R-levocetirizine, the racemic switch of (±)-cetirizine, is a potent H1-receptor antagonist. R-levocetirizine is currently marketed as oral dosage forms used to prevent and treat allergic conditions. In the present study, a stereoselective HPLC method was developed and optimized for the determination of cetirizine enantiomers in bulk and formulations by employing a chemometric tool. Chromatographic separation was carried out on a Chiralpak AS-3R analytical column (150 × 4.6 mm i.d., 3 µm). D-optimal mixture design was employed to study the effect of solvent composition, viz. acetonitrile (75–85%, v/v), MeOH (5–15%, v/v) and water (5–15%, v/v), by keeping other factors constant, such as mobile phase additives, flow rate (1.0 mL/min) and wavelength (235 nm) on output responses: retention factor and resolution of S- and R-enantiomers and analysis time. The mobile phase system comprising of acetonitrile−methanol−water−acetic acid−diethylamine (85 : 10 : 5 : 0.1 : 0.1, v/v) was selected as optimal by a graphical optimization trade-off technique. The proposed method was validated according to the ICH guidelines and successfully applied for the quantitation of cetirizine enantiomers in pharmaceutical dosage forms. The present study resulted in a fast and efficient liquid chromatographic method for the enantiopurity assessment of R-levocetirizine in real samples.