Experimental Determination of Drug Diffusion Coefficients in Unstirred Aqueous Environments by Temporally Resolved Concentration Measurements

Abstract

The diffusion coefficient (also known as diffusivity) of an active pharmaceutical ingredient (API) is a fundamental physicochemical parameter that affects passive diffusion through biological barriers and, as a consequence, bioavailability and biodistribution. However, this parameter is often neglected, and it is quite difficult to find diffusion coefficients of small molecules of pharmaceutical relevance in the literature. The available methods to measure diffusion coefficients of drugs all suffer from limitations that range from poor sensitivity to high selectivity of the measurements or the need for dedicated instrumentation. In this work, a simple but reliable method based on time-resolved concentration measurements by UV-visible spectroscopy in an unstirred aqueous environment was developed. This method is based on spectroscopic measurement of the variation of the local concentration of a substance during spontaneous migration of molecules, followed by standard mathematical treatment of the data in order to solve Fick's law of diffusion. This method is extremely sensitive and results in highly reproducible data. The technique was also employed to verify the influence of the environmental characteristics (i.e., ionic strength and presence of complexing agents) on the diffusivity. The method can be employed in any research laboratory equipped with a standard UV-visible spectrophotometer and could become a useful and straightforward tool in order to characterize diffusion coefficients in physiological conditions and help to better understand the drug permeability process.

Impact of Mucin on Drug Diffusion: Development of a Straightforward In Vitro Method for the Determination of Drug Diffusivity in the Presence of Mucin

“…The analytical method developed by di [12] was employed to study the influence of mucin on the passive diffusion of drugs with different physicochemical characteristics (Table 1). Briefly, a conventional double beam VWR UV-6300 PC UV-visible spectrophotometer (VWR International, Radnor, PA, USA) was utilized, together with two Hellma ® Suprasil ® quartz absorption cuvettes with a chamber volume of 700 µL and path length of 10 mm (Sigma-Aldrich, Steinheim, Germany).…”

“…Thus, in the present study we investigated the applicability of a previously established method based on UV-visible localized spectroscopy [12] for the assessment of drug-mucin interaction, in order to experimentally quantify the relative diffusivity (D rel ) of drugs in mucin. This versatile and easy-to-implement method allows the measurement of very precise diffusivities (D, experimental error below 5% and fitting error below 1% [12]) for all chemical entities containing a chromophore by employing a standard UV-visible spectrophotometer. Empirical data are then analyzed by standard non-linear data fitting procedure in order to find diffusivity values.…”

Transient Incomplete Separation Facilitates Finding Accurate Equilibrium Dissociation Constant of Protein–Small Molecule Complex

“…[17] We created av irtual ACTIS setup in COMSOL Multiphysics software and used this virtual setup to simulate separagrams under conditions similar to realistic ones in an envisioned proof-ofprinciple experiment. [18] Thevalues of l and Q in avirtual setup were 50 times smaller than in ar eal one to reduce computational time from hours to minutes.T his reduction was possible given scalability of these parameters and the results,a sl ong as Equation (3) is satisfied. (3)].…”

“…Weused m L % 500 mm 2 s À1 for L as asmall-molecule drug. [18] Thevalues of l and Q in avirtual setup were 50 times smaller than in ar eal one to reduce computational time from hours to minutes.T his reduction was possible given scalability of these parameters and the results,a sl ong as Equation (3) is satisfied. [19] Ther emaining parameters (diffusion coefficients,r ate constants,c oncentrations,a nd capillary radius) were set at realistic values.…”

Transient Incomplete Separation Facilitates Finding Accurate Equilibrium Dissociation Constant of Protein–Small Molecule Complex

“…(3)]. We used μ L ≈500 μm 2 s −1 for L as a small‐molecule drug . The values of l and Q in a virtual setup were 50 times smaller than in a real one to reduce computational time from hours to minutes.…”