Levofloxacin, a quinolone used in the treatment of an upper respiratory tract infection is involved in skin reactions. Patch tests are helpful for the aetiological diagnosis. Unfortunately, they are known for their lack of sensibility. A weak transcutaneous penetration due to the physicochemical characteristics of tested drugs is a well-known cause of false negative results but these characteristics are usually unknown. The aim of the present study was to evaluate physicochemical parameters by potentiometric method in order to optimize the transcutaneous penetration of a preparation for patch test. This method is applicable to a medium such as the preparation for patch test. In order to value the stability of the preparation, qualitative and quantitative analysis of the level of levofloxacin was effectuated. In our study, spectrophotometric and potentiometric data at T0 and six months later with the preparation showed the stability of the preparation. The proportions of non-ionized (45%) and ionized levofloxacin (55%) were not modified. Further experiments are needed to choose the minimal efficient dose of the suspected molecule able to detect a positive reaction in cases of allergic reaction. In addition, this method could be useful in qualitative and quantitative control of preparations made at hospital.