J Agric Food Chem. 2015 Jan 14;63(1):262-8. doi: 10.1021/jf504176r.
Chlorogenic acid and its derivatives (CADs) are valuable bioactive plant secondary metabolites with many health benefits. In the present study, Stevia rebaudiana hairy root cultures were established, and the culture conditions for the production of CADs were optimized. The hairy roots were induced by coculture of S. rebaudiana leaves and Agrobacterium rhizogenes (C58C1) after infection, which were further verified by PCR detection of rolB and rolC genes. HPLC-MS and HPLC analysis showed that chlorogenic acid (3-caffeoylquinic acid, 3-CQA), 3,5-dicaffeoylquinic acid (3,5-CQA), and 4,5-dicaffeoylquinic acid (4,5-CQA) were the major CADs in the hairy roots. Eight single roots with rapid growth rate were selected. Among them, T3 had the highest yield of CADs. B5 medium supplemented with 40 g/L sucrose was more suitable for the production of CADs than others. Under optimal culture conditions, the total content of these three compounds reached 105.58 mg/g and total yield was 234.40 mg/100 mL.
绿原酸及其衍生物（CADs）是具有多种健康益处的有价值的植物次生代谢物。本研究建立了甜菊叶外植体毛状根培养体系，并对 CADs 的生产条件进行了优化。毛状根通过叶与发根农杆菌（C58C1）共培养诱导，经 rolB 和 rolC 基因 PCR 检测进一步验证。HPLC-MS 和 HPLC 分析表明，毛状根中主要的 CADs 为绿原酸（3-咖啡酰奎宁酸，3-CQA）、3,5-二咖啡酰奎宁酸（3,5-CQA）和 4,5-二咖啡酰奎宁酸（4,5-CQA）。选择了 8 株生长速度较快的单根，其中 T3 的 CADs 产量最高。B5 培养基添加 40 g/L 蔗糖比其他培养基更适合 CADs 的生产。在最佳培养条件下，这三种化合物的总量达到 105.58 mg/g，总收率达到 234.40 mg/100 mL。
