高效液相色谱法测定大鼠血浆及组织中多柔比星含量

目的：建立高效液相色谱法测定大鼠血浆及组织样品中多柔比星的含量,并研究其在大鼠体内的分布特征。方法：生物样品经过液-液萃取后,采用HPLC-FLD法,色谱柱为Zorbax SB-C1（8150mm×4.6mm,5μm）,流动相为乙腈和0.01mol·L-1磷酸二氢钾缓冲液（pH2.3）,梯度洗脱,流速为1mL·min-1,柱温为35℃;荧光检测器,激发波长为237nm,发射波长为555nm。结果：该法测定血浆中多柔比星的线性范围为2.5-500ng·mL-1,r=0.9991。其他组织的分析方法也均满足生物样品的测定要求。静脉给予5mg·kg-1多柔比星,结果显示其在大鼠体内分布广泛且持久,肾脏和脾脏及毒性靶器官心脏中浓度较高。结论：该方法准确度、灵敏度高,可以测定72h内多柔比星在主要组织中的分布,可用于研究心脏毒性与多柔比星＂量＂之间相关性。     

Development and validation of rapid and sensitive HPLC method for the quantitative determination of doxorubicin in human plasma

An isocratic reversed-phase high-performance liquid chromatographic method with ultraviolet detection at 254?nm has been developed for the determination of doxorubicin in human plasma. Plasma samples were extracted by a selective one-step liquid–liquid extraction using dichloromethane. Doxorubicin and the internal standard epirubicin were separated using a column packed with C₁₈ material, using a mobile phase consisting of water:acetonitrile (75:25v/v). The calibration graph for doxorubicin was linear in the range 0.2–10?μg/mL, with a correlation coefficient, R²?=?0.9986. Lower limit of quantitation was 0.2?μg/mL, using 1?mL plasma samples. The extraction recovery ranged from 98.5–101.1%, and the recovery rate was consistent for drug and internal standard examined at each level. The interday and intraday precision values ranged between 0.5–2%. Validation data showed that the assay for doxorubicin is sensitive, selective, accurate, and reproducible. The assay has been used in population pharmacokinetics study.     

Indirect determination of paracetamol in pharmaceutical formulations by inhibition of the system luminol-H2O2-Fe(CN)3-(6) chemiluminescence

After a large drug scanning, the system Luminol-H2O2-Fe(CN)6(3-) is proposed for first time for the indirect determination of paracetamol. The method is based on the oxidation of paracetamol by hexacyanoferrate (III) and the subsequent inhibitory effect on the reaction between luminol and hydrogen peroxide. The procedure resulted in a linear calibration graph over the range 2.5-12.5 microg ml(-1) of paracetamol with a sample throughput of 87 samples h(-1). The influence of foreign compounds was studied and, the method was applied to determination of the drug in three different pharmaceutical formulations.     

RP-HPLC测定大鼠血浆中的水飞蓟宾

目的 采用RP-HPLC法测定大鼠血浆中的水飞蓟宾.方法 以穿心莲内酯为内标,采用Kromaasil C_(18)色谱柱(150mm ×4.6 mm,5μm),流动相为乙腈-水(35:65,10%磷酸调pH4.5),流速1.0 mL·min~(-1),检测波长288 nm,柱温35℃,进样量为20μL.结果 水飞蓟宾0.5～100.0 μg·mL~(-1)与内标峰面积比的线性关系良好(r~2=0.9999);高、中、低浓度的平均回收率分别为100.04%、99.82%、98.76%;日内和日间RSD均小于15%.结论 所建方法简单、灵敏、准确可靠,可用于水飞蓟宾的药物动力学研究.     

A simple and sensitive gradient elution liquid chromatography with tandem mass spectrometry (LC-MS/MS) method for the quantification of doxorubicin in rabbit plasma

In the present study, we developed and validated a simple and sensitive gradient elution liquid chromatography tandem mass spectrometry (LC-MS/MS) method for quantification of doxorubicin in rabbit plasma. Daunorubicin was used as an internal standard (IS). The doxorubicin and IS were extracted with ethyl acetate from plasma samples. The chromatographic separations were achieved on a C₁₈ column (2.1 mm×50 mm, 2.5 μm) configured with a C₁₈ guard column (2.1 mm×10 mm, 2.5 μm). The mobile phase of 0.1% formic acid-water solution and acetonitrile was delivered using a gradient elution program at a flow rate of 0.4 mL/min. The temperature for column was maintained at 40 ºC. The electrospray ionization (ESI) source was operated in the positive ion mode, and the quantification was conducted using multiple reaction monitoring (MRM) of the transitions m/z 544.07→396.96 and m/z 528.06→321.05 for doxorubicin and IS, respectively. The calibration curve of doxorubicin was linear (r > 0.999) within the range of 2-600 ng/mL. The lower limit of quantification was 2 ng/mL. The relative errors of intra­day and inter-day accuracies ranged from -2.48% to 0.18% and from -3.78% to 1.94%, respectively. The relative standard deviations of intra­day and inter-day precisions were less than 8.65% and 6.41%, respectively. The method exhibited satisfactory results in terms of specificity, sensitivity, matrix effect, recovery and stability. The newly developed LC-MS/MS method was reliable to monitor doxorubicin concentrations in rabbit plasma.     

Determination and pharmacokinetics of DT-13 in rat plasma by LC-MS

A sensitive and specific LC-MS assay for DT-13 in rat plasma was developed. DT-13 is an active steroidal saponin present in Liriopes Radix and is developed as an anti-tumor drug candidate. The samples were extracted by acetonitrile-mediated plasma protein precipitation. The chromatographic separation was carried out using a Ultimate C₁₈ column (250 mm × 4.6 mm, i.d., 5 μm) with a mobile phase composed of acetonitrile: 5 mmol/L aqueous ammonium acetate (60:40, v:v). The method was validated and the specificity, linearity (r² = 0.9980 within 10-1000 ng/mL), lower limit of quantitation (LLOQ, 10 ng/mL), precision (intra- and inter-day <12.3%), accuracy (93.4-106.3%), recovery (91.0 ± 4.7%) and stability were determined. The method was applied to the pharmacokinetic study of DT-13 in rat plasma after intravenous and intragastric administration. The results showed DT-13 underwent a prolonged absorption and slow elimination with a low oral bioavailability (5.51%) in rats.     

液相色谱-串联质谱法测定大鼠血浆中左旋泮托拉唑钠的浓度及其毒代动力学研究

目的 建立液相色谱-串联质谱法测定大鼠血浆中左旋泮托拉唑钠的浓度并研究其毒代动力学.方法 大鼠血浆样本以乙腈沉淀蛋白后,经Chiralcel OJ-RH色谱柱(4.6 mm×150 mm,5 μm);流动相为乙腈∶水=28∶72,流速为0.6 mL·min-1,柱温为35 ℃;采用Agilent 6430三重四极杆串联质谱仪,离子化方式:电喷雾-正离子(API-ES),监测方式:多反应监测,左旋泮托拉唑监测离子对384.0/199.8,非那西丁监测离子对180.0/110.0,用作内标.结果 本法专属性良好;左旋泮托拉唑钠在50～30 000 ng·mL-1范围内线性关系符合要求;准确度均在85%～115%,精密度在15%内.SD大鼠连续4周静脉注射给予注射用左旋泮托拉唑钠低、中、高3个剂量,左旋泮托拉唑钠动力学参数AUC0～4 h增长与剂量增长均呈正相关.首次和4周给药后测定无明显差异.结论 本法经过方法学验证,适用于大鼠血浆中左旋泮托拉唑钠浓度的测定,可用于左旋泮托拉唑钠大鼠体内毒代动力学研究.     

Early metabolomics changes in heart and plasma during chronic doxorubicin treatment in B6C3F1 mice

The present study aimed to identify molecular markers of early stages of cardiotoxicity induced by a potent chemotherapeutic agent, doxorubicin (DOX). Male B6C3F₁ mice were dosed with 3 mg kg^?1 DOX or saline via tail vein weekly for 2, 3, 4, 6 or 8 weeks (cumulative DOX doses of 6, 9, 12, 18 or 24 mg kg^?1, respectively) and euthanized a week after the last dose. Mass spectrometry‐based and nuclear magnetic resonance spectrometry‐based metabolic profiling were employed to identify initial biomarkers of cardiotoxicity before myocardial injury and cardiac pathology, which were not noted until after the 18 and 24 mg kg^?1 cumulative doses, respectively. After a cumulative dose of 6 mg kg^?1, 18 amino acids and four biogenic amines (acetylornithine, kynurenine, putrescine and serotonin) were significantly increased in cardiac tissue; 16 amino acids and two biogenic amines (acetylornithine and hydroxyproline) were significantly altered in plasma. In addition, 16 acylcarnitines were significantly increased in plasma and five were significantly decreased in cardiac tissue compared to saline‐treated controls. Plasma lactate and succinate, involved in the Krebs cycle, were significantly altered after a cumulative dose of 6 mg kg^?1. A few metabolites remained altered at higher cumulative DOX doses, which could partly indicate a transition from injury processes at 2 weeks to repair processes with additional injury happening concurrently before myocardial injury at 8 weeks. These altered metabolic profiles in mouse heart and plasma during the initial stages of injury progression due to DOX treatment may suggest these metabolites as candidate early biomarkers of cardiotoxicity. Published 2016. This article is a U.S. Government work and is in the public domain in the USA     

铁氰化钾—鲁米诺化学发光体系测定吩噻嗪类药物

目的:确定以铁氰化钾-鲁米诺化学发光体系分析测定吩噻嗪类药物的新方法。方法:在强碱性条件下,盐酸异丙嗪和盐酸氢丙嗪对铁氰化钾-鲁米诺体系的发光有强烈的增强作用。从而试验以流动注射化学发光法分析测定盐酸异丙嗪和盐酸氯丙嗪。结果:在优化的实验条件下,化学发光强度在一定范围内与上述药物浓度成正比。对盐酸异丙嗪的检出限为3.0ng·mL~(-1);对盐酸氢丙嗪的检出限为5.0ng·mL~(-1)。结论:本方法快捷、简便且具有很高的灵敏度,本法用于相应药片中的上述药物的分析,并与药典规定的方法进行对照,结果令人满意。     

Determination of Huperzine A in rat plasma by high-performance liquid chromatography with a fluorescence detector

A simple reversed-phase high-performance liquid chromatography (HPLC)-fluorescence method for the determination of Huperzine A in rat plasma was developed and validated. Separation was achieved on Kromasil C(8) column (5 microm, 150 mm x 4.6mm i.d.). The mobile phase, methanol-water-triethanol amine (45:55:0.05, v/v/v), was delivered at a flow rate of 1.0 ml/min. The eluent was monitored by a fluorescence detector with excitation wavelength at 310 nm and emission wavelength at 370 nm. No interfering peaks were observed in rat blank plasma. The relationship between Huperzine A concentration and peak-area ratio of Huperzine A to the IS was linear over the range of 2.5-250 ng/ml. The intra- and inter-day coefficients of variation were 相似文献   

Determination of limonin in rat plasma by liquid chromatography-electrospray mass spectrometry

A simple, sensitive and selective liquid chromatography coupled with electrospray ionization mass spectrometry (LC/ESI/MS) method for determination of limonin (LM) in rat plasma has been developed and validated. The method had advantages of a single liquid–liquid extraction with ether and high sensitivity. Analyses were conducted at a flow rate of 0.2 ml/min by a gradient elution. The detection utilized selected ion monitoring in the negative ion mode at m/z 460.00 and 423.15 for the deprotonated molecular ions of LM and the internal standard, respectively. The quantitation limit for LM in rat plasma was 1.0 ng/ml. The linearity was also excellent over the concentration range of 1.9–500 ng/ml of LM. The intra- and inter-day precision (relative standard deviation (R.S.D.%)) was lower than 10% and accuracy ranged from 90 to 110%, showing a good reproducibility. This developed method was successfully applied to analysis of LM in biological fluids.     

HPLC荧光和质谱两种检测器测定大鼠血浆及组织中的表阿霉素

目的 建立测定大鼠血浆及组织中表阿霉素浓度的方法.方法 采用RP-HPLC荧光和质谱两种检测器,Lima C18(2)分析柱(150 mm×4.6 mm,5μm),流动相为0.01 mol·L-1乙酸铵-乙腈(60:40,甲酸调pH3.5),流速0.6 ml·min-1,样品在碱性条件下,用乙酸乙酯漩涡混合提取浓集后进样,λEx=450 am,λEm=530 nm,质谱以MRM模式测定,内标法定量.结果 荧光检测标准曲线在2.44～2.50×103μg·L-1有良好线性,定量限2.44μg·L-1,日内RSD小于5.0%,日间RSD小于8.6%,方法回收率99%～113%,萃取回收率86.8%～89.7%;质谱检测标准曲线在0.49～2.00×103μg·L-1有良好线性,定量限0.49μg·L-1,日内RSD小于6.5%,日间RSD小于8.7%,方法回收率98～5%.结论 所建方法快速简便、灵敏准确,适用于血浆及组织中表阿霉素的测定及药物动力学研究.     

Determination of deoxyschizandrin in rat plasma by LC-MS

A simple, rapid and sensitive LC-MS method was developed for quantification of deoxyschizandrin in rat plasma. A 50 miccrol plasma sample was extracted by ether and performed on Elite Hypersil C(18) column (200 mm x 4.6 mm, 5 microm) with the mobile phase of methanol-water (84:16, v/v) in a run time of 6.5 min. The analyte was monitored with positive atmospheric pressure chemical ionization (APCI) by selected ion monitoring (SIM) mode. A good linear relationship was obtained over the range of 1.0-50.0 ng/ml and the validated method was successfully applied for the pharmacokinetic studies of deoxyschizandrin in rat. After oral administration of 4 mg/kg deoxyschizandrin and Schisandra extract which contained the same dose of deoxyschizandrin to male rats, the C(max) of deoxyschizandrin were 15.8+/-3.1 and 34.3+/-16.8 ng/ml, T(max) were 0.51+/-0.13 and 3.83+/-1.83 h, T(1/2) were 5.3+/-2.2 and 6.5+/-3.4h.     

流动注射化学发光法测定药物制剂中的诺氟沙星含量

目的建立药物制剂中诺氟沙星的流动注射化学发光测定法。方法选择氢氧化钠溶液为介质,选择鲁米诺,高碘酸钾为反应剂,采用流动注射化学发光法测定诺氟沙星。结果选择鲁米诺浓度5．0×10-5mol／L,高碘酸钾浓度6．0×10mol／L,可在试剂消耗最少的情况下得到最强发光信号。诺氟沙星浓度与化学发光信号范围线性为5．0×10-8～2．0×10-6mol／L,检出限为1．8×10mg／L,回收率96．0％～101．5％,相对标准偏差1．59％～2．04％。结论该方法简便快捷、灵敏度高,可用于药物制剂中诺氟沙星的含量检测。     

多柔比星纳米粒在大鼠体内的靶向性分布研究

目的 研究多柔比星纳米粒（NDXRB）经肝动脉给药后在大鼠体内靶向性分布。方法 SD大鼠30只,随机分为两组,每组各15只,经肝动脉按2mg/kg的剂量分别注入NDXRB或DXRB水溶液,于给药后的1、5、15h各处死5只大,分别提取心、肝、脾、肺、肾和血浆样品,以高效液相色谱法测定DXRB的浓度。结果15h以内NDXRB组大鼠肝和脾中DXRB浓度均极显著高于DXRB组（P＜0．01）,而血浆、心和肺中DXRB浓度极显著低于DXRB组（P＜0．01）。肾组织中DXRB组浓度在5h以内显著高于NDXRB组（P＜0．05）,15h时两组间无显著性差异（P＞0．05）。各时间点均以心脏药物浓度为最低。结论 NDXRB肝动脉给药后改变了DXRB的体内分布特征,在对肝脏和脾脏表现出显著靶向性的同时在心脏的分布显著减少。     

Determination of a novel low-voltage-activated calcium channel blocker (HYP-10) in rat plasma by liquid chromatography-mass spectrometry

A novel T-type calcium channel blocker, 4-amino-1-{4-[(4-chloro-phenyl)-phenyl-methyl]-piperazin-1-yl}-butan-1-one (HYP-10) has been synthesized, and the compound has shown promise as both a nociceptive and inflammatory pain reliever as well as an analgesic in a rat neuropathic pain model. A quantification method was developed for the determination of HYP-10 in rat plasma. After simple protein precipitation with methanol, HYP-10 and the internal standard, methaqualone were chromatographed on a reversed-phase column and detected by liquid chromatography/tandem mass spectrometry with electrospray ionization. The accuracy and precision of the assay were in accordance with FDA regulations for validation of bioanalytical methods. This method was applied to measure the plasma HYP-10 concentration after a single intravenous administration of the compound in rats.     

HPLC-MS/MS determination of a peptide conjugate prodrug of doxorubicin, and its active metabolites, leucine-doxorubicin and doxorubicin, in dog and rat plasma

A HPLC-MS/MS Electrospray (ESI) method was developed and validated to quantify a peptide conjugate prodrug of doxorubicin (Dox-Con) and its active metabolites leucine-doxorubicin (Leu-Dox) and doxorubicin (Dox) in dog and rat plasma. The analytes were extracted from plasma by solid-phase extraction on a Bond Elut® C8 cartridge and eluted with chloroform–methanol (2:1). Eluates were evaporated and reconstituted in acetonitrile–5 μM sodium trifluoroacetate in 0.1% aqueous formic acid (20:80) and injected onto a Waters Oasis® HLB column. Analytes were eluted from the column with a solvent gradient into the mass analyzer. The ions were quantified in the selected reaction-monitoring mode (SRM), using positive ions, on a triple quadrupole mass spectrometer. The lower limits of quantification for Dox-Con, Leu-Dox, and Dox in plasma, were approximately 5, 1 (dog)/6 (rat), and 0.5 ng/ml, respectively. Intra- and inter-assay accuracy (% of nominal concentration) and precision (%CV) for all analytes were within 15 and 16%, respectively.     

HPLC-electrospray mass spectrometric assay for the determination of (R,R)-fenoterol in rat plasma

A fast and specific liquid chromatography-mass spectrometry method for the determination of (R,R)-fenoterol ((R,R)-Fen) in rat plasma has been developed and validated. (R,R)-Fen was extracted from 125 microl of plasma using solid phase extraction and analyzed on Atlantis HILIC Silica 3 microm column. The mobile phase was composed of acetonitrile:ammonium acetate (pH 4.1; 20mM) (85:15, v/v), at a flow rate of 0.2 ml/min. The lower limit of detection (LLOD) was 2 ng/ml . The procedure was validated and applied to the analysis of plasma samples from rats previously administered (R,R)-Fen in an intravenous bolus.     

RP-HPLC法测定兔眼各组织中多柔比星的浓度

目的建立测定兔眼各组织中多柔比星浓度的RP-HPLC法。方法采用RP-HPLC法,色谱柱为DiamonsilC18柱(250mm×4.6mm,5μm),流动相为乙腈∶0.2mol/L磷酸二氢钾缓冲液∶三乙胺(30∶70∶0.2),磷酸调pH到4.0;检测波长234nm;内标为柔红霉素;甲醇沉淀蛋白,固相萃取富集。结果在兔眼各组织中,多柔比星和内标保留时间合适,分离效果良好;在0.1~10μg/ml的浓度范围内线性关系良好,r>0.999;各组织中日内与日间精密度分别小于3.70%和3.39%,低、中、高浓度,各组织回收率接近100%,提取率、定量限和检测限均符合要求。结论所建立的RP-HPLC法测定兔眼各组织中多柔比星的含量准确可靠。     

HPLC测定大鼠血浆中的山奈酚

目的建立测定大鼠血浆中山奈酚的方法。方法以黄芩素为内标,无水乙醚为萃取溶剂,采用HPLC法,Shimm-pack VP-ODS柱为分析柱,乙腈-0.5%冰醋酸(33.3:66.7)为流动相,流速为1.0 ml.min-1,检测波长为370 nm,柱温为40℃。结果山奈酚0.050~5.301μg.ml-1与峰面积比(山奈酚/内标)的线性关系良好,最低定量限为0.020μg.ml-1,4种浓度的平均回收率为96.48%~108.33%,日内RSD≤7.39%,日间RSD≤5.88%,3种浓度的平均萃取回收率为93.67%、98.37%、101.83%。结论所建方法简单、灵敏、准确可靠,可用于山奈酚的药物动力学研究。