Paul Peiris

Encapsulation of probiotic bacteria with alginate–starch and evaluation of survival in simulated gastrointestinal conditions and in yoghurt

A modified method using calcium alginate for the microencapsulation of probiotic bacteria is reported in this study. Incorporation of Hi-Maize starch (a prebiotic) improved encapsulation of viable bacteria as compared to when the bacteria were encapsulated without the starch. Inclusion of glycerol (a cryo-protectant) with alginate mix increased the survival of bacteria when frozen at -20 degrees C. The acidification kinetics of encapsulated bacteria showed that the rate of acid produced was lower than that of free cultures. The encapsulated bacteria, however, did not demonstrate a significant increase in survival when subjected to in vitro high acid and bile salt conditions. A preliminary study was carried out in order to monitor the effects of encapsulation on the survival of Lactobacillus acidophilus and Bifidobacterium spp. in yoghurt over a period of 8 weeks. This study showed that the survival of encapsulated cultures of L. acidophilus and Bifidobacterium spp. showed a decline in viable count of about 0.5 log over a period of 8 weeks while there was a decline of about 1 log in cultures which were incorporated as free cells in yoghurt. The encapsulation method used in this study did not result in uniform bead size, and hence additional experiments need to be designed using uniform bead size in order to assess the role of different encapsulation parameters, such as bead size and alginate concentration, in providing protection to the bacteria.

Application of RBGR: a simple way for screening of oxygen tolerance in probiotic bacteria

Oxygen toxicity is a major problem in the survival of probiotic bacteria in dairy foods. High levels of oxygen in the product are detrimental to the viability of these predominantly anaerobic bacteria. Screening probiotic bacteria for oxygen tolerance before their incorporation could ensure high cell counts in food products during storage. Reported techniques have focused only on qualitative estimations of oxygen tolerance in probiotic bacteria. To characterize the oxygen tolerance of a large number of organisms, a quantitative measurement is essential. For the first time, the oxygen tolerance of several probiotic strains was measured quantitatively using an index known as Relative Bacterial Growth Ratio (RBGR). The tolerance to oxygen varied between organisms, and this technique can therefore be applied for screening probiotic bacteria for oxygen tolerance.

Inhibition of Non-proteolytic Clostridium botulinum with Lactic Acid Bacteria in Extended Shelf-Life Cook-Chill Soups

The possibility of protecting cook-chill foods with microbial cultures against the risk of botulism was demonstrated. Three commercial soups were incubated with Clostridium botulinum 17B (103 spores/g) and protective cultures (PCs) during 10–15 days at 10°C. The PCs populations were enumerated on M17, MRS and maltose tryptic soy agar, C. botulinum—on sorbitol tryptic soy agar, botulinal toxin was detected by the immunoassay, bacteriocins—by well diffusion assay. C. botulinum did not grow in two soups with low pH (5.2–5.5) and was unaffected by the PCs. In seafood chowder (pH 6.2) C. botulinum populations reached 108 cfu/g. The co-incubation with the PCs, nisin-producing Lactococcus lactis (107 cfu/g) or pediocin-producing Pediococcus pentosaceus (3×108 cfu/g) singularly and as a mixture, prevented toxigenesis as well as reduced the product pH to 4.8–5.0 and C. botulinum populations to undetectable levels. Color, mouth-feel, texture, flavor and the overall acceptability of seafood chowder was not affected by the presence of the PCs.

Simple and robust monitoring of ethanol fermentations by capillary electrophoresis

Free‐solution capillary electrophoresis (CE), or capillary zone electrophoresis, with direct UV detection was used for the first time for the determination of mono‐ and disaccharides, sugar alcohols, and ethanol in fermentation broths. Sample preparation proved to be minimal: no derivatization or specific sample purification was needed. The CE conditions can be adapted to the type of fermentation by simply altering the background electrolyte (BGE). KOH (130 mM) or NaOH (130 mM) as the BGE led to the fastest analysis time when monitoring simple fermentations. A mixture of 65 mM NaOH and 65 mM LiOH led to a 19% improvement in resolution for a complex mixture of carbohydrates. Quantification of a simple carbohydrate fermentation by CE showed values in close agreement with that of high‐performance anion exchange chromatography and high‐performance liquid chromatography (HPLC) on a cation exchange resin. For complex fermentations, quantification of carbohydrates by HPLC and CE led to similar results, whereas CE requires an injection volume of only 10–20 nL. Analysis of an ethanol fermentation of hydrolyzed plant fiber demonstrated the robustness of the separation and detection of carbohydrates, as well as ethanol. Ethanol determination is achieved by coupling the CE method to pressure mobilization, using the same instrument and the same sample.

Characterization of the exopolysaccharide produced by a whey utilizing strain of Klebsiella oxytoca

Physical, chemical and rheological properties of a polysaccharide produced by an isolate of Klebsiella oxytoca were characterized. Freeze dried samples of the polysaccharide were neutral and were completely soluble in water. Samples did not form gels even in the presence of salt treatments. The major monosaccharide constituents of the polysaccharide were rhamnose (37%, w/w) and glucose (34%, w/w). Residues of cellobiose were detected, suggesting that the polysaccharide had a cellulose backbone. The gum was more comparable to broth apparent viscosities of xanthan gum than to gellan gum. The K. oxytoca polysaccharide (KOP) produced high solution viscosity at low concentrations. At a gum concentration 0.5% (w/v), an apparent viscosity of 400 cP at 24 s-1 was obtained. Rheological behavior showed that the KOP formed non newtonian fluids, indicating that it is a pseudoplastic biopolymer. Although the KOP solutions displayed pseudoplastic behavior, increases in shearing time did not result in significant changes on the apparent viscosity. This indicated that the gum is neither thixotropic nor rheopectic. The conclusion reached about the potential application of the gum was that it could be suitable for use as a stabilizing or suspending agent rather than a gelling agent.

Involvement of a plasmid in exopolysaccharide production by Klebsiella oxytoca

A Klebsiella oxytoca isolate which can produce significant levels of an exopolysaccharide using whey as a growth substrate has been reported. The plasmid profile of this isolate was shown to be different from that of the non-exopolysaccharide-producing K. oxytoca ATCC 43863. Irreversible curing of the single plasmid in the K. oxytoca isolate was achieved using 30 μg acriflavin/ml. The ability to produce the exopolysaccharide was lost with the curing of the plasmid (parent strain produced a medium viscosity of 1260 cP at 1 s−1 compared to 1.6 cP at 200 s−1 produced by the cured strain). However, the ability to metabolize lactose was not significantly affected by curing, and both the parent and the cured strain produced similar levels of viable cells (~109 cfu/ml) after 62 h growth on lactose-rich medium. The exopolysaccharide-producing ability of the isolate was stable for at least 139 generations.

Rheological characteristics of an exopolysaccharide produced by a strain of Klebsiella oxytoca

Rheological characteristics of Klebsiella oxytoca exopolysaccharide (KOP) were investigated. Syneresis was 10% significantly lower (P<0.01) in KOP treated yoghurt than in the control. KOP yoghurt was highest in gumminess 80 and hardness 182. KOP and commercial binder treated sausages did not differ (P>0.01) in firmness, however, 0.25% (w/w) of the KOP replaced 17% (w/w) of commercial binder.

Short Communication: Optimization of bioprocess conditions for exopolysaccharide production by Klebsiella oxytoca

Optimization of bioprocess conditions increased exopolysaccharide production by a strain of Klebsiella oxytoca from 6g/l to 15g/l; this corresponded to an increase in medium viscosity from 36cP at 12s−1 to 20,000 cP at 0.6 s−1. A combination of equal proportions of tryptone nitrogen and urea nitrogen proved to be the best nitrogen source. Lactose was shown to be the preferred carbon source. At an optimum temperature of 25°C, a pH of 7 was found to be the best for exopolysaccharide production. The concentration of exopolysaccharide produced on whey, enriched whey, enriched whey permeate and lactose-rich medium was comparable.