The effects of slow and rapid warming on the viability of marine microalgae cryopreserved at different cooling rates were studied. Of five species, Rhodomonas baltica was the only one that did not survive slow warming following cryopreservation at cooling rates of 6 and 16#&176;C min-1. This alga tolerated slow warming when the cooling rate employed to freeze the alga was 0.5#&176;C min-1. However, significant lower viabilities (17.2% ± 15.4 in a salinity of 20 parts per thousand (ppt or g/liter) and 15% Me2SO and 7.9% ± 6.5 in a salinity of 36 ppt and 15% Me2SO) were observed in comparison to cells rapidly thawed (28.8% ± 16.7 in 20 ppt salinity-15% Me2SO and 17% ± 11.5 in 36 ppt salinity-15% Me2SO). Chaetoceros gracilis cryopreserved with 15% Me2SO tolerated slow and rapid warming achieving similar viabilities when the cooling rate was 0.5#&176;C min-1 (average 26.8% ± 5.6 in a salinity of 20 ppt and 31.0% ± 7.2 in a salinity of 36 ppt). At a higher cooling rate (6#&176;C min-1), viability decreased with slow warming and was zero at 16#&176;C min-1. Although viabilities were below 6% for Tetraselmis chuii frozen in salinities either of 20 or 36 ppt (both with 15% Me2SO), it could tolerate slow warming when the cooling rate was 16#&176;C min-1. Under all cooling rates tested, rapid thawing under the same conditions of salinity and cryoprotectant concentration enabled this alga to recover with viabilities ranging from 79.2% ± 20 to 99.4 ± 22.2. The cryopreservation of Nannochloropsis gaditana using a salinity of 20 ppt and a cooling rate of 0.5#&176;C min-1 was the only case where slow warming produced higher viabilities (36.7% ± 5.4 for absence of cryoprotectant and 38.9% ± 2.4 for 15% Me2SO) than those achieved after rapid thawing (17.7% ± 4.8 for absence of cryoprotectant and 25.1% ± 6.2 for 15% Me2SO). This alga, together with Nannochloris atomus, showed higher viability levels when slowly thawed after cryopreservation using a cooling rate of 16#&176;C min-1. However, significantly higher levels of viability were always observed when the algae were thawed rapidly.
Document Type: Research Article
Centro de Investigacion y Cultivo de Especies Marinas "El Toruno,", Junta de Andalucia, El Puerto de Santa Maria, Cadiz, 11500, Spain 2:
Instituto de Ciencias Marinas de Andalucia, CSIC, Puerto Real, Cadiz, 11510, Spain