Holistically benchmarked optimization-assisted green spectrophotometric–chemometric approach for simultaneous determination of piroxicam and venlafaxine in pharmaceutical formulations and spiked human plasma

The recent clinical introduction of the piroxicam (PIR) and venlafaxine hydrochloride (VEN) combination has highlighted the need for analytical methodologies capable of simultaneously quantifying both drugs while minimizing the environmental burden of routine pharmaceutical analysis. To date, only a chromatographic method has been reported for this purpose, which is associated with relatively high organic solvent consumption, greater instrumental complexity, and increased energy requirements. A green UV spectrophotometric platform integrated with chemometric modeling was therefore established for the concurrent determination of PIR and VEN in pharmaceutical formulations and as a proof-of-concept application to spiked human plasma. Spectral measurements were performed using a water–ethanol (1:1, v/v) solvent system, whereas methanol was employed solely for plasma protein precipitation. Experimental design efficiency was enhanced by combining Brereton’s multilevel calibration design with a Maximin Distance Design (MMD) for validation sample selection, enabling representative calibration and validation datasets to be generated with a reduced number of experimental mixtures. Severe spectral overlap between the two analytes was effectively resolved using six well-established chemometric approaches: CLS, PCR, PLS, GA-PLS, FA-PLS, and MCR-ALS. Among the investigated models, MCR-ALS showed the best overall numerical performance, achieving correlation coefficients of up to 0.9998 for pharmaceutical formulations and 0.9996 for spiked human plasma. The corresponding limits of detection were 0.125 and 0.029 µg mL⁻¹ for VEN and PIR, respectively, in pharmaceutical formulations, and 0.245 and 0.058 µg mL⁻¹ in spiked human plasma. Across the investigated concentration ranges, the proposed method also exhibited satisfactory accuracy and precision. The sustainability performance of the analytical platform was evaluated using multiple complementary metrics, with the pharmaceutical assay compared with the previously reported chromatographic procedure according to the evaluated criteria. The proposed pharmaceutical spectrophotometric assay showed a more favorable sustainability profile according to the evaluated criteria, including environmental and practical applicability dimensions. The application to spiked human plasma provided a proof-of-concept assessment of matrix applicability. Nevertheless, additional investigations involving incurred clinical specimens and full bioanalytical validation are required before the methodology can be considered suitable for routine clinical practice or pharmacokinetic studies.