Effects of Compound troxerutin and poreine cerebroside injection on cytochrome P450 enzyme in vivo and in vitro / 中国药房

OBJECTIVE To investigate the effects of Compound troxerutin and poreine cerebroside injection on the activity of cytochrome P450 （CYP450） enzyme in vivo and in vitro. METHODS Human liver microsomes were incubated with Compound troxerutin and poreine cerebroside injection （volume fraction 0.05%-10%） and the specific probe substrates of CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6 and CYP3A4 for 30 min. The production of corresponding metabolites was detected by ultra-high performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS）， and the half inhibitory concentration （IC50） was calculated. The relative mRNA expression （i.e. induction multiple） of CYP450 enzyme was determined by real-time fluorescence quantitative PCR after human primary hepatocytes were incubated with Compound troxerutin and poreine cerebroside injection （volume fraction 0.05%-10%） or 3 positive inducers of CYP1A2， CYP2B6， CYP3A4 for 48 hours. Male SD rats were randomly divided into control group （normal saline+probe substrates of CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6， CYP3A4 8， 2， 1， 1， 10， 10， 8 mg/kg） and experimental group （Compound troxerutin and poreine cerebroside injection 0.9 mL/kg+probe substrates of CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6， CYP3A4 8， 2， 1， 1， 10，10， 8 mg/kg）， with 6 rats in each group. The pharmacokinetic parameters of probe substrates were detected by UPLC-MS/MS and Cocktail probe drug method. RESULTS After the lzqpharm@126.com treatment of 0.05%-10% Compound troxerutin and poreine cerebroside injection， the activities of CYP2B6， CYP2C8 and CYP2C19 in human liver microsomes had no significant change， and IC50 could not be fitted； IC50 of CYP1A2， CYP2C9， CYP2D6 and CYP3A4 were 419.90%， 97.78%， 176.00%， 19.42%， respectively. After the treatment of 0.05%-10% Compound troxerutin and poreine cerebroside injection， the average induction multiple of CYP3A4 mRNA in human primary hepatocytes （No. MHK） was 4.88 （and the average induction multiples of 2 concentration points were higher than 2）. After the treatment of Compound troxerutin and poreine cerebroside injection， AUC0－t and AUC0－∞ of CYP2C8， CYP2C9 and CYP2C19 substrates were increased significantly， CL of CYP2C8 and CYP2C19 substrates were decreased significantly， while t1/2 of CYP2C9 substrate was prolonged significantly （P＜0.05）. CONCLUSIONS Compound troxerutin and poreine cerebroside injection has no obvious inhibitory effect on CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6 and CYP3A4 in human liver microsomes in vitro， but can induce the mRNA expression of CYP3A4 in human primary hepatocytes in vitro， and can inhibit the activities of CYP2C8， CYP2C9 and CYP2C19 in rats in vivo.

Green synthesis of gold nanaoparticles using Delphinium Chitralense tuber extracts, their characterization and enzyme inhibitory potential

Abstract Green synthesis has been introduced as an alternative to chemical synthesis due to the serious consequences. Metal nanoparticles synthesized through green approach have different pharmaceutical, medical and agricultural applications. The present study followed a green and simple route for the preparation of potentially bioactive gold nanoparticles (Au NPs). Au NPs were prepared via green synthesis approach using crude basic alkaloidal portion of the tuber of Delphinium chitralense. The green synthesized Au NPs were characterized by transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray diffraction (XRD) fourier transform infrared (FTIR), and UV-Visible spectrophotometer. Morphological analysis shows that Au NPs have cubic geometry with different sizes. UV-Vis spectroscopic analysis confirmed the synthesis of Au NPs while XRD proved their pure crystalline phase. The Au NPs showed promising dose dependent inhibition of both AChE and BChE as compared to the crude as well as standard drug.

Resumo A síntese verde foi introduzida como uma alternativa à síntese química devido às graves consequências. As nanopartículas metálicas sintetizadas através da abordagem verde têm diferentes aplicações farmacêuticas, médicas e agrícolas. O presente estudo seguiu uma rota verde e simples para a preparação de nanopartículas de ouro potencialmente bioativas (Au NPs). As NPs de Au foram preparadas via abordagem de síntese verde usando a porção alcaloide básica bruta do tubérculo de Delphinium chitralense. As NPs de Au sintetizadas verdes foram caracterizadas por microscopia eletrônica de transmissão (TEM), microscopia eletrônica de varredura (MEV), difração de raios X (DRX), infravermelho com transformada de Fourier (FTIR) e espectrofotômetro UV-Visível. A análise morfológica mostra que as NPs de Au possuem geometria cúbica com tamanhos diferentes. A análise espectroscópica UV-Vis confirmou a síntese de Au NPs enquanto a XRD provou sua fase cristalina pura. O Au NPs mostrou inibição dependente da dose promissora de AChE e BChE em comparação com a droga bruta e padrão.

Green synthesis of gold nanaoparticles using Delphinium Chitralense tuber extracts, their characterization and enzyme inhibitory potential / Síntese verde de nanaopartículas de ouro utilizando extratos de tubérculos Delphinium chitralense, sua caracterização e potencial inibitório enzimático

Greeen synthesis has been introduced as an alternative to chemical synthesis due to the serious consequences. Metal nanoparticles synthesized through green approach have different pharmaceutical, medical and agricultural applications. The present study followed a green and simple route for the preparation of potentially bioactive gold nanoparticles (Au NPs). Au NPs were prepared via green synthesis approach using crude basic alkaloidal portion of the tuber of Delphinium chitralense. The green synthesized Au NPs were characterized by transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray diffraction (XRD) fourier transform infrared (FTIR), and UV-Visible spectrophotometer. Morphological analysis shows that Au NPs have cubic geometry with different sizes. UV-Vis spectroscopic analysis confirmed the synthesis of Au NPs while XRD proved their pure crystalline phase. The Au NPs showed promising dose dependent inhibition of both AChE and BChE as compared to the crude as well as standard drug.

A síntese verde foi introduzida como uma alternativa à síntese química devido às graves consequências. As nanopartículas metálicas sintetizadas através da abordagem verde têm diferentes aplicações farmacêuticas, médicas e agrícolas. O presente estudo seguiu uma rota verde e simples para a preparação de nanopartículas de ouro potencialmente bioativas (Au NPs). As NPs de Au foram preparadas via abordagem de síntese verde usando a porção alcaloide básica bruta do tubérculo de Delphinium chitralense. As NPs de Au sintetizadas verdes foram caracterizadas por microscopia eletrônica de transmissão (TEM), microscopia eletrônica de varredura (MEV), difração de raios X (DRX), infravermelho com transformada de Fourier (FTIR) e espectrofotômetro UV-Visível. A análise morfológica mostra que as NPs de Au possuem geometria cúbica com tamanhos diferentes. A análise espectroscópica UV-Vis confirmou a síntese de Au NPs enquanto a XRD provou sua fase cristalina pura. O Au NPs mostrou inibição dependente da dose promissora de AChE e BChE em comparação com a droga bruta e padrão.

The Influence of Different Extraction Methods/Solvents on Composition, Biological Activities and ADMET Predictions of Phenolics in Tribulus terrestris

Abstract Tribulus terrestris is a plant that has medical importance in treating diseases such as cancer, urinary tract infections, kidney stones, rheumatism, and hypertension. The aim of this study is to examine the effects of extraction methods/solvents on the biological activity and chemical content of the plant and then to determine ADMET predictions of phenolics that were analysed quantitatively using HPLC-DAD in the plant. Maceration methanol (IC50:0.277 mg/mL) and chloroform (0.263 mMFeSO4/mg extract) extracts were found to have the strongest DPPH radical scavenging and iron (III) ion reducing activity, respectively. It was determined that Soxhlet methanol (0.0225 mMTE/mg extract) and Soxhlet chloroform (1.360 mMTE/mg extract) extracts exhibited stronger radical cation (ABTS.+) scavenging and cupric ion reducing activity compared to other extracts, respectively. It was found that ultrasonic bath methanol extracts showed the highest anti-urease (21.47%) and calcium oxalate anti-crystallization (71.54%, 80.52%) activity. It was also found that Soxhlet methanol extract (66.763%) has strong acetylcholinesterase enzyme inhibition capacity compared to other extracts. Pyrocatechol, vanillic acid, vanillin, rutin and rosmarinic acid were analysed by HPLC-DAD in the plant. Moreover, it was determined that methanol extracts obtained using soxhlet, ultrasonic bath, and maceration contained the highest amount of rutin, pyrocatechol, and rutin, respectively. ADMET predictions of phenolics show that these compounds are easily absorbed and do not have toxic effects, suggesting that this species may be used as a natural medicinal and nutritional source in the future after detailed analysis tests.

Phenolic Content and Enzyme Inhibition Activities Barbarea auriculata var. paludosa, B. integrifolia and B. plantaginea (Brassicaceae)'s

Barbareagenus has been presented about 12 species, 9 taxon are endemic, in Turkey. In this sudy, enzyme inhibition was carried out on methanolic extract, chloroform extract, ethyl acetate extract, and the remaining aqueous phases from the aerial parts of B. auriculata var paludosa, B. integrifolia, andB. plantagineaspecies and HPLC studies were carried on their methanolic extract in the present study for the first time. Phenolic compounds were determined using reverse phase-high performance liquid chromatography (RP-HPLC). p-OH benzoic acid, vanilic acid, syringaldehyde, coumaric acid, synapic acid and benzoic acid were detected as major phenolic compounds in the species. Assay of enzyme inhibition activitieswere done using spectrophotometric methods. Results of these studies reveal that the extracts from these species have moderate tyrosinase, AChE and BuChE inhibitory activity. In the biological activity studies, it was observed that B. integrifoliawas thehighest activity

Neuropharmacological evaluation on four traditional herbs used as nervine tonic and commonly available as Shankhpushpi in India

Background: Shankhpushpi is an Ayurvedic drug, widely used for its actions on the central nervoussystem, especially to improve intellect and boost memory. Four botanicals viz. Canscora decussata Schult.(CD), Clitorea ternatea Linn. (CT), Convolvulus pluricaulis Choisy. (CP) and Evolvulus alsinoides Linn. (EA)are considered as sources of Shankhpushpi by Indian practitioners on the basis of their morphologicaldescriptions given in ancient texts.Objective: The present study was undertaken to evaluate the neuropharmacological effect of four herbscommonly identified as source of Shankhpushpi.Materials and methods: Methanol extracts of all four varieties were tested and evaluated in vitro and invivo for their neuropharmacological effects. Experiments such as protection against b-amyloid inducedneurotoxicity on brain cell line (Neuro 2A), antioxidant potential, AchE (acetylcholinesterase enzyme)inhibition, and 5-LOX (lipoxygenase) enzyme inhibition were conducted for in vitro evaluation. For in vivoevaluation, scopolamine (0.3 mg/kg i.p.) induced memory retrieval using pole climbing apparatus andMorris water maze were performed in rat models.Results: It was found that protective effects of EA and CD against b-amyloid induced neurotoxicity inNeuro 2A cells were significantly higher than CT and CP. EA proved to be superior than other varieties onthe basis of antioxidant activity, AchE inhibitory and LOX inhibitory activities. The preventive activity ofEA on scopolamine induced memory retrieval in pole climbing and Morris water maze task in rats wasfound to be higher than that of CD, CT and CP.Conclusion: EA has remarkable neuropharmacological effect as compared to other three varietiesof Shankhpushpi. This effect may be attributed due to the presence of steroids (stigmasterol and betulinicacid), coumarins (scopoletin) and flavonoids (b-carotene and chlorogenic acid). Hence it can be used as apromising lead in development and management of neuronal disorders including Alzheimer's disease.© 2017 Transdisciplinary University, Bangalore and World Ayurveda Foundation. Publishing Services byElsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)

Value of the Filtration Fraction Assessed by Dynamic ⁹⁹mTc-Diethylenetriaminepentaacetic Acid Renal Scintigraphy After Angiotensin-Converting Enzyme Inhibition for the Diagnosis of Renovascular Hypertension / 대한핵의학회잡지

PURPOSE: This study aimed to determine the diagnostic value of the relative filtration fraction (RFF) assessed by dynamic ⁹⁹mTc-diethylenetriaminepentaacetic acid (⁹⁹mTc-DTPA) renal scintigraphy after angiotensin-converting enzyme (ACE) inhibition for renovascular hypertension (RVHT) diagnosis.METHODS: ⁹⁹mTc-DTPA captopril renal scintigraphy performed in adolescents or adults (≥ 10 years) with suspected RVHT was retrospectively reviewed. The RFF of the affected kidney was qualitatively assessed as the relative glomerular filtration rate during the 2 to 3-min period compared with the relative perfusion during the first 60 s (qualitative RFF) and scored from 1 (definitely same) to 5 (definitely decreased). The quantitative RFF of the affected kidney was obtained by dividing the percentage of glomerular filtration rate by the percentage of renal perfusion.RESULTS: Overall, 173 patients (high probability, n = 15; and low probability, n = 158) were included based on conventional captopril renal scintigraphic criteria. An abnormal qualitative RFF was observed in 12 patients with high probability, and the diagnostic sensitivity was 80.0% (95% CI, 51.9–95.7). The RFF was normal in 152 patients with low probability, and the diagnostic specificity was 96.2% (95% CI, 91.9–98.6). The RFF was lower in patients with high probability than in those with low probability (0.79 ± 0.15 vs. 1.02 ± 0.11, P < 0.0001).CONCLUSIONS: The RFF assessed by dynamic ⁹⁹mTc-DTPA renal scintigraphy after ACE inhibition can detect patients with high probability for RVHT. The RFF after ACE inhibition might be a useful diagnostic criterion especially when baseline scintigraphy is not available for evaluating ACE inhibition-induced changes.

Inhibición de la a-amilasa por medio de extractos de plantas medicinales como tratamiento complementario/alternativo de la diabetes y la caries / Inhibition of a a-amylase by means of extracts of medicinal plants as complementary/alternative treatment of diabetes and caries

Los inhibidores selectivos de la α-amilasa salival y pancreática humana, son un medio eficaz para controlar los niveles de azúcar en la saliva y la sangre, durante el control de la caries y la diabetes. En la presente revisión, después de identificar las principales funciones de la enzima, se discutieron exponen algunas de las respuestas observadas después de la exposición de la α-amilasa a las plantas, en escala molecular y entera. Diferentes tipos de moléculas de plantas medicinales actúan como inhibidores de la α-amilasa, como una terapia alternativa o complementaria en el tratamiento de la diabetes en y en el control de uno de los factores de la formación la caries (dieta) (AU)

Selective inhibitors of human salivary and pancreatic α-amylase are an effective means of controlling saliva and blood sugar levels in the management of caries and diabetes. In the present review, after identifying the main functions of the enzyme, it was exposed some of the observed responses after exposure to α-amylase at the molecular and whole plants scale. Different types of medicinal plants molecules have been found to act as α-amylase inhibitors, as an alternative or complementary therapy in the management of diabetes and control to one of the predisposing factors to caries (diet) (AU)

Inhibition of cytochrome P450 3A enzyme by Millettia aboensis: its effect on the pharmacokinetic properties of efavirenz and nevirapine

ABSTRACT The chronic and comorbid nature of HIV infection necessitate the use of multiple drugs including herbs to relieve symptoms with a possible increase in herb–drug interaction cases. This study was designed to evaluate the effect of Millettia aboensis (Hook. f.) Baker, Fabaceae, on cytochrome P450 3A isoenzyme and the influence of this effect on the bioavailability of two antiretroviral agents. In vitro effect of ethanol extract of M. aboensis on intestinal and liver microsomes extracted from female rats was assessed using erythromycin-N-demethylation assay method while in vivo effects were determined by estimating simvastatin plasma concentrations in rats. The effect of the extract on pharmacokinetic parameters of orally administered efavirenz (25 mg/kg) and nevirapine (20 mg/kg) was determined in rats divided into groups (n = 5). Plasma drug concentrations were assayed using HPLC and pharmacokinetic parameters determined through a non-compartmental analysis as implemented in WinNonlin pharmacokinetic program. The extract inhibited both intestinal and liver microsomal cytochrome P450 3A isoenzyme activities in vitro and enhanced simvastatin absorption in vivo with possible inhibition of metabolizing enzymes as indicated by significant (p < 0.05) increase in maximal concentration, area under curve and mean resident time of the drug. However, further in vivo interaction studies in animal model did not produce significant (p > 0.05) changes in the pharmacokinetic parameters of efavirenz and nevirapine. HPLC fingerprinting indicated the presence of quercetin and kaempferol in the extract. These findings revealed M. aboensis as an inhibitor of cytochrome P450 3A enzyme but, with no significant effect on the bioavailability of orally administered nevirapine and efavirenz.

Inhibition of regorafenib against UDP-glucuronosyltransferases / 药学学报

This study was designed to investigate the inhibitory effects of regorafenib (REG) on the catalytic activities of 12 kinds of human UGT isoforms and human liver microsomes (HLM) in vitro. The broader potential of REG to perpetrate drug-drug interactions (DDI) arising from UGT enzyme inhibition is predicted by in vitro-vivo extrapolation (IV-IVE). Fifty mixed HLM and 12 kinds of recombinant UGTs were utilized as enzyme sources to evaluation the inhibitory effects of REG against UGTs. 4-Methylumbelliferone (4-MU) as a nonselective substrate of UGTs except for UGT1A4, N-(3-carboxypropyl)-4-hydroxy-1,8-napht-halimide (NCHN) and N-butyl-4-(4-hydroxyphenyl)-1,8-naphthalimide (NPHN) as the specific fluorescent substrate of UGT1A1, and trifluoperazine (TFP) as the specific substrate of UGT1A4. The half maximal inhibitory concentration (IC50) was calculated via the nonlinear regression analysis using Graphpad Prism 6.0, the inhibition kinetic types were selected and evaluated based on the intersection location of Lineweaver-Burk plot and Dixon plot, and Ki values were determined by the second plot of slopes. The potential DDI risk based on UGT1A1 inhibition was also evaluated through the in vitro parameters. The results demonstrated that REG displayed strong inhibitory effects against UGT1A1, 1A7, 1A9, and 2B7. The IC50 values were from 0.15 to 6.6 μmol·L-1 and Ki values from 0.027 to 14 μmol·L-1. The REG exerted competitive inhibition against UGT1A1-mediated 4-MU-O-glucuronidation and UGT1A1-mediated NPHN-O-glucuronidation, while the inhibition of NCHN-4-O-glucuronide by REG was suited to noncompetitive inhibition in both HLM and recombinant UGT1A1. Likewise, REG exhibited a mixed efficacy in inhibition of UGT1A7-, UGT1A9-, and UGT2B7-catalyzed 4-MU-O-glucuronidation. The AUC ratio of UGT1A1 specific substrates NPHN and NCHN can be increased by 101% to 302% and 13% to 109%, respectively. These results suggest that much caution should be exercised when REG is co-administered with UGT1A1 substrates.

In vitro and in vivo cytochrome P450 3A enzyme inhibition by Aframomum melegueta and Denniettia tripetala extracts

Objective To evaluate the in vitro and in vivo inhibitory effects of two commonly used herbs, Aframomum melegueta (A. melengueta) and Dennettia tripetala (D. tripetala) on CYP 3A enzymes. Methods In vitro inhibition of the enzymes were assessed with microsomes extracted from female albino rats using erythromycin-N-demethylation assay (EMND) method while their in vivo effects were measured by estimating simvastatin plasma concentrations in rats. Pharmacokinetic parameters were determined using non-compartmental analysis as implemented in WinNonlin pharmacokinetic program. Results EMND assay with intestinal microsomes indicated that aqueous extracts of D. tripetala and A. melengueta significantly (P < 0.05) inhibited intestinal CYP 3A activity at both 50 μg and 100 μg concentrations. Petroleum ether extract of D. tripetala and ethanol extracts of A. melengueta inhibited intestinal CYP3A activity at 100 μg but not at 50 μg concentrations. All the extracts showed an in vitro dose dependent CYP 3A inhibition with liver microsomes. In vivo analysis showed that pre-treatment with the extracts enhanced systemic absorption of simvastatin with reductions in metabolizing enzymes activity as indicated in significant increases in maximal concentration, area under curve, area under moment curve and mean resident time of simvastatin (P < 0.05). Conclusions Herbal preparations containing these plants' extracts should be used with caution especially in patients on CYP450 3A substrate medications.

In vitro and in vivo cytochrome P450 3A enzyme inhibition by Aframomum melegueta and Denniettia tripetala extracts

OBJECTIVE@#To evaluate the in vitro and in vivo inhibitory effects of two commonly used herbs, Aframomum melegueta (A. melengueta) and Dennettia tripetala (D. tripetala) on CYP 3A enzymes.@*METHODS@#In vitro inhibition of the enzymes were assessed with microsomes extracted from female albino rats using erythromycin-N-demethylation assay (EMND) method while their in vivo effects were measured by estimating simvastatin plasma concentrations in rats. Pharmacokinetic parameters were determined using non-compartmental analysis as implemented in WinNonlin pharmacokinetic program.@*RESULTS@#EMND assay with intestinal microsomes indicated that aqueous extracts of D. tripetala and A. melengueta significantly (P < 0.05) inhibited intestinal CYP 3A activity at both 50 μg and 100 μg concentrations. Petroleum ether extract of D. tripetala and ethanol extracts of A. melengueta inhibited intestinal CYP3A activity at 100 μg but not at 50 μg concentrations. All the extracts showed an in vitro dose dependent CYP 3A inhibition with liver microsomes. In vivo analysis showed that pre-treatment with the extracts enhanced systemic absorption of simvastatin with reductions in metabolizing enzymes activity as indicated in significant increases in maximal concentration, area under curve, area under moment curve and mean resident time of simvastatin (P < 0.05).@*CONCLUSIONS@#Herbal preparations containing these plants' extracts should be used with caution especially in patients on CYP450 3A substrate medications.

Estudos estruturais e funcionais de Tsa1 de Saccharomyces cerevisiae: um modelo biológico para o estudo de inibidores de crescimento celular para leucemia linfoide aguda / Functional and structural evaluation of Tsa1 from Saccharomyces cerevisiae: a biological model for study of cellular growth inhibitors for acute lymphocytic leukemia

As peroxirredoxinas (Prx) são enzimas antioxidantes que se destacam pela capacidade de decompor uma grande variedade de hidroperóxidos com elevada eficiência (106-108M-1s-1), mantendo essas moléculas em níveis adequados à homeostase celular. Entretanto, já foi demonstrado que em diversos tipos tumorais os níveis de Prx são extremamente aumentados e experimentos envolvendo sua inativação resultam na diferenciação ou apoptose de células tumorais. Recentemente, foi descoberto um diterpenóide denominado adenantina que seria o primeiro inibidor para as Prx1 e Prx2 de humanos e foi demonstrada que sua aplicação em células de leucemia mieloide aguda promoveu diferenciação ou apoptose dessas células. Nesse contexto, o presente trabalho apresenta duas vertentes: 1) A caracterização das alterações estruturais e funcionais promovidas pela ligação da adenantina ao sítio ativo das Prx utilizando Tsa1 de Saccharomyces cerevisiae como modelo biológico, em função da sua alta similaridade com Prx2 de humanos; 2) Avaliação da atividade antitumoral dose dependente de adenantina sobre as linhagens celulares REH e MOLT-4 de leucemia linfoide aguda. No que concerne à primeira linha de investigação, nossos resultados revelam que Tsa1 é suscetível à inibição por adenantina, uma vez que o tratamento reduziu em ~66 % a velocidade de decomposição de peróxido de hidrogênio. Adicionalmente, a mutação da Thr44 de Tsa1, pertencente à chamada tríade catalítica, por uma Ser resultou em uma proteína mais suscetível a alterações na estrutura secundária e à inibição da atividade peroxidásica em função da ligação com adenantina, apresentando uma diminuição de ~85% na velocidade de reação. Características semelhantes foram observadas para a proteoforma Tsa2 de S. cerevisiae, que carreia naturalmente a substituição da Thr44 pela Ser. Análises de sequências de Prx em bancos de dados revelaram que majoritariamente proteínas contendo Ser são encontradas em organismos procariotos, muitos deles patogênicos. Finalmente, demonstramos por meio de ensaios citotoxicidade que as bactérias Staphylococcus aureus e Staphylococcus epidermidis, que possuem uma Ser na tríade catalítica, têm seu crescimento inibido pelo tratamento com adenantina (IC50 de 460µM e 77µM, respectivamente), enquanto que para Escherichia coli, que possui Thr nessa posição, a toxicidade da adenantina foi bastante baixa (não foi possível determinar o IC50 nas condições utilizadas). Dessa forma, os dados apresentados neste trabalho demonstram o potencial da utilização da adenantina tanto como antibiótico quanto como antileucêmico

Peroxiredoxins (Prx) are antioxidant enzymes which stand out due the ability to decompose a wide variety of hydroperoxides with high efficiency (106-108M-1s-1) maintaining these molecules at suitable levels to cellular homeostasis and participating in several signaling events. However, it has been shown that, in many tumor types, Prx levels are extremely increased and experiments involving its inactivation have resulted in differentiation or apoptosis of tumor cells. It was recently found a diterpenoid, called adenanthin, that would be the first human Prx1 and Prx2 inhibitor and it was demonstrated that its application in acute myeloid leukemia cells was able to promote differentiation or apoptosis. In this context, this work presents two lines of research: 1) Characterization of structural and functional changes promoted by adenanthin binding to Prx active site using Tsa1 from Saccharomyces cerevisiae as biological model, due to its high similarity to human Prx2. 2) Evaluation of adenanthin dose-dependent antitumor activity over the acute lymphoid leukemia cell lines REH and MOLT-4. As regards the first line of research, our result reveal that Tsa1 is susceptible to inhibition by adenanthin, since the treatment with this binder reduced the hydrogen peroxide decomposition velocity in ~ 66%. In addition, the replacement of Thr44 from Tsa1, aminoacid belonging to the so-called catalytic triad, by a Ser resulted in a protein more susceptible to alterations in secondary structure and to peroxidase activity inhibition in function of adenanthin binding, presenting ~85% of decrease in reaction velocity. Similar characteristics were observed for Tsa2 proteoform from S. cerevisiae, which naturally carries the substitution of Thr44 by Ser. Prx sequences analyzes in databases revealed that mostly Ser-containing proteins are found in prokaryotic organisms, many of them pathogenic ones. Finally, we demonstrate through cytotoxicity assays that the bacteria Staphylococcus aureus and Staphylococcus epidermidis, which have a Ser in catalytic triad, have their growth inhibited by adenanthin treatment (IC50 of 460µM and 77µM, respectively), whereas for Escherichia Coli, which has Thr at that position, the tocyxicity of adenanthin was quite low (it was not possible to determine the IC50 under the used conditions). Regarding the second line of investigation, we found that adenanthin is able to induce the death of leukemic cell lines REH and MOLT-4, and for the last one, there was an unexpected proliferation of cells treated by the longest incubation period (72 hours), characterizing a possible indication of differentiation process. In this sense, the data presented here demonstrate the potential of adenanthin use in both antibiotic and antileukemic treatment

In Vitro Metabolism of Two Novel Tacrine Derivates ST09 and ST10 / 中国药学杂志

OBJECTIVE: To determin the in vitro metabolic stability of ST09 and ST10 in human liver microsomes (HLMs), and to evaluate their potential inhibitions on five HLM cytochrome P450 isoforms. METHODS: A liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to assess remaining concentration of ST09 and ST10 at designed time points during the HLM incubation. Six major metabolites of cytochrome P450 were simultaneously measured with LC-MS/MS, and the inhibitory effects of ST09 and ST10 were respectively evaluated with IC50 values. RESULTS: ST09 was extremely unstable in vitro, and t1/2 was less than 1 min. However, ST10, the major metabolite of ST09, was NADPH-independent metabolized in HLMs, while its t1/2 and microsomal intrinsic clearance (CLint) were 32 min and 0.043 mL·min-1·mg(protein)-1, respectively. IC50 values of ST09 and ST10 on CYP3A4 (midazolam as substrate), CYP3A4 (testosterone as substrate), CYP1A2, CYP2C9, CYP2C19 and CYP2D6 were 0.42/0.25, 1.27/0.81, 24.92/18.21, 36.53/54.34, 67.64/144.90, 6.43/5.30 μmol·L-1, respectively. CONCLUSION: ST09 and ST10 are extensively metabolized in vitro and both compounds had significant inhibition on CYP3A4 and CYP2D6.

Anti-pepsin activity of silicon dioxide nanoparticles / Actividad antipepsina de nanopartículas de dióxido de silicio / Atividade antipepsina de nanopartículas de dióxido de silício

SiO2NPs as an inhibitor of pepsin enzyme for treatment of gastro-esophageal reflux disease (GERD) were investigated. Silicon dioxide nanoparticles (pepsin coated SiO2NPs) are among the safest nanoparticles that can be used inside the human body. The activity of pepsin before and after the addition of certain amounts of the NPs to the reaction mixture was measured spectrophotometrically. Furthermore, these experiments were repeated at different temperatures, different weights of NPs, and different ionic strengths. The kinetic parameters (Km & Vmax) of the pepsin-catalyzed reactions were calculated from the Lineweaver-Burk plots. The results showed that there is a significant reduction of pepsin activity by SiO2NPs (Vmax of free pepsin = 4.82 U and Vmax of the immobilized pepsin = 2.90 U). The results also indicated that the presence of ionic strength causes remarkable reduction of pepsin activity. It can be concluded the best condition for inhibition of pepsin activity is by using a combinationof SiO2NPs and high concentration NaCl at 37 °C.

Se usaron nanopartículas de dióxido de silicio como inhibidores de la pepsina para el tratamiento del reflujo gastroesofágico (GERD). Estas nanopartículas (SiO2NPs recubiertas de pepsina) son unas de las más seguras y pueden usarse en el cuerpo humano. Se midió a través de espectrofotometría la actividad de la pepsina antes y después de añadir cierta cantidad de NPs a la mezcla reactante. Adicionalmente, se repitieron estas pruebas a diferentes temperaturas, variando el peso de las NPs y la fuerza iónica. Se calcularon los parámetros cinéticos (Km y Vmax) de las reacciones catalizadas con pepsina a través de las gráficas de Lineweaver-Burk. Los resultados mostraron que, usando SiO2NPs (Vmax de pepsina libre = 4.82 U y Vmax de pepsina inmovilizada = 2.90 U) y a través de la presencia de fuerza iónica, la actividad enzimática se reduce significativamente. Se concluye que la mejor condición para inhibir la actividad enzimática es usando una combinación de SiO2NPs y una alta concentración de NaCl a 37 °C.

Foram usadas nanopartículas de dióxido de silício como inibidores da pepsina para o tratamento do refluxo gastroesofágico (GERD). Estas nanopartículas (SiO2NPs cobertas de pepsina) são uma das mais seguras e podem usar-se no corpo humano. Foi medida a atividade da pepsina mediante espectrofotometria antes e depois de agregar certa quantidade de NPs à mistura de reação. Adicionalmente, repetiram-se estas provas a diferentes temperaturas, variando o peso das NPs e a força iónica. Foram calculados os parâmetros cinéticos (Km e Vmax) das reações catalisadas com pepsina a través das gráficas de Lineweaver-Burk. Os resultados mostraram que, usando SiO2NPs (Vmax de pepsina livre = 4.82 U e Vmax de pepsina imobilizada = 2.90 U) e a través da presença de força iónica, a atividade enzimática se reduze significativamente. Foi concluído que a melhor condição para inibir a atividade enzimática é usando uma combinação de SiO2NPs e uma alta concentração de NaCl a 37 °C.

Effects of caffeine, tea polyphenol and daidzein on the pharmacokinetics of lansoprazole and its metabolites in rats

abstract The aim of this study was to evaluate the effects of caffeine, tea polyphenol and daidzein on the pharmacokinetics of lansoprazole and its metabolites. Rats were intragastrically administered caffeine (30 mg·kg-1, once per day), tea polyphenol (400 mg·kg-1, once per day) or daidzein (13.5 mg·kg-1, once per day) for 14 days, followed by an intragastric administration of lansoprazole (8 mg·kg-1) on the 15th day. The plasma concentrations of lansoprazole and its two primary metabolites, 5-hydroxylansoprazole and lansoprazole sulfone, were determined by high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). Tea polyphenol significantly elevated the Area Under the Curve (AUC) of lansoprazole from 680.29 ± 285.99 to 949.76 ± 155.18 μg/L.h and reduced that of lansoprazole sulfone from 268.82 ± 82.37 to 177.72 ± 29.73 μg/L.h. Daidzein increased the AUC of lansoprazole from 680.29 ± 285.99 to 1130.44 ± 97.6 μg/L.h and decreased that of lansoprazole sulfone from 268.82 ± 82.37 to 116.23 ± 40.14 μg/L.h. The pharmacokinetics of 5-hydroxylansoprazole remained intact in the presence of tea polyphenol or daidzein. Caffeine did not affect the pharmacokinetics of lansoprazole and its metabolites. The results imply that tea polyphenol and daidzein may inhibit the in vivo metabolism of lansoprazole by suppressing CYP3A.

resumo O objetivo deste estudo foi avaliar os efeitos da cafeína, do polifenol do chá e da daidzeína na farmacocinética do lansoprazol e de seus metabólitos. Administraram-se, intragastricamente, aos ratos cafeína (30 mg·kg-1, uma vez ao dia), polifenol do chá(400 mg·kg-1, uma vez ao dia) ou daidzeína (13,5 mg·kg-1, uma vez ao dia), por 14 dias, seguindo-se a administração de lansoprazol (8 mg·kg-1) no 15º. dia. As concentrações plasmáticas do lansoprazol e de seus dois metabólitos primários, 5-hidroxilansoprazol e sulfona de lansoprazol, foram determinadas por cromatografia líquida de alta eficiência acoplada com espectrometria de massas (CLAE-EM/EM). O polifenol do chá elevou, significativamente, a Área Sob a Curva (ASC) do lansoprazol de 680,29 ± 285,99 para 949,76 ± 155,18 μg/L.h e reduziu a da sulfona de lansoprazol de 268,82 ± 82,37 para 177,72 ± 29,73 μg/L.h. A daidzeína aumentou a ASC do lansoprazol de 680,29 ± 285,99 para 1130,44 ± 97,6 μg/L.h e reduziu a da sulfona de lansoprazol de 268,82 ± 82,37 para 177,72 ± 29,73 μg/L.h. A farmacocinética do 5-hidroxilansoprazol permaneceu intacta na presença de polifenol do chá ou daidzeína. A cafeína não afetou a farmacocinética do lansoprazol e de seus metabólitos. Os resultados sugerem que o polifenol do chá e a daidzeína podem inibir o metabolismo in vivo do lansoprazol por supressão da CYP3A.

In vitro inhibitory effects of plumbagin, the promising antimalarial candidate, on human cytochrome P450 enzymes

OBJECTIVE@#To investigate the propensity of plumbagin to inhibit the three isoforms of human cytochrome P450 (CYP), i.e., CYP1A2, CYP2C19, and CYP3A4 using human liver microsomes in vitro.@*METHODS@#Inhibitory effects of plumbagin on the three human CYP isoforms were investigated using pooled human liver microsomes. Phenacetin O-deethylation, omeprazole hydroxylation and nifedipine oxidation were used as selective substrates for CYP1A2, CYP2C19 and CYP3A4 activities, respectively. Concentrations of paracetamol, 5-hydroxyomeprazole, and oxidized nifedipine were determined in microsomal incubation mixture using high-performance liquid chromatography.@*RESULTS@#Plumbagin showed significant inhibitory effects on all CYP isoforms, but with the most potent activity on CYP2C19-mediated omeprazole hydroxylation. The IC50 (concentration that inhibits enzyme activity by 50%) values of plumbagin and nootkatone (selective inhibitor) for CYP2C19 were (0.78 ± 0.01) and (27.31 ± 0.66) μM, respectively. The inhibitory activities on CYP1A2-mediated phenacetin O-deethylation and CYP3A4-mediated nifedipine oxidation were moderate. The IC50 values of plumbagin and α-naphthoflavone (selective inhibitor) for CYP1A2 were (1.39 ± 0.01) and (0.02 ± 0.36) μM, respectively. The corresponding IC50 values of plumbagin and ketoconazole (selective inhibitor) for CYP3A4 were (2.37 ± 0.10) and (0.18 ± 0.06) μM, respectively.@*CONCLUSIONS@#Clinical relevance of the interference of human drug metabolizing enzymes should be aware of for further development scheme of plumbagin as antimalarial drug when used in combination with other antimalarial drugs which are metabolized by these CYP isoforms.

Inhibitory and inductive effects of Polygonum capitatum water extract on CYP450 / 中国药理学通报

Aim To evaluate the inhibitive and induc-tive effects of Polygonum capitatum water extract on main cytochrome P450 isoforms in human and liver mi-crosomes of mouse in vitro for predicting the herb-drug interactions in clinical application. Methods The in vitro inhibitory effect was evaluated by incubating Po-lygonum capitatum water extract with the probe sub-strates of main phase I metabolic enzymes in human liver microsomes, including CYP1A2, CYP2E1, CYP2C9,CYP2C19 and CYP3A4. Mice were adminis-tered with Polygonum capitatum water extract at dosage of 0 . 58 g · kg-1 and 1 . 16 g · kg-1 by gastric lavage for successive 7 days and 14 days, then the cocktail-LC-MS/MS method was applied to assess the inductive effect of main CYP450 isoforms in mouse liver micro-somes. Results The IC50 values of Polygonum capita-tum water extract on main CYP450 isoforms in human liver microsomes were from 849 . 6 mg · L-1 to 2 287 mg·L-1 . Compared with the blank control group, the activites of CYP2C9 and CYP3A4 in 1. 16 g·kg-1 7 d group were about 49 . 9 % and 21. 1 % higher ( P <0. 01, P < 0. 05 ) respectively, the activities of CYP2C9 and CYP3A4 in 0. 58 g·kg-1 7 d group were 27. 6 % and 15. 5 % higher ( P <0. 01 , P <0. 05 ) respectively, the activities of CYP2C9 and CYP3A4 in 1. 16 g·kg-1 14 d group were 67. 5 % and 32. 1 %higher (P<0. 01) respectively, while the activities of CYP1 A2 , CYP2 E1 and CYP2 C19 were not increased significantly in Polygonum capitatum treatment group. Conclusions Polygonum capitatum water extract do not show the inhibitory effect on main CYP450 in hu-man liver microsomes. There is induction on CYP2C9 and CYP3 A4 in mouse liver microsomes by Polygonum capitatum water extract.

In vitro inhibitory effects of plumbagin, the promising antimalarial candidate, on human cytochrome P450 enzymes

Objective: To investigate the propensity of plumbagin to inhibit the three isoforms of human cytochrome P450 (CYP), i.e., CYP1A2, CYP2C19, and CYP3A4 using human liver microsomes in vitro. Methods: Inhibitory effects of plumbagin on the three human CYP isoforms were investigated using pooled human liver microsomes. Phenacetin O-deethylation, omeprazole hydroxylation and nifedipine oxidation were used as selective substrates for CYP1A2, CYP2C19 and CYP3A4 activities, respectively. Concentrations of paracetamol, 5-hydroxyomeprazole, and oxidized nifedipine were determined in microsomal incubation mixture using high-performance liquid chromatography. Results: Plumbagin showed significant inhibitory effects on all CYP isoforms, but with the most potent activity on CYP2C19-mediated omeprazole hydroxylation. The IC

Phenolic composition and inhibitory activity of Mangifera indica and Mucuna urens seeds extracts against key enzymes linked to the pathology and complications of type 2 diabetes

Objective: To investigate the phenolic compounds composition and the inhibitory activity ofMangifera indica (M. indica) and Mucuna urens (M. urens) seeds extracts against some key enzymes (α-amylase, α-glucosidase and aldose reductase) implicated in the pathology and complications of type 2 diabetes in vitro. Methods: Reverse phase chromatographic quantification of the major flavonoids and phenolic acids in the seeds extracts was carried out using high performance liquid chromatography coupled with diode array detection. The inhibitory activities of the seeds extracts against α-amylase andα-glucosidase were estimated using soluble starch and ρ-nitrophenylglucopyranoside as their respective substrates. Inhibition of aldose reductase activity by the extracts was assayed using partially purified lens homogenate of normal male rat as source of enzyme; inhibition of Fe2+-induced lipid peroxidation by extracts was tested in rat pancreas homogenate.Results:The chromatography result revealed that extracts of both seeds had appreciable levels of some major flavonoids and phenolic acids of pharmacological importance, including gallic acid, chlorogenic acid, caffeic acid, ellagic acid, catechin, rutin, quercitrin, quercetin and kaempferol. Extracts of both seeds effectively inhibited α-amylase, α-glucosidase and aldose reductase activities in a dose-dependent manner, having inhibitory preference for these enzymes in the order of aldose reductase>α-glucosidase>α-amylase. With lower half-maximal inhibitory concentrations (IC50) against α-amylase, α-glucosidase, and aldose reductase, M. indica had stronger inhibitory potency against these enzymes than M. urens. Extracts of both seeds also inhibited Fe2+-induced lipid peroxidation in a dose-dependent pattern, with M. indica being more potent than M. urens.Conclusions:The results obtained provide support for a possible use of M. indica and M. urens seeds in managing hyperglycemia and preventing the complications associated with it in type 2 diabetes.