Diethylammonium chloride

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

NOAEL (oral, rat, OECD 422) = 200 mg/kg bw/day (RA from CAS 75-50-3)

Link to relevant study records

Reference

Reference 1

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

(Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

(no data on detailed clinical observation, sensory reactivity to stimuli, assessment of grip strength, and motor activity)

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

yes

Remarks:

no data on detailed clinical observation, sensory reactivity to stimuli, assessment of grip strength, and motor activity

Qualifier:

according to guideline

Guideline:

other: OPPTS 870-3650

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

-Age: 9 weeks

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

Males: 42 days; Females: 2 weeks prior to breeding, continuing through breeding (2 weeks), gestation (3 weeks), lactation (4 days), and until the day of necropsy (test day 40 or 54).

Details on mating procedure:

Male/female per cage maximum for 2 weeks beginning evening of dosing day 15. Mating confirmed by existence of sperm in the vaginal plug  and vaginal smear every morning.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

Once daily

Frequency of treatment:

daily

Dose / conc.:

8 mg/kg bw/day

Dose / conc.:

40 mg/kg bw/day

Dose / conc.:

200 mg/kg bw/day

No. of animals per sex per dose:

13

Control animals:

yes, concurrent vehicle

Parental animals: Observations and examinations:

General condition was observed at least once a day during breeding and at least twice a day before and after dosing over the administration period. Body weights for males were determined on days 1, 7, 14, 21, 28, 35, 42 and on the day of necropsy. Body weights were determined for all females on days 1, 7, 14. Females who took time before mating were weighed on days 35 and 42. Females who copulated were weighed on pregnancy days 0, 7, 14 and 20. Females who delivered were weighed on nursing days 0, 4, and on the day of necropsy. Females who copulated but did not deliver were weighed on the equivalent of pregnancy day 25 (day of necropsy).
Food consumption was measured on days 1, 7, 13, 29, 35 and 41 for males, and on days 1, 7, and 13 for all females. Females with unconfirmed copulation were measured for food consumption on days 29, 35 and 41.

Oestrous cyclicity (parental animals):

-Estrus Cycle: Estrus cycle was determined from the vaginal smear and the mean number of days to mating season was recorded.

Litter observations:

-Number of delivered pups:  The number of delivered pups (live pups +  stillborns) was counted on nursing day 0, and the delivery rate ((number  of delivered pups/number of implantation traces) x 100) and the live  birth rate ((number of delivered live pups/number of implantation traces)  x 100) were calculated. With the delivered pups, external malformation and the sex were checked and the sex ratio ((number of male live pups/number of female live pups) x 100) was calculated.
-Number of dead pups: The number of dead pups was checked daily, and the  birth rate ((number of delivered live pups/number of delivered pups) x  100) and the viable rate of new-born pups on day 4 ((number of live pups  on nursing day 4/number of live pups on nursing day 0) x 100) were  calculated. Dead pups were autopsied and abnormality and internal organs were observed.

Postmortem examinations (parental animals):

-Urinalysis was conducted on 5 rats/sex/dose level at week 6.
-Hematology and clinical chemistry: Males prior to necropsy and on the  following day after day 42 administration; Females prior to necropsy:  females who delivered-following nursing day 4, females who mated but did  not deliver-equivalent of pregnancy day 25, and females who did not mate-  following day 54 of administration

ORGANS EXAMINED AT NECROPSY (MACROSCOPIC AND MICROSCOPIC):
-Macroscopic: organ weights: testes and epididymides; females, ovaries and uteri were extracted, the pregnancy corpora lutea number of the ovary was counted under the stereoscopic microscope, the implantation number of the uterus was counted, and the implantation rate ((implantation number/pregnancy corpora lutea number) x 100) was calculated.
-Microscopic: 5 animals/sex/control and high dose group- sperm and prostrate ventral lobes of all males and vagina, ovaries and uteri of all females; also testes, epididymides, ovaries found to be abnormal during pathologic examinations were all examined histopathologically.

Statistics:

Fisher's Exact Test- mating and conception rate,
Mann-Whitney U Test (2-tailed) and Fisher's Exact Test (1-tailed): histopathological examinations,
Dunnett's Multiple Comparison Test (significance level=5%): body weight, food consumption, hematology, clinical chemistry and organ weights

Reproductive indices:

-Copulation: When mating was confirmed that day was reckoned as pregnancy  day 0.  From the results of copulation, mating rate ((number of animals  mated/number of animals cohabited) x 100), conception rate ((number of  animals conceived/number of animals mated) x 100), number of days needed  for mating from the start of cohabitation and the number of times of  estrus which recurred during that time were calculated.

-Observation of delivery: With all cases where delivery was confirmed,  the pregnancy period (number of days from pregnancy day 0 to delivery  day) was calculated and the birth rate ((number of females who delivered  live pups/number of animals conceived) x 100) for each group was found. 

Offspring viability indices:

-Number of delivered pups:  The number of delivered pups (live pups +  stillborns) was counted on nursing day 0, and the delivery rate ((number  of delivered pups/number of implantation traces) x 100) and the live  birth rate ((number of delivered live pups/number of implantation traces)  x 100) were calculated. With the delivered pups, external malformation  and the sex were checked and the sex ratio ((number of male live  pups/number of female live pups) x 100) was calculated.
-Number of dead pups: The number of dead pups was checked daily, and the  birth rate ((number of delivered live pups/number of delivered pups) x  100) and the viable rate of new-born pups on day 4 ((number of live pups  on nursing day 4/number of live pups on nursing day 0) x 100) were  calculated. Dead pups were autopsied and abnormality and internal organs  were observed

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

-Clinical signs prior to death
200 mg/kg bw/day: The male that died on Day 25 showed salivation, emaciation, abnormal breathing noise and dyspnea from administration day 19, and vulval peripheral fur soiled and loose passage were observed from the day before the death. The male that died on Day 42 showed salivation, emaciation, abnormal breathing noise, dyspnea, a drop in body temperature, faded auricle, tottering and brown soil around the nose were observed from administration day 10, although discontinuously. The female rat that died on pregnancy day 22, was observed with salivation and abnormal breathing noise sporadically from administration day 11.

-Clinical signs in surviving animals:
200 mg/kg bw/day:
Males: salivation 10/13, abnormal breathing noise 3/13, and decreased contact response 1/13;
Females: salivation 10/13, abnormal breathing noise 3/13, and emaciation 1/13.

40 mg/kg/day: no abnormalities

Mortality:

mortality observed, treatment-related

Description (incidence):

-Mortality and time to death: 1 male given 200 mg/kg/day died on day 25, 1 male given 200 mg/kg/day died on day 42 and 1 female died on pregnancy day 22 (administration day 38)

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Males
No significant differences in body weights at 200 mg/kg bw/day; however, body weight gains were decreased when compared to controls. There were no significant differences in body weight or body weight gains in males administered 40 mg/kg bw/day.
Females
No significant differences in body weight or body weight gains.

Food consumption and compound intake (if feeding study):

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

40 mg/kg bw/day: significant increase in urea nitrogen

Urinalysis findings:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Other effects:

not specified

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

Daily oral administration of trimethylamine by gavage resulted in the death of two males and 1 female administered 200 mg/kg bw/day. Abnormal breathing noise, salivation immediately after the administration, ulcers and inflammatory changes in the stomach and intestinal tracts, squamous hyperplasia and edema in submucosa were observed in both males and females in the 200 mg/kg bw/day group. An inhibitory tendency in body weight increase, decrease in food consumption, total protein concentration and albumin concentration were also observed in the males in the same group.
There was no effect of trimethylamine administration on body weights and food consumption of the females and on organ weights, urine examination and hematological examination results in the males and females. Therefore it was inferred that the general toxicological NOAEL (No Observed Adverse Effect Level) is 40 mg/kg bw/day.
Adverse and treatment-related effects on reproductive performance were not observed at 200 mg/kg bw/day in the dams, furthermore no embryo-/fetotoxicity was observed in the offspring. No abnormality which can be considered the effects of trimethylamine administration was observed in the reproductive organs, and no effect on the observation results of estrous cycle was found. Also no effect was found on the mating rate, conception rate, the birth rate, pregnancy period, and nursing state, the number of corpora lutea, the number and rate of implantation, the viability of the delivered pups, sex ratio, body weight and form. From the above, it was considered that reproductive/developmental toxicity NOEL is 200 mg/kg bw/day for both males and females and also for delivered pups.

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

40 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Treatment-related and adverse effects (clinical signs and mortality) observed at 200 mg/kg bw/day

Key result

Dose descriptor:

NOEL

Remarks:

reproduction

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No treatment-related adverse effects (the mating rate, conception rate, the birth rate, pregnancy period, and nursing state, the number of corpora lutea, the number and rate of implantation) observed at the highest tested dose level

Critical effects observed:

not specified

Clinical signs:

not specified

Mortality / viability:

not specified

Body weight and weight changes:

not specified

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Histopathological findings:

not specified

no details given

Key result

Dose descriptor:

NOEL

Remarks:

developmental

Generation:

F1

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No treatment-related adverse effects (the viability of the delivered pups, sex ratio, body weight and form) observed at the highest tested dose level

Key result

Critical effects observed:

not specified

Key result

Reproductive effects observed:

not specified

Conclusions:

Adverse and treatment-related effects on reproductive performance were not observed at 200 mg/kg bw/day in the dams, furthermore no embryo-/fetotoxicity was observed in the offspring. No abnormality which can be considered the effects of trimethylamine administration was observed in the reproductive organs, and no effect on the observation results of estrous cycle was found. Also no effect was found on the mating rate, conception rate, the birth rate, pregnancy period, and nursing state, the number of corpora lutea, the number and rate of implantation, the viability of the delivered pups, sex ratio, body weight and form. From the above, it was considered that reproductive/developmental toxicity NOEL is 200 mg/kg bw/day for both males and females and also for delivered pups.

The NOAEL for systemic toxicity was considered to be 40 mg/kg bw/day based on abnormal breathing noise and salivation immediately after the administration, ulcers and inflammatory changes in the stomach and intestinal tracts, squamous hyperplasia and edema in submucosa, which were observed in both males and females in the 200 mg/kg bw/day group. These local effects (ulcers, inflammation) at sites of contact (stomach, intestines) were considered to be due to corrosive properties of substance.

Reference 2

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

(Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

refer to analogue justification provided in IUCLID section 13

Reason / purpose for cross-reference:

read-across source

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

40 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Treatment-related and adverse effects (clinical signs and mortality) observed at 200 mg/kg bw/day

Remarks on result:

other: Source: CAS 75-50-3, Takashima, 2003

Key result

Dose descriptor:

NOEL

Remarks:

reproduction

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No treatment-related adverse effects (the mating rate, conception rate, the birth rate, pregnancy period, and nursing state, the number of corpora lutea, the number and rate of implantation) observed at the highest tested dose level

Remarks on result:

other: Source: CAS 75-50-3, Takashima, 2003

Key result

Critical effects observed:

not specified

Key result

Dose descriptor:

NOEL

Remarks:

developmental toxicity

Generation:

F1

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No treatment-related adverse effects (the viability of the delivered pups, sex ratio, body weight and form) observed at the highest tested dose level

Remarks on result:

other: Source: CAS 75-50-3, Takashima, 2003

Key result

Critical effects observed:

not specified

Reproductive effects observed:

no

Conclusions:

A screening for reproductive / developmental toxicity test with the source substance trimethylamine (CAS 75-50-3) had no effect on reproductive performance leading to a NOEL of 200 mg/kg bw/day for reproduction and developmental toxicity. Applying the RA-approach similar results are expected for the target substance.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

200 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

The available information comprises an adequate and reliable study (Klimisch score 2) from a reference substance with similar structure and intrinsic properties. Read-across is justified based on structural similarities (refer to endpoint discussion for further details). The selected study is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.7, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Effect on fertility: via inhalation route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

49 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

The available information comprises an adequate and reliable study (Klimisch score 2) from a reference substance with similar structure and intrinsic properties. Read-across is justified based on structural similarities (refer to endpoint discussion for further details). The selected study is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.7, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Justification for read-across

There are no data regarding reproduction toxicity available for diethylammonium chloride (CAS 660-68-4). In order to fulfil the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.7, read-across from appropriate substances is conducted in accordance with Regulation (EC) No 1907/2006, Annex XI, 1.5. Structural similarities and similarities in properties and/or activities of the source and target substance are the basis of read-across. A detailed justification for the analogue read-across approach is provided in the technical dossier (see IUCLID Section 13).

As no data is available regarding developmental toxicity for diethylammonium chloride (CAS 660-68-4) reliable information from the analogue substance trimethylamine (CAS 75-50-3) is taken into account to fulfil the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.7. In addition, supporting information from two reliable inhalation repeated dose toxicity studies (90-day) in rats and mice conducted with diethylamine (CAS 109-89-7) were taken into account.

CAS 75-50-3

Reliable data is available for the source substance trimethylamine (CAS 75-50-3). Trimethylamine was tested in a combined repeated dose oral toxicity study with the reproduction/developmental toxicity screening study according to OECD Guideline 422 (Takashima et al., 2003). 13 Sprague Dawley rats per sex and dose were treated via gavage with the test substance at doses of 8, 40 and 200 mg/kg bw/day, respectively. The control group received the vehicle water. Males were treated for 42 days. Females were treated for 2 weeks prior to breeding, continuing through breeding (2 weeks), gestation (3 weeks), lactation (4 days), and until the day of necropsy.

Clinical observations, food consumption, body weights, hematology, clinical chemistry, urinalysis, organ weights and gross examination at necropsy, microscopic examination of various organs and tissues and assessment of various reproductive and developmental parameters, e.g. estrous cycle, duration of gestation, copulation index, fertility index for females, gestation index, live birth index, viability index, pup mortality, sex ratio and implantation index were used for detecting the effects of treatment.

At 200 mg/kg bw/day, one male died on day 25 and on day 42, respectively. One high-dose female died on pregnancy day 22 (administration day 38). Clinical signs such as abnormal breathing noise and salivation were observed in the surviving animals and the animals of the high-dose group that died, respectively. In addition, emaciation and dyspnea were noted in the high-dose males that died. No abnormalities on reproductive organs were observed after trimethylamine administration, and no effects on the observation results of estrous cycle were found. Examination of reproductive parameters such as mating rate, conception rate, the birth rate, pregnancy period, and nursing state, the number of corpora lutea, the number and rate of implantations revealed no treatment-related and adverse effects by the test substance. Based on the results of this study, the NOAEL systemic was derived to be 40 mg/kg bw/day and the NOEL for reproductive performance was derived to be 200 mg/kg bw/day.

CAS 109-89-7

In the study, groups of 10 male and 10 female rats were exposed to diethylamine vapor at concentrations of 0, 8, 16, 32, 62, or 125 ppm (corresponding to 24, 49, 97, 188 and 379 mg/m³, respectively), 6 hours plus T90 (12 minutes) per day, 5 days per week for 14 weeks. All rats survived to the end of the study. Mean body weights of all exposed groups were similar to those of the chamber control groups. There were significant exposure concentration-related decreases in sperm motility in 32, 62, and 125 ppm males, however no histopathological correlate was noted; there were no significant differences in the lengths of estrous cycles between chamber control and exposed groups of females. From the data a systemic NOAEC of 16 ppm (corresponding to 49 mg/m3) could be established. Considering the fertility effects, a NOAEC of 16 ppm for males based on the effect on sperm motility was derived.

In a second study, groups of 10 male and 10 female mice were exposed to diethylamine vapor at concentrations of 0, 8, 16, 32, 62, or 125 ppm (corresponding to 24, 49, 97, 1882 and 379 mg/m³, respectively), 6 hours plus T90 (12 minutes) per day, 5 days per week for 14 weeks. All mice survived to the end of the study. The mean body weights of 125 ppm males and females were significantly less than those of the chamber controls. There were significant exposure concentration-related decreases in sperm motility in males exposed to 32, 62, or 125 ppm; the estrous cycle of 125 ppm females was significantly longer than that of the chamber controls but only by half a day. However, no histopathological correlate was noted. Histopathologic changes were noted primarily in the nasal cavity and involved both the respiratory and olfactory epithelium of males and females principally in the 62 or 125 ppm groups. These lesions included suppurative inflammation, squamous metaplasia of the respiratory epithelium, olfactory epithelial atrophy, and necrosis of the turbinates. From the data a systemic NOAEC of 16 ppm (corresponding to 49 mg/m3) could be established. Considering the fertility effects, a NOAEC of 16 ppm for males based on the effect on sperm motility and of 62 ppm for females based on changes in estrous cycling was derived, respectively.

The effects on the estrous cycle cannot be regarded as a clear effect on reproduction toxicity, since the estrous cycle is a sensitive parameter that can be influenced by stress, e.g. the inflammation of the olfactory epithelium in the mid- and high-dose group can be regarded as a stress-mediated response. Sperm motility is the endpoint which exhibits the lowest variation and highest sensitivity of all sperm parameters (Mangelsdorf, 2002). Further, fertility is closely correlated with sperm motility. The effects on sperm motility were dose-related and were observed in rats and mice, therefore an influence on the male fertility cannot be clearly excluded. However, there is no further evidence on the reduction of the fertility from the experimental data, and in addition no histopathological correlate was reported, therefore these effects are considered not to be sufficient for classification.

Effects on developmental toxicity

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Justification for read-across

There are no data regarding reproduction toxicity available for diethylammonium chloride (CAS 660-68-4). In order to fulfil the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.6, read-across from appropriate substances is conducted in accordance with Regulation (EC) No 1907/2006, Annex XI, 1.5. Structural similarities and similarities in properties and/or activities of the source and target substance are the basis of read-across. A detailed justification for the analogue read-across approach is provided in the technical dossier (see IUCLID Section 13).

As no data is available regarding developmental toxicity for diethylammonium chloride (CAS 660-68-4) reliable information from the analogue substance trimethylamine (CAS 75-50-3) is taken into account to fulfill the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.6.

Developmental Toxicity

CAS 75-50-3

Reliable data is available for the source substance trimethylamine (CAS 75-50-3). Trimethylamine was tested in a combined repeated dose oral toxicity study with the reproduction/developmental toxicity screening study according to OECD Guideline 422 (Takashima et al., 2003). 13 Sprague Dawley rats per sex and dose were treated via gavage with the test substance at concentrations of 8, 40 and 200 mg/kg bw/day, respectively. The control group received the vehicle water. Males were treated for 42 days. Females were treated for 2 weeks prior to breeding, continuing through breeding (2 weeks), gestation (3 weeks), lactation (4 days), and until the day of necropsy.

Clinical observations, food consumption, body weights, hematology, clinical chemistry, urinalysis, organ weights and gross examination at necropsy, microscopic examination of various organs and tissues and assessment of various reproductive and developmental parameters, e.g. estrous cycle, duration of gestation, copulation index, fertility index for females, gestation index, live birth index, viability index, pup mortality, sex ratio and implantation index were used for detecting the effects of treatment.

At 200 mg/kg bw/day, one male died on day 25 and on day 42, respectively. One high-dose female died on pregnancy day 22 (administration day 38). Clinical signs such as abnormal breathing noise and salivation were observed in the surviving animals and the animals found dead of the high-dose group, respectively. In addition, emaciation and dyspnea were noted in the high-dose males found dead. Examination of developmental parameters such as the viability of the delivered pups and viability of pups on day 4, sex ratio and body weight revealed no treatment-related and adverse effects by the test substance. Based on the results of this study, the NOAEL systemic was derived to be 40 mg/kg bw/day and the NOEL for developmental toxicity was derived to be 200 mg/kg bw/day. The available study does not indicate a developmental toxic potential of the test substance, however a final conclusion on the developmental toxicity cannot be made based on the results of a screening study.

Justification for classification or non-classification

According to Article 13 of Regulation (EC) No. 1907/2006 "General Requirements for Generation of Information on Intrinsic Properties of substances", information on intrinsic properties of substances may be generated by means other than tests e.g. from information from structurally related substances (grouping or read-across), provided that conditions set out in Annex XI are met. Annex XI, "General rules for adaptation of this standard testing regime set out in Annexes VII to X” states that “substances whose physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity may be considered as a group, or ‘category’ of substances. This avoids the need to test every substance for every endpoint". Since the analogue concept is applied to diethylammonium chloride, data will be generated from information on reference source substances to avoid unnecessary animal testing. Additionally, once the analogue read-across concept is applied, substances will be classified and labelled on this basis.

 

Applying the RA approach, the available data on fertility / developmental toxicity do not meet the classification criteria according to Regulation (EC) No. 1272/2008.