Amelia Farrés

Yogurt production from reconstituted skim milk powders using different polymer and non-polymer forming starter cultures

Using polymer producing (ropy) strains of lactic acid bacteria it was possible to reduce considerably the syneresis of yogurt, even with 12% total milk solids. The viscosities obtained with these strains were also similar to those obtained using normal strains and milk with 17% total solids content. The concentration of milk and the polymer produced by ropy starters had a synergic effect in increasing viscosity. Polymer production was not affected in most cases by milk concentration. One type of ropy culture (Wiesby) seemed to produce a different kind of polymer as it could not be determined by alcohol precipitation, in spite of being able to reduce syneresis and increase viscosity in yogurt. A limited number of yogurts were evaluated organoleptically, one prepared with a ropy starter strain (NCFB at 12, 14·5 and 17% total solids) and one prepared with a non-ropy strain (LL-I at 17% total solids). The results suggest that the ropy strain yogurts had different mouthfeel from the non-ropy strain yogurts; the most acceptable product overall was the ropy strain made with 12% total solids.

Lactic acid bacterial diversity in the traditional mexican fermented dough pozol as determined by 16S rDNA sequence analysis.

The lactic acid bacteria diversity of pozol, a Mexican fermented maize dough, was studied using a total DNA extraction and purification procedure and PCR amplification of 16S rDNA for gram-positive and related bacterial groups. Thirty-six clones were obtained and sequenced to 650 nucleotides. These partial sequences were identified by submission to the non-redundant nucleotide database of NCBI. The identified sequences were aligned with reference sequences of the closest related organisms. This analysis indicated that only 14 sequences were unique clones and these were identified as Lactococcus lactis, Streptococcus suis, Lactobacillus plantarum, Lact. casei, Lact. alimentarium, and Lact. delbruekii and Clostridium sp. Two non-ribosomal sequences were also detected. Unlike other environments analyzed with this molecular approach where many unidentified microorganisms are found, the identity of most sequences could be established as lactic acid bacteria, indicating that this is the main group among the gram-positive bacteria in pozol. Use of this molecular method permitted detection of lactic acid bacteria different from those previously isolated and identified by culture techniques

Enzymes involved in carbohydrate metabolism and their role on exopolysaccharide production in Streptococcus thermophilus

The role of the enzymes uridine‐5′‐diphospho‐(UDP) glucose pyrophosphorylase and UDP galactose 4‐epimerase in exopolysaccharide production of Gal− ropy and non‐ropy strains of Streptococcus thermophilus in a batch culture was investigated. Growth of the ropy and non‐ropy strains was accompanied by total release of the galactose moiety from lactose hydrolysis in modified Bellinker broth with lactose as the only carbon source. This was associated with a greater exopolysaccharide production by the ropy strain. The polymer produced by both strains in cultures with lactose or glucose as carbon sources contained glucose, galactose and rhamnose, indicating that glucose was used as a carbon source for bacterial growth and for exopolysaccharide formation. UDP‐glucose pyrophosphorylase activity was associated with polysaccharide production during the first 12 h in a 20 h culture in the ropy strain, but not in the non‐ropy strain. UDP‐galactose 4‐epimerase was not associated with exopolysaccharide synthesis in any strain. The evidence presented suggests that the glucose moiety from lactose hydrolysis is the source of sugar for heteropolysaccharide synthesis, due to a high UDP‐glucose pyrophosphorylase activity.

Medium optimization by a fractional factorial design for lipase production by Rhizopus delemar

Abstract The aim of the present work is to improve production of extracellular lipase by Rhizopus delemar using industrial grade carbon and nitrogen sources and performing statistical optimization of the production medium. Soya paste and dextrin were selected in the first phase of this work as the best carbon and nitrogen sources and were used in the following experiments. Optimization of the medium composition was carried out with a fractional factorial design in two steps. Lipolytic activity at 48 h was 12-fold greater than with the initial medium. The results show that it is necessary to modify the carbon source-nitrogen source ratio, to reduce olive oil concentration and to add Tween 80 to the medium to achieve maximum production.

Purification and characterization of an extracellular lipase from Penicillium candidum

Penicillium candidum produces and secretes a single extracellular lipase with a monomer molecular weight of 29 kDa. However, this enzyme forms dimers and higher molecular weight aggregates under nondenaturing conditions. The lipase from P. candidum was purified 37-fold using Octyl-Sepharose CL-4B and DEAE-Sephadex columns. The optimal assay conditions for lipase activity were 35°C and pH 9. The lipase was stable in the pH range of 5–6 with a pl of 5.5, but rapid loss of the enzyme activity was observed above 25°C. Tributyrin was found to be the best substrate for the P. candidum lipase, among those tested. Metal ions such as Fe2+ and Cu2+ inhibited enzymatic activity and only Ca2+ was able to slightly enhance lipase activity. Ionic detergents inhibited the activity of the enzyme, whereas nonionic detergents stimulated lipase activity.

Kinetic studies of Gly28:Ser mutant form of Bacillus pumilus lipase: Changes in kcat and thermal dependence

Lipases are useful catalysts for a wide variety of industrial purposes. Herein we report the stability and thermal dependence of the activity of wild-type Bacillus pumilus lipase (BplA) and four site-directed mutants designed to improve its thermal stability. The Gly28:Ser mutation produces a dramatic four-fold increase in its k(cat) and a remarkable increase in its stability. While the increase in k(cat) is temperature-independent, the increase in stability shows that the resultant interactions of this mutation have a strong enthalpic component. Thermal dependence of stability, k(cat), K(M) and k(cat)/K(M) were analysed to gain insight on the structural effects of mutations on BplA. Our results are consistent with a gain in enzyme mobility for those mutants displaying enhanced catalytic properties; the analysis of thermal dependence of kinetic parameters indicates that the mutations did not change either the catalytic mechanism or the rate-limiting step of catalysis.

Development of a liquid nutritional supplement using a Sesamum indicum L. protein isolate

This work presents some functional properties and the potential use of a new protein isolate from sesame seed flour, Sesaprot® (SP), as protein source in a liquid nutritional supplement. It was compared to a commercial soybean isolate and results showed that its emulsifying properties are better than those of its soybean counterpart. Lysine content is lower than FAO recommendations for children, but is adequate for adults. Other essential amino acids, however, are present in adequate amounts for any kind of consumer. Osmolality, pH and emulsion stability of an experimental formulation were similar to those of commercial beverages. A consumer sensory test indicated that the product was preferred to one prepared with a soybean isolate and to a commercial brand that was assayed.

ANCUT2, an Extracellular Cutinase from Aspergillus nidulans Induced by Olive Oil

Cutinases are versatile carboxylic ester hydrolases with great potential in many biocatalytic processes, including biodiesel production. Genome sequence analysis of the model organism Aspergillus nidulans reveals four genes encoding putative cutinases. In this work, we purified and identified for the first time a cutinase (ANCUT2) produced by A. nidulans. ANCUT2 is a 29-kDa protein which consists of 255 amino acid residues. Comparison of the amino acid sequence of ANCUT2 with other microbial cutinase sequences revealed a high degree of homology with other fungal cutinases as well as new features, which include a serine-rich region and conserved cysteines. Cutinase production with different lipidic and carbon sources was also explored. Enzyme activity was induced by olive oil and some triacylglycerides and fatty acids, whereas it was repressed by glucose (1%) and other sugars. In some conditions, a 22-kDa post-translational processing product was also detected. The cutinase nature of the enzyme was confirmed after degradation of apple cutin.

Influence of water activity on the fermentation of yogurt made with extracellular polysaccharide-producing or non-producing starters

Abstract Ropy and non-ropy strains of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus were studied with respect to their resistance to reduction of water activity in milk used to produce yogurt. Glycerol reduced the a w of the milk considerably, but it was less inhibitory to acid production than sucrose. The ropy strains did not show any advantage over nonropy ones when they were used to produce yogurt in milk with reduced a w ; their performance was similar to non-producer strains in terms of lactic acid production.

The starch-binding domain as a tool for recombinant protein purification

Recombinant protein purification with affinity tags is a widely employed technique. One of the most common tags used for protein purification is the histidine tag (Histag). In this work, we use a tandem starch-binding domain (SBDtag) as a tag for protein purification. Four proteins from different sources were fused to the SBDtag, and the resulting fusion proteins were purified by affinity chromatography using the Histag or the SBDtag. The results showed that the SBDtag is superior to the Histag for protein purification. The efficient adsorption of the fusion proteins to raw corn starch was also demonstrated, and two fusions were selected to test purification directly using raw starch from rice, corn, potato, and barley. The two fusion proteins were successfully recovered from crude bacterial extract using raw starch, thus demonstrating that the SBDtag can be used as an efficient affinity tag for recombinant protein purification on an inexpensive matrix.