Polarimetric second harmonic generation microscopy: An analytical tool for starch bioengineering

Abstract

Second harmonic generation (SHG) is a nonlinear optical process that inherently generates signal in non-centrosymmetric materials, such as starch granules, and therefore can be used for label-free imaging. Both intensity and polarization of SHG are determined by material properties that are characterized by the nonlinear susceptibility tensor, x (2) . Examination of the tensor is performed for each focal volume of the image by measuring the outgoing polarization state of the SHG signal for a set of incoming laser beam polarizations. Mapping of nonlinear properties expressed as the susceptibility ratio reveals structural features including the organization of crystalline material within a single starch granule, and the distribution of structural properties in a population of granules. Isolated granules, as well as in situ starch, can be analyzed using polarimetric SHG microscopy. Due to the fast sample preparation and short imaging times, polarimetric SHG microscopy allows for a quick assessment of starch structure and permits rapid feedback for bioengineering applications. This article presents the basics of SHG theory and microscopy applications for starch-containing materials. Quantification of ultrastructural features within individual starch granules is described. New results obtained by polarization resolved SHG microscopy of starch granules are presented for various maize genotypes revealing heterogeneity within a single starch particle and between various granules.

Physical, Chemical and Rheological Characterization of Tuber and Starch from Ceiba aesculifolia subsp. parvifolia

“…Figure 5 depicts the FT-IR spectra corresponding to CAP tuber starch. Starch granules consist of two different water types, according to their location; one of these refers to water as a part of the hydrogen bonding network within the crystalline structure and another that is more mobile and considers hydration water [ 76 ]. Bands in the region 3000−3800 cm −1 may be attributed to O−H stretching regions of water molecules hydrogen-bonded [ 77 ], whereas the band observed around 2100 cm −1 is assigned to –OH bending vibration of free water molecules due to its hygroscopic nature [ 78 ].…”

Study of a bi-axial (KTP) crystal using Double Stokes Mueller Polarimetry

“…[16,28]. The susceptibility ratios determined using DSMP give information about the structural properties of crystalline material in single starch granules as well as a lump of starch granules [16,29]. The entropy of the susceptibility tensors is determined for z-cut quartz crystal and collagen of rat tail tendon.…”

Dual‐ and single‐shot susceptibility ratio measurements with circular polarizations in second‐harmonic generation microscopy

“…Second‐harmonic generation (SHG) microscopy is a powerful technique for imaging noncentrosymmetric biomaterials such as collagen fibers , myosin filaments , microtubules , and starch granules . The SHG efficiency is characterized by the second‐order susceptibility tensor χ 2 whose elements depend on the composition and the 3D organization of the SHG‐active molecules.…”

“… is the susceptibility tensor element for which the generated second harmonic electric field and incident light electric field are along the C 6 symmetry axis of the molecular fiber, such as myofibril or collagen fiber ( z ) and is the susceptibility tensor element for which the SHG electric field is along z ‐axis and the incident light electric field is perpendicular to it. The R ‐ratio has been previously used to characterize different biomaterials such as tendon , bone , muscle , and starch . The R ‐ratio was further used to differentiate between normal and cancerous tissues in various organs such as lung , breast , thyroid , and pancreas .…”