Extraction of 15 microcystins and nodularin using immunoaffinity columns

Microcystins (MCYSTs) were isolated from surface water using reusable immunoaffinity columns. Individual MCYST were determined by high performance liquid chromatography equipped with a photo-diode array detector (HPLC-PDA, 200-300 nm). Subsequent analysis of the samples by liquid chromatography-electrospray ionization mass spectrometry (LC-ESMS) provided molecular weight information, which was used to tentatively identify individual MCYST variants for which standards were not available. Results obtained using immunoaffinity columns (IAC)-HPLC-PDA were compared to those obtained using solid phase extraction (SPE) Oasis(R) HLB-HPLC-PDA. This is the first report of the extraction of 15 microcystins and nodularin using immunoaffinity columns. Whereas previous reports demonstrates the use of IAC for four microcystins, we found that IAC selectively extracted the following microcystins: MCYST-RR, [D-Asp(3)]MCYST-RR, MCYST-YR, MCYST-LR, 3 MCYST-LR variants, MCYST-AR, MCYST-FR, MCYST-WR, MCYST-LA, MCYST-LA variant, the less polar microcystins such as MCYST-LF, MCYST-LW and nodularin. The IAC extracts were free of interferences which enabled better detection and identification of MCYSTs. Based on the amount loaded to the cartridges, the method detection limit was 10-14 ng when using IAC and 25 ng for SPE of each MCYST-RR, MCYST-YR and MCYST-LR. Reproducibilities expressed as relative standard deviation were 6-10% for SPE and 4-17% for IAC. Crown Copyright (C) 2003 Published by Elsevier Ltd. All rights reserved.