Bioautography for Rapid Detection, Isolation, and Identification of Anti-Tuberculosis Leads

Classical “bioactivity-guided isolation” has major shortcomings with regard to structure dereplication, potency and selectivity of the active principle. The present project is aimed at overcoming these constraints by developing a new 3-D approach using TLC-MS-Bioautography, which allows the early detection, isolation and identification of anti-TB natural products from cultured actinomycete strains. By aligning high-performance TLC with LC-MS and superimposing the structural information with a new agar-overlay bioassay specifically developed for TB, the active principle(s) can be chemically and biologically classified at an early stage of the isolation process. Gaining chemical information at a much earlier stage allows a more targeted process which utilizes counter-current chromatography and significantly shortens the isolation procedure of the active component(s) to 2 – 3 steps. State-of-the-art technologies are used for structure de-replication including detailed resolution of complex ¹H NMR spin systems of recurring structural elements and assessment by quantitative NMR, which allows calculation of quantitative purity-activity relationships concurrent with structural analysis of the active principles. Establishing the proof of concept is demonstrated with the identification of highly anti-TB active cyclic peptides from cultured actinomycete strains.