TOXICOLOGY AND APPLIED PHARMACOLOGY 77, 187-200 (1985)



## Acute Toxicity and Recovery in the Hemopoietic System of Rats after Treatment with Ethylene Glycol Monomethyl and Monobutyl Ethers

DEREK 

### GRANT, SUE SUESH, 

### Huw B. JONES, SHARAT D. GANGOLLI,

AND WILLIAM H. BUTLER

The British Industrial Biological ResearchAssociation, Woodmansteme Road,

Carshalton. Surrey, SM5 4DS, England

Received May 10, 1984; acceptedAugust 27, 1984

Acute Toxicity and Recovery in the Hemopoietic System of Rats after Treatment with Ethylene Glycol Monomethyl and Monobutyl Ethers. GRANT, D., SULSH,S.,JONES,H. B., GANGOLLI, S. D., AND BUTLER, W. H. (1985). Toxicol. Appl. Pharmacol. 77, 187-200. Male ratswere given ethylene glycol monomethyl ether (EGM) or ethylene glycol monobutyl ether (EGB) po for4consecutivedaysatdosesof 100or 500 mg/kg body wt/day for EGM, and 500
or 1000mg/kg body wt/day for EGB. Animals were killed on Days 1,4, 8,and 22 atIer the final treatment. Both EGM and EGB produced thymic atrophy and lymphocytopenia and, in the cam of EGM, neutropenia also. Hemolytic anemia induced by EGB resulted in splenic extramedullary hemopoicsis, hypcrplasia ofboth spleen and bone marrow, and rcticulocytosis. Apart from residual slight increases in spleen weight, mean red cell volume, and mean corpuscular hemoglobin atthe end ofthe recovery period, other effectswere reversible. With EGM, reduction in the numbers ofcirculating red cellswas only slight. Treatment with EGM alsoabolished splenicextramedtdkuy hemopoiesis which pamally recoveredon Day4,followed byamarked responseon Day8,and return tothemoderate control valueson Day22.Femoral bone marrow was hemorrhagic 1 day after treatment with EGM which appeared to be associated with sinus endothelial cell damage. By Day 4 the histologic appearance of the
marrow was normal. Testicular atrophy was also produced in EGM-treated animals which persisted for the duration of the experiment. It is concluded that EGM and EGB differ considerably in thespectrum oftoxic changesinduced, and apart from testicular atrophy, these changes were.largely reversible within ashort time of the end oftreatment. 8 1985 Academic Pras. Inc.



### Ethylene glycol monoalkyl ethers are colorless volatile liquids with widespread industrial applications owing to their miscibility with many organic solvents, and with water- and oil-based formulations. The presence of ethylene glycol ethers in paints, lacquers, dyes, inks, and cleaning agents has generated considerable interest in their toxicology. Several
case histories have been reported in which adverse effectswere produced in individuals who had absorbed ethylene glycol ethers through the skin by direct contact (Ohi and Wegmann, 1978), via the lung after inhaling vapors (Avon, 1963), or through the gut when mistaken for alcohol and ingested (Nitter-Hauge, 1970). The spectrum of toxicity induced by the ethylene glycol ethers in various species include disorders of the hemopoietic, reproductive, and central nervous systems, with the site of action and severity of toxicity depending upon the particular ethylene glycol ether and species involved (ECETOC, 1982). In view of the reported differences in toxicity between closely related ethylene glycol ethers (Werner etal., 1943; Carpenter etal.,

187 0041-008X/85 $3.00 Cowi@ 8 1985 by Academic Press, Inc. All rights of npmduction in any form -4.

### 188 GRANT



### 1956; Nagano 

### et al., 1979), a comparative study was initiated using ethylene glycol monomethyl ether(EGM) andethyleneglycol monobutyl ether (EGB) to characterize the differences in toxicity in the hemopoietic systemandthe reversibility of sucheffectsin the period following treatment.

METHODS

Animals. Male F344 rats,agedapproximately 4weeks,
were obtained from Charles River U.K. Ltd., Margate, Kent. They were housed in a room designed to provide
atemperature of 20 + 2°C 12 to 15 air changes per hour, a relative humidity of 40 to 602, and alighting cycleof 12hr light: 12hr dark. Animals were allowed freeaccessto feed(R& M No. 1expanded, Special Diet Services, Witham, Essex) and domestic mains water throughout. Materials. EGM was obtained from Sigma Chemical Company, Poole, Dorset,and EGB from Aldrich Chemical Company, Gillingham, Dorset. The purity ofboth test materials was 99.9% asdetermined by gas-liquid chromatography. Design and treatment. On receipt, ratswere randomly assignedto fivegroups with the aid of random number tables (Fisher, 1965). Eachgroup contained 24 animals, housed 6 to a cage and allowed to acclimatize for 1 week. Animals weredosedpowith water (control), EGM (100 or 500 m&kg/day), or EGB (500 or 1000 mg/kg/ day)daily for4consecutivedays.The glycoletherswere diluted in water and given atadosevolume of5ml/kg bodywt. Thesedosesof EGM and EGB were chosenon the basisofeffects

obtained in preliminary range finding studies. Six animals from each group were bled, then killed, and autopsied on Days 1,4, 8,and 22 after the lasttreatment. Hematology. Animals were warmed in a container heated to approximately 40°C and bled from a lateral caudal vein, and the following were determined: red blood cell count (RBC), white blood cell count (WBC), hemoglobin concentration (Hb), mean red cell volume (MCV), hematocrit (Hct), reticulocyte count, and differential white cell count. The RBC, WBC, Hct,and MCV
were determined on a Coulter Counter (model ZF) coupled to a Coulter MCV/Hct accessory.During the period of reticulocytosis in EGB-treated animals, the WBC was determined by a manual method using an improved Neubauer counting chamber. Hemoglobin concentrations were measured on a Coulter Hemoglobinometer. Necropsy and histopathology. After overnight fasting, animals were weighed and killed by ip injection with sodium pentobarbitone. The spleen,thymus, liver,testes, and kidneys were weighed, and together with cervical ET AL.

and mesenteric lymph nodes,thesewere fixed in neutral buffered Formalin forexamination by light microscopy. Femoral bone marrow was immersed in a solution containing 4% formaldehyde and 1% glutaraldehyde adjusted to pH 7.2 with 0.1 M phosphate buffer (McDowell and Trump, 1976), a fixation method suitable for both light and electron microscopy. Samples of
marrow for electron microscopy were postfixed in 1% osmium tetroxide, dehydrated through graded ethanols, and embedded in agar lOO/araldite. Sectionswere cutat athickness of 1pm and stained with toluidine blue for light microscopy, or atathickness of60 to 90 nm and stained with lead citrate and uranyl acetatefortransmissionelectron microscopy.Other samplesofbone marrow and the remainder of the tissueswere processed into paraffin wax,cut at5pm, and stained with hematoxylin and eosin. Statistical analyses. Data were analyzed by one-way analysis of variance; means were compared by least significant difference criteria (Snedecor and Cochran, 1968).



### RESULTS


### Body Weights and Organ Weights

The mean weights of the animals were similar in all groupsbefore the commencement oftreatment. Treatment-related reductions in the ratesof body weight gain were seenonly in the high-doseEGB group (Table 1). Reducedrelative weights(g/100gbodywt) of kidneys, spleen,testes,and thymus were seenin EGM-treated ratson Day 1of recovery (Table 1).With the exception of weights ofkidneys,thesereductionsweredoserelated. The weightsof spleensrecovered to exceed control values by Day 8. By Day 22, all organweightsin EGM-treated animalswere similarto or slightly higherthan thoseofthe control group apart from weights of testes which remained lowerthroughout. In contrast, EGB causeddose-relatedincreases in therelative weightsofspleen,liver, and kidneys on Day 1 of recovery, but in
common with EGM, thymic weights were reduced (Table 1).Theseweight changesin EGB-treated animalsreturned to normal by Day 22apart from the weightsof livers and spleens which remained slightly raised.ACUTE TOXICITY OF GLYCOL ETHERS 

### 189

TABLE 1

RELATIVE ORGAN WEIGHTS OF MALE F344 RATS DOSED WITH ETHYLENE GLYCOL MONOMETHYL ETHER (EGM) OR ETHYLENE GLYCOL MONOBUTYL ETHER (EGB)

Dose bWkg/day) Body wt (9) Liver Relative organ weight (g/lOOg body w-t)

Spleen Kidneys Testes Thymus

CON

EGM

EGB

CON

EGM

EGB

CON

EGM

EGB

CON

EGM

EGB 0

100

500

500

1000

0

100

500

500

1000

0

100

500

500

1000

0

100

500

500

1000 62.0



### (2.6) 68.S (5.2) 64.5 (3.8) 64.0 

### (5.6) 53.722 (5.3)

16.2 (7.4) 14.2 

### (8.1) 72.7 (4.9) (81.5) (7.4) 56.4*** (7.5)

92.0 (6.4) 105.5* (9.0) 84.5 (14.6) 102.5 (10.1) 71.5’ (11.1)

113.2 (5.8) 171.8 (7.1) 141.8*** (15.0) 162.0 (10.3) 147.5** (17.8) Day 1 recovery

3.869 0.326 (0.157) (0.010) 3.628 0.305 (0.216) (0.0 18) 3.67 1 0.237*** (0.182) (0.028) 4.480*** 0.610*** (0.113) (0.079) 4.921**’ 0.669*** (0.314) (0.030)

Day 4 recovery
3.620 0.337 (0.040) (0.032) 3.542 0.306 (0.165) (0.026) 3.490 0.296 (0.137) (0.028) 4.001** 0.435*** (0.165) (0.018) 4.381*** 0.532*** (0.438) (0.055)

Day 8 recovery 3.789 0.335 (0.115) (0.022) 3.766 0.354 (0.112) (0.011) 3.653 0.431*** (0.161) (0.0 14) 4.019* 0.394** (0.094) (0.018) 4.244+++ 0.436*** (0.277) (0.056)

Day 22 recovery 3.312 0.284 (0.148) (0.021) 3.355 0.211 (0.094) (0.030) 3.446 0.330* (0.191) (0.04 1) 3.315 0.307 (0.112) (0.028) 3.550* 0.338** (0.171) (0.025) I.06 1.051 (0.030) (0.199) 0.969++ 1.016 (0.040) (0.156) 0.997* 0.630** (0.033) (0.100) 1.036 0.954 (0.041) (0.225) 1.191*** 0.891 (0.080) (0.338)

0.913 (0.035) 0.926 (0.031) 0.903 (0.03 1) 0.946 (0.054) 1.085*** (0.059) 1.080

(0.098) 0.946 (0.286) 0.624*** (0.101) 1.152 (0.169) 1.174 (0.126)

0.885 (0.018) 0.863 (0.06 1) 0.908 (0.029) 0.916 (0.042) 1.OOO* (0.113) 1.123

(0.323) 0.909 (0.314) 0.633** (0.034) 1.280 (0.05 1) 1.234 (0.334)

0.719 (0.028) 0.726 (0.033) 0.787** (0.040) 0.720 (0.030) 0.703 (0.192) 1.332

(0.059) 1.356 (0.054) 0.615*** (0.079) 1.377 (0.055) 1.261 (0.268) 0.262

(0.040) 0.214* (0.024) 0.096*** (0.030) 0.261 (0.032) 0.164*** (0.036)

0.309 (0.012) 0.251* (0.030) 0.115*** (0.024) 0.294 (0.023) 0.187*** (0.09 1)

0.298 (0.02 1) 0.258* (0.025) 0.117*** (0.014) 0.275 (0.047) 0.2 13*** (0.039)

0.205 (0.022) 0.198 (0.022) 0.238 (0.043) 0.210 (0.0 17) 0.219 (0.039)

Note. Values are the means of six animals and those marked with an asterisk differ significantly from the control values using least significant difference criteria (*p < 0.05, 

### **p < 0.01, 

### ***p < 0.001). Figures in parentheses
are SD.ACUTE TOXICITY OF GLYCOL ETHERS 

### 191

Histopathology

In the high-dose EGM group, femoral bone marrow wasseverely hemorrhagic, with loss of much of the normal nucleated tissue (Fig. 1).Electron microscopy revealed extensivethinning ofthe sinus endothelial cells of
marrow sinuses which had cytoplasmic processes extending into ‘the lumen (Fig. 2). Large discontinuities which often contained erythrocytes were frequently seen in the endothelial cell layer in the sinuses. These breaks were not at the point of cell to cell contact asthe zonulae adherens were intact. Pinocytotic activity wasnormal asjudged by the numbers and appearance of vesicles, although some cells contained asmall number of autophagosomes and secondary lysosomes. There was also slightly increased electron density and crenation ofsome nuclei. Despite the severity of the initial hemorrhage, the normal architecture of the marrow was restored by Day 4. Only one ofthe six animals in the low-dose group had hemorrhagic marrow on Day 1. In the spleen the moderate amount of extramedulhuy hemopoiesis (EMH) normally present in animals of this age wasabolished bytreatment with 500 mg EGM/kg body wt, and reduced in most of the lOO-mg EGM/ kg body wt group (Fig. 3). This reduced amount of EMH returned to normal or near normal values in the lowdose EGM group within 8 days, whereas in animals of the high-dose group there was partial recovery on Day 4 and a marked increase on Day 8. The degree of EMH had returned to normal by Day 22. Testicular atrophy wassevere in the highdose EGM group with disruption of the architecture of the seminiferous tubules and degenerative changes in the spermatocytes similar to those described by Greasy and Foster (1984). This change wasreflected in the absence ofspermatozoa in the epididymides. Testes were of normal histologic appearance in the low-dose EGM group. Treatment with EGB produced marrow hyperplasia with no evidence of hemorrhage at any time. Splenic EMH was marked in both EGB-treated groups on Day 1 but this change had returned to normal by Day 8. Unlike EGM, treatment with EGB had no discemable effectson the testes. Histologic changes common to ratstreated with either EGM or EGB at the high doses
wereslight hepatic EMH in the first week of recovery and lymphocyte depletion of the thymic cortex on Day 1(Erg. 4). This lymphocyte depletion wasonly transient asthe appearance of thymic tissue was normal by Day 4. Kidneys and lymph nodes were of normal appearance in all groups. Hematology
Changes in the circulating blood wereproduced by both EGM and EGB, but there
were qualitative and quantitative differences in the responses seen. EGM (Table 2) produced slight reductions of the BBC, Hct, and Hb values at Day 4 and 8 posttreatment, resulting in a mild normochromic, normocytic anemia. The reticulocyte count was slightly depressed in these EGM-treated. animals on Days 1 and 4, whereas on Day 8 the numbers of circulating reticulocytes were signScantly increased (p < 0.05). The leukocyte count wasmarkedly reduced in the high-dose EGM group on Day 1 due to reduced numbers of neutrophils and lymFIG. 1. Femoral bone marrow morphology-Day 1recovery. (a) Control rat. Note the well-ordered nucleated marrow tissuesurrounding asinus. (b) Rattreated with 500 mg EGM/k&Iay for4consecutive days.The sinus endothelium is morphologically abnormal with an irregular outline and large gaps in which red blood cellscanbeseen.Much ofthenormal nucleated marrow tissue ismissing, and there are large numbers of mature red cells occupying the extravascular areas. Resin embedded, I-srn sections, stained with toluidine blue stain, X1460.

### 192 GRANT ET AL.

FIG,2.Sinus endothelial cell ultrastructure-Day 1recovery. (a) Control rat. The nuclei and luminal surfacesof the endothelium are smooth in outline. (b) Rat treated with 500 mg EGM/kg/day for 4 consecutivedays.The endothelial cell nuclei areveryirregular in outline and the cytoplasm variesfrom extreme thinness toareasofthickening with frequent knob-like processes.Lead citrate and uranyl acetate staining, x2030. Insetshows irregular endothelium with intact zonulae adherens, x7190.



### phocytes. These leukocyte counts subse- Animals treated with the lowerdoseof EGM quently increasedbut failed to reach control had slightly reduced leukocyte counts on values by the end of the recovery period. Day 1.ACUTE TOXICITY OF GLYCOL ETHERS 

### 193

In contrast, EGB (Table 2) caused pronounced dose-dependent effectson circulating red and white cells. Animals treated with the highest dose of EGB (1000 mg/kg body wt/ day) had reduced RBC, Hct, Hb, and markedly elevated MCV, reticulocyte counts, and
mean corpuscular hemoglobin (MCH) atthe end of treatment. Most of these changes in the red cells disappeared over the recovery period, although the MCV and MCH remained slightly elevated at Day 22. The reticulocytosis was particularly striking on Days 1and 4, but by Day 8the reticulocyte counts had returned to normal. Examination of Romanowski-stained blood films taken from this group revealed increased numbers of circulating nucleated RBCs (normoblasts), pronounced anisocytosis, polychromasia, and the presence of Howell Jolly bodies (Fig. 5). These changes had resolved by Day 8. Leukocyte counts were also depressed on Day 1 in the high-dose EGB group, due principally to a reduction in the numbers of circulating lymphocytes. The leukocyte numbers gradually increased, although, in common with EGM-treated animals, they did not reach control values by the end of the recovery period. These effects on red and white cells
were also seen in animals treated with the lower dose of EGB (500 mg/kg body w-t/ day), although the severity of the changes
waslessmarked.

DISCUSSION

The two ethylene glycol ethers, EGM and EGB, which differ only in their alkyl moieties, produced several markedly dissimilar changes in circulating blood and selected tissues after acute treatment. The major hematological effect of EGM
wasleukopenia, whereas EGB produced both leukopenia and anemia. The pronounced leukopenia in EGM-treated animals appeared to be associated with reductions in both lymphocytes and neutrophils, asopposed to those treated with EGB in which lymphocytes were preferentially affected. With both ethylene glycol ethers there was also a loss of lymphocytes from the thymic cortex and reductions in thymic weights, but whether this change indicates direct toxicity to the thymus or is mediated by possible effects on circulating lymphocytes or bone marrow is unclear. Since lymphocytopenia was common to animals treated with either of the ethylene gIyco1 ethers, but marrow toxicity wasconfined to those treated with EGM, the lymphocytopenia does not appear to be related to bone marrow toxicity. There does, however, seem to be some association between neutropenia and bone marrow toxicity in EGM-treated animals. The impairment of granulopoiesis in damaged marrow would be expected to lead to arapid neutropenia owing to the short half-life of neutrophils in the circulating blood (Wade and Mandell, 1983). Full recovery from the leukopenia produced by the ethylene glycol ethers was equivocal owing to the raised number of circulating leukocytes in control animals at the end of the recovery period. The reductions in RBC, Hct, Hb, and marginally lower reticulocyte counts during the initial period of recovery from EGM treatment, together with reduced splenic EMH and bone marrow toxicity, suggest an inhibitory action on erythropoiesis. If continued this inhibition would eventually lead to anemia by the failure to replace effete red cells. Indeed, it hasbeen reported that anemia
occurs in rats and several other specieswhen the duration of exposure to EGM is more prolonged (Miller et al., 1981; Doe et al., 1983). On Day 8 the markedly elevated splenic EMH and the modest increase in the numbers of circulating reticulocytes suggest recovery from the inhibition of erythropoiesis. In contrast, the various changes seen in circulating red cells after EGB treatment are typical of hemolytic anemia with an associated reticulocytosis and hyperplasia of both bone marrow and spleen. This observation isin agreement with the findings ofCarpenter

### 194 GRANT ET AL.ACUTE TOXICITY OF GLYCOL ETHERS 

### 195

FIG.3.Splenic extramedullary hemopoiesis (EMH)-Day 1recovery. (a)Control rat. Islands of EMH
areclearlydistinguishable inthe red pulp area.(b) Rattreatedwith 500 mg EGM/kg/day for4consecutive days.The red pulp isdevoid ofanyovert EMH. (c) Rattreated with 1000 mg EGB/kg for 4consecutive days. Note the marked increase in EMH which occupies amajor proportion ofthe red pulp, H & E, X290.

et al. 

### (1956) who have demonstrated hemolysis and increased osmotic fragility in rats treated with EGB. The majority of these effects on the red cell population were reversible, although increasesin MCH and MCV weremore per& tent, probably due to the selective hemolytic action of EGB on aged red cells (Carpenter et al., 

### 1956), leaving a population of young erythrocytes. In addition, erythrocytes which
are produced during remission from severe acute anemia tend to be macro&c and contain more Hb (Schalm et al., 

### 1975),factors which may also contribute to the persistence of these changes. The long-lasting spleen weight increases produced by these ethylene glycol ethers may be a residual effect of the elevated concentration ofsplenic EMH. In contrast to the proliferative response in the marrow of EGB-treated animals, those given EGM had severely hemorrhagic marrows. The major site of toxicity relating to hemorrhage appeared to be endothelial cells lining the sinuses, many of which had evidence of damage. The presence of mature erythrocytes in the large gaps of the sinus walls and their abundance in the substance ofthe marrow tissue suggestthat the integrity ofthe sinus endothelium wasimpaired. This, together with the loss of the resident population of nucleated cells, agreeswith previous reports of marrow hypocellularity after treatment with EGM (Miller et al., 

### 1981). The pathogenesis of nucleated marrow cell loss is unclear at present.

Testicular atrophy similar to that described by Foster et al. 

### (1983) and Creasyand Foster (1984) was evident in animals treated withTABLE 2

HEMATOL~CY OF MALE F344 RATS DOSED WITH ETHYLENE GLYCOL MONOMETHYL ETHER (EGM) OR ETHYLENE GLYCOL MONOBUTYL ETHER (EGB)

Dose RBC b%.Wday) (lo’2/1) MCV (fl) Hct (%) Hb (g/d]) Reticulocytes

(IRBC) MCH

(PP) Total Leukocytes (109/1)

Neutrophils Lymphocytes

CON

EGM

EGB

CON

EGM

EGB 0

100

500

500

loo0

0

100

500

500

1000 6.55 60.2 38.7

(0.15) 

### (1.2) (1.5) 6.58 59.7 38.8 (0.28) (0.5) (1.3) 6.67 59.3 39.2 (0.2 1) 

### W-V (1.3) 5.04*** 74.5*** 37.3 (0.50) 

### (2.2) (2.7) 3.34*** 82.7*** 27.8*** (0.6 I) (2.5) (4.3)

6.65 59.2 (0.3 1) 

### 03.8) 6.64 57.7 (0.47) 

### W3) 6.22 57.5 (0.17) 

### (0.6) 6.06* 68.7*** (0.28) (2.9) 4 80*** ” (0:72) 77 6*- ’

(4:2) 38.7

(1.5) 37.7 (2.7) 35.3* (1.4) 41.2 

### (1.2) 37.2” (4.4) Day 1 recovery

15.7 4.2h (0.3) 

### (1.8) 15.7 3.9 

### (0.6) W) 15.4 2.7 (0.4) (1.0) 14.0* 25.2*** (1.1) (9.3) 10.6*** 32.6**+ (1.9) 

### (6.9)

Day 4 recovery
16.3 10.4 (0.5) (3.6) 15.9 9.1 

### (0.6) (5.1) 14.4*** 6.6” (0.4) (5.9) 16.7 22.4+ (0.5) (9.9) 15.3*” 51 3*** U 

### (1.2) (11:8) 24.0 4.93 0.55 4.37 (0.4) (1.19) (0.26) (1.03) 23.8 3.85; 0.5 1 3.33** (0.5) (0.31) (0.09) (0.31) 23.1 1.52-f 0.13**‘ I.30*** ‘ 0 (0.5) (0.32) (0.02) (0.17) F 27.9*** 2.68*** 0.48 2.16*** 5 (1.0) (0.61) (0.16) (0.54) 3I.8*** 1.75-e 0.46 1.29”* $ 

### (1.6) (0.39) (0.29) (0.19) F

24.5 5.23 0.54 

### (0.6) (0.52) (0.2 I) 24.0 6.04** 0.37 (1.4) (0.60) (0.08) 23.2 3.00*** 0.10*** (0.5) (0.31) (0.07) 27.6** 4.59* 0.59 

### (1.8) (0.48) (0.29) 32.1*** ” 2 36*** a

(0140) 0.32*” (2.3) (0.12) 4.58 (0.47) 5.60*** (0.56) 2.90*** (0.26) 3.95* (0.29) 1 77+++ ” (0:36)CON

EGM

EGB

CON

EGM

EGB 0

100

500

500

1000

0

100

500

500

1000 6.94

(0.29) 6.99 (0.28) 6.46. (0.25) 6.23*** (0.30) 6.16*** (0.39)

7.63 (0.25) 7.72 (0.12) 7.39 (0.70) 7.46 (0.2 1) 7.36 (0.15) 60.5 41.5



### (0.6) (2.1) 59.2 41.0 

### (1.2) (2.2) 60.5 38.5* (1.1) 

### (2.1) 65.2*** 40.3 

### (1.2) (2.4) 71 o***” (3:2) 42.8”

(2.3)

58.7 43.8 (1.4) 

### (2.1) 57.3f 43.5 (0.5) 

### (0.6) 58.7 42.7 

### (0.8) (5.0) 59.0 43.4 (0.9) 

### (1.6) 61.2*** 44.5 

### (0.8) (0.6) Day 8 recovery

16.7 5.5 (0.7) (1.9) 16.1 (0.3) (3) 15.5** 

### (0.8) ,;:;; 16.5 (0.7) (K) 17.4 6.9 (0.7) 

### (2.6)

Day 22 recovery
16.7 3.2 (0.5) (1.1) 16.5 3.8 

### (0.2) (1.3) 16.9 3.2 (1.3) (1.0) 17.2 (0.4) ,::,“, 17.3 

### (0.2) (2) 24.0 6.13 0.72 5.32 (0.5) (1.08) (0.16) (1.04) 23.1 5.39 0.54 4.81 

### (0.8) (0.85) (0.24) (0.69) 23.9 4.49- 0.52 3.91** 

### (0.6) (0.58) (0.18) (0.52) 26.5*** 5.52 1.10 4.39’ (0.9) (0.47) (0.30) (0.27) 29.4*** 5.55” 1.29** 4.24” 

### (1.2) (0.82) (0.56) (0.51)

21.9 9.90 1.22 8.61 (0.3) (2.97) (0.45) (2.70) 21.3’ 8.43 0.84* 6.04*+ 

### (0.2) (3.97) (0.28) (0.46) 22.9 6.25*+ 1.00 5.20*** (0.7) (0.8 1) (0.19) (0.75) 23.0*** 6.06** 1.26 4.76+++ (0.3) (0.41) (0.19) (0.32) 23.5*** 5.30** 1.06 4.18*** (0.5) (0.42) (0.28) (0.43)

Note. Values are the means of six animals and those marked with an asterisk differ significantly from the control values using least significant difference criteria (*p 4 0.05, 

### **p < 0.01, 

### l**p -C 0.001). Figures in parentheses are SD. ” Mean value of five animals. hMean value of four animals. ’Mean value of three animals.FIG. 4. Thymus morphology-Day I recovery. (a) Control rat. Note the clear distinction between the outer cortex and inner medulla. (b) Rat treated with 500 mg EGM/kgjday for 4 consecutive days. There is considerable depletion of lymphocytes from the cortex and an increased lymphocyte population in the medulla. The demarcation between the two areas is less distinct, H & E, X60.

198ACUTE TOXICITY OF GLYCOL ETHERS 

### 199

FIG. 5. Blood film from a rat given 1000 mg EGB/kg/day for 4 consecutive days-Day I recovery. Note the pronounced polychromasia, anisocytosis,and nucleated red blood cell (normoblast) containing
a Howell Jolly body. May-Grunwald-Giemsa stain, X1460.



### 500 mg EGM/kg body wt. This atrophy persisted for the duration of the recovery period, whereas EGB had no discemable effect on either testis weights or on their histologic appearance at doses which were clearly toxic to the hemopoietic system. The studies by Foster et al. 

### (1983) have shown that a longer period of recovery, equivalent to one maturation cycle (7to 8weeks), after dosing rats with 500 mg EGM/kg body wt permits the resumption ofspermatogenesis. In conclusion, it is clear that there are considerable differences between EGM and EGB in many of the toxic changes induced, and that, apart from testicular atrophy, most of the other histologic and hematologic changes resolve within 3weeks or lessofthe cessation of treatment. The reasons for the differential toxicity between the two glycol ethers are unknown, but it is likely that events related to the metabolism of the two compounds are involved asthe acetic acid metabolites appear to be largely responsible for the toxic effects induced by the ethylene glycol ethers (Miller et al., 

### 1982, 1983).

ACKNOWLEDGMENTS

Wethank Mrs. L. Beechforthe preparation ofspecimens for electron microscopy, and Mr. A. Dagnall for photographic assistance.This work ispart ofa research program on acute/chronic toxicity relationships, and we thank the Food and Drink Industries Council fortheir support.


### REFERENCES

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