A new N-butyl-4-amino-1,8-naphthalimide-based colorimetric and ratiometric fluorescent probe for the detection of biothiols (cysteine, homocysteine, and glutathione) was designed and synthesized. The probe exhibited good selectivity and sensitivity toward biothiols over other non-thiol-bearing amino acids and common anions with significant changes in both color (from colorless to yellow) and fluorescence