ABSTRACT

This chapter explains in addition to the methods based on reversible acid-base equilibria, there are several cases in pharmaceutical analysis in which an irreversible rearrangement or elimination reaction or even the cleavage of the molecule takes place by acid catalysis, and this reaction involves an increase in spectrophotometric activity. Although with the use the 4-nitro-derivative of the reagent the reaction can be used in direct spectrophotometry, too, it is generally applied in high performance column chromatography with UV detection as a standard derivatization technique of fatty acids, bile acids, etc. If acylation is carried out with an acid chloride, it is worth running the reaction in the presence of an acid binding tertiary amine to make it quantitative. Ion pair formation between heparin, containing acidic groups, and various dyes may also form the basis of spectrophotometric methods, which were used for the determination of heparin in biological samples since the results were, to a certain extent, correlated with anticoagulant activity.