A rapid, sensitive and specific liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method for the determination of clotrimazole in human plasma was developed and validated. The plasma samples were extracted with a mixed solvent of methyl tert-butyl ether–dichloromethane (4:1, v/v), and estazolam was selected as the internal standard. Then, the separation was carried out on a Phenomenex Luna CN column (2.0 × 150 mm, 5 μm), using a mobile phase of methanol-0.1% aqueous formic acid solution (85:15, v/v). A triple quadrupole mass spectrometer with the positive ionization mode was used for the determination of target analytes. The monitored ion transitions were m/z 276.9 → 165.1 for clotrimazole and m/z 294.9 → 266.9 for estazolam, respectively. The calibration curve of clotrimazole was established over the range of 0.01563–1.000 ng mL−1 (r2 = 0.9935). The intra- and inter-day precisions were less than 10% and all the biases were not more than 9%. The mean extraction recovery of clotrimazole was greater than 68.4% and no significant matrix effect was detected. The LLOQ of 0.01563 ng mL−1 is sensitive enough to perform pharmacokinetic studies after clotrimazole administration.

A simple and sensitive method of gas chromatography–mass spectrometry for quantification of β-elemene in human plasma was developed and validated. An n-hexane-based liquid–liquid extraction assisted by saturated sodium chloride solution was used for sample preparation. Naphthalene was chosen as the internal standard. The calibration curve (r2 = 0.9997) of β-elemene was established at the range of 0.0655–40.00 μg mL−1 and the lower limit of quantification (LLOQ) was 0.0655 μg mL−1 using 200 μL of plasma, respectively. The accuracy was in a range of 87.1–101.7 %, and intra- and inter-day precisions (RSD %) at three QC levels were all less than 10 %. The LLOQ of 0.0665 μg mL−1 is sensitive enough to perform pharmacokinetic studies after β-elemene administration.

A novel, simple and reliable bioanalytical method of high performance liquid chromatography for simultaneous quantification of nelarabine and its active metabolite 9-β-d-arabinofuranosyl-guanine (Ara-G) in human plasma was developed and validated. The calibration curves for nelarabine and Ara-G were linear over concentration ranges of 0.1562–10 and 0.6250–40 μg mL−1, respectively. Intra- and inter-day precision and accuracy results satisfied the acceptable criteria for bioanalytical method validation. This method has been applied to a pharmacokinetic study involving 20 T-cell lymphoblastic leukemia/lymphoma patients.

•We established a LC-ESI-MS/MS method to determine simotinib, a novel TKIs.•The method was applied to a pharmacokinetic study in 12 healthy Chinese volunteers.•The clinical pharmacokinetic parameters of simotinib were reported for the first time.Simotinib is a novel epidermal growth factor receptor tyrosine kinase inhibitor. This study presented a sensitive and specific liquid chromatography-electrospray ionization-mass spectrometry method using erlotinib as internal standard for the determination of simotinib in human plasma. The method involved a simple liquid–liquid extraction using diethyl ether. The analytes were separated with isocratic gradient elution on an Agilent TC-C18 column (4.6 × 150 mm, 5 μm). Mass spectrometric detector equipped with electrospray ionization source was carried out in the mode of multiple reaction monitoring (MRM). The monitored transitions were m/z 501.2 → 182.1 for simotinib and m/z 394.4 → 278.1 for erlotinib. The calibration curve of simotinib was established over the range of 2.058–3000 μg L−1 (r2 = 0.9924). The intra- and inter-day precisions were all less than 10%, and all the biases were not more than 7%. This validated method was then successfully applied to a pharmacokinetic study involving twelve healthy Chinese volunteers. The mean Cmax and Tmax for simotinib were 254.79 ± 98.30 μg L−1 and 1.71 ± 0.48 h, respectively. Plasma concentrations declined with a t1/2 of 5.37 ± 2.32 h. AUC0–t and AUC0→∞ values obtained were 1262.59 ± 501.41 μg L−1 h and 1329.95 ± 517.42 μg L−1 h, respectively.

A novel simple bioanalytical method of high performance liquid chromatography (HPLC) for simultaneous quantification of morinidazole and its carbonylation metabolite (M1) in human plasma was developed and validated. The calibration curves for morinidazole and M1 were linear over concentration ranges of 23.99–1.464 × 104 μg L−1 and 5.407–3300 μg L−1, respectively. Intra- and inter-day precision and accuracy results satisfied the acceptable criteria for bioanalytical method validation. This method could be a useful method for clinical pharmacokinetic studies of morinidazole and its carbonylation metabolite, and it has been applied to a pharmacokinetic study involving 11 renal insufficiency patients and 11 healthy volunteers.Graphical abstractA HPLC method for simultaneous determination of morinidazole and its carbonylation metabolite in human plasma was established and applied to a clinical pharmacokinetic study involving 11 renal insufficiency patients (Group A) and 11 healthy volunteers (Group B).Highlights► We established a method to determine morinidazole and its carbonylation metabolite. ► The method was applied to a clinical study involving renal insufficiency patients. ► We compared pharmacokinetic parameters of morinidazole in two groups of subjects.

Biological sample pretreatment is an important step in biological sample analysis. Due to the diversity of biological matrices, the analysis of target substances in these samples presents significant challenges to sample processing. To meet these emerging demands on biopharmaceutical analysis, this paper summarizes several new techniques of on-line biological sample processing: solid phase extraction, solid phase micro-extraction, column switching, limited intake filler, molecularly imprinted solid phase extraction, tubular column, and micro-dialysis. We describe new developments, principles, and characteristics of these techniques, and the application of liquid chromatography–mass spectrometry (LC–MS) in biopharmaceutical analysis with these new techniques in on-line biological sample processing.This paper summarizes several new techniques of on-line biological samples processing in recent years (such as solid phase extraction, solid phase micro-extraction, column switching technique, limit intake filler technique, molecular imprinted solid phase extraction technique, tubular column technique and micro-dialysis technique), and describes their developments, principles, characteristics and application matched with liquid chromatography–mass spectrometry (LC–MS) in biopharmaceutical analysis in on-line biological sample processing field.Download full-size image

Zidovudine (AZT), the first drug approved by the US Food and Drug Administration for the treatment of human immunodeficiency virus (HIV) infection, is metabolized in the host cells to 5′-AZT triphosphate (AZT-TP) which inhibits HIV reverse transcriptase. As the pharmacokinetics of AZT and its phosphorylated metabolites in human peripheral blood mononuclear cells (hPBMCs) is limited, the aim of this study was to determine the pharmacokinetic parameters of AZT and its phosphorylated metabolites in hPBMCs from 12 healthy Chinese male subjects after a single oral dose of 600 mg of AZT. Blood samples were collected prior to drug administration, then at 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8 and 10 h after drug administration. Mononuclear cells collected by Ficoll-Hypaque density gradient centrifugation were used for determination of AZT and metabolites [AZT monophosphate (AZT-MP), AZT diphosphate (AZT-DP) and AZT-TP] and the plasma was used to evaluate the pharmacokinetics of AZT. Plasma concentration of AZT peaked within 0.583 h and intracellular concentrations of AZT, AZT-MP, AZT-DP and AZT-TP peaked within 1.083, 1.500, 1.417 and 1.583 h, respectively. AZT in plasma was eliminated rapidly with t1/2 of 2.022 h, and AZT-MP, AZT-DP and AZT-TP were eliminated with t1/2 of 13.428, 8.285 and 4.240 h, respectively. The plasma concentration of the phosphorylated metabolites was not quantifiable.A pharmacokinetic experiment has been performed to evaluate the pharmacokinetics of zidovudine in plasma and pharmacokinetics of zidovudine and its phosphate metabolites in human peripheral blood mononuclear cells.Download full-size image

A rapid, specific and sensitive HPLC–MS/MS method was developed and validated for the determination of 20(S)-protopanaxadiol (PPD) in human plasma. PPD and the internal standard PD were extracted from plasma by liquid–liquid extraction with cyclohexane–methylene dichloride (2:1, v/v). The separation was performed on a HyPURIYTY C18 column using methanol–5 mM ammonium formate (90:10, v/v) as mobile phase at a flow rate of 0.35 mL/min. Mass spectrometric detection was carried out by electrospray ionization (ESI) in the positive ion mode using multiple reaction monitoring (MRM). The monitored transitions were m/z 425.4→217.2 for PPD and at m/z 461.4→425.5 for PD. The method was linear over the range 0.512–100 ng/mL with a lower limit of quantification (LLOQ) of 0.512 ng/mL. The mean extraction recovery of PPD was greater than 78.2% and no significant matrix effect was detected. The intra- and inter-day precisions were less than 10% and the biases below 4% for PPD. The validated method was applied to a three-level single-dose clinical pharmacokinetics study of 12 healthy Chinese volunteers and the main pharmacokinetic parameters of PPD were obtained.An LC–ESI-MS/MS method for determination of 20(S)-protopanaxadiol in human plasma was established and validated. It is further applied to a clinical pharmacokinetic study of 12 healthy Chinese volunteers after a single oral dose of 20(S)-protopanaxadiol. Download full-size image

A rapid, sensitive and specific liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method for the determination of clotrimazole in human plasma was developed and validated. The plasma samples were extracted with a mixed solvent of methyl tert-butyl ether–dichloromethane (4:1, v/v), and estazolam was selected as the internal standard. Then, the separation was carried out on a Phenomenex Luna CN column (2.0 × 150 mm, 5 μm), using a mobile phase of methanol-0.1% aqueous formic acid solution (85:15, v/v). A triple quadrupole mass spectrometer with the positive ionization mode was used for the determination of target analytes. The monitored ion transitions were m/z 276.9 → 165.1 for clotrimazole and m/z 294.9 → 266.9 for estazolam, respectively. The calibration curve of clotrimazole was established over the range of 0.01563–1.000 ng mL−1 (r2 = 0.9935). The intra- and inter-day precisions were less than 10% and all the biases were not more than 9%. The mean extraction recovery of clotrimazole was greater than 68.4% and no significant matrix effect was detected. The LLOQ of 0.01563 ng mL−1 is sensitive enough to perform pharmacokinetic studies after clotrimazole administration.