Optimization of saccharification for Obushera production using commercial enzymes
Application of starter cultures in Obushera production has had some draw backs because of low residual sugars resulting into less sweet but sour tasting product that is not so appealing to consumers. This necessitated the optimisation of a mashing process that increases the sugar content prior to starter culture fermentation of Obushera. In this study, response surface methodology (RSM) was used to optimise the effect of four commercial enzymes ( protease, β-glucanase, α-amylase and amyloglucosidase) on reducing sugars, Free Amino Nitrogen (FAN), total soluble solids (Brix) and residual starch using two local sorghum varieties (Seredo and Sekedo). Using optimised enzyme combinations obtained in this study, sugar levels were increased prior to fermentation. Fermentation was then carried out for 24 h to assess the effects of starter cultures composed of lactic acid bacteria: Lactococcus lactis and Lactobacillus planturum, and the yeast, Saccharomyces cerevisiae on the physico-chemical properties of Obushera. The Obushera was prepared using Sekedo and Seredo sorghum varieties. Additonally, the effect of the starters on sensory acceptability of the Obushera was determined. In a further step, this study compared linear, 2FI, quadratic and cubic model based on coefficient of determination (R2), lack of fit (LOF), PRESS and probability value (p-value). The results indicated that the reduced quadratic models fitted better than other models for prediction of reducing sugars, FAN, total soluble solids and residual sugars for both sorghum varieties. Protease, α-amylase and amyloglucosidase were the significant (p<0.05) terms of the reduced quadratic models while β-glucanase had a negligible role (p>0.05) in mashing. This study indicated that application of commercial enzymes significantly (p<0.05) increases mashing parameters (reducing sugars, total solube solids and FAN). The optimum FAN, reducing sugars, total soluble solids, and residual starch levels achieved were 108.3 mg/100 g, 73.9 g/100 g, 18 Brix and 1.46 g/100 g, respectively for Sekedo variety when 0.08 AU-N of protease (at 50°C for 40 min), 40,000 U of α-amylase (at 75°C for 20 min) and 14,000 U of amyloglucosidase (at 55°C for 20 min) were applied to 250 ml of 20% malt sorghum slurry. While for Seredo variety, optimum FAN, reducing sugars, total soluble solids and residual starch levels obtained were 99.3 mg/100 g, 74.8 g/100 g, 17.9 Brix and 1.07 g/100 g respectively when 0.08 AU-N of Protease (at 50°C for 40 min), 40,000 U of α-amylase (at 75°C for 20 min) and 18,000 U of amyloglucosidase (at 55°C for 20 min) were applied to 250 ml of 20% malt sorghum slurry. The pH of enzyme saccharified Obushera prepared from both Sekedo and Seredo variety decreased from 6.3 to 3.7, while lactic acid increased from 1.65 to 8.5 gkg-1 after 24 h of fermentation with LAB used individually or in combination with yeast. Combinations of LAB and yeast starters depleted sugars much faster (p<0.05) than LAB starters alone. Changes in total soluble solids were insignificant (p>0.05) for all samples (17 to 16 °Brix for LAB starters without yeasts and 11 to 10 °Brix for sorghum malt fermented samples) except for samples inoculated with combination of LAB and yeast (17 to 10 °Brix). Sensory evaluation showed that Obushera fermented for 10 h with combination of LAB and yeast starters was more acceptable than that fermented with LAB only or traditionally fermented product obtained after 24 h of fermentation. The results of this study showed that commercial enzymes (protease, α-amylase and amyloglucosidase) can be used to increase fermentable sugars prior to inoculation with starter cultures for production of a more acceptable Obushera product. Ultimately, a safe and more acceptable Obushera product of consistent quality can be obtained under controlled fermentation using a starter combination of LAB (Lactococcus lactis and Lactobacillus. planturum) and the yeast, Saccharomyces cerevisiae.