多糖部分降解产物在板蓝根颗粒多糖成分分析中的应用

doi:10.3969/j.issn.1000-1565.2021.01.006

摘要/Abstract

摘要： 板蓝根多糖的药理活性与板蓝根的功效一致,其部分降解产物的毛细管区带电泳(CZE)图谱特征性强,可用于板蓝根颗粒中多糖类成分的定性分析.首先采用酶法去除板蓝根颗粒中糊精等辅料的干扰,所得板蓝根多糖在盐酸浓度0.5 mol/L,温度80 ℃条件下部分降解4 h,以1-苯基-3-甲基-5-吡唑啉酮(PMP)为衍生试剂,采用毛细管区带电泳法测定其部分降解产物图谱.电泳条件:未涂层弹性石英毛细管柱(70 cm×50 μm, i.d,有效长度61 cm),运行缓冲液为40 mmol/L硼砂缓冲液(pH 10.1),检测波长245 nm,运行电压17 kV,室温下重力进样10 cm×15 s,进而运用聚类分析(CA)对该产物的CZE谱图进一步探讨.结果表明:CZE图谱在低聚糖部分具有差异,而单糖组成相对稳定,均含木糖、阿拉伯糖、葡萄糖、鼠李糖、甘露糖和半乳糖等,结合板蓝根药材相应谱图,从低聚糖和单糖上进行比对可达到鉴别目的,且发现不同厂家产品之间多糖含量存在较大差异.板蓝根颗粒中多糖部分降解产物CZE图谱的建立完善了该药品的质量控制,同时也为其药效物质研究及其他相关中药制剂中多糖的质量控制提供参考.

关键词: 板蓝根颗粒, 多糖, 部分降解, 鉴别, 毛细管区带电泳, 聚类分析

Abstract: The pharmacological activities of Radix isatidis polysaccharide are consistent with the efficacy of R. isatidis. As the capillary zone electrophoresis(CZE)spectrum of partial degradation products of R. isatidis polysaccharide has obvious characteristics, and can be used for the qualitative analysis of polysaccharide from Banlangen Granules. In this paper, the interference of pharmaceutical excipients such as dextrin from Banlangen Granules was firstly removed by enzymatic method. Then the obtained R. isatidis polysaccharide was partially degraded for 4 h at a hydrochloric acid concentration of 0.5 mol/L and a temperature of 80 ℃, and the spectra were determined by CZE using 1-phenyl-3-methyl-5-pyrazolone(PMP)as- DOI:10.3969/j.issn.1000-1565.2021.01.006多糖部分降解产物在板蓝根颗粒多糖成分分析中的应用张建,卢凯,石晶,郭怀忠(河北大学药学院,河北省药物质量分析控制重点实验室,河北保定 071002)摘要:板蓝根多糖的药理活性与板蓝根的功效一致,其部分降解产物的毛细管区带电泳(CZE)图谱特征性强,可用于板蓝根颗粒中多糖类成分的定性分析.首先采用酶法去除板蓝根颗粒中糊精等辅料的干扰,所得板蓝根多糖在盐酸浓度0.5 mol/L,温度80 ℃条件下部分降解4 h,以1-苯基-3-甲基-5-吡唑啉酮(PMP)为衍生试剂,采用毛细管区带电泳法测定其部分降解产物图谱.电泳条件:未涂层弹性石英毛细管柱(70 cm×50 μm, i.d,有效长度61 cm),运行缓冲液为40 mmol/L硼砂缓冲液(pH 10.1),检测波长245 nm,运行电压17 kV,室温下重力进样10 cm×15 s,进而运用聚类分析(CA)对该产物的CZE谱图进一步探讨.结果表明:CZE图谱在低聚糖部分具有差异,而单糖组成相对稳定,均含木糖、阿拉伯糖、葡萄糖、鼠李糖、甘露糖和半乳糖等,结合板蓝根药材相应谱图,从低聚糖和单糖上进行比对可达到鉴别目的,且发现不同厂家产品之间多糖含量存在较大差异.板蓝根颗粒中多糖部分降解产物CZE图谱的建立完善了该药品的质量控制,同时也为其药效物质研究及其他相关中药制剂中多糖的质量控制提供参考.关键词:板蓝根颗粒;多糖;部分降解;鉴别;毛细管区带电泳;聚类分析中图分类号:O657.8 文献标志码:A 文章编号:1000-1565(2021)01-0040-07Application of partial degradation products of polysaccharide in analysis of polysaccharide from Banlangen GranulesZHANG Jian, LU Kai, SHI Jing, GUO Huaizhong(Key Laboratory of Pharmaceutical Quality Control of Hebei Province,College of Pharmacy, Hebei University, Baoding 071002, China)Abstract: The pharmacological activities of Radix isatidis polysaccharide are consistent with the efficacy of R. isatidis. As the capillary zone electrophoresis(CZE)spectrum of partial degradation products of R. isatidis polysaccharide has obvious characteristics, and can be used for the qualitative analysis of polysaccharide from Banlangen Granules. In this paper, the interference of pharmaceutical excipients such as dextrin from Banlangen Granules was firstly removed by enzymatic method. Then the obtained R. isatidis polysaccharide was partially degraded for 4 h at a hydrochloric acid concentration of 0.5 mol/L and a temperature of 80 ℃, and the spectra were determined by CZE using 1-phenyl-3-methyl-5-pyrazolone(PMP)as- 收稿日期:2019-12-24 基金项目:国家自然科学基金资助项目(21575033);河北省自然科学基金资助项目(H2016201221;B2018201270) 第一作者:张建(1993—),男,河南信阳人,河北大学在读硕士研究生.E-mail:zjian201709@163.com 通信作者:郭怀忠(1968—),男,内蒙古商都人,河北大学教授,主要从事中药质量控制及其药效学研究.E-mail: ghuaizh@aliyun.com第1期张建等:多糖部分降解产物在板蓝根颗粒多糖成分分析中的应用a derivatizing reagent. The electrophoresis conditions were as follows: uncoated fused silica capillary column(70 cm×50 μm i.d., 61 cm effective length), 40 mmol/L borax solution(pH 10.1)was used as the running buffer, the detection wavelength was set at 245 nm, the operation voltage was 17 kV, hydrodynamic pressure injection was employed(10 cm×15 s). The CZE spectra of partial degradation products were further explored using cluster analysis(CA). The results showed that the CZE spectra had some differences in oligosaccharide composition, while monosaccharide composition was relatively stable, including xylose, arabinose, glucose, rhamnose, mannose and galactose. Combined with the corresponding spectrum of R. isatidis, we could achieve identification purpose through comparison between their oligosaccharides and monosaccharides, and there were significant differences in polysaccharide contents among the products of different manufacturers. The establishment of CZE spectrum of the partial degradation products of polysaccharide from Banlangen Granules has improved the quality control of the drug, and also provided reference for the study of the pharmacodynamic substances and the quality control of polysaccharide in other related Chinese medicine preparations.

Key words: Banlangen Granules, polysaccharide, partial degradation, identification, capillary zone electrophoresis, cluster analysis

中图分类号:

O657.8

张建,卢凯,石晶,郭怀忠. 多糖部分降解产物在板蓝根颗粒多糖成分分析中的应用[J]. 河北大学学报(自然科学版), 2021, 41(1): 40-46.

ZHANG Jian, LU Kai, SHI Jing, GUO Huaizhong. Application of partial degradation products of polysaccharide in analysis of polysaccharide from Banlangen Granules[J]. Journal of Hebei University(Natural Science Edition), 2021, 41(1): 40-46.

参考文献

[1] 国家药典委员会. 《中华人民共和国药典》一部[M]. 北京:中国医药科技出版社,2015:1033.
[2] 马莉,孙琴,李友,等. HPLC法测定板蓝根药材及制剂中靛蓝和靛玉红含量[J]. 药物分析杂志,2010,30(09):1642-1645. DOI: 10.16155/j.0254-1793.2010.09.032.
[3] 辛敏通,傅欣彤,陈有根,等. 柱前衍生化UPLC测定板蓝根颗粒中主要游离氨基酸的含量[J]. 中国中药杂志,2011,36(23):3306-3309. DOI: 10.4268/cjcmm20112319.
[4] 肖春霞,黄晓婧,文永盛,等. HPLC法同时测定板蓝根颗粒中腺苷和(R,S)-告依春的含量[J]. 中国药物评价,2017,34(2):81-85. DOI: 10.3969/j.issn.2095-3593.2017.02.001.
[5] 王宁莉,周振旗,贾万仓. 板蓝根饮片、板蓝根颗粒中(R,S)-告依春的含量测定[J]. 中国药业,2018,27(3):30-32. DOI:10.3969/ j.issn.1006-4931.2018.03.009.
[6] 陈吉业,郭怀忠,马会英,等. 板蓝根颗粒中多糖的提取优化及单糖组成分析[J]. 中国医药工业杂志,2014,45(9):836-839. DOI: 10.16522/j.cnki.cjph.2014.09.004.
[7] 高艳梅,谷俊峰. HPLC测定板蓝根颗粒中表告依春的含量[J]. 中国民族民间医药,2019,28(2):17-18.
[8] WANG C M, RUAN S M, GU X M, et al. Antiviral activities of Radix Isatidis polysaccharide against type II herpes simplex virus in vitro [J]. Food Science and Technology, 2018, 38(1): 180-183. DOI: 10.1590/1678-457X.19317.
[9] 贾建,曹荣安,杜亿华,等. 板蓝根多糖研究进展[J]. 食品工业科技,2016,37(18):378-383. DOI: 10.13386/j.issn1002-0306.2016.18.064.
[10] 赵春霞,余河水,王春华,等. 中药中糖类定性及定量研究进展[J]. 中成药,2019,41(9):2184-2192. DOI: 10.3969/j.issn.1001-1528.2019.09.034.
[11] 李晓霞,王迪,王桂臻,等. 基于内切-1,4-β-半乳聚糖酶水解的黄芪糖指纹图谱的建立及不同种质资源黄芪的鉴别[J].中药材,2018,41(6):1306-1311. DOI: 10.13863/j.issn1001-4454.2018.06.012.
[12] JING P, ZHAO S J, LU M M, et al. Multiple-Fingerprint analysis for investigating quality control of Flammulina velutipes fruiting body polysaccharides [J]. Journal of Agricultural and Food Chemistry, 2014, 62(50): 12128-12133. DOI: 10.1021/jf504349r.
[13] 郭怀忠,吴芳,张伟泉,等. 聚类分析辅助中药寡糖电泳分析鉴定中药[J]. 色谱,2013,31(10):1001-1004. DOI: 10.3724/SP.J.1123.2013.03033.
[14] YANG F F, GUO H Z, ZHANG L S, et al. Identification of confusable herbal medicines by mapping of partial degradation products from Herbal medicine polysaccharides [J]. Analytical Sciences, 2018, 34(11): 1291-1296. DOI: 10.2116/analsci.18P058.
[15] 张秀媛,袁永俊,何扩. 糖化酶的研究概况[J]. 食品研究与开发,2006,27(9):163-166. DOI: 10.3969/j.issn.1005-6521.2006.09.054.
[16] 崔施展,贾东升,谢晓亮,等. 酶法测定黄芪含片中黄芪多糖的研究[J]. 食品研究与开发,2015,36(21):143-146. DOI: 10.3969/j.issn.1005-6521.2015.21.036.
[17] 贾东升,崔施展,谢晓亮,等. 含麦芽糊精枸杞提取物干粉中多糖含量测定方法[J]. 国际药学研究杂志,2015,42(4):527-530. DOI: 10.13220/j.cnki.jipr.2015.04.018.
[18] 贾东升,温春秀,赵江丽,等. 固体制剂中多糖含量测定的样品预处理方法研究[J]. 药物分析杂志,2015,35(3):409-413. DOI: 10.16155/j.0254-1793.2015.03.007.
[19] 张利沙,孙玮玮,徐自慧,等. 复方南板蓝根颗粒中多糖的含量测定及单糖组成分析[J]. 药物分析杂志,2017,37(10):1845-1850. DOI: 10.16155/j.0254-1793.2017.10.15.
[20] 郭怀忠,张斌,徐相涛,等. 板蓝根多糖的降解和单糖组成的毛细管区带电泳测定研究[J]. 中草药,2010,41(5):744-747.
[21] 杨飞飞,郭怀忠,王晓欢,等. 采用寡糖CZE指纹图谱鉴定北沙参和南沙参[J]. 中南药学,2015,13(5):455-457. DOI: 10.7539/j.issn.1672-2981.2015.05.002.
[22] 郭怀忠,董露,杨飞飞,等. 1-苯基-3-甲基-5-吡唑啉酮糖衍生物在反相高效液相色谱法分离中保留行为的探讨[J]. 中南药学,2017,15(04):511-514. DOI: 10.7539/j.issn.1672-2981.2017.04.029. (