Excitation wavelength-specific changes in lignocellulosic autofluorescence

Lignocellulose (paper sheets of thermomechanical pulp) was irradiated with specific wavelengths in an epifluorescence microscope equipped with filter cubes with excitation and emission filters commonly used for excitation of fluorophores with blue, green, and red emissions. The irradiation was shown to affect the relative autofluorescence intensity over time. Shorter wavelength excitation (lambda(ex) = 340-380 nm) induced photoyellowing that increased the autofluorescence. Filter cubes allowing irradiation at longer wavelengths (lambda(ex) = 450-490 nm and lambda(ex) = 515-560, respectively) caused photobleaching and a decrease in lignocellulosic autofluorescence. The increase and decrease in autofluorescence was reversible; prephotoyellowed samples could be photobleached and prephotobleached samples regained autofluorescence by shorter wavelength irradiation. Thus, the specificity of the excitation wavelength and the time-dependent autofluorescence might affect long-term imaging experiments of lignocellulose-based samples.