Separation techniques have become unavoidable in the drug development process. All samples must be considered to consist of several components. It is important to verify that a compound developed in the initial research phase primarily contain the expected compound and not a number of major impurities that may be responsible for effect or side effects, but even a “pure” API do always contain minor impurities which have to be controlled. The formulated drug products contain a lot of excipients and have to be tested for stability where new impurities may be formed. In the clinical trials samples of plasma, urine, tissue etc. also have to be analysed for the drug substance and its metabolites often in very low concentrations. Therefore all these samples must be considered as complex samples and separation is needed in order to determine each component in a selective and reliable way.
Liquid chromatography is most useful for such qualitative and quantitative analysis and is performed using high performance liquid chromatography (HPLC) or ultra high performance liquid chromatography equipment (UHPLC). Using these techniques high efficiency separations are achieved, and in this chapter these techniques are described in detail.
High performance liquid chromatography (HPLC) Ultrahigh performance liquid chromatography (UHPLC) Separation theory Separation selectivity Separation efficiency Detection API purity Drug stability Sample preparation Validation
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