Preliminary Studies On Secondary Metabolites Of Four Fungal Strains With Siderophore Activity

Siderophore is a kind of low molecular iron chelate with high affinity to Fe3+produced by most microorganisms. According to the literature survey for nearly10years, the study of eukaryotic microorganism siderophore is limited; The study of siderophore from terrestrial alkaline environment microbial has not been reported yet. Therefore, for the siderophore from terrestrial alkaline environment fungi, the preliminary work was carried on.In this paper, based on the principle of the CAS screening procedure,99fungi strains isolated from Cheng hai (an alkaline lakes in yunnan province) and DaTun alkaline tailings (Ge jiu, yunnan province) were screened at different levels. After primary screening, total36strains with the Fe3+chelating activity were selected and the4strains (strain "FEDT-866","FEDT-145","FECH-998" and "FECH-595") with the high siderophore activity were determined for further study after the secondary screening.Later, by the methods as "CAS double-layer plates","TLC-CAS in-situ chromogenic reaction" and "spectrophotometric detection", the siderophore production ability of the above4strains was verified; The effect of "resin in-situ adsorption" for siderophore activity substances during the liquid cultivation process of4strains was examined and the preliminary molecular identification of the strains was carried on.Based on the results of the preliminary molecular identification, the strains FEDT-866(with higher similarity with Aspergillus tubigensis) and the strains FECH-988(with higher similarity with Aspergillus nomius) were identified as the microorganism Aspergillus. The strain FECH-595(with higher similarity with Penicillium svalbardense) and the strain FEDT-145(with higher similarity with Penicillium chrysogenum) were identified as the microorganism Penicillium.In compound separation stage, the strain FECH-595, strain FECH-998,and strain FEDT-145were cultivated with Czapek’s media without iron addition to20L respectively; The strain FEDT-866were cultivated with Czapek’s media without iron addition to1L. After series of procedures as silica gel chromatography, CHP20P adsorption, Sephadex-LH20chromatography, crystallization and recrystallization, HPLC detection and HPLC preparation,30compounds were obtained from the cultivation crude extract or cultivation supernatant. Among these compounds, the structure of14were identified and8of them were determined as the novel ones. From the cultivation supernatant of strain FED-866, two fractions (FS-01and FS-02) with the strong siderophore activity were obtained and the structure elucidation is ongoing.From the cultivation crude extract of strain FECH-998, one pure compounds (FS-03) with the strong siderophore activity were obtained and the structure was determined as the kojic acid.From the cultivation crude extract of strain FECH-595,13compounds were isolated. The structures of6compounds were elucidated:Compound FS-07(4-(2-hydroxyethyl)phenol);Compound FS-08(O-methyl (dihydroxyacetyl)prop-1-en-2-ylcarbamothioate); Compound FS-10(4-methyl-8-azabicyclo[3.2.1]oct-5(8)-ene-2,6-dione);Compound FS-12(methyl (2-acetyl-3,-dihydroxy-4-methoxyphenyl)acetate);compound FS-13(3-(3-hydroxypropyl)furan-2(5H)-one) and compound FS-15(2,3,4-trimethyl-3-[(1E)-3-oxobut-1-en-1-yl]cyclohexanone).Among them, the compounds FS-08, FS-10, FS-12and FS-13were determined as the novel ones. The compound FS-08and compound FS-12has a weak activity of CAS.From the cultivation crude extract of strain FEDT-145,14compounds were isolated. The structures of7compounds were elucidated:Compound FS-04(Uridine); Compound FS-05(11S,14S-cyclo-L-Trp-L-Phe);Compound FS-06(WIN64821);Compound FS-09(7-methoxy-2-benzofuran-1(3H)-one);Compound FS-11(6-methyl-2-oxopyridine-1(2H)-carboxamide); compound FS-14(5-hydroxymethyl-1H-1,3-diazepine-4,7-dione) and compound FS-16(3-benzyl-6-[hydroxy(2-hydroxyphenyl)methyl]piperazine-2,5-dione)Among them, the compounds FS-09, FS-11and FS-14were determined as the novel ones. The compound FS-14has a strong activity of CAS, the compound FS-11and the compound FS-16have a weak strong activity of CAS.This work lays the foundation for the further research of siderophore type secondary metabolites of fungi from the similar environment.