Development of a FIA system with immobilized enzymes for specific post-column detection of purine bases and their nucleosides separated by HPLC column.
A sensitive and highly selective method for the simultaneous determination of purine bases and their nucleosides is proposed. An amperometric flow-injection system with the two immobilized enzyme reactors (guanase immobilized reactor and purine nucleoside phosphorylase/xanthine oxidase co-immobilized reactor) is used as the specific post-column detection system of HPLC, to convert compounds separated by a reversed-phase. HPLC column to electroactive species (hydrogen peroxide and uric acid) which can be detected at a flow-through platinum electrode. The proposed detection system is specific for a group of purine bases and purine nucleosides and does not respond for purine nucleotides and pyrimidine bases. The linear determination ranges are from 10 pmol to 5 nmol for four purine bases (hypoxanthine, xanthine, guanine, and adenine) and four purine nucleosides (inosine, xanthosine, guanosine, and adenosine). The detection limits are 1.2-5.5 pmol.
Department of Applied Chemistry, College of Engineering, University of Osaka Prefecture, Sakai, Japan.
T
T
Yao
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Y
Matsumoto
T
T
Wasa
D002851:Chromatography, High Pressure Liquid; D004800:Enzymes, Immobilized; D006148:Guanine Deaminase; D006863:Hydrogen-Ion Concentration; D011684:Purine Nucleosides / Q000032:analysis*; D011683:Purine-Nucleoside Phosphorylase; D011687:Purines / Q000032:analysis*; D014969:Xanthine Oxidase
D004800:Enzymes, Immobilized; D011684:Purine Nucleosides; D011687:Purines; D014969:Xanthine Oxidase; D011683:Purine-Nucleoside Phosphorylase; D006148:Guanine Deaminase