The vulnerable plant Dalbergia tonkinensis Prain, is a rare species, native to Vietnam. Phytochemical investigations and biological evaluations of this species are quite limited. Antimicrobial screening has suggested that, at the low dose of 1.0 mg/mL, the methanol extracts of the leaf, stem bark, and root, as well as chloroform fraction of heartwood can serve as useful sources against seven gram-positive skin microbacteria, Bacillus cereus (ATCC27522), Escherichia coli (JM109), Staphylococcus aureus (ATCC25923), Staphylococcus epidermidis (ATCC14990), Streptococcus pneumonia (ATCC49619), Streptococcus pyogenes (ATCC12344), and Vibrio parahaemolyticus (RIMD2210010), and four oral gram-positive microbacteria Streptococcus mutans (ATCC 25175), Streptococcus mitis (ATCC903), Streptococcus sobrinus (ATCC 33478), and Porphyromonas gingivalis (ATCC33277), with an inhibitory percentage of 60-80% growth for several strains Bacillus cereus, Escherichia coli, and Streptococcus pneumonia. We further assessed antimicrobial activities of the enriched diverse flavonoids from chloroform fraction of its heartwood. Extensive structural activity relationship studies showed structure function closely related to the antimicrobial activity, in which methoxylation at C-2’, and 4’ in isoflavanones, hydroxylation at C-3’ in flavones, substitution at C-5 in isoflavones, and lactone opened ring in neoflavonoids were found to increase the effective inhibitions. In the second antimicrobial assessment, the isolated flavonoid liquiritigenin showed the MIC values of 50, and 100 μg/mL against the microbacterial strains Staphylococcus aureus subsp. aureus (ATCC 11632), and Aspergillus niger (439), respectively, whereas the MIC value of 100 μg/mL was assignable to biochanin A against microbacterium Staphylococcus aureus subsp. aureus.