Vasopressin V2 receptor antagonists of the type first described by Manning et al. are distinguished by a β,β-alkylidene-β-mercaptopropionic acid moiety at position 1, and the disulfide bond is crucial for their metabolic stability—the cyclic hexapeptide is resistant to proteolysis (e.g., chymotrypsin) when the disulfide is intact but becomes susceptible to endopeptidases (e.g., papain) upon reduction. To examine the role of the disulfide bond and prepare analogues with increased metabolic stability, we developed dicarbavasopressin antagonist analogues in which the disulfide bridge is replaced with two methylene groups. We synthesized optically pure Boc-L-Pas(OBzl) (suitable for solid-phase peptide synthesis) via Kolbe electrochemical coupling of the monobenzyl ester of cyclohexanediacetic acid with Boc-L-glutamic acid α-benzyl ester, followed by selective hydrolysis of the less hindered α-benzyl ester. Linear peptide intermediates were prepared by solid-phase synthesis on benzhydrylamine resin, cleaved from the resin with anhydrous liquid HF, converted to their hydrochloride salts, and cyclized in good yield with diphenyl phosphorazidate and triethylamine. Peptides were purified by preparative HPLC (Hamilton PRP-1 polystyrene column), confirmed to be homogeneous by HPLC and TLC, and their structures verified by FAB mass spectrometry. A comparison of antagonist potency (Table I) showed each dicarba analogue was approximately equipotent with its corresponding disulfide congener in three tail-modified analogues, indicating the 20-membered ring can present an equivalent antagonist pharmacophore despite the slightly altered geometry of the ethylene bridge versus the disulfide. Assessment of agonist activity (Table II) revealed the disulfide antagonist 4 exhibited full agonist activity in conscious, water-loaded dogs pretreated with indomethacin (comparable to AVP, though at higher doses), while its dicarba congener 5 showed much reduced agonist activity. A similar trend was observed for analogues 6 (disulfide) and 7 (dicarba), where 7 did not exhibit any agonist activity at a dose of 100 µg/kg. Thus, dicarbavasopressin antagonists retain full in vitro and in vivo antagonist potency but have reduced agonist activity compared to their disulfide congeners. Combined with the expected increase in metabolic stability (due to resistance to ring opening), these analogues represent more viable potential therapeutic candidates than their disulfide counterparts.