Cancionero popular de la provincia de Granada. Volumen V

Capillary electrophoresis (CE) was developed from combining several features of different methods, including the principle of gel electrophor- esis, the fused silica capillary of gas chromatography and the highly sen- sitive detectors of HPLC (Suntornsuk, 2007). CE was introduced in the early 1980s. It is an automated analytical and separation technique driven under the influence of the electric field that occurs in a capillary filled with buffer. Separation is based on the differential movement of ions under the applied electric field. The size, charge, degree of ionization, viscosity, etc., of the analyte controls the speed of movement. Three mechanisms are accounted for in separation: (i) differences in analyte mobilities; (ii) differ- ences in analyte isoelectric points; and (iii) differences in analyte conduct- ivity. Instrumentation for CE consists of electrodes, sample introduction systems, a capillary, voltage supply, detector and liquid-handling system. Detection in CE can be performed directly on a capillary. UV-diode array, electrochemical and spectroflurometer are the common online detectors, while a mass spectrometer can be used as an external detector. Liquid- handling systems include an auto sampler, buffer replenishment, buffer levelling and fraction collector.

CE is promising for the separation and analysis of active ingredients in natural products as it needs only small amounts of standards and can analyse samples rapidly. It is also a good tool for generating chemical fin- gerprints because it has similar technical characteristics to liquid chro- matography. Quantitative analysis is done mainly by the comparison of the mobility or migration time of the compounds of interest with those of the standards. Quantitative analysis is calculated from the peak height or peak area based on a calibration curve of a standard. CE is also recom- mended in several pharmacopeia.

The situation of CE analysis in hyphenation development is somewhat like HPLC analysis. The hyphenated CE instruments such as CE-PDA, CE-MS and CE-NMR have already been reported. Online coupling of CE with MS and other spectrometry allows both efficient separation by CE and specific and sensitive detection to be achieved.

The mass spectra of the separated analyte are recorded using mass spectrometric detection. Selectivity and sensitivity of this hyphenated technique are high. The ESI-MS detection method is generally used in most of the CE-MS techniques, since ESI is considered to be appropriate for providing information about molecular weight and structural characteris- tics (Dunayevskiy et al., 1996). The coupling of CE with MS creates some complexity as MS cannot accept CE solvents or electrolytes, and conven- tional gas-phase ionization in MS is not suitable for compounds separated by CE, which are thermally labile, polar or have a high molecular weight. The coupling of CE-MS was introduced successfully in 1988 (Smith and Udseth, 1988). Much progress has been made since then to make CE-MS applicable for analysis of a wide variety of analytes. ESI can be performed

by either liquid support or non-liquid support. Sheath liquid is chosen to overcome the volatility and conductivity problems of CE buffer, which usu- ally has poor electrospray capability. Sheath liquid is usually composed of organic solvents (acetonitrile, methanol and 2-propanol) and acidic or basic buffer to promote the ionization of positive and negative ions, respectively. CE can be coupled to different mass analysers such as quadruple, ion trap (IT) and TOF. IT and TOF offer high speed and sensitivity. IT can be used for MSn _{experiments, whereas TOF gives high mass resolution and the pos-}

sibility of measured mass accuracy.

CE is important for drug discovery, manufacturing and other aspects such as determination of active ingredients, enantiomeric separation, etc. Although HPLC is still predominant, CE will continue to serve as a com- plementary technique in natural product research. It is an ideal technique to be miniaturized on a microchip (Suntornsuk, 2007).

6.8 Conclusion

Smaller, faster and high-resolving instruments for high-throughput ana- lysis in a short time and with low cost are desirable. Hyphenated methods create unprecedented opportunities and their main advantages include extremely low limits of detection and quantification, high precision and repeatability of the determinations. On the other hand, hyphenated methods have their own limitations, such as high price and complexity of the apparatus. Consequently, they are not in general laboratory usage. The application of a hyphenated method requires perfect understanding of analytical methodologies and apparatus. These systems are expensive and are used in scientific research rather than for routine analysis.

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7

Non-destructive Techniques

7.1 Introduction

Rapid analytical techniques have been developed as alternatives to the traditional methods of analysis. The application of quick, solvent-free and on-field analytical techniques present an enormous advantage over the conventional wet chemistry approaches (Pedro and Ferreira, 2005). In par- ticular, fast, reliable and non-destructive analysis with minimal sample preparation is required for the quantification of valuable components in screening a large number of samples. These techniques are also finding application in online, semi-continuous monitoring of composition, as sample preparation is minimal in these non-destructive techniques. Furthermore, conventional chemical analysis such as high-performance liquid chromatography (HPLC) or gas chromatography (GC) may take sev- eral hours to generate results, which may delay the validation of quality control. Therefore, analytical techniques generating rapid analytical re- sults as compared to conventional methods are required.