Ingredient Analysis

Study on polysaccharide in Rehmanniae Radix and Rehmanniae Radix Praeparata by high performance liquid chromatography with pre-column derivatization

Expand
  • 1. National Institutes for Food and Drug Control, Beijing 102629, China;
    2. Infinitus (China) Co., Ltd., Jiangmen 529100, China

Revised date: 2023-03-30

  Online published: 2024-06-25

Abstract

Objective: To determine the monosaccharide composition and contents of polysaccharides in Rehmanniae Radix and Rehmanniae Radix Praeparata, and analyze the differences between them. Methods: The extracted polysaccharide were hydrolyzed by trifluoroacetic acid, and the monosaccharides were derivatized by 1-phenyl-3-methyl-5-pyrazolone(PMP). Waters Symmetry-C18 column (250 mm×4.6 mm, 5 μm) was adopted, with gradient elution of 0.05 mol·L-1 phosphate buffer (pH 6.8, mobile phase A) and acetonitrile (mobile phase B). The flow rate was 1.0 mL·min-1, the detection wavelength was 250 nm, and the column temperature was 30 ℃. Principal component analysis and variance analysis were used to analyze the contents and ratios of monosaccharides after hydrolysis of Rehmanniae Radix and Rehmanniae Radix Praeparata. Results: Weights of galacturonic acid, glucose, galactose and arabinose showed a good linear relationship with the peak areas in the range of 0.02-1.82、0.10-3.91、0.09-7.08 and 0.03-1.69 μg, respectively (r>0.999 3). The recovery rates were from 97.6% to 103.7%. The RSDs (n=6) for repeatability tests were from 3.2% to 4.7%. The intermediate precision met the requirements. After hydrolysis, the polysaccharides in Rehmanniae Radix and Rehmanniae Radix Praeparata both contained the four monosaccharides above with no significant difference between them. But the ratio of glucose content to galactose content in Rehmanniae Radix was greater than 0.4, and that in Rehmanniae Radix Praeparata was basically less than 0.4. Conclusion: The method is reproducible and accurate, and suitable for the determination of monosaccharides which are the hydrolysates of polysaccharides in Rehmanniae Radix and Rehmanniae Radix Praeparata. There are some differences between Rehmanniae Radix and Rehmanniae Radix Praeparata on the ratio of glucose content to galactose content.

Cite this article

LI Ming-hua, SUN Hong-mei, CHENG Xian-long, WEI Feng, MA Shuang-cheng . Study on polysaccharide in Rehmanniae Radix and Rehmanniae Radix Praeparata by high performance liquid chromatography with pre-column derivatization[J]. Chinese Journal of Pharmaceutical Analysis, 2023 , 43(4) : 602 -611 . DOI: 10.16155/j.0254-1793.2023.04.09

References

[1] 中华人民共和国药典2020年版. 一部[S].2020: 129, 130
ChP 2020. Vol Ⅰ[S].2020: 129, 130
[2] 王志江, 魏国栋, 马思缇. 地黄多糖的化学和药理作用研究进展[J].中国实验方剂学杂志, 2015, 21(16): 231
WANG ZJ, WEI GD, MA ST. Chemical and pharmacological effects of Rehmanniae Radix polysaccharides[J].Chin J Exp Tradit Med Form, 2015, 21(16): 231
[3] 祁小妮, 林楠楠, 李振亮, 等. 生地黄多糖提取工艺的优化及抗氧化活性研究[J].食品与发酵工业, 2016, 42(2): 231
QI XN, LIN NN, LI ZL, et al. Optimization the extraction process of polysaccharide by response surface methodology from the Rehmannia and its antioxidant activity[J].Food Ferment Ind, 2016, 42(2): 231
[4] 崔瑛, 潘贝, 史银玥, 等. 生地黄及生地黄多糖在制备抗焦虑药物中的应用:中国, 201910454786.7[P].2019-08-02
CUI Y, PAN B, SHI YY, et al. Application of Rehmannia glutinosa Polysaccharides in Preparation of Anti-anxiety Drugs: China, 201910454786.7[P].2019-08-02
[5] 张副兴, 裘生梁, 祝雨田, 等. 熟地黄多糖提取技术及药理作用研究进展[J].浙江中西医结合杂志, 2013, 23(2): 153
ZHANG FX, QIU SL, ZHU YT, et al. Research progress on extraction technology and pharmacological action of polysaccharide from Radix Rehmanniae Praeparata[J].Zhejiang J Integr Tradit Chin West Med, 2013, 23(2): 153
[6] 王赵, 赵剑锋, 王莹, 等. 注射用黄芪多糖单糖鉴别及游离单糖检查方法研究[J].中国药业, 2018, 27(11): 21
WANG Z, ZHAO QF, WANG Y, et al. Study on method of identification of monosaccharide and inspection of free monosaccharide of Astragalus polysaccharide for injection[J].China Pharm, 2018, 27(11): 21
[7] 杨毅, 李文燕, 郭丽, 等. HPGPC联合HPLC-ELSD测定樟芝子实体中多糖分子量、多糖组成和单糖含量[J].中药材, 2018, 41(5):1145
YANG Y, LI WY, GUO L, et al. HPGPC combined HPLC-ELSD to determine polysaccharide molecular weight, polysaccharide composition and monosaccharide content in Ganoderma lucidum fruiting body[J].J Chin Med Mater, 2018, 41(5): 1145
[8] 常青. 基于高效液相色谱法优化测定7种混合单糖的研究[J].福建轻纺, 2018(10): 25
CHANG Q. Study on optimal determination of 7 kinds of mixed monosaccharides based on high performance liquid chromatography[J].Fujian Text, 2018(10): 25
[9] 孙红梅, 李振, 彭喜春, 等. 不同产地枸杞子中多糖含量的比较研究[J].中国食物与营养, 2021, 27(3): 58
SUN HM, LI Z, PENG XC, et al. Comparative study on polysaccharide content in Lycii Fructus from different habitats[J].Food Nutrit China, 2021, 27(3): 58
[10] 吴诗云, 钟益宁, 张焱, 等. 熟地黄多糖含量测定方法研究[J].安徽农业科学, 2016, 44(35): 154
WU SY, ZHONG YN, ZHANG Y, et al. Study on the determination methods of polysaccharide content of Radix Rehmanniae Preparata[J].J Anhui Agric Sci, 2016, 44(35): 154
[11] 张艳萍, 俞远志, 张虹. 气相色谱分析生地黄多糖的单糖组成及其含量[J].中国中药杂志, 2009, 34(4): 419
ZHANG YP, YU YZ, ZHANG H. Analysis of monosaccharides in Radix Rehmanniae by GC[J].China J Chin Mater Med, 2009, 34(4): 419
[12] USP 43-NF 38[S].2020
[13] 张婷, 孙红梅, 李振, 等. 响应面法优化水提醇沉玄参多糖工艺及其含量分析[J].中国现代中药, 2021, 23(4): 679
ZHANG T, SUN HM, LI Z, et al. Response surface methodology for optimization of water extract and alcohol precipitate and content analysis of Scrophulariae Radix polysaccharides[J].Mod Chin Med, 2021, 23(4): 679
[14] 符梦凡, 赵一帆, 阎卫东. 柱前衍生化HPLC法分析枸杞多糖中单糖组成[J].食品科学, 2018, 39(18): 186
FU MF, ZHAO YF, YAN WD. Analysis of monosaccharide composition of polysaccharides from Lycium barbarum L. by HPLC with precolumn derivatization[J].Food Sci, 2018, 39(18): 186
[15] 成显波. 柱前衍生化高效液相色谱法测定多糖水解液中葡萄糖含量[J].轻工科技, 2021, 37(9): 3
CHENG XB. Study on the determination methods of glucose content of polysaccharide hydrolysis by HPLC with precolumn derivatization[J].Light Ind Sci Technol, 2021, 37(9): 3
[16] 马定远, 陈君, 李萍, 等. 柱前衍生化高效液相色谱法分析多糖中的单糖组成[J].分析化学, 2002, 30(6): 702
MA DY, CHEN J, LI P, et al. Analysis of monosacchaaride compositions in polysaccharides by pre-column derivatization high performance liquid chromatography[J].Chin J Anal Chem, 2002, 30(6): 702
[17] 赵孟欣, 王泽岚, 孟哲, 等. 温和条件下柱前标记-高效液相色谱-质谱法测定枸杞多糖中单糖组成[J].色谱, 2019, 37(11): 1162
ZHAO MX, WANG ZL, MENG Z, et al. Analysis of monosaccharide composition in polysaccharides from Lycium barbarum L. by precolumn labeling under mild conditions and high-performance liquid chromatography coupled with mass spectrometry[J].Chin J Chromatogr, 2019, 37(11): 1162
[18] 朱彩平, 张声华. 枸杞多糖的提取及其组成的气相色谱分析[J].现代食品科技, 2009, 25(11): 1327
ZHU CP, ZHANG SH. Extraction and composition of Lycium barbarum polysaccharide by GC[J].Mod Food Sci Technol, 2009, 25(11): 1327
[19] 林好, 桂蜀华, 于兵兵, 等. 不同炮制工艺熟地黄多糖单糖分析及其对卵巢颗粒细胞的影响[J].中成药, 2019, 41(12): 2958
LIN H, GUI SH, YU BB, et al. Analysis of polysaccharides and monosaccharides from Rehmannia glutinosa by different processing technology and effects on ovarian granulosa cells[J].Chin Tradit Pat Med, 2019, 41(12): 2958
[20] 张晓亚, 徐金娣, 许军, 等. 整合糖组与代谢组学方法比较蒸制和酒炖熟地黄化学成分[J].中草药, 2021,52(6): 1591
ZHANG XY, XU JD, XU J, et al. Integrating glycomics and metabolomics to compare chemical profiles of steaming and wine-stewing processed Rehmanniae Radix Praeparata[J].Chin Tradit Herb Drugs, 2021, 52(6): 1591
[21] 李军, 张丽萍, 张振凌, 等. 熟地黄清蒸和酒炖不同时间还原糖含量测定[J].中成药, 2006, 28(4):513
LI J, ZHANG LP, ZHANG ZL, et al. Determination of reducing sugar content in cooked Rehmannia glutinosa steamed and stewed with wine at different times[J].Chin Tradit Pat Med, 2006, 28(4): 513
[22] 孟祥龙, 王勃, 胡聪, 等. 基于HPLC-MS及网络药理学探讨熟地黄炮制过程与美拉德反应的相关性[J].中药材, 2020,43(1):61
MENG XL, WANG B, HU C, et al. Study on the correlation between the processing process of Radix Rehmanniae Praeparata and Maillard reaction based on HPLC-MS and network pharmacology[J].J Chin Med Mater, 2020, 43(1): 61
Outlines

/