N-benzyl-N-C16-18 (even numbered)-alkyl-N-methyl-C16-18 (even numbered)-alkyl-1-aminium chloride

Endpoint summary

Description of key information

The potential toxicity of BDHTMAC (see Toxicological information section for the justification of read across) following repeated administration was assessed using:
- two subacute (28-day) oral (gavage) studies performed in rats according to OECD guideline 407 and Good Laboratory Practices with the test substance, BDHTMAC (Queudot, 2010a) or with dimethyldioctadecylammonium chloride (DODMAC), the major active component of the ditallowdimethylammonium chloride (DHTDMAC)(Hoechst, 1990).
- two subchronic (90-day) feeding studies performed with the analogue substance DHTDMAC,  in Beagle-dogs or in rats according to a method similar to OECD guideline 409 and 408, respectively (Lindberg, 1971).
The results of the four studies were coherent and indicated that the substance was of moderate toxicity by repeated gavage administration. Based on some limitations of the studies perfomed with the analogue substances, the valid 28-day subacute toxicity study in rats performed with the test substance (Queudot, 2010a) is taken for risk characterisation. According to the results of this study, the NOAEL with regard to systemic oral toxicity was established at 100 mg/kg body weight per day. 
 In an earlier study predating official test guidelines and Good Laboratories Practices, rabbits received technical grade of DODMAC (75% active in isopropanol/water) via the dermal route of exposure. Twenty applications (5 per week for 4 consecutive weeks) of 2 mL of 0, 0.2 and 2% aqueous test substance solutions (corresponding to about 0, 4 and 40 mg/kg body weight per day) induced mild local dermal effects but no clinical or morphological signs of substance related systemic toxicity. Based on an analogue approach, The NOAEL for the dermal systemic effects of the substance was considered to be greater than 40 mg/kg body weight per day.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion

Dose descriptor:

NOAEL

100 mg/kg bw/day

Study duration:

subacute

Species:

rat

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Dose descriptor:

NOAEL

40 mg/kg bw/day

Study duration:

subacute

Species:

rabbit

Additional information

Repeated dose toxicity: oral

 

The objective of the study of Queudot (CIT, 2010) was to evaluate the potential toxicity of the test substance (N-benzyl-N-C16-18(even numbered)-alkyl-N-methyl-C16-18(even numbered)-alkyl-1-aminium chloride (BDHTMAC) following daily oral administration (gavage) to rats for 4 weeks according to OECD guideline 407 and EU method B.43. The study was conducted in compliance with the principles of Good Laboratory Practice regulations.

Three groups of five male and five female Sprague-Dawley rats received the test item by oral administration (gavage) at dose‑levels of 50, 100 or 250 mg/kg/day for 4 weeks. A constant dosage-volume of 10 mL/kg was used.

In addition, five males and five females received the vehicle only (corn oil) under the same experimental conditions and acted as controls.

Samples of dosage forms administered on day 1 and on a day 22 were analyzed for test item concentration.

The animals were checked daily for mortality and clinical signs. In addition, a detailed clinical examination was performed once before the beginning of the treatment period and then once a week until the end of the study. A Functional Observation Battery was performed at the end of the treatment period (day 27).

Body weight was recorded once before beginning of the treatment period, then on the first day of treatment and once a week until the end of the study. Food consumption was recorded once a week during the study. Hematology, blood biochemistry and urine investigations were performed on day 29.

On completion of the treatment period, the animals were euthanized and submitted to a full macroscopicpost-mortemexamination. Designated organs were weighed and selected tissue specimens were preserved. A microscopic examination was performed on designated tissues from the control and high-dose groups. According to the microscopic results of the highest-dose group the adrenals of low- and intermediate‑dose animals were examined.

 The test item concentrations in the administered dosage forms analyzed in weeks 1 and 4 remained within an acceptable variation range of -7.7% to +1.2% when compared to the nominal values.

No mortalities occurred during the study.

At 250 mg/kg bw/ day, loud and abdominal breathing were noted in 2/5 females during the second part of the treatment period. A statistically significant increase in alanine aminotransferase activity was noted in males and females (2.5 -fold and 1.7 -fold, respectively).There was a statistically significant increase in adrenal weights in both sexes which correlated with a slight hypertrophy (grade 2 on a maximum scale of 4) of adrenal cortical cells in 4 out 5 males and in 1 out of 5 females. All the remaining animals of this dose-group showed a minimal hypertrophy (grade 1).

At 50 and 100 mg/kg/day, no adverse test item-related effects were noted. A slight but not dose-related increase in adrenal weights was observed. Minimal hypertrophy of adrenal cortical cells (grade 1) was noted in some animals. One female of the control group showed also a minimal hypertrophy of adrenal cortical cells (grade 1). In the absence of associated degeneration necrosis, this change was not considered to be adverse.

Consequently, under the experimental condition of this study, the No Observed Adverse Effect Level (NOAEL) was set at 100 mg/kg/day.

 

 

The oral repeated dose toxicity of quaternary ammonium compound, di-C16 -18 -alkyldimethyl, chloride was also investigated as described below in several subacute and subchronic studies performed in rats and dogs using either technical grade or pure dioctadecyldimethylammonium chloride (DODMAC) or dihydrogenated tallowalkyldimethylammonium chloride (DHTDMAC).

In a subacute (28-day) oral toxicity study (Hoechst, 1990), dioctadecyldimethylammonium chloride (DODMAC), 90% active in 5% isopropanol and 5% water, was administered by gavage in sesame oil at doses of 0, 20, 100, and 500 mg/kg to Wistar rats. The study was performed according to OECD guideline 407 in compliance with the principles of Good Laboratory Practices. Dosages were calculated on nominal concentration values.

Beginning at the 8th day (females) and 14th day (males) of treatment some high dose animals showed squatting position, abnormal gait, disregular and noisy breathing. Reduced spontaneous activity, retracted flanks and distended abdomen were also seen in some high dose females.

Body weight gain was slightly (non-significantly) lower in high dose males and females compared to the control values.

Hematology revealed significantly reduced reticulocyte counts in high dose males which were within the normal range for this species, sex and age, whereas no other red cell parameter was changed. Mean values for segmented neutrophile ratio were increased in high dose males and females due to abnormal rates in two males and three females. Lower concentrations of albumin and the albumin-globulin ratio and higher gamma-globulin values were observed in males of the high dose group (all changes were significant).

Absolute and relative organ weights of the adrenals were determined to be significantly increased in high dose males. Adrenal weights of high dose females were also higher than controls, however adrenals from three females only were weighted. Macroscopically, enlargement of the adrenals was seen in one female and discoloration of the adrenal surface was evident in three females of the high dose group. Corresponding to these observations in the adrenals, two females had cortical necrosis with peripheral granulocytic infiltration, one of this was hemorrhagic. Furthermore in a single high dose female ulceration of the forestomach was found. The NOAEL from this study was considered to be 100 mg/kg bw/d.

In a subchronic (90-day) feeding study (Lindberg,1971), 4 male and 4 female Beagle dogs per group received dihydrogenatedtallow dimethyl ammonium chloride (DHTDMAC) of unknown purity at dose levels of 0,14,140 and 2800 ppm via the diet. The study included investigations on clinical signs, food consumption, body and organ weights, selected parameters on haematology, clinical chemistry and urinalysis on days 45 and 90, and histopathology of numerous organs / tissues. No test substance-related effects were seen following 90 days of administration. The NOAEL of DHTDMAC in Beagle dogs from this study was established at 2800 ppm in the diet, corresponding to about 756 mg/kg body weight per day in males and 935 mg/kg body weight per day in females.

 

In another subchronic (90 -day) feeding study combined to a reproductive assay (Smith, 1971), dihydrogenatedtallowdimethylammonium chloride (DHTDMAC) of unknown purity was administered to rats at dose levels of 0, 7, 140 and 2800 ppm. No treatment-related mortalities were observed.

There was a slight depression in weight gain of the treated rats compared to controls, however there was no dose-response relationship. Food consumption was lower for all treated groups compared to controls but it appears that the rats were not the same age at the start of the study and there were large differences in the starting weights and subsequent weights because of this. This may explain the differences in weight gain, and the differences in food consumption as food consumption data were not corrected for body weight.

There were no differences in haematology, clinical chemistry or urinalysis between 2800 ppm rats and controls.Lesions noted at histopathological examination were those of spontaneous disease and not unusual for rats. The most frequent findings were lesions in the trachea and lungs, indicating chronic murine pneumonia. Lesions occurred in both treated and control rats.The NOAEL of DHTDMAC in rats from this study was established at 2800 ppm in the diet.

 

 

The results of the four studies were coherent however the two last studies (Lindberg and smith, 1971). were conducted by Industrial Bio-Test Laboratories Inc.(IBT) and no information was available as to whether these studies had been audited. Based on section 3.1.8 of the Manual for investigation of HPV chemicals regarding the acceptance and use of IBT studies (OECD, last update on december 2005), when a study has not been audited by either EPA or FDA (this is the assumption here), and if the findings of the IBT study were consistent with a study conducted at a later date in another laboratory, then the data could be used but should be considered only as a supportive evidence. in this case, the IBT studies are used in support with another independent references (Queudot, 2010a and Hoechst, 1990) and consolidate the toxicological profile of the substance indicating its moderate toxicity by repeated gavage administration.

Based on these limitations, the valid 28-day subacute toxicity study in rats performed with the test substance (Queudot, 2010a) is taken for risk characterisation. According to the results of this study, the NOAEL with regard to systemic oral toxicity was established at 100 mg/kg body weight per day.

 

 

Repeated dose toxicity: dermal

in a dermal repeated dose study (Hoechst, 1974), technical grade dioctadecyldimethylammonium chloride (DODMAC) containing 75% active in isopropanol /water was applied onto rabbit skin. The study predated official test guidelines and GLP but give some information on the potential systemic toxicity of quaternary ammonium compound, di-C16 -18 -alkyldimethyl, chloride via the dermal route of exposure.

Groups of 3 male and 3 female rabbits (strain "Gelbsilber") received 20 dermal applications (5 days per week for 4 consecutive weeks) of aqueous solutions containing 0, 0.2 and 2% DODMAC (corresponding to about 0, 4 and 40 mg/kg body weight per day). General behaviour, general health condition, food consumption were not influenced by the treatment. Haematology, clinical chemistry and urinalysis revealed no significant findings. Gross pathology of the animals at study termination as well as histopathological investigations revealed no substance related changes. Local skin effects in form of slight redness and foldings were observed in some of the high dose animals.

Based on the results of this study the NOAEL for systemic dermal effects was greater 40 mg/kg body weight per day (2% (v/v) aqueous test substance solution).

Based on the limitations of the 90-day feeding studies, the subacute oral 28 -day studies in rats are considered the most appropriate for evaluation purposes especially the oral 28-day study performed with the substance of interest one subacute (28-day) oral (gavage) study on the test substance, N-benzyl-N-C16-18(even numbered)-alkyl-N-methyl-C16-18(even numbered)-alkyl-1-aminium chloride. This view is in line with the conclusions of the existing EU risk assessment on Dioctadecyldimethylammonium chloride (DODMAC).

Repeated dose toxicity: via oral route - systemic effects (target organ) digestive: liver; glandular: adrenal gland

Justification for classification or non-classification

Based on the results of the Queudot study (2010) and on all available data of the quaternary ammonium compounds, di-C16 -18 -alkyldimethyl, chloride and according to the criteria laid down in EU regulation (EC) n° 1272/2008/EC (CLP) and EU directive67/548/EEC (DSD), the substance is not classified for repeated dose toxicity.