液相色谱-质谱联用法测定人血浆中多潘立酮

目的:建立测定人血浆中多潘立酮的液相色谱-电喷雾串联质谱(LC/ESI-MS/MS)法。方法:待测血浆0.1 mL 用甲醇沉淀蛋白,取离心后的上清液进样10μL在 Phenomenex Gimim-C_(18)柱(2.00 mm×50 mm,5 μm)上分离,流动相为甲醇-水(40:60,v/v,含0.3%醋酸),流速0.2 mL·min~(-1),LC/ESI-MS/MS 采用多离子反应监测,正离子模式,用于定量分析的离子反应分别为 m/z426→m/z175(多潘立酮)和 m/z 379→m/z 264(氨溴索,内标)。结果:血浆中的内源性物质不干扰测定,每个样品分析时间约2 min;本法线性范围为0.3～100 ng·mL~(-1),最低定量浓度为0.3 ng·mL~(-1);日内、日间 RSD 分别小于7.1%和13.3%,相对误差小于5.4%。结论:该法操作简便、快速、准确,灵敏度高,可用于多潘立酮临床治疗剂量的药物动力学研究。     

HPLC-MS-MS法测定人血浆中托吡酯的浓度

目的:建立测定人血浆中托吡酯浓度的HPLC-MS-MS方法。方法:采用液液萃取法,以甲基叔丁基醚为提取溶剂;HPLC采用Hypersil ODS C18色谱柱(150 mm×4.6 mm,5μm),流动相为5 mmol·L-1醋酸铵溶液-甲醇(25∶75),流速0.5 mL·min-1;质谱采用ESI源,负离子检测模式,定量分析离子为托吡酯m/z338.1[M-H]-→78.0,双氯芬酸钠m/z294.1[M-H]-→249.7。结果:人血浆中托吡酯的标准曲线范围为10.00~3 600.00 ng.mL-1(n=7),r>0.999;日内和日间精密度均小于8.9%;提取回收率(低、中、高)分别为(75.5±3.1)%,(77.0±1.6)%和(72.8±1.5)%,内标提取回收率为(76.1±3.5)%;基质效应(低、中、高)分别为(99.2±5.6)%,(98.9±4.2)%和(99.4±3.8)%,内标基质效应为(100.4±1.3)%。结论:该方法灵敏、快速、专属性强,可用于人血浆中托吡酯浓度的测定。     

高效液相色谱-质谱法测定大鼠血浆中硫酸酯型脱氢表雄酮和孕烯醇酮

建立同时测定 SD 大鼠血浆中脱氢表雄酮硫酸酯(DHEAS)和孕烯醇酮硫酸酯(PREGS)的高效液相色谱-质谱法。方法:选择雌酮硫酸酯(ES)作为内标。雄性 SD 大鼠的血浆使用乙腈沉淀除去蛋白后,经固相萃取、水解及衍生化后,以高效液相色谱质谱检测器测定脱氢表雄酮硫酸酯和孕烯醇酮硫酸酯的浓度。色谱柱为 Zorbax SB C_(18)柱温40℃,流动相为乙腈水,使用梯度洗脱;采用大气压化学离子源(HPLC/APCI-MS),负离子检测方式,用于选择离子检测(SM)定量分析的离子为 m/z 490.0(DHEAS)和 m/z 518.0(PREGS)及 m/z 472.1(ES)[M-H]~-。结果:血浆中脱氢表雄酮硫酸酯和孕烯醇酮硫酸酯线性范围分别为:0.05～2.0ng·mL~(-1),0.03～2.0 ng·mL~(-1),r 分别为0.9990和0.9979。低、中、高浓度血样的提取回收率在66.8%～82.3%之间,相对回收率为:98.4%～111.6%,日内、日间变异均小于15%。正常雄性 SD 大鼠的血浆脱氢表雄酮硫酸酯和孕烯醇酮硫酸酯分别为0.070±0.020 ng·mL~(-1)、0.13±0.031 ng·mL~(-1)。结论:本实验方法具有灵敏度高、准确度好及变异较小的特点,适合测定雄性 SD 大鼠血浆中脱氢表雄酮硫酸酯和和孕烯醇酮硫酸酯的含量。     

HPLC／MS／MS测定恒河猴血浆中纳曲酮及其代谢物纳曲醇的浓度

目的:建立高效液相色谱-质谱联用法测定恒河猴血浆中纳曲酮及其代谢物纳曲醇的浓度。方法:液相色谱:色谱柱为Nucleocil ODS(5μm,50 mm×2.0 mm),流动相为乙腈-甲醇-水-冰醋酸(20:20:60:1),流速为0.2 mL·min-1;质谱:离子源为Turbo Ionspray源,采用多反应监测(MRM)方式进行检测,m／z342.3→324.3(纳曲酮)、m／z 344.3→326.6(纳曲醇,代谢产物)、m／z 328.4→310.4(纳洛酮,内标)。样品经有机相提取纯化后进样。结果:纳曲酮和纳曲醇的线性范围分别为0.098-150 ng·mL-1和0.781-25 ng·mL-1,萃取回收率均大于75％,日内误差(RSD)均小于9.3％,日间误差(RSD)均小于12％。结论:本方法准确、快速、灵敏、专属性好,适用于动物试验及临床上测定血浆中纳曲酮及其代谢物纳曲醇的浓度和药代动力学研究。     

液质联用测定人血浆中泮托拉唑的血药浓度

目的建立液质联用测定人血浆泮托拉唑药物浓度的方法。方法采用WatersAtlantisTMC18柱(2.1mm×100mm,3.0μm),流动相:0.1%甲酸溶液-乙腈(40∶60),流速0.2mL·mL-1,柱温30℃,进样量5μL,电喷雾电离源(ESI),MRM法检测,泮托拉唑母离子[M+H]m/z384.2,子离子m/z200.5,内标盐酸帕罗西汀母离子[M+H]m/z330.1,子离子m/z192.1。结果线性范围为10~5000ng·mL-1,最低检测浓度为1.0ng·mL-1,三种浓度的平均回收率均在90%以上,日内RSD为1.8%~4.0%(n=5),日间RSD为1.2%~1.7%(n=5)。结论该法操作简便,灵敏度高,分析快速,适用于临床上人血浆中泮托拉唑浓度测定。     

LC/MS/MS法测定人血浆中愈创木酚甘油醚浓度及药动学研究

目的:建立以高效液相色谱-质谱联用法测定人血浆中愈创木酚甘油醚浓度的方法,并研究愈创木酚甘油醚片的人体药动学。方法:碱化的血浆药品经乙酸乙酯萃取后,以甲醇-1%甲酸(70:30)为流动相,对乙酰氨基酚为内标,采用Aquasil C18柱分离。通过液相色谱-串联质谱联用仪,以选择反应监测方式进行检测,定量分析的离子反应分别为m/z199→m/z125(愈创木酚甘油醚)和m/z152→m/z110(对乙酰氨基酚)。结果:愈创木酚甘油醚检测浓度在5·0～2500ng·mL-1范围内线性关系良好(r=0·9953),定量下限为5·0ng·mL-1。20名受试者口服愈创木酚甘油醚片0·2g后的主要药动学参数Cmax为(754·6±190·5)ng·mL-1、t1/2为(0·97±0·12)h、tmax为(0·63±0·22)h、AUC0～t为(1435·8±441·9)ng·h·mL-1、AUC0～∞为(1444·9±449·3)ng·h·mL-1。结论:本方法简便、快速、灵敏,可用于愈创木酚甘油醚的临床药动学研究。     

高效液相色谱法同时测定人血浆中单硝酸异山梨酯、阿司匹林和水杨酸

目的:建立同时测定人血浆中单硝酸异山梨酯、阿司匹林及其代谢物水杨酸浓度的高效液相色谱法。方法:以茶碱为内标,血浆经酸化后,乙酸乙酯提取,高效液相色谱质谱检测器(MSD)和二极管阵列检测器(DAD)串联在线测定血浆中单硝酸异山梨酯、阿司匹林和水杨酸的浓度;分析柱:Zorbax DB-C18(5μm,2.1 mmx150mm),保护柱:Zorbax DB-C8(5μm,2.1mm×12.5 mm),柱温30℃;流动相为A:3 mmol·L-1乙酸铵溶液(含冰乙酸0.022％),流速为0.17 mL·min-1,B:甲醇-乙腈(85:15),流速为0.03 mL·min-1。单硝酸异山梨酯和阿司匹林用高效液相-电喷雾质谱(HPLC/ESI-MS)选择离子检测,水杨酸用DAD检测。结果:血浆中单硝酸异山梨酯、阿司匹林和水杨酸线性范围分别为0.00625-1.2 μg·mL-1,0.025-1.2μg·mL-1,0.0625-12μg·mL-1;相对回收率分别为101.3％-103.0％,99.8％-119.7％,98.4％-104.0％;提取回收率均高于80％;日内、日间精密度均小于8％。结论:方法简便、快速、重现性好,可用于临床血药浓度监测和复方制剂的药代动力学和生物利用度研究。     

HPLC-MS法研究氯雷他定片人体相对生物利用度及药代动力学

目的:建立准确高灵敏的HPIC-MS法测定氯雷他定在血浆中的浓度,以研究氯雷他定片在健康华人志愿受试者中的药代动力学和相对生物利用度。方法:对20例健康华人男性志愿受试者进行二交叉试验,随机单剂量口服氯雷他定(20 mg)试验或参比片后,按设定时间采集肘静脉血;分取血浆1.0 mL,经0.1 mol·L-1氢氧化钠溶液0.1 mL碱化和5 mL乙酸乙酯萃取处理;采用Zorbax-Ext-ODS柱,甲醇-乙腈-2.0％醋酸溶液(62:10:28)流动相,HPLC-MS内标(地西泮,[MH]+,m/z=285)法选择性正离子检测法测定氯雷他定([MH]+,m/z=383)血浆浓度。结果:HPLC-MS法测定血浆中氯雷他定的最低检测限为0.1 ng·mL-1,在0.2-20 ng·mL-1范围内线性关系良好;萃取绝对回收率为79.3％-84.0％;血药浓度测定日内、日间RSD均小于:15％。测得口服氯雷他定(20 mg)试验和参比片后的主要药代动力学参数Cmax(ng·mL-1)分别为8.3±3.7和6.8±5.4;Tmax(h)分别为1.3±0.6和1.3士0.4;tl/2(h)分别为4.6±1.4和5.1±1.4;AUCO→∞(ng·h·mL-1)分别为28.4±21.2和25.6±16.2,相对生物利用度F为(112.9±24.7)％。结论:建立的HPLC-MS方法灵敏、准确,测定结果可靠;统计分析表明氯雷他定试验和参比片生物等效。     

LC-MS/MS法同时测定人血浆中氯吡格雷及其羧酸代谢物的浓度

目的建立快速测定人血浆中氯吡格雷及其羧酸代谢物SR26334含量的LC-MS/MS法。方法乙醚-正己烷(4:1,V:V)2次液-液提取法(中性和酸化条件下),采用Teknokroma C_(18)色谱柱,以那格列奈和吡格列酮为内标,同时测定血浆中氯吡格雷和SR26334的浓度。流动相:甲醇-0.1%甲酸(80:20,V:V);流速:0.2mL·min~(-1);以多反应离子监测方式检测:氯吡格雷[M+H]~+,m/z 322.1→212.1;那格列奈[M+H]~+,m/z 318.3→166.2;氯吡格雷羧酸代谢物SR26334[M+H]~+,m/z 308.1→q98.1;吡格列酮[M+H]~+,m/z 357.2→134.2。结果氯吡格雷和内标那格列奈的保留时间分别在4.4和3.7min,SR26334和内标吡格列酮的保留时间分别在1.3和1.7min,氯吡格雷的线性范围为5～5000ng·L~(-1);SR26334的线性范围为20～2500μg·L~(-1)。提取回收率大于75%,方法回收率大于90%,日内、日间RSD小于10%(n=5)。结论本方法简便快速,适用于氧吡格雷制剂的新药临床研究和临床长期治疗病人血药浓度的常规监测。     

UPLC-QTRAP-MS/MS法同时测定大鼠血浆中洛伐他汀和活性代谢产物洛伐他汀酸的浓度及其药动学研究

摘要 目的：建立测定大鼠血浆中洛伐他汀及其活性代谢产物洛伐他汀酸浓度的超高效液相三重四级杆线型离子阱串联质谱检测方法,并将其应用于药动学研究。方法：采用乙酸乙酯萃取血浆样品,以辛伐他汀为内标,采用Agilent ZORBAX C18 2.1 mm × 50 mm, 1.8 μm）为色谱柱,以乙腈（A）-0.1%甲酸（B）为流动相,梯度洗脱流速为0.4 mL·min-1,柱温40°C。以正离子多反应离子监测模式,用于定量分析的离子对分别为m/z 405.1?225.1（洛伐他汀）,m/z 423.2?199.2（洛伐他汀酸）,m/z 419.2?199.1（辛伐他汀）。结果：洛伐他汀和洛伐他汀酸的线性范围分别为0.08-80.0 ng·mL-1和1.60-1600.0 ng·mL-1,其定量下限（LLOQ）分别为0.08 ng·mL-1和1.60 ng·mL-1;洛伐他汀和洛伐他汀酸的日内和日间精密度均小于9.0%,准确度在-5.18%-6.83%范围内;洛伐他汀和洛伐他汀酸的提取回收率均大于72.26%,RSD均小于10.83%;洛伐他汀和洛伐他汀酸的基质效应在91.11%-105.73%,RSD均小于7.46%;洛伐他汀和洛伐他汀酸的Tmax分别为1.58 ± 0.11 h和2.15 ±0.26 h,Cmax分别为136.22 ± 30.25 ng·mL-1和212.57 ± 33.92 ng·mL-1,t1/2分别为35.42 ± 12.67 h和17.86 ± 6.15 h,AUC0?24 h分别为279.92 ± 44.18 ng h·mL-1和390.34 ±20.15 ng h·mL-1。结论：该方法快速、灵敏、准确,可用于洛伐他汀和洛伐他汀酸在大鼠体内的药动学研究。     

Efficacy of gut lavage,hemodialysis, and hemoperfusion in the therapy of paraquat or diquat intoxication

Clinical and in vitro investigations were carried out to test the efficacy of gut lavage, hemodialysis, and hemoperfusion in the treatment of poisoning with paraquat or diquat. In a patient suffering from diquat intoxication 130 times more diquat was removed by gut lavage 30 h after ingestion than was removed by complete aspiration of the gastric contents.Determination of in vitro clearances for paraquat and diquat by hemodialysis showed that, at serum concentrations of 1–2 ppm, such as are frequently encountered in poisoning in man, toxicologically relevant quantities of herbicide cannot be removed from the body. At a concentration of 20 ppm, on the other hand, hemodialysis proved to be effective, the clearance being 70 ml/min at a blood flow rate of 100 ml/min. The efficacy of hemoperfusion with coated activated charcoal was on the whole better. Especially at concentrations around 1–2 ppm, the clearance values for hemoperfusion were some 5–7 times higher than those for hemodialysis.In a patient suffering from paraquat poisoning, both hemodialysis as well as hemoperfusion were carried out. The in vitro results could be confirmed: At serum concentrations of paraquat less than 1 ppm no clearance could be obtained by hemodialysis while by hemoperfusion with activated charcoal quite high clearance values were measured and the serum level dropped down to zero.

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Zusammenfassung Klinische Untersuchungen und Laboratoriumsversuche wurden durchgeführt, um die Wirksamkeit von Darmspülung, Hämodialyse und Hämoperfusion bei Paraquat- und Deiquat-Vergiftungen zu prüfen.Bei einem Patienten wurde 30 Std nach Deiquat-Aufnahme durch Darmspülung 130mal mehr Deiquat entfernt als durch vollständige Aspiration des Mageninhaltes. In vitro-Versuche ergaben, daß bei Blutserumkonzentrationen von 1–2 ppm, die bei Vergiftungen oft gemessen werden, durch Hämodialyse keine toxikologisch relevanten Paraquat- oder Deiquat-Mengen entfernt werden können. Dagegen erwies sich die Hämodialyse bei 20 ppm und einer Blutumlaufgeschwindigkeit von 100 ml/min mit einer Clearance von 70 ml/min als wirksam. Die Hämoperfusion mit beschicheter Aktivkohle war in diesen Versuchen aber eindeutig überlegen, denn insbesondere bei Konzentrationen um 1–2 ppm waren die Clearance-Werte 5–7mal höher als bei der Hämodialyse.Die in vitro-Ergebnisse wurden bei einem Patienten mit einer Paraquat-Vergiftung bestätigt: Bei Konzentrationen unter 1 ppm war die Hämodialyse wirkungslos, während durch Hämoperfusion relativ hohe Clearance-Werte erreicht wurden, so daß der Serumspiegel rasch unter die Nachweisgrenze abfiel.

     

In vitro studies on sterically stabilized liposomes (SL) as enzyme carriers in organophosphorus (OP) antagonism

This study describes a new approach for organophosphorous (OP) antidotal treatment by encapsulating an OP hydrolyzing enzyme, OPA anhydrolase (OPAA), within sterically stabilized liposomes. The recombinant OPAA enzyme was derived from Alteromonas strain JD6. It has broad substrate specificity to a wide range of OP compounds: DFP and the nerve agents, soman and sarin. Liposomes encapsulating OPAA (SL)* were made by mechanical dispersion method. Hydrolysis of DFP by (SL)* was measured by following an increase of fluoride ion concentration using a fluoride ion selective electrode. OPAA entrapped in the carrier liposomes rapidly hydrolyze DFP, with the rate of DFP hydrolysis directly proportional to the amount of (SL)* added to the solution. Liposomal carriers containing no enzyme did not hydrolyze DFP. The reaction was linear and the rate of hydrolysis was first order in the substrate. This enzyme carrier system serves as a biodegradable protective environment for the recombinant OP-metabolizing enzyme, OPAA, resulting in prolongation of enzymatic concentration in the body. These studies suggest that the protection of OP intoxication can be strikingly enhanced by adding OPAA encapsulated within (SL)* to pralidoxime and atropine.     

Synthesis,Cytotoxicity and Antileishmanial Activity of Some N-(2-(indol-3-yl)ethyl)-7-chloroquinolin-4-amines

We report herein the condensation of 4,7-dichloroquinoline (1) with tryptamine (2) and D-tryptophan methyl ester (3) . Hydrolysis of the methyl ester adduct (5) yielded the free acid (6) . The compounds were evaluated in vitro for activity against four different species of Leishmania promastigote forms and for cytotoxic activity against Kb and Vero cells. Compound (5) showed good activity against the Leishmania species tested, while all three compounds displayed moderate activity in both Kb and Vero cells.     

Steroidal sapogenin contents in some domestic plants

In order to find out the values of the steroid resources for the future use. the compositions and contents of steroidal sapogenins from 13 domestic plants have been investigated. As a result,Dioscorea nipponica, D. quinqueloba andSmilax china were found to have large amount of diosgenin. And pennogenin inTrillium kamtschaticum andParis verticillata, yuccagenin inAllium fistulosum, hecogenin inAgave americana and neochlorogenin inSolanum nigum were appeared to be major steroidal sapogenins.     

Aquatic Insects and Trace Metals: Bioavailability,Bioaccumulation, and Toxicity

Abstract

The uptake of metals from food and water sources by insects is thought to be additive. For a given metal, the proportions taken up from water and food will depend both on the bioavailable concentration of the metal associated with each source and the mechanism and rate by which the metal enters the insect. Attempts to correlate insect trace metal concentrations with the trophic level of insects should be made with a knowledge of the feeding relationships of the individual taxa concerned. Pathways for the uptake of essential metals, such as copper and zinc, exist at the cellular level, and other nonessential metals, such as cadmium, also appear to enter via these routes. Within cells, trace metals can be bound to proteins or stored in granules. The internal distribution of metals among body tissues is very heterogeneous, and distribution patterns tend to be both metal and taxon specific. Trace metals associated with insects can be both bound on the surface of their chitinous exoskeleton and incorporated into body tissues. The quantities of trace meals accumulated by an individual reflect the net balance between the rate of metal influx from both dissolved and particulate sources and the rate of metal efflux from the organism. The toxicity of metals has been demonstrated at all levels of biological organization: cell, tissue, individual, population, and community. Much of the literature pertaining to the toxic effects of metals on aquatic insects is based on laboratory observations and, as such, it is difficult to extrapolate the data to insects in nature. The few experimental studies in nature suggest that trace metal contaminants can affect both the distribution and the abundance of aquatic insects. Insects have a largely unexploited potential as biomonitors of metal contamination in nature. A better understanding of the physico-chemical and biological mechanisms mediating trace metal bioavailability and exchange will facilitate the development of general predictive models relating trace metal concentrations in insects to those in their environment. Such models will facilitate the use of insects as contaminant biomonitors.