液相色谱-质谱法测定人体血浆中非那雄胺含量

建立了测定人体血浆中非那雄胺药物浓度的液相色谱-质谱分析方法。色谱柱为Hypersil-Keystone C_(18)柱,流动相为乙腈∶水(含0.2%三氯乙酸)=55∶45(V/V),质谱选择电喷雾正离子源(ESI+),选择离子模式(SIM)监测,非那雄胺和内标卡马西平质荷比分别为m/z373和m/z237。血浆样品经乙腈去蛋白、离心和吹干后,残渣用流动相溶解。当非那雄胺的浓度在0.5~120ng·mL~(-1)范围时线性关系良好(r=0.999),检测限(S/N=3)为0.02ng·mL~(-1);日间和日内测定的精密度(RSD)分别在6.4%~2.2%和5.5%~1.7%之间;加标0.5ng·mL~(-1)、40ng·mL~(-1)和80ng·mL~(-1)非那雄胺的平均回收率(n=6)分别为108.5%、101.2%和98.7%。方法快速、准确和灵敏,已成功用于人血浆中非那雄胺浓度的测定和人体药代动力学研究。     

Determination and pharmacokinetics of manidipine in human plasma by HPLC/ESIMS

A sensitive HPLC/ESIMS method was established for the determination of manidipine in human plasma and pharmacokinetics study. After basified plasma with ammonia, manidipine and the internal standard (IS) (felodipine) were extracted with n-hexane and separated on a Hypersil ODS2 column with a mobile phase of methanol-5 mm ammonium acetate solution containing 0.1% acetic acid (85:15, v/v). MS determination was performed by electrospray ionization in the selected ion monitoring mode. Manidipine was monitored at m/z 611.4 and IS at m/z 384. The assay had a calibration range from 0.2 to 20 ng/mL and a lower limit of quantification of 0.1 ng/mL. The method has been successfully applied to the pharmacokinetic study in healthy volunteers.     

Development and validation of a hydrophilic interaction liquid chromatography-tandem mass spectrometric method for the analysis of paroxetine in human plasma

A sensitive, simple, fast and rugged hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method for the determination of paroxetine was developed and validated over curve range 0.050-50 ng/mL using only 0.4 mL plasma. This is the first published LC-MS/MS method and the low limit of quantitation of this method is 10-fold lower than previously published methods. A simple liquid-liquid extraction method using methyl-tert butyl ether (MTBE) as the extraction solvent was used to extract paroxetine and the internal standard (IS) fentanyl-d(5) from plasma. The extract was evaporated to dryness, reconstituted and injected onto a silica column using a low aqueous-high organic mobile phase. The chromatographic run time was 2.0 min per injection, with retention times of 1.1 and 1.2 min for paroxetine and IS, respectively. The detection was by monitoring paroxetine at m/z 330 --> 192 and IS at m/z 342 --> 188, respectively. The inter-day precision and accuracy of the quality control (QC) samples were <5.0% relative standard deviation (RSD) and <2.9% relative error (RE). This method can be used for supporting therapeutical drug monitoring and pharmacokinetic or drug-drug interaction studies.     

Determination of testosterone propionate in human plasma by gas chromatography-mass spectrometry

A specific, sensitive and accurate quantitative analysis of testosterone propionate in human plasma was developed using gas chromatography-mass spectrometry-selected-ion monitoring. For the calculation of testosterone propionate in plasma, peak height ratios were measured by selected-ion monitoring performed on the molecular ions of the trifluoroacetyl derivative of testosterone propionate (m/z 440) and testosterone propionate-19,19,19-d3 (m/z 443). The sensitivity of the method was judged from the lower limit of the detection of the mass spectrometer which was at 20 pg. The inter-assay coefficients of variation and relative error at a concentration of 1.31 ng/ml of plasma were 5.47% and -2.3%, respectively. The method described was applied to the determination of plasma concentrations of testosterone propionate-19,19,19-d3 following an intramuscular dose of testosterone propionate-19,19,19-d3 in a healthy male volunteer.     

Simultaneous determination of spironolactone and its active metabolite canrenone in human plasma by HPLC-APCI-MS

A sensitive and specific liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) method for the simultaneous determination of spironolactone and its active metabolite canrenone in human plasma has been developed and validated. After the addition of estazolam as the internal standard (IS), plasma samples were extracted with methylene chloride : ethyl acetate mixture (20 : 80, v/v) and separated by high-performance liquid chromatography (HPLC) on a reversed-phase C18 column with a mobile phase of methanol-water (57 : 43, v/v). Analytes were determined in a single quadrupole mass spectrometer using an atmospheric pressure chemical ionization (APCI) source. LC-APCI-MS was performed in the selected-ion monitoring (SIM) mode using target ions at m/z 341.25 for spironolactone and canrenone, m/z 295.05 for estazolam. The method was proved to be sensitive and specific by testing six different plasma batches. Calibration curves of spironolactone and canrenone were linear over the range 2-300 ng/ml. The within- and between-batch precisions (relative standard deviation (RSD)%) were lower than 10% and the accuracy ranged from 85 to 115%. The lower limit of quantification (LLOQ) was identifiable and reproducible at 2 ng/ml. The proposed method was successfully applied to study the pharmacokinetics of spironolactone and its major metabolite in healthy male Chinese volunteers.     

Determination of azasetron hydrochloride in rabbit plasma by liquid chromatography tandem mass spectrometry and its application

A sensitive and selective liquid chromatography tandem mass spectrometry method for determination of azasetron hydrochloride in rabbit plasma was developed. After addition of doxapram hydrochloride as internal standard (IS), protein precipitation by 10% trichloroacetic acid was used as sample preparation. Chromatographic separation was achieved on a Zorbax SB-C(18) (2.1 × 50 mm, 3.5 μm) column with acetonitrile-water as mobile phase with gradient elution. An electrospray ionization source was applied and operated in positive ion mode; multiple reaction monitoring mode was used to quantification using target fragment ions m/z 349.9 → 223.5 for azasetron hydrochloride and m/z 378.9 → 291.8 for the IS. Calibration plots were linear over the range of 6-1000 ng/mL for azasetron hydrochloride in plasma. The lower limit of quantitation for azasetron hydrochloride was 6 ng/mL. The mean recovery of azasetron hydrochloride from plasma was in the range 85.6-92.7%. The RSDs of intra-day and inter-day precision were both less than 12%. This method is simple and sensitive enough to be used in pharmacokinetic research for determination of azasetron hydrochloride in rabbit plasma.     

Comparison of pyrolysis-mass spectrometry with high performance liquid chromatography for the analysis of vancomycin in serum

Mass spectrometry coupled with a pyrolysis inlet system (Pyr-ms) is compared with high performance liquid chromatography (HPLC) for the determination of vancomycin and its crystal degradation products (CDP-Is) in human serum. Quantitative analysis of Pyr-ms was performed by selected ion monitoring (SIM) method at 108 mass:charge (m/z) of pyrolysis product of vancomycin. 3-Nitroaniline (138 m/z) was used as an internal standard. A mu-Bondapak C(18) column and the gradient mobile phase of triethylamine buffer (pH 6.2), acetonitrile and tetrahydrofuran and a photometric detection at 205 nm are found to be the optimum conditions for the HPLC determination of vancomycin and CDP-Is. The limit of detection (LOD=1 ng ml(-1)), linearity (1 ng ml(-1)-10 mg ml(-1)), relative standard deviation (R.S.D.=1%), time analysis (1/2 h) and sample volume (50 mul) of Pyr-ms are far better than of the HPLC method. However, the HPLC method can individually determine the concentration of vancomycin and its degradation products.     

Selective and sensitive determination of bis(7)-tacrine, a high erythrocyte binding acetylcholinesterase inhibitor, in rat plasma by high-performance liquid chromatography-tandem mass spectrometry

The current study aims to develop a specific and sensitive LC-MS/MS method for determination of bis(7)-tacrine (B7T) in rat plasma. A 100 microL plasma sample was extracted with ethyl acetate. B7T and the internal standard (IS), pimozide, in the samples were then analyzed with LC-MS/MS in positive electrospray ionization condition. Chromatographic separation of B7T and IS was achieved in a C(18) reversed-phase HPLC column (150 x 2.1 mm i.d.) by isocratic elution with a mobile phase consisting of 0.05% formic acid in water and acetonitrile (1:1, v/v) at a flow rate of 0.35 mL/min. Multiple-reaction monitoring (MRM) mode was employed to measure the ion transitions: m/z 247 to 197 for B7T and m/z 462 to m/z 328 for IS, respectively. The method was linear over the studied ranges of 100-5000 and 10-100 ng/mL. The intra-day and inter-day variations of the analysis were less than 6.8% with standard errors less than 9.0%. The detection limit of B7T in rat plasma was 1 ng/mL. The developed method was successfully applied to the pharmacokinetic study of B7T after intravenous administration of 1 mg/kg B7T and further proved to be readily utilized for determination of B7T in rat plasma samples.     

Simultaneous quantitation of nicorandil and its denitrated metabolite in plasma by LC-MS/MS: application for a pharmacokinetic study

A liquid chromatography-electrospray ionization tandem mass spectrometry method was developed and validated for the simultaneous quantitation of nicorandil and its denitrated metabolite, N-(2-hydroxyethyl)-nicotinamide, in rat plasma. After a liquid-liquid extraction step, chromatographic separation was performed on a ShinPack C(18) column with an isocratic mobile phase composed of methanol and 2 mM aqueous ammonium acetate containing 0.03% (v/v) formic acid (33:67 v/v). Procainamide was used as an internal standard (IS). Selected reaction monitoring was performed using the transitions m/z 212 → m/z 135, m/z 166 → m/z 106 and m/z 236 → m/z 163 to quantify nicorandil, its denitrated metabolite and IS, respectively. Calibration curves were constructed over the range of 5-15,000 ng.ml(-1) for both nicorandil and its metabolite. The mean relative standard deviation (RSD%) values for the intra-run precision were 5.4% and 7.3% and for the inter-run precision were 8.5% and 7.3% for nicorandil and its metabolite, respectively. The mean accuracy values were 100% and 95% for nicorandil and its metabolite, respectively. No matrix effect was detected in the samples. The validated method was successfully applied to a pharmacokinetic study after per os administration of nicorandil in rats.     

Detection of Anabolic Steroids By Gc/Sim/Ms With Trifluoroacetylation in Equine Plasma and Urine

ABSTRACT

A method to detect three anabolic steroids (boldenone, nandrolone and mesterolone) is presented. The anabolic steroids are isolated from equine plasma and urine by extraction with diethyl ether and C₁₈ Sep-Pak cartridge adsorption, respectively. The extracts obtained were derivatized with trifluoroacetyl anhydride and analyzed by GC/SIM/MS. The selected ion monitoring (SIM) mode was applied to increase the sensitivity and, when possible, the higher m/z ions were selected to improve identification. Stability of derivatives was good and compounds having hydroxy and conjugated ketone groups produced trifluoroacetyl ester derivatives that were also stable. Repeatability of the chromatographic analysis was evaluated on the basis of area repeatability, and the coefficient of variation obtained was lower than 4.4. The detection limit was 1 and 5 ng/ml for all the anabolic steroids studied in equine plasma and urine, respectively.     

Liquid chromatography-tandem mass spectroscopic method for the determination of zerumbone in human plasma and its application to pharmacokinetics

A rapid, sensitive, specific and selective LC-MS/MS method for the determination of zerumbone (ZER) in human plasma using 2,4-diamino-6-(4-methoxyphenyl)-1,3,5-triazine (DMTZ) as an internal standard (IS) has been developed and validated. ZER was chromatographed on C8 column using a mobile phase of acetonitrile/water (80:20, v/v) at a flow rate of 0.25 ml min(-1) . Quantitation was achieved using ESI+ interface, employing multiple reaction monitoring (MRM) mode at m/z 219 > 81 and 218 > 134 for ZER and IS, respectively. The calibration standards were linear over a range of 5-3000 ng ml(-1) (r(2)=0.9994) with an LLOQ of 5 ng ml(-1) (RSD %; 11.4% and bias%; 9.5%). Intra- and inter-day precision of ZER assay ranged from 0.18 to 3.56% with accuracy (bias) that varied between -5.09 and 4.3%, demonstrating good precision and accuracy. Recoveries of ZER and the IS from human plasma were above 85%. The developed method was validated for the determination of ZER in rat plasma. Linearity, stability of ZER and the ME on rat plasma were discussed. The applicability of the developed method was demonstrated by measuring ZER in rat plasma samples following intravenous and intraperitoneal administration of ZER prepared in hydroxypropyl-β-cyclodextrin (HPβCD) and sodium carboxymethyl cellulose (CMC), respectively, in 20 mg kg(-1) and this study indicated a clear significant difference (p<0.05) in pharmacokinetic parameters of ZER in ZER/HPβCD complex compared with ZER in CMC preparation.     

Simultaneous quantification of a non-nucleoside reverse transcriptase inhibitor efavirenz, a nucleoside reverse transcriptase inhibitor emtricitabine and a nucleotide reverse transcriptase inhibitor tenofovir in plasma by liquid chromatography positive ion electrospray tandem mass spectrometry

A high-performance liquid chromatography/positive ion electrospray tandem mass spectrometry method for the simultaneous quantification of efavirenz, emtricitabine and tenofovir was developed and validated with 100 microL human plasma. Following solid-phase extraction, the analytes were separated using a gradient mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H]+ ions, m/z 316 to 168 for efavirenz, m/z 248-130 for emtricitabine and m/z 288-176 for tenofovir, m/z 482-258 for rosuvastatin (IS), m/z 260-116 for propranolol (IS). The method exhibited a 100-fold linear dynamic range for all the three analytes in human plasma (20-2000, 2-200 and 20-2000 ng/mL for efavirenz, emtricitabine and tenofovir respectively). The lower limit of quantification was 2 ng/mL for emtricitabine and 20 ng/mL for both efavirenz and tenofovir with a relative standard deviation of less than 11%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The total chromatographic run time of 4 min for each sample made it possible to analyze more than 250 human plasma samples per day. The method is precise and sensitive enough for its intended purpose. The method is also successfully applied to quantify efavirenz, emtricitabine and tenofovir concentrations in a rodent pharmacokinetic study.     

Determination of bergenin in human plasma after oral administration by HPLC-MS/MS method and its pharmacokinetic study

A highly sensitive, simple and selective high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and applied to the determination of bergenin concentration in human plasma. Bergenin and the internal standard (IS) thiamphenicol in plasma were extracted with ethyl acetate, separated on a C(18 )reversed-phase column, eluted with mobile phase of acetonitrile-water, ionized by negative ion pneumatically assisted electrospray and detected in the multi-reaction monitoring mode using precursor --> product ions of m/z 327.1 --> 192 for bergenin and 354 --> 185.1 for the IS, respectively. The linear range of the calibration curve for bergenin was 0.25-60 ng mL(-1), with the lowest limit of quantification of 0.25 ng mL(-1), and the intra/inter-day relative standard deviation (RSD) was less than 10%. The method is suitable for the determination of low bergenin concentration in human plasma after therapeutic oral doses, and has been first and successfully used for its pharmacokinetic studies in healthy Chinese volunteers.     

同位素稀释-气相色谱-质谱法检测大鼠血浆中的内源性胍丁胺

建立了大鼠血浆中内源性胍丁胺的同位素稀释-气相色谱-负化学电离质谱定量分析方法。大鼠血浆样品经蛋白沉淀并蒸干后,用六氟乙酰丙酮衍生化,采用Florisil固相萃取柱净化,以稳定同位素标记的d₈-胍丁胺为内标,在气相色谱-质谱仪上采用负化学电离方式电离,选择离子模式检测。胍丁胺标准溶液的检出限为0.0057 ng/mL。血浆中添加的胍丁胺在1.14~57.0 ng/mL范围内呈良好的线性关系（r=0.997）,方法回收率介于92.3%~109.8%之间,日内和日间精密度均小于15%。大鼠血浆中胍丁胺平均含量水平为（22±9）ng/mL,雌、雄大鼠血浆中的胍丁胺水平未见显著性差异（p>0.05）。该方法特异性好、灵敏度高,为生物体内胍丁胺的生理功能研究提供了高灵敏的分析方法。     

Simultaneous measurements of endogenous and deuterium-labelled tracer variants of androstenedione and testosterone by capillary gas chromatography-mass spectrometry

A capillary gas chromatographic-mass spectrometric method for the simultaneous determination of androstenedione and testosterone in human plasma using [19,19,19-2H3]androstenedione and [19,19,19-2H3]testosterone as internal standards is described. For calculation of plasma androstenedione and testosterone, peak heights were measured by selected-ion monitoring of the molecular ions of the heptafluorobutyryl derivatives of androstenedione and [2H3]androstenedione (m/z 482 and 485) and of testosterone and [2H3]testosterone (m/z 680 and 683). The isotope dilution method needed no complex corrections for contributions and provides a sensitive and reliable technique with good accuracy, precision and reproducibility.     

Gas chromatographic-mass spectrometric determination of plasma selegiline using a deuterated internal standard.

A gas chromatographic-mass spectrometric method is described for the determination of plasma selegiline. Tetradeuteroselegiline was synthesized and served as the internal standard. Human plasma samples (1 ml) containing 1-6 ng of selegiline were acidified, washed with diethyl ether-hexane, then alkalinized and extracted with heptane-isoamyl alcohol. Analytical separations were performed on a dimethylsilicone capillary column. Detection was by selected ion monitoring of the electron impact generated m/z 96 and 100 alpha-cleavage fragments of drug and internal standard, respectively.     

Simultaneous quantification of atorvastatin and active metabolites in human plasma by liquid chromatography-tandem mass spectrometry using rosuvastatin as internal standard

A simple, sensitive, selective and rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantification of atorvastatin and its active metabolites ortho-hydroxyatorvastatin and para-hydroxyatorvastatin in human plasma using rosuvastatin as internal standard (IS). Following simple liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reversed-phase C18 column and analyzed by MS in the multiple reaction monitoring mode using the respective [M+H]+ ions, m/z 559/440 for atorvastatin, m/z 575/466 for ortho-hydroxyatorvastatin, m/z 575/440 for para-hydroxyatorvastatin and m/z 482/258 for the IS. The assay exhibited a linear dynamic range of 0.1-20 ng/mL for atorvastatin and its two metabolites in human plasma. The lower limit of quantification was 100 pg/mL with a relative standard deviation of less than 8%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The average absolute recoveries of atorvastatin, ortho-hydroxyatorvastatin, para-hydroxyatorvastatin and the IS from spiked plasma samples were 54.2 +/- 3.2, 50.1 +/- 3.8, 65.2 +/- 3.6 and 71.7 +/- 2.7%, respectively. A run time of 2.5 min for each sample made it possible to analyze more than 300 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies.     

Liquid chromatography/electrospray ionization mass spectrometry method for the quantification of valproic acid in human plasma

A simple, sensitive and rapid liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) method was developed and validated for the quantification of valproic acid, an antiepileptic drug, in human plasma using benzoic acid as internal standard (IS). Following solid-phase extraction, the analytes were separated using an isocratic mobile phase on a reversed-phase C18 column and analyzed by MS in the single ion monitoring mode using the respective [M-H]- ions, m/z 143 for valproic acid and m/z 121 for the IS. The assay exhibited a linear dynamic range of 0.5-60 microg/mL for valproic acid in human plasma. The lower limit of quantification was 500 ng/mL with a relative standard deviation of less than 10%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The average absolute recoveries of valproic acid and the IS from spiked plasma samples were 96.1+/-4.2 and 95.6+/-2.7%, respectively. A run time of 4.5 min for each sample made it possible to analyze more than 250 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability and bioequivalence studies.     

Rapid liquid chromatography-tandem mass spectrometry method for quantification of ziprasidone in human plasma

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of ziprasidone (ZIP) in human plasma was developed. ZIP and N-methyl ziprasidone as internal standard (IS) were extracted from alkalinized plasma using tert- butyl methyl ether. Separation was performed isocratically on a C8 column with 90% acetonitrile containing 2 mmol/L ammonium acetate as a mobile phase with a total run time of 2.5 min. MS/MS transitions of m/z 413 --> 194 and m/z 427 --> 177 of the analyte and internal standard were used for quantification. Confirmatory ions of m/z 413 --> 177 and m/z 427 --> 180 were collected as well. The calibration curve based on peak-area ratio was linear up to at least 200 ng/mL with a detection limit of 0.1 ng/mL. The method showed satisfactory reproducibility with a coefficient of variation of less than 5%. The method was successfully applied to the analysis of ZIP in spiked human plasma.     

Simultaneous determination of docetaxel and ketoconazole in rat plasma by liquid chromatography/electrospray ionization tandem mass spectrometry

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method has been developed and validated for simultaneous quantification of docetaxel and ketoconazole in rat plasma with paclitaxel as internal standard (IS). The analytes were extracted from rat plasma by using a liquid-liquid extraction technique with ethyl acetate and the LC separation was performed on a Cosmosil-C(18) analytical column (150 mm x 2.0 mm i.d., Nacalai Tesque Inc., Japan). The extracted samples were analyzed with LC/MS/MS, operating in selected reaction monitoring (SRM) mode. The SRM transitions of precursor ions to product ions were 830.3-->549.1 (m/z) for docetaxel, 531.2-->489.3 (m/z) for ketoconazole, and 876.7-->307.9 (m/z) for the IS. The calibration curves were linear over the range of 2-500 ng/mL for docetaxel and 50-20 000 ng/mL for ketoconazole, with coefficients of correlation above 0.999. The limits of quantification for docetaxel and ketoconazole were both 2 ng/mL. The limit of detection for each analyte was 1 ng/mL. The intra- and inter-day precision (CV) of analysis were within 7%, and the accuracy ranged from 95 to 110%. The overall recoveries for docetaxel and ketoconazole were about 89.0% and 91.1%, respectively. The total analysis time was only 9.0 min. This quantitation method was successfully applied to the simultaneous determination of docetaxel and ketoconazole in rat plasma and some potential interaction was found in the current coadministration pharmacokinetic study. This established method was also utilized in the in vitro and in vivo drug-drug interaction study of docetaxel and ketoconazole (to be published).