A sensitive and selective liquid chromatography–tandem mass spectrometric (LC ? MS/MS) method was established to determine 2-oxo-clopidogrel, a crucial intermediate metabolite in human plasma. A chromatographic separation was performed on a Sapphire C18 column following a liquid–liquid extraction sample preparation with methyl t-butyl ether. Detection was carried out on a triple quadrupole mass spectrometer operated in multiple reaction monitoring (MRM) with an electrospray ionization (ESI) mode. The method was validated in terms of specificity, accuracy, precision and limit of quantification. The calibration curves ranged from 0.50 to 50.0 ng/mL with good linearity. The stability was fully validated with addition of 1,4-dithio-DL-threitol (DTT) into the plasma sample prior to and in the preparation procedure. The validated method was proved to be suitable for use in pharmacokinetic study after single oral administration of 75 mg clopidogrel tablets in human subjects, which could make contribution to intensive study of the clinical drug–drug interactions of clopidogrel and individual treatment.

A highly sensitive, rapid and selective liquid chromatography-tandem mass spectrometry(LC-MS/MS) method for the quantitative determination of retinamido-ester in rat plasma was developed and validated. A simplified protein precipitation with acetonitrile was employed for the sample preparation.The separation was carried out on an Agilent TC C18 column ( 150 mm×4. 6 mm ID, 5 μm particle size)with the mobile phase consisted of methanol-water-formic acid (93 : 7 : 0.1). Simvastatin was used as internal standard. The detection was performed on a trap-quadrupele tandem mass spectrometer by selected reaction monitoring (SRM) scan mode via atmospheric pressure chemical ionization (APCI). The range of and inter-day precision values were between 95.97% and 104. 43%, and RSD was between 4. 63% and 10. 69%, respectively. This method was applied to determine the pharmacokinetic parameters. The main pharmacokinetic parameters of retinamido-ester after oral administration via gastric gavage of 2. 5, 5, 10mg·kg-1 were as follows,T1/2;(11.28±7.23),(8.90±3.82),(8.01±5.56)h; AUC0-∞:(103.41±61.46),(190.23±74.99),(421.66±299.20)ng·h·mL-1;MRT:(6.31±0.75),(5.98±0.71), (6.18±0.97) h; CL/F: (30. 10 ± 13.67), (29.58±10.59), (31.18 ±17.51)L·h-1·kg-1;Vd/F:(414.94±159.82),(356.16±139.85),(369.28±322.73)L·kg-1,respectively.

The UPLC fingerprint of colistimethate sodium was established for the study of quality consistency.The chromatographic column was Acquity UPLC? Peptide CSH C18 (2.1 mm × 150 mm, 1.7 μm).The mobile phase A was phosphate buffer-acetonitrile (19∶1), and the mobile phase B was phosphate buffer-acetonitrile (1∶1).The mobile phase was in gradient elution at a flow rate of 0.3 mL/min.The column temperature was set at 30 °C and the detection wavelength was 210 nm.The similarity of the fingerprints was analyzed with the Similarity Evaluation System for Chromatographic Fingerprint of Tradition Chinese Medicine (Version 2012) in combination with content determination of multiple index components to evaluate the quality consistency of imported and domestic bulk drugs.The result showed that both the original and generic bulk drugs met the specified limit requirements in the European Pharmacopoeia standards, and that their UPLC fingerprints were highly similar, indicating that the quality of the two substances was consistent.Establishing a fingerprint for similarity evaluation and combining it with the results of indicator component content determination as a comprehensive evaluation method for the study of drug quality consistency of complex components has the characteristics of fast, accurate, and comprehensive, which is helpful for drug quality evaluation and provides ideas for the evaluation of antibiotic quality consistency of complex components.

Objective:To learn about the knowledge, attitude and behavior of brucellosis and brucellosis infection in Tongyu County, Jilin Province, and to provide a reference for the formulation of prevention and control measures in relocated poverty alleviation and relocation areas.Methods:Using a multi-stage sampling method, three townships, Xianghai Township, Wulanhua Town and Xinhua Town were selected in Tongyu County, where the incidence of brucellosis is high in Jilin Province; then Miren, Huimin, Longjing, Xinfeng and Dayou villages from the three townships were surveyed. A face-to-face questionnaire survey was conducted among the villagers to collect the knowledge, attitude and behavior of brucellosis; and blood samples were collected from the villagers according to the principle of informed consent for brucellosis serological tests, and according to the "Diagnostic criteria for Brucellosis (WS 269-2007)" for diagnosis.Results:A total of 274 questionnaires were distributed and 244 valid questionnaires were collected with a recovery rate of 89.05%. Among them, 233 people had heard of brucellosis, accounting for 95.49%. The total awareness rate of knowledge related to brucellosis prevention and control was 39.29% (2 205/5 612), of which the total awareness rate of "knowledge related to brucellosis prevention" was 71.99% (527/732), and the total awareness rate of "knowledge related to human infection with brucellosis" was 38.08% (1 115/2 928), and the total awareness rate of "knowledge about brucellosis in animals" was 28.84% (563/1 952). A total of 90.13% (21/233) people supported the brucellosis prevention and control plan (such as free screening for brucellosis, publicity and education, immunization, culling, etc.), and 61.54% (48/78) of the people would take the initiative to buy protective equipment. The contact rates of livestock through assisted feeding and slaughtering were 27.46% (67/244) and 11.48% (28/244), respectively, and the total protection rates of sheep pens cleaning and slaughtering were 30.91% (102/330) and 21.43% (30/140), respectively. In terms of brucellosis infection, a total of 1 confirmed case, 1 latent infection and 37 suspected cases were found.Conclusions:The awareness rate of "knowledge related to brucellosis prevention" in Tongyu County is generally good, but the awareness rate of "knowledge about brucellosis in animals" is low, and the protection rate of some brucellosis prevention and control behaviors is poor. Health and epidemic prevention departments should carry out targeted health education, improve and optimize propaganda methods in order to improve the protection level of the general population and reduce the risk of brucellosis infection.

OBJECTIVETo investigate the effects of reducing APE/Ref1 expression in the cultures of rat spiral ganglion cells with oxidative damage induced by H(2)O(2).

METHODSPrimary cultured rat spiral ganglion cells were infected with small interfering RNA to APE/Ref1 (Ape1siRNA) for 72 h, followed by treating with H(2)O(2) (0, 10, 25, 50, 100 and 300 µmol/L) for 1 h , and then cultured in normal medium for 24 h. Western blot were used to detect the level of APE/Ref1 protein and phosphorylation of histone protein H2AX in the infected cells. The caspase3 activation was tested by spectrophotometric method . The cell viability was determined by MTT and the apoptosis of spiral ganglion cells was determined by terminal-deoxynucleotidyl transferase mediated nick and labeling (TUNEL).

RESULTSWestern blot showed that infection with Ape1siRNA resulted in APE/Ref1 reduced expression in the spiral ganglion cells. Exposing spiral ganglion cultures with reduced expression of APE/Ref1 to H(2)O(2) (50, 100, 300 µmol/L) for 1 h resulted in increasing in the phosphorylation of histone protein H2AX. The reduction in APE/Ref1 significantly reduced cell viability in cultures 24 h after 1 h expression to 50-300 µmol/L H(2)O(2). The apoptosis of cells and caspase 3 activity was detected significantly improved.

CONCLUSIONSThe induced of APE/Ref1 results in significantly decrease in spiral ganglion cells viability in oxidative stress. The repairing function of APE/Ref1 is necessary for optimal levels of neuronal rat spiral ganglion cells survival.

Animals ; Cells, Cultured ; DNA-(Apurinic or Apyrimidinic Site) Lyase ; genetics ; Gene Silencing ; Hydrogen Peroxide ; Oxidation-Reduction ; Oxidative Stress ; RNA, Small Interfering ; Rats ; Rats, Sprague-Dawley ; Spiral Ganglion ; cytology ; metabolism