ABSTRACT
5F-EDMB-PICA is a newly emerged synthetic cannabinoid which has been characterized in relevant literature in recent years. Although phase-I metabolites of 5F-EDMB-PICA have been partly reported, the phase-II metabolism of this synthetic cannabinoid has not been studied yet. In this study, we established a phase-I and phase-II metabolism model in vitro by using pooled human liver microsomes, NADPH regeneration system, and UGT incubation system, with 1 mg/ml 5F-EDMB-PICA added and incubated at 37 °C for 60 min. The metabolites were analyzed by Q Exactive™ Hybrid Quadrupole-Orbitrap™ Mass Spectrometer, via which we discovered and identified 14 phase-I metabolites and 4 phase-II metabolites of 5F-EDMB-PICA, involving pathways such as ester hydrolysis, dehydrogenation, hydrolytic defluorination, hydroxylation, dihydroxylation, glucuronidation, and combinations of the pathways mentioned above. We recommend considering the monohydroxylation metabolites (M9, M10) with higher content and intact ester and 5-fluoropentyl structures as potential biomarkers of 5F-EDMB-PICA.
Subject(s)
Cannabinoids , Microsomes, Liver , Humans , Microsomes, Liver/metabolism , Cannabinoids/metabolism , Glucuronosyltransferase/metabolism , Metabolic Networks and Pathways , Metabolic Detoxication, Phase I , Metabolic Detoxication, Phase II , NADP/metabolism , HydroxylationABSTRACT
Adequate antioxidant supply is essential for maintaining metabolic homeostasis and reducing oxidative stress during detoxification. The emerging evidence suggests that certain classes of phytonutrients can help support the detoxification process by stimulating the liver to produce detoxification enzymes or acting as antioxidants that neutralize the harmful effects of free radicals. This study was designed to examine the effects of a guided 28-day metabolic detoxification program in healthy adults. The participants were randomly assigned to consume a whole food, multi-ingredient supplement (n = 14, education and intervention) or control (n = 18, education and healthy meal) daily for the duration of the trial. The whole food supplement contained 37 g/serving of a proprietary, multicomponent nutritional blend in the form of a rehydratable shake. Program readiness was ensured at baseline using a validated self-perceived wellness score and a blood metabolic panel, indicating stable emotional and physical well-being in both groups. No significant changes or adverse effects were found on physical or emotional health, cellular glutathione (GSH) and the GSH:GSSG ratio, porphyrin, and hepatic detoxification biomarkers in urine. The intervention was positively associated with a 23% increase in superoxide dismutase (p = 0.06) and a 13% increase in glutathione S-transferase (p = 0.003) activities in the blood. This resulted in a 40% increase in the total cellular antioxidant capacity (p = 0.001) and a 13% decrease in reactive oxygen species (p = 0.002) in isolated PBMCs from participants in the detoxification group. Our findings indicate that consuming a whole food nutritional intervention as a part of the guided detoxification program supported phase II detoxification, in part, by promoting enhanced free radical scavenging and maintaining redox homeostasis under the body's natural glutathione recycling capacity.
Subject(s)
Antioxidants , Metabolic Detoxication, Phase II , Adult , Humans , Healthy Volunteers , Glutathione , Dietary SupplementsABSTRACT
Aflatoxins are toxic secondary metabolites produced by Aspergillus fungi. The most toxic among them is Aflatoxin B1 (AFB1) which is known to have genotoxic, immunotoxic, teratogenic, carcinogenic, and mutagenic toxic effects (amongst others). The mechanisms responsible for its toxicity include the induction of oxidative stress, cytotoxicity, and DNAdamage. Studies have found that natural anti-oxidants can reduce the damage that AFB1 inflicts on the body by alleviating oxidative stress and inhibiting the biotransformation of AFB1. Therefore, this review outlines the latest progress in research on the use of natural anti-oxidants as antidotes to aflatoxin poisoning and their detoxification mechanisms. It also considers the problems that may possibly arise from their use and their application prospects. Our aim is to provide a useful reference for the prevention and treatment of AFB1 poisoning.
Subject(s)
Aflatoxin B1 , Antidotes , Antioxidants , Metabolic Detoxication, Phase II , Aflatoxin B1/metabolism , Aflatoxin B1/poisoning , Aflatoxin B1/toxicity , Antidotes/pharmacology , Antioxidants/pharmacology , Carcinogens/metabolism , Carcinogens/toxicity , Animals , ChickensABSTRACT
OBJECTIVES: Acetaminophen (APAP) (paracetamol) is a widely used non-prescription drug for pain relief and antipyretic effects. The clearance of APAP is mainly through phase-2 biotransformation catalysed by UDP-glucuronosyl transferases (UGT). Dasabuvir is an anti-hepatitis C drug reported to inhibit several UGT isoforms. The study evaluated the in-vitro inhibitory capacity of dasabuvir versus APAP glucuronidation. METHODS: Procedures included human liver microsomal incubations with APAP and isoform-selective probe substrates. KEY FINDINGS: Dasabuvir inhibited APAP metabolism by a reversible, mixed-type (competitive and non-competitive) partial inhibition, with an inhibition constant Ki = 3.4 µM. The index constant 'a' was 6.7, indicating the relative contribution of competitive and non-competitive inhibition. The enzyme-inhibitor complex was still able to catalyse the reaction by 12% of the control capacity. Dasabuvir produced strong partial inhibition effect of UGT1A1 and UGT1A9 and relatively complete inhibition of UGT1A6. CONCLUSIONS: Consistent with previous reports, dasabuvir inhibits the activity of 3 UGT isoforms associated with APAP metabolism. In-vitro to in-vivo scaling by 2 different approaches showed identical results, predicting an increased AUC of APAP by a factor of 1.3-fold with coadministration of dasabuvir. Until the findings are confirmed in clinical drug interaction studies, APAP dosage should not exceed 3 g per day in dasabuvir-treated patients to avoid potentially hepatotoxic APAP exposures.
Subject(s)
2-Naphthylamine/pharmacokinetics , Acetaminophen/pharmacokinetics , Glucuronosyltransferase/metabolism , Sulfonamides/pharmacokinetics , Uracil/analogs & derivatives , Antipyretics/pharmacokinetics , Antiviral Agents/pharmacokinetics , Area Under Curve , Drug Interactions , Glucuronosyltransferase/antagonists & inhibitors , Humans , Isoenzymes/antagonists & inhibitors , Metabolic Detoxication, Phase II , Microsomes, Liver , UDP-Glucuronosyltransferase 1A9/antagonists & inhibitors , Uracil/pharmacokineticsABSTRACT
The t-type trichothecene producers Fusarium sporotrichioides and Fusarium graminearum protect themselves against their own mycotoxins by acetylating the C-3 hydroxy group with Tri101p acetylase. To understand the mechanism by which they deal with exogenously added d-type trichothecenes, the Δtri5 mutants expressing all but the first trichothecene pathway enzymes were fed with trichodermol (TDmol), trichothecolone (TCC), 8-deoxytrichothecin, and trichothecin. LC-MS/MS and NMR analyses showed that these C-3 unoxygenated trichothecenes were conjugated with glucose at C-4 by α-glucosidic linkage. As t-type trichothecenes are readily incorporated into the biosynthetic pathway following the C-3 acetylation, the mycotoxins were fed to the ΔFgtri5ΔFgtri101 mutant to examine their fate. LC-MS/MS and NMR analyses demonstrated that the mutant conjugated glucose at C-4 of HT-2 toxin (HT-2) by α-glucosidic linkage, while the ΔFgtri5 mutant metabolized HT-2 to 3-acetyl HT-2 toxin and T-2 toxin. The 4-O-glucosylation of exogenously added t-type trichothecenes appears to be a general response of the ΔFgtri5ΔFgtri101 mutant, as nivalenol and its acetylated derivatives appeared to be conjugated with hexose to some extent. The toxicities of 4-O-glucosides of TDmol, TCC, and HT-2 were much weaker than their corresponding aglycons, suggesting that 4-O-glucosylation serves as a phase II xenobiotic metabolism for t-type trichothecene producers.
Subject(s)
Fusarium/metabolism , Glucose/metabolism , Metabolic Detoxication, Phase II , Trichothecenes/metabolism , Acetylation , Chromatography, Liquid , Tandem Mass SpectrometryABSTRACT
Nicotinamide N-methyltransferase (NNMT) has progressed from being considered merely a Phase II metabolic enzyme to one with a central role in cell function and energy metabolism. Over the last three decades, a significant body of evidence has accumulated which clearly demonstrates a central role for NNMT in cancer survival, metastasis, and drug resistance. In this review, we discuss the evidence supporting a role for NNMT in the progression of the cancer phenotype and how it achieves this by driving the activity of pro-oncogenic NAD+-consuming enzymes. We also describe how increased NNMT activity supports the Warburg effect and how it promotes oncogenic changes in gene expression. We discuss the regulation of NNMT activity in cancer cells by both post-translational modification of the enzyme and transcription factor binding to the NNMT gene, and describe for the first time three long non-coding RNAs which may play a role in the regulation of NNMT transcription. We complete the review by discussing the development of novel anti-cancer therapeutics which target NNMT and provide insight into how NNMT-based therapies may be best employed clinically.
Subject(s)
Energy Metabolism/genetics , Neoplasms/genetics , Nicotinamide N-Methyltransferase/genetics , Drug Resistance, Neoplasm/genetics , Gene Expression Regulation, Neoplastic/genetics , Humans , Metabolic Detoxication, Phase II/genetics , NAD/metabolism , Neoplasms/drug therapy , Neoplasms/pathology , Protein Processing, Post-Translational/geneticsABSTRACT
Advances in genomics have revealed many of the genetic underpinnings of human disease, but exposomics methods are currently inadequate to obtain a similar level of understanding of environmental contributions to human disease. Exposomics methods are limited by low abundance of xenobiotic metabolites and lack of authentic standards, which precludes identification using solely mass spectrometry-based criteria. Here, we develop and validate a method for enzymatic generation of xenobiotic metabolites for use with high-resolution mass spectrometry (HRMS) for chemical identification. Generated xenobiotic metabolites were used to confirm identities of respective metabolites in mice and human samples based upon accurate mass, retention time and co-occurrence with related xenobiotic metabolites. The results establish a generally applicable enzyme-based identification (EBI) for mass spectrometry identification of xenobiotic metabolites and could complement existing criteria for chemical identification.
Subject(s)
Cytochrome P-450 Enzyme System/metabolism , Mass Spectrometry/methods , Microsomes, Liver/enzymology , Xenobiotics/metabolism , Animals , Cytochrome P-450 Enzyme System/genetics , Gene Expression , Humans , Isoenzymes/genetics , Isoenzymes/metabolism , Isotope Labeling , Liver/enzymology , Metabolic Detoxication, Phase I/genetics , Metabolic Detoxication, Phase II/genetics , MiceABSTRACT
BACKGROUND: The interplay between phase II enzymes and efflux transporters leads to extensive metabolism and low systemic bioavailability of flavonoids. OBJECTIVE: In this study, the dynamic interplay between multiple UGTs and multiple efflux transporters that occur inside the cells was fully investigated. METHODS: A new HeLa-UGT1A9-MRP3 cell was established to overexpress two dominant efflux transporters MRP3 and BCRP, and two UGT isoforms UGT1A9 and UGT1A3. The metabolism and glucuronides excretion for a model flavonoid genistein were determined in HeLa-UGT1A9-MRP3 cells and HeLa-UGT1A9-Con cells that overexpressed one UGT (1A9) and one efflux transporter (BCRP). RESULTS: The excretion rate grew nearly 6-fold, cellular clearance of glucuronides increased about 3-fold, and fraction of genistein metabolized (fmet) increased (14%, p<0.01) in the new cells. Small interfering (siRNA)-mediated MRP3 functional knockdown resulted in marked decreases in the excretion rates (26%-78%), intracellular amounts (56%-93%), and cellular clearance (54%-96%) in both cells, but the magnitude of the differences in HeLa- UGT1A9-Con cells was relatively small. Reductions in fmet values were similarly moderate (11%-14%). In contrast, UGT1A9 knockdown with siRNA caused large decreases in the excretion rates (46%-88%), intracellular amounts (80%-97%), cellular clearance (80%-98%) as well as fmet value (33%-43%, p<0.01) in both UGT1A9 cells. Comparisons of the kinetic parameters and profiles of genistein glucuronidation as well as UGT mRNA expression suggest that HeLa-UGT1A9-MRP3 has increased expression of both MRP3 and UGT1A3. CONCLUSION: The newly engineered HeLa-UGT1A9-MRP3 cells is an appropriate model to study the kinetic interplay between multiple UGTs and efflux transporters, and a promising biosynthetic tool to obtain flavonoid glucuronides of high purity.
Subject(s)
ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Cell Engineering/methods , Genistein/pharmacology , HeLa Cells , Multidrug Resistance-Associated Proteins/metabolism , Neoplasm Proteins/metabolism , UDP-Glucuronosyltransferase 1A9/metabolism , Biosynthetic Pathways , Flavonoids/pharmacology , Humans , Metabolic Detoxication, Phase II , Metabolic Flux AnalysisABSTRACT
In this study, we present, for the first time in Spain, the levels of 19 mycotoxins in plasma samples from healthy and sick children (digestive, autism spectrum (ASD), and attention deficit hyperactivity (ADHD) disorders) (n = 79, aged 2-16). The samples were analyzed by liquid chromatography-mass spectrometry (triple quadrupole) (LC-MS/MS). To detect Phase II metabolites, the samples were reanalyzed after pre-treatment with ß-glucuronidase/arylsulfatase. The most prevalent mycotoxin was ochratoxin A (OTA) in all groups of children, before and after enzyme treatment. In healthy children, the incidence of OTA was 92.5% in both cases and higher than in sick children before (36.7% in digestive disorders, 50% in ASD, and 14.3% in ADHD) and also after the enzymatic treatment (76.6 % in digestive disorders, 50% in ASD, and 85.7% in ADHD). OTA levels increased in over 40% of healthy children after enzymatic treatment, and this increase in incidence and levels was also observed in all sick children. This suggests the presence of OTA conjugates in plasma. In addition, differences in OTA metabolism may be assumed. OTA levels are higher in healthy children, even after enzymatic treatment (mean OTA value for healthy children 3.29 ng/mL, 1.90 ng/mL for digestive disorders, 1.90 ng/mL for ASD, and 0.82 ng/mL for ADHD). Ochratoxin B appears only in the samples of healthy children with a low incidence (11.4%), always co-occurring with OTA. Sterigmatocystin (STER) was detected after enzymatic hydrolysis with a high incidence in all groups, especially in sick children (98.7% in healthy children and 100% in patients). This supports glucuronidation as a pathway for STER metabolism in children. Although other mycotoxins were studied (aflatoxins B1, B2, G1, G2, and M1; T-2 and HT-2 toxins; deoxynivalenol, deepoxy-deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol; zearalenone; nivalenol; fusarenon-X; neosolaniol; and diacetoxyscirpenol), they were not detected either before or after enzymatic treatment in any of the groups of children. In conclusion, OTA and STER should be highly considered in the risk assessment of mycotoxins. Studies concerning their sources of exposure, toxicokinetics, and the relationship between plasma levels and toxic effects are of utmost importance in children.
Subject(s)
Attention Deficit Disorder with Hyperactivity/blood , Autism Spectrum Disorder/blood , Digestive System Diseases/blood , Mycotoxins/blood , Adolescent , Age Factors , Attention Deficit Disorder with Hyperactivity/diagnosis , Autism Spectrum Disorder/diagnosis , Biological Monitoring , Biomarkers/blood , Case-Control Studies , Child , Child, Preschool , Chromatography, Liquid , Digestive System Diseases/diagnosis , Female , Humans , Male , Metabolic Detoxication, Phase II , Mycotoxins/adverse effects , Ochratoxins/blood , Risk Assessment , Spain , Spectrometry, Mass, Electrospray Ionization , Sterigmatocystin/blood , Tandem Mass SpectrometryABSTRACT
Endocrine disrupting compounds (EDCs) are ubiquitous environmental pollutants that alter endocrine system function, induce birth defects, and a myriad of other negative health outcomes. Although the mechanism of toxicity of many EDCs have been studied in detail, little work has focused on understanding the mechanisms through which pregnant dams and fetuses protect themselves from EDCs, or if those protective mechanisms are sexually dimorphic in fetuses. In this study, we examined proteomic alterations in the livers of mouse dams and their male and female fetuses induced by vinclozolin, a model antiandrogenic EDC. Dam livers upregulated nine phase I and phase II detoxification pathways and pathway analysis revealed that more pathways are significantly enriched in dam livers than in fetal livers. Phase I and II detoxification proteins are also involved in steroid and steroid hormone biosynthesis and vinclozolin likely alters steroid levels in both the dam and the fetus. The response of the fetal liver proteome to vinclozolin exposure is sexually dimorphic. Female fetal livers upregulated proteins in xenobiotic metabolism pathways, whereas male fetal livers upregulated proteins in oxidative phosphorylation pathways. These results suggest that female fetuses increase protective mechanisms, whereas male fetuses increase ATP production and several disease pathways that are indicative of oxidative damage. Females fetuses upregulate proteins and protective pathways that were similar to the dams whereas males did not. If this sexually dimorphic pattern is typical, then males might generally be more sensitive to EDCs.
Subject(s)
Androgen Antagonists/toxicity , Endocrine Disruptors/toxicity , Liver/drug effects , Oxazoles/toxicity , Proteome , Adenosine Triphosphate/metabolism , Androgen Antagonists/metabolism , Animals , Endocrine Disruptors/metabolism , Female , Liver/embryology , Liver/metabolism , Male , Maternal Exposure , Metabolic Detoxication, Phase I , Metabolic Detoxication, Phase II , Mice , Oxazoles/metabolism , Oxidative Phosphorylation , Pregnancy , Proteomics , Sex Characteristics , Sex FactorsABSTRACT
Nonalcoholic steatohepatitis (NASH), the progressive form of nonalcoholic fatty liver disease, is increasing in prevalence. NASH-related alterations in hepatic protein expression (e.g., transporters) and in overall physiology may affect drug exposure by altering drug disposition and elimination. The aim of this study was to build a physiologically-based pharmacokinetic (PBPK) model to predict drug exposure in NASH by incorporating NASH-related changes in hepatic transporters. Morphine and morphine-3-glucuronide (M3G) were used as model compounds. A PBPK model of morphine with permeability-limited hepatic disposition was extended to include M3G disposition and enterohepatic recycling (EHR). The model captured the area under the plasma concentration-time curve (AUC) of morphine and M3G after intravenous morphine administration within 0.82-fold and 1.94-fold of observed values from 3 independent clinical studies for healthy adult subjects (6, 10, and 14 individuals). When NASH-related changes in multidrug resistance-associated protein 2 (MRP2) and MRP3 were incorporated into the model, the predicted M3G mean AUC in NASH was 1.34-fold higher compared to healthy subjects, which is slightly lower than the observed value (1.63-fold). Exploratory simulations on other physiological changes occurring in NASH (e.g., moderate decreases in glomerular filtration rate and portal vein blood flow) revealed that the effect of transporter changes was most prominent. Additionally, NASH-related transporter changes resulted in decreased morphine EHR, which could be important for drugs with extensive EHR. This study is an important first step to predict drug disposition in complex diseases such as NASH using PBPK modeling.
Subject(s)
Analgesics, Opioid/pharmacokinetics , Liver/metabolism , Models, Biological , Morphine Derivatives/pharmacokinetics , Non-alcoholic Fatty Liver Disease/metabolism , Administration, Intravenous , Analgesics, Opioid/administration & dosage , Case-Control Studies , Computer Simulation , Humans , Metabolic Detoxication, Phase II , Morphine Derivatives/administration & dosage , Multidrug Resistance-Associated Protein 2 , Multidrug Resistance-Associated Proteins/metabolismABSTRACT
In animal health risk assessment, hazard characterisation of feed additives has been often using the default uncertainty factor (UF) of 100 to translate a no-observed-adverse-effect level in test species (rat, mouse, dog, rabbit) to a 'safe' level of chronic exposure in farm and companion animal species. Historically, both 10-fold factors have been further divided to include chemical-specific data in both dimensions when available. For cats (Felis Sylvestris catus), an extra default UF of 5 is applied due to the species' deficiency in particularly glucuronidation and glycine conjugation. This paper aims to assess the scientific basis and validity of the UF for inter-species differences in kinetics (4.0) and the extra UF applied for cats through a comparison of kinetic parameters between rats and cats for 30 substrates of phase I and phase II metabolism. When the parent compound undergoes glucuronidation the default factor of 4.0 is exceeded, with exceptions for zidovudine and S-carprofen. Compounds that were mainly renally excreted did not exceed the 4.0-fold default. Mixed results were obtained for chemicals which are metabolised by CYP3A in rats. When chemicals were administered intravenously the 4.0-fold default was not exceeded with the exception of clomipramine, lidocaine and alfentanil. The differences seen after oral administration might be due to differences in first-pass metabolism and bioavailability. Further work is needed to further characterise phase I, phase II enzymes and transporters in cats to support the development of databases and in silico models to support hazard characterisation of chemicals particularly for feed additives.
Subject(s)
Animal Feed/toxicity , Cytochrome P-450 Enzyme System/metabolism , Food Contamination , Glucuronosyltransferase/metabolism , Xenobiotics/pharmacokinetics , Animals , Cats , Metabolic Detoxication, Phase I , Metabolic Detoxication, Phase II , No-Observed-Adverse-Effect Level , Rats , Risk Assessment , Species Specificity , Substrate Specificity , Uncertainty , Xenobiotics/administration & dosage , Xenobiotics/toxicityABSTRACT
The aim of this study was to investigate the sensitivity and specificity of endogenous glycochenodeoxycholate and glycodeoxycholate 3-O-glucuronides (GCDCA-3G and GDCA-3G) as substrates for organic anion transporting polypeptide 1B1 (OATP1B1) in humans. We measured fasting levels of plasma GCDCA-3G and GDCA-3G using liquid chromatography-tandem mass spectrometry in 356 healthy volunteers. The mean plasma levels of both compounds were ~ 50% lower in women than in men (P = 2.25 × 10-18 and P = 4.73 × 10-9 ). In a microarray-based genome-wide association study, the SLCO1B1 rs4149056 (c.521T>C, p.Val174Ala) variation showed the strongest association with the plasma GCDCA-3G (P = 3.09 × 10-30 ) and GDCA-3G (P = 1.60 × 10-17 ) concentrations. The mean plasma concentration of GCDCA-3G was 9.2-fold (P = 8.77 × 10-31 ) and that of GDCA-3G was 6.4-fold (P = 2.45x10-13 ) higher in individuals with the SLCO1B1 c.521C/C genotype than in those with the c.521T/T genotype. No other variants showed independent genome-wide significant associations with GCDCA-3G or GDCA-3G. GCDCA-3G was highly efficacious in detecting the SLCO1B1 c.521C/C genotype with an area under the receiver operating characteristic curve of 0.996 (P < 0.0001). The sensitivity (98-99%) and specificity (100%) peaked at a cutoff value of 180 ng/mL for men and 90 ng/mL for women. In a haplotype-based analysis, SLCO1B1*5 and *15 were associated with reduced, and SLCO1B1*1B, *14, and *35 with increased OATP1B1 function. In vitro, both GCDCA-3G and GDCA-3G showed at least 6 times higher uptake by OATP1B1 than OATP1B3 or OATP2B1. These data indicate that the hepatic uptake of GCDCA-3G and GDCA-3G is predominantly mediated by OATP1B1. GCDCA-3G, in particular, is a highly sensitive and specific OATP1B1 biomarker in humans.
Subject(s)
Glucuronides/metabolism , Glycochenodeoxycholic Acid/metabolism , Liver-Specific Organic Anion Transporter 1/metabolism , Liver/metabolism , Adult , Biomarkers/metabolism , Chromatography, Liquid , Female , Genome-Wide Association Study , Genotype , Glucuronides/blood , Glycochenodeoxycholic Acid/blood , HEK293 Cells , Healthy Volunteers , Humans , Liver-Specific Organic Anion Transporter 1/deficiency , Liver-Specific Organic Anion Transporter 1/genetics , Male , Metabolic Detoxication, Phase II , Oligonucleotide Array Sequence Analysis , Pharmacogenomic Variants , Phenotype , Polymorphism, Single Nucleotide , Tandem Mass Spectrometry , Young AdultABSTRACT
Understanding internal dose metrics is integral to adequately assess effects environmental contaminants might have on aquatic wildlife, including fish. In silico toxicokinetic (TK) models are a leading approach for quantifying internal exposure metrics for fishes; however, they often do not adequately consider chemicals that are actively biotransformed and have not been validated against early-life stages (ELS) that are often considered the most sensitive to the exposure to contaminants. To address these uncertainties, TK models were parameterized for the rapidly biotransformed chemical benzo[a]pyrene (B[a]P) in embryo-larval and adult life stages of fathead minnows. Biotransformation of B[a]P was determined through measurements of in vitro clearance. Using in vitro-in vivo extrapolation, in vitro clearance was integrated into a multi-compartment TK model for adult fish and a one-compartment model for ELS. Model predictions were validated using measurements of B[a]P metabolites from in vivo flow-through exposures to graded concentrations of water-borne B[a]P. Significantly greater amounts of B[a]P metabolites were observed with exposure to greater concentrations of parent compound in both life stages. However, when assessing biotransformation capacity, no differences in phase I or phase II biotransformation were observed with greater exposures to B[a]P. Results of modelling suggested that biotransformation of B[a]P can be successfully implemented into in silico models to accurately predict life stage-specific abundances of B[a]P metabolites in either whole-body larvae or the bile of adult fish. Models developed increase the scope of applications in which TK models can be used to support environmental risk assessments.
Subject(s)
Benzo(a)pyrene/toxicity , Cyprinidae/metabolism , Larva/drug effects , Models, Biological , Water Pollutants, Chemical/toxicity , Animals , Benzo(a)pyrene/metabolism , Biological Transport , Computer Simulation , Cyprinidae/growth & development , Larva/metabolism , Metabolic Detoxication, Phase I , Metabolic Detoxication, Phase II , Toxicokinetics , Water Pollutants, Chemical/metabolismABSTRACT
Zebrafish (Danio rerio) larvae have gained attention as a valid model to study in vivo drug metabolism and to predict human metabolism. The microinjection of compounds, oligonucleotides, or pathogens into zebrafish embryos at an early developmental stage is a well-established technique. Here, we investigated the metabolism of zebrafish larvae after microinjection of methyl 2-(1-(5-fluoropentyl)-1H-pyrrolo[2,3-b]pyridine-3-carboxamido)-3,3-dimethylbutanoate (7'N-5F-ADB) as a representative of recently introduced synthetic cannabinoids. Results were compared to human urine data and data from the in vitro HepaRG model and the metabolic pathway of 7'N-5F-ADB were reconstructed. Out of 27 metabolites detected in human urine samples, 19 and 15 metabolites were present in zebrafish larvae and HepaRG cells, respectively. The route of administration to zebrafish larvae had a major impact and we found a high number of metabolites when 7'N-5F-ADB was microinjected into the caudal vein, heart ventricle, or hindbrain. We further studied the spatial distribution of the parent compound and its metabolites by mass spectrometry imaging (MSI) of treated zebrafish larvae to demonstrate the discrepancy in metabolite profiles among larvae exposed through different administration routes. In conclusion, zebrafish larvae represent a superb model for studying drug metabolism, and when combined with MSI, the optimal administration route can be determined based on in vivo drug distribution.
Subject(s)
Cannabinoids/administration & dosage , Cannabinoids/metabolism , Models, Biological , Zebrafish/metabolism , Animals , Cannabinoids/chemistry , Cell Line , Drug Administration Routes , Humans , Larva , Metabolic Detoxication, Phase I , Metabolic Detoxication, Phase II , Metabolic Networks and Pathways , Metabolome , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Zebrafish/embryologyABSTRACT
Troglitazone and its major metabolite troglitazone sulfate were intravenously administered to chimeric mice with different ratios of liver replacement by human hepatocytes. Total clearances were converted to hepatic intrinsic clearances normalized to their liver weight, with the assumption that extra-hepatic elimination of these compounds was negligible. These values were plotted against the replacement indices, and postulated values for virtual 100% chimeric mice were assumed to be equivalent to those in humans. Metabolic formation ratio was estimated by comparing AUCs of troglitazone sulfate after separate administration of troglitazone and troglitazone sulfate. Liver to plasma concentration ratios were obtained from direct measurement. These parameters were extrapolated to 100% chimeric mice and subjected to semi-physiological pharmacokinetic modeling using pharmacokinetic parameters for oral administration taken from literature. Our simulated plasma concentration-time profile of troglitazone agreed well with observed values obtained in clinical study. However, the profile of troglitazone sulfate was far below the reported values. Although the possible reasons for this discrepancy remains unsolved, the combination of chimeric mice with semi-physiological PK modeling proved to be a useful tool in understanding the function of each PK parameter in human pharmacokinetics of troglitazone and its conjugated metabolite.
Subject(s)
Hepatocytes/enzymology , Hypoglycemic Agents/pharmacokinetics , Liver/enzymology , Models, Biological , Sulfuric Acid Esters/pharmacokinetics , Troglitazone/pharmacokinetics , Animals , Computer Simulation , Hepatocytes/transplantation , Humans , Hypoglycemic Agents/blood , Male , Metabolic Detoxication, Phase II , Mice, Transgenic , Sulfuric Acid Esters/blood , Transplantation Chimera , Troglitazone/bloodABSTRACT
OBJECTIVE: To explore the effects of aerobic exercise combined with huwentoxin-I (HWTX-I)-mediated Keap1-Nrf2-ARE pathway on phase II detoxification enzymes HO-1 and NQO1 and their protective effects against obstructive jaundice (OJ)-induced central nervous system injury in mice. METHODS: 50 male KM mice were randomly divided into blank group (GO), model group (M), aerobic exercise group (T), HWTX-I group (H), and aerobic exercise combined with HWTX-I group (TH). Mouse models of OJ were established with surgical suture for 72 h in the mice in all the groups except for the blank control group. The mice received interventions by aerobic exercise and tail vein injection of HWTX-I (0.05 µg/g) and were assessed by behavioral observation, Clark's neurological function scores, enzyme-linked immunosorbent assay (ELISA), brain tissue Nissl staining, hippocampal tissue Western blotting, and liver tissue mRNA expression profiling and sequencing. RESULTS: The mice in group M had obvious jaundice symptoms after the operation with significantly increased Clark's neurological score (P < 0.01). Compared with those in group M, the mice in group T, group H, and group TH showed significantly decreased serum levels of ALT, AST, TBIL, and TBA (P < 0.01) with increased contents of 5-HT and BDNF and decreased contents of S100B and NSE in the hippocampus (P < 0.01). Synergistic effects between aerobic exercise and HWTX-I were noted on the above parameters except for the liver function indicators. Interventions with aerobic exercise and HWTX-I, alone or in combination, obviously lessened pathologies in the brain tissue induced by OJ, and the combined treatment produced the strongest effect. The treatment also increased the expression levels of Nrf2, HO-1, and NQO1 mRNA and protein in brain tissues (P < 0.01 or 0.05) with a synergistic effect between aerobic exercise and HWTX-I. Illumina high-throughput sequencing showed that the differentially expressed factors participated mainly in such neural regulatory pathways as neuroactive ligand-receptor interaction, GABAergic synapses, dopaminergic synapses, synaptic vesicle circulation, and axon guidance, involving tissue cell neuronal signal transduction, apoptosis inhibition, immune response, and toxicity. Aerobic exercise and HWTX-I synergistically increased the accumulation of the signal pathways related with neuron damage repair and proliferation. CONCLUSIONS: Aerobic exercise combined with HWTX-I can up-regulate the expression of phase â ¡ detoxification enzymes HO-1 and NQO1 through the Keap1-Nrf2-ARE pathway to protect the central nervous system against OJ-induced damage in mice.
Subject(s)
Jaundice, Obstructive , Trauma, Nervous System , Animals , Kelch-Like ECH-Associated Protein 1 , Male , Metabolic Detoxication, Phase II , Mice , NF-E2-Related Factor 2 , Physical Conditioning, Animal , Reptilian Proteins , Spider VenomsABSTRACT
A physiologically based pharmacokinetic (PBPK) model for di-isononyl phthalate (DiNP) was developed by adapting the existing models for di(2-ethylhexyl) phthalate (DEHP) and di-butylphthalate (DBP). Both pregnant rat and human time-course plasma and urine data were used to address the hydrolysis of DiNP in intestinal tract, plasma, and liver as well as hepatic oxidative metabolism and conjugation of the monoester and primary oxidative metabolites. Data in both rats and humans were available to inform the uptake and disposition of mono-isononyl phthalate (MiNP) as well as the three primary oxidative metabolites including hydroxy (7-OH)-, oxo (7-OXO)-, and carboxy (7-COX)-monoisononyl phthalate in plasma and urine. The DiNP model was reliable over a wide range of exposure levels in the pregnant rat as well as the two low exposure levels in humans including capturing the nonlinear behavior in the pregnant rat after repeated 750 mg/kg/day dosing. The presented DiNP PBPK model in pregnant rat and human, based upon an extensive kinetic dataset in both species, may provide a basis for assessing human equivalent exposures based upon either rodent or in vitro points of departure.
Subject(s)
Environmental Pollutants/pharmacokinetics , Phthalic Acids/pharmacokinetics , Plasticizers/pharmacokinetics , Animals , Female , Humans , Intestines , Liver/metabolism , Metabolic Detoxication, Phase II , Models, Animal , Oxidation-Reduction , Plasma/metabolism , Pregnancy , RatsABSTRACT
OBJECTIVES: Isobavachin is a phenolic with anti-osteoporosis activity. This study aimed to explore its metabolic fates in vivo and in vitro, and to investigate the potential drug-drug interactions involving CYPs and UGTs. METHODS: Metabolites of isobavachin in mice were first identified and characterized. Oxidation and glucuronidation study were performed using liver and intestine microsomes. Reaction phenotyping, activity correlation analysis and relative activity factor approaches were employed to identify the main CYPs and UGTs involved in isobavachin metabolism. Through kinetic modelling, inhibition mechanisms towards CYPs and UGTs were also explored. KEY FINDINGS: Two glucuronides (G1 - G2) and three oxidated metabolites (M1 - M3) were identified in mice. Additionally, isobavachin underwent efficient oxidation and glucuronidation by human liver microsomes and HIM with CLint values from 5.53 to 148.79 µl/min per mg. CYP1A2, 2C19 contributed 11.3% and 17.1% to hepatic metabolism of isobavachin, respectively, with CLint values from 8.75 to 77.33 µl/min per mg. UGT1As displayed CLint values from 10.73 to 202.62 µl/min per mg for glucuronidation. Besides, significant correlation analysis also proved that CYP1A2, 2C19 and UGT1A1, 1A9 were main contributors for the metabolism of isobavachin. Furthermore, mice may be the appropriate animal model for predicting its metabolism in human. Moreover, isobavachin exhibited broad inhibition against CYP2B6, 2C9, 2C19, UGT1A1, 1A9, 2B7 with Ki values from 0.05 to 3.05 µm. CONCLUSIONS: CYP1A2, 2C19 and UGT1As play an important role in isobavachin metabolism. Isobavachin demonstrated broad-spectrum inhibition of CYPs and UGTs.
Subject(s)
Enzyme Inhibitors/pharmacology , Flavonoids/pharmacology , Glucuronosyltransferase/antagonists & inhibitors , Plant Extracts/pharmacology , Psoralea , Animals , Cytochrome P-450 Enzyme Inhibitors/isolation & purification , Cytochrome P-450 Enzyme Inhibitors/metabolism , Cytochrome P-450 Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/isolation & purification , Enzyme Inhibitors/metabolism , Flavonoids/isolation & purification , Flavonoids/metabolism , Glucuronides/metabolism , Glucuronosyltransferase/metabolism , Humans , Isoenzymes , Kinetics , Male , Metabolic Detoxication, Phase I , Metabolic Detoxication, Phase II , Mice , Microsomes, Liver/drug effects , Microsomes, Liver/enzymology , Oxidation-Reduction , Plant Extracts/isolation & purification , Plant Extracts/metabolism , Psoralea/chemistry , Species SpecificityABSTRACT
This study aimed to investigate the detoxification metabolism responses in scallop Chlamys farreri exposed to phenanthrene (PHE), chrysene (CHR), benzo[a]pyrene (B[a]P) and PHE + CHR + B[a]P for 15 days under laboratory conditions. The mRNA expression levels of AhR signaling pathway (AhR, HSP90, XAP2 and ARNT), detoxification system (phase I: CYP1A1 and CYP1B1; phase II: SULTs, UGT and GSTs) and ATP-binding cassette transporters (phase 0: ABCB1 and phase III: ABCC1, ABCG2) in digestive glands of scallops exposed to PHE (0.7, 2.1 µg/L), CHR (0.7, 2.1 µg/L), B[a]P (0.7, 2.1 µg/L), and PHE + CHR + B[a]P (0.7 + 0.7 +0.7, 2.1 + 2.1 + 2.1 µg/L) were detected. In present study, key genes (AhR, HSP90, XAP2 and ARNT) of the AhR signaling pathway can be significantly induced by pollutants, suggesting that the AhR/ARNT signaling pathway plays a role directly or indirectly. AhR, HSP90 and ARNT reached the maximum value on day 6, which can be preliminarily understood as the synchronization of their functions. Besides, the results also indicated that different genes had specific response to different pollution exposure. CYP1B1, GST-2, GST-omega and GST-microsomal could be potional indexes to PHE, ARNT, GST-sigma 2 and GST-3 were sensitive to CHR exposure, HSP90, GST-theta and ABCG2 were considered as potional indexes to BaP while CYP1A1 and UGT were possible to be indexes for monitoring the mix exposure of these three PAHs. These findings in C. farreri suggested that phase II detoxification metabolic enzymes isoforms played an essential role in detoxification mechanisms and mRNA expression levels of specific SULTs, UGTs and GSTs were potentially to be ideal indexes in PAHs pollution research. In summary, this study provides more valuable information for the risk assessments of different rings of PAHs.