Molecular Orientation in Dry and Hydrated Cellulose Fibers: A Coherent Anti-Stokes Raman Scattering Microscopy Study

Abstract

Coherent anti-Stokes Raman scattering (CARS) microscopy is combined with spontaneous Raman scattering microspectroscopy and second harmonic generation (SHG) microscopy to interrogate the molecular alignment in dry and hydrated cellulose fibers. Two types of cellulose were investigated: natural cellulose I in cotton fibers and regenerated cellulose II in rayon fibers. On the basis of the orientation of the methylene symmetric stretching vibration, the molecular alignment of cellulose microfibrils is found to be conserved on the micrometer scale. Whereas the molecular orientation in cotton shows modest variability along the fiber, the alignment of the cellulose units in rayon is highly consistent throughout the fiber. The ordered alignment is retained upon fiber hydration. Upon hydration of the cellulose fibers, an anisotropic electronic contribution is observed, which indicates an ordered incorporation of water molecules into the fiber structure. The third-order and second-order electronic polarizability of cellulose I are directed along the axis of the polyglucan chain. No second-order optical response is observed in cellulose II, supporting the antiparallel arrangement of the polyglucan chains in regenerated cellulose.

Visualization of structural changes in cellulosic substrates during enzymatic hydrolysis using multimodal nonlinear microscopy

“…Glucan chains lack inversion symmetry and have previously been shown to give rise to SHG in starch (Brackmann et al 2011;Slepkov et al 2010) and cellulosic substrates of different origin Brown et al 2003;Glas et al 2015;Pohling et al 2014;Slepkov et al 2010;Zimmerley et al 2010). In order to interpret the SHG images of cellulosic substrates, the coherent nature of the SHG signal and the supramolecular structure of cellulose must be taken into account.…”

“…The excitation power at the sample was approximately 40 mW for each of the excitation beams. CARS experiments were conducted at a Raman shift of 2880 cm -1 , which is characteristic of the C-H stretching vibration of the CH 2 group in cellulose (Zimmerley et al 2010). The 1064 nm output was used as the Stokes beam and the output of the OPO was tuned to 814 nm and used as the pump beam, resulting in CARS emission at 659 nm.…”

“…This allows intrinsic 3D optical sectioning, since the signal is effectively generated only in the most intense part of the focused beam. SHG can be used to probe non-centrosymmetric structures, and has been used to image cellulose in cotton (Slepkov et al 2010;Zimmerley et al 2010) and rayon fibres (Zimmerley et al 2010), cellulose derived from the bacterium Acetobacter xylinum Brown et al 2003) and the algae Valonia ventricosa (Brown et al 2003). Multiphoton excited fluorescence (MPEF) can be used to probe the intrinsic fluorescence of the cellulosic substrate, i.e.…”

“…Coherent Raman scattering techniques, coherent anti-Stokes Raman scattering (CARS) and stimulated Raman scattering, can probe molecular vibrations, allowing chemically sensitive imaging, for example to image carbohydrates and lignin in plant cell walls. Lignin is generally imaged by probing aromatic ring stretching vibration at 1600 cm -1 (Chen et al 2010;Ding et al 2012;Pohling et al 2014;Saar et al 2010;Zeng et al 2010), while polysaccharides can be imaged either by probing C-C and C-O stretching vibrations at *1100 cm -1 (Chen et al 2010;Saar et al 2010) or C-H stretching vibrations at *2900 cm -1 (Ding et al 2012;Pohling et al 2014;Zimmerley et al 2010). If full Raman spectrum is recorded, lignin and different types of polysaccharides can be distinguished using spectral fitting (Pohling et al 2014).…”

Spectrally-broad coherent anti-Stokes Raman scattering hyper-microscopy utilizing a Stokes supercontinuum pumped at 800 nm

“…Additionally, a nearly-identical image is obtained via contrast for benzonitrile in the "silent region" at 2240 cm 1 (not shown), however, we chose 3074 cm 1 and 2934 cm 1 as the two CARS contrasts to highlight the spectral sensitivity of our SF-CARS system. Stimulated vibrational imaging of cellulose, both at the fingerprint [17,30] and CH-OH frequency ranges [31][32][33], has been previously demonstrated. In our sample, the CARS signal from cellulose at 1100 cm 1 is detected, but the signal is largely swamped by the large NRB from the solvents that soak the fibre.…”