Liquid chromatography/ultraviolet/mass analysis

This LC-MS experimental approaches used in NEF metabolite identification are also routinely employed for in vitro metabolism comparisons across species, in which liver microsomes or hepatocytes from humans and animal species are used. In addition, liver microsomal incubations followed by metabolite identification and quantitative estimation using liquid chromatography-ultraviolet/ mass spectrometry (LC-UV/MS) is an approach commonly taken to determine metabolic soft spots, where a major metabolic reaction takes place (Table 6.10). Use of UV detection allows for quantitative analysis of major metabolites in the absence of chemical standards, with the assumption that the metabolic reaction did not disturb the molecule s UV chromophore. 

C.W. Huck, M.R. Buchmeiser and G.K. Bonn, Fast analysis of flavonoids in plant extracts by liquid chromatography-ultraviolet absorbance detection on poly(carboxylic acid)coated silica and electrospray ionization tandem mass spectrometric detection. J. Chromatogr.A 943 (2002) 33-38. 

Cheng Y.F. Lu Z. Neuc U. Ultrafast liquid chromatography/ultraviolet and liquid chromatography/tandem mass spectrometric analysis. Rapid Communications in Mass Spectrometry, 2001, 15, 141-151. 

Is the method specific and stability indicating as shown by analysis of samples subjected to stressed stability studies (pH, light, heat, oxidation) Normally, specificity is determined through peak purity using ultraviolet (UV) diode array or liquid chromatography (LC)/mass spectrometry (MS) analysis. In methods for analysis of drug products, placebo formulations (and stressed placebos) must yield blank chromatographic baselines. The development... 

Cheng, Y.-R Lu, Z. Neue, U. Ultrafast Liquid Chromatography/Ultraviolet and Liquid Chromatography/Tandem Mass Spectrometric Analysis, Rapid Commun. Mass Spectrom. 15, 141-151 (2001). 

The quality control analyses of these chemicals are performed using almost the whole range of trace analysis techniques available. Among the most important are atomic absorption spectrophotometry in all its forms, ICP emission spectrometry, and ICP mass spectroscopy, ion chromatography, gas and liquid chromatography, ultraviolet and visible absorption spectrophotometry, voltammetry, and spectro-fluorimetry. 

Very little in the way of advances has occurred since 1971 in the applications of ultraviolet or infrared spectroscopy to the analysis of fluonnated organic compounds Therefore, only gas-liquid chromatography, liquid chromatography, mass spectrometry, and electron scattering for chemical analysis (ESCA) are discussed The application of nuclear magnetic resonance (NMR) spectroscopy to the analysis of fluonnated organic compounds is the subject of another section of this chapter... 

Cuyckens F and Claeys M. 2002. Optimization of a liquid chromatography method based on simultaneous electrospray ionization mass spectrometric and ultraviolet photodiode array detection for analysis of flavonoid glycosides. Rapid Commun Mass Spectrom 16(24) 2341—2348. 

Chromatographic methods are also often used as part of systems that are called hyphenated methods, (see Chapter 15) where the output of the chromatographic section is used as the input for an identification method such as mass spectrometry. These hyphenated methods are also most often referred to by their acronyms, for example, GC-MS—gas chromatography-mass spectrometry and HPLC-MS—high-performance liquid chromatography-mass spectrometry. Note that although ultraviolet-visible (UV-Vis) is hyphenated, it is not a hyphenated method in that it does not consist of two different methods of analysis. Hyphenated methods will be discussed fully in Chapter 15. 

A wide array of laboratory techniques and instrumentation is used in forensic studies. This includes ultraviolet, infrared, and visible spectrophotometry neutron activation analysis gas chromatography and mass spectrophotometry high pressure liquid chromatography and atomic absorption spectrophotometry. The techniques and instrumentation chosen depend on the type of sample or substance to be examined. 

High-pressure liquid chromatography (HPLC) with fluorescence detection, ion-trap mass spectrometry (ITMS), and capillary electrophoresis (CE) with ultraviolet detection, three novel detection techniques for the analysis of cocaine in hair, have been evaluated by Tagliaro et aU°- and Traldi et al. The HPLC technique was highly sensitive, capable of detecting 0.015 ng/mg of cocaine in hair. The CE method was sensitive and highly selective. ITMS analysis of hair readily demonstrated the presence of cocaine in hair, but cocaine metabolites were more difficult to identify. 

Reversed-phase high-performance liquid chromatography (RP-HPLC) is the usual method of choice for the separation of anthocyanins combined with an ultraviolet-visible (UV-Vis) or diode-array detector (DAD)(Hebrero et al., 1988 Hong et ah, 1990). With reversed-phase columns the elution pattern of anthocyanins is mainly dependent on the partition coefficients between the mobile phase and the Cjg stationary phase, and on the polarity of the analytes. The mobile phase consists normally of an aqueous solvent (water/carboxylic acid) and an organic solvent (methanol or acetonitrile/carboxylic acid). Typically the amount of carboxylic acid has been up to 10%, but with the addition of a mass spectrometer as a detector, the amount of acid has been decreased to as low as 1 % with a shift from trifluoroacetic acid to formic acid to prevent quenching of the ionization process that may occur with trifluoroacetic acid. The acidic media allows for the complete displacement of the equilibrium to the fiavylium cation, resulting in better resolution and a characteristic absorbance between 515 and 540 nm. HPLC separation methods, combined with electrochemical or DAD, are effective tools for anthocyanin analysis. The weakness of these detection methods is a lack of structural information and some nonspecificity leading to misattribution of peaks, particularly with electrochemical... 

Methods in the analysis of drug impurities (e.g., ultraviolet, UV Fourier transform infrared, FT-IR nuclear magnetic resonance, NMR mass spectrometry, MS) are used to separate, identify, and quantify impurities, as well as establish their structure. Currently the most efficient methods seem to be combined techniques such as GC-MS, LC-MS, liquid chromatography-diode-array detection-mass spectrometry (LC-DAD-MS), LC-NMR, LC-DAD-NMR-MS, etc. [18-20]. 

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