Chemical Composition And The Sedative And Hypnotic Effects Of The Fruit Pulp From Ziziphus Jujuba Ver. Spinosa

Suan-Zao [Ziziphus jujuba Mill. var. spinosa (Bunge) Hu ex H. F. Chow], the Rhamnaceae family, is widely distributed throughout China with a rich amount. Suan-Zao-Ren, the Kernels of this plant, is a well-known traditional Chinese medicine with sedative and hypnotic effects. As a by-product of processing Suan-Zao-Ren, the Suan-Zao pulp (SZP) is usually used for production of juice, while it is not found medicinal use. Therefore, a series of investigation on SZP were performed in the thesis, including the isolation and identification of triperpenes; establishment of a HPLC method for determination of triterpenes and comparison of their contents in the samples collected from different regions; studies on the sedative and hypnotic activities using animal model; development of a novel LC-MS/MS method for determination of7kinds of neuro-transmeters and determination of these neuro-transmeters in mice brain after oral administration of SZP80%ethanol extract. The purpose of the studies was to provide scientific evidences for the further development of this natural resource.Using silica Gel column chromatography,12triterpenes were isolated from the ethyl acetate fraction of SZP80%ethanol extract. The chemical structures of7of these compounds were identified as3-O-cis-p-Coumaroyl alphitolic acid,3-O-cis-p-Coumaroyl maslinic acid,3-O-trans-p-Coumaroyl alphitolic acid,3-O-trans-p-Coumaroyl maslinic acid, Oleanolic acid, Ursolic Acid and3-O-one-Ursolic Acid, based on their physico-chemical properties and various spectrometry data. Others need further identification.A LC-UV method was firstly developed for simultaneous determination of the contents of12compounds isolated from SZP and comparison of their HPLC profiles of SZP collected from different regions of China. SZP sample was immersed in80%ethanol for the extraction under the sonic condition. The extract solution was then chromatographied on a C18HPLC column. The results showed that the method achieved excellent linearity, precision, reproducibility and extraction recovery and was suitable for analysis of the compounds in SZP. The analytical results showed that the chemical components were similar among the different SZP collected from different regions. However, their contents were varied among the samples from different regions. It was found that the contents of the triterpenes in SZP collected from Luo-Nin county of Henan province were relatively higher than others. The results indicate that the method is easy to conduct, accurate and reliable, and could be used for quality control purpose.The effects of SZP80%ethanol extract on the behaviors of spontaneous motion and sleeping, and the contents of brain neurotransmitters of mice after oral dosing were investigated. The results both from open-field and light-dark transition tests showed that the extract (10g/kg) could markedly inhibit the spontaneous motion, shortened the total distance of motion, and decreased the frequency of light-dark transition in dose-dependent. The extract could increase the number of sleeping, shorten the sleeping latency time and prolong the sleeping time of mice, which sleeping was produced by i.p. injection of pentobarbital sodium. The effect of the extract on the contents of mice brain neurotransmitters was similar with that of estazolam. The brain content of5-HT, an inhibitory neurotransmitter was significantly increased after oral dose of SZP extract. These results indicate that SZP has a remarkable sedative-hypnotic activity.A novel sensitive and reliable LC-MS/MS method was developed for simultaneous determination of7kinds of neurotransmitters, namely dopamine (DA), epinephrine(E), Noradrenaline Bitartrate (NE),5-Hydroxytryptamine (5-HT), acetylcholine (Ach), y-aminobutyrate (GABA) and glutamic acid (Glu) in mice brain. The validation data indicated that the method showed excellent sensitivity, linearity, precision and accuracy. The method was proved suitable to determination of the neurotransmitters in animal brain by assay of mice brain samples after i.p. injection of donepezil and scopolamine.