Rapid HPLC Method for Determination of Parachloroaniline in Chlorhexidine Antiseptic Agent in Mouthrinses, Ophthalmic and Skin Solution


We described a simple and rapid method to quantify simultaneously chlorhexidine (CHD) and its major metabolite, para Chloroaniline (pCA) by HPLC with UV detection without the additional need of mobile-phase amine modifiers or ion-pairing reagents, with good resolution between pCA and CHD, symmetry peak of the compound and short run time. HPLC-UV analyses were performed using a Dionex® Summit liquid chromatograph (Dionex Corp, Sunnyvale, CA, USA). Chromatographic separations were carried out on a Luna® 150 mm×3 mm i.d. column packed with 3 µm CN (cyano) particles (Phenomenex®), guarded by an on-line filter. Mobile phase consist of methanol:water with sodium chloride with 0.02% of formic acid (55:45). Wavelengths for pCA and for CHD are 238 and 255 nm respectively. Influence of methanol and of sodium chloride content in the eluant has been studied. Linearity of CHD is very good, from 0.5 up to 21.2 µg/l while linearity of pCA is in the range of 0.05 to 10 µg/l with correlation coefficients above 0.999. Resolution between the components is above 4, asymmetry is about 1.3 and 1.7 for pCA and CHD respectively and the run time is less than 5 minutes. This method has been applied to CHD solution of different medical devices. No interference has been reported, and the analysis of direct injection of solution, without any treatment is achieved in less than five minutes.In conclusion, we present a validated method for dosage of CHD and its major impurity pCA, known to be carcinogen, available into medical products or medicinal device for in-vitro diagnostic.

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A. Nicolay, E. Wolff, M. Vergnes, J. Kaloustian and H. Portugal, "Rapid HPLC Method for Determination of Parachloroaniline in Chlorhexidine Antiseptic Agent in Mouthrinses, Ophthalmic and Skin Solution," American Journal of Analytical Chemistry, Vol. 2 No. 4, 2011, pp. 422-428. doi: 10.4236/ajac.2011.24051.

Conflicts of Interest

The authors declare no conflicts of interest.


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