Feifan Xie

In vitro dissolution similarity factor (f2) and in vivo bioequivalence criteria, how and when do they match? Using a BCS class II drug as a simulation example

The present study examined the agreement between in vitro dissolution f2 similarity and in vivo bioequivalence criteria for BCS class II drugs. Dissolution test profiles were generated using the First-order model with varied dissolution parameters around the standard values of a reference profile. The in vivo curves were derived from in vitro dissolution profiles with the drugs pharmacokinetics parameters by numerical convolution method. The Cmax, Tmax, AUC0-t and AUC0-∞ obtained from in vivo test and reference concentration-time curves were compared, and the CmaxR (Cmax ratio), TmaxDif (Tmax difference), AUC0-tR (AUC0-t ratio) and AUC0-∞R (AUC0-∞ ratio) were determined. The relationships between CmaxR, AUC0-tR, AUC0-∞R, f2 and the First-order model parameters demonstrated that the Similarity Region 1 enclosed by the f2 contour line labeled 50 was completely within the Bioequivalence Region enclosed by the contour lines labeled 0.80 and 1.20 of AUC0-tR, AUC0-∞R, and CmaxR, and the Similarity Region 2 enclosed by the f2 contour line labeled 35 was nearly overlapped with the Bioequivalence Region, but did not exactly match. The results indicate that the public standard for in vitro dissolution f2 similarity criterion (f2⩾50) is probably slightly conservative and may be widened to an appropriate lower critical value.

New techniques of on-line biological sample processing and their application in the field of biopharmaceutical analysis

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.

Simultaneous determination of bendamustine and its active metabolite, gamma-hydroxy-bendamustine in human plasma and urine using HPLC-fluorescence detector: Application to a pharmacokinetic study in Chinese cancer patients

A simple, sensitive and cost-effective assay based on reversed phase high performance liquid chromatography (RP-HPLC) with isocratic mode for simultaneous determination of bendamustine (BM) and its active metabolite, gamma-hydroxy-bendamustine (γ-OH-BM) in human plasma and urine was developed and validated. Sample preparation involved protein precipitation by 10% perchloric acid-methanol solution. The peaks were recorded by using fluorescence detector (excitation wavelength 328 nm and emission wavelength 420 nm). The calibration curves were linear over concentration ranges of 8.192-10,000 ng mL(-1) and 5-1,000 ng mL(-1) for BM in human plasma and urine as well as 10-1,000 ng mL(-1) and 5-1,000 ng mL(-1) for γ-OH-BM in human plasma and urine, respectively. Intra- and inter-run precisions of BM and γ-OH-BM were less than 15% and the bias were within ± 15% for both plasma and urine. This validated method was successfully applied to a pharmacokinetic study enrolling 10 Chinese patients with indolent B-cell non-Hodgkin lymphoma and chronic lymphocytic leukemia administered a single intravenous infusion of 100 mg m(2) bendamustine hydrochloride.

Pharmacokinetic study of benfotiamine and the bioavailability assessment compared to thiamine hydrochloride

Benfotiamine is a lipid‐soluble thiamine precursor which can transform to thiamine in vivo and subsequently be metabolized to thiamine monophosphate (TMP) and thiamine diphosphate (TDP). This study investigated the pharmacokinetic profiles of thiamine and its phosphorylated metabolites after single‐ and multiple‐dose administration of benfotiamine in healthy Chinese volunteers, and assessed the bioavailability of orally benfotiamine administration compared to thiamine hydrochloride. In addition, concentration of hippuric acid in urine which is produced in the transformation process of benfotiamine was determined. The results showed that thiamine and its phosphorylated metabolites exhibited different pharmacokinetic characteristics in plasma, blood and erythrocyte, and one‐compartment model provided the best fit for pharmacokinetic profiles of thiamine. The transformation process of benfotiamine to thiamine produced large amount of hippuric acid. No accumulation of hippuric acid was observed after multiple‐dose of benfotiamine. Compared to thiamine hydrochloride, the bioavailability of thiamine in plasma and TDP in erythrocyte after oral administration of benfotiamine were 1147.3 ± 490.3% and 195.8 ± 33.8%, respectively. The absorption rate and extent of benfotiamine systemic availability of thiamine were significantly increased indicating higher bioavailability of thiamine from oral dose of benfotiamine compared to oral dose of thiamine hydrochloride.

Intracellular pharmacokinetic study of zidovudine and its phosphorylated metabolites

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.

Development and validation a LC-MS/MS method for the simultaneous determination of agomelatine and its metabolites, 7-desmethyl-agomelatine and 3-hydroxy-agomelatine in human plasma: Application to a bioequivalence study.

A novel sensitive and selective LC-MS/MS method for the determination of agomelatine, 7-desmethyl-agomelatine and 3-hydroxy-agomelatine in human plasma was developed and validated. After simple protein precipitation, the analytes were separated on a Phenomenex ODS3 column (4.6×150 mm, 5μm, Phenomenex, USA) with mobile phase consisted of methanol and 5mM ammonium formate solution (containing 0.2% formic acid) at a ratio of 70:30 (v/v) with a flow rate of 0.8mL/min. The MS acquisition was performed in multiple reactions monitoring (MRM) mode with a positive electrospray ionization source. The mass transitions monitored were m/z 244.1→185.1, m/z 230.1→171.1, m/z 260.1→201.1 and m/z 180.1→110.1 for agomelatine, 7-desmethyl-agomelatine, 3-hydroxy-agomelatine and internal standard (phenacetin), respectively. The method was validated for specificity, linearity and lower limit of quantification, precision and accuracy, extraction recovery, matrix effect and stability. The calibration curves for agomelatine, 7-desmethyl-agomelatine and 3-hydroxy-agomelatine in human plasma were linear over concentration ranges of 0.0457-100ng/mL, 0.1372-300ng/mL and 0.4572-1000ng/mL, respectively. Intra- and inter-day precisions and accuracies data met the acceptance criteria of FDA guideline for bioanalytical method validation. The developed method has been successfully applied to a bioequivalence study in healthy Chinese volunteers.

Development and validation of a simple HPLC assay for the quantitation of SM-1, a novel derivative of the PAC-1 anticancer agent, and an initial pharmacokinetics study in rats

A simple and specific HPLC-UV method was developed and validated for the determination of SM-1 in rat plasma. The bioanalytical procedure involved extraction of SM-1 and gefitinib (internal standard, IS) from a 100 μL plasma aliquot with simple protein precipitation with methanol. Chromatographic separation was achieved using an Agilent TC-C18 column (4.6 mm × 150 mm, 5 μm) with an isocratic mobile phase consisting of 10 mM potassium hydrogen phosphate solution (pH 7.0, adjusted by using 10% phosphoric acid)–acetonitrile (37 : 63, v/v) at a flow-rate of 1.0 mL min−1. The UV detection wavelength was 282 nm. SM-1 and IS eluted at 3.6 and 7.0 min, respectively, with a total run time of 8 min. Method validation was performed according to US Food and Drug Administration bioanalytical guidelines and the results met the acceptance criteria. The calibration curve of SM-1 in rat plasma was linear over the concentration range of 0.1–20 μg mL−1. Intra- and inter-run precisions of SM-1 were less than 6.1% and the biases were within ±10.0%. The method was successfully applied to the pharmacokinetics study after intravenous and oral administration of SM-1 in rats.

A sensitive LC-ESI-MS/MS method for the determination of clotrimazole in human plasma

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.

LC-ESI-MS/MS determination of simotinib, a novel epidermal growth factor receptor tyrosine kinase inhibitor: Application to a pharmacokinetic study

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) (r(2)=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.

Determination of Strong Acidic Drugs in Biological Matrices: A Review of Separation Methods

Strong acidic drugs are a class of chemical compounds that normally have high hydrophilicity and large negative charges, such as organophosphatic compounds and organosulphonic compounds. This review focuses on sample preparation and separation methods for this group of compounds in biological matrices in recent years. A wide range of separation techniques, especially chromatographic method, are presented and critically discussed, which include liquid chromatography (e.g., ion-pair and ion-exchange chromatography), capillary electrophoresis (CE), and other types. Due to the extremely low concentration level of target analytes as well as the complexity of biological matrices, sample pretreatment methods, such as dilute and shoot methods, protein precipitation (PP), liquid-liquid extraction (LLE), solid-phase extraction (SPE), degradation, and derivatization strategy, also play important roles for the development of successful analytical methods and thus are also discussed.