Study Of Tooth Leaves Exochorda Leaf Chemical Constituents And Pharmacological Effects

Objective:To investigate the effects of leaves of Exochorda serratifolia S.Moore on the glucose tolerance in normal and diabetic mice and to study the hypoglycemic effect on STZ induced diabetic mice and study the chemical constituents of the leaves of Exochorda serratifolia S.Moore. To establish the assay method of the content of apigenin-7-O-β-D- glucopyranoside and apigenin-7-O-β-D-neospheroside at one time . To establish the assay method of the content of total flavonoids of Exochorda serratifolia S.Moore and assay the content of total flavonoids in different parts and different batches of the leaves with this method.Analyzing the difference of the chemical constituents between the leaves and other parts of Exochorda serratifolia S.Moore using the HPLC. To study the enrichment of the leaves of total flavonoids of Exochorda serratifolia S.Moore resulted from macroporous resin. Materials and methods:Materials:Leaves which were from Lingyuan, Chaoyang city and were identified as the Exochorda serratifolia S.Moore by professor Wang bing who comes from Plants office of Liaoning University of TCM.Methods:1. First,diabetic mice model was induced by STZ(150mg·kg-1). Normal or diabetic mice were given a single dose of the water extract from leaves and the 80% EtOH extract from leaves,and the changes of glucose tolerance were observed respectively;Second,observe the effect about the mice's blood glucose level of the water extract and the 80% EtOH extract meformin hydrochloride (333mg·kg-1) respectively.2. Isolation and purification were carried out on the chromatography of silica gel and Sephadex LH-20. Their structures were elucidated on basis of physicochemical properties and spectral data; Preparation of liquid relative to compounds was used for the preparation and purification and their structures were elucidated on basis of physicochemical properties and spectral data.3. It was used to test the content of apigenin-7-O-β-D- glucopyranoside and apigenin-7-O-β-D-neospheroside in the leaves of Exochorda serratifolia S.Moore with HPLC; Ultraviolet spectrophotometry was used at 267nm with rutin as the reference to detect the content of total flavonoids of leaves.4. HPLC and gradient elution were used to analyse the chemical constituents of different parts;Ultraviolet spectrophotometry was used at 267nm with rutin as the reference to detect the content of total flavonoids of different parts.5. Leaves of Exochorda serratifolia S.Moore were extracted with 50% EtOH by reflux extraction three times,one hour one time.And purified with D101 macroreticular resin.Results:1.The water extract and the 80% EtOH extract could lower the serum glucose levels 1h and 2h after glucose administration in normal or diabetic mice. And the water extract could lower the the serum glucose levels 2h after given glucose in diabetic mice(P<0.05);The blood glucose level was lower remarkably in the hyperglycemia mice that were administrated at high dose of the water extract of leaves of Exochorda serratifolia S.Moore (P<0.05).2. 23 Compounds were isolated from this plant for the first time.,19 Compounds structures were elucidated on basis of physicochemical properties and spectral data.(the rest four compounds were Long-chain unsaturated fatty acids ):β-sitostero(l1),18 carbonated single gc-ms(2),oleanolic acid(3),ursolic acid(4),apigenin(5),protocatechuic acid(6),luteolin(7),quercetin(8),daucosterol(9),apigenin-7-O-β-D- glucopyranoside(10),quercetin-3-O-β-D-glucopyranoside(11),apigenin-7-O-β-D-neospheroside(12),trans-Hydroxyl cinnamic acid(13),cis- Hydroxyl cinnamic acid(14),(5-oxo-2,5-dihydrofuran-3-yl) methyl (2E)-3-(4-hydroxyphenyl) acrylate(15),(5-oxo-2,5-dihydrofuran-3-yl) methyl (2Z)-3-(4-hydroxyphenyl) acrylate(16),caffeic acid(17),rutin(18),quercetin -7-O-β-D-neospheroside(19).3.The liner ranges of apigenin-7-O-β-D-neospheroside and apigenin-7-O-β-D- glucopyranoside were 1.424-5.696μg(r=0.9996)和0.17-1.02μg(r=0.9995),respectively.The recoveries of apigenin-7-O-β-D-neospheroside and apigenin-7-O-β-D- glucopyranoside were 98.1%和101.3%. The liner ranges of rutin was 5.6-22.4μg·mL^-1(r=0.9998),the recoveries was 97.77%, RSD=1.78%(n=3).4. Ultraviolet spectrophotometry was used at 267nm with rutin as the reference to detect the content of total flavonoids of different parts,the content of total flavonoids in flowers and leaves were higher than that of roots and stems. Through HPLC gradient elution to different parts,the difference of effective ingredient between apigenin-7-O-β-D-neospheroside and apigenin-7-O-β-D- glucopyranoside is obvious differences.5. The content of total flavonoids in the extract is 59.44%.Conclusion:1. The leaves of Exochorda serratifolia S.Moore could improve the glucose tolerance in normal and diabetic mice induced by STZ and also had an hypoglycemic effect on STZ induced diabetic mice.2. 23 Compounds were isolated from this plant for the first time. Among them there were two pairs of isomers,contains a pair of new compouds.3. The method of using HPLC and gradient elution to determine the chemical constituents of apigenin-7-O-β-D-neospheroside and apigenin-7-O-β-D-glucopyranoside of leaves of Exochorda serratifolia S.Moore is accurate and reliable. It can be used for the quality control of the leaves of Exochorda serratifolia S.Moore. The method of Ultraviolet spectrophotometry is accurate and reliable that can be used to determine the content of total flavonoids of Exochorda serratifolia S.Moore.4. To analysis the different parts and determine their content of total flavonoids, we can get the differences of the chemical constituents and differences of the contents of total flavonoids.5. D101 macroreticular resin fits in purification of the total flavonoids in leaves of Exochorda serratifolia S.Moore.