Agricultural Uses of Antibiotics - ACS Publications - American

11

Antibiotic Residues in Food: Regulatory Aspects

Robert C. Livingston

Downloaded by NATL UNIV OF SINGAPORE on November 8, 2017 | http://pubs.acs.org Publication Date: September 18, 1986 | doi: 10.1021/bk-1986-0320.ch011

Center for Veterinary Medicine, U.S. Food and Drug Administration, Rockville, MD 20857

The Food and Drug Administration has the r e s p o n s i b i l i t y of ensuring that residues of drugs i n animal-derived food are safe for human consumption. The permitted residue levels and the conditions of use of each i n d i v i d u a l drug are determined by t o x i c o l o g i c a l and chemic a l studies. The studies required for a n t i b i o t i c s vary according to the drug and the proposed application. T h i s paper d i s c u s s e s the f o l l o w i n g p o i n t s : (1) the requirements for approval of a new a n t i b i o t i c as well as those for a new use of an approved a n t i b i o t i c , (2) the e f f e c t s of recent changes i n the requirements on the amount of r e s i d u e s permitted i n food, and (3) d e f i c i e n c i e s i n current methods for determining a n t i b i o t i c residues i n food.

The Food & Drug Administration has the r e s p o n s i b i l i t y for the premarket clearance of a l l animal drugs. The 1958 food additive amendment to the Federal Food, Drug & Cosmetic Act requires sponsors to demonstrate the safety of their products. The Kefauver-Harris amendment of 1962 requires the sponsors to demonstrate, i n addition to safety, the e f f i c a c y of t h e i r drugs. Safety implies safety to the animal as well as to the consumers of animal products. The role of the Center f o r Veterinary Medicine i n the premarket approval process i s to e s t a b l i s h conditions of drug use and to e s t a b l i s h the allowable tolerances for drug residues i n animal-derived food products. The two major questions concerning the use of a n t i b i o t i c s i n agriculture are the safety of the residues i n the animal-derived food and the a n t i b i o t i c resistance that may develop from the use of these drugs i n animals. I w i l l not talk about a n t i b i o t i c resistance as Mr. Frappaolo discusses this issue i n a separate paper. The residue issue can be further divided into the t o x i c i t y and the a l l e r g i c reaction to the drug residues. There i s s u f f i c i e n t concern f o r the a l l e r g i c reaction to p e n i c i l l i n that i t s tolerance i s based upon this concern; however, the rest of the a n t i b i o t i c s have tolerances based on t o x i c i t y other than the a l l e r g i c reaction. This chapter not subject to U.S. copyright. Published 1986, American Chemical Society

Moats; Agricultural Uses of Antibiotics ACS Symposium Series; American Chemical Society: Washington, DC, 1986.

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LIVINGSTON

Antibiotic Residues in Food

129

Drugs are used i n agriculture to promote growth, improve feed e f f i c i e n c y and to control disease. Modern methods of producing animal-derived food depend heavily on the use of a n t i b a c t e r i a l substances. In discussing the regulatory concerns for a n t i b i o t i c s i n agriculture, one needs to review how tolerances have been established for drug residues i n animal products. Table 1 gives the


TABLE I.

DEFINITIONS OF TOLERANCES AND TOXICITY TESTS FOR REQUESTED TOLERANCE WITH SAFETY FACTORS

Toxicity test required

Tolerance

Definition

Negligible tolerance*

Toxicologically insignificant residue

90-Day subacute study i n rat and dog (preferably i n utero for rat)

Finite tolerance

Measureable amount of residue

Lifetime studies i n rat and mouse; 6-month study i n dog; 3-generation reproduction study with teratologic phase

Safety factors 2,000

100+

- Residue must be < 0.1 ppm i n meat and < 10 ppb i n milk and eggs. + - If teratogenic a c t i v i t y i s demonstrated, the safety factor i s 1,000; may also be < 100 when human exposure data are available or when a sensitive measurement i s used to set a no-effect concentration.

d e f i n i t i o n of two types of tolerances that have been used i n regulating animal drugs since 1966. A n e g l i g i b l e tolerance has a value of 0.1 part per m i l l i o n (ppm) i n meat. Negligible tolerances were obtained by drug sponsors by conducting two ninety-day subacute studies generally one i n the rat and one i n the dog. A safety factor of 2000 was used to calculate tolerances based on these two studies. If the calculated tolerance exceeded 0.1 ppm, the t o l e r ance was a r b i t r a r i l y set at 0.1 ppm; consequently, most a n t i b i o t i c s have tolerances of 0.1 ppm. If a sponsor desired a higher tolerance than 0.1 ppm, additional t o x i c o l o g i c a l studies were required. To obtain a f i n i t e tolerance, that i s a tolerance above 0.1 ppm, l i f e time studies i n the rat and mouse were required; i n addition, a s i x month study i n the dog and a three generation reproduction study with a t e r a t o l o g i c a l phase were also required. Because of the chronic nature of these studies, the safety factor was reduced from 2000 to 100. The equation for c a l c u l a t i n g a tolerance based upon the toxi c i t y studies i s presented i n Table 2. The various t o x i c i t y studies are examined to determine the lowest no-effect l e v e l i n each of the species. The no-effect l e v e l i n the most sensitive species i s used to determine the tolerance. The tolerance i s equal to the no-effect


130

AGRICULTURAL USES OF ANTIBIOTICS

TABLE I I .

CALCULATION OF TOLERANCE FOR A DRUG RESIDUE

TOLERANCE

NEL X 60 KG (SF) (FF) (0.5 KG/DAY)


NEL - NO-EFFECT LEVEL IN THE MOST SENSITIVE TEST SPECIES SF

- SAFETY FACTOR

FF

= FOOD FACTOR

l e v e l times the average weight of a person (60 kg) divided by the safety factor, a consumption factor and 0.5 kg, the estimated consumption of meat per day. The consumption factor i s an acknowledgement that organ meats, such as l i v e r and kidney, are not consumed to the same extent as muscle tissue. The consumption factors f o r the various edible products of the different species are given i n Table 3. For example, the consumption factor for muscle i n a l l species i s

TABLE III.

RELATIVE FACTORS FOR ASSIGNING NEGLIGIBLE TOLERANCES MAJOR SPECIES CATEGORIES Tissue

Beef

Pork

Muscle Liver Kidney Skin Fat

1 2 3

1 3 4 4 4

* Not used f o r human

-* 4

Sheep 1 5 5

-* 5

Poultry 1 3

-* 2 2

food.

1. The consumption factor for beef l i v e r i s 2. Because of this doubl i n g of the consumption factor, the tolerance i n l i v e r can be twice the value of the tolerance f o r the drug i n muscle. The consumption factor f o r pork l i v e r i s 3, indicating that pork l i v e r i s consumed less than beef l i v e r . Because of these consumption factors, the tolerances i n the Code of Federal Regulations (CFR) d i f f e r depending on what edible tissue i s being described. However, some of the older tolerances i n the CFR give the same value f o r a l l edible t i s sues. These drugs were regulated before the use of consumption factors were developed. The v i o l a t i o n rate f o r a n t i b i o t i c s , as determined by USDA, also needs to be examined i n order to discuss the regulatory concerns for a n t i b i o t i c residues. Table 4 l i s t s the v i o l a t i v e residue rates f o r a n t i b a c t e r i a l s i n several species f o r the years 1979 through 1983.


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TABLE IV.


VIOLATIVE RESIDUE RATES FOR ANTIBACTERIALS

1979 MATURE CATTLE CHICKENS TURKEYS BOB VEAL SWINE Downloaded by NATL UNIV OF SINGAPORE on November 8, 2017 | http://pubs.acs.org Publication Date: September 18, 1986 | doi: 10.1021/bk-1986-0320.ch011

131

2.2 nil 2.4 7.8 10

1980

3.9 6.8

1981

7.3 8.7

(%)

1982

6.1 7

1983 .2 nil .01 7.7 9.2

The residue v i o l a t i o n rate i n mature c a t t l e , chickens, and turkeys, i s very low, 0.2% or less. In f a c t , chickens have almost a zero v i o l a t i o n rate. This i s due to the highly integrated chickenproducing operations i n this country, whereas turkeys are raised more by independent producers. However, the v i o l a t i o n rate for t u r keys i s s t i l l very low. The v i o l a t i o n rate i s not low for a l l species. Bob veal through 1979 to 1983 has had a very large v i o l a t i o n rate r e l a t i v e to the other species. This i s due to the fact that bob veal are given drugs to keep them a l i v e u n t i l they are marketed. As bob veal are marketed as young as 10 days of age, the l i k e l i h o o d of withdrawal periods being followed for bob veal i s not high. Some of the drugs used i n bob veal require more than 10 days to deplete to below their established tolerances. The v i o l a t i o n rate i n swine i s also r e l a t i v e l y high. This i s primarily due to residues of sulfamethazine. The high v i o l a t i o n rate for sulfamethazine i n swine i s due to several factors. Powdered sulfamethazine i s e l e c t r o s t a t i c and tends to adhere to mixing equipment. This effect leads to contamination of nonmedicated feed. Studies indicate that average levels of contamination as high as 3 ppm can occur. These levels i n the withdrawal feed of swine can cause v i o l a t i v e residue l e v e l s . Another problem i s i n the husbandry of swine. Pigs are coprophagic and as l i t t l e as 2-3 ppm of s u l f a methazine i n the feces w i l l also result i n v i o l a t i v e residues. Compounding the problem has been the r e f u s a l of some producers to f o l low the withdrawal period. A recent publication of USDA indicates that half of the v i o l a t i o n s of sulfamethazine i n swine are the result of producers not following the withdrawal period (1). Based on the conservative nature of the n e g l i g i b l e tolerance concept and the low v i o l a t i o n rate of a n t i b a c t e r i a l s , the regulatory concern for a n t i b i o t i c s i s not large. The Center for Veterinary Medicine i s no longer using the concept of a n e g l i g i b l e tolerance i n i t s approval process. The current procedures for c a l c u l a t i n g tolerances for drug residues have the potential of further reducing our regulatory concern for many of the approved animal drugs. Table 5 l i s t s the minimal t o x i c o l o g i c a l t e s t ing for an animal drug by today's standards. These tests e s s e n t i a l l y replace the studies required to obtain a n e g l i g i b l e tolerance, i . e . , the two ninety-day feeding studies. The f i r s t thing that i s required i s a battery of genetic t o x i c i t y tests to help assess potential carcinogenicity of an animal drug. The second requirement i s the


132


TABLE V.

0 0


0

MINIMUM TOXICOLOGICAL TESTING FOR AN ANIMAL DRUG

A BATTERY OF GENETIC TOXICITY TESTS A 90-DAY FEEDING STUDY BOTH IN A RODENT SPECIES (USUALLY THE RAT) AND IN A NON-RODENT MAMMALIAN SPECIES (USUALLY THE DOG). A TWO-GENERATION REPRODUCTION STUDY WITH A TERATOLOGY COMPONENT IN RATS.

ninety-day feeding studies i n both a rodent species, usually the rat and i n a non-rodent mammalian species, usually the dog. The t h i r d requirement i s a two-generation reproduction study with a teratology component i n rats. Although the minimum t o x i c o l o g i c a l studies required by today's standards are more extensive, the 0.1 ppm cap for the tolerance has been raised to 1.0 ppm i n the t o t a l d a i l y diet of an i n d i v i d u a l . Assuming that one-third of the d a i l y diet i s composed of meat products, the 1 ppm i n the diet means that a tolerance of up to 3 ppm i n the meat can be obtained based on these studies. The 3 ppm i s the tolerance i n muscle t i s s u e . Using the consumption factors previously discussed, the t o l e r ance i n kidney, l i v e r , and skin/fat can be several multiples higher than 3 ppm. Most drugs that we see i n the program today would not require a tolerance higher than 3 ppm because their residue levels are usually much less than 3 ppm i n muscle t i s s u e . In fact, several drugs have tissue residues i n the ppb range at zero withdrawal. I f a drug requires an assigned tolerance greater than 3 ppm to obtain approval, i f the residues bioaccumulate, or i f i t i s a suspect carcinogen, additional t o x i c o l o g i c a l tests are required. Table 6 l i s t s

TABLE VI.

TOXICOLOGICAL TEST REQUIRED WHEN RESIDUE LEVELS EXCEED 3PPM,THE DRUG IS A SUSPECT CARCINOGEN, OR THE DRUG IS EXPECTED TO BIOACCUMULATE

0

Chronic bioassays for oncogenicity/chronic t o x i c i t y i n each of two rodent species.

0

A chronic bioassay (one year) i n a non-rodent mammalian species (usually the dog).

0

A teratology study i n a second species.

0

Other specialized t e s t i n g i f necessary.



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133

the t o x i c o l o g i c a l studies required under these conditions. In addition to the subchronic studies, chronic studies are required i n two rodent species and a non-rodent species, usually the dog. Also, a teratology study i s required i n a second species and depending on s p e c i f i c concerns other specialized testing may be required. A l i b e r a l i z i n g aspect of the new t o x i c o l o g i c a l requirements i s that the safety factor for subchronic studies has been reduced from 2000 to 1000. The recent change i n the method of calculating tolerances within the animal drug program i n CVM w i l l have quite a dramatic effect on the permitted tolerances supported by subchronic studies. Table 7 l i s t s a few representative drugs that are currently regulated i n food-producing animals. The f i r s t column i s the no-effect

TABLE VII.

SELECTED ANTIBIOTICS APPROVED FOR USE IN FOOD PRODUCING ANIMALS

Drug Apramycin Bacitracin (Zn, MD) Erythromycin Gentamicin Oleandomycin Oxytetracycline Tylosin

NEL (mg/kg) 25 >50 25 60 200 365 40

CFR T o l . (ppm) 0.1 0.5 0.1 0.1 0.15 1.0 0.2

Possible T o l . (ppm) 3.0 6.0 3.0 7.2 24.0 438.0 48.0

l e v e l for the drug that was used to determine the current tolerance as l i s t e d i n the CFR. The second column l i s t s the current tolerance. The t h i r d column indicates the possible tolerance based upon the formula given i n Table 2. Apramycin, for example, would have a poss i b l e tolerance of 3 ppm. This i s asubstantial increase over the present tolerance of 0.1 ppm. S i m i l a r i l y , the current tolerance of 0.5 ppm for b a c i t r a c i n could possibly be j u s t i f i e d as 6 ppm based upon the no-effect l e v e l alone. However, current policy would l i m i t the tolerance i n muscle to 3 ppm. The tolerance for erythromycin would increase from 0.1 up to 3 ppm. Gentamicin could possibly jump from 0.1 to 7.2, but again would be limited to 3.0 based upon our p o l i c y . The no-effect l e v e l of the l a s t three drugs l i s t e d i n the table, oleandomycin, oxytetracycline, and t y l o s i n , indicate the safety of these compounds. These drugs would a l l be candidates f o r a revised tolerance of 3 ppm based on conventional t o x i c i t y . These tolerances are not a l l being automatically revised i n the CFR f o r several reasons: (1) the subchronic studies used to calculate the current tolerances may not meet todays standards, (2) most of these drugs were regulated on the basis of residues of parent drug only, (3) the o f f i c i a l methods for monitoring the residues may not meet present standards, and (4) some of the drugs have safety concerns that are not s a t i s f i e d by subchronic studies.



134


Present standards require that drugs be regulated on the basis of t o t a l residues. Total residues r e s u l t i n g from drug administration to an animal consist of the parent drug and a l l compounds derived from i t , i . e . , metabolites, conjugates, and residues bound to b i o l o g i c a l macromolecules. Total residues are t y p i c a l l y determined i n a l l edible tissues by dosing the animal under proposed use conditions with a radiolabeled drug. Several animals are usually employed i n such a study to permit their s e r i a l s a c r i f i c e after the drug has last been administered. From such an experiment, the depletion of t o t a l residues i n each of the tissues can be followed. Figure 1 represents a t y p i c a l depletion curve for t o t a l residues of a drug. The withdrawal period i s approximated by the point i n time where the t o t a l residue curve intersects the safe concentration l e v e l , previously referred to as the tolerance, as determined by the toxicol o g i c a l studies and the formula i n Table 2. To ensure compliance with the withdrawal period, an assay i s needed to monitor t o t a l residues i n the edible tissues. Because i t i s impractical to develop assays for each residue i n each of the edible tissues, the concept of a marker residue and a target tissue i s introduced. The marker residue i s a selected analyte whose l e v e l i n a p a r t i c u l a r tissue has a known relationship to the l e v e l of the t o t a l residue of t o x i c o l o g i c a l concern i n a l l edible tissues. Therefore, i t can be taken as a measure of the t o t a l residue of interest i n the target animal. The information obtained from studies of the depletion of the radiolabeled t o t a l residue can be used to calculate a l e v e l of the marker residue that must not be exceeded i n a selected tissue (the target tissue) i f the t o t a l residue of toxicol o g i c a l concern i n the edible tissues of the target animal i s not to exceed i t s safe concentration. In the example depicted i n Figure 1, the safe concentration i s 2.0 ppm. The marker residue i s at a l e v e l of 1.0 ppm when the safe concentration i s 2.0 ppm. The method i s developed f o r the marker residue at 1.0 ppm and the tolerance for the drug i s 1.0 ppm of the marker residue. The o v e r a l l effect of regulating on t o t a l residues as opposed to the parent drug i s a lowering of the tolerance. The amount by which the tolerance decreases depends on the proportion of the parent drug to the t o t a l residues. For some a n t i b i o t i c s the parent drug i s a good approximation of t o t a l residues because they are not metabolized. For other drugs the parent drug i s a vanishingly small f r a c t i o n of the t o t a l residue and the parent drug would not serve as a marker residue for the t o t a l residue. In the l a t t e r case, the tolerances would be greatly reduced based upon the low percentage of parent drug. A few of the currently regulated a n t i b i otics would not require t o t a l residue studies to support requests for new uses. The tetracyclines are not s i g n i f i c a n t l y metabolized and the parent drug i s a good approximation of the t o t a l residues. We do recognize that degradation to the epi form may occur to a small extent. The aminoglycosides undergo limited metabolism and t h e i r absorption from the GI tract i s low. Residue studies on gentamicin using microbiological assays, radiotracers and RIA techniques a l l gave the same r e s u l t s . The lack of absorption also has been demonstrated with the polypeptides b a c i t r a c i n and bambermycins. Another reason that the tolerance i s not automatically raised i n accordance with our new policy, i s the question of adequate


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LIVINGSTON



SAFE CONCENTRATION = 2.0 PPM TOLERANCE = 1.0 PPM

TIME (DAYS)

Figure 1. Typical depletion curve f o r t o t a l residues and a marker residue i n an edible tissue.



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methods to monitor residues. Most of the a n t i b i o t i c s have microbiol o g i c a l assays as methods for monitoring residues. These methods are not s p e c i f i c and are limited to measuring b i o l o g i c a l l y active residues. The adequacy of the extraction procedures for these methods has recently been questioned. If the method does not measure a l l of the drug residue, the withdrawal periods w i l l be too short. The need for chemical methods for many of the a n t i b i o t i c s was the subject of a recent Association of O f f i c i a l A n a l y t i c a l Chemists symposium. Sulfamethazine i s an example of a drug where the tolerance would not be raised based only on subchronic studies because of i t s possible carcinogenicity. FDA i s presently conducting chronic studies i n both rats and mice as well as a t o t a l residue study at the National Center for Toxicological Research. These studies are to be completed next year. As the Director of CVM stated i n a speech at the Food Editors Conference i n Dallas l a s t June, these studies " w i l l either exonerate sulfamethazine or w i l l incriminate i t to a point incompatible with continued approval." In either case, the v i o l a t i o n problem disappears. In summary, the regulatory concerns for residues of regulated a n t i b i o t i c s i s not large. This i s due to the conservative procedures for s e t t i n g most tolerances and the low v i o l a t i o n rates. Another reason f o r the lack of concern of residues of regulated a n t i b i o t i c s i s the number of new a n t i b i o t i c s that q u a l i f y for zero withdrawal periods. The fact that t h e i r r e l a t i v e safety enables them to obtain zero withdrawal periods places competitive pressure on sponsors to also develop safer drugs. There are some s p e c i f i c problems but they are being addressed and the future v i o l a t i o n rates should even be lower than present l e v e l s . Literature Cited 1.

Fed. Regis. 50:

20796 (May 20,

1985)

R E C E I V E D June 10, 1986


Agricultural Uses of Antibiotics - ACS Publications - American

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