LC-MS/MS法测定人血浆中特比萘芬浓度及其生物等效性评价

目的:建立人血浆中特比萘芬浓度的LC-MS/MS测定方法,并评价特比萘芬的药动学特征及2种制剂的人体生物等效性比较.方法:采用LC-MS/MS法测定特比萘芬的浓度,色谱柱为Diamonsil C18(2)(4.6 mm×150 mm,5 μm),流动相为甲醇-20mmol·L-1醋酸铵溶液(含0.1%甲酸)=80:20,质谱采用MRM模式,检测离子为特比萘芬292.2→140.8,克霉唑为277.2→164.9.20名男性健康受试者随机分成2组,分别交叉给予受试制剂和参比制剂各0.25 g,估算特比萘芬的药动学参数并评价2种制剂的人体生物等效性.结果:人血浆中特比萘芬在2～2 000 ng·ml-1范围内线性关系良好,定量下限为2.0 ng·ml-1,批内及批间精密度RSD＜15%.受试制剂与参比制剂的各主要药动学参数:tmax分别为(1.2±0.6)和(1.7±0.8) h,Cmax分别为(904.4±272.7)和(831.3±307.0) ng·ml-1,t1/2分别为(20.9±3.4)和(27.1±6.6) h,用梯形法计算AUC(0～∞)分别为(3 951±894.7)和(4 764±2 501) ng·h·ml-1.结论:该方法适用于血浆中特比萘酚的检测.结果显示2种制剂生物等效.     

LC-MS/MS法测定健康人血浆中的拉呋替丁

本文建立了一种快速测定人血浆中拉呋替丁血药浓度的LC-MS/MS法,采用CN柱(4.6 mm×150 mm,5μm),柱温30℃,流动相为甲醇-水(含20 mmol.L-1乙酸铵,0.2%甲酸)(97:3);流速1.0 mL.min-1;气动辅助电喷雾离子化(ESI),正离子检测,多反应离子检测(MRM)拉呋替丁和内标氯苯那敏分别为:m/z 432.20→351.15;m/z 275.10→230.05。拉呋替丁在1.98～396 ng.mL-1范围内线性关系良好(r=0.9991),定量限1.98 ng.mL-1。拉呋替丁低、中、高三个浓度批内及批间变异、准确度、绝对回收率均符合方法学要求,无显著基质效应。该方法专属性强,分析周期短,适合于临床上拉呋替丁血浆含量的测定。     

HPLC-MS/MS法测定人血浆中奥昔布宁和去乙基奥昔布宁的质量浓度

目的 建立HPLC-MS/MS法测定人血浆中奥昔布宁和去乙基奥昔布宁的质量浓度.方法 选用Waters-Atlantis dC18色谱柱,以0.01％甲酸溶液-乙腈为流动相进行梯度洗脱,采用正离子多反应监测方式测定样品质量浓度.用于定量分析的离子对分别为m／z358.2→142.2(奥昔布宁),m/z330.2→96.2(去乙基奥昔布宁),m/z268.2→152.2(奥昔布宁D10),m/z 335.2→101.2(去乙基奥昔布宁D5).结果 血浆样品中,奥昔布宁在0.05～25 ng·mL-1线性关系良好(r=0.996 5),最低定量浓度为0.05 ng·mL-1;去乙基奥昔布宁在0.05～25 ng·mL-1线性关系良好(r=0.998 5),最低定量浓度为0.05 ng· mL-1.两者日内与日间RSD均＜15％,提取回收率＞75％,且稳定性均较好.结论 本方法简便快速、灵敏准确、特异性强,适用于奥昔布宁和去乙基奥昔布宁的体内药物动力学研究.     

LC-MS/MS方法测定Beagle犬血浆中比卡鲁胺的浓度

目的 建立灵敏的液相色谱-串联质谱法测定狗血浆中比卡鲁胺的浓度.方法 血浆样品采用乙腈蛋白沉淀方法,以0.2％甲酸-乙腈(35∶65,v/v)为流动相;采用Zorbax SB C18柱(150 mm×4.6mm,5 μm)分离,通过电喷雾电离源,以选择多反应监测(MRM)方式进行负离子检测,用于定量分析的离子反应分别为m/z428.9→254.7(比卡鲁胺)和m/z 269→169.6(内标,甲苯磺丁脲).结果 建立的体内比卡鲁胺测定方法线性范围为5～2 000 ng·mL-1,定量下限可达1 ng·mL-1.日内、日间精密度(RSD)均＜10％.结论 该方法预处理操作简洁、灵敏、专属性强,可方便用于比卡鲁胺药物的体内药动学研究.     

HPLC-/MS/MS法测定健康人血浆中普拉克索浓度

目的 建立高效液相串联质谱法测定人血浆中普拉克索浓度的方法.方法 在碱性条件下,用乙酸乙酯提取浓缩后,进样用LC-MS/MS,固定相为AQ-C18柱(4.6 mm× 150 mm,10 μm),流动相为乙腈-20 mmol·L-1醋酸铵水溶液=60:40,质谱条件为电喷雾离子源、正离子方式、多级离子反应监测,离子反应分别为m/z:212.2→152.9(普拉克索)和m/z 273.2→109.6(吡西卡尼).结果 普拉克索的血浆浓度在5 ～ 1000 pg·mL-1内线性关系良好,Y=1.33×103X+0.05(r=0.9979),定量下限可达5 pg·mL-1.结论 建立的检测方法准确、稳定,可满足血浆中普拉克索含量测定.     

LC-MS/MS法同时测定人血浆中咪达唑仑和 1-羟基咪达唑仑的浓度

目的:建立同时测定人血浆样本中咪达唑仑和1-羟基咪达唑仑浓度的LC-MS/MS方法.方法:以d4-咪达唑仑为内标,血浆样本加入含内标的乙腈处理后进样分析.色谱柱为Kinetex C18(2.1mm×50 mm,2.6μm),流动相为乙腈-1 mmol· L-1甲酸铵溶液(含0.1％甲酸),梯度洗脱,流速为0.3 mL·min-1,进样量为3.0 μL.质谱条件采用电喷雾正离子源,扫描方式为多重反应监测,用于定量分析的检测离子分别为m/z 325.9→291.0(咪达唑仑)、m/z 342.0→324.1(1-羟基咪达唑仑)、m/z 329.9→295.0(d4-咪达唑仑).结果:咪达唑仑和1-羟基咪达唑仑分别在0.300～200 ng· mL-1和0.150～100 ng·mL-1线性关系良好,定量下限分别为0.300 ng· mL-1和0.150ng·mL-1,批内和批间精密度均小于15％,准确度在-4.7％～9.9％,基质效应为95.5％～101.4％,提取回收率为100.6％～102.4％.血浆样本室温放置12h、3次冻融循环及-40℃冰冻12个月稳定性良好,待测溶液进样器放置24h稳定.结论:本研究建立的分析方法灵敏、准确,样本前处理简便、快捷,可用于人血浆样本中咪达唑仑和1-羟基咪达唑仑浓度的测定.     

LC-MS/MS法同时测定人血浆中的辛伐他汀和辛伐他汀酸

目的 采用LC-MS/MS法同时测定人血浆中的辛伐他汀和辛伐他汀酸.方法 采用BEH C18色谱柱(50 mm ×2.1mm,1.7 μm),流动相为乙腈-0.01 mol·L-1醋酸铵(72∶28),流速0.15 mL·min-1,柱温40℃,进样量8μL.辛伐他汀、辛伐他汀酸及内标洛伐他汀的检测离子对分别为:m/z 419.43→199.12、437.38→303.26、405.45→199.14.结果 辛伐他汀、辛伐他汀酸的线性范围分别为0.241 ～61.76 ng·mL-1(r =0.999)、0.344 ～ 88.16 ng·mL-1(r =0.997).在人血浆基质中,高、中、低浓度(0.482、3.86、30.88 ng· mL-1)的日内、日间RSD均小于15％,方法回收率分别为95％～104％、97％ ～108％.样品预处理方法对血浆中的辛伐他汀和辛伐他汀酸测定无干扰.结论 所用方法处理简单、灵敏、特异性高,定量准确,可为辛伐他汀制剂的药动学研究提供方法.     

LC-MS/MS方法同时检测人血浆中异烟肼、乙胺丁醇和吡嗪酰胺的浓度

目的:建立同时检测人血浆中3种抗结核药物异烟肼、乙胺丁醇和吡嗪酰胺浓度的LC-MS/MS方法,用于肺结核患者及临床试验中三药血药浓度的测定。方法:以对乙酰氨基酚为内标,血浆样品经乙腈沉淀蛋白等处理后检测。采用AgilentZORBAX SB-Aq色谱柱(2.1 mm×100 mm,3.5μm)为分析柱,ZORBAX SB-Aq柱(2.1 mm×12.5 mm,5μm)为保护柱,以乙腈-5 mmol·L-1甲酸铵水溶液(含0.1%甲酸)(8:92,v/v)为流动相,使用电喷雾离子源(ESI),以正离子多反应监测(MRM)方式进行检测,异烟肼m/z 138.2→121.0,乙胺丁醇m/z 205.2→116.1,吡嗪酰胺m/z 124.1→81.1,对乙酰氨基酚m/z152.0→110.0。分析时间为5 min。结果:血浆中内源性物质对测定无干扰,异烟肼线性范围为0.1～6.0μg.mL-1,定量下限(LLOQ)为0.1μg.mL-1,乙胺丁醇线性范围为0.1～5.0μg.mL-1,定量下限(LLOQ)为0.1μg.mL-1,吡嗪酰胺线性范围为1.0～50.0μg.mL-1,定量下限(LLOQ)为1.0μg.mL-1。日内、日间精密度(RSD)均小于10%,准确率为90.4%～108.7%。结论:本方法特异性强,灵敏度高,测定结果可靠,适用于临床血浆样品的高通量分析。     

LC-MS/MS法测定人血浆中替利定和去甲替利定的质量浓度

目的:建立LC-MS/MS法测定人血浆中替利定和去甲替利定的质量浓度。方法:选用Agilent-Zorbax-Eclipse-XDB-C18色谱柱,以甲醇-1 mmol·L-1醋酸铵(75∶25)为流动相,采用正离子,多反应监测方式测定样品质量浓度。用于定量分析的离子对分别为[M+H]+m/z 274.3→m/z 155.1(替利定),[M+H]+m/z 260.2→m/z 155.1(去甲替利定)和[M+H]+m/z 284.8→m/z 192.9(地西泮)。结果:血浆样品中,替利定在0.5～250 ng·mL-1范围内线性关系良好(r=0.9936),最低定量质量浓度为0.5 ng·mL-1;去甲替利定在1～500 ng·mL-1范围线性关系良好(r=0.9948),最低定量质量浓度为1 ng·mL-1。二者日内与日间RSD均小于15%,平均回收率高,且稳定性均较好。结论:本方法简便快速、灵敏准确、特异性强,适用于盐酸替利定和去甲替利定的体内药代动力学研究。     

HPLC-MS/MS测定人体血浆和尿液中罗通定的浓度

目的 建立一种简便、灵敏的测定人体血浆和尿液中罗通定浓度的高效液相色谱串联质谱( HPLC-MS/MS)方法.方法 血浆、尿液样品分别采用乙腈沉淀处理后,进样分析.采用Agilent-ECLIPSE-C18柱(2.1mm×150 mm,5μm),以乙腈-0.2％甲酸溶液=85∶15为流动相,采用正离子,多反应监测方式测定样品浓度.检测离子为m/z356.3→192.3(罗通定)和m/z 285.0→193.0(地西泮,内标).结果 罗通定血浆及尿样均在2.5～1000 ng· mL-1与峰面积线性关系良好(r=0.999 4);最低定量浓度均为2.5 ng·mL-1.日内与日间RSD均＜10％,血浆样品回收率在91.4％～109.2％,尿样回收率在88.63％～115.8％.结论 本方法简便快速、灵敏准确,适用于罗通定在人体内的药物动力学研究.     

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.     

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.     

Pharmacological treatment of COVID-19: an opinion paper

The precocity and efficacy of the vaccines developed so far against COVID-19 has been the most significant and saving advance against the pandemic. The development of vaccines has not prevented, during the whole period of the pandemic, the constant search for therapeutic medicines, both among existing drugs with different indications and in the development of new drugs. The Scientific Committee of the COVID-19 of the Illustrious College of Physicians of Madrid wanted to offer an early, simplified and critical approach to these new drugs, to new developments in immunotherapy and to what has been learned from the immune response modulators already known and which have proven effective against the virus, in order to help understand the current situation.     

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.