RESUMO
The use of medicinal herbs has increased significantly. However, the presence of pesticide residues and mycotoxins in medicinal herbs has generated constant discussion and concern among regulatory agencies. Developing and validating an analytical method for determining pesticides and mycotoxins in medicinal plants is challenging due to the naturally occurring substances in these plants. The purpose of this work was to develop and to optimize a sensitive, accurate, precise, effective QuEChERS method for simultaneous determination of over 160 pesticide and mycotoxin residues in complex medicinal plant matrices using LC-TQ-MS/MS. A comprehensive comparison of clean-up procedures and other parameters was conducted to achieve this goal. The validation procedure was performed according to SANTE 11312/2021. More polar analytes, such as acephate, methamidophos and omethoate, presented a higher negative matrix effect in both Melissa officinalis L. and Malva sylvestris L. However, other molecules, such as spirodiclofen, showed a 24% signal enhancement in M. officinalis and a 46% signal suppression in M. sylvestris, indicating that a representative matrix-matched calibration would lead to inaccurate quantification of the analyte. Accuracy and precision were satisfactory according to SANTE 11312/2021 for 157 pesticide residues and mycotoxins in M. officinalis and for 152 molecules in M. sylvestris. LOQs at 10 µg kg-1 were achieved for 117 pesticides in M. officinalis and 99 pesticides in M. sylvestris. Among the mycotoxins, all four aflatoxins (B1, B2, G1 and G2) presented LOQs of 5 µg kg-1, and ochratoxin A had an LOQ of 10 µg kg-1 in M. officinalis. The same LOQ values were shown for these mycotoxins in M. sylvestris, except for aflatoxin B2 and ochratoxin A, which had LOQs of 20 µg kg-1. Moreover, in Southern Brazil, there has been no previous study on mycotoxin and pesticide contamination in medicinal herbs. Therefore, the application of this method was assessed through the analysis of forty-two real samples. Imidacloprid was found in M. officinalis, and methyl pirimiphos was found in M. sylvestris. The proposed method not only serves as a helpful tool for routine monitoring but also offers a basis for further research on risk assessment and control in food safety.
Assuntos
Micotoxinas , Plantas Medicinais , Espectrometria de Massas em Tandem , Espectrometria de Massas em Tandem/métodos , Micotoxinas/análise , Micotoxinas/isolamento & purificação , Plantas Medicinais/química , Brasil , Cromatografia Líquida/métodos , Limite de Detecção , Resíduos de Praguicidas/análise , Praguicidas/análise , Reprodutibilidade dos TestesRESUMO
Starches-rich and protein-rich cereal samples commonly need tedious sample preparation steps before instrumental analysis. This study developed a miniaturized centrifugal integrated cold-induced phase separation (CIPS) method for convenient sample preparation. A small-sized centrifuge tube (2 mL) and a low-temperature centrifuge, both of which are easily accessible, make up the basic components of the system. Unlike conventional sample preparation methods that need a step-by-step extraction, enrichment, purification, and centrifugation, this centrifugal integrated CIPS method can be performed by a one-step combination protocol under a low-temperature centrifuge. As a proof-of-concept study, satisfactory recoveries and enrichment factors were demonstrated for the extraction of fumonisins and ochratoxins from cereals. A sensitive and selective quantification method was yielded by combining LC-HRMS using tSIM acquisition mode, with good linearity (R2 > 0.998), accuracy (82.9-106.5%), and precision (<13.4%). This strategy is convenient, low-cost, repeatable, and easy to semi-automate, further expanding the extraction potential for other acidic mycotoxins.
Assuntos
Grão Comestível , Contaminação de Alimentos , Fumonisinas , Grão Comestível/química , Contaminação de Alimentos/análise , Fumonisinas/análise , Fumonisinas/isolamento & purificação , Centrifugação , Estudo de Prova de Conceito , Micotoxinas/isolamento & purificação , Micotoxinas/análise , Micotoxinas/química , Cromatografia Líquida de Alta Pressão , Separação de FasesRESUMO
Biscogniauxia rosacearum, recognized for the first time as a pathogen involved in grapevine trunk diseases in Paveh (west of Iran) vineyards, produced meso-2,3-butanediol (1) as the only phytotoxin. Nectriapyrone (2), (3R)-5-methylmellein (3), (3R)-5-methyl-6-methoxymellein (4), and tyrosol (5) were instead produced as phytotoxins from a strain of the same fungus isolated from oak trees in Zagros forests of Gilan-e Gharb, Kermanshah Province. They were identified comparing their 1H and 13C NMR, ESIMS, and specific optical rotation data with those already reported in the literature. The phytotoxicity of metabolites (1-5) was estimated by leaf puncture assay on Quercus ilex L. and Hedera helix L., and by leaf absorption assay on grapevine (Vitis vinifera L.) at a concentration of 5 × 10-3 and 10-3 M. Tested on grapevine, meso-2,3-butanediol (1) and (3R)-5-methyl-6-methoxymellein (4) resulted to be the most phytotoxic compounds. On Q. ilex, nectriapyrone (2) and tyrosol (5) showed severe necrosis at the highest concentration while none of the compounds (1-5) was active on H. helix. Furthermore, the phytotoxicity of compounds 3 and 4 was also compared with that of some related natural melleins to perform a structure-activity relationship (SAR) study. The results of this study were also discussed.
Assuntos
Ascomicetos/patogenicidade , Micotoxinas/toxicidade , Doenças das Plantas/microbiologia , Ascomicetos/metabolismo , Carvão Vegetal , Irã (Geográfico) , Micotoxinas/química , Micotoxinas/isolamento & purificação , Quercus/microbiologia , Relação Estrutura-Atividade , Vitis/microbiologiaRESUMO
Environmental pollutants, such as mycotoxins, pesticides, and pharmaceuticals, are a group of contaminates that occur naturally, while others are produced from anthropogenic sources. With increased research on the adverse ecological and human health effects of these pollutants, there is an increasing need to regularly monitor their levels in food and the environment in order to ensure food safety and public health. The application of magnetic nanomaterials in the analyses of these pollutants could be promising and offers numerous advantages relative to conventional techniques. Due to their ability for the selective adsorption, and ease of separation as a result of magnetic susceptibility, surface modification, stability, cost-effectiveness, availability, and biodegradability, these unique magnetic nanomaterials exhibit great achievement in the improvement of the extraction of different analytes in food. On the other hand, conventional methods involve longer extraction procedures and utilize large quantities of environmentally unfriendly organic solvents. This review centers its attention on current applications of magnetic nanomaterials and their modifications in the extraction of pollutants in food commodities.
Assuntos
Imãs/química , Micotoxinas/isolamento & purificação , Nanoestruturas/química , Praguicidas/isolamento & purificação , Preparações Farmacêuticas/isolamento & purificação , Extração em Fase Sólida/métodos , Animais , Poluentes Ambientais/análise , Poluentes Ambientais/isolamento & purificação , Contaminação de Alimentos/análise , Inocuidade dos Alimentos , Humanos , Micotoxinas/análise , Praguicidas/análise , Preparações Farmacêuticas/análise , Extração em Fase Sólida/instrumentaçãoRESUMO
Magnetic covalent organic framework nanocomposite denoted as Fe3O4@TAPB-Tp with core-shell structure was fabricated via a simple template-mediated precipitation polymerization method at mild conditions. The polyimine network shell was created through the polymerization of 1,3,5-tris(4-aminophenyl)-benzene (TAPB) and 1,3,5-triformyl-phloroglucinol (Tp) in tetrahydrofuran (THF) by the Schiff-base reaction. Featuring with large specific surface area (163.19 m2 g-1), good solution dispersibility, and high stability, the obtained Fe3O4@TAPB-Tp exhibited high adsorption capacities and fast adsorption for zearalenone and its derivatives (ZEAs). The adsorption isotherms showed multilayer adsorption dominated at low concentration and monolayer adsorption at high concentration between the interface of ZEAs and Fe3O4@TAPB-Tp. With the Fe3O4@TAPB-Tp as sorbent, a magnetic solid-phase extraction-ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was established for simultaneous adsorption and detection of five ZEAs in complex samples. The proposed method displayed favorable linearity, low limits of detection (0.003 ~ 0.018 µg kg-1), and good repeatability (2.37~10.4%). The developed method has been applied for real sample analysis, with recoveries of 81.27~90.26%. These results showed that Fe3O4@TAPB-Tp has a good application potential for the adsorption of ZEAs in food samples. Magnetic covalent organic framework nanocomposite (Fe3O4@TAPB-Tp) were quickly fabricated at mild conditions and used as effective adsorbent for magnetic solid-phase extraction of zearalenone and its derivatives (ZEAs) from food samples prior to ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analysis.
Assuntos
Nanopartículas de Magnetita/química , Estruturas Metalorgânicas/química , Micotoxinas/análise , Zearalenona/análise , Adsorção , Animais , Derivados de Benzeno/química , Cromatografia Líquida de Alta Pressão , Ovos/análise , Contaminação de Alimentos/análise , Limite de Detecção , Fenômenos Magnéticos , Leite/química , Micotoxinas/química , Micotoxinas/isolamento & purificação , Nanocompostos/química , Floroglucinol/análogos & derivados , Floroglucinol/química , Polimerização , Extração em Fase Sólida/métodos , Espectrometria de Massas em Tandem , Zea mays , Zearalenona/análogos & derivados , Zearalenona/isolamento & purificaçãoRESUMO
Edible insects are widely consumed in Africa, Asia, Oceania and Latin America, but less commonly so in Western countries. Since the turn of the millennium, however, entomophagy has aroused growing interest worldwide in response to the increasing scarcity of food resources. In fact, edible insects can be a source of high-quality protein, and also of fat, energy, minerals and vitamins. However, the lack of regulatory guidelines for microbiologically or chemically hazardous agents potentially present in these new foods (e.g., mycotoxins) may make their consumption unsafe. In this work, we developed an environmentally friendly analytical method using natural deep eutectic solvents (NADES or natural DES) in combination with ultra-high performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS) for the simultaneous determination of six mycotoxins of great concern owing to their toxic effects on humans and animals (namely, fumonisin B1, fumonisin B2, T-2 toxin, HT-2 toxin, ochratoxin A and mycophenolic acid) in insect-based food products. The target mycotoxins were co-extracted from cricket flour by using the optimum DES composition (namely, a mixture of choline chloride and urea, in a 1:2 mole ratio, containing 15% water which resulted in the highest extraction recoveries for all toxins). An experimental design method (Fractional Factorial Design (FFD) was used to examine the influence of the operational variables DES volume and water content, amount of sample, extraction time and extraction temperature on the extraction efficiency for each mycotoxin. Under optimum conditions, extraction recoveries were close to 100% except for fumonisin B2 (70%) and T-2 toxin (50%), with relative standard deviations (RSDs) below 13% in all cases. The proposed NADES-UHPLC-MS/MS method was validated in accordance with the European Commission 2002/657/EC and 2006/401/EC decisions, and used to determine the target compounds in cricket flour, silkworm pupae powder and black cricket powder.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Insetos Comestíveis/química , Micotoxinas/isolamento & purificação , Solventes/química , África , Animais , Humanos , Espectrometria de Massas em Tandem/métodosRESUMO
The pressurized liquid extraction (PLE) technique was used, for the first time, to obtain protein extracts with antioxidant activity from side streams (muscle, heads, viscera, skin, and tailfins) of gilthead sea bream (Sparus aurata) in order to give added value to these underutilized matrices. Extraction conditions previously optimized for sea bass (Dicentrarchus labrax) side streams were applied. Protein recovery percentages were 22% (muscle), 33% (heads), 78% (viscera), 24% (skin), and 26% (tailfins), which represented an increase of 1.2-4.5-fold compared to control samples (extraction by stirring). The SDS-PAGE profiles revealed that PLE-assisted extraction influenced protein molecular weight distribution of the obtained extracts. PLE conditions also allowed increasing the antioxidant capacity measured by both Trolox equivalent antioxidant capacity (TEAC; 1.3-2.4 fold) and oxygen radical absorbance capacity (ORAC; 1.9-6.4) assays for all fish extracts. Inductively coupled plasma mass spectrometry (ICP-MS) and high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (LC-ESI-qTOF-MS) were used to investigate the presence of toxic metals and mycotoxins in sea bream side streams. The levels of As, Hg, Cd, and Pb were below those established by authorities for fish muscle for human consumption (except for Cd in viscera samples). Through a nontargeted screening approach, no mycotoxins or related metabolites were detected for all sea bream side streams. This study contributes to the research on the valorization of fish processing side streams using environmentally friendly technology.
Assuntos
Antioxidantes/farmacologia , Proteínas de Peixes/farmacologia , Extração Líquido-Líquido , Dourada/metabolismo , Animais , Antioxidantes/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Eletroforese em Gel de Poliacrilamida , Proteínas de Peixes/isolamento & purificação , Manipulação de Alimentos , Metais/isolamento & purificação , Peso Molecular , Micotoxinas/isolamento & purificação , Capacidade de Absorbância de Radicais de Oxigênio , Pressão , Espectrometria de Massas por Ionização por Electrospray , ResíduosRESUMO
Ustilaginoidins, a group of bis-naphtho-γ-pyrones, are one of the major mycotoxins produced by Ustilaginoidea virens. This group of bis-naphtho-γ-pyrone mycotoxins has been demonstrated to have antibacterial and immunological inhibitory activities and strong cytotoxicity to human oral epidermoid carcinoma. However, little is yet known about the toxicity of ustilaginoidins to animals or toxicity mechanisms. In this study, toxicity assays to zebrafish larvae show that ustilaginoidin D is highly toxic to zebrafish with an LC50 of â¼7.50 µM. Ustilaginoidin D causes an obvious yolk sac absorption delay and liver damage in zebrafish, which is indicated by liver atrophy and the increased alanine and aspartate transaminase activities. Interestingly, different doses of ustilaginoidin D can alter zebrafish movement behavior in a distinct manner. Transcriptome analyses show that global gene expression profiling in zebrafish is significantly changed in response to ustilaginoidin D exposure. KEGG pathway analyses reveal that differentially expressed genes are enriched in the pathways related to lipid metabolism and hyperbilirubinemia, which are indicators of severe liver injury. Consistently, the expression of the marker genes for hepatotoxic responses is significantly induced by ustilaginoidin D. The findings indicate that ustilaginoidin D induces lipid metabolism disorders and hepatotoxicity in zebrafish larvae and poses a potential risk to food safety.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Hypocreales , Larva/efeitos dos fármacos , Larva/metabolismo , Locomoção/efeitos dos fármacos , Micotoxinas/toxicidade , Animais , Doença Hepática Induzida por Substâncias e Drogas/patologia , Relação Dose-Resposta a Droga , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/fisiologia , Locomoção/fisiologia , Micotoxinas/isolamento & purificação , Peixe-ZebraRESUMO
Cordyceps fumosorosea, an insect pathogenic fungus, produces different toxins/secondary metabolites which can act as pest control agents. This study reports the extraction and characterization of crude mycelial extracts of C. fumosorosea isolate SP502 along with their bio-efficacy against Bemisia tabaci and Aphis craccivora. Fourier transform infrared spectroscopy, liquid chromatography, mass spectrometery and nuclear magnetic resonance analysis of C. fumosorosea isolate SP502 extracts showed the presence of five major compounds-Trichodermin, 5-Methylmellein, Brevianamide F, Enniatin and Beauvericin-which all may potentially be involved in insecticidal activity. The HPLC analysis of C. fumosorosea mycelial extracts and Beauvericin standard showed similar chromatographic peaks, with the content of Beauvericin in the crude toxin being calculated as 0.66 mg/ml. The median lethal concentrations of C. fumosorosea mycelial extracts towards first, second, third and fourth instar nymphs of A. craccivora were 46.35, 54.55, 68.94, and 81.92 µg/mL, respectively. The median lethal concentrations of C. fumosorosea mycelial extracts towards first, second, third and fourth instar nymphs of B. tabaci were 62.67, 72.84, 77.40, and 94.40 µg/mL, respectively. Our results demonstrate that bioactive compounds produced by C. fumosorosea isolate SP502 have insecticidal properties and could, therefore, be developed into biopesticides for the management of B. tabaci and A. craccivora.
Assuntos
Afídeos/efeitos dos fármacos , Agentes de Controle Biológico/farmacologia , Cordyceps/metabolismo , Hemípteros/efeitos dos fármacos , Micotoxinas/farmacologia , Animais , Afídeos/crescimento & desenvolvimento , Agentes de Controle Biológico/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Cordyceps/patogenicidade , Hemípteros/crescimento & desenvolvimento , Espectrometria de Massas , Micotoxinas/isolamento & purificação , Metabolismo Secundário , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
Liver samples from finisher pigs were collected at the slaughterhouses for the analysis of zearalenone (ZEA), alfa-/beta-zearalenone (α-ZE, ß-ZE), zearalanone (ZA), alfa-/beta-ZA (α-ZA, ß-ZA), aflatoxin B1 (AFB1) and aflatoxin M1, fumonisin B1 (FB1), ochratoxin A (OTA) and ochratoxin B, deoxynivalenol and deepoxi-deoxynivalenol (DOM-1). For the analysis liquid chromatography-triple quadrupole coupled with mass spectrometry was applied. Liver samples with detected FB1 were further histopathologically evaluated after hematoxylin and eosin staining. Various levels of liver mycotoxins were detected in all farms. Pig livers with 2.91-8.30 µg/kg of FB1 were detected in three farms, estimate of 850-2400 µg/kg of FB1 intake, whereas 0.54 µg/kg of OTA was detected in one farm, estimate of 75 µg/kg of OTA intake. Moreover, pig livers with 0.30 µg/kg of ZEA, 1.87 µg/kg of α-ZE, and 0.63 µg/kg of ß-ZE were detected in one farm, estimate with of 300 µg/kg of ZEA intake. The histopathological analysis revealed that the lesions' grading and necrosis grading were analogously increased when FB1 concentration increased from 2.91 to 4.36-8.30 µg/kg. The severity of megalocytosis was analogously increased with FB1 detection levels and particularly in levels of 4.36-8.3 µg/kg. However, the increased FB1 detection levels did not show analogous behavior with the severity of hepatic cell vacuolization. Results showed that FB1 remained the most critical risk factor in the Greek pig industry, whereas ZEA and AFB1 were also prevalent. The OTA contamination in pig farms raised a high risk for animal and human health.
Assuntos
Exposição Ambiental/análise , Fumonisinas/isolamento & purificação , Micoses/veterinária , Micotoxinas/isolamento & purificação , Doenças dos Suínos/microbiologia , Matadouros , Animais , Biomarcadores/análise , Cromatografia Líquida , Exposição Ambiental/efeitos adversos , Fígado/microbiologia , Espectrometria de Massas , SuínosRESUMO
Research on the ability of lactic acid bacteria (LAB) to bind aflatoxin B1 (AFB1) has mostly been focusing on lactobacilli and bifidobacteria. In this study, the AFB1 binding capacities of 20 Enterococcus strains belonging to E. casseliflavus, E. faecalis, E. faecium, E. hirae, E. lactis, and E. mundtii, 24 Pediococcus strains belonging to species P. acidilactici, P. lolii, P. pentosaceus, and P. stilesii, one strain of Lactococcus formosensis and L.garviae, and 3 strains of Weissella soli were investigated in MRS broth at 37 °C at 0.2 µg/mL mycotoxin concentration. According to our results, among non-lactobacilli LAB, the genera with the best AFB1 binding abilities were genus Pediococcus, with a maximum binding percentage of 7.6% by P. acidilactici OR83, followed by genus Lactococcus. For AFB1 bio-detoxification purposes, beside lactobacilli, pediococci can also be chosen, but it is important to select a strain with better binding properties than the average value of its genus. Five Pediococcus strains have been selected to compare their sterigmatocystin (ST) binding abilities to AFB1 binding, and a 2-3-fold difference was obtained similar to previous findings for lactobacilli. The best strain was P. acidilactici OR83 with 18% ST binding capacity. This is the first report on ST binding capabilities of non-Lactobacillus LAB strains.
Assuntos
Aflatoxina B1/metabolismo , Animais de Zoológico/metabolismo , Animais de Zoológico/microbiologia , Lactobacillales/metabolismo , Lactobacillus , Esterigmatocistina/metabolismo , Aflatoxina B1/genética , Aflatoxina B1/isolamento & purificação , Animais , Cromatografia Líquida de Alta Pressão/métodos , Lactobacillales/genética , Lactobacillales/isolamento & purificação , Micotoxinas/genética , Micotoxinas/isolamento & purificação , Micotoxinas/metabolismo , Ligação Proteica/fisiologia , Esterigmatocistina/isolamento & purificaçãoRESUMO
Tricholoma ustale, a poisonous member of the Tricholomataceae family, causes gastrointestinal symptoms such as diarrhea and vomiting. In Japan, 86 cases (affecting a total of 347 patients) of poisoning with Tricholoma ustale have been reported between 1989 and 2010. Ustalic acid is one of the primary toxic components in Tricholoma ustale. In the present study, the quantitative analysis of the ustalic acid content in mushroom and food samples was conducted by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Mushroom and food samples were extracted using methanol containing 0.5% formic acid and 50% aqueous methanol, respectively. Purification using SAX solid-phase extraction (SPE) was conducted prior to LC-MS/MS analysis, which was performed in the ESI negative mode using a C18 column. The method developed for the LC-MS/MS analysis of ustalic acid was extremely sensitive. The limits of quantitation calculated at a signal-to-noise ratio of 10 were 10ng/g (shiitake mushroom) and 0.40ng/g (miso soup). The accuracies of quantitation in the shiitake mushroom and miso soup samples ranged from 99.8%-105% and 98.8%-102%, respectively. This method was applied to leftover mushroom samples from a food poisoning case; here, ustalic acid was detected at 0.57, 3.7µg/g. This analytical method using LC-MS/MS could be useful in food poisoning cases involving mushrooms. This is the first report in which the ustalic acid content was determined using the leftovers of a food poisoning case.
Assuntos
Agaricales/química , Análise de Alimentos , Intoxicação Alimentar por Cogumelos/diagnóstico , Micotoxinas/isolamento & purificação , Cromatografia Líquida , Humanos , Masculino , Extração em Fase Sólida , Espectrometria de Massas em TandemRESUMO
Hsp90 is a promising drug target for cancer therapy. However, toxicity and moderate effect are limitations of current inhibitors owing to broad protein degradation. The fungal mycotoxin penisuloxazin A (PNSA) belongs to a new epipolythiodiketopiperazines (ETPs) possessing a rare 3H-spiro[benzofuran-2,2'-piperazine] ring system. PNSA bound to cysteine residues C572/C598 of CT-Hsp90 with disulfide bonds and inhibits Hsp90 activity, resulting in apoptosis and growth inhibition of HCT116 cells in vitro and in vivo. We identified that analogues PEN-A and HDN-1 bound to C572/C597 and C572 of CT-Hsp90α respectively, with binding pattern very similar to PNSA. These ETPs exhibited different effects on ATPase activity, dimerization formation and selectivity on client protein of Hsp90, indicating client recognition of Hsp90 can be exactly regulated by different sites of Hsp90. Our findings not only offer new chemotypes for anticancer drug development, but also help to better understand biological function of Hsp90 for exploring inhibitor with some client protein bias.
Assuntos
Produtos Biológicos/metabolismo , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Proteínas de Choque Térmico HSP90/metabolismo , Micotoxinas/metabolismo , Células A549 , Animais , Sítios de Ligação/efeitos dos fármacos , Sítios de Ligação/fisiologia , Produtos Biológicos/isolamento & purificação , Produtos Biológicos/farmacologia , Células HCT116 , Células HL-60 , Células HeLa , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Micotoxinas/isolamento & purificação , Micotoxinas/farmacologia , Venenos/isolamento & purificação , Venenos/metabolismo , Venenos/farmacologia , Estrutura Secundária de Proteína , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Mycotoxin management in agriculture is an essential challenge for maintaining the health of both animals and humans. Choosing the right adsorbent is still a question for many breeders and an important criterion for feed manufacturers. New adsorbents are still being sought. Graphene oxide is a promising material in the field of nanotechnology, which excels in its adsorption properties. Presented in vitro study investigates graphene oxide for the binding of mycotoxins from crushed wheat. The results show that graphene oxide has an adsorption capacity for aflatoxin 0.045 mg/g, zearalenone 0.53 mg/g and deoxynivalenol 1.69 mg/g at 37° C. In vitro simulation of crushed wheat digestion showed rapid adsorption during the gastric phase. Of the minerals, Mg, Cu and Zn were the most adsorbed. The applied dose of graphene oxide of 10 mg/g caused only a slight inhibition of the digestive enzymes α-amylase and trypsin compared to pepsin and gastric lipase. In vitro results indicated the suitability of graphene oxide in the adsorption of the aflatoxin, zearalenone and deoxynivalenol.
Assuntos
Grafite/química , Micotoxinas/isolamento & purificação , Adsorção , Aflatoxina B1/isolamento & purificação , Aflatoxina B1/toxicidade , Ração Animal/análise , Ração Animal/toxicidade , Animais , Digestão , Contaminação de Alimentos/análise , Contaminação de Alimentos/prevenção & controle , Absorção Gastrointestinal , Humanos , Técnicas In Vitro , Micotoxinas/toxicidade , Nanoestruturas/química , Tricotecenos/isolamento & purificação , Tricotecenos/toxicidade , Triticum/química , Triticum/toxicidade , Zearalenona/isolamento & purificação , Zearalenona/toxicidadeRESUMO
A new type of immunomagnetic bead based on the metal-organic framework materials (MOFs) and the magnetic core (Fe3O4) was prepared for analysis of the mycotoxins in food samples. The MOF conjugated with the monoclonal antibodies (McAbs) was coated on the surface of Fe3O4 beads by reversed-phase microemulsion method, which could purify deoxynivalenol (DON), zearalenone (ZEN), T-2, and HT-2 mycotoxins at the same time. The composite (Fe3O4@AMP&ZnCl2@McAbs) was characterized by Transmission electron microscopy (TEM), Fourier-transform infrared spectroscopy (FTIR), and Energy dispersive spectroscopy (EDS) mapping. The results showed that the synthesis of the composite was successful. The maximum toxin adsorption capacity per 100 mg of composite was DON 688.26 ng, ZEN 864.98 ng, and T-2/HT-2 2801.80 ng, and in adding recovery experiment, the recovery of four mycotoxins decreased slightly with the increase of usage times, but still maintained a high adsorption rate and stability. For effectiveness comparison and evaluation, the composite and commercial DZTMS-PREP immune affinity column were used to treat three samples of corn, wheat, and oat flour, and the purification effect of the two pretreatment methods on the four toxins was similar.
Assuntos
Contaminação de Alimentos/análise , Separação Imunomagnética/métodos , Estruturas Metalorgânicas/química , Micotoxinas , Adsorção , Anticorpos Monoclonais/química , Anticorpos Monoclonais/metabolismo , Grão Comestível/química , Limite de Detecção , Modelos Lineares , Nanopartículas de Magnetita/química , Micotoxinas/análise , Micotoxinas/isolamento & purificação , Micotoxinas/metabolismoRESUMO
This study describes a smart analysis platform capable of quantitative measurements using a multiplex lateral flow strip. Using the multi-mycotoxin strip, five fungal toxins were simultaneously and quantitatively detected in naturally contaminated wheat. First, a matrix-based standard curve was established for the detection of aflatoxin B1 (AFB1), fumonisin B1 (FB1), T-2, deoxynivalenol (DON), and zearalenone (ZEN). Established on an open android system, the platform is able to read 6 lines on the strip simultaneously. The platform is equipped with a Quick Response code scanning model, which reads the established standard curves, and then rapidly quantify mycotoxins in naturally contaminated wheat. All the data and sample information are stored on a central server through the platform which is linked to the cloud. The limits of detection (LOD) for AFB1, FB1, T-2, DON, and ZEN in wheat were 4, 20, 10, 200, and 40 µg/kg and the visual cut off values was 20, 1000, 200, 4000, and 400 µg/kg, separately. To validate the platform and the multi-mycotoxin detection method, 10 wheat samples were analyzed and the results were in a good agreement with those obtained by LC-MS/MS. The platform will be a powerful tool for crop monitoring services.
Assuntos
Contaminação de Alimentos/análise , Imunoensaio/métodos , Micotoxinas/análise , Triticum/metabolismo , Aflatoxina B1/análise , Aflatoxina B1/imunologia , Aflatoxina B1/isolamento & purificação , Anticorpos/química , Anticorpos/imunologia , Fumonisinas/análise , Fumonisinas/imunologia , Fumonisinas/isolamento & purificação , Ouro/química , Limite de Detecção , Nanopartículas Metálicas/química , Micotoxinas/imunologia , Micotoxinas/isolamento & purificação , Sistemas Automatizados de Assistência Junto ao Leito , Triticum/química , Zearalenona/análise , Zearalenona/imunologia , Zearalenona/isolamento & purificaçãoRESUMO
Fungi produce several toxins active against plants, animal or humans. Among them, ribotoxins are enzymes that specifically attack ribosomes irreparably compromising protein synthesis, useful as insecticides or as anticancer agents. Here, a novel ribotoxin from the edible mushroom Pleurotus ostreatus has been purified and characterized. This ribotoxin, named Ostreatin, is a specific ribonuclease releasing α-fragment when incubated with yeast or rabbit ribosomes. Ostreatin shows IC50 of 234 pM in rabbit reticulocyte lysate, and metal dependent endonuclease activity. Following the completion of Ostreatin primary structure, we ascertained that this toxin is homologous to Ageritin, the first ribotoxin-like protein from the basidiomycete Agrocybe aegerita, with which it shares 38.8% amino acid sequence identity. Ostreatin consists of 131 amino acid residues with an experimental molecular mass of 14,263.51 Da ([M+H+]+). Homology modeling revealed that Ostreatin and Ageritin share a similar fold in which the common catalytic triad is conserved. Purified Ostreatin lacks N-terminal and C-terminal peptides, which instead are present in the Ostreatin coding sequence. Such peptides are probably involved in protein sorting and for this they could be removed. Our findings confirm the presence of ribotoxin-like proteins in basidiomycetes edible mushrooms, that we propose as novel tool for biotechnological applications.
Assuntos
Carpóforos/enzimologia , Micotoxinas/química , Pleurotus/enzimologia , Ribonucleases/química , Agaricales , Sequência de Aminoácidos , Ascomicetos/genética , Sequência de Bases , Cromatografia em Gel , Ativação Enzimática , Expressão Gênica , Modelos Moleculares , Micotoxinas/genética , Micotoxinas/isolamento & purificação , Micotoxinas/metabolismo , Conformação Proteica , Proteínas Recombinantes , Ribonucleases/genética , Ribonucleases/isolamento & purificação , Ribonucleases/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Relação Estrutura-AtividadeRESUMO
While complex sample pooling strategies have been developed for large-scale experiments with robotic liquid handling, many medium-scale experiments like mycotoxin screening by Enzyme-Linked Immunosorbent Assay (ELISA) are still conducted manually in 48- and 96-well plates. At this scale, the opportunity to save on reagent costs is offset by the increased costs of labor, materials, and risk-of-error caused by increasingly complex pooling strategies. This paper compares one-dimensional (1D), two-dimensional (2D), and Shifted Transversal Design (STD) pooling to study whether pooling affects assay accuracy and experimental cost and to provide guidance for when a human experimentalist might benefit from pooling. We approximated mycotoxin contamination in single corn kernels by fitting statistical distributions to experimental data (432 kernels for aflatoxin and 528 kernels for fumonisin) and used experimentally-validated Monte-Carlo simulation (10,000 iterations) to evaluate assay sensitivity, specificity, reagent cost, and pipetting cost. Based on the validated simulation results, assay sensitivity remains 100% for all four pooling strategies while specificity decreases as prevalence level rises. Reagent cost could be reduced by 70% and 80% in 48- and 96-well plates, with 1D and STD pooling being most reagent-saving respectively. Such a reagent-saving effect is only valid when prevalence level is < 21% for 48-well plates and < 13%-21% for 96-well plates. Pipetting cost will rise by 1.3-3.3 fold for 48-well plates and 1.2-4.3 fold for 96-well plates, with 1D pooling by row requiring the least pipetting. Thus, it is advisable to employ pooling when the expected prevalence level is below 21% and when the likely savings of up to 80% on reagent cost outweighs the increased materials and labor costs of up to 4 fold increases in pipetting.
Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Programas de Rastreamento/métodos , Micotoxinas/isolamento & purificação , Humanos , Método de Monte Carlo , Micotoxinas/química , Micotoxinas/genética , Zea mays/genética , Zea mays/microbiologiaRESUMO
Phytotoxins are widely found in plant pathogens. In recent years, many diseases caused by Neopestalotiopsis clavispora have been reported. To better understand the pathogenicity of N. clavispora, a solid fermentation strategy was employed to isolate and identify virulence factors afritoxinone B, afritoxinone A and oxysporone. The phytotoxic activities of these toxins were evaluated. Oxysporone exhibited high levels of phytotoxic activity after 72 h and the lesion area ranged from 21.5-84.3 mm2 after 9 days of treatment. The phytotoxic activities of the other two compounds were lower than that for oxysporone. The phytotoxic activity towards non-host organisms was also assessed for the three analyzed compounds; phytotoxic activity was observed in each case. Based on these results, we conclude that oxysporone is the main virulence factor in N. clavispora. We also suggest that each of the three compounds were non-host-specific toxins (NHST). To our knowledge, this is the first study to analyze phytotoxins produced by N. clavispora.
Assuntos
Ascomicetos/patogenicidade , Kadsura/microbiologia , Micotoxinas/metabolismo , Doenças das Plantas/microbiologia , Fatores de Virulência/metabolismo , Ascomicetos/metabolismo , Dioxanos/metabolismo , Espectroscopia de Ressonância Magnética , Micotoxinas/isolamento & purificação , Pironas/metabolismo , Fatores de Virulência/isolamento & purificaçãoRESUMO
In this study, 15 different mycotoxins were estimated in three staple cereals from selected agro-ecological regions in Nigeria using a 'novel' green extraction method, pressurized hot water extraction (PHWE) in comparison to a conventional solvent extraction method. Discrimination of the results of PHWE and solvent extraction using principal component analysis (PCA) and orthogonal projection to latent structures discriminate analysis (OPLS-DA) did not yield any differential clustering patterns. All maize samples (n = 16), 32% (n = 38) of sorghum and 35% (n = 37) of millet samples were positive for at least one of the 15 tested mycotoxins. Contamination levels for the cereals were higher in the warm humid rain forest region and gradually decreased towards the hot and arid region in the north of the country. The results demonstrate the applicability of PHWE as a possible alternative extraction method to conventional methods of extraction, which are solvent based.