Studies On Separation, Purification And Biological Activities Of Polysaccharides From Platycodonis Radix And Schisandrae Chinensis Fructus

Platycodonis Radix and Schisandrae Chinensis Fructus are famous medicinal plants inChina. Platycodonis Radix has been used in traditional oriental medicine as an expectorantand antitussive and used to treat coughs, colds, upper respiratory infections, sore throats,tonsillitis, and chest congestion. According to reports, the the polysaccharides of Platycodongrandiflora has immunomodulatory and anti-tumor effects. Schisandrae Chinensis Fructushas the function of replenishing and promoting production of body fluid, tonifying thekidney and relieving mental strain. It is reported that the polysaccharides of Schisandra hasanti-fatigue, hypoxia, lowering blood sugar and immunomodulatory effects. So far thestudies on Platycodonis Radix and Schisandrae Chinensis Fructus have been focused onsaponin and lignans. So we need to systematic study on the polysaccharides of them.Thesubject studied on extraction, separation, purification and biological activities ofpolysaccharides from Platycodon Grandiflorum and Schisandra Chinensis.This subject is divided into six parts. The first part is to set up and optimize the ways ofextracted crude polysaccharides from Platycodonis Radix and Schisandrae ChinensisFructus.The second part is the purification of them. This part also studies on the properties ofthem. The third part is that polysaccharides from Platycodonis Radix (PRPS) acts on the ratasthma model. The forth part is that observe the effect of PRPS on the HASMCsproliferation model which is sensitized by endothelin-1(ET-1). In the fifth part,polysaccharides from Schisandrae Chinensis Fructus (ASPS) acts on CCl4acute hepaticinjury mice of model. The last part is to abserve antioxidation activity of them.The first part of the experiment used the hot reflux method to extract the the crudepolysaccharides from Platycodonis Radix, and used room-temperature immersion method ofalkaline to extract crude polysaccharide from Schisandrae Chinensis Fructus. In order tofurther optimize the extraction process of the polysaccharides, the experimental designedtwo orthogonal experiments of L9(34). The results showed that the yield of crudepolysaccharides from Platycodonis Radix was12.66%at the conditions of85%ethanolrecipitation,1:10solid-liquid ratio and2h for3times for extraction.The yield of crudepolysaccharides from Schisandrae Chinensis Fructus was1.18%at the conditions of90% ethanol recipitation,1:6solid-liquid ratio,2h for extraction and the concentration of NaOHwas2M. In the two extraction process, the energy consumption was low, the conditions weremild. They were efficient extraction methods. Polysaccharides of Platycodonis Radix andSchisandrae Chinensis Fructus have a variety of pharmacological activities. The optimization of theextraction process laid the foundation for the further development and application of polysaccharides.And it provided a rich source of polysaccharides to separate and purify.In the second part, the crude polysaccharides were purified by sevag deprotained. Theseparation and purification of the polysaccharides were used the means of columnchromatography. In the end, we gained five polysaccharides called PRPS-a, PRPS-b,PRPS-c, ASPS-a-1, ASPS-b-2and ASPS-b-3. They were isolated from Platycodonis Radixand Schisandrae Chinensis Fructus for the first time. Their molecular weights were2567Da,439746Da,2839Da,103300Da,61600Da,34700Da tested by HPSEC. The uronic acidcontent of ASPS-a-1, ASPS-b-2and ASPS-b-3were7.8%,39.4%and43.5%. There was nouronic acid in PRPS-a, PRPS-b, PRPS-c. The monosaccharide composition of them wasanalyzed by GC. PRPS-a was composed of fructose, glucose and galactose with a molarpercentage ratio of41:35:24. PRPS-b was composed of arabinose, fructose and glucose witha molar percentage ratio of35:43:22. PRPS-c was composed of arabinose, fructose andgalactose with a molar percentage ratio of28:39:33. The monosaccharide composition ofASPS-a-1, ASPS-b-2and ASPS-b-3were Ara:Xyl:Glu:GalA=22:15:51:6,Ara:Xyl:Glu:GalA=14:18:33:35,Ara:Xyl:Glu:GalA=16:9:32:41.In the third and forth parts, anti-asthma activity of PRPS was tested in vivo and in vitro.In vitro experiment, the rat asthmatic model was established by sensitizing and challengingSD rats with aerosolized ovalbumin (OVA) and the intervention of PRPS was performed.The changes of the levels of IL-4, IFN-γ, EOS in broncho alveolar lavage fluid (BALF) andserum were observed. The expression level of Eotaxin1and ICAM-1were detected. Theresults showed that PRPS can significantly inhibit the expression of Eotaxin1, ICAM-1inasthmatic rats and reduced the number of EOS in blood and airway. In addition, PRPS cansignificantly reduce the level of IL-4and increase the level of IFN-γ in blood and serum inrat asthmatic model. This proved that PRPS can regulate the disorder in the balance betweenTh1and Th2cytokines which makes it has anti-asthma activity. In vivo experiment, the cellsproliferating model of human ASMCs were established by ET-1and the intervention ofPRPS-a, PRPS-b and PRPS-c was performed. MTT method was used to detect inhibition toHASMCs. The experimental results showed that PRPS-b and PRPS-c could inhibit HASMCs proliferation obviously. The inhibitory activity of PRPS-a wasn't evident. Theactivity difference depends on the monosaccharide composition of PRPS-a, PRPS-b andPRPS-c. The polysaccharide has powerful inhibitory activity which containing arabinose andfructose. This part ultimately proved that PRPS has anti-asthma activity which consistentwith the medicinal efficacy of relieving cough and asthma of Platycodonis Radix. So thepolysaccharides are the active site of Platycodonis Radix.In the last two parts, antihepatitis and antioxidant activity of alkaline S. chinensispolysaccharides (ASPS) were studied. The model of acute liver injury was induced by CCl4and the intervention of ASPS was performed. The activities of T-AOC, NO and SOD, thelevel of Alb and MDA, as well as the pathological changes of the liver were determined. Theresults showed that different doses of ASPS can reduce the weight of liver tissue, the level ofMDA and the activities of NO. It can also increase the level of Alb and the activities ofT-AOC, SOD. The pathological sections of liver showed that the swelling of liver cells, fattychange and inflammatory cells infiltration were alleviated by ASPS. These results suggestedthat ASPS can role in the resistance to the liver injury. The free radical scavenging and metalchelate activity of ASPS-a-1, ASPS-b-2and ASPS-b-3were detected. The results provedthat they have strong antioxidant activity. In addition, the antioxidant activity of them relatedto the content of furfural acid. These two chapters determined that ASPS can protect the liverby inhibiting free radicals generation and radical scavenging from injury caused by CCl4.In conclusion, this project took polysaccharides of Platycodonis Radix and SchisandraeChinensis Fructus as the research object. It determined their extraction and purificationprocess, studied on their physicochemical properties and pharmacological activity. Itconfirmed the PRPS, PRPS-a, PRPS-b and PRPS-c have antiasthmatic activity. It alsoproved that ASPS, ASPS-a-1, ASPS-b-2and ASPS-b-3have antihepatitis activity andantioxidant activity.These results provide the data of polysaccharides from PlatycodonisRadix and Schisandrae Chinensis Fructus to establish the material basis of traditionalChinese medicine efficacy and material resources library. The object provides a theoreticalbasis for the polysaccharides becoming. effective drugs against the disease, the Schisandrapolysaccharide as a hepatoprotective drug and antioxidant provides a theoretical basis.