Streptomyces sp. No. 4 produce two forms of amylase that attack raw cassava starch. Both forms, amylase‐1 and amylase‐2, were purified by starch adsorption, affinity chromatography, and ion exchange chromatography. The molecular weights of amylase‐1 and amylase‐2 as determined by SDS‐PAGE were 56 kD and 77 kD, respectively. Optimal enzyme activities occurred at pH 5.5 and at 50°C for amylase‐1 and at 45°C for amylase‐2. The activation energy of amylase‐1 and amylase‐2 were 67 and 42 kJ/mol, respectively. Hg2+ and pCMB inhibited both enzymes, whereas 2‐mercaptoethanol activated only amylase‐2. EDTA inhibited amylase‐1 but activated amylase‐2. The main product of hydrolysis of raw cassava starch by amylase‐1 was maltose, followed by maltotriose, maltotetraose and dextrin. Amylase‐2 cleaved raw cassava starch to produce glucose and maltose as main products. Both amylase‐1 and amylase‐2 are α‐amylases, as shown by the fast disappearance of iodine staining, the corresponding reaction products and the ability of both enzymes to hydrolyze crosslinked blue starch.
Cassava pulp, a low cost solid byproduct of cassava starch industry, has been proposed as a high potential ethanolic fermentation substrate due to its high residual starch level, low ash content and small particle size of the lignocellulosic fibers. As the economic feasibility depends on complete degradation of the polysaccharides to fermentable glucose, the comparative hydrolytic potential of cassava pulp by six commercial enzymes were studied. Raw cassava pulp (12% w/v, particle size <320 µm) hydrolyzed by both commercial pectinolytic (1) and amylolytic (2) enzymes cocktail, yielded 70.06% DE. Hydrothermal treatment of cassava pulp enhanced its susceptibility to enzymatic cleavageas compared to non-hydrothermal treatment raw cassava pulp. Hydrothermal pretreatment has shown that a glucoamylase (3) was the most effective enzyme for hydrolysis process of cassava pulp at temperature 65 °C or 95 °C for 10 min and yielded approximately 86.22% and 90.18% DE, respectively. Enzymatic pretreatment increased cassava pulp vulnerability to cellulase attacks. The optimum conditions for enzymatic pretreatment of 30% (w/v) cassava pulp by a potent cellulolytic/ hemicellulolytic enzyme (4) was achieves at 50 °C for 3, meanwhile for liquefaction and saccharification by a thermo-stable α-amylase (5) was achieved at 95 °C for 1 and a glucoamylase (3) at 50 °C for 24 hours, respectively, yielded a reducing sugar level up to 94,1% DE. The high yield of glucose indicates the potential use of enzymatic-hydrothermally treated cassava pulp as a cheap substrate for ethanol production.
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