Chapter 2 High performance liquid chromatography (HPLC): Principles and applications

This chapter focuses on high performance liquid chromatography (HPLC), which is an instrumental analytical method that gained increased acceptance mainly because it met two basic factors: (1) the need for a wide range of rapid analyses for nutrients, and (2) the need for methods that can be easily automated. HPLC is applied to several categories of substances: carbohydrates, lipids, vitamins, additives, synthetic colorings, natural pigments, contaminants, and amino acids. HPLC is a form of liquid chromatography, where separation occurs between a mobile phase and a stationary phase. Depending on the nature of the stationary phase, the separation process can be of four different modes: adsorption chromatography, partition chromatography, ion-exchange chromatography, and size exclusion chromatography. The choice of the solvent depends on the nature of the operation mode, that is, isocratic or gradient elution. The polarity for such an elution medium can, therefore, vary from buffered aqueous solutions to hydrocarbons.