Empirical method for rapid quantification of intrinsic fluorescence signals of key metabolic probes from optical spectra measured on tissue-mimicking turbid medium

Significance: Optical fluorescence spectroscopy technique has been explored extensively to quantify both glucose uptake and mitochondrial metabolism with proper fluorescent probes in small tumor models in vivo. However, it remains a great challenge to rapidly quantify the intrinsic metabolic fluorophores from the optically measured fluorescence spectra that contain significant distortions due to tissue absorption and scattering. Aim: To enable rapid spectral data processing and quantify the in vivo metabolic parameters in real-time, we present an empirical ratio-metric method for rapid fluorescence spectra attenuation correction with high accuracy. Approach: A first-order approximation of intrinsic fluorescence spectra can be obtained by dividing the fluorescence spectra by diffuse reflectance spectra with some variable powers. We further developed this approximation for rapid extraction of intrinsic key metabolic probes (2-NBDG for glucose uptake and TMRE for mitochondrial function) by dividing the distorted fluorescence spectra by diffuse reflectance intensities recorded at excitation and emission peak with a pair of system-dependent powers. Tissue-mimicking phantom studies were conducted to evaluate the method. Results: The tissue-mimicking phantom studies demonstrated that our empirical method could quantify the key intrinsic metabolic probes in near real-time with an average percent error of ∼5%. Conclusions: An empirical method was demonstrated for rapid quantification of key metabolic probes from fluorescence spectra measured on a tissue-mimicking turbid medium. The proposed method will potentially facilitate real-time monitoring of key metabolic parameters of tumor models in vivo using optical spectroscopy, which will significantly advance translational cancer research.