Reverse phase method development sample matrix

For biological samples where the analytes often are non-volatile and/or occur in an aqueous matrix, the reversed-phase mode, using a hydrophobic stationary phase and an aqueous mobile phase is extremely useful. The usefulness and popularity of HPLC was further increased by the possibility to automate and computerize the systems providing unattended operations and high sample capacities. Many Nobel Prize awards have been based upon work in which chromatography played an important role [5], Most recently, the 2002 Nobel Prize in chemistry was awarded to the development of methods for identification and structure analyses of biological macromolecules" in which HPLC and Mass spectrometry (MS) were used [6],... 

Was the published method developed in distilled water or in a real sample matrix Distilled water has unusual properties with respect to mechanisms of ion exchange and to some degree for reversed phase. It is critical that the method is robust and that it is capable of working across a wide range of pH and ionic strengths. 

An understanding of simple methods development is crucial to developing effective environmental applications of solid-phase extraction (SPE). The lour mechanisms outlined in Chapter 2 (reversed phase, normal phase, ion exchange, and mixed mode) are sufficient for the majority of SPE applications in environmental analysis. The molecule s structure and the sample matrix are (he main factors that will determine which mechanism of isolation and separation will be the most appropriate. The fundamental approach to selection of sorbents will be a key topic and many examples are given. This chapter will also discuss applications of SPE to environmental matrices. These include water, soil, and air, for a variety of compounds. 

An electrostatic ion chromatographic method was developed for the direct determination of iodide, bromide and nitrate in seawater (Hu et ai, 1999). An octadecylsifica column modified with a zwitterionic surfactant 3- NJV-dimethylmyristylammonio)propane-suifonate was used as the stationary phase, and an electrolytic solution was used as the eluent. The matrix species (such as chloride and sulfate) were retained weakly, and showed iitde or no interference. The method was applied to the determination of iodide, bromide, and nitrate in artificial seawater, giving detection fimits of 0.8 p,g 1 for iodide, 0.75 p.g 1 for bromide, and 0.52 p,g 1 for nitrate, and relative standard deviations of <1.2%. The real seawater samples were also analyzed successfully. Later, another electrostatic ion chromatographic method was developed for the determination of iodide in seawater by the same research group (Hu et ai, 2002). A reversed-phase ODS column was... 

Solid-phase extraction (SPE) using small, disposable cartridges, columns, or disks is employed for isolation and cleanup of pesticides from water and other samples prior to TLC analysis, especially using reversed-phase (RP) octa-decyl (C-18) bonded silica gel phases. Microwave-assisted extraction (MAE) is a time- and solvent-saving method for removing residues from samples such as soils. Supercritical fluid extraction (SEE) has been used for sample preparation in the screening of pesticide-contaminated soil by conventional TLC and automated multiple development (AMD). Ultrasonic solvent extraction (USE) and videodensitometry have been combined for quantification of pesticides in sod. Matrix solid-phase dispersion (MSPD) with TLC and GC has been used to determine diazinon and ethion in nuts. 

Separation and pre-concentration. A method first suggested by Knapp et al. (1975), developed further by Prange (1983) for seawater analysis using TXRF and later slightly modified by Schmidt et al. (1993) combines matrix removal with enrichment by a factor of about 50. The elements V, Mn, Fe, Co, Ni, Cu, Zn, Se, Mo, Cd, Pb and U are chelated using sodium-dibenzyldithiocarbamate (Na-DBDTC). The water-insoluble chelates are adsorbed on a column of a reversed-phase silica gel. Alkaline and alkaline earth elements elute unchelated with the aqueous phase and are discarded. The chelate complexes are removed from the dried column with a mixture of chloroform and methanol (methanol is needed for the quantitative elution of Mo). Some 100/tL of this extract are evaporated on quartz sample carriers and measured for around 1000 s. 

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