Organic solvent resistant microorganism which secretes lipolytic enzyme

MARTINEZ, M ALEJANDRA; TORRES, SEBASTIÁN; DEL CASTILLO, LOURDES; CASTRO, GRUILLERMO R

Orlando, Florida

Congreso; 101st General Meeting; 2001 - American Society for Microbiology (ASM); 2001

American Society for Microbiology (ASM)

Control/Tracking Number: GM01-A-33690-ASMActivity: AbstractOrganic Solvent Resistant Microorganism which Secretes Lipolytic EnzymeLipases are hydrolases acting on water poor soluble substrates such as triglycerides and esters. Non-aqueous enzymology has become an area of interest to chemical and pharmaceutical industries. Recent reports have shown that enzymes produced by organic solvent resistant microorganisms were stable in presence of organic solvents. The aim of this work was to isolate thermoresistant and organic solvent resistant microorganisms that produce lipolytic enzyme.Heat shocked soil samples (80oC, 10 min.) were cultured in LB medium supplemented with NH4NO3 and tributyrin at 55oC. Screening for microorganisms lipolytic activity was performed on tributyrin agar. Organic solvent resistant bacteria were select in synthetic liquid medium with isoamyl alcohol (from 0,1 to 1%) at 55oC. Enzyme activity was assayed using p-nitrophenyl derivatives. Effects of organic solvents on hydrolytic activity were carried out using six hydrophylic solvents. Strain characterization was made using biochemical and molecular approaches (API system and PCR-based techniques: 16S amplification, sequence analysis, ITS-PCR and t-RNA-PCR).Considering the highest lipolytic activity and organic solvent resistance (it was able to grow in presence of 0.6 % of isoamyl alcohol) strain S86 was selected. From morphological, biochemical and molecular characteristics, strain S86 was identified as Bacillus licheniformis. Optimal growth temperature was 50oC (m = 3.12 h-1 in medium without solvent; m = 1.52 h-1 in medium with 0.4 % of isoamyl alcohol). Enzyme production was maximal at the stationary phase in both media (with and without isoamyl alcohol), and it was the highest in the medium with 0.4% organic solvent at 50oC (5.37 U/mg). Relative enzyme activities at 50 % solvent composition were in glycerol (81%), ethanol (54%), isopropanol (50%), 1,3 propanediol (38%), acetone (22%) and propanol (4%). Optimal temperature for enzyme activity was between 60 to 70oC.Because of its organic solvent resistance and high activity, the enzyme from Bacillus licheniformis S86 can be used as biocatalytic in non-aqueous media.