毛细管电色谱手性配体交换法检测生物样品中的硝基精氨酸异构体研究

目的：建立毛细管电色谱（CEC）分离检测生物样品中硝基精氨酸异构体的方法。方法：采用CEC手性配体交换模式检测血浆样品中D-硝基精氨酸（D—NNA）和L-硝基精氨酸（L—NNA），流动相为50mmol／L醋酸缓冲液[pH5．0，含2mmol／Laspartame，1mmol／L Cu^2＋和5％（v／v）甲醇]；流速为0．02mL／min；操作压为1000psi；检测波长为UV280nm。结果：L—NNA和D—NNA在0．025～0．75mmol／L的浓度范围内峰面积／浓度的相关系数均可达0．99以上，日内变异系数均小于3．0％，日间变异系数分别为3．1％和3．4％。结论：本研究方法具有快速、高分辨、检测样品量和流动相用量少等优点。     

肾脏是体内N~G-D-硝基精氨酸单向手性转化的主要器官

目的　研究NG D 硝基精氨酸 (NG nitro D arginine,D NNA)在体内发生手性转化的的部位。方法　考察肾脏结扎对于D NNA诱导大鼠动脉压升高活性的变化 ,并运用毛细管电色谱 (capillaryelectrochromatography ,CEC)分析技术测定D NNA在体内手性转化的情况。结果　肾脏结扎使大鼠完全丧失对D NNA (32mg·kg-1)升高血压反应 ,但不影响由NG L 硝基精氨酸 (NG nitro L arginine ,L NNA)(16mg·kg-1)引起的升压反应。而D NNA/肾匀浆孵育液(32mg·kg-1)则可使肾脏结扎大鼠产生升压反应。CEC血药检测证实D NNA在大鼠体内转化生成L NNA ,而L NNA在体内未被代谢生成D NNA ;肾脏结扎则使D NNA的手性转化大量减少。结论　D NNA在大鼠体内可单向代谢转化L NNA ,肾脏是D NNA发生手性转化的主要器官     

布洛芬精氨酸注射及口服给药在大鼠体内的立体选择性药代动力学

利用手性高效液相色谱法研究大鼠注射及口服布洛芬精氨酸之后布洛芬对映体的药代动力学。布洛芬精氨酸注射及口服给药后, 布洛芬对映体的药代动力学呈现立体选择性, 且口服给药后立体选择性程度更高。与系统前转化相比, R-布洛芬至S-布洛芬的系统转化在口服给药后立体选择性药代动力学中起更重要的作用。布洛芬精氨酸口服给药后, 优势对映体S-布洛芬迅速吸收, S-布洛芬与R-布洛芬的绝对生物利用度分别为100% 和80%。基于研究发现的S-布洛芬体内系统性暴露, 可以推断布洛芬精氨酸注射及口服给药后的药理作用相似, 仅在作用的起始阶段存在差异。     

毛细管电谱法分离手性药物

毛细管电色谱（CEC）有机地结合了高效液相色谱的多选择性和毛细管电泳的高效性,是一种新型的微柱分离方法。本文综述了毛细管电色谱在手性分离方面的分离方式及其应用,并对影响手性分离的主要因素作了介绍。     

手性药物代谢动力学的立体选择性

手性是生物体系的一个基本特征。手性大分子物质,如酶、载体、受体、血浆蛋白和多糖等构成了生物体的手性内环境。在体内,手性药物对映体与生物大分子间相互识别、相互作用的立体选择性导致了手性药物在药代动力学方面的立体选择性,其主要表现在手性药物的吸收、分布、代谢和排泄四个过程的差异。     

手性药物对映体的药物代谢动力学

目前世界上,临床应用的药物中具有光学活性的超过60%,而这些具有手性特征的药物中,绝大多数人工合成的药物通常是以外消旋体方式给药的[1～4]。药物的外消旋体引入体内后,其对映体分子均由体内具手性的蛋白质、酶和受体以两个不同的分子处理。因而,药物对映体在体内可具有不同的代谢途径和药理作用。随着现代先进的分析技术的发展,手性拆分技术得到进一步的提高,可以对生物体液中的手性药物的异构体进行分析测定,这使得研究手性药物在生物体内的药物代谢动力学过程成为可能,而这对深入研究手性药物的药理和毒理作用具有重大意义。手性药物的…     

分子印记技术在毛细管电色谱手性分离中的应用

分子印记技术是制备具有预定选择性分离介质的有效途径，其应用于毛细管电色谱(CEC)手性分离具高效及独特功用。综述分子印记技术的基本原理及其在CEC中的应用模式和实例。     

一叶萩碱的高效毛细管电泳手性分离及其大鼠体内立体选择性代谢研究

目的建立一叶萩碱(SE)的高效毛细管电泳手性分离方法。方法以羟丙基-β-环糊精(HP-β-CD)为手性选择剂,测定条件为:分离介质32 mmol·L^-1 HP-β-CD的Tris-H₃PO₄缓冲液(40 mmol·L^-1,H₃PO₄调至pH 6.0);分离电压15 kV,柱温16℃,压力进样6 s,检测波长254 nm;大鼠各生物样品碱化后乙酸乙酯萃取。结果测定条件下SE基本达到基线分离,大鼠生物样品测定不受内源及代谢物干扰。大鼠ip SE经胆汁、尿和粪排泄以D型为主,具有立体选择性。结论本法简便可靠,可适用于SE在大鼠体内立体选择性代谢研究。     

一叶萩碱的高效毛细管电泳手性分离及其大鼠体内立体选择性代谢研究

目的　建立一叶碱 (SE)的高效毛细管电泳手性分离方法。方法　以羟丙基 β 环糊精 (HP β CD)为手性选择剂 ,测定条件为 :分离介质 3 2mmol·L- 1 HP β CD的Tris H3PO4 缓冲液 ( 4 0mmol·L- 1 ,H3PO4 调至pH 6 0 ) ;分离电压 15kV ,柱温 16℃ ,压力进样 6s ,检测波长 2 5 4nm ;大鼠各生物样品碱化后乙酸乙酯萃取。结果　测定条件下SE基本达到基线分离 ,大鼠生物样品测定不受内源及代谢物干扰。大鼠ipSE经胆汁、尿和粪排泄以d型为主 ,具有立体选择性。结论　本法简便可靠 ,可适用于SE在大鼠体内立体选择性代谢研究。     

Chiral pharmacokinetics and inversion of N^G-nitro-arginine in the rat

AIM: To explore pharmacokinetics of N^G-nitro-D-arginine (D-NNA) and N^G-nitro-L-arginine (L-NNA) in conscious rats.METHODS: The plasma concentration of D-NNA and L-NNA were determined by chiral ligand exchange method with capillary electrochromatography (CEC). Pharmacokinetic parameters were estimated using non-compartment model and were fitted using a computer program DAS. Chiral inversion rate of D-NNA to L-     

Chiral pharmacokinetics and inversion of enantiomers of a new quinoxaline topoisomerase IIbeta poison in the rat.

XK469 (NSC 697887; (+/-)-2-[4-(7-chloro-2-quinoxaliny)oxy]phenoxy propionic acid), an analog of the herbicide Assure, which possesses antitumor activity, especially against murine solid tumors and human xenografts, has recently been found to be the first topoisomerase IIbeta poison. Both R(+) and S(-) isomers are cytotoxic, although the R-isomer is more potent. Using a chiral high-performance liquid chromatography assay, pharmacokinetics of R(+)-, S(-)-, and (+/-)-XK469 in Fischer-344 rats were investigated following their separate i.v. administrations. S(-)-XK469 was found to be predominantly converted to the R-isomer in circulation when the S-isomer was administered either alone or as a racemic mixture. No trace of the S-isomer was found in circulation or in urine or feces, following the R-isomer administration, up to 72 h. In the rat, the plasma concentration-time profiles for both isomers follow a two-compartment pharmacokinetics with the mean t(1/2beta) for the R-isomer of 24.7 h being significantly longer than 4.2 h, the mean t(1/2beta) for the S-isomer. The mean total clearance of the S-isomer was over 200-fold more rapid than that of the R-isomer, and the major clearance route of the S-enantiomer was inversion to its antipode, as estimated by the fractional formation clearance of R(+)-XK469 of 0.93. Protein binding for both enantiomers was in the range of 95 to 98%. Urinary and fecal elimination in 72 h as the intact drug were 7 to 10% and 8% of the administered dose, respectively, either administered as the individual enantiomers or as a racemate. Cumulative biliary elimination in 7 h was about 3% of the dose. No evidence of enantiomeric interaction at the pharmacokinetic level was detected.     

Biological activation of NG-nitro-D-arginine by kidney homogenate

N^G-nitro-L-arginine (L-NNA) and D-NNA have been shown to inhibit endothelium-dependent relaxation. This study examined if the inhibitory effect of L-NNA or D-NNA on relaxation is increased following incubation of the drug with the supernatant of tissue homogenates. Acetylcholine (ACh) caused concentration-dependent relaxation of pre-constricted rat aortic rings with maximum relaxation of 95%. Maximum relaxations to ACh were reduced to 71 and 37% in the presence of D-NNA (40 μM) and L-NNA (1 μM), respectively. Relaxation to ACh was further reduced to 18% in the presence of D-NNA that was incubated for 1 h with the supernatant of kidney homogenate, but unaffected by D-NNA incubated with the supernatant of trichloroacetic acid-denatured kidney homogenate. Incubation of L-NNA (1 μM) with either kidney supernatant or denatured kidney supernatant for 1 h did not affect its inhibitory effect on ACh-induced relaxation. Neither 1 h’s incubation with plasma, or supernatants of liver, lungs or aorta homogenates affected the inhibitory action of D-NNA (40 or 120 μM) on ACh-induced relaxation. After D-NNA was incubated in kidney supernatant, its inhibitory effect on ACh-induced relaxation of the aorta was abolished by pretreatment of the aorta with L-arginine (L-Arg) but not D-Arg suggesting involvement of the L-Arg pathway. The results suggest that D-NNA is converted by the kidney to a compound that acts similar to L-NNA. There appears to be little conversion of L-NNA to D-NNA. Received: 21 April 1997 / Accepted: 20 June 1997     

左旋奥硝唑在人体内的构型转化及药动学研究

目的:研究左旋奥硝唑在健康人体内的构型转化及单次口服左旋奥硝唑的人体药代动力学。方法:24名健康志愿者,单剂量口服左旋奥硝唑片500 mg后72 h采集血浆样品,用手性拆分色谱柱分离、检测血浆中左旋奥硝唑与右旋奥硝唑;以高效液相色谱法测定血药浓度,用DAS ver 2.0软件计算药动学参数。结果:在给药后的健康志愿者体内未检出右旋奥硝唑。左旋奥硝唑人体内主要药动学参数分别为:Cmax=(28.86±8.77)mg.L-1,AUC0→t=(453.9±184.4)mg.h.L-1,Tmax=(0.57±0.36)h,V1/F=(31.7±15.4)L,CL/F=(2.42±0.69)L.h-1,t1/2β=(14.7±2.1)h,MRT=(17.6±1.8)h。结论:健康志愿者口服左旋奥硝唑后,在体内未发生构型转化。因而在对左旋奥硝唑血浆样品进行检测时,可以不必考虑消旋化带来的影响。     

Stereoselective pharmacokinetics and inversion of suprofen enantiomers in humans.

The stereoselective pharmacokinetics of suprofen enantiomers has been studied in humans by means of stable isotope-labeled pseudoracemate-diastereomer methodology. After a single oral dose of a near equimolar mixture of unlabeled-(R)-(-)- and [2H3]-(S)-(+)-suprofen [or unlabeled-(S)- and [2H3]-(R)-suprofen] to three healthy male subjects, the plasma concentrations of drug were determined by a stereospecific gas chromatography-mass spectrometry method. Racemic [2H7]suprofen was used as an internal standard. The method involved chiral derivatization with (S)-(-)-1-(naphthyl)ethylamine to form the diastereomeric amide. The plasma concentrations were consistently higher for the (R)-isomer than the (S)-isomer. No significant difference in the elimination half-life of the enantiomers was observed. An average of 6.8% of an administered dose of the (R)-isomer was stereospecifically inverted to the (S)-isomer. There was no measurable inversion of the (S)- to (R)-isomer. The present stable isotope-labeled pseudoracemate-diastereomer methodology has made it possible to evaluate the pharmacokinetics of each enantiomer, including the estimation of chiral inversion after administration of the racemic mixture.     

灯盏花乙素苷元不同给药剂量的大鼠药动学比较

目的:比较灯盏花乙素苷元不同剂量给药的大鼠药动学。方法:12只大鼠随机均分为2组,分别单剂量灌胃给予灯盏花乙素苷元在20和200 mg·kg~(-1),用高效液相-电化学色谱法(HPLC-ECD)测定给药后不同时间血浆中灯盏花乙素苷元及其代谢物灯盏花乙素的浓度,计算AUC_(0-24h)。色谱柱为Kromasil C_(18)柱(150 mm×4．6 mm,5μm);流动相为pH 2．6的50 mmol·L~(-1)磷酸盐缓冲液-甲醇-四氢呋喃(60:40:10);流速为1．0 mL·min~(-1);分析电压为100 mV。结果:灯盏花乙素苷元20 mg·kg~(-1)剂量给药时,原型药完全被代谢,血浆中只检测到代谢产物灯盏花乙素;200 mg·kg~(-1)剂量给药时,血浆中代谢产物灯盏花乙素和原型药灯盏花乙素苷元共存。灯盏花乙素苷元200 mg·kg~(-1)剂量时,代谢产物灯盏花乙素的AUC值相仅较20 mg·kg~(-1)剂量时提高3倍(1934．83 vs 536．51 ng·mL~(-1)·h)。结论:大小剂量下灯盏花乙素苷元的药动学行为呈非线性。     

N-硝基-L-精氨酸对海洛因依赖性大鼠催促戒断的治疗作用及对生化指标的影响

目的:研究一氧化氮合酶抑制剂N-硝基-L-精氨酸对海洛因依赖性大鼠催促后出现的戒断综合征的治疗效果.方法:采用剂量递增法,皮下注射海洛因从1 mg·kg-1增至8 mg·kg-1,并维持该剂量至42 d,经纳洛酮催促后得到理想的大鼠海洛因依赖模型.在该模型的基础上选用3,9,18,27 mg·kg-1 4种N-硝基-L-精氨酸剂量给雄性大鼠灌胃.结果:发现4个剂量组与自然戒断组比较,经t检验后均有统计学意义.27 mg·kg-1的剂量组与自然戒断组评分比较有极显著性差异(P＜0.01), 其余3个剂量组与自然戒断组评分比较差异有显著性(P＜0.05).实验结果还发现,催促戒断的雄性大鼠在27 mg·kg-1 N-L-Arg的保护下,其腹泻症状和激惹症状与自然戒断组差异有显著性,分别为P＜0.01和P＜0.05.血清中NO、皮质醇和超氧化物歧化酶(SOD)活性测定结果显示:NOS抑制剂1,2 mg·kg-1可明显降低NO含量,而4 mg·kg-1·d-1无作用;长期使用NOS抑制剂N-硝基-L-精氨酸,SOD的活性升高,但各组间的皮质醇含量无明显差异.结论:实验结果表明,N-硝基-L-精氨酸对海洛因戒断综合征有明显的治疗作用,且呈剂量相关趋势,尤其对大鼠的腹泻和激惹症状有很好的疗效.     

Pressor effects of L and D enantiomers of NG-nitro-arginine in conscious rats are antagonized by L- but not D-arginine.

The effects of NG-nitro-L-arginine (L-NNA) and NG-nitro-D-arginine (D-NNA) on mean arterial pressure (MAP) were studied in conscious, unrestrained rats. I.v. bolus of either L-NNA (1-64 mg/kg) or D-NNA (2-64 mg/kg) dose dependently increased MAP to similar maximum values of 55 +/- 7 and 52 +/- 4 mm Hg and with ED50 values of 4.0 +/- 0.9 and 8.9 +/- 1.2 mg/kg (P less than 0.05), respectively. The time course of the MAP response to a single dose (32 mg/kg i.v. bolus) of L-NNA and D-NNA were also obtained. The pressor effects of L-NNA and D-NNA each lasted greater than 2 h with the rise phase t 1/2 of 5 and 27 min (P less than 0.05), respectively. I.v. infusions (10 mg/kg per min) of L-arginine (L-Arg) and D-arginine (D-Arg) did not alter the pressor response to noradrenaline nor angiotensin II. L-Arg but not D-Arg attenuated the pressor responses to both L-NNA and D-NNA. Therefore, both L-NNA and D-NNA are efficacious and long-lasting pressor agents; the pressor effects of both can be antagonized by L-Arg but not D-Arg. Our results suggest that the pressor effects of both L-NNA and D-NNA involve the L-Arg/nitric oxide pathway.     

Chiral separation of ibuprofen and chiral pharmacokinetics in healthy Chinese volunteers.

A rapid, sensitive and stereoselective HPLC method based on chiral column analysis was developed and fully validated for the simultaneous determination of the two enantiomers of ibuprofen in human plasma. Using this method, a chiral pharmacokinetic study of two different ibuprofen tablets, i.e. dexibuprofen tablets and racemic ibuprofen tablets, was carried out on 20 healthy Chinese male volunteers according to a single-dose (400 mg), two-way, cross-over randomized design. When a 'non-chiral calculation method' was used, the statistical analysis showed no significant difference for the pharmacokinetic parameters (AUC0-infinity, AUC0-t, Cmax and tmax) between the two oral formulations, suggesting that they were pharmaceutically bioequivalent. Considering that the pharmacological activity of ibuprofen resides exclusively in the S(+)-enantiomer, and that the unidirectional inversion of the R(-) to the S(+)-enantiomer is incomplete and might be race-dependent, the pharmacokinetic parameters for only the S(+)-enantiomer were further compared and the inversion ratio calculated. It was found that only 25% of R(-)-ibuprofen in the racemic ibuprofen tablets was inverted into S(+)-ibuprofen in the Chinese population, which suggested that dexibuprofen might possess a much stronger pharmacological activity than that of racemic ibuprofen when administered at the same dose.