"The businessman and investor who wants to come, we must be hospitable. They must come and leave Iran easily," Rouhani said in a speech to promote trade in Tehran.
The moderate president's comments came just two days after the judiciary, which is controlled by more hardline elements of the regime, announced that it had sentenced business consultant Siamak Namazi and his 80-year-old father Baquer Namazi to 10 years each for "espionage and collaboration with the American government".
Both are dual nationals with extensive connections in both countries, having spent years trying to promote closer trade and diplomatic ties between the US and Iran.
Signup to our newsletter and follow us on Facebook and Twitter!
Hardliners in Iran allowed Rouhani's moderate government to sign a nuclear deal with world powers last year that lifted international sanctions, but have strongly opposed any further efforts to improve ties with the West.
"All the efforts... of the government in the last three years including in the (nuclear deal) have been designed to remove obstacles in the way of businessmen, entrepreneurs, exporters of goods and services, entry of capital and banking relations," said Rouhani.
"Our aim was to remove Iranophobia. The government has done a great job in this regard. Iran is shouting with a loud voice that I am... kinder than how I have been portrayed in the world, I want peaceful coexistence with the region and the world," he added.
"I ask and request all factions and groups (within the country): let's hear Iran's voice and respond positively."