Evaluation of dermorphin metabolism using zebrafish water tank model and human liver microsomes

2021 ◽  
Vol 22 ◽  
Author(s):  
Juliana de L. Castro ◽  
Henrique M. G. Pereira ◽  
Valéria P. de Sousa ◽  
Maria Elvira P. Martucci
Keyword(s):  
Human Liver ◽  
High Resolution Mass Spectrometry ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Model Systems ◽  
Water Tank ◽  
Doping Agent ◽  
Tank Model ◽  
Metabolic Pattern ◽  
Kinetics Of

Background: Dermorphin is a heptapeptide with an analgesic potential higher than morphine that does not present the same risk for the development of tolerance. These pharmacological features make dermorphin a potential doping agent in competitive sports and is already prohibited for racehorses. For athletes, the development of an efficient strategy to monitor for its abuse necessitates an investigation of the metabolism of dermorphin in humans. Methods: Here, human liver microsomes and zebrafish were utilized as model systems of human metabolism to evaluate the presence and kinetics of metabolites derived from dermorphin. Five hours after its administration, the presence of dermorphin metabolites could be detected in both models by liquid chromatography coupled to high resolution mass spectrometry. Results: Although the two models showed common results, marked differences were also observed in relation to the formed metabolites. Six putative metabolites, based on their exact masses of m/z 479.1915, m/z 501.1733, m/z 495.1657, m/z 223.1073, m/z 180.1017 and m/z 457.2085, are proposed to represent the metabolic pattern of dermorphin. The major metabolite generated from the administration of dermorphin in both models was YAFG-OH (m/z 457.2085), which is the N-terminal tetrapeptide previously identified from studies with rats. Conclusion: Its extensive characterization and commercial availability suggests that it could serve as a primary target analyte for the detection of dermorphin misuse. The metabolomics approach also allowed the assignment of other confirmatory metabolites.

scholarly journals Nontargeted Metabolomics by High-Resolution Mass Spectrometry to Study the In Vitro Metabolism of a Dual Inverse Agonist of Estrogen-Related Receptors β and γ, DN203368

Pharmaceutics ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 776
Author(s):  
Sin-Eun Kim ◽  
Seung-Bae Ji ◽  
Euihyeon Kim ◽  
Minseon Jeong ◽  
Jina Kim ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
High Resolution ◽  
Human Liver ◽  
High Resolution Mass Spectrometry ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Rat Liver Microsomes ◽  
High Resolution Mass ◽  
Resolution Mass

DN203368 ((E)-3-[1-(4-[4-isopropylpiperazine-1-yl]phenyl) 3-methyl-2-phenylbut-1-en-1-yl] phenol) is a 4-hydroxy tamoxifen analog that is a dual inverse agonist of estrogen-related receptor β/γ (ERRβ/γ). ERRγ is an orphan nuclear receptor that plays an important role in development and homeostasis and holds potential as a novel therapeutic target in metabolic diseases such as diabetes mellitus, obesity, and cancer. ERRβ is also one of the orphan nuclear receptors critical for many biological processes, such as development. We investigated the in vitro metabolism of DN203368 by conventional and metabolomic approaches using high-resolution mass spectrometry. The compound (100 μM) was incubated with rat and human liver microsomes in the presence of NADPH. In the metabolomic approach, the m/z value and retention time information obtained from the sample and heat-inactivated control group were statistically evaluated using principal component analysis and orthogonal partial least-squares discriminant analysis. Significant features responsible for group separation were then identified using tandem mass spectra. Seven metabolites of DN203368 were identified in rat liver microsomes and the metabolic pathways include hydroxylation (M1-3), N-oxidation (M4), N-deisopropylation (M5), N,N-dealkylation (M6), and oxidation and dehydrogenation (M7). Only five metabolites (M2, M3, and M5-M7) were detected in human liver microsomes. In the conventional approach using extracted ion monitoring for values of mass increase or decrease by known metabolic reactions, only five metabolites (M1-M5) were found in rat liver microsomes, whereas three metabolites (M2, M3, and M5) were found in human liver microsomes. This study revealed that nontargeted metabolomics combined with high-resolution mass spectrometry and multivariate analysis could be a more efficient tool for drug metabolite identification than the conventional approach. These results might also be useful for understanding the pharmacokinetics and metabolism of DN203368 in animals and humans.

Effect of Commonly Used Organic Solvents on the Kinetics of Cytochrome P450 2B6- and 2C8-Dependent Activity in Human Liver Microsomes

2007 ◽  
Vol 35 (11) ◽  
pp. 1990-1995 ◽  
Author(s):  
Ragini Vuppugalla ◽  
Shu-Ying Chang ◽  
Hongjian Zhang ◽  
Punit H. Marathe ◽  
David A. Rodrigues
Keyword(s):  
Cytochrome P450 ◽  
Organic Solvents ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Cytochrome P450 2B6 ◽  
Dependent Activity ◽  
Kinetics Of

scholarly journals Kinetics of tris (1-chloro-2-propyl) phosphate (TCIPP) metabolism in human liver microsomes and serum

Chemosphere ◽  
2016 ◽  
Vol 144 ◽  
pp. 1299-1305 ◽  
Author(s):  
Nele Van den Eede ◽  
Gregg Tomy ◽  
Fang Tao ◽  
Thor Halldorson ◽  
Stuart Harrad ◽  
...  
Keyword(s):  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Kinetics Of

scholarly journals Kinetics of Trihalogenated Acetic Acid Metabolism and Isoform Specificity in Liver Microsomes

2011 ◽  
Vol 30 (5) ◽  
pp. 551-561 ◽  
Author(s):  
Shakil A. Saghir ◽  
Burhan I. Ghanayem ◽  
Irvin R. Schultz
Keyword(s):  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Water Disinfection ◽  
Metabolic Clearance ◽  
Interspecies Differences ◽  
P450 Reductase ◽  
Cyp 2E1 ◽  
Chemical Inhibitors ◽  
Kinetics Of

This study determined the metabolism of 3 drinking water disinfection by-products (halogenated acetic acids [HAAs]), bromodichloroacetic acid (BDCAA), chlorodibromoacetic acid (CDBAA), and tribromoacetic acid (TBAA), using rat, mouse, human liver microsomes, and recombinant P450. Metabolism proceeded by reductive debromination forming a di-HAA; the highest under nitrogen >>2% oxygen > atmospheric headspaces. Vmax for the loss of tri-HAA was 4 to 5 times higher under nitrogen than atmospheric headspace. Intrinsic metabolic clearance was TBAA>CDBAA>>BDCAA. At the high substrate concentrations, tri-HAA consumption rate was 2 to 3 times higher than the formation of di-HAA. Liberation of Br− from TBAA corresponded to the expected amount produced after DBAA formation, indicating retention of Br− by additional metabolite/metabolites. Subsequent experiments with CDBAA detected negligible formation of chlorodibromomethane (CDBM) and failed to account for the missing tri-HAA. Carbon monoxide and especially diphenyleneiodonium ([DPI] P450 reductase inhibitor) blocked CDBAA metabolism. Other chemical inhibitors were only partially able to block CDBAA metabolism. Most effective were inhibitors of CYP 2E1 and CYP 3A4. Immunoinhibition studies using human liver microsomes and anti-human CYP 2E1 antibodies were successful in reducing CDBAA metabolism. However, CDBAA metabolism in wild-type (WT) and CYP 2E1 knockout (KO) mouse liver microsomes was similar, suggesting significant interspecies differences in CYP isoform in tri-HAA metabolism. Additional assessment of CYP isoform involvement was complicated by the finding that recombinantly expressed rat and human P450 reductase was able to metabolize CDBAA, which may be a contributing factor in interspecies differences in tri-HAA metabolism.

Metabolic Characteristics of SM-1, a Novel PAC-1 Derivative, in Human Liver Microsomes

2021 ◽  
Vol 17 ◽  
Author(s):  
Ya Gong ◽  
Peiqi Wang ◽  
Jianming Li ◽  
Jinsong Ding
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Enzyme Kinetics ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Spectrometry Method ◽  
Mass Spectrometry Method ◽  
Metabolic Characteristics ◽  
Kinetics Of

Background and Objectives: SM-1 is a new synthetic small molecule compound with antitumor activity. The metabolism of SM-1 is a key parameter that needs to be evaluated to provide further insight into drug safety and efficacy in the early phases of drug development. Methods and Results: In this study, the biotransformation process of SM-1 including the metabolic pathways and major metabolites was investigated based on a liquid chromatography-mass spectrometry method. Upon incubation of SM-1 with human liver microsomes, five metabolites were identified, namely dihydrodiol formation (R1), hydroxylation (R2, R3 and R5), and debenzylation (R4) of SM-1, with R1 and R4 being the major metabolites. The enzyme kinetic parameters of SM-1 were determined by a liquid chromatography tandem mass spectrometry method. The enzyme kinetics of SM-1 obeyed the Michaelis-Menten equation. The Vmax, Km, and CLint of SM-1 in HLMs were 14.5 nmol/mg protein/h, 6.32 μM, and 2.29 mL/mg protein/h, respectively. The chemical inhibition studies showed that CYP450 isoenzymes were responsible for SM-1 metabolism in HLMs and CYP3A4 was the major CYP450 isoenzyme involved in the metabolism of SM-1; these findings were confirmed by using the human recombinant CYP3A4. Conclusions : Through identification of the biotransformation pathways and enzyme kinetics of SM-1, the metabolic enzymes for SM-1 in HLMs are characterized.

scholarly journals In vitro biotransformation of pyrrolizidine alkaloids in different species: part II—identification and quantitative assessment of the metabolite profile of six structurally different pyrrolizidine alkaloids

2020 ◽  
Vol 94 (11) ◽  
pp. 3759-3774
Author(s):  
Ina Geburek ◽  
Dieter Schrenk ◽  
Anja These
Keyword(s):  
Human Liver ◽  
Pyrrolizidine Alkaloids ◽  
High Resolution Mass Spectrometry ◽  
Liver Microsomes ◽  
Metabolite Profile ◽  
Human Liver Microsomes ◽  
Rat Liver Microsomes ◽  
Metabolite Profiles ◽  
Fragmentation Patterns

Abstract Pyrrolizidine alkaloids (PA) exert their toxic effects only after bioactivation. Although their toxicity has already been studied and metabolic pathways including important metabolites were described, the quantification of the latter revealed a large unknown portion of the metabolized PA. In this study, the qualitative and quantitative metabolite profiles of structurally different PAs in rat and human liver microsomes were investigated. Between five metabolites for europine and up to 48 metabolites for lasiocarpine were detected. Proposals for the chemical structure of each metabolite were derived based on fragmentation patterns using high-resolution mass spectrometry. The metabolite profiles of the diester PAs showed a relatively good agreement between both species. The metabolic reactions were summarized into three groups: dehydrogenation, oxygenation, and shortening of necic acid(s). While dehydrogenation of the necine base is considered as bioactivation, both other routes are considered as detoxification steps. The most abundant changes found for open chained diesters were dealkylations, while the major metabolic pathway for cyclic diesters was oxygenation especially at the nitrogen atom. In addition, all diester PAs formed several dehydrogenation products, via the insertion of a second double bond in the necine base, including the formation of glutathione conjugates. In rat liver microsomes, all investigated PAs formed dehydropyrrolizidine metabolites with the highest amount formed by lasiocarpine, whereas in human liver microsomes, these metabolites could only be detected for diesters. Our findings demonstrate that an extensive analysis of PA metabolism can provide the basis for a better understanding of PA toxicity and support future risk assessment.

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