| The impact of granule hydration and starch granule-associated proteins (SGAPs) on the reactivity of maize and wheat starches (including both normal and waxy genotypes) was investigated. The fluorescent probe, 5-(4,6-dichlorotriazinyl)aminofluorescein (DTAF), was utilized to derivatize starches in granular form. The granular locale of reaction and molecular reactivity of starch chains (i.e., amylose [AM] and amylopectin [AP] branch chains, including long [LC], intermediate [IC], and short [SC] chains) were highlighted by confocal laser scanning microscopy (CLSM) and size-exclusion chromatography (SEC) coupled to refractive index (RI) and fluorescence (FL) detection, respectively. Progressing from limited (≤ 25.56 g moisture/100 g starch) to full (≥ 32.64 g moisture/100 g starch) starch hydration, reaction evolved from granule surfaces (including those of channels and cavities) into the granule matrix, resulting in dramatic increases in starch chain reactivities. This phenomenon was attributed to the occurrence of a glass transition, by which waxy (relative to normal) starches exhibited greater extents of hydration/swelling (i.e., plasticization), more homogeneous granular reaction patterns, and greater overall molecular extents of reaction. Given sufficient hydration (i.e., aqueous reaction system), AP-LC was the starch chain fraction responsible for majority of reaction (followed by AM, AP-IC, and AP-SC), indicating that AP-LC are located within granule amorphous regions, which are most susceptible to reaction. AM participated in the reaction only when starch granules were hydrated to a critical degree, suggesting that AM may be intertwined with AP crystalline domains and/or oriented differently (i.e., radial arrangement) within granules relative to AP-LC.;SGAP removal also facilitated reagent diffusion into granule interiors to subsequently enhance the overall molecular extent of reaction (with the exception of normal wheat starch, due to its relatively inaccessible channel structures), though this effect was somewhat compensated for by sufficient hydration of starch granules (i.e., facilitating reaction within the granule matrix). Despite similar molecular reactivities for protease and non-proteasetreated starches reacted in fully aqueous slurries, a greater modification effect (i.e., cross-linking) was achieved with starches first subjected to proteolysis, particularly for waxy starches that lacked GBSSI matrix proteins GBSSI. Mechanisms for this finding are likely attributable to a greater extent of reaction within granule interiors rather than at exteriors and/or more focused reaction of starch chains, rather than protein molecules. |
