Anwendung einer enzymatischen Methode zur Charakterisierung von Huminstoffen Enzymatic Characterization of Humic Substances

Abstract

Summary: A method is described for the specific hydrolysis of amino acids from humic substances by using the proteolytic enzyme "Pronase E". The amino acids are analyzed by a fluorescence detector after precolumn derivatization and HPLC-separation. The pattern and the concentration of amino acids after the enzymatic hydrolysis are set up for humic substances of different origin. Due to the results the enzymatic hydrolysis is suitable for the characterization of humic substances with a nitrogen content higher than 3%. As shown in a kinetic study of the hydrolysis of a fulvic acid the influence of humic substances on the enzyme activity increases in case of lower nitrogen content. Due to the specificity of the proteolytic enzyme the operationally defined method is limited as shown by a protein of a known amino acid content. The amino acids leucine, valine, tyrosine and histidine are released to the highest extent. The determined total extent for the hydrolysis of Insulin B is 26.34% and for Casein 38.37%. By comparison of the enzymatic hydrolysis with the classic acid hydrolysis the advantages of the biochemical method are the simple experimental handling and the mild reaction conditions. Amino acids which are very unstable like glutamine and asparagine can only be analyzed by the enzymatic hydrolysis.

Degradation Pathway of the Photochemical Oxidation of Ethylenediaminetetraacetate (EDTA) in the UV/H2O2-process

“…A Hewlett Packard (HP) 1090 liquid chromatograph with a HP 79883 B diode array detector and a HP 1046 A fluorescence detector were used. Glycinate was determined by a HPLC-method especially developed for amino acids [17]. FeEDTA, oxamate, and glyoxylate were determined by isotachophoresis [10], oxalate, formate, nitrate, nitrite, cyanate by HPIC [16].…”

Detection of glucosamine in the acid hydrolysis solution of humic substances

“…Released amino acids and glucosamine were determined by a fluorescence detector (Hewlett Packard 1046 A) after precolumn derivatization (o-phthalaldehyde, 9-fluorenylmethylchloroformate) and HPLC-separation (Hewlett Packard 1090 A). A detailed description of the method was given in [5].…”

“…The total nitrogen content was determined with the method of Dumas following the combustion of the samples. The gaseous products were detected after an reducing step with an apparatus from Heraeus (Hanau, Germany) [5].…”

“…The hydrolysis procedure was carried out with hydrochloric acid for 15 h at 100°C as described in [5].…”

“…They are polyfunctional refractory substances, which are fairly resistant to biological decomposition [1,2]. Because HS contribute significantly to the biogeochemically important bound nitrogen in the biosphere, there is a great interest to characterize the bound nitrogen forms of these compounds [3][4][5][6][7]. The nitrogen content of soil humic acids (HA) amounts to 6%, while soil fulvic acids (FA) only contain 3% at most [4].…”