School of Agriculture, Food and Ecosystem Sciences - Theses

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Start date: 2021 × Type: PhD thesis × Subject: Bifidobacterium × Subject: Gut microbiota ×

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    Healthy Chocolates Enriched with Microencapsulated Probiotics
    HOSSAIN, MD NUR ( 2022)
    Probiotics are live microorganisms that can modulate the gastrointestinal environment and confer health benefits to the host when ingested in sufficient numbers. However, maintaining probiotic viability with the minimum recommended number during processing, storage, and exposure to the harsh gastrointestinal tract is very challenging in food and pharmaceutical product development. Encapsulation has been considered an effective technique to improve probiotic viability, and various polymers including, alginate, casein, pectin, chitosan, xanthan, and gelatin have been used as encapsulants. In this study, the probiotics were encapsulated using a complex admixture of cocoa mass along with Sodium alginate and fructooligosaccharides to extend the probiotic’s storage viability and survivability during the gastrointestinal transit. Cocoa powder or chocolates is a complex mixture of polysaccharides, lipids, polyphenols, and other organic compounds that are not readily digestible and absorbed in the gastrointestinal tract. The aim of this PhD work was to investigate cocoa powder as an encapsulating ingredient to overcome the major challenges of processing and storage conditions and adverse gastrointestinal conditions. Furthermore, the project was involved with the formulation of healthy probiotic chocolates which can modulate the gut environment to produce some essential metabolites. Two different types of chocolates (70 and 45 percent cocoa chocolates) and eight different probiotics named Lactobacillus casei 431, Lactobacillus rhamnosus, Lactobacillus plantarum UALp 05, Lactobacillus sanfranciscensis JCM5668, Lactobacillus acidophilus, Bifidobacterium animalis subsp. lactis, Streptococcus thermophilus UASt 09, and Lactobacillus delbrueckii subsp. bulgaricus 12315 were used in the entire study. The results revealed excellent improvement in the survivability of probiotics and demonstrated their role in modulating the gut environment and the production of short chain fatty acids and vitamin B12 and the bioconversion of polyphenols during the in vitro gastrointestinal digestion and colonic fermentation. The coca powder admixture was used to encapsulate probiotics and compared with commonly used encapsulating materials such as whey protein concentrate, hi maize resistant starch, and skim milk powder with an emulsion based freeze drying technique. The encapsulation efficiency of the cocoa powder along with sodium alginate and fructooligosaccharides showed very promising results as compared with whey protein concentrate, hi maize resistant starch, skim milk, and sodium alginate. Encapsulating Lactobacillus rhamnosus, Lactobacillus casei 431, Lactobacillus plantarum UALp 05, Lactobacillus sanfranciscensis JCM5668, and Bifidobacterium animalis subsp. lactis using cocoa powder along with sodium alginate and fructooligosaccharides demonstrated a very good yield and a high rate of survivability. The best viabilities of encapsulated probiotics in chocolates during storage were found to be 120 days at 4 degrees celsius and 90 days at room temperature. Additionally, a high survivability rate of the cocoa powder encapsulated probiotics was detected at 60 degrees celsius which could be excellent findings to scale up the production of probiotic chocolates industrially. The in vitro gastrointestinal digestion and colonic fermentation of two types of probiotic chocolates with 70 and 45 percent cocoa chocolates contents showed positive effects on the gut environment modulating activity. All tested probiotic chocolates demonstrated very promising results concerning the bioaccessible polyphenols and bioconversion activity, production of fecal metabolites such as short-chain fatty acids (acetic, propionic, isobutyric, butyric, and isovaleric acid), and biosynthesis of vitamin B12 (Lactobacillus sanfranciscensis and Lactobacillus plantarum). However, the interaction among the various tested probiotics and chocolate polyphenols (45 and 70 percent cocoa) enriched with these encapsulated probiotics during the in vitro gastrointestinal digestion and colonic fermentation revealed different levels of bioconversion of polyphenols (bioaccessible polyphenols). For example, Lactobacillus sanfranciscensis JCM5668 and Steptococcus thermophilus UASt 09 demonstrated better conversion activities for epicatechin, procyanidin B1, and procyanidin B2 in probiotic chocolate containing 70 perccent cocoa, while Lactobacillus casei 431, Lactobacillus rhamnosus, Lactobacillus plantarum UALp 05 showed better capacities with 45 percent cocoa chocolates. These findings concluded that chocolates could be utilized by probiotics for their metabolic activities and modulate the gut environment which improved the functionality of chocolates. Furthermore, investigating the impact of in vitro gastrointestinal digestion and colonic fermentation of probiotic chocolate on the production of short chain fatty acids revealed a direct relationship between the probiotics and the released short chain fatty acids. The acetic acid production rate was much higher for Lactobacillus plantarum UALp 05 and Bifidobacterium animalis subsp. lactis than other probiotics at 24 to 48 hours. While Lactobacillus sanfranciscensis JCM5668 and Streptococcus thermophilus UASt 09 produced fewer amounts of acetic acids. Similarly, different amounts of propionic acid were produced by different probiotics after 48 hours of colonic fermentation and ranked from largest to smallest as follows: Bifidobacterium animalis> Lactobacillus acidophilus> Streptococcus thermophilus> Lactobacillus sanfranciscensis> Lactobacillus bulgaricus> Lactobacillus casei> Lactobacillus rhamnosus> Lactobacillus plantarum irrespective of cocoa content. The best isobutyric acid efficacy was estimated in probiotic-chocolate fortified with Streptococcus thermophilus followed by Lactobacillus sanfranciscensis, Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus casei containing 45 percent cocoa chocolates at 24 hours of colonic fermentation. Lactobacillus acidophilus was the most efficient producer of butyric acid and released significantly higher amounts than all other probiotics throughout the colonic fermentation. However, Lactobacillus acidophilus produced a smaller amount of both isobutyric and isovaleric acids. Surprisingly, 70 percent of dark chocolate samples produced a higher amount of branched short chain fatty such as isobutyric and isovaleric acid. These secreted short chain fatty acids during the in vitro gastrointestinal digestion and colonic fermentation of probiotic chocolates may indicate that chocolate could act as prebiotics for the gut microbiota, which then produced more short chain fatty acids. Vitamin B12 was the other assessed metabolite. All eight tested probiotics with both types of chocolates (45 and 70 percent cocoa) were investigated for the biosynthesis of vitamin B12. Results showed that in vitro colonic fermentation produced vitamin B12 only in the presence of Lactobacillus plantarum UALp 05, Lactobacillus sanfranciscensis JCM5668. Additionally, these probiotics produced more B12 in the presence of 45 percent cocoa chocolate than 70 percent dark chocolate. The sensory tasting using a 9 point hedonic scale and five sensory attributes (taste, color, flavor, texture, and overall acceptability) of the formulated probiotic chocolates containing 45 percent and 70 percent cocoa powder showed insignificant differences between these two types of probiotic chocolates. Consequently, it was concluded that both probiotic chocolate with 45 percent and 70 percent cocoa contents could be attractive to consumers. Such probiotic chocolate will offer consumers a new source of healthy snacks with additional health benefits. This thesis presents promising research on probiotics with chocolate as a carrier and expands the research window for probiotic functional products using non dairy chocolates as carriers. It is recommended that conducting a follow up study on probiotic chocolate at an industrial scale with in vivo applications will provide stronger proof of the functional properties and health benefits of such a healthy chocolate snack.