Rapid High-performance Liquid Chromatography-tandem Mass Research Articles

Comparative Study on the Pharmacokinetics of Paeoniflorin, White Peony Root Water Extract, and Taohong Siwu Decoction After Oral Administration in Rats

Taohong Siwu Decoction (TSD) is a classic traditional Chinese medicine (TCM) compound with pharmacological effects such as vasodilation and hypolipidemia. Paeoniflorin (PF) is one of the active ingredients of TSD. The aim of this study was to evaluate the pharmacokinetics of PF in herbal extracts and their purified formsin rats. A sensitive and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS-MS) method for the determination of PF in rat plasma was developed. Rats were divided into three groups, and given PF solution, water extract of white peony root (WPR), or TSD by gavage. At different predetermined timepoints after gavage, blood was collected from the orbital vein. The pharmacokinetic parameters of PF in the plasma of rats in the three groups was determined. The pharmacokinetic studies showed that the time to reach maximum concentration (Tmax) of PF in the purified forms group was relatively high, while the half-lives (T½) of PF in the TSD and WPR groups were longer. Among the three groups, PF in the purified forms group had the maximum area under the concentration-time curve (AUC0-t=732.997µg/L·h) and the largest maximum concentration (Cmax=313.460µg/L), which showed a significant difference compared with the TSD group (P<0.05). Compared with the purified group, the clearance (CLz/F=86.004L/h/kg) and the apparent volume of distribution (Vz/F=254.787L/kg) of PF in the TSD group increased significantly (P<0.05). A highly specific, sensitive, and rapid HPLC-MS-MS method was developed and applied for the determination of PF in rat plasma. It was found that TSD and WPR can prolong the action time of paeoniflorin in the body.

Pharmacokinetic and brain distribution study of an anti-glioblastoma agent in mice by HPLC-MS/MS.

Previously compound I showed great anti-glioblastoma activity without toxicity in a mouse xenograft study. In this study, a sensitive and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and validated to investigate the pharmacokinetics and brain distribution of compound I in mice. The protein precipitation method was applied to extract the compound from mouse plasma and brain homogenates, and it was then separated using a Kinetex C18 column with a mobile phase consisting of acetonitrile-0.1% formic acid water (50:50, v/v). The analytes were detected with multiple reaction monitoring for the quantitative response of the compounds. The inter- and intra-day precisions were <8.29 and 3.85%, respectively, and the accuracy range was within ±7.33%. The method was successfully applied to evaluate the pharmacokinetics of compound I in mouse plasma and brain tissue. The peak concentration in plasma was achieved within 1h. The apparent elimination half-life was 4.06 h. The peak concentration of compound I in brain tissue was 0.88 μg/g. The results indicated that compound I was rapidly distributed and could cross the blood-brain barrier. The pharmacokinetic profile summarized provides valuable information for the further investigation of compound I as a potential anti-glioblastoma agent.

Pharmacokinetic study of an anti-trypanosome agent with different formulations and administration routes in mice by HPLC-MS/MS.

Previously compound 12 showed great anti-trypanosome activity without toxicity in an in vivo study. In the current study, a sensitive and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and validated to investigate its pharmacokinetics in mouse plasma. A protein precipitation method was applied to extract the compound, and it was then separated using a Kinetex C18 column with mobile phase consisting of acetonitrile-0.1% formic acid water (50:50, v/v) at a flow rate of 300 μl/min. The analytes were detected with the multiple reaction monitoring in negative electrospray ionization source for quantitative response of the compounds. Compound 12 was detected at m/z 477.0 → 367.2, while the internal standard compound 14 was detected at m/z 499.2 → 268.2. Inter- and intra-day precision was <5.22 and 2.79% respectively, while the accuracy range was within ±9.65%. The method was successfully applied to evaluate the pharmacokinetics of compound 12 in mouse plasma with two formulations (20% Cremophor EL or sesame oil) and drug administration routes (oral and intraperitoneal injection). We observed a better drug serum concentration with the Cremophor formulation, and the two different drug administration routes did not show significant differences from the drug distribution.

Determination of finerenone – a novel, selective, nonsteroidal mineralocorticoid receptor antagonist – in human plasma by high-performance liquid chromatography-tandem mass spectrometry and its application to a pharmacokinetic study in venous and capillary human plasma

A straightforward and rapid high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) assay allowing the sensitive and selective quantitation of finerenone (BAY 94–8862) in lithium heparin human plasma is described. Finerenone is a novel, selective, nonsteroidal mineralocorticoid receptor antagonist that is in phase III clinical trials for the treatment of chronic kidney disease. Finerenone quantitation is performed after addition of its stable isotope-labelled internal standard (ISTD) by protein precipitation with acidified acetonitrile followed by HPLC–MS/MS separation and detection. The determination of finerenone concentrations was validated for a plasma volume of 0.100 mL and subsequently also for a lower plasma volume of 0.010 mL, collected e.g. in paediatric studies. The analytical range was from 0.100 µg/L (lower limit of quantification) to 200 µg/L (upper limit of quantification). Inter-day accuracy was 99.7–105.0% for the plasma volume of 0.100 mL and 101.1–104.5% for the plasma volume of 0.010 mL. Inter-day precision was ≤ 7.0%, independent of the extracted plasma volume. A moderate, concentration-independent matrix effect on ionisation was observed for both finerenone and its ISTD of 0.535–0.617, which is fully compensated by the ISTD (ISTD-normalised matrix factors were 0.98–1.03). The assay was successfully applied with both validated plasma volumes to a clinical phase I study in which the pharmacokinetics of 20 mg finerenone were compared in capillary plasma (0.010 mL) and venous plasma (0.100 mL) in a concentration range from the lower limit of quantification to 310 µg/L (capillary plasma) and 252 µg/L (venous plasma). The area under the plasma concentration versus time curve was similar in both matrices, while maximum concentrations were 37% higher in capillary plasma. In conclusion, capillary sampling should not bias pharmacokinetic exposure estimates compared with venous plasma values, if limited to sampling times in the distribution and elimination phases of finerenone.

Rapid and sensitive liquid chromatography–tandem mass spectrometric method for the quantitative determination of potentially harmful substance 5,5′-oxydimethylenebis (2-furfural) in traditional Chinese medicine injections

With the rapid development and wide application of traditional Chinese medicine injection (TCMI), a number of adverse events of some TCMIs have incessantly been reported and have drawn broad attention in recent years. Establishing effective and practical analytical methods for safety evaluation and quality control of TCMI can help to improve the safety of TCMIs in clinical applications. In this study, a sensitive and rapid high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method has been developed and validated for the quantitative determination of potentially harmful substance 5,5′-oxydimethylenebis (2-furfural, OMBF) in TCMI samples. Chromatographic separation was performed on a C18 reversed-phase column (150mm×2.1mm, 5µm) by gradient elution, using methanol–water containing 0.1% formic acid as mobile phase at the flow rate of 0.3mL/min. MS/MS detection was performed on a triple quadrupole mass spectrometer with positive electrospray ionization in the multiple reaction-monitoring mode. The method was sensitive with a limit of quantification of 0.3ng/mL and linear over the range of 0.3–30ng/mL (r=0.9998). Intra- and inter-day precision for analyte was <9.52% RSD with recoveries in the range 88.0–109.67% at three concentration levels. The validated method was successfully applied to quantitatively determine the compound OMBF in TCMIs and glucose injections. Our study indicates that this method is simple, sensitive, practicable and reliable, and could be applied for safety evaluation and quality control of TCMIs and glucose injections.

Analysis of prasugrel active metabolite R-138727 in human plasma: a sensitive, highly selective and fast LC-MS/MS method.

A cost effective, sensitive, simple, and rapid high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the quantification of the prasugrel metabolite in human plasma. Following solid phase extraction, the analyte (prasugrel active metabolite; R-138727) and internal standard (emtricitabine) were separated using a mobile phase in an isocratic elution mode on a reverse phase C18 column and were analyzed by an LC-MS/MS in the multiple reaction monitoring mode using the respective [M+H](+) ions, m/z 498.3-206.0 for R-138727 and m/z 248.2-130.1 for the internal standard. The assay exhibited a linear dynamic range of 0.2-120 ng/mL. The lower limit of quantification was 0.2 ng/mL with a relative standard deviation of 5.0%. This LC-MS/MS method was validated with intra-batch and inter-batch precision and accuracy. Results for precision and accuracy are in range of 3.9-9.6% and 95.2-102.2% respectively. This validated method is simple and repeatable to use in bioequivalence/pharmacokinetic studies.

A rapid high-performance liquid chromatography-tandem mass spectrometry assay for unambiguous detection of different milk species employed in cheese manufacturing

The aim of the study was to investigate the possibility to differentiate the 4 most important species in Italian dairy industry (cow, buffalo, sheep, and goat), applying a bottom-up proteomic approach to assess the milk species involved in cheese production. Selective peptides were detected in milk to use as markers in cheese products. Trypsin-digested milk samples of cow, sheep, goat, and buffalo, analyzed by HPLC-tandem mass spectrometry provided species-specific peptides, some of them recognized by Mascot software (Matrix Science Ltd., Boston, MA) as derived from well-known species specific proteins. A multianalyte multiple reaction monitoring method, built with these specific peptides, was successfully applied to cheeses with different composition, showing high specificity in detection of species involved. Neither aging nor production method seemed to affect the response, demonstrating that chosen peptides well act as species markers for dairy products.

Validation of an HPLC-MS-MS assay for determination of morellic acid in rat plasma: application to pharmacokinetic studies.

A selective and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS-MS) was developed for the quantification of morellic acid in rat plasma. HPLC was performed using a Capcell MG C18 (50 × 4.6 mm, i.d., 5 µm) column, and isocratic elution with water-acetonitrile (20:80, v/v) at a flow rate of 0.5 mL/min. Sample preparation of analyte and internal standard (gambogic acid) involved liquid-liquid extraction using ethyl acetate-isopropanol (1:1, v/v) from 50 µL plasma. The precursor → production transitions for analyte and IS were m/z 559.4 → 471.3, and m/z 627.3 → 583.3, respectively, and were monitored on a triple-quadrupole mass spectrometer, operating in negative ion scan mode. The method was validated across the dynamic concentration range of 20-7,500 ng/mL for morellic acid, with a fast run time of 6.0 min. The analytical method measured concentrations of morellic acid with accuracy (% bias) of ≤6.4% and precision (% RSD) of ≤14.0%. Morellic acid was stable during the battery of stability studies. Finally, the applicability of this assay has been successfully demonstrated in vivo pharmacokinetic studies in Sprague-Dawley rats. This method will therefore be useful for further preclinical and clinical pharmacokinetic studies of morellic acid.

Selective and rapid determination of raltegravir in human plasma by liquid chromatography–tandem mass spectrometry in the negative ionization mode

A selective and rapid high-performance liquid chromatography–tandem mass spectrometry method was developed and validated for the quantification of raltegravir using raltegravir-d3 as an internal standard (IS). The analyte and IS were extracted with methylene chloride and n-hexane solvent mixture from 100µL human plasma. The chromatographic separation was achieved on a Chromolith RP-18e endcapped C18 (100mm×4.6mm) column in a run time of 2.0min. Quantitation was performed in the negative ionization mode using the transitions of m/z 443.1→316.1 for raltegravir and m/z 446.1→319.0 for IS. The linearity of the method was established in the concentration range of 2.0–6000ng/mL. The mean extraction recovery for raltegravir and IS was 92.6% and 91.8%, respectively, and the IS-normalized matrix factors for raltegravir ranged from 0.992 to 0.999. The application of this method was demonstrated by a bioequivalence study on 18 healthy subjects.

Determination of gymnemagenin in rat plasma using high‐performance liquid chromatography–tandem mass spectrometry: application to pharmacokinetics after oral administration of Gymnema sylvestre extract

A sensitive and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method has been developed and validated for the determination of gymnemagenin (GMG), a triterpene sapogenin from Gymnema sylvestre, in rat plasma using withaferin A as the internal standard (IS). Plasma samples were simply extracted using liquid-liquid extraction with tetra-butyl methyl ether. Chromatographic separation was performed on Luna C(18) column using gradient elution of water and methanol (with 0.1% formic acid and 0.3% ammonia) at a flow rate of 0.8 mL/min. GMG and IS were eluted at 4.64 and 4.36 min, ionized in negative and positive mode, respectively, and quantitatively estimated using multiple reaction monitoring (MRM) mode. Two MRM transitions were selected at m/z 505.70 → 455.5 and m/z 471.50 → 281.3 for GMG and IS, respectively. The assay was linear over the concentration range of 5.280-300.920 ng/mL. The mean plasma extraction recoveries for GMG and IS were found to be 80.92 ± 8.70 and 55.63 ± 0.76%, respectively. The method was successfully applied for the determination of pharmacokinetic parameters of GMG after oral administration of G. sylvestre extract.

Simultaneous quantification of metformin and glipizide in human plasma by high‐performance liquid chromatography–tandem mass spectrometry and its application to a pharmacokinetic study

A selective, sensitive and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and validated to determine metformin and glipizide simultaneously in human plasma using phenacetin as internal standard (IS). After one-step protein precipitation of 200 μL plasma with methanol, metformin, glipizide and IS were separated on a Kromasil Phenyl column (4.6 × 150 mm, 5 µm) at 40°C with an isocratic mobile phase consisting of methanol-10 mmol/L ammonium acetate (75:25, v/v) at a flow rate of 0.35 mL/min. Electrospray ionization source was applied and operated in the positive mode. Multiple reaction monitoring using the precursor → product ion combinations of m/z 130 → m/z 71, m/z 446 → m/z 321 and m/z 180 → m/z 110 were used to quantify metformin, glipizide and IS, respectively. The linear calibration curves were obtained over the concentration ranges 4.10-656 ng/mL for metformin and 2.55-408 ng/mL for glipizide. The relative standard deviation of intra-day and inter-day precision was below 10% and the relative error of accuracy was between -7.0 and 4.6%. The presented HPLC-MS/MS method was proved to be suitable for the pharmacokinetic study of metformin hydrochloride and glipizide tablets in healthy volunteers after oral administration.

Development and validation of a rapid high-performance liquid chromatography-tandem mass spectrometry method for the determination of WJ-38, a novel aldose reductase inhibitor, in rat plasma and its application to a pharmacokinetic study

WJ-38 is an aldose reductase inhibitor that is being developed for the treatment of diabetic complications. The present paper describes a sensitive and specific liquid chromatography-tandem mass spectrometry method for the determination of WJ-38 in rat plasma. Partial denaturation of plasma proteins with methanol followed by liquid–liquid extraction using ethyl acetate was used to extract strongly protein-bound WJ-38 from rat plasma. Chromatographic separation was performed on an Inertsil ODS-3 column with an isocratic mobile phase consisting of acetonitrile, water and formic acid (75:25:0.125, v/v/v). Mass spectrometric detection was achieved by a triple-quadrupole mass spectrometer equipped with an ESI interface operating in positive ionization mode. Quantitation was performed using selected reaction monitoring of precursor-product ion transitions at m/z 392→246 for WJ-38 and m/z 446→321 for glipizide (internal standard). A linear calibration curve was obtained over the concentration range of 10.0–10,000ng/mL for WJ-38 in rat plasma. The intra- and inter-day precisions were less than 13.6% and the accuracy was within ±5.3%. The extraction recovery of WJ-38 from rat plasma was over 66.0%. The validated method has been successfully applied to a pharmacokinetic study in rats after intragastrical administration of WJ-38.

LC‐ESI‐MS/MS determination of paclitaxel on dried blood spots

A simple and rapid high-performance liquid chromatography-tandem mass spectrometric assay for determination of paclitaxel on rat dried blood spots was developed and validated. The extracted sample was chromatographed without further treatment using a reverse-phase Oyster ODS3, 4.6 × 50 mm, 3 µm column with mass spectrometry detection. The mobile phase comprised of acetonitrile-water, 60:40 v/v, with a flow rate of 0.4 mL/min was used. The calibration was linear over the range 0.2-20 ng/mL. The limits of detection and quantification were 0.08 and 0.2 ng/mL, respectively. The intra- and inter-day precision (CV%) and accuracy (relative error %) were less than 10 and 12%, respectively.

Development and Validation of a Sensitive and Rapid Method to Determine Naratriptan in Human Plasma by LC-ESI-MS-MS: Application to a Bioequivalence Study

A simple, sensitive, selective, and rapid high-performance liquid chromatography-tandem mass spectrometry method is developed and validated for the quantitation of naratriptan, using sumatriptan as internal standard (IS). The method included liquid-liquid extraction of naratriptan and IS with methyl-tert-butyl ether and dichloromethane mixture from 100 μL human plasma. The chromatographic separation is achieved on ACE C18 (50 mm × 2.1 mm, 5 μm) analytical column under isocratic conditions, using 0.1% acetic acid and acetonitrile (15:85, v/v) at a flow-rate of 0.4 mL/min. The precursor → product ion transitions for naratriptan (m/z 336.10 → 98.06) and IS (m/z 296.09 → 251.06) were monitored on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring (MRM) and positive ion mode. The linearity of the method for naratriptan is determined in the range of 103-20690 pg/mL with the analysis time of 1.5 min. The method is fully validated according to USFDA guidelines. A systematic post-column infusion study is conducted for ion-suppression due to endogenous matrix components. The process efficiency of analyte (96%) and IS (93%) from spiked plasma samples was consistent and reproducible. The application of the method is demonstrated by a bioequivalence study of 2.5 mg naratriptan tablet formulation in 31 healthy volunteers under fasting conditions.

Simultaneous determination of 14 β-lactam antibiotics in cosmetic products by liquid chromatography tandem mass spectrometry method

Abstract In this study, a simple and rapid high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method was established and validated to determine the 14 β -lactam antibiotics in cosmetic products, including 1 (ceftazidime), 2 (cefaclor), 3 (cefdinir), 4 (ampicillin), 5 (cefalexin), 6 (ceftezole), 7 (cefotaxim), 8 (cefradine), 9 (cefuroxime), 10 (cephazoline), 11 (cefathiamidine), 12 (cefoperazone), 13 (cafalotin), 14 (piperacillin).

Rapid high-performance liquid chromatography–tandem mass spectrometry method for determination of pentoxifylline and its active metabolites M1 and M5 in human plasma and its application in bioavailability study

A new rapid, sensitive and selective liquid chromatography coupled with mass spectrometry method was developed and validated for the simultaneous quantification of pentoxifylline (PTX) and two major active metabolites in human plasma (M1 and M5). After a deproteinization step, chromatographic separation of the selected analytes was performed on a RP-C18 column. The detection of target compounds was in multiple reaction monitoring mode using an ion trap mass spectrometer equipped with an electrospray ion source. The method was validated and proved to be linear, accurate and precise over the range 5.08–406.14, 10.08–806.40 and 20.15–1611.60 ng/mL in case of PTX, M1 and M5, respectively. The major advantages of this method are the small sample volume, simple sample processing technique, the high sensitivity and the very good selectivity guaranteed by the MS/MS (in case of PTX) or MS/MS/MS (in case of M1 and M5) detection. The validated method has been successfully applied to a bioequivalence study.

Assay of Colistin and Colistin Methanesulfonate in Plasma and Urine by Liquid Chromatography-Tandem Mass Spectrometry

A rapid high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed for the routine quantification of colistins A and B and their prodrugs, colistin methanesulfonate (CMS) A and CMS B, respectively, in human plasma and urine by using polymyxin B1 as the internal standard (IS). CMS concentrations were determined indirectly by subtracting the colistin concentrations determined in biological samples from the whole colistin concentrations determined after sample treatment with sulfuric acid in order to hydrolyze CMS into colistin. After extraction on a solid-phase extraction column, the colistins were separated on an XBrigde C(18) column with isocratic elution (run time, 3.8 min). The mobile phase was 0.1% (vol/vol) formic acid in acetonitrile-0.1% (vol/vol) formic acid in water (20:80, vol/vol), run at a 0.2-ml/min flow rate. Ions were detected in the turbo-ion-spray-positive and multiple-reaction-monitoring modes. The ions monitored (precursor [M + 2H](2+) to product ions) were m/z 585.5/101.2 for colistin A, m/z 578.5/101.2 for colistin B, and m/z 602.5/241.2 for IS. Prevalidation studies demonstrated the stability of CMS in biological samples and extracts, a key point for the reliable quantification of colistin and CMS. The assay was accurate and reproducible for the quantification of colistins A and B and CMSs A and B in plasma samples over concentration ranges appropriate for pharmacokinetic studies: 0.024 to 6.144, 0.015 to 3.856, 0.029 to 7.492, and 0.010 to 2.508 microg/ml, respectively. In urine samples, the assay was validated over the same concentration ranges for colistins and over concentration ranges of 0.058 to 7.492 microg/ml and 0.020 to 2.508 microg/ml for CMSs A and B, respectively.

Rapid high‐performance liquid chromatography–tandem mass spectrometry method for simultaneous measurement of venlafaxine and O‐desmethylvenlafaxine in human plasma and its application in comparative bioavailability study

A rapid high-performance liquid chromatography-tandem mass spectrometry method has been developed and validated for simultaneous measurement of venlafaxine and O-desmethylvenlafaxine in human plasma using fluoxetine as an internal standard. In the liquid-liquid extraction method, compounds and internal standard were extracted from plasma using methyl tertiary butyl ether as an extraction solvent. The HPLC separation of the analytes was performed on a Zorbax SB-C(18), 50 x 4.6 mm, 5 microm column, using a isocratic elution program using a mobile phase consisting of HPLC-grade methanol: 5 mm ammonium acetate (80:20 v/v) at a flow-rate of 1.0 mL/min with a total runtime of 3.0 min. The proposed method has been validated with a linear range of 4-400 ng/mL for venlafaxine and 5-500 ng/mL for O-desmethyl venlafaxine. The method was applied for a bio-equivalence study of 75 mg tablets formulation in 32 Indian male healthy subjects under fasting conditions.

A High-Performance Liquid Chromatography-Mass Spectrometry Method Using a Novel Atmospheric Pressure Chemical Ionization Approach for the Rapid Simultaneous Measurement of Tacrolimus and Cyclosporin in Whole Blood

Concentration monitoring and dose individualization is required to optimize either tacrolimus or cyclosporin therapy. In this study, the validation of a simple, rapid high-performance liquid chromatography-tandem mass spectrometry method for the simultaneous measurement of tacrolimus and cyclosporin in whole blood is reported. Blood samples (100 microL) were prepared by protein precipitation with zinc sulphate followed by acetonitrile (containing the internal standards ascomycin and cyclosporin D). The chromatographic run time was 1.5 minutes per sample. Mass spectrometric detection was by selected reaction monitoring with an atmospheric pressure chemical ionization source in negative ionization mode (tacrolimus: m/z 802.5 --> 560.6, cyclosporin: m/z 1200.8 --> 1088.4). The assay had an analytical range of 1.0 to 30 mug/L (r > 0.998, n = 6) for tacrolimus and 25 to 2000 microg/L (r > 0.999, n = 6) for cyclosporin. Tacrolimus inter- and intraday inaccuracy and precision [coefficient of variation (CV)] using quality control samples (2.5, 12.5, 25 microg/L) was less than +/-10.0% and CV less than 5.0%, respectively (n = 5). Similarly, cyclosporin inter- and intraday inaccuracy and precision using quality control samples (70, 400, 1500 microg/L) was less than +/-2.0% and CV less than 5.0%, respectively (n = 5). The lower limit of quantification for tacrolimus was 1.0 mug/L and cyclosporin 25 microg/L. The assay had an absolute mean recovery of 86.7% for tacrolimus and 89.0% for cyclosporin (n = 15). Intersubject variability, as a measure of potential matrix effects on results, was less than 6.0% CV for both analytes (n = 15). Extracted samples were stable for at least 20 hours. Results obtained from external proficiency testing samples measured by this method compared with the mean of all liquid chromatography-tandem mass spectrometry methods used by scheme participants revealed a strong correlation and good agreement for tacrolimus (r = 0.993, mean bias = -10.3%, n = 19) and cyclosporin (r = 0.996, mean bias = 3.0%, n = 20). In conclusion, this is the first reported high-throughput method that uses negative atmospheric pressure chemical ionization for the simultaneous measurement of tacrolimus and cyclosporin in whole blood.

Rapid and sensitive liquid chromatography tandem mass spectrometry method for the quantification of ambroxol in human plasma

A sensitive, specific and rapid high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was described and validated for the quantification of ambroxol in human plasma using enalaprilat as the internal standard (IS). Chromatographic separation was performed on a Lichrospher CN column with a mobile phase of methanol and water (containing 0.1% formic acid) (70:30, v/v). The total run time was 5.0 min for each sample. The analytes was detected by mass spectrometry with electrospray ionization source in positive selected reaction monitoring mode. The precursor-fragment ion reaction for ambroxol was m/z 378.9 --> 263.8, and for IS was m/z 349.0 --> 205.9. The linearity was established over the concentration range of 1.56-400.00 ng/mL. The inter-day and the intra-day precisions were all within 10%. A simple protein precipitation with methanol was adopted for sample preparation. The extraction recoveries of ambroxol and IS were higher than 90.80%. The validated method was successfully applied in pharmacokinetic study after oral administration of 90 mg ambroxol to 24 healthy volunteers.