SEMICONTINUOUS CULTURE SYSTEM FOR LEMNA GIBBA BIOASSAY: FUNCTIONING AND THEORY OF OPERATION

Abstract

To overcome the problems associated with high concentrations of nutrient and test chemicals in laboratory Lemna gibba bioassays, a semi-continuous culture system was adopted, and specific equipment was designed and constructed to simulate steady-state conditions of the most natural aquatic systems. The equipment for Lemna semicontinuous culture consists of a growth chamber and a circulation control unit. Lemna gibba is kept in the growth chamber in a growth pot (with 1 mm diameter perforation at about 10 cm above the bottom), which retains the plant but allows an efficient exchange of the growth medium. Flow rate and composition of the medium is therefore varied independently of the L. gibba population density. The media are kept in circulation, and recharges or discharge of the media are controlled. The system took into consideration chemical processes such as O2 and CO2 exchange and maintained a degree of turbulence through continuous shaking of the growth chambers. It is also possible to investigate growth under different supply of O2 and CO2, and pH control with dissolved CO2 or reducing conditions with N2 gases. Theoretical analysis of growth in the steady state shows that integrated activation time of the dilution pump is proportional to the growth rate of L. gibba. Theoretical analysis was also used to determine the minimum flow-rate and nutrient concentration of the medium to cover the requirements of L. gibba. Experiments were carried out that demonstrated that the steady state growth could be attained and be controlled by the nutrient concentration, flow rate, and recharge intervals. The cultures could be kept at steady state over 21 days of the Lemna test period. Keywords. Lemna gibba, steady state, semicontinuous culture, batch culture, nutrient media