International Journal of Molecular Sciences

Research

Jump to: Review

23 pages, 5748 KiB

Open AccessArticle

Evaluation of the Multimycotoxin-Degrading Efficiency of Rhodococcus erythropolis NI1 Strain with the Three-Step Zebrafish Microinjection Method

by Edina Garai, Anita Risa, Emese Varga, Mátyás Cserháti, Balázs Kriszt, Béla Urbányi and Zsolt Csenki

Int. J. Mol. Sci. 2021, 22(2), 724; https://doi.org/10.3390/ijms22020724 - 13 Jan 2021

Cited by 13 | Viewed by 2701

Abstract

The multimycotoxin-degrading efficiency of the Rhodococcus erythropolis NI1 strain was investigated with a previously developed three-step method. NI1 bacterial metabolites, single and combined mycotoxins and their NI1 degradation products, were injected into one cell stage zebrafish embryos in the same doses. Toxic and [...] Read more.

The multimycotoxin-degrading efficiency of the Rhodococcus erythropolis NI1 strain was investigated with a previously developed three-step method. NI1 bacterial metabolites, single and combined mycotoxins and their NI1 degradation products, were injected into one cell stage zebrafish embryos in the same doses. Toxic and interaction effects were supplemented with UHPLC-MS/MS measurement of toxin concentrations. Results showed that the NI1 strain was able to degrade mycotoxins and their mixtures in different proportions, where a higher ratio of mycotoxins were reduced in combination than single ones. The NI1 strain reduced the toxic effects of mycotoxins and mixtures, except for the AFB1+T-2 mixture. Degradation products of the AFB1+T-2 mixture by the NI1 strain were more toxic than the initial AFB1+T-2 mixture, while the analytical results showed very high degradation, which means that the NI1 strain degraded this mixture to toxic degradation products. The NI1 strain was able to detoxify the AFB1, ZEN, T-2 toxins and mixtures (except for AFB1+T-2 mixture) during the degradation experiments, which means that the NI1 strain degraded these to non-toxic degradation products. The results demonstrate that single exposures of mycotoxins were very toxic. The combined exposure of mycotoxins had synergistic effects, except for ZEN+T-2 and AFB1+ZEN +T-2, whose mixtures had very strong antagonistic effects. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

18 pages, 6121 KiB

Open AccessArticle

Characterization of In Vitro and In Vivo Metabolism of Antazoline Using Liquid Chromatography-Tandem Mass Spectrometry

by Joanna Giebułtowicz, Natalia Korytowska, Roman Piotrowski, Piotr Kułakowski, Gniewomir Latacz, Ewa Szymańska, Barbara Wiśniowska and Sebastian Polak

Int. J. Mol. Sci. 2020, 21(24), 9693; https://doi.org/10.3390/ijms21249693 - 18 Dec 2020

Cited by 3 | Viewed by 2395

Abstract

Antazoline (ANT) was recently shown to be an effective and safe antiarrhythmic drug in the termination of atrial fibrillation. However, the drug is still not listed in clinical guidelines. No data on ANT metabolism in humans is available. We used liquid chromatography coupled [...] Read more.

Antazoline (ANT) was recently shown to be an effective and safe antiarrhythmic drug in the termination of atrial fibrillation. However, the drug is still not listed in clinical guidelines. No data on ANT metabolism in humans is available. We used liquid chromatography coupled with tandem mass spectrometry to identify and characterize metabolites of ANT. We analyzed plasma of volunteers following a single intravenous administration of 100 mg of ANT mesylate and in in vitro cultures of human hepatocytes. We revealed that ANT was transformed into at least 15 metabolites and we investigated the role of cytochrome P450 isoforms. CYP2D6 was the main one involved in the fast metabolism of ANT. The biotransformation of ANT by CYP2C19 was much slower. The main Phase I metabolite was M1 formed by the removal of phenyl and metabolite M2 with hydroxyl in the para position of phenyl. Glucuronidation was the leading Phase II metabolism. Further study on pharmacokinetics of the metabolites would allow us to better understand the activity profile of ANT and to predict its potential clinical applications. Ultimately, further investigation of the activity profile of the new hydroxylated M2 metabolite of ANT might result in an active substance with a different pharmacological profile than the parent molecule, and potentially a new drug candidate. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

13 pages, 1691 KiB

Open AccessArticle

Identification of In Vitro Metabolites of Synthetic Phenolic Antioxidants BHT, BHA, and TBHQ by LC-HRMS/MS

by Ons Ousji and Lekha Sleno

Int. J. Mol. Sci. 2020, 21(24), 9525; https://doi.org/10.3390/ijms21249525 - 15 Dec 2020

Cited by 32 | Viewed by 7429

Abstract

Butylated hydroxytoluene (BHT) and its analogs, butylated hydroxyanisole (BHA) and tert-butyl-hydroquinone (TBHQ), are widely used synthetic preservatives to inhibit lipid oxidation in the food, cosmetic and pharmaceutical industries. Despite their widespread use, little is known about their human exposure and related biotransformation products. [...] Read more.

Butylated hydroxytoluene (BHT) and its analogs, butylated hydroxyanisole (BHA) and tert-butyl-hydroquinone (TBHQ), are widely used synthetic preservatives to inhibit lipid oxidation in the food, cosmetic and pharmaceutical industries. Despite their widespread use, little is known about their human exposure and related biotransformation products. The metabolism of these compounds was investigated using in vitro incubations with human and rat liver fractions. Liquid chromatography coupled to high-resolution tandem mass spectrometry was employed to detect and characterize stable and reactive species formed via oxidative metabolism, as well as phase II conjugates. Several oxidative metabolites have been detected, as well as glutathione, glucuronide, and sulfate conjugates, many of which were not previously reported. A combination of accurate mass measurements, MS/MS fragmentation behavior, and isotope-labeling studies were used to elucidate metabolite structures. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Graphical abstract

22 pages, 4771 KiB

Open AccessArticle

Diclofenac Degradation—Enzymes, Genetic Background and Cellular Alterations Triggered in Diclofenac-Metabolizing Strain Pseudomonas moorei KB4

by Joanna Żur, Artur Piński, Danuta Wojcieszyńska, Wojciech Smułek and Urszula Guzik

Int. J. Mol. Sci. 2020, 21(18), 6786; https://doi.org/10.3390/ijms21186786 - 16 Sep 2020

Cited by 20 | Viewed by 4141

Abstract

Diclofenac (DCF) constitutes one of the most significant ecopollutants detected in various environmental matrices. Biological clean-up technologies that rely on xenobiotics-degrading microorganisms are considered as a valuable alternative for chemical oxidation methods. Up to now, the knowledge about DCF multi-level influence on bacterial [...] Read more.

Diclofenac (DCF) constitutes one of the most significant ecopollutants detected in various environmental matrices. Biological clean-up technologies that rely on xenobiotics-degrading microorganisms are considered as a valuable alternative for chemical oxidation methods. Up to now, the knowledge about DCF multi-level influence on bacterial cells is fragmentary. In this study, we evaluate the degradation potential and impact of DCF on Pseudomonas moorei KB4 strain. In mono-substrate culture KB4 metabolized 0.5 mg L⁻¹ of DCF, but supplementation with glucose (Glc) and sodium acetate (SA) increased degraded doses up to 1 mg L⁻¹ within 12 days. For all established conditions, 4′-OH-DCF and DCF-lactam were identified. Gene expression analysis revealed the up-regulation of selected genes encoding biotransformation enzymes in the presence of DCF, in both mono-substrate and co-metabolic conditions. The multifactorial analysis of KB4 cell exposure to DCF showed a decrease in the zeta-potential with a simultaneous increase in the cell wall hydrophobicity. Magnified membrane permeability was coupled with the significant increase in the branched (19:0 anteiso) and cyclopropane (17:0 cyclo) fatty acid accompanied with reduced amounts of unsaturated ones. DCF injures the cells which is expressed by raised activities of acid and alkaline phosphatases as well as formation of lipids peroxidation products (LPX). The elevated activity of superoxide dismutase (SOD) and catalase (CAT) testified that DCF induced oxidative stress. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

17 pages, 3124 KiB

Open AccessArticle

Metabolomics Fingerprint Induced by the Intranigral Inoculation of Exogenous Human Alpha-Synuclein Oligomers in a Rat Model of Parkinson’s Disease

by Federica Murgia, Luigi Atzori, Ezio Carboni, Maria Laura Santoru, Aran Hendren, Augusta Pisanu, Pierluigi Caboni, Laura Boi, Giuliana Fusco and Anna R. Carta

Int. J. Mol. Sci. 2020, 21(18), 6745; https://doi.org/10.3390/ijms21186745 - 14 Sep 2020

Cited by 4 | Viewed by 2723

Abstract

Parkinson’s disease (PD) is considered a synucleinopathy because of the intraneuronal accumulation of aggregated α-synuclein (αSyn). Recent evidence points to soluble αSyn-oligomers (αSynO) as the main cytotoxic species responsible for cell death. Given the pivotal role of αSyn in PD, αSyn-based models are [...] Read more.

Parkinson’s disease (PD) is considered a synucleinopathy because of the intraneuronal accumulation of aggregated α-synuclein (αSyn). Recent evidence points to soluble αSyn-oligomers (αSynO) as the main cytotoxic species responsible for cell death. Given the pivotal role of αSyn in PD, αSyn-based models are crucial for the investigation of toxic mechanisms and the identification of new therapeutic targets in PD. By using a metabolomics approach, we evaluated the metabolic profile of brain and serum samples of rats infused unilaterally with preformed human αSynOs (HαSynOs), or vehicle, into the substantia nigra pars compacta (SNpc). Three months postinfusion, the striatum was dissected for striatal dopamine (DA) measurements via High Pressure Liquid Chromatography (HPLC) analysis and mesencephalon and serum samples were collected for the evaluation of metabolite content via gas chromatography mass spectrometry analysis. Multivariate, univariate and correlation statistics were applied. A 40% decrease of DA content was measured in the HαSynO-infused striatum as compared to the contralateral and the vehicle-infused striata. Decreased levels of dehydroascorbic acid, myo-inositol, and glycine, and increased levels of threonine, were found in the mesencephalon, while increased contents of fructose and mannose, and a decrease in glycine and urea, were found in the serum of HαSynO-infused rats. The significant correlation between DA and metabolite content indicated that metabolic variations reflected the nigrostriatal degeneration. Collectively, the metabolomic fingerprint of HαSynO-infused rats points to an increase of oxidative stress markers, in line with PD neuropathology, and provides hints for potential biomarkers of PD. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Graphical abstract

22 pages, 2924 KiB

Open AccessArticle

Biotransformation of Methoxyflavones by Selected Entomopathogenic Filamentous Fungi

by Mateusz Łużny, Tomasz Tronina, Ewa Kozłowska, Monika Dymarska, Jarosław Popłoński, Jacek Łyczko, Edyta Kostrzewa-Susłow and Tomasz Janeczko

Int. J. Mol. Sci. 2020, 21(17), 6121; https://doi.org/10.3390/ijms21176121 - 25 Aug 2020

Cited by 20 | Viewed by 3106

Abstract

The synthesis and biotransformation of five flavones containing methoxy substituents in the B ring: 2′-, 3′-, 4′-methoxyflavones, 2′,5′-dimethoxyflavone and 3′,4′,5′-trimethoxyflavone are described. Strains of entomopathogenic filamentous fungi were used as biocatalysts. Five strains of the species Beauveria bassiana (KCh J1.5, J2.1, J3.2, J1, [...] Read more.

The synthesis and biotransformation of five flavones containing methoxy substituents in the B ring: 2′-, 3′-, 4′-methoxyflavones, 2′,5′-dimethoxyflavone and 3′,4′,5′-trimethoxyflavone are described. Strains of entomopathogenic filamentous fungi were used as biocatalysts. Five strains of the species Beauveria bassiana (KCh J1.5, J2.1, J3.2, J1, BBT), two of the species Beauveria caledonica (KCh J3.3, J3.4), one of Isaria fumosorosea (KCh J2) and one of Isaria farinosa (KCh KW 1.1) were investigated. Both the number and the place of attachment of the methoxy groups in the flavonoid structure influenced the biotransformation rate and the amount of nascent products. Based on the structures of products and semi-products, it can be concluded that their formation is the result of a cascading process. As a result of enzymes produced in the cells of the tested strains, the test compounds undergo progressive demethylation and/or hydroxylation and 4-O-methylglucosylation. Thirteen novel flavonoid 4-O-methylglucosides and five hydroxy flavones were isolated and identified. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

21 pages, 7458 KiB

Open AccessArticle

Revealing Metabolic Perturbation Following Heavy Methamphetamine Abuse by Human Hair Metabolomics and Network Analysis

by Suji Kim, Won-Jun Jang, Hyerim Yu, Jihyun Kim, Sang-Ki Lee, Chul-Ho Jeong and Sooyeun Lee

Int. J. Mol. Sci. 2020, 21(17), 6041; https://doi.org/10.3390/ijms21176041 - 21 Aug 2020

Cited by 24 | Viewed by 5104

Abstract

Methamphetamine (MA) is a highly addictive central nervous system stimulant. Drug addiction is not a static condition but rather a chronically relapsing disorder. Hair is a valuable and stable specimen for chronic toxicological monitoring as it retains toxicants and metabolites. The primary focus [...] Read more.

Methamphetamine (MA) is a highly addictive central nervous system stimulant. Drug addiction is not a static condition but rather a chronically relapsing disorder. Hair is a valuable and stable specimen for chronic toxicological monitoring as it retains toxicants and metabolites. The primary focus of this study was to discover the metabolic effects encompassing diverse pathological symptoms of MA addiction. Therefore, metabolic alterations were investigated in human hair following heavy MA abuse using both targeted and untargeted mass spectrometry and through integrated network analysis. The statistical analyses (t-test, variable importance on projection score, and receiver-operator characteristic curve) demonstrated that 32 metabolites (in targeted metabolomics) as well as 417 and 224 ion features (in positive and negative ionization modes of untargeted metabolomics, respectively) were critically dysregulated. The network analysis showed that the biosynthesis or metabolism of lipids, such as glycosphingolipids, sphingolipids, glycerophospholipids, and ether lipids, as well as the metabolism of amino acids (glycine, serine and threonine; cysteine and methionine) is affected by heavy MA abuse. These findings reveal crucial metabolic effects caused by MA addiction, with emphasis on the value of human hair as a diagnostic specimen for determining drug addiction, and will aid in identifying robust diagnostic markers and therapeutic targets. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

12 pages, 869 KiB

Open AccessArticle

The Identification of Metabolites and Effects of Albendazole in Alfalfa (Medicago sativa)

by Lucie Raisová Stuchlíková, Martina Navrátilová, Lenka Langhansová, Kateřina Moťková, Radka Podlipná, Barbora Szotáková and Lenka Skálová

Int. J. Mol. Sci. 2020, 21(16), 5943; https://doi.org/10.3390/ijms21165943 - 18 Aug 2020

Cited by 5 | Viewed by 3283

Abstract

Albendazole (ABZ), a widely used anthelmintic drug, enters the environment mainly via livestock excrements. To evaluate the environmental impact of ABZ, the knowledge of its uptake, effects and metabolism in all non-target organisms, including plants, is essential. The present study was designed to [...] Read more.

Albendazole (ABZ), a widely used anthelmintic drug, enters the environment mainly via livestock excrements. To evaluate the environmental impact of ABZ, the knowledge of its uptake, effects and metabolism in all non-target organisms, including plants, is essential. The present study was designed to identify the metabolic pathway of ABZ and to test potential ABZ phytotoxicity in fodder plant alfalfa, with seeds and in vitro regenerants used for these purposes. Alfalfa was chosen, as it may meet manure from ABZ-treated animals in pastures and fields. Alfalfa is often used as a feed of livestock, which might already be infected with helminths. The obtained results showed that ABZ did not inhibit alfalfa seed germination and germ growth, but evoked stress and a toxic effect in alfalfa regenerants. Alfalfa regenerants were able to uptake ABZ and transform it into 21 metabolites. UHPLC-MS/MS analysis revealed three new ABZ metabolites that have not been described yet. The discovery of the parent compound ABZ together with the anthelmintically active and instable metabolites in alfalfa leaves shows that the contact of fodder plants with ABZ-containing manure might represent not only a danger for herbivorous invertebrates, but also may cause the development of ABZ resistance in helminths. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Graphical abstract

12 pages, 805 KiB

Open AccessArticle

Degradation Mechanism of 2,4-Dichlorophenol by Fungi Isolated from Marine Invertebrates

by Efstratios Nikolaivits, Andreas Agrafiotis, Eirini Baira, Géraldine Le Goff, Nikolaos Tsafantakis, Suchana A. Chavanich, Yehuda Benayahu, Jamal Ouazzani, Nikolas Fokialakis and Evangelos Topakas

Int. J. Mol. Sci. 2020, 21(9), 3317; https://doi.org/10.3390/ijms21093317 - 7 May 2020

Cited by 12 | Viewed by 4001

Abstract

2,4-Dichlorophenol (2,4-DCP) is a ubiquitous environmental pollutant categorized as a priority pollutant by the United States (US) Environmental Protection Agency, posing adverse health effects on humans and wildlife. Bioremediation is proposed as an eco-friendly, cost-effective alternative to traditional physicochemical remediation techniques. In the [...] Read more.

2,4-Dichlorophenol (2,4-DCP) is a ubiquitous environmental pollutant categorized as a priority pollutant by the United States (US) Environmental Protection Agency, posing adverse health effects on humans and wildlife. Bioremediation is proposed as an eco-friendly, cost-effective alternative to traditional physicochemical remediation techniques. In the present study, fungal strains were isolated from marine invertebrates and tested for their ability to biotransform 2,4-DCP at a concentration of 1 mM. The most competent strains were studied further for the expression of catechol dioxygenase activities and the produced metabolites. One strain, identified as Tritirachium sp., expressed high levels of extracellular catechol 1,2-dioxygenase activity. The same strain also produced a dechlorinated cleavage product of the starting compound, indicating the assimilation of the xenobiotic by the fungus. This work also enriches the knowledge about the mechanisms employed by marine-derived fungi in order to defend themselves against chlorinated xenobiotics. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Graphical abstract

Review

Jump to: Research

30 pages, 5686 KiB

Open AccessReview

Legacy and Emerging Per- and Polyfluoroalkyl Substances: Analytical Techniques, Environmental Fate, and Health Effects

by Richard A. Brase, Elizabeth J. Mullin and David C. Spink

Int. J. Mol. Sci. 2021, 22(3), 995; https://doi.org/10.3390/ijms22030995 - 20 Jan 2021

Cited by 156 | Viewed by 13365

Abstract

Due to their unique chemical properties, per- and polyfluoroalkyl substances (PFAS) have been used extensively as industrial surfactants and processing aids. While several types of PFAS have been voluntarily phased out by their manufacturers, these chemicals continue to be of ecological and public [...] Read more.

Due to their unique chemical properties, per- and polyfluoroalkyl substances (PFAS) have been used extensively as industrial surfactants and processing aids. While several types of PFAS have been voluntarily phased out by their manufacturers, these chemicals continue to be of ecological and public health concern due to their persistence in the environment and their presence in living organisms. Moreover, while the compounds referred to as “legacy” PFAS remain in the environment, alternative compounds have emerged as replacements for their legacy predecessors and are now detected in numerous matrices. In this review, we discuss the historical uses of PFAS, recent advances in analytical techniques for analysis of these compounds, and the fate of PFAS in the environment. In addition, we evaluate current biomonitoring studies of human exposure to legacy and emerging PFAS and examine the associations of PFAS exposure with human health impacts, including cancer- and non-cancer-related outcomes. Special focus is given to short-chain perfluoroalkyl acids (PFAAs) and ether-substituted, polyfluoroalkyl alternatives including hexafluoropropylene oxide dimer acid (HFPO-DA; tradename GenX), 4,8-dioxa-3H-perfluorononanoic acid (DONA), and 6:2 chlorinated polyfluoroethersulfonic acid (6:2 Cl-PFESA; tradename F-53B). Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

46 pages, 4768 KiB

Open AccessReview

A Review on the Fate of Legacy and Alternative Antimicrobials and Their Metabolites during Wastewater and Sludge Treatment

by Timothy Abbott, Gokce Kor-Bicakci, Mohammad S. Islam and Cigdem Eskicioglu

Int. J. Mol. Sci. 2020, 21(23), 9241; https://doi.org/10.3390/ijms21239241 - 3 Dec 2020

Cited by 22 | Viewed by 4253

Abstract

Antimicrobial compounds are used in a broad range of personal care, consumer and healthcare products and are frequently encountered in modern life. The use of these compounds is being reexamined as their safety, effectiveness and necessity are increasingly being questioned by regulators and [...] Read more.

Antimicrobial compounds are used in a broad range of personal care, consumer and healthcare products and are frequently encountered in modern life. The use of these compounds is being reexamined as their safety, effectiveness and necessity are increasingly being questioned by regulators and consumers alike. Wastewater often contains significant amounts of these chemicals, much of which ends up being released into the environment as existing wastewater and sludge treatment processes are simply not designed to treat many of these contaminants. Furthermore, many biotic and abiotic processes during wastewater treatment can generate significant quantities of potentially toxic and persistent antimicrobial metabolites and byproducts, many of which may be even more concerning than their parent antimicrobials. This review article explores the occurrence and fate of two of the most common legacy antimicrobials, triclosan and triclocarban, their metabolites/byproducts during wastewater and sludge treatment and their potential impacts on the environment. This article also explores the fate and transformation of emerging alternative antimicrobials and addresses some of the growing concerns regarding these compounds. This is becoming increasingly important as consumers and regulators alike shift away from legacy antimicrobials to alternative chemicals which may have similar environmental and human health concerns. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

17 pages, 618 KiB

Open AccessReview

Application of In Vitro Metabolism Activation in High-Throughput Screening

by Masato Ooka, Caitlin Lynch and Menghang Xia

Int. J. Mol. Sci. 2020, 21(21), 8182; https://doi.org/10.3390/ijms21218182 - 31 Oct 2020

Cited by 30 | Viewed by 6726

Abstract

In vitro methods which incorporate metabolic capability into the assays allow us to assess the activity of metabolites from their parent compounds. These methods can be applied into high-throughput screening (HTS) platforms, thereby increasing the speed to identify compounds that become active via [...] Read more.

In vitro methods which incorporate metabolic capability into the assays allow us to assess the activity of metabolites from their parent compounds. These methods can be applied into high-throughput screening (HTS) platforms, thereby increasing the speed to identify compounds that become active via the metabolism process. HTS was originally used in the pharmaceutical industry and now is also used in academic settings to evaluate biological activity and/or toxicity of chemicals. Although most chemicals are metabolized in our body, many HTS assays lack the capability to determine compound activity via metabolism. To overcome this problem, several in vitro metabolic methods have been applied to an HTS format. In this review, we describe in vitro metabolism methods and their application in HTS assays, as well as discuss the future perspectives of HTS with metabolic activity. Each in vitro metabolism method has advantages and disadvantages. For instance, the S9 mix has a full set of liver metabolic enzymes, but it displays high cytotoxicity in cell-based assays. In vitro metabolism requires liver fractions or the use of other metabolically capable systems, including primary hepatocytes or recombinant enzymes. Several newly developed in vitro metabolic methods, including HepaRG cells, three-dimensional (3D) cell models, and organ-on-a-chip technology, will also be discussed. These newly developed in vitro metabolism approaches offer significant progress in dissecting biological processes, developing drugs, and making toxicology studies quicker and more efficient. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

22 pages, 1110 KiB

Open AccessReview

Metabolomic Approaches to Study Chemical Exposure-Related Metabolism Alterations in Mammalian Cell Cultures

by Aneta Balcerczyk, Christian Damblon, Bénédicte Elena-Herrmann, Baptiste Panthu and Gilles J. P. Rautureau

Int. J. Mol. Sci. 2020, 21(18), 6843; https://doi.org/10.3390/ijms21186843 - 18 Sep 2020

Cited by 20 | Viewed by 4430

Abstract

Biological organisms are constantly exposed to an immense repertoire of molecules that cover environmental or food-derived molecules and drugs, triggering a continuous flow of stimuli-dependent adaptations. The diversity of these chemicals as well as their concentrations contribute to the multiplicity of induced effects, [...] Read more.

Biological organisms are constantly exposed to an immense repertoire of molecules that cover environmental or food-derived molecules and drugs, triggering a continuous flow of stimuli-dependent adaptations. The diversity of these chemicals as well as their concentrations contribute to the multiplicity of induced effects, including activation, stimulation, or inhibition of physiological processes and toxicity. Metabolism, as the foremost phenotype and manifestation of life, has proven to be immensely sensitive and highly adaptive to chemical stimuli. Therefore, studying the effect of endo- or xenobiotics over cellular metabolism delivers valuable knowledge to apprehend potential cellular activity of individual molecules and evaluate their acute or chronic benefits and toxicity. The development of modern metabolomics technologies such as mass spectrometry or nuclear magnetic resonance spectroscopy now offers unprecedented solutions for the rapid and efficient determination of metabolic profiles of cells and more complex biological systems. Combined with the availability of well-established cell culture techniques, these analytical methods appear perfectly suited to determine the biological activity and estimate the positive and negative effects of chemicals in a variety of cell types and models, even at hardly detectable concentrations. Metabolic phenotypes can be estimated from studying intracellular metabolites at homeostasis in vivo, while in vitro cell cultures provide additional access to metabolites exchanged with growth media. This article discusses analytical solutions available for metabolic phenotyping of cell culture metabolism as well as the general metabolomics workflow suitable for testing the biological activity of molecular compounds. We emphasize how metabolic profiling of cell supernatants and intracellular extracts can deliver valuable and complementary insights for evaluating the effects of xenobiotics on cellular metabolism. We note that the concepts and methods discussed primarily for xenobiotics exposure are widely applicable to drug testing in general, including endobiotics that cover active metabolites, nutrients, peptides and proteins, cytokines, hormones, vitamins, etc. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

17 pages, 2401 KiB

Open AccessReview

Coumarin-Based Profluorescent and Fluorescent Substrates for Determining Xenobiotic-Metabolizing Enzyme Activities In Vitro

by Hannu Raunio, Olli Pentikäinen and Risto O. Juvonen

Int. J. Mol. Sci. 2020, 21(13), 4708; https://doi.org/10.3390/ijms21134708 - 1 Jul 2020

Cited by 29 | Viewed by 5454

Abstract

in vivo methods, such as spectrophotometric, fluorometric, mass spectrometric,
and radioactivity-based techniques. In fluorescence-based assays, the reaction produces a fluorescent
product from a nonfluorescent substrate or vice versa. Fluorescence-based enzyme assays are
usually highly sensitive and specific, allowing measurements on small specimens of [...] Read more.

in vivo methods, such as spectrophotometric, fluorometric, mass spectrometric,
and radioactivity-based techniques. In fluorescence-based assays, the reaction produces a fluorescent
product from a nonfluorescent substrate or vice versa. Fluorescence-based enzyme assays are
usually highly sensitive and specific, allowing measurements on small specimens of tissues with
low enzyme activities. Fluorescence assays are also amenable to miniaturization of the reaction
mixtures and can thus be done in high throughput. 7-Hydroxycoumarin and its derivatives are
widely used as fluorophores due to their desirable photophysical properties. They possess a large -
conjugated system with electron-rich and charge transfer properties. This conjugated structure leads
to applications of 7-hydroxycoumarins as fluorescent sensors for biological activities. We describe in
this review historical highlights and current use of coumarins and their derivatives in evaluating
activities of the major types of xenobiotic-metabolizing enzyme systems. Traditionally, coumarin
substrates have been used to measure oxidative activities of cytochrome P450 (CYP) enzymes. For this
purpose, profluorescent coumarins are very sensitive, but generally lack selectivity for individual CYP
forms. With the aid of molecular modeling, we have recently described several new coumarin-based
substrates for measuring activities of CYP and conjugating enzymes with improved selectivity. Full article

(This article belongs to the Special Issue Identification of Metabolites of Xenobiotics)

► Show Figures

Figure 1

Journal Menu

Journal Browser

Identification of Metabolites of Xenobiotics

Share This Special Issue

Special Issue Editor

Special Issue Information

Keywords

Benefits of Publishing in a Special Issue

Related Special Issue

Published Papers (14 papers)

Research

Review

Further Information

Guidelines

MDPI Initiatives

Follow MDPI