Reversed-phase liquid chromatography organic solvent

Khaledi, M. G., J. K. Strasters, A. H. Rodgers, and E. D. Breyer. 1990. Simultaneous enhancement of separation selectivity and solvent strength in reversed-phase liquid chromatography using micelles in hydro-organic solvents nal. Chem62 130-136. 

Bielejewska, A., Duszcyk, K., Sybilska,D. (2001) Influence of organic solvent on the behavior of camphor and a-pinene enantiomers in reversed-phase liquid chromatography systems with a-cyclodextrin as chiral additive, J.Chromatog. A 931, 81-93 ... 

Reversed phase liquid chromatography (RPLC) allows the separation of analytes with different hydrophobicity and polarity characteristics. It has good selectivity mobile phases used in the technique contain organic solvents and small amounts of inorganic salts [149]. The effectiveness of the process depends on the hydrophobicity of the separated analytes. Charged substances must first be transformed into neutral derivatives (e.g., by adding appropriate anti-ions into the mobile phase). 

Micelles and cyclodextrins are the most common reagents used for this technique. Micellar electrokinetic capillary chromatography (MECC or MEKC) is generally used for the separation of small molecules [6], Sodium dodecyl sulfate at concentrations from 20 to 150 mM in conjunction with 20 mM borate buffer (pH 9.3) or phosphate buffer (pH 7.0) represent the most common operating conditions. The mechanism of separation is related to reversed-phase liquid chromatography, at least for neutral solutes. Organic solvents such as 5-20% methanol or acetonitrile are useful to modify selectivity when there is too much retention in the system. Alternative surfactants such as bile salts (sodium cholate), cationic surfactants (cetyltrimethy-lammonium bromide), nonionic surfactants (poly-oxyethylene-23-lauryl ether), and alkyl glucosides can be used as well. 

The use of positive-ion ESI with reverse-phase liquid chromatography is common for most methods. Mobile phases are often a combination of either methanol or acetonitrile combined with water that contains additives such as acids or volatile ammonium buffers. Optimization between increased sensitivity with a higher percent of organic solvent in the mobile phase and adequate retention on the column is important for LC-MS/MS detection. 

Alternatively, the following empirical relationships can be used to estimate the equivalent solvent composition of isoeluotropic binary solvent mixtures given that experimental conditions have been established using methanol as the organic modifier in reversed-phase liquid chromatography... 

Typical water miscible organic solvents (e.g. methanol, acetonitrile, etc.) affect selectivity in micellar electrokinetic chromatography in a manner similar to reversed-phase liquid chromatography. The range of composition variation, however, is restricted by instability of the micelles at moderate concentrations of organic solvents, and... 

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