Determination of glassy state by cryo-SEM and DSC in cryopreservation of mint shoot tips by encapsulation–dehydration

Abstract

Cryopreservation of mint shoot tips grown in vitro (Mentha 9 piperita) was performed after encap- sulation in alginate beads. Encapsulated shoot tips were first precultured in sucrose enriched medium (0.75 M) and then dried under a sterile air flow (0-6 h). After cooling in liquid nitrogen and warming in a warm water bath, alginate beads were transferred to solid culture medium for 4 weeks. The effect of dehydration time of the encapsu- lated shoots was evaluated for water content, cooling and warming rates, ice crystal formation and cellular vitrifica- tion, by using low temperature scanning electron micros- copy and differential scanning calorimetry. Viability of the recovered material showed a close relation between the dehydration time, cooling and warming rates, ice formation avoidance and tissue vitrification. At short drying periods (up to 3 h), ice crystals were formed and the viability was low or absent. After longer drying periods (5 and 6 h), both beads and specimens became vitrified.