Study of carotenoid production by Iranian halophilic bacterial isolates and effects of various factors on production level

Abstract

Introduction: Carotenoids are organic pigments with significant commercial interests, which are used in neutraceuticals, pharmaceuticals, cosmetics and foods. Considering the importance of carotenoids, in the present study we aimed to study carotenoid producing microorganisms from Urmia hypersaline and Gorigol fresh water lakes in North-western of Iran and evaluating effects of various factors on production level. Methods: In the first stage totally 30 halophilic prokaryotic microorganisms isolated from Urmia hypersaline and Gorigol fresh water lakes in North-western of Iran, were cultivated in culture media and incubated at appropriate conditions. Then single colonies were cultivated in broth media. In order to study the carotenoid production and also the production profile, total carotenoid was extracted using acetone-methanol solution (7:3 v/v). Then, carotenoid content of the extracts was evaluated using UV spectroscopy and confirmed by thin layer chromatography (TLC) in the presence of antimony pentachloride (SbCl5). The production profile was analyzed using liquid-chromatography mass spectroscopy (LC-MS) techniques. The biochemical and molecular characterization of the isolates was studied by the conventional methods used for bacterial systematic and 16S rRNA gene sequence analysis. Finally, the analysis of environmental factors (temperature, pH and salinity) through response surface methodology (RSM) on the biomass and carotenoid production of three isolates was done. In addition the effect of light on cell growth and carotenoid production in optimized condition was evaluated. Results: The isolated strain TBZ112 exhibited the highest carotenoid-producing ability followed by TBZ110 and TBZ123. On the basis of their phenotypic and chemotaxonomic characteristics, all 3 strains were assigned to the genus Halorubrum. Total carotenoid extracts from all selected strains exhibited the same absorption spectra and chromatographic carotenoid profiles. The LC-MS analytical results indicated that the produced carotenoids that were bacterioruberin, lycopene, and β-carotene, among which bacterioruberin were predominant. The optimum conditions for both cell growth and carotenoid production in TBZ112, TBZ110 and TBZ126 cultures determined by RSM, were about temperature 31 ºC & 32 ºC, pH 7.5 & 7.9, and NaCl concentration (w/v) 18.33% & 20%, respectively. In addition the study indicates that all strains synthesize higher amounts of biomass and total carotenoids in the light than in the dark. Conclusion: As a consequence, all identified strains isolated from Urmia Lake have high capacity in the production of carotenoids. These extremely halophilic archaea could be considered as prokaryotic candidates for carotenoid production source for future studies. In addition, using optimization with RSM and light as an inducing factor in this study, 38%, 70% and 45% increase in total carotenoid production by TBZ110, TBZ112 and TBZ126 respectively was obtained as compared to the un-optimized medium.