Hydrocotyle asiatica, ext.

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Hydrocotyle asiatica, ext.

EC Number:

283-640-5

EC Name:

Hydrocotyle asiatica, ext.

Cas Number:

84696-21-9

Molecular formula:

not applicable

IUPAC Name:

Hydrocotyle asiatica, ext.

Test material form:

solid

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

28-day

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Control (conr oil)

Doses / concentrations 2

Dose / conc.:

100 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Examinations

Observations and examinations performed and frequency:

The wet weight of a subset of tissues was taken and a set of organs/tissues was preserved.
A full histopathological evaluation of the tissues was performed on high dose and control animals. Any gross lesion macroscopically identified was examined microscopically in all animals.

Results and discussion

Results of examinations

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

No clinical signs were observed in all animals of the male and female control and LD group. All animals of the female MD group were seen without clinical findings during the treatment period. For two males in the MD group and all males and females in the HD group polydipsia/polyuria was recorded directly after test item application on several days in the first half of the treatment period. As no other clinical findings were noted and clinical biochemistry, haematology or histopathological evaluation showed no test item-related findings, the transiently observation polydipsia/polyuria was not considered to be an adverse systemic toxicological finding and might be a local reaction on test item treatment.
Weekly detailed clinical observation showed a statistical significant decrease of 100% below control for animal sleeps and an increase of animal moving in cage (400 % above control) in the male HD group in week 3. In females, a statistical significant decrease was found for animal sleeps (60 % below control) and presence of grooming in the MD group in treatment week 1.
Furthermore, the parameter animal sleeps was statistically significantly decreased in the MD group (100 % below control) and the HD group (80 % below control) and for the presence of animal moving in cage in the MD and HD group in week 2. The findings were seen transiently in single weeks and without dose relation. Considering the absence of systemic clinical findings the statistical significances are considered to be without toxicological relevance.
In males and females, no toxicological relevant effects were observed in any of the parameters of the functional observation battery before and at the end of the treatment period when compared with the controls.
Statistical significances were seen in in the week before the administration period (in males for the absence of animal sleeps and the presence of moving in cage in the HD group, an increase of response to handling in the MD group (25 % above control), a decrease of 30 % below control for head touch in the LD group and rearing supported in the MD group (75.76 % below control), an increase of equilibrium reflex in the male LD and MD group (40 % above control), in females for the response to handling in the LD group (19.05 % above control)) and in the last week of treatment in females for the absence of animal sleeps in the LD, MD and HD group, in all dose groups for animal moving in cage (400 % above control), response to handling in all dose groups (16.67% below control in all dose groups), rearing supported (113.33 % above control in the LD group) and in the female MD group for the absence of urination.
As the statistical significances were found in the week before the first administration but not in the last treatment week or were found without dose dependency and additionally, clinical observation showed no systemic findings, the differences were not considered to be of toxicological relevance. There were no biologically relevant differences in body temperature between the groups.
With exception of a moderate decrease in body weight change in all female dose groups in week 3 of the treatment period (LD 61.17 %, MD 76.47 % and HD 67.06 % below control), no statistical significant changes were found for mean body weight and mean body weight gain in all male and female dose groups. Mean body weight increased during the entire treatment period in all male and female dose and control groups. There was no test item-related effect on body weight development in this study.

Mortality:

no mortality observed

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

There was no test item-related effect on food consumption observed. No statistical significances were seen in all male and female dose groups.

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

The test item had no effect on haematological parameters and coagulation parameters determined at the end of the treatment period.
In males, statistical significant differences compared to the control group were seen for the slight increases of mean WBC count (63.36 % above control), mean HGB (9.11% above control) and mean HCT (7.89 % above control) in the MD group. A slight decrease of mean monocytes was found with statistical significance in the LD and MD group (LD: 31.77 % and MD 34.12 % below control). Mean MCHC was slightly and statistically significantly increased in the female MD and HD group (MD 4.11 % and HD 4.36 % above control).
No dose dependency was noted for the statistical significant parameters in males, the differences were seen in one gender and histopathological evaluation showed no test item-related effects on the examined organs. Therefore, a toxicological effect of the test item was not considered.
There was no test item-related effect on the coagulation parameters considered. No statistical significant changes were found for coagulation parameters in all male and female dose groups and mean values were comparable to the corresponding control groups.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

The test item had no effect on parameters of clinical biochemistry determined at the end of the treatment period.
Mean ASAT was slightly and statistically significantly decreased in the female HD group when compared to control (30.67 % below control). A decrease between 26.33 % (male LD) and 35.88 % (male MD) was found in male groups but without dose dependency and statistical significance. In the female HD group, a slight statistical significant decrease was noted for Urea. The mean value was found with a decrease of 23.78% below control. In males, a decrease of urea was found with 11.29 % below control and without dose dependency or statistical significance. The statistical significant changes were found in one gender and as no toxicological effect was found at microscopic evaluation, a toxicological effect of the test item was not considered.

Urinalysis findings:

no effects observed

Description (incidence and severity):

No test item-related effect on urinary parameters was found.
Macroscopically, a cyst at the greater curvature of the stomach was seen in one male control animal and a bladder stone was found in the urinary bladder of one HD male animal. No other macroscopic findings were noted for all other animals in the study. As the findings were seen in one control animal or in one single HD animal and under consideration of histopathological assessment, the gross lesions were considered to be incidental and not induced by the treatment with the test item.

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

Absolute weight of pituitary gland was slightly and statistically significantly increased in the female MD group (19.66 % above control), but no dose dependent increase was seen for absolute and relative organ weight. Under consideration of the absence of microscopic test item-related findings, the increase is not considered to be of toxicological relevance. No adverse effects of the test item on organ weights were found.

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No changes related to the test item administration were observed at histopathological evaluation. The histopathological NOEL (no observed effect level) may be established at 1000 mg/kg bw.

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Target system / organ toxicity

Key result

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

The test item Centella asiatica dry ext. did not cause indicators of toxicity under the conditions of this study. No adverse effects of the test item were found at all tested dose levels. The NOAEL may be established at 1000 mg/kg bw.

Executive summary:

On the basis of this 28-day Repeated Dose Oral Toxicity study with Centella asiatica dry ext. in male and female Wistar rats with dose levels of 100, 300, and 1000 mg/kg body weight day the following conclusions can be made.

The test item Centella asiatica dry ext. did not cause indicators of toxicity under the conditions of this study. No adverse effects of the test item were found at all tested dose levels. The NOAEL may be established at 1000 mg/kg bw.