A convenient synthesis of new optically active 2-oxaisocephems with (ω-alkylpyridinium-ω-thio)methyl groups at the 3-position and their biological activities are described. In particular, 10b and 10d having [2-(2-aminothiazol-4-yl)-2-(Z)-cyclopentyloxyimino]acetamido group at the 7-position were found to possess high in vitro potency and showed excellent in vivo efficacy. As part of our study to find more effective anti-infectives, we required a prompt and efficient synthesis of optically active 2-oxaisocephem class of β-lactam antibiotics. A synthesis of optically pure 2-oxaisocephems with different substituents at the 3- and the 7-position is considered to be one of the most attractive subjects because of the expected enhancement of antibacterial activity. Although 2-oxaisocephems have been reported to have only partial antibacterial activity, most of previous reports were concerned on racemic compounds. And the existing enantioselective syntheses are not appropriate since the introduction of various substituents into the 3-position is limited. Therefore, we intended to search more effective antibiotics which show broad spectrum of antibacterial activity by the synthesis of optically active 2-oxaisocephems with (ω-alkylpyridinium-ω-thio)methyl groups at the 3-position and 2-aminothiazol-4-yl moiety at the 7-position. In particular, we attempted to improve antibacterial activity against Methicillin-resistant Staphylococcus aureus (MRSA) which is recently a major pathogen in hospitals and has been associated with an increasing number of infections since 1961. We describe herein the synthesis of new optically active 2-oxaisocephems and their biological activities.