Development and validation of a copper ligand-exchange chromatographic method for the estimation of D-lactic acid in Ringer-lactate solution.

In this study, the chromatographic conditions for separation and determination of L- and D-lactic acid enantiomers by copper ligand exchange chromatography have been examined and optimized statistically using a response surface methodology (RSM). The chromatographic variables: copper sulfate, acetic acid and organic modifier were screened by operating a 2-level full factorial design (FFD). The significant effect of independent chromatographic variables was analyzed using the analysis of variance (ANOVA). Variables proved significant (p < 0.05) were cautiously tuned using RSM with a face-centered central composite design. Moreover, a D-optimality design was employed to minimize the variation in the regression coefficients of the fitted model. The proposed model represented an excellent example of fulfilling the efficiency of factorial designs in optimizing the chromatographic conditions and maximizing the output. The chromatographic separation was achieved on Supelco Astec CLC-D chiral bidentate ligand (5.0 µm, 150.0 × 4.6 mm). The isocratic mobile phase composition was 7 mM anhydrous copper sulfate in 1.0 mM acetic acid containing 4% methanol. A photodiode array detector was used to determine the optimal detection wavelength, which was at 236 nm. A linear calibration curve was obtained in the range of 30.0-3600 µg mL-1 with a high value for the coefficient of determination (R2 ≥ 0.999). The optimized method has been successfully applied to the determination of lactate in the commercial Ringer-lactate solution for injection. The results obtained were in excellent agreement with the label claim with no interference from other additives commonly co-formulated with the drug. Intra- and inter-day precision, detection and quantification limits, as well as percent coefficient of variation, have been estimated according to ICH guidelines for assessment of analytical procedures.