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20 páginas, 8 figuras, 3 tablas, Many lactic acid bacteria (LAB) produce metabolites with applications in the food industry, such as dextran-type exopolysaccharides (EPS) and riboflavin (vitamin B2). Here, 72 bacteria were isolated from sourdoughs made by Spanish bread-makers. In the presence of sucrose, colonies of 22 isolates showed a ropy phenotype, and NMR analysis of their EPS supported that 21 of them were dextran producers. These isolates were identified by their random amplified polymorphic DNA (RAPD) patterns and their rrs and pheS gene sequences as LAB belonging to four species (Weissella cibaria, Leuconostoc citreum, Leuconostoc falkenbergense and Leuconostoc mesenteroides). Six selected strains from the Leuconostoc (3) and Weissella (3) genera grew in the absence of riboflavin and synthesized vitamin B2. The EPS produced by these strains were characterized as dextrans by physicochemical analysis, and the L. citreum polymer showed an unusually high degree of branching. Quantification of the riboflavin and the EPS productions showed that the W. cibaria strains produce the highest levels (585–685 μg/and 6.5–7.4 g/L, respectively). Therefore, these new LAB strains would be good candidates for the development of fermented foods bio-fortified with both dextrans and riboflavin. Moreover, this is the first report of riboflavin and dextran production by L. falkenbergense, This research was funded by the Spanish Ministry of Science, Innovation and Universities, (grants RTI2018-097114-B-I00 and PCIN-2017-075), by the Basque Government Industry and Education Department (grant PIBA_2020_1_0032) and by the University of the Basque Country (General Grant to Research Groups (GIU 19/014))

Transparent and colorless citric acid-incorporated fish gelatin/chitosan composite films were prepared in this work. The addition of citric acid into film forming formulations slowed down the swelling rate and swelling values of the films, which maintained their integrity and resulted in flexible hydrated films. Furthermore, citric acid acted as plasticizer, increasing the elongation at break of the films. Additionally, films showed good UV barrier properties and the incorporation of citric acid and chitosan reduced the E. coli growth, especially for the films prepared with 20 wt % citric acid, highlighting the potential use of these films as active food packaging. These results were related to the changes observed by Fourier transform infrared (FTIR) analysis, which revealed the different protein structure achieved as a function of citric acid and chitosan contents.

Acrolein occasionally appears in cider, completely spoiling its quality due to its bitter taste. It is crucial to detect it in the early steps, before the taste is severely affected, to apply the appropriate treatment. A simple and rapid analytical method to determine this compound in cider is therefore desirable. In this work, a quantitative determination method of acrolein in cider is proposed using the proton nuclear magnetic resonance technique (1H NMR). Acrolein produces a doublet signal in the spectrum at 9.49 ppm, whose area is used to determine the concentration of this compound. Importantly, 3-(Trimethylsilyl)-2,2,3,3-d4-propionic acid sodium salt is added to the cider as a reference for 0.00 ppm and 1,3,5-benzenetricarboxylic acid as an internal standard for acrolein determination. The method is validated by gas chromatography (GC). There is a good correlation between the acrolein concentrations obtained by 1H NMR and by gas chromatography in different commercial ciders (Pearson coefficient 0.9994). The 95% confidence interval for the intercept is 0.15 &plusmn, 0.49 (includes 0) and for the slope is 0.98 &plusmn, 0.03 (includes 1). When applying the paired t test, no significant difference is observed. The proposed method is direct, and no prior derivatization is needed.

Two exopolysaccharide (EPS)-producing lactic acid bacteria (LAB) strains, Liquorilactobacillus (L.) sp CUPV281 and Liquorilactobacillus (L.) mali CUPV271, were isolated from Spanish apple must. Each of the strains produced a dextran, with different branching degrees, to be incorporated into soy protein isolate (SPI) film-forming formulations. Films were prepared by compression molding, a more rapid processing method than solution casting and, thus, with a greater potential for scaling-up production. Thermal analysis showed that SPI and EPS start the degradation process at temperatures above 190 &deg, C, confirming that the compression temperature selected (120 &deg, C) was well below the corresponding degradation temperatures. Resulting films were transparent and homogeneous, as shown by UV-Vis spectroscopy and SEM, indicating the good compatibility between SPI and EPS. Furthermore, FTIR analysis showed that the interactions between SPI and EPS were physical interactions, probably by hydrogen bonding among the polar groups of SPI and EPS. Regarding antifungal/fungistatic activity, LAB strains used in this study showed an inhibitory effect on germination of fungal spores.

The contribution of the individual polyphenols of Basque cider to the antioxidant activity and browning has been determined. Nineteen monovarietal musts were obtained using different varieties of cider apples throughout three seasons. Fermentation of these musts was monitored throughout 4 or 5 samplings, until their transformation into cider. Partial least squares regression was done to relate individual polyphenols with the parameters studied. The polyphenols which contributed most to the antioxidant activity were chlorogenic acid, 4-p-coumaroylquinic acid, (−)-epicatechin, phloretin 2′-O-xyloglucoside and isoquercitrine. The main contributors to browning were 4-p-coumaroylquinic acid and phloretin 2′-O-xyloglucoside. The antioxidant activity of Basque ciders was compared with that of other beverages, concluding that it was similar to that of red wines and greater than that of orange juices.

36 p.-6 fig.-1 tab., Dextrans synthesised by three Leuconostoc mesenteroides strains, isolated from mammalian milks, were studied and compared with dextrans produced by Lc. mesenteroides and Lactobacillus sakei strains isolated from meat products. Size exclusion chromatography coupled with multiangle laser light scattering detection analysis demonstrated that the dextrans have molecular masses between 1.74 × 108 Da and 4.41 × 108 Da. Rheological analysis of aqueous solutions of the polymer revealed that all had a pseudoplastic behaviour under shear conditions and a random, and flexible, coil structure. The dextrans showed at shear zero a difference in viscosity, which increased as the concentration increased. Also, the purified dextrans were able to immunomodulate in vitro human macrophages, partially counteracting the inflammatory effect of Escherichia coli O111:B4 lipopolysaccharide. During prolonged incubation on a solid medium containing sucrose, dextran-producing bacteria showed two distinct phenotypes not related to the genus or species to which they belonged. Colonies of Lc. mesenteroides CM9 from milk and Lb. sakei MN1 from meat formed stable and compact mucoid colonies, whereas the colonies of the other three Leuconostoc strains became diffuse after 72 h. This differential behaviour was also observed in the ability of the corresponding strains to bind to Caco-2 cells. Strains forming compact mucoid colonies showed a high level of adhesion when grown in the presence of glucose, which decreased in the presence of sucrose (the condition required for dextran synthesis). However no influence of the carbon source was detected for the adhesion ability of the other Lc. mesenteroides strains, which showed variable levels of binding to the enterocytes., This work was supported by the Spanish Ministry of Economy and Competitiveness (grant AGL2015-65010-C3-1-R).

Ropy strains of heterolactic Lactobacillus spp. were isolated from bottled spoiled ciders produced in the Basque Country. The isolates were able to produce slime experimentally on glucose containing medium and apple must. The production of ropy slime occurred during exponential growth and the increase in viscosity was correlated with production of polysaccharide. They were characterized with regards to growth at different pH, sulfur dioxide (SO2), ethanol and temperatures and showed similar behavior. All ropy strains grew in the presence of high ethanol concentrations (15%) and at low temperatures (10 and 15°C). In addition, these were tolerant to 10, 25 and 50 mg/l of total SO2 (pH 3.8), but with longer lag times as the concentration of SO2, increased. However, the ropy lactobacilli studied were affected by the pH of the medium and in general pH below 3.7 limited their growth seriously.

Bifidobacterium longum INIA P132 and Bifidobacterium infantis INIA P731, isolated from infant-faeces, were investigated in this work. Regarding the probiotic and technological potential of the bifidobacteria, both were resistant to gastrointestinal tract simulated conditions. B. longum showed high survival upon freezing and thawing as well as lyophilisation and was able to grow in milk. B. infantis had higher adhesion capacity to human Caco-2 cells than the commercial probiotic Bifidobacterium animalis BB12 strain. Moreover, both bacteria secrete heteropolysaccharides (HePS) composed of rhamnose, galactose and glucose. In a dextran sodium sulphate-induced enterocolitis model in zebra fish larvae, treatment with each HePS preparation resulted in a decrease of the larval mortality. In addition, the HePS from B. longum immunomodulated in vitro human macrophages treated with the inflammatory Escherichia coli O111:B4 lipopolysaccharide. Thus, both studied bifidobacteria and their HePS have potential beneficial effects on health and thus, to their application in functional foods., This work was supported by a grant from de Department of Environment, Territorial Planification, Agriculture and Fishing and the project no. IT 866-13 from the Basque Government, by the Spanish Ministry of Science and Innovation (AGL 2015-65010-C3-1-R) and by the Spanish Ministry of Economy and Competitiveness (RM12-00004-00-00 and RTA2013-00029-00-00).

Chitosan films with antioxidant and antimicrobial properties were prepared by the incorporation of gallic acid as a bioactive. The films obtained were homogeneous, transparent and colorless with high mechanical resistance. Results indicated that films had antimicrobial activity against E. coli, especially those films plasticized with 15 wt % glycerol and containing 20 wt % gallic acid, which showed the biggest inhibition halo. Additionally, gallic acid-incorporated chitosan films exhibited a great antioxidant activity with DPPH scavenging capacity values of 99% and a bioactive release of 33% after 4 days, suggesting the potential suitability of these modified chitosan films as active films for food or pharmaceutical purposes.

24 p.-2 fig.-2 tab.-1 fig. supl.-1 tab.supl., Lactic acid bacteria (LAB) are naturally occurring bacterial populations in traditional cider from the Basque Country (Spain), which is spontaneously fermented without starters. Some LAB are able to produce exopolysaccharides (EPS) thus causing viscosity, a cider spoilage called “ropiness” or “oilness”. A total of 41 isolates recovered between 1992 and 2009 from ropy ciders of different cider factories were included in the present study. Identification by 16S rRNA gene sequencing, biochemical traits (API), RAPD-PCR using four universal primers (M13, M13V, P1 and P2) and 16S-23S ISRs amplification profiles revealed that the spoiler species were L. collinoides, L. diolivorans, L. sicerae and L. suebicus. EPS production in L. collinoides and L. sicerae strains approached by PCR amplification of eps genes encoding for homopolysaccharide and heteropolysaccharide revealed that these strains synthesize heteropolysacccharides and in addition, L. collinoides CUPV231 synthesizes a 2-branched (1,3)-β-D-glucan. The molecular weight of their EPS determined by HP-SEC revealed the presence of three polymers with average molecular weights (MW) of around 106 g/mol, 104 g/mol and 103 g/mol. The middle-weight fraction was the most abundant except in L. sicerae CUPV261 which mainly produced the high Mw polymer., This study was supported by grants AGL2012-40084-C03 (co-funded by FEDER), AGL2015-65010-C3-1-R, IT866-13 from the Basque Government and PROMETEO/2012/040 from the Generalitat Valenciana.

A quantitative determination method of lactic and acetic acids in cider is proposed by means of the proton nuclear magnetic resonance (1H NMR) technique. Lactic acid gives a doublet signal at 1.42 ppm in the spectrum and acetic acid a singlet signal at 2.09 ppm, whose area is used to determine the concentration of these compounds. 3-(Trimethylsilyl)-2,2,3,3-d4-propionic acid sodium salt is added to the cider with a double purpose: as a reference for 0.00 ppm and as an internal standard for 1H NMR measurement. Enzymatic analysis methods are used to validate the method. Good correlations are found between the lactic and acetic acids concentrations obtained by 1H NMR and those obtained by enzymatic methods in different commercial ciders. For lactic acid, the 95% confidence interval for the slope is 1.05 ± 0.08 (includes 1) and for the intercept 0.27 ± 0.42 (includes 0). For acetic acid, these intervals are 1.01 ± 0.06 for the slope and 0.03 ± 0.08 for the intercept. The described method is fast and direct.

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. © 2014 Society of Chemical Industry

30 p.-1 tab.-5 fig.-2 fig. supl., Exopolysaccharides synthesized by lactic acid bacteria have prebiotic properties and contribute to the rheology and texture of fermented foods. Here, we have standardized an immunological method for the specific detection of 2-substituted (1,3)β-d-glucans. The method allows direct detection and quantification of this exopolysaccharide in culture supernatants containing other mono- and poly-saccharides. Moreover, it allows specific detection of the biomolecules synthesized in vitro in enzymatic reactions. Thus, this method allows the fast identification of producing bacteria, as well as biochemical characterization of the glycosyltransferases responsible for their synthesis., This work was supported by the Spanish Ministry of Economy and Competiveness grants AGL2009-12998 and AGL2012-40084.

1p.International Symposium on Immunonutrition 2017 Madrid, 17th–19th July, 2017 10th Anniversary

Chitosan-based films for food packaging applications were prepared by casting and dried at room temperature or heat-treated in order to study functional properties and antimicrobial activity. In all cases, films were flexible and transparent, regardless of chitosan molecular weight, glycerol content, and temperature. Regarding antimicrobial activity, chitosan film forming solutions showed antimicrobial behaviour against Escherichia coli and Lactobacillus plantarum. It was also observed that the bacteriostatic property of chitosan-based films against bacteria employed in this study was notably affected by temperature. Moreover, temperature produced significant variation in the functional properties of chitosan-based films, such as colour, wettability, resistance against UV light and mechanical properties. In good agreement with this behaviour, total soluble matter (TSM), fourier transform infrared (FTIR) spectroscopy, thermo-gravimetric analysis (TGA) and X-ray diffraction (XRD) results suggested a change in the chemical structure of chitosan films, possibly due to Maillard reaction when heat treatment was used.

Lactobacillus collinoides CUPV237 is a strain isolated from a Basque cider. Lactobacillus collinoides is one of the most frequent species found in cider from Spain, France, or England. A notable feature of the L. collinoides CUPV237 strain is its ability to produce exopolysaccharides.

28 p.-5 fig, Fermented oat-based foods offer attractive prospects within the market of non-dairy functional products, since they are suitable substrates for the delivery of probiotic microorganisms, and are significant sources of dietary fiber, both insoluble and soluble such as β-glucan, good quality fat and other phytochemicals important for human health., In the present work, whole oat flour was fermented with probiotic Lactobacillus plantarum strains to produce new functional foods with improved nutritional and technological features. Viability of the probiotic and the main technological, physico-chemical, nutritional and sensorial parameters were monitored at 7, 14 and 21 days of cold storage. The microbial survival was higher than 5x108 cfu g−1 at the end of the shelf life. After the fermentation step, viscosity was higher in products inoculated with the exopolysaccharide-producing L. plantarum strain Lp90. However, a subsequent viscosity reduction was detected in all the samples throughout the storage period, consistent with the observed concentration decrease of the oat β-glucan. Vitamin B2 content was about two-fold higher in products fermented by the riboflavin-overproducing LpB2, and in these samples the riboflavin concentration further increased during cold storage., P.L., G.S. and M.T. are supported by the Spanish Ministry of Economy and Competitiveness grant AGL2012-40084-C03.

14 páginas, 4 figuras -- PAGS nros. 6026-6039, Probiotics, prebiotics and synbiotics are frequently-used components for the elaboration of functional food. Currently, most of the commercialized probiotics are limited to a few strains of the genera Bifidobacteria, Lactobacillus and Streptococcus, most of which produce exopolysaccharides (EPS). This suggests that the beneficial properties of these microorganisms may be related to the biological activities of these biopolymers. In this work we report that a 2-substituted-(1,3)-β-D-glucan of non-dairy bacterial origin has a prebiotic effect on three probiotic strains. Moreover, the presence of this β-D-glucan potentiates in vitro adhesion of the probiotic Lactobacillus plantarum WCFS1 to human intestinal epithelial cells., This work was funded by the Italian Ministry for the Development in the framework of the projects named ―Industria 2015, Bando Nuove Tecnologie per il Made in Italy—Realizzazione di una innovativa pasta alimentare funzionale arricchita di componenti bioattivi e probiotici‖ and ―Pro. Ali.Fun.‖ as well as by the Spanish Ministery of Innovation and Science grant AGL2009-12998-C03, We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI).

The ability to produce a 2-branched (1,3)-beta-D-glucan was screened in 147 lactic acid bacteria strains recovered from cider. Among them, 32 identified as Pediococcus parvulus exhibited a ropy character and were PCR positive for the presence of the gtf gene, related to the synthesis of the beta-glucan. Half of the strains produced more than 100 mg L(-1) of exopolysaccharide (EPS). (1)H NMR spectra of the crude EPSs were identical to that previously described for P. parvulus 2.6, indicating that all are 2-branched (1,3)-beta-D-glucans. The EPSs from two of the isolates were subjected to acid hydrolysis and methylation analysis, confirming the NMR results. Size exclusion chromatography (SEC) showed in all crude EPSs the presence of two different molecular mass fractions of about 10(7) Da and 10(4) Da, whose relative proportions varied among strains. EPS amounts and concentrations of high molecular fraction are linearly correlated. Intraspecific diversity of isolates was determined by RAPD profiles. Based on genotypic and phenotypic characteristics, two strains were selected to be further studied as probiotics.

Some physicochemical and rheological properties of the exopolysaccharide (EPS) produced by Pediococcus parvulus 2.6 were examined. Structural characterization by NMR ((1)H and 2D-COSY) showed that the same EPS, a 2-substituted (1,3)-beta-D-glucan, was synthesized irrespective of sugar source used for growth (glucose, fructose, or maltose). The molecular masses of these beta-glucans were always very high (10(6) Da) and influenced by the culture medium or sugar source. The steady shear rheological experiments showed that all concentrations of the beta-glucan aqueous solutions exhibited a pseudoplastic behavior at high shear rates. Viscoelastic behavior of beta-glucan solutions was determined by dynamic oscillatory analysis. A critical concentration of 0.35% associated with the appearance of entanglements was calculated. The beta-glucan adopts an ordered hydrogen bond dependent helical conformation in neutral and slightly alkaline aqueous solutions, which was partly denatured under more alkaline conditions.

Background. In Mexico and other developing countries, few reports of the survival of children with acute leukaemia exist.Objective. We aimed at comparing the disease-free survival of children with acute myeloid leukaemia who, in addition to being treated with the Latin American protocol of chemotherapy and an autologous transplant, either underwent early intensified chemotherapy or did not undergo such treatment.Procedure. This was a cohort study with a historical control group, forty patients, less than 16 years old. Group A (20 patients), diagnosed in the period 2005–2007, was treated with the Latin American protocol of chemotherapy with an autologous transplant plus early intensified chemotherapy: high doses of cytarabine and mitoxantrone. Group B (20 patients), diagnosed in the period 1999–2004, was treated as Group A, but without the early intensified chemotherapy.Results. Relapse-free survival for Group A was 90% whereas that for Group B it was 60% (P=0.041). Overall survival for Group A (18, 90%) was higher than that for Group B (60%). Complete remission continued for two years of follow-up.Conclusions. Relapse-free survival for paediatric patients treated with the Latin American protocol of chemotherapy with an autologous transplant plus early intensified chemotherapy was higher than that for those who did not receive early intensified chemotherapy.

Our aim in this paper is to describe the results of treatment of acute lymphoblastic leukaemia (ALL) in Mexican children treated from 2006 to 2010 under the protocol from the Dana-Farber Cancer Institute (DFCI) 00-01. The children were younger than 16 years of age and had a diagnosis of ALL denovo. The patients were classified as standard risk if they were 1–9.9 years old and had a leucocyte count 9/L, precursor B cell immunophenotype, no mediastinal mass, CSF free of blasts, and a good response to prednisone. The rest of the patients were defined as high risk. Of a total of 302 children, 51.7% were at high risk. The global survival rate was 63.9%, and the event-free survival rate was 52.3% after an average follow-up of 3.9 years. The percentages of patients who died were 7% on induction and 14.2% in complete remission; death was associated mainly with infection (21.5%). The relapse rate was 26.2%. The main factor associated with the occurrence of an event was a leucocyte count >100 × 109/L. The poor outcomes were associated with toxic death during induction, complete remission, and relapse. These factors remain the main obstacles to the success of this treatment in our population.

Biogenic amines play an important physiological role in mammals, and high amounts of some exogenous amines in human diet may contribute to a wide variety of toxic effects. These amines are commonly found in many foodstuffs, particularly in fermented products such as cheese, meat products, beer, wine, and ciders. Here, the level of biogenic amines in some natural ciders was examined. Twenty-four samples of cider purchased from commercial sources were analyzed by reverse-phase high-performance liquid chromatography and fluorescence detection after precolumn derivatization with o-phthaldialdehyde. Amine levels were variable, ranging from not detected to 23 mg/liter. The average level of total biogenic amines in ciders was 5.94 +/- 8.42 mg/liter. Putrescine, histamine, and tyramine were the prevailing amines being present in 50.0, 37.5, and 33.3% of the ciders studied; very small amounts of ethylamine and phenylethylamine were observed in only one sample. Other cider parameters were analyzed to determine whether they affect the biogenic amine content in ciders, and the results were evaluated by applying cluster analysis and principal component analysis. Ciders that showed lower glycerol contents and higher amounts of 1,3-propanediol had much higher levels of histamine, tyramine, and putrescine, suggesting a high activity of lactic acid bacteria during cider making and thus the need for effective control of lactic acid bacteria.

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.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.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.

The thermodegradative behavior of an exopolysaccharide (EPS), produced by Pediococcus damnosus 2.6 from a strain isolated from a ropy Basque Country cider, was studied. Two endothermic processes at 118 and 163°C were detected by differential scanning calorimetry and considered as sub and main glass transitions, respectively, for the EPS. Dynamic thermogravimetry experiments showed: a first weight loss, below 100°C, due to alcohol molecules trapped in the structure of the EPS, a main degradation process involving both dehydration reactions, which yield levoglucosan structures, and scissions of C–O bonds, either in the inner glucan rings or between rings, going on with depolymerization reactions up to 350°C, and char-forming reactions at higher temperatures. From isothermal degradation processes, at short reaction time, an activation energy of 33 kJ/mol was determined. This value changed to 50.4 kJ/mol for conversion ranges of 20–35%, rising to 138 kJ/mol at 50% (determined by dynamic methods). Analyzing th...

Aplastic anemia (AA) is a hematological disease characterized by the deficient production of blood cells. The incidence of AA worldwide is low (1-5 new cases per 10(6) individuals per year). In contrast to other countries, no current reports exist on the incidence of this disorder in Mexico. In the present study, we have determined the incidence of AA in a defined subpopulation from Mexico City during the period 1996-2000. For the purpose of this study, we focused on the experience from a single medical institution: the Mexican Institute of Social Security (IMSS), which covers around 50% of Mexico's population. The incidence of AA was determined based on the actual number of patients diagnosed with this disease at the IMSS in Mexico City in a given year and the total number of individuals registered at the IMSS in Mexico City in the same year. Considering the IMSS population as a whole, the annual incidence of AA was 3.9 new cases per 10(6) individuals per year. In the pediatric population, the annual incidence was 4.2 new cases per 10(6) individuals per year, whereas in people 15-years-old and older the incidence was 3.8 new cases per 10(6) individuals per year. These incidences were higher than those reported in most studies from the USA, Europe and Israel. Compared to the incidence in Thailand, the incidence we observed in children was considerably higher, whereas the one in adults was similar to the one in that country. The results of the present study suggest that the incidence of AA in Mexico City is one of the highest worldwide, particularly in terms of the pediatric population; however, these results must be taken with caution since this study comprises only a subpopulation from Mexico City and not the entire population. Thus, further studies including a broader population, both in Mexico City and other urban and rural areas of this country, will be necessary in order to obtain better and more complete estimates of the actual incidence of AA in Mexico.

29 p.-4 fig., Lactobacillus fermentum isolated from sourdough was able to produce riboflavin. Spontaneous roseoflavinresistant mutants were obtained by exposing the wild strain (named L. fermentum PBCC11) to increasing concentrations of roseoflavin. Fifteen spontaneous roseoflavin-resistant mutants were isolated, and the level of vitamin B2 was quantified by HPLC. Seven mutant strains produced concentrations of vitamin B2 higher than 1 mg L−1. Interestingly, three mutants were unable to overproduce riboflavin even though they were able to withstand the selective pressure of roseoflavin.Alignment of the rib leader region of PBCC11 and its derivatives showed only point mutations at two neighboring locations of the RFN element. In particular, the highest riboflavinproducing isolates possess an A to G mutation at position 240,while the lowest riboflavin producer carries a T to A substitution at position 236. No mutations were detected in the derivative strains that did not have an overproducing phenotype. The best riboflavin overproducing strain, named L. fermentum PBCC11.5, and its parental strain were used to fortify bread. The effect of two different periods of fermentation on the riboflavin level was compared. Bread produced using the coinoculum yeast and L. fermentum PBCC11.5 led to an approximately twofold increase of final vitamin B2 content., This work was founded by the Italian Ministry for Development in the framework of the “Industria 2015 Bando Nuove Tecnologie per il Made in Italy - Realizzazione di una innovativa pasta alimentare funzionale arricchita di componenti bioattivi e probiotici” and by the Spanish Ministry of Economy and Competitiveness grants AGL2012-40084-C03-01, AGL2012-40084-C03-03, and IT886-13 from the Basque Government.

We reported the experience of hemophilia camps which was accomplished with patients from hospitals of the Instituto Mexicano del Seguro Social. The aim was to prepare the families and patients regarding the disease treatment, in order to promote the self sufficiency and to know the impact of the program on the course of the disease. Surveys were applied about treatment items and personal opinions were collected. The results of the national hemophilia camp were: group of 56 patients, average 14 years, 2 % women, 51 % severe hemophilia and 43 % had hemophilic brothers. Benefits: patients increased their knowledge about earlier bleeding identification and the self-infusion method; they became aware on their responsibility in self care, timely treatment and duties at home. Hemophilia camps with patients are an option for attitude change before disease complications. Social network creation and the increase in self-sufficiency are other benefits.Se informa la experiencia con los campamentos de hemofilia que incluyen a pacientes de un hospital del Instituto Mexicano del Seguro Social en la Ciudad de México y de otros hospitales en el país, realizados con el objetivo de capacitar a los enfermos y a sus familiares en los cuidados de la enfermedad y de promover la autosuficiencia. Se aplicaron cuestionarios sobre los temas tratados y se recabaron las opiniones personales. De la experiencia con el primer campamento nacional de hemofilia se observó que en el grupo de 56 pacientes (promedio de edad de 14 años, 2 % mujeres, 51 % con hemofilia severa, 43 % con hermanos con hemofilia) se incrementaron los conocimientos en la identificación de los sangrados y la técnica de autoinfusión. De igual forma, los pacientes se concientizaron acerca de su responsabilidad en el autocuidado, el tratamiento oportuno y los deberes en el hogar. De tal forma, se señala que los campamentos son una opción para el cambio de actitud ante la enfermedad, para crear redes sociales y favorecer la autosuficiencia.

"Se informa la experiencia con los campamentos de hemo fi lia que incluyen a pacientes de un hospital del Instituto Mexicano del Seguro Social en la Ciudad de México y de otros hospitales en el país, realizados con el objetivo de capacitar a los enfermos y a sus familiares en los cuidados de la enfermedad y de promover la auto- su fi ciencia. Se aplicaron cuestionarios sobre los temas tratados y se recabaron las opiniones personales. De la experiencia con el primer campamento nacional de hemo fi lia se observó que en el grupo de 56 pacientes (promedio de edad de 14 años, 2 % mujeres, 51 % con hemo fi lia severa, 43 % con hermanos con hemo fi lia) se incrementaron los conocimientos en la identi fi cación de los sangrados y la técnica de autoinfusión. De igual forma, los pacientes se concientizaron acerca de su responsabilidad en el autocuidado, el tratamiento opor- tuno y los deberes en el hogar. De tal forma, se señala que los campamentos son una opción para el cambio de actitud ante la enfermedad, para crear redes socia- les y favorecer la autosu fi ciencia."

The evolution of biogenic amines and other nitrogen compounds during the elaboration period of natural ciders in two successive seasons and two types of presses was studied. The harvest year affects the concentrations of most of the amino acids, while few of them were affected by the type of press. Asparagine and aspartic acid were the most abundant amino acids in fresh musts followed by α-alanine, α-aminobutyric acid, glutamine and glutamic acid. The mean concentrations of these amino acids in the fresh musts from the two harvest years were 12.35, 11.12, 6.45, 6.29, 5.28 and 4.87 mg L−1, respectively. During cidermaking, a decrease in the sum of all amino acids was detected (from 128 ± 54.311 mg L−1 to 22.5 ± 4.863 mg L−1 in 2005 and from 72.2 ± 15.256 mg L−1 to 6.5 ± 4.112 mg L−1 in 2006). Furthermore, significant differences in the concentration of most amino acids related to harvest year were observed. Concerning the biogenic amine content, putrescine was the main and the only amine present in all musts (3.72 ± 1.68 mg L−1) and ciders (3.59 ± 1.83 mg L−1). Histamine was the second biogenic amine of quantitative importance in final ciders (1.15 ± 0.69 mg L−1), and tyramine was only detected in one of the cidermaker cellars at the end of the elaboration period. The low concentrations of biogenic amines in ciders were attributed to the low contents of the corresponding precursor amino acids and do not affect to cider quality, This study was supported by the Spanish Ministerio de Ciencia e Innovación (Projects, PET2007-0134 and AGL2006n-04514). The work at the UPV was partially supported by the Diputación de Gipuzkoa, Programa Red Gipuzkoana de Ciencia, Tecnología e Innovación (co-financed by the European Union) and UPV/EHU (EHU06/93) and Gobierno Vasco (IT335-10). Gaizka Garai-Ibabe acknowledges the Gobierno Vasco (Dpto. Agricultura, Pesca y Alimentación) for the pre-doctoral fellowship.

Wheat contains various essential nutrients including the B group of vitamins. However, B group vitamins, normally present in cereals-derived products, are easily removed or destroyed during milling, food processing or cooking. Lactic acid bacteria (LAB) are widely used as starter cultures for the fermentation of a large variety of foods and can improve the safety, shelf life, nutritional value, flavor and overall quality of the fermented products. In this regard, the identification and application of strains delivering health-promoting compounds is a fascinating field. Besides their key role in food fermentations, several LAB found in the gastrointestinal tract of humans and animals are commercially used as probiotics and possess generally recognized as safe status. LAB are usually auxotrophic for several vitamins although certain strains of LAB have the capability to synthesize water-soluble vitamins such as those included in the B group. In recent years, a number of biotechnological processes have been explored to perform a more economical and sustainable vitamin production than that obtained via chemical synthesis. This review article will briefly report the current knowledge on lactic acid bacteria synthesis of vitamins B2, B11 and B12 and the potential strategies to increase B-group vitamin content in cereals-based products, where vitamins-producing LAB have been leading to the elaboration of novel fermented functional foods. In addition, the use of genetic strategies to increase vitamin production or to create novel vitamin-producing strains will be also discussed.

The evolution of yeasts and lactic acid bacteria was examined during two methods of cider making: the traditional employed unwashed apples of different varieties and the modified used a sole washed acidic apple variety with temperature control during the cider making process. The alcoholic fermentation was carried out mainly by Kloeckera apiculata and Saccharomyces cerevisiae species, and their distributions were similar in the two methods. In the traditional, the malolactic fermentation proceeded at the same time as alcoholic fermentation and the «piqure lactique» occurred. In the modified method, no malolactic fermentation took place. In all musts, a fundamentally heterofermentative lactic flora was found, with Lactobacillus species most abundant in the traditional process

28 p.-1 fig.-4 tab., Ropiness in natural cider is a relatively frequent alteration, mainly found after bottling, leading to consumer rejection. It is derived from the production of exopolysaccharides (EPS) by some lactic acid bacteria most of which synthesize a 2-branched (1,3)-β-D-glucan and belong to the genera Pediococcus, Lactobacillus and Oenococcus. This polysaccharide synthesis is controlled by a single transmembrane glycosyltransferase (GTF). In this work, a method based on quantitative PCR (qPCR) and targeting the gtf gene was developed for detection and quantification of these bacteria in cider. The newly designed primers GTF3/GTF4 delimit a 151 bp fragment within the 417 bp amplicon previously designed for conventional PCR. The inclusivity and exclusivity of the qPCR assay were assessed with 33 cider isolates belonging to genus Lactobacillus, Oenoccocus and Pedioccocus, together with reference strains of 16 species and five genera including β-glucan,α-glucan and heteropolysaccharide (HePS) producing strains and non-EPS producers. The qPCR assay,followed by the melting curve analysis, confirmed the generation of a single PCR product from the β-glucan producers with a Tm of 74.28± 0.08 and CT values (10 ng DNA) ranging between 8.46 and 16.88 (average 12.67± 3.5). Some EPS− LAB strains rendered CT values ranging from 28.04 to 37.75 but they were significantly higher (P(CTb28.54)= 0.05) than those of the β-glucan producers. The assay showed a wide quantification range of 5 log units using calibrated cell suspensions of Pediococcus parvulus 2.6 and Oenococcus oeni I4. The linearity was extended over 7 log orders when calibration curves were obtained from DNA. The detection limit for β-glucan producing LAB in artificially contaminated cider was about 3× 102 CFU per ml. The newly developed qPCR assay was successfully applied to monitor the cidermaking process, in 13 tanks from two cider factories, revealing a decrease in CT values derived from an increase in β-glucan producing LAB populations. In addition, 8 naturally spoiled bottled cider were tested for the quantification of these organisms using the five standard curves constructed: P. parvulus 2.6 genomic DNA and gtf amplicon (417 bp), calibrated cell suspensions of Pediococcus parvulus 2.6, Lactobacillus diolivorans G77 and Oenococcus oeni I4 and results were compared to LAB total counts on MRS. Levels obtained from the different approaches were within a log range and showed no significant differences. Therefore, the amplicon-derived standard curve is proposed for the routine estimation of gtf+populations in cider., This work was supported by the Ministerio de Educación y Ciencia (projects AGL2006-11932 and CSD2007-00063), Universidad del País Vasco (UPV/EHU) (EHU08/37) and the Diputación Foral de Gipuzkoa and the Generalitat Valenciana (GVACOMP2009-257). Programa Red Gipuzkoana de Ciencia, Tecnología e Innovación (co-financed by the European Union).

35 p.-4 fig.-2 fig. supl., We have isolated three lactic acid bacteria (Lactobacillus suebicus CUPV221, Pediococcus parvulus CUPV1 and P. parvulus CUPV22) that produced high levels of 2-substituted (1,3)-β-d-glucans which increased the viscosity of the growth media. The (1,3)-β-d-glucan consisted of two main molecular species, with masses of approximately 107 and 104 Da, whose proportions varied among the strains. The three strains survived exposure to saliva and simulated gastric conditions at pH 5, with P. parvulus CUPV22 surviving at pH 3.1, and L. suebicus CUPV221 surviving at pH 1.8. All strains were resistant to pancreatin and bile salts. P. parvulus CUPV22 exhibited the highest adhesion (10.5%) to Caco-2 cells, which decreased to 1.2% after washing the cells. Finally, P. parvulus CUPV22 and L. suebicus CUPV221 induced the production of inflammation-related cytokines by polarized macrophages, and interestingly, L. suebicus stimulated the production of cytokine IL-10. These results indicate that the three strains have potential utility for the production of functional foods, This work was supported by the Ministerio de Ciencia e Innovación (Projects AGL2006-11932 and AGL2009-12998, Subprojects 01 and 02), the Universidad del País Vasco (UPV/EHU) (EHU08/37) and the Diputación Foral de Gipuzkoa, Programa Red Gipuzkoana de Ciencia, Tecnología e Innovación (co-financed by the European Union). Gaizka Garai-Ibabe acknowledges the Gobierno Vasco (Dpto. Agricultura, Pesca y Alimentación) for the pre-doctoral fellowship

Exopolysaccharides have prebiotic potential and contribute to the rheology and texture of fermented foods. Here we have analyzed the in vitro bioavailability and immunomodulatory properties of the 2-substituted (1,3)-β- d -glucan-producing bacterium Pediococcus parvulus 2.6. It resists gastrointestinal stress, adheres to Caco-2 cells, and induces the production of inflammation-related cytokines by polarized macrophages.

BACKGROUND Childhood hemangiomas are benign tumors of endothelial cells, characterized by a rapidly proliferating initial phase and followed by a slow involution. However, some grow and may reach a massive size, threatening a patient's functions or life. These require immediate medical treatment. OBJECTIVE The objective was to determine the therapeutic effectiveness of interferon (IFN)-α-2b in children with hemangiomas threatening the patient's functions or life. MATERIALS AND METHODS All patients were treated with IFN-α-2b at a dosage of 3 million U/m2 corporal surface, applied subcutaneously, 5 days a week for the first 6 months and subsequently three times a week for 6 to 24 months. RESULTS The study included 20 patients with hemangiomas localized in different sites and with diverse functional alterations: ages varied between 3 and 48 months (median, 12.8 months), and 8 were male and 12 female. An excellent response was observed in 17 (85%) patients. Side effects were slight and transitory; there was a follow-up from 7 to 10 years, and no late toxicity was observed. CONCLUSIONS We can conclude that IFN-α-2b is an effective option for treating alarming hemangiomas that are resistant to steroids and that endanger proper functioning of the affected organ or the patient's life.

International audience; 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 °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.

Aims: To study the occurrence of histidine, tyrosine and ornithine decarboxylase activity in lactic acid bacteria (LAB) isolated from natural ciders and to examine their potential to produce detrimental levels of biogenic amines. Methods and Results: The presence of biogenic amines in a decarboxylase synthetic broth and in cider was determined by reversed-phase high-performance liquid chromatography (RP-HPLC). Among the 54 LAB strains tested, six (five lactobacilli and one oenococci) were biogenic amine producers in both media. Histamine and tyramine were the amines formed by the LAB strains investigated. Lactobacillus diolivorans were the most intensive histamine producers. This species together with Lactobacillus collinoides and Oenococcus oeni also seemed to produce tyramine. No ability to form histamine, tyramine or putrescine by Pediococus parvulus was observed, although it is a known biogenic amine producer in wines and beers. Conclusions: This study demonstrated that LAB microbiota growing in ciders had the ability to produce biogenic amines, particularly histamine and tyramine, and suggests that this capability might be strain-dependent rather than being related to a particular bacterial species. Significance and Impact of the Study: Production of biogenic amines by food micro-organisms has continued to be the focus of intensive study because of their potential toxicity. The main goal was to identify the microbial species capable of producing these compounds in order to control their presence and metabolic activity in foods.

Aims: To study the influence of medium constituents on growth, and exopolysaccharide (EPS) production by a strain of Oenococcus oeni. The structure of one of the EPSs has also been characterized. Methods and Results: EPS concentration was estimated by the phenol/sulfuric acid method. After purification and fractionation of crude EPSs, the sugar composition was determined by GLC-MS of the TMS methyl glycosides. The major polysaccharide is 2-substituted-(1–3)-β-d-glucan. This structure was determined by methylation analysis and conventional 1H- and 13C-nuclear magnetic resonance spectroscopy. In addition, O. oeni synthesized two heteropolysaccharides, although a lesser proportion, constituted by galactose and glucose, and one of them also showed rhamnose. The sugar source has a clear influence on growth and EPS synthesis, and EPS production was not enhanced by adding ethanol or increasing the nitrogen source. EPS biosynthesis starts in the exponential growth phase, and continued during the stationary growth phase. Conclusions: Higher EPS yields were obtained on cultures grown on glucose + fructose. O. oeni produces a β-glucan, as the predominant EPS, and it is also able to produce two heteropolysaccharides. Significance and Impact of the Study: This work provides a better understanding of EPS synthesis by O. oeni and shows the first EPS structure described for this species.