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Extending storage life of fresh-cut apples using natural products and their derivatives.

Prevention of browning of apples slices has been difficult to achieve because of the rapidity of the enzymatic oxidation of phenolic substrates even under reduced atmospheric pressure storage. Combinations of enzymatic inhibitors, reducing agents, and antimicrobial compounds containing calcium to extend storage life were tested to decrease the browning of Red Delicious apple slices stored at 5 and 10 degrees C under normal atmospheric conditions. Treatments were devised to prevent browning for up to 5 weeks at 5 degrees C with no apparent microbial growth using dipping solutions of compounds derived from natural products consisting of 4-hexylresorcinol, isoascorbic acid, a sulfur-containing amino acid (N-acetylcysteine), and calcium propionate. Analyses of organic acids and the major sugars revealed that the slices treated with the combinations of antibrowning compounds retained higher levels of malic acid and had no deterioration in sugar levels at 5 and 10 degrees C, indicating that higher quality was maintained during storage.

Biological Products↗

Organic acids in post-mortem cerebrospinal fluid.

Organic acids of cerebrospinal fluid (CSF) have been determined by gas chromatography-mass spectrometry in 29 post-mortem samples obtained from infants and 10 samples obtained from hospitalized children, as controls. Though the organic acids profile was similar in the two groups, eight organic acids not observed in CSF from live infants were inconstantly found in post-mortem CSF and for three of them, malic acid, lactyllactic acid and uracile, concentrations were correlated with the delay in sampling.

Acids↗

Reversible enzymic protection of the alpha-amino group of amino acid derivatives using an aminopeptidase A.

In previous papers we have reported that an aminopeptidase A (EC 3.4.11.7) purified from Staphylococcus chromogenes was able to catalyse the introduction of L-malic acid at the N-terminus of Tyr and Phe derivatives. We now show that this enzyme can be used for selective alpha-amino protection of derivatives of probably all amino acids, except Gly and Pro, by the malyl group. The following L-malyl derivatives were synthesized in thermodynamically controlled reactions with yields ranging from 4 to 47%: L-malyl-Tyr-OEt, -ALA-OMe, -Ser-OEt, -Lys-OEt, -Phe-OMe, -Met-NH2, -Glu-MH2, Arg-NH2, -Tye-NH2, -Val-NH2, -Ala-Phe and -Ala-Phe-NH2 (OEt and OMe are ethyl and methyl esters respectively). The reactions were monitored by reverse-phase h.p.l.c.; the products were quantified by amino acid analysis, and their structure was confirmed by m.s. No synthesis was obtained with Gly and Pro derivatives as nucleophiles. The effects of pH, temperature, enzyme concentration, nucleophile concentration, reaction time and addition of an organic co-solvent were studied. An important shift towards synthesis was obtained by carrying out the reactions at 55 degrees C in the presence of 55% organic co-solvent Triglyme (2,4,8,11-tetraoxadodecane) [8-60-fold increase in Ksyn. ([product] [acyl-donor]-1 [nucleophile]-1)].

Amino Acid Sequence↗

Changes in acidity and in proton transport at the tonoplast of grape berries during development.

As in many fruits, the induction of grape berry (Vitis vinifera L.) ripening results in intense breakdown of malic acid. Using membrane fractions, we tested the hypothesis that changes in acidity resulted from malate vacuolar decompartmentation. The hydrolytic activities of the two primary vacuolar pumps inorganic pyrophosphatase (V-PPase; EC 3.6.1.1) and vacuolar ATPase (V-ATPase; EC 3.6.1.3) increased throughout development with an acceleration during ripening, as confirmed by Western blotting and analysis of transcript expression. The ratio of V-PPase activity to V-ATPase activity was always in favour of V-PPase and reached its maximum value at véraison. The rate of anion transport strongly increased during ripening. Before ripening, tonoplast passive permeability was low, but rose during ripening. Our data indicate that tonoplast leakage dramatically increased during ripening. This leakage is probably the prime cause of malate decompartmentation, amplified by the incapacity of oxidative phosphorylation to face increased energy demand.

Adenosine Triphosphate↗

pH-responsive swelling behavior of collagen gels prepared by novel crosslinkers based on naturally derived di- or tricarboxylic acids.

The aim of this study was to compare the physicochemical properties of alkali-treated collagen (AlCol) gels prepared using two kinds of naturally derived crosslinkers made from citric and malic acids (CAD and MAD, respectively) that we have developed. From the crosslinking reaction between active ester groups and amino groups of AlCol, we successfully obtained AlCol gels, named AlCol-CAD and AlCol-MAD, prepared using CAD and MAD, respectively. The gelation time of the AlCol solution containing CAD initially decreased with increasing CAD concentration up to 70 mM, and then increased as the CAD concentration increased further. The gelation time reached its minimum and began to increase. On the other hand, for AlCol-MAD solution, gelation occurred within 40s at any MAD concentration. Moreover, the residual amino groups in AlCol-CAD and AlCol-MAD were found to decrease with increasing CAD or MAD concentrations, whereas increased residual carboxyl groups were detected only in the case of AlCol-CAD. The swelling ratio of AlCol-CAD significantly increased at CAD concentrations above 50mM. On the other hand, AlCol-MAD showed little increase in swelling ratio with increasing MAD concentration. Also, AlCol-CAD was swollen when the gels were immersed in a solution with high pH. On the other hand, no significant increase in swelling ratio was observed when AlCol-MAD was immersed in a similar solution. These results suggest that the different amounts of carboxyl groups in AlCol-CAD affected the swelling behavior of gels and that this pH-responsive AlCol-CAD has potential for drug delivery systems and tissue engineering.

Collagen↗

Carbohydrate metabolism in two apple genotypes that differ in malate accumulation.

In the apple variety 'Usterapfel', there are two known genotypes, which differ in malic acid content. One hundred days after full bloom, low-acid fruit (LA-fruit) contained 125 micromolg(-1) dry matter (DW) of malate, while the high-acid genotype (HA-fruit) reached levels up to 627 micromolg(-1) DW. There was no difference in the catalytic activity of enzymes involved in malate metabolism, such as PEPcarboxylase, malate dehydrogenase, and NADP malic enzyme. After [14C]glucose incorporation into the excised tissue of either genotype, the organic acid fraction was labeled to approximately the same extent. Furthermore, uptake of [14C]malate was significantly lower in excised tissue of LA-fruit. These findings suggest that low malate content in LA-fruit is the result of a restricted ability to accumulate malate in apple parenchyma cells. The different ability to accumulate malate had a pronounced effect on overall carbon partitioning. However, the rate of respiration and the rate of malate synthesis was similar in both genotypes. In HA-fruit, the glycolytic flux through pyruvate kinase was increased to compensate for the carbon that accumulated in the vacuole as malate. Since malate storage in the LA-fruit was restricted, it was more easily available for gluconeogenesis, and was correlated with a three-times higher activity of PEPcarboxykinase. LA-fruit showed higher concentrations of ATP, which stimulated Glc6P and fructose-6-phosphate formation. The elevated hexosephosphate content led to an enhanced partitioning of carbon into starch (+40%), hemicellulose (+104%), and sucrose (+40%) in more mature fruit. The activation of carbohydrate synthesis resulted in a significant drop in glucose-1-phosphate (Glc1P). To meet the increased demand for Glc1P, the activities of neutral and acid invertase, hexokinase, and phosphoglucomutase were higher in LA-fruit. Glucose was a more versatile substrate for this metabolic route than was fructose. It was also evident that glycolytic flux in apple was dependent on glucose level, and that the reaction catalysed by phosphoglucomutase contributed to the regulation of carbon partitioning between malate and carbohydrate polymers.

Carbohydrate Metabolism↗

Uptake and transformation of pesticide metabolites by duckweed (Lemna gibba).

Uptake and transformation of 14C-labeled metabolites from several pesticides, 3-methyl-4-nitrophenol (1), 3,5-dichloroaniline (2), 3-phenoxybenzoic acid (3), (R,S)-2-(4-chlorophenyl)-3-methylbutanoic acid (4), and (1RS)-trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropanecarboxylic acid (5), were examined by using duckweed (Lemna gibba) in Hoagland's medium. More uptake into duckweed from the exposure water at pH 7.0 was observed for non-ionized 1 and 2 than for 3-5 in an ionized form, and their hydrophobicity accounted for these differences. While carboxylic acids 4 and 5 were scarcely transformed in duckweed, 1-3 mainly underwent phase II conjugation with glucose for 1 and 2, malic acid for 3, glutamic acid for 2, and malonylglucose for 3, the chemical identities of which were confirmed by various spectrometric analyses (LC-MS, LC-MS/MS, and NMR) and/or HPLC cochromatography with reference synthetic standards.

Araceae↗

The Relation between Photosynthesis, Respiration, and Crassulacean Acid Metabolism in Leaf Slices of Aloe arborescens Mill.

Leaves and leaf slices from Aloe arborescens Mill. were used to study the interrelations between Crassulacean acid metabolism, photosynthesis, and respiration. Oxygen exchange of leaf slices was measured polarographically. It was found that the photosynthetic utilization of stored malic acid resulted in a net evolution of oxygen. This oxygen production, and the decrease in acid content of the leaf tissue, were completely inhibited by amytal, although the rate of respiratory oxygen uptake was hardly affected by the presence of this inhibitor of mitochondrial electron transport. Other poisons of respiration (cyanide) and of the tricarboxylic acid cycle (trifluoroacetate, 2-diethyl malonate) also were effective in preventing acid-dependent oxygen evolution. It is concluded that the mobilization of stored acids during light-dependent deacidification of the leaves depends on the operation of the tricarboxylic acid cycle and of the electron transport of the mitochondria.A comparison of enzyme activities in extracts from Aloe leaves and from other plants and studies of leaf anatomy and chloroplast morphology revealed typical characteristics of C(3)-, as well as C(4)-, plants in Aloe.

Journal Article↗

Addition of fumaric acid and sodium benzoate as an alternative method to achieve a 5-log reduction of Escherichia coli O157:H7 populations in apple cider.

A study was conducted to develop a preservative treatment capable of the Food and Drug Administration-mandated 5-log reduction of Escherichia coli O157:H7 populations in apple cider. Unpreserved apple cider was treated with generally recognized as safe acidulants and preservatives before inoculation with E. coli O157:H7 in test tubes and subjected to mild heat treatments (25, 35, and 45 degrees C) followed by refrigerated storage (4 degrees C). Fumaric acid had significant (P < 0.05) bactericidal effect when added to cider at 0.10% (wt/vol) and adjusted to pH 3.3, but citric and malic acid had no effect. Strong linear correlation (R2 = 0.96) between increasing undissociated fumaric acid concentrations and increasing log reductions of E. coli O157:H7 in apple cider indicated the undissociated acid to be the bactericidal form. The treatment that achieved the 5-log reduction in three commercial ciders was the addition of fumaric acid (0.15%, wt/vol) and sodium benzoate (0.05%, wt/vol) followed by holding at 25 degrees C for 6 h before 24 h of refrigeration at 4 degrees C. Subsequent experiments revealed that the same preservatives added to cider in flasks resulted in a more than 5-log reduction in less than 5 and 2 h when held at 25 and 35 degrees C, respectively. The treatment also significantly (P < 0.05) reduced total aerobic counts in commercial ciders to populations less than those of pasteurized and raw ciders from the same source (after 5 and 21 days of refrigerated storage at 4 degrees C, respectively). Sensory evaluation of the same ciders revealed that consumers found the preservative-treated cider to be acceptable.

Anticarcinogenic Agents↗

Changes in physicochemical characteristics and volatile constituents of strawberry (Cv. Cigaline) during maturation.

Changes in the volatile composition of strawberries (cv. Cigaline) at six stages of maturity, from 28 to 44 days after anthesis, were investigated over two harvesting seasons. Volatiles were isolated by direct solvent extraction and analyzed by means of GC-FID and GC-MS, with special attention to the quantification of furanones. Firmness, skin color, soluble solids (SS), titratable acidity (TA), SS/TA ratio, organic acids, and sugars were also determined. With increasing maturity, soluble solids, SS/TA ratio, Minolta a value, and levels of sucrose, glucose, fructose, and malic acid increased, whereas Minolta L value, hue angle (), titratable acidity, and levels of citric acid decreased. Furanones and esters were generally not detected before half-red fruits, whereas C(6) compounds were the main compounds in immature fruits. During maturation, levels of these so-called green components decreased drastically, whereas levels of key flavor compounds increased significantly and were closely correlated with skin color development. Maximum volatile production was observed in fully red fruits.

Chemical Phenomena↗

Effects of monocarboxylic and dicarboxylic acids on myosin ATPase activity tested by luminometric procedure.

L(+)Lactic acid enhances myosin ATPase in vitro. Different organic acids were tested for activation of myosin ATPase activity. L(+)Lactic is more effective in stimulating ATPase than D(-)Lactic. D(+) and L(-)Malic acids were also effective at the concentration of 2.5 x 10(-2)-5.0 x 10(-2) mmoles/l. At 3.0 x 10(-2) mmoles/l concentration the following acids are activators: acetic, oxalic, malonic, oxaloacetic, pyruvic, glyoxylic, glycolic; succinic is an inhibitor and acetoacetic is without effect. The activation is not in relation with the pKa of these acids. The inhibitory effects of organic acids are evident at the concentration of 5.0 x 10(-2) mmoles/l. This inhibitory effect is linearly increasing with their pKa. The results are discussed in connection with the possible role of these metabolites in controlling not only ATPase activity towards splitting of ATP, but also in controlling the removal of its hydrolytic products.

Animals↗

Atmospheric carbonaceous aerosols over grasslands of central Europe and a Boreal forest.

A labour-intensive analytical technique was applied to atmospheric particulate matter samples collected in a German urban/industrial influenced grassland location (Melpitz) and in a Finnish forest area (Hyytiälä) in order to achieve a detailed chemical speciation of the organic content. The representative nature of the solvent and water-extractable fractions was determined. The organic compounds identified in the solvent extracts are represented by primary compounds with both anthropogenic and biogenic origin, which mainly derive from the vegetation waxes and from petrogenic sources. Secondary products resulting from the oxidation of volatile organic compounds were also detected. The German meadow presented the highest levels of sugars and acidic compounds in the water extracts, whilst polyols were the most abundant class in the Finnish forest. The major compounds of these classes were malic acid, mannitol, arabitol, glucose and sucrose. Levoglucosan was also found in the water extract.

Aerosols↗

rpoB gene: a target for identification of LAB cocci by PCR-DGGE and melting curves analyses in real time PCR.

Lactic acid bacteria (LAB) are essential in the quality of many fermented beverages like beer, cider and wine. In the two later cases, they convert malic acid into lactic acid during the malolactic fermentation. After fermentation, microbial stabilization is needed to prevent the development of spoilage bacteria species. Among them, cocci lead to different alterations: Pediococcus sp., and some strains of Leuconostoc mesenteroides and Oenococcus oeni can produce exopolysaccharides which modify wine viscosity and lead to ropiness. They also can produce acetic acid, biogenic amine, ethyl carbamate and volatile phenols. Therefore detection and identification are crucial. Results of phenotypic tests and DNA-DNA probes are not accurate enough. 16S RNA gene which is currently used for bacterial species identification presents intraspecies heterogeneity. The rpoB gene is an alternative to this limitation. However previous PCR targeting partial sequence of rpoB gene could not delimit cocci species. Therefore we compared the rpoB gene sequence of the six main cocci species found in fermented beverages: P. damnosus, P. dextrinicus, P. parvulus, P. pentosaceus, L. mesenteroides and O. oeni. The most discriminating partial sequence of the rpoB gene was chosen for designing primers. By PCR-DGGE the reliability of these primers was verified. It was controlled in a mixture of several cocci and other lactic acid bacteria (Lactobacillus sp.). Then we adapted the primers and the PCR conditions in order to achieve the identification of cocci species by real time PCR program including the fluorescent dye SYBR Green I, which gives faster results. PCR melt curves were established and a specific T(m) was attributed to each species.

Animals↗

Responses of primate cortical neurons to unitary and binary taste stimuli.

1. The responses of 126 neurons in primary gustatory cortices of two rhesus monkeys were recorded during sapid stimulation of the tongue with 18 taste stimuli. Ten of these stimuli were dissolved in distilled water (DW): 1.0 M sucrose (Suc), 0.1 M and 0.03 M sodium chloride (NaCl), 0.003 M hydrochloric acid (HCl), 0.001 M quinine hydrochloride (QHCl), 0.03 M monosodium glutamate (MSG), 0.03 M polycose, 0.3 M glycine, 0.1 M proline, and 0.1 M malic acid. Seven other stimuli were dissolved in 0.03 M MSG; the last stimulus was a mixture of 1.0 M Suc and 0.03 M NaCl. 2. The average spontaneous rate (2.2 +/- 0.2 spikes/s, mean +/- SE) and response to DW (2.5 +/- 0.2) of these 126 neurons was low but within the range previously reported for neurons in primate taste cortex. Suc was the most effective stimulus for 24.1% of the neurons tested followed by NaCl (15.7%), QHCl (14.8%), HCl (11.1%), MSG (10.2%), and other miscellaneous unitary gustatory stimuli (8.3%). Binary taste mixtures were the most effective stimuli for 15.7% of the sample. The net responses (corrected for DW, in spikes/s) for Suc-best (3.3), NaCl-best (4.3), HCl-best (3.4), QHCl-best (2.3), and MSG-best (4.1) were sluggish, but comparable with that reported previously. 3. The response breadth of the 82 neurons that responded best to either Suc, NaCl, HCl, or QHCl measured with the entropy coefficient indicated a moderate response breadth for these neurons (mean = 0.79; range = 0.30-0.98). According to the response criteria adopted in this experiment (water response +/- 1.96 SD), however, 81 of these 82 neurons (98.1%) responded to only one or two of the four basic taste stimuli. The disparity between the entropy- and criterion-based measures of response derive from the nature of the two statistics. Adjustments that would make the entropy statistic less inclusive and the definition of a response according to statistical criteria less exclusive would increase their concordance. 4. Three multivariate statistics (cluster, principal axis factor, and multidimensional analysis) were used to analyze the data. Cluster analysis enabled us to divide the 82 taste neurons into groups on the basis of response similarity. Each of the four largest groups was dominated by neurons that responded best to one of the four basic taste stimuli: Suc, NaCl, QHCl, and HCl (ranked in descending order); the fifth largest cluster contained neurons that responded best to MSG. Principal axis factor analysis demonstrated that 80.8% of the total variance could be accounted for by three factors. Neurons responding best to Suc, NaCl, and QHCl each were closely associated with one of those three factors, but the loadings of the HCl-best neurons were evenly distributed across all three factors. The communality coefficient of these three factors was > 80% for the Suc-, NaCl-, HCl-, and QHCl-best neurons; the MSG-best neurons, by comparison, had very few high loadings on any of these three factors and a correspondingly low communality coefficient of 40.4%, a difference that was statistically significant from the other four groups. Thus the three factors related to Suc-, NaCl-, HCl-, and QHCl-best neurons are not relevant to MSG-best neurons. We used multidimensional analysis to arrange the neurons that responded best to Suc, NaCl, HCl, QHCl, and MSG into five loosely arranged and partially overlapping clusters. A multidimensional space based on stimulus similarity showed that MSG was as different from the four basic taste stimuli as they were from one another. 5. Mixture suppression, a common observation in human psychophysical experiments, was examined at the neurophysiological level by including binary tastants in the stimulus battery. The average response of 19 Suc-best neurons to 1.0 M Suc (4.1 spikes/s) decreased to near 0 when the solvent was changed from DW to either 0.03 M MSG or 0.03 M NaCl. Similar decrements were observed in NaCl- and MSG-best neurons tested with Suc/NaCl mixtures.

Animals↗

Biodegradation of potato slops from a rural distillery by thermophilic aerobic bacteria.

A study has been made of thermophilic aerobic biodegradation of the liquid fraction of potato slops (distillation residue) from a rural distillery. The COD of this fraction ranged from 49 to 104 g O2/l, the main contributions to the COD coming from organic acids, reducing substances, and glycerol. It was found that biodegradation could be divided into the following stages: organic acids were removed first, followed by reducing substances and glycerol. The extent of removal varied according to the process temperature. At 50 degrees C, acetic and malic acids were removed completely, but the amount of isobutyric acid increased. At 60 degrees C, organic acid removal ranged from 51.2% (isobutyric acid) to 99.6% (lactic acid). Removals of glycerol and reducing substances were 86.2% and 87.4%, respectively. COD reduction was also temperature dependent, the highest removal efficiency (76.7%) being achieved at 60 degrees C. Dissolved oxygen may have limited the biodegradation process, as indicated by the DOT-versus-time profile.

Acetic Acid↗

Flagella-driven chemotaxis towards exudate components is an important trait for tomato root colonization by Pseudomonas fluorescens.

Motility is a major trait for competitive tomato root-tip colonization by Pseudomonas fluorescens. To test the hypothesis that this role of motility is based on chemotaxis toward exudate components, cheA mutants that were defective in flagella-driven chemotaxis but retained motility were constructed in four P. fluorescens strains. After inoculation of seedlings with a 1:1 mixture of wild-type and nonmotile mutants all mutants had a strongly reduced competitive root colonizing ability after 7 days of plant growth, both in a gnotobiotic sand system as well as in nonsterile potting soil. The differences were significant on all root parts and increased from root base to root tip. Significant differences at the root tip could already be detected after 2 to 3 days. These experiments show that chemotaxis is an important competitive colonization trait. The best competitive root-tip colonizer, strain WCS365, was tested for chemotaxis toward tomato root exudate and its major identified components. A chemotactic response was detected toward root exudate, some organic acids, and some amino acids from this exudate but not toward its sugars. Comparison of the minimal concentrations required for a chemotactic response with concentrations estimated for exudates suggested that malic acid and citric acid are among major chemo-attractants for P. fluorescens WCS365 cells in the tomato rhizosphere.

Amino Acids↗

Capillary electrophoresis for evaluating orange juice authenticity: a study on Spanish oranges.

Fruit juices have very distinct organic acid profiles that can be used as fingerprints for establishing possible adulteration. Recently, our group developed and validated a capillary electrophoresis method using UV detection for determining citric, isocitric, tartaric, and malic acids in natural and commercial orange juices. Sample treatment consisted of only dilution and centrifugation or filtration. This method has been applied to evaluate these acids and their ratios in 63 samples of Navelina, the most common variety of Spanish oranges, over a three month period. This evaluation has been conducted to establish ranges of acid concentrations and to compare them with those found in commercial juices. The more reliable parameter, because of the lower variability in fresh samples, was found to be the citrate/isocitrate ratio with a value of 113 (RSD = 10%). Only one of nine ramdonly selected commercial juices presented values within the range of those of the population of just-pressed Navelina orange juice. Moreover, three of them had measurable tartrate values, which is not a natural component of orange juice, showing mixtures with cheaper fruits.

Beverages↗

Induction of a Crassulacean acid like metabolism in the C(4) succulent plant, Portulaca oleracea L.: physiological and morphological changes are accompanied by specific modifications in phosphoenolpyruvate carboxylase.

The induction of a Crassulacean acid like metabolism (CAM) was evidenced after 21-23 days of drought stress in the C(4) succulent plant Portulaca oleracea L. by changes in the CO(2) exchange pattern, in malic acid content and in titratable acidity during the day-night cycle. Light microscopy studies also revealed differences in the leaf structure after the drought treatment. Following the induction of the CAM-like metabolism, the regulatory properties of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31), the enzyme responsible for the diurnal fixation of CO(2) in C(4) plants but nocturnal in CAM plants, were studied. The enzyme from stressed plants showed different kinetic properties with respect to controls, notably its lack of cooperativity, higher sensitivity to L-malate inhibition, higher PEP affinity and lower enzyme content on a protein basis. In both conditions, PEPC's subunit mass was 110 kDa, although changes in the isoelectric point and electrophoretic mobility of the native enzyme were observed. In vivo phosphorylation and native isoelectrofocusing studies indicated variations in the phosphorylation status of the enzyme of samples collected during the night and day, which was clearly different for the control and stressed groups of plants. The results presented suggest that PEPC activity and regulation are modified upon drought stress treatment in a way that allows P. oleracea to perform a CAM-like metabolism.

Journal Article↗