Biosynthetic nano-selenium (bio-SeNP), as a plant growth regulator, has better bioavailability and lower toxicity than selenite and selenate. This study investigated the beneficial role of bio-SeNP in mitigating the adverse effects of multiple heavy metals (HMs, e.g., Cd, Pb, and Hg) on growth and yield of pak choi (Brassica chinensis) grown in slightly or heavily polluted (SP or HP) soil by regulating metabolic and antioxidant systems. The results revealed that foliar application of bio-SeNP (5, 10, 20 mg L^-1 Se) at the 6-leaf stage greatly reduced the levels of Cd, Pb, and Hg in shoots and roots of pak choi. Application of 5 mg L^-1 bio-SeNP significantly (p < 0.05) decreased the translocation factor (TF) of Cd, Pb, and Hg from root to shoot by 9.83%, 44.21%, and 46.99% for SP soil, 24.17%, 56.00%, and 39.36% for HP soil, respectively. Meanwhile, all bio-SeNP treatments led to a significant improvement in plants growth by enhancing the antioxidant defense system (e.g., AsA-GSH) and promoting chlorophyll synthesis as well as suppressed the lipid peroxidation products contents (MDA) in shoots. Moreover, the enhanced levels of mineral nutrient elements (e.g., Ca, Mg, Fe, or Zn) and organic selenium (e.g., selenocystine, Se-methylselenocysteine, and selenomethionine) in the edible shoots of bio-SeNP-treated pak choi plant under multiple HMs stress indicated the positive impacts of bio-SeNP on the improvement of shoot quality and nutritional values. Collectively, our results indicated that bio-SeNP play an important role in the management of multiple HMs-induced adverse effects on pak choi. Foliar application of bio-SeNP at appropriate concentration (≤ 5 mg L^-1 Se) can be considered as a promising agronomic measure for safety leafy vegetable production in multiple HMs polluted soils when bio-SeNP application.