Study Of Bioactive Components Extracted From Sea Buckthorn And Cardiac Protective Effect On Hearts

Sea buckthorn(Hippopha? rhamnoides L.) is a deciduous shrub, which fruit is considered to be a rich source of biologically active substances like vitamin, microelement, flavonoid, polyphenol, fatty acid etc. It also has lots of physiological functions, such as lower cholesterol, blood pressure and blood sugar. In this thesis, extract yield and antioxidant capacities were used as indexes to choose suitable extraction solvent by ultrasound-assisted solvent extraction method. Then through single-factor experiments, the optimum technological parameters of purification were established. Experiments on the isolated rat hearts were carried out according to improved Langendorff retrograde perfusion technology to investigate the protective of bioactive components from sea buckthorn(BCSB) against myocardial ischemia reperfusion injury(MIRI). This research revealed the mechanism of BCSB against MIRI, which may be attributed to the inhibition of autophagy. At last, these studies offered theoretical and experiment basis for the development and utilization of sea buckthorn. This thesis mainly contained two chapters as follows:1. To choose suitable extraction solvent, extract yield and antioxidant capacities(scavenging DPPH and hydroxyl radical) were used as indicators. Different volume fraction of ethanol, acetone and water was used to extract BCSB by ultrasound-assisted solvent extraction method. What's more, we found that polyphenols were the main ingredients in extracts. So polyphenols were regarded as the reference during purification experiments. Then through single-factor experiments, the optimum technological parameters of purification were established. Firstly, during five tested macroporous resins, NKA-2 was the most desirable adsorption and desorption material for polyphenol from sea buckthorn. Secondly, when 0.20 mg/mL sample was flowed through NKA-2 resin column at a speed of 1.0 mL/min, the adsorption reached dynamic equilibrium. After that, the adsorbed extracts could be eluted with 90 mL of 60% ethanol. Lastly, the extracts were analyzed by ultra-high performance liquid chromatograph. Our results show that the ingredients of the extracts including gallic acid,(+)-catechin, caffeic acid, chlorogenic acid, quercetin, kaempferol and isorhamnetin etc.2. The cardiac protective effect and mechanism of BCSB against MIRI on isolated rat hearts were studied by improved Langendorff retrograde perfusion technology. Through monitoring heart rate index, we found that a certain amount of BCSB pretreatment could significantly improve rat myocardial functions. The results of the physical and chemical testing showed that BCSB could reduce the extent of myocardial injury through detecting the leakage of lactate dehydrogenase(LDH) and creatine kinase isoenzyme(CK-MB) in coronary effluent. The results of western blotting revealed that treatment with BCSB inhibited the expression levels of Beclin-1, ratio of microtubule-associated protein 1 light chain 3(LC3) II to I, which may be attributed to the inhibition of autophagy. These studies provided pharmacology experimental basis for the comprehensive development and utilization of sea buckthorn to treat coronary heart disease.