Simultaneous determination of aflatoxins B1, B2, G1 and G2 in commercial rices using immunoaffinity column clean-up and HPLC-MS/MS.

Rice is frequently contaminated with aflatoxins, that are highly toxic fungal substances and strongly involved on hepatic cancer. In this work, different extraction and clean-up methods were evaluated for the simultaneous extraction and clean-up of aflatoxins B1, B2, G1 and G2 from rice. Favourable results were obtained by using methanol - water (80:20, v/v) extraction followed by immunoaffinity columns for clean-up, with recoveries of 86-92%, standard deviations between 5 and 11%, LOD ranged between 0.09 and 0.32 µg/kg, and LOQ between 0.31 and 1.06 µg/kg. Method validation and sample analysis were performed by using HPLC-MS/MS. Nine rice samples from different origin, varieties and specific characteristics, acquired in Spanish supermarkets were analysed. In two basmati samples from the same batch aflatoxin B1 was detected at (1.62 ± 0.08) µg/kg and (0.77 ± 0.03) µg/kg, both lower than the levels established by European Regulation for aflatoxin B1 in cereals.

Selenium bio-enrichment of Mediterranean fruit juices through lactic acid fermentation.

This work was carried out to elaborate selenium (Se) bio-enriched fermented Mediterranean fruit juices. To this purpose, pomegranate and table red grape juices were added with sodium selenite (Na2SeO3) and fermented by Levilactobacillus brevis CRL 2051 and Fructobacillus tropaeoli CRL 2034 individually or combined. To better evaluate the effect of selenite addition and starter strain inoculums on the total bacterial community of the fruit juices, fermentation trials were performed with raw and pasteurized fruit juices. No statistical significant differences were observed for total mesophilic microorganisms (TMM) and rod-shaped lactic acid bacteria (LAB) levels among raw and pasteurized juices inoculated with the starter strains, while significant differences between those juices with and without selenite were registered. LAB cocci, Pseudomonadaceae and yeasts were detected only for the raw juice preparations. The dominance of L. brevis CRL 2051 and F. tropaeoli CRL 2034 was confirmed by randomly amplified polymorphic DNA (RAPD)-PCR analysis. After fermentation, pH dropped for all inoculated trials and control raw juices. The soluble solid content (SSC) levels of the raw juices were higher than the corresponding pasteurized trials. The thermal treatment affected consistently yellowness of grape juice trials and redness of pomegranate juices. No microbial Se accumulation was registered for pomegranate juices, while F. tropaeoli CRL 2034 accumulated the highest amount of Se (65.5 µg/L) in the grape juice. For this reason, only trials carried out with raw grape juices were investigated by metagenomics analysis by Illumina MiSeq technology. Non-inoculated grape juices were massively fermented by acetic acid bacteria while Fructobacillus and Lactobacillus (previous genus name of Levilactobacillus) represented the highest operational taxonomy units (OTUs) relative abundance % of the trials inoculated with the starter strains as confirmed by this technique.

Biotransformation of Selenium by Lactic Acid Bacteria: Formation of Seleno-Nanoparticles and Seleno-Amino Acids.

Selenium (Se) is an essential micronutrient for the majority of living organisms, and it has been identified as selenocysteine in the active site of several selenoproteins such as glutathione peroxidase, thioredoxin reductase, and deiodinases. Se deficiency in humans is associated with viral infections, thyroid dysfunction, different types of cancer, and aging. In several European countries as well as in Argentina, Se intake is below the recommended dietary Intake (RDI). Some lactic acid bacteria (LAB) can accumulate and bio-transform selenite (toxic) into Se-nanoparticles (SeNPs) and Se-amino acids (non-toxic). The microbial growth, Se metabolite distribution, and the glutathione reductase (involved in selenite reduction) activity of Se-enriched LAB were studied in this work. The ninety-six assayed strains, belonging to the genera Lactococcus, Weissella, Leuconostoc, Lactobacillus, Enterococcus, and Fructobacillus could grow in the presence of 5 ppm sodium selenite. From the total, eight strains could remove more than 80% of the added Se from the culture medium. These bacteria accumulated intracellularly between 1.2 and 2.5 ppm of the added Se, from which F. tropaeoli CRL 2034 contained the highest intracellular amount. These strains produced only the seleno-amino acid SeCys as observed by LC-ICP-MS and confirmed by LC-ESI-MS/MS. The intracellular SeCys concentrations were between 0.015 and 0.880 ppm; Lb. brevis CRL 2051 (0.873 ppm), Lb. plantarum CRL 2030 (0.867 ppm), and F. tropaeoli CRL 2034 (0.625 ppm) were the strains that showed the highest concentrations. Glutathione reductase activity values were higher when the strains were grown in the presence of Se except for the F. tropaeoli CRL 2034 strain, which showed an opposite behavior. The cellular morphology of the strains was not affected by the presence of Se in the culture medium; interestingly, all the strains were able to form spherical SeNPs as determined by transmission electron microscopy (TEM). Only two Enterococcus strains produced the volatile Se compounds dimethyl-diselenide identified by GC-MS. Our results show that Lb. brevis CRL 2051, Lb. plantarum CRL 2030, and F. tropaeoli CRL 2034 could be used for the development of nutraceuticals or as starter cultures for the bio-enrichment of fermented fruit beverages with SeCys and SeNPs.

Residual brewing yeast as a source of polyphenols: Extraction, identification and quantification by chromatographic and chemometric tools.

A method combining aqueous extraction, reversed-phase high-performance capillary liquid chromatography with photodiode array detection (cLC-DAD) and chemometric tools, was developed to determine phenolic compounds in residual brewing yeast. The effect of temperature, nature of extraction solvent and method for separation of extract solution were studied to optimize the extraction conditions on the basis of total phenolic content (TPC), total flavonoids content (TFC) and antioxidant capacity. Polyphenols were determined by cLC-DAD. Flavonols as rutin and kaempferol, flavonoids as naringin, phenolic acids as gallic acid and antioxidants as trans-ferulic and p-coumaric acids were found and quantified in the brewing residue. Data were subjected to evaluation using multifactor ANOVA and principal component analysis (PCA), both showing that lyophilization pretreatment affects the content of individual polyphenols and that residual brewing yeast contains higher polyphenol amounts than the liquid beer waste. The obtained results suggest that residual brewing yeast could be a source of polyphenols.

Establishment of selenium uptake and species distribution in lupine, Indian mustard, and sunflower plants.

Selenium has been recognized as essential for all mammals; therefore, its concentration level and speciation are of great concern. Plants are one of the main sources of selenium in the diet. Thus, inorganic selenium uptake and its transformation in different species were evaluated in Indian mustard (Brassica juncea), sunflower (Helianthus annus), and white lupine (Lupinus albus). More than 1.2 g x kg(-)(1) (dry matter) of Se was found in the aerial part of Indian mustard when growing on 1 mg x L(-)(1) of Se as Na(2)SeO(4), and approximately half this amount was determined in the leaves of the lupine, which is still quite high. Selenomethionine was the main selenium-containing amino acid identified in most of the extracts by HPLC-ICP-MS. The higher values were 6.8 and 14.5 mg x kg(-)(1) (expressed as Se in dry matter) in the leaves of lupine and sunflower, respectively. This is of great importance because some authors have considered the combination of this enriched material with non-enriched food as a source of selenium supplementation.