Determining Corn Germ and Pericarp Residual Starch by Acid Hydrolysis

Corn (Zea mays) is the primary feedstock for fuel ethanol production in the United States. In 2007, ≈58 million metric tons were processed by the dry-grind industry to produce ethanol and distillers dried grains with solubles (DDGS), a coproduct used as animal diet ingredient. A limited market depressed the price of DDGS, which could be eased if coproducts were diversified (Rausch and Belyea 2006). Fractionation processes (modified dry-grind processes, as they are referred to when adjunct to ethanol production) have been developed to produce higher value coproducts such as corn germ and fiber (Singh et al 2005). Removing germ and fiber could increase fermentation throughput by concentrating starch in the fermentation mash (Singh et al 2005). However, ethanol yields typically are reduced due to starch lost to germ and fiber fractions during fractionation. Maximizing starch recovery from these fractions is important for process economics. A common way to measure the effectiveness of fractionation is by determining the residual starch in fractionated germ and fiber. The enzymatic method is the established way of measuring total starch in cereal products (McCleary et al 1994). The procedure consisted of incubating finely milled samples with thermostable α-amylase at high temperatures to liquefy granular starch before digestion with glucoamylase and quantifying glucose by colorimetric assay (e.g., glucose oxidase-peroxidase-chromogen assay). As a tool for measuring residual starch in samples that have gone through enzymatic processes (e.g., starch hydrolysis and fermentation), a method using another enzyme (thermostable α-amylase) does not appear ideal. For example, such a method would be unlikely to be capable of hydrolyzing resistant starch (McCleary et al 2002), which, expectedly, could grow in proportion to the total residual starch as the extent of enzyme reaction increases. To a lesser degree, methods relying on acid hydrolysis have been used to determine total starch in plant tissues (Pirt and Whelan 1951; Ebell 1969). One potential disadvantage of assays based on acid hydrolysis is overestimation of starch content, apparently due to breakdown of nonstarch carbohydrates (Rose et al 1991; Chow and Landhausser 2004). In contrast, McCleary et al (1994) reported that acid hydrolysis gave comparable and sometimes lower starch concentrations in whole cereal products when compared with the enzymatic method. The applicability of acid hydrolysis to residual starch analysis in fractionated grain components has not been the subject of any published study. It would be useful to examine whether such a method could accurately estimate starch contents in these samples (i.e., corn germ and fiber). The use of an assay based on acid hydrolysis to measure starch in corn germ and pericarp fiber was evaluated. The assay, referred to as HCl assay, was adopted with modification from a procedure developed by Ebell (1969), using a more dilute HCl concentration and higher temperature. The use of more dilute HCl and higher temperature (thus allowing for shorter incubation time) could minimize nonstarch degradation. The results of this assay on fractionated germ and fiber samples were compared with the enzymatic method (Approved Method 76-13, AACC International 2000) using the Total Starch Kit (Megazyme, Bray, Co. Wicklow, Ireland), which shall be referred to as MZYM assay.