1-Methoxy-agroclavine from Penicillium sp. WC75209, a novel inhibitor of the Lck tyrosine kinase.

A high-throughput screen was developed and implemented to identify inhibitors of the Lck tyrosine kinase. This report describes the identification of a specific inhibitor of this enzyme from the solid fermentation culture of the Penicillium sp., WC75209. The active compound was isolated and structurally characterized as 1-methoxy-5R, 10S-agroclavine, a new member of the ergot alkaloid family.

Bripiodionen, a new inhibitor of human cytomegalovirus protease from Streptomyces sp. WC76599.

Bripiodionen (1), a new natural product, was isolated from Streptomyces sp. WC76599 during the screening of microbial fermentation extracts for their ability to inhibit human cytomegalovirus protease. The structure of 1 was elucidated by spectroscopic methods. Compound 1 displayed inhibitory activity against human cytomegalovirus protease with an IC50 value of 30 microM.

BMS-192548, a tetracyclic binding inhibitor of neuropeptide Y receptors, from Aspergillus niger WB2346. II. Physico-chemical properties and structural characterization.

The structure of BMS-192548, a tetracyclic binding inhibitor of neuropeptide Y receptors, was established by spectroscopic methods. The compound has an unusual B-C-D ring beta-diketone moiety.

BMS-192548, a tetracyclic binding inhibitor of neuropeptide Y receptors, from Aspergillus niger WB2346. I. Taxonomy, fermentation, isolation and biological activity.

During the screening of microbial fermentation extracts for their ability to inhibit the binding of 125I-peptid YY (PYY) to the neuropeptide Y (NPY) receptor using the scintillation proximity assay (SPA), BMS-192548 was isolated from the extract of Aspergillus niger WB2346 by bioassay-guided fractionation. BMS-192548 showed the inhibitory activity against 125I-PYY binding to SK-N-MC and SMS-KAN cells, which express NPY1 and NPY2 receptors, respectively, with IC50 values of 24 microM in Y1 and 27 microM in Y2 receptor binding. BMS-192548 demonstrated weak cytotoxicity against murine tumor cell line M-109 with an IC50 value of 240 microM.

Manumycins E, F and G, new members of manumycin class antibiotics, from Streptomyces sp.

Three new manumycin class antibiotics, namely manumycins E, F and G, were isolated from the culture broth of Streptomyces sp. strain WB-8376. Their structures were established by spectroscopic methods, and the S configuration of C-4 in the epoxycyclohexenone moiety was determined by CD exciton chirality method for each of the three compounds. Manumycins E, F and G are active against Gram-positive bacteria, and have moderate inhibitory effects on the farnesylation of p21 ras protein. They demonstrated weak cytotoxic activity against human colon tumor cell HCT-116.

Metabolism of levamisole, an anti-colon cancer drug, by human intestinal bacteria.

1. Anaerobic incubation of levamisole with human intestinal flora resulted in the formation of three thiazole ring-opened metabolites, namely, levametabol-I, II and III. These new hydroxamic lactam-type metabolites were isolated and characterized by various spectroscopic methods. 2. Various pure cultures of human intestinal bacterial strains were shown, by quantitative h.p.l.c. analysis, to have ring-opening ability. Strong metabolizers include Bacteroides and Clostridium spp. Bacterial mixtures prepared from human faeces showed much greater ability to transform levamisole (74% in 48 h) than any pure strain culture. 3. Greatly decreased levamisole-transforming activity was observed with autoclaved bacterial cultures, and no activity was found with broth medium alone. This indicates that metabolism requires the presence of anaerobic bacteria and involves, at least in part, a non-enzymic process.

Metabolism of 1,4-dinitro-2-methylpyrrole, a mutagen formed by a sorbic acid-nitrite reaction, by intestinal bacteria.

1,4-Dinitro-2-methylpyrrole (DNMP), a mutagenic product formed by the interaction of two common food additives, sorbic acid and sodium nitrite, was transformed to 1-nitro-2-methyl-4-aminopyrrole (NMAP) by human fecal mixtures and various intestinal bacterial strains. Under anaerobic conditions the cell suspensions of Actinomyces, Bacteroides, Clostridium, Eubacterium, Fusobacterium, and Peptostreptococcus spp. demonstrated the nitroreduction activity. Under aerobic conditions, only Actinomyces and Bacteroides spp. showed activity, and this was at a decreased level. In cell suspensions of Bacteroides thetaiotaomicron VPI 5482, NAD(P)H and glucose accelerated the reduction rate, whereas dicoumarol and heat significantly inhibited the rate, and flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) did not affect the rate. With cell-free preparations of the same strain, reduction required NAD(P)H as a cofactor in a dose-dependent fashion and was inactivated by air and heat.

Cis-flupentixol metabolites in rat plasma and bile. The first proof of glutathione conjugation at the exocyclic double bond.

The plasma and biliary metabolites of cis-flupentixol (cis-FPT) were studied after ip administration to rats. Cis-FPT sulfoxide was found to be the major phase-I metabolite in plasma and bile. Five biliary metabolites were isolated by gradient elution HPLC and characterized by various spectroscopic methods: F1, 1'-S-glutathionyl-10,1'-dihydroFPT sulfoxide; F2, cis-FPT sulfoxide sulfate; F3, 8-O-(or 7-O) glucuronyl-cis-FPT; F4, cis-FPT sulfoxide; and F5, cis-FPT glucuronide. A novel non-enzymatic addition of glutathione (GSH) onto the exocyclic double bond was demonstrated to occur for the first time with not only flupentixol and certain of its metabolites, but also with other psychotropic drugs with a tricyclic nucleus and an exocyclic double bond. Specifically, these nonenzymatic additions of GSH were observed with cis-FPT, trans-FPT, cis-FPT sulfoxide, cis-, trans-dealkylFPT, cis-FPT N-oxide, cis-chlorprothixene, cis-thiothixene, and cyclobenzaprine. Among them, cis-FPT sulfoxide showed the most potent adduct formation activity, and the product was characterized to be identical with the in vivo metabolite F1 of cis-FPT.

The identification of urinary metabolites of doxepin in patients.

The metabolism of doxepin was investigated in three patients who provided cumulative urine samples on each of 3 successive days. These samples were examined by means of stereoselective HPLC or HPLC combined with mass spectrometry through a plasmaspray interface (LCMS). In addition, sufficient quantities of the major metabolites were isolated from the urine by column chromatography. The isolated metabolites were examined by HPLC, proton nuclear magnetic resonance spectroscopy (1H-NMR), and chemical ionization and/or electron impact mass spectrometry (EIMS). The resultant spectra and chromatographic properties were compared with authentic reference standards. The metabolites were identified as (E)-2-hydroxydoxepin, (E)-2-hydroxy-N-desmethyldoxepin, (Z)- and (E)-N-desmethyldoxepin, and (Z)- and (E)-doxepin N-oxide. There was no evidence of hydroxylation at the oxymethylene bridge. A further metabolite previously unreported was tentatively identified by LCMS and EIMS as an aromatic hydroxy-N-desmethyldoxepin hydrated at the exocyclic double bond. This metabolite was present in very low amounts, precluding its analysis by 1H-NMR. Moreover, this type of compound dehydrated readily in vitro, and numerous attempts to synthesize reference materials were unsuccessful. The tentative identification of this hydrated metabolite lends significant support to a possible mechanism responsible for the enrichment of cis-N-desmethyldoxepin over time in plasma.

Chronotropic Effect of the Methanolic Extracts of the Plants of the Paris Species and Steroidal Glycosides Isolated from P. vietnamensis on Spontaneous Beating of Myocardial Cells1.

Rhizomes of five identified plants of the PARIS species, Liliaceae, and Rhizoma Paridis which are sold as a crude drug named "Zao Xiu", "Qiyeyizhihua" or other names in nine different markets in China were tested for their effects on cultured cardiomyocytes. In the standard medium, eight methanol extracts out of sixteen at a concentration of 0.2 mg/ml stopped the spontaneous beating of myocardial cell sheets, but these extracts significantly increased the beating rate when the concentration was reduced to one half. In the culture medium with a low calcium concentration, 0.5 mM, the beating rate of the cells decreased to about 60% of that of the control in the standard medium. The addition of five of the extracts to the low calcium medium at a concentration of 0.1 mg/ml caused a stop of cell beating, but the other extracts increased beating rate at least by 10%. Three steroidal glycosides isolated from the rhizomes of P. VIETNAMENSIS (Takht.) H. Li also stimulated cell beating. Among them, diosgenin-3- O-alpha- L-rhamnopyranosyl-(1-->2) - (alpha- L-arabinofuranosyl-(1-->4))- D-glucopranoside (compound 1) was the most effective stimulant for cell beating as well as calcium uptake by the myocardial cells.

Metabolism of swertiamarin from Swertia japonica by human intestinal bacteria.

The biotransformation of swertiamarin [1, a seco-iridoid glucoside isolated from Swertia japonica (Schult.) Makino] by human intestinal bacteria was investigated. Three metabolites were isolated and identified as erythrocentaurin (2), 5-hydroxymethylisochroman-1-one (3), and gentianine (4) by spectroscopic methods. Through screening of various defined strains of intestinal bacteria (25 species), it was found that all these species had the ability to metabolize 1 to 2 and 3, whereas only a few species had the ability to produce 4. This is the first report to show that one of the metabolic intermediates of the secoiridoid compound is further transformed to a nitrogen-containing compound through metabolic processes by human intestinal bacteria.

Metabolism of homoorientin by human intestinal bacteria.

As a part of our studies on the metabolism of bioactive compounds from oriental medicines by intestinal flora, homoorientin, a C-glycosylflavonoid, was anaerobically incubated with a human intestinal bacterial mixture. Homoorientin was transformed to 6-C-glucosyleriodictyol, (+/-)-eriodictyol, luteolin, 3,4-dihydroxyphenylpropionic acid, and phloroglucinol. A novel cleavage of the C-glycosyl bond was discovered for the first time by using intestinal bacteria.