#TITLE_ALTERNATIVE#

• Main Author: ROSTIYANTI (NIM: 20514301), DEIS
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/26462
• Institution: Institut Teknologi Bandung

Utilization of microorganism in the application of biotechnology to industrial scale is growing. One type of microorganisms that are often exploited because of its several advantages are halophile bacteria. Halophilic bacteria is one type of extremophile microorganisms that can survive in environments with high salinity. One strategy of halophilic bacterial adaptation in the face of environments with high salinity is by accumulating osmolyte in the cytoplasm in the form of the organic compound with a low molecular weight to offset the osmotic pressure exerted from outside of the cells. Ectoine is one osmolyte found in some types of halophilic bacteria that function as a stabilizer of biomolecules against dehydration, freezing, heating dan drought. In addition, ectoine is also known to maintain three-dimensional structure of macromolecules. <br /> <br /> This study focused on the isolation, characterization and application of ectoine produced by halophilic bacterial isolates from Bledug Kuwu mud crater, Purwodadi, Central Java. Selection of seven Bledug Kuwu-bacterial isolates obtained one halophilic Gram-negative bacteria, which have been identified as Halomonas elongata BK AB4-DR, as the most potential ectoin producer. The bacteria was then grown for 25 hours at a temperature of 37°C with 150 rpm of aeration in MM63 media containing 9% NaCl but without the addition of yeast extract. After this growing period, 85 mg of dry cell weight containing 22 mg of ectoin (26% of yield) was obtained. The molecular weight and chemical structure of the isolated ectoine samples were identified and characterized using HPLC, FTIR, LC-MS, and 1H-NMR. FTIR spectra of the samples showed a pattern of vibrational spectrum for NH, CN and C = N groups, which is a functional group for ectoin, that observed at the wave number of 1404 cm-1, 1622 cm-1 and 1656 cm-1. Further elucidation of the structure was carried out by 1H-NMR and LCMS, and for the comparison, we also carried out the structural analysis for a commercial ectoin. The elucidation results verified that the sample was ectoin with a molecular weight of 142 g/mol and its IUPAC name is 1,4,5,6-tetrahydro-2-methyl-4-pyrimidinecarboxylic acid. For applications, we evaluated the role of ectoin as a protective structure by studying the stability of Serum Bovine Albumin (BSA) under thermal perturbation. Spectroscopic analysis using fluorescence spectroscopy showed that the addition of ectoin with a final concentration of 1 x 10-3 M did not significantly improved the thermal stability of BSA. Further analysis using Circular Dichroism (CD), however, revealed that the presence of ectoin can maintain thermal stability of BSA structure at the level of secondary structure. Our study thus showed that ectoine is playing role to protect secondary structure of proteins by preventing aggregation due to thermal perturbations.