CN115368420B - 具有抗炎活性的化合物及其制备方法和应用 - Google Patents
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Abstract
本发明属于化学医药领域，具体涉及一种具有抗炎活性新化合物及其制备方法和用途。具体是以十字花科植物萝卜的干燥成熟种子莱菔子为原料，经一定的制备步骤分离得到一个新的化合物，根据NMR分析，阐明了具体结构，该化合物为首次报道。本发明公开了一种新化合物的具体结构，并且对其抗炎活性进行评估，结果表明该化合物具有明显的抗炎活性。
Description
技术领域
本发明属于化学医药领域，具体涉及莱菔子中1个具有抗炎活性新化合物及其制备方法。
背景技术
莱菔子(Raphani semen)为十字花科植物萝卜Raphanus sativus.L.的干燥成熟种子。莱菔子作为药食同源中药，在我国已有1400多年药用历史。主产于我国的河北、河南、浙江、湖北、四川等省。莱菔子性平，味辛甘，归肺、脾、胃经。具有消食除胀，降气化痰的功效，用于饮食停滞，脘腹胀痛，大便秘结，积滞泻痢，痰壅喘。目前从莱菔子中分离得到的化合物包括：生物碱、硫代葡糖苷和异硫氰酸盐、黄酮、挥发油、脂肪酸类、多糖和蛋白质等。研究表明，莱菔子具有平喘、镇咳、祛痰，抗氧化，降血压、降血脂，抗菌，抗炎，增强胃肠道动力等作用药理作用。
中药成分快速分离纯化技术对于阐明中药复杂的物质基础、质量控制以及发现潜在的活性物质具有重要意义。高压制备液相系统在分析型高效液相色谱的基础上，在保证样品分离度的前提下，大幅度提高载样量，从而快速获得高纯度的目标化合物。利用传统的柱色谱分离结合高压制备液相快速获得单体化合物。
发明内容
本发明的目的之一在于从莱菔子中发现了如下式I的1个新化合物，化学结构如下：
本发明的目的之二在于公开新化合物的制备方法，包括以下步骤：
(1)取十字花科植物萝卜的干燥成熟种子莱菔子，加入石油醚回流脱脂，合并滤渣；在滤渣中加入70％甲醇，冷浸提取，合并提取液，减压浓缩，得粗提物；
(2)将粗提物溶于蒸馏水得混悬液，依次用正己烷、乙酸乙酯、正丁醇萃取，保留乙酸乙酯萃取相和正丁醇萃取相，得乙酸乙酯萃取液和正丁醇萃取液，将乙酸乙酯萃取液和正丁醇萃取液减压浓缩，得乙酸乙酯萃取物和正丁醇萃取物；
(3)将正丁醇萃取物过AB-8大孔吸附树脂柱分离，以乙醇-水(0:100,1:4,3:7,1:1,7:3, 4:1,100:0)梯度洗脱，共分为7个组分，分别命名为Cr.A-G。Cr.D经制备液相色谱，55％甲醇-水洗脱，得到式I化合物，命名为Raphanused A。
在本发明的上述步骤中，其中：
步骤(1)中，所述莱菔子与石油醚的质量体积比为1：4。
步骤(1)中，莱菔子与70％甲醇水的质量体积比为1：10，质量体积比的单位为kg/L。例如，质量体积比为1:10，即1kg莱菔子需要加入10L的70％甲醇水；冷浸时间为15天；重复次数为3次。
步骤(2)中，所述莱菔子粗提物与蒸馏水的质量体积比为1：1；所述莱菔子粗提物与正己烷、乙酸乙酯、正丁醇各自的质量体积比均为1：1。
步骤(3)中，液相色谱中色谱柱规格为Innocal ODS-2，10×250mm，5μm。流速为3mL/min，柱温为25℃，检测为双波长检测，分别为254nm和220nm，保留时间为35min。
本发明的目的之三还涉及所述式I化合物制备抗炎药物中的应用。其中，所述化合物抑制LPS诱导的RAW264.7炎症细胞中NO的产生。结果显示，式I化合物的IC50为16.54±2.11μmol/L，略低于地塞米松的IC50(8.27±1.21)μmol/L，展现出抗炎作用。
附图说明
图1式I化合物HREIMS图
图2式I化合物核磁共振1H谱图
图3式I化合物核磁共振13C谱图
图4式I化合物核磁共振1H-1H COSY谱图
图5式I化合物核磁共振HMBC谱图
图6式I化合物核磁共振HSQC谱图
具体实施方式
下面结合实施例对本发明作进一步阐述。必须说明下述实施例是用于说明本发明而不是对本发明的限制。根据本发明的实质对本发明进行的简单改进都属于本发明要求保护的范围。
下述实施例所使用的部分仪器和试剂为：METTLER AE240型电子天平(梅特勒--托利多(上海)有限公司)，KQ-250B超声波清洗器(昆山市超声仪器有限公司)。细胞培养箱(力新仪器上海有限公司)；96孔细胞培养板；酶标仪(Thermo)；DEME培养基，胎牛血清(FBS，Gibco公司)；LPS，DMSO(Sigma公司)；CCK8试剂盒；细胞计数器；其他试剂为分析纯，水为蒸馏水。其他仪器均是实验室常用仪器；其他试剂也均是常规试剂，均可以市售或者实验室获得。
实施例1式I化合物提取
通过如下方法制备式I化合物：
(1)取粉碎后的莱菔子11kg，加四倍量石油醚(60-90℃)脱脂3次，挥干石油醚，待无石油醚味时，加入10倍量的70％甲醇，室温条件下浸泡15天，过滤得70％甲醇水提取液，反复浸提3次，合并提取液，浓缩，即得莱菔子的70％甲醇水粗提物785.6g。
(2)取785.6g 70％甲醇水粗提物溶于785.6mL蒸馏水中，置于分液漏斗中，加入正己烷785.6mL，萃取3次，之后依次用500mL的乙酸乙酯和正丁醇萃取3次。合并各级萃取液，浓缩，得到165.11g正己烷萃取物、25.79g乙酸乙酯萃取物、41.9g正丁醇萃取物，水部位为452.8g。
(3)将正丁醇萃取物过AB-8大孔吸附树脂柱分离，以乙醇-水(0:100,1:4,3:7,1:1,7:3, 4:1,100:0)梯度洗脱，共分为7个组分，分别命名为Cr.A-G。Cr.D经制备液相色谱(检测为双波长检测，分别为254nm和220nm)，55％甲醇-水洗脱(Innocal ODS-2，10×250mm， 5μm；流速为3mL/min，柱温为25℃，保留时间为35min)，得到式I化合物，即Raphanused A。
实施例2式I化合物鉴定
对得到的化合物进行结构鉴定，鉴定结果如下：
式I化合物：黄色无定形粉末。ESI-MS测得分子离子峰为m/z:989.2843[M-H]-，结合氢谱碳谱数据推测分子式为C46H54O24。氢谱中低场区苯环上的δH 7.02(2H,s)，以及双键上的 7.55(1H,d,J＝15.8Hz)和6.52(1H,d,J＝15.8Hz)以及碳谱中δC 166.75、145.45、138.33、124.38、 114.75、106.16提示该化合物包含芥子酰基母核。氢谱中高场区，在δH5.35-3.98区间有14 个连氧氢信号，同时碳谱在δC 102.81-62.39区间有12个连氧碳信号，推测该化合物有两个糖结构。HSQC谱中，δH 5.22(1H,d,J＝3.7Hz)与δC 90.82直接相关，根据耦合常数及化学位移推测δH 5.22为端基氢信号，根据1H-1H COSY判断氢氢之间的连接方式结合碳谱，推断出其中一个糖为葡萄糖；HMBC谱中，δH 5.35(fru-3)与102.81相关，结合δC83.14，77.54，72.59， 63.15，62.39推断出该糖为果糖。葡萄糖的苷键构型，通过端基氢的耦合常数5.22(1H,d,J＝3.7 Hz)确定为α构型。结合莱菔子中化学成分的特点确定为蔗糖(葡萄糖加果糖)。通过与微谱数据库比对，化合物的NMR谱图与化合物dimeth-yl(±)-thomasidioate相似，且dimeth-yl(±)- thomasidioate为木脂素衍生物。提示该化合物主要包含芥子酰基、蔗糖和木脂素衍生物三个片段。三个片段的连接方式主要通过HSQC及HMBC谱来判断，HMBC谱中δH 3.67与δC166.58相关，证明该甲氧基连接在9位羰基碳上，而9’羰基碳δC 170.87与δH 5.22相关，证明9’羰基与果糖的3位相连形成苷。同时芥子酰基上9”位羰基碳δC 166.75与δH 4.37,4.20相关，证明芥子酰基与葡萄糖6位相连。化合物结构如式I所示，HMBC与HSQC相关如图5、 6所示，化合物为未被文献报导的新化合物，将其命名为Raphanused A其核磁共振信号归属如下。1H-NMR (600MHz,DMSO-d6)δH 7.75(1H,s,H-7),7.55(1H,d,J＝15.8Hz,H-7”),7.07(1H,s,H-6),7.02 (2H,s,H-2”,6”),6.52(1H,d,J＝15.8Hz,H-8”),6.16(2H,s,H-2’,6’),5.35(1H,d,J＝7.9Hz, fru-3),5.22(1H,d,J＝3.7Hz,glc-1),4.86(1H,s,H-7’),4.37(1H,d,J＝10.7Hz,glc-6a),4.20(1H, dd,J＝10.7,7.0Hz,glc-6b),4.11(1H,d,J＝5.6Hz,glc-5),4.06–3.98(1H,m,glc-3),3.93(1H,d, J＝0.9Hz,H-8’),3.84(3H,s,5-OCH3),3.76(6H,s,3’,5’-OCH3),3.67(3H,s,9-OCH3),3.58(6H,s, 3”,5”-OCH3),3.49(3H,s,3-OCH3)。13C-NMR(151MHz,DMSO-d6)δC 170.87(C-9’),166.75(C-9”),166.58(C-9),148.06(C-3’,5’),147.70(C-5),145.45(C-7”),145.36(C-3),141.86(C-4), 138.39(C-7),138.33(C-4”),134.46(C-4’),132.50(C-8),124.38(C-1”),122.89(C-2),122.40 (C-1),121.19(C-1’),114.75(C-8”),108.93(C-6),106.16(C-2”,6”),104.91(C-2’,6’),102.81 (fru-2),90.82(glc-1),83.14(fru-5),77.54(fru-3),72.96(glc-5),72.59(fru-4),71.36(glc-2),70.28(glc-3),70.18(glc-4),64.25(glc-6),63.15(fru-6),62.39(fru-1),59.78(3-OCH3),56.07(3”, 5”-OCH3),56.06(3’,5’-OCH3),56.00(5-OCH3),51.71(9-OCH3),46.47(C-8’),40.05(C-7’)。
所述化合物HREIMS如图1所示。
所述化合物核磁共振1H谱图如图2所示。
所述化合物核磁共振13C谱图如图3所示。
所述化合物核磁共振1H-1H COSY谱图谱图如图4所示。
所述化合物核磁共振HMBC谱图如图5所示。
所述化合物核磁共振HSQC谱图如图6所示。
确定其结构式为：
实施例3式I化合物抗炎活性研究
供试品配制：取式I化合物，配制成5、25、50、100、200μmol/L的供试品药液。
细胞培养：RAW264.7细胞用含10％FBS DMEM培养，置5％CO2、37℃、饱和湿度的细胞培养箱中培养。待细胞生长至对数生长期进行传代，2～3d传代1次。取至少3代后的细胞用于实验。
化合物对RAW264.7细胞NO生成的影响：取对数生长期的RAW264.7细胞，制成浓度为1×105/mL单细胞悬液，接种至96孔培养板中，每孔100μL，置于37℃、5％CO2的细胞培养箱中培养。培养24h后，分别设立空白对照组、LPS组、阳性药地塞米松组、受试药物组 (5、25、50、100、200μmol/L)，细胞同时用1μg/mL LPS和不同浓度药物在37℃、5％CO2、饱和湿度条件下培养24h。收集细胞上清液，以Griess法用酶标仪检测540nm波长处OD值，检测NO的释放量，计算IC50。
抑制率＝(C2-C1)/(C2-C0)×100％
式中：C0、C1、C2分别为540nm下测得的空白对照组、不同浓度的样品组、LPS组的吸光度。
该化合物能显著抑制LPS诱导的RAW264.7产生NO，其中式I化合物的IC50为(16.54±2.11)μmol/L，略低于地塞米松的IC50(8.27±1.21)μmol/L，表明式I化合物具有明显的抗炎作。
Claims (4)
1.一种化合物，其特征在于，其化学结构式如式I所示：
。
2.一种权利要求1所述式I化合物的制备方法，其特征在于，包括以下步骤：(1)取十字花科植物萝卜的干燥成熟种子莱菔子，加入石油醚回流脱脂，合并滤渣；在滤渣中加入70％甲醇，室温条件下浸泡15天，反复浸提3次，合并提取液，减压浓缩，得粗提物；其中，所述莱菔子与石油醚的质量体积比为1：4；所述莱菔子与70％甲醇水的质量体积比为1：10kg/L；(2)将粗提物按质量体积比为1：1溶于蒸馏水得混悬液，依次用质量体积比为1：1的正己烷、乙酸乙酯、正丁醇萃取，保留正丁醇萃取相，得正丁醇萃取液，将正丁醇萃取液减压浓缩，得正丁醇萃取物；(3)将正丁醇萃取物过AB-8大孔吸附树脂柱分离，以乙醇-水在0:100,1:4,3:7,1:1,7:3,4:1,100:0梯度下进行洗脱，共分为7个组分，分别命名为Cr.A-G；Cr.D经制备液相色谱，55％甲醇-水洗脱，得到式I化合物，液相色谱中色谱柱规格为Innocal ODS-2，10×250mm，5μm；流速为3mL/min，柱温为25℃，检测为双波长检测，分别为254nm和220nm，保留时间为35min。
3.权利要求1所述的式I化合物或者权利要求2制备方法获得的式I化合物在制备抗炎药物中的应用。
4.根据权利要求3所述的应用，其特征在于，所述式I化合物抑制LPS诱导的RAW264.7炎症细胞中NO的产生。
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102234297A (zh) * | 2011-05-06 | 2011-11-09 | 南京泽朗医药科技有限公司 | 一种从莱菔子中提取葡萄糖莱菔子苷的方法 |
| CN113893271A (zh) * | 2021-11-15 | 2022-01-07 | 广东工业大学 | 莱菔子提取物的新用途 |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102234297A (zh) * | 2011-05-06 | 2011-11-09 | 南京泽朗医药科技有限公司 | 一种从莱菔子中提取葡萄糖莱菔子苷的方法 |
| CN113893271A (zh) * | 2021-11-15 | 2022-01-07 | 广东工业大学 | 莱菔子提取物的新用途 |
Non-Patent Citations (4)
| Title |
|---|
| Intestinal anti-in fl ammatory activity of the seeds ofRaphanus sativus L. in experimental ulcerative colitis models;Ki-Choon Choi，等;《Journal of Ethnopharmacology》;第179卷;55 – 65 * |
| Phenylpropanoids from Brachybotrys paridiformis Maxim. ex Oliv. and their anti-HBV activities;Si-Tong Wu, 等;《Phytochemistry 》;第197卷;第113114页 * |
| Sucrose die ster of aryldihydronaphthale ne-type lignans fromEchium angustifoliumMill. and their antitumor activity;Ahmed Ramadan El-Rokh，等;《Phytochem istry》;第149卷;第155-160页 * |
| 超高效液相色谱– 四极杆静电场轨道阱高分辨 质谱法分析莱菔子化学成分;冯俊杰，等;化学分析计量;第30卷(第11期);第14-22页 * |
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