Probiotic Food Products Research Articles

Cholesterol-Lowering Probiotics as Potential Biotherapeutics for Metabolic Diseases

Cardiovascular diseases are one of the major causes of deaths in adults in the western world. Elevated levels of certain blood lipids have been reported to be the principal cause of cardiovascular disease and other disabilities in developed countries. Several animal and clinical trials have shown a positive association between cholesterol levels and the risks of coronary heart disease. Current dietary strategies for the prevention of cardiovascular disease advocate adherence to low-fat/low-saturated-fat diets. Although there is no doubt that, in experimental conditions, low-fat diets offer an effective means of reducing blood cholesterol concentrations on a population basis, these appear to be less effective, largely due to poor compliance, attributed to low palatability and acceptability of these diets to the consumers. Due to the low consumer compliance, attempts have been made to identify other dietary components that can reduce blood cholesterol levels. Supplementation of diet with fermented dairy products or lactic acid bacteria containing dairy products has shown the potential to reduce serum cholesterol levels. Various approaches have been used to alleviate this issue, including the use of probiotics, especially Bifidobacterium spp. and Lactobacillus spp.. Probiotics, the living microorganisms that confer health benefits on the host when administered in adequate amounts, have received much attention on their proclaimed health benefits which include improvement in lactose intolerance, increase in natural resistance to infectious disease in gastrointestinal tract, suppression of cancer, antidiabetic, reduction in serum cholesterol level, and improved digestion. In addition, there are numerous reports on cholesterol removal ability of probiotics and their hypocholesterolemic effects. Several possible mechanisms for cholesterol removal by probiotics are assimilation of cholesterol by growing cells, binding of cholesterol to cellular surface, incorporation of cholesterol into the cellular membrane, deconjugation of bile via bile salt hydrolase, coprecipitation of cholesterol with deconjugated bile, binding action of bile by fibre, and production of short-chain fatty acids by oligosaccharides. The present paper reviews the mechanisms of action of anti-cholesterolemic potential of probiotic microorganisms and probiotic food products, with the aim of lowering the risks of cardiovascular and coronary heart diseases.

The science and regulations of probiotic food and supplement product labeling

Presented by the New York Academy of Sciences, the U.S. Food and Drug Administration (FDA), and the U.S. Office of Dietary Supplements of the National Institutes of Health, the symposium "Probiotic Foods and Supplements: The Science and Regulations of Labeling," was held on June 12, 2010 at the New York Academy of Sciences, New York, NY, the goals of which were to facilitate the exchange of ideas regarding labeling and substantiation of claims for probiotics among academic, industry, and regulatory professionals, and to discuss ways to translate and communicate research results in a truthful way to the consumer and to such health professionals as physicians, pharmacists, and dieticians. The target audience for this symposium included academicians interested in conducting research on the health benefits of probiotics; scientists; communications personnel, and regulatory specialists from companies involved in, or interested in, the marketing of probiotics; U.S. government regulatory experts tasked with oversight of probiotic foods and dietary supplement products; and other experts in the field interested in the development of probiotics for the U.S. market.

Progress in the field of probiotics: year 2011

The interest and scope for research in the field of probiotics has significantly widened in recent years. This brief review covers the significant advances in the field of probiotics. These include conclusive evidence for the benefits of probiotics in preventing all cause mortality and necrotizing enterocolitis (NEC) in preterm very low birth weight (VLBW) neonates, understanding the role of probiotics as vaccine adjuvants, and in modulating inflammatory bowel diseases, bowel cancer, type 1 diabetes mellitus, obesity, high cholesterol levels, and bacterial resistance. Other areas of progress include understanding the role of probiotics in oral health and ageing. Current evidence will lead to routine probiotic supplementation to prevent all cause mortality and NEC in preterm VLBW neonates. Probiotics may also become novel agents as vaccine adjuvants, and in dealing with major public health issues such as obesity, type I diabetes mellitus, and poor oral health. Research in applications of probiotics in food products will rationalize product development, and health claims. As new frontiers continue to be explored the challenges to the basis of the hygiene hypothesis will influence further developments in the field of probiotics.

Effect of probiotic bacteria on chemical composition and sensory quality of fermented sausages

Probiotic food products are very popular on domestic and international markets. The application of probiotics in dairy products is quite frequent, while their application in meat products is still being explored. Three variants of Sremska- type fermented sausages were produced using starter culture Bactoferm T-SPX (Chr. Hansen): 1. control sausage variant; 2. variant with Lactobacillus helveticus RO52 (Lallemand, France); 3. variant with Bifidobacterium longum RO175 (Lallemand, France). Through 40 days of fermented sausage ripening, the survival of probiotic bacteria, the changes of starter bacteria counts, as well as the chemical composition, pH values and sensory evaluation were examined. During the first period of ripening, Lactobacillus helveticus RO52 and Bifidobacterium longum RO175 counts were 106 cfug−1, after which they increased to the level of 108 cfug−1 and remained there until the end of ripening. Starter bacteria counts were within the range typical for fermented sausages. The chemical composition and pH values of fermented sausages produced with probiotic bacteria did not significantly differ from the control variant. Sensory evaluation has shown that all variants of fermented sausages had an acceptable sensory quality.Based on the survival of probiotic bacteria during 40 days of fermented sausage ripening, it can be concluded that probiotics can be successfully used in the production of fermented sausages, without affecting the sensory quality of the sausage.

Evaluation of Sensory Properties of Probiotic Yogurt Containing Food Products with Prebiotic Fibresin Mwanza, Tanzania

Yogurt becomes a functional food upon incorporating probiotics-live microorganisms which when adequately administered confer health benefits. Prebiotics are fermentable fibres that nourish beneficial gastrointestinal microflora enhance the functionality of probiotics. This research aimed to improve the acceptability and functionality of probiotic yogurt produced in Mwanza, Tanzania by incorporating probiotic food ingredients. The probiotic culture Lactobacillus rhamnosus GR-1 and standard yogurt cultures Lactobacillus delbrueckii bulgaricus and Steptococcus thermophilus were used to manufacture yogurt, then locally available prebiotic food ingredients containing fructooligosaccha- ride/inulin were incorporated. A nine-point facial hedonic scale was used to evaluate five yogurt samples. A mean score between one and three indicated that the sample product was well accepted. Probiotic yogurt containing onions, garlic and sweet potato received a score of 1.6 ± 0.84 (p 0.90), and plantains, 5.3 ± 2.56 (p > 0.90) were not well accepted. Sweet, mildly flavored prebiotic ingredients were most successfully incorpo- rated into probiotic yogurt in Mwanza.

Optimization of Process Parameters for Spray Drying of Fermented Soy Milk

Generally used as an inexpensive source of high-quality protein with many other health-promoting properties, soy milk offers an interesting alternative when fermented. Fermentation with lactic acid bacteria brings about the value addition to soy milk, making a nutritious probiotic food product. Dehydration helps to achieve longer shelf life and easier transportation and storage, enabling wider distribution of the product. The present work pertains to optimization of process parameters for spray drying of fermented soy milk using response surface methodology (RSM). The process parameters studied include inlet air temperature, aspirator speed (air flow rate), feed flow rate, and atomization pressure. The experiments were designed using the central composite design tool. Residual moisture content was found to be low at higher inlet air temperature, lower feed flow rate, higher atomization pressure, and higher air flow rate. Porosity reduced with increased atomization pressure. Higher product yield was obtained at high air flow rate and low feed flow rate. The increase in the atomization pressure increases the cohesiveness between particles, resulting in reduced flowability. An increase in the inlet air temperature greatly reduces the viability and the isoflavone aglycone content in the product. Protein denaturation during the process was found to reduce the product solubility.

Enhancement of Biological Properties of Soymilk by Fermentation

Soymilk has attracted much interest in the food industry because of its health- promoting properties. Fermentation with lactic acid bacteria is known to provide value addition to soymilk by reducing the beany flavor and content of indigestible oligosaccharides and by enhancing the bioavailability of isoflavones, resulting in a nutritious probiotic food product. In the present study, Soymilk fermentation was studied using Lactobacillus acidophilus strain isolated from ragi, and effect of soymilk supplementation on the fermentation characteristics was also investigated. Viable count of 1.99 × 109 CFU/mL sample and 630.1 U/g of β-glucosidase activity were determined in soymilk after 12 h of fermentation. Soymilk supplementation with skimmed milk powder at 5% level gave best results with viable counts of 22.9 × 109 CFU/ml sample or 212.9 × 109 CFU/g solids; 764 U/g of β-glucosidase activity, 1.44% la titratable acidity and pH of 3.9. Daidzein and genistein profile in fermented soymilk showed that 45.8% and 57.5% of respective isoflavones existing as aglycone form, indicating enhancement of the biological property of soymilk through improvement of health-relevant bioactive forms.

Sensory Analysis: Relevance for Prebiotic, Probiotic, and Synbiotic Product Development.

Sensory analysis represents a decisive step during the various stages of food product development. For probiotic, prebiotic, and synbiotic foods, which have shown continuous and significant consumption in the functional food category, the choice of an appropriate technique allows obtaining relevant sensory information that contributes to consumer acceptance. This review focuses on the importance of sensory analytical techniques in prebiotic and probiotic food product development. Examples of the most known sensory methodologies applied to these important functional foods are presented, as well as some considerations about consumer attitudes that can influence acceptance of these products. Moreover, applications of such techniques on functional product evaluation are provided.

Nutritional altruism and functional food: lay discourses on probiotics

This exploratory study set out to analyse discourse about 'friendly bacteria' by scrutinising data from focus groups convened to discuss participants' reasons for choosing probiotic food products and to set their use of probiotics in context in relation to other food choices. In discussing food choice and dietary patterns, participants in the five focus groups reported here drew on a major interpretive repertoire - that of 'nutritional altruism' in which participants described making choices and preparing food which they believed would benefit others. Those caring for children or shopping for a family member with an illness or disability expressed this imperative to make decisions which would benefit the other most acutely. So far, much research has focused upon the sensory qualities and perceived health benefits for individual consumers, but this study begins to explore the discourses drawn upon when people describe why they might buy food for others. The findings have implications for health education, but also reflect a broader socio-political trend in that risks and benefits are devolved and managed at the level of individual citizens and through processes of individual choice and consumption.

In vitro cholesterol-lowering activity ofLactobacillus plantarumandLactobacillus paracaseistrains isolated from the Italian Castelmagno PDO cheese

The discovery of several healthy beneficial effects of the consumption of dairy products fermented with some bacterial strains led to the investigation of the functional properties of these microorganisms. One of the most studied properties is the cholesterol-lowering activity of bacteria with probiotic characteristics, mostly isolated from human gut. In this work, eight Lactobacillus plantarum and five Lactobacillus paracasei strains isolated from cheese were studied in vitro for their cholesterol-lowering action and their acid and bile tolerance. The ability of these strains to remove cholesterol was assessed in de Man, Rogosa and Sharpe (MRS) medium, supplied with cholesterol, and in ultra-high temperature (UHT) whole homogenized milk. Among all tested strains, two L. plantarum and three L. paracasei strains gave rise to a significant reduction of the cholesterol level in MRS broth; in particular, L. plantarum strains lowered the cholesterol content by an average of 19.4%, whereas L. paracasei strains lowered by an average of 6.8%. The two L. plantarum strains possessing the highest cholesterol-lowering activity in MRS broth were also tested in milk. Results showed that L. plantarum strains maintained this activity because after 24 h the cholesterol decrease ranged from about 5.0% to 8.2% without significant variations between the two strains. Results from the binding assay suggested that cholesterol was mainly removed through the adsorption on the cell wall. Data from acid and bile tolerance assays showed that the L. plantarum dairy isolates were able to maintain viability at pH 2 and to grow in a medium with bile salts, and therefore were regarded as probiotics or dairy starters for new probiotic or functional food production.

Use of mixed cultures for the fermentation of cereal-based substrates with potential probiotic properties

The production of a new cereal-based probiotic foods with suitable aroma, flavor and pH using mixed culture fermentation has been investigated. This required the selection of suitable types of cereal grains and probiotic microorganisms. In a medium of 5% (w/v) malt suspension the effects of yeast presence on the fermentation of a lactic acid bacterium (LAB), Lactobacillus reuteri, was studied. With different inoculum ratios between the yeast and the LAB, the characteristics of the fermentation broth including pH and the contents of free amino nitrogen (FAN), reducing sugar, lactic acid and ethanol were investigated. It was found that LAB growth was enhanced by the introduction of the yeast. In mixed culture broth pH was lowered and the production of lactic acid and ethanol were increased in comparison against pure LAB culture.

Intestinal survival and persistence of probiotic Lactobacillus and Bifidobacterium strains administered in triple-strain yoghurt

The aim of the present study was to investigate the intestinal survival and persistence of probiotic strains Lactobacillus F19, Lactobacillus acidophilus NCFB 1748, and Bifidobacterium animalis subsp. lactis Bb-12 consumed in a yoghurt (ABC product), and also their effect on the intestinal microbiota. Based on the results of culture studies and strain-level analysis by randomly amplified polymorphic DNA (RAPD) fingerprinting Lactobacillus F19 and B. animalis subsp. lactis Bb-12 survived well through the human gastrointestinal tract; they were detected in reasonable numbers in the faeces of 100% and 79% of the study subjects, respectively. Ingestion of the probiotic yoghurt increased transiently the numbers of bifidobacteria and lactobacilli. For lactobacilli the increase was due to the detection of the ingested probiotic strains in faeces, while in bifidobacteria the increase was likely caused by the increase of indigenous bifidobacteria since the ingested Bifidobacterium strain did not comprise the predominant part of bifidobacterial population during the intervention. Probiotic strains were infrequently detected in mucosal biopsy samples. The present study indicates that developing probiotic food products with multiple probiotic strains is feasible.

A Washed-Curd Goat's Cheese as a Vehicle for Delivery of a Potential Probiotic Bacterium: Lactobacillus delbrueckii subsp. lactis UO 004

This study characterizes the probiotic properties of Lactobacillus delbrueckii subsp. lactis UO 004 and examines its suitability for making cheese. This strain was isolated from infant feces and shows interesting features, such as acid and bile tolerance, adherence to intestinal epithelial cells, and inhibition of the growth of certain enteropathogens, that support its potential use as a probiotic strain. In this regard, the suitability of a washed-curd cheese (Vidiago type) made with goat's milk as a delivery system for this probiotic strain was assessed. Lactobacillus delbrueckii subsp. lactis UO 004 was incorporated into a starter culture (IPLA 001). Changes in the overall composition of control and experimental cheeses were determined during ripening through bacteriological, chemical, high-performance liquid chromatography, and gas chromatography analyses. Slight changes in the gross composition and appreciable differences in the flavor compounds profile were observed between control and experimental cheeses. This strain was capable of surviving at high cell numbers (108 to 109 CFU/g) in cheeses after 28 days of ripening without adversely affecting sensory criteria or appearance of the cheese, thus satisfying the criteria for a probiotic food product.

Effect of a Probiotic Infant Formula on Infections in Child Care Centers: Comparison of Two Probiotic Agents

To investigate the effect of 2 different species of probiotics in preventing infections in infants attending child care centers. A double-blind, placebo-controlled, randomized trial was conducted from December 1, 2000, to September 30, 2002, at 14 child care centers in the Beer-Sheva area of Israel in healthy term infants 4 to 10 months old. Infants were assigned randomly to formula supplemented with Bifidobacterium lactis (BB-12), Lactobacillus reuteri (American Type Culture Collection 55730), or no probiotics. Duration of feeding, including follow-up, for each participant was 12 weeks. All infants were fed only the assigned formula and were not breastfed due to parental decision before recruitment to the study. Probiotic or prebiotic food products or supplements were not allowed. Main outcome measures were number of days and number of episodes with fever (>38 degrees C) and number of days and number of episodes with diarrhea or respiratory illness. Participants (n = 201) were similar regarding gestational age, birth weight, gender, and previous breastfeeding. The controls (n = 60), compared with those fed B lactis (n = 73) or L reuteri (n = 68), had significantly more febrile episodes (mean [95% confidence interval]: 0.41 [0.28-0.54] vs 0.27 [0.17-0.37] vs 0.11 [0.04-0.18], respectively). The controls also had more diarrhea episodes (0.31 [0.22-0.40] vs 0.13 [0.05-0.21] vs 0.02 [0.01-0.05], respectively) and episodes of longer duration (0.59 [0.34-0.84] vs 0.37 [0.08-0.66] vs 0.15 [0.12-0.18] days, respectively). The L reuteri group, compared with BB-12 or controls, had a significant decrease of number of days with fever, clinic visits, child care absences, and antibiotic prescriptions. Rate and duration of respiratory illnesses did not differ significantly between groups. Child care infants fed a formula supplemented with L reuteri or B lactis had fewer and shorter episodes of diarrhea, with no effect on respiratory illnesses. These effects were more prominent with L reuteri, which was also the only supplement to improve additional morbidity parameters.

Numbers and strains of lactobacilli in some probiotic products

The numbers and types of Lactobacillus strains in probiotic feed or food products for sale in Europe were compared with the information stated on the product labels. The labels of all 10 products gave information on the genus and species of Lactobacillus, and seven gave information on the numbers of a specified strain(s). Lactobacilli were recovered using a recently validated method, developed as part of the European Community Project SMT4 CT98-2235 for the official control of probiotic microorganisms used as feed additives. Polymerase chain reaction (PCR)-based methods were used to assess the accuracy of labelling with regard to genus and species, and pulsed-field gel electrophoresis (PFGE) was used to identify strains. Five products were mislabelled with respect to the numbers and three with respect to species of lactobacilli. In four cases, the specified strains were not detected. Four fermented milks sold under three trademarks contained the same strain, which was named differently on each label. As safety and functionality of probiotics are strain dependent, these results demonstrated the need to control lactobacilli present in commercially probiotic human food products, not only at the species but also at the strain level, to ensure their quality and protect the consumer. The usefulness of the official methods developed for animal feed additives to identify lactobacilli in probiotic foods for humans was demonstrated.

Differentiation of Enterococcus faecium from Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains by PCR and dot-blot hybridisation

Variations in length and sequence of the 16S/23S spacer region of Enterococcus faecium provided the basis for development of simple PCR and dot-blot hybridisation assays that enabled the differentiation of potentially probiotic Enterococcus faecium strains from Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus. Such assays may be useful for differentiation of yoghurt starter cultures and enterococcal strains when they are simultaneously present in probiotic food products.

Probiotics in foods not containing milk or milk constituents, with special reference to Lactobacillus plantarum 299v

Lactic acid fermentation is the simplest and safest way of preserving food and has probably always been used by humans. Species such as Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus paracasei, Lactobacillus acidophilus, and Lactobacillus salivarius are common in the human mucosa, from the mouth to the rectum. In food, L. paracasei and L. rhamnosus are usually associated with dairy products whereas L. plantarum is found in fermented foods of plant origin. A probiotic food product containing no milk constituent was launched in Sweden in 1994. The product is a lactic acid fermented oatmeal gruel that is mixed in a fruit drink. It contains approximately 5 x 10(10) colony-forming units of L. plantarum 299v/L. The strain L. plantarum 299v originates from the human intestinal mucosa and has been shown in rats to decrease translocation, improve mucosal status, improve liver status, improve the immunologic status of the mucosa, and reduce mucosal inflammation. In humans, L. plantarum 299v can increase the concentration of carboxylic acids in feces and decrease abdominal bloating in patients with irritable bowel disease. It can also decrease fibrinogen concentrations in blood. Should probiotics be administrated through foods, the probiotic organism must remain vigorous in the food until consumption and the food must remain palatable, ie, the food carrier and the organism must suit each other. L. plantarum 299v not only affects the bacterial flora of the intestinal mucosa but may also regulate the host's immunologic defense. The mechanisms involved need to be clarified.

Dietary Bifidobacterium lactis (HN019) enhances resistance to oral Salmonella typhimurium infection in mice.

The ability of a newly identified probiotic lactic acid bacterial strain, Bifidobacterium lactis (HN019), to confer protection against Salmonella typhimurium was investigated in BALB/c mice. Feeding mice with B. lactis conferred a significant degree of protection against single or multiple oral challenge with virulent S. typhimurium, in comparison to control mice that did not receive B. lactis. Protection included a ten-fold increase in survival rate, significantly higher post-challenge food intake and weight gain, and reduced pathogen translocation to visceral tissues (spleen and liver). Furthermore, the degree of pathogen translocation showed a significant inverse correlation with splenic lymphocyte proliferative responses to mitogens, blood and peritoneal cell phagocytic activity and intestinal mucosal anti-S. typhimurium antibody titers in infected mice; all of these immune parameters were enhanced in mice fed B. lactis. Together, these results suggest that dietary B. lactis can provide a significant degree of protection against Salmonella infection by enhancing various parameters of immune function that are relevant to the immunological control of salmonellosis. Thus dietary supplementation with B. lactis provides a unique opportunity for developing immune-enhancing probiotic dairy food products with proven health benefits.

Antiproliferative effects of homogenates derived from five strains of candidate probiotic bacteria.

Unheated and heat-treated homogenates were separately prepared from candidate probiotic bacteria, including Lactobacillus rhamnosus GG, Bifidobacterium lactis, Lactobacillus acidophilus, Lactobacillus delbrueckii subsp. bulgaricus, and Streptococcus thermophilus. We compared the phytohemagglutinin-induced proliferation of mononuclear cells in the presence of homogenates and in the presence of a control containing no homogenate by assessing thymidine incorporation in cell cultures. All homogenates suppressed proliferation, whether the enzymatic activity was inactivated or not inactivated by heating. When the proliferation assays were repeated with cytoplasmic and cell wall extracts derived from the homogenate of L. rhamnosus GG, the cytoplasmic extract but not the cell wall extract was suppressive. These findings indicate that candidate probiotic bacteria possess a heat-stable antiproliferative component(s). These bacteria may be used to generate microbiologically nonviable yet immunologically active probiotic food products that are easier to store and have a longer shelf life.

In VitroStudy of Bile Salt Hydrolase (BSH) Activity of BSH IsogenicLactobacillus plantarum80 Strains and Estimation of Cholesterol Lowering through Enhanced BSH Activity

Growth and bile salt hydrolase (BSH) activity of the isogenic Lactobacillus plantarum 80 (LP80) strains were studied in vitro . In pure culture experiments viability and growth performance of the BSH - LP80 strain was negatively affected by the presence of conjugated bile salts. The LP80 wild type (WT) and BSH overproducing LP80 (pCBHl) strains did not show a die-off upon supplementation of bile salts. The latter strains hydrolysed glyco-conjugated deoxycholate (GDCA) more readily than tauro-conjugated deoxycholate (TDCA), indicating substrate specificity of the enzyme. BSH activities towards TDCA of LP80 WT and LP80 (pCBHl) stationary phase cells were 0.17 and 1.02 μmol/mg CDW.h respectively; activities towards GDCA of the respective strains were 3.52 and 54.80 μmol/mg CDW.h respectively. The study of BSH activity as a function of growth revealed a marked difference in behaviour between LP80 WT and LP80 (pCBHl) with LP80 WT hydrolysing GDCA when reaching the exponential phase, whereas LP80 (pCBHl) immediately started to hydrolyse GDCA. TDCA hydrolysis increased after GDCA hydrolysis was completed. BSH activity of LP80 (pCBHl) in a mixed microbial association, resembling that of the small intestine, was comparable to that determined under pure culture conditions, indicating that BSH activity will probably not be influenced by the presence of the normal intestinal microbiota. Based on the BSH activity of LP80 (pCBHl) and on physiological data on the bile salt-cholesterol metabolism interrelationship, it was calculated that a daily intake of a realistic amount of highly BSH active Lactobacillus cells, e.g. in the form of yoghurt, might lead to a significant reduction of cholesterol. Hence, this in vitro study indicates that altering BSH activity can be a valid (micro) biological alternative treatment for patients with severe hypercholesterolaemia. Keywords - Bile salt hydrolysis, Lactobacilli, Cholesterol, Hypercholesterolaemia, Probiotic food products.