摘要
在东北地区水域采集含有浮游微藻的水样,用纱布过滤去掉水体中较大的沉淀物,取100 mL滤液转入500 mL的三角摇瓶中,加入100 mL BG11培养基,在25~28℃、光照强度为3000 Lux,光暗比为14∶10下富集培养,采用稀释涂布法分离纯化至明显的绿色。对筛出微藻在200 mL柱式光照反应器中进行光照通气培养与评价,其中生长最快的微藻生物量达到1.8305 g·L^(-1),最高的油脂含量达到34.69%,最高油脂产率达29 mg·L^(-1)·d^(-1)。取5株(一株小球藻属Chlorella、两株单针藻属Monoraphidium两株栅藻属Scenedesmus)生长快速、含油量高的藻种新鲜藻液分别离心(4000 r·min^(-1),3 min),去离子水洗两次后,加入新鲜培养基重悬,取出部分微藻加入5种保护剂:甘油(GLY)、二甲基亚砜(DMSO)、蔗糖(SUC)、聚乙二醇(PEG)和羟二基淀粉(HES),分别置于-18℃和-80℃保藏。一个月后,取出保藏微藻,解冻,摇匀,离心,用生理盐水洗两次,去除保护剂后一部分检测细胞活性评价,一部分接种到新鲜培养基进行生长评价。选取确定保护剂后进一步考察保护剂浓度(10%、20%、30%、40%和50%)和保藏时间(3个月、6个月和9个月以及1年)。结果表明:采用二醋酸荧光素法能快速反应细胞冷冻后细胞活力,采用-18℃添加20%甘油或-80℃添加10%甘油冷冻保藏,适用于大多数含油微藻。该方法可保藏含油微藻至少1年且细胞生物量和油脂含量稳定。
Water samples containing planktonic microalgae were collected in the waters in the Northeast China,filtered with gauze to remove the larger sediments,and then 100 mL of the filtrate was transfered into a 500 mL triangle shaker flask,and added 100 mL of BG11 medium at 25℃-Enriched culture at 28℃,light intensity of 3000 Lux,light to dark ratio of 14∶10 by dilution coating method for separation and purification.The screened microalgae were cultured and evaluated by light aeration in a 200 mL column-type light reactor,with the maximal microalgae biomass of 1.8305 g·L^(-1),oil content of 34.69%,and oil yield of 29 mg·L^(-1)·d^(-1).Five strains including 2 species in Scenedesmus,3 species in Monoraphidium,and 1 species in Chlorella,of fast-growing alga species with high oil content were centrifuged(4000 r·min^(-1),3 min),deionized after washing twice with water,added fresh medium to resuspend,removed part of the microalgae and stored in 5 kinds of protective agents:glycerol(GLY),dimethyl sulfoxide(DMSO),sucrose(SUC),polyethylene glycol(PEG)and hydroxyl dibasic starch(HES),at-18℃and-80℃,respectively.The results showed that PEG and DMSO as penetrating cryprotectants were the more effective than other 3 cryoprotectants.When the concentration of the 2 cryprotectants exceeded 20%,the cell activity of most microalgae was not affected.Adding 20%of GLY at-18℃or adding 10%of GLY at-80℃were adopted for frozen preservation of most of oil microalgae at least 1 year and kept microalgae physiological characteristics stable,such as biomass and oil content.
作者
李澜鹏
廖莎
王鹏翔
孙启梅
师文静
李晓姝
张霖
彭绍忠
LI Lanpeng;LIAO Sha;WANG Pengxiang;SUN Qimei;SHI Wenjing;LI Xiaoshu;ZHANG Lin;PENG Shaozhong(Dalian Research Institute of Petroleum and Petrochemicals,SINOPEC,Dalian 116045,China)
出处
《水产学杂志》
CAS
北大核心
2021年第3期34-41,共8页
Chinese Journal of Fisheries
基金
中国石化科技开发项目(214054).
关键词
含油微藻
细胞相对活性
保护剂
冷冻保藏
oil-rich microalga
relative cell activity
cryoprotectant
cryopreservation
