Characterization of an Alkaline Amylase and a Pectinase From Alkaliphilic Bacillus Halodurans Lbw 5117 Isolated From Lake Bogoria in Kenya and Demonstration of Their Applications in Bio-processing Woven Cotton

Abstract

De-sizing is a key textile wet processing step that is carried out on sized woven cotton to remove the starch impurity from cloth. On the other hand, scouring is done on de-sized fabrics to remove the natural pectins and other non-cellulosic impurities from the cotton fibers of the cloth. Both processes are done to obtain sufficiently hydrophilic cellulosic fibers that will allow the cloth to be evenly bleached, mercerized, and dyed/printed in the subsequent processes. Both desizing and scouring are commercially carried out using conventional chemicals or enzymes. Although the chemicals are cheap and effective in removing impurities, they are environmentally unfriendly, and can also damage the fabrics. On the other hand, using enzymes offers better advantages e.g., they produce higher-quality fabrics and generate fewer pollution loads. However, most of the enzymes operate at or near neutral pH, making their use risky due to possible damage to the fabrics by cellulases from neutrophilic contaminants. In addition, the requisite enzymes are expensive because they are often high-temperature enzymes that have to be imported from China, Denmark, and the US. To address these challenges, low-temperature alkaline enzymes from indigenous alkaliphilic microorganisms should be sought and utilized as an alternative to both the costly commercial enzymes and the fabric-damaging and environmentally unfriendly chemicals. The study aimed to characterize an alkaline protease, pectinase, and amylase obtained from an alkaliphilic bacterium that was isolated from Lake Bogoria (soda lake), Kenya, and to demonstrate their potential to desize (amylase) and scour (pectinase or protease) woven cotton. The study began with the isolation and identification of alkaliphilic microbial soda lake isolates that exhibit protease-, pectinase- and amylase- activities. 16 isolates were identified as B. halodurans and 2 as B. pseudofirmus. All the B. halodurans and B. pseudofirmus isolates produced amylases while pectinases were produced by all the B. halodurans isolates. On the other hand, proteases were produced by 12 B. halodurans and all the B. pseudofirmus isolates. Among the isolates, Bacillus halodurans LBW 5117 was ranked as one of the most potent producers of both amylases (0.32 U/ml) and pectinases (0.09 U/ml). This bacterium was

therefore cultured for the individual production and characterization of both enzymes (amylase = Amy LBW 5117 and pectinase = PGase LBW 5117). Amy LBW 5117 was an endo-α-1-4-amylase while PGase LBW 5117 was an endo-polygalacturonase.The optimum storage condition for Amy LBW 4117 was in liquid form at 4 oC for six weeks while that for PGase LBW 5117 was at 4 - 30 oC for one year. On the other hand, the optimum operating conditions for Amy LBW 5117 was pH 10.0 at 60 oC in the presence of 1.0 mM Ca2 and 0.05 mM Tween 20 while that for PGase LBW 5117 was pH 10.5 at 50 oC in the presence of 1.5 mM Ca2+ and 0.05 mM Tween 20. In addition, these additives also enhanced the thermo-stability property of both enzymes, with Amy LBW 5117 retaining 96 % of its original activity after 3 h of incubation at 60 C and PGase LBW 5117 retaining 200 % of its activity after 8 h of incubation.at 50 oC. Furthermore, metal ions that are commonly found in cotton fibers and tap water either stimulated the catalytic activities of both enzymes or had no significant effect on them. Moreover, Amy LBW 5117 was cellulase-free and could hydrolyze the different types of starch products that are used to make textile sizing agents. Similarly, PGase LBW 5117 was cellulase-free and could hydrolyze different pectins with various degrees of methyl esters that are found in cotton fibers. Indeed, a preliminary desizing application study of Amy LBW 5117 revealed that it could desize woven cotton that contained starch as the size and yield a fabric with a commercially acceptable amount of residual starch (0.0725%) (TEGEWA rating 7.5). Similarly, PGase LBW 5117 could remove pectin and other non-cellulosic impurities from an Amy LBW 5117 desized fabric and yield a cloth with improved wettability (drop test = 10 sec) and dye-ability (capillary rise test = 28 mm after 30 min) properties. This study demonstrates that Amy LBW 5117 and PGase LBW 5117 have great potential to be effective textile desizing and scouring agents due to their good operational properties.