A series of novel 4-(3,5-dimethyl-1H-pyrazol-1-yl)-2-substituted phenyl/Heterocyclic Thieno[2,3-d] Pyrimidine (8 a-j) derivatives were synthesized by a facile Five-step procedure that afforded advantages of mild reaction conditions, simple protocol and good yields. The structures of the final compounds were confirmed by IR, NMR, EI-MS. The final compounds were screened for their anti-bacterial activity against Bacillus subtilis and Staphylocouccus aureus from Gram positive group of bacteria and Escherichia coli and Klebsiella pneumonia from Gram negative group of bacteria and antifungal activity against Candida albicans and Aspergillus flavus. Anti-bacterial and anti-fungal activities were Evaluated and compared with the standard drugs Such as Amoxicillin and Ketoconazole. From anti-bacterial and antifungal activity screening results, it has been observed that compounds 8i, 8h, 8e and 8j possess good activity.