LC/MS/MS法测定人血浆中头孢克肟含量

目的 建立人血浆中头孢克肟的LC/MS/MS法.方法 血浆样品经蛋白沉淀提取,采用C8色谱柱分析,以乙腈:水:甲酸(40:60:0.5,v/v/v)为流动相,三重四级杆质谱检测器,正离子多反应监测模式(MRM),监测离子分别为:m/z 454.3→m/z 285.2(头孢克肟),m/z 282.2→m/z 212.2(...     

New ultra-performance liquid chromatography-tandem mass spectrometry method for the determination of irbesartan in human plasma

With the objective of reducing analysis time and maintaining good efficiency, there has been substantial focus on high-speed chromatographic separations and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) is a preeminent analytical tool for rapid biomedical analysis. In this study a simple, rapid, sensitive, and specific ultra-performance liquid chromatography-MS/MS method was developed and validated for quantification of the angiotensin II receptor antagonist, irbesartan (IRB), in human plasma. After a simple protein precipitation using methanol and acetonitrile, IRB and internal standard (IS) telmisartan were separated on Acquity UPLC BEH C18 column (50 mm × 2.1 mm, i.d. 1.7 μm, Waters, Milford, MA, USA) using a mobile phase consisted of acetonitrile: methanol: 10 mM ammonium acetate (70: 15: 15 v/v/v) with a flow rate of 0.4 mL/min and detected MS/MS in negative ion mode. The ion transitions recorded in multiple reaction monitoring mode were m/z 427.2→193.08 for IRB and m/z 513.2→469.3 for IS. The assay exhibited a linear dynamic range of 2–500 ng/mL for IRB in human plasma with good correlation coefficient of (0.995) and with a lower limit of quantitation of 2 ng/mL. The intra- and interassay precisions were satisfactory; the relative standard deviations did not exceed 9.91%. The proposed UPLC-MS/MS method is simple, rapid, and highly sensitive, and hence it could be reliable for pharmacokinetic and toxicokinetic study in both animals and humans.     

A rapid and sensitive liquid chromatography-tandem mass spectrometry method for determination of olopatadine concentration in human plasma

A sensitive and rapid method based on liquid chromatography- tandem mass spectrometry (MS-MS) was developed for the determination of olopatadine in human plasma. Sample preparations were carried out by protein precipitation with the addition of acetonitrile followed by liquid-liquid extraction with ethyl acetate/dichloromethane after internal standard (IS, amitriptyline) spiked. After evaporation to dryness, the resultant residue was reconstituted in mobile phase. Separation of olopatadine and IS from the interferences was achieved on a C(18) column followed by MS-MS detection. The analytes were monitored in the positive ionization mode with a TurboIonspray source. Multiple reaction monitoring using the transition of m/z 338 → 165 and m/z 278 → 91 was performed to quantify olopatadine and IS, respectively. The method had a total chromatographic run time of 3.5 min and linear calibration curves over the concentration range of 0.2-100 ng/mL. The lower limit of quantification was 0.2 ng/mL. For each QC concentration level the intra- and interday precisions were less than 11.4%, and relative errors ranged between -6.40% and 9.26%. The validated method was successfully applied to the quantification of olopatadine concentration in human plasma after administration of olopatadine at an oral dose of 5 mg in order to evaluate the pharmacokinetics.     

Development and validation of a liquid chromatography-tandem mass spectrometry method for the determination of zofenopril and its active metabolite zofenoprilat in human plasma

A novel, sensitive and rapid liquid chromatographic-electrospray ionization mass spectrometric method was developed and validated for the determination of zofenopril and its active metabolite zofenoprilat in human plasma. The method was based on a single extraction step using methyl tert-butyl ether and did not require chemical derivatization. The chromatographic conditions were optimized; separation was performed on a phenyl-hexyl column (5μm, 250mm×4.6mm i.d.) with a mobile phase consisting of a solution of methanol and water (95:5, v/v) that also contained 0.1% of formic acid. A flow rate of 1.0mL/min was used. Zofenopril, zofenoprilat and the internal standard (IS) fosinopril sodium were measured using an electrospray ion source in a positive reaction monitoring mode. Linear calibration curves were generated for zofenopril concentrations between 0.1052 and 1052ng/mL and for zofenoprilat concentrations between 0.2508 and 2508ng/mL. In both cases, the coefficients of determination were greater than 0.995. The extraction recovery for zofenopril was 93.5% on average. It was 92.5% for zofenoprilat. The inter- and intra-batch precision and accuracy for both zofenopril and zofenoprilat were higher than 14%. The method was applied to measure the concentrations of zofenopril and zofenoprilat in plasma samples.     

Quantitative determination of spinosin in rat plasma by liquid chromatography-tandem mass spectrometry method

A sensitive method for the quantitative determination of spinosin in rat plasma was developed and validated using high-performance liquid chromatographic separation with tandem mass spectrometric detection. The analytes of interest were extracted from rat plasma samples by methyl tert-butyl ether (MTBE) after acidification with 1.0% acetic acid aqueous solution. Chromatographic separation was achieved on an Agilent Zorbax SB-C(18) (50 mm x 4.6 mm, 5 microm) using a isocratic mobile phase consisting of acetonitrile-water (30:70, v/v) with 1% isopropyl alcohol and 0.01% heptafluorobutyric acid. The flow rate was 0.2 ml/min. The column temperature was maintained at 25 degrees C. Detection was performed on a triple quadrupole tandem mass spectrometer by selected reaction monitoring (SRM) mode via electrospray ionization (ESI). The calibration curve was linear over the range of 1.00-400 ng/ml in rat plasma, with 1.00 ng/ml of the lower limit of quantification (LLOQ). The inter- and intra-day precisions and accuracy for all samples were satisfactory. The validated method was successfully applied for the pharmacokinetic study of spinosin in rat. After oral administration of 20mg/kg spinosin to rats, the main pharmacokinetic parameters of T(max), C(max), T(0.5) and AUC(0-T) were 5.33+/-0.58 h, 132.2+/-10.6 ng/ml, 4.89+/-0.37 h, 1.02+/-0.09 microg h/l, respectively.     

Simultaneous determination of sildenafil and its active metabolite UK-103,320 in human plasma using liquid chromatography-tandem mass spectrometry

A liquid chromatography-tandem mass spectrometric method for the simultaneous determination of sildenafil and its active N-demethylated metabolite, UK-103,320 in human plasma was developed. Sildenafil, UK-103,320 and the internal standard (DA-8159) were extracted from human plasma with dichloromethane at basic pH. A reverse-phase LC separation was performed on Luna phenylhexyl column with the mixture of acetonitrile-ammonium formate (10 mM, pH 6.0) (60:40, v/v) as mobile phase. The detection of analytes was performed using an electrospray ionization tandem mass spectrometry in the multiple reaction-monitoring mode. The lower limits of quantification for sildenafil and UK-103,320 were 2.0 ng/ml. The method showed a satisfactory sensitivity, precision, accuracy, recovery and selectivity.     

Ultra-performance liquid chromatography-tandem mass spectrometry method for the analysis of amphetamines in plasma

A fast and selective ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) method for the determination of amphetamines (amphetamine, methamphetamine, methylenedioxymethamphetamine, methylenedioxyamphetamine, methylenedioxyethylamphetamine, ephedrine, and p-methoxyamphetamine) in plasma has been developed and validated. Sample preparation was performed by liquid-liquid extraction using ethyl acetate. For optimized chromatographic performance with repeatable retention times, narrow and symmetrical peaks, and focusing all analytes at the column inlet, a gradient start, with acid mobile phase consisting of 0.1% formic acid and methanol was chosen. Positive electrospray ionization MS-MS detection was performed with two multiple reaction monitoring transitions for each analyte. Deuteriumlabeled internal standards were used for five of the analytes. The limit of detection was in the range 0.25-1.25 ng/mL, and the limit of quantification was fixed at the lowest calibrator of 2.5 ng/mL for all of the compounds. The RSD values of the intra- and interassay precision and accuracy were lower than 11% at four concentration levels, including two external quality controls. No or only minor matrix effects were observed, and the extraction method presented recoveries higher than 93% for all the compounds. Total run time, including equilibration, was 12 min. The method is routinely used at the National Institute of Criminalistics and Criminology for quantitative determination of the main amphetamines in plasma from forensic and driving under the influence cases.     

Hydrophilic interaction liquid chromatography-tandem mass spectrometry for the determination of levofloxacin in human plasma

A rapid, sensitive and selective hydrophilic interaction liquid chromatography-tandem mass spectrometric (HILIC-MS/MS) method for the determination of levofloxacin in human plasma was developed. Levofloxacin and ciprofloxacin (internal standard) were extracted from human plasma with dichloromethane and analyzed on an Atlantis HILIC Silica column with the mobile phase of acetonitrile-ammonium formate (100 mM, pH 6.5) (82:18 v/v). The analytes were detected using an electrospray ionization tandem mass spectrometry in the multiple-reaction-monitoring mode. The standard curve was linear (r>0.999) over the concentration range of 10.0-5000 ng/ml. The lower limit of quantification for levofloxacin was 10.0 ng/ml using 20 microl plasma sample. The coefficient of variation and relative error for intra- and inter-assay at four QC levels were 2.9-7.8% and -7.3% to -2.2%, respectively. The recoveries of levofloxacin and ciprofloxacin were 55.2% and 77.3%, respectively. This method was successfully applied to the pharmacokinetic study of levofloxacin in humans.     

Liquid chromatography-tandem mass spectrometry method for the determination of trimetazidine in human plasma

A sensitive liquid chromatography-tandem mass spectrometry (LC/MS/MS) method was developed and validated for the determination of trimetazidine (CAS 13171-25-0) in human plasma, using pseudoephedrine as internal standard (IS). Plasma samples were simply pretreated with methanol for deproteinization. The chromatographic separation was performed on a C18 column with a mobile phase of 3 mmol/L ammonium acetate solution-methanol (15:85, v/v) at a flow rate of 0.3 mL/min. The chromatographic separation was achieved in less than 3.2 min. The linearity was established over the concentration range of 1-100 ng/mL. Both intra- and inter-batch standard deviations were less than 9.5%. The method was successfully applied to study the relative bioavailability of trimetazidine hydrochloride tablets in healthy Chinese volunteers and the pharmacokinetic parameters of the reference and test tablets were compared.     

In vitro plasma protein binding determination of flunarizine using equilibrium dialysis and liquid chromatography-tandem mass spectrometry

A highly sensitive method was developed and validated for determining the free fraction of flunarizine in human plasma. Equilibrium dialysis was used for the separation of free (unbound) drug and liquid chromatography/tandem mass spectrometry (LC-MS/MS) was used for quantitation. Post-dialysis plasma or buffer samples of 0.2 mL were extracted using a liquid-liquid extraction procedure and analyzed using a high performance liquid chromatography electrospray tandem mass spectrometer system. The compounds were eluted isocratically on a Supelco Supelcosil ABZ+Plus column, ionized using a positive ion atmospheric pressure electrospray ionization source, and analyzed using multiple reaction monitoring. The ion transitions monitored were m/z 405-->203 for flunarizine and m/z 409-->207 for flunarizine-d4 (internal standard, IS). The chromatographic run time was 3.5 min per injection, with retention times of 2.1 min for both flunarizine and IS. The calibration curve for flunarizine was linear over the concentration range of 0.25-2000 ng/mL (r(2)>0.9989) in the combined matrix of human plasma and isotonic sodium phosphate buffer (1:1, v/v) with the lower limit of quantitation of 0.25 ng/mL. The inter-assay coefficient of variability (CV) for the quality control samples was less than 13.5%, and the inter-assay percent nominal was greater than 98.2%. In vitro protein binding of flunarizine was determined at concentrations of 5, 10 and 100 microg/mL using the validated method. Flunarizine was extensively bound to plasma protein with a 0.083+/-0.005% overall percent free drug in plasma and a CV value less than 7.8%. This validated method will be used for the ex vivo assessment of flunarizine protein binding in human plasma from a drug-drug interaction clinical study.     

Selective,sensitive, and rapid liquid chromatography-tandem mass spectrometry method for determination of Guaifenesin in human plasma

A simple, sensitive, selective, and rapid liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed and validated for quantitation of guaifenesin in human plasma using guaifenesin-d3 as internal standard (IS). Following solid phase extraction, the analyte was chromatographed using an isocratic mobile phase on a reversed-phase C₁₈ column and analyzed by MS in multiple reaction-monitoring (MRM) mode (positive ion mode). The limit of quantitation for this method was 8?ng/mL and the linear dynamic range was 8–2200?ng/mL.     

Validated liquid chromatography-tandem mass spectrometric method for the quantitative determination of daidzein and its main metabolite daidzein glucuronide in rat plasma

A highly selective and sensitive liquid chromatographic-tandem mass spectrometric (LC-MS-MS) method was developed and validated to determine daidzein and its main metabolite daidzein glucuronide in rat plasma. The analytes and internal standard genistein were extracted from plasma samples by n-hexane-diethyl ether (1:4, v/v), and separated on a C18 column. The mobile phase consisted of acetonitrile-water-formic acid (80 : 20: 1, v/v/v). Detection was performed on a triple quadrupole tandem mass spectrometer by selected reaction monitoring (SRM) mode via atmospheric pressure chemical ionization (APCI) source. The method has a limit of quantification of 0.24 ng/ml. The linear calibration curves were obtained in the concentration range of 0.24-1000 ng/ml. The intra- and inter-day precisions were lower than 13.2% in terms of % RSD. The accuracy ranged from -0.5% to 2.4% in terms of % RE (relative error). This method was successfully applied to the determination of plasma concentration of daidzein and its main metabolite daidzein glucuronide in rats after an oral administration of 20 mg/kg daidzein.     

液相色谱-串联质谱法测定人血浆中氟氯西林的含量

目的为临床制定安全有效的给药方案提供参考。方法采用液相色谱-串联质谱(LC-MS-MS)法。血浆样品经液-液萃取处理后,以乙腈-水(体积比25∶75)为流动相,采用Zorbax Ec lipseXDB C8柱(150 mm×4.6 mm,5μm)分离;通过电喷雾三重四级杆串联质谱,以负离子、选择反应监测方式进行检测,用于定量的离子反应分别为m/z452→m/z311(氟氯西林)和m/z468→m/z327(内标双氯西林)。结果氟氯西林线性范围为20.0~15000μg.L-1,定量下限为20.0μg.L-1,提取回收率均大于70%,日内、日间精密度均小于7%。结论该法适用于注射用氟氯西林钠的单次肌内注射给药的药物动力学研究。     

Quantitative determination of methylnaltrexone in human serum using liquid chromatography-tandem mass spectrometry

Methylnaltrexone (MNTX) is a novel peripherally acting μ-opioid antagonist that prevents peripheral side effects of opioid drugs such as constipation without affecting the analgesia. We developed a selective and sensitive assay to measure MTNX concentrations in human serum.The drug was measured after protein precipitation with perchloric acid using naltrexone as internal standard and liquid chromatography-tandem mass spectrometry (LC-MS/MS) for detection. The chromatography was performed isocratically on a RP18 column using 25 mM ammonium acetate buffer (pH 4)/acetonitrile (90%/10%; flow rate 200 μl/min) as mobile phase. The MS/MS analysis was performed in positive ionization mode monitoring the m/z transitions 356.4/284.2 for MNTX and 342.4/324.2 for naltrexone.The method was validated according to selectivity, linearity, accuracy, precision, recovery, matrix effects and stability. The validation range for MNTX in serum was 0.5-250 ng/ml. The developed LC-MS/MS was shown to be valid and successfully applied to measure serum-concentration-time curves of MNTX in a pilot study in healthy volunteers.     

液相色谱-串联质谱法测定头发中11种阿片类生物碱

建立头发中海洛因、吗啡、单乙酰吗啡等11种阿片类生物碱的液相色谱-串联质谱测定方法,并考察海洛因滥用者头发中阿片类组分的存在情况。头发经冷冻研磨后加入硼酸缓冲液超声30 min,用氯仿-异丙醇(9∶1)提取。用Allure PFP丙基柱,以乙腈-乙酸铵(0.1%甲酸)梯度洗脱分离,采用二级质谱多反应监测模式(MRM)检测11种阿片类生物碱。头发中海洛因、吗啡、单乙酰吗啡等11种阿片类生物碱在对应质量浓度范围内线性良好(r>0.996 0);检测限(LOD)均小于0.05 ng.mg-1;回收率范围为47.2%～110%;日内精密度和日间精密度均小于14%。21例海洛因滥用者头发中均检出了海洛因、单乙酰吗啡、吗啡、可待因、乙酰可待因、氢可酮等主要组分。所建方法灵敏度高、选择性好,适用于同时分析头发中海洛因等11种阿片类生物碱组分,可有效鉴别海洛因滥用与阿片类药物或食品的摄取。     

Quantitative determination of sufentanil in human plasma by liquid chromatography-tandem mass spectrometry

A sensitive and specific method for the quantification of sufentanil in human plasma by liquid chromatography coupled with tandem mass spectrometry has been developed. Fentanyl was used as the internal standard. Rapid sample preparation involved purification on a C(18) solid-phase extraction column. Chromatographic separation of the analytes was obtained using an RP-C(18) mu-HPLC column. LC-MS-MS detection was performed by atmospheric pressure ionisation (API) source equipped with an ionspray (IS) interface operating in the positive ion mode. For unambiguous substance confirmation, three analyte precursor-product ion combinations were monitored during multiple reaction monitoring (MRM) LC-MS-MS analysis. The method's performance characteristics were evaluated in blank and spiked control plasma samples. Overall accuracy (relative error, R.E., %), repeatability (relative standard deviations, R.S.D., %) and within-laboratory reproducibility (R.S.D., %) ranged from -9.28 to -2.71%, from 6.42 to 2.82% and from 13.52 to 6.06%, respectively, for sufentanil. The limit of quantification for sufentanil in human plasma samples was 0.3 ng ml(-1). Due to its high sensitivity and specificity, the method was successfully employed for sufentanil determination in maternal plasma samples collected immediately before epidural administration of a single sufentanil dose to women in labour, 20 min after drug administration, and at birth in arterial and venous umbilical cord plasma samples from the newborn babies. Research is in progress to adopt the method for performance of complete pharmacokinetic studies of sufentanil in human plasma.     

Development and validation of a method for quantitative determination of valsartan in human plasma by liquid chromatography-tandem mass spectrometry

A sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of valsartan in human plasma was developed and validated. A 0.5 ml aliquot was extracted using solid-phase extraction in an Empore^® high performance extraction disk plate, universal resin 96-well format. The estimated calibration range of the method was 2–2000 ng/ml.

The method was fully validated with intra-day mean accuracy and precision of 94.8–107% and 2.19–5.40% and inter-day mean accuracy and precision of 93.5–105% and 1.87–5.67%, respectively. No significant loss of valsartan in processed samples was confirmed in processed samples for up to 24 h at 10 °C. Sample dilution up to 50-fold with blank human plasma provided acceptable analyses. No interference peaks or matrix effects were observed. No effect of QC sample location results was observed in a 96-well plate. This LC-MS/MS technique was found to improve quantitative determination of valsartan allowing its pharmacokinetic evaluation with clinically relevant doses.     

Liquid chromatography-tandem mass spectrometry detection of the quaternary ammonium compound mebezonium as an active ingredient in t61

Quaternary ammonium compounds pose an analytical challenge. Mebezonium, a muscle-relaxing agent contained in veterinary euthanasia solution T61, was analyzed in body fluids, organs, and injection sites of a veterinarian by liquid chromatography-tandem mass spectrometry (LC-MS-MS) method. Additionally, embutramide and tetracaine, which are two other active ingredients contained in T61, methadone, xylazine, and analgesics were detected by LC-MS-MS and high-performance liquid chromatography-ultraviolet detection methods. For detection of mebezonium a solid-phase extraction (SPE) combined with ionpairing reagent heptafluorobutyric acid was developed. Separation was achieved on Phenomenex Synergi Hydro RP C(18) column combined with ammonium formate buffer and acetonitrile (pH 3.5). To enrich other drugs, liquid-liquid extraction procedures were used. Most of these drugs were separated on a Restek Allure PFP Propyl column using the mentioned mobile phase. Mebezonium and embutramide were detected in femoral vein serum in concentrations of 10.9 and 2.0 mg/L, respectively. The concentration of xylazine and methadone in serum was 2.0 and 0.4 mg/L, respectively. The LC-MS-MS method with SPE combined with an ion-pairing reagent allowed the quantitation of mebezonium. Methadone was detected in toxic concentrations and was, in combination with xylazine and T61, considered to be the cause of death.