Fabrication Process Research Of MC-LR

Blue-green algal bloom aggravated in recent years causing by the pollution of freshwater lakes, in which the main harmful substances are microcystins (microcystins, MCs). As the main isomers of MCs, MC-LR is the one of the standard samples in the detecting of drinking water quality. Many literatures reported the research of the extraction, separation and preparation of MC-LR including solid phase extraction, high performance liquid chromatography and thin-layer chromatography et al., but fewer concerning the large scale preparation of MCs especially in land. Most of isomers standard samples were exported from abroad.MCs isomers were extracted by using algae from Dianchi, Yunnan province as raw materials. The extracting conditions were discussed. MC-LR was then prepared from the extracts and characterized by HPLC-MS.Different solvents, extraction methods and extraction time of MCs isomers extraction process were studied and the best extraction procedure was obtained as follows: The algae was stirred with magnet iced stirring for 1.5h and then extracted with ultrasonic for 20min in the solvent of methanol/water (65% v/v) at room temperature. The crude extracts were pre-purified by filtering with the 0.45μm of microfiltration membrane and eluted with solid phase extraction by using 70% methanol/water as eluent.MCs isomers were separated and purified by using high performance liquid chromatography. The three main isomers of MCs could be separated successfully with mobile phase of acetonitrile/water (80% v/v), detection wavelength of 238nm, flow rate of 0.5mL/min and injection volume of 2.0μL.MCs isomers were prepared by using preparative high performance liquid chromatography. The ten main isomers were gained with mobile phase of acetonitrile/water (50% v/v), detection wavelength of 238nm, flow rate of 2mL/min and injection volume of 100μL. MC-LR was characterized by HPLC-MS, in which the molecular ion peaks and characteristics peaks matched with the MC-LR structure. Finally, MC-LR was prepared with algae aeruginosa cultured in our laboratory. The results were the same as the preparation MC-LR from nature algae.The preparation methods of MCs were simple and easy to operate in our research. The studies have a good guidance to the preparation of other MCs isomers.