[Effect of origin, tree age, and harvesting time on content of flavonoids and terpene lactones in Ginkgo Folium].

The content of total flavonol glycosides in Ginkgo Folium in the planting bases was determined by high performance liquid chromatography(HPLC).The samples were extracted by reflux with methanol-25% hydrochloric acid.The HPLC conditions were as follows: Agilent ZORBAX SB-C_(18) column(4.6 mm×250 mm, 5 µm), isocratic elution with mobile phase of 0.4% phosphoric acid solution-methanol(45∶55), flow rate of 1 mL·min~(-1), column temperature of 30 ℃, detection wavelength of 360 nm, and injection vo-lume of 10 µL.A method for the determination of terpene lactones in Ginkgo Folium was established based on ultra-high performance liquid chromatograph-triple-quadrupole/linear ion-trap tandem mass spectrometry(UPLC-QTRAP-MS/MS).The UPLC conditions were as below: gradient elution with acetonitrile-0.1% formic acid, flow rate of 0.2 mL·min~(-1), column temperature of 30 ℃, sample chamber temperature of 10 ℃, and injection volume of 10 µL.The ESI~+and multiple reaction monitoring(MRM) were adopted for the MS.The above methods were used to determine the content of total flavonol glycosides and terpene lactones in 99 batches of Ginkgo Folium from 6 planting bases, and the results were statistically analyzed.The content of flavonoids and terpene lactones in Ginkgo Folium from different origins, from trees of different ages, harvested at different time, from trees of different genders, and processed with different methods was compared.The results showed that the content of total flavonol glucosides in 99 Ginkgo Folium samples ranged from 0.38% to 2.08%, and the total content of the four terpene lactones was in the range of 0.03%-0.87%.The method established in this study is simple and reliable, which can be used for the quantitative analysis of Ginkgo Folium.The research results lay a basis for the quality control of Ginkgo Folium.

[Effect of heavy metal lead on anticoagulant activity of leech based on label-free quantitative proteomics].

Whitmania pigra is the most widely distributed species of leeches in the market. In this study, the effect of heavy metal lead pollution on the anticoagulant activity of Wh. pigra was studied and the potential mechanism was explored. Pb(NO_3)_2 was used to contaminate the breeding soil which was then used to rear Wh. pigra for 50 days(lead-contaminated group, LC group), and meanwhile the blank control group(CG group) was set. Proteins were extracted from the obtained leech samples, and the differentially expressed proteins between LC and CG groups were analyzed by label-free proteomics technology. In this study, a total of 152 differentially expressed proteins were screened out, of which 93 proteins were up-regulated and 59 proteins were down-regulated in LC group. Bioinformatics analysis showed that the biological processes enriched with the differentially expressed proteins were mainly vesicle-mediated transport and transport positive regulation; the enriched cell components were mainly endocytosis vesicles and apical plasma membrane; the enriched molecular functions mainly included carbohydrate binding. The differentially expressed proteins were enriched in 76 KEGG pathways, which mainly involved metabolic pathways, biosynthesis of secondary metabolites, and bacterial invasion of epithelial cells. In this study, two differentially expressed proteins with Antistasin domain were presumed, which provides reference for further exploring the regulatory mechanism and signal transduction underlying the effect of lead pollution on the anticoagulant activity of leech.