Comparative Chemical and Bioactivity Studies of Intra- and Extracellular Metabolites of Endophytic Bacteria, Bacillus subtilis NCIB 3610

Endophytic bacteria are able to produce unique bioactive compounds for various biotechnological applications. The intracellular and extracellular extracts of the endophytic bacterium, Bacillus subtilis NCIB 3610 were investigated for chemical composition as well as evaluated for antiviral, antimicrobial, antifungal and cytotoxic activities. The GC/MS analysis of intracellular and extracellular hexane extracts revealed that 2,6-di-t-butyl-4-methyl phenol was found as main compound in intracellular hexane extract, while pentacosane and hexacosane as well as 14-methyl hexadecanoic methyl ester were found as major metabolites in extracellular hexane extract. The intracellular and extracellular polysaccharides were isolated from aqueous extract which characterized as heteropolysaccharides bounded with protein. The yield of intracellular polysaccharide fraction (FC1) (42.17% +/- 1.21) was higher than that of the extracellular polysaccharides fractions (FM1, FM2 and FM3) (12.68%). The GLC analysis of fractions revealed the presence of 8, 8, 6 and 9 monosaccharides, respectively, the fractions FC1, FM2 and FM3 composed mainly of galactose, mannose and glucose with weight-average molecular weight (Mw) 295.5, 165.5 and 129.8 kDa, respectively. The amino acid analysis of protein bounded to intra- and extracellular polysaccharide fractions revealed the presence of 17 amino acids. Glutamic acid and alanine were found as predominant amino acids in FC1, while glycine, glutamic and aspartic acids were existed as dominant amino acids in exopolysaccharide fractions. The anti-bacterial activity displayed that intracellular polysaccharide (FC1) has a promising antibacterial activity against Staphylococcus aureus and Streptococcus pneumoniae which gave 51.6 and 37% relative inhibition activity of Doxycyclin Hydrchloride. Moreover, the most fractions of intracellular and extracellular showed broad spectrum antifungal activity. The evaluation of cytotoxic activity of intracellular and extracellular fractions on HCT116, HepG2 and MCF7 human cell lines showed that intracellular chloroform, ethyl acetate and methanol fractions have a potent cytotoxic activity. Overall, the bioactive fractions obtained from Bacillus subtilis NCIB 3610 can be useful for biotechnological and pharmaceutical application as they showed cytotoxic and antimicrobial activities.