Title: High-throughput micro-plate HCL-vanillin assay for screening tannin content in sorghum grain Authors
Submitted to: Journal of the Science of Food and Agriculture
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 17, 2013
Publication Date: January 1, 2014
Citation: Herald, T.J., Gadgil, P., Perumal, R., Bean, S. and Wilson, J.D. 2014. High-throughput micro-plate HCL-vanillin assay for screening tannin content in sorghum grain. Journal of the Science of Food and Agriculture. 94:2133-2136. DOI: 10.1002/jsfa.6538. Interpretive Summary: Sorghum that contains tannin is reported to offer health promoting potential above those offered by many fruits such as blueberries. Currently, the test used to determine the amount of tannin present in sorghum is time consuming and not efficient for screening large sample sets. The objective of this research was to develop a rapid and accurate method for determining tannin in sorghum grain. The new test method was validated against a currently reliable method on 25 suspected tannin sorghum lines. The method was approximately eight times faster, as accurate and used fewer supplies than the current method.
Technical Abstract: Sorghum contains tannin which is a phenolic compound that offers health promoting antioxidant capacity. The HCl-vanillin assay is a common and time consuming method for determining tannin content, but is not efficient for screening large sample sets as seen in association mapping panels or breeding nurseries. The objective was to develop a sensitive and reproducible high-throughput 96-well plate micro-titer platform assay for use as a diagnostic tool for rapid screening of large samples sets of sorghum for tannin content. A known tannin containing sorghum, Sumac, was selected to show proof of concept of the high throughput assay and a larger sample set of 25 sorghum containing tannin were used to validate the assay. As a point of reference a HCl-vanillin assay and bleach test were included in the study. The high-throughput 96-well platform was more rapid than the conventional assay. Approximately 30 measurements per day were completed using the HCl-vanillin conventional assay compared to 224 measurements using the high-throughput 96-well platform. The 96-well platform correlated with the conventional assay. The Coefficient of Variation (CV %) was 3.54% and 3.21% for the high-throughput 96-well platform and HCl-vanillin method, respectively. The 96-well assay exhibited good reproducibility with the inter plate CV% between 2.77%-4.85%. The high-throughput 96-well platform method proved to be as robust and reproducible as the conventional method for determining tannin content in sorghum grain. The high-throughput micro-titer platform assay is usable for routine screening of a large number of samples.