The chemical oxytocin helped men in romantic relationships keep their distance from strangers they might find attractive.
They stayed about four to six inches further away when approaching or being approached by good-looking women than those given a dummy drug.
Dubbed the 'cuddle drug', oxytocin is naturally made in the body and is involved in sex, sexual attraction, trust and confidence.
It is released into the blood during labour - triggering the production of breast milk - and floods the brain during breastfeeding, helping mother and baby bond.
Researchers said their findings published in The Journal of Neuroscience suggest oxytocin could promote fidelity. In contrast oxytocin had no effect on single men.
Dr René Hurle*mann, of Bonn University in Germany, said: "Previous animal research in prairie voles identified oxytocin as major key for monogamous fidelity in animals.
"Here we provide the first evidence that oxytocin may have a similar role for humans."
In the study his team administered oxytocin or a placebo via a nasal spray to fifty-seven healthy and heterosexual men, about half of whom were in monogamous relationships.
Forty-five minutes later the participants were introduced to a female experimenter they later described as "attractive".
As the woman moved towards or away from the volunteers the men were asked to indicate when she was at an "ideal distance" as well as when she moved to a place that felt "slightly uncomfortable."
Dr Hurlemann said: "Because oxytocin is known to increase trust in people we expected men under the influence of the hormone to allow the female experimenter to come even closer - but the direct opposite happened."
The effect of oxytocin on the monogamous men was the same regardless of whether the beauty maintained eye contact or averted her gaze - or if the men were the ones approaching or withdrawing from her.
Oxytocin also had no effect on the men's attitude towards the woman - both those who received the hormone and the placebo rated her as being equally attractive.
In a separate experiment the researchers found oxytocin had no effect on the distance men kept between themselves and a male experimenter.
They said future studies are needed to determine exactly how oxytocin might act on the brain to affect behaviour.
Psychiatrist Professor Larry Young, of Emory University in Atlanta who was not involved in the study, said the hormone could be nature's way of encouraging fathers not to stray.
He said: "In monogamous prairie voles we know oxytocin plays an important role in the formation of the pair bond.
"This study suggests the general role of oxytocin in promoting monogamous behaviour is conserved from rodents to man."
