acid

108 phenol

109 phenoxyethanol

110 phenyl-1-propanol, 2-

111 phenylalanine

112 potassium sorbate

113 propyl gallate

114 propylene glycol

115 resorcinol

116 retinol

117 riboflavin

118 sodium benzoate

119 sodium erythorbate

120 sodium lauryl glyceryl ether sulfonate

121 sodium lauryl trioxyethylene sulfate

122 sodium stearoyl lactylate

123 sorbic acid

124 stearyl tartrate

125 styrene

126 sucrose monopalmitate

127 sucrose monostearate

128 tartaric acid

129 tertiary butyl hydroquinone

130 tocopherol, alpha131 tolualdehyde
132 toluene

133 triethylene glycol

134 triethylene glycol monomethyl ether

135 trimethylamine

136 undecalactone, gamma137 vanillin

138 xylitol

Cramer et al. (1978) Structural Class II

1 acrylic acid

2 allyl alcohol

3 allyl heptanoate

4 allyl hexanoate

5 butylated hydroxytoluene

6 caffeine

7 carotene, beta8 carvone

9 carvone, d10 diacetyl

11 diallyl phthalate

12 diketopiperazine

13 ethyl maltol

14 ethyl vanillin

15 ethylhexyl phthalate, mono-2

16 etretinate

17 fenthion

18 furfural

19 isobornyl acetate

20 isophorone

21 maltol

22 methyl amyl ketone

23 methyl anthranilate

24 piperidine

25 piperonal

26 propargyl alcohol

27 pyridine

28 thujone

Cramer et al, (1978) Structural Class III

1 (1-naphthyl)ethylene--diamine dihydro chloride, N2 (2-chloroethyl)trimethyl-ammonium chloride

3 (8-beta)N-cyclohexyl-6-methyl- 1 -(1 -methyl

ethyl)ergoline-8 -carboxamide

4 (chloroacetyl)-acetanilide, 4'-

5 1,1'-(2,2,2-trichloroethylidene)bis(4-chloro)-benzene

6 11-oxo-11H-pyrido(2,1-b)quinazoline-2-carboxylic acid

7 2(2,4,5-trichlorophenoxy) propionic acid

8 2-(2-methyl--4-chlorophenoxy)propionic acid

9 4-(2-methyl-4-chlorophenoxy) butyric acid

10 acenaphthene

11 acephate

12 acesulfame potassium

13 acetaminophen

14 acetoacetamide

15 acetoacetamide-N-sulfonic acid

16 acetochlor

17 acetonitrile

18 acifluorin sodium

19 acrylamide

20 acrylonitrile

21 alachlor

22 albendazole

23 albendazole sulfoxide

24 aldicarb

25 aldicarb sulfone

26 alinidine hydrobromide

27 allyl isovalerate

28 amaranth

29 ameltolide

30 ametryn

31 amino-4-ethoxy-acetanilide, 3-

32 amino-4-nitrophenol, 2-

33 amino-5-nitrophenol, 2-

34 aminophenol, p35 amitraz

36 ammonium carmine

37 amphetamine sulfate, dl38 ampicillin trihydrate

39 anethole, trans40 anilazine

41 anisidine hydrochloride, p42 anthranilic acid

43 arochlor 1254

44 aspartame

45 asulam

46 atrazine

47 avermectin B1

48 azaperone
49 azinphos methyl

50 azorubine

51 azuletil (KT1-32)

52 baythroid

53 benomyl

54 bentazon

55 benzofuran

56 benzoic acid, 2-[[[[N-(4-methoxy-6-methyl

-1,2,3-triazin-2-yl)-N-methylamino]carbonyl]

amino]sulfonyl] methyl ester

57 benzoin

58 benzyl violet 4B

59 benzyl-p-chlorophenol, o60 betaxolol

61 bidrin

62 biphenthrin

63 biphenyl, 1,1-

64 biphenylamine hydrochloride, 2-

65 bis(2-chloro-1-methyl-ethyl)ether Technical Grade

66 bis(2-chloroisopropyl)ether

67 bisphenol A

68 bromoacetic acid, 2-

69 bromodichloromethane

70 bromomethane

71 bromoxynil

72 bromoxynil octanoate

73 brown FK

74 butyl chloride, n75 butylate

76 calcium cyanamide

77 canthaxanthin

78 caprolactam

79 captafol

80 captan

81 carazolol

82 carbaryl

83 carbendazim

84 carbofuran

85 carbon tetrachloride

86 carbosulfan

87 carboxin

88 carmoisine

89 chloramben

90 chlordane

91 chlorendic acid

92 chlorimuron-ethyl

93 chloro-p-toluidine, 3-

94 chloroacetic acid

95 chloroaniline hydrochloride, p96 chlorobenzene

97 chlorobenzilate

98 chlorodibromomethane

99 chloroform

100 chlorofructose, 6-

101 chloronaphthalene, beta102 chlorophenol, 2-

103 chloropropylate

104 chlorothalonil

105 chlorotoluene, o106 chlorpheniramine maleate

107 chlorpromazine

108 chlorsulfuron

109 chlorthal dimethyl

110 chocolate brown HT

111 C.I. Acid Orange 12

112 C.I. Acid Orange 3

113 C.I. Acid Red 18

114 C.I. Disperse Blue 1

115 C.I. acid red 14

116 C.I. disperse yellow 3

117 C.I. pigment red 23

118 C.I. solvent yellow 14

119 cimetidine

120 clofentezine

121 clonitralid

122 closantel

123 codeine

124 coumaphos

125 coumarin

126 crufomate

127 curcumin

128 cyclodextrin, beta129 cyclohexylamine

130 cyclophosphamide

131 cyhalothrin

132 cypermethrin

133 cyromazine

134 daminozide

135 decabromodiphenyl oxide
136 deoxyquinine

137 diamino-2,2'-stilbenedisulfonic acid, 4,4'-, disodium sa

138 diaminophenol dihydrochloride, 2,4-

139 diazinon

140 dibenzo-p-dioxin

141 dibromobenzene, 1,4-

142 dicamba

143 dichloro-2-propanol, 1,3-

144 dichloro-p-phenylenediamine, 2,6-

145 dichlorobenzene, 1,2-

146 dichlorobenzene, 1,4-

147 dichlorobenzilic acid

148 dichlorodifluoromethane

149 dichloroethane, 1,1-

150 dichloroethane, 1,2-

151 dichloroethylene, 1,1-

152 dichloroethylene, trans- 1,2-

153 dichloromethane

154 dichlorophenol, 2,4-

155 dichlorophenoxyacetic acid, 2,4-

156 dichloropropanol, 2,3-

157 dichloropropene, 1,3-

158 dichlorvos

159 dicofol

160 dieldrin

161 diethyldithiocarbamate

162 diethylthiourea, N,N'-

163 difenzoquat

164 diflubenzuron

165 dihydroavermectin-B1 a, 22,23-

166 dihydroavermectin-B1 b, 22,23-

167 dihydrocoumarin, 3,4-

168 diiodomethyl p-tolyl sulfone

169 dimethipin

170 dimethoate

171 dimethoxyaniline hydrochloride, 2,4-

172 dimethoxybenzidine-4,4'-diisocyanate, 3,3'-

173 dimethyl hydrogen phosphite

174 dimethyl methylphosphonate

175 dimethyl morpholinophosphoramidate

176 dimethylaniline, N,N'-

177 dinitrobenzene, m178 dinitrotoluene, 2,4-

179 dinocap

180 dinoseb

181 diphenamid

182 diphenhydramine hydrochloride

183 diphenylamine

184 diphenylhydantoin, 5,5'-

185 diphenylhydantoin, 5,5-

186 diquat

187 disulfoton

188 dithiobiurea, 2,5-

189 diuron

190 dodecylguanidine acetate, n191 EDTA, disodium

192 endothall

193 ephedrine sulfate

194 epichlorohydrin

195 erythromycin stearate

196 erythrosine

197 ethalfuralin

198 ethephon

199 ethion

200 ethyl dipropylthiocarbamate, s201 ethyl p-nitrophenyl phenylphosphorothioate

202 ethylene chlorohydrin

203 ethylene thiourea

204 ethylmethylphenylglycidate

205 fast green FCF

206 febantel

207 fenamiphos

208 fenbendazole

209 fenchlorphos

210 fluometuron

211 fluoranthene

212 fluorene

213 fluridone

214 flurprimidol

215 flutolanil

216 fluvalinate

217 folpet

218 fonofos

219 fosetyl-al

220 glufosinate-ammonium

221 glyphosate

222 haloxyfop-methyl

223 HC blue no. 1

224 HC blue no. 2
225 HC yellow 4

226 heptachlor

227 heplachlor epoxide

228 hexabromobenzene

229 hexachlorobenzene

230 hexachlorobutadiene

231 hexachlorocyclohexane, gamma232 hexachlorocyclopentadiene

233 hexachloroethane

234 hexachlorophene

235 hexahydro-1,3,5-trinitro-1,3,5-triazine

236 hexamethylenetetramine

237 hexazinone

238 hexythiazox

239 hydralazine

240 hydrazobenzene

241 hydrochlorothiazide

242 hydroxypropyl methanethiolsulfonate, 2-

243 hydroxyquinoline, 8-

244 imazalil

245 imazaquin

246 imazethapyr

247 iodinated glycerol

248 ipazilide

249 iprodione

250 ipronidazole

251 isopropalin

252 isoxaben

253 jervine

254 lactofen

255 lansoprazole

256 levamisole

257 linamarin

258 linuron

259 londax

260 malaoxon

261 malathion

262 maleic anhydride

263 maleic hydrazine

264 manidipine hydrochloride

265 melamine

266 mepiquat chloride

267 mercaptobenzothiazole, 2-

268 merphos

269 merphos oxide

270 meso-2,3-dimercaptosuccinic acid

271 metalaxyl

272 methamidophos

273 methidathion

274 methomyl

275 methoxychlor

276 methoxypsoralen, 8-

277 methyl carbamate

278 methyl ethyl ketoxime

279 methyl parathion

280 methyl-4-chlorophenoxyacetic acid, 2-

281 methyl-N-methylanthranilate

282 methyldopa sesquthydrate, alpha283 methylolacrylamide, N284 metolachlor

285 metribuzin

286 metsulfuron methyl

287 mexacarbate

288 miporamicin

289 mirex

290 molinate

291 monodiethanolamine salt of riboflavin-5'-phosphate

292 monuron

293 myclobutanil

294 naled

295 nalidixic acid

296 naphthoxy acetic acid, 2-

297 napropamide

298 natamycin

299 nitro-o-toluidine, 5-

300 nitro-p-phenylenediamine, 2-

301 nitroaniline, p302 nitroanthranilic acid, 4-

303 nitrofurantoin

304 nitrofurazone

305 nitroguanidine

306 nitronaphthalene, 1-

307 nitrosodiphenylamine, N308 norflurazon

309 ochratoxin A

310 octabromodiphenyl ether

311 octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine

312 olaquindox

313 oryzalin

314 oxadiazon
315 oxamyl

316 oxfendazole

317 oxyfluorfen

318 oxytetracycline

319 oxytetracycline hydrochloride

320 oxythioquinox

321 paclobutrazol

322 paraquat

323 parathion

324 patulin

325 pendimethalin

326 penicillin VK

327 pentabromodiphenyl ether

328 pentachloroethane

329 pentachloronitrobenzene

330 pentachlorophenol

331 pentaerythritol tetranitrate

332 permethrin

333 phenformin

334 phenyl-2-naphthylamine, n335 phenyl-3-methyl-5-pyrazolone, 1-

336 phenylbutazone

337 phenylenediamine, m338 phenylephrine hydrochloride

339 phosmet

340 phosphamidon

341 photodieldrin

342 phthalamide

343 phthalic anhydride

344 picloram

345 pilsicainide hydrochloride

346 pirimiphos-methyl

347 pravadoline

348 probenecid

349 prochloraz

350 proflavine monohydrochloride hemihydrate

351 promethazine hydrochloride

352 prometon

353 prometryn

354 pronamide

355 propachlor

356 propanil

357 propargite

358 propazine

359 propham

360 propiconazole

361 propiverine hydrochloride (P-4)

362 propoxur

363 propylene dichloride

364 pydrin

365 pyrazinamide

366 pyrene

367 pyrimidinone

368 quercetin

369 quinalphos

370 quinine hydrochloride

371 quinofop ethyl

372 red 2G

373 reserpine

374 resmethrin

375 thudamine 6G

376 ronidazole

377 rotenone

378 saccharin

379 sethoxydim

380 simazine

381 sodium 2,2-dichloropropionate

382 sodium cyclamate

383 sodium fluoroacetate

384 sodium naphthionate

385 sorbitan monolaurate

386 sorbitan monostearate

387 succinic anhydride

388 sucrose acetate isobutyrate

389 sulfadimidine

390 sulfisuxazole

391 sulfolene, 3-

392 sunset yellow FCF

393 suplatast tosilate

394 tebuthiuron

395 terbacil

396 terbutryn

397 tert-butyl-2-chlorophenol, 4-

398 tetrachlorobenzene, 1,2,4,5-

399 tetrachloroethane, 1,1,1,2-

400 tetrachloroethane, 1,1,2,2-

401 tetrachloroethylene

402 tetrachlorophenol, 2,3,4,6-

403 tetrachloropyridine, 2,3,5,6-
404 tetrachlorvinphos

405 tetracycline hydrochloride

406 tetraethyldithiopyrophosphate

407 tetraethylthiourea disulfide

408 tetrahydrocannabinol, delta-9-

409 tetrakis(hydroxymethyl)phosphonium chloride (THPC)

410 tetrakis(hydroxymethyl)phosphonium sulphate (THPS)

411 thiabendazole

412 thiameturon methyl

413 thiobencarb

414 thiophanate-methyl

415 thiram

416 tocopheryl acetate, dl-alpha417 toluenediamine hydrochloride, 2,6-

418 toluenediamine sulfate, 2,5-

419 tralomethrin

420 trenbolone acetate

421 trenbolone hydroxide, 17-alpha422 triadimefon

423 triallate

424 triamterene

425 tribromomethane

426 trichloroacetonitrile

427 trichlorobenzene, 1,2,4-

428 trichloroethane, 1,1,2-

429 trichloroethane, 1,1,1-

430 trichloroethylene

431 trichloroethylene, 1,1,2-

432 trichlorogalactosucrose

433 trichlorophenol, 2,4,5-

434 trichlorophenoxyacetic acid, 2,4,5-

435 tridiphane

436 triethylene tetramine dihydrochloride

437 trifluralin

438 trimethylaniline, 2,4,5-

439 trinitrotoluene, 2,4,6-

440 tris-(2-chloroethyl) phosphate

441 trisulfuron

442 tylosin

443 vinclozolin

444 vinyl chloride

445 zatosetron maleate

446 zearalenone

447 zeranol

See Also:

Toxicological Abbreviations