Changes in cell water relations during precultures were followed in an attempt to understand the mechanism of cell hardening for cryopreservation by vitrification. Medium containing 0.4 M sucrose (psiw=-1.45 MPa) and containing 5 mg L-1 of ABA (MT psiw=-0.73 MPa and MS psiw=-0.48 MPa) were used to harden cell suspensions of orange and carrot. Preculture in these medium did not cause a significant decrease of cell viability, however, it improved the cell survival to PVS2 and liquid nitrogen expositions. When cells were inoculated into medium containing 0.4 M sucrose, the cell psiw decreased rapidly until reach the medium psiw , and turgor was also severely reduced or disappeared. Subsequently, cell psis began to decrease and after 48 h of treatment in low water potential medium, turgor was recovered by osmoregulation to almost the same values that cells had at the beginning of the experiment. Preculture in medium containing ABA also caused a decrease of cell psis but in much less extent than the preculture in low water potential medium. Since both precultures improved significantly the survival to PVS2 pretreatment and to LN exposure, this increment of survival to osmotic shock and vitrification can not be attribute solely to quantitative solute accumulation. The results suggest that qualitative accumulation of determined solutes would be the events which contributed for the improvement of tolerance to PVS2 exposure and thence tolerance to liquid nitrogen.
Preculture; cell suspension; sucrose; ABA; cryopreservation
