Tree peony seeds, traditionally used for edible oil production, are rich in α-linolenic acid (ALA). However, little attention is paid to their development as a healthcare food due to their bitter and astringent taste. The aim of this study was to optimize the debittering process of peony seeds on the basis of maintaining nutritional value and to identify the compounds that cause the taste of bitterness. We first optimized the debittering process by orthogonal experiments which reduced the polyphenol content by 90.25%, and we measured the main nutritional value of fatty acid composition, indicating that the high content of ALA is not affected by debittering. Then, we identified and determined the types and content of polyphenols, the metabolites causing bitter taste, in the samples based on LC-ESI-QQQ-MS. Principal component analysis (PCA) and orthogonal partial least square discriminant analysis (OPLS-DA) were used to compare and analyze the seeds at different stages of debittering. Thirty-eight key metabolites were identified, of which paeoniflorin, taxifolin, alibiflorin, protocatechuic acid, benzoyl paeoniflorin, quercetin-3-galactoside, and oxpaeoniflorin were significantly compared, and most of them were positively correlated with bitter taste. These results are conducive to the exploration and study of the bitter taste and nutritional value of tree peony seeds in the future.

Fatty acids play critical roles in plant growth and stress adaptation, primarily through modulating membrane fluidity. This study combined bioinformatics (genome-wide identification and chromosomal localization) with experimental techniques (RT-qPCR, VIGS, and GC) to investigate the ARF family in Paeonia rockii. Seventeen PrARF genes were identified, showing evolutionary collinearity with Arabidopsis thaliana and Vitis vinifera and uneven chromosomal distribution. Among these, PrARF9 was specifically and highly expressed during late seed development, exhibiting a pattern highly consistent with the fatty acid synthesis key gene PrFAD3 and the accumulation trend of α-linolenic acid (ALA). In Nicotiana benthamiana, transient overexpression of PrARF9 upregulated its homologous gene NbFAD3, resulting in increased total fatty acid content and elevated lipid droplet accumulation. In contrast, in Paeonia rockii, silencing of PrARF9 downregulated PrFAD3 expression and reduced fatty acid levels, whereas overexpression of PrARF9 produced the opposite effect. We present a comprehensive analysis of the ARF gene family in P. rockii, combined with functional verification of a candidate gene regulating lipid synthesis. In summary, PrARF9 positively regulates PrFAD3, thereby participating in oil accumulation and ALA synthesis in P. rockii.