Tens of thousands of faithful gathered in a sun-drenched St Peter's Square to attend the mass in which Francis formally bestowed the sainthoods, which had been approved by his predecessor Benedict XVI.
The new saints include Italian cobbler Antonio Primaldo, who was killed along with an estimated 800 other Italians, who remain anonymous, in 1480 by Ottoman forces in the southern part of the country for refusing to convert to Islam.
"While we venerate the martyrs of Otrante, ask God to support the many Christians who still suffer from violence and give them the courage and fate and respond to evil with goodness," he said, referring to the persecution of Christians.
Francis used the occasion to take a firm stance against abortion for the first time since his election in March, calling for the "protection of the embryo".
He called for legislation to "protect all human beings from the first moment of their existence," lending his support to a pro-life march of some 30,000 that also took place in Rome on Sunday.
The new pope also canonised Colombia's Laura di Santa Caterina da Siena Montoya y Upeguila, and Mexico's Maria Guadalupe Garcia Zavala, who both died in the 20th century.
The two women founded religious orders and devoted their lives to helping the poor, the sick and native peoples.
The pope paid homage to di Santa Caterina da Siena Montoya y Upeguila, or "Madre Laura" -- Colombia's first saint -- for respecting the local culture and traditions while teaching natives during her mission.
He also praised Guadalupe Garcia Zavala, whom he referred to as "Madre Lupita", for "kneeling in front of the sick on the hospital pavement to treat them with tenderness."
Madre Lupita is Mexico's second saint.
"There must be no shame, fear or disgust to touch the flesh of Christ," said Francis, the first Latin American pope, underscoring the importance of charity and to help others.
Hundreds of Colombians and Mexicans travelled to the Vatican for the event, with some Mexican nuns in the crowd sporting wide-brimmed sombreros. Many parents had also brought their children along in the hope that the pope would bless them.
Maria Rosales Gomez from Mexico said she was convinced that Madre Lupita had worked a miracle 14 years ago, when her sick baby boy recovered from an illness and survived.
Speaking before Colombia's President Juan Manuel Santos, the pope also underscored his support for reconciliation efforts between the Colombian government and leftist guerrillas and called for an end to "violence and insecurity" in Mexico, where thousands of people each year fall victim to drug violence.