| Human body will be damaged inevitably due to application of nuclear weapons and improper use of nuclear technology(nuclear power plant leakage).While wearing radiation suits can play a role in physical protection to body,but for cancer patients undergoing radiotherapy or accidental exposure to radiation damage,taking chemical radiation protectants is more important.Searching for natural antioxidants from natural products has become the focus of radiation protection research,because of side effects of many synthetic radiation protectants(such as amistine)are obvious,and many natural antioxidants(such as tea polyphenols)as a radiation protectant have a high efficiency and low toxicity characteristics.In this study,active tracer method was used to prepare/isolate the different activities radioprotective component of Pinus koraiensis bark(RCPKB).The optimal RCPKB was obtained by spleen cell injury protection model,and then gamma ray irradiated ICR mouse model was constructed.The protective effect of RCPKB on ?-ray induced damage was studied comprehensively in vitro and in vivo.In this study,three different polar solvents(95% ethanol,40% acetone and water)were used to prepare the extracts,respectively.95% ethanol extract of Pinus koraiensis was the best extract solvent after the antioxidant activity tracing and spleen lymphocyte proliferative activity analysis in vitro.The component of petroleum ether and dichloromethane was found to be cytotoxic,while the ethyl acetate and n-butanol components had spleen lymphocyte proliferation active through activity tracking from 95% ethanol extract.Then the in vitro antioxidant effects of different fraction products of n-butanol were evaluated by active tracing.It was found that NBEPKB-50 ME had the best anti-radiation activity,reduced the content of malondialdehyde,increase the activity of superoxide dismutase(SOD)and catalase in spleen cells after irradiation through active tracing of n-butanol extracts from 95% ethanol extracts of Pinus koraiensis.And protective effect of NBEPKB-50 ME on DNA damage in splenocytes was proved by single cell gel electrophoresis.39 compounds were identified by GC / MS based on active tracing results.The main components were aspartene(6.92%),dibutyl phthalate(6.40%),ethyl linolenate(6.38%),ethyl(E)-9-octadecenoate(12.70%),Ethyl stearate(3.46%),(1R)marinol(4.45%).The methylene chloride fraction was separated by a mixed solvent of petroleum ether and ethyl acetate in different proportions,and 6monomers were obtained by recrystallization,included E-ferulic acid hexacosyl ester(1),?-sitosterol(2)21-epi-serratenediol beta palmitate(3),ferulic acid(4),ursolic acid(5),Kaempfer Ol 3-O-beta-glucopyranosyl(1-4)--L-rhamnopyranosyl(1-6)alpha-beta-D-glucopyranoside-7-O-[2-O(E)-feruloyl]-beta-D-glucopyranoside(6).The content of flavonoids in the NBEPKB-50 ME fraction was 92.67%,while 77.49% was the procyanidins.The LC-MS results showed that some of procyanidins were B-type.Seven kinds of procyanidins belonging to two,four and pentamer types were deduced,and the relative contents of these procyanidins accounted for 57.34% of the total peak area.The ICR mouse injury model was established by 60 Co.It was found that each dose group of NBEPKB-50 ME dose group had anti-radiation effect.The proliferation of splenic lymphocytes was promoted by NBEPKB-50 ME in mice and rats,and the reduced glutathione(GSH)was increased,activity of myeloperoxidase(MPO)and level of malondialdehyde(MDA)were decreased significantly,while the activities of antioxidant enzymes [superoxide dismutase(SOD),catalase(CAT),glutathione peroxidase(GSH-Px)] were significantly increased in some organs of ICR mice.The results were proved that NBEPKB-50 ME activated the antioxidant enzymes effectively,inhibited oxidase system,inhibited lipid peroxidation,reduced cell membrane damage,reduced bone marrow micronucleus and chromosome aberration rate.NBEPKB-50 ME can restored hematopoietic system effectively and improve the body's immune function.These results also suggests that the NBEPKB-50 ME anti-radiation mechanism is associated with free radical scavenging,antioxidant enzyme levels,cell proliferation,glutathione levels,and lipid peroxidation.Five days after exposed to radiation,the expression of p-ERK,ERK Nrf2,AKt and HO-1 were down-regulated in intestine of radiation control group mice,however up-regulated in medium dose group.Compared with the irradiated control group,ERK down-regulated 26.12 + 2.73%,AKt decreased 94.69+ 9.31%,p-AKt up-regulated 78.08 + 7.62%,Nrf2 increased 32.55 + 3.08%,while HO-1 up-regulated 57.45 + 5.43%.These results show that NBEPKB-50 ME promoted the repair process of spleen after injury through up-regulated the expression of proteins in survival pathway.Therefore,the study of protective effects of NBEPKB-50 ME on oxidative stress induced by radiation has a certain theoretical significance and development prospect for Anti mutagenic and anti radiation effects of radiation exposure persons. |
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