Studies On The Secondary Metabolites Of Halotolerant Talented Strains Under Salt Stress

Halotolerant microorganism has been considered as an important and promising type of extremophiles, possessing special ecology, specific physiological mechanism and extensive utilization. The special hypersaline environment where they lived could active silent gene and induces other specific metabolic pathway. Therefore, we planned to obtain more novel bioactive compounds from the secondary metabolites of halotolerant talented strains under salt stress. Studies include the screening for halotolerant talented strains, isolation and structural elucidation of secondary metabolites, preliminary bioactivity evaluation, and the salt effects on the chemodiversity of secondary metabolites.1. Screening for the halotolerant talented strains.120 strains of halotolerant fungi have been isolated from salt sediments in Putian sea salt field, Fujian and mangrove environment, Hainan. The EtOAc extracts of these fungi were evaluated with the combinatory method of chemical and bioactive screening.4 of them were determined as halotolerant talented strains of cytotoxicity or antibiotic activities.2. Study on bioactive constituents of four halotolerant talented strains under salt stressAfter optimization of fermentation conditions under salt stress, these strains were fermented, respectively. The whole broths were extracted with ethyl acetate to give active extracts, which were subjected to extensive silica gel column chromatography, Sephadex LH-20 and HPLC.86 compounds were isolated from four halotolerant talented strains under salt stress,80 structures of which (see Fig.0-1) were determined by means of spectral analysis (UV, IR, NMR, MS, X-Ray) and chemical methods. Among them,32 compounds were new ones, including two novel cyclic hexapeptides (1 and 2) obtained from Aspergillus sclerotiorum PT06-1 cultured in a nutrient-limited medium containing 10% salts, twelve cyclic tripeptides (10-21) and a dipeptide derivative (23) from PT06-1 cultured in a nutrient-suffient medium containing 10% salts, two anthranilate condensation compound (29 and 30) and butyrolactone I analog (33) from Aspergillus terreus PT06-2 cultured in a hypersaline medium, three steroids (47-49) and a red pyrrole-containing pigment (51) from Aspergillus flocculosus PT05-1 cultured with 10% sea salt, ten drimane-type sesquiterpenes (58,59,66-73) from Aspergillus candidus 085241 cultured with 1% sea salt. Compounds 2,3,32,47 and 48 exhibited weak cytotoxicity against HL-60 cell line with IC50 values of 56,36,58,14, and 12μM, respectively; 48 also had weak cytotoxicity against BEL-7402 cell line with an IC50 value of 18μM. Compounds 1,2, 10,13,16,18,20,29,30,31,47,48 and 49 showed antibiotic activities against Candida albicans, Pseudomonas aeruginosa, Enterobacter aerogenes or Staphylococcus aureus with MIC values ranged from 1.6μM to 64μM. Additionally, two known compounds 3 and 43 were evaluated with significant antiviral activity against H1N1 virus in vitro with IC50 values of 4.6 and 6.6μM, respectively.3. Preliminary exploration of halide salt effects on chemodiversity of secondary metabolites from halotolerant strainsTo further explore the halide salt effects on chemodiversity of secondary metabolites of halotolerant talented strains, secondary metabolites from 9 halotolerant talented strains under different culture conditions containing different halide salts at different concentration were analyzed by comparing their weight, TLC, HPLC and bioactivities. The strain HK01 was found to mainly produce bioactive penicillenol (81) cultured in a hypersaline medium. In addition, we established that the different halide salts were also able to influence the secondary metabolites of halotolerant strains.In a word, novel bioactive compounds can be obtained from secondary metabolites of halotolerant strains under salt stress, and more nitrogen-containing compounds produced when stains cultured in the hypersaline media. It was supposed that the specific products probably be metabolized by halotolerant microorganisms to relieve or resist the salt stress from enviroment. Above studies provided a good example for obtaining bioactive novel compounds from halotolerant strains.