Guided by the observation that fluorine substitution at the 5-position of the base or the 2'-position of the sugar ring enhances the biological activity and acid stability of nucleoside analogs, and the fact that 2'-vinylfluoro-containing nucleosides have been reported in the D-series but not in the L-series, this study describes the synthesis and evaluation of a series of 2'-fluorinated-2',3'-dideoxy-2',3'-didehydro-(L)-nucleosides (2-5). Key intermediates 2'-fluorinated acetals 12 and 13 were synthesized via stereospecific introduction of a 2'-phenylseleno moiety, fluorination, oxidative elimination, reduction, and acylation. With these intermediates, N-glycosylation reactions with bis(trimethylsilyl) nucleobases (5-fluorocytosine, thymine, 5-iodouracil, adenine) followed by desilylation afforded the target nucleosides, which were separated into cis- and trans-isomers via reverse-phase HPLC or silica gel chromatography. The anomeric configuration of the isomers was assigned using proton NMR, where α-anomers showed lower-field H-4' signals and higher-field H-5' signals compared to β-anomers. Acid stability assessment revealed that the 2'-fluorinated nucleosides were stable at pH 4.3, 7.1, and 9.1 for 1 week, with half-lives of approximately 21-22 days at pH 2.0 (significantly longer than β-L-FD₄C, which had a half-life of less than 30 min at pH 2.0). Biological evaluation in CEM-SS cells showed that among the eight nucleosides, the 5-fluorocytosine-bearing nucleoside 2a exhibited the best activity: its anti-HIV activity in HIV-infected CEM-SS cells was comparable to AZT, and its cytotoxicity was at least sevenfold lower than β-L-FD₄C. However, 2a was about 40-fold less potent than β-L-FD₄C against HBV. In summary, a general method for synthesizing cis- and trans-2'-vinylfluoro-containing L-nucleosides was developed, and 2a showed a favorable biological profile, although 2'-fluorination improved the cytotoxicity of β-L-FD₄C without enhancing its antiviral activity.