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United States Department of Agriculture

Agricultural Research Service

Research Project: INDUSTRIALLY ROBUST ENZYMES AND MICROORGANISMS FOR PRODUCTION OF SUGARS AND ETHANOL FROM AGRICULTURAL BIOMASS Title: Anaerobic conversion of lignocellulosic corn fiber to butyric acid, a substrate for microbial butanol production

Authors
item Agler, Matthew - CORNELL UNIV
item Iten, Loren
item Qureshi, Nasib
item Cotta, Michael
item Dien, Bruce
item Angenent, Largus - CORNELL UNIV

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: July 22, 2009
Publication Date: N/A

Technical Abstract: Many factors, including sharply fluctuating fuel prices and questions regarding the sustainability of fuel produced from potential food crops, have bolstered interest in renewable fuels from alternative feedstocks. We tested pretreated and nonpretreated corn fiber for its susceptibility to hydrolysis and subsequent fermentation in 5-L thermophilic (55 deg C) anaerobic reactors by nondefined mixed cultures. The three methods of fiber pretreatment were all performed in fluidized sand bed reactors at 15% corn fiber w/v in water at 160 deg C for 20 min, and 0.5% w/v H2SO4; 1:10 CaO to biomass ratio; and hot water only. Four identical anaerobic sequencing batch reactors (ASBRs) were fed every two days, each always receiving one of the three pretreated substrates or an untreated substrate. The reactors are operated at environmental conditions selected to "direct" electron flow towards butyric acid production while maximizing hydrolysis. A low pH of 5.5 disrupts methanogenesis and is near the optimum for butyric acid formation. We have shown that the levels of nitrates in the pretreated substrates, presumably formed during pretreatment, also contribute to inhibition of methane generation from acetate. In addition, we found that butyric acid formation begins after inhibition of hydrogenotrophic methanogenesis (at pH 5.5, inhibition occurred when total volatile fatty acids reached ~8500 mg as CH3COOH/L), when hydrogen levels become measurable in the headspace, and continues to rise correlated to hydrogen evolution. This coincides with the finding from thermodynamic modeling that NADH oxidation by H+ reduction can only occur at very low hydrogen concentrations (e.g., during hydrogen uptake by methanogens). Thus far, we have achieved butyrate levels of 3.25, 3.57, 2.76, and 1.06 g/L at total volatile fatty acid levels of 8.2, 11.6, 7.0, and 6.3 g as CH3COOH/L in reactors fed acid, base, hot water, and untreated substrates, respectively. Thermodynamic analysis also shows that acetic acid reduction to ethanol with hydrogen as electron donor may occur (negative delta G) at the conditions in the reactors. Indeed, significant quantities of ethanol have been found (4.60, 5.35, 4.58, and 1.49 g/L in reactors treating acid, base, hot water, and untreated substrate, respectively), although much of it likely comes directly in the pathway through acetyl-CoA. Thermodynamic calculations also show that under conditions of lower butyrate concentrations, acetate and ethanol could be further converted to butyrate by the pathway employed by certain microorganisms common in mixed cultures, including Clostridium kluyveri. Because of the consistent environmental pressure on the systems, we expect development of microbial communities possessing optimum characteristics for degradation of substrate. The diverse communities may have the ability to remove any biologically inhibiting compounds formed during pretreatment (usually derived from lignin), and tests are currently being conducted to evaluate this possibility.

Last Modified: 4/16/2014