Glycollic acid

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Guideline compliant Commission Directive 87/302/EEC, U.S. EPA Pesticide Assessment Guidlines Subdivisoin F, 82-1 ( 1982), OECD Test Guideline 407 , Maff Japan NohSan No. 4200 ( 1985). This study was selected as the key study because the information provided for the hazard endpoint is sufficient for the purpose of classification and labelling and/or risk assessment.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPP 82-1 (90-Day Oral Toxicity)

Deviations:

yes

Remarks:

Ophthalmological findings and gross pathological lesion incidences were not evaluated by statistics (EPA FIFRA).

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Commission Directive 87/302/EEC

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: MAFF Japan NohSan 40 4200

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 70% solution

Test animals

Species:

rat

Strain:

other: Crl:CD (SD) IGS.BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, North Carolina.
- Age at study initiation: 48 days. Birthdated 18 August 1998. Rats were circa 5 weeks old at time of receipt.
- Group mean bodyweight range at time of allocation to study was 229.5 to 232.6g for males and 163.7 to 165.0g for females.
- Housing: With the exception of some portions of the reproductive study, all rats were housed one per cage, sexes separate, in stainless steel, wire-mesh cages suspended above cage boards.
- Diet (e.g. ad libitum): All rats were fed PMI Nutrition International, Inc. Certified Rodent Checkers LabDiet@ 5002 ad libitum.
- Water (e.g. ad libitum): Tap water was provided ad libitum.
- Acclimation period: Upon arrival at Haskell Laboratory, the rats were quarantined for six days of the 13-day pretest period.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23" +/- 1°C.
- Humidity (%): 50% +/- 10%.
- Photoperiod (hrs dark / hrs light): 12 hours light, 12 hours dark, with fluorescent light.
IN-LIFE DATES: From: October 5, 1998 To: March 1, 1999

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

Dose solutions were prepared at 15, 30 and 60 mg/mL on a daily basis and administered on same day by gavage in a dose volume of 10 mL/kg bw to achieve dose levels of 150, 300, and 600 mg/kg per day, based on the most recently recorded weight. For pregnant rats, from gestation day 18 until delivery, dose volumes were based on the gestation day 18 body weights. Dosing solutions were stored refrigerated until used. Dosing solutions stored beyond 14 days after preparation were not administered to animals. Control animals were dosed with commercially-supplied water only.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

HPLC: Hewlett-Packard 1090; UV 210 nm; column: Zorbax® SB-C18, 4.6 mm x 150 mm; 40°C mobile phase: 2.0% acetonitrile/98.0% 3.1 mM H3P04; 1.0ml/min., Injection volume: 4.0 µl
From each dosing solution sample, 1 mL was aliquoted and diluted to 10 mL with high-performance liquid chromatography (HPLC) grade water, then mixed. The 0 mg/mL and 15 mg/mL solutions were analyzed without further dilution. The 30 mg/mL and 60 mg/mL samples were further diluted with HPLC grade water to an expected concentration of 1.5 mg/mL active ingredient (a.i.) prior to analysis. Samples submitted for analysis were analyzed the day the solutions were prepared by the testing group.

Duration of treatment / exposure:

90 days with continuation of treatment through to test day 131 (males) or day 21 of lactation (females) for animals in reproduction phase.

Frequency of treatment:

daily administration

Doses / concentrations

Remarks:

Doses / Concentrations:
150, 300 or 600 mg/kg bw ; 15, 30 and 60 mg/mL administered in 10 mL/kg bw volume of water

No. of animals per sex per dose:

40 per sex per dose level. Each dosage group was divided into subchronic toxicity, immunotoxicity, neurotoxicity, and reproductive toxicity subsets (10 per sex per subset per concentration).

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose levels for this study were selected based on results from a developmental toxicity study in which glycolic acid was administered by gavage to Crl:CD%R female rats (25 per group) on days 7-21 of gestation at daily dose levels of 0, 75, 150, 300, or 600 mg/kg per day.
- Rationale for animal assignment (if not random): Rats were selected for study use on the bases of adequate body weight gain, freedom from any
clinical signs of disease or injury, and a body weight within 20% of the mean within a sex. In error, two rats with ophthalmological abnormalities were assigned to study groups in the reproduction subset (see Examinations below.) The selected rats were distributed by computerized, stratified randomization so that there were no statistically significant differences among group body weight means within a sex. See study design table in "other information section".

Positive control:

A positive control study involved collecting sera from animals previously injected with SRBC and the immunosuppressive agent, Cyclophosphamide. Serum was analysed for SRBC-specific IgM antibody.

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes.
- Time schedule: Daily.
DETAILED CLINICAL OBSERVATIONS: Yes.
- Time schedule: Cage-site examinations to detect moribund or dead rats and abnormal behavior and appearance among rats were conducted at least once daily throughout the study. Moribund and dead rats were submitted for a gross and microscopic examination. At every weighing, each rat was individually handled and examined for abnormal behavior and appearance. Rats designated for neurotoxicity evaluations had cage-site examinations approximately one to two hours after dosing on test day 1, and approximately one to two hours after dosing on one day during the weeks that the functional observational battery was conducted.

BODY WEIGHT: Yes.
- Time schedule for examinations: All rats were weighed once per week during the 90-day feeding phase of the study. In addition, the neurotoxicity sub study rats were weighed on the days of neurotoxicity evaluation. During the reproduction sub-study, male rats were weighed on a weekly schedule and female rats were weighed during gestation on days 0, 7, 14, 18, and 21 and weighed during lactation on days 0, 7, 14 and 21. Female rats without a known start of gestation and female rats that copulated but did not deliver a litter continued to be weighed weekly.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)/ FOOD EFFICIENCY
The amount of food consumed by each rat over each weighing interval was determined throughout the study. Food consumption was determined for each female rat designated for reproductive assessment on gestation days 0, 7, 14 and 21. From these determinations and body weight data, mean daily food consumption and mean food efficiency were calculated.

OPHTHALMOSCOPIC EXAMINATION: Yes.
- Time schedule for examinations: Two ophthalmological examinations were conducted by a veterinary ophthalmologist. Both eyes were examined by focal illumination and indirect ophthalmoscopy. The examinations were conducted under subdued lighting after mydriasis had been produced with a 1% tropicamide solution. On test day 9, the initial examination was performed on all rats received for the study, prior to selection and grouping. In error, two rats with ophthalmological abnormalities were assigned to study groups in the Reproduction subset. Since animals in the reproductive toxicity subset did not undergo ophthalmological examination during the study, the pre-existing abnormalities had no impact on the study. All surviving rats designated for subchronic toxicity were examined on test day 86 prior to the scheduled sacrifice.

HAEMATOLOGY: Yes. Blood cell counts, hemoglobin concentration, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration were determined on a Serono Baker 9000@ hematology analyzer. Differential cell counts were determined on a Hematrak@Automated Differential System cell counter. Absolute values for the various types of leukocytes were calculated from the leukocytic data.
- Time schedule for collection of blood: Test days 46 and 93 for male rats and test days 47 and 94 for female rats.
- Anaesthetic used for blood collection: Carbon dioxide.
- Animals fasted: Yes, for 16 hours.
- How many animals: 10 per sex per dose.
- Parameters examined were: Red blood cell count (RBC), neutrophil count (Neut), white blood cell count (WBC), band neutrophil count (Band), platelet count (PLT), lymphocyte count (Lymph), hemoglobin concentration (HGB), atypical lymphocyte count (Alym), hematocrit (HCT), monocyte count (Mono), mean corpuscular volume (MCV), eosinophil count (Eosin), mean corpuscular hemoglobin (MCH), basophil count (Baso) and mean corpuscular hemoglobin concentration
(MCHC).
CLINICAL CHEMISTRY: Yes.
Clinical chemical parameters were measured on a Boehringer Mannheim Mtachi 717 clinical
chemistry analyzer using Boehringer Mannheim reagents.
- Time schedule for collection of blood: Test days 46 and 93 for male rats and test days 47 and 94 for female rats.
- Animals fasted: Yes, for 16 hours.
- How many animals: 10 per sex per dose.
- Parameters examined were: Alkaline phosphatase activity (ALP), alanine aminotransferase activity (ALT), aspartate aminotransferase activity (AST), sorbitol dehydrogenase activity (SDH), bilirubin concentration (BILRN), cholesterol concentration (CHOL), total protein concentration (TPROT), albumin concentration (ALBMN), globulin concentration (GLOBN), glucose concentration (GLUCO), urea nitrogen concentration (BUN), creatinine concentration (CREAT),
phosphate concentration (PHOS), calcium concentration (CALC), sodium concentration (Na), potassium concentration (K) and chloride concentration (Cl).

URINALYSIS: Yes.
Osmolality was determined on a Precision Systems model Multi-OsmetteTM2 430 osometer. Urine biochemical constituents were measured on a ClinitekTM2 00 urine chemistry analyzer using Ames MultistixTMurine chemistry dipsticks. Urine appearance (color and transparency) was recorded and the sediment from each specimen was microscopically examined.
- Time schedule for collection of urine: One day prior to each bleeding time, an overnight (approximately 16 hour) urine specimen was collected from each rat to determine:
- Metabolism cages used for collection of urine: Yes.
- Animals fasted: Yes, 16 hrs.
- Parameters examined were volume (VOL), glucose, osmolality (OSMOL), urobilinogen (UROBL), PH, ketone (acetoacetic acid), hemoglobin or blood (BLOOD), protein and bilirubin.

NEUROBEHAVIOURAL EXAMINATION: Yes.
- Time schedule for examinations: Test days 3 and 4 before substance administration and test days 9 and 10, 45 and 46, and 86 and 87 before substance administration.
- Dose groups that were examined: All dose groups.
- Battery of functions tested: Functional observational battery test.

OTHER: Immunotoxicity Evaluation:
- Humoral Immune Fuction and Immune Organ Weights.
- Test day 23, 10 animals per sex per dose were injected IV with Sheep Red Blood Cells.
- Test day 29 animals sacrificed.
- Spleen and Thymus removed and weighed.
- Serum collected and analyzed for IgM antibody.

OTHER: Reproductive Assessment:
- Test day 97, females housed 1:1 ratio with random males of same dose level.
- In post-partum examination live and dead pups were counted, live pups were weighed and each pup was examined for abnormal behavior and appearance.
- Examinations took place on postpartum days 0, 4, 7, 14, and 21.

Sacrifice and pathology:

Ophthalmoscopic examinations: on day -9, surviving rats on day 86
Haematology, clinical chemistry, urinalysis: 10 per sex /group, day 46/47 and 93/94
Sacrifice and pathology: Organ Weights, Gross and histopathology, Immunotoxicology evaluations
GROSS PATHOLOGY: Yes- The liver, kidneys, adrenal glands, testes(male), ovaries(female) and brain were all weighed at sacrifice. Gross lesions were perserved.
HISTOPATHOLOGY: Yes - All collected tissues ( except nasal tissue without gross lesions) from all animals in the control and high concentration groups were processed and received a full histopathological examination.

Other examinations:

Haematology, clinical chemistry, urinalysis: 10 / sex /group; day 46/47 and 93/94
Immunotoxicology Evaluations - Using the Humoral Immune Function and Immune Organ Weights.
Neurotoxicity Evaluations - Functional Observation battery and Motor activity tests were used.
Reproductive Assessment - Breeding, gestation and lactation procedures

Statistics:

The various statistical analyses applied to each of the study phases are fully detailed in the study report and included, as appropriate, one way analysis of variance followed by Dunnett’s test or linear contrasts; Kruskal-Wallis test followed by Dunn’s test; Cochrane-Armitage trend test; Jonckhere-Terpstra trend test; linear contrast by least square means; Levene’s test for homogeneity, Shapiro-Wilk test for normality, Bartlett’s test.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

effects observed, treatment-related

Endocrine findings:

not examined

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

Samples of glycolic acid 70% solution, as supplied for the study, were analysed near the beginning and end of the study to determine stability over the course of study duration. The analysis results confirmed suitable long term stability. The average percent active ingredient on test day 2 was 69.5% and on test day 143 was 68.1%.

The dosing solutions were analysed on test day 1, the range was 95 to 109.3% of nominal, and the mean values for each test group were 98.7%, 98.3% and 107.5% for low, intermediate and high concentrations. Further analyses were completed for test days 37, 86, 99 and 142. Results were consistently within the acceptable range of nominal (92.0 to 102.7% were recorded values).

Male and female rats given 300 and 600 mg/kg per day had lower mean body weight, overall body weight gain, food consumption and food efficiency. No adverse clinical signs indicative of systemic toxicity and no test substance-related ophthalmological findings were observed. Toxicologically significant increased neutrophil levels in male rats dosed with 300 mg/kg per day or 600 mg/kg per day and increased urea nitrogen, phosphorus, and creatine and decreased urine concentration in male rats dosed with 300 mg/kg per day or 600 mg/kg per day were observed. Mean kidney weight of male rats in the 300 mg/kg per day and 600 mg/kg per day were significantly higher than that of the control group. Gross findings of renal pelvis dilatation were observed and correlated with microscopic findings of oxalate crystal nephrosis and unilateral hydronephrosis in males dosed with 300 or 600 mg/kg per day.

No toxicologically important changes in immunotoxicity parameters were measured. No toxicologically important changes in the behavourial parameters of neurotoxicity were measured. No compound-related gross lesions or microscopic findings observed in tissues of the nervous system or skeletal muscle examined. No toxicologically important changes in the measures of reproductive function. No compound-related gross observations were noted in the P1 females or the F1 weanlings. Compound-related gross lesions in the kidneys of male rats in the 600 mg/kg per day group similar to those found in subchronic toxicity evaluation were noted.

Effect levels

1

Dose descriptor:

NOEL

Remarks:

systemic toxicity

Effect level:

150 mg/kg bw/day (actual dose received)

Sex:

male

Basis for effect level:

histopathology: non-neoplastic

2

Dose descriptor:

NOEL

Remarks:

systemic toxicity

Effect level:

600 mg/kg bw/day (actual dose received)

Sex:

female

Basis for effect level:

other: highest dose tested

3

Dose descriptor:

NOEL

Remarks:

neurotoxicity

Effect level:

600 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: Neurotoxicity The FOB evaluations showed no treatment-related effects in either males or females dosed at 150, 300 or 600 mg/kg bw/day.

4

Dose descriptor:

NOEL

Remarks:

immunotoxicity

Effect level:

600 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: There were no treatment-related adverse effects on spleen or thymus and glycolic acid did not affect the primary humoral immune response to SRBC.

5

Dose descriptor:

LOAEL

Effect level:

300 mg/kg bw/day (actual dose received)

Sex:

male

Basis for effect level:

histopathology: non-neoplastic

6

Dose descriptor:

NOAEL

Remarks:

overall

Effect level:

150 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: effect observed in males at ≥300 mg/kg

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Any changes observed in rats treated with 150 mg glycolic acid/kg per day were either considered not to be compound related or not adverse. Under the conditions of this study, the overall NOEL for both male and female rats was 150 mg/kg per day.

The NOELs for this study are based on 100% glycolic acid dosed (adjusted for 70% purity of the test substance).

Applicant's summary and conclusion

Conclusions:

In the subchronic phase of the study, administration of the higher dose levels (300 and 600 mg/kg bw/day) were associated in males with reduced bodyweight gain, reduced food conversion efficiency and neutrophilia (various lesions consistent with an inflammatory response to irritation of the lungs or trachea were considered secondary to aspiration of the acidic test material and not resulting from systemic exposure to glycolic acid). The primary target organ was the kidney. Renal effects included weight changes, clinical pathology consistent with reduced glomerular filtration rate and reduced ability to concentrate urine, and microscopic evidence for oxalate crystal nephropathy for males dosed at 300 and 600 mg/kg bw/day. Similar effects were not present among the treated females.

The NOEL for male rats exposed to glycolic acid 70% solution was found to be 150 mg/kg bw/day based on renal oxalate crystal nephropathy observed at higher doses. The NOEL for females in this study was 600 mg/kg bw/day, the highest dose tested, based on the absence of primary findings in the target organ.

Neurotoxicity: The FOB evaluations showed no treatment-related effects in either males or females dosed at 150, 300 or 600 mg/kg bw/day. The NOEL for neurobehavioural effects was 600 mg/kg bw/day.

Immunotoxicity: There were no treatment-related adverse effects on spleen or thymus and glycolic acid 70% solution did not affect the primary humoral immune response to SRBC. The immune system does not appear to be a primary target for glycolic acid activity. The NOEL for immunotoxicological endpoints in male and female rats was 600 mg/kg bw/day.

Reproductive toxicity: The NOEL for reproductive toxicity was 600 mg/kg bw/day, based on the absence of treatment related effects on reproductive function. The NOEL for reproductive organ pathology in both the P1 generation and the F1 weanlings was 600 mg/kg bw/day, based on the absence of gross pathological changes.

The NOELs for this study are based on 100% glycolic acid dosed (adjusted for 70% purity of the test substance).

Executive summary:

In the subchronic phase of the study, administration of the higher dose levels (300 and 600 mg/kg bw/day) were associated in males with reduced bodyweight gain, reduced food conversion efficiency and neutrophilia (various lesions consistent with an inflammatory response to irritation of the lungs or trachea were considered secondary to aspiration of the acidic test material and not resulting from systemic exposure to glycolic acid). The primary target organ was the kidney. Renal effects included weight changes, clinical pathology consistent with reduced glomerular filtration rate and reduced ability to concentrate urine, and microscopic evidence for oxalate crystal nephropathy for males dosed at 300 and 600 mg/kg bw/day. Similar effects were not present among the treated females.

 

The NOEL for male rats exposed to glycolic acid 70% solution was found to be 150 mg/kg bw/day based on renal oxalate crystal nephropathy observed at higher doses. The NOEL for females in this study was 600 mg/kg bw/day, the highest dose tested, based on the absence of primary findings in the target organ.

 

Neurotoxicity:

The FOB evaluations showed no treatment-related effects in either males or females dosed at 150, 300 or 600 mg/kg bw/day. The NOEL for neurobehavioural effects was 600 mg/kg bw/day

 

Immunotoxicity:

There were no treatment-related adverse effects on spleen or thymus and glycolic acid 70% solution did not affect the primary humoral immune response to SRBC. The immune system does not appear to be a primary target for glycolic acid activity. The NOEL for immunotoxicological endpoints in male and female rats was 600 mg/kg bw/day.

 

Reproductive toxicity:

The NOEL for reproductive toxicity was 600 mg/kg bw/day, based on the absence of treatment related effects on reproductive function. The NOEL for reproductive organ pathology in both the P1 generation and the F1 weanlings was 600 mg/kg bw/day, based on the absence of gross pathological changes. 

The NOELs for this study are based on 100% glycolic acid dosed (adjusted for 70% purity of the test substance).