Qualitative Colorimetric Tests for Solid Phase Synthesis

This chapter provides an overview of the most widely used qualitative tests including instructions on reagent preparation and storage, experimental protocol, and the scope and limitations of each test. The majority of these tests can be performed in less than 10 minutes with simple laboratory equipment and minimal reagent preparation. The chapter also reports the results of experiments to determine the functional group response of each test using amino acids as representative of organic compounds. To determine the utility of each test in the presence of multiple functional groups, each of the summarized colorimetric tests was applied against a broad range of amino acids. It was performed to determine the universality of each test, and to identify and report those cases with unusual results. The amino acids were tested for the presence of each functional group under all possible levels of amine and lateral chain protection, thus enabling the determination of the extent to which the chemical interference by other functional groups could affect each test. The chapter also investigates the use of each test at medium (approximately 0.5 mmol/g) and low (approximately 0.025 mmol/g) resin loading. Due to the numerous factors that can influence colorimetric test results (like test reagent stability, resin type, functional group interference, and lability of protecting group), performing a positive and a negative control for any test applied to a new synthesis is recommended. To minimize false results due to the lability of the resin–product bond or the product itself (such as Fmoc-protected amino acids), colorimetric tests should be performed with utmost immediacy in regards to the completion of the step to be monitored.