Analysis of Polyphenols in Green Tea using the LCMS-IT-TOF

Significance of the Topic

Green tea contains a variety of polyphenolic compounds, notably catechins, that exhibit strong antioxidant properties and potential health benefits such as blood pressure regulation, anti-inflammatory effects, and chemopreventive activity. As consumer and industry interest in nutraceuticals grows, reliable analytical methods for characterizing these bioactive molecules are essential.

Goals and Study Overview

This study aimed to develop and demonstrate a workflow combining liquid chromatography and ion trap–time of flight mass spectrometry (LCMS-IT-TOF) to identify and characterize major polyphenols in green tea. The objectives included efficient sample cleanup, chromatographic separation, structural elucidation via MSn, and accurate mass determination to support formula prediction.

Methodology and Instrumentation

Sample Preparation involved brewing one commercial tea bag in 250 mL water, dividing the cooled infusion across six C18 SPE columns, washing, and eluting polyphenols with an acetone–water–acetic acid solvent. Pooled eluates were centrifuged and diluted 1:10 for analysis.

Used Instrumentation

The green tea extract was analyzed on a Shimadzu LCMS-IT-TOF system under the following conditions:

• Column: Shim-pack VP-ODS (2.0 × 150 mm)
• Mobile phases: 0.5% acetic acid in water (A) and acetonitrile (B)
• Gradient: 5–12% B (0–5 min), 12–40% B (5–30 min), 40–70% B (30–35 min), hold at 70% B to 36 min, return to 5% B by 38 min, re-equilibrate to 42 min
• Flow rate: 0.2 mL/min, injection volume: 2 µL
• ESI probe voltage: 4.50 kV; temperatures: 200 °C (CDL and block heater); nebulizing gas: 1.5 L/min
• MS scan ranges: MS1 100–1500 m/z, MS2 50–1100 m/z (CID 35%), MS3 50–800 m/z (CID 75%)

Main Results and Discussion

Chromatographic separation produced distinct peaks for catechins and related glycosides, with total ion chromatogram peaks corresponding to expected m/z values. MSn fragmentation confirmed structural features of quercetin glycosides, and Composition Formula Predictor ranked the correct catechin formulas at highest confidence. Observed mass accuracies for major polyphenols were all within 5 ppm, demonstrating reliable mass assignments.

Advantages and Practical Applications

The LCMS-IT-TOF platform offers integrated MSn capability and high mass accuracy, enabling confident identification of polyphenols in complex botanical extracts. This approach benefits quality control in nutraceutical production and supports research into health-promoting compounds.

Future Trends and Applications

Future developments may include high-throughput screening of diverse plant extracts, incorporation of ion mobility separation for isomer differentiation, and data-driven workflows using predictive analytics to accelerate compound discovery.

Conclusion

Combining solid-phase extraction, robust LC separation, MSn fragmentation, and accurate time-of-flight mass measurements provides a comprehensive method for green tea polyphenol analysis. The workflow ensures precise formula assignment and structural confirmation, meeting the needs of both research and industry quality assurance.

References

• Shimadzu Scientific Instruments. Analysis of Polyphenols in Green Tea using the LCMS-IT-TOF Application Note.