| As a renewable boilogical resource,deer antler base has been used since 2000 years ago in Traditional Chinese Medicine.Large yied and easy to be obtained are its advantages.Moreover,the antitumor activity of deer antler base is still focused by many researchers.However,the antitumor activity site is unclear,the mechanism of antitumor is unknown,and due to its high degree of ossification and hard texture,it should not be used directly.Base on these reasons above,the utilization and development of deer antler base was greatly limited.And,it was usually decocted by the water,which was the only method.Therefore,it is imperative to study the proccesed method and the antitumor effect of deer antler base.In this experiment,we extracted the deer antler base with acid,alkaline,salt,alcohol and water to obtain five kinds of extraction site of deer antler base and optimized the conditions of extraction.Then,the pre-column derivatization RP-HPLC method was used to determine the content and coposition of 21 kinds of amino acid and explore the basis of its antitumor effect.Finally,the H22 tumor bearing mice was selected to compare the anti-liver cancer activity of five kinds of extraction of deer antler base,screen the active site of antitumor and explore mechanism according to the immunological theory.The results were:1.The optimal conditions of extraction site of acid solution,alkaline solution,salt solution,alcohol solution and water were: temperature 90?,time 130 min,ratio of solid to liquid 1:25;temperature 90?,time 130 min,ratio of solid to liquid 1:20;temperature 80?,time 130 min,ratio of solid to liquid 1:25;temperature 70?,time 150 min,ratio of solid to liquid 1:25;temperature 100?,time 130 min,ratio of solid to liquid 1:26.The order of extraction rates of the five kinds of extraction were in the order: extraction of acid solution?extraction of alkaline solution?extraction of water?extraction of salt solution?extraction of alcohol,and there was no significant difference between extraction of salt solution and extraction of alcohol solution(P>0.05).2.In the five kinds of extraction and powdered of deer antler base,the content of medical amino acid account for total amino acid was more than 60%.In the extraction of alkaline,seven kinds of amino acid(Tyr,Leu,Lys,Asp,Phe,Met and Glu)were the highest(P<0.05).In the extraction of water,five kinds of amino acid(Asp,Gly,Met,Glu and Arg)were the highest(P<0.05).In the extraction of alcohol,five kinds of amino acid(Gly,Arg,Phe,Met and Glu)were the highest(P<0.05).In the extraction of alkaline solution,the E/T value and E/N value were higher than the the collagen of sea cucumber,donkey-hide gelatin.The content of total amino acid in five kinds of extraction was in the follow order: extraction of water?extraction of alkaline solution?extraction of alcohol?extraction of acid solution?extraction of salt solution,and there were no significant difference among extraction of water,extraction of alkaline solution and extraction of alcohol(P>0.05).3.Compared with the model group,the powdered deer antler base and its five kinds of extraction can significantly inhibit tumor growth and promote the apoptosis of tumor cell(P<0.05);protect the spleen tissue structure,induce tumor necrosis;improve immune organ index,stimulation index of splenic lymphocytes,the levels of IL-6,IL-12,TNF-? and IFN-? in the serum(P<0.05),the CD4+/CD8+ value of the spleen and peripheral blood(P<0.05);and decrease the the level of immunosuppressive cytokine TGF-?(P<0.05),increase the level of cellular immunity of H22 tumor-bearing mice.These also can increase the levels of IgE,IgG and IgM in the serum of H22 tumor-bearing mice(P<0.05)and increse the level of humoral immunity of H22 tumor-bearing mice.And the anti-tumor actity of extraction of alkali solution was the best. |
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