Development of a method for the genetic identification of mussel species belonging to Mytilus, Perna, Aulacomya, and other genera

Legislation regarding the labeling of processed products is an important issue in the protection of consumer rights. This labeling is especially important in products that cannot be identified on the basis of their morphological characters, because these are removed from the animal in the transformation process. The goal of this study was the identification of mussel species using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) and Forensically Informative Nucleotide Sequencing (FINS) methodologies. The molecular marker selected was 18S rDNA (nuclear small-subunit rDNA gene), which allows identification at the genus level and at the species level in some cases. The genera included in this study were Mytilus, Perna, Aulacomya, Semimytilus, Brachidontes, Choromytilus, and Perumytilus. Different markers were used for genetic identification at the species level. To identify the species included in the genus Perna and Choromytilus, a fragment of ITS 1 (Internal Transcribed Spacer 1) was amplified by multiplex PCR and digested with restrictases. The species of Mytilus were identified by length polymorphism and RFLP of the polyphenolic adhesive protein gene. This methodology was validated with products manufactured in the authors' pilot plant and applied to commercial samples. Therefore, this sequential method can be completely or partially used to determine the mussel genus or species present in any food product.     

Antibacterial activity of coffee extracts and selected coffee chemical compounds against enterobacteria

The in vitro antimicrobial activity of commercial coffee extracts and chemical compounds was investigated on nine strains of enterobacteria. The antimicrobial activity investigated by the disc diffusion method was observed in both the extracts and tested chemical compounds. Even though pH, color, and the contents of trigonelline, caffeine, and chlorogenic acids differed significantly among the coffee extracts, no significant differences were observed in their antimicrobial activity. Caffeic acid and trigonelline showed similar inhibitory effect against the growth of the microorganisms. Caffeine, chlorogenic acid, and protocatechuic acid showed particularly strong effect against Serratia marcescens and Enterobacter cloacae. The IC(50) and IC(90) for the compounds determined by the microtiter plate method indicated that trigonelline, caffeine, and protocatechuic acids are potential natural antimicrobial agents against Salmonella enterica. The concentrations of caffeine found in coffee extracts are enough to warrant 50% of the antimicrobial effect against S. enterica, which is relevant to human safety.     

Analysis of herbicides in olive oil by liquid chromatography time-of-flight mass spectrometry

The application of liquid chromatography time-of-flight mass spectrometry (LC/TOF-MS) for the identification and quantitation of four herbicides (simazine, atrazine, diuron, and terbuthylazine) in olive oil samples is reported here. The method includes a sample treatment step based on a preliminary liquid-liquid extraction followed by matrix solid-phase dispersion (MSPD) using aminopropyl as a sorbent material. A final cleanup step is performed with florisil using acetonitrile as an eluting solvent. The identification by LC/TOF-MS is accomplished with the accurate mass (and the subsequent generated empirical formula) of the protonated molecules [M + H]+, along with the accurate mass of the main fragment ion and the characteristic chlorine isotope cluster present in all of them. Accurate mass measurements are highly useful in this type of complex sample analyses since they allow us to achieve a high degree of specificity, often needed when other interferents are present in the matrix. The mass accuracy typically obtained is routinely better than 2 ppm. The sensitivity, linearity, precision, mass accuracy, and matrix effects are studied as well, illustrating the potential of this technique for routine quantitative analyses of herbicides in olive oil. Limits of detection (LODs) range from 1 to 5 microg/kg, which are far below the required maximum residue level (MRL) of 100 microg/kg for these herbicides in olive oil.     

Contents of carboxylic acids and two phenolics and antioxidant activity of dried portuguese wild edible mushrooms

The organic acids and phenolics compositions of nine wild edible mushrooms species (Suillus bellini, Tricholomopsis rutilans, Hygrophorus agathosmus, Amanita rubescens, Russula cyanoxantha, Boletus edulis, Tricholoma equestre, Suillus luteus, and Suillus granulatus) were determined by HPLC-UV and HPLC-DAD, respectively. The antioxidant potential of these species was also assessed by using the DPPH* scavenging assay. The results showed that all of the species presented a profile composed of at least five organic acids: oxalic, citric, malic, quinic, and fumaric acids. In a general way, the pair of malic plus quinic acids were the major compounds. Only very small amounts of two phenolic compounds were found in some of the analyzed species: p-hydroxybenzoic acid (in A. rubescens, R. cyanoxantha, and T. equestre) and quercetin (in S. luteus and S. granulatus). All of the species exhibited a concentration-dependent scavenging ability against DPPH*. T. rutilans revealed the highest antioxidant capacity.     

Phenylphenalenone type compounds from the leaf fibers of abaca (Musa textilis)

A series of phenylphenalenone type compounds, known to play a role as phytoalexins in plants of the Musaceae family, have been identified for the first time in the leaf fibers of abaca (Musa textilis). Among the phenylphenalenone type compounds identified, the structure of a novel compound, (1R)-2,3-dihydro-4,9-dihydroxy-8-methoxy-1-phenylphenalene, has also been described in abaca fibers. Its structure was elucidated by analysis of one- and two-dimensional NMR (correlation spectroscopy, heteronuclear single quantum correlation, and heteronuclear multiple bond correlation) spectroscopic data.     

Influence of harvesting method and washing on the presence of pesticide residues in olives and olive oil

The influence of the preliminary steps in olive oil production (harvesting and washing) on pesticide residues in olives and olive oil has been investigated. Analyses were performed by GC-MS/MS and revealed that endosulfan sulfate and two herbicides (diuron and terbuthylazine) were the most frequently found residues in olives and olive oil. The harvesting method has a decisive influence on herbicide concentrations found in olives. Thus, 16 and 48% of the olive samples harvested on the ground after falling from the tree presented concentrations higher than the maximum residue limit (MRL) for diuron and terbuthylazine, respectively. In olives harvested directly from the tree, diuron was not found at concentrations higher than MRL and terbuthylazine was found in only 10% of the samples. The washing step performed routinely in olive mills was effective in removing the superficial contamination by herbicides present in olives harvested on the ground. Nevertheless, even after washing, the olive oil obtained from ground olives showed herbicide residue concentrations higher than those obtained from tree olives.     

Changes in the free amino acid contents of honeys during storage at ambient temperature

This study was carried out to establish the changes in the free amino acid contents of floral honeys, honeydew honeys, and blend honeys during storage at room temperature and to test the capacity of the amino acids to distinguish the origin of the honeys after storage. For this purpose, 54 artisanal honeys (39 floral, 5 honeydew, and 10 blend) were studied. Samples were taken from recently collected honeys and at 3, 6, 9, 12, 16, 20, and 24 months after harvesting. The contents of most of the free amino acids were found to decrease with storage time, with the greatest reduction observed in the first 9 months. The contents of the amino acids aspartic acid, beta-alanine, and proline increased in the first few months after storage, reaching maximum values at 6 months, suggesting the possible existence of enzymatic activities. The application of stepwise discriminant analysis to the free amino acid content data demonstrated that the contents of the amino acids valine, beta-alanine, gamma-aminobutyric acid, serine, isoleucine, alpha-alanine, ornithine, and glutamine correctly assigned 87% of honeys to their group of origin: floral, honeydew, or blend.     

Production and composition of cider spirits distilled in "alquitara"

The capacity of alquitara (a traditional distillation system) to produce cider brandies is evaluated. To do so, the chemical composition of 12 fractions obtained during the distillation process and the cider brandies obtained from five ciders were analyzed (alcohol strength, methanol, volatile substances, furfural, and metals), taking into account European and Spanish legislation. During the course of distillation, an important increase in methanol, furfural, 2-phenylethanol, and metals in the last fractions was observed, while fusel oils were more abundant in the first fractions collected. Only acetaldehyde behaved differently, showing a minimum concentration in the middle fractions that might be explained by its formation on the surface of alquitara. On the other hand, the final distillates obtained by means of this method complied with the considered regulations. Worth highlighting in this regard are the low levels of a potential toxin such as methanol, as well as the detection of a constant ratio for methanol, ethanol, and fusel oil for the pairs of cider/spirits analyzed, which could be interpreted as an indication of good uniformity in the distillation system and method, thus guaranteeing product quality.     

Influence of glycosidic linkages and molecular weight on the fermentation of maltose-based oligosaccharides by human gut bacteria

A structure-function study was carried out to increase knowledge of how glycosidic linkages and molecular weights of carbohydrates contribute toward the selectivity of fermentation by gut bacteria. Oligosaccharides with maltose as the common carbohydrate source were used. Potentially prebiotic alternansucrase and dextransucrase maltose acceptor products were synthesized and separated into different molecular weights using a Bio-gel P2 column. These fractions were characterized by matrix-assisted laser desorption/ionization time-of-flight. Nonprebiotic maltooligosaccharides with degrees of polymerization (DP) from three to seven were commercially obtained for comparison. Growth selectivity of fecal bacteria on these oligosaccharides was studied using an anaerobic in vitro fermentation method. In general, carbohydrates of DP3 showed the highest selectivity towards bifidobacteria; however, oligosaccharides with a higher molecular weight (DP6-DP7) also resulted in a selective fermentation. Oligosaccharides with DPs above seven did not promote the growth of "beneficial" bacteria. The knowledge of how specific structures modify the gut microflora could help to find new prebiotic oligosaccharides.     

Evaluation of the health of free-ranging greater rheas (Rhea americana) in Argentina

The health of 22 free-ranging adult rheas (Rhea americana) examined and sampled during a translocation/reintroduction project and six juvenile rheas kept in semicaptivity was investigated, and details of their haematology and plasma biochemistry are presented. Serological testing for antibodies to infectious agents was negative for infectious laryngotracheitis, avian adenovirus, avian influenza, avian reovirus, infectious bursal disease, infectious bronchitis virus, paramyxovirus types 1, 2, and 3, fowlpox and Salmonella Pullorum. Antibodies to Chlamydophila species were found in 25 of 27 of the birds, and 22 of 25 had antibodies to Aspergillus species. Ova of gastrointestinal nematodes of the genus Capillaria were identified, and the anoplocephalid cestode Monoecocestus cf rheiphilus was identified in R americana for the first time.