以球等鞭金藻(Isochrysis galbana)3011为对象,通过研究降低培养温度后其超氧化物歧化酶(superoxide dismutase,SOD)、过氧化物酶(peroxidase,POD)和过氧化氢酶(catalase,CAT)活性以及还原型谷胱甘肽(glutathione,GSH)、脂质过氧化产物丙二醛(malondialdehyde,MDA)、活性氧(reactive oxygen species,ROS)和二十二碳六烯酸(docosahexaenoic acid,DHA)含量的变化,以阐明低温环境对球等鞭金藻细胞抗氧化系统和DHA含量的影响。用流式细胞术结合荧光染色法测定低温环境对球等鞭金藻细胞内ROS水平的影响;并采用气相色谱法检测球等鞭金藻细胞内DHA含量。结果表明:在21、18、15℃低温环境处理下,球等鞭金藻细胞SOD、CAT和POD活性均随培养时间延长而呈现先升高后降低的趋势;温度越低,峰值出现越早,且峰值越大,峰后酶活性下降越快;GSH含量的变化趋势与上述酶活性的变化相似,MDA含量则持续增加;ROS水平随着温度的降低而呈现出较为复杂的变化,15℃和18℃诱导16 h出现ROS水平爆发,20 h时达到峰值,分别为(14.11±0.11)%和(14.74±0.58)%(P〈0.05);经18℃低温诱导24 h后,球等鞭金藻细胞内DHA含量为0.105 mg/g,比对照组高0.06 mg/g。因此,低温环境可以作为提高代谢物产量的诱导子,使球等鞭金藻细胞产生主动防御反应,引起清除活性氧相关的酶活性的升高,同时也提高了DHA产量。
Effects of low environmental temperature on the activities of superoxide dismutase(SOD), peroxidase(POD) and catalase(CAT), and the contents of glutathione peroxidase(GSH), malondialdehyde(MDA), docosahexaenoic acid(DHA) and reactive oxygen species(ROS) in Isochrysis galbana 3011 were investigated for exploring the relationship among antioxidant enzyme activities, ROS levels and DHA contents at different temperatures. The enzyme activities were determined by corresponding kits. ROS levels were detected by flow cytometry and inverted fluorescence microscope, and DHA contents were determined by gas chromatography(GC). The results showed that activities of SOD, CAT and POD increased firstly and then decreased with increasing culture time at 15, 18 and 21 ℃. Lower temperature could result in earlier and higher enzyme activity peaks followed by a faster decline. The profile of GSH was similar to that of antioxidant enzyme activities, while content of MDA increased continuously. ROS levels showed a complex change with declining temperature, and ROS burst was appeared after induction for 16 h at 15 and 18 ℃, with maximum values of(14.11 ± 0.11)%and(14.74 ± 0.58)% at 20 h, respectively(P 0.05). DHA was 0.105 mg/g at 18 ℃ for 24 h, which was higher than that at 24 ℃ for 24 h by 0.06 mg/g. Therefore, low environmental temperature can improve the levels of metabolic products by activating microalgae defense responses, increasing ROS-related enzyme activities, ROS levels and DHA contents.