Absorption, Distribution and Excretion
Five Cr1:CD BR VAF/Plus rats (half male and half female) were administered 2 mg/kg of folothiazolamide by gavage (using 2 mL/kg corn oil as a carrier) daily for 14 consecutive days. Subsequently, the rats received 2 mg/kg of labeled folothiazolamide and were sacrificed at 24 hours or 168 hours, respectively, and tissues (including blood) were collected for radiolabeling analysis. Urine, feces, and exhaled gases were collected periodically after administration. The purity of unlabeled folothiazolamide was 99%, and the purity of ring-labeled folothiazolamide was >99%. Pretreatment had no significant effect on metabolism…excretion patterns and tissue residues over time were similar to those of a single dose. Folothiazolamide is rapidly absorbed, metabolized, and excreted. Urine is the primary route of excretion, while feces and exhaled gases are secondary routes. No sex-related differences in excretion pathways or rates were observed. Excretion was essentially completed within 24 hours after administration. At 24 hours, 75% to 82% of the administered dose was eliminated. 168 hours after the experiment ended, 70% and 13.2% of the administered dose were excreted in urine and feces, respectively. Approximately 5.3% was excreted as carbon dioxide. Five Cr1:CD BR VAF/Plus rats (half male and half female) were administered a single dose of 2 mg/kg or 20 mg/kg folothiazole ester via gavage (dissolved in 2 mL/kg corn oil). Unlabeled folothiazole ester had a purity of 99%, and ring-labeled folothiazole ester had a purity >99%. Excremental and exhaled gas samples were collected during the 7-day post-exposure observation period. Marker analysis was performed on major tissues at the end of the experiment. Regardless of sex or dose level, exhaled gas contained 5–6% of the administered dose (presumably CO₂ in a 1 N NaOH trap). Most exhaled gas was collected within the first 12 hours after administration. The concentration of volatile organic compounds collected downstream of the CO₂ trap was extremely low. Urinary metabolites accounted for 66-67% of the administered dose in males and 71-73% in females. No significant differences were observed between dose levels. In all cases, more than half of the urinary markers were collected within 6 hours of administration. Fecal metabolites accounted for 11-12% of the administered dose regardless of sex or dose. Most fecal markers were collected within 48 hours of administration. The percentage of total administered dose detected in all body tissues was 11.0% in male rats in the 2 mg/kg/day group and 9.4% in male rats in the 20 mg/kg/day group; for female rats, it was 8.2% and 6.8%, respectively, indicating significant tissue retention of the drug. One week after treatment, tissue markers were widely distributed. The highest concentrations of markers in the 2 mg/kg group were found in the liver, lungs, and heart. Radioactive material was not concentrated at the site of drug entry (gastrointestinal tract), in the circulatory system (blood), or in adipose tissue. This significant tissue retention, coupled with the large amount of carbon dioxide produced from labeled carbon metabolism, indicates that a large amount of carbon from the Fosthiazate ring is absorbed into the body's carbon pool. …
Ten Cr1:CD BR VAF/Plus rats (each sex) were administered unlabeled Fosthiazate once daily by gavage (dissolved in 2 mL/kg corn oil). On day 15, the rats were given 2 mg/kg of S-sec-butyl-labeled Fosthiazate. Five rats from each sex were sacrificed 24 hours after administration, and the remaining five rats from each sex were sacrificed 7 days later. …Pretreatment had no significant effect on distribution and excretion patterns. In this study, the distribution of biomarkers in rats over 7 days (expressed as a percentage of the administered dose) was as follows: urine (73% and 74% in males and females, respectively), feces (8% and 9% in males and females, respectively), exhaled carbon dioxide (9% and 8% in males and females, respectively), exhaled volatile organic compounds (0.5% and 0.6% in males and females, respectively), and the sum of all tissues (1.8% and 1.2% in males and females, respectively). ... At sacrifice on day 7, the radioactivity of all sampled organs did not exceed twice that of whole blood. Tissue samples were taken 24 hours after sacrifice (excluding the gastrointestinal tract, as the labeled intestinal contents did not have sufficient normal transit time). The liver showed the highest radioactivity ratio relative to whole blood (4.4 in male rats and 2.3 in female rats), followed by the lungs (2.0 in male rats and 1.6 in female rats). The kidneys and adrenal glands showed slightly lower radioactivity ratios than the lungs. The radioactivity ratios of most organs and tissues were similar to or lower than those of whole blood. ...

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Metabolism/Metabolites
Seven Cr1:CD BR VAF/Plus rats (half male and half female) were administered 18 mg/kg of folothiazole ester via gavage (2.5 mL/kg corn oil solvent) as a single dose. The unlabeled folothiazole ester had 50% of the carbon atoms on the butyl methyl group labeled with 13C, with a purity of 99.3%. The purity of S-sec-butyl-2-14C-labeled flothiazolyl ester was 97.7% (mass spectrometry analysis). Researchers assessed the metabolism of flothiazolyl ester in tissues and excrement over 48 hours and characterized the major metabolites in urine. The labeling distribution of the S-sec-butyl-2-14C-labeled group in excrement in this study was largely consistent with the results of another study using ring-labeled 14C. This indicates that both the ring and butyl substituents undergo degradation, releasing significant amounts of CO2 (approximately 10% of the administered dose). Identified high-molecular-weight metabolites (excluding glutathione products) underwent ring-opening reactions, typically accompanied by methylation of the cyclic sulfur. Oxidation products of this cyclic sulfur, such as sulfonic acids, sulfoxides, and sulfones, accounted for approximately 20% of the administered dose. Several S-sec-butyl hydrolysis residues were observed, which also exhibited sulfur oxidation, with or without methylation. Some S-sec-butyl residues were glutathione-bound and existed as N-acetylcysteine products (not further characterized). ...
Before identifying the major metabolites, seven Cr1:CD BR VAF/Plus rats (half male and half female) were administered 22 mg/kg cyclo-14C-Fosthiazate by gavage once (dissolved in 2.2 mL/kg corn oil). ...Nine peaks were observed migrating near the solvent front, forming the largest cluster of radiolabeled components, but none were characterized. Based on their migration on the reversed-phase HPLC column and the minimal effect of the basic ion pair reagent on their migration, these peaks were considered to be small molecules, polar, and significantly uncharged. These metabolites accounted for 42% of the male dose and 27% of the female dose. Considering the significant release of labeled carbon dioxide, the large amount of labeled residue retained in tissues, and the presence of many small molecular weight labeled residues in urine, the cyclic carbon appears to have been largely assimilated into the carbon library. Three metabolites retained the S-butyl substituent and underwent ring-opening of the thiazolyl ring. These metabolites account for approximately 7% of the administered dose in men and approximately 18% in women. The most abundant is (RS)-S-sec-butyl O-ethyl N-(2-methanesulfonylethyl)thiophosphoramide ester (BESxP). Some similar metabolites lack the S-sec-butyl group; notably, O-ethyl S-hydro N-2-(methanesulfonyl)ethylthiophosphoramide ester (DBSoS) (4.6% of the administered dose in men and 3.0% in women). Acetamide (a rat liver carcinogen, at concentrations approximately 1000 times higher than those observed in this study) accounted for 3% and 2% of the administered dose in men and women, respectively. ...
Phospoxonase (PON1) is a key enzyme in organophosphate metabolism. PON1 can inactivate certain organophosphates through hydrolysis. PON1 hydrolyzes active metabolites in various organophosphate pesticides and nerve agents such as soman, sarin, and VX. The presence of PON1 polymorphism leads to differences in the enzyme level and catalytic efficiency of this esterase, which in turn suggests that different individuals may be more susceptible to the toxic effects of organophosphate exposure.

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Biological Half-Life
Five Cr1:CD BR VAF/Plus rats (half male and half female) were administered 2 mg/kg or 20 mg/kg of folothiazol ester via gavage (2 mL/kg corn oil carrier) as a single dose. The purity of unlabeled folothiazol ester was 99%, and the purity of ring-labeled folothiazol ester was >99%. …The median estimated initial half-life (defined as time to peak plasma concentration within 12 hours after administration) for male and female rats in the low-dose group was 14.9 hours and 11.5 hours, respectively. The estimated initial half-life for both male and female rats in the high-dose group was 8.6 hours. In the later stage from 18 to 168 hours, the median half-life for males and females in the 2 mg/kg dose group was 76 hours and 66 hours, respectively; the median half-life for males and females in the 20 mg/kg dose group was 92 hours and 87 hours, respectively.

Toxicity Summary
Acibenzolar are cholinesterase or acetylcholinesterase (AChE) inhibitors. Cholinesterase inhibitors (or "anticholinesterases") inhibit the activity of acetylcholinesterase. Because acetylcholinesterase plays a vital physiological role, chemicals that interfere with its activity are potent neurotoxins; even low doses can cause excessive salivation and lacrimation, followed by muscle spasms and ultimately death. Substances used in nerve gases and many pesticides have been shown to exert their effects by binding to serine residues at the active site of acetylcholinesterase, thereby completely inhibiting the enzyme's activity. Acetylcholinesterase breaks down the neurotransmitter acetylcholine, which is released at the neuromuscular junction, causing muscle or organ relaxation. The mechanism of action of acetylcholinesterase inhibitors is to allow acetylcholine to accumulate and exert its sustained effect, ensuring the continuous transmission of nerve impulses and preventing muscle contraction from ceasing. The most common acetylcholinesterase inhibitors are phosphorus-containing compounds designed to bind to the enzyme's active site. Its structural requirements include a phosphorus atom with two lipophilic groups, a leaving group (such as a halogen or thiocyanate group), and a terminal oxygen atom.

[1]. Chiral organophosphorous pesticide fosthiazate: absolute configuration, stereoselective bioactivity, toxicity, and degradation in vegetables. J Agric Food Chem. 2020 Jul 22;68(29):7609-7616.

Fosthiazate is a phosphonate, organophosphonate, and organothiophosphate insecticide. It can be used as an EC 3.1.1.7 (acetylcholinesterase) inhibitor, an agricultural chemical, and a nematicide. There are reports and data regarding Fosthiazate in capparis spinosa. Fosthiazate belongs to the organophosphate insecticide or nematicide class and is used to control nematodes on tomatoes.

C9H18NO3PS2

283.35

283.046

98886-44-3

91758

White to off-white <25°C powder,>25°C liquid

1.3±0.1 g/cm3

371.3±25.0 °C at 760 mmHg

178.3±23.2 °C

0.0±0.8 mmHg at 25°C

1.541

0.94

0

5

6

16

301

0

CCC(C)SP(=O)(N1CCSC1=O)OCC

DUFVKSUJRWYZQP-UHFFFAOYSA-N

InChI=1S/C9H18NO3PS2/c1-4-8(3)16-14(12,13-5-2)10-6-7-15-9(10)11/h8H,4-7H2,1-3H3

3-[butan-2-ylsulfanyl(ethoxy)phosphoryl]-1,3-thiazolidin-2-one

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

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.)