Glycollic acid

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Study initiated on 16 October 1995. Study completed on 20 June 1996.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Reason / purpose for cross-reference:

reference to other study

Remarks:

Test doses in the present study determined after pilot study summarised in the cross-reference

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Reference 2

Reference Type:

publication

Title:

Developmental Toxicity Study of Glycolic Acid in Rats

Author:

Munley SM, Kennedy GL, Hurtt ME

Year:

1989

Bibliographic source:

Drug and Chemical Toxicology, 22(4), 569-582

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EU Directive 87/302/EEC (OJ No L133 31.5.88)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPP 83-3 (Prenatal Developmental Toxicity Study)

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: MAFF NohSan 59, No. 4200.

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Glycollic acid

EC Number:

201-180-5

EC Name:

Glycollic acid

Cas Number:

79-14-1

Molecular formula:

C2H4O3

IUPAC Name:

2-hydroxyacetic acid

Details on test material:

Glycolic acid >99%

Specific details on test material used for the study:

99.6% purity.

Test animals

Species:

rat

Strain:

other: Crl:CD.BR

Remarks:

The rat was selected to provide data in a rodent species. The Crl:CD BR strain was chosen because historical control data are available from the literature, the supplier, and previous studies at the laboratory.

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, North Carolina.
- Age at study initiation: Males were 77 days old on arrival and females were 63 days old on arrival.
- Weight at study initiation: Males weighed between 306.1 and 371.6 g on arrival and females weighed between 180.6 and 230.5 g on arrival.
- Fasting period before study: N/A
- Housing: The rats were housed individually in suspended, wire-mesh, stainless steel cages. Nesting material was not provided because the dams were euthanized prior to expected parturition.
- Diet (e.g. ad libitum): The rats were provided Purina Certified Rodent Chow@ #5002 (Meal) ad libitum.
- Water (e.g. ad libitum): The rats were supplied with tap water from United Water Delaware ad libitum.
- Acclimation period: All animals were quarantined for at least 6 days and then released for the study by the laboratory veterinarian.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 +/- 1°C.
- Humidity (%): 50 +/- 10%.
- Photoperiod (hrs dark / hrs light): 12 hours light, 12 hours dark with fluorescent lighting.

IN-LIFE DATES: From: 16 October 1995 (first day of dosing) To: 4 December 1995 (last fetal evaluation).

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Solutions of the test material in deionized water were prepared weekly during the study and stored in the refrigerator. Sufficient amounts of solution needed for dose administration were removed from the refrigerated containers daily.
Glycolic acid was administered by gavage because the oral route is a potential route of accidental human exposure. The dose volume was 10 mL/kg and individual dosages were based on the most recently recorded body weights.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples (-30 mL each) of each test solution were taken three times during the study: October 16, October 30, and November 6, 1995. Analysis of the first sampling addressed concentration and stability. For the second and third samplings, analyses addressed concentration. Samples were submitted to the Analytical Group of Environmental Sciences (ES) at Haskell Laboratory.

At the first sampling, two samples from the vehicle and each test solution were collected. One set of samples was submitted and analysed shortly after submission and the remaining samples were refrigerated for 7 days and then analysed for stability. At the second and third samplings, one sample of the vehicle and each test solution were analysed shortly after submission, to verify concentration. Analysis of glycolic acid in dosing formulations was conducted according to the following method.

Aliquots of the supplied dosing samples were pipetted into a beaker containing phenyl violet indicator/ice slurry. The volume of the aliquots varied based on the concentration of the supplied dosing samples. Targeted concentrations were 75 or 150 mg of glycolic acid for each titration. The aliquots were titrated with a standardised volumetric sodium hydroxide solution (0.251N - Baker Chemical) to the free acid equivalence point of the 99% glycolic acid. All glycolic acid samples were analysed the day they were prepared and submitted.

Details on mating procedure:

- Impregnation procedure: cohoused.
- M/F ratio per cage: 1:1.
- Length of cohabitation: until copulation was confirmed. Mating began on October 9, 1995, with Gestational Day (GD) 1 occurring from October 10-13, 1995 and October 16-19, 1995.
- Proof of pregnancy: presence of a copulation plug in the vagina or on the cage board referred to as day 1 of gestation.

Duration of treatment / exposure:

On days 7-21 of gestation (14 days). Females were euthanized on gestation day 22.

Frequency of treatment:

daily

Duration of test:

14 days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

600 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

300 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

150 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

75 mg/kg bw/day (nominal)

No. of animals per sex per dose:

25 females

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose concentrations were based on the results of a rat pilot developmental toxicity study. In the pilot study, test concentrations of 70% technical solution were administered to groups of 8 bred rats over gestational days 7-21. Dose concentrations were 0, 125, 250, 500, and 1000 mg/kg bw. Maternal toxicity was demonstrated at 500 and 1000 mg/kg bw/day. At 1000 mg/kg bw/day, maternal effects included mortality, significantly reduced maternal body weights, weight changes and reduced food consumption. Dose-related increases of the following clinical observations were observed: abnormal gait and mobility, lung noise, salivation, stained and wet fur. At 500 mg/kg bw/day, similar, yet markedly less severe evidence of maternal toxicity was demonstrated and there was a slight but significant reduction in maternal weight gain. The incidences of lung noise and wet fur were significantly increased as well. Developmental toxicity was evident at 500 and 1000 mg/kg bw/day. At 1000 mg/kg bw/day, mean foetal weight was significantly reduced. Embryolethality was significantly increased and among the surviving foetuses, malformations and variations were significantly increased. At 500 mg/kg bw/day, the significant reduction in foetal weight persisted as did the increase in foetal variations. There was no evidence of either maternal or developmental toxicity at 250 or 125 mg/kg bw/day.

- Rationale for animal assignment (if not random): Before dosing began, females selected for the study that copulated during the first week of mating were ranked by their body weights on gestational day 1 and randomly assigned to control or experimental groups. Females selected from the second week of mating were similarly assigned. The randomization resulted in a distribution in which the mean body weights for all groups were not statistically different (p=0.9744). In addition to random assignment to groups, bias was controlled by coding all females prior to scheduled sacrifice. They remained coded during the collection of the post-mortem and foetal data.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS / DETAILED CLINICAL OBSERVATIONS: Yes
- Observations for morbidity and mortality were made daily.
- Individual clinical signs were recorded each morning on Days 1-22G and each afternoon on Days 7-21G (the dosing period).

BODY WEIGHT: Yes
- Time schedule for examinations: Females were weighed on Days 1 and 7-22G.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food was weighed on Days 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21 and 22G.
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 22
- Organs examined: the organs of the thoracic and abdominal cavities were examined for gross pathologic changes.

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other:
- the types of implants (live and dead foetuses, and resorptions) were counted and their relative positions were recorded.
- the empty uterus was weighed.

Fetal examinations:

Live foetuses were weighed, sexed, and examined for external alterations.

To identify stunted foetuses for each litter, the maximum stunted weight (MSW) was calculated by subtracting the lightest weight from the total weight, dividing by the remaining number of foetuses, and multiplying by 0.666. A foetus weighing less than or equal to the MSW was considered stunted. If the lightest foetus was determined to be stunted, the procedure was repeated until all remaining foetal weights were in excess of the MSW.

Retarded renal development was classified using the schema of Woo and Hoar.

- External examinations: Yes: all per litter.
- Soft tissue examinations: Yes: half per litter. The first live foetus and thereafter each alternate one removed subsequently was decapitated and examined for visceral alterations.
- Skeletal examinations: Yes: all per litter. All foetuses were fixed, stained (Alizarin red S) and examined for skeletal alterations.
- Head examinations: Yes: all per litter.

Statistics:

Sequential trend testing was applied to the developmental toxicity data for each parameter as listed below. If a significant dose-response was detected, data from the top dose group was excluded and the test repeated until no significant trend was detected. For litter parameters, the proportion of affected foetuses per litter or the litter mean was the experimental unit for statistical evaluation. The level of significance selected was p<0.5. Where the data were tied and the standard large sample version of Jonckheere's test was not applicable, exact p values were calculated using permutation methodology.

Maternal weight, weight changes and food consumption: Linear contrast of means from ANOVA.

Live foetuses, dead foetuses, resorptions, nidations, copora lutea, incidence of foetal alterations: Jonckheere's test.

Incidence of pregnancy, clinical observations, maternal mortality, females with total resorptions early deliveries: Cochran-Armitage test.

Foetal weight, sex ratio: Linear contrast of least square means from ANCOVA.

Indices:

N/A

Historical control data:

N/A

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

At 600 mg/kg bw/day, there were significantly increased incidences of adverse clinical observations; abnormal gait/mobility was observed in 5 of 25 dams at this level. Lung noise (wheezing and/or rattling after dosing which persisted into the afternoon and occasionally until the following day) and/or irregular respiration was seen in 8 of 25 dams. Two dams were lethargic. Lung noise similar to that observed at 600 mg/kg was observed in 2 of 25 dams dosed at 300 mg/kg. This incidence approached statistical significance (p=0.0553). No remarkable clinical observations were observed at 150 or 75 mg/kg bw/day.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Maternal body weights and weight changes were reduced at 600 mg/kg bw/day. Final body weight (Day 22G) and the final weight adjusted for the products of conception were significantly reduced. Statistically significant reductions in body weight changes were observed over Days 7-9,17-19, 19-21,21-22, and 7-22G. Weight changes calculated using the adjusted final body weight (final body weight minus the products of conception) were also significantly reduced (Days 7-22 and 1-22G). Maternal body weights, weight changes, and adjusted final body weights were unaffected at dose levels of 300 mg/kg bw/day and lower.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Maternal food consumption was significantly reduced at 600 mg/kg bw/day over Days 21-22G. Maternal food consumption was unaffected at dose levels of 300 mg/kg bw/day and lower.

Food efficiency:

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no significant post-mortem findings at any dose level.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Maternal developmental toxicity

Number of abortions:

not specified

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not specified

Changes in number of pregnant:

not specified

Other effects:

no effects observed

Description (incidence and severity):

There were no dose-related effects on reproductive outcome parameters: dams with total resorptions, mean corpora lutea, mean number of implantations, litter size or sex ratio.

Details on maternal toxic effects:

N/A

Effect levels (maternal animals)

1

Dose descriptor:

LOAEL

Effect level:

300 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOEL

Effect level:

150 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

Maternal abnormalities

Abnormalities:

no effects observed

Results (fetuses)

Fetal body weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean foetal weight was significantly reduced at 600 mg/kg bw/day.

Reduction in number of live offspring:

not specified

Changes in sex ratio:

not specified

Changes in litter size and weights:

not specified

Anogenital distance of all rodent fetuses:

not examined

Changes in postnatal survival:

not specified

External malformations:

not specified

Skeletal malformations:

effects observed, treatment-related

Description (incidence and severity):

The incidence of specific malformations was significantly increased at 600 mg/kg/day. These malformations were fused and absent ribs, fused and hemivertebra, and abnormally fused and cleft/non-fused sternebra. At 300 mg/kg, there was a slight increase in the incidence of two skeletal malformations, fused ribs and fused vertebra. The incidences for these findings, 2 foetuses from 2 litters, approached statistical significance (p=0.0555). There were no compound-related malformations at 150 or 75 mg/kg. The incidence of variations was increased at 600 mg/kg bw/day. The specific variations were misaligned and incompletely ossified sternebra and incompletely ossified vertebra. There were no increased foetal variations at 300 mg/kg bw/day and lower.

Visceral malformations:

not specified

Details on embryotoxic / teratogenic effects:

N/A

Effect levels (fetuses)

1

Dose descriptor:

LOAEL

Effect level:

300 mg/kg bw/day

Basis for effect level:

other: embryotoxicity & teratogenicity

2

Key result

Dose descriptor:

NOEL

Effect level:

150 mg/kg bw/day

Basis for effect level:

other: embryotoxicity & teratogenicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

yes

Lowest effective dose / conc.:

300 mg/kg bw/day (nominal)

Treatment related:

yes

Relation to maternal toxicity:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Any other information on results incl. tables

Table 6.8.1-01a   Summary of reproductive outcomes

Dose (mg/kg bw/day)	0	75	150	300	600
Corpora lutea  (mean /litter)	16.3	16.3	15.8	17.4	17.0
Implantations  (mean /litter)	15.4	15.0	14.7	15.3	15.9
Total number of resorptions  (mean /litter)	0.7	0.8	0.7	0.6	0.6
Total number of foetuses  (mean /litter)	14.6	14.2	14.0	14.7	15.3
Total number of live foetuses  (mean /litter)	14.6	14.2	14.0	14.7	15.3
Mean foetal weight	5.02	5.20	5.17	5.05	4.38
Sex ratio  (No. of male foetuses /No. live foetuses)	0.44	0.49	0.50	0.50	0.48

Table 6.8.1-01b   Summary of skeletal anomalies

Dose (mg/kg bw/day)	0	75	150	300	600
Skeletal examination number examined (No. of litters)	366  (25)	354  (25)	321  (23)	339  (23)	351  (23)
Fused rib  (No. of litters)	0  (0)	0  (0)	0  (0)	2  (2)	9  (9)
Absent ribs  (No. of litters)	0  (0)	0  (0)	0  (0)	0  (0)	3  (3)
Fused vertebra  (No. of litters)	0  (0)	0  (0)	0  (0)	2  (2)	6  (6)
Hemi-vertebra  (No. of litters)	0  (0)	1  (1)	0  (0)	1  (1)	8  (8)
Abnormally fused sternebra  (No. of litters)	0  (0)	0  (0)	0  (0)	0  (0)	3  (3)
Cleft/non-fused sternebra (No. of litters)	1  (1)	0  (0)	0  (0)	1  (1)	6  (5)
Misaligned sternebra (No. of litters)	1  (1)	1  (1)	0  (0)	1  (1)	13  (9)
Incompletely ossified sternebra (No. of litters)	14  (6)	4  (2)	13  (2)	12  (3)	60  (14)
Incompletely ossified vertebra (No. of litters)	95  (19)	119  (21)	68  (17)	114  (22)	204  (21)

Applicant's summary and conclusion

Conclusions:

There was no evidence of either maternal or developmental toxicity at either 150 or 75 mg/kg bw/day. Thus, the maternal and developmental no-observed-effect level (NOEL) was 150 mg/kg bw/day. Therefore, the results indicate that glycolic acid is not likely to be uniquely toxic to the rat conceptus, developmental effects were only apparent at maternally toxic doses.

Executive summary:

Glycolic acid was administered by gavage to groups of 25 Crl:CD BR female rats on days 7-21 of gestation at daily dose levels of 0, 75, 150, 300, or 600 mg/kg.

Compound-related, adverse maternal and developmental toxicity was observed at 600 mg/kg. Maternal effects included significant reductions in maternal weight changes, body weights, and food consumption. Adverse clinical observations were significantly increased at this level as well as included abnormal gait/staggering, lung noise (wheezing and/or rattling), irregular respiration, and lethargy. There were no remarkable post-mortem findings in the dams. Developmental toxicity was evident at this level as well as significantly reduced mean fetal weight and a significant increase in the incidence of fetal malformations and variations.

Marginal evidence was detected at 300 mg/kg bw/day. Regarding maternal toxicity, lung noise similar to that observed at 600 mg/kg bw/day was observed in 2 of 25 dams (p=0.0553). Developmental toxicity was evident only as a slight but not statistically significant (p=0.0555) increase in the incidence of skeletal malformations. No other maternal or developmental parameters were affected.

There was no evidence of either maternal or developmental toxicity at either 150 or 75 mg/kg bw/day. Thus, the maternal and developmental no-observed-effect level (NOEL) was 150 mg/kg bw/day. Therefore, the results indicate that glycolic acid is not likely to be uniquely toxic to the rat conceptus, developmental effects were only apparent at maternally toxic doses.