Cryopreserved Marine Microalgae Grown Using Different Freezing Methods

Marine microalgae are a key diet component in finfish and shellfish aquaculture. Cryopreservation of the microalgae is suggested by many other studies as the best method for long-term storage. To test cryopreservation efficacy, 19 taxas of marine microalgal species were examined. In the first experiment we compared dimethylsulfoxide ($Me_2SO$) and glycerol, which are most widely used as cryoprotectant agents (CPAs). The cryopreservation comprised two freezing procedures. Firstly, the samples containing the CPAs were kept at $4^{\circ}C$ for 10 min before being plunged into liquid nitrogen ($-196^{\circ}C$). Secondly, samples containing CPAs were pre-cooled ($-1^{\circ}C$ $min^{-1}$ to $-80^{\circ}C$ before being plunged into liquid nitrogen. Most of the species were successfully cryopreserved using $Me_2SO$, whereas the Prasinophyceae (T. striata and T. suecica) were successfully cryopreserved using glycerol. In general, the cooling method had no influence on the survival of the microalgae except in the case of the Tetraselmis species. In the second experiment, the cultured solution was divided before cryopreservation into concentrated and non-concentrated groups to identify the effect of cell density during cryopreservation. After 12 months of storage, the samples were again divided into centrifugation and non-centrifugation groups to learn the effect of $Me_2SO$ on the culture. Viability and growth of the microalgae were not influenced by cell density or the centrifugal removal of the $Me_2SO$ after thawing.