Research Project: Grain Composition Traits Related to End-Use Quality and Value of Sorghum

Location: Grain Quality and Structure Research

2022 Annual Report

Objectives
OBJECTIVE 1: Determine and quantify grain components linked to ‘health-promoting’ benefits and commercial quality of sorghum foods and feed. • Subobjective 1.A. Determine the mechanism related to the reduced protein quality of cooked sorghum flour. • Subobjective 1.B. Characterize protease inhibitors in sorghum and their role in modulating digestibility in sorghum flour. • Subobjective 1.C. Evaluate and identify bioactive compounds in sorghum linked to anti-cancer and other health promoting properties. OBJECTIVE 2: Develop and improve methodologies for rapid prediction and measurement of sorghum grain attributes linked to valuable end-use quality traits. • Subobjective 2.A. Utilize UHPLC (ultra-high-performance liquid chromatography) size exclusion for characterizing sorghum polymeric protein complexes related to end-use quality of sorghum. • Subobjective 2.B. Develop near infrared spectroscopic methods to predict grain composition and quality traits of sorghum.

Approach
Sorghum is an important drought tolerant crop in the central U.S. where water is limited and rainfall unpredictable. Sorghum has been primarily used for animal feed in the U.S. and is consistently used by the biofuel industry and increasingly used in human foods. As for any cereal, grain composition plays an important role in its utilization. To support utilization of sorghum grain, research is needed that identifies grain components linked to functional and nutritional quality of sorghum products. One issue for sorghum utilization is how processing, especially cooking, impacts sorghum flour nutritional and functional properties. It is known that heating increases sorghum protein cross-linking, which in turn affects both protein and starch functionality and digestibility. The exact mechanism of how this occurs is not known; nor is it known how protein and starch changes influence the role of digestive inhibitory compounds in sorghum. Identifying the mechanism behind these changes will provide avenues to improve sorghum flour quality as well as provide new targets to improve sorghum grain composition at the genetic level. Likewise, sorghum is known to have high levels of bioactive compounds that have potential human-health promoting benefits. However, much of the past research on bioactive compounds in sorghum has been based on chemical assays. To further define and identify the health-promoting benefits of sorghum, research using additional methods such as cellular based assays are needed. Such research will help define the value of sorghum in human foods and provide targets for the genetic improvement of sorghum.

Progress Report
The overall goal of this project is to identify grain composition traits related to end-use quality and value of sorghum. The project is completing the second year of its current 5-year plan. Progress this year includes completing analysis of data on changes to sorghum proteins when heated which directly relates to Objective 1.A. Related research on protein digestibility has also been completed including development a novel fermented drink using sorghum grain and the impact of fermentation on the digestibility of the grain left after fermentation. Analysis of protein content, composition, and protein digestibility on a second year of samples treated with different levels of nitrogen fertilization has been completed. Samples from sorghum lines that varied in planting date were also analyzed for protein content, composition, and digestibility to determine how planting time impacts grain composition and protein quality. Research has also investigated the use of hydrolyzing sorghum flour with granular starch hydrolyzing enzyme to prepare a high digestible protein for food applications. Preliminary results showed protein digestibility increased by approximately 50% in the residual material left after fermentation. Related to Objective 1.C sorghum brans have been processed and analyzed for an upcoming mouse obesity study. A study evaluating expression of polyphenols in sorghum plant tissue throughout various phenotypes is currently being conducted. Sorghum polyphenols have been evaluated in an inflammation model and have been shown to attenuate signal transducer and activator of transcription 3 (STAT3) signaling. Substantial research related to objective 2.B has been completed again this year with near-infrared spectroscopy calibration curves maintained and improved for protein content, starch content, amylose content, lysine content, crude fat, as well as improvement in discrimination models for presence or absence of tannin in sorghum. Over 4,400 sorghum grain samples were scanned by near infrared spectroscopy this year for the sorghum breeding community, geneticists, seed industry and stakeholders. Calibration curves were also developed for predicting protein content in sorghum grain using a handheld battery powered near infrared spectroscopy instrument. In addition, research this year has been completed on development of a polymerase chain reaction (PCR) method to determine the content of sorghum flour in corn-sorghum flour mixtures. The use of sorghum bran to produce gold coated nano particles was also completed.

Accomplishments
1. Evaluation of handheld near infrared spectroscopy instrumentation for predicting protein content in sorghum grain. Protein content can be an important end-use quality in cereal grains for both animal feed and human foods. Several methods are available to measure protein content in grains including near infrared spectroscopy (NIRS). NIRS is a rapid, high-throughput technique, especially when used to analyze whole grain. However, traditional NIRS instrumentation can be expensive and not suited for use in field environments. Because of these reasons, ARS researchers in Manhattan, Kansas, evaluated the use of a relatively inexpensive, small battery powered handheld micro NIRS instrument for predicting protein content in sorghum grain. A diverse set of 87 sorghum samples was used to generate a calibration model for protein content. The handheld NIRS was able to effectively predict protein content. This compared very closely to the calibration developed with the same set of samples using the larger, more expensive benchtop NIRS instrument. Developing high protein sorghum lines is of increasing interest of stakeholders and the sorghum industry to help meet demand for high protein food products and animal feeds and development of high protein sorghum varieties could help increase the demand for U.S. sorghum. This research demonstrates that the battery powered handheld NIR can be successfully used to predict protein content in whole sorghum grain and could be used in the field saving breeders time and money in identifying samples to harvest for future research and as a less expensive option for breeding programs actively breeding sorghum for increased protein content compared to benchtop NIRS instruments.

2. Use of real-time polymerase chain reaction assay for analysis of corn and sorghum flour mixtures. Some bioethanol plants in the United States use a blend of corn and sorghum grains as feedstocks sources depending on availability and price. Knowledge of the grain composition of milled feedstock is beneficial for maximizing process efficiency, bioethanol marketing and for potential future regulatory requirements. ARS researchers in Manhattan, Kansas, developed an assay for determining the composition of corn-sorghum flour mixtures. The results showed that the developed assay for sorghum had a good performance and could determine the percentage of sorghum in flour mixtures within 2.23%. For corn, the assay did not perform as well and overestimated the amount of corn in the mixtures. The method would be suitable to use to determine the amount of sorghum in flour mixtures for the biofuel industry where ethanol is sold into markets desiring to know the composition of the feedstocks the ethanol was produced from.

3. Development of near infrared spectroscopy calibrations for starch content and composition in sorghum grain. Starch is the major component of all cereal grains and starch content impacts the end-use quality of grains. For sorghum grain, starch content is an important attribute in food, biofuel, and animal feed markets. Starch is made up of two different types, amylose, and amylopectin. The ratio of these two types (often expressed as percent amylose) also greatly influences the functionality of sorghum flour. To rapidly determine the starch and amylose content in sorghum grain, researchers at ARS in Manhattan, Kansas, developed near infrared spectroscopy (NIRS) calibrations for intact sorghum grain. NIRS was able to predict starch content and amylose content. This method allows for approximately 100 grain samples per day to be analyzed for starch content and composition and can be used by the sorghum breeding community to select germplasm with desired starch properties and in genetic studies investigating genes involved in starch development in sorghum. The development of low amylose (i.e. “waxy”) sorghum varieties is of increasing interesting to stakeholders for developing and expanding overseas markets for U.S. sorghum where waxy sorghum is desired and creating new demand in domestic specialty food markets as well as feed and biofuel markets where rapidly digestible starch is desired.

4. Evaluation of sorghum cookies as inclusions in ice cream. One of the most popular ice cream types is cookies-and-cream which includes the addition of traditional wheat-flour cookies into the product. The use of wheat-flour cookies, however, introduces gluten into the ice cream. As an option for a gluten-free product, researchers from ARS in Manhattan, Kansas, and Kansas State University formulated and investigated two types of sorghum-flour cookies (chocolate and non-chocolate) for use as inclusions in ice cream. The sorghum-flour cookies had similar width, spread and thickness as wheat-flour cookies. The sorghum-flour cookies also had similar hardness to wheat-flour cookies when frozen. Chocolate sorghum-flour cookies were found to produce similar properties as chocolate wheat-flour cookies when crumbled, an important aspect for consideration as ice cream inclusions. The results of this study demonstrated that chocolate sorghum-flour cookies could be used as a gluten-free alternative for ice cream inclusions and opens new markets for use of sorghum flour.

5. Identification of quality trait loci for sorghum grain composition traits. Grain composition is the major factor determining the end-use quality and value of cereal grains. To identify genetic regions related to the control of grain composition in sorghum, scientists at ARS in Manhattan, Kansas, and Kansas State University analyzed grain from an inbred sorghum population grown at six environments in Kansas. Multiple genetic regions (quality trait loci) were found related to sorghum grain composition including seven regions related to protein content, 10 regions for starch content and 10 regions for amylose content. Candidate gene analysis indicated that the genetic regions linked to protein and starch content were related to transcription factors that regulate starch and protein accumulation in grain. These results provide the starting point for development of genetic markers for use in breeding programs and accelerating genetic improvement of sorghum grain quality.

Review Publications
Lee, H., Amarakoon, D., Wei, C., Choi, K., Smolensky, D., Lee, S. 2021. Adverse effect of polystyrene microplastics (PS-MPs) on tube formation and viability of human umbilical vein endothelial cells. Food and Chemical Toxicology. 154:112356. https://doi.org/10.1016/j.fct.2021.112356.
Xu, X., Bean, S.R., Wu, X., Shi, Y. 2022. In vitro digestibility of starch in sorghum differing in endosperm hardness and flour particle size. Food Chemistry. 383. Article 132635. https://doi.org/10.1016/j.foodchem.2022.132635.
Shen, Y., Wu, X., Li, Y. 2022. Modulating molecular interactions in pea protein to improve its functional properties. Food Hydrocolloids. 8. Article 100313. https://doi.org/10.1016/j.jafr.2022.100313.
Peiris, K.H., Wu, X., Bean, S.R., Perez-Fajardo, M.A., Hayes, C.M., Yerka, M., Jagadish, K.S., Ostmeyer, T., Aramouni, F.M., Tesso, T., Perumal, R., Rooney, W.L., Kent, M., Bean, B. 2021. Near infrared spectroscopic evaluation of starch properties of breeding populations of grain sorghum. Processes. 9(11). Article 1942. https://doi.org/10.3390/pr9111942.
Li, J., Zhao, R., Xu, Y., Wu, X., Bean, S.R., Wang, D. 2022. Fuel ethanol production from starchy grain and other crops: An overview on feedstocks, affecting factors, and technical advances. Renewable Energy. 188:223-239. https://doi.org/10.1016/j.renene.2022.02.038.
Wu, X., Maghirang, E.B., Armstrong, P.R. 2022. Predicting single kernel moisture and protein content of mushroom popcorn using NIR spectroscopy: Tool for detecting their effect on popping performance. Applied Engineering in Agriculture. 38(3):469-476. https://doi.org/10.13031/aea.14875.
Hein, N.T., Impa, S.M., Wagner, D., Kumar, R., Tiwari, M., Prasad, V., Bheemanahalli, R., Tilley, M., Wu, X., Neilsen, M., Jagadish, K. 2021. Grain micronutrient composition and yield components in field-grown wheat are negatively impacted by high night-time temperature. Field Crops Research. 99(3):615-624. https://doi.org/10.1002/cche.10523.
Cetinkaya, T., Mendes, A.C., Jacobsen, C., Ceylan, Z., Chronakis, I.S., Bean, S.R., Garcia-Moreno, P.J. 2020. Development of kafirin-based nanocapsules by electrospraying for encapsulation of fish oil. LWT - Food Science and Technology. https://doi.org/10.1016/j.lwt.2020.110297.