Gut Microbiota and Nutrition: Strategies for the Prevention and Treatment of Type 2 Diabetes.

The prevalence of diabetes has increased in last decades worldwide and is expected to continue to do so in the coming years, reaching alarming figures. Evidence have shown that patients with type 2 diabetes (T2D) have intestinal microbial dysbiosis. Moreover, several mechanisms link the microbiota with the appearance of insulin resistance and diabetes. Diet is a crucial factor related to changes in the composition, diversity, and activity of gut microbiota (GM). In this review, the current and future possibilities of nutrient-GM interactions as a strategy to alleviate T2D are discussed, as well as the mechanisms related to decreased low-grade inflammation and insulin resistance. A bibliographic search of clinical trials in Pubmed, Web of Science, and Scopus was carried out, using the terms "gut microbiota, diet and diabetes." The data analyzed in this review support the idea that dietary interventions targeting changes in the microbiota, including the use of prebiotics and probiotics, can improve glycemic parameters. However, these strategies should be individualized taking into account other internal and external factors. Advances in the understanding of the role of the microbiota in the development of metabolic diseases such as T2D, and its translation into a therapeutic approach for the management of diabetes, are necessary to allow a comprehensive approach.

Use of Mustard Extracts Fermented by Lactic Acid Bacteria to Mitigate the Production of Fumonisin B1 and B2 by Fusarium verticillioides in Corn Ears.

Corn (Zea mays) is a worldwide crop subjected to infection by toxigenic fungi such as Fusarium verticillioides during the pre-harvest stage. Fusarium contamination can lead to the synthesis of highly toxic mycotoxins, such as Fumonisin B1 (FB1) and Fumonisin B2 (FB2), which compromises human and animal health. The work aimed to study the antifungal properties of fermented yellow and oriental mustard extracts using nine lactic acid bacteria (LAB) in vitro. Moreover, a chemical characterization of the main phenolic compounds and organic acids were carried out in the extracts. The results highlighted that the yellow mustard, fermented by Lactiplantibacillus plantarum strains, avoided the growth of Fusarium spp. in vitro, showing Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC) values, ranging from 7.8 to 15.6 g/L and 15.6 to 31.3 g/L, respectively. Then, the lyophilized yellow mustard fermented extract by L. plantarum TR71 was applied through spray-on corn ears contaminated with F. verticillioides to study the antimycotoxigenic activity. After 14 days of incubation, the control contained 14.71 mg/kg of FB1, while the treatment reduced the content to 1.09 mg/kg (92.6% reduction). Moreover, no FB2 was observed in the treated samples. The chemical characterization showed that lactic acid, 3-phenyllactic acid, and benzoic acid were the antifungal metabolites quantified in higher concentrations in the yellow mustard fermented extract with L. plantarum TR71. The results obtained confirmed the potential application of fermented mustard extracts as a solution to reduce the incidence of mycotoxins in corn ears.

Extraction of Phenolic Compounds from Fresh Apple Pomace by Different Non-Conventional Techniques.

Red Delicious apple pomace was produced at laboratory scale with a domestic blender and different non-conventional extraction techniques were performed to isolate phenolic compounds, such as ultrasound-assisted extraction (UAE), ultraturrax extraction (UTE), accelerated solvent extraction (ASE) and pulsed electric field (PEF) extraction pre-treatment. Total phenolic content (TPC) was determined by Folin-Ciocalteu assay. Phloridzin, the main phenolic compound in apples, was determined by chromatographic analysis Q-TOF-LC/MS. The results obtained with these techniques were compared in order to identify the most efficient method to recover polyphenols. The highest value of TPC (1062.92 ± 59.80 µg GAE/g fresh apple pomace) was obtained when UAE was performed with EtOH:H2O (50:50, v/v), while ASE with EtOH:H2O (30:70, v/v) at 40 °C and 50% of flush was the most efficient technique in the recovery of phloridzin. The concentration of the main phenolic compounds ranged from 385.84 to 650.56 µg/g fresh apple pomace. The obtained results confirm that apple pomace represents an interesti-ng by-product, due to the presence of phenolic compounds. In particular, phloridzin could be considered a biomarker to determine the quality of numerous apple products. Therefore, this research could be a good starting point to develop a value-added product such as a functional food or nutraceutical.

Multi-mycotoxin contamination of green tea infusion and dietary exposure assessment in Moroccan population.

Green tea infusion is one of the most widely drunk beverages worldwide due to its health benefits associated with microelements, essential oils, and polyphenols, etc. Several studies have reported that green tea is subjected to contamination by various toxigenic fungi. Thus, this work aims to investigate the co-occurrence of 15 mycotoxins [four aflatoxins (AFB1, AFB2, AFG1, AFG2), ochratoxin A (OTA), beauvericin (BEA), four enniatins (ENA, ENA1, ENB, ENB1), zearalenone (ZEN), alternariol (AOH), tentoxin (TENT), T-2 and HT-2 toxins] in green tea samples available in Morocco by liquid chromatography tandem mass spectrometry method. Analytical and consumption data were then used to assess the dietary exposure for the population. Out of 111 total green tea samples, 62 (56%) were contaminated by at least one mycotoxin. The most found mycotoxins in samples were AOH (40%), ZEN (35%), AFG1 (2%), AFB2 (2%), ENB (2%) and TENT (1%). The highest level was found for ZEN with 45.8 ng/g. There is no sample that exceeded the recommended levels set by European Pharmacopoeia for certain mycotoxins in plant material. Although multi-mycotoxin co-occurred in samples (33%), the probable estimated daily intake values show that the intake of mycotoxins through the consumption of green tea does not represent a risk for the population.

Occurrence of Free and Conjugated Mycotoxins in Aromatic and Medicinal Plants and Dietary Exposure Assessment in the Moroccan Population.

Aromatic and medicinal plants (AMPs), as herbal material, are subjected to contamination by various mycotoxin-producing fungi, either free and conjugated. Such a problem is associated with poor storage practices, and lack of adopting good agricultural practices and good harvesting practices. Nevertheless, AMPs are poorly investigated. The purpose of this study was to investigate the co-occurrence of 15 mycotoxins (four aflatoxins (AFB1, AFB2, AFG1, and AFG2), ochratoxin A (OTA), beauvericin (BEA), four enniatins (ENA, ENA1, ENB, and ENB1), zearalenone (ZEN), alternariol (AOH), tentoxin (TENT), T-2, and HT-2 toxins) in 40 samples of AMPs frequently consumed in Morocco by using liquid chromatography tandem mass spectrometry. Evaluation of conjugated mycotoxins and their identification using liquid chromatography coupled to time-of-flight mass spectrometry with ion mass exact was also carried out. Results showed that 90% of the analyzed samples presented at least one mycotoxin, and 52% presented co-occurrence of them. Mycotoxins detected were: AOH (85%), ZEN (27.5%), ß-ZEL (22%), AFG1 (17.5%), TENT (17.5%), ENB (10%), AFG2 (7.5%), α-ZEL (5%), ENA1 (2.5%), and HT-2 (2.5%), while the conjugated mycotoxins were ZEN-14-Glc (11%) and ZEN-14-Sulf (9%). The highest observed level was for AOH, with 309 ng/g. Ten samples exceeded the recommended levels set by the European Pharmacopoeia for AF mycotoxins in plant material (4 ng/g), and three samples exceeded the maximum limits for AFs (10 ng/g) in species established by the European Commission. Although the co-occurrence of several mycotoxins in AMP samples was observed, the dietary exposure assessment showed that the intake of mycotoxins through the consumption of AMP beverages does not represent a risk for the population.

Coffee Silverskin and Spent Coffee Suitable as Neuroprotectors against Cell Death by Beauvericin and α-Zearalenol: Evaluating Strategies of Treatment.

Coffee silverskin and spent coffee have been evaluated in a neuroblastoma cell line (SH-SY5Y cells) against beauvericin (BEA) and α-zearalenol (α-ZEL)-induced cytotoxicity with different strategies of treatment. First, the direct treatment of mycotoxins and coffee by-products extracts in SH-SY5Y cells was assayed. IC50 values for α-ZEL were 20.8 and 14.0 µM for 48 h and 72 h, respectively and, for BEA only at 72 h, it was 2.5 µM. Afterwards, the pre-treatment with spent coffee obtained by boiling water increased cell viability for α-ZEL at 24 h and 48 h from 10% to 16% and from 25% to 30%, respectively; while with silverskin coffee, a decrease was observed. Opposite effects were observed for BEA where an increase for silverskin coffee was observed at 24 h and 48 h, from 14% to 23% and from 25% to 44%, respectively; however, a decrease below 50% was observed for spent coffee. Finally, the simultaneous treatment strategy for the highest concentration assayed in SH-SY5Y cells provided higher cytoprotection for α-ZEL (from 44% to 56% for 24 h and 48 h, respectively) than BEA (30% for 24 h and 48 h). Considering the high viability of coffee silverskin extracts for SH-SY5Y cells, there is a forthcoming promising use of these unexploited residues in the near future against mycotoxins effects.

Reducing the effect of beauvericin on neuroblastoma SH-SY5Y cell line by natural products.

In the present work, different natural compounds from coffee by-product extracts (coffee silverskin and spent coffee) rich in polyphenols, was investigated against beauvericin (BEA) induced-cytotoxicity on SH-SY5Y cells. Spent coffee arise as waste products through the production of instant coffee and coffee brewing; while the silverskin is a tegument which is removed and eliminated with toasting coffee grains. First of all, polyphenol extraction methods, measurement of total polyphenols content and its identification were carried out. Afterwards evaluating in vitro effects with MTT assay on SH-SY5Y cells of coffee by-product extracts and mycotoxins at different concentrations and exposure times was performed. TPC in silverskin coffee by-product extracts was >10 times higher than in spent coffee by-product extracts. Chlorogenic acid was the majority polyphenol detected. Viability for BEA reached IC50 values at 72h (2.5 µM); boiling water silverskin coffee extract reached the highest viability also in pre-treatment BEA exposure and compared with MeOH and MeOH:H2O (v/v, 50:50) extracts. These results in SH-SY5Y cells highlight the use of such residues as supplements or bioactive compounds in the future.

Chemoprotective effect of carotenoids from Lycium barbarum L. on SH-SY5Y neuroblastoma cells treated with beauvericin.

Goji berry has recently been introduced in Mediterranean diet and its consumption is increasing. This study aims to determine cytoprotection of lutein (LUT), zeaxanthin (ZEAX) and goji berry extract (GBE) rich in carotenoids against Beauvericin (BEA)-induced cytotoxicity on SH-SY5Y neuroblastoma cells. Both carotenoids and GBE showed cytoprotective effects. Cytoprotection was evaluated by simultaneous combination of the two xanthophylls LUT and ZEAX with BEA, as well as using pre-treatment assays. The highest protective effect occurred in 16%, 24% and 12% respectively for LUT, ZEAX and LUT + ZEAX incubating simultaneously with BEA, while by pre-treatment assay LUT showed a cytoprotection effect over 30% and ZEAX alone or LUT + ZEAX promoted only a slight cytoprotection (<10%). Pre-treatment assays with GBE, showed a cytoprotection, between 3 and 20%, for BEA concentrations ranging from 0.1 to 6.25 µM, whereas no protective effect was observed when the cells were simultaneously incubated with GBE and BEA. Finally, by means of CI-isobologram method, the interaction between LUT, ZEAX and BEA were evaluated, and the results showed an synergism effect for almost all combinations tested. The data presented shows a option of using goji berries to potentially mitigate the toxicity of beauvericin eventually present in foods.

Cytoprotective effects of carotenoids-rich extract from Lycium barbarum L. on the beauvericin-induced cytotoxicity on Caco-2 cells.

In this work, the cytotoxicity of Beauvericin (BEA), lutein (LUT), zeaxanthin (ZEAX) and goji berries extract (GBE) rich in carotenoids, was investigated, as well as cytoprotective effects of these carotenoids against BEA induced-cytotoxicity on Caco-2 cells. Cytotoxicity was carried out using MTT and protein content (PC) assays during 24 and 48 h of exposure. Only BEA showed cytotoxic effect obtaining a reduction in cell proliferation range from 6.5 to 92.8%. Simultaneous combination of LUT and ZEAX with BEA slightly increased cell proliferation compared to BEA tested alone. LUT, ZEAX and GBE showed cytoprotective effects against cytotoxicity induced by BEA on Caco-2 cells. Pre-treatment assays showed the highest cytoprotection effect at the highest dose of BEA assayed (2.5 µM) in 29%, 31% and 35% for LUT, ZEAX and LUT + ZEAX, respectively; GBE showed a cytoprotection of 20%, for the same dose of BEA. The interaction between LUT, ZEAX and BEA studied by means of CI-isobologram method showed a synergism and antagaonism effect for all the combinations tested. These findings highlight that food containing high level of carotenoids, as goji berries, could contribute to reduce the toxicological risk that natural contaminant as BEA mycotoxin in diet can produce to the humans.

Evaluating the impact of supercritical-CO2 pressure on the recovery and quality of oil from "horchata" by-products: Fatty acid profile, α-tocopherol, phenolic compounds, and lipid oxidation parameters.

The effect of supercritical carbon dioxide (SC-CO2) (10-40 MPa) and conventional extraction (CE) to recover oil from by-products obtained during "horchata" production was assessed. To evaluate both extraction techniques, the fatty acid composition, polyphenols, α-tocopherol, antioxidant capacity and lipid oxidation parameters of the extracts obtained were analysed. A linear relationship between extraction pressure and oil yield was observed. However, the highest oil yield was obtained under conventional extraction. The by-products from "horchata" presented a profile where monounsaturated fatty acids (MUFA) were the predominant, representing ≈ 70% of total fatty acids. The amount of saturated fatty acids (SFA) and polyunsaturated fatty acids (PUFA) was higher and MUFA lower at 10 MPa samples compared to the oils extracted using SC-CO2 at 20, 30 or 40 MPa, where no differences were detected. The content of α-tocopherol was significantly higher after SC-CO2 treatments compared to conventional extraction, independently of the applied treatment. On the other hand, the values of phenolic compounds and total antioxidant activity (TAC) increased as the pressure conditions of the SC-CO2 extraction increased, presenting a linear adjustment of the data. Regarding lipid oxidation, the lower oxidation indexes were obtained when the SC-CO2 pressure increased. Finally, our results confirmed that the application of SC-CO2 could be a potential alternative to conventional extraction in order to obtain oils from "horchata" by-products rich in high-added value compounds without the use of organic solvents which can be toxic.

Occurrence of Mycotoxins in Botanical Dietary Supplement Infusion Beverages.

The aim of the present work was to study the occurrence of mycotoxins [aflatoxins (1-4), 3-acetyldeoxyniavlenol (5), 15-acetyldeoxynivalenol (6), nivalenol (7), HT-2 (8), T-2 (9), ochratoxin A (10), zearalenone (11), enniatin A (12), enniatin A1 (13), enniatin B (14), enniatin B1 (15), and beauvericin (16)] present in potable products derived from herbal teas. Analysis was carried out by liquid chromatography coupled to ion-trap tandem mass spectrometry (LC-MS/MS-IT) after a dispersive liquid-liquid microextraction procedure (DLLME) was conducted. The DLLME method was applied to 52 commercial samples of chamomile, chamomile with anise, chamomile with honey, linden, pennyroyal mint, thyme, valerian, and horsetail beverages. The results obtained showed that the following mycotoxins were detected in the samples: 2 (19.1 to 134.7 µg/L), 3 (below the limit of quantification), and 4 (2.2 to 13.5 µg/L). Also, 6 was detected in one sample at 112.5 µg/L, and 14 was detected only in two samples, although at very low concentration levels. Pennyroyal mint and thyme showed the highest concentration levels of mycotoxins. A risk assessment, however, showed negative results regarding the consumption of herbal tea beverages and the presence of mycotoxins.

Amylase-Trypsin Inhibitors in Wheat and Other Cereals as Potential Activators of the Effects of Nonceliac Gluten Sensitivity.

Nonceliac gluten sensitivity (NCGS) is a gluten-related gastrointestinal disorder distinct from celiac disease (CD) and gluten allergy that is not easy to diagnose due to the lack of biomarkers. It is characterized by intestinal symptoms and extraintestinal manifestations with the consumption of gluten-containing foods. In contrast to CD, NCGS patients do not present a genetic predisposition or intestinal villi atrophy. Recent studies question the proinflammatory triggering activity of α-gliadin fraction contained in wheat, since it has been demonstrated that the amylase-trypsin inhibitors (ATIs) exert a strong activating effect on the innate immune response. We aimed to analyze the role of ATIs in the activation of innate immunity and in the development of the symptoms characteristic of NCGS. A systematic literature search was made using databases such as MEDLINE, SciELO, Science Direct, and Scopus, with focus on key words such as "amylase-trypsin inhibitors," "wheat," "gluten," and "celiac." Many studies are available on the structure, inhibition mechanism, and immune system effects of ATIs, mainly focused on IgE-mediated reactions. Recently, with the increase of NCGS interest, has increased the literature on the capacity of ATIs contained in wheat to activate the innate immune system. Literature published to date questions the relationship between activation of the innate immune system and gluten in NCGS. ATIs may have acted as interfering contaminant of gluten and appear as potential activator of innate immunity in NCGS patients. In view of their potential impact, more interventional studies are needed to demonstrate the proinflammatory effect of ATIs.

Multimycotoxin LC-MS/MS Analysis in Tea Beverages after Dispersive Liquid-Liquid Microextraction (DLLME).

The aim of the present study was to develop a multimycotoxin liquid chromatography tandem mass spectrometry (LC-MS/MS) method with a dispersive liquid-liquid microextraction procedure (DLLME) for the analysis of AFs, 3aDON, 15aDON, NIV, HT-2, T-2, ZEA, OTA, ENNs, and BEA in tea beverages and to evaluate their mycotoxin contents. The proposed method was characterized in terms of linearity, limits of detection (LODs), limits of quantification (LOQs), recoveries, repeatability (intraday precision), reproducibility (interday precision), and matrix effects to check suitability. The results show LODs in the range of 0.05-10 µg/L, LOQs in the range of 0.2-33 µg/L, and recoveries in the range of 65-127% (RSD < 20%). The method developed in this study was applied to 44 commercial samples of black tea, red tea, green tea, and green mint tea. The results show that, of the analyzed mycotoxins, AFB2, AFG2, 15aDON, AFG1, and ENB were detected in the samples. AFB2 (14.4-32.2 µg/L) and 15aDON (60.5-61 µg/L) presented the highest levels. Green mint tea contained the highest concentration of mycotoxins. The risk assessment study shows that the population is not much exposed to mycotoxins through the consumption of tea beverages.

Influence of the antimicrobial compound allyl isothiocyanate against the Aspergillus parasiticus growth and its aflatoxins production in pizza crust.

Aflatoxins (AFs) are secondary metabolites produced by different species of Aspergillus, such as Aspergillus flavus and Aspergillus parasiticus, which possess mutagenic, teratogenic and carcinogenic activities in humans. In this study, active packaging devices containing allyl isothiocyanate (AITC) or oriental mustard flour (OMF) + water were tested to inhibit the growth of A. parasiticus and AFs production in fresh pizza crust after 30 d. The antimicrobial and anti-aflatoxin activities were compared to a control group (no antimicrobial treatment) and to a group added with commercial preservatives (sorbic acid + sodium propionate). A. parasiticus growth was only inhibited after 30 d by AITC in filter paper at 5 µL/L and 10 µL/L, AITC sachet at 5 µL/L and 10 µL/L and OMF sachet at 850 mg + 850 µL of water. However, AFs production was inhibited by all antimicrobial treatments in a dose-dependent manner. More importantly, AITC in a filter paper at 10 µL/L, AITC sachet at 10 µL/L, OMF sachet at 850 mg + 850 µL of water and sorbic acid + sodium propionate at 0.5-2.0 g/Kg completely inhibited AFs formation. The use of AITC in active packaging devices could be a natural alternative to avoid the growth of mycotoxinogenic fungi in refrigerated bakery products in substitution of common commercial preservatives.

Determination of mycotoxins in bee pollen by gas chromatography-tandem mass spectrometry.

Bee pollen, promoted as a natural food supplement, is consumed increasingly by people to maintain a healthy diet. Depending on environmental conditions, pollen can also be an optimum medium for growth of molds such as Fusarium and Penicillium . A quick, easy, cheap, rapid, and safe (QuEChERS) extraction procedure followed by a gas chromatography-tandem mass spectrometry (GC-MS/MS) determination of eight selected Fusarium toxins in bee pollen was developed and optimized. Recovery studies at 20, 80, and 1000 µg/kg showed values between 73 and 95% with relative standard deviations (RSDs) of <15% for all studied mycotoxins. Limits of quantitation (LOQ) ranged from 1 to 4 µg/kg. The proposed method was applied to the analysis of 15 commercial samples. Two of 15 samples showed quantifiable values for neosolaniol and nivalenol.

Optimization of Matrix Solid-Phase Dispersion method for simultaneous extraction of aflatoxins and OTA in cereals and its application to commercial samples.

A method based on Matrix Solid-Phase Dispersion (MSPD) has been developed for the determination of 5 mycotoxins (ochratoxin A and aflatoxins B and G) in different cereals. Several dispersants, eluents and ratios were tested during the optimization of the process in order to obtain the best results. Finally, samples were blended with C(18) and the mycotoxins were extracted with acetonitrile. Regarding to matrix effects, the results clearly demonstrated the necessity to use a matrix-matched calibration to validate the method. Analyses were performed by liquid chromatography-triple quadrupole-tandem mass spectrometry (LC-QqQ-MS/MS). The recoveries of the extraction process ranged from 64% to 91% with relative standard deviation lower than 19% in all cases, when samples were fortified at two different concentrations levels. Limits of detection ranged from 0.3 ng g(-1) for aflatoxins to 0.8 ng g(-1) for OTA and the limits of quantification ranged from 1 ng g(-1) for aflatoxins to 2 ng g(-1) for OTA, which were below the limits of mycotoxins set by European Union in the matrices evaluated. Application of the method to the analysis of several samples purchased in local supermarkets revealed aflatoxins and OTA levels.

Apple-products phytochemicals and processing: a review.

Epidemiological studies have linked the consumption of apples with reduced risk of some cancers, cardiovascular disease, asthma, and diabetes. Extensive research exists on apples and the health benefits of their beverages and phytochemicals. The purpose of this paper is to review the most recent literature in this area focusing on phytochemicals, phytochemical bioavailability and antioxidant behavior.

Biocatalyzed acidolysis of soybean oil triacylglycerols to increase oleic acid content.

Lipase catalyzed acidolysis of triacylglycerols (TAG) of soybean oil with oleic acid in organic solvent was studied; immobilized lipase from Rhizomucor miehei was used and the effects of reaction time, incubation temperature and enzyme load on TAG total and positional fatty acid (FA) percentage compositions were investigated. The results show that oleic acid incorporation was high after 24 and 48 h, while after 72 h a lower level of oleic acid in TAG was observed. Moreover, for the reactions carried out at 30 and 40 degrees C, it was observed that the oleic acid level was about 46.5% while ligtly higher values (about 49%) were observed at 50 degrees C; however, under this last condition, the modification of sn-2 position FA composition was higher. Finally, the variable enzyme load resulted also important on the incorporation of oleic acid; in particular, even if the value of 10% (w/w) of enzyme load resulted in a lightly lower incorporation of oleic acid in soybean oil TAG (about 45%), in this situation a minimal modification of sn-2 position FA composition was obtained.

Off-line solid-phase microextraction and capillary electrophoresis mass spectrometry to determine acidic pesticides in fruits.

A method based on solid-phase microextraction (SPME) and capillary electrophoresis/mass spectrometry (CE/ MS) is described for determining simultaneously five acidic pesticides (o-phenylphenol, ioxynil, haloxyfop, acifluorfen, picloram) in fruits. The CE device is coupled to an electrospray interface by a commercial sheath-flow adapter. Emphasis is placed on fulfillment of the speed and sensitivity requirements. The best separation is achieved using 32 mM ammonium formate/acid formic buffer at pH 3.1, with a working voltage of 25 kV. The MS detection of the five pesticides was performed in negative ionization mode. Full-scan spectra with base peaks corresponding to [M-H]- were obtained except for acifluorfen, which gives [M-H-CO2]- as most abundant ion. Compared with the conventional EC-UV, the limits of detection were lower for acifluorfen, haloxyfop, ioxynil, and picloram, by a factor of 20, 20, 50, and 2, respectively. Extraction involved fruit sample homogenization with an acetone-water solution (5:1), filtration, and acetone evaporation prior to fiber extraction. SPME conditions such as time, pH, ion strength, stationary phase of the fiber, sample matrix, and desorption solvents were examined. The recovery of the analytes ranged from 7 to 94%, and the relative standard deviation was between 3 and, 13%. The method was found to be linear between 0.02 and 500 mg kg(-1) with correlation coefficients ranging from 0.992 to 0.997. The limits of quantification were from 0.02 to 5 mg kg(-1). The optimized method was successfully applied to the analysis of acid pesticides in fruit samples.