Foliar Spraying of Brassinolide Affects Leaf Quality and Secondary Metabolite Profiles of Cold-Stressed Tea Plants

Cold stress is an important limiting factor affecting spring tea quality. This study analyzed the effects of foliar spraying of brassinolide (BR) at concentrations of 0.05, 0.1, 0.5, and 1 mg/L on the chlorophyll index, nitrogen balance index, quality, antioxidant defense system, and secondary metabolite profiles in leaves of Camellia sinensis cv. Fuding-dabaicha grown at 4 °C for 48 h. All exogenous BR treatments significantly increased leaf nitrogen balance index, with the highest effect at 0.1 mg/L, which also significantly increased leaf chlorophyll index. BR treatments distinctly increased tea polyphenol, catechin, amino acid, and caffeine levels at cold stress, with the greatest effect at 0.1 mg BR/L. Foliar spraying of BR showed no effect on the expression of CsGOGAT at cold stress, but it differentially regulated the expression of CsHMGR, CsGDH, and CsGs, accompanied by their expression being up-regulated under 0.1 mg BR/L treatment. BR-treated plants exhibited a low level of oxidative damage at cold stress based on malondialdehyde levels, which was associated with higher glutathione levels and CsCAT and CsSOD gene expression levels under BR concentrations of 0.05 mg/L and 0.1 mg/L. Non-targeted metabolomics found a total of 26,175 metabolites, the majority of which were lipids and lipid-like molecules (8.97%) and organic heterocyclic compounds (8.97%). BR treatments with 0.05, 0.1, 0.5, and 1 mg/L triggered 1181, 1997, 2414, and 1455 differential metabolites, respectively, accompanied by more differential metabolites being up-regulated. Among them, 18 differential metabolites were associated with tea quality. The enriched pathways of differential metabolites were mainly caffeine metabolism, amino acid synthesis and metabolism, alkaloid synthesis and metabolism, and flavonoid synthesis, depending on the BR concentrations used. Caffeine metabolism was an BR-inducible differential metabolite pathway. Taken together, foliar spraying of BR (0.1 mg/L) improved leaf antioxidant capacity and quality as well as modulated secondary metabolites and their pathways in cold-stressed tea.