The synthesis and in vitro antibacterial and beta-lactamase inhibitory activity of the 2-azetidinone-1-oxysulfonic acids having a substituent at C-4 position of the beta-lactam ring are described. The influence of C-4 substituents on the antibacterial activity was examined for the compounds having alpha-ureidoacetyl or alpha-oxyiminoacetyl group as acyl side chain at C-3 position. The antibacterial activity is correlated with the C-4 substituents and acyl side chain. Especially, 4(R)-methyl substituted derivatives exhibited excellent activity against Gram-negative bacteria and 4-dimethyl substituted derivatives exhibited strong activity against resistant Gram-negative bacteria except for Pseudomonas aeruginosa. 39 and 40 showed strong inhibitory activity against cephalosporinase of Enterobacter cloacae H-27.