Disodium (Z)-4-(9-octadecenylamino)-4-oxo-2(or 3)-sulphonatobutyrate

Repeated dose toxicity: oral

Administrative data

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:

1976

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The study was not performed under GLP, although the study was conducted according to standards of that time.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1976

Report date:

1976

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Principles of method if other than guideline:

The age of the animals is very young by which theymay be more sensitive. In addition, the validity of the study is uncertain as there was no analytics on the feed and as body weights decreased more extensively than feed intake. It may have been possible that feed palatability led to feed wastage.

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Butanoic acid, 4-amino-4-oxosulfo-, N-tallow alkyl derivs., disodium salts

IUPAC Name:

Butanoic acid, 4-amino-4-oxosulfo-, N-tallow alkyl derivs., disodium salts

Constituent 2

Reference substance name:

68988-69-2

Cas Number:

68988-69-2

IUPAC Name:

68988-69-2

Test material form:

liquid: viscous

Details on test material:

- Name of test material (as cited in study report): Aerosol 18
- Physical state: White, thick liquid substance with a faint aromatic odor; aqueous paste
- Analytical purity: 35%
- Impurities (identity and concentrations): See confidential details
- Composition of test material, percentage of components: See confidential details
- Purity test date: Not provided
- Lot/batch No.: #W50704022
- Expiration date of the lot/batch: Not provided
- Stability under test conditions: Not provided
- Storage condition of test material: Not provided

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Wilmington, Mass.
- Age at study initiation: < 5 weeks (deduced from body weights)
- Weight at study initiation: 52-111g
- Fasting period before study: Not provided
- Housing: individual drawer-type wire mesh suspended cages
- Diet (e.g. ad libitum): Charles River, Rat, Mouse and Hamster Meal
- Water (e.g. ad libitum): ad libitum
- Acclimation period: quarantined 1 week prior to the initiation of the experiment

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24.4°C
- Humidity (%): 45-65%
- Air changes (per hr): 10/hour
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

PREPARATION OF FEED PREPARATIONS: The diet was prepared by mixing, on a weight basis, the compound with the basic diet in a Patterson-Kelley twin shell blender, equipped with a high speed mixing bar having slanted blades. Calculations were adjusted to 100% activity.

DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with (Type of food): Charles River, Rat, Mouse and Hamster Meal
- Storage temperature of food: Not provided

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

90 days

Frequency of treatment:

daily

Doses / concentrations

Remarks:

Doses / Concentrations:
0.50,2.00 and 8.00 (first 6 weeks) and 4.00 g/kg bw/day (week 7 through termination)
Basis:
nominal in diet

No. of animals per sex per dose:

20

Control animals:

yes, plain diet

Examinations

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: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): g/rat/week
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY: change in bw(g)/feed(g)/rat/week
- Body weight gain in g/range period (first 6 weeks; second 7 weeks): Yes

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at six weeks and at the termination of the study
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: 40 (5 male and 5 female rats from each group)
- Parameters examined: hematocrit percentage, hemoglobin concentration, total leucocyte counts, differential leucocyte counts

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the conclusion of the experiment
- Animals fasted: No data
- How many animals: the same rats used for hematology when possible
- Parameters examined: blood glucose, blood urea nitrogen, serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, gamma glutamyl transpeptidase

URINALYSIS: Yes
- Time schedule for collection of urine: at six weeks and prior to sacrifice
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
ORGAN WEIGHTS: Yes

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

four rats from the high dose group died during the sixth week; the level was subsequently lowered to 4.00 g/kg/day for the remaining seven weeks

Mortality:

mortality observed, treatment-related

Description (incidence):

four rats from the high dose group died during the sixth week; the level was subsequently lowered to 4.00 g/kg/day for the remaining seven weeks

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

reduced body weight gain in the rats of all levels; this depression appeared more pronounced in the male than in the female rats

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

depressed food consumption at various dose levels; more in male rats than in female rats

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

mid and high dosed males showed a different feed efficiency

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

some rats with increased hemoglobin were observed at the highest dose

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

some female rats with increased SGOT & SGPT observed at the highest dose

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

some rats of the medium and high dose showed hematuria

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Zonal fatty infiltration of the liver in het high dosed males and females.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
Four rats from the high dose group died during the sixth week. The level was subsequently lowered to 4.00 g/kg/day for the remaining seven weeks.

BODY WEIGHT AND WEIGHT GAIN
The male control group gained an average of 397.8 g during the course of the experiment, the 0.50 g/kg/day group gained an average of 321.9 g, the 2.00 g/kg/day group gained an average of 207.9 g and the 8.00g/kg/day-4.00 g/kg/day group (three survivors) gained an average of 134.2 g. The female control group gained an average of 166.6 g, the 0.50 g/kg/day group gained an average of 155.3 g, the 2.00 g/kg/day group gained an average of 113.1 g while the 8.00g/kg/day-4.00 g/kg/day group (six survivors) gained an average of 93.8 g.
From the above data it becomes obvious that Aerosol 18 depresses significantly the rate of body weight gain in the rats of all levels under the conditions of this experiment. This depression appears more pronounced in the male than in the female rats.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
The mean feed consumption data in g/rat/week for the male control group was 178.4 g, for the 0.50 g/kg/day group 171.5 g, for the 2.00 g/kg/day group 159.9g and for the 8.00g/kg/day-4.00 g/kg/day group (three survivors) 144.0 g. The corresponding mean feed consumption for the female control group was 144.3 g, for the 0.50 g/kg/day group 147.1 g, for the 2.00 g/kg/day group 147.5 g and for the 8.00g/kg/day-4.00 g/kg/day group (six survivors) 141.1 g.
During different weeks the feed consumption of the test animals was significantly depressed more so in the male rats than in the female rats.

FOOD EFFICIENCY
The mean feed efficiency (change in body weight (g)/ feed(g)/rat/week) for the male control group was 0.18, for the 0.50 g/kg/day group 0.15, for the 2.00 g/kg/day group 0.10 and for the 8.00g/kg/day-4.00 g/kg/day group (three survivors) 0.07. The corresponding feed efficiency for the female control group was 0.09, for the 0.50 g/kg/day group 0.08, for the 2.00 g/kg/day group 0.06 and for the 8.00g/kg/day-4.00 g/kg/day group (six survivors) 0.06.
The feed efficiency of the male mid dose and male high dose test groups is decidedly different from that of the male control group. The female groups, however, do not differ from the female control group to the same extent as the male test groups.

HAEMATOLOGY
A. Hematocrit
All rats had normal hematocrit concentrations for the duration of the experiment.
B. Hemoglobin
All rats had normal hemoglobin concentration for the duration of the experiment except for rats #4927, #4935 and #4930 (8.00g/kg/day-4.00 g/kg/day group) which had hemoglobin values of 18.1, 18.3 and 18.1 g%, respectively at six weeks. Rat #4935 had a normal hemoglobin at three months; rat #4927 died of acute, toxic cystitis and rat #4930 died of toxic gastritis before three month data could be taken.
C. White Blood Cell Count
Rats #4891 and #4910 (2.00 g/kg/day group) at six weeks had WBC counts of 36600 and 30900 cells/mL, respectively, but by three months their WBC was normal. Rat #4827 (control group) at six weeks had a WBC of 27800 cells/mLand 25100 cells/mL at three months, but its bone marrow was histologically normal.
D. Differential White Blood Cell Count
The albino rat in our experience displays lymphocytosis normally throughout its lifetime. All rats examined at six and 13 weeks displayed lymphocytosis.
E. RBC Morphology
The RBC’s of all animals were morphologically normal at all time.

CLINICAL CHEMISTRY
A. Blood Glucose
All rats had normal blood glucose values.
B. Blood Urea Nitrogen
All rats examined had normal BUN values.
C. Serum Glutamic Oxaloacetic Transaminase
Female rats #4924, #4940 and #4946 (8.00g/kg/day-4.00 g/kg/day group) had SGOT values at three months of 200, 230 and 250 units/liter, respectively.
D. Serum Glutamic Pyruvic Transaminase
Rats #4924, #4940 and #4946 (8.00g/kg/day-4.00 g/kg/day group) has SGPT values at three months of 112, 125 and 140 units/liter, respectively.
E. Gamma Glutamyl Transpeptidase
All rats examined had normal GGTP values.

URINALYSIS
Some of the rats in all groups displayed hematuria to a greater or lesser degree. Of the animals which were sacrificed at the end of the study, rat #4855 (0.50 g/kg/day group) displayed chronic renal disease, usually seen in this strain of rat but at a later age. The urinary system of the other sacrificed rats which displayed hematuria was not remarkable histologically.

ORGAN WEIGHTS
The weight of the parenchymal organs (heart, liver and kidneys) of the mid dose and high dose male rats and the mid dose group of female rats, are significantly lower than the corresponding weight of the relevant control groups. The heart and kidneys of the high dose female rats are also significantly lower. This parallels the decrease in mean body weight of these same groups. In addition, there is a significant decrease in the mean thyroid weight in the mid dose and high dose rats and in the mean adrenal weight of the mid dose and high dose male groups as well as of all female test groups. Finally there is a significant decrease in the mean ovarian weight of the mid dose and high dose group.

GROSS PATHOLOGY
Zonal fatty infiltration of the liver in het high dosed males and females.

HISTOPATHOLOGY: NON-NEOPLASTIC : The 8.00 g/kg/day rats which died halfway through the experiment displayed classical acute toxic findings of the gastrointestinal and urinary system as described in the appropriate necropsy reports. Some of the high dose test rats which were sacrificed at the end of the experiment displayed zonal fatty infiltration of the liver. The rest of the examined rats from the terminal sacrifice displayed normal histology.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
No effects

Effect levels

1

Dose descriptor:

NOEL

Effect level:

< 500 mg/kg diet

Based on:

act. ingr.

Sex:

male/female

2

Dose descriptor:

NOAEL

Effect level:

500 mg/kg diet

Based on:

act. ingr.

Sex:

male/female

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Table 1. Terminal Body Weight and Organ Weights for male and female rats receiving Aerosol 18 in their diet for 13 weeks

	Diet	Terminal	Organ Weights (grams)/Body Weight (kilograms)
	Conc.	Body
Sex	g/kg	Weight(g)	Thyroids	Heart	Liver	Adrenals	Kidneys	Gonads	Pituitary	Brain
Male	0	466.0	0.030	1.163	17.315	0.061	3.132	3.425	0.012	2.091
	0.50	392.0*	0.029	1.031	14.484	0.056	2.656*	3.035	0.013	2.075
	2.00	285.7*	0.024*	0.766*	11.169*	0.052*	1.990*	3.569	0.012	2.058
	4.00(a)	219.0*	0.023	0.537*	8.120*	0.050*	1.483*	2.880	0.010	1.817
Female	0	270.8	0.031	0.797	11.051	0.088	2.069	0.151	0.016	1.987
	0.50	246.4	0.028	0.746	10.387	0.071*	1.804*	0.146	0.012	1.980
	2.00	200.7*	0.022*	0.573*	7.406*	0.060*	1.470*	0.079*	0.010*	2.013
	4.00(b)	182.2*	0.023*	0.517*	8.850	0.058*	1.308*	0.070*	0.012	1.850*

*Statistically significant from control at P=0.05.

 (a) Three survivors at termination

(b) Six survivors at termination

 

Table 2. Organ-to-Body Weight Ratio data for male and female rats receiving Aerosol 18 in their diet for 13 weeks (grams of organ weight/ kilograms of body weight)

	Diet	Terminal	Organ Weights (grams)/Body Weight (kilograms)
	Conc.	Body
Sex	g/kg	Weight(g)	Thyroids	Heart	Liver	Adrenals	Kidneys	Gonads	Pituitary	Brain
Male	0	466.0	0.064	2.496	37.157	0.131	6.721	7.350	0.026	4.487
	0.50	392.0	0.074	2.630	36.949	0.143	6.776	7.742	0.033	5.293
	2.00	285.7	0.084	2.681	39.093	0.182	6.965	12.492	0.042	7.203
	4.00(a)	219.0	0.105	2.452	37.078	0.228	6.772	13.151	0.046	8.297
Female	0	270.8	0.114	2.943	40.809	0.325	7.640	0.558	0.059	7.338
	0.50	246.4	0.144	3.028	42.155	0.288	7.321	0.593	0.049	8.036
	2.00	200.7	0.110	2.855	36.901	0.299	7.324	0.394	0.050	10.030
	4.00(b)	182.2	0.126	2.838	45.573	0.318	7.179	0.384	0.066	10.154

 (a) Three survivors at termination

(b) Six survivors at termination

Applicant's summary and conclusion

Conclusions:

From the data presented in this study it appears that the NOEL for Aerosol 18 in the rat is < 500 mg active ingredient/kg feed/day when orally administered; NOAEL is considered 500 mg/kg bw/day.

Executive summary:

The purpose of this study was to investigate the toxic effects, if any, of test item containing 35% active ingredient when fed to albino rats for ninety days at doses of 0.50, 2.00 and 8.00 g act.ingr./kg bw/day. The rats were carefully observed for the duration of the experiment, body weight and feed intake /conversion were measured and pertinent haematological and biochemical parameters were conducted after 6 weeks and after termination. At the conclusion of the experiment all rats were necropsied and all organs and/or tissues were examined histologically from ten rats (five male and five female). Due to mortality of 4 rats at 8 g/kg bw/day, dosing was reduced to 4 g/kg feed/day after 6 weeks. The study further showed decreased body weight gain and food intake at all levels and acute gastro-enteritis and/or acute nephritis within six weeks at 8.00 g/kg/day. When this dose was decreased to 4.00 g/kg/day and given for seven weeks some rats displayed zonal fatty infiltration in the liver. At the lowest dose level, decreases in body weight (gain) and feed intake were minimal (<10%), and were not considered relevant. Decreased organ weights were observed for heart, liver, kidneys, adrenal an thyroid weight at the medium and high doses. From the data presented in this study it appears that the NOEL was < 500 mg act. ingr./kg bw/day, however the NOAEL can be considered to be 500 mg/kg bw/day.