| Transfer factor is biological multicomponents with immune activities, extracted from animal spleen, containing peptides, nucleic acids and other active substances. Currently, Transfer factor is mainly extracted by physical methods, such as in the way of freeze-thaw, dialysis or any other way. They are mainly from pig spleen, cow spleen or chicken spleen but spotted deer spleen. From ancient times, spotted deer has been precious, for its medicinal value and healthcare functions of their organs. Spotted deer’s organs have been all developed except from the spleen. It is very wasteful to abandon the deer spleen. Shuangyang Deer town, a deer breeding base is near to our school, so we can get healthy deer spleen. For these reasons, we developed methods to extract activities from deer spleen, optimized the extraction method by enzymatic hydrolysis method, and developed oral liquid and researched its activities on mouse’s behavior and immune competence. The mainly study parts are following:1. Study the extraction process, on the basis of the content of polypeptide and ribose, as the indicator. After researching information, we determined to use papaya protease and 5’-phosphdies-terase to extract deer spleen. On the basis of the theory of one factor optimization, we chose significant factors from the ratio of liquid to solid, reaction temperature, enzyme hydrolysis period, pH conditions. By response surface design, optimize the extraction process. The optimum extraction was as follows:the ratio of liquid to solid(mL/g)for 3.97:1, Grind in a colloid mill for 2-5 min directly, adjust pH to 5.08, add 025% Papain and keep 66.08℃ for 30min, following adding 1% 5’-phosphdies-terase kept for 2 hours, then keep 80℃ for 30 min. After cooling down, the sample is centrifuged at 8000 g for 10 min and supernatant is ultrafiltrated in the 8KD ultrafiltration membrane conditions.2. Study the preparation process for oral deer spleen liquid. Deer spleen extraction is diluted 10 times as oral deer spleen liquid. We chose natural sweeteners, citric acid and preservatives, ensuring oral deer spleen liquid is healthy. According to the organoleptic evaluation of 20 people, the formula of oral deer spleen liquid was as follows:maltitol 6% and critic acid 0.8%. vitamin C was chosen as an anti-corrosion agent because it preserved the liquid and made the efficacious contents effective.3. Study the quality standards and stability of oral deer spleen liquid. The pH=5.0-7.0; polypeptide concentration above 1.0mg/mL; ribose concentration above 100ug/mL. Microbial limits and heavy metal concentration are less than the provisions restricted by "healthy food general standard GB 16740-1997 ". And the indicators of oral deer spleen liquid were no significant changes in the long-term stability test. But some changes happened in the light accelerated test and high temperature accelerated test. The oral liquid should preserve in the dark and keep it low temperature.4. Study the activity of oral deer spleen liquid on rats’behavior and immunology activities. The experimental results showed that there was no significant in each dose group and blank group in the dependent experiment and rotarod test. In the weight loading swimming experiment,the high dose group and low dose group could improve the antifatigue activity compared to control group,but not significant. The result of running test showed that the high-dose group could improve the antifatigue activity. In the immunity test, the high-dose group could significantly increased thymus index, but spleen index has no significance. Some dose group significantly increased IL-2,IL-6,IL-10, INF-y level in the serum, compared to the control group. However, there was no significance in TNF-α and IL-12 level in the serum between dose group and control group. The results showed that oral deer spleen had the potential for improving fatigue effect and enhancing immunological activity to a certain extent. |
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