| FDA水解酶分析法表征近海泥滩微生物活性 |
| 摘要点击 1671 全文点击 1258 投稿时间：2013-01-19 修订日期：2013-04-26 |
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| 中文关键词 海洋泥滩沉积物 微生物活性 FDA水解酶分析 荧光光度法 |
| 英文关键词 coastal intertidal mudflat microbial activity FDA hydrolase analysis fluorescence spectrometry |
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| 中文摘要 |
| 针对近海潮间带环境高盐、有机质组成复杂和微生物活性低的特点,根据微生物酶解荧光素双醋酸酯(FDA)产生荧光素的原理,建立了适于泥滩类潮间带沉积环境中微生物活性检测的荧光光度法. 从样品浸提液、反应产物检测仪器、预处理方法、实验条件等不同方面进行优化,确定泥滩类潮间带沉积环境中微生物活性的最优检测方法和条件如下:沉积物湿样以灭菌陈海水为介质,加入吐温-80分散剂,充分振荡后静置,使较大颗粒自然沉降; 取上层悬浊液,经无菌滤膜(1.2 μm,多次煮沸灭菌)过滤,得到待测菌液; 取待测菌液加入适量FDA溶液,在25~30℃ 下避光反应180 min,以丙酮为终止剂终止反应,25 min内以分子荧光光度计(激发波长488 nm,发射波长530 nm)测定反应产物的荧光强度,该方法的检测范围(以干重计)是3.0×103~1.1×105个·g-1. 微生物活性以单位质量样品的荧光素含量表示(μg·g-1,干重). |
| 英文摘要 |
| A method based on fluorescence spectrometry was developed to detect the microbial activities in marine mudflat sediment, where is characterized by high salinity, complex organic compounds and low microbial biomass. This paper optimized the sample extracts, the detection equipment for reaction products, the pretreatment methods, and the experimental conditions. The optimal procedure is described as following. Fresh sediment was first extracted with sterilized and aged seawater, followed by the addition of Tween-80 solution, then uniformly dispersed by thorough oscillating, and kept steady for precipitation. After filtration through a sterilized membrane (1.2 μm, sterilized in boiling water repeatedly), the supernatant was supplemented with an appropriate amount of FDA solution and allowed to react in dark for 180 min at temperature ranged 25-30℃. The reaction was terminated by the addition of acetone, and the fluorescence intensity of the reaction mixture was measured within 25 min using a molecular fluorescence photometer at an excitation wavelength of 488 nm and an emission wavelength of 530 nm, and the detection range of this method (dry weight) was 3.0×103-1.1×105 ind·g-1. The microbial activity was reported as fluorescence content in per unit sediment mass (μg·g-1, dry weight). |
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