灵芝菌丝体多糖提取工艺优化及其对慢性酒精肝损伤的保护作用

Optimization of Extraction Technology of Polysaccharides from Ganoderma lingzhi Mycelium and Its Protective Effect on Alcoholic Liver Injury

本研究以灵芝菌丝体为材料,采用水提醇沉法提取灵芝菌丝体多糖,在单因素实验基础上,结合响应面分析法优化灵芝菌丝体多糖提取工艺,通过Sevage法及透析法去除蛋白和小分子,获得初步纯化灵芝菌丝体多糖(SGP)。动物模型:将60只小鼠随机分成空白对照组、模型组、阳性对照组和给药低、中、高组(125、250、500 mg/kg·bw),除空白组外,各组按照0.1 mL/10 g·bw剂量灌胃56°北京红星二锅头,建立慢性酒精肝损伤模型。第15 d开始,各给药组造模4 h后,分别灌胃水飞蓟宾(200 mg/kg·bw)和不同剂量的SGP溶液,连续2周后,解剖。采用试剂盒法测定小鼠血清及肝脏组织中谷草转氨酶(AST)、谷丙转氨酶(ALT)、过氧化氢酶(CAT)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)、丙二醛(MDA)、总胆固醇(TC)、甘油三酯(TG)和白介素-6(IL-6)、白介素-1β(IL-1β)和肿瘤坏死因子α(TNF-α)水平,并观察小鼠的肝脏病理切片。结果表明:灵芝菌丝体多糖最优提取条件为提取温度89℃、提取时间2.5 h和液料比85:1(m L/g),在此条件下,多糖得率为3.44%。动物实验结果表明,与模型组相比,SGP中、高剂量组小鼠肝脏指数极显著下降(P<0.01);SGP中、高剂量组肝脏和血清中的CAT、SOD和GSH-PX活力显著上升(P<0.05),SGP高剂量组ALT、AST活力及MDA、TC、TG、IL-6、IL-1β、TNF-α含量极显著下降(P<0.01);病理组织切片结果显示,SGP可明显改善肝脏受损情况。说明,SGP可以通过改善小鼠肝脏氧化应激水平,脂质代谢水平并降低小鼠肝脏细胞炎症因子含量,进而对慢性酒精性肝损伤小鼠发挥保护作用。

In this study,Ganoderma lingzhi mycelium powder was used as material to extract polysaccharides from Ganoderma lingzhi mycelium by water extraction and alcohol deposition method.On the basis of single factor experiment,response surface methodology was combined to optimize the extraction process of polysaccharides from Ganoderma lingzhi mycelium.Proteins and small molecules were removed by Sevage method and dialysis method,then,the preliminary purification of Ganoderma lingzhi mycelium polysaccharide(SGP)was obtained.Animal model:60 mice were randomly divided into blank control group,model group,positive control group and low,medium and high administration groups(125,250 and 500 mg/kg·bw).Except for blank group,each group was given 56°Beijing Red Star Erguotou by gavage at a dose of 0.1 mL/10 g·bw to establish a chronic alcohol liver injury model.Starting from the 15th day,4 h after modeling,each administration group was given intragastric administration of silybin(200 mg/kg·bw)and different doses of SGP solution,respectively,and dissected after 2 consecutive weeks.The levels of aspartate amino transferase(AST),aspartate transaminase(ALT),catalase(CAT),superoxide dismutase(SOD),glutathione peroxidase(GSH-PX),malondialdehyde(MDA),total cholesterol(TC),triglyceride(TG),interleukin-6(IL-6),interleukin-1β(IL-1β),and tumor necrosis factor-α(TNF-α)in serum and liver tissues of mice were determined by kit method,and the liver pathological sections of mice were observed.The results showed that the optimal extraction conditions of polysaccharides from Ganoderma lingzhi mycelium were 89℃,2.5 h and liquid/solid ratio 85:1 mL/g.Under these conditions,the yield of polysaccharides was 3.44%.Animal experiment results showed that the liver index of mice in SGP medium and high dose groups was significantly decreased(P<0.01).The activities of CAT,SOD and GSH-PX in liver and serum of SGP medium and high dose groups were significantly increased(P<0.05),the activities of ALT and AST and the contents of MDA,TC,TG,IL-6,IL-1βand TNF-αwere significantly decreased in SGP high-dose group(P<0.01).The results of pathological tissue sections showed that SGP could significantly improve liver damage.These results indicate that SGP can play a protective role in mice with chronic alcoholic liver injury by improving the oxidative stress level,lipid metabolism level and reducing the content of inflammatory factors in mouse liver cells.