Production of rosmarinic acid and lithospermic acid B in Ti transformed Salvia miltiorrhiza cell suspension cultures

Abstract

A Ti transformed cell culture of S. miltiorrhiza demonstrated the ability to produce rosmarinic acid (RA) and lithospermic acid B (LAB) as determined by HPLC. A time course of biomass accumulation, phenolic compound formation, medium pH and conductivity as well as sugar, nitrate, ammonium and phosphate uptake was examined both in flask and in bioreactor cultures using hormone free B5 medium containing 2% sucrose. In flasks, the stationary phase of growth was reached after about 16 days of cultivation; the maximum biomass concentration was about 10.5 g l-1 dry weight. The maximum production of RA (4.59% of dry cell weight or 481.5 mg l-1) appeared on the 20th day of cultivation, at which time the production of LAB was 0.81% of dry weight or 85.0 mg l-1. A linear relationship with an intercept between the dry cell weight increase and the medium conductivity decrease was observed which may be used for monitoring cell growth rate. Initial medium pH over a wide range (3.4-8.5) had little effect on cell growth but had significant influences on rosmarinic acid formation, although the trend was irregular. In bioreactor cultivation, the maximum dry Cell biomass accumulation obtained was 7.52 g l-1 on the 15th day of cultivation. The lower performance of cell growth in the bioreactor was due to the poor mixing of the culture.