Target: Protoporphyrinogen oxidase (PPO). Pyraflufen-ethyl acts as a contact herbicide. Upon application, it is absorbed by plant foliage and inhibits PPO, leading to the accumulation of protoporphyrin IX, which generates reactive oxygen species (ROS) upon light exposure, causing rapid cell membrane disruption and desiccation of susceptible weeds.

In vitro, Pyraflufen-ethyl inhibits PPO enzyme activity extracted from plant chloroplasts, preventing the conversion of protoporphyrinogen IX to protoporphyrin IX. No specific in vitro activity in mammalian cell systems is reported for the parent compound, though it is highly effective at low application rates (grams per hectare) in plants.

In vivo (in plants), Pyraflufen-ethyl is rapidly absorbed by foliage and translocated within the plant. It causes rapid desiccation and necrosis of treated tissues, with visible symptoms (wilting, browning) appearing within 1-2 days of application. It is selective, controlling broad-leaved weeds and grasses in crops.

For cell-free PPO inhibition assay: plant chloroplasts or etioplasts are isolated from susceptible weed species and homogenized. The PPO enzyme is extracted and incubated with varying concentrations of Pyraflufen-ethyl and the substrate protoporphyrinogen IX in a 96-well plate. Fluorescence is measured (excitation 410 nm, emission 630 nm) to detect accumulated protoporphyrin IX. IC50 is calculated from the dose-response curve.

No mammalian cell-based assays are typically performed for this herbicide, as it is not intended for human therapeutic use. However, in plant cell suspension cultures, Pyraflufen-ethyl induces rapid cell death and electrolyte leakage from protoplasts, confirming its herbicidal mechanism of action.

For in vivo animal studies: Pyraflufen-ethyl is not used therapeutically in animals. Ecotoxicological studies are conducted on aquatic and terrestrial organisms (fish, daphnia, bees) to assess environmental safety. Mammalian toxicity studies in rats are required for regulatory registration but are not conducted for pharmacological research.

Absorption, Distribution and Excretion
Following a single or multiple oral dose of 5 mg/kg Pyraflufen-ethyl, it is rapidly absorbed and excreted within 96 hours (plasma half-life of 3 to 3.5 hours). However, absorption saturates at doses up to 500 mg/kg, and the Cmax values do not reflect a 100-fold dose difference (2.7 to 2.8 Fg eq/g in the low-dose group and 100 to 107 Fg eq-hr/g in the high-dose group). Following a single or multiple oral dose of low-dose (5 mg/kg) Pyraflufen-ethyl, urinary excretion accounts for 27% to 33% of the administered radioactivity, indicating that multiple dosing regimens do not affect the absorption/excretion process. After a single 500 mg/kg dose, urinary excretion decreases to only 5% to 7%. In all treatment groups, the remaining radioactive material is excreted in feces. Bile excretion analysis following a single 5 mg/kg dose showed that 36% of the administered dose appeared in bile. Based on excretion data, the overall bioavailability of the low-dose group was approximately 56%. Due to the lack of bile excretion data for the high-dose group, its bioavailability could not be definitively assessed. No sex difference was observed in excretion patterns. However, plasma and blood clearance were faster in women than in men, as evidenced by the plasma/blood radioactivity time course and higher AUC values in men (32.3 vs 18.4 Fg eq-hr/g for the low-dose group and 2,738 vs 1,401 Fg eq-hr/g for the high-dose group). Radioactivity concentrations showed that at 96 hours post-administration, radioactivity concentrations in all tissues were at or near the limit of detection (generally <0.01 Fg eq/g and never exceeding 0.02 Fg eq/g). Therefore, Pyraflufen-ethyl and its metabolites did not appear to have significant retention. Tissue load data after compound administration did not reveal specific targets other than the liver and kidneys (tissues associated with the absorption and elimination of orally administered exogenous substances). [EPA 40 CFR Part 180]

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Metabolism/Metabolites
Five Sprague-Dawley rats of each sex were administered 5 mg/kg of [phenyl-U-14C]ET-751 via a single gavage. Each animal was immediately transferred to a glass metabolic cage after administration. Urine and feces were collected every 24 hours for 96 hours. Metabolites in urine and feces were evaluated using high-performance liquid chromatography (HPLC) and two-dimensional thin-layer chromatography (2-D TLC). Metabolites were identified in urine and feces. Similar to [pyrazole-5-14C]ET-751, the hypothesized metabolic pathway is ester hydrolysis and N-demethylation of the pyrazole ring at position 1, followed by hydrolysis of the ether bond at position 5 of the benzene ring to phenol, and methylation to methoxy group. Twenty male Sprague-Dawley rats were administered unlabeled ET-751 technology via gavage for 14 consecutive days at a dose of 5 mg/kg/day. On day 15, they were given a single gavage dose of [pyrazole-5-14C] ET-751 at a dose of 5 mg/kg. Metabolites in urine, feces, plasma, and gastrointestinal contents were identified using high-performance liquid chromatography (HPLC). After 96 hours, the total values in urine and feces were 26.66% and 64.44%, respectively. …After 96 hours, the concentrations in all samples were below or equal to the limits of detection. Metabolites in urine, feces, and plasma were identified…The hypothesized metabolic pathway is ester hydrolysis and N-demethylation at the 1-position of the pyrazole ring, and hydrolysis of the ether bond at the 5-position of the benzene ring to generate phenol followed by methylation to generate a methoxy group. Six male Sprague-Dawley rats were administered a single oral gavage dose of 5 mg/kg of [Pyrazole-5-14C] ET-751 via bile duct cannulation. Metabolites were identified using high-performance liquid chromatography (HPLC). Within 48 hours of administration, 17.90% (feces), 19.66% (urine), and 36.09% (bile) of the radioactive material were excreted. After 48 hours, 13.86% of the radioactive material was present in the gastrointestinal contents, and 2.86% remained in the gastrointestinal tract. The estimated oral absorption rate was 55.75% (total urine and bile). Metabolites were identified in urine, feces, the gastrointestinal tract, and gastrointestinal contents. The hypothesized metabolic pathway involves ester hydrolysis and N-demethylation at the 1-position of the pyrazole ring, and hydrolysis of the ether bond at the 5-position of the benzene ring to generate a phenolic hydroxyl group. Five male and five female Sprague-Dawley rats in each group were injected with 5 or 500 mg/kg of [pyrazole-5-14C]ET-751 and sacrificed at 3, 6, 9, 24, 96, or 168 hours later, respectively. ...Metabolites were identified in urine, plasma, and feces...The hypothesized metabolic pathway is ester hydrolysis and N-demethylation at the 1-position of the pyrazole ring, and hydrolysis of the ether bond at the 5-position of the benzene ring to generate phenol, followed by methylation to generate a methoxy group.

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PK properties in mammals: after oral administration, Pyraflufen-ethyl is rapidly absorbed and extensively metabolized, primarily by ester hydrolysis to its free acid form (pyraflufen), followed by conjugation. The parent compound has low systemic bioavailability, and excretion occurs mainly via feces and urine.

Toxicity profile: Pyraflufen-ethyl has low acute oral toxicity in rats (LD50 > 5000 mg/kg). Chronic exposure may cause long-term adverse effects in the aquatic environment. It is not classified as a human carcinogen or mutagen. Eye and skin irritation may occur upon direct contact.

[1]. Parthenium hysterophorus L. control in response to pyraflufen-ethyl application. Crop Protection, Volume 57, March 2014, Pages 35-37.

Pyraflufen-ethyl is an ethyl ester formed by the condensation of the carboxyl group of pyrflufenicol and ethanol. As a prodrug of pyrflufenicol, it is used to control broadleaf and grassy weeds in various crops. It has multiple functions, including acting as an EC 1.3.3.4 (protoporphyrinogen oxidase) inhibitor, a prodrug, and an agrochemical. It belongs to the pyrazole, biaryl, ethyl ester, aromatic ether, monochlorobenzene, and monofluorobenzene classes of compounds. Its function is similar to that of pyrflufenicol. Pyraflufen-ethyl is a selective contact herbicide used to control broadleaf and grassy weeds in crops.

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Pyraflufen-ethyl is a registered herbicide, not a human pharmaceutical. It is approved for agricultural use in many countries (including the US EPA and EU) for post-emergence control of broadleaf weeds in cereals, corn, and soybeans. It is not FDA-approved for human use. The compound is also used as an analytical standard for pesticide residue testing.

C15H13N2O4F3CL2

413.17592

383.989

129630-19-9

182951

Light yellow to light brown solid powder

1.6±0.1 g/cm3

468.9±40.0 °C at 760 mmHg

126-127 °C

237.4±27.3 °C

0.0±1.2 mmHg at 25°C

1.575

2.73

0

8

8

26

480

0

APTZNLHMIGJTEW-UHFFFAOYSA-N

InChI=1S/C15H13Cl2F3N2O4/c1-3-24-11(23)6-25-10-4-7(9(18)5-8(10)16)13-12(17)14(22(2)21-13)26-15(19)20/h4-5,15H,3,6H2,1-2H3

ethyl 2-[2-chloro-5-[4-chloro-5-(difluoromethoxy)-1-methylpyrazol-3-yl]-4-fluorophenoxy]acetate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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

DMSO : ~100 mg/mL (~242.03 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (6.05 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 2: ≥ 1.25 mg/mL (3.03 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 12.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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