广枣果皮总原花青素纯化工艺及成分鉴定研究

Purification and component identification of total proanthocyanidins in Choerospondias axillaris pericarp

该文以工业废弃物广枣果皮为研究对象,首先根据高效液相色谱法(HPLC)和质谱图谱比对结果,确定了广枣果皮总原花青素纯化工艺考察过程中的定量标志物。基于2个稳定存在的原花青素类成分为标志物,优化总原花青素的纯化工艺,考察了5种不同大孔吸附树脂的吸附和解吸附能力、NKA-Ⅱ树脂的静态吸附动力学曲线、最大上样量和洗脱梯度等参数,制备了广枣果皮总原花青素,并采用超高效液相色谱-四极杆飞行时间质谱联用技术(UPLC-Q-TOF-MS/MS)对其进行定性分析。结果表明,NKA-Ⅱ树脂对广枣果皮总原花青素的吸附和解吸附能力均较强。取广枣皮70%乙醇粗提物溶于水中,质量浓度为50 mg·mL^(-1),上样7倍柱体积(BV),静态吸附12 h,用8 BV蒸馏水和6 BV 10%乙醇除杂,然后用8 BV 50%乙醇洗脱,减压浓缩并干燥,即得纯化后的总原花青素粉末。用香草醛盐酸法测得总原花青素的纯度为47.67%,转移率为59.92%,优化工艺可行。UPLC-Q-TOF-MS/MS定性分析鉴定出了总原花青素中16个原花青素类成分。该研究优选的原花青素纯化工艺简单,成分鉴定与检测方法准确,可适用于难以分离纯化的某大类成分的工艺考察,可为中药废弃物中有用成分的综合利用提供技术支持和研究思路。

The present study determined the quantitative markers of total proanthocyanidins in the purification of the industrial waste Choerospondias axillaris pericarp based on the comparison results of high-performance liquid chromatography(HPLC)and mass spectrometry(MS)and optimized the purification process with two stable procyanidins as markers.The adsorption and desorption of five different macroporous adsorption resins,the static adsorption kinetics curve of NKA-Ⅱresin,the maximum sample load,and the gradient elution were investigated.The UPLC-Q-TOF-MS/MS was employed for qualitative analysis of the newly-prepared total proanthocyanidins of C.axillaris pericarp.As revealed by the results,NKA-Ⅱresin displayed strong adsorption and desorption toward total proanthocyanidins.The sample solution(50 mg·mL^(-1))was prepared from 70%ethanol crude extract of C.axillaris pericarp dissolved in water and 7-fold BV of the sample solution was loaded,followed by static adsorption for 12 h.After 8-fold BV of distilled water and 6-fold BV of 10%ethanol were employed to remove impurities,the solution was eluted with 8-fold BV of 50%ethanol,concentrated,and dried under reduced pressure,and purified total proanthocyanidin powder was therefore obtained.Measured by vanillin-hydrochloric acid method,the purity and transfer rate of total proanthocyanidins were 47.67%and 59.92%,respectively,indicating the feasibi-lity of the optimized process.UPLC-Q-TOF-MS/MS qualitative analysis identified 16 procyanidins in C.axillaris total proanthocyanidins.The optimized purification process is simple in operation and accurate in component identification,and it can be applied to the process investigation of a class of components that are difficult to be separated and purified.It can also provide technical support and research ideas for the comprehensive utilization of industrial waste.