Release studies of Lactobacillus casei strain Shirota from chitosan-coated alginate-starch microcapsules in ex vivo porcine gastrointestinal contents.

AIMS: To investigate the release characteristics of Lactobacillus casei strain Shirota (LCS) from chitosan-coated alginate-starch (CCAS) capsule in different regions of ex vivo porcine gastrointestinal (GI) contents. METHODS AND RESULTS: A 0.1 g of CCAS encapsulated bacteria (containing c. 10(8) CFU of LCS) were incubated in different sections of ex vivo porcine GI contents (10 ml) anaerobically at 37 degrees C. Samples were taken from different GI contents at different time intervals (up to 24 h) and estimated for the release of LCS from capsules. There was almost a complete release (1.1 x 10(8) CFU) of LCS from capsules within 8 h of incubation in ileal contents, while it took nearly 12 h to release the completely encapsulated bacteria in colon content under similar conditions. There was only a partial release of encapsulated LCS, incubated in duodenal and jejunal contents, while there was no significant (P > 0.05) release of encapsulated bacteria in gastric contents even after 24 h of incubation. CONCLUSIONS: The capsules were able to release viable probiotic cells completely in ex vivo porcine ileal and colon contents. SIGNIFICANCE AND IMPACT OF THE STUDY: CCAS capsule can be an effective way of protecting probiotic bacteria from adverse gastric conditions and delivering viable bacteria to the host intestinal systems.

Alginate-pectin microcapsules as a potential for folic acid delivery in foods.

Most naturally occurring folate derivatives in foods are highly sensitive to temperature, oxygen, light and their stability is affected by processing conditions. Folic acid incorporated microcapsules using alginate and combinations of alginate and pectin polymers were prepared to improve stability. Folic acid stability was evaluated with reference to encapsulation efficiency, gelling and hardening of capsules, capsular retention during drying and storage. Use of alginate in combination with pectin produced more robust capsules and contributed to greater encapsulation efficiency. The capsules lost their spherical shape as a consequence of increased pectin. The high alginate capsules, A100:P0 (100% alginate: 0% pectin) and A80:P20 (80% alginate: 20% pectin) were of regular spherical shape, while those with more pectin, A70:P30 (70% alginate: 30% pectin) and A60:P40 (60% alginate: 40% pectin) formed irregular spheres. The loading efficiency, expressed as a percentage of the actual loading to theoretical loading, varied from 55-89% with the composition of the mixed polymers. After 11 weeks of storage at 4 degrees C, folic acid retention in freeze-dried capsules was 100% (A70:P30 and A60:P40), 80% (A80:P20) and 30% (A100:P0). The blended alginate and pectin polymer matrix increased folic acid encapsulation efficiency and reduced the leakage from the capsules compared to those made with alginate alone and showed higher folic acid retention after freeze drying and storage.

An improved method of microencapsulation and its evaluation to protect Lactobacillus spp. in simulated gastric conditions.

An improved method of microencapsulation was developed to increase the efficacy of capsules in protecting the encapsulated bacteria under simulated gastric conditions. Lactobacillus acidophilus CSCC 2400 was encapsulated in calcium alginate and tested for its survival in simulated gastric conditions. The effects of different capsule sizes (200, 450, 1000 microm), different sodium alginate concentrations (0.75%, 1%, 1.5%, 1.8% and 2% w/v) and different concentrations of calcium chloride (0.1, 0.2, 1.0 M) on the viability of encapsulated bacteria were investigated. The viability of the cells in the microcapsules increased with an increase in alginate capsule size and gel concentration. There was no significant difference (p>0.05) in the viability of encapsulated cells when the concentration of calcium chloride was increased. Increase in cell load during encapsulation increased the number of bacterial survivors at the end of 3-h incubation in simulated gastric conditions. Hardening the capsule in calcium chloride solution for a longer time (8 h) had no impact on increasing the viability of encapsulated bacteria in a simulated gastric environment. The release of encapsulated cells at different phosphate buffer concentrations was also studied. When encapsulated L. acidophilus CSCC 2400 and L. acidophilus CSCC 2409 were subjected to low pH (pH 2) and high bile concentration (1.0% bile) under optimal encapsulation conditions (1.8% (w/v) alginate, 10(9) CFU/ml, 30 min hardening in 0.1 M CaCl(2) and capsule size 450 microm), there was a significant increase (p<0.05) in viable cell counts, compared to the free cells under similar conditions. Thus the encapsulation method described in this study may be effectively used to protect the lactobacillus from adverse gastric conditions.

A Review of Oxygen Toxicity in Probiotic Yogurts: Influence on the Survival of Probiotic Bacteria and Protective Techniques.

Oxygen toxicity is considered a significant factor influencing the viability of probiotic bacteria such as Lactobacillus and Bifidobacterium spp. in yogurts. This review assesses the involvement of oxygen during the manufacture of yogurt and its diffusion into the final product. The oxygen sensitivity of Lactobacillus acidophilus and Bifidobacterium spp. is discussed. The impact of dissolved oxygen of the product on the survival of these probiotic bacteria is highlighted. Additionally, microbiological, chemical, and packaging techniques recommended to protect probiotic bacteria from oxygen toxicity in dairy products are reviewed.

Metabolic and biochemical responses of probiotic bacteria to oxygen.

The interaction between oxygen and probiotic bacteria was studied by growing Lactobacillus acidophilus and Bifidobacterium spp. in 0, 5, 10, 15, and 21% oxygen in a hypoxic glove box. The metabolic responses of each probiotic strain in the different oxygen environments were monitored by measuring the levels of lactic acid and determining the lactate-to-acetate ratio. Biochemical changes induced by oxygen were examined by monitoring the specific activities of NADH oxidase, NADH peroxidase, and superoxide dismutase. In addition, the ability to decompose hydrogen peroxide and the sensitivity of each strain to hydrogen peroxide was also determined. With an increase in oxygen percentage, levels of lactic acid in L. acidophilus strains decreased, whereas the lactate-to-acetate ratio reduced in all the bifidobacteria tested. At 21% oxygen, the specific activities of NADH oxidase and NADH peroxidase, and the hydrogen peroxide decomposing ability of five probiotic strains was significantly higher than at 0% oxygen. The sensitivity of the probiotic strains to hydrogen peroxide however, remained unaffected in all the different oxygen percentages. Superoxide dismutase levels did not reveal any conclusive trend. In both L. acidophilus and Bifidobacterium spp., NADH oxidase and NADH peroxidase functioned optimally at pH 5. Growth in the various oxygen environments did not change this optimum pH.

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.

Survival and therapeutic potential of probiotic organisms with reference to Lactobacillus acidophilus and Bifidobacterium spp.

The present paper provides an overview on the use of probiotic organisms as live supplements, with particular emphasis on Lactobacillus acidophilus and Bifidobacterium spp. The therapeutic potential of these bacteria in fermented dairy products is dependent on their survival during manufacture and storage. Probiotic bacteria are increasingly used in food and pharmaceutical applications to balance disturbed intestinal microflora and related dysfunction of the human gastrointestinal tract. Lactobacillus acidophilus and Bifidobacterium spp. have been reported to be beneficial probiotic organisms that provide excellent therapeutic benefits. The biological activity of probiotic bacteria is due in part to their ability to attach to enterocytes. This inhibits the binding of enteric pathogens by a process of competitive exclusion. Attachment of probiotic bacteria to cell surface receptors of enterocytes also initiates signalling events that result in the synthesis of cytokines. Probiotic bacteria also exert an influence on commensal micro-organisms by the production of lactic acid and bacteriocins. These substances inhibit growth of pathogens and also alter the ecological balance of enteric commensals. Production of butyric acid by some probiotic bacteria affects the turnover of enterocytes and neutralizes the activity of dietary carcinogens, such as nitrosamines, that are generated by the metabolic activity of commensal bacteria in subjects consuming a high-protein diet. Therefore, inclusion of probiotic bacteria in fermented dairy products enhances their value as better therapeutic functional foods. However, insufficient viability and survival of these bacteria remain a problem in commercial food products. By selecting better functional probiotic strains and adopting improved methods to enhance survival, including the use of appropriate prebiotics and the optimal combination of probiotics and prebiotics (synbiotics), an increased delivery of viable bacteria in fermented products to the consumers can be achieved.

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.