Evaluating efficacy of non-thermal microbial load reduction treatments on native resistant pulse starch sources for human gut microbiome research
Open Access
- Author:
- Paff, Andy
- Graduate Program:
- Food Science
- Degree:
- Master of Science
- Document Type:
- Master Thesis
- Date of Defense:
- October 03, 2022
- Committee Members:
- Robert Roberts, Program Head/Chair
Darrell William Cockburn, Thesis Advisor/Co-Advisor
Edward G Dudley, Committee Member
Greg Ziegler, Committee Member - Keywords:
- Pulse
microbiome
gut
human
bleach
hydrogen peroxide
alcohol
16S rRNA - Abstract:
- In vitro simulated colonic fermentation offers a cost-effective option for screening the impact of foods and drugs on the gut microbiota prior to clinical studies. A drawback of the model is increased susceptibility to microbial contamination which may influence outcomes and conclusions. Although a simulated gastric and intestinal digestion may address some concern through pH and dilution, viable microbes may remain as contaminants. Autoclaving is generally an option for sterilization, however, not when the item of concern is sensitive to moist heat. In this study, liquid chemical sterilant alternatives to moist heat sterilization were explored for treating pulse flours for use in in vitro fermentations with and without human gut microbiota derived from a human donor’s feces. Key attributes considered in chemical selection were accessibility, safety, cost, and effectiveness of treatments. Three chemicals were evaluated (bleach, alcohol, and hydrogen peroxide) at varying concentrations, as well as dry autoclaving, on green lentil, field pea, chickpea, or sprouted field pea flours. Production of flours and sprouting of green lentil was performed in lab. In vitro simulated colonic fermentations were performed in a 96-deep well plate format with or without agitation for 24 hours. Acetate and butyrate from fermentation supernatants were measured by high pressure liquid chromatography as an indicator of community function. 16S rRNA sequencing was used to evaluate community structure in pulse fermentation with and without feces inclusion. In fecal-free control fermentations, Erwinia comprised over 50% of the relative abundance of microbes present at >0.1% relative abundance in three of the flours and was also the only pulse contaminant found at significant levels in fecal fermentation with pulse flour. All chemical treatments greatly decreased Erwinia, demonstrating effectiveness in addressing main contaminants in the pulse flours evaluated. Anaerobic plate counts were used to evaluate effectiveness of the sterilization treatments in reducing microbial load in pulse flours. While all treatments reduced viable counts, only hydrogen peroxide and bleach treatments reduced microbial load below detectable levels. Scanning electron microscopy was used to evaluate the impact of sterilization treatments on the starches and were generally found to have minimal effects on starch granule structure. The results suggest there are a variety of chemical treatments useful in microbial load reduction in pulse flours without adversely affecting behavior of simulated colon fermentation.