## 3 Results and Discussion
### Morphology and Particle Size of Insulin-Loaded MPs  
The morphologies of the MPs were studied by scanning electron microscope (SEM). Several studies have demonstrated that the morphology and pore size of matrixes intended for drug delivery play an important role in the control of the release kinetics [11]. SEM images of batches A1 and A2 revealed small nonspherical particles that clumped together and mixed with little needle-like crystals (Figure 2). The morphology of batch A3 was characterized by large lumpy crystals, while mixtures of lumpy and small crystals were identified in batches A4 and A5 (Figure 2). In general, the MPs showed rough porous surfaces and irregular shapes. Batches A1 and A2 had almost identical morphologies with agglomerated particles, while batches with higher concentration of PEG-4000 (i.e., batches A3-A5) showed a clearer distribution of small and large MPs.  
Figure 1: DSC thermograms of insulin-loaded PEG/mucin matrixes in superposition. Batch description: A0 = mucin/PEG/insulin (1:1:0), A1 = mucin/PEG/insulin (1:1:15), A2 = mucin/PEG/insulin (1:2:15), A3 = mucin/PEG/insulin (1:3:15), A4 = mucin/PEG/insulin (1:4:15), A5 = mucin/PEG/insulin (1:5:15). Note: mucin and PEG are measured in grams, while insulin is measured in milliliters.  
Moreover, particle size may be affected by one or more factors, including, among others, the degree of homogenization, homogenization speed, composition ratio, rate of particle size growth, and crystal habit of the particle [24]. During Ostward ripening, the particles tend to move towards one another, which leads to the formation of aggregates due to the kinetic advantage gain by larger crystals as a result of an increase in the concentration [23; 24]. In this study, the particle size was found to increase with increasing PEG/mucin ratio. Therefore, batch A5, having the highest PEG-4000/mucin ratio, displayed a significantly (\(p<0.05\)) higher particle size (\(398\pm 0.21\) m) as compared to batch A1 with a particle size of \(311\pm 0.22\) m, while unloaded MPs (batch A0) showed a particle size of \(276\pm 0.03\) m (Table 2). Thus, the particle size tends to increase after drug loading due to the intensification of interparticle attractive interactions (van der Waals forces) leading to particle aggregation.  
\begin{table}
\begin{tabular}{c c c c c} \hline \hline
**Batch 2** & **EE (\%)** & **DL (\%)** & **PS (\(\mu\)m)** & **RV (\%)** \\ \hline A0 & – & – & \(276\pm 0.03\) & \(66\pm 0.14\) \\ A1 & \(82\pm 0.12\) & \(18\pm 0.01\) & \(311\pm 0.22\) & \(75\pm 0.13\) \\ A2 & \(81\pm 0.11\) & \(26\pm 0.24\) & \(321\pm 0.13\) & \(80\pm 0.14\) \\ A3 & \(83\pm 0.23\) & \(28\pm 0.04\) & \(328\pm 0.11\) & \(83\pm 0.17\) \\ A4 & \(84\pm 0.16\) & \(35\pm 0.17\) & \(346\pm 0.05\) & \(86\pm 0.13\) \\ A5 & \(92\pm 0.12\) & \(39\pm 0.12\) & \(398\pm 0.21\) & \(89\pm 0.12\) \\ \hline \hline \end{tabular}
\end{table}
Table 2: Physicochemical parameters of insulin-loaded MPs (\(n=5\)) 1.  
Figure 2: SEM of representative batches of insulin-loaded microparticles (MPs) containing different ratios of mucin to PEG. Formulations: A1 = mucin/PEG/insulin (1:1:15), A2 = mucin/PEG/insulin (1:2:15), A3 = mucin/PEG/insulin (1:3:15), A4 = mucin/PEG/insulin (1:4:15), A5 = mucin/PEG/insulin (1:5:15). Control A0 corresponds to the formulation containing no insulin (mucin/PEG, 1:1). Each formulation is indicated in the top left corner of the corresponding microscopy image.