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1.
Electrophoresis ; 45(11-12): 1033-1040, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38456379

RESUMEN

Imazamox (IMX), a chiral herbicide used in cereals and oilseed crops to control weeds, is commonly sold as a racemic mixture. Its enantiomers, being chiral compounds, may exhibit unique properties when exposed to chiral environments. While IMX enantiomers have been reported to degrade differently in soil and be toxic to some species, their effects on human systems remain poorly understood. This study utilized Caco-2 (human colon adenocarcinoma cell line) cells to assess the in vitro permeability of a racemic mixture of IMX and its isolated enantiomers. Additionally, the study aimed to evaluate whether the metabolite imazamox-O-desmethyl (IMX-D) forms during the permeability process. An enantioselective chromatographic method was developed, fully validated, and the apparent permeability values were obtained. The apparent permeability of rac-IMX, (+)-IMX, and (-)-IMX was determined to be 4.15 × 10-5, 5.78 × 10-5, and 7.33 × 10-5 cm s-1, respectively. These findings suggest that IMX exhibits high intestinal permeability, with an enantioselective absorption for (-)-IMX as compared to (+)-IMX. Finally, the permeability study in Caco-2 cells revealed that the metabolite IMX-D was not generated.


Asunto(s)
Permeabilidad , Humanos , Células CACO-2 , Estereoisomerismo , Imidazoles/química , Imidazoles/metabolismo , Reproducibilidad de los Resultados , Límite de Detección , Modelos Lineales , Cromatografía Líquida de Alta Presión/métodos , Plaguicidas/química , Plaguicidas/metabolismo
2.
Toxicol Lett ; 351: 1-9, 2021 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-34407455

RESUMEN

Tebuconazole (TEB) is a chiral triazole fungicide worldwide employed to control plant pathogens and preserve wood. People can be exposed to TEB either through diet and occupational contamination. This work investigates the in vitro inhibitory potential of rac-TEB, S-(+)-TEB, and R-(-)-TEB over the main cytochrome P450 enzymes (CYP450) using human liver microsomes to predict TEB in vivo inhibition potential. The IC50 values showed that in vitro inhibition was enantioselective for CYP2C9, CYP2C19, and CYP2D6, but not for CYP3A4/5. Despite enantioselectivity, rac-TEB and its single enantiomers were always classified in the same category. The inhibition mechanisms and constants were determined for rac-TEB and it has shown to be a mixed inhibitor of CYP3A4/5 (Ki = 1.3 ± 0.3 µM, αKi = 3.2 ± 0.5 µM; Ki = 0.6 ± 0.3 µM, αKi = 1.3 ± 0.3 µM) and CYP2C9 (Ki = 0.7 ± 0.1 µM, αKi = 2.7 ± 0.5 µM), and a competitive inhibitor of CYP2D6 (Ki = 11.9 ± 0.7 µM) and CYP2C19 (Ki = 0.23 ± 0.02 µM), respectively, suggesting that in some cases, rac-TEB has a higher or comparable inhibitory potential than well-known strong inhibitors of CYP450 enzymes, especially for CYP2C9 and CYP2C19. In vitro-in vivo extrapolations (IVIVE) were conducted based on the results and data available in the literature about TEB absorption and metabolism. R1 values were estimated based on the Food and Drug Administration guideline and suggested that in a chronic oral exposure scenario considering the acceptable daily intake dose proposed by the European Food and Safety Authority, the hypothesis of rac-TEB to inhibit the activities of CYP3A4/5, CYP2C9, and CYP2C19 in vivo and cause pesticide-drug interactions cannot be disregarded.


Asunto(s)
Inhibidores Enzimáticos del Citocromo P-450/farmacología , Sistema Enzimático del Citocromo P-450/metabolismo , Plaguicidas/farmacología , Triazoles/química , Triazoles/farmacología , Inhibidores Enzimáticos del Citocromo P-450/química , Humanos , Estructura Molecular , Plaguicidas/química , Relación Estructura-Actividad
3.
Food Chem Toxicol ; 146: 111826, 2020 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33127494

RESUMEN

Fenamiphos (FS) is a chiral organophosphate pesticide that is used to control nematodes in several crops. Enantioselective differences may be observed in FS activity, bioaccumulation, metabolism, and toxicity. Humans may be exposed to FS through occupational and chronic (food, water, and environmental) exposure. FS may cause undesirable CYP450 pesticide-drug interactions, which may impact human health. Here, the CYP450 isoforms involved in enantioselective FS metabolism were identified, and CYP450 inhibition by rac-FS, (+)-FS, and (-)-FS was evaluated to obtain reliable information on enantioselective FS risk assessment in humans. CYP3A4 and CYP2E1 metabolized FS enantiomers, and CYP2B6 may participate in rac-FS metabolism. In addition, rac-FS, (+)-FS, and (-)-FS were reversible competitive CYP1A2, CYP2C19, and CYP3A4/5 inhibitors. High stereoselective inhibition potential was verified; rac-FS and (-)-FS strongly inhibited and (+)-FS moderately inhibited CYP1A2. Stereoselective differences were also detected for CYP2C19 and CYP3A4/5, which were strongly inhibited by rac-FS, (+)-FS, and (-)-FS. Our results indicated a high potential for CYP450 drug-pesticide interactions, which may affect human health. The lack of stereoselective research on the effect of chiral pesticides on the activity of CYP450 isoforms highlights the importance of assessing the risks of such pesticides in humans.


Asunto(s)
Inhibidores Enzimáticos del Citocromo P-450/farmacología , Sistema Enzimático del Citocromo P-450/metabolismo , Isoenzimas/metabolismo , Compuestos Organofosforados/metabolismo , Plaguicidas/metabolismo , Sistema Enzimático del Citocromo P-450/química , Sistema Enzimático del Citocromo P-450/genética , Interacciones Farmacológicas , Humanos , Isoenzimas/antagonistas & inhibidores , Isoenzimas/química , Isoenzimas/genética , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/metabolismo , Compuestos Organofosforados/toxicidad , Plaguicidas/toxicidad , Proteínas Recombinantes
4.
J Pharm Biomed Anal ; 187: 113349, 2020 Aug 05.
Artículo en Inglés | MEDLINE | ID: mdl-32413833

RESUMEN

Ethofumesate (ETO) is a chiral herbicide that is marketed as a racemic mixture in the European Union and the United States. The growing consumption of pesticides in the world, along with their presence in water and food, has increased human exposure to these chemicals. Another issue concerning these compounds is that each enantiomer of a chiral pesticide may interact with biomolecules differently. For this reason, this study aimed to investigate the in vitro metabolism of ethofumesate (the racemic mixture as well as the isolated enantiomers) by human liver microsomes (HLM) and to explore the in vitro-in vivo correlation. Before the kinetics was determined, the method was fully validated by evaluating its selectivity, linearity, precision, accuracy, carryover, and stability. All the evaluated parameters agreed with the European Medicines Agency guideline. The enzyme kinetic parameters and the in vitro-in vivo correlation demonstrated that there was no enantioselective difference for the metabolism and bioavailable fraction of each enantiomer. The enzyme kinetics was biphasic; the KM1 values were 15, 5.8, and 5.6 for rac-ETO, (+)-ETO, and (-)-ETO, respectively. The total in vitro intrinsic clearance was 0.10 mg mL min-1 mg-1 for rac-ETO and its enantiomers. The enantiomer (-)-ETO was only metabolized by CYP2C19, while (+)-ETO was metabolized by both CYP2C19 and CYP3A4. CYP2C19 polymorphism and/or inhibition may represent a risk for humans exposed to this pesticide.


Asunto(s)
Benzofuranos/metabolismo , Herbicidas/metabolismo , Mesilatos/metabolismo , Microsomas Hepáticos/metabolismo , Benzofuranos/química , Citocromo P-450 CYP2C19/metabolismo , Citocromo P-450 CYP3A/metabolismo , Herbicidas/química , Humanos , Técnicas In Vitro , Mesilatos/química , Reproducibilidad de los Resultados , Estereoisomerismo
5.
Planta Med ; 86(6): 415-424, 2020 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-32126582

RESUMEN

Ocotea fasciculata presents yangambin (YAN) and its isomer epi-yangambin (EPI-YAN) as major lignans, which are employed as the plant markers for quality control purposes and as potential pharmacological compounds. However, a gap between the pure isomers and safety and efficacy protocols is faced by the scientific community. In this context, this work aimed to report (i) a new and advantageous purifying process in a semi-preparative scale for YAN and EPI-YAN isolation from Ocotea fasciculata, and (ii) an in vitro cytotoxicity study to estimate, for the first time, the LD50 values of the isolated epimers, as well as the influence of albumin concentration in cell culture medium. The best condition for epimers isolation was achieved in normal-phase liquid chromatography. The lignan fraction (LF), previously obtained from the plant ethanolic extract, was purified yielding 17% and 29% of YAN and EPI-YAN, respectively. The in vitro study demonstrated that YAN and EPI-YAN were safe, and only at the highest concentration studied, a decrease on cell viability was observed. The estimated LD50 value was higher than 1612 mg/kg for both epimers. The LF, on the other hand, demonstrated an estimated LD50 of 422 mg/kg. Lignan cytotoxicity studies also evidenced that the higher cell viability was related to the higher concentration of fetal bovine serum as a source of albumin in medium. This is the first time the LD50 and safety of the isolated epimers were estimated, opening up great perspectives of success in in vivo studies.


Asunto(s)
Furanos , Lignanos , Ocotea , Extractos Vegetales
6.
Toxicol Lett ; 313: 196-204, 2019 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-31278966

RESUMEN

Fipronil is a chiral insecticide employed worldwide in crops, control of public hygiene and control of veterinary pests. Humans can be exposed to fipronil through occupational, food, and environmental contamination. Therefore, the risk assessment of fipronil in humans is important to protect human health. Fipronil sulfone is the major metabolite formed during fipronil metabolism by humans. Since the CYP450 enzymes are the main ones involved in drug metabolism, the evaluation of their inhibition by fipronil and its main metabolite is important to predict drug-pesticide interactions. The aim of this work was to investigate the inhibition effects of rac-fipronil, S-fipronil, R-fipronil and fipronil sulfone on the main human CYP450 isoforms. The results showed that CYP2D6 is the only CYP450 isoform inhibited by these xenobiotics. In addition, no enantioselective differences were observed in the inhibition of CYP450 isoforms by fipronil and its individuals' enantiomers. Rac-fipronil, S-fipronil and R-fipronil are moderate CYP2D6 inhibitors showing a competitive inhibition profile. On the other hand, the metabolite fipronil sulfone showed to be a strong inhibitor of CYP2D6 also by competitive inhibition. These results highlight the importance of metabolite evaluation on pesticide safety since the metabolism of fipronil into fipronil sulfone increases the risk of pesticide-drug interactions for drugs metabolized by CYP2D6.


Asunto(s)
Inhibidores del Citocromo P-450 CYP2D6/toxicidad , Citocromo P-450 CYP2D6/metabolismo , Plaguicidas/toxicidad , Pirazoles/toxicidad , Citocromo P-450 CYP2D6/química , Inhibidores del Citocromo P-450 CYP2D6/química , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Humanos , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Modelos Biológicos , Plaguicidas/química , Conformación Proteica , Pirazoles/química , Medición de Riesgo , Relación Estructura-Actividad
7.
J Pharm Biomed Anal ; 147: 89-109, 2018 Jan 05.
Artículo en Inglés | MEDLINE | ID: mdl-28844369

RESUMEN

The consumption of pesticides worldwide has been growing in recent decades, and consequently the exposure of humans and other animals to them as well. However, even though it is known that chiral pesticides can behave stereoselectively, the knowledge about the risks to human health and the environment is scarce. Among the pesticides registered to date, approximately 30% have at least one center of asymmetry, and just 7% of them are currently marketed as a pure stereoisomer or as an enriched mixture of the active stereoisomer. There are several in vitro, in vivo, and in silico models available to evaluate the enantioselective metabolism of chiral pesticides aiming ecotoxicological and risk assessment. Therefore, this paper intends to provide a critical view of the metabolism of chiral pesticides in non-target species, including humans, and discuss their implications, as well as, conduct a review of the analytical techniques employed for in vitro and in vivo metabolism studies of chiral pesticides.


Asunto(s)
Plaguicidas/metabolismo , Animales , Cromatografía Líquida de Alta Presión , Humanos , Plaguicidas/química , Estereoisomerismo
8.
Planta Med ; 83(8): 737-745, 2017 May.
Artículo en Inglés | MEDLINE | ID: mdl-28073118

RESUMEN

Artepillin C, a natural product present in the Brazilian green propolis, has several biological properties. Among these properties, the antitumor action of this product is noteworthy and makes it a promising drug candidate for the treatment of several types of cancer. This paper describes the in vitro metabolism of Artepillin C in rat and human liver microsomes. The rat model suggested a sigmoidal profile for the metabolism, adapted to the Hill's kinetic model. The enzymatic kinetic parameters were as follows: maximal velocity = 0.757 ± 0.021 µmol/mg protein/min, Hill coefficient = 10.90 ± 2.80, and substrate concentration at which half-maximal velocity of a Hill enzyme is achieved = 33.35 ± 0.55 µM. Based on these results, the calculated in vitro intrinsic clearance for Artepillin C was 16.63 ± 1.52 µL/min/mg protein. The in vitro metabolism assay conducted on the human model did not fit any enzymatic kinetic model. Two novel metabolites were formed in both mammal microsomal models and their chemical structures were elucidated for the first time. The main human cytochrome P450 isoforms involved in Artepillin C metabolism had been identified, and the results suggest a majority contribution of CYP2E1 and CYP2C9 in the formation of the two metabolites.


Asunto(s)
Microsomas Hepáticos/metabolismo , Fenilpropionatos/metabolismo , Animales , Antineoplásicos/metabolismo , Citocromo P-450 CYP2C9/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Humanos , Própolis/química , Ratas , Ratas Sprague-Dawley
9.
J Chromatogr A ; 1467: 326-334, 2016 Oct 07.
Artículo en Inglés | MEDLINE | ID: mdl-27554030

RESUMEN

The distinct activity and toxicity of enantiomers has increased concern about the use of chiral pesticides. The chiral pesticide Fenamiphos (FS) is employed as a racemic mixture to control nematode pests. Although recent studies revealed that FS enantiomers possess different toxicity, the toxicokinetics and liver metabolism of these enantiomers in humans remain unclear. This study characterizes the in vitro metabolism of rac-FS, (+)-FS, and (-)-FS by human liver microsomes and predicts some toxicokinetic parameters. First, a new enantioselective HPLC method was developed to analyze FS and its metabolites [fenamiphos sulfoxide (FSO) and fenamiphos sulfone (FSO2)]. Chiral separation of the stereoisomers was accomplished in an in-line coupled achiral-chiral column (Lichrosorb Si60 - Chiralpak AS-H); hexane: ethanol: methanol (85:12:3, v/v/v) was used as mobile phase at a flow rate of 1.2mLmin-1. Then, the HPLC method was fully validated. All the evaluated parameters agreed with the European Medicines Agency guideline. Finally, the enantioselective kinetic parameters were determined for CYP450 enzymes. The predicted toxicokinetic parameters showed that the liver exclusively eliminated FS without stereoselectivity.


Asunto(s)
Microsomas Hepáticos/metabolismo , Compuestos Organofosforados/metabolismo , Plaguicidas/metabolismo , Cromatografía Líquida de Alta Presión/métodos , Humanos , Compuestos Organofosforados/química , Plaguicidas/química , Estereoisomerismo
10.
J Pharm Biomed Anal ; 128: 528-537, 2016 Sep 05.
Artículo en Inglés | MEDLINE | ID: mdl-27381871

RESUMEN

A new capillary electrophoresis method for the enantioselective analysis of cis- and trans- dihydrotetrabenazine (diHTBZ) after in vitro metabolism by human liver microsomes (HLMs) was developed. The chiral electrophoretic separations were performed by using tris-phosphate buffer (pH 2.5) containing 1% (w/v) carboxymethyl-ß-CD as background electrolyte with an applied voltage of +15kV and capillary temperature kept at 15°C. Dispersive liquid-liquid microextraction was employed to extract the analytes from HLMs. Dichloromethane was used as extraction solvent (75µL) and acetone as disperser solvent (150µL). The method was validated according to official guidelines and showed to be linear over the concentration range of 0.29-19.57µmolL(-1) (r=0.9955) for each metabolite enantiomer. Within- and between-day precision and accuracy evaluated by relative standard deviation and relative error were lower than 15% for all enantiomers. The stability assay showed that the analytes kept stable under handling, storage and in metabolism conditions. After method validation, an enantioselective in vitro metabolism and in vivo pharmacokinetic prediction was carried out. This study showed a stereoselective metabolism and the observed kinetic profile indicated a substrate inhibition behavior. DiHTBZ enantiomers were catalyzed mainly by CYP2C19 and the predicted clearance suggests that liver metabolism is the main route for TBZ elimination which supports the literature data.


Asunto(s)
Electroforesis Capilar/métodos , Microextracción en Fase Líquida/métodos , Microsomas Hepáticos/metabolismo , Tetrabenazina/farmacocinética , Citocromo P-450 CYP2C19/metabolismo , Estabilidad de Medicamentos , Humanos , Técnicas In Vitro , Reproducibilidad de los Resultados , Estereoisomerismo , Tetrabenazina/análogos & derivados , Tetrabenazina/metabolismo
11.
Talanta ; 154: 511-9, 2016 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-27154708

RESUMEN

In this study, a novel method combining dispersive liquid-liquid microextraction (DLLME) and fast liquid chromatography coupled to mass spectrometry (LC-MS/MS) was developed and validated for the extraction and determination of bisphenol A (BPA) and six bisphenol analogues, namely bisphenol S (BPS), bisphenol F (BPF), bisphenol P (BPP), bisphenol Z (BPZ), bisphenol AP (BPAP) and bisphenol AF (BPAF) in human urine samples. Type and volume of extraction and disperser solvents, pH sample, ionic strength, and agitation were evaluated. The matrix-matched calibration curves of all analytes were linear with correlation coefficients higher than 0.99 in the range level of 0.5-20.0ngmL(-1). The relative standard deviation (RSD), precision, at three concentrations (1.0, 8.0 and 15.0ngmL(-1)) was lower than 15% with accuracy ranging from 90 to 112%. The biomonitoring capability of the new method was confirmed with the analysis of 50 human urine samples randomly collected from Brazilians. BPA was detected in 92% of the analyzed samples at concentrations ranging

Asunto(s)
Microextracción en Fase Líquida , Compuestos de Bencidrilo , Brasil , Cromatografía Liquida , Humanos , Fenoles , Espectrometría de Masas en Tándem
12.
J Pharm Biomed Anal ; 109: 192-201, 2015 May 10.
Artículo en Inglés | MEDLINE | ID: mdl-25778930

RESUMEN

Zopiclone (ZO) is a chiral drug that undergoes extensive metabolism to N-desmethylzopiclone (N-Des-ZO) and zopiclone-N-oxide (N-Ox-ZO). Pharmacological studies have shown (S)-N-Des-ZO metabolite presents anxiolytic activity and a patent for this metabolite was requested for anxiety treatment and related disorders. In this context, biotransformation employing fungi may be a promising strategy to obtain N-Des-ZO. To perform the biotransformation study in this work, an enantioselective method based on capillary electrophoresis (CE) and dispersive liquid-liquid microextraction (DLLME) was developed. CE analyses were carried out in sodium phosphate buffer (pH 2.5; 50mmolL(-1)) containing 0.5% (w/v) carboxymethyl-ß-CD, at a constant voltage of +25kV. DLLME was conducted using 2mL of liquid culture medium pH 9.5. Chloroform (100µL) and methanol (300µL) were employed as extraction and disperser solvent, respectively. After CE and DLLME optimization, the analytical method was fully validated. The method was linear over a concentration range of 90-6000ngmL(-1) for each ZO enantiomer (r>0.999) and 50-1000ngmL(-1) for each N-Des-ZO enantiomer (r>0.998). Absolute recovery of 51 and 82% was achieved for N-Des-ZO and ZO, respectively. The accuracy and precision results agreed with the EMA (European Medicines Agency) guideline, and so did the stability study. Application of the developed method in a biotransformation study was conducted in order to investigate the ability of fungi, belonging to the genus Cunninghamella, in metabolizing ZO chiral drug. Fungi Cunninghamella elegans ATCC 10028B and Cunninghamella echinulata var elegans ATCC 8688A demonstrated to be able to enantioselectively biotransform ZO to its active metabolite, N-Des-ZO. Therefore, the proposed goals of this work, i.e. a fast DLLME-CE method and an outstanding strategy to obtain N-Des-ZO, were successfully attained.


Asunto(s)
Compuestos de Azabiciclo/análisis , Compuestos de Azabiciclo/metabolismo , Cunninghamella/metabolismo , Hipnóticos y Sedantes/análisis , Hipnóticos y Sedantes/metabolismo , Piperazinas/análisis , Piperazinas/metabolismo , Biotransformación , Electroforesis Capilar , Indicadores y Reactivos , Microextracción en Fase Líquida , Estereoisomerismo
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