Interaction of stereoisomers of barbiturates with [H]α-dihydropicrotoxinin binding sites

Racemic depressant barbiturates inhibit the binding of gamma-aminobutyric acid antagonist [³H]α-dihydropicrotoxinin (DHP) to rat brain membranes with IC₅₀ values ranging from 5 to 50 μM. The (−) isomers of pentobarbital and secobarbital were three to four-fold more potent than their (+) isomers in inhibiting [³H]DHP binding. In contrast, the (+) isomer of hexobarbital was a better inhibitor than (−) hexobarbital. The stereoisomers of 1-methyl-5-phenyl-5-propyl barbituric acid (MPPB), which show opposite pharmacological activity, inhibited [³H]DHP binding in a biphasic manner with a plateau at 4–100 nM MPPB. These results suggest heterogeneity of DHP binding sites. The possibility that depressant and convulsant barbiturates may act at the level of DHP site at the GABA synapse is discussed.     

手性药物的质量控制研究

文中结合研发实例重点解读了《手性药物药学研究技术指导原则》中的药学研究思路与质量控制研究及稳定性研究等章节的内容,并归纳总结了在手性药物研发中的关键问题与常见问题。     

Carboxy-terminal extension stabilizes the topological stereoisomers of guanylin

The peptide hormone guanylin constitutes two topological stereoisomers, which are connected through an equilibrium of interconversion. To investigate the importance of amino acid residues in the central region between the inner cysteines and at the carboxy terminus for this isomerism, synthetic derivatives of guanylin were compared by HPLC, 2D ¹H NMR spectroscopy and by their guanylyl cyclase-C (GC-C)-activating potency. An increase in the central sterical bulk by introduction of diiodo-Tyr⁹ had virtually no effect on the isomerization kinetics. Compared to guanylin, carboxy-terminal amidation did not affect the equilibrium between the two isoforms either. In contrast, two significantly stabilized isomers were obtained by extending the carboxy terminus of guanylin with one additional leucine resembling the characteristic of human uroguanylin isomers. This effect was intensified by a further Lys-Lys extension, thus revealing that the conformational exchange between the guanylin isomers is dependent on the extent of the sterical hindrance in the carboxyterminal region of this peptide. Demonstrated by 2D NMR spectroscopy, the separated isomers of the carboxy-terminally extended derivatives of guanylin exhibit unambiguously closely related structures as found originally for guanylin isomers, which are only detectable as a mixture. Because only one of the stabilized guanylin isomers activates guanylyl cyclase-C, the three-dimensional structure of the GC-C-activating guanylin isomer is now defined. The stabilized isoforms of guanylin described in this study represent suitable tools for the separate functional investigation of the GC-C-agonistic isomer of guanylin as well as of its isomeric counterpart.     

In vitro and in vivo assessment of the effect of impurities and chirality on methamidophos-induced neuropathy target esterase aging.

In vitro and in vivo studies evaluated neuropathy target esterase (NTE) inhibition and aging (i.e., loss of reactivation potential) by analytical and technical grade racemic and resolved L-(-) and D-(+) isomers of methamidophos (O,S-dimethyl phosphoramidothioate). For studies in vitro, microsomal protein from phenobarbital-induced livers was isolated from chick embryos and NTE inhibition assays were performed using chick embryo brain homogenate treated with 1 or 5 mM methamidophos (with and without metabolic enzymes); for studies in vivo, hens received 30 to 35 mg/kg methamidophos injected into the pectoral muscle. NTE aging in hens was assessed 24 h later or after 30 min to 1 h incubation in vitro using solutions of potassium fluoride (KF) reactivator. Technical methamidophos produced significantly higher levels of aged-inhibited NTE than analytical methamidophos or isolated optical isomers. In vivo, technical methamidophos produced 61% total NTE inhibition with 18% aged and 43% unaged NTE; hens receiving analytical grade averaged 6% aged, 52% unaged, and 58% total NTE inhibition. Results for 1 mM analytical methamidophos in vitro were 5% aged, 54% unaged, and 59% total inhibition; for 1 mM technical methamidophos, values averaged 11% aged, 50% unaged, and 60% total NTE inhibition. The degree of NTE aging obtained both in vivo and in vitro for the isolated D-(+) and L-(-) isomers never exceeded that obtained using analytical grade. These data indicate that impurities in methamidophos could contribute to OPIDN potential. The in vitro methodology described could be applied to first tier screening for detection of NTE inhibition and aging, thus reducing the need for whole-animal testing for OPIDN.     

梭曼异构体气相色谱分析及在小鼠脑及膈肌中的浓度测定

目的 :建立灵敏、可靠的梭曼光学异构体气相分析方法以测定小鼠皮下染毒后脑与膈肌中游离梭曼的浓度。方法 :大进样量气相色谱仪及手性毛细管柱实现梭曼 4个异构体的分离 ;用二异丙基氟磷酸酯作内标 ,测定小鼠皮下染毒后脑与膈肌中游离C(± )P(- )梭曼。结果 :梭曼 4个异构体在确定的气相条件下得到了较好的分离 ;C(± )P(- )梭曼在小鼠脑匀浆中及膈肌匀浆中 ,0 .5～ 10ng/ 2ml浓度范围的标准曲线线性关系良好 ,r >0 .999。日间、日内精密度在 10 %以内 ,提取方法的回收率为 5 3.6%～ 60 .2 %。用气相色谱分析方法考察了小鼠皮下染毒后 ,脑与膈肌中游离C(± )P( )梭曼的浓度 ,方法重复性好。结论 :大进样量气相色谱仪及手性毛细管柱测定小鼠皮下染毒后脑与膈肌中游离C(± )P(- )梭曼的浓度 ,方法灵敏、可靠 ,重复性好     

Stereospecific inhibition of monoamine uptake transporters by meta-hydroxyephedrine isomers

Summary. Meta-hydroxyephedrine (HED) comprises four stereoisomers consisting of two enantiomeric pairs related to ephedrine and pseudoephedrine. HED is transported into adrenergic neurons and radiolabeled HED has been employed in positron emission tomography (PET) to image adrenergic neurons in vivo. To extend structure-activity analyses of binding sites within monoamine transporters and to determine which stereoisomer displayed the best selectivity for PET imaging applications, we tested the HED compounds for their abilities to inhibit [³H]neurotransmitter uptake into platelets, transfected cells, and chromaffin vesicles. We hypothesized that the HED compounds would be most potent at the norepinephrine transporter (NET) compared to the serotonin or dopamine transporters and that the 1R diastereomers would be more effective than 1S diastereomers. Supporting the hypotheses, all stereoisomers were most potent at the NET and the 1R,2S stereoisomer was the most potent inhibitor overall. However, the 1S,2R isomer may be preferred for PET applications because of better selectivity among the transporters and reduced neuronal recycling. Received October 22, 2001; accepted January 14, 2002 Published online June 28, 2002     

诺美孕酮乙酸酯3—（O—羧甲基）肟的合成及其异构体的分离

诺美孕酮乙酸酯与羧甲氧基胺半盐酸盐在含少量吡啶的甲醇中室温反应,反－诺美孕酮乙酸酯３－（Ｏ－羧甲基）肟的两种立体异构体的混合物。经层析分离以２：１的比例得到两种立体异构体。     

依帕司他原料药有关物质的HPLC测定方法改进研究

目的 采用改进的高效液相色谱法测定依帕司他原料药有关物质。方法 采用依利特Hypersil BDS C₁₈色谱柱（250 mm×4.6 mm,5 μm）,以乙腈-pH6.5磷酸盐缓冲液（0.025 mol·L^-1 KH₂PO₄+0.025 mol·L^-1 Na₂HPO₄,用H₃PO₄调节pH至6.5）为流动相,梯度洗脱,流速为1.0 mL·min^-1,检测波长为396 nm和280 nm,柱温为30℃,进样量为20 μL。结果 依帕司他与相邻杂质能较好地分离。立体异构体杂质在浓度0.010~30 μg·mL^-1内线性关系良好,检测限和定量限分别为0.80,2.4 ng·mL^-1。精密度、稳定性、准确度、耐用性均符合要求。结论 该方法可用于依帕司他原料药有关物质的检查。     

EFFECTS OF ALBUMIN ON THE DISPOSITION OF MORPHINE AND MORPHINE-3-GLUCURONIDE IN THE RAT ISOLATED PERFUSED LIVER

1. The effect of albumin on the disposition of morphine and hepatically generated morphine-3-glucuronide (M3G) was investigated in the single-pass rat isolated perfused liver. 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.