北方农业学报 ›› 2024, Vol. 52 ›› Issue (5): 59-70.doi: 10.12190/j.issn.2096-1197.2024.05.07

• 分子生物学·生理生化 • 上一篇    下一篇

鹰嘴豆黄酮类化合物反相高效液相色谱法的建立及其体外药效评价

海婷玉1,2, 卫丁一1,2, 丁昱1,2, 熊青山1,2, 何子涵1,2, 戴小华1,2, 吴星霖1   

  1. 1.新疆农业大学,新疆 乌鲁木齐 830052;
    2.新疆草食动物新药研究与创制重点实验室,新疆 乌鲁木齐 830052
  • 收稿日期:2024-06-11 出版日期:2024-10-20 发布日期:2025-01-14
  • 通讯作者: 吴星霖(1976—),男,副教授,硕士,主要从事基础兽医方面的研究工作。戴小华(1976—),女,副教授,博士,主要从事兽医药理学与毒理学及天然产物功能性方面的研究工作。
  • 作者简介:海婷玉(1998—),女,硕士研究生,研究方向为兽医药理学与毒理学及天然产物功能性。
  • 基金资助:
    国家自然科学基金项目(31960718); 2021年新疆农业大学研究生科研创新项目(XJAUGRI2021032)

Establishment of reversed phase high performance liquid chromatography for flavonoids in chickpeas and evaluation of their in vitro efficacy

HAI Tingyu1,2, WEI Dingyi1,2, DING Yu1,2, XIONG Qingshan1,2, HE Zihan1,2, DAI Xiaohua1,2, WU Xinglin1   

  1. 1. Xinjiang Agriculture University,Urumqi 830052,China;
    2. Xinjiang Key Laboratory of New Drug Study and Creation for Herbivorous Animal,Urumqi 830052,China
  • Received:2024-06-11 Online:2024-10-20 Published:2025-01-14

摘要: 【目的】建立鹰嘴豆黄酮类化合物有效成分的反相高效液相色谱方法,并通过抑菌试验评价鹰嘴豆和鹰嘴豆总黄酮的体外药效。【方法】运用反相高效液相色谱分析方法,采用 Zorbax SB-C18(4.6 mm×150.0 mm,5 μm)色谱柱,流动相A(甲醇)∶B(0.5%磷酸溶液)=45∶55,以0.8 mL/min流速进行等度洗脱,柱温30 ℃,利用高效液相色谱仪对精密度、稳定性、重复性及加样回收率进行验证。通过滤纸片扩散法、牛津杯法、微量肉汤稀释法评价鹰嘴豆(100 mg/mL)和鹰嘴豆总黄酮(20 mg/mL)的抑菌效果。【结果】等度洗脱后,共分离出芦丁、柚皮苷二氢查尔酮、槲皮素3种黄酮成分,其色谱峰在30 min内均分离度良好,线性范围为0.003 75~0.060 00 mg/mL(R2为0.996~0.999);稳定性和重复性试验结果表明,芦丁含量为0.032 mg/g,柚皮苷二氢查尔酮含量为0.052 mg/g,槲皮素未检出。抑菌效果试验表明,滤纸片扩散法和牛津杯法检测鹰嘴豆和鹰嘴豆总黄酮均未出现抑菌圈,微量肉汤稀释法的孔板均浑浊。【结论】本试验选用的反相高效液相色谱法具有一定可靠性,适用于鹰嘴豆中芦丁和柚皮苷二氢查尔酮2种黄酮类化合物的分离测定;通过3种抑菌方法验证,鹰嘴豆和鹰嘴豆总黄酮均未出现抑菌作用。

关键词: 鹰嘴豆, 总黄酮, 反相高效液相色谱法, 抑菌效果

Abstract: 【Objective】To establish a reversed phase high performance liquid chromatography method for the effective components of chickpea flavonoids,and evaluate the in vitro pharmacological effects of chickpeas and chickpea total flavonoids through antibacterial tests.【Methods】Reversed phase high performance liquid chromatography was used for analysis,Zorbax SB-C18(4.6 mm×150.0 mm,5 μm)column was used,with mobile phase A(methanol)∶B(0.5% phosphoric acid aqueous solution)=45∶55. The flow rate was 0.8 mL/min for equal elution,and the column temperature was 30 ℃. The precision,stability,repeatability and sample recovery were verified by HPLC. Filter paper diffusion method,Oxford cup method and micro broth dilution method were used to investigate the antibacterial effect of chickpea(100 mg/mL) and chickpea total flavonoids(20 mg/mL).【Results】After equal elution,three flavonoids,rutin,naringin dihydrochalcone,and quercetin were well separated,and their chromatographic peaks showed good separation within 30 min,with a linear range of 0.003 75-0.060 00 mg/mL(R2=0.996-0.999). The stability and repeatability test results indicate that,the content of rutin was 0.032 mg/g,and the content of naringin dihydrochalcone was 0.052 mg/g,quercetin was not detected. In the antibacterial effect experiment,the filter paper diffusion method and Oxford cup method did not appear bacteriostatic circle,and the pore plates of the micro broth dilution method bacteriostatic experiment were muddy.【Conclusion】The reversed phase high performance liquid chromatography method used in this experiment has certain reliability and is suitable for the separation and determination of two flavonoids,rutin and naringin dihydrochalcone in chickpeas. Through the verification of three antibacterial methods,chickpea and chickpea total flavonoids did not have obvious antibacterial effect.

Key words: Chickpeas, Total flavonoids, Reversed phase high performance liquid chromatography, Antibacterial effect

中图分类号: 

  • TQ28