A fully automated synthesis of N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB) was carried out by a convenient three-step, one-pot procedure on the modified TRACERlab FXFN synthesizer, including [18F]fluorination of ethyl 4-(trimethylammonium triflate)benzoate as the precursor, saponification of the ethyl 4-[18F]fluorobenzoate with aqueous tetrapropylammonium hydroxide instead of sodium hydroxide, and conversion of 4-[18F]fluorobenzoate salt ([18F]FBA) to [18F]SFB treated with N,N,N′,N′-tetramethyl-O-(N-succinimidyl)uranium tetrafluoroborate (TSTU). The purified [18F]SFB was used for the labeling of Tat membrane-penetrating peptide (containing the Arg-Lys-Lys-Arg-Arg-Arg-Arg-Arg-Arg-Arg-Arg-Pro-Leu-Gly-Leu-Ala-Gly-Glu-Glu-Glu-Glu-Glu-Glu-Glu sequence, [18F]CPP) through radiofluorination of lysine amino groups. The uncorrected radiochemical yields of [18F]SFB were as high as 25–35% (based on [18F]fluoride) (n=10) with a synthesis time of∼40 min. [18F]CPP was produced in an uncorrected radiochemical yields of 10–20% (n=5) within 30 min (based on [18F]SFB). The radiochemical purities of [18F]SFB and [18F]CPP were greater than 95%. Copyright © 2010 John Wiley & Sons, Ltd.