Abstract: A novel and mild synthesis method of thymine-¹⁵N₂ was designed. Ethyl formate and ethyl propionate as raw materials, the intermediate ethyl 2-formylpropionate was condensed under sodium metal, the target compound thymine-¹⁵N₂ was obtained by condensation with isotope urea-¹⁵N₂ under acid catalysis. Synthesis route had the advantages of simple operation, short process flow and less side product. The yield was more than 80% and isotope ¹⁵N abundance did not diluted. The product was characterized by HPLC, IR, MS, ¹H NMR and ¹³C NMR. The chemical purity was more than 99% and ¹⁵N enrichment was more than 98 atom%.