An Electron Diffraction Study on Litnerized Potato Starch

Starch is arranged in a crystalline manner within its storage granules and should thus give sharp X-ray diagrams. Unfortunately most of the common starch granules have sizes between 1 and 100μm, making them too small for an X-ray study on individual grains. There is only one instance where an oriented X-ray diagram could be obtained on one sector of an individual giant starch granule. Despite their small size, starch granules are still too thick to be studied by electron diffraction with a transmission electron microscope. The only reported study on starch ultrastructure using electron diffraction on frozen hydrated material was made on small fragments. The present study has been realized on thin sectioned granules previously litnerized to improve the signal to noise ratio.Potato starch was hydrolyzed for 10 days in 2.2N HCl at 35°C, dialyzed against water until neutrality and embedded in Nanoplast. Sectioning was achieved with a commercially available low-angle “35°” diamond knife (Diatome) after a very carefull trimming and a pre-sectioning with a classical “45°” diamond knife. Sections obtained at a final sectioning angle of 42.2° (compared with the usual 55-60°) and at a nominal thickness of 900Å were collected on a Formvar-carbon coated grid. The exact location of the starch granules in their sections was recorded by optical microscopy on a Zeiss Universal polarizing microscope (Fig. 1a). After rehydration at a relative humidity of 95% for 24 hours they were mounted on a Philips cryoholder and quench frozen in liquid nitrogen before being inserted under frozen conditions in a Philips EM 400T electron microscope equipped with a Gatan anticontaminator and a Lhesa image intensifier.