Effect of pullulan nanoparticle surface charges on HSA complexation and drug release behavior of HSA-bound nanoparticles

Abstract

Nanoparticle (NP) compositions such as hydrophobicity and surface charge are vital to determine the presence and amount of human serum albumin (HSA) binding. The HSA binding influences drug release, biocompatibility, biodistribution, and intercellular trafficking of nanoparticles (NPs). Here, we prepared 2 kinds of nanomaterials to investigate HSA binding and evaluated drug release of HSA-bound NPs. Polysaccharides (pullulan) carboxyethylated to provide ionic derivatives were then conjugated to cholesterol groups to obtain cholesterol-modified carboxyethyl pullulan (CHCP). Cholesterol-modified pullulan (CHP) conjugate was synthesized with a similar degree of substitution of cholesterol moiety to CHCP. CHCP formed self-aggregated NPs in aqueous solution with a spherical structure and zeta potential of -19.9 ± 0.23 mV, in contrast to -1.21 ± 0.12 mV of CHP NPs. NPs could quench albumin fluorescence intensity with maximum emission intensity gradually decreasing up to a plateau at 9 to 12 h. Binding constants were 1.12 × 10(5) M(-1) and 0.70 × 10(5) M(-1) to CHP and CHCP, respectively, as determined by Stern-Volmer analysis. The complexation between HSA and NPs was a gradual process driven by hydrophobic force and inhibited by NP surface charge and shell-core structure. HSA conformation was altered by NPs with reduction of α-helical content, depending on interaction time and particle surface charges. These NPs could represent a sustained release carrier for mitoxantrone in vitro, and the bound HSA assisted in enhancing sustained drug release.

Figure 1. Schematic illustration for synthesis of CHCP conjugate.

Figure 2. Infrared spectra of (a) pullulan, (b) CEP, and (c) CHS.

Figure 3. Nuclear magnetic resonance spectra of (a) pullulan, (b) CEP, and (c) CHCP.

Figure 4. Size distribution, zeta potential, and transmission electron microscope images of (a) CHCP nanoparticles and (b) cholesterol-modified pullulan (CHP) nanoparticles.

Figure 5. The fluorescence spectra of human serum albumin (HSA, 1.5×10⁻⁵ mol/L) (A) in the absence and (B) presence of the mixture of CHCP, and (C) CHP with the same concentration (4.2×10⁻⁶ mol/L).

Figure 6. Emission intensity of HSA upon interaction with CHCP nanoparticles (—○—) and CHP nanoparticles (—□—) at 342 nm as a function of time.

Figure 7. Fluorescence spectra of HSA (1.5×10⁻⁵ mol/L) in the presence of (A) CHP and (B) CHCP with different concentrations: (a) 0, (b) 2.07×10⁻⁷, (c) 3.31×10⁻⁷, (d) 4.14×10⁻⁷ , (e) 8.28×10⁻⁷, (f)20.7×10⁻⁷, and (g) 41.4×10⁻⁷ mol/L.

Figure 8. Plots (n = 6) of Fo/(Fo −F) versus 1/[CHP] (—▪—) and 1/[CHCP] (—•—).

The concentration of HSA was 1.5×10⁻⁵ mol/L.

Figure 9. CD spectra of HSA in the (a) without (b and c) with CHP and CHCP nanoparticles in solution at 37°C.

Figure 10. The release of mitoxantrone in phosphate buffered saline at 37°C in vitro (□, free mitoxantrone; ○, CHP; ◊, CHCP; ▹, HSA-mitoxantrone; ▽, CHCP-HSA; Δ, CHP-HSA).

Figure 11. Effect of HSA complexation on drug released from nanoparticles.