Development of correlation spectroscopy (COS) method for analyzing fluorescence excitation emission matrix (EEM): A case study of effluent organic matter (EfOM) ozonation.

Two-dimensional correlation spectroscopy (2DCOS) has been used as a powerful tool for analyzing spectral features, but it has never been applied to fluorescence excitation-emission matrix (EEM) data due to the incompatible dimensions. This study first investigated EEM-COS by reducing the dimensions of the EEM (using parallel factor analysis, PARAFAC) for fitting to 2DCOS (EEM-PARAFAC-COS). The fluorescence changes of effluent organic matter (EfOM) during ozonation were studied using EEM-COS and synchronous fluorescence (SF)-2DCOS. The conventionally used SF-2DCOS proved to be biased due to the intrinsic drawback of SF, while the EEM-PARAFAC-COS gave accurate and trustworthy results. Homo-EEM-PARAFAC-COS indicated that the fluorescence protein-like and fulvic-like substances in EfOM were preferentially ozonated compared to humic-like substances. Hetero-EEM-PARAFAC-COS analyses on the EEM, FTIR, UV-vis absorbance, and size-exclusion chromatography showed that the fluorescence protein-like and fulvic-like substances in EfOM were associated with lower molecular weight (MW, ∼0.95 kDa), UV absorbance at ∼280 nm, and more electron-enriched aromatics (with amide and phenolic groups), which explained their ozonation preference, while humic-like substances were related to carboxylic groups, UV absorbance at ∼255 nm, and organics at MW of ∼4.50 kDa. This work demonstrated the great potential of EEM-PARAFAC-COS in studying fluorescence change and correlating fluorescence with other spectra.