摘要
目的:建立四妙凉血颗粒中5种成分的含量测定方法,探讨四妙凉血颗粒对急性痛风性关节炎(acute gouty arthritis,AGA)大鼠的抗炎作用机制。方法:采用高效液相色谱法测定四妙凉血颗粒中5种成分含量;大鼠足踝关节注射尿酸钠悬液诱导AGA,采用容积法检测足踝关节肿胀程度;采用酶联免疫吸附试验法测定大鼠血清及关节液中白细胞介素1β(interleukin-1β,IL-1β)、IL-6和肿瘤坏死因子α(tumor necrosis factorα,TNF-α)含量;采用免疫组织化学和图像分析方法观测大鼠踝关节软骨组织中TNF-α蛋白表达水平。结果:方法学考察结果显示各项指标符合规定,四妙凉血颗粒样品中落新妇苷、盐酸小檗碱、山柰酚、黄芩素和汉黄芩素的平均回收率为97.14%~102.67%,RSD值均小于3.0%。尿酸钠混悬液注射后,与空白对照组比较,模型组不同时间点大鼠足踝关节肿胀度,关节液中IL-1β、IL-6、TNF-α含量,血清IL-6水平及TNF-α蛋白表达均增高(P<0.05);空白对照组大鼠软骨细胞可见少量TNF-α蛋白表达。与模型组比较,阳性组(秋水仙碱0.65 mg/kg)及四妙凉血颗粒高、中、低剂量(6、3、1.5 g/kg)组大鼠第3~5天足踝关节肿胀度,关节液中IL-1β、IL-6、TNF-α含量及血清IL-6、TNF-α水平均降低(P<0.05);四妙凉血颗粒低、中、高剂量组大鼠TNF-α蛋白积分吸光度降低(P<0.05)。与阳性组比较,四妙凉血颗粒高、中、低剂量组大鼠足踝关节肿胀度,关节液中IL-1β、IL-6、TNF-α含量及血清IL-6、TNF-α表达水平和关节组织TNF-α蛋白积分吸光度均变化不明显,差异无统计学意义(P>0.05)。结论:本实验建立了四妙凉血颗粒含量测定方法,该方法专属性强、重复性好、准确度高,可用于四妙凉血颗粒样品中落新妇苷、盐酸小檗碱、山柰酚、黄芩素和汉黄芩素的含量测定;四妙凉血颗粒治疗AGA的作用机制可能与其降低关节组织炎症因子IL-1β、IL-6和TNF-α水平有关。
Objective:To establish the determination methods for the contents of five components in Simiao Liangxue granules and to discuss the anti-inflammatory mechanism of the medicine on rats with acute gouty arthritis(AGA).Methods:HPLC was used to detect the contents of five components in the medicine;AGA rat models were prepared by injecting sodium urate suspension into foot and ankle,the swelling degree of ankle joint was detected by volume method;ELISA method was adopted to measure the contents of IL-1β,IL-6 and TNF-αin the serum and joint fluid;immunohistochemical and image analysis methods were utilized to observe the expressions of TNF-αprotein in cartilage tissue of ankle.Results:The results of methodological examination showed that all the indicators conformed to the regulations,average recovery rates of astilbin,berberine hydrochloride,kaempferol,baicalein and scutellarin ranged between 97.14%and 102.67%in the samples of the medicine,RSD values were smaller than 3.0%.After the injection of sodium urate suspension,compared with blank control group,more significant swelling of ankle joint could be seen,the levels of IL-1β,IL-6 and TNF-αin the joint fluid,the expressions of IL-6 and TNF-αin the serum were elevated in the model group at different time points(P<0.05);small amounts of TNF-αprotein expressions could been found in the blank control group.Compared with the model group,the swelling of ankle joints in the positive group(colchicin,0.65 mg/kg),high,moderate and low doses groups of Simiao Liangxue granules(6,3 and 1.5 g/kg)was alleviated from the third to the fifth day,the contents of IL-1β,IL-6 and TNF-αin joint fluid,the levels of IL-6 and TNF-αin the serum were reduced(P<0.05);TNF-αprotein integral absorbance was lowered in the low,moderate and high doses groups of the medicine(P<0.05).Compared with the positive group,the swelling of ankle joints,the contents of IL-1β,IL-6 and TNF-αin joint fluid,the levels of IL-6 and TNF-αin the serum,as well as TNF-αprotein integral absorbance in the joint tissue were changed insignificantly,and the difference had no statistical meaning(P>0.05).Conclusion:The determination method for the contents of the granules established in the study,exclusive,reproducible and accurate,could be used for the content determination of astilbin,berberine hydrochloride,kaemp ferol,baicalein and scutellarin in the samples of the granules;the mechanism of treating AGA with the medicine may be related to the reduction in the levels of local inflammation factors IL-1β,IL-6 and TNF-αin the joints.
作者
唐春丽
农必华
冯茵怡
林忆龙
魏江存
罗远
TANG Chunli;NONG Bihua;FENG Yinyi;LIN Yilong;WEI Jiangcun;LUO Yuan(The First Affiliated Hospital of Guangxi University of Chinese Medicine,Nanning 530023,China;Guangxi International Zhuang Medicine Hospital,Nanning 530201,China)
出处
《西部中医药》
2025年第2期1-5,共5页
Western Journal of Traditional Chinese Medicine
基金
国家中医药管理局“十二五”中医药重点学科(民族药学)壮药学资质项目
全国中药特色技术传承人才培训项目(20184828005)
广西自然科学基金项目(2020GXNSFAA259059)
广西中医药大学高层次人才培育创新团队资助项目(2022A008)。
作者简介
唐春丽(1980-),女,硕士学位,副主任中药师。研究方向:中药及其制剂质量分析;通讯作者:魏江存(1989-),男,硕士学位,助理研究员。研究方向:中药、民族药质量分析和药效机制研究。Email:960837714@qq.com。
