Carotenoids and Chlorophylls in Vegetables Using a Thermo Scientific™ Acclaim™ C30 Column

Significance of the Topic

Carotenoids and chlorophylls are key pigments in plants that drive photosynthesis and protect against photooxidative damage. In human nutrition certain carotenoids serve as provitamin A and antioxidants. Accurate profiling of these compounds in vegetables supports food quality control, nutritional assessment and research into plant physiology.

Study Objectives and Overview

This study demonstrates a high-performance liquid chromatography method for simultaneous separation of six major carotenoids and two chlorophylls in vegetable extracts. The goal is to achieve excellent selectivity and resolution using a Thermo Scientific Acclaim C30 column, enabling reliable quantification in complex plant matrices.

Methodology and Instrumentation

Sample Preparation:

• Extracts of maize, spinach and carrot in acetone

Chromatographic Conditions:

• Column: Acclaim C30, 5 µm, 4.6 × 150 mm
• LC System: Dionex UltiMate 3000 RS
• Mobile phases: A – acetonitrile; B – methanol/ethyl acetate (1:1 v/v); C – 200 mM acetic acid in water
• Gradient: initial 85%A/14.5%B/0.5%C held for 2 min, linear to 65%A/34.5%B/0.5%C by 15 min, held to 25 min
• Flow rate: 1.50 mL/min; Column temperature: 30 °C; Injection volume: 8 µL
• Detector: Diode array at 450 nm, spectral range 260–800 nm

Main Results and Discussion

The protocol achieved baseline separation of eight target analytes with distinct retention order: lutein, zeaxanthin, chlorophyll b, alpha-cryptoxanthin, beta-cryptoxanthin, chlorophyll a, alpha-carotene and beta-carotene. Resolution between critical pairs exceeded industry guidelines, ensuring accurate peak identification in all three vegetable matrices. Diode array spectra enabled confirmation of compound identity based on characteristic absorption maxima.

Practical Benefits and Applications

• Reliable quantification of nutritionally important pigments in foodstuffs for quality control and labeling
• Versatile method applicable to diverse plant matrices including grains, leafy greens and roots
• Robust gradient and detection settings reduce analysis time while maintaining resolution
• Feasible integration into routine laboratory workflows for research, manufacturing and regulatory testing

Future Trends and Potential Applications

Advances in chromatographic media and ultra-high-performance systems may further shorten run times and enhance peak capacity. Coupling C30 separations with mass spectrometry could expand identification to minor carotenoids and degradation products. Emerging applications include monitoring of antioxidant stability in functional foods and precision breeding of crops with tailored pigment profiles.

Conclusion

The presented Acclaim C30 method offers a robust, high-resolution approach for simultaneous analysis of major carotenoids and chlorophylls in vegetables. Its strong selectivity, reproducibility and adaptability make it a valuable tool for nutritional research, food quality assurance and plant science studies.

Reference

Rodriguez-Amaya DB and Kimura M HarvestsPlus Handbook for Carotenoid Analysis International Food Policy Research Institute 2004