Isolation of inhibitors of adenylate cyclase from dan-shen, the root of Salvia miltiorrhiza

The effect of dan-shen extract, the root of Salvia miltiorrhiza, on adenylate cyclase was investigated in both rat brain and rat erythrocytes. The EtOAc fraction of the MeOH extract was proved to have significant inhibitory activity. Potent inhibitory principles in the EtOAc fraction were isolated and identified as 4 polyphenolic acids, rosemarinic acid, lithospermic acid, and their methyl ester derivatives.     

Screening for molluscicidal activity in crude drugs

Thirty-four extracts of crude drugs and medicinal plants have been screened for activity against Oncomelania nosophora, the intermediate host of the Japanese strain of Schistosoma japonicum. Strong molluscicidal activity was found in the MeOH extract of Anemarrhenae Rhizoma. Although timosaponin A-III, one of the main saponins of Anemarrhenae Rhizoma, showed very strong killing activity, markogenin-3-O-beta-D-glucopyranosyl (1-2)-O-beta-D-galactopyranoside having the same glycosidic linkage was found to be inactive.     

Preparation of deuterated methyl and dimethyl substituted nicotinoylating agents for derivatization of the N-terminal of protein

Methyl groups of 6-methylnicotinic acid and 2,6-dimethylnicotinic acid were deuterated by an H-D exchange reaction under conditions of 1% NaOD/D(2)O on heating. With a condensation reaction between the D-labeled nicotinic acid derivative and N-hydroxysuccinimide with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride, the nicotinoylating agents, 1-(6-methyl[D(3)]nicotinoyloxy)succinimide (2c) and 1-(2,6-dimethyl[D(6)]nicotinoyloxy)succinimide (2f) were prepared. Both D-labeled nicotinoylating agents and their unlabeled counterparts quantitatively modified the N-terminal of protein.     

Chemical derivatization of peptides containing phosphorylated serine/threonine for efficient ionization and quantification in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

We describe a useful method for the efficient ionization and relative quantification of peptides containing serine/threonine phosphorylation sites. This method is based on beta-elimination of the phosphate group from serine/threonine phosphorylation sites under alkaline conditions, followed by Michael addition reaction with N-(2-mercaptoethyl)-6-methylnicotinamide (MEMN). As a result of the derivatization reaction, the negatively charged phosphate group is substituted with the nicotinoyl moiety to improve the ionization efficiency of the derivatized peptide. The combination of d(3)-labeled MEMN (d(3)-MEMN) and MEMN (d(0)-MEMN) generates a 3 Da mass difference between d(3)-MEMN-labeled and d(0)-MEMN-labeled peptides, which is a useful signature for the identification of peptides containing serine/threonine phosphorylation sites in the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrum. Moreover, the mass difference is useful for the quantitative analysis of serine/threonine phosphorylation in proteins. In this paper, we describe the synthesis of d(0)/d(3)-labeled MEMN and an application of our approach to model peptides and proteins.     

Efficient identification and quantification of proteins using isotope-coded 1-(6-methylnicotinoyloxy)succinimides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

We describe a convenient and useful method for the identification and relative quantification of proteins using light and heavy reagents, 1-(6-methylnicotinoyloxy)succinimides (6-CH(3)-Nic-NHS and 6-CD(3)-Nic-NHS, respectively). This method is based on the chemical derivatization of amino groups of tryptic peptides with these reagents, i.e., the basic moiety of the reagents thus incorporated into both the N-terminal amino group and the epsilon-amino group of the lysine residue would improve the ionization efficiency of tryptic peptides. An increase in protein sequence coverage is achieved by derivatization with these reagents or by combination of mass values before and after derivatization. Since a combination of 6-CH(3)-Nic-NHS and d(3)-labeled reagent (6-CD(3)-Nic-NHS) generates a 3 Da mass difference per reaction site, the d(3)-labeled reagent shifts the mass values of d(0)-labeled peptides according to the number of reactive amino groups in the peptides. In the case of tryptic peptides, the mass values of C-terminal arginine and lysine peptides are shifted by 3 and 6 Da, respectively. Further, the 3 Da mass difference between 6-CH(3)-Nic-NHS and 6-CD(3)-Nic-NHS offers a means for the relative quantification of protein by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.     

Inhibitory effect of 2-(E-2-alkenoylamino)ethyl alkyl sulfides on gastric ulceration in rats. II. Structure and activity relationships of 2-(E-n or Z-n-Decenoylamino)ethyl alkyl sulfides

The analogues of 2-(E-n or Z-n-decenoylamino)ethyl carbamoylmethyl sulfide, including the modifications of sulfide portion, double bond in decenoyl chain and alkyl sulfide moiety, were synthesized and their inhibitory effects on stress-induced ulceration in rats were compared. Replacing the sulfura atom by methylene group or oxygen atom reduced the effect of potency. Saturation of the double bond in the decenoyl chain tended to reduce the anti-ulcerogenic activity in rats. There was no relationship between the position of double bond in decenoyl chain and the pharmacological activity. On the other hand, compounds with E-configuration showed stronger anti-ulcer activity than the corresponding Z-type of compounds. Among 9 kinds of S substituted alkyl groups for carbamoylmethyl, 2-(E-2-decenoylamino)ethyl 2-cyclohexylethyl sulfide showed the most potent anti-ulcerogenic activity in rats and also showed the lowest acute toxicity in mice.     

Enhanced cytotoxicity in simultaneous and sequential drug-heat treatments of cultured Chinese hamster V79 cells

Under hyperthermal conditions, some genotoxic drugs such as bleomycin, paraquat, and N-alkyl-N-nitrosoureas exhibit increased cytotoxicity toward cultured Chinese hamster V79 cells. Sequential combinations of heat and drug treatments, regardless of whether drug-exposure precedes or follows hyperthermia, also induce synergistic cytotoxicity to some extent. This may be attributed not only to the relationship of temperature and chemical injury as defined by the Arrhenius law, but also to a lethal interaction between the biological consequences of chemical injury and thermal damage. Ethanol, dimethylsulfoxide (DMSO), ethylenediaminetetraacetic acid (EDTA), and urea, which are known to affect cell membrane and protein, also exert synergistic cytotoxicity at 43 degrees C at a dose range that is nontoxic at 37 degrees C. When used sequentially with thermal treatment, they also proved to be synergistic. Glycerol, however, protected cells against thermal damage when used in a simultaneous chemical-thermal combination. But when treatments were carried out sequentially, glycerol proved destructive.     

Product analyses of ozone mediated nitration of benzimidazole derivatives with nitrogen dioxide: formation of 1-nitrobenzimidazoles and conversion to benzotriazoles

Several benzimidazole derivatives having electron-withdrawing or -donating substituent(s) at the benzene moiety were used as models of the imidazole moiety of purine bases and their nitration with nitrogen dioxide and ozone (so-called Kyodai nitration) were examined. Products were extracted from the reaction mixture with AcOEt and their structures were analyzed. 1-Nitrobenzimidazole derivatives and unexpected 1-nitrobenzotriazole derivatives were identified. Although the yields of 1-nitrobenzimidazole derivatives were quite low, these were all new compounds that could be obtained only by Kyodai nitration. It was speculated that benzotriazoles were formed via 1-nitrobenzimidazoles and subsequent nitration toward benzotriazoles resulted in the formation of 1-nitrobenzotriazoles.