Chemical permeation enhancers (CPEs) can enable antibiotic flux across the tympanic membrane. Here we study whether combinations of CPEs (sodium dodecyl sulfate, limonene, and bupivacaine hydrochloride) are synergistic and whether they could increase the peak drug flux. Synergy is studied by isobolographic analysis and combination indices. CPE concentration-response (i.e. trans-tympanic flux of ciprofloxacin) curves are demonstrated for each CPE, isobolograms constructed for pairs of CPEs, and synergy demonstrated for all three pairs. Synergy is much greater at earlier (6 h) than later (48 h) time points, although the effect sizes are greater later. Synergy is also demonstrated with the three-drug combination. Combinations of CPEs also greatly enhance the maximum drug flux achievable over that achieved by individual CPEs.