A thin-layer chromatography-bioautographic method for detecting dipeptidyl peptidase IV inhibitors in plants

A thin-layer chromatography (TLC)-bioautographic method was developed with the aim to detect dipeptidyl peptidase IV (DPP IV) inhibitors from plant extracts. The basic principle of the method is that the enzyme (DPP IV) hydrolyzes substrate (Gly-Pro-p-nitroaniline) into p-nitroaniline (pNA), which diazotizes with sodium nitrite, and then reacts with N-(1-naphthyl) ethylenediamine dihydrochloride in turn to form a rose-red azo dye which provides a rose-red background on the TLC plates. The DPP IV inhibitors showed white spots on the background as they blocked enzymolysis of the substrate to produce pNA. The method was validated with respect to selectivity, sensitivity, linearity, precision, recovery, and stability after optimizing key parameters including plate type, time and temperature of incubation, concentration of substrate, enzyme and derivatization reagents, and absorption wavelength. The results showed good lineary within amounts over 0.01–0.1 μg range for the positive control, diprotin A, with the coefficient of determination (r2) = 0.9668. The limits of detection (LOD) and quantification (LOQ) were 5 and 10 ng, respectively. The recoveries ranged from 98.9% to 107.5%. The averages of the intra- and inter-plate reproducibility were in the range of 4.1–9.7% and 7.6–14.7%, respectively. Among the nine methanolic extracts of medicinal herbs screened for DPP IV inhibitors by the newly developed method, Peganum nigellastrum Bunge was found to have one white active spot, which was then isolated and identified as harmine. By spectrophotometric method, harmine hydrochloride was found to have DPP-IV inhibitory activity of 32.4% at 10 mM comparing to that of 54.8% at 50 μM for diprotin A

Determination of the number of J/ψ events with inclusive J/ψ decays

A measurement of the number of J/ψ events collected with the BESIII detector in 2009 and 2012 is performed using inclusive decays of the J/ψ. The number of J/ψ events taken in 2009 is recalculated to be (223.7 ± 1.4) × 106, which is in good agreement with the previous measurement, but with significantly improved precision due to improvements in the BESIII software. The number of J/ψ events taken in 2012 is determined to be (1086.9 ± 6.0) × 106. In total, the number of J/ψ events collected with the BESIII detector is measured to be (1310.6 ± 7.0) × 106, where the uncertainty is dominated by systematic effects and the statistical uncertainty is negligible