The effects of UV-B radiation on the nutritional composition of Antarctic phytoplankton

The actual piece of equipment used was an International Light IL 1700Radiometer equipped with broad band detectors to measure PAR, UV-A and erythemal UV-B. The effects of UV-B radiation on the fatty acid, total lipid and sterol composition and content of three Antarctic marine phytoplankton were examined in a preliminary culture experiment. Exponential growth phase cultures of the diatoms Odontella weissflogii and Chaetoceros simplex and the Haptophyte Phaeocystis antarctica were grown at 2 (plus or minus 1)degrees C and exposed to 16.3 (plus or minus 0.7) W.m-2 photosynthetically active radiation (PAR). UV-irradiated treatments were exposed to constant UV-A (4.39 (plus or minus 0.20) W.m-2) and low (0.37 W.m-2) or high UV-B (1.59 W.m-2). UV-B treatments induced species specific changes in lipid content and composition. The sterol, fatty acid and total lipid content and profiles for O. weissflogii changed little under low UV-B when compared with control conditions (PAR alone), but showed a decrease in the lipid content per cell under high UV-B treatment. In contrast, when P. antarctica was exposed to low UV-B irradiance, storage lipids were reduced and structural lipids increased indicating that low UV-B enhanced cell growth and metabolism. P. antarctica also contained a higher proportion of polyunsaturated fatty acids under low UV-B in comparison with PAR irradiated control cultures. The flagellate life stage of P. antarctica died under high UV-B irradiation. However, exposure of P. antarctica to high UV-B irradiance increased total lipid, triglyceride and free fat.The effect of UV-B irradiances on the lipid content of Antarctic marine phytoplankton is species specific. Changes in ambient UV-B may alter the nutritional quality of food available to higher trophic levels.EXPERIMENTALAll measurements of irradiance were made with an International Light IL 1700 Radiometer equipped with broad band detectors to measure PAR, UV-A and erythemal UV-B [14]. A National Institute of Standards and Technology intercomparison package (NIST Test #534/240436-88) was used to calibrate each light sensor.Unialgal cultures of the diatoms Odontella weissflogii and Chaetoceros simplex were isolated from sea ice collected in Prydz Bay, Antarctica during the 1990/91 austral summer. Phaeocystis antarctica was isolated from Prydz Bay in 1982/83 summer. Cultures of diatoms and Phaeocystis antarctica were maintained in 2 l glass flasks using f/2 growth medium [32] and GP5 medium [33] respectively at a temperature of 2 plus or minus 1 degrees C. Cool white fluorescent lights provided photosynthetically active radiation (PAR) intensity of 17.08 J.m-2.s-1 (84.7 micro E.m-2.s-1), with no UV-B enhancement, on a 12 h light : 12 h dark cycle. Immediately before experimental irradiation, three replicate subsamples of approximately 15 ml were obtained from each parental culture and fixed with Lugols iodine, a known sample volume sedimented, and cells counted over 15 replicate fields using a Labovert inverted microscope. Mean cell concentration and standard deviation were then computed. Each exponential growth phase parental culture was thoroughly mixed and 3 replicate 300 ml Costar polystyrene culture flasks (which completely absorbed wavelengths below 295 nm) established for each light treatment (control, low and high UV exposures). Cultures were irradiated for 24 hours in a 48 hour experimental period (6 h light : 12 h dark : 12 h light : 12 h dark : 6 h light) [14, 23]. Exposures were conducted in a Thermoline controlled environment cabinet at 2 plus or minus 1 degrees C with cool white fluorescent tubes to provide PAR and UV-A (320-400 nm), with UV-B provided by FS20T 12 UV-B Westinghouse sunlamps. PAR and UV-A irradiances were 16.3 plus or minus 0.7 W.m-2 (81.3 plus or minus 3.4 micro E.m-2.s-1) and 4.39 plus or minus 0.20 W.m-2 respectively. The spectral distribution and UV-B irradiance were varied by attenuation with glass filters [5] to provide low (0.37 W.m-2) or high UV-B (1.59 W.m-2). Sensors were each covered by an attenuating glass screen and a single layer of Costar culture flask to measure the experimental irradiances to which the algae were exposed. UV-B irradiances were chosen to reflect less than (74%) and greater than (318%) peak UV-B exposure as measured at an Antarctic coastal site (Casey station, 66 degrees S, [34]).Following irradiation each culture was well mixed and approximately 15 ml was fixed with Lugols Iodine for subsequent estimation of cell concentration (as above). Chlorotic and greatly vesicularised cells were considered to be dead [23]. The remainder of each experimental culture was filtered through Whatman GF/F filters. On completion of filtration, the filters were stored at -20C overnight before extraction of lipids the following day.本研究使用的实际设备为International Light IL 1700辐射计（Radiometer），配备宽带探测器以测量光合有效辐射（Photosynthetically Active Radiation, PAR）、UV-A及红斑量UV-B。本预培养实验探究了UV-B辐射对3株南极海洋浮游植物的脂肪酸、总脂及甾醇组成与含量的影响。实验所用的指数生长期培养物包括硅藻威氏角毛藻（Odontella weissflogii）、简单角毛藻（Chaetoceros simplex）以及定鞭藻类（Haptophyte）南极棕囊藻（Phaeocystis antarctica），培养温度为2(±1)℃，光合有效辐射强度为16.3(±0.7) W·m⁻²。UV辐照组持续暴露于UV-A（4.39±0.20 W·m⁻²）以及低强度（0.37 W·m⁻²）或高强度（1.59 W·m⁻²）UV-B。UV-B处理可引发物种特异性的脂质含量与组成变化。与仅施加PAR的对照组相比，威氏角毛藻（O. weissflogii）在低UV-B辐照下的甾醇、脂肪酸、总脂含量及组分变化极小，但在高UV-B处理下其单位细胞脂质含量出现下降。与之相反，南极棕囊藻（P. antarctica）在低UV-B辐照下储存脂质减少、结构脂质增加，表明低UV-B可促进细胞生长与代谢；相较于仅受PAR辐照的对照培养物，该藻在低UV-B条件下还含有更高比例的多不饱和脂肪酸。南极棕囊藻的鞭毛生活阶段在高UV-B辐照下发生死亡，但在高UV-B辐照处理后，其总脂、甘油三酯及游离脂肪含量均有所提升。UV-B辐照度对南极海洋浮游植物脂质含量的影响具有物种特异性。环境UV-B的变化可能会改变高营养级生物可获取食物的营养品质。 ### 实验方法 所有辐照度测量均使用配备宽带探测器（可测量PAR、UV-A及红斑量UV-B）的International Light IL 1700辐射计完成[14]。使用美国国家标准与技术研究院（National Institute of Standards and Technology, NIST）比对包（NIST测试编号#534/240436-88）对每个光传感器进行校准。硅藻威氏角毛藻与简单角毛藻的单种培养物分离自1990/91年南极夏季普里兹湾的海冰样本，南极棕囊藻分离自1982/83年夏季的普里兹湾海域。硅藻与南极棕囊藻的培养物分别置于2L玻璃烧瓶中，使用f/2培养基[32]与GP5培养基[33]进行维持，培养温度为2±1℃。采用冷白色荧光灯提供光合有效辐射，光照强度为17.08 J·m⁻²·s⁻¹（84.7 μE·m⁻²·s⁻¹），无UV-B增强，光照周期设置为12h光照:12h黑暗。实验辐照前，从每个亲本培养物中获取3份约15ml的重复子样本，使用卢戈氏碘液（Lugols iodine）固定，将已知体积的样本沉降后，使用Labovert倒置显微镜对15个重复视野内的细胞进行计数，随后计算平均细胞浓度与标准差。将每份指数生长期的亲本培养物充分混匀，为每个光照处理（对照组、低UV-B组与高UV-B组）各设置3个300ml的Costar聚苯乙烯培养瓶（可完全吸收295nm以下波长的光线）。实验周期为48小时，其中辐照时长为24小时，光照周期设置为6h光照:12h黑暗:12h光照:12h黑暗:6h光照[14,23]。辐照实验在Thermoline可控环境箱中开展，培养温度维持在2±1℃，冷白色荧光灯管提供PAR与UV-A（320-400nm），UV-B由FS20T 12型Westinghouse紫外线太阳灯提供。PAR与UV-A的辐照度分别为16.3±0.7 W·m⁻²（81.3±3.4 μE·m⁻²·s⁻¹）与4.39±0.20 W·m⁻²。通过玻璃滤片衰减来调整光谱分布与UV-B辐照度，以获得低（0.37 W·m⁻²）或高（1.59 W·m⁻²）强度的UV-B。每个传感器均覆盖衰减玻璃屏与单层Costar培养瓶，以测量藻类实际受到的实验辐照度。所选UV-B辐照度分别对应南极沿海站点（凯西站，南纬66°，[34]）测得的峰值UV-B暴露量的74%以下与318%以上。辐照结束后，将每个培养物充分混匀，取约15ml样本用卢戈氏碘液固定，用于后续细胞浓度估算（方法同上）。萎黄且大量囊泡化的细胞被视为死亡细胞[23]。将每个实验培养物的剩余样本通过Whatman GF/F滤膜过滤，过滤完成后，将滤膜置于-20℃过夜储存，次日进行脂质提取。