PHYTOCHEMISTRY AND ANTIMYCOBACTERIAL ACTIVITIES OF THE LEAF EXTRACTS OF PAVETTA CRASSIPES K. SCHUM

Abstract

Tuberculosis (TB), an infectious disease mainly caused by Mycobacterium tuberculosis is the second greatest contributor among infectious diseases to adult mortality, causing approximately two million deaths annually worldwide. Effective treatment of TB has been hampered by the emergence of drug resistant strains of M. tuberculosis. There is therefore an increasing need for the development of new antituberculosis drugs to combat this disease. Pavetta crassipes (Rubiaceae) is a plant which has been claimed as a traditional cure for TB and other respiratory diseases in Northern Nigeria. The aims of this study were to isolate, identify the constituents responsible for the reported biological activities of P. crassipes leaves against M. tuberculosis and subsequently evaluate the activity of the pure isolated compounds against this organism. Pavetta crassipes leaf was collected at Suleja and its identity was confirmed in the herbarium of the National Institute for Pharmaceutical Research and Development, Abuja. Cold maceration was employed in the successive extraction of air-dried leaves of P. crassipes using hexane, ethyl acetate and methanol. Separation and isolation of the plant constituents were achieved by column chromatography, preparative high performance liquid chromatography and preparative thin layer chromatography. Structures of the isolated compounds were established by spectroscopic methods (infra-red spectroscopy, ultraviolet spectroscopy, mass spectrometry, 1D and 2D- nuclear magnetic resonance spectroscopy) and chemical methods. The antimycobacterial activity of P. crassipes extracts, fractions and constituents was determined against M. tuberculosis H37Rv using the broth microdilution method, with isoniazid as the reference compound. Isolated compounds were also tested for antimycobacterial activity using the green fluorescence reporter assay method. Ursolic acid, ?-sitosterol, 3-caffeoyl 1-ethyl quinate, 3-caffeoyl 1-methyl quinate, quercetin 3-O-?-rutinoside, D-mannitol and a novel monoterpene iridoid glucoside, pavetoside were isolated and structurally elucidated. The ethyl acetate and methanol extracts exhibited antimycobacterial activities with Minimum Inhibitory Concentration (MIC) of 250 and 521 ?g/mL, respectively against M. tuberculosis. Ursolic acid, 3-caffeoyl 1-methyl quinate, and 3-caffeoyl 1-ethyl quinate were found active against M. tuberculosis with MIC of 200, 100, and 50 ?g/mL, respectively. ?-sitosterol, quercetin 3-O-?-rutinoside, D-mannitol and pavetoside did not exhibit any significant activity against M. tuberculosis. The antimycobacterial activity of Pavetta crassipes was attributed to ursolic acid, 3-caffeoyl 1-ethyl quinate, and 3-caffeoyl 1-methyl quinate. The presence of antimycobacterial terpenoid and quinate esters in leaves of Pavetta crassipes provides scientific evidence for the ethnomedicinal use of this plant as a traditional antituberculosis remedy.