Ph. Morin

Borate complexation of flavonoid-O-glycosides in capillary electrophoresis: I. Separation of flavonoid-7-O-glycosides differing in their flavonoid aglycone

Capillary electrophoresis was found to give significantly greater efficiency, selectivity and speed compared with high-performance liquid chromatography for the separation of mixtures of flavonoid-O-glycosides. The migration behaviour of selected flavonoid-O-glycosides and their flavonoid aglycones in free solution capillary electrophoresis and micellar electrokinetic capillary chromatography (MECC) was investigated. A mixture of flavonoid-7-O-glycosides and their flavonoid aglycones was resolved by MECC. A 70 cm × 50 μm I.D. fused-silica capillary column and a 50 mM sodium dodecyl sulphate-20 mM Tris-CHl running buffer (pH 7.1) were used for these electrophoretic separations. Under these neutral conditions, the retention mechanism is based on hydrophobic interactions betweeen flavonoids and the hydrophobic core of the micelles. Free zone capillary electrophoresis of borate complexes was more appropriate for the separation of a mixture of flavonoid-O-glycosides. In that case, the migration behaviour of these selected flavonoid-O-glycosides was explained by ionization of some hydroxyl groups and mainly by borate complexation of the sugar moiety. The identity of each flavonoid-O-glycoside was determined by using a fast-scanning multiple-wavelength UV detector and by recording its on-column UV spectrum in the range of 190–360 nm.

Borate complexation of flavonoid-O-glycosides in capillary electrophoresis. II. Separation of flavonoid-3-O-glycosides differing in their sugar moiety.

Capillary electrophoresis was found to give significantly higher efficiency, selectivity and speed than high-performance liquid chromatography for the separation of a mixture of flavonoid-3-O-glycosides differing in their sugar moiety. Boric acid running buffer (0.2 M, pH 10.5) was used for this electrophoretic separation. The migration order in free solution capillary electrophoresis (CE) and the selectivity of these flavonoid-3-O-glycosides can be mainly explained by in situ borate complexation of both the sugar moiety and the cis-1,2-hydroxyl groups on the flavonoid skeleton and, to a lesser extent, by the ionization of hydroxyl groups on the flavonoid skeleton due to alkaline pH conditions. The correlation of the electrophoretic mobilities with the configuration and conformation of the compounds is discussed.

Identification of phosphonic acids by capillary electrophoresis–ionspray mass spectrometry

The identification of alkylphosphonic acids in spiked tap water has been investigated by on-line capillary electrophoresis-UV spectrometry-mass spectrometry (CE-UV-MS) in negative-ion mode. The 5 mM sorbic acid-ammonia electrolyte (pH 6.5) allows simultaneous indirect UV and MS (ionspray) detection. Several parameters (electrolyte pH, make-up chemical composition and make-up flow-rate) have been optimized and 5 mg/l limit of detection has been reached for these analytes in selected ion monitoring MS detection. MS-MS detection has also been investigated to reach a low detection limit (100 micrograms/l) for alkyl alkylphosphonic acids in spiked tap water. The mass spectra of these compounds exhibit a very abundant negative ion, [M-H]- (MS) with characteristic fragmentation (MS-MS) of acids and monoesters. Quantitative analysis achieved with CE-UV shows good correlation coefficients and allows accurate quantification of the analytes.

Separation of transition metal cations by capillary electrophoresis optimization of complexing agent concentrations (lactic acid and 18-crown-6)

Abstract The simultaneous separation of alkali, alkaline earth and transition metal cations by capillary electrophoresis was achieved using an electrolyte containing imidazole as indirect UV co-ion and lactic acid and 18-crown-6 as complexing agents with indirect UV detection. The addition of lactic acid to the imidazole electrolyte allows the separation of transition metal cations, whereas 18-crown-6 allows one to resolve ammonium and potassium and also sodium and lead. An electrolyte containing 10 mM imidazole, 5 mM lactic acid and 0.5 mM 18-crown-6 at pH 4.5 and with the separation temperature fixed at 30°C allow the simultaneous separation of alkali, alkaline earth and transition metal cations by capillary electrophoresis. By increasing the hydrodynamic injection time to 15 s, a linear relationship between corrected peak area and cation concentration in the 0.1–1 ppm range was obtained for transition metal cations. Further, the separation of inorganic cations at a low concentration level (100 ng ml−1) was achieved without any loss of resolution.

Synthesis and study of a molecularly imprinted polymer for the specific extraction of indole alkaloids from Catharanthus roseus extracts.

Two molecularly imprinted polymers (MIP) for catharanthine and vindoline have been synthesized in order to specifically extract these natural indole alkaloids from Catharanthus roseus by solid-phase extraction (SPE). Each MIP was prepared by thermal polymerisation using catharanthine (or vindoline) as template, methacrylic acid (or itaconic acid) as functional monomer, ethylene glycol dimethacrylate (EDMA) as cross-linking agent and acetonitrile (or acetone) as porogenic solvent. For catharanthine-MIP, a SPE protocol (ACN-AcOH 99/1 washing and MeOH-AcOH 90/10 elution) allows a good MIP/NIP selectivity (imprinting factor 12.6). The specificity of catharanthine-MIP versus related bisindole alkaloids was assessed by cross-reactivity study. The catharanthine-MIP specifically retained catharanthine and its N-oxide analogue but displayed a weak cross-reactivity for other Vinca alkaloids (vinorelbine, vincristine, vinblastine, vindoline, vinflunine). It appears that the catharanthine-like unit of these molecules are hardly trapped in catharanthine cavities located in the MIP, probably due to the sterical hindrance of the vindoline moiety. Finally, the MIP-SPE applied to C. roseus extract enabled quantitative recovery of catharanthine (101%) and the total removal of vindoline. Its capacity was determined and was equal to 2.43 μmol g(-1). Vindoline is a weaker base than catharanthine, so the vindoline-MIP was achieved with a strong acidic monomer (itaconic acid) to increase vindoline-monomer interactions and a modified washing solvent (ACN-HCOOH 99/1) to reduce non-specific interactions. The influence of the amount of HCOOH (protic modifier) percolated during the washing step upon the elution yield and the imprinting factor for vindoline was investigated. This preliminary optimisation of the washing step, and in particular the number of moles of acid percolated, seems useful to emphasize the use of MIP in conditions of high selectivity or high yield. A compromise was obtained with an imprinting factor equal to 7.6 and an elution recovery of 33%. However MIP-vindoline failed to achieve a specific extraction of vindoline since catharanthine was also extracted probably because of strong non-specific interactions occurring between catharanthine and the sorbent.

Liquid chromatography analysis of phosphonic acids on porous graphitic carbon stationary phase with evaporative light-scattering and mass spectrometry detection.

The analysis of several phosphonic acids has been investigated by liquid chromatography (LC), using porous graphitic carbon as stationary phase with mass spectrometry (MS) or evaporative light-scattering detection (ELSD). In both detection modes (MS and ELSD), the mobile phase must be volatile and, due to the porous graphitic carbon (PGC) properties, should promote electronic interactions. Among the various hydrogeno- and perfluorocarboxylic acids tested, trifluoroacetic acid (0.1%, v/v) was selected as electronic competitor for solute retention. The baseline resolution of a phosphonic acids mixture required a trifluoroacetic-acetonitrile gradient elution. This methodology was then applied to the identification of phosphonic acids in a spiked tap water sample. Quantitative analyses are successfully achieved with a good correlation coefficient.

Determination of weak (2.0–2.5) dissociation constants of non-UV absorbing solutes by capillary electrophoresis

SummaryCapillary electrophoresis (CE) has proved to be a fast and convenient method for the determination of the dissociation constants of non-UV absorbing solutes in the acidic pKA range (2.0–2.5).The electroosmotic flow was reversed by washing the capillary with 0.2% polybren aqueous solution. A series of background electrolytes was prepared with phenylphosphonic acid (pKA=1.29) and β-alanine (pKA=3.55) with the same ionic strength and a high buffer capacity in order to improve the repeatability (0.1–0.2 %) of the electrophoretic mobility and to determine the values of pKA accurately.This procedure was applied to the determination of the dissociation constants of several alkyl-alkylphosphonic acids whose pKA values have not yet been published in the literature. In this work, their dissociation constants have been found to vary between 1.91 and 2.34 for alkyl-methylphosphonic acids and between 2.10 and 2.38 for alkyl-ethylphosphonic acids.

Factors influencing the separation of ionic and non-ionic chemical natural compounds in plant extracts by capillary electrophoresis

Abstract Capillary electrophoresis represents an alternative to the common liquid chromatographic technique in natural product research because of the high-resolution separations obtained by several migration modes. Natural molecules such as flavonoids, flavonoid-O-glycosides, coumarins, phenolic acids, glucosinolates or desulphoglucosinolates, have been resolved by several electrophoretic techniques (free solution capillary electrophoresis with a no-complexing or complexing electrophoretic buffer and by micellar electrokinetic capillary chromatography with a cationic or anionic surfactant). In free solution capillary electrophoresis (CE), the selectivity of the flavonoid-O-glycosides, using a borate buffer, may be explained by in-situ borate complexation of both the sugar moiety and the cis-1, 2-hydroxyl groups on the flavonoid skeleton, and also by the ionization of hydroxyl groups on the flavonoid skeleton in such alkaline pH conditions. In particular, the magnitude of the borate complexation depends o...

Capillary electrophoresis analysis of chemical warfare agent breakdown products I. Counterelectroosmotic separation of alkylphosphonic acids and their monoester derivatives

An analytical method has been developed for the analysis of chemical warfare agents by capillary zone electrophoresis (CZE). Sorbate anion has been selected as indirect UV-absorbing agent and decamethonium cation reduces the electroosmotic flow. The optimized electrolyte (5 mM sorbic acid, 0.1 mM decamethonium bromide pH 6) allows the simultaneous separation of alkylphosphonic acids and their monoester derivatives in less than 15 min. The measurement of phosphonic solutes has been achieved in spiked soils using calibration curves in the 5-50 mg/l concentration range.

Comparison of native, alkylated and charged cyclodextrins for the chiral separation of labetalol stereoisomers by capillary electrophoresis

A capillary electrophoresis method for the enantioresolution of labetalol was developed using CDs as chiral selectors and uncoated capillaries. Various native (α-, β- and γ-CD), alkylated (hydroxy propyl-β- and γ-CD, methyl-β-CD) and anionic (sulfated-β-CD and sulfobutylether-γ-CD) cyclodextrins were tested and operational parameters such as buffer pH, concentration of CD were investigated. Propranolol was also studied as a model compound. Uncharged γ-CDs were more effective than β-CDs to separate the enantiomers of labetalol but no complete resolution of the four isomers was obtained. The use of charged cyclodextrins led to a combination of hydrophobic inclusion and ion-pairing interaction in the chiral recognition mechanism. Thus, a complete resolution of the four enantiomers of the labetalol was attained using 7.7 g/l sulfated-β-CD in a 30 mM phosphate buffer, pH 6.5.