Application of Sustainable analytical chemistry concepts has become crucial in order to remove the environmentally harmful impacts originating from the routine use of analytical techniques. Here, a new LC method is developed and its parameters are analyzed, depending on a mixed micellar mobile phase. This was primarily aimed at getting rid of the use of organic solvents in conventional routine analyses. Combinations of tazobactam (TZB) with piperacillin (PPC) or cefepime (CFM) are commonly used as effective antimicrobial therapies, especially for resistant strains. Therefore, the three drugs were separated and quantified using an organic solvent-free mobile phase. The mixed micellar mobile phase was comprised of 15 mM Brij-35 with 38 mM SDS, adjusted to pH 3.5. Separation was performed by HPLC on monolithic RP-C18 column Chromolith® Performance RP-18e (100 mm × 4.6 mm) at a rate of 1 mL per minute of flow in conjunction with a measurement wavelength 210 nm. The method was found valid and applicable in accordance of precision, and accuracy within ranges of 5–100 µg mL−1 for PPC and CFM and of 0.625–12.5 µg mL−1 for TZB. The quality-by-design technique was used to analyze the effect of modifying the mixed micellar ratios on separation efficiency and conclude their behavior. Finally, the suggested approach was assessed applying the green analytical procedure index against the greenest published methodology to show superiority.