多花黄精水提物成分分析及抗肿瘤活性研究  

Compositional Analysis and Antitumor Activity of Aqueous Extracts of Polygonatum Cyrtonema Hua

作　　者：宋露 耿春叶 邢陈昱 王倩 郭瑶瑶 陈艳君 王芳 李国四 王威 高雷雷 刘东[2] 韩邦兴[2] SONG Lu;GENG Chunye;XING Chenyu;WANG Qian;GUO Yaoyao;CHEN Yanjun;WANG Fang;LIGuosi;WANG Wei;GAO Leilei;LIU Dong;HAN Bangxing(School of Pharmacy,Anhui University of Chinese Medicine,Hefei 230012 Anhui,China;Generic Technology Research Center for Anhui Traditional Chinese Medicine Industry,Anhui Engineering Research Center for Eco-agriculture of Traditional Chinese Medicine,Anhui Dabie Mountain Chinese Academy of Medicine,West Anhui University,Luʼan 237012 Anhui,China)

机构地区：[1]安徽中医药大学药学院,安徽合肥230012 [2]皖西学院安徽省现代中药产业共性技术研究中心安徽省中药生态农业工程研究中心安徽省大别山中医药研究院,安徽六安237012

出　　处：《中药新药与临床药理》2024年第7期952-962,共11页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：安徽省教育厅项目自然科学重点项目(KJKJ2020A0628,KJ2021A0951,2022AH051673,2022AH051687);安徽省大别山中医药研究院开放课题(TCMADM-2023-04);安徽省现代中药产业共性技术研究中心开放课题(AHTCMGTRC-2023-03)。

摘　　要：目的分析多花黄精水提物成分,并对其体内外抗肿瘤活性进行评价,为进一步开发利用多花黄精提供理论基础。方法(1)通过水提法制备多花黄精水提物,利用UPLC-Q-TOF/MS、苯酚硫酸法等方法分析多花黄精水提物化学成分,并通过CCK-8法检测多花黄精水提物对多种肿瘤细胞增殖抑制活性,采用流式细胞术检测细胞周期和凋亡情况,用Western Blot法检测凋亡相关蛋白Bcl-2和Bax的表达情况,利用多花黄精水提物含药血清检测入血成分对细胞增殖抑制活性。(2)构建荷瘤小鼠模型,随机分为模型组(予以生理盐水)、阳性药环磷酰胺组(50 mg·kg^(-1))以及多花黄精水提物低、中、高剂量组(55.9、111.8、223.6 mg·kg^(-1)),灌胃治疗7 d。治疗期间,观察小鼠体质量及肿瘤体积变化情况。治疗结束后处死小鼠,采用苏木素-伊红(HE)染色观察心、肝、脾、肺、肾以及肿瘤组织病理学变化;免疫组化(IHC)检测肿瘤组织Bcl-2及Bax蛋白表达情况。结果多花黄精水提物多糖含量达到(10.07±1.3)%,黄酮含量为(0.044±0.004)%,并且通过UPLC-Q-TOF/MS检测出39种成分,包含黄酮类的黄芩素、槲皮素、木犀草素、芦丁,有机酸类的阿魏酸及多酚类的没食子酸等抗肿瘤成分。多花黄精水提物对Hela、A549、4T1、B16、MFC和HepG2细胞均有抑制作用(P<0.001),其中对B16细胞抑制作用最为明显,且多花黄精水提物可诱导B16细胞周期阻滞于G0/G1期(P<0.001)。流式双染和Western Blot结果显示多花黄精水提物明显促进B16细胞凋亡,降低Bcl-2的表达,且促进Bax的表达(P<0.01,P<0.001)。多花黄精水提物入血成分对B16细胞也具有抑制作用(P<0.001)。此外,多花黄精水提物体内活性实验结果显示,与模型组比较,不同剂量的多花黄精水提物能够抑制肿瘤生长,诱导肿瘤细胞坏死,降低Bcl-2表达,提高Bax的表达。结论多花黄精水提物成分丰富,含有多种抗肿瘤活性成分,能抑�Objective To analyze the composition of the aqueous extract of Polygonatum Cyrtonema Hua(PCHE)and evaluate its antitumor activity in vitro and in vivo.Our aim is to provide a theoretical basis for the further development and utilization of Polygonatum Cyrtonema Hua.Methods(1)PCHE was prepared by aqueous extraction,and the chemical composition of PCHE was analyzed by UPLC-Q-TOF/MS and phenol-sulfuric acid method.The inhibitory activity on tumor cells proliferation of PCHE was detected by CCK-8 assay.Cell cycle and apoptosis were detected by flow cytometry,and the expression of apoptosis-related proteins Bcl-2 and Bax was detected by Western Blot.The inhibitory activity of PCHE-containing serum on cell proliferation was detected.(2)A B16 tumor-bearing mice model was constructed and model mice were randomly divided into the model group(saline),the positive drug group(CTX:50 mg·kg^(-1)),and PCHE low-,medium-,and high-dose groups(55.9,111.8,223.6 mg·kg^(-1)),and treated by gavage for 7 days.Changes in body weight and tumor volume of mice were observed during the treatment period.The mice were executed after the treatment,and the histopathological changes of heart,liver,spleen,lung,kidney and tumor were observed by hematoxylin-eosin(HE)staining.The protein expression of Bcl-2 and Bax in tumor tissues was detected by immunohistochemistry(IHC).Results The polysaccharide content of PCHE reached(10.07±1.3)%,and the flavonoid content was(0.044±0.004)%,and thirty-nine components were detected by UPLC-Q-TOF/MS,which contained antitumor components such as flavonoids(baicalein,quercetin,luteolin and rutin),organic acids(ferulic acid)and polyphenols(gallic acid),etc.PCHE exhibited the inhibitory effects on Hela,A549,4T1,B16,MFC and HepG2 cells,among which the inhibitory effect on B16 cells was the most significant(P<0.001),and PCHE induced cell cycle arrest at G0/G1 phase in B16 cells(P<0.001).The results of double-staining flow cytometry and Western Blot showed that PCHE significantly promoted apoptosis of B16 cells,decreas

关 键 词：多花黄精 水提物 抗肿瘤 黑色素瘤B16细胞 凋亡 小鼠 大鼠 

分 类 号：R285.5[医药卫生—中药学]