Simple, Reliable HPLC Analyses of Organic Acids
Using Water-Compatible Allure or Ultra C18 Columns
- Use 100% aqueous mobile phases without losing retention.
- Simple, isocratic method.
- Complete resolution of critical organic acids, including quinic and tartaric.
Organic acids are common components in foods and beverages, and play a critical role in product characteristics like taste and aroma. They can be tested for in many food products including fruits, cheeses, and various beverages such as juices and wines. Organic acids can originate in the foods themselves (e.g. cranberries) or can be produced by food processing (e.g. alcoholic fermentation). A method that allows resolution of organic acids, as well as their quantification, can help determine product quality and authenticity.
Reversed phase HPLC coupled with UV-Vis detection is a popular technique for organic acid analysis because it is easy to use. One common method, AOAC method 986.13, stipulates reversed phase HPLC using two C18 stationary phase columns in series. Because organic acids are low in molecular weight, and have polar functionalities, 100% aqueous buffer is the mobile phase of choice. A low pH buffer is used to ensure that all acidic groups are protonated (otherwise, the organic acids are neutral), thus allowing the best interaction between the organic acids and the C18 stationary phase. However, using a 100% aqueous mobile phase can cause the C18 chain in conventional C18 columns to collapse. Phase collapse results in loss of retention, and the column must be flushed with organic mobile phase, a time consuming step, to restore chain structure and column performance.
Three Restek columns—the Ultra Aqueous C18 column, the AllureAqueous C18 column, and the Allure Organic Acids column—were designed using aqueous-compatible C18 phases that combat phase collapse. The advantage of using these columns is demonstrated in Figure 1 by the fast analysis of organic acids on a Shimadzu Prominence 20A system. Here, we compared the ability of the Ultra Aqueous C18 phase and a conventional C18 phase to withstand phase collapse. The Ultra Aqueous C18 phase resolves organic acids in a 100% aqueous mobile phase without loss of retention. In comparison, the conventional C18 phase suffers a complete loss of retention following phase collapse when used under the same conditions. Thus, in an analysis that requires, or is improved by, a mobile phase with a high aqueous content, an Ultra Aqueous C18 column is the superior choice.
In analyses of organic acids, specifically, under high aqueous mobile phase conditions, the Allure Organic Acids column is the column of choice. We have developed a method using a 300mm Allure Organic Acids column to separate critical organic acids: tartaric, quinic, malic, citric and fumaric acids. This method uses 100% aqueous mobile phase as recommended by AOAC method 986.13. In addition to allowing repeated injections in 100% aqueous mobile phase without the severe lose of retention observed with conventional C18 column technology, the Allure Organic Acids column is prepared and tested specifically for separating organic acids. Figure 2 shows that tartaric and quinic acids are resolved to baseline; Figure 3 shows typical analyses under the conditions we recommend.
Figure 2 Excellent resolution of organic acids, including tartaric and quinic acids, on an Allure Organic Acids column.
Figure 3 Sharp, easily differentiated organic acid profiles for cranberry juice cocktail on an Allure Organic Acids column.