The Extraction And Purification And Injection Preparation Of Polysaccharide From Agaricus Blazei Murill

Agaricus blazei Murill (ABM) is a kind of new edible and medical fungus in recent years. This mushroom has been attracting attention as a source of immunomodulators, anticarcinogens, antimutigens and antibacterial substances. It contains rich biological active components and exhibits diverse bioactivities so that it is exactly important to extract and purify its active components. The methods of extraction and purification polysaccharide from ABM have been reported more at home and abroad, but there are a few of reports on tonic or drug made from polysaccharide of ABM. In this dissertation, injection made from polysaccharide of ABM is developed, which will provide evidences for applying on clinical research as an adjuvant cancer chemotherapy drugs.To make the polysaccharide injection from ABM safety, we chose an economical, safe and efficient process to isolate, purify, concentrate and dry according to Pharmacopoeia of the People's Republic of China and the relevant provisions of extraction and purification of TCM, natural medicine. First, removing protein by Sevage method. According to literature, it had best efficiency when polysaccharide liquid 2mg·mL^-1 was treated with chloroform: n-butanol (3:1) and repeated for 3 times. Second, moving pigment with H₂O₂. In order to shorten the time and prevent the broken of structure of polysaccharide, the experiment was operated at 37℃. The following were dialyzation, concentration(60℃) and freeze-drying. Then, a prepared product called AB which was brown and powder was obtained. It consisted of 85.4% polysaccharide and 14.2% protein. AB was derided into three fractions by DEAE-cellulose ion-exchange chromatography with NaCl in 0, 0.1, 0.3mol.L^-1. AB-â… was neutral polysaccharide, AB-â…¡and AB-â…¢were acidic ones. AB-â… , AB-â…¡and AB-â…¢were further fractionated by Sepharose CL-4B gel filtration. AB-â… showed single and symmetrical peak, a light yellow powder was obtained after dialyzation and freeze-drying. AB-â…¡and AB-â…¢were devided into two parts, collected the main peak, we gained AB-â…¡-b which was yellow powder and AB-â…¢-b which was brown powder. AB-â… , AB-â…¡-b and AB-â…¢-b were identified to be polysaccharide of uniform molecular weight and consisted of 91.8%, 90.4%, 89.2% polysaccharide and 7.1%, 8.8%, 9.3% protein, respectively.To observe the immune pharmacological effct of AB, AB-â… , AB-â…¡-b and AB-â…¢-b in normal mice and mice with immunosuppression, we treated mice with cyclophosphamide (CTX) and investigated the difference of immunological function around polysaccharide-treated. The examinations were designed by immunological organ weight test, hemogram test, phagocytic function of peritoneal macrophage test, hemolysin production test, E-rosettes formation test and lymphocytes proliferation test to investigate immunological function from nonspecific immunity, humoral immunity and cell imminity. The results of tests showed four kinds of polysaccharide had an antagonistic effect on the damage induced by CTX in mice, some even exceeded the saline-treated level; polysaccharide-treated in normal mice were significantly increased compared with saline-treated(P＜0.01) except lymphocytes proliferation test; AB-â…¢-b-treated group was significantly difference compared with AB, AB-â… and AB-â…¡-b -treated group in immunological organ weight test and hemolysin production test(P＜0.01), that indicated AB-â…¢-b had the best immunoregulation effect.On the basis of the study above, we designed the technical process on polysaccharide injection of ABM and obtained injection products. According to Pharmacopoeia of the People's Republic of China and the relevant technical provisions of TCM, natural medicine, injection intermediates were tested by quality and safety tests. The results of acute toxicity test, local muscle stimulation test, sensitivity test, hemolysis test proved the injection of polysaccharide extracted from ABM was safe and can be used via intramuscular injection and intravenous injection.Above all, polysaccharide of ABM is an immunoregulator and has an antagonistic effect on the damage induced by cyclophosphamide. It can be accompanied with chemotherapy drugs and increase efficacy. Injection of polysaccharide from ABM may be developed into a new adjuvant cancer chemotherapy drugs to benefit mankind.