Analysis of ascorbic acid, citric acid and benzoic acid in orange juice

Significance of the topic

The determination of antioxidants and preservatives in fruit juices is essential for ensuring product safety, regulatory compliance, and shelf-life stability. Ascorbic acid (vitamin C) contributes both nutritional value and antioxidant protection, while citric and benzoic acids act as preservatives to inhibit microbial growth. Accurate quantification of these compounds supports quality control in the food industry and addresses health concerns such as potential benzene formation from benzoic and ascorbic acid interactions.

Objectives and Study Overview

This study aimed to develop and validate a reliable liquid chromatography method for the simultaneous quantification of ascorbic acid, citric acid, and benzoic acid in orange juice. It compared a conventional HPLC approach with a rapid UHPLC alternative to demonstrate improvements in analysis time without compromising sensitivity or accuracy.

Methodology and Instrumentation

• Sample preparation: Simple extraction from orange juice samples with common solvents.
• Chromatographic conditions: Initial method on Agilent 1260 Infinity LC using a Poroshell EC-C18 column, followed by transfer to Agilent 1290 Infinity UHPLC for faster separations.
• Detection: UV absorbance monitoring, optimized wavelengths for each analyte.
• Validation: Recovery, linearity, precision, limit of detection (LOD), and limit of quantification (LOQ) assessed across relevant concentration ranges.

Main Results and Discussion

• Recovery: Exceeded 90% for all analytes, indicating efficient extraction and minimal matrix interference.
• Linearity: Calibration curves demonstrated strong linear response over typical regulatory concentration ranges.
• Sensitivity: Both HPLC and UHPLC methods achieved comparable LODs, with benzoic acid detection unchanged despite five-fold faster UHPLC runtime.
• Throughput: UHPLC reduced analysis time by nearly 80%, enhancing laboratory productivity.

Benefits and Practical Applications

• Versatility: Applicable to routine quality control in juice manufacturing.
• Efficiency: Short runtime and simple sample prep minimize resource consumption.
• Regulatory compliance: Meets requirements for monitoring preservative levels and nutritional content.

Future Trends and Opportunities

Advances may include coupling with mass spectrometry for confirmatory analysis, utilization of green solvents and micro-scale extraction, integration into automated high-throughput platforms, and development of online monitoring tools for real-time quality assurance.

Conclusion

The presented HPLC and UHPLC methods offer robust, accurate, and rapid quantification of key additives in orange juice. Transitioning to UHPLC notably enhances throughput while preserving analytical performance, supporting efficient quality control in the food industry.

Used Instrumentation

• Agilent 1260 Infinity Binary LC with diode array detector
• Agilent 1290 Infinity UHPLC system
• ZORBAX Poroshell 120 EC-C18 column (2.7 μm, 4.6×100 mm)