Solid-phase synthesis of [8-arginine]-vasopressin through a crystalline protected nonapeptide intermediate and biological properties of the hormone

Abstract

The neurohypophyseal hormone [8-arginine]-vasopressin has been synthesized by the solid-phase method. The protected nonapeptide intermediate, S-benzyl-TV-tosyl-L-cysteinyl-L-tyrosyl-L-phenylalanyl-L-glutaminyl-L-asparaginyl-S-benzyl-L-cysteinyl-L-prolyl-TV°-tosyl-L-arginylglycinamide, has been obtained in crystalline form. Removal of the protecting groups from the crystalline protected nonapeptide followed by oxidative cyclization provided highly active hormone preparations in excellent yields. Further purification by ion exchange chromatography afforded [8-arginine]-vasopressin possessing 503 ± 53 units/per mg of rat antidiuretic activity, 487 ±15 units/mg of rat pressor activity, 12.0 ± 0.2 units/mg of rat oxytocic activity, 30-120 units/mg of rabbit milkejecting activity, and 100 ± 5 units/mg of avian vasodepressor activity.The solid-phase method of peptide synthesis6 hag been successfully used for the preparation of several naturally occurring neurohypophyseal peptides such as oxytocin,6 [8-lysine]-vasopressin,7 and glumitocin.8This paper describes the application of this method to the synthesis of the mammalian antidiuretic principle, [8-arginine]-vasopressin,9 Figure 1. Chiefly through the crystallization of the protected nonapeptide (S-benzyl-TV-tosyl-L-cysteinyl-L-tyrosyl-lphenylalanyl-L-glutaminyl-L-asparaginyl-S'-benzyl-Lcysteinyl -L-prolyl-TVG-tosyl-L-arginylglycinamide, I, Figure 2) the hormone was obtained in excellent yield and with high biological potency.Earlier syntheses of [8-arginine]-vasopressin gave preparations exhibiting considerable variation in the * To whom correspondence should be addressed.

Inhibition of Vasopressin-Stimulated Urea Transport Across the Toad Bladder by Thiourea

“…In this approached the molarity of the mucosal fluid. In other experiment urea-linked volume flux increased 24-fold words, the urea-linked volume flux across the bladder in the presence of vasopressin, whereas net urea flux in-wall was initially a hypertonic volume flux, and only afcreased only 13-fold. The molarity of the urea solution ter a lag period became an isotonic volume flux.…”

Unequivocal assignments of NH proton magnetic resonance bands in oxytocin using 2H‐ and 15N‐substituted molecules

“…The protected nonapeptide amide was liberated from the resin by ammonolysis in a mixture of dimethylformamide and ethanol (1:1 v/v) saturated with ammonia at -5°C, stirring the suspension at room temperature for 48 hr [14].…”

“…The remaining free thiol groups were oxidised by air at pH 6.8 [14]. The solution was acidified to pH 4 with acetic acid and freeze-dried.…”

Synthesis of Tritium Labelled [8-Arginine]-vasopressin

“…Position 3 was chosen as the one to have the radioactive residue since it was the last Dosition to be filled tide were carried out according to the procedure of Meienhofer et a/. (5).…”