Exotic Corn Lines with Increased Resistant Starch and Impact on Starch Thermal Characteristics

Cereal Chem. 87(3):190–193 Ten parent corn lines, including four mutants (dull sugary2, amyloseextender sugary2, amylose-extender dull, and an amylose-extender with introgressed Guatemalen germplasm [GUAT ae]) and six lines with introgressed exotic germplasm backgrounds, were crossed with each other to create 20 progeny crosses to increase resistant starch (RS) as a dietary fiber in corn starch and to provide materials for thermal evaluation. The resistant starch 2 (RS2) values from the 10 parent lines were 18.3–52.2% and the values from the 20 progeny crosses were 16.6–34.0%. The %RS2 of parents was not additive in the offspring but greater RS2 in parents was correlated to greater RS2 in the progeny crosses (r = 0.63). Differential scanning calorimetry (DSC) measured starch thermal characteristics, revealing positive correlations of peak gelatinization temperature and change in enthalpy with %RS2 (r = 0.65 and r = 0.67, P ≤ 0.05); however, % retrogradation (a measure of RS3) and retrogradation parameters did not correlate with %RS2. The %RS2 and onset temperature increased with the addition of the ae gene, likely because RS delays gelatinization. Four types of resistant starch (RS) have been defined. RS1 is resistant because of the surrounding food matrix; RS2 is present in ungelatinized, raw starches; RS3 is created by retrogradation; and RS4 is produced through chemical alteration (Englyst et al 1996). Incorporation of RS into the diet provides many health benefits: it serves as a prebiotic, or fermentable substrate, for the growth of probiotics; lowers the pH of the colon (Cherrington et al 1991); increases mineral absorption (Courdray et al 1997); and increases cell turnover (Young et al 2005). Cholesterol metabolism may be down-regulated by RS, by production of short-chain fatty acids that may either suppress cholesterol synthesis in the liver (Hara et al 1999) or decrease cholesterol absorption (Vahouny et al 1988). Corn endosperm mutants can affect appearance of the kernel or underlying component quality, while double mutants may synergistically affect endosperm appearance and quality. Corn starch properties in the germplasm can be modified by traditional plant breeding methods using major (e.g., naturally occurring mutant genes), or minor (modifying genes) genetic factors (Ji et al 2004) where the effect is enhanced by crossing the lines. Exotic corn lines may provide unusual traits of interest including increased %RS through the presence of modifying genes. High-amylose (amylose-extender, ae) corn lines provide greater amounts of RS2 than normal corn through a major (mutant) gene. Thus, crossing ae and exotic corn types could increase the RS, provide unique materials for food use, and possibly provide cooking properties better than ae corn lines used alone. Whereas four types of RS have been defined, only RS2, RS3, and RS4 are routinely measured. The process of extraction may alter RS1 because it destroys the surrounding food matrix. There are several options available for measuring RS2, RS3, and RS4. The Megazyme RS kit measures RS2 effectively using Approved Method 32-40 (AACC International 2010) and is designed to screen large numbers of samples (McCleary and Monaghan 2002). However, most starch is not eaten in an ungelatinized form; the starch is generally cooked. Thus, the Megazyme kit, which includes no gelatinization step, may not be an accurate measure of RS as eaten. Differential scanning calorimetry (DSC) measures starch gelatinization characteristics, including retrogradation, the cause of RS3. Thus, starches with a high percentage of retrogradation should have high RS3 percentages (Haralampu 2000). The starch properties from many corn mutants, including sugary2 (su2), amylose extender (ae), and amylose dull (ae du), were examined by DSC (Tziotis et al 2005). Less work has been done on double mutants, and especially on other mutants that vary in RS. An ae starch might provide a large amount of both RS2 and RS3. The ae starches do not completely gelatinize under boiling temperatures (Champ 1992), leaving some of the RS2 intact; the starch that does gelatinize would be available for retrogradation and, thus, a source of RS3. Thermal characterization of starches from corn lines with elevated levels of RS has not been done, especially in relation to RS2 measured with the Megazyme kit and RS3 measured with DSC. These evaluations would be useful in predicting behavior of the corn starches in food products. The objectives of this study were to identify new corn breeding crosses containing high %RS2 by crossing four mutants and six lines with introgressed exotic backgrounds with each other and to relate the percentage of RS2 in the starches measured by using the Megazyme Resistant Starch kit (RS2), with the %RS3, and other thermal characteristics measured on DSC. MATERIALS AND METHODS Corn Materials Ten corn lines (Zea mays L.), four mutants and six lines with introgressed exotic backgrounds (Table I), termed ‘parents’, were crossed with each other to create 20 progeny crosses (Table II). The GUAT ae parent is the first public 70% amylose line (Campbell et al 2007). For easier discussion and comparison, the progeny crosses are separated into four groups and identified by the first parent, creating the mutant groupings of dull sugary2 (du su2), amylose-extender sugary2(ae su2), amylose-extender dull (ae du), and an amylose-extender with introgressed Guatemalan germplasm (GUAT ae). All progeny crosses were grown at Juana Diaz, Puerto Rico, in 2006 and 2007 and were the second-selfed generation in the process of developing inbred lines. Ears were harvested at full maturity, and dried at 37.5°C to ≈12% moisture. Seeds were stored at 4°C and 10% rh until needed for starch extraction. Seeds from individual ears were pooled and 15 kernels were randomly selected for each starch extraction. Commercial cornstarch (Sigma, St. Louis, MO) was used as a typical corn starch control. A high-amylose control (High Am-C, amylogel 03001, 50% apparent amylose, Cargill, Cedar Rapids, IA) was used as an ae control in the analyses. 1 Graduate student and professor, respectively, E262 Lagomarcino, College of Human Sciences and Department of Food Science and Human Nutrition, Iowa State University, Ames, IA 50011. 2 Research geneticist, USDA-ARS, Corn Insects and Crop Genetics Research Unit, Department of Agronomy, Iowa State University, Ames, IA 50011. 3 Corresponding author. 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