Polysaccharide ASPF from Acanthopanax senticosus Fructification: Structural Characterization and Its Anti-lung Cancer Activity in Vitro

doi:10.11924/j.issn.1000-6850.casb2021-0779

Abstract

Abstract:

Homogenous molecular weight polysaccharide ASPF was purified from the fructification of Acanthopanax senticosus, and its structure characterization and in vitro anti-lung cancer activity were explored. The extracted crude polysaccharides were separated by DEAE-52 column chromatography to obtain 3 components, after screening for in vitro anti-lung cancer activity, ASPB-3 was obtained by Sephadex G-200 gel column chromatography to get ASPF. GPC, IR and HPLC methods were selected to analyze the structure of the polysaccharide, and the CCK-8 method was used to study its effect on the survival rate of lung cancer cells in vitro. The content of uronic acid in ASPF was 22.9%. It was an acidic pectin-like structure polysaccharide. Its relative molecular mass M_W was 9.7462×10⁴. IR scanning results showed that it had a pyran ring structure, which was composed of rhamnose and galacturonic acid. The results of monosaccharide determination showed that it was composed of six monosaccharides: rhamnose, galacturonic acid, glucose, xylose, galactose, and arabinose. There was an inhibitory effect on A549 cells, and its mechanism might be related to the effect on the apoptosis gene Bcl-2/Bax. In summary, ASPF has a certain in vitro inhibitory effect on human lung cancer cells.

Key words: Acanthopanax senticosus fructification, polysaccharide, purification, structural characterization, anti-lung cancer

CLC Number:

R284.2

CUI Xuejiao, TONG Xiaoyu, ZHANG Yanlong, ZENG Weimin. Polysaccharide ASPF from Acanthopanax senticosus Fructification: Structural Characterization and Its Anti-lung Cancer Activity in Vitro[J]. Chinese Agricultural Science Bulletin, 2022, 38(22): 157-164.

Figures/Tables 14

References 26

[1]	叶强, 张阳, 张玉姣, 等. 国内五加属植物资源人工栽培的研究现状及展望[J]. 黑龙江农业科学, 2018(11):163-166.
[2]	SHI X L, YANG Y, REN H X, et al. Identification of multiple components in deep eutectic solvent extract of Acanthopanax senticosus root by ultra-high-performance liquid chromatography with quadrupole orbitrap mass spectrometry[J]. Phytochemistry letters, 2020, 35(4):175-185. doi: 10.1016/j.phytol.2019.11.017 URL
[3]	HAO B H, HAI P L, HAI M L, et al. Phytochemical and chemotaxonomic investigations on Acanthopanax leucorrhizus Harms[J]. Biochemical systematics and ecology, 2018, 81(3):21-26. doi: 10.1016/j.bse.2018.08.004 URL
[4]	LI X J, ZOU Q P, WANG X, et al. Lupane triterpenes from the leaves of Acanthopanax gracilistylus[J]. Molecules, 2018, 23(1):87. doi: 10.3390/molecules23010087 URL
[5]	DANIEL Z, MARTA O, AGNIESZKA G, et al. Phytochemical content and pharma-Nutrition study on Eleutherococcus senticosus fruits intractum[J]. Oxidative medicine & cellular longevity, 2016, 12(11):10-15.
[6]	MENG Q, PAN J, LIU Y, et al. Anti-tumour effects of polysaccharide extracted from Acanthopanax senticosus and cell-mediated immunity[J]. Experimental and therapeutic medicine, 2018, 15(2):1694.
[7]	QING L, ZHI T C, ZHI L X, et al. Binding of the polysaccharide from Acanthopanax giraldii Harms to toll-like receptor 4 activates macrophages[J]. Journal of ethnopharmacology, 2019, 241(12):112011. doi: 10.1016/j.jep.2019.112011 URL
[8]	HUANG L Z, HUANG B K, YE Q, et al. Bioactivity-guided fractionation for anti-fatigue property of Acanthopanax senticosus[J]. Journal of ethnopharmacology, 2010, 133(1):213-219. doi: 10.1016/j.jep.2010.09.032 URL
[9]	ZHAO Z Y, XU X J, YE Q W, et al. Ultrasound extraction optimization of Acanthopanax senticosus polysaccharides and its antioxidant activity[J]. International journal of biological macromolecules, 2013, 59(8):290-294. doi: 10.1016/j.ijbiomac.2013.04.067 URL
[10]	HU H B, LIANG H P, WU Y. Isolation, purification and structural characterization of polysaccharide from Acanthopanax brachypus[J]. Carbohydrate polymers, 2015, 127(5):97-100.
[11]	XIA Y G, HUANG Y X, LIANG J, et al. Comparable studies of two polysaccharides from leaves of Acanthopanax senticosus: Structure and antioxidation[J]. International journal of biological macromolecules, 2020, 147:350-362. doi: 10.1016/j.ijbiomac.2019.12.244 URL
[12]	丛登立, 王浩天, 高笑一, 等. 刺五加果的抗疲劳作用[J]. 吉林大学学报:医学版, 2010, 36(5):891-894.
[13]	YAN Z W, ZHOU M J, LIU J P, et al. Effects of total saponins from the pulp of Acanthopanax senticosus (Ruper. Et Maxim) Harms on blood lipidin mice with hyperlipidemia[J]. Special wild economic animal and plant research, 2009, 31(4):35-36.
[14]	LIANG C, XIU L X, HUI F, et al. Isolation and identification of anthocyanin component in the fruits of Acanthopanax sessiliflorus (Rupr. & Maxim.) Seem. by means of high speed counter current chromatography and evaluation of its antioxidant activity[J]. Molecules, 2020, 25(8):20-34. doi: 10.3390/molecules25010020 URL
[15]	HE W B, XU J, SHAO X R, et al. Study on recipe and technology of Changbai mountain Acanthopanax fruit cake[J]. Science and technology of food industry, 2012, 33(5):289-291.
[16]	YANG Z H, HAN Y H, MENG X, et al. Optimization in phenol-sulfuric acid method for determination of content of Paulowniae flos polysaccharide by response surface methodology[J]. Chemistry & bioengineering, 2020, 37(7):62-68.
[17]	侴晓玉, 王树成, 高娃, 等. 粮食中蛋白质含量的测定[J]. 广东化工, 2017, 44(4):114-115.
[18]	LI Y P, ZHOU H L. Progress on determination content of uronic acid in polysaccharides[J]. Food research and development, 2019, 49(17):207-211.
[19]	HELFER B, BOSWELL B C, FINLAY D, et al. Caspase-8 promotes cell motility and calpain activity under nonapoptotic conditions[J]. Cancer research, 2006, 66(8):4273-4278. doi: 10.1158/0008-5472.CAN-05-4183 URL
[20]	柳叶, 陈龙云. 黄芪多糖对人肺癌顺铂耐药株A549/DDP细胞耐药性的影响及其机制[J]. 吉林大学学报:医学版, 2020, 46(6):1162-1168.
[21]	肖小龙, 莫观, 廉云, 等. 大分子灵芝多糖对非小细胞肺癌细胞增殖、迁移和侵袭能力的抑制作用[J]. 上海中医药大学学报, 2018, 32(6):56-60.
[22]	赵俊霞, 闫永鑫, 赵娟, 等. 刺五加多糖诱导人小细胞肺癌H446细胞凋亡[J]. 细胞生物学杂志, 2008(2):239-242.
[23]	AHMAD B F R, ALI S S, AL-SHALI R A, et al. Vitamin E and 5-amino salicylic acid ameliorates acrylamide-induced peripheral neuropathy by inhibiting caspase-3 and inducible nitric oxide synthase immunoexpression[J]. Journal of chemical neuroanatomy, 2021, 113:406-413.
[24]	KULYAR M F, YAO W, DING Y, et al. Cluster of differentiation 147 (CD147) expression is linked with thiram induced chondrocyte's apoptosis via Bcl-2/Bax/Caspase-3 signalling in tibial growth plate under chlorogenic acid repercussion[J]. Ecotoxicology and environmental safety, 2021, 213:104-121.
[25]	NOORI A R, TASHAKOR A, NIKKHAH M, et al. Loss of WD2 subdomain of Apaf-1 forms an apoptosome structure which blocks activation of caspase-3 and caspase-9[J]. Biochimie, 2021, 180(3):46-85.
[26]	丁浩淼, 洪嘉瑶, 陈雪佳, 等. 羊栖菜多糖抗肿瘤及其作用机制研究进展[J]. 海洋科学, 2020, 44(3):129-137.

基因名称		序列(5'→3')
Bcl-2	上游	GTGGAGGAGCTCTTCAGGGA
Bcl-2	下游	CCGGTTCAGGTACTCAGTCATC
Bax	上游	AGGATGATTGCCGCCGT
Bax	下游	GGCCTTGAGCACCAGTTTG
β-actin	上游	TAGTTGCGTTACACCCTTTCTTG
β-actin	下游	TCACCTTCACCGTTCCAGTTT

基因名称		序列(5'→3')
Bcl-2	上游	GTGGAGGAGCTCTTCAGGGA
Bcl-2	下游	CCGGTTCAGGTACTCAGTCATC
Bax	上游	AGGATGATTGCCGCCGT
Bax	下游	GGCCTTGAGCACCAGTTTG
β-actin	上游	TAGTTGCGTTACACCCTTTCTTG
β-actin	下游	TCACCTTCACCGTTCCAGTTT

单糖	标准品保留时间/min	样品保留时间/min
Rha	26.707	26.755
Gal UA	35.985	36.011
Glc	42.225	42.253
Xyl	48.826	48.851
Gal	51.373	51.413
Ara	53.829	53.866

单糖	标准品保留时间/min	样品保留时间/min
Rha	26.707	26.755
Gal UA	35.985	36.011
Glc	42.225	42.253
Xyl	48.826	48.851
Gal	51.373	51.413
Ara	53.829	53.866