Ammonium 2-amino-4-(hydroxymethylphosphinyl)butyrate

Repeated dose toxicity: dermal

Administrative data

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Oct 1983 - May 1985

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1985

Report date:

1985

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)

Version / remarks:

May 1981

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPP 82-2 (Repeated Dose Dermal Toxicity -21/28 Days)

Version / remarks:

Nov 1982

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Ammonium 2-amino-4-(hydroxymethylphosphinyl)butyrate

EC Number:

278-636-5

EC Name:

Ammonium 2-amino-4-(hydroxymethylphosphinyl)butyrate

Cas Number:

77182-82-2

Molecular formula:

C5H12NO4P.H3N

IUPAC Name:

ammonium 2-amino-4-[hydroxy(methyl)phosphoryl]butanoate

Test animals

Species:

rat

Strain:

Wistar

Remarks:

Hoe: WISKf (SPF71)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Hoechst AG, Pharma Forschung Toxikologie, Kastengrund, Germany
- Age at study initiation: about 8-10 weeks
- Weight at study initiation: males: 143-154 g; females: 144-154 g
- Housing: single
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 8/9 days

DETAILS OF FOOD AND WATER QUALITY:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): 55 +/- 10
- Air changes (per hr): 16
- Photoperiod (hrs dark / hrs light): 12/12 (illumination: 7 am to 7 pm)

IN-LIFE DATES: From: 17.10.1983 To: 08/09.12.1983

Administration / exposure

Type of coverage:

occlusive

Vehicle:

other: physiological saline

Details on exposure:

TEST SITE
- Area of exposure: nape (shaved)
- % coverage: 10
- Type of wrap if used: bandage (aluminium foil, Fixomull-Strech adhesive plaster)
- Time intervals for shavings or clipplings: at least once weekly

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes, washed
- Time after start of exposure: Exposure for 6 h. After removal of bandage, treated skin was washed.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1.3 ml/kg bw
- Concentration (if solution): 7.5, 22.5, 75.0% solution (w/v)
- Constant volume or concentration used: yes
- For solids, paste formed: no, solution in deionized water

VEHICLE
- Justification for use and choice of vehicle (if other than water): deionized water

USE OF RESTRAINERS FOR PREVENTING INGESTION: not specified

Duration of treatment / exposure:

30 days (21 applications)

Frequency of treatment:

Monday-Friday, 6 h/d

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

100 mg/kg bw/day

Doses / concentrations 2

Dose / conc.:

300 mg/kg bw/day

Doses / concentrations 3

Dose / conc.:

1 000 mg/kg bw/day

No. of animals per sex per dose:

6 (5 for recovery groups; control and high dose)

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
- Rationale for animal assignment (if not random): random
- Fasting period before blood sampling for clinical biochemistry:
- Rationale for selecting satellite groups:
- Post-exposure recovery period in satellite groups: 14 days
- Section schedule rationale (if not random):
- Other:

Positive control:

not included

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: Twice weekly

FOOD CONSUMPTION: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Time schedule: Twice weekly

WATER CONSUMPTION: Yes
- Time schedule for examinations: Once weekly

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the end of the study
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: All animals
- Parameters checked: hemoglobin, erythrocytes, leucocytes, hematocrit, reticulocytes, differential blood count*, thrombocytes, coagulation time, erythrocyte sedimentation rate, thromboplastin time, activated partial thromboplastin time, methemoglobin*. MCV, MCH, and MCHC were also calculated. (* no evaluation for 100 and 300 mg/kg bw/day groups, since the findings for 1000 mg/kg bw/day were normal)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At the end of the study
- Animals fasted: No
- How many animals: All animals
- Parameters checked: sodium, potassium, inorg. phosphorus, uric acid, total bilirubin, direct bilirubin, creatinine, serum glucose, urea nitrogen, calcium, chloride, SGOT, SGPT, AP, LDH, cholesterol, total lipids, total protein, serum electrophoresis
- In view of the results at the end of the treatment period, the only parameters examined at the end of the recovery period were chloride and uric acid in the males and sodium, potassium and chloride in the females.

URINALYSIS: Yes
- Time schedule for collection of urine: At the end of the study (In the night from Day 28 to 29)
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked: Appearance, color, protein, glucose, hemoglobin, bilirubin, pH, sediment. No urinalysis was carried out at the end of the recovery period, since there were no substance-related changes

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
- Autopsy included skin, orifices, eyes, teeth, oral mucosa, and internal organs
- Organs weighed: heart, lungs, liver, kidneys, spleen, brain, testes (without epididymes)/ovaries, adrenals, pituitary, thyroid

HISTOPATHOLOGY: Yes
heart, urinary, bladder, pancreas, lungs, testes, adrenals, liver, epididymides, thymus, kidneys, prostate, pituitary, spleen, seminal vesicles, brain, stomach, ovaries, eye with optic nerve, jejunum, uterus, bone marrow (femoral), colon, thyroid, treated skin areas, untreated skin areas

Statistics:

Dunnett's test, Sidak test, Nemenyi/Dunnett, Nemenyi/Sidak

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

- 1000 mg/kg bw/day: 4 males and 2 females showed piloerection. One male animal refused its food almost entirely from the beginning of the
second week of treatment and had to be removed from the study in a cachectic condition on day 16; encrusted blood was also observed around the eyes and the mouth/nose of this animal.
- 300 mg/kg bw/day: one male animal showed notably aggressive behaviour, squatting position, pilo-erection and convulsive jumping and rolling spasms at the end of the treatment period.

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

On days 5 and 6 of the study, slight signs of irritation appeared on the treated skin areas of the females. During the further course of the study no
signs of dermal irritation were observed in either the females or the males.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

- 1000 mg/kg bw/day: One male was humanely killed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

- 300 mg/kg bw/day: decreased APTT in males, increased APTT in females. Decreased leucocytes in males

Since these changes were not dose-related and occurred only to a minimal extent and without further toxicological correlates, they must be considered as non-substance-related and of no toxicological relevance.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

- 1000 mg/kg bw/day: In males, a slight increase in uric acid and potassium; Decreases in sodium in females and chloride in both sexes
- 300 mg/kg bw/day: In males, a slight increase in uric acid and alpha3 globulin; decrease in sodium in females
- 100 mg/kg bw/day: Increase in gamma1 globulin in females

Effects cannot be considered as toxicologically relevant (independent of dose level and only to a minimum extent). Uric acid levels were still within range of normal biological variation and were thus considered not toxicologically relevant.

Urinalysis findings:

no effects observed

Behaviour (functional findings):

effects observed, treatment-related

Description (incidence and severity):

- 1000 mg/kg bw/day: 4 males and 2 females showed either timid or aggressive behavior, increased motor excitation (especially followign tactile stimuli), or squatting position
- 300 mg/kg bw/day: one male showed notably aggressive behavior, squatting position, convulsive jumping, and rolling spasms

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Effect levels

Dose descriptor:

NOEL

Effect level:

100 mg/kg bw/day

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

behaviour (functional findings)

clinical signs

Remarks on result:

other: NOEL comparable to NOAEL

Target system / organ toxicity

Critical effects observed:

yes

Lowest effective dose / conc.:

300 mg/kg bw/day

System:

central nervous system

Organ:

brain

Treatment related:

yes

Dose response relationship:

yes

Any other information on results incl. tables

Table 1: Clinical findings

Dose group (mg/kg bw/day)	Sex	Day of treatment	Symptoms
control	Male	1-30	No symptoms
	Female	1-30	No symptoms*
100	Male	1-30	No symptoms
	Female	1-30	No symptoms
300	Male
	Animal No 19	10-15 10-19	aggressive behavior easily startled
	Animal No 23	24-30 27-30	Aggressive behavior, ‘kangaroo’ position, piloerection Convulsive jumpings
	Female	1-30	No symptoms
1000	Male
	Animal No 24	7-16 8-16 13-16	Right upper rodent tooth broken, ‘kangaroo’ position, crusted eyes, blood crusted nose Narrowed eye opening Piloerection, hyporeflexia, decreased respiratory rate, abdominal position
	Animal No 26	16-23 20-23 23	Aggressive behavior Easily startled piloerection
	Animal No 27	10-15 10-26	Aggressive behavior Easily startled
	Animal No 28	20-26	Lesion on the left foreleg
	Animal No 30	7-12 7-26	Aggressive behavior Hyperactivity following tactile stimulus
	Animal No 33	10-16 10-22	Aggressive behavior Easily startled
	Female
	Animal No 64	6-15	Hyperactivity following tactile stimulus
	Animal No 68	12-19 12-32	‘kangaroo’ position piloerection

* Four females showed lesions; however, they were due to removing the occlusive bandage by test animal