Malic acid

Toxicity to reproduction

Administrative data

Endpoint:

two-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Experimental data published in peer reviewed journal

Data source

Reference

Reference Type:

review article or handbook

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)

Deviations:

yes

Remarks:

2 rather than 3 treatment groups investigated

GLP compliance:

no

Remarks:

Study pre-dates implementation of GLP regulations

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Malic acid

EC Number:

230-022-8

EC Name:

Malic acid

Cas Number:

6915-15-7

Molecular formula:

C4H6O5

IUPAC Name:

2-hydroxybutanedioic acid

Details on test material:

No data

Test animals

Species:

rat

Strain:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: No data
- Age at study initiation: (P) x approximately 3 wks; (F1) x 14 wks (at mating)
- Weight at study initiation: (P) Males: no data; Females: no data; (F1) Males: no data; Females: no data
- Fasting period before study: No data, assumed not applicable
- Housing: No data
- Diet (e.g. ad libitum): No data, assumed ad libitum
- Water (e.g. ad libitum): No data, assumed ad libiutum
- Acclimation period: No data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data
- Humidity (%): No data
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data

Administration / exposure

Route of administration:

oral: feed

Vehicle:

not specified

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): No data
- Mixing appropriate amounts with (Type of food): No data
- Storage temperature of food: No data

Details on mating procedure:

- M/F ratio per cage: No data
- Length of cohabitation: No data
- Proof of pregnancy: No data, referred to as day ? of pregnancy
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility. No data
- Further matings after two unsuccessful attempts: No data
- After successful mating each pregnant female was caged: No data

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

From 9 weeks prior to mating of P1 animals through to weaning of 2nd generation animals

Frequency of treatment:

Continuous (in feed)

Details on study schedule:

- F1 parental animals not mated until approximately 100 days after selected from the F1 litters.
- Selection of parents from F1 generation when pups were 21 days of age.
- Age at mating of the mated animals in the study: 14 weeks

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 1000 and 10000 ppm
Basis:
nominal in diet

No. of animals per sex per dose:

10 males / 20 females / group

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: No data
- Rationale for animal assignment (if not random): No data

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: No data

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

Oestrous cyclicity (parental animals):

No data

Sperm parameters (parental animals):

No data

Litter observations:

No data
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: Yes but no details
- If yes, maximum of 8 pups/litter; excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2] offspring: number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities

GROSS EXAMINATION OF DEAD PUPS: No data

Postmortem examinations (parental animals):

No data

Postmortem examinations (offspring):

No data
SACRIFICE
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed at 21 days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGTHS
No data

Statistics:

No data

Reproductive indices:

No details available

Offspring viability indices:

No details available

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weight gain of males slightly reduced prior to mating

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Body weight gain of males slightly reduced prior to mating

Organ weight findings including organ / body weight ratios:

not specified

Histopathological findings: non-neoplastic:

not specified

Other effects:

not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Details on results (P0)

Body weight gains of males of the P1 generation groups treated with malic acid were slightly decreased compared to control animals prior to mating. Body weights in females were comparable (with controls).
Feed consumption was similar for test and control animals.
Survival was similar for test and control animals.
Appearance and behaviour were similar for treated and control rats of the P1 generation.
None of the P1 animals died during the FIA or F1B phase.

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

10 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: overall effects

Results: F1 generation

General toxicity (F1)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Laboured respiration / wheezing during weaningk

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings:

no effects observed

Details on results (F1)

For all litters, the various indices, litter sizes, and pup body weights were comparable among test and control animals.
Necropsied pups from three of the low-dose F1A litters had rough surfaces on the spleen.
The number of pups that were weak or had laboured respiration during lactation was increased in the high-dose group F2A litters. No abnormal findings were reported at necropsy.

The P2 test and control animals were similar throughout the study. Wheezing was observed in all groups during the F2B phase.
At necropsy of the F2A litters, renal discoloration was noted in two animals, dark renal medullas were noted in four animals, rough surfaces on the spleen were noted in four animals and white foci on the spleen were seen in three weanlings of the low-dose group. Renal discoloration (three animals), dark red corticomedullary zones (three animals), dark renal medullas (three animals), rough surfaces on the spleen (two animals), and a firm, enlarged, irregularly-shaped caecum with a hole penetrating it (one animal) were observed in high-dose weanling animals.
In the F2B litters, weakness and laboured respiration were observed in a few low-dose pups, and the renal pelvis of one high-dose pup was dilated at necropsy. The animals of the F2B generation delivered by caesarean section exhibited no meaningful differences between test and control animals in the number and placement of implantation and resorption sites or in the number, weight, or length of live neonates, and none of the neonates died.
The skeletal development of the F2B neonates was similar between test and control animals.
Those slight differences in developmental indices that were observed were considered to be within the range of normal variations in foetal development.

Results: F2 generation

Effect levels (F2)

Dose descriptor:

LOAEL

Generation:

F2

Effect level:

10 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: clinical signs

Overall reproductive toxicity

Reproductive effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Administration of malic acid over 2 generations had no clear effect on fertility or development. The LOAEL from the study may be regarded as 10000 ppm.

Executive summary:

Reproductive toxicity has been investigated with a 2-generation study using methods similar to those described in OECD TG 416. Administration of malic acid over 2 generations had no clear effect on fertility or development. The LOAEL from the study may be regarded as 10000 ppm.