Determination of d-Mannose in Plasma by HPLC

Abstract

d-Mannose is an essential monosaccharide in the structure of glycoproteins, cell-surface glycoconjugates, and glycosylated phosphatidylinositide anchors. Several stud-ies have focused on plasma or serum mannose concentrations in patients with invasive candidiasis (1)(2), diabetes mellitus (3)(4)(5)(6)(7)(8), or congenital disorders of glycosylation type 1 (9). There are various methods to determine mannose concentrations in human plasma or serum: enzymatic methods (4)(5)(7)(10), gas-liquid chromatography (2)(11), high-resolution liquid chromatography (12), gas-liquid chromatography–mass spectrometry (6), and capillary electrophoresis (13). None of these methods is fully suitable for routine use for various reasons, e.g., the incomplete or time-consuming elimination of the ∼100-fold excess of blood glucose, the use of instruments with limited availability, and the need for a large sample volume. We investigated whether a HPLC assay using an anion-exchange column would be appropriate for plasma mannose determinations. With the procedure described below, mannose could be rapidly and accurately determined in small amounts of plasma. Boric acid, guanidine hydrochloride, sodium metaperiodate, and acetonitrile were purchased from Wako Pure Chemicals Co. Ltd. d-Mannose was obtained from Sigma Chemical Co. We obtained blood samples from healthy individuals and from nondiabetic and diabetic patients after receiving their informed consent for this study. After collection of venous blood in tubes containing disodium EDTA (an anticoagulant) and NaF (a glycolysis inhibitor), plasma was separated by centrifugation. The plasma was mixed with an equal volume of 0.6 mol/L perchloric acid, a protein-precipitating reagent. The mixture was centrifuged at 8000 g for 10 min at 4 °C, and the protein-free supernatant was used for the analysis of mannose. We used a HPLC system (Gulliver system with a JASCO PU-980 pump) equipped …