Comparison with CZE

The separation of endogenous 17- or 18-hydroxylated corticosteroids of the 21-hydroxylated 4-pregnen series was obtained by capillary electrophoresis of their charged borate chelate complexes (323). Aldosterone, 18-hydroxycorti-costerone, 18-hydroxy-deoxycorticosterone, cortisone, cortisol and 11-deoxycor-tisol are separated and resolved with 400 mM borate buffer at pH 9.0. The corticosteroid/borate chelation complex as indicated by CE data correlated well with 11 B-NMR. The separation of corticosteroids and benzothiazin analogs were studied by MEKC and a comparison with CZE was made (324). Bile salts, which have a similar carbon skeleton to the corticosteroids, were used for the separation of these steroids. A short analysis time, 15 min, and a high number of theoretical plates (150,000-350,000) were obtained. Sodium cholate was found to be very effective. The MEKC method was applied to the determination of the drug substance in tablets and cream formulations. An internal standard method was used for quantitation. The purity of the drug substance was also determined. 

Plasma protein binding is also an important parameter in the pharmacokinetic field. Frontal analysis combined with capillary zone electrophoresis (CZE-FA) (67-69) is a powerful technique for high-throughput assay, because it is relatively rapid and easy to automate, in comparison with conventional methods such as dialysis, ultrafiltration, and ultracentrifugation. Recently, we introduced the EKC approach with ionic CDs to frontal analysis for anionic drugs that cannot be analyzed by conventional CZE-FA (70). In this approach, ionic CDs work as an EKC pseudostationary not for proteins but for small solutes. 

Figure 8.5 CZE profiles (pH 9.2) of the retained (A) and unretained (B) fractions obtained after passing the crude extract through Bio-Rad AG 50W-X8 filter, and of proteins retained on the cation-exchange cartridge from the alcohol-treated rat hair preparation (C). High-sulfur proteins were recovered in the filtrate, while low-sulfur proteins were released by 0.1 M triethylamine. Comparison with the two-dimensional gel separations is visualized under each CZE run. The position of the fraction of low-sulfur proteins occurring in alcohol-treated animals but absent in controls is indicated by an arrow. (From Jelfnkov et al., 1995, with permission.)...

Comparison of CZE with HPLC for the det. of additives in food stuffs Caffeine, aspartame benzoic acid in soft drinks sweetening powders Cola degassed, diluted powders dissolved, filtered and diluted... 

Preservation of extracts with CHCI3, confirmation with CZE Preservation of extracts with CHCI3, confirmation with lEC Comparison of different samphng techniques Filters stored frozen... 

Various modes of CE have been used for impurity determination with CE. Hansen described the comparison of CZE (FSCE), MEKC, MEEKC, and NACE for the determination of impurities in bromazepam [193] and found that NACE provided the best technique for the poorly water-soluble compounds with impurities determined to be 0.05%. Chiral CE methods can be used to determine enantiomeric impurities [191]. Readers are referred to a paper by Sokoliess and Roller [192] describing method development for chiral purity testing in CE. FSCE is the most widely used as most drugs and impurities are acidic or basic. Low pH buffers are used for basic drug impurities and high pH buffers for acidic compounds. 

Another study using CZE for the analysis of CDT as a marker of alcohol abuse was carried out by Giordano et al. [202]. They did not detect asialo-Tf in sera from alcoholic people, but they detected a significant increase in the disialo-Tf (related to total Tf) in those samples in comparison with control (nonalcoholic subjects) samples. In the same direction, and by establishing the Tf index as % area of disialo-Tf in relation to tetrasialo-Tf, Lanz et al. [185] found that a good classification of sera could be performed on the basis of this index increasing in sera of alcoholic individuals. 

FIGURE 36.6 Comparison of CZE analysis of partially denatured human sera using run buffer containing 0.04% NanoOrange dye. Sera were diluted 1 500 in 0.5% SDS, with 1% dye included in the sample buffer. Separation conditions 100 mM borate, 3 mM diaminobutane, pH 8.5. (Reprinted from Giordano, B. C., et al,... 

In ID, the natural isotopic abundance ratio of Cd is altered in the sample by spiking it with an exact and known amount of Cd-emiched isotope (the so-called spike , with a different isotopic abvmdance ratio than natural cadmium). The reference isotope is usually the isotope of highest natural abundance ( " Cd), while the spike isotope is one of the lesser abundant natural isotopes (normally Cd, Cd, or Cd). As a result of the spiking process, the measurement by ICP-MS of the new isotope ratio (e.g., " Cd/ Cd) and its comparison with the natural isotope ratio offers the original Cd concentration in the sample. If the isotope dilution is performed online in an LC- or CZE-ICP-MS experiment, quantification of Cd in each of the isolated species can be accurately achieved by integration of each chromatographic/electrophore-tic peak after transformation of the data into mass flow by means of the ID equation. 

FIGURE I Comparison of the separation of a protein digest by (a) HPLC and (b) CZE, reprinted with permission from reference 25. 

Fig. 10.22. Comparison of separation of drug mix by CEC, HPLC and CZE. Column, Spherisorb ODS-1, 3 pm, 250 (335) mm x 100 pm i.d. mobile phase, acetonitrile-25 mM phosphate, 0.2% hexylamine, pH 2.5 (80 20 v/v) voltage (CEC), 25 kV pressure (HPLC), 200 bar CZE, uncoated capillary 250 (335) mm x 75 pm i.d. Peak identification 1, procaine 2, timolol 3, ambroxolol 4, metoclopramide 5, thiourea 6, naproxene 7, antipyrine. Reproduced with permission from Dittman et al. [56],...

RhEPO is another complex sialoglycoprotein, with three N-linked and one 0-linked glycosylation sites, that has been extensively characterized by CZE and cIEF as described in previous sections. The comparison of rhEPO and uEPO has been reported on uncoated capillaries [40] and on capillaries dynamically coated with ionene [62]. Although increases in resolution and sensitivity are desirable, and the samples of uEPO corresponded to purified standards, these works showed the potential of discriminating between exogenous and endogenous EPO. 

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