高速液体クロマトグラフィーによる天然水中の溶存金属酵素アルカリホスファターゼの同定

概要

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Metalloenzyme (alkaline phosphatase) dissolved in lake water has been identified by means of high performance liquid chromatography (HPLC). Water samples were collected from Shinobazu-no-ike Pond (Tokyo) and Lake Kasumigaura (Ibaraki), and filtered with a 0.45 μm Millipore filter just after sampling. Filtered water was concentrated to (500〜1000) fold by ultrafiltration, where ultrafiltration membrane (Amicon UM-10) which concentrated the components of molecular weight larger than 10000 was used. The enzymatic activity of alkaline phosphatase was measured using p-nitrophenyl phosphate as a substrate. Addition of EDTA (ethylenediamine tetraacetic acid) to the concentrated lake water did not show any enzymatic activity, while the enzymatic activity was recovered with addition of zinc ion. The HPLC chromatograms of the concentrated lake water were examined, where an aqueous porous gel column W-71 from Shimadzu Co. Ltd., was used for separation. According to the chromatorams, the efHuent at 12.2 min was corresponding to alkaline phosphatase. The final identification was made by adding E. coli alkaline phosphatase to the concentrated lake water and by measuring the liquid chromatogram. Zinc was also found in the fraction of the effluent between 11.3 and 12.3 min. More details of the experiments will be published in near future.
社団法人日本分析化学会の論文
1980-08-05