目的 研究不同栽培基质对近野生铁皮石斛Dendrobium officinale药效成分积累的调控机制，为其品质差异的分子基础及栽培基质优化提供依据。方法 以石生和木生栽培基质的铁皮石斛样品为材料，采用非靶向代谢组学鉴定并比较2种栽培模式下次生代谢物谱以及通过转录组学测序用于筛选差异表达基因，并结合KEGG等数据库进行通路富集分析，并与代谢物变化进行整合分析。结果 代谢组共鉴定2 878个代谢物，其中354个在2种模式间显著差异，主要涉及苯丙素类、类黄酮、生物碱、脂质及芳香族氨基酸等物质。石生样品中黄酮类、生物碱类及部分氨基酸类代谢物整体积累水平较高，木生样品中脂质及类固醇类物质相对富集。转录组测序筛选出2 966个差异表达基因，KEGG富集分析显示苯丙素类生物合成（ko00940）、类黄酮生物合成（ko00941）、苯丙氨酸/酪氨酸/色氨酸生物合成（ko00400）及托品烷、哌啶和吡啶生物碱生物合成（ko00960）等通路显著分化。类黄酮合成关键酶[如苯丙氨酸解氨酶（phenylalanine ammonia lyase，PAL）、查耳酮合成酶（chalcone synthase，CHS）、关黄烷酮3-羟化酶（flavanone 3-hydroxylase，F3H）]基因在石生样品中上调表达，与相关代谢物积累趋势一致。联合分析表明，栽培基质可能通过调控苯丙素-类黄酮-生物碱代谢相关基因表达，改变代谢通量分配方向，形成不同代谢表型。结论 栽培基质通过调控苯丙素、类黄酮及生物碱等相关代谢通路基因的表达，影响铁皮石斛活性成分的种类与积累水平，从而造成石生与木生栽培品质量的差异。系统阐明了铁皮石斛品质差异的分子机制，为栽培基质优化和药材分级利用提供理论依据。

Objective To investigate the regulatory mechanisms by which different cultivation substrates influence the accumulation of pharmacologically active components in near-wild Tiepishihu (Dendrobium officinale), and provide a basis for elucidating the molecular mechanisms underlying quality differences and for optimizing cultivation substrates. Methods D. officinale samples grown on stone-based and wood-based substrates were used as experimental materials. Non-targeted metabolomics was employed to identify and compare the secondary metabolite profiles under the two cultivation modes, while transcriptome sequencing was conducted to screen differentially expressed genes (DEGs). KEGG and other databases were used for pathway enrichment analysis, which was then integrated with metabolite variation data. Results A total of 2 878 metabolites were identified by metabolomics, of which 354 showed significant differences between the two cultivation modes. These metabolites were mainly involved in phenylpropanoids, flavonoids, alkaloids, lipids, and aromatic amino acids. Flavonoids, alkaloids, and certain amino acid-derived metabolites exhibited overall higher accumulation in stone-grown samples, whereas lipids and steroid-related compounds were relatively enriched in wood-grown samples. Transcriptome sequencing identified 2 966 DEGs. KEGG enrichment analysis indicated significant differentiation in pathways including phenylpropanoid biosynthesis (ko00940), flavonoid biosynthesis (ko00941), phenylalanine/tyrosine/tryptophan biosynthesis (ko00400), and tropane, piperidine and pyridine alkaloid biosynthesis (ko00960). Genes encoding key enzymes in flavonoid biosynthesis [such as phenylalanine ammonia lyase (PAL), chalcone synthase (CHS), flavanone 3-hydroxylase (F3H)] were upregulated in stone-grown samples, consistent with the accumulation patterns of the corresponding metabolites. Integrated analysis suggested that cultivation substrates may regulate the expression of genes involved in phenylpropanoid-flavonoid-alkaloid metabolism, thereby altering the direction of metabolic flux allocation and ultimately shaping distinct metabolic phenotypes. Conclusion Cultivation substrates influence the types and accumulation levels of active constituents in D. officinale by regulating the expression of genes in phenylpropanoid, flavonoid, and alkaloid biosynthetic pathways, resulting in quality differences between stone-grown and wood-grown materials. This study systematically elucidates the molecular mechanisms underlying quality variation in D. officinale and provides a theoretical basis for cultivation substrate optimization and graded utilization of medicinal materials.

R282

国家自然科学基金区域创新发展联合基金项目（U25A20163）；2025年浙江省大学生科技创新活动计划（2025R406A037）