Abstract
BackgroundThe main challenge of cassava-based biobutanol production is to enhance the simultaneous saccharification and fermentation with high hyperamylolytic activity and butanol yield. Manipulation of cofactor [e.g., Ca2+ and NAD/(P)H] levels as a potential tool to modulate carbon flux plays a key role in the cassava hydrolysis capacity and butanol productivity. Here, we aimed to develop a technology for enhancing butanol production with simultaneous hydrolysis of cassava (a typical model as a non-cereal starchy material) using a cofactor-dependent modulation method to maximize the production efficacy of biobutanol by Clostridium sp. stain BOH3.ResultsSupplementing CaCO3 to the medium containing cassava significantly promotes activities of α-amylase responsible for cassava hydrolysis and butanol production due to the role of Ca2+ cofactor-dependent pathway in conversion of cassava starch to reducing sugar and its buffering capacity. Also, after applying redox modulation with l-tryptophan (a precursor as de novo synthesis of NADH and NADPH), the levels of cofactor NADH and NADPH increased significantly by 67 % in the native cofactor-dependent system of the wild-type Clostridium sp. stain BOH3. Increasing availability of NADH and NADPH improved activities of NADH- and NADPH-dependent butanol dehydrogenases, and thus could selectively open the valve of carbon flux toward the more reduced product, butanol, against the more oxidized acid or acetone products. By combining CaCO3 and l-tryptophan, 17.8 g/L butanol with a yield of 30 % and a productivity of 0.25 g/L h was obtained with a hydrolytic capacity of 88 % towards cassava in a defined medium. The metabolic patterns were shifted towards more reduced metabolites as reflected by higher butanol–acetone ratio (76 %) and butanol–bioacid ratio (500 %).ConclusionsThe strategy of altering enzyme cofactor supply may provide an alternative tool to enhance the stimulation of saccharification and fermentation in a cofactor-dependent production system. While genetic engineering focuses on strain improvement to enhance butanol production, cofactor technology can fully exploit the productivity of a strain and maximize the production efficiency.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-015-0351-7) contains supplementary material, which is available to authorized users.
Highlights
The main challenge of cassava-based biobutanol production is to enhance the simultaneous saccharification and fermentation with high hyperamylolytic activity and butanol yield
Through using a Ca2+ probe fluorescence assay, we investigated if calcium exists in the α-amylase as a metal cofactor and there are increases in intracellular Ca2+ levels in Clostridium sp. strain BOH3 when it is growing in Ca2+-supplemented medium compared to the control medium
Ca2+ was detected in pure α-amylase secreted by culture BOH3 without Ca2+ treatment, suggesting Ca2+ is a metal cofactor for extracellular α-amylase of culture BOH3 (Fig. 1a)
Summary
The main challenge of cassava-based biobutanol production is to enhance the simultaneous saccharification and fermentation with high hyperamylolytic activity and butanol yield. We aimed to develop a technology for enhancing butanol production with simultaneous hydrolysis of cassava (a typical model as a non-cereal starchy material) using a cofactor-dependent modulation method to maximize the production efficacy of biobutanol by Clostridium sp. Li et al Biotechnol Biofuels (2015) 8:166 as a substrate (i.e., substrate cost), and the potential for culture degeneration, which made the industrial-scale fermentation-derived butanol less competitive when compared to the petroleum-based butanol production [1, 5]. To improve the economic competitiveness of biobutanol production, from fermentation substrate point of view, the availability of an inexpensive raw material is essential as the cost of the substrate is one of the most important factors affecting the market price of butanol [6, 7]. Cassava represents an alternative cheap carbon source for industrial production of butanol, which is promising and attractive in both economic and geographical considerations
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