Increasing resistance to current fungicides is a clinical problem that leads to the need for new treatment strategies. Clove oil (CO) has already been described as having antifungal action. However, it should not be applied directly to the skin as it may be irritating. One option for CO delivery and suitable topical application would be nanoemulsions (NEs). NEs have advantages such as decreased irritant effects and lower dose use. The purpose of this work was the development of NEs containing CO and in vitro evaluation against Candida albicans and Candida glabrata. The NEs were produced by an ultrasonic processor with different proportions of CO and Pluronic® F-127. In order to determine the best composition and ultrasound amplitude, an experimental design was performed. For the evaluation, droplet size and polydispersity index (PdI) were used. After the stability study, in vitro activity against C. albicans and C. glabrata was evaluated. NEs selected for the stability study, with diameter <40 nm and PdI <0.2, remained stable for 420 d. Activity against Candida spp. was improved when the CO was nanoemulsified, for it possibly leads to a better interaction between the active and the microorganisms, mainly in C. albicans.