In this work, the effect of ascorbic acid on luminescence of porous silicon nanoparticles is studied. Porous nanoparticles with a multi-sized distribution are produced from a porous silicon free standing film and filtered through a 450 nm membrane. The highly luminescent and water-soluble nanoparticles are dispersed in deionized water. The stability of photoluminescence signal and the dimensions of porous silicon nanoparticles are characterized before adding ascorbic acid into the porous nanoparticles solution. The increasing of photoluminescence, before stabilization, has been correlated to oxidation of nanoparticles in water, evaluated by the technique of Attenuated Total Reflection. Once the nanoparticle photoluminescence signal is stabilized, ascorbic acid is loaded into the solution and its presence and real absorption onto the porous silicon nanoparticles are evaluated from their emission quenching. The quenching of photoluminescence is studied with different concentrations of ascorbic acid by Stern–Volmer plot for confirming the quenching law. This study shows that luminescence quenching of porous nanoparticles could be used for determining the presence of ascorbic acid in a solution.