Abstract

Total luminescence spectra are representations of luminescence intensity as a function of excitation wavelength (λ<sub>ex</sub>) on one axis and emission wavelength (λ<sub>em</sub>) on the other; they are also referred to as excitation-emission matrices (EEM). A ridge due to first-order scattered light will appear on a diagonal across the spectrum, with the peak intensity centered at λ<sub>em</sub> = λ<sub>ex</sub>. Diagonal ridges due to higher-order scattered light may also appear (λ<sub>em</sub> = <i>n</i>λ<sub>ex</sub> for <i>n</i>th-order scattered light). The scattered light contribution is easily recognized but not easily removed from the EEM. Highly scattering samples such as protein-containing biological samples, micellar systems, environmental samples, and microbial samples are all at risk of having important fluorescence information obscured by the presence of the scattered light peaks. Scattered light will appear as a component in the factor analysis of EEMs, adding to the complexity of the data analysis and interpretation. Spectral fingerprinting of highly scattering samples may be hampered by the loss of information near the scattered light peaks. For example, the emission spectra of polycyclic aromatic hydrocarbons (PAH) are often shifted from the excitation spectra by only a small amount, so that there is a substantial amount of information in the region of the scattered light ridge for PAH-containing samples.

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