Changing mechanical properties of photopolymerized, dityrosine-crosslinked protein-based hydrogels

Abstract

Hydrogels based on renewable resources are a promising class of materials for future applications in pharmaceutics, drug delivery and personalized medicine. Thus, optional adjustments of mechanical properties such as swelling behavior, elasticity and network strength are desired. In this context, hydrogels based on the biological raw materials bovine serum albumin and casein were prepared by dityrosine-crosslinking of their tyrosine residues through visible light-induced photopolymerization. Changing the tyrosine accessibility by urea addition before photopolymerization increased the storage modulus of the hydrogels by 650% while simultaneously being more elastic. Furthermore, contributions of the buffer system composition, variation of protein concentration and storage medium towards mechanical properties of the hydrogel such as storage moduli, elasticity, fracture strain, compressive strength and relative weight swelling ratio are discussed. It could be shown, that changes in precursor solution and storage medium characteristics are crucial parameters towards tuning the mechanical properties of protein-based hydrogels.

Keywords: BSA—bovine serum albumin; casein; protein unfolding; protein-based hydrogels; urea; visible-light induced photopolymerization.

FIGURE 1

Schematic overview of the experimental workflow. Pre-processed protein, photoinitiator and co-factor stock solution were mixed to generate a precursor solution. Polymerization was achieved through illumination in a mold. Hydrogels were stored under different conditions and mechanical properties were determined either by oscillatory rheometry, swelling studies or uniaxial compression tests.

FIGURE 2

Characterization of hydrogels containing 100 mg/ml bovine serum albumin prepared in SPB. (A) Shear stress dependent (ω = 25 rad s⁻¹, 22°C) and (B) frequency-dependent [τ = 10 Pa, highlighted by a vertical line in (A), 22°C] oscillatory shear test of hydrogels prepared and stored in a 20 mM SPB at pH 8 without urea and with 3 M urea present in the precursor solution and storage medium. (n = 3) (C) Hydrogel storage modulus (G’) and loss factor (tan δ) in dependency of the urea content in the precursor solution and storage medium. (n = 3) Abbreviations: n.s, not significant, *p < 0.05.

FIGURE 3

Characterization of hydrogels containing 40, 60, 80, and 100 mg/ml bovine serum albumin prepared in MCB pH 7 with 0 or 4 M urea. All samples were stored in formulation buffer prior analysis. (A) Storage modulus (G’) and (B) loss factor (tan δ) determined by frequency-dependent oscillatory shear rheology. (n = 3) (C) Fracture strain (ε_max) and (D) compressive strength (σ_max) of those hydrogels determined by uniaxial compression tests. Abbreviations: n.s, not significant, *p < 0.05.

FIGURE 4

Characterization of hydrogels containing 100 mg/ml bovine serum albumin (pH 7 and 8) or casein (pH 6) prepared in MCB containing 0, 2 or 4 M urea. (A) Storage modulus (G’) and (B) loss factor (tan δ) as determined by frequency-dependent oscillatory shear rheology. All samples were stored in DPBS (n = 3) (C,D): Relative weight swelling ratio (m_rel) of BSA- and casein-based hydrogels stored in (C) DPBS or (D) MCB without urea at formulation buffer pH. (n = 3).