The development of methods using capillary electrophoresis (CE) to measure and characterize intracellular nucleotide metabolites and endogenous nucleotide pools continues. Our previous work has shown that a more than 100-fold signal enhancement can be obtained for the CE analysis of nucleotides by using quantitative on-line sample-stacking. These methods have been applied to characterize and study mixtures of synthetic nucleotide derivatives. Biologically derived mixtures of nucleotides, however, show a marked deterioration in signal and resolution upon sample-stacking and CE analysis. Sample preparation methods and analysis strategies to overcome this effect have been explored and have provided some success, resulting in a 20- to 30-fold signal enhancement for biological samples. This investigation continues. We are also evaluating whole-column sample stacking for the micropreparative CE isolation of trace nucleotide metabolites for structural analysis by MALDI mass spectrometry. This will have application for the determination and measurement of the active intracellular nucleotide metabolites of the nucleoside-based reverse transciptase inhibitors that used in AIDS therapy. The National Cancer Institute (NCI) Chemical Database, which contains over half a million unique structures acquired over the past 50 years, is a valuable resource for biological testing, for drug development and for chemical information applications. Many investigators use this database to identify new lead compounds for anticancer and antiviral drug development. The validity of this approach depends on the selected compounds possessing the stated structures. Our recently completed targeted analytical evaluation of the structural integrity and purity of compounds in the open NCI Chemical Database revealed that a substantial fraction (30%) of them may be unacceptable for the above purposes. These compounds have no spectral evidence for the presence of the desired structure, or contaminants exceed 50% of the sample. Because of this, we continue to analyze and evaluate the structures and purities of any compounds obtained from NCI Database on a case-by-case basis.
