基于HPLC-Q-Orbitrap-MS槐白皮水提物的化学成分测定及其对溃疡性结肠炎的治疗作用研究

Study on Determination of Chemical Constituents in Sophora Japonica Water Extract by HPLC-Q-Orbitrap-MS and Its Therapeutic Effect on Ulcerative Colitis

关键词:  槐白皮  高效液相色谱-四级杆/静电场轨道阱高分辨质谱  溃疡性结肠炎  NF-κB P65  TNF-α  IL-1β

Key words:  Sophora japonica  HPLC-Q-Orbitrap-MS  ulcerative colitis  NF-κB P65  TNF-α  IL-1β

摘要:

目的 分析槐白皮水提物的化学成分，探讨其对溃疡性结肠炎的治疗作用及机制。方法 采用高效液相色谱-四级杆/静电场轨道阱高分辨质谱（HPLC-Q-Orbitrap-MS）技术对槐白皮水提物的化学成分进行定性鉴定，与数据库（mzCloud、mzVault、ChemSpider）检索比对。42只♂ BALB/c小鼠随机分为正常组、模型组、阳性对照组和槐白皮水提物低、中、高剂量组6组，每组7只。正常组饮用蒸馏水，其他组均饮用4%葡聚糖硫酸钠溶液，采用灌胃给药（10 mL·kg^–1），连续给药11 d。给药期间观察小鼠体征变化，进行疾病活动指数（disease activity index，DAI）评估，给药结束后HE染色观察小鼠结肠组织病理学改变，ELISA测定小鼠血清中炎症因子TNF-α、IL-1β含量，免疫组化法检测结肠组织中NF-κB P65的表达。结果 通过mzCloud数据库对提取物中未知化合物进行分析和鉴定，在数据库中共匹配到323个化合物，其中匹配度>90%的化合物有22个。与模型组相比，槐白皮水提物中、高剂量组能明显抑制葡聚糖硫酸钠诱导的小鼠结肠缩短，显著降低DAI指数，明显降低血清中TNF-α、IL-1β含量和结肠组织中NF-κB P65的表达。结论 槐白皮水提取物可通过抑制肠道炎症反应，减轻葡聚糖硫酸钠诱导的BALB/c小鼠溃疡性结肠炎损伤。

Abstract:

OBJECTIVE To analyze the chemical compositions of Sophora japonica water extract, and to explore its therapeutic effects on ulcerative colitis mice and related mechanisms. METHODS HPLC-Q-Orbitrap-MS was used to qualitatively identify the chemical compositions, the data was analyzed and compared with database(mzCloud, mzVault, ChemSpider). Forty-two male BALB/c mice were randomly divided into 6 groups:normal group, model group, low, medium and high-dose groups of water extract of Sophora japonica, and positive control group, 7 in each group. The normal group was given drinking water, the other groups were administrated with 4% dextran sulphate sodium aqueous solution. All groups were given orally by gavage for 11 d at a dose of 10 mL·kg^-1. The changes of disease activity index(DAI) and microscopic pathological examination of colon tissue were observed. The levels of TNF-α and IL-1β in plasma were determined by ELISA. The expression of NF-κB p65 was detected by immunohistochemistry. RESULTS Three hundred twenty three compounds were matched in the database mzCloud, of which 22 were more than 90% matched. Compared with the model group, the colon length was longer and DAI index was lower in Sophora japonica water extract groups. The expression of NF-κB p65 in colon tissue was decreased, and the contents of TNF-α and IL-1β in plasma were decreased in a dose-dependent manner. CONCLUSION The Sophora japonica water extract can alleviate ulcerative colitis injury induced by dextran sulphate sodium in mice by inhibiting intestinal inflammatory reaction.