作 者 ：冀晓青,韩笑天,杨佰娟,俞志明
期 刊 ：生态学报 2012年 32卷 6期 页码：1745~1754
关键词：强壮前沟藻去藻滤液；粗提物；化感物质；溶藻效应；GC/MS分析；
Keywords:cell-free filtrate of A. carterae, crude extraction, allelochemicals, dissolving algae, GC/MS,
摘 要 ：微藻化感作用是一种极其复杂的生理、生态学现象。选取强壮前沟藻指数生长初期Ⅰ和平台生长初期Ⅱ两个阶段的滤液对中肋骨条藻、海洋原甲藻、锥状斯氏藻及球等鞭金藻生长的影响进行了研究,并萃取了阶段Ⅱ的粗提物,抑藻检测表明其具有"杀藻"效应,通过GC/MS分析该粗提物中具有潜在化感作用的物质种类。研究发现强壮前沟藻两个生长阶段的滤液对中肋骨条藻均产生强烈致死效应(phaseⅠ: F=15.18475, P=0.00298<0.05; phaseⅡ:F=6.24559, P=0.03149<0.05);锥状斯氏藻在强壮前沟藻滤液中生长,实验结束时两个阶段中的细胞密度分别是对照组的79.3% 和 68.9%;海洋原甲藻在强壮前沟藻生长阶段Ⅱ滤液实验的最后3d,其生长受到显著抑制(F=4.84438, P=0.04925<0.05);而等鞭金藻在强壮前沟藻两个生长阶段滤液中被抑制现象不明显(P>0.05)。强壮前沟藻滤液实验表明,强壮前沟藻能够向微环境中分泌代谢产物来抑制中肋骨条藻和海洋原甲藻的生长,并且这种抑制效应具有种类特殊对应性。上述实验结果还表明,强壮前沟藻生长阶段Ⅱ的滤液具有的生长抑制作用较为明显。采用乙酸乙酯萃取强壮前沟藻生长阶段Ⅱ滤液中的代谢产物,检测发现其代谢粗提物具有溶藻效应,GC/MS分析结果表明粗提物中存在4种可能产生化感抑制作用的物质,其中二丁基羟基甲苯(Butylated Hydroxytoluene BHT)被认为具有抗滤过性病原体和抗微生物活性。
Abstract:Algal allelopathy is an ecological or physiological phenomenon that has focused attention on the interactions among algae and the production of algal toxins. A species of marine bloom-forming microalga: Amphidinium carterae was selected in the present study to estimate its potentially allelopathic effects on other species of microalgae. The cell-free filtrates of A. carterae in two growth phases: exponential grwoth phase (abbreviated as growth phase Ⅰ) and stationary growth phase (abbreviated as growth phase Ⅱ) were prepared, and its inhibitory effects on growth of other species of marine microalgae were studied under stable environmental conditions (temperature, light and illuminating periodicity, etc.). The cell-free culture filtrates were prepared firstly when A. carterae were cultured phaseⅠand phaseⅡ, which was about the the growth 7th and 17th respectively, and enriched with f/2 medium. The above prepared culture filtrates were used to culture 4 species of marine microalgae: Skeletonema costatum、Prorocentrum micans、Scrippsiella trochoidea and Isochrysis golbana. Moreover, the crude extractions in the cell-free filtrate of A. carterae of growth phaseⅡ were extracted by GC/MS, and the major components were analyzed. Results showed that: in the enriched cell-free filtrates, S. costatum were obviously inhibited in either filtrates of phase I (F=15.18475, P=0.00298<0.05) or phase Ⅱ(F=6.24559, P=0.03149<0.05) as compared to that in the control which in f/2 enriched medium, but the cell numbers of S. trochoidea were only about 79.3% and 68.9% in filtrates prepared from phaseⅠand phaseⅡ as compared to the control by the end of experiment. As to P. micans, its growth was obviously suppressed (F=4.84438, P=0.04925<0.05) in the filtrate prepared from phaseⅡduring the last 3 days. Little inhibition was observed in I. golbana in either filtrates prepared from either phases (P>0.05). Cell-free filtrate cultures experiment of A. carterae is one of the basic approaches on allelopathy research. Results in the present study suggested that A. carterae secreted or released some substances into the culture medium, which inhibited the co-cultured S. costatum and P. micans, and these inhibitory effects could not caused by light, nutrients or cell contact to gain a competitive advantage. Many algae could release allelochemicals into the environment which may alter phytoplankton growth to enhance its predominance. Besides, growth of these two species was more inhibited in enriched cell-free filtrate prespred from growth phaseⅡ than that from phaseⅠ. It also proved that the allelochemical effects on algae were species-specific because growth of S. costatum and P. micans was inhibited, but exerted little effects on S. trochoidea and I. golbana. Another experiment was performed to further prove the allelopathic effect of A. carterae.The crude extractions from cell-free filtrates of A. carterae at different concentrations were added into S. costatum and P.micans culture media, found that cells dissolved in higher concentration after many hours. It is noted that the crude extractions had the function of dissolving some algal cells. Then the crude extractions were analyzed by GC/MS. Four components in the crude exactions were suspected to have the allelopathic effects, they were Butylated Hydroxytoluene, Butyl citrate, Benzene, 1-Chlorodifluoromethoxy-4-nitro-, and Benzyl butyl phthalate, among with Butylated Hydroxytoluene (BHT) was reported with oxidative characteristic and was used as an antioxidant additive. Results in the present study brought light on the hypothesis that A. carterae could realese allelochemical(s); however, deep research should carry out to explain the possible inhibiting mechanism.