studies of tricresyl phosphate - National Toxicology Program

321 Pages·2000·24.19 MB·English

NATIONAL TOXICOLOGY PROGRAM
Technical Report Series
No. 433
TOXICOLOGY AND CARCINOGENESIS

STUDIES OF TRICRESYL PHOSPHATE

(CAS NO. 1330-78-5)

IN F344/N RATS AND B6C3F, MICE

(GAVAGE AND FEED STUDIES)
FOREWORD

The National Toxicology Program (NTP) is made up of four charter agencies of the U.S. Department of
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the National Instituteof Environmental Health Sciences (NIEHS), National Institutes of Health; the
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The studies described in this Technical Report were performed underthe 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 compliancew ith Food and Drug Administration (FDA) Good
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NTP TECHNICAL REPORT
ON THE
TOXICOLOGYANDCARCINOGENESIS .
STUDIES OF TRICRESYL PHOSPHATE

(CAS NO. 1330-78-5)
IN F344/N RATS AND B6C3F, MICE
(GAVAGE AND FEED STUDIES)
NATIONAL TOXICOLOGY PROGRAM
P.O. Box 12233
ResearchTrianglePark, NC 27709
September 1994
NTPTR 433
NIII I’ublication No. 94-3164
U.S. DEPARTMENT OF HEALTH AND HUMANSERVICES
Public HealthService
National Institutes of Health
2 Tricresyl Phosphate, NTP TR 433
CONTRIBUTORS

National Toxicology Program NTP Pathology Working Group
Evaluated and irltcryreted results atid report firldirrgs E\*ollratcd slicks, prcl>arcdp a tho log^ rcport ot1 rats
(5,Ocrohcr 1991)
K.M. Abdo, Ph.D.
C.J. Alden, Ph.D. P.K. Hildcbrandt, D.V.M., Chair
PATHCO, Inc.
G.A. Boorman, D.V.M., Ph.D.
W.W. Carlton, D.V.M., Ph.D.
D.A. Bridge, B.S.
Purdue University
J.R. Bucher, Ph.D.
J.R. Hailey, D.V.M.
S.L.Eustis, D.V.M., Ph.D.
National Toxicology Program
T.J. Goehl, Ph.D. M.M. McDonald, D.V.M., Ph.D.
J.R. Hailey, D.V.M. National ToxicologyProgrill11
J.K. Haseman, Ph.D. R.C. Sills, D.V.M., Ph.D.
R.D. Irwin, Ph.D. Natiollal Toxicology Program
G.N. Rao, D.V.M., Ph.D. K. Yosh,itomi, D.V.M., Ph.D.
J.H. Roycroft, Ph.D. Experimental PatholokT Laboratories
B.A. Schwetz, D.V.M., Ph.D.
C.C. Shackelcord, D.V.M., M.S., Ph.D.
D.B. Waiters, Ph.D.
K.L. Witt, M.S., OakRidgeAssociatedUniversities
J.R. Leininger, D.V.M., Ph.D., Chair
Pathology Associates, Inc.
Battelle Columbus Laboratories
E. Gaillard, D.V.M.
Conducted 16-day and 13-wceksntdics, cvalrratcd
ExpcrimentalPathology hboratories
pathologv findings
R.A. Herbert, D.V.M., Ph.D.
A.C. Peters, D.V.M., PrincipalInvestigator NationalTosicologyProgram
M.P. Jokinen, D.V.M.
M.J. Ryan, D.V.M., Ph.D.
Niltional Tosicology I’rogr;~m
P.C. Stromberg, D.V.M., Ph.D.
J. Kanno, M.D., Ph.D.
Tokyo Mcdical and Dcntal University
Conducted ?-year ShrdieS, evalitarcd M.M. McDonald, D.V.M., Ph.D.
potholop jhdings
National Toxicology Program
R. Miller, D.V.M.
P.J. Kurtz, Ph.D., PrincipalInvestigator
Chemical Industry Institute of Tosicology
M.J. Ryan, D.V.M., Ph.D.
A.W. Singer, D.V.M.
Biotechnical Services, Inc.
I’rq>ared Tcchical Reyort
Experimental Pathology Laboratories, Inc.
Provided pathology qrtaliy ossltrmce D.D. Lambright, Ph.D., l’rincipalInvestigator
J.R. Bcverly, B.A.
J.F. Hardisty, D.V.M., Principal Investigator P. Chaffin, B.S.E.
K. Yoshitomi, D.V.M., Ph.D. G. Gordon, M.A.
E. Gaillard, D.V.M.
E.S. Rathman, MS.
Dynamac Corporation
Prlyared qttali? asswance audits
S. Brecher, Ph.D., PrincipalInvestigator
3
CONTENTS

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

...............
EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVI‘lY 10

...................................
TECIINICAL REPORTS REVIEW SUBCOMMITTEE 11

...........
SUMMARY OF TECIINICAL REPORTS REVIEW SUBCOhIMIT?’EE CORIhIENTS 12

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

....................................................
hlATERMLSANDhlETIIODS 21

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

................................................
DISCUSSION AND CONCLUSIONS 67

................................................................
REFERENCES 73

APPENDIXA Summary of Lesionsin Male Rats in the 2-YearF eedStudy

..............................................
of Tricresyl Phosphate 79

APPENDIXB Summary of Lesionsin Fem:lle Rats in the 2-Yew Feed Study

..............................................
of Tricresyl Phosphate 111

APPENDIXC Summary of LesionsinMale Mice in the 2-Year Feed Study

..............................................
of Tricresyl Phosphate 145

APPENDIXD Summary of LesionsinFemaleMicein the 2-YearF eed Study

..............................................
of Tricresyl Phosphate 185

.................................................

APPENDIXE GenetTicoxicology 229

....................
APPENDIXF Organ Weights and Org:rn-Weight-to-Ilotly-\Yei~l~tRatios 237

..............................
APPENDIXG IIematology and ClinicaCl hemistry I<esolts 263

.............................................
APPENDIXI€ Neurobehavioral Studies 275

...................
APPENDIXI Chemical Characterization and Ilose Formulation Studies 293

APPENDIXJ Feed and Compound Consumption in the 2-YearF eed Studies

..............................................
of Tricresyl Phosphate 309

APPENDIXK Ingredients. Nutrient Composition. and Cont:rminant I.evels

in NIII-07 Rat and Mouse Ration ...................................... 315

APPENDIXL Sentinel Animal Program ............................................ 321
ABSTRACT

TRICRESYL PI-IOSPIIATE

CAS NO. 1330-78-5

Chemical Formula: C,IH210,P Molecular Weight: 368.36
Tricresyl phosphate is an organophosphatep lasticizer aspermatogenesis occurred in the testes of male rats
widely used in vinyl plastics and as a fire retardant that received 2,900 and 5,800 mgntg. Changesin
additive for hydraulic fluids. Toxicology and carcino- neurobehavioral parameters in groups that received
genesis studies wereconducted by administeringa 1,450, 2,900, or 5,800 mgkg werec onfounded by
mixed isomer preparationof 79% tricresyl phosphate mortality and reduced body weights andw ere not
esters (consisting of 21% tri-nz-cresyl phosphate, 4% attributed to a direct neurotoxic response.
tri-p-cresyl phosphate, less than 1% tri-o-cresyl
phosphate, and otheurnidentified tricresyl phosphate
esters) by gavage to groups of F344/N ratsa nd 16-DAY GAVAGESTUDY IN MICE
B6C3Fl mice for 16 days and 13weeks, and in feed to Groups of 10 malea nd 10 female mice received
groups of F344/N rats and B6C3F1 mice for 13weeks
tricresyl phosphate in corn oil by gavage at doses of
and2 years. Genetic toxicology studiesw erecon­
0,360, 730, 1,450, 2,900,or 5,800 mgntg body weight,
ducted in Salmonella t),phinzurium ancdultured
5 days per week, for a total of 13 or 14 doses in a
Chinese hamster ovary cells.
16-day period. Five males anda ll females that
received 1,450 mgkag l,l mice that received
2,900 mg/kg, andf our males and one female that
16-DAY GAVAGESTUDY IN RATS received 5,800 mgkg died before the end of the
Groups of 10male and 10 female rats received study. Final mean body weights of male mice that
tricresyl phosphate in corn oil by gavage at doses of received 1,450 and 5,800 mg/kg were significantly
0,360, 730, 1,450, 2,900,or 5,800 mgkg body weight, lower thant hat of the controls.Finalmean body
5 days per week, for a total of 13 or 14 doses in a weights of female mice that received 360, 730, or
16-day period. One female receiving 1,450 mgkg and 5,800 mgntg were significantly greater than thatof the
five males and eight females receiving 2,900 mgkg controls. Necrosis of the mandibular lymph node,
died before the end of the study. Final mean body thymus, ands p leenoccurred primarily in mice
weights of male and female rats that received 1,450, receiving 2,900 and 5,800 mgntg. Hindlimbgrip
2,900, or 5,800 mgkg were significantly lower than strengths of male mice that received 360 and
those of the controls. Necrosis of the mandibular 1,450 mgkg and male and female mice that received
lymph node, spleen, andt hymus occurred primarily in 730 and 5,800 mgkg were significantly lower than
rats receiving 2,900 and 5,800 mag. Diffuse those of the controls at the end of the study.
6 Tricresyl Phosphate, NTP TR 433
13-WEEK GAVAGESTUDY IN RATS 13,000ppm tricresylphosphatewereestimatedto
deliver daily doses of 55, 120, 220,430, or 750 mgkg
Groups o1f 0maleand10femaleratsreceived
bodyweight (malesa)nd651,202,304,30, or
tricresyl phosphate in corn oil by gavage at doses of
770 mgkg (females).Therewerenobiologically
0, 50, 100,200,400, or 800 mg/kg bodyweight. All
significant changesin neurobehavioral parameters in
ratssurvivedto the endo f the study.F inalmean
rats.
bodyweightsofmaleratsreceiving200,400, and
SO0 mgkg were significantly lower than that of the
Cytoplasmicvacuolization of the adrenacl ortex
controls.Cytoplasmicvacuolization of the adrenal
occurred in all exposed groups of rats. Hyperplasia
cortex occurred in all dosed groups and the severity
of ovarian interstitial cells and inflammation of the
increased with dose.Ovarianinterstitialcellhyper­
ovarian interstitium occurred ianll exposed groups of
trophyoccurredinalldosedgroups offemales.
females.Renalpapillaryedemaandrenalpapillary
Atrophy of the seminiferous tubules occurreidn male
necrosis occurred in 13,000ppmmalesandfemales
rats that received 400 and SO0 mgkg. There were no
and in 6,600 ppm females. Basophilic hypertrophyo f
biologicallysignificantchanges in neurobehavioral
parameters in rats. the pituitary gland parsdistalis and atrophy of the
seminiferotuubsuolecscurred in 6,6a0n0d
13,000 ppm males.
13-WEEK GAVAGESTUDY IN MICE Dose selection for the 2-year study in rats was based
Groups of 10m ale and 10 femalemicereceived olnowemreabnodwyeighttso;xricesponses
tricresyl phosphate in corn oil by gavage at doses of observed in the kidney, pituitary gland, and testis of
0, 50, 100,200, 400, or 800 m&/kg bodyweight.All males and the kidney of females exposedto 6,600 and
micesurvived to the endo f thestudy.Final mean 13,000 ppm; the presence of cytoplasmicvacuol­
body weights of male mice receiving 20m0@ g and of ization of the adrenal cortex in exposedmalesand
male and femalemicereceiving400 and SO0 m@g females;and the occurrenceo f ovarian interstitial cell
weresignificantlylower than those of the controls. hyperplasia in females exposed to 900 and 1,700 ppm.
Cytoplasmicvacuolization of theadrenaclortex
occurred in all dosed groups of mice and the severity
increased with dose. Ovarian interstitialcellhyper­ 13-WEEK FEEDSTUDY IN MICE
trophy was present in all dosed groups of female
Groups of 10 male and 10 female mice were feddiets
miceM. ultifocal degeneration of the spinal cord
containing 0, 250, 500, 1,000, 2,100, or 4,200 ppm of
occurred in males and females that received 100,200,
tricresyl phosphate. All mice survived to the end of
400, and SO0 mgkg, and multifocal degeneration of
the study. Meanbodyweights of 4,200 ppm males
the sciatic nerve occurredin males that received 200,
and of females exposed to 2,100 and 4,200 ppm were
400, and SO0 mgkg and females that received100,
lower than those of controls throughout the study.
200,400, and SO0 mg/kg. Hindlimb grip strengths of
Feed consumptionby females exposed to 1,000,2,100,
male mice that received 200, 400, or 800 mgkg were
or 4,200 ppm was lower than that by controls during
significantlylower than that of the controls at the
week12.Dietarylevelsof250, 500, 1,000,2,100, or
end of the study.
4,200 ppm tricresylphosphatewereestimatedto
deliveraverage daily dosesof45,110, lS0, 380, or
900 mg/kg body weight (males) and6 5,130,230,530,
13-WEEK FEEDSTUDY IN RATS o1r,050 mgkg (females). Interpretation of grip
Groups of 10 male and 10 female rats were fed diets strength changes observed in groups receiving 2,100
containing 0, 900, 1,700, 3,300, 6,600,or 13,000 ppm or 4,200 ppm were confounded by the reduced body
of tricresyl phosphate. All rats survived to the end of weights of these groups.
the study.F inalmeanbodyweightsofmalesand
females exposed to 6,600 and 13,000 ppm and females Cytoplasmicvacuolization of the adrenacl ortex
exposed to 3,300 ppm were significantly lower than occurred in all exposedgroupsofmale and female
those of controls.F eed consumption by male and mice with the exception of 250 ppm males. Papillary
femaleratsexposed lo 13,000 ppm waslower than hyperplasiaof thegallbladdermucosaoccurredin
that by controls during thefirstweekof the study. malemiceexposedto 500 ppm or moreandin
Dietarleyvels of 900, 1,7030,,3060,,6 0o0r female mice exposed to 1,000 ppm or more. Axonal
Tricresyl Phosphete, NTI' TR 433 7
degeneration occurredin males and females exposed &YEAR FEE11s1'UI)Y IN MICE
to 2,100a nd4,200 ppm and femalesexposedto Groups of 95 male and 95 female mice were fed diets
1,000 ppm. Renal tubule regeneration occurreidn all containing 0, 60, 125, or 250 ppm of tricresyl phos­
4,200 ppm male mice.
phate. After 3, 9, and 15monthsofchemicalexpo­
sure, up to 15 males and 15 females per group were
Dose selection fort he 2-year study in mice was based evaluatedforforelimb and hindlimb grip strength,
on the presenceo f axonal degenerationat concentra­ thennecropsied and evaluatedforhistopathologic
tions of 1,000 ppm or more and Lytoplasmic vacuoli­ lesions.
zation of the adrenalc ortex at concentrations of
500 ppm or more in males and in all exposed groups Survival, Mean Body Weights, and
of females. Feed Co1zsrrn~ptiort
Survival of exposed groups of male and female mice
was similar to that of the controls.The final mean
&YEAR FEEDSTUDY IN RA'I'S body weights of males and females receiving tricresyl
phosphatewere similar tothose of controls. Feed
Groups of 95 male and 95 female rats were fed diets
consumption by exposed groups of male and female
containing 0, 751,50, or 300 ppm otfricresyl
micewas similar to that by thecontrols.Dietary
phosphate. An additionalgroup o9f5male and
levels of 60,125, or 250 ppm tricresyl phosphate were
95 female rats were fed diets containing 600 ppm of
estimated to deliver average daily dosesof 7, 13, or
tricresylphosphatefor 22 weeks and thenreceived
27 mukg body weight (males) and S, lS, or 37 mgkg
only controlf eed.After 3, 9, and 15 months of
(females).
chemical exposure,up to 15 males and 15 females per
group were evaluated for forelimband hindlimb grip
Pathology Firzdings
strength, thennecropsied and evaluated for histo­
Thcre were no chemical-related increased incidences
pathologic lesions.
of neoplasms in mice.Ceroidpigmentationof the
Survival, Mean Body Weights, adrenaclorteoxccurred in all groups omfice
and Feed Consumption throughout most of the 2-year study,with the excep-
Survivaolefxposedratwsas similar to that of tionof60 and 125 ppm females at the 3-month
controls. The final mean body weights of all exposcd interim evaluatiohno; wever, the severiwtyas
markedly incrcascdin female mice receiving 250 ppm.
groups of males and females were similar to those of
the controls. Feedconsumption by exposedgroups Incidences of clear cell foci, Fdtty change, and ceroid
pigmentation of the liver were significantly increased
ofmale and femaleratswas similar to that by the
controlsD. ietarylevelso7f51,50, or 300 ppm in male mice that received125 or 250 ppm.
tricresyl phosphate were cstimated to deliver average
daily doses of 3, 6, or 13 mgkg body weight (males)
and 4, 7, or 15 mg/kg (females). GENE'I'IC TOXICOLO~Y
Tricresyl phosphatewas not mutagenic in Salnzonella
Pathology Findings fyphinzwi~~mstrains TA9T8 ,A 10T0,A153.o5r,
There were no chemical-related increased incidences TA1537, nor did it induce chromosomal aberrations
of neoplasms in rats.Cytoplasmic vacuolization of or sisterchromatidexchanges in culturedChinese
the adrenal cortexoccurred in 600 ppm males and hamstcrovarycells.These in tin-o assayswere all
1503,00, and 600 ppm females at the3-month conducted with and withoutexogenousmetabolic
interim evaluation. At 9 and 15 months, Lytoplasmic activation (S9).
vacuolization occurred only in fernale rats, primarily
in the 300 ppm group. Cytoplasmic vacuolization of
the adrenal cortex and ovarian interstitial cell hyper­ CONCLUSIONS
plasiaoccurred in female rats exposed to 300 ppm Undertheconditions of these 2-yearfeedstudies,
throughoutthe 2-year study and theincidence and there was no evidence of carcinogenicactivity* of
severity weresignificantly increased at the end of the tricrcsylphosphate in maleorfemaleF344/Nrats
study. that received 75, 150,or300 ppm. There was no
8 Tricresyl Phosphate, NTP TR 433
evidence ofcarcinogenic acrivir?,of tricresyl phosphate of the adrenal cortex and ovarian interstitial cell
in male or female B6C3Fl mice that received 60, 125, hyperplasia in female rats, increased incidences of
or 250 ppm. clfeoacru cse, ll fatty change, and ceroid pigmentation
of the liver in male mice, and increased severity of
Nonneoplastic lesions associated with exposure to ceroid pigmentation of the adrenal cortex in female
tricresylphosphateincludedcytoplasmicvacuolization mice.
l Explanation of Levels of Evidence of Carcinogenic Activity is on page10. A summary of the Technical Reports Review Subcommittee
comments and the public discussion on this report appears on page 12.
Tricresyl Phosphate, NTP TR 433 9
Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies of Tricresyl Phosphate
Variable Male
Female
nktle Female
F344/N Rats
F344/N Rats
156C3F,Mice 156C3F, Mice
Doses 0, 75, 150, or 0, 75, 150. or 0, 60, 125, or 0, 60, 125, or
300 ppm in feed 300 ppm in fecd 250 ppm in feed 250 ppm in feed
(Approximately 3, 6, (Approximately 4,7, (Approximately 7, 13, (Approximately8,
or 13 m@g) or 15 mghqg) or 27 m@g) 18, or 37 m&g)
Body weights Exposed groups Exposed groups &posed groups Exposed groups
similar to controls similar to controls similar to controls similar to controls
2-Year survival 32/51, 30/50, 35/50, 34/51, 38/53, 30/50, 43/51. 13/19, 44/19. 41/50, 3/50, 42/48,
rates 28/50 26/49 42/50 45/51
Nonneoplastic None Adrenal cortex: Liver:ceroid Adrenal cortex:
effects cytoplasmic pigmcntation (0152, ceroid pigmentation
vacuolimtion (14/51. 0/49. 30/49, 28/50); (severity grades -1.2,
12/53, 16/50. 36/50); clear cell Cocus (5/52, 1.6, 2.5, 3.9)
Ovary:interstitial 8/49,17/49,12/50);
hyperplasia (0/51, fatty change (6152,
0/53,0/50, 15/50) 10/49, 23/19, 22/50)
Neoplastic effects None None None None
Level of evidence No evidence No evidence No evidence No evidence
of carcinogenic
activity
Genetic toxicology
sa/motle//al )!phifntlrictmge nemutation:Negative with and witllout S9 in strains TA98, TA100, TA1535, and TA1537

Sister chromatid exchanges

Chinese hamster ovary cells in rirro: Negative with and without S9

Chromosomal aberrations

Chinese hamster ovary cellsin vitro: Negative with and without SF)