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The IHSS soil humic acid (HA) standard and two HAs from soils of very different origin (Chernozem and Ranker) were fractionated by tandem size-exclusion chromatog raphy−polyacrylamide gel electrophoresis. From each HA, three fractions with different molecular sizes (MSs) and electrophoretic mobilities were obtained and investigated for their fluorescence properties and ability to photoinduce the transformation of 2,4,6-trimethylphenol and herbicide fenuron. Regardless of the source of the HA, the two high MS fractions were found to be very weakly fluorescent. They photoinduced the degradations of fenuron and 2,4,6-trimethylphenol less efficiently than the bulk HA (10−50-fold and 1.4−5.3-fold, respectively). In contrast, the low MS fraction was proved to be fluorescent and to photoinduce the transformation of probes as least as efficiently than the bulk HA. These results show that (i) most of fluorophores and a great part of photoinductive chromophores are located in the low MS fractions of soil HAs and (ii) this distribution of photochemically active constituents may be characteristic across broad soil types.