The engineering of biomaterials provides an opportunity for developing new polymeric composites. In this context, the objective of this work is to produce and evaluate fiber obtained from mixtures of corn straw waste and polylactic acid (PLA)in vitro. Treated corn straw fiber (TCSF) was made by cutting corn straw waste and applying alkali, acid, and silane treatments. Corn straw fiber (CSF) or TCSF was combined with PLA or modified PLA (MPLA) in an electrospinning technique, to manufacture a 100-500 nm nanofiber mat. We studied morphology, mechanical properties, water resistance, biodegradability, and cytocompatibility properties of the electrospun PLA/CSF and MPLA/TCSF nanofibers. The resulting tensile properties and morphological characterizations indicated enhanced adhesion between TCSF and MPLA in the nanofiber, as well as improved water resistance and tensile strength, compared to PLA/CSF nanofiber. Regarding biocompatibility, only viable cells were found for all of the compositions, and the biocompatibility of the materials was validated by morphological analysis of cultured cells. Morphology proliferation assays showed that human keratinocyte cells grow well on the surfaces of PLA, MPLA, and their composite nanofibers. The weight loss of PLA/CSF and MPLA/TCSF composites buried in soil indicated that both were biodegradable, especially at high levels of CSF or TCSF substitution.