Rheological Properties of Starch Gels from Wheat Mutants with Reduced Amylose Content Tomoko Sasaki,1,2 Takeshi Yasui,3 Chikako Kiribuchi-Otobe,4 Takashi Yanagisawa,5 Masaya Fujita,6 and Kaoru Kohyama1 ABSTRACT
Cereal Chem. 84(1):102–107
Wheat lines with reduced amylose content were recently produced by single and double mutation from a low-amylose line, Kanto 107. They are appropriate for clarifying the influence of amylose content on starch gel properties because of their similar genetic background. When measured using the concanavalin A method (ConA), the total amylose content of isolated starches from Kanto 107 and three mutants (K107Afpp4, Tanikei A6599-4, K107Wx2) was 24.8, 18.5, 7.1, and 1.7%, respectively. Results of differential scanning calorimetry (DSC) showed that the difference in amylose content strongly affected gelatinization conclusion temperature and enthalpy. We prepared 30 and 40% starch gels and measured their
dynamic shear viscoelasticity using a rheometer with parallel plate geometry. Compressive and creep-recovery tests were conducted under uniaxial compression. The storage shear modulus correlated highly with the amylose content of starch in 30 and 40% starch gels. The creeprecovery test showed a clear distinction in creep curves among starch samples. When the compressive force required for 50, 80, and 95% strains was compared, starch gels with lower amylose content showed lower compressive force at 50% strain. Waxy starch gel (K107Wx2) showed higher compressive force at strain >80% than other samples due to its sticky property.
Starch is the main component of wheat flour. When starch granules are heated in excess water, granules swell and gelatinization takes place, disrupting the molecular order within starch granules. The process induces changes such as native crystallite melting, birefringence loss, and starch solubilization. When cooling a sufficiently concentrated suspension of gelatinized starch, retrogradation occurs involving changes such as starch molecule reassociation and crystallization, which results in gel formation. The rheological properties of starch gel contribute to the texture and acceptability of wheat flour products (Ross et al 1997; Seib 2000; Sasaki et al 2004). Starches consist of amylose and amylopectin. Amylose is an essentially linear molecule and amylopectin is highly branched. The ratio of amylose to amylopectin strongly influences the physical properties of starch (Parovuori et al 1997; Czuchajowska et al 1998; Yuryev et al 1998; Demeke et al 1999; Ortega-Ojeda et al 2004). Amylose and amylopectin play different roles in gel formation. The short-term development of retrogradation in starch gels is attributed to the gelation and crystallization of the solubilized amylose fraction (Miles et al 1985), while the long-term changes in starch granules during storage of starch gels are attributed to the recrystallization of the amylopectin fraction (Abd Karim et al 2000). It has been difficult to clarify the relationship between the amylose content and the rheological properties of wheat starch because the variation of amylose content in native wheat starches is not wide. Wheat lines with low and high amylose content have been developed to determine the effect of amylose content on starch properties such as pasting (Araki et al 2000; Yamamori and Quynh 2000; Yamamori et al 2000; Miura et al 2002; Mangalika et al 2003). However, the relationship has not been clarified between various rheological properties and amylose content using
wheat starches with widely distributed amylose content. Wheat lines with reduced amylose content were recently produced by single or double mutation from a low-amylose line, Kanto 107 (Yasui et al 1997, 2002; Kiribuchi-Otobe et al 1998). The amount of amylose synthesis depends on the expression of three structural genes that encode isoforms of granule-bound starch synthase (GBSSI) (Tsai 1974; Echt and Schwartz 1981). Lines with one or more null alleles have reduced amylose content compared with lines with three functional alleles on three waxy loci. Kanto 107 is considered a low-amylose line, carrying null alleles on Wx-A1 and Wx-B1 loci. K107Wx2 is a waxy line lacking the three waxy proteins Wx-A1, Wx-B1, and Wx-D1. K107Wx2 line was induced by mutation using ethyl methane sulphonate (EMS) treatment of seeds from Kanto 107 and has a Wx-D1d allele (Yasui et al 1997, 1998). Another wheat mutant line induced by EMS treatment from Kanto 107 is K107Afpp4, which contained much lower amylose content than Kanto 107 and showed an altered flour pasting profile (Yasui et al 2002). K107Afpp4 carries null Wx-A1 and WxB1 alleles and has a Wx-D1g allele (Yasui 2004). Tanikei A6599-4 was produced by double mutation from Kanto 107. Tanikei A6599-4 contained a small amount of amylose and showed a stable hot paste viscosity (Kiribuchi-Otobe et al 1998). This line also had null alleles on Wx-A1 and Wx-B1 loci and a mutated allele at the Wx-D1 locus, which was designated Wx-D1e (KiribuchiOtobe et al 2001; Yanagisawa et al 2001). This allelic designation was subsequently changed to Wx-D1f (McIntosh et al 2003). Tanikei A6599-4 is assumed to be a waxy line containing little amylose but not amylose-free because the starch granules of Tanikei A6599-4 stained dark brown, quite different from the purple of the nonwaxy line. These mutant lines from Kanto 107 are appropriate for evaluating the influence of amylose content on starch gel properties because their genetic background is similar. We compared the gelatinization properties and various rheological parameters of starch gels prepared from wheat mutants with reduced amylose content using a range of deformations within the linear viscoelastic region and beyond this region because the differences in rheological properties between wheat lines could differ in scale and type of deformation. Rheological properties of concentrated starch gels (30 and 40%) were determined using a dynamic mechanical test under shear deformation, creep-recovery, and compression tests under uniaxial compression. Because starch concentration is high in wheat products, rheological properties in the concentrated system could provide useful information on food quality.
1 National
Food Research Institute, Kannondai, Tsukuba, Ibaraki 305-8642, Japan. author. Fax: +81-29-838-7996. Phone: +81-29-838-8031. E-mail:
[email protected] 3 National Agricultural Research Center for Western Region, Nishifukatsucho, Fukuyama, Hiroshima 721-8514, Japan. 4 National Agriculture and Food Research Organization, Kannondai, Tsukuba, Ibaraki 305-8517, Japan. 5 National Agricultural Research Center for Western Region, Zentsuji, Kagawa 7658508, Japan. 6 National Institute of Crop Science, Kannondai, Tsukuba, Ibaraki 305-8518, Japan. 2 Corresponding
DOI: 10.1094 / CCHEM-84-1-0102 © 2007 AACC International, Inc.
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CEREAL CHEMISTRY
MATERIALS AND METHODS Samples Four wheat lines (Triticum aestivum L.), Kanto 107, K107Afpp4, Tanikei A6599-4, and K107Wx2, were grown during 2001-2002 at the National Institute of Crop Science. Wheat grains were tempered overnight to 14% moisture on a dry basis, then milled using a test mill (Quadrumat Jr. Brabender, Duisburg, Germany) fitted with a screen of 236-μm openings. Starch was isolated by the dough-ball method of Wolf (1964). Flours were fractionated into prime and tailing starches. After excluding tailing starches, the prime starch was washed with 0.1M NaCl and freeze-dried. Starch recovery from wheat flour was 40.8–49.4%. Amylose Content and α-Amylase Activity Total amylose content of starch was determined by the method of Gibson et al (1997) using an amylopectin-amylose assay kit (Megazyme International Ireland Ltd.). α-Amylase activity in isolated starch was determined by the method of McCleary and Sheehan (1987) using an α-amylase assay kit (Megazyme); one unit of activity was defined as a Ceralpha unit. Differential Scanning Calorimetry Measurement Differential scanning calorimetry (DSC) measurement was conducted using a SSC5200H system with a DSC120 module (Seiko Instruments, Tokyo, Japan) calibrated with indium. Starch (15.0 mg, db) was weighed into a silver pan with 35 μL of distilled water. A pan containing 50 μL of distilled water was used as a reference (Kohyama and Nishinari 1991). After sealing, pans were scanned at 1°C/min from 40 to 120°C. Preparation of Starch Gels Starch suspensions of 30 and 40% (w/w) on a dry basis were prepared. The suspension was stirred continuously at 500 rpm for 30 min using a magnetic stirrer, then heated in a water bath at 55°C for 2.5–4.5 min with continuous stirring at 500 rpm until the suspension became thick enough to prevent starch from settling. The paste was placed between two glass plates with a 1.0 mm spacer, sealed in an airtight bag, and heated at 100°C for 15 min, followed by storage at 5°C for 1 hr. Immediately before measurement, gels were cut from the center of the gel into disks 35 mm in diameter for dynamic viscoelasticity measurement and 20 mm in diameter for creep-recovery and compression measurements. Rheological Meaurement of Starch Gels The dynamic viscoelasticity of gels was measured in a frequency range of 0.01–10 Hz at 25°C and constant stress (50 Pa) using a rheometer (RheoStress RS75, Haake, Germany) with parallel plates (35 mm in diameter, gap 1.0 mm). At this stress level, all samples showed linear viscoelastic behavior in the preliminary stress sweep test. Silicone oil was applied to the exposed surfaces of samples to prevent evaporation during the experiment. A creep meter (Rheoner RE-33005, Yamaden Co., Tokyo, Japan) with a 2 kgf load cell was used for compressive creep-recovery and compression measurements. Creep-recovery measurement was performed at 25°C using a stick-like probe (5 mm diameter). A
starch gel disk (20 mm in diameter) was placed in the center of a base place and uniaxial stress was applied at 10 mm/sec to compress the starch gels and maintained for 1 min. Starch gel was allowed to creep for 1 min. The stress then was suddenly removed and gels were allowed to recover for 1 min. The applied force was determined for a variety of samples in the preliminary stress sweep test. Because all starch gels showed linear viscoelastic behavior at
