Measurements of phosphatase activity in peatlands are made difficult by the low levels of activity and the characteristically high concentrations of dissolved organic matter within the sediments. These materials may cause high background absorbances in colorimetric assays, and quenching interference in fluorimetric assays. This review describes the development of a new approach which allows such problems to be overcome by using HPLC to separate the interferences from the products of enzymic hydrolysis. This approach is applicable to various environmental samples such as peat, wetland sediment, and sludge which may contain a large amount of interfering organic matters.