Environmental Health Perspectives

Vol. 57, pp. 199-206, 1984

Further Studies on the Toxicology

of the Glycol Ethers with Emphasis on

Rapid Screening and Hazard Assessment

by J. E. Doe*

The discovery that ethylene glycol monomethyl ether (EGME) could affect the testis and the developing

fetus in laboratory animals prompted further work to understand the effect of EGME and to examine

additional glycol ethers to see if they showed EGME's reprotoxicity.

Propylene glycol monomethyl ether (PGME) was shown not to cause testicular atrophy or to affect the

development of rats at 600 ppm by inhalation, whereas EGME caused testicular atrophy at 300 ppm and

showed teratogenic potential at 100 ppm.

Diethylene glycol monomethyl ether (diEGME) was found to show no teratogenic potential when

administered subcutaneously in rats at up to 1000 ,iL/kg, whereas EGME had effects at 40 ,iL/kg.

EGME has been shown to cause effects on the testis in rats after a single exposure to 600 ppm or above for

4 hr. The effects can be seen as little as 24 hr after exposure. Ethylene glycol monoethyl ether (EGEE) also

causes a reduction in testicular weight following a single exposure to saturated vapor for 3 hr (17 mg/I), but

ethylene glycol monoisopropyl ether (EGPE) at 15 mg/l and ethylene glycol monobutyl ether (EGBE) at 4

mg/l showed no effect on the testis.

Introduction

The reproductive toxicology of glycol ethers has been

the subject of much concern in recent years. This

concern was initiated by a report (1) describing testicular atrophy in mice dosed with ethyelene glycol monomethyl ether (EGME) and ethylene glycol monoethyl

ether (EGEE). Moreover, both EGME (2) and EGEE

(3) have been shown to cause fetotoxicity and/or teratogenicity in laboratory animals. These observations lead

to a general reappraisal of the glycol ethers as a class

and this paper describes some of the work which was

carried out within Imperial Chemical Industries PLC in

order to determine the reproductive toxicology of

several members of the glycol ethers group which were

of proven or potential commercial importance. None of

the studies was conducted to full classical protocols, but

each one was designed to answer a specific question to

aid in either product development or in the development

of safe handling. In retrospect, this series of experiments can be seen as a way in which toxicology can

provide useful data even though the studies are not

carried out to a regulatory protocol.

*Imperial Chemical Industries PLC, Central Toxicology Laboratory,

Alderley Park, Macclesfield, Cheshire, UK.

Comparative Reproductive Toxicology

by Inhalation in Rats of Ethylene

Glycol Monomethyl (EGME) and

Propylene Glycol Monomethyl Ether

(PGME)

This study was carried out at a time when the effects

of EGME upon both the testes by inhalation (4) and its

effects of the fetus (2) were known. However, there were

no data available on the toxicology of PGME, and this

study was designed to assess whether PGME was

capable of causing the same effects on reproductive

toxicology as EGME. It was therefore important to use

EGME as a positive control for both the testicular

effects and the teratogenic effects. The study was

designed in two parts: effect on testis and hematology

and determination of teratogenic potential.

Effect on Testis and Hematology

Ten male rats of the Alpk/Ap (Wistar-derived) strain

were exposed to the following concentrations: 100 ppm

EGME, 300 ppm EGME, or 200 ppm PGME or 600 ppm

PGME for 6 hr/day for 10 consecutive days in 3.4 m3

inhalation chambers (5). During the exposure period
J. E. DOE

the animals were periodically weighed and observed for

changes in their clinical condition or other behavioral

changes. On the day after the 10th exposure the animals

were killed by exposure to Fluothane. Blood samples

were taken for hematological assessment and the testis

weighed and removed and processed, along with the

thymus, into slides for subsequent histopathological

evaluation.

The body weight gain and absolute weights of the rats

exposed to 300 ppm EGME were significantly reduced;

however, 100 ppm EGME and 200 or 600 ppm PGME

had no effect on body weight. None of the treatments

caused any changes in clinical observations. At postmortem there were no effects at 100 ppm EGME or at 200

and 600 ppm PGME, however, the thymus and the

testes of the rats exposed to 300 ppm EGME were

reduced in size (Fig. 1). In the blood samples taken at

postmortem 300 ppm of EGME was shown to cause a

reduction in total white blood cells, the number of red

blood cells and the hemaglobin concentration of the red

blood cells. There were no effects at 100 ppm EGME or

with either concentration of PGME. Histological examination of the testis revealed a few seminiferous tubules

with disordered spermatogenesis with a similar incidence in the controls and in rats exposed to 100 ppm

EGME, 200 ppm PGME or 600 PGME. Pronounced

tubular atrophy was seen in the testis from all the rats

exposed to 300 ppm EGME, where 70 to 100% of the

seminiferous tubules were affected.

Determination of Teratogenic Potential

In order to provide a rapid assessment of the effect of

PGME and EGME on the developing embryo when rats

were exposed by inhalation, an adaptation of the

procedure described by Chernoff and Kavlock (6) was

employed. Pregnant female rats, approximately 11 to 13

weeks of age, of the Alpk/Ap (Wistar-derived) strain

were used. Groups of 20 rats per group were exposed to

air, 100 or 300 ppm EGME and 200 or 600 ppm of PGME

for 6 hr/day on days 6 to 17 (inclusive of gestation, day of

mating being day 0). After the last exposure period the

rats were removed from the exposure chamber and

housed singly. The rats were allowed to deliver their

litters which were observed for 3 days. The number of

live and dead pups in each litter and the weight of each

litter were determined on days 1 and 3 postpartum. Any

rats not producing litters by day 24 of gestation were

killed, and a gross postmortem examination was carried

out to determine the status of pregnancy. All the rats

were pregnant: all the control rats and all the rats

exposed to PGME produced litters. Exposure to 300

ppm EGME caused a significant reduction in body

weight gain throughout the study, and none of these

rats produced litters. Only 9 of the 20 rats exposed to

100 ppm produced litters and in these rats the gestation

period was significantly increased to a mean value of

23.6 days from a control value of 22 days. The comparative litter data are shown in Thble 1. There was a

significant reduction in the proportion of live pups

initially and the proportion surviving for 3 days postpartum in the litters of dams exposed to 100 ppm EGME.

The weight of the pups in this group on day 1 was

similar to the controls, although they had gained

significantly less weight by day 3. All the pups were

normal externally. There was no evidence of any effect

in either of the groups exposed to PGME.

L

.2

,uV

L R L K 1. R K

Conlrol l(X)ppm F(iME C M[ ( )ppm (1M 2o(1)ppill l(iMI (o)(pIp IMA,\I

FIGURE 1. Effect on testis weights of the inhalation of EGME and PGME for 10 consecutive days in male rats. Asterisks denote significantly

different from controls: (**) p < 0.01; (*) p < 0.05.

200
SCREENING AND HAZARD ASSESSMENT

Table 1. Comparative effect of EGME and PGME by inhalation on pregnant rats: litter data.

Group mean no. of Proportion of Proportion of Group mean Live pup

live and dead pups live pups pups surviving per litter, weight on

Treatment on day 1 (n) on day 1 to day 3 day 1 (n) day 3, g (n)

Control 11.5 ± 3.0 (20) 226/229 (98.7%) 223/226 (98.7%) 6.0 ± 0.5 (20) 7.2 ± 0.8 (20)

100 ppm EGME 3.3 ± 2.4 (9)t 23/30"t (76.7%) 16/23 (69.6%) 6.1 ± 0.6 (7) 6.3 ± 1.1 (4)

300 ppm EGME

200 ppm PGME 11.7 ± 2.0 (20) 231/233 (99.1%) 226/231 (97.8%) 6.0 ± 0.4 (20) 7.1 ± 0.7 (20)

600 ppm PGME 11.7 ± 2.6 (20) 228/233 (97.9%) 227/228 (99.6%) 6.0 ± 0.4 (20) 7.0 ± 0.7 (20)

tStatistically significantly different from the control group at the 1% level (Fisher's exact test, two-sided).

*Statistically significantly different from the control group at the 1% level (t-test, two-sided).

Conclusions

This experiment demonstrated that 300 ppm of

EGME caused hematological changes, testicular atrophy and thymic atrophy in male rats and complete

embryo lethality in pregnant female rats.A 100 ppm

level caused fetal mortality, fetotoxicity and the impairment of postnatal development. This spectrum of effects

has been defined as describing teratogenic potential (6).

In contrast, neither 200 nor 600 ppm of PGME caused

any effects in either males or pregnant females. The

results of this preliminary study therefore indicate

clearly that PGME does not share the spectrum of

activity shown by EGME upon the reproductive process

at significant dose levels. The study has been described

previously (7).

Teratogenic Potential of Diethylene

Glycol Monomethyl Ether (diEGME) in

the Postnatal Development Test in Rats

Few or no data existed at the time this study was

carried out on the effect of diethylene glycol ethers on

the developing embryo. This study was carried out to

assess the effect of diethylene glycol monomethyl ether

(diEGME) on the rat fetus in order to determine

whether diEGME showed the same spectrum of activity as EGME, which would have meant reconsideration

of the way in which the compound is handled. The

postnatal development test (6) was used, as EGME had

already been shown to be positive in this test by

inhalation. As diEGME is relatively nonvolatile, human

exposure is more likely to be by dermal contact;

consequently diEGME was administered by the subcutaneous route as this represents the worst case,

assuming the complete breakdown of the barrier function of the skin. Pregnant female Alpk/Ap (Wistarderived) rats 11 to 13 weeks of age were mated and the

day on which spermatozoa were detected was taken as

day 0 of pregnancy. Groups of 15 rats were used and

they were injected subcutaneously with 2 mL of sterile

water containing the appropriate compounds on days 6

to 20 of gestation inclusive. The groups were controls,

250 puL/kg diEGME, 500 ,LL/kg diEGME, 1000 ,LL/kg

diEGME, 40 ,uL/kg EGME, 250 p1L/kg EGME. The

rats were observed daily for changes in clinical condition and weighed on day 1 and on each day between day

6 and 20. Each rat was allowed to litter and the

offspring weighed and the number of dead and live pups

were recorded on days 1 and 4 postpartum. The female

rats and the pups were then killed. All animals which

failed to produce a litter by day 24 of pregnancy were

killed and the status of pregnancy determined.

No abnormalities in clinical condition were seen in

any of the rats treated with diEGME; however, some

of the rats treated with 250 p1L/kg EGME showed

piloerection and vaginal bleeding. The body weight gain

of these rats treated with 250 ,uL/kg EGME was

significantly reduced compared with controls. One control animal did not produce a litter and was found not to

be pregnant. None of the rats treated with 250 ,uL/kg

EGME produced litters, although they were all found to

have been pregnant.

The litter data are shown in Table 2, where it will be

seen that 40 puL/kg of EGME reduced the number of

litters as well as the number of live pups per litter and

the percentage of pups surviving for 4 days. EGME

therefore clearly showed its fetotoxic effect in this

study. diEGME, on the other hand, caused no maternal

toxicity and little or no effect on the development of the

young at 250 or 500 puL/kg. There was a slight, but not

statistically significant, decrease in the survivability in

the offspring of the rats treated with 1000 puL/kg, and

this may be indicative of a slight fetotoxic effect at this

high dose level. However, diEGME certainly does not

share the marked fetotoxicity shown by EGME, and, as

the route of administration was chosen to maximize the

possible effects of diEGME, it is unlikely that dermal

exposure to diEGME in humans would present a

significant risk. These data were reported previously

(8).

Effect on Route of Administration

on Teratogenic Potential of EGME

EGME has been shown to cause testicular atrophy

and/or fetotoxicity by a variety of routes such as

201
J. E. DOE

Table 2. Effect of subcutaneous diEGME and EGME in pregnant rats: litter data.

diEGME diEGME diEGME EGME EGME

Control 250 ,uL/kg 500 ,L/kg 1000 ,uL/kg 40 ,uL/kg 250 ,uL/kg

No. pregnant 14/15 15/15 15/15 15/15 15/15 15/15

No. of litters 14 15 15 15 7 0

No. of live pups per litter ( ± SD) 11.7 ± 2.7 12.0 ± 1.9 10.9 ± 3.1 10.5 ± 3.6 3.7 ± 4.2k

Pup weight at day 1, g ( ± SD) 5.9 ± 0.6 6.0 ± 0.5 6.3* ± 0.4 6.2 ± 0.5 6.1 ± -/5

Pup weight at day 5, g ( ± SD) 8.5 ± 1.1 8.8 ± 1.2 9.3 ± 1.1 8.6 ± 1.1 10.4 ± 0.7*

Weight gain per pup, g ( ± SD) 2.5 ± 0.8 2.8 ± 0.8 3.0 ± 1.0 2.4 ± 0.9 3.6 ± 0.2

No. live pups per litter day 5 ( ± SD) 11.2 ± 2.9 11.3 ± 1.9 10.6 ± 2.5 9.1 ± 3.3 2.1 ± 4.5t -

Surviving on day 5, % 95.2 91.4 89.9 85.6 46.9

No. of litters with deaths day 1-5 4/14 6/15 8/15 10/15 6/7

No. of litters with more than one death day 1-5 2/15 2/15 2/15 7/15 4/7

*Statistically significantly different from the control group means at 95% level Student t test.

'Statistically significantly different from the control group means at 99% level, Student t test, two-sided.

Table 3. Effect of route of administration on teratogenic potential of EGME.

Mean pup Mean pup

Number Number of Mean litter weight, Mean litter Survival, weight, % Weight

Treatment and route pregnant litters size, day 0 day 0, g size, day 4 % day 4, g gain (pups)

10 mL/kg saline IP 15 12 7.6 5.3 5.4 64 8.8 43

200 mg/kg EGME IP 15

10 mL/kg saline PO 15 15 10.7 6.1 10.7 99 9.0 48

200 mg/kg EGME PO 15 0

2 mL/kg saline SC 15 15 10.8 6.0 10.5 91 9.25 52

40 mg/kg EGME SC 15 7 4.6* 6.0 2.1* 23* 10.7 31

250 mg/kg EGME SC 15 0

2 mL/kg saline dermally 15 15 13.2 5.6 12.7 96 9.2 59

40 mg/kg EGME dermally 15 15 12.7 5.9 12.3 97 9.4 98

Control 20 20 11.5 6.0 11.5 99 7.0 17

100 ppm EGME, inhalation 20 9 3.3* 6.2 4.0* 49* 6.5 - 15*

300 ppm EGME, inhalation 20 0

*Statistically significantly different from control (p < 0.05).

inhalation (4) and ingestion (2). The work reported here

forms a comparison of the results of using EGME as

a positive control in several studies to determine the

teratogenic potential of a variety of compounds. EGME

has been given by the following routes during the course

of these studies: intraperitoneally, orally, subcutaneously, dermally and by inhalation. The results are shown

in Table 3. It will be seen that 200 to 250 mg/kg of

EGME caused complete loss of litters by the intraperitoneal route, by the oral route, and when given

subcutaneously; 300 ppm of EGME by inhalation, which

corresponds approximately to a dose of 350 mg/kg

assuming 100% retention, caused a similar result. At

lower doses, EGME causes a loss of approximately half

the litters, with reduced litter size and survivability in

the pups. This is shown by 40 mg/kg EGME subcutaneously and 100 ppm EGME by inhalation (which corresponds to an approximate dose of 115 mg/kg assuming

100% retention). Interestingly, a similar dose administered dermally, i.e., 40 mg/kg EGME, had no effect on

the developing fetus. This result indicates that the skin

has some barrier function with regard to EGME.

Effect of Ethylene Glycol

Monoisopropyl Ether (EGPE) on the

Testis and Blood of Rats

Groups of 10 male Alpk/Ap (Wistar-derived) rats

were exposed to air, 300 or 1000 ppm of EGPE for 6

hr/day for 5 days of week 1 and for 4 days of week 2. On

the day following the last exposure, the rats were killed,

blood samples were taken, and the testes were weighed.

Rats in the 1000 ppm EGPE group showed marked

hematuria on the first day of exposure; subsequently, no

more hematuria was seen. The skin of these rats was

pale in appearance up to and including day 4; subsequently their clinical condition appeared comparable to

the controls. The body weight gains of the rat in the

1000 ppm EGPE group were statistically significantly

lower than the controls, while the rats in the 300 ppm

groups had weight gains similar to the control group. At

postmortem the appearance and weights of the testis

from both the 1000 and 300 ppm groups were similar to

the controls.

202
SCREENING AND HAZARD ASSESSMENT

At 1000 ppm, statistically significant reductions were

seen in hemoglobin concentration, red blood cell count

and mean corpuscular hemoglobin concentrations. In

addition, mean cell volumes and mean cell hemoglobin

were statistically significantly higher in the 1000 ppm

group. These changes are consistent with the initial

hematuria observed in these rats. There were no

changes in the white blood cell counts, and apart from a

slight increase in mean cell volume there were no

hematological effects in the rats exposed to 300 ppm

EGPE. This study indicates that EGPE behaves in a

similar manner to ethylene glycol monobutyl ether

(EGBE) (9), i.e., the major effect appears to be

hemolysis, followed by replacement with resistant cells

which are younger and larger. There was no suggestion

of the typical response seen with EGME of testicular

atrophy and leukocytopenia.

sure and throughout a subsequent 14-day observation

period. On day 15 the animals were killed by over

exposure to halothane and subjected to gross macroscopic postmortem examination, which included the

weighing of the testes. The results are shown in Table 4.

Both EGME and EGEE caused reductions in testis

weight when compared with the controls on day 15,

EGME causing an approximately 50% reduction and

EGEE a 20% reduction. Neither EGPE nor EGBE

caused a reduction in testes weight. Clinical observations also revealed another sign associated with the

glycol ethers, hematuria in the animals treated with

EGEE, EGPE or EGBE. EGME did not cause hematuria. These limited data indicate that both EGME

and EGEE can cause testicular atrophy after a single,

albeit, very high, exposure by inhalation. However, even

the saturated vapor from EGPE and EGBE did not

cause testicular atrophy.

Effect of a Single Inhalation Exposure

to Ethylene Glycol Monoalkyl Ethers Effect of a Single Exposure to

(EGME, EGEE, EGPE, EGBE) EGME on the Testis of Male Rats

Ethylene glycol monomethyl ether (EGME) has been

shown to cause testicular atrophy following 90 days

exposure (10) and 10 days exposure (4,7). In these

studies, 300 ppm (= 960 mg/mr3) caused testicular

atrophy. The work reported here was designed to assess

whether a single very high exposure to EGME would

also cause testicular atrophy. In order to determine

whether there is a clear structure activity relationship,

ethylene glycol monoethyl (EGEE), monoisopropyl

(EGPE) and monobutyl (EGBE) ethers were also

examined in the same experiment. Male 250 g Alpk/Ap

(Wistar-derived) rats were exposed for 3 or 4 hr to the

saturated vapor generated by blowing air through the

relevant test material contained in a sintered glass

bubbler. The undiluted vapor was led into 1 L glass

exposure chambers, each containing five rats housed

individually. The atmosphere concentrations were estimated by dividing the weight loss from the bubbler by

the total airflow. The rats were observed during exponble 4. Comparative data on effect of exposure to saturated

vapor of glycol ethers on testis weights at day 15.

Testis weight, % of body weight

Hematuria

Compound and (Present

concentration 3-hr exposure Control or absent)

EGME, 24 mg/L 0.49 ± 0.08 1.06 ± 0.06

(7500 ppm)

EGEE, 17 mg/L 0.78 ± 0.07 1.06 ± 0.06 +

(4500 ppm)

EGPE, 15 mg/L 0.95 0.092 0.98 ± 0.10 +

(3500 ppm)

EGBE, 4 mg/L 0.95 ± 0.06 0.93 ± 0.05 +

(800 ppm)

'4-hr exposure.

*Statistically different from control (Student's t-test, p < 0.05).

The previous study had shown that EGME is capable

of causing testicular weight loss after a single exposure

to a saturated vapor. These studies were designed to

determine the dose-response relationship of the effect

of the single exposure on the testes, to determine the

histological effect, and to determine the time course of

the onset of testicular damage.

In the first study, groups of 20 male Alpk/AP rats

were exposed to 5000, 2500, 1250, 625, 300 or 150 ppm

EGME. A further group of rats was exposed to air only.

Following exposure which took place in 3.4 m3 whole

body exposure chambers, they were returned to their

holding cages for 13 days, and 14 days after treatment,

were killed with an overdose of halothane. At autopsy

the left and right testis were weighed separately and

stored in Bouin's fixative for subsequent processing and

microscopic examination. The caudal epididymides were

weighed together and preserved for processing and

microscopic examination. The accessory glands, seminal vesicle and ventral prostate were weighed and later

examined histopathologically.

There was a dose-related fall in body weight gain

as shown by the body weight immediately before

postmortem. These were accompanied by a dose-related

fall in testes weight which was significant down to 1250

ppm (Fig. 2). There were reductions in the caudal

epididymidal weight and an increase in the accessory

gland weight at 5000 ppm of EGME. Histological

examination of the testes revealed severe bilateral

tubular atrophy with disordered spermatogenesis in

rats exposed to 5000 ppm EGME for 4 hr, with many

tubules showing only stem cells and Sertoli cells.

Similar, though less marked changes, were seen in rats

exposed to 2500 and 1250 ppm EGME. Although testes

weight was not reduced at 625 ppm EGME, maturing

spermatids showed evidence of damage.

203
J. E. DOE

BODY WEIGHT IN -day 13(BWG)

1101

150 12SO

Exposure level E.G.M.E. (ppm - log scale)

TESTIS WEIGHT - day 14 (Right)

5(1

14----%LbeVa l3U

Exposure level E.G.M.E. (ppm - log scale)

CAU~ EPIDIULMAL WEIGHT - y 14 ( )

- -- -'-'' - - - - - - - - - - - - - -- - - - - - - - -

in

a

-_-_- 625 23

Exposure level E.G.M.E. (ppm - log scale)

ACCESSORY GLANDW WErIGHT day 14

--Ep

W"

_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _~

- 30 625 125 250

Exposure level E.G.M.E. (ppm - log stale)

FIGURE 2. Effect on rat testis weight of a single exposure of EGME. Significance as in Figure 1.

In the second study, groups of 90, 8-week-old Alpk/AP

rats were exposed to either 2500 or 1000 ppm EGME

for a single 4-hr period. A further group of 90 rats was

sham-exposed to air. Ten rats per group were killed on

each of days 1, 2, 3, 4, 5, 8, 10, 15 and 19 following

exposure. Individual body weights were recorded on

the day before exposure and subsequently on days 4, 8,

15 and 19. Testes, epididymides and accessory gland

weights were recorded at autopsy. Samples of these

tissues were preserved in Bouin's fixative prior to

examination. The results are shown in Figure 3.

In the second study, testis weight reduction was seen

in exposed rats 48 hr after exposure to both 2500 and

1000 ppm, and testes weights were lower than control

values in both the exposed groups at subsequent time

points. Histological examination of the testes revealed

damage to the germinal epithelium 24 hr after exposure

with primary spermatocytes as the target cells for

EGME. Recovery was not evident at day 19 following

exposure to 2500 ppm EGME, and the germinal epithelium remained disordered. Electron microscopy revealed changes in the Sertoli cells, with cytoplasmic

retraction and swollen mitochondria evident 4 days after

exposure to EGME. There was a biphasic response in

BUG

Cont,'I o

means

110o

2500

Ioo -

3000

Ut. 90

X 0

Cont'l

means 70

6C

2500

110

Ut. oo

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Ut. I

cont ' 1

mans

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Eoh,ft ah%f%^ I

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SCREENING AND HAZARD ASSESSMENT

TESTIS WEIGHT - (Right)

(Values for right and left testes were similar)

8 10

DAYS POST EXPOSURE

CAUDA EPIDIDYMAL WEIGHT - (Paired)

8 10

DAYS POST EXPOSURE

ACCESSORY GLAND WEIGHT

*

Nb-~ **

**

1 2 3 4 5 8 10

DAYS POST EXPOSURE

FIGURE 3. Time course of the effect of a single exposure of EGME in male rats. Significance as in Figure 1.

exposed animals with regard to epididymal weight. The

increased weight at day 4 probably reflects the passage

through the epididymides at this point of large numbers

of immature germ cells shed from the testes following

the initial insult. Subsequently, epididymal weights

were reduced when compared to control values, again

reflecting the situation in the testes where there was a

marked reduction in the number of spermatozoa produced following exposure. EGME would appear to be

exerting its effect primarily on rapidly dividing cells,

with primary spermatocytes being affected initially.

Subsequently, Sertoli cells showed signs of damage and

this may account for the resulting marked loss of

immature germ cells.

These studies indicate that even a relatively short

exposure to EGME vapor can caused marked testicular

atrophy although at high concentrations. Even at concentrations comparable with those causing testicular

atrophy over a 90-day period (i.e., 300 ppm versus

625 ppm), there were subtle effects on spermatogenesis highlighting the need for further study of the

effect of EGME on the testes. The results were

reported previously (11).

General Conclusion

The work reported in this paper demonstrates very

clearly the role of toxicology examinations which do not

110

100

Wt. 90

8C

Cont'l 70

means 6o 2 3 4 5 15 19

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130

120

Wt.

110

^,100

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Cont'l 90

means 80

70

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2 3 4 5 15

**

110

19

Wt.

Contr'

means

100

90

80

15 19

205
206 J. E. DOE

fall clearly into the type of testing currently done for

regulatory purposes. Both the testicular effect of EGME

and the effect on the developing fetus can be seen in

relatively short term and/or inexpensive experiments.

The effect of EGME on the testis has been shown after

10 days exposure or indeed after a single exposure. This

has enabled the effect of other glycol ethers to be

examined very quickly. These data were in many cases

generated before the effects of the other glycol ethers in

regulatory type studies were available and where this

has occurred these studies have accurately predicted

the effect in the regulatory studies. Thus, EGME has

been shown to affect the testis (10). EGBE on the other

hand, does not appear to affect the testis in a similar

manner (9). It is therefore reasonable to assume that

EGPE which did not affect the testes weights after

either 10 days exposure at 1000 ppm or 4 hr exposure to

a saturated vapor would not cause testicular atrophy in

a longer regulatory study.

The effect of EGME upon the developing embryo is

dramatically revealed using the test for teratogenic

potential (6).

At higher levels EGME causes complete resorption of

all embryos and at lower levels all three of the factors

suggested by Chernoff and Kavlock (6) affected, i.e.,

numbers of litters, numbers of pups, their survivability

to day 4. Again, these results are in agreement with the

results of full teratology studies which have been

carried out with EGME. EGME has been shown to be

fetotoxic and teratogenic in the rabbit (10) and in

the mouse (2). This activity of EGME gives added

confidence to the results found with PGME and with

diEGME in this test. Even though EGME has such a

dramatic effect, neither PGME nor diEGME caused

any significant changes in the parameters listed by

Chernoff and Kavlock (6). It is important to remember,

however, that in both the studies reported here the dose

of test compound was not so high that there was severe

maternal toxicity. It is important when trying to

interpret the results of this type of study that extravagantly high doses are not used. These studies give a fair

amount of confidence in predicting that neither diEGME

nor PGME will be significantly teratogenic.

Taken overall, these studies indicate that even with a

limited resource in terms of time and availability of

experimental facilities, significant knowledge can be

gained on the toxicology of a group of compounds,

provided that a good positive control substance, such as

EGME, is available.

The studies reported here were carried out as a collaborative effort

by I. S. Chart, D. Hart, L. K. Head, S. Millward, L. Pinto, D. M.

Samuels, D. J. Tinston, G. A. Wickramaratne and D. R. Woodcock.

Thanks are also due to Miss L. Pritchard for typing this manuscript.

REFERENCES

1. Nagano, K., Nakayama, E., Oobayashi, H., Yamada, T., Adachi,

H., Nishizawa, T., Ozana, H., Nakaichi, M., Okuda, H., Minami,

K., and Yamazaki, K. Embryotoxic effects of ethylene glycol

monomethyl ether in mice. Toxicology 39: 335-343 (1981).

2. Nagano, K., Nakayama, E., Hoyano, M., Dobayski, H., Adachi,

H. and Yamada, T. Testicular atrophy of mice induced by ethylene

glycol monoalkyl ethers. Japan. J. Ind. Health 21: 29-25 (1979).

3. Andrew, F D., Buschbom, R. L., Cannon, W C., Miller, R. A.,

Montgomery, L. F., Phelps, D. W, and Sikov, M. R. Teratologic

assessment of ethylbenzene and 2-ethoxyethanol. Batelle Pacific

Northwest Laboratories, report to NIOSH, 1981.

4. Miller, R. R., Ayres, J. A., Calhoun, L. L., Young, J. T., and

McKenna, J. M. Comparative short-term inhalation toxicity of

ethylene glycol monomethyl ether and propylene glycol monomethyl ether in rats and mice. Toxicol. Appl. Pharmacol. 61:

368-377 (1981).

5. Doe, J. E., and Tinston, D. J. Novel chambers for long term

inhalation studies. In: Proceedings of the Inhalation Toxicology

and Technology Symposium (B. K. J. Leong, Ed.), Ann Arbor

Science Publishers, Ann Arbor, MI, 1981.

6. Chernoff, N. and Kavlock, R. J. A potential in vivo screen for

the determination of teratogenic effects in mammals. Teratology 21 (2): 33A (1980).

7. Doe, J. E., Samuels, D. M., Tinston, D. J., and Wickramaratne,

G. A. Comparative aspects of the reproductive toxicology by

inhalation in rats of ethylene glycol monomethyl ether and

propylene glycol monomethyl ether. Toxicol. Appl. Pharmacol. 69:

43-47 (1983).

8. Doe, J. E., Hart, D., and Wickramaratne, G. A. The teratogenic

potential of diethylene glycol monomethyl ether (DGME) as

assayed in the post natal development test by the subcutaneous

route in rats. Toxicologist 3: 70 (1983).

9. Dodd, D. E., Snellings, W M., and Ballantyne, B. Ethylene

glycol monobutyl ether (EGBE) vapor 9 day and 90 day inhalation

studies in Fischer 344 rats. Toxicologist 2: 159 (1982).

10. Miller, R. R., Ayres, J. A., Young, J. T. and McKenna, M. J.

Ethylene glycol monomethyl ether. I. Subchronic vapor inhalation

study with rats and rabbits. Fundam. Appl. Toxicol. 3: 49-54

(1983).

11. Samuels, D. M., Doe, J. E., and Tinston, D. J. The effects on the

rat testis of single inhalation exposures to ethylene glycol alkyl

ethers, in particular ethylene glycol monomethyl ethers. Paper

presented at European Society of Toxicology Meeting, Rome,

March 1983.