Outdoor Cultivation Of Two Marine Microalgae As Live Feed And Nutrient Loss Evaluation Of The Concentrates After Cryopreservation

Marine microalgae, as primary productivity of marine ecosystem, are large groupof species with plenty of nutrition and have been the main living feed in aquaculture.With the increasing development of aquaculture, the demand of living microalgae alsohas being increased. Therefore, it is particularly important to strengthen the researchon cultivation and preservation technology for marine microalgae. In the present study,the technology for cultivation of two marine microalgae, Chaetoceros muelleri andIsochrysis zhangjiangensis, and cryopreservation was investigated. The mainexperimental results were as follows.1． Microalgae cultivation system with CO2supplement coupled with pHfeedback control was designed and constructed. It could achieve the double effects ofon-line CO2addition and pH control in medium. During the cultivation ofChaetoceros muelleri, compared to the control culture, this system showed anincreased growth rate of cells at1.85times and enhanced ability of resistance to theoutdoor environment. While, total fatty acid content increased14.5%, in which theunsaturated fatty acids and EPA increased17.1%and22.1%, respectively. Thecontent of C18:3was increased significantly at57.4%.2．In the cultivation system mentioned above, the effects of three nitrogensources (sodium nitrate, urea and ammonium bicarbonate) on the outdoor growth andfatty acid content of Chaetoceros muelleri were investigated. The result showed thatby using ammonium bicarbonate, the highest specific growth rate and maximal dryweight concentration were achieved at0.23d-1and0.39g/L respectively. By usingsodium nitrate, more total fatty acids could be accumulated and the content could beup to96.95mg/g. The highest contents of PUFA and EPA were obtained at27.18mg/g (31.15%TFA) and11.76mg/g (13.48%TFA) using ammonium bicarbonate. Theresult suggests that ammonium bicarbonate is the best nitrogen source for the growthof Chaetoceros muelleri and accumulation of PUFA.3．Effects of nitrogen sources on the growth, pigment and fatty acid content ofIsochrysis zhangjiangensis were investigated comparatively. The result showed that by using ammonium bicarbonate, the maximal specific growth rate and biomassconcentration reached at0.24d-1and0.262g L-1respectively. The highest chlorophylla and fucoxanthin content were obtained at16.58mg/g and13.9mg/g in the secondday using sodium nitrate, respectively.The highest content of total fatty acid was upto137.22mg/g by using urea, PUFA content were34.54mg/g (31.24%TFA),53.68mg/g (39.12%TFA) and44.34mg/g (32.63%TFA), and DHA content were17.55(15.87%TFA),29.58(21.56%TFA) and24.97(18.38%TFA) by using sodium nitrate,urea and ammonium bicarbonate. The result suggested that ammonium is the bestnitrogen source for DHA accumulation.4．By using ammonium bicarbonate, the effects of three salinities (20‰,25‰,30‰) on the growth, pigment content and fatty acid content of Isochrysiszhangjiangensis were investigated. The result showed that the highest cell density, dryweight concentrate, pigment and DHA contents were achieved at20‰salinity. Thecell density and biomass concentrate were5.5×106/ml and0.227g L-1respectively.While, chlorophyll a and fucoxanthin content were up to the maximum at19.30mg/g和14.52mg/g, DHA content at24.63mg/g (17.29%TFA). The highest specificgrowth rate and biomass concentration were0.25d-1and0.39gL-1, respectively. At20‰salinity, the content of total fatty acid and PUFA were up to124.31mg/g and47.96mg/g. With the rise of salinity, the total fatty acid contents in algal cells alsogradually increased but the levels of PUFA reduced, showing a negative correlationwith salinity.5. The nutrient loss rate of Chaetoceros muelleri concentrate aftercryopreservation under-18℃and4℃was studied. The result showed with cellconcentration at1010/ml (A) and109/ml (B), the control group without anycryprotectant had the minimum loss rate of fatty acid content when cryopreservated at-18℃. The loss rate of EPA, PUFA and TFA (fast freezing group referred as Aconcentrate) were1.73%,2.13%and2.32%. Fatty acid loss rate of A concentrate wassignificantly lower than that in B concentrate, slow freezing group (cooling to-18℃at the rate of1℃/min) was lower than that of fast freezing (freezing at-18℃directly) group. During cryopreservation at4℃, loss rate of fatty acid was similar to that-18℃with a slightly higher value.6. The protein content in living Chaetoceros muelleri accounted for34.7%of thecell dry weight. After cryopreservation of concentrates for2months at-18℃, the lossrate of fatty acid content were7.39%,8.39%,18.79%and16.11%in the four groups,Af group (fast freezing for A concentrate), As group (slow freezing for Aconcentrate), Bf group (fast freezing for B concentrate), and Bs group (slow freezingfor B concentrate), respectively. The loss rate of fatty acids in A concentrate wassignificantly lower than that in B concentrate. After cryopreservation of concentratesfor2months at4℃, the loss rate of protein was higher than that at-18℃. The lossrate of A concentrate was18.12%and that of B concentrate was24.71%, the formermarkedly lower than the latter.