Irradiation in the Production, Processing, and Handling of Food, 48057-48073 [05-16279]

Agencies

• Food and Drug Administration
• DEPARTMENT OF HEALTH AND HUMAN SERVICES

[Federal Register Volume 70, Number 157 (Tuesday, August 16, 2005)]
[Rules and Regulations]
[Pages 48057-48073]
From the Federal Register Online via the Government Printing Office [www.gpo.gov]
[FR Doc No: 05-16279]


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DEPARTMENT OF HEALTH AND HUMAN SERVICES

Food and Drug Administration

21 CFR Part 179

[Docket No. 1999F-4372]


Irradiation in the Production, Processing, and Handling of Food

AGENCY: Food and Drug Administration, HHS.

ACTION: Final rule.

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SUMMARY: The Food and Drug Administration (FDA) is amending the food 
additive regulations to provide for the safe use of ionizing radiation 
for control of Vibrio species and other foodborne pathogens in fresh or 
frozen molluscan shellfish (e.g., oysters, mussels, clams, etc.). This 
action is in

[[Page 48058]]

response to a petition filed by the National Fisheries Institute and 
the Louisiana Department of Agriculture and Forestry.

DATES: This rule is effective August 16, 2005. Submit written or 
electronic objections and requests for a hearing by September 15, 2005. 
See section VI of this document for information on the filing of 
objections.

ADDRESSES: You may submit written or electronic objections and requests 
for a hearing identified by Docket No. 1999F-4372, by any of the 
following methods:
     Federal eRulemaking Portal: https://www.regulations.gov. 
Follow the instructions for submitting comments.
     Agency Web site: https://www.fda.gov/dockets/ecomments. 
Follow the instructions for submitting comments on the agency Web site.
     E-mail: fdadockets@oc.fda.gov. Include Docket No. 1999F-
4372 in the subject line of your e-mail message.
     FAX: 301-827-6870.
     Mail/Hand delivery/Courier [For paper, disk, or CD-ROM 
submissions]: Division of Dockets Management (HFA-305), Food and Drug 
Administration, 5630 Fishers Lane, rm. 1061, Rockville, MD 20852.
    Instructions: All submissions received must include the agency name 
and docket number for this rulemaking. All objections received will be 
posted without change to https://www.fda.gov/ohrms/dockets/default.htm, 
including any personal information provided. For detailed instructions 
on submitting objections, see the ``Objections'' heading of the 
SUPPLEMENTARY INFORMATION section of this document.
    Docket: For access to the docket to read background documents or 
comments received, go to https://www.fda.gov/ohrms/dockets/default.htm 
and insert the docket number, found in brackets in the heading of this 
document, into the ``Search'' box and follow the prompts and/or go to 
the Division of Dockets Management, 5630 Fishers Lane, rm. 1061, 
Rockville, MD 20852.

FOR FURTHER INFORMATION CONTACT: Lane A. Highbarger, Center for Food 
Safety and Applied Nutrition (HFS-255), Food and Drug Administration, 
5100 Paint Branch Pkwy., College Park, MD 20740, 301-436-1204.

SUPPLEMENTARY INFORMATION:

Table of Contents

I. Background
II. Safety Evaluation
    A. Analyses of Data by the World Health Organization
    B. Radiation Chemistry
    C. Assessment of Potential Toxicity
    D. Microbiological Profile of Molluscan Shellfish
    E. Nutritional Considerations
III. Comments
    A. Studies Reviewed in the 1999 FAO/IAEA/WHO Report on High-Dose 
Irradiation
    B. Review Article
    C. Irradiated Strawberry
    D. Reproduction Performance
    E. Mutagenicity Studies
    F. International Opinions
    G. Alkylcyclobutanones
    H. Promotion of Colon Cancer
    I. Indian National Institute of Nutrition Studies
    J. Toxicity Data
    K. Failure to Meet Statutory Requirements
    L. Trans Fatty Acids
    M. Elevated Hemoglobin
    N. Dangers of Radiation
    O. Nutritional Deficiency
IV. Conclusions
V. Environmental Impact
VI. Objections
VII. References

I. Background

    In a notice published in the Federal Register of October 19, 1999 
(64 FR 56351), FDA announced that a food additive petition (FAP 9M4682) 
had been filed by the National Fisheries Institute, 1901 North Fort 
Myer Dr., Arlington, VA 22209, and the Louisiana Department of 
Agriculture and Forestry, P.O. Box 3334, Baton Rouge, LA 70821. The 
petition proposed that the food additive regulations in part 179, 
Irradiation in the Production, Processing, and Handling of Food (21 CFR 
part 179), be amended to provide for the safe use of approved sources 
of ionizing radiation for control of Vibrio and other foodborne 
pathogens in fresh or frozen molluscan shellfish.

II. Safety Evaluation

    Under section 201(s) of the Federal Food, Drug, and Cosmetic Act 
(the act) (21 U.S.C. 321(s)), a source of radiation used to treat food 
is defined as a food additive. The additive is not added to food 
literally, but is rather a source of radiation used to process or treat 
food such that, analogous to other food processing technologies, its 
use can affect the characteristics of the food. In the subject 
petition, the intended technical effect is for control of foodborne 
pathogens, including but not limited to Vibrio bacteria, that might be 
present in fresh or frozen molluscan shellfish.
    In evaluating the safety of a source of radiation to treat food 
intended for human consumption, the agency must identify the various 
effects that may result from irradiating the food and assess whether 
any of these effects pose a public health concern. In this regard, the 
following three areas of concern need to be addressed: (1) Potential 
toxicity, (2) nutritional adequacy, and (3) potential microbiological 
risk from the treated food. Each of these areas is discussed in detail 
in this document. FDA has fully considered the data and studies 
submitted in the subject petition as well as other data and information 
relevant to safety.

A. Analyses of Data by the World Health Organization

    Based on a joint FAO/IAEA/WHO\1\ Committee's conclusion on the 
toxicological, microbiological safety and nutritional adequacy of 
irradiated foods, the Codex Alimentarius Commission (Codex) published 
its standard for irradiated foods in 1983 (revised in 2003) for 
adoption by Codex member countries (Refs. 1 and 2). This standard was 
based on the conclusion that the irradiation of any food commodity at 
an overall average dose of up to 10 kiloGray (kGy) presents no 
concerns. The newly revised standard (2003) states that the
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    \1\ FAO is the Food and Agriculture Organization of the United 
Nations; IAEA is the International Atomic Energy Agency; and WHO is 
the World Health Organization.
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    [m]inimum absorbed dose should be sufficient to achieve the 
technological purpose and the maximum absorbed dose should be less 
than that which would compromise consumer safety, wholesomeness [of 
the food] or would adversely affect structural integrity, functional 
properties, or sensory attributes. The maximum absorbed dose 
delivered to a food should not exceed 10 kGy, except when necessary 
to achieve a legitimate technological purpose.
    (Ref. 2) The original version of the standard explains in a 
footnote that ``wholesomeness [in the context of the standard] refers 
to safety for consumption of irradiated foods from the toxicological 
point of view * * * and that irradiation up to an overall average dose 
of 10 kGy introduces no special nutritional or microbiological 
problems.''
    FDA did not adopt the 1983 Codex recommendations because, at that 
time, it had not sufficiently analyzed the issues of nutritional 
adequacy and microbiological safety for all foods at all doses, nor had 
the agency pursued the analysis of toxicity beyond the examination of 
individual studies (62 FR 64107 at 64112, December 3, 1997).
    At the request of one of its member states, WHO conducted a 
subsequent review and analysis of the safety data on irradiated food 
(Ref. 3). WHO

[[Page 48059]]

considered the extent to which data on one type of food can be 
extrapolated to other foods and the extent to which individual studies 
of irradiated foods can be integrated into a single database to be 
evaluated as a whole, as opposed to separate evaluations of a series of 
individual studies (62 FR 64107 at 64112). This review included all of 
the studies in FDA's files considered to be reasonably complete by the 
agency, as well as those studies that appeared to be acceptable but had 
some deficiencies interfering with interpretation of the data (51 FR 
13376 at 13378, April 18, 1986). WHO's review also included data from 
the U.S. Department of Agriculture (USDA) and from the Federal Research 
Centre for Nutrition at Karlsruhe, Germany (62 FR 64107 at 64112). WHO 
concluded that while levels of some vitamins are decreased when food is 
irradiated at doses relevant for food irradiation, few vitamins are 
severely affected, with the exception of thiamine and vitamin E. 
However, these losses are small (on the order of 10 to 20 percent or 
less) at or below an overall average absorbed dose of 10 kGy and are 
comparable to losses seen with other forms of food processing, such as 
thermal processing and drying (Ref. 3).

B. Radiation Chemistry

    Scientists have compiled a large body of data regarding the effects 
of ionizing radiation on different foods under various conditions of 
irradiation. These data indicate that the effects of ionizing radiation 
on the characteristics of treated foods are a direct result of the 
chemical reactions induced by the absorbed radiation. The types and 
amounts of products generated by radiation-induced chemical reactions 
(``radiolysis products'') depend on both the chemical constituents of 
the food and on the specific conditions of irradiation. The principles 
of radiation chemistry also govern the extent of change, if any, in 
both the nutrient levels and the microbial load of irradiated foods. 
For a detailed discussion and evaluation of radiation chemistry, 
nutrition, toxicology, and microbiology related to irradiation of 
flesh-based foods under various conditions of use, see the agency's 
final rule permitting the irradiation of meat (62 FR 64107). In the 
current rulemaking, FDA has reviewed relevant data and information 
regarding radiation chemistry as it applies specifically to fresh or 
frozen molluscan shellfish irradiated at absorbed doses not to exceed 
5.5 kGy.
    The major components of fresh or frozen molluscan shellfish are 
water, protein, and lipid. Irradiation of water produces reactive 
hydroxyl and hydrogen radicals. These radicals can either recombine to 
form water, hydrogen gas, or hydrogen peroxide, or react with other 
components of molluscan shellfish. While the most significant effect of 
radiation-processing on the protein and lipid components of fresh or 
frozen molluscan shellfish results from the chemical reactions induced 
by hydroxyl radicals generated from the radiolysis of the water, 
radiolysis products of protein and lipid may also result from directly 
absorbed radiation. These radiolysis products, however, form in very 
small amounts and are usually the same as compounds found in foods that 
have not been irradiated (Ref. 4).
    The amounts of radiolysis products generated in a particular food 
are directly proportional to the radiation dose. Therefore, FDA can 
draw conclusions about the amounts of radiolysis products expected to 
be generated at radiation doses relevant to the subject petition by 
extrapolating from data obtained at higher doses for foods of similar 
composition irradiated under similar conditions. In general, the types 
of products generated by irradiation are similar to those products 
produced by other methods of food processing, such as canning, cooking, 
etc., because all chemical reactions caused by the addition of energy 
must follow the laws of chemistry. The radiation chemistry of food is 
also strongly influenced by the physical state of the food (solid, 
liquid, dry, or frozen) during irradiation. For example, the extent of 
chemical change that occurs in a particular food in the dry or frozen 
state will be less than the change that occurs in the same food when 
liquid water is present, all other conditions (including dose and 
ambient atmosphere) being equal, because indirect reaction products 
from water will be minimized (Ref. 5).
    During the course of reviewing chemical effects of irradiation as 
part of the evaluation of this and other petitions, FDA became aware of 
a reference that suggested that irradiating apple juice may produce 
furan (Ref. 6). Because furan has been shown to cause cancer in 
laboratory animals, FDA initiated research on whether the referenced 
report was accurate and whether furan was a common radiolysis product 
in food. FDA has confirmed that certain foods form furan in low 
quantities when irradiated and also that some foods form furan when 
heated. Studies on the irradiation of molluscan shellfish show that if 
furan is formed when molluscan shellfish are irradiated, it is formed 
at levels that are undetectable, or below the background levels of 
natural furan formation (Ref. 7). Therefore, the consumption of 
irradiated molluscan shellfish will not increase the amount of furan in 
the diet and is not an issue with this petition.
    In the Federal Registers of May 2, 1990 (55 FR 18538), and December 
3, 1997 (62 FR 64107), FDA issued final rules permitting the use of 
ionizing radiation for the control of foodborne pathogens in poultry 
and meat, respectively (referred to henceforth as the poultry and meat 
final rules). In the poultry final rule, the agency concluded that 
poultry irradiated at a dose not to exceed 3 kGy was safe. In the meat 
final rule, the agency concluded that refrigerated uncooked meat, meat 
byproducts, and meat food products, as defined in Title 9 of the Code 
of Federal Regulations (CFR), irradiated at doses up to 4.5 kGy are 
safe, and that frozen meat, meat by-products, and meat food products 
irradiated at doses up to 7.0 kGy are safe. Because meat is high in 
protein, lipid, and water, the radiation chemistry of proteins, lipids, 
and water (in both the liquid and frozen state) was extensively 
discussed in the meat final rule. The radiation chemistry of proteins 
and lipids discussed in the meat final rule is also relevant to other 
flesh foods, including foods such as poultry and fish, that may be 
referred to as ``meat'' in common usage, but that do not conform to the 
definition of meat in Title 9 of the CFR. Molluscan shellfish, 
depending on the species, differ from other flesh foods in that they 
contain between 2 and 6 percent carbohydrate, up to 20 percent protein, 
and up to 10 percent fat; the remainder is primarily water. While the 
carbohydrate level is higher than in other flesh foods, the level is 
still low.
1. Protein
    With respect to proteins, several types of reactions can occur as a 
result of irradiation. One type of reaction is the breaking of a small 
number of peptide bonds to form polypeptides of shorter length than the 
original protein. Radiation-induced aggregation or cross-linking of 
individual polypeptide chains can also occur; these processes result in 
protein denaturation. In irradiated flesh foods, most of the radiolytic 
products derived from proteins have the same chemical composition 
regardless of the protein sources, but are altered in their secondary 
and tertiary structures. These changes are similar to those that occur 
as a result of heating, but in the case of irradiation, such changes 
are far less pronounced and the amounts of reaction products generated 
are far lower (Refs. 4 and 8). Studies have established that

[[Page 48060]]

there is little change in the amino acid composition of fish irradiated 
at doses below 50 kGy (Ref. 9), which is well above the petitioned 
maximum absorbed dose for molluscan shellfish. Therefore, no 
significant change in the amino acid composition of fresh or frozen 
molluscan shellfish is expected to occur under the conditions set forth 
in this regulation.
2. Carbohydrate
    The main effects of ionizing radiation on carbohydrates in foods 
have been reviewed previously in the literature and by WHO (Refs. 5, 
10, and 11). One of the main effects of ionizing radiation is the 
abstraction of hydrogen from the carbon-hydrogen bonds of the 
carbohydrate, resulting in directly ionizing and exciting the 
carbohydrate molecule. Carbohydrate radicals may result from ionization 
of monosaccharides such as glucose or polysaccharides such as starch. 
Radiolysis products formed from starches of different origin are 
reported to be qualitatively similar (Refs. 5 and 11). In 
polysaccharides, the glycosidic linkages between constituent 
monosaccharide units may be broken, resulting in the shortening of 
polysaccharide chains and reduction in the viscosity of polysaccharides 
in solution. Starch may be degraded into dextrins, maltose, and 
glucose. Sugar acids, ketones, and other sugar monosaccharides may also 
be formed as a result of ionizing radiation. Irradiation of 
carbohydrates at doses up to 10 kGy has minimal effect on the 
carbohydrate functionality. The overall effects of ionizing radiation 
are the same as those caused by cooking and other food processing 
treatments. Carbohydrates that are present as a component of food are 
less sensitive to the effects of irradiation than pure carbohydrates 
(Ref. 5). No significant change in the carbohydrate composition of 
fresh or frozen molluscan shellfish is expected to occur under the 
conditions set forth in this regulation, i.e., a maximum absorbed dose 
of 5.5 kGy.
3. Lipid
    The meat final rule also discussed the radiation chemistry of 
lipids (predominantly triglycerides in meat). A variety of radiolysis 
products derived from lipids have been identified, including fatty 
acids, esters, aldehydes, ketones, alkanes, alkenes, and other 
hydrocarbons (Refs. 12 and 13). Identical or analogous compounds, 
however, are also found in foods that have not been irradiated. In 
particular, heating food produces the same types of compounds, but in 
amounts far greater than the trace amounts produced from irradiating 
food (Refs. 4 and 14). In addition, alkylcyclobutanones (ACBs), which 
are formed in small quantities when fats are exposed to ionizing 
radiation, have been identified in meat and poultry. The specific ACBs 
formed will depend on the fatty acid composition of the food. For 
example, 2-dodecylcyclobutanone (2-DCB) has been reported to be formed 
from palmitic acid in amounts from 0.3 to 0.6 microgram per gram lipid 
per kGy (microg/g lipid/kGy) from irradiated chicken (Ref. 15). Other 
researchers have found that (2--DCB) is formed at significantly lower 
rates, 0.04 microg/g lipid/kGy from ground beef (Ref. 16). For 
comparison, ground beef tallow contains approximately 25 percent 
palmitic acid and chicken fat contains approximately 22 percent 
palmitic acid.
    One major difference between fish (including shellfish and finfish) 
and other flesh foods is the predominance of polyunsaturated fatty 
acids (PUFAs) in the lipid phase of fish. PUFAs are a subclass of 
lipids that have a higher degree of unsaturation in the hydrocarbon 
chain than the saturated (e.g., stearic acid) or monounsaturated (e.g., 
oleic acid) fatty acids. Due to the higher level of unsaturation, PUFAs 
are generally more readily oxidized than saturated fatty acids. 
Therefore, PUFAs could be more radiation-sensitive than other lipid 
components, as observed in some studies of irradiated oil. However, 
evidence from meat studies suggests that the protein component of meat 
may protect lipids from oxidative damage (Ref. 5). Because the lipid 
fraction of meat consists primarily of saturated and monounsaturated 
fatty acids with negligible quantities of PUFAs, FDA did not explicitly 
address the radiation chemistry of PUFAs in its previous reviews.
    The effects of irradiation on PUFAs in fish have been described in 
several studies reviewed by FDA. Adams et al. studied the effects of 
radiation on the concentration of PUFAs in herring and showed that 
irradiation of herring fillets at sterilizing doses (50 kGy), well 
above the petitioned maximum dose for molluscan shellfish, had no 
effect on the concentration of PUFAs (Ref. 17). Similarly, Armstrong et 
al. conducted research on the effects of radiation on fatty acid 
composition in fish and concluded that no significant changes occurred 
in the fatty acid profiles upon irradiation at 1, 2, or 6 kGy (Ref. 
18). The authors also concluded that variations in fatty acid 
composition between individual samples were greater than any radiation-
induced changes.
    Sant'ana and Mancini-Filho studied the effects of radiation on the 
distribution of fatty acids in fish (Ref. 19). They studied two 
monounsaturated fatty acids and seven PUFAs (including three different 
omega-3 fatty acids) before and after irradiation at doses up to 3 kGy. 
The authors observed insignificant changes in the concentration of 
total monounsaturated fatty acids and an approximately 13 percent 
decrease in total PUFAs at the highest dose, largely attributable to a 
loss of the long chain PUFAs, including docosahexaenoic acid. The 
overall change for essential fatty acids (e.g., linoleic and linolenic 
acids) was minimal (less than 3 percent). The authors also observed an 
increase in lipid oxidation based on levels of thiobarbituric acid 
reactive substances, but noted that antioxidants such as tocopherol 
protect against lipid oxidation (Ref. 4).
    In addition, a study summarized in an International Consultative 
Group on Food Irradiation monograph compared the fatty acid composition 
of unirradiated and irradiated herring oil (Ref. 20). The profile for 
12 fatty acids was compared to controls 1 day and 28 days after 
irradiation. Only two fatty acids appeared to have decreased by day 28 
following irradiation at 50 kGy (Ref. 4).
    Research conducted by FDA on various species of seafood also 
demonstrated that the concentrations of PUFAs are not significantly 
affected by irradiation (Refs. 21 and 22). Therefore, based on the 
totality of evidence, the agency concludes that no significant loss of 
PUFAs is expected to occur in the diet under the conditions of 
irradiation set forth in this regulation. In summary, FDA's review of 
the radiation chemistry of proteins and lipids in the subject petition 
raises no issues that have not been considered previously in the meat 
and poultry final rules (Ref. 4).

C. Assessment of Potential Toxicity

    In the safety evaluation of irradiated meat and poultry, the agency 
examined all of the available data from toxicological studies relevant 
to the safety of irradiated flesh-based foods, including studies on 
fish high in PUFAs. These included 24 long-term feeding studies, 10 
reproduction/teratology studies, and 15 genotoxicity studies with 
flesh-based foods irradiated at doses from 6 to 74 kGy. No 
toxicologically significant adverse effects attributable to irradiated 
flesh foods were observed in any of the studies (62 FR 64107 at 64112 
and 64114).

[[Page 48061]]

    The proposed maximum absorbed dose of 5.5 kGy for fresh and frozen 
molluscan shellfish in the subject petition is somewhat higher than the 
currently permitted maximum dose for the irradiation of non-frozen 
meat. However, FDA previously evaluated the long-term toxicological 
studies of flesh foods fed at a range that includes absorbed doses that 
are either similar to or considerably higher than the absorbed dose 
requested in this petition. In addition, the absorbed dose exceeded 50 
kGy in many studies with no adverse effects reported. Therefore, these 
data demonstrate that molluscan shellfish irradiated at levels up to 
the dose proposed in this petition will not present a toxicological 
hazard (Ref. 8).
    In summary, FDA has reviewed a large body of data relevant to the 
assessment of potential toxicity of irradiated foods. While all of the 
studies are not of equal quality or rigor, the agency concludes that 
the quantity and breadth of testing and the number and significance of 
endpoints assessed would have identified any real or meaningful risk. 
The overwhelming majority of studies showed no evidence of toxicity. On 
those few occasions when adverse effects have been reported, FDA finds 
that those effects have not been consistently produced in related 
studies conducted at a higher dose or longer duration, as would be 
expected if the effects were attributable to irradiation (62 FR 64107 
at 64112 and 64114). Therefore, based on the totality of evidence, FDA 
concludes that irradiation of fresh and frozen molluscan shellfish 
under the conditions proposed in this petition does not present a 
toxicological hazard.

D. Microbiological Profile of Molluscan Shellfish

    Vibrio bacteria predominate in estuarine environments, and 
consequently, are naturally present in most finfish and shellfish (Ref. 
23). Most cases of reported diseases attributed to Vibrio species are 
associated with consumption of raw molluscan shellfish, particularly 
raw oysters. Although Vibrio species from shellfish infect relatively 
few individuals, they can cause severe illness, including mortality. Of 
the 12 Vibrio species known to cause human infections, 8 have been 
associated with consumption of food. V. parahaemolyticus and V. 
vulnificus are most commonly isolated from oysters. V. vulnificus is 
associated with 95 percent of all seafood-related deaths in the United 
States (Ref. 24).
    In general, the subject petition relies on published or other 
publicly available information or material from previous food additive 
petitions to address microbiological issues. The petitioner has 
documented that Vibrio species in uncooked molluscan shellfish provide 
a significant public health risk. Vibrio bacteria are highly sensitive 
to ionizing radiation and are usually eliminated by doses as low as 0.5 
kGy. Published D10 values\2\ for V. parahaemolyticus and 
other Vibrio species range from 0.02 to 0.4 kGy (Ref. 25).
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    \2\ D10 is the absorbed dose of radiation required to 
reduce a bacterial population by 90 percent.
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    Control of contaminating Salmonella or Listeria generally requires 
higher doses than for Vibrio species, because the D10 values 
are higher, about 0.5 to 1.0 kGy and 0.4 to 0.6 kGy, respectively (Ref. 
26). Several publications referenced in the subject petition state that 
these three genera can be eliminated by doses well under 10 kGy. 
Numerous studies demonstrate that a dose of 5 kGy will reduce a 
population of Salmonella serotypes, Staphylococcus aureus, Shigella, 
and Vibrio by at least six log cycles. Other studies report 5-log 
reductions for Listeria and Salmonella at 2.3 kGy and 2.8 kGy. In 
addition, D10 values for irradiation cited in published 
literature for several Salmonella serotypes in various fresh foods 
ranged from 0.2 to 0.9 kGy. Therefore, irradiation at doses up to the 
dose limit in the regulation could significantly reduce the populations 
of these organisms (Ref. 25).
    Clostridium botulinum (C. botulinum) type E can sometimes be found 
in seafood. Because this organism is relatively resistant to radiation, 
as compared to non-spore forming bacteria, the petitioner provided data 
regarding the likelihood that C. botulinum would grow and produce toxin 
in irradiated molluscan shellfish. Included in the petition's 
references is an in-depth discussion of the likelihood for outgrowth 
and toxin production by C. botulinum type E in fish (Ref. 27). The 
author cites studies conducted in his laboratory on the effect of 
storage temperature and irradiation on toxin production by C. botulinum 
type E in fish. In these studies, no toxin was detected after 
incubation with fish of up to 10\5\ organisms at 0 degrees Celsius for 
8 weeks, well beyond the shelf life of these products. At 5 degrees 
Celsius, no toxin was produced for up to 6 weeks of storage in 
inoculated fish that had not been irradiated or for up to 7 weeks when 
irradiated at 2 kGy. Thus, it took longer for toxin to be produced in 
the irradiated fish than in fish that were not irradiated. 
Additionally, the time required for toxin production, 7 weeks, is far 
beyond the shelf life of fresh seafood. Therefore, irradiation would 
not increase the risk from botulinum toxin.
    Current Hazard Assessment and Critical Control Point plans in 
effect for molluscan shellfish require storage under proper conditions, 
including maintenance at controlled temperatures. Therefore, 
irradiation can serve as an effective method for the primary intended 
use of eliminating populations of Vibrio species and other pathogens in 
molluscan shellfish without adding a significant risk from the growth 
of and toxin production by C. botulinum type E (Ref. 25).
    The subject petition includes data and information that support the 
effectiveness of the proposed irradiation of fresh and frozen molluscan 
shellfish at a maximum absorbed dose of 5.5 kGy to control Vibrio 
species and other foodborne pathogens. While the data show that 
irradiation is effective in reducing the levels of Vibrio species and 
other bacteria in fresh and frozen molluscan shellfish, the data also 
show that irradiation will not increase the risk of toxin production 
from germinated spores of C. botulinum type E.
    Based on the available data and information, FDA concludes that 
irradiation of fresh or frozen molluscan shellfish conducted in 
accordance with current good manufacturing practices will reduce or 
eliminate bacterial populations with no increased microbial risk from 
pathogens that may survive the irradiation process.

E. Nutritional Considerations

    Lipids are a component of molluscan shellfish contributing 
approximately 20 to 30 percent to the caloric value of molluscan 
shellfish. PUFAs are a significant source of omega-3 and omega-6 fatty 
acids and are therefore nutritionally important components of the fat 
of molluscan shellfish. As noted in section II.A of this document, PUFA 
levels were not reduced significantly by ionizing radiation. 
Additionally, the amount of omega-3 and omega-6 PUFAs can vary widely 
within a single species and between species of molluscan shellfish. The 
omega-3 fatty acid content among most species varies within a factor of 
2, and the total PUFA content can vary by more than a factor of 10 
(omega-3 and omega-6 PUFAs) within an individual species. Furthermore, 
molluscan shellfish are only one of several fish sources of long chain 
PUFAs. Because of the variety of seafood sources of long chain PUFAs, 
the variation of fatty acid content in molluscan shellfish, and the 
observed insensitivity of PUFAs to irradiation, FDA concludes that 
irradiation of fresh

[[Page 48062]]

and frozen molluscan shellfish under the conditions proposed will not 
adversely affect the nutritional adequacy of the diet with respect to 
PUFAs (Ref. 8).
    Molluscan shellfish contain several B-vitamins including thiamine, 
niacin, vitamin B6, and vitamin B12.\3\ Individual food intake data is 
available from nationwide surveys conducted by the USDA. These surveys 
were designed to monitor the types and amounts of foods eaten by 
Americans and food consumption patterns in the U.S. population. FDA 
routinely uses these data to estimate exposure to various foods, food 
ingredients, and food contaminants. The relative contribution of the 
food category ``shellfish and fish (excluding canned tuna)'' is less 
than 3 percent of the dietary intake for thiamine, niacin, and vitamin 
B6 (Ref. 28). Fish and shellfish are, however, significant contributors 
to vitamin B12 intake among U.S. adults, contributing to approximately 
20 percent of the total vitamin B12 intake.
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    \3\ Dietary sources of nutrients have been evaluated using the 
1994/1996 Continuing Survey of Food Intakes by Individuals database.
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    Irradiation of any food, regardless of the dose, has no effect on 
the levels of minerals that are present in trace amounts (Ref. 5). 
Levels of certain vitamins, on the other hand, may be reduced as a 
result of irradiation. The extent to which this reduction occurs 
depends on the specific vitamin, the type of food, and the conditions 
of irradiation. Not all vitamin loss is nutritionally significant, 
however, and the extent to which a reduction in a specific vitamin 
level is significant depends on the relative contribution of the food 
in question to the total dietary intake of the vitamin. While thiamine 
is among the most radiation sensitive, the more nutritionally 
significant vitamin in fish and shellfish, vitamin B12, is extremely 
resistant to radiation.
    Based on the available data and information, FDA concludes that 
irradiation of fresh or frozen molluscan shellfish under the conditions 
set forth in the regulation in this document will have no adverse 
impact on the nutritional adequacy of the diet.

III. Comments

    FDA has received numerous letters, primarily form letters, from 
individuals that state their opinions regarding the potential dangers 
and unacceptability of irradiating food. None of these letters contain 
any substantive information that can be used in a safety evaluation of 
irradiated molluscan shellfish.
    Additionally, FDA received several comments from Public Citizen 
(PC) and the Center for Food Safety (CFS) requesting the denial of this 
and other food irradiation petitions. The comments were largely of a 
general nature and not necessarily specific to the petitioned requests. 
Some of the comments specifically questioned a report of a Joint FAO/
IAEA/WHO Study Group on the wholesomeness of foods irradiated with 
doses above 10 kGy. Because the comments were addressed to the Docket 
for this rulemaking, the comments and FDA's response are discussed as 
follows:

A. Studies Reviewed in the 1999 FAO/IAEA/WHO Report on High-Dose 
Irradiation

     (1) One comment states that the petition should be denied because 
there are four positive studies mentioned but mischaracterized in the 
1999 FAO/IAEA/WHO report on high-dose irradiation. The comment states:
    The 1999 FAO/IAEA/WHO report is the most detailed recent review 
of food irradiation safety. CFS [Center for Food Safety] anticipates 
that FDA will seek to rely on it. It is critical that FDA understand 
the defects in that report before making a determination on the 
above-referenced additive petition...the four studies were 
incorrectly classified as ``negative for high-dose irradiation 
effect, possible effect of nutrition or diet.''* * *
    The 1999 FAO/IAEA/WHO report acknowledged the Anderson et al. 
study (on laboratory animal diets) showed ``evidence of weakly 
mutagenic effect'' with one diet that was irradiated, yet it 
classified the study as ``negative for high-dose irradiation effect, 
possible effect of nutrition or diet'' (p. 117). However, no 
indication exists that the irradiated standard PRD laboratory diet 
that produced the mutagenic effect was otherwise deficient. Further, 
the unirradiated control PRD diet did not produce the mutagenic 
effect. Anderson et al. found irradiation of the diet produced the 
effect. The 1999 FAO/IAEA/WHO report's classification of the study 
as ``negative'' was unfounded. (Emphasis in original.)
    In the study performed by Anderson et al. (1981) mice were fed four 
laboratory diets irradiated at 10 kGy, 25 kGy, and 50 kGy (Ref. 29). 
Mice were also fed unirradiated diets as a negative control. 
Additionally, mice were injected intraperitoneally with a known 
mutagen, cyclophosphamide, at 200 mg per kg of body weight (mg/kg body 
weight) as a positive control. The study report stated that mice 
consuming one diet (PRD diet)\4\ irradiated at 50 kGy resulted in a 
slight increase in post-implantation deaths over the unirradiated diet 
when compared to the positive control. The other three irradiated diets 
showed no significant increases in early post-implantation death. The 
comment provides no information to explain why the Anderson et al. 
study on radiation-sterilized laboratory diets should be considered 
relevant to the conditions proposed in this petition for the 
irradiation of molluscan shellfish to a maximum absorbed dose that will 
not exceed 5.5 kGy. Moreover, the comment provides no analysis of the 
study and no information to demonstrate that the ``weakly mutagenic 
effect'' associated with the laboratory diet irradiated at 50 kGy is 
attributable to irradiation of the diet.
---------------------------------------------------------------------------

    \4\ The PRD diet is a formulation of 5.125 g/100 g Barley, 10.0 
g/100 g maize meal, 18.125 g/100 g oats (Sussex Ground), 20.0 g/100 
g wheat, 20.0 g/100 g wheat feed, 5.0 g/100 g white fish meal (crude 
protein 66 percent), 2.5 g/100 g yeast, 10.0 g/100 g soya extract, 
7.5 g/100 g dry skimmed milk (crude protein 33), 0.75 g/100 g salt 
(NaCl), and a 1.0 percent vitamin mineral supplement.
---------------------------------------------------------------------------

    (2) The comment states that ``[a] thorough discussion of the 
Bugyaki et al. study in a 1970 FAO/IAEA/WHO Expert Committee report 
highlighted it as a significant positive finding.'' The comment goes on 
to state:
    The 1999 FAO/IAEA/WHO report admitted that Bugyaki et al. showed 
``chromosomal abnormalities in germ cells due to formation of 
peroxides and radicals,'' but - without explanation - classified the 
study as ``negative for high-dose irradiation effect, possible 
effect of nutrition or diet'' (p. 118). That is plain inconsistency; 
the `peroxides and radicals' resulted from the irradiation (see 
Bugyaki et al., at p. 118: ``... some of the changes produced by 
radiation -- the free radicals for example -- will disappear with 
time.'' [translated from French]). Further, the same Expert 
Committee agreed 29 years earlier that Bugyaki et al. demonstrated 
``certain disturbing effects'' of high dose irradiation. That 
Committee did not discount the effects as artifacts of nutrition or 
diet, as the 1999 Committee did. The 1999 FAO/IAEA/WHO report's 
classification of this study as `negative' again lacks a rational 
foundation. (Emphasis in original.)
    In Bugyaki et al., a 1968 report on irradiated wheat, mice were fed 
a diet containing 50 percent freshly irradiated wheat meal (50 kGy); 
the balance was basic food powder (the basic food powder was described 
by the author to contain 55 percent vegetable matter, 35 percent animal 
matter, and 10 percent complementary nutrients) (Ref. 30). Control 
animals were fed a diet containing 50 percent wheat that had not been 
irradiated with the balance being the basic food powder. Because the 
authors were concerned that compression into pellets may affect the 
irradiated foods, the animals were fed the food in powder form. The 
authors note that there were readily observable

[[Page 48063]]

physical and chemical changes in the wheat meal irradiated at 50 kGy.
    The authors state that both the treated and untreated animals 
developed tumors. However, the tumors found in the treated animals were 
different than the tumors found in the untreated animals. The authors 
note that the treated animals had a slight increase in anatomic-
pathological lesions; however, they go on to state that there was no 
well defined damage. Additionally, they state that there were 
alterations in the meiotic chromosomes of the treated animals. The 
authors conclude that animals consuming a large part of their diet 
irradiated at doses as high as 50 kGy may deserve special attention.
    The comment provides no information to demonstrate why the Bugyaki 
et al study on freshly irradiated wheat at 50 kGy is relevant to the 
conditions proposed in this petition for the irradiation of molluscan 
shellfish to a maximum absorbed dose that will not exceed 5.5 kGy. 
Foods irradiated at such a high dose often require careful control of 
temperature and atmosphere to prevent compositional changes that would 
make them unsuitable for food use. The agency notes that several long 
term feeding studies using foods irradiated under appropriate 
conditions at doses greater than 50 kGy demonstrated no toxicological 
effects that could be attributed to the irradiated foods.
    (3) The comment states:
    The 1999 FAO/IAEA/WHO report states the study performed by 
Moutschen-Dahmen et al. showed ``increased pre-implantation 
embryonic deaths; not confirmed by cytological analysis'' and 
classified the study as ``negative for high-dose irradiation effect, 
possible effect of nutrition or diet'' (p. 115). The suggestion of 
an effect of nutrition or diet is unsupported. (Emphasis in 
original.)
    The agency has previously addressed the study by Moutschen-Dahmen 
et al. (51 FR 13376 at 13387) and noted:
    There was no increase in post-implantation losses. Post-
implantation losses, determined by counting dead embryos, are 
believed to be the most reliable and sensitive indicator of dominant 
lethality. The authors found only pre-implantation losses, which are 
much less sensitive than post-implantation losses and merely a 
measure of total implants dead or alive subtracted from the total 
number. In addition to the possibility that results of the study 
could be spurious, any number of factors other than dominant 
lethality may cause pre-implantation losses, such as a decrease in 
the number of eggs ovulated.
    If these effects were real, one would expect to see some effect 
on post implantation losses at a lower dose because post-
implantation losses are a much more sensitive indicator than pre-
implantation losses, as mentioned previously.
    The agency concluded:
    Although the findings reported may be statistically significant, 
the authors were uncertain as to what to attribute these results. 
They concluded that the most probable mechanism by which these 
effects could be produced would be via chromosomal aberration. The 
studies necessary to establish an association between these effects 
and chromosomal aberrations were not conducted. Additional treatment 
levels below that conducted as mentioned previously to detect post-
implantation losses or examinations of the 24 to 48 hour fertilized 
eggs could have proved better evidence of causality, but these 
studies were not conducted. Thus, although pre-implantation losses 
were observed, FDA concludes that there is no biological 
significance to this observation because it was not reproducible.
    The comment provides no information to demonstrate why the 
Moutschen-Dahmen et al. (Ref. 31) study (1970) in which mice were fed a 
laboratory chow diet, of which 50 percent was irradiated at 50 kGy is 
relevant to the conditions proposed in this petition for the 
irradiation of molluscan shellfish to a maximum absorbed dose that will 
not exceed 5.5 kGy. The study was designed to look for mutations that 
would be lethal to the animals. Further, the comment provides no 
information to demonstrate that the pre-implantation deaths were caused 
by dominant lethal mutations that were induced by the consumption of 
irradiated food. Finally, the comment provides no evidence to refute 
the agency's previous conclusion.
    (4) With regard to another study (Ref. 32), the comment states 
that:
    The 1999 FAO/IAEA/WHO report admits the study showed 
``significant increase in the mutation frequency induced by the high 
dose irradiated foods,'' but nevertheless classified the study as 
``negative for high-dose irradiation effect, possible effect of 
nutrition or diet'' (p. 115). This is patently contradictory; the 
`negative' classification again lacks explanation. (Emphasis in 
original.)
    In the study performed by Johnston-Arthur et al. (1975), Swiss 
albino mice were starved for 36 hours and then fed normal and 
irradiated ( 7.5 kGy, 15 kGy, and 30 kGy) laboratory chow for 7 hours 
(Ref. 32). The mice were then injected intraperitoneally with 
Salmonella typhimurium TA 1530 and the bacteria were incubated in the 
mice for 3 hours. The mice were then sacrificed and the bacteria were 
harvested and tested using the host-mediated assay test for 
mutagenicity. The results indicated a significant increase in the 
mutation frequency in the bacteria that were exposed to the 30 kGy-
sterilized food. No significant differences were observed in the 
bacteria that were harvested from the mice fed the 7.5 kGy and 15 kGy 
diet when compared with the control.
    The comment provides no information to demonstrate why the 
Johnston-Arthur et al. study on the irradiation sterilization of lab 
chow at 30 kGy is relevant to the irradiation of molluscan shellfish to 
a maximum absorbed dose that will not exceed 5.5 kGy. Moreover, 
mutation studies with S. typhimurium are intended to screen for 
possible mutations affecting animals that can be tested in long term 
animal studies. However, several properly conducted long term feeding 
studies performed on animals fed with foods irradiated at higher doses 
(up to 56 kGy) have shown no mutagenic effects to the subject animals.
    Finally, the agency notes that the subject of this regulation is 
the petition (FAP 9M4682) regarding shellfish and not the 1999 FAO/
IAEA/WHO report on high-dose irradiation. In its review of the 
published literature on the safety of irradiated foods, the agency 
finds that properly conducted animal feeding studies showed no evidence 
of toxicity attributable to irradiated food. On the few occasions when 
studies reported adverse effects, the effects were not consistently 
reproduced in related studies conducted with similar foods irradiated 
to doses equal to or higher than those for which the adverse effects 
were reported, as would be expected if the reported effect were a toxic 
effect caused by a radiolysis product (62 FR 64107 at 64112 and 64114).

B. Review Article

    One comment submitted a paper (Kevesan and Swaminathan, 1971) that 
reviewed studies performed in the 1950s and 1960s on irradiated 
substrates and irradiated foods (Ref. 33). The comment states that 
numerous studies from the 1950s and 1960s found a variety of toxic 
effects in animal feeding and in vitro studies, which on the whole cast 
doubt on the safety of the technology. The comment asks FDA to ``take a 
closer look at the host of past positive studies cited therein.''
    The comment further states:
    [A]ttempts to discount all of the past positive findings as 
aberrations, products of chance, or artifacts of diet will no longer 
suffice. These studies need further FDA review particularly in view 
of the 2003 Codex Alimentarius standard revision that allowed for 
higher absorbed doses of radiation than previously permitted.
    The agency notes that the subject of FAP 9M4682 is the irradiation 
of molluscan shellfish to a maximum absorbed does of 5.5 kGy, not the 
recently revised Codex standard. Furthermore, the authors of the paper 
referenced by the comment do not come to the conclusion that the 
comment implies. Rather, the study's authors

[[Page 48064]]

(Kevesan and Swaminathan) conclude that ``major deficiencies in the way 
some of the experiments have been designed and conducted coupled with 
inadequacy of genetic data urgently necessitates further investigations 
before concluding that the irradiated food materials `can be consumed 
with impunity'.''
    FDA agrees with the conclusions of the review article in the 
context of studies performed prior to 1970. However, many properly 
conducted studies have been performed after this review was written. As 
previously noted in this document, the agency finds that properly 
conducted animal feeding studies showed no evidence of toxicity 
attributable to irradiated food. On the few occasions when studies 
reported adverse effects, the effects were not consistently reproduced 
in related studies conducted with similar foods irradiated to doses 
equal to or higher than those for which the adverse effects were 
reported, as would be expected if the reported effect were a toxic 
effect caused by a radiolysis product (62 FR 64107 at 64112 and 64114). 
The comment provides no additional information that would cause the 
agency to change its conclusion on the safety of irradiated food.

C. Irradiated Strawberry

    One comment submitted a paper (Verschuuren, Esch, and Kooy, 1971) 
describing the effects of feeding rats irradiated strawberry-powder and 
irradiated strawberry-juice (Ref 34). The comment states that rats fed 
``irradiated strawberry powder supplement showed a statistically 
significant growth deficit compared to the control animals fed the same 
diet, including the powder supplement, but which was unirradiated.'' 
The comment goes on to state:
    FDA's internal reviewers in 1981 and 1982 (reviews are attached 
to study) twice classified the Verschurren (sic) et al. study as one 
the agency should ``accept'' without reservations, only to be later 
overridden by a third reviewer who was able to reclassify the study 
as ``reject.'' This change was based on the third reviewer's 
suggestion that the study was hampered by ``inadequate diet and 
restricted food intake,'' a surprising suggestion as nothing in the 
study supported that conclusion
    The comment misrepresents the conclusion of one of the reviewers 
who did the initial review of the study. Initially, the study was 
accepted by two reviewers. However, upon further review by one of the 
initial reviewers and a third reviewer, this paper was rejected in the 
secondary review because of inadequate diet and restricted food intake. 
The comment provides no information that would alter the agency's 
conclusion that some of the diets were incomplete and restricted. 
Moreover, the comment provides no information that explains why the 
consumption of irradiated strawberry-powder is relevant to the 
consumption of irradiated molluscan shellfish with a maximum absorbed 
dose of 5.5 kGy.

D. Reproduction Performance

    One comment states that a study conducted at Columbia University in 
1954 ``supports other studies that yielded adverse health effects, 
which our organizations have previously submitted to this docket.''
    The comment submitted part of a report, ``Termination Report--Part 
1, Food Irradiation and Associated Studies, September 15, 1954,'' which 
was conducted at Columbia University for the U.S. Atomic Energy 
Commission. The report compares the fertility of ``Professor Sherman's 
high generation rats'' that were fed either ``Sherman diet 16'' or a 
``modified Sherman diet''\5\ (milk powder was replaced by skim milk 
powder and irradiated butterfat). The report concluded that there was a 
significant decrease in the fertility of the rats fed the irradiated 
diet. The report also mentions that there is significant vitamin E 
destruction; however, the comment did not include the entire results 
and discussion section with the authors' discussion.
---------------------------------------------------------------------------

    \5\ The control diet was ``Sherman diet 16,'' consisting of 1000 
g ground whole wheat, 200 g whole milk powder, and 20 g salt. The 
``irradiated diet'' consisted of 1000 g ground whole wheat, 147 g 
skim milk powder, 53 g irradiated butterfat, and 20 g salt.
---------------------------------------------------------------------------

    FDA reviewers have previously reviewed a subsequent publication of 
a report of this study (Ref. 35). At the time of the study, it was not 
well recognized that irradiation of fat in the presence of air can 
stimulate oxidation leading to rancidity and high levels of peroxides. 
Such rancidity can lead to nutritional deficiencies due to the animals 
reducing their food consumption and destruction of vitamins. FDA 
reviewers concluded that it appears that littermates were mated and 
that the females were mated almost continually, allowing little time 
for rest between litters. If there was a nutritional or oil 
peroxidation and palatability problems with the diet, it would be 
exacerbated by the continuous breeding of the females. Considering the 
report's mention of considerable vitamin E destruction, the effects 
seen appear to be the result of a nutritionally inadequate diet, not 
toxicity, and would not be relevant to irradiation of molluscan 
shellfish.

E. Mutagenicity Studies

    One comment states that the petition should be denied because the 
number of positive mutagenicity studies (including those discussed 
previously that were identified by the comment as mischaracterized or 
ignored) compares favorably with the number of negative studies. The 
comment states that ``[m]ore than one-third of both in vivo and in 
vitro studies are positive'' for mutagenicity, suggesting there is 
``bias in the official posture in support of the safety of 
irradiation.''
    The suggestion of the comment that FDA showed a ``bias in the 
official posture'' on the safety of the consumption of irradiated food 
is not supported by any substantive information.
    The Bureau of Foods Irradiated Foods Committee (BFIFC) recommended 
that foods irradiated at a dose above 1 kGy be evaluated using a 
battery of mutagenicity tests to assess whether long-term feeding 
studies in animals were necessary (Ref. 36). Mutagenicity studies are 
primarily used to screen for potential mutagenic effects. Animal 
feeding studies are more reliable for determining the true mutagenic 
potential of a compound that is consumed in food (Ref 37). Moreover, 
one cannot draw valid conclusions from data simply by summing positive 
and negative results without fully evaluating the individual studies 
and assessing what conclusions such studies support and considering the 
totality of evidence. If the occasional report of a mutagenic effect 
were valid and significant to health, one should have seen consistent 
adverse toxicological effects in the many long term and reproduction 
studies with animals. This has not been the case.

 F. International Opinions

    The comment states that the petition should be denied because ``[a] 
majority of Parliamentary Members voted for a provision that the EU's 
list of foods authorised (sic) for irradiation should not be 
expanded,'' and ``[a] working group of the Codex Alimentarius 
Commission's Contaminants and Food Additives Committee in November, 
2002, recommended against approval of a Codex proposal to remove the 
present 10 kiloGray radiation dose cap, which would allow any foods to 
be irradiated at any dose -- regardless of how high. (Emphasis in 
original.)''
    The agency notes that the subject of this regulation is the 
petition (FAP 9M4682) to permit irradiating shellfish at a dose up to 
5.5 kGy, not whether the maximum dose in the Codex General Standard for 
Irradiated Foods should be

[[Page 48065]]

raised above 10 kGy. The act requires FDA to issue a regulation 
authorizing safe use of an additive when safety has been demonstrated 
under the proposed conditions of use. FDA notes that the Codex General 
Standard for Irradiated Foods has recently been revised (Codex 2003) by 
supplanting reference to a maximum overall average dose of 10 kGy with 
the statement that ``[t]he maximum absorbed dose delivered to a food 
should not exceed 10 kGy, except when necessary to achieve a legitimate 
technological purpose.'' (Ref. 2). The comment fails to demonstrate why 
the debate within Codex leading up to this change is relevant to the 
conditions proposed in this petition for the irradiation of molluscan 
shellfish to a maximum absorbed dose that will not exceed 5.5 kGy.
    One comment states that the petition should be denied because of a 
report published by the Organisation for Economic Co-Operation and 
Development (OECD) which states:
    Hitherto available data indicate, however, that increased rates 
of mutation and chromosomal aberration will probably be induced in 
certain cases. Although experiments indicate that the genetical 
(sic) effect, in cases where it is induced, is relatively small 
compared to the effect of direct exposure of animals to radiation, 
the same experiments indicate that the possible effect will not be 
negligible.
    The comment goes on to state that ``[r]ather than being refuted by 
subsequent evidence, the OECD's statement regarding likely induction of 
mutations and chromosomal aberration has been confirmed in many 
studies, cited in this and our earlier comments.''
    The 1965 OECD report, entitled ``Steering Committee for Nuclear 
Energy Study Group on Food Irradiation,'' reflects scientific 
understanding at the time it was written (Ref. 38). The document is a 
compendium of published and unpublished (at the time) reports on the 
effect of irradiated substances on a variety of organisms. The report 
concluded that ``it is impossible to arrive at any definite conclusion 
as to the presence or absence of genetic effects if irradiated food 
were used for human consumption or for animal feeding.'' Furthermore, 
the report states that more rigorous studies should be performed and 
when contradictory results are found, the reasons should be determined. 
Since the report was compiled in 1965 numerous studies have been 
performed on the effects of consuming irradiated foods in multiple 
animal species and in humans. Starting in the 1980's, FDA has reviewed 
these and other studies, and while many of these studies cannot 
individually establish safety, they still provided important 
information that, when evaluated collectively, supports a conclusion 
that there is no reason to believe that irradiation of flesh foods 
presents a toxicological hazard. The comment provides no evidence to 
refute the agency's conclusion.

G. Alkylcyclobutanones

    One comment states that ``certain chemical by-products formed in 
food that has been irradiated, known as cyclobutanones, could be toxic 
enough to cause significant DNA damage, potentially leading to 
carcinogenic and mutagenic effects.'' In addition, the comment states 
that ``[t]wo major international food safety groups -- CCFAC (Codex 
Committee on Food Additives and Contaminants), and SCF (The Scientific 
Committee on Food of the European Commission) -- deemed the indications 
of toxicity strong enough to necessitate considerable additional 
study.''
    2-ACBs have been reported as radiolysis products of fats (Refs. 39a 
and 39b). Studies performed by researchers have reported that certain 
alkylcyclobutanones can cause single strand DNA breaks detectable by 
the COMET\6\ assay (Ref. 40). Several animal feeding studies have been 
conducted with fat-containing foods irradiated at doses far higher than 
would be used on molluscan shellfish. If 2-ACBs, at the level present 
in irradiated foods, were of sufficient toxicity to cause significant 
DNA damage, one would expect to have seen adverse effects in those 
studies where animals were fed meat as a substantial part of their 
diet. Moreover, the COMET assay has not yet reached the level of 
reliability and reproducibility that is needed to be considered a 
standard procedure for testing potential genotoxins. At present, the 
assay is of value primarily in basic research of cellular response to 
DNA damage and repair, in both in vitro and in vivo systems (Ref. 41).
---------------------------------------------------------------------------

    \6\ Single cell gel electrophoresis or `Comet assay' is a rapid 
and very sensitive fluorescent microscopic method to examine DNA 
damage and repair at individual cell level.
---------------------------------------------------------------------------

    Also, contrary to what is implied by the comment, the Scientific 
Committee on Foods of the European Commission concluded, in July 2002, 
``[a]s the adverse effects noted refer almost entirely to in vitro 
studies, it is not appropriate, on the bases of these results, to make 
a risk assessment for human health associated with the consumption of 
2-ACBs present in irradiated fat-containing foods.'' The genotoxicity 
of 2-ACBs has not been established by the standard genotoxicity assays 
nor are there any adequate animal feeding studies in existence to 
determine no-observed-adverse-effect levels (NOAELs) for various 
alkylcyclobutanones. Reassurance as to the safety of irradiated fat-
containing food can be based on the large number of feeding studies 
carried out with irradiated foods which formed the basis for the 
wholesomeness assessments of irradiated foods published by FAO/IAEA/
WHO.
    Moreover, researchers have recently demonstrated that 2-DCB does 
not induce mutations in the Salmonella mutagenicity test or 
intrachromosomal recombination in Saccharomyces cerevisiae or the 
Escherichia coli tryptophan reverse mutation assay (Refs. 42 and 43). A 
further study, published in 2004, has demonstrated that the Ames assay 
showed no difference between 5 concentrations of 2-DCB and the 
controls, including samples incubated with S9. The results indicate 
that 2-DCB does not produce point or frameshift mutations in Salmonella 
and is not activated by S9. The study also investigated the toxicity of 
2-DCB and concluded ``that the potential risk from 2-DCB, if any, is 
very low'' (Ref. 44).
    One comment states that 2-DCB is a unique radiolysis byproduct of 
palmitic acid, and ``[b]ecause palmitic acid appears in molluscan 
shellfish in varying quantities and high percentages, the FDA should 
refrain from considering the petition until potential cytotoxicity and 
genotoxicity of 2-DCB in each type of shellfish covered by the petition 
is thoroughly studied.''
    FDA agrees that 2-DCB is a radiation by-product of triglycerides 
with esterified palmitic acid and that molluscan shellfish contain 
significant amounts of such triglycerides. FDA previously reviewed 
studies in which animals were fed diets containing irradiated meat, 
poultry, and fish which contain triglycerides with palmitic acid (62 FR 
64107 at 64113), and concluded that no adverse effects were associated 
with the consumption of these irradiated flesh foods. The comment 
provides no evidence to refute the agency's conclusion regarding the 
irradiation of molluscan shellfish to a maximum absorbed dose that will 
not exceed 5.5 kGy.
    One comment states that two studies by Delinc[eacute]e et al. on 
the potential genotoxicity of 2-DCB were mischaracterized in the 1999 
FAO/IAEA/WHO report. The comment states that while ``[t]he 1999 FAO/
IAEA/WHO report properly labeled Study 5 as demonstrating a `possible 
effect of high-dose irradiation.'* * * it rationalized this by saying 
the level of the lipid

[[Page 48066]]

present in the experiment was three orders of magnitude greater than 
the normal lipid level in chicken meat.'' In addition, the comment 
states that ``[s]tudy 6 did not, in fact, use an `extremely high level' 
of 2-DCB as claimed in the WHO Secretariat's proof note. The level of 
2-DCB, according to the researchers, was carefully calibrated and 
multiplied by the appropriate toxicological safety factor, to determine 
the safety of chicken irradiated for shelf sterilization.'' In summary, 
the comment states that ``Delinc[eacute]e et al. conclude that applying 
the standard toxicological safety factor of 100 below the `no-effect 
level' means that 2-DCB failed the standard safety test'' and should be 
denied under Sec.  170.22 (21 CFR 170.22).
    In the first study cited, Delinc[eacute]e et al. incubated rat and 
human colon cells for 30 minutes in solutions containing 0.3-1.25 mg/ml 
2-DCB and determined by the COMET assay that there were single strand 
DNA breaks (Ref. 45). The authors also state that they observed a 
cytotoxic effect at increased concentration. Cytotoxicity can confound 
the results of the COMET assay such that standard protocols attempt to 
use concentrations below that producing cytotoxicity (Ref. 46). 
Delinc[eacute]e notes that the 2-DCB concentration in the lipid 
fraction of chicken irradiated at 58 kGy (Raltech study) is 17 microg/g 
lipid (Refs. 45 and 47). Thus, the concentration of 2-DCB used in the 
assay was 17 to 73 times higher than that in the lipid fraction of 
radiation sterilized chicken. As the average dose