2.3. Hydrogel Characterization

The swelling ratio of the hydrogel at equilibrium was determined by measuring the weight of the hydrated gel and that of the dried gel. The hydrogels were immersed in DI water at room temperature and for 24 h to ensure the hydrogels were fully hydrated [16]. The degree of swelling (Q), defined as the reciprocal of the volume fraction of a polymer in a hydrogel (v₂), was calculated from the following Equation (1),

where ρ_s is the density of water, ρ_p is the density of polymer and Q_m is the swelling ratio, the mass ratio of swelled gel to the dried gel.

The average pore size (ξ) of hydrogel was calculated from the polymer volume fraction (v_2,s) and the unperturbed mean-square end-to-end distance of the monomer unit (r_o⁻²) using Equations (2) and (3):

where l is the average value of the bond length between C–C and C–O bonds in the repeatable unit of PEG [–O–CH₂–CH₂–], which is taken as 1.46 Å; M_c is the average molecular mass between cross-links in the network; M_r is the molecular mass of the PEG repeating unit; n is the number of repeat unit, which is taken as 7 (PEGDA M_n of 575 g/mol) and C is the characteristic ratio for poly(ethylene glycol), which is taken here as 4 [17].

The inner micro-structures of the hydrophobically-modified hydrogels were analyzed by SEM (FE-Scanning electron microscope, JEOL-7001F, JEOL Ltd., Tokyo, Japan) with an acceleration voltage of 5 kV. The hydrogels were lyophilized and covered with a platinum (Pt) layer produced by a vapor deposition. Values of the water contact angle (θ_w) on the surfaces of the hydrogels were measured by depositing a drop of water (4.0 μL) under atmospheric condition with DSA100 (KRÜSS, Hamburg, Germany). The contact angle was measured immediately after placing the water drop on the hydrogel surface [18]. Bovine serum albumin (BSA, Sigma) was also used as a model protein to evaluate the protein release rate from the hydrophobically-modified hydrogels [19]. The PEGDA and PPGMA stock solution were dissolved in DI water to prepare the pre-gelled solution at 10 wt %. The pre-gelled solution was mixed with BSA and photo-initiator, the mixture immediately cast between glass plates with spacers of 1 mm thickness, and exposed by UV lamp about 10 min. After 10 min, gel disks with a diameter of 8 mm were punched. Each hydrogels containing BSA was suspended in 400 μL of Dulbecco’s phosphate buffered saline (PBS, biowest) and incubated at 37 °C. At a designated time point, each PBS immersed hydrogel containing BSA was removed. Then, hydrogels were filled with fresh PBS again thereby keeping the sink condition. BSA amount released from the hydrogel was quantitatively measured using Pierce^TM BCA protein assay kit (Thermo Scientific, USA) according to the manufacturer’s instructions. The absorbance was measured at 562 nm using ELISA (Multiskan GO, Thermo Scientific, USA). The BSA release profile obtained was fitted to the Ritger-Peppas equation [20].

where M_t is the cumulative amount of protein released at the time, t; M_∞ is the total amount of protein in the hydrogels; k is the kinetic rate constant and n is the exponent related to the release mechanism.