Fluorescence Inner-Filtering Correction for Determining the Humification Index of Dissolved Organic Matter

TLDR

Fluorescence spectrometry has been proposed as a simple method to assess natural organic matter humification by measuring the red‑shift of fluorescence emission, but indices calculated without standardizing dissolved organic matter concentration vary with concentration due to inner‑filtering effects. This study aimed to evaluate how dissolved organic matter concentration affects humification index determination using samples from field corn extract, soil:water extract, and soil fulvic acid. Humification indices were calculated as the ratio of fluorescence intensity at longer wavelengths to that at shorter wavelengths, and corrected for inner‑filtering either by extrapolating to 100 % transmittance or by applying explicit primary and secondary correction factors, yielding concentration‑invariant values when absorbance at 254 nm was below ~0.3. The study found that index values are linearly related to solution transmittance at 254 nm, that defining the index as the 300–345 nm intensity divided by the sum of 300–345 nm and 435–480 nm intensities is statistically advantageous, and that correcting for inner‑filtering is essential for quantitative comparison across studies.

Abstract

The use of fluorescence spectrometry has been suggested as a simple method to determine the extent of natural organic matter humification by quantifying the red-shifting of fluorescence emission that occurs with increasing humification. Humification indices are calculated by dividing fluorescence intensity at longer wavelengths by intensity at shorter wavelengths. These indices calculated without any specific efforts to standardize dissolved organic matter (DOM) concentration will result in index values that vary with DOM concentration due to fluorescence inner-filtering effects. This study critically evaluated the effect of DOM concentration on humification index determination using organic matter isolated from field corn extract, soil:water extract, and soil fulvic acid. The results show that humification index values are sensitive to DOM concentration of the solution and are linear with respect to transmittance of the solution at the 254 nm used as the excitation wavelength. An approximate correction for DOM is to exploit the linear nature of the regression fit and to determine index values at the extrapolated 100% transmittance value. An exact correction using explicit correction factors for both primary and secondary inner-filtration effects was shown to give humification index values that are concentration invariant when absorbance of the solution at 254 nm was less than approximately 0.3 unit. Defining the humification index as the fluorescence intensity in the 300→345 nm region divided by the sum of intensity in the 300→345 nm and 435→480 nm regions was statistically advantageous. This study suggests that for quantitative results which can be used to compare humification of natural organic matter across different studies, correction of the fluorescence emission spectra for inner-filtration effects is needed.