绿原酸清洁提取及其体外生物活性评价

Eco-Friendly Extraction of Chlorogenic Acid and its Biological Activity Evaluation in Vitro

摘要:

植物纤维组织致密结构阻碍天然生物活性成分绿原酸的溶出，为减少绿原酸溶出的传质阻力，选取复配酶（纤维素酶和木质素酶）催化水解金银花叶溶浸获得绿原酸，并采用DPPH自由基清除法、铁氰化钾还原法评价提取物绿原酸的体外抗氧化性。分析酶解金银花叶后的SEM照片发现，酶能有效破坏植物纤维组织结构，有助于形成内部疏松结构，而金属离子对酶催化水解活性作用依次为Mg2+＞Na+＞Ca2+＞K2+＞Cu2+；选取纤维素酶?鄄木质素酶酶活比为3∶2的复配酶，在水浴温度45 ℃、pH 4的条件下，当复配酶投加量为50 U/g时，在Mg2+诱导催化水解2 g金银花叶11 h后，绿原酸浸提率达94.68%，比超声醇提法提高了19.60%，所得绿原酸具备高抗氧化性和还原性，为资源化利用金银花叶以及绿原酸生产提供了新途径。

Abstract:

The chlorogenic acid was a natural bioactive ingredient which was usually hindered in the dense structure of plant fibers and difficult to be extracted. An enzymatic cell wall-breaking process was thus developed using the combination of cellulase and ligninase to reduce transfer resistance during extraction and to improve the extraction efficiency of chlorogenic acid from Lonicera japonica leaves. The antioxidant activity in vitro of chlorogenic acid was evaluated by DPPH free radical scavenging method and ferricyanide reduction analysis. The product of enzymatic hydrolysis by the dual-enzyme complex was characterized by SEM. The complex was confirmed to effectively destruct the plant fiber structure and help to form the internal loose structure. The effect of various metal ions on the enzyme activity were weakened in the order of Mg2+，Na+，Ca2+，K+，and Cu2+. The optimized extraction condition was determined using 3∶2 of cellulase to ligninase ratio （U/U） with 50 U/g of enzymes/substrate and Mg2+ at 45 ℃ and pH 4.0 for 11 h. Under the optimal condition，the extraction rate of chlorogenic acid could reach as high as 94.68%，which was 19.60% higher than that of ultrasound-assisted ethanol extraction. The anti-oxidative of reductive activities of chlorogenic acid were improved. This study provides a theoretical foundation to the comprehensive utilization of Lonicera japonica leaves and industrialization of chlorogenic acid.