Title: Analysis of dideoxyadenosine triphosphate by capillary electrophoresis with fluorescence detection. I. Derivatization through the phosphate group
Authors: Mallampati, Swapna ×
Van Aerschot, Arthur
Hoogmartens, Jos
Van Schepdael, Ann #
Issue Date: Nov-2007
Series Title: Electrophoresis vol:28 issue:21 pages:3948-3956
Abstract: A CZE method was developed, which separates 2',3'-dideoxyadenosine-5'-triphosphate (ddATP) from other metabolites and endogenous nucleotides at high concentrations (20-200 mug/mL) to allow UV detection. To enhance sensitivity, fluorescence detection which requires prior derivatization of compounds was examined. Precapillary derivatization of ddATP in the presence of N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDAC) with dansyl ethylenediamine (dansyl EDA) was faster and stable compared to that of 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl ethylenediamine (BODIPY FL EDA). Reaction conditions, reagent concentrations and detection parameters were optimized and highest derivatization efficiency was achieved in 0.1 M 1-methylimidazole buffer (pH 8.0) with 140 mM EDAC in 1-methylimidazole buffer and 30 mM dansyl EDA in DMF for 90 min at 60 degrees C. Dansyl EDA derivatives of ddATP, 2'-deoxyadenosine-5'-triphosphate (dATP) and ATP were comigrating with the CZE method; therefore, a MEKC method was developed and optimized for repeatable separations. Upon dansylation, sensitivity of ddATP with fluorescence detection (LOQ = 12 ng/mL) was 160 times higher than UV detection (LOQ = 1.9 mug/mL).
ISSN: 0173-0835
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Pharmaceutical Analysis
Medicinal Chemistry (Rega Institute)
× corresponding author
# (joint) last author

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