NATION& TOXICOLOGY PROGRAM 

Technical Report Series 

No. 455 

TOXICOLOGY AND CARCINOGENESIS 

STUDIES OF 

CODEINE 

(CAS NO. 76-57-3) 

~. . . - . 

IN F344/NRATS AND B6C3Fl MICE 

(FEED STUDIES) 

U.S. DEPART” OF HEALTH AND HUMAN SERVICES 

Pnblic Health Service 

National Institutes of Health 
FOREWORD 

The National Toxicology Program (NTP) is made up of four charter agencies of the U.S. Department of 

Health and Human Services (DHHS): the National Cancer Institute (NCI), National Institutes of Health; 

the National Institute of Environmental Health Sciences (NIEHS), National Institutes of Health; the 

National Center for Toxicological Research (NCTR), Food and Drug Administration; and the National 

Institute for Occupational Safety and Health (NIOSH), Centers for Disease Control. In July 1981, the 

Carcinogenesis Bioassay Testing Program, NCI, was transferred to the NIEHS. The NTP coordinates the 

relevant programs, staff, and resources from these Public Health Service agencies relating to basic and 

applied research and to biological assay development and validation. 

The NTP develops, evaluates, and disseminates scientific information about potentially toxic and 

hazardous chemicals. This knowledge is used for protecting the health of the American people and for the 

primary prevention of disease. 

The studies described in this Technical Report were performed under the direction of the NIEHS and were 

conducted in compliance with NTP laboratory health and safety requirements and must meet or exceed all 

applicable federal, state, and local health and safety regulations. Animal care and use were in accordance 

with the Public Health Service Policy on Humane Care and Use of Animals. The prechronic and chronic 

studies were conducted in compliance with Food and Drug Administration (FDA) Good Laboratory 

Practice Regulations, and all aspects of the chronic studies were subjected to retrospective quality 

assurance audits before being presented for public review. 

These studies are designed and conducted to characterize and evaluate the toxicologic potential, including 

carcinogenic activity, of selected chemicals in laboratory animals (usually two species, rats and mice). 

Chemicals selected for NTP toxicology and carcinogenesis studies. are chosen priniarily on the bases of 

human exposure, level of production, and chemical structure. The interpretive conclusions presented in 

this Technical Report are based only on the results of these NTP studies. Extrapolation of these results to 

other species and quantitative risk analyses for humans require wider analyses beyond the purview of these 

studies. Selection per se is not an indicator of a chemical’s carcinogenic potential. 

These NTP Technical Reports are available for sale from the National Technical Information Service, 

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copies of this Technical Report are available without charge while supplies last from NTP Central Data 

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Listings of all published NTP reports and ongoing studies are also available from NTP Central Data 

Management. The Abstracts and other study information for 2-year studies are also availableon the 

NTP’sWorldWideWebsite: http://ntp-server.niehs.nih.gov. 
NTP TECHNICAL REPORT 

ON THE 

TOXICOLOGY AND CARCINOGENESIS 

STUDIES OF 

CODEINE 

(CAS NO. 76-57-3) 

IN F344/N RATS AND B6C3Fl MICE 

(FEED STUDIES) 

NATIONAL TOXICOLOGY PROGRAM 

P.O. Box 12233 

ResearchTrianglePark,NC27709 

August 1996 

NTP TR 455 

NIH Publication No. 96-3360 

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES 

Public Health Service 

National’hstitutes of Health 
2 Codeine, NTP TR 455 

CONTRIBUTORS 

National Toxicology Program 

Evaluated and interpreted results and reported jindings 

J.K. Dunnick, Ph.D., StudyScientist 

G.A. Boorman, D.V.M., Ph.D. 

D.A. Bridge, B.S. 

J.R. Bucher, Ph.D. 

L.T. Burka, Ph.D. 

M.R. Elwell, D.V.M., Ph.D. 

J.R. Hailey, D.V.M. 

J.K. Haseman, Ph.D. 

D.S. Marsman, D.V.M., Ph.D. 

G.N. Rao, D.V.M., Ph.D. 

J.H. Roycroft, Ph.D. 

G.S. Travlos, D.V.M. 

D.B. Walters, Ph.D. 

K.L. Witt, M.S., Oak RidgeAssociatedUniversities 

Microbiological Associates, Inc. 

Conducted 14-day and 13-week studies, 

evaluated pathology findings 

H.S. Lilja, Ph.D., PrincipalInvestigator 

M. Hagopian, Ph.D. 

D.S. Wyand, D.V.M. 

Conducted 2-year studies, evaluated pathology jindings 

M.L. Wenk, Ph.D., StudyDirector 

L.H. Brennecke, D.V.M. (rats) 

M.A. Stedham, D.V.M. (mice) 

Experimental Pathology Laboratories, Inc. 

Provided pathology quality assurance 

J.F. Hardisty, D.V.M., PrincipalInvestigator 

K. Yoshitomi, D.V.M., Ph.D. 

Dynamac Corporation 

Prepared quality assurance audits 

S. Brecher, Ph.D., PrincipalInvestigator 

NTP Pathology Working Group 

Evaluated slides, prepared pathology report on rats 

(6October 1993) 

P.K. Hildebrandt, D.V.M., Chair 

PATHCO, Inc. 

R.C. Cattley, M.S., V.M.D., Ph.D. 

Chemical Industry Institute of Toxicology 

A. Enomoto, D.V.M. 

National Toxicology Program 

J.R. Hailey, D.V.M. 

National Toxicology Program 

A. Radovsky, D.V.M., Ph.D. 

National Toxicology Program 

R.C. Sills, D.V.M., Ph.D. 

National Toxicology Program 

K. Yoshitomi, D.V.M., Ph.D. 

Experimental Pathology Laboratories, Inc. 

Evaluated slides, prepared pathology report on mice 

(14 December 1993) 

P.K. Hildebrandt, D.V.M., Chair 

PATHCO, Inc. 

J.M. Cullen, V.M.D., Ph.D. 

North Carolina State University 

D. Dixon, D.V.M., Ph.D. 

National Toxicology Program 

R.A. Herbert, D.V.M., Ph.D. 

National Toxicology Program 

A. Radovsky, D.V.M., Ph.D. 

National Toxicology Program 

R.C. Sills, D.V.M., Ph.D. 

National Toxicology Program 

K. Yoshitomi, D.V.M., Ph.D. 

Experimental Pathology Laboratories, Inc. 

Analytical Sciences, Inc. 

Provided Statistical analyses 

R.W. Morris, M.S., PrincipalInvestigator 

N.G. Mintz, B.S. 

S. Rosenblum, M.S. 

Biotechnical Services, Inc. 

Prepared Technical Report 

S.R. Gunnels, M.A., PrincipalInvestigator 

G. Gordon, M.A. 

L.M. Harper, B.S. 

S.M. Swift, B.S. 
3 

CONTENTS 

ABSTRACT ......................................................... 5 

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTMTY ........... 9 

TECHNICAL REPORTS REVIEW SUBCOMMI'ITEE ............................. 10 

SUMMARY OF TECHNICAL REPORTS REVEW SUBCOMMITTEE COMMENTS......... 11 

INTRODUCTION ..................................................... 13 

MATERIALSANDMETHODS ............................................ 23 

RESULTS .......................................................... 33 

DISCUSSION AND CONCLUSIONS ......................................... 57 

REFERENCES ....................................................... 61 

APPENDIX A Summary of Lesions in Male Rats in the 2-Year Feed Study 

ofcodeine ............................................... 71 

APPENDIXB Summary of Lesions in .Female Rats in the 2-Year.Feed Study 

ofcodeine ............................................... 107 

AppENDnr C Summary of Lesions in Male Mice in the 2-Year Feed Study 

ofcodeine ............................................... 145 

APPENDIXD Summary of Lesions in Female Mice in the 2-Year Feed Study 

ofcodeine ................................................ 177 

APPENDIXE GeneticToxicology .......................................... 217 

APPENDIXF Organ Weights and Organ-Weight-to-Body-Weight Ratios ................ 227 

AppENDnr G Hematology. Clinical Chemistry. and Urinalysis Results .................. 239 

ApPJm.DEH Reproductive Tissue Evaluations 

and Estrous Cycle Characterization ............................... 245 

APPENDIX I Chemical Characterization and Dose Formulation Studies ................. 249 

APPENDIXJ Feed and Compound Consumption in the 2-Year Feed Studies of Codeine ...... 263 

APPENDIXK Ingredients. Nutrient Composition. and Contaminant Levels 

in NIH-07 Rat and Mouse Ration ................................ 269 

APPENDIX L Sentinel Animal Program ..................................... 273 

APPENDIX M Codeine Toxicokinetics in Rats During a Two-Year Dosed Feed Study ......... 277 
5 

ABSTRACT 

CODEINE 

CAS NO.76-57-3 

ChemicalFormula: C18HZIN03 Molecular Weight: 299.36 

Synonyms: 7,8didehydro4,5-epoxy-3-methoxy-17-methylmorphinan-6-ol;methylmorphine;3-0-methylmorphinemonohydrate; 

N-methylnorcodeine; morphine-3-methyl ether; morphine monomethyl ether 

Trade names: Codeinum,Codicept,Coducept, Metilmorfii 

Codeineisusedin a variety of pharmaceuticals 

includinganalgesics,sedatives,hypnotics,antiperistaltics, and antitussive agents. The National Cancer 

InstituteandtheFoodandDrugAdministration 

nominatedcodeine for study becauseitis a widely 

useddruganditisrepresentative ofthemorphine 

classofcompounds, for which chronic carcinogenicitystudieshadnotbeenconducted.Theoral 

route of administration was selected because it is the 

primary route of human exposure. Male and female 

F344/NratsandB6C3F,miceweregivencodeine 

(99% pure)infeed for 14days,13weeks,or 

2 years. Genetictoxicologystudieswereconducted 

in Salmonella typhimurium andculturedChinese 

hamster ovary cells. 

14-DAY STUDY IN RATS 

Groups of five maleandfivefemaleF344/Nrats 

weregiven 0, 1,562, 3,125, 6,250, 12,500, or 

25,000 ppm codeineinfeed for 14 days,which 

resultedindailydosesofapproximately125,250, 

450,650, or 750 mg codeine/kg bodyweightto 

malesand125,250, 500, 700, or 300'mg/kg to 

females.Onefemaleexposed to 6,250 ppm, one 

male and three females exposed to 12,500 ppm, and 

allmalesandfemalesexposedto25,000ppmdied 

during the study. Final mean body weights and mean 

bodyweightgainsofallexposed groupsexcept 

1,562 ppm femalesweresignificantlylowerthan 

those of the controls. 

No chemical-related gross lesions were observed in 

ratsatnecropsy.Thickeningoftheforestomach 

mucosa and and (hyperplasiahyperkeratosis) 

lymphoid depletion of the thymus in exposed males 

andfemalesandtesticulardegenerationinexposed 

males, primarilythe and observed in12,500 

25,000 ppm groups, were associated with decreased 

survival and increased morbidity in these groups. 

14-DAY STUDY IN MICE 

Groups of fivemaleandfivefemaleB6C3Flmice 

were given 0, 781, 1,562, 3,125, 6,250, or

12,500 ppm codeineinfeed for 14 days,which 

resultedindailydoses of approximately 150,300, 

600, 1,300, or 3,000 mg codeine/kg body weight to 

malesand 200, 400, 750, 1,500, or 3,000 mg/kg to 

females. All mice survived to the end of the study. 

The final mean body weight of 3,125 ppm females 

was significantly greater than thatof the controls; the 

final mean body weight of 12,500 ppm females and 

themeanbodyweightgainsof 12,500ppmmales 

andfemalesweresignificantlylowerthanthoseof 

the controls. 
6 

Absolute and relative liver weights of 3,125, 6,250, 

and 12,500 ppmmalesandof 12,500 ppmfemales 

and the absolute and relative right kidney weights of 

12,500ppmmalesweresignificantlylowerthan 

thoseofthecontrols.Nogrossorhistopathologic 

lesions were attributed to codeine exposure. 

H-WEEK STUDY IN RATS 

Groups of 10 male and 10 female F344/N rats were 

given 0, 390,781,1,562,3,125, or 6,250ppm

codeine in feed for13 weeks, which resulted in daily 

doses of approximately25, 50, 100, 200, or 450 mg 

codeine/kgbodyweighttomalesand 25, 50, 100, 

250, or 500 mg/kgtofemales.Therewereno 

chemical-related deaths during the study. Final mean 

bodyweightsandmeanbodyweightgainsofall 

groups of exposed males and of females exposed to 

1,562, 3,125, or 6,250 ppm were significantly lower 

thanthoseofthecontrols.Feedconsumptiondecreasedincreasing concentration with exposure 

during the first week of the study; however, by the 

end of the study, feed consumptionby most exposed 

groupswassimilartothatbythecontrols.There 

werealterationsof prious hematology andclinical 

chemistry parameters at the end of the study. There 

was a mild dose-dependent lymphopenia in females 

receiving 1,562ppmandaboveandin 6,250 ppm 

males.Therealsowas a minimal tomildmacrocytosis that occurred in all exposed groups of males 

and in females exposed to781, 3,125,or 6,250 ppm. 

Nosignificantdifferencesbetweencontroland 

exposed rats were observed in sperm morphology or 

vaginal cytology parameters. 

Absoluteandrelativeadrenalglandweightsof 

exposed males and of 3,125 and 6,250 ppm females 

were significantly greater than those of the controls. 

Absolute and relative liver weights of exposed males 

were significantly lower than those of the controls. 

Relativethymusweightsof 3,125 and 6,250ppm 

malesweresignificantlylowerthanthatofthe 

controls. No chemical-related grossorhistopathologic lesions were observed in male or female rats. 

13-WEEK STUDY IN MICE 

Groups of 10 male and 10 female B6C3F, mice were 

given 0, 390, 781, 1,562,3,125, or 6,250 ppm

codeine in feed for13 weeks, which resulted in daily 

dosesofapproximately 60, 120, 260, 460, or 

1,000 mg codeine/kg body weight to males and 60, 

Codeine, NTP TR 455 

130,280,530, or 1,200 mg/kgtofemales.Two 

malemiceinthe 3,125 ppmgroupdiedduring

week 7. Allothermicesurvivedtotheend of the 

study.Finalmeanbodyweightsofexposedmales 

andfemalesweresimilartothoseofthecontrols. 

Feed consumption by exposed males and females was 

similartothat bythe controls.Abnormalposture 

was observed in all exposed groups of males. There 

werenosignificantdifferences in hematology or 

urinalysisparametersinmaleorfemalemice. 

Minor,sporadicchangesoccurredinafewofthe 

clinical chemistry parameters; they were not consideredbiologicallysignificant.Nosignificantdifferencesinspermmorphology or vaginalcytology 

parameters were attributed to codeine exposure. 

Absoluteandrelativekidneyweightsof 3,125 and 

6,250 ppmmaleswerelowerthanthoseofthe 

controls. No chemical-related differencesinorgan 

weightswereobservedinfemales.Nochemicalrelated gross or histopathologic lesions were observed 

in male or female .mice. 

%YEAR STUDY IN RATS 

Groups of 60 male and 60 female F344/N rats were 

feddietscontaining 0, 400, 800, or1,600 ppm 

codeine for up to 106 weeks, with 9 or 10 rats per 

group evaluated at 15 months. These exposure concentrations resulted in average daily doses of approximately 15, 30, and 70 mg codeine/kg body weight to 

males and 15, 40, and 80 mg/kg to females. 

Survival, Body Weights, Feed Consumption, 

and Clinical Findings 

Survival of 400 ppm females was significantly greater 

thanthatofthecontrols;survivalof all groups of 

exposedmalesandof 800 and 1,600 ppmfemales 

wassimilartothatofthecontrols.Therewasan 

exposure-related decrease in mean body weights of 

malesandfemales.Thefinalmeanbodyweightof 

1,600 ppm males was 88% that of the controls, and 

the f& meanbodyweightof 1,600 ppmfemales 

was 89%that of the controls. Feed consumption by 

exposed groups was similar to that by the controls. 

Chemical-relatedclinicalfindingswerelimitedto 

ocular discharge in exposed males and females. 

Pathology Findings

Absoluteandrelativeadrenalglandweightsof800 

and 1,600 ppm males were significantly greater than 

those of thecontrolsat 15 months.Therewereno 
Codeine, NTP TR 455 7 

increasedincidencesofneoplasmsattributableto 

codeine exposure at any site. At 2 years, there were 

exposure-relateddecreasesintheincidencesof 

adrenalmedullahyperplasiainmalesandfemales. 

There was an exposure-related decrease in the incidence of benign pheochromocytomas in males, and 

theincidencesinexposedmalesweresignificantly 

lowerthanthatinthecontrols.In 1,600 ppm 

femalestheincidences ofmammarygland fibroadenomas and of fibroadenomas or adenocarcinomas 

inallexposedgroupsofmiceweresignificantly 

greater than those in the controls, but there were no 

increases in thyroid gland follicular cell neoplasms. 

Theincidence of centrilobular fattychangeinthe 

liverof 3,000 ppmmaleswassignificantlylower 

thanthatinthecontrolsat15months,andthe 

decreased incidence appeared to be related to exposure level. At 2 years, the incidences of eosinophilic 

foci, foci of fatty change, centrilobular cytomegaly, 

andcentrilobularfattychangein 3,000 ppm males 

(combined) were significantly lower than those in thewere lower than those in the controls. The incidence 

controls. The decreased incidences of benign pheochromocytomasinmalesandmammaryglandneoplasms in females wereconsidered toberelatedto 

codeine exposure. 

%YEAR STUDY IN MICE 

Groups of 60 male and 60 female B6C3F, mice were 

feddietscontaining 0, 750,1,500, or 3,000 ppm 

codeine for up to 106 weeks, with 9 or 10 mice per 

group evaluated at 15 months. These exposure concentrations resulted in average daily doses of approximately 100, 200, or 400 mg codeine/kg body weight 

to males and females. 

Survival, Body Weights, Feed Consumption, 

and Clinical Findings 

Survival of exposed males and females was similar to 

that of the controls. Mean body weights of 750 and 

1,500 ppm males and females were similar to those 

of the controls throughout most of the study. Mean 

body weights of 3,000 ppm males and females were 

less than those of the controls from about week 13, 

andthefinalmeanbodyweights ofthesegroups 

were 86% and 82% those of the respective controls. 

Feed consumption by exposed groups was similar to 

that by the controls. 

Pathology Findings 

Therewerenoincreasedincidences ofneoplasms 

attributable to codeine xposureatanysite. At 

15 months, the incidence of thyroid gland follicular 

cell hyperplasia in3,000 ppm males was significantly 

greater than that of the controls, and this lesion was 

observedin 1,500 and 3,000ppmfemales.At 

2 years, the incidences of follicular cell hyperplasia 

of hepatocellular adenomasandtheincidenceof 

hepatocellular adenomasor carcinomas (combined) in 

3,000 ppm malesandfemalesweresignificantly

lower than those in the controls;this was considered 

to be related to lower body weights in these groups. 

GENETIC TOXICOLOGY 

Codeine phosphate was not mutagenic in any of four 

strains of Salmonellatyphimunum, with or without 

S9 metabolic activation enzymes, In cytogenetic tests 

withculturedChinesehamster ovarycells,codeine 

phosphateinduceddose-relatedincreasesinsister 

chromatidexchanges,withandwithout S9, onlyat 

concentration levels that caused cell cycle delay. No 

induction of chromosomal aberrations was noted in 

culturedChinesehamsterovarycellstreatedwith 

codeine phosphate, with or without S9. 

CONCLUSIONS 

Undertheconditionsofthese2-yearfeedstudies, 

there was no evidence of carcinogenicactivity* of 

codeineinmaleorfemaleF344/Nratsexposed to 

400, 800, or 1,600 ppm. Therewas no evidence of 

carcinogenicactivity of codeine in male or female 

B6C3F, mice exposed to 750, 1,500, or 3,000 ppm. 

Thyroidglandfollicularhyperplasia cell was 

increased in exposed male and female mice. 

Decreased incidences of benign pheochromocytomas

ofthe adrenalmedullainmaleratsandmammary 

glandfibroadenomasandfibroadenomas or adenocarcinomas (combined) in female rats were related to 

codeine exposure. 

* Explanation of Levels of Evidence of Carcinogenic Activity ison page 9. A summary of the Technical Reports Review Subcommittee 

comments and the public discussion on this Technical Repoa appearson page 1 1. 
8 Codeine, NTP TR 455 

Summary of the %Year Carcinogenesis and Genetic Toxicology Studies of Codeine 

Female Male 

F344lN Rats 

Doses 0,400,800,or 

1,600ppm

(approximately 15, 30, 

or 70 mglkg in feed) 

Body weights 1,600ppm group

lower than controls 

2-Year survival 29/50. 20/50, 21/50, 

rates 20150 

Nonneoplastic None 

effects 

Neoplastic effects None 

Decreased Adrenal medulla: 

neoplasm benign 

pheochromocytoma incidences 

(16149, 6/50. 6/50. 

3/50) 

Level of evidence No evidence 

of carcinogenic

activity 

Genetic toxicology 

Salmonella typhimurium gene mutations: 

Sister 'chromatid exchanges 

Cultured Chinese hamster ovary cellsin vitro: 

Chromosomal aberrations 

Cultured Chinese hamster ovary cells in vifro: 

Female 

MiceB6C3F, F344lN B6C3F, MiceRats

0,400,800,or 0, 750, 1,500, or 0,750. 1,500,or 

1,600ppm 3,000ppm 3,OOO ppm (approximately 15,40, (approximately 100, (approximately 100, or 80mglkg in feed) 200,or 400 mglkg in 200,or 400mglkg in 

feed) feed) 

1,600ppm group 3,000ppm group 3,OOO ppm grow 

lower than controls lower than controls lower than controls 

28/50. 38/50. 29/51,41/50, 38/50, 45/50,36/50, 36/51, 43/51,

32/51 43/50 35/50 

None Thyroid gland: Thyroid gland: 

follicular cell follicular cell 

hyperplasia (7/49, hyperplasia (14/48, 

25/50. 29/50. 34/50)29151, 42151, 44/50) 

None None None 

Mammary gland: None None 

fibroadenoma (27150, 

21/50. 27/51, 8/51); 

fibroadenoma or 

adenocarcinoma 

(30150, 23/50, 29/51, 

8/51) 

No evidence No evidence No evidence 

Negative in strains TA97, TA98, TA100, and TA1535 with and without S9

Positive with and without S9 

Negative with and without S9 
Codeine, NTP TR 455 9 

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY 

The National Toxicology Program describes the results of individual experiments on a chemical agent and notes the strength of the evidence 

for conclusions regarding each study. Negative results, in which the study animals do not have a greater incidence of neoplasia than control 

animals, do not necessarily mean that a chemical is not a carcinogen, inasmuch as the experiments are conducted under a limited set of 

conditions. Positive results demonstrate that a chemical is carcinogenic for laboratoryanimals under the conditions of the study and indicate 

that exposure to the chemical has the potential for hazard to humans. Other organizations, such as the International Agency for Research on 

Cancer, assign a strength of evidence for conclusions based on an examination of all available evidence, including animal studies such as 

those conducted by the NTP, epidemiologic studies, and estimates of exposure. Thus, the actual determination of risk to humans from 

chemicals found to be carcinogenic in laboratory animals requires a wider analysis that extends beyond the purview of these studies. 

Five categories of evidence of carcinogenic activity are used in the Technical Report series to summarize the strength of the evidence 

observed in each experiment: two categories for positive results (clear evidence and some evidence); one category for uncertain fiidings 

(equivocal evidence); one category forno observable effects (no evidence); and one category for experiments that cannot be evaluated 

because of major flaws (inadequate study). These categories of interpretative conclusions were first adopted in June 1983 and then revised 

in March 1986 for use in the Technical Report series to incorporate more specifically the concept of actual weight of evidence of 

carcinogenic activity. For each separate experiment (male rats, femalerats, male, mice, female mice), one of the following five categories is 

selected to describe the fimdings. These categories refer to the strength of the experimental evidence and not to potency or mechanism. 

l Clear evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a dose-related 

(i) increase of malignant neoplasms, (ii) increase of a combination of malignant and benign neoplasms, or (iii) marked increase 

of benign neoplasms if there is an indication from this or other studies of the ability of such tumors to progress to malignancy. 

l Some evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a chemical-related 

increased incidence of neoplasms (malignant, benign, or combined) in which the strength of the response is less than that 

required for clear evidence. 

l Equivocal evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a marginal increase 

of neoplasms that may be chemical related. 

l No evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing no chemical-related 

increases in malignant or benign neoplasms. 

l Inadequate study of carcinogenic activity is demonstrated by studies that, because of major qualitative or quantitative 

limitations, cannot be interpreted as valid for showing either the presence or absence of carcinogenic activity. 

When a conclusion statement for a particular experiment is selected, consideration must be given to key factors that would extend the actual 

boundary of an individual category of evidence. Such consideration should allow for incorporation of scientific experience and current 

understanding of long-term carcinogenesis studies in laboratory animals, especially for those evaluations that may be on the borderline 

between two adjacent levels. These considerations should include: 

l adequacy of the experimental design and conduct; l occurrence of common versus uncommon neoplasia; l progression (or lack thereof) from benignto malignant neoplasia as well as from preneoplastic to neoplastic lesions; l some benign neoplasms have the capacity to regress but others (of the same morphologic type) progress. At present, it is 

impossible to identify the difference. Therefore, where progression is known to be a possibility, the most prudent course 

is to assume that benign neoplasms of those types have the potential to.become malignant; 

l combining benign and malignant tumor incidence known or thoughto represent stages of progression in the same organ or 

tissue; 

l latencyintumorinduction; 

l multiplicityinsite-specificneoplasia; 

l metastases; 

l supporting information from proliferative lesions (hyperplasia) in the same site of neoplasia or in other experiments (same 

lesion in another sex or species); l presence or absence of dose relationships; l statistical significance of the observed tumor increase;. l concurrent control tumor incidence as well as the historical control rate and variability for a specific neoplasm; l survival-adjusted analyses and false positive or false negative concerns; l structure-activity correlations; and l in some cases, genetic toxicology. 
10 Codeine, NTP TR 455 

NATIONAL TOXICOLOGY PROGRAM BOARD OF SCIENTIFIC COUNSELORS 

TECHNICAL REPORTS REVIEW SUBCOMMITTEE 

The members of the Technical Reports Review Subcommittee who evaluated the draft NTP Technical Report on codeine on June 20, 1995, 

are listed below. Subcommittee members serve as independent scientists, not as representatives of any institution, company, or 

governmental agency. In this capacity, subcommittee members have five major responsibilities in reviewing NTP studies: 

l to ascertain that all relevant literature data have been adequately cited and interpreted, l to determine if the design and conditions of the NTP studies were appropriate, l to ensure that the Technical Report presents the experimental results and conclusions fully and clearly, l to judge the significance of the experimental results by scientific criteria, and l to assess the evaluation of the evidence of carcinogenic activity and other observed toxic responses. 

Arnold L. Brown, M.D., Chair 

University of Wisconsin Medical School 

Madison, WI 

Thomas L. Goldsworthy, Ph.D. 

Department of Experimental Pathology and Toxicology 

Chemical Industry Institute of Toxicology 

Research Triangle Park, NC 

Meryl H. Karol, Ph.D. 

Department of Environmental Occupational Health 

University of Pittsburgh 

Pittsburgh, PA 

Curtis D. Klaassen, Pl1.D. 

Department of Pharmacology and Toxicology 

University of Kansas Medical Center 

Kansas City, KS 

Claudia S. Miller, M. D.,Principal Reviewer 

University of Texas Health Sciences Center 

San Antonio, TX 

Janardan K.Reddy, M.D. 

Department of Pathology 

Northwestern University Medical School 

Chicago; IL 

* Didnotattend 

Irma Russo, M.D.* 

Fox Chase Cancer Center 

Philadelphia, PA 

Louise Ryan, Ph.D. 

Division of Biostatistics 

Dana-Farber Cancer Institute 

Boston, MA 

Robert E. Taylor, M.D., Ph.D., principalReviewer 

Department of Pharmacology 

Howard University College of Medicine 

Washington, DC 

Mary JO Vodicnik, Ph.D., PrincipalReviewer 

. Lilly Research Laboratories 

Greenfield, IN 

Jerrold M. Ward, D.V.M., Ph.D. 

National Cancer Institute 

Frederick, MD 
Codeine, NTP TR 455 11 

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS 

OnJune 20, 1995 theTechnicalReportonthe Dr.Miller,thesecondprincipalreviewer,agreed 

toxicologyandcarcinogenesistudies of codeine with the proposed conclusions. She asked for clarifireceivedpublicreviewbytheNationalToxicology cationregardingthesignificanceofdose-related 

Program’s Board of Scientific Counselors’ Technical increases in sister chromatid exchanges occurring at 

Reports Review Subcommittee. The review meeting concentrations that caused cell cycle delay in cultured 

was held at the National Institute of Environmental Chinesehamsterovarycells.Dr.Dunnicksaidshe 

Health Sciences, Research Triangle Park, NC. would clarify these findings (page 56). 

Dr.Vodicnik,thethirdprincipalreviewer,agreed Dr. J.K. Dunnick, NIEHS, introduced the toxicology with the proposed conclusions. She was pleased that and carcinogenesis studies of codeine by discussing toxicokinetic data were included (Appendix M). thechemical’susesandtherationale for study, 

describingtheexperimentaldesign,reportingon Dr. A. Turturro, NationalCenter for Toxicological survival and body weight effects, and commenting onResearch,asked ifsomequantificationcouldbe compound-relatednonneoplasticlesionsinmice. addedrelatinglowerbodyweightswithdecreased Dr. Dunnick also reported on toxicokinetic studies in incidencesofmammaryglandandadrenalgland rats and mice performed in collaboration with investi- neoplasmsinrats. Dr. J. Haseman, NIEHS, re- gatorsBurroughsWellcome.proposed at The sponded that incidences of adrenal gland neoplasms conclusions for the 2-year studies were noevidence were decreased at all three concentrations, including of carcinogenicactivity in male or female F344/N two that had no body weight differences. However, rats or in male or female B6C3F, mice. he said that decreased incidences of mammary gland 

neoplasms could in part be explained by lower body 

Dr.Taylor,aprincipalreviewer,agreed withthe weight, and perhaps some quantification of the extent 

proposed conclusions. He observed that as noted in of this association could be added (page59). 

the Technical Report, the analgesic actionof codeine 

depends on 0-demethylation to morphine, a reaction Dr.VodicnikmovedthattheTechnicalReporton 

that is mediated in humans by cytochrome Pdso IID6. codeine be accepted with the revisions discussed and 

Heprovidedadditionalreferencesandsaidthe withtheproposedconclusionsaswritten for male 

discussion on human metabolism of codeine should andfemaleratsandmice, no evidence of carcinobeexpanded.Dr.Dunnicksaidtheadditional genic activity. Dr.Millersecondedthemotion, 

references would be included. which was accepted unanimously with nine votes. 
13 

INTRODUCTION 

CODEINE 

CAS NO. 76-57-3 

ChemicalFormula: C18HZ1N03 Molecular Weight: 299.36 

Synonyms: 7,8didehydro-4,5-epoxy-3-methoxy-17-methylmorph~-6-ol; methylmorphine;3-0-methylmorphinemonohydrate; 

N-methylnorcodeine; morphine-3-methylether; morphine monomethyl ether 

Trade names: Codeinum, Codicept, Coducept, Metilmorfii 

CHEMICAL AND PHYSICAL PROI%RTlFS 

Codeineisabittertasting,odorless,colorless to 

white material that occurs as a powderor crystallizes 

as monohydrate into orthorhombic, sphenoidal rods 

or octahedral tablets. It has a specific gravity of 1.32 

(20/4" C), a boiling point of 250" C, and a melting 

pointof 154" to 156" C;itsublimesat 140" to 

145" C at a pressure of 1.5 mmHg.Codeineis 

slightly soluble in water(1: 120),sparingly soluble in 

ether (1 :18), very soluble in chloroform (13 , and 

freely soluble in alcohol (1:2), and has a pK of 6.05 

at 15" C (Haley, 1983; MerckZnda, 1989; RTECS, 

1991;Willette, 1991). 

PRODUCTION, USE, 

AND HUMANEXPOSURE 

Codeineisthemostwidelyused ofthemany 

alkaloids that occur naturally in opium, the air-dried 

milky ofpoppy exudate plant the (Papaver

somniferum). It belongs to the group ofphenantoosmalltobeofcommercialimportance;yields 

vary from 0.8% to 3.5 %, depending on the country 

of origin (Swinyard, 1980). Commercialcodeineis 

prepared from morphine by methylating the phenolic 

hydroxylgrouporfromthebainebyappropriate 

reductionanddemethylation(Tyler et al., 1981; 

Willette, 1991). 

Codeine and its phosphate and sulfate salts are used 

in a variety of pharmaceuticals as analgesics, sedatives, antiperistaltics,antitussive hypnotics, and 

agents. In 1981, approximately 58 millionprescriptions for codeine-containing analgesicswere 

writtenintheUnitedStates,makingcodeine analgesics the most 'prescribed class of therapeutic drugs 

(DHHS, 1982). In 1993, combinations of codeine 

andcodeinephosphatewithacetaminophenwere 

amongthetop 100 prescriptiondrugsdispensedin 

theUnitedStates(AmericanDruggist, 1994). The 

American Hospital Formulary ServiceDrug Information (AHFS, 1995) lists two productscontaining 

codeine and 

thebaine. As aconstituent of opium, codeinehas 88 containing codeine phosphate. Codeine phosphate 

beenused for centuries; ithasbeenavailable as a is the most widely used form of codeine because of 

pure alkaloid since the middle ofthe 19th century. itsgreaterwatersolubility(Shanahan et al., 1983; 

Codeine is extracted from opium in amounts usually Hunskaar and Dragsund, 1985). 

threne derivatives, which also includes morphine andcodeine,threecontaining sulfate, 
14 Codeine, NTP TR 455 

For adults, the usual prescribed analgesic dosage is 

15 to 60 mgtakenorallyevery 4 hours, and the 

usualprescribedantitussivedosageis 10 to 20 mg 

every 4 to 6 hours;thepeakantitussiveffect 

usuallyoccurswithin 1 to 2 hours(AHFS, 1995). 

Thetypicaldailyadulttherapeuticdoseis 1 to 

4 mg/kgbodyweightperday or 40 to 160 mg/m2 

bodysurfacearea;thedailypediatricdoseis 

generally 3 mg/kgor 100 mg/m2in six equally 

dividedoral,subcutaneous,orintramusculardoses 

(JaffeandMartin, 1990; AHFS, 1995). Although 

thepotentialforabuseofcoughpreparationsand 

analgesics containing codeine led to their regulation 

under Schedule I1 of the Comprehensive Drug Abusequantifying codeine metabolism in various species are 

PreventionandControlActof 1970 (Swinyard, difficult to compare because the routes of administra1980;21 CFRParts 1300-1316), insomestates tion,thespecies,andtheroutesofrecoveryvary,

codeine is available without prescription as a compo-and the codeine conjugates are not always identified. 

nent of over-the-counter cough medicines containing 

up to 10 mgof codeine in an average5 mL dose. In 

otherstateshowever,thesecoughmedicinesare 

available only by prescription (Willette, 1991). 

AccordingtotheInternationalNarcoticsControl 

Board, 54,000 kgofcodeinewasmanufacturedat 

threeUnitedStatesfacilitiesin 1986; nospecific 

federal standards for workplace exposure to narcotic 

dustssuchascodeineexist,andexposurecontrol 

defaults to the nuisance dust standard of10 mg/m3as 

an 8-hour, time-weighted average(Biagini e?al., 

1990). TheNationalOccupationalExposureSurvey 

estimatedthatthepotentialworkplaceexposureto 

codeine,codeinephosphate,andcodeinesulfate 

combinedduring 1981 through 1983 was 36,000 

workers, 33,000 of whom were women in the health 

care services. The majority of the workers, 31,000, 

were potentially exposed to codeine phosphate, 1,000 

tocodeine,andlessthan 6,OOO tocodeinesulfate 

(NIOSH, 1990). The Drug Abuse Warning Network 

reported 2,482 emergency room mentions of codeine 

abuse from January 1985 to September 1988 (Cone 

et al., 1991). 

N-demethylationtonorcodeine,andconjugationof 

morphine and norcodeine with glucuronic acid (Adler 

andLatham, 1950; Phillipson e?al., 1978) 

(Figure 1). Studieswith rats, mice, dogs, hamsters, 

guinea pigs, rabbits, cats, and monkeys have shown 

that, while qualitatively the metabolic pathways are 

similaramongthespecies,therearequantitative 

differences in the amount of each metabolite formed 

(Adler and Latham, 1950; Woods e? al., 1956; Yeh 

andWoods, 1970,1971; BechtalandSinterhauf, 

1978; Phillipson e?al., 1978; Cone e?al., 1979; 

Suzuki e?al., 1988; Oguri e?al., 1990; Yuan e?al., 

1994; AppendixM).However,publishedstudies 

BiologicaldispositionstudiesbyYehandWoods 

(1969) demonstrated that 74% of a 2 mg/kg subcutaneousdoseofN-['4C]-methyl-codeine(base)was 

excretedbymale rats Sprague-Dawleywithin 

24 hours as free codeine, free morphine, and conjugatedmorphineviapulmonary,biliary,intestinal, 

and urinary routes.Infemalerats, 86% was 

excretedviathesameroutes.Hepaticclearanceof 

codeineaccounted forabouthalfofthesystemic 

clearance, indicating significant extrahepatic elimination. Only 2% of the administered radioactivity was 

retained in the body 24 hours after injection. 

Morphine formed by metabolic conversion of codeine 

hasbeenfoundinplasma,urine,bile,andfeces 

(J6hannessonandWoods, 1964; YehandWoods, 

1969; Yoshimura e?al., 1970) and in thebrain of 

rats(J6hannessonandSchou, 1963; Dahlstromand 

Paalzow, 1976; Chen e?al., 1990). Morphine in the 

brainoriginatesprimarilyby0-demethylationof 

codeineintheliver.However,codeinecrossesthe 

blood-brainbarrierfaster than morphinedue to its 

greater lipid solubility (Jaffe and Martin, 1990) and 

is also 0-demethylated to morphine within the brain 

(Chen e?al., 1990). 

PHARMACOKINETICS,ABSORPTION, 

DISTRIBUTION, METABOLISM, 

AND EXCRETION 

Experimental Animals 

Codeinemetabolizedby with is conjugation 

glucuronic 0-demethylationmorphine, acid, to 

Interspeciesdifferencesinconjugationofcodeine 

with glucuronic acid, 0-demethylation to morphine, 

and N-demethylation to norcodeine in vivo appear to 

originate from different activities of the microsomal 

enzyme,uridine diphosphate-glucuronyltransferase 

(UDPGT)andcytochromeP450.Phillipson e? al. 
Introduction 15 

Odemethylation Memethylation / Conjugation 

I\ 

I 

Conjugation 

COOH YQ 

f Y 6-giucuronide 

Morphine-3-glucuronide HO Norcodeine-&giucuronide

Morphine-6-glucuronide OH 

FIGURE 1 

Metabolic Pathways of Codeine in Rats (Findlay et al., 1977) 

(1978) demonstrated conversion of codeine into 

norcodeine (10%), morphine (8%), normorphine 

(2%), and codeine-N-oxide (20%) by liver microsomal preparations from guinea pigs; 60% of the 

dose was recovered as unchanged codeine. Hanioka 

et al. (1990) reported that UDPGTactivity (as 

measured by conjugation of codeine/metabolite) was 

higher in the liver microsomal preparations from 

guinea pigs (2.87 nmol/min per mg protein) and 

rabbits (2.34 nmol/min per mg) than that from rats 

(0.32 nmol/min per mg) or mice (0.30 nmol/min per 

mg).AlthoughtheUDPGTactivityinmice was 

similar to that of rats, urinary excretion of codeine 

glucuronide by mice was nine-fold higher. 

The 0-demethylation of codeine to morphine is a 

function of a polymorphic cytochrome P450 enzyme 

(CYPP,,,IID6 in manandCYPP,JIDlin rat) 

(Mikus et al., 1991, 1994). The female Dark-Agouti 

rat is an animal model for the human poor metabolizer phenotype (individuals who are poor hydroxylators of debrisoquine),while the Sprague-Dawley rat 

is a model for the human extensive metabolizer 

phenotype (individuals who readily metabolize both 

debrisoquineandcodeine). In an in vitro studyof 

codeine 0-demethylation by liver microsomes, the 

intrinsiclearanceofcodeine to morphine was 

tenfoldlower in female Dark-Agouti rats than in 

female Sprague-Dawley rats (Mikus et al. , 1991). 
16 Codeine, NTP TR 455 

The pharmacokinetics of codeine have been shown to 

be linear in a dose-ranging study inmaleSpragueDawley rats administered single intravenous bolus 

dosesof 1,1.5, 3,or 4 mg codeine phosphate/kg 

(Shah and Mason, 1991). A 3 mg/kg bolus dose of 

codeine phosphate delivered intravenously to male 

Sprague-Dawley rats was rapidlymetabolizedand 

excreted (Shah and Mason, 199Oa). In addition, the 

meanvolumeofsystemic distribution was greater 

than the total blood volume, suggesting that codeine 

sequesters out of the blood into various organs and 

tissues like most other organic bases. Following oral 

administration of codeine phosphate, the half life of 

codeine or morphine in rat plasma is estimated at one 

hour (Shah and Mason, 199Oa). Enterohepatic

recirculation may cause secondary morphine peaks 

60 minutes or more after administration of codeine 

phosphate in the rat (Shah and Mason, 1991). Up to 

30% of intravenously administered morphine undergoes enterohepatic recirculation in the rat (Walsh and 

Levine, 1975). The mean time residence of

morphine in the body is more than two times that of 

codeine (J6hannesson and Woods, 1964; Spector, 

1971). 

InF344/N rats, the bioavailability ofcodeinewas 

approximately 25% when administered in feed at a 

concentration of 1,600 ppm (Yuan et al., 1994; 

Appendix M), andthebioavailabilityofcodeine 

phosphate was approximately 8%when administered 

by gavage at 5 mg/kg to Sprague-Dawley rats (Shah 

and. Mason, 199Oa). Bioavailabilityofcodeine 

increasedwith increasing concentration in feed. As 

the F344/N rats aged, exposure resulted in decreased 

plasma concentrations of codeine and increased total 

Appendix M) . morphine concentrations (Yuan et al., 1994; 

The metabolic pathwaysofcodeineinhumans 

Humans 

are 

similar to those in the rat - i.e., partial conjugation 

with glucuronic acid, 0-demethylation to morphine, 

andN-demethylation to norcodeine;themethyl 

moiety is then converted to CO, via formaldehyde 

andexhaled (Adler ef al., 1955).However, after 

oral administration, 0-demethylation to morphine is 

a minor pathway in humans, as evidenced by an 

AUC,,h,:AUC,,,,e ratio of approximately 0.03 in 

humans (Shah and Mason 199Ob)compared to a ratio 

of 0.9 in rats (Shah and Mason, 1991). The mean 

codeine C, achieved in healthy humans following a 

60 mg oral dose of codeine phosphate (88 ng/mL) 

(Shahnd Mason, 199Ob) or codeine sulfate

(173 ng/mL) (Guay et al., 1988) were comparable to 

the C, in F344/N rats exposed to 800 ppm codeine 

(estimated to deliver 30 to 50 mg/kg per day) in feed 

(128 ng/mL) (Yuan ef al., 1994; Appendix M). Peak 

plasma concentrations for morphine in humans given 

an oral dose of 60 mg codeine phosphate were lower 

than that of rats exposed to 800 ppm of codeine in 

feed (Yuan et al., 1994; Appendix M). 

The major metabolic pathway of codeine in humans 

isconjugationwith glucuronic acid to codeine-6- 

glucuronide. In a studywithealthymaleand 

female volunteers given a single oral dose of 30 mg 

codeine phosphate (22 mg codeine base), the half-life 

of codeine was 1.8 hours and itwasdetected in 

plasma for 5 hours; the half-life of codeine-6-

glucuronide was 3 hours and it was detected in the 

plasma for at least 15 hours. Following an oral dose 

ofcodeinephosphate (using AUC analysis), it is 

estimated that in plasma 90% of the dose is codeine6-glucuronide, 4% is codeine, and 6% is morphine 

or other metabolites (Vree and Verwey-van Wissen, 

1992).Other researchers have also shown that the 

main metabolite of codeine in humans is codeine-6- 

glucuronide (Chen et al., 1991; Yue et al., 1991a). 

In humans, codeine is readily absorbed after oral 

ingestion, injection, or rectal administration, and the 

half-life in plasma is 2 to 4 hours (Jaffe and Martin, 

1990).Following oral or subcutaneous administration, peak concentrations in plasma and the brain are 

attainedwithin 15 to 30 minutes, which correlates 

with the onset of action (AHFS, 1995). Since 

codeine is a lipophilic opioid, it is absorbed transdermallyand from the nasal and buccal mucosa. It 

is distributed in the lung, liver, kidney, and spleen 

(Jaffe and Martin, 1990). 

In a study of the biological disposition of codeine 

phosphate in humans, 10% of the dose was excreted 

unchanged, 40% as conjugated codeine, 10% as 

norcodeine, and 10% as morphine (Parke, 1968). 

Urinary excretion profiles of four healthymale 

volunteers with a history of heroin abuse showed free 

codeine (7%), conjugated codeine (38%), free 

morphine ( < 1%), and conjugated morphine (4%) 

(Cone et al., 1991). Most of the free and bound 
Introduction 17 

codeine (95%), free morphine (88%), and bound 

morphine (70%) was excretedwithin 24 hours. In 

another study, both unchanged and codeine 

norcodeine were found in the urine of healthy male 

volunteers given single 30 mg oral doses of codeine. 

As in the guinea pig, hydrocodone and its metabolites, 6a-hydrocodol, 6P-hydrocodo1,nor- ad

hydrocodone, were also detected in the urine (Cone 

et al., 1979). The main urinary metabolite identified 

in the Vree and Verwey-van Wissen (1992) study of 

eight volunteers codeine-6-glucuronide, was 

accounting for 81.8% of the dose; 4.4% of the dose 

wasexcreted as free codeine, 0.6% as morphine, 

2.1 % as morphine-3-glucuronide,and 0.8% as 

morphine-6-glucuronide. 

Interindividual variations in the ability to glucuronidate codeine, related to UDPGT activities,have 

been demonstrated bothin vivo and in vitro in human 

liver and kidneymicrosomes(Yue et al., 1989a,b, 

1990). Furthermore, interethnic differencesinboth 

the glucuronidation and demethylationofcodeine 

have been noted in a number of studies, in particular 

thoseofYue et al. (1989a, 1991b) comparing 

SwedishCaucasianswithHanChinese.Caucasians 

excreted significantly less codeine and morphine and 

significantly more codeine-6-glucuronide and

morphine-6-glucuronide; the glucuronidation efficiency of Chinese subjects was consistently lower. 

Studieshaveshownthat the analgesic activity of 

codeine depends to a large extent on o-demethylation 

to morphine, a reaction that is mediated bycytochrome P450 IID6. The analgesic effect may be 

diminished in segments of the population that are 

poor metabolizers of codeine or are exposed to other 

environmental factors that affect codeine metabolism 

(Mortimer et al., 1990;Desmeules et al., 1991; 

Sindrup et al., 1991). 

Codeinehasbeenidentified in human breastmilk. 

Codeine concentrations in the milk of two lactating 

women given single oral doses of a compound analgesiccontaining 60 mgof codeinephosphatewere 

1.5 to 2.4 times as high as codeine concentrations in 

maternalplasma at the sametimeintervalsafter 

administration (Findlay et al., 1981). Metabolically 

producedmorphine levels in breastmilkwerelow 

but exceeded those in maternal plasma after 1 hour. 

Codeineand morphine were alsodetectedinthe 

plasma of newborns 1 to 4 hours after nursing when 

their mothers received a single oral dose of 60 mg 

codeine sulfate 1 hour before nursing (Meny et al., 

1993). Codeine plasma levels ranged from less than 

0.8 to 4.5 ng/mL, and morphine plasma levels 

ranged from less than 0.5to2.2 ng/mL. Free 

codeine levels in breast milk 20 to 240 minutes after 

administration ranged from 33.8 to 3 14 ng/mL and 

free morphine levels ranged from 1.9 to 20.5 ng/mL. 

PHARMACOLOGY 

Codeinehaspharmacologiceffects on the central 

nervoussystem (e.g., analgesia, drowsiness, mood 

changes,respiratory depression, nausea, and dysfunction of the endocrineandautonomicnervous 

systems) and the gastrointestinal tract (e. g .,decreased 

gastrointestinal motility). Alkaloids of this type tend 

to cause physical dependence, tolerance, and addiction (Jaffe and Martin, 1990). These pharmacologic 

effects (and recent research to understand the mechanisms involved) have been summarized (Almeida and 

Shippenberg,1991; Herz, 1993a,b). 

Opioid receptors belong to a larger family of receptors that share the ability to modify the function of 

membraneenzymes or ion channelsbyinteracting 

with one or more guaninenucleotide regulatory 

proteins in the cell membrane (Costa et al., 1991), 

and the amino acid sequences of rat opioid receptors 

havebeendeducedbycloningandsequencing 

cDNAs(Evans et al., 1992;Kieffer et al., 1992; 

Chen et al., 1993; Fukuda et al., 1993). 

Thegeneralpharmacologicactionofcodeine is 

similar to that of morphine; however, its analgesic 

potency,degreeof respiratory depression, toxicity, 

andpotential for addiction are less thanthoseof 

morphine. In rats, subcutaneous dosesof 60 mg 

codeinephosphate/kgandintraperitoneal doses of 

30mg/kgwereequianalgesicwith 5 and 10 mg 

morphine sulfate/kg, respectively; depending on the 

route of administration, the relative analgesic potenciesofcodeineandmorphinevaried as much as 

fourfold (Jbhannesson and Woods, 1964). However, 

the bioavailability of orally administered codeine in 

man is about 60%whereas that of morphine is about 

25% (Jaffe and Martin, 1990). Codeine undergoes 

lessfirst-passmetabolisminthe liver than does 

morphine,resultingin an oral-parenteral potency 

ratio of 1: 1.5 (Wallenstein et al., 1967). 
18 Codeine, NTP TR 455 

Codeine has been used in the treatment of pain for 

more than a century; about 60%of the clinical useof 

codeine is for pain relief (Shah and Mason, 199Oa). 

The binding affinity of codeine to opioid receptors is 

relativelyweak, approximately 1/3,000 that of 

morphine (Chen et al., 1990), and it is thought the 

analgesic effect of codeine is due to its biotransformation to morphine. Codeine exerts a drying 

effect on respiratory tract mucosaandincreases 

viscosityof bronchial secretions (AHFS, 1995). 

Along with other opiates, codeine causes the release 

ofhistamine through degranulation of mastcells, 

whichmay cause peripheral vasodilatation with 

related hypotension (Casale et al., 1984). 

TOXICITY 

Experimental Animals 

For rats, the oral LD,, is 427 mg codeine/kg body 

weight, the intraperitoneal LD50 is 130 mg/kg, the 

subcutaneous LD50 is 229 mg/kg, and the intravenous 

LD,, is 75 mglkg; for mice, therespective LD,, 

values are 250, 60, 84, and 54 mg/kg. The intramuscular LD,o in mice is 290 mg/kg. The intravenous LD50 in dogs is 69 mg/kg, and in rabbits it is 

34 mg/kg (RTECS, 1991). 

Induction of physicaldependence on codeine has 

been demonstrated in male Sprague-Dawley rats 

given 0.5 mg codeine/g food. Abruptcodeine 

withdrawal in rats treated for more than 2 days was 

associatedwithbodyweight loss, diarrhea, ptosis, 

and vocalization, which increased in intensity as the 

duration of codeine treatment increased(Suzuki 

et af., 1984). In a study of three inbred strains of 

mice, the severity of withdrawal signs precipitated by 

the opioid antagonist indicated naloxone that

C57BW6 mice developed greater physical dependence on codeine in feed thandid C3H/He and 

DBA/2 mice (Suzuki et af., 1991). In addition, the 

role of genotype in determining the degree of preference for morphine and codeine was demonstrated 

in a study in which Lewis rats (a strain derived from 

Sprague-Dawley rats) displayed a significantly greater 

preference for both codeine and morphine in feed 

than did F344 rats (Suzuki et al., 1988). 

Codeinecausescytotoxicityinisolated rat hepatocytes as measured by a time- anddose-dependent 

leakageoflactatedehydrogenase(Ellingtonand 

Rosen, 1987). At concentrations of0.5 or 1.25 mM, 

celldeathbegan after 60 minutesandviability 

decreased to less than 10% after 120 to 150 minutes. 

Hepatotoxicitywasinhibitedbythe addition of 

metyrapone, an inhibitor of cytochrome P450 metabolism, indicating that the cytotoxicity was caused by a 

P4,0-generated metabolite of codeine. 

Humans 

In sporadic cases, therapeutic doses of opioids may 

be associated with side effects, including respiratory 

depression, nausea, vomiting, dizziness, mental 

clouding, dysphoria, constipation, increased pressure 

inthebiliary tract, urinary retention, and hypotension. Acuteopioidtoxicity, circulatory collapse, 

respiratory paralysis, coma, and death have been 

reported from overdosage. The exact amount of any 

opioid that is toxic or lethal in humans has not been 

determined, although older literature suggests that a 

normal, pain-free adult is not likely to die after oral 

morphine doses of less than 120 mg, or to have 

serious toxic effects withless than 30 mg parenterally 

(Jaffe and Martin, 1990). 

Opioids have been implicated in allergic phenomena 

andhypersensitivityreactionsrelated to histamine 

release and manifested as itching, swelling, urticaria 

and other types of skin rashes such as fixed eruptions, exfoliative dermatitis with eosinophilia, 

pruritus, and scarlatiniform rashes (Feldberg and 

Paton, 195 1; Jaffe and Martin, 1990). Wheals at the 

site of codeine or morphine injection are also probably related to release of histamine (Shanahan et al., 

1983; Hunskaar and Dragsund, 1985). 

Rare anaphylactoid reactions have beenreported after 

intravenous administration of codeine and morphine, 

and three cases of severe adverse reactions to intravenous codeine phosphate reported in children were 

attributed to histamine release and thought to be dose 

related (Shanahanet af.,1983). These three children 

had no adversereaction to codeine administered 

intramuscularly. In three adults undergoing surgical 

anesthesia, hypotension life-threatening followed 

intravenous bolus injections of 10 to 60 mg codeine 

phosphate (Parke et al., 1992). Because the amount 

of histamine released from mast cells depends on the 

speed as well as the dose, the reactions were attributed to the fact that the dose was administered as a 

bolusinjection.Hypotension,which is increased 
Introduction 19 

undergeneralanesthesia,resultsfromvasodilation thefetusesand 19% of the litters in the 150 mg/kg 

and peripheral pooling of blood, and appears after a hamstergroup.Theoralno-observable-adverselatencyof15 to 20 seconds, atypicalfindingwith effect-level (NOAEL) for developmental toxicity was 

histaminereleasers(FeldbergandPaton,1951). 10 mg codeine/kg in hamsters and 75 mg/kg in mice; 

Codeine is amorepotenthistaminereleaserin theNOAEL for maternal toxicitywas 50 mg/kg in 

equianalgesic doses than morphine, and the Americanhamsters and 150 mg/kg in mice. 

MedicalAssociation’sDrugEvaluationsstatethat 

codeineshouldnot be administered intravenously 

(AMA,1986). 

Opioids should be used with caution in patients with 

hepatic disease, since increased bioavailability after 

oral administration or cumulative effects may occur. 

Renal disease greatly alters the pharmacokinetics of 

opioids(Barnes et al., 1985;Matzke et al., 1986); 

cases have been reported of renal failure and severe 

narcosisassociatedwithcodeineadministrationor 

abuse (Murray, 1973; Park et al., 1989; Hill et al., 

1991).active morphine-6- Themetabolite, 

glucuronide, may accumulate during repeated administration of codeine to patients withimpairedrenal 

function, resulting in symptoms ofopioidoverdose 

includingprofoundnarcosis(Matzke et al., 1986, 

Barnes et al., 1985; Hill et aZ., 1991). 

REPRODUCTIVE 

AND DEVELOPMENTALTOXICITY 

Experimental Animals 

Oraladministration of codeine a to twiceday 

hamstersandmiceduringmajororganogenesishas 

been shown to result in developmental toxicity in the 

presence and absence of maternal toxicity. The doses 

atwhichtheseeffectsareseenareusuallyseveral 

times higher than estimated human exposures when 

codeine is prescribed (Williams et al., 1991). 

Administration of codeine in water to LVG Syrian 

hamsters (0, 10, 50, or 150 mg/kg)twice aday on 

gestation days 5 through 13 and Swiss CD-1 mice(0, 

37.5, 75, 150,or300mg/kg)ongestationdays6 

through 15 increased thepercentage of resorptions 

per dam, and the number of dams with 100% resorptionswasincreasedin150mg/kghamstersand 

300mg/kgmice;however,thenumbers of live 

fetuses per litter wereunaffected(Williams et al., 

1991).Meanfetalbodyweightsweresignificantly 

Rats given 10, 35, or 120 mg codeine base/kg body 

weightorallyongestationdays6to15andrabbits 

given 5, 12.5, or 30 mg/kg on gestation days 6 to 18 

showednoteratogeniceffects,but120mg/kg was 

embryotoxictoratswhengivenatthetime of 

implantation (Lehman, 1976). 

A 50% reduction inserumtestosteronelevel was 

observed in male Sprague-Dawley rats 4 hours after 

a single subcutaneous injectionof 8.6 mg codeine/kg 

(Cicero, 1977). Male rats administered subcutaneous 

injections of morphine sulfate at 50 mg/kg per day 

for up to 9 weeks showed decreased serum luteinizinghormoneandtestosteronelevelsandreduced 

seminalvesicleandprostateglandweights;these 

effects were reversed within the 13-week withdrawal 

period (James et al. , 1980). 

Codeine was not teratogenic in the chicken embryo 

but was shownto beateratogeninrodentsafter 

subcutaneous injection (Ancel and Scheiner, 195 1). 

InLakeview goldenhamsters,single outbred a 

subcutaneous injection of codeine phosphate (73 mg 

codeine base/kg) on day8 of gestation caused cranioschisisin 6% of 12-day-oldfetuses(Geberand 

Schramm, 1975). Administration of 1 10 mg codeine 

phosphate/kg body weight to JBT/Jd mice on day 9 

of gestation causedhydrocephalicdilation of the 

fourth brain ventricle in 15 % of 13-day-old fetuses 

and was much less teratogenic than heroin (Jurand, 

1980).InCF-1albinomice,subcutaneousinjection 

of 100 mg/kg codeine sulfate on gestation days 8 and 

9 produced delayed ossification of the supraoccipital 

bone,paws,xiphoidandsternebraealongwith 

checkerboard and in sternebraepolysternebrae 

18-day-old fetuses (Zellers and Gautieri, 1977). 

Humans 

There have been several studies on the reproductive 

and developmental toxicity of codeine in humans, but 

decreased in 50 and 150 mg/kg hamsters and 150 andthe interpretation, of these studies is complicated by

300mg/kgmice.Althoughnotstatisticallysignifi- many factors, including multiple drug intake and the 

cant, meningoencephalocele was observed in 3% of clinicalconditionrequiringcodeinetreatment. For 
20 Codeine, NTP TR 455 

example,thecollaborative project perinatal of 

Heinonen et al. (1977) didnotshowanyincreased 

risk of malformationfromanalgesic or antipyretic 

use of codeine,whileothershavereportedan 

associationbetweenmaternalintake of opiates, 

primarily codeine, and cleft lipand palate in children 

(Sax&, 1975; LourwoodandRiedlinger, 1989). 

Opiates cross the placental barrier (Villee,1965) and 

arepresentintheneonatalbodyfluids(Waddell, 

1972) andarefoundinmaternalmilk(Welchand 

Findlay, 198 1). Somestudiessuggestthatcodeine 

In vivo, noincreaseinthefrequency of micronucleatedpolychromaticerythrocyteswasobserved 

in the bone marrow of female mice administered100 

to 500 mg/kg (252 to 1258 pmol/kg)codeine 

phosphate by intraperitoneal injection once a day for 

5 days and sacrificed4 hours after the final injection 

(BruceandHeddle, 1979). However,becauseonly 

1,000 polychromaticerythrocyteswerescoredper 

dose group(333 cells in each of three mice compared 

to the usual2,000 polychromatic erythrocytes in each 

of three to five mice per dose group), the validity of 

thesedataisquestionable. A second bonemarrow 

exposure can produce long-lasting effects on postnatal micronucleusstudywithmaleandfemale NMRI 

development (Mellin, 1964; Johnston, 1972;

Rothman et al., 1979; Sonderegger and Zimmerman, 

1979; LourwoodRiedlinger, and 1989). It is 

difficult to establish the role of codeine in producing 

congenital anomalies versus a variety of other drugs 

takeninaddition to codeine duringpregnancyand 

often without medical supervision. 

CARCINOGENICITY 

No information onthecarcinogenicpotentialof 

codeineinexperimentalanimalsorinhumans was 

found in the literature. 

GENETIC TOXICITY 

Codeinehashownlittleevidence ofmutagenic 

potentialinany of the standard in vitro or in vivo 

genotoxicity assays. It wasmutagenic not in

Salmonellatyphimurium (BruceandHeddle, 1979; 

King et al., 1979; Zeiger et al., 1992), Escherichia 

coli (King et al., 1979), oringermcells of 

Drosophila melanogaster (Nasrat et al., 1977; King 

et al., 1979). It did not induce chromosomal aberrations in cultured Chinese hamster ovary cells, but it 

did induce a significant increase in sister chromatid 

exchanges in cultured Chinese hamster ovary cells in 

boththepresenceandabsence of S9 metabolic 

activation enzymes (NTP, unpublished; AppendixE). 

Thereisonereport of increased frequencies of 

mice, at a lower total dose than that used by Bruce 

and Heddle (1979), also gave negative results (King 

et al., 1979). Inthisinvestigation,micewere 

administeredcodeinephosphateindosesup to 

0.25 mmol/kg by intraperitoneal injection or up to 

0.5 mmol/kg by gavage, once a day for 2 days, and 

sacrificed 6 hoursafterthefinaldose.Although 

1,000 polychromaticerythrocyteswerescoredin 

each of four mice per group, no data were presented. 

Becausetheoptimumtimeintervalbetweenfinal 

dosingandharvestisusually 24 hours,theshort 

intervalbetweenthetimesoffinaldosingandthe 

sacrifice times in these two studies effectively eliminated the potential effectsof the final doses from the 

analyses. 

In conclusion, the only indications of codeine-induced 

mutagenicityaretheresults of an in vitro sister 

chromatid exchange test in cultured Chinese hamster 

ovary cells that showed increases in sister chromatid 

exchanges both with and withoutS9. 

Morphine, a major metabolite of codeine, has been 

tested in a variety of mutagenicity assays. Morphine 

induced significant, dose-dependent increases in DNA 

damage (single-strand breaks, detected by single-cell 

microgel electrophoresis) in cultured human T-cells 

treatedwith 0.001 to 0.03 pg/mL for 1, 3, 4, or 

7 days(Shafer et al., 1994). In addition, a highly 

significant increase in the frequency of mutations at 

micronuclei in cultured rat kidney fibroblasts exposedthe HPRT locus of these human T-cells was observed 

to a combination ofcodeinephosphateandpara- aftertreatmentwith 0.01 pg/mL morphine for 

cetamol(Dunn et al., 1987); however, a similar 4 days. Knapp and Kramers (1976) tested morphine 

increasein micronucleatedcells was observed after in the Drosophila melanogaster sex-linked recessive 

treatmentwithparacetamolalone,suggestingthat lethalassayandfoundnoincreaseingermcell 

paracetamol was solely responsible for the increased mutati6ns in males. No increase in the frequency of 

frequency of micronucleated cells. chromosomalaberrationswasobservedinhuman 
Introduction 21 

cord blood treated lymphocxtes with 0.2 to

6.0 pg/mL morphine for 24, 48, or 68 hours (Falek 

et al., 1972); however, this report, which presented 

data from several studies, did not present data from 

this particular experiment. Badr and Rabouh (1983) 

reported the results of dominant lethal and spermatocyte cytogenetic tests in male mice administered 10 

to 60 mg/kg morphine. Totalembryoniclosses 

appearedmarkedlyincreasedweeks 1 to 3 posttreatment for all dose groups, indicating generalized 

toxicity to spermatids and mature sperm. It appears 

from their data that morphine may induce dominant 

lethalityinlate sperm cell stages.However,these 

data are difficult to interpret because control data 

from individual matingperiods(weeks)werenot 

presentedandit appears that mostoftheobserved 

effectsresulted from preimplantation losses (an 

endpointnotscoredintheanalysisofdominant 

lethality). The cytogenetic data from diakinesismetaphase I spermatocytes scored45 to 50 days after 

treatmentshowed extraordinary levels oftranslocationmultivalents, e.g., approximately 25% of 

cellsscored in the 40 and 60 mg/kg groups were 

aberrant. Such an observation impliesmorphineinduced chromosomal damagein spermatogonial stem 

cellstreatedmice. of Even the most potent

alkylating germ cell mutagens are not capableof 

inducingsuchdamage.Additional experiments are 

needed to confirm these results and, if confirmed, to 

further assess the germ cell of mutagenicity

morphine. 

STUDY k4TIONALE 

The National Cancer Institute and the Food and Drug 

Administration nominated codeine for study because 

it is a widely used drug and becauseit is a representativeofthemorphineclassof compounds, for 

which chronic carcinogenicity studieshadnotbeen 

conducted.Thefeed rather than gavage exposure 

route was selected to more closely approximate oral 

exposure in humans whencodeine is taken 3 to 4 

times per day. It was expected that animals would be 

able to tolerate higher exposure levels in feed than in 

oral bolus dosing, and 25,000 ppm wasselected as 

the high dose for rats in the 14-day study (estimated 

deliveryof 2,000 mg codeine/kg body weight per 

day), and 12,500 ppm was selected as the high dose 

for mice (3,000 mg/kg per day). 
22 Codeine, NTP TR 455 
23 

MATERIALS AND METHODS 

PROCUREMENT AND 

CHARACTERIZATION OF CODEINE 

CodeinewasobtainedfromPenickCorporation 

(Newark, NJ) in two lots and (471NFN003

471NINOOUA). Lot471NFN003 wasusedduring 

the14-dayand13-weekstudies,lot 471NINOOUA 

was used during the 2-year studies. Identity, purity, 

and stability analyses were conducted by the analytical chemistry laboratory, Midwest Research Institute 

(KansasCity,MO)(Appendix I). Reportson 

analyses performed in support of the codeine studies 

are on file at the National Institute of Environmental 

Health Sciences (NIEHS). 

Thechemical, a white powder,wasidentifiedas 

codeinebyinfrared, ultraviolethisible, and nuclear 

magnetic resonance spectroscopy. Purity of each lot 

was determined by elemental analyses, Karl Fischer 

wateranalysis,functional grouptitration, thin-layer 

chromatography, gas and chromatography.

Elementalanalyses for carbon, and hydrogen

nitrogenwereinagreementwiththetheoretical 

values for codeine when corrected for water content. 

Karl water indicated Fischeranalysis 

1.98% f 0.07% waterinlot471NFN003and 

4.66% f 0.05% waterinlot 471NINOOVA. Functionalgrouptitrationindicated a purity (anhydrous 

basis)of 100.0% f 0.5% for lot 471NFN003and 

99.8% f 0.8% for lot 471NINOOVA. Thin-layer 

chromatography for both lots indicated a major spot 

and no impurities (system 1) ora major spot and one 

traceimpurity(system 2). Gaschromatography 

usingsystem A indicatedonemajorpeakandone 

impurity with areas greater than 0.1 % relative to the 

major peak for both lots. The overall purity of each 

lot was determined to be at least 99% on an anhydrousbasis.Eachlot ofthechemical also met the 

United States PharmacopeiaXX standards for identity 

and purity. 

when stored protected from light at temperatures up 

to 25" C. To ensure stability, the bulk chemical was 

stored at approximately 20" C, in the dark, in sealed 

amber glass bottles in a secure cabinet. 

Stability was monitoredbythestudylaboratory 

during the lCday, 13-week, and 2-year studies using 

functional group titration and/or gas chromatography. 

No degradation of the bulk chemical was detected. 

PREPARATIONAND ANALYSIS 

OF DOSEFORMULATIONS 

Thedoseformulationswerepreparedweeklyby 

mixingcodeinewithfeed(Table11).Homogeneity 

and stability studies of a 160 ppm and a 30,000 ppm 

(homogeneity only) formulation were performed by 

theanalyticalchemistrylaboratoryusinghighperformanceliquidchromatography.Homogeneity 

was confirmed, and the stability of the formulations 

was confirmed for up to 3 weeks when stored in the 

dark at room temperature. 

Periodic analyses of the dose formulations of codeine 

were conducted at the study laboratory and analytical 

chemistrylaboratoryusinghigh-performanceliquid 

chromatography.Duringthe14-daystudies,formulations were analyzed at the beginning of the study 

(Table12).Forthe13-weekstudies,formulations 

were analyzed at the beginning, midpoint, and end of 

thestudy(Table13).Duringthe2-yearstudies, 

formulationswereanalyzedevery 6 to 10weeks 

(Table 14). Of the dose formulations analyzed for the 

rat study, 97% (86/89) were within 10% of the target 

concentration.Forthemousestudy, 100% (40140) 

were within 10% of the target concentration. Results 

of periodic referee analyses performed by the analyticalchemistrylaboratoryagreedwiththeresults 

obtained by the study laboratory (Table 15). 

Stability studies of the bulk chemical were performed 

by the analyticalchemistrylaboratoryusinggas 14-DAY STUDIES 

chromatography.studies that These indicated Male and female F344/N rats and B6C3F, mice were

codeine was stable as a bulk chemical for 2 weeks obtainedfromFrederickCancerResearchFacility 
24 Codeine, NTP TR 455 

(Frederick, MD). On receipt, the rats and mice were 

4 weeks old. Ratswerequarantined for 12 days 

(males) or 14 days (females) and mice were quarantined for 12 days; rats and mice were 6 weeks old on 

the first day of the studies. Before initiation of the 

studies, two male and two female rats and mice were 

randomly selected for parasite evaluation and gross 

observation for evidence ofdisease.Groupsof 

five male and five female rats werefediets 

containing 0, 1,562, 3,125, 6,250, 12,500, or 

25,000 ppm codeine. Groups of five male and 

five female mice werefed diets containing 0, 781, 

1,562, 3,125, 6,250, or 12,500 ppm codeine.Feed 

and water were available ad libitum. Rats and mice 

werehoused five per cage. Clinicalfindingswere 

recorded daily for rats and mice. Feed consumption 

was recorded daily by cage. The animals were 

weighed initially, after the first week, and at the end 

of the studies. Details of the study design and animal 

maintenance are summarized in Table 1. 

A necropsy was performed on all rats and mice. The 

brain, heart, right kidney, liver, lung, right testis, 

and thymus were weighed. Complete histopathologic 

examinations were performed on 0, 6,250 (female 

only), 12,500, and 25,000 ppm (maleandfemale) 

rats and 0 and 12,500 ppm mice.,Additionally,the 

stomach, testis, and thymus were examined in 1,562 

and 3,125 ppm male and female rats andin 

6,250 ppm male rats. Table 1 lists the tissuesand 

organs examined. 

ISWEEK STUDIES 

The 13-week studies ‘were conducted to evaluate the 

cumulative toxic effects of repeated exposure to 

codeine and to determine the appropriate doses to be 

used in the 2-year studies. 

Male and female F344/N rats and B6C3F1 mice were 

obtained from Simonsen Laboratories (Gilroy, CA). 

On receipt, the rats and micewere 4 weeksold. 

Animals were quarantined for 8 days(females) or 

12 days (males) and were 6 weeksold on the first 

dayof the studies. Beforeinitiation of thestudies, 

five male and five female rats andmicewere 

randomly selected for parasite evaluation and gross 

observation for evidence of disease. Attheend of 

the studies, serologic analyses wereperformed on 

five male and five female control rats ,and mice using 

the protocols of the NTP Sentinel Animal Program 

(Appendix L). 

Groups of 10 male and 10 female rats and mice were 

feddietscontaining 0, 390, 781, 1,562, 3,125,or 

6,250 ppm codeine.Feed and water were available 

ad libitum. Ratsandmice were housed five per 

cage.Clinical findings were recorded weekly for 

rats and mice.Feed consumption was recorded 

weekly by cage. The animals were weighed initially, 

weekly, and at the end of the studies. Details of the 

study design and animal maintenanceare summarized 

in Table 1. 

Atthe end of the13-week studies, samples were 

collected for sperm morphology and vaginal cytology 

evaluations from male andfemale rats and mice in 

the 0, 1,562,3,125, and 6,250 ppm groups. The 

parameters evaluated are listed in Table 1. Methods 

usedwerethose described in the NTP’s sperm 

morphologyandvaginal cytology evaluations protocol (NTP, 1983). For 7 consecutive days before 

the scheduled terminal sacrifice, the vaginal vaults of 

the females were moistened with saline, if necessary, 

and samples of vaginal fluid and cells were stained. 

Relative numbers of leukocytes, nucleated epithelial 

cells, and large squamous epithelial cells .were 

determined and used to ascertain estrous cycle stage 

(i.e., diestrus, proestrus, estrus, and metestrus). 

Male animals were evaluated for sperm morphology, 

count, and motility. The right epididymis and right 

testiswereisolatedandweighed. The tail of the 

epididymis(cauda epididymis) was then removed 

from the epididymalbody (corpus epididymis) and 

weighed.Testyolk (rats) or modified Tyrode’s 

buffer (mice) wasapplied to slides, and a small 

incision was made at the distal border of the cauda 

epididymis. The sperm effluxing from the incision 

weredispersedin the buffer on the slides, and the 

motile and nonmotile spermatozoa were counted for 

five fields per slide by two observers. Following 

completion of sperm motility estimates, each right 

caudaepididymiswasplaced in buffered saline 

solution. Caudae were finely minced, and the tissue 

wasincubated in the saline solution and then heat 

fixed at 65 O C. Sperm density was then determined 

microscopically with the aid of a hemacytometer. To 

quantifyspermatogenesis,the testicular spermatid 

head count was determinedby removing the tunica 

albugineaandhomogenizing the right testis in 
Materials and Methods 25 

phosphate-bufferedsalinecontaining 10% dimethylwasexaminedin 3,125 ppm female rats. Table 1 

sulfoxide. Homogenization-resistantspermatidnucleiliststhetissuesand organs routinely examined. 

werecountedwith a hemacytometer. Four sperm 

morphology slides were prepared for each animal 

evaluated.--An aliquot of killed sperm suspension was 

stained in a test tube, spread on a microscope slide, 

and examined. 

Atthe end of the13-week studies, rats andmice 

were anesthetized with a short-acting barbiturate aid 

blood was collected from the tails of all rats and mice 

for hematologyanalysesand from the xternal 

jugular vein for clinical chemistryanalyses.Blood 

for hematology determinations was placed in tubes 

containing potassium EDTA as the anticoagulant. 

Blood for serum analyses was collected in containers 

withoutanticoagulant,allowedto clot atroom 

temperature, and centrifuged, and serum was separated. Hematologyparametersweremeasured on a 

Baker Series 7000 Cell Counter (Baker Instrument 

Corp., Allentown, PA). Plateletcountsweredetermined on a Baker Series 810 PlateletAnalyzer 

(Baker Instruments Corp., Allentown, PA). Clinical 

chemistry parameters were measured on a Gemini 

centrifugal analyzer (Electro Nucleonics Inc., 

Fairfield, NJ). The hematologyclinical and 

chemistry parameters measured are listed in Table 1. 

Urinalysis studies were performed after week 12 of 

the studies; rats and mice were housed individually in 

metabolismcages for a 16-hourcollection period. 

Urine collection containers were immersed in an icewater bath during the samplingperiod to minimize 

evaporation and to suppress bacterial growth. Feed 

was withheld, but waterwasavailable ad libitum 

during the collection period. Urinevolumewas 

measured and specific gravity was determined with a 

refractometer (Table 1). 

A necropsy was performed on all animals. The 

adrenal glands, brain, heart, right kidney, liver, lung, 

right testis, thymus, andspleenwereweighed. 

Tissues for microscopic examination were fixed and 

preserved in 10% neutral buffered formalin,

processedand trimmed, embeddedin paraffin, 

sectioned to a thickness of 5 to 6 pm, and stained 

withematoxylinand eosin. A complete histopathologic examination wasperformed on 0 and 

6,250 ppm rats and miceandallmicethatdied 

before the end of the study. Additionally, theliver 

YEAR STUDIES 

study ~~~i~ 

Groups of 60 male and 60 female rats were fed diets 

con&ning 0, 400, 800, or 1,600 ppm codeine. 

Groups of 60 male and60 female mice were fed diets 

containing 0,750,1,500, or 3,000 ppm codeine. Ten 

maleand 9 or 10 female rats and mice from each 

group were evaluated at 15 months for histopathology 

and organ weights. 

Source and Specification of Animals 

Male and female F344/N rats and B6C3F, mice were 

obtained from Taconic Farms (Germantown, NY) for 

use in the 2-year studies. Rats were quarantined for 

15daysandmicewerequarantined for 13 days 

before the beginningofthe studies. Five male and 

five female rats and mice were selected for parasite 

evaluationand gross observation disease. of

Serology samples were collected for viral screening. 

Rats and mice were approximately6 weeks old at the 

beginning of the studies. The health of the animals 

was monitored during the studies according to the 

protocolsoftheNTPSentinelAnimal Program 

(Appendix L). 

Animal Maintenance 

Ratswerehoused five per cage and micewere 

housedindividually.Feedandwaterwereavailable 

ad libitum. Feed was consumptionmeasured 

monthlybycage.Cagesandracks were rotated 

every 2 weeks. Further details of animalmaintenance are given inTable 1. Information on feed 

compositionadcontaminants is provided in 

Appendix K. 

Clinical Examinations and Pathology 

Allanimalswereobservedtwice daily. Clinical 

findings were recorded every4 weeks. Body weights 

werecordedinitially,weekly for 13 weeks, 

monthly thereafter, and .at the end of the studies. 

A complete necropsyandmicroscopic examination 

wereperformed on all rats andmice.Atthe 

15-monthinterimevaluation necropsy, the adrenal 

gland, kidney, and liver of rats and mice were 

weighed. At necropsy, all organs and tissueswere 
26 Codeine, N"TR 455 

examined for grossly visible lesions, andallmajor 

tissues were fixed and preserved in 10% neutral 

buffered formalin, processed and trimmed, embedded 

in paraffin, sectioned to a thickness of 5 to 6 pm, 

and stained with hematoxylinand eosin for microscopic examination. For all paired organs (i.e., 

adrenal gland, kidney, ovary), samples from each 

organ are examined. Tissues micro- examined 

scopically are listed in Table 1. 

Microscopic evaluations were completed by the study 

laboratory pathologist, and the pathology data were 

entered into theToxicologyDataManagement 

System. The microscopic slides, paraffin blocks, and 

residual wet tissues were sent to the NTP Archives 

for inventory, slide/block match, andwettissue 

audit. The slides, individual animal data records, 

andpathology tables were evaluatedby an independent quality assessment laboratory. Theindividual animal records and tables were compared for 

accuracy, the slide and tissue counts were verified, 

and the histotechnique was evaluated. For the 2-year 

studies, a quality assessment pathologist reviewed the 

adrenal gland (cortex and medulla),lymphnodes 

(mandibular), forestomach, nose, and other tissues of 

interest for rats and thyroid gland, kidney, liver, and 

other tissues of interest for mice. 

The qualityassessment report andslideswere 

submitted to the NTP Pathology WorkingGroup 

(PWG) chair, who reviewed the selected tissues and 

any other tissues for which a disagreement in diagnosis between the laboratory and quality assessment 

pathologists existed. Representativehistopathology 

slides containing examples of lesionsrelated to 

chemical administration, examples of disagreements 

in diagnoses between the laboratory and quality 

assessment pathologist, or lesions of general interest 

were presented by the chair to the PWG for review. 

The PWG consistedof the quality assessment pathologist and other pathologists experienced in rodent 

toxicologic pathology. This group examined the 

tissueswithoutanyknowledgeof dose groups or 

previously rendered diagnoses. When the PWG 

consensus differed from the opinion of the laboratory 

pathologist, the diagnosis was changed. Thus, the 

final diagnoses represent a consensus of contractor 

pathologists and the PWG. Detailsofthesereview 

procedures have ben described, part, in by

Maronpot and Boorman (1982) and Boorman et al. 

(1985). For subsequent analysesof the pathology 

data, the diagnosed lesions for each tissue type were 

evaluatedseparately or combined according to the 

guidelines of McConnell et al. (1986). 

STATISTICALMETHODS 

Survival Analyses 

The probability of survival was estimated by the 

product-limit procedure of Kaplan and Meier (1958) 

and is presented in the form of graphs. Animals 

found dead of other than natural causes or missing 

were censored from the survival analyses; animals 

dying from natural causes were not censored. 

Statisticalanalyses for possible dose-related effects 

on survival used Cox's (1972) method for testing two 

groups for equality and Tarone's (1975) life table test 

to identify dose-related trends. All reported P values 

for the survival analyses are two sided. 

Calculation of Incidence 

The incidences of neoplasmsor nonneoplastic lesions 

as presented in Tables Al, A5, B1, B5, C1, C5, Dl, 

and D5 are given as the number of animals bearing 

such lesions at a specific anatomic site and the 

numberofanimalswith that site examined microscopically. For calculation of statistical significance, 

theincidencesofmostneoplasms (Tables A3, B3, 

C3, and D3) and all nonneoplastic lesions are given 

as number of animals affected at each site examined 

microscopically. However, when macroscopic 

examinationwas required to detect neoplasms in 

certain tissues (e.g., skin, intestine, harderian gland, 

and mammary gland) before microscopic evaluation, 

or when neoplasmshadmultiple potential sites of 

occurrence (e.g., leukemia or lymphoma), the 

denominatorsconsistof the number of animals on 

which a necropsy was performed. Tables A3, B3, 

C3, and D3 also give the survival-adjusted neoplasm 

rate for each group and each site-specific neoplasm, 

i.e., theKaplan-Meier estimate of the neoplasm 

incidence that would have been observed at the end 

of the study in the absence of mortality from all other 

competing risks (Kaplan and Meier, 1958). 

Analysis of Neoplasm Incidences 

The majority ofneoplasmsinthese studies were 

considered to be incidental to the cause of death or 

notrapidly lethal. Thus, the primary statistical 

method used was logistic regression analysis, which 
Materials and Methods 27 

assumed that the diagnosed neoplasmsweredis- testwas used, a procedure based on the overall 

covered as the result of death from an unrelated proportion of affected animals. 

cause and thus did notaffect the risk of death. In 

this approach, neoplasm prevalence was modeled as 

a logistic function of chemical exposure and time. 

Both linear and quadratic terms in time were incorporatedinitially,and the quadratic term was eliminatedifthe fit ofthemodel .was not significantly 

enhanced. The neoplasm incidences of exposed and 

control groups were compared on the basisofthe 

likelihood score test for the regression coefficient of 

dose. This method of adjusting for intercurrent 

mortality is the prevalence analysisof Dime and 

Lagakos (1983), further described and illustrated by 

Dime and Haseman (1986). Whenneoplasms are 

incidental, this comparison of thetime-specific 

neoplasm prevalences also provides a comparison of 

Analysis of Continuous Variables 

Twoapproacheswereemployed to assess the significance of pairwise comparisons between exposed 

and control groups in the analysis of continuous 

variables. Organ and body weight data, which have 

approximatelynormal distributions, were analyzed 

using the parametric multiple comparison procedures 

ofDunnett (1955) andWilliams (1971,1972). 

Clinical chemistry, hematology, spermatid, and 

epididymal spermatozoal data,which have typically 

skewed distributions, were analyzed using the nonparametric multiple comparison methods of Shirley 

(1977) and Dunn (1964). Jonckheere’s test

(Jonckheere, 1954) wasused to assess the signifithe time-specific neoplasm incidences (McKnight andcance of the dose-related trends and to determine 

Crowley, 1984). 

In addition to logistic regression, other methods of 

statistical analysis were used, and the results of these 

tests are summarized in theappendixes.These 

methodsincludethe life table test (Cox, 1972; 

Tarone, 1975), appropriate for rapidly lethal

neoplasms,and the Fisher exacttestandthe 

Cochran-Armitagetrendtest(Armitage, 1971; Gart 

et al., 1979), procedures based on the overall proportion of neoplasm-bearing animals. 

Tests of significance included pairwise comparisons 

of each exposed group with controls and a test for an 

overall dose-related trend. Continuity-corrected tests 

were used in the analysis of neoplasm incidence, and 

reported P values are one sided. Theprocedures 

described in the preceding paragraphs were also used 

to evaluate selected lesions. nonneoplastic For 

further discussion of these statistical methods, refer 

to Haseman (1984). 

Analysis of Nonneoplastic Lesion Incidences 

Because all nonneoplastic lesions in this study were 

considered to be incidental to the cause of death or 

not rapidly lethal, the primary statistical analysis used 

wasa logistic regression analysis in whichnonneoplastic lesion prevalence was modeled as a logistic 

function of chemical exposure and time. For lesions 

detected at the interim evaluation, the Fisher exact 

whether a trend-sensitive test (Williams’ or Shirley’s 

test) was more appropriate for pairwise comparisons 

than a test that does not assume a monotonic doserelatedtrend(Dunnett’s or DUM’S test). Prior to 

analysis, extreme values identified by the outlier test 

of Dixon and Massey(1951) were examined by NTP 

personnelandimplausiblevalueswereeliminated 

from the analysis.Averageseverity values were 

analyzed for significance using the Mann-Whitney U 

test(HollanderandWolfe, 1973). Becausethe 

vaginal cytology data are proportions (the proportion 

oftheobservationperiodthat an animal wasina 

given estrous stage), an arcsine transformation was 

used to bring the data into closer conformance with 

normality Treatment assumptions. effects were

investigatedbyapplyingamultivariateanalysisof 

variance (Morrison, 1976) to the transformed data to 

test for simultaneous equality of measurementsacross 

exposure levels. 

Historical Control Data 

Although the concurrent control group is always the 

first and most appropriate control group used for 

evaluation, historical control data can be helpful in 

theoverallassessment of neoplasmincidencein 

certain instances. Consequently, neoplasm incidences 

from the NTP historical control database (Haseman 

et al., 1984,1985) are included in the NTP reports 

for neoplasms appearing to show compound-related 

effects. 
28 Codeine, NTP TR 455 

QUALITY ASSURANCE METHODS 

The 13-week and 2-year studies wereconductedin 

compliance with Food and Drug Administration Good 

Laboratory Practice Regulations (21 CFR, Part 58). 

In addition, as records from the 2-year studies were 

submitted to the NTP Archives, thesestudieswere 

audited retrospectively by an independentquality 

assurance contractor. Separate audits covering

completenessandaccuracyofthepathology data, 

pathologyspecimens, final pathology tables, and a 

draft of this NTP Technical Report were conducted. 

Audit procedures and findings are presented in the 

reports and are on file at NIEHS. The audit findings 

were reviewed andassessedbyNTP staff, so all 

commentshadbeenresolved or wereotherwise 

addressed during the preparation of this Technical 

Report. 

GENETIC TOXICOLOGY 

The genetic toxicity of codeine was assessedby 

testing the ability of the chemical to induce mutations 

in various strains of Salmonellatyphimurium and 

sister chromatid exchangesand chromosomal 

aberrations in cultured Chinese hamster ovary cells. 

The protocols for these studies- and the results are 

given in Appendix E. 

The genetic toxicity studies of codeine are part of a 

larger effort by the NTP to develop a database that 

would permit the evaluation ofcarcinogenicityin 

experimental animals from the structure and 

responses of the chemical in short-term in vitro and 

in vivo genetictoxicitytests.These genetic toxicity 

tests were originally developed to study mechanisms 

of chemically inducedDNAdamageand to predict 

carcinogenicity in animals, based on the electrophilic 

theoryofchemical carcinogenesis and thesomatic 

mutationtheory (Miller and Miller, 1977; Straus, 

1981; Crawford, 1985). 

There is a strong correlation between a chemical’s 

potentialelectrophilicity (structural alert to DNA 

reactivity), mutagenicity in Salmonella, and carcinogenicityin rodents. The combination of electrophilicityand Salmonella mutagenicity is highly 

correlatedwith the induction of carcinogenicity in 

rats and mice and/or at multiple tissue sites (Ashby 

andTennant, 1991). Other in vitro genetic toxicity 

tests do not correlate well with rodent carcinogenicity 

(Tennant et al., 1987; Zeiger et al., 1990), although 

these other tests can provide information on the types 

of DNA and chromosome effects that can be induced 

by the chemical being investigated. Data from NTP 

studies show that a positive response in Salmonella is 

currently the most predictive in vitro test for rodent 

carcinogenicity (89% of the Salmonella mutagens 

were rodent carcinogens), and that there is no 

complementarity among the in vitro genetic toxicity 

tests. That is, no batteryoftests that included the 

Salmonella test improvedthe predictivity of the 

Salmonella test alone. The predictivity for 

carcinogenicity of a positive response inbone 

marrow chromosome aberration or micronucleus tests 

is not yet defined. 
Materials and Methods 29 

TABLE1 

Experimental Design and Materials and Methodsin the Feed Studies of Codeine 

14-Day Studies 

Study Laboratory 

EG&G Mason Research Institute 

(Worcester, MA) 

Strain and Species 

Rats: F344lN 

Mice: B6C3FI 

Animal Source 

Frederick Cancer Research Facility 

(Frederick, MD) 

Time Held Before Studies 

Rats:12 days (males) or 14 days 

(females) 

Mice:12 days 

Average Age When Studies Began 

6 weeks 

Date of First Dose 

Rats: 2 April (males) 

or 4 April (females) 1984 

Mice: 9 April (males) 

or 16 April (females) 1984 

Duration of Dosing 

14 days 

Date of Last Dose 

Rats: 16April(males) or 

18 April (females) 1984 

Mice:23April(males) or 

30 April (females) 1984 

Necropsy Dates 

Rats:16April(males) or 18April 

(females)1984 

Mice:23April(males) 

or 30 April (females) 1984 

Average Age at Necropsy 

8 weeks 

13-Week Studies 

EG&G Mason Research Institute 

(Worcester, MA) 

Rats:F344lN 

Mice:B6C3Fl 

Simonsen Laboratories 

(Gilroy, CA) 

Rats: 12 days (males) or 8 days 

(females) 

Mice:12 days (males) or 8 days 

(females) 

6 weeks 

Rats: 23October(males) or 

19 October (females) 1984 

Mice:13November(males) 

9 November (females) 1984 

13 weeks 

Rats: 23 or 25 January (males) 

16 or 18 January (females) 1985 

Mice: 13 or 15February(males) 

6 or 8 February (females) 1985 

Rats: 23 or 25 January (males) 

or 16 or 18 January (females) 1985 

Mice:13 or 15February(males) 

or 6 or 8 February (females) 1985 

19 weeks 

2-Year Studies 

Microbiological Associates, Inc. 

(Bethesda, MD) 

Rats: F344lN 

Mice:B6C3F1 

Taconic Farms 

(Germantown, NY) 

Rats: 15days 

Mice: 13days 

7 weeks 

Rats:14April1989 

Mice: 29 March 1989 

Rats: 105-106 wet:ks 

Mice:105-106weeks 

Rats: 19 April 1991 

Mice: 3 April 1991 

Rats: 15-Monthinterimevaluation -

12-13July1990 

Terminal -

15-19 April 1991 

Mice:15-Monthinterimevaluation -

27-28 June 1990 

Terminal -

27 March-3 April 1991 

15-Month interim evaluation -

72 weeks 

Terminal -11 1-1 12 weeks 
30 ' Codeine, NTP TR 455 

TABLE1 

Experimental Design and Materials and Methodsin the Feed Studies of Codeine (continued) 

14-Day Studies 

Sue of Study Groups 

5 males and 5 females 

Method of Distribution 

Animals of similar weight classes were 

assigned to cages using a computergenerated list of random numbers; 

cages were assigned to test groups 

using an additional computer-generated 

list of random numbers 

Animals per Cage 

5 

Method of Animal Identification 

Toe clip 

Diet 

NM-07 open formula meal diet 

(Zeigler Brothers, Inc., Gardners, PA), 

availablead libitum, changed weekly. 

WaterDistribution 

Tap water (Worcester municipal 

supply) via automatic watering system 

(Edstrom Industries, Waterford,WI), 

availablead libitum 

Cages 

See-Through I1 System (Lab Products, 

Rochelle Park, NJ), changed twice per 

week. 

Bedding 

Beta-Chip (Northeastern Products 

Corp., Warrensburg, NY), changed 

twice per week 

Cage Filters 

Non-woven fiber (Snow Filtration, 

Cincinnati, OH), changed once every 

two weeks' 

Racks 

See-Through I1 System Products, as (Lab Same 

Rochelle Park, NJ), changed once 

every two weeks 

13-WeekStudies 

10 males and 10 females 

Same as 14day studies 

5 

Toe clip 

Same as 14day studies 

Same as 14day studies 

Same as 14day studies 

Same as 14day studies 

Same as 14day studies 

14day studies 

2-Year Studies 

15-Month interim evaluation10 males and 10 females 

Terminal -50 males and 50 females 

Same as 14day studies 

Rats: 5 

Mice: 1 

Tail tattoo 

NM-07 open formula powdered diet 

(Zeigler Brothers, Inc., Gardners, PA), 

available ad libitum, changed weekly. 

Tap water (Washington Suburban 

Sanitary Commission, Potomac plant) 

via automatic watering system 

(Edstrom Indusmes, Waterford, WI), 

available ad libitum 

Same as 14day studies, except mice 

cages changed weekly. 

Sani-Chips (P.J. Murphy Forest 

Products, Montville, NJ), changed 

twice per week (rats)or once per week 

(mice) 

Spun-Bonded Polyester (Snow 

Filtration, Cincinnati, OH), changed 

once every two weeks 

Same as 14-day studies 
Materials and Methods 31 

TABLE1 

Experimental Design and Materials and Methods in the Feed Studies of Codeine (continued) 

14-Day Studies 

Animal Room Environment 

Temperature:, 21.1"to 23.9"C 

Relative humidity: 28%to 60% 

Fluorescent light: 12 hourslday 

Room air: at least 10 changes per hour 

Doses 

Rats: 0,1,562, 3,125, 6,250, 12,500, 

or 25,000 ppm in feed, available 

ad libitum 

Mice: 0,781, 1,562, 3,125, 6,250, or 

12.500 ppm in feed, available 

ad libitum 

Type and Frequency of Observation 

Observed twice daily; animals were 

weighed initially, after the first week, 

and at the end of the studies; clinical 

observations were recorded daily. 

Feed consumption was recorded daily 

by cage. 

Method of Sacrifice 

Sodium thiamylal asphyxiation 

Necropsy 

Necropsy performed on all animals. 

Organs weighed were brain, heart, 

right kidney, liver, lung, right testis, 

and thymus. 

Clinical Pathology 

None 

13-Week Studies 

Temperature: 20.5"to 25.0"C 

Relative humidity: 34%to 55%

Fluorescent light: 12 hourslday 

Room air: at least 10 changes per hour 

0, 390, 781, 1,562, 3,125, or 

6,250 ppm in feed, available ad libitum 

Observed twice daily; animals were 

weighed initially, weekly, and at the 

end of the studies; clinical observations 

were recorded weekly. Feed consumption was recorded weekly by cage. 

Carbon dioxide asphyxiation 

Necropsy performed on all animals. 

Organs weighed were adrenal gland, 

brain, heart, right kidney, liver, lung, 

spleen, right testis, and thymus. 

Blood was collected from the tail of all 

animals for hematology and from the 

external jugular vein for clinical chemistry. Urine was collected from 

animals housed individually in metabolism cages for 16 hours. 

Hematology: Hematocrit, hemoglobin, 

erythrocytes, reticulocytes, nucleated 

erythrocytes, mean cell volume, mean 

cell hemoglobin, mean cell hemoglobin 

concentration, platelets, and leukocytes 

and differentials 

Clinical Chemistry: urea nitrogen, 

creatinine, glucose, total protein, 

albumin, alkaline aminotransferase, 

alkaline phosphatase, and aspartate 

aminotransferase 

Urinalysis: specific gravity and 

volume 

2-Year Studies 

Temperature: 19.4"to 26.7" C (rats), 

20" to 25" C (mice) 

Relative humidity: 25%to 95%(rats), 

23 % to 94%(mice) 

Fluorescent light: 12 hourslday 

Room air: at least 10 changes per hour 

Rats: 0,400, 800, or 1.600 ppm in 

feed, available ad libitum 

Mice: 0,750, 1,500, or 3,000 ppm in 

feed. available ad libitum 

Observed twice daily and clinical 

observations were recorded every 

4 weeks; animals were weighed 

initially, weekly for 13 weeks, monthly 

thereafter, and at the end of the 

studies. Feed consumption was 

recorded monthly by cage. 

Carbon dioxide asphyxiation 

Necropsy performed on all animals. 

Organs weighed at the 15-month 

interim evaluation were adrenal gland, 

right kidney, and liver. 

None 
32 Codeine, NTF’ TR 455 

TABLE1 

Experimental Design and Materials and Methods in the Feed Studies of Codeine (continued) 

14-Day Studies 

Histopathology 

Complete histopathology was 

performed on all animals in the 0, 

6,250 (females only), 12,500, and 

25,OOO ppm (rats) and 0 and 

12,500 ppm (mice). In addition to 

gross lesions and tissue masses, the 

tissues examined included: adrenal 

gland, brain, clitoral gland (rats only), 

esophagus, femur with marrow and 

epiphysis, heart, large intestine (cecum, 

colon and rectum), small intestine 

(duodenum, jejunum, and ileum), 

kidney, liver, lungs and mainstem 

bronchi, lymph nodes (mandibular and 

mesenteric), mammary gland, nose, 

ovary, pancreas, parathyroid gland, 

pituitary gland, preputial gland (rats 

only), prostate gland, salivary gland, 

skin, spleen, stomach (forestomach and 

glandular), testis, thymus, thyroid 

gland, trachea, urinary bladder, and 

uterus. In addition, the stomach, 

testis, and thymus were examined in 

1,562 and 3,125 ppm male and female 

rats and 6,250 ppm male rats. 

13-Week Studies 

Complete histopathology was 

performed on all 0 and 6,250 ppm rats 

and mice and all mice that died before 

the end of the study. In addition to 

gross lesions and tissue masses, the 

tissues examined included: adrenal 

gland, brain, clitoral gland (rats only), 

esophagus, heart, large intestine 

(cecum, colon and rectum), small 

intestine (duodenum, jejunum, and 

ileum), kidney, liver, lungs and 

mainstem bronchi, lymph nodes 

(mandibular and mesenteric), mammary 

gland, nose, ovary, pancreas, 

parathyroid gland, pituitary gland, 

preputial gland, prostate gland, salivary 

gland, skin, sternebrae with marrow, 

spleen, stomach (forestomach and 

glandular), testis, thymus, thyroid 

gland, trachea, urinary bladder, uterus, 

and vagina. Additionally, the liver was 

examined in 3,125 ppm female rats. 

Sperm Morphology and Vaginal Cytology Evaluations 

None Rats and mice in the 0, 1,562, 3,125, 

and 6,250 ppm groups were evaluated. 

Sperm samples were collected at the 

end of the studies and evaluated for 

sperm morphology, count, and 

motility. The right cauda, epididymis, 

and testis were weighed. Vaginal 

samples were collected for 7 consecutive days before the end of the studies 

and evaluated for the relative frequency 

of estrous stages and for estrous cycle 

length. 

2-Year Studies 

Complete histopathology was 

performed on all rats and mice. In 

addition to gross lesions and tissue 

masses, the tissues examined included: 

adrenal gland, brain, clitoral gland, 

esophagus, femur with marrow and 

epiphysis, heart, large intestine (cecum, 

colon and rectum), small intestine 

(duodenum, jejunum, and ileum), 

kidney, liver, lungs and mainstem 

bronchi, lymph nodes (mandibular and 

mesenteric), mammary gland, nose, 

ovary, pancreas, parathyroid gland, 

pituitary gland, preputial gland, 

prostate gland, salivary gland, skin, 

spleen, stomach (forestomach and 

glandular), testis, thymus, thyroid 

gland, trachea, urinary bladder, and 

Uterus. 

None 
33 

RATS 

I$-DAY STUDY 

Onefemaleexposed to 6,250 ppm, onemaleand 

three females exposed to 12,500ppm, and all males 

and females exposed to 25,000 ppm died during the 

study (Table 2). Final mean body weights and mean 

bodyweightgainsofallexposed groups except 

1,562 ppmfemalesweresignificantlylowerthan 

thoseof the controls. Initialfeedconsumptionby 

exposed groups was lower than that by the controls. 

Although feed consumption by exposed groups had 

improved by day 14, feed consumption by all groups 

ofexposedmalesandby 12,500 ppmfemales 

remained lower than that bythe controls. Dietary 

levels of 1,562, 3,125, 6,250, 12,500, or

25,000 ppmcodeineresultedindailydoses of 

approximately 125,250,450,650, or 750 mg 

codeine/kg body weight to males and 125, 250, 500, 

700, or 300 mg/kg to females. 

The absoluterelative weights and thymus of

12,500ppm males ad females and the absolute and 

relativetestisweightsof 12,500ppmmaleswere 

significantlyowerthan those ofthe controls 

(Table Fl); reduced organ weights wereconsidered 

secondary to reduced body weights. 

No chemical-related gross lesions were observed in 

ratsnecropsy.Nonneoplastic at lesions were

observedprimarilyinthe 12,500 and 25,000 ppm 

groups and were associated with decreased survival 

andincreasedmorbidity in these groups. These 

lesionsincludedthickeningoftheforestomach 

mucosa consisting of hyperplasia (males: 0 ppm, 015; 

1,562ppm, 0/5; 3,125 ppm, 2/5; 6,250 ppm, 1/5; 

12,500ppm, 2/5;25,000 ppm, 1/5; females: 015, 

0/5, 0/5, 0/5, 1/4,2/4) and hyperkeratosis (males: 

05, 0/5, 1/5, 0/5, 2/5, 1/5; females: 0/5, 0/5, 0/5, 

1/5,1/4, 2/4), lymphoiddepletionof the thymus 

(males: 0/5, 0/5, 0/5,0/5, 115, 515; females: 015, 

0/5, 0/5, 1/5, 3/3, 4/4), andtesticulardegeneration 

(0/5, 0/5, 0/5, 015, 315, 215). 

Because of the lower body weights in the12,500 ppm 

groups, the high dose selected for the 13-week study 

was 6,250 ppm. 
34 Codeine, NTF’ TR 455 

TABLE2 

Survival, Mean Body Weights, and Feed Consumption of Rats in the 14-Day Feed Study of Codeine 

Final Weight 

Mean Bodv Weightb (g) Feed Relative 

ChangeFinal Initial Dose Survivala 

@Pd (%I Day 1 Day 14 

Male 

3,125 

1.562 

0 

515 

515 

515 

123 f 3 

125 f 2 

123 f 2 

165 f 4** 

172 f 3** 

193 f 5 

42 f 1** 

46 f 2** 

70 f 3 

85 

89 

13.6 

0.4 

0.3 

15.9 

13.5 

13.3 

6,250 515 126 f 3 151 f 2** 25 f 1**78 0.4 12.8 

12,500 4Pid 125 f 3 119 f 5** -4 f 3** 62 0.3 11.2 

25.OOO Otie 128 f 2

Female 

0 515 113 f 2 141 f 3 28 f 1 10.6 11.3 

1,562 515 115 f 3 140 f4 25 f 2 99 5.1 12.8 

3,125 515 113 f 2 126 * 5** 13 f 3** 89 7.3 11.5 

6,250 415f 114 f 3 128 f 3* 13 f 0** 90 3.4 13.3 

12,500 2l5g 115 f 2 112 f 7** -6 f 3** 80 7.8 10.0 

25,OOO 015h 113 f 2 

* Significantly different(PSO.05) from the control group by Williams’ or Dunnett’s test 

** PSO.01 

a Numberof animals survivinglnumber initially in group 

Weights and weight changes are given as mean f standard error. Subsequent calculations are based on animals surviving to the end of 

the study. No fmal mean body weights were calculated for groups with 100%mortality. 

Feed consumption data are based on 10 animals per group and expressed as grams of feed consumed per animal per day. No feed 

consumption data available for25,OOO ppm rats. 

Dayofdeath: 12 

e Dayofdeath: 10, 12, 12, 12, 13 

Dayofdeath: 4 

Dayofdeath: 7,9, 9 

Dayofdeath: 7, 7, 8, 9, 9 
Results 35 

13-WEEK STUDY 

Onemalerat in the 390 ppmgroupdiedduring 

week 2; all other rats survived until the end of the 

study (Table3). Final mean body weights and mean 

body weight gains of all groups of males exposed to 

codeine and of females exposed to 1,562, 3,125, and 

observationsincludedruffled fur, alopecia,and 

abnormal posture in 3,125 and 6,250ppm females. 

As showninTable G1, therewerealterations of 

various hematology, clinical chemistry, and urinalysis 

parametersattheend of the 13-week ratstudy. 

6,250ppm were significantly lower than those of theThere was mildose-dependentleukopeniaand 

controls. Feed consumption decreased with increas- lymphopenia, evidenced by decreased leukocyte and 

ing exposure level during thefust week of the study; lymphocytecountsinfemalesreceiving 1,562ppm 

however, by the end of the study, feed consumption and above and in 6,250ppm males. The decreased 

by most exposed groups was similar to that by the leukocyte were to counts attributeddecreased 

controls.Dietarylevels of 390, 781, 1,562, 3,125,lymphocytenumbers. A decreaseinlymphopoiesis 

or 6,250ppmcodeineresulted in dailydoses of isoftenassociatedwithlymphopeniaandhasbeen 

approximately 25, 50, 100, 200, or 450 mg associated with such conditions as lower body weight 

codeine/kgbodyweighttomalesand 25, 50, 100, gain or malnutrition. In this study,theexposed 

250, or 500 mg/kgtofemales.Alteredclinical animalsgainedmuchlessweightthancontrols, 

TABLE3 

Survival, Mean Body Weights, and Feed Consumption of Rats in the 13-Week Feed Study of Codeine 

Final Weight 

Mean Bodv Weightb k) Feed Relative 

Dose Survivala Initial Consumption‘ Final Controls to Change 

@Pd (%I Week 1 Week 13d 

Male 

0 10110130 

128 10110

f 3 379 f 7 249 f 6 11.8 19.2 

130 390 9110e f 3 352 f 6** 223 f 6** 93 11.3 19.3

781 f 3 348 f 6** 220 f 5** 92 11.2 18.6 

1,562 10110 132 f 3 330 f 7** 198 f 6** 87 10.7 17.0

3,125 , 10110 129 f 3 311 f 5** 182 f 5** ’ 82 9.8 17.5 

6,250 10110 131 f 3 309 f 7** 178 f 6** 82 8.3 19.2 

Female 

0 10110 92*2 195 f 4 103 f 4 10.2 8.9 

390 10110 92f2 193 f 3 101 f 3 99 8.4 10.4 

781 10110 92f2 190 f 4 97 8.410.0 99 f 4 

1,562 10110 91 *2 180 f 6* 88 f 6* 92 7.7 10.7 

3,125 10/10 89 f 2 173 f 5** 84 f 5** 89 7.8 11.9 

6.250 10110 91 f2 170 f 3** 79 f 3** 87 6.0 12.1 

* Significantly different (Pi;O.OS)fromthecontrolgroupbyWilliams’orDunnett’stest 

** P40.01 

a Numberofanimalssurvivinglnumberinitiallyingroup 

Weights and weight changes are given as mean f standard error. Subsequent calculations are based on animals surviving to the end of 

the study. 

Feed consumption is expressed as grams of feed consumed per animal per day. 

Feed consumption data for females from week 12 

e Week of death: 2 
36 Codeine, NTP TR 455 

indicating a decreased nutritionalstatus,andthere No significant differences in sperm morphology or 

was a dose-related decrease inthymusweight.vaginalcytology parameters between control and 

Therefore, the lymphopenia would be consistent with exposed rats were observed (Table Hl). 

decreased related lymphopoiesis to decreased

nutrition and possibly compounded by a stress-related 

increase in endogenous steroid production. There 

also was a minimal to mild macrocytosis, evidenced 

by increased mean cell volumes, that occurred in all 

exposed groups of males and in females exposed to 

781,3,125, or 6,250ppm. The presence of larger 

red cells in the absence of an erythropoietic response 

would be consistent with altered iron metabolism or 

hemoglobin production and could also be related to 

the decreased body weights andalterednutritional 

status. Most other changes wereminimalandonly 

occurred in male rats exposed to 1,562ppmand 

above. For example, there were minimaldecreases 

Absoluteandrelativeadrenalglandweights of all 

groups of exposed males and of3,125and 6,250ppm 

females were significantly greater than those of the 

controls (Table F2). Absoluteand relative liver 

weightsofexposedmales were significantly lower 

than those of the controls. Relative thymus weights 

of 3,125 and 6,250ppmmales were significantly 

lowerthanthoseof the controls. No chemicalrelated gross or histopathologic lesions were observed 

in male or female rats. 

Dose SelectionRationale: Based on markedly lower 

in red cell counts, reticulocyte counts, serum glucose final mean body weights in groups exposed to 3,125 

concentrations, and alkalinephosphataseactivities;and 6,250ppm, codeine exposure levels selected for 

eachofthese parameters can beaffectedby a the2-yearfeedstudy in male andfemale rats were 

decreased nutritional status or feed intake. 400,800,and 1,600ppm.
Results 37 

%YEAR STUDY 

Survival 

Estimatesof 2-year survival probabilities for male 

andfemale rats are shown in Table 4 andinthe 

Kaplan-Meier survival curves (Figure 2). Survival of 

400 ppm females was significantly greater than that 

of the controls; survival of all groups of exposed 

males and of800and 1,600 ppm females was similar 

to that of the controls. 

Body Weights, 

Feed and Compound Consumption, 

and Clinical Findings 

There was an exposure-related decrease in mean body 

weights of males and females (Figure 3 and Tables 5 

TABLE4 

Survival of Rats in the 2-YearFeed Study of Codeine 

Male 

Animals initially in study 

15-Month interim evaluationa 

Moribund 

Natural deaths 

Animals surviving to study termination 

Percent probability of survival at endof studyb 

Mean survival (days)' 

Survival analysisd 

Female 

Animals initially in study 

15-Month interim evaluationa 

Moribund 

Natural deaths 

Animals surviving to study termination 

Percent probability of survival at endof study 

Mean survival (days) 

Survival analysis 

a Censoredfromsurvivalanalyses 

0 PPm 

60 

10 

14 

7 

29 

59 

629 

P=O.244 

60 

10 

15 

7 

28 

56 

628 

P=0.782N 

and 6). Mean body weights of 1,600 ppm males and 

females were lower than those of the controls from 

week 2, and the final mean body weights were 88% 

(males)and 89% (females) those of the controls. 

Feed consumption by exposed groups was similar to 

thatbythe controls (Tables J1 and 52). Dietary 

levels of 400,800,and 1,600 ppm codeine resulted 

in daily doses of approximately 15, 30, and 70 mg 

codeine/kgbodyweight to males and 15, 40,and 

80 mg/kg to females.Chemical-relatedclinical 

findings were limited to ocular discharge in exposed 

males and females. 

400 ppm 800 ppm 1,600 ppm 

60 60 60 

10 10 10 

23 24 23 

7 5 7 

20 21 20 

40 42 40 

635 643 628 

P=O.216 P=O.319 P=O.193 

60 60 60 

10 9 9 

9 17 14 

3 5 5 

38e 2% 32 

76 57 63 

665 659 663 

P=0.035N P=0.718N P=0.369N 

Kaplan-Meier determinations basedon the number of animals alive on the first day of terminal sacrifice ' Mean ofall deaths (uncensored, censored, and terminalsacrifice) 

The result of the life table trend test (Tarone, 1975) is in the control column, and the results of the life table pairwise comparisons (Cox, 

1972) with the controls are in the exposure columns. A negative trend or a lower mortality in an exposure group is indicated by N. 

e Includes one animal that died during the last week of the study. 
38 Codeine, NTP TR 455 

0 15 so 45 60 75 90 105 

WEEKS ON STUDY 

0 15 45 60 75 90 105 D 

WEEKS ON STUDY 

FIGURE 2 

Kaplan-Meier Survival Curves for Rats Administered Codeine in Feed for 2 Years 
Results 39 

0 8 15 30

, 4s 

I 

60 

1 

75 

I 

' 90 

I 

105 

I 

!O 

WEEKS ON STUDY 

I I I 

0 15 so 4s 60 75 90 165 0 

WEEKS ON STUDY 

FIGURE 3 

Growth Curves for Rats Administered Codeine in Feed for 2 Years 
40 Codeine, NTP TR 455 

TABLE5 

Mean Body Weights and Survival of Male Rats in the 2-Year Feed Study of Codeine 

Weeks 0 ppm 400 ppm 800 ppm 1,600 ppm 

on Av.Wt. No. of Av.Wt.Wt. (% of No. ofAv.Wt.Wt. (% of No. of Av.Wt.Wt. (% of No. of 

Study (8) Survivors (8) controls) Survivors (8) controls) Survivors (8) controls) Survivors

1 154 60 157 102 60 154 100 60 155 100 60 

2 199 60 187 94 60 182 92 60 179 90 60 

3 234 60 22 1 95 60 216 92 60 211 90 60 

4 257 60 243 95 60 238 93 60 235 92 60 

5 274 60 260 95 60 254 93 60 250 91 60 

6 292 60 279 96 60 272 93 60 268 92 60 

7 302 60 294 97 60 286 95 60 279 92 60 

8 316 60 305 97 60 299 95 60 293 93 60 

9 325 60 314 97 60 307 95 60 298 92 60 

10 341 60 327 96 60 320 94 60 315 92 60 

11 347 60 333 96 60 325 94 60 318 92 60 

12 352 60 340 96 60 333 95 60 330 94 60 

13 360 60 346 96 60 339 94 60 337 94 60 

14 368 60 355 96 60 348 94 60 346 94 60 

17 392 60 370 95 60 363 93 60 354 90 60 

21 407 60 396 97 60 382 94 60 366 90 60 

25 430 60 414 96 60 398 93 60 387 90 60 

29 438 60 426 97 60 408 93 60 397 91 60 

33 444 59 429 97 60 414 93 60 394 89 60 

37 452 59 437 97 60 420 93 60 402 89 60 

41 45 8 59 442 96 60 428 93 60 405 88 60 

45 465 59 452 97 60 434 93 60 408 88 60 

49 466 58 452 97 59 435 93 60 405 87 60 

53 478 58 465 97 59 444 93 60 407 85 59 

57 477 58 468 98 59 450 94 60 409 86 58 

61 470 58 459 98 58 443 94 60 404 86 58 

65 471 56 458 97 57 444 94 60 400 85 58 

69a 463 46 454 98 47 439 95 50 403 87 48 

73 459 44 453 99 47 435 95 50 397 87 47 

77 458 43 448 98 47 429 94 48 395 86 46 

81 45 3 42 444 98 46 423 94 45 393 87 41 

85 444 41 439 99 44 423 95 42 393 89 38 

89 447 39 434 97 41 424 95 41 396 89 36 

93 437 37 428 98 37 415 95 36 390 89 35 

97 427 33 412 97 33 408 96 32 382 89 31 

101 433 30 418 97 26 40 1 93 29 376 87 28 

105 41 1 29 405 99 20 390 95 21 362 88 22 

Mean for weeks 

1-13 289 277 96 27 1 94 267 93 

14-52 432 417 97 403 93 386 90 

53-105 452 442 98 426 94 393 87 

a Interim evaluationoccurredduringweek 65. 
Results 41 

TABLE6 

Mean Body Weights and Survival of Female Rats in the 2-Year Feed Study of Codeine 

Weeks 0 ppm 400 PPm 800 ppm 1,600 pDm 

on Av. Wt. No. of Av. Wt.Wt. (% of No. of Av. Wt.Wt. (% of No. of Av. Wt.Wt. (% of No. of 

Study (9) Survivors (9) controls) Survivors (9) controls) Survivors (9) controls) Survivors 

1 121 60 121 100 60 121 100 60 120 99 60 

2 140 60 136 97 60 136 97 60 133 95 60 

3 155 60 150 97 60 149 96 60 147 95 60 

4 167 60 158 95 60 156 94 60 155 93 60 

5 175 60 166 95 60 165 94 60 163 93 60 

6 182 60 175 96 60 171 94 60 173 95 60 

7 190 60 183 96 60 179 95 60 180 95 60 

8 193 60 187 97 60 184 95 60 184 95 60 

9 195 60 187 96 60 184 94 60 182 93 60 

10 201 60 192 95 60 191 95 60 188 93 60 

11 204 60 196 96 60 193 95 60 191 94 60 

12 209 60 199 95 60 197 94 60 197 94 60 

13 212 60 196 93 60 198 93 60 194 91 60 

14 215 60 205 95 60 203 95 60 198 92 60 

17 224 60 211 95 60 206 92 60 204 91 60 

21 233 60 223 96 60 221 95 60 217 93 60 

25 242 60 228 94 60 227 94 60 223 92 60 

29 244 60 233 96 60 232 95 60 232 95 60 

33 255 60 237 93 60 235 92 60 234 92 60 

37 260 60 240 93 60 243 93 60 245 94 60 

41 273 60 251 92 60 251 92 60 249 91 60 

45 286 60 259 91 60 258 90 60 255 89 60 

49 297 59 266 89 60 262 88 60 256 86 60 

53 310 59 277 89 60 270 87 60 264 85 60 

57 320 59 288 90 60 284 89 59 271 85 59 

61 328 59 295 90 59 292 89 59 279 85 59 

65 336 58 306 91 59 302 90 58 283 84 58 

69a 331 46 308 93 49 304 92 49 284 86 48 

73 339 44 312 92 48 307 91 49 290 86 48 

77 342 43 317 93 48 315 92 49 297 87 48 

81 342 41 319 93 46 317 93 49 298 87 48 

85 341 36 325 95 46 323 95 48 307 90 48 

89 338 35 326 96 45 325 96 47 304 90 48 

93 343 34 333 97 45 327 95 43 309 90 43 

97 345 33 334 97 45 324 94 40 307 89 42 

101 349 30 336 96 42 324 93 32 303 87 39 

105 342 28 335 98 39 315 92 29 303 89 32 

Mean for weeks 

1-13 180 173 96 171 95 170 94 

14-52 253 235 93 234 93 231 92 

53-105 336 315 94 309 92 293 87 

a Interim evaluation occurred during week 66. 
42 Codeine, NTP TR 455 

Pathology and Statistical Analyses 

This section describes the statisticallysignificant or 

biologically noteworthy changes in the incidences of 

neoplasms and/or nonneoplastic lesions of the adrenal 

gland, clitoral/preputial gland, mammary gland, 

pituitary gland, respiratory system, and liver. 

Summaries of the incidencesofneoplasmsand 

nonneoplastic lesions, individual tumor animal

diagnoses, statistical analyses of primary neoplasms 

that occurred with an incidence of at least 5% in at 

least one animal group, and historical incidences for 

the neoplasms mentioned in this section are presented 

inAppendix A for male rats and Appendix B for 

female rats. 

Adrenal Gland: Absolute and relative adrenal gland 

weightsof 800 and 1,600 ppm malesweresignificantly greater than those of the controls at 15 months 

(Table F3). In histological sections, the overall 

cross-sectional area of the adrenal gland was similar 

between controls and exposure groups. However, 

when compared to that of the controls, there was 

generally a smalleramountof adrenal medulla 

relative to the amount of adrenal cortex in 800 and 

1,600 ppm males. In these exposure groups, the 

zona reticularis appeared to be slightly thickened and 

the cells of the inner portion of the zona fasicularis 

wereslightlysmaller than those of the controls. 

Vasculardilatation in the medullaand inner cortex 

wasalsoslightly more prominent in 800 and 

1,600 ppm males. These effects were not present in 

400 ppm males or in any group of exposed females. 

At 2 years, there were exposure-related decreased 

incidencesofadrenalmedulla hyperplasia in males 

andfemales (Tables 7, A5, and B5). There was an 

exposure-related decrease in the incidence of benign 

pheochromocytomain males, and the incidences in 

exposed males were significantly lower than that in 

the controls. The incidence of benign pheochromocytoma in 1,600 ppm males was lower than had been 

seen in any historical control group (Tables 7 and 

A4b). These decreases were considered to be related 

to codeine exposure. 
Results 43 

TABLE7 

Incidences of Neoplasms and Nonneoplastic Lesionsof the Adrenal Medulla in Rats 

in the 2-Year Feed Studyof Codeine 

400 ppm 800 ppm 1,600 ppm 

Male 

Number Examined Microscopically 49 50 50 50 

Hyperplasiaa 25 (l.8)b 12**(1.8) 10**(1.5) 9**(1.9) 

Benign Pheochromocytoma' 

Overall rated 16/49 (33%) 6/50 (12%) 6/50 (12%) 3/50 (6%) 

Adjusted ratee 49.1% 24.9% 21.7% 13.6% 

Terminal ratef 12/28 (43%) 3/20 (1 5 %) 3/21 (14%) 2/20 (10%) 

First incidence (days) 599 678 608 708 

Logistic regression testg P=O.WlN P=0.013N P=O.OlON P=O.WlN 

Female 

Number Examined Microscopically 50 50 50 51 

Hyperplasia 8 (1.4) 5 (1.4) 2* (1.5) 2* (1.5) 

Benign Pheochromocytomah 

Overall rate 2/50 (4%) 5/50 (10%) 1/50 (2%) 1/51 (2%) 

Adjusted rate 6.6% 12.2% 2.7% 3.1% 

Terminal rate 1/28 (4%) 3/38 (8%) 0128 (0%) 1/32 (3%) 

First incidence (days) 677 689 686 733 (T) 

Logisti' regression test P=0.158N P=O.296 P=0.472N P=0.437N 

* Significantly different (P50.05) from the control group by the logistic regression test 

** PSO.01 

a Number of animalswithlesion 

Average severity of lesions in affected animals: l=minimal, 2=mild, 3=moderate, 4=marked ' Historical incidence for 2-year NTP feed studies with untreated control groups (mean f standard deviation): 379/1,182 

(32.1% f 11.7%); range 10%-63% 

Number of animals with neoplasm per numberof animals with adrenal medulla examined microscopically 

e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence in animals surviving until the end of the study 

g In the control column are the P values associated with the trend test. In the exposure group columns are. the P values corresponding to 

the painvise comparison between the controls and that exposure group. The logistic regression test regards lesions in animals dying 

prior to terminal kill as nonfatal. A negative trend or a lower incidence in an exposure. group is indicated by N. 

Historicalincidence:4911175 (4.17% f 2.50%); range 0%-8% 
44 Codeine, N" TR 455 

ClitoraVPreputial Gland: In 1,600 ppm females, the 1,600 ppm, 11/50)andtheincidence of preputial 

incidence of clitoral gland ectasia was significantly glandectasiain400ppmmales (1/50, 9/50, 6/48, 

greater than that in the controls (Tables 8 and B5). 4/50)weresignificantlygreaterthanthoseinthe 

The incidenceof clitoral gland adenoma or carcinoma,controls at 2 years (Table A5). 

(combined)in 1,600 ppm females was significantly 

lower than that in the controls (Tables 8 and Bl), and 

it was at the low end of the historical control range Mammary Gland: The incidences of mammary gland 

(2 %-21% , Tables 8 and B4a).Theincidence of fibroadenoma and fibroadenoma or adenocarcinoma 

preputialglandhyperplasiain 1,600 ppm males (combined) in 1,600 ppm females were significantly 

(0 ppm, 3/50; 400ppm, 1/50; 800 ppm, 4/48; lower than those in the controls (Table 9). 

TABLE8 

Incidences of Neoplasms and Nonneoplastic Lesionsof the Clitoral Gland in Female Rats 

in the 2-YearFeed Study of Codeine 

0 PPm 400 ppm 800 ppm 1,600 ppm 

Number Examined Microscopically 47 47 50 51 

Ectasiaa 5 (2.2)b 9 (2.7) 20**(2.3) 7 (2.4) 

Inflammation 0 0 1 (3.0) 3 (3.0) 

Adenoma 

Overall rate' 5/47 (1 1%) 6/47 (13%) 12/50 (24%) 1/51 (2%) 

Adenoma or Carcinomad 

Overall rate 6/47 (13X) 6/47 (1 3 X) 12/50 (24%) 1/51 (2%) 

Adjusted ratee 19.0% 15.7% 37.8% 3.1% 

Terminal ratef 3/27 (11%) 5/36 (14%) 10129 (34%) 1/32 (3X)

First incidence (days) 587 546 662 733 (T)

Logistic regression testg P=0.052N ,P=0.568N P=O.189 P=0.033N 

(T) Terminalsacrifice 

** Significantly different (PSO.01) from the control group by the logistic regression test 

Number of animals with lesion 

Average severity of lesions in affected animals: l=minimal, 2=mild, 3=moderate, 4=marked 

Number of animals with neoplasm per numberof animals with clitoral gland examined microscopically 

Historical incidence for 2-year NTF' feed studies with untreated control groups (mean f standard deviation): 119/1,117 

(10.7% f 5.6%); range 2%-21% 

Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence in animals surviving until the endof the study 

In the control column are the P values associated with the trend test. In the exposure group columns are the P values corresponding to 

the pairwise comparison between the controls and that exposure group. The logistic regression test regards lesions in animals dying 

prior to terminal kill as nonfatal. A negative trend or a lower incidence in an exposure group is indicated by N. 
Results 45 

TABLE9 

Incidences of Mammary Gland Neoplasms in Female Rats in the 2-Year Feed Study of Codeine 

0 PPm 400 ppm 800 ppm 1,600 ppm 

Fibroadenomaa 

Overdl rateb 27/50 (54%) 21/50 (42%) 27/51 (53%) 8/51 (16%) 

Adjusted rate' 70.3% 47.4% 68.2% 23.0% 

Terminal rated 17/28 (61X) 15/38 (39%) 17/29 (59%) 6/32 (1 9%) 

First incidence (days) 475 400 641 645 

Logistic regression teste P<0.001N P=0.087N P=0.359N P<0.001N 

Adenocarcinomaf 

Overall rate 3/50 (6%) 2/50 (4%) 3/51 (6%) 0151 (0%)

Adjusted rate 8.5% 5.3% 10.3% 0.0% 

Terminal rate 1/28 (4%) 2/38 (5%) 3/29 (10%) 0132 (0%)

First incidence (days) 495 733 (T) 733 (T) 3 

Logistic regression test P=0.102N P=0.533N P=0.650N P=0.147N 

Fibroadenoma or Adenocarcinomah 

Overall rate 30/50 (60%) 23/50 (46 %) 29/51 (57%) 8/51 (16%) 

Adjusted rate 74.4% 52.0% 73.5% 23.0% 

Terminal rate 18/28 (64%) 17/38 (45%) 19/29 (66%) 6/32 (19%) 

First incidence (days) 475 400 641 645 

Logistic regression test P<0.001N P=0.063N P=0.284N P<0.001N 

a Historical incidence for 2-year NTP feed studies with untreated control groups (mean f standard deviation): 46511.202 

(38.7%f 12.7%);range 8%-58% 

Number of animals with neoplasm per number of animals with mammary gland examined microscopically 

Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence in animals surviving until the end of the study 

e In the control column are the P values associated with the trend test. In the exposure group columns are the P values corresponding to 

the pairwise comparison between the controls and that exposure group. The logistic regression test regards lesions in animals dying 

prior to terminal kill as nordad. A negative trend or a lower incidence in an exposure group is indicated by N. 

Historical incidence (carcinoma): 32/1,202 (2.7% f 2.9%);range 0%-10% 

Not applicable; no neoplasms in animal group 

Historical incidence (includes data for adenoma): 507/1,202 (42.2% f 13.5%);range 8%-64% 

PituitaryGland: Theincidence of pituitary gland RespiratorySystem: Theincidences of cytoplasmic 

(parsdistalis)focalhyperplasiain 400 ppmmales alteration of theolfactoryepitheliumin 800 ppm 

was greater than that in the controls at2 years (5/50, males and 1,600 ppm males and females were greater 

14/50, 5/48, 3/47; Table A5). Theincidencesof than those in the controls at 2 years (males: 12/50, 

pituitary gland (pars distalis) angiectasis in 400 and 16/50, 28/50, 47/50; females: 38/50, 45/50, 42/51, 

1,600 ppmfemaleswerelowerthanthatinthe 49/51;Tables A5 and B5). 

controls (13/46, 7/50, 10/50, 7/51; Table B5). The 

incidence of pituitary glandadenoma or carcinoma Liver: Absolute and relative liver weights of exposed 

(combined)in 1,600 ppmfemaleswassignificantly malesweresignificantlylowerthanthoseofthe 

lower than that in the controls (17/46, 18/50, 19/50, controls at 15 months(Table F3). At 2 years, there 

12/51 ; Tables B1 and B3), andtheincidencewas was an exposure-related decrease in the incidence of 

lowerthanthehistoricalcontrolrange (30%-80%; cystic degeneration of the liver (14/50, 5/50, 1/50, 

Table B4c). 0/50; Table A5). 
46 Codeine, NTP TR 455 

MICE 300, 600, 1,300, or 3,000 mg/kgbodyweight to 

males and 200, 400, 750, 1,500, or 3,000 mg/kg to 14-DAY STUDY females.

All mice survived to the end of the study (Table 10). 

The final mean body weight of 3,125 ppm females 

was significantly greater than that Absolute and relative liver weights of 3,125, 6,250, of the controls; the and12,500ppmmalesandof12,500ppmfemales final mean body weight of 12,500 ppm females and and the absolute and relative right kidney weights of themeanbodyweightgainsof12,500ppmmales 12,500 ppm malesweresignificantlylowerthan andfemalesweresignificantlylowerthanthose of 

those of the controls (Table F4). No gross or histo- thecontrols.Initially,feedconsumption by 6,250 pathologiclesionswere attributed to codeine and 12,500 ppm males and females was lower than 

exposure. that by the controls. At day 14, feed consumption by 

exposedmalesandfemales,except 12,500 ppm 

females, was similar to that by the controls. Dietary Becauseofthelowermeanbodyweightgainsof 

levelsof 781, 1,562, 3,125, 6,250, or 12,500 ppm 12,500 ppm malesandfemales,thehighdose 

codeine resulted in daily dosesof approximately 150, selected for the 13-week study was 6,250 ppm. 

TABLE10 

Survival, Mean Body Weights, and Feed Consumption of Mice in the 14-Day Feed Study of Codeine ~ ~~ _____~ 

DoseSurvivalaInitial 

@P@ 

Final toControlsConsumDtion‘ 

MeanBodyWeightb (e) Feed 

Change 

(%I 

Relative 

Final Weight 

Day 1 Day 14 

Male 

0 515 21.1 f 0.6 25.2 f 0.9 4.1 f 0.4 4.0 4.1 

781 515 21.2 f 0.6 25.7 f 1.0 4.5 f 0.4 102 4.2 4.6 

1,562 515 21.5 f 1.0 25.9 f 1.2 4.4 f 0.3103 4.1 

3,125 515 20.9 f 0.6 24.4 f 1.0 3.4 f 0.5 97 3.6 

5.0 

4.9 

6,250 515 21.8 f 0.5 24.7 f 0.8 2.9 98f 0.3 2.4 3.8 

12,500 515 20.9 f 0.5 22.8 f 0.7 1.9 f 1.1* 90 2.2 3.5 

Female 

0 515 16.7 f 0.4 18.7 f 0.3 2.0 f 0.3 4.5 3.9 

78 1 515 16.8 f 0.4 18.8 f 0.2 

1,562 515 14.8 f 0.9 18.2 f 0.4 

3,125 515 17.0 f 0.4 20.1 f OS* 

2.0 f 0.2101

3.4 f 0.7*97

3.1 f 0.2108

4.3 4.1 

4.4 4.7 

4.2 4.2 

12.500 

6,250 

515 

515 

16.4 f 0.4 

17.0 f 0.3 

16.8 f 0.3** 

19.0 f 0.2 

0.4 f 0.3* 

2.0 f 0.2 

90 

102 2.8 

2.3 

3.1 

2.9 

a Numberof animals survivinglnumber initially in group 

** P40.01 

* Significantly different(P 40.05) from the control group by Williams’ or Dunnett’s test 

Feed consumption dataare based on 25 animals per group and expressed as grams of feed consumed per animal per day. 

Weights and weight changes are given as mean f standard error. 
Results 47 

13-WEEK STUDY 

Two male mice in the 3,125 ppm group died during 

week 7 (Table 1 l), and these deaths were not related 

to codeine administration. All other mice survived to 

theendofthestudy.Finalmeanbodyweightsof 

exposed males and females were similar to those of 

the controls, although the final mean body weight of 

6,250 ppm maleswas 9% lowerandthatof 

6,250 ppm females was 6% lower thanthatofthe 

respectivecontrols.Feedconsumptionbyexposed 

males and females was similar to that by the controls. 

Dietarylevels of 390,781, 1,562, 3,125, or 

6,250 ppm codeine resultedin daily doses of approximately 60, 120, 260,460,or1,000 mg codeinelkg 

bodyweighttomalesand60,130,280,530,or 

1,200 mg/kg tofemales.Abnormalposture was 

observed in all exposed groups of males. 

Therewereno chemical-related in differences

hematologyorurinalysisparametersinmaleor 

femalemice(TableG2).Minor,sporadicchanges 

TABLE11 

occurredin a few oftheclinicalchemistryparameters; they were not considered biologically significant. 

Sperm motility in 1,562 ppm males was significantly 

lessthanthatofthe controls (Table H2); however, 

this finding was not observed at 3,125 or 6,250 ppm. 

No significantdifferencesinspermmorphologyor 

vaginal cytology were attributed to codeine exposure. 

Absoluteandrelativekidneyweightsof 3,125 and 

6,250 ppm males were significantly lower than those 

ofthecontrols(Table F5). No chemical-related 

differencesinorganweightswereobservedin 

females. No chemical-relatedgross or histopathologic lesions were observed in male or female mice. 

Dose Selection Rationale:Because of the lower body 

weights at 6,250 ppm and above, the exposure levels 

selected for the 2-year feed study in mice were 750, 

1,500, and 3,000 ppm. 

Survival, Mean Body Weights, and Feed Consumptionof Mice in the 13-Week Feed Study of Codeine 

Final Weight 

MeanBodvWeightb M Feed Relative 

ngeFinal SurvivalaDose Initial 

(PPm) (W Week 1Week 13d 

Male 

0 10110 22.5 f 0.3 30.7 f 0.7 8.2 f 0.6 2.8 4.6 

390 10110 22.0 f 0.3 30.3 f 0.3 8.3 f 0.4 99 2.9 4.4 

781 10110 22.7 f 0.2 27.8 f 1.3 5.1 f 1.3 91 3.1 5.3 

1,562 10110 21.5 f 0.2 29.7 f 0.8 8.2 f 0.7 97 3.1 5.2 

3,125 22.2 f 0.3 28.8 f 0.9 6.7 f 0.9 94 2.9 4.4 

6,250 10110 22.8 f 0.2 27.9 f 0.6 5.2 f 0.4 91 2.8 5.0 

Female 

0 10110 16.9 f 0.3 22.5 f 0.7 5.6 f 0.7 2.6 6.6 

390 10110 17.2 f 0.4 24.6 f 0.4 7.3 f 0.2 109 2.5 4.4 

78 1 10/10 16.6 f 0.5 24.5 f 0.5 7.9 f 0.5 109 2.4 4.7 

1,562 10110 17.1 f 0.2 23.5 f 0.8 6.3 f 0.8 104 2.6 4.3 

3,125 10110 17.1 f 0.3 23.0 f 0.7 5.9 f 0.7 102 2.5 4.3 

6,250 10110 17.5 f 0.3 21.2 f 0.8 3.7 f 0.8 94 2.4 5.4 

a Numberofanimals survivinglnumber initiallyin group 

Weights and weight changes are given as mean f standard error. Subsequent calculations are based on animals survivingto the end of 

the study. Differences from the control group were not significant by Williams’ or Dunnett’s test. 

. Feed consumption is expressed as grams of feed consumed per animal per day. 

Feed consumption data for females from week 12 

e Week of death: 7, 7 
48 Codeine, NTP TR 455 

&YEAR STUDY 

Survival 

Estimatesof2-yearsurvivalprobabilitiesformale 

andfemalemiceareshowninTable12andinthe 

Kaplan-Meier survival curves (Figure4). Survival of 

exposed males and females was similar to that of the 

controls. 

Body Weights, 

Feed and Compound Consumption, 

and Clinical Findings 

Mean body weights of 750 and 1,500 ppm males and 

Mean body weights of 3,000 ppm males and females 

werelowerthanthoseofthecontrols fromabout 

week 13, and the final mean body weights of these 

groupswere 86% and 82% those oftherespective 

controls. Feed consumption by exposed groups was 

similartothat by the controls(Tables 53 and54). 

Dietary levels of 750, 1,500, and 3,000 ppm codeine 

resultedindailydosesof approximately 100, 200, 

and400mgcodeine/kgbodyweight.Clinical 

findings included alopecia on the ventral body surface 

in 1,500 ppm malesand 3,000 ppmmalesand 

femalesbeginning on day 57. andruffled fur in 

females were similar to those of the controls through-1,500 ppm males and 3,000 ppm males and females 

out most of the study (Tables 13 and 14, Figure 5). from day 85. 

TABLE12 

.Survivalof Mice in the 2-YearFeed Study of Codeine 

Male 

AnimalsinitiallyinStudy . 

15-Month interim evaluationa 

Accidental deatha 

Moribund 

Natural deaths 

Animals surviving to study termination 

Percent probability of survival at end of studyb 

Mean survival (days)c 

survival analysisd 

Female 

Animals initially in study 

15-Month interim evaluationa 

Missinga 

Moribund 

Natural deaths 

Animals surviving to study termination 

Percent probability of survival at end of study 

Mean survival (days) 

Survival analysis 

a Censoredfromsurvivalanalyses 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

60 60 60 60 

10 10 10 10 

0 1 0 0 

4 4 2 1 

5 7 3 6 

41 38 45 43 

83 79 90 87 

656 650 669 657 

P=0.417N P=O.810 P=0.350N P=0.766N 

60 60 60 60 

10 9 9 9 

0 0 0 1 

7 9 6 4 

7 6 2 11 

36e 36 43 39 

73 72 85 71 

641 627 662 641 

P=0.979N P=O.945 P=O.lIJN P=l.Ooo 

Kaplan-Meier determinations based on the number ofanimals alive on the first day of terminal sacrifice 

Mean of all deaths (uncensored, censored,and terminal sacrifice) * The result of the life table trend test (Tarone, 1975) is in the control column, and the results of the life table pairwise comparisons (Cox, 

1972) with the controls are in the exposed columns. A negative trend or a lower mortality in an exposure group is indicated by N. e Includes one animal that died during the last week of the study. 
Results 49 

1.a 

..... .......... .................. I .................. I ................. i.................. I ....... 

A lso6PPU 

0.4 I I I 1 1 I I 

15 50 45 60 75 90 1os 

WEEKS ON STUDY 

0.6 I I 1 I 

0 0 1s 30 45 60 75 90 1os 

WEEKS ON STUDY 
50 Codeine, NTP TR 455 

TABLE13 

Mean Body Weights and Survival of Male Mice in the 2-Year Feed Study of Codeine 

Study 

Weeks 

on 

0 Survivors 

Av. Wt. No. of 

0 DDUl 

(g) controls) Survivors 

Av. Wt.Wt. (% of No. of 

750 DDUI 

(g) controls) survivors 

Av. Wt.Wt. (96of No. of 

1.500 DDUI 

(g) controls) Survivors 

Av. Wt. Wt. (% of No. of 

3.000 DDU~ 

1 23.2 60 23.1 100 60 22.6 97 60 23.3 100 60 

2 23.8 60 23.6 99 60 23.4 98 60 22.8 96 60 

3 25.6 60 24.9 97 59 25.2 98 59 24.7 97 59 

4 26.7 60 25.9 97 59 25.8 97 59 24.8 93 59 

5 27.7 60 27.3 99 59 26.6 96 59 26.5 96 59 

6 28.1 60 27.7 99 59 27.2 97 59 26.9 96 59 

7 29.7 60 29.1 98 59 28.6 96 59 27.9 94 59 

8 30.6 60 30.0 98 59 29.3 96 59 28.4 93 59 

9 31.6 60 30.2 96 59 29.9 95 59 29.2 92 59 

10 32.4 60 31.1 96 59 30.5 94 59 29.8 92 59 

11 33.4 60 32.2 96 59 31.6 95 59 30.6 92 59 

12 34.1 60 33.0 97 59 32.2 94 59 31.5 92 59 

13 34.6 60 33.3 96 59 32.5 94 59 31.2 90 59 

14 35.2 60 34.0 97 59 32.8 93 59 31.7 90 59 

17 38.1 60 36.9 97 59 35.0 92 59 33.3 87 59 

21 41.6 60 40.0 96 59 37.2 89 59 34.9 84 59 

25 42.3 60 41.1 97 59 39.3 93 59 36.6 87 59 

29 44.9 60 44.8 100 59 42.3 94 59 38.9 87 59 

33 46.1 60 45.1 98 59 42.5 92 59 38.5 84 59 

37 48.3 59 46.8 97 59 44.2 92 59 40.4 84 59 

41 48.9 59 48.9 100 59 45.7 94 59 41.7 85 59 

45 49.5 59 49.9 101 59 47.O. 95 59 43.3 88 59 

49 49.4 59 50.0 101 59 47.4 96 59 43.8 89 59 

53 50.0 59 51.3 103 59 48.7 97 59. 44.6 89 59 

57 50.4 59 51.6 102 59 49.2 98 59 45.2 90 59 

61 50.7 59 51.7 102 58 49.4 97 59 45.4 90 58 

65 50.3 59 51.5 102 58 49.0 97 ,59 45.0 90 58 

69a 50.6 48 51.3 101 47 49.5 98 49 44.7 88 48 

73 49.4 47 50.5 102 47 48.6 98 49 43.4 88 47 

77 49.2 47 50.4 102 47 48.5 99 49 43.4 88 47 

81 49.9 46 50.6 101 46 47.5 95 49 43.7 88 46 

85 49.5 45 50.3 102 45 47.5 96 49 43.8 89 46 

89 50.2 44 51.4 102 44 48.1 96 49 44.2 88 46 

93 48.5 43 50.3 104 42 45.9 95 48 41.8 86 46 

97 49.0 43 51.0. 104 42 46.0 94 47 42.2 86 45 

101 49.0 41 50.3 103 41 45.1 92 47 41.7 85 44 

104 48.2 41 50.4 105 39 44.3 92 45 41.4 86 43 

Mean for weeks 

1-13 29.3 28.6 98 28.1 96 27.5 94 

14-52 44.4 43.8 98 41.3 93 38.3 87 

53-104 49.6 50.9 103 47.7 96 43.6 88 

a Interim evaluation occurred during week 66. 
Results 51 

TABLE 14 

Mean Body Weights and Survival of Female Mice in the 2-YearFeed Study of Codeine 

Weeks 0 DDm 750 DUXU 1.500 DDm 3.000 DDm 

on Av. Wt. No. of Av. Wt. Wt. (96of No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of 

study (9) Survivors (g) controls) Survivors (g) controls) Survivors (g) controls) Survivors 

1 18.3 60 18.5 101 60 18.4 101 60 18.3 100 60 

2 19.5 ' 60 19.6 101 59 19.2 99 59 18.9 97 58 

3 21.2 59 21.2 100 58 21.4 101 59 21.0 99 58 

4 22.1 59 22.3 101 58 22.3 101 59 21.7 98 58 

5 23.5 59 23.6 100 58 23.4 100 59 23.1 98 58 

6 24.5 59 24.5 100 58 24.1 98 59 23.6 96 58 

7 25.9 59 25.9 100 58 25.5 99 59 25.3 98 58 

8 26.7 59 26.7 100 58 26.3 99 59 26.0 97 58 

9 27.6 59 26.9 98 58 26.6 96 59 25.8 94 58 

10 28.3 59 27.7 98 58 27.3 97 59 26.6 94 58 

11 29.3 59 28.4 97 58 27.7 95 59 26.8 92 58 

12 30.4 59 29.0 95 58 28.0 92 59 27.5 91 58 

13 29.8 59 28.6 96 58 27.7 93 59 26.9 90 58 

14 31.1 59 30.0 97 58 29.0 93 59 28.1 90 58 

17 34.5 59 33.3 97 58 31.2 90 59 29.5 86 58 

21 39.1 59 36.9 94 58 34.0 87 59 31.7 81 58 

25 40.7 59 38.8 95 58 36.6 90 59 33.6 83 58 

29 42.5 59 41.7 98 58 38.9 92 59 35.1 83 58 

33 44.6 59 42.4 95 58 39.8 89 59 35.5 80 58 

37 46.2 59 44.6 97 58 41.4 90 59 36.5 79 58 

41 47.5 58 47.2 99 98 43.8 92. 59 38.5 81 58 

45 48.9 58 49.0 100 58 45.3 93 59 39.9 82 58 

49 48.7 58 49.5 102 58 46.5 96 57 40.4 83 58 

53 50.2 58 51.0 102 56 48.3 96 57 42.2 84 58 

57 51.2 58 52.7 103 55 50.4 98 57 43.6 85 58 

61 52.5 58 54.3 103 55 51.7 99 57 44.4 85 57 

65 52.4 58 54.0 103 55 51.3 98 57 44.2 84 56 

69a 52.6 48 53.5 102 46 52.5 100 48 44.2 84 47 

73 50.9 47 53.2 105 45 52.4 103 48 44.3 87 47 

77 49.8 46 53.2 107 44 52.3 105 48 43.7 88 46 

81 51.4 45 53.9 105 43 53.2 104 48 44.3 86 45 

85 53.0 42 54.2 102 43 54.1 102 48 44.1 83 45 

89 53.2 41 53.5 101 42 53.7 101 48 43.6 82 44 

93 51.3 41 52.1 102 40 52.4 102 47 41.7 81 42 

97 52.5 39 53.2 101 37 52.5 100 47 42.4 81 42 

101 51.6 38 53.5 104 36 51.3 99 47 41.4 80 41 

104 49.6 36 51.3 103 36 49.1 99 44 40.6 82 37 

Mean for weeks 

1-13 25.2 24.8 99 24.5 98 24.0 96 

14-52 42.4 41.3 97 38.7 91 34.9 83 

53.1 103 51.8 100 43.2 84 

a Interim evaluation occurred during week 66. 

53-10451.6 
52 

0 15 so 45 60 75 90 105 

WEEKS ON STUDY 

I I I I I I 

0 15 30 45 60 75 90 I5 

WEEKS ON STUDY 

FIGURE 5 

Growth Curves for Mice Administered Codeine in Feed €or2 Years 
Results 53 

Pathology and Statistical Analyses 

This section describes the statistically significant or 

biologically noteworthy changes in the incidences of 

neoplasms and/or nonneoplastic lesions of the thyroid 

gland, liver, and otherorgans.Summaries ofthe 

incidences of neoplasmsandnonneoplasticlesions, 

controls, and follicular cell hyperplasia was observed 

in 1,500 and 3,000 ppm females but not in controls 

(Tables 15, C5, and D5). At 2 years, the incidences 

of follicular cell hyperplasia in all exposed groups of 

malesandfemalesweresignificantlygreaterthan 

those in the controls. Hyperplasia was characterized 

individual animal tumor diagnoses, statistical analysesbyan increase in the number of follicular epithelial 

of primary neoplasms that occurred with an incidencecellsliningthethyroidfollicles. This changewas 

of at least 5% inatleastoneanimalgroup,and 

historical incidences for the neoplasms mentioned in 

thissectionarepresentedinAppendixC for male 

mice and Appendix D for female mice. 

Thyroid Gland: At 15 months,theincidence of 

thyroid gland follicular cell hyperplasia in3,000 ppm 

maleswasignificantlygreaterthanthatinthe 

TABLE15 

minimaltomildinseverity,generallyfocalin 

distribution,andinvolvedone to threeadjacent 

follicles.Theseverity was similar amongexposed 

and control groups. Mild papillary infolding of the 

epithelium was sometimes present within the follicle. 

Cellular atypia and evidence of local invasion were 

notpresent. No effectonthyroidglandneoplasia 

was observed in mice. 

Incidences of Nonneoplastic Lesionsof the Thyroid Gland in Mice in the 2-YearFeed Study of Codeine 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

Male 

15-Month Interim Evaluation 

Follicular Cell Hyperplasiaa 

Number Examined Microscopically 

0 

10 

0 

10 

0 

10 10 

6**(1 .2)b 

2-Year Study 

Follicular Cell Hyperplasia 

Number Examined Microscopically 49 

7 (1.1) 25**(1.2) 

50 

29**(1.3) 

50 

34**(1.5) 

50 

Female 

15-MonthInterimEvaluation 

Number Examined Microscopically 10 9 9 9 

Follicular Cell Hyperplasia 0 0 1 (1.0) 1 (1.0) 

2-Year Study 

Number Examined Microscopically 48 51 51 50 

Follicular Cell Hyperplasia 14(1.4) 29**(1.6) 42**(1.7) 44**(1.9) 

** Significantly different (PSO.01) from the control group by the Fisher exact test (interimevaluation) or the logistic regression test 

(2-year study) 

a Number of animalswithlesion 

Average severity grade of lesion in affected animals: l=minimal, 2=mild, 3=moderate, 4=marked 
54 

Liver:Theincidenceofcentrilobularfattychange, 

which normally occurs in the liver of ad libitum-fed 

mice,wassignificantlylowerin3,000ppmmales 

than in the controls at 15 months, and the decreased 

incidenceappearedtoberelatedtoexposurelevel 

(Tables16andC5).At 2 years,theincidences of 

eosinophilic focus, foci of fatty change, centrilobular 

cytomegaly,andcentrilobularchange fatty in 

3,000 ppm maleswerelowerthanthoseinthe 

controls,andtheincidence of clear cellfocusin 

3,000 ppm males was significantly lower than that in 

thecontrols.Similareffectsintheincidences of 

nonneoplasticliverlesionswerenotobservedin 

female mice. There were exposure-related decreases 

in the incidences of hepatocellular adenomas and the 

TABLE16 

Codeine, NTP TR 455 

incidences of hepatocellular adenomas or carcinomas 

(combined) in males and females, and the incidences 

in the 3,000 ppm groups were lower than those in the 

controls(Tables16, C3, and D3).However,the 

incidence of hepatocellular adenomas or carcinomas 

(combined) in all groups of males and females were 

inthemid-rangeofthehistoricalcontrols(males, 

10%-68%; females, 3%-56%; Tables 16, C4,

and D4). Decreasedincidencesofhepatocellular 

neoplasmshavebeenassociatedwithlowbody 

weightsin studies. the previous Becauselow 

neoplasm incidences in3,000 ppm males and females 

areassociatedwithreducedbodyweighteffectsin 

this study, there is no clear anticarcinogenic effect. 

Incidences of Neoplasms and NonneoplasticLesions of the Liver in Mice in the 2-YearFeed Study 

of Codeine 

Male 

15-MonthInterimEvaluation 

Number Examined Microscopically 

Eosinophilic Focusa 

Foci of Fatty Change 

Centrilobular Cytomegaly 

Centrilobular Fatty Change 

2-Year Study 

Number Examined Microscopically 

Clear Cell Focus 

Eosinophilic Focus 

Foci of Fatty Change 

Centrilobular Cytomegaly 

Centrilobular Fatty Change 

Hepatocellular Adenoma 

Overallrate' 

Hepatocellular Carcinoma 

Overallrate 

Hepatocellular Adenoma or Carcinomad 

Overallrate 

Adjusted ratee 

Terminal ratef 

First incidence (days) 

Logistic regression testg 

(continued) 

~~~~ 

-

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

10 10 10 10 

1 0 0 0 

0 0 0 0 

0 0 1 (l.O)b 0 

10 (1.4) 10 (1.5) 6 (1.0) 1**(1.0) 

50 50 50 50 

7 4 2 0** 

8 12 5 4 

4 4 0 0 

3 

3 

(2.0) 

(1.0) 

8 

0 

(1.1) 0 

0 

0 

0 

23/50 (46%) 25/50 (50%) 16/50 (32%) 10/50** (20%) 

13/50 (26%) 8/50 (16%) 13/50 (26%) 7/50 (14%) 

29/50 (58 %) 29/50 (58%) 23/50 (46%) 16/50 (32%) 

63.0% 62.8% 47.9% 34.0% 

24/41 (59%) 21/38 (55%) 20/45 (44%) 12/43 (28%) 

497 396 635 500 

P=0.002N P=O.567 P=0.118N P=0.007N 
Results 55 

TABLE16 

Incidences of Neoplasms and NonneoplasticLesions of the Liver in Mice in the 2-Year Feed Study 

of Codeine (continued) 

Female 

2-Year Study 

Number Examined Microscopically 

Clear Cell Focus 

Eosinophilic Focus 

Foci of Fatty Change 

Hepatocellular Adenoma 

Overall rate 

Hepatocellular Carcinoma 

Overall rate 

Hepatocellular Adenomaor Carcinomah 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Logistic regression test 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

50 51 51 50 

0 0 1 0 

6 7 9 2 

0 0 1 0 

12/50 (24%) 10151 (20%) 13/51 (25%) 5/50* (10%) 

5/50 (10%) 6/51 (12%) 6/51 (12%) 4/50 (8 %) 

16/50 (32%) 15/51 (29%) 15/51 (29%) 8/50 (16%) 

40.3% 37.3% 34.9% 21.4% 

13/36 (36%) 11/36 (31 %) 15/43 (35%) 6/35 (17%) 

497 616 731 720 

P=0.028N P=0.520N P=0.413N P=0.047N 

* Significantly different(PsO.05) from the control group by the Fisher exact test (interim evaluation) or the logistic regression test 

(2-year study)

** P50.01 

a Number of animalswithlesion 

Average severity grade of lesion in affected animals: l=minimal, 2=mild, 3=moderate, 4=marked 

Number of animals with neoplasm per number of animals with liver examined microscopically 

Historical incidence for 2-year NTP feed studies with untreated control groups (mean f standard deviation): 509/1,316 

(38.7%f 13.9%);range 10%-68% 

e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence inanimals surviving until the end of the study 

In the control column are the P values associated with the trend test. In the exposure group columns are the P values corresponding to 

the painvise comparison between the controls and that exposure group. The logistic regression test regards lesions in animals dying 

prior to terminal kill as nonfatal. A negative trend or a lower incidence in an exposure group is indicated by N. 

Historicalincidence: 260/1,312 (19.8% f 12.8%);range 3%-56% 

Other Organs: The incidences of malignant 

lymphomawere lower in exposedmalesthanin 

controls (0 ppm, 7/50;750ppm, 1/50; 1,500ppm, 

1/50; 3,000 ppm, 0/50;Table C3) but not in exposed 

females (7150, 5/51,6/51, 8/50; Table D3). Except 

in 3,000ppmmales, the incidences ofmalignant 

lymphomas in the control and exposed groups of 

males were within the historical control range from 

NTP feed studies (2% to 24%; Table C4b); therefore, the decreasedincidencescouldnot be clearly 

attributed to codeine exposure. 

By statistical analysis, the incidences of nonneoplastic 

lesions at severalsites in 3,000ppmmales and 

femalesweresignificantly less than those of the 

controls. These included mineralization of the brain 

(males: 33/50, 21/50, 18/50; 14/50,females: 23/50, 

18/51,21/51, 10150;Tables C5 and D5), chronic 

inflammationofthe preputial gland (25/50, 22/50, 

23/50,15/50), chronic inflammation of the salivary 

glandsinmales (22150, 21/50, 15/50, 13/50), 

chronic inflammation ofthe clitoral gland (3/41, 

4/43, 5/45,10/44), and hyaline degeneration 
56 Codeine, NTP TR 455 

(cytoplasmic alteration) of the olfactory epithelium in 

females (27/50, 32/51, 27/50, 41/50). 

GENETICTOXICOLOGY 

Codeine phosphate (100 to 10,000 pg/plate) was not 

mutagenic in Salmonella typhimurium strains TA97, 

TA98, TA100, or TA1535, with or without induced 

liver S9 enzymes (Table El ; Zeiger et al., 1992). 

Dose-relatedincreasesinthefrequencyof sister 

chromatid exchanges wereinducedbycodeine 

phosphate in cultured Chinese hamster ovary cells, 

with and without S9 (Table E2), but these 

significantly increased frequencies were noted only at 

doses that caused marked cell cycle delay, indicative 

of a high levelofcytotoxicity.The role of 

confounding factors, such as toxicity-related changes 

in DNA metabolism, needs to be considered in the 

evaluation of cytogenetic data. Slowlycyclingcells 

undergo prolonged exposure to 5-bromodeoxyuridine, 

which can also result in an increased level of sister 

chromatidexchanges. A clear association between 

cytotoxicity and sister chromatid exchange frequencieshasnotbeen shown. There are numerous 

examples of chemicalsthatinducecell cycle delay 

without inducing sister chromatid exchanges

(Galloway et al., 1987). Acidic pH shifts were noted 

in cultures exposed to higher concentrations of 

codeine phosphate; in some trials, addition of buffer 

N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic 

acid)stabilized the pH. Sister chromatid exchange 

frequencies did not appear to be affected by the pH 

shifts; increases were noted both in acidic and neutral 

cultures. Noincreasesinthe frequency of chromosomal aberrations were noted in cultured Chinese 

hamster ovary cells treated withcodeine phosphate at 

dosessimilar to those used in the sister chromatid 

exchange test (Table E3); codeine phosphate-induced 

cell cycle delay was also observed in these cultures. 
57 

DISCUSSION AND CONCLUSIONS 

Codeine (methylmorphine) is used orally and parenterally as an analgesic and as a respiratory depressant.Two-yearstudieswereconductedbecause no 

previous carcinogenesis bioassays of codeineor other 

opioid compounds had beenreported in the literature. 

Codeine, codeine phosphate, and codeine sulfate are 

available in tablets of 15 to 60 mg, and are usually 

taken four times a day (60 to 240 mglday or 1 to 

4 mglkg per day for a 60 kg person). 

In the 14-day study, all rats receiving 25,000 ppm 

died and finalmeanbodyweightsof 12,500 ppm 

males and females were 62 % and 80% those of the 

controls, respectively. There were no chemicalrelated .deaths or histopathologic lesions in mice in 

the 14-day study; however, final mean body weights 

of 12,500 ppm males and females were90%those of 

the controls. 

Although the cause of death in the 25,000 ppm rats 

could not be determined, the central and peripheral 

neuroendocrine effects of the chemical are believed 

to be important factors in the events leading to the 

animals’ death. Endogenous and codeineits

metabolite, morphine, are found in various tissues of 

the rat, includingthe brain, spinal cord, heart, 

adrenal glands, and liver (Donnerer et al., 1986). 

0-Demethylation of codeine to morphine has been 

demonstrated in rat brain homogenates and it has 

been suggested that morphine formation in the brain 

(or transfer of morphine across theblood-brain 

barrier) may be an important mechanism for codeine 

pharmacologic effects (Chen et al., 1990). This 

hypothesis is further supported by the low affinity of 

codeine for the p-opiate receptor whencompared 

with that of morphine (Pert and Snyder, 1973; Shah 

and Mason, 1991). 

In the 13-week studies, there were no chemicalrelated deaths, histopathologic lesions, or reproductive effects (as measured bythe sperm morphology 

and vaginal cytology protocol) in either rats or mice. 

Absoluteandrelative adrenal gland weights of all 

groups of exposedmale rats andof 3,125 and 

6,250 ppm female rats were significantly greater than 

those of the controls. Body weights in rats exposed 

to3,125or6,250 ppmandinmiceexposed to 

6,250 ppm were lower than those of the controls, and 

reduced body weights have been reported previously 

as a codeine-related effect (Thornhill et al., 1978). 

Based on the body weight effects, the high doses 

selected for the 2-year studies were 1,600 ppm for 

rats and 3,000 ppm for mice. 

One of the characteristicsof the physiological and 

behavioral effects of repeated exposure to codeine or 

morphine drugs is tolerance, which is measured as an 

increase over time in the amount of drug needed to 

produce an analgesic effect (Dambisya et al., 1991). 

Tolerance to the body weight effects of codeine have 

not been reported previously (Koga, 1976; Thornhill 

et al., 1978; Suzuki et al., 1984) and were not seen 

in the current 2-year studies. 

In the 2-year feed studies of codeine in rats and 

mice, there were no chemical-related effects .on 

survival. Mean body weights of 800 and 1,600 ppm 

male and female rats ranged from 4% to 16% lower 

thanthoseofthe controls throughout most of the 

study. Mean body weights of 1,500 and 3,000 ppm 

mice ranged from 1% to 16% lower than those of the 

controls in males and from 5% higher to 21 % lower 

than controls in females. Feed consumption by 

exposed groups was similar to that by the controls 

(Table 17). 

Codeine metabolism in humans and rodents follows 

the same metabolic pathways, including conjugation 

with glucuronic acid, 0-demethylation to morphine 

(Mikus et al., 1991, 1994), N-demethylation to norcodeine, and conjugation ofmorphine and norcodeine 

with glucuronic acid (Cone et al., 1979; Chen et al., 

1991). The amount of each metabolite varies across 

and within species (Hanioka et al., 1990; Mortimer 

et al., 1990; Yue et al., 1990). The exposure levels 

used in the rat studies gave plasma codeine levels that 

overlapped plasma levels for codeine reported in humans (Yuan et al., 1994; Appendix M). O-Demethylation to morphine is a minor pathway and conjugationofcodeine is a majorpathwayinhumans. 
58 Codeine, NTP TR 455 

Comparison of Doses in Codeine 2-YearFeed Studiesa 

TABLE17 

Males Females 

Rats 

Dose in ppm 0400 1,600800 1,600 0 400800 

Grams feedlday 16.4 16.8 18.5 12.1 12.2 16.5 13.1 11.5 

mg codeinelkg body weight 

mg/mz 

. Mice 

Dose in ppm 

Grams feedlday 

mg codeinelkg body weight 

mglm’ 

Humans (Male or Female) 

1-4 mglkg 

37-148mglm’ 

0 14 30 73 0 18 39 70 

0 73 156 380 0 91 203 364 

0 1,500 3OOO750 0 750 1,500 3,OOO

4.7 4.9 5.1 5.0 5.2 5.5 5.6 5.5 

0 I2 156 337 0 81 174 389 

0 216 468 1,011 0 243 522 1,167 

a The dose is calculated as an average at 1 year. Calculation for body surface area dose based on Freireich et al., 1966;mg/m’ = K,,,x 

(dose in mglkg) where K,,,is 37 for humans, 5.2 for rats, and 3.0 for mice. (K,,, is a conversion based on average height-to-body-weight 

ratios.) 

Morphineplasmalevelsinmicewerealsogreater 

than those found in humans, and codeine and morphine plasma levels generally overlapped in rats and 

mice atthedosesusedintheNTPstudies(NTP, 

unpublished data). 

Exposuretocodeineincreasedtheadrenalgland 

weights in male ratsat 15 months;however,the 

mechanism for this effect is not clear. Dose-related 

increases in adrenal gland weight were also present 

at 13 weeks in male and female rats, but males were 

morestronglyaffected.There wasnomorphologic 

alteration(hyperplasia,hypertrophy)toexplainthe 

increased adrenal gland weight in exposed males. It 

is possible that the increase in adrenal gland weight 

may have been related to vascular dilatation and/or 

hypertrophy of cells of the inner cortex, which is not 

evidentfollowingtissueprocessing.Theredid 

appear to be a decrease in the amount of medullary 

tissuerelative to cortextissueinadrenalgland 

sections. This findingwasconsistentwithe 

findings at 2 years;theincidences of proliferative 

lesions of the adrenal medulla in exposed male rats 

were significantly lower than those in the controls. 

Historically,enhancedcellproliferation in the 

adrenalmedullahas beenassociated withincreased 

incidences of adrenalglandneoplasms(Tischler 

et al., 1991). Theadrenalglandnormallycontains 

thehighestendogenoustissueconcentration of 

codeine in therat(Donnerer et al., 1987). Opioid 

peptide immunostaining is seen in some cells of the 

adrenalmedullabutthisexpressionisenhancedin 

human pheochromocytomas (Bostwicket al., 1987). 

Thehighexogenouscodeinelevelsinrats in this 

study may have resulted in a decreased requirement 

or expression of endogenous opioidpeptidesand 

possibly was associated with the decrease in spontaneous proliferative lesions at this site. 

The incidences of thyroid gland follicular cell hyperplasiaweresignificantlygreaterthanthoseinthe 

controls in3,000 ppm male mice at15 months and in 

all exposed groups of males and females at 2 years; 

however,therewerenochemical-relatedthyroid 
Discussion and Conclusions 59 

gland in Other have neoplasms studies mice. 

indicated that morphine can suppress the release of 

thyrotrophic hormone-releasing (which hormone

induces release of thyroid stimulating hormone) and 

reduceserumthyroidstimulatinghormonelevels 

under resting conditions at 15 and 50 mg/kg in the 

rat (Ruzsis and Mess, 1983), but a similar effect was 

not reported in mice given daily500 pg injections for 

5 days (approximately 20 mg codeine or morphine/ 

kg)(Redding et al., 1966). Differencesinplasma 

levels of codeine or its metabolites between rats and 

mice do not appearto completely explainthis thyroid 

effect, and differences in brain levels might provide 

ever,thereducedbodyweightsdonotcompletely 

explain the decreased incidences of mammary gland 

andadrenalglandneoplasmsinratsinthisstudy. 

Forexample,themaximummeanbodyweight 

achieved by female rats in the 1,600 ppm group in 

this study averaged 309 g (Table 6). There are four 

untreated control groups in the current NTP historical 

control database with similar maximum mean body 

weights(averaging 307 g). Themammarygland 

tumor incidence in these historical control groups is 

35% (71/201),compared with the rate of16% (8/51) 

observed in the1,600 ppm codeine group. Similarly, 

theSeilkop (1995) logisticmodelpredicts a mamone basis for the difference in species response in themary gland tumor incidence of37%for controls with 

thyroid. There was no treatment-related increase for 

proliferative lesions of the pituitary gland in male or 

female mice. 

Theincidence of cytoplasmicalteration ofthe 

olfactoryepitheliumwasincreasedinexposedrats 

and 3,000 ppmfemalesmiceafter 2 years.This 

lesion may be consideredanadaptiveresponseto 

chemicalexposure(Monticello et al., 1990). The 

cause of this effect in the current studywas not clear; 

however,it was probably notrelatedtothelocal 

irritatiodinhalation effect. While codeine is used as 

a depressant for the respiratory system inman and to 

inhibitcoughandreflexbronchoconstriction,there 

havebeennoreportsintheliterature to suggest 

codeine-related lesions of the respiratory system after 

codeineadministrationtohumans(Karlsson et al., 

1990; Shook et al., 1990; Biagini et al., 1992). In 

the guinea pig model, interaction of codeine (rather 

thanitsmetabolites)withreceptorsinthetracheobronchialtree mayberesponsible for inhibition of 

the cough reflex (Karlsson et al., 1990). 

Inthecurrent 2 yearstudies,theincidences of 

52-week body weight and survival rates equivalent to 

thoseobservedinthefemaleratgroupreceiving 

1,600 ppm codeine. 

Theadrenalglandneoplasmswerereducedinall 

exposed groups of male rats, even at exposure concentrations where there were no significant decreases 

inbodyweight.Thesedecreasedneoplasmincidences may have been due in part to the pharmacologiceffectsofcodeine,andhavebeenseenpreviously with other chemicals having neuroendocrine 

effects (such as dl-amphetamine sulfate and methylphenidatehydrochloride)(NTP, 1991; NTP, 1995). 

Thedecreasedincidences of hepatic neoplasmsin 

3,000 ppmmicemayhavebeenrelatedtothe 

decreasedbodyweights (10% to 15 % lowerthan 

those of the controls) that were seen during the last 

year of thestudy.Rous (1914) first observed that 

tumor growth is retarded in animals consuming less 

feed, with concomitantreducedbodyweightgain. 

The relationship between feed restriction and tumor 

growthhasbeeninvestigatedusingtransplantedor 

inducedtumors(Sylvester et al., 1981; Grossand 

includingmammaryglandfibroadenomainfemale Dreyfuss, 1984; Ershler et al., 1986) or by studying 

rats,adrenal gland pheochromocytomain malerats the effects of feed restriction on the development of 

several spontaneously occurring neoplasms decreased, 

andhepatocellularneoplasmsinmaleandfemale 

mice. 

It has been reported that reduced body weight gain is 

closelyassociatedwithreducedneoplasmincidence 

at certain sites in rodents (Schneider and Reed,1985; 

Rao et al., 1987; NTP, 1996); reduced body weight 

and feed restriction are also associated with reducing 

theincidenceofchemicallyinducedcancer. How-ofthe 

naturally occurring tumors. A lifespan study in SPF 

Fischer 344 rats, in which the feed restricted group 

received 60%ofthefeedreceivedbythe controls, 

resulted in decreased incidences of testicular interstitialcelltumors,bileducthyperplasia,myocardial 

fibrosis,andmyocardialdegeneration(Yu et al., 

1982). It is estimatedthatthefinalbodyweightof 

the feed-restricted groupwas approximately 70% that 

controls. 
60 Codeine, NTP TR 455 

In a study using Charles River rats, dietary restric- CONCLUSIONS 

tionoverthelifespan oftheanimalsresultedin Undertheconditionsofthese2-yearfeedstudies, reduced incidences of benign mesenchymal neoplasms therewas no evidence of carcinogenic activity* of

andneoplasmsofepithelialtissue(lung,pancreas, 

pituitarygland,andthyroid gland)(Ross andBras, 

1971). A 20% restriction of feedinWistarratsor 

Swissmice for 24 monthscauseddecreasesinthe 

incidencesofneoplasmsofthemammarygland, 

pituitary gland, andskin in rats and pituitary gland in 

mice (Tucker, 1979). Spontaneous lymphomas were 

inhibited by feed restriction in mice (Weindruch and 

Walford,1982). 

Codeine is nongenotoxic in theSalmonella assay and 

in other genotoxicity tests, andinthesecarcinogenesis bioassays there was no evidence that codeine 

had carcinogenic activity in either rats or mice. 

codeineinmaleorfemale F344/Nratsexposedto 

400, 800, or 1,600 ppm. There was no evidence of 

carcinogenic activity of codeine inmaleorfemale 

B6C3F, mice exposed to 750, 1,500, or 3,000 ppm. 

Thyroidglandfollicularhyperplasia cell was 

increased in exposed male and female mice. 

Decreased incidences of benign pheochromocytomas 

of the adrenalmedullainmaleratsandmammary 

glandfibroadenomasandfibroadenomas or adenocarcinomas (combined) in female rats were related to 

codeine exposure. 

* Explanation of Levels of Evidence of CarcinogenicActivityisonpage 9. A summary of theTechnicalReportsReviewSubcommittee 

comments and the public discussion on this Technical Report appears on page 11. 
61 

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Pharmacol.Biochem.Behav. 9, 433-438. 

the enterohepatic circulation of morphine in the rat. 

J. Pharmacol.Exp.Ther. 195, 303-310. 

Weindruch, R., and Walford,R.L. (1982). Dietary 

restriction in mice beginning at1 year of age: Effect 

Science 215, 1415-1418. 

onlife-spanandspontaneouscancerincidence. 

Metab.Rev. 12, 261-277. 

Excretion of drugs inhumanbreastmilk. 

Welch, R.M., and Findlay, J.W.A. 

Drug 

(1981). 

Willette, R.E. (1991). Analgesicagents.In Wilson 

and Gisvold's Textbook of OrganicMedicinaland 

PharmaceuticalChemistry (J.N. Delgado and 

W.A. Remers, Ms.), 9th ed., pp. 629-646. 

J.B. Lippincott Company, Philadelphia. 
References 69 

Williams, D.A. (1971). A test for differences 

betweentreatmentmeanswhenseveral dose levels 

are compared with a zero dose control. Biometrics 

27,103-1 17. 

Williams, D. A. (1972). The comparison of several 

dose levels with a zero dose control. Biometrics 

28,519-531. 

Williams, J., Price, C.J., Sleet, R.B., George, J.D., 

Marr, M.C., Kimmel, C.A., and Morrissey, R.E. 

(199 1). Codeine: toxicity Developmental in 

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401-413. 

Woods, L.A., Muehlenbeck, H.E., and Mellett, L.B. 

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metabolitesinthedogandmonkey. J. Pharmacol. 

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Yeh, S.Y., and Woods, L.A. (1969). Physiologic 

disposition of N-C'4-methyl-codeineinthe rat. 

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Yeh, S.Y., and Woods, L. A. (1970). Isolation and 

characterization of codeine-6-glucuronide from dog 

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Yeh, S.Y., and Woods, L.A. (1971). Excretion of 

codeine and its metabolites by dogs, rabbits and cats. 

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Yoshimura, H., Mori, M.-A., Oguri, K., and 

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Yu, B.P., Masoro, E.J., Murata, I., Bertrand, H.A., 

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71 

APPENDIX A 

SUMMARY OF LESIONS IN MALE RATS 

IN THE 2-YEAR FEED STUDY 

OF CODEINE 

TABLEA1 Summary of the Incidence of Neoplasms in Male Rats 

in the 2-Year Feed Study of Codeine ................................. 73 

TABLEA2 Individual Animal Tumor Pathology of Male Rats 

TABLEA3 Statistical Analysis of Primary Neoplasms in Male Rats 

TABLE A4 Historical Incidence of Benign Pheochromocytomas 

TABLEA5 Summary of the Incidence of Nonneoplastic Lesions in Male Rats 

in the 2-Year Feed Study of Codeine ................................. 78 

in the 2-Year Feed Study of Codeine ................................. 94 

of the Adrenal Medulla in Untreated Male F344lN Rats ..................... 98 

in the 2-Year Feed Study of Codeine ................................. 99 
Lesions in Male Rats 73 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of Codeinea 

Disposition Summary 

Animals initially in study 

PS-Month interim evaluarion 

Early deaths 

Moribund 

Natural deaths 

Survivors 

Terminal sacrifice 

Animals examined microscopically 

B%Month Interim Evaluation 

AlimentarySystem 

Intestine large, colon 

Polyp adenomatous 

Liver 

Mesentery 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Pheochromocytoma benign 

Pituitary gland 

Pars distalis, adenoma 

Genital System 

Epididymis 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

RespiratorySystem 

Lung 

Alveolar/bronchiolar adenoma 

SystemicLesions 

Multiple organsb 

Leukemia mononuclear 

Mesothelioma malignant 

0 PPm 400 ppm 

60 60 

10 10 

14 23 

7 7 

29 20 

60 60 

(10) (10) 

(10) (10) 

2 (20%) 6 (60%) 

4 (40%) 4 (40%) 

(10) (9) 

1 (10%) 

800 ppm 

60 

10 

24 

5 

21 

60 

(10) 

(10) 

3(30%) 

4 (40%) 

1,600 ppm 

60 

10 

23 

7 

20 

60 

(10) 

(10) 

8 (80%) 

1 (10%) 
74 Codeine, NTP TR 455 

TABLEA1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

15-Month Interim Evaluation (continued) 

Systems Examined With No Neoplasms Observed 

Cardiovascular System 

General Body System 

HematopoieticSystem 

IntegumentarySystem 

Musculoskeletal System 

Nervous System 

Special Senses System 

Urinary System 

2-Year Study 

Alimentary System 

Esophagus 

Intestine large, colon 

Polyp adenomatous, multiple 

Intestine large, rectum 

Intestine large, cecum 

Adenoma 

Intestine small, duodenum 

Sarcoma, metastatic, mesentery 

Intestine small, jejunum 

Adenocarcinoma 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 

Sarcoma, metastatic, mesentery 

Mesentery 

Sarcoma 

Pancreas 

Sarcoma, metastatic, mesentery 

Salivary glands 

Stomach, forestomach 

Sarcoma, metastatic, mesentery 

Stomach, glandular 

Tongue 

Squamous cell papilloma 

Cardiovascular System 

Blood vessel (49) 

Heart (50)

Alveolarlbronchiolar carcinoma, metastatic, 

1 (2%) lung 
Lesions in Male Rats 75 

TABLEA1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of Codeine (continued) 

400 ppm 800 ppm 1,600 ppm 

&Year shdy (continued) 

Endocrine System 

Adrenal medulla 

Adrenal cortex 

Pheochromocytoma malignant 

(50) 

(49) 

1 (2%) 

Pheochromocytoma complex 1 (2%) 

Pheochromocytoma benign 13 (27%) 6 (12%) 3 (6%) 

Bilateral, pheochromocytoma malignant 

Bilateral, pheochromocytoma benign 3 (6%) 

Adenoma 

Islets, pancreatic (50) 

Parathyroid gland 

Carcinoma, metastatic, thyroid gland 

Pars distalis, adenoma 

Pituitary gland (48) 

6 (13%) 

Pars intermedia, adenoma 

Bilateral, C-cell, adenoma 

Thyroid gland (50) 

1 (2%) 

C-cell, adenoma 1 (2%) 

C-cell, carcinoma 3 (6%) 

Follicular cell, adenoma 

Follicular cell, carcinoma 1 (2%) 

General Body System 

None 

GenitalSystem 

Penis 

Epididymis (49) 

Preputial gland 

Adenoma 

(50) 

3 (6%) 

(50) 

6 (12%) 

(50) 

2 (4%) 

Carcinoma 

Bilateral, adenoma 

Prostate 

Seminal vesicle 

Sarcoma, metastatic, mesentery 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

HematopoieticSystem 

Bone marrow 

Pheochromocytoma malignant, metastatic, 

(50) 

adrenal medulla 

Lymph node 

Mediastinal, alveolarhronchiolar carcinoma, 

(20) 

metastatic, lung 1 (6%) 

Mediastinal, sarcoma, metastatic, mesentery 

Lymph node, mesenteric 

Lymph node, mandibular 

(50) 

(50) 
76 Codeine, NTP TR 455 

TABLEAI 

SmmaO' of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

2-Year Study (continued) 

Hematopoietic System (continued) 

Spleen (50)

Sarcoma, metastatic, mesentery 

Thymus (46) 

IntegumentarySystem 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Lipoma 

Squamous cell carcinoma 1 (2%) 

Trichoepithelioma 1 (2%) 

Foot, squamous cell papilloma 1 (2%)

Lip, basosquamous tumor benign 1 (2%) 

Subcutaneous tissue, fibroma 5 (10%) 2 (4%) 3 (6%) 1 (2%) 

Subcutaneous tissue, sarcoma 1 (2%) 

Tail, squamous cell papilloma 1 (2%) 

Musculoskeletal System 

Bone (50) (50) 

Rib, osteosarcoma 

Skeletal muscle (1) 

Abdominal, sarcoma, metastatic, mesentery 

Back, lipoma 

Thoracic, alveolarhronchiolar carcinoma, 

metastatic, lung 1 (100%) 

Nervous System 

Brain 

Astrocytoma malignant 

Meningioma malignant 

Spinal cord 

Hemangioma 

RespiratorySystem 

Lung (50)

Alveolarhronchiolar adenoma 1 (2%) 

Alveolarhronchiolar carcinoma 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 

Sarcoma, metastatic, mesentery 

Schwannoma malignant, metastatic, ear 

Mediastinum, alveolarhronchiolar carcinoma, 

metastatic, lung 

Nose 

Adenoma 

Chondroma 

Trachea 
Lesions in Male Rats 77 

TABLEA1 

Summary of the Incidence of Neoplasms in MaleRats in the 2-YearFeed Study of Codeine (continued) 

400 ppm 800 ppm 

2-Year Study (continued) 

Special Senses System 

Ear 

Schwannoma malignant Pinna.schwannoma malignant 

Urinary System 

Kidney (50) (50)

Sarcoma, metastatic, mesenteq 

Renal tubule, adenoma 

Renal tubule, carcinoma 

Urinary bladder (50) (49) 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

Neoplasm Summary 

Total animals with primary neoplasms' 

15-Month interim evaluation 7 10 9 

2-Year Study 50 50 50 

Total primary neoplasms 

15-Month interim evaluation 9 12 10 

2-Year study 131 117 127 

Total animals with benign neoplasms 

15-Month interim evaluation 7 10 7 

2-Year study 49 47 50 

Total benign neoplasms 

15-Month interim evaluation 8 12 8 

2-Year study 91 81 80 

Total animals with malignant neoplasms 

15-Month interim evaluation 1 2 

2-Year study 36 33 39 

Total malignant neoplasms 

15-Month interim evaluation 1 2 

2-Year study 40 36 47 

Total animals with metastatic neoplasms 

2-Year study 2 2 

Total metastatic neoplasms 

2-Year study 5 4 

a Number of animals examined microscopically at the siteand the number of animals with neoplasm 

Number of animals with any tissue examined microscopically 

primary neoplasms: all neoplasms except metastatic neoplasms 

1,600 ppm 

9 

50 

10 

115 

9 

48 

10 

66 

41 

49 

4 

13 
78 Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 0 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Polyp adenomatous, multiple 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tongue 

Squamous cell papilloma 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma malignant 

Pheochromocytoma complex 

Pheochromocytoma benign 

Bilateral, pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell, carcinoma 

General Body System 

None 

+: Tissue examined microscopically 

A:Autolysisprecludesexamination 

2344455555666666666777777 

0344902569034556788023333 

3469505479163272014002222 

0000000000000000000000000 

1001420110325554350200111 

4125688204705063189257179 

......................... 

......................... 

+++++++++++++++++A+++++++ 

A+++++++++++++++A+++++++ 

A++++A+++++++++++++++++++ 

A++++A+++++A+++++A+++++++ 

+++++A+++++A+++++A+A+++++ 

......................... 

+ + + ++ ......................... 

......................... 

......................... 

......................... + 

X 

......................... 

......................... 

......................... 

......................... X 

X x x xx x x 

......................... 

+++M+M+++++M+++MM+M++++MM 

......................... X X 

......................... X 

M:Missingtissue X: Lesionpresent 

I: Insufficienttissue Blank:Notexamined 
Lesions in Male Rats 79 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Polyp adenomatous, multiple 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tongue 

Squamous cell papilloma 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma malignant 

Pheochromocytoma complex 

Pheochromocytoma benign 

Bilateral, pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell,. carcinoma 

General Body System 

None 

1111111111111111117111111 

3333333333333333333333333 

2222222222222222333333333 

0000000000000000000000000 Total 

2222333444445556012233345 Tissues/ 

1349068012582390685625991 Tumors 

......................... 50 

......................... 50 

X 1 

......................... 49 

......................... 41 

......................... 48 

......................... 46 

......................... 46 

......................... 50 

X 1 

X 1 

X 1 

++ + + + 10 

......................... 50 

......................... 50 

......................... 50 

........................ 49 

1 

1 

++++++++++M++++++++++++++ 49 

......................... 50 

......................... 50 

++++++++++M++++++++++++++ 49 

1 

X 1 

X X xx X X 13 

X X X 3 

......................... 50 

M+M++++M+++M+++++++++++++ 38 

......................... 50 

X X xxx xx 9 

......................... 50 

X X 3 

X X 2 

X 1 
Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Genital System 

Epididymis 

Preputial gland 

Adenoma 

Prostate 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

IntegumentarySystem 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Lip, basosquamous tumor benign 

Subcutaneous tissue, fibroma 

Musculoskeletal System 

Bone U 

Nervous System 

Brain 

Meningioma malignant ~_____ 

Respiratory System 

Lung 

Alveoladbronchiolar adenoma 

Nose 

Trachea 

Special Senses System 

EY e 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

2344455555666666666777777 ~ ~~ 

0344902569034556788023333 

3469505479163272014002222 

0000000000000'000000000000 

1001420110325554350200111 

4125688204705063189257179 

......................... 

.......................... X 

+++++++++M+++++++++++++++ 

......................... 

......................... X x x x x xx x x x x x x x x 

X x x x x xx X 

.. 

......................... ++ +++++ +++ + ++ ......................... 

......................... 

.......................... 

+++M++++++M+++M+++M++++++ 

+,++++M+++M+++++++++M+++++ 

......................... X 

X X 

......................... 

......................... 

X 

......................... X 

......................... 

......................... 

+ 

......................... 

......................... 

......................... xx x xxxxxxxxxxxxxxx 

X 
Lesions in Male Rats 81 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Genital System 

Epididymis 

Preputial gland 

Adenoma 

Prostate 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell. adenoma 

HematopoieticSystem 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

IntegumentarySystem 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Lip, basosquamous tumor benign 

Subcutaneous tissue, fibroma 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Meningioma malignant 

Respiratory System 

Lung 

Alveolarhronchiolar adenoma 

Nose 

Trachea 

Special Senses System 

EYe 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

1111111111111111111111111 

3333333333333333333333333 

2222222222222222333333333 

0000000000000000000000000 Total 

2222333444445556012233345 Tissues/ 

1349068012582390685625991 Tumors 

......................... 50 

......................... 50 

x x 3 

......................... 49 

......................... 50 

......................... 50 

xxxxxxxxxxxxxxxxxxxxxxx 38 

X 9 

......................... 50 

+ + + + + + + 20 

......................... 50 

......................... 50 

......................... 50 

......................... 46 

......................... 41 

X 1 

......................... 50 

1 

X 1 

X X X 5 

......................... 50 

......................... 50 

1 

......................... 50 

1 

......................... 50 

......................... 50 

1 

......................... 50 

......................... 50 

......................... 50 

x xxx xx x xx x x x xx 32

1 
82 Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 400 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Adenoma 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Alveolarhronchiolar carcinoma, 

metastatic, lung 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Adenoma 

Prostate 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

HematopoieticSystem 

Bone marrow 

3445556666666666666666667 

3024780112234555577788990 

6469468554721117888944351 

1101110110010011001000000 

0160118019807611871989899 

1359651487632619070183954 

......................... 

......................... 

+++++++++++A+++++++++++++ 

+++++++++++A+++++++++++++ 

+++A+++++++++++++++++++++ 

+++++++++++++++++++++A+++ 

+++A+++++++A++++++++++A++ 

......................... + + + + + + ......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

X 

......................... 

......................... X 

......................... X X 

+++M++++++++++++++++++MM+ 

......................... X X 

X 

......................... X 

X 

......................... 

......................... X X X 

++++++++++M++++++++++++++ 

......................... 

......................... x x x x xxxxxxxxx 

X xxx x xx 

......................... 
Lesions in Male Rats 83 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 400 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Adenoma 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Alveolar/bronchiolar carcinoma, 

metastatic, lung 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Adenoma 

Prostate 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

HematopoieticSystem 

Bone marrow 

7777777777777777777777777 

0012233333333333333333333 

6710922222222222222333333 

1110000000000000000000001 Total 

lo07666667777778889688991 Tissues/ 

4726314780134584576223292 Tumors 

......................... 50 

......................... 50 

++A++++++++++++++++++++++ 48 

++A++++++++++++++++++++++ 48 

X 1 

......................... 49 

++++A++++++++++++++++++++ 48 

++A++++++++++++++++++++++ 46 

......................... 50 

++++ ++++ + ++ +++ + 21 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

1 

......................... 50 

......................... 50 

x x x x 6 

......................... 50 

X 3 

++++++M++++++++++M+++++++ 45 

......................... 50 

x x 4 

1 

......................... 50 

X X 3 

X X 2 

1 

......................... 50 

......................... 50 

X X X 6 

......................... 49 

......................... 50 

......................... 50 

x x x x x x x x xx x x x x x x x x x x x x x x 39 

X 8 

......................... 50 
84 Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 400 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System (continued) 

Lymph node 

Mediastinal, alveoladbronchiolar 

carcinoma, metastatic, lung 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Keratoacanthoma 

Lipoma 

Foot, squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Skeletal muscle 

Thoracic, alveoladbronchiolar 

carcinoma, metastatic, lung 

Nervous System 

Brain 

Astrocytoma malignant 

Spinal cord 

~ ~~ 

Respiratory System 

Lung 

Alveoladbronchiolar carcinoma 

Schwannoma malignant, metastatic, ear 

Mediastinum, alveoladbronchiolar 

carcinoma, metastatic, lung 

Nose 

Adenoma 

Chondroma 

Trachea 

Special Senses System 

Ear 

Pinna, schwannoma malignant 

EY e 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

3445556666666666666666667 

3024780112234555577788990 

6469468554721117888944351 

1101110110010011001000000 

0160118019807611871989899 

1359651487632619070183954 

+ ++ ++ ++ ++++ 3. + 

X 

......................... 

......................... 

......................... 

+++++++M+++++MM+M++++++M+ 

......................... 

......................... 

X 

X 

......................... 

+ 

X 

......................... 

-t 

......................... 

X 

X 

X 

......................... X 

X 

......................... 

+ 

X 

......................... 

......................... 

......................... x xx xxxxxxxx xxxx x x

X X 
Lesions in Male Rats 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 400 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System (continued) 

Lymph node 

Mediastinal, alveolar/bronchiolar 

carcinoma, metastatic, lung 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

Integumentary System 

Mammary gland 

Skin 

Keratoacanthoma 

Lipoma 

Foot, squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Skeletal muscle 

Thoracic, alveolarhonchiolar 

carcinoma, metastatic, lung 

Nervous System 

Brain 

Astrocytoma malignant 

Spinal cord 

Respiratory System 

Lung 

Alveolar/bronchiolar carcinoma 

Schwannoma malignant, metastatic, ear 

Mediastinum, alveolar/bronchiolar 

carcinoma, metastatic, lung 

Nose 

Adenoma 

Chondroma 

Trachea 

Special Senses System 

Ear 

Pinna, schwannoma malignant 

Eye 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

7777777777777777777777777 

0012233333333333333333333 

6710922222222222222333333 

1110000000000000000000001 Total 

loo7666667777778889688991 Tissues/ 

4726314780134584576223292 Tumors 

+ + + + 17 

1 

+M+++++++++++++++++++++++ 49 

......................... 50 

......................... 50 

M+++++++M+M++++++M+++++++ 41 

++++++++++M++++++++++++++ 49 

......................... 50 

xx 2 

X 1 

X 1 

X 2 

1 

......................... 50 

1 

1 

......................... 50 

X 1 

1 

......................... 50 

X 2 

1 

1 

......................... 50 

X 2 

1 

......................... 50 

1 

1 

+ 1 

......................... 50 

......................... 49 

......................... 50 

xx x X xx x x X X 27 

Mesothelioma malignant 2 
86. Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 800 ppm 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Adenoma 

Intestine small, duodenum 

Intestine small, jejunum 

Adenocarcinoma 

Intestine small, ileum 

Liver 

Pheochromocytoma malignant, 

metastatic, adrenal medulla 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Bilateral, pheochromocytoma malignant 

Bilateral, pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

General Body System 

None 

Genital System 

Epididymis 

Penis 

Preputial gland 

Adenoma 

Carcinoma 

Bilateral, adenoma 

Prostate 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

55555.55566666666666667777 

1245567803333355568890012 

9660974084588858857756720 

1111111111111111111111111 

7562426373533473834476655 

74111887849188'30033603037 

......................... 

......................... 

......................... 

+++++++++A+++++++++++++++ 

......................... 

+++++++++A++++++++++A++++ 

X 

+++++++++A+++++++++++++AM 

......................... 

X 

+ ++++ ......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... X X X 

X 

......................... 

+M++++++++++M+++++M++++M+ 

......................... X X 

......................... 

X 

M++++++++++++++++++++++++ 

+ ......................... X X 

X 

......................... 

......................... 

......................... 

X xxxxxxxxxxxxxxxxxxx x x

X X 
Lesions in Male Rats 87 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 800 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Adenoma 

Intestine small, duodenum 

Intestine small, jejunum 

Adenocarcinoma 

Intestine small, ileum 

Liver 

Pheochromocytoma malignant, 

metastatic, adrenal medulla 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Bilateral, pheochromocytoma malignant 

Bilateral, pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

C-cell. adenoma 

C-cell, carcinoma 

General Body System 

None 

Genital System 

Epididymis 

Penis 

Preputial gland 

Adenoma 

Carcinoma 

Bilateral, adenoma 

Prostate 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

1771711717111111111111111 

2222333333333333333333333 

0266222222222222222333333 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

1426223344556661111223551 Tissues/ 

4031692547055691569279262 Tumors 

......................... 50 

......................... 50 

......................... 50 

......................... 49 

X 1 

........................... 50 

+++A+++++++++++++++++++++ 41 

1 

......................... 41 

......................... 50 

1 

++++++++ +++ + 11 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

x x 5 

1 

X 1 

......................... 50 

++M++++++++M+++++++M+M+++ 42 

++M+++++++++++++++++++++M 48 

x x X X 6 

......................... 50 

X X 2 

X X X 4 

......................... 49 

1 

++M++++++M+++++++++++++++ 48 

X xx 5 

X 1 

X 2 

......................... 50 

......................... 50 

......................... 50 

x xxxxxxxxxxxxxxxxxxxxxxx 46 

X 3 
88 Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 800 ppm (continued) 

5555555566666666666667777 

Number of Days on Study 1245567803333355568890012 

9660974084588858857756720 

1111111111111111111111111 

Carcass ID Number 7562426373533473834476655 

7411188784918830033603037 

HematopoieticSystem 

Bone marrow ......................... Pheochromocytoma malignant, 

metastatic, adrenal medulla X 

Lymph node + + ++ + +++++ + 

Lymph node, mandibular ......................... Lymph node, mesenteric ......................... Spleen ......................... Thymus +++++++++++++++MM++M++++M 

IntegumentarySystem 

Mammary gland ......................... Fibroadenoma X 

Skin ......................... Keratoacanthoma 

Squamous cell carcinoma 

Trichoepithelioma X 

Subcutaneous tissue, fibroma X 

Musculoskeletal System 

Bone ......................... Skeletal muscle 

Back, lipoma 

Nervous System 

Brain ......................... 

Respiratory System 

Lung ......................... Pheochromocytoma malignant, 

metastatic, adrenal medulla X 

Schwannoma malignant, metastatic, ear X 

Nose ......................... Trachea ......................... 

Special Senses System 

Ear + 

Pinna, schwannoma malignant X 

Eye + 

Urinary System 

Kidney ......................... Renal tubule, adenoma 

Urethra + 

+++++++++A+++++++++++++A+ 

Systemic Lesions 

Multiple organs ......................... Leukemia mononuclear xxxxxxxxxxxxxxxxxxxxxx 

Mesothelioma malignant 
Lesions in Male Rats 89 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 800 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Pheochromocytoma malignant, 

metastatic, adrenal medulla 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

IntegumentarySystem 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Squamous cell carcinoma 

Trichoepithelioma 

Subcutaneous tissue, fibroma 

Musculoskeletal System 

Bone 

Skeletal muscle 

Back, lipoma 

Nervous System 

Brain 

Respiratory System 

Lung 

Pheochromocytoma malignant, 

metastatic, adrenal medulla 

Schwannoma malignant, metastatic, ear 

Nose 

Trachea 

Special Senses System 

Ear 

Pinna, schwannoma malignant 

EY e 

Urinary System 

Kidney 

Renal tubule, adenoma 

Urethra 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

1111117111111111111111111 

2222333333333333333333333 

0266222222222222222333333 

1111111111111111111111111 Total 

1426223344556661111223551 Tissues1 

4031692541055691569219262 Tumors 

......................... 50 

1 

+ ++ 14 

+++++++++++++++M+++++++++ 49 

......................... 50 

......................... 50 

+MMM++++++++M+++M++++++++ 41 

......................... 50 

1 

......................... 50 

X X 2 

X 1 

1 

x x 3 

......................... 50 

+ 1 

X 1 

......................... 50 

......................... 50 

1 

1 

......................... 50 

.......................... 50 

1 

1 

t 2 

......................... 50 

X 1 

1 

......................... 48 

......................... 50 

X x x xxxx x xxx x xx 36 

x x 2 
90 Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 1,600 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Sarcoma, metastatic, mesentery 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular adenoma 

Sarcoma, metastatic, mesentery 

Mesentery 

Sarcoma 

Pancreas 

Sarcoma, metastatic, mesentery 

Salivary glands 

' Stomach, forestomach 

Sarcoma, metastatic, mesentery 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Carcinoma, metastatic, thyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

Bilateral, C-cell, adenoma 

C-cell, adenoma 

C-cell, carcinoma 

General Body System 

Tissue NOS 

Genital System 

Coagulating gland 

Epididymis 

Preputial gland 

Adenoma 

Prostate 

Seminal vesicle 

Sarcoma, metastatic, mesentery 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

3345555555555666666667777 

5883344557889025577880000 

5340734091254319900370155 

2222112221112212222211222 

2132983228981491223388101 

5738840391195028746156691 

......................... 

......................... 

......................... 

......................... 

++++++A++++++++++++++++++ 

X 

++++++A++++++++++A+++A+++ 

+++++++++++++++++++++++A+ 

......................... 

X 

+++ + + + + + 

X 

......................... X ......................... 

......................... X 

.......................... 

......................... 

......................... 

......................... 

........................... 

......................... 

++M+++++++++++M++++++++++ 

++++++++++++++.M++++++++++ 

X X 

......................... 

X 

X X 

+ 

++M++++++++++++++++++++++ 

......................... X 

......................... 

......................... X ......................... xxxxxxxxxxxxxxxxxxxxxx 

X 
Lesions in Male Rats 91 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 1,600 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Sarcoma, metastatic, mesentery 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular adenoma 

Sarcoma, metastatic. mesentery 

Mesentery 

Sarcoma 

Pancreas 

Sarcoma, metastatic, mesentery 

Salivary glands 

Stomach, forestomach 

Sarcoma, metastatic, mesentery 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Carcinoma, metastatic, thyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

Bilateral, C-cell. adenoma 

Ccell, adenoma 

Ccell. carcinoma 

General Body System 

Tissue NOS 

Genital System 

Coagulating gland 

Epididymis 

Preputial gland 

Adenoma 

Prostate 

Seminal vesicle 

Sarcoma, metastatic, mesentery 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

7777777777777777777777777 

0123333333333333333333333 

85901222222222223333333-33 

1221212222222222111122222 Total 

8228090001112333899901333 Tissues/ 

7618091470292789206764245 Tumors 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

A++++++++++++++++++++++++ 48 

1 

A++++++++++++++++++++++++ 46 

A++A+++++++++++++++++++++ 47 

......................... 50 

X 1 

1 

+ ++ ++ + + + + 17 

1 

......................... 50 

1 

......................... 50 

......................... 50 

1 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

X xx 3 

......................... 50 

+++++++++M++++++++++++M++ 46 

X 1 

........................ 47 

xx 4 

......................... 50 

X 1 

1 

X 3 

+ 1 

1 

......................... 49 

......................... 50 

x 2 

......................... 50 

......................... 50 

1 

......................... 50 

xxxxxxxxxxxxxxxxxxxxxxxxx 47 

1 
92 Codeine, NTP TR 455 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of Codeine: 1,600 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone.marrow 

Lymph node 

Mediastinal, sarcoma, metastatic, 

mesentery 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Sarcoma, metastatic, mesentery 

Thymus 

IntegumentarySystem 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Subcutaneous tissue, fibroma 

Tail, squamous cell papilloma 

Musculoskeletal System 

Bone 

Rib, osteosarcoma 

Skeletal muscle 

Abdominal, sarcoma, metastatic, mesentery 

Nervous System 

Brain 

Astrocytoma malignant 

Spinal cord 

Hemangioma 

RespiratorySystem 

Lung 

Sarcoma, metastatic, mesentery 

Schwannoma malignant, metastatic, ear 

Nose 

Trachea 

Special Senses System 

Ear 

Schwannoma malignant 

Pinna, schwannoma malignant 

Urinary System 

Kidney 

Sarcoma, metastatic, mesentery 

Renal tubule, carcinoma 

Urinary bladder 

SystemicLesions 

Multiple organs 

Leukemia mononuclear 

3345555555555666666667777 

5883344557889025577880000 

5340734091254319900370155 

2222112221112212222211222 

2132983228981491223388101 

5738840391195028746156691 

......................... + + + + +++++++ ++ 

X 

++++++++++ ++++++++++++++ ......................... 

......................... X 

++++++++MM++++MM+M++++++M 

......................... 

......................... X 

X 

......................... X 

+ 

X 

......................... X 

+ + 

X 

......................... X 

X ......................... 

......................... 

+ 

X 

......................... X 

......................... 

......................... 

X x x x x x x xx x x x x x x x x xx x x 

Mesothelioma malignant X X X 
Lesions in Male Rats 93 

TABLEA2 

Individual Animal Tumor Pathology of Male Rats in the 2-YearFeed Study of Codeine: 1,600 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Lymph node 

Mediastinal, sarcoma, metastatic, 

mesentery 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Sarcoma, metastatic, mesentery 

Thymus 

Integumentary System 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Subcutaneous tissue, fibroma 

Tail, squamous cell papilloma 

Musculoskeletal System 

Bone 

Rib, osteosarcoma 

Skeletal muscle 

Abdominal, sarcoma,metastatic,mesentery 

Nervous System 

Brain 

Astrocytoma malignant 

Spinal cord 

Hemangioma 

RespiratorySystem 

Lung

Sarcoma, metastatic, mesentery 

Schwannoma malignant, metastatic, ear 

Nose 

Trachea 

Special Senses System 

Ear 

Schwannoma malignant 

Pinna, schwannoma malignant 

Urinary System 

Kidney 

Sarcoma, metastatic, mesentery 

Renal tubule, carcinoma 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

7777777777777777777777777 

0123333333333333333333333 

8590122222222222333333333 

1221212222222222111122222 Total 

8228090001112333899901333 Tissues/ 

7618091470292789206764245 Tumors 

......................... 50 

+ ++ ++ +++ + 22 

1 

.......................... 49 

......................... 50 

......................... 50 

1 

M+++++++M++++++++++++++M+ 41 

++M+M++++++++++++++++++++ 48 

X 1 

......................... 50 

X 2 

1 

X 1 

......................... 50 

1 

1 

1 

......................... 50 

1 

2 

1 

......................... 50 

1 

X 2 

......................... 50 

......................... 50 

+ 2 

1 

X 1 

......................... 50 

1 

X 1 

++M++++++++++++++++++++++ 49 

......................... 50 

x x x x x x x x x xx x xxx X 37

3 
- 

- 

- 

- 

- 

- 

- 

- 

94 Codeine, NTP TR 455 

TABLEA3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-YearFeed Study of Codeine 

0 PPm 400 ppm 800 ppm 1,600 ppm 

AdrenalMedulla:BenignPheochromocytoma 

Overall ratea 16/49 (33%) 6/50 (12%) 6/50 (12%) 3/50 (6%) 

Adjusted rateb 49.1 % 24.9% 21.7% 13.6% 

Terminal rate' 12/28 (43%) 3/20 (15%) 3/21 (14%) 2/20 (10%) 

First incidence (days) 599 678 608 708 

Life table testd P=0.005N P=0.070N P=0.051N P=0.007N 

Logistic regression testd P=0.001N P=0.013N P=O.OlON P=0.001N 

Cochran-Armita e testd P<0.001N

Fisher exact test % P=0.012N P=0.012N P<0.001N 

AdrenalMedulla:Benign,Complex,orMalignantPheochromocytoma 

Overall rate 16/49 (33~%) 6/50 (12%) 7/50 (14%) 3/50 (6%) 

Adjusted rate 49.1% 24.9% 24.5% 13.6% 

Terminal rate 12/28 (43%) 3/20 (15%) 3/21 (14%) 2/20 (10%) 

First incidence (days) 599 678 608 708 

Life table test P=0.007N P=0.070N P=0.089N P=0.007N 

Logistic regression test P=0.001N P=0.013N P=0.020N P=0.001N 

Cochran-Annitage test P-0.001N 

Fisher exact test P=0.012N P=O.O24N P<0.001N 

Liver:HepatocellularAdenoma or Carcinoma 

Overall rate 3/50 (6%) 0150 (0%) 0/50 (0%) 1/50 (296) 

Adjusted rate 10.3% 0.0% 0.0% 5.0% 

Terminal rate 3/29 (10%) 0120 (0%) 0/21 (0%) 1/20 (5%) 

First incidence (days) 732 (T) -C - 732 (T) 

Life table test P=0.327N P=0.192N P=0.182N P=0.445N 

Logistic regression test P=0.327N P=O. 192N P=O.l82N P=0.445N 

Cochran-Annitage test P=0.247N 

Fisher exact test P=0.121N P=0.121N P=0.309N 

PancreaticIslets:Adenoma 

Overall rate 0150 (0%) 3/50 (6%) 0150 (0%) 0150 (0%) 

Adjusted rate 0.0% 11.3% 0.0% 0.0% 

Terminal rate 0/29 (0%) 1 /20 (5 %) 0/21 (0%) 0/20 (0%) 

First incidence (days) - 658

Life table test P=0.339N P=O.O94 

Logistic regression test P=0.317N P=O.119 

Cochran-Amitage test P=0.311N 

Fisher exact test P=O.121 

PituitaryGland(Pars Distalis): Adenoma 

Overall rate 9/50 (18%) 4/50 (8%) 6/48 (13%) 4/47 (9%) 

Adjusted rate 29.1 I 14.6% 24.4% 13.0% 

Terminal rate 8/29 (28 46) 1/20 (5%) 4/20 (20%) 0/20 (0%) 

First incidence (days) 449 632 526 594 

Life table test P=0.253N P=O.259N P=0.516N P=0.249N 

Logistic regression test P=0.177N P=O.l16N P=0.307N P=0.151N 

Cochran-Annitage test P=0.167N 

Fisher exact test P=O.l17N P=0.318N P=O.l42N 
Lesions in Male Rats 95 

TABLEA3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400800PPm ppm 1,600 ppm 

PreputialGland:Adenoma 

Overall rate 3/50 (6%) 6/50 (12%) 7/48 (15%) 2/50 (4%) 

Adjusted rate 9.1 % 26.8%20.6% ' 7.4% 

Terminal rate 2/29 (7%) 2/20 (10%) 3/20 (15%) 1/20 (5%) 

First incidence (days) 655 

Life table test P=O.O90 P=O.171 P=0.463N P=0.604N 

Logistic regression test P=0.351NP=O.142 P=O.245 P=0.501N 

Cochran-Armitage test P=0.342N 

Fisher exact test P=O.142 P=O.243 P=0.500N 

PreputialGland:AdenomaorCarcinoma 

Overall rate 3/50 (6%) 6/50 (12%) 8/48 (17%) 2/50 (4%) 

586 567 582 

Adjusted rate 9.1 % 20.6% 31.1% 7.4% 

Terminal rate 2/29 (7%) 2/20 (10%) 4/20 (20%) 1/20 (5%) 

First incidence (days) 655 

Life table test P=0.491NP=O.O49 P=O.171 P=0.604N 

Logistic regression test P=O.O85 P=O.245 P=0.373N P=0.501N 

Cochran-Armitage test P=0.360N 

Fisher exact test P=O.O87 P=O.243 P=0.500N 

Skin:SquamousCellPapilloma,Keratoacanthoma,Trichoepithelioma,orSquamousCellCarcinoma 

586 567 582 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Skin (SubcutaneousTissue):Fibroma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Skin (SubcutaneousTissue):FibromaorSarcoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

1/50 (2%) 3/50 (6%) 4/50 (8%) 3/50 (6%) 

3.4% 14.3% 17.2% 12.4% 

732 (T) 

1/29 (3%) 

729 

2/20 (10%) 

706 

3/21 (14%) 

594 

2/20 (10%) 

P=O.182 P=O.189 P=O.109 P=O.207 

P=O.248 P=O.200 P=O.146 P=O.293 

P=O.282 

P=O.309 P=O.181 P=O.309 

5/50 (10%) 2/50 (4%) 3/50 (6%) 1/50 (2 46) 

15.8% 8,4% 11.8% 3.0% 

4/29 (14%) 1/20 (5%) 2/21 (10%) 0120 (0%) 

567 693 635 670 

P=0.153N P=0.336N P=0.487N P=0.173N 

P=0.099N P-0.220N P=0.346N P=0.108N 

P=0.095N 

P=0.218N P=0.357N P=0.102N 

5/50 (10%) 3/50 (6%) 3/50 (6%) 1/50 (2%) 

15.8% 10.3% 11.8% 3.0% 

4/29 (14%) 1 /20 (596) 2/21 (10%) 0/20 (0%) 

567 426 635 670 

P=0.129N P=O.489N P=0.487N P=0.173N 

P=0.079N P=O.356N P=O.346N P=0.108N 

P=0.079N 

P=0.357N P=0.357N P=0.102N 
96 Codeine, NTP TR 455 

Testes:Adenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

ThyroidGland (Ccell): Adenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

ThyroidGland (Ccell): Carcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

ThyroidGland (Ccell): Adenoma or Carcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

AllOrgans:MononuclearCellLeukemia 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-armitage test 

Fisher exact test 

0 PPm 

47/50 (94%) 

97.9% 

28/29 (9796) 

334 

P=O.O83 

P=O.495 

P=O.343 

3/50 (6%) 

9.7% 

2/29 (7%) 

68 1 

P=0.472N 

P=0.404N 

P=0.371N 

2/50 (4%) 

6.9% 

2/29 (7%) 

732 (T) 

P=O.260 

P=O.319 

P=O.357 

5/50 (10%) 

16.4% 

4/29 (14%) 

68 1 

P=O.416 

P=O.513 

P=O.563 

32/50 (64%) 

72.7% 

17/29 (59%) 

495 

P=O.o44 

P=O.112 

P=O.O65 

400 ppm 

47/50 (94%) 

100.0% 

20120 (100%) 

426 

P=O.101 

P=0.500N 

P=0.661N 

3/50 (6%) 

12.0% 

1/20 (5%) 

684 

P=O.543 

P=O.645 

P=0.661N 

2/50 (4%) 

10.0% 

2/20 (10%) 

732 (T) 

P=O.555 

P=O.555 

P=0.691N 

5/50 (10%) 

21.3% 

3/20 (15%) 

684 

P=O.437 

P=O.572 

P=0.630N 

27/50 (54%) 

64.296 

7/20 (35%) 

404 

P=0.553N 

P=0.083N 

P=0.208N 

800 ppm 

49/50 (98%) 

100.0% 

21/21 (100%) 

519 

P=O.O97 

P=O.650 

P=O.309 

2/50 (4%) 

9.5% 

2/21 (10%) 

732 (T) 

P=0.619N 

P=O.526N 

P=0.500N 

4/50 (8%) 

17.3% 

3/21 (14%) 

707 

P=O.216 

P=O.282 

P=O.339 

5/50 (10%) 

21.9% 

4/21 (19%) 

707 

P=O.455 

P=O.585 

P=0.630N 

36/50 (72%) 

78.8% 

12/21 (57%) 

519 

P=O.141 

P=0.507N 

P=O.260 

1,600 ppm 

48/50 (96%) 

100.0% 

20120 (100%) 

484 

P=O.O71 

P=O.676 

P=O.500 

2/50 (4%) 

8.5% 

1/20 (5%) 

705 

P=0.614N 

P=0.538N 

P=0.500N 

3/50 (6%) 

9.9% 

0120 (0%) 

659 

P=O.411 

P=O.479 

P=O.500 

5/50 (10%) 

17.6% 

1/20 (5 %) 

659 

P=O.483 

P=O.593 

P=O.63ON 

37/50 (74%) 

81.5% 

12/20 (60%) 

355 

P=O.O71 

P=O.299 

P=O.194 
Lesions in Male Rats 97 

TABLEA3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study of Codeine (continued) 

~ ~~~~ 

0 PPm 400 ppm 800 ppm 1,600 ppm 

All Organs:MalignantMesothelioma 

Overall rate 1/50 (2%) 2/50 (4%) 2/50 (4%) 3/50 (6%) 

Adjusted rate 3.4% 5.7% 8.3% 7.7% 

Terminal rate 1/29 (3%) 0120 (0%) 0121 (0%) 0120 (0%) 

First incidence (days) 732 (T) 574 722 484 

Life table test P=O.208 P=O.459 P=O.415 P=O.274 

Logistic regression test P=O.232 P=O.500 P=O.465 P=O.296 

Cochran-Armitage test P=O.232 

Fisher exact test P=O.500 P=O.500 P=O.309 

All Organs:BenignNeoplasms 

Overall rate 50150 (100%) 49/50 (98I) 50150 (100%) 48/50 (96%) 

Adjusted rate 100.0% 100.0% 100.0% 100.0% 

Terminal rate 29/29 (100%) 20/20 (100%) 21/21 (100%) 20/20 (100%) 

First incidence (days) 203 336 519 484 

Life table test P=O.175 P=O.134 P=O.160 P=O.148 

Logistic regression test P=0.032N P=0.357N P=0.068N 

Cochran-Armitage test P=0.140N 

Fisher exact test P=0.500N P= 1.000N P=0.247N 

All Organs:MalignantNeoplasms 

Overall rate 36/50 (72%) 33/50 (66%) 39/50 (78%) 41/50 (82%) 

Adjusted rate 79.9% 74.5% 82.3% 83.6% 

Terminal rate 20/29 (69%) 10/20 (50%) 13/21 (62%) 12/20 (60%) 

First incidence (days) 203 336 519 355 

Life table test P=O.O53 P=O.386 P=O.156 P=O.O67 

Logistic regression test P=O.O77 P=0.337N P=O.324 P=O.171 

Cochran-Armitage test P=O.O73 

Fisher exact test P=0.333N P=O.322 P=O.171 

All Organs:Benign or MalignantNeoplasms 

Overall rate 50150 (100%) 50150 (100%) 50150 (100%) 50/50 (100%) 

Adjusted rate 100.01 100.0% 100.0% 100.0% 

Terminal rate 29/29 (100%) 20/20 (100%) 21/21 (100%) 20/20 (looX) 

First incidence (days) 203 336 519 355 

Life table test P=O.122 P=O.108 P=O.160 P=O.O97 

Logistic regression test -

Cochran-Annitage test 

Fisher exact test P= 1.000N P= 1.WN P= 1.000N 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animalslnumber of animals examined. Denominator is number of animals examined microscopically for adrenal gland, liver, 

pancreatic islets, pituitary gland, preputial gland, testes, and thyroid gland; for other tissues, denominator is number of animals necropsied. 

Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to 

pairwise comparisons between the controls and that exposed group. The life table test regards neoplasms inanimals dying prior to terminal kill as being 

(directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests 

compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposed group is indicated by N. 

Not applicable;no neoplasms in animal group 

Value of statistic cannot be computed. 
98 Codeine, NTP TR 455 

TABLEA4 

Historical Incidence of Benign Pheochromocytomas of the Adrenal Medulla in Untreated Male F344/NRatsa 

Incidence in Controls 

~ ~ ~~ ~~~ 

Historical Incidence at Microbiological Associates, Inc. 

&Amphetamine Sulfate 

C.I. Direct Blue 218 

23/49 

15/49 

Overall Historical Incidence 

Total 379/1,182 (32.1%) 

Standard deviation 11.7% 

Range 10%-63% 

a Data as of 17 June 1994 
Lesions in Male Rats 99 

TABLEA5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-YearFeed Study of Codeinea 

0 PPm 400 PPm 800 ppm 1,600 ppm 

DispositionSummary 

Animals initially in study 

15-Month interim evalualion 

Early deaths 

Moribund 

Natural deaths 

Survivors 

Terminal sacrifice 

Animals examined microscopically 

15-Month Interim Evaluation 

AlimentarySystem 

Intestine small, ileum 

cyst 

Liver 

Basophilic focus 

Clear cell focus 

Eosinophilic focus 

Hepatodiaphragmatic nodule 

Inflammation, chronic. focal 

Mixed cell focus 

Necrosis 

Vacuolization cytoplasmic, focal 

Mesentery 

Fat, necrosis 

Pancreas 

Accessory spleen 

Acinar cell, atrophy 

Cardiovascular System 

Heart 

cardiomyopathy 

Endocrine System 

Adrenal cortex 

Hyperplasia, focal 

Hypertrophy

Vacuolization cytoplasmic, focal 

Adrenal medulla 

Hyperplasia 

Pituitary gland 

Pars distalis, hyperplasia, focal 

Thyroid gland 

C-cell, hyperplasia 

Follicle, cyst 

60 60 

10 10 

14 23 

7 7 

29 20 

60 60 

(10) 

2(20%) 

1 (10%) 

1(10%) 1 (10%) 

1 (10%) 

2 (20%) 

1 (10%) 

1 (10%) 

(3) 

3 (100%) 

(10) 

1(10%) 

5 (50%) 4 (40%) 

1(10%) 

3 (30%) 

(10) 

60 

10 

24 

5 

21 

60 

(10) 

5 (50%) 

1 (10%) 

1 (10%) 

(7) 

6 (86%) 

(10) 

1 (10%) 

5 (50%) 

60 

10 

23 

7 

20 

60 

2 (20%) 

5 (50%) 

a Number of animals examined microscopically at the site and the number ofanimals with lesion 
100 Codeine, NTP TR 455 

TABLEA5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

15-Month InterimEvaluation (continued) 

Genital System 

Epididymis 

Edema, focal 

(10) 

Penis 

Hemorrhage 

Ectasia 

Preputial gland (9) 

Hyperplasia 

Inflammation 

Atrophy 

Prostate 

1 (11%) 

(10) 

Miltration cellular, focal, lymphocyte 

Inflammation 

Depletion cellular 

Seminal vesicle 

Interstitial cell, hyperplasia, focal 

Congestion, focal 

Testes 

2 (20%) 

(10) 

(10) 

3 (30%) 

HematopoieticSystem 

Bone marrow 

Lymph node, mandibular 

Hyperplasia 

Thymus 

Hyperplasia, plasma cell 

Atrophy

Hemorrhage 

Respiratory System 

Lung 

Hemorrhage 

(10) 

1 (10%) 

(9) 

1 (11%) 

(10) 

2 (20%) 

Infiltration cellular, focal, histiocyte 1 (10%) 

Inflammation 1 (10%) 

Edema, focal 

Nose (10) (10) 

1 (10%) 

(10) 

Foreign body 2 (20%) 1 (10%) 

Fungus 2 (20%) 1 (10%) 

Hemorrhage 1 (10%) 1 (10%) 

Infiltration cellular, focal, lymphocyte 2 (20%) 

Olfactory epithelium, atrophy, focal 

Goblet cell, hyperplasia, focal 

Inflammation 

2 (20%) 

4 (40%) 

2 (20%) 

1 (10%) 

1 (10%) 

3 (30%) 

1 (10%) 

2 (20%) 

2 (20%) 

Olfactory epithelium, cytoplasmic alteration 8(80%) 7 (70%) 8 (80%) 10 (100%) 

Olfactory epithelium, vacuolization 

Olfactory epithelium, hyperplasia, focal 1 (10%) 

cytoplasmic, focal 1 (10%) 

Respiratory epithelium, cytoplasmic 

alteration, focal 1 (10%) 

Special Senses System 

Eye 

Lens, cataract 

(1) 

1(100%) 
Lesions in Male Rats 101 

TABLE A5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

I5-Month Interim Evaluation (continued) 

Urinary System 

Kidney 

Nephropathy, chronic 

(10) 

10 (100%) 

(10)

9 (90%) 

(10) 

8 (80%) 

(10) 

8 (80%) 

~~~~~~ ~ 

Systems Examined With No Lesions Observed 

General Body System 

Integumentary System 

Musculoskeletal System 

Nervous System 

2- Study 

Alimentary System 

Intestine large, colon 

Amyloid deposition 

(50) (50) 

1 (2%) 

Inflammation 1 (2%) 

Pigmentation 

Ulcer 

Inflammation 

Intestine large, rectum (49) (48) 

1 (2%) 

Pigmentation 1 (2%) 

Ulcer 1 (2%) 

Diverticulum 

Intestine small, jejunum (47) 

Inflammation, chronic 

Abscess 

Liver (50) 

1 (2%) 

Angiectasis 3 (6%) 

Atrophy 1 (2%) 

Basophilic focus 17 (34%) 17 (34%) 21 (42%) 16 (32%) 

Clear cell focus 2 (4%) 2 (4%) 1 (2%) 

Congestion 2(4%) 1 (2%) 

Degeneration 3(6%) 1 (2%) 2 (4%) 

Degeneration, cystic 14 (28%) 5 (10%) 1 (2%) 

Developmental malformation 2 (4%) 1 (2%) 

Eosinophilic focus 7 (14%) 1(2%) 5 (10%) 4 (8%) 

Fatty change 5 (10%) 1 (2%) 1 (2%) 

Fibrosis 3 (6%) 1(2%) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 

Hemorrhage 1(2%) 1 (2%) 

Hyperplasia, focal 

Hepatodiaphragmatic nodule 2 (4%) 3 (6%) 2 (4%) 

1 (2%) 

1 (2%) 

Infiltration cellular, lymphocyte 1 (2%) 1 (2%) 1 (2%) 

Inflammation, chronic, focal 1 (2%) 1 (2%) 

Leukocytosis 1 (2%) 

Mixed cell focus 2 (4%) 1 (2%) 

Thrombosis 

Necrosis 

1 (2%) 

1 (2%) 

1 (2%) 

3(6%) 

4(8%) 

1 (2%) 

1 (2%) 

1 (2%) 

Bile duct, cyst 1 (2%) 

Bile duct, fibrosis 1 (2%) 

Bile duct, hyperplasia 1 (2%) 5 (10%) 3(6%) 1(2%) 
102 Codeine, NTP TR 455 

TABLEA5 

SmmaV of the Incidence.Of Nonneoplastic Lesions in Male Rats.in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 

2-Year Study (continued) 

Alimentary System (continued) 

Mesentery 

Fat, necrosis 

Pancreas 

Amyloid deposition 

Accessory spleen 

Ectopic liver 

Edema 

Necrosis 

Infdttation cellular, lymphocyte 

Artery, inflammation 

Acinar cell, atrophy 

Amyloid deposition 

Salivary glands 

1 (2%) 

29 (58%) 

1 (2%) 

(50) 

Inflammation, chronic 

Infiittation cellular, lymphocyte

Atrophy 

1 (2%) 

1 (2%) 

Hyperkeratosis, focal 

Edema 

Stomach, forestomach (50) 

1 (2%) 

Hyperplasia 

Ulcer 

Stomach, glandular 

Amyloid deposition 

Hyperkeratosis, focal 

Ulcer 

Necrosis 

2 (4%) 

1 (2%) 

Cardiovascular System 

Heart 

Cardiomyopathy 

(50)

24 (48%) 

Atrium, thrombosis 

Atrium, dilatation 

Mineraliition, focal 

2 (4%) 

1 (2%) 

1 (2%) 

5 (10%) 

1 (2%) 

Endocrine System 

Adrenal cortex 

Angiectasis 

Cytologic alterations 

Hemorrhage 3 (6%) 

Hyperplasia, focal 2 (4%) 

Hypertrophy

Necrosis 

1 (2%) 

Vacuolization cytoplasmic, focal 

Adrenal medulla 

Islets. pancreatic 

Hyperplasia 

Parathyroid gland 

Hyperplasia 

Hyperplasia 

1,600 ppm 

22 (44%) 

2 (4%) 

4 (8%) 

1 (2%) 

4 (8%) 

(50) 

20 (40%) 

2 (4%) 

5 (10%) 

1 (2%) 

1 (2%) 

(46) 

1 (2%) 
Lesions in Male Rats 103 

TABLEA5 

Summary of the Incidence of Nonneoplastic Lesions inMale Rats in the 2-Year Feed Study of Codeine (continued) 

&Year Study (continued) 

Endocrine System (continued) 

Pituitary gland 

Congestion 

Pars distalis, angiectasis 

Pars distalis, cyst 

Pars distalis, degeneration 

Pars distalis, hyperplasia, focal 

Pars intermedia, angiectasis 

Pars nervosa, cyst 

Pars nervosa, developmental malformation 

Thyroid gland 

C-cell, hyperplasia 

Follicle, atrophy 

Follicle, dilatation 

General Body System 

Tissue NOS 

Mediastinum, iditration cellular, 

lymphocyte 

Genital System 

Epididymis 

Granuloma sperm 

Hypoplasia 

Hypospermia 

Penis 

Angiectasis 

Inflammation 

Preputial gland 

Atrophy 

Depletion cellular 

Ectasia 

Hyperplasia 

Inflammation 

Prostate 

Atrophy

Cytoplasmic alteration 

Dilatation 

Fibrosis, focal 

Hemorrhage 

Hyperplasia, focal 

Infiltration cellular, focal, lymphocyte 

Inflammation 

Seminal vesicle 

Depletion cellular 

Dilatation 

Ectasia 

Infitration cellular, lymphocyte 

Testes 

Atrophy 

Bilateral, atrophy 

Interstitial cell, hyperplasia, focal 

0 PPm 400 ppm 800 ppm 1,600 ppm 

(48) (47) 

3 (6%) 

2 (4%) 

14 (28%) 

(50) 

4 (8%) 

1 (2%) 1 (2%) 

(1) 

1 (100%) 

(49) (49) 

33 (66%) 34 (69%) 

9 (18%) 4 (8%) 

1 (2%) 11 (22%) 

3 (6%) 9 (18%) 

(49) (50) 

5 (10%) 

1 (2%) 

4 (8%) 2 (4%) 

5 (10%) 2 (4%) 

(50) 

35 (70%) 

(50) 

42 (84%) 

(50) 

1 (2%) 

1 (2%) 
104 Codeine, NTP TR 455 

2-Yeur Stud' (continued) 

HematopoieticSystem 

Bone marrow 

Atrophy

Congestion 

Hemorrhage 

Hyperplasia 

Myelofibrosis 

Necrosis 

Erythroid cell, hyperplasia 

Myeloid cell, atrophy 

Myeloid cell, hyperplasia 

Lymph node 

Lumbar, pigmentation 

Mediastinal, hemorrhage 

Mediastinal, pigmentation 

Pancreatic, lymphocyte, hyperplasia 

Lymph node, mandibular 

Amyloid deposition 

cyst 

Hemorrhage 

Hyperplasia. lymphoid 

Hyperplasia, plasma cell 

Lymphocyte, hyperplasia 

Lymph node, mesenteric 

Amyloid deposition 

cyst 

Cyst, multiple 

Erythrophagocytosis 

Hemorrhage 

Hyperplasia, plasma cell 

Infitration cellular, histiocyte 

Pigmentation 

Lymphocyte, hyperplasia 

Spleen 

Congestion 

Developmental malformation 

Fibrosis 

Hematopoietic cell proliferation 

Witration cellular, histiocyte 

Necrosis 

Pigmentation 

Lymphocyte, hyperplasia 

Thymus 

Atrophy

Depletion cellular 

Hemorrhage 

Witration cellular, lymphocyte, 

0 PPm 400 ppm 800 ppm 1.,600 ppm 

8 (16%) 

1 (2%) 

1 (2%) 

(41) 

23 (56%) 

1 (2%) 

1 (2%) 
Lesions inMale Rats 105 

TABLEA5 

Summary of the Incidence of Nonneoplastic Lesions in MaleRats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400PPm PPm 1,600 ppm 

2-Year Shrdy (continued) 

IntegumentarySystem 

Mammary gland (47) (49)

Fibrosis 

Galactocele 

Hemorrhage 

Hyperplasia 3 (6%)

Infdtration cellular, lymphocyte 

Infiltration cellular. histiocyte 

Lactation 

Pigmenfation 

Skin 

Acanthosis 

Hemorrhage 

Inflammation 

Ulcer 

Hair follicle, atrophy 

Subcutaneous tissue. inflammation 

Tail, acanthosis, focal 

Tail, inflammation 

Tail. necrosis 

Musculoskeletal System 

Bone (50)

Hyperostosis 1 (2%) 

Rib, developmental malfonnation. focal 1 (2%) 

Nervous System 

Brain 

Compression 

Hemorrhage 

Necrosis 

Cerebellum, necrosis, focal 

Respiratory System 

Lung

Fibrosis, focal 

Hemorrhage 

Mitration cellular, focal, histiocyte 

InfYtration cellular. lymphocyte 

Inflammation 

Leukocytosis 

Alveolar epithelium, hyperplasia, focal 

(50) (50) 
106 Codeine, NTP TR 455 

TABLEA5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the2-YearFeed Study of Codeine (continued) 

2-Year Study (continued) 

Respiratory System (continued) 

Nose 

Angiectasis 

Edema, focal 

Foreign body 

Fungus 

Inflammation 

Inflammation, acute 

Thrombosis 

Cartilage, dysplasia 

Glands, dilatation, focal 

Goblet cell, hyperplasia, focal 

Nasolacrimal duct, Mammation 

Nasopharyngeal duct, metaplasia, squamous 

Olfactory epithelium, atrophy, focal 

Olfactory epithelium, cytoplasmic alteration 

Olfactory epithelium, degeneration 

Olfactory epithelium, necrosis, focal 

Respiratory epithelium, cytoplasmic 

alteration, focal 

Respiratory epithelium, hyperplasia 

Trachea 

Infiitration cellular, focal, lymphocyte 

Inflammation 

Special Senses System 

Eye

Phthisis bulbi 

Lens, cataract 

Retina, degeneration 

Urinary System 

Kidney 

Congestion 

Infarct 

Infiitration cellular, lymphocyte 

Nephropathy, chronic 

Pigmentation 

Bilateral, hydronephrosis 

Glomerulus, degeneration 

Transitional epithelium, hyperplasia, focal 

Urethra 

Inflammation 

Transitional epithelium, hyperplasia 

Urinary bladder 

Amyloid deposition 

Dilatation 

Hemorrhage 

Witration cellular, lymphocyte 

Inflammation 

Transitional epithelium, hyperplasia 

0 PPm 400 ppm 

(50) 

1 (2%) 

4 (8%) 

11 (22%) 

1 (2%) 

16 (32%) 

(50) 

1 (2%) 

(50) 

1 (2%) 

40 (80%) 

1 (2%) 

1 (2%) 

(50) 

1 (2%) 

800 ppm 1,600 ppm 

(50) 

1 (2%) 

1 (2%) 

1 (2%) 

5 (10%) 

1 (2%) 

1 (2%) 

2(4%) 

47 (94%) 

2 (4%) 

1 (2%) 

(50) 

(2) 

1 (50%) 

1 (2%) 

1 (2%) 

1 (2%) 
107 

APPENDIX B 

SUMUMARY OF LESIONS IN FEMALE RATS 

IN THE 2-YEAR FEED STUDY 

OF CODEINE 

TABLEB1 Summary of the Incidence of Neoplasms in Female Rats 

in the 2-Year Feed Study of Codeine ................................. 109 

TABLEB2 Individual Animal Tumor Pathology of Female Rats 

in the 2-Year Feed Study of Codeine ................................. 114 

TABLE B3 Statistical Analysis of Primary Neoplasms in Female Rats 

in the 2-Year Feed Study of Codeine .................................. 130 

TABLEB4a Historical Incidence of Benign Pheochromocytomas of the Adrenal Medulla 

in Untreated Female F344/N Rats ................................... 135 

TABLEB4b Historical Incidence of Clitoral Gland Neoplasms 

in Untreated Female F344/N Rats ................................... 135 

TABLEB4c Historical Incidence of Mammary Gland Neoplasms 

in Untreated Female F344/N Rats ................................... 136 

TABLE B4d Historical Incidence of Pituitary Gland (Pars Distalis) Neoplasms 

in Untreated Female F344/N Rats ................................... 136 

TABLEB5 Summary of the Incidence of Nonneoplastic Lesions in Female Rats 

in the 2-Year Feed Study of Codeine ................................. 137 
Lesions in Female Rats 109 

0 PPm 400 ppm 800 ppm 1,600 ppm 

Early deaths 

IS-Monfh inferim evdwn 

Animals initially in study 

Disposition Summary 

Terminal sacrifice 

Died last week of study 

Natural deaths 

Moribund 

Survivors 

10 

60 

7 

15 

28 

10 

60 

3 

9 

37 

1 

9 

60 

5 

17 

28 

1 

9 

60 

5 

14 

32 

Animals examined microscopically 60 60 60 60 

Pituitary gland 

Endocrine System 

15-Month Interim Evaluation 

Pars distalis, adenoma 

(10) 

1 (10%) 

(10) 

1 (10%) 

(9) 

1 (11%) 

(9) 

Polyp stromal 

uterus 

Genital System 

Tail, keratoacanthoma 

Skin 

Integumentary System 

Multiple organsb 

Systemic Lesions 

Leukemia mononuclear 

Urinary System 

Special Senses System 

Respiratory System 

Nervous System 

Musculoskeletal System 

HematopoieticSystem 

General Body System 

Cardiovascular System 

Alimentary System 

Systems Examined With No Neoplasms Observed 
110 Codeine, NTP TR 455 

TAB^ B1 

Summary of the Incidence 'of Neoplasms in Female Rats in'the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

2-Year Study 

Alimentary Sy$tem 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Sarcoma, metastatic, uncertain primary site 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Adenocarcinoma 

Intestine small, ileum 

Liver 

Histiocytic sarcoma 

Mesentery 

Oral mucosa 

Buccal, squamous cell carcinoma 

Pancreas 

Salivary glands 

Carcinoma, metastatic, Zymbal's gland 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Pheochromocytoma malignant 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Carcinoma 

Parathyroid gland 

Adenoma 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Thyroid gland 

Bilateral, follicular cell, carcinoma 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell, adenoma 

Follicular cell. carcinoma 
Lesions in Female Rats 111 

TABLEB1 

Summary of the Incidence of Neoplasms in FemaleRats in the 2-YearFeed Study of Codeine (continued) 

2-Year Study (continued) 

General Body System 

None 

Genital System 

Clitoral gland 

Adenoma 

carcinoma 

Bilateral, adenoma 

Ovary 

Granulosa-theca tumor malignant 

Luteoma 

Oviduct 

Utems 

Polyp stromal 

Sarcoma 

Sarcoma stromal 

Vagina 

Squamous cell papilloma 

Hematopoietic System 

Bone marrow 

Histiocytic sarcoma 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Carcinoma, metastatic, Zymbal’s gland 

Lymph node, mesenteric 

Spleen 

Histiocytic sarcoma 

Thymus 

Thymoma benign 

IntegumentarySystem 

Mammary gland 

Adenoacanthoma 

Adenocarcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Keratoacanthoma 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibroma, multiple 

Subcutaneous tissue, fibrosarcoma 

Subcutaneous tissue, sarcoma 

Tail, schwannoma malignant 

(50) 

(12) 

(48) 

(50) 

(50) 

(44) 

1 (2%) 

1 (2%) 

1 (2%) 
112 Codeine, NTP TR 455 

TABLEB1 

Summary of the Incidence of Neoplasms in Female Rats in the2-YearFeed Study of Codeine (continued) 

2-Year Sdy (continued) 

Musculoskeletal System 

Skeletal muscle 

Sarcoma 

Nervous System 

Brain 

Astrocytoma benign 

Carcinoma, metastatic, pituitary gland

Spinal cord 

Respiratory System 

Lung 

Alveolarhronchiolar adenoma 

Alveolarhronchiolar carcinoma 

Chordoma, metastatic, uncertain primary site 

Sarcoma, metastatic, uncertain primary site 

Nose 

Carcinoma, metastatic, Zymbal’s gland 

Sarcoma 

Nasolacrimal duct, squamous cell carcinoma 

Trachea 

Special Senses System 

Ear 

Pinna, schwannoma malignant 

Zymbal’s gland 

Carcinoma 

Urinary System 

Kidney 

Lipoma 

Nephroblastoma 

Renal tubule, carcinoma 

Urinary bladder 

SystemicLesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear 

Lymphoma malignant 

0 PPm 400 ppm 800 ppm 1,600 ppm 

(1) 

1 (100%) 

(50) (50) (51) 

1 (2%) 

1 (2%) 1 (2%) 

(2) 

(50) 

1 (2%) 

(50) 

(50) 

(50) (50) (51) (51) 

1 (2%) 

14 (28%) 16 (32%) 18 (35%) 12 (24%) 

1 (2%) 
Lesions in Female Rats 113 

TABLEB1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study of Codeine (continued) 

0 PPm 400 PPm 800 ppm 

Neoplasm Summary 

Total animals with primary neoplasms' 

15-Month interim evaluation 2 2 3 

2-Year study 46 44 47 

Total primary neoplasms 

15-Month interim evaluation 2 2 3 

2-Year study 91 84 112 

Total animals with benign neoplasms 

15-Month interim evaluation 1 2 3 

2-Year study 38 40 41 

Total benign neoplasms 

15-Month interim evaluation 1 2 3 

2-Yearstudy . 63 63 77 

Total animals with malignant neoplasms 

15-Month interim evaluation 1 

2-Year study 27 19 28 

Total malignant neoplasms 

15-Month interim evaluation 1 

2-Year study 28 21 35 

Total animals with metastatic neoplasms 

2-Year study 1 3 1 

Total metastatic neoplasms 

2-Year study 1 5 2 

Total animals with malignant neoplasms 

of uncertainprimary site 

2-Year study 1 1 

a Number of animals examined microscopically at the site and the number of animals with neoplasm 

Number of animals with any tissue examined microscopically

' primary neoplasms:allneoplasmsexceptmetastaticneoplasms 

1,600 ppm 

37 

51 

27 

32 

17 

19 
Codeine, NTP TR 455 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 0 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Adenocarcinoma 

Intestine small, ileum 

Liver 

Mesentery 

Oral mucosa 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma malignant 

Pheochromocytoma benign 

Islets, pancreatic 

Carcinoma 

Parathyroid gland 

Adenoma 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland 

Adenoma 

Carcinoma 

+: Tissue examined microscopically 

A:Autolysisprecludesexamination 

3444445555555556666677777 

1377993447888893677812333 

3255052785116711337925333 

2222232222222222222222222 

6647809898459675869854677 

6911909784656230514628747 

......................... 

......................... 

......................... 

......................... 

.......................... 

++++++++++++++++++++++++M 

X .......................... 

......................... + 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

X ......................... 

++M++++++++++++++++++++++ 

X 

+++++++++++++++++M++MM+++ 

X x xx 

......................... X X 

X 

X 

++++++++M++++++++M++++f++ 

X X 

X 

M:Missing tissue X: Lesionpresent 

I: Insufficienttissue Blank:Notexamined 
Lesions in Female Rats 115 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Adenocarcinoma 

Intestine small, ileum 

Liver 

Mesentery 

Oral mucosa 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma malignant 

Pheochromocytoma benign 

Islets, pancreatic 

Carcinoma 

Parathyroid gland 

Adenoma 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland 

Adenoma 

Carcinoma 

7777777777777777777777777 

3333333333333333333333333 

3344444444444555555666666 

2222222222222222222222222 Total 

8955677788999445589446679 Tissues/ 

3378425812257471381395890 Tumors 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 49 

1 

......................... 50 

......................... 50 

+ + ++ + 6 

+ + + 3 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

X 1 

X 2 

......................... 50 

X 1 

++++MM+++++++M+++++++++M+ 45 

1 

++++++++++++M++++++++++++ 46 

X xxx X xxxxxx X 16 

A 1 

......................... 50 

X X 4 

X 2 

1 

+++++M+++++++++++++++++++ 47 

X X X 5 

1 
116 Codeine, NTP TR 455 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Genital System (continued) 

Ovary 

Granulosa-theca tumor malignant 

Luteoma 

uterus 

Polyp stromal 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

Thymoma benign 

IntegumentarySystem 

Mammary gland 

Adenocarcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibrosarcoma 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Astrocytoma benign 

Carcinoma, metastatic, pituitary gland 

RespiratorySystem 

Lung 

Alveolarhronchiolar carcinoma 

Nose 

Trachea 

Special Senses System 

EYe 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Lymphoma malignant 

3444445555555556666677777 

1377993447888893677812333 

3255052785116711337925333 

2222232222222222222222222 

6647809898459675869854677 

6911909784656230514628747 

......................... 

.......................... 

......................... 

+ + + + 

++++++++++++++++++++M++++ 

.......................... 

......................... 

M+++++++++++++M+M+++++++M 

x # 

......................... X X 

xxx x xx x x xx x 

X X 

......................... 

X X 

X 

......................... 

......................... X 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... x x X xx x xxx xx 
Lesions in Female Rats 117 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

7117171771171777111777177 

Number of Days on Study 3333333333333333333333333 

3344444444444555555666666 

2222222222222222222222222 Total 

Carcass ID Number 8955671788999445589446619 Tissues/ 

3318425812257471381395890 Tumors 

Genital System (continued) 

Ovary ......................... 50 

Granulosa-theca tumor malignant X 1 

Luteoma X 1 

uterus ......................... 50 

Polyp stromal X 1 

HematopoieticSystem 

Bone marrow ......................... 50 

Lymph node + + + + + +++ 12 

Lymph node, mandibular +++++++++++++++M+++++++++ 48 

Lymph node, mesenteric ......................... 50 

Spleen ......................... 50 

Thymus ++++++M+++++++++++++++M++ 44 

Thymoma benign 1 

Integumentary System 

Mammary gland ......................... 50 

Adenocarcinoma X 3 

Fibroadenoma x xx xx X x x x xx 22 

Fibroadenoma, multiple X X X 5 

Skin ......................... 50 

Squamous cell papilloma X 1 

Subcutaneous tissue, fibroma 2 

Subcutaneous tissue, fibrosarcoma 1 

MusculoskeletalSystem 

Bone ......................... 50 

Nervous System 

Brain ......................... 50 

Astrocytoma benign 1 

Carcinoma, metastatic, pituitary gland X 1 

~ ~~ ~~ ~ ~~~ ~~~ ~~ ~~ ~~ ~ ~ ~~ 

Respiratory System 

Lung ......................... 50 

Alveolar/bronchiolar carcinoma X 1 

Nose ......................... 50 

Trachea ......................... 50 

Special Senses System 

EY e + 1 

Urinary System 

Kidney ......................... 50 

Urinary bladder ......................... 50 

Systemic Lesions 

Multiple organs ......................... 50 

Leukemia mononuclear xx X 14 

Lymphoma malignant X 1 
118 Codeine, NTP TR 455 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 400 ppm 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Pancreas 

Salivary glands 

Carcinoma, metastatic, Zymbal’s gland 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Healt 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Thyroid gland 

C-cell, adenoma 

C-cell, carcinoma 

General Body System 

Tissue NOS 

Genital System 

Clitoral gland 

Adenoma 

Bilateral, adenoma 

Ovary 

Uterus 

Polyp stromal 

4455666677777777777777777 

0744088800133333333333333 

0906619957213333334444444 

3333333333333333333333333 

1402431540030235550011222 

4410990578302932794937136 

......................... 

......................... 

......................... 

+++++A+++++A+++++++++++++ 

......................... 

+++++++++++A+++++++++++++ 

+++++AA++++++++++++++++++ 

......................... ++ + ......................... 

......................... X 

......................... 

......................... 

......................... 

......................... 

......................... 

......................... X X ......................... X 

+M+M++++M+++++++M+++MM+++ 

......................... X X xx xx x x 

X ......................... x x X 

X 

+ 

+++++++++++M+++M+++++++++ 

X X X 

X 

......................... 

......................... X xx x X 
Lesions in Female Rats 119 

Tmq B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: -400ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small. duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Pancreas 

Salivary glands 

Carcinoma, metastatic, Zymbal's gland 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Pheochromocytomabenign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis. adenoma 

Pars distalis, carcinoma 

Thyroid gland 

C-cell, adenoma 

C-cell. carcinoma 

General Body System 

Tissue NOS 

Genital System 

Clitoral gland 

Adenoma 

Bilateral, adenoma 

Ovary 

UkNS 

Polyp stromal 

7777777777777777777777777 

3333333333333333333333333 

4444444455555555555666666 

3333333333333333333333333 Total 

2333444400112344555133455 Tissues/ 

7678125667588508038224346 Tumors 

......................... 50 

......................... 50 

......................... 50 

......................... 48 

......................... 50 

......................... 49 

......................... 48 

......................... 50 

+ + 5 

......................... 50 

++++++++M++++++++++++++++ 49 

1 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

X 1 

......................... 50 

X x x 5 

......................... 50 

1 

++MMM++M+++M+++++M+++M+++ 37 

......................... 50 

X X xxx x x x x 17 

1 

......................... 50 

3 

1 

1 

+++++++++++++++++++++++M+ 47 

X X 5 

1 

......................... 50 

......................... 50 

X 6 
120 Codeine, NTP TR 455 

TABLEB2 

Individual Animal Tumor Pathologyof Female Rats in the 2-YearFeed Study of Codeine: 400 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Carcinoma, metastatic, Zymbal’s gland 

Lymph node, mesenteric 

Spleen 

Thymus 

Integumentary System 

Mammary gland 

Adenocarcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Keratoacanthoma 

Subcutaneous tissue, fibroma, multiple 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Carcinoma, metastatic, pituitary gland 

Spinal cord 

Respiratory System 

Lung 

Chordoma, metastatic, uncertain primary site 

Nose 

Carcinoma, metastatic, Zymbal’s gland 

Trachea 

Special Senses System 

Zymbal’s gland 

Carcinoma 

Urinary System 

Kidney 

Lipoma 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

4455666677777777777777777 

0744088800133333333333333 

0906619957213333334444444 

3333333333333333333333333 

1402431540030235550011222 

4410990578302932794937136 

.......................... ++ +++ + + + ......................... X 

++++++++ ++++++++++++++++ ......................... 

+++++++++++++++M+++++++++ 

......................... X 

X x x x x x x x xx x 

......................... 

......................... 

......................... X 

+ 

......................... X 

......................... X 

......................... 

+ 

X 

......................... 

......................... 

......................... x x x xx x x x X X X
Lesions in Female Rats 121 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 400 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Carcinoma, metastatic, Zymbal’s gland 

Lymph node, mesenteric 

Spleen 

Thymus 

~ ~~~ 

IntegumentarySystem 

Mammary gland 

Adenocarcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Keratoacanthoma 

Subcutaneous tissue, fibroma, multiple 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Carcinoma, metastatic, pituitary gland 

Spinal cord 

Respiratory System 

Lung 

Chordoma, metastatic, uncertain primary site. 

Nose 

Carcinoma, metastatic, Zymbal’s gland 

Trachea 

Special Senses System 

Zymbal’s gland 

Carcinoma 

Urinary System 

Kidney 

Lipoma 

Urinary bladder 

SystemicLesions 

MultipIe organs 

Leukemia mononuclear 

7777777777777777777777777 

3333333333333333333333333 

4444444455555555555666666 

3333333333333333333333333 Total 

2333444400112344555133455 Tissues/ 

7678125667588508038224346 Tumors 

......................... 50 

+ + 10 

++++++++M++++++++++++++++ 49 

1 

......................... 49 

......................... 50 

M++M++++M++++++++++++++++. 46 

......................... 50 

X 2 

xx x xx xx 20 

X 1 

......................... 50 

X 1 

X 1 

......................... 50 

......................... 50 

1 

+ 2 

......................... 50 

1 

......................... 50 

1 

......................... 50 

1 

1 

......................... 50 

X 1 

+++++++++++M+++++++++++++ 49 

......................... 50 

X X X 16 
122 Codeine, NTP TR 455 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 800 ppm 

34.56666666666666667777777 

Number of Days on Study 6280344456777888890002333 

5746511392037466740771333 

3343333434344334444334333 

Carcass ID Number 7706977081601862000781668 

2823936623'931510048119580 

AlimentarySystem 

Esophagus ......................... Intestine large, colon ......................... Intestine large, rectum ......................... Sarcoma, metastatic, uncertain primary site X 

Intestine large, cecum ++++++++++++++++++++A++++ 

Intestine small, duodenum .......................... Intestine small, jejunum ++++A++++++++++A++++AA+++ 

Intestine small, ileum ++A+A+++++++++++++++A++++ 

Liver .......................... Histiocytic sarcoma X 

Mesentery + ++ 

Pancreas ......................... Pharynx + 

Salivary glands ......................... Stomach, forestomach .......................... Stomach, glandular ......................... 

Cardiovascular System 

Blood vessel ......................... Heart ......................... 

Endocrine System 

Adrenal cortex ......................... Adenoma X 

Adrenal medulla ......................... Pheochromocytoma benign X 

Islets, pancreatic ......................... Adenoma 

Parathyroid gland M.++M++++++M++++M+++++M+++ 

Pituitary gland ++M++++++++++++++++++++++ 

Pars distalis, adenoma X X xx x x x x xx 

Thyroid gland .......................... C-cell, adenoma X 

Follicular cell, adenoma X 

Follicular cell, carcinoma X 

General Body System 

None ~~ ~~~~ 

Genital System 

Clitoral gland +++++++M+++++++++++++++++ 

Adenoma X X 

Bilateral, adenoma 

Ovary ......................... Oviduct + 

Utems .......................... Polyp stromal X x x 

Sarcoma X 

Sarcoma stromal X 

Vagina + 

Squamous cell papilloma X 
Lesions in Female Rats 123 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 800 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Sarcoma, metastatic, uncertain primary site 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Histiocytic sarcoma 

Mesentery 

Pancreas 

Pharynx 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

PiNitary gland 

Pars distalis, adenoma 

Thyroid gland 

C-cell, adenoma 

Follicular cell, adenoma 

Follicular cell, carcinoma 

General Body System 

None 

Genital System 

Clitoral gland 

Adenoma 

Bilateral, adenoma 

ovary

Oviduct 

UeNS 

Polyp stromal 

Sarcoma 

sarcoma stromal 

Vagina 

Squamous cell papilloma 

~~ 

7~777~777~777777777777177?777 

33333333333333333333333333 

34444444555555555556666666 

43333444333333333443333444 Total 

16778111778999999007889011 Tissues/ 

64043278796012467195893504 Tumors 

.......................... 51 

.......................... 51 

.......................... 51 

1 

.......................... 50 

.......................... 51 

.......................... 47 

.......................... 48 

.......................... 51 

1 

3. + + 6 

.......................... 51 

1 

.......................... 51 

.......................... 51 

.......................... 51 

.......................... 51 

.......................... 51 

.......................... 51 

1 

+++++++M++++++++++++++++++ 50 

1 

.......................... 51 

X 1 

++++++M++++M+++M+M+++M++++ 41 

........................... 50 

X x x x x x x X 19 

.......................... 51 

xx X X 1 

1 

.......................... 50 

X xxx X x x x X 11 

X 1 

.......................... 51 

1 

.......................... 51 

x x X 6 

1 

1 

+ 2 

1 
124 Codeine, NTP TR 455 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 800 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Histiocytic sarcoma 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Adenocarcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, sarcoma 

Tail, schwannoma malignant 

Musculoskeletal System 

Bone 

Skeletal muscle 

Sarcoma 

Nervous System 

Brain 

Respiratory System 

Lung 

Sarcoma, metastatic, uncertain primary site 

Nose 

Nasolacrimal duct, squamous cell carcinoma 

Trachea 

Special Senses System 

Ear 

Pinna, schwannoma malignant 

Eye

Harderian gland 

Zymbal’s gland 

Carcinoma 

Urinary System 

Kidney 

Nephroblastoma 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear 

3456666666666666667777777 

6280344456777888890002333 

5746511392037466740771333 

3343333434344334444334333 

7706977081601862000781668 

2823936623931510048119580 

......................... X 

+ ++++ 

X ......................... 

++++++++++M+++++++++M++++ 

......................... X 

++++++++++++++++++++M++++ 

++++++++++++++++++++M++++ 

xxx x xx X X 

xx X 

++++++++++++++++M++++++++ 

xx X 

X 

X 

......................... + 

X 

......................... 

......................... X ......................... 

......................... 

.+ 

X 

+ + + 

X X X 

......................... 

X 

......................... 

......................... X 

X X X x xxx 
Lesions in Female Rats 125 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 800 ppm (continued)' 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Histiocytic sarcoma 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Adenocarcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, sarcoma 

Tail, schwannoma malignant 

Musculoskeletal System 

Bone 

Skeletal muscle 

Sarcoma 

Nervous System 

Brain 

RespiratorySystem 

Lung 

Sarcoma, metastatic, uncertain primary site 

Nose 

Nasolacrimal duct, squamous cell carcinoma 

Trachea 

Special Senses System 

Ear 

Pinna, schwannoma malignant 

EY e Harderian gland 

Zymbal's gland 

Carcinoma 

Urinary System 

Kidney 

Nephroblastoma 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

117171111111711171171~1111 

33333333333333333333333333 

34444444555555555556666666 

43333444333333333443333444 Total 

16118111118999999007889011 Tissues/ 

64043218196012467195893504 Tumors 

.......................... 51 

1 

+ + + + 9 

1 

......................... 49 

.......................... 49 

.......................... 51 

1 

+++++++M++++++M++++++++M++ 41 

.......................... 50 

X X X 3 

X xxxx x x x x x xx x 21 

x x X 6 

.......................... 50 

3 

1 

1 

.......................... 51 

1 

1 

.......................... 51 

.......................... 51 

1 

.......................... 51 

X 1 

.......................... 51 

+ 

X 

+ 

+ 

.......................... 51 

1 

+++++++++++++M++++++++++++ 50 

.......................... 51 

1 

Leukemia mononuclear xx X X X xxx x 18
126 Codeine, NTP TR 455 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 1,600 ppm 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Oral mucosa 

Buccal, squamous cell carcinoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tongue 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

Bilateral, follicular cell, carcinoma 

C-cell, adenoma 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland 

Adenoma 

Ovary 

Uterus 

Polyp stromal 

Vagina 

3446666666667777777771777 

8471233468990011122333333 

3437418544181248905333334 

4444444444444444444444444 

2167664254252743426334562 

9308610159322762452782438 

......................... 

......................... 

......................... 

+++++++++++++A+++++++++++ 

++++++++++++++++A++++++++ 

++++++++++++++++A++++++++ 

+++A+++++A+++A++A++++++++ 

......................... + + 

+ 

X ......................... 

......................... 

A++++++,++++++++++++++++++ 

......................... + 

......................... 

......................... 

......................... 

......................... 

......................... 

++++M+++M++++++++++++++++ 

......................... X X 

......................... 

X X 

......................... 

......................... 

......................... X X X 
Lesions in Female Rats 127 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 1,600 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Oral mucosa 

Buccal, squamous cell carcinoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tongue 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Thyroid gland 

Bilateral, follicular cell, carcinoma 

C-cell, adenoma 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland 

Adenoma 

Ovary 

Uterus 

Polyp stromal 

Vagina 

77777777777777777777777777 

33333333333333333333333333 

44444455555555555566666666 

44444444444444444444444444 Total 

33456622344456677723345567 Tissues/ 

45804846613717901971957854 Tumors 

.......................... 51 

.......................... 51 

.......................... 51 

.......................... 50 

.......................... 50 

.......................... 50 

.......................... 47 

.......................... 51 

+ 3 

1 

1 

.......................... 51 

+++++++++++++++++++++++++M 50 

.......................... 50 

........................... 51 

1 

.......................... 51 

.......................... 51 

.......................... 51 

.......................... 51 

X 1 

.......................... 51 

+++++++++++++M++M+++++++M+ 46 

.......................... 51 

X xxx xx X xxx 12 

.......................... 51 

X 1 

X 3 

X 1 

.......................... 51 

X 1 

.......................... 51 

.......................... 51 

3 

+ 1 
128 Codeine, NTP TR 455 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-YearFeed Study of Codeine: 1,600 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

Thymoma benign 

IntegumentarySystem 

Mammary gland 

Adenoacanthoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Subcutaneous tissue, fibroma 

Musculoskeletal System 

Bone 

Nervous System

Brain 

Respiratory System 

Lung

Alveolarhronchiolar adenoma 

Alveolarhronchiolar carcinoma 

Nose 

Sarcoma 

Trachea 

Special Senses System 

EY e Harderian gland 

Zymbal's gland 

Carcinoma 

Urinary System 

Kidney 

Renal tubule, carcinoma 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

3446666666667777777777777 

8471233468990011122333333 

3437418544181248905333334 

4444444444444444444444444 

2767664254252743426334562 

9308610159322762452782438 

......................... +++ ++ + + '+ 

+++++++++M+++++++++++++++ 

......................... 

......................... 

+++++++++++++M+++++++++++ 

~~~~ ~ ~~ ~ ~ 

......................... 

X X X 

......................... 

......................... 

.......................... 

......................... X 

......................... X 

......................... 

+ + + 

+ 

+ 

X 

......................... X 

+M+++++++++++++++++++++++ 

......................... xxx xxx X
Lesions in Female Rats 129 

TABLEB2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of Codeine: 1,600 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

Thymoma benign 

Integumentary System 

Mammary gland 

Adenoacanthoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Subcutaneous tissue, fibroma 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Respiratory System 

Lung 

Alveolarlbronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Nose 

Sarcoma 

Trachea 

Special Senses System 

EY e 

Harderian gland 

Zymbal’s gland 

Carcinoma 

Urinary System 

Kidney 

Renal tubule, carcinoma 

Urinary bladder 

~~ ~ 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

77777777777777777777777777 

33333333333333333333333333 

44444455555555555566666666 

44444444444444444444444444 Total 

33456622344456677723345567 Tissues/ 

45804846613717901971957854 Tumors 

.......................... 51 

+ + + 11 

+++++++++++++++++++++++++M 49 

.......................... 51 

.......................... 51 

++++++++++M++M+M+M++++++++ 46 

X 1 

.......................... 51 

X 1 

X X X 6 

xx 2 

.......................... 51 

X 1 

.......................... 51 

.......................... 51 

.......................... 51 

1 

X 1 

.......................... 51 

1 

.......................... 51 

+ + 

.......................... 51 

1 

.......................... 50 

.......................... 51 

X 12 
130 Codeine, NTP TR 455 

TABLEB3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-YearFeed Study of Codeine 

400 ppm 

5/50 (10%) 

12.2% 

3/38 (8%) 

689 

P=O.362 

P=O.296 

P=O.218 

5/50 (10%) 

12.2% 

3/38 (8%) 

689 

P=O.535 

P=O.474 

P=O.357 

6/47 (13%) 

15.7% 

5/36 (14%) 

546 

P=0.566N 

P=O.596 

P=O.500 

6/47 (13%) 

15.7% 

5/36 (14%) 

546 

P=0.430N 

P=0.568N 

P=0.621N 

21/50 (42%) 

47.4% 

15/38 (39%) 

400 

P=0.019N 

P=0.087N 

P=0.158N 

800 ppm 

1/50 (2%) 

2.7% 

0128 (0%) 

686 

P=0.457N 

P=0.472N 

P=0.500N 

1/50 (2%) 

2.7% 

0128 (0%) 

686 

P=0.275N 

P=0.266N 

P=0.309N 

12/50 (24%) 

37.8% 

10/29 (34%) 

662 

P=O.O81 

P=O.115 

P=O.O71 

12/50 (24%) 

37.8% 

10129 (34%) 

662 

P=O.145 

P=O.189 

P=O.123 

27/5 1 (53 %) 

68.2% 

17/29 (59%) 

641 

P=0.419N 

P=0.359N 

P=0.537N 

1,600 ppm 

1/51 (2%) 

3.1% 

1/32 (3X) 

733 (T) 

P=0.438N 

P=0.437N 

P=0.492N 

1/51 (2%) 

3.1% 

1/32 (3%) 

733 (T) 

P=0.249N 

P=0.235N 

P=0.301N 

1/51 (2%) 

3.1% 

1/32 (3%) 

733 (T) 

P=0.071N 

P=0.054N 

P=0.085N 

1/51 (2%) 

3.1% 

1/32 (3%) 

733 (T) 

P=0.037N 

P=0.033N 

P=O.WN 

8/51 (16%) 

23.0% 

6/32 (19%) 

645 

P<0.001N 

P<0.001N 

P<0.001N 

AdrenalMedulla:BenignPheochromocytoma 

Overall ratea 

Adjusted rateb 

Terminal rate' 

First incidence (days) 

Life table testd 

Logistic regression test' 

Cochran-Armita e testd 

Fisher exact test %

AdrenalMedulla:Benign 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

ClitoralGland:Adenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

2/50 (4%) 

6.6% 

1/28 (4%) 

677 

P=0.171N 

P=0.158N 

P=0.188N 

or Malignant Pheochromocytoma 

3/50 (6%) 

10.0% 

2/28 (7%) 

677 

P=0.099N 

P=0.087N 

P=O.llON 

5/47 (1 1 %) 

16.7% 

3/27 (1 1 X) 

663 

P=0.115N 

P=0.078N 

P=O. 115N 

ClitoralGland:AdenomaorCarcinoma 

Overall rate 6/47 (13%) 

Adjusted rate 19.0% 

Terminal rate 3/27 (1 1%) 

First incidence (days) 587 

Life table test P=0.075N 

Logistic regression test P=0.052N 

Cochran-Amitage test P=0.075N 

Fisher exact test 

MammaryGland: fibroadenoma 

Overall rate 27/50 (54%) 

Adjusted rate 70.346 

Terminal rate 17/28 (61A) 

First incidence (days) 475 

Life table test P<0.001N 

Logistic regression test P<0.001N 

Cochran-Annitage test P<0.001N 

Fisher exact test 
Lesions in Female Rats 131 

TABLEB3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

MammaryGland:Adenocarcinoma 

Overall rate 3/50 (6%) 2/50 (4%) 3/51 (6%) 0151 (0%) 

Adjusted rate 8.5% 5.3% 10.3% 0.0% 

Terminal rate 1/28 (4%) 2/38 (5%) 3/29 (10%) 0132 (0%) 

First incidence (days) 495 733 (T) -e 733 0 

Life table test P=0.388NP=O.lOlN 

Logistic regression test P=0.533NP=0.102N 

Cochran-Armitage test P=0.107N 

Fishertest exact . 

MammaryGland:Fibroadenoma,Adenoacanthoma,orAdenocarcinoma 

Overall rate 30/50 (60%) 

Adjusted rate 74.4% 

Terminal rate 18/28 (64%) 

First incidence (days) 475 

Life table test P<0.001N 

Logistic regression test P<0.001N 

Cochran-Armitage test P<0.001N 

Fisher exact test 

PituitaryGland(ParsDistalis):Adenoma 

Overall rate 16/46 (35%) 

Adjusted rate 50.6% 

Terminal rate 12/27 (44%) 

First incidence (days) 495 

Life table test P=O.l42N 

Logistic regression test P=0.072N 

Cochran-Armitage test P=0.129N 

Fisher exact test 

PituitaryGland(Pars Distalis): AdenomaorCarcinoma 

Overall rate 17/46 (37%) 

Adjusted rate 53.9% 

Tenninal rate 13/27 (48%) 

First incidence (days) 495 

Life table test P=0.094N 

Logistic regression test P=0.041N 

Cochran-Armitage test P=0.082N 

Fisher exact test 

Skin (SubcutaneousTissue):Fibroma 

Overall rate 2/50 (4%) 

Adjusted rate 5.8% 

Terminal rate 1/28 (4%) 

First incidence (days) 547 

Life table test P=0.445N 

Logistic regression test P=0.475N 

Cochm-Armitage test P=0.465N 

Fisher exact test 

P=0.500N 

23/50 (46%) 

52.0% 

17/38 (45%) 

400 

P=O.OllN 

P=0.063N 

P=0.115N 

17/50 (34%) 

42.2% 

15/38 (39%). 

546 

P=0.239N 

P=0.279N 

P=0.553N 

18/50 (36%) 

43.7% 

15/38 (39%) 

546 

P=0.224N 

P=0.257N 

P=0.545N 

1/50 (2%) 

2.6% 

1/38 (3X) 

733 cr)

P=0.422N 

P=0.526N 

P=0.500N 

P-0.629N 

P=0.650N 

P=0.652N 

29/51 (57%) 

73.5% 

19/29 (66%) 

641 

P=0.344N 

P=O.284N 

P=0.453N 

19/50 (38%) 

49.3% 

11/29 (38%) 

427 

P=O.484 

P=0.559N 

P=O.455 

19/50 (38%) 

49.3% 

11/29 (38%) 

427 

P=O.565 

P=0.456N 

P=O.542 

3/51 (6%) 

7.8% 

0129 (0%) 

659 

P=O.557 

P=O.484 

P=O.509 

P=0.100N 

P=0.147N 

P=O.l18N 

8/51 (16%) 

23.0% 

6/32 (19%) 

645 

P<O.OOlN 

P<O.OOlN 

P<O.OOlN 

12/51 (24%) 

34.3% 

10132 (31%) 

63 1 

P=0.105N 

P=0.054N 

P=0.159N 

12/51 (24%) 

34.3% 

10/32. (31%) 

63 1 

P=0.069N 

P=0.031N 

P=O.lllN 

1/51 (2%) 

3.1 I 

1/32 (3%) 

733 (T) 

P=0.456N 

P=0.508N 

P=0.492N 
132 Codeine, NTP TR 455 

TABLEB3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

Skin (Subcutaneous Tissue): Fibroma,Fibrosarcoma,orSarcoma 

Overall rate 3/50 (6%) 1/50 (2%) 4/51 (8%) 1/51 (2%) 

Adjusted rate 7.7% 2.6% 10.7% 3.1% 

Terminal rate 1/28 (4%) 1/38 (3%) 0/29 (0%) 1/32 (3%) 

First incidence (days) 313 733 (T) 659 733 (T)

Life table test P=0.316N P=0.250N P=O.555 P=0.273N 

Logistic regression test P=0.403N P=0.488N P=O.386 P=0.436N 

Cochran-Armitage test P=0.334N 

Fisher exact test P=0.309N P=O.511 P=0.301N 

ThyroidGland(C-cell):Adenoma 

Overall rate 4/50 (8%) 3/50 (6%) 5/51 (10%) 3/51 (6%) 

Adjusted rate 12.6% 6.9% 16.1% 8.7% 

Terminal rate 3/28 (1 1 X) 1/38 (3%) 4/29 (14%) 2/32 (6%) 

First incidence (days) 475 606 686 702 

Life table test P=0.444N P=0.360N P=O.531 P=0.430N 

Logistic regression test P=0.448N P=0.538N P=O.564 P=0.461N 

Cochran-Armitage test P=0.469N 

Fisher exact test P=0.500N P=O.513 P=0.489N 

ThyroidGland(C-cell):AdenomaorCarcinoma 

Overall rate 6/50 (12%) 4/50 (8%) 5/51 (10%) 3/51 (6%) 

Adjusted rate 18.9% 9.4% 16.1% 8.7% 

Terminal rate 4/28 (14%) 2/38 (5%) 4/29 (14%) 2/32 (6%) 

First incidence (days) 475 606 686 702 

Life table test P=0.198N P=0.219N P=0.470N P=0.183N 

Logistic regression test P=0.189N P=0.353N P=0.417N P=0.191N 

Cochran-Armitage test P=0.215N 

Fisher exact test P=0.370N P=0.486N P=0.234N 

Uterus:Stromal Polyp 

Overall rate 1/50 (2%) 6/50 (12%) 6/51 (12%) 3/51 (6%) 

Adjusted rate 3.6% 15.0% 19.2% 6.9% 

Terminal rate 1/28 (4%) 5/38 (13%) 5/29 (17%) 0132 (0%) 

First incidence(days) 733 (T) 546 662 473 

Life table test P=O.463 P=O.114 P=O.O68 P=O.368 

Logistic regression test P=O.454 P=O.O76 P=O.O82 P=O.256 

Cochran-Armitage test P=O.444 

Fisher exact test P=O.O56 P=O.O59 P=O.316 

Uterus:StromalPolyp orStromalSarcoma 

Overall rate 1/50 (2%) 6/50 (12%) 7/51 (14%) 3/51 (6%) 

Adjusted rate 3.6% 15.0% 21.3% 6.9% 

Terminal rate 1/28 (4%) 5/38 (13%) 5/29 (17%) 0132 (0%) 

First incidence (days) 733 (T) 546 662 473 

Life table test P=O.455 P=O.114 P=O.O43 P=O.368 

Logistic regression test P=O.438 P=O.O76 P=O.O51 P=O.256 

Cochran-Armitage test P=O.429 

Fisher exact test P=O.O56 P=O.O32 P=O.316 
Lesions in Female Rats 133 

TABLEB3 

Statistical Analysisof Primary Neoplasms in Female Rats in the 2-YearFeed Study of Codeine (continued) 

Zymbal’s Gland: Carcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Annitage test 

Fisher exact test 

All Organs:MononuclearCell Leukemia 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

All Organs: Benign Neoplasms 

Overall rate 

Adjusted rate 

Terminal mte 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Annitage test 

Fisher exact test 

All Organs: Malignant Neoplasms 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

0150 (0%) 

0.0% 

0/28 (0%) -

P=O.401 

P=O.341 

P=O.358 

14/50 (28%) 

35.3% 

5/28 (18%) 

432 

P=0.267N 

P=0.353N 

P=0.311N 

39/50 (78%) 

92.7% 

25/28 (89%) 

475 

P=O.o06N 

P<0.001N 

P=0.001N 

27/50 (54%) 

61.7% 

12/28 (4396) 

313 

P=O.O6ON 

P=0.076N 

P=0.051N 

400 PPm 

1/50 (246) 

2.4% 

0138 (0%) 

705 

P=O.567 

P=O.525 

P=O.500 

16/50 (32%) 

34.3% 

8/38 (21%) 

479 

P=0.441N 

P=O.272 

P=O.414 

40/50 (80%) 

83.3% 

30138 (79%) 

400 

P=0.059N 

P-0.460N 

P=O.500 

20/50 (40%) 

41.6% 

10138 (26%) 

479 

P=0.027N 

P=0.228N 

P=O. 115N 

800 ppm 

3/51 (6%) 

7.1 96 

0/29 (0%) 

606 

P=O.169 

P=O.102 

P=O.125 

18/51 (35%) 

52.196 

13/29 (4546) 

635 

P=O.369 

P10.305 

P=O.283 

41/51 (80%) 

90.9% 

25/29 (86%) 

427 

P=0.484N 

P=0.444N 

P=O.480 

28/51 (55%) 

66.1% 

15/29 (52%) 

365 

P=0.471N 

P=O.484 

P=O.543 

1,600 ppm 

1/51 (2%) 

2.8% 

0132 (0%) 

718 

P=O.543 

P=O.539 

P=O.505 

12/51 (24%) 

27.3% 

2/32 (6%) 

617 

P=0.248N 

P=0.587N 

P=0.387N 

27/51 (53%) 

65.1% 

18/32 (56%) 

473 

P=0.003N 

P=0.001N 

P=0.007N 

17/51 (33%) 

36.9% 

4/32 (13%) 

444 

P=0.021N 

P=0.061N 

P=0.029N 
134 Codeine, NTP TR 455 

TABLEB3 

Statistical Analysisof Primary Neoplasms in Female Rats in the2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

All Organs: Benign or Malignant Neoplasms 

Overall rate 46/50 (92%) 44/50 (88A) 47/51 (92%) 37/51 (73%) 

Adjusted rate 95.8% 88.0% 95.9% 75.5% 

Terminal rate 26/28 (93%) 32/38 (84%) 27/29 (9396) 20/32 (63%) 

First incidence (days) 313 400 365 444 

Life table test P=0.041N P=0.019N P=0.382N P=0.018N 

Logistic regression test P=0.004N P=0.422N P=0.633N P=0.012N 

Cochran-Armitage test P=0.003N 

Fisher exact test P=0.370N P=O.OlONP=O.631 

(“)Terminal sacrifice 

a Number of neoplasm-bearing animaldnumber of animals examined. Denominator is number of animals examined microscopically for adrenal gland, 

clitoral gland, mammary gland, pituitary gland, thyroid gland, and uterus; for other tissues, denominator is number of animals necropsied. 

Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to 

pairwise comparisons between the controls and that exposed group. The life table test regards neoplasms in animals dying prior to terminal kill as being 

(directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests 

compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasmsin animal group 
Lesions in Female Rats 135 

TABLEB4a 

Historical Incidence of Benign Pheochromocytomas of the Adrenal Medulla in Untreated FemaleF344/NRatsa 

Incidence in Controls 

Historical Incidence at Microbiological Associates, Inc. 

C.I. Direct Blue 218 1/49 

dl-Amphetamine Sulfate 2/49 

Overall Historical Incidence 

Total 4911,175 (4.2%) 

Standard deviation 2.5% 

Range 0%-8% 

a Dataasof 17 June 1994 

TAB^ B4b 

Historical Incidence of Clitoral Gland Neoplasms in Untreated Female F344/N Ratsa 

Incidence in Controls 

Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Microbiological Associates, Inc. 

C.I. Direct Blue 218 2/47 0147 2/47 

dl-Amphetamine Sulfate 2/39 0/39 2/39 

Overall Historical Incidence 

Total 92/1,117 (8.2%) 29/1,117 (2.6%) 119/1,117 (10.7%) 

Standard deviation 4.3% 3.8% 5.6% 

Range 0%-19% 0%-15% 2%-21% 

a Dataasof 17 June 1994 
136 Codeine, NTP TR 455 

TABLEB4c 

Historical Incidence of Mammary Gland Neoplasms in Untreated Female F344/NRatsa 

Incidence in Controls ~ ~~~ ~ ~~~ ~~ ~ ~~ 

Fibroadenoma Carcinoma Fibroadenoma, 

Adenoma, or Carcinoma 

Historical Incidence at Microbiological Associates, Inc. 

C.I. Direct Blue 218 

dl-Amphetamine Sulfate 

Overall Historical Incidence 

Total 

Standard deviation 

Range 

a Data as of 17 June 1994 

TABLEB4d 

19/50 3/50 22/50 

2 1/50 3/50 25/50 

46Y1.202 (38.7%) 3211,202 (2.7%) 50711,202 (42.2%) 

12.7% 2.9% 13.5% 

876-5846 0%-10% 8%-64% 

Historical Incidence of Pituitary Gland (Pars Distalis) Neoplasms in Untreated Female F344/NRatsa 

Incidence in Controls 

Adenoma CarcinomaAdenomaor Carcinoma 

Historical Incidence at Microbiological Associates, Inc. 

C.I. Direct Blue 218 28/50 2/50 30150 

dl-Amphetamine Sulfate 31/50 0150 3 1/50 

Overall Historical Incidence 

Total 65211,196 (54.5%) 1311,196 (1.1%) 66511,196 (55.6%) 

Standard deviation 12.5% 1.4% 13.0% 

Range 30%-76% OW% 30%-80% 

a Data as of 17 June 1994; includesdataforunspecifiedsite . , 
Lesions in Female Rats 137 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study of Codeinea 

0 PPm 400 ppm 800 ppm 1,600 ppm 

DispositionSummary 

Animals initially in study 

15-Month interim evaluation 

Early deaths 

Moribund 

Natural deaths 

Survivors 

Died last week of study 

Terminal sacrifice 

Animals examined microscopically 

15-Month Interim Evaluation 

AlimentarySystem 

Liver 

Basophilic focus 

Developmental malformation 

Eosinophilic focus 

Hepatodiaphragmatic nodule 

Hyperplasia, focal 

Inflammation 

Necrosis 

Bile duct, hyperplasia, focal 

Mesentery 

Fat, necrosis 

Pancreas 

Infiltration cellular, lymphocyte 

Acinar cell, atrophy 

Cardiovascular System 

Heart 

Cardiomyopathy 

Endocrine System 

Adrenal cortex^ 

Hemorrhage 

Hyperplasia, focal 

Adrenal medulla 

Inffltration cellular, lymphocyte 

Pituitary gland 

Pars distalis, angiectasis 

Pars distalis, cyst 

Pars distalis, hyperplasia 

Pars distalis, hyperplasia, focal 

Pars distalis, pigmentation 

60 

10 

15 

7 

28 

60 

1 (10%) 

1 (10%) 

3 (30%) 

3 (30%) 

2 (20%) 

60 

10 

9 

3 

1 

37 

60 

1 (10%) 

3 (30%) 

60 60 

9 9 

17 14 

5 5 

1 

28 32 

60 60 

1 (11%) 

1 (11%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
138 Codeine, NTP TR 455 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 

15-Month Interim Evaluation (continued) 

Genital System 

Clitoral gland (10)

Ectasia 1 (10%) 

Hyperplasia 

Inflammation 1(10%) 

Ovary (10) 

cyst 1 (10%) 

UteNS (10)

Dilatation 1(10%) 

HematopoieticSystem 

Spleen 

Developmental malformation 

Thymus 

Atrophy

Hemorrhage 

IntegumentarySystem 

Mammary gland 

Dilatation 

Respiratory System 

Lung 

Inffltration cellular, histiocyte 

Inflammation 

Alveolar epithelium, hyperplasia 

Nose 

Edema, focal 

Fungus 1(10%) 

Hemorrhage 

Infiltration cellular, focal, lymphocyte 

Inflammation 2 (20%) 

Olfactory epithelium, cytoplasmic alteration 8 (80%) 

Special Senses System 

Eye

Lens, cataract 

Retina, atrophy 

Urinary System 

Kidney (10) 

Infarct 

Mineralization 1(10%) 

Nephropathy, chronic 7 (70%) 

400 ppm 

(10) 

1(10%) 

(10) 

2(20%) 

(10) 

1(10%) 

2 (20%) 

10 (100%) 

(10) 

1(10%) 

1 (10%) 

10 (100%) 

800 ppm 

(9) 

1 (11%) 

(9) 

1 (11%) 

(9) 

1 (11%) 

(9) 

2 (22%) 

4 (44%) 

1(11%) 

(9) 

3 (33%) 

9 (100%) 

(9) 

9 (100%) 

1,600 ppm 

(9) 

1 (11%) 

(9) 

(9) 

1 (11%) 

(9) 

2(22%) 

2 (22%) 

(9) 

1(11%) 

1(11%) 

1(11%) 

8 (89%) 

(9) 

5 (56%) 
Lesions in Female Rats 139 

400 ppm 

(50) 

1 (2%) 

(48) 

(50) 

36 (72%) 

4 (8%) 

2 (4%) 

2 (4%) 

4 (8%) 

5 (10%) 

1 (2%) 

1 (2%) 

(5) 

1 (20%) 

3 (60%) 

(50) 

1 (2%) 

12 (24%) 

1 (2%) 

(49) 

. 800 ppm 

(51) 

(50) 

(5 1) 

31 (61%) 

1 (2%) 

3 (6%) 

4 (8%) 

1 (2%) 

4 (8%) 

2(4%) 

10 (20%) 

1 (2%) 

1 (2%) 

3 (6%) 

1 (2%) 

(6) 

5 (83%) 

(51) 

1 (2%) 

1(2%) 

20 (39%) 

(1) 

1 (100%) 

(51) 

1,600 ppm 

(51) 

(50) 

1 (2%) 

(51) 

29 (57%) 

2 (4%) 

3 (6%) 

1 (2%) 

1 (2%) 

3 (6%) 

1 (2%) 

2(4%) 

9 (18%) 

1 (2%) 

(3) 

3 (100%) 

(1) 

(51) 

1 (2%) 

24 (47%) 

(50) 

0 PPm 

15-Month InterimEvaluation (continued) 

Systems Examined With No Lesions Observed 

General Body System 

Musculoskeletal System 

Nervous System 

2-Year Study 

Alimentary System 

Intestine large, colon 

Inflammation 

Intussusception 

Intestine large, cecum 

Inflammation 

Inflammation, granulomatous 

Liver 

Basophilic focus 

Clear cell focus 

Congestion 

Degeneration 

Developmental malformation 

Eosinophilic focus 

Fatty change 

Hematopoietic cell proliferation 

Hepatodiaphragmatic nodule 

Infiltration cellular, lymphocyte 

Inflammation 

Leukocytosis 

Mixed cell focus 

Necrosis 

Centrilobular, degeneration 

Mesentery 

Infiltration cellular, lymphocyte 

Fat, inflammation 

Fat, necrosis 

Lymphatic, cyst

Oral mucosa 

Gingival, cyst 

Pancreas 

Accessory spleen 

Infdtration cellular, lymphocyte 

Inflammation 

Acinar cell, atrophy 

Acinar cell, hyperplasia 

Artery, inflammation 

Pharynx 

Palate, hyperplasia 

Salivary glands 

Atrophy, focal 

(50) 

1 (2%) 

(50) 

1 (2%) 

(50) 

38 (76%) 

5 (10%) 

1 (2%) 

3 (6%) 

5 (10%) 

10 (20%) 

2(4%) 

2 (4%) 

2(4%) 

4 (8%) 

1 (2%) 

2 (4%) 

(6) 

1(17%) 

5 (83%) 

1 (17%) 

(3) 

1 (33%) 

(50) 

1 (2%) 

2(4%) 

24 (48%) 

1 (2%) 

(50) 

1 (2%) 
140 Codeine, NTP TR 455 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in FemaleRats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

2-Year Study (continued) 

Alimentary System (continued) 

Stomach, forestomach 

Hyperplasia 

Inflammation 

Ulcer 

Stomach, glandular 

Fibrosis, focal 

Necrosis 

Ulcer 

Tongue 

Inflammation 

Cardiovascular System 

Heart 

Cardiomyopathy 

HYPe*oPhY 

Infiltration cellular, lymphocyte 

Atrium, dilatation 

Amum, thrombosis 

CoronaIy artery, inflammation 

Valve, inflammation, chronic 

Valve, pigmentation 

EndocrineSystem 

Adrenal cortex 

Accessory adrenal cortical nodule 

Angiectasis 

Congestion 

Cytologic alterations 

Cytoplasmic alteration 

Degeneration 

Fibrosis 

Hematopoietic cell proliferation 

Hemorrhage 

Hyperplasia, focal 

Hypertrophy

Necrosis 

Thrombosis 

Vacuolization cytoplasmic 

Vacuolization cytoplasmic, focal 

Capsule,inflammation 

Adrenal medulla 

Hyperplasia 

Hyperplasia,focal 

Infiltration cellular, lymphocyte 

Islets, pancreatic 

Hyperplasia 

Parathyroid gland 

Hyperplasia 

(50) 

12 (24%) 

1 (2%) 

(50) 

13 (26%) 

1 (2%) 

2 (4%) 

7 (14%) 

(50) 

8 (16%) 

1 (2%) 

(50) 

(45) 

(50) 

7 (14%) 

1 (2%) 

(50) 

6 (12%) 

9 (18%) 

1(2%) 

6 (12%) 

(50) 

5 (10%) 

(50) 

(37) 

3(8%) 

(51) 

15 (29%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

(51) 

6 (12%) 

1 (2%) 

1(2%) 

1 (2%) 

1 (2%) 

8 (16%) 

1 (2%) 

1(2%) 

4(8%) 

(50) 

2(4%) 

(51) 

(41) 

(51) 

9 (18%) 

(51) 

1(2%) 

3(6%) 

2(4%) 

1 (2%) 

1(2%) 

3(6%) 

1 (2%) 

1 (2%) 

2(4%) 

8 (16%) 

1(2%) 

(51) 

2 (4%) 

1(2%) 

(51) 

1 (2%) 

(46) 
Lesions in Female Rats 141 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

2-Year Study (continued) 

Endocrine System (continued) 

Pituitary gland 

Developmental malformation 

Pars distalis, angiectasis 

Pars distalis, cyst 

Pars distalis, hyperplasia 

Pars intermedia, angiectasis 

Thyroid gland 

C-cell, hyperplasia 

General Body System 

None 

GenitalSystem 

Clitoral gland 

Ectasia 

Hyperplasia 

Inflammation 

Ovary 

cyst 

Inflammation, granulomatous 

Corpus luteum, hypertrophy 

UkNS 

cyst 

Dilatation 

Fibrosis 

Hemorrhage 

Endometrium, hyperplasia 

Vagina 

Inflammation 

Prolapse 

HematopoieticSystem 

Bone marrow 

Hemorrhage 

Hyperplasia 

Infiltration cellular, histiocyte 

Myelofibrosis 

Erythroid cell, atrophy 

Erythroid cell, hyperplasia 

Myeloid cell, hyperplasia 

Lymph node 

Iliac, hyperplasia, plasma cell 

Mediastinal, cyst 

Mediastinal, hemorrhage 

Mediastinal, pigmentation 

Pancreatic, pigmentation 

Renal, hemorrhage 

Renal, pigmentation 

(46) 

13, (28 96) 

13 (28%) 

13 (28%) 

(50) 

4 (8%) 

(47) 

5 (11%) 

9 (19%) 

2 (4%) 

(50) 

7 (14%) 

16 (32%) 

18 (36%) 

(50) 

11 (22%) 

(47) 

9 (19%) 

7 (15%) 

2 (20%) 

2 (20%) 

1 (10%) 

(50) 

10 (20%) 

16 (32%) 

13 (26%) 

1 (2%) 

(51) 

5 (10%) 

(50) 

7 (14%) 

8 (16%) 

1 (2%) 

(51) 

4 (8%) 

6 (12%) 

1 (2%) 

2 (4%) 

(2) 

1 (50%) 

8 (16%) 

2 (22%) 

2 (22%) 

2 (22%) 

1 (11%) 

(51) 

1 (2%) 

7 (14%) 

14 (27%) 

14 (27%) 

(51) 

5 (10%) 

4 (8%) 

1 (2%) 

2 (4%) 

(1) 

1 (100%) 

1 (100%) 

1 (2%) 

4 (36%) 

2 (18%) 
142 Codeine, NTP TR 455 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Studyof Codeine (continued) 

study (continued) 

Hematopoietic System (continued) 

Lymph node, mandibular 

Congestion 

Edema 

Hemorrhage 

Hyperplasia, lymphoid 

Hyperplasia, plasma cell 

Infiltration cellular, histiocyte 

Pigmentation 

Lymph node, mesenteric 

Edema 

Hemorrhage 

Infiltration cellular, histiocyte 

Spleen 

Congestion 

Depletion lymphoid 

Hematopoietic cell proliferation 

Hyperplasia, lymphoid 

Necrosis 

Pigmentation ’ Thymus 

Amyloid deposition 

Atrophy

Congestion 

cyst 

Hemorrhage 

IntegumentarySystem 

Mammary gland 

Galactocele 

Hyperplasia 

Infiltration cellular, histiocyte 

Inflammation 

Lactation 

Skin 

Acanthosis 

Subyaneous tissue, cyst epithelial 

inclusion 

Tail, fibrosis 

Musculoskeletal System 

Bone 

Cranium, hyperostosis 

Femur, hyperostosis 

Maxilla, hyperostosis 

Turbinate, hyperostosis 

0 PPm 

2(4%) 

2(4%) 

1(2%) 

1(2%) 

(50) 

1(2%). 

1 (2%) 

1 (2%) 

(50) 

1(2%) 

1(2%) 

5 (10%) 

29 (66%) 

1 (2%) 

2 (5%) 

(50) 

1 (2%) 

8(16%) 

400 ppm 

(49) 

1(2%) 

3 (6%) 

(49) 

2(4%) 

(50) 

1(2%) 

2 (4%) 

1 (2%) 

1(2%) 

(46) 

1 (2%) 

29 (63%) 

1 (2%) 

(50) 

2 (4%) 

7 (14%) 

1 (2%) 

1 (2%) 

800 ppm 

(49) 

1(2%) 

4 (8%) 

1(2%) 

1(2%) 

(49) 

3 (6%) 

(51) 

5 (10%) 

1(2%) 

2(4%) 

(47) 

23 (49%) 

(50) 

3 (6%) 

7 (14%) 

3 (6%) 

1(2%) 

24 (48%) 

(50) 

1,600 ppm 

(49) 

3 (6%) 

3. (6%) 

6 (12%) 

1 (2%) 

1(2%) 

2(4%) 

(46) 

23 (50%) 

1(2%) 

2 (4%) 

(51) 

8 (16%) 

16 (31%) 

(5 1) 

1(2%) 
Lesions in FemaleRats 143 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

2-Year Study (continued) 

Nervous System 

Brain (50)

Compression

(51) (50) (51) 

Degeneration, focal 

8 (16%) 4 (8%) 10 (20%) 4 (8%) 

1 (2%) 

Meninges, thrombosis, focal 

Meninges, inflammation, focal 

Necrosis 

Hydrocephalus 

Hemorrhage 

2 (4%) 

1 (2%) 

1 (2%) 

2 (4%) 

4 (8%) 

2 (4%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

Respiratory System 

Lung 

Congestion 

(51) 

2 (4%) 

(51) 

1 (2%) 

Mitration cellular, lymphocyte 

Hemorrhage 

1 (2%) 

3 (6%) 

1 (2%) 

Infitration cellular, histiocyte 7 (14%) 15 (29%) 

Inflammation 4 (8%) ’ 13 (25%) 

Leukocytosis 

Alveolar epithelium, hyperplasia 1 (2%) 

Cytoplasmic alteration 

Nose 

1 (2%) 

(51) (51) 

Foreign body 

Edema, focal 

1 (2%) 

2 (4%) 

Thrombosis 

Inflammation 5 (10%) 

1 (2%) 

4 (8%) 

Nasolacrimal duct, inflammation 

Nares, hyperplasia 

3 (6%) 3 (6%) 

1 (2%) 

Olfactory epithelium, atrophy, focal 2 (4%) 1 (2%) 

Olfactory epithelium, cytoplasmic alteration 42 (82%) 49 (96%) 

Respiratory epithelium, hyperplasia 

Infitration cellular, lymphocyte 

Trachea 

2 (4%) 

(51) (5 1) 

1 (2%) 

Special Senses System 

Eye 

Phthisis bulbi 

Hemorrhage 

(1) 

1 (100%) 

(5) 

1 (20%) 

1 (20%) 

Cornea, inflammation 1 (20%) 

Lens, cataract 1 (100%) 2 (40%) 

Retina, degeneration 

Lens, hyperplasia 1 (100%) 

Harderian gland 

Infiltration cellular, lymphocyte 

Inflammation 
144 Codeine, NTP TR 455 

TABLEB5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the2-YearFeed Study of Codeine (continued) 

0 PPm 400 ppm 800 ppm 1,600 ppm 

Urinary System 

Kidney 

cyst 

(50) (51) 

1 (2%) 

Infarct 1 (2%) 3 (6%) 1 (2%) 

Infiltration cellular, lymphocyte 1 (2%) 1 (2%) 

Inflammation 1 (2%) 

Mineralization 3 (6%) 1 (2%) 1 (2%) 

Necrosis 1 (2%) 2 (4%) 

Nephropathy,chronic 39 (78%) 41 (82%) 40 (78%) 40 (78%) 

Bilateral, hydronephrosis 1 (2%) 

Transitional epithelium, hyperplasia, focal 

Infiltration cellular, lymphocyte 

Urinary bladder (50) (49) 

1 (2%) 

(50) (50) 

1 (2%) 
145 

APPENDIX C 

SUMMARY OF LESIONS IN MALE MICE 

IN THE 2-YEAR FEED STUDY 

OF CODEINE 

TABLEC1 Summary of the Incidence of Neoplasms in Male Mice 

in the 2-Year Feed Study of Codeine ................................. 146 

TABLEC2 Individual Animal Tumor Pathology of Male Mice 

in the 2-Year Feed Study of Codeine ................................. 150 

TABLEC3 Statistical Analysis of Primary Neoplasms in Male Mice 

in the 2-Year Feed Study of Codeine ................................. 166 

TABLEC4a Historical Incidence of Hepatocellular Neoplasms 

in Untreated Male B6C3Fl Mice .................................... 169 

TABLEC4b Historical Incidence of Malignant Lymphoma 

in Untreated Male B6C3Fl Mice .................................... 169 

TABLE CS Summary of the Incidence of Nonneoplastic Lesions in Male Mice 

in the 2-Year Feed Study of Codeine ................................. 170 
146 Codeine, NTP TR 455 

TABLEC1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-YearFeed Study of Codeinea 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

Disposition Summary 

Animals initially in study 60 60 60 60 

15-iUonth interim cvalualion 10 10 10 10 

Early deaths 

Accidental death 1 

Moribund 4 4 2 1 

Natural deaths 5 7 3 6 

Survivors 

Terminal sacrifice 41 38 45 43 

Animals examined microscopically 60 60 60 60 

15-Month Interim Evaluation 

Alimentary System 

Liver (10) (10). (10) (10)

Hepatocellular carcinoma 1 (10%) 

Hepatocellular adenoma 3 (30%) 1 (10%) 2 (20%) 1 (10%) 

Hematopoietic System 

Bone marrow 

Fibroma, osseous 

Respiratory System 

Lung (10) (10)

Alveolar/bronchiolar adenoma 1 (10%) 

Systems Examined With No Neoplasms Observed 

Cardiovascular System 

Endocrine System 

General Body System 

Genital System 

Integumentary System 

Musculoskeletal System 

Nervous System 

Special Senses System 

Urinary System 
Lesions in Male Mice 147 

TABLEC1 

Summary of the Incidence of Neoplasms in Male Mice in the2-Year Feed Study of Codeine (continued) 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

2-Year Study 

AlimentarySystem 

Gallbladder 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Carcinoma 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hemangiosarcoma, multiple 

Hepatoblastoma 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Histiocytic sarcoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Subcapsular,adenoma 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Pituitary gland 

Pars intermedia, carcinoma 

Thyroid gland 

Follicular cell, adenoma 

Follicular cell, adenoma, multiple 

General Body System 

None 

~~ 

Genital System 

Epididymis 

Prostate 

Seminal vesicle 

Adenoma 

Testes 

Interstitial cell, adenoma 

1 (2%) 

1 (2%) 

10 (20%) 8 (16%) 13 (26%) 7 (14%) 

3 (6%) 

12 (24%) 15 (30%) 11 (22%) 

11 (22%) 10 (20%) 5 (10%) 

(1) (3) (3) 

(49) 

(49) 
148 Codeine, NTP TR 455 

2-Year Study (continued) 

HematopoieticSystem 

Bone marrow 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Spleen 

Hemangiosarcoma 

Histiocytic sarcoma 

Thymus 

Histiocytic sarcoma 

IntegumentarySystem 

Skin 

Subcutaneous tissue, histiocytic sarcoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

None 

Nervous System 

Brain 

Respiratory System 

Lung 

Alveolar/bronchiolar adenoma 

Alveolarhronchiolar adenoma, multiple 

Alveolarhronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Special Senses System 

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Hepatocellular carcinoma, metastatic, liver 

Urinary bladder 

Systemic Lesions 

Multiple organsb 

Histiocytic sarcoma 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed 

0 PPm 

(50) 

2 (4%) 

(50) 

11 (22%) 

4 (8%) 

1 (2%) 

4 (8%) 

(50) 

(50) 

(50) 

3 (6%) 

4 (8%) 

750 ppm 

(50) 

1 (2%) 

1 (2%) 

(50) 

8 (16%) 

2 (4%) 

2 (4%) 

(50) 

1 (2%) 

1,500 ppm 

(50) 

(50) 

13 (26%) 

1 (2%) 

2 (4%) 

3 (6%) 

(49) 

1 (2%) 

3,000 ppm 

(44) 

1 (2%) 

(50) 

1 (2%) 

(50) 

7 (14%) 

1 (2%) 

2 (4%) 

2 (4%) 

(50) 

(3)

3 (100%) 

(50) 

2 (4%) 
Lesions in Male Mice 149 

TABLEC1 

Summary of the Incidence of Neoplasms in Male Mice in the2-YearFeed Study of Codeine (continued) 

0 PPm 750 ppm 

Neoplasm Summary 

Total animals with primary neoplasms' 

15-Month interim evaluation 3 1 

2-Year study 41 39 

Total primary neoplasms 

15-Month interim evaluation 4 1 

2-Year study 71 58 

Total animals with benign neoplasms 

15-Month interim evaluation 3 1 

2-Year study 34 32 

Total benign neoplasm 

15-Month interim evaluation 4 1 

2-Year study 45 42 

Total animals with malignant neoplasms 

15-Month interim evaluation 

2-Year study 20 15 

Total malignant neoplasms 

15-Month interim evaluation 

2-Year study 26 16 

Total animals with metastatic neoplasms 

2-Year study 4 2 

Total metastatic neoplasms 

2-Year study 4 3 

a ' Number of animals examined microscopically at the site and the number of animals with neoplasm 

Number of animals with any tissue examined microscopically ' Primary neoplasms: all neoplasmsexceptmetastaticneoplasms 

1,500 ppm 

2 

36 

2 

55 

2 

30 

2 

38 

15 

17 

3 

3 

3,000 ppm 

4 

28 

4 

35 

3 

21 

3 

24 

1 

10 

1 

11 

2 

2 
150 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 0 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma, multiple 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach ,

Stomach, glandular 

Tooth 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars intermedia, carcinoma 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

Tissue NOS 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Adenoma 

Testes 

+: Tissue examined microscopically 

A:Au,tolysis precludesexamination 

2445566667777777777777777 

2594702892222222222222223 

707699993999999,9999999990 

0000000000000000000000000 

3511421330011223344445550 

5059081091502594624672562 

......................... 

++AM+++MMM++++++M++MMM++M 

+M+++++++++++++++++++++++ 

......................... 

......................... 

+++++A+++++++++++++++++++ 

......................... 

+++++A+++++++++++++++++++ 

......................... 

x xx X X 

X X 

X x x X 

+ 

x x x X X 

+++++++++++M+++++++++++++ 

......................... 

......................... 

......................... + + 

......................... 

......................... 

........................... 

......................... 

......................... X X 

MMM++++MM++M+M++++++M+MM+ 

M++++++M+++++++++++++++M+ 

X 

+M+++++++++++++++++++++++ 

X 

.......................... 

+++++++M+++++++++M+++++++ 

+++++++++M+++++++M+++++.+M 

......................... 

......................... 

M:Missingtissue X: Lesionpresent 

I: Insufficienttissue Blank: Not examined 
Lesions in Male Mice 151 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma, multiple 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars intermedia, carcinoma 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

Tissue NOS 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Adenoma 

Testes 

11111111111.11111171717111 

3333333333333333333333333 

0000000000000000011111111 

0000000000000000000000000 Total 

0001122223334455601234455 Tissues/ 

3684112311211514096033838 Tumors 

+++M++++++M++++M+++++++++ 47 

++M++M+++++M+++M+M+++++M+ 34 

......................... 49 

......................... 50 

......................... 50 

......................... 49 

......................... 50 

......................... 49 

......................... 50 

X 1 

X x x X X 10 

X 3 

X X x x X X 12 

X X X x xx 11 

1 

......................... 49 

......................... 50 

......................... 50 

M++++++++++++++++++++++++ 49 

+ + ++ + + + 9 

......................... 50 

......................... 50 

+++++++++++++M+++++++++++ 49 

++++++++++.+++M+++++++++++ 49 

......................... 50 

2 

M+++++++++,+++++M+M++++M++ 36 

......................... 46 

1 

......................... 49 

1 

+ 1 

......................... 50 

......................... 48 

++++++M++++++++++++++++++ 46 

......................... 50 

X 1 

......................... 50 
152 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

~ ~ ~~ 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Hemangiosarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

~~ 

Respiratory System 

LungAlveolarhronchiolar adenoma 

Alveolarhronchiolar adenoma, 

multiple 

Alveolarlbronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

EY e Hanlerian gland 

Adenoma 

Zymbal's gland 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed 

2445566667777777777777777 

2594702892222222222222223 

7076999939999999999999990 

0000000000000000000000000 

3511421330011223344445550 

5059081091502594624672562 

......................... + ++ + 

+++MM++M++MMM+++++++M++++ 

+++M+++++++++++++++++++++ 

......................... X 

+++M+++M++++M++++++++++M+ 

MMMMMMMMMMMMMMMMMMMMMMMMM 

.......................... 

X X 

......................... 

......................... 

......................... X X X 

x x X X 

X 

x x X ......................... 

+M+++++++++++++++++++++++ 

+ 

+ 

X 

......................... 

......................... 

......................... X X 

X x 
Lesions in Male Mice 153 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Hemangiosarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Subcutaneous tissue, sarcoma 

~ ~~~ 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Respiratory System 

Lung 

Alveolarhronchiolar adenoma 

Alveoladbronchiolar adenoma, 

multiple 

Alveolarhronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

Eye

Harderian gland 

Adenoma 

Zymbal’s gland 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed 

7177777777777777777777777 

3333333333333333333333333 

0000000000000000011111111 

0000000000000000000000000 Total 

0001122223334455601234455 Tissues/ 

3684712371271514096033838 Tumors 

......................... 50 

+ + 6 

++M+++++M++++++++M+++++++ 40 

......................... 49 

......................... 50 

1 

++++++++++++M+++++++++MM+ 43 

MM+MMMMMMMMMMMMMMMMMMMM+M 2 

......................... 50 

2 ~ ~ ~~ ~~ ~ ~ 

......................... 50 

......................... 50 

......................... 50 

X x x x X xx x 11 

4 

1 

X 4 

......................... 50 

......................... 49 

1 

+ + 3 

X X 3 

+ 1 

......................... 50 

......................... 50 

......................... 50 

X 3 

X X 4 
154 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathologyof Male Mice in the 2-Year Feed Study of Codeine: 750 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatoblastoma 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

Cardiovascular Sy7tem 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Subcapsular, adenoma 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Testes 

Interstitial cell, adenoma 

03455666677’7177777777.7777 

0975602391222222222222222 

8649632376459999999999999 

1000101100000000000001111 

0688071067976678889990011 

8148102567092761363470379 

+++++++++M+++++++++++++++ 

++++++++M+++M++M++++++M++ 

......................... 

......................... 

.......................... 

......................... 

......................... 

......................... 

......................... X 

X 

X xx x xx 

xx x x X X X 

X x x 

+ ......................... 

......................... 

......................... 

......................... + + + 

......................... 

.......................... 

+++++++++I++++M++++++++++ 

+++++++++I++++”+++++++++ 

X 

......................... X 

+++++MM++M+MM+++++M+M++M+ 

......................... 

......................... X 

......................... 

......................... 

......................... 

......................... 

......................... X 
Lesions in Male Mice 155 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatoblastoma 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

~~~~~~ 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Subcapsular, adenoma 

Adrenal medulla 

Pheochromocytoma benign 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

pituitary gland 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Testes 

Interstitial cell, adenoma 

7777777777777777777777777 

3333333333333333333333333 

0000000011111111111111111 

0000000100000000001111111 Total 

6778999166777888990001112 Tissues/ 

5459589689138025124670140 Tumors 

+++++++++M+++++++++++++++ 48 

+++++M++M+++++++M+++M+M++ 41 

++++++++++++++M++++++++++ 49 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

1 

1 

X X 8 

xx xx X xx x 15 

X X xx X X X 10 

++ 3 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

+ + + + + 8 

......................... 50 

......................... 50 

......................... 48 

X 1 

......................... 47 

1 

......................... 50 

1 

+M+++M++++M+M+M+++M+++++M 35 

++++++++++++++M++++++++++ 49 

......................... 50 

X 2 

......................... 50 

......................... 50 

+++++++++++++++++++M+++++ 49 

......................... 50 

......................... 50 

1 
156 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Spleen 

Histiocytic sarcoma 

Thymus 

Integumentary System 

Mammary gland 

Skin 

Subcutaneous tissue, histiocytic 

sarcoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Skeletal muscle 

Nervous System 

Brain 

Respiratory System 

Lung 

Alveolar/bronchiolar adenoma 

Alveolarhronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Hepatocellular carcinoma, metastatic, 

liver 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignantmixed 

0345566667777777777777777 

0975602391222222222222222 

8649632376459999999999999 

1000101100000000000001111 

0688071067976678889990011 

8148102567092761363470379 

......................... + 

X 

+++++++++M+M++++M++++++++ 

......................... X 

......................... X 

M++M+M+M+MM+M+++++M+++++M 

MMMMMMMMMMMMMMMMMMMMMM+MM 

......................... 

X 

X 

......................... + 

.......................... 

......................... X x x X 

X X 

X X 

......................... 

......................... 

+ + 

x x 

......................... 

X 

......................... 

......................... X X 
Lesions 157 in Male Mice 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Spleen 

Histiocytic sarcoma 

Thymus 

Integumentary System 

Mammary gland 

Skin 

Subcutaneous tissue, histiocytic 

sarcoma 

Subcutaneous tissue, sarcoma 

MusculoskeletalSystem 

Bone 

Skeletal muscle 

Nervous System 

Brain 

RespiratorySystem 

Lung 

Alveolarhronchiolar adenoma 

Alveolarlbronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

Harderian gland 

Adenoma 

~ ~~ ~~ ~ 

Urinary System 

Kidney 

Hepatocellular carcinoma, metastatic, 

liver 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant mixed 

7777777777777777777777777 

3333333333333333333333333 

0000000011111111111111111 

0000000100000000001111111 ,Total 

6778999166777888990001112 Tissues/ 

5459589689138025124670140 Tumors 

......................... 50 

1 

1 

+++++++M++++++++M+++++++M 44 

+++++++++++M+++++++++++++ 49 

1 

......................... 50 

1 

++M+++M++++MM++++++MM++M+ 34 

MMMMMMMMMMMMMMMMMMMMMMMMM 1 

......................... 50 

1 

1 

......................... 50 

1 

.......................... 50 

......................... 50 

X x x x. 8 

2 

2 

......................... 50 

......................... 50 

+ 3 

X 3 

......................... 50 

1 

......................... 50 

......................... 50 

2 

X 1 
158 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 1,500 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

lntestine large, colon 

intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum . Intestine small, jejunum 

Carcinoma 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Thyroid gland 

Follicular cell, adenoma 

Follicular cell, adenoma, multiple 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Testes 

0.667777777777777777777777 

0361222222222222222223333 

9578799999999999999990000 

1111111111111111111111111 

2763722234444555667772333 

6647123523679027683578056 

......................... 

+M+++++M+++++M+M++M++++++ 

......................... 

......................... 

......................... 

+++A+++++++++++++++++++++ 

+++A+++++++++++++++++++++ 

X 

+++A+++++++++++++++++++++ 

......................... x xxx X xx X X 

X X X 

X X 

+ ........................... 

......................... 

......................... 

......................... ++ + 

......................... 

.......................... 

......................... X 

+++M++ ++++++++++++++++++ ......................... 

+M+M+++++++M++++++++++++M 

+M+M+++++++++++++++++++++ 

......................... 

......................... 

......................... 

++++++++++++++++++M++++++ 

......................... 

......................... 
Lesions in Male Mice 159 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 1,500 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Carcinoma 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Thyroid gland 

Follicular cell, adenoma 

Follicular cell, adenoma,multiple 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Testes 

7777777777777777777777777 

3333333333333333333333333 

0000000000011111111111111 

1111111111111111111111111 Total 

4444555666722233345566777 Tissues1 

012,5569237814714841415024 Tumors 

......................... 50 

+++++++++++++++++++M++M+M 42 

......................... 50 

++++++++++++++++++++++++M 49 

......................... 50 

......................... 49 

......................... 49 

1 

......................... 49 

......................... 50 

X X xx 13 

x xxxx x xx 11 

xx X 5 

+ + 3 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

+ + ++ 7 

......................... 50 

......................... 50 

......................... 50 

1 

......................... 48 

......................... 50 

+++++++++M++-++++++++++M+M 43 

......................... 48 

......................... 50 

X 1 

X 1 

......................... 50 

......................... 50 

M++++++++++++++++++++++++ 48 

......................... 50 

......................... 50 
160 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 1,500 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Respiratory System 

Lung

Alveolarhronchiolar adenoma 

Alveolarhronchiolar adenoma. 

multiple 

Alveolarhronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

Eye

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Lymphoma malignant mixed 

0667777777777777777777777 

0361222222222222222223333 

9578799999999999999990000 

1111111111111111111111111 

2763722234444555667772333 

6647123523679027683578056 

......................... 

+++M+++++++++++++++++++++ 

+++M+M+++++++++++++++++++ 

......................... 

+M+M++++++++++++++M+++++M 

MMMMMMMMMMMMMMMMMMM+M+MMM 

......................... 

......................... 

......................... 

.......................... X xxx X X 

X 

X 

x x 

+M+++++++++++++++++++++++ 

......................... 

+ 

+ 

X 

......................... 

......................... 

......................... 
Lesions in Male Mice 161 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 1,500 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Thymus 

Integumentary System 

Mammary gland 

Skin 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Respiratory System 

Lung 

Alveolar/bronchiolar adenoma 

Alveolarlbronchiolar adenoma, 

multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

EY e Harderian gland 

Adenoma 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Lymphoma malignant mixed 

7777777777777777777777777 

3333333333333333333333333 

0000000000011111111111111 

1111111111111111111111111 Total 

4444555666722233345566777 Tissues/ 

0125569237814714841415024 Tumors 

......................... 50 

++++++++++++++++M+++++++M 47 

......................... 48 

+++++++M+++++++++++++++++ 49 

M+M++++M+++++++++++++++M+ 42 

MMMMMMMMMMM+MMMMMMMMMMMMM 3 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

xx x x X X X 13 

1 

X 2 

X 3 

......................... 49 

......................... 50 

1 

t + + + 5 

X X X X 5 

......................... 50 

......................... 50 

......................... 50 

X 1 
Codeine, NTP TR 455 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Histiocytic sarcoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Testes 

0455667777777777777777777 

0003592222222222223333333 

9107276999999999990000000 

1212122112222222221111122 

9091822990000122338889901 

2538921696789339071685845 

......................... 

A++AM++++M+++M+++++M++MM+ 

......................... 

......................... 

A++A+++++++++++++++++++++ 

A++A+++++++++++++++++++++ 

A+++A++++++++++++++++++++ 

A++A++++++++++++M++++++++ 

......................... xx x xx 

x x X 

X 

+ 

X ......................... 

......................... 

......................... 

A++++++++++++++++++++++++ 

+ 

......................... 

......................... 

......................... 

+++M+++++++++++++++++++++ 

......................... 

+M++MM+++++++M++++++++++M 

M++M++M++++++++++++++++++ 

......................... 

......................... 

......................... 

......................... 

A++++++++++++++++++++++++ 

......................... 
Lesions in Male Mice 163 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-YearFeed Study of Codeine: 3,000 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Histiocytic sarcoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tooth 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis 

Preputial gland 

Prostate 

Seminal vesicle 

Testes 

7777777777777777777777777 

3333333333333333333333333 

0000000111111111111111111 

22222221'11111222222222222 Total 

2223334889999001111122333 Tissues/ 

0782490340147010146'946158 Tumors 

......................... 50 

........................ 40 

......................... 50 

+++++++++++++++++M+++++++ 49 

......................... 48 

......................... 48 

.......................... 48 

......................... 47 

......................... 50 

X X 7 

X X xx X X 9 

1 

1 

1 

......................... 50 

......................... 50 

......................... 50 

......................... 49 

+ 2 

......................... 50 

......................... 50 

......................... 50 

X 1 

......................... 49 

......................... 50 

++++M++MMM+++++++++++++MM 39 

+++++++++++++M+++++++++++ 46 

......................... 50 

X X 2 

......................... 50 

.......................... 50 

......................... 50 

......................... 49 

......................... 50 
164 Codeine, NTP TR 455 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 3,000 ppm (continued) 

0455667777777777717777777 

Number of Days on Study 0003592222222222223333333 

9107276999999999990000000 

1212122112222222221111122 

Carcass ID Number 9091822990000~122338889901 

2538921696789339071685845 

HematopoieticSystem 

Bone marrow ......................... Lymph node + 

Lymph node, mandibular ........................ Lymph node, mesenteric A++A+++++++++++++++++++++ 

Histiocytic sarcoma X 

Spleen ......................... Thymus +++++MM+M++++++++++++++++ 

Histiocytic sarcoma X 

IntegumentarySystem 

Mammary gland MMMMMMMMMMM+MMMMMMMMMMMMM 

Skin ......................... Subcutaneous tissue, histiocytic 

sarcoma 

Musculoskeletal System 

Bone ......................... 

Nervous System 

Brain ......................... ~ ~~~~~~~~~~~~ ~ ~ 

RespiratorySystem 

Lung ......................... Alveolarlbronchiolar adenoma X X X 

Alveolarhronchiolar adenoma, 

multiple X 

Alveolarhronchiolar carcinoma x x 

Hepatocellular carcinoma, metastatic, 

liver xx 

Nose ......................... Trachea .......................... 

Special Senses System 

E Ye +

Harderian gland ++ 

Adenoma xx 

Urinary System 

Kidney ......................... Urinary bladder +++A+++++++++++++++++++++ 

SystemicLesions 

Multiple organs ......................... Histiocytic sarcoma X 
Lesions in Male Mice 165 

TABLEC2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of Codeine: 3,000 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node. mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Spleen 

Thymus 

Histiocytic sarcoma 

Integumentary System 

Mammary gland 

Skin 

Subcutaneous tissue. histiocytic 

Sarcoma 

~~~~~~ 

Musculoskeletal System 

Bone 

Nervous System 

Brain 

Respiratory System

J-uns 

Alveolarhmnchiolar adenoma 

Alveolarhronchiolar adenoma. 

multiple 

Alveolarhronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose 

Trachea 

Special Senses System 

Eye

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

7777777777777777777777777 

3333333333333333333333333 

0000000111111111111111111 

2222222111111222222222222 Total 

2223334889999001111122333 Tissues/ 

0782490340147010146946158 

......................... + 

+MM+M++++++++++++++++++++ 

......................... 

......................... 

++++++++++++MM+M+++++++++ 

MMMMMMMMMMMMMMMMMMMMMMMMM 

......................... 

X ~ 

......................... 

......................... 

......................... 

x x xx 

......................... 

......................... 

+ 

X 

......................... 

......................... 

......................... X 

Tumors 

50 

2 

45 

48 I 

1 

50 

44 

1 

1 

50 

1 

50 

50 

50 

7 

1 

2 

2 

50 

50 

1 

3 

3 

50 

49 

50 

2 
166 Codeine, NTP TR 455 

TABLEC3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-YearFeed Study of Codeine 

0 PPm 

HarderianGland:Adenoma 

Overall ratea 3/50 (6%) 

Adjusted rateb 7.3% 

Terminal rate' 3/41 (7%) 

First incidence (days) 729 

Life table testd P=0.563N 

Logistic regression testd P=0.563N 

Cochran-Armita e testd P10.537 3 Fisher exact test 

Liver:HepatocellularAdenoma 

Overall rate 23/50 (46%) 

'Adjusted rate 53.3% 

Terminal rate 21/41 (51%) 

First incidence (days) 497 

Life table test P<0.001N 

Logistic regression test P<0.001N 

Cochran-Armitage test P<0.001N 

Fisher exact test 

Liver:HepatocellularCarcinoma 

Overall rate 13/50 (26%) 

Adjusted rate 28.2% 

Terminal rate 8/41 (20%) 

First incidence (days) 497 

Life table test P=0.121N 

Logistic regression test P=0.150N 

Cochran-Armitage test P=0.150N 

Fisher exact test 

Liver:HepatocellularAdenomaorHepatocellularCarcinoma 

Overall rate 29/50 (58%) 

Adjusted rate 63.0% 

Terminal rate 24/41 (59%) 

First incidence (days) 497 

Life table test P-0.002N 

Logistic regression test P=O.OOZN 

Cochran-Armitage test P-0.002N 

Fisher exact test 

Liver:HepatocellularCarcinoma or Hepatoblastoma 

Overall rate 13/50 (26%) 

Adjusted rate 28.2% 

Terminal rate 8/41 (20%) 

First incidence (days) 497 

Life table test P=0.108N 

Logistic regression test P=0.133N 

Cochran-Armitage test P=0.132N 

Fisher exact test 

750 ppm 

3/50 (6%) 

7.9% 

3/38 (8%) 

729 

P=O.628 

P=O.628 

P=0.661N 

25/50 (50%) 

57.8% 

20/38 (53%) 

474 

P=O.309 

P=O.402 

P=O.421 

8\50 (16%) 

18.0% 

3/38 (8%) 

396 

P=0.214N 

P=0.154N 

P=0.163N 

29/50 (58I) 

62.8% 

21/38 (55%) 

396 

P=O.447 

P=O.567 

P=0.580N 

9/50 (18%) 

19.9% 

3/38 (8%) 

396 

P=0.290N 

P=0.222N 

P=0.235N 

1,500 ppm 

5/50 (10%) 

11.1% 

5/45 (1 1 X) 

729 

P=O.408 

P=O.408 

P=O.357 

16/50 (32%) 

35.6% 

16/45 (36%) 

729 

P=0.054N 

P=0.065N 

P=O.l09N 

13/50 (26%) 

27.1% 

10/45 (22%) 

635 

P=0.480N 

P=0.589N 

P=0.590N 

23/50 (46%) 

47.9% 

20/45 (44%) 

635 

P=0.086N 

P=O.l18N 

P=O.l58N 

13/50 (2696) 

27.1% 

10145 (22%) 

635 

P=0.480N 

P=0.589N 

P=0.590N 

3,000 ppm 

3/50 (6%) 

7.0% 

3/43 (7%) 

729 

P=0.641N 

P=0.641N 

P=0.661N 

10150 (20%) 

22.0% ' 

8/43 (19%) 

500 

P=0.004N 

P=0.005N 

P=0.005N 

7/50 (14%) 

15.6% 

5/43 (12%) 

697 

P=0.099N 

P=0.105N 

P=0.105N 

16/50 (32%) 

34.0% 

12/43 (28%) 

500 

P=0.008N 

P-0.007N 

P=0.008N 

7/50 (14%) 

15.6% 

5/43 (12%) 

697 

P=0.099N 

P=0.105N 

P=0.105N 
Lesions in Male Mice 167 

TABLEC3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-YearFeed Study of Codeine (continued) 

Liver:HepatocellularAdenoma,HepatocellularCarcinoma,orHepatoblastoma 

Overall 29/50 (58%) 30/50 (60%) 23/50 (46%) 16/50 (32%) 

Adjusted 63.0% 63.6% 47.9% 34.0% 

Terminal 24/41 (59%) 21/38 (55%) 20/45 (44%) 12/43 (28%) 

396 497 635 500 

Life P=0.001N P=O.381 P=0.086N P=0.008N

P=O.487 

P=O.500 

8/50 (16%) 

20.1% 

7/38 (18%) 

474 

P=0.106N 

P=0.079N 

P=0.077N 

9/50 (18%) 

22.7% 

8/38 (21 I) 

474 

P=0.162N 

P=0.124N 

P=O.l21N 

1/50 (2%) 

2.6% 

1/38 (3 96) 

729 

P=0.043N 

P=0.029N 

P=0.030N 

32/50 (6446) 

74.2% 

27/38 (71 46) 

474 

' 

P=0.438N 

P=0.417N 

P=0.118N 

P=0.158N 

14/50 (28%) 

30.4% 

13/45 (29%) 

727 

P=0.381N 

P=0.349N 

P-0.500N 

16/50 (32%) 

34.8% 

15/45 (33%) 

727 

P=0.548N 

P=0.516N 

P=O.500 

1/50 (2%) 

2.2% 

1/45 (2%) 

729 

P=0.028N 

P=0.034N 

P=0.030N 

30750 (60%) 

63.8% 

28/45 (62%) 

635 

P=O.lllN 

P=0.160N 

P=0.266N 

P=0.007N 

P=0.008N 

8/50 (16%) 

18.0% 

7/43 (16%) 

401 

P=0.059N 

P=0.071N 

P=0.077N 

10150 (20%) 

22.5% 

9/43 (21 %) 

401 

P=0.143N 

P=0.166N 

P=0.178N 

0150 (0%) 

0.0% 

0143 (0%) -e

P=O.OlON 

P=O.OOSN 

P=0.006N 

21/50 (42%) 

45.4% 

18/43 (42%) 

401 

P=0.005N 

P=0.007N 

P=O.OOSN 

Logistic 

Cochran-Armitage 

Fisher 

Lung:AhreolarhronchiolarAdenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Lung: Alveolarhronchiolar Adenoma orCarcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

AU Organs: MalignantLymphoma(LymphocyticorMixed) 

First 

e 

sion 

Overall 

Adjusted 

rate Terminal 

First 

Life 

ssion Logistic 

Cochran-Armitage 

Fisher 

AU Organs: BenignNeoplasms 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

P=0.002N 

P=0.002N 

15/50 (30%) 

36.6% 

15/41 (37%) 

729 

P-0.089N 

P=0.119N 

P=0.132N 

15/50 (30%) 

36.6% 

15/41 (37%) 

729 

P=0.195N 

P=0.248N 

P=0.272N 

7/50 (14%) 

15.4% 

4/41 (10%) 

450 

P=0.003N 

P=0.003N 

P=0.003N 

34/50 (68%) 

77.2% 

31/41 (76%) 

497 

P=O.563 P-0.001N

P=0.002N 

P=O.WN 
168 Codeine, NTP TR 455 

TABLEC3 

Statistical Analysis of Primary Neoplasms in Male Micein the 2-YearFeed Study of Codeine (continued) 

750 ppm 

AU Organs: Malignant Neoplasms 

Overall rate 20/50 (40%) 15/50 (30%) 15/50 (30%) 10/50 (20%) 

Adjusted rate 40.05% 31.4% 31.2% 22.2% 

Terminal rate 11/41 (27%) 6/38 (16%) 12/45 (27%) 8/43 (19%) 

First incidence (days) 227 3% 635 697 

Life table test P=0.023N P=O.28ON P=O.l53N P=O.O3ON 

Logistic regression test P=0.056N P=0.169N P=0.255N P=O.O24N 

Cochran-Armitage test P=0.024N 

Fisher exact test P=O.ZOlN P=0.2OlN P-0.024N 

All Organs:Benign or Malignant Neoplasms 

Overall rate 41/50 (82%) 39/50 (78%) 36/50 (72%) 28/50 (56%) 

Adjusted rate 82.0% 81.2% 75.0% 58.3% 

Terminal rate 32/41 (7896) 29/38 (76%) 33/45 (73%) 23/43 (53%) 

First incidence (days) 227 396 635 401 

Life table test P=0.002N P=O.542 P=O.O79N P=O.OlON 

Logistic regression test P=0.003N P=0.401N P=0.162N P=0.005N 

Cochran-Armitage test P=0.001N 

Fisher exact test P=0.402N P=0.171N P=0.004N 

a Number of neoplasm-bearing animalslnumber of animals examined. Denominator is number of animals examined microscopically for liver and lung; for 

other tissues, denominator is numberof animals necropsied. 

Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminalkill 

Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to 

pairwise comparisons between the controls and that exposed group. The life table test regards neoplasmsin animals dying priorto terminal kill as being 

(directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests 

compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. 

e Not applicable;noneoplasmsinanimalgroup 
Lesions in Male Mice 169 

TABLEC4a 

Historical Incidence of Hepatocellular Neoplasms in Untreated Male B6C3Fl Micea 

Incidence in Controls 

Adenoma Carcinoma Adenoma or Carcinoma 

~~ 

Historical Incidence at Microbiological Associates, Inc. 

(2.1. Direct Blue 218 16/50 7/50 21/50 

dl-Amphetamine Sulfate lO/SO 4/50 14/50 

Overall Historical Incidence 

Total 344/1,316 (26.1%) 220/1,316 (16.7%) 509/1,316 (38.7%) 

Standard deviation 13.2% 7.2% 13.9% 

Range 4%-60% 3%-29% 10%48% 

a Dataasof 17 June 1994 

TABLEC4b 

Historical Incidence of Malignant Lymphoma in Untreated Male B6C3FlMicea 

Incidence inControls 

Historical Incidence at Microbiological Associates, Inc. 

(2.1. Direct Blue 218 2/50 

dl-Amphetamine Sulfate 4/50 

Overall Historical Incidence 

Total 117/1,324 (8.8%) 

Standard deviation 6.2% 

Range 2%-24% 

a Data as of 17 June 1994;includes data for histiocytic, lymphocytic, mixed, unspecified, or undifferentiated cell types 
170 Codeine, NTP TR 455 

TABLEC5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the2-YearFeed Study of Codeinea 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

DispositionSummary 

Animals initially in study 60 60 60 60 

15-Month interim evaldn 10 10 10 10 

Early deaths 

Accidental death 1 

Moribund 4 4 2 1 

Natural deaths 5 7 3 6 

Survivors 

Terminal sacrifice 41 38 45 43 

Animals examined microscopically 60 60 60 60 

15-Month Interim Evaluation 

Alimentary System 

Liver 

Basophilic focus 

Eosinophilic focus 

Inflammation, chronic active 2 (20%) 

Mixed cell focus 2 (20%) 

Centrilobular, cytomegaly 1 (10%) 

Centrilobular, fatty change 10 (100%) 10 (100%) 6 (60%) 

Mesentery 

Fat, necrosis 

Pancreas 

Inflammation, chronic 

Acinar cell, vacuolization cytoplasmic 

Salivary glands 

Inflammation, chronic 

Stomach, glandular 

Developmental malformation 

Tooth 

Developmental malformation 

Endocrine System 

Adrenal cortex 

Subcapsular, hyperplasia 

Islets, pancreatic 

Hyperplasia 

Pituitary gland 

Pars distalis, hyperplasia 

Thyroid gland 

Follicular cell, hyperplasia, focal 

a Number of animals examined microscopically at the site and the number of animals with lesion 
Lesions in Male Mice 171 

TABLECS 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-YearFeed Study of Codeine (continued) 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

15-Month Interim Evaluation (continued) 

Genital System 

Epididymis (10)

Inflammation, chronic active 

Preputial gland (10)

Dilatation 9 (90%)

Inflammation, chronic 6 (60%) 

Prostate (10)

Inflammation, chronic 1(10%) 

Testes (10)

Seminiferous tubule, degeneration 

Hematopoietic System 

Lymph node, mandibular 

Hyperplasia, lymphoid 

Lymph node, mesenteric 

Hyperplasia, lymphoid 

Spleen 

Hyperplasia, lymphoid 

Hyperplasia, reticulum cell 

Thymus 

Hyperplasia, lymphoid 

Integumentary System 

Skin (10)

Inflammation, chronic active 

Epidermis, hyperplasia 

Nervous System 

Brain 

Mineralization 

Respiratory System 

Lung

Congestion 

Infiltration cellular, lymphocyte 5 (50%) 

Alveolar epithelium, hyperplasia 

Nose 

Foreign body 

Olfactory epithelium, degeneration, hyaline 2 (20%) 

Respiratory epithelium, degeneration, hyaline 1(10%) 

Respiratory epithelium, hyperplasia, 

glandular 1(10%) 

Respiratory epithelium, inflammation, 

chronic active 2 (20%) 

Respiratory epithelium, metaplasia 1(10%) 
172 Codeine, NTP TR 455 

TABLEC5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the2-Year Feed Study of Codeine (continued)’ 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

15-Month Interim Evaluation(continued) 

Urinary System 

Kidney 

cyst 

Inflammation, chronic 

(10) 

3 (30%) 

2 (20%) 

(10) 

6 (60%) 

(10) 

8(80%) 

(10) 

7(70%) 

Infiltration cellular, lymphocyte 

Urinary bladder 

Nephropathy 7 (70%) 

3 (30%) 

(10) 

5 (50%) 

4 (40%) 

(10) 

3 (30%) 

3 (30%) 

(10) 

2(20%) 

1 (10%) 

(10) 

Systems Examined With No Lesions Observed 

Cardiovascular System 

General Body System 

MusculoskeletalSystem 

Special Senses System 

2-Year Study 

AlimentarySystem 

Gallbladder 

Degeneration, hyaline 

(34) 

Dilatation 

Inflammation, chronic 

Inflammation, chronic active 

Intestine large, cecum 

Hyperplasia, lymphoid 

Hyperplasia, lymphoid 

Intestine small, jejunum 

Hyperplasia, lymphoid 

Intestine small, ileum (49) 

Hyperplasia, plasma cell 1 (2%) 

Inflammation, chronic active 

Inflammation, chronic active 

Infarct 

Hematopoietic cell proliferation 

Fatty change, focal 

Eosinophilic focus 

cyst 

Clear cell focus 

Basophilic focus 

Liver (50) 

4 (8%) 

8 (16%) 

7(14%) 

3 (6%) 

2(4%) 

2 (4%) 

(50) 

4(8%) 

12 (24%) 

1 (2%) 

4 (8%) 

4 (8%) 

4 (8%) 

3 (6%) 

5 (10%) 

1 (2%) 

3 (6%) 

(50) 

3 (6%) 

4 (8%) 

1 (2%) 

2 (4%) 

2 (4%) 

Mitotic alteration 1 (2%) 1 (2%) 

Necrosis 

Mixed cell focus 

2 (4%) 

1 (2%) 

3 (6%) 

1 (2%) 1 (2%) 

Centrilobular, cytomegaly 

Pigmentation 

3 (6%) 

1 (2%) 1 (2%) 

Centrilobular, fatty change 3 (6%) 8 (16%) 

Fat, necrosis 

Artery, inflammation, chronic active 

Hemorrhage 

Mesentery (1) (3) 

3 (100%) 

(3) 

2(67%) 

1 (33%) 

1 (33%) 
Lesions in Male Mice 173 

TABLEC5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-YearFeed Study of Codeine (continued) 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

%Year Study (continued) 

Alimentary System (continued) 

Pancreas 

Inflammation, chronic 

Acinar cell, atrophy 

Artery, inflammation, chronic active 

Duct, cyst 

Salivary glands 

Inflammation, chronic 

Stomach, forestomach 

Hyperplasia 

Inflammation, chronic active 

Necrosis 

Stomach, glandular . Dilatation 

Inflammation, chronic active 

Necrosis 

Tooth 

Developmental malformation 

Cardiovascular System 

Heart 

Fibrosis 

Artery, inflammation, chronic active 

Atrium, thrombosis 

Ventricle, thrombosis 

Endocrine System 

Adrenal cortex 

Hypertrophy, focal 

Extra adrenal tissue, inflammation, 

chronic active 

Subcapsular. hyperplasia 

Islets, pancreatic 

Hyperplasia 

Pituitary gland 

Pars distalis, cyst 

Pars distalis, hyperplasia 

Thyroid gland 

Inflammation, chronic active 

C-cell, hyperplasia, focal 

Follicle, cyst 

Follicular cell, hyperplasia, focal 

(50) (50) 

(50) 

1 (2%) 

1 (2%) 

(49) (50) (49) 

(49) (50) 

23 (47%) 24 (48%) 

26 (52%) 

(50) 

1 (2%) 

(46) 

1 (2%) 

7 (14%) 34 (68%) 

General Body System 

None 
174 Codeine, NTP TR 455 

TABLEC5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the2-YearFeed Study of Codeine (continued) 

2-Yf?W shcdy (continued) 

GenitalSystem 

Epididymis 

Granuloma sperm 

Inflammation, chronic active 

Spermatocele 

Preputial gland 

Abscess 

Dilatation 

Hyperplasia 

Inflammation, chronic 

Inflammation, chronic active 

Prostate 

Hyperplasia 

Inflammation, acute 

Inflammation, chronic 

Inflammation, chronic active 

Seminal vesicle 

ArnPhY 

Dilatation 

Inflammation, chronic active 

Testes 

Seminiferous tubule, degeneration 

HematopoieticSystem 

Bone marrow 

Congestion 

Myeloid cell, hyperplasia 

Lymph node 

Hemorrhage 

Pancreatic, hyperplasia, lymphoid 

Lymph node, mandibular 

Hyperplasia, lymphoid 

Pigmentation, hemosiderin 

Lymph node, mesenteric 

Angiectasis 

Granuloma 

Hematopoietic cell proliferation 

Hemorrhage 

Hyperplasia, histiocytic 

Hyperplasia, lymphoid 

Spleen 

Hematopoietic cell proliferation 

Hyperplasia, lymphoid 

Inflammation. chronic active 

Thymus 

Atrophy 

cyst 

Ectopic parathyroid gland 

Necrosis 

0 PPm 

2 (4%) 

23 (48%) 

(46) 

11 (24%) 

1 (2%) 

(50) 

10 (20%) 

1(2%) 

(6) 

(49) 

5 (10%) 

13 (27%) 

(50) 

14 (28%) 

1(2%) 

(43) 

5 (12%) 

750 ppm 

4 (8%) 

2(4%) 

20 (40%) 

(49) 

12 (24%) 

7 (14%) 

(50) 

1(2%) 

(49) 

1 (2%) 

3(6%) 

9 (18%) 

1 (2%) 

2 (4%) 

(50) 

13 (26%) 

(34) 

2(6%) 

1(3%) 

1(3%) 

1,500 ppm 

4(8%) 

1(2%) 

(50) 

37 (74%) 

11 (23%) 

2(4%) 

(50) 

1 (2%) 

6 (12%) 

1 (2%) 

(50) 

3(6%) 

(50) 

1(2%) 

1 (2%) 

9 (19%) 

2 (4%) 

(49) 

15 (31%) 

1 (2%) 

(42) 

7 (17%) 

3 (7%) 

3,000 ppm 

(50) 

1 (2%) 

(50) 

1 (2%) 

37 (74%) 

1 (2%) 

1 (2%) 

14 (28%) 

(50) 

1 (2%) 

1 (2%) 

11 (22%) 

(49) 

1(2%) 

(50) 

1 (2%) 

(2) 

1 (50%) 

1 (50%) 

(45) 

2(4%) 

(48) 

1(2%) 

1(2%) 

10 (21%) 

(44) 

3(7%) 
Lesions in Male Mice 175 

TABLEC5 

Summary of the Incidence of Nonneoplastic Lesionsin Male Mice in the 2-YearFeed Study of Codeine (continued) 

2-Year Study (continued) 

Integumentary System 

Skin 

Foreign body 

Inflammation, chronic active 

Ulcer 

Epidermis, hyperplasia 

Prepuce, inflammation, chronic active 

Subcutaneous tissue, abscess 

Subcutaneous tissue, edema 

Subcutaneous tissue, inflammation, chronic 

active 

MusculoskeletalSystem 

Skeletal muscle 

Inflammation, chronic 

Nervous System 

Brain 

Compression 

cyst 

Mineralization 

Respiratory System 

Luw 

Congestion 

Infiitration cellular, lymphocyte 

Infiitration cellular, histiocyte 

Inflammation, chronic active 

Alveolar epithelium, hyperplasia 

Artery, inflammation, chronic active 

Bronchiole, hyperplasia 

Interstitium, inflammation, chronic active 

Pleura, inflammation, chronic 

Nose 

Exudate 

Olfactory epithelium, degeneration, hyaliie 

Respiratory epithelium, hyperplasia, 

glandular 

Respiratory epithelium, proliferation 

connective tissue 

Special Senses System 

EY e 

Cornea, inflammation, chronic active 

Harderian gland 

Inflammation, chronic active 

Zymbal’s gland 

Hypertrophy 

(50) 

1 (2%) 

2 (4%) 3 (6%) 1 (2%) 

1 (2%) 

1 (2%) 1 (2%) 4 (8%) 

1 (2%) 

1 (2%) 

(1) 

1 (100%) 

10 (20%) 

2 (4%) 

1 (2%) 

1 (2%) 

(49) 

24 (49%) 

1 (2%) 

(1) 

(3) (3) 

(1) 

1 (100%) 
176 Codeine, N'I" TR 455 

TABLEC5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the2-YearFeed Study of Codeine (continued) 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

2-Year Study (continued) 

Urinary System 

Kidney 

cyst 

Hydronephrosis 

Infarct 

Inflammation, chronic 

Inflammation, chronic active 

Metaplasia, osseous 

Nephropathy 

Pigmentation 

Artery, inflammation, chronic active 

Bilateral, hydronephrosis 

Capsule, fibrosis 

Renal tubule, degeneration, hyaline 

Renal tubule, hyperplasia 

Renal tubule, necrosis 

Renal tubule, pigmentation 

Urinary bladder 

Dilatation 

Infiltration cellular, lymphocyte 

dammation, acute 

Inflammation, chronic active 

1 (2%) 

2 (4%) 

(50) 

2 (4%) 

10 (20%) 

3 (6%) 

43 (86%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

(50) 

2 (4%) 

4 (8%) 

3 (6%) 

1 (2%) 

1 (2%) 

41 (82%) 

1 (2%) 

(50) 

2 (4%) 

6 (12%) 

1 (2%) 

1 (2%) 

1 (2%) 

7 (14%) 

37 (74%) 

(49) 

3 (6%) 

4 (8%) 

1(2%) 
177 

APPENDIX D 

SUMMARY OF LESIONS IN FEMALE MICE 

IN THE 2-YEAR FEED STUDY 

OF CODEINE 

TABLEDl Summary of the Incidence of Neoplasms in Female Mice 

in the 2-Year Feed Study of Codeine ................................. 179 

TABLE D2 Individual Animal Tumor Pathology of Female Mice 

in the 2-Year Feed Study of Codeine ................................. 184 

TABLE D3 Statistical Analysis of Primary Neoplasms in Female Mice 

in the 2-Year Feed Study of Codeine ................................. 204 

TABLED4 Historical Incidence of Hepatocellular Neoplasms 

in Untreated Female B6C3Fl Mice ................................... 208 

TABLE D5 Summary of the Incidence of NoMeoplastic Lesions in Female Mice 

in the 2-Year Feed Study of Codeine ................................. 209 
Lesions in Female Mice 179 

TABLEDl 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Feed Study of Codeinea 

0 PPm 

Disposition Summary 

Animals initially in study 60 

IS-Month interim evaluation 10 

Early deaths 

Moribund 7 

Natural deaths 7 

Survivors 

Died last week of study 1 

Terminal sacrifice 35 

Missing 

Anis examined microscopically 60 

15-Month Interim Evaluation 

Alimentary System 

Intestine large, cecum (10) 

Intestine small, jejunum (10) 

Liver (10) 

Hepatocellular carcinoma 

Hepatocellular adenoma 2 (20%) 

Endocrine System 

Thyroid gland 

Follicular cell, adenoma 

Respiratory System 

Lung (10)

Alveolar/bronchiolar adenoma 2 (20%) 

Systemic Lesions 

Multiple organsb 

Lymphoma malignant mixed 

Systems Examined With No Neoplasms Observed 

Cardiovascular System 

General Body System 

Genital System 

Hematopoietic System 

Integumentary System 

Musculoskeletal System 

Nervous System 

Special Senses System 

Urinary System 

750 ppm 

60 

9 

9 

6 

36 

60 

(9) 

(9) 

(9) 

1(11%) 

1 (11%) 

(9) 

1 (11%) 

1,500 ppm 

60 

9 

6 

2 

43 

60 

(9) 

(9) 

(9) 

(9) 

3,000 ppm 

60 

9 

4 

11 

1 

34 

1 

59 
180 Codeine, NTP. TR 455 

TABLEDl 

Summary of the Incidence of Neoplasms in Female Mice in the 2-YearFeed Study of Codeine (continued) 

2-Year Smdy 

Alimentary 'System 

Intestine large, rectum 

Adenocarcinoma, metastatic, pancreas 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Histiocytic sarcoma 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatocellular Carcinoma 

' Hepatocellular carcinoma,' multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Adenocarcinoma, metastatic, pancreas 

Fibrosarcoma 

Hemangioma 

Pancreas 

Histiocytic sarcoma 

Acinar cell, adenocarcinoma 

Salivary glands 

Stomach, forestomach 

Squamous cell carcinoma 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Osteosarcoma,metastatic, skin 

EndocrineSystem 

Adrenal cortex 

Adenoma 

Extra adrenal tissue, adenocarcinoma, 

metastatic, pancreas 

Islets, pancreatic 

Adenoma 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Pars intermedia, adenoma 

Thyroid gland 

Histiocytic sarcoma 

Follicular cell, adenoma 

General Body System 

None 

0 PPm 

5 (10%) 

7 (14%) 

5 (10%) 

1 (2%) 

(9) 

(50) 

(50) 

750 ppm 1,500 ppm 3,000 ppm 

6 (12%) 3 (6%) 

1 (2%) 

9 (18%) 11 (22%) 4 (8%) 

1 (2%) I 2 (4%) 1 (2%)

2 (4%) 1 (2%) 

(9) (5) 

1 (20%) 

1 (11%) 

1 (11%) 

(51) 

(50) 

1 (2%) 

1 (2%) 

(50) 

4 (8%) 
Lesions in Female Mice 181 

TABLEDl 

Summary of the Incidence of Neoplasms in Female Mice in the 2-YearFeed Study of Codeine (continued) 

750 ppm 1,500 ppm 3,000 ppm 

2-Year Stud’ (continued) 

Genital System 

Ovary (50)

Adenocarcinoma, metastatic, pancreas 1 (2%) 

Adenoma 2 (4%) 

Cystadenoma 1 (2%) 

Granulosa cell tumorbenign 

Histiocytic sarcoma 

Teratoma maliinant 

UkNS 

Adenocarcinoma, metastatic, pancreas 

Hemangioma 

Hemangiosarcoma 

Histiocytic sarcoma 1 (2%) 

Leiomyosarcoma 

Polyp stromal 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Hematopoietic System 

Bone marrow 

Histiocytic sarcoma 

Lymph node 

Histiocytic sarcoma 

Teratoma malignant, metastatic 1 (11%) 

Mediastinal, histiocytic sarcoma 1 (11%) 

Lymph node, mandibular (49) (49)

Histiocytic saxom 1 (2%) 

Lymph node, mesenteric (49) (47) (48)

Adenocarcinoma, metastatic, pancreas 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Spleen 

Hemangiosarcoma 

Histiocytic sarcoma 1 (2%) 

Thymus (41) 

IntegumentarySystem 

Mammary gland (45)

Carcinoma 

Skin (50)

Subcutaneoustissue, fibrosarcoma 

Subcutaneous tissue, hemangiosarcoma 1 (2%)

Subcutaneous tissue, liposarcoma 

Subcutaneous tissue, osteosarcoma 

Subcutaneous tissue, sarcoma 1 (2%) 
182 

2-Year Study (continued) 

Musculoskeletal System 

Bone 

Chondrosarcoma 

Osteosarcoma 

Skeletal muscle 

Adenocarcinoma, metastatic, pancreas 

Hemangiosarcoma 

Nervous System 

None 

Respiratory System 

Lung

Adenocarcinoma, metastatic, pancreas 

Alveolarhronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolarhronchiolar carcinoma 

Carcinoma, metastatic, harderian gland 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Osteosarcoma,metastatic, skin 

Mediastinum, osteosarcoma, metastatic, skin 

Nose 

Trachea 

Special Senses System 

Harderian gland 

Adenoma 

Carcinoma 

Bilateral. adenoma 

Urinary System 

Kidney 

Adenocarcinoma, metastatic, pancreas 

Histiocytic sarcoma 

Squamous cell carcinoma, metastatic, 

uncerrain primary site 

Urinary bladder 

Adenocarcinoma, metastatic, pancreas 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed 

0 PPm 

(50) 

(1) 

(50) 

1 (2%) 

1 .(2%) 

1 (2%) 

(50) 

(50) 

(2) 

1 (50%) 

(50) 

(50) 

(50) 

2 (4%) 

2 (4%)

5 (10%) 

Codeine, NTP TR 455 

750 ppm 1,500 ppm 3,000 ppm 

(4) (3) (3) 

2 (50%) 1 (33%) 2 (67%) 

1 (25%) 

1 (25%) 

(51) (51) (50) 

2 (4%) 1 (2%) 

1 (2%) 

4 (8%) 6 (12%) 8 (16%) 
Lesions in Female Mice 183 

TABLEDl 

Summary of the Incidence of Neoplasms in Female Mice in the2-YearFeed Study of Codeine (continued) 

750 ppm 

Total animals with primary neoplasms' 

Neoplasm Summary 

15-Month interim evaluation 4 3 

2-Year study 34 36 

Total primary neoplasms 

15-Month interim evaluation 6 3 

2-Year study 

Total animals with benign neoplasms 

51 60 

15-Month interim evaluation 4 2 

2-Year study 26 25 

Total benign neoplasms 

15-Month interim evaluation 5 2 

2-Year study 34 35 

Total animals with malignant neoplasms 

15-Month interim evaluation 1 1 

2-Year study 15 23 

Total malignant neoplasms 

15-Month interim evaluation 1 1 

2-Year study 17 25 

Total animals with metastatic neoplasms 

2-Year study 3 

Total metastatic neoplasms 

2-Year study 7 

Total animals with malignant neoplasms 

of uncertain primary site 

2-Year study 1 

a Number'of animals examined microscopically at the site and the numberof animals with neoplasm 

Number of animals with any tissue examined microscopically ' primary neoplasms:allneoplasmsexceptmetastaticneoplasms 

1,500 ppm 3,000 ppm 

38 31 

69 48 

33 19 

48 27 

17 19 

21 21 

5 

14 
184 Codeine, NTP TR 455 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

~~~~~ ~ ~ ~ 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

General Body System 

None 

Genital System 

Clitoral gland 

Ovary 

Adenoma 

Granulosa cell tumor benign 

Oviduct 

UkNS 

Histiocytic sarcoma 

Polyp stromal 

+: Tissue examined microscopically 

A: Autolysisprecludesexamination 

0245555556667777777777777 

0793467895591233333333333 

9270450211953212444444444 

222222222222-2222222222222 

5995678858544578444466789 

7300658395193686256849041 

M+++++M++++++++++++++++++ 

++AM+++++++++M+++++++++++ 

......................... 

......................... 

......................... 

++A++++++M+++++++++++++++ 

......................... 

++A+++++++++++++++M++++++ 

......................... xx 

X xx 

X 

X 

+ + + + + ......................... 

......................... 

......................... 

.......................... 

......................... 

......................... 

......................... 

++++++++++I++++++++++++++ 

......................... X 

++M+M+M++M++M+M+++++M++++ 

M++M+++++++++++++++++++++ 

xxx x x X 

X 

++++++++++++++M+++f++++++ 

++M++++++++MM++MMM++++M++ 

+++++++++++M+++++++++++++ 

+ ++ ......................... X 

X X 

M: Missingtissue X: Lesionpresent 

I: hfficient.tissue Blank: Not examined 
Lesions in Female Mice 185 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Islets,pancreatic . 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

General Body System 

None 

GenitalSystem 

Clitoral gland 

Ovary 

Ademma 

Granulosa cell tumor benign 

Oviduct 

uterus 

Histiocytic sarcoma 

Polyp stromal 

7777777771777711777777777 

3333333333333333333'333333 

5555555555555556666666666 

2222222222222222222222223 Total 

4556777788899994556668890 Tissues/ 

4587146912767891231280940 Tumors 

......................... 48 

+++MM+++++++++M+++M++++++ 43 

......................... 50 

......................... 50 

......................... 50 

......................... 48 

......................... 50 

......................... 48 

......................... 50 

X xx 5 

X X xx 7 

X X X X 5 

1 

+ + + + 9 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

......................... 50 

.......................... 50 

......................... 50 

......................... 49 

......................... 50 

1 

+MM+++M++M+++++++++M+M++M 36 

......................... 48 

X X X xx 11 

1 

+++++++++++++++++++++I+++ 48 

........................ 41 

......................... 49 

X X 2 

X 1 

3 

......................... 50 

1 

xx 4 
Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Histiocytic sarcoma 

Lymph node 

Histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Spleen 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Subcutaneous tissue, hemangiosarcoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Skeletal muscle 

Nervous System 

Brain 

Respiratory System 

Lung 

Alveolarhronchiolar adenoma 

Alveolarhronchiolar carcinoma 

Histiocytic sarcoma 

Nose 

Trachea 

Special Senses System 

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Urinary bladder 

0245555556667777777777777 

0793467895591233333333333 

9270450211953212444444444 

2222222222222222222222222 

5995678858544578444466789 

7300658395193686256849041 

......................... X 

+ ++ + 

X 

M++++.++M+++++++++++++++++ 

......................... X ......................... X 

M++MM+++++IM+M++M++M+++M+ 

+++++++++++++++++++++++M+ 

......................... X 

X 

......................... + ~~~ ~ ~ 

......................... 

......................... X 

X ......................... 

.......................... ~ 

+ 

......................... 

......................... 

SystemicLesions 

Multiple organs ......................... Histiocytic sarcoma X X 

Lymphoma malignant lymphocytic X X 

Lymphoma malignantmixed x xx 
Lesions in Female Mice 187 

TAB- D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of Codeine: 0 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

HematopoieticSystem 

Bone marrow 

Histiocytic sarcoma 

Lymph node 

Histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Spleen 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Subcutaneous tissue, hemangiosarcoma 

Subcutaneous tissue, sarcoma 

MusculoskeletalSystem 

Bone 

Skeletal muscle 

Nervous System 

Brain 

Respiratory System 

Lung

Alveolarhronchiolar adenoma 

Alveolarhronchiolar carcinoma 

Histiocytic sarcoma 

Nose 

Trachea 

Special Senses System 

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant lymphocytic 

Lymphoma malignantmixed 

7777777777777777777777777 

3333333333333333333333333 

5555555555555556666666666 

2222222222222222222222223 Total 

4556777788899994556668890' Tissues1 

4587146912767891231280940 Tumors 

......................... 50 

1 

4 

1 

+M+++++++++++++M++++++++M 45 

+++++++++++++++I+++++++++ 49 

1 

......................... 50 

1 

M+++++++++++++M++M+++++++ 38 

++M+++++++++MM+++++M+++++ 45 

......................... 50 

1 

1 

......................... 50 

1 

......................... 50 

......................... 50 

1 

X 1 

1 

.......................... 50 

......................... 50 

+ 2 

X 1 

......................... 50 

......................... 50 

......................... 50 

2 

2 

X X 5 
188 Codeine, NTP TR 455 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Fibrosarcoma 

Hemangioma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Osteosarcoma, metastatic, skin 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

Tissue NOS 

Genital System 

Clitoral gland 

Ovary 

Adenoma 

Histiocytic sarcoma 

Oviduct 

00333455,66666667777777777 

0045871312346773333333333 

8889892965374271444444444 

3333333333333333333333333 

4453424335224120000111223 

6804335292572184128267241 

M++++++++++++++++++++++++ 

++A+++++M++M++A++M++M++++ 

......................... 

......................... 

M+++++++++++++A++++++++++ 

A+++++++++++++A++++++++++ 

A++++++++++++A++M++++++++ 

A+A+++++++++++A++++++++++ 

......................... X X X 

X xx X 

X 

X X 

+ + 

......................... 

......................... 

......................... 

........................... 

......................... 

......................... X 

......................... 

......................... 

......................... 

M++M+++M++M++M+++M++M+++M 

.M++++++++++++++++++++++++ 

X X X 

X ......................... X 

+ 

MM+++++M++++M++++.+++++M++ 

......................... xx 

X 

+ 
Lesions in Female Mice 189 

TABLED2 

Individual Animal Tumor Pathologyof Female Mice in the 2-Year Feed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Fibrosarcoma 

Hemangioma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Osteosarcoma, metastatic, skin 

Endocrine System 

Adrenal cortex 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

Tissue NOS 

Genital System 

Clitoral gland 

Ovary 

Adenoma 

Histiocytic sarcoma 

Oviduct 

77777777777777777777777777 

33333333333333333333333333 

44444444555555555555555566 

33333333333333333333333333 Total 

33344455000011111222555624 Tissues/ 

56707914356903489169789004 Tumors 

.......................... 

+++M+++++++++++++M++M+++++ 

.......................... 

.......................... 

.......................... 

........................... 

.......................... 

.......................... 

.......................... X X X 

X X X x x 

++ + +++ + 

X 

X .......................... 

+M++++++++++++++++++++++++ 

.......................... 

.......................... 

.......................... 

.......................... 

.......................... 

M+++++++++++++++++++++++++ 

.......................... X X 

MM+M++++++++++++++++++++M+ 

++++++++++ +++++++++M+++M+ 

x x X X X X 

.......................... X xx 

~~ 

+M++++++++++++++M++++++M++ 

.......................... X 

50 

42 

51 

51 

49 

49 

48 

48 

51 

6 

9 

1 

2 

9 

1 

1 

51 

50 

51 

51 

51 

51 

1 

51 

50 

51 

2 

39 

47 

9 

1 

51 

4 

1 

43 

51 

3 

1 

1 
190 Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Genital System (continued) 

Uterus 

Hemangiosarcoma 

Histiocytic sarcoma 

Leiomyosarcoma 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

HematopoieticSystem 

Blood 

Bone marrow 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Histiocytic sarcoma 

Lymph node, mesenteric 

Histiocytic sarcoma .

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Spleen 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Subcutaneous tissue, fibrosarcoma 

Subcutaneous tissue, liposarcoma 

Subcutaneous tissue, osteosarcoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Osteosarcoma 

Nervous System 

Brain 

Respiratory System 

~~ 

Lung 

Alveolarhronchiolar adenoma 

Alveoladbronchiolar carcinoma 

Carcinoma, metastatic, harderian 

gland 

Histiocytic sarcoma 

Osteosarcoma, metastatic, skin 

Mediastinum, osteosarcoma, 

metastatic, skin 

Nose 

Trachea 

0033345,566666667777777777 

0045871312346773333333333 

8889892965374271444444444 

3333333333333333333333333 

4453424335224120000111223 

6804335292572184128267241 

......................... X 

X 

......................... + + 

X 

MM++M++++++++++++++++M+++ 

X ........................ X X 

......................... X 

+++++++++++++M+++MM+++MMM 

......................... 

......................... X X 

X 

X 

X 

......................... 

......................... 

~ 

......................... X X 

X 

X X 

X 

X ......................... 

......................... 
Lesions in Female Mice 191 

TABLE D2 

Individual Animal Tumor Pathologyof Female Mice in the 2-Year Feed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Genital System (continued) 

uterus 

Hemangiosarcoma 

Histiocytic sarcoma 

Leiomyosarcoma 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

HematopoieticSystem 

Blood 

Bone marrow 

Lymph node 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Histiocytic sarcoma 

Lymph node, mesenteric 

Histiocytic sarcoma 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Spleen 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Skin 

Subcutaneous tissue, fibrosarcoma 

Subcutaneous tissue, liposarcoma 

Subcutaneous tissue, osteosarcoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Osteosarcoma 

NeNOUSSystem 

Brain 

Respiratory System 

Lung 

Alveolarhronchiolar adenoma 

Alveolarhronchiolar carcinoma 

Carcinoma, metastatic, harderian 

gland 

Histiocytic sarcoma 

Osteosarcoma, metastatic, skin 

Mediastinum, osteosarcoma, 

metastatic, skin 

Nose 

Trachea 

17711717177171171717177117 

33333333333333333333333333 

44444444555555555555555566 

33333333333333333333333333 Total 

33344455000011111222555624 Tissues/ 

56101914356903489169189004 Tumors 

.......................... 51 

X 

X 1 

+ 1 

.......................... 51 

2 

1 

+M++++++++++++++++++++++++ 46 

1 

+M++++++++++++++++++++++++ 48 

2 

X 1 

.......................... 51 

1 

+M+++++M++++++++++M++++++M 41 

.......................... 51 

.......................... 51 

X 

.......................... 51 

X 1 

.......................... 51 

.......................... 51 

2 

X 1 

1 

2 

1 

1 

.......................... 51 

.......................... 51 
192 Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Special Senses System 

EY e Harderian gland 

Adenoma 

Carcinoma 

Bilateral, adenoma 

Urinary System 

Kidney 

Histiocytic sarcoma 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed 

0033345566666667777777777 

0045871312346773333333333 

8889892965374271444444444 

3333333333333333333333333 

4453424335224120000111223, 

680433529'2572184128267241 

+ 

+ + 

X 

X 

......................... X 

M++++++++++++++++++++++++ 

......................... X X 

X 

X 
Lesions in Female Mice 193 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 750 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Special Senses System 

EY e Harderian gland 

Adenoma 

Carcinoma 

Bilateral, adenoma 

Urinary System 

Kidney 

Histiocytic sarcoma 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Urinary bladder 

SystemicLesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed 

1111,1111111111111111111111 

33333333333333333333333333 

44444444555555555555555566 

33333333333333333333'333333 Total 

33344455000011111222555624 Tissues/ 

56101914356903489169189004 Tumors 

1 

+ + 4 

X X 2 

1 

1 

.......................... 51 

1 

X 1 

.......................... 50 

.......................... 51 

2 

1 

xxx 4 
194 Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 1,500 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Pars intermedia. adenoma 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland 

Ovary 

Utems 

Hemangioma 

Polyp stromal 

0336777777777777777777777 

0224022233333333333333333 

8393347714444444444444455 

4333333443333333334444433 

0998989016677788880011166 

2066732521534824590316848 

+++++++++M+++++++++++++++ 

++++++++++++++++++++++M++ 

......................... 

......................... 

......................... 

A +++++A+++++++++++++++++ 

+ ....................... A +M+++++++++++++++++++++ 

......................... 

X X 

X x x 

+ + + ......................... 

......................... 

......................... 

......................... 

......................... 

......................... 

+++++M+++++++++++++++++++ 

X 

+++++M+++++++++++++++++++ 

......................... X 

M+++M+++++++++++++M+M+++M 

M++++++++++++++++++++++++ 

X X x x x x xx x x 

X 

X X 

......................... X 

+++++M++++++++++++++M++++ 

++M++++++++++++++++++++++ 

++M++++++++++++++++++++++ 

X 

X 
Lesions in Female Mice 195 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of Codeine: 1,500 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

AlimentarySystem 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Adenoma 

Adrenal medulla 

Islets, pancreatic 

Adenoma 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars distalis, carcinoma 

Pars intermedia, adenoma 

Thyroid gland 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland 

Ovary 

u@NS 

Hemangioma 

Polyp stromal 

11111117111171111111111111 

33333333333333333333333333 

55555555555555555666666666 

33333333334444444333333444 Total 

71188899990000011666719112 Tissues/ 

12101113594618931369598050 Tumors 

++++++++++++++++++M+++++++ 49 

++++M+++M++++++++++++M++++ 47 

.......................... 51 

++++++++++++M+++++++++++++ 50 

.......................... 51 

.......................... 48 

.......................... 50 

.......................... 48 

.......................... 51 

X 1 

X X X X 6 

xxx X xx X' X 11 

X X 2 

+ ++ + I 

.......................... 51 

.......................... 51 

.......................... 51 

........................... 51 

+++++++++++++M++++++++++++ 50 

.......................... 51 

.......................... 50 

1 

.......................... 50 

.......................... 51 

1 

+M+++M++MM++++M+++M+++++++ 40 

+++++++++++++++++M++++++++ 49 

X X X X xxx 11 

1 

X 3 

.......................... 51 

X X 3 

+++++++M+,++++++++M+M+M++++ 45 

.......................... 50 

.......................... 50 

1 

X 2 
196 Codeine, NTP TR 455 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Hemangiosarcoma 

Thymus 

~ ~ ~~-

Integumentary System 

Mammary gland 

Carcinoma 

Skin 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Chondrosarcoma 

Skeletal muscle 

Hemangiosarcoma 

Nervous System 

Brain 

spinal cord 

Respiratory System 

Lung 

keolarhronchiolar adenoma 

Alveolarhronchiolar adenoma, 

multiple 

Alveolarhronchiolar carcinoma 

Nose 

Trachea 

Special Senses System 

Ear 

EY e Harderian gland 

Adenoma 

Urinary System 

Kidney 

Ureter 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Lymphoma malignantmixed 

033677777777777777'7777777 

0224022233333333333333333 

8393347714444444444444455 

4333333443333333334444433 

0998989016677788880011166 

2066732521534824590316848 

......................... + + ......................... 

M+M+++++M++++++++M+++++++ 

......................... X 

+++MM+++++++++++++++++++M ~ ~ ~ ~~ ~~ ~ ~ ~ 

......................... X X ......................... X 

......................... X 

+ 

X 

......................... + 

......................... X X 

X 

......................... 

......................... 

+ 

+ 

+ 

......................... 

......................... 

......................... xxxx. X 
Lesions in Female Mice 197 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 1,500 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Hematopoietic System 

Bone mamw 

Lymph node 

Lymph node, mandibular 

Lymph node, mesenteric 

Spleen 

Hemangiosarcoma 

Thymus 

Integumentary System 

Mammary gland

Carcinoma 

Sb 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Chondrosarcoma 

Skeletal muscle 

Hemangiosarcoma 

Nervous System 

Brain 

spinal cord 

Respiratory System 

Lung

Alveolarlbronchiolar adenoma 

Alveolarlbronchioly adenoma, 

multiple 

Alveolarhronchiolar carcinoma 

Nose 

Trachea 

Special Senses System 

Ear 

Eye

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Ureter 

Urinary bladder 

Systemic Lesions 

Multiple organs 

Lymphoma malignant mixed 

77777777777777777777777777 

33333333333333333333333333 

55555555555555555666666666 

33333333334444444333333444 Total 

77788899990000011666779112 Tissues/ 

12701713594678937369598050 Tumors 

.......................... 51 

2 

+++M++++++++++M+++++++++++ 49 

.......................... 47 

.......................... 51 

1 

+++M++++++++++++++++++++++ 47 

M+++++++++++++++++++++++++ 50 

2 

.......................... 51 

1 

.......................... 51 

.......................... 51 

1 

.......................... 51 

x x X 5 

X 1 

1 

+++++++++++++++++M++++++++ 50 

.......................... 51 

+ 

+ + 

X 

.......................... 51 

+ 1 

........................... 51 

.......................... 51 

X 6 
198 Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 3,000 ppm 

Number of Days on Study 

Carcass ID Number 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Adenocarcinoma, metastatic, pancreas 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Histiocytic sarcoma 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Adenocarcinoma, metastatic, pancreas 

Pancreas 

Histiocytic sarcoma 

Acinar cell, adenocarcinoma 

Salivary glands 

Stomach,forestomach 

Squamous cell carcinoma 

Stomach, glandular 

Cardiovascular System 

Blood vessel 

Heart 

Endocrine System 

Adrenal cortex 

Extra adrenal tissue, adenocarcinoma, 

metastatic, pancreas 

Adrenal medulla 

Islets, pancreatic 

Parathyroid gland 

Pituitary gland 

Pars distalis, adenoma 

Pars intermedia, adenoma 

Thyroid gland 

Histiocytic sarcoma 

Follicular cell, adenoma 

General Body System 

None 

003555666777777777777777 

009059138112222333333333 

887668910110278144444444 

444444444444444444444444 

576323242235655633334444 

826480443191342735681369 

........................ 

A+++M++++M++MM+++M+++M++ 

........................ 

+++++++M++++++++++++++++ 

X , . 

M+++++++++++++++++++++++ 

A+++++++++++A+++++++++++ 

........................ X 

A+++++++++++A+++++++++++ 

........................ xx X 

X 

X 

X A 

+ ++ 

X ......................... A

v 

X 

........................ 

........................ 

A+++++++++++++++++++++++ 

+++++++++M++++++++++++++ 

......................... 

........................ 

X 

+++++++++M++++++++++++++ 

........................ 

+M+M+MMMMMM+++++++++MM++ 

M++++++M++++++++++++++++ 

x x X 

X ........................ X 
Lesions in Female Mice 199 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 3,000 ppm (continued) 

~~~ ~~ ~~ ~~~~~~~~~~~~~~~~~~ 

17117117171717771777777177 

Number of Days on Study 33333333333333333333333333 

44444444444455555555556666 

44444444444444444444444444 Total 

Carcass ID Number 55666171777722344555672246 Tissues1 

05129014678959128369832604 Tumors 

Alimentary System 

Esophagus ++M++++++++++++++++M++++++ 48 

Gallbladder ++M++++++++++++++++++++M++ 41 

Intestine large, colon .......................... 50 

Intestine large, rectum .......................... 49 

Adenocarcinoma, metastatic, pancreas 1 

Intestine large, cecum .......................... 49 

Intestine small, duodenum .......................... 48 

Intestine small, jejunum .......................... 50 

Histiocytic sarcoma 1 

Intestine small, ileum .......................... 48 

Liver .......................... 50 

Hepatocellular carcinoma 3 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma xx x 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery + + 

Adenocarcinoma, metastatic, pancreas 

Pancreas .......................... 50 

Histiocytic sarcoma 1 

Acinar cell, adenocarcinoma 1 

Salivary glands .......................... 50 

Stomach, forestomach .......................... 50 

Squamous cell carcinoma X 1 

Stomach, glandular .......................... 49 

Cardiovascular System 

Blood vessel .......................... 49 

Heart .......................... 50 

Endocrine System 

Adrenal cortex .......................... 50 

Extra adrenal tissue, adenocarcinoma, 

metastatic, pancreas 1 

Adrenal medulla +++++++++++M++++++++++++++ 48 

Islets, pancreatic .......................... 50 

Parathyroid gland +++++++M++++++++M+++MMMMM+ 33 

Pituitary gland +++M+++++++++++++M++++++++ 46 

Pars distalis, adenoma xx x X X X X 10 

Pars intermedia, adenoma 1 

Thyroid gland .......................... 50 

Histiocytic sarcoma 1 

Follicular cell, adenoma X X 2 

General Body System 

None 
200 Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 3,OOO ppm (continued) 

003555666777777777777777 

Number of Days on Study ' 0090'5913811222233333333,3 

887668910110278144444444 

444444444444444444444444 

Carcass ID Number 576323242235655633334444 

8264804431913427356813'69 

Genital System 

Clitoral gland ++MM++M++++M+M+M++++++++ 

Ovary ........................ Adenocarcinoma, metastatic, pancreas X 

Adenoma 

Cystadenoma X 

Histiocytic sarcoma X 

Teratoma malignant X 

Oviduct + 

UeNS ........................ Adenocarcinoma, metastatic, pancreas X 

Histiocytic smoma X 

Vagina + 

Hematopoietic System 

Bone marrow ........................ Lymph node + ++ + + + 

Teratoma malignant, metastatic X 

Mediastinal, histiocytic sarcoma X 

Lymph node, mandibular +++++++M++++++++++++++++ 

Histiocytic sarcoma X 

Lymph node, mesenteric M++M++++++++++++++++++++ 

Adenocarcinoma, metastatic, pancreas X 

Histiocytic sarcoma X 

Spleen ........................ Histiocytic sarcoma X 

Thymus +++++M+++MM+MM+M++++++++ 

Integumentary System 

Mammary gland ........................ Carcinoma 

Skin ........................ Subcutaneous tissue, hemangiosarcoma X 

Subcutaneous tissue,sarcoma X 

~~ 

Musculoskeletal System 

Bone ........................ Skeletal muscle + 

Adenocarcinoma, metastatic, pancreas X 

Nervous System 

Brain ........................ 

Respiratory System 

Lung ........................ 

Adenocarcinoma, metastatic, pancreas X 

Alveolarhronchiolar adenoma X X 

Hepatocellular carcinoma, metastatic, 

liver X X X 

Histiocytic sarcoma X 

Nose ........................ Trachea ........................ 
Lesions in Female Mice 201 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of Codeine: 3,000 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Genital System 

Clitoral gland 

Ovary 

Adenocarcinoma, metastatic, pancreas 

Adenoma 

Cystadenoma 

Histiocytic sarcoma 

Teratoma malignant 

Oviduct 

uterus 

Adenocarcinoma, metastatic, pancreas 

Histiocytic sarcoma 

Vagina 

Hematopoietic System 

Bone marrow 

Lymph node 

Teratoma malignant, metastatic 

Mediastinal, histiocytic sarcoma 

Lymph node, mandibular 

Histiocytic sarcoma 

Lymph node, mesenteric 

Adenocarcinoma, metastatic, pancreas 

Histiocytic sarcoma 

Spleen ' 

Histiocytic sarcoma 

Thymus 

IntegumentarySystem 

Mammary gland 

Carcinoma 

Skin 

Subcutaneous tissue, hemangiosarcoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone 

Skeletal muscle 

Adenocarcinoma, metastatic, pancreas 

Nervous System 

Brain 

Respiratory System 

Lung 

Adenocarcinoma, metastatic, pancreas 

Alveolarhronchiolar adenoma 

Hepatocellular carcinoma, metastatic, 

liver 

Histiocytic sarcoma 

Nose 

Trachea 

11711711711117117117111171 

3333333,3333333333333333333 

44444444444455555555556666 

44444444444444444444444444 Total 

55666711111722344555612246 Tissues1 

05129014618959128369832604'Tumors 

........................... 44 

.......................... 50 

1 

X X 2 

1 

1 

1 

1 

.......................... 50 

1 

1 

1 

.......................... 50 

+ + + 9 

1 

1 

.......................... 49 

1 

.......................... 48 

1 

1 

.......................... 50 

1 

......................... 42 

.......................... 50 

xx 2 

.......................... 50 

1 

X 2 

.......................... 50 

1 

1 

.......................... 50 

.......................... 50 

1 

X X 4 

3 

1 

.......................... 50 

.......................... 50 
202 Codeine, NTP TR 455 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-YearFeed Study of Codeine: 3,000 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Special Senses System 

EY e Harderian gland 

Adenoma ~~ - ~~ ~ ~ 

Urinary System 

Kidney 

Adenocarcinoma, metastatic, pancreas 

Histiocytic sarcoma 

Urinary bladder 

Adenocarcinoma, metastatic, pancreas 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant mixed 

003555666777777777777777 

009059138112222333333333 

887668910110278144444444 

444444444444444444444444 

576323242235655633334444 

826480443191342735681369 

+ 

+ 

X 

........................ X 

X ........................ X 

........................ X 

x x x X X 
Lesions in Female Mice 203 

TABLED2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of Codeine: 3,000 ppm (continued) 

Number of Days on Study 

Carcass ID Number 

Special Senses System 

Eye

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Adenocarcinoma, metastatic, pancreas 

Histiocytic sarcoma 

Urinary bladder 

Adenocarcinoma,metastatic,pancreas 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant mixed 

11111111111111111111111111 

33333333333333333333333333 

44444444444455555555556666 

44444444444444444444444444 Total 

55666111111122344555612246 Tissues1 

05129014618959128369832604 Tumors 

+ + 3 

+ + 3 

X 2 

.......................... 50 

1 

1 

.......................... 50 

1 

.......................... 50 

1 

X X X 8 
204 Codeine, NTP TR 455 

HarderianGland:Adenoma 

Overall ratea 

Adjusted rateb 

Terminal rate' 

First incidence (days) 

Life table testd 

Logistic regression testd 

Cochran-Armita e tese 

Fisher exact test %

HarderianGland:Adenoma or Carcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Liver:HepatocellularAdenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Liver:HepatocellularCarcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Liver:HepatocellularAdenomaorCarcinoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Amitagetest 

Fisher exact test 

1/50 (2%) 

2.8% 

1/36 (3%) 

73 1 

P=O.524 

P=O.524 

P=O.524 

1/50 (2%) 

2.8% 

1/36 (346) 

73 1 

P=0.597N 

P=0.581N 

P=O.595 

12/50 (24%) 

32.0% 

11/36 (3 1 %) 

497 

P=0.054N 

P=0.045N 

P=0.061N 

5/50 (10%) 

12.5% 

3/36 (8%) 

565 

P=0.382N 

P=O.376N 

P=0.395N 

16/50 (32%) 

40.3 I 

13/36 (36%) 

497 

P=0.035N 

P=0.028N 

P=O.O39N 

750 ppm 

3/51 (6%) 

8.3% 

3/36 (8%) 

73 1 

P=O.305 

P=O.305 

P=O.316 

4/5 1 (8 %) 

10.5% 

3/36 (8%) 

616 

P=O.184 

P=O.173 

P=O.187 

10151 (20%) 

25.3% 

7/36 (19%) 

616 

P=0.408N 

P=0.424N 

P=0.385N 

6/51 (12%) 

15.7% 

4/36 (1 1 %) 

647 

P=O.492 

P=O.499 

P=O.514 

15/51 (29%) 

37.3% 

11/36 (31X) 

616 

P=0.506N 

P=0.520N 

P=0.474N 

1,500 ppm 

1/51 (2%) 

2.3% 

1/43 (2%) 

731 

P=0.722N 

P=0.722N 

P=0.748N 

1/51 (2%) 

2.3% 

1/43 (2%) 

73 1 

P=0.722N 

P=0.722N 

P=0.748N 

13/51 (25%) 

30.2% 

13/43 (30%) 

73 1 

P=0.488N 

P=0.565N 

P=O.523 

6/51 (12%) 

14.0% 

6/43 (14%) 

73 1 

P=O.610 

P=O.529 

P=O.514 

15/51 (29%) 

34.9% 

15/43 (35%) 

73 1 

P=0.288N 

P-0.413N 

P=0.474N 

3,000 ppm 

2/50 (4%) 

5.7% 

2/35 (6%) 

73 1 

P=O.490 

P=O.490 

P=O.500 

2/50 (4%) 

5.7% 

2/35 (6%) 

73 1 

P=O.490 

P=O.490 

P=O.500 

5/50 (10%) 

14.3% 

5/35 (14%) 

73 1 

P=0.059N 

P=0.047N 

P=0.054N 

4/50 (8%) 

10.5% 

2/35 (6%) 

720 

P=O.JOlN 

P=O.499N 

P=0.500N 

8/50 (16%) 

21.4% 

6/35 (17%) 

720 

P=0.060N 

P=0.047N 

P=0.050N 
Lesions in Female Mice 205 

TABLED3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-YearFeed Study of Codeine (continued) 

750 ppm 1,500 ppm 3,000 ppm 

Lung: Alveolarhronchiolar Adenoma 

Overall rate 1/50 (2%) 2/51 (4%) 6/51 (12%) 4/50 (8%) 

Terminal rate 

Adjusted rate 

1/36 (3%) 

2.8% 

2/36 (6%) 

5.6% 

5/43 (12%) 

13.5% 

2/35 (6%) 

10.4% 

First incidence (days) 73 1 731 724 711 

Life table test P=O.113 P-0.500 P=O.O96 P=O.185 

Logistic regression test P~O.130 P=O.500 P=O.O94 P=O.191 

Cochran-Armitage test P=O.111 

Fisher exact test P=O.508 P=O.O59 P=O.181 

Overall rate 

Lung: Alveolarhronchiolar Adenoma or Carcinoma 

2/50 (4%) 3/51 (6%) 7/51 (14%) 4/50 (8%) 

Adjusted rate .5.6% 8.3% 15.8% 10.4% 

Terminal rate 2136 (6%) 3/36 (8%) 6/43 (14%) 2/35 (6%) 

Life table test 

First incidence (days) 

P10.241 

73 1 

P=O.500 

73 1 

P=O.134 

724 

P=O.339 

711 

Logistic regression test P~O.276 P=O.500 P=O.135 P=O.357 

Cochran-Armitage test P~O.237 

Fisher exact test P=O.509 P=O.O85 P=O.339 

Ovary:Adenoma 

Overall rate 2/49 (4%) 3/51 (6%) 0150 (0%). 2/50 (4%) 

Adjusted rate 5.6% 8.3% 0.0% 5.7% 

First incidence (days) 

Terminal rate 

731 

2/36 (6%) 

731 

3/36 (8 96) 0/43 (0%) -e 731 

2/35 (6%) 

Life table test P=0.476N P=O.500 P=0.200N P=O.685 

Logistic regression test P=O.476N P=O.500 P=O.200N P=O.685 

Cochran-Armitage test P=O.47ON 

Fisher exact test P=O.519 P=0.242N P=0.684N 

PituitaryGland(Pars Distalis): Adenoma 

Overall rate 11/48 (2396) 9/47 (19%) 17/49 (35 96) 10146 (22%) 

Adjusted rate 29.7% 25.9% 39.4% 30.3% 

Terminal rate 10136(28%) 8/33 (24%) 16/42 (38%) 10133 (30%) 

First incidence (days) 722 5 12 703 73 1 

Life table test P=O.473 P=0.482N P=O.264 P=0.586N 

Logistic regression test P=O.Z39N P=0.498N P=O.269 P=0.528N 

Cochran-Armitage test P=O.472 

Fisher exact test P=0.422N P=O.146 P=O.544N 

Terminal rate 

Adjusted rate 

Overall rate 

PituitaryGland(ParsDistalis):Adenoma or Carcinoma 

10136 (2846) 

29.7 46 

11/48 (2396) 

8/33 (24%) 

25.9% 

9/47 (19%) 

17/42 (40%) 

41.7 % 

18/49 (37%) 

10133 (30%) 

30.3% 

10146 (22%) 

Life table test 

First incidence (days) 

P=O.460 

722 

P=0.482N 

512 

P=O.202 

703 

P=O.586N 

73 1 

Fisher exact test 

Cochran-Armitage test 

Logistic regression test 

P=O.460 

P=O.544 P=0.498N 

P-0.422N 

P=O.206 

P=O.103 

P=0.528N 

P=0.544N 
PituitaryGland (ParsIntermedia): Adenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

Skin (SubcutaneousTissue):FibrosarcomaorSarcoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

ThyroidGland(FollicularCell):Adenoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exacttest 

Uterus:StromalPolyp 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

AllOrgans:HemangiomaorHemangiosarcoma 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

I/48 (2 X) 

2.3% 

0136 (0%) 

570 

P=O.556 

P=O.545 

P=O.544 

1/50 (2%) 

2.8% 

1/36 (3%) 

731 

P=0.59JN 

P=0.577N 

P=O.595 

0/48 (0%) 

0.0% 

0/34 (0%) -

P=O.399 

P=O.399 

P=O.398 

4/50 (8%) 

10.6% 

3/36 (8%) 

65 1 

P=0.057N 

P=0.055N 

P=0.061N 

1/50 (2%) 

2.6% 

0136 (0%) 

713 

P=0.562N 

P=0.579N 

P=0.579N 

1/47 (2%) 

2.7% 

0133 (0%) 

677 

P=O.752 

P=0.744N 

P=O.747 

4/51 (8%) 

10.6% 

3/36 (8%) 

647 

P=O.180 

P=O.172 

P-0.187 

4/51 (8%) 

11.1% 

4/36 (1 1 X) 

731 

P=O.O70 

P=O.O70 

P=O.O66 

0151 (0%) 

0.0% 

0136 (0%) -

P=0.066N 

P-0.065N 

P=0.056N 

2/51 (4%) 

5.3% 

1/36 (3%) 

664 

P=O.494 

P=O.495 

P=O.508 

3/49 (6%) 

7.1% 

3/42 (7%) 

73 1 

P=O.357 

P=O.303 

P=O.316 

1/51 (2%) 

2.2% 

0143 (0%) 

727 

P=0.715N 

P=0.723N 

P=0.748N 

3/51 (6%) 

7.0% 

3/43 (7%) 

731 

P-0.166 

P~O.166 

P-0.133 

2/51 (4%) 

4.7% 

2/43 (5%) 

731 

P=0.263N 

P=0.296N 

P=0.329N 

3/51 (6%) 

6.7% 

2/43 (5%) 

703 

P=O.377 

P=O.338 

P=O.316 

1/46 (2%) 

3.0% 

1/33 (3%) 

73 1 

P=O.753 

P=O.765 

P=O.742 

2/50 (4%) 

5.7% 

2/35 (6%) 

731 

P=O.490 

P=O.490 

P=O.500 

2/50 (4%) 

5.7% 

2/35 (6%) 

731 

P=O.245 

P=O.245 

P=O.258 

0150 (0%) 

0.0% 

0/35 (0%) -

P=0.067N 

P=0.061N 

P=0.059N 

1/50 (2%) 

2.1 % 

0135 (0%) 

506 

P=0.757N 

P=0.759N 

P=0.753N 
Lesions in Female Mice 207 

TABLED3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-YearFeed Study of Codeine (continued) 

0 PPm 750 PPm 1,500 ppm 3,000 ppm 

All Organs: Malignant Lymphoma(Lymphocytic or Mixed) 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

All Organs:Benign 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

All Organs:Malignant 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

7/50 (14%) 5/51 (10%) 6/51 (12%) 8/50 (16%) 

17.4% 12.4% 12.8% 19.6% 

4/36 (1 1%) 3/36 (8X) 2/43 (5%) 4/35 (1 1 %) 

544 348 703 556 

P=O.385 P=0.386N P=0.369N P=O.503 

P=O.360 P=0.362N P=0.469N P=O.500 

P30.357 

P=0.366N P=0.485N P=O.500 

Neoplasms 

All Organs:BenignorMalignant 

Overall rate 

Adjusted rate 

Terminal rate 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

26/50 (52%) 25/51 (49%) 33/51 (65%) 19/50 (38%) 

64.7 96 60.8% 73.3% 51.1% 

22/36 (6146) 20/36 (56%) 31/43 (72%) 17/35 (49%) 

497 512 703 711 

P=O.llON P=0.509N P=O.430 P=0.129N 

P=0.065N P=0.547N P=O.257 P=0.080N 

P=O.l26N 

P=0.460N P=O.137 P=O. 114N 

Neoplasms 

15/50 (30%) 24/51 (47%) 17/51 (33%) 19/50 (38%) 

34.2% 52.0% 34.7% 41.7% 

8/36 (22%) 14/36 (39%) 11/43 (26%) 9/35 (26%) 

530 348 329 397 

P=O.468 PtO.077 P=0.530N P=O.293 

P=O.269 P=O.o60 P=O.187 P=O.162 

P~O.424 

PEO.060 P=O.442 P=O.263 

Neoplasms 

34/50 (6846) 36/5 1 (7 1 46) 38/51 (75%) 31/50 (62%) 

73.7% 76.5% 77.6% 67.2I 

24/36 (67%) 25/36 (69%) 32/43 (74%) 20/35 (57%) 

497 348 329 397 

P=0.284N P=O.420 P=0.430N P=0.392N 

P=0.332N P=O.421 P=O.358 P=0.336N 

P=0.271N 

P=O.474 P=O.308 P=0.338N 

a Number of neoplasm-bearing animalslnumber of animals examined. Denominator is number of animals examined microscopically for liver, lung, ovary, 

pituitary gland, thyroid gland, and uterus; for other tissues, denominator is number of animals necropsied. 

Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to 

pairwise comparisons between the controls and that exposed group. The life table test regards neoplasms inanimals dying prior to terminal kill as being 

(directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests 

compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. 

e Not applicable;noneoplasmsin animal group 
208 Codeine, NTP TR 455 

TABLED4 

Historical Incidence of Hepatocellular Neoplasms in Untreated Female B6C3Fl Micea 

Incidence inControls 

Adenoma CarcinomaAdenomaor Carcinoma 

Historical Incidence at Microbiological Associates, Inc. 

C.I. Direct Blue 218 7/49 5/49 10/49 

dl-Amphetamine Sulfate 5/50 0/50 5/50 

Overall Historical Incidence 

Total 194/1,312 (14.8%) 90/1,312 (6.9%) 260/1,312 (19.8%) 

Standard deviation 10.5% 6.1% 12.8% 

Range 2%-50% 0%-20% 3%-56% 

a Data as of 17 June 1994 
Lesions 209 in Female Mice 

750 ppm 1,500 ppm 3,000 ppm 

Disposition Summary 

Animals initially in study 60 60 60 60 

IS-MonUI interim evduafion 10 9 9 9 

Early deaths 

Moribund 7 9 6 4 

Natural deaths 7 6 2 11 

Survivors 

Terminal sacrifice 

Died last week of study 

3s 

1 

36 43 34 

1 

Missing 1 

Animals examined microscopically 60 60 60 59 

15-Month Interim Evaluation 

Alimentary System 

Liver 

Basophilic focus 

(9) 

Mixed cell focus 

Inflammation, chronic active 3 (33%) 

1 (11%) 

2 (22%) 2 (22%) 

Mesentery 

Fat, necrosis 

Pancreas 

Inflammation, chronic 

Salivary glands 

Inflammation, chronic 

Dilatation 

Stomach, glandular 

Inflammation, chronic active 1 (11%) 

Endocrine System 

Adrenal cortex 

Subcapsular, hyperplasia 

Adrenal medulla 

Hyperplasia 

Pituitary gland 

Thyroid gland 

Pars distalis, hyperplasia 

Follicular cell, hyperplasia, focal 

Genital System 

Clitoral gland 

Dilatation 

Ovary 

cyst 

UkNS 

Dilatation 

Hyperplasia, cystic 

a Number of animals examined microscopically at the site and the number of animals with lesion 
210 Codeine, NTP TR 455 

TABLED5 

Summary of the Incidence of Nonneoplastic Lesionsin Female Mice in the 2-Year Feed Study of Codeine (continued) 

750 ppm 

(9)

1 (11%) 

(9) 

1 (11%) 

(9)

6 (67%) 

(9)

7 (78%) 

6 (67%) 

1 (11%). 

(9) 

1 (11%) 

7 (78%) 

1 (11%) 

(9)

7 (78%) 

1,500 ppm 

(9) 

(9) 

1 (11%) 

(9) 

9 (100%) 

(9) 

8 (89%) 

8 (89%) 

1 (11%) 

(9) 

7 (78%) 

(9) 

9 (100%) 

3,000 ppm 

(9) 

(9) 

(9) 

4 (44%) 

(9) 

9 (100%) 

8 (89%) 

1 (11%) 

(9) 

5 (56%) 

(9)

6 (67%) 

0 PPm 

15-Month Interim Evaluation (continued) 

Hematopoietic System 

Lymph node, mandibular 

Hyperplasia, lymphoid 

Thymus 

Hyperplasia, lymphoid 

Integumentary System 

Mammary gland 

Intiitration cellular, lymphocyte 

Skin 

Inflammation, chronic active 

Musculoskeletal System 

Bone 

Dysplasia 

Nervous System 

Brain 

Mineralization 

Respiratory System 

Lung 

Infiltration cellular, lymphocyte 

Nose 

Olfactory epithelium, degeneration, hyaline 

RespiratoIyepithelium,degeneration,hyaline 

Respiratory epithelium, inflammation, 

chronic active 

Urinary System 

Kidney 

cyst 

Inflammation, chronic 

Renal tubule, regeneration 

Urinary bladder 

Infiltration cellular, lymphocyte 

(9) 

(10) 

(10) 

(10)

2 (20%) 

(10) 

9 (90%) 

(10) 

9 (90%) 

7 (70%) 

1 (10%) 

(10) 

5 (50%) 

2 (20%) 

(10) 

8 (80%) 

Systems Examined WithN o Lesions Observed 

Cardiovascular System 

General Body System 

Special Senses System 
Lesions in Female Mice 211 

TABLED5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-YearFeed Study of Codeine (continued) 

2-Year Study 

Alimentary System 

Gallbladder 

Inflammation, chronic active 

Intestine small, duodenum 

Ulcer 

Intestine small, jejunum 

Hemorrhage 

Hyperplasia, lymphoid 

Liver 

Angiectasis 

Atrophy

Basophilic focus 

Clear cell. focus 

Congestion 

cyst 

Eosinophilic focus 

Fatty change, focal 

Hematopoietic cell proliferation 

Infarct 

Inflammation, chronic active 

Mineralization 

Mitotic alteration 

Mixed cell focus 

Necrosis 

Bile duct, hyperplasia 

Centrilobular, inflammation, chronic 

Centrilobular, necrosis 

Centrilobular, pigmentation 

Hepatocyte, necrosis 

Kupffer cell, hypertrophy 

Mesentery 

Inflammation, chronic active 

Artery, inflammation, chronic active 

Fat, necrosis 

Pancreas 

Inflammation, chronic 

Inflammation, chronic active 

Acinar cell, atrophy 

Acinar cell, depletion secretory 

Duct, cyst 

Salivary glands 

Inflammation, chronic 

Stomach, forestomach 

Hyperplasia 

Inflammation, acute 

Inflammation, chronic active 

Stomach, glandular 

cyst 

Dysplasia 

Inflammation, chronic active 

Necrosis 

0 PPm 

(43) 

1 (2%) 

(48) 

(50) 

(50) 

1 (2%) 

6 (12%) 

3 (6%) 

7 (14%) 

2 (4%) 

1 (2%) 

(9) 

2 (22%) 

5 (56%) 

(50) 

1 (2%) 

2 (4%) 

2 (4%) 

1 (2%) 

1 (2%) 

(50) 

28 (56%) 

(50) 

1 (2%) 

2 (4%) 

(50) 

1 (2%) 

3 (6%) 

2 (4%) 

750 ppm 

(42) 

(49) 

(48) 

1 (2%) 

(51) 

1 (2%) 

7 (14%) 

5 (10%) 

7 (14%) 

1 (2%) 

2 (4%) 

1 (2%) 

(9) 

8 (89%) 

(5 1) 

1 (2%) 

1 (2%) 

(50) 

28 (56%) 

(51) 

1 (2%) 

(5 1) 

9 (18%) 

1 (2%) 

1,500 ppm 

(47) 

(48) 

1 (2%) 

(50) 

1 (2%) 

(51) 

1 (2%) 

1 (2%) 

1 (2%) 

2 (4%) 

9 (18%) 

1 (2%) 

2 (4%) 

12 (24%) 

1 (2%) 

2 (4%) 

2 (4%) 

1 (2%) 

1 (2%) 

1 (2%) 

(7) 

1 (14%) 

6 (86%) 

(5 1) 

1 (2%) 

1 (2%) 

(51) 

27 (53%) 

(51) 

2 (4%) 

2 (4%) 

(51) 

1 (2%) 

3,000 ppm 

(41) 

1 (2%) 

(48) 

(50) 

1 (2%) 

1 (2%) 

(50) 

1 (2%) 

1 (2%) 

1 (2%) 

2 (4%) 

3 (6%) 

1 (2%) 

4 (8%) 

1 (2%) 

1 (2%) 

2 (4%) 

1 (2%) 

(5) 

1 (20%) 

2 (40%) 

(50) 

2 (4%) 

2 (4%) 

2 (4%) 

(50) 

23 (46%) 

(50) 

1 (2%) 

1 (2%) 

3 (6%) 

(49) 

1 (2%) 

6 (12%) 
212 

2-Year SWdy (continued) 

Cardiovascular System 

Blood vessel 

Aorta, inflammation, chronic active 

Heart 

Fibrosis 

Inflammation, chronic active 

Mineralization 

Thrombosis 

Artery, inflammation, chronic active 

Endocrine System 

Adrenal cortex 

Angiectasis 

cyst 

Cytoplasmic alteration, focal 

Hematopoietic cell proliferation 

Hypertrophy, focal 

Inflammation, chronic active 

Subcapsular, hyperplasia 

Adrenal medulla 

Hyperplasia 

Pituitary gland 

Craniopharyngeal duct, hyperplasia 

Pars distalis, angiectasis 

Pars distalis, cyst 

Pars distalis, hyperplasia 

Pars intermedia. hyperplasia 

Thyroid gland 

Inflammation, chronic active 

Follicle, cyst 

Follicular cell, hyperplasia, diffuse 

Follicular cell, hyperplasia, focal 

General Body System 

None 

Genital System 

Clitoral gland 

Dilatation 

Inflammation, chronic active 

Pigmentation 

Ovary 

Abscess 

cyst 

Hemorrhage 

Infiltration cellular, histiocyte 

Inflammation, chronic active 

Penovarian tissue, infiltration cellular, 

lymphocyte 

0 PPm 

47 (94%) 

(49) 

2(4%) 

11 (23%) 

1 (2%) 

(48) 

2 (4%) 

3 (6%) 

14 (29%) 

2 (4%) 

750 ppm 

(51) 

1(2%) 

2 (4%) 

(47) 

1 (2%) 

1 (2%) 

12 (26%) 

(51) 

6 (12%) 

1 (2%) 

29 (57%) 

11 (22%) 

1,500 ppm 

1(2%) 

1 (2%) 

11 (22%) 

(51) 

10 (20%) 

5 (10%) 

1 (2%) 

42 (82%) 

12 (24%) 

1 (2%) 

2(4%) 

Codeine, NTP TR 455 

3,000 ppm 

(50) 

1 (2%) 

3 (6%) 

45 (90%) 

(48) 

2 (4%) 

(46) 

2(4%) 

2 (4%) 

10 (22%) 

1 (2%) 

(50)

4 (8%) 

3 (6%) 

44 (88%) 

(44) 

41 (93%) 

10 (23%) 

1(2%) 

(50) 

3 (6%) 

6 (12%) 

2 (4%) 

1(2%) 
Lesions in Female Mice 213 

TABLEDS 

Summary of the Incidence of Nonneoplastic ~IOMin Female Mice in the2-YearFeed Study OF Codeine (continued) 

0 PPm 750 ppm 1,500 ppm 3,000 ppm 

2-Yew Study (continued) 

Genital System (continued) 

Oviduct 

cyst 

Dilatation 

Mammation. chronic active 

UaNS 

Atrophy

Dilatation 

Hemorrhage 

Hyperplasia, cystic 

Inflammation, chronic active 

Cervix, Mammation, chronic active 

Lymphatic, ectasia 

Hematopoietic System 

Bone marrow 

Atrophy

Congestion 

Myeloid cell, hyperplasia 

Lymph node 

Mac. hyperplasia, lymphoid 

Iliac, inflammation. chronic active 

Inguinal, pigmentation 

Mediastinal, hyperplasia, plasma cell 

Lymph node, mandibular 

Hemorrhage 

Hyperplasia, lymphoid 

Lymph node, mesenteric 

cyst 

Fibrosis 

Hematopoietic cell proliferation 

Hemorrhage 

Hyperplasia, lymphoid 

Infitrationcellular, histiocyte 

Intlammation, chronic active 

Spleen 

Atrophy

Fibrosis 

Hematopoietic cell proliferation 

Hyperplasia, lymphoid 

Mitration cellular, plasma cell 

Pigmentation, hemosiderin 

Capsule, inflammation, chronic active 

Thymus 

Atrophy

Ectopic parathyroid gland 

Ectopic thyroid 

Hyperplasia, lymphoid 

(3) 

1 (33%) 

1(33%) 

(50) 

10 (20%) 

36 (72%) 

1 (25%) 

1 (50%) 

1 (11%) 

(49) (49) 

3(6%) 

(47) 
214 Codeine, NTP TR 455 

750 ppm 3,000 ppm 

IntegumentarySystem 

Mammary gland 

Hyperplasia 

Inflammation, chronic active 

Skin 

Developmental malformation 

Ulcer 

Inflammation, chronic active 6 (12%) 5 (10%) 

Epidermis, hyperplasia 

Subcutaneous tissue, abscess 

Subcutaneous tissue, inflammation, 

chronic active 

Subcutaneous tissue, pigmentation 

Subcutaneous tissue, fat, atrophy 

Mu&uloskeletal System 

Bone 

Artery, inflammation, chronic active 

Fracture 

Dysplasia 

(50) 

11(22%) 

(51) 

5 (10%) 

(51) 

1 (2%) 

1 (2%) 

8 (16%) 

(50) 

6 (12%) 

Nervous System 

Infitration cellular, histiocyte 

Hemorrhage 

Degeneration 

Compression 

Brain (50) 

1 (2%) 

3 (6%) 

(51) 

1 (2%) 

1(2%) 

Artery, inflammation, chronic active 

Mineralization 23 (46%) 18 (35%) 

Meninges, inflammation, chronic 1 (2%) 

Respiratory System 

Lung

Congestion 

Hemorrhage 

Infiitration cellular, lymphocyte 

Infiitration cellular, histiocyte 

Pigmentation, hemosiderin 

Alveolar epithelium, hyperplasia 

Nose 

Respiratory epithelium, hyperplasia, 

Olfactory epithelium, degeneration, hyaline 

glandular 
Lesions in Female Mice 215 

TABLED5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-YearFeed Study of Codeine (continued) 

0 PPm 750 ppm 1,500 ppm 

2-Year Study (continued) 

Special Senses System 

EY e (2) 

Atrophy

Inflammation, chronic active 1 (50%) 

1 (50%) 

Cornea, inflammation, chronic active 

Inflammation, chronic active 

Hyperplasia, focal 

Harderian gland (2) 

1 (50%) 

(4) (3) 

2 (67%) 

Kidney 

Urinary System 

Amyloid deposition 

Glomerulosclerosis

Atrophy 

Hemorrhage 

Hydronephrosis 

Infarct 

Metaplasia, osseous 

Inflammation, chronic 12 (24%) 

Capsule, inflammation, chronic active 

Nephropathy 3 (6%) 

Glomerulus. inflammation, chronic 

Perirenal tissue, inflammation, 

chronic active 

Renal tubule, degeneration, hyaline 

Renal tubule, pigmentation 

Renal tubule, karyomegaly 1 (2%) 

Renal tubule, regeneration 

Infiitrationcellular, lymphocyte 

Ureter 

Dilatation 

Urinary bladder 

19 (38%) 27 (53%) 

(50) (5 1) 

(1) 

31 (61%) 

1 (100%) 

Inflammation, chronic active 

Infiitration cellular, lymphocyte 38 (76%) 37 (73%) 

Artery, inflammation, chronic active 1 (2%) 

3,000 ppm 

(3) 

2 (67%) 

1 (33%) 

(3) 

1 (33%) 

1 (2%) 

1 (2%) 

1 (2%) 

2(4%) 

13 (26%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

1 (2%) 

19 (38%) 

(50) 

36 (72%) 

1 (2%) 
217 

APPENDIX E 

GENETIC TOXICOLOGY 

SALMOIVELLAMUTAGENICITY TEsr PROTOCOL . . . . . . .. . . . . . . . . . . . . . . .. . . . . . . . . . . . 218 

CHINESEHAMSTER OVARY CELLCYTOGENETICS pRoToco~s . ..... . . . . . . .. . . . . . . . . . . . 218 

RESULTS ......................................................... 219 

TABLEEl Mutagenicity of CodeinePhosphatein Salmonella typhimurium ... . . . . . . . . . . . . . 221 

TABLE E2 Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells 

byCodeinePhosphate . . . . . . . , . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . 222 

TABLEE3 Induction of ChromosomalAberrationsinChineseHamsterOvaryCells 

byCodeinePhosphate . . . . . ...... . . . . . . . . . . . . . . . . . . . .. . . . .. . .. . . . 224 
218 Codeine, NTP TR 455 

GENETIC TOXICOLOGY 

SALMONELLAMUTAGENICITY TEST PROTOCOL 

Testing was performed as reported by Zeiger et al. (1992). Codeine phosphate was sent to the laboratory 

as a coded aliquot from Radian Corporation (Austin, TX). It was incubated with the Salmonella 

typhimurium tester strains (TA97, TA98, TA100, or TA1535) either in buffer or S9 mix (metabolic 

activation enzymes and cofactors from Aroclor 1254-induced male Sprague-Dawley rat or Syrian hamster 

liver) for 20 minutes at 37" C. Top agar supplemented with I-histidine and d-biotin was added, and the 

contents of the tubes were mixed and poured onto the surfaces, of minimal glucose agar plates. Histidineindependent mutant colonies arising on these plates were counted following incubation for 2 days at 

37" c. 

Each trial consisted of triplicate plates of concurrent positive and negative controls and five doses of 

codeine phosphate. In the absence of toxicity, 10,000 pg/plate wasselectedasthehigh dose. All trials 

were repeated. 

In this assay, a positive response is defined as a reproducible, dose-related increase in histidineindependent (revertant) colonies in any one strain/activation combination. An equivocal response is 

defined as an increase in revertants that is not dose related, not reproducible, or is of insufficient 

magnitude to support a determination of mutagenicity. A negative response is obtained when no increase 

in revertant colonies is observed following chemical treatment. There was no minimum percentage or fold 

increase required for a chemical to be judged positive or weakly positive. 

CHINESEHAMSTER OVARY CELLCYTOGENETICS PROTOCOLS

Testing was performed as reported by Galloway et al. (1987). Codeine phosphate was sent to the 

laboratory as a coded aliquot by Radian Corporation. It was tested in cultured Chinese hamster ovary 

(CHO) cells for induction of sister chromatid exchanges(SCEs) and chromosomal aberrations (Abs), both 

in the presence and absence of Aroclor 1254-induced male Sprague-Dawley rat liver S9 and cofactor mix. 

Cultures were handled under gold lights to prevent photolysisof bromodeoxyuridine-substituted DNA. 

Each test consisted of concurrent solvent and positive controls and of at least three doses of codeine 

phosphate; the high dose in the absence of S9 was limited by toxicity. In the presense of S9, the high 

dose was limited to 10,000 mg/mL. A single flask per dose was used, and tests yielding equivocal or 

positive results were repeated. In both the SCE and Abs tests, higher doses of codeine phosphate 

produced an acidic pH shift in the cultures; some experiments employedN-(2-hydroxyethyl)piperazine-N'-

(Zethanesulfonic acid) (HEPES) buffer to counter this effect. 

Sister Chromatid Exchange Test: In the SCE test without S9, CHO cells were incubated for a minimum of 

25.6 hours with codeine phosphate in McCoy's 5A medium supplemented with fetal bovine serum, 

1-glutamine,andantibiotics.Bromodeoxyuridine(BrdU)wasadded 2 hours after culture initiation. After 

approximately 26 hours, the medium containing codeine phosphate was removed and replaced with fresh 

medium plus BrdU and Colcemid, and incubation was continued for 2 hours. Cells were then harvested 

by mitotic shake-off, fiied, and stained with Hoechst 33258 and Giemsa. In the SCE test with S9, cells 

were incubated with codeine phosphate, serum-free medium, and S9 for 2 hours. The medium was then 

removed and replaced with medium containing serum and BrdU and no codeine phosphate and incubation 

proceeded for an additional 25.4 hours or more, with Colcemid present for the final 2 hours. Harvesting 

and staining were the same as for cells treated without S9. All slides were scored blind and those from a 

single test were read by the same person. For trial 1, both with and without S9, fifty second-division 

metaphase cells were scored for frequency of SCEs/cell from each dose level. Because of the positive 
Genetic Toxicology 219 

response, 20 second-division metaphase cells were scored in trials 2 and 3. Because significant chemicalinduced cell cycle delay was seen, incubation time was lengthened to ensure a sufficient number of 

scorable (second-division metaphase) cells. 

Statistical analyses were conducted on the slopes of the dose-response curves and the individual dose 

points (Galloway et al., 1987). An SCE frequency 20%. above the concurrent solvent control value was 

chosen as' a statistically conservative positive response. The probability of this level of difference 

occurring by chance at one dose point is less than 0.01; the probability for such a chance occurrence at 

two dose points is less than 0.001. A n increase of 20% or greater at any single dose was considered weak 

evidence of activity; increases at two or more doses resulted in a determination that the trial was positive. 

A statistically significant trend(P<0.005) in the absence of any responses reaching 20% above 

background led to a call of equivocal. 

Chromsomal Aberrations Test: In the Abs test without S9, cells were incubated in McCoy's 5A medium 

with codeine phosphate for 18.5 or 18.6 hours; Colcemid was added and incubation continued for 2 hours. 

The cells were then harvested by mitotic shake-off, fixed, and stained with Giemsa. For the Abs test with 

S9, cells were treated with codeine phosphate and S9 for 2 hours, after which the treatment medium was 

removed and the cells were incubated for 18.5 hours in fresh medium, with Colcemid present for the final 

2 hours. Cells were harvested in the same manner as for the treatment without S9. The harvest time for 

the Abs test was based on the cell cycle information obtained in the SCE test; cell cycle delay was 

anticipated, and the incubation period was extended. 

Cells were selected for scoring on the basis of good morphology and completeness of karyotype 

(21 f 2 chromosomes). All slides were scored blind and those from a single test were read by the same 

person. One hundred first-division metaphase cells were scored at each dose level. Classes of aberrations 

included simple (breaks and terminal deletions), complex (rearrangements and translocations), and other 

(pulverized cells, despiralized chromosomes, and cells containing 10 or more aberrations). 

Chromosomal aberration data are presented as percentage of cells with aberrations. To arrive at a 

statistical call for a trial, analyses were conducted on both the dose response curve and individual dose 

points. For a single trial, a statistically significant (PSO.05) difference for one dose point and a 

significant trend (PSO.015) were considered weak evidence for a positive response; significant differences 

for two or more doses indicated the trial was positive. A positive trend test in the absence of a 

statistically significant increase at any one dose resulted in an equivocal call (Gallowayet al., 1987). 

Ultimately, the trial calls were based on a consideration of the statistical analyses as well as the biological 

information available to the reviewers. 

RIZSULTS 

Codeine phosphate (100 to 10,000 pglplate) was not mutagenic in Salmonella typhimrium strains TA97, 

TA98, TA100, or TA1535, with or without induced liver S9 enzymes (Table El; Zeiger et al., 1992). 

Dose-related increases in the frequency of SCEs were induced by codeine phosphate in cultured CHO 

cells, with and without S9 (Table E2), but these significantly increased frequencies were noted only at 

doses that caused marked cell cycle delay, indicative of a high level of cytotoxicity. The role of 

confounding factors, such as toxicity-related changes in DNA metabolism, needs to be considered in the 

evaluation of cytogenetic data. Slowly cycling cells undergo prolonged exposure to 5-bromodeoxyuridine, 

which can also result in an increased level of SCEs. A clear association between cytotoxicity and SCE 

frequencies has not been shown. There are numerous examples of chemicals that induce cell cycle delay 

without inducing SCEs (Galloway et al., 1987). Acidic pH shifts were noted in cultures exposed to higher 

concentrations of codeine phosphate; addition of HEPES buffer in some of the trials stabilized the pH. 
220 Codeine, NTP TR 455 

SCE frequencies did not appear to be affected by the pHshifts; increases were noted both in acidic and 

neutral cultures. No increases in the frequency of Abs were noted in cultured CHO.cells treated with 

codeine phosphateat doses similar to those used in theSCE test (Table E3); codeine phosphate-induced 

cell cycle delay was also observed in these cultures. 
Genetic Toxicology 221 

TABLEEl 

Mutagenicity of Codeine Phosphate in Salmonella @phimunuma 

Revertants/plateb 

Strain Dose 439 +hamster S9 +rat S9

Trial(&plate) 1 Trial 2 

TAlOO 0 102 f 5.7 98 f 6.7 110 f 8.7 120 f 6.1 107 f 4.0 115 f 8.1 

100 100 f 6.1 97 f 5.6 102 f 9.9 120 f 5.1 89 f 8.3 101 f 5.9 

333 87 f 3.0 90 f 7.0 112 f 10.9 113 f 7.2 95 f 3.3 116 f 3.8 

10,m 

3,333 

1,m 

107 f 3.2 

97 f 1.7 

89 f 7.6 

93 f 3.5 

92 f 4.4 

104 f 13.9 

114 f 11.3 

97 f 2.4 

111 f 5.8 

114 f 7.1 

115 f 1.5 

119 f 10.6 

104 f 8.7 

100 f 2.6 

86 f 5.0 

107 f 13.1 

110 f 3.8 

121 f 9.2 

Positive control' 

Trial summary 

558 f 19.2 

Negative 

365 f 14.2 

Negative 

1.649 f 10.3 

Negative 

549 f 4.8 

Negative 

1,260 f 14.2 

Negative 

380 f 18.7 

Negative 

TA1535 0 23 f 2.0 8 f 2.0 7 f 0.7 10 f 1.9 10 f 1.2 12 f 1.2 

100 19 f 3.5 5 f 0.9 8 f 1.0 6 f 1.2 10 f 0.3 8 f 0.9 

333 24 f 3.0 7 f 1.5 9 f 2.3 9 f 1.7 5 f 0.3 10 f 1.5 

1,000 26 f 1.2 4 f 0.3 8 f 1.5 10 f 3.2 10 f 2.3 10 f 0.3 

3,333 26 f 2.3 4 f 1.5 5 f 1.5 8 f 1.2 8 f 0.7 7 f 1.5 

10,000 30 f 0.9 8 f 1.0 8 f 2.2 7 f 0.3 6 f 0.9 7 f 0.3 

Trial summary Negative Negative Negative Negative Negative Negative 

Positive control 485 f 17.1 274 f 5.5 407 f 16.7 132 f 4.5 281 f 3.8 155 f 3.5 

TAW 

0 136 f 6.6 124 f 14.5 175 f 6.0 160 f io.2 180 f 18.7 170 f 16.3 

100 137 f 2.6 121 f 8.5 149 f 15.5 154 f 5.6 176 f 7.4 215 f 10.4 

333 115 f 8.6 110 f 8.0 141 f 3.6 174 f 25.4 190 f 14.9 199 f 6.5 

1,000 138 f 6.4 134 f 10.4 146 f 16.0 194 f 22.8 198 f 12.1 192 f 9.0 

3,333 128 f 0.9 121 f 15.9 152 f 2.3 170 f 14.5 193 f 10.3 194 f 18.4 

10,m 148 f 12.5 141 f 14.8 153 f 16.5 173 f 25.5 196 f 6.0 199 f 7.6 

Trial summary Negative Negative Negative Negative Negative Negative 

Positive control 1,931 f 109.1 2,102 f 134.1 1,346 f 93.5 1,302 f 28.0 1,070 f 28.0 558 f 19.4 

TA98 0 13 f 1.9 18 f 3.5 27 f 5.5 19 f 2.6 28 f 3.2 18 f 1.2 

100 13 f 1.0 15 f 1.3 30 f 2.3 22 f 3.4 32 f 0.6 20 f 2.8 

333 19 f 3.8 12 f 3.5 28 f 2.6 21 f 6.6 32 f 2.5 23 f 1.3 

1,Ooo 19 f 1.2 15 f 1.5 36 f 0.9 21 f 1.5 30 f 5.3 16 f 5.7 

3,333 17 f 3.8 17 f 0.9 32 f 3.0 26 f 2.7 32 f 1.7 27 f 5.5 

10,Ooo 26 f 1.2 17 f 3.7 39 f 2.0 18 f 5.9 36 f 2.0 20 f 4.6 

Trial summary Equivocal Negative Negative Negative Negative Negative 

Positive control 1,278 f 52.0 1,824 f 19.1 1.029 f 20.0 338 f 7.9 823 f 16.7 142 f 10.5 

a Study performed at SRI, International. The detailed protocol and these data are presented in Zeiger et al. (1992). 

Revertants are presented as mean f standard error from three plates. ' The positive controls in the absence of metabolic activation were sodium azide(TA100 and TA1535). 9-aminoacridine (TA97); and 

4-nitro-o-phenylenediamine(TA98). The positive control for metabolic activation with all strains was 2-aminoanthracine. 

10%30% 10% 30% 
222 Codeine, NTP TR 455 

TABLEE2 

Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells by Codeine Phosphatea 

No. of scEs/ Relative 

DoseTtalChromo- No. of Chromo-SCEs/HrsChange of SCEs/ 

Compound(CrglmL)Cels somes SCEssomeCell in BrdUChromosomeb 

(%I 

439 

Trial 1 

Summary:Positive 

Dimethylsulfoxide 50 1,0230.42

Mitomycin-C 0.0015 50 1,027 753 0.73 15.1 25.7 72.43 

0.0100 5 103 265 2.57 53.0 25.7 505.07 

phosphate Codeine 1,ooo 50 1,031 541 0.52 10.8 30.9' 23.40*

1,500 50 1.029 560 0.54 11.2 30.9' 27.99* 

2,ooo 50 1.012 574 0.56 11.5 30.9' 33.39* 

2,500 0 

P<0.001d 

Trial 2 

Summary:Positive 

8.7 435 25.7 

4090.39

Mitomycin-C 0.0015 20 404 289 0.71 14.5 25.6 79.49 

0.0100 5 105 252 2.40 50.4 25.6 502.21 

phosphate Codeine 1,O@J 20 400 173 0.43 8.7 25.6 8.52

Dimethylsulfoxide 20 8.2 163 25.6 

409 1.500 224 0.54 11.2 32.3' 37.42*

2,ooo 20 417 236 0.56 11.8 32.3' 42.01* 

2,500 0 

P<O.001 

Trial 3 

Summary:Positive 

20 

Dimethylsulfoxide 20 8.7 4050.42 

412 301 

173 25.6 

Mitomycin-C 0.0015 20 0.73 15.1 25.6 71.03

0.0100 5 103 217 2.10 43.4 25.6 393.2 1 

Codeine phosphate 1,ooo 20 408 178 0.43 8.9 25.6 2.13

1,500 20 402 278 0.69 13.9 32.2' 61.89* 

2,ooo 20 414 305 0.73 15.3 32.2' 72.47* 

2,500 0 

P<O.ool 

* Positive response (220% increase over solvent control) a Study performed at Limn Bionetics, Inc. A detailed description of the protocol is presented in Galloway et al. (1987). SCE=sister 

chromatid exchange; BrdU=bromodeoxyuridine. 

SCEs/chromosome in treated cells versus SCEs/chromosome in solventcontrol cells. ' Because codeine phosphate induced a delay in the cell division cycle, harvest time was extended to maximize the number of seconddivision cells available for analysis. 
Genetic Toxicology 223 

TABLEE2 

Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cellsby Codeine Phosphate(continued) 

Compound(CcglmL)CelssomesSCEssome 

, DoseTtalChromoNo. of No. of 

SCEsl 

Chromo-SCEsl 

Cell 

Relative 

in BrdUChromosome 

Hrs Change of SCEs/ 

(%I 

Summary:Positive 

Trial 1 

+s9 

Dimethylsulfoxide 50 1,029 462 0.4425.4 9.2 

Cyclophosphamide 

0.4 50 1.028 733 0.71 14.7 25.4 58.81 

2.0 5 103 177 1.71 35.4 25.4 282.75 

Codeine phosphate 

500 50 1,033 553 0.53 11.1 25.4 19.24 

1,700 50 1,008 603 0.59 12.1 33.9c 33.24* 

5,me 50 1,015 742 0.73 14.8 33.9c 62.82* 

P<0.001 

Trial 2 

Summary:Positive 

Dimethylsulfoxide 0.46 20

Cyclophosphamide 

25.8 9.4 403 188 

0.4 20 415 295 0.71 14.8 25.8 52.38 

2.0 5 102 208 2.03 41.6 25.8 337.13 

Codeine phosphatee 

8,OOO 20 413 248 0.60 12.4 31 .Oc 28.72* 

9,@33 20 414 269 0.64 13.5 31.0' 39.28* 

10,Ooo 20 410 280 0.68 14.0 31 .Oc 46.39* 

P<O.OOl 

Trial 3 

Summary:Weaklypositive 

Dimethylsulfoxide 11.6 25.6 0.57 20

Cyclophosphamide 

401 232 

0.4 20 401 267 0.66 13.4 25.6 15.09 

2.0 5 103 192 1.86 38.4 25.6 222.20 

Codeine phosphatef 

9.000 20 402 269 0.66 13.5 25.6 15.66 

9,500 20 407 279 0.68 14.0 25.6 18.49 

10,Ooo 20 401 309 0.77 15.5 32.2 33.19* 

P<O.001 

Significance of relative SCEslchromosome tested by the linear regression trend test vs. logof the dose 

e AcidicpH shift 

HEPES buffer used;no acidic pH shift noted at any of the doses tested 
224 Codeine, NTP TR 455 

TABLEE3 

Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells by Codeine Phosphatea 

439 +s9 

Dose Total No. of Abs/ CeUs with Dose Total No. of Abs/ Cells with 

(ccg/mL) CeUs Abs Cell Abs (96) (CcglmL) Cells Abs Cell Abs (%) 

Trial 1- Harvest time: 20.5 hoursb Trial 1- Harvest time: 20.5 hours 

Summary: Negative Summary: Negative 

Dimethylsulfoxide Dimethylsulfoxide 

3.03 

Mitomycin-C Cyclophosphamide 

0.0026 100 15 0.15 14.0 2.5 ' 100 7

100 0.03 100 1 0.01 1.o 

7.0 0.07 

8 24.0

Codeine phosphate Codeine phosphate 

2,o0o 100 0 0.00 0.0 9,000c 100 4 0.04 4.0 

2,500 100 4 0.04 4.0 9,500 100 3 0.03 3.0 

3,000c 100 4 0.04 3.0 lo.m 100 3 0.03 3.0 

P=0.271d P=O.247 

Trial 2 - Harvest time: 20.6 hours Trial 2 - Harvest time: 20.5 hours 

Summary: Negative Summary: Negative 

Dimethylsulfoxide Dimethylsulfoxide 

0.0620 25 13 0.52 40.0 25 12.5 0.32 

3.03 100 0 0.00 0.0 

Mitomycin-C Cyclophosphamide 

0.0250 100 16 0.16 14.0 2.5 100 1 0.01 1.o 

100 0.03 

0.0625 25 6 0.24 24.0 25 12.5 6 ' 0.24 24.0

Codeine phosphate Codeine phosphate' 

1 0.01 1.o 9,m 100 2 0.02 2.0 

0.02 2.0 9,500 100 2 0.02 2.0 

2,500 100 

3,000c 100 2 

3,500 0 lo.m 100 1 0.01 1.o

P-0.693 P=O.271 
Genetic Toxicology 225 

TABLEE3 

Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells by Codeine Phosphate (continued) 

49 

Dose Total No. of Abs/ Cells with 

(pg/mL) Cells Abs Cell Abs (%) 

Trial 3 - Harvest time: 20.6 hours 

Summary:Negative 

Dimethylsulfoxide 

100 3 ' 0.03 3.0

Mitomycin-C 

0.0250 100 9 0.09 9.0 

0.0625 25 . 5 0.20 20.0 

Codeine phosphate 

3,500 100 3 0.03 3.0 

4,ooo 100 4 0.04 2.0 

4,500 100 4 0.04 4.0 

5,Ooo 0 

P=O.404 

a StudyperformedatLittonBionetics,Inc.ThedetailedprotocolispresentedinGalloway et al. (1987). Abs=aberrations. 

Because of significant chemical-induced cell cycle delay, incubation time prior to addition of Colcemid was lengthened to provide 

sufficient metaphase cells at harvest. 

Acidic pH shift at doses of 3,000 pglmL and higher 

Significance of percent cells with aberrations tested by the linear regression trend test vs. log of the dose 

e HEPES buffer used; no pH shift noted at any doses in this trial 
227 

APPENDIX F 

ORGAN WEIGHTS 

AND ORGAN-WEIGHT-TO-BODY-WEIGHT RATIOS 

TABLEF1 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

in the 14-Day Feed Study of Codeine ................................ 228 

TABLE F2 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

in the 13-Week Feed Study of Codeine ............................... 230 

TABLE F3 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

at the 15-Month Interim Evaluation in the 2-Year Feed Study 

ofcodeine ................................................. 232 

TABLEF4Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice 

in the 14-Day Feed Study of Codeine ................................ 233 

TABLEF5 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice 

in the 13-Week Feed Study of Codeine ............................... 235 

TABLEF6Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice 

at the 15-Month Interim Evaluation in the 2-Year Feed Study 

ofcodeine ................................................. 237 
228 Codeine, NTP TR 455 

TABLEF1 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats in the 14-Day Feed Study of Codeinea 

0 PPm 1,562 ppm 3,125 ppm 6,250 ppm 12,500 ppm 25,000 ppm 

Male 

n 5 5 5 5 4 

Necropsy body wt 193 f 5 172 f 3** 171 f 4** 151 f 2** 119 f 4** 

Brain 

Absolute 1.719 f 0.017 1.802 f 0.111 1.724 f 0.030 1.654 f 0.028 1.618 f 0.017 

Relative 8.92 f 0.18 10.53 f 0.74* 10.13 f 0.30* 10.93 f 0.11** 13.64 f 0.42** 

Heart 

Absolute 0.633 f 0.018 0.598 f 0.021 0.570 f 0.015** 0.511 f 0.004** 0.415 f 0.012** 

Relative 3.28 f 0.07 3.49 f 0.11 3.36 f 0.15 3.38 f 0.05 3.49 f 0.08 

Absolute 

R. Kidney 

0.691 f 0.022 0.727 f 0.110 0.594 f 0.011 0.571 f 0.016 0.456 f 0.014** 

Relative 3.58 f 0.07 4.26 f 0.68 3.49 f 0.12 3.77 f 0.09 3.84 f 0.15 

Liver 

Absolute 8.199 f' 0.258 6.679 f 0.234** 6.596 f 0.152** 6.336 f 0.153** 5.163 f 0.276** 

Relative 42.46 f 0.42 38.90 f 0.95* 38.79 f 1.42* 41.87 f 0.68 43.29 f 0.78 

Absolute

Lung 

1.033 f 0.098 1.527 f 0.415 0.831 f 0.015 0.827 f0.008 0,658 f 0.020 

Relative 5.35 f 0.50 8.78 f 2.28 4.89 f 0.15 5.47 f 0.13 5.53 f 0.07 

R. Testis 

Absolute 1.007 f 0.062 0.813 f 0.064*' 0.743 f 0.040** 0.814 f 0.049** 0.398 f 0.062** 

Relative 5.22 f 0.32 4.81 f 0.41' 4.35 f 0.18 5.37 f 0.26 3.31 f 0.44** 

Absolute 

Thymus 

0.383 f 0.024 0.420 f 0.011 0.411 f 0.012 0.322 f 0.030 0.096 f 0.016** 

Relative 1.98 f 0.11 2.45 f 0.07 2.41 f 0.11 2.13 f 0.20 0.80 f 0.11** 
Organ Weight Analyses 229 

TABLEF1 

Organ Weights and Organ-Weight-to-Body-Weight Ratiosfor Rats in the 14-Day Feed Study of Codeine (continued) 

0 PPm 1,562 ppm 3,125 ppm 6,250 ppm 12,50025,OOO ppmppm 

Female 

n 5 5 5 4 2 

Necropsy body wt 141 f 3 140 f 4 126 f 5** 128 f 3* 112 f 7** 

Brain 

Absolute 1.627 f 0.019 1.655 f 0.021 1.584 f 0.037 1.605 f 0.009 1.568 f 0.138 

Relative 11.53 f 0.12 11.86 f 0.15 12.67 f 0.56* 12.61 f 0.29* 13.94 f 0.40** 

Heart 

Absolute 0.461 f 0.012 0.492 f 0.015 0.448 f 0.027 0.449 f 0.009 0.410 f 0.026 

Relative 3.27 f 0.06 3.52 f 0.07 3.55 f 0.11 3.53 f 0.113.65 f 0.02 

R. Kidney 

Absolute 0.501 f 0.020 0.512 f 0.025 0.459f 0.019 0.476 f 0.012 0.480 0.033 

Relative 3.54 f 0.08 3.66 f 0.11 3.66 f 0.09 3.74 f 0.034.27 f OM** 

Liver 

Absolute 5.572 f 0.194 5.915 f 0.217 4.981 f 0.292 ,5593f 0.123 5.338 f 0.059 

Relative 39.43 f 0.79 42.32 f 0.68 39.54 f 1.31 43.88 f 0.61** 47.68 f 2.32** 

Luns 

Absolute 0.723 f 0.035 0.761 f 0.026 0.709 f 0.029' 0.731 f 0.033 0.663 f 0.066 

Relative 5.12 f 0.19 5.45 f 0.14 5.76 f 0.19' 5.74 f 0.245.89 f 0.23 

Thymus 

Absolute 0.350f 0.015 0.340f 0.011 0.265 f 0.061 0.259 f 0.016 0.124 f 0.044** 

Relative 2.48 f 0.10 2.44 f 0.07 2.05 f 0.42 2.04 f 0.151.08 f 0.32* 

* Significantly different(PsO.05)from the control group by Williams' or Dunuea's test 

** PSO.01

* Organ weights and body weights are given in grams; organ-weight-to-body-weight ratios are given as mg organ weightlg body weight 

(mean f standard error). 

No data calculated due to 100% mortality in this exposure group 

n=4 
230 Codeine, NTP TR 455 

TABLEF2 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats in the 13-Week Feed Study of Codeinea 

0 PPm 390ppm781ppm 1,562 ppm 3,125 ppm 6,250 ppm 

Male 

n 10 9 10 10 10 10 

Necropsy body wt 370 f 7 343 f 7* 335 f 6** 316 f 7** 300 f 6** 292 f 9** 

Adrenal Gland 

Relative 

Absolute 

0.12 f 0.00 

0.045 f 0.002 0.052 f 0.002* 

0.15 f 0.01** 

0.058 f 0.002** 

0.17 f 0.01** 0.19 f om** 

0.059 f 0.002** 0.068 f 0.002** 

0.23 f 0.00** 

0.069 f 0.003** 

0.23 f 0.01** 

Brain 

Absolute 1.992 f 0.013 1.960 f 0.027 1.933 f 0.034 1.946 f 0.021 1.946 f 0.0261.980 f 0.023 

Relative 5.40 f 0.09 5.74 f 0.12 5.78 f 0.10* 6.19 f 0.12** 6.49 f 0.11**6.83 f 0.17** 

Heart 

Absolute 1.063 f 0.033 0.934 f 0.020**0.940 f 0.015** 0.851 f 0.020** 0.875 f 0.031**0.871 f 0.035** 

Relative 2.88 f 0.10 2.73 f 0.082.81 f 0.05 2.70 f 0.03 2.91 'f0.062.98 f 0.04 

Absolute 

R.Kidney 

1.263 f 0.033 1.103 f 0.030**1.043 f 0.031** 0.968 f 0.021** 0.990f 0.024** 0.963 f 0.038** 

Relative 3.42 f 0.08 3.22 f 0.09 3.11 f 0.06** 3.07 f 0.04** 3.30 f 0.06 3.29 f 0.05 

Liver 

Absolute 13.828 f 0.356 11.453 f 0.426** 10.466 f 0.331** 9.306 f 0.289** 9.538 f 0.402** 9.537 f 0.593** 

Relative . 37.36 4 0.47 33.36 f 0.76** 31.22 f 0.66** 29.44 f 0.45** 31.65 f 0.80** 32.45 f 1.17** 

Absolute

Lung 

1.380 f 0.046 1.273 f 0.023 1.187 f 0.044** 1.168 f 0.049** 1.162 f 0.035**b 1.139 f 0.044** 

Relative 3.73 f 0.11 3.73 f 0.103.54 f 0.11 3.70 f 0.15 3.90 f O.Ottb 3.91 f 0.10 

Absolute 

Spleen 

0.777 f 0.016 0,720 f 0.0230.709 f 0.018* 0.636 f 0.017** 0.611 f 0.027**0.568 f 0.017** 

Relative 2.10 f 0.01 2.10 f 0.05 2.11 f 0.02 2.02 f 0.04 2.03 f 0.06 1.95 f 0.03* 

R. Testis 

Absolute 1.430 f 0.020 1.472 f 0.0151.431 f 0.034 1.369 f 0.039 1.402 f 0.021.416 f 0.032b 

Relative 3.87 f 0.04 4.30 f 0.07**4.28 f 0.11** 4.34 f 0.11** 4.67 f OM** 4.86 f 0.12**b 

Absolute 

Thymus 

0.303 f 0.017 0.287 f 0.0140.285 & 0.014 0.238 f 0.016** 0.167 f 0.012**0.198 f 0.007** 

Relative 0.82 f 0.05 0.84 f 0.040.85 f 0.04 0.76 f 0.06 0.55 f 0.04** 0.69 f 0.04** 
Organ Weight Analyses 231 

TABLEF2 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats in the 13-Week Feed Study of Codeine (continued) 

Female 

n 10 10 10 10 10 10 

Necropsy body wt 194 f 5 189 f 3 192 f 3 181 f 3* 175 f 5** 170 f 3** 

Adrenal Gland 

Absolute 0.051 f 0.002 0.049 f 0.002 0.054f 0.003 0.055 f 0.001 0.061f 0.003** 0.064f 0.002** 

Relative 0.26 f 0.01 0.26 f 0.01 0.28 f 0.01 0.31 f 0.01** 0.35 f 0.01** 0.38f 0.01** 

Brain 

Absolute 1.771 f 0.020b 1.790f 0.018 1.812 f 0.023 1.809 f 0.019 1.810f 0.018 1.839 f 0.015 

Relative 9.05 f 0.16b 9.48 f 0.17 9.44 f 0.16 10.01 f 0.16** 10.44f 0.27** 10.85f 0.16** 

Heart 

Absolute 0.585 f 0.013 0.573 f 0.011 0.590 f 0.012 0.571 f 0.012 0.613f 0.019 0.586 f 0.009 

Relative 3.03 f 0.04 3.03 f 0.04 3.07 f 0.06 3.16 f 0.05 3.52 f 0.06** 3.46 f 0.05** 

R.Kidney 

Absolute 0.668 f 0.014 0.636 f 0.017 0.647 f 0.009 0.644 f 0.012 0.649 f 0.021 0.643 f 0.009 

Relative 3.45 f 0.04 3.36.f 0.07 3.37 f 0.05 3.56 f 0.06 3.72 f 0.07** 3.79 f 0.04** 

Liver 

Absolute 6.401 f 0.212 6.063 f 0.142 6.337 f 0.160 5.904 f 0.187 6.002f 0.336 6.325 f 0.228 

Relative 33.00 f 0.45 32.08 f 0.71 32.94 f 0.57 32.55 f 0.62 34.16 f 0.97 37.19f 0.98** 

Lung

Absolute 0.917 f 0.030 0.883 f 0.032b 0.889 f 0.025 0.880 f 0.011 0.843f 0.017b 0.881 f 0.027 

Relative 4.74 f 0.14 4.67 f 0.17b 4.62f 0.12 4.87 f 0.10 4.91 f O.lOb 5.20f 0.17 

Spleen 

Absolute 0.482 f 0.016 0.448 f 0.012 0.477 f 0.014 0.435 f 0.010* 0.416f 0.016** 0.400f 0.017** 

Relative 2.48 f 0.05 2.37 f 0.05 2.48 f 0.04 2.40 f 0.03 2.38 f 0.022.35 f 0.07 

Thymus 

Absolute 0.221 f 0.003 0.217 f 0.012 0.214f 0.008 0.216 f 0.006 0.186f 0.013** 0.178f 0.005** 

Relative 1.15 f 0.03 1.15 f 0.07 1.11 f 0.03 1.19 f 0.03 1.06 f 0.061.05 f 0.03 

* Significantly different (PSO.05)from the control group by Williams’ or Dunnett’s test 

** PSO.01 

a Organ weights and body weights are given in grams; organ-weight-to-body-weightratios are given as mg organ weightlg body weight 

(mean f standarderror). 

n=9 
232 Codeine, NTP TR 455 

TABLEF3 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats at the 15-Month Interim Evaluation 

in the 2-YearFeed Study of Codeine 

0 PPm 400 ppm 800 ppm 1,600 ppm 

Male . 

n 10 10 10 10 

Necropsy body wt 478 f 6 448 f 6* 432 f 9** 392 f 12** 

Adrenal Gland 

Absolute 0.059 f 0.004 0.061 f 0.002 0.071 f 0.003* 0.076 f 0.005** 

Relative 0.12 f 0.01 0.14 f 0.00 0.17 f 0.01** 0.19 f 0.01** 

R. Kidney 

Absolute 1.610 f 0.042 1.617 f 0.042 1.560f 0.032 1.577f 0.042 

Relative 3.36 f 0.07 3.61 f 0.10 3.62 f 0.11 4.04 f om** 

Liver 

Absolute 16.732 f 0.281 14.531 f 0.442** 13.384 f 0.384** 12.223 f 0.568** 

Relative 34.98 f 0.26 32.44 f 0.99* 31.08 f 1.05** 31.13 f O.%** 

Female 

n 10 10 9 9 

Necropsy body wt 334 f 3 313 f 8* 313 f 5* 283 f 7** 

Adrenal Gland 

Absolute 0.065 f 0.003 0.061 f 0.003 0.066 f 0.003 0.059f 0.004 

Relative 0.19 f 0.01 0.19 f 0.01 0.21 f 0.01 0.21 f 0.01 

R.Kidney 

Absolute 1.079f 0.027 1.164f 0.125 1.077 f 0.021 0.993 f 0.029 

Relative 3.23 f 0.07 3.72 f 0.38 3.45 f 0.05 3.52 f 0.05 

Liver 

Absolute 9.487 f 0.159 10.038f 0.246 9.790f 0.169 8.922 f 0.419 

Relative 28.43 f 0.48 32.17 f 0.96** 31.35 f O.M* 31.53 f 1.20* 

* Significantly different(PsO.05) from the control group by Williams’ or Dunnett’s test 

** PSO.01 

a Organ weights and body weights are given in grams; organ-weight-to-body-weightratios are given as mg organ weighflg body weight 

(mean f standard error). 
Organ Weight Analyses 233 

TABLEF4 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 14-Day Feed Study of Codeinea 

0 PPm781ppm 1,562 ppm 3,125 ppm 6,250 ppm 12,500 ppm 

Male 

n 5 5 5 5 5 5 

Necropsy body wt 25.2 f 0.9 25.7f 1.0 25.9 f 1.2 24.4 f 1.024.7 f 0.8 22.8 f 0.7 

Brain 

Absolute 0.450f 0.016 0.448 f 0.014 0.474f 0.006 0.466 f 0.011 0.457 f 0.009 0.445 f 0.010 

Relative 17.83 f 0.20 17.45 f 0.50 18.41 f 0.79 19.19 f 0.44 18.54f 0.47 19.62 f 0.92 

Heart 

Absolute 0.114f 0.005 0.125f 0.008 0.135 f 0.009 0.121f 0.006 0.122 f 0.006 0.119 f 0.005 

Relative 4.52 f 0.12 4.88 f 0.21 5.22 f 0.17 4.97 f 0.104.93 f 0.16 5.24 f 0.28* 

Absolute 

R. Kidney 

0.202 f 0.009 0.210f 0.009 0.207 f 0.007 0.203 f 0.012 0.192 f 0.010 0.157f 0.006** 

Relative 8.01 f 0.22 8.14 f 0.17 7.99 f 0.17 8.34f 0.34 7.76 f 0.18 6.94f 0.43* 

Liver 

Absolute 1.366 f 0.069 1.299 f 0.038 1.249 f 0.059b 1.162 f 0.060** 1.092f 0.046** 0.947 f 0.016** 

Relative 54.07 f 1.31 50.59 f 0.80* 49.37 f 47.65f 0.80** 44.14 f 0.46** 41.67 f 1.47** 

Absolute 

Lung 

0.184f 0.016 0.169f 0.010 0.189 f 0.010 0.160f 0.003 0.165 f 0.006 0.153 f 0.006 

, Relative 7.25 f 0.52 6.57f 0.32 7.30 f 0.27 6.62 f 0.26 6.69 f 0.03 6.75 f 0.33 

R. Testis 

Absolute 0.097 f 0.005 0.096 f 0.005 0.103 f 0.006 0.098 f 0.003 0.133 f 0.031 0.096f 0.004 

Relative 3.83 f 0.13 3.74 f 0.06 3.98f 0.09 4.05 f 0.095.47 f 1.38 4.23 f 0.27 

Absolute 

Thymus 

0.038 f 0.008 0.048 f 0.009 0.055 f 0.002 0.045 f 0.006 0.043 f 0.003 0.034 f 0.004 

Relative 1.54f 0.34 1.88 f 0.39 2.14 f 0.18 1.89 f 0.271.76 f 0.16 1.50f 0.19 
234 Codeine, NTP TR 455 

TABLEF4 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 14-Day Feed Study of Codeine (continued) 

0 PPm ppm781 1,562 ppm 3,125 ppm 6,250 ppm 12,500 ppm 

Female 

n 5 5 5 5 5 5 

Necropsy body wt 18.7f 0.3 18.8 f 0.2 18.2 f 0.4 20.1 f 0.5* 19.0f 0.2 16.8f 0.3** 

Brain 

Absolute 0.444f 0.006 0.433 f 0,010 0.450 f 0.007 0.449 f 0.011 0.447f 0.008 0.450 f 0.005 

Relative 23.78 f 0.51 23.05 f 0.59 24.78 f 0.27 22.37 f 0.44 23.50 f 0.15 26.78 f 0.61** 

Heart 

Absolute 0.094f 0.003 0.093 f 0.004 0.093f 0.007 0.098 f 0.004 0.099f 0.004 0.084f 0.002 

Relative 5.01 f 0.11 4.94 f 0.16 5.09 f 0.28 4.86f 0.14 5.23 f 0.19 5.00 f 0.19 

R. Kidney 

Absolute 0.128 f 0.006 0.138 f 0.006 0.134 f 0.006 0.143 f 0.002 0.134f 0.002 0.119f 0.002 

Relative 6.87 f 0.29 7.32 f 0.26 7.36f 0.20 7.11 f 0.12 7.05 f 0.13 7.05 f 0.11 

Liver 

Absolute 0.941 f 0.039 0.943 f-0.026 0.861 f 0.036 0.954f 0.040 0.902 f 0.030 0.749 f 0.011** 

Relative 50.29 f 1.33 50.17 f 1.36 47.33 f 1.07 47.41 f 1.29 47.46 f 1.07 44.55f 0.89** 

Lung 

Absolute 0.168 f 0.007 0.154f 0.008 0.140f 0.004** 0.154f 0.004 0.147f 0.006 0.147f 0.006 

Relative 9.00f 0.25 8.19 f 0.45 7.70 f 0.23* 7.69f 0.27* 7.75 f 0.24* 8.75 f 0.41 

Thymus 

Absolute 0.055 f 0.006 0.063 f 0.003 0.062 f 0.001 0.061 f 0.002 0.062 f 0.004 0.056f 0.003 

Relative 2.98 f 0.36 3.34 f 0.15 3.44f 0.11 3.06 f 0.11 3.27 f 0.19 3.34 f 0.19 

* Significantly different(PsO.05) fromthecontrolgroupbyWilliams’ or Dunnett’stest 

** PSO.01 

a Organ weights and body weights are given in grams; organ-weight-to-body-weightratios are given as mg organ weighug body weight 

(mean f standarderror). 

n=4 
Organ Weight Analyses 235 

TABLEF5 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 13-Week Feed Study of Codeinea 

0 PPm 390 ppm 781 ppm 1,562 ppm 3,125 ppm 6,250 ppm 

Male 

n 10 10 10 10 8 10 

Necropsy body wt 30.8 f 0.5 31.3 f 0.431.8 f 0.539 f 0.631 f 0.5 29.7 f 0.4 

Adrenal Gland 

Absolute 0.005 f 0.001 0.008 f 0.001**0.006 f 0.Mx) 0.007 f 0.001 0.006 f 0.0010.006 f o*ooO 

Relative 0.17 f 0.02 0.24 f 0.02*0.18 f 0.000.21 f 0.020.21 f 0.02 0.21 f 0.01 

Brain 

Absolute 0.460f 0.006 0.464 f 0.005 0.458 f 0.005 0.462 f 0.005 0.466 f 0.003 0.467 f 0.004 

Relative 14.99 f 0.24 14.81 f 0.25 14.44 f 0.21 14.97 f 0.25 15.50 f 0.27 15.74 f 0.18* 

Heart 

Absolute 0.140 f 0.005 0.143 f 0.003 0.138 f 0.004 0.142 f 0.004 0.145 f 0.004 0.142 f 0.003 

Relative 4.54 f 0.11 4.55 f 0.07 4.36 f 0.10 4.58 f 0.08 4.81 f 0.10 4.78 f 0.06 

.Absolute 

R.Kidney 

0.273 f 0.006 0.286 f 0.0080.259 f 0.0060.257 f 0.0060.251 f 0.007*0.232 f 0.005** 

Relative 8.89 f 0.14 9.13 f 0.238.15 f 0.18*8.32 f 0.13*8.34 f 0.13*7.80 f 0.15** 

Liver 

Absolute 1.463 f 0.041 1.543 f 0.042 1.549 f 0.027 1.535 f 0.029 1.477 f 0.049 1.461 f 0.027 

Relative 47.51 f 0.67 49.21 f 1.08 48.75 f 0.78 49.71 f 0.96 49.04 f 1.17 49.26 f 0.91 

Lung 

Absolute 0.179 f 0.005 0.182 f 0.005 0.179 f 0.004 0.190 f 0.007 0.183 f 0.008 0.171 f 0.003 

Relative 5.83 f 0.15 5.80 f 0.17 5.63 f 0.12 6.16 f 0.20 6.11 f 0.26 5.77 f 0.16 

Absolute 

Spleen 

0.083 f 0.009 0.098 f 0.005 0.092 f 0.003 0.089 f 0.006 0.099 f 0.011 0.094 f 0.005 

Relative 2.69 f 0.26 3.15 f 0.18 2.89 f 0.09 2.84 f 0.16 3.30 f 0.37 3.15 f 0.14 

R. Testis 

Absolute 0.110 f 0.002 0.111 f 0.002 0.114 f 0.002 0.112 f 0.005 0.110 f 0.003 0.104 f O.Wb 

Relative 3.58 f 0.07 3.55 f 0.08 3.60 f 0.06 3.60 f 0.12 3.65 f 0.09 3.51 f O.OSb 

Absolute 

Thymus 

0.029 f 0.003 0.026 f 0.002 0.020 f 0.0030.029 f 0.0030.028 f 0.0030.033 f 0.003 

Relative 0.96 f 0.08 0.82 f 0.070.63 f 0.10*0.95 f 0.080.93 f 0.091.12 f 0.08 
236 Codeine, NTP TR 455 

TABLEF5 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 13-Week Feed Study of Codeine (continued) 

0 PPm 390 ppm 781 ppm ppm 1,562 3,1256,250 ppmpm

+ 

Female 

n 10 9 10 10 

Necropsy body wt 23.9 f 0.4‘25.2 f 0.4 24.4 f 0.4238 f 0.3 24.2 f 0.321.3 f 0.6** 

Adrenal Gland 

Absolute 0.010 f o.oO0 0.011 f 0.001 0.011 f 0.001 0.010 f 0.001 0.010 f 0.001 0.009 f 0.001 

Relative 0.41 f 0.02 0.46 f 0.03 0.44f 0.03 0.44f 0.03 0.41 f 0.03 0.43 f 0.03 

Bliiin 

Absolute 0.472 f 0.004 0.476 f 0.004 0.464 f 0.005 0.468 f 0.005 0.475 f 0.007 0.473 f 0.005 

Relative 19.78 f 0.40 18.94 f 0.34 19.11 f 0.45 19.66 f 0.29 19.64 f 0.26 22.32 f 0.57** 

Heart 

Absolute 0.109 f 0.002 0.115 f 0.002 0.107 f 0.003 0.112 f 0.002 0.109 f 0.002 0.104 f 0.003 

Relative 4.57 f 0.12 4.59 f 0.09 4.40 f 0.08 4.71 f 0.10 4.51 f 0.08 4.91 f 0.13 

Absolute 

R. Kidney 

0.177 f 0.003 0.193 f 0.005 0.180 f 0.005 0.177 f 0.004 0.177 f 0.003 0.166 f 0.003 

Relative 7.42 f 0.13 7.64 f 0.14 7.40 f 0.27 7.40 f 0.12 7.31 f 0.13 7.80 f 0.18 

Liver 

Absolute 1.103 f 0.014 1.217 f 0.023** 1.152 f 0.027 1.106 f 0.030 1.145 f 0.022 1.065 f 0.025 

Relative 46.19 f 0.58 48.29 f 0.66 47.29 f 1.03 46.34 f 0.91 47.28 f 0.66 50.07 f 0.49** 

Absolute

Lug 

0.159 f 0.005 0.161 f 0.006 0.164 f 0.005 0.159 f 0.006 0.168 f 0.004 0.163 f 0.005 

Relative 6.64 f 0.23 6.38 f 0.17 6.75 f 0.23 6.66 f 0.20 6.93 f 0.14 7.66 f 0.19** 

Spleen 

Absolute 0.073 f 0.002 0.078 f 0.003 0,070 f 0.0030.070 f 0.0020.076 f 0.0030.070 f 0.006 

Relative 3.05 f 0.073.09 f 0.08 2.87 f 0.112.93 f 0.07 3.12 f 0.13 3.24 f 0.22 

Thymus 

Absolute 0.031 f 0.0030.032 f 0.0030.029 f 0.0040.025 f 0.0020.025 f 0.0030.022 f 0.003 

Relative 1.27 f 0.141.25 f 0.111.20 f 0.151.07 f 0.101.03 f 0.12 1.04 f 0.14 

* Significantly different(PSO.05) from the control group by Williams’ or Dunnett’s test 

** PSO.01 

a Organ weights and body weights are given in grams; organ-weight-to-body-weightratios are given as mg organ weightlg body weight 

(mean f standard error). 

n=9 

n=lO 
Organ Weight Analyses 237 

TABLEF6 

Organ Weights and Organ-Weight-to-Body-Weight Ratiosfor Mice at the 15-Month Interim Evaluation 

in the 2-Year Feed Study of Codeine 

Male 

n 

Necropsy body wt 

Adrenal Gland 

Absolute 

Relative 

R.Kidney 

Absoluie 

Relative 

Liver 

Absolute 

Relative 

n 

Necropsy body wt 

Adrenal Gland 

Absolute 

Relative 

R.Kidney 

Absolute 

Relative 

Liver 

Absolute 

Relative 

* Significantly different (P

** PSO.01 

0 PPm 750 ppm 1,500 ppm 

9 10 9 

51.3 f 0.6 52.6 f 0.7 48.1 f 1.3* 

0.008 f 0.001 0.007 f 0.001 0.006 f 0.001 

0.16 f 0.01 0.14 f 0.02 0.13 f 0.01 

0.448 f 0.012 0.427 f 0.011 0.382 f 0.009** 

8.74 f 0.23 8.12 f 0.15 7.97 f 0.20 

2.443 f 0.093 2.305 f 0.084 2.230 f 0.193 

47.64 f 1.78 43.82 f 1.41 46.20 f 3.38 

10 9 9 

55.1 f 1.8 56.0 f 1.5 50.9 f 2.8 

0.011 f 0.001 0.015 f 0.001 0.013 f 0.001 

0.21 f 0.02 0.26 f 0.03 0.27 f 0.03 

0.302 f 0.010 0.316 f 0.007 0.2% f 0.007 

5.51 f 0.17 5.66 f 0.15 5.92 f 0.24 

2.110 f 0.227 2.079 f 0.062 1.987 f 0.081 

38.14 f 3.60 37.15 f 0.97 39.34 f 1.00 

50.05) from the control group by Williams’ or Dunnett’s test 

3,000 ppm 

10 

43.9 f 1.3** 

0.009 f 0.001 

0.20 f 0.02 

0.362 f 0.011** 

8.27 f 0.27 

2.048 f 0.181 

46.37 f 3.35 

9 

45.8 f 1.8** 

0.013 f 0.001 

0.29 f 0.02 

0.274f 0.007* 

6.04f 0.21 

1.886 f 0.089 

41.32 f 1.48 

a Organ weights and body weights are given in grams; organ-weight-to-body-weightratios are given as mg organ weightlg body weight 

(mean f standard error). 
239 

APPENDIX G 

HEMATOLOGY, CLINICAL CHEMISTRY, 

AND URINALYSIS RESULTS 

TABLE G1 Hematology, Clinical Chemistry, and Urinalysis Data for Rats 

in the13-WeekFeedStudy of Codeine . . ... ..... . ... . . . . . . . . . . . . ... 240 

TABLE 62 Hematology,ClinicalChemistry,andUrinalysisData for Mice 

in the 13-Week FeedStudy of Codeine .. . . . . . . ... . . . . . . . . . . . . ...... 242 
240 Codeine, NTP TR 455 

TABLEG1 

Hematology, Clinical Chemistry, and Urinalysis Data for Rats in the 13-Week Feed Study of Codein@ 

390 ppm 781ppm 1,562 ppm 3,125 ppm 6,250 ppm 

Hematology 

n 10 9 10 10 10 9 

Reticulocytes (106/pL)0.27 

Erythrocytes (106/pL)8.92 

Hemoglobin (g/dL) 

Hematocrit (%) 

f 0.020.24 

f 0.138.51 

44.8 f 0.6 

16.2 f 0.1 

f 0.02 

f 0.08* 

45.3 f 0.4 

16.5 f 0.1 

0.20 f 0.01* 

8.41 f 0.10** 

16.5 f 0.1 

45.5 f 0.7 

0.19 f 0.02* 

8.41 f 0.05** 

16.6 f 0.1** 

45.9 f 0.2 

0.22 f 0.03* 

8.24 f 0.24** 

16.2 f 0.4 

45.4 f 1.2 

0.20 f 0.01** 

8.31 f 0.09** 

16.3 f 0.2 

45.7 f 0.5 

Nucleated erythrocytes 

Meancellhemoglobin (pg) 18.1 f 0.2 

Meancellvolume (a) 50.3 f 0.353.1 

(10~1~~) 0.01 f 0.010.04

, 19.4 f 0.2** 

f 0.4** 

f 0.02 

19.6 f 0.2** 

54.0 f 0.4** 

0.07 f 0.02 0.04 f 0.02 

19.8 f 0.1** 

54.6 f 0.2** 

19.7 f 0.1** 

55.1 f 0.2** 

0.05 f 0.02 

19.6 f 0.2** 

55.0 f 0.2** 

0.05 f 0.03 

Mean cell hemoglobin 

Leukocytes (103/pL)6.78 

Platelets (~o~/~L) 

concentration(g/dL) 

f 0.346.66 

540.5 f 14.8531.4 

36.1 f 0.436.4 

f 0.33 

f 13.8 

f 0.4 

6.29 f 0.24 

503.8 f 10.Sb 

36.3 f 0.6 

6.05 f 0.37 

498.0 f 9.5* 

36.2 f 0.3 

6.56 f 0.16 

487.4 f 9.6** 

35.7 f 0.2 

5.44 f 0.27** 

471.7 f 14.9** 

35.7 f 0.3 

Segmented neutrophils 

Eosinophils (103/pL)0.13 

Monocytes (103/pL) . 

Lymphocytes (103/pL)5.24 

(10~1~~) 

f 0.030.07 

0.25 f 0.06 

f 0.235.11 

1.16 f 0.11.11 

f 0.01 

0.36 f 0.05 

f 0.24 

f 0.12 

0.12 f 0.04 

0.30 f 0.04 

4.89 f 0.20 

0.96 f 0.05 

0.07 f 0.02 

0.21 f 0.03 

4.62 f 0.23 

1.14 f 0.16 1.31 f 0.13 

4.86 f 0.21 

0.29 f 0.04 

0.09 f 0.03 

0.91 f 0.08 

4.20 f 0.32** 

0.26 f 0.04 

0.06 f 0.02 

Clinical Chemistry 

n 5 4 5 5 5 5 

Albumin (g/dL) 

Total protein (g/dL) 

Glucose (mgldL) 

Creatinine (mg/dL) 

Urea nitrogen (mg/dL) 

4.5 f 0.1 

6.3 f 0.1 

185 f 1 

0.56 f 0.02 

20.4 f 1.3 

4.8 f 0.1 

6.9 f 0.2 

186 f 13 

0.63 f 0.06 

21.5 f 0.9 

4.7 f 0.0 

6.3 f 0.1 

168 f 4* 

0.50 f 0.06 

21.0 f 1.5 

4.7 f 0.1 

6.3 f 0.2 

178 f 9 

0.58 f 0.04 

20.0 f 1.8 

4.6 f 0.1 

6.4 f 0.2 

156 f 4** 

0.54 f 0.02 

20.2 f 1.8 

4.7 f 0.1 

6.2 f 0.3 

157 f 5** 

0.56 f 0.02 

23.4 f 1.5 

Alanine aminotransferase 

(IU/L) 73 f 6 65 f 3 57 f 3* 52 f 6* 56 f 4* 39 f 3** 

aminotransferase (IU/L) 

Aspartate

WL) 

Alkaline phosphatase 

248 f 14 

143 f 12 102 f 13 

258 f 24 210 f 11 

103 f 7 

192 f 13* 

102 f 6* 

203 f 14* 

117 f 7 

167 f 16** 

97 f 8* 

Urinalysis 

n 10 9 10 10 10 10 

Specificgravity 

Vobw (a16 hr) 

1.036 f 0.0061.048 

‘5.1 f 0.9 

f 0.005 

3.4 f 0.9 

1.030 f 0.002 

5.1 f 0.5 

1.040 f 0.002 

4.2 f 0.4 5.0 f 0.7 

1.043 f 0.0031.043 

4.9 f 0.7 

f 0.003 
Hematology, Clinical Chemistry, and Urinalysis 241 

TABLEG1 

Hematology, Clinical Chemistry, and Urinalysis Data for Rats in the 13-WeekFeed Study of Codeine (continued) 

0 PPm 390ppm . ppm 7811,562 ppm 3,125 ppm 6,250 ppm 

Female 

Hematology 

n 10 9 10 10 10 10 

Hematocrit (R) 43.6 f 0.6 45.3 f 0.7 43.9 f 0.6 44.3 f 0.6 43.4 f 0.6 44.2 f 0.5 

Hemoglobin16.1 (gIdL) f 0.1 16.5 f 0.2 16.1 f 0.1 16.1 f 0.2 15.9 f 0.1 16.0 f 0.1

Erythrocytes(1061pL) . 8.01 f 0.10’ 8.26 f 0.16 7.96 f 0.12 8.06 f 0.11 7.80 f 0.13 7.85 f 0.10 

Reticulocytes(1061pL)0.20 f 0.02 0.22 f 0.01 0.23 f 0.02 0.19 f 0.01 0.21 f 0.02 0.16 f 0.01 

Nucleated erythrocytes 

(10~1~~) 0.01 f 0.01 0.01 f 0.01 0.00 f 0.00 0.01 f 0.01 0.02 f 0.01 0.04 f 0.02b 

Meancellvolume (fi) 54.5 f 0.2 55.0 f 0.3 55.3 f 0.3* 54.9 f 0.3 55.5 f 0.4* 56.4 f 0.3** 

Mean cell hemoglobin @g) 20.2 f 0.3 20.1 f 0.2 20.3 f 0.3 20.0 f 0.3 20.4 f 0.3 20.4 f 0.2 

Mean cell hemoglobin 

concentration 37.0 (gIdL) f 0.6 36.5 f 0.3 36.8 f 0.5 36.4 f 0.5 36.7 f 0.4 36.3 f 0.3

Platelets (10~1~~) 562.4 f 28.5 591.0 f 17.8 589.0 f 13.8 553.4 f 12.4 515.5 f 12.1 578.8 f 18.5 

Leukocytes (103/pL) 5.64 f 0.29 5.50 f 0.32 5.94 f 0.42 4.64 f 0.21* 4.41 f 0.22** 3.76 f 0.26**b

Segmented neutrophils 

(10~4~) 1.13 f 0.12 1.03 f 0.07 1.33 f 0.14 0.79 f 0.11 0.71 f 0.07* 0.64 f 0.06**b 

Lymphocytes(103/pL)4.32 f 0.24 4.18 f 0.30 4.36 f 0.33 3.65 f 0.13 3.53 f 0.20* 2.95 f 0.25**b 

Monocytes 0.15 (1031pL) f 0.03 0.20 f 0.03 0.22 f 0.04 0.14 f 0.02 0.12 f 0.02 0.12 f 0.02b

~ o ~ i n ~ p m( 1 0 ~ 1 ~ ~ ) 0.05 f 0.01 0.07 f 0.02 0.03 f 0.01 0.07 f 0.02 0.05 f 0.01 0.05 f O.Olb

Cliical Chemistry 

n 5 5 5 5 5 5 

Urea nitrogen (mgIdL) 18.4 f 0.7 20.2 f 1.2 21.8 f 1.5 19.4 f 1.6 17.8 f 0.7 21.8 f 0.6 

Creatinine (mgIdL) 0.40 f 0.04’ 0.46 f 0.08 0.42 f 0.05 0.46 f 0.05 0.44 f 0.02 0.48 f 0.02 . 

Glucose (mgIdL) 169 f 4 159 f 4 147 f 4* 154 f 6 168 f 6 156 f 5 

Total protein (gIdL) 5.6 f 0.1’ 5.5 f 0.1 5.8 f 0.2 5.7 f 0.2 5.6 f 0.1 5.9 f 0.1 

Albumin (gIdL) 4.4 f 0.1c 4.3 f 0.0 4.5 f 0.1 4.6 f 0.1 4.4 f 0.1 4.8 f 0.1 

Alanine aminotransferase 

(IUIL) 52 f 4 58 f 5 53 f 3 49 f 5 50 f 3 42 f 4 

Alkaline phosphatase 

(IuW 

Aspartate 

216 f 18 197 f19112 f 11 216 f 10 260 f 21 215 f 14 

aminotransferase (IUIL) 97f7 103 f 2 88 f 4 93 f 4 104 f 4 83 f 7 

Urinalysis 

n 9 10 10 10 10 10 

Volume (mLI16 hr) 

Specific gravity 

3.1 f 0.6 

1.027 f 0.004 

2.5 f 0.419 

1.035 f 0.0031.039 

f 0.2 

f 0.003*b 

1.8 f 0.433 

1.050 f 0.005**1.045 

f 1.4 

f 0.005** 

2.7 f 0.6 

1.044 f 0.003** 

* Significantly different (PSO.05) from the control group by Dun” or Shirley’s test 

** P zso.01 

a Mean f standarderror.Statisticaltestswereperformedonunroundeddata. 

n=9 ‘ n=4 
242 Codeine, NTP TR 455 

TABLE62 

Hematology, Clinical Chemistry, and Urinalysis Data for Mice in the 13-Week Feed Study of Codeinea 

0 PPm 390ppm781ppm 1,562 ppm 3,125 ppm 6,250 ppm 

Male 

n 9 10 10 10 8 10 

Hematology 

Hematocrit (X) 46.8 f 0.8 46.2 f 0.9 45.8 f 0.6 48.1 f 1.0 45.7 f 1.1 46.0 f 0.8 

Hemoglobin(gIdL)15.7 f 0.2 15.7 f 0.3 15.5 f 0.3 16.2 f 0.4 15.7 f 0.4 15.6 f 0.3 

Erythrocytes(1061pL)9.32 f 0.15 9.20 f 0.19 9.18 f 0.12 9.36 f 0.14 9.10 f 0.14 8.98 f 0.12 

Reticulocytes(1061pL)0.24 f 0.02 0.25 f 0.02b 0.18 f 0.03 0.29 f 0.03 0.27 f 0.04 0.29 f 0.03 

Nucleated erythrocytes 

Mean cellhemoglobin(pg)16.8 f 0.2 

Meancellvolume (a) 50.3 f 0.3 

(10~4~) 0.01 f 0.01

17.1 f 0.2 

50.3 f 0.5 

0.01 f 0.01 

16.9 f 0.2 

50.0 f 0.3 

0.02 f 0.01 0.00 f 0.00 

51.3 f 0.5 

17.3 f 0.2 

0.04 f 0.04 

50.1 f 0.7 

17.2 f 0.2 

0.01 f 0.01 

51.3 f 0.3 

17.4 f 0.2 

Mean cell hemoglobin 

concentration(gIdL)33.5 f 0.3 34.1 f 0.2 33.7 f 0.3 33.7 f 0.2 34.4 f 0.3 33.9 f 0.2 

Leukocytes(103/pL)4.06 

Platelets (10~1~~) 

f 0.41 

893.6 f 35.5 

3.97 f 0.42 

964.1 f 45.4 

4.74 f 0.99 

873.0 f 61.2 976.9 f 40.9 

4.68 f 0.59 

904.6 f 39.1 

4.60 f 0.83 

939.7 f 46.1 

3.09 f 0.22 

Segmented neutrophils 

(~o~I~L) 1.01 f 0.23 1.31 f 0.18b 2.58 f 1.00 1.34 f O.Ub 1.74 f 0.66 0.86 f 0.08 

Monocytes(1031pL) 0.08 f 0.03 

Lymphocytes(1031pL) 2.84 f 0.24 

~o~in~~m (10~1~~) 0.12 f 0.03 0.13 f 0.03 

0.10 f 0.03 

2.15 f 0.23 

0.09 f 0.02b 

0.10 f 0.03 

1.92 f 0.34 2.64 f 0.27 

0.09 f 0.03 

0.12 f 0.03 

2.65 f 0.50 

0.05 f 0.03 

0.16 f 0.05 

2.09 f 0.20 

0.03 f 0.01 

0.11 f 0.02 

Clinical Chemistry 

Creatinine (mgIdL) 

Urea nitrogen (mgIdL) 

0.31 f 0.03 

20.6 f 1.1 22.1 f l.lb 

0.28 f 0.02b 0.29 f 0.03 

22.0 f 1.9 19.1 f 2.2 

0.28 f 0.03 

18.1 f 2.0 

0.27 f 0.04' 

26.8 f 1.1* 

0.34 f 0.04 

Glucose (mgIdL) 193 f 4 178 f 7 192 f 9 190 f 5 197 f 7 160 f 8* 

Albumin (gIdL) 

Total protein (g/dL) 

3.4 f 0.1 

4.7 f 0.1 

3.3 f 0.1 

4.8 f 0.1 

3.7 f 0.2 

4.8 f 0.1 4.7 f 0.1 

3.3 f 0.1 

4.8 f 0.2 

3.6 f 0.1 

4.8 f 0.2 

3.5 f 0.1 

Alanine aminotransferase 

(IUIL) 45 f 4 38 f 5 53 f 8 40f2 32 f 2 42 f 6 

(IUIL) 

Alkaline phosphatase 

61 f2 59 f 3 59 f 3 67 f 3 67 f 7 74 f 2** 

aminotransferase (IUIL) 

Aspartate 

134 f 28 89 f 24 109 f 20 94 f 17 63 f 11* 69 f 9 

Urinalysis 

n 9 10 10 10 6 8 

Volume (mLI16 hr) 

Specific gravity 

1.0 f 0.1 

1.045 f 0.004 1.045 f 0.002 

1.2 f 0.1 

1.032 f 0.003 

1.3 f 0.1 0.7 f 0.2 

1.045 f 0.006' 

0.9 f 0.2 

1.046 f 0.007 

0.8 f 0.2 

1.035 f 0.004' 
Hematology, Clinical Chemistry, and Urinalysis 243 

TABLE62 

Hematology, Clinical Chemistry, and Urinalysis Data for Mice in the 13-Week Feed Study of Codeine (continued) 

0 PPm 390 ppm 781 ppm 6,2503,125 ppmpmpm1,562 

Female 

n 9 10 10 10 10 10 

Hematology 

Hematocrit (X) 49.1 f49.31.1 f 1.0 51.8 f 1.0 49.8 f 0.7 51.3 f 0.8b 47.9 f 0.8 

Hemoglobin(gldL)16.7 f 0.217.0 f 0.2 17.4 f 0.3 17.1 f 0.2 17.4 f 0.3 16.0 f 0.3 

Erythrocytes(106/pL)9.81 f 0.16 9.68 f 0.21 10.14 f 0.17 9.77 f 0.07 9.87 f O.llb 9.45 f 0.13 

Reticulocytes(106/pL)0.19 f 0.03 0.25 f 0.05 0.26 f 0.03 0.23 f 0.03 0.24 f 0.03b 0.30 f 0.06 

Nucleated erythrocytes 

Meancellhemoglobin(pg) 17.0 f 0.2 

Meancellvolume (a) 50.2 f 0.5 

(~o~/~L) 0.00 f 0.00

17.7 f 0.4 

'51.0 f 0.3 

0.01 f 0.01b 

17.2 f 0.3 

50.9 f 0.5 

0.02 f 0.02 

17.5 f 0.2 

51.1 f 0.6 

0.00 f 0.00 

17.4 f 0.2b 

51.9 f 0.4b 

0.01 f 0.01 

16.9 f 0.1 

50.7 f 0.3 

0.00 f 0.00 

Mean cell hemoglobin 

concentration(g/dL)34.0 f 0.5 34.6 f 0.7 33.6 f 0.5 34.4 f 0.4 33.5 f 0.3b 33.3 f 0.2 

Leukocytes(103/pL)2.88 

Platelets (10~1~~) 

f 0.174.19 

720.3 f 22.6d782.4 

f 0.59b 

f 19.1 

3.08 f 0.54 

799.9 f 10.1*b 

3.26 f 0.32 

767.6 f 12.2b 

3.63 f 0.47 

786.8 f 16.4b 

3.25 f 0.36 

801.4 f 20.7 

Segmented neutrophils 

(~o~/~L) 0.37 f 0.06 0.69 f 0.13b 0.31 f 0.04 0.50 f 0.10 0.85 f 0.12* 0.61 f 0.12 

Monocytes (103/pL)0.03 

Lymphocytes(103/pL)2.41 

f 0.01 

f 0.153.35 

0.08 f 0.03b 

f 0.47b 

0.06 f 0.01 

2.65 f 0.51 

0.06 f 0.01 

2.60 f 0.30 

0.05 f 0.02 

2.67 f 0.36 

0.07 f 0.03 

2.50 f 0.29 

~ o ~ i n ~ ~ h i l ~( ~ o ~ / ~ L ) 0.060.02 0.07 f 0.02b 0.07 f 0.02 0.10 f 0.02 0.06 f 0.02 0.07 f 0.02 

Clinical Chemistry 

Urea nitrogen (mg/dL) 19.0 f 1.8 15.0 f 0.8 17.3 f 1.8b 13.3 f 1.0* 13.8 f 1.0* 16.6 f 3.9*' 

Creatinine (mg/dL) 0.31 f 0.02 0.28 f 0.02 0.25 f 0.03d 0.29 f O.Olb 0.34 f 0.02 0.26 f 0.02' 

Glucose (mg/dL) 191 f 3 194 f 4 187 f 8 194 f 10 188 f 6 199 f Sd 

Total protein (g/dL) 4.77 f 0.05 4.60 f 0.07 4.53 f 0.16 4.74 f 0.13 4.63 f 0.10 4.90 f 0.09e 

Albumin (g/dL) 3.7 f 0.1 3.8 f 0.1 3.6 f 0.2 3.9 f 0.1 3.9 f 0.1 4.1 f 0.1**' 

Alanine 

aminotransferase (IUIL) 44f6 52 f 12 42 f lob 27 f 3 29 f 3 42 f 10 

Alkaline phosphatase 

Aspartate 

(KJW 80 f 3 72 f 3 73 f 7 84 f 6 74 f 3 89 f Sb 

aminotransferase (IUIL) 98 f 1275 f 11 80 f 14b 67 f 5 71 f 1472 f gd 

Urinalysis 

n 10 10 10 8 9 8 

Volume (mL/16 hr) 0.4 f 0.1 0.5 f 0.1 0.3 f 0.0 0.5 f 0.1 0.4 f 0.1 0.4 f 0.1 

Specific gravity 1.036 f 0.005d 1.033 f 0.004b 1.036 f O.Wd 1.027 f 0.003 1.034 f 0.004' 1.028 f 0.006 

* Significantly different(PSO.05) from the control group by Dunn's or Shirley's test 

** PSO.01 

a Mean f standard error. Statisticaltestswereperformed on unroundeddata. 

n=9 

n=7 

n=8 

e n=6 
245 

AND ESTROUS CYCLE CHARACTERIZATION 

REPRODUCTIVE TISSUE EVALUATIONS 

APPENDIX H 

TABLE €I2 

TABLE H1 

SummaryofReproductiveTissueEvaluationsandEstrousCycleCharacterization 

SummaryofReproductiveTissueEvaluationsandEstrousCycleCharacterization 

for Rats inthe 13-Week Feed Study of Codeine .. . .. . . . . . .. . . . . . . . . . . . . . 

for Mice inthe 13-Week Feed Study of Codeine .. . . . . . . . . . . . .. . . .. . . . .. . 

246 

247 
246 Codeine, NTP TR 455 

TABLEH1 

Summary of Reproductive Tissue Evaluations and Estrous Cycle Characterizationfor Rats 

in. the 13-WeekFeed Study of Codeinea 

0 PPm 1,562 ppm 3,125 ppm 

Male 

n 10 10 10 

Weights (g) 

Necropsy body wt 370 f 7 316 f 7** 300 f 6** 

R. cauda 0.211 f 0.005 0.204 f 0.006 0.208 f 0.005 

R. epididymis 0.458 f 0.008 0.436 f 0.013 0.453 f 0.007 

R. testis 1.430 f 0.020 1.369 f 0.039 1.402 f 0.021 

Concentration 

(106/g caudaepididymaltissue) 628.9 f 55.3 629.1 f 40.8 552.6 f 43.8 

Motility (%) 74.16 f 3.20 83.94 f 1.75 70.89 f 4.32 

Epididymal spermatozoal parameters (per500 sperm) 

Normal 495.5 f 1.0 495.9 f 0.5 495.1 f 0.9 

Amorphous 0.500 f 0.307 0.200 f 0.133 0.700 f 0.213 

Excessive hook 0.800 f 0.249 0.500 f 0.224 1.Oo0 f 0.333 

No hook 2.40 f 0.54 2.50 f 0.50 2.90 f 0.66 

Pin-head 0.m f 0.m o.Oo0 f 0.m 0.m f o.Oo0 

Short-headed 0.800 f 0.291 0.900f 0.277 0.200 f 0,133 

Two tails or heads 0.m f o.Oo0 o.Oo0 f 0.m o.Oo0 f 0.m 

Abnormal (%) 0.900f 0.209 0.820 f 0.092 0.980 f 0.180 

Female 

n 10 10 10 

Necropsy body wt (9) 194 f 5 181 f 3* 175 f 5** 

Estrous cycle length (days) 4.88 f 0.30' 4.71 f 0.42d 4.60 f 0.24e 

Estrous stage (% of cycle) 

Diestrus 32.9 34.3 40.0 

Proestrus 22.9 21.4 20.0 

Estrus 30.0 28.6 22.9 

Metestrus 14.3 15.7 14.3 

Uncertain diagnoses 0.0 0.0 2.9 

* Significantly different (P40.05) from the control group by Williams' test (body weight) ** P40.01 a Data are presented as mean f standard error. 

n=9 

Estrous cycle longer than7 days or unclear in 2 of 10 animals 

Estrous cycle longer than7 days or unclear in 3 of 10 animals 

e Estrous cycle longer than 7 days or unclear in 5 of 10 animals 

6,250 ppm 

10 

292 f 9** 

0.190 f 0.008 

0.439 f 0.008 

1.416 f 0.032b 

566.3 f 61.1 

51.58 f 6.62 

493.7 f 0.8 

0.600f 0.221 

0.800 f 0.327 

3.90 f 0.67 

0.m f o.Oo0 

1.Oo0 f 0.258 

o.Oo0 f 0.m 

1.260 f 0.161 

10 

170 f 3** 

4.20 f 0.49e 

32.9 

28.6 

27.1 

11.4 

0.0 
Sperm Morphology and Vaginal Cytology Evaluations 247 

TABLEH2 

Summary of Reproductive Tissue Evaluations andEstrous Cycle Characterizationfor Mice 

in the 1fWeek Feed Study of Codeinea 

1,562ppm 3,125 ppm 6,250ppm 

Male 

n 9 10 7 9 

Weights (9) 

Necropsy body wt 30.7 f 0.6 30.9 f 0.6 30.1 f OSb 29.7 f 0.4' 

R. cauda 0.015 f 0.001 0.015 f 0.001 0.016 f O.OOlb 0.014 f 0.001'

R. epididymis 0.041 f 0.002 0.042 f 0.002 0.040 f 0.002b 0.040 f 0.002'

R. testis 0.110 f 0.002' 0,112 f 0.005 0.110 f 0.003b 0.104 f 0.002

Concentration 

(106/gcaudaepididymaltissue) 1,221 f 1,122 121 f 1,164 84 f 1,202 161 f 81' 

Motility (46) 78.44 f 5.94 65.34 f 5.19** 79.63 f 1.43b 66.20 f 6.55'

Epididymal spermatozoal parameters(per 500 sperm) 

Normal 494.4 f 493.6 1.0 f l.ld 494.0 f 1.0 493.4 f 1.0

Amorphous 2.11 f1.67 0.42 f 0.33d 2.29 f2.22 0.29 f 0.62

Blunt hook 0.667 f 0.289 0.556 f 0.377d0.429 f 0.2970.778 f 0.465 

Ba" 2.33 f3.33 0.65 f 0.41d 2.43 f2.44 0.69 f 0.50

pin-head 0.111 f 0.111 o.Oo0 f o.Oo0d o.Oo0 f o.Oo0 o.Oo0 f 0.000 

Shea-headed 0.222 f 0.147 0.778 f 0.364d 0.857 f 0.553 1.Oo0 f 0.289

Two tails or heads 0.m f o.Oo0 0.111 f O.llld o.Oo0 f o.Oo0 o.Oo0 f 0.000 

Abnormal (46) 1.11 f1.37 0.20 f1.20 0.21 f1.32 0.21 f 0.17'

Female 

n 10 10 10 10 

Necropsy body wt (9) 23.9 f 0.4d 23.8 f 0.3 24.2 f 0.3 21.3 f 0.6** 

Estrous cycle length (days) 4.70 f 0.26 5.57 f 0.20e 5.33 f 0.42* 5.43 f 0.20e 

Estrous stage (46 of cycle) 

Diestrus 17.1 31.4 41.4 34.3 

Proestrus 30.0 15.7 15.7 18.6 

Estrus 35.7 34.3 27.1 32.9 

Metestrus 17.7 14.3 15.7 14.3 

Uncertain diagnoses 0.0 4.3 0.0 0.0 

** Significantly different(PSO.01) from the control group by Williams' (body weight) or Dunn's test (motility) a Data are presented as mean f standard error. 

n=8 

n=10 * n-9 

e Estrous cycle longer than 7 days or unclear in 3 of 10 animals 

Estrous cycle longer than 7 days or unclear in 4 of 10 animals 
249 

APPENDIX I 

CHEMICAL CHARACTERIZATION AND 

DOSE FORMULATION STUDIES 

PROCUREMENT AND CHARACTERIZATION OF CODEINE . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . 250 

PREPARA~oNAND ANALYSIS OF DOSE FORMULATIONS . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . 251

FIGURE I1 Infrared Absorption Spectrum of Codeine . . . . . . . ... . . . . . . .. . . .. . . .. . . . 253 

FIGURE I2 Nuclear Magnetic Resonance Spectrum of Codeine . . . . . . . . . . . . . . . . . . . . . . . 254 

TABLE I1 Preparationand Storage of Dose Formulations in the Feed Studies of Codeine .. . . . 255 

TABLE I2 Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 14-Day Feed Studies of Codeine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 256 

TABLEI3 Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 13-Week Feed Studies of Codeine . . . . . . . . .... . . . . . . . . . . . . . . . . . . 257 

TABLE I4 Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 2-Year Feed Studies of Codeine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 258 

TABLEI5 Results of Referee Analysis of Dose Formulations Administered to Rats and Mice 

in the 13-Week and 2-Year Feed Studies of Codeine .. . . . . . . . . . . . . . . . . . . . . 262 
250 Codeine, NTP TR 455 

CHEMICAL CHARACTERIZATION AND 

DOSE FORMULATION STUDIES 

PROCUREMENT AND CHARACTERIZATION OF CODEINE 

Codeine was obtained from Penick Corporation (Newark, NJ) in two lots (471NFN003 and 

471NINOOUA). Lot 471NFN003 was usedduringthe14-dayand13-week studies, and lot 471NINOOUA 

was used during the 2-year studies. Identity, purity, and stability analyses were conducted by the 

analytical chemistry laboratory, Midwest Research Institute (Kansas City, MO). Reports on analyses 

performed in support of the codeine studies are on file at the National Institute of Environmental Health 

Sciences (NIEHS). 

Both lots of the chemical, a white powder, were identified as codeine by infrared, ultraviolet/visible, and 

nuclear magnetic resonance spectroscopy. All spectra were consistent with those expected for the 

structure and with the literature spectra (SudtZer Standurd Spectra; Jacobson et al., 1972; CRC, 1975) of 

codeine (Figures I1 and 12). The observed melting point ranges, 155.4" to 156.8" C for lot 

471NINOOUAand 154" to 158" C for lot 471NFN003, and the specific optical activities, 

-117.1 f 0.1" for lot 471NINOOUA and [a]<*=-120.0 f 0.6" for lot 471NFN003, were 

consistent with the literature reference (Merck Index, 1983). 

The purity of each lot was determined by elemental analyses, Karl Fischer water analysis, functional 

group titration, thin-layer chromatography (TLC), and gas chromatography. For functional group 

titration, samples of codeine were dissolved in 25 mL glacial acetic acid and titrated with 0.1 N perchloric 

acid in glacial acetic acid and monitored potentiometrically using a combination pH/mV electrode filled 

with 4 M potassium chloride. TLC was performed on Silica Gel 60 F-254 plates with two solvent 

systems: 1) methanol and 2) carbon tetrachloride:n-butanol:methanol:6 N ammonium hydroxide 

(40:30:30:2). Caffeine was used as a reference standard. Plates were examined with visible and 

ultraviolet light (254 and 366 nm) and with a spray of iodoplatinate reagent. Gas chromatography was 

performed using a flame ionization detector with a nitrogen carrier gas at a flow rate of 70 mL/minute. 

Two systems were used: 

A) 1% SP-2100 on 100/120 Supelcoport (1.8 m X 4 mm ID) glass column, with an oven 

temperature program of 100" C for 5 minutes, then 100 " to 250" C at 10" C per minute, 

and 

B) 1% Dexsil 300 on 100/120 Supelcoport (1.8 m x 4 mm ID) glass column, with an oven 

temperature program of 100" C for 5 minutes, then 100" to 350" C at 10" C per minute. 

Generally, elemental analyses for carbon, hydrogen, and nitrogen were in agreement with the theoretical 

values for codeine; however, the hydrogen value for lot 471NFN003 was higher than the theoretical value 

and the carbon value was slightly lower. Karl Fischer water analysis indicated 1.98% * 0.07% water in 

lot 47 1NFN003 and 4.66 % f 0.05% water in lot 471NINOOUA. Functional group titration indicated a 

purity (anhydrous basis) of 100.0% f 0.5% for lot 471NFN003 and 99.8% f 0.8% for lot 

471NINOOUA. For both lots, TLC by system 1 indicated one major spot and no impurities, and TLC by 

system 2 indicated one major spot and one slight trace impurity. Gas chromatography using system A 

indicated one major peak and one impurity with an area greater than 0.1 % relative to the major peak for 

both lots, and system B indicated one major peak and no impurities. Concomitant gas chromatography 

with a reference sample of lot 47 1NFN003 using system A with an oven temperature of 170" C to 250" C 

at 10" C per minute indicated that lot 47 1 NIN001/A had a purity of 97.4 % f 0.4 % relative to lot 

471NFN003. The overall purity of each lot was determined to be at least 99% on an anhydrous basis. 
Chemical Characterization and Dose Formulation 251 

The analytical chemistry laboratory analyzed lot 471NFN003 to determine if it met United States 

Pharmacopeia XX (USP) purity requirements. The complete battery of USP analyses was performed as a 

supplement to the chemical characterization of codeine. The chemical met the USP requirements for the 

identification tests, which included ultravioletlvisible absorption spectroscopy, a selenious acid color test, 

and an organic nitrogenous bases test. The melting range was determined to be 154.1 " to 156.1 " C. 

Weight losson drying was 1.90% f 0.01%. Theresidueonignition was 0.022% f 0.006%. Test for 

readily carbonizable substances and morphine met USP specifications. The titrimetric purity assay 

indicated that lot 47 1NFN003 contained 100.0% f 0.6% codeine, which met USP requirements for 

purity. 

Stability studies of the bulk chemical (lot 471NFN003) were performed by the analytical chemistry 

laboratory. Gas chromatography was performed using system A for the purity analysis except with an 

oven temperature program of 170" to 250" C at 10" C per minute. These studies indicated that codeine 

was stable as a bulk chemical for 2 weeks when stored protected from light at temperatures up to 25 O C. 

To ensure stability, the bulk chemical was stored at approximately 20" C, in the dark, in sealed amber 

glass bottles in a secure cabinet. The stability of the bulk chemical was monitored by the study 

laboratories prior to the start of the 1Cday and 13-week studies, once during the 13-week studies, and at 

least every 4 months during the 2-year studies using functional group titration and/or gas chromatography. 

No degradation of the bulk chemical was observed. 

PREPARATION AND ANALYSIS OF DOSEFORMULATIONS 

The dose formulations were prepared weekly by mixing codeine with feed (Table 11). Mixtures were 

made by preparing a codeinelfeed premix by hand, which was then blended with feed for 15 minutes using 

an intensifier bar. Formulations were stored in double plastic bags at 4" C (14-day, 13-week, and 

beginning of the 2-year studies) or at room temperature (end of 2-year study) for up to 3 weeks. Bags in 

use were transferred to the study room in covered plastic buckets. 

Homogeneity and stability studies of a 160 and a 30,000 ppm (homogeneity only) formulation were 

performed by the analytical chemistry laboratory. Extracts were prepared by mixing samples in a 

methano1:water solution (70:30) containing 0.9%hydrochloric acid and clarified by centrifugation. For 

the 30,000 ppm dose formulation, aliquots of the extracts were mixed with 4 mL of internal standard 

solution (3 mg propiophenone/mL methanol) and diluted with a water:methanol solution (37:63) containing 

1% glacial acetic acid and 0.005 M sodium heptane sulfonate. High-performance liquid chromatography 

(HPLC) was performed with a Brownlee RP-18 (10 p, 100 mm X 4.6 mm ID) column using ultraviolet 

detection (280 nm) and a mobile phase solvent system of water:methanol (63:37) containing 1 % glacial 

acetic acid and 0.005 M sodium heptane sulfonate. The flow rate was 1.OmL/minute. For the 160 ppm 

dose formulation, 20 mL aliquots of the extracts were mixed with 20 mL water and 20 mL methylene 

chloride and centrifuged to separate the phases. Aliquots (25 mL) of the top layers were mixed with 2 mL 

portions of 1 N sodium hydroxide and 20mL methylene chloride and centrifuged after which the top 

layers were removed by aspiration. Aliquots of the methylene chloride layers were evaporated. The 

residues were dissolved in 3 mL of internal standard solution (0.12 mg propiophenonelml methanol) and 

mixed with 7 mL of water:glacial acetic acid solution (99: 1) containing0.005 M sodium heptane 

sulfonate. The solutions were then analyzed using the HPLC system described previously. Homogeneity 

was confmed, and the stability of the dose formulations was confirmed for up to 3 weeks when stored in 

the dark at room temperature. 

Periodic analyses of the dose formulations of codeine were conducted at the study laboratory and 

analytical chemistry laboratory using HPLC. Dose formulations were analyzed at the beginning of the 

14-day studies (Table 12) and at the beginning, midpoint, and end of the 13-week studies (Table 13). All 
252 Codeine, NTP TR 455 

dose formulations for the 14-day and 13-week studies were within 10% of the target concentrations. 

During the 2-year studies, dose formulations were analyzed every 6 to 10 weeks (Table 14). Of the dose 

formulations analyzed for the 2-year rat study, 97% (86/89)were within 10% of the target concentration. 

Two dose formulations were remixed, and no dose formulation morethan 10% from the target 

concentration was used. Of the dose formulations analyd for the 2-year mouse study, 100% (40/40) 

were within 10% of the target concentration. Results of periodic referee analyses performed by the 

analytical chemistry laboratory indicated good agreementwith the results obtained by the study laboratory 

(Table 15). 
Micrometers 

2.5 3.0 4.0 5.0 6.0 8.0 10 16 20 25 50 

100 

80 

20 

01 I I I I I I 

4000 Wavenumber (cm-1) 2500 2000 1800 1600 1400 Wavenumber (cm-1) 600 400. 200

ABSCISSA ORDINATE SCAN TIME 12min SCAN REP. - SINGLEBEAM - EXPANSION 1 EXPANSION 1 RESPONSE 1 TIME DRIVE - PRESAMPLECHOP a 

SUPPRESSION Off %T 0-1ooo/o ABS - SUT PROGRAM Normal 16) OPERATOR T. Pedenon DATE 7/9/86

SAMPLE Codeine REMARKSReference beam SOLVENT - CEU PATH 

Batch No.: 03 attenuated with IR die 

Lot No.: 471 NIN Wl/A CONCENTRATION 2% (WM) in KBr REFERENCE 250 N 

Task No.: RE-1768 

Projecl No.: 8403-68 

N

8 
- -- 

Codeine 

Lot No.: 471 NIN 0011A 

Batch No.: 03 

Project No.: 8403-68 

Instrument: Nicolet NT-300 INB FF-NMR 

Nucleus: .Proton 

Solvent: Deuterated Chlorofon ONE PULSE SEQUENCE 

Internal Reference: Tetramefhyfsilane P2 = 7.00 USEC 

Sample Temperature: Ambient D5= 10.00 SEC 

Operator:D.Taylor NA= 16 

Date: 7109186 SIZE= 16384 

AT = 2.05 SEC 

Integration OPDON= 1 

Assignments (bppm) J Obsewed Theoretical ABC ON 

BUl-rERWORTH FILTER ON 

(a) ca. 1.81 -2.1 4 2 DBAlT.= 3 

(b) ca. 2.24-2.71 ADC = 12 BiTS 

(c) ca. 2.38-2.47 7 Ai= 4 

(d) 2.43 sw = +/- 2000.00 

(e) 2.59 DW = 250 

(0 3.04 Jc.l i: 18.7 HZ 0.99 1 RG = 100 USEC 

1.11 1 DE = 250 USEC (9) 3.35 

(h) 3.82 2.83 3 

n HIGH POWER ON 

(i) 4.18 0.92 1 f-F2 = 300.054777

0) 4.79 

1.02 1 BB MODULATION ON 

(k) 4.88 1.02 1 OF = 1299.70

(I) 5.28 1.02 1. SF= 300.054484 

(m)5.70 1.02 1 EM= .IO 

(n) or (0) 6.56 J, = 8.2 HZ 1.02 1 PA= 134.6 

1.02 1 PB= 70.5 (a) or (n) 6.65 

Impurities: 

(p) ca. 2.67-2.86 Trace 

(9)ca. 3.309.47 Trace 

(r) Chloroform, 7.29 

10 8 6 4 2 0 PPM
Chemical Characterization and Dose Formulation 255 

TABLEI1 

Preparation and Storage of Dose Formulations in the Feed Studies of Codeine ~~~~ ~ ~ 

14-Day Studies 13-WeekStudies 

~~ 

2-Year Studies 

~ __ 

Preparation 

A premix of feed and codeine was A premix of feed and codeine was 

prepared, then layered into the prepared, then layered into the 

remaining feed and blended in a remaining feed and blended in a 

Patterson-Kelly V-blender with the Patterson-Kelly twin-shell blender with 

intensifier bar on for 15 minutes. the intensifier bar on for 15 minutes. 

Doses were prepared weekly. Doses were prepared weekly. 

Chemical Lot Number 

471NFN003 471NFN003 471NIN001/A 

MaximumStorageTime 

3 weeks 3 weeks 3 weeks 

Stored in double plastic bags at 4' C 

Storage Conditions 

Stored in double plastic bags at4" C 

or at room temperaNre. Bags in use 

Stored in double plastic bags at 4" C 

stored in covered plastic buckets in the 

study room. 

EG&G Mason Research Institute 

Study Laboratory 

EG&G Mason Research InstiNte Microbiological Associates, Inc. 

(Worcester, MA) (Worcester, MA) (Bethesda, MD) 

Midwest Research Institute 

Referee Laboratory 

Midwest Research Institute Midwest Research Institute 

(Kansas City, MO) (Kansas City, MO) (Kansas City, MO) 
256 Codeine, NTP TR 455 

TABLEI2 

Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 14-Day Feed Studies of Codeine 

Determined Target % Difference 

DatePreparedDateAnalyzedConcentrationConcentrationafromTarget 

(PPd @Pm) 

Rats 

26 March 1984 29 March 1984 1,562 1,564 0 

3,125 3,170 +1 

30 March 1984 6,250 6,310 +1 

12,500 12,OOO -4 

25,OOO 24.700 -1 

Mice 

26 March 1984 29 March 1984 781 740 -5 

1,562 1,564 0 

3,125 3.170 +1 

30 March 1984 

12,500 

6,250 

12,000 

6.310 

-4 

+1 

a Results of duplicate analyses 
Chemical Characterization and Dose Formulation 257 

TABLEI3 

Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 13-Week Feed Studies of Codeine 

ConcentrationDate Date Analyzed Prepared

10 October 1984 12 October 1984 

17 October 1984 

5 December 1984 6 December 1984 

30 January 1985 31 January 1985 

5 February 1985 

a Resultsofduplicateanalyses 

Sample selection, from top leftof V-blender 

Sample selection from top right of V-blender 

Sample selection from bottom of V-blender 

10 December3,125 1984 

Target 

@Pm) 

390 

390 

390 

781 

1,562 

3,125 

6,250 

6,250 

6.250 

390 

781 

1.562 

6,250 

390 

78 1 

1.562 

3,125 

6,250 

Determined 

Concentrationa Target from

@Pd 

399b 

418' 

368* 

780 

1,490 

3.020 

6,230b 

6,520' 

6,010' 

384 

773 

1,550 

3.050 

6,210 

394 

789 

1,480 

3,100 

6.070 

% Difference 

+2 

+7 

-6 

0 

-5 

-3 

0 

+4 

-4 

-2 

-1 

-1 

-2 

-1 

+1 

+1 

-5 

-1 

-3 
TABLEI4 

Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 2-Year Feed Studies of Codeine 

Determined Target

Concentration Analyzedfrom Concentrationa TargetDat Date Prepared

@Pm) @Pd 

Rats 

3 April 1989 5-7 April 1989 381 

409 

772 

760 

1,490 

1.620 

15 May 1989 16-18 May 1989 . 408 

b 

379 

776 

751 

1,530 

1,560 

10 July 1989 10-14 July 1989 b 

470 

811 

839 

1,760 

1,770 

18 July 1989 400 378C9d 

17 July 1989 18 July 1989 400 404e 

1,600 1,480 

5 September 1989 I 6-8 September 1989 406 

422 

855 

820 

1,660 

1,690 

30 October 1989 14 November 1989 424 

419 

789 

737 

'b 

1,700 

7 November 1989 1,770 

8 November 1989 422'.* 

8 November 1989e9 November 1989 1,600 1.670 

Codeine, NTP TR 455 

% Difference 

-5 

+2 

-3 

-5 

-7 

+1 

+2 

-6 

-3 

-6 

-4 

-2 

+18 

+1 

+5 

+10 

+11 

-5 

+1 

-7 

+2 

+6 

+7 

+3 

+4 

+6 

+6 

+5 

-1 

-8 

+6 

+11 

+6 

+4 
Chemical Characterization andDose Formulation 259 

TABLEI4 

Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 2-YearFeed Studies of Codeine (continued) 

Determined Target % Difference

ConcentrationConcentration Analyzed TargetPrepared DatefromDate

Rats (continued) 

18 December 1989 19-21 

26 February 1990 

16 April 1990 

11 June 1990 

20 August 1990 

1 October 1990 

December 1989 400 

21-22December 1989 

27 February - 3 March 1990 

17-19April 1990 

12-14June 1990 

21-23August 1990 

2-4 October 1990 

@Pd 

400 

800 

800 

1,600 

1,600 

400 

800 

1,600 

400 

400 

800 

800 

1,600 

1.600 

400 

400 

800 

800 

1,600 

1,600 

(PPd 

398 

b 

813b 

743 

b 

1.490 

390' 

787'*d 

1,520' 

379 

375 

771 

769 

1,560 

1,500 

381 

406 

799 

780 

1,540 

1,560 

397 

380 

792 

776 

1.620 

1,540 

416 

407 

792 

801 

1,590 

1,600 

381 

405 

855 

841 

1,540 

1,570 

0 

+2 

-7 

-7 

-2 

-2 

-5 

-5 

-6 

-4 

-4 

-2 

-6 

-5 

+2 

0 

-2 

-4 

-2 

-1 

-5 

-1 

-3 

+1 

-4 

+4 

+2 

-1 

0 

-1 

0 

-5 

+1 

+7 

+5 

-4 

-2 
Codeine, NTP TR 455 

TABLEI4 

Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 2-YearFeed Studies of Codeine (continued) 

Determined Target % Difference

ConcentrationConcentration Analyzed TargetPrepared Datefrom Date

Rats (continued) 

26 November 1990 

21 January 1991 

25 March 1991 

Mice 

20 March 1989 

15 May 1989 

10July 1989 

5 September 1989 

30 October 1989 

27-30November 1990 

22-24January 1991 

25 January 1991 

26-28March 1991 

21-27March 1989 

16-18May 1989 

10-14July 1989 

6-8 September 1989 

1-4November 1989 

@Pm) 

400 

400 

800 

800 

1,600 

1,600 

800 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

750 

1,500 

3,ooo 

750 

1,500 

3,ooo 

750 

1,500 

3,ooo 

@Pd 

402 

399 

793 

782 

1,570 

1,630 

386 

368 

yll 

1,530 

1,570 

788' 

373 

362 

742 

748 

1.480 

1,440 

706 

1,460 

2.930 

778 

1.470 

2.980 

724 

1,560 

3.100 

807 

1,640 

3,010 

711 

1490 

2940 

+1 

0 

-1 

-2 

-2 

+2 

-3 

-8 

+1 

-4 

-2 

-1 

-7 

-9 

-7 

-6 

-7 

-10 

-6 

-3 

-2 

+4 

-2 

-1 

-3 

+4 

+3 

+8 

+9 

0 

-5 

-1 

-2 
Chemical Characterization and Dose Formulation 261 

TABLEI4 

Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 2-YearFeed Studies of Codeine (continued) 

ConcentrationDate Date Analyzed Prepared Target 

@Pm) 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

750 

1,500 

3.00 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

750 

1,500 

3,Ooo 

Determined 

Concentration Target from

(PPm) 

792b 

b 

b 

771egd 

1,500' 

3,Ooo' 

72 1 

1,400 

2.880 

790 

1.430 

2,830 

708 

1,510 

2,890 

70 1 

1,490 

2,990 

77 1 

1,480 

2.990 

738 

1,500 

3,220 

718 

1,440 

2,950 

% Difference 

+6 

+3 

0 

0 

-4 

-7 

-4 

+5 

-5 

-6 

-6 

fl 

-4 

+1 

-1 

0 

+3 

-1 

0 

-2 

0 

+7 

-4 

-4 

-2 

Mice (continued) 

18 December 1989 

18 December 1989 

26 Febmary 1990 

16 April 1990 

11 June 1990 

20 August 1990 

1 October 1990 

26 November 1990 

21January1991 

0 

a Results of duplicateanalyses 

Repeat analysis required 

Results of reanalysis 

19-21 December 1989 

21-22 December 1989 

27 February -

3 March 1990 

17-19 April 1990 

12-14 June 1990 

21-23 August 1990 

2-4 October 1990 

27-30 November 1990 

22-24 January 1990 

Results of triplicate analyses 

e Results of remix 
262 Codeine, NTP TR 455 

TABLEI5 

Results of Referee Analysis of Dose Formulations Administeredto Rats and Mice 

in the 13-Week and 2-Year Feed Studies of Codeine 

DeterminedConcentration bpm) 

Target Concentration Referee Study 

Date Prepared (PPm) Laboratoryb Laboratorya

13-Week Studies (EG&G Mason Research Institute) 

10 October 1984 78 1 780 774 f 1.2 

2-Year Studies (MicrobiologicalAssociates,Inc.) 

Rats 

3 April 1989 800 772 808 f 14 

Mice 

20 March 1989 3,Ooo 2,930 3,050 f 220 

5 September 1989 1,500 1,640 1,610 f 70 

26 Februay 1990 750 72 1 762 f 22 

a Results of duplicateanalyses 

Results of triplicate analyses (mean f standarderror) 
263 

APPENDIX J 

FEED AND COMPOUND CONSUMPTION 

IN THE 2-YEAR FEED STUDIES 

OF CODEINE 

TABLE J1 Feed and CompoundConsumptionbyMale Rats in the 2-Year FeedStudy 

ofcodeine ................................................. 264 

TABLE 52 Feed and CompoundConsumptionbyFemale Rats in the 2-Year FeedStudy 

ofcodeine ................................................. 265 

TABLE 53 Feed q"ompound ConsumptionbyMaleMiceinthe 2-Year FeedStudy 

ofcodeine ................................................. 266 

TABLE 54 Feed and CompoundConsumptionbyFemaleMice in the 2-Year FeedStudy 

ofcodeine ................................................. 267 
264 , Codeine, NTP TR 455 

TABLEJ1 

Feed and Compound Consumptionby Male Rats in the 2-YearFeed Study of Codeine 

2 16.7 199 14.9 187 32 14.9 182 66 14.8 179 132 

6 16.9 292 17.2 279 25 17.2 272 50 18.1 268 108 

10 18.1 341 17.2 327 21 16.7 320 42 17.6 315 89 

14 16.5 368 16.4 355 18 16.9 348 39 17.0 346 79 

17 15.8 392 16.5 370 18 16.1 363 36 18.1 354 82 

21 16.9 407 16.5 396 17 16.5 382 34 17.3 366 76 

25 16.6 430 17.0 414 16 16.9 398 34 16.9 387 70 

29 14.4 438 14.1 426 13 15.3 408 30 17.1 397 69 

33 17.1 444 16.9 429 16 16.0 414 31 16.3 394 66 

37 14.7 452 14.3 437 13 15.3 420 29 16.9 402 67 

41 16.8 458 17.1 442 16 16.5 428 31 15.7 405 62 

45 14.6 465 15.4 452 14 15.7 434 29 17.1 408 67 

49 16.4 466 15.7 452 14 15.5 435 28 16.2 405 64-

53 16.5 478 16.4 465 14 16.8 444 30 18.5 407 73 

57 15.0 477 14.9 468 13 15.3 450 27 16.4 409 64 

61 15.0 470 14.4 459 13 15.2 443 28 17.8 404 71 

65 15.0 471 15.5 458 14 15.2 444 27 15.8 400 63 

69 13.6 463 13.6 454 12 13.6 439 25. 17.6 403 70 

73 14.7 459 14.3 453 13 13.8 435 25 16.4 397 66 

77 14.0 458 13.4 448 12 13.9 429 26 16.9 395 68 

81 13.7 453 13.9 444 12 14.3 423 27 16.1 393 65 

85 13.8 444 13.3 439 12 14.3 423 27 15.8 393 64 

89 12.7 447 . 13.1 434 12 14.9 424 28 15.1 396 61 

93 12.7 437 12.4 428 12 13.8 415 27 14.0 390 57 

97 13.0 427 11.9 412 12 13.3 408 26 13.9 382 58 

101 13.0 433 12.3 418 12 12.0 401 24 11.9 376 51 

Means for weeks 

1-13 17.2 277 16.4 264 26 16.3 258 53 16.8 254 110 

14-52 16.0 432 16.0 417 15 16.1 403 32 16.9 386 70 

53-101 14.0 455 13.8 445 12 14.3 429 27 15.8 396 64 

a Grams of feed consumed per animal per day 

Milligrams of codeine consumed per kilogram body weight per day 
Feed and Compound Consumption 265 

TABLE52 

Feed and Compound Consumption by Female Rats in the2-YearFeed Study of Codeine 

2 12.0 140 10.8 

6 11.5 182 11.3 

11.0 136

12.2 175

192 

32 136 65 10.6 133 128 

26 171 57 12.3 173 114 

10 12.4 201 11.1 12.0 23 191 50 11.8 188 101 

14 11.4 215 12.6 12.0 205

17 11.6 224 10.9 211 21 12.0 206 47 12.2 204 96 

25 203 47 11.6 198 94 

21 10.8 233 10.8 11.8 223

11.0 228

11.3 233

10.7 237

37 11.4 260 10.8 240 18 ' 11.0 243 36 10.7 245 70

19 221 43 12.3 217 91 

25 10.5 242 11.0 19 227 39 11.1 223 79 

29 11.1 244 10.3 18 232 39 11.1 232 77 

33 11.0 255 10.3 17 235 36 11.4 234 78 

41 11.5 273 10.7 10.9 251 

11.7 259

11.1 266

13.1 277

12.3 288

12.6 295

12.0 306

12.0 308

12.6 312

12.1 317

12.0 319

85 10.9 341 11.3 325 14 ' 11.6 323 29 11.2 307 59 

11.7 326

12.1 333

10.8 334

10.4 336

17 25 1 35 11.5 249 74 

45 11.8 286 11.6 18 258 36 10.6 255 66

49 12.0 297 11.0 17 262 34 11.6 256 72 

53 12.1 3 10 12.2 18 270 39 11.5 264 70 

57 12.2 320 12.0 17 284 35 12.0 271 71

61 12.6 328 12.5 17 292 34 11.2 279 64

65 11.1 336 11.9 16 302 32 11.8 283 67

69 11.4 33 1 11.4 15 304 32 11.1 284 62 

73 12.1 339 11.9 15 307 33 12.0 290 66 

77 11.4 342 11.1 14 315 31 11.1 297 60

81 11.8 342 11.7 15 317 30 11.7 298 63 

89 10.8 338, 11.6 14 325 29 11.0 304 58

93 11.5 343 12.3 15 327 30 11.4 309 59

97 12.5 345 11.5 14 324 27 11.1 307 58

101 11.6 349 11.2 13 324 26 11.4 303 60

Mew for weeks 

1-13 12.0 175 11.7 167

11.4 235

53-101 11.7 336 11.7 313 15 ' 11.9 309 31 11.4 292 63 

a Grams of feed consumed per animal perday 

Milligrams of codeine consumed per kilogram body weight per day 

11.1 27 166 57 11.6 164 114

14-52 11.3 253 11.0 19 234 39 11.4 23 1 80 
266 Codeine, NTP TR 455 

TABLE53 

Feed and Compound Consumption by Male Mice in the 2-YearFeed Study of Codeine 

0 DDUl 750 DRUI 1.500 PDm 3.000 DPIII 

Body Feed Dose/Body Feed Body Feed 

Weight (glday) Day(%day)Weight WeightDayb Day(glday) Weight 

Week (9) (9) (mdkglday) 0 (mglkglday) (9) (mdkglday) 

2 4.4 23.8 3.8 23.6 121 3.9 23.4 251 4.1 22.8 537 

6 4.7 28.1 4.5 27.7 123 4.4 27.2 245 4.5 26.9 498 

10 5.0 32.4 5.1 31.1 122 5.1 30.5 25 1 4.9 29.8 497 

14 5.1 35.2 5.5 34.0 121 5.4 32.8 246 5.2 31.7 492 

17 5.0 38.1 5.1 36.9 104 5.2 35.0 223 5.1 33.3 460 

21 4.6 41.6 4.8 40.0 90 4.9 37.2 196 4.8 34.9 414 

25 4.7 42.3 5.0 41.1 91 5.1 39.3 196 5.2 36.6 422 

29 4.7 44.9 4.7 44.8 79 4.8 42.3 169 4.7 38.9 360 

33 4.9 46.1 5 .O 45.1 83 5.1 42.5 180 5.2 38.5 404 

37 4.4 48.3 4.7 46.8 76 4.9 44.2 165 4.7 40.4 348 

41 4.6 48.9 4.8 48.9 73 4.8 45.7 156 5.0 41.7 357 

45 4.5 49.5 4.6 49.9 69 5.0 47.0 160 5.0 43.3 345 

49 4.6 49.4 4.8 50.0 72 5.1 47.4 161 5.0 43.8 341 

53 4.7 50.0 4.9 51.3 72 5.1 48.7 156 5.0 44.6 337 

57 4.9 50.4 5.1 51.6 74 5.0 49.2 153 5.2 45.2 342 

61 4.9 50.7 5.0 51.7 72 5.1 49.4 155 5.1 45.4 339 

65 5.0 50.3 5.0 51.5 72 5.1 49.0 156 4.9 45.0 326 

69 4.7 50.6 4.7 51.3 69 4.7 49.5 143 4.7 44.7 313 

73 4.8 49.4 4.9 50.5 73 5.1 48.6 157 4.9 43.4 337 

77 4.7 49.2 4.9 50.4 72 5.0 48.5 156 5.0 43.4 343 

81 5.1 49.9 5.O 50.6 74 5.4 47.5 169 5.1 43.7 353 

85 4.7 49.5 4.9 50.3 73 5.0 47.5 157 5.0 43.8 344 

89 4.6 50.2 4.7 51.4 69 5.1 48.1 160 5.0 44.2 338 

93 4.9 48.5 5.1 50.3 76 5.2 45.9 171 5.1 41.8 368 

97 4.9 49.0 4.8 5 1.O 71 5.0 46.0 163 4.9 42.2 348 

101 4.8 49.0 4.9 50.3 74 5.2 45.1 173 5.1 41.7 365 

Meam for weeks 

1-13 4.7 28.1 4.5 27.5 122 4.5 27.0 249 4.5 26.5 511 

14-52 4.7 44.4 4.9 43.8 86 5.0 41.3 185 5.0 38.3 394 

53-101 4.8 49.7 4.9 50.9 72 5.1 47.9 159 5.0 43.8 342 

a Grams of feed consumedper animal perday 

Milligrams of codeine consumed per kilogram body weight per day 
Feed and Compound Consumption 267 

TABLE54 

Feed and Compound Consumptionby Female Mice in the 2-Year Feed Study of Codeine ~~~~ ~ ~ 

0 DDm 750 DDIU 1.500 DDIU 3,000 DDIU 

Body Feed Body Feed Body Feed Dose/ Body Feed Dose/ Dose/

DayWeight(plday) (glday) WeightWeight Dayb (glday) WeigM Day Week 0 0 (mdkg/day) (8) (mdkgldaj.) 0 (md4my) 

2 4.3 19.5 4.5 19.6 174 4.4 19.2 345 4.1 18.9 651 

6 4.6 24.5 4.5 24.5 138 4.5 24.1 217 4.4 23.6 564 

10 5.0 28.3 5.0 27.7 137 5.1 27.3 282 5.0 26.6 568 

14 5.2 31.1 5.5 30.0 137 5.5 29.0 283 5.2 28.1 560 

17 5.4 34.5 5.3 33.3 119 5.4 31.2 258 5.2 29.5 528 

21 4.8 39.1 5.1 36.9 103 5.1 34.0 223 5.0 31.7 470 

25 5.2 40.7 5.3 38.8 103 5.2 36.6 214 5.2 33.6 469 

29 5.2 42.5 5.1 41.7 93 5.2 38.9 201 5.0 35.1 432 

33 5.0 44.6 5.4 42.4 95 5.4 39.8 205 5.6 35.5 474 

37 5.0 46.2 5.5 44.6 92 5.5 41.4 199 5.6 36.5 457 

41 5.1 47.5 5.3 47.2 84 5.3 43.8 183 5.3 38.5 415 

45 5.1 48.9 5.3 49.0 80 5.2 45.3 174 5.4 39.9 403 

49 5.2 48.7 5.4 49.5 82 5.5 46.5 177 5.6 40.4 412 

53 5.2 50.2 5.5 51.0 81 5.6 48.3 174 5.5 42.2 389 

57 5.3 51.2 5.5 52.7 78 5.5 50.4 163 5.6 43.6 384 

61 5.2 52.5 5.5 54.3 75 5.5 51.7 159 5.6 44.4 376 

65 5.4 52.4 5.6 54.0 77 5.4 51.3 157 5.5 44.2 370 

69 5.4 52.6 5.5 53.5 78 5.7 52.5 163 5.6 44.2 377 

73 5.2 50.9 5.5 53.2 78 5.7 52.4 164 5.7 44.3 385 

77 5.2 49.8 5.4 53.2 76 5.5 52.3 158 5.4 43.7 372 

81 5.2 51.4 5.5 53.9 77 5.5 53.2 155 5.6 44.3 376 

85 5.1 53.0 5.2 54.2 72 5.4 54.1 150 5.5 44.1 371 

89 5.2 53.2 5.3 53.5 75 5.6 53.7 157 5.8 43.6 397 

93 5.3 51.3 5.4 52.1 77 5.4 52.4 155 5.6 41.7 404 

97 4.8 52.5 5.2 53.2 73 5.3 52.5 152 '5.4 42.4 384 

101 5.1 51.6 5.5 53.5 77 5.5 51.3 162 5.7 41.4 411 

Means for weeks 

1-13 4.6 24.1 4.7 23.9 150 4.7 23.5 301 4.5 23.0 595 

42.4 5.3 41.3 99 5.3 38.7 212 5.3 34.9 462 

51.7 5.4 53.3 77 5.5 52.0 159 5.6 43.4 384 

a Grams of feedconsumedper animal perday 

Milligrams of codeine consumed per kilogram body weight per day 

14-52 5.1 

5.2 53-101
269 

APPENDIX K 

INGREDIENTS, NUTRIENT COMPOSITION, 

AND CONTAMINANT LEVELS 

IN NIH-07 RAT AND MOUSE RATION 

TABLEK1 Ingredients of NIH-07 Rat and Mouse Ration ........................... 270 

TABLEK2 Vitamins and Minerals inNIH-07 Rat and Mouse Ration ................... 270 

TABLEK3 Nutrient Compositionof NIH-07 Rat and Mouse Ration .................... 271 

TABLEK4 ContaminantLevels in NIH-07 Rat and Mouse Ration ..................... 272 
270 Codeine, NTP TR 455 

TABLEK1 

Ingredients of NM-07 Rat and Mouse Rationa 

Ingredientsb 

Ground #2 yellow shelled corn 

Ground hard winter wheat 

Soybean meal (49% protein) 

Fish meal (60% protein) 

Wheat middlings 

Dried skim milk 

Alfalfa meal (dehydrated, 17% protein) 

Corn gluten meal (60% protein) 

soy oil 

Dried brewer’s yeast 

Dry molasses 

Dicalcium phosphate 

Ground limestone 

Salt 

Premixes (vitamin and mineral) 

a NCI, 1976;NM, 1978 

Percent by Weight 

24.50 

23.00 

12.00 

10.00 

10.00 

5.00 

4.00 

3.OO 

2.50 

2.00 

1 SO 

1.25 

0.50 

0.50 

0.25 

Ingredients were ground to pass through a U.S. Standard Screen No. 16 before being mixed. 

TABLEK2 

Vitamins and Minerals in NIH-07 Rat and Mouse Rationa 

Vitamins 

A 

D3 

K3 

d-or-Tocopheryl acetate 

Choline 

Folic acid 

Niacin 

d-Pantothenic acid 

Riboflavin 

Thiamine 

B12. 

Pyndoxine 

Biotin 

Minerals 

Iron 

Manganese 

Zinc 

Copper 

Iodine 

Cobalt 

Amount 

5,500,OooIU 

4,600,OooIU 

2.8 g 

20,OooIu 

560.0 g 

2.2g 

30.0 g 

18.0g 

3.4g 

10.0g 

4.Ooo 

1.7 g 

140.0 mg 

120.0g 

60.0g 

16.0g 

4.0 g 

1.4 g 

0.4 g 

Source 

Stabilized vitamin A palmitate or acetate 

D-activated animal sterol 

Menadione 

Choline chloride 

&Calcium pantothenate 

Thiamine mononitrate 

Pyridoxine hydrochloride 

d-Biotin 

Iron sulfate 

Manganous oxide 

Zinc oxide 

Copper sulfate 

Calcium iodate 

Cobalt carbonate 

a Per ton (2,OOOlb) of finished product 
Feed Analyses 271 

TABLEK3 

Nutrient Compositionof NIH-07 Rat and Mouse Ration 

Nutrient 

Protein (R by weight) 

Crude fat (% by weight) 

Crude fiber(96by weight) 

Ash (96by weight) 

Amino Acids (% of total diet) 

Arginine 

Cystine 

Glycine 

Histidine 

Isoleucine 

Leucine 

Lysine 

Methionine 

Phenylalanine 

Threonine 

Tryptophan 

Tyrosine 

Valine 

Essential Fatty Acids (% of total diet) 

Linoleic 

Linolenic 

Vitamins 

Vitamin A (IUkg) 

Vitamin D (IU/kg) 

a-Tocopherol (ppm) 

Thiamine (ppm) 

Riboflavin (ppm) 

Niacin (ppm) 

Pantothenic acid (ppm) 

Pyridoxine (ppm) 

Folic acid (ppm) 

Biotin (ppm) 

Vitamin BIZ(ppb) 

Choline (ppm) 

Minerals 

Calcium (I) 

Phosphorus (%) 

Potassium (X) 

Chloride (X) 

Sodium (%) 

Magnesium (I) 

Sulfur (%) 

Iron (ppm)

Manganese (ppm) 

Zinc (ppm) 

Copper (ppm)

Iodine (ppm) 

Chromium (ppm) 

Cobalt (ppm) 

Mean f Standard 

Deviation 

-23.46f 0.78 

5.23 f 0.23 

3.64 f 0.51 

6.52 f 0.18 

1.287 f 0.084 

0.306 3 0.075 

1.160f 0.050 

0.580 f 0.024 

0.917 f 0.034 

1.972 f 0.052 

1.273 f 0.051 

0.437 f 0.115 

0.994 f 0.125 

0.896 f 0.055 

0.223 f 0.160 

0.677 f 0.105 

1.089 f 0.057 

2.389 f 0.233 

0.277f 0.036 

7,084f 1.650 

4,450 f 1,382 

36.92 f 9.32 

18.29 f 1.59 

7.92 f 0.93 

100.95 f 25.92 

30.30 f 3.60 

9.25 f 2.62 

2.51 f 0.64 

0.267 f 0.049 

40.14 f 20.04 

3,068 f 314 

1.21 f 0.10 

0.95 f 0.04 

0.887f 0.067 

0.526f 0.092 

0.315 f 0.034 

0.168 f 0.008 

0.274 f 0.063 

356.2f 90.0 

92.24 f 5.35 

58.14 f 9.91 

11.50 f 2.40 

3.70 f 1.14 

1.71 f 0.45 

0.797f 0.23 

Range 

21.30 - 25.20 

4.80 - 5.80 

2.60 - 4.80 

6.12- 6.97 

1.100- 1.390 

0.181 - 0.400 

1.060- 1.220 

0.531 - 0.608 

0.867 - 0.965 

1.850- 2.040 

1.200 - 1.370 

0.306 - 0.699 

0.665 - 1.110 

0.824 - 0.985 

0.107 - 0.671 

0.564 - 0.794 

0.962- 1.170 

1.830- 2.570 

0.210- 0.320 

4,290- 12,540 

3,OOO - 6,300 

22.5 - 48.9 

15.0 - 22.0 

6.10- 9.00 

65.0- 150.0 

23.0 - 34.6 

5.60 - 14.0 

1.80- 3.70 

0.19 - 0.35 

10.6 - 65.0 

2,400- 3,430 

0.90- 1.37 

0.88 - 1.03 

0.772- 0.971 

0.380 - 0.635 

0.258 - 0.370 

0.151 - 0,180 

0.208- 0.420 

255.0- 523.0 

81.70- 99.40 

46.10 - 81.60 

8.090- 15.39 

1.52 - 5.83 

0.85 - 2.09 

0.490 - 1.150 

Number of Samples 

26 

26 

26 

26 

10 

10 

10 

10 

10 

10 

10 

10 

10 

10 

10 

10 

10 

9 

9 

26 

4 

9 

26 

10 

9 

10 

10 

10 

10 

10 

9 

26 

26 

8 

8 

10 

10 

10 

10 

10 

10 

10 

10 

9 

6 
272 Codeine, NTP TR 455 

T~BLE~4 

ContaminantLevels in NIH-07Rat and Mouse Rationa 

Mean f Standard 

Deviationb Range 

Contaminants 

Arsenic (ppm) 0.34 f 0.17 0.06 - 0.60 

Cadmium (ppm) 0.07f 0.03 0.05 - 0.12 

Lead (ppm) 0.25 f 0.18 0.10- 0.90 

Mercury (ppm) 0.03 f 0.02 0.02 - 0.08 

Selenium (ppm) 0.33 f 0.10 0.10- 0.52 

Aflatoxins (ppb)' <5.0 

Nitrate nitrogen (ppm)d 14.10 f 4.40 5.90 - 21.0 

Nitrite nitrogen (ppm)d 0.23 f 0.21 0.10- 1.00 

BHA (ppmle 1.44 f 0.87 1 .00- 4.00 

BHT (ppmle 1.44 f 1.23 1.00- 7.00 

Aerobic plate count (CFUlg) 10,802f 83,872 4,700- 380,ooO 

Coliform (MPN/g) 25.64 f 27.5 3.00- 93.00 

Escherichia coli (MPN/g) 3.31 f 1.20 3.00 - 9.0 

Total nitrosoamines (ppb)f 7.20f 2.48 2.00 - 13.70 

N-Nitrosodimethylamine(p b)f 5.45 f 2.00 1.00 - 11.00 P N-Nitrosopyrrolidine (ppb) 1.75 f 1.02 1.00 - 4.30 

Pesticides(ppm) 

(u-BHC <0.01 

fl-BHC <0.02 

y-BHC <0.01 

&BHC <0.01 

Heptachlor <0.01 

Aldrin <0.01 

Heptachlor epoxide <0.01 

DDE <0.01 

DDD <0.01 

DDT <0.01 

HCB <0.01 

Mirex <0.01 

Methoxychlor <0.05 

Dieldrin <0.01 

Endrin <0.01 

Telodrin <0.01 

Chlordane <0.05 

Toxaphene <0.1 

Estimated PCBs <0.2 

Ronnel <0.01 

Ethion <0.02 

Trithion <0.05 

Diazinon <0.1 

Methyl parathion <0.02 

Ethyl parathion <0.02 

Malathion 0.28 f 0.26 <0.05 - 1.00 

Endosulfan I <0.01 

Endosulfan I1 <0.01 

Endosulfan sulfate <0.03 

a CFU = colony forming units, MPN = mostprobablenumber, BHC is hexachlorocyclohexane 

For values less than the limit of detection, the detection limit is given as the mean. 

No aflatoxin measurement was recorded for the lot milled2 October 1989. 

Sources of contamination: alfalfa, grains, and fish meal. 

e Sources of contamination: soy oil and fish meal; no BHA or BHT measurementswererecorded 

All values were corrected for percent recovery. 

Number of Samples 

26 

26 

26 

26 

26 

25 

26 

26 

26 

26 

26 

26 

26 

26 

26 

26 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

31 

26 

31 

31 

31 

or benzene hexachloride. 

for the lot milled 1 November 1989. 
273 

APPENDIX L 

SENTINEL ANIMAL PROGRAM 

METHODS ............................................................ 274 

RESULTS .......................................................... 276 
274 Codeine, NTP TR 455 

SENTINEL ANIMAL PROGRAM 

MJ3THODS 

Rodents used in the Carcinogenesis Program of the National Toxicology Program are produced in 

optimally clean facilities to eliminate potential pathogens that may affect study results. The Sentinel 

Animal Program is part of the periodic monitoring of animal health that occurs during the toxicologic 

evaluation of chemical compounds. Under this program, the disease state of the rodents is monitored via 

serology on sera from extra (sentinel) animals in the study rooms. These animals and the study animals 

are subject to identical environmental conditions. The sentinel animals come from the same production 

source and weanling groups as the animals used for the studies of chemical compounds. 

Serum samples were collected from randomly selected rats and mice during the 13-week and 2-year 

studies. Blood from each animal was collected and allowed to clot, and the serum was separated. The 

samples were processed appropriately and sent to Microbiological Associates, Inc. (Bethesda, MD), for 

determination of antibody titers. The laboratory serology methods and viral agents for which testing was 

performed are tabulated below; the times at which blood was collected during the studies are also listed. 

Method and Test Time of Analvsis 

RATS 

13-WeekStudy 

ELISA 

Mycoplasma arthritidis Study termination 

Mycoplasma pulmnis Study termination 

PVM (pneumonia virus of mice) Study termination 

RCV/SDA (rat coronavirus/sialodacryoadenitisvirus) Study termination 

Sendai Study termination 

Hemagglutination Inhibition 

H-1 (Toolan’s H-1 virus) Study termination 

KRV (Kilham rat virus) Study termination 

2-Year Study 

ELISA 

M. arthritidis Study termination 

M.pulmnis Study termination 

PVM 6, 12,and18months,studytermination 

RCV/SDA 6, 12,and18months,studytermination 

Sendai 6, 12,and18months,studytermination 

Hemagglutination Inhibition 

H- 1 6, 12,and18months,studytermination 

KRV 6, 12,and18months,studytermination 
Sentinel Animal Program 215 

MICE 

13-W~kStudy 

Complement Fixation 

LCM (lymphocytic choriomeningitis virus) 

ELISA 

Ectromelia virus 

GDVII (mouse encephalomyelitis virus) 

Mouse adenoma virus 

MHV (mouse hepatitis virus) 

M.arthritidis 

M.pulmonis 

PVM 

Reovirus 3 

Sendai 

Immunofluorescence Assay 

EDIM (epizootic diarrhea of infant mice) 

Hemagglutination Inhibition 

K (papovavirus) 

MVM (minute virus of mice) 

Polyoma virus 

2-Year Study 

ELISA 

Ectromelia virus 

EDIM 

GDVII 

LCM 

MVM 

Mouse adenoma virus 

Mouse adenoma virus - FL 

MHV 

M.arthritidis 

M.pulmonis 

PVM 

Reovirus 3 

Sendai 

Immunofluorescence Assay 

EDIM 

LCM 

MVM 

Mouse adenoma virus 

Reovirus 3 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

Study termination 

6, 12, and 18 months, study termination 

18 months 

6, 12, and '18 months, study termination 

6, 12, and 18 months, study termination 

6 months 

6 months 

18 months, study termination 

6, 12, and 18 months, study termination 

Study termination 

Study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months,study termination 

Study termination 

12 months 

12 months 

18 months, study termination 
276 Codeine, NTP TR 455 

2-Year Study (continued) 

Hemagglutination Inhibition 

K 6, 12, and 18 months, study termination 

MVM 18 months, study termination 

Polyoma virus 6, 12, and 18 months, study termination 

RESULTS 

For the 13-week feed studies with rats and mice, all serology test results were negative. One rat and one 

mouse had positive titers to M. arthritidis at the end of the 2-year studies. 

Further evaluation of serum positive for M. arthritidis by immunoblot and Western blot procedures 

indicated that the positive titers may be due to cross reaction with antibodies of nonpathogenic 

Mycoplasma or other agents. Only sporadic samples were positive, and there were no clinical findings or 

histopathologic changes of M. arthritidis infection in animals with positive titers. Accordingly, sporadic 

M. arthritidis-positive titers were considered to be false positives. 
277 

APPENDIX M 

CODEINE TOXICOKINETICS IN RATS 

DURING A TWO-YEAR DOSED FEED STUDY 

Jinhua Yuan, June K. Dunnick, Evelyn R. Barnes, and John W.A. Findlay 

Drug Metabolism and Disposition 22, 14-20 (1994) 

(Reproduced with permission of the 

American Society for Pharmacology and Applied Therapeutics) 
278 Codeine, NTP TR 455 

0090-95S6/94/22014014$02.00/0

DRUGMETABOLISM AND DISPOSITION Vol. 22, No. I 

Copyright(D 1994 by The American SocietyTor Pharmacology and Experimental Therapeutics Printed in U.S.A. 

CODEINETOXlCOKlNETlCS IN RATS DURING A TWO-YEARDOSEDFEEDSTUDY 

JlNHUAYUAN,JUNE K. DUNNICK,EVELYN R. BARNES,ANDJOHN W. A. FINDLAY 

National Instiluteof Environmental Health Sciences (J.Y., J.K.D.); and the Division of Pharmacokinetics and Drug Metabolism, 

Burroughs WellcomeCo. (E.R.B.,J.W.A.F.) 

(Received March 15,1993; accepted July 7,1993) 

ABSTRACT: 

Codeine toxicokinetics In F344 rats of both sexes were determined 

during a 2-year chronic toxicology study using dosed feed as the 

exposure route with a 12-hr light/dark cycle starting at ROO a.m. 

Rats were allowed to access to dosed feed formulations ad libitum 

with codeine concentrations at 0, 000, 800, and 1600 ppm. Blood 

samples were collected from Individual rat on days 7, 21, and 90 at 

200 p.m., 11:OO p.m., 390 a.m., and ROO a.m. Additional samples 

were collected at 16 and 24 months between 6:OO-8:OO a.m. Plasma 

concentrations of codeine and morphine were determined directly 

by radioimmunoassay. Concentrations of their conjugates were determined indirectly by measuring the total amount of free codeine 

and morphine released after samples were treated with 8-glucuronCodeine is one of the opioid alkaloids that has been used as 

an effective analgesic and antitussive drug for centuries (1-3). 

However, little is known about the carcinogenic potential of 

codeine and the related morphine group of chemicals. Because 

of the widespread use of codeine and the lack of knowledge of 

its carcinogenic potential, the National Toxicology Program has 

conducted long-term rodent studies of codeine. In the 2-year 

chronic rodent studies, codeine was administered in the feed to 

F344 rats of both sexes at concentrations of 0, 400, 800, and 

1600 ppm. (This dose range was selected to include a low dose 

with little or no toxic effects and a high dose that was -100times 

the human dose on a mukg basis and was considered the 

maximum dose that animals could tolerate over the 2-year dosing 

period.) Because humans may ingest codeine-containing drugs 

3-4 times/day, the feed route was chosenfor the rodent bioassay. 

Comparison of the metabolism and toxicokinetics in the experimental studies and humans serves as one basis for extrapolating the results from animal studies to humans. This study 

reports the findings on the resulting plasma concentrations of 

codeine and its various metabolites in rats during 2 years of 

dosed-feed exposure and compares the results to those obtained 

in human pharmokinetic studies. 

Materials and Methods 

Chemicals. Codeine (lot 471-NIN-001/A, purity > 99%) was manufactured by Penick Corporation (Newark, NJ). [I-’HlCodeine (specific 

activity, 20 Ci/mmol) and[ 1,6,7-’H]dihydromorphine (specific activity, 

81 Ci/mmol) were purchased from Amersham Corp. (Arlington Heights, 

IL). Details of antisera production and of other necessary chemicals can 

be found elsewhere (5). 

Formulations. Dosed feed formulations were preparedbyblending 

codeine with NIH-07 feed (Zeigler Brothers, Gardnen, PA) to concenSend reprint requerlr to: Dr.Jinhua Yuan, K221. Pharmacokinetics. Eioanalytical and Radiochemistry Function, Searle. 4901 We Parkway,Skokie, IL 

60077. 

idase. Results indicated that plasma concentrations of both codeine 

and morphine steadily decreased from day 7 to 16 months and then 

rebounded at 24 months. Results also indicated that plasma concentrations of both codeine and morphine correlated well with the 

amounts of codeine added to the feed. Bioavailability of codeine 

using the dosed feed route increased with dose, varying from 10% 

to 25%. which was somewhat higher than the previously reported 

-8% bioavailabilily using the gavage route. Concentrations of conjugated codeine were very low, whereas concentrations of conjugated morphine were very high. These results suggested that de 

methylation of codeine to morphine in rats Is the main metabolic 

pathway and was maintained over the course of the study. 

trations of0,400,800, and 1600 ppm. Stability and homogeneity of the 

prepared formulations,as well as the concentrations of codeine in feed, 

wereconfirmedby takingsamplesfromthe feed formulations and 

analyzing the codeine content prior to use.Feedsamples (10 g) were 

extracted for I5 min with 50 ml of acidified aqueous methanol solution 

(18 ml concentrated HCI and 1387 ml methanol in 595 ml water). The 

extractwas clarified by centrifugation for5 min at 1500 rpm. An aliquot 

of the supernatant (15 ml) wasfirstdiluted with water (I5 ml), then 

washed withmethylene chloride (IS mi).Afterdiscarding the organic 

phase, the upper aqueous layer (25 ml) was made basic by adding I N 

NaOH (2 ml) and was reextracted with methylene chloride (20ml). The 

resultingorganic phase (10 ml) was evaporated to dryness, and the residue 

was reconstituted in 3 mlof methanol containing propiophenone (200 

pg/ml) as internalstandard,and 7 mlof a diluting solution (1.01 g 

sodium heptane sulfonate and 5 ml acetic acid in 495 mlwater). The 

reconstitutedsolutionwasfiltratedandtheninjected (23 pl) into the 

HPLC system. The HPLC system contained a UV detector operated at 

280nm and a Brownlee RP-18 column (IO pm, 100 mm X 4.6 i.d.. 

AlltechAssociates,Inc.,Deerfield, IL) preceded by a guard column 

(Brownlee RP-18, 30 mm X 4.6 id.). The mobile phase (1 ml/min) was 

watecmethanol(63:37, v/v) containing glacial acetic acid(I %, v/v) and 

sodium heptane sulfonate (0.005 M).The standard curve was obtained 

by analyzing spiked feed samples. 

Animals. Groups of male and female Fischer 344 rats (4-week old) 

were obtained from Taconic Farms (Germantown,NY) and were quarantined for 15 days before the start of the study. A light cycleof7:00 

a.m.-7:00 p.m. was used in the animal facility. Rats of both sexes were 

randomized by bodyweight and were assignedto a specific blood 

collection period: days 7,21,90, and months 16 (65 weeks) and months 

24(104weeks).Ratswereexposed to codeine via thedosedfeed od 

libitum. Concentrations of codeine (freebase) in feed were 0,400.800, 

and 1600ppm. Body weights and feed consumption were recorded during 

the study. On days 7, 2 I, and 90, groups of 5 male and 5 female rats 

from each of the codeine-treated groups were bled via the orbital sinus, 

under C02anesthesia (70% COl, 30%01), at 7:00 p.m., 1 1:00 p.m., 3:00 

a.m. (next day), and7:00 a.m. (next day) of the dark cycle of the animal 

facility (7:OO p.m.-7:OOa.m.). Control animals were bled only at I1:00 

p.m. on days 7. 21, and 90. At 16 and 24 months, blood samples wcre 

collected using the same procedure from 5 rats of each of the codeine14 
279 

Codeine Toxicokineticsin Rats During a Two-year Dosed Feed Study 

CODEINE TOXlCOKlNETlCS IN F344 RATS 15 

TABLE 1 

Daily feed and codeine consumption of ratsin chronic codeine dosedfied studies' 

Melc Female 

Dou 

Day 1 Day 21 Day90 Month 16 24 Month Day 7 Day 2 I Day 90 16 Month 24 Month

ppm 

Body Weight (9) 

0 151 f 28 232 f 22 361 f 13 471 f 24 41 1 f 43 124 f 13 148 f 2 217 f I5 336 f 19 342 f 33 

400 153 f 24 230 t 21 361 f 24 458 f 22 405 f 36 120 f 13 144 f 9 201 f 12 306 f 22 335 f 40 

800 I55 f 23 212 f 26 355 f 16 444 f 24 390 f 29 I19 f 13 147 f 9 204f II 302 f 21 315 f 40 

1600 146 f 23 203 f 20 330 f 18 400f24 362f 33 llOf 14 142 f 10 198 f 13 283 f 20 303 f 23 

Food Consumption (g/animal/day)b 

0 14.4 f 1.3 18.1 f 2.8 18.5 f 1.2 15.0 f 0.7 9.8 f 2.3 11.6 f 1.5 10.8 f 0.8 11.3 f 1.4 11.1 f 0.6 10.2 f 0.5 

400 13.0 f 2.5 18.2 f 4.3 18.6 f 2.6 15.5 f 1.2 9.2 f 1.9 10.7 f 1.3 I 1.7 f 2.6 11.7 f 1.6 1 1.9 f 0.4 9.7 f 2.0 

800 13.4 f 1.7 17.0 f 3.7 17.7 f 2.8 15.2 f 1.6 11.6 f 1.3 10.0 f 1.3 11.7 f 3.2 11.3 f 2.1 12.0 f 0.7 10.0 f 0.8 

1600 12.3 f 1.3 17.2 f 4.0 17.7 f 3.8 15.7 f 1.0 10.3 f 1.3 9.Of 1.4 12.2 f 2.9 12.3 f 2.0 11.8 f 0.6 10.8 f 1.5 

Codeine lntake (mg/day/kg body weight) 

0 - - - - - - - - - 400 34.Of 8.4 31.7 f 8.0 20.7 f 3.2 10.8 f 1.0 7.3 f 1.6 35.6 f 5.6 32.4 f 7.5 23.2 f 3.5 12.4 f 1.0 9.3 f 2.2 

800 69.1 f 13.5 64.Ofl5.8 40.1f6.7 21.9f2.6. 19.Of2.6 68.6f 11.3 63.5f 18.0 44.2f8.6 25.Of2.4 20.3f3.0 

I600 133.9 f 25.0 135.1 f 34.0 86.Of 19.0 50.2f4.436.4f 5.7 130.2 f26.9 137.2f 34.4 99.5 f 17.6 53.4f4.6 45.6f 7.2 

Values are expressed as the meanf SD of 5 rats (0 ppm) and20 rats (400.800, and 1600ppm) for day7 to day 90; and of 60 rats for 16 months 

and of 30 rats for 24 months. 

Rats were housed S/cage and feed consumption for an individual rat was calculated based on the feed consumption of per cage. 

Day Ngld IL 

21 Ihy 21 

I 

I 

I I 

I 

I 

L

I 

I 1 4 1- T 

7AM 7PM 7w 7PM 7AM 7PM 7AM 

Time of Day 

FIG.I. Plasma concentration projib of unconjugafed codeine in male 

F344 rats during the dark cycleon days 7, 21, and 90 in a codeine 

dosedfied study. 

Curvesfor day 7 weregeneratedusing a dosedfeedmodel. 400 

ppm; 0,800 ppm; and0, 1600 ppm. 

treated groups at 6:00-8:00 a.m. Each rat was sampled only once. All 

blood'samples were drawn through heparinized microhematocrit tubes 

into EDTA-treated 5-mlVenoject blood collectiontubes,andwere 

centrifuged under refrigerated conditions to separate the plasma. The 

harvested plasma samples were stored at -70% until they were analyzed.

7AM 7PM 7AM 7PM 7AM 

lime of Day 

FIG.2. Plmma concentration projles of unconjugated codeine infide 

F344 rats during the dark cycleon days 7, 21. and 90 in a codeine 

dosedfied study. 

Curves for day 7 weregeneratedusing a dosed feedmodel. 4 so0 

ppm; 0.800 ppm; and 0, 1600 ppm. 

total morphine concentrations, which are the plasma conckntrationsd 

codeine and morphine after samples being treated with fi-glucuronidase. 

Modeling. Plasma concentrations of unconjugated codeine on day 7 

were fit to a dosed feedmodelassuming linearpharmacokinetics as 

previously described by Yuan (4). Briefly, the modeltreatsa dmed feed 

study as a series ofconsecutivegavagestudies,withvariable dosa 

Analyzing Plasma Samples.Plasma samples were analyzed for codeinerepeated at a small time interval suchas 0.5 hr. The theoretical gavage

and morphine by radioimmunoassay procedures as previously described dose at each time interval was determined from the daily toul chemical 

(5). The concentrations of their conjugates were determined indirectly intake (experimentally determined), aswell as the percentage ofthe daily

by measuring the total amount of fret codeine and morphine released feeding activity occurring during the same time interval. The plasma

after samples were treated with ,9-glucuronidase using the same radio- concentrations of test chemical from each theoretical small gavagedosc 

immunoassay procedures(5). Results are expressedas total codeine and at each time interval were calculatedbased on the small gavage dw and 
Codeine, NTP TR 455 280 

16 YUAN ET AL. 

TABLE 2 

EstimatedA UC valuesfor codeine in rat dosedfeed codeine studieso 

Day 7 Day 21 Day 90 Dose Sex in Feed Codeineb Total Codeine' Codeine Total CodeineCodeine Total Codeine 

PPm ng. hrJml ng.hrJml ng.hrlml ng.hrJml ng.hrjml ng.hrJml 

Male 400350 f 30 358 * 33 143 f 21 175 f 37 224 f 26289 f 26 

800 1,566 f I17 1,567 f 120 635 f 49 632 f 46 1,237 f 118 1,492 f 144 

1600 4,894 f 336 4,958 f 387 2,668 f 236,689 f 234 1,681 f 136 3,042 f 150 

Female 400 572 f 41 581 368 f 35 394 f 24 395 f 26 f 29 639 f 48

1,633 800 f 90 1,676 f 104 1,319 f 77 1,380 f98584 f 65 1,547 f 101 

1600 4,820 f 2924,870 f 270 3,631 f 3533,727 

'Unconjugated and conjugated codeine. 

" AUC values are expressed as mean f SD for the 12-hr dark cycle period.

Unconjugated codeine. 

lay 21 

I

Day Nirn

I

Day Nghl 

lay 90 

f 3803,350 f 391 3,876 f 395 

Day

L

90 

Day NiM 

I 

7AM 7PM 7AM 7PM 

lo 1 

?AM 7PM 7AM 7PM 7AM 7PM ?AM

Time of Day 

FIG.3. Plasma concentrationprojiles of unconjugated and conjugated 

morphine in maleF344 rats during the dark cycleon days 7. 21. and 90 

in a codeine dosedfied study. 

Curves for day 7 of unconjugated morphine were generated using a 

dosed feed model. Filled symbols represent unconjugated morphine, and 

open symbol.^ represent total morphine. .,400 ppm; 0, 800 ppm; and 

A, I600 ppm. 

the unit impulse response function, then summed to yield the plasma 

concentrationsof test chemical profiles in a dosed feed study. The unit 

impulse response function, defined as the plasma concentration profiles

obtained after a bolus gavage dose of I unit of test chemical, can be 

experimentally determined after bolus gavage doses. Specifically,the unit 

impulse response funnion for codeine is basedon the followingequation: 

with DO= I pg/kg, (I) 

Time of Day 

FIG.4. Plasma concentrationof unconjugared and conjugated morphine 

profilesinfemale F344 rats during the darkcwle on days 7.21. and 90 

in a codeine dosedfeed study. 

Filled symbols represent unconjugated morphine, and open symbols 

represent total morphine. W, 400 ppm; 0,800 ppm; and A, 1600 ppm. 

where K,and K. are the elimination and absorption rate constants and 

are equal to I .26 and 33 hP, respectively; V,,is the apparent volume of 

distribution and is equal to 3.8 litenfig, assuming codeine is evenly

distributed between plasma and red blood cells (6). The value for bioavailability (4was initially set to be 8%and was finally determined by 

the model. The dose unit (&kg) was necessary to express c6(l) in ng/

ml, a concentration unit used throughout this study. 

As previously stated, the model used a daily feeding activity curve to 

calculate the theoretical gavagedose at each time interval (input function)

that assumesthat feedingactivity was directlyproportional to the amount 

of feed consumed. Ideally, the feeding activity curve should be obtained 

from the treated animals. However, because of the experimental dimculty, the feeding activity curve collected from control animals (4. 7) was 
Codeine Toxicokinetics in Rats During a Two-year Dosed Feed Study 281 

Dou Sex 

in Feed 

PPm 

Male 400 

800 

1600 

Female 400 

800 

1600 

CODEINETOXlCOKlNETlCS IN F344 RATS 17 

TABLE 3 

Estimated AUC values for morphine in rat dosedfeed codeine studies" 

Day 7 Day 2 I Day 90 

Morphineb Total Morphine' Morphine Total Morphine Morphine Total Morphine 

ng. hrlml ng. hrlml mg. hrlml ng. hrlml ng. hrlml ng. hrlml 

427 f 17 3,752 f 296 472 f 15 3,422 f 122 368 f 18 3,206 f 368 

I ,O I8 f 32 13,728 f 1,427 676 f 22 3,944 rt 180 764 f 30 7,050 f 49 I 

2,069 f 155 24,970 f 1,357 1,429 f 135 15,692 f 1,883 1,449 f 35 13,102 f 1,195 

382 f 8 3,638 f 143 394 f 16 2,900 f 87 275 f 25 5,084 f 5 14 

867 f 76 18,236 f 1,014 840 f 36 3,778 it 238 609 f 27 12,002 f 470 

1,447 f 97 22,506 f 1,325 1,508 f 105 13,944 f 1,362 1,079 f 89 24,254 f 1,647 

a AUC values are exoressed as mean f SD for the 12-hr dark cvcle -. wriod.

Unconjugated morphine.

Unconjugated and conjugated morphine. 

Months of Exposure 

FIG.5. Plasma concentrationsof codeine and morphine over the2-year rat codeine dosedfeed study. 

Samples were collected at600-8:00 a.m. A,male rat; 0,female rat; -, 

actuallyusedin the model. Thisisjustifiedbecause concentration of 

codeine in diet was low and the presence of codeine had little impact on 

the feed palatability at early stages (tableI). Moreover, model simulation 

indicated that slight changes in the feeding activity curve or even the 

absorption half-life had little effea toward the achievable plasma concentrations(4). 

Chootinga unit impulse response function for unconjugated morphinetions of codeine and morphine were evaluated by four-way ANOVA as was more difficult than for codeine, because morphine is a metabolite of a function of dose, sex, the day of sampling, and the time of sampling

codeine.From a theoretical point of view, theunitimpulseresponse using the program Data Desk@(Odesta Corp., Nonhbrook, IL). 

function for unconjugated morphine shouldat least contain three exponential terms representing absorptionof codeine, convenion of codeine Results 

to morphine, and elimination of morphine, respectively. However, the Some dose-related decreases in body weight gain were observed experimentallyobtainedplasmaconcentrations ofmetabolitesrarely

allowthese threeexponentialcomponents to be separated. Because on days 7, 21, and 90 and were more obvious at 24 months 

codeine is rapidly absorbed and metabolized, these two processes can be (table I), which might be related to opiate dependence. Based on

mathematically expressedas one exponential term. Hence,eq. I can also the feed consumption data in table 1, codeine at the doses used 

be used as the unit impulse response function for unconjugated morphine had very minor impact on thepalatability of the dosed feed. The 

except that K. should be viewed as the apparent absorption-formation 

rate constant. The values of K,,K., and Vd used in the model were 0.77 

hr-l, 1.26 hfl. and 10.8 liten/kg based on literature (5, 8), respectively. 

The bioavailability was initially setto be 10% and was finally determined 

from the model. Again, the feed activity curve from control animals(4, 

7) was used in the model. 

400 ppm; . . . . ., 800 ppm; ---, 1600 ppm. 

Data Analysis. AUC1 values of codeine, morphine, and their conjugatesover the 12-hr bloodsamplingperiodweredeterminedby the 

trapezoidal rule. The standard deviation of AUC was estimated based 

again on the trapezoidal rule (9). T,. and C,, were directly determined 

from the plasma concentration data. Rest was used to determine if the 

calculated AUC values weresex- or dosedependent. Plasma concentradaily intake of codeineincreased proportionally with the codeine 

concentration in feed. Daily intakemy weight was sex-independent (ANOVA, p > 0.05) and was gradually decreased over 

' Abbreviations used are: AN, area under the cum;T-, time to reach peek 

concentration;C-, maximum concentration;ANOVA. analysisof variance. 
282 Codeine, NTP TR 455 

18 YUAN €T AL. 

the course of study because of the increase in the body weight of 

the animal. 

Plasma concentrations of unconjugated codeine increased with 

the daily amount of codeine consumed and fluctuated over the 

course of the study. The peak plasma concentrations of codeine 

occurred at I1:00 p.m. on day 7 (figs. 1 and 2). Overall, plasma 

concentration profiles of unconjugated codeine were similar in 

both sexes. ANOVA analysis indicated that codeine plasma 

concentrations were dosedependent ( p < 0.01). In most cases, 

female rats had higher plasma concentrations of codeine and its 

metabolites than males. Plasma concentrations of unconjugated 

codeine consistently decreased from day 7 to day 90 (ANOVA, 

p < 0.01), which may be related to the decrease ofcodeine intake 

over the course of the study (table 1). However, because the drug 

intake on day 7 and day 21 were comparable, the reduction in 

plasma codeine concentrations may also have resulted from 

enzyme induction. 

The calculated AUC values of unconjugated codeine are listed 

in table2. AUC values increased with dose (F-test, p < 0.01) and

3 varied considerably throughout the study. However, the calcu3 lated AUC values generally decreased as the study progressed, e which indicated that codeine did not accumulate during the 

d study (table 2). Female rats tended to have larger AUC values .- -=0.01). 0 

.+ (ANOVA,p -

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s 3 3 3 e phine was a major metabolite of codeine in the rat. Peak concentrations of unconjugated morphine in the 1600 ppm dose group 

reached 254 ng/ml on day 7. Plasma concentrations of unconjugated morphine decreased from day 7 to day 90, although the 

extent of the reduction was less dramatic than for those of the 

parent drug. The unconjugated morphine AUC data (table 3)

indicated that exposure to morphine in the plasma of these rats 

was generally somcwhat less that to codeine (table 2). Unconjugated morphine AUC values approximately increased linearly 

Theoretical plasma concentration profiles on day 7 predicted 

by the model are also shown in figs. 1 and 2. The prediction was 

reasonably consistent with the study results. Based on the model 

results from day 7,bioavailability of codeine from the diet was 

estimated to be 10, 24, and 25% for 400, 800, and 1,600 ppm 

dose respectively. groups,

Plasma concentrations of conjugated codeine (presumably codeine-6-glucuronide) were also determined. Results indicated 

that most codeine was present in plasma as the unconjugated 

form, and theconcentrations of conjugated codeine were very 

low. Plasma concentrations of total codeine (unconjugated and 

conjugated) were generally higher in females than in males, but 

the difference was less consistent than for free codeine. The time 

courses of plasma concentrations of total codeine (data not 

presented) were very similar to those shown in fig. 1. As expected, 

the estimated AUC values of total codeine were slightly higher 

than the AUC values of unconjugated codeine (table 2) and also 

increasedwith dose (F-test, p < 0.01). 

Unconjugated morphine plasma concentration data are summarized in figs. 3 and 4. Plasma concentrations of unconjugated 

morphine were generally higher than those of unconjugated 

codeine in the 400 ppm dose group. but lower than the free 

codeine concentrations at the twohigher dose groups. This 

finding is also in agreement with earlier observations that mor3 3 3 1 6 7 withdosefrom 400 to 1600 ppm. s as 3m For male rats, morphine plasma concentration profiles on day 8

1 %& 8 8 2 8 7 were also predicted by the model. Again, the prediction was 

0 * c 5 b 4 reasonablyclose tothe experimentally determined concentrations (fig. 3, day 7).Based on the modeling results, the bioavail- 
Codeine Toxicokinetics in Rats During a Two-year Dosed Feed Study 283 

CODEINETOXlCOKlNETlCS IN F344 RATS 19 

Morphine-3glucuronide 

Morphine-Gglucoronide "1

Norcodeine-Gglucuronide 6glucoronide 

l3H 

FIG.6. Merabolicpalhwap of codeine in rats (14). 

I

ability of morphine from codeine diet was estimated to be 15, of total morphine peaked at 16 months and were still relatively 

20, and 20% for 400, 800, and 1600 ppm dose groups, respec- high at 24 months. In mostcases,femaleratsshowedhigher 

tively. For female rats, the prediction was not conducted becauseplasma concentrations ofcodeineand its metabolites than males. 

of unavailability of the estimated values for K., K, and Vd (5). Discussion Total morphine plasma concentrations are also summarized 

in figs. 3 and 4. It is immediately clear that conjugated morphine Pharmacokinetic studies of codeinein rats and humans have 

(presumably morphine-3-glucuronide) circulatedin rat plasma been reported (5, 6, 10-1 3). and the main metabolic pathways 

at much higher concentrations than either codeine or the primary of codeine in rats and humans were also identified (14). Based 

metaboIite,morphine, in their unconjugated forms.Thiswas on the literature andthe current findings, the metabolic pathways 

also consistent with earlier studies. Mean total morphine concen- of codeine in rats are summarized in fig. 6. The rate constants 

trationsfrequentlyexceededthecorrespondingunconjugated 

morphine concentrations by 10- to 20-fold or more. Peak. concentrations of total morphineranged up to 3 162 ng/ml, indicating considerably greater exposure of rats to conjugated morphine. 

Total morphine AUC data are also presentedin table 3. These 

valuesweremuchhigher than the AUC values for any other 

compound measured in the study, again confirming that exposure of these rats to morphine andits conjugated metabolite was 

muchgreaterthan to the parentdrug,codeine.Thus,total 

morphine AUC values were all multiples of the corresponding 

data for total codeine with the higher multiples being noted at 

the lower dose groups.

The distribution of codeine and codeinederived metabolites 

in plasma (mean f SE, N = 360) based on data from day 7 to 

day 90 was as follows: free codeine (1 1.8 f 0.47%), conjugated 

codeine(1.42 f O.l6%), free morphine(9.56 f 0.30%).and 

conjugated morphine (77.19 f 0.63%). These data further indicated a high exposure to conjugated morphine in rats. 

Plasma concentrations of codeine and its metabolites betweenand after enzyme treatment of plasma samples) was much less 

7:OO-8:00 a.m. over the course of the 2-year study are shownin 

fig. 5. Plasma concentrations of codeineand total codeine steadily decreased from day 7 to 16 months and then rebounded at 

24 months. Plasma concentration of morphineshowed a steady 

decrease from day 7 to 16 months then held relatively constant 

between16 and 24 months. In contrast, plasma concentrations 

assigned for various pathways were based on the publisheddata 

(5, 6). A comparison of CmLIand T- of plasma codeine and 

morphine in various studies in humans and rats are shown in 

table 4. The differences ofC, in reported human and SpragueDawley ratstudies might berelated to the different analytical

methods used in thestudies [Le. HPLC with electrochemical 

detection (6, 12) vs. radioimmunoassay(5, 15)) It can beseen 

that C,, for codeine in rats receiving 800 ppm codeine in the 

feed is comparable with the mean of C- achieved in humans 

receiving an oral dose of 60 mg codeine phosphate or codeine 

sulfate. On the otherhand, because theC- values ofthe codeine 

metabolite, morphine, obtainedin the rat study are much higher 

than the values obtained in the human study, Odemethylation 

must be the main metabolic pathway in rats but only a minor 

route in humans (15, 16).This difference may further suggest 

that the rate of Odemethylation of codeine in humans is much 

slower than in rats. Exposure of rats to codeine-6-glucuronide 

(indicated by the minor differences in codeine measured before 

than that in humans. In contrast, exposure of rats to morphine 

and its conjugates was much greater than that in humans (table 

4).

Thehigherplasmaconcentrationsofconjugatedmorphine 

maybe attributed to its smaller Vd because of itsenhanced 

aqueous solubility. Like other glucuronide conjugates, it is also 
Codeine, NTP TR 455 284 

20 YUAN ET AL. 

possible that conjugated morphine may be subject to enterohepatic circulation (1 7). The presence of high plasma concentrations of morphine conjugates might have a significant impact. on 

the interpretation ofthe rat codeine chronic study results, because 

it has been demonstrated that morphine conjugates also possess 

opioid pharmacological effects in humans (2). 

Study results also indicate that the bioavailability of codeine 

in feed is greater than the -8% bioavailability of codeine obtained after gavage administration of codeine phosphate (6). This 

is especially true when the concentration of codeine in the diet 

is high (e.g. the estimated bioavailability of codeine is -25% at 

1600 ppm). This enhanced bioavailability of codeine might be 

explained by the increase in hepatic perfusion rate as the results 

of the presence of food in the gastrointestinal tract. It is well 

known that an increase in hepatic perfusion rate will decrease 

hepatic transit time, which in turn reduces the time available for 

Odemethylation of codeine in liver during the first pass (1 8). 

Therefore,thebioavailability of codeine at higher doses was 

improved. 

The changes of plasma concentration of codeine and its metabolites over the course of this study (fig. 5) suggest that metabolism of codeine and morphine might be increased between day 

7 and 16 months. This is evident, because plasma concentrations 

of codeine and total codeine were lowest at 16 months, whereas 

the total morphine concentrations were highest despite the decreased intake of chemical on a unit body weight basis (table I). 

At 24 months, the metabolicrate for rats is expectedto be slower 

because of the effects of aging on the metabolismof many 

compounds. This is also consistent with the observation of the 

rebound of plasma concentrations of codeine and morphine at 

24 months. 

In conclusion, exposure of rats to codeine through the dosed 

feed route did not changethe metabolic pathways, with 0-

demethylation of codeine remaining the main metabolic pathway. At 800 ppm, theC,, of codeine in rats is comparable with 

the C, of codeine in humans observed after a therapeutic dose 

of codeine. Accumulation of codeineor its metabolites was not 

observed over the courseof the study. Administering codeine via 

the diet improved the bioavailability of codeine and bioavailability increased with the concentration of codeine in the diet. 

Plasma concentrationsof codeine and its metabolites increased 

with the codeine concentration in dosed feed. Peak concentrations of codeine and its metabolites occurred between 11 p.m. 

and 7:00 a.m. Prolonged exposure of rats to codeine resulted in 

decreased plasma concentrations of codeine and increased total 

morphine concentrations. 

Acknowledgments. Special thanks to Microbiological Associates, Inc., for dosing and collecting the samples. Thanks also to 

Dr. H. B. Matthews. Dr. L. T. Burka,Dr. K. Dembyatthe 

National Institute of Environmental Health Sciences, and Dr. K. 

Dix at CIIT for reviewing this manuscript. 

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