Establishment and chemical analysis of hairy roots of Eucommia ulmoides.

摘要

The bark of Eucommia ulmoides Oliver (Eucommiaceae) has been used over thousands of years as a folk remedy. Extracts from the bark exhibit multiple pharmacological functions, especially in lowering blood pressure. Geniposidic acid (GA), pinoresinol diglucoside (PG) and chlorogenic acid (CA) are three compounds isolated from the bark of the Eucommia tree, which display enormous bioactivities.; As a type of plant tissue culture, the hairy root culture system displays prospective application over traditional cell or callus cultures, which are characterized by rapid growth, and stable biochemical and genetic capacity. The present dissertation discusses the establishment and chemical analysis of E. ulmoides hairy roots.; After molecular identification, including PCR and Southern blotting, hairy roots were subjected to various physical and chemical treatments to investigate their impacts on growth and secondary metabolites. Results showed that light, initial pH, and culture volume did not affect the growth remarkably, while medium type, medium strength, sucrose concentration and auxin influenced the growth significantly. Biomass of E. ulmodies hairy roots increased 30% in modified culture conditions. Initial pH only had effect on PG content, while culture volume showed some effect on CA content. Other factors demonstrated varied impacts on content and yield of secondary metabolites. Zero to 50 muM methyl jasmonic acid (MeJA) elicited PG synthesis but inhibited GA production. A low concentration of salicylic acid (SA) reduced secondary metabolic synthesis, while a high concentration of it accelerated the exudation of the metabolites into medium. Exposure to light tended to improve synthesis of secondary metabolites, especially GA and CA. In contrast to other sources of E. ulmoides, hairy roots could synthesize high amounts of secondary metabolites, even specific compounds.; In order to increase secondary metabolite production, Vitreoscilla hemoglobin (VHb) was introduced into E. ulmoides hairy root. Molecular characterization, i.e. PCR, Southern blotting and Northern blotting, confirmed the integration and expression of the VHb gene. The transformed hairy root showed improved growth and enhanced synthetic capacity of secondary metabolites.; This research will contribute to exploit a new approach to produce PG, GA and CA from the special tissue culture, hairy root culture system.; Keywords. chlorogenic acid; Eucommia ulmoides ; geniposidic acid; hairy root; methyl jasmonate; pinoresinol diglucoside; salicylic acid; tissue culture; Vitreoscilla hemoglobin.