Fiber Evaluation for Prebiotic Effects and Fermentation Kinetics - Sustainable Sources and Commercial Products

Abstract

Dietary fibers are a broad subset of non-digestible carbohydrates that have many health benefits to consumers. When consumed in adequate amounts they can protect against cardiovascular disease, decrease the risk of diabetes, aid in weight loss and numerous other health benefits. One unique attribute that dietary fibers have is that when they reach the distal intestine or colon, some of these fibers can be fermented by the hundreds of species of bacteria present. When these fibers provide growth to the beneficial bacteria present in the gastrointestinal tract, they are referred to as prebiotics, or prebiotic dietary fibers. As the understanding of the gut micobiota and the gut microbiome expand, the importance of stimulating the beneficial microbiota present in the gut becomes increasingly more important. Because the human diet is so complex, understanding the fermentation process of individual compounds can be a difficult task. The use of in vitro fermentation models is one way to understand the fermentation of dietary fibers, and whether or not they support the growth of beneficial taxa, and very closely mimics the function of the colon in that regard. Incubating fecal bacteria from human donors, exposing them to specific dietary fibers, and measuring their fermentation differences provides a snapshot of how these compounds may ferment in vivo. The primary objective of this research was to conduct a preliminary in vitro analysis of two emerging dietary fibers (wheat dextrin and partially hydrolyzed guar gum) to test their prebiotic capacity (Chapter 2), and to compare new dietary fibers (Oatwell, xylooligosaccharides, beta-glucans) to an established prebiotic dietary fiber (inulin) to measure key fermentation differences in vitro (Chapter 5). The secondary objective of this research was to compare fermentation differences between donors exposed to these same compounds by measuring differences in the production of short chain fatty acids (Chapter 3), and the microbiota that were stimulated (Chapter 4). The final objective of this research was to determine if conventionally or organically grown plants in Minnesota provided a significant amount of dietary fiber, if it was fermentable, and whether or not the growing differences in these plants affected key nutrients and compounds, including: vitamin C, potassium, iron, sodium, calcium and nitrate-nitrogen. Both wheat dextrin and partially hydrolyzed guar gum stimulated growth of two beneficial genera of bacteria, Lactobacillus and Bifidobacterium, indicating that both fibers are bifidogenic and lactogenic within 24 h of fermentation (Chapter 2). At 12 h wheat dextrin was significantly more bifidogenic (9.50 CFU log10/mL) than partially hydrolyzed guar gum (PHGG) (9.30 CFU log10/mL) (p=0.052), and also at 24 h wheat dextrin (9.41 CFU log10/mL) compared with PHGG (9.27 CFU log10/mL) (p=0.043). Wheat dextrin produced less total short chain fatty acids at both 12 h and 24 h than partially hydrolyzed guar gum, and produced significantly lower amounts of gas at 12 h and 24 h (p<0.001, p<0.001), a key marker for gastrointestinal tolerance. Changes in short chain fatty acid concentration (SCFA) due to the fermentation of dietary fibers in the colon has been widely studied, but there are limited studies analyzing the differences in this production across multiple individuals (ages 23-68) exposed to the same dietary fiber (Chapter 3). The objective of this project was to look at fermentation differences from six fecal donors all exposed to partially hydrolyzed guar gum in an in vitro model. With the six donors analyzed in this study, gas production varied from 59-80 mL/0.5g fiber at 12 h and 85-93 mL/0.5g fiber at 24 h between the six donors. At 12 h butyrate concentrations varied from 6.99 µmol/mL to 23.84 µmol/mL and from 8.78 µmol/mL to 22.84µmol/mL at 24 h. Total SCFA concentration at 24 h ranged from 42.85 µmol/mL to 91.17 µmol/mL. With over a 2-fold difference in SCFA production, significant differences were found between healthy individuals exposed to partially hydrolyzed guar gum in vitro. Following this analysis, 16s rRNA sequencing was used to identify the fecal microbiota responsible for the fermentation of partially hydrolyzed guar gum (Chapter 4) comparing samples at baseline, 12 h and 24 h post-exposure. Parabacteroides increased from 3.48% of sequence reads to 10.62% of sequence reads after 24 h (p = 0.0181) and Bacteroidetes increased from 45.89% of sequence reads to 50.29% of sequence reads (p = 0.0008). Partially hydrolyzed guar gum stimulates growth of Parabacteroides, a genus of bacteria that have been inversely associated with irritable bowel syndrome (IBS) and ulcerative colitis. PHGG provides stimulation of beneficial Bacteroidetes (Bacteroides and Parabacteroides), which may be correlated with many positive health markers and outcomes in vivo. Five other dietary fibers were analyzed to test their prebiotic capacity and fermentation differences, including: Oatwell, inulin, beta-glucan, xylooligosaccharide and a dried chicory root blend. Oatwell had the highest production of propionate at 12 h (4.76 μmol/mL) compared to inulin, dried chicory root blend and xylooligosaccharide samples (p<0.03). Its effect was similar to those of the beta-glucan samples. Oatwell and beta- glucan samples promoted the highest mean propionate production at 24 h. Xylooligosaccharide resulted in a significant increase in the genus Bifidobacterium after 24 h of fermentation (0 h: 0.67 OTUs; 24 h: 5.22 OTUs; p = 0.038). Inulin and the dried chicory root blend increased the beneficial genus Collinsella, consistent with findings in clinical studies. All prebiotic dietary fibers studied promoted the formation of beneficial markers due to the fermentation of each specific compound. All compounds provided different, significant fermentation patterns, and all provided beneficial effects that would promote host health in vivo. Produce can provide a substantial amount of dietary fiber in the consumer diet, along with many other nutrients. Arugula (Eruca sativa), mizuna (Brassica rapa var. nipponsinca), red giant mustard (Brassica juncea) and spinach (Spinacia oleracea ‘Tyee’) are fresh produce crops high in nutritive value and provide shortfall and high interest nutrients addressed in the 2015 U.S. Dietary Guidelines. The primary objective of this project was to evaluate fertility treatments unique to these crops that optimize their nutritional capacity. Plants were grown using five different fertility treatments, including four organic treatments and one conventional control. The plant treatment combinations were replicated three times and the entire experiment was duplicated. Fertility treatments had a high impact on vitamin C (with over a 3-fold difference in treatments in the first experiment), nitrate (over 10-fold difference among fertility treatments in some species) and potassium concentrations (over 5-fold difference among fertility treatments in some species) in analyzed plant tissue. No consistent differences were found for fiber, calcium, iron and sodium concentrations in tissue analyzed. This is the first study to analyze the impact that different production treatments can have on multiple deficient nutrients and compounds addressed by the U.S. Dietary Guidelines for high-impact, highly-consumed produce crops. Based on in vitro assays, partially hydrolyzed guar gum, wheat dextrin, and xylooligosaccharides stimulate the growth of Lactobacilli and Bifidobacteria, the two most beneficial genera of bacteria in the gastrointestinal tract. All dietary fibers analyzed resulted in significant amounts of short chain fatty acids being produced from their fermentation, which was found in all three in vitro studies. Although they are all fermentable by fecal bacteria, within healthy donors there can be over a 2-fold total difference in acetate, propionate and butyrate production within 24 h of fermentation for these dietary fibers. For consumers who prefer to consume dietary fiber in whole foods instead of supplements or fortified products, red giant mustard and mizuna both offer between 2-4g fiber/serving, which is a “good source of dietary fiber” for the consumer. This includes both organic and conventionally grown red giant mustard and mizuna. Like any other dietary component, moderation and variety is still the most important factor to consider. Different dietary fibers all support different functions and roles in the body, and may all be fermented differently depending on the consumer. Whole foods, fortified foods and supplements may all play a critical role in developing a healthy gut microbiome, and may all be needed for consumers to meet their recommended daily intake.