Environmental Health Perspectives Vol. 86, pp. 27-36, 1990



# Inhalation Toxicology and 

# Carcinogenicity of 

# 1,3-Butadiene in 

# B6C3F1 

# Mice Following 65 

# Weeks of 

# Exposure by 

# Ronald L. 

# Melnick,* J. 

# E. Huff,* Joseph 

# H. 

# Roycroft,* Billy J. 

# Chou,t and Rodney 

# A. 

# Millert

1,3-Butadiene, a large-production volume chemical usedmainly in the manufacture of synthetic rubber,

was found to induce multiple-organ carcinogenicity in male and female B6C3F1 mice at exposure concentrations (625 and 1250ppm) equivalent toandbelowtheOSHAstandardof1000ppm. Sincethis studywas terminated after 60 weeks of exposure because of reduced survival due to fatal tumors, and because dose-responserelationships for1,3-butadiene-induced neoplastic andnonneoplastic lesionswerenotclearly established, a second long-term inhalation study of 1,3-butadiene in B6C3F1 mice was conducted at lower exposure concentrations, ranging from 6.25 to 625 ppm. Both the histopathological findings from animals dyingthroughweek 65 andtheresults ofevaluations ofanimals exposedfor40 and 65weeks arepresented in this report. Exposure to 1,3-butadiene caused a regenerative anemia at concentrations of 62.5 ppm and higher. Testicularatrophywas inducedat 625ppm, andovarianatrophywas observedat 20 ppmandhigher. Duringthe first 50weeks ofthe study, lymphocytic lymphomawas themajorcause ofdeathofmice exposed to 625 ppm 1,3-butadiene. Neoplasms of the heart, forestomach, lung, Harderian gland, mammary gland, ovary, andliverwerefrequentlyobserved in 1,3-butadiene-exposedmicethatdiedbetweenweek40 andweek 65 of the study. Studies in which exposure to 1,3-butadiene was stopped after limited periods were also included to assess the relationship between exposure levels and duration of exposures on the outcome of 1,3-butadiene-inducedcarcinogenicity. Inthese studies, lymphocytic lymphomas were inducedinmalemice exposed to 625 ppm 1,3-butadiene for only 13 weeks. The incidence oflymphocytic lymphoma inmale mice exposed to 625 ppm 1,3-butadiene for 26 weeks was two times that inmice exposed to 625 ppm for 13 weeks. However, when the exposure concentration was reduced by half to 312 ppm and the exposure duration extendedto 52weeks, the incidence oflymphocytic lymphomawasreducedby90%. Thus, themultipleofthe exposure concentrationtimes the exposuredurationdidnotpredictthe incidenceoflymphocytic lymphoma in mice. The early mortalities resulting from lymphocytic lymphomas in male mice exposed to 625 ppm 1,3-butadiene limited the expression of tumors at other sites. A clearer dose-response for 1,3-butadieneinduced neoplasia should be apparent from experiments in mice exposed to lower concentrations of this chemical for 2 years. 

## Introduction 1,3-Butadiene is a 

### colorless, 

### flammable, noncorrosive gas 

### (bp: -4.4 

### °C, Vp: 1900 

### mmHg at 20 

### °C) used

### mainly in 

### the 

### production of 

### synthetic rubber and 

### thermoplastic resins 

### (1). In

### production

### volume, 1,3-butadiene is one of 

### the top 25 

### organic chemicalsmanufactured in

### the United 

### States 

### (2), with an annual 

### production level of rubber

### grade 

### 1,3-butadiene in 

### the 

### U.S. of 

### approximately 2.5 

### billion 

### pounds (3). This gas 

### has also been identified in 

### automobile 

### exhaust, cigarette smoke, and in 

### incineration 

### products of fossil fuels 

### (4). 1,3-Butadiene was

*National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709. tBattelle Pacific Northwest Laboratories, Richland, WA 99352. Address reprint requests to R. Melnick, NIEHS, P.O. Box 12233, Research Triangle Park, NC 27709. 

### selectedforevaluationofits

### toxicologicand

### carcinogenic 

### potential because of its 

### large production 

### volume, its 

### potential forhumanexposure in

### facilities that

### produce or process 

### this 

### chemical, and 

### the lack of

### long-term toxicology

### and 

### carcinogenicitydata for 

### this 

### compound (5). The National 

### Institute for 

### Occupational Safety and Health 

### estimates that 

### about 65,000 workers are 

### potentially 

### exposed to 

### 1,3-butadiene (6), and

### the 

### U.S. Environmental Protection

### Agency estimates that between 5300 and 8200 

### workers are 

### exposed to 

### 1,3-butadiene in 

### facilities that either 

### produce the monomer or process the monomer 

### into 

### polymers (7). Recent surveys 

### conducted 

### by 

### NIOSH at monomer 

### and

### polymermanufacturingplants indicate 

### that most 

### occupational exposures to 

### 1,3-butadiene are less 

### than 20 ppm; 

### however, excursions in 

### certain 

### jobs were as 

### high as 374 ppm 

### (8). 1,3-Butadiene has 

### longbeenconsidered to

### have a 

### low,MELNICK ETAL.



### noncumulative toxicity in animals and 

### humans. The LC50 for 1,3-butadiene was 

### reported to be 270,000 

### mg/m3 (122,000 

### ppm) in mice 

### exposed for 2 hr and 285,000 

### mg/m3 (129,000 ppm) inrats 

### exposedfor4hr (9). 

### Carpenter et al. 

### (10) 

### exposedrats, 

### guinea

### pigs, rabbits, and dogs to 0, 600, 

### 2300, or 6700 ppm 

### 1,3-butadiene 7.5 

### hr/day, 6 

### days/week, for 8 

### months. 

### Slight growth re

### tardation and 

### cloudy swelling in 

### hepatocytes were observed in the 

### highest dose group; 

### there were no apparent 

### treatment-related 

### pathologic 

### changes in

### the adrenal gland, heart, 

### kidneys, 

### lungs, skeletal muscle, pancreas, 

### spleen, testes, or 

### ovares. 

### Exposure of

### humans to 2000 ppm 

### 1,3-butadiene or 

### higher for 6 to 8 hr caused 

### slight irritation of the eyes (9). 

### The National 

### Toxicology 

### Program (NTP) studied 

### the long-termeffects of

### inhalationexposureto 

### 1,3-butadiene inmice 

### (5,11). 

### Groups of 50 

### male and 50 

### female 

### B6C3F1 mice were 

### exposed 6 

### hr/day, 5 

### days/week to air con

### taining 0 

### (chamber 

### control), 625, or 1250 ppm 

### 1,3-butadiene. This 

### study, intended to 

### last for 103 

### weeks, was ended after 60 to 61 weeks 

### because of 

### cancer-related 

### mortalityin

### both sexes atbothexposure

### concentrations. There were 

### early induction and 

### increased incidences of 

### malignant lymphomas, 

### hemangiosarcomas of

### the 

### heart, alveolar-bronchiolar 

### neoplasms, and squamous cell neo

### plasms of the forestomach in 

### males and 

### females; and 

### acinarcell carcinomas ofthemammary

### gland, granulosa cell 

### neoplasms of the ovary, 

### and 

### hepatocellular neo

### plasms in 

### females. The 

### observation of

### high incidences of hemangiosarcomas of the heart was a 

### particularly un

### usual 

### finding, since

### the

### historical rate

### forthisuncommon 

### endothelial cell tumor is 

### about 0.04% in 

### untreated 

### B6C3F1 mice 

### and has 

### rarely been induced in 

### B6C3F1 mice in 

### long-term studies 

### (12,13). In 

### male 

### mice, doseresponses for 

### neoplasms of the 

### heart, 

### lung, and 

### forestomachwere not 

### clearlydemonstrated sincebothdoses 

### produced 

### similarresponses, whereas in

### female

### mice, the rate 

### of 

### malignant 

### lymphomas was 

### lower than 

### that in 

### males, and the 

### dose-responses for 

### other 

### neoplastic lesions were more 

### clearly defined. 

### Nonneoplastic lesions associated with exposure 

### of 

### B6C3F1 mice to 

### 1,3-butadiene included 

### epithelialhyperplasia ofthe 

### forestomach, endothelial 

### hyperplasia ofthe 

### heart, alveolar 

### epithelial 

### hyperplasia, hepatocellular necrosis, testicular 

### atrophy, ovarian 

### atrophy, and 

### nasal 

### cavity lesions 

### (chronic 

### inflammation, 

### fibrosis, and osseous 

### and 

### cartilagenous metaplasia in 

### the 

### olfactory re

### gion) in 

### high-dose males 

### (5,11). 

### Stimulated 

### by the unusual 

### effects of 

### 1,3-butadiene in 

### mice, the fact that the studies were 

### terminated 

### early, the sometimes unclear 

### dose-response relationships for various 

### lesions, andthe 

### importance of

### 1,3-butadiene as a 

### high-production volume 

### chemical with 

### potential human exposure, 

### additional 

### toxicology/carcinogenicity studies 

### ofvarious 

### durations, 

### including a 

### 2-year exposure, were 

### initiated to better 

### characterize dose-response relation

### ships for 

### 1,3-butadiene-induced 

### nonneoplastic and neo

### plastic lesions in 

### B6C3F1 mice. Data from 

### the first 65 weeks 

### (15

### months) ofthat

### study (earlydeathanimalsand 

### interim evaluations at 40 and 65 

### weeks) are 

### presented. Thedata

### fromthe 2-yearexposures arenotyetavailable and will 

### be reported 

### separately.


## Materials and 

## Methods
Chemical and Exposure 

### System Liquified 

### 1,3-butadiene of 99+% 

### purity, containing trace amounts of 

### t-butyl 

### catechol, a 

### peroxide inhibitor,

was obtained from Phillips Chemical Co. 

### (Borger, TX). Concentrations of the dimer, 

### 4-vinyl-1-cyclohexene, in the cylinder headspace were determined 

### by gas chro

### matographic analysis. Cylinders were not 

### retained for
use if the dimer content was greater than 150 ppm. 

### 1,3-Butadiene gas was 

### metered to 

### the exposure chambersviaadistributionsystemanddilutedinthe

### freshair chamber inlets. Chamber concentrations of 

### 1,3-butadiene were 

### monitored 

### continuously 

### during the exposure periods with a 

### Hewlett-Packard 5840 gas chro

### matograph equipped with a 

### flame ionization 

### detector 

### (oven temperature: 

### 120°C; 

### GC column: 12" x 1/8" nickel column

### packed with 1% 

### SP-1000 on 60/80 

### Carbopack B; carrier gas: 

### nitrogen, 20 

### mL/min). The 

### dailymean concentrations of 

### 1,3-butadiene, distributed 

### uniformly in the 

### chambers, were within 2% 

### of the target concen

### trations. 

### Animal Maintenance Five-week-old male and female 

### B6C3F1 mice, obtained from Frederick 

### Cancer Research Facility (Fred

### erick, MD), were 

### quarantined and 

### acclimatized for 10 

### days prior to 

### the start of the study. 

### Stainless steel 

### Hazelton2000 

### chambers (2.3

### m3) were

### used, and

### animals

were exposed in 

### individual wire 

### mesh cage units, each 

### havinga

### floorarea

### of106

### cm2. The chamberenvironment

was 

### maintained at 75 + 3°F 

### and 55 ± 15% 

### relative 

### humidity, with a 

### chamber air flow of 15 air 

### changes/hr. Fresh, 

### softened tap water and NIH-07 diet were available ad 

### libitum, except 

### when the feed was 

### removed 

### during the exposure 

### periods and 

### overnight 

### prior to 

### the 40- and 

### 65-week 

### necropsies.



### Treatment 

### Groups Groups of 70 male and 70 female 

### B6C3F1 mice were 

### exposed6

### hr/day, 5

### days/weekfor

### periodsupto 65

### weeks to 0, 6.25, 20, 

### 62.5, or 200 ppm 

### 1,3-butadiene; groups of 90

### maleand 90 

### femalemicewere

### similarlyexposedto 625 ppm 

### 1,3-butadiene. The additional animals in 

### the 

### 625-ppm exposure group were 

### included out 

### of concern 

### that 

### high mortality rates 

### observed 

### previously (5,11) at 

### this exposure concentration 

### might interfere with 

### the interim 

### evaluations at 

### weeks 40 and 65. 

### Fifty animals pergroupwere 

### includedfor

### evaluationof

### 1,3-butadieneinduced 

### carcinogenicity after exposure 

### for up to 103 

### weeks. The 

### 625-ppm concentration 

### corresponds to the 

### low exposure 

### level in 

### the 

### previous inhalation study 

### (5,11), while 6.25 ppm 

### and 20 ppm more 

### closely represent 

### occupational exposure levels to 

### 1,3-butadiene (8). 28INHALATION TOXICOLOGY OF 1,3-BUTADIENE IN MICE



### Animals were 

### observed twice 

### daily for

### moribundityand 

### mortality, and

### body

### weights were measured

### weekly for the first 13 

### weeks of the 

### study and at 4-week intervals 

### thereafter. Studies in which the exposure to 

### 1,3-butadiene was stopped 

### after limited exposure 

### periods (stop-exposure 

### studies, 

### SE) were included to assess the 

### relationship between exposure level and duration 

### ofexposure on the outcome of 

### 1,3-butadiene-induced carcinogenicity. Groups of 50 male mice were 

### exposed to one of the 

### following regimens: a) 625 ppm for 13 weeks 

### (SE 13 

### WK), 

### b) 200 ppm for 40 

### weeks (SE 40 

### WK), c) 625 ppm 

### for26

### weeks 

### (SE 26

### WK), or

### d) 312ppm

### for52weeks 

### (SE 52 

### WK). After the exposures were 

### terminated, these groups 

### of animals were 

### placed in control chambers for theremainderof

### the 104-week

### study. Thetotal exposure to 

### 1,3-butadiene 

### (concentration

### multipliedbydurationof 

### exposure, 

### CT) was 

### approximately equivalent for the first and secondtreatments (8000 

### and 8125 ppm-weeks), 

### andit

### providedabouthalfthetotal exposure

### giveninthe third and fourth treatments (16,224 and 16,250 ppm 

### weeks). 

### Hematology Blood 

### samples were 

### obtained at

### weeks 40 

### and 65 

### from the 

### supraorbital sinus 

### of 

### C02-anesthetized mice and 

### collected into 

### tubes 

### containing EDTA. Red 

### blood cell count 

### (RBC), whitebloodcell count 

### (WBC), hemoglobin concentration 

### (HGB), volume of 

### packed red cells 

### (VPRC), mean 

### corpuscular volume 

### (MCV), mean cor

### puscular 

### hemoglobin 

### (MCH), mean 

### corpuscular hemoglobin concentration 

### (MCHC), and

### platelet counts were 

### determined with an 

### Ortho E

### LT-8/ds 

### hematology 

### analyzer. 

### Histopathology
All

### moribund animals 

### and those 

### sacrificedat 40 and 65 

### weeks were 

### killed by 

### CO2 asphyxiation and then im

### mediatelynecropsied. Tissue 

### samples, preserved in 10% 

### neutral buffered 

### formalin, were 

### embedded in 

### paraffin, 

### sectioned, and stained with

### hematoxylin and eosin. The 

### following tissues were 

### examined 

### microscopically: gross 

### lesions and tissue masses, 

### bronchial, mediastinal, man

### dibular, and 

### mesenteric 

### lymph nodes; salivary gland; sternebrae 

### (including marrow); 

### thyroid; parathyroid; small 

### intestine; large 

### intestine; liver; gall bladder; prostate, testes, 

### epididymis, and 

### seminal 

### vesicles; ovaries; 

### lungs and mainstem bronchi; nasal cavity and tur

### binates; 

### heart; 

### esophagus; stomach; uterus; 

### brain; thymus; 

### larynx; trachea; pancreas; 

### spleen; 

### kidneys; adre

### nals; urinary bladder; pituitary; and mammary 

### gland.


### Statistics
Mean 

### body weights, organ 

### weights, organ 

### weight/ 

### body 

### weight ratios, and 

### hematology results of treated groups were compared to those of 

### control groups using 

### Dunnett's t-test. 

### The minimum level of

### probability ac

### cepted for 

### significance was p<0.05. 

## Results Survival, Body 

### Weight, and 

### Organ Weights Exposure 

### of B6C3F1 mice to 625 ppm 1,3-butadiene causeda

### sharpdecrease in survival ofmales 

### and females (Fig. 1). Increased 

### mortality was apparent in exposed males at about week 25 and in exposed females at about week30. Survivalwas reduced 50%byweek36 formales andby

### week46 

### forfemales. All

### mice exposedto 625ppm 1,3-butadiene were dead by

### week 65. Exposure ofmice to 200 ppm 

### 1,3-butadiene also caused a 

### reduction in 

### survival; 

### however, the rate of

### mortalityatthis exposure 

### levelwasmuchless than

### thatat 625ppm. 

### Atweek65 the survival 

### ofmice 

### exposed to 200 ppm 

### 1,3-butadiene was 43 of50 

### formales and 31 

### of50 for 

### females. Inthe 

### control and 

### lower exposure concentration groups, survival was 

### generally greater than 90% at week 65. 

### Survival data from the stop-exposure groups 

### (Fig. 2) indicate that although mortality due to exposure to 1,3-butadiene was concentrationrelated, themultiple of exposure 

### concentration times durationofexposure 

### (CT) did not predict the 

### probability of survival. Mortalities 

### occurred at a greater rate 

### formale mice exposed to 625 ppm 

### 1,3-butadiene for 26 

### weeks, compared to 312 ppm for 52 weeks 

### (equivalent total exposure), 

### and for male 

### mice exposed to 625 ppm for 13 

### weeks, compared to 200 ppm 

### for 40 

### weeks. Exposure of

### male or 

### female mice to 6.25 to 625 ppm 

### 1,3-butadiene for 65 weeks did not 

### produce any 

### significant 

### adverse effects on mean 

### body weight 

### gain. The initial mean

### bodyweight ofmale mice was 22.8 + 0.3 g and of 

### female mice was 18.5 + 0.2 g. 

### At week 65, the

mean 

### body

### weight ofcontrol male mice was 49.3 g, 

### and the mean 

### body weights of 

### the groups 

### of male mice 

### exposed to 

### 1,3-butadiene 

### ranged from46.0 to 47.4 g; 

### the

mean

### bodyweight ofcontrol 

### femalemicewas 43.5g, and 

### the mean 

### body weights of the groups 

### of female mice 

### exposed to 

### 1,3-butadiene 

### ranged from 42.9 to 49.7 g. At 

### the 

### interim evaluations at weeks 40 

### and 

### 65, 

### liver, brain, 

### kidney, spleen, 

### lung, heart, 

### thymus, and testis 

### weights were

### measured. The 

### onlysignificant concentra

### tion-related effect was a decrease in testis 

### weight for males 

### exposed to 62.5 ppm and 

### higher (Table 1). The testis

### weight/bodyweightratiowas decreasedin

### the 200 and 625 ppm exposure groups, 

### compared to controls. A 

### similar dose-related decrease in testis 

### weight was ob

### served at 

### the 65-week evaluation. 

### Hematology Hematologic indices were measured in

### blood samples taken frommale and femalemice at the 40- and 65-week 

### interim evaluations. For male mice at week 40, 

### significant 

### decreases were 

### observed in red blood cell counts, 

### hemoglobin

### concentration, and

### packedredcellvolume at exposure 

### concentrations of

### 62.5, 200, or 625ppm 

### 1,3-butadiene 

### (Table 2). Similar decreases 

### (data not 

### shown)

were 

### observed in 

### female mice 

### exposed to 625 ppm 

### 1,3-butadiene and furtherdemonstrate the dose-related anemia 

### caused

### by 1,3-butadiene inmice. These 

### changes 29MELNICK ET AL.

Males, 24 10o

0,9 00Oppm

08-

0.7 ,200 ppm i

> 06

o 05

-r m

- 0.4-

0

a.

- 0.2

0.1

-

30 40 50 60 70
WEEKS ON STUDY
FIGURE 1. Survival curves formale and female B6C3F1 mice exposed
to 200 or 625 ppm 1,3-butadiene for 65 weeks.

in male and female mice were not accompanied by increases in reticulocyte counts or in the frequency of 

### polychromatophilic cells in 

### peripheral blood, and they

mayindicate apartial orpoorlyregenerative response in the bone marrow of 

### 1,3-butadiene-exposed mice to reTable 1. Testis weights and testis weight/body weight ratios (x 1000) for male mice exposed to 1,3-butadiene for 40 weeks.
Exposure concentration, Testis weight, Testis weight/body weight, ppm mg, ratioa
0 116 ± 6 2.89 ± 0.19 6.25 117 ± 9 2.93 ± 0.28 20 114 9 2.78 0.25 62.5 103 ± 12* 2.87 ± 0.36 200 102 ± 6* 2.54 ± 0.18* 625 60 ± 10* 1.60 + 0.07* 

### aResults are given as means ± SD; n = 10. *Different compared to control, p < 0.05. 10 20 30 40 50 60 70
WEEKS ON STUDY
FIGURE 2. Survival curves for male B6C3F1 mice exposed to 1,3- butadiene: stop-exposure groups. Exposure to 1,3-butadiene was stopped after 13 (SE 13 wk), 26 (SE 26 wk), 40 (SE 40 wk), or 52 weeks (SE 52 wk). After these limited exposure periods, animals

were held in control chambers.

duced levels of circulating erythrocytes. Another change caused by exposure ofmale and female mice to 1,3-butadiene was an increase in mean corpuscular volume and in mean corpuscular hemoglobin. A similar profile ofhematologic changes was observed inmale and female mice exposed to 625 ppm 1,3-butadiene for 65 weeks. 

### 40-Week Evaluation
Gross andmicroscopic evaluations ofearly-deathmice at week 40 (i.e., those which died between the start of the study andweek 40), indicated that the predominant

cause ofdeath in the 625-ppm exposure group was lymphocytic lymphoma (Table 3). The cumulativenumberof deaths by week 40 at the lower exposure levels was too

Table 2. Hematologic changes in peripheral blood of male mice exposed to 1,3-butadiene by inhalation for 40 weeks.a
Exposure concentration, RBC HGB, VPRC, MCV, MCH, ppm x 106/p.L g/dL mL/dL fL pg 0 10.4 ± 0.3 16.5 ± 0.4 48.1 ± 1.5 46.3 ± 0.8 15.9 ± 0.2 6.25 10.3 ± 0.3 16.4 ± 0.5 47.8 ± 1.7 46.4 ± 1.0 16.0 ± 0.3 20 10.4 ± 0.4 16.7 ± 0.7 48.2 ± 2.2 46.3 ± 0.8 16.1 ± 0.4 62.5 9.9 ± 0.4* 15.9 ± 0.6* 45.9 ± 2.1* 46.7 ± 1.2 16.1 ± 0.3 200 9.6 ± 0.5* 15.6 ± 0.9* 45.4 + 2.7* 47.2 ± 1.0 16.2 ± 0.2* 625 7.6 ± 1.2* 13.5 ± 1.8* 39.9 5.3* 53.2 ± 2.9* 17.9 ± 0.8*

an= 10. *Different from control 

### (0 ppm), p < 0.05. -J

U, Ui. 0

-i

0. 30INHALATION TOXICOLOGY OF 1,3-BUTADIENE IN MICE

Table 3. Incidence of microscopic lesions in early death mice (through week-40) exposed to 625 ppm 1,3-butadiene.

SE SE Malesa Femalesa week 13b week 26 Target: lesion n = 49 n = 21 n = 10 n = 20 Lymphocytic lymphoma 43 16 9 19 Heart: Endothelial hyperplasia 2 0 0 0 Forestomach: Epithelial hyperplasia 22 11 3 9 Squamous cell neoplasm 1 2 0 1 Lung: Alveolar epithelial hyperplasia 3 1 0 0 Alveolar-bronchiolar neoplasm 1 2 0 1 Gonad: Atrophy 35 15 2 4
aNumberofearlydeathanimals: males, n = 49; females, n = 21. There were
no deaths in either the male or female control groups. bSE, stop-exposure groups (males).

few (3/group or

### less) todemonstrate any

### potential trend. However, 

### these deaths were 

### generally due to 

### lymphocytic 

### lymphoma and included threemice at 200 ppmand

onemouse at 62.5ppm. 

### Malemiceweremore 

### susceptible to 

### 1,3-butadiene-induced lymphoma than 

### females; by week 40, 

### lymphocytic lymphoma was observed in 43 

### of 49 

### early-death males and 16 

### of 21 

### early-death females 

### exposed to 625 ppm 

### 1,3-butadiene. Lymphocytic 

### lymphomas occurred in most early

### death mice in 

### the stop-exposure 

### (SE) groups 

### (Table 3). Ten of 50 male mice 

### exposed to 625 ppm 

### 1,3-butadiene for 13 

### weeks and 20 

### of 50 male mice 

### exposed to 625 ppm 

### 1,3-butadiene for 26 weeks died 

### by week 40. Twenty

### eight of these deaths were 

### due to 

### lymphocytic lymphoma. The 

### mortalityrate 

### and the incidence of lympho

### cytic 

### lymphoma was 

### markedly reduced in those groups 

### exposed to 

### lower concentrations of 

### 1,3-butadiene for a 

### longer duration 

### (comparable total exposures); lympho

### cytic 

### lymphoma was present in 

### only two male mice 

### exposed to 200 ppm 

### 1,3-butadiene for 40 weeks and one 

### male mouse 

### exposed to 312 ppm. 

### The incidence 

### of other tumor types, typically later 

### appearing than 

### lymphocytic lymphoma, were too few to 

### clearlydemonstrate an

### exposure-related effect. The low incidences of squamous cell neoplasms of the fore

### stomach and 

### alveolar-bronchiolar 

### neoplasms observed 

### by week 40 are 

### noteworthy since 

### they are uncommon, 

### especially in 46 to 

### 47-week-old 

### B63CF1, and 

### because 

### they had been observed at increased incidences in the 

### previous inhalation 

### study in mice 

### (5,11). 

### Epithelial hy

### perplasiaof

### the forestomachwas observedinover50% of themice

### dyingbyweek40 thatwere 

### exposedto 625ppm 

### 1,3-butadiene. Testicular and ovarian 

### atrophy, lesions observedat

### highrates inthe

### previous study (5,11), were 

### prominent in 

### this study as 

### well. The 

### incidence of selected 

### histopathologic lesions in groups 

### of 10

### male or 10 

### femalemicekilledat

### week40 are 

### shown in 

### Table 4. 

### Lymphocytic 

### lymphomas were 

### observed in two

### males and one 

### female 

### exposed to 625 ppm 1,3-butadiene. The low incidence of lymphocytic lymphoma in these animals, compared to the early-death mice 

### (Table 3), 

### reflects the 

### rapid onset of a moribund condition in 

### lymphoma-bearing mice. 

### Proliferative le

### sions of the 

### forestomach 

### (epithelial hyperplasia and squamous 

### cell

### papilloma) werepresent in

### bothmales and females 

### exposed to 625 ppm 

### 1,3-butadiene. Epithelial 

### hyperplasia of the 

### forestomach was 

### observed in one female mouse 

### exposed to 200 ppm 

### 1,3-butadiene. The incidences of alveolar 

### epithelial hyperplasia and alve

### olar-bronchiolar neoplasms were increased in 

### male and 

### female mice 

### exposed to 625 

### and 200 ppm 

### 1,3-butadiene, butnot to 62.5ppm

### 1,3-butadiene. Testicular

### atrophy, of 

### minimal to mild severity, was present in 6 of 10 males 

### exposed to 625 ppm 

### 1,3-butadiene. Ovarianatrophywas marked in

### femalemice 

### exposedto 200 or625ppm 1,3-bu

### tadiene, while ovaries of mice 

### exposed to 62.5 ppm 

### appeared 

### normal. The 

### atrophic ovaries had no identifia

### ble oocytes, 

### follicles, or corpora lutea. A 

### granulosa cell neoplasmofthe ovarywas observedinone femalemouse in the 200-ppm dose group. 

### Hemangiosarcomas of the heart were not observed in animals sacrificed at the 40-week interim evaluation; 

### however, endothelial hy

### perplasia, a 

### proliferative vascular lesion of the 

### heart,

was 

### observed in one

### male and one 

### female 

### exposed to 625 ppm 

### 1,3-butadiene for 40 

### weeks and two 

### early death male mice 

### exposed to 625 ppm 

### 1,3-butadiene.



### 65-Week 

### Evaluation

Data 

### from the 65-week interim sacrifice ofmale mice 

### (Table 5) show that exposure to 1,3-butadiene resulted in 

### dose-related increases in the incidences of prolifera

### tive lesions ofthe

### heart, forestomach, lung, andHarderian 

### gland (a 

### site not identified in the 

### previous study). 

### Only 7 

### ofthe 90

### malemice 

### exposedto 625ppm

### butadiene survived until week 65. 

### Proliferative responses were 

### induced inmalemice at concentrations as low as 62.5 or 20 ppm 

### after 65 

### weeks of exposure. Testicular atrophy

was 

### observed 

### only in 

### the 625-ppm exposure group. The interim sacrifice 

### offemalemice atweek 65 

### (Table 

### 6) also revealed 

### 1,3-butadiene-induced proliferative

Table 4. Incidence of microscopic lesions in mice exposed to

1,3-butadiene for 40 weeks.a

Exposure concentration, ppm Males Females Target: lesion 0 200 625 0 200 625 Lymphocytic lymphoma 0 0 2 0 0 1 Forestomach: Epithelial hyperplasia 0 0 6 0 1 6 Squamous cell papilloma 0 0 3 0 0 2 Lung. Alveolar epithelial hyperplasia 0 3 3 0 0 1 Alveolar-bronchiolar neoplasm 1 2 4 0 3 1 Gonad: Atrophy 0 0 6 0 9 8

an = 10, exceptforfemalemiceexposedto625ppm1,3-butadienewheren = 8. 31MELNICK ETAL.

Table 5. Incidence of microscopic lesions in male mice exposed

to 1,3-butadiene for 65 weeks.a

Exposure concentration, ppm Target: lesion 0 6.25 20 62.5 200 625 Lymphocytic lymphoma 0 0 0 0 0 2 Heart: Endothelial hyperplasia 0 NEb 0 1 1 1 Hemangiosarcoma 0 NE 0 0 1 3 Forestomach: Epithelial hyperplasia 0 0 0 1 0 2 Squamous cell neoplasm 0 0 0 1 3 4 Lung: Alveolar epithelial hyperplasia 0 0 1 1 3 2 Alveolar-bronchiolar neoplasm 0 0 0 2 4 5 Harderian gland: Adenoma 0 0 2 4 3 3 Liver: Hepatocellular neoplasm 3 1 5 3 4 6 Testis: Atrophy 0 NE NE NE 0 4

an = 10, except for mice exposed to 625 ppm 1,3-butadiene where

n = 7. bNE, not examined microscopically.

Table 6. Incidence of microscopic lesions in female mice exposed to 1,3-butadiene for 65 weeks.a

Exposure concentration, ppm Target: lesion 0 6.25 20 62.5 200 625 Lymphocytic lymphoma 1 0 0 0 3 0 Heart: Endothelial hyperplasia 0 NEb NE 0 4 0 Hemangiosarcoma 0 NE NE 0 1 2 Forestomach: Epithelial hyperplasia 0 0 0 0 2 1 Squamous cell neoplasm 0 0 0 1 2 1 Lung: Alveolar epithelial hyperplasia 0 0 2 1 4 0 Alveolar-bronchiolar neoplasm 0 0 0 3 4 2 Liver: Hepatocellular neoplasm 1 1 1 1 3 1 Ovary: Atrophy 0 0 2 9 8 2

an = 10, except for mice exposed to 625 ppm

n = 2. bNE, not examined microscopically. 1,3-butadiene where

changes in the heart, forestomach, and lung, especially at 200 and 62.5 ppm 1,3-butadiene. Onlytwo femalemice exposed to 625 ppm 1,3-butadiene survived until week 65. Ovarian atrophy was observed at exposure levels of 20 ppm and higher. The cumulative incidence of microscopic lesions in early-death mice through week 65 shows that lymphocytic lymphoma was the major cause ofdeath at the 625

ppm exposure level, especially for male mice (Table 7). Betweenweek 40 andweek 65, 24 male mice exposed to 625 ppm 1,3-butadiene died; 18 had lymphocytic lymphoma. Increased incidences of hemangiosarcomas of the heart, forestomach neoplasms, alveolar-bronchiolar neoplasms, Harderianglandadenomas, andhepatocellularneoplasms werealsoevident inearly-deathmalemice exposed to 625 ppm 1,3-butadiene. The number ofearly deaths in male mice exposed to 62.5 or 200 ppm 1,3- Table 7. Incidence of microscopic lesions in early-death mice (through week 65) exposed to 1,3-butadiene.

Exposure concentration, ppm Males Females 62.5 200 625 62.5 200 625 Target: lesion (4)a (7) (73) (6) (19) (80) Lymphocytic lymphoma 1 2 61 1 6 36 Heart: Endothelial hyperplasia 0 0 6 1 4 9 Hemangiosarcoma 0 0 3 0 3 24 Forestomach: Epithelial hyperplasia 0 1 39 3 5 41 Squamous cell neoplasm 0 0 6 0 0 26 Lung: Alveolar epithelial hyperplasia 0 0 9 1 4 10 Avelolar-bronchiolar neoplasm 1 3 3 1 5 22 Harderian gland: Adenoma 0 1 4 0 5 6 Mammary gland: Adenocarcinoma 1 3 11 Ovary: Granulosa cell neoplasm 0 2 5 Liver: Hepatocellular neoplasm 1 1 5 0 4 2 Gonad: Atrophy 0 1 53 4 15 68 

### aNumbers inparentheses equal the number of early death animals in each group. Only one control male mouse and one control female

mouse died during the first 65 weeks of the study; hepatocellular adenoma was observed in the male mouse.



### butadiene was too low to 

### ascertain any

### potential carcin

### ogenic

### effects. 

### Fifty-nine femalemice thatwere 

### exposed to 625 ppm 

### died between week 40 

### and week 65; 20 had 

### lymphocytic lymphoma. The 

### incidence of other

### primary tumors was 

### highinthese 59 mice: 24

### hemangiosarcomas of the 

### heart, 24 squamous 

### cell 

### neoplasms of the fore

### stomach, 20 

### alveolar-bronchiolar 

### neoplasms, 6 Harder

### ian 

### gland adenomas, 11 mammary 

### gland adenocarcinomas, 

### and 5 

### granulosa 

### cell neoplasms of the ovary. Elevated 

### incidences of neoplasms at 

### these sites were 

### also apparent in

### early-death femalemice 

### exposed to 200 ppm 

### 1,3-butadiene. The 

### development of 

### neoplasms in 

### the 

### heart, forestomach, and 

### lung were 

### supported by high incidences of

### hyperplasias in 

### these organs in both 

### males and

### females. Testicular

### atrophywas 

### observedat a 

### high incidence in 

### male mice 

### exposed to 625 ppm, while ovarian

### atrophywas 

### observed infemalemice 

### exposed to 

### 62.5, 200, or 625 ppm 

### 1,3-butadiene. The tumor 

### incidence 

### profiles in the stop-exposure groups 

### (Table 8) indicate that at 

### comparable total exposures, 

### the occurrence of

### lymphocytic 

### lymphoma is more 

### likely with exposure to a 

### higher concentration 

### of 

### 1,3- butadiene for a short 

### time, 

### compared to exposure to a lower 

### concentration for an 

### extended duration. This is 

### evident 

### by

### comparingthe incidence of

### lymphocytic 

### lymphoma in the 625 ppm 

### SE 13 WKgroup 

### (15 

### lymphocytic 

### lymphomas) withthat inthe 200ppm

### SE 40

### WKgroup 

### (3 lymphocytic lymphomas), and 

### comparing the incidence in

### the 625ppm

### SE 26

### WKgroup 

### (30

### lymphocytic 

### lymphomas) with 

### that in 

### the 312 ppm 

### SE 52 

### WK group 

### (3 lymphocytic 

### lymphomas). An 

### obscuring effect of a 

### high 32INHALATION TOXICOLOGY OF 1,3-BUTADIENE INMICE

incidence 

### of 

### lymphocytic 

### lymphoma on 

### 1,3-butadieneinduced neoplasia at other 

### primary sites is evident by comparing 

### the tumor 

### profile in the 312 ppm 

### SE 52 WK group with that in the 625 ppm 

### SE 26 WK group. The incidences 

### of

### hemangiosarcomas of the 

### heart, alveolar

### bronchiolar 

### neoplasms, and 

### hepatocellular neoplasms

were 

### higher in the group with the lower incidence of 

### lymphocytic 

### lymphoma (312 ppm 

### SE 52 

### WK), 

### indicating that 

### longer survival allows these other 

### chemically in

### duced tumors to manifest. 

### Graphic presentation of the incidences oflymphocytic 

### lymphomas and

### hemangiosarcomas 

### ofthe heart versus 

### weeks on 

### study for these two stop-exposure groups 

### (Fig. 3) shows that 

### hemangiosarcomas 

### ofthe heart are more 

### likely to 

### be observed when 

### the incidence of 

### early 

### lymphocytic lymphomas is 

### low; conversely, when the rate of 

### early lymphocytic lymphoma is 

### high, the incidence 

### of 

### 1,3-butadiene-induced hemangiosarcoma of 

### the 

### heart is low.


## Discussion 1,3-Butadiene has 

### been shown to induce multiple organ 

### carcinogenicity in rats and mice (5,11,14). In 

### B6C3F1 mice, exposure to 625 or 1250 ppm 1,3- 

### butadiene caused 

### early induction and 

### significantly in

### creased incidences of 

### malignant lymphomas, hem

### angiosarcomas of the 

### heart, alveolar-bronchiolar neoplasms, squamous 

### cell 

### neoplasms of the 

### forestomach, acinarcell carcinomas 

### ofthemammary

### gland, granulosa cell 

### neoplasms of the ovary, and 

### hepatocellular neoplasms (5,11); 

### Sprague-Dawley rats 

### exposed to 1000 or 8000 ppm 

### 1,3-butadiene had 

### increased incidences at 

### multiple sites: 

### pancreatic exocrine 

### adenomas, uterine
sarcomas, 

### Zymbal gland 

### carcinomas, mammary 

### gland neoplasms, 

### thyroid follicular

### cell

### neoplasms, and 

### Leydig cell 

### neoplasms ofthe testis 

### (14). 

### Morphological descripTable 8. Incidence ofmicroscopic lesions in early-death male mice: stop-exposure (SE) groups through week 65.

Exposure concentration, ppma 200 625 312 625 SE 40 SE 13 SE 52 SE 26 Target: lesion (8000)a (8125) (16224) (16250) Lymphocytic lymphoma 3 15 3 30 Heart: Endothelial hyperplasia 0 1 1 5 Hemangiosarcoma 1 0 12 4 Forestomach: Epithelialhyperplasia 2 5 12 13 Squamous cell neoplasm 0 3 3 9 Lung: Alveolareipthelialhyperplasia 1 0 3 4 Alveolar-bronchiolarneoplasm 1 3 9 5 Harderian gland: Adenoma 1 1 3 2 Liver: Hepatocellularneoplasm 2 0 8 4 Testis: Atrophy 0 2 1 5 aNumberofearly-death

### aninals ineachgroup (n): 

### 200ppm, SE40, n = 7; 625 ppm, SE 13, n = 20; 312 ppm, SE 52, n = 21; 625 ppm, SE 26, n = 36. nTotal exposure expressed asppm-weeks. 

### tions and illustrations 

### of the neoplastic 

### lesions induced by 

### 1,3-butadiene in B6C3F1 mice are presented in the 

### followingpaper (15). The 

### largenumberofprimaryorgan sites of tumor induction 

### by 1,3-butadiene was unusual 

### for chemicals studiedbythe NTP; only 1.9% ofapproximately 360 chemicals evaluated showed evidence ofcar

### cinogenicity at six or more 

### primary sites. 

### Although the previous mouse study was ended after 

### only 60 weeks 

### ofexposure because of

### highrates ofearly 

### mortality (5,11), the 

### doses used in that study are not considered excessive for 

### evaluation of carcinogenicity, 

### since survival was not 

### reduced from toxic 

### effects other 

### than from the induction of 

### fetal neoplasms. Furthermore, 

### those concentrations (625 and 1250 ppm) are 

### equivalent toand

### belowthe 

### OSHAstandardof1000ppm 

### for occupational exposure to 1,3-butadiene (16). Announcements have beenmade to lower this level consid

### erably (17). The results ofthe studies of 1,3-butadiene inmice are unusual inthatuncommontumors, suchas

### hemangiosarcoma of the 

### heart and squamous cell 

### neoplasms of the 

### forestomach, were 

### inducedafter

### inhalationexposure. In untreated 

### B6C3F1 mice 110 to 112 weeks of age, the 

### historical rate 

### for 

### hemangiosarcomas of the heart is 

### about 0.04% (1 in 2500), while that of squamous 

### cell neoplasms of the forestomach is about 0.5% (1 in 200) 

### (12). In 

### the present 

### studies, Harderian gland adenomas

were 

### observed in 

### early-death mice 

### (through week 65) 

### exposed to 200 or 625 ppm 

### 1,3-butadiene and at 

### the 65-weekinterim

### evaluation. Theseobservations indicate

an 

### additional site of tumor 

### induction by 

### 1,3-butadiene;

_ o Lymphoma, 625ppmSE 26wk

* Lymphoma, 312 ppm SE52wk I o Hemongiosorcoma, 625 ppm SE 26wk 60 - a Hemangiosarcoma, 312 ppm SE 52 wk

0

C,) -Ja

### a. 0

w llJ z

L-) z w a

z 50

40

30

20

10 .10
0

30 40 70

WEEKS ON STUDY

FIGURE 3. Cumulative incidence oflymphocytic lymphomas orhemangiosarcomas of the heart versus weeks on study in two stopexposure groups for which the total exposures to 1,3-butadiene

were comparable (625 ppm SE 26 wk and 312 ppm SE 52 wk). 33MELNICK ETAL.

the 

### historical rate 

### of Harderian 

### gland neoplasms in un

### treated 110- to 

### 112-week-old 

### B6C3F1 mice is 

### approximately 2% (12). The present 

### inhalation studies of 

### 1,3-butadiene were 

### performed to 

### better characterize 

### dose-response rela

### tionships for butadiene-induced 

### neoplastic and nonneoplastic lesions in 

### B6C3F1 mice at various intervals and 

### after 2 years 

### ofexposure. 

### The 

### following conclusions are 

### based onthe results ofthe 

### 40-weekand 65-weekevaluations. 

### 1,3-Butadiene is a 

### gonadal toxicant in 

### mice, in

### ducing testicular 

### atrophy in males at 625 ppm, and 

### ovarian 

### atrophy in 

### females at 20 ppm and 

### higher. This gas was 

### also toxic to 

### thebonemarrow

### of

### mice, causinga 

### partially or

### poorlyregenerative 

### dose-dependent anemia at exposure 

### concentrations of 62.5 ppm

### and

### higher. The increase in 

### MCV and 

### MCH at 625 ppm 

### 1,3-butadiene indicates that 

### although 1,3-butadiene caused suppression 

### of

### hematopoiesis in 

### the bone marrow, young 

### larger cells may

### have been 

### released into the 

### blood from extra

### medullary sites. It has 

### been 

### reported that exposure of 

### B6C3F1 mice to 1250 ppm 

### 1,3-butadiene produces a 

### macrocytic-megaloblastic anemia 

### (18) and promotes ex

### tramedullary hematopoiesis in the 

### spleen (19). For 

### male or 

### female mice 

### exposed to 625 ppm 1,3- 

### butadiene, lymphocytic 

### lymphoma was the 

### major cause 

### of

### death in

### the first 50 

### weeks ofthe 

### study. Priorto

### week 65, 

### the lowest exposure 

### concentration at

### which 

### lymphocytic 

### lymphomas were 

### observed was 62.5 ppm; 

### lymphocytic 

### lymphoma was observed in one 

### male mouse ex

### posed to this 

### concentration of 

### 1,3-butadiene for 29 weeks. 

### Lymphocytic lymphomas were 

### also induced in 

### male mice 

### exposed to 625 ppm 

### 1,3-butadiene for 

### only 13 

### weeks 

### (i.e., stop-exposure 

### study). The 

### incidence of

### lymphocytic 

### lymphoma was two 

### times 

### higher in male mice 

### exposed to 625 ppm 

### 1,3-butadiene for 26 

### weeks, com

### pared to male mice 

### exposed to 625 ppm for 13 weeks. When 

### the exposure concentration was reduced to 312 ppm 

### and the exposure 

### duration was 

### extended to 52 

### weeks (312 ppm 

### SE 52 

### WK), the 

### incidence of 

### lymphocytic 

### lymphoma was 10% 

### ofthe incidence inthe group 

### of mice 

### that received the same total exposure to 1,3- 

### butadiene 

### (625 ppm 

### SE 26 

### WK). 

### Thus, the 

### multiple of exposure 

### concentration times exposure 

### duration does not 

### predict the incidence of 

### lymphocytic lymphoma in 

### mice. At 

### equivalent total exposures, 

### the likelihood for induction of 

### lymphocytic 

### lymphoma is greater with exposure to a 

### higher concentration of 1.3-butadiene for a 

### shorter time than for exposure to a 

### lower concentration of 

### 1,3-butadiene over an extended duration. 

### The 

### earlymortalities due to 

### lymphocytic lymphomas, 

### especially in 

### male mice 

### exposed to 625 ppm 

### 1,3- 

### butadiene, appear to 

### limit the 

### expression of

### neoplasms at other 

### sites. Increased 

### incidences of 

### hemangiosarcomas 

### of the 

### heart, squamous 

### cell 

### neoplasms of the 

### forestomach, alveolar-bronchiolar 

### neoplasms, Harderian 

### gland 

### adenomas, 

### adenocarcinomas of the mammary 

### gland, 

### granulosa cell 

### neoplasms ofthe ovary, 

### and

### hepatocellular 

### neoplasms were 

### frequently observed in 

### 1,3-butadiene-exposedmice that

### diedbetweenweeks 45 and 65 

### ofthe 

### study. Since earlylymphocytic lymphomas

were not as 

### prevalent at 

### concentrations of 6.25 to 200 ppm, it is likely that a 

### clearer dose response for 1,3-butadiene-induced 

### neoplasia at these sites will be obtained over this concentration range. The lack of a clear 

### dose response 

### for some of 

### the 

### neoplastic and nonneoplastic lesions observed in the previous inhalation 

### study of 

### 1,3-butadiene in 

### B6C3F1 mice (5,11) was likely due to 

### reduced survival caused

### byearlydevelopment of 

### malignant lymphomas. Proliferative, nonneoplastic le

### sions of the 

### forestomach, 

### heart, and 

### lung which were 

### observed in 

### early-death mice 

### and at 

### the 40-week and 65-week 

### evaluations may represent treatment related 

### preneoplastic changes at these target sites. The carcinogenicity studies of 1,3-butadiene in rats and in mice demonstrate a species difference with respect to 

### sites of tumor 

### induction and magnitude of response. Irons

### andcoworkers (20,21) have suggested

### that activation 

### of an 

### endogenous ecotropic retrovirus may play a critical role in butadiene-induced thymic 

### lymphomalleukemiain

### B6C3F1mice. 

### However, thefinding that exposure to 1250 ppm 1,3-butadiene for up to 1 year also caused a 14% incidence 

### of 

### thymic lymphoma/ leukemia in NIH 

### Swiss mice, a strain that does not express the 

### endogenous 

### ecotropic retrovirus recovered 

### fromtissues of

### B6C3F1 mice. 

### Further, Swissmice

### have

a 

### background rate 

### of zero 

### for this 

### neoplasm, indicating that 

### 1,3-butadiene can induce 

### thymic 

### lymphoma inde

### pendently of this activated 

### retrovirus. Butadiene monoxide (1,2-epoxybutene-3) has been 

### identified as 

### the primary 

### metabolite of 1,3-butadiene 

### biotransformation 

### by rat liver 

### microsomal monooxygenase 

### (22). This intermediate may be 

### further metabolized 

### by liver monooxygenase or 

### epoxide hydrolase to 

### diepoxybutane or 

### 3,4-epoxy-1,2-butanediol, respec

### tively (23). Butadiene monoxide and 

### diepoxybutane 

### have been shown to be 

### mutagenic to 

### Salmonella typhimurium strains, which 

### are sensitive to base 

### pair substitution mutagens 

### (24,25), and to 

### induce local neo

### plasmswhen

### appliedto

### the skinofmiceorwhenadministered to rats or mice 

### by subcutaneous 

### injection (26,27). 

### Since 

### 1,3-butadiene is 

### mutagenic to 

### Salmonella 

### ty

### phimurium only when incubated with 

### liver S-9 activation systems 

### (28), epoxide intermediates may 

### be the 

### ultimate 

### carcinogenic forms ofthis 

### chemical. Butadiene monoxide has been 

### identified in 

### the expired air 

### of 

### Sprague-Dawley rats 

### exposed to 

### 1,3-butadiene (29). 

### Although the rate 

### ofbutadiene metabolismis greater in 

### B6C3F1 mice than 

### Sprague-Dawley rats at 

### equivalent exposure 

### concentrations 

### (30-32), the 

### magnitude of 

### this difference at 

### carcinogenic exposure 

### levels in these species does not seem to be sufficient to account 

### for the 

### species difference in 

### carcinogenicity of 

### 1,3-butadiene. The present studies 

### indicate that 200 ppm butadiene is 

### carcinogenic to 

### the 

### heart, lung, Harderian 

### gland, mammary

### gland, and ovary of

### B6C3F1 mice; however, other than for the mammary 

### gland, neoplasms were not in

### duced at 

### these sites in 

### Sprague-Dawley rats 

### exposed to 1000 or 8000 ppm 

### 1,3-butadiene. The metabolism of 34INHALATION TOXICOLOGY OF 1,3-BUTADIENE IN MICE 35

1,3-butadiene is greater in 

### Sprague-Dawley rats ex

### posed to 1000 ppm 

### 1,3-butadiene than in 

### B6C3F1 mice 

### exposed to 200 ppm 

### 1,3-butadiene (30,31). 

### Perhaps steady-state tissue concentrations 

### of these intermediates at target sites are 

### sufficiently different to account 

### for the 

### species difference in 

### carcinogenicity of 

### 1,3-butadiene. The inhalation 

### carcinogenicity studies of 1,3-butadiene in rats and mice have raised concerns of risk for 

### humans exposed to this 

### chemical. Results of epidemi

### ology studies indicate an association 

### between occu

### pational exposure to 

### 1,3-butadiene andthe

### development of 

### lymphatic and 

### hematopoietic neoplasms (33-36). These findings are consistent with the 

### results of the 

### inhalation studies of 

### 1,3-butadiene in mice.

NOTE ADDED IN PROOF: Data from the 2-year exposure studies 

### clearly showed that the incidence of 

### alveolar-bronchiolar 

### neoplasms in 

### female 

### B6C3F1, mice

was 

### increasedatall exposure

### concentrations ofof1,3-bu

### tadiene, including 6.25ppm, 

### compared to controls 

### (R.L. Melnick et 

### al., Cancer 

### Res., in 

### press).

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