Establishment of Hairy Root Cultures by Agrobacterium Rhizogenes Mediated Transformation of Isatis Tinctoria L. for the Efficient Production of Flavonoids and Evaluation of Antioxidant Activities

• Published in: PloS one Vol. 10; no. 3; p. e0119022
• Main Authors: Gai, Qing-Yan, Jiao, Jiao, Luo, Meng, Wei, Zuo-Fu, Zu, Yuan-Gang, Ma, Wei, Fu, Yu-Jie
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 18.03.2015; Public Library of Science (PLoS)
• Subjects: Agrobacterium - genetics; Agrobacterium - metabolism; Analysis; Antioxidants; Antioxidants (Nutrients); Antioxidants - chemistry; Antioxidants - metabolism; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; beta-Glucosidase - genetics; beta-Glucosidase - metabolism; Biphenyl Compounds - antagonists & inhibitors; Biphenyl Compounds - metabolism; Carrier Proteins - genetics; Carrier Proteins - metabolism; Chinese medicine; Collaboration; Culture Media - chemistry; Culture Media - pharmacology; Design optimization; Factor Analysis, Statistical; Flavonoids; Flavonoids - biosynthesis; Flavonoids - chemistry; Food; Forestry; Gene Expression; Genetic transformation; Hairy root; Isatis - genetics; Isatis - metabolism; Isatis - microbiology; Isatis tinctoria; Isoflavones; Kaempferol; Liquiritigenin; Metabolites; Old fields; Pharmaceuticals; Phytochemicals; Picrates - antagonists & inhibitors; Picrates - metabolism; Plant ecology; Plant Roots - genetics; Plant Roots - metabolism; Plant Roots - microbiology; Plants, Genetically Modified; Quercetin; Roots; Rutin; Seeds; Tandem Mass Spectrometry; Tissue Culture Techniques - methods; Traditional Chinese medicine; Transformation; Transformation, Genetic; Beijing China; China
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0119022

In this work, Isatis tinctoria hairy root cultures (ITHRCs) were established as an alternative source for flavonoids (FL) production. I. tinctoria hairy root line V was found to be the most efficient line and was further confirmed by the PCR amplification of rolB, rolC and aux1 genes. Culture parameters of ITHRCs were optimized by Box-Behnken design (BBD), and eight bioactive FL constituents (rutin, neohesperidin, buddleoside, liquiritigenin, quercetin, isorhamnetin, kaempferol and isoliquiritigenin) were quali-quantitatively determined by LC-MS/MS. Under optimal conditions, the total FL accumulation of ITHRCs (24 day-old) achieved was 438.10 μg/g dry weight (DW), which exhibited significant superiority as against that of 2 year-old field grown roots (341.73 μg/g DW). Additionally, in vitro antioxidant assays demonstrated that ITHRCs extracts exhibited better antioxidant activities with lower IC₅₀ values (0.41 and 0.39, mg/mL) as compared to those of field grown roots (0.56 and 0.48, mg/mL). To the best of our knowledge, this is the first report describing FL production and antioxidant activities from ITHRCs.