Iñaki Berregi

Quantitative determination of caffeine, formic acid, trigonelline and 5-(hydroxymethyl)furfural in soluble coffees by 1H NMR spectrometry.

A quantitative method for the determination of caffeine, formic acid, trigonelline and 5-(hydroxymethyl)furfural (5-HMF) in soluble coffees by applying the proton nuclear magnetic resonance technique ((1)H NMR) is proposed. Each of these compounds records a singlet signal at the 7.6-9.5 ppm interval of the spectrum, and its area is used to determine the concentration. 3-(Trimethylsilyl)-2,2,3,3-tetradeuteropropionic acid is added in an exact known concentration as a reference for delta=0.00 ppm and as an internal standard. The method is applied to commercial soluble coffees and satisfactorily compared with results obtained by standard methods. The limits of detection and the coefficients of variation (N=10) are, respectively, 1.32 mg/g of solid product and 4.2% for caffeine, 0.45 mg/g and 2.6% for formic acid, 0.58 mg/g and 2.4% for trigonelline, and 0.30 mg/g and 7.3% for 5-HMF. The described method is direct and no previous derivatization is needed.

Glycerol metabolism and bitterness producing lactic acid bacteria in cidermaking.

Several lactic acid bacteria were isolated from bitter tasting ciders in which glycerol was partially removed. The degradation of glycerol via glycerol dehydratase pathway was found in 22 out of 67 isolates. The confirmation of glycerol degradation by this pathway was twofold: showing their glycerol dehydratase activity and detecting the presence of the corresponding gene by a PCR method. 1,3-propanediol (1,3-PDL) and 3-hydroxypropionic acid (3-HP) were the metabolic end-products of glycerol utilization, and the accumulation of the acrolein precursor 3-hydroxypropionaldehyde (3-HPA) was also detected in most of them. The strain identification by PCR-DGGE rpoB showed that Lactobacillus collinoides was the predominant species and only 2 belonged to Lactobacillus diolivorans. Environmental conditions conducting to 3-HPA accumulation in cidermaking were studied by varying the fructose concentration, pH and incubation temperature in L. collinoides 17. This strain failed to grow with glycerol as sole carbon source and the addition of fructose enhanced both growth and glycerol degradation. Regarding end-products of glycerol metabolism, 1,3-PDL was always the main end-product in all environmental conditions assayed, the only exception being the culture with 5.55 mM fructose, where equimolar amounts of 1,3-PDL and 3-HP were found. The 3-HPA was transitorily accumulated in the culture medium under almost all culture conditions, the degradation rate being notably slower at 15 degrees C. However, no disappearance of 3-HPA was found at pH 3.6, a usual value in cider making. After sugar exhaustion, L. collinoides 17 oxidated lactic acid and/or mannitol to obtain energy and these oxidations were accompanied by the removal of the toxic 3-HPA increasing the 1,3-PDL, 3-HP and acetic acid contents.

Creatinine, creatine and protein in cooked meat products

The accuracy of the results obtained with a flow injection (FI) system for the simultaneous determination of creatinine and creatine in cooked meat products was evaluated in 30 samples by comparison with those obtained by reference methods. The FI method was then applied to 52 different samples (cooked ham, 14; frankfurters, 11; wieners, 9; chopped, 10; mortadella, 8) and the protein content was also determined. From % creatinine of total creatine (creatine + 1.159 creatinine) an estimation of the heat treatment applied in the processing was made. Cooked ham, mortadella and chopped had the highest values, indicating that the cooking conditions were more severe for these products than those used in the sausages. The ratio of total creatine/protein permitted an estimation of muscular protein to be made. According to the mean value for this ratio, the products were ordered as: cooked ham, 21.9 > chopped, 17.7 > wieners, 15.9 > mortadella, 13.0 > frankfurters, 12.3mg total creatine per gram of protein.

Production and partial characterization of exopolysaccharides produced by two Lactobacillus suebicus strains isolated from cider

Many lactic acid bacteria synthesize extracellular polysaccharides (exopolysaccharides, EPSs) with a large variation in structure and potential functional properties. Although EPS production can produce detrimental effects in alcoholic beverages, these polymers play an important role in the rheological behavior and texture of fermented products. In this work, EPS production by two Lactobacillus suebicus strains, which were isolated from ropy ciders, was examined in a semidefined medium. The existence of priming glycosyltransferase encoding genes was detected by PCR. In addition, the preliminary characterization of the polymers was undertaken. Molecular masses were determined by size exclusion chromatography revealing the presence of two peaks, corresponding to polymers of high- and low-molecular-weight in all fractions. The composition of the EPS fractions was analyzed by gas chromatography-mass spectrometry after acid hydrolysis, revealing that they contained glucose, galactose, N-acetylglucosamine and phosphate, although in different ratios, suggesting that a mixture of polysaccharides is being synthesized. We also examined the influence of the sugar source (glucose, ribose, xylose, or arabinose) and pH conditions on growth and EPS production.

Ripening and Changes in Chemical Composition of Seven Cider Apple Varieties

Changes in composition of seven Basque cider apples (Malus x domestica Borkh.) varieties were studied for 5 weeks before harvesting, while the apples were still on the tree. The evolution of 21 parameters was measured. Special emphasis was made on the sugars, acidity and total phenolic compounds, responsible, respectively, for the sweet, acid and bitter taste of apples and their juices. The whole composition data throughout the ripening process are given. Correlation analysis was used to verify the usefulness of polyphenol index (absorbance at 280nm) for estimating the total phenolic content in apple juices. A high correlation was found between this index and the total phenolic content, either for each apple variety or for all the varieties together, demonstrating the usefulness of polyphenol index to carry out the estimation in these samples. Cluster analysis was used to find patterns of clustering between the apple varieties studied. The apple samples were classified into seven groups, coincident with the seven varieties studied. This proved that the ripening state did not affect this classification, in spite of the changes produced in the composition of apples. Principal component analysis was used to summarise the information of the data matrix in a more reduced way. Acidity (PC1) and phenolic compounds (PC2) were the most relevant parameters for discrimination of apple varieties.

A new sample-injection/sample-dilution system for the flow-injection analytical technique

A new sample-injection/sample-dilution system for the flow-injection analytical technique is proposed. The system is based on two variable-speed computer-controlled peristaltic pumps and a specially designed commutator located at the merging point. One of the pumps delivers a sample and the other a carrier solution. The pumps are run in such a way that the total flow rate after the merging point remains permanently constant. The volume of the injected sample is determined by the time the sample delivering pump is on, whilst the dilution of the sample is determined by relative flow rates of sample and carrier solutions. Two additional possibilities are offered by the proposed system. Firstly, the volume of the injected sample plug can be varied in a simple way, and secondly, for the first time, the injection of a sample plug having an exactly predetermined concentration profile for the measured analyte at the merging point can be carried out. By introducing a third pump delivering a standard solution of the measured species, the system is able to perform a quantitative determination of an analyte, either by the standard addition method or by forming a calibration curve using only one standard solution.

Quantitative determination of carboxylic acids, amino acids, carbohydrates, ethanol and hydroxymethylfurfural in honey by (1)H NMR.

A method using (1)H NMR spectroscopy has been developed to quantify simultaneously thirteen analytes in honeys without previous separation or pre-concentration steps. The method has been successfully applied to determine carboxylic acids (acetic, formic, lactic, malic and succinic acids), amino acids (alanine, phenylalanine, proline and tyrosine), carbohydrates (α- and β-glucose and fructose), ethanol and hydroxymethylfurfural in eucalyptus, heather, lavender, orange blossom, thyme and rosemary honeys. Quantification was performed by using the area of the signal of each analyte in the honey spectra, together with external standards. The regression analysis of the signal area against concentration plots, used for the calibration of each analyte, indicates a good linearity over the concentration ranges found in honeys, with correlation coefficients higher than 0.985 for the thirteen quantified analytes. The recovery studies give values over the 93.7-105.4% range with relative standard deviations lower than 7.4%. Good precision, with relative standard deviations over the range of 0.78-5.21% is obtained.

Simultaneous spectrofluorimetric determination of the rare earths with calcein.

The reactivity of the fluorescent reagent calcein with the trivalent cations of the rare earths has been spectrofluorimetrically studied in aqueous solution. Optimum excitation and emission wavelengths were 492-497 and 519-522 nm, respectively. Optimum pH was in the range 6.0-9.2. The stoichiometry of the complexes was 1:1. A direct, rapid and sensitive method for the determination of rare earth mixtures has been proposed with a detection limit of 4.49x10(-8) M and a coefficient of variation of 0.82%.

Fluorimetric determination of histamine in wine and cider by using an anion-exchange column-FIA system and factorial design study

A study has been performed of the conditions for the reaction of histamine with o-phthaldehyde in a flow injection analysis system employing three channels, using an anion-exchange column to eliminate sample matrix interferences. Factorial design was used to determine which operational parameters should be included in the optimization and their optimal values were found. The method developed shows good selectivity for histamine determination in alcoholic beverages. A linear response of up to 2.0mgl(-1) was observed and the detection and quantification limits were 30 and 101mugl(-1), respectively. The repeatability, measured by the R.S.D. for 10 replicate injections, was 0.84 and 0.52% for histamine solutions of 0.20 and 2.0mgl(-1), respectively. The recoveries obtained in wine and cider samples were close to 100% and a sample frequency of 24 samples per hour was achieved.

Polyphenolic profile in cider and antioxidant power

BACKGROUND The aim of this work was to find the effect of polyphenolic compounds in Basque ciders on the following parameters: antioxidant activity, browning, protein-precipitating capacity, turbidity and reduction potential. These five parameters are highly important, as they affect the taste, the visual aspect and the preservation of cider, and are mainly related to polyphenolic compounds. RESULTS Procyanidin B1 and procyanidin B2 showed a significant positive effect on antioxidant activity. p-Coumaric acid, (-)-epicatechin and hyperin had a significant positive effect on protein-precipitating capacity. Tyrosol had a significant negative effect on reduction potential. CONCLUSION Procyanidin B1 and procyanidin B2 are the most powerful antioxidants in Basque cider, while p-coumaric acid, (-)-epicatechin and hyperin are those with greatest capacity to precipitate proteins. Ciders with higher tyrosol concentration will have less reduction potential and higher antioxidant reservoir.