The present study reports the use of ultravioletevisible absorption, steady-state fluorescence, and time-resolved fluorescence to determine the water content in ethanol by using Rhodamine B as an optical probe. The experiments were performed by preparing water/ethanol blends with different percentages of water in the presence of the optical probe. The absorbance, fluorescence intensity, and fluorescence lifetime values were linearly dependent on the water content in the blends in the range between 0 and 10% (w/w). The results also revealed that the three techniques have a limit of detection for water <= 1.4% (w/w). Besides, molecular dynamics simulations were carried out to evaluate the interaction of RhB with water and ethanol molecules when subjected to different water content in the blends. The simulations revealed that water molecules perform well-oriented dipole-dipole interactions (hydrogen bonding) with chromophores/fluorophores groups of Rhodamine B, affecting its absorption and emission characteristics, and altering the microenvironment density and viscosity. The present findings point out that common optical techniques can be used to develop a simple, rapid, portable, and precise approach to monitor water in ethanol as far as RhB be used as a probe. (C) 2020 Elsevier Ltd. All rights reserved.