Pre-Column (HPLC) Fluorescence Labelling of Glucuronides

Lingeman, H.; Meussen, G. W. M.; Zouwen, C. van der; Underberg, W.J.M.; Hulshoff, A.

doi:10.1007/978-1-4757-1892-8_24

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“…18 This derivatization reaction was performed in order to allow a sensitive detection of the carboxylic acid. Figure 1 shows the typical chromatograms of the reaction mixtures obtained after derivatization of benzoic acid with SN 23-518 (Fig, la) and SN 23-082 (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Liquid Chromatography of Basic Solutes on Silica Gel and Aluminum Oxide

1988

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: To whom correspondence should be directed.Chromatography of basic solutes has been described on non-modified silica and alumina using aqueous buffers with suitable modifiers. On these packing materials selectivity and retention of the compounds are influenced in various ways; by the modifier content, the electrolyte concentration and the pH. Generally, only cationic species are retained on these sorbents. Their retention decreases with increasing electrolyte concentration. The relationship between retention and modifier content is more complicated and, moreover, different for the two packing materials. The differences in selectivity will be illustrated by comparing the chromatography of combinations of basic drugs (e.g. phenylethylamines, alkaloids), the analysis of basic drugs in pharmaceutical formulations ( e.g. prednisolone, physostigmine) and the detection of basic drug metabolites (e.g. amphoteric glucuronides) on silica and alumina. The aqueous nature of the eluent allows the combination of these packing materials and the use of column switching systems.Keywords liquid chromatography, non-modified silica, basic solutes, pharmaceutical formulationsThe liquid chromatographic (LC) analysis of basic solutes with the usual reversed-phase (RP) systems on modified silica frequently suffers from badly tailing peaks. ' To prevent these problems, the column packing materials2, the use of ion-pair chromatography3, the addition of tailing-suppressing agents4 or the application of normal-phase chromatography5 have been advocated. However, these systems are sometimes still inadequate.An interesting development for the separation of amines is the application of non-modified polar stationary phases, such as aluminum oxide (alumina) and silica gel (silica), in combination with aqueous solvent mixtures."6-10 The retention behavior of these amines, primarily based on ion-exchange (IE) mechanisms, appears to be controlled predominantly by the pH and the nature and concentration of the organic modifier.The nature and concentration of the competing ions and the buffer components present in the mobile phase also exert some additional influence on the retention.Alumina, amphoteric in nature, exhibits anion-exchange properties at pH values below the zero point of charge (ZPC) and shows cationexchange behavior above this value10, requiring the pH of the mobile phase to be higher than the ZPC for the analysis of protonated amines and quaternary ammonium ions. In contrast, silica shows only cationexchange properties and its IE capacity strongly increases with increasing pH."Characteristic for both systems is that neutral molecules and anions are hardly retained. Increasing the hydrophobic chain lengths of the competing ions in both IE systems results in a larger capacity ratio (k') of the solute molecules12, whereas an increase in the concentration of the counter ion hardly influences k' in the two IE systems. This leads to the conclusion that the influence of added salt is to be solely attributed to the competing effect of the ...

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Section: Resultsmentioning

confidence: 99%

“…For the analysis of this derivative a two column IE/ RP system was employed. 18 The non-modified silica column in the IE mode holds up the excess of the amine (SN 23-518). The derivative, however, is eluted unretained on this column.…”

Section: Resultsmentioning

confidence: 99%