Distinguishing chemically induced NADPH- and NADH-related metabolic responses using phasor analysis of UV-excited autofluorescence.

NADPH and NADH are well known for their role in antioxidant defense and energy metabolism, respectively, however distinguishing their cellular autofluorescence signals is a challenge due to their nearly identical optical properties. Recent studies applying spectral phasor analysis to autofluorescence emission during chemically induced metabolic responses showed that two-component spectral behavior, , spectral change acting as a superposition of two spectra, depended on whether one or multiple metabolic pathways were affected. Here, we use this property of spectral behavior to show that metabolic responses primarily involving NADPH or NADH can be distinguished.