Modification of nanoscale polymeric adsorbent for the preparative separation and purification of tacrolimus from fermentation broth of Streptomyces tsukubaensis.

Tacrolimus is an important immunosuppressant produced by microbial fermentation. In this study, a modified nanoscale polymeric adsorbent, Ag+-exchanged resin, was prepared and studied for the preparative separation and purification of tacrolimus from fermentation broth of Streptomyces tsukubaensis. The performance and absorption characteristics of the modified nanoscale polymeric adsorbent namely Ag-NPS was evaluated. Notably, Ag-NPS resin displayed the pronounced separation capacities for tacrolimus and its equilibrium adsorption data was well-fitted to the Langmuir isotherm. Moreover, the dynamic adsorption and desorption tests was carried out to obtain the operational parameters for further purification of tacrolimus. Finally, tacrolimus and the two major impurities, ascomycin and dihydrotacrolimus, were separated well in the scale-up purification process. The purity and recovery of tacrolimus was recorded to be 99.12±0.25% and 90.41±2.05%. In conclusion, this method displayed a high potential for separation and purification of tacrolimus and other unsaturated bioactive compounds in high yield from the fermentation broth.

Improvement of tacrolimus production in Streptomyces tsukubaensis by mutagenesis and optimization of fermentation medium using Plackett-Burman design combined with response surface methodology.

OBJECTIVE: This study was conducted to enhance the production of tacrolimus in Streptomyces tsukubaensis by strain mutagenesis and optimization of the fermentation medium. RESULTS: A high tacrolimus producing strain S. tsukubaensis FIM-16-06 was obtained by ultraviolet mutagenesis coupled with atmospheric and room temperature plasma mutagenesis.Then, nine variables were screened using Plackett-Burman experimental design, in which soluble starch, peptone and Tween 80 showed significantly affected tacrolimus production. Further studies were carried out employing central composite design to elucidate the mutual interaction between the variables and to work out optimal fermentation medium composition for tacrolimus production. The optimum fermentation medium was found to contain 61.61 g/L of soluble starch, 20.61 g/L of peptone and 30.79 g/L of Tween 80. In the optimized medium, the production of tacrolimus reached 1293 mg/L in shake-flask culture, and reached 1522 mg/L while the scaled-up fermentation was conducted in a 1000 L fermenter, which was about 3.7 times higher than that in the original medium. CONCLUSIONS: Combining compound mutation with rational medium optimization is an effective approach for improving tacrolimus production, and the optimized fermentation medium could be efficiently used for industrial production.

Identification and antimicrobial properties of a new alkaloid produced by marine-derived Verrucosispora sp. FIM06-0036.

The actinomycete strain FIM06-0036 was isolated from marine sponge sample collected from the East China Sea and identified as Verrucosispora sp. based upon the results of 16S rRNA sequence analysis. One new alkaloid, 2-ethylhexyl 1H-imidazole-4-carboxylate (1), together with a known alkaloid butyl 1H-imidazole-4-carboxylate (2) was obtained from the fermentation products of this strain, the structures of compounds 1 and 2 were determined by their detailed analysis of 1 D, 2 D NMR and HR-ESI-MS data, along with literature data analysis. Compounds 1 and 2 were evaluated for their antimicrobial activity with MIC (minimum inhibitory concentration) values ranging from 8 to 256 µg · mL-1 against Helicobacter pylori, Klebsiella Pneumonia, Staphylococcus aureus and Enterococcus faecalis.

FW0622, a new siderophore isolated from marine Verrucosispora sp. by genomic mining.

Using the draft genome sequence of Verrucosispora sp. FIM060022, we have identified a new desferrioxamine-like siderophore, FW0622. This is the first chemically characterized siderophore obtained from Verrucosispora. The structure was elucidated by extensive spectral analyses. The biosynthetic pathway of FW0622 was proposed to occur via the non-ribosomal peptide synthetase (NRPS)-independent (NIS) synthetase pathway based on the putative biosynthetic siderophore gene cluster in FIM060022. The results demonstrate that marine-derived Verrucosispora species deserve recognition as an important source of new natural products. Furthermore, this study verified that genome mining is an effective way to identify compounds that may be overlooked by traditional methods.

Isolation, purification and identification of two new alkaloids metabolites from marine-derived Verrucosispora sp. FIM06025.

Chemical investigation of a marine-derived actinomycete strain Verrucosispora sp. FIM06025 isolated from a marine sponge sample collected from the East China Sea, resulted in the discovery of two new alkaloids, (2-(hydroxymethyl)-3-(2-(hydroxymethyl)-3-methylaziridin-1-yl) (2-hydroxyphenyl) methanone (1) and 2-(1-hydroxyethyl)-3,4-dihydrobenzo [f] [1,4]oxazepin-5(2H)-one (2). The structures of compounds 1 and 2 were determined by the detailed analysis of 1D, 2D NMR and HR-TOF-MS data, along with literature data analysis. The bioefficacy investigations revealed that compound 1 exhibited a broad spectrum of antimicrobial activity with MIC (minimum inhibitory concentration) values ranging from 3.4 to 200 µg·mL-1 against H. pylori, P. aeroginosa, A. baumanniiin, E. coli and K. pneumonia, S. aureus, C. albicans and E. faecium, however, compound 2, up to 200 µg/mL, displayed no antibacterial activity against these bacteria.

Structure elucidation and antitumour activity of a new macrolactam produced by marine-derived actinomycete Micromonospora sp. FIM05328.

Chemical investigation of a marine-derived actinomycete strain Micromonospora sp. FIM05328 isolated from a soil sample collected from the East China Sea, resulted in the discovery of a new 26-membered polyene macrolactam metabolite FW05328-1 (1), together with a known polyene with pyridone ring compound aurodox (2). The structures of compounds 1 and 2 were determined by the detailed analysis of 1D, 2D NMR and HR-TOF-MS data, along with literature data analysis. 1 and 2 exhibited excellent antiproliferative activities against KYSE30, KYSE180 and EC109 human tumour cell lines, but displayed no antibacterial activities against bacteria or fungi were tested.

Pericoterpenoid A, a new bioactive cadinane-type sesquiterpene from Periconia sp.

Pericoterpenoid A (1), a new cadinane-type sesquiterpene, was isolated from an endolichenic fungal strain Periconia sp. (No. 19-4-2-1). Its structure was characterized by analyzing the spectroscopic data (IR, MS, 1D- and 2D-NMR). The antimicrobial activity against Escherichia coli, Staphylococcus aureus, Aspergillus niger, and Candida albicans was evaluated. Pericoterpenoid A showed moderate antimicrobial activity against A. niger and weak activity against C. albicans. This is the first report of the presence of cadinane-type sesquiterpene in Periconia sp.

Nodulisporipyrones A-D, new bioactive α-pyrone derivatives from Nodulisporium sp.

Four new α-pyrone derivatives, nodulisporipyrones A-D (1-4), were isolated from the extract of an endolichenic fungal strain Nodulisporium sp. (65-12-7-1) that was fermented with rice. The structures of 1-4 were elucidated by extensive spectroscopic analysis, and the absolute configurations were determined by modified Mosher's method and electronic circular dichroism experiments. Their antimicrobial activities against Staphylococcus aureus 209P, Escherichia coli ATCC0111, Aspergillus niger R330, and Candida albicans FIM709 were evaluated using a paper disk diffusion method. Nodulisporipyrones A-D (1-4) are the first α-pyrone derivatives from Nodulisporium fungi.

Pericocins A-D, New Bioactive Compounds from Periconia sp.

One new dihydroisocoumarin, pericocin A (1), one new chromone, pericocin B (2), and two new α-pyrone derivatives, pericocins C-D (3-4), together with two known compounds, 3-(2-oxo-2H-pyran-6-yl)propanoic acid (5) and (E)-3-(2-oxo-2H-pyran-6-yl)acrylic acid (6), were isolated from the culture of the endolichenic fungus Periconia sp.. Their structures were elucidated by spectroscopic methods. All these compounds are derived from the polyketone biosynthetic pathway. Compound 1 was obtained as a mixture of enantiomers. The antimicrobial activity of compounds 1-5 was tested against Escherichia coli, Staphylococcus aureus, Aspergillus niger, and Candida albicans. Compounds 1-5 showed moderate antimicrobial activity against A. niger and weak activity against C. albicans.

New phenethylamine derivatives from Arenibacter nanhaiticus sp. nov. NH36A(T) and their antimicrobial activity.

Five new phenethylamine (PEA) derivatives (1-5) were isolated from the strain of Arenibacter nanhaiticus sp. nov. NH36A(T) derived from the marine sediment of the South China Sea by bioassay-guided fractionation. Their structures were elucidated by spectroscopic methods including UV, IR, HR-MS and NMR. Interestingly, compounds 1-4 existed as enantiomers, which were resolved by chiral liquid chromatography. The resolved configuration of each enantiomer was assigned by the Marfey's method. Of these compounds, 5 showed weak antimicrobial activity against Staphylococcus aureus and Bacillus subtilis with MIC values of 0.50 and 0.25 mg ml(-1), respectively.

[Actinobacterial diversity of marine sediment samples from Chile].

OBJECTIVE: The aim of this study was to investigate actinobacterial diversity in Chilean marine sediments. METHODS: Actinobacterial diversity in these sediments was investigated by selective isolation method, culture-independent method and phylogenetic analysis based on 16S rRNA gene sequences. Six selective media were used to isolate actinomycetes from sediment samples. The primers for the class Actinobacteria were used for Actinobacterial 16S rRNA gene amplification and then a clone library was constructed for the sediment sample btt. Twenty-two strains with different culture characteristics and 59 clones from sample btt were selected for 16S rRNA gene sequences analysis. To determine requirement for seawater each strain was grown on oatmeal agar prepared with deionized water and with seawater, respectively. Strains were screened for antibiotic activity against bacteria and fungi. RESULTS: In total 328 actinomycetes were obtained. Twenty-two strains which were selected belonged to Streptomyces, Micromonospora, Polymorphospora, Aeromicrobium and Brachybacterium. Fifty-nine clones (40 OTUs) were sequenced, and 60% OTUs belonged to Actinobacteridae, Acidimicrobidae and Rubrobacteridae. The other 40% OTUs, which formed several distinct clades in phylogenetic tree among phylum Actinobacteria may represent new taxonomical groups. 50% of the 47 sea water dependant strains and nineteen strains out of the above 22 strains exhibited antimicrobial activity. CONCLUSION: There was abundant actinobacterial diversity in the marine sediments of Chile, and the result implied that there were large numbers of unknown actinobacterial groups in the sediments. Actinomycetes from Chilean marine sediments had the potential of producing bioactive secondary metabolites.

Synthesis and biological evaluation of 4-fluoroproline and 4-fluoropyrrolidine-2-acetic acid derivatives as new GABA uptake inhibitors.

Preparation for the N-alkylated derivatives of enantiomerically pure (2S)-4-fluoroproline and (2S)-4-fluoropyrrolidine-2-acetic acid is described. The final compounds were evaluated as potential GAT-1 uptake inhibitors via cultured cell lines expressing mouse GAT-1. Compared with their corresponding 4-hydroxy compounds, these derivatives exhibited slight improvement on their inhibitory potency, but still much weaker than their corresponding compounds with no substituents at the C-4 of the pyrrolidine moiety, with the most potent affinity being about 1/15 fold as that of Tiagabine. The drastic decrease of their affinity may arise from sharp reduction of their basicity due to strong inductive effect of the 4-fluorine. However the configuration of the C-4 linking fluorine did not have much influence on their affinity for GAT-1.