High performance column liquid chromatographic analysis of selected carboxylic acids in Biomphalaria glabrata patently infected with Schistosoma mansoni
Daniel R. Massa, Michael J. Chejlava, Bernard Fried, and Joseph Sherma. Lafayette College, Easton, PA
A high performance ion exclusion column liquid chromatography (HPLC) method was developed for determination of the effects of a patent Schistosoma mansoni infection on four biologically important carboxylic acids in the digestive gland gonad complex (DDG) and hemolymph (blood) of Biomphalaria glabrata snails. Acids were extracted from DDGs with 50% Locke's solution, and extracts were cleaned up by ion exchange solid phase extraction (SPE) on a Bond Elute-SAX column with elution by 0.50 M sulfuric acid. Determination was carried out by use of HPLC with a Biorad Aminex HPX-87H column, 5.0 mM sulfuric acid mobile phase, and ultraviolet (UV) detection. Hemolymph was applied directly to the SPE column for extraction and cleanup of the acids. Acetic, fumaric, malic, and pyruvic acids were identified by comparison of retention times with standards, confirmed by automatic spectral analysis with the UV diode array detector during the separation, and quantified at concentrations ranging from 7.9 to 280 ppm in the DGG and 124 to 9000 ppm in hemolymph. Infection with S. mansoni caused a significant reduction (Student's t-test, P < 0.05) in the concentrations of the four acids in the DGG but not the hemolymph of B. glabrata compared to uninfected cohort snails. The significant reduction of the acids in patently infected snails suggests that the acids were utilized by the sporocysts and cercariae in the snail tissue.