Submitted to: Journal of Chromatography
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 1, 2003
Publication Date: January 1, 2004
Citation: Robbins, R.J., Bean, S. 2004. Development of a measurement system for phenolic acids: quantitative high-performance liquid chromatography with photodiode array detection.. Journal of Chromatography. A1038:97-105 Interpretive Summary: Purification and measurement system was developed for naturally occurring compounds (called phenolic acids) found in plants based foods. Phenolic acids are linked to healthful benefits. To understand how these acids are healthful, we need to know how much are present in the foods we eat everyday. A measurement system needs to be developed to carry this out. The purification was performed by liquid chromatography (LC) and the detection and measurements were performed by ultraviolet (UV) absorption. Several different purification conditions were tested. The best was finally chosen for our measurements. Measurements were carried out on 3 different wine samples. To test our method, samples were analyzed in our as well as in a collaborating laboratory. The results from both the laboratories show good agreement.
Technical Abstract: A high performance liquid chromatography with photodiode array detection method was developed to determine the levels of phenolic acids expected to be found in commonly consumed foods. Separation of 16 phenolic acids simultaneously was achieved. Six columns (varied stationary phase, particle sizes and column lengths), several solvent systems (CH3CN, CH3OH, H2O containing either acetic, formic or trifluoroacetic acids) were investigated. Resolution was achieved with a high purity silica Phenomenex Luna C18 column (4.6 x 150, 5 m). Mobile phase is a binary gradient consisting of CH3OH-H2O (with 0.1% formic acid), at 25°C, with a flow rate at 0.7 mL/min. Acids were monitored at selected wavelength representing the lowest energy max for individual acids. The conditions of the mobile phase were chosen as to be compatible with mass spectral detection (e.g. acid additive and flow rate). Quantitative studies used the external calibration method. Phenolic acids demonstrated linear behavior over a wide range (0-100mg/L). The correlation coefficients ranged from 0.9991-1. The utility of our measurement system was show by running several wine samples. Initial quantification experiments on red wines yielded values consistent with literature reports. To confirm our quantitative results, identical samples were analyzed both in-house and by a collaborating laboratory. Correlation of data from two laboratories generated linear regression equations that approached unity (0.93- 0.98) and with R2 values ranging from 0.990-0.999.