title: Two-step separation of nostotrebin 6 from cultivated soil cyanobacterium (Nostoc sp.) by high performance countercurrent chromatography.
abstract: High performance countercurrent chromatography (HPCCC) was successfully applied for the separation of nostotrebin 6 from cultivated soil cyanobacteria in a two-step operation. A two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (4:5:4:5, v/v/v/v) was employed for the HPCCC separation. In the first-step operation, its neutral upper phase was used as stationary phase and its basic lower phase (1% NH3 in lower phase) was employed as mobile phase at a flow rate of 1 mL/min. In the second operation step, its neutral upper phase was used as stationary phase, whereas both its neutral lower phase and basic lower phase were employed as mobile phase with a linear gradient elution at a flow rate of 0.8 mL/min. The revolution speed and temperature of the separation column were 1,000 rpm and 30 °C, respectively. Using HPCCC followed by clean-up on Sephadex LH-20 gel, 4 mg of nostotrebin 6 with a purity of 99% as determined by HPLC/DAD-ESI-HRMS was obtained from 100 mg of crude extract. The chemical identity of the isolated compound was confirmed by comparing its spectroscopic data (UV, ESI-HRMS, ESI-HRMS2) with those of an authentic standard and data available in the literature.
authors: Department of Phototrophic Microorganisms-ALGATECH, Institute of Microbiology, Academy of Sciences of the Czech Republic, Opatovický mlýn, Třebon 379 81, Czech Republic. jcheel@email.cz.
authors: José
authors: J
authors: Cheel
authors: Department of Phototrophic Microorganisms-ALGATECH, Institute of Microbiology, Academy of Sciences of the Czech Republic, Opatovický mlýn, Třebon 379 81, Czech Republic. kucerova@alga.cz.
authors: Petra
authors: P
authors: Kučerová
authors: Brunel Institute for Bioengineering, Brunel University, Kingston Lane, Uxbridge, Middlesex UB8 3PH, UK. Ian.Garrard@brunel.ac.uk.
authors: Ian
authors: I
authors: Garrard
authors: Brunel Institute for Bioengineering, Brunel University, Kingston Lane, Uxbridge, Middlesex UB8 3PH, UK. svetlana.ignatova@brunel.ac.uk.
authors: Svetlana
authors: S
authors: Ignatova
authors: Department of Phototrophic Microorganisms-ALGATECH, Institute of Microbiology, Academy of Sciences of the Czech Republic, Opatovický mlýn, Třebon 379 81, Czech Republic. hrouzekp@gmail.com.
authors: Pavel
authors: P
authors: Hrouzek
authors: Department of Phototrophic Microorganisms-ALGATECH, Institute of Microbiology, Academy of Sciences of the Czech Republic, Opatovický mlýn, Třebon 379 81, Czech Republic. kopecky@alga.cz.
authors: Jiří
authors: J
authors: Kopecký
mesh_terms: D000085:Acetates / Q000737:chemistry; D002800:Cholinesterase Inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D003377:Countercurrent Distribution; D003517:Cyclopentanes / Q000737:chemistry / Q000302:isolation & purification*; D006586:Hexanes / Q000737:chemistry; D000432:Methanol / Q000737:chemistry; D046937:Nostoc / Q000737:chemistry*; D012988:Soil Microbiology; D012997:Solvents / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D014867:Water / Q000737:chemistry
chemical_list: D000085:Acetates; D002800:Cholinesterase Inhibitors; D003517:Cyclopentanes; D006586:Hexanes; D012997:Solvents; C551503:nostotrebin 6; D014867:Water; C026385:n-hexane; C007650:ethyl acetate; D000432:Methanol