When Murad died in 1595, Safiye arranged for her son Mehmed to succeed as sultan, and she became the Valide Sultan—one of the most powerful in Ottoman history. Until her son's death in 1603, Ottoman politics were determined by a party headed by herself and Gazanfer Ağa, chief of the white eunuchs and head of the enderun (the imperial inner palace).

Safiye eventually enjoyed an enormous stipend of 3,000 aspers a day during the latter part of her son's reign.When Mehmed III went on the Eger campaign in Hungary in 1596, he gave his mother great power over the empire, leaving her in charge of the treasury. During her interim rule she persuaded her son to revoke a political appointment of the judgeship of Istanbul and to reassign to the grand vizierate to Damat Ibrahim Pasha, her son-in-law.

During this period, the secretary of the English ambassador reported that while in the palace, Safiye "espied a number of boats upon the river [the Bosphorus] hurrying together. The Queen Mother sent to enquire of the matter [and] was told that the Vizier did justice upon certain chabies [kahpe], that is, whores. She, taking displeasure, sent word and advised [the Vizier] that her son had left him to govern the city and not to devour the women; [thus] commanding him to look well to the other business and not to meddle any more with the women till his master's return."

The greatest crisis Safiye endured as valide sultan stemmed from her reliance on her kira, Esperanza Malchi. A kira was a non-Muslim woman (typically Jewish) who acted as an intermediary between a secluded woman of the harem and the outside world, serving as a business agent and secretary. In 1600, the imperial cavalry rose in rebellion at the influence of Malchi and her son, who had amassed over 50 million aspers in wealth. Safiye was held responsible for this, along with the debased currency the troops were paid with, and nearly suffered the wrath of the soldiers, who brutally killed Malchi and her son. Mehmed III was forced to say "he would counsel his mother and correct his servants." To prevent the soldiers from suspecting her influence over the Sultan, Safiye persuaded Mehmed to have his decrees written out by the Grand Vizier, instead of personally signing them.

Interior design of Yeni Mosque in Eminönü, Istanbul. The construction began during Safiye's regency.

Safiye was instrumental in the execution of her grandson Mahmud in 1603, having intercepted a message sent to his mother by a religious seer, who predicted that Mehmed III would die in six months and be succeeded by his son. According to the English ambassador, Mahmud was distressed at "how his father was altogether led by the old Sultana his Grandmother & the state went to Ruin, she respecting nothing but her own desire to get money, & often lamented thereof to his mother," who was "not favored of the Queen mother." The sultan, suspecting a plot and jealous of his son's popularity, had him strangled.