High-Speed and Simultaneous Analysis of Active Ingredients CPC and GK2 in Mouthwash

Significance of the Topic

Oral care products such as mouthwashes contain multiple active ingredients that require reliable, rapid analysis to ensure product quality and consumer safety. Quaternary ammonium compounds like cetylpyridinium chloride (CPC) often exhibit peak tailing in reversed-phase HPLC due to interactions with residual silanol groups. Developing a high-speed method that avoids ion-pairing reagents or perchlorate salts can streamline routine quality control and minimize solvent use.

Objectives and Study Overview

This study aims to establish a rapid, simultaneous HPLC method for quantifying CPC and dipotassium glycyrrhizinate (GK2) in mouthwash formulations. Key goals include:

• Completing separation within four minutes
• Avoiding ion-pairing reagents and perchlorate salts
• Achieving sharp peak shapes and high repeatability

Methodology and Instrumentation

A Shimadzu Nexera XR system equipped with a Shim-pack Arata C18 column (75 × 3.0 mm I.D., 2.2 µm) was used. The mobile phase consisted of 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B). Gradient elution was programmed from 40% B at 0 min to 50% B at 4 min, then returned to 40% B by 4.5 min. Flow rate was 1.0 mL/min, column temperature 40 °C, and injection volume 1 µL. Detection was performed at 254 nm using a photodiode array detector. Real samples were prepared by tenfold dilution in ultrapure water followed by 0.2 µm filtration.

Main Results and Discussion

• Both CPC and GK2 were fully resolved within four minutes without peak tailing.
• Retention time and peak area repeatabilities were ≤0.6% RSD for a 10 mg/L standard mix over six injections.
• Calibration curves from 5 to 200 mg/L exhibited excellent linearity (r² ≥ 0.9999).
• Analysis of two commercial mouthwashes yielded CPC at 23.1 and 44.7 mg/L, and GK2 at 143.6 and 22.4 mg/L (post-dilution concentrations).
• UV spectra obtained by the PDA detector confirmed peak identity through matching absorption maxima (CPC at 212/258 nm; GK2 at 250 nm).

Benefits and Practical Applications

• Rapid throughput supports high-volume quality control in oral care manufacturing.
• Eliminating ion-pairing reagents simplifies mobile phase preparation and reduces potential matrix effects.
• The end-capped C18 column effectively suppresses silanol interactions, enhancing peak symmetry for basic analytes.

Future Trends and Potential Applications

• Extending the approach to other quaternary ammonium and basic pharmaceutical compounds.
• Coupling with mass spectrometric detection for enhanced sensitivity and specificity.
• Adapting the method to green solvents and ultra-high-performance LC for further speed and environmental benefits.
• Implementing in automated, multi-analyte screening workflows for consumer products.

Conclusion

A fast, reliable HPLC method for simultaneous determination of CPC and GK2 in mouthwash was developed using a Nexera XR system and Shim-pack Arata C18 column. By omitting ion-pairing reagents and perchlorates, the procedure achieves sharp peaks, high repeatability, and linear calibration over a broad concentration range, making it well suited for routine quality assurance in oral care product analysis.