Physiological effects of temporary immersion on Hevea brasiliensis callus

In vitro culture by temporary immersion generates potentially stressful conditions for explants that may differ from those associated with classic methods. In order to evaluate the effects of these conditions on physiological changes in explants, different parameters of metabolic activity were investigated for a friable embryogenic callus of Hevea brasiliensis (Mull. Arg.), in response to 1 min, and 1, 12 and 24 h per day of immersion, using semi-solid and agitated liquid media as controls. The relative growth rate of the callus was not significantly different for the 1 min immersion treatment and the controls, but it decreased by about 60% for the 1-, 12- and 24-h immersion treatments. During the immersed stage, the rate of respiration of the callus was comparable for all the treatments. However, during the emersed stage, the respiration rate increased by 140 and 164% for the 12- and 24-h immersion treatments, respectively. Meanwhile, the total adenylate nucleotide concentration and the ratio of ATP/ADP remained almost constant, or even decreased. The adenylate energy charge was comparable for all the treatments, averaging 0.88. The superoxide dismutase activity and the lipid peroxidation increased with the immersion duration, and were significantly higher for the 12- and 24-h immersion treatments than for controls. However, after 24 h in emersed stage, there was no lipid peroxidation, regardless of previous immersion duration. It appears from these results that the immersed stage induced a substantial oxidative stress, which was not associated with the callus immersion per se.