A new G-triplex-based strategy for sensitivity enhancement of the detection of endonuclease activity and inhibition.

EcoRI is an important biomacromolecule in live cells and protects bacterial cells against foreign DNA. In this work, we developed a simple and convenient G-triplex (G3) (5'-TGGGAAGGGAGGGAATTCCCT-3')-based colorimetric assay for the rapid and selective detection of EcoRI activity and inhibition. The sequence specifically responds to EcoRI in the presence of K+ and hemin to form a G-triplex/hemin complex. Taking advantage of G-triplex, EcoRI activity was investigated under the optimized conditions. The absorption intensity ratio displayed a linear relationship against the concentration of EcoRI in the range 0 to 100 U mL-1, and the detection limit was 5.7 U mL-1. Furthermore, G3 showed good selectivity, and the ability to be used to screen for EcoRI inhibitors, indicating its potential in detection and analysis applications.

[Reversed-phase liquid chromatography separation and mass spectrometry fragmentation analysis of mitiglinide and three isomers].

The separation of mitiglinide and its three isomer impurities was achieved by reversed-phase ultra-performance liquid chromatography-high resolution mass spectrometry. An ACQUITY UPLC HSS T3 column (100 mm×2.1 mm, 1.8 µm) was used as the stationary phase and water-acetonitrile-n-pentanol (75:25:1, v/v/v; formic acid was added to adjust pH to 1.8) was used as the mobile phase with a flow rate of 0.4 mL/min. According to the exact mass and high resolution mass spectrometry fragmentation (Q Exactive), significant differences were observed in the fragment ion abundance in the secondary mass spectra of mitiglinide and its three isomer impurities. Two of these isomer impurities were newly discovered. The possible fragmentation mechanisms of mitiglinide and its three isomer impurities were also deduced. The limit of detection of the developed method was 1 µg/L. The linearity of the developed method was good from the limit of quantitation (2 µg/L) to 10000 µg/L with a correlation coefficient of 0.9999. The relative standard deviation of the peak area was 2.0%. On the basis of these results, the sources of the mitiglinide isomer impurities were discussed. Isomer impurity 1 was degraded at high temperature, while isomer impurities 2 and 3 were determined to be synthetic impurities. In addition, samples of mitiglinide calcium raw materials were analyzed.

Distribution and accumulation of caffeine in rat tissues and its inhibition on semicarbazide-sensitive amine oxidase.

Wistar rats were treated with caffeine or 2-bromoethylamine, the effect of caffeine on the activity of semicarbazide-sensitive amine oxidase (SSAO) in rat serum and tissues was studied using various LC-MS methods. Caffeine was found to present in all tissues after administration for 10 days and accumulated for 25 days. The level of caffeine was high in brain and liver, and the SSAO activity in all tissues was found to be inhibited by caffeine. As the concentration of caffeine increased, the SSAO activity decreased. The inhibition ratio was correlated to the levels of caffeine present. We presume that caffeine may treat with SSAO activity associated diseases.

[Simultaneous determination of 15 anti-obesity drugs in blood by high performance liquid chromatography-tandem mass spectrometry].

An analytical method for the simultaneous determination of 15 anti-obesity drugs (caffeine, sibutramine, phenformin, etc.) in blood sample was developed using high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). After a simple protein precipitation step, the HPLC separation was performed on an UltimateXB-C18 column with methanol and 20 mmol/L ammonium acetate (containing 0.1% (v/v) of glacial acetic acid) as the mobile phases in a gradient elution mode. The MS/MS detection was achieved by electrospray ionization in both positive and negative modes by rapid switching with selective reaction monitoring (SRM). The results showed that the limits of quantification of all anti-obesity drugs were in the range of 0.001 -0.05 mg/L. The calibration curves of all anti-obesity drugs showed good linearity and the correlation coefficients were more than 0.99. The recoveries of all antiobesity drugs at 3 spiked levels were in the range of 77.3% - 110.8% with the intra-day and inter-day precisions less than 12.3%. The mass spectrum characterizations of 15 anti-obesity drugs were studied. The method is sensitive and reproducible for the detection of the 15 anti-obesity drugs in blood, and can also be applied to the determination of the anti-obesity drugs in pharmaceuticals or foods.

[Simultaneous identification of fifteen glucocorticoids in Chinese traditional medicine preparations by liquid chromatography-mass spectrometry].

To monitoring illegal abuse of these drugs, a liquid chromatography-mass spectrometry (LC-MS) method for identification of glucocorticoids in Chinese traditional medicine preparations was developed. Samples were extracted using different methods according to the nature of products. The analyses were performed on a Diamond C18 column (5 microm, 250 mm x 4.6 mm) with gradient elution using water-tetrahydrofuran (100 : 1, v/v) as mobile phase A and acetonitrile-water-tetrahydrofuran (80 : 20 : 1, v/v) as mobile phase B. Prednisone acetate in the preparation used to cure dermatitis was identified by comparing the retention time, full scan MS, and full scan MS2.

[Determination of five components in compound hypotensive tablet by HPLC].

AIM: To establish a method for the determination of the five components (reserpine, chlordiazepoxide, hydrochlorothiazide, dihydralazine sulfate, triamterene) in compound hypotensive tablet. METHODS: The chromatography was performed using a CN column with acetontrile-0.1 mol L(-1) sodium heptasulfonate solution (7:3) and (5:5) as the mobile phases. The detection wavelength was 267 nm for reserpine, chlordiazepoxide and hydrochlorothiazide, 310 nm for dihydralazine sulfate, 360 nm for triamterene. RESULTS: The linear range of each component was tested, and the recovery and stability of each component was satisfactory, three lots of samples were determined using the method. CONCLUSION: This is an accurate and credible quality control method for compound hypotensive tablet.

Rapid determination of aristolochic acid I and II in Aristolochia plants from different regions by beta-cyclodextrin-modified capillary zone electrophoresis.

Chinese herbs nephropathy (CHN) is a kind of severe kidney disease caused by excessively taking aristolochic acid (AA). Hence, it is essential for health security and quality control of related herbal medicines to develop an efficient method for separation and determination of these two important components in Traditional Chinese Medicines. In this study, a rapid capillary zone electrophoresis (CZE) method using 120 mM sodium borate buffer containing 10 mM beta-cyclodextrin (beta-CD) as modifier was firstly developed for the analysis of AA-I and AA-II within 4min in some medicinal plant samples. The separation conditions including pH of running buffer, CD content in the buffer system, applied voltage and capillary temperature were systematically optimized, and two kinds of aristolochic acids in 37 herbal samples of Aristolochia plants were successfully determined with high separation efficiency, satisfactory sensitivity, repeatability and recovery. The result indicated high variability in the contents of aristolochic acids due to different species and regions. The comparison of CZE method with high performance liquid chromatography (HPLC) was also discussed.