Productivity in P limited peatlands is regulated in part by the turnover of organic phosphates, which is influenced by the chemical nature of the compounds involved. We used solution 31P nuclear magnetic resonance (NMR) spectroscopy to quantify organic and inorganic phosphates in benthic floc (a mixture of plant detritus and algae) and underlying soil from sites along P gradients in hard water and soft water areas of the northern Florida Everglades, USA. Phosphorus-enriched sites were dominated by cattail (Typha spp.), while unenriched sites included sawgrass (Cladium jamaicense Crantz) ridges and open-water sloughs. Phosphorus extracted in a solution containing 0.25 M NaOH and 50 mM EDTA (ethylenediaminetetraacetate) included phosphate, phosphate monoesters, DNA, and pyrophosphate. Signals from phosphate monoesters were consistent with those from alkaline hydrolysis products of RNA and phospholipids formed during extraction and analysis, whereas phytic acid (myo-inositol hexakisphosphate), the most abundant organic phosphate in most soils, was not detected. Phosphorus composition was similar among sites, although neither DNA nor pyrophosphate were detected in extracts of benthic floc from a calcareous slough. DNA was a greater proportion of the P extracted from soil compared to benthic floc, while the opposite was true for pyrophosphate. Research on the cycling of organic phosphates in wetlands focuses conventionally on the turnover of phosphate monoesters, but our results suggest strongly that greater emphasis should be given to understanding the role of phosphate diesters and phosphodiesterase activity.