Absorption, Distribution and Excretion
Eight-week-old male and female ICR mice were orally administered 5 mg/kg body weight of [(14)C-4-methyl]-tocrofus-methyl (radiochemical purity 99%) dissolved in corn oil, and the radiocarbon content in urine, feces, and exhaled breath was monitored for 7 days post-administration. Within 24 hours, 69-76% of the administered radiolabeled material was excreted in urine, 4-6% in feces, and less than 1% in exhaled breath. Seven days after administration, the total residual radiocarbon in the body was less than 1% of the administered dose. Six-week-old Sprague-Dawley rats were orally administered 5 mg/kg body weight of [(14)C-4-methyl]-tocrofus-methyl (radiochemical purity 99%) dissolved in corn oil, and the radiocarbon content in urine, feces, and exhaled breath was monitored for 7 days post-administration. Within 24 hours, 62-67% of the administered dose was excreted in urine, 16-21% in feces, and less than 1% in exhaled air. Seven days after administration, the total residual radiocarbon in the body was less than 1% of the administered dose. Whole-body autoradiography performed 1 hour and 6 hours after treatment showed the highest accumulation of radiolabeled material in the stomach and intestines, followed by the kidneys and liver. Male and female Sprague-Dawley rats were given a single oral dose of 5 or 200 mg/kg body weight of tocrofos-methyl, a compound uniformly labeled with 14C on the benzene ring (radiochemical purity >99%). Another group of animals were given unlabeled tocrofos-methyl orally for 14 consecutive days at a dose of 5 mg/kg body weight, followed by a single oral dose of 5 mg/kg body weight of [14C-phenyl]-tocrofos-methyl. The administered radiocarbon was readily excreted, with over 95% of the dose excreted in urine and feces within 48 hours. Seven days later, 85-91% of the drug was excreted in urine. Fecal excretion at that time was as follows: continuous dose group, 9.3% in men and 12% in women; low dose group, 20% in men and 19% in women; high dose group, 20% in men and 12% in women. Less than 0.1% of the administered dose was excreted as 14C-carbon dioxide in all groups. Peak 14C concentrations were reached in almost all tissues within 2 hours. Following low-dose administration, the highest 14C concentrations were observed in the kidneys; expressed as tocrofos methyl equivalents, the 14C concentration in male kidney tissue was 4700 ng/g and in female kidney tissue was 3450 ng/g; plasma 14C concentrations were 1140 ng/mL in men and 1270 ng/mL in women. The concentration of (14)C in the liver of male rats was 1240 ng/g tissue, and in the liver of female rats it was 1220 ng/g tissue; the concentration of (14)C in the blood of male rats was 736 ng/mL, and in the blood of female rats it was 835 ng/mL. 72 hours after administration, the concentration of (14)C in each organ was ≤ 5% of its respective peak concentration. After 7 days, the residual radioactivity was less than 1% of the administered dose. In male and female rats with bile duct cannulation, the cumulative excretion of (14)C within 48 hours was: 5.8-12% in bile, 47-59% in urine, and 42-24% in feces.

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Metabolism/Metabolites
Male ICR mice were orally administered 5 mg/kg body weight of [(14)C-4-methyl]-tocrofus-methyl; metabolites were isolated from feces and urine by chromatography and identified by cochromatographic and/or spectroscopic analysis with standards. Major metabolites detected in excrement included: 2,6-dichloro-4-methylphenol (9% of the dosage label), O,O-dimethyl-O-(2,6-dichloro-4-carboxyphenyl) phosphate (11%), O-methyl-O-hydro-O-(2,6-dichloro-4-carboxyphenyl) phosphate (12%), 3,5-dichloro-4-hydroxybenzoic acid (12%), and 3,5-O-dichloro-4-hydroxybenzylglycine (13%). The main biotransformation reactions included: oxidative desulfurization to produce oxidized compounds and their derivatives; oxidation of 4-methyl to produce alcohols and acids; cleavage of PO-aryl and PO-methyl bonds; and conjugation of the resulting acid with glycine. Metabolites found in mice were similar to those found in rats, except for 3,5-O-dichloro-4-hydroxybenzylglycine. Male and female Sprague-Dawley rats were orally administered 5 or 200 mg/kg body weight of toclofos-methyl, labeled with 14C at uniformly distributed sites on the 4-methyl or benzene ring. Some rats were pretreated with unlabeled toclofos-methyl at 5 mg/kg body weight/day for 14 consecutive days. Metabolites were isolated from feces, urine, bile, and major tissues by chromatography and identified by co-chromatographic and/or spectroscopic analysis with standards. More than 10 metabolites were detected in excrement. No significant differences were observed related to sex or dose. The major metabolites detected in excrement were O-methyl-O-hydro-O-(2,6-dichloro-4-methylphenyl) phosphate (10-26% of urine (14)C), O-methyl-O-hydro-O-(2,6-dichloro-4-hydroxymethylphenyl) thiophosphate (12-25%), O-methyl-O-hydro-O-(2,6-dichloro-4-carboxyphenyl) thiophosphate (11-35%), and O-methyl-O-hydro-O-(2,6-dichloro-4-methylphenyl) thiophosphate (12-44%). In cannulated rats, most of the radiolabeled substances excreted into bile within 24 hours after administration were associated with polar metabolites; the major metabolites in bile were O-methyl-O-hydro-O-(2,6-dichloro-4-hydroxymethylphenyl) thiophosphate and 2,6-dichloro-4-methylphenol glucuronide. Radiocarbons excreted in feces within 24 hours of administration were associated only with the parent compound. Two hours after oral administration, the major metabolites in the blood, liver, and kidneys were O,O-dimethyl-O-(2,6-dichloro-4-carboxyphenyl)thiophosphate, 3,5-dichloro-4-hydroxybenzaldehyde, O-methyl-O-hydro-O-(2,6-dichloro-4-methylphenyl)thiophosphate, and O-methyl-O-hydro-O-(2,6-dichloro-4-hydroxymethylphenyl)thiophosphate. Only trace amounts of the parent compound were detected in the liver. The main biotransformation reactions were oxidative desulfurization to produce oxidized compounds and related derivatives, 4-methyl oxidation to produce alcohols and acids, cleavage of PO-aryl and PO-methyl bonds, and the conjugation of the resulting acids and phenols with glucuronic acid.

Toxicity Summary
Identification and Uses: Methyltocrophos is an agricultural fungicide. Human Studies: In in vitro human estrogen activity assays, methyltocrophos induced a weak response. In in vitro non-programmed DNA synthesis assays in human cancer cells (HeLa), the results were negative. Animal Studies: In rabbits, methyltocrophos showed minimal eye irritation and no skin irritation. In guinea pigs, it did not cause skin sensitization. Acute exposure to methyltocrophos in rats, mice, and dogs resulted in decreased spontaneous movement, dyspnea, piloerection, urinary incontinence, and ataxia. These symptoms resolved completely by day 10. In dogs receiving a dose of 1000 mg/kg body weight, brain cholinesterase activity was lower after 16 days of treatment compared to dogs receiving lower doses. In dogs at a concentration of 2000 ppm, plasma cholinesterase activity in female dogs decreased by 19–26% throughout the study period, but no significant decrease in erythrocyte cholinesterase activity was observed in either male or female dogs, or in plasma cholinesterase activity in male dogs. Brain cholinesterase activity was unaffected by treatment. In rabbits administered transdermally, erythrocyte cholinesterase activity in male rabbits was lower than in the control group, but no dose-response relationship was observed. In animals at doses of 300 and 1000 mg/kg body weight/day, plasma cholinesterase activity in both male and female dogs was lower than in the control group (reduced by 22–29%). In female dogs at a dose of 1000 mg/kg body weight/day, relative kidney weight increased by 20%. At doses up to 3000 mg/kg body weight/day, tocrofos-methyl was not teratogenic in rabbits but was toxic to female rabbits. At doses up to 50 mg/kg body weight/day, tocrofos-methyl showed no embryotoxicity, fetal toxicity, or teratogenicity in rats. In vitro reverse mutation assays against Salmonella Typhimurium TA98, 100, 1535, 1537, and 1538 were all negative, regardless of metabolic activation. In vivo genotoxicity studies in rats and mice were also negative.

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Non-human toxicity values
Oral LD50 in male rats: approximately 5000 mg/kg
Dermal LD50 in male rats: >5000 mg/kg
Intraperitoneal LD50 in male rats: approximately 5000 mg/kg
Subcutaneous LD50 in male rats: >5000 mg/kg
For more complete non-human toxicity data for tocrofos-methyl (18 in total), please visit the HSDB record page.

Methyl tocrophos is an organothiophosphate ester, chemically named 2,6-dichloro-4-methylphenol, in which the hydrogen atom on the hydroxyl group is replaced by a dimethoxyphosphothiophosphate group. Methyl tocrophos is a phospholipid biosynthesis inhibitor and fungicide used to control soil-borne diseases caused by Typhula incarnata, Corticium rolfsii, Typhula ishikariensis, and Rhizoctonia solani. It is an antifungal pesticide. It is an organothiophosphate ester and a dichlorobenzene compound.

C9H11O3PSCL2

301.12664

299.954

57018-04-9

91664

White crystals from methanol
Colorless crystals
White cyrstalline solid

1.4±0.1 g/cm3

338.5±52.0 °C at 760 mmHg

78-80°C

158.5±30.7 °C

0.0±0.7 mmHg at 25°C

1.563

4.03

0

4

4

16

260

0

S=P(OC)(OC1=C(Cl)C=C(C)C=C1Cl)OC

OBZIQQJJIKNWNO-UHFFFAOYSA-N

InChI=1S/C9H11Cl2O3PS/c1-6-4-7(10)9(8(11)5-6)14-15(16,12-2)13-3/h4-5H,1-3H3

(2,6-dichloro-4-methylphenoxy)-dimethoxy-sulfanylidene-λ5-phosphane

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ≥ 36 mg/mL (~119.55 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)