Synthesis of peptide inhibitors of human spleen fibrinolytic proteinase (SFP) and human leukocyte elastase-like proteinase (ELP)

Stereoisomers of specific substrates for SFP and ELP, Suc-L-Tyr-L-Leu-L-Val-pNA and Suc-L-Ala-L-Tyr-L-Leu-L-Val-pNA, were synthesized by the conventional solution method. Suc-L-Tyr-D-Leu-D-Val-pNA was found to be an effective and specific inhibitor of SFP and ELP. Suc-L-Tyr-L-Leu-L-Val-Pipe and Suc-L-Tyr-D-Leu-D-Val-Pipe inhibited only SFP very slightly and showed no inhibitory effect on ELP. Both Dan-L-Tyr-L-Leu-L-Val-pNA and Dan-L-Tyr-D-Leu-D-Val-pNA exhibited an inhibitory effect on both SFP and ELP.     

The effects of (±)-idazoxan and its stereoisomers on mouse vas deferens motility in vitro. A comparison with yohimbine

The effects of idazoxan (IDZ) and its stereoisomers were compared to that of a classical alpha 2-antagonist, yohimbine (YOH), on para-aminoclonidine (PAC)- and norepinephrine (NE)-mediated inhibition of the twitch response evoked in the mouse vas deferens by low-frequency (0.1 Hz) field stimulation. (+/-)-IDZ and (+)-IDZ antagonized the inhibitory effects of PAC, (+)-IDZ being twice as potent as (+/-)-IDZ; in contrast, antagonism by (-)-IDZ failed to meet Schild criteria for a competitive mechanism. YOH completely reversed the inhibition of twitch response induced by NE, but not that induced by PAC; in the latter case, residual inhibition was almost fully reversed by (+)-IDZ and to a lesser extent by (+/-)-IDZ, while (-)-IDZ proved ineffective. These results provide pharmacological evidence of alpha 2-receptor heterogeneity at the vas deferens level. A possible additional mechanism involving imidazoline binding sites is discussed.     

A new analytical method for stereoisomers of methamphetamine and amphetamine and its application to forensic toxicology

We have developed a rapid and sensitive high-performance liquid chromatographic method for the determination of low concentrations of the stereoisomers of methamphetamine (MAMP) and the demethylated metabolite, amphetamine (AMP). The acetyl derivatives of the stereoisomers (d and l-form) of MAMP and AMP smuggled by gangsters or extracted from biological specimens (hair) of drug addicts were clearly separated on two stereoisomer-analytical columns (Chiralcel OB and OJ, Daicel Ind. Co., Japan) connected in series at 50 degrees C. The mobile phase was a mixture of n-hexane and isopropanol (9:1, v/v) and the UV detector was set at 220 nm. The practical limit of sensitivity for the analysis was 62.5 ng of the stereoisomers of MAMP and AMP. Our analytical method for MAMP and AMP in human hair is very useful for the diagnosis of abuse of the drug. The determination of the d/l ratio of MAMP and AMP (the fingerprint of MAMP and AMP) is applicable to the investigation of smuggling routes of these drugs.     

Correlation between chemical structure and biological activities of Porphyromonas gingivalis synthetic lipopeptide derivatives

We recently separated a PG1828-encoded triacylated lipoprotein (Pg-LP), composed of two palmitoyl and one pentadecanoyl groups at the N-terminal of glycerocysteine from Porphyromonas gingivalis, a periodontopathic bacteria, and found that Pg-LP exhibited definite biological activities through Toll-like receptor (TLR) 2. In the present study, we synthesized 12 different Pg-LP N-terminal peptide moieties (PGTP) using four combinations of glyceryl (R and S) and cysteinyl (l and d) stereoisomers, and three different acyl group regioisomers, N-pentadecanoyl derivative (PGTP1), S-glycero 2-pentadecanoyl derivative (PGTP2) and S-glycero 3-pentadecanoyl derivative (PGTP3). All the PGTP compounds (RL, SL, SD, RD) tested showed TLR2-dependent cell activation. The activating capacities of the PGTP-R compounds were more potent than those of the PGTP-S compounds, whereas there were no differences between the PGTP-L and -D compounds. Furthermore, the production of interleukin (IL)-6 following stimulation with the PGTP1-RL, PGTP2-RL and PGTP3-RL compounds was impaired in peritoneal macrophages from TLR2 knock-out (KO), but not those from TLR1 KO or TLR6 KO mice. These results suggest that P. gingivalis triacylated lipopeptides are capable of activating host cells in a TLR2-dependent and TLR1-/TLR6-independent manner, and the fatty acid residue at the glycerol position in the PGTP molecule plays an important role in recognition by TLR2.     

One peptide,two topologies: structure and interconversion dynamics of human uroguanylin isomers

The peptide hormone uroguanylin stimulates chloride secretion via activation of intestinal guanylyl cyclase C (GC-C). It is characterized by two disulfide bonds in a 1-3/2-4 pattern that causes the existence of two topological stereoisomers of which only one induces intracellular cGMP elevation. To obtain an unambiguous structure-function relationship of the isomers, we determined the solution structure of the separated uroguanylin isoforms using NMR spectroscopy. Both isomers adopt well-defined structures that correspond to those of the isomers of the related peptide guanylin. Furthermore, the structure of the GC-C-activating uroguanylin isomer A closely resembles the structure of the agonistic Escherichia coli heat-stable enterotoxin. Compared with guanylin isomers, the conformational interconversion of uroguanylin isomers is retarded significantly. As judged from chromatography and NMR spectroscopy, both uroguanylin isoforms are stable at low temperatures, but are subject to a slow pH-dependent mutual isomerization at 37°C with an equilibrium isomer ratio of approximately 1:1, The conformational exchange is most likely under the sterical control of the carboxy-terminal leucine. These results imply that GC-C is activated by ligands exhibiting the molecular framework corresponding to the structure of uroguanylin isomer A.     

STEREOCHEMISTRY IN ANAESTHESIA

1. Interest in the pharmacokinetic and pharmacodynamic properties of the enantiomers of chiral drugs has greatly increased in recent years. This is particularly so for agents used in anaesthesia. 2. Chiral compounds are those that can exist in two non-superimposable forms. Each form is termed an enantiomer or stereoisomer. Two naming systems are in use: one uses the terms (+) and (–) to indicate the direction the compound will rotate polarized light, while the other system, based on the absolute three-dimensional structure of the enantiomers, uses the terms R and S. 3. Investigation of the stereoisomers of the volatile anaesthetic agent isoflurane is increasing our understanding of the mechanism of general anaesthesia. Current evidence suggests a protein, rather than a lipid, receptor site. 4. Investigation of the stereoisomers of local anaesthetics is increasing the safety of these drugs. 5. For bupivacaine, a widely used amide local anaesthetic, important enantiomeric differences can be found for toxicity, clinical effect and pharmacokinetics. In particular S-(–)-bupivacaine has an improved central nervous system and cardiac safety profile. This is partly explained by the pharmacokinetic differences. 6. Based on these differences, ropivacaine, a propyl homo-logue of bupivacaine, has been produced solely as the S-(–)-enantiomer. The available evidence suggests significantly improved safety for this agent over racemic bupivacaine.     

Chiral separation of racemate CPU86017, an anti-arrhythmic agent, produces stereoisomers possessing favourable ion channel blockade and less alpha-adrenoceptor antagonism

1. CPU86017 is an effective anti-arrhythmic agent of the Class III complex that has two chiral centres, 7N and 13aC. As a promising anti-arrhythmic agent, the blockade on I(Kr), I(Ks) and calcium influx may be modulated to be mild, moderate and potent, with less a-adrenoceptor blockade. In order to improve activity at ion channels, four stereoisomers, namely SS ((+)-7S,13aS-CPU86017), SR ((-)-7S,13aR-CPU86017), RR ((-)-7R,13aR-CPU86017) and RS ((+)-7R,13aS-CPU86017), have been separated. In the present study, the effects of these four isomers on I(Kr) and I(Ks), calcium channels and a-adrenoceptors were compared with the effects of the racemate CPU86017. 2. In the present study, I(Kr) and I(Ks) were measured as tail currents (I(Kr.tail) and I(Ks.tail), respectively) using the whole-cell patch-clamp technique. Antagonism of receptor-operated calcium channels and voltage-dependent calcium channels (VDC) in vascular smooth muscle by CPU86017 and the four isomers were tested as suppression of phenylephrine- or KCl-induced contractions of aortic rings, respectively. 3. For I(Kr.tail) inhibition, the IC(50) of SS, SR, RR, RS and CPU86017 was 2.86 +/- 1.20, 39.4 +/- 8.5, 3.48 +/- 0.80, 7.65 +/- 1.50 and 12.5 +/- 7.8 x 10(-9) mol/L, respectively; for I(Ks.tail) inhibition IC(50) values were 16.9 +/- 4.0, 20.0 +/- 2.1, 99.1 +/- 5.9, 160 +/- 81 and 65.0 +/- 4.7 x 10(-9) mol/L, respectively. The SR isomer showed balanced blockade of I(Kr) and I(Ks) that was associated with a loss of a-adrenoceptor antagonism but enhanced VDC blockade. 4. Configuration of 13aC critically determines I(Kr) blockade and the Ca(2+) antagonism of the isomers of CPU86017. The SR isomer exhibits mild blockade of I(Kr), moderately enhanced blockade of I(Ks) and Ca(2+) influx and less a-adrenoceptor antagonism compared with the racemate and may be promising as an anti-arrhythmic.     

大鼠血浆中抗精神分裂症候选新药SIPI6360立体异构体的LC-MS/MS法测定

建立了液相色谱-串联质谱法拆分并测定大鼠血浆中SIPI6360的立体异构体.采用Daicel Chiralpak IA手性柱,以甲醇为流动相,采用ESI源正离子模式、多反应监测进行定量分析.结果显示,(R)-(+)-SIPI6360与(S)-(-)-SIPI6360立体异构体分离良好,在0.5～100 ng/ml浓度范围内线性关系良好.方法回收率为94.0％～102.8％,批内和批间RSD小于5.2％和11.4％.     

Simultaneous High-Performance Liquid Chromatographic Analysis of Cefprozil Diastereomers in a Pharmacokinetic Study

Cefprozil, a new oral cephalosporin, consists of a 90:10 cis:trans isomer mixture. Sensitive, specific and reproducible high performance liquid chromatographic methods have been developed for the simultaneous quantification of the two stereoisomers of cefprozil in plasma and urine samples from human and rats. Cephalexin acted as the internal standard. Plasma protein was precipitated with acetonitrile and trichloracetic acid with subsequent extraction of acetonitrile. After vortexing and centrifuging, the aqueous phase was injected onto a reverse phase C8 column. Urine samples were acidified with sodium acetate buffer (pH 3.8) and then directly injected onto a reverse phase C18 column. The detector was set at 280 nm. These methods were applied to determine protein binding of both isomers in human and rat sera, and to perform a pharmacokinetic study in human. Results showed that both isomers bound moderately to serum proteins with no interference by the other isomer. The pharmacokinetic study in human indicated that cefprozil was well absorbed and the cis and trans isomers have similar pharmacokinetics.