Optimization of Methods for Determination of the Encapsulation Efficiency of Doxorubicin in the Nanoparticles Based on Poly(lactic-co-glycolic acid) (PLGA)

Introduction. Drug encapsulation efficiency (EE) is the important parameter of the nanoparticle-based drug formulations. Generally, the methods for evaluation of the EE are based on separation of the free and NP-bound fractions of the drug; however, applicability of these methods for a particulate formulation needs careful consideration.

Aim. The purpose of the study was to optimize the procedure for evaluation of the EE for a nanoparticle-based drug formulation taking doxorubicin loaded in the PLGA nanoparticles (PLGA-Dox NP) as a model formulation.

Materials and methods. The PLGA-Dox NP were prepared by a “double emulsion” method at pH of the external aqueous phase of 7.4 or 6.4. The NP size and size distribution (PDI) were determined by photon correlation spectroscopy (PCS) and transmission electron microscopy (TEM). For the EE evaluation of doxorubicin, the NP were separated by centrifugation (CF), ultrafiltration (UF), or gel filtration. The efficiency of NP separation by the CF method was evaluated by the PLGA content in the supernatant by capillary electrophoresis (CZE).

Results and discussion. The average hydrodynamic diameter of the PLGA-Dox/7.4 and PLGA-Dox/6.4 NP (FCS) was 103 ± 10 nm and 141 ± 8 nm, respectively. According to the TEM data, the main fraction of the NP averaged 50 ± 16 nm. The EE of doxorubicin, determined after the NP separation by centrifugation at 48254.g, was 78.9 ± 1.8 % for the PLGA-Dox/6.4 and 91.5 ± 0.9 % for the PLGA-Dox/7.4 NP with the residual PLGA content in the supernatant of ~5 %. At lower acceleration the NP separation was incomplete leading to underestimation of the EE. Also, the ultrafiltration method using the filters with the NMWL of 50 and 100 kDa enabled the reliable NP separation with the minimal doxorubicin adsorption on the filter (<4 %). Separation of the NP by gel filtration led to the underestimation of the EE due to considerable desorption of doxorubicin from the NP surface.

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