Enzymic Cleavage Of In Vivo Formed Maillard-Type Compounds Involved In Haemostasis

1981 ◽  
Author(s):  
L Mester ◽  
L Szabados ◽  
M Mester
Keyword(s):  
Enzyme System ◽  
Basal Membrane ◽  
Enzyme Concentration ◽  
Cytochrome P450 Enzyme ◽  
Microsomal Preparation ◽  
Chemical Hydrolysis ◽  
P450 Enzyme ◽  
Liver Microsomal Preparation

Desoxyfructose derivatives of serotonin (Mester et al.,1975), of haemoglobin (Flückiger and Winterhalter, 1976), of poly-L-lysine (Mester et al., 1975) and of lysine rich histones (Kertesz-Crisba, 1977) are easily formed in vivo by a simple Maillard-type chemical reaction. Some of these compounds interfere with platelet functions (Mester et al.,1976) or contribute to the thickening of the basal membrane of blood vessels (Cerami et al., 1979).While the chemical synthesis of Maillard-type compounds proceeds readily even in vivo, the chemical cleavage of them needs sever conditions which certainly do not exist in vivo (Gottschalk, 1952). However, a slow liberation of serotonin from desoxyfructo-serotonin is observed in vivo, suggesting the existence of an enzyme system for the cleavage of Maillard-type sugar-amine derivatives. In vitro, using a sheep liver microsomal preparation rich in Cytochrome P450 enzyme, the liberation of serotonin is in linear correlation with the enzyme concentration. The cleavage of desoxyfructo-serotonin is activated by NADPH having its optimum at pH=7.4, excluding definitely the occurence of a chemical hydrolysis.Factors interfering with the enzyme system involved in the cleavage of Maillard-type compounds, may also interfere with haemostasis.

scholarly journals Chlorogenic Acid as a Model Compound for Optimization of an In Vitro Gut Microbiome-Metabolism Model

Proceedings ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 31 ◽  
Author(s):  
Olivier Mortelé ◽  
Elias Iturrospe ◽  
Annelies Breynaert ◽  
Christine Lammens ◽  
Xavier Basil Britto ◽  
...  
Keyword(s):  
Chlorogenic Acid ◽  
Gut Microbiome ◽  
Model Compound ◽  
In Vitro Models ◽  
Cytochrome P450 Enzyme ◽  
Flight Mass Spectrometry ◽  
P450 Enzyme ◽  
Analytical Phase

It has been believed that the metabolism of xenobiotics occurred mainly by the cytochrome P450 enzyme system in the liver. However, recent data clearly suggest a significant role for the gut microbiota in the metabolism of xenobiotic compounds. This microbiotic biotransformation could lead to differences on activation, inactivation and possible toxicity of these compounds. In vitro models are generally used to study the colonic biotransformation as they allow easy dynamic and multiple sampling over time. However, to ensure this accurately mimics communities in vivo, the pre-analytical phase requires optimization. Chlorogenic acid, a polyphenolic compound abundantly present in the human diet, was used as a model compound to optimize a ready-to-use gut microbiome biotransformation platform. Samples of the in vitro gastrointestinal dialysis-model with colon stage were analyzed by liquid chromatography coupled to high resolution time-of-flight mass spectrometry. Complementary screening approaches were also employed to identify the biotransformation products.

Effects of 31 recombinant CYP2C19 variants on clomipramine metabolism in vitro

2021 ◽  
pp. 026988112110505
Author(s):  
Tian Lan ◽  
Ya-Qing Ma ◽  
Ya-Min Dang ◽  
Chen-Chen Wang ◽  
Ren-Ai Xu ◽  
...  
Keyword(s):  
High Performance ◽  
Intrinsic Clearance ◽  
Cytochrome P450 Enzyme ◽  
Liquid Chromatography Mass Spectrometry ◽  
Wild Type ◽  
Catalytic Activities ◽  
Enzyme Superfamily ◽  
P450 Enzyme

Background: CYP2C19 is an important member of the cytochrome P450 enzyme superfamily. We recently identified 31CYP2C19 alleles in the Han Chinese population; studying the effects of CYP2C19 on drug metabolism can help reduce adverse drug reactions and therapeutic failure. Aim: The aim of this study was to assess the catalytic activities of 31 allelic isoforms and their effects on the metabolism of clomipramine in vitro. Methods: The wild-type and 30 CYP2C19 variants were expressed in insect cells, and each variant was characterized using clomipramine as the substrate. Reactions were performed at 37°C with 5–150 μmol/L substrate for 30 min. By using ultra-high-performance liquid chromatography-mass spectrometry to detect the products, the kinetic parameters Km, Vmax, and intrinsic clearance (Vmax/Km) of N-desmethyl clomipramine were determined. Results: Among the CYP2C19 variants tested, CYP2C19*29, L16F, and T130M showed extremely increased intrinsic clearance of clomipramine, CYP2C19*3C, and N277K showed similar intrinsic clearance (Vmax/Km) values with CYP2C19*1, while the intrinsic clearance values of other variants were significantly decreased (from 0.65% to 63.28%). In addition, CYP2C19*3 and 35FS could not be detected because they have no detectable enzyme activity. Conclusions: As the first report of 31 CYP2C19 alleles for clomipramine metabolism, our study could provide corresponding reference for clomipramine for further studies in vivo and offer valuable information relevant to the personalized medicine for CYP2C19-metabolized drug.

scholarly journals Endocannabinoid turnover in GtoPdb v.2021.3

2021 ◽  
Vol 2021 (3) ◽  
Author(s):  
Stephen P.H. Alexander ◽  
Patrick Doherty ◽  
Christopher J. Fowler ◽  
Jürg Gertsch ◽  
Mario Van der Stelt
Keyword(s):  
Cytochrome P450 ◽  
Phospholipase D ◽  
Cytochrome P450 Enzyme ◽  
Facilitated Diffusion ◽  
In Vitro Experiments ◽  
Diacylglycerol Lipase ◽  
P450 Enzyme ◽  
Key Enzyme

The principle endocannabinoids are 2-acylglycerol esters, such as 2-arachidonoylglycerol (2-AG), and N-acylethanolamines, such as anandamide (N-arachidonoylethanolamine, AEA). The glycerol esters and ethanolamides are synthesised and hydrolysed by parallel, independent pathways. Mechanisms for release and re-uptake of endocannabinoids are unclear, although potent and selective inhibitors of facilitated diffusion of endocannabinoids across cell membranes have been developed [28]. FABP5 (Q01469) has been suggested to act as a canonical intracellular endocannabinoid transporter in vivo [17]. For the generation of 2-arachidonoylglycerol, the key enzyme involved is diacylglycerol lipase (DAGL), whilst several routes for anandamide synthesis have been described, the best characterized of which involves N-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD, [70]). A transacylation enzyme which forms N-acylphosphatidylethanolamines has been identified as a cytosolic enzyme, PLA2G4E (Q3MJ16) [62]. In vitro experiments indicate that the endocannabinoids are also substrates for oxidative metabolism via cyclooxygenase, lipoxygenase and cytochrome P450 enzyme activities [5, 23, 72].

scholarly journals Endocannabinoid turnover (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

2019 ◽  
Vol 2019 (4) ◽  
Author(s):  
Stephen P.H. Alexander ◽  
Patrick Doherty ◽  
Christopher J. Fowler ◽  
Jürg Gertsch ◽  
Mario Van der Stelt
Keyword(s):  
Cytochrome P450 ◽  
Phospholipase D ◽  
Cytochrome P450 Enzyme ◽  
Facilitated Diffusion ◽  
In Vitro Experiments ◽  
Diacylglycerol Lipase ◽  
P450 Enzyme ◽  
Key Enzyme

The principle endocannabinoids are 2-acylglycerol esters, such as 2-arachidonoylglycerol (2-AG), and N-acylethanolamines, such as anandamide (N-arachidonoylethanolamine, AEA). The glycerol esters and ethanolamides are synthesised and hydrolysed by parallel, independent pathways. Mechanisms for release and re-uptake of endocannabinoids are unclear, although potent and selective inhibitors of facilitated diffusion of endocannabinoids across cell membranes have been developed [19]. FABP5 (Q01469) has been suggested to act as a canonical intracellular endocannabinoid transporter in vivo [12]. For the generation of 2-arachidonoylglycerol, the key enzyme involved is diacylglycerol lipase (DAGL), whilst several routes for anandamide synthesis have been described, the best characterized of which involves N-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD, [49]). A transacylation enzyme which forms N-acylphosphatidylethanolamines has recently been identified as a cytosolic enzyme, PLA2G4E (Q3MJ16) [43]. In vitro experiments indicate that the endocannabinoids are also substrates for oxidative metabolism via cyclooxygenase, lipoxygenase and cytochrome P450 enzyme activities [4, 16, 51].

scholarly journals Reconstitution of theIn VitroActivity of the Cyclosporine-Specific P450 Hydroxylase from Sebekia benihana and Development of a Heterologous Whole-Cell Biotransformation System

2015 ◽  
Vol 81 (18) ◽  
pp. 6268-6275 ◽  
Author(s):  
Li Ma ◽  
Lei Du ◽  
Hui Chen ◽  
Yue Sun ◽  
Shan Huang ◽  
...  
Keyword(s):  
Redox System ◽  
Cytochrome P450 Enzyme ◽  
Regeneration System ◽  
Nadph Regeneration ◽  
Site Specific ◽  
Whole Cell ◽  
Content Type ◽  
P450 Enzyme

ABSTRACTThe cytochrome P450 enzyme CYP-sb21 fromSebekia benihanais capable of catalyzing the site-specific hydroxylation of the immunosuppressant cyclosporine (CsA), leading to the single product γ-hydroxy-N-methyl-l-Leu4-CsA (CsA-4-OH). Unlike authentic CsA, this hydroxylated CsA shows significantly reduced immunosuppressive activity while it retains a side effect of CsA, the hair growth stimulation effect. Although CYP-sb21 was previously identified to be responsible for CsA-specific hydroxylationin vivo, thein vitroactivity of CYP-sb21 has yet to be established for a deeper understanding of this P450 enzyme and further reaction optimization. In this study, we reconstituted thein vitroactivity of CYP-sb21 by using surrogate redox partner proteins of bacterial and cyanobacterial origins. The highest CsA site-specific hydroxylation activity by CYP-sb21 was observed when it was partnered with the cyanobacterial redox system composed ofseFdx andseFdR fromSynechococcus elongatusPCC 7942. The best bioconversion yields were obtained in the presence of 10% methanol as a cosolvent and an NADPH regeneration system. A heterologous whole-cell biocatalyst usingEscherichia coliwas also constructed, and the permeability problem was solved by usingN-cetyl-N,N,N-trimethylammonium bromide (CTAB). This work provides a useful example for reconstituting a hybrid P450 system and developing it into a promising biocatalyst for industrial application.

scholarly journals Unraveling the Metabolic Routes of Retapamulin: Insights into Drug Development of Pleuromutilins

2018 ◽  
Vol 62 (4) ◽  
Author(s):  
Feifei Sun ◽  
Huiyan Zhang ◽  
Gerard Bryan Gonzales ◽  
Jinhui Zhou ◽  
Yi Li ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Metabolic Pathways ◽  
Enzyme Inhibitors ◽  
Side Chain ◽  
Cytochrome P450 Enzyme ◽  
Cytochrome P450 Enzymes ◽  
Short Term ◽  
P450 Enzyme

ABSTRACT Retapamulin, a semisynthetic pleuromutilin derivative, is exclusively used for the topical short-term medication of impetigo and staphylococcal infections. In the present study, we report that retapamulin is adequately and rapidly metabolized in vitro via various metabolic pathways, such as hydroxylation, including mono-, di-, and trihydroxylation, and demethylation. Like tiamulin and valnemulin, the major metabolic routes of retapamulin were hydroxylation at the 2β and 8α positions of the mutilin moiety. Moreover, in vivo metabolism concurred with the results of the in vitro assays. Additionally, we observed significant interspecies differences in the metabolism of retapamulin. Until now, modifying the side chain was the mainstream method for new drug discovery of the pleuromutilins. This approach, however, could not resolve the low bioavailability and short efficacy of the drugs. Considering the rapid metabolism of the pleuromutilins mediated by cytochrome P450 enzymes, we propose that blocking the active metabolic site (C-2 and C-8 motif) or administering the drug in combination with cytochrome P450 enzyme inhibitors is a promising pathway in the development of novel pleuromutilin drugs with slow metabolism and long efficacy.

scholarly journals The role of cytochrome P450 enzymes in carcinogen activation and detoxication: an in vivo–in vitro paradox

2018 ◽  
Vol 39 (7) ◽  
pp. 851-859 ◽  
Author(s):  
Lindsay Reed ◽  
Volker M Arlt ◽  
David H Phillips
Keyword(s):  
Cytochrome P450 ◽  
Cytochrome P450 Enzyme ◽  
Cytochrome P450 Enzymes ◽  
Predominant Role ◽  
Enzyme Systems ◽  
P450 Enzyme

Cytochrome P450 enzyme systems have been widely used in vitro to determine the pathways of activation of procarcinogens, but paradoxically, these same enzymes can play a more predominant role in carcinogen detoxification in vivo.

Enzyme system of Clostridium stercorarium for hydrolysis of arabinoxylan: reconstitution of the in vivo system from recombinant enzymes

Microbiology ◽  
2004 ◽  
Vol 150 (7) ◽  
pp. 2257-2266 ◽  
Author(s):  
Helmuth Adelsberger ◽  
Christian Hertel ◽  
Erich Glawischnig ◽  
Vladimir V. Zverlov ◽  
Wolfgang H. Schwarz
Keyword(s):  
Extracellular Enzymes ◽  
Enzyme System ◽  
Thermophilic Bacterium ◽  
Recombinant Enzymes ◽  
Type Mode ◽  
Complete Set ◽  
First Time ◽  
Hydrolysis Of

Four extracellular enzymes of the thermophilic bacterium Clostridium stercorarium are involved in the depolymerization of de-esterified arabinoxylan: Xyn11A, Xyn10C, Bxl3B, and Arf51B. They were identified in a collection of eight clones producing enzymes hydrolysing xylan (xynA, xynB, xynC), β-xyloside (bxlA, bxlB, bglZ) and α-arabinofuranoside (arfA, arfB). The modular enzymes Xyn11A and Xyn10C represent the major xylanases in the culture supernatant of C. stercorarium. Both hydrolyse arabinoxylan in an endo-type mode, but differ in the pattern of the oligosaccharides produced. Of the glycosidases, Bxl3B degrades xylobiose and xylooligosaccharides to xylose, and Arf51B is able to release arabinose residues from de-esterified arabinoxylan and from the oligosaccharides generated. The other glycosidases either did not attack or only marginally attacked these oligosaccharides. Significantly more xylanase and xylosidase activity was produced during growth on xylose and xylan. This is believed to be the first time that, in a single thermophilic micro-organism, the complete set of enzymes (as well as the respective genes) to completely hydrolyse de-esterified arabinoxylan to its monomeric sugar constituents, xylose and arabinose, has been identified and the enzymes produced in vivo. The active enzyme system was reconstituted in vitro from recombinant enzymes.

Drug Interactions of Macrolides: Emphasis on Dirithromycin

1997 ◽  
Vol 31 (3) ◽  
pp. 349-356 ◽  
Author(s):  
Vish S Watkins ◽  
Ron E Polk ◽  
Jennifer L Stotka
Keyword(s):  
Cytochrome P450 ◽  
Drug Interactions ◽  
Ethinyl Estradiol ◽  
Enzyme System ◽  
Cytochrome P450 Enzyme ◽  
Cytochrome P450 Enzymes ◽  
Kidney Transplant Recipients ◽  
P450 Enzyme ◽  
Cytochrome P450 Enzyme System ◽  
Clinically Significant

Objective To describe the drug interactions of dirithromycin, a new macrolide, and to compare them with those of other macrolides. Data Sources A literature search was performed using MEDLINE to identify articles published between January 1980 and July 1995 concerning the drug interactions of macrolides. Published abstracts were also examined. All studies using dirithromycin were performed under the sponsorship of Eli Lilly and Company. Data Synthesis Erythromycin, the first macrolide discovered, is metabolized by the cytochrome P450 enzyme system. By decreasing their metabolism, erythromycin can interact with other drugs metabolized by the cytochrome P450 enzymes. The lack of such interactions would be a desirable feature in a newer macrolide. We describe studies performed to detect any interactions of dirithromycin with cyclosporine, theophylline, terfenadine, warfarin, and ethinyl estradiol. The studies showed that dirithromycin, like azithromycin, is much less likely to cause the interactions detected with clarithromycin and erythromycin. A review of the literature showed differences among macrolides in their abilities to inhibit cytochrome P450 enzymes and, thus, to cause drug–drug interactions. Erythromycin and clarithromycin inhibit cytochrome P450 enzymes, and have been implicated in clinically significant interactions. Azithromycin and dirithromycin neither inhibit cytochrome P450 enzymes nor are implicated in clinically significant drug–drug interactions. Conclusions Dirithromycin, a new macrolide, does not inhibit the cytochrome P450 enzyme system. The concomitant use of dirithromycin with cyclosporine, theophylline, terfenadine, warfarin, or ethinyl estradiol was studied in pharmacokinetic and pharmacodynamic studies. In vitro, dirithromycin did not bind cytochrome P450. In healthy subjects, erythromycin increases the clearance of cyclosporine by 51%, whereas dirithromycin causes no significant changes in the pharmacokinetics of cyclosporine. In kidney transplant recipients, administration of dirithromycin was associated with a significant (p < 0.003) decrease of 17.4% in the clearance of cyclosporine. In patients taking low-dose estradiol, the administration of dirithromycin caused a significant (p < 0.03) increase of 9.9% in the clearance of ethinyl estradiol; escape ovulation did not occur. Unlike erythromycin and clarithromycin, dirithromycin had no significant effects on the pharmacokinetics of theophylline, terfenadine, or warfarin. The alterations typical of drug interactions that are based on inhibition of the cytochrome P450 system occurring with erythromycin and clarithromycin were not observed with dirithromycin.

Biosynthesis of Nitrogenous Phospholipids in Spinach Leaves

1974 ◽  
Vol 52 (6) ◽  
pp. 469-482 ◽  
Author(s):  
M. O. Marshall ◽  
M. Kates
Keyword(s):  
Microsomal Fraction ◽  
Enzyme System ◽  
Supernatant Fraction ◽  
Exchange Reactions ◽  
Methylation Pathway ◽  
Ethanolamine Kinase ◽  
Spinach Leaves

Pathways for biosynthesis of phosphatidylserine (PS), phosphatidylethanolamine (PE), and phosphatidylcholine (PC), in spinach leaves have been studied both in vivo (whole leaves and leaf slices) and in vitro (cell-free leaf fractions). Biosynthesis of PS was shown to occur by the action of a particle-bound CDP-diglyceride: serine phosphatidyltransferase, and PE by the action of a PS-decarboxylase localized in the 100 000 × g supernatant fraction. PE was also formed by the operation of the CDP-ethanolamine:diglyceride phosphorylethanolamine transferase, localized in the microsomal fraction. The presence of ethanolamine kinase required for formation of phosphorylethanolamine was demonstrated in vitro, but not the presence of CTP:phosphorylethanolamine cytidyltransferase; however, the latter is presumed present on the basis of in vivo results. Operation of the methylation pathway for biosynthesis of PC was established in vivo, and direct methylation of phosphatidyl-N-methylethanolamine to phosphatidyl-N,N-dimethylethanolamine (PE-diMe) and of PE-diME to PC by S-adenosylmethionine was demonstrated with a particulate enzyme system localized in the microsomal fraction; direct methylation of PE itself could not be shown in this system. PC was also synthesized by the CDP-choline:diglyceride phosphorylcholine transferase system localized in the microsomal fraction. Synthesis of PE and PC by Ca2+-stimulated exchange reactions with ethanolamine and choline, respectively, could be demonstrated, but at low rates. However, no synthesis of PS by exchange reactions with serine could be detected.

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