HPLC method validation and application for organic acid analysis in wine after solid-phase extraction

Violeta Ivanova-Petropulos, Krste Tašev, Marina Stefova


A solid-phase extraction method followed by reverse phase high-performance liquid chromatography (RP-HPLC) was optimized and validated for the quantitative determination of tartaric, malic, shikimic, lactic, citric and succinic acids in wine. Solid-phase extraction was carried out with C18 cartridges and extraction recoveries for all acids ranging from 98.3 to 103% were obtained. HPLC separation was performed with isocratic elution on a LiChrosorb RP-18 column (250 × 4.6 mm I.D., 5 µm) protected with the appropriate guard column. The mobile phase was a 5 mM solution of H3PO4 with pH 2.1 at a flow rate of 1 ml/min. Detection of the organic acids was performed at 210 nm. The developed method was validated by checking its linearity, limit of detection (LOD), limit of quantification (LOQ), precision and recovery. The method was applied to the analysis of organic acids in Macedonian red and white wines.


wine; organic acids; separation; HPLC method; validation

Full Text:



J. H. Swiegers, P. J. Chambers, I. S. Pretorius, Olfaction and taste: Human perception, physiology and genetics. Australian Journal of Grape and Wine Research, 11, 109–113 (2005).

P. Ribґereau-Gayon, Y. Glories, A. Maujean, D. Dubourdieu, Handbook of Enology, The Chemistry of Wine and Stabilization and Treatments, Volume 2. John Wiley & Sons, Ltd. (2006).

T. Henick-Kling, Control of malolactic fermentations in wine: energetics, flavor modification and methods of starter culture preparation. Journal of Applied Bacteriol-ogy, 79, 29–37 (1995).

C. Tessini, C. Mardones, L. Rivas, D. von Baer, Measurement uncertainty of shikimic acid in red wines produced in Chile. Accreditation and Quality Assurance, 14, 381–387 (2009).

C. Mardones, A. Hitschfeld, A Contreras, K. Lepe, L. Gutiérrez, D. von Baer, Comparison of shikimic acid de-termination by capillary zone electrophoresis with direct and indirect detection with liquid chromatography for varietal differentiation of red wines. Journal of Chroma-tography A, 285–292 (2005).

A. Schneider, V. Gerbi, M. Redoglia, A Rapid HPLC Method for Separation and Determination of Major Organic Acids in Grape Musts and Wines. American Journal of Enology and Viticulture, 38, 151–155 (1987).

D. Tusseau, C. Benoit, Routine high-performance liquid chromatographic determination of carboxylic acids in wines and Champagne. Journal of Chromatography, 395, 323–333 (1987).

E. Falque-Lopez, E. Fernández-Gómez, Simultaneous determination of the major organic acids, sugars, glycerol, and ethanol by HPLC in grape musts and white wines. Journal of Chromatographic Science, 35, 254–257 (1996).

M. Kordiš-Krpež, V. Abram, M. Kač, S. Ferjančič, Determination of Organic Acids in White Wines by RP-HPLC. Food Technology and Biotechnology, 39, 93–99 (2001).

I. Mato, S. Suarez-Luque,J. Huidobro, Review of the analytical methods to determine organic acid in grape juices and wines. Food Research International, 38, 1175–1188 (2005).

A. Hakan Aktas, S. Sen, M. Yilmazer, E. Cubuk, Determination of Carboxylic Acids in Apple Juice by RP HPLC, Iranian Journal of Chemistry and Chemical Engineering, 24, 1–6 (2005).

C. R. Davis, D. Wibowo, G. H. Fleet, T. H. Lee, Properties of wine lactic acid bacteria: their potential oenological significance. American Journal of Enology and Viticulture, 39, 137–142 (1988).

Official Gazette of Republic of Macedonia, No. 50, 2010.

A. Zotu, Z. Lokou, O. Karava, Method Development for the Determination of Seven Organic Acids in Wines by Reversed-Phase High Performance Liquid Chromatog-raphy. Chromatographia, 60, 39–44 (2004).

V. I. Esteves, S. S. F. Lima, D. L. D. Lima, A. C. Duarte, Using capillary electrophoresis for the determinationof organic acids in Port wine. Analytica Chimica Acta, 513, 163–167 (2004).

R. G. Peres, E. P. Moraes, G. A. Micke, F. G. Tonin, M. F. M. Tavares, D. B. Rodriguez-Amaya, Rapid method for the determination of organic acids in wine by capillary electrophoresis with indirect UV detection, Food Control, 20, 548–552 (2009).

DOI: http://dx.doi.org/10.20450/mjcce.2016.1073


  • There are currently no refbacks.

Copyright (c) 2016 Violeta Ivanova-Petropulos, Krste Tašev, Marina Stefova

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.