Second order retention effects in reversed-phase liquid chromatography. 1. Influence of solute size

Retention times or volumes in reversed-phase liquid chromatography are substantially influenced by partial steric exclusion of solutes from the pore space of the usual octadecyl silica column packings. Contrary to the common view that exclusion effects become appreciable only with “large” molecules, they are clearly observable even with solutes of a size similar to or even smaller than eluent molecules. The extent of exclusion was directly determined from elution volume versus carbon number plots of n-alkanes with n-pentane eluent. Using high precision (relative standard deviation <0,1%) retention data with methanol eluent, it was found that the “effective” dead volume depends on solute chain length. If such data is corrected for partial exclusion, corresponding log (capacity factors) as functions of carbon number are absolutely linear which is equivalent to perfectly constant methylene selectivity, α, within the n-alkane series. Since this observation was made on various columns with thousands of data, it may be regarded as a case of experimental proof of Martin's postulate of additivity of retention increments of molecular constituents.