WO2006102789A1 - Ferula fukanensis extract, the preperation method and the use in producing medicine thereof - Google Patents

Abstract

The present invention provides Ferula fukanensis extract, the preparation method and the use in producing medicine thereof. The method of extracting the active compounds includes:1) The Ferula fukanensis is extracted with methanol, the extract solution is concentrated under reduced pressure and dried,then extract is obtained; 2) The extract is extracted with chloroform and ethyl acetate,successively; 3) The chloroform fraction is subjected to column chromatography, eluted with the mixed solution of n-hexane and ethyle acetate; 4) The eluent is further seperated and purified by High Performance Liquid Chromatography, 11 active compounds are obtained. The compounds or the salt, enantiomer, racemer, tautomer or physiological derivative thereof which are extracted from Ferula fukanensis, can be used to prepare the medicine which can treat or prevent inflammation or tumor.

Description

阜康阿魏提取物、 其制备方法及其在制备药物中的应用 技术领域  Extract of Fukang Awei, preparation method thereof and application thereof in preparing medicine

本发明涉及阜康阿魏提取物、 其制备方法及其在制备药物 中的应用， 特别是涉及从阜康阿魏中提取具有抗炎活性和抗肿 瘤活性的化合物。 背景技术  The present invention relates to a extract of Frucco sinensis, a preparation method thereof and use thereof in the preparation of a medicament, and in particular to a compound which has anti-inflammatory activity and anti-tumor activity extracted from Fukang Awei. Background technique

困扰人们的日常炎性疾病包括， 关节炎症例如骨关节炎、 类风湿性关节炎、 类风湿性脊推炎、 痛风关节炎等； 炎性皮肤 病例如湿疹、 牛皮癣、 皮炎等； 炎性眼疾例如眼色素层炎、 结 膜炎等； 肺部疾病例如哮喘、 支气管炎、 急性呼吸窘迫综合症 等； 菌血症、 那毒素血症、 口疮溃疡、 龈炎、 胰腺炎等； 胃肠 道疾病例如节段性回肠炎、 萎缩性胃炎、 溃疡性结肠炎、 腹腔 炎、 消化性溃疡、 应激性肠道综合症例如由幽门螺旋杆菌感染 而引起的粘膜炎症或者由非甾体类抗炎药引起的肠胃病等等。  Daily inflammatory diseases that plague people include joint inflammation such as osteoarthritis, rheumatoid arthritis, rheumatoid arthritis, gout arthritis, etc.; inflammatory skin diseases such as eczema, psoriasis, dermatitis, etc.; inflammatory eye diseases such as Uveitis, conjunctivitis, etc.; pulmonary diseases such as asthma, bronchitis, acute respiratory distress syndrome, etc.; bacteremia, toxemia, aphthous ulcer, gingivitis, pancreatitis, etc.; gastrointestinal diseases such as festivals Segmental ileitis, atrophic gastritis, ulcerative colitis, abdominal inflammation, peptic ulcer, irritable bowel syndrome such as mucosal inflammation caused by H. pylori infection or caused by non-steroidal anti-inflammatory drugs Gastrointestinal diseases and so on.

有关各种炎症致病作用的机理是： 已知体内一氧化氮 （ N 〇 ) 是介导细胞免疫和炎症的毒性物质。 其前体是左旋精氨酸 (L-arg),L-arg在 N 0合成酶（N 0 S )的作用下生成 N 0。 目前 已经分离出三种不同类型的 NOS，包括内皮 NO合成酶（eNOS), 神经元 NO合成酶（nNOS)及诱导型 NO合成酶（iNOS)。 目前已 知，巨噬细胞、 肝细胞、 平滑肌细胞、 腺癌细胞以及上皮细胞均 能表达 iNOS。某些炎性细胞因子和微生物产物如脂多糖（ LPS ) 则可诱导 iNOS 表达。 iNOS —旦被诱导即可表达出高度活性, 产生大量 NO , 进而导致细胞损伤， 造成炎症恶化， 甚至导致癌 症。 因此， 长时间以来， 人们致力于寻找 NO 合成酶的抑制剂 来治疗与之相关的炎性疾病。 但这些抑制剂大多局限于化学合 成的物质， 副作用较大。 有关的科学工作者始终不遗余力的试 图开发新型的疗效确切、 副作用少、 研制成本低的抗炎活性化 合物。 加深对中草药的研究可能是实现这个愿望的良好载体。 The mechanism of various inflammatory pathogenic effects is: It is known that nitric oxide (N 〇 ) is a toxic substance that mediates cellular immunity and inflammation. Its precursor is L-arginine (L-arg), and L-arg produces N 0 under the action of N 0 synthase (N 0 S ). Three different types of NOS have been isolated, including endothelial NO synthase (eNOS), neuronal NO synthase (nNOS) and inducible NO synthase (iNOS). It is known that macrophages, hepatocytes, smooth muscle cells, adenocarcinoma cells, and epithelial cells all express iNOS. Certain inflammatory cytokines and microbial products such as lipopolysaccharide (LPS) induce iNOS expression. iNOS is highly active when it is induced, producing a large amount of NO, which leads to cell damage, inflammation, and even cancer. Therefore, for a long time, efforts have been made to find inhibitors of NO synthase to treat inflammatory diseases associated therewith. However, most of these inhibitors are limited to chemically synthesized substances with large side effects. Relevant scientists have spared no effort to develop new types of anti-inflammatory active compounds with definite curative effects, few side effects, and low development costs. Deepening the study of Chinese herbal medicines may be a good vehicle to achieve this desire.

癌症是当前危害人类健康的严重疾病，抗癌化学药物在杀灭 癌细胞的同时，对人体正常细胞也产生毒副作用，因此从天然的 动、 植、 矿物中寻找疗效高、 毒性低的抗癌成分是近年来国内 外学者非常重视的课题。如美国国立癌症研究所自 1957年以来， 每年评价 1500 ~ 5000种植物提取物的抗癌活性成分，至今被筛 选的植物在 30,000 种以上，被评价的提取物在 100, 000种以上。 据美国 《 Annual Re2port s of Medicinal Chemistry》 报道，在 1984 - 1995年间被 FDA批准生产的 31 种新抗癌药中（不包含 生物制品）， 61 %来源于天然产物、半合成品或以天然产物为模型 的全合成品。 1994年可供临床使用的 87种抗癌药中， 62 %以天 然产物为基源。 目前在除对 2000多种植物进行了抗癌活性成分 的筛选外，也对传统中药进行了大量的研究，从复方、 单方、 单一 成分、 有效部位及提取物等中寻找有效成分、 新的药物。  Cancer is a serious disease that currently harms human health. Anticancer chemical drugs also have toxic side effects on normal human cells while killing cancer cells. Therefore, it is necessary to find anti-cancer with high efficacy and low toxicity from natural animals, plants and minerals. The ingredients are very important topics for scholars at home and abroad in recent years. For example, since 1957, the National Cancer Institute has evaluated 1,500 to 5,000 plant extracts of anticancer active ingredients each year. More than 30,000 plants have been screened so far, and more than 100,000 extracts have been evaluated. According to the US Annual Re2port s of Medicinal Chemistry, 31 new anticancer drugs (excluding biological products) approved by the FDA between 1984 and 1995, 61% are derived from natural products, semi-synthetic products or natural products. For the full synthesis of the model. Of the 87 anticancer drugs available for clinical use in 1994, 62% were based on natural products. At present, in addition to the screening of anti-cancer active ingredients for more than 2,000 plants, a large number of studies have been carried out on traditional Chinese medicines, and active ingredients, new drugs have been sought from compounds, unilateral, single ingredients, effective parts and extracts. .

阜康阿魏 Ferula fukanensis、 是主要分布在中亚的干燥 陆地，以前的研究通过 GC-MS(CI/EI)分析了这种植物的聚硫烷 ( polysulfanes ) ，且许多小组研究了阿魏属 （伞形科） 植物的 化学成分。 该属常见的化合物是倍半萜烯 （特别是胡萝 卜烷、 蛇麻烷和愈创木烷）、 倍半萜烯香豆素和倍半萜烯色酮。  Ferula fukanensis, which is mainly distributed in the dry land of Central Asia, was previously analyzed by GC-MS (CI/EI) for the polysulfanes of this plant, and many groups studied the genus (Umbelliferae) The chemical composition of plants. Common compounds of this genus are sesquiterpenes (especially carbosilane, hopsane and guaiacene), sesquiterpene coumarin and sesquiterpene chromone.

由于植物药副作用小， 研究和制取的成本低， 所以从中草 药中发现有确切疗效的药物活性成分， 将是一个可取的尝试。 发明内容 Due to the small side effects of botanicals and the low cost of research and preparation, it is a desirable attempt to find a pharmaceutically active ingredient with a definite therapeutic effect from Chinese herbal medicine. Summary of the invention

本发明的发明 目 的是通过***研究阜康阿魏中的活性成 分， 来寻找可用于制备抗炎药物的有效活性成分以及可用于制 备抗肿瘤药物的有效活性成分。  SUMMARY OF THE INVENTION The object of the present invention is to find an effective active ingredient which can be used for the preparation of an anti-inflammatory drug and an effective active ingredient which can be used for the preparation of an antitumor drug by systematically studying the active ingredient in the Frucco.

本发明的另一个目的是提供一种从阜康阿魏植物中提取活 性成分的方法。  Another object of the present invention is to provide a method for extracting an active ingredient from a F. sylvestris plant.

本发明还提供了从阜康阿魏中提取的化合物用于制备治疗 有关炎性疾病的药物用途。  The present invention also provides a compound for use in the preparation of a medicament for the treatment of an inflammatory disease.

本发明还提供了从阜康阿魏中提取的化合物用于制备治疗 肿瘤的药物用途。  The invention also provides a pharmaceutical use of a compound extracted from Fukang Awei for the preparation of a tumor.

在本发明中提供了从阜康阿魏中提取的抗炎活性化合物 An anti-inflammatory active compound extracted from Fukang Awei is provided in the present invention.

( 1 )、 （ 2)、 （ 3)、 （ 4)、 ( 5) ( 6) 结构式如下： (1), (2), (3), (4), (5) (6) The structural formula is as follows:

在本发明中提供了从阜康阿魏中提取的抗肿瘤活性化合物

In the present invention, an antitumor active compound extracted from Fukang Awei is provided.

( 7)、 （ 8)、 （ 9)、 ( 10) ( 11) 结构式如下： (7), (8), (9), (10) (11) The structural formula is as follows:

化合物 （ 11 ) 本发明从阜康阿魏中提取活性化合物的方法包括如下步 骤：

Compound (11) The method for extracting an active compound from Fukang Awei of the present invention comprises the following steps:

1 ) 阜康阿魏经曱醇提取后的提取液减压浓缩、 干燥得到萃取 物；  1) The extract of the ruthenium ferulate is concentrated under reduced pressure and dried to obtain an extract;

2 ) 将萃取物用氯仿、 乙酸乙酯依次萃取；  2) extracting the extract sequentially with chloroform and ethyl acetate;

3 ) 将氯仿的萃取物经层析柱用正己烷与乙酸乙酯的混合液洗 脱；  3) The chloroform extract is eluted through a chromatography column with a mixture of n-hexane and ethyl acetate;

4 ) 对步骤 3 ) 收集到的洗脱液经高压液相色谱进一步分离精 制， 得到活性化合物。  4) The eluate collected in step 3) is further separated and purified by high pressure liquid chromatography to obtain an active compound.

本发明的另一个方面是提供从阜康阿魏中提取的抗炎活性 化合物或其药学上可接受的盐、 对映体、 外消旋体或互变异构 体在用于治疗或预防抑制 NO 生成活性是有益的疾病的药物制 备中的用途。  Another aspect of the invention provides an anti-inflammatory active compound, or a pharmaceutically acceptable salt, enantiomer, racemate or tautomer thereof, for use in the treatment or prophylaxis of an anti-inflammatory active compound or a pharmaceutically acceptable salt thereof NO production activity is a useful drug preparation in the manufacture of diseases.

本发明的另一个方面是提供从阜康阿魏中提取的抗炎活性 化合物或其药学上可接受的盐、 对映体、 外消旋体、 互变异构 体或生理官能衍生物在用于治疗或预防炎性疾病的药物制备中 的用途。  Another aspect of the present invention provides the use of an anti-inflammatory active compound or a pharmaceutically acceptable salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, which is extracted from Fukang Use in the preparation of a medicament for the treatment or prevention of an inflammatory disease.

适宜的盐包括与有机和无机酸或碱形成的盐； 药学上可接 受的盐包括与下列酸形成： 盐酸、 氢溴酸、 硫酸、 柠檬酸、 酒 石酸、 磷酸、 乳酸、 丙酮酸、 乙酸、 三氟乙酸、 琥珀酸、 草酸、 富马酸、 马来酸、 丁酮二酸、 曱磺酸、 乙磺酸、 对曱苯磺酸、 苯磺酸、 羟乙磺酸； 药学上可接受的碱式盐包括铵盐、 碱金属 盐、 如钠盐和钾盐、 碱土金属盐如钙盐和镁盐以及与有机碱如 二环己基胺和 N - 曱基 - D -葡糖胺形成的盐。 Suitable salts include those formed with organic and inorganic acids or bases; pharmaceutically acceptable salts include those formed with hydrochloric acid, hydrobromic acid, sulfuric acid, citric acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, Fluoroacetic acid, succinic acid, oxalic acid, fumaric acid, maleic acid, butanone diacid, sulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, isethionate; pharmaceutically acceptable base Salts include ammonium salts, alkali metal salts such as sodium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, and organic bases such as A salt formed from dicyclohexylamine and N-decyl-D-glucosamine.

生理官能衍生物的实例包括酯、 酰胺、 氨基曱酸酯， 优选 酯和酰胺。  Examples of physiologically functional derivatives include esters, amides, amino phthalates, preferably esters and amides.

本发明的化合物还可有利地与第二种药学抗炎活性物质， 尤其是可诱导的环氧酶（ COX - 2 ) 同工酶的选择性抑制剂联合 用药， 具体的本发明提供的化合物或其药学上可接受的盐、 对 映体、 外消旋体或互变异构体与 COX - 2抑制剂联合用药以治 疗炎症、 炎性疾病以及与炎症相关的疾病的用途。  The compounds of the invention may also be advantageously administered in combination with a second pharmaceutical anti-inflammatory active, in particular a selective inhibitor of an inducible cyclooxygenase (COX-2) isoenzyme, specifically a compound provided herein or Use of a pharmaceutically acceptable salt, enantiomer, racemate or tautomer thereof in combination with a COX-2 inhibitor for the treatment of inflammation, inflammatory diseases, and diseases associated with inflammation.

炎性疾病及临床病症包括关节疾病， 特别是关节炎 （例如 类风湿性关节炎、 骨关节炎）、 或胃肠道疾病（如溃疡性结肠炎、 胃炎、 以及其它感染引起的粘膜炎症、 由非甾体抗炎药引起的 肠病）、 肺部疾病 （如成人呼吸道窘迫症、 哮喘、 嚢纤维化、 慢 性阻塞性肺部疾病）、 心脏病 （如心肌炎）、 神经组织疾病 （如 多发性硬化病）、 胰腺疾病 （如糖尿病及并发症）、 肾疾病 （如 肾小球性肾炎）、 皮肤病 （如皮炎、 牛皮癣、 湿疹、 荨麻疹）、 眼部疾病 （如青光眼）、 移植器官疾病 （如排斥反应）、 多器官 疾病 （如***性红斑狼疮） 及病毒和细菌感染后的炎性后遗症。  Inflammatory diseases and clinical conditions include joint diseases, especially arthritis (such as rheumatoid arthritis, osteoarthritis), or gastrointestinal diseases (such as ulcerative colitis, gastritis, and other infections caused by mucosal inflammation, by Intestinal diseases caused by non-steroidal anti-inflammatory drugs), lung diseases (such as adult respiratory distress, asthma, sputum fibrosis, chronic obstructive pulmonary disease), heart disease (such as myocarditis), neurological diseases (such as multiple Sclerosis), pancreatic diseases (such as diabetes and complications), kidney diseases (such as glomerulonephritis), skin diseases (such as dermatitis, psoriasis, eczema, urticaria), eye diseases (such as glaucoma), transplant disease (eg rejection), multiple organ diseases (eg systemic lupus erythematosus) and inflammatory sequelae following viral and bacterial infections.

此外， 本发明提供的化合物可以有助于预防或治疗与 HIV 感染有关的淋巴细胞的丟失、 增加放射治疗期间肿瘤细胞的放 射性敏感性、 降低肿瘤的生长、 肿瘤发展、 血管生成和肿瘤转 移。  Furthermore, the compounds provided by the present invention can help prevent or treat lymphocyte loss associated with HIV infection, increase radiosensitivity of tumor cells during radiation therapy, reduce tumor growth, tumor development, angiogenesis, and tumor metastasis.

本发明提供的化合物还可以用于制备保健食品、 饮料、 或 饲料等。  The compounds provided by the present invention can also be used in the preparation of health foods, beverages, or feeds and the like.

通过本发明， 提供拥有 NO 生成抑制作用的新规氧杂萘邻 酮诱导体化合物成为可能。 还能够提供抑制 IL- 6、 IL-1 的 mRNA化合物。 本发明的另一个方面是提供从阜康阿魏中提取的抗肿瘤活 性化合物或其药学上可接受的盐、 对映体、 外消旋体或互变异 构体在用于治疗或预防肿瘤的药物制备中的用途。 According to the present invention, it is possible to provide a novel oxaphthalene ketone inducer compound having an inhibitory effect on NO production. It is also possible to provide an mRNA compound that inhibits IL-6 and IL-1. Another aspect of the present invention provides an antitumor active compound or a pharmaceutically acceptable salt, enantiomer, racemate or tautomer thereof for extracting or treating a tumor, or a pharmaceutically acceptable salt, enantiomer, racemate or tautomer thereof for use in treating or preventing a tumor Use in the preparation of a drug.

本发明的另一个方面是提供从阜康阿魏中提取的抗肿瘤活 性化合物或其药学上可接受的盐、 对映体、 外消旋体、 互变异 构体或生理官能衍生物在用于治疗或预防肿瘤的药物制备中的 用途。  Another aspect of the present invention provides an antitumor active compound or a pharmaceutically acceptable salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, which is extracted from Fukang Use in the preparation of a medicament for the treatment or prevention of tumors.

适宜的盐包括与有机和无机酸或碱形成的盐； 药学上可接 受的盐包括与下列酸形成： 盐酸、 氢溴酸、 硫酸、 柠檬酸、 酒 石酸、 磷酸、 乳酸、 丙酮酸、 乙酸、 三氟乙酸、 琥珀酸、 草酸、 富马酸、 马来酸、 丁酮二酸、 曱磺酸、 乙磺酸、 对曱苯磺酸、 苯磺酸、 羟乙磺酸； 药学上可接受的碱式盐包括铵盐、 碱金属 盐、 如钠盐和钾盐、 减土金属盐如钙盐和镁盐以及与有机碱如 二环己基胺和 N -甲基 - D -葡糖胺形成的盐。  Suitable salts include those formed with organic and inorganic acids or bases; pharmaceutically acceptable salts include those formed with hydrochloric acid, hydrobromic acid, sulfuric acid, citric acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, Fluoroacetic acid, succinic acid, oxalic acid, fumaric acid, maleic acid, butanone diacid, sulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, isethionate; pharmaceutically acceptable base Salts include ammonium salts, alkali metal salts such as sodium and potassium salts, reduced earth metal salts such as calcium and magnesium salts, and salts with organic bases such as dicyclohexylamine and N-methyl-D-glucosamine. .

. 生理官能衍生物的实例包括酯、 酰胺、 氨基曱酸酯， 优选 酯和酰胺。  Examples of physiologically functional derivatives include esters, amides, amino phthalates, preferably esters and amides.

本发明的化合物还可有利地与第二种药学抗肿瘤活性物 质， 尤其是与抑制拓朴异构酶活性药物联合用药， 具体的本发 明提供的化合物或其药学上可接受的盐、 对映体、 外消旋体或 互变异构体与拓朴异构酶抑制剂联合用药肿瘤疾病的用途。  The compounds of the invention may also be advantageously administered in combination with a second pharmaceutical antitumor active, especially in combination with a medicament for inhibiting topoisomerase activity, specifically a compound provided herein, or a pharmaceutically acceptable salt, enantiomer thereof, Use of a racemic or tautomer in combination with a topoisomerase inhibitor for the treatment of neoplastic diseases.

在本发明中提供从阜康阿魏的根茎提取、 离析的新型倍半 萜烯香豆素、 倍半萜烯色酮、 倍半萜烯苯丙基类衍生物为有效 成分的抗肿瘤剂。 此抗肿瘤剂具有确切的诱导 HL - 60 细胞调 亡的活性， 此抗肿瘤剂诱导骨髓芽球性白血病细胞凋亡作用具 有非常好的效果， 对于恶性肿瘤等的治疗非常有效。 有望制成 副作用较少的抗肿瘤药物。 本发明的化合物为提供新的抗过敏剂、 抗炎症剂、 抗肿瘤 剂成为可能。 附图说明 In the present invention, an antitumor agent containing a novel sesquiterpene coumarin, a sesquiterpene ketone, and a sesquiterpene phenylpropyl derivative extracted and isolated from the rhizome of F. sylvestris is provided as an active ingredient. The antitumor agent has an exact activity for inducing apoptosis of HL-60 cells, and the antitumor agent has a very good effect on inducing apoptosis of myeloid leukemia cells, and is very effective for the treatment of malignant tumors and the like. It is expected to be made into anti-tumor drugs with fewer side effects. The compounds of the present invention are possible to provide new anti-allergic agents, anti-inflammatory agents, and anti-tumor agents. DRAWINGS

图 1发现抑制 IL-lb， IL-6的 mRNA作用 具体实施方式  Figure 1 shows the inhibition of IL-lb, IL-6 mRNA action.

实施例 1: 提取物的制备与化合物结构的确定  Example 1: Preparation of extract and determination of compound structure

将 5.9kg的阜康阿魏根粉碎，加入 80%曱醇（曱醇：水 = 8:2) 38升，在室温中放置 12个小时。经过一个小时左右的加热循环， 得到萃取液。 萃取后的阜康阿魏再度用 80%的曱醇 38L提取， 与之前得到的萃取液混合。此溶液 40°C的条件下进行减压浓缩， 得到 711 的褐色粉末萃取物。  5.9 kg of Fukang Aweigen was pulverized, and 80 liters of 80% sterol (sterol: water = 8:2) was added and allowed to stand at room temperature for 12 hours. After an hour or so of heating, an extract is obtained. The extracted 阜康阿魏 was again extracted with 80% sterol 38L and mixed with the previously obtained extract. This solution was concentrated under reduced pressure at 40 ° C to give a brown powder extract of 711.

取粉末萃取物 448.8 g溶解于 3L水中， 然后用氯仿、 乙酸 乙酯各 3L,依次提取三次。将其在 40°C的条件下进行减压浓缩， 得到氯仿馏分 272.4g、 乙酸乙酯馏分 142.0g。  448.8 g of the powder extract was dissolved in 3 L of water, and then 3 L of each of chloroform and ethyl acetate was successively extracted three times. This was concentrated under reduced pressure at 40 ° C to obtain 272.4 g of chloroform fraction and 142.0 g of ethyl acetate fraction.

取氯仿萃取物 70g, 用硅胶层析柱（ Wako gel C一 300和光 纯药， 12xl7cm)进行分离。 用正己烷：乙酸乙酯进行洗脱萃取， 依次以浓度为 0%、 2%、 5%、 10%、 20%、 40%、 60%、 100%的乙 酸乙酯 3L洗脱， 各分离出 200ml的馏分 1~ 11， 其中使用 10%浓度洗 脱时分別收集 1升和 2升洗脱液的熘分， 类似的 20%、 40%浓度也收集 2个馏分。  70 g of a chloroform extract was taken and separated by a silica gel column (wako gel C-300 and koji, 12 x 17 cm). The extract was eluted with n-hexane:ethyl acetate, and eluted sequentially with 0%, 2%, 5%, 10%, 20%, 40%, 60%, 100% ethyl acetate (3 L). 200 ml fractions 1 to 11, respectively, were collected using 1% and 2 liters of eluate at 10% concentration, and 2 fractions were collected at similar concentrations of 20% and 40%.

正己烷：乙酸乙酯 =90: 10溶液， 在 40°C的条件下减压浓缩。 所得浓缩物用 （6.5 g) 层析柱 （Wako gel C一 300和光纯药、 3x20cm)进行分离， 用正己烷：乙酸乙酯 = 100:0、 95:5、 90:10、 85:15、 80:20、 70:30、 60:40、 50:50、 0:100溶液洗脱， 在 40 °C 的条件下进行减压浓缩， 得 1.2 g浓缩物。 浓缩物用正向 HPLC (正已烷-乙酸乙酯， 85:15， 流速 9 ml/分， UV检测器设置在 254 nm )分离并进一步纯化，产生化合物（ 1 )( 13.7 mg, BtlS.S 分）， 通过反向 HPLC ( 乙氰 -水， 87:13， 流速 9 ml/分， UV检 测器设置在 210 nm) 产生化合物 （ 2 ) ( 13.2 mg, Εέ14.5分）、 化合物 （ 3 ) ( 8,0 mg, Rt 15.3分）、 化合物 （ 4 ) ( 23.0 mg, Rt 21.0分） 和化合物 （ 5 ) ( 9.7 mg, Rt 26.5分）， 通过反向 HPLC ( 乙氰 -水， 72:28, 流速 9 ml/分， UV检测器设置在 210 nm) 产生化合物 （ 6) ( 18.3 mg, Rt 11.1分）。 N-hexane: ethyl acetate = 90: 10 solution, and concentrated under reduced pressure at 40 °C. The obtained concentrate was separated with a (6.5 g) column (Wako gel C-300 and Wako Pure Chemical Industries, 3×20 cm) using n-hexane:ethyl acetate = 100:0, 95:5, 90:10, 85:15, 80:20, 70:30, 60:40, 50:50, 0:100 solution elution at 40 °C Concentration under reduced pressure gave 1.2 g of a concentrate. The concentrate was separated by forward HPLC (n-hexane-ethyl acetate, 85:15, flow rate 9 ml/min, UV detector set at 254 nm) and further purified to yield compound (1) (13.7 mg, BtlS.S a compound ( 2 ) ( 13.2 mg, Εέ 14.5 min), compound ( 3 ) by reversed phase HPLC (acetonitrile-water, 87:13, flow rate 9 ml/min, UV detector set at 210 nm) (8,0 mg, Rt 15.3 min), compound (4) (23.0 mg, Rt 21.0 min) and compound (5) (9.7 mg, Rt 26.5 min) by reverse HPLC (acetonitrile-water, 72:28) , flow rate 9 ml/min, UV detector set at 210 nm) yields compound (6) ( 18.3 mg, Rt 11.1 min).

实施例 2: 确定理化参数及化合物的结构  Example 2: Determining physical and chemical parameters and structure of compounds

化合物 （ 1 ) 为阜康阿魏 （ Fukanemarin) B(l): 黄色油。  Compound (1) is Fukanemarin B(l): yellow oil.

[_a]23_D±o。 （c 0.23, MeOH)。 UV (MeOH) _max(log ε) nm:310 (4.00)， 245 (3.90), 212 (4.37)。 IR (KBr) v_max cm.i:3408, 2974, 2935, 1767, 1707, 1616, 1512, 1444, 1379, 1277, 1249, 1195, 1159, 1110, 1026, 945, 838, 775 cnT Ή和 ^C-NMR数据见 表 1。 FABMS m/z 409 [M+H] +。 HRFABMS m/z 409.20130 [M+H]+ (计算为 C2₅H2₉0₅409.20147 )。 [ _a ] 23 _{D ±} o. (c 0.23, MeOH). UV (MeOH) _max (log ε) nm: 310 (4.00), 245 (3.90), 212 (4.37). IR (KBr) v _max cm.i: 3408, 2974, 2935, 1767, 1707, 1616, 1512, 1444, 1379, 1277, 1249, 1195, 1159, 1110, 1026, 945, 838, 775 cnT Ή and ^C - NMR data are shown in Table 1. FABMS m/z 409 [M+H] +. HRFABMS m/z 409.20130 [M+H]+ (calculated as C2 ₅ H2 ₉ 0 ₅ 409.20147 ).

化合物（ 2)为阜康阿魏 E(2): 无色油。 [a]²3_D- 2.0。（c 0.36, MeOH)。 UV (MeOH) _max(log ε) nm:332 (3.89), 317 (4.00), 288 (3.68), 244 (3.75), 225 (4.03), 207 (4.37). IR (KBr) v_max cm"¹:3429, 2970, 2929, 1717, 1638, 1614 1562, 1517, 1450, 1415, 1382, 1333, 1273, 1198， 1159, 1105, 1025, 983, 943, 843, 768 cm— i。 iH和 c-NMR数据见表 2和 3。EIMS m/z 408 [M]+。 HREIMS m/z 408.19387 [M]⁺ ( 计算为 C₂₅H2₈0₅ 408.19365 )。 The compound (2) is a ruthenium E (2): a colorless oil. [a] ² 3 _D - 2.0. (c 0.36, MeOH). UV (MeOH) _max (log ε) nm: 332 (3.89), 317 (4.00), 288 (3.68), 244 (3.75), 225 (4.03), 207 (4.37). IR (KBr) v _max cm" ¹ : 3429, 2970, 2929, 1717, 1638, 1614 1562, 1517, 1450, 1415, 1382, 1333, 1273, 1198, 1159, 1105, 1025, 983, 943, 843, 768 cm—i. iH and c-NMR The data are shown in Tables 2 and 3. EIMS m/z 408 [M] + HREIMS m/z 408.19387 [M] ⁺ (calculated as C ₂₅ H2 ₈ 0 ₅ 408.19365 ).

化合物（ 3 )为阜康阿魏 F(3):无色油。 [α]²³ο+41.7° (c0.14， MeOH)。 UV (MeOH) _max(log ε) nm:332 (3.89), 319 (4.09)， 289 (3.77), 244 (3.86), 225 (4.15)， 207 (4.45)。 IR (KBr) v_max cm'¹:3423, 2925, 1718， 1634, 1614, 1516, 1450， 1416, 1382， 1333， 1273, 1198, 1155, 1025, 984, 943， 841， 767 cm^-1。 The compound (3) is a ruthenium F (3): a colorless oil. [α] ²³ ο+41.7° (c0.14, MeOH). UV (MeOH) _max (log ε) nm: 332 (3.89), 319 (4.09), 289 (3.77), 244 (3.86), 225 (4.15), 207 (4.45). IR (KBr) v _max cm' ¹ : 3423, 2925, 1718, 1634, 1614, 1516, 1450, 1416, 1382, 1333, 1273, 1198, 1155, 1025, 984, 943, 841, 767 cm ^-1 .

和 13C-NMR数据见表 2和 3。 EIMS /23/Ζ408 [M] +。 HREIMS 408.19365 [M]+ (计算为 C25H28O5408.19365 )。 And 13C-NMR data are shown in Tables 2 and 3. EIMS /23/Ζ408 [M] +. HREIMS 408.19365 [M]+ (calculated as C25H28O5408.19365).

化合物（ 5 )为阜康阿魏 G(5) 无色油。 [a]23_D-8.9。（c0.18， MeOH)。 UV (MeOH) _max(log ε) nm: 325 (3.98), 316 (4.09), 288 (3.75), 244 (3.91)， 225 (4.18), 207 (4,40)。 IR (KBr) v_max cm"¹:3418, 2973, 2938, 1768, 1726, 1638, 1614, 1561, 1517, 1449, 1413, 1378， 1331， 1273, 1194, 1156, 1112， 1026,985, 948, 837, 762 cnT iH和 C- NMR数据见表 2和 3。EIMS m/z 408 [M] +。 HREIMS m/z 408.19365 [M]+ (计算为 C₂₅H₂80₅ 408.19365 )。 The compound (5) is a ruthenium G (5) colorless oil. [a] 23 _{D -} 8.9. (c0.18, MeOH). UV (MeOH) _max (log ε) nm: 325 (3.98), 316 (4.09), 288 (3.75), 244 (3.91), 225 (4.18), 207 (4,40). IR (KBr) v _max cm" ¹ : 3418, 2973, 2938, 1768, 1726, 1638, 1614, 1561, 1517, 1449, 1413, 1378, 1331, 1273, 1194, 1156, 1112, 1026,985, 948, 837, 762 cnT iH and C-NMR data are shown in Tables 2 and 3. EIMS m/z 408 [M] + HREIMS m/z 408.19365 [M]+ (calculated as C ₂₅ H ₂ 80 ₅ 408.19365 ).

表 1 化合物（1 )的 NMR 光谱数据 H-NMR - 30 0 M ， 13C-NMR- 75MH z , CDCl₃，TMS，S(ppm),c/=Hz)a Table 1 NMR spectrum data of the compound (1) H-NMR - 30 0 M , 13C-NMR- 75MH z , CDCl ₃ , TMS, S (ppm), c/= Hz)a

s ：一重线、 d: 二重线、 t : 三重线、 m: 多重线 s : one heavy line, d: double line, t : triple line, m: multiple line

a 赋值通过解偶联 iH-iHCOSY，HMQC,HMBC 和不同的 光谱证实。 0398 表 2化合物 （ 2) ， （ 3 ) ， （ 5) 的 iHNMR光谱数据（30 0 MH z , CDCl₃,TMS,6(ppm),_e/=Hz) ^ The assignment was confirmed by uncoupling iH-iHCOSY, HMQC, HMBC and different spectra. 0398 Table 2 iHNMR spectral data of compounds (2), (3), (5) (30 0 MH z , CDCl ₃ , TMS, 6 (ppm), _e /= Hz) ^

a 赋值通过解偶联 iH- iHCOSY'HMQC'HMBC 和不同的 NOE光谱证实。  The assignment was confirmed by uncoupling iH-iHCOSY'HMQC'HMBC and different NOE spectra.

b 重叠信号。 b Overlapping signals.

表 3化合物 （ 2) ， （ 3) ， （ 5) 的 ¹³CNMR光谱数据（75M , CDCl₃,TMS,S(ppm)) ^a Table ^{13 13} CNMR spectral data of compounds ( 2), ( 3) , ( 5) (75M , CDCl ₃ , TMS, S (ppm)) ^a

a 赋值通过解偶联

和不同的 光谱证实。 ^b 重叠信号。 阜康阿魏 B(l)呈黄色油状，根据快原子轰击质谱（ FABMS) 数据，分子量为 408,该数据显示在阴极振荡型的 m/z409( M+H) +处有质子化的分子离子吸收峰， 而在 m/z407 ( M-H) ⁺处有去 质子化的分子离子吸收峰。 这些数据， 连同 iH和 i³CNMR光 谱数据 （表 1), 提示分子化学式为 C₂₅H₂₈0₅， 这得到阳极振荡 型的 HRFABMS的支持 （ C2₅H280₅, m/z409.20147 )。 a assignment by uncoupling

And confirmed by different spectra. ^b overlapping signals. 阜康阿魏 B(l) is yellow oil. According to fast atom bombardment mass spectrometry (FABMS) data, the molecular weight is 408. This data shows protonated molecular ions at the cathode oscillation type m/z409(M+H)+. The peak is absorbed, and there is a deprotonated molecular ion absorption peak at m/z 407 (MH) ⁺ . These data, together with the iH and i ³ C NMR spectral data (Table 1), suggest that the molecular formula is C ₂₅ H ₂₈ 0 ₅ , which is supported by the anodic oscillation type HRFABMS (C2 ₅ H280 ₅ , m/z 409.20147 ).

化合物 （ 1 ) 的 iH NMR光谙数据证明 δ_Η7.62 ( 1H， d, J = 8.8Hz, H-5 )、 δ_Η6.81( 1H， dd, J= 8.8,2.6Hz, H-6 )、 δ_Η6.76 ( 1H, d, J= 2.6Hz, H-8) 处存在 1， 2， 4-三取代苯环， 在 δ_Η3.85处有一个甲氧基团， 以及根据 iH- iH COSY, HMQCH 和 HMBC光谱的相关性确定的以倍半萜烯单位为特征的其它信 号的存在， 化合物 （ 1) 其余的 iH和 isCNMR数据除了这些倍 半萜烯单位外， 还表明一个 7-氧-取代 香豆素（ 7-氧杂萘邻酮） 化合物的存在。 δ_Η3.85( OMe )与 δ_Η6.81( Η-6 )及 δ_Η6.76 (H-8) 在 NOE实验的微分中的相关性提示甲氧基团连接于 07。 The iH NMR pupil data of the compound (1) proved that δ _Η 7.62 ( 1H, d, J = 8.8 Hz, H-5 ), δ _Η 6.81 ( 1H, dd, J = 8.8, 2.6 Hz, H-6 ), δ _Η 6.76 ( 1H, d, J = 2.6Hz, H-8) There is a 1, 2, 4-trisubstituted benzene ring, a methoxy group at δ _Η 3.85, and according to iH- iH COSY, HMQCH and The correlation of HMBC spectra determines the presence of other signals characterized by sesquiterpene units. Compound (1) The remaining iH and isCNMR data, in addition to these sesquiterpene units, also indicate a 7-oxo-substituted coumaric The presence of a compound (7-oxaphthalenone). The correlation between δ _Η 3.85 ( OMe ) and δ _Η 6.81 ( Η -6 ) and δ _Η 6.76 (H-8) in the differential of the NOE experiment suggests that the methoxy group is attached to 07.

在化合物 （ 1) 的 HMBC光 i普中， δ_Η3.79 ( H-1' ) 与 C-2、 C—3、 C-4、 C— 2'和 C-3'， δ_Η1.36( H— 15')与 C-3、 C— Γ和 C— 2'， 以及 δ_Η7.71 ( OH) 与 C- 3、 C-4和 C-10的相关性证明倍半萜 烯基团与香豆素单位的 C-3连接，并且有一个羟基与 C- 4相连。 C- 3和 C- 4的化学移位与来自 F.pallida.¹⁶⁾的相似化合物的化 学移位的报道非常接近。 此外， δ_Η3.29 (Η-7') 与 C-5'、 C- 6'、 C- 8'及 C-9'， δ_Η5.91 (Η-9') 与 C-8'、 C- 10'及 Oil', δ_Η7.07 ( Η-ΙΙ') 与 C_8'、 C-9'及 C-10', δ_Η9.8 ( Η-12') 与 C—9'、 C-10'及 C-ll'的 HMBC相关性证实化合物 （ 1) 有 7-取代- 4- 甲基- 2-呋喃基体系。 倍半萜烯单位的两个双键结构象是根据 NOE试验的微分确定的。 化合物 （ 1) 显示了 H- 4'和 H- 13'及 H-14', H- 3'和 H-5'、 H- 5'和 H- 7'之间显著的 NOE相关性， 表明化合物 （ 1) 中倍半萜烯单位的两个双键具有 E构型。 这样 化合物 （ 1 ) 是 4-羟基- 7-曱氧基 -3-[1, 2， 6-三甲基 - 7- ( 4_ 甲基 -2-呋喃） -七氯 -2 ( E)， 5 ( E) -二烯] -香豆素。 In the HMBC luminescence of the compound (1), δ _Η 3.79 ( H-1 ' ) and C-2, C-3, C-4, C-2' and C-3', δ _Η 1.36 (H- 15') and C-3, C-Γ and C-2', and the correlation of δ _Η 7.71 ( OH) with C-3, C-4 and C-10 demonstrates sesquiterpene group and coumarin The unit is C-3 linked and has a hydroxyl group attached to C-4. The chemical shifts of C-3 and C-4 are very close to those reported by similar compounds from F. pallida. ¹⁶⁾ . In addition, δ _Η 3.29 (Η-7') and C-5', C-6', C-8' and C-9', δ _Η 5.91 (Η-9') and C-8', C-10 'And Oil', δ _Η 7.07 (Η-ΙΙ') with C_8', C-9' and C-10', δ _Η 9.8 (Η-12') and C-9', C-10' and C- The HMBC correlation of ll' confirms that the compound (1) has a 7-substituted-4-methyl-2-furanyl system. The two double bond structures of the sesquiterpene unit are determined according to the differential of the NOE test. Compound (1) shows H-4' and H-13' and Significant NOE correlation between H-14', H-3' and H-5', H-5' and H-7', indicating that the two double bonds of the sesquiterpene unit in compound (1) have E structure. Thus, the compound (1) is 4-hydroxy-7-methoxy-3-[1,2,6-trimethyl-7-(4-methyl-2-furan)-heptachloro-2(E), 5 (E) -Diene] - Coumarin.

得到的阜康阿魏 E(2)是无色油状。 化合物 （ 2) - ( 5) 的质 谱数据表明有同样的分子离子 （见实验部分）， 这表示同分异构 化合物的存在。  The obtained 阜康阿魏 E(2) is a colorless oil. The mass spectral data of compounds (2) - (5) indicate the same molecular ion (see experimental section), which indicates the presence of isomeric compounds.

化合物 （ 2) 的 iH和 CNMR光谱类似于 2, 3-二氢 -7- 羟基 -2S*,3R*-二甲基 -3-[4 -曱基 -5- 4-曱基- 2-呋喃） -3 ( E) -戊烯基]-呋喃 [3,2- c]香豆素，除了 C-7存在的曱氧基（ δ_Η3.86; δ_Η55.5 ) 以夕卜。 δ_Η3.86 ( OMe ) 与 δ_Η6.82 ( Η - 6 和 Η- 8 ) 在 ΝΟΕ实验微分中的相关性提示曱氧基团连接于 C- 7。 The iH and CNMR spectra of the compound (2) are similar to 2,3-dihydro-7-hydroxy-2S*,3R*-dimethyl-3-[4-indolyl-5-4-indole-2-furan -3 (E)-pentenyl]-furan [3,2-c]coumarin, in addition to the decyloxy group present in C-7 (δ _Η 3.86; δ _Η 55.5 ). The correlation between δ _Η 3.86 ( OMe ) and δ _Η 6.82 ( Η - 6 and Η - 8 ) in the experimental differentiation of ΝΟΕ suggests that the oxime group is attached to C-7.

在化合物 （ 2) 的 HMBC光 i瞽中， δ_Η1·44 ( 2-Me ) 与 C- 2 和 C— 3， 以及 δ_Η1.28 ( 3-Me ) 与 C— 1'、 C— 2、 C—3和 C— 3a的 相关性提示 C- Γ与 C-3相连。 C-2通过醚键与 C-9b相邻是根 据 C- 2 (8_C89.3)和 C-9b ( Scl64.9)化学移位的不饱和值推断 的 i³⁾。 C-2 和 C-3 处二甲基二氢呋喃成分的相对构型是根据 NOE实验微分确定的。化合物（ 2)显示了 2- Me和 3- Me之间、 H-2和 H- Γ之间显著的 NOE相关性， 表明 2-Me和 3- Me (曱 基） 之间的顺式关系。 根据 4'- Me 的化学移位 （其相对向高磁 场移位 （ 8cl5.9) )， ¹⁷⁾倍半萜浠单位双键的构型被命名为 4Έ。 这样 2被确定为 2，3-二氢- 7-曱氧基 -2S*，3R*-二曱基 - 3- [4-曱 基 -5-4-曱基 -2-呋喃） -3 ( E) -戊烯基] -呋喃 [3，2-c]香豆素。 In the HMBC photoinhibition of compound (2), δ _Η 1·44 ( 2-Me ) and C-2 and C-3, and δ _Η 1.28 ( 3-Me ) and C-1 ', C-2 The correlation between C-3 and C-3a suggests that C-Γ is linked to C-3. The C-2 adjacent to C-9b through the ether bond is inferred from the unsaturated value of the chemical shift of C-2 (8 _C 89.3) and C-9b (Scl64.9 ⁾ . The relative configuration of the dimethyl dihydrofuran component at C-2 and C-3 was determined based on the NOE experimental differential. Compound (2) shows a significant NOE correlation between 2-Me and 3-Me, H-2 and H- ,, indicating a cis relationship between 2-Me and 3-Me (thiol). According to the chemical shift of 4'-Me (which is relatively high magnetic field shift (8cl5.9)), ¹⁷⁾ the configuration of the sesquiterpene unit double bond is named 4Έ. Thus 2 was identified as 2,3-dihydro-7-nonyloxy-2S*,3R*-dimercapto-3-[4-indolyl-5-4-mercapto-2-furan)-3 ( E) -pentenyl]-furan [3,2-c] coumarin.

阜康阿魏 F(3)是无色油状。 HMBC 实验提示化合物 （ 3) 的结构与化合物 （ 2) 类似。 不过， 化合物 ( 3 ) 的 NMR光谱与 化合物（ 2 )稍有不同，特别是在 3- Me( 3为 Sc23.3, 2为 S_c19.1 ) 和 C- 2 ( 3为 Sc92.7， 2为 Sc89.3 ) 处， 提示化合物 （ 3) 可能 是化合物 （ 2)在手性中心 C-2和 C-3处的非对映异构体。 C-2 和 C- 3处二曱基二氢呋喃成分的相对构型是根据 NOE实验微分 确定的。 化合物 （ 3) 显示了 2-Me和 H-1'之间、 3_Me和 H-2 之间显著的 NOE相关性，表明 2-Me和 3-Me之间的反式关系。 因此，化合物（ 3 )是 2,3-二氢-7-甲氧基-28*，31 *-二曱基-3-[4- 甲基- 5- 4-曱基 -2-呋喃） -3 ( E) -戊烯基] -呋喃 [3,2- c]香豆素。 Fukang Awei F (3) is a colorless oil. The HMBC experiment suggests that the structure of compound (3) is similar to compound (2). However, the NMR spectrum of the compound (3) (2) is slightly different, especially in the 3- Me (3 to Sc23.3, 2 to S _c 19.1) And C-2 (3 is Sc92.7, 2 is Sc89.3), suggesting that compound (3) may be a diastereomer of compound (2) at the chiral centers C-2 and C-3. The relative configuration of the dimercaptodihydrofuran component at C-2 and C-3 is determined by NOE experimental differentiation. Compound (3) showed a significant NOE correlation between 2-Me and H-1', between 3_Me and H-2, indicating a trans relationship between 2-Me and 3-Me. Thus, the compound (3) is 2,3-dihydro-7-methoxy-28*,31*-dimercapto-3-[4-methyl-5- 4-mercapto-2-furan- 3 (E)-pentenyl]-furan [3,2-c]coumarin.

阜康阿魏 G(5)是无色油状。 HMBC 实验提示化合物 （ 5) 的结构与化合物 （ 4) 类似。 不过， 化合物 （ 5) 的 NMR光谱与 化合物 （ 4)稍有不同， 特别是在 C-1'和 2- Me处， 表明化合物 ( 5) 可能是化合物 （ 4)在手性中心 C-2和 C-3处的非对映异 构体。 C-2 和 C-3 处二曱基二氢呋喃成分的相对构型是根据 NOE实验微分确定的。 化合物 （ 5)显示了 2- Me和 H-1'之间、 3-Me和 H-2之间显著的 NOE相关性， 表明 2和 3_Me之间的 反式关系。 因此， 化合物 （ 5)是 2,3-二氢- 7-曱氧基 -2S*，3R*- 二曱基 -2-[4-曱基- 5- 4-曱基- 2-呋喃] -3 ( E ) -戊烯基] -呋喃 [3，2- c]香豆素。  Ji Kang A Wei G (5) is a colorless oil. The HMBC experiment suggests that the structure of the compound (5) is similar to that of the compound (4). However, the NMR spectrum of the compound (5) is slightly different from the compound (4), especially at C-1' and 2-Me, indicating that the compound (5) may be the compound (4) at the chiral center C-2 and Diastereomer at C-3. The relative configuration of the dimercaptodihydrofuran component at C-2 and C-3 is determined by NOE experimental differentiation. Compound (5) shows a significant NOE correlation between 2-Me and H-1', 3-Me and H-2, indicating a trans relationship between 2 and 3_Me. Therefore, the compound (5) is 2,3-dihydro-7-decyloxy-2S*,3R*-dimercapto-2-[4-indolyl-5-4-indolyl-2-furan]- 3 ( E ) -pentenyl] -furan [3,2- c]coumarin.

化合物 （ 4) 和 （ 6) 是已知化合物， 其结构通过与文献报 道比较得以阐明。 化合物的结构式确定如下：  Compounds (4) and (6) are known compounds, and their structures are clarified by comparison with literature reports. The structural formula of the compound is determined as follows:

化合物（1) 化合物（2) Compound (1) Compound (2)

化合物（5) 化合物（6)  Compound (5) Compound (6)

实施例 3 : 抑制 NO生成试验  Example 3: Inhibition of NO production test

由于干扰素 r 以及脂多糖刺激而产生的巨噬细胞对 NO 生 成的阻碍效果 （抑制 NO 生成的效果） 通过下面的实验方法求 得。并且根据阻碍效果的 IC50(um)来评价 NO生成的抑制效杲。  The inhibitory effect of macrophages on NO production by interferon-r and lipopolysaccharide stimulation (the effect of inhibiting NO production) was obtained by the following experimental method. Further, the inhibitory effect of NO production was evaluated based on the IC50 (um) of the barrier effect.

所用材料：  Materials used:

RAW 264.7细胞 （大日本制药）  RAW 264.7 cells (Greater Japan Pharmaceuticals)

N-1-盐酸蔡乙二胺 （ lg 和光纯药）  N-1-hydrochloric acid, Cai ethylene diamine ( lg and pure medicine)

磺胺 （ 500g 和光纯药）  Sulfonamide (500g and pure medicine)

Ham's F12 培养基 （ SIGMA N488 500mL )  Ham's F12 Medium (SIGMA N488 500mL)

IFN-γ ( Geneyme/Techne lOO g )  IFN-γ (Geneyme/Techne lOO g )

月旨多糖 （ LPS , 055 : B5 lOmg, Sigma )  Moon Polysaccharide ( LPS , 055 : B5 lOmg , Sigma )

磷酸 （ 500ml 和光纯药）  Phosphoric acid (500ml and pure medicine)

DMSO ( 500ml 和光纯药）  DMSO (500ml and pure medicine)

96孔微滴定板 （ 50/盒 住友电木，商品名 〔8096R〕） 试-险方法：  96-well microtiter plate (50/box Sumitomo Bakelite, trade name [8096R]) Trial-risk method:

RAW264.7 细胞， 是在 5%的 C02、 37°C的温度、 含有 10%FBS的 Ham's F12的培养基中培育出来的。 将处在对数生 长期的 RAW264.7 细胞浓度调整为 1.2X10⁶个 / ml, 将细胞在 C02恒温箱中培养 2 小时后。 添加了各种浓度的萃取物或离析 成分。 同时添加 LPS ( 100ng/ml), INF-γ ( lOU/ml), 样品 0.4 L。 在 C02 恒温箱中培养一段时间后， 取培养上清， 将 RAW264.7 细胞的培养上清 lOO L 放进平底 96 孔板中， 用 Griess法定量 NO的氧化体 N0₂-。 使 DMSO的浓度在培养基 中成为 0.2%以下， 加入 0.1%萘二胺溶液 50 L、 1%对氨基苯碩 酰胺溶液 50 L， 室温中避光放置 10分钟。 用分光光度计测定 570nm (对照 655nm ) 的吸光度 O.D.。 根据 NaN0₂标准溶液 作成的检量线， 使用的是亚硝酸钠溶液（ 100, 50, 20, 10, 5, 2, 1, ΟμΜ), 定量培养基中的 Ν0₂-。 RAW264.7 cells, at 5% CO2, 37 ° C temperature, containing 10% FBS was cultured in the medium of Ham's F12. The concentration of RAW264.7 cells in the logarithmic growth phase was adjusted to 1.2× ^{10 6} cells/ml, and the cells were cultured in a CO 2 incubator for 2 hours. Various concentrations of extract or segregation components were added. Add LPS (100 ng/ml), INF-γ (lOU/ml), and 0.4 L of sample. After culturing for a while in a C02 incubator, the culture supernatant was taken, and the culture supernatant of RAW264.7 cells was placed in a flat-bottom 96-well plate, and the NO oxidant N0 ₂ - was quantified by the Griess method. The concentration of DMSO was made 0.2% or less in the medium, and 50 L of a 0.1% naphthalene diamine solution and 50 L of a 1% p-aminobenzene amide solution were added, and the mixture was allowed to stand in the dark for 10 minutes at room temperature. The absorbance OD at 570 nm (control 655 nm) was measured with a spectrophotometer. According to the calibration curve prepared from the NaN0 ₂ standard solution, sodium nitrite solution (100, 50, 20, 10, 5, 2, 1, ΟμΜ) was used to quantify Ν0 ₂ - in the medium.

细胞毒性通过 ΜΤΤ法、 由镜检确定。 ΜΤΤ法是已知的常 规方法。 即， 于 96孔微滴定板,每孔 200μ1细胞浓度为 l.OxlO⁵ 细胞 /ml， 添加不同浓度的提取物或单体组分， 将细胞培养 16 小时， 加入 MTT 试剂 ， 再培养 4 小时。 弃上清， 添加 150pLDMSO, 将生成的曱臜 （ formazane) 完全溶解， 测定 570nm下的吸光度。 Cytotoxicity was determined by sputum method and by microscopic examination. The method is a known conventional method. That is, in a 96-well microtiter plate, the cell concentration of 200 μl per well was 1.0×× ¹⁰ cells/ml, and different concentrations of extract or monomer components were added, and the cells were cultured for 16 hours, added with MTT reagent, and cultured for further 4 hours. The supernatant was discarded, 150 pL of DMSO was added, and the resulting formazane was completely dissolved, and the absorbance at 570 nm was measured.

活性评价  Activity evaluation

计算出 N0₂—的量， 代入下面的公式求出抑制效果 Calculate the amount of N0 ₂ — and substitute the following formula to find the suppression effect.

抑制率 （％) ={1- (X-Y) / ( Z-Y ) }X 100  Inhibition rate (%) = {1- (X-Y) / ( Z-Y ) }X 100

X： 在实验化合物存在下， 由 IFN-γ 和 LPS 诱导产生的 N0₂—的量， X: the amount of N0 ₂ - induced by IFN-γ and LPS in the presence of the test compound,

Υ; 在实验化合物、 IFN- γ和 LPS均不存在的情况下， 诱 导产生的 ΝΟ 的量，  Υ; the amount of ruthenium induced by the test compound, IFN-γ, and LPS,

Z: IFN- γ和 LPS诱导产生的 N0₂—的量。 化合物 （ 1 ) 的 NO生成抑制效果的 IC₅o如下所示： 当 RAW264.7与 LPS和 IFN-γ温育时， NO生成急剧上升。 这些化合物表现出抑制活性 （ 1 : ICso=30.2±1.7 μΜ ; 2 : IC₅₀=29.0士 1.0 μΜ; 3: IC₅₀=30.7±0.9 μΜ; 4: IC₅o=27.8±4.6 μΜ; 5: IC₅o=27.3±2.3 μΜ; 6: IC₅o=87.5±11.7 μΜ; n=3 )。 利用 MTT 测定法测量了这些化合物的细胞毒性效果。 在 LPS/IFN-Y处理 24小时后， 这些化合物 （ 3-100 μΜ ) 不表现 处任何显著的细胞毒性。 通过比较这些化合物的抑制活性， 我 们鉴定了可能影响活性水平的有意思的特点。 4的抑制活性比 6 强， 提示曱氧基团增强倍半萜烯香豆素的抑制活性。 NO是被称 之为 NOS的家族酶合成， 其在 NO的产生中利用精氨酸作为底 物。 其活性被细胞因子以及通过细胞暴露于免疫和炎性刺激物 在转录水平调控。 化合物 （ 1 )、 （ 2 ) 和 （ 4 ) 以剂量依赖性的方 式抑制 iNOS mRNA的表达 （图 1 )。 Z: The amount of N0 ₂ - induced by IFN-γ and LPS. The IC ₅ o of the NO production inhibitory effect of the compound (1) is as follows: When RAW264.7 is incubated with LPS and IFN-γ, NO production sharply rises. These compounds exhibited inhibitory activity (1: ICso = 30.2 ± 1.7 μΜ; 2: IC ₅₀ = 29.0 ± 1.0 μΜ; 3: IC ₅₀ = 30.7 ± 0.9 μΜ; 4: IC ₅ o = 27.8 ± 4.6 μΜ; 5: IC ₅ o = 27.3 ± 2.3 μΜ; 6: IC ₅ o = 87.5 ± 11.7 μΜ; n = 3). The cytotoxic effects of these compounds were measured by MTT assay. These compounds (3-100 μΜ) did not exhibit any significant cytotoxicity after 24 hours of LPS/IFN-Y treatment. By comparing the inhibitory activities of these compounds, we identified interesting features that may affect the level of activity. The inhibitory activity of 4 is stronger than that of 6, suggesting that the oxime group enhances the inhibitory activity of sesquiterpene coumarin. NO is a family enzyme synthesis called NOS, which utilizes arginine as a substrate in the production of NO. Its activity is regulated at the transcriptional level by cytokines as well as by exposure of cells to immune and inflammatory stimuli. Compounds (1), (2) and (4) inhibited the expression of iNOS mRNA in a dose-dependent manner (Fig. 1).

实施例 4: iNOS mRNA的反转录聚合酶链式反应分析 RAW264.7 细胞以 1.2X 10⁶细胞 /ml 在 96 孔平底板中于 37°C培养 2 小时， 然后将测试化合物与 LPS (100 ng/ml)和 IFN-γ (0.33 ng/ml)同时添加到培养物中。细胞于 37°C温育大约 8小时。 利用 RNA分离试剂盒 （ QIAGEN, Hilden, Germany ) 从细胞沉淀中分离总 RNA。 通过寡聚（ dT ) 12- 18引物将总 RNA ( 250 ng ) 反转录为 cDNA。 PCR样品含 30 μΐ反应混合物， 由 50 mM KC1、 5 mM MgCl₂、 0.2 mM dNTP、 0.6单位 Ampli Taq GOLD (Applied Biosystems, CA， U.S. A)和 0.4 μπιοΐ有义 和 反 义 引 物 组 成 。 iNOS 的 正 义 引 物 为 5'-ACCTACTTCCTGGACATTACGACCC-3'， 而反义引物为 5'-AAGGGAGCAATGCCCGTACCAGGCC-3'₀ 甘油醛 -3-磷酸 脱 氢 酶 （ GAPDH ) 的 正 义 引 物 为 5'-ACCACAGTCCATGCCATCAC-3' , 而 反 义 引 物 为 5'-TCCACCACCCTGTTGCTGTA-3'„ PCR反应在如下条件下 进行： 25个循环的 94°C变性 1分钟、 60°C退火 1分钟和 72°C 延伸 1.5分钟， 利用热循环仪 （ GeneAmp PCR Systems 9770; PE Applied Biosystems, U.S.A)。 PCR产物在 2%琼脂糖凝胶 上跑样， 并通过溴乙锭染色观察。 然后对凝胶中的奈带进行照 相。 Example 4: Reverse transcription polymerase chain reaction analysis of iNOS mRNA RAW264.7 cells were cultured at 1.2X 10 ⁶ cells/ml in a 96-well flat bottom plate at 37 ° C for 2 hours, and then the test compound was combined with LPS (100). Ng/ml) and IFN-γ (0.33 ng/ml) were simultaneously added to the culture. The cells were incubated at 37 ° C for approximately 8 hours. Total RNA was isolated from the cell pellet using an RNA isolation kit (QIAGEN, Hilden, Germany). Total RNA (250 ng) was reverse transcribed into cDNA by oligo (dT) 12-18 primers. The PCR sample contained 30 μΐ of the reaction mixture consisting of 50 mM KCl, 5 mM MgCl ₂ , 0.2 mM dNTP, 0.6 units of Ampli Taq GOLD (Applied Biosystems, CA, US A) and 0.4 μπιοΐ sense and antisense primers. The sense primer for iNOS is 5'-ACCTACTTCCTGGACATTACGACCC-3', and the antisense primer is 5'-AAGGGAGCAATGCCCGTACCAGGCC-3' ₀ glyceraldehyde-3-phosphate The sense primer for dehydrogenase (GAPDH) is 5'-ACCACAGTCCATGCCATCAC-3', and the antisense primer is 5'-TCCACCACCCTGTTGCTGTA-3' „ PCR reaction is carried out under the following conditions: 25 cycles of denaturation at 94 ° C for 1 minute, Annealing at 60 ° C for 1 minute and 72 ° C for 1.5 minutes, using a thermal cycler (GeneAmp PCR Systems 9770; PE Applied Biosystems, USA). The PCR product was run on a 2% agarose gel and stained with ethidium bromide. Observation. The nep in the gel was then photographed.

发现 COX- 2，IL- 1 β ,IL-6,TNF-a的 mRNA抑制作用 （图 It was found that mRNA inhibition of COX-2, IL-1β, IL-6, and TNF-a (Fig.

1 ) 1 )

FFE (阜康阿魏） 中的化合物、 IFN- γ(10 U/mL)及 LPS ( 100ng/mL) 共存的条件下， 将 RAW264.7 细胞 （ 1.2x 106 细胞 /mL)培养 6小时。 RNA的提取及 RT- PCR的条件与 6相 同。 只是 COX-2 的引物中使用了 5'-TCAAAAGAAGTGCTG GAAAAGGTT-3'[a] 、 5'-TCTACCTGAGTGTCTTTGACT GTG-3'[a] , IL-1 β 的 引 物 中 使用 5'-TGAAGGGCTGCT TCCAAACCTTTGACC-3'ts] 、 5'-TGTCCATTGAGGTGG AGAGCTTTCAGC-3'[a], IL-6 的 引 物 中 使 用 5'— TGG AGTCACAGAAGGAGTGGCTAAG-3'[s] 5'-TCTGACCAC AGTGAGGAATGTCCAC-3'[a], TNF- a 的 引 物 中 使 用 5'_CAGCCTCTTCTCATTCCTGCTTG-3'[s] 5'-GTCTTT GAGATCCATGCCGTTG-3'[a] o  RAW264.7 cells (1.2 x 106 cells/mL) were cultured for 6 hours in the presence of a compound, IFN-γ (10 U/mL) and LPS (100 ng/mL) in FFE (阜康阿魏). The conditions for RNA extraction and RT-PCR were the same as those for 6. Only 5'-TCAAAAGAAGTGCTG GAAAAGGTT-3'[a], 5'-TCTACCTGAGTGTCTTTGACT GTG-3'[a] was used as a primer for COX-2, and 5'-TGAAGGGCTGCT TCCAAACCTTTGACC-3'ts was used for the primer of IL-1 β. ], 5'-TGTCCATTGAGGTGG AGAGCTTTCAGC-3'[a], IL-6 primer used 5'- TGG AGTCACAGAAGGAGTGGCTAAG-3'[s] 5'-TCTGACCAC AGTGAGGAATGTCCAC-3'[a], primer for TNF-a Use 5'_CAGCCTCTTCTCATTCCTGCTTG-3'[s] 5'-GTCTTT GAGATCCATGCCGTTG-3'[a] o

如图 1浓度依赖的抑制已被认定。 实施例 5 Concentration-dependent inhibition as shown in Figure 1 has been identified. Example 5

取实施例 1中的氯仿提取物 70g， 用硅胶层析柱（ Wako gel 0200、 和光纯药制、 6Φ X 17cm)进行分离， 洗脱剂为正乙烷： 乙酸乙酯系， 依次以浓度为 0%、 2%、 5%、 10%、 20%、 40%、 60%、 100。/。的乙 酸乙酯 3L洗脱， 各分离出 200ml的馏分 1~ 11， 其中使用 10%浓度洗 脱时分别收集 1升和 2升洗脱液的馏分， 类似的 20%、 40%浓度也收集 2个馏分。  70 g of the chloroform extract of Example 1 was separated and separated on a silica gel column (wako gel 0200, manufactured by Wako Pure Chemical Industries, Ltd., 6 Φ X 17 cm). The eluent was n-ethane: ethyl acetate, and the concentration was 0%, 2%, 5%, 10%, 20%, 40%, 60%, 100. /. The ethyl acetate 3L was eluted, and each of the 200 ml fractions 1 to 11 was separated, and the fractions of 1 liter and 2 liters of the eluate were separately collected at a concentration of 10%, and similar concentrations of 20% and 40% were also collected. Distillate.

实施例 6  Example 6

从实施例 5中取得的 1~11馏分中， 对馏分 6, 将其减压浓缩物用硅 胶层析柱（4φ X 23cm)进行分离， 洗脱剂为正乙烷： 乙酸乙酯系， 依 次以浓度为 0%、 2%、 5%、 10%、 20%、 40%、 60%、 100%的乙酸 乙酯 3L洗脱， 各分离出 200ml的馏分 1~ 11, 其中使用 10%浓度洗脱 时分别收集 1升和 2升洗脱液的馏分， 类似的 20%、 40%浓度也收集 2 个馏分。  From the 1 to 11 fraction obtained in Example 5, the fraction 6 was subjected to separation on a silica gel column (4 φ X 23 cm), and the eluent was n-ethane: ethyl acetate. Eluate with a concentration of 0%, 2%, 5%, 10%, 20%, 40%, 60%, 100% ethyl acetate 3L, and separate 200 ml fractions 1 to 11, each using a 10% concentration wash Fractions of 1 liter and 2 liters of eluate were separately collected at the time of desorption, and 2 fractions were collected at similar concentrations of 20% and 40%.

实施例 7  Example 7

对实施例 6中利用硅胶层析柱得到的洗脱液组分 6的，用正相 HPLC 对其进行分离。使用 YMC-PAK SIL-06层析柱（ 20 φ χ 150mm )( YMC 公司制）， 流动相使用 70%正己烷（正己烷： 乙酸乙酯 =70: 30), 流速 8.0ml/min进行洗脱，利用 UV(254nm)分离出在保留时间 21分 18秒洗 脱峰。  The fraction 6 of the eluate obtained in the silica gel column of Example 6 was separated by normal phase HPLC. A YMC-PAK SIL-06 column (20 φ χ 150 mm) (manufactured by YMC Corporation) was used, and the mobile phase was eluted with 70% n-hexane (n-hexane: ethyl acetate = 70: 30) at a flow rate of 8.0 ml/min. The elution peak at a retention time of 21 minutes and 18 seconds was separated by UV (254 nm).

实施例 8  Example 8

将实施例 7中利用正相 HPLC得到的、 21分 18秒洗脱峰， 再利用 反相 HPLC进行分离。 使用 YMC-PAK ProCl8 (10φ χ 150mm)层 析柱， 流动相使用 56%乙腈（乙腈： 水 =56: 44), 流速 3.0ml/min (室 温）进行洗脱。  The elution peak of 21 minutes and 18 seconds obtained by normal phase HPLC in Example 7 was separated and subjected to reverse phase HPLC. A YMC-PAK ProCl8 (10φ χ 150mm) chromatography column was used, and the mobile phase was eluted using 56% acetonitrile (acetonitrile: water = 56: 44) at a flow rate of 3.0 ml/min (room temperature).

将保留时间为 12分 42秒、 16分 22秒的洗脱馏分进行正相 HPLC 分离。 Normal phase HPLC was performed on the eluted fraction with a retention time of 12 minutes, 42 seconds, and 16 minutes and 22 seconds. Separation.

对保留时间 12分 42秒洗脱馏分， 使用 YMC-PAK SIL-06层析柱 ( 10Φ X 150mm), 流动相使用 68%正己烷（正己烷： 乙酸乙酯 =68: 32), 流速 3.0ml/min (室温）进行洗脱， 利用 UV(254nm)分离出在保 留时间 15分 57秒洗脱峰， 得到化合物 （ 8 )。  The fraction was eluted for a retention time of 12 minutes and 42 seconds, using a YMC-PAK SIL-06 column (10 Φ X 150 mm), and a mobile phase using 68% n-hexane (n-hexane: ethyl acetate = 68:32) at a flow rate of 3.0 ml. Elution was carried out at /min (room temperature), and a peak eluted at a retention time of 15 minutes and 57 seconds was separated by UV (254 nm) to give a compound (8).

对保留时间 16分 22秒洗脱馏分， 使用 YMC-PAK SIL-06层析柱 ( 10Φ X 150mm), 流动相使用 70%正己烷（正己烷： 乙酸乙酯 =70: 30)， 流速 3.0ml/min (室温）进行洗脱， 利用 UV(254nm)分离出在保 留时间 15分 39秒洗脱峰， 得到化合物 （7)。  The fraction was eluted for a retention time of 16 minutes and 22 seconds, using a YMC-PAK SIL-06 column (10 Φ X 150 mm), and the mobile phase was 70% n-hexane (n-hexane: ethyl acetate = 70: 30) at a flow rate of 3.0 ml. Elution was carried out at /min (room temperature), and a peak eluted at a retention time of 15 minutes and 39 seconds was separated by UV (254 nm) to obtain a compound (7).

实施例 9  Example 9

在实施例 5中利用硅胶层析柱得到的 1~11中， 合并其中的馏分 8 和 9, 取其减压浓缩物， 用硅胶层析柱（3φχ20αη)进行分离， 洗脱剂 为正乙烷： 乙酸乙酯系， 以乙酸乙酯浓度为 0%、 2%、 5%、 10%、 20 %、 40%、 60%、 100%的洗脱剂 3L依次洗脱， 得出各为 200ml的组 分 1 ~ 10，其中使用 40%浓度洗脱时分别收集 1升和 2升洗脱液的组分， 类似的 60 %浓度也收集 2个组分。用反相 HPLC对组分 6进行分离。使 用 YMC- PAK Pro C18 (20φ χ 150mm)层析柱、 流动相使用 56%乙 氰（乙氰： 水 = 56: 44), 以 9.0ml/min (室温）的流速洗脱， 利用 UV ( 210nm )分离出在保留时间 13分 42秒洗脱的峰（peak)。 该保留时 间 13分 42秒洗脱的馏分，用正相 HPLC进一步分离。使用 YMC-PAK SIL-06层析柱（ 10 Φ X 150mm )， 流动相使用 48%正己烷（正己烷： 乙 酸乙酯 =48: 52), 流速 3.0ml/min (室温）进行洗脱， 利用 UV(254nm) 分离出在保留时间 15分 24秒洗脱峰， 得到化合物 （9)。  In Example 1 using the silica gel column, 1 to 11 was combined, and the fractions 8 and 9 were combined, and the reduced-concentration was taken, and separated by a silica gel column (3φχ20αη), and the eluent was n-hexane. : Ethyl acetate, eluted with 3L of ethyl acetate at 0%, 2%, 5%, 10%, 20%, 40%, 60%, 100% eluent to give 200ml each. Components 1 to 10, in which 1 liter and 2 liters of eluent components were separately collected using a 40% concentration elution, and 2 components were also collected at a similar 60% concentration. Fraction 6 was isolated by reverse phase HPLC. Using a YMC-PAK Pro C18 (20φ χ 150mm) column, the mobile phase was eluted with 56% acetonitrile (acetonitrile: water = 56: 44) at a flow rate of 9.0 ml/min (room temperature) using UV (210 nm) The peak eluted at a retention time of 13 minutes and 42 seconds was separated. The fraction eluted at 13 minutes and 42 seconds was further separated by normal phase HPLC. The YMC-PAK SIL-06 column (10 Φ X 150 mm) was used, and the mobile phase was eluted with 48% n-hexane (n-hexane: ethyl acetate = 48: 52) at a flow rate of 3.0 ml/min (room temperature). UV (254 nm) separated the elution peak at a retention time of 15 minutes and 24 seconds to obtain a compound (9).

实施例 10  Example 10

在实施例 5中得到的 1~11馏分中， 对馏分 3, 将其减压浓缩物， 用 硅胶层析柱（5.5cl xl3cm)进行分离， 洗脱剂为正乙烷： 乙酸乙酯系， 以乙酸乙酯浓度为 0%、 2%、 5%、 10%、 20%、 40%、 60%、 100% 的洗脱剂依次洗脱， 各得 200ml组分 1~ 7， 其中使用 0%、 2%浓度洗 脱时收集的组分合为一个。 In the fractions 1 to 11 obtained in Example 5, the fraction 3 was subjected to a reduced pressure concentrate and separated on a silica gel column (5.5cl x 13 cm). The eluent was n-hexane: ethyl acetate. Eluent eluted with ethyl acetate at 0%, 2%, 5%, 10%, 20%, 40%, 60%, 100%, each obtained 200ml of components 1~7, of which 0% was used. The components collected at the 2% concentration elution were combined into one.

得到的组分中，对组分 3利用反相 HPLC进行分离。使用 YMC-PAK Pro C18 (20Φ X 150mm)层析柱、 流动相使用 85%乙氰（乙氰： 水= 85: 15), 以 9.0ml/min (室温）的流速洗脱， 利用 UV ( 210nm )分离 出在保留时间 19分 58秒和 21分 23秒洗脱峰。  Of the obtained components, component 3 was separated by reverse phase HPLC. Using a YMC-PAK Pro C18 (20Φ X 150mm) column, the mobile phase was eluted with 85% acetonitrile (acetonitrile: water = 85: 15) at a flow rate of 9.0 ml/min (room temperature) using UV (210 nm) The peaks were eluted at retention times of 19 minutes 58 seconds and 21 minutes 23 seconds.

对保留时间 19分 58秒的洗脱峰， 使用 YMC-PAK SIL-06层析柱 For the elution peak with a retention time of 19 minutes and 58 seconds, use a YMC-PAK SIL-06 column.

( 10Φ X 150mm), 流动相使用 90%正己烷（正己烷： 乙酸乙酯 =90: 10)， 流速 3.0ml/min (室温）进行洗脱， 利用 UV(254nm)分离出在保 留时间 9分 14秒洗脱峰， 得到化合物（ 10 )。 (10Φ X 150mm), the mobile phase was eluted with 90% n-hexane (n-hexane: ethyl acetate = 90: 10), flow rate 3.0 ml/min (room temperature), separated by UV (254 nm) at retention time of 9 min. The peak was eluted in 14 seconds to give the compound (10).

对保留时间 21分 23秒的洗脱峰， 使用 YMC- PAK SIL-06层析柱 For the elution peak with a retention time of 21 minutes and 23 seconds, use the YMC-PAK SIL-06 column.

(20Φ X 150mm), 流动相使用 88%正己烷（正己烷： 乙酸乙酯 =88: 12), 流速 9.0ml/min (室温）进行洗脱， 利用 UV(254nm)分离出在保 留时间 10分 55秒洗脱峰， 得到化合物 （11)。 (20Φ X 150mm), the mobile phase was eluted with 88% n-hexane (n-hexane: ethyl acetate = 88: 12), flow rate 9.0 ml/min (room temperature), separated by UV (254 nm) at retention time 10 min. The peak was eluted in 55 seconds to give the compound (11).

3d 68.Ζ0Ϊ/900Ζ OAV 化合物 （ 9 ) 的 NMR光谱

3d 68.Ζ0Ϊ/900Ζ OAV NMR spectrum of compound ( 9 )

关于化合物 （8)、 （10)、 （11), 由于是已知化合物参照论文如下： 化合物 （8) Isaka K., Nagatsu A.， et. AL, Chem. Pharm. Bull.(9), pp.1072-1076, 2001

Regarding the compounds (8), (10), (11), since it is a known compound, the reference paper is as follows: Compound (8) Isaka K., Nagatsu A., et. AL, Chem. Pharm. Bull. (9), pp .1072-1076, 2001

化合物 （ 10 )， (11) Kojima K.， Isaka K, et. Al., Chem. Pharm. Bull., 46, pp. 1781-1784, 1998 Compound (10), (11) Kojima K., Isaka K, et. Al., Chem. Pharm. Bull., 46, pp. 1781-1784, 1998

化合物结构式如下：  The structural formula of the compound is as follows:

化合物 （ 11 )

Compound ( 11 )

上述从阜康阿魏植物种提取、 分离的化合物分别属于倍半 萜烯香豆素、 倍半萜烯色酮、 倍半萜烯苯丙基类衍生物。  The above compounds extracted and isolated from the F. sylvestris plant species are sesquiterpene coumarin, sesquiterpene ketone, and sesquiterpene phenylpropyl derivatives, respectively.

实施例 11 : 诱导白血病细胞凋亡实验：  Example 11: Induction of apoptosis in leukemia cells:

实验按照 Vermes 等的方法 ( Journal of Immunological Methods 184卷 39-51 页 1995年 ) 进行。 HL-60及 Jurkat 细胞， 使其在 RPMI1640培养基中浮游， 成为 1 - 3 x 106细胞 /mlo 在此细胞浮游液 500μ1中， 加入被检物或培养基， 使浓度 成为 10pg/ml， 在 37。C、 5 % C0₂的条件下培养 24小时。 离心 洗涤后， 加入膜联蛋白 （ annexins ) V结合緩冲液， 再添加 5μ1 的膜联蛋白 V-FITC。 通过流式细胞计， 将膜联蛋白 V阳性细 胞为诱导凋亡的细胞， 以对全部细胞的百分比进行评价。 The experiment was carried out in accordance with the method of Vermes et al. (Journal of Immunological Methods, Vol. 184, pp. 39-51, 1995). HL-60 and Jurkat cells were allowed to float in RPMI1640 medium to become 1 - 3 x 106 cells/mlo. In this cell suspension 500 μl, the test substance or medium was added to make the concentration. Become 10pg/ml, at 37. C, 5% C0 ₂ was incubated for 24 hours. After centrifugation, annexin V binding buffer was added and 5 μl of annexin V-FITC was added. The annexin V positive cells were induced to apoptosis by flow cytometry, and the percentage of all cells was evaluated.

MBP激酶活性实验： 实验按照 De Souza等的方法（Blood 99卷 3432 - 3438页 2002年 ) 进行。 HL-60及 Jurkat细胞， 使其在使其在 RPMI1640培养基中浮游， 成为 1 - 3 X 106细胞 /ml。 在此细胞浮游液 500μ1中， 加入被检物或培养基， 使浓度 成为 10pg/ml, 在 37 °C、 5 % C02的条件下培养 4、 或者 6小时。 离 心洗涤后 ， 添加可溶緩冲液以溶解细胞， 利用 含有 MBP5mg/ml的凝胶电泳。对凝胶进行脱变性、再变性处理之后， 作用 32P-ATP进行 3小时的磷酸化反应。 对凝胶进行干燥后， 用分析器分析放射活性。 若 36kDa产生带， 那么 MBP激酶活 性即为阳性 （ + )。  MBP kinase activity assay: The experiment was carried out according to the method of De Souza et al. (Blood 99, pp. 3432-3438, 2002). HL-60 and Jurkat cells were allowed to float in RPMI1640 medium to become 1 - 3 X 106 cells/ml. In the cell suspension 500 μl, the test substance or the medium was added to have a concentration of 10 pg/ml, and cultured at 37 ° C, 5 % C02 for 4 or 6 hours. After centrifugation, a soluble buffer was added to dissolve the cells, and gel electrophoresis was carried out using MBP 5 mg/ml. After the gel was subjected to de-denaturation and re-denaturation treatment, 32P-ATP was subjected to phosphorylation reaction for 3 hours. After the gel was dried, the radioactivity was analyzed using an analyzer. If the band is produced at 36 kDa, the MBP kinase activity is positive (+).

其结果如表 4 所示。 化合物 （ 7 ) ~ ( 11 ) 具有强 HL-60 细胞凋亡诱导活性， 伴随 MBP激酶活性。  The results are shown in Table 4. Compounds (7) ~ (11) have strong apoptosis-inducing activity of HL-60 cells, accompanied by MBP kinase activity.

表 4  Table 4

实施例 12: 制备药物剂型 当本发明提供的化合物或其药学上可接受的盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物单独给药时， 优选以 药用制剂提供， 该制剂包括本发明提供的化合物或其药学上可 接受的盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物 其药学上可接受的载体或赋形剂以及可任选的一种或多种其它 的治疗成份。 Example 12: Preparation of a pharmaceutical dosage form When the compound provided by the present invention, or a pharmaceutically acceptable salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, is administered alone, it is preferably provided in a pharmaceutical preparation, the preparation including the present A pharmaceutically acceptable carrier or excipient of the compound or a pharmaceutically acceptable salt, enantiomer, racemate, tautomer or physiologically functional derivative thereof, and optionally one Or a variety of other therapeutic ingredients.

所述制剂包括口服给药制剂、 胃肠外给药制剂 （ 包括皮内 注射、 肌内注射、 静脉注射以及关节腔内注射）、 吸入制剂 （包 括由不同剂量的加压气雾器、 喷雾器、 吹入器中产生的微粒粉 剂或雾剂 ）、 直肠和局部给药制剂 （包括皮肤给药、 口腔给药、 舌下给药、 眼内给药）。 最适合的给药途径取决于用药患者的状 况和疾病。 制剂通常以单位剂型提供， 并且可以通过药学领域 中已知的任何方法进行制备。 所有的方法都包括将活性成份与 载体混合的步骤， 载体由一个或多个辅助成分构成。 通常情况 下通过使活性成分与液体或精细固体载体或两者均勾紧密结 合， 随后根据需要将所述产物制成制剂； 适宜口服给药的本发 明制剂可以以独立的单位如胶嚢剂、 扁嚢剂或片剂提供， 而每 单位含有预定量的活性成分； 散剂或颗粒剂； 水溶性液体或悬 浮液； 或水包油型乳剂或油包水型乳剂； 也可以是大丸剂、 药 糖剂或糊剂。  The preparations include oral administration preparations, parenteral administration preparations (including intradermal injection, intramuscular injection, intravenous injection, and intra-articular injection), inhalation preparations (including pressurized aerosol devices, sprayers, and different doses). Microparticle powders or aerosols produced in insufflators, rectal and topical formulations (including dermal, buccal, sublingual, intraocular). The most suitable route of administration will depend on the condition and disease of the patient being administered. The formulations are usually provided in unit dosage form and may be prepared by any methods known in the art of pharmacy. All methods include the step of mixing the active ingredient with a carrier which is comprised of one or more accessory ingredients. The preparation is usually formulated by bringing the active ingredient into a liquid or fine solid carrier or both, and then preparing the product as needed; the preparation of the invention suitable for oral administration can be in a separate unit such as a capsule, A sputum or tablet is provided, and each unit contains a predetermined amount of the active ingredient; a powder or granule; a water-soluble liquid or suspension; or an oil-in-water emulsion or a water-in-oil emulsion; or a granule or a medicine Sugar or paste.

通过任选与一种或多种辅助成分一起压片或塑型可以制备 片剂； 通过在适当的机器上压制自由流动形式如粉末或颗粒（任 选与粘合剂、 润滑剂、 惰性稀释剂、 表面活性剂或分散剂混合） 中的活性组分可以制备压制片剂； 所述片剂可任选为包衣或压 痕的， 并且配成制剂以提供活性成分的緩释或控释。  Tablets may be prepared by tableting or molding, optionally with one or more accessory ingredients; by compressing free-flowing forms such as powders or granules on a suitable machine (optional with binders, lubricants, inert diluents) The active component of the surfactant, or dispersion of the dispersing agent can be used to prepare compressed tablets; the tablets may optionally be coated or indented and formulated to provide sustained or controlled release of the active ingredient.

胃肠外给药的制剂包括水性和非水性无菌注射液， 该注射 液可以含有抗氧化剂、 緩冲剂、 抗菌剂及使制剂与患者血液等 张的溶盾， 水性和非水性的无菌悬浮液， 该悬浮液可以含有悬 浮剂和增稠剂； 所述制剂可以以单位剂量或多剂量包装， 例如 用密封的安剖管和管制瓶提供； 并且可以在冷冻千燥的条件下 储存， 自使用前仅需提供无菌的液体载体如盐水或注射用水； 可以用散剂、 颗粒剂及前面描述的各种片剂制备临时注射溶液 和悬浮液。 Formulations for parenteral administration include aqueous and non-aqueous sterile injectable solutions, the injection The solution may contain an antioxidant, a buffer, an antibacterial agent, and a solution to dissolve the preparation with the patient's blood, an aqueous and non-aqueous sterile suspension, and the suspension may contain a suspending agent and a thickening agent; Unit dose or multi-dose packaging, for example, provided with sealed ampoule and control bottles; and can be stored under freezing conditions, only need to provide a sterile liquid carrier such as saline or water for injection before use; , granules and various tablets as described above are prepared for the preparation of temporary injection solutions and suspensions.

直肠给药的制剂可用常见的载体 （如可可脂或聚乙二醇） 的栓剂形式提供。  Formulations for rectal administration may be presented as a suppository in a conventional carrier such as cocoa butter or polyethylene glycol.

用于口腔的局部给药的制剂 （如口腔含化剂或舌下给药 剂）， 包括在调味基质 （如蔗糖、 ***糖） 中的活性成份的锭 剂。  A preparation for topical administration to the oral cavity (e.g., an oral or sublingual agent), and a tablet comprising the active ingredient in a flavoring base such as sucrose or arabinose.

可以理解， 除上面特别提到的成分外， 本发明的制剂还包 括本领域相关制剂类型的其它常规成分。 下面以化合物 （ 1 ) 为 例列举的实施例， 同样适用于本发明中的其它化合物。  It will be understood that in addition to the ingredients specifically mentioned above, the formulations of the present invention also include other conventional ingredients of the type of formulation in the art. The following examples, exemplified by the compound (1), are equally applicable to the other compounds in the present invention.

① 550mg的化合物 （ 1 )、 10550mg的乳糖、 4500mg的淀 粉均勾混合， 添加羟基丙基纤维素溶液 0.15ml , 炼合制造软块， 利用造粒机制造颗粒， 进行干燥。 干燥后的颗粒中加入 300mg 的硬脂酸镁均匀混合后， 利用压片机制造片剂。  1 550 mg of the compound (1), 10550 mg of lactose, and 4500 mg of starch were mixed, and 0.15 ml of a hydroxypropylcellulose solution was added thereto, and a soft block was produced by refining, and granules were produced by a granulator to be dried. After the dried granules were uniformly mixed with 300 mg of magnesium stearate, tablets were produced by a tableting machine.

②将从阜康阿魏提取的化合物 （ 1 ) 10g与玉米淀粉 40g混 合， 加水制成软材， 过 12 目歸造粒、 干燥， 得到颗粒剂， 在本 颗粒剂中每 500mg中含化合物 （ 1 ) 100mg。  2 10 g of compound ( 1 ) extracted from Fukang Awei and 40 g of corn starch were mixed with water to make soft material. After 12 mesh, granulation and drying were carried out to obtain granules, and the compound was contained in 500 mg of the granules. 1) 100mg.

③将从阜康阿魏提取的化合物 （ 1 ) 40g与乳糖 100g、 硬脂 酸镁 10g混合， 以每 600mg填充肠溶胶嚢， 本肠溶胶嚢剂中， 每个胶嚢含化合物 （ 1 ) 160mg。  3 Compound (1) 40g extracted from Fukang Awei is mixed with 100g of lactose and 10g of magnesium stearate to fill the intestinal sol with 600mg. In this intestinal sol agent, each capsule contains compound (1) 160mg. .

④将从阜康阿魏提取的化合物 （ 1 ) 25g 以普通的注射剂制 造法， 用加热至 60 摄氏度的注射用蒸馏水 1000ml 溶解， 用 NaCL调解等张，封入安剖瓶。本注射液 10ml中含有化合物（ 1 ) 250mg。 4 Compound ( 1 ) 25g extracted from Fukang Awei is prepared by ordinary injection The method was dissolved in 1000 ml of distilled water for injection heated to 60 ° C. The isotonic sheets were adjusted with NaCL and sealed in an ampoule. The injection (10 ml) contained 250 mg of the compound (1).

⑤将从阜康阿魏提取的化合物 （ 7 ) 10g与玉米淀粉 40g混 合， 加水制成软材， 过 12 目薛造粒、 干燥， 得到颗粒剂， 在本 颗粒剂中每 500mg中含化合物 （ 7 ) 100mg。  5 10 g of compound ( 7 ) extracted from Fukang Awei and 40 g of corn starch were mixed with water to make a soft material, granulated and dried by 12 mesh, to obtain granules, and the compound was contained in 500 mg of the granules. 7) 100mg.

⑥将从阜康阿魏提取的化合物 （ 7 ) 40g与乳糖 100g、 硬脂 酸镁 10g混合， 以每 600mg填充肠溶胶嚢， 本肠溶胶囊剂中， 每个胶嚢含化合物 （ 7 ) 160mg。  6 40 g of compound ( 7 ) extracted from Fukang Awei, mixed with 100 g of lactose and 10 g of magnesium stearate, and filled with intestinal sol per 600 mg. In this enteric capsule, each capsule contains compound ( 7 ) 160 mg .

⑦将从阜康阿魏提取的化合物 （ 7 ) 25g 以普通的注射剂制 造法， 用加热至 60 ¾氏度的注射用蒸馏水 1000ml 溶解， 用 NaCL调解等张，封入安剖瓶。本注射液 10ml中含有化合物 ( 7 ) 250mg。  7 Compound (7) 25 g extracted from Fukang Awei was dissolved in 1000 ml of distilled water for injection heated to 60 3⁄4 ° C by ordinary injection preparation, and the isotope was adjusted with NaCL, and sealed into an ampoule. The injection (10 ml) contains 250 mg of the compound (7).

实施例 13、 制备食品  Example 13, preparation of food

本发明提供的化合物也可以以食品的形式提供， 优选的食品 形式包括粉末、 颗粒、 糊剂、 胶状物等， 颗粒形式可以添加糖 (如乳糖） 来增加甜味； 也可以制备成饮料的形式； 这些食物 或饮料除阜康阿魏提取物外， 还可以加入维生素、 无机元素， 如钙、 醇类、 去味剂， 如多酚。 这些食物包括特殊的健康食品、 医用食品等种类。 参考文献：  The compounds provided by the present invention may also be provided in the form of a food. Preferred food forms include powders, granules, pastes, gels, etc., and the granule form may be added with a sugar (such as lactose) to increase the sweetness; or may be prepared into a beverage. In addition to these extracts, these foods or beverages may also contain vitamins, inorganic elements such as calcium, alcohols, deodorants, such as polyphenols. These foods include special health foods, medical foods, and the like. references:

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Claims

权利要求 Rights request

1、 一种从阜康阿魏中提取的活性化合物 （ 1) 或其盐、 对 映体、 外消旋体、 互变异构体或生理官能衍生物，

1. An active compound (1) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, extracted from Fukang

化合物 （ 1 )  Compound ( 1 )

2、 一种从阜康阿魏中提取的活性化合物 （ 2) 或其盐、 对 映体、 外消旋体、 互变异构体或生理官能衍生物，

2. An active compound (2) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, extracted from Fukang

化合物 （ 2)  Compound ( 2)

3、 一种从阜康阿魏中提取的活性化合物 （ 3) 或其盐、 对 映体、 外消旋体、 互变异构体或生理官能衍生物，

3. An active compound (3) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, extracted from Fukang

化合物 （ 3)  Compound ( 3)

4、 一种从阜康阿魏中提取的活性化合物 （ 5) 或其盐

映体、 外消旋体 ，

化合物 （ 5) 4. An active compound (5) or a salt thereof extracted from Fukang

Aporoid, racemate,

Compound (5)

5、 根据权利要求 1、 2、 3或 4所述的活性化合物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在 于用于制备治疗或预防炎性疾病药物的用途。 5. The active compound according to claim 1, 2, 3 or 4, or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, for use in the preparation of a treatment or prevention The use of inflammatory disease drugs.

6、 根据权利要求 5所述的活性化合物或其盐、 对映体、 外 消旋体、 互变异构体或生理官能衍生物， 其特征在于所述炎性 疾病及临床病症包括关节炎、 或胃肠道疾病、 肺部疾病、 心脏 病、 神经组织疾病、 胰腺疾病、 腎疾病、 皮肤病、 眼部疾病、 移植器官疾病、 多器官疾病及病毒和细菌感染后的炎性后遗症。  The active compound or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof according to claim 5, characterized in that the inflammatory disease and clinical condition include arthritis, Or gastrointestinal tract disease, lung disease, heart disease, nervous tissue disease, pancreatic disease, kidney disease, skin disease, eye disease, transplant organ disease, multiple organ disease, and inflammatory sequelae after viral and bacterial infection.

7、 一种从阜康阿魏中提取的活性化合物 （ 4 ) 或其盐、 对 映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于 用于制备治疗或预防炎性疾病药物组合物的用途，  7. An active compound (4) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, extracted from Fukang, which is characterized by being used for the preparation of a treatment or prevention Use of a pharmaceutical composition for an inflammatory disease,

化合物 （ 4 )  Compound ( 4 )

8、 一种从阜康阿魏中提取的活性化合物 （ 6 ) 或其盐、 对 映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于 用于制备治疗或预防炎性疾病药物组合物的用途，

化合物 （ 6 ) 8. An active compound (6) or a salt, an enantiomer, a racemate, a tautomer or a physiologically functional derivative thereof, which is extracted from Fukang, and is characterized in that it is used for the preparation of a treatment or prevention Use of a pharmaceutical composition for an inflammatory disease,

Compound ( 6 )

9、 根据权利要求 1、 2、 3、 4、 7或 8所述的活性化合物或 其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其 特征在于适宜的盐包括与有机和无机酸或碱形成的盐。 9. The active compound according to claim 1, 2, 3, 4, 7 or 8 or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized in that it is suitable Salts include salts with organic and inorganic acids or bases.

10、 根据权利要求 1、 2、 3、 4、 7或 8所述的活性化合物 或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于生理官能衍生物包括酯、 酰胺、 氨基甲酸酯。 . 10. The active compound according to claim 1, 2, 3, 4, 7 or 8 or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized by physiological function Derivatives include esters, amides, and carbamates. .

11、 根据权利要求 1、 2、 3、 4、 7或 8所迷的活性化合物 或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于可与第二种药学活性物质联合用于制备治疗或预防 炎性疾病药物的用途。  11. The active compound according to Claim 1, 2, 3, 4, 7 or 8 or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized in that it is compatible with The use of a second pharmaceutically active substance in combination for the manufacture of a medicament for the treatment or prevention of an inflammatory disease.

12、 根据权利要求 11所述的活性化合物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于炎性疾 病及临床病症包括关节疾病， 特别是关节炎、 或胃肠道疾病、 肺部疾病、 心脏病、 神经组织疾病、 胰腺疾病、 肾疾病、 皮肤 病、 眼部疾病、 移植器官疾病、 多器官疾病及病毒和细菌感染 后的炎性后遗症。  The active compound or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof according to claim 11, characterized in that the inflammatory disease and the clinical condition include joint diseases, in particular Arthritis, or gastrointestinal disease, lung disease, heart disease, nervous tissue disease, pancreatic disease, kidney disease, skin disease, eye disease, transplant organ disease, multiple organ disease, and inflammatory sequelae after viral and bacterial infection .

13、 根据权利要求 1、 2、 3、 4、 7或 8所述的活性化合物 或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于用于制备保健食品、 饮料、 或饲料。  The active compound according to claim 1, 2, 3, 4, 7 or 8 or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized in that it is used for Prepare health foods, beverages, or feeds.

14、 一种从阜康阿魏中提取活性化合物的方法， 包括如下 步骤：  14. A method of extracting an active compound from a scorpion, comprising the steps of:

1 ) 阜康阿魏经曱醇提取后的提取液减压浓缩、 干燥得到萃 取物；  1) The extract of the scorpion argan extract after sterol extraction is concentrated under reduced pressure and dried to obtain an extract;

2 ) 将萃取物用氯仿、 乙酸乙酯依次萃取；  2) extracting the extract sequentially with chloroform and ethyl acetate;

3 ) 将氯仿的萃取物经层析柱用正己烷与乙酸乙酯的混合液 洗脱；  3) extracting the chloroform extract through a chromatography column with a mixture of n-hexane and ethyl acetate;

4 ) 对步骤 3 ) 收集到的洗脱液经高压液相色谱进一步分离 精制， 得到活性化合物。  4) The eluate collected in step 3) is further separated and purified by high pressure liquid chromatography to obtain an active compound.

15、 一种从阜康阿魏中提取的化合物 （ 7 ) 活性化合物或其 盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 15. A compound (7) active compound extracted from Fukang Awei or its active compound or a salt, an enantiomer, a racemate, a tautomer or a physiologically functional derivative,

化合物 （ 7)

Compound (7)

16、 一种从阜康阿魏中提取的化合物 （9) 活性化合物或其 盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物，

16. A compound (9) active compound or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, which is extracted from Fukang.

化合物 （ 9)  Compound ( 9)

17、 根据权利要求 1或 2所述的化合物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于用于制 备治疗或预防肿瘤药物的用途。  17. A compound according to claim 1 or 2, or a salt, enantiomer, racemate, tautomer or physiologically functional derivative thereof, for use in the manufacture of a medicament for the treatment or prevention of a tumor.

18、 根据权利要求 17所述的活性化合物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于肿瘤疾 病包括骨髓芽球性白血病。  The active compound or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof according to claim 17, characterized in that the tumor disease comprises myeloid leukemia.

19、 一种从阜康阿魏中提取的活性化合物 （ 8) 或其盐、 对 映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于 用于制备治疗或预防肿瘤药物组合物的用途，  19. An active compound (8) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, which is extracted from Fukang, and is characterized in that it is used for the preparation of a treatment or prevention Use of a tumor drug composition,

化合物 （8)

Compound (8)

20、 一种从阜康阿魏中提取的活性化合物 （ 10 ) 或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在 于用于制备治疗或预防肿瘤药物组合物的用途，

20. An active compound (10) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, which is extracted from Fukang, and is characterized in that it is used for the preparation of a treatment or prevention Use of a tumor drug composition,

化合物 （ 10 )  Compound ( 10 )

21、 一种从阜康阿魏中提取的活性化合物 （ 11 ) 或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在 于用于制备治疗或预防肿瘤药物组合物的用途，

An active compound (11) or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, which is extracted from Fukang, and is characterized in that it is used for the preparation of a treatment or prevention Use of a tumor drug composition,

化合物 ( 11 )  Compounds ( 11 )

22、 根据权利要求 15、 16、 19、 20或 21所述的活性化合 物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于适宜的盐包括与有机和无机酸或碱形成的盐。  22. The active compound according to claim 15, 16, 19, 20 or 21 or a salt, enantiomer, racemate, tautomer or physiologically functional derivative thereof, characterized in that suitable salts include Salts formed with organic and inorganic acids or bases.

23、 根据权利要求 15、 16、 19、 20或 21所述的活性化合 物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于生理官能衍生物包括酯、 酰胺、 氨基曱酸酯。  The active compound according to claim 15, 16, 19, 20 or 21 or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized by a physiologically functional derivative Including esters, amides, amino phthalates.

24、 根据权利要求所述 15、 16、 19、 20或 21的活性化合 物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于可与第二种药学抗肿瘤活性物质联合用于制备治疗 或预防肿瘤药物的用途。  24. The active compound according to claim 15, 16, 19, 20 or 21 or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized by The use of a pharmaceutical antitumor active substance in combination for the preparation of a medicament for treating or preventing a tumor.

25、 根据权利要求 24所述的活性化合物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于肿瘤包 括骨髓芽球性白血病。 The active compound according to claim 24, or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof, characterized by a tumor pack Including bone marrow globulin leukemia.

26、 根据权利要求 15、 16、 19、 20或 21所述的活性化合 物或其盐、 对映体、 外消旋体、 互变异构体或生理官能衍生物， 其特征在于用于制备保健食品、 饮料、 或饲料。  The active compound or a salt, enantiomer, racemate, tautomer or physiological functional derivative thereof according to claim 15, 16, 19, 20 or 21, which is characterized in that it is used for the preparation of health care Food, drink, or feed.

27、 一种抗肿瘤药物组合物， 其特征在于有效成分包括从 阜康阿魏的根茎提取、 分离的倍半萜烯香豆素、 倍半萜烯色酮、 倍半萜烯苯丙基类衍生物。  An antitumor pharmaceutical composition, characterized in that the active ingredient comprises sesquiterpene coumarin, sesquiterpene ketone, sesquiterpene phenyl propyl group extracted and isolated from the rhizome of Fukang Awei derivative.