Recombinant H2 relaxin (serelaxin) has gained considerable attention as a new vasoprotective drug, largely due to its potential therapeutic effects in heart failure and fibrosis. However, serelaxin is laborious and costly to produce. A single-chain peptidomimetic, B7-33, has been developed to overcome these problems but little is known about its biological actions in the vascular system. This study first compared the rapid vascular effects of an acute bolus injection of B7-33 compared with serelaxin. Male Wistar rats received a tail vein injection of placebo (20 mM sodium acetate), B7-33 (13.3 μg/kg) or serelaxin (26.6 μg/kg). Three hours later vascular function in the mesenteric artery, small renal artery and abdominal aorta was assessed by wire myography. B7-33 and serelaxin selectively enhanced bradykinin-mediated endothelium-dependent relaxation in the rat mesenteric artery by increasing endothelium-derived hyperpolarization, but had no overall effects on relaxation in the small renal artery or aorta. We then compared the actions of B7-33 and serelaxin in an ex vivo model of vascular disease using virgin female mouse mesenteric arteries pre-incubated in placental trophoblast conditioned media to induce endothelial dysfunction characteristic of preeclampsia. Co-incubation of these arteries in trophoblast conditioned media with B7-33 or serelaxin (15, 30 nM) prevented the development of endothelial dysfunction. In conclusion, equimolar doses of B7-33 replicated the acute beneficial vascular effects of serelaxin in rat mesenteric arteries and also prevented endothelial dysfunction induced by placental trophoblast conditioned media in mouse mesenteric arteries. Therefore, B7-33 should be considered as a cost-effective vasoactive therapeutic in cardiovascular diseases, including preeclampsia.