A laminated polymer film formulation for enteric delivery of live vaccine and probiotic bacteria

Live bacterial cells (LBC) are administered orally as attenuated vaccines, to deliver biopharmaceutical agents, and as probiotics to improve gastrointestinal health. However, LBC present unique formulation challenges and must survive gastrointestinal antimicrobial defenses including gastric acid after administration. We present a simple new formulation concept, termed Polymer Film Laminate (PFL). LBC are ambient dried onto cast acid-resistant enteric polymer films that are then laminated together to produce a solid oral dosage form. LBC of a model live bacterial vaccine and a probiotic were dried directly onto a cast film of enteric polymer. The effectiveness at protecting dried cells in a simulated gastric fluid (pH 2.0) depended on the composition of enteric polymer film used, with a blend of ethylcellulose plus Eudragit L100 55 providing greater protection from acid than Eudragit alone. However, although PFL made from blended polymers films completely released low molecular weight dye into intestinal conditions (pH 7.0), they failed to release LBC. In contrast, PFL made from Eudragit alone successfully protected dried probiotic or vaccine LBC from simulated gastric fluid for 2h, and subsequently released all viable cells within 60min of transfer into simulated intestinal fluid. Release kinetics could be controlled by modifying the lamination method.

Dysregulated circulating dendritic cell function in ulcerative colitis is partially restored by probiotic strain Lactobacillus casei Shirota

BACKGROUND: Dendritic cells regulate immune responses to microbial products and play a key role in ulcerative colitis (UC) pathology. We determined the immunomodulatory effects of probiotic strain Lactobacillus casei Shirota (LcS) on human DC from healthy controls and active UC patients. METHODS: Human blood DC from healthy controls (control-DC) and UC patients (UC-DC) were conditioned with heat-killed LcS and used to stimulate allogeneic T cells in a 5-day mixed leucocyte reaction. RESULTS: UC-DC displayed a reduced stimulatory capacity for T cells (P < 0.05) and enhanced expression of skin-homing markers CLA and CCR4 on stimulated T cells (P < 0.05) that were negative for gut-homing marker β7. LcS treatment restored the stimulatory capacity of UC-DC, reflecting that of control-DC. LcS treatment conditioned control-DC to induce CLA on T cells in conjunction with β7, generating a multihoming profile, but had no effects on UC-DC. Finally, LcS treatment enhanced DC ability to induce TGFβ production by T cells in controls but not UC patients. CONCLUSIONS: We demonstrate a systemic, dysregulated DC function in UC that may account for the propensity of UC patients to develop cutaneous manifestations. LcS has multifunctional immunoregulatory activities depending on the inflammatory state; therapeutic effects reported in UC may be due to promotion of homeostasis.

Effect of seasonal variation on the properties of whipping cream, soft cheese and skim milk powder in the UK

Whipping cream, skim milk powder and soft cheese were produced throughout the year. Whipping cream manufactured in spring and winter produced significantly higher overrun and better serum stability, and whipping time was related to buffering capacity of raw milk. Heat stability of reconstituted skim milk powder (RSMP) at 9% TS was greater in summer and autumn, and greater than 25% TS throughout the year. It was positively related to the protein content of raw milk, but negatively with fat. In contrast to other dairy products, no significant effect of season on the properties of soft cheese was found.

In vitro evaluation of the fermentation properties and potential probiotic activity of Lactobacillus plantarum C4 in batch culture systems

Lactobacillus plantarum C4 has been tested in in vitro pH-controlled anaerobic faecal batch cultures as compared to Lactobacillus rhamnosus GG to determine changes caused to the composition of faecal bacteria. Effects upon major groups of the microbiota and levels of short-chain fatty acids (SCFA) were assessed over 24 h. Concomitantly, hydrophobic character and ability of both bacterial cells to adhere in vitro to Caco-2 cells were investigated. Quantitative analysis of bacterial populations revealed that there was a significant increase in Lactobacillus/Enterococcus numbers in vessels with probiotic supplemented with fructooligosaccharides (FOS), compared to the negative control. L. plantarum C4 showed to have more hydrophilic behaviour and fulfilled better adhesive properties, compared to L. rhamnosus GG. Thus, L. plantarum C4 can modulate the intestinal microbiota in vitro, promoting changes in some numerically and metabolically relevant microbial populations and shifts in the production of SCFA.

Effect of blending Jersey and Holstein-Friesian milk on Cheddar cheese processing, composition and quality

The aim of this study was to investigate the effects of numerous milk compositional factors on milk coagulation properties using Partial Least Squares (PLS). Milk from herds of Jersey and Holstein- Friesian cattle was collected across the year and blended (n=55), to maximise variation in composition and coagulation. The milk was analysed for casein, protein, fat, titratable acidity, lactose, Ca2+, urea content, micelles size, fat globule size, somatic cell count and pH. Milk coagulation properties were defined as coagulation time, curd firmness and curd firmness rate measured by a controlled strain rheometer. The models derived from PLS had higher predictive power than previous models demonstrating the value of measuring more milk components. In addition to the well-established relationships with casein and protein levels, CMS and fat globule size were found to have as strong impact on all of the three models. The study also found a positive impact of fat on milk coagulation properties and a strong relationship between lactose and curd firmness, and urea and curd firmness rate, all of which warrant further investigation due to current lack of knowledge of the underlying mechanism. These findings demonstrate the importance of using a wider range of milk compositional variables for the prediction of the milk coagulation properties, and hence as indicators of milk suitability for cheese making.

Improving survival of probiotic bacteria using bacterial poly-γ-glutamic acid

A major hurdle in producing a useful probiotic food product is bacterial survival during storage and ingestion. The aim of this study was to test the effect of γ-PGA immobilisation on the survival of probiotic bacteria when stored in acidic fruit juice. Fruit juices provide an alternative means of probiotic delivery, especially to lactose intolerant individuals. In addition, the survival of γ-PGA-immobilised cells in simulated gastric juice was also assessed. Bifidobacteria strains (B. longum, B. breve), immobilised on 2.5 % γ-PGA, survived significantly better (P < 0.05) in orange and pomegranate juice for 39 and 11 days respectively, compared to free cells. However, cells survived significantly better (P < 0.05) when stored in orange juice compared to pomegranate juice. Moreover, both strains, when protected with 2.5 % γ-PGA, survived in simulated gastric juice (pH 2.0) with a marginal reduction (<0.47 log CFU/ml) or no significant reduction in viable cells after four hours, whereas free cells died within two hours. In conclusion, this research indicates that γ-PGA can be used to protect Bifidobacteria cells in fruit juice, and could also help improve the survival of cells as they pass through the harsh conditions of the gastrointestinal tract (GIT). Following our previous report on the use of γ-PGA as a cryoprotectant for probiotic bacteria, this research further suggests that γ-PGA could be used to improve probiotic survival during the various stages of preparation, storage and ingestion of probiotic cells.

Expert consensus document: the International Scientific Association for Probiotics and Prebiotics consensus statement on the scope and appropriate use of the term probiotic.

An expert panel was convened in October 2013 by the International Scientific Association for Probiotics and Prebiotics (ISAPP) to discuss the field of probiotics. It is now 13 years since the definition of probiotics and 12 years after guidelines were published for regulators, scientists and industry by the Food and Agriculture Organization of the United Nations and the WHO (FAO/WHO). The FAO/WHO definition of a probiotic—“live microorganisms which when administered in adequate amounts confer a health benefit on the host”—was reinforced as relevant and sufficiently accommodating for current and anticipated applications. However, inconsistencies between the FAO/WHO Expert Consultation Report and the FAO/WHO Guidelines were clarified to take into account advances in science and applications. A more precise use of the term 'probiotic' will be useful to guide clinicians and consumers in differentiating the diverse products on the market. This document represents the conclusions of the ISAPP consensus meeting on the appropriate use and scope of the term probiotic.

Jersey milk suitability for Cheddar cheese production: process, yield, quality and financial impacts

The aim of this study was to first evaluate the benefits of including Jersey milk into Holstein-Friesian milk on the Cheddar cheese making process and secondly, using the data gathered, identify the effects and relative importance of a wide range of milk components on milk coagulation properties and the cheese making process. Blending Jersey and Holstein-Friesian milk led to quadratic trends on the size of casein micelle and fat globule and on coagulation properties. However this was not found to affect the cheese making process. Including Jersey milk was found, on a pilot scale, to increase cheese yield (up to + 35 %) but it did not affect cheese quality, which was defined as compliance with the legal requirements of cheese composition, cheese texture, colour and grading scores. Profitability increased linearly with the inclusion of Jersey milk (up to 11.18 p£ L-1 of milk). The commercial trials supported the pilot plant findings, demonstrating that including Jersey milk increased cheese yield without having a negative impact on cheese quality, despite the inherent challenges of scaling up such a process commercially. The successful use of a large array of milk components to model the cheese making process challenged the commonly accepted view that fat, protein and casein content and protein to fat ratio are the main contributors to the cheese making process as other components such as the size of casein micelle and fat globule were found to also play a key role with small casein micelle and large fat globule reducing coagulation time, improving curd firmness, fat recovery and influencing cheese moisture and fat content. The findings of this thesis indicated that milk suitability for Cheddar making could be improved by the inclusion of Jersey milk and that more compositional factors need to be taken into account when judging milk suitability.

Optimisation of oat milk formulation to obtain fermented derivatives by using probiotic Lactobacillus reuteri microorganisms

Functional advantages of probiotics combined with interesting composition of oat were considered as an alternative to dairy products. In this study, fermentation of oat milk with Lactobacillus reuteri and Streptococcus thermophilus was analysed to develop a new probiotic product. Central composite design with response surface methodology was used to analyse the effect of different factors (glucose, fructose, inulin and starters) on the probiotic population in the product. Optimised formulation was characterised throughout storage time at 4 ℃ in terms of pH, acidity, β-glucan and oligosaccharides contents, colour and rheological behaviour. All formulations studied were adequate to produce fermented foods and minimum dose of each factor was considered as optimum. The selected formulation allowed starters survival above 107/cfu ml to be considered as a functional food and was maintained during the 28 days controlled. β-glucans remained in the final product with a positive effect on viscosity. Therefore, a new probiotic non-dairy milk was successfully developed in which high probiotic survivals were assured throughout the typical yoghurt-like shelf life.

Stability of probiotic Lactobacillus plantarum in dry microcapsules under accelerated storage conditions

This study investigated the stability of freeze dried and fluid bed dried alginate microcapsules coated with chitosan containing model probiotic bacteria, Lactobacillus plantarum, during storage for up to 45 days at different water activities (0.11, 0.23, 0.40 and 0.70) and temperatures (4, 30 and 37 °C). The loss in cell viability was around 0.8 log in the case of fluid bed drying and around 1.3 in the case of freeze drying, with the former method resulting in dried capsules of smaller size (~ 1 mm vs 1.3 mm), more irregular shape, and with a rougher surface. In both cases, the water activity and water content were less than 0.25 and 10% w/w, respectively, which favours high storage stability. The storage stability studies demonstrated that as the water activity and temperature decreased the survival of the dried encapsulated cells increased. Considerably better survival was observed for fluid bed dried encapsulated cells compared to freeze dried encapsulated cells and freeze dried free cells with 10% sucrose (control), and in some cases, e.g. at 4 and 30 °C at water activities of 0.11, 0.23 and 0.40, there was more than 1 log difference after 45 days, with concentrations higher than 108 CFU/g after 45 days of storage. The results indicate that fluid bed drying is an effective and efficient manufacturing method to produce probiotic containing capsules with enhanced storage stability.

Short communication: estimation of the financial benefit of using Jersey milk at different inclusion rates for Cheddar cheese production using partial budgeting

Partial budgeting was used to estimate the net benefit of blending Jersey milk in Holstein-Friesian milk for Cheddar cheese production. Jersey milk increases Cheddar cheese yield. However, the cost of Jersey milk is also higher; thus, determining the balance of profitability is necessary, including consideration of seasonal effects. Input variables were based on a pilot plant experiment run from 2012 to 2013 and industry milk and cheese prices during this period. When Jersey milk was used at an increasing rate with Holstein-Friesian milk (25, 50, 75, and 100% Jersey milk), it resulted in an increase of average net profit of 3.41, 6.44, 8.57, and 11.18 pence per kilogram of milk, respectively, and this additional profit was constant throughout the year. Sensitivity analysis showed that the most influential input on additional profit was cheese yield, whereas cheese price and milk price had a small effect. The minimum increase in yield, which was necessary for the use of Jersey milk to be profitable, was 2.63, 7.28, 9.95, and 12.37% at 25, 50, 75, and 100% Jersey milk, respectively. Including Jersey milk did not affect the quantity of whey butter and powder produced. Althoug further research is needed to ascertain the amount of additional profit that would be found on a commercial scale, the results indicate that using Jersey milk for Cheddar cheese making would lead to an improvement in profit for the cheese makers, especially at higher inclusion rates.

Selective effects of Lactobacillus casei Shirota on T cell activation, natural killer cell activity and cytokine production

Modulation of host immunity is an important potential mechanism by which probiotics confer health benefits. This study was designed to investigate the effects of a probiotic strain, Lactobacillus casei Shirota (LcS), on immune function, using human peripheral blood mononuclear cells (PBMC) in vitro. In addition, the role of monocytes in LcS-induced immunity was also explored. LcS promoted natural killer (NK) cell activity and preferentially induced expression of CD69 and CD25 on CD8+ and CD56+ subsets in the absence of any other stimulus. LcS also induced production of IL-1β, IL-6, TNF-α, IL-12 and IL-10 in the absence of lipopolysaccharide (LPS). In the presence of LPS, LcS enhanced IL-1β production, but inhibited LPS-induced IL-10 and IL-6 production, and had no further effect on TNF-α and IL-12 production. Monocyte-depletion significantly reduced the impact of LcS on lymphocyte activation, cytokine production and NK cell activity. In conclusion, LcS preferentially activated cytotoxic lymphocytes in both the innate and specific immune system, which suggests that LcS could potentiate the destruction of infected cells in the body. LcS also induced both pro-inflammatory and anti-inflammatory cytokine production in the absence of LPS, but inhibited LPS-induced cytokine production in some cases. Monocytes play an important role in LcS-induced immunological responses.

Input usage, output mix and industry deregulation: an analysis of the Australian dairy manufacturing industry

In this paper we estimate a Translog output distance function for a balanced panel of state level data for the Australian dairy processing sector. We estimate a fixed effects specification employing Bayesian methods, with and without the imposition of monotonicity and curvature restrictions. Our results indicate that Tasmania and Victoria are the most technically efficient states with New South Wales being the least efficient. The imposition of theoretical restrictions marginally affects the results especially with respect to estimates of technical change and industry deregulation. Importantly, our bias estimates show changes in both input use and output mix that result from deregulation. Specifically, we find that deregulation has positively biased the production of butter, cheese and powders.