Ortho-substituted polymethoxydiarylazolopyrimidines were synthesized using polymethoxysubstituted benzaldehydes and acetophenones as starting material. X-ray crystallography data clearly confirmed that the subsequent cyclization of 3-amino-1,2,4-triazole with ketoaldehydes yielded polymethoxyphenylsubstituted 6,7-diaryl-[1,2,4]triazolo[1,5-a]pyrimidines as single isomers. All compounds were evaluated in vivo using phenotypic sea urchin embryo assay. 6-(4-Methoxyphenyl)-7-(3,4,5-trimethoxyphenyl)pyrazolo[1,5-a]pyrimidine showed antimitotic microtubule destabilizing activity. The importance of aryl rings substituents in diaryltriazolopyrimidines for their antiproliferative antitubulin effect has been suggested. © ARKAT USA, Inc.