Estimating the helical pitch angle of amylopectin in starch using polarization secondharmonic generation microscopy

Abstract

Starch granules are among the brightest natural second harmonic generation (SHG)converters. They basically consist of amylose and amylopectin molecules and the source ofthe SHG signal is still undetermined. In the present study we perform polarizationsensitive SHG (PSHG) imaging of wheat starch granules and we fit the SHGsignal variation of each pixel of the PSHG images into a generalized biophysicalmodel. By assuming that the SHG source molecule is a helix with cylindricalsymmetry along its long axis, the model extracts the helical pitch angle of the SHGsource for every pixel of the image. By displaying the pixel histogram representingthis helical pitch angle we found a highly peaked histogram with maximum atθe = 36.1° and awidth of Δθe = 9.3°. This pitch angle corresponds to the strand of the parallel double helical structure, calledamylopectin (as measured by a small angle x-ray scattering technique). This demonstratesamylopectin (and not amylose) as the source of SHG in starch. According to ourknowledge, this is the first estimation of the amylopectin’s pitch angle in starch usingPSHG.