A simplified technique to detect variations of in leaf chlorogenic acid levels between within plants caused by maturation or biological stress

Authors: Baker, Con; Mock, Norton; AVERYANOV, ANDREY - Russian Institute Of Phytopathology

Submitted to: Physiological and Molecular Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/28/2018
Publication Date: 5/31/2018
Citation: Baker, C.J., Mock, N.M., Averyanov, A.A. 2018. A simplified technique to detect variations of in leaf chlorogenic acid levels between within plants caused by maturation or biological stress. Physiological and Molecular Plant Pathology. 103:114-121. 10.1016/j.pmpp.2018.05.007.
DOI: https://doi.org/10.1016/j.pmpp.2018.05.007

Interpretive Summary: Bacterial plant diseases cause major damage to crops each year and the cost of controlling them adds greatly to production costs and often involves antibiotics which are a public concern. The plant leaf apoplast, which is the cell wall region just outside the plant cell itself, is the first line of defense against most aerial pathogens. In this manuscript we are reporting a simplified but very sensitive procedure to detect chlorogenic acid leakage from the cells which identifies the time point when first damage to the plant cell wall occurs in pathogenic interactions. This has allowed us to distinguish between chemical events in the apoplast that relate to early recognition responses or to post recognition responses. This information defines the time by which control measures need to be engaged, and will benefit plant scientists and breeders who are devising new strategies to improve disease resistance in plants as well as decrease the use of chemical pesticides and antibiotics.

Technical Abstract: Most methods used for the quantification of chlorogenic acids (CGA) in plant tissue use a multi-step organic solvent extraction, which improves accuracy but is laborious and time-consuming. We needed a simpler technique that would allow us to compare the effects of various bacterial treatments on leaf tissue CGA levels over time. Here we describe such a technique that was rapid, used less tissue, and was highly reproducible. The final technique involved grinding a small amounts of leaf tissue in acidified water, which helped stabilize the phenolics, and after clarification by centrifugation could directly be analyzed by Ultra High Performance Liquid Chromatography with Ultraviolet and Mass spectrophotometer detectors (UHPLC-UV-MS). Using this method, we found that CGA levels can vary greatly with leaf age, location on the leaf, and between plants. Therefore in order to observe the more subtle changes caused by bacterial treatments, it was essential to first be aware of the CGA variations in plants to find the most homogenous tissue to use. Ultimately, we found that leaf halves were most homogenous and adjacent panels on a same side of the midrib could be used for multiple sampling of a treatment over time. During this study we also found some interesting relationships regarding CGA distribution in the plant.