UPLC-MS/MS检测大鼠血浆中阿帕替尼的浓度及其药动学研究

目的 采用UPLC-MS/MS建立快速检测大鼠血浆中阿帕替尼浓度的方法,并应用于药动学研究。方法 大鼠血浆样本用乙腈沉淀蛋白,液质联用技术检测浓度,流动相为乙腈-水（含0.1%甲酸）,梯度洗脱,流速为0.3 mL·min^-1,柱温40℃,内标为氯唑沙宗;质谱条件：电喷雾离子化源（ESI）,负离子监测模式,检测离子对阿帕替尼为m/z 396.2→210.0和m/z 396.2→158.0,氯唑沙宗m/z 168.0→132.0。结果 阿帕替尼和内标氯唑沙宗的保留时间分别为1.07 min和1.40 min,线性范围为10~2 000 ng·mL^-1（r²=0.993）,检测限为1 ng·mL^-1,准确度为90.65%~111.50%,基质效应为89.14%~104.65%,平均回收率>86%,日内、日间精密度RSD均<10%。常温下放置24 h、冻融2次和-80℃冻存30 d的RSD均<10%。药动学研究结果显示,大鼠单次灌胃阿帕替尼76.5 mg·kg^-1,AUC_（0-t）为（6 114.41±645.99）ng·mL^-1·h,CL_z/F为（12.21±1.08）L·h^-1·kg^-1,V_z/F为（75.70±38）L·kg^-1,T_1/2为（4.23±1.94）h,T_max为（2±0.71）h,C_max为（1 377.7±284.54）μg·L^-1。结论 该法操作简便,重复性好,准确可靠,适用于大鼠血浆中阿帕替尼的浓度检测及其药动学研究。     

UPLC-MS/MS测定大鼠血浆中柴胡皂苷B2、B4的含量及药动学研究

目的 建立UPLC-MS/MS测定大鼠血浆中柴胡皂苷B2、B4,并对柴胡皂苷B2、B4静脉给药后的药动学进行研究。方法 以柴胡皂苷F为内标,采用UPLC BEH C₁₈（2.1 mm×50 mm,1.7 μm）柱,柱温为40℃。流动相为乙腈-0.1%甲酸水溶液,梯度洗脱,流速为0.4 mL·min^-1,洗脱时间为4 min,使用氮气作为去溶剂化气体（800 L·h^-1）和锥形气体（50 L·h^-1）。MRM模式对柴胡皂苷B2 m/z 825.4→617.5、柴胡皂苷B4 m/z 957.6→649.6和柴胡皂苷F（内标）m/z 973.7→119.0进行定量分析。大鼠血浆用乙腈沉淀法去除蛋白。结果 在5~5 000 ng·mL^-1内,大鼠血浆中柴胡皂苷B2、B4线性良好（r>0.995）,定量下限为5 ng·mL^-1。柴胡皂苷B2日内精密度RSD<12%,日间精密度RSD<15%。柴胡皂苷B4日内精密度RSD<14%,日间精密度RSD<15%。柴胡皂苷B2、B4的准确度范围为89.3%~110.2%。结论 该分析方法灵敏、快速、选择性良好,并成功应用于柴胡皂苷B2、B4静脉给药后的大鼠药动学研究。     

UHPLC-MS/MS同时检测大鼠血浆中5种CYP450探针药物

目的 采用UHPLC-MS/MS同时检测大鼠血浆中非那西丁、甲苯磺丁脲、奥美拉唑、美托洛尔、咪达唑仑的血药浓度。方法 血浆样品经乙腈沉淀,采用Agilent ZORBAX Eclipse Plus C₁₈色谱柱（2.1 mm×50 mm,1.8 μm）;流动相为乙腈-含0.1%甲酸的水,梯度洗脱,流速为0.4 mL·min^-1。检测采用电喷雾离子源,多反应监测。非那西丁：[M+H]⁺,m/z 180.1→109.9;甲苯磺丁脲：[M+H]⁺,m/z 271.1→91.0;奥美拉唑：[M+H]⁺,m/z 346.1→135.9;美托洛尔：[M+H]⁺,m/z 268.2→115.0;咪达唑仑：[M+H]⁺,m/z 326.1→290.8;内标卡马西平：[M+H]⁺,m/z 237.1→194.0。6只♂ SD大鼠,单剂量口服灌胃10 mg·kg^-1非那西丁,1 mg·kg^-1甲苯磺丁脲,10 mg·kg^-1奥美拉唑,10 mg·kg^-1美托洛尔和10 mg·kg^-1咪达唑仑,分别在给药后多点尾静脉采血。用DAS计算药动学参数。结果 血浆中非那西丁、甲苯磺丁脲、奥美拉唑、美托洛尔和咪达唑仑在各自浓度范围内线性关系良好。日内及日间RSD均<15%,提取回收率>75%,稳定性考察结果良好。非那西丁的AUC_0-t为（5 868.30±2 062.87）ng·mL^-1·h;甲苯磺丁脲的AUC_0-t为（58 056.34±15 569.16）ng·mL^-1·h;奥美拉唑的AUC_0-t为（14 181.67±4 085.40）ng·mL^-1·h;美托洛尔的AUC_0-t为（1 123.67±180.469）ng·mL^-1·h;咪达唑仑的AUC_0-t为（946.91±322.03）ng·mL^-1·h。结论 该方法灵敏度高、操作方便、结果准确,可作为CYP450酶活性及相关研究的测定方法。     

HS-GC-MS/MS测定富马酸丙酚替诺福韦中微量的基因毒性杂质N-亚硝基二甲胺

目的 采用顶空进样气相色谱三重四级杆质谱联用（HS-GC-MS/MS）法测定富马酸丙酚替诺福韦中微量的基因毒性杂质N-亚硝基二甲胺（NDMA）。方法 采用三重四极杆GC-MS/MS，Agilent VF-WAX ms（30 m×0.25 mm，1 μm）色谱柱，载气：氦气；恒流模式1.0 mL·min^-1；程序升温，进样口温度230℃，顶空温度130℃；质谱采用电子轰击电离源（EI），电离能量为70 eV，离子源温度230℃，多反应监测（MRM）模式进行检测，溶剂为N-甲基吡咯烷酮（NMP）。进行专属性、系统适用性、检测限与定量限、线性与范围、准确度、精密度、溶液稳定性、耐用性考察。结果 NDMA与相邻色谱峰之间分离效果良好；NDMA在7.0～105.0 ng·mL^-1线性关系良好，检测限为3.5 ng·mL^-1，定量限为7.0 ng·mL^-1；NDMA低、中、高质量浓度（56、70、84 ng·mL^-1）回收率为95.6%～109.3%，RSD为4.0%（n=9）；重复性试验NDMA质量浓度RSD为6.5%，中间精密度RSD为6.1%；对照品溶液室温放置24 h稳定，供试品溶液室温放置90 h内溶液稳定；保持其他条件不变，分别改变进样口温度（230、225、235℃）、离子源温度（230、225、235℃）、载气体积流量（1.0、0.9、1.1 mL·min^-1）、顶空温度（130、128、132℃），方法耐用性良好。结论 所建立的方法准确度好、灵敏度高、简便可靠，对仪器污染小，可用于富马酸丙酚替诺福韦中NDMA的质量控制。     

青蒿琥酯自微乳在大鼠体内的药动学研究

目的 建立大鼠血浆中青蒿琥酯的HPLC-MS/MS测定方法,并研究青蒿琥酯自微乳在大鼠体内的药动学特征。方法 12只SD大鼠随机分为2组,单剂量分别灌胃（50 mg·kg^-1）青蒿琥酯自微乳和青蒿琥酯原料药,以格列吡嗪为内标,用LC-MS/MS测定给药后血浆中的药物浓度,并计算药动学参数。结果 青蒿琥酯血浆样品的线性范围为1.0~1 000.0 ng·mL^-1,回归方程为A=294.74C-439.33（r=0.999 6）,定量下限为1.0 ng·mL^-1。日内、日间变异系数（RSD）均<10%,符合生物样品的分析要求。青蒿琥酯原料药和青蒿琥酯自微乳的药动学参数C_max、t_1/2和AUC_0→t分别为：（87.6±8.80）ng·mL^-1,（1.88±0.33）h和（43.3±1.74）h·ng·mL^-1;（421±41.6）ng·mL^-1,（1.48±0.17）h和（282±17.7）h·ng·mL^-1。其中,C_max和AUC_0→t存在显著性差异（p<0.01）。结论 该方法简便灵敏,可用于血浆中青蒿琥酯的含量测定,经灌胃给药后,与原料药比较,青蒿琥酯自微乳能显著提高生物利用度。     

珍菊降压片在大鼠体内药动学研究

目的 建立同时测定大鼠血浆中芦丁和氢氯噻嗪的LC-MS/MS方法,并研究珍菊降压片在大鼠体内的药动学。方法 采用蛋白质沉淀法处理血浆样品,色谱柱为Pntulips BP-C₁₈柱（2.1 mm×50 mm,5μm）,流动相为乙腈-水梯度洗脱,柱温为30℃;流速为0.45 mL·min^-1。采用电喷雾离子源,选择离子反应监测模式。采用DAS 2.0软件计算药动学参数。结果 方法学验证结果表明内源性杂质不干扰待测物和内标的测定,芦丁和氢氯噻嗪的线性范围分别为5~1 000 ng·mL^-1（r²=0.997 1）和2.5~500 ng·mL^-1（r²=0.995 8）。芦丁和氢氯噻嗪药动学参数：AUC_（0-t）为（107 157.31±38 056.63）,（130 387.28±46 306.69）ng·mL^-1·min^-1;T_1/2z为（108.65±20.95）,（240.86±46.44）min;T_max为（34.25±16.34）,（120.00±0.00）min;C_max为（683.44±254.03）,（368.45±136.95）ng·mL^-1。结论 该方法准确度、精密度、回收率和基质效应均符合生物基质样品测试要求,适用于珍菊降压片在大鼠体内的药动学研究。     

GC-MS测定富马酸卢帕他定中痕量杂质偶氮二异丁腈

目的 采用气相色谱-质谱联用法测定富马酸卢帕他定中偶氮二异丁腈（azodiisobutyronitrile,AIBN）杂质的含量。方法 采用DB-624毛细管柱（30 m×0.25 mm,1.4 μm）,程序升温,初始温度为70℃,以12℃·min^-1升到250℃,以氦气为载气,流速为0.6 mL·min^-1,采用电子轰击电离源（EI）,在单离子检测模式（SIM）下选择m/z 69、m/z 54和m/z 41进行检测。结果 AIBN浓度在0.155 2~3.103 μg·mL^-1内与峰面积线性关系良好（r²=0.999 8）;检测限为0.045 μg·mL^-1,定量限为0.15 μg·mL^-1,样品中杂质AIBN测定结果的重复性良好,RSD（n=6）为0.31%;杂质AIBN低、中、高浓度的加样回收率（n=3）分别为103.9%,101.0%,99.4%,RSD（n=3）分别为0.03%,0.11%和0.08%。经检测,3批富马酸卢帕他定供试品中AIBN均未检出。结论 本方法操作简便,结果准确,灵敏度高,可用于富马酸卢帕他定中AIBN的检测。     

HS-GC-MS同时测定沙芬酰胺中磺酸酯类基因毒性杂质

目的 建立同时测定沙芬酰胺原料药中5种磺酸酯类基因毒性杂质（甲磺酸甲酯、甲磺酸乙酯、甲磺酸异丙酯、甲磺酸丙酯、甲磺酸丁酯）的方法。方法 采用顶空进样气相色谱-质谱联用法,在线衍生,RESTEK Rxi-624Sil毛细管柱（30 m×0.25 mm,1.4 μm）,四级杆质量检测器,离子化模式为电子轰击离子化（EI）模式,采集模式为选择离子监测。结果 5种杂质检测限为0.3~1.3 ng·mL^-1,定量限为0.9~4.3 ng·mL^-1;精密度、稳定性、重复性试验的RSD均<5%;浓度在5~300 ng·mL^-1内,5种杂质的峰面积与其相对应的浓度有良好的线性关系,相关系数（r²）均>0.995;回收率为99.4%~100.6%,RSD为0.8%~2.6%（n=9）。结论 该法操作简便,重复性好,结果准确可靠,可用于沙芬酰胺中磺酸酯类基因毒性杂质的测定。     

基质补偿-电感耦合等离子体质谱法测定脂肪乳注射液中铝元素含量

目的 建立脂肪乳注射液中铝元素含量的电感耦合等离子体质谱法（ICP-MS）测定方法。方法 采用基质饱和系统，直接稀释制样，在线内标引入方式，以电感耦合等离子体质谱法进行含量分析。结果 铝离子浓度在1~30 ng·mL^-1线性关系良好，相关系数（r）为0.999 4，仪器精密度、重复性的RSD均<3%，方法检测限为1.04 μg·L^-1，加样回收率为102.0%~113.3%，RSD为3.6%（n=9）。结论 本方法准确、灵敏高、简便可行，可为脂肪乳注射液铝元素含量测定提供有力技术保障。     

UPLC-MS/MS同时检测消癌平注射液中通关藤苷A、I、H的含量

目的 建立UPLC-MS/MS同时测定消癌平注射液中通关藤苷A、通关藤苷I、通关藤苷H的含量。方法 采用Phenomenex Kinetex XB-C₁₈色谱柱（2.1 mm×50 mm,2.6 μm）,以0.1%甲酸水-乙腈为流动相,梯度洗脱分离,流速为0.2 mL·min^-1,通过电喷雾离子源,多反应监测（MRM）,正离子模式。结果 通关藤苷A、I、H分别在0.05~10 ng·mL^-1,0.025~10 ng·mL^-1,0.025~10 ng·mL^-1浓度内呈良好的线性关系,r>0.998,检测限为0.012 5~0.025 ng·mL^-1,平均回收率分别为97.9%,95.7%,96.1%,RSD<3.4%。结论 方法简单快速,准确灵敏,可用于消癌平注射液中通关藤苷A、I、H的定量测定。     

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.     

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.     

Alzheimer’s disease – current trends in basic and clinical research. Highlights from the 16th ECNP

Advances in the molecular biological knowledge of neuronal nicotinic acetylcholine receptors (nAChRs) have led to a growing interest by the pharmaceutical industry in the development of novel compounds that selectively modulate nAChR function. The ability of (-)-nicotine, an activator of nAChRs, to enhance attentional aspects of cognition in animals and humans, to exert neuroprotective and anxiolytic-like effects, and presumably to mediate the negative correlation between smoking and Alzheimer's (and Parkinson's) Disease, has focused interest on the potential therapeutic utility of modulators of nAChR function for treatment of some of the deficits associated with these progressive, neurodegenerative conditions. Numerous compounds are known which activate nAChRs and which might serve as lead compounds toward the development of such agents. The pharmacologic diversity of neuronal nAChR subtypes suggests the possibility of developing selective compounds which would have more favourable side-effect profiles than existing agents. This broader class of agents, collectively called cholinergic channel modulators (ChCMs), is anticipated to encompass compounds which would have more favourable side-effect profiles than existing agents, which generally exhibit low selectivity. This selectivity may be achieved by preferentially activating some subtypes of nAChRs (i.e., Cholinergic Channel Activators, ChCAs) or inhibiting the function of other subtypes (Cholinergic Channel Inhibitors, ChCIs). An overview of the biology of nAChRs and the rationale for the use of ChCMs for the treatment of dementia related to neurodegenerative diseases are presented, followed by a discussion of lead compounds and compounds under consideration for clinical evaluation.     

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.