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Superior Resolution and Peak Symmetry for Organic Acids in Fruit Juices
by Rebecca Wittrig, Ph.D., HPLC Product Marketing Manager and Vernon Bartlett, HPLC Innovations Team Manager
Abstract
Organic acids in fruit juices can be separated and quantified by reversed phase HPLC, and the resulting profiles used to identify a juice or verify its purity. Because several of the target acids are extremely difficult to resolve, a 100% aqueous mobile phase is needed to maximize the interactions between the acids and the C18 stationary phase. Novel binding chemistry in an application-specific HPLC column prevents the alkyl chains from collapsing in this aqueous environment, enhancing retention and selectivity. A single 30-cm Allure™ Organic Acids column effectively and reproducibly resolves key organic acids, including tartaric and quinic acids, under these challenging conditions.
Products of the fruit juice industry are worth over $12 billion per year in the US alone.1 As in other arenas in which there is a large potential for profit, unscrupulous producers and traders have found ways to replace or extend more valuable juices. Typically this is done by diluting high value juices with lower value ones (e.g., white grape juice or pear juice), or with sugar water, or by substituting sugars for juice solids. Quality assurance laboratories are charged with detecting these adulterations.
Organic acid profiles vary among different fruit juices, and organic acid composition and concentrations can be used to identify a juice or verify its purity. For example, malic acid is a major component of the organic acid profile of apple juice. If apple juice has been diluted, the malic acid content will be reduced. Similarly, cranberry juice contains quinic, malic, and citric acids, but negligible amounts of tartaric acid. A "cranberry juice" that contains measurable amounts of tartaric acid likely has been diluted with grape juice, which contains relatively high levels of tartaric acid.
Many laboratories involved in authenticity testing follow AOAC method 986.13 in determining the organic acid content of fruit juices.2 In this procedure, reversed phase HPLC with UV detection is used to quantify the organic acids including quinic, malic, citric, and tartaric acids. Because several of the target acids are extremely difficult to resolve, the procedure specifies a 100% aqueous, phosphate-buffered mobile phase, to maximize the interaction between the acids and the C18 stationary phase. C18 chains collapse in a totally aqueous environment, however, and separations and peak shapes are poor. To compensate, the analysis requires two columns in series.
The Allure™ Organic Acids column enhances retention and selectivity in this challenging application.3 Novel binding chemistry enables the alkyl chains to remain extended in a 100% aqueous environment; retention is stable and reproducible. Consequently, a single column effectively resolves key organic acids, such as tartaric and quinic acids, under the conditions specified in AOAC method 986.13. In a separation of typical fruit juice organic acids, tartaric and quinic acids are resolved to baseline and all peaks are sharp and symmetric (Figure 1). This superior chromatography makes interpretation of analytical data more reliable.
In evaluations of fruit juice quality, the unique Allure Organic Acids column helps ensure trouble-free analysis and accurate results.3
References
1. Authenticity of Apple Juice Technical Bulletin #2, Analytical Chemical Services of Columbia, Inc., 1996.
2. Official Methods of Analysis, AOAC International, 17th edition, method #986.13, 2000.
3. Single-Column Method for HPLC Analysis of Organic Acids in Fruit Juices, Using an Allure Organic Acids Column Restek Corporation, Bellefonte, PA, 2003 (lit.cat.# 59530).
Figure 1. Excellent separation and peak symmetry for fruit juice organic acids: Allure Organic Acids column.
Column: Allure Organic Acids, 300 × 4.6 mm, 5 μm particles, 60 Å pores; mobile phase: 100 mM phosphate buffer, pH 2.5; flow: 0.5 mL/min.; temperature: ambient; detection: UV, 226 nm.
Peaks: 1) tartaric acid, 2) quinic acid, 3) malic acid, 4) citric acid, 10 μg each on column; 5) fumaric acid, 0.05 μg on column. LC_0238
Restek Corp.
110 Benner Circle
Bellefonte, PA 16823
Restek Corporation
110 Benner Circle
Bellefonte, PA, 16823
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