2，4-D對高氏柴胡葉片癒合組織誘導、增殖與柴胡皂苷含量之影響

Abstract

Effect of 2, 4-dichlorophenoxyacetic acid (2, 4-D) on leaf callus induction, proliferation and formation of saikosaponin along with callus culturing of Bupleurum kaoi, a native medicinal herb of Taiwan, have been investigated. A seventy-five percent primary callus induction rate was obtained from the explants cultured on half-strength Murashige and Skoog (MS) medium containing 4 mg L^(-1) 2, 4-D for 8 weeks in darkness. Among the varied 2, 4-D concentrations (0 to 0.8 mg L^(-1)) tested for callus proliferation, the highest callus weight was observed on half-strength MS medium containing lower concentration (0.1 to 0.2 mg L^(-1)) of 2, 4-D and increasing of 2, 4-D concentration did not further improve callus proliferation. In addition, time course of callus growth and saikosaponins content were also established. There was linearly increasing in callus fresh weight from week 1 to 10 before a slowly growth occurred from week 10 to 12. The highest biomass of callus was obtained at week 10, where callus browning started. Therefore, the period for a proper growth of callus on a proliferation medium should be no longer than 10-week-incubation. Saikosaponin a, c and d (SSa, SSc, SSd) contents of callus were analyzed at two-week intervals in a total of 12 weeks span using a high performance liquid chromatography (HPLC). The total amount of SSa, SSc, and SSd (SSa+SSc+SSd), 1.12mg g^(-1) dry wt, was founded reaching the peak at interval from week 8 to week 10 and diminished hereafter. The contents of SSa, SSc, and SSd in various sources of plant materials including the aerial parts and roots of eighteen-month-old field-grown plants of B. kaoi and market crude drug (Radix Bupleuri-roots of B. Chinense), were also determined by HPLC. Although the total amount of SSa, SSc, and SSd of the cultured callus was lower than other sources of tested samples, it is thought that by modifying several important factors for callus culture of B. kaoi could make this system more efficient. Cell suspension culture in a long run has a great potential and advantage to produce secondary metabolites stably year round comparing with the field-growth plants for pharmaceutical utilization.