| Summary: | Browning is a major problem in the tissue culture of woody plants. Previous studies have shown that nitric oxide (NO) plays a role in regulating plant responses to stress, but its effect on browning in the tissue culture of Malus remains unclear. This study aimed to investigate the impact of exogenous NO donor sodium nitroprusside (SNP) on the browning of <i>Malus sieversii</i> stem tip explants. The results demonstrated that the addition of 50 μM SNP significantly reduced explant browning. Further analysis revealed that exogenous NO decreased the browning index (BD) and levels of malondialdehyde (MDA), hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>), and superoxide anion radical O<sub>2</sub><sup>−</sup>. Additionally, NO treatment increased the activity of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX). NO treatment also enhanced the activity of phenylalanine ammonia lyase (PAL), which contributed to the accumulation of flavonoids and inhibited the activity of polyphenol oxidase (PPO) and peroxidase (POD), which are key enzymes involved in the browning process. Furthermore, 50 μM SNP significantly promoted the accumulation of non-enzymatic antioxidants such as proline (Pro), soluble sugar (SS), and soluble protein (SP). Therefore, the results suggest that NO is able to counteract excessive reactive oxygen species (ROS) damage by enhancing both the enzymatic and non-enzymatic antioxidant systems, resulting in a reduction in browning in stem tip explants. Consequently, an improvement in the in vitro propagation efficiency of <i>Malus sieversii</i> shoot tip explants can be achieved.
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