血叶兰化学成分及其生物活性研究

Chemical constituents and their biological activities of Ludisia discolor（Orchidaceae）

关键词:  血叶兰属，血叶兰，化学成分，抗炎活性，抗氧化活性

Key words:  Ludisia, Ludisia discolor, chemical constituent, anti-inflammatory activity, antioxidant activity

摘要:

为探究血叶兰（Ludisia discolor）的化学成分和生物活性，该研究利用气相色谱-质谱（GC-MS）分析乙醇提取物石油醚萃取相的化合物；利用硅胶、Sephadex LH-20和半制备高效液相等多种色谱技术对乙酸乙酯萃取相进行分离纯化，运用质谱和核磁共振波谱分析鉴定化合物结构；然后对分离得到的化合物进行抗炎和抗氧化活性测试。结果表明：（1）从石油醚萃取相中共鉴定出17个化合物，其中棕榈酸甲酯、亚油酸甲酯、亚麻酸甲酯、2-单棕榈酸甘油4个化合物具有较高的相对含量。（2）从乙酸乙酯萃取相中分离得到15个化合物，其结构分别鉴定为罗汉松脂素（1）、(+)-松脂素（2）、callyspongidipeptide A（3）、环-[(S)-脯氨酸-(R)-亮氨酸]（4）、epi-boscialin（5）、对苯二甲酸二丁酯（6）、对香豆酸甲酯（7）、对羟基苯乙酮（8）、对羟基苯甲醇（9）、对羟基苯甲醛（10）、香草醛（11）、4-(甲氧基甲基)苯酚（12）、β-谷甾醇（13）、肉豆蔻酸（14）和棕榈酸（15），化合物1-15均为首次从血叶兰中分离得到。（3）化合物1和2具有一定抑制LPS诱导RAW 264.7产生NO的作用，其IC50分别为（37.76±2.68）和（53.14±1.63） μmol?L-1 [阳性对照槲皮素IC50 值为（9.32±0.36） μmol?L-1]；抗氧化结果显示化合物1和2对DPPH自由基具有一定清除能力，IC50值分别为（51.22±1.07）和（79.22±7.44） μg?mL-1 [阳性对照维生素C IC50值为（6.01±0.17） μg?mL-1]；同时这两个化合物对ABTS?＋自由基表现出强于阳性对照trolox [IC50 =（34.65±0.53） μg?mL-1]的清除能力，其IC50值分别为（2.21±0.01）和（3.58±0.17） μg?mL-1。该研究丰富了血叶兰的化学成分，进一步明确了其抗氧化和抗炎活性成分，为后续药理活性的深入研究提供了化学结构基础。

Abstract:

In order to explore the chemical constituents and their biological activities of Ludisia discolor, gas chromatography-mass spectrometry (GC-MS) was carried out to analysis of the chemical components of petroleum ether fraction of ethanol extract. Meanwhile, the compounds were isolated from the ethyl acetate fraction by using various column chromatography including silica gel, Sephadex LH-20, and semi-preparative HPLC. MS and NMR spectroscopic data were used to identify the structure of the compounds. And then the isolated compounds were tested for anti-inflammatory with RAW 264.7 cells, and antioxidant activities were carried out with DPPH and ABTS?＋ free radical scavenging ability.The results showed as follows: (1) A total of 17 compounds were identified from the petroleum ether fraction of L. discolor, in which, methyl palmitate, methyl linoleate, methyl linolenic acid, and 2-monopalmitin had a relatively high content. (2) 15 compounds were isolated from the ethyl acetate fraction, their structures were identified as matairesinol (1), (＋)-pinoresinol (2), callyspongidipeptide A (3), cyclo-[(S)-Pro-(R)-Leu] (4), epi-boscialin (5), dibutyl terephthalate (6), methyl 4-hydroxycinnamate (7), 1-(4-hydroxyphenyl) ethanone (8), 4-hydroxybenzyl alcohol (9), p-hydroxy-benzaldehyde (10), vanillin (11), 4-(methoxym-ethyl) phenol (12), β-sitosterol (13), tetradecanoic acid (14), and palmitic acid (15), respectively. Compounds 1-15 were isolated from L. discolor for the first time. (3) Phenylpropanoid compounds 1 and 2 showed moderate inhibition on NO production from RAW264.7 cells induced by LPS, with IC50 values of (37.76±2.68) μmol?L-1 and (53.14±1.63) μmol?L-1 (quercetin as the positive control, with IC50 value of (9.32±0.36) μmol?L-1), respectively. Antioxidant results showed that compounds 1 and 2 had moderate DPPH free radical scavenging ability, with IC50 values of (51.22±1.07) μg?mL-1 and (79.22±7.44) μg?mL-1 [vitamin C as the positive control, with IC50 value of (6.01±0.17) μg?mL-1], respectively. Meanwhile, they had stronger ABTS?＋ free radical scavenging ability than the positive control trolox [IC50 = (34.65±0.53) μg?mL-1], with IC50 values of (2.21±0.01) μg?mL-1 and (3.58±0.17) μg?mL-1, respectively. Overall, this study enriched the chemical composition of L. discolor, and it was further clarified that phenolic compounds are the active antioxidant components in L. Discolor, and phenylpropanoid compounds have certain anti-inflammatory activity, which provides a chemical structure basis for subsequent in-depth pharmacological research.