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Administrative data

Description of key information

No valid repeated dose toxicity studies on salicylic acid are available. A read-across approach is therefore proposed from studies on Methyl salicylate (MeS) which is readily metabolised to salicylic acid (See section.7.1.1).

A set of studies was conducted on MeS by Webb & Hansen (1963), consisting of oral feeding studies of duration 17 weeks and 2 years in rats, studies in dogs by capsule administration of duration 59 days and 2 years, and a dermal study in rabbits.  These studies did not examine all the parameters recommended in the current OECD guidelines 409 and 452, however they were conducted according to good scientific principles by US FDA.  The chronic studies in rat and dog are therefore proposed as key studies for this endpoint, with the subchronic studies in rats and dogs as supporting study. A second set of subacute oral studies was conducted on MeS by Abbott & Harrisson (1978) to further investigate the effects on bone in rats and liver in dogs, identified in the studies by Webb & Hansen.  These are considered to be acceptable as supporting studies.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
chronic toxicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
no data are available
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: This is an old study predating GLP however the protocol and results were reported in adequate detail and included hematological studies, gross pathology, and limited histopathological examinations of key organs and tissues.
Reason / purpose:
reference to same study
Qualifier:
no guideline available
Principles of method if other than guideline:
MeS was blended with diet and adminstered daily for a period of 2 years.
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Osborne-Mendel
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: no data
- Age at study initiation: weanling
- Weight at study initiation: ~ 50 g
- Fasting period before study: no data
- Housing: no data
- Diet: ad libitum
- Water: no data
- Acclimation period: no data


ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod: no data


IN-LIFE DATES: no data
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:


DIET PREPARATION
- Rate of preparation of diet (frequency): every other week
- Mixing appropriate amounts with (Type of food): no data
- Storage temperature of food: all diets were stored in sealed cans under refrigeration
- Other: 10% additional methyl salicylate was added at time of mixing to compensate for evaporation

VEHICLE: none
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
none
Duration of treatment / exposure:
2 years
Frequency of treatment:
daily
Remarks:
Doses / Concentrations:
0, 0.1, 0.5, 1 and 2% (0, 50, 250, 500, and 1000 mg/kg bw/day)
Basis:
nominal in diet
No. of animals per sex per dose:
25/sex/dose (except for 24 males, 26 females in 2% group)
Control animals:
yes
Details on study design:
- Dose selection rationale: the data reported in the subchronic study in rats by the same authors (methyl salicylate/ dietary administration for 17 weeks in rats).

Positive control:
no data are available
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: No


DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: The rats were weighed weekly.


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No


FOOD EFFICIENCY: No



OPHTHALMOSCOPIC EXAMINATION: No


HAEMATOLOGY: Yes
- Time schedule for collection of blood: 3, 11, 17 and 22 months.
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes
- How many animals: 10 rats/dose



CLINICAL CHEMISTRY: No


URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Other examinations:
none
Statistics:
not reported
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
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:
CLINICAL SIGNS AND MORTALITY
Animals of the 1.0% and 2.0% groups developed rough hair coats. In the high-dose group, half of the animals died by week 8 and all of the animals
died by week 49 of the study.
BODY WEIGHT AND WEIGHT GAIN:
Animals of the 1.0% and 2.0% groups had statistically significant growth inhibition.
HAEMATOLOGY
no hematological effects were observed.ORGAN WEIGHTSAverage organ weights were similar for all animals. however, relative organ to body weight
ratios for the testes of male animals and for the heart and kidneys of the female animals of the 1.0% groups were significantly increased.
GROSS PATHOLOGY
gross lesions of the pituitary gland were observed in 10 animals of the 0.5% group as compared to four animals in the control
group.In the 2% group, 29 of the 50 animals had pneumonia, which appeared to be more acute than regularly observed.
HISTOPATHOLOGY: NON-NEOPLASTIC
Microscopically, the only induced lesion was a pronounced change in the bones of the rats on the 2.0% diet.
Cancellous bone in the metaphysis was increased as compared to same-age controls; this was observed to a moderate degree in five and a marked
degree in four of the nine bones examined from animals of the 2.0% group. Bone lesions were slight in 2 of 11 and 1 of 11 bones examined from
animals of the 1.0% and 0.5% groups, respectively. The affected bones had fewer osteoclasts, and the number was inversely proportional to the
degree of change.
HISTOPATHOLOGY: NEOPLASTIC
The data are limited:Malignant pituitary tumors occurred in 1 male and 2 female rats on the 0.5% diet. Mammary tumors occurred in females rats on
all diets.
Dose descriptor:
NOAEL
Effect level:
50 mg/kg diet
Sex:
male/female
Basis for effect level:
other: Increased amount of cancellous bone from 250 mg/kg
Critical effects observed:
not specified

The results for a supplemental study:

One male test animal died on day 11, the two others died on day 19. The females died on days 31, 40 and 71. Rough hair coat and growth inhibition was observed for all test animals, with some animals having labored respiration.

Grossly, four of the six treated rats had slight to moderate lung damage. Focal gastric hemorrhages were present in the glandular portion of three of the test animals. Bone growth was affected.

Long bones of the limbs showed reduced length and diameter compared to controls. Density of the hypophysis of long bones was increased.

Conclusions:
Under the test conditions, growth retardation and bone lesions were reported in rats at levels of 1.0 and 2.0% MeS in the diet, with minimal effects at 0.5%. Based on this study and the subchronic oral study, the NOAEL /oral/rat is 0.1% (50 mg/kg body weight/day, equivalent to 45.4 mg/kg bw/day as SA).
Executive summary:

Webb and Hansen (1963) administered methyl salicylate in the diet to groups of 24 -25 male and 25 -26 female Osborne-Mendel rats at dietary concentrations of 0, 0.1%, 0.5%, 1.0% or 2.0% in the diet providing doses of approximately 0, 50, 250, 500, and 1000 mg/kg body weight/day for two years. All rats in the 1000 mg/kg group died by the 49th week. Body weight of both sexes were significantly decreased in both the 500 and 1000 mg/kg group body weight/day groups.

An increased amount of cancellous bone was present in the metaphyses in rats treated at either 500 or 1000 mg/kg body weight/day, with a more marked effect at the highest dose level. A slight increase was reported in one rat at 250 mg/kg. The relative testes weights of males were significantly increased as were the relative weights of the heart and kidneys of females in the 500 mg/kg body weight/day group. Gross pituitary gland lesions were found in 10 rats at 250mg/kg bw/day compared to four animals in the control groups. while based on chronic and subchronic oral studies in rat by Webb and Hansen (1963), the NOAEL value is 50 mg/kg body weight/day (equivalent to 45.4 mg/kg bw/day as SA).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
45.4 mg/kg bw/day
Study duration:
chronic
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
no data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Single concentration tested. No data on substance purity.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
GLP compliance:
no
Species:
rat
Strain:
other: Alderley Park
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Alderley Park
- Age at study initiation: no data
- Weight at study initiation: mean 200g
- Fasting period before study: no data
- Housing: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: no data
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Remarks on MMAD:
MMAD / GSD: Not applicable, vapour
Duration of treatment / exposure:
4 weeks
Frequency of treatment:
7 hours per day, 5 days per week
Remarks:
Doses / Concentrations:
635 mg/m3
Basis:
nominal conc.
No. of animals per sex per dose:
4
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: Daily

FOOD CONSUMPTION: - No data

FOOD EFFICIENCY:- No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: No data
- Time schedule for collection of blood: No data
- Animals fasted: No data
- How many animals: No data

CLINICAL CHEMISTRY: No data

URINALYSIS: Yes
- Time schedule for collection of urine: Overnight after last exposure
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data

NEUROBEHAVIOURAL EXAMINATION: No data
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
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:
NOEC
Effect level:
700 mg/m³ air
Sex:
female
Basis for effect level:
other: overall effects no adverse effects
Critical effects observed:
not specified

No toxic signs and no macroscopic or microscopic effects were seen at necropsy following exposure to a saturated atmosphere of methyl salicylate for 7 hours per day, 5 days per week for 4 weeks.

Conclusions:
Methyl salicylate showed no evidence of toxicity by inhalation when tested at an effectively saturated concentration for 7 hours per day for four weeks. Although no conclusion can be drawn for potential local effects of SA dust inhalation, it can be predicted that systemic toxicity would be low.
Executive summary:

4 female Alderley Park rats weighing an average of 200 g were exposed to a dynamic atmosphere (atmosphere continuously generated and passed through the exposure chamber) containing a saturated atmosphere (700 mg/m3) of methyl salicylate for 7 hours per day, 5 days per week for 4 weeks. Food and water were available ad libitum. Animals were weighed each day, and their condition and behaviour were recorded throughout the exposure period. Urine was collected overnight after the last exposure day for biochemical testing. Gross necropsy was conducted as well as microscopic examination of organs. Methyl salicylate at 700 mg/m3 (120 ppm), 635 mg/m3 as SA, the maximum achievable atmosphere, did not cause any adverse effects (Gage, 1970). Although this study does not allow any conclusion to be drawn for the possibility of local effects from inhalation of SA dust, it can be predicted that systemic toxicity from SA inhalation would be low.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
635 mg/m³
Study duration:
subacute
Species:
rat

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

No valid repeated dose toxicity studies on salicylic acid are available. A read-across approach is therefore proposed from studies on Methyl salicylate (MeS) which is readily metabolised to salicylic acid.

Justification for Read-across from Methyl salicylate:

The oral absorption, distribution and metabolism of MeS, NaS and ASA have been compared in rats, dogs and humans (Davison, 1961). In rats and dogs, MeS, NaS and ASA were all rapidly absorbed following oral administration even at high concentrations. Absorption of MeS in humans was somewhat slower than for ASA, with total salicylate plasma concentration at 90 minutes approximately half that from ASA. Salicylic acid has also been shown to be rapidly absorbed after oral administration in rats (Rainsford at al., 1980).

Plasma analysis in rats showed rapid hydrolysis to free salicylate for MeS, NaS and ASA, resulting in comparable plasma concentrations of salicylate at 60 minutes post dosing, with no measurable parent compound. In humans, hydrolysis of MeS was slower and less complete, with 30% MeS remaining unhydrolysed at 15 minutes, and 21% at 90 minutes (Davison, 1961).

These results indicate that following absorption, the initial metabolic step for all these salicylates (MeS, NaS and ASA) is hydrolysis to free salicylate. Since free salicylate is the principal species circulating in plasma following absorption of MeS and SA, it follows that data from methyl salicylates are acceptable for read across to SA for all systemic toxicological endpoints.

(See also section.7.1.1)

MeS-Subchronic toxicity studies

1- Oral studies:

Subchronic toxicity oral studies have been conducted on MeS: two in rats and two in dogs. In the 17 -week study in Osborne-Mendel rats reported by Webb and Hansen, 1963, a NOAEL of 0.1% in the diet, equivalent to ~ 50 mg/kg bw/day, was identified. Bone lesions and growth retardation were observed in rats fed MeS at 1% and 2% in the diet. The results of 6 to 12 week experiments in SD rats reported by Abbott and Harrisson (1978), suggest a NOAEL of 0.3% in the diet (180 mg/kg/bw/day) based on reduced bodyweight at 0.63%. In dogs, administered MeS orally in capsules, a NOAEL of 50 mg/kg bw/day was identified from a study of duration 59 days (Webb & Hansen, 1963) and a NOAEL of 170 mg/kg bw/day was reported from studies of duration approximately 6 months (Abbott and Harrisson;1978). Liver enlargement and growth retardation were reported in dogs given capsules with 150 and 350 mg/kg/day of MeS. Although these studies were limited in endpoints evaluated, they were well conducted and reported (reliability: 2).

Given these results, a systemic NOAEL of 50 mg/kg bw/day MeS can be identified, equivalent to 45.4 mg/kg bw/day salicylic acid

2) Dermal

Webb and Hansen (1963), applied MeS dermally to groups of three rabbits of mixed sex at 0.5, 1.0, 2.0 and 5.0 mL/kg (590, 1180, 2360, and 4720 mg/kg) body weight/day, 5 days per week for up to 96 days. All 3 high-dose animals had died by day 28 of exposure, following weight loss and depressed activity. In one of these animals, local effects on the skin and lesions in a number of organs were reported. The only effect reported in the other high dose animal examined was a suggestion of an effect on the distal portion of nephrons. At 2.0 ml/kg (2360 mg/kg), slight sloughing of epidermal scales was observed in 2/3 rabbits. No dermal effects were noted in rabbits exposed to 0.5 or 1.0 ml/kg (590 and 1180 mg/kg body weight/day). The NOAEL for local effects was 1.0 ml/kg/day (1180 mg/kg bw/day). This value cannot be read across to SA however, since the lower pH of the acid than the ester predicts a higher potential for irritation for SA on repeated administration, even thougyh neither substance is classified as irritant on single administration. No clear NOAEL was determined for systemic effects due to the reported increased incidence in spontaneous nephritis and mild hepatitis over that in rabbits in other experiments. This dermal study at very high doses was limited in design and in reporting detail and is not useful for risk assessment for systemic toxicity.. It is not possible to determine whether the effects reported on kidney and other organs in one high dose animal represent systemic toxicity or a secondary effect due to skin and muscle damage.

3) Inhalation (sub-acute study)

4 femalerats were exposed to a saturated atmosphere (700 mg/m3) of MeS for 7 hours per day, 5 days per week for 4 weeks. Animals were weighed each day, and their condition and behaviour were recorded throughout the exposure period. Urine was collected overnight after the last exposure day for biochemical testing. Gross necropsy and microscopic examination of organs was carried out. MeS at 700 mg/m3 (120 ppm), equivalent to 635 mg/m3 as salicylic acid, the maximum achievable atmosphere, did not cause any adverse effects (Gage, 1970). This study does not allow any conclusion to be drawn on potential for local effects of SA on the respiratory system, but provides limited data suggesting the SA would not cause significant systemic toxicity.

MeS-Chronic toxicity studies

Chronic toxicity studies have been conducted on MeS in rats and in dogs for 2 years (Webb and Hansen, 1963). Although the studies are relatively old and are limited in the endpoints evaluated, the protocol and results were reported in adequate detail and included hematological studies (reliability: 2)

Webb and Hansen (1963) administered methyl salicylate in rats at dietary concentrations of 0, 0.1%, 0.5%, 1.0% or 2.0% in the diet (equivalent to 0, 50, 250, 500, and 1000 mg/kg bw/day) for two years. Body weight of both sexes were significantly decreased in both the 500 and 1000 mg/kg group body weight/day groups and an increased amount of cancellous bone was present in the metaphyses in rats treated at either 500 or 1000 mg/kg body weight/day.

In dogs, the same authors administered MeS in capsule form at doses of 0, 50, 150, or 350 mg/kg body weight/day, 6 days/week for 2 years. One high dose animal died of hepatitis unrelated to MeS. Hematological analyses and necropsy examination were normal, except that dogs treated at 150 and 350 mg/kg body weight/day had enlarged livers with hepatocellular swelling. No other pathology was reported in any of the animals. Reduced body weight was reported in the 350 and 150 mg/kg body weight/day groups.

The 2 year oral toxicity data for MeS are consistent with the oral subchronic toxicity data from the same laboratory.

NOAEL value for MeS is 50 mg/kg bw/day in both rats and dogs, equivalent to 45.4 mg/kg bw/day as SA.

Conclusion:

Given these results, a systemic NOAEL of 45.4 mg/kg bw/day can be used for quantitative human health risk assessment.

NB:

Restrospective studies of children receiving salicylate therapy in the management of juvenile rheumatoid arthritis did not reveal any cases in which either bone lesions or hepatomegaly, as seen in rats and dogs, could be associated with massive daily doses of salicylate over prolonged periods of time. The reviews of human case histories (secondary reference, Abbott and Harrisson, 1978) suggest that the salicylate-related bone lesion in rats and hepatomegaly in dogs are not relevant for human risk assessment.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Reliability 2 study on several species, with protocol and results reported in adequate detail and including hematological studies, gross pathology, and limited histopathological examinations of key organs and tissues.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
Only study available on MeS, reliability 2

Repeated dose toxicity: via oral route - systemic effects (target organ) other: bone

Justification for classification or non-classification

The guidance values for classification of a substance as harmful on repeated exposure by the oral route according to the criteria of Annex VI Directive 67/748/EEC(R48/22) relate to severe effects reported at 50 mg/kg bw/day or below in a 90-day study. The equivalent guidance values for classification according to EU/GHS criteria (STOT Category 2) relate to effects reported at 10-100 mg/kg bw/day in a 90-day study.

For SA, the no-effect level in both 17-week and 2-year oral dietary studies read across from MeS (Webb & Hansen, 1963) was 45.4 mg/kg bw/day as SA (50 mg/kg bw/day as MeS). Slight effects on bone density were reported at the LOAEL of 500 mg/kg bw/day in the subchronic study, not seen in human juvenile arthritis terament with ASA. The limited data available on repeat dose toxicity by dermal or inhalation exposure do not indicate a significant hazard by either route.

SA is not classified for repeat-dose toxicity according to the criteria of Annex VI Directive 67/748/EECor UN/EU GHS.