Semester of Degree Completion

Spring 2018

Thesis Director

Steven L. Daniel

Thesis Committee Member

Gary A. Bulla

Thesis Committee Member

Kai Hung

Abstract

Bile acids (cholate and chenodeoxycholate) are synthesized from cholesterol in the human liver, and secreted in small intestine for food digestion. However, a portion of these primary bile acids is converted in the colon to toxic secondary bile acids, deoxycholate and lithocholate. Secondary bile acid formation is the result of 7α-dehydroxylating anaerobes, and one of the key players in this conversion is Clostridium scindens. Interestingly, little is known about the basic physiology and nutrition of C. scindens. Therefore, the goal of this research was to understand the nutritional requirement, and determine the 7α-dehydroxylation activity and end products from glucose fermentation in different growth conditions by C. scindens ATCC 35704. C. scindens was maintained in anaerobic defined medium (DM) containing 25 mM glucose, minerals, trace metals, amino acids, vitamins, bicarbonate, 100% CO2 gas phase, and sodium sulfide. To determine the amino acid and vitamin requirements of C. scindens, the "leave one [amino acid group] out" technique and "leave one [vitamin] out" techniques were used to eliminate the non-essential amino acids and vitamins. With these techniques, it was found that an amino acid, tryptophan, and three vitamins (riboflavin, pantothenate, and pyridoxal) were required for growth of C. scindens. C. scindens was able to convert cholate to deoxycholate along with some bile acid-derived products in defined as well as in minimal growth conditions. Also, C. scindens fermented glucose mainly to ethanol, acetate, formate, and H2. The ratio of these end products varied with growth conditions. Understanding the metabolic profile of C. scindens ATCC 35704 may help in treatment of disease associated to secondary bile acids.

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