| Background and objectivesBee pollen is known as the traditional natural nutritious food, which the bee collected from the honey plants within the blossom, and joined the special glands secretion(nectar and saliva), and contains rich nutrients. In addition to contains rich nutrients, bee pollen contain various bioactive substances, such as polysaccharides,active peptide, flavonoids, nucleoside, steroids, aldehyde, glycosides, unsaturated fatty acid, and so on. Bee pollen is the important resources in the new drug, health food research.Polysaccharide is an indispensable part of living organisms, and a kind of important biological macromolecules except protein and nucleic acid. Compared with proteins and nucleic acids, carbohydrates is the most ideal information carrier in the process of life, it can bearing most biological information whit the smallest structural,and play an important role in the process of cell recognition, cell signaling transduction, intercellular substance transportation, cell transformation and cell apoptosis. Research shows that polysaccharide from different bee pollen have different functions, such as immunity enhancement, antitumor, anti-inflammatory,antiviral, anti thrombus, reducing blood lipid, regulate blood sugar, antioxidant.Polysaccharide has become one of the forefront of new drug research and development.Polysaccharide is one of the main active components of bee pollen, It has many biological activities. The rape bee pollen is the most production in various of kinds bee pollen of china. Due to the influence of hydrology, geography, sampling time,gathering season, the contention of polysaccharide, monosaccharide composition,structure, biological activity and pharmacological are different in the bee pollen polysaccharide from different regions. At present, the researches of rape bee pollen are all about the rape bee pollen from Qinghai, Gansu, Xinjiang, Neimenggu, Sichuan,Hubei, et al, but the studies of Yunnan rape bee pollen polysaccharide has not to be reported yet. For comprehensive exploitation and utilization of Yunnan rape bee pollen new drug resources, 1 This study extracted the total polysaccharides from Yunnan rape bee pollen with the method of water extract and alcohol precipitation,measured the polysaccharide contention, and explored the optimal method of remove the protein from the Rape Bee pollen polysaccharide. 2 This study observed the actions of Yunnan rape bee pollen polysaccharide on anti-oxidation, anti-fatigue ability, the effect on immune function and the protective effect of acute liver injury.This study will provide experimental basis in further research on rape bee pollen polysaccharide.Method1. The study of the optimal method of removal the protein from the Rape Bee pollen polysaccharide. The total polysaccharides is extracted from Yunnan rape bee pollen with the method of water extract and alcohol precipitation. The seveag, trichloroacetic acid method(TAC) and enzyme methods were used to remove the protein from the Rape Bee pollen polysaccharide, and investigated optimal preparation conditions of each method.2. Research the actions of Yunnan rape bee pollen polysaccharide(RBPP) on anti-oxidation and anti-fatigue. Forty healthy KM mice were randomly divided into 4groups: high dose RBPP group, middle dose RBPP group, low dose RBPP group and the blank control group. The high,middle and low dose RBPP groups were given RBPP 300mg/kg.d, 150mg/kg.d, and 100mg/kg.d respectively by intragastric administration for 14 days.While the blank control group was given the same volume physiological saline. After the 12 hs of last administration, the related indicators were tested.3. Effects of RBPP on immune function. Fifty healthy KM mice(body weight 15-22g)were randomly divided into 5 groups with 10 mice in each group and half male and half female. The high, middle and low dose RBPP groups were given RBPP300mg/kg.d, 150mg/kg.d, and 100mg/kg.d respectively by intragastric administration for 7 days. in the second day and the fifth day, the RBPP groups and the modelcontrol group were given cyclophosphamide(CY) 80mg/kg by intraperitoneal injection to establish the model of immune suppression mice. While the blank control group was given the same volume physiological saline. After 24 h of the last treatment,using the conventional methods to test the related indicators.4. Study the protective effects of RBPP on acute liver injury in mice. Fifty healthy KM mice(body weight 15-22g) were randomly divided into 5 groups with 10 mice in each group and half male and half female. The five groups were: RBPP high, medium,low dose groups, blank control group and model control group. The RBPP groups were received 300 mg/ kg.d, 150 mg/ kg.d and 100 mg/ kg.d respectively by intragastric administration for 14 days. While the blank control group and the liver injury model control group were given same volume physiological saline. The liver injury model group and the RBPP high, medium, low dose groups were received0.1%CCL4 by intraperitoneal injection after 2h of the last administration. Then all mice were fasted for 12h(didn’t fasted water) and were executed to test the related indicators.Result1. The study of the optimal method of removal the protein from the Rape Bee pollen polysaccharide. The results showed that four times of removal protein is the best time of seveag method, 10% of TCA is the optimal concentration of TCA method, the optimum technological conditions of enzyme method were enzyme usage 2%,temperature at 50℃, deproteinization time 1h and p H value 6.0.2. Research the actions of Yunnan rape bee pollen polysaccharide(RBPP) on anti-oxidation and anti-fatigue. The results showed that compared with blank control group, the high(300mg/kg), middle(150mg/kg) and low(100mg/kg) doses of RBPP could remarkably lengthen the load swimming time, increase muscle and hepatic glycogen content. On the other hand, RBPP 300mg/kg dose could significantly increase the activity of serum LDH. In addition, the results also showed that with the dosage of 300mg/kg, 150mg/kg and 100mg/kg, RBPP could obviously increase the activity of SOD in liver tissue and decrease the content of MDA in liver tissue. Theresults suggest that RBPP may has the effects of anti-oxidation and anti-fatigue.3. Effects of RBPP on immune function. The results showed that compared with blank control group, the model group’s viscera index, the level of white blood cells of mice obviously decrease. It proved the model was established. Compared with model mice, the high, middle and low dose of RBPP have the antagonistic effects on the immunosuppressive action caused by cyclophosphamid. Compared with immune suppression mice group, RBPP could significantly increase the viscera index, the level of white blood cells of the mice. Meanwhile, with the dosage of 300mg/kg,150mg/kg and 100mg/kg, RBPP could obviously promote the function of macrophage cell in abdominal cavity and the formation of serum hemolysin.4. Study the protective effects of RBPP on acute liver injury in mice. The results showed that compared with blank control group, the liver injury model group’s activates of serum AST and ALT increased significantly, and the liver tissue MDA content increased and the liver SOD activity decreased significantly of the model mice. It proved the liver injury model was established. Compared with model group,the study found that the activity of serum ALT of RBPP 300 mg/kg, 150 mg/kg and100 mg/kg groups reduced significantly, and the activity of serum AST of RBPP 150mg/kg and 300 mg/kg groups reduced remarkably. In addition, the liver tissue MDA content reduced and the liver tissue SOD activity were increased in RBPP 300 mg/kg,150 mg/kg and 100 mg/kg groups. On the other hand, the results of histpoathollgical examination of liver showed that the RBPP 300 mg/kg and 150 mg/kg were able to make the swelling, hyperemia, necrosis of liver cells significantly reduced, but the100mg/kg dose was no obviously improving effect.Conclusion1.The effect of deproteinization with different methods were evaluated in terms of the ratio of protein removal and ratio of polysaccharide loss. The results showed that the optimal method of Yunnan rape bee pollen polysaccharide was the enzymatic method,its ratio of protein removal was 85.89% and the ratio of polysaccharide loss was25.34%. 2. RBPP may has the effects of anti-oxidation and anti-fatigue. 3. RBPP hasgood effect of improvement on immune function on immune suppression mice induced by cyclophosphamide. 4. RBPP has good effect of protect the liver injury. |
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