固相萃取亲水作用色谱法测定人体血浆中表阿霉素

目的:建立固相萃取-亲水作用色谱法(SPE—HILIC)测定人体血浆中的表阿霉素。方法:血浆样品中加入表柔红霉素作为内标,经 Oasis HLB 固相萃取(SPE)小柱萃取后进样测定,色谱柱为 Kromasil KR100—5SIL 硅胶色谱柱(250 mm×4.6mm,5μm),乙腈-甲酸铵缓冲液(40 mmol·mL~(-1),pH 2.9)(90:10)为流动相,检测波长为254 nm。结果:血浆中表阿霉素的线性范围为0.05～2.5μg·mL~(-1)(r~2=0.9991);最低检测限为0.05μg·mL~(-1);样品的回收率高于89.4%;日内及日间精密度RSD 小于7.0%。结论:方法简便,准确可靠,流动相与质谱检测器兼容,适用于血浆表阿霉素浓度测定及药代动力学研究。     

固相萃取高效液相色谱法测定人血浆中硫酸茚地那韦浓度

目的:建立硫酸茚地那韦血药浓度的高效液相色谱分析方法。方法:以对硝基苯胺为内标,血浆样品用固相萃取,色谱柱:Kromasil C_(18)色谱柱(150 mm×4.6mm,5μm),在线过滤器;流动相为0.1%三乙胺水溶液(用20%磷酸调节 pH 为4.3)-乙腈(69.5:30.5);检测波长为210 nm;流速1.0 mL·min~(-1),柱温为25℃。结果:硫酸茚地那韦在0.05～25.60μg·mL~(-1)的范围内有良好的线性关系(r=0.9995),最低检测浓度为0.01μg·mL~(-1)(S/N>3),该方法的相对回收率为97.4%～105.0%(n=5),绝对回收率为93.9%～105.6%(n=5);日内 RSD 为2.6%～3.7%,日间 RSD 为1.4%～4.0%。结论:本方法简便,准确,灵敏,特异性强,重现性好,可用于血浆中硫酸茚地那韦的测定及人体内药代动力学研究。     

HPLC法测定兔血浆中利多卡因和亚甲蓝浓度

目的:建立利多卡因和亚甲蓝同时给药后两药血药浓度的 HPLC 测定方法。方法:血浆样品用氯仿-环己烷-异丙醇(60:30:10)萃取,采用 Shim-pack VP-ODS 分析柱(150 mm×4.6 mm,5μm),以醋酸盐缓冲液-甲醇-乙腈(45:45:10)为流动相,流速1.0 mL·min~(-1),检测波长235 nm,以布比卡因为内标,测定血浆样品中利多卡因;血浆样品以乙腈沉淀蛋白,采用 Sphericorb NH_2分析柱(150 mm×4.6 mm,5 μm),以磷酸盐缓冲液-乙腈(40:60)为流动相;流速1.0 mL·min~(-1),检测波长600 nm,测定血浆样品中亚甲蓝浓度。结果:利多卡因血药浓度测定:线性范围0.16～10.08μg·mL~(-1),绝对回收率大于73%,方法回收率98.49%～107.2%,日内、日间精密度 RSD 均小于8%;亚甲蓝血药浓度测定:线性范围为0.052～3.328μg·mL~(-1),绝对回收率大于72%,方法回收率95.19%～104.3%,日内和日间精密度 RSD 均小于9%。结论:该方法简便、准确,重复性好,可用于利多卡因和亚甲蓝同时给药后两药的家兔体内药代动力学研究。     

固相萃取RP-HPLC法同时测定人全血中艾司洛尔和代谢物对映体

目的:建立人全血中艾司洛尔及其代谢物对映体的测定方法。方法:样品经固相萃取小柱纯化浓缩后,采用手性衍生化试剂2,3,4,6-四-O-乙酰基-β-D-吡喃葡萄糖异硫氰酸酯(GITC)对艾司洛尔和代谢物进行柱前手性衍生化。衍生产物以乙腈-0.01 mol·L~(-1)磷酸二氢钾(pH 4.5)缓冲液(55:45)为流动相,用 Aglient Zorbax C_(18)(4.6mm×25mm,5 μm)色谱柱分离,流速为0.80 mL·min~(-1),检测波长为248 nm。结果:艾司洛尔对映体在0.05～10μg·mL~(-1)范围内线性关系良好,代谢物对映体在0.03～10μg·mL~(-1)范围内线性关系良好。艾司洛尔和代谢物的萃取回收率均大于75%,日内、日间 RSD 均小于15%。结论:该方法结果准确,重现性好,灵敏度高,可同时测定人全血中艾司洛尔及其代谢物对映体。     

大鼠体内米托蒽醌的血药浓度测定及药动学研究

目的:建立测定大鼠体内米托蒽醌血药浓度的方法,研究米托蒽醌在大鼠体内的药动学。方法:取SD大鼠6只,尾静脉注射米托蒽醌5 mg/kg,分别于给药前和给药后5、10、20、40、60、120、240、480、720 min取尾静脉血0.3 mL,置于肝素化EP管中,离心分离血浆,加入硅胶充分研磨混匀后,再加入含0.5 mol/L盐酸的甲醇溶液沉淀蛋白,研磨混匀,离心取上清液,氮气吹干后加流动相复溶。采用高效液相色谱法测定米托蒽醌的血药浓度,色谱柱为ZORBAX SB-C_(18),流动相为20 mmoL/L乙酸铵水溶液(用稀盐酸调pH至2.0)-甲醇(65∶35,V/V),流速为1.0 mL/min,检测波长为244 nm,柱温为30℃,进样量为20μL,应用DAS 3.0软件计算药动学参数。结果:米托蒽醌检测质量浓度的线性范围为200～10 000μg/L(r=0.999 6,n=6),定量下限为200μg/L,检测限为150μg/L;日内、日间精密度和稳定性试验的RSD均<8.0%(n分别为5、3、6);提取回收率为(85.64±3.93)%～(92.31±1.68)%(n=3);准确度试验中的回收率为(93.58±1.42)%～(113.92±2.74)%(n=6)。米托蒽醌在大鼠体内的药动学参数AUC0-720 min为(5 247.1±474.6)μg·h/L,t_(1/2z)为(24.88±6.94)h,CLZ为(0.46±0.09)L/(h·kg),Vz为(11.07±2.64)L/kg。结论:该方法提取回收率高、重复性好,适用于米托蒽醌血药浓度的测定和药动学研究。     

HPLC法同时测定人体血浆中氟尿嘧啶及其前体药替加氟的浓度

目的:建立同时测定氟尿嘧啶及其前体药替加氟在人体血浆中浓度的 HPLC 方法。方法:以阿昔洛韦为内标,血浆样品用硝酸银(10%)沉淀蛋白质,采用 Zorbax-ODS C_(18)分析柱(4.6 mm×250 mm,5μm),检测波长为265 nm,流动相:0.01 mol·L~(-1)磷酸盐(用2 mol·L~(-1)氢氧化钠溶液调节 pH 为8.0)-甲醇(97:3),流速1.0 mL·min~(-1),柱温为45℃。结果:氟尿嘧啶、替加氟分别在0.1～20μg·mL~(-1)(r=0.9998)和0.2～40μg·mL~(-1)(r=0.9998)浓度范围内线性关系良好,最低检测浓度分别为5和10 ng·mL~(-1),方法回收率分别为97.2%～101.3%和98.3%～102.6%,日内、日间 RSD 小于6%。结论:方法灵敏、快速、准确,并适用于临床上氟尿嘧啶及前体药替加氟浓度监测。     

液相色谱-电喷雾串联质谱法测定人血浆中辛伐他汀浓度

目的:建立液相色谱-电喷雾串联质谱联用测定人血浆中辛伐他汀浓度的方法。方法:色谱条件为 Discovery ODS C_(18)(2.1 mm×100 mm,3μm)色谱柱;流动相为乙腈-0.1%甲酸溶液(65:35,v/v);柱温30℃;流速0.2 mL·min~(-1);进样量5μL。质谱条件为电喷雾电离源(ESI),选择性检测定量离子为 m/z 441.3/325.0(辛伐他汀)和 m/z 427.2/325.0(洛伐他汀,内标)带正电荷的离子峰。样品用乙酸乙酯提取。结果:血浆中辛伐他汀浓度在0.20～20.0 ng·mL~(-1)内呈良好线性关系,r=0.9996。提取回收率为90.0%～92.4%(RSD 6.0%～14.9%),方法回收率在85%～115%之内,日内和日间精密度试验的RSD 均<15%,最低检测浓度为0.2 ng·mL~(-1)。结论:该测定方法经全面考察符合血浆样品的测定要求,可以应用于辛伐他汀的人体药动学研究和生物等效性评价。     

固相萃取HPLC测定人血浆中头孢西酮及在药动学研究中的应用

目的:建立反相高效液相色谱办法测定人血浆中头孢西酮浓度,并用于注射用头孢西酮钠人体药代动力学研究。方法:采用高效液相色谱紫外检测法,血浆中加入内标后经固相萃取,色谱柱为 Apollo C_(18)(5μm,250 mm×4.6 mm)。流动相为乙腈-0.02 mol·L~(-1)醋酸铵溶液(18∶82)(pH 5.0),流速1 mL·min~(-1),检测波长为278 nm。结果:本方法线性检测范围为0.5～250μg·mL~(-1),线性关系良好(r=0.9996);最低检测浓度为0.5μg·mL~(-1);方法绝对回收率为67.2%～84.0%,相对回收率为91.1%～102.1%;日内、日间 RSD 均小于8%。结论:本方法灵敏度高,操作简便,可用于人血浆中头孢西酮的浓度测定及临床药代动力学研究。     

液-液萃取RP—HPLC法测定人血浆中佐米曲普坦的浓度

目的:建立RP-HPLC法测定人血浆中佐米曲普坦浓度。方法:以替硝唑为内标,血浆样品经液-液萃取前处理后,选用Hypersil C_(18)色谱柱(5μm,200mm×5.0mm),乙腈-磷酸盐缓冲液(0.05mol·L~(-1)磷酸二氢钠,用磷酸调pH至5.0)(15∶85)为流动相,室温下在223nm处进行测定。结果:测定佐米曲普坦血药浓度线性范围为1.92-66.14ng·mL~(-1),最低定量浓度1.92ng·mL~(-1);方法回收率为90.62%-105.7%,萃取回收率为71.70%-114.75%。结论:本法分离效果良好,灵敏度高,回收率高,日内、日间误差小,能满足于人体药代动力学及生物利用度研究。     

HPLC法同时测定肿痛安胶囊中5种成分的含量

目的建立同时测定肿痛安胶囊中:天麻素、升麻素苷、5-O-甲基维斯阿米醇苷、欧前胡素和异欧前胡素含量的HPLC法。方法色谱柱:Phenomenex Luna C_(18)柱(250mm×4.6mm,5μm);流速:1.0mL·min~(-1);柱温:25℃;流动相:甲醇-水梯度洗脱;检测波长:230nm;进样量:5μL。结果天麻素质量浓度在12.46～199.30μg·mL~(-1)范围内线性关系良好,r_1=0.999 5,平均回收率为101.2%,RSD值为1.6%(n=6);升麻素苷质量浓度在12.07～193.20μg·mL~(-1)范围内线性关系良好,r_2=0.999 7,平均回收率为100.1%,RSD值为1.8%(n=6);5-O-甲基维斯阿米醇苷质量浓度在8.12～129.90μg·mL~(-1)范围内线性关系良好,r_3=0.999 8,平均回收率为99.1%,RSD值为1.7%(n=6);欧前胡素质量浓度在12.52～200.40μg·mL~(-1)范围内线性关系良好,r_4=0.999 6,平均回收率为98.6%,RSD值为1.4%(n=6);异欧前胡素质量浓度在12.65～202.40μg·mL~(-1)范围内线性关系良好,r_5=0.999 6,平均回收率为99.0%,RSD值为0.7%(n=6)。结论该方法简单、准确,可同时测定5种成分的含量,可用于肿痛安胶囊的质量控制。     

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.