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Velocity-difference induced focusing (V-DIF) of nucleotides was achieved by using a dynamic pH junction in capillary electrophoresis (CE) with UV detection. The influence of specific analyte properties, such as nucleotide base structure, sugar structure, and degree of phosphorylation, is examined. The pKa values and borate complexation with vicinal diols are important factors that caused the focusing. Therefore, the pH and borate content in the sample and background electrolyte can be adjusted to optimize the focusing effect. This method allows the injection of large volumes of sample (approximately 300 nL), resulting in at least 50-fold improvement in concentration sensitivity. The detection limit of 4.0 x 10(-8) M for nucleotides can be achieved in favorable conditions. V-DIF can be also applied to nucleotide pool analysis from cell extracts to improve the concentration sensitivity of CE and to reduce the time-consuming steps of desalting and off-line preconcentration that are often required for assays of nucleotides from biological samples.

Understanding the separation process in capillary electrophoresis (CE) leads to the unification of the theories for separation science. While the separation of analytes is governed by equilibria in chromatography, and by (centrifugal) field in ultracentrifugation, the separation in CE is governed by both equilibria and (electric) field. Therefore, a comprehensive separation theory that describes the separation process of analytes in CE should be able to describe the separation processes in both chromatography and ultracentrifugation. In this paper, we propose that individual capacity factors for each analyte species be used to describe the migration behavior of an analyte. The effect of field on each analyte species, as well as the effect of equilibria are considered in deriving a generalized equation that is applicable for all separation techniques. The separation factor defined at present does not directly relate to the migration rates of the analytes, and therefore can not be used in a generalized theory. We propose that the ratio of the migration rates of a pair of analytes (gamma) should be used as the separation factor, instead of the ratio of the two capacity factors. When gamma is used to describe the separation of two closely migrating analytes, all separation techniques have the same resolution equation.

A comprehensive theory is proposed to describe the migration behavior of analytes in capillary electrophoresis (CE) when one or more additives are present in the buffer solution. This theory amalgamates and extends the previous work done by others. The capacity factor (k') in this theory is defined as the product of the equilibrium constant and the additive concentration, thus, k' changes linearly with additive concentration. The net electrophoretic mobility of an analyte is a function of k', therefore, it can be changed by varying the additive concentration. Three parameters are needed to predict the mobility of an analyte in a one-additive CE system: the mobility of the free analyte, the mobility of the complex, and the equilibrium constant for the analyte-additive interaction (which determines the fraction of the free analyte at different additive concentrations). When additives are used, the change in viscosity obscures this relationship, therefore, a viscosity correction factor is required to convert all mobilities to an ideal state where the viscosity remains constant. The migration behavior of an analyte in a solution with multiple additives can be predicted and controlled, once the equilibrium constants of the interactions between the analyte and each of the additives are obtained separately. beta-Cyclodextrin and hydroxypropyl-beta-cyclodextrin are used as additives and the migration behavior of phenol, p-nitrophenol, and benzoic acid are studied as a model system to verify this theory. When the necessary viscosity correction factor is included, the net electrophoretic mobilities of the analytes obtained from experimental results agree with the values predicted by the theory based on dynamic complexation. Although only experiments with one and two additives were carried out to verify the theory, the equations apply to situations when more than two additives are used. The relationship between the theories of electrophoresis and chromatography is clarified.

T2 weighed spin echo magnetic resonance images (MRI) of the intervertebral disks of 4 lumbar spines were graded and the nuclei pulposi were analyzed for water, collagen and proteoglycan. The brightness of the nuclear image correlated directly with the proteoglycan concentration, but not with the water or collagen. The dark midnuclear cleft had a collagen concentration slightly higher and a water concentration slightly lower than the adjacent zones; no corresponding differences in proteoglycan were seen, although the relationship with MRI grade was confirmed.

An assay for keratan sulfate in papain digests of human intervertebral disc and other tissues has been developed. The digest is applied to the acetate form of a tertiary amine acrylic anion-exchange resin, the oligosaccharide hexose is removed by washing the resin with 0.2 M sodium acetate buffer pH 5.0, then the keratan sulfate is eluted quantitatively with 1.0 M pyridinium sulfate pH 2.5 and assayed for hexose by the anthrone reaction. The keratan sulfate content of human intervertebral disc tissues ranged from 7 to 78 mumole galactose equivalents/g fresh weight; the root mean square error was 2 mumole/g; 10-25 mg of tissue were required. The separation of oligosaccharides from keratan sulfate was confirmed by gel permeation chromatography, sugar composition, ester sulfate analysis, and nuclear magnetic resonance.

The presence of acetate and pyruvate groups in Klebsiella capsular polysaccharides may be demonstrated and estimated quantitatively by running the proton magnetic resonance spectrum of the polysaccharide (as sodium salt) in deuterium oxide at 95 C. Such spectra also permit an assessment to be made of the number of alpha- and beta-linkages in the repeat unit of the polysaccharide structure.