Isolation, Characterization Of Polysaccharides From Ganoderma Lucidum Fruiting Bodies And Correlation Aanlysis Of The Chromatographic Fingerprints-immunocompetence

Ganoderma lucidum is the treasure of traditional chinese medicine, and has thereputation for functional food. Because of its prominent biological effects such asimmunocompetence, its research and development has always been a hit point. Recently,Ganoderma lucidum products industry has gained development rapidly, however Ganodermalucidum has been intermingled by some bad sellers so that it’s difficult to control and identify.The main reason is that the composition of Ganoderma lucidum fruiting bodies, spores andmycelium is very complicated, while the detection method is still so weak that there is noeffective system for quality control and quality supervision of Ganoderma lucidum products.Ganoderma lucidum polysaccharides is the main active ingredient of Ganoderma lucidum,Chinese pharmacopoeia also evaluates the quality of Ganoderma lucidum by the content ofGanoderma lucidum polysaccharides. In order to establish effective method of detecting thequality of Ganoderma lucidum fruiting bodies and their products, this paper was supposed tostudy the profile-effect relationship between high performance size exclusion chromatography,partial hydrolysis-precoloumn derivatization reversed-phase high performance liquidchromatography(RP-HPLC) and immune activity.Taking Ganoderma lucidum fruiting body as the experimental material, extracted it bythe way of water bath reflux, and decolorized it with resin. After that, using Sevag method todeprotein it and passed it through DEAE Sepharose Fast Flow ion exchange coloumnchromatography coloumn to isolate the Ganoderma lucidum polysaccharide. Finally, we gotfive kinds of Ganoderma lucidum polysaccharide fractions which were called: GLPJD1,GLPJD2, GLPLD1, GLPLD2, GLPLD3. HPSEC results indicated that the purity of fiveGanoderma lucidum polysaccharide fractions can reach up to90％. Moreover, their relativemolecular mass were respectively4.41x106,4.70x106,7.86x103,7.38x103,1.77x104Da.Infrared analysis results suggested that the anomeric carbon of Ganoderma lucidum fruitingbody polysaccharides were beta type, the characteristic absorption peak at1750cm-1alsoimplied that Ganoderma lucidum fruiting body polysaccharides might have uronic acid. Inaddition, monosaccharide composition of five kinds of Ganoderma lucidum polysaccharideswere determined after complete acid hydrolysis, it was found that five polysaccharides didcontain a small amount of uronic acid. SEC-LLS analysis results showed that the the absolutemolecular weight of five polysaccharide fractions were repectively12.18106、7.086106、0.012106、0.017106、0.019106Da. One thing that hasn’t changed was that the purity ofthis five polysaccharides fractions were consistently very high.With the purpose of better characterizing profile-effect relationship between Ganodermalucidum polysaccharides and their immunomodulatory effects, we choosed the experiment ofdetecting polysaccharides’ normal mouse macrophages promoting ratios. The result indicatedthat the activities of promoting the macrophage proliferation ratios were different amongGanoderma lucidum polysaccharides collected from different sources or planted withdifferent methods. We can tell from the results that the14th Ganoderma lucidumpolysaccharides collected from Jiangxi province and the7th as well as13th Ganoderma lucidum polysaccharides planted in Zhejiang province standed out in their mousemacrophages promoting activities. The9th Ganoderma lucidum polysaccharides extractedfrom Liaoning province Ganoderma lucidum fruiting body also had better immunostiulatingactivities. Additionly, by comparing five polysaccharide fractions’ immunomodulatory effect,it wasn’t hard to tell that the macrophage promoting activities of GLPJD1,GLPJD2weremuch higher than GLPLD1、GLPLD2、GLPD3.This paper analyzed the HPSEC and PMP RP-HPLC fingerprint chromatography of29kinds of polysaccharides extracted form Ganoderma lucidum fruiting bodies and theirimmunocompetence on promoting normal mice macrophage proliferation activity. Taking onthe preliminary exploration as well as establishing the profile-effect relationship between twodifferent kinds of fingerprint chromatography and Ganoderma lucidum polysaccharidesimmunoregulating activity.HPSEC profile-effect correlation analysis results shows that the fraction with Mw <5000Da was basically not immunostimulating activity related. However, Mw＞1×106Da and Mw1×105～5×105Da fractions were positively related with normal mouse macrophageproliferation activity. On the other hand, PMP-RP HPLC profile-effect correlation resultsdemonstrated that its positive correlation peak was mainly coming from characteristic peak19and the GlcUA peak. These two positive correlation peaks and Mw＞1×106Da as well as Mw1×105～5×105Da positive correlation peaks can be the important part of the main indicatorswhich were used as a judgement of Ganoderma lucidum fruiting bodies and their productsquality. Furthermore, it serves as study reference for further study and proof.