Preparation And Protective Effect On ⁶⁰Co-γ Radiation Damage Of Chitosan Loaded With Pinus Koraiensis Polyphenols Micropheres

Pine polyphenol is a kind of natural product derived from Pinus koraiensis, which had a good anti-radiation function. But because of its unique polyhydroxy molecular structures, which is vulnerable to light, high temperature, oxygen, extreme p H and other harmful environmental factors, which could made the functional composition degradate and even turned into toxic byproducts. Microsphere as a new technology of drug embedding, which had great potential in targeting and sustained drug delivery. Chitosan as a kind of natural carrier material, which not only had great anti-radiation function, but also played a protective role and could sustained release core drug. In this study, chitosan as a kind of drug carriers, embedding pine polyphenols, prepared an anti-radiation drug system, which not only solved the problem of poor stability of polyphenols, but also enhanced the effectivity of the drug system, which guaranteed this composite drug delivery system safe, non-toxic and high efficacy. This is a bright new spot in this field, which has a strong practical significance for the research of anti-radiation drugs.In this study we used emulsification cross-linking technology, chitosan was wall material, pine polyphenols was core material, glutaraldehyde was cross-linking agent, and explore the best pine polyphenols microspheres preparation conditions. Then chosed the factor of chitosan concentration, crosslinking time, crosslinking agent amount, stirring speed, crosslinking temperature and emulsifying time as the single factor respectively, set the embeding rate and drug loading rate as response, detect the influence of various factors to the polyphenol microsphere embeding rate and drug loading rate, and observed the microspheres shape and appearance through electron microscope photograph. Based on these, selected four obvious influence factors, Design expert7.0.0 designed 29 group of tests, whose result showed that the best preparation key parameters of polyphenols microspheres were as following: chitosan concentration was 2%, crosslinking agent amount was 7.19 ml, stirring speed was 660.98 r/min, crosslinking temperature was 41.18 ℃, emulsifying time was 198.65 min, crosslinking time was 80 min;After optimized, the morphology of microspheres observed by scanning electron microscopy(SEM) was good, the rate of qualified balls was high, the size of the spherical distribution is uniform; the core-shell structure is obviously; the microspheres particle size is 3um measured by laser granulometer; the embeding rate and drug loading rate were 73.57% and 7.47% respectively. Through simulated human gastric and intestinal conditions, the microsphere release situation was: microsphere under the condition of the acidity stomach is stable, which could transit to intestinal smoothly, and released stably, whose slow release performance proved that the microspheres is successful.This project used KM mouse, which weight 19-21 g. And devided into four groups: the normal control group, radiation model groups, pine polyphenol groups(low, middle and high dose), pine polyphenol microsphere groups(low, middle and high dose), each group had half male and half female. After fed 14 days, we used 60 Co gamma ray as radiation source, 6.0 Gy doses, radiated the each group mice except the normal control group, after 24 h executed, testing all the indexes, the result showed that: 1) compared with normal control group, radiation model group, pine polyphenol group and pine polyphenols microsphere group female and male mice immune organ have different degree of injury, serum SOD activity reduced and MDA level increased, the bone marrow DNA content decreased, the spleen lymphocyte proliferation ability also weakened, and monocyte-macrophage phagocytosis ability had reduced, bone marrow cell micronucleus number increased, illustrated that 60 Co gamma ray damaged the body immune system in mice; 2) compared with model group, pine polyphenol group and pine polyphenols microsphere group female and male mice immune organ index increased with the drug dosage increased, serum SOD activity increased with the increasing of drug dosage, the MDA level reduced with the increasing of drug dosage, bone marrow DNA content, spleen lymphocyte proliferation ability and monocyte-macrophage phagocytosis increased with the increasing of the drug dose, at the same time, the increased dosage could make the bone marrow micronucleus decreased, which showed that pine polyphenols and pine polyphenols microspheres could protect the body immune system from the 60 Co gamma rays damage to mice; 3) Compared with low, medium and high dosage three dose groups of pine polyphenols, low, middle and high dosage group of pine polyphenols microspheres male and female mice immune organ index improved in different degree, serum SOD activity are much higher and MDA level reducd, bone marrow DNA content, the spleen lymphocyte proliferation ability and monocyte-macrophage phagocytosis ability had been enhanced, at the same time, the bone marrow cells micronucleus number reduced, which showed that the protect effect of pine polyphenols microspheres to 60 Co gamma ray radiation is better than pine polyphenols monomer.Various index showed that pine polyphenols and pine polyphenols microspheres achieved the role of protect mice from radiation injury mainly through the following ways: Firstly, removed the harmful free radicals in body, which is producted by radiation stress; Secondly, improved the activity of endogenous antioxidant enzymes and enhance the body’s antioxidant capacity, immunity and anti-stress ability; Thirdly, enhanced the body immunity, promote the immune cell proliferation and reduced the damage.In addition, we found that in the detected indexes of animal experiment, the difference between the result of female mice and male mice is significant: the immune organ index, serum SOD and MDA, spleen lymphocyte proliferation ability, mononuclear macrophage phagocytosis test and marrow DNA content, except the damage brougut by stress itself, female mice showed a stronger ability to resist the radiation damage, while in bone marrow micronucleus, male mice had the strong superiority. This suggested that different gender in fighting the same stress environment, the body would appear differentiation, it’s not only because of different gender, peculiar to the different hormone and its receptor, even some important immune cells in different gender polarization performance are also related, and the key differences between protein expression needed further research.