Analysis of cell suspension cultures raised in shake flasks
i. Kinetics of cell growth and nutrient uptake
The specific growth rate (μ) of the suspended cells was found to be 0.1072 d-1. It was observed that the cultures remained in the lag phase till the 2nd day. Biomass increased till the 10th day following which the stationary phase started (Figure 11.5) . The pH of the medium underwent variation during different stages of culture. It was observed that after showing a slight decrease in the value, the pH dropped sharply between 4-6 days, which dropped further after 10 days resulting in poor growth and stationary phase of cells. The drop of pH may be attributed to preferential uptake of NH4+ ions which in turn resulted in decreased pH due to liberation of H+ ions; pH tends to increase if NO3- is utilized faster than NH4+. Uptake of nitrate was observed at slower rate. It was present in the culture medium till the last days of cultivation (16th day). The concomitant synthesis of acid triterpenes in the medium was found to be growth associated and showed an increase with the increase in biomass. Conductivity was shown to have an inverse relationship with growth. Among the major inorganic nutrients, it was invariably observed that phosphate was almost completely consumed by the 10th day of culture. Its utilization was very fast in the initial days than in the later stages of growth. Hence, it may be concluded that complete utilization of phosphate and a marked drop in the pH of the culture medium resulted in the onset of stationary phase in cell suspension cultures.
Figure 11.5: Kinetics of cell growth and nutrient uptake in cell suspension cultures of L. camara .