CYP51A(Kd=0.5 nM)

In this study, the DMI fungicide mefentrifluconazole exhibited excellent inhibitory activity against the mycelial growth of S. rolfsii, with a mean EC50 value of 0.21 ± 0.11 mg L-1 and a range of 0.02 to 0.55 mg L-1 for 261 isolates collected from Hebei, Henan and Shandong provinces. Mefentrifluconazole significantly reduced the biomass of mycelia and affected the morphology of hyphae. Although sclerotia were more tolerant to mefentrifluconazole than mycelial growth, mefentrifluconazole greatly inhibited the formation and germination of sclerotia. In addition, sclerotia produced by mefentrifluconazole-treated mycelia were deficient in nutrients (e.g., protein, carbohydrate and lipid). These results indicated that mefentrifluconazole may reduce the population of S. rolfsii in the following year. In greenhouse experiments, mefentrifluconazole showed control efficacy and good persistence against peanut S. rolfsii. The preventative and curative activities of mefentrifluconazole at 200 mg L-1 against southern blight still reached 95.36% and 60.94%, respectively, after 9 days of application. No correlation was observed for the sensitivity of S. rolfsii to mefentrifluconazole and the tested DMI, quinone outside inhibitor and succinate dehydrogenase inhibitor fungicides. Conclusion: All data indicated that mefentrifluconazole could provide favorable control efficacy against S. rolfsii from peanuts and reduce the infection and population of S. rolfsii in the following year. © 2023 Society of Chemical Industry[2].

Mefentrifluconazole is put through a rigorous toxicity testing regimen that includes numerous studies on reproductive toxicity. Studies on the long-term repeated dose toxicity and/or carcinogenicity in rats, mice, and dogs have been carried out. Systemic toxicity occurs at the highest dose level that has been studied in each species[1]. In the mefentrifluconazole acute and repeat dose toxicity studies. Rats receiving a single dose have an LD50 of greater than 2000 mg/kg bwt when administered orally, more than 5000 mg/kg bwt when applied topically, and more than 5.314 mg/L when inhaled as dust aerosol. Mefentrifluconazole is not a phototoxicant in vitro, nor is it irritating to the skin or eyes[1]. Mefentrifluconazole (oral administration; 2000 mg/kg bwt; single dose) reduces body weight gain and transient neurobehavioral effects only on the day of treatment (unsteady gait, decreased motor activity, decreased forelimb grip strength, and increased distance between the hind limbs in the landing foot-splay test) in the acute neurotoxicity study conducted in rats[1]. For all three of the species under investigation, the liver is the target organ in the repeated-dose toxicity studies. Higher doses in rats (oral diets; 383/334 mg/kg/bwt/d (4000 ppm)) and mice (C57BL/6JRj; 61 mg/kg bwt/d (300 ppm)) cause changes in clinical chemistry parameters, reduce food intake and body weight gain, and increase liver weight, which is accompanied by either liver cell necrosis or liver cell hypertrophy. Increases in liver weight are thought to be an adaptive response to treatment when they occur at low doses and are not linked to any histopathological changes[1].

In vitro screening assays for candidate selection [1]
The screening procedure used for candidate selection included computer modelling of binding to fungal CYP51 and human CYP19, glasshouse efficacy studies, in vitro binding experiments with Zymoseptoria tritici CYP51 (ZstCYP51), and in vitro aromatase inhibition assays using human recombinant CYP19 supersomes. The methodology of the CYP51 and CYP19 assays is briefly described below. [1]
ZstCYP51was expressed in E. coli and the recombinant protein subsequently purified. Azole binding to the heme iron of ZstCYP51 was measured in the absence of reductase by difference spectroscopy as a “type II” spectrum characterized by a maximum at ∼430 nm and a minimum at ∼410 nm. Each binding experiment usually consisted of eight wavelength scans in the 400–450 nm range with rising concentration of azole, against a DMSO control. The delta absorption was plotted against the azole concentration and the binding constant Kd was determined by nonlinear regression. [1]
The aromatase (CYP19) inhibition assay was conducted utilizing the recombinant human aromatase (Corning Supersomes Human CYP19 aromatase + reductase) and the fluorometric artificial substrate dibenzylfluorescein (DBF) (Sigma D-7191) as described by Stresser et al. (2000). Cofactors in this assay were 1.3 mM NADP+ , 0.4 mU Glucose-6-phosphat-dehydrogenase, 3.3 mM Glucose-6-phosphat and 3,3 mM MgCl2. Test compounds were dissolved in DMSO (final DMSO concentration 1%). Enzymatic reaction was started with enzyme/substrate mix (4 pmol/ml enzyme, 0.4 μM DBF) and incubated for 30 min at 37 °C. The reaction was stopped by addition of 75 μl 2 M NaOH and incubated for another 2 h at 37 °C. Measurement was conducted at 490 nm excitation wavelength, 530 nm emission wavelength and 515 nm cut-off. Dose response analyses were made using a log-logistic 4-parameter model. More recently, the above method was adapted for assessing the inhibition of rat aromatase utilizing recombinant rat aromatase (Corning Rat CYP19 + P450 Reductase SUPERSOMES™). [1]
The results from the CYP in vitro assays were compared against marketed azole data (CYP19 and CYP51 data from in-house experiments and published toxicological information). As a conclusion from this screening exercise, a tentative threshold human aromatase inhibition IC50 value of 1 μM was defined for compound candidates for consideration of promotion to field testing and in vivo regulatory studies. Additionally, a quotient of target(CYP51) and off-target(CYP19) inhibition was calculated for candidate and market agrochemical azoles and used as selection criterion.

Rabbit [1]
\nFour groups each of 27–33 nulliparous female New Zealand White rabbits (age range 10–15 weeks on receipt) were acclimatized for 5 days and then injected with 0.2 ml of a synthetic gonadotrophin releasing hormone (Receptal®). Approximately one hour later the females were inseminated with semen from stock male rabbits of the same source and strain. The day of insemination was designated gestation day (GD) 0. Bodyweights of females on GD 0 ranged between 2747 and 3819 g. Administration of mefentrifluconazole (MFZ) by oral gavage commenced on GD 6 and continued once daily until GD 28. Dose levels were 0, 5, 15 and 25 mg/kg bwt/day as a suspension in 1% w/v aqueous carboxymethylcellulose, at a dose volume of 10 ml/kg bwt. The high dose level in this study was set at half the lethal dose, as determined from the results of a preliminary study in non-pregnant female rabbits, in which severe toxicity was observed at 50 mg/kg bwt/day and two out of three animals at this dose were killed in extremis. At 150 mg/kg bwt/day and above all females died or were killed in extremis. [1]
\n\nThroughout the study, clinical signs and food consumption were monitored daily and body weights were recorded at 2- to 3-day intervals. Prior to termination on GD 29 blood samples were taken from the marginal ear vein of each female for assessment of hematology and clinical chemistry parameters. On GD 29 all surviving females were killed by intravenous injection of pentobarbital (Narcoren® 2.0 ml per animal). The females were necropsied in random order, the uterus and ovaries were removed from each female and the intact uterus was weighed. All other gestational parameters were evaluated “blind” viz. numbers of corpora lutea, implantation sites, live and dead fetuses, early and late resorptions. Individual fetuses and placentae were weighed, and all fetuses were examined externally. Heads were removed from approximately half of the fetuses from each litter, fixed in Bouin's fluid and sectioned and examined, according to the method of Wilson (1965). The torsos and intact fetuses were examined internally, hearts and kidneys were sectioned and evaluated, and fetal sexes were recorded. Following examination, all fetuses and torsos were eviscerated, and the skeletons were processed using a modified method of Kimmel and Trammell (1981), prior to examination of ossified and cartilaginous structures.\n

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\n\nRat [1]
\nNulliparous female Wistar rats (CRL:WI(Han)) were mated by the breeder. Mating was confirmed by vaginal plug/sperm-positive vaginal smear and the day of mating was designated GD 0. The mated females were shipped and received at the test facility on the same day. Body weight on GD 0 ranged between 144.2 and 185.7 g. On arrival, the females were randomly assigned to four groups of 25 females. Administration of mefentrifluconazole (MFZ) by oral gavage commenced on GD 6 and continued once daily until GD 19. Dose levels were 0, 50, 150 and 400 mg/kg bwt/day as a suspension in 1% w/v aqueous carboxymethylcellulose, at a dose volume of 10 ml/kg bwt. The high dose level was selected based upon the results from a 14-day dose range-finding study in non-pregnant rats, where oral administration of 250 and 500 mg/kg bwt/day gave rise to reductions in food consumption and body weight gain, and from a dose-range finding study in pregnant rats in which the NOAEL was found to be > 200 mg/kg bwt/day. On GD 20 the females were killed by cervical dislocation under isoflurane anesthesia. They were necropsied in random order; the uterus and ovaries were removed from each female and the intact uterus was weighed. All other gestational parameters were evaluated “blind”, viz. numbers of corpora lutea, implantation sites, live and dead fetuses, early and late resorptions. Any uterus or single uterine horn with no obvious implantation sites was stained using the Salewski technique (1964). Individual fetuses and placentae were weighed, and all fetuses were examined and sexed externally. Approximately half of the fetuses from each litter were fixed in Harrison's fluid and were subsequently examined according to the Barrow and Taylor method (1969). The remaining fetuses from each litter were eviscerated and processed using a modified method of Kimmel and Trammell (1981), prior to examination of ossified and cartilaginous skeletal structures.\n

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\n\nTwo-generation reproduction toxicity study in rats [1]
\nThe study was conducted according to OECD TG 416 (2001). Wistar rats (Crl:WI(Han), aged 28 days ± 1 day on arrival were acclimatized for approximately 6 days and were then randomly assigned on a body weight basis to four groups of 25 males and 25 females per group to form the F0 generation. Males and females were derived from different litters and animals were housed individually except during pairing or during lactation, when the female and litter were housed together until weaning on postnatal day (PND). Environmental conditions were similar to those of the prenatal studies. In addition, nesting material was provided to pregnant females from late gestation. At commencement of treatment males were in the weight range 97.5–123.0 g and females 90.3–112.2 g. [1]
\nmefentrifluconazole (MFZ) was administered via the diet at target dose levels of 0, 25, 75 and 200 mg/kg bwt/day. To achieve these target doses the concentration of mefentrifluconazole (MFZ) in the diet was adjusted weekly according to the animals’ group mean food intake, except during gestation when the females received the same dietary concentration as the last pre-mating week and during lactation when the concentration was reduced by 50% to allow for the increase in food intake. Food consumption and body weights were recorded weekly except during the mating periods. During lactation, food consumption was recorded for PND 1–4, 4–7, 7–14, and 14–22. The F0 and F1 animals were continuously exposed for a minimum of 75 days prior to mating until the study was terminated at weaning of the F1 or F2 litters.

Mefluconazole fungicide for poultry (Colinus virginianus, 22 weeks 14 days): LD50 816 mg/kcal
Mefluconazole fungicide for poultry (Colinus virginianus, ErlyLf, 21 weeks): LOEL 531 ppm
Mefluconazole fungicide for fish (Oncorhynchus mykiss, 1.7 g, 96-hour LC50 536 ppm
Mefluconazole fungicide for fish (Danio rerio, 0.3 g, 96-hour LC50 823 ppm
Mefluconazole fungicide for crustaceans (Daphnia pulex LifCyc, 21-day LOEC 40.6 ppm)

[1]. Innovative selection approach for a new antifungal agent mefentrifluconazole (Revysol®) and the impact upon its toxicity profile. Regul Toxicol Pharmacol. 2019 Aug;106:152-168.

[2]. Bioactivity of the DMI fungicide mefentrifluconazole against Sclerotium rolfsii, the causal agent of peanut southern blight. Pest Manag Sci. 2023 Jun;79(6):2126-2134. d

2-[4-(4-chlorophenoxy)-2-(trifluoromethyl)phenyl]-1-(1H-1,2,4-triazol-1-yl)prop-2-ol is an aromatic ether, a diphenyl ether, substituted at position 1 with 2-hydroxy-1-(1H-1,2,4-triazol-1-yl)prop-2-yl, at position 2 with trifluoromethyl, and at position 4 with chlorine. It belongs to the monochlorobenzene, triazole, tertiary alcohol, aromatic ether, and (trifluoromethyl)benzene classes. Mefenofibrate (trade name: Revysol®) is the active ingredient of a novel isopropanol-triazole fungicide with highly selective bactericidal activity. Comprehensive toxicity testing conducted according to OECD guidelines has shown that mefenofibrate (MFZ) is non-genotoxic and non-carcinogenic. Repeated-dose studies in rats, mice, and dogs have identified the liver as the primary target organ. Prenatal developmental toxicity studies in rats and rabbits showed no treatment-related embryotoxicity or teratogenicity at the highest tested dose levels. In a two-generation diet study in rats, the high-dose group showed reduced food intake and weight gain throughout the administration period. Mating ability and fertility, estrous cycles, gestation, and prenatal and postnatal survival of offspring were largely unaffected, and there was no evidence of masculinization of female pups or feminization of male pups. The screening strategy aimed to find candidate drugs that had both high bactericidal activity and minimized adverse side effects caused by aromatase inhibition, and MFZ was ultimately selected. The selection strategy for MFZ was confirmed by the absence of any endocrine disruption potential in toxicity studies. [1]

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Background: The pathogen of southern peanut blight, Sclerotium rolfsii, is becoming increasingly prevalent and causing serious damage in China, resulting in severe economic losses to the peanut industry. To effectively control southern peanut blight, this study evaluated the bioactivity of the new-generation sterol demethylation inhibitor (DMI) fungicide mefenoxam against Phytophthora insicae. Results: This study showed that the DMI fungicide mefenoxam exhibited excellent inhibitory activity against Phytophthora insicae mycelial growth. The average EC50 value of 261 strains collected from Hebei, Henan, and Shandong provinces was 0.21 ± 0.11 mg L⁻¹, ranging from 0.02 to 0.55 mg L⁻¹. Mefenoxam significantly reduced mycelial biomass and affected mycelial morphology. Although sclerotia were more tolerant to mefenoxam than mycelia, mefenoxam still significantly inhibited sclerotia formation and germination. Furthermore, the sclerotia produced by mefenoxam-treated mycelia were deficient in nutrients (such as protein, carbohydrates, and lipids). These results suggest that mefenoxam may reduce the population size of Phytophthora insicae in the following year. Greenhouse experiments showed that mefenflurazole had good control and persistence against Phytophthora indicum in peanuts. At a concentration of 200 mg L⁻¹, after 9 days of application of mefenflurazole, the prevention and treatment rates against Phytophthora indicum reached 95.36% and 60.94%, respectively. No correlation was observed between the sensitivity of S. rolfsii to mefenflurazole and the tested DMI, quinone-based inhibitors, and succinate dehydrogenase inhibitors. Conclusion: All data indicate that mefenflurazole has good control effect against S. rolfsii in peanuts and can reduce the infection rate and population size of S. rolfsii in the following year. © 2023 China Chemical Industry Association. [2]

C18H15CLF3N3O2

397.78

397.08

C, 54.35; H, 3.80; Cl, 8.91; F, 14.33; N, 10.56; O, 8.04

1417782-03-6

71230671

Off-white to light yellow solid powder

4.65

1

7

5

27

491

0

ClC1C([H])=C([H])C(=C([H])C=1[H])OC1C([H])=C([H])C(=C(C(F)(F)F)C=1[H])C(C([H])([H])[H])(C([H])([H])N1C([H])=NC([H])=N1)O[H]

JERZEQUMJNCPRJ-UHFFFAOYSA-N

InChI=1S/C18H15ClF3N3O2/c1-17(26,9-25-11-23-10-24-25)15-7-6-14(8-16(15)18(20,21)22)27-13-4-2-12(19)3-5-13/h2-8,10-11,26H,9H2,1H3

2-(4-(4-chlorophenoxy)-2-(trifluoromethyl)phenyl)-1-(1H-1,2,4-triazol-1-yl)propan-2-ol

Mefentrifluconazole; BAS 750 F; BAS 750F; 1417782-03-6; Revysol; 2-(4-(4-Chlorophenoxy)-2-(trifluoromethyl)phenyl)-1-(1H-1,2,4-triazol-1-yl)propan-2-ol; D9EYN9N7UI; Mefentrifluconazole [ISO]; 2-[4-(4-chlorophenoxy)-2-(trifluoromethyl)phenyl]-1-(1,2,4-triazol-1-yl)propan-2-ol; 1H-1,2,4-Triazole-1-ethanol,alpha-(4-(4-chlorophenoxy)-2- (trifluoromethyl)phenyl)-alpha-methyl; BAS750F;

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 : ~100 mg/mL (~251.40 mM)

Solubility in Formulation 1: ≥ 6.25 mg/mL (15.71 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 62.5 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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Solubility in Formulation 2: ≥ 6.25 mg/mL (15.71 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 62.5 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 3: ≥ 6.25 mg/mL (15.71 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 62.5 mg/mL clear DMSO stock solution to 900 μL corn oil and mix evenly.

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