The Development of a Biorefining Strategy for the Conversion of Wheat Straw and Sorghum Bran to Value Added Products

Renewable energy, such as biofuel has been highlighted as a future fuel that could replace fossil fuels. The conflict between biofuel and food security has encouraged the research on the conversion of lignocellulosic biomass into biofuels. Although lignocellulosic biomass is abundant, the presence of lignin and the cost of enzymes have caused several major issues in regards to the commercialization of lignocellulosic biofuels.

The aim of this study was to investigate the feasibility of using wheat straw and sorghum bran to produce value added products, such as enzymes and bioethanol. The utilization of wheat straw for cellulase production and the subsequent hydrolysis was investigated. Six fungal strains (Aspergillus niger N402, Aspergillus niger N403, Aspergillus niger CKB, Trichoderma reesei R32, Trichoderma reesei R33 or Rhizomucor variabilis RS) were investigated using both solid-state fermentation (SSF) and submerged fermentation (SmF).

In SSF, cellulase production increased from 3.2±0.05 FPU/g to 8.1±0.3 FPU/g (Filter Paper Unit) when wheat straw was modified using alkali treatment. The addition of starch improved the cellulase production with a cellulase activity of 23.14±0.09 FPU/g being obtained when 0.04% starch was added. The inoculum and reactor size also affected cellulase production. A. niger N402 with an inoculation ratio of 1x107 spores/g resulted in the highest cellulase activity of 55.93 FPU/g and 30.43 FPU/g in SSF using Petri Dish and 250 mL shake flask, respectively.

The optimisation of cellulase production using a newly isolated fungal strain, R. variabilis (RS) was performed in both submerged fermentation (SmF) and solid-state fermentation (SSF). The impact of various parameters, including pH, mineral addition, nitrogen source, temperature, and substrate concentration, was investigated for SmF and incubation time, pH, temperature, inoculation size, moisture content, and nitrogen source were investigated for SSF. An optimum fermentation condition was determined to be: pH 6.5, 0.03% tryptone and fermentation for 3 days for SmF, a cellulase activity of 18.44 FPU/g was obtained. Similarly, an optimum fermentation condition for SSF was determined to be: pH 7, 28°C, inoculation size of 1×107 spores per g substrate, 0.03% tryptone and fermentation for 5 days. The cellulase activity was 30.19 FPU/g. Response Surface Methodology (RSM) was used to further optimize cellulase activity in SmF and SSF. This approach resulted in cellulase activity of 23.81 FPU/g for SmF and 24.80 FPU/g for SSF. Two rounds of physical mutagenesis of RS strain were carried out using UV lights and microwave heat. A mutant strain MW15-03 was obtained, which showed 21.6% higher cellulase production capacity in comparison with the parent strain.

Sorghum bran, a starch rich food processing waste, was investigated for the production of glucoamylase in SmF and SSF. The fermentation parameters, such as cultivation time, substrate concentration, pH, aeration rate, inoculation ratio, temperature, nitrogen source, and mineral addition were investigated for SmF. The glucoamylase activity was improved from 1.90 U/mL in an initial test to 19.26 U/mL at 10% substrate concentration, pH 6, fermentation volume 200 mL in 500 mL shaking flask and fermentation of 3 days. RSM was used to further optimize glucoamylase activity in SmF and glucoamylase activity of 59.03 U/mL was achieved at the following conditions: substrate concentration 8%, pH 6, yeast extract concentration 5 g/L and fermentation volume 100 mL in 250 mL shaking flasks. Larger scale production of glucoamylase enzyme in 2 L bioreactors under the optimum condition resulted in 21.67 U/mL of glucoamylase activity at 72 hours of fermentation, while further increasing sorghum bran concentration to 12.5% gave an improved glucoamylase activity of 37.55 U/mL at 115 hours of the fermentation.

The crude glucoamylase solution was used for the enzymatic hydrolysis of the sorghum bran. A sorghum bran hydrolysis carried out at 200 rpm, 55°C for 48 hours at a substrate loading ratio of 80 g/L resulted in 11.74 g/L glucose, which was comparable to that obtained using a commercial enzyme (12.72 g/L). Larger scale sorghum bran hydrolysis in 2 L bioreactors with crude glucoamylase enzyme resulted in a glucose concentration of 38.7 g/L from 200 g/L sorghum bran.

Wheat straw hydrolysate, sorghum processing wastewater and sorghum bran hydrolysate were used as substrates for the production of bioethanol. The addition of minerals accelerated the rate of yeast fermentation. Marine yeast strain W. anomalus M15 resulted in a very high ethanol yield of 49.79%. Upto 19.3 g/L bioethanol was obtained. Autoclaved wheat straw at 121°C for 15 minutes gave the highest ethanol yield of 16.95% using the marine yeast W. anomalus M15.