In this study, in vitro cytotoxic evaluation was carried out for a hierarchical web of carbon micronanofibers applied as an adsorbent and a support for metal catalysts for several environmental and biological applications. The web was prepared by growing carbon nanofibers (CNFs) on substrate-activated carbon microfibers (ACFs) by chemical vapor deposition (CVD) using Cu/Ag nanoparticles as the catalyst. The cytotoxicity of the prepared ACF/CNF was ascertained with respect to human lymphocytes using MTT assays and fluorescence microscopy analysis. The DNA damage was assessed using DNA profiling. The tests were also carried out for several common commercial carbonaceous and noncarbonaceous adsorbents, such as carbon nanotubes (CNTs), granular activated carbon (GAC), silica, alumina, and zeolite, for comparison. This study reveals that the cytotoxicity of the ACF/CNFs is comparable to or less than that of the commercial adsorbents. Therefore, the ACF/CNFs prepared in this study may be safely used in environmental and biological applications.