Content overflows the browser. Try zooming out (Ctrl + −) to see the full page. PDF Download
User Benefits Application News High-Speed and High Sensitivity Analysis of Active Ingredients in Mouthwash Natsuki Iwata High Performance Liquid Chromatograph Nexera™ XR Fig. 1 Chromatograms of Mixed Standard Solution (5 mg/L each) Repeatability Table 2 shows the repeatability (%RSD) of the retention time and the peak area in six repeated analyses of a mixed standard solution (0.5 mg/L each). The repeatability of the retention time and the peak area was good for all compounds. 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min 0.0 2.5 5.0 7.5 10.0 12.5 15.0 mAU 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min 0.0 2.5 5.0 7.5 10.0 12.5 15.0 mAU 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min 0.0 2.5 5.0 7.5 10.0 12.5 mAU 1 3 2 Peak 3. IPMP Peak 2. CPC Peak 1. GK2 250 nm 258 nm 280 nm Table 1 Analytical Conditions System: Nexera XR Column: Shim-pack Arata C18 *1 (75 mm × 3.0 mm I.D., 2.2 µm) Flowrate: 0.8 mL/min Mobile Phase: A) 20 mmol/L (Sodium) phosphate buffer (pH 2.6) B) Methanol Time Program: 60 % B (0 min)→80 % B (4.00-4.50 min) →60 % B (4.51-7.00 min) Mixer: 180 µL Column Temp.: 45 °C Injection Volume: 5 µL Vial: SHIMADZU LabTotal™ for LC 1.5 mL, Glass *2 Detection (PDA): 250 nm (GK2), 258 nm (CPC), 280 nm (IPMP) (STD cell for SPD-M40) *1 P/N: 227-32802-02 *2 P/N: 227-34001-01 Enables high sensitivity analysis of three types of active ingredients within 4 minutes. Basic compounds such as quaternary ammonium salts can be analyzed with good repeatability without peak tailing. The photodiode array (PDA) detector allows chromatograms of any wavelength to be obtained with a single analysis. Introduction Commercial oral care products (mouthwashes) contain multiple active ingredients, which are generally analyzed by HPLC 1) . When the quaternary ammonium salt CPC (cetylpyridinium chloride) is analyzed with a C18 column, interactions between the CPC and residual silanol groups remaining on the surface of the silica gel packing material are known to cause adsorption and peak tailing. Application News No. 01-00116A describes high-speed and simultaneous analysis of active ingredients CPC and GK2 (dipotassium glycyrrhizinate) in mouthwash using a Shim-pack Arata™ C18 column. It was shown that using that column, satisfactory peak shapes were obtained without addiing an ion-pairing reagent or perchlorate to an acidic mobile phase. Furthermore, if high sensitivity analysis is desired, it is effective to use a mobile phase with low UV absorption. This article describes high-speed and high sensitivity analysis of CPC, GK2, and isopropyl methylphenol (IPMP) in mouthwash using a phosphate buffer-based mobile phase. Analysis of Mixed Standard Solution CPC, GK2, and IPMP are added to oral care products due to purported bactericidal and anti-inflammatory effects. Fig. 1 shows the chromatograms from a mixed standard solution of CPC, GK2, and IPMP (5 mg/L of each, prepared with 50 % methanol solution). Table 1 shows the analytical conditions. With a typical C18 column, adsorption of basic compounds can be suppressed by using an acidic mobile phase that contains an ion-pairing reagent or perchlorate. However, with a Shim-pack Arata C18 column, the addition of ion-pairing reagents to the mobile phase was unnecessary and the three target compounds could be analyzed quickly.
Application News www.shimadzu.com/an/ Shimadzu Corporation © Shimadzu Corporation, 2023 For Research Use Only. Not for use in diagnostic procedures. This publication may contain references to products that are not available in your country. Please contact us to check the availability of these products in your country. The content of this publication shall not be reproduced, altered or sold for any commercial purpose without the written approval of Shimadzu. See http://www.shimadzu.com/about/trademarks/index.html for details. Third party trademarks and trade names may be used in this publication to refer to either the entities or their products/services, whether or not they are used with trademark symbol “TM” or “”. Shimadzu disclaims any proprietary interest in trademarks and trade names other than its own. The information contained herein is provided to you "as is" without warranty of any kind including without limitation warranties as to its accuracy or completeness. Shimadzu does not assume any responsibility or liability for any damage, whether direct or indirect, relating to the use of this publication. This publication is based upon the information available to Shimadzu on or before the date of publication, and subject to change without notice. First Edition: May 2023 01-00505-EN Fig. 2 Calibration Curves Calibration Curves Fig. 2 shows the calibration curves of the three target compounds for concentrations from 0.1 to 10 mg/L each. Excellent linearities were obtained for all compounds, with coefficients of determination (r 2 ) of 0.999 or greater. In addition, high sensitivity analysis was achieved by using a phosphate buffer as a mobile phase with low UV absorption. Table 3 shows the limit of quantitation calculated from the lowest calibration curve concentrations for the three target compounds (calculation method: ASTM). Conc. Conc. Table 3 Limit of Quantitation Fig. 3 Chromatograms of Mouthwash (Solid line: Sample, Broken line: Standard solution) Table 2 Repeatability (%RSD) in Six Repeated Analyses 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min 0.0 2.5 5.0 7.5 10.0 12.5 mAU 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min 0.0 2.5 5.0 7.5 10.0 12.5 15.0 mAU 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min 0 1 2 3 4 5 6 7 mAU 1 3 2 Peak 3. IPMP Peak 2. CPC Peak 1. GK2 250 nm 258 nm 280 nm Compound Concentration (mg/L) %RSD GK2 1.65 0.27 CPC 4.25 0.24 IPMP 2.92 0.15 (mg/L) CPC GK2 0.0 5.0 0.0 0.5 1.0 1.5 2.0 2.5 3.0 (×10,000) 0.0 5.0 0.0 1.0 2.0 3.0 4.0 (×10,000) 0.0 5.0 0.0 1.0 2.0 3.0 4.0 (×10,000) (mg/L) (mg/L) IPMP Compound Retention time Peak area GK2 0.11 1.01 CPC 0.12 0.48 IPMP 0.12 0.64 Compound LOQ (mg/L) GK2 0.079 CPC 0.076 IPMP 0.053 Peak area Peak area Peak area Conc. Conclusion A high-speed and high sensitivity method for analyzing CPC, GK2, and IPMP, which are active ingredients in mouthwash, was developed. In combination with a Shim-pack Arata C18 column and a phosphate buffer as the mobile phase, tailing of the peaks of basic compounds was suppressed, enabling the quantification of trace compounds in a short period of time. Analysis of Mouthwash A commercial mouthwash was used as the sample. Analytes were prepared by diluting the mouthwash 100-fold with 50 % methanol solution and filtration with a 0.2 µm membrane filter. Fig. 3 shows the chromatograms of the mouthwash, and Table 4 shows the concentrations of the three compounds in the mouthwash. Note that these concentrations shown in Fig. 4 are average concentrations obtained from six repeated analyses of samples after pretreatment. Table 4 Analysis Results (n=6) Nexera, Shim-pack Arata and SHIMADZU LabTotal are trademarks of Shimadzu Corporation or its affiliated companies in Japan and/or other countries. References 1)U. S. Pharmacopeia 41-NF 36, 2017, Official Monographs “Cetylpyridinium Chloride” Related Applications 1. High-Speed and Simultaneous Analysis of Active Ingredients CPC and GK2 in Mouthwash, Application News No. 01-00116A
Related Products Some products may be updated to newer models. Nexera series Ultra High Performance Liquid Chromatograph Shim-pack Arata LC Columns HPLC Column Related Solutions Cosmetics & Personal Care Products Safe and Secure Price Inquiry Product Inquiry Technical Service / Support Inquiry Other Inquiry Powered by TCPDF (www.tcpdf.org)