Benzoic acid, 4-hydroxy-, C18-22-alkyl esters

Endpoint summary

Description of key information

NOAEL oral (subchronic), rat ≥ 1200 mg/kg bw/day

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

26 Jan - 20 Jul 1972

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Basic data given. Publication is written in Japanese.

Principles of method if other than guideline:

The study was conducted before appropriate test guidelines and GLP were developed. The test item was administered orally to rats in the diet for 25 weeks. Clinical signs, food consumption and body weights and blood parameters were observed. At study termination, gross necropsy and histopathological examinations were conducted.

GLP compliance:

no

Limit test:

no

Species:

rat

Strain:

other: SD-JCL

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:
- Age at study initiation: 5 weeks
- Diet: CE-2, ad libtum
- Water: ad libtum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 1 °C
- Humidity (%): 50 ± 5%

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
- Mixing appropriate amounts with (Type of food): 0.2, 1, 2%

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

25 weeks

Frequency of treatment:

daily

Remarks:

Doses / Concentrations:
0.2, 1 and 2%
Basis:
other:

Remarks:

Doses / Concentrations:
120, 600, and 1200 mg/kg bw/day
Basis:
other: calculated based on 50 - 60 g/kg bw day food intake

No. of animals per sex per dose:

6, 5 and 5 males in 2%, 1% and 0.2% group
5 females in each group

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: based on LD50 in mice

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: once a week

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- 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

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Yes

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: after last administration
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all surviving animals
- Examined parameters: RBC, Hb, Ht, WBC, WBCP

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: after last administration
- Animals fasted: No data
- How many animals: all surviving animals
- Examined parameters: GOT and GPT, alkaliphosphate, cholinesterase, protein, A/G ratio, total chlesterol, blood sugar

URINALYSIS: Yes
- Time schedule for collection of urine: after last administration
- Metabolism cages used for collection of urine: No
- Animals fasted: No data
- Examined parameters: protein, sugar

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes: heart, lung, liver, spleen, kidney and other main organs

HISTOPATHOLOGY: Yes: heart, lung, liver, spleen, kidney

Statistics:

t-test or chi-square test

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Significant higher body-weight was observed in males in 0.2% group. Significant lower body weight was observed in males in 1 and 2% group.

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not examined

Dose descriptor:

NOAEL

Effect level:

>= 1 200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: corresponding to 2% test material in diet, calculation based 50 - 60 g/kg bw day food intake

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises adequate, reliable (Klimisch score 2) and consistent studies from reference substances with similar structure and intrinsic properties. Read-across is justified based on common functional groups, common precursors/breakdown products and similarities in physicochemical and toxicological properties (refer to endpoint discussion for further details).
The selected study is thus sufficient to fulfil the standard information requirements set out in Annex VIII-IX, 8.6, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Justification for grouping of substances and read-across

No data are available on the acute toxicity of Benzoic acid, 4-hydroxy-, C18-22-alkyl esters (CAS 201305-16-0). In order to fulfil the standard information requirements set out in Annex VIII-IX in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006, read-across from structurally related substances is conducted.

In accordance with Article 13 (1) of Regulation (EC) No 1907/2006, "information on intrinsic properties of substances may be generated by means other than tests, provided that the conditions set out in Annex XI are met.” In particular for human toxicity, information shall be generated whenever possible by means other than vertebrate animal tests, which includes the use of information from structurally related substances (grouping or read-across).

Having regard to the general rules for grouping of substances and read-across approach laid down in Annex XI, Item 1.5, of Regulation (EC) No 1907/2006, whereby physicochemical, toxicological and ecotoxicological properties may be predicted from data for reference substance(s) by interpolation to other substances on the basis of structural similarity, the substances depicted in the table below are selected as source substances for assessment.

The read-across is based on the identified structural similarities and the likelihood of common breakdown products by biological processes (metabolism). A detailed analogue approach justification is provided in the technical dossier (see IUCLID Section 13).

CAS	201305-16-0 TARGET SUBSTANCE	99-76-3	120-47-8	94-13-3	94-26-8
Chemical Name	Benzoic acid, 4-hydroxy-, C18-22-alkyl esters	Methyl 4-hydroxybenzoate	Ethyl 4-hydroxybenzoate	Propyl 4-hydroxybenzoate	Butyl 4-hydroxybenzoate
MW	390.60-446.71 g/mol	152.15 g/mol	166.18 g/mol	180.2 g/mol	194.23 g/mol
Repeated dose toxicity: oral	WoE RA: CAS 99-76-3 RA: CAS 120-47-8 RA: CAS 94-13-3 RA: CAS 94-26-8	Experimental result: NOAEL (rat, chronic) male/female ≥ 5500-5900 mg/kg bw/day NOAEL (dog, subchronic) male/female ≥ 1000 mg/kg bw/day	Experimental result: NOAEL (rat, subchronic) male/female ≥ 1200 mg/kg bw/day NOAEL (rat, subchronic) male/female 900-1200 mg/kg bw/day	Experimental result: NOAEL (rat, chronic) male/female ≥ 5500-5900 mg/kg bw/day NOAEL (dog, subchronic) male/female ≥ 1000 mg/kg bw/day	Experimental result: NOAEL (rat, subchronic) male/female 900-1200 mg/kg bw/day

 

Discussion

Repeated dose toxicity

Oral

Subchronic

No data are available on the subchronic oral toxicity of Benzoic acid, 4-hydroxy-, C18-22-alkyl esters (CAS 201305-16-0), but there are reliable studies on the structurally related substances Methyl 4-hydroxybenzoate (CAS 99-76-3), Ethyl 4-hydroxybenzoate (CAS 120-47-8), Propyl 4-hydroxybenzoate (CAS 94-13-3) and Butyl 4-hydroxybenzoate (CAS 94-26-8) available, which are used for read-across based on the analogue approach.

A study was conducted to assess the subchronic oral toxicity of Ethyl 4-hydroxybenzoate (CAS 120-47-8) (Sado et al., 1973). The study was performed before appropriate test guidelines and GLP were developed. The test item was daily administered to 5 SD-JCL rats per sex per dose in the diet at concentrations of 0.2, 1 and 2% (corresponding to 120, 600, and 1200 mg/kg bw/day) for 25 consecutive weeks. Concurrent control animals received plain diet. The doses administered were selected on the basis of the LD50 previously determined in mice. Clinical signs, food consumption, body weights and blood parameters were examined. At study termination, gross necropsy and histopathological examinations on heart, lung, liver, spleen and kidneys were conducted. No mortalities occurred, and no clinical signs indicating systemic toxicity were noted throughout the study period. During the course of the study, significant higher body-weights were observed in males of the low dose group (0.2%), and significant lower body weights were observed in males fed 1 and 2% test substance in diet. However, no effects on food consumption and food efficiency were reported. Haematological examination and urinalysis revealed no adverse effects, and no treatment related changes were observed at gross necropsy and microscopical examination. The determination of organ weights revealed no test material-associated effects. Based on the results of this study, a NOAEL value of ≥ 1200 mg/kg bw was determined.

Supporting data are available for Ethyl 4-hydroxybenzoate (CAS 120-47-8) (Matthews et al., 1956). The study was conducted before appropriate test guidelines were developed. The test item was administered to groups of 12 Wistar rats per sex per dose via diet at concentrations of 2 and 8% (corresponding to 900-1200 and 5500-5900 mg/kg bw/day) for 12 consecutive weeks. Concurrent control animals received plain diet. Clinical signs, food consumption, compound intake and body weight development were recorded. At study termination, all animals were submitted to pathological examination, and particular attention was paid to kidneys, liver, heart, lung, spleen and pancreas. Animals administered 8% test substance in diet showed signs of toxicity characterised by depression, decreased motor activity and deaths within the first few weeks of the study. Body weight gain of these animals was decreased as compared to control animals receiving plain diet. Hereby, females were observed to be less sensitive than males; and no such effects were observe on animals dosed with 2% test item in diet. At gross necropsy and microscopical examination, no treatment related effects were noted. Based on the results of this study, a NOAEL value of 900-1200 mg/kg bw was determined.

Further supporting data are available for Butyl 4-hydroxybenzoate (CAS 94-26-8) (Matthews et al., 1956). The study was conducted before appropriate test guidelines and GLP were developed. The test item was administered to groups of 12 Wistar rats per sex per dose via diet at concentrations of 2 and 8% (corresponding to 900-1200 and 5500-5900 mg/kg bw/day) for 12 consecutive weeks. Concurrent control animals received plain diet. Clinical signs, food consumption, compound intake and body weight development were recorded. At study termination, all animals were submitted to pathological examination, and particular attention was paid to kidneys, liver, heart, lung, spleen and pancreas. Animals administered 8% test substance in diet showed signs of toxicity characterised by depression, decreased motor activity and deaths within the first few weeks of the study. In fact, all males dosed with 8% test substance in diet died before the end of the study period. Females were observed to be less sensitive than males, however, the group of females dosed with 8% test substance also had many early deaths and depression. Body weight gain of animals administered 8% test substance in diet was decreased as compared to control animals receiving plain diet. The effect was more apparent in males than in females. No such effects as described above were observed on animals dosed with 2% test item in diet. At gross necropsy and microscopical examination, no treatment related effects were noted. Based on the results of this study, a NOAEL value of 900-1200 mg/kg bw was determined.

Additional data on subchronic oral toxicity on non-rodents are available for Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), respectively (Matthews et al., 1956). The studies were conducted before appropriate test guidelines and GLP were developed. Young mongrel pups were administered capsules containing 500 and 1000 mg/kg bw of Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), respectively, for up to 422 days. Concurrent control animals remained untreated. Animals were examined for clinical signs, body weight, and changes in blood and urine parameters. Approximately one month before the end of the study all animals receiving 1000 mg/kg bw test material were studied for possible cumulation of the compound in the blood. Just prior to regular daily dosing a control blood sample was taken an analysed for the test substance and its metabolic byproducts. After the usual dose had been administered, blood samples were collected over the next 24 h and analysed for the test substance and its metabolic byproducts. In addition, urine was quantitatively collected over the same period of time and was analysed the same way. At termination of the study the dogs were sacrificed and pathological and histopathological examinations were performed. One of the females receiving 500 mg/kg bw/day of Methyl 4-hydroxybenzoate (CAS 99-76-3) was mated toward the end of the experiment. The findings for both, Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), were entirely the same. No treatment-related mortalities occurred and no toxic symptoms were observed in any of the animals throughout the study period. At the end of the study all animals appeared to be in excellent condition as far as general appearance was concerned. The body weights of the animals showed respectable increases. Since these animals were of varying antecedents it was expected that the final weights would vary greatly. All findings in haematology, urinalysis, and gross and microscopical examinations were entirely normal. At zero time (24 h following the previous treatment) a small amount of assayable test compound was still present in the blood, chiefly as conjugated end products. However, the plasma concentration-time curve resembled closely that of dogs not previously treated. 96-100% of the daily administered dose of the test item was accounted for in the urine samples, chiefly in the form of conjugated end products. Both the results from the urine and plasma analyses therefore furnish strong evidence of the lack of cumulation of the test item when orally administered at a dose of 1000 mg/kg bw/day for 1 year. The mated female dosed wit 500 mg/kg bw/day Methyl 4-hydroxybenzoate (CAS 99-76-3) delivered a litter of healthy pups, which were later raised in the authors' laboratory. Based on the results of these studies, NOAEL values of ≥ 1000 mg/kg bw /day each were determined for Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), respectively.

 

Chronic

Studies were conducted to assess the chronic oral toxicity of Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), respectively (Referenz). The studies were conducted before appropriate test guidelines and GLP were developed. The test item was administered to groups of 12 Wistar rats per sex per dose via diet at concentrations of 2 and 8% (corresponding to 900-1200 and 5500-5900 mg/kg bw/day) for 96 consecutive weeks. Concurrent control animals received plain diet. Clinical signs, food consumption, compound intake and body weight development were recorded. At study termination, all animals were submitted to pathological examination, and particular attention was paid to kidneys, liver, heart, lung, spleen and pancreas. The findings for both, Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), were entirely the same. No fatalities occurred and no clinical signs were observed throughout the study period. Food intake and hence drug intake remained fairly constant throughout the study period. Body weight gain of animals administered 8% test substance in diet was decreased as compared to control animals receiving plain diet. The effect was more striking at the beginning of the study and disappeared with progressing study duration. Moreover, this effect was more apparent in males than in females. No such effect was observed for the animals receiving 2% test substance in diet. No test material related changes were noted at pathological evaluation. Based on the results of these studies, NOAEL values of ≥ 5500-5900 mg/kg bw /day each were determined for Methyl 4-hydroxybenzoate (CAS 99-76-3) and Propyl 4-hydroxybenzoate (CAS 94-13-3), respectively.

 

Inhalation

No data on repeated dose toxicity via the inhalation route are available.

 

Dermal

No data on repeated dose toxicity via the dermal route are available.

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Hazard assessment is conducted by means of read-across from structural analogues/surrogates. The selected study is the most adequate and reliable study based on the identified similarities in structure and intrinsic properties between source and target substances and overall assessment of quality, duration and dose descriptor level (refer to the endpoint discussion for further details).

Justification for classification or non-classification

Based on read-across from structurally similar substances, the available data on oral repeated dose toxicity do not meet the classification criteria according to Regulation (EC) No 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.