We report ground-state formation of 3-hydroxyflavone (3HF) anion in DMSO. According to the literature this process is possible only in hydrogen bonding donor (HBD) solvents. Our results show that a solvent with zero HBD acidity but sufficiently high hydrogen bonding acceptor basicity can also give anion formation. A possible new mechanism is proposed. Furthermore, theoretical calculations show evidence for the formation of a 3HF center dot center dot center dot DMSOH+ complex. As 3HF anion is a promising biophysical fluorescent probe, our results can help to understand the factors governing its ground-state formation and how its spectral properties are related to its microenvironment in biological systems. (C) 2008 Elsevier B. V. All rights reserved.