响应面法优化超声辅助低共熔溶剂提取葛根药渣总黄酮及其活性研究

Optimization of Ultrasound-Assisted Extraction of Total Flavonoids from Puerariae Radix Residue Using Deep Eutectic Solvents by Response Surface Methodology and Its Bioactivity Evaluation

关键词:  葛根  低共熔溶剂  工艺优化  抗炎活性  抗氧化活性

Key words:  Pueraria lobata  deep eutectic solvent  process optimization  anti-inflammatory activity  antioxidant activity

摘要:

目的 为了充分利用中药葛根药渣资源,实现效益最大化和生态保护。方法 采用超声辅助低共熔溶剂提取葛根药渣总黄酮；单因素试验和Box-Behnken法优化提取工艺；利用D101大孔吸附树脂回收溶剂中黄酮成分,开展低共熔溶剂回收再利用实验；采用液膜制片,检测三种提取溶剂傅里叶变换红外吸收光谱；DPPH法和ABTS法研究葛根总黄酮体外抗氧化活性；构建小鼠大脑中动脉阻塞(Middle Cerebral Artery Occlusion,MCAO)模型,ELISA法检测小鼠大脑皮质白细胞介素-1β(Interleukin-1 beta,IL-1β)、白细胞介素-6(Interleukin-6,IL-6)、肿瘤坏死因子-α(Tumor Necrosis Factor-alpha,TNF-α)炎症因子水平。结果 葛根药渣总黄酮最佳提取条件：氯化胆碱和乙二醇配比1 : 3.906、含水率为41.18%、液料比1 : 31.41、超声时间19.52 min,葛根药渣黄酮的平均提取率为4.557%；DES溶剂回收可循环利用三次；红外光谱DES吸收峰发生位移；DES提取液的自由基清除率显著高于水提取液和乙醇提取液；与Model组相比,DES组中小鼠大脑皮质IL-1β、IL-6、TNF-α水平显著降低(P<0.05)。结论 超声辅助低共熔溶剂能够有效提高葛根药渣总黄酮提取率和体内外活性,可实现绿色环保与高效利用,为后续中药葛根资源开发利用提供思路。

Abstract:

OBJECTIVE To fully utilize the herb residue resources of Pueraria lobata and maximize economic benefits while achieving ecological protection. METHODS Ultrasound-assisted deep eutectic solvent (DES) was adopted to extract total flavonoids from Pueraria lobata herb residues. Single-factor experiments and the Box-Behnken design were applied to optimize the extraction process. D101 macroporous adsorption resin was used to recover flavonoid components from the solvent, and experiments on the recovery and reuse of deep eutectic solvents were carried out. Liquid film sample preparation was employed to determine the Fourier transform infrared (FTIR) absorption spectra of the three extraction solvents. The in vitro antioxidant activities of total flavonoids from Pueraria lobata were evaluated by DPPH and ABTS radical scavenging assays. A mouse model of middle cerebral artery occlusion (MCAO) was established, and enzyme-linked immunosorbent assay (ELISA) was performed to detect the levels of interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in the cerebral cortex of mice. RESULTS The optimal extraction conditions for total flavonoids from Pueraria lobata herb residues were as follows: the ratio of choline chloride to ethylene glycol was 1:3.906, water content was 41.18%, liquid-to-material ratio was 1:31.41, and ultrasonic time was 19.52 min. Under these conditions, the average extraction rate of flavonoids from Pueraria lobata herb residues reached 4.557%. The recovered DES could be recycled for three times. The absorption peaks of DES showed a displacement phenomenon in the infrared spectrum. The radical scavenging rate of the DES extract was significantly higher than those of the water extract and ethanol extract. Compared with the model group, the levels of IL-1β, IL-6 and TNF-α in the DES group were significantly decreased (P<0.05). CONCLUSION Ultrasound-assisted deep eutectic solvent can effectively improve the extraction rate of total flavonoids from Pueraria lobata herb residues. Moreover, the flavonoid extract obtained by this method exhibits significantly higher anti-inflammatory and antioxidant activities than those extracted by traditional methods. This approach realizes the green, environmentally friendly and efficient utilization of herb residues, which provides a feasible strategy for the further development and utilization of Pueraria lobata resources.