Quality differentiation and salidroside biosynthesis in Rhodiola crenulata, R. fastigiata, and intergrades

Rhodiola crenulata , the only authentic medicinal species recorded in the Chinese Pharmacopoeia, is increasingly threatened by resource depletion and adulteration, raising concerns regarding its quality and safety. Here, an integrative multi-analysis combining phenotypic traits, anatomical characteristics, metabolite profiling, and transcriptome sequencing was conducted to systematically compare R. crenulata , R. fastigiata , and naturally occurring intergrades. R. crenulata exhibited a lower rhizome yield (34.9 g) than R. fastigiata (55.6 g) and intergrades (39.5 g), but displayed a larger stem diameter (3.9 mm) and leaf width (1.3 cm), along with a shorter stem length (14.8 cm). Anatomically, it possessed a double-layered vascular cambium with an irregular arrangement, accompanied by significantly larger vessel elements (324.2 μm²) and storage parenchyma cells (1882.5 μm²). Metabolite analysis showed that the salidroside content in R. crenulata (0.81%) was 20-fold higher than in R. fastigiata and 4-fold higher than in intergrades, while tyrosol (0.45‰) and total flavonoids (7.2%) remained the highest. Transcriptomic analysis revealed extensive differential gene expression among genotypes and along different axial positions of the rhizome, with the phenylpropanoid biosynthesis and tyrosine metabolism pathways being significantly enriched. Co-expression and molecular docking identified six candidate UDP-glycosyltransferases with strong in silico substrate-binding affinity, among which RcUGT5914 showed species-specific expression. Collectively, this research provides a multi-dimensional framework for the accurate authentication and quality evaluation of R. crenulata , and establishes a molecular basis for its conservation and genetic improvement.