We systematically investigate the pore structure of cellulose triacetate (CTA) and polyamide (PA) forward osmosis (FO) membranes with positron annihilation lifetime spectroscopy (PALS) and fractional rejection (FR) method to better understand the relationship between membrane pores (atomic-scale free volume) and separation performance. The results reported in this study suggest a range of membrane pore sizes and inner structures appropriate for the osmotic membrane process regardless of membrane materials. The PALS data suggest that the active layer of both FO membranes is mainly composed of generally uniform pores of larger size (0.29-0.30 nm) compared to seawater reverse osmosis (SWRO) (0.20-0.24 nm) membranes. The membrane pore size distribution (PSD) deduced by the FR method suggest that the PSD of both FO membranes are significantly shifted to smaller sizes under pressurized conditions, possibly due to the overall compression of the FO membranes. The effects of applied pressure on the structure and performance of active layer should be further investigated along with other properties such as thickness, swelling and sorption properties.