Abstract Panax quinquefolius L., with a history of over 300 years in traditional Chinese medicine, is notably rich in ginsenosides—its primary bioactive components. Although our previous study found that biochar application could enhance the content of ginsenoside Re, Rg and other contents in P. quinquefolius, its effect on the overall secondary metabolism of P. quinquefolius and its mechanism are still unclear. In this paper, the correlation between plant microbiome and secondary metabolites was studied from the perspective of plant rhizosphere microorganisms and endophytes, and the mechanism of biochar-induced metabolic reprogramming of P. quinquefolius was revealed. The results showed that biochar treatment significantly increased the accumulation of various substances in P. quinquefolius, including nucleosides, glycerophosphocholines, fatty acyls, steroidal glycosides, triterpenoids, and other bioactive compounds. Additionally, biochar treatment significantly enriched beneficial rhizosphere microorganisms such as Bacillus, Flavobacterium, and Devosia, while reducing the relative abundance of harmful fungi like Fusarium. Furthermore, it promoted endophytic Flavobacterium, Acaulospora, and Glomus, and suppressed pathogenic genera such as Plectosphaerella, Cladosporium, and Phaeosphaeria. These shifts in rhizosphere microbial community and endophytes structure and function were closely linked to the accumulation of secondary metabolites (e.g. ginsenosides Rg3, F2) in P. quinquefolius. Overall, our findings suggest that biochar may influence key endophytes and rhizosphere microorganisms to regulate the accumulation of secondary metabolites in P. quinquefolius. Therefore, this study provides valuable insights into the potential application of biochar in Chinese medicine agriculture.