目的 分析醋炙、黑豆制、甘草制、麸炒4种炮制方法对窄竹叶柴胡Bupleurum marginatum var. stenophyllum化学成分的影响，基于“成分-毒性”关联筛选肝毒性相关成分，探究减毒内涵。方法 以正常小鼠构建体内评价模型，通过检测血清中天冬氨酸氨基转移酶（aspartate aminotransferase，AST）、丙氨酸氨基转移酶（alanine aminotransferase，ALT）活性，结合肝脏组织病理学切片观察，明确治疗剂量下窄竹叶柴胡生品的肝毒性风险。以人正常肝细胞L02构建体外肝毒性评估模型，对比4种炮制品与生品对AST、ALT活性的影响，筛选最佳减毒炮制方法。采用超高效液相色谱-四级杆飞行时间串联质谱技术对不同产地炮制前后化学成分进行表征；通过主成分分析（principal component analysis，PCA）、偏最小二乘判别分析（partial least squares-discriminant analysis，PLS-DA）及正交偏最小二乘判别分析（orthogonal partial least squares-discriminant analysis，OPLS-DA）比较炮制前后成分变化。通过皮尔森相关性分析、灰色关联度分析建立共有成分峰面积与L02细胞AST、ALT水平的相关性，筛选潜在肝毒性成分。以斑马鱼为模型，考察柴胡皂苷a（saikosaponin a，SSa）、柴胡皂苷d（saikosaponin d，SSd）、柴胡皂苷b₁（saikosaponin b₁，SSb₁）、柴胡皂苷b₂（saikosaponin b₂，SSb₂）的肝毒性差异。结果 血清生化指标检测结果表明，与空白对照组相比，窄竹叶柴胡生品高、中剂量组的AST、ALT水平显著升高（P＜0.01、0.05），组织病理学结果显示窄竹叶柴胡并未引起病理性肝损伤。醋炙品和黑豆制品组的AST、ALT水平较生品组显著降低（P＜0.05、0.01），甘草制组次之，而麸炒组无显著差异。生品和不同炮制品共鉴别出37个成分，其中共有成分27个，醋炙、黑豆制、甘草制后化学成分发生了变化。SSa、3"-O-乙酰基-SSa、4"-O-乙酰基-SSa、SSd、丙二酰基-SSa、3"-O-乙酰基-SSd为窄竹叶柴胡的潜在肝毒性成分。SSd潜在肝毒性大于SSa，转化后的SSb₂、SSb₁的毒性降低。结论 窄竹叶柴胡生品对肝功能指标有一定的影响，醋炙、黑豆制是有效减毒方法，其减毒的核心机制与促进SSa和SSd转化为低毒性皂苷有关。

Objective To analyze the effects of four processing methods (vinegar stir-frying, black soybean processing, glycyrrhiza processing and bran stir-frying) on the chemical components of Bupleurum marginatum var. stenophyllum (BM), screen its hepatotoxic components based on the “ingredient-toxicity” correlation system, and explore the connotation of attenuation. Methods Normal mice were used to establish an in vivo evaluation model. The activities of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in serum were detected, and the liver histopathological sections were observed to clarify the risk of liver toxicity of the crude products at therapeutic doses. Human normal liver cell L02 was used to construct an in vitro hepatotoxicity evaluation model, and the effects of four processed products and raw products on AST and ALT activities were compared to screen the best attenuation processing method. Ultra-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry was used to characterize the chemical components before and after processing in different regions. Principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were used to compare the changes of components before and after processing. Pearson correlation analysis and grey correlation analysis were used to establish the correlation between the peak area of common components and the levels of AST and ALT in L02 cells, and to screen potential hepatotoxic components. Zebrafish was used as a model to investigate the differences in hepatotoxicity of saikosaponin a (SSa), saikosaponin d (SSd), saikosaponin b₁ (SSb₁), and saikosaponin b₂ (SSb₂). Results The results of serum biochemical index detection indicated that compared with the blank control group, the levels of AST and ALT in the high-dose and medium-dose groups of raw B. marginatum var. stenophyllum were significantly increased (P < 0.01, 0.05). The histopathological results showed that B. marginatum var. stenophyllum did not cause pathological liver injury.AST and ALT levels in vinegar stir-frying and black soybean processing groups were significantly lower than those in raw product group (P < 0.05, 0.01), followed by glycyrrhiza group, while there was no significant difference in bran stir fried products. A total of 37 components were identified in the raw products and different processed products, including 27 components. The chemical composition changed after vinegar stir-frying, black soybean processing, and licorice processing. SSa, 3"-O-acetyl-SSA, 4"-O-acetyl-SSa, SSd, malonyl-SSa, and 3"-O-acetyl-SSd were the potential hepatotoxic components of BM. The potential hepatotoxicity of SSd was greater than that of SSa, and the toxicity of transformed SSb₂ and SSb₁ was reduced. Conclusion The crude products of BM have certain effects on liver function indexes, and the preparation with vinegar stir-frying and black soybean is an effective method for attenuation. The core mechanism of attenuation is related to promoting the conversion of SSa and SSd into low toxic saponins.

R284.1

黑龙江省自然科学基金联合引导项目(PL2024H196);黑龙江省普通本科高等学校“优秀青年教师基础研究支持计划”项目(YQJH2024086);黑龙江省高等学校基本科研业务费项目(2024-KYYWF-1018);龙江科技英才春雁支持计划(CYCX24009);鸡西市“揭榜挂帅”科技攻关项目(JKJB2024H03)