Xiaotian Li

Metabonomic profiling in studying anti-osteoporosis effects of strontium fructose 1,6-diphosphate on estrogen deficiency-induced osteoporosis in rats by GC/TOF-MS

A novel strontium salt compound strontium fructose 1, 6-diphosphate (FDP-Sr) has been proved to have highly effective for bone loss via dual effects of stimulating bone formation and suppressing bone absorption. In the present study, metabolomic approach was used to identify and study potential biomarkers associated with the effect and safety of FDP-Sr. The metabolomic profiles of bone loss induced by estrogen deficiency in a rat model was described to attain a system-level map of the shift on the metabolic response in plasma using GC/TOF-MS, after FDP-Sr was orally administered at the dose of 110 mg/kg/day for the prevention and 220 mg/kg/day for the treatment. Meanwhile, bone turnover biomarkers and bone mineral density were investigated to identify the specific changes of potential anti-osteoporosis effects of FDP-Sr. The differences in metabolic profiles between osteoporosis rats and FDP-Sr treated rats were well observed by the partial least squares-discriminant analysis (PLS-DA) to the MS spectra. Some metabolites including homocysteine, arachidonic acid, alanine, and hydroxyproline, which significantly changed during osteoporosis progression could be effectively reversed after FDP-Sr therapy. Of course some metabolites such as uric acid, glyceric acid, octadecadienoic acid, docosahexaenoic acid, oleic acid, and hexadecanoic acid were not found to reverse significantly after FDP-Sr administration. These results delineated the FDP-Sr effects-related metabolic alterations in the bone loss rats, suggesting that metabonomic analysis could provide helpful information on the new potential biomarkers relating to the mechanism of anti-osteoporosis action and side effects of FDP-Sr against estrogen deficiency induced bone loss.

Simultaneous determination of Eleutheroside B and Eleutheroside E in rat plasma by high performance liquid chromatography–electrospray ionization mass spectrometry and its application in a pharmacokinetic study

Eleutheroside B and Eleutheroside E, two kinds of the major bioactive saponins of Eleutherococcus senticosus, play a pivotal role in biologic activity. In this study, a specific and sensitive high performance liquid chromatography-electrospray ionization-tandem mass spectrometry method (HPLC-MS/MS) was developed and validated for simultaneous determination of Eleutheroside B and Eleutheroside E in rat plasma. The analytes were extracted from rat plasma via a simple protein precipitation procedure with methanol and polygonin was used as internal standard. Chromatographic separation was achieved on a C18 column using a gradient elution program with acetonitrile and water containing 0.1% ammonium hydroxide solution as the mobile phase, with a flow rate of 0.2mL/min. The detection was performed on a triple-quadrupole tandem mass spectrometer by multiple reactions monitoring (MRM) mode in a negative ion mode via electrospray ionization (ESI). The transition monitored were m/z 371 [M-H](-)→209 for Eleutheroside B, m/z 741[M-H](-)→579 for Eleutheroside E and m/z 389[M-H](-)→277 for internal standard. Linear calibration curves were obtained in the concentration range of 1-2000ng/mL for both (Eleutheroside B and Eleutheroside E), with a lower limit of quantification of 1ng/mL. Extraction recovery was over 80% in plasma. The intra- and inter-day precision (RSD) values were below 12% and accuracy (RE) was -2.80 to 5.70% at three QC levels for both. The assay was successfully applied to study pharmacokinetics behavior in rats after oral and intravenous administration of the single substances (Eleutheroside B and Eleutheroside E). And further research was performed by comparing the difference in pharmacokinetic behavior between the single substances and an aqueous extract of E. senticosus after oral administration. Significant difference in pharmacokinetic characteristics between the single substances and an aqueous extract was found in rat, which would be beneficial for the pre-clinical research and clinical use of E. senticosus.

Pharmacokinetics and tissue distribution of Aucubin, Ajugol and Catalpol in rats using a validated simultaneous LC-ESI-MS/MS assay.

To enable an investigation of pharmacokinetics and tissue distribution of Aucubin, Ajugol and Catalpol in rats, a high-performance liquid chromatography-electro spray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) method was developed for the simultaneous quantitative determination of the three compounds. Biological samples were prepared by a simple protein precipitation with methanol (containing 0.05% formic acid). The analytes were separated by a C18 reversed phase column and detected with a triple quadrupole tandem mass spectrometer in the multiple-reaction monitoring (MRM) mode to monitor the precursor-to-product ion transitions of m/z 364.3→149.0 for Aucubin, m/z 366.5→151.1 for Ajugol, m/z 380.0→183.3 for Catalpol and m/z 530.3→183.1 for Picroside-II (IS) in positive ionization. Good linearity of each calibration curve was produced over the concentration range of 1-1000ng/mL. The lower limit of quantification (LLOQ) was 1ng/mL for the three analytes. This method was successfully applied to the pharmacokinetic and tissue distribution studies of Aucubin, Ajugol and Catalpol in rat. The current results revealed pharmacokinetic behaviors of the herb compound and provided novel evidence of the presence of Aucubin and Catalpol in rat brain. The acquired data would be helpful for the clinical application and further studies of Traditional Chinese Medicines (TCM) with active ingredients of Iridoid Glycosides.

Determination of antazoline hydrochloride in rat plasma and excreta by reversed-phase ion-pair chromatography and its application to pharmacokinetics

A reversed-phase ion pair chromatography method with liquid-liquid extraction analytical method was developed and validated for the determination of antazoline hydrochloride in plasma and excreta of rat. The aim of our study was to characterize the preclinical pharmacokinetics and excretion profiles of antazoline hydrochloride in rats after intravenous injection at the dose of 10 mg/kg. Plasma and excreta samples were extracted with ethyl acetate, and phenacetin was used as the internal standard. The result showed that the method is suitable for the quantification of antazoline hydrochloride in plasma and excreta samples. Analysis of accuracy (90.89-112.33%), imprecision (<7.1%) and recovery (>82.5%) showed adequate values. After a single intravenous administration at 10 mg/kg to rats, plasma concentration profile showed a relative fast elimination proceeding with a terminal elimination half-life of 3.53 h. Approximately 61.8 and 14.2% of the administered dose were recovered in urine and bile after 72 and 24 h post-dosing respectively; 5.9% of the administered dose was recovered in feces after 72 h post-dosing. The above results show that the major elimination route is urinary excretion.

Determination of antazoline hydrochloride in Beagle dog plasma by HPLC-UV and its application to pharmacokinetics.

In order to evaluate the pharmacokinetics characteristic of antazoline hydrochloride in Beagle dogs, a sensitive and specific HPLC method was developed and validated using phenacetin as the internal standard (IS). The analyte and the IS were extracted from dog plasma by ethyl acetate under the basic condition. The analyte was separated by a C18 column and detected with a variable wavelength UV-detector. The mobile phase consisted of methanol-5mmolL(-1) tetrabutyl ammonium bromide (45:55, v/v) containing 0.5% glacial acetic acid in a flow rate of 1.0mLmin(-1). Standard calibration graph for antazoline was linear over a curve range of 20-1600ngmL(-1) (R>0.99) and the lower limit of quantification was 20ngmL(-1) using a plasma sample of 500μL. The intra- and inter-day precision values were less than 14.3% relative standard deviation (RSD). The intra-day assay accuracy was in the range of 98.1-100.6% and the inter-day assay accuracy in the range of 99.2-101.1%. The extraction recoveries were on the average of 88.4% for antazoline and 76.8% for IS. Plasma samples were stable at least for 1 month at -20°C. This method was successfully applied to pharmacokinetics study of antazoline after intravenous administration to Beagle dogs.

Simultaneous determination of Guanfu base G and its active metabolites by UPLC-MS/MS in rat plasma and its application to a pharmacokinetic study.

To establish a rapid and sensitive ultra performance liquid chromatography mass spectrometry (UPLC-MS/MS) method for the determination of concentration of guanfu base G (GFG) and its active metabolites in rat plasma. The GFG and its active metabolites and the internal standard (phenacetin) were separated on an Acquity UPLC(®) BEH C18 chromatography column (2.1mm×50mm I.D., 1.7μm) using gradient elution with a mobile phase of methanol and ultrapure water at a flow rate of 0.4mL/min. The detection was performed on a Xevo triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode to monitor the precursor-to-product ion transitions of m/z 472.26→m/z 310.03 for GFG and m/z 180.00→m/z 109.99 for phenacetin (IS) using a positive electrospray ionization interface. The lower limit of quantification (LLOQ) was 1ng/mL, the limit of detection (LOD) was 0.3ng/mL, and the linear calibration curve was obtained over the concentration range of 1-200ng/mL. The intra-day and inter-day assay variations were measured to be below 10.97%, and the accuracy values (relative error) ranged from 95.4% to 103.6%. After validation, this method was successfully applied to a pharmacokinetic study where rats were intravenous administration of 5mg/kg GFG. A simple, rapid, sensitive, and accurate method for the determination of the concentration of GFG and its metabolites in rat plasma was developed and validated.

LC–MS-MS for the Determination of Ponicidin in Dog Plasma

A highly specific and sensitive liquid chromatography-tandem mass spectrometry method has been developed and validated for the determination of ponicidin in dog plasma. The plasma samples were prepared using liquid-liquid extraction with ethyl acetate as the extraction solvent. Chromatographic separation was accomplished on a Waters XTerra MS C18 column. The extracted ponicidin and the internal standard, oridonin, were detected by tandem mass spectrometry in positive electrospray ionization mode with multiple reaction monitoring. The optimized mass transition ion pairs (m/z) for quantitation were 363.08-345.08 for ponicidin and m/z 365.10-347.06 for the internal standard. The lower limit of quantification was 5 ng/mL. The linear range of the method was from 5 to 5,000 ng/mL. The intra-day and inter-day precision measurements were lower than 5.3 and 6.0% in terms of relative standard deviation and the accuracy was within ±8.4% in terms of relative error. Additionally, no significant matrix ionization suppression or enhancement was observed. The validated method was successfully applied in a pharmacokinetic study of ponicidin in dogs. The primary pharmacokinetic parameters in dogs were: terminal elimination half-life, 8.14 ± 1.35 h; mean residence time, 12.30 ± 2.08 h; area under the plasma concentration-time curve from time zero to the last measurable concentration, 14.34 ± 1.37 µg/h/mL; area under the plasma concentration-time curve from time zero to infinity, 15.75 ± 1.44 µg/h/mL; apparent volume of distribution, 4.79± 1.68 L/kg; total body clearance, 0.41 ± 0.08 L/kg/h.

Pharmacokinetic Study of Guanfu Base G in Rats by LC–ESI-MS

A sensitive and simple liquid chromatography -: electrospray ionization mass spectrometry method has been established and validated for the quantification of Guanfu base G in rats. Phenoprolamine hydrochloride was selected as the internal standard. Sample preparation involved simple liquid-liquid extraction by ethylacetate with high efficiency. The chromatographical separation was performed on a Shimadzu C18 column (150 × 2.0 mm, 5 µm) with a gradient elution of 0.2% acetic acid-acetonitrile (30:70, v/v). The method was sensitive with the lowest limit of detection at 1 ng/mL (S/N ≥ 3) in 100 µL of rat plasma. Good linearity (r = 0.9996) was obtained covering a concentration of 5-2000 ng/mL. The intra- and interday assay precision ranged from 4.3 to 6.1% and 5.4 to 8.3%, respectively. In addition, the stability, extraction recovery and matrix effect involved in the method were also validated. After intravenous dosing, rat plasma Guangfu base G (GFG) concentration declined in a biphasic manner with a terminal elimination half-life of 3.72 h. The total plasma clearance values were 1.15 L/h/kg. After oral dosing, the plasma GFG concentration reached a maximum within 0.5 h. The absolute bioavailability of GFG was 83.06%.