Thiram

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2 Aug - 17 Sep 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Remarks:

thionyl-14C

Species:

human

Duration of exposure:

6 h

Details on study design:

DOSE PREPARATION
- Method for preparation of dose suspensions: labelled stock solution was evaporated to dryness, dry unlabelled thiram was added and taken up with 200 mL acetone. Radioactivity was measured and the solvent was evaporated to a small volume. Aliquots were transferred to vials and formulated to give a Thiram 80WG formulation that was mixed with water to give a workable slurry.

VEHICLE
- Justification for use and choice of vehicle (if other than water): water
- Amount(s) applied (volume or weight with unit): 6.4 µL
- Concentration (if solution): high dose: 617 g/L (nominal 500 g/L); low dose: 1.46 g/L (nominal 1.6 g/L)

TEST SITE
- Area of exposure: 0.64 cm²

REMOVAL OF TEST SUBSTANCE
- Washing procedures and type of cleansing agent: swabbing with 1% Tween 80 in distilled water on cotton wool buds until no further radioactivity was removed
- Time after start of exposure: 6 h

SAMPLE COLLECTION
- Receptor fluid: collected at hourly intervals for the duration of the experiment (24 hours)
- Terminal procedure: skin samples were tape stripped to remove residual surface dose and stratum corneum

ANALYSIS
- Method type(s) for identification: LSC

Details on in vitro test system (if applicable):

SKIN PREPARATION
- Source of skin: Skin samples were obtained from human donors post mortem and were supplied by the International Institute for the Advancement of Medicine.
- Ethical approval if human skin:
- Type of skin: full thickness (see Table 1)
- Preparative technique: dermatome
- Thickness of skin (in mm): 0.2-0.4
- Membrane integrity check: with tritiated water
- Storage conditions: Skin samples were stored at ca. -20°C
- Justification of species, anatomical site and preparative technique: specified in OECD TG 428

PRINCIPLES OF ASSAY
- Diffusion cell: Scott-Dick flow-through diffusion cell, flow rate 1.5 mL/h
- Receptor fluid: 0.01 M phosphate-buffered saline, supplemented with 5% bovine serum albumin
- Solubility of test substance in receptor fluid: solubility was verified for the high-dose level
- Test temperature: 32°C
- Humidity: 41.2%-54.1% (high dose); 49.8%-56.4% (low dose)
- Occlusion: open

Absorption in different matrices:

see Table 2

Total recovery:

see Table 2

Time point:

24 h

Dose:

500 mg/mL

Parameter:

percentage

Absorption:

2.82 %

Remarks on result:

other: 24 h

Remarks:

Flux: 0.201 µg/cm²/h

Time point:

24 h

Dose:

1.6 mg/mL

Parameter:

percentage

Absorption:

11.9 %

Remarks on result:

other: 24 h

Remarks:

Flux: 0.048 µg/cm²/h

Conversion factor human vs. animal skin:

not applicable

Table 2: Summary of results

	Recovery of [^14C]-Thiram [% of dose, mean ± SD]
	Slurry of commercial formulation (500 g/L) n=6	In-use dilution (1.6 g/L) n=4
Skin wash	90.24 ± 3.54	70.88 ± 8.22
Donor compartment wash	2.22 ± 2.56	0.13 ± 0.25
Receptor fluid (0-24 h)	0.06 ± 0.02	3.77 ± 2.27
Receptor compartment wash	0.06 ± 0.00	0.04 ± 0.02
Skin^1)	nd	1.02 ± 0.97
Stratum corneum
Tape strips (1 + 2)	2.28 ± 0.73	14.49 ± 6.80
Tape strips (3 – last)	1.53 ± 1.01	2.16 ± 1.68
Absorbed dose^2)	0.12 ± 0.16	4.82 ± 3.24
Potentially absorbed dose^3)	1.65 ± 1.17	6.98 ± 4.92
Dermal absorption value	1.65 + 1.17 = 2.82 =3	6.98 + 4.92 = 11.9 =12
Total recovery	96.39 ± 1.58	92.47 ± 4.49
^1)Skin without stratum corneum ^2)The absorbed dose is the amount in receptor fluid plus receptor compartment wash plus skin (excluding tape strips, i.e. stratum corneum) ^3)The potentially absorbed dose is the amount in receptor fluid plus receptor compartment wash plus skin and tape strips (excluding the first 2 tape strips) nd: not detected

Conclusions:

The dermal absorption values for the test substance when formulated as Thiram 80 WG were estimated to be 3% and 12%, respectively, for the commercial formulation and the 1.6 g/L in-use dilution. For DNEL calculation, the result on the commercial formulation was considered.

Reference 2

Endpoint:

basic toxicokinetics

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Objective of study:

other: Absorption, distribution, metabolism and excretion

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Version / remarks:

adopted in 2010

Qualifier:

according to guideline

Guideline:

EPA OPP 85-1 (Metabolism and Pharmacokinetics)

Deviations:

no

GLP compliance:

yes

Radiolabelling:

yes

Remarks:

(14C)-test substance

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Wilmington, MA, USA (Experiment A), Charles River, Portage, MI, USA (Experiment B), Taminco Farms, Germantown, NY, USA (Experiment C)
- Age at study initiation: 6 – 8 weeks
- Weight at study initiation: means were 207.3 -223.8 g (females), 217.9 – 227.3 g (males), Experiment A, 209.6 – 246.9 g (Experiment B) 200 -250 g (Experiment C)
- Housing: individually in suspended stainless steel, screen-bottom cages, in metabolism cages for sample collection (Experiment A, B, C)
- Diet: certified Rodent Chow #5002 (Purina Mills, Inc, Experiment A, B, C), Agway Pro Lab 1000 (Agway, Waltham, MA, Experiment C), all ad libitum
- Water: tap water, ad libitum
- Acclimation period: 10-11 days (Experiment A, B)

ENVIRONMENTAL CONDITIONS (from Expeirment C)
- Temperature (°C): 20 - 30
- Humidity (%): 46 - 78
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: 10 Aug – 17 Aug 1987 (Experiment A), 23 Jul 1990 – 2 Oct 1990 (Experiment C)

Route of administration:

oral: gavage

Vehicle:

polyethylene glycol

Remarks:

400

Details on exposure:

Amount applied: 4.75 mL/kg bw, 1.7×107 dpm/mL

Duration and frequency of treatment / exposure:

Single application (Experiment A and B) or 14 unlabelled doses followed by one labelled dose (Experiment C)

Dose / conc.:

1.9 mg/kg bw/day

Remarks:

low dose, Experiment A

Dose / conc.:

125 mg/kg bw/day

Remarks:

high dose, Experiment A

Dose / conc.:

2 mg/kg bw/day

Remarks:

Experiment B (single dose) and C (for 14 days unlabelled, followed by a single dose of labelled test material)

No. of animals per sex per dose / concentration:

5 (Experiment A)
3 (males only, Experiment B)
5 (Experiment C)

Control animals:

yes

Positive control reference chemical:

No

Details on dosing and sampling:

Experiment A:
Sampling time
Urine and faeces: 4, 8, 12, 24 h, 2, 3, 4, 5, 6 and 7 days
Blood: at sacrifice
Tissues: bone (femur), brain, fat (peritoneal), gonads, heart, intestine w/contents, kidney, liver, lung, muscle (thigh), spleen, stomach w/ contents
Method: fecel samples and tissues were homogenized in distilled water, bone samples were ground. All samples were oxidized. Radioactivity was analysed with liquid scintillation counting



Experiment B:
Sampling time for expired air (trapping solution): 0, 2, 4, 6, 8, 11, 14, 16, 24, 28, 32, 48, 52, 72, 76, 80 and 96 h
Method: liquid scintillation counting, Beckman LS 9800 liquid scintillation system
Counting time: 10 min

In experiment B, the exit gases from the metabolism cages were passed through a scrubbing tower containing Harvey Carbon-14 Cocktail, then through a solution of diethylamine/ethanol (1:1) and finally through a solution of Viles CS2 reagent. The Harvey Cocktail is a very efficient trapping agent for CO2. The chemical reactivity of the cocktail would also make it an effective trapping agent for CS2 and COS, since it is capable of forming carbamates and/or thiocarbamates with CO2, CS2 or COS.

Experiment C:
Sampling time
Urine and faeces: 4, 8, 12, 24, 32, 48, 72 and 96 h
Blood and tissues: at sacrifice (96 h after administration)
Tissues: bone (femur), brain, adipose tissues, gonads, heart, kidney, liver, lung, skeletal muscle, spleen
Expired air (trapping solution): 4, 8, 12, 24, 32, 48, 72 and 96 h


Faeces were homogenized prior to measurement. Liver, kidney, heart, lung, brain, testes and spleen samples were finely minced. Adipose tissue, skeletal muscle and ovaries were not pretreated for combustion, blood cells were vortexed. Bone marrow was flushed from the bone prior to combustion.
Method: Tracor Analytic Mark III scintillation system.

Experiment D
Urine samples analysed
From Experiment B: 24 h samples from the low (1.9 mg/kg bw) and high (125 mg/kg bw) dose
From Experiment C: 4, 8, 12 and 24 h
For analyzation, samples from animals from the same sex and dose were pooled.

Method: HPLC with a LDC/Milton Roy CM4000 pump, LDC spectroMonitor variable wavelength UV detector and a FLO-ONE radioactivity detector. The analytical column was a Baxter Burdick & Jackson C-18. (5 µm, 4.6 x 250 mm) with a Brownlee precolumn (C-18, 7 µm). Initial mobile phase was 100% phosphoric acid buffer, increased to 30% acetonitrile , increased to 100% acetonitrile.

The amount of radioactivity in the HPLC fractions was analysed with Beckman LS 9800 liquid scintillation system.

Preliminary studies:

Total recoveries of radioactivity were 32.1 and 27.0% for males and females of the single low dose group, respectively, not accounting for volatile residues. In the single high dose group, the recovery (without volatiles) was 35.7% (♂) and 32.0% (♀). In the repeated-dose study (Experiment C), volatile residues were accounted for. Accordingly, the recoveries were much higher with 85.4% (♂) and 92.9% (♀) (see Table A6_2-1 in the attached background material).

Type:

absorption

Results:

Based on the the high excretion via air and urine (total 83 - 87%), high absorption is assumed, but no specific experiments were conducted.

Type:

excretion

Results:

[14C]-test substance-derived radioactivity was excreted in the urine (ca. 35—40% of the dose within 96 hr.), faeces (Ca. 2-5%) and expired air (ca. 47-48%). Most of the radioactivity was excreted in the first 12 hours after dosing.

Type:

distribution

Results:

Distributions in tissues were low, the highest percentage of the dose was seen in blood (0.9-1.99 %), liver (0.4-1.8%), muscle (0.05-0.6%), bone (± 0.5%) and kidney (0.07- 0.16%).

Type:

metabolism

Results:

The test substance is extensively and almost completly metabolized to more polar products, diethylthiocarbamic acid being formed as the principal metabolite. Other metabolic products include carbon disulfide, methyldiethyldithiocarbamate, and sulfate.

Details on absorption:

No designated experiments were conducted to examine the absorption of the test susbtance. However, the majority of the applied radioactivity is either excreted in urine or exhaled (total 83-87%, see Table A6_2-1 in the attached background material). This demonstrates a high degree of absorption.

Details on distribution in tissues:

Four to seven days after oral exposure to single or multiple doses, trace levels of test-substance-derived radioactivity were detected in all tissues. Total recovery was low (from 1.8 to 4.2%): the highest percentage of the dose was seen in blood (0.9-1.99 %), liver (0.4-1.8%), muscle (0.05-0.6%), bone (± 0.5%) and kidney (0.07- 0.16%). All other tissues, in both sexes, contained less than 0.1% of the dose. (Table A6_2-2 under "Any other informations on results incl. tables"). This would indicate that the test substance did not accumulate in the rat upon repeated dose administration.

Details on excretion:

The majority of the dose (at least 83.7% for males and 89.6% for females) was eliminated from the body within four days following dosing (Table A6_2-1 under "Any other informations on results incl. tables").
As far as urinary and faecal excretion is concerned, the following data were obtained.

After a single dose, maximal urinary excretion occurred within 4 to 24 h and accounted for 6% and 14 % for a low or high dose respectively. Faecal excretion was low, occurring within 48 and 96 h and representing approximately 3.7% of the dose. Seven days after exposure, total urinary and faecal excretion ranged from 24 to 34% of the administered dose, mostly present in urine. There were no significant differences in total recovery between dose levels or between sexes. In these experiments, a majority of the material was unaccounted for (Table A6_2-3 under "Any other informations on results incl. tables").

After repeated oral administration of a low dose, urinary excretion became more important, approximately 33-35%. Faecal excretion was low, only 0.5% of the dose. No significant differences were found between male and female. (Table A6_2-4 under "Any other informations on results incl. tables"). While urinary excretion trends to increase with the dose, faecal excretion decreased. These effects are in agreement with the quantitative differences occurring in the metabolism of thiram suggesting saturation of a metabolic route.

Total 14C test substance-derived exhaled radioactivity over 96 h was 47.0 and 48.2% for male and female rats respectively. Exhaled radioactivity was collected in KOH traps (likely as CO2 and CS2), mainly within the first 12 hours, and in Viles CS2 traps (likely CS2), mainly excreted within the first 8 hours. The extent and rate of exhalation of 14C test substance-derived radioactivity were similar in male and female rats. These data also indicate that the test substance is rapidly metabolized by the rat. After multiple doses, exhalation is slightly lower (Table A6_2-5 under "Any other informations on results incl. tables").

Metabolites identified:

yes

Details on metabolites:

In general, the metabolism of dialkyldithiocarbamates such as the test substance in mammals is straightforward, diethylthiocarbamic acid being formed as the principal metabolite. Other metabolic products include carbon disulfide, methyldiethyldithiocarbamate, and sulfate.
The metabolic decomposition of ethylene bisdithiocarbamates such as nabam, maneb and zineb is complex and results in the formation of carbon disulfide, hydrogen sulfide, ethylene diamine, ethylene bisthiuram disulfide, 5,6-dihydro-3H-imidazo(2,1-C)-1,2,4-dithiazole-3-thione, ethylene diisocyanate, ethylene thiourea, ethylene urea and 2-imidazoline (Table A6_2-6 under "Any other informations on results incl. tables").
The test substance is extensively metabolized in rats to more polar products. The metabolic pathway involved a reduction of the disulfide bond and subsequent reactions of the thiol moiety to form oxidative and conjugative polar products. No unchanged test substance was detected in the urine.
Among the five metabolites identified in the urine, the alanine conjugate of dimethyldithiocarbamate is one of the major metabolites, occurring both after single or repeated, and low or high dose administration.
Quantitative differences occurred for the other metabolites: dithiocarbamate thiosulfenic acid was the second major metabolite after subchronic or high dose, while in the low dose animals 2-thioxo¬thiazolidine-4-carboxylic acid was the second major metabolite.
Three additional urinary metabolites, P1, P2 and P3, were not identified. P1 and P2 constitute together less than 5% of the radioactivity present in the urine of subchronically treated rats. P3 constituted 12-21% of the radioactivity present in urine of low dose animals. None of these three were present in the urine of high dose rats.
Sexual difference occurred in the metabolism after low dose exposure: males had almost twice as much P3 as the females, whereas females had ten times more dithiocarbamate thiosulfenic acid than the males (Table A6_2-6 under "Any other informations on results incl. tables").
After subchronic exposure, a major portion of the dose (ca.47-48%) was exhaled in air. It is likely that radioactivity collected in the KOH trap was 14CO2 and/or 14CSO (carbamyl sulfide, carbon thioxide), since it has been reported that CSO is soluble in KOH. Radioactivity collected in the Viles CS2 traps is likely to be CS2, as it was reported earlier that CS2 is a metabolite of the test substance. (Table A6_2-4 under "Any other informations on results incl. tables").

Summarized results of Experiment A, B, C and D can be found in Attachment 1, 2, 3 and 4 in the attached background material, respectively. An overview of relevant results can be found below. 

 

Table A6_2-1:   Recovery of radioactivity [%] in rats following single oral administration of (^14C)-test substance*
Sample [hour]	50 ppm		500 ppm		1000 ppm
	Male	Female	Male	Female	Male	Female
Urine
0-6	8.09	17.26	11.60	25.69	12.03	15.54
6-12	0.00	3.01	2.14	7.65	4.37	0.79
12-24	5.44	2.06	2.94	3.93	3.45	4.65
24-48	0.73	0.79	0.48	1.01	0.66	0.43
48-72	0.37	0.46	0.18	0.36	0.28	0.31
72-96	0.24	0.30	0.17	0.25	0.17	0.20
Total Urine	14.87	23.88	17.51	38.89	20.96	21.93
Faeces
0-6	0.00	0.01	0.01	0.00	0.02	0.00
6-12	0.02	0.00	0.01	0.00	0.00	0.00
12-24	5.80	3.82	2.14	0.77	0.93	0.12
24-48	2.31	2.45	1.52	1.30	1.16	0.82
48-72	0.28	0.56	0.27	0.31	0.66	0.46
72-96	0.07	0.18	0.06	0.07	0.22	0.17
Total Faeces	8.47	7.00	3.99	2.45	2.99	1.57
Cage
Wipes	0.05	0.14	0.02	0.08	0.01	0.06
Ringe	1.21	3.24	1.83	1.23	2.27	5.85
Wash	0.07	0.16	0.05	0.10	0.06	0.13
Total Cage	1.33	3.54	1.89	1.40	2.34	6.04
Total Elimination	24.66	34.42	23.38	42.74	26.28	29.54
Tissue
Muscle	0.28	0.56	0.10	0.37	0.24	0.25
Fat	0.04	0.08	0.03	0.04	0.03	0.03
Lung	0.03	0.05	0.02	0.04	0.02	0.02
Spleen	0.01	0.01	0.01	0.02	0.01	0.01
Kidney	0.11	0.13	0.06	0.08	0.06	0.06
Liver	0.97	0.87	0.55	0.50	0.55	0.62
Stomach	0.02	0.03	0.02	0.02	0.01	0.00
Stomach content	0.03	0.06	0.03	0.05	0.03	0.04
Intestine	0.01	0.01	0.00	0.00	0.00	0.00
Intestine content	0.02	0.03	0.02	0.03	0.08	0.07
Blood	0.85	1.35	0.73	1.55	0.74	0.91
Total Tissue	2.35	3.15	1.54	2.68	1.76	2.01
Total Recovery**	27.01	37.56	24.92	45.42	28.04	31.55
*   Data are presented as group mean values with n = 2 for each group except females - 1000 ppm where n =1 ** Total recovery does not include volatile^14C-residues in expired air which was not collected in this study.

 

Table A6_2-2:    Tissue distribution of radioactivity from ^14C-test substance
dose - mg/kg bw (frequency)	1.9 (1 x)		2 (15 x)		125 (1x)
day of sacrifice after treatment	7	7	4	4	7	7
sex	♂	♀	♂	♀	♂	♀
bone	0.55	0.48	-	-	0.476	0.42
blood	0.975	1.632	-	-	1.236	1.99
plasma + blood cells	-	-	0.04+0.5	0.04+1.43	-	-
brain	0.0075	0.0076	0.01	0.01	0.01	0.01
fat	0.015	0.0084	0.06	0.08	0.014	0.022
gonads	0.01	0	0.01	0.00	0.01	0
heart	0.03	0.045	0.02	0.03	0.016	0.014
intestine + content	0.097	0.127	-	-	0.072	0.078
kidney	0.127	0.165	0.07	0.10	0.048	0.068
liver	1.862	1.479	0.69	0.94	0.44	0.376
lung	0.065	0.077	0.03	0.04	0.03	0.038
muscle	0.087	0.136	0.39	0.58	0.05	0.11
spleen	0.01	0.01	0.01	0.03	0.008	0.018
stomach + content	0.032	0.065	-	-	0.016	0.024
total	3.867	4.23	1.83	3.27	2.42	3.168

 

Table A6_2-3:    Mean urinary and faecal excretion as a function of time after a single low or high dose
Dose [mg/kg bw]	Urine				Faeces
	1.9		125		1.9		125
Sex	♂	♀	♂	♀	♂	♀	♂	♀
Time [h]
0	0	0	0	0	0	0	0	0
4	5.46	6.48	0.95	1.07	0.13	0	0	0.188
8	3.6	3	0.58	0.95	0.022	0	0	0
12	3.8	2.7	2.21	2.27	0.19	0.465	0.16	0.168
24	2.4	2.8	10.8	8.65	1.18	1.01	0.204	0.28
48	2.2	1.19	10.4	8.12	1.56	1.62	0.77	0.492
72	0.84	0.79	2.31	3.4	0.54	0.49	1.08	0.244
96	0.66	0.42	0.86	1.11	0.25	0.482	0.56	0.606
120	0.48	0.5	0.7	0.62	0.18	0.288	0.39	0.3
144	0.26	0.47	0.85	0.33	0.114	0.354	0.35	0.238
168	0.17	0.44	0.59	0.43	0.128	0.242	0.29	0.278
total	19.87	18.79	30.25	26.95	4.3	4.95	3.8	2.79

 

 

Table A6_2-4:          Excretion of radioactivity after 2 doses: comparison with excretion after 1 dose
Dose [mg/kg bw]	Urine						Faeces
	1.9			125			1.9		125
Sex		♂	♀		♂	♀	♂	♀	♂	♀
Time [h]
0		0	0				0	0
4		5.46	6.48		16.3	16.4	0.13	0	0.1	0
8		3.6	3		6.4	6.4	0.022	0	0	0
12		3.8	2.7		4.2	4.4	0.19	0.465	0	0.3
24		2.4	2.8		4.2	4.4	1.18	1.01	0.3	0.2
48		2.2	1.19		0.6	0.6	1.56	1.62	0	0.1
72		0.84	0.79		0.9	1.4	0.54	0.49	0.9	1.8
96		0.66	0.42		0.4	0.9	0.25	0.482	0.3	0.8
120		0.48	0.5		0.3	0.6	0.18	0.288	0.9	2.1
144		0.26	0.47				0.114	0.354
168		0.17	0.44				0.128	0.242
total		19.87	18.79		33.3	35.2	4.3	4.95	2.6	5.3

 

Table A6_2-5:          Exhalation of volatile ^14C [% of administered dose]
Dose [mg/kg bw]	1.9 (2 rats)	2.0
frequency	1 x	15 x
Time [h]
2	9.08
4	16.31	24.3
6	9.79
8	6.13	15.6
11	7.98
14	3.78
16	1.01
24	4.10	1.7
28	1.105
32	0.605	0.3
48	1.165
52	0.25
56	0.095
72	0.395	0.3
76	0.115
80	0.085
96	0.337	0.2
total	62.86	47.0

 

Table A6_2-6:          Metabolites of ^14C-test substance in rat urine [% of radioactivity in urine]
Dose [mg/kg bw]	1.9		125		2 (x 15)
Sex	♂	♀	♂	♀	♂	♀
Metabolites
2-thioxothiazolidine-4-carboxylic acid	33.6	30.2	12.6	7.8	2.4	2.1
dimethyldithiocarbamate glucuronide	3.2	4.2	5.6	8.7	6.6	6.9
dimethyldithiocarbamate thiosulfenic acid derivative	0.9	9.1	37.3	42.4	44.4	48.1
dimethyldithiocarbamate ethyl ester	ND	ND	3.4	2.4	0.8	0.8
dimethyldithiocarbamate alanine	34.8	42.1	33.9	36.5	41.8	38.3
P1 unknown	ND	ND	ND	ND	1.3	1.8
P2 unknown	ND	ND	ND	ND	2.8	2.1
P3 unknown	20.8	12.2	ND	ND	ND	ND

Conclusions:

The test substance, studied at doses between 2 and 125 mg/kg bw in the rat, is well absorbed from the GI tract. It is eliminated, for most part within 24 hours, mainly via urine (33%) and exhaled air (47%), and does not accumulate in the organism. 96 hours after dosing, the highest tissue levels are found in blood, liver, muscle, bone and kidney, representing about 3% of the dose administered. It is extensively metabolised, ~85% of the administered dose was detected as metabolites in urine, faeces (5%) and expired air. The metabolic pathway involves a reduction of the disulfide bond and subsequent reactions of the thiol moiety to form oxidative and conjugative polar products. No unchanged test substance was detected in the urine.
In general, the metabolism of dialkyldithiocarbamates such as the test substance in mammals is straightforward, diethylthiocarbamic acid being formed as the principal metabolite. Other metabolic products include carbon disulfide, methyldiethyldithiocarbamate, and sulfate.

Reference 3

Endpoint:

basic toxicokinetics

Type of information:

experimental study

Adequacy of study:

key study

Study period:

3 Aug - 9 Oct 1987

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Objective of study:

distribution

excretion

Qualifier:

according to guideline

Guideline:

EPA OPP 85-1 (Metabolism and Pharmacokinetics)

Deviations:

yes

Remarks:

The amount of test substance should be measured until about 95% of the administered dose is excreted or for seven days. Distribution was not studied as a function of time.

GLP compliance:

yes

Radiolabelling:

yes

Remarks:

14C

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Charles River Laboratories, Portage, MI, USA
- Age at study initiation: 5 weeks
- Weight at study initiation: means were 132.4 – 142.4 g (males), 137.6 – 139.4 g (females)
- Housing: individually in suspended stainless steel, screen-bottom cages
- Diet: certified Rodent Chow #5002 (Purina Mills, Inc.ad libitum
- Water: tap water, ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.6 – 23.9
- Humidity (%): 20 - 70
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12/12
- Fasting period

IN-LIFE DATES: From: 3 Aug 1987 To: 9 Oct 1987

Route of administration:

other: oral: diet (9-week period)/gavage (single application)

Vehicle:

polyethylene glycol

Details on exposure:

DIET PREPARATION (non- radiolabelled)
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with: basal diet
- Storage temperature of food: room temperature

The radiolabelled test material was dissolved in polyethylene glycol 300 (PEG300).

Duration and frequency of treatment / exposure:

Repeated unlabelled doses for 9 weeks followed by one labelled dose.

Dose / conc.:

10.25 other: µCi

Remarks:

single dose, adminsitered to all animals of all dose groups after the subchronic feeding period.

Dose / conc.:

50 ppm

Remarks:

9-week study period

Dose / conc.:

500 ppm

Remarks:

9-week study period

Dose / conc.:

1 000 ppm

Remarks:

9-week study period

No. of animals per sex per dose / concentration:

2

Control animals:

yes

Positive control reference chemical:

No

Details on study design:

Post Exposure Period: 96 h

Clinical signs and mortality were examined daily.

Body weight was determined pre-dose, at study initiation, weekly through week 8 and before and 96 h after the animals was given the radio-labelled test material.

Food consumption was monitored weekly from study initiation through week 8.

Samples of urine, faeces, blood and cage rinse were taken. Sampling time (0 h = start of application): Urine and faeces: 0-6, 6-12, 12-24, 24-48, 48-72, 72-96h; Blood: 1, 2, 4, 8, 24, 48, 72, 96h; Cage rinse: 24 h-intervals

Pathology: 96 h after receiving the radiolabelled substance;
Tissues: Fat (perigonadal), intestine and contents, kidney, liver, lung, muscle (thigh), spleen, stomach and contents

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, blood, plasma, serum or other tissues, cage washes
- Time and frequency of sampling: Samples of urine, faeces, blood and cage rinse were taken. Sampling time (0 h = start of application): Urine and faeces: 0-6, 6-12, 12-24, 24-48, 48-72, 72-96h; Blood: 1, 2, 4, 8, 24, 48, 72, 96h; Cage rinse: 24 h-intervals

ANALYTICAL METHOD
For analysis of radioactivity in tissues of the rats after sacrifice and in feces, the samples were homogenized with deionized water. No sample preparation was needed for stomach contents, intestine contents and blood samples.

Total 14 C-residues in tissues and feces were determined by liquid scintillation counting after oxidative combustion.

Cage wipes were analysed by adding methanol to the cage wipes for extraction (5 h).

Liquid scintillation counting: Beckman LS 9800 liquid scintillation system
Counting time: 10 min
Limit of sensitivity: twice the background counting rate (here 232 dpm/g)

Cage wash, cage rinse and urine were also analysed without sample preparation.
Liquid scintillation counting: Beckman LS 7500 liquid scintillation system
Counting time: 5 min

Preliminary studies:

The total recovery of 14C-test substance equivalents as a percent of the administered dose in tissue and excreta for all groups and sex are summarized in Table A6_2-1 in the attached background material. For females, an average of 38.2% of the administered dose was recovered in the tissues and excreta, whereas, for the males, this was about 26.7% of the administered dose. Approximately 76% of total 14C-recovered was found in the urine. Faecal recovery was substantial only in animals dosed at 50 ppm. In a separate study (Gay et al., 1991), it was found that about 61% of the administered 14C-test substance was eliminated in the expired air. In the present study, expired air was not collected and there fore, substantial loses were expected. If the 61% is added to the total observed recovery in the current study, the calculated total recovery is about 87% for the males and about 99% for the females.

Type:

absorption

Results:

Experiments were conducted to examine the absorption of the test substance (area under curve), but this part of the report is not presented by the applicant.

Type:

distribution

Results:

Tissues contained 1.2 - 3.8% of the dose, mainly in liver and kidney. Blood contained circa 1% of the dose.

Type:

metabolism

Results:

Metabolites were not examined.

Type:

excretion

Results:

The levels of 14C-residues in the faeces peaked between 12-48 h after administration, the levels in urine after 6 h. Females excreted more radioactivity via urine than males.

Details on absorption:

Experiments were conducted to examine the absorption (area under the curve), but this part of the report is not presented by the applicant.

Details on distribution in tissues:

For all groups, the liver contained the highest concentration of 14C-residues, followed by the kidneys and the lungs (see Table A6_2-1 in the attached background material). Altogether the tissues contained only 1.2-3.8% of the dose for all treated groups whereas the blood contained about 1% of the dose.

Details on excretion:

The levels of 14C-residues in the faeces peaked between 12-48 h after administration. The total recoveries from the 50 ppm dose group (average 7.7%) were more than twice from those seen in animals receiving 500 ppm (average 3.2%) or 1000 ppm (average 2.5%).
The levels of 14C-residues in the urine peaked at 6 h after dose administration for all dose groups in both sexes. Thereafter excretion rates declined rapidly and were virtually complete at 24 h. Females eliminated substantially more 14C-Thiram equivalents in urine than did males.

Metabolites identified:

not measured

Remarks:

A suggested metabolic pathway can be found under "Illustrations".

Enzymatic activity measured:

not measured

There were no treatment-related antemortem observations. One female in the 1000 ppm group died due to technical error during the 4 h-postdose sample collection. Body weight (gain) and food consumption were lower throughout the study in the 500 and 1000 ppm group.

 

Summarized results of Experiment A, B and C can be found in Attachment 2, 3 and 4 in the attached background material, respectively. An overview of relevant results extracted from the study report can be found in Attachment 1. 

Conclusions:

The studies were conducted similar to guidelines and under GLP conditions. The test substance was excreted mainly via expired air with the liver being the tissue which contains the highest concentration of 14C-test substance residues.

Description of key information

The test substance is well and rapidly absorbed from the gastro intestinal tract. The majority of the dose is eliminated from the body within four days after dosing, for most part within 24 hours, either via urine (~34%) or the expired air (~48%), and does not accumulate in the organism.
The highest tissue levels of the test substance are found in blood and liver, independent of the administered dose and the elapsed time, but the amount of radioactivity in this tissues was low in all studies.
The test substance is rapidly degraded by rats to more polar products. The urine, which held ca. 30% of the [14C]-test substance-derived radioactivity, contained virtually no unchanged [14C]-test substance. Five urinary metabolites were detected and isolated by HPLC and identified by mass spectrometry.
The dermal absorption values for the test substance when formulated as 80 WG were estimated to be 3% and 12%, respectively, for the commercial formulation and the 1.6 g/L in-use dilution.
Thus, 3% is the assumed dermal absorption for the neat test substance.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Absorption rate - oral (%):

100

Absorption rate - dermal (%):

3

Additional information

Toxicokinetics

Toxicokinetic behavior of the test substance was investigated in several GLP-compliant studies in rats, which were both conducted similar to OECD 417 and are considered as key studies.

 

After single oral and repeated-dose administration of the radiolabelled test substance to rats, no absorption studies were performed but based on the high excretion via air and urine (total 83 - 87%), high absorption is assumed.

Distribution was rapid but no bioaccumulation in tissues was observed. The highest percentage of the dose was seen in blood, liver, muscle, bone and kidney. The highest percentage of radioactivity in tissues was a total of 4.24%. Metabolism of absorbed doses is extensive. The most important fraction of the test material was metabolised into volatile compounds excreted in air. These volatile metabolites, collectively accounted for ca 47-48% of the administered dose, and consisted of CS2, (COS) and CO2. The principle route is hydrolysis to dimethyldithiocarbamate, which further decomposed to CS2, COS and CO2 exhaled as volatile metabolites or conjugated. In urine, only trace amounts of the parent compound were detected, suggesting complete metabolisation of the test material. The metabolic pathway in the rat is characterised by the reduction of disulfide bound leading to dimethyl dithiocarbamic acid (DDC). DDC is further oxidized mainly to dimethyl-thiosulfenic acid after high doses, or to thioxothiazolidine carboxylic acid after low doses. The metabolite M1, dimethylamino-4-thiazoline carboxylic acid, has also been identified in the urine of rats after treatment with a single oral dose of the test substance. 

Excretion via urine was less compared to expired air and excretion via faeces was minor. Following oral administration, the majority of radioactivity was excreted as volatiles (CS2, CO2) in expired air (up to approximately 48%) within 24 h. Excretion via urine was 35 – 40%, in feces the maximum was 7.7%. Lower doses of test substance administration. After repeated exposure, recovery in faeces was higher than for the high dose group. After repeated oral administration of a low dose, urinary excretion became more important, approximately 33-35%. Faecal excretion was low, only 0.5% of the dose. The rate of elimination was relatively fast; the majority of the radioactivity was eliminated within 96 h after dosing

 

 

Dermal Absorption

Dermal absorption was investigated in vitro on human (Report number: PFX0008/053512). The study was conducted according to GLP and OECD guideline 428. Following topical application of a low (1.6 g mg/mL) and a high concentration (500 mg/mL) of the 14C-labelled test material to human skin, the total absorbed dose was 11.9% and 2.82% of the applied dose, respectively after 24 h.