Toxin Metabolites Research Articles

AFB1 Toxicity in Human Food and Animal Feed Consumption: A Review of Experimental Treatments and Preventive Measures.

The current review aims to outline and summarize the latest research on aflatoxin, with research studies describing natural, herbal and chemical compound applications in animal (pig) models and in vitro cellular studies. Aflatoxin, a carcinogenic toxin metabolite, is produced by Aspergillus flavus in humid environments, posing a threat to human health and crop production. The current treatment involves the prevention of exposure to aflatoxin and counteracting its harmful toxic effects, enabling survival and research studies on an antidote for aflatoxin. To summarize current research prospects and to outline the influence of aflatoxin on animal forage in farm production, food and crop processing. The research application of remedies to treat aflatoxin is undergoing development to pinpoint biochemical pathways responsible for aflatoxin effects transmission and actions of treatment. To underline the environmental stress of aflatoxin on meat and dairy products; to describe clinical syndromes associated with aflatoxicosis on human health that are counteracted with proposed treatment and preventive interventions. To understand how to improve the health of farm animals with feed conditions.

Formation mechanism and environmental drivers of Alexandrium catenella bloom events in the coastal waters of Qinhuangdao, China

In the last 5 years, paralytic shellfish toxins (PSTs) have been recurrently detected in mollusks farmed in the mussel culture area of Qinhuangdao city, along with the occurrence of toxic outbreaks linked to dinoflagellate species of the Alexandrium genus. To understand the formation mechanism and variation of these events, continuous and comprehensive PSTs monitoring was carried out between 2017 and 2020. Through the analysis of both phytoplankton and cysts via light microscopy and quantitative polymerase chain reaction, it was shown that Alexandrium catenella was responsible for the production of PSTs, which consisted mainly of gonyautoxins 1,4 (GTX1/4, 87%) and GTX2/3 (13%). During bloom events in 2019, mussels accumulated the highest PSTs value (929 μg STX di-HCl eq·kg−1) in conjunction with the peak of cell abundances, and toxin profiles were consistent with high distributions of GTX1/4, GTX2/3, and Neosaxitoxin. Toxin metabolites vary in different substances and mainly transferred to a stable proportion of α-epimer: β-epimers 3:1. The environmental drivers of Alexandrium blooms included the continuous rise of water temperature (>4 °C) and calm weather with low wind speed and no significant precipitation. By comparing toxin profiles and method sensitivity, it was found that dissolved toxins in seawater are more useful for early warning. These results have important implications for the effective monitoring and management of paralytic shellfish poisoning outbreaks.

Variations in the toxicity and condition index of five bivalve species throughout a red tide event caused by Alexandrium catenella: A field study

Harmful red tides in China have caused paralytic shellfish toxins (PSTs) pollution and led to severe socioeconomic effects in shellfish aquaculture. Although shellfish can survive harmful algal blooms, the effects on their Condition Index (CI) have been underestimated. This study sought to evaluate the effects of the profiles and levels of paralytic shellfish toxins on variations in the CI in bivalves under natural blooming conditions. We observed clear soft tissue lesions to varying degrees except in Mytilus galloprovincialis after toxin exposure. Among the five species of shellfish exposed in situ, only M. galloprovincialis accumulated PSTs content above the maximum permitted level (800 μg STX di-HCl eq./kg). The highest toxin content in all sample tissues was observed in Patinopecten yessoensis. Significant interspecies differences in PSTs accumulation among the five bivalve species were observed in the hepatopancreas. A total of nine PSTs components and four new C-11 hydroxyl metabolites (so-called M-toxins) toxins were detected, and detoxification diversity was observed among bivalves. We observed a higher proportion of M-toxin in early stages, and the proportions changed only slightly over time in M. galloprovincialis and Magallana gigas, thus accounting for the significantly higher metabolism rate. Notably, the CI in M. gigas and Argopecten irradians was positively correlated with lowest toxin accumulation of PSTs content, but significantly inhibited. In conclusion, our results revealed a significant inhibitory effect on the CI in shellfish, in a species specific manner, with distinct levels of inhibition correlated with different toxin metabolites. Our study revealed the toxin content of different bivalves exposed to a natural red tide environment and the consequent effects on growth, thus building a foundation for research on the mechanisms underlying the effects of PSTs on growth. These data establish the ecological and economic significance of the effects of harmful algal blooms on bivalves.

Characterization of NRPS and PKS genes involved in the biosynthesis of SMs in Alternaria dauci including the phytotoxic polyketide aldaulactone

Alternaria dauci is a Dothideomycete fungus, causal agent of carrot leaf blight. As a member of the Alternaria genus, known to produce a lot of secondary metabolite toxins, A. dauci is also supposed to synthetize host specific and non-host specific toxins playing a crucial role in pathogenicity. This study provides the first reviewing of secondary metabolism genetic basis in the Alternaria genus by prediction of 55 different putative core genes. Interestingly, aldaulactone, a phytotoxic benzenediol lactone from A. dauci, was demonstrated as important in pathogenicity and in carrot partial resistance to this fungus. As nothing is known about aldaulactone biosynthesis, bioinformatic analyses on a publicly available A. dauci genome data set that were reassembled, thanks to a transcriptome data set described here, allowed to identify 19 putative secondary metabolism clusters. We exploited phylogeny to pinpoint cluster 8 as a candidate in aldaulactone biosynthesis. This cluster contains AdPKS7 and AdPKS8, homologs with genes encoding a reducing and a non-reducing polyketide synthase. Clusters containing such a pair of PKS genes have been identified in the biosynthesis of resorcylic acid lactones or dihydroxyphenylacetic acid lactones. AdPKS7 and AdPKS8 gene expression patterns correlated with aldaulactone production in different experimental conditions. The present results highly suggest that both genes are responsible for aldaulactone biosynthesis.

Severe Inhibition of Long-Chain Acyl-CoA Enoylhydratase (EC 4.2.1.74) in a Newborn Foal Suffering From Atypical Myopathy.

In horses, congenital defects of energy production from long-chain fatty acids have not been described so far. In contrast, inhibition of fatty acid degradation caused by the toxins hypoglycin A and methylenecyclopropylglycine from various maple species are observed frequently. These non-proteinogenic aminoacids are passed on placentally to fetuses or with collostrum or milk to newborn foals. Nevertheless, newborn foals become very rarely symptomatic. Vertical transmission apparently is not sufficient to induce clinical disease without a particular genetic constellation being present. One of these rare cases was investigated here using samples from a mare and her foal. Intoxication by hypoglycin A and methylenecyclopropylglycine is also of interest to human pathology, because these toxins have caused fatal poisonings after consumption of certain fruits many times, especially in children. Maple toxins, their metabolites and some short-chain acyl compounds were quantified by ultrahigh-pressure liquid chromatography/tandem mass spectrometry. An comprehensive spectrum of long-chain acylcarnitines was prepared using electrospray ionization tandem mass spectrometry. Organic acids and acylglycines were determined by gas chromatography mass spectrometry. For evaluation, results of other horses poisoned by maple material as well as unaffected control animals were used. In the serum of the foal, hypoglycin A was detected at a low concentration only. Toxin metabolites reached <3.5% of the mean of a comparison group of horses suffering from atypical myopathy. The spectrum of acylcarnitines indicated enzyme inhibition in short-chain and medium-chain regions typical of acer poisoning, but the measured concentrations did not exceed those previously found in clinically healthy animals after maple consumption. The values were not sufficient to explain the clinical symptoms. In contrast, a remarkably strong enrichment of tetradecenoylcarnitine and hexadecenoylcarnitine was observed. This proves a blockade of the long-chain enoyl-CoA hydratase (EC 4.2.1.74). Vertical transfer of maple toxins to a newborn foal is sufficient for induction of clinical disease only if there is an additional specific reactivity to the active toxins. This was found here in an inhibition of long-chain enoyl-CoA hydratase. Isolated dysfunction of this enzyme has not yet been reported in any species. Further studies are necessary to prove a specific genetic defect.

Histopathology, toxin and secondary metabolites of Alternariaster helianthi in sunflower

Histopathology, toxin and secondary metabolites of Alternariaster helianthi in sunflower

In Vitro Metabolism of Azaspiracids 1-3 with a Hepatopancreatic Fraction from Blue Mussels (Mytilus edulis).

Azaspiracids (AZAs) are a group of biotoxins produced by the marine dinoflagellates Azadinium and Amphidoma spp. that can accumulate in shellfish and cause food poisoning in humans. Of the 60 AZAs identified, levels of AZA1, AZA2, and AZA3 are regulated in shellfish as a food safety measure based on occurrence and toxicity. Information about the metabolism of AZAs in shellfish is limited. Therefore, a fraction of blue mussel hepatopancreas was made to study the metabolism of AZA1-3 in vitro. A range of AZA metabolites were detected by liquid chromatography-high-resolution tandem mass spectrometry analysis, most notably the novel 22α-hydroxymethylAZAs AZA65 and AZA66, which were also detected in naturally contaminated mussels. These appear to be the first intermediates in the metabolic conversion of AZA1 and AZA2 to their corresponding 22α-carboxyAZAs (AZA17 and AZA19). α-Hydroxylation at C-23 was also a prominent metabolic pathway, producing AZA8, AZA12, and AZA5 as major metabolites of AZA1-3, respectively, and AZA67 and AZA68 as minor metabolites via double-hydroxylation of AZA1 and AZA2, but only low levels of 3β-hydroxylation were observed in this study. In vitro generation of algal toxin metabolites, such as AZA3, AZA5, AZA6, AZA8, AZA12, AZA17, AZA19, AZA65, and AZA66 that would otherwise have to be laboriously purified from shellfish, has the potential to be used for the production of standards for analytical and toxicological studies.

Profiling of Brevetoxin Metabolites Produced by Karenia brevis 165 Based on Liquid Chromatography-Mass Spectrometry.

In this study, Karenia brevis 165 (K. brevis 165), a Chinese strain, was used to research brevetoxin (BTX) metabolites. The sample pretreatment method for the enrichment of BTX metabolites in an algal culture medium was improved here. The method for screening and identifying intracellular and extracellular BTX metabolites was established based on liquid chromatography-time-of-flight mass spectrometry (LC-ToF-MS) and liquid chromatography triple quadrupole tandem mass spectrometry (LC-QqQ-MS/MS). The results show that the recovery rates for BTX toxins enriched by a hydrophilic–lipophilic balance (HLB) extraction column were higher than those with a C18 extraction column. This method was used to analyze the profiles of extracellular and intracellular BTX metabolites at different growth stages of K. brevis 165. This is the first time a Chinese strain of K. brevis has been reported that can produce toxic BTX metabolites. Five and eight kinds of BTX toxin metabolites were detected in the cell and culture media of K. brevis 165, respectively. Brevenal, a toxic BTX metabolite antagonist, was found for the first time in the culture media. The toxic BTX metabolites and brevenal in the K. brevis 165 cell and culture media were found to be fully proven in terms of the necessity of establishing a method for screening and identifying toxic BTX metabolites. The results found by qualitatively and quantitatively analyzing BTX metabolites produced by K. brevis 165 at different growth stages show that the total toxic BTX metabolite contents in single cells ranged between 6.78 and 21.53 pg/cell, and the total toxin concentration in culture media ranged between 10.27 and 449.11 μg/L. There were significant differences in the types and contents of toxic BTX metabolites with varying growth stages. Therefore, when harmful algal blooms occur, the accurate determination of BTX metabolite types and concentrations will be helpful to assess the ecological disaster risk in order to avoid hazards and provide appropriate disaster warnings.

Oral exposure to environmental cyanobacteria toxins: Implications for cancer risk

BackgroundAreca nut/betel quid (AN/BQ) chewing, a prevalent practice in parts of the Pacific and Asia, is an independent cause of cancers of the oral cavity and esophagus and may be linked to liver cancer. The mechanisms of AN/BQ-associated carcinogenesis are unclear. In a Guam population, we previously demonstrated that AN/BQ chewing alters the oral bacterial microbiome including in chewers with oral premalignant lesions. Enrichment of specific taxa, including Cyanobacteria, was observed. ObjectivesWe undertook an investigation to evaluate Areca catechu and/or Piper betle plants as potential sources of Cyanobacteria and cyanotoxins in AN/BQ chewers in Guam. MethodsWe evaluated bacterial 16S rRNA with Illumina MiSeq in 122 oral samples and 30 Areca catechu nut and Piper betle leaf samples. Cyanobacteria sequences were interrogated using the NCBI database to identify candidate species and their reference sequences were evaluated for secondary metabolite toxins using AntiSMASH 5.0. Selected toxins were measured by ELISA in extracts from 30 plant samples and in a subset of 25 saliva samples. ResultsCyanobacteria was the predominant taxa in Areca catechu and Piper betle plants, comprising 75% of sequences. Cyanobacteria was detected at low levels in oral samples but 90-fold higher in current AN/BQ chewers compared to former/never chewers (p = 0.001). Microcystin/nodularin was detected in saliva (15 of 25 samples) and Piper betle leaves (6 of 10 samples). Cylindrospermopsin was detected in all saliva and leaf samples and 7 of 10 nut/husks. Salivary cylindrospermopsin levels were significantly higher in current chewers of betel quid (i.e., crushed Areca catechu nut wrapped in Piper betle leaf) compared to those chewing Areca nut alone. Anabaenopeptin was detected in saliva (10 of 25 samples), all leaf samples, and 7 of 10 nut/husks. Salivary anabaenopeptin concentration was weakly, albeit significantly, correlated with oral Cyanobacteria relative abundance. DiscussionOur study demonstrates that Cyanobacteria can contaminate AN/BQ plants and expose chewers to potent hepatotoxins. With worldwide increases in climate-related overgrowth of Cyanobacteria, our findings have broad implications for cancer risk across populations.

STUDY OF AFLATOXINS IN CASHEW NUTS PRODUCED IN COTE DIVOIRE BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY (LCHP)

The objective of this study is to isolate, identify and quantify four types of aflatoxins noted AFB1, AFB2, AFG1, and AFG2 that can be found in cashews grown in Cote dIvoire. These carcinogenic mycotoxins (AF) are secondary metabolite toxins produced by Aspergillus molds in plant foods.This work involved eleven (11) samples of 500 g of cashew nuts from eleven (11) cities of Cote dIvoire for the 2018-2019 campaign. These cities are: Beoumi, Bondoukou, Dabakala, Daloa, Douekoue, Ferkessedougou, Korhogo, Mignignan, Odienne, Sinematiali, and Touba. The test were carried out by high performance liquid chromatography (HPLC) after extraction of the four (4) mycotoxins on an immunoaffinity column at a flow rate of 3 mL / minute. These aflatoxins were identified and quantified from the following pairs of Retention time (Rt) in minutes and Limit of Detection (LD) in µg / kg: (13.777 0.00143), (10.583 0.00136), (9.901 0.00151), and (8.184 0.00564) respectively for AFB1, AFB2, AFG1, and AFG2. Our results show that all eleven (11) samples from these eleven (11) different cities contain aflatoxin (AFB1, AFB2, AFG2 and AFG1) contents below the national standard (2 µg / kg), that of the CODEX Alimentarius (1.4 µg / kg) and that of the European Union (2 µg / kg) indicating that cashews produced in Cote dIvoire comply with international standards and their consumption does not pose any risk to human health caused by the studied aflatoxins.

A data-driven integrative platform for computational prediction of toxin biotransformation with a case study

Recently, biogenic toxins have received increasing attention owing to their high contamination levels in feed and food as well as in the environment. However, there is a lack of an integrative platform for seamless linking of data-driven computational methods with ‘wet’ experimental validations. To this end, we constructed a novel platform that integrates the technical aspects of toxin biotransformation methods. First, a biogenic toxin database termed ToxinDB (http://www.rxnfinder.org/toxindb/), containing multifaceted data on more than 4836 toxins, was built. Next, more than 8000 biotransformation reaction rules were extracted from over 300,000 biochemical reactions extracted from ~580,000 literature reports curated by more than 100 people over the past decade. Based on these reaction rules, a toxin biotransformation prediction model was constructed. Finally, the global chemical space of biogenic toxins was constructed, comprising ~550,000 toxins and putative toxin metabolites, of which 94.7% of the metabolites have not been previously reported. Additionally, we performed a case study to investigate citrinin metabolism in Trichoderma, and a novel metabolite was identified with the assistance of the biotransformation prediction tool of ToxinDB. This unique integrative platform will assist exploration of the ‘dark matter’ of a toxin’s metabolome and promote the discovery of detoxification enzymes.

Study on the Metabolic Effects of Repeated Consumption of Canned Ackee.

Ackee fruits (Blighia sapida), an important food source in some tropical countries, can be the cause of serious poisoning. Ackees contain hypoglycin A and methylenecyclopropylglycine. Experiments were undertaken by a volunteer to elucidate the metabolic details of poisoning. Rapid intestinal absorption of the toxins was followed by their slow degradation to methylenecyclopropylacetyl and methylenecyclopropylformyl conjugates. Impairment of the metabolism of branched chain amino acids and ß-oxidation of fatty acids was found. Reduced enzyme activities were observed for several days after ingestion. A defined dose of fruit material caused significantly higher concentrations of metabolites when consumed 24 h after a previous ingestion than when consumed only once. The accumulation of toxins, toxin metabolites, and products of the intermediate metabolism after repeated consumption may, at least partly, explain the high frequency of fatal cases observed during harvesting. No inhibition of enzymes that degrade long-chain acyl compounds was observed in the experiments.

Toxin Degradation by Rumen Microorganisms: A Review.

Animal feeds may contain exogenous compounds that can induce toxicity when ruminants ingest them. These toxins are secondary metabolites originating from various sources including plants, bacteria, algae and fungi. Animal feed toxins are responsible for various animal poisonings which negatively impact the livestock industry. Poisoning is more frequently reported in newly exposed, naïve ruminants while ‘experienced’ ruminants are observed to better tolerate toxin-contaminated feed. Ruminants can possess detoxification ability through rumen microorganisms with the rumen microbiome able to adapt to utilise toxic secondary metabolites. The ability of rumen microorganisms to metabolise these toxins has been used as a basis for the development of preventative probiotics to confer resistance against the poisoning to naïve ruminants. In this review, detoxification of various toxins, which include plant toxins, cyanobacteria toxins and plant-associated fungal mycotoxins, by rumen microorganisms is discussed. The review will include clinical studies of the animal poisoning caused by these toxins, the toxin mechanism of action, toxin degradation by rumen microorganisms, reported and hypothesised detoxification mechanisms and identified toxin metabolites with their toxicity compared to their parent toxin. This review highlights the commercial potential of rumen inoculum derived probiotics as viable means of improving ruminant health and production.

VenoMS-A Website for the Low Molecular Mass Compounds in Spider Venoms.

Spider venoms are highly complex mixtures. Numerous spider venom metabolites are uniquely found in spider venoms and are of interest concerning their potential use in pharmacology, agriculture, and cosmetics. A nontargeted ultra-high performance high-resolution electrospray tandem mass spectrometry (UHPLC-HR-ESI-MS/MS) approach offers a resource-saving way for the analysis of crude spider venom. However, the identification of known as well as the structure elucidation of unknown low molecular mass spider venom compounds based on their MS/MS spectra is challenging because (1) acylpolyamine toxins are exclusively found in spider and wasp venom, (2) reference MS/MS spectra are missing in established mass spectrometry databases, and (3) trivial names for the various toxin metabolites are used in an inconsistent way in literature. Therefore, we introduce the freely accessible MS website for low molecular mass spider venom metabolites, venoMS, containing structural information, MS/MS spectra, and links to related literature. Currently the database contains the structures of 409 acylpolyamine toxins, 36 free linear polyamines, and 81 additional spider venom metabolites. Implemented into this website is a fragment ion calculator (FRIOC) that allows us to predict fragment ions of linear polyamine derivatives. With three metabolites from the venom of the spider Agelenopsis aperta, it was demonstrated how the new website can support the structural elucidation of acylpolyamines using their MS/MS spectra.

Identifikasi Aspergillus flavus pada Kue Pia yang Di Jual Di Dusun Warurejo Kabupaten Pasuruan

Warurejo village that located in Pasuruan district is the center of pia cake business expecially green beans pia. Pia cake is a cake that can’t durable cause by microorganism that come from the less clear and wraping in production. It makes contamination of mycotoxins from spesies of the fungus is Aspergillus flavus . Aspergillus flavus can produce a toxin metabolite compund that is aflatoxin. If aflatoxin is continuous consumtion will cause liver cancer. This research aim to determine presence of Aspergillus flavus on pia cake that produce by center of pia cake business in Warurejo village. It will be research with storage time of 0, 5 and 10 days. This study is descriptive, the number of sample taken randomly as many as 30 samples from 33 stores. Research this is obtained by macroscopically and microscopically. In macroscopically colonis that, look yellowish greeen and on the bottom is yellow to brown while microscopically rounded vesicles and conidi are round. 73% of Aspergillus flavus has positive contaminated fungus in 0 days and 5 days. 97% positive was Aspergillus flavus with 10 days storage time. In day 10 found more Aspergillus flavus fungus that on days 0 and 5 days.

Chemical assessment of mycotoxin contaminants and veterinary residues in Costa Rican animal feed

BackgroundWhen present in animal feedstuff, mycotoxins contaminants and antibiotic residues can have negative implications for animal production and Public Health, including the transmission of carcinogenic compounds and the selection of antibiotic resistant bacteria, respectively. So far there are no available methods in which both mycotoxins and antibiotic residues are analyzed using a parallel extraction approach. To address this issue, we developed a LC/MS methodology with high sensitivity (0.005 to 6.42 and 24.55 to 132.73 μg kg− 1 for mycotoxins and antimicrobials) and specificity (unique target ion mass/charge) that allows the detection of 26 mycotoxins and 23 antibiotic residues in animal feedstuff and validated it through the determination of these analytes in 294 animal feed and feed ingredient samples in the framework of a country-wide surveillance program. Two hundred and five of these samples were analyzed for mycotoxins and 89 for antibiotics.FindingsFumonisin was the most frequently toxin found, with FB1 and FB2 presenting prevalences of 50 and 52% and maximum concentrations of 14,927.61 and 8646.67 μg kg− 1, respectively. Other toxins, including diacetoxyscirpenol n = 4/101 (3.96%), fusarenon-X n = 2/101 (1.98%), citrinin n = 2 (1.98%), and patulin n = 1 (0.99%) were rarely found. Toxicologically relevant concentrations of toxin metabolites, such as HT-2 (6.38–485.49 μg kg− 1) and 3−/15-acetoxydeoxynivalenol (877.89–3236.56/5.44–1685.3 μg kg− 1), were also found. Few samples exceeded threshold mycotoxin concentrations defined in current EU guidelines. Dairy cattle and swine feeds included the higher number of samples exceeding guideline values (n = 6 and n = 5, respectively). From the total of samples analysed for antibiotics, 7.7% (n = 7/89) were classified as medicated for poultry and pigs. Unexpectedly, 57% of these medicated samples contained no detectable antibiotics (n = 4/7). The remaining 43% of the samples (n = 3/7) presented inconsistencies regarding the concentration of analytes declared on the labels or the antibiotics found. Likewise 74.6% (n = 50/67) of the non-medicated feed samples analyzed had antibiotic residues. Additionally, we analyzed commercial monensin standards for purity and evaluate batch-to-batch flushing feed industry practices.ConclusionsHerein we report the results for a year-wide analysis for mycotoxins and antibiotics in feed samples. Mycotoxins, several metabolites, and the occurrence of these emerging contaminants were evaluated and antibiotic residues in non-medicated feed samples were found using a targeted MS-based LC approach. This validated multi-analyte method is expected to facilitate the monitoring and surveillance of contaminants, from natural and anthropogenic origin, in animal feed.

Single-cell electro-phenotyping for rapid assessment of Clostridium difficile heterogeneity under vancomycin treatment at sub-MIC (minimum inhibitory concentration) levels

Current methods for measurement of antibiotic susceptibility of pathogenic bacteria are highly reliant on microbial culture, which is time consuming (requires > 16 h), especially at near minimum inhibitory concentration (MIC) levels of the antibiotic. We present the use of single-cell electrophysiology-based microbiological analysis for rapid phenotypic identification of antibiotic susceptibility at near-MIC levels, without the need for microbial culture. Clostridium difficile (C. difficile) is the single most common cause of antibiotic-induced enteric infection and disease recurrence is common after antibiotic treatments to suppress the pathogen. Herein, we show that de-activation of C. difficile after MIC-level vancomycin treatment, as validated by microbiological growth assays, can be ascertained rapidly by measuring alterations to the microbial cytoplasmic conductivity that is gauged by the level of positive dielectrophoresis (pDEP) and the frequency spectra for co-field electro-rotation (ROT). Furthermore, this single-cell electrophysiology technique can rapidly identify and quantify the live C. difficile subpopulation after vancomycin treatment at sub-MIC levels, whereas methods based on measurement of the secreted metabolite toxin or the microbiological growth rate can identify this persistent C. difficile subpopulation only after 24 h of microbial culture, without any ability to quantify the subpopulation. The application of multiplexed versions of this technique is envisioned for antibiotic susceptibility screening.

Metabolic alterations in the bone tissues of aged osteoporotic mice

Age-related osteoporosis is characterized by reduced bone mineralization and reduced bone strength, which increases the risk of fractures. We examined metabolic changes associated with age-related bone loss by profiling lipids and polar metabolites in tibia and femur bone tissues from young (5 months old) and old (28 months old) male C57BL/6J mice using ultra-performance liquid chromatography quadrupole-time-of-flight mass spectrometry. Partial least-squares discriminant analysis showed clear differences in metabolite levels in bone tissues of young and old mice. We identified 93 lipid species, including free fatty acids, sphingolipids, phospholipids, and glycerolipids, that were significantly altered in bone tissues of old mice. In addition, the expression of 26 polar metabolites differed significantly in bone tissues of old mice and young mice. Specifically, uremic toxin metabolite levels (p-cresyl sulfate, hippuric acid, and indoxylsulfate) were higher in bone tissues of old mice than in young mice. The increase in p-cresyl sulfate, hippuric acid, and indoxylsulfate levels were determined using targeted analysis of plasma polar extracts to determine whether these metabolites could serve as potential osteoporosis biomarkers. This study demonstrates that LC-MS-based global profiling of lipid and polar metabolites can elucidate metabolic changes that occur during age-related bone loss and identify potential biomarkers of osteoporosis.

Peroxiredons: Tryparedoxin Peroxidase from Leishmania major

Background: Leishmania parasites causative agents of leishmaniasis are aerotolerant or microaerophilic and consume oxygen to some extent. Leishmania parasites produce hydrogen peroxide as a metabolism product in certain circumstances and are naturally exposed to the toxin metabolites such as Reactive Oxygen Species generated by macrophages. To stand oxidative stress, trypanosomatids evolved a unique dithiol trypanothione pathway which consists of a cascade of low molecular weight thiol specific oxidoreductases acting in the order of trypanothione reductase, trypanothione, tryparedoxin, and tryparedoxin peroxidase to detoxify peroxides. Tryparedoxin peroxidases belong to a widespread family of peroxiredoxins. Keywords: Hydrogen peroxide, kinetoplastida, Leishmania, peroxiredoxin, ROS, tryparedoxin peroxidase.

Tracing the metabolism of HT-2 toxin and T-2 toxin in barley by isotope-assisted untargeted screening and quantitative LC-HRMS analysis.

An extensive study of the metabolism of the type A trichothecene mycotoxins HT-2 toxin and T-2 toxin in barley using liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS) is reported. A recently developed untargeted approach based on stable isotopic labelling, LC-Orbitrap-MS analysis with fast polarity switching and data processing by MetExtract software was combined with targeted LC-Q-TOF-MS(/MS) analysis for metabolite structure elucidation and quantification. In total, 9 HT-2 toxin and 13 T-2 toxin metabolites plus tentative isomers were detected, which were successfully annotated by calculation of elemental formulas and further LC-HRMS/MS measurements as well as partly identified with authentic standards. As a result, glucosylated forms of the toxins, malonylglucosides, and acetyl and feruloyl conjugates were elucidated. Additionally, time courses of metabolite formation and mass balances were established. For absolute quantification of those compounds for which standards were available, the method was validated by determining apparent recovery, signal suppression, or enhancement and extraction recovery. Most importantly, T-2 toxin was rapidly metabolised to HT-2 toxin and for both parent toxins HT-2 toxin-3-O-β-glucoside was identified (confirmed by authentic standard) as the main metabolite, which reached its maximum already 1 day after toxin treatment.Graphical Isotope-assisted untargeted screening of HT-2 toxin and T-2 toxin metabolites in barleyElectronic supplementary materialThe online version of this article (doi:10.1007/s00216-015-8975-9) contains supplementary material, which is available to authorized users.