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SIMULTANEOUS EXTRACTION AND SEPARATION OF CHLOROPHYLL AND PHYCOCYANIN WITH A BIPHASIC HYDROPHOBIC-HYDROPHILIC SYSTEM; TESTING DIFFERENT FILTERS AND SOLVENTS:

Abstract

Octanol-water extraction of chlorophyll and phycocyanine from cyanobacteria provides the possibility of a high-throughput methodology for monitoring water bodies for harmful algal blooms (HABs). However, problems encountered with this methodology include denaturation of the phycocyanin with hydrophobic solvents, debris from the filters, and incomplete lysis of the bacteria. In this report we use toluene, mineral oil, chloroform and isomers of octanol compared with 1-octanol in a biphasic extraction system of water and an organic solvent. A strategy for reducing background from filter debris is to collect the algae onto filters that allow the cells to be washed free of the filter, thus eliminating the homogenization of the filter along with the cells. We present data showing that Pall filters Super 800 and glass fiber filters GF/E disintegrate during homogenization, and Pall Versapore, Millipore, Nuclepore, and Durapore filters remained relatively intact. Nuclepore filters are very thin membranes that dissolve in toluene. Filters that remain intact can be removed after homogenization of the cells, facilitating removal of sample from the two layers of the biphasic extraction system and eliminating filter debris and the need to filter the samples prior to reading the O.D. or fluorescence. This work is supported by the East Georgia State College Baccalaureate Biology Program.

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