Lihua Zuo

Rapid determination of 30 bioactive constituents in XueBiJing injection using ultra high performance liquid chromatography-high resolution hybrid quadrupole-orbitrap mass spectrometry coupled with principal component analysis

&NA; Xuebijing injection (XBJ) is a traditional Chinese herbal prescription widely used in the treatment of sepsis. Extensive chemical studies revealed that XBJ injection contains amino acids, phenolic acids, flavonoid glycosides, terpeneglycosides and phthalides. In this study, the applicability of ultra high performance liquid chromatography coupled with high resolution hybrid quadruple‐orbitrap mass spectrometry (UHPLC‐Q‐Orbitrap MS) for the simultaneous quantitative analysis of 30 bioactive constituents in XueBiJing injection (XBJ) was investigated. The mass spectrometer was operated in full MS scan mode. The use of 70,000 FWHM mass resolution and narrow mass windows (5 ppm) could effectively improve the selectivity of the method. Separation was achieved on a Waters ACQUITY UPLC® HSS C18 column (2.1 mm × 100 mm, 1.8 &mgr;m) with a gradient mobile phase consisting of acetonitrile‐water (containing 10 mM ammonium acetate) at a flow rate of 0.2 mL/min. Satisfactory linearity was achieved within wide linear range and all correlation coefficients (r) of analytes were more than 0.9996. The limits of detection (LODs) were in the range of 0.1180–27.82 ng/mL for different analytes. The relative standard deviations (RSDs) of inter‐ and intra‐day precisions were less than 3.0% and the recoveries of the assay were in the range of 98.5%–101.5%. The validated method was successfully applied for simultaneous determination of 30 bioactive compounds in XueBiJing injection from 10 batches samples by UHPLC‐Q‐Orbitrap MS within 10 min. Moreover, the results were evaluated principal component analysis and two compounds might be the most important chemical markers for chemical quality control of XBJ injection. The novel Q‐Orbitrap mass spectrometry has been proved to be a very promising and powerful tool for routine screening of bioactive compounds in traditional Chinese medicine injection, ensuring drug safety and public health. Graphical abstract Figure. No caption available. HighlightsThe report describes the validation with high precision and accuracy of an UHPLC‐Q‐orbitrap MS method for the determination of 30 bioactive constituents in XueBiJing injection for the first time.The high resolution hybrid quadrupole‐orbitrap mass spectrometry possess high accuracy and stability, and so the full MS scan mode with positive/negative ion swing was performed.This method was simple and time saving.The determination results were analyzed by multivariate analysis in order to discover the markers for quality control of XBJ injection.

Regulation of aerobic glycolysis by long non-coding RNAs in cancer.

As with miRNAs a decade ago, long non-coding RNAs (lncRNAs) are emerging as a new class of RNAs involved in physiological and pathological processes. Recent evidences have shown that lncRNAs play a role in cancer metabolism. The relationship between lncRNAs and aerobic glycolysis provides new strategies for the treatment of cancer. Here we discuss recent findings on the role of lncRNAs in aerobic glycolysis and provide insights into their mechanisms of action. In addition, we explore the potential challenges in using lncRNAs as targets for cancer therapy.

Development and validation of a sensitive UHPLC-MS/MS method for quantitation of prucalopride in rat plasma and its application to pharmacokinetics study.

A rapid, sensitive, selective and accurate ultra high performance liquid chromatography with tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for the quantitation of prucalopride in rat plasma using carbamazepine as an internal standard (IS). Separation was achieved on a Waters ACQUITY UHPLC(®) HSS C18 column (2.1mm×50mm, 1.8μm) column with a gradient mobile phase consisting of acetonitrile-water (containing 0.1% formic acid) as mobile phase at a flow rate of 0.2mL/min. Prucalopride and IS were monitored using positive electrospray triple quadrupole mass spectrometer (Waters Xevo TQD) via multiple reaction monitoring (MRM) mode. The monitored transitions were set at m/z 367.99→195.89 and m/z 236.97→194.04 for prucalopride and IS, respectively. The achieved lower limit of quantitation was 0.1ng/mL. The validated method had an excellent linearity in the range of 0.1-100ng/mL (r>0.996). The intra- and inter-day precisions were both≤7.8% for prucalopride and IS, and the average intra- and inter-day accuracies ranged from -3.0% to 8.5%. Extraction recoveries at three levels QC concentrations were in the range of 90.0-110.0% for prucalopride and 99.6% for IS. Matrix effects were found to be acceptable. The validated assay was successfully applied to a pharmacokinetic study of prucalopride following oral administration of 0.25, 0.5, 1.0mg/kg to female and male rats respectively.

Chemical profiling and quantification of ShenKang injection, a systematic quality control strategy using ultra high performance liquid chromatography with Q Exactive hybrid quadrupole orbitrap high-resolution accurate mass spectrometry

ShenKang injection is traditional Chinese medicine used to treat chronic renal failure in China. It is a compound preparation that consists of four herbs: Rhubarb, Salvia miltiorrhiza, Safflower and Radix Astragali. We developed an ultra high pressure liquid chromatography coupled with Q Exactive hybrid quadrupole-orbitrap high resolution accurate mass spectrometry tandem mass spectrometry method to analyze its chemical compositions, and a total of 90 compounds were identified from ShenKang injection. Among them, 19 major compounds were unambiguously detected by comparing with reference standards. Meanwhile, 13 representative compounds selected as quality control markers were simultaneously quantified in ShenKang injection samples. Chromatographic separation was accomplished on a Waters ACQUITY HPLC® HSS C18 column using gradient elution. The method was validated with respect to linearity, sensitivity, accuracy and precision, reproducibility and stability. And the validated method was successfully applied for simultaneous determination of 13 bioactive compounds in ShenKang injection from ten batches of samples by liquid chromatography with mass spectrometry. The results were analyzed by principal components analysis method, and three compounds had a significant relationship with the quality control of ShenKang injection. This research established a rapid and reliable method for the integrating quality control, including qualitation and quantification of ShenKang injection.

Simultaneous determination and pharmacokinetic study of twelve bioactive compounds in rat plasma after intravenous administration of Xuebijing injection by UHPLC-Q-Orbitrap HRMS

HIGHLIGHTSDeveloping and verifying of a high precision, accuracy and rapid UHPLC‐Q‐Orbitrap HRMS method for the determination of twelve bioactive compounds in rat plasma.The pharmacokinetic study of Xuebijing injection following intravenous administration of different dosages to rats is reported for the first time.The analysis time of twelve compounds and IS was less than 4 min, which offered shorter analysis time over existing methods.Extraction performed with a simple and effective protein precipitation preparation procedure for Xuebijing injection plasma samples. ABSTRACT Xuebijing injection (XBJ) is a traditional Chinese herbal prescription widely used in the treatment of sepsis. Extensive studies revealed that the major bioactive constituents of XBJ injection, including phenolic acids, flavonoids, monoterpene glycosides, lactones and organic acids, play an important role in the treatment. In this study, a rapid, sensitive and accurate ultra high performance liquid chromatography – Q Exactive hybrid quadrupole – orbitrap high‐resolution accurate mass spectrometry (UHPLC‐Q‐Orbitrap HRMS) method was developed for simultaneous determination of twelve bioactive compounds in rat plasma after intravenous administration of XBJ injection. A gradient elution for separation was achieved on a waters ACQUITY UHPLC® BEH C18 column (2.1 mm × 50 mm, 1.7 &mgr;m) column with acetonitrile‐water (containing 0.1% formic acid) as mobile phase at a flow rate of 0.2 mL/min. All compounds and IS were monitored by parallel reaction monitoring assay both in positive and negative ion mode. The developed method was validated for intra‐day and inter‐day accuracy and precision whose values fell in the acceptable limits. Extraction recoveries at three levels of QC concentrations were all more than 80% for all compounds and IS, and matrix effects were found in the range of 80.0–120.0%. Stability results showed that all analytes were stable at the investigated conditions. The validated method was successfully applied to a pharmacokinetic study of Xuebijing injection following intravenous administration of 2.5, 5.0, 10.0 mL/kg to rats respectively. And the result indicated that the pharmacokinetic behavior of XBJ injection is positively related to dosage at the range of 2.5–10 mg/kg. This study will provide a meaningful basis for evaluating the rationality of XBJ injection for clinical application.

Nonlinear relationship between enteric-coated mycophenolate sodium dose and mycophenolic acid exposure in Han kidney transplantation recipients

The aim of the research was to investigate the pharmacokinetics (PK) of enteric-coated mycophenolate sodium (EC-MPS) by quantification of the active metabolite of mycophenolic acid (MPA) after multiple escalating oral doses in Han kidney transplant recipients. A total of 28 Han postoperative kidney transplant recipients were given a multiple-dose of 540, 720 or 900 mg of EC-MPS two times a day in combination with tacrolimus for 6 days. Blood specimens were collected at each time point from 0 to 12 h after EC-MPS administration. MPA plasma concentrations were measured by UPLC—UV. The relationship between the EC-MPS dose and its PK parameters was assessed. In the range from 540 to 900 mg, Cmax and AUC0—12h did not increase with dose escalation. The AUC0—12h, Cmax, C0 and Tmax for the 540 720 and 900 mg doses were not significantly different, respectively (P >0.05). AUC0—12 h and Cmax were increased less than proportionally with increasing EC-MPS dose levels. Inter-individual variability in AUC0—12h, Cmax and C0 were considerable. Nonlinear PK relationships were found from the doses of 540–900 mg of EC-MPS.

A new approach to rapid determination of 18 bioactive constituents in Alpinia oxyphylla using UPLC-MS/MS with positive–negative conversion multiple reaction monitor (+/−MRM) technology

A new approach was developed for the simultaneous separation and determination of 18 active components (including one new component) in traditional Chinese medicines, Fructus Alpinia oxyphylla, by ultra high-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) with positive–negative conversion multiple reaction monitor (+/−MRM) technology. The chromatographic separation was performed on a Waters ACQUITY UPLC® HSS C18 column (2.1 mm × 50 mm, 1.8 μm) with gradient elution using 0.1% formic acid aqueous solution (A) and acetonitrile (B) at a flow rate of 0.3 mL min−1. This is the first report of the simultaneous determination of 18 bioactive constituents in Alpinia oxyphylla extract by UPLC-MS/MS. Good linear behaviors over the investigated concentration ranges were observed with the values of r higher than 0.998 for all the analytes. The method was reproducible with intra-and inter-day variation less than 3.0% and the recovery of the assay was in the range of 95.5% to 104.6%. The validated method was successfully applied for the analysis of 18 bioactive compounds in Fructus Alpinia oxyphylla extract from different sources and the analysis results were classified by hierarchical cluster analysis (HCA). This study provides a powerful and practical guidance to be used for the comprehensive quality control of Alpinia oxyphylla and lays the foundation for further research of Alpinia oxyphylla.

Metabolomics approach in lung tissue of septic rats and the interventional effects of Xuebijing injection using UHPLC-Q-Orbitrap-HRMS

&NA; Sepsis is the dysregulated host response to an infection which leads to life‐threatening organ dysfunction. Metabolomic profiling in bio‐fluid or tissue is vital for elucidating the pathogenesis of sepsis and evaluating therapeutic effects of medication. In this study, an untargeted metabolomics approach was applied to study the metabolic changes in lung tissue of septic rats induced by cecal ligation and puncture (CLP) and investigate the treatment effects of Xubijing injection (XBJ). Metabolomics analyses were performed on ultra‐high performance liquid chromatography‐Q Exactive hybrid quadrupole‐orbitrap high‐resolution accurate mass spectrometry (UHPLC‐Q‐Orbitrap‐HRMS) together with multivariate statistical analysis. A total of 26 differential metabolites between CLP and sham‐operated group were identified. The altered metabolic pathways included energy metabolism, amino metabolism, lipid metabolism, fatty acid metabolism and hormone metabolism. Among the 26‐varied metabolites, 15 were significantly regulated after XBJ treatment. The metabolic pathway network of sepsis was drawn to interpret the pathological feature of lung damage caused by sepsis and the underlying regulating mechanism of XBJ on the molecular levels. Our findings display that LC‐MS‐based metabolomics is a useful tool for uncovering the underlying molecular mechanism of sepsis, and XBJ may exert therapeutic effect by regulating multiple metabolic pathways.

Antiseptic Activity of Ethnomedicinal Xuebijing Revealed by the Metabolomics Analysis Using UHPLC-Q-Orbitrap HRMS

Xuebijing (XBJ) injection is an ethnomedicinal formula that has been widely used in the therapy of sepsis in China. However, the underlying theraputic mechanisms remain uninvestigated. In this research, a metabolomic method based on UHPLC-Q-Orbitrap HRMS was applied to make a holistic evaluation of XBJ on septic rats which were induced by the classical cecal ligation and puncture (CLP) operation. The plasma metabolic changes were profiled and evaluated by multivariate analytical (MVA) methods. In the results, a total of 41 differential metabolites were identified between CLP-operated group and sham-operated group, which were mainly involved in amino acid metabolism and lipid metabolism. After pathway analysis, it was finally discovered that the majority of the influenced metabolic pathways caused by sepsis mainly involved in energy metabolism, oxidative stress, and inflammation metabolism. When intervened by XBJ injection, 32 of the 41 disordered metabolites had been adjusted in reverse, which suggested that XBJ could mediate the abnormal metabolic pathways synergistically. In conclusion, the present study systematically investigated the efficacy and its underlying therapeutic mechanisms of XBJ on sepsis, while offering a new insight for the subsequent relevant exploration of other Chinese medicine at the same time.

Pharmacokinetics of Mycophenolate Mofetil and Development of Limited Sampling Strategy in Early Kidney Transplant Recipients

The mycophenolate mofetil (MMF) dose management for optimization of post-transplant treatment especially the early postoperative phase has been well recognized. MMF is a pro-drug of mycophenolic acid (MPA) and is widely used in Chinese renal transplant patients. Until now, the pharmacokinetic (PK) characteristics and model for the area under the concentration–time curve for the 12-h (h) of exposure (AUC0-12 h) of MPA (MPA-AUC0-12 h) estimation were lacking for the new formulation of MMF dispersible tablet in renal transplant patients. The aims of the study were to investigate the PK characteristics of MMF dispersible tablet by detecting the active metabolite of MPA and to establish an accuracy and precision equation for calculating MPA-AUC0-12 h by limited sampling strategy (LSS) in Chinese kidney transplant patients. A total of 60 postoperative kidney transplant recipients were given a multiple-dose of MMF dispersible tablet twice daily combination with tacrolimus (Tac) and steroids. On the 5th day post-transplantation, blood specimens were collected before drug administration and up to 12 h after MMF dispersible tablet administration. Non-compartmental PK analysis was used to determine the data obtained from individual patients. Multivariate stepwise regression analysis was used to develop models for predicting MPA-AUC0-12 h. The 3- and 4-point sampling models using 2 h, 4 h, 8 h and 1 h, 2 h, 4 h and 8 h, respectively, allowed accurate estimation of MPA-AUC0-12 h. PK parameters of MMF dispersible tablet were obtained and the 4-point LSS is the best model for accurate and precise estimation of MPA-AUC0-12 h.