Screening of Factors Influencing Keratinase Fermentation from Bacillus Haynesii BK1H using The Plackett-Burman Design (PBD)

Departement of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Malang, Malang, East Java 65145, Indonesia

^* Corresponding author: suharti.fmipa@um.ac.id

Abstract

Keratinase is a class of proteases that degrade keratin into polypeptides and amino acids by breaking peptide and disulfide bonds in keratinous proteins. Protease is one of the largest industrial enzymes, the global protease market is increasing rapidly every year. In previous studies, a keratinase-producing microbe was isolated from Bleduk Kuwu which was identified as Bacillus haynesii BK1H. However, further study needs to be done to optimize the production by observing microenvironmental factors that influence keratinase production. In this study, screening some microenvironmental factors is reported. Screening factors including carbon sources, type and concentration of metal ions, agitation speed, amount of inoculum, pH, and temperature. Screening for these factors was started with the One Factor at A Time (OFAT) method and followed by the Plackett-Burman Design (PBD) method. In this study, sequential work was done: (1) Regeneration of Bacillus haynesii BK1H Microbes, (2) Preparation of Tyrosine Standard Series Solutions, (3) Screening of Factors Affecting Protease Fermentation of Bacillus haynesii BK1H using the One Factor at A Time (OFAT) method. ), and (4) Maintaining Significantly Influential Factors by Using Plackett-Burman Design (PBD). The results of the OFAT approach showed that the best condition for keratinase production was achieved at rice husk concentration, additional carbon source, of 1%; pH of 7; a temperature of 35°C; the amount of inoculum of 1%; agitation speed of 150 rpm; magnesium sulfate concentration of 0.04 g/mL, and calcium chloride concentration of 0.0005 g/m. Justification of those factors using PBD confirmed that only additional rice husk, magnesium sulfate and calcium chloride concentration, and agitation speed were significantly important toward keratinase production at selected experiment level limits.