Influence of the conditions of cultivation on the growth of callus and cell cultures ofAjuga turkestanica and the biosynthesis of ecdysterone

Abstract

The influence of light on the total productivity of a tissue culture of Ajuga turkestanica has been studied. When calluses are grown in the absence of light the yield of biomass and its ecdysterone content increase. In a study of the influence of c~-naphthylacetic acid on the productivity of a suspension cell culture it has been shown that with an increase in the concentration of the auxin the yield of crude mass rises but the ecdysterone content falls. The activation of the biosynthesis of the secondary product takes place with a retardation of the processes of growth and cell division.Under the conditions of a decrease in the natural reserves of useful plants, work on the production of biologically active compounds with the aid of tissue and plant cell cultures is becoming urgent.We have previously obtained callus tissue of Ajuga turkestanica retaining the biosynthetic activity of ecdysterone under in vitro conditions [1]. With the aim of studying the possibility of a physiological regulation of the growth processes of tissue and cell cultures and of the synthesis of ecdysterone in them, we have investigated the influence of the illumination regime on the growth of A. turkestanica callus tissue and the yield of product and have considered the dependence of the growth of a suspension cell culture and of the accumulation of ecdysterone on the amount of an auxin in a liquid nutrient medium and also the nature of its accumulation in the dynamics of the growth of the cell culture.We investigated callus tissue being passaged for five years. Samples of tissue grown under conditions of illumination (6000 lux with a 16-hour photoperiod) and in the dark were taken on the 30th day of cultivation. In the tissue culture grown in the light it was impossible to detect the presence of ecdysterone by the HPLC method because it was present in only trace amounts. The yield of crude mass from one flask was 11.46 g, the dry matter amounting to 0.42 g. Growing the callus tissue in the dark had a favorable effect not only on the yield of biomass but also on the biosynthesis of ecdysterone. Thus, the yields of crude mass and dry matter were 13.62 and 0.54 g, respectively, and the ecdysterone content was 0.069%.The influence of an auxin on productivity was studied on a suspension cell culture obtained by transferring A. turkestanica callus tissue into a liquid nutrient medium with the following composition: 1/2 part of Murashige-Skoog mineral salts [2]; sucrose, 30 g/liter; inositol, 100 mg/liter; thiamine-HCl, 0.4 mg/liter. By the repeated selection of a finely aggregated fraction we obtained a viable, actively growing suspension. Media with different amounts of c~-naphthylacetic acid (NAA) were tested: 1) with no added auxin; 2) with 1 mg/liter of NAb,; 3) with 2.5 mg/liter of added NAA. Ecdysterone levels were determined in 14-day dry samples.The results of the experiment showed that ta-naphthylacetic acid affects the growth of the culture and the yield of product differently (Table 1).Thus, in the absence of an auxin the growth of ...