Preparation of 
Food Contact Substance Notifications 
(Toxicology Recommendations): 
Guidance for Industry 
 
Additional copies are available from: 
Office of Food Additive Safety 
Division of Food Contact Substances HFS-275 
Center for Food Safety and Applied Nutrition 
Food and Drug Administration 
5001 Campus Drive 
College Park, MD  20740 
(Tel) 240-402-1175 
 
https://www.fda.gov/FoodGuidances 
You may submit electronic or written comments regarding this guidance at any time.  Submit 
electronic comments to https://www.regulations.gov.  Submit written comments to the 
Dockets Management Staff (HFA-305), Food and Drug Administration, 5630 Fishers Lane, 
rm. 1061, Rockville, MD 20852.  All comments should be identified with docket number 
FDA-1999-D-0062 and with the title of the guidance document.   
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Food Safety and Applied Nutrition  
 
October 2021 
 
OMB Control No. 0910-0495 
Current expiration date available at https://www.reginfo.gov/public/do/PRAMain 
*See additional PRA statement in Section XII of this guidance 
 
 
 
This is a revision to this guidance, which was originally issued in April 2002.  Revisions are 
noted by date at the end of the guidance. 
  Contains Nonbinding Recommendations 
Table of Contents 
 
 
 
 
I.  Introduction ......................................................................................................................... 3 
II.  Exposure Estimates ............................................................................................................. 4 
III.  Test Substance .................................................................................................................... 4 
IV.  Safety Testing Recommendations....................................................................................... 5 
A.  Minimum Testing Recommendations ..................................................................... 5 
1.  Incremental exposure at or less than 0.5 parts per billion (ppb) ..................... 5 
2.  Cumulative exposure greater than 0.5 ppb but not exceeding 50 ppb  ........... 5 
3.  Cumulative exposure between 50 ppb and 1 part per million (ppm) .............. 6 
4.  Cumulative exposure at or greater than 1 ppm ............................................... 7 
B.  Safety Testing Protocols ......................................................................................... 7 
C.  Application of the Testing Recommendations to Biocides..................................... 7 
D.  Genetic Toxicity Testing Recommendations .......................................................... 7 
E.  Flexibility in Applying FDAs Recommendations ................................................. 8 
V.  Organization of the Safety Information .............................................................................. 8 
VI.  Safety Narrative (SN) ......................................................................................................... 9 
VII.  Comprehensive Toxicology Profile (CTP) ....................................................................... 10 
A.  Preparation of Study Summaries for the CTP....................................................... 11 
B.  Determination of No-Observed-Effect Level (NOEL) ......................................... 12 
C.  Risk Assessment for Carcinogenic Constituents .................................................. 12 
D.  Bibliography ......................................................................................................... 13 
VIII.  FDAs Views of the Relevance of Various Safety Studies in Notifications..................... 13 
IX.  Evaluation of Structural Similarity to Known Toxicants ................................................. 15 
X.  Prenotification Consultation (PNC) Meetings .................................................................. 16 
XI.  Additional Toxicological Considerations in Deciding to Submit a Notification .............. 16 
XII.  Paperwork Reduction Act of 1995 .................................................................................... 17 
XIII.  References ......................................................................................................................... 17 
Document History ......................................................................................................................... 18 
  Contains Nonbinding Recommendations 
Preparation of  
Food Contact Substance Notifications 
(Toxicology Recommendations): 
1
Guidance for Industry
 
 
This guidance represents the current thinking of the Food and Drug Administration (FDA or we) 
on this topic.  It does not establish any rights for any person and is not binding on FDA or the 
public.  You can use an alternative approach if it satisfies the requirements of the applicable 
statutes and regulations.  To discuss an alternative approach, contact the FDA staff responsible 
for this guidance at the phone number listed on the title page. 
 
 
 
I.  Introduction 
Section 409 of the Federal Food, Drug, and Cosmetic Act (the FD&C Act) establishes a food 
contact substance notification (FCN) process as the primary means by which FDA regulates food 
additives that are food contact substances (FCSs).  An FCS is any substance that is intended for 
use as a component of materials used in manufacturing, packing, packaging, transporting, or 
holding food if the use is not intended to have any technical effect in the food (section 409(h)(6) 
of the FD&C Act). 
 
An FCS that is a food additive must be regulated for its intended use in 21 CFR Parts 173-178, 
be exempted from regulation under our Threshold of Regulation Policy (21 CFR 170.39), or be 
the subject of a notification under section 409(h) of the FD&C Act that is effective (section 
409(a)(3) of the FD&C Act).  Both FCNs and food additive petitions (FAPs) for FCSs must 
contain sufficient scientific information to demonstrate that the substance that is the subject of 
the notification or petition is safe for the intended use (sections 409(h)(1) and 409(b) of the 
FD&C Act).  Section 409(b) of the FD&C Act sets forth the statutory requirements for data in a 
FAP to establish the safety of a food additive.  These requirements include full reports of 
investigations made with respect to the safety of the additive.  Because the safety standard is the 
same for all food additives, whether subject to the FCN process or the petition process, the data 
and information that should be included in an FCN or FAP are comparable. 
 
 
1 This guidance has been prepared by the Office of Food Additive Safety, Division of Food Contact Substances in 
the Center for Food Safety and Applied Nutrition at the U.S. Food and Drug Administration. 
3 
  Contains Nonbinding Recommendations 
The contents of this document do not have the force and effect of law and are not meant to bind 
the public in any way, unless specifically incorporated into a contract.  This document is 
intended only to provide clarity to the public regarding existing requirements under the law.  
FDA guidance documents, including this guidance, should be viewed only as recommendations, 
unless specific regulatory or statutory requirements are cited. 
 
II.  Exposure Estimates 
The level of safety testing that is recommended to support an FCN for an FCS is largely 
determined by the cumulative estimated daily intake (CEDI) of the FCS.  The CEDI is the sum 
of the estimated daily intakes (EDIs) of the FCS that may result from the application of the 
substance described in the notification and any other regulated food uses of the substance.  For 
information on estimating human dietary exposures, refer to the document entitled Guidance for 
Industry: Preparation of Premarket Submissions for Food Contact Substances (Chemistry 
Recommendations). 
 
In some cases, limitations in the submitted chemistry information could affect the magnitude of 
an exposure estimate, and thereby affect the toxicological testing recommendations.  Therefore, 
FDA recommends that a notifier provide adequate information on the level of the FCS expected 
in foods in order for an estimate of the CEDI to reflect probable consumer exposure to the FCS 
and to ensure that the appropriate level of safety testing is conducted. 
 
FDA recognizes that the use of CEDI in this guidance appears to differ from the approach of 
FDAs Threshold of Regulation (TOR) process (21 CFR 170.39).  However, the two approaches 
are, in fact, consistent.  Under TOR, indirect food additive uses that result in incremental 
exposures at or less than 0.5 ppb in the diet are eligible for exemption from the FAP requirement.  
At the time the TOR process was established, FDA determined that, because of the conservative 
assumptions ordinarily applied in estimating exposure, the cumulative exposure from a limited 
number of trivial food additive uses is not likely to be more than negligible.  Accordingly, in the 
case of the TOR exposure levels, it was not necessary to utilize cumulative exposure levels.  
FDA believes that the determination made in establishing its TOR is still sound. 
 
III.  Test Substance 
FDA generally recommends that the test substance for safety studies be identical to the substance 
that is expected to migrate to food.  Ordinarily, the appropriate test substance is the FCS itself.  
In some cases, however, appropriate test substances may include various constituents of the FCS, 
such as minor components, materials used in manufacturing, or decomposition products, if these 
constituents are expected to migrate to food.  For example, for an FCS that is a polymer, FDA 
recommends testing low-molecular weight oligomers for toxicity, but not the polymer itself, as 
the oligomers may be expected to be the primary migrant to food from the FCS. 
 
Some FCSs decompose to other substances that exert technical effects either during the 
manufacture of food contact materials (e.g., slimicides) or in food contact materials themselves 
(e.g., phosphorus-based antioxidants in which phosphorus oxidizes to phosphates and 
phosphites).  Other FCSs decompose as a consequence of imparting their technical effect or are 
4 
  Contains Nonbinding Recommendations 
known to decompose during processing, in storage, and in food or food-simulating solvents (e.g., 
antioxidants in polymers).  In such cases, decomposition products of the FCSs may be 
appropriate test substances for safety studies. 
 
Test and control substances used in the safety studies should be characterized and handled in 
accordance with the Good Laboratory Practice (GLP) regulations for non-clinical laboratory 
studies in 21 CFR 58.105 (Test and Control Articles).  In all cases, the composition of the test 
substance used in safety studies should be known.  Notifiers should provide the names, structural 
formulae, and quantities of major components and other constituents of the test substance, and 
the approximate total quantity of unidentified material.  If available, both common names and 
trade names should be provided.  A single batch of a test substance should be used for a safety 
study, if possible.  If more than one batch is used, the strength, composition, purity, and other 
characteristics of each batch should be approximately the same. 
 
Additional information on the chemical identity of the FCS and its constituents is contained in 
the document Guidance for Industry: Preparation of Premarket Submissions for Food Contact 
Substances (Chemistry Recommendations).  For guidance on safety studies for specific test 
substances, notifiers are advised to contact FDA. 
 
IV.  Safety Testing Recommendations 
A.  Minimum Testing Recommendations 
FDA recommends studies to assess the safety of an FCS and its constituent(s) if 
appropriate, on the basis of the CEDI (see II).  These recommendations are consistent 
with the general principle that the potential risk of a substance is likely to increase as 
exposure increases. 
 
FDA recommends that notifiers submit, as a minimum, the following studies and other 
information to assess the safety of an FCS (and each constituent as appropriate): 
 
1.  Incremental exposure at or less than 0.5 parts per billion (ppb) (i.e., 1.5 
micrograms (g/person/day) in the diet 
a.  No safety studies are recommended for an FCS (or a constituent, as 
appropriate) if exposure for a single use is at or less than 0.5 ppb. 
b.  Available information on the potential carcinogenicity of such substances 
should be discussed in a comprehensive toxicology profile (CTP) (e.g., 
carcinogenicity studies, genetic toxicity studies, or information on 
structural similarity to known mutagens or carcinogens (see IX)). 
c.  For a carcinogenic constituent of an FCS, the CTP should contain an 
estimate of the potential human cancer risk from the constituent due to the 
proposed use of the FCS (see VII. C.). 
2.  Cumulative exposure greater than 0.5 ppb (i.e., 1.5 g/person/day) but not 
exceeding 50 ppb (i.e., 150 g/person/day) 
a.  The potential carcinogenicity of an FCS (and/or a constituent, if 
appropriate) with a cumulative exposure between 0.5 ppb and 50 ppb 
5 
  Contains Nonbinding Recommendations 
should be evaluated using genetic toxicity tests.  The recommended 
genetic toxicity tests include: (1) a test for gene mutations in bacteria and 
(2) an in vitro test with cytogenetic evaluation of chromosomal damage 
using mammalian cells or an in vitro mouse lymphoma thymidine kinase 
(tk) gene mutation assay.  FDA prefers the mouse lymphoma tk gene 
mutation assay because this assay measures heritable genetic damage in 
living cells and is capable of detecting chemicals that induce either gene 
mutations or chromosomal aberrations, including genetic events associated 
with carcinogenesis.  In performing the mouse lymphoma tk assay, either 
the soft agar or the microwell method should be used. 
b.  Additional information on the potential carcinogenicity of such a 
substance should be discussed, as appropriate, in CTPs (e.g., 
carcinogenicity studies, genetic toxicity studies, information on structural 
similarity to known mutagens and carcinogens (see IX), etc.). 
c.  For a carcinogenic constituent of an FCS, the CTP should estimate the 
potential human cancer risk from the constituent due to the proposed use 
of the FCS (see VII.C.). 
3.  Cumulative exposure between 50 ppb (i.e., 150 g/person/day) and 1 part per 
million (ppm) (i.e., 3 mg/person/day) 
a.  The potential carcinogenicity of an FCS (and/or a constituent, if 
appropriate) with an estimated cumulative exposure between 50 ppb and 1 
ppm should be evaluated using genetic toxicity tests.  The recommended 
genetic toxicity tests include: (1) a test for gene mutations in bacteria; (2) 
an in vitro test with cytogenetic evaluation of chromosomal damage using 
mammalian cells or an in vitro mouse lymphoma tk gene mutation assay 
(the mouse lymphoma assay is preferred); and, (3) an in vivo test for 
chromosomal damage using rodent hematopoietic cells.  In performing the 
mouse lymphoma tk gene mutation assay, either the soft agar or the 
microwell method should be used. 
b.  Additional information on the potential carcinogenicity of such a 
substance should be discussed, as appropriate, in CTPs (e.g., 
carcinogenicity studies, genetic toxicity studies, information on structural 
similarity to known mutagens and carcinogens (see IX), etc.). 
c.  For a carcinogenic constituent of an FCS, the CTP should estimate the 
potential human risk from the constituent due to the proposed use of the 
FCS (see VII.C.). 
d.  The potential toxicity of an FCS (and/or a constituent, if appropriate) 
should be evaluated by two subchronic oral toxicity tests, one in a rodent 
species and one in a non-rodent species.  The studies should provide an 
adequate basis for determining an acceptable daily intake (ADI) for the 
FCS or a constituent in the indicated range of CEDIs.  In addition, the 
results of these studies will help determine whether longer-term or 
specialized safety tests (e.g., metabolism studies, teratogenicity studies, 
reproductive toxicity studies, neurotoxicity studies, and immunotoxicity 
studies) should be conducted to assess the safety of these substances. 
6 
  Contains Nonbinding Recommendations 
4.  Cumulative exposure at or greater than 1 ppm (i.e., 3 mg/person/day) 
When the estimated exposure to an FCS or a constituent is 1 ppm or greater, FDA 
recommends that a FAP be submitted for the FCS (see XI). 
B.  Safety Testing Protocols 
FDA provides general guidance on the conduct of standard toxicity tests, other than 
genetic toxicology tests, and it is relevant to toxicity testing of FCSs and their 
constituents.  (See Guidance for Industry and Other Stakeholders: Toxicology Principles 
for the Safety Assessment of Food Ingredients (Redbook 2000).) 
 
The Redbook 2000 sections available on FDAs internet site include guidelines on the 
conduct of certain genetic toxicity tests.  For genetic toxicity tests not yet found on this 
website, FDA recommends that notifiers consult the testing guidelines published by 
the Organization for Economic Co-operation and Development (OECD) or the guidelines 
of the United States Environmental Protection Agency (EPA) and the genotoxicity 
guidelines of the International Conference on Harmonization (ICH) of Technical 
Requirements for Registration of Pharmaceuticals for Human Use. 
 
Alternative procedures for conducting safety tests may be used.  In such cases, FDA 
recommends that notifiers consult us on proposed deviations from recommended safety 
test protocols before the tests are conducted. 
 
All safety studies should be conducted according to FDAs GLP regulations, or the GLP 
guidelines of the EPA, or the guidelines of the OECD.  If a study was not conducted in 
compliance with these regulations or guidelines, a brief statement of the reason for 
noncompliance should be given.  For a safety study conducted after 1978 that does not 
comply with FDA GLP regulations, FDA recommends that notifiers include a report of a 
data audit by an independent third party auditor if the study is pivotal to assessing the 
safety of the FCS. 
 
C.  Application of the Testing Recommendations to Biocides 
Biocides are a class of FCSs that are toxic by design.  Consequently, FDA recommends 
that notifiers apply FDA's minimum testing recommendations (see IV.A.) to biocides at 
CEDIs that are 1/5 the value of the CEDIs used to determine the appropriate level of 
safety testing for other types of FCSs.  FDA considers these lower exposure cutoffs 
appropriate for FCSs used primarily for their antimicrobial or fungicidal effects. 
D.  Genetic Toxicity Testing Recommendations 
For an FCS with a cumulative exposure greater than 0.5 ppb, FDA recommends that 
genetic toxicity testing be done.  This is because carcinogenicity is an ongoing health 
concern at low levels of exposure and genetic toxicity testing is the most reliable 
experimental indicator of potential carcinogenicity, with the exception of full-scale 
chronic animal carcinogenicity studies. 
7 
  Contains Nonbinding Recommendations 
 
In some cases, genetic toxicity testing may not be useful, or the recommendations that are 
provided above may need to be modified.  For example, FDA believes that genetic 
toxicity testing of polymers is unnecessary and that testing of oligomers and other 
constituents that can migrate into foods is more appropriate. 
 
E.  Flexibility in Applying FDAs Recommendations 
The information and recommendations provided in this document are intended to help 
ensure that sufficient safety information is available on an FCS and its constituent(s) to 
determine whether the substance is safe under its intended conditions of use.  Although 
the information contained in this document represents FDAs current thinking on the 
safety information needed to establish the safety of an FCS and its constituents, an 
alternative approach may be used by a notifier if the approach satisfies the applicable 
statute and regulations. 
 
The information and recommendations discussed in this document permit notifiers to 
exercise their own judgment in selecting safety tests to be performed for FCSs.  The level 
of testing and types of safety information needed to determine the safety of a particular 
FCS or its constituent(s) should be evaluated on a case-by-case basis.  Intended use, 
potential acute and chronic toxicity (e.g., signs/symptoms of neurotoxicity and 
hyperplasia, respectively), and structural alerts are some of the factors that should be 
considered. 
V.  Organization of the Safety Information 
FDA recommends that the notifier organize the safety information into two parts.  The first part 
of the safety information should be provided in Part III of FDA Form 3480.  The second part of 
the safety information is the safety data package attached to FDA Form 3480. 
 
Part III of FDA Form 3480 is the safety summary.  The safety summary in Part III of FDA Form 
3480 is divided into three sections: Section A- Safety Narrative, Section B- Comprehensive 
Toxicology Profile(s), and Section C- Relevant Toxicity Studies.  Detailed information on 
preparing the safety narrative (SN) (Section A of FDA Form 3480) is provided in this guidance 
document (see VI). 
 
The second part of the safety information in the notification is the safety data package.  FDA 
recommends that the notifier organize the safety data package as follows: 
 
  Section I. Comprehensive Toxicology Profile(s) 
  Section II. Original Reports of Safety Studies 
  Section III. Published literature 
  Section IV. Appendices 
 
Detailed information for preparing the comprehensive toxicology profile(s) (Section I of safety 
data package) is provided in this guidance document (see VII). 
8 
  Contains Nonbinding Recommendations 
 
Section II of the safety data package should contain the original reports of safety studies and 
Section III should contain the published literature (i.e., data or information that the notifier relied 
upon to prepare Section I).  When available, full study reports, including the primary data (i.e., 
individual animal data, plate counts, etc.), should be submitted for all of the recommended safety 
studies, cancer bioassays, and other pivotal studies on the FCS and its constituents, as 
appropriate.  The original study reports should be included in the safety data package whether 
conducted by the notifier or by a third party.  It is particularly important that notifiers submit full 
study reports of studies and related information that are used quantitatively, for example, to 
conduct risk assessments or set no-observed-effect levels (NOELs).  For clarification or to 
determine if the full study report for a specific safety study should be included in an FCN, 
notifiers are advised to contact FDA. 
 
Section IV of the safety data package should include appendices with data and other information 
not addressed in other sections of the safety data package.  Such data typically would have been 
considered by the notifier and judged to be supplementary.  The inclusion of such information in 
this section is intended to permit FDA to make an independent assessment of the utility of such 
information.  In particular, FDA recommends that notifiers include abstracts of available studies 
not discussed in the CTP in this section with a statement regarding the notifier's rationale for 
their exclusion.  If such studies and information are voluminous, FDA recommends that the 
notifier contact FDA before preparing such an appendix.  In addition, the appendix should 
include the results of all literature searches conducted and information relevant to the searches 
(e.g., names of selected databases, the period of years searched, the specific search terms used, 
etc.) under a separate heading.  Other information in Section IV might include material safety 
data sheets, book chapters, review articles, etc. 
 
VI.  Safety Narrative (SN) 
Each notification should contain a safety narrative (SN).  A SN is a concise summary of the 
scientific basis for a safety decision.  Ordinarily, the SN should reference the estimated human 
exposure and potential toxicity of the FCS and its constituent(s) and should be based on 
chemistry and safety information and analyses described in detail in other sections of the 
notification.  In the SN, the notifier should be explicit in reporting all effects of an FCS, 
including those considered adverse or physiologic.  The SN should also include conclusions 
regarding the mutagenic and carcinogenic potential of the FCS and any toxicologically relevant 
constituents, as appropriate.  Furthermore, the SN should provide the appropriate worst-case, 
upper-bound, lifetime risk levels for carcinogenic constituents associated with the FCS.  
However, a detailed quantitative risk assessment procedure for carcinogenic constituents of FCSs 
is not needed in this section. (See VII.C.)  If an ADI for the FCS is determined, it should be 
justified in terms of the most relevant study and end-point chosen, the animal species selected, 
and the safety (or uncertainty) factor applied.  Generally, an ADI for an FCS with a CEDI below 
50 ppb is not available.  In cases where appropriate studies are available, an ADI may be 
calculated.  If a previously established ADI supports the new intended use of an FCS, this should 
be discussed in the SN. 
 
9 
  Contains Nonbinding Recommendations 
To calculate an ADI, the NOEL for each identified adverse effect from all relevant safety studies 
should be multiplied by an appropriate safety factor.  Information on determining the NOEL is 
given in Section VII.B of this guidance document.  In general, FDA recommends that the notifier 
use a safety factor of 1/1000 if NOELs are derived from subchronic studies and 1/100 for 
NOELs derived from chronic studies.  For reproduction and developmental endpoints, FDA 
recommends that the notifier use a safety factor of 1/1000 if the observed effects are severe or 
irreversible (e.g., a missing limb or decrease in the number of pups born live); otherwise, FDA 
recommends a safety factor of 1/100.  Additional adjustments may be appropriate when 
considered on a case-by-case basis. 
 
Traditionally, the lowest ADI would be chosen as the definitive ADI, unless there is scientific 
rationale not to do so (e.g., if a toxicological effect seen in animals is shown not to occur in 
humans). 
 
VII.  Comprehensive Toxicology Profile (CTP) 
Each notification should include a CTP of all unpublished and published safety studies and 
related information relevant to the safety assessment of the FCS.  If there are constituent(s) of the 
FCS that are expected to migrate to food, then a CTP for each constituent of potential 
toxicological concern should also be provided in the notification. 
 
In preparing a CTP, all safety studies that identify adverse effects of the substance or that bear 
significantly on the determination of an ADI for the substance should be addressed.  FDA's 
views on the relevance, in general, of various types of safety studies are discussed below (see 
VIII) and should be considered in preparing the CTP. 
 
If the test substance in a specific study that is addressed in the CTP differs from the FCS, its 
relationship to the FCS should be clearly indicated.  For example, the test substance should be 
identified as a constituent of the FCS (e.g., monomer, oligomer, decomposition product, side 
reaction product or impurity, as appropriate). 
 
FDAs recommendations on preparing key components of the CTP, including study summaries, 
determination of NOELs, risk assessments, and bibliography are provided below. 
   
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  Contains Nonbinding Recommendations 
A.  Preparation of Study Summaries for the CTP 
1.  Study Summaries for Genetic Toxicity Studies 
The potential for genetic toxicity is an important consideration in the safety 
evaluation of FCSs.  Information on the genetic toxicity of the FCS and its 
constituents should be described in detail in the CTPs.  In evaluating the safety of the 
FCS and its constituents, notifiers should consider all published and unpublished 
genetic toxicity data. 
 
In summarizing genetic toxicity studies, FDA recommends that the notifier: 
  Group the available data by test systems (e.g., gene mutations in bacteria, 
o
gene mutations in cultured mammalian cells, chromosomal aberrations in 
vitro, chromosomal aberrations in vivo, etc.).  Individual studies within the 
same test system should be presented in chronological order. 
  Prepare a table of the genetic toxicity data for the FCS and its constituents, if 
o
appropriate. 
  Formulate and justify an overall conclusion regarding the genotoxic potential 
o
of the FCS and its constituents, if appropriate. 
2.  Study Summaries for in vivo Toxicity Tests 
Standard in vivo toxicity tests of the FCS and its constituents should be described in 
detail in the CTP.  Both unpublished and published safety data should be included 
and presented in an organized fashion.  Study reports and published articles of the 
same study type (i.e., subchronic, chronic, reproductive, etc.) should be grouped by 
species (e.g., mouse, rat, dog, etc.), then summarized in chronological order within 
each grouping.  The following is one example of an outline that a notifier could 
follow to organize the studies within the CTP: 
  Acute toxicity studies (may be presented in tabular form) 
o
  Short-term toxicity studies 
o
  Subchronic toxicity studies 
o
  Mouse 
  Rat 
  Dog 
  Other species 
  Reproductive and developmental studies 
o
  Chronic studies (by species). 
o
  Carcinogenicity studies 
o
  Special studies (including in vitro studies, as appropriate) 
o
FDA recommends that each individual study summary include the following 
minimum information: 
  Identity of test substance 
o
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  Contains Nonbinding Recommendations 
  Animal species and strain(s) tested 
o
  Number of animals/sex/dose and control groups 
o
  Route of administration 
o
  Doses milligram/kilogram bodyweight/day (mg/kg bw/d), frequency and 
o
duration of dosing, and dosing vehicle(s), if any 
  Other elements of study design, as appropriate (e.g., recovery phase, culling 
o
method, interim sacrifice, etc.) 
  Parameters measured (e.g., clinical signs, clinical laboratory tests, organ 
o
weights, histopathology, etc.) and the frequency of measurements 
  Significant, compound-related effects (including doses at which effects were 
o
observed, incidences of animals with effects, etc.) 
  Highest dose(s) at which no substance-related effect(s) were observed (i.e., 
o
NOEL for each effect.) 
B.  Determination of No-Observed-Effect Level (NOEL) 
A NOEL should be determined by the most sensitive, non-neoplastic adverse effect 
identified from relevant safety studies.  The NOEL should be expressed in terms of mg 
per kg body weight per day of the test animal. 
If the levels of the FCS or constituents given to test animals in a study are expressed as 
percent or parts per million in the diet, the notifier should report the NOEL using these 
units and also calculate intake on a mg/kg bw/d basis. In these cases, the notifier should 
indicate if actual food consumption data were used in such calculations.  A summary 
table of the adverse effects observed, if any, should be prepared by study type (i.e., 
subchronic, chronic, reproductive, etc.) to facilitate the evaluation and determination of 
NOELs for all of the substance-related effects. 
 
C.  Risk Assessment for Carcinogenic Constituents 
The CTP should include risk assessments for carcinogenic constituents of FCSs, as 
appropriate.  The Delaney clause of the FD&C Acts food additive provisions prohibits 
the approval of carcinogenic food additives including FCSs (section 409(c)(3)(A)).  
Importantly, however, the Delaney clause applies to the additive itself and not to 
constituents of the additive.  Therefore, if a food additive that is an FCS, has not been 
shown to cause cancer but contains a carcinogenic constituent, FDA evaluates the 
constituent under the general safety standard (section 409 (c)(3)(A) of the FD&C Act) 
using quantitative risk assessment procedures. 
 
If the results of epidemiology studies or rodent carcinogenicity studies on the constituent 
are either positive or equivocal, the notifier ordinarily should calculate an extreme-case, 
upper-bound, lifetime risk to humans from exposure to the constituent.  A notifier may 
use another approach to estimate the risk presented by a carcinogenic constituent, and 
should present convincing scientific evidence to justify the alternative approach to 
estimate the risk.  In calculating the risk, the notifier should: 
 
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  Contains Nonbinding Recommendations 
1.  Use the tumor data from the most sensitive species, strain, sex, and study; 
2.  Assume that tumors arising at multiple sites are independent of each other and 
add their risks; and 
3.  Calculate the extreme-case, upper-bound, lifetime risk by multiplying the unit 
cancer risk by the estimated human exposure to the constituent based on its use 
level in the notification.  The unit cancer risk is defined as the slope of a straight 
line drawn from the lowest apparent effect dose to zero.  FDA has calculated the 
unit risk for some constituents of FCSs; these are available upon request. 
 
General information on FDAs approach to risk assessment is contained in publications 
by Kokoski et al. (1990) and Lorentzen (1984).  For more specific information on the 
Center for Food Safety and Applied Nutrition's quantitative risk assessment procedures, 
notifiers should contact FDA. 
D.  Bibliography 
The CTP should include a bibliography with all references listed alphabetically.  All 
published and unpublished studies and information presented in the CTP should be 
referenced appropriately in the text by citing the author(s) and year of publication.  Each 
published reference should include the names of all authors, the year of publication, the 
full title of the article, pages cited, and name of publication.  For a book, the reference 
also should include the title of the book, the edition, the editor(s), and the publisher.  
Reference to unpublished studies should identify all authors, the sponsor of the study, the 
laboratory conducting the study, the final report date, the full title of the final report, the 
report identification number, and inclusive page numbers.  References to government 
publications should include the department, bureau or office, title, location of publisher, 
publisher, year, pages cited, publication series, and report number or monograph number. 
VIII.      FDAs Views of the Relevance of Various Safety Studies in 
Notifications 
With the exception of acute studies, FDA considers safety studies in which the test substance is 
given via the oral route most relevant to the safety assessment of substances in food.  The data 
collected from studies using other routes of administration, including inhalation and dermal 
studies, may be of value if systemic effects at distal sites are observed.  Only studies and 
information that are relevant to the safety assessment of a substance in food need be discussed in 
the CTP. 
 
Below, FDAs views on the relevance of various types of toxicological studies to the safety 
assessment of an FCS are discussed in brief. 
 
A.  Acute Toxicity Studies 
Acute toxicity data, including median lethal dose (LD ) values, rarely are used in the overall 
50
safety assessment of FCSs to which long-term repeated exposure of consumers is expected.  
It is not necessary to discuss acute studies individually.  An exception may be where there is 
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significant and useful information that is provided by the acute toxicity study that may 
provide clues as to the potential target organs for the compounds adverse effect(s).  
Otherwise, the results of acute toxicity studies should be summarized in a table. 
 
B.  Genetic Toxicity Studies 
FDA believes that information on the genetic toxicity of a substance is critical to the safety 
assessment of that substance because, in the absence of carcinogenicity data, genetic toxicity 
studies may be used to draw conclusions about its potential carcinogenicity. 
 
Factors that should be considered in determining whether results of genetic toxicity studies 
indicate a potential safety concern for an FCS include: 
 
1.  Other available safety data such as bioassays; 
2.  The quality of the genetic toxicity studies; 
3.  The array of positive and negative genetic toxicity test results; and 
4.  The chemical structure of the substance (see IX). 
C.  Short-term Toxicity Studies 
Short-term toxicity studies in animals, usually only 7 to 28 days in duration, should not be 
used to establish an ADI for an FCS.  However, individual summaries of short-term studies 
should be included in the CTP.  The summary of these studies should discuss endpoints or 
target organs potentially associated with toxicity and dose levels that would be appropriate 
for longer-term toxicity studies. 
D.  Subchronic Toxicity Studies 
NOELs from subchronic toxicity studies often are the basis for determining ADIs for FCSs.  
In such cases, it is important to provide complete summaries of subchronic studies, including 
detailed discussions of the study results in the CTP.  If the primary objective of a subchronic 
study is to identify the target organ or select doses for a longer study, it may be appropriate to 
emphasize these objectives in the study summary.  If subchronic studies are available in 
different species, species differences, if any, should be discussed. 
E.  Reproductive and Developmental Toxicity Studies 
NOELs from reproductive and developmental toxicity studies may be the basis for 
determining ADIs for FCSs.  Therefore, a summary and detailed discussion of the results of 
each study should be provided.  For both parental animals and their offspring in each 
generation, NOELs should be identified for all substance-related changes.  The summaries 
should state which effect(s) were used to derive NOELs.  The toxicological relevance of any 
reported changes should be evaluated and, if observed, the impact of concurrent maternal 
toxicity on the results of the study should be addressed. 
   
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F.  Chronic Toxicity Studies 
If chronic toxicity studies are available, the results of these studies will ordinarily supersede 
subchronic studies results for the purpose of assessing the safety of an FCS.  Due to the 
longer duration of these studies, toxic effects may be identified that would not be detected in 
shorter-term studies.  In the CTP, the results of chronic rodent or non-rodent studies should 
be summarized and discussed in detail. 
G.  Carcinogenicity Studies 
Carcinogenicity studies are relevant to the safety assessment of FCSs and their constituents.  
When such studies are available, all neoplastic and non-neoplastic study observations should 
be discussed.  Summary tables of treatment-related neoplastic and non-neoplastic lesions at 
any organ/tissue site should be prepared.  The incidence of test animals with benign and 
malignant tumors at a specific organ site, both separately and combined, should be provided 
as appropriate (McConnell et al., 1986; National Toxicology Program (NTP) Guidelines).  If 
available, a detailed morphological description of any significant lesions should be included.  
Statistical trend tests should be performed in addition to tests of significance between dose 
and control groups.  In addition, all effects observed should be evaluated for potential 
biological relevance.  Related histopathological information, such as time to tumor formation 
and historical tumor data from performing laboratories, should be discussed.  Reports 
prepared by NTP provide good examples of how the histopathological data requested above 
should be presented.  The CTP should state clearly whether the FCS was associated with 
neoplastic or pre-neoplastic changes and discuss whether the incidence, location, and type of 
tumors observed in this study demonstrate any carcinogenic effects attributable to the FCS or 
its constituents, as appropriate.  Note that the detailed information described above is 
particularly important to support a conclusion that no carcinogenic effects were observed in a 
study. 
H.  Special Studies 
Special studies include metabolism and pharmacokinetic studies, and other studies designed 
to test specific types of toxic effects in animals (e.g., neurotoxicity, immunotoxicity).  
Clinical studies, and observations reported in humans are also considered special studies.  
Ordinarily, clinical studies are not a part of the testing paradigm for FCSs.  However, if 
clinical studies are available, individual study summaries should be provided in the CTP.  
The results of clinical studies may affect the ADI determination for an FCS. 
IX.  Evaluation of Structural Similarity to Known Toxicants 
It is reasonable to expect that the chemical structure and physicochemical properties of FCSs and 
their constituent(s) are potential determinants of toxicity.  To the extent feasible, discussions or 
explanations that predict toxicity based on structure/activity relationships may be incorporated 
into the safety assessment of FCSs and their constituent(s).  When appropriate, expert analysis, 
decision-tree procedures (e.g., Cramer et al., 1978), or computer-assisted quantitative 
structure/activity techniques may be used to relate the chemical structure of a substance with a 
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toxicological endpoint of interest.  Such information should not be considered as a substitute for 
actual data, but may be useful in developing an overall strategy for assessing the safety of a 
substance and interpreting the results of carcinogenicity and other types of safety studies. 
 
X.  Prenotification Consultation (PNC) Meetings 
A notifier may request a PNC meeting regarding a notification for an FCS.  Many notifications 
will not require PNC interactions between FDA and the notifier.  Such interactions will occur at 
the discretion of the notifier and are intended to facilitate the submission of successful 
notifications since notifications without adequate scientific support will not be accepted.  FDA 
considers all PNC meetings consultative in nature.  PNC meetings should not be considered 
determinative with respect to FDAs decision to accept or object to a notification submitted to 
FDA subsequent to a PNC meeting. 
 
One example of when a PNC meeting might be helpful is when the ADI/CEDI ratio is less than 
five.  In such cases, the notifier may wish to request a meeting before submitting a notice to 
discuss possible interpretive differences in establishing a NOEL to calculate an ADI.  Because 
dosing levels in safety studies are often spaced by a factor of three, the determination of the 
NOEL would seldom be expected to differ by more than a single dose.  Therefore, FDA believes 
that when the ADI/CEDI ratio is less than five, a PNC meeting should be considered. 
 
PNC meetings may also be helpful when there are questions regarding the carcinogenicity of an 
FCS, significant risk potentially associated with a carcinogenic constituent, or when there are 
equivocal mutagenicity data. 
 
XI.  Additional Toxicological Considerations in Deciding to Submit 
a Notification 
FDAs experience in evaluating the safety of FCSs and their constituents indicates that situations 
may arise in which an FCN will be appropriate for the use of an FCS even if the cumulative 
exposure for the FCS or its constituents is at or greater than 1 ppm, or 200 ppb in the case of 
biocides.  Examples of such cases are provided below. 
 
An FCN may be appropriate for an FCS, even if the estimated cumulative exposure is greater 
than 1 ppm, or 200 ppb for biocides, when: 
 
There is an existing ADI for the FCS and its constituent(s).  In such a case, the notifier 
should contact FDA to determine the applicability of the ADI for the FCS, before 
submitting an FCN. 
 
A large database is available on a close structural analog of the FCS and its 
constituent(s), which analog has been approved by FDA.  In such cases, the following 
toxicological tests are recommended to demonstrate the degree of toxicological and 
metabolic similarity between the FDA-regulated analog and the FCS and its 
constituent(s): 
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a.  One 90-day oral toxicity study in a rodent or non-rodent species; and 
b.  Comparative absorption, distribution, metabolism, and elimination studies. 
 
The FCS and/or its constituent(s) is poorly absorbed or is not absorbed from the 
gastrointestinal tract.  Such assertions should be supported by relevant scientific 
information or data. 
 
The FCS undergoes chemical or metabolic transformation solely to products known to be 
of little toxicological concern at the estimated level of CEDI.  Such assertions should be 
supported by relevant in vivo or in vitro data. 
 
XII.  Paperwork Reduction Act of 1995  
This guidance contains information collections that are subject to review by the Office of 
Management and Budget (OMB) under the Paperwork Reduction Act of 1995 (44 U.S.C. 3501-
3521).   
 
The time required to complete this information collection is estimated to average 25 to 150 hours 
per response, including the time to review instructions, search existing data resources, gather the 
data needed, and complete and review the information collection.  Send comments regarding this 
burden estimate or suggestions for reducing this burden to: 
 
Office of Food Additive Safety 
Division of Food Contact Substances, HFS-275 
Center for Food Safety and Applied Nutrition 
Food and Drug Administration 
5001 Campus Drive 
College Park, MD 20740 
 
 
An agency may not conduct or sponsor, and a person is not required to respond to, a 
 
collection of information unless it displays a currently valid OMB control number.  
 
The OMB control number for this information collection is 0910-0495 (To find the 
 
current expiration date, search for this OMB control no. available at 
 
https://www.reginfo.gov/public/do/PRAMain). 
 
 
 
XIII.  References   
The following references marked with an asterisk (*) are on display at the Dockets Management 
Staff, (HFA-305), Food and Drug Administration, 5630 Fishers Lane, Rm. 1061, Rockville, MD 
20852, 240-402-7500 and are available for viewing by interested persons between 9 a.m. and 4 
p.m., Monday through Friday; they are also available electronically at 
https://www.regulations.gov.  References without asterisks are not on public display at 
https://www.regulations.gov because they have copyright restriction.  Some may be available at 
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  Contains Nonbinding Recommendations 
the website address, if listed.  References without asterisks are available for viewing only at the 
Dockets Management Staff.  FDA has verified the website addresses, as of the date this 
document publishes, but websites are subject to change over time. 
 
1.  Cramer, G.M., Ford, R.A., and Hall, R.L. 1978. Estimation of toxic hazard - A decision tree 
approach. Food Cosmet. Toxicol. 16:255-276. 
2.  Kokoski, C.J., Henry, S.H., Lin, C.S., and Ekelman, K.B. Methods used in safety evaluation. 
In: Food Additives, edited by Branen, A.L., Davidson, P.M., and Salminen, S., Marcel 
Dekker, Inc., New York, p. 579-616, 1990. 
3.  Lorentzen, R. 1984. FDA Procedures for carcinogenic risk assessment. Food 
Technology 38(10):108-111.* 
4.  McConnel E.E., Solleveld, H.A., Swenberg, J.A., and Boorman, G.A. 1986. Guidelines for 
combining neoplasms for evaluation of rodent carcinogenesis studies. J. Natl. Canc. 
Inst. 76:283-289. 
Document History 
  April 2002  First edition of guidance was issued. 
  October 2021  The guidance was updated to include Paperwork Reduction Act 
information and non-substantive formatting or editorial revisions. 
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