An experimental methodology for the determination of the obstruction factor in the expression for mesopore diffusion in Zorbax Eclipse Plus C18 reversed-phase particles is proposed . The method uses peak parking experiments conducted on particles that were previously stripped of their stationary phase by flushing the column with trifluoroacetic acid at a temperature of 60°C . Further using pure organic solvents as the mobile phase, any potential retention or surface diffusion effect is omitted . To avoid interference between the parked peaks and baseline disturbances typically occurring when switching on and off the flow, peak parking experiments were carried out in a set-up wherein two identical columns were used in parallel . This set-up allowed to maintain the flow through the detector at all times, by redirecting the flow from one column to the other during the peak parking experiments . Several tracer molecules (ionic and deuterated tracers) were compared and it was found that the use of deuterated molecules provides the best possible coverage of the accessible space of the mesopore volume . Interpreting the peak parking responses obtained with these tracers with a model based on the effective medium theory (EMT) subsequently provided an estimate of the value of the mesopore diffusion obstruction factor Î³mp . Taking the well-established pore hindrance factor F (λ) correction into account, the obtained experimental Î³mp-values are more in agreement with the tortuous and constricted diffusion paths one can expect in the void space within a structure resembling a monolithic skeleton with tetrahedral connectivity rather than in the void space formed by a packing of nanospheres . This is also more in line with the measured internal porosity values lying around Îµpz=0.5, whereas a packing of nanospheres would rather correspond to an Îµpz of 0.4 . As such, the presented protocol provides a means to infer the internal mesopore structure of reversed-phase particles.