Study on the racemization of synephrine by off-column chiral high-performance liquid chromatography

In this study, the racemization kinetic parameters of R-(-)-synephrine, the active phenethylamine alkaloid of Citrus aurantium L., were determined by means of an off-column HPLC method. Enantioseparation was carried out in different buffer solutions and solvents on a chiral stationary phase (CSP) with cellobiohydrolase as the chiral selector (Chiral-CBH, 100 mm x 4.0 mm id., 5 mu m). The mobile phase was 10 mM sodium phosphate buffer (pH 6.0)-2-propanol (95:5, w/w), with 50 mu M disodium EDTA, at 0.8 mL/min. The column was thermostatted at 20 degrees C and detection was set at 225 nm. The influence of pH value, ionic strength, temperature and addition of organic modifier on the rate constant, the half-life of racemization and the free energy barrier of racemization of R-(-)-synephrine were determined. Among the different chemical and physical parameters evaluated as affecting the racemization of naturally occurring R-(-)-synephrine, pH, temperature and addition of an organic co-solvent appear to have the strongest effect, while ionic strength does not exert a significant influence on the racemization rate. The results of the present study indicated that synephrine racemization is possible at high temperature at both acidic and basic pH values: therefore, the extraction procedure of R-(-)-synephrine from the plant material should be carried out under specific conditions to preserve the stereochemical integrity and the biological activity of this secondary metabolite. (C) 2010 Elsevier B.V. All rights reserved.