A selective, simple, rapid, green, cost-effective, and non-destructive assay was proposed for the simultaneous quantitation of Fampridine (FPN), Dexamethasone (DMS), and Fluoxetine (FLX) in spiked human plasma and commercial dosage forms without interference from common drugs excipients. The solid state of a tertiary mixture of FPN, DMS, and FLX was determined by Fourier transformation infrared (FTIR) Spectroscopy. The calibration curves were linear in the ranges of 1.0–8.0, 0.9–8.0, and 1.2–10.0 µg/mg for FPN, DMS, and FLX, respectively. The limits of detection (LODs) were 0.34, 0.30, and 0.40, and the limits of quantitation (LOQs) were 1.0, 0.9, and 1.2 µg/mg for FPN, DMS, and FLX respectively. The suggested method was validated using the ICH guidelines. The presented assay was properly used for the analysis of the cited drugs in spiked human plasma and pharmaceutical formulations.