Biotransformation of Carboxylic Acids to Alcohols: Characterization of Thermoanaerobacter Strain AK152 and 1-Propanol Production via Propionate Reduction

Abstract

Thermoanaerobacter strains have recently gained interest because of their ability to convert short chain fatty acids to alcohols using actively growing cells. Thermoanaerobacter thermohydrosulfuricus strain AK152 was physiologically investigated for its ethanol and other alcohol formation. The temperature and pH optimum of the strain was 70 °C and pH 7.0 and the strain degraded a variety of compounds present in lignocellulosic biomass like monosaccharides, disaccharides, and starch. The strain is highly ethanologenic, producing up to 86% of the theoretical ethanol yield form hexoses. Strain AK152 was inhibited by relatively low initial substrate (30 mM) concentration, leading to inefficient degradation of glucose and levelling up of all end-product formation. The present study shows that the strain produces alcohols from most of the tested carboxylic acids, with the highest yields for propionate conversion to propanol (40.7%) with kinetic studies demonstrating that the maximum conversion happens within the first 48 h of fermentation. Various physiological tests were performed to maximize the acid conversion to the alcohol which reveals that the optimum pH for propionate conversion is pH 6.7 which affords a 57.3% conversion. Kinetic studies reveal that propionate conversion is rapid, achieving a maximum conversion within the first 48 h of fermentation. Finally, by using ¹³C NMR, it was shown that the addition of propionate indeed converted to propanol.

Keywords: biocatalysis; bioreduction; carboxylic acids; extremophile; fusel alcohols; thermophile; volatile fatty acids.

Figure 1

Growth characteristics of Thermoanaerobacter strain AK152; influence of cultivation temperature at pH 7.0 (A) and pH at 70 °C (B).

Figure 1

Growth characteristics of Thermoanaerobacter strain AK152; influence of cultivation temperature at pH 7.0 (A) and pH at 70 °C (B).

Figure 2

End product formation after 5 days by Thermoanaerobacter strain AK152 for selected carbon sources (65 °C, pH 7.0) at a concentration of 20 mM with the exception of polymeric substrates which were provided at 0.2% w/v. Values represent average of triplicate fermentations with standard deviation presented at error bars.

Figure 3

Influence of initial glucose concentration (A) and liquid-gas phase ratio (using 20 mM glucose) (B) on fermentation by Thermoanaerobacter strain AK152. Fermentations were performed at 65 °C and pH 7.0 with end products being quantified after 5 days. Values represent average ± standard deviation (n = 3).

Figure 4

Conversion of short-chain fatty acids by Thermoanaerobacter strain AK152 using glucose (20 mM) as a source of reducing potential. Values represent average ± standard deviation (n = 3).

Figure 5

Influence of initial propionate concentration at 65 °C and pH 7.0; (A) and initial pH with 20 mM propionate concentration; (B) during glucose (20 mM) fermentation by Thermoanaerobacter strain AK152 (65 °C. Values represent the average of triplicates with standard deviation presented as error bars.

Figure 6

Fermentation of 20 mM glucose (A), 20 mM glucose + 20 mM propionate (B), 20 mM glucose + 50 mM propionate (C), 20 mM glucose + 100 mM (D) by Thermoanaerobacter strain AK152. Values represent the average of triplicates ± standard deviation.

Figure 6

Fermentation of 20 mM glucose (A), 20 mM glucose + 20 mM propionate (B), 20 mM glucose + 50 mM propionate (C), 20 mM glucose + 100 mM (D) by Thermoanaerobacter strain AK152. Values represent the average of triplicates ± standard deviation.

Figure 7

¹³C NMR spectra for the reduction of ¹³C1-labeled propionate by Thermoanaerobacter strain AK152 after 5 days of fermentation with glucose (20 mM) at 65 °C, pH 7.0.