Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1986
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Acceptably documented study which meets basic scientific principles and contains sufficient detail to be able to judge the results reliable as a contribution to the understanding of the repeat dose toxicity of this substance. There were a number of deviations from current guidelines noted, but none that render the results unreliable.
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
publication
Title:
A sub-chronic dermal exposure study of diethylene glycol monomethyl ether and ethylene glycol monomethyl ether in male guinea pigs
Author:
Hobson DW, D'Addario AP, Brunser RH, Uddin DE
Year:
1986
Bibliographic source:
Fund Appl Toxicol 6, 339-48

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
: no opthalmology; males only, n=6; no opthalmology
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): diethylene glycol monomethyl ether
- Physical state: liquid
- Analytical purity: 99%
- Supplier: Aldrich Chemical cCompany Inc, Milwaukee.

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals, Scottdale, Pa
- Age at study initiation: 6-8 weeks
- Weight at study initiation: 521-530g
- Housing: 2 per polycarbonate cage, with hardwood, coarse grade bedding.
- Diet (ad libitum): Purina guinea pig chow 5025, Ralston Purina Inc, St Louis, Mo
- Water ad libitum
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 70
- Humidity (%): 40
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST SITE
- Area of exposure: shaved back.
- % coverage: 2x2cm guaze patches.
- Type of wrap if used: hypoallergenic surgical tape (Blenderm) held in place with a Stockinette bandage.
- Time intervals for shavings or clipplings: no data.

REMOVAL OF TEST SUBSTANCE: none reported

USE OF RESTRAINERS FOR PREVENTING INGESTION: none reported
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
13 weeks. Dermal exposures for 6hours per day.
Frequency of treatment:
5 days/week
Doses / concentrationsopen allclose all
Dose / conc.:
40 mg/kg bw/day
Dose / conc.:
200 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
6 (7 for control)
Control animals:
other: yes, 0.9% saline solution
Details on study design:
- Dose selection rationale: Based on an adequate range to cover anticipated exposures through use as a jet fuel deicer plus a substantial safety margin with a 5 fold difference in doses to give a range for evaluation of potential toxic responses elicited by the guinea pig at exposure levels greatly in excess of any anitipcated human exposure.
- Rationale for study: Since the study was designed to look for an alternative to methoxyethanol, for which the critical effect is testicular toxicity, only males were used in the study.
Positive control:
2-methoxyethanol at a dose of 1g/kg

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
Time schedule for examinations: daily prior to dosing
- Cage side observations: signs of intoxication.

DERMAL IRRITATION (if dermal study): Yes

BODY WEIGHT: Yes
- Time schedule for examinations: daily prior to dosing

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Following euthenasia at end of study
- Anaesthetic used for blood collection: Yes (identity) / No / No data
- Animals fasted: Yes
- How many animals: ot specified, presumed all.
- Parameters checked: RBC, Hgb, Hct, MCV, MCH, MCHC, WBC count. Differential WBC count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Following euthenasia at end of study
- Animals fasted: Yes
- How many animals: not specified, presumed all.
- Parameters checked: ALT, AST, BUN, ALP, CK, G-GTP, LDH. Also serum testosterone

URINALYSIS: Yes
- Time schedule for collection of urine: last 24 hours of exposure
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked: ALP, Ca, creatinine, ALP, ALT and G-GTP (latter 3 to detect kidney damage), specific gravity. Creatinine clearance was also calculated

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
Animals sacrificed by anaesthetic overdose (Halothane)

GROSS PATHOLOGY: Yes. adrenals, intestines ( duodenum, jejunum, ileum, colon), kidneys, liver, lung, lymph node, nerve, pancreas, seminal vesicles, muscle, skin, spleen, stomach, salivary gland, testes, thyroid, thymus, trachea, urinary bladder, urocyst. Testes preserved in Bouin's solution, all other tissues from organs above in neutral buffered formalin.

HISTOPATHOLOGY: Yes. Tissues examined by embedding in parrafin, sectioning and examination after H&E staining by light microscopy
Statistics:
One way analysis of variance: Serum and urinary clinical chemistry, terminal body weights, organ weights, haematology values. Differences between groups analysed using Duncan's multiple range test. Daily body weights evalued using two way ANOVA, with dose and experimental day as variables tested.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Dermal irritation:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Positive control animals showed body weight gain reductions only.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Significant adverse changes were seen in the positive control animals. In animals dosed with 2-(2-methoxyethoxy)ethanol, the only change noted was a slight but statistically significant decrease in MCHC in the high dose animals. It was noted that the actual mean figure for the mid dose group was lower than for the high dose group but the bigger standard deviation for the mid dose group did not render it significant. The biological significance of this finding is not likely to be significant.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
The high dose animals showed a significant increase in serum LDH values (at p=0.05 but not p=0.01), similar to the mean levels seen with the positive control. However, whilst a dose reponse was seen, the standard deviation was high which brings into question the significance of the result. The biological significance of these findings is not clear.
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
All treated animals, including the positive control animals, showed increased urinary calcium excretion with no clear dose response. thuis change was seen without any evidence of renal mineralisation or other damage can can be rationalised as a secondary consequence of the elimination of the acid metabolite of the test substance. It is believed that they result in the formation of calcium salts, which then leads to increased urinary calcium levels by decreasing the number of calcium ions available for tubular reabsorption by the kidney. No other changes were seen.
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Animals in the mid and high dose groups showed both relative and absolute decreased spleen weights (at p<0.05 but not p<0.01). These changes were even more pronounced in the positive control animals.
Gross pathological findings:
no effects observed
Description (incidence and severity):
The only treatment related lesions were seen in the positive control animals.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
No testicular lesions or evidence of toxicity to the lymphoid system were seen other than in the positive control animals. The only significant changes notedin animals exposed to 2-(2-methoxyethoxy)ethanol were mild, periportal, hepatocellular vacuolar fatty change to the liver. Whilst these finding were regarded as evidence for mild, reversible hepatotoxicity and seen at all doses, it was noted that the changes did not show a clear dose/response relationship. The evaluation of these observed changes is difficult to rationalise as they were seen in the absence of any other effects that would be expected, such as increased liver weight. In addition, as the guinea pig is not commonly used for sub-chronic tests, there is no background data on fatty changes to the liver. It is therefore unclear if these change is an adverse change or not.
Histopathological findings: neoplastic:
not examined

Effect levels

Dose descriptor:
NOAEL
Effect level:
40 mg/kg bw/day (nominal)
Sex:
male
Basis for effect level:
organ weights and organ / body weight ratios
Remarks on result:
other: Fatty vacuolisation in the liver, particularly incidence and severity seen in low dose group, are considered to be of minimal significance and Ca levels in urine are secondary effects.

Target system / organ toxicity

Critical effects observed:
not specified
Lowest effective dose / conc.:
200 mg/kg bw/day
System:
haematopoietic
Organ:
spleen
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified

Any other information on results incl. tables

Statistically significant changes reported (standard deviation in brackets

 End point  Negative control  Positive control  High dose  Mid dose  Low dose
 Spleen weight (absolute) (g)  1.38 (0.25)  0.82 (0.26)*  1.02 (0.19)*  0.99 (0.15) *  1.27 (0.39)
Serum LDH (IU/liter)   53.7 (29.5)  144.2 (52.8)*  134.5 (100.4)*  68.8 (31.1)  55.4 (33.5)
 Blood MCHC (%)  34.9 (0.7)  34.5 (1.0)  33.8 (0.6)*  33.3 (1.8)  34.7 (0.4)
 Urine Calcium (mg/day)  1.4 (0.6)  3.3 (1.1)*  4.2 (2.2)*  3.8 (1.9)*  3.9 (1.0)*
 Fatty liver changes (# animals)  0/7  - 6/6   6/6  2/6

Applicant's summary and conclusion

Conclusions:
A number of the statistically significant changes seen in this study are difficult to explain biologically, especially when considering the relativel low doses at which they were observed.
Executive summary:

In a dermal sub-chronic toxicity study which followed the basic requirements of a guideline study, guinea pigs were exposed occlusively to 2 -(2 -methoxyethoxy)ethanol at doses of 40, 200 and 1000mg/kg for 13 weeks. Exposed animals showed decreased splenic weights in the high and medium dose groups. Isolated but significant changes were also seen in the high dose groups clinical chemistry (LDH level) and haematology (MCHC level). Changes were seen in all animals urine chemistry (calcium concentration) but this was attributed to be a secondary consequence of the elimination of metabolites. Some minor fatty changes were seen in the livers of some animals in all dose groups, but it was difficult to explain the biological significance of these in the absence of any changes to any other liver parameters (eg weight) and were therefore considered to be of minimal significance. Based on this analysis, the no effect level from this study is 40mg/kg, although the suspicion that this is likely to be a very conservative finding (effects seen are not very much more severe at 1000mg/kg) and the effects may not be 'adverse' and certainly not 'significant toxic effects', facts that need to be borne in mind in selecting appropriate assessment factors for the risk characterisation. Synopsis: NOAEL (dermal, guinea pig) = 40mg/kg