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Alcohol (Fayetteville, N.Y.) Nov 2021Alcohol abuse has become a serious health issue worldwide. Ketamine can reduce addiction risk among patients with alcohol use disorders. This study aimed to determine...
BACKGROUND
Alcohol abuse has become a serious health issue worldwide. Ketamine can reduce addiction risk among patients with alcohol use disorders. This study aimed to determine the effects of alcohol on the pharmacokinetics of ketamine during long-term alcohol exposure.
METHOD
An ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for determination of ketamine and norketamine was developed and validated. A total of 15 rats were given 40% alcohol for 3 weeks. The pharmacokinetics of ketamine were measured at time zero, 1 week, 2 weeks, and 3 weeks after alcohol exposure. The metabolic capability of liver CYP450 was evaluated using three probe drugs: metoprolol, phenacetin, and tolbutamide.
RESULTS
During drinking of 40% alcohol, the AUC(0-t), AUC(0-∞), and Cmax of ketamine and norketamine significantly increased, while V and CL significantly decreased with time (p < 0.001). The pharmacokinetic changes of norketamine were highly consistent with ketamine. Additionally, the concentration ratio of norketamine/ketamine in sample time also decreased over time. However, there were no pharmacokinetic changes of three probe drugs, which indicated there was no significant change of liver CYPs activities.
CONCLUSION
Alcohol significantly increases plasma concentration of ketamine and norketamine. The effect of alcohol on pharmacokinetics of ketamine should be considered in clinical therapy.
Topics: Alcoholism; Animals; Chromatography, Liquid; Humans; Ketamine; Rats; Tandem Mass Spectrometry
PubMed: 33549609
DOI: 10.1016/j.alcohol.2021.01.008 -
European Journal of Pharmaceutical... Jun 2021The cynomolgus monkey is a nonhuman primate that is often used for pharmacokinetic and toxicokinetic studies of new chemical entities. Species differences in drug...
The cynomolgus monkey is a nonhuman primate that is often used for pharmacokinetic and toxicokinetic studies of new chemical entities. Species differences in drug metabolism are obstacles for the extrapolation of animal data to humans. This study aimed to characterize hydrolase activities for typical compounds by cynomolgus monkey liver microsomes and recombinant monkey carboxylesterases (CES1 and CES2) and arylacetamide deacetylase (AADAC) compared with the activities in humans. To estimate the contribution of each hydrolase, the ratios of the expression level of each hydrolase in the liver microsomes and recombinant systems were used. For almost all of the tested human CES1 substrates, hydrolase activities in cynomolgus monkey liver microsomes tended to be lower than those in human liver microsomes, and recombinant cynomolgus monkey CES1 showed catalytic activity, but not for all substrates. For human CES2 substrates, hydrolase activities in cynomolgus monkey liver were higher than those in human liver microsomes, and recombinant monkey CES2 was responsible for their hydrolysis. Among human AADAC substrates, phenacetin was mainly hydrolyzed by monkey AADAC, whereas indiplon and ketoconazole were hydrolyzed by AADAC and other unknown enzymes. Flutamide was hydrolyzed by monkey CES2, not by AADAC. Rifamycins were hardly hydrolyzed in monkey liver microsomes. In conclusion, this study characterized the hydrolase activities of cynomolgus monkeys compared with those in humans. The findings would be helpful for pharmacokinetic or toxicokinetic studies of new chemical entities whose main metabolic pathway is hydrolysis.
Topics: Animals; Carboxylic Ester Hydrolases; Flutamide; Hydrolases; Hydrolysis; Liver; Macaca fascicularis; Microsomes, Liver
PubMed: 33722734
DOI: 10.1016/j.ejps.2021.105807 -
Biopharmaceutics & Drug Disposition Jan 2021CYP1A2 is one of the main Cytochrome P450 enzymes in the human liver associated with the metabolism of several xenobiotics. CYP1A2 is especially involved in the...
CYP1A2 is one of the main Cytochrome P450 enzymes in the human liver associated with the metabolism of several xenobiotics. CYP1A2 is especially involved in the metabolic activation of different procarcinogens. Therefore, the development of cancer may be inhibited by inhibiting CYP1A2 activity. Here, the inhibitory effect of HYIpro-3-1 and its derivatives on CYP1A2 activity in human liver microsomes (HLM) was studied through LC-MS/MS using a cocktail assay. Among the four compounds, HYIpro-3-1 showed the most selective and strongest inhibitory effect on CYP1A2 at IC values of 0.1 µM in HLMs and inhibition was confirmed using purified human CYP1A2. It was determined that inhibition is reversible because the inhibitory effect of HYIpro-3-1 is not dependent on preincubation time. HYIpro-3-1 showed a typical pattern of competitive inhibition for CYP1A2-catalyzed phenacetin O-deethylation, based on the Lineweaver-Burk plot, with a Ki value of 0.05 μM in HLMs; the secondary plot also showed a linear pattern. In our study, HYIpro-3-1 was proposed as a novel inhibitor with the capacity to selectively inhibit CYP1A activity in HLMs.
Topics: Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP1A2 Inhibitors; Humans; Microsomes, Liver
PubMed: 33386627
DOI: 10.1002/bdd.2259 -
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi =... Aug 2022Primary human hepatocytes (PHH) are the gold standard of human liver model for drug screening. However, a problem of culturing PHH is the rapid decline of cytochrome...
Primary human hepatocytes (PHH) are the gold standard of human liver model for drug screening. However, a problem of culturing PHH is the rapid decline of cytochrome P450 (CYP450) activity, which plays an important role in drug metabolism. In this study, thermo-responsive culture dishes were used to explore the conditions for murine embryonic 3T3-J2 fibroblasts to form cell sheet. Based on the cell sheet engineering technology, a three-dimensional (3D) "sandwich" co-culture system of 3T3-J2 cell sheet/PHH/collagen gel was constructed. The tissue structure and protein expression of the model section were observed by hematoxylin eosin staining and immunofluorescence staining respectively. Phenacetin and bupropion were used as substrates to determine the activity of CYP450. The contents of albumin and urea in the system were determined by enzyme linked immunosorbent assay (ELISA). The results showed that the complete 3T3-J2 cell sheet could be obtained when the cell seeding density was 1.5×10 /dish (35 mm dish) and the incubation time at low temperature was 60 min. Through cell sheet stacking, a 3D liver model was developed. Compared with the two-dimensional (2D) model, in the 3D model, the cell-cell and cell-matrix connections were tighter, the activities of cytochrome P450 CYP1A2 and cytochrome P450 CYP2B6 were significantly increased, and the secretion levels of albumin and urea were increased. These indexes could be maintained stably for 21 d. Therefore, cell sheet stacking is helpful to improve the level of liver function of 3D liver model. This model is expected to be used to predict the metabolism of low-clearance drugs in preclinical, which is of great significance for drug evaluation and other studies.
Topics: Albumins; Animals; Cytochrome P-450 Enzyme System; Hepatocytes; Humans; Liver; Mice; Urea
PubMed: 36008342
DOI: 10.7507/1001-5515.202108056 -
Xenobiotica; the Fate of Foreign... Dec 2021Pirfenidone is a first-line drug for the treatment of idiopathic pulmonary fibrosis. The primary metabolic pathways of pirfenidone in humans are 5-hydroxylation and...
Pirfenidone is a first-line drug for the treatment of idiopathic pulmonary fibrosis. The primary metabolic pathways of pirfenidone in humans are 5-hydroxylation and subsequent oxidation to 5-carboxylpirfenidone. The aims of this study were to determine the cytochrome P450 isoforms responsible for pirfenidone 5-hydroxylation and to evaluate their contributions in human liver microsomes (HLM).Among the recombinant P450 isoforms, CYP1A2, CYP2D6, CYP2C19, CYP2A6, and CYP2B6 were shown to catalyse the 5-hydroxylation of pirfenidone. Pirfenidone 5-hydroxylase activity by HLM was inhibited by α-naphthoflavone (by 45%), 8-methoxypsolaren (by 84%), tranylcypromine (by 53%), and quinidine (by 15%), which are CYP1A2, CYP1A2/CYP2A6/CYP2C19, CYP2A6/CYP2C19, and CYP2D6 inhibitors, respectively.In 17 individual HLM donors, pirfenidone 5-hydroxylase activity was significantly correlated with phenacetin -deethylase ( = 0.89, < 0.001) and -mephenytoin 4'-hydroxylase activities ( = 0.51, < 0.05), which are CYP1A2 and CYP2C19 marker activities, respectively.By using the relative activity factors, the contributions of CYP1A2, CYP2C19, and CYP2D6 to pirfenidone 5-hydroxylation in the human liver were 72.8%, 11.8%, and 8.9%, respectively.In conclusion, we clearly demonstrated the predominant P450 involved in pirfenidone 5-hydroxylation in the human liver is CYP1A2, with CYP2C19 and CYP2D6 playing a minor role.
Topics: Catalysis; Cytochrome P-450 CYP1A2; Cytochrome P-450 CYP2C19; Cytochrome P-450 CYP2D6; Cytochromes; Humans; Hydroxylation; Liver; Microsomes, Liver; Pyridones
PubMed: 34779706
DOI: 10.1080/00498254.2021.2007553 -
PloS One 2023GL-V9, a new synthetic flavonoid derived from wogonin, has shown beneficial biological functions. In this study, accurate and sensitive UPLC-MS/MS methods were developed...
GL-V9, a new synthetic flavonoid derived from wogonin, has shown beneficial biological functions. In this study, accurate and sensitive UPLC-MS/MS methods were developed and validated for the quantification of GL-V9 and its glucuronide metabolite (5-O-glucuronide GL-V9) in Beagle dog plasma. The chromatographic separation was performed on a C8 column (ACE Excel 5 C8 50×3.0 mm) using 0.1% formic acid and acetonitrile were used as mobile phase. Mass detection was performed on a triple quadrupole tandem mass spectrometer equipped with an electrospray ionization (ESI) interface operating in positive ion mode. Quantitative analysis was performed in multiple reaction monitoring (MRM) mode with the transitions of m/z 410.2→126.1 for GL-V9, m/z 586.3→410.0 for 5-O-glucuronide GL-V9 and m/z 180.0→110.3 for phenacetin (internal standard), respectively. The calibration curves for GL-V9 and 5-O-glucuronide GL-V9 showed excellent linearity over the concentration range of 0.5-500 ng/mL with correlation coefficient greater than 0.99. The intra- and inter-day accuracies were within 99.86% to 109.20% for GL-V9 and 92.55% to 106.20% for 5-O-glucuronide GL-V9, respectively. The mean recovery was 88.64% ± 2.70% for GL-V9, and 92.31% ± 6.28% for 5-O-glucuronide GL-V9, respectively. The validated method was successfully applied to the pharmacokinetic study in Beagle dogs after oral and intravenous administration. The oral bioavailability of GL-V9 was approximately 2.47%~4.35% in Beagle dogs and reached steady state on the fifth day after repeated dosing.
Topics: Dogs; Animals; Tandem Mass Spectrometry; Chromatography, Liquid; Chromatography, High Pressure Liquid; Glucuronides; Flavonoids; Reproducibility of Results
PubMed: 37285365
DOI: 10.1371/journal.pone.0286467 -
Journal of Pharmacological and... 2020Ex vivo studies of human fetal hepatic drug metabolism are uncommon as it requires access to functional liver tissue and therefore raises practical and ethical concerns....
INTRODUCTION
Ex vivo studies of human fetal hepatic drug metabolism are uncommon as it requires access to functional liver tissue and therefore raises practical and ethical concerns. Large animal models provide an alternative opportunity to study changes in cytochrome P450 (CYP) activity in the mother and fetus during pregnancy. We aimed to develop methods to determine the activity of CYP1A2, CYP2C9, CYP2D6 and CYP3A4 in sheep hepatic microsomes.
METHODS
We identified optimal conditions to determine the activity of CYP1A2 (using the probe drug phenacetin), CYP2C9 (diclofenac), CYP2D6 (dextromethorphan) and CYP3A4 (midazolam) by varying techniques for microsome extraction, probe drug concentration, incubation time and microsome concentration. The specificity of each probe drug was assessed by determining the rate of metabolism when specific CYP enzyme inhibitors were included in the reaction.
RESULTS
The optimum incubation time and probe drug concentration was six hours with 5 μM phenacetin (CYP1A2), four hours with 10 μM diclofenac (CYP2C9), 30 min with 1 μM of midazolam (CYP3A4) and 10 min with 1 μM dextromethorphan (CYP2D6). For both CYP2D6 and CYP3A4 reactions required 20 μg of microsomal protein, whereas for CYP1A2 and CYP2C9, reactions required 40 μg of microsomal protein. Metabolism of phenacetin, dextromethorphan and midazolam was reduced by specific enzyme inhibitors, but the specific CYP2C9 inhibitor sulfaphenazole did not substantially inhibit diclofenac metabolism.
DISCUSSION
This study identifies the optimal conditions for determining CYP activity in maternal sheep hepatic microsomes. In doing so, we have developed a standardised protocol for assessment of microsomal activity of CYP3A4, CYP1A2 and CYP2D6, but we were unable to optimise conditions for assessment of CYP2C9. This approach can be applied to investigate the impact of pregnancy complications on maternal and fetal hepatic drug metabolism.
Topics: Animals; Cell Fractionation; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Dextromethorphan; Diclofenac; Dose-Response Relationship, Drug; Enzyme Assays; Feasibility Studies; Female; Maternal-Fetal Exchange; Microsomes, Liver; Midazolam; Phenacetin; Pregnancy; Pregnancy Complications; Sheep
PubMed: 33080390
DOI: 10.1016/j.vascn.2020.106934 -
Xenobiotica; the Fate of Foreign... Sep 2021Leonurine hydrochloride (LH) is derived from an ingredient of Houtt which is widely used for diseases in women.The influence of LH on the activity of cytochrome P450...
Leonurine hydrochloride (LH) is derived from an ingredient of Houtt which is widely used for diseases in women.The influence of LH on the activity of cytochrome P450 (CYPs) enzymes was investigated in this study.The effect of LH on CYPs enzyme activities were studied using the enzyme-selective substrates phenacetin (1A2), coumarin (2A6), diclofenac (2C9), S-mephenytoin (2C19), paclitaxel (2C8), dextromethorphan (2D6), chlorzoxazone (2E1) and testosterone (3A4). The IC value was calculated to express the strength of inhibition. The inhibition of CYPs was fitted with competitive or non-competitive inhibition models and corresponding parameters were also obtained.LH exerted inhibitory effects on the activity of CYP1A2, 2D6, and 3A4 with the IC values of 18.05, 15.13, and 20.09 μM, respectively. The obtained results showed that LH inhibited the activity of CYP1A2 and CYP2D6 via competitive manners ( = 8.667 μM and = 7.805 μM, respectively), while LH attenuated the CYP3A4 activity via a non-competitive manner ( = 9.507 μM). Moreover, LH showed time-dependent inhibition on CYP3A4 with the value of 4.31/0.044 min·μM.The inhibition of CYP1A2, CYP2D6, and CYP3A4 by LH, demonstrated , indicated the potential herb-drug interaction. Therefore, pharmacokinetic interactions involving LH and CYP1A2 or CYP2D6 or CYP1A2 substrates are likely to occur.
Topics: Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Gallic Acid; Humans; Microsomes, Liver
PubMed: 34176447
DOI: 10.1080/00498254.2021.1947544 -
International Immunopharmacology Jun 2020Interleukin (IL)-22 is a cytokine up-regulated in inflammatory situations and known to exert various hepatic effects. The potential impact of IL-22 towards liver drug...
Interleukin (IL)-22 is a cytokine up-regulated in inflammatory situations and known to exert various hepatic effects. The potential impact of IL-22 towards liver drug detoxifying proteins remains nevertheless unknown, but may be important to determine owing to the well-established alterations of liver detoxification occuring during inflammation. The present study was therefore designed to analyze the effects of IL-22 towards drug metabolizing enzyme and drug transporter expression and activity in cultured human hepatic cells. Exposure of differentiated hepatoma HepaRG cells or primary human hepatocytes to 10 ng/mL IL-22 was found to repress mRNA expression of cytochrome P-450 (CYP) 1A2, CYP3A4, CYP2B6 and CYP2C9 and of the sinusoidal sodium-taurocholate co-transporting polypeptide (NTCP); such IL-22 effects were concentration-dependent for CYP3A4 (IC = 1.7 ng/mL), CYP2B6 (IC = 0.9 ng/mL) and NTCP (IC = 1.8 ng/mL). Activity of CYP1A2 (phenacetin O-deethylation), CYP3A4 (midazolam hydroxylation) and CYP2B6 (bupropion hydroxylation), as well as that of NTCP (taurocholate uptake) were concomitantly decreased in IL-22-treated HepaRG cells; by contrast, activity of organic anion transporter polypeptides (OATPs) (estrone-3-sulfate uptake) and of organic cation transporter (OCT) 1 (tetra-ethylammonium uptake) remained unchanged. IL-22 was next found to activate the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) 3 pathway, whose inhibition by the JAK inhibitor ruxolitinib fully prevented the IL-22-mediated CYP3A4, CYP2B6 and NTCP repression in HepaRG cells. This JAK-dependent down-regulation of hepatic drug detoxifying proteins, notably of CYPs, by IL-22 may contribute to alteration of pharmacokinetics in patients suffering from acute and chronic inflammatory diseases and may be the source of drug-drug interactions.
Topics: Cytochrome P-450 CYP1A2; Cytochrome P-450 CYP2B6; Cytochrome P-450 CYP3A; Gene Expression Regulation; Hep G2 Cells; Hepatocytes; Humans; Inactivation, Metabolic; Interleukins; Janus Kinases; Nitriles; Organic Anion Transporters; Organic Anion Transporters, Sodium-Dependent; Pyrazoles; Pyrimidines; STAT3 Transcription Factor; Signal Transduction; Symporters; Interleukin-22
PubMed: 32234672
DOI: 10.1016/j.intimp.2020.106439 -
Environmental Science and Pollution... Feb 2023Phenacetin (PNT) is one of the most frequently detected nonsteroidal anti-inflammatory drugs in the water ecosystems, which poses a potential risk to environmental...
Phenacetin (PNT) is one of the most frequently detected nonsteroidal anti-inflammatory drugs in the water ecosystems, which poses a potential risk to environmental aquatic organisms. Acid-washed zero-valent aluminum (ZVAl) as a highly efficient activator for persulfate (PS) process was investigated to degrade PNT from the aqueous solution. The results indicated that acid-washed pretreatment for ZVAl could efficiently increase the degradation efficiency of PNT in the PS treatment. The degradation efficiency of PNT (50 μM) was up to 90% in 4 hours with the addition of 0.2 g/L acid-washed ZVAl and 8 mM PS at pH 6.8 and 25 °C. The PNT degradation followed pseudo-first order kinetics in the present system. High activator dosage, PS concentration, and reaction temperature could enhance the PNT degradation. The presence of inorganic anions (i.e., NO, HCO) and humic acid (HA) showed inhibitory effects on the PNT degradation. The reuse results illustrated the acid-washed ZVAl material would have continuous and efficient activation performance for PS to degrade the PNT. Radical scavenger experiments and electron paramagnetic resonance indicated that both SO and •OH were major reactive species during the PNT degradation. The possible degradation pathways of PNT mainly included the break of C-N and C-O bonds and further oxidation.
Topics: Aluminum; Phenacetin; Ecosystem; Water Pollutants, Chemical; Oxidation-Reduction; Water
PubMed: 36229732
DOI: 10.1007/s11356-022-23473-z