DL-malic acid

Repeated dose toxicity: oral

Administrative data

Endpoint:

chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not stated

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Peer-reviewed study comparable to OECD guideline 452.

Cross-reference

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1947

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 452 (Chronic Toxicity Studies)

Principles of method if other than guideline:

Overview of chronic exposure studies completed prior to introduction of experimental test guidelines. The study design was similar in principle to the method subsequently published as OECD 452

GLP compliance:

no

Remarks:

Study pre-dates establishment of GLP guidelines

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Fumaric acid

EC Number:

203-743-0

EC Name:

Fumaric acid

Cas Number:

110-17-8

IUPAC Name:

but-2-enedioic acid

Details on test material:

No details provided

Test animals

Species:

rat

Strain:

Osborne-Mendel

Sex:

male

Details on test animals or test system and environmental conditions:

Male weanling rats (21-days) were fed diets containing fumaric acid for two years. The basic diet consisted of ground commercial rat biscuits with 1% added cod-liver oil. Fumaric acid was mixed with the basic diet using a rotary batch mixer. All animals were kept in individual cages in a room with temperature and humidity controlled for the duration of the experiment. Animals were given free access to their respective diets and water.

Administration / exposure

Route of administration:

oral: feed

Vehicle:

other: cod liver oil added to moisten diet

Details on oral exposure:

The basic diet consisted of ground commercial rat biscuits with 1% added cod-liver oil. Fumaric acid was mixed with the basic diet using a rotary batch mixer. Animals were given free access to their respective diets.

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No data

Duration of treatment / exposure:

2 years

Frequency of treatment:

Daily in feed

Doses / concentrations

Remarks:

Doses / Concentrations:
0.5, 1.0, and 1.5%
Basis:
nominal in diet

No. of animals per sex per dose:

7- 12 male rats per dose, 10-12 females per group

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: In a previous experiment, male and female rats were fed diets containing 0.1, 0.5, 0.8 and 1.2% fumaric acid for two years.
This study was initiated to examine the toxicity of fumaric acid more closely.

Positive control:

No data

Examinations

Observations and examinations performed and frequency:

Individual animal weights and food consumption were determined at weekly intervals.

Sacrifice and pathology:

Gross pathology and histopathology of lung, heart, liver, spleen, pancrease, stomach, small intestine, kidney, adrenal and testis.
Additional structures examined by histopathology include colon, bone marrow, leg bones, leg muscles, lymph nodes, uterus, ovary, thyroid and parathyroid.

Other examinations:

No details

Statistics:

No details

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Mortality in the high dose group was significant but low mortality rates evident in lower dose groups

Mortality:

mortality observed, treatment-related

Description (incidence):

Mortality in the high dose group was significant but low mortality rates evident in lower dose groups

Body weight and weight changes:

no effects observed

Description (incidence and severity):

weight gains recorded for males and females at each dose level showed no treatment related effects

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Food consumption over te first 26 weeks of the study andsecond 26 weeks were compared or each treatment group. No statistically significant changes observed.

Food efficiency:

not specified

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Sporadic findings reported but no treatment relationship established

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

At two years, there were only 2 animals living on 1.5% fumaric acid.; other dose levels had no effect on mortality rate. Animals fed 1.5% fumaric acid showed more atrophy of the testis and 2 rats fed 0.5% and 1.0% fumaric acid showed phlegmonous gastritis.

Effect levels

Dose descriptor:

NOAEL

Effect level:

ca. 600 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

1.2% test material inclusion level

Sex:

male

Basis for effect level:

other: Low incidence of mortality seen at a dose level of 1.5%

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Mean gain in weight of rats fed fumaric acid for a year (data from a previous experiment is also included)

Test material	Dosage (%)	No. of animals	Sex	Mean gain in weight (g)	Standard error of mean (+ g)
Fumaric acid	0.1	11	M	452.9	19.0
	0.1	10	F	288.0	10.0
	0.5	10	M	444.4	20.7
	0.5	12	F	269.7	9.6
	0.8	9	M	429.6	11.0
	0.8	12	F	265.2	5.7
	1.0	10	M	468.8	25.9
	1.2	11	M	466.9	14.6
	1.2	12	F	280.0	10.2
	1.5	7	M	459.9	23.2
Control		31	M	464.9	13.4
		22	F	274.4	9.8

Applicant's summary and conclusion

Conclusions:

Signs of toxic effects were observed when rats were fed fumaric acid at high dietary concentrations - 1.5%.
Toxic effects occurred in rats fed diets containing 1.5% fumaric acid for two years. An increase in mortality rate and more atrophy of the testis were seen in rats fed 1.5% fumaric acid. Inanition seems at least partly responsible for atrophy of the testis.
No adverse effects on reproductive organs were reported in a previous study using female rats administered up to 1.2% fumaric acid in the diet for 2 years.
The NOAEL was 1.2% or circa 600 mg/kg bw/day.
A previous study conducted in a similar manner with female rats showed no adverse effects on reproductive organs after administration of up to 1.2% fumaric acid in the diet for 2 years. Based on the low incidence of mortality of male rats, 1.2% is very near a NOAEL for chronic exposure to fumaric acid (600 mg/kg bw). The 1.2% NOAEL (600 mg/kg bw/day) derived from the available long term rat toxicity data was confirmed as the appropriate point of departure by two reviews - The International Programme on Chemical Safety (WHO 1975 Food Additive Series 6) report and the findings ofthe European Commission DG C Report of the Scientific Committee on animal nutrition on the safety of fumaric acid (SCAN, 2003).
In the SCAN report, a safe use was established in piglets in the field at circa 1000 mg/kg bw/day and circa 2000 mg/kg bw/day in experimental data. For human psoriatic patients the safe use was 0.9g fumaric acid equivalent/d or circa 15 mg/kg bw/day.
The WHO report contains confirmation of the study in which groups of rats were dosed at 1.2% fumaric acid without adverse toxicological effects.

Executive summary:

In a two-year dietary study using male rats, a very slight increase in mortality rate and some testicular atrophy was observed after administration of 1.5% fumaric acid (approximately 750 mg/kg bw/day). Gross and microscopic examination of major organs revealed no abnormalities. The authors of this study concluded that inanition was partly responsible for testicular atrophy. A previous study conducted in a similar manner with female rats showed no adverse effects on reproductive organs after administration of up to 1.2% fumaric acid in the diet for 2 years. Based on the low incidence of mortality of male rats, 1.2% is very near a NOAEL for chronic exposure to fumaric acid (600 mg/kg bw). The 1.2% NOAEL (600 mg/kg bw/day) derived from the available long term rat toxicity data was confirmed as the appropriate point of departure by two reviews - The International Programme on Chemical Safety (WHO 1975 Food Additive Series 6) report and the findings of the European Commission DG C Report of the Scientific Committee on animal nutrition on the safety of fumaric acid.