Ultrasensitive capillary electrophoresis of sulfated disaccharides in chondroitin/dermatan sulfates by laser-induced fluorescence after derivatization with 2-aminoacridone.
An ultrasensitive capillary electrophoretic method for separating the variously sulfated chondroitin/dermatan sulfate-derived delta-disaccharides after digestion with chondro/dermatolyases and derivatization with the fluorophore 2-aminoacridone is described. All known mono-, di- and tri-sulfated delta-disaccharides were completely separated using 15 mM orthophosphate buffer (pH 3.0) at 20 kV without any interference of the excess derivatizing reagent. They were detected at the anode (reversed polarity) using either an Ar-ion laser-induced fluorescence (LIF) detector (excitation wavelength 488 nm) or a UV detector. The sensitivity obtained by LIF (0.51 pmol/l) was at least 100 and 10 times higher as compared to those obtained by UV detection at 232 nm of underivatized delta-disaccharides and at 254 nm of those derivatized with aminoacridone, respectively. The method has been easily applied to the analysis of chondroitin/dermatan sulfates from various tissues at the attogram level, including chondrotin/dermatan sulfates from normal and aneurysmal human abdominal aortas.
Lamari
F
F
Department of Chemistry, University of Patras, Greece.
Theocharis
A
A
Hjerpe
A
A
Karamanos
N K
NK
D000609:Aminoacridines / Q000737:chemistry; D002809:Chondroitin Sulfates / Q000737:chemistry*; D003871:Dermatan Sulfate / Q000737:chemistry*; D004187:Disaccharides / Q000032:analysis*; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D007202:Indicators and Reagents / Q000737:chemistry; D007834:Lasers; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence
D000609:Aminoacridines; D004187:Disaccharides; D007202:Indicators and Reagents; D003871:Dermatan Sulfate; C072515:2-aminoacridone; D002809:Chondroitin Sulfates