Mycobacterium neoaurum has the ability to produce steroidal intermediates known as 22-hydroxy-23, 24-bisnorchol-4-en-3-one (BA) upon the knockout of the genes for either the hydroxyacyl-CoA dehydrogenase (Hsd4A) or acyl-CoA thiolase (FadA5). In a previous study, we discovered a novel metabolite in the fermentation products when the fadA5 gene was deleted. This research aims to elucidate the metabolic pathway of this metabolite through structural identification, homologous sequence analysis of the fadA5 gene, phylogenetic tree analysis of M. neoaurum HGMS2, and gene knockout. Our findings revealed that the metabolite is a C23 metabolic intermediate, named 24-norchol-4-ene-3, 22-dione (designated as 3-OPD). It is formed when a thioesterase (TE) catalyzes the formation of a β-ketonic acid by removing CoA from the side chain of 3, 22-dioxo-25, 26-bisnorchol-4-ene-24-oyl CoA (22-O-BNC-CoA), followed by spontaneously undergoing decarboxylation. These results have the potential to contribute to the development of novel steroid intermediates.
Mycobacterium/metabolism* ; Phylogeny ; Steroids/metabolism* ; Metabolic Networks and Pathways ; Sterols/metabolism*

OBJECTIVETo investigate the chemical constituents of Nauclea latifolia, which is an African folk medicine and collected from Guinea.

METHODThe chemical constituents were isolated by silica gel and Sephadex LH-20 column chromatographies, and structurally elucidated by spectral evidence together with physiochemical properties.

RESULTTwelve compounds including 8 triterpenes and 4 sterols were isolated from the roots of N. latifolia, and their structures were defined as 24-en-cycloartenone (1), ursolic aldehyde (2), quinovic acid (3), rotundic acid (4), 3beta,19alpha,23,24-tetrahydroxyurs-12-en-28-oic acid (5), pyrocincholic acid 3beta-O-beta-D-fucopyranoside (6), quinovic acid 3beta-O-beta-D-glucopyranoside (7), quinovic acid-3b-O-D-glucopyranosyl-(28-->1)-beta-D-glucopyranosyl ester (8), beta-sitosterol (9), stigmastan-3,6-dione (10), stigmast-4-en-6beta-ol-3-one (11) and daucosterol (12) .

CONCLUSIONAll compounds except for 4,9, andl2 are isolated from this plant for the fist time, while compounds 2, 6, 10, and 11 are isolated from the genus Nauclea for the first time.

Chromatography ; methods ; Guinea ; Magnetic Resonance Spectroscopy ; methods ; Plant Extracts ; chemistry ; isolation & purification ; Plant Roots ; chemistry ; metabolism ; Plants, Medicinal ; chemistry ; Rubiaceae ; chemistry ; metabolism ; Sterols ; chemistry ; isolation & purification ; Triterpenes ; chemistry ; isolation & purification

Antifungal drug resistance is a significant clinical problem, and antifungal agents that can evade resistance are urgently needed. In infective niches, resistant organisms often co-existed with sensitive ones, or a subpopulation of antibiotic-susceptible organisms may evolve into resistant ones during antibiotic treatment and eventually dominate the whole population. In this study, we established a co-culture assay in which an azole-resistant Candida albicans strain was mixed with a susceptible strain labeled with green fluorescent protein to mimic in vivo conditions and screen for antifungal drugs. Fluconazole was used as a positive control to verify the validity of this co-culture assay. Five natural molecules exhibited antifungal activity against both susceptible and resistant C. albicans. Two of these compounds, retigeric acid B (RAB) and riccardin D (RD), preferentially inhibited C. albicans strains in which the efflux pump MDR1 was activated. This selectivity was attributed to greater intracellular accumulation of the drugs in the resistant strains. Changes in sterol and lipid compositions were observed in the resistant strains compared to the susceptible strain, and might increase cell permeability to RAB and RD. In addition, RAB and RD interfered with the sterol pathway, further aggregating the decrease in ergosterol in the sterol synthesis pathway in the MDR1-activated strains. Our findings here provide an alternative for combating resistant pathogenic fungi.
ATP-Binding Cassette Transporters ; genetics ; metabolism ; Antifungal Agents ; chemistry ; metabolism ; pharmacology ; Azoles ; pharmacology ; Biosynthetic Pathways ; drug effects ; genetics ; Candida albicans ; chemistry ; drug effects ; metabolism ; Cell Membrane ; chemistry ; metabolism ; Coculture Techniques ; Drug Resistance, Fungal ; drug effects ; Ergosterol ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Lipids ; chemistry ; Molecular Structure ; Permeability ; Phenyl Ethers ; chemistry ; metabolism ; pharmacology ; Sterols ; chemistry ; metabolism ; Stilbenes ; chemistry ; metabolism ; pharmacology ; Triterpenes ; chemistry ; metabolism ; pharmacology

This study is to investigate the effect of the C21 sterols on inducing apoptosis of hepatocellular cancer cells and its potential mechanism. The transplanted model of hepatoma substantiality (Heps) was established in mice, and the mice were divided into four groups: negative controls group and C21 sterols groups (10, 20, 40 mg x kg(-1)) , treated with drugs separately once a day for 9 days. Then the mice were sacrificed, the tumor growth inhibition rate (IR) was calculated and tumor tissue samples were taken and examined under electron microscope. The tumor cells were harvested and cell viability or apoptosis was analyzed by acridine orange and ethidium bromide (AO/EB) stain. B-cell lymphoma/leukemia-2 gene (bcl-2) in tumor cells was inspected by immunohistochemistry. After treatment with C21 sterols (10, 20, 40 mg x kg(-1)), inhibitory effect on the transplanted Heps was observed. The IR was 34.79%, 47.08% and 50.23%, respectively. Apoptosis induced by the C21 sterols was observed, low growth density and some apoptotic cells were observed in tumor under the electron microscope. The expression of bcl-2 gene on tumor cells decreased in the C21 sterols groups, but the percentage of positive area is higher in 40 mg x kg(-1) group than that in 20 mg x kg(-1) group, which differed from apoptosis results. Inhibiting the excessive expression of bcl-2 gene to promote apoptosis may be one of anti-tumor mechanisms for the C21 sterols in Baishouwu.
Animals ; Antineoplastic Agents, Phytogenic ; isolation & purification ; pharmacology ; Apoptosis ; drug effects ; Cynanchum ; chemistry ; Female ; Gene Expression Regulation, Neoplastic ; Genes, bcl-2 ; Liver Neoplasms, Experimental ; metabolism ; pathology ; ultrastructure ; Male ; Mice ; Mice, Inbred ICR ; Neoplasm Transplantation ; Plants, Medicinal ; chemistry ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Random Allocation ; Sterols ; isolation & purification ; pharmacology ; Tumor Burden ; drug effects

Animals ; Antineoplastic Agents, Phytogenic ; isolation & purification ; pharmacology ; Cholesterol ; biosynthesis ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; HIV Protease ; metabolism ; HIV Protease Inhibitors ; pharmacology ; Humans ; Plants, Medicinal ; chemistry ; Platelet Aggregation ; drug effects ; Polysaccharides ; isolation & purification ; pharmacology ; Reishi ; chemistry ; Sterols ; isolation & purification ; pharmacology ; Triterpenes ; isolation & purification ; pharmacology