Gas-chromatography-mass-spectrometry revealed the presence of various bioactive compounds with anticancer properties in protocorm-like-body (PLB) cultures of a Dendrobium hybrid orchid (Dendrobium Enopi x Dendrobium Pink Lady). Pre-illumination of red fluorescent light lessened the stimulating effects of light-emitting diodes (LEDs) on secondary metabolites production among in vitro PLB cultures, possibly due to habituation. The highest flavonoid content of 16.79 μmol/ g of fresh weight (FW) was achieved under blue-red (1:1) LED for PLBs pre-treated with white LED for more than 3 subculture cycles. Phenolics content significantly reduced as PLBs pre-cultured under red fluorescent light for 2 subculture cycles were exposed to LED illuminations, where far red LED resulted in the lowest total phenolic content (18.85 μmol/ g FW). High intensity green LED (16.9 μmol/s) enhanced the accumulation of phenolics while amino acids such as L-leucine, glycine and proline exhibited no significant stimulating effect for secondary metabolites production.
A droplet-vitrification cryopreservation protocol has been successfully developed for Aranda Broga Blue orchid hybrid using protocorm-like bodies (PLBs). However, maximum growth regeneration percentage was recorded at 5% only based on previous report. Thus, to improve growth recovery of cryopreserved PLBs, cryopreservation stages were supplemented with ascorbic acid, tested at 50, 100 and 150 mg/L. However, results demonstrated that exogenous ascorbic acid was not favorable in regeneration of cryopreserved explants (maximum value of 1.67 % with 50 mg/L ascorbic acid supplementation). Total soluble protein and various antioxidant enzyme activities such as catalase (CAT), superoxide dismutase (SOD) and ascorbate peroxidase (APX) were evaluated after each cryopreservation stages in conjunction with the application of exogenous ascorbic acid. Addition of antioxidant must be carefully evaluated and its application may not guarantee successful growth recovery. RAPD and SCoT molecular analysis confirmed the genetic stability of regenerated cryopreserved PLBs as no polymorphism was detected compared to control PLBs culture.