Determination of molecular formulas and structural regularities of low molecular weight fulvic acids by size-exclusion chromatography with electrospray ionization quadrupole time-of-flight mass spectrometry.

ABSTRACT

Size-exclusion chromatography was coupled to a quadrupole time-of-flight mass spectrometer using electrospray ionization in the negative ion mode to analyze Suwannee River fulvic acid. About 220 exact masses of low molecular weight fulvic acids (FAs) were measured in the range from m/z 190 to 350, and molecular formulas could be deduced for all these molecular masses. All molecular formulas can be described by the superimposition of three homologous series in the FA mixture (a) a series of hydrogen homologs constituted of five to six members for a given number of carbons and oxygens; (b) a series of alkyl chain homologs with stepwise addition of one methylene group; and (c) isobaric compounds that differ by the formal replacement of an oxygen atom by a methane group. Product ion spectra of the FA species reflect the repeated parallel losses of carbon dioxide and water. The minimum number of carboxylate groups and the maximum number of hydroxy moieties could be determined from the product ion spectra. Furthermore, it was obvious that the structural differences between homologs in the three series are located in the carbon backbone of the FA rather than in its extremities that are expelled as neutral fragments. These structural regularities reduce the complexity of FA mixtures to a certain set of yet unknown basic structures.