Progress 06/01/24 to 05/31/25
Outputs
Target Audience:The target audience includes academic and government researchers, as well as industry members. The research updates were disseminated through peer-reviewed manuscripts, conference presentations, and industry outreach events at the University of California, Davis. In addition, collaboration was developed with the US-BARD project with Volcani Institute in Isreal focused on exploring a byproduct of plant products for the delivery of probiotics. Changes/Problems:We are requesting a one-year no-cost extension to complete the pending tasks as described in the plan for the coming reporting year and the manuscripts. What opportunities for training and professional development has the project provided?The project has partially supported the training of three postdoctoral fellows and two graduate students (one MS and one PhD student). The project has also supported the training of three undergraduate students in diverse aspects of this project including chemical and microbial analysis as well as development of biomaterial scaffolds. How have the results been disseminated to communities of interest?The research results were disseminated to communities of interest through a peer-reviewed manuscriptand presentations to industry stakeholders at theUC Davis campus. In addition, the USDA-NIFA funding also enabled a collaboration with Volcani Institute in Isreal. This collaborative effort has been supported through the US-Isreal BARD project. What do you plan to do during the next reporting period to accomplish the goals?1. In-vivo study to evaluate the influence of the probiotic biofilms in food-derived 3D scaffolds for reducing high-fat diet-induced inflammation 2. In-vivo evaluation of the in-situ fermentation-derived 3d gel scaffolds for the delivery of the probiotics 3. Complete and submitthree manuscripts that are in progress based on accomplishments in this reporting period.
Impacts
What was accomplished under these goals?
3D scaffolds for enhanced stability and survivability of Probiotics:3D scaffolds for plant by-products to enhance stability and delivery of probiotics: The research developed a 3D scaffold made from pomegranate mesocarp and evaluated the survival of encapsulated probiotic bacteria through the distinct phases of digestion--gastric digestion, intestinal digestion, and colonic fermentation.Lacticaseibacillus rhamnosusGG (LGG) andLactiplantibacillus plantarumwere encapsulated into scaffolds made of either decellularized tissue or untreated pomegranate pomace tissue.The scaffolds were further incubated in MRS to increase total cell volume and encourage the growth of biofilm.Cell viability results ranged from 8 to 9 Log CFU/g post-digestion, suggesting that a pomegranate tissue-based scaffold could effectively enhance the survival of probiotic bacteria in the GI tract. The survival of bacteria in combination with the plant pomacewas at least 4 orders of magnitude higher compared to the cells without encapsulation. The developed scaffold suggests a promising approach for enhancing the delivery of probiotic bacteria to the gut. Future studies will evaluate the in-vivo potential of the plant by-product derived scaffold. In-vivo validation of the apple-derived 3D scaffold for enhancing delivery and persistence of probiotics: In our previous report we described the in-vitro assessment of apple-derived 3D scaffold for enhancing the gastro-intestinal stability of probiotic cells. In this study, the probiotic cells were cultured to form in-situ biofilm in an apple-derived 3D scaffold.During this year, we have validated the effectiveness of the apple-derived 3D scaffold to enhance the delivery and persistence of probiotic cells in vivo. The results of the in-vivo study demonstrate that the probiotic biofilms in apple tissue enhanced the persistence of probiotic cells compared to unencapsulated controls by at least 3logs (3 orders of magnitude higher) across the small intestine, cecum, and colon after 72 hours of the initial feeding. These probiotic cells delivered in the form of biofilms on apple scaffoldalso achieved enhanced attachment to the mucosal tissues in the cecum and colon, indicating the potential for these probiotic cells to influencegut health. A future in-vivo study will be conducted to evaluate the influence of this composition on the high-fat diet-induced inflammation. Role of probiotic cell interactions with the food grade 3D scaffold in improving survivability during simulated GI delivery and storage stability: We havedeveloped an in-situ fermentation process for the encapsulation of Lacticaseibacillus rhamnosus GG (LGG) in a mixed protein/starch gel, as an approach to protect the bacterium during storage and digestion and as a model to investigate the effect of cell-to-encapsulation matrix interactions on such protection. This studyevaluated the role of the pili of LGG in both gel formation and the effectiveness of the gel in maintaining cell viability during simulated digestion, as well as the ability of the hydrated gels to stabilize the bacteria during unrefrigerated storage. A rapid in-situ gel formation occurred at room temperature after inoculating the polymeric mixture with LGG, regardless of the pilation of the cells. Our results suggest that the pili-dependent retention of LGG within the gels partially explains the protection that the gels provided to the encapsulated cells during simulated gastrointestinal digestion. These results enhance our understanding of how bacterial-to-encapsulation matrix interactions may influence the survival of probiotics during delivery. Finally, the hydrated gels sustained high counts of LGG during two months of unrefrigerated storage. This, to our knowledge, is the first study to demonstrate enhanced stability of probiotics in a hydrated state using food-grade biopolymers.
Publications
- Type:
Peer Reviewed Journal Articles
Status:
Published
Year Published:
2025
Citation:
Influence of the pili of Lacticaseibacillus rhamnosus GG on its encapsulation and survival in mixed protein-starch gels assembled by in-situ fermentation, Applied Environmental Microbiology
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Progress 06/01/23 to 05/31/24
Outputs
Target Audience:The results were disseminated through peer-reviewed publications and meeting presentations, including the Gordon Research Conference and CoFE. In addition, the PI also shared the results with industry members of companies through the campus venture catalyst group and events such as Natural Product Expo. Changes/Problems:There are delays due to the recruitment of postdoctoral scholars to support this project. We plan to submita no-cost extension request. What opportunities for training and professional development has the project provided?The project provided training opportunities to a graduate student, twopostdoctoral fellows, and one project scientist. In addition 4 undergraduate researchers have been engaged in research activities during this period. How have the results been disseminated to communities of interest?The research results were shared through scientific conferences, peer-reviewed publications, and engagement with industries through public events such as Natural product expo presentations by the PI. What do you plan to do during the next reporting period to accomplish the goals?The goals of the next phase are: 1. Complete the pending manuscripts from the year two studies 2. Assessment of the ability of 3D scaffolds and soluble oligosaccharides derived from black beans spent fractions to promote growth and mucoadhesion of probiotics in a simulated colonic environment 3. Optimization of the probiotic biofilm formation on plant-based scaffolds and evaluation of in-vivo persistence
Impacts
What was accomplished under these goals?
Novel food products rich in probiotics and prebiotics have significant potential to impact human health. However, the limited viability of probiotics during delivery to the gut and the lack of persistence of these bacteria in the gut significantly limits the success of these novel products. The harsh gastrointestinal conditions reduce the delivery of viable probiotics in food products, and the lack of persistence and growth is constrained by the poor mucoadhesion and availability of selective prebiotics, respectively. To address these challenges our interdisciplinary team is developing plant derived 3D scaffolds for the delivery of probiotics and enhance their persistence in-vivo. In this project period, our interdisciplinary team has made progress in the following goals: (a) Compositional analysis of probiotic biofilm in apple tissue: In our previous report we reported on our discovery of the unique potential of 3D scaffolds with probiotic biofilm to significantly enhance both the survivability and persistence of probiotic cells in-vitro and in-vivo. In this period, we have evaluated the composition of these probiotic biofilms using FTIR spectroscopy and LC-QQQ MS. The results of this analyses reveal significant increase in microbial biomass and associated extracellular biopolymer. The biopolymer fraction was significantly enriched in polysaccharide fraction and the protein fractions based on the results of IR measurement. For the LC-QQQ MS, the team has been optimizing the extraction and characterization process, and developing the analytical standards and the derivatization steps with 1-pheny-3-methyl-5-pyrazolone (PMP) methanolic solution for analysis of the carbohydrate composition of these biofilms. Fourteen monosaccharide standards were successfully resolved chromatographically in a 25 min binary gradient after derivatization, covering the full range of potential building blocks for the biopolymer. Mass spectrometry analysis revealed a varied composition, dominated by glucose, followed by galactose and xylose in comparable quantities, alongside with arabinose and mannose, and lastly, rhamnose and fucose found in lower abundances. (b) Mechanistic understanding of probiotic delivery and the role of structural properties: Many of the probiotic delivery approaches have been evaluated using empirical approaches. To develop a more fundamental understanding, our team has initiated mechanistic modeling of transport, persistence, and attachment of bacteria in the gut. The current model simulates the coupled movement of chyme and the mass transport of bacteria in the gut, including factors such as bulk mobility due to peristalsis and binding interactions with mucin and epithelial tissues. Imaging measurements were conducted to characterize the residual fractions or structure of the 3D scaffold in the gut. For this characterization, the samples of intestinal and cecal isolates from an animal study were imaged. The results reveal a distinct fiber-like morphology in the 3D scaffold in the small intestine, with distributed bacteria along the fiber length. In our next steps, we plan to evaluate the role of these fiber-like morphologies in influencing the distribution of the bacteria in the gut. (c) Optimization of extraction processes from beans and characterization of the spent fractions: One of the goals of this project is to utilize other by-products from the plant protein extraction industries to develop scaffolds for the delivery of probiotics. In this regard our team has been optimizing the extraction process from ground black beans and chickpeas. When these fractions were extracted using a conventional alkaline extraction method (AEP) widely used in the industry to obtain protein concentrates and/or isolates. The TPE of the black bean and chickpea extractions were 19.2 ± 1.5% and 25.3 ± 1.7%, demonstrating that a majority of the pulse proteins remained in the insoluble fraction. The poor extractability is likely due to the large particle size of the ground pulses used in this study. Comparatively, when black bean and chickpea flour were used and extraction was performed under the same conditions, protein extraction yields of 75% (black bean) and 63% (chickpea) were achieved. The residual fractions rich in carbohydrates and residual protein will be further evaluated to develop 3D scaffolds.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Nitin N, Plant Biomaterials for Pathogen Control and Promoting Delivery of Probiotics
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2024
Citation:
Biofilm scaffolds for the delivery of probiotics, COFE, 2024
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2024
Citation:
3d scaffolds for the delivery of probiotics, Gordon Research Conference, Lactic acid bacteria,
- Type:
Journal Articles
Status:
Published
Year Published:
2024
Citation:
1. Yang, J. S., Dias, F. F. G., Pham, T. T. K., Barile, D., & de Moura Bell, J. M. L. N. (2024). A sequential fractionation approach to understanding the physicochemical and functional properties of aqueous and enzyme-assisted aqueous extracted black bean proteins. Food Hydrocolloids, 146, 109250. https://doi.org/10.1016/j.foodhyd.2023.109250
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Progress 06/01/22 to 05/31/23
Outputs
Target Audience:The target audience includes food manufacturing andnutritional product companies. In addition, the target audience also includes researchers in academic and government laboratories. In this period, the target audiences reached by our efforts include both industry participants and researchers in academic and government laboratories. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project provided training opportunities to a graduate student and a postdoctoral fellow. The postdoctoral fellow was promoted to project scientist in this review period also. How have the results been disseminated to communities of interest?The results were disseminated through peer-reviewed publications and presentations at meetings, including COFE and ACS meetings. In addition,the PI also shared the results with industry members of companies, including Kraft Heinz and Nestle, during their visit to the UCD campus. What do you plan to do during the next reporting period to accomplish the goals?Aim 2:Assessment of the ability of 3D scaffoldsand soluble oligosaccharides derived from apples and black beans spent fractions to promote growth and mucoadhesion of probiotics in a simulated colonic environment Aim 3: Evaluation ofin-vivopersistence and growth of probiotics delivered using the optimizedcompositions of 3Dscaffolds containing probiotics and oligosaccharides and their influence on high-fat diet-induced inflammation
Impacts
What was accomplished under these goals?
Assessment of the enhanced survivability of probiotics during simulatedin vitrodigestion using 3D scaffolds derived from apple and black beans extraction byproducts and also promote growth of bacteria in-vitro. In the first year, we focused on evaluating the enhanced survivability of probiotics using 3D scaffolds derived from plant by- products. Our first set of efforts was focused on Apple by-products. This study develops a novel low-cost microbial delivery system by transforming common food materials such as apple tissue into a 3D scaffold. Apple tissue scaffold was constructed by decellularization of intact tissue using a minimal amount of sodium dodecyl sulfate (0.5 % w/v). Vacuum-assisted infusion of model probiotic Lactobacillus cells led to a high encapsulation yield of probiotic cells (1010 CFU/g of scaffold) in 3D scaffolds on a wet basis. The bio-polymer coated 3D scaffolds with infused cells significantly enhanced the survivability of infused probiotic cells during simulated gastric and intestinal digestions. In addition, imaging and plate counting results validate the growth of the infused cells in the 3D scaffold after 1-2 days of fermentation in MRS media, while cells without infusion in the scaffold had limited attachment with the intact apple tissue. Overall, these results highlight the potential of the apple tissue-derived 3D scaffold to deliver probiotic cells and include the biochemical compositions to support the growth of delivered microbial cells in the colon.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Apple-derived 3D scaffold for improving gastrointestinal viability and in-situ growth of probiotics
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Plant derived compositions for improved delivery and persistence of probiotics in the gut, COFE, 2022
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