Antiprotozoal activity of bicycles featuring a dimethylamino group at their bridgehead

Several dimethylamino-derivatives of the new compound-class 3-azabicyclo[3.2.2]nonanes were prepared. For better comparison of activity also a few analogues of bicyclo[2.2.2]octanes and 2-azabicyclo[3.2.2]nonanes were synthesized. Their activities were examined in vitro against the multiresistant K₁ strain of Plasmodium falciparum and against Trypanosoma brucei rhodesiense (STIB 900). A couple of the newly synthesized compounds showed promising antiprotozoal activity and selectivity. The results of the biological tests of the novel compounds were compared with the activities of already synthesized compounds and of drugs in use. Structure–activity relationships were discussed.     

Determination of a new proton pump inhibitor, YH1885, in human plasma and urine by high-performance liquid chromatography

A high-performance liquid chromatographic method was developed for the determination of a new proton pump inhibitor, YH1885 (I), in human plasma and urine, and rat blood and tissue homogenate using fenticonazole as an internal standard. The sample preparation was simple: a 2.5 volume of acetonitrile was added to the biological sample to deproteinize it. A 50-μl aliquot of the supernatant was injected onto a C₈ reversed-phase column. The mobile phase employed was methanol-0.005 M tetrabutylammonium dihydrogenphosphate (77:23, v/v), and it was run at a flow-rate of 1.0 ml/min. The column effluent was monitored using an ultraviolet detector at 270 nm. The retention times for I and the internal standard were 9.0 and 10.3 min, respectively. The detection limits for I in human plasma and urine, and in rat tissue homogenate (including blood) were 50, 100 and 100 ng/ml, respectively. The coefficients of variation of the assay (within-day and between-day) were generally low (below 8.84%) for human plasma and urine, and for rat tissue homogenate. No interferences from endogenous substances were found.     

产γ_内酰胺水解酶菌株的筛选及发酵条件研究

(-)γ-内酰胺是合成两种抗艾滋病药物(-)carbovir和(-)abacavir的重要原料,具有重要的应用价值,微生物酶法拆分( /-)γ-内酰胺生产(-)γ-内酰胺的方法具有良好的应用前景。以N-乙酰苯丙氨酸为唯一碳源,从土壤中筛选得到了69株具有酰胺水解酶活性的菌株,利用液相色谱分析方法确定了其中20株有较高的酰胺水解酶活性,利用手性色谱分析的方法进一步得到了一株具有较高立体选择性,能拆分( /-)γ-内酰胺而获得(-)γ-内酰胺的菌株L29-9,对该菌株的产酶培养基的碳、氮源及初始pH值等进行了研究。结果表明:当选择碳源柠檬酸2g/L、氮源酵母提取物5g/Lp、H 7.0、培养时间40h时,通过完整细胞转化,在30℃下经过12h的反应,产物(-)γ-内酰胺产率达40%,ee值99.5%。     

朗德鹅胆汁的化学成分及金属蛋白酶抑制活性研究

应用多种色谱技术进行分离纯化,从朗德鹅胆汁85％乙醇提取物中分离得到6个化合物。经理化性质和光谱数据分析鉴定为苯乙酸（1）、鹅去氧胆酸（2）、鹅去氧胆酸乙酯（3）、棕榈酸-α-单甘油酯（4）、顺-6-十八碳烯酸（5）、（4E）-2-[2＇-hydroxyhexadecanoylamino]-4-octadecane-1,3-diol（6）。化合物1、3、4和6为首次从该属动物胆汁中分得,其中化合物6为首次从陆生动物胆汁中分得的一种神经酰胺类成分。首次对化合物2、4和5进行抑制金属蛋白酶活性的实验,评价了三个化合物的生物活性。     

Isolation and identification of a humanTRPV1 activating compound from soy sauce

Transient receptor potential vanilloid 1 (TRPV1) was identified as a receptor of capsaicin, which is a pungent ingredient in hot red peppers. Due to its relevance for nociception, a physiological and pharmacological study of TRPV1 has also been developed. Therefore, it is important to enrich scientific knowledge regarding the TRPV1 activating or inhibiting compounds. In this study, we fractionated soy sauce based on the human TRPV1 (hTRPV1) activity using column chromatography and purified 5-(9H-pyrido[3,4-b]indol-1-yl)-2-furanmethanol (perlolyrine) as an hTRPV1-activating compound. Additionally, perlolyrine activates the human transient receptor potential ankyrin 1 (hTRPA1). The EC₅₀ of hTRPV1 and hTRPA1 were 2.87 and 1.67 μmol L^?1, respectively. HPLC quantification of soy sauces showed that they contain 2.22–12.13 μmol L^?1 of perlolyrine. The sensory evaluation revealed that perlolyrine has taste modification effect. The results of this study, for the first time, suggest that perlolyrine induces the activation of hTRPV1 and hTRPA1.     

Involvement of 5-HT receptors in the development and expression of methamphetamine-induced behavioral sensitization: 5-HT receptor channel and binding study

Methamphetamine (MAP) is one of the most commonly abused drugs in Asia, and previous studies suggest that serotonin 3 receptors (5-HT(3)) are involved in MAP-induced locomotion and reward. However, little is known about the role of 5-HT(3) receptors in MAP-induced behavioral sensitization. Here, we measured the effects of MDL 72222, a 5-HT(3) antagonist, and SR 57227 A, a 5-HT(3) agonist, on the development and expression of MAP-induced behavioral sensitization, and alternations of 5-HT(3) receptor binding labeled with the 5-HT(3)-selective antagonist, [(3)H]GR65630, in mice. In addition, we investigated the effects of MAP on 5-HT(3A) receptor channel activity in Xenopus laevis oocytes expressing 5-HT(3A) receptors. We found that MDL 72222 attenuated both the development and expression of behavioral sensitization to MAP (1.0 mg/kg, i.p.), and that this attenuating effect of MDL 72222 was reversed by pre-treatment with SR 57227 A. In oocytes expressing 5-HT(3A) receptor, MAP exhibited a dual modulation of 5-HT(3A) receptor channel activity, i.e. pre-treatment with a low dose of MAP (0.1 microm) enhanced 5-HT-induced inward peak current (I(5-HT)) but a high dose of MAP (100 microm) inhibited I(5-HT). The acute administration of MDL 72222 with MAP decreased [(3)H]GR65630 binding versus MAP alone in the mouse striatum. Our results suggest that MDL 72222 attenuates MAP-induced behavioral sensitization via 5-HT(3) receptors in the caudate putamen, and that 5-HT(3) receptor antagonists like MDL 72222 have potential as novel anti-psychotic agents for the treatment of MAP dependence and psychosis.     

Contribution to the determination of the chemotherapeutic drug G1 in biological fluids

A sensitive gas chromatographic method for the quantitative determination of the new antibacterial and antifungal drug G1, 1-(5-bromofuran-2-yl)-2-bromo-2-nitroethene, has been optimized. The method involves a fast and single extraction step from spiked serum and urine samples. The G1 drug was quantified using an internal standard method and by means of a nitrogen-selective detector. The results are statistically significant and show that mean levels of G1 as low as 1 μg ml⁻¹ can be measured accurately.     

Determination of 1-[5-(2-cyclopropyl-5,7-dimethyl-imidazo[4,5-b]pyridin-3-ylmethyl)thiophen-2-yl]cyclopent-3-enecarboxylic acid (CP-191,166), an angiotensin II antagonist, in dog and rat plasma by high-performance liquid chromatography

A simple and precise high-performance liquid chromatographic (HPLC) assay was developed and validated for the determination of a novel angiotensin II antagonist, 1-[5-(2-cyclopropyl-5,7-dimethyl-imidazo[4,5-b]pyridin-3-ylmethyl)thiopen-2-yl)cyclopent-3-enecarboxylic acid (CP-191,166, I), in dog and rat plasma. The internal standard (II, a saturated derivative of I) and analyte were extracted by liquid-liquid extraction using methyl tert.-butyl ether. Samples were analyzed by reversed-phase HPLC using a Zorbax C₈ narrow-bore column with ultraviolet detection at 289 nm. The quantitation limit of I was 10 ng/ml and the calibration curve was linear over the range of 0.01–10.0 μg/ml (r²>0.99). In dog and rat plasma, intra- and inter-assay precision ranged from 0.00 to 3.36% and 0.00 to 4.95%, respectively. The average recoveries were similar (73%) for both I and II and the upper limit of quantification of I can be as high as 500 μg/ml. The method described has been successfully applied to the quantification of I in about 2000 dog and rat plasma samples over a nine-month period.     

A Pitfall of the 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethonyphenol)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) Assay due to Evaporation in wells on the Edge of a 96 well Plate

The 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethonyphenol)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) calorimetric assay is replacing the traditional 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as a fast, one-step assay of cell viability. We have observed that evaporation of the outer wells of a 96 well plate increases the absorbancy by 52% compared to the inner wells. Filling the outer 2 rows of wells with media and replacement of the media prior to addition of the MTS reagent will, however, correct this inaccuracy.     

Participation of peroxisomal beta-oxidation system in the chain-shortening of a xenobiotic acyl compound

A drug, (E)-3-[4-(1-imidazolylmethyl)phenyl]-2-propenoic acid, was metabolized to 4-(1-imidazolylmethyl)benzoic acid in isolated hepatocytes of rats, which was enhanced markedly by the pretreatment of rats with clofibrate. With liver homogenates, the formation of the CoA-ester of this drug and its subsequent chain-shortening were demonstrated. In the series of these reactions, acyl-CoA synthetase, CoA, ATP and NAD were required, whereas cyanide did not inhibit the reaction. These results indicate that peroxisomes are capable of shortening the acyl side-chains of drugs by the beta-oxidation, giving an additional suggestion on the functions of peroxisomes.     

Theoretical research in structure characteristics of different inhibitors and differences of binding modes with CBP bromodomain

The CBP (CREB (cAMP responsive element binding protein) binding protein) bromodomain (BRD) could recognize and bind with acetyl K382 of human tumor suppressor protein p53 which the mutation of encoding gene might cause human cancers. CBP-BRD serves as a promising drug target for several disease pathways and a series of effective drug have been discovered. In this study, molecular dynamics (MD) simulations and molecular mechanics generalized born surface area (MM-GB/SA) approaches were performed to investigate the different binding modes between five inhibitors with CBP-BRD. Based on the energy and conformation analyses, a potent core fragment is chosen to act as the starting point for new inhibitor design by means of LUDI and rational drug design approaches. Then, T.E.S.T and molinspirition were applied to evaluate oral bioavailability and drug promiscuity of the new molecules. These results shed light on the idea for further inhibitor design.     

Deacylation of carcinogenic 5-nitrofuran derivatives by mammalian tissues

The deacylations of N-[4-(5-nitro-2-furyl)-2-thiazolyl] fonnamide (FANFT), N-[4-(5-nitro-2-furyl)-2-thiazolyl] acetarnide (NFTA) and formic acid 2-[4-(5-nitro-2-furyl)-2-thiazolyl] hydrazide (FNT) by liver, kidney, small intestines and stomach of mouse, rat, hamster and guinea pig were investigated. FANFT was deformylated to 2-amino-4-(5-nitro-2-furyl)thiazole (ANFT). FANFT formamidase activity was higher in the liver and small intestines of mouse, hamster and guinea pig, and small intestines and stomach of rat. There was no detectable FANFT formamidase activity in the stomach of the mouse and hamster. Neither NFTA nor FNT was deacylated by the rodent tissue homogenates studied. It is suggested that (1)4 ANFT is a metabolite of FANFT but not NFTA; (2) 2-hydrazino-4-(5-nitro-2-furyl)thiazole (HNFT) may not be a metabolite of FNT; and (3) the induction of tumors by FANFT, NFTA and FNT may not be due to a common carcinogenic metabolite, although these chemicals demonstrate similar organ specificities in some of these rodents.     

Preparative Biorganic Chemistry Part 10 Asymmetric Reduction of Bicyclo[2.2.2]Octane-2,6-Diones with Baker's Yeast

Reduction of bicyclo[2.2.2]octane-2,6-dione with baker's yeast gave (lR,4S,6S)-6-hydroxybi-cyclo[2.2.2]octan-2-one (95% e.e.) contaminated with 8% of its (1S,4R,6S)-isomer. Similarly, the yeast reduction of 1-methylbicyclo[2.2.2]octane-2,6-dione furnished (1R,4S,6S)-6-hydroxy-1-methylbi-cyclo[2.2.2.]octan-2-one (99.5% e.e.) in 59% yield. The yeast reduction of 4-methylbi-cyclo[2.2.2.]octane-2,6-dione afforded (1R,4S,6S)-6-hydroxy-4-methylbicyclo[2.2.2.]octan-2-one (98% e.e.) contaminated with 3% of its (1S,4R,6S)-isomer in 58% yield.     

一种改进的小鼠局灶性脑缺血神经症状定量评价方法

本文旨在建立一种客观评价小鼠局灶性脑缺血神经症状的定量方法。在大脑中动脉阻塞诱导局灶性脑缺血后24h,采用悬挂试验分别测定转动的平均角和优势角以及转动次数,并用爬板试验测定小鼠攀爬角度;分析定量测定指标与脑梗死体积、神经元密度的相关性,并与经典的行为学评价方法比较。还以此法观察抗脑缺血药{pranlukast,4-氧-8-[对-(4-苯丁氧基)苯甲酰氨基]-2-(5-四氮基)-4H-1-苯并吡喃半水化合物}(ONO-1078)的作用。结果显示,脑缺血小鼠各项指标均有显著改变,定量评价总分与脑梗死体积、神经元密度减少密切相关,与经典行为学评分之间也密切相关。ONO-1078可显著抑制缺血损伤,并降低定量评价总分。因此,本方法可反映局灶性脑缺血神经症状变化,具有客观、定量、操作简便、无损伤的优点,并能用于评价抗脑缺血药物的作用。     

Allosteric interactions among drug binding sites on calmodulin

Felodipine is a fluorescent dihydropyridine Ca²⁺-antagonist. It binds to calmodulin in a Ca²⁺-dependent manner, and undergoes a fluorescence increase which allows us to monitor its interaction with calmodulin. Hydrophobic ligands including the calmodulin antagonist, R24571 and Ca²⁺ antagonists, prenylamine and diltiazem, bind to calmodulin and potentiate felodipine binding by as much as 20 fold. These studies suggest that allosteric interactions occur among different drug binding sites on calmodulin. Our results are discussed in terms of the mechanism of action of calmodulin.     

Lilac alcohol epoxide: a linalool derivative in Actinidia arguta flowers

Lilac alcohol epoxide (2-(5-methyl-5-(oxiran-2-yl)-tetrahydrofuran-2-yl)propan-1-ol), a previously unreported monoterpene, was identified in the solvent extract of the flowers of seven Actinidia arguta genotypes. The diastereomeric lilac alcohol epoxides co-occurred with the lilac aldehydes and alcohols. Another compound, the lilac diol (2-(5-(1-hydroxyethyl)-5-methyl-tetrahydrofuran-2-yl)propan-1-ol) was synthesised as part of our efforts to identify the lilac alcohol epoxide.     

Photoaffinity Labeling of Adenylate Cyclase-Linked Serotonin Receptors in Aplysia Neurons

Serotonin stimulated adenylate cyclase in Aplysia neurons with a Kact of 0.7 microM. Under the same conditions, 1-[2-(4-aminophenyl)ethyl]4-(3-trifluoromethylphenyl)piperazine stimulated adenylate cyclase with a Kact of 20 microM. The azido derivative of this compound, 1-[2-(4-azidophenyl)ethyl]4-(3-trifluoromethylphenyl)piperazine, or of serotonin, (4-amino, 3-nitrophenylazido-serotonin), also stimulated the cyclase in the dark, but with lower efficiency (Kact greater than 10(-4) M). Irradiation of the membranes in the presence of 100 microM 1-[2-(4-azidophenyl)ethyl]4-(3-trifluoromethylphenyl)piperazine abolished 75% of the cyclase activity stimulated by 5 microM serotonin. Under the same conditions, 100 microM 4-amino, 3-nitrophenylazido-serotonin did not inhibit serotonin-stimulated adenylate cyclase activity. When [3H]1-[2-(4-azidophenyl)ethyl]4-(3-trifluoromethylphenyl)piperazine (20 microM) was irradiated with membranes for 5 min at 4 degrees C, a dozen peptides were labeled, as revealed by a fluorogram of sodium dodecyl sulfate-polyacrylamide gels. Among them, the labeling of five polypeptides (molecular weights of 45,000, 55,000, 63,000, 80,000, and 94,000) was protected by the presence of 0.2 mM serotonin during photolysis. These peptides may be related to serotonin receptors.     

Analysis of a novel antiinflammatory agent, 1-(7-tert.-butyl-2,3-dihydro-3,3-dimethylbenzo[b]furan-5-yl)-4-cyclopropylbutan-1-one (PGV-20229), in plasma matrices by stable-isotope-dilution gas chromatography-mass spectrometry

A sensitive and selective GC-MS method was developed for the determination of low levels of a novel antiinflammatory agent, 1-(7-tert.-butyl-2,3-dihydro-3,3-dimethylbenzo[b]furan-5-yl)-4-cyclopropylbutan-1-one (I), in small volumes of animal plasma. The method involved the addition of ¹³C₆-labeled-I to plasma samples, followed by a simple liquid-liquid extraction with hexane to isolate the analytes from matrix components. The levels of I in the sample extracts were determined by isotope-dilution GC-MS analysis using selected-ion monitoring. The method was linear over three orders of magnitude, with a limit of quantitation of 1.8 ng/ml I, using plasma sample volumes of 0.1 ml. The method was utilized to determine the pharmacokinetic parameters of I in rats and dogs, following intravenous administration.     

Antifungal antibiotics and Siba inhibit 1-aminocyclopropane-1-carboxylic acid synthase activity

The antifungal antibiotics Sinefungin and A9145C isolated from Streptomyces griseolus and the synthetic nucleoside Siba, which are analogs of S-adenosylmethionine, inhibit the activity of 1-aminocyclopropane 1-carboxylic acid synthase from tomato fruits. Sinefungin and Siba were shown to be more potent inhibitors than A9145C. In extracts of green fruits, the enzyme activity was inhibited by Sinefungin with an I50 of 1 microM, which was similar to that caused by aminoethoxyvinylglycine, and by Siba with an I50 of 100 microM; in extracts from red tomatoes, the I50's were 25 microM and 100 microM, respectively. The inhibition of ACC synthase by these analogs could be reversed by gel filtration chromatography.     

The effect of cross-linking spectrin-actin complexes with band 4.1 on the state of polymerization of the actin

The polymerization of actin in the presence of spectrin tetramers and band 4.1 isolated from the human erythrocyte has been measured using a fluorescence energy transfer technique. The results show that the cross-linking of spectrin-actin complexes by band 4.1 results in a limited depolymerization of actin filaments and a concomitant rise in the critical actin concentration. The phenomenon may explain in part the existence of actin in the erythrocyte cytoskeleton as short oligomers rather than as long filaments.