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Pharmaceutical Biology Dec 2017Codeine, also known as 3-methylmorphine, is an opiate used to treat pain, as a cough medicine and for diarrhoea. No study on the effects of codeine on the metabolic...
CONTEXT
Codeine, also known as 3-methylmorphine, is an opiate used to treat pain, as a cough medicine and for diarrhoea. No study on the effects of codeine on the metabolic capacity of CYP enzyme is reported.
OBJECTIVE
In order to investigate the effects of codeine on the metabolic capacity of cytochrome P450 (CYP) enzymes, a cocktail method was employed to evaluate the activities of CYP2B1, CYP2D1, CYP1A2, CYP3A2 and CYP2C11.
MATERIALS AND METHODS
Sprague-Dawley rats were randomly divided into codeine group (low, medium, high) and control group. The codeine group rats were given 4, 8, 16 mg/kg (low, medium, high) codeine by continuous intragastric administration for 14 days. Five probe drugs bupropion, metroprolol, phenacetin, midazolam and tolbutamide were given to rats through intragastric administration, and the plasma concentrations were determined by UPLC-MS/MS.
RESULTS AND CONCLUSION
The pharmacokinetic parameters of bupropion and metroprolol experienced obvious change with AUC, C increased and CL decreased for bupropion in medium dosage group and midazolam low dosage group. This result indicates that the 14 day-intragastric administration of codeine may inhibit the metabolism of bupropion (CYP2B1) and midazolam (CYP3A2) in rat. Additional, there are no statistical differences for albumin (ALB), alkaline phosphatase (ALP), creatinine (Cr) after 14 intragastric administration of codeine, while alanine aminotransferase (ALT), aspartate aminotransferase (AST), uric acid (UA) increased compared to control group. The biomedical test results show continuous 14 day-intragastric administration of codeine would cause liver damage.
Topics: Animals; Bupropion; Codeine; Cytochrome P-450 Enzyme System; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Activation; Isoenzymes; Male; Random Allocation; Rats; Rats, Sprague-Dawley; Tolbutamide
PubMed: 28253826
DOI: 10.1080/13880209.2017.1297466 -
Journal of Advanced Pharmaceutical... 2017A lot of pharmaceutical substances have a poor solubility that limits their absorption and distribution to the targeted sites to elicit the desired action without...
A lot of pharmaceutical substances have a poor solubility that limits their absorption and distribution to the targeted sites to elicit the desired action without causing untoward effects on healthy cells or tissues. For such drugs, new modes of delivery have to be developed for efficient and effective delivery of the drug to the target site. Formation of magnetically active solid dispersion of such drugs could be a useful approach to addressing this problem because they combine targeted delivery and good solubility. In this work, the distribution of superparamagnetic nanoparticles in the solid dispersion of polyethylene glycol with average molecular weight 950-1050 g/mol and phenacetin was studied using atomic force and magnetic force microscopy. The distribution of nanoparticles was found to be uniform in studied composites. Magnetically active solid dispersions may find application in the production of the capsulated drug delivery systems with enhanced solubility parameters.
PubMed: 28217547
DOI: 10.4103/2231-4040.197331 -
Advanced Biomedical Research 2016A very simple, sensitive, and accurate high-performance liquid chromatography (HPLC) method with ultraviolet detector was developed and applied to determine...
A simple and sensitive high-performance liquid chromatography method for determination of ciprofloxacin in bioavailability studies of conventional and gastroretentive prolonged-release formulations.
BACKGROUND
A very simple, sensitive, and accurate high-performance liquid chromatography (HPLC) method with ultraviolet detector was developed and applied to determine ciprofloxacin in human plasma following administration of a gastroretentive formulation developed in our laboratory.
MATERIALS AND METHODS
HPLC analysis was performed on a C μ-Bondapack column (250 mm × 3.9 mm) using acetonitrile: potassium dihydrogen phosphate solution 0.1 M (20:80, v/v, pH 3) at a flow rate of 1.5 ml/min and eluate was monitored at 276 nm. After addition of phenacetin as internal standard, plasma samples were treated with 0.1 M phosphate buffer (pH: 7) and followed by extraction with dichloromethane. The method was validated for linearity, precision, accuracy, limit of quantitation (LOQ), robustness, stability, and applied in bioavailability studies of our developed gastroretentive formulation in healthy volunteers.
RESULTS
The calibration curves were linear over the concentration range 0.025-4 μg/ml with the detection limit of 15 ng/ml. Accuracy % were within 93-115 and the coefficient of variance % ranged from 0.20 to 12.8. The very low LOQ (25 ng/ml) allowed avoiding fluorometric detection which is more expensive and is not available in all laboratories. Ciprofloxacin was stable in samples with no evidence of degradation during 3 freeze-thaw cycles and 3 months storage at -70°C.
CONCLUSION
This validated HPLC method was successfully used for the determination of ciprofloxacin in human plasma following oral administration of controlled release formulation, conventional immediate-release tablets and when administered concomitantly with divalent and trivalent cations such as aluminum-, magnesium-, or calcium-containing products under which the bioavailability of ciprofloxacin is significantly reduced.
PubMed: 27995102
DOI: 10.4103/2277-9175.190995 -
The Journal of Pharmacology and... Jan 2017In this work, we first describe the population variability in hepatic drug metabolism using cryopreserved hepatocytes from five different donors cultured in a perfused...
In this work, we first describe the population variability in hepatic drug metabolism using cryopreserved hepatocytes from five different donors cultured in a perfused three-dimensional human liver microphysiological system, and then show how the resulting data can be integrated with a modeling and simulation framework to accomplish in vitro-in vivo translation. For each donor, metabolic depletion profiles of six compounds (phenacetin, diclofenac, lidocaine, ibuprofen, propranolol, and prednisolone) were measured, along with metabolite formation, mRNA levels of 90 metabolism-related genes, and markers of functional viability [lactate dehydrogenase (LDH) release, albumin, and urea production]. Drug depletion data were analyzed with mixed-effects modeling. Substantial interdonor variability was observed with respect to gene expression levels, drug metabolism, and other measured hepatocyte functions. Specifically, interdonor variability in intrinsic metabolic clearance ranged from 24.1% for phenacetin to 66.8% for propranolol (expressed as coefficient of variation). Albumin, urea, LDH, and cytochrome P450 mRNA levels were identified as significant predictors of in vitro metabolic clearance. Predicted clearance values from the liver microphysiological system were correlated with the observed in vivo values. A population physiologically based pharmacokinetic model was developed for lidocaine to illustrate the translation of the in vitro output to the observed pharmacokinetic variability in vivo. Stochastic simulations with this model successfully predicted the observed clinical concentration-time profiles and the associated population variability. This is the first study of population variability in drug metabolism in the context of a microphysiological system and has important implications for the use of these systems during the drug development process.
Topics: Cryopreservation; Cytochrome P-450 Enzyme System; Hepatocytes; Humans; L-Lactate Dehydrogenase; Liver; Perfusion; Pharmaceutical Preparations; Phenotype; Serum Albumin; Tissue Culture Techniques; Tissue Distribution
PubMed: 27760784
DOI: 10.1124/jpet.116.237495 -
Genes and Environment : the Official... 2016Toxicogenomics is a rapidly developing discipline focused on the elucidation of the molecular and cellular effects of chemicals on biological systems. As a collaborative... (Review)
Review
Toxicogenomics is a rapidly developing discipline focused on the elucidation of the molecular and cellular effects of chemicals on biological systems. As a collaborative study group of Toxicogenomics/JEMS·MMS, we conducted studies on hepatocarcinogens in rodent liver in which 100 candidate marker genes were selected to discriminate genotoxic hepatocarcinogens from non-genotoxic hepatocarcinogens. Differential gene expression induced by 13 chemicals were examined using DNA microarray and quantitative real-time PCR (qPCR), including eight genotoxic hepatocarcinogens [o-aminoazotoluene, chrysene, dibenzo[a,l]pyrene, diethylnitrosamine (DEN), 7,12-dimethylbenz[a]anthracene, dimethylnitrosamine, dipropylnitrosamine and ethylnitrosourea (ENU)], four non-genotoxic hepatocarcinogens [carbon tetrachloride, di(2-ethylhexyl)phthalate (DEHP), phenobarbital and trichloroethylene] and a non-genotoxic non-hepatocarcinogen [ethanol]. Using qPCR, 30 key genes were extracted from mouse livers at 4 h and 28 days following dose-dependent gene expression alteration induced by DEN and ENU: the most significant changes in gene expression were observed at 4 h. Next, we selected key point times at 4 and 48 h from changes in time-dependent gene expression during the acute phase following administration of chrysene by qPCR. We successfully showed discrimination of eight genotoxic hepatocarcinogens [2-acetylaminofluorene, 2,4-diaminotoluene, diisopropanolnitrosamine, 4-dimethylaminoazobenzene, 4-(methylnitsosamino)-1-(3-pyridyl)-1-butanone, N-nitrosomorpholine, quinoline and urethane] from four non-genotoxic hepatocarcinogens [1,4-dichlorobenzene, dichlorodiphenyltrichloroethane, DEHP and furan] using qPCR and principal component analysis. Additionally, we successfully identified two rat genotoxic hepatocarcinogens [DEN and 2,6-dinitrotoluene] from a nongenotoxic-hepatocarcinogen [DEHP] and a non-genotoxic non-hepatocarcinogen [phenacetin] at 4 and 48 h. The subsequent gene pathway analysis by Ingenuity Pathway Analysis extracted the DNA damage response, resulting from the signal transduction of a p53-class mediator leading to the induction of apoptosis. The present review of these studies suggests that application of principal component analysis on the gene expression profile in rodent liver during the acute phase is useful to predict genotoxic hepatocarcinogens in comparison to non-genotoxic hepatocarcinogens and/or non-carcinogenic hepatotoxins.
PubMed: 27482301
DOI: 10.1186/s41021-016-0041-0 -
Basic & Clinical Pharmacology &... Jan 2017Conventional in vitro human hepatic models for drug testing are based on the use of standard cell lines derived from hepatomas or primary human hepatocytes (PHHs).... (Comparative Study)
Comparative Study
Human Hepatic HepaRG Cells Maintain an Organotypic Phenotype with High Intrinsic CYP450 Activity/Metabolism and Significantly Outperform Standard HepG2/C3A Cells for Pharmaceutical and Therapeutic Applications.
Conventional in vitro human hepatic models for drug testing are based on the use of standard cell lines derived from hepatomas or primary human hepatocytes (PHHs). Limited availability, interdonor functional variability and early phenotypic alterations in PHHs restrict their use, whilst standard cell lines such as HepG2 lack a substantial and variable set of liver-specific functions such as CYP450 activity. Alternatives include the HepG2-derivative C3A cells selected as a more differentiated and metabolically active hepatic phenotype. Human HepaRG cells are an alternative organotypic co-culture model of hepatocytes and cholangiocytes reported to maintain in vivo-like liver-specific functions, including intact Phase I-III drug metabolism. In this study, we compared C3A and human HepaRG cells using phenotypic profiling, CYP450 activity and drug metabolism parameters to assess their value as hepatic models for pre-clinical drug testing or therapeutics. Compared with C3As, HepaRG co-cultures exhibit a more organotypic phenotype, including evidence of hepatic polarity with the strong expression of CYP3A4, the major isoform involved in the metabolism of over 60% of marketed drugs. Significantly greater CYP450 activity and expression of CYP1A2, CYP2E1 and CYP3A4 genes in HepaRG cells (comparable with that of human liver tissue) was demonstrated. Moreover, HepaRG cells also preferentially expressed the hepatic integrin α β - an important modulator of cell behaviour including growth and survival, differentiation and polarity. Drug metabolite profiling of phenacetin (CYP1A2) and testosterone (CYP3A4) using LC-MS/MS and HPLC, respectively, revealed that HepaRGs had more intact (Phase I-II) metabolism profile. Thus, HepaRG cells significantly outperform C3A cells for the potential pharmaceutical and therapeutic applications.
Topics: Animal Testing Alternatives; Bile Ducts; Cell Differentiation; Cell Line; Coculture Techniques; Cytochrome P-450 Enzyme System; Drug Evaluation, Preclinical; Epithelial Cells; Gene Expression Regulation, Enzymologic; Hep G2 Cells; Hepatocytes; Humans; Metabolic Detoxication, Phase I; Metabolic Detoxication, Phase II; Phenacetin; Testosterone
PubMed: 27285124
DOI: 10.1111/bcpt.12631 -
Molecules (Basel, Switzerland) Apr 2016Aschantin is a bioactive neolignan found in Magnolia flos with antiplasmodial, Ca(2+)-antagonistic, platelet activating factor-antagonistic, and chemopreventive...
Aschantin is a bioactive neolignan found in Magnolia flos with antiplasmodial, Ca(2+)-antagonistic, platelet activating factor-antagonistic, and chemopreventive activities. We investigated its inhibitory effects on the activities of eight major human cytochrome P450 (CYP) and uridine 5'-diphospho-glucuronosyltransferase (UGT) enzymes of human liver microsomes to determine if mechanistic aschantin-enzyme interactions were evident. Aschantin potently inhibited CYP2C8-mediated amodiaquine N-de-ethylation, CYP2C9-mediated diclofenac 4'-hydroxylation, CYP2C19-mediated [S]-mephenytoin 4'-hydroxylation, and CYP3A4-mediated midazolam 1'-hydroxylation, with Ki values of 10.2, 3.7, 5.8, and 12.6 µM, respectively. Aschantin at 100 µM negligibly inhibited CYP1A2-mediated phenacetin O-de-ethylation, CYP2A6-mediated coumarin 7-hydroxylation, CYP2B6-mediated bupropion hydroxylation, and CYP2D6-mediated bufuralol 1'-hydroxylation. At 200 µM, it weakly inhibited UGT1A1-catalyzed SN-38 glucuronidation, UGT1A6-catalyzed N-acetylserotonin glucuronidation, and UGT1A9-catalyzed mycophenolic acid glucuronidation, with IC50 values of 131.7, 144.1, and 71.0 µM, respectively, but did not show inhibition against UGT1A3, UGT1A4, or UGT2B7 up to 200 µM. These in vitro results indicate that aschantin should be examined in terms of potential interactions with pharmacokinetic drugs in vivo. It exhibited potent mechanism-based inhibition of CYP2C8, CYP2C9, CYP2C19, and CYP3A4.
Topics: Benzodioxoles; Cytochrome P-450 Enzyme Inhibitors; Enzyme Inhibitors; Glucuronosyltransferase; Humans; Lignans; Microsomes, Liver; Time Factors
PubMed: 27128896
DOI: 10.3390/molecules21050554 -
Acta Pharmacologica Sinica May 2016The substrate cocktail is frequently used to evaluate cytochrome P450 (CYP) enzyme-mediated drug interactions and potential interactions among the probe substrates....
AIM
The substrate cocktail is frequently used to evaluate cytochrome P450 (CYP) enzyme-mediated drug interactions and potential interactions among the probe substrates. Here, we re-optimized the substrate cocktail method to increase the reliability and accuracy of screening for candidate compounds and expanded the method from a direct CYP inhibition assay to a time-dependent inhibition (TDI) assay.
METHODS
In the reaction mixtures containing human liver microsome (0.1 mg/mL), both the concentrations of a substrate cocktail (phenacetin for 1A2, coumarin for 2A6, bupropion for 2B6, diclofenac for 2C9, dextromethorphan for 2D6, and testosterone for 3A4) and the incubation time were optimized. Metabolites of the substrate probes were simultaneously analyzed by multiple-reaction monitoring (MRM) using a routine LC/MS/MS. Direct CYP inhibition was validated using 7 inhibitors (α-naphthoflavone, tranylcypromine, ticlopidine, fluconazole, quinidine, ketoconazole and 1-ABT). The time-dependent inhibition was partially validated with 5 inhibitors (ketoconazole, verapamil, quinidine, paroxetine and 1-ABT).
RESULTS
The inhibition curve profiles and IC50 values of 7 CYP inhibitors were approximate when a single substrate and the substrate cocktail were tested, and were consistent with the previously reported values. Similar results were obtained in the IC50 shifts of 5 inhibitors when a single substrate and the substrate cocktail were tested in the TDI assay.
CONCLUSION
The 6-in-1 substrate cocktail (for 1A2, 2A6, 2B6, 2C9, 2D6 and 3A) is reliable for assessing CYP inhibition and time-dependent inhibition of drug candidates.
Topics: Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Drug Evaluation, Preclinical; Drug Interactions; Humans; Microsomes, Liver; Substrate Specificity; Time Factors
PubMed: 27063220
DOI: 10.1038/aps.2016.10 -
Chinese Medicine 2016Sodium tanshinone IIA sulfonate (STS) is a water-soluble derivative of tanshinone IIA for treating cardiovascular disorders. The roles of cytochrome P450 enzymes (CYPs)...
BACKGROUND
Sodium tanshinone IIA sulfonate (STS) is a water-soluble derivative of tanshinone IIA for treating cardiovascular disorders. The roles of cytochrome P450 enzymes (CYPs) in the metabolism of STS have remained unclear. This study aims to screen the main CYPs for metabolism of STS and study their interactions in vitro.
METHODS
Seven major CYPs were screened for metabolism of STS by human liver microsomes (HLMs) or recombinant CYP isoforms. Phenacetin (CYP1A2), coumarin (CYP2A6), tolbutamide (CYP2C9), metoprolol (CYP2D6), chlorzoxazone (CYP2E1), S-mephenytoin (CYP2C19), and midazolam (CYP3A4) were used as probe substrates to determine the potential of STS in affecting CYP-mediated phase I metabolism in humans. Enzyme kinetic studies were performed to investigate the modes of inhibition of the enzyme-substrate interactions by GraphPad Prism Enzyme Kinetic 5 Demo software.
RESULTS
Sodium tanshinone IIA sulfonate inhibited the activity of CYP3A4 in a dose-dependent manner by the HLMs and CYP3A4 isoform. The K m and V max values of STS were 54.8 ± 14.6 µM and 0.9 ± 0.1 nmol/mg protein/min, respectively, for the HLMs and 7.5 ± 1.4 µM and 6.8 ± 0.3 nmol/nmol P450/min, respectively, for CYP3A4. CYP1A2, CYP2A6, CYP2C9, CYP2D6, CYP2E1, and CYP2C19 showed minimal or no effects on the metabolism of STS.
CONCLUSION
This in vitro study showed that STS mainly inhibited the activities of CYP3A4.
PubMed: 27006687
DOI: 10.1186/s13020-016-0083-z -
Evidence-based Complementary and... 2016Bakuchicin is a furanocoumarin isolated from Psoralea corylifolia and shows several biological activities. Although there have been studies on the biological effects of...
Bakuchicin is a furanocoumarin isolated from Psoralea corylifolia and shows several biological activities. Although there have been studies on the biological effects of bakuchicin, its modulation potency of CYP activities has not been previously investigated. Here, we investigated the inhibitory effects of bakuchicin on the activities of CYP isoforms by using a cocktail of probe substrates in pooled human liver microsomes (HLMs) and human recombinant cDNA-expressed CYP. Bakuchicin strongly inhibited CYP1A-mediated phenacetin O-deethylation with an IC50 value of 0.43 μM in HLMs. It was confirmed by human recombinant cDNA-expressed CYP1A1 and CYP1A2 with a K i value of 0.11 μM and 0.32 μM, respectively. A Lineweaver-Burk plot indicated that the inhibition mechanism of bakuchicin was competitive inhibition. Overall, this is the first study to investigate the potential CYP1A1 and CYP1A2 inhibition associated with bakuchicin and to report its competitive inhibitory effects on HLMs.
PubMed: 26977174
DOI: 10.1155/2016/5198743