The role of arginine residues in the structure-activity relationships of alpha-scorpion neurotoxins was studied. Toxins I and II from Androctonus australis Hector (north African scorpion), containing respectively 2 and 3 arginines, were modified by phenylglyoxal or p-hydroxyphenylglyoxal. Modified derivatives were purified by reverse-phase HPLC and/or ion exchange HPLC. Subsequent bioassays showed that toxin I (AaH I) derivatives with single modifications on Arg 2 and Arg 60 had low activity (25 and 14% of residual activity, assessed in receptor binding experiments). Doubly modified (Arg 2, Arg 60) AaH I had 7% residual activity while further derivatization of the alpha-amino group led to an almost inactive derivative. These results agree with the involvement of arginines 2 and 60, as well as the alpha-amino group, of AaH I in the toxin/receptor interaction, probably via electrostatic interactions. Consistent with the role of N-terminal residues, the selective removal of the N-terminal dipeptide Val-Arg of toxin III from the same scorpion resulted in low activity (7% residual activity). The arginine residue in position 56 of toxin II was important for bioactivity since the derivative modified by phenylglyoxal on Arg 56 exhibited low residual activity (20%). Arg 62 and Arg 18, on the other hand, can be modified without any great effect on the pharmacological activity of AaH II. These results furnish a more precise picture of those residues involved in the "toxic region", which appears to be composed of residues belonging to the conserved hydrophobic surface and to the C-terminal and N-terminal sequences.