Quantitative derivatization and high-performance liquid chromatographic analysis of cyanobacterial heterocyst-type...

Procedures are described for the rapid and quantitative analysis of cyanobacterial heterocyst-type glycolipids (HGs) by normal-phase HPLC of their per-O-benzoylated derivatives. Total lipids are obtained from 1 ml of nitrogen-fixing cyanobacterial culture by triplicate extraction with chloroform/methanol, 1/1 (v/v), and the HGs are isolated from other complex lipids by preparative silica gel TLC. A C18 solid-phase extraction cartridge is used to ensure quantitative salt-free recovery of the HGs, and the purified glycolipids are then rendered uv-absorbing by a per-O-benzoylation derivatization reaction for which optimal conditions have been established. Derivatives are analyzed within 12 min on a 3-microns silica HPLC column using a linear gradient of 2-propanol in n-hexane and uv monitoring at 230 nm. The reaction product was also used to determine the relative proportions of the glucosyl and galactosyl epimers of individual members of this class of glycolipid.