基于生物活性导向分离及灰色关联度分析的山楂核干馏液抗炎和抗菌作用物质基础研究

目的 研究山楂核干馏液抗炎和抗菌的主要物质基础。方法 使用干馏釜制备山楂核干馏液；通过大孔吸附树脂柱色谱对干馏液进行分离；基于气相色谱-质谱法对山楂核干馏液进行分析；以脂多糖诱导的小鼠单核巨噬细胞RAW264.7生成一氧化氮模型评价山楂核干馏液的抗炎作用，采用微稀释法评估其体外抑制白色念珠菌生长活性，采用灰色关联度分析法建立其谱效关系，以评估其抗炎和抗菌活性，并筛选主要活性化合物。结果 山楂核干馏液及其4个亚流分具有明显的抗炎作用和抑制白色念珠菌生长作用，其中Fr.B的抗炎和抗菌作用均较强。从山楂核干馏液4个流分的色谱图中确定12个共有峰，其中发挥主要抗炎作用的成分为4、6、2、9号峰，质谱分析鉴定4个峰分别对应2，6-二甲氧基-4-甲基苯酚、1，2，5-三甲氧基-3-甲基苯、2，6-二甲氧基苯酚和乙酰丁香酮；发挥主要抗菌作用的成分为4、2、9、11号峰，质谱分析鉴定这4个峰分别对应2，6-二甲氧基-4-甲基苯酚、2，6-二甲氧基苯酚、乙酰丁香酮和二氢芥子醇。结论 山楂核干馏液具有较好的抑制脂多糖诱导的RAW264.7细胞生成一氧化氮的作用，同时对白色念珠菌的生长具有显著的抑制作用，明确了山楂核干馏液中2，6-二甲氧基-4-甲基苯酚、1，2，5-三甲氧基-3-甲基苯、2，6-二甲氧基苯酚和乙酰丁香酮4个成分对其抗炎作用贡献最大，2，6-二甲氧基-4-甲基苯酚、2，6-二甲氧基苯酚、乙酰丁香酮和二氢芥子醇对其抗菌作用贡献最大。研究结果为山楂核资源的开发利用及相关产品质量标准的制定提供参考。

Anti-inflammatory and Anti-bacterial Effects of Destructive Distillation Extracts of Hawthorn Seed Based on Bioactivity-oriented Separation and Grey Relational Analysis

Objective To investigate the material basis of the anti-inflammatory and anti-bacterial effects of destructive distillation extracts of hawthorn seed.Methods Destructive distillation extracts of hawthorn seed were prepared in a distillation retort and separated by macroporous adsorption resin column chromatography, followed by gas chromatography-mass spectrometry analysis. The anti-inflammatory effects of destructive distillation extracts of hawthorn seed were evaluated by lipopolysaccharide (LPS)-induced nitric oxide (NO) production model in mouse mononuclear macrophage RAW264.7 cells. The microdilution method was used to evaluate its in vitro inhibitory effect on the growth activity of Candida albicans. The spectrum-effect relationship established by grey relational analysis was used to evaluate its anti-inflammatory and anti-bacterial activities and to screen the main active compounds.Results Destructive distillation extracts of hawthorn seed and four fractions, especially Fr.B, possessed obvious anti-inflammatory effects and inhibitory effects against the growth of C. albicans. From the chromatograms of the four fractions, twelve common peaks were identified. Among them, the top four peaks showing potent anti-inflammatory effects were peaks 4, 6, 2, and 9, respectively, which were identified by MS as 2,6-dimethoxy-4-methylphenol, 1,2,5-trimethoxy-3-methylbenzene, 2,6-dimethoxyphenol, and acetosyringone. The top four peaks showing potent anti-bacterial effects were peaks 4, 2, 9, and 11, respectively, which were identified by MS as 2,6-dimethoxy-4-methylphenol, 2,6-dimethoxyphenol, acetosyringone, and dihydrosinapyl alcohol.Conclusion Destructive distillation extracts of hawthorn seed have good inhibitory effects on LPS-induced NO production in RAW264.7 cells and significant inhibitory effects on the growth activity of C. albicans. The 2,6-dimethoxy-4-methylphenol, 1,2,5-trimethoxy-3-methylbenzene, 2,6-dimethoxyphenol, and acetosyringone in destructive distillation extracts of hawthorn seed make great contributions to the anti-inflammatory efficacy, while 2,6-dimethoxy-4-methylphenol, 2,6-dimethoxyphenol, acetosyringone, and dihydrosinapyl alcohol to anti-bacterial efficacy. The findings of this study are expected to provide references for the development and utilization of hawthorn seeds and the formulation of related product quality standards.