Abstract Background Research on aging is growing as the elderly make up a greater share of the population, focusing on reversing and inhibiting the aging process. The exhaustion and senescence of stem cells are the fundamental drivers behind aging. β-Carotene has been depicted to have many biological functions, and we speculate that it may have an anti-aging effect. Methods Firstly, the anti-aging property of β-carotene was investigated in vitro using mesenchymal stem cells (MSCs) induced by H₂O₂. The anti-aging effect was characterized using Western-bloting, confocal laser scanning microscopy, indirect immunofluorescence, and immunohistochemistry. The anti-aging property was also tested in vivo using aged mice. Results The in vitro experiment revealed that β-carotene could relieve the aging of MSCs, as evidenced by a series of aging marker molecules such as p16 and p21. β-Carotene appeared to inhibit aging by regulating the KAT7-P15 signaling axis. The in vivo experiment revealed that β-carotene treatment has significantly down-regulated the aging level of tissues and organs. Conclusions In this work, we explored the anti-aging effect of β-carotene in vivo and in vitro. The experimental results indicate that β-carotene may be an important potential anti-aging molecule, which can be used as a drug or in functional food to treat aging in the future.