The formation of a colored charge-transfer (CT) complex between atorvastatin calcium (ATR-Ca) as a n-electron donor and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) as a π-electron acceptor was investigated, for the first time. The spectral characteristics of the CT complex have been described, and the reaction mechanism has been proved by computational molecular modeling. The reaction was employed in the development of a novel microwell-based spectrophotometric assay for determination of ATR-Ca in its pharmaceutical formulations. The proposed assay was carried out in 96-microwell plates. The absorbance of the colored-CT complex was measured at 460 nm by microwellplate absorbance reader. The optimum conditions of the reaction and the analytical procedures of the assay were established. Under the optimum conditions, linear relationship with good correlation coefficient (0.9995) was found between the absorbance and the concentration of ATR-Ca in the range of 10-150 μg/well. The limits of detection and quantitation were 5.3 and 15.8 μg/well, respectively. No interference was observed from the additives that are present in the pharmaceutical formulation or from the drugs that are co-formulated with ATR-Ca in its combined formulations. The assay was successfully applied to the analysis of ATR-Ca in its pharmaceutical dosage forms with good accuracy and precision. The assay described herein has great practical value in the routine analysis of ATR-Ca in quality control laboratories, as it has high throughput property, consumes minimum volume of organic solvent thus it offers the reduction in the exposures of the analysts to the toxic effects of organic solvents, and reduction in the analysis cost by 50-fold. Although the proposed assay was validated for ATR-Ca, however, the same methodology could be used for any electron-donating analyte for which a CT reaction can be performed.