Protein is one of the most important fluorophores and it exists in most intracellular and extracellular substances. In fact, many biomolecules, including proteins, enzymes, coenzymes, pigments and primary or secondary metabolites (e.g., fulvic and humic acids), have been found to exhibit a characteristic fluorescence 3. The use of fluorescence is based on the fact that all microorganisms contain natural intracellular and extracellular fluorophores, whose concentrations depend on the physiological state of the cells 1, 2. Three-dimensional excitation–emission matrix (EEM) fluorescence spectroscopy as a rapid, selective and sensitive technique is very helpful for bacterial metabolism monitoring. Successful biological wastewater treatment process depends largely on high bacterial activity. A specific method for fluorescence monitoring of anammox reactor were put forward, which included typical fluorescence indexes and their possible values for different operation phases.

Correlation analysis between different fluorophores discovered that the two peaks in each of these three groups-two intracellular protein-like peaks, two humic acid-like peaks and the two extracellular protein-like peaks had strong intercorrelation, which gave evidence of their homology. The two intracellular protein-like peaks had high correlation with MLVSS and MLVSS growth rate. Correlation analysis revealed that intracellular 420 peak and humic-like peak had strong correlation with nitrogen removal rate. Two main protein-like fluorophores were identified from the extracellular substances. Four main fluorophores were identified from the intracellular substances.

In this work, excitation–emission matrix (EEM) fluorescence spectroscopy was used to characterize the intracellular and extracellular substances of anammox sludge during reactor operation of 276 days. Anammox is a cost-effective process to treat nitrogenous wastewater.

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