Dimethyl disulphide

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:

1989-04-24 to 1989-05-23

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

yes

Remarks:

see below: Any other information ...

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Dimethyl disulphide

EC Number:

210-871-0

EC Name:

Dimethyl disulphide

Cas Number:

624-92-0

Molecular formula:

C2H6S2

IUPAC Name:

(methyldisulfanyl)methane

Details on test material:

Source: Atochem
Batch: 6437 from lot 82551
Purity: 99.88%

Test animals

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ORGANISMS:
- Source: ENKI-Konijnenfarm, someren, the Netherlands
- Age at reception: 12 weeks old - Weight at the start of the treatment: no data
- Number of animals:  The control and top-dose group comprised 10 males  and 10 females, whereas the low- and mid-dose group comprised 5 males and  5 females.
- Aclimatation period: 13 days

HOUSING
The animals were housed individually in suspended, galvanized cages,  fitted with a wire-mesh floor and front.

FOOD and WATER
- Food: standard laboratory rabbit diet ad libitum
- Water: tap water, ad libitum 

ENVIRONMENTAL CONDITIONS
- Temperature : 18 ± 3°C
- Relative humidity : at least 40%
- Light/dark cycle : 12h/12h
- Ventilation : at least 10 changes/hour

Administration / exposure

Type of coverage:

occlusive

Vehicle:

unchanged (no vehicle)

Details on exposure:

Doses were applied by volume, viz. 1.0, 0.1, and 0.01 ml/kg body weight  for the top-dose, the mid-dose, and the low-dose, respectively. The  respective amounts of the test substance were applied topically to the  intact, shaven skin area by means of a syringe fitted with a blunt  needle. Upon contact with the shaven skin, the test substance immediately  spread spontaneously, so covering application sites of circa 2 x 2 cm, 6  x 6 cm, and 15 x 15 cm in the low-, mid-, and top-dose group,  respectively. Immediately after application, the test site of each rabbit  was covered with porous gauze dressing fixed onto a non-irritating tape.  The entire trunk of each rabbit was wrapped with this tape to maintain  the gauze dressing in position and to retard evaporation of volatile  substances. The animals of the control group were sham-treated with the patches only.  The dermal exposure period was approximately 6 hours per day and 5 days  per week for a period of 4 weeks for the control, low-dose, and mid-dose  group. Because of the high mortality that occurred in the top-dose group  in week 3 of the study, the animals of this group were treated only  during the first 2 1/2 week of the study, i.e. 13 exposures to DMDS. 

Details on analytical verification of doses or concentrations:

Not appropriate

Duration of treatment / exposure:

0.01 and 0.1 ml/kg/d groups were treated 5 days/week during a four-week period, whereas the 1 ml/kg/d group was treated with for 2 1/2 weeks (i.e. 13 days of treatment).

Frequency of treatment:

DMDS was administered daily, by dermal occlusive application (6 hours daily) to four groups of albino rabbits.

Doses / concentrations

Remarks:

Doses / Concentrations:
0.01, 0.1, 1 ml/kg/day (10.63, 106.3 and 1063 mg/kg bw/d)
Basis:
nominal per unit body weight

No. of animals per sex per dose:

The control and 1.0 ml/kg/d group consisting of 10 males and 10 females, and the 0.01 and 0.1 ml/kg/d group consisting of 5 males and 5 females.

Control animals:

other: sham treated with the occlusive dressing

Details on study design:

After nominal day 16 of the study, the surviving animals  of the top-dose group  were kept without further treatment during the remainder of the study. 

Positive control:

Not appropriate

Examinations

Observations and examinations performed and frequency:

- Clinical signs: twice a day on exposure days and once a day on  non-exposure days.
- Mortality: twice a day.
- Dermal reactions: At the start of the study and prior to each daily administration,  individual skin reactions were evaluated in all (four) groups by the  method of Draize et al. (J. Pharmacol. Exp. Ther. 82 (1944) 377-390). In  addition, on day 28 and day 29 of the study just prior to autopsy, again  skin effects were recorded.
- Body weight: at the start of the study, twice a week thereafter, and on  the day of autopsy, i.e. on day 0, 3, 7, 10, 14, 17, 21, 24, 28, and day  29 of the study.
- Food consumption: on day 0-3, day 3-7, day 7-10, day 10-14, day 14-17,  day 17-21, day 21-24, and day 24-28.
- Ophthalmoscopic examination: no
- Blood examinatrions: On nominal day 23 for males and on nominal day 24 for females (Le. 7 and  8 days after the last treatment to DMDS for males and females of the  top-dose group, respectively), haematology and clinical chemistry  determinations were conducted in blood or plasma of the animals, which  were deprived of food for approximately 24 hours prior to the time of  blood sampling * Haematology: Hemoglobin, hematocrit, red blood cell count, white blood cell count,  differential leukocyte count, platelet count, mean cell volume, mean cell  haemoglobin concentration, mean cell haemoglobin * Biochemistry:  . Electrolytes: calcium, chloride, phosphorous, potassium, sodium,  . Enzymes: alkaline phosphatase, alanine-aminotransferase,  aspartate-aminotransferase, gamma-glutamyl-transferase  . Other: albumin, blood creatinine, blood urea nitrogen,  albumin/globulin, glucose, total bilirubin, total cholesterol, total  serum protein, bile acids
- Urinanalysis: no

Sacrifice and pathology:

On day 28 and 29 of the study (i.e. 12 and 13 days after the last  treatment to DMDS for males and females of the top-dose group), animals  were sedated by intravenous injection with Nembutaltm and subsequently  killed by opening the abdominal aorta. Next, the animals were examined  grossly for pathological changes.
- Weighted organs: adrenals, brain, heart, kidneys, liver, lungs,  ovaries, spleen, testes, thyroid and thymus.
- Microscopic examinations:  Histopathological examination was done on the skin (treated and  untreated), heart, brain, lungs, trachea, liver, kidneys, adrenals,  spleen, testes, ovaries and thymus of the animals of the control group,  the mid-dose and the high-dose groups. Examination of the bone-marrow  (sternum), popliteal, submandibular and mesenteric lymph nodes, spinal  cord and sciatic nerve were carried out in the rabbits of the control-  and the top-dose group. The skin, the thymus and the heart were also examined in the low- dose group because of possible  treatment-related effects. In addition, histopathological examination was  also done on the brain of the animals of the control group, the mid-dose,  and the top-dose group.

Statistics:

Body-weight: Covar + Dunnetts tests (two-sided)
Food intake: Anova + Dunnetts tests (two-sided)
Haematology: Anova + Dunnetts tests (two-sided) and Kruskal-Wallis Anova + Mann Whitney u-test
Blood chemistry: Anova + Dunnetts tests (two-sided)
Organ weight: Anova + Dunnetts tests (two-sided)
Histophatology: pairwise Fisher's test

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

After each daily exposure to DMDS, approximately 10-15 minutes after administration, all animals of the mid dose group were slightly lethargic whilst animals in the high dose group had distinct or severe lethargy or were even unconscious (4/10 animals on day 11). Symptoms disappeared during week-end days (d5-6 and d12-13) when no treatment occurred.
In addition, a number of high dose males and females repeatedly showed clear signs of pain (screaming) after application of DMDS. At the end of the second week, one male and three females of the high dose group showed spasms after treatment and two of the females also showed spasms during the subsequent weekend when there was no treatment. On the last exposure day for the high dose animals (study day 16) all remaining males and females showed spasms after treatment.

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

DMDS was absorbed by the skin or had disappeared otherwise within one minute after application. After removal of the patches, no remains of DMDS were present on the skin or on the gauze dressing of the covering patches.
During the four-week test period none of the controls showed any signs of irritation on the skin area treated with the control patches. Repeated dermal administration of DMDS caused severe, dose-dependent skin irritation in all dose groups.
During week 1 the following dermal effects were observed in DMDS treated animals:
- low-dose group: very slight, well-defined or moderate erythema, very slight or slight oedema, and ischemic necrosis
- mid dose group: well-defined, moderate or severe erythema, slight, moderate, or severe oedema, ischemic necrosis, and slight encrustation
- high dose group: well-defined, moderate, or severe erythema, moderate or severe oedema, ischemic necrosis, haemorrhages, and very slight, slight, or moderate encrustation.

During the second week of exposure, the treated skin of all test animals showed incrustation, which almost completely covered the treated skin area. The degree of erythema and oedema could only be scored on the small areas of the application site that did not show incrustation, or at the periphery. At the end of the second week, upon regeneration of the skin underneath, the crusts loosened. In addition, due to the inflexibility, the formed crusts cracked, especially in the animals of high dose group. Administration of DMDS to the newly formed skin underneath these cracks caused small wounds. During the third and fourth week of exposure, the severity of the encrustation in most animals of the three dose groups was such that scoring of erythema and oedema was no longer possible. After study day 16 (i.e. exposure day 13) it was decided to stop treatment of the animals of the high dose group, because of the high mortality that had occurred. Furthermore, it was decided not to record skin effects of this group on a daily basis. Instead, skin effects were recorded only on study day 21 (i.e exposure day 16) and on study day 28 or 29, just prior to autopsy in order to determine the reversibility or irreversibility of the lesions.

In the animals of the high dose group the severity of the skin effects had decreased during the recovery period. However, the 12 to 13-day recovery period was clearly not long enough to fully evaluate the reversibility or irreversibility of the dermal effects observed.

Mortality:

mortality observed, treatment-related

Description (incidence):

Two males in the control group were prematurely sacrificed due to a traumatic injury. One male in the low dose group was prematurely sacrificed due to a poor general condition. One male in the top dose group was prematurely sacrificed due to a traumatic injury. These 4 deaths were not considered to be treatment-related.
Five females and 4 males of the top-dose group were found dead. These 9 deaths were considered to be treatment-related.
Control group.
On nominal day 17 of the study, one male (A6) of the control group was found with a broken hindleg, probably because the leg had been caught between the wire mesh floor and the side panel of the cage. The animal was killed; subsequent gross observation at autopsy did not reveal any macroscopical changes.
One nominal day 24 of the study, one male (A20) of the control group was found with paralyzed hindquarters. It was decided to kill this animals. At autopsy it appeared to have a broken backbone. The cause of this injury could not be found.
Low dose group
One male (B50) of the low-dose group showed very poor defecation from nominal day 10 of the study. On day 14 of the study, the general condition of this animal was very poor, therefore, it was decided to kill the animal.
Top dose group
One nominal day 14 of the study, one male (D84) of the top dose group was found with paralyzed hindquarters. On day 15, it was decided to kill this animals. At autopsy it appeared to have a broken backbone. The cause of this injury could not be found.
One nominal day 8 of the study, one female (D91) of the top dose group was found with the lower body part paralyzed and on day 9 it was decided to kill this animals. However, at autopsy indications of a broken backbone were not found.
Five females of the top dose group were found dead between 45 minutes and 6 hours after treatment on days 8 (D89), 10 (D81), 11 (D97), 15 (D87) and 16 (D83)). Four males (D86, D94, D98 and D100) of the top-dose group were found dead on nominal day 15 at 6 hours after treatment. These deaths were considered to be treatment-related. Therefore, it was decided to no longer treat the animals of the top-dose group after day 16 of the study.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

High dose group males had reduced body weights in comparison to controls up to study day 17. Following termination of treatment after study day 16, males of this group regained weight and at necropsy, no significant differences were observed. Body weights of the low and mid dose groups were unaffected by treatment. In females, there were no clear differences in mean body weights between the groups.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

In males, food intake in the high dose group animals was somewhat reduced during the first two weeks of the study. There were no indications for a treatment-related effect on food intake of females in any of the dose groups.

Food efficiency:

not specified

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):

Treatment related haematological changes were confined to males in the high dose groups. Red blood cell count (RBC) and haemoglobin concentration (Hb) were decreased. This was accompanied by an increased mean corpuscular volume (MCV), a decreased mean corpuscular haemoglobin concentration (MCHC) and an increased number of reticulocytes (retics). Furthermore, Heinz bodies were seen in erythrocytes of most animals in this group. This group also showed a decreased number of white blood cells (WBC) due to a decrease in the number of lymphocytes (lymph).
The combination of decreased RBC and haemoglobin with an increased number of reticulocytes and the presence of Heinz bodies indicates a haemolytic anaemia, accompanied by increased erythropoiesis as compensation. The increased MCV also indicates the presence of a relatively large number of young red blood cells, which are larger than old erythrocytes. Haemolytic anaemia is also observed in ruminants fed with Brassica oleraceae (like kale, cabbage etcetera). These plants contain large amouts of S-methylcysteine sulphoxide which is converted in ruminants to DMDS (R.H. Smith, The veterinary Record, July 5, 1980, page 12-15).

The effects on the white blood cells, especially the lymphocytes in males of the high dose group may be related to the severe effects of treatment on the skin of the animals. The effects on plasma sodium and creatinine concentration in males of the high dose group may be due to their poor health.
The occurrence of these effects in males and their complete absence in females indicate either a sex-dependent difference in sensitivity or in metabolism of the test compound.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Males of the high dose group showed decreased plasma creatinine and sodium concentrations when compared with the control group. There were no treatment related findings in the mid and low dose groups. None of the findings in females were considered to be of toxicological significance.

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, treatment-related

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

The absolute and organ weights relative to brain weight measured at necropsy did not show statistically significant differences; however group mean organ weight adjusted to mean necropsy body weight was not calculated in the study report.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Macroscopy of the animals that died or were killed when moribund during the experimental period did not reveal a distinct treatment-related cause of death. At the scheduled necropsy examination, with the exception of the application sites, there were no findings that were related to DMDS treatment.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Histopathological examination identified treatment-related changes in the treated skin and the heart. The dermal lesions were seen in the treated skin of nearly all rabbits treated with topical DMDS. The changes mainly consisted of acanthosis, hyper- and/or parakeratosis, subcutaneous infiltrates of mononuclear cells and/or polymorphonuclear inflammatory cells, oedema and incidentally congestion. The severity of some of the lesions was slightly higher in the mid dose than in the low dose animals. In the high dose female rabbits that survived the experimental period, acanthosis and mononuclear cell infiltrate in the subcutis were somewhat more pronounced than in intercurrent deaths, whereas parakeratosis, subcutaneous oedema and polymorphonuclear inflammatory cell infiltrate were less pronounced. In the males of the high dose group the skin lesions of survivors and intercurrent deaths were comparable. The untreated skin of test and control animals did not reveal any abnormalities.

In the heart myocarditis and myocardial degeneration were observed in one male rabbit and in 8 female rabbits of the high dose group. This finding was most pronounced in the animals that died during the study; in severe cases it may have caused the death of the animal.

The clinically-observed neurological changes were not accompanied by histopathological changes in the brain, the spinal cord or the sciatic nerve.

Histopathological findings: neoplastic:

not examined

Other effects:

not specified

Effect levels

1

Dose descriptor:

NOEL

Remarks:

systemic toxicity

Effect level:

10.63 mg/kg bw/day

Sex:

male/female

Basis for effect level:

other: no effect

2

Key result

Dose descriptor:

NOAEL

Remarks:

systemic toxicity

Effect level:

106 mg/kg bw/day

Sex:

male/female

Basis for effect level:

clinical signs

3

Key result

Dose descriptor:

LOAEL

Remarks:

local irritation

Effect level:

8.5 mg/cm² per day (nominal)

Sex:

male/female

Basis for effect level:

dermal irritation

Target system / organ toxicity

Critical effects observed:

no

Any other information on results incl. tables

Table 1 Mortality

 

Days Dose (mg/kg bw)	Number of death (killed because moribound or found dead)
	Males	Females
0	2/10	0/10
10.6	1/5	0/5
106.3	0/5	0/5
1063	5/10	6/10

Table 2 Summary of dermal reactions in rabbits, draize scores

 

Day of test	Dose level (mg/kg)
	Males				Females
	0	10.6	106.3	1063	0	10.6	106.3	1063
Erythema
2	0	1.8	2.8	3.9	0	2.4	3.2	3.3
7	0	2.2	2.6	2.3	0	2.6	3.2	3.4
14	0	?	?	-^1	0	?	?	-^1
Autopsy (21)	0	?	?	1^2	0	2.6	?	2.25^2
Oedema
2	0	1.4	2.6	3	0	2	3	3
7	0	3	4	4	0	2.6	3.8	4
14	0	?	?	-^1	0	?	?	-^1
Autopsy (21)	0	?	?	1^2	0	2.8	?	1.5^2
Incrustation
4	0	0	0.4	1.8	0	0	0.4	1
7	0	1.8	2.8	3.7	0	1.8	2.8	3.1
14	0	3	4	-^1	0	3.4	3.8	-^1
Autopsy (21)	0	2	3.8	3^2	0	2.8	4	1.75^2

? Because of the degree of incrustation present on the entire area of the application site erythema and/or oedema could not be observed

¹animals of the high dose group were no longer treated with DMDS; skin effects not recorded

²skin reaction were recorded in order to determine the reversibility or the irreversibility of the skin lesions (4/10 females and 5/10 males examined)

Table 3 Group mean haematology (selected parameters – males- Day 23)

 

Dose mg/kg bw/day		RBC 10^12/L	Hb mmol/L	PCV l/l	MCV fl	MCH fmol	MCHC mmol/L	Retics /1000	Heinz /1000	WBC 10^9/L	Lymph 10^9/L
0	Mean sem n	5.85 0.09 9	7.9 0.1 9	0.368 0.004 9	62.9 1.0 9	1.35 0.02 9	21.4 0.2 9	31.8 5.4 9	0.0 0.0 9	11.0 0.7 9	6.4 0.4 9
10.6	Mean sem n	6.05 0.17 4	8.1 0.3 4	0.384 0.013 4	63.6 1.2 4	1.34 0.03 4	21.1 0.1 4	28.5 5.0 4	0.0 0.0 4	12.3 1.9 4	5.5 0.7 4
106.3	Mean sem n	6.26 0.26 5	7.8 0.2 5	0.365 0.009 5	58.5 1.6 5	1.25* 0.03 5	21.3 0.4 5	23.2 4.1 5	0.0 0.0 5	11.5 0.8 5	5.3 0.6 5
1063	Mean sem n	4.84** 0.24 5	6.6** 0.4 5	0.349 0.018 5	72.2** 2.0 5	1.36 0.03 5	18.9** 0.5 5	102.8*** 18.3 5	9.6 5.7 5	7.7* 0.7 5	3.6** 0.7 5

Statistics: * p<0.05, **P<0.01, ***P<0.001

sem – standard error of the mean

Table 4       Group mean clinical chemistry (selected parameters) – day 23

 

Dose mg/kg bw/day		Creatinine µmol/L	Sodium mmol/L
0	Mean sem n	133.1 4.6 9	140.6 0.4 9
10.6	Mean sem n	129.8 9.4 4	141.6 1.0 4
106.3	Mean sem n	131.5 3.1 5	141.6 0.4 5
1063	Mean sem n	108.0* 6.5 5	138.4* 0.3 5

Table 5 Incidence of selected histopathological findings - terminal kill

 

Observation   Dose (ppm)	Nasal cavity findings (10 animals/group)
	Males				Females
	0	10.6	106.3	1063	0		10.6	106.3	1063
Heart myocarditis
Very slight	0	0	0	0	0		0	0	2
Slight	0	0	0	0	0		0	0	3
Moderate	0	0	0	0	0		0	0	2
severe	0	0	0	0	0		0	0	1
Total incidence	0	0	0	0	0		0	0	8***
Myocardial degeneration
slight	0	0	0	1	0		0	0	3
Moderate	0	0	0	0	0		0	0	1

***p < 0.001

 

Applicant's summary and conclusion

Conclusions:

In this study, dermal application of DMDS resulted in treatment-related mortality in the males and females of the high dose group. In addition, dose-dependent clinical effects on the central nervous system (slight to severe lethargy) were induced by DMDS in the males and the females of the mid and high dose group. In addition, a number of high dose males and females repeatedly showed clear signs of pain (screaming) after application of DMDS. However, the observed neurological changes were of a temporary nature and were not accompanied by histopathological changes in the brain, spinal cord or sciatic nerve.

Severe dose-dependent dermal effects were observed in the males and females of all three dose groups, whereas adverse effects on body weight were only observed in males of the high dose group.
Dermal application of DMDS in this study resulted in clear effects on the red blood cell system in males of the high dose group. Microscopical examination revealed dose-dependent changes in the treated skin of the males and the females of all three dose-groups, confirming the macroscopic findings observed during the study and at necropsy. Treatment-related changes were also observed in the heart of one male and several females of the top-dose group, which died during the study or were killed at necropsy.

The no-adverse-effect level of DMDS for systemic toxicity is 106.3 mg DMDS/kg body weight/day, when applied dermally. The overall no-adverse-effect level of DMDS including local skin effects is lower than 10.63 mg DMDS/kg body weight/day.

Executive summary:

In a study performed according to the OECD guideline # 410, dimethyl disulphide was administered daily, by dermal occlusive application (6 hours daily) to four groups of albino rabbits. The dose levels applied were 0, 0.01, 0.1, and 1.0 ml/kg body weight/day, which is equivalent to 0, 10.63, 106.3, and 1063 mg/kg body weight/day, respectively. The control and 1.0 ml/kg/d group consisting of 10 males and 10 females, and the 0.01 and 0.1 ml/kg/d group consisting of 5 males and 5 females.. The animals of the 0.01 and 0.1 ml/kg/d group were treated five days a week during a four-week period, whereas animals of the 1 ml/kg/d group were treated with DMDS for 2 1/2 weeks (i.e. 13 days of treatment).

After each daily treatment with DMDS, temporary effects on the central nervous system (CNS) were observed in the animals of the 0.1 and 1 ml/kg/d group. The observed behavioural effects consisted of slight lethargy in the 0.1 ml/kg/d group and distinct to severe lethargy and unconscinousness in the 1 ml/kg/d group. At the end of each daily exposure these effects were no longer observed. During the four-week test period, treatment-related signs of abnormal behaviour were not observed in the animals of the 0.01 ml/kg/d group or in the controls. During the second and third week of the study treatment-related mortality occurred in males and females of the 1 ml/kg/d group. Therefore, it was decided to discontinue the treatment of the 1 ml/kg/d group on nominal day 16 of the study, i.e. after 13 days of treatment.

Repeated dermal administration of DMDS caused severe, dose-dependent skin irritation in all dose groups.

During the treatment period, decreased body weights were observed in males of the 1 ml/kg/d group. Food intake of males was somewhat decreased during the same period in the 1 ml/kg/d group. Mean body weight and food intake figures of the 0.01 and 0.1 ml/kg/d group were comparable with those of the controls.

Haematology and clinical chemistry examinations of the 1 ml/kg/d males, 7 days after the last exposure to DMDS, revealed treatment-related changes in several red blood cell variables and in the number of white blood cells, and treatment-related changes in plasma creatinine and sodium concentrations. In the females no effects on clinical chemistry variables were observed that could be ascribed to treatment with DMDS.

The absolute and relative organ weights measured at autopsy did not show statistically significant differences that could be ascribed to treatment. However, the mean thymus weight of both males and females tended to be lower in the two higher dose groups than in the controls.

Macroscopic examination at autopsy did not reveal any treatment-related changes other than the dermal lesions induced during the treatment with DMDS. Microscopic examination revealed treatment-related changes in the heart of males and females of the 1 ml/kg/d group and in the treated skin of all three dose-groups, confirming the macroscopic lesions observed during the study and at autopsy. No treatment-related changes were found in the brain, spinal cord, sciatic nerve, or thymus.

The dose of 1.0 ml/kg/day (1063 mg/kg bw/day) is a clear effect level and exceeds the maximal tolareted dose. The no-adverse-effect level (NOAEL) of DMDS for systemic toxicity is 0.1 ml/kg/day (106.3 mg/kg bw/day) and the NOAEL for local irritation is less than 0.01 ml/kg/day (10.63 mg/kg bw/day).