The effect of sodium nitroprusside (SNP) of different concentrations (0.05, 0.1, 0.2 mM) on biomass and extracellular polyphenol accumulation and antioxidant defense system of Lachnum YMU50 had been studied in this article. Results showed that, when the NO donor was SNP, both 0.05 and 0.1 mM SNP could promote the biomass accumulation of Lachnum YMU50 and activate the phenylalanine ammonia lyase, thus promoting its extracellular polyphenol accumulation and activate and induce its antioxidant defense system by activating the activities of catalase, superoxide dismutase, and ascorbate peroxidase, and 0.1 mM SNP has a stronger effect than 0.05 mM SNP. SNP 0.2 mM was adverse to the accumulation of the biomass and extracellular polyphenol of Lachnum YMU50, decreasing the activities of catalase, superoxide dismutase, and ascorbate peroxidase and increasing the contents of hydrogen peroxide and malondialdehyde, which indicated that 0.2 mM SNP damaged the antioxidant defense system of Lachnum YMU50.