Over the past decades, limited strategies have been developed to study nanomechanical properties. The in-depth study on the nanomechanical properties of electrospun scaffolds can provide better insights at greater scales. Atomic force microscopy (AFM) nanoindentation is an authoritative method that can characterize mechanical properties. In this research, theoretical solubility studies of polyhydroxybutyrate (PHB) in different solvents were initially carried out. Then, three series of PHB-based mats were prepared, including PHB/lignin, PHB/cellulose nanofiber (CNF), and PHB/lignin/CNF. The theoretical modulus of samples was studied from two different standpoints, through the use of nanoindentation on a single nanofiber. First, scaffolds were considered as a series of short fiber reinforced nanocomposites (NCs). In the second viewpoint, the geometrical structure of each scaffold was considered similar to an open-cell foam (OCF). Consequently, modeling outputs verified that among NC models, isotropic Halpin-Tsai presented plausible predictions with '93.36 to 99.77% accuracy. In addition, Tetrakaidecahedron indicated more accurate predictions among OCF models for the mats ('78.99 to 96.15% adaption). Indeed, the theoretical NC and OCF models were able to provide acceptable forecasts for the application range of electrospun fibers. Based on this, nanoscale investigation of mechanical properties can turn a new page to estimate the further application of mats.