The free-volume of a matrix is a fundamental parameter that relates to its molecular and bulk characteristics, such as crystalline change and glass transition behavior. In starch-based bionanocomposite, we investigated the effect of the addition of montmorillonite nanoclay (MMT) and food plasticizers (glycerol and sorbitol) on changes of molecular pore size (including pore volume and pore distribution) using the Positron Annihilation Lifetime Spectroscopy (PALS) method. The results demonstrated counter-intuitive impact of MMT on the total free-volume where the total free-volume increased within the polymeric matrix. When compared to the pure matrix free-volume, the addition of MMT also resulted in the appearance of a broader distribution of the void sizes. The plasticizers, on the other hand, apparently occupied the void spaces, and therefore decreased the free-volume of the matrix. Further, together with the small angle X-ray scattering (SAXS) analysis, we concluded that this is a result of interplay between the plasticizer-plasticizer interactions and the polymer-plasticizer interactions. For example, in the starch/glycerol/MMT system, the pore radii slightly decrease upon the increasing of glycerol amount (OG210 = O.27 nm and OG220 = 0.26 nm), but the relative weight did increase with the increasing glycerol concentration. However, increasing the sorbitol amount increased the pore size from 0.23 nm(OS210) to 0.28 nm(OS220). Furthermore, the addition MMT in the OS010 system, promote the emergence of a new dateable pore radius(0.90 nm), and the total weight significantly increased from 13.70 (OS010) to 19.5% (OS210). We suggest that the pore variation (size and distribution) due to the MMT and plasticizers are reflected in the polymer glass transition and crystallinity because ultimately, the, total free-volume is a reflection of level of interactions existing within the bulk of these nanocomposites.