Laboratory capillary electrophoresis

Thompson, L. Veening, H. Strain, T. G. Capillary Electrophoresis in the Undergraduate Instrumental Analysis Laboratory Determination of Common Analgesic formulations, /. Chem. Educ. 1997, 74, 1117-1121. 

Capillary electrophoresis employing chiral selectors has been shown to be a useful analytical method to separate enantiomers. Conventionally, instrumental chiral separations have been achieved by gas chromatography and by high performance liquid chromatography.127 In recent years, there has been considerable activity in the separation and characterization of racemic pharmaceuticals by high performance capillary electrophoresis, with particular interest paid to using this technique in modem pharmaceutical analytical laboratories.128 130 The most frequently used chiral selectors in CE are cyclodextrins, crown ethers, chiral surfactants, bile acids, and protein-filled... 

E. Jackim and L. Watts Jackim (eds), Capillary Electrophoresis Procedures Manual A Laboratory User s Aid for Quick Starts, Elsevier Science Publishers, Amsterdam (1996). 

Capillary electrophoresis systems are also likely to play an increasingly prominent analytical role in the QC laboratory (Figure 7.2). As with other forms of electrophoresis, separation is based upon different rates of protein migration upon application of an electric field. 

Some coupled systems allow measurement of the main N and P forms (nitrate, ammonia and orthophosphates) [22,27,29], among which is a system based on membrane technology in combination with semi-micro continuous-flow analysis (pCFA) with classical colorimetry. With the same principle (classical colorimetry), another system [30] proposes the measurement of phosphate, iron and sulphate by flow-injection analysis (FIA). These systems are derived from laboratory procedures, as in a recent work [31] where capillary electrophoresis (CE) was used for the separation of inorganic and organic ions from waters in a pulp and paper process. Chloride, thiosulphate, sulphate, oxalate,... 

Many of the more established techniques have been validated through collaborative studies which becomes of greater importance as laboratories seek to become accredited via ISO, EN or related systems where the use of official or well validated methods is mandatory. New instrumental techniques are constantly being reported in the literature but it often requires many years before procedures are introduced, validated and then applied within the food industry. Recent techniques that can be included in this category are capillary electrophoresis and liquid chromatography-mass spectrometry (LC-MS). In time procedures based on these techniques will also become accepted as routine methods and are likely to be adopted by some of the official international bodies like the AOAC International, CEN, ISO, etc. 

Tim Wehr is Staff Scientist at Bio-Rad Laboratories in Hercules, California. He has more than 20 years of experience in biomolecule separations, including development of HPLC and capillary electrophoresis methods and instrumentation for separation of proteins, peptides, amino acids, and nucleic acids. He has also worked on development and validation of LC-MS methods for small molecules and biopharmaceuticals. He holds a B.S. degree from Whitman College, Walla Walla, Washington, and earned his Ph.D. from Oregon State University in Corvallis. 

With capillary electrophoresis (CE), another modern primarily analytically oriented separation methodology has recently found its way into routine and research laboratories of the pharmaceutical industries. As the most beneficial characteristics over HPLC separations the extremely high efficiency leading to enhanced peak capacities and often better detectability of minor impurities, complementary selectivity profiles to HPLC due to a different separation mechanism as well as the capability to perform separations faster than by HPLC are frequently encountered as the most prominent advantages. On the negative side, there have to be mentioned detection sensitivity limitations due to the short path length of on-capillary UV detection, less robust methods, and occasionally problems with run-to-run repeatability. Nevertheless, CE assays have now been adopted by industrial labs as well and this holds in particular for enantiomer separations of chiral pharmaceuticals. While native cyclodextrins and their derivatives, respectively, are commonly employed as chiral additives to the BGEs to create mobility differences for the distinct enantiomers in the electric field, it could be demonstrated that cinchona alkaloids [128-130] and in particular their derivatives are applicable selectors for CE enantiomer separation of chiral acids [19,66,119,131-136]. 

Over the past 40 years, capillary electrophoresis (CE) has advanced significantly as a technique for biomolecular characterization. It has not only passed the transition from a laboratory curiosity to a mature instrumental-based method for micro-scale separation, but also emerged as an indispensable tool in the biotech and pharmaceutical industries. CE has become a method of choice in research and development (R D) for molecular characterization, and in quality control (QC) for the release of the therapeutic biomolecules.In the biopharmaceutical industry, more and more CE methods have been validated to meet International Conference on Harmonization (ICH) requirements. In this chapter, we present real industrial examples to demonstrate the role of CE in R D of pharmaceutical products. The focus in this chapter is on method development analytical control for manufacturing and release of therapeutic proteins and antibodies. 

Capacity Another consideration is the capacity of the laboratory to handle the work involved in a given method. For example, a capillary electrophoresis method would not be chosen if the laboratory does not have the instrument. It is also important to look at such factors as other equipment needed, the supplies needed, etc., or whether the laboratory can follow the required health and safety regulations, etc. Sometimes there may not be enough manpower or equipment to handle the sample work load. In that case, storage and refrigeration can also be a problem. 

F. Huber, Good Laboratory Practice A Primer for High Performance Liquid Chromatography, Capillary Electrophoresis, and UV-Visible Spectroscopy, Hewlett-Packard Publ. 12-5091-6259E, 1993. 

Because of these ever-widening interests, the measurement of plasma tHcy is undertaken in many clinical chemistry and routine laboratories. Various methods are employed, including high-performance liquid chromatography (HPLC) assays, conventional amino acid analysis, capillary electrophoresis, gas chromatography with or without mass spectrometry, liquid chromatography with tandem mass spectrometry, and in many routine clinical chemistry laboratories immunoassays. In this chapter, those methods that are often available in laboratories involved in the investigation of inborn errors of metabolism are described, namely HPLC and tandem mass spectrometry. 

Emerging separation technologies such as capillary electrophoresis may lead to the improved and cost-effective resolution of complex mixtures of drug residues. This is a valuable tool that is finding its place as a workhorse technique in the analytical laboratory. Capillary electrophoresis is expected to provide a wide variety of new methods that may well prove superior to currently available technology. 

Experimental setup for capillary electrophoresis. Courtesy of Bio-Rad Laboratories, Life Science Group, Hercules, CA. 

Extremely low level detection work is being performed in analytical chemistry laboratories Detection of rhodamine 6G at 50 yoctomole (50 > . 10-24 mol) has been reported using a sheath flow cuvette for fluorescence detection following capillary electrophoresis. This represents 30 molecules of rhodamine, a highly fluorescent molecule. See also Electrophoresis. 

Finally, equipment for assaying and analyzing the preparations is needed. Most such equipment is fairly standard in biochemical laboratories and includes spectrophotometers, scintillation counters, analytical gel and capillary electrophoresis apparatuses, immunoblot-ting materials, and immunochemical reagents. 

Automation in the Clinical Laboratory Biosensor Design and Fabrication Capillary Electrophoresis in Clinical Chemistry DNA Arrays Preparation and Application Drugs of Abuse, Analysis of Molecular Biological Analyses and Molecular Pathology in Clinical Chemistry Nucleic Acid Analysis in Clinical Chemistry Phosphorescence, Fluorescence, and Chemiluminescence in Clinical Chemistry Product Development for the Clinical Laboratory... 

Several instruments have been developed in various laboratories since the late 1970s. Currently, several companies have introduced the capillary electrophoresis commercially (for example Microphoretic Systems, Sunnyvale, California Bio-Rad, Richmond, California Applied Biosystems, Inc., Foster City, California and Beckman Instruments, Inc., Palo Alto, California). Although the instruments have many practical features for the separation and analysis of analytes, several new features need to be incorporated (for routine use) by protein chemists. 

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