Avaliação de aspectos bioquímicos da crioinjúria e da capacidade biossintética de Passiflora suberosa L. após a criopreservação

Abstract

Passiflora suberosa L., commonly known as maracujazinho-cortiça is a wild species of passion fruit with agronomic, ornamental and medicinal potential. Thus, strategies aiming at the conservation of this species, such as cryopreservation, are considered of great relevance. Since the cryopreservation process can be associated with oxidative damage that causes injuries and results in low survival rates, the objective of this study was to evaluate biochemical aspects associated with cryoinjury, aiming at identifying the critical steps of the cryopreservation protocol for P. suberosa shoot tips using the V-Cryoplate technique. For the evaluation of oxidative injuries, membrane lipid peroxidation and the activity of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX) were quantified at different stages of the cryopreservation protocol. In addition,different concentrations of ascorbic acid and glutathione were added at the critical steps of the cryogenic process in order to reduce the oxidative stress. The metabolic stability of plants derived from cryopreserved shoot tips was analyzed by Thin Layer Chromatography, High Performance Liquid hromatography and by the evaluation of the antioxidant potential. The malondialdehyde content (subproduct of lipid peroxidation) in the explants was significantly higher after treatment with the cryoprotectant solution PVS3 and immersion into nitrogen (LN), with values up to 2.5 times higher than those observed in the controls. However, these rates were reduced during the regrowth stage. All antioxidant enzymes analyzed (SOD, CAT, APX) showed an expressive increase in their activities during the early stages of the cryopreservation protocol, although a significant reduction of the specific activities of SOD and CAT was observed after osmoprotection. Based on these results, osmoprotection and dehydration were considered the most critical stages of the cryopreservation protocol for P. suberosa shoot tips using V-Cryo-plate. However, the application of ascorbic acid and glutathione in these stages, aiming at reducing the oxidative stress, did not lead to an increase in plant recovery rates. In the investigation of the metabolic stability after cryopreservation, the chromatographic analyzes did not show differences between leaves of plants submitted or not to cryopreservation. Likewise, a high antioxidant potential was observed in all extracts, both by the direct scavenge of free radicals and by the ability to chelate metallic ions. The antioxidant activity of the different materials was also associated with the presence of flavonoids, phenolic acids and saponins in the extracts. These results contribute to a better understanding of the cellular responses of tropical plants to the stressful conditions induced by cold and dehydration. Furthermore, maintenance of biosynthetic capacity in plants derived from cryopreserved shoot tips provides relevant information for the study of the functioning of secondary metabolism during and after cryopreservation. These data confirm the effectiveness of cryopreservation for long-term conservation of P. suberosa.