The synthesis and biological activities of arginine-vasopressin analogues are described, where p-azido-L-phenylalanine [Phe(pN3)] or p-(bromoacetylamino)-L-phenylalanine [Phe-(pNHCOCH2Br)] replace Tyr2 or Phe3. The hormone analogues are prepared via precursors containing p-aminophenylalanine [Phe(pNH2)] in position 2 or 3. During peptide synthesis the p-amino group of [Phe(pNH2)] is protected by the tert-butyloxycarbonyl or the benzyloxycarbonyl group, the side chains of cysteine and arginine by the acetamidomethyl residue and the tosyl group, respectively. The amino and guanidino protecting groups are removed from the nonapeptides by trifluoromethanesulfonic acid yielding the S-protected derivatives which are cyclized by means of iodine. The ring closure by disulfide formation is confirmed by Edman degradation, CD and 1H-NMR spectroscopy. Modification at the p- and alpha-amino groups result in [Phe(pN3)2]-vasopressin, [Phe(pNHCOCH2Br)2]vasopressin, Nalpha-dansyl-[Phe(pN3)2]vasopressin, [Phe2,Phe-(pN3)3]vasopressin and [Phe2,Phe(pNHCOCH2-Br)3]vasopressin. The analogues modified only in position 2, [Phe(pN3)2]vasopressin stimulate the adenylate cyclase derived from bovine kidney inner medulla to similar maximal velocities as arginine vasopressin and show high apparent affinities for enzyme activation. The Nalpha-dansyl derivative and the analogues with reactive groups in position 3 have reduced maximal velocities and apparent affinities for vasopressin-sensitive adenylate cyclase. These results suggest that especially the derivatives with reactive groups in position 2 are useful for the labelling of vasopressin receptors in plasma membranes and for studies of covalent hormone-receptor complexes.