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  • Chemical profiling of Callicarpa nudiflora and its effective compounds identification by compound-target network analysis
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-15
    Yong-Sheng Wu; Liu Shi; Xin-Guang Liu; Wei Li; Rui Wang; Sheng Huang; Yi Li; Dong-Lan Yan; Hui-Ying Wang; Yuan Tian; Yan-Ming Chen; Hua Yang

    Callicarpa nudiflora, belonging to the family Verbenaceae, is widely used to treat inflammation caused by bacterial infection.However, the underlying active substances of C. nudiflora against inflammation remains obscure. In this work, an ultra high-performance liquid chromatography (UHPLC) coupled with quadrupole time-of-flight mass spectrometry method was developed to characterize the ingredients in C. nudiflora, and a validated UHPLC coupled with triple quadrupole tandem mass spectrometry method was applied to quantify major components. As a result, a total of 96 chemical compounds were identified in C. nudiflora, and 26 compounds of them were further quantified in 34 batches of C. nudiflora. Based on the identified components from C. nudiflora, a compound-target network for the anti-inflammation effect was constructed by reverse docking target prediction, disease associated genes screening in DisGeNET and the protein-protein interaction from STRING. The compound-target network showed that C. nudiflora might exert anti-inflammation effect on the target of complement 3 and 5 in the pathway of cells and molecules involved in local acute inflammatory response, and 16 effective candidate compounds were found such as catalpol, acteoside, rutin, etc. This study provided an opportunity to deepen the understanding of the chemical composition and the potential anti-inflammatory mechanism of C. nudiflora.

    更新日期:2020-01-15
  • Spectrochemical identification of kanamycin resistance genes in artificial microbial communities using Clover-assay
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-15
    Naifu Jin; Camilo L.M. Morais; Francis L. Martin; Dayi Zhang
    更新日期:2020-01-15
  • Development, validation and application of a new HPLC-DAD method for simultaneous quantification of apixaban, dabigatran, edoxaban and rivaroxaban in human plasma
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-14
    Filipa Gouveia; Joana Bicker; Joana Santos; Marília Rocha; Gilberto Alves; Amílcar Falcão; Ana Fortuna
    更新日期:2020-01-14
  • Simultaneous determination of fluoxetine, venlafaxine, vortioxetine and their active metabolites in human plasma by LC-MS/MS using one-step sample preparation procedure
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-11
    Martin Kertys; Michaela Krivosova; Igor Ondrejka; Igor Hrtanek; Ingrid Tonhajzerova; Juraj Mokry

    The aim of antidepressant therapy is to induce remission and prevent relapses of major depressive disorder with minimum adverse effects during the treatment. Due to high variability in metabolism, therapeutic drug monitoring is recommended as a useful tool for individualisation of the therapy. For this purpose, we have developed simple and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for quantification of fluoxetine (FLX), venlafaxine (VEN), vortioxetine (VTX) and their active metabolites norfluoxetine (NFLX) and O-desmethylvenlafaxine (ODV). After one-step extraction procedure using Waters OSTRO plate, analytes were separated by gradient elution on Acquity UPLC BEH C18 (50 × 2.1 mm, 1.7 μm) with runtime 4.2 min. The detection was done on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode with transitions at m/z 310.23 → 148.20 for FLX, m/z 296.23 → 134.20 for NFLX, m/z 278.31 → 121.13 for VEN, m/z 264.31 → 107.14 for ODV and m/z 299.19 → 150.05 for VTX using a positive electrospray ionisation interface. The method was successfully validated according to the European Medicine Agency guideline for the selectivity, linearity and lower limit of detection, precision and accuracy, matrix effect, extraction recovery, carry over, dilution integrity and stability over a concentration range of 1–300 ng/mL for FLX, NFLX, VEN, ODV and 0.2–100 ng/mL VTX. Extraction recovery for each analyte was > 80%, and no significant matrix effects were observed. The developed method was employed for quantification of antidepressants in clinical samples from patients treated with either FLX, VEN, or VTX.

    更新日期:2020-01-13
  • 更新日期:2020-01-11
  • The Simultaneous Quantification of Phytosterols And Tocopherols in Liposomal Formulations Using Validated Atmospheric Pressure Chemical Ionization- Liquid Chromatography –Tandem Mass Spectrometry
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-09
    Asmita Poudel; George Gachumi; Ildiko Badea; Zafer Dallal Bashi; Anas El-Aneed

    A novel liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed and validated to simultaneously quantify phytosterols (brassicasterol, campesterol, stigmasterol and β-sitosterol) and tocopherols (alpha, beta, gamma and delta) entrapped in the lipid bilayer of a liposomal formulation. Atmospheric pressure chemical ionization (APCI) was employed due to the enhanced ionization of phytosterols and tocopherols in comparison to electrospray ionization. Unlike published work, the chromatographic conditions were modified to simplify the analytical approach. For the first time, a simple isocratic elution (acetonitrile:methanol 99:1 v/v) was utilized for the separation of four phytosterols and four tocopherols in a single run. A substantially better baseline separation of phytosterols were obtained in comparison to reported methods by using poroshell C18 column. The method has a total run time of 7 minutes, which is the shortest run time among all reported quantitative methods for the simultaneous determination of four phytosterols and four tocopherols. Calibration curves for all phytosterols were linear in the range of 0.05-10 µg/mL. In the case of tocopherols, alpha tocopherol showed linear response in the range of 0.25-10 µg/mL. However, gamma and delta tocopherols exhibited quadratic relationship in the same concentration range (0.25-10 µg/mL). Validation parameters met the International Conference on Harmonization (ICH) guidelines in terms of selectivity, accuracy, precision, repeatability, sensitivity, matrix effects, dilution integrity and stability. The method was, for the first time, successfully applied for the quantifying phytosterols and tocopherols entrapped inside liposomes. An interesting chromatographic phenomenon was observed during sample analysis. Alpha tocopherol (entrapped in the liposomal lipid bilayer) was found to elute at two retention times, 2.53 minutes and 3.60 minutes. Such dual separation was not observed in calibration standards and quality controls. It was concluded that the chiral recognition ability of liposomes made up of phosphatidylcholine separated the enantiomers of alpha tocopherol, giving rise to two peaks at two different retention time. To sum, the reported novel LC-MS/MS method addresses three major analytical shortcomings, namely i)longer run time, ii)complex gradient elution and iii)poor baseline separation of phytosterols and tocopherols.

    更新日期:2020-01-09
  • Spectroscopic characterisation of a series of Salmonella Typhi Vi-diphtheria toxoid glycoconjugate antigens differing in polysaccharide-protein ratio
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-09
    Christopher Jones; So Jung An; Yeon Kyung Yoon; Sudeep Kothari; Sushant Sahastrabuddhe; Rodney Carbis

    Glycoconjugate vaccines consisting of the Salmonella enterica subsp. enterica serovar Typhi (S. Typhi) Vi capsular polysaccharide (PS) covalently attached to a suitable carrier protein have become available to support mass paediatric vaccination campaigns against typhoid. One developmental vaccine from the International Vaccine Institute (IVI) uses diphtheria toxoid (DTx) as the carrier protein. Several investigational conjugates with different PS:protein ratios were prepared, as previously reported by the IVI group, for physicochemical and immunochemical characterisation. We describe here the further spectroscopic characterisation of this series of glycoconjugate immunogen bulks using NMR spectroscopy, circular dichroism and absorption spectroscopy. We have used several mathematical approaches to extract information from the spectroscopic data not previously applied to glycoconjugates. These complementary approaches provide information on (i) the integrity of the carrier protein, (ii) consistency between batches of vaccine components, (iii) the polysaccharide: protein ratio (iv) the O-acetylation of the Vi in the conjugate (v) the stability of the O-acetylation of the Vi, and (vi) the presence of residual process reagents in the bulk. The utility of the data analysis approaches is discussed. Together, these analytical methods provide important characterisation of Vi-DTx conjugates to support development and quality control of commercial products.

    更新日期:2020-01-09
  • Stability of Sodium Ascorbyl Phosphate in the Water-Glycerol System
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-09
    Xiaohui Dong; Ting Zhang; Hongyuan Wei; Leping Dang

    Sodium ascorbyl phosphate is a hydrophilic derivative of ascorbic acid with better stability compared to the parent compound. However, sodium ascorbyl phosphate is not as stable in solution as it is in the solid state, and it has been found to degrade, with accompanying discoloration, under the influence of different conditions. Here, the degradation mechanism of sodium ascorbyl phosphate in the water-glycerol system was revealed and the thermal degradation kinetics was shown to follow second-order kinetics. A thermal degradation prediction model was established and successfully fitted to the experimental data. In addition, the stability of sodium ascorbyl phosphate in the water-glycerol system during storage was investigated under different conditions, including changes in concentration, temperature, pH, light and oxygen, and metal ions. Sodium ascorbyl phosphate content was quantitatively measured via HPLC, and the color and pH values of the sample were qualitatively measured using a spectrophotometer and a pH meter, respectively. It was found that temperature and pH are the most important factors affecting the stability of sodium ascorbyl phosphate.

    更新日期:2020-01-09
  • Simultaneous determination of a promising anti-brain tumor agent CAT3 and its two major metabolites in mouse plasma and brain by a LC-MS/MS method
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-09
    Shengyu Zhao; Ru-Bing Wang; Jie Bai; Xiaoqing Fan; Minwan Hu; Baolian Wang; Jinping Hu; Yan Li

    A rapid and reproducible method with high selectivity was developed for simultaneous determination of a promising anti-brain tumor agent CAT3 and its two metabolites PF403 and GLU-PF403 in mouse plasma and brain. An economic deproteinization with septuple acetonitrile (v/v) was applied to pretreat the samples in this study. All analytes were well retained and separated on a CAPCELL CORE PC (2.7 μm, 2.1 mm I.D. × 150 mm, SHISEIDO Technologies) column with an eluting solvent of acetonitrile /water containing 0.1% formic acid (v/v) at the flow rate of 0.2 mL per minute. The detection was carried out on a Q Exactive high resolution mass spectrometer equipped with a HESI ion source in parallel reaction monitoring (PRM) mode. The corresponding transitions for quantitation were 434.23→ 70.07 for CAT3, 350.17→70.07 for PF403, 526.21→70.07 for GLU-PF403, 364.19→70.07 for IS-1 and 625.18→317.07 for IS-2, respectively. A well-linear fit curve was achieved among the range of 0.1∼50 ng/mL for CAT3, 0.2∼100 ng/mL for PF403 and 2.5∼600 ng/mL for GLU-PF403 both in mouse plasma and brain homogenate. The intra-/inter-day accuracies of three analytes were within ±14.5% and precisions were below to 13.44 %. The mean values of recovery of three compounds in mouse plasma and brain homogenate were among 98.06 ∼ 118.63% and 81.04∼108.69%. The analytes in NaF-treated ice cold blood of mouse was stable within tested 30 minutes. Plasma and brain homogenate samples had no obvious changes during all storage, sample treatment and analytic process of mouse plasma sample. The reproducible and reliable method was well employed to the research of CAT3 pharmacokinetic characteristics in mouse plasma and brain after a single intragastric administration at dose of 10 mg/kg.

    更新日期:2020-01-09
  • Simplification of affinity macroporous monolith microfluidic column synthesis and its ability for protein separation
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-08
    Ashish Khaparde; Kishore K.R. Tetala

    A generic multi-component approach was designed to perform simultaneous in situ polymerization and ligand immobilization to develop affinity porous polymer based chromatography resin in a facile mode. This strategy exploits the regioselective ring opening reaction between epoxy group of monomer and native functional group of ligand (i.e. amine) under aqueous condition (pH 9.7). As a proof-of-concept, reaction of iminodiacetic acid (IDA) with allyl glycidyl ether (AGE) in presence of other monomer (HEMA) and crosslinkers (DATD, PDA) for 4 h via thermal initiation process (temperature of 65 °C) was shown. Successful polymerization (both ex situ & in situ) was confirmed by visual observation, surface morphology of the polymer by scanning electron microscope and ligand immobilization by FT-IR analysis. Chelation of the metal-ion i.e. copper (Cu (II)) with IDA in the monolith showed IgG adsorption capacity (27.8 mg/g monolith) over IDA-monolith without metal-ion. The affinity column has shown efficient capture of high abundant proteins such as IgG, transferrin and albumin from human plasma.

    更新日期:2020-01-09
  • Characterization of bromelain indicates a molar excess of inhibitor vs enzyme molecules, a Jacalin-like lectin and Maillard reaction products
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-08
    Peter Gross; Holger Seelert; Peter Meiser; Rolf Müller

    The phytotherapeutic bromelain is a heterogeneous protein mixture, extracted from pineapple stem, with high proteolytic activity based on cysteine proteases. Its global protein chemical composition was analyzed qualitatively and quantitatively by SDS-PAGE and RP-HPLC. A SDS-PAGE method with elaborate sample pretreatment was developed, to cope with the bromelain's self-digestion properties and the hypothetical disulfide scrambling during electrophoresis. Both can produce misleading results, if not considered. RP-HPLC was applied for its high separation power for bromelain proteinaceous compounds. A peak identification and assignment to different protein classes in bromelain was done by enzyme kinetics and MS. The method was successfully applied for the quantitative determination of the molar ratio between inhibitor and enzyme and resulted to be approximately 3:2. Bromelain contains, from a molar point of view, inhibitor molecules as major component, which thus might be considered as a natural pharmaceutical excipient in Bromelain, because it protects the enzymes against autolysis. We described two methods to separate the inhibitor fraction from the enzyme fraction, RP-HPLC and size exclusion chromatography. A pineapple derived Jacalin-like-lectin, herein called ‘Anlec’, was identified and quantified by RP-HPLC-MS in bromelain and its content was determined to be 5 %, related to all proteins in bromelain. Anlec binds specifically to mannose-containing glycans and is discussed in literature to possess anti-HIV medical potential. Bromelain could therefore be a possible and economic source for the production of Anlec. An isolation strategy of Anlec from bromelain, in high purity, is shown is in this work. The presented RP-HPLC results are comprehensive in chemical information, and the method is expedient to provide appropriate bromelain protein isolations but also to accomplish quality control, covering all relevant protein components. It is furthermore shown, that proteins in bromelain may react with reducing sugars in a Maillard reaction to form glycated proteins. Maillard reaction products in bromelain are detected and characterized and could be responsible for the limited stability and storage times at room temperature of bromelain. Even the active center thiol group could be potentially glycated.

    更新日期:2020-01-08
  • On-Flow LC-MS/MS Method for Screening of Xanthine Oxidase Inhibitors
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-07
    Marili V.N. Rodrigues; Caio Rodrigues-Silva; Sinésio Boaventura; Adriana S.S. Oliveira; Susanne Rath; Quezia B. Cass

    The screening of compounds is the initial step in research for the development of new drugs. For this reason, the availability of fast and reliable tools for the screening of a large number of compounds becomes essential. Among the therapeutic targets, the enzyme xanthine oxidase (XO) is of great interest for its importance as a biological source of superoxide radicals, which contribute to the oxidative stress on organisms and are involved in many pathological processes. In the present study, we validated a new method using an immobilized capillary enzyme reactor in an LC system directly coupled to triple quadrupole mass spectrometry to screen for XO ligands. The use of mass spectrometry provided selectivity and speed to the system, eliminating the analytical separation step. The Michaelis-Menten constant (KM) value determined for the immobilized enzyme was 14.5 ± 0.4 μmol L-1, which is consistent with the value previously reported for the XO-ICER with UV detection in a 2D LC method. The on-line approach was successfully applied to assay the XO inhibitory activities of thirty isolated compounds from different classes of natural products and provided greater productivity (288 analysis/day) than 2D LC method (84 analysis/day) of screened samples.

    更新日期:2020-01-07
  • A sensitive nanocomposite design via carbon nanotube and silver nanoparticles: Selective probing of Emedastine Difumarate
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-03
    Hamideh Imanzadeh; Nurgul K Bakirhan; Biuck Habibi; Sibel A Ozkan
    更新日期:2020-01-04
  • Assessment method for deamidation in proteins using carboxylic acid derivatization-liquid chromatography-tandem mass spectrometry
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-03
    Shimba Kawasue; Yohei Sakaguchi; Reiko Koga; Hideyuki Yoshida; Hitoshi Nohta
    更新日期:2020-01-04
  • Chemical Analysis of Saffron by HPLC based Crocetin Estimation
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-03
    Chilakala Nagarjuna Reddy; Sandip B. Bharate; Ram A. Vishwakarma; Sonali S. Bharate
    更新日期:2020-01-04
  • Development and investigation of a QuEChERS-based method for determination of phthalate metabolites in human milk
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-02
    Jinyoung An; Ye-Yeong Kim; Hyun-Deok Cho; Junhee Kim; Ji-Youn Lee; Yeoreum Lee; Eunji Jo; Jaeick Lee; Sangwon Cha; Sang Beom Han

    Phthalates are commonly used as plasticizers and are known as risk factors toward several conditions such as cancer, birth defects, and endocrine disruption. Biomonitoring of phthalates is necessary to assess the potentially harmful effects of long-term exposure. In this work, we have developed a novel QuEChERS method to determine eight phthalate metabolites—mono-(3-carboxypropyl) phthalate, mono-(2-ethyl-5-carboxypentyl) phthalate, mono-(2-ethyl-5-hydroxyhexyl) phthalate, mono-(2-ethyl-5-oxohexyl) phthalate, mono-n-butyl phthalate, mono-benzyl phthalate, mono-(carboxyloctyl) phthalate, and mono-(carboxynonyl) phthalate—in human milk. The extraction process was optimized by comparing three different QuEChERS methods, and a further purification step was used to eliminate interferential lipid. In this process, several factors, such as the pH based on QuEChERS additive salts, acid dissociation constant, and distribution coefficient of the analyte, were found to have a significant effect on the extraction efficiency of the QuEChERS method. Target compounds were determined using liquid chromatography–tandem mass spectrometry equipped with electrospray ionization in the multiple-reaction monitoring mode. The developed method was verified by evaluating the selectivity, linearity, lower limit of quantification, accuracy, precision, and recovery, and applied to monitor real milk samples from 26 people. It is expected that the established method can be utilized not only to monitor phthalate metabolites in biological samples but also to identify the correlation between phthalate concentrations observed for the mother and the newborn.

    更新日期:2020-01-02
  • LC-MS/MS assay coupled with carboxylic acid magnetic bead affinity capture to quantitatively measure cationic host defense peptides (HDPs) in complex matrices with application to preclinical pharmacokinetic studies
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2020-01-02
    Maura J. O’Neill; King Chan; Jesse M. Jaynes; Zachary Knotts; Xia Xu; Abisola Abisoye-Ogunniyan; Theresa Guerin; Jerome Schlomer; Dandan Li; Jeffrey W. Cary; Kanniah Rajasekaran; Clayton Yates; Serguei Kozlov; Thorkell Andresson; Udo Rudloff

    Synthetic host defense peptides (HDP) are a new class of promising therapeutic agents with potential application in a variety of diseases. RP-182 is a 10mer synthetic HDP design, which selectively reduces M2-like tumor associated macrophages via engagement with the cell surface lectin receptor MRC1/CD206 and is currently being developed as an innate immune defense regulator to improve anti-tumor immunity in immunologically cold tumors. Herein, we describe a sensitive and specific liquid chromatography (LC) coupled to quadrupole electron spray tandem mass spectrometry method to measure positively charged HDPs and HDP peptide fragments in complex biological matrices. Carboxylic acid magnetic beads were used as an affinity-capturing agent to extract the positively charged RP-182 from both mouse plasma and tissue homogenates. Beads were eluted with 0.1% (v/v) formic acid and chromatographic separation was achieved on a Waters 2.1 x 100 mm, 3.5 µm XSelect Peptide CSH C18 column with a Vanguard pre-column of the same phase. MS/MS was performed on a Thermo TSQ Quantiva triple quadrupole mass spectrometer operating in Selected Reaction Monitoring (SRM) mode fragmenting the plus three parent ion 458.9+3 and monitoring ions 624.0+2, 550.5+2, and 597.3+1 for RP-182 and 462.4+3 > 629.1+2, 555.5+2, and 607.3+1 for isotopic RP-182 standard. The assay had good linearity ranging from 1 ng to 1000 ng in mouse plasma with the lower limit of detection for RP-182 at 1 ng in mouse plasma with good intra- and inter-sample precision and accuracy. Recovery ranged from 66% to 77% with minimum matrix effects. The method was successfully applied to an abbreviated pharmacokinetic study in mice after single IP injection of RP-182. The method was successfully tested on a second HDP, the 17mer D4E1, and the cationic human peptide hormone ghrelin suggesting that it might be a general sensitive method applicable to quantifying HDP peptides that are difficult to extract.

    更新日期:2020-01-02
  • Metabolomic study of raw and bran-fried Atractylodis Rhizoma on rats with spleen deficiency
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-30
    Bei-xue Zhang; Xiao-jie Qi; Qian Cai

    Atractylodis Rhizoma, a classical Chinese medicine, exhibits unambiguous therapeutic effect on spleen deficiency in China for decades. The aim of the present study was to explore the different effects on the composition and level of endogenous metabolites in rats with spleen deficiency after oral administration of raw and bran-fired Atractylodis Rhizoma, and to explain the mechanism of pharmacodynamic enhancement of the bran-fried Atractylodis Rhizoma from the perspective of metabolomics. With this purpose, spleen deficiency model was established by diet, excessive fatigue and bitter cold diarrhea. Then, Enzyme-linked immunosorbent assay (ELISA) was used to determine the contents of vasoactive intestinal peptide (VIP), Somatostatin (SS), substance P (SP) and succinodehydrogenase (SDH) in rats of each group, and to compare the contents of VIP, SS, SP and SDH among groups. UPLC-Q-TOF-MS based metabolomics was adopted to analyze the plasma from spleen deficiency rats and control rats. Principle component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) were utilized to identify differences of metabolic profiles in rats among the control group and the model group;The OPLS-DA were used to analyze the effects of raw and bran-fried Atractylodis Rhizoma on the same metabolites. The results showed that compared with the control group, the contents of VIP, SS, SP and SDH in the plasma of model group decreased, which proved the success of the model group. Compared with model group, the contents of VIP, SS, SP and SDH in the plasma of raw and bran-fried Atractylodis Rhizoma increased, and the effect of bran-fried Atractylodis Rhizoma was better than that of raw Atractylodis Rhizoma. Metabolomics results showed that seventeen different metabolites of spleen deficiency were screened out in the plasma of rats with spleen deficiency compared with the control group. Among them, Nicotinic acid, Dihydrofolic acid, Pantetheine 4'-phosphate and Photophatidylcholine (PC) were the metabolites significantly associated with spleen deficiency, and bran-fried Atractylodis Rhizoma had better intervention and regulation. Through the analysis of metabolic pathways related to these different metabolites of spleen deficiency, and primarily involved in glucosamine metabolism, one carbon pool by folate and so on. This study showed that Atractylodis Rhizoma could provide satisfactory therapeutic effects on spleen deficiency and metabolomics study can be utilized to further understand the molecular mechanisms.

    更新日期:2019-12-30
  • A quality control system for ligand-binding assay of plasma renin activity: Proof-of-concept within a pharmacodynamic study
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-28
    Fabian Konstantin Suessenbach; Nina Makowski; Martin Feickert; Tanja Gangnus; Jutta Tins; Bjoern Bengt Burckhardt

    While the role of plasma renin activity (PRA) in heart failure have been widely studied in adults, comprehensive data on pediatric heart failure remain lacking. This drawback is increasingly being addressed by academic research. Nevertheless, such pediatric investigations are commonly conducted only once due to ethical constraints. Therefore, the quality of bioanalytical data must be ensured to acquire meaningful insights into maturing humoral parameters. However, appropriate post-validation assessment of bioanalytical runs is currently underrepresented by regulatory guidance. Thus, for applications in an academic environment, an easy-to-handle six-step bioanalytical quality control system was designed based on regulatory guidelines (e.g. U.S. Food and Drug Administration) combined with international recommendations (e.g. Clinical and Laboratory Standards Institute) and current scientific discussion. Its applicability to an enzyme-linked immunosorbent assay for determination of PRA was investigated within three pediatric trials of the EU-funded “Labeling of Enalapril in Neonates up to Adolescents” project. This quality control system identified 15% bioanalytical runs as non-compliant to the predefined specifications and ensured the reliable quantification of 940 pharmacodynamic samples. The inter-run assessment of quality controls was able to demonstrate the comparability of the study results. Furthermore, 86% of incurred sample reanalysis pairs complied with regulatory requirements (>67%), thus underlining the long-term reproducibility of the utilized ligand-binding assay. Successful participation in interlaboratory testing confirmed the accuracy of the applied method throughout the entire study period. Further investigations showed no notable differences between the five applied lots of the PRA assay. The applicability of this quality control system was proven in an academic environment and ensured reliable results for PRA over the entire 24-month study period.

    更新日期:2019-12-29
  • Measurement of NLG207 (formerly CRLX101) nanoparticle-bound and released camptothecin in human plasma
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-27
    Keith T. Schmidt; Cody J. Peer; Alwin D.R. Huitema; Monique D. Williams; Susan Wroblewski; Jan H.M. Schellens; Ravi A. Madan; William D. Figg

    Camptothecin (CPT), a potent inhibitor of topoisomerase I and HIF-1α, failed to demonstrate utility as an anti-cancer agent in early clinical trial investigations, primarily due to limited clinical activity and significant toxicity attributable to unfavorable physicochemical properties (e.g. low plasma solubility, pH-labile lactone ring). NLG207 (formerly CRLX101), a nanoparticle-drug conjugate (NDC) of CPT designed to optimize plasma pharmacokinetics and facilitate drug delivery to tumors, is included as part of combination treatment in two Phase II clinical trials ongoing at the National Cancer Institute (NCT02769962 and NCT03531827). To better understand the potential for drug-drug interactions and to correlate drug exposure to clinical outcomes and pharmacodynamic biomarkers, a robust analytical method was developed to measure CPT in human plasma. Two sample processing methods were developed to quantify both NDC-bound CPT and free CPT, primarily via alteration of pH conditions. A solid-phase extraction recovered >79% of CPT prior to quantitative analysis by ultra HPLC-MS/MS. Dynamic calibration ranges of 10 to 10,000 ng/mL and 1 to 1,000 ng/mL for total and free CPT, respectively were expected to capture clinical ranges. NLG207 NDCs demonstrated significant rates of CPT release in human plasma at room temperature after 2 hours but were shown to be stable at 4 °C for 24 hours and through 4 freeze/thaw cycles. This assay was used to quantitate CPT plasma concentrations in clinical samples to confirm clinical utility following NLG207 treatment in a subject with advanced prostate cancer.

    更新日期:2019-12-27
  • Metabolomics study to identify Plasma Biomarkers in Alzheimer Disease: ApoE genotype effect
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-26
    carmen peña-bautista; marta roca; rogelio lópez-cuevas; miguel baquero; máximo vento; consuelo cháfer-pericás

    Alzheimer Disease (AD) is the main cause of dementia, and it has a great social and economic impact worldwide. It is a complex multifactorial disease, and we still do not know enough about its causes. For this reason, omics studies could be a useful tool for the search for new biomarkers and for enhancing the knowledge of different metabolic pathways that may be altered in the initial stages of the disease. Metabolomic analysis was carried out for plasma samples from early AD patients and healthy controls. Obtained data were normalized and analyzed by volcano plot and supervised orthogonal-least-squares-discriminant analysis. Fifteen variables were selected as the most important variables for the groups’ discrimination, and the different levels of 6 identified metabolites could discriminate between patients with different ApoE4 genotypes (ε4-carriers and non ε4-carriers). In conclusion, ApoE4 genotype is associated with changes in lipid metabolomics profile in AD patients, and it could be relevant for the development of AD since early stages.

    更新日期:2019-12-27
  • Rapid and highly sensitive quantification of the anti-tuberculosis agents isoniazid, ethambutol, pyrazinamide, rifampicin and rifabutin in human plasma by UPLC-MS/MS
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-26
    Lingjie Wu; Zhenjie Ye; Hui Liu; Hongliang Guo; Jing Lin; Ling Zheng; Nannan Chu; Xiaolong Liu
    更新日期:2019-12-27
  • 更新日期:2019-12-27
  • Metabolic profiling of chronic obstructive pulmonary disease model rats and the interventional effects of HuaTanJiangQi decoction using UHPLC-Q-TOF/MSE
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-24
    Wei Fang; ChenHui Li; QingQing Wu; ZhaoMin Yao; Jie Wu; Peng Huang; DianLei Wang; ZeGeng Li

    The occurrence of chronic obstructive pulmonary disease (COPD) will lead to physiological and pathological variations and endogenous metabolic disorders. A traditional Chinese medicine formula, HuaTanJiangQi decoction (HTJQ), exhibits an unambiguous therapeutic effect on COPD in China. Nevertheless, the mechanism of its therapeutic effect on COPD is not clear. With this purpose, pulmonary function, histopathological and the inflammatory factors in bronchoalveolar lavage fluid (BALF) in rats model of COPD were investigated. Then, ultra high-performance liquid chromatography quadrupole-time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) analysis and multivariate statistical analysis were used to further reveal the mechanism of HTJQ therapeutic effect on COPD via metabolomics study. The results showed that the characteristics of lung tissues were significantly reversed, the concentration of LTB4 and LTC4 were gradually decreased, and the lung function began to recover after HTJQ treatment. These typical indicators of COPD in HTJQ intervention group were reversed similar to the control group, suggested that HTJQ has a therapeutic effect on COPD. Moreover, 32 dysregulated metabolites, including Thromboxane a2, Sphingosine 1-phosphate, PC(18:2(9Z,12Z)/18:1(11Z)), Leukotriene B4, Glutathione, Arachidonic acid, Sphingosylphosphocholine acid, N-Acetyl-leukotriene e4, Lysopc(18:1(11Z)), L-Cysteine, and Guanosine diphosphate. All the altered metabolites were associated with the onset and development of COPD, and involved in glycerophospholipid metabolism, sphingolipid metabolism, glutathione metabolism, and arachidonic acid metabolism, which were significantly changed in rats model with COPD. Generally, these findings provide a systematic view of metabolic changes linked to the onset and development of COPD, also indicated that HTJQ could provide satisfactory therapeutic effects on COPD and metabolomics study can be utilized to further understand the molecular mechanisms.

    更新日期:2019-12-25
  • Recent developments of bioanalytical methods in determination of neurotransmitters in vivo
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-24
    Joanna Matys; Barbara Gieroba; Krzysztof Jóźwiak
    更新日期:2019-12-25
  • Circulatory Glutamine/Glucose ratio for evaluating disease activity in Takayasu arteritis: A NMR based serum metabolomics study
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-24
    Umesh Kumar; Avinash Jain; Anupam Guleria; Venkatesh Kumar R; Durga P Misra; Ruchika Goel; Debashish Danda; Ramnath Misra; Dinesh Kumar

    Quantitative assessment of disease activity is important for effective care of patients with Takayasu arteritis (TA). Activated glutaminolysis and reduced glycolytic flux is the hallmark of active inflammation. Based on this, we hypothesize that the circulatory Glutamine/Glucose ratio (QGR) can serve as an indicant of active inflammation in TA. To probe this hypothesis, the serum samples were collected from 45 active and 53 inactive TA patients fulfilling American College of Rheumatology (ACR) criteria and assessed for disease activity according to Indian Takayasu Clinical Activity Score (ITAS) using acute phase reactant–erythrocyte sedimentation rate [ITAS-A (ESR)]. The quantitative profiles of circulatory metabolites implicated in glutaminolysis (Glutamine and Glutamate) and those which estimate glycolytic flux (i.e. glucose and lactate) were measured using high filed (800 MHz) NMR spectroscopy. The recorded spectra were analyzed using CHENOMX NMR Suite and the estimated concentration profiles were compared and evaluated for their diagnostic potential using Metaboanalyst. Compared to inactive-TA patients, the sera of active-TA patients were characterized by significantly decreased serum levels of glutamine and lactate suggesting that these patients exhibit activated glutaminolysis and reduced glycolytic activity. This is further supported by significantly decreased QGR and lactate to glucose ratio (LGR) levels in active compared to inactive TA patients. The receiver operating characteristic (ROC) curve analysis revealed satisfactory accuracy, sensitivity and specificity for QGR [with area under ROC curve (AUROC) = 0.76 and 95% confidence interval (CI) = 0.66-0.84) compared to that for LGR (with AUROC = 0.67 and CI = 0.561-0.77). Therefore, we believe that the circulatory QGR has the potential to serve as surrogate marker for the assessment of disease activity in TA patients. However, the use of this ratio in clinical settings will require future studies on large patient cohorts and procedural optimization as well to improve accuracy.

    更新日期:2019-12-25
  • UPLC–MS/MS method for the simultaneous quantification of bictegravir and 13 others antiretroviral drugs plus cobicistat and ritonavir boosters in human plasma
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-24
    Hélène Gouget; Gaëlle Noé; Aurélie Barrail-Tran; Valérie Furlan

    A sensitive and rapid ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method has been developed and validated for 14 antiretroviral drugs and 2 boosters in human plasma. Plasma (100 µL) was precipitated with a solution of acetonitrile containing labelled internal standards. The compounds were separated with a total chromatic run time of 6 min using an Acclaim TM RSLC 120 C18 column (2.1 x 100 mm, 2.2 µm). The method was fully validated according to the European Medecines Agency guidelines. Linearity of all analytes concentrations was validated up to 5000 ng/mL. Lower limits of quantification were ranged from 2.5 ng/mL to 10 ng/mL according to compounds. Intra-day and inter-day precision ranged from 0.2% to 8.9% and accuracies were below 13%. This UPLC-MS/MS method can be applied to clinical pharmacology research and therapeutic drug monitoring in patients living with HIV.

    更新日期:2019-12-25
  • The elution behavior of cyclosporine congeners in a developed HPLC system reflects the lipophilicity
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Kumiko Sakai-Kato; Kohki Yoshida

    Recently, the development of cyclic peptide drugs has accelerated. To develop an analytical method to determine the physicochemical properties of these lipophilic drug candidates, we investigated the separation mechanism of cyclosporine congeners A, B, C, and D using HPLC with a column packed with 2-μm nonporous octadecylsilyl silica particles at high temperature. The four congeners were eluted with good repeatability in terms of retention time, peak area, and theoretical plate number. A difference of one amino acid in the eleven amino-acid sequence of the cyclosporine congeners was able to be recognized by our system within 4 min by isocratic elution, and the resolution was greater than 1.68. The calculated logP values of these congeners were well correlated with the retention factors with a correlation coefficient of 0.991. We could elucidate the separation mechanism of cyclosporine congeners on the high-temperature HPLC system. These results show that this method using HPLC on a column packed with 2-μm nonporous octadecylsilyl silica particles can be used for studying the lipophilicity of cyclosporine congeners.

    更新日期:2019-12-23
  • Simultaneous determination of di(2-ethylhexyl) phthalate and diisononylcyclohexane-1,2-dicarboxylate and their monoester metabolites in four labile blood products by liquid chromatography tandem mass spectrometry
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Amandine Descat; Marie Lecoeur; Mostafa Kouach; Laurence Goossens; Aurélie Thelliez; Pascal Odou; Bertrand Decaudin; Jean-François Goossens
    更新日期:2019-12-23
  • 更新日期:2019-12-23
  • Drug-gut microbiota metabolic interactions: The case of UniPR1331, selective antagonist of the Eph-ephrin system, in mice
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Francesca Ferlenghi; Riccardo Castelli; Laura Scalvini; Carmine Giorgio; Miriam Corrado; Massimiliano Tognolini; Marco Mor; Alessio Lodola; Federica Vacondio

    The interest on the role of gut microbiota in the biotransformation of drugs and xenobiotics has grown over the last decades and a deeper understanding of the mutual interactions is expected to help future improvements in the fields of drug development, toxicological risk assessment and precision medicine. In this paper, a microbiome drug metabolism case is presented, involving a lipophilic small molecule, N-(3β-hydroxy-Δ5-cholen-24-oyl)-L-tryptophan, UniPR1331, active as antagonist of the Eph–ephrin system and effective in vivo in a murine orthotopic model of glioblastoma multiforme (GBM). Following the administration of a single 30 mg/kg dose (p.o.) to mice, maximal plasma levels were reached 30 min after dosing and rapidly declined thereafter. To explain the observed in vivo behaviour, in vitro phase I and II metabolism assays were conducted employing mouse and human liver subcellular fractions and profiling main metabolites by means of tandem (HPLC-ESI-MS/MS) and high resolution mass spectrometry (HPLC-ESI-HR-MS). In the presence of in vitro mouse liver fractions, UniPR1331 showed a low phase I metabolic clearance, despite the identification of a 3-oxo and several hydroxylated metabolites. Conversely, after oral administration of UniPR1331 to mice, a novel isobaric metabolite was detected that (i) was subjected, as parent UniPR1331, to enterohepatic circulation (ii) had not been previously identified in vitro in mouse liver microsomes and (iii) was not observed forming after intraperitoneal (i.p.) administration of UniPR1331. An in vitro faecal fermentation assay produced the same chemical entity supporting a major role of gut microbiota in the in vivo clearance of UniPR1331.

    更新日期:2019-12-23
  • Optimized one-pot derivatization and enantioseparation of cysteine: application to the study of a dietary supplement
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Lucia Pucciarini; Giorgio Saluti; Roberta Galarini; Andrea Carotti; Antonio Macchiarulo; Serge Rudaz; Roccaldo Sardella

    Cysteine is a sulfur-containing amino acid which plays an outstanding role in many biological pathways in mammals. The analysis and quantification of native cysteine remains a critical issue due to its highly reactive thiol group evolving to the disulfide cystine derivative through oxidation reaction. Aimed at improving the derivative stability, cysteine was labelled with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), which reacts with both amino and thiol groups. The derivatization was optimized and the chemical identity of the reaction product was assessed via high-resolution mass spectrometry. The NBD-cysteine derivative resulted stable for 10 days. This derivative was enantioresolved (α and RS equal to 1.25 and 2.70, respectively) thanks to a (R,R)-Whelk-O1 phase with the following chromatographic setting: eluent, MeOH/water-90/10 (v/v) with 15 mM ammonium formate ( pwsH 6.0); column temperature, 35 °C; flow rate, 1.0 mL/min. The developed method was validated following the ICH guidelines and applied for the quality control of a L-cysteine containing dietary supplement.

    更新日期:2019-12-23
  • Untargeted Metabolomics Reveals novel serum biomarker of Renal Damage in Rheumatoid Arthritis
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Lili Song; Qingshen Yin; Mingqin Kang; Ningning Ma; Xin Li; Zhen Yang; Hua Jin; Mengya Lin; Pengwei Zhuang; Yanjun Zhang

    Rheumatoid arthritis (RA) is a chronic progressive disease, it often involves kidney, lung, heart, and other systems.Renal damage is quite common in RA. Exploring of biomarkers of renal damage in the course of RA progression is of significant importance for disease diagnosis and treatment. We use type II Collagen-Induced Arthritis(CIA) Model. Serums were collected at the 4th, 6th, 8th, and 10th week after the first immunization. An untargeted metabonomic strategy based on UPLC-Q/TOF/MS with support vector machine(SVM) was developed to discover the biomarkers in the rats’ serum samples between the RA stage(4-6 weeks in RA model, at which time the kidneys are not affected) and renal damage in RA stage(8-10 weeks in RA model, and the kidneys are affected). Principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) were used to analyze the metabolic profiles of rat serum. The support vector machine (SVM) method was used to screen the specific markers of renal damage in RA. Following multivariate statistical and integration analysis, 5 specific markers of renal damage in RA were screened and found. After the analysis of these metabolites, pentose and glucuronate interconversions are closely related to the pathogenesis of RA renal damage. The present study first use untargeted dmetabonomics combined with the pathological features in the different phases of CIA model rats. This will provide a basis for the choice of treatment drugs for patients with RA who may be complicated by renal damage.

    更新日期:2019-12-23
  • Spectrofluorimetric cytosensing of colorectal cancer cells using terbium-doped dendritic fibrous nano-silica functionalized by folic acid
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Jafar Soleymani; Mohammad Hasanzadeh; Nasrin Shadjou; Mohammad Hossein Somi; Abolghasem Jouyban
    更新日期:2019-12-23
  • 更新日期:2019-12-23
  • Using HPLC to analyze (S)-oxiracetam and four related substances in the bulk drug of (S)-oxiracetam
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Chao Wang; Hong Dong; Honghui Liu; Zhaoyi Sun; Ahu Yuan; Jinhui Wu; Yiqiao Hu

    (S)-oxiracetam is undergoing clinical trials as an active ingredient in the racemic oxiracetam. Here, we report a specific analytical method for analyzing (S)-oxiracetam and four related impurities in the bulk drug of (S)-oxiracetam by using high-performance liquid chromatography (HPLC) system. The chromatographic system included a Capcell pak NH2 analytical column, a mobile phase containing acetonitrile-water (95:5, v/v; pH adjusted to 2.0 with trifluoroacetic acid) at a flow rate of 1.0 mL/min, column temperature at 35 ℃ and the UV detection wavelength is set at 210 nm. This analytical method has shown effective and specific analysis for (S)-oxiracetam and four related substances. Moreover, the molecular weight and chemical structure preliminarily speculated of related substances were characterized by mass spectrometry. The methodology was verified by HPLC and results collected of the method validation included the system suitability, specificity sensitivity, linearity and accuracy, good linear correlation coefficient R2 was more than 0.9991. The analytical method developed and verified in the study, as far as we know, is the most exhaustive HPLC determination report which could be applied for the quality control and stability monitor purposes of the bulk drug of (S)-oxiracetam in the routine pharmaceutical analysis.

    更新日期:2019-12-23
  • LC-ESI-MS/MS Assay Development and Validation of a Novel Antidiabetic Peptide PSTi8 in Mice Plasma using SPE: an Application to Pharmacokinetics
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Guru R. Valicherla; Mohammed Riyazuddin; Sudhir Shahi; Anand P. Gupta; Anees A. Syed; Athar Husain; Jiaur R. Gayen
    更新日期:2019-12-23
  • Metabolic and lipidomic characterization of malignant pleural effusion in human lung cancer
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Zhiyi Yang; Zhengbo Song; Zhongjian Chen; Zhenyu Guo; Hangbiao Jin; Cheng Ding; Yanjun Hong; Zongwei Cai

    Malignant pleural effusion (MPE) is an important hallmark for late-stage lung cancer with metastasis. Current clinical diagnosis methods require tedious work to distinguish MPE from benign pleural effusion (BPE). The objective of this study was to characterize the metabolic signatures in MPE of lung cancer, and identify potential metabolite biomarkers for diagnosis of MPE. MPE from lung cancer (n = 46) and BPE from tuberculosis patients (n = 32) were investigated by liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based global metabolomic and lipidomic profiling. Multivariate partial least-square discriminative analysis models exhibited distinct metabolic profiles between MPE and BPE. A total of 25 ether lipids, including phosphatidylcholines (PC), lysophosphatidylcholines (LPC) and phosphatidylethanolamines (PE), were observed to be significantly downregulated in MPE with excellent diagnostic potential. Plasmalogen PC(40:3p) showed highest AUC value of 0.953 in receiver operating characteristic (ROC) model. Oxidized polyunsaturated fatty acids (PUFA) were upregulated in MPE. The obtained results implied a high oxidative stress and peroxisome disorder in lung cancer patients. Combined metabolomic and lipidomic profiling have discovered potential biomarkers in MPE with excellent clinical diagnostic capability. Dysregulated ether lipids and oxidized PUFAs have implied an aberrant redox metabolism, which provides novel insights into the pathology of MPE in lung cancer.

    更新日期:2019-12-23
  • Characterization of chemical profile and quantification of representative components of DanLou tablet, a traditional Chinese medicine prescription, by UHPLC-Q/TOF-MS combined with UHPLC-TQ-MS
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-23
    Pei Lin; Qi Wang; Yuehe Liu; Zifei Qin; Hao Gao; Min Ye; Hongcai Shang; Xinsheng Yao; Zhihong Yao

    DanLou tablet (DLT), a famous traditional Chinese medicine prescription (TCMP) consisting of 10 herbal medicines, is extensively used for the treatment of angina pectoris and acute coronary syndrome in China. However, active chemical constituents responsible for the therapeutic effects still remains unclear, due to the fact that the complex composition in DLT have not been holistically clarified. Therefore, this study aimed to characterize the chemical profile and simultaneously quantify the representative components in DLT. First, 157 chemical constituents including flavonoids, triterpenoids, tanshinones, lactones, phenolic acids, paeoniflorins and the other types of components were detected, among which 39 were exactly identified by comparing their retention times and MS fragmentation behaviors with those of authentic standards by ultra-high performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UHPLC-Q/TOF-MS). Moreover, 33 representative components were simultaneously quantified by ultra-high performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UHPLC-TQ-MS), which were selected based on following three principles: qualitative and quantitative markers in the Chinese Pharmacopeia (2015 edition), bioactive components possessing cardiovascular-related in vivo or in vitro activities and those derived from 10 consisted herbs in DLT with a diversity of representative structure types. The method was validated in terms of linearity, precision, repeatability and recovery and successfully applied for the quality evaluation of 20 batches of DLT samples. Further chemometric analysis indicated that danshensu and salvianolic acid B were the most significant quantitative markers for the content fluctuation of DLT. In summary, the chemical profiles of DLT were systematically characterized and a practical quantitative method combined with chemometrics was developed to evaluate the intrinsic quality of multiple DLT samples in this study. The present work would be helpful for guaranteeing the safety, efficacy, and controllability in clinical medication of DLT.

    更新日期:2019-12-23
  • The relationship between serum clozapine concentrations and hematological parameters by a validated mass spectrometric method
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-20
    Karam Mazin Kamil Gharab; Duygu Eryavuz Onmaz; Sedat Abusoglu; Memduha Aydin; Abdullah Sivrikaya; Oguzhan Tok; Gulsum Abusoglu; Ali Unlu

    Objective Clozapine is one of the most effective drugs for resistant schizophrenia, but its severe metabolic and hematological side effects limit the use of clozapine. It has been reported that clozapine blood concentrations should be maintained between 350-600 ng/mL. Our aim was to develop a determination method for clozapine and its main metabolites norclozapine and clozapine-N-oxide, to perform validation studies and to investigate the change of various biochemical parameters in patients using clozapine. Methods A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for clozapine measurement. Thus, blood samples were collected from 38 patients with schizophrenia and 32 healthy volunteers. Biochemical and hematological parameters were measured by Beckman-Coulter AU 5800 (Beckman Coulter, Brea, USA) and Beckman Coulter LH 780 analyzer (Beckman Coulter, Miami, FL, USA), respectively. Hormone levels were analyzed using Cobas 6000 analyzer (Roche Diagnostics, Germany). Results The LC-MS/MS method was linear between 1.22-2500 ng/mL (r2 = 0.9971) for clozapine. The retention times of clozapine, norclozapine and clozapine-N-oxide were 0.92, 0.89 and 0.95, respectively. Blood glucose (GLU) (p = 0.025), low density lipoprotein (LDL-cholesterol) (p = 0.015), triglyseride (TG) (p = 0.042) and total cholesterol (TC) (p = 0.024) levels were higher; hemoglobin (HGB) (0.015), mean corpuscular hemoglobin (MCH) (0.036), red blood cell count (RBC) (0.020), neutrophil (NEU) (0.034), and platelet (PLT) (P = 0.005) levels were lower in the clozapine group. Conclusions This LC-MS/MS method was rapid, simple, cost-effective and suitable for the routine clozapine monitoring. Furthermore, norclozapine and clozapine-N-oxide were also determined. Monitoring of metabolic and hematological parameters with clozapine levels is very important. However, the limitations of the study were that the method was not validated for norclozapine and clozapine-N-oxide, so the validation parameters were not evaluated for these two metabolites.

    更新日期:2019-12-20
  • Relationship between cancer tissue derived and exhaled volatile organic compound from colorectal cancer patients. Preliminary results
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-20
    Nicoletta De Vietro; Antonella Aresta; Maria Teresa Rotelli; Carlo Zambonin; Catia Lippolis; Arcangelo Picciariello; Donato Francesco Altomare
    更新日期:2019-12-20
  • Method transfer of a near-infrared spectroscopic method for blend uniformity in a poorly flowing and hygroscopic blend
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-20
    Bárbara B. Alvarado-Hernández; James V. Scicolone; Carlos Ortega-Zuniga; Andrés D. Román-Ospino; Yleana M. Colón; Efrain Aymat; Eric Sánchez; Fernando J. Muzzio; Rodolfo J. Romañach
    更新日期:2019-12-20
  • The role of solid state properties on the dissolution performance of flufenamic acid
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-20
    Francesca Maestrelli; Patrizia Rossi; Paola Paoli; Enrico De Luca; Paola Mura
    更新日期:2019-12-20
  • Boron and Nitrogen Codoped Carbon Dots as Fluorescence Sensor for Fe3+ with Improved Selectivity
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-20
    Hao Wu; Lan-Fang Pang; Meng-Jie Fu; Xiao-Feng Guo; Hong Wang
    更新日期:2019-12-20
  • Near infrared analysis of pharmaceutical powders with empirical target distribution optimization (ETDO)
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-20
    Troels Pedersen; Jukka Rantanen; Kaisa Naelapää; Erik Skibsted

    Near infrared (NIR) spectroscopy is a well-established method for analysis of pharmaceutical products, and especially useful for process monitoring and control of continuous production due to high sample throughput. In this work, a previously established method called empirical target distribution optimization (ETDO) wherein reference sample values using information from model prediction of the calibration data was used as a tool to improve the performance of NIR partial least squares (PLS) models. Model performance was assessed using root mean square error (R2), bias and accuracy in prediction of test samples. A target value selection threshold was tested to assess the ETDO procedure for NIR analysis of powder samples. The amount of specific variation captured by the model was examined and compared for models calibrated with and without ETDO. The results reported in this work suggests that PLS models optimized with ETDO of reference values can provide more specific PLS models for NIR analysis for complex powder mixtures. In addition, the model optimization method could also be applied as a tool to verify the necessary amount of PLS components to produce robust models. The ETDO method presented in this work is an approach that could be applied in the development of continuous blending or tableting processes where robust in-line quantitative analysis of powder samples is needed.

    更新日期:2019-12-20
  • Identification of quality control markers in Suhuang antitussive capsule based on HPLC-PDA fingerprint and anti-inflammatory screening
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-19
    Xingdong Wu; Qinyan Liu; Dong Chen; Weiwei Qin; Bingyun Lu; Qirui Bi; Zhen Wang; Yuning Jia; Ninghua Tan
    更新日期:2019-12-19
  • Attenuated total reflection Fourier transform infrared spectroscopy based methods for identification of chromatography media formulations used in downstream processes
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-19
    Éva Szabó; László Zoltán Baranyai; Zoltán Sütό; András Salgó; Szilveszter Gergely

    Chromatographic media play a crucial role in the downstream processing of biotechnology products. The physical and chemical properties of these processing aids are mostly monitored by expensive and time‐consuming preparative tests, but spectroscopic techniques may also be used to measure chromatographic media samples. In this study, chromatographic media formulations used in downstream processes were investigated using attenuated total reflection Fourier transform infrared (ATR FT-IR) spectroscopy. Samples were measured both in original suspension form and after drying to examine the possibilities of a potential spectroscopic method without sample preparation. Principal component analysis (PCA) was employed to identify the spectral differences among the formulations with distinct support matrices and functional groups and soft independent modeling of class analogy (SIMCA) was performed to creating classification models for identification of chromatography media. To increase the number of samples in the SIMCA, simulated spectra were generated based on the experimental spectra. PCA models indicated that spectra of samples in original suspension form and after drying contained similar information about the chemical properties of chromatographic media samples. Moreover, during the classification of spectra based on SIMCA, both measurement techniques gave high sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) results. These results show that ATR FT-IR could be applied as a simple alternative method for monitoring the chromatography media samples. This technique is also feasible without sample preparation. Thereby the multi-hours drying steps may be omitted, the measurements can be performed in a few minutes, and the potential effects of sample preparations can be eliminated.

    更新日期:2019-12-19
  • An enantiomeric quantitative LC-MS/MS method for tolvaptan and its monohydroxylates in human plasma using a reversed-phase separation procedure
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-19
    Shunta Akutsu; Takafumi Naito; Kohei Oshawa; Masao Saotome; Yuichiro Maekawa; Junichi Kawakami
    更新日期:2019-12-19
  • 更新日期:2019-12-19
  • Lifecycle management in pharmaceutical analysis: How to establish an efficient and relevant continued performance monitoring program
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-19
    Joachim Ermer; Debora Aguiar; Annette Boden; Bei Ding; Daniel Obeng; Michael Rose; Julie Vokrot

    It is the objective of a systematic and holistic Quality-by-Design approach to demonstrate and ensure that an analytical procedure is fit for its intended purpose over its entire lifecycle. Such a lifecycle approach, as proposed for a new USP General Information Chapter includes the three stages Procedure Design and Development, Procedure Performance Qualification, and Continued Procedure Performance Verification, in alignment to manufacturing process validation. A decisive component of this approach is the Analytical Target Profile, which defines the performance requirements for the measurement of a Quality Attribute as the target for selection, development and optimization of the respective analytical procedures. Although the most benefit can be gained by a comprehensive Quality-by-Design approach establishing the Analytical Target Profile in the very beginning of a drug development project, it may also be established retrospectively for analytical procedures long in routine use, in order to facilitate future lifecycle activities such as continual improvements, transfers, monitoring and periodic performance evaluations. In contrast to the first two stages of the analytical lifecycle with usually limited amount of data, the Continued Procedure Performance Verification stage offers the possibility to utilize a much more reliable data base to collect, analyze, and evaluate data that relate to analytical procedure performance. This monitoring program should be aligned as far as possible with other quality systems already in place and may include performance indicators such as Conformity (i.e. out-of specification test results with analytical root-cause), Validity (i.e. failure to meet method acceptance criteria, e.g. system suitability tests), and (numerical) analytical performance parameters (e.g. ranges for replicate determinations, control sample results, etc). In addition to the monitoring of analytical control parameters by means of control charts, average (pooled) performance parameters can be calculated. Over time, a large number of data can be included and thus the reliability of these estimates is increased tremendously. Such reliable estimates for the true performance parameters, e.g. repeatability or intermediate precision are essential to identify systematic effects (also called special cause variation) with good confidence. The intent of the analytical procedure performance evaluation is to identify substandard performance, identify root cause through investigations, and determine when additional activities are required to improve it. Examples are provided for the monitoring and evaluation of performance parameters for the compendial API Furosemide and for biopharmaceutical applications.

    更新日期:2019-12-19
  • Ketamine analogues: Comparative toxicokinetic in vitro–in vivo extrapolation and quantification of 2-fluorodeschloroketamine in forensic blood and hair samples
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-18
    Anders Bork Davidsen; Marie Mardal; Niels Bjerre Holm; Anna Katrine Andreasen; Sys Stybe Johansen; Carolina Noble; Petur Weihe Dalsgaard; Kristian Linnet

    Recently, the new psychoactive substance (NPS) ketamine analogue 2-fluoro-deschloroketamine (2FDCK) was observed in driving-under-the-influence-of-drugs whole blood samples and in a forensic hair investigation case in Denmark. The molecular structure variations among the NPS subgroups may alter the metabolic fate and drug potency, thereby posing a threat for drug users. This study reports quantification of 2FDCK in whole blood samples and forensic hair and compares the following toxicokinetic parameters: unbound fraction (fu) and in vitro–in vivo-extrapolation (IVIVE) of hepatic clearance for ketamine, norketamine, 2FDCK, methoxetamine and deschloroketamine. The fu was investigated with ultrafiltration, while clearance studies were conducted at 1 µM with pooled human liver microsomes. Samples were analysed by liquid chromatography and mass spectrometry. For the first time, 2FDCK was determined in a concentration range between 0.005 and 0.48 mg/kg in blood samples. Segmental hair analysis demonstrated 2FDCK at concentrations from 0.007–0.034 ng/mg throughout the three investigated segments. Toxicokinetic comparison of the five compounds gave a fu between 0.54 and 0.84, with ketamine being the most bound and deschloroketamine being the least bound, in accordance with the logP of the compounds. Conversely, a negative correlation was observed between the molecular weight of the halogen in the ortho-position and IVIVE hepatic clearance. The IVIVE of hepatic clearance, CLparallel-tube, gave values from 18.1 to 5.44 mL/min/kg for ketamine and methoxetamine, respectively. The deschloroketamine IVIVE was disregarded due to low drug elimination under the experimental conditions used. This study provides a basis for toxicokinetic understanding of ketamine analogues.

    更新日期:2019-12-19
  • Toxicological Effects of Some Antiparasitic Drugs on From Equine Liver Glutathione S-Transferase Enzyme Activity
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-17
    Fikret Turkan; Mehmet Harbi Calimli; Ayhan Akgun; Fulya Gulbagca; Fatih Sen

    Benzimidazoles are antiparasitic drugs having an extensive application field like agriculture, medicine, and especially in veterinary medicine. In this study, we report the effect of some benzimidazole drugs such as ricobendazole (RBZ), thiabendazole (TBZ), albendazole (ALBA) and oxfendazole (OFZ) on glutathione s-transferase (GST) enzyme activity. The kinetics studies, IC50 and Ki values of the tested drugs on GSTs enzyme activity were investigated. The obtained ranking of IC50 values were found to be approximately RBZ (53.31 µM, r2: 0.9778) < OFZ (57.75 µM, r2: 0.9630) < ALBA (63.00 µM, r2: 0.9443) < TBZ (69.30 µM, r2: 0.9491). And the obtained ranking of Ki values of the tested drugs (RBZ, TBZ, ALBA, and OFZ) for GSTs enzyme activity was found to be approximately 26.37 ± 2.96, 44.01 ± 5.74, 39.82 ± 3.98 and 30.14 ± 3.03 µM, respectively. Experimental results showed that tested the benzimidazoles drugs have some significant inhibitory effect on GSTs enzyme activity. And also, it was determined that RBZ, ALBA, OFZ are competitive inhibition, but TBZ is non-competitive inhibitors on GSTs enzyme activity. RBZ drug showed the best inhibitory effect with the lowest Ki value.

    更新日期:2019-12-18
  • Development of an on-line immobilized α-glucosidase microreactor coupled to liquid chromatography for screening of α-glucosidase inhibitors
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-17
    Beibei Qiu; Yuqiong Shi; Luyun Yan; Xiangrong Wu; Jinhua Zhu; Dongbao Zhao; Zaved H. Khan; Xiuhua Liu
    更新日期:2019-12-18
  • A novel UPLC/MS/MS method for rapid determination of murrayone in rat plasma and its pharmacokinetics
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-16
    Shengyi Zhai; Xiaoxiao Deng; Chen Zhang; Yu Zhou; Huanzhang Xie; Zhou Jiang; Lee Jia

    Murraya paniculata (L.) is a traditional Chinese medicine (TCM) wildly grown in southeast China, and used for abortion in folk. Murrayone, a coumarin-containing compound extracted from M. paniculata, is the most bioactive substance in this species and is being developed as a novel cancer metastasis chemopreventive agent based on its unique pharmacological properties. In the present study, a novel rapid and sensitive method for quantitative analysis of murrayone in rat plasma and for determining its pharmacokinetics in rats was developed and validated using UPLC/MS/MS. Plasma samples were subjected to protein precipitation and then directly analyzed by UPLC/MS/MS. Both murrayone and coumarin as an internal standard (I.S.) were carried on a C18 column with a gradient mobile phase consisting of acetonitrile and water at a flow rate of 0.3 mL/min. Several gradient elution procedures were evaluated to achieve effective chromatography resolution and a sensitive response to murrayone and the I.S.. Mass spectrometry was carried out using a triple-quadrupole system via positive electrospray ionization and multiple reaction monitoring (MRM). Good linearity (r 2 = 0.9987) was achieved over a linear range of 4.0 - 1600 ng/mL with a lower limit of quantitation (LLOQ) of 4.0 ng/mL for murrayone. The inter- and intraday accuracy and precision ranged from 90.0 to 99.7% and 1.1 to 12.3% at four quality control concentrations, respectively. The average absolute recoveries of murrayone and the I.S. were determined to be 85.9 - 92.4% and 86.5 - 90.7%, respectively, at 10.0, 80.0, and 800 ng/mL. Murrayone was stable under a variety of storage and processing conditions that may be routinely encountered in laboratories based on all the stability tests. This newly developed method was successfully applied to the pharmacokinetic study of murrayone in rats for the first time, and the current assay methodology could provide important insights into potential therapeutics and facilitate further pharmacodynamic explorations of murrayone.

    更新日期:2019-12-17
  • Metabolic profiling of tyrosine kinase inhibitor nintedanib using metabolomics
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-16
    Zi-Meng Zhou; Yi-Kun Wang; Dong-Mei Yan; Jian-He Fang; Xue-Rong Xiao; Ting Zhang; Yan Cheng; Kang-Ping Xu; Fei Li

    Nintedanib is a promising tyrosine kinase inhibitor for clinically treating idiopathic pulmonary fibrosis (IPF). Some clinical cases reported that nintedanib treatment can cause hepatotoxicity and myocardial toxicity. U. S. FDA warns the potential drug-drug interaction when it is co-administrated with other drugs. In order to understand the potential toxicity of nintedanib and avoid drug-drug interaction, the metabolism of nintedanib was systematically investigated in human liver microsomes and mice using metabolomics approach, and the toxicity of metabolites was predicted by ADMET lab. Nineteen metabolites were detected in vivo and in vitro metabolism, and 8 of them were undescribed. Calculated partition coefficients (Clog P) were used to distinguish the isomers of nintedanib metabolites in this study. The major metabolic pathways of nintedanib majorly included hydroxylation, demethylation, glucuronidation, and acetylation reactions. The ADMET prediction indicated that nintedanib was a substrate of the cytochrome P450 3A4 (CYP3A4) and P-glycoprotein (P-gp). And nintedanib and most of its metabolites might possess potential hepatotoxicity and cardiotoxicity. This study provided a global view of nintedanib metabolism, which could be used to understand the mechanism of adverse effects related to nintedanib and its potential drug-drug interaction.

    更新日期:2019-12-17
  • Determination of buprenorphine, naloxone and phase I and phase II metabolites in rat whole blood by LC-MS/MS
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-16
    Camille Cohier; Sophie Salle; Anne Fontova; Bruno Mégarbane; Olivier Roussel

    Buprenorphine and buprenorphine/naloxone combination are maintenance treatments used worldwide. However, since their marketing, despite ceiling respiratory effects, poisonings and fatalities have been attributed to buprenorphine misuse and overdose. Therefore, to better understand the mechanisms of buprenorphine-related toxicity in vivo, experimental investigations have been conducted, mainly in the rat. We developed a liquid chromatographic-tandem mass spectrometric (LC-MS/MS) method with electrospray ionization for the simultaneous quantification of buprenorphine, naloxone and their metabolites (norbuprenorphine, buprenorphine glucuronide, norbuprenorphine glucuronide and naloxone glucuronide) in rat whole blood. Compounds were extracted from whole blood by protein precipitation and chromatographically separated using gradient elution of aqueous ammonium formate and methanol in a Raptor Biphenyl core-shell column (100 mm x 3,0 mm x 2,7 µm). Following electrospray ionization, quantification was carried out in the multiple reaction monitoring (MRM) mode by the tandem mass spectrometer API 3200 system. The LC-MS/MS method was validated according to the currently accepted criteria for bioanalytical method validation. The method required small sample volumes (50 µL) and was sensitive with limits of quantification of 6.9, 6.2, 3.6, 3.3, 1.3 and 57.7 ng/mL for buprenorphine, norbuprenorphine, buprenorphine glucuronide, norbuprenorphine glucuronide, naloxone and naloxone glucuronide respectively. The upper limit of quantification was 4,000 ng/ml for all the studied compounds. Trueness (88-115%), repeatability and intermediate precision (both <15%) were in accordance with the international recommendations. The procedure was successfully used to quantify these compounds in the whole blood sample from one rat 24 hours after the intravenous administration of buprenorphine/naloxone (30.0/7.5 mg/kg).

    更新日期:2019-12-17
  • Development and validation of an LC-MS/MS method for the quantification of goserelin in a Pheroid® formulation simulated intestinal fluid
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-12
    John Takyi-Williams; Linnè Erasmus; Rose Hayeshi; Anne Grobler

    The purpose of this reported study was to develop and validate an LC-MS/MS method for the quantification of goserelin in a Pheroid® formulation simulated intestinal fluid. Biopharmaceuticals are formulated in drug delivery systems to improve their gastrointestinal stability. Goserelin, a peptide drug was formulated in Pheroid® delivery system and its gastrointestinal stability assessed using simulated intestinal fluid, which required an assay to determine the varying amounts of goserelin remaining after a specific time. Several extraction methods and solvents investigated to extract goserelin from complex matrix led to either poor recovery, peak shape or high background interference. A rapid gradient reversed-phase method coupled to tandem mass spectrometry detection was optimized for the separation and quantification of the extracted peptide. A simple, reproducible and good recovery extraction procedure for goserelin quantification was achieved through simultaneous acetonitrile protein precipitation and water-saturated n-butanol liquid-liquid extraction with water dilution. The method was found to be rapid, specific, precise and accurate, and successfully applied to determine goserelin remaining content in a simulated intestinal fluid, with potential use in other lipid-based formulation evaluated in simulated intestinal fluids.

    更新日期:2019-12-13
  • Sensitive Quantitation of Low Level Free Polysaccharide in Conjugate Vaccines by Size Exclusion Chromatography-Reverse Phase Liquid Chromatography with UV Detection
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-11
    Deanna C. Schuchmann, Weiying Hou, Joshua Creahan, Yan He, Michael T. Jones

    The level of free polysaccharide is a critical quality attribute of polysaccharide-protein conjugate vaccines. The work presented describes a simple and sensitive method for the determination of low level free polysaccharides in multiple polysaccharide-protein conjugates. The method utilizes a reverse phase (RP) column to perform a size exclusion chromatography (SEC) separation of free polysaccharide and a reverse phase liquid chromatography (RPLC) separation of free protein and protein-polysaccharide conjugate. The use of phosphate buffer in the mobile phase enables the universal and sensitive detection of low level free polysaccharides at UV 200 nm. The method has been validated to monitor low level free polysaccharide (< 1%) in multiple polysaccharide-protein conjugates. The limit of quantitation is 2 µg/ml or 0.3 % free polysaccharide in 0.6 mg/ml polysaccharide-protein conjugate. The accuracy is in the range of 94.1.0 – 108.5 %.

    更新日期:2019-12-11
  • Development of a Mass Spectrometry-Based Pseudotargeted Metabolomics strategy to analyze Hormone-Stimulated Gastric Cancer Cells
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-10
    Juan Li, Qingli Wang, Yichao Zheng, Piao Zhou, Xia Xu, Xueqi Liu, Longfei Zhao, Hongmin Liu

    Gastric cancer (GC) is the third most common cause of cancer death worldwide, and the incidence of GC is higher in males than females. To investigate the gastric cellular response to hormone therapy, we developed a cell pseudotargeted metabolomics method based on liquid chromatography-hybrid triple quadrupole linear ion trap mass spectrometry (LC-QTRAP MS). Chromatographic separation, sample analysis and metabolite extraction were optimized in an integrated manner. The established pseudotargeted method, which combined nontargeted and targeted analyses, exhibited high selectivity, good repeatability and wide metabolome coverage. The method was then applied to discover differential metabolites from hormone-stimulated gastric cancer cells compared with the controls for the first time. The results demonstrated that hormone had subtle but phenotypically important alterations in nucleotide metabolism, amino acid metabolism, glycolysis, tricarboxylic acid (TCA) cycle, aminoacyl-tRNA biosynthesis and so on, which indicate that the developed method is a powerful tool for effective screening of endogenous polar metabolites in cell samples.

    更新日期:2019-12-11
  • Development and validation of an LC-MS/MS method for the quantitative analysis of the anti-influenza agent camphecene in rat plasma and its application to study the blood-to-plasma distribution of the agent
    J. Pharmaceut. Biomed. Anal. (IF 2.983) Pub Date : 2019-12-10
    Alina A. Okhina, Artem D. Rogachev, Olga I. Yarovaya, Mikhail V. Khvostov, Tatyana G. Tolstikova, Andrey G. Pokrovsky, Veniamin A. Khazanov, Nariman F. Salakhutdinov

    A method of quantitative determination of camphecene, a new anti-influenza agent, in rat blood plasma based on LC-MS/MS was developed, validated and used to study the distribution of the agent between blood cells and blood plasma. The method was validated according to FDA and EMA recommendations in terms of selectivity, linearity, accuracy, precision, recovery, stability and carry-over. Plasma samples were precipitated with methanol followed by the addition of a methanolic solution of 2-adamantylamine hydrochloride (internal standard). HPLC analysis was performed on a reversed-phase column; the total time of analysis was 11 min, including column equilibration. MS/MS detection was performed on a 3200QTRAP triple quadrupole mass spectrometer in multiple reaction monitoring (MRM) mode. Transitions 196.4 → 122.2/153.3 and 152.2 → 93.1/107.2 were monitored for camphecene and the internal standard, respectively. The calibration curve was built in the concentration range of 10-5000 ng/ml. The intra-day and inter-day accuracy and precision, carry-over and recovery were within the acceptable limits. It was found that, after spiking blood with camphecene and separating plasma, the concentration of the substance in the latter was close to its initial concentration in the blood. This property of the substance may be useful for clinical trials of the agent. It has also been established that the process of camphecene distribution (adsorption) between blood cells and blood plasma is reversible, and the amount of adsorbed substance is linearly dependent on its initial concentration in the blood for a wide range of concentrations, temperatures and hematocrit values.

    更新日期:2019-12-11
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