Fasciculins are peptides present in the venom of green and black mamba snakes, with potent inhibitory activity towards acetylcholinesterase. In order to determine the role of fasciculin loop II in the acetylcholinesterase inhibition, two fasciculin fragments were synthesized by the solid phase procedure using N-alpha-Boc protected amino acids. The two peptides, Fas-A and Fas-B, span the 26-32 and 22-35 sequences of fasciculin and a disulfide bridge links each peptide end, thus ensuring the formation of a looped structure. Both peptides were characterized chemically, structurally and functionally. Circular dichroism indicated the existence of 19.4 and 24.9% of beta-sheet for Fas-A and Fas-B, respectively; SDS-PAGE patterns and mass spectrometry disclosed the intramolecular disulfide formation in both peptides. An inhibitory effect on eel acetylcholinesterase was observed with the longer peptide (Ki = 15.1 microM), without reaching the affinity level of the parent native toxin (Ki = 0.3 nM). This study confirms that fasciculin central loop residues strongly contribute to toxin interaction with acetylcholinesterase.