The PZS-HGMC were measured for diameter with a micrometer screw gauge, and the mean was then calculated (Zeeb et al. 2015).

The SW% of the HGMC was investigated via the immersing method (Anal and Stevens 2005) in detail. The SW study was conducted in a water bath shaker at 37±0.5 °C. Previously weighted HGMC were immersed in 0.1N HCl for two hours, resulting in the HGMC swelling. The swollen HGMC were wiped with filter paper to eliminate excessive buffer and then weighed. All significant shifts in weight were recorded. The SW% was calculated by the following formula (Taher et al. 2019):

$\text{SW}\%=\frac{\text{Weight of swollen MCs}-\text{Weight of dry MCs}}{\text{Weight of dry MCs}}\times 100$

The drug content of the PZS-HGMG was directly measured. The PZS-HGMG (200 mg) was carefully weighed and crushed. The PZS-HGMG were mixed with deionized water (10 mL) for a full day at 25±0.5 °C. Then, the solution was filtered through a nano-filter syringe. The PZS concentration was detected spectrophotometrically using the UV-visible spectrophotometer (Shimadzu, Japan) at 283 nm. The EE% was detected using the following equation (Umaredkar et al. 2020):

$(\%\mathrm{EE})=\frac{\text{RA}}{\text{TA}}\times 100$

RA and TA are the resulting and theoretical amounts of PZS found in the PZS-HGMG, respectively. The results were repeated in triplicate.

The morphologies and surface of the PZS-HGMG were investigated using SEM (Agilant 8500 FE, Japan) (Thanos et al. 2006). PZS-HGMG samples were covered with gold and photographed at a voltage of 30 kV before being placed on aluminum stubs for examination.

Before submitting samples for FTIR evaluation, a pestle and mortar were used to reduce the PZS-HGMG to a powder. The FTIR probe examined the sample (JASCO FTIR Model 6600, Tokyo, Japan). The range was selected from 400 to 4000 cm^-1 with 95% transmittance. Interactions can be detected, and functional group alterations can be identified using FTIR spectral analysis (FTIR spectrophotometer at a scanning number of 8 with the KBr sampling method) (Moghaddam et al. 2015).

Some differential scanning calorimetry (DSC) studies were applied using DSC 60 TASYS Software (Shimadzu, Japan). The dry MCs used in the DSC trial varied from 3 to 7 mg. The sample was analyzed in a nitrogen flow with a 25–300 °C temperature, and the scan rate was 10 °C/min. The DSC detects crystallinity changes, melting points, and drug-excipient-adsorbent interactions (Honary et al. 2009).

The resulting PZS-HGMG had diameters ranging from 1.3 to 2.5 mm by vernier caliper (Gulia et al. 2023), as displayed in Table 3.

Table 3 illustrates the swelling behavior of the PZS-HGMG at pH 1.2; the obtained SW% ranges from 410.0% to 840.0%.

The PZS-HGMG showed swelling, resulting in MC with a small amount of SAT bursting after two hours of testing. The Na⁺ ion in the used buffer and the Ca²⁺ ion in the cavity of Ca-alginate microcapsules are involved in the ion exchange mechanism (Elnashar et al. 2010). One possible cause of swelling involves the exposure of the dry PZS-HGMG to water, and the hydrophilic SAT and CTN groups caused them to swell (Khan et al. 2023).

The EE% was 87.0 to 94.0%; the greater EE% of the SAT–CTN microcapsules may be attributed to the more porous structure (Dhurke et al. 2013). Since Ca⁺² and -NH₃⁺ of CTN compete, they can react with –COO– of SAT (Akyuz et al. 2017). The estimated values of EE are illustrated in Table 3.

By fixing the effects of one parameter, one can examine the effects of the other two parameters with PS; thus, by holding the CTN concentration constant, SAT has a direct relationship with PS, and CaCl₂ did not affect PS (Fig. 1A).

Figure 1. 

Response surface graphs show variables’ effects on (A, B, C) particle size.

By holding the CaCl₂ concentration constant, PS appeared to be directly associated with SAT, while CTN did not affect PS (Fig. 1B).

By holding the SAT concentration constant, both CTN and CaCl₂ have a direct relationship with PS (Fig. 1C).

The overall adjusted R² of the model was 99.11%, with SAT showing 84.75% contribution of the final model (p-value < 0.001), with SAT (r = 0.5, p-value < 0.001), CaCl₂ (r = -0.0625, p-value = 0.006), SAT*SAT (r = -0.2, p-value < 0.001), CaCl₂*CaCl₂ (r = 0.175, p-value < 0.001), and SAT*CaCl₂ (r = 0.075, p-value = 0.012) are the significant coefficients in the final model:

Particle size (mm) = 2.787 + 1.038 Alginate − 1.338 CaCl2 − 1.437 Chitosan − 0.2000 Alginate * Alginate + 0.1750 CaCl2 * CaCl2 + 0.625 Chitosan * Chitosan + 0.0750 Alginate * CaCl2 + 0.1250 Alginate * Chitosan + 0.2500 CaCl2 * Chitosan

By fixing the effects of one parameter, one can examine the effects of the other two parameters with SW%; thus, by holding the CTN concentration constant, SAT has a direct relationship with SW%, and CaCl₂ did not affect SW% (Fig. 2A).

Figure 2. 

Response surface graphs show variables’ effects on (A, B, C) swelling%.

By holding the CaCl₂ concentration constant, SW% appeared directly associated with SAT, while CTN did not affect SW% (Fig. 2B).

By holding the concentration of SAT constant, both CTN and CaCl₂ have a direct relationship with SW% (Fig. 2C).

The overall adjusted R² of the model was 97.0%, with SAT showing a 78.92% contribution to the final model (p-value < 0.001), with SAT (r = 160.63, p-value < 0.001), and SAT*SAT (r = 108.1, p-value < 0.001) as the significant coefficients in the final model:

Swelling (%) = 509 − 313.1 Alginate + 113.1 CaCl2 − 378 Chitosan + 108.1 Alginate * Alginate − 16.9 CaCl2 * CaCl2 + 422 Chitosan * Chitosan + 3.8 Alginate * CaCl2 + 100.0 Alginate * Chitosan - 0.0 CaCl2 * Chitosan

By fixing the effects of one parameter, one can examine the effects of the other two parameters with EE%. By holding the CTN concentration constant, SAT and CaCl₂ have a direct relationship with EE% (Fig. 3A).

Figure 3. 

Response surface graphs show variables’ effects on (A, B, C) encapsulation efficiency (%).

Holding the CaCl₂ concentration constant, EE% appeared to be directly associated with SAT and CTN (Fig. 3B).

By holding the concentration of SAT constant, both CTN and CaCl₂ have a direct relationship with EE% (Fig. 3C).

The overall adjusted R² of the model was 87.13%, with SAT showing an 82.77% contribution to the final model (p-value < 0.001), with SAT (r = 2.25, p-value < 0.001), and CaCl₂ (r = 0.625, p-value = 0.046) as the significant coefficients in the final model:

Encapsulation efficacy (%) = 82.75 + 1.88 Alginate + 2.62 CaCl2 − 4.38 Chitosan − 0.000 Alginate * Alginate − 0.250 CaCl2 * CaCl2 − 6.25 Chitosan * Chitosan − 0.250 Alginate * CaCl2 + 3.75 Alginate * Chitosan + 0.00 CaCl2 * Chitosan

The concentration of SAT increased, and the diameter increased (Fig. 1A, B). This result is compatible with the outcomes of (Patil et al. 2012). The model graphs indicated that SAT significantly impacts the SW of various formulations (Fig. 2A, B). When the content of SAT grew, the formulations swelled, as represented by (Swain et al. 2014), who discovered a relation between the rising swelling and diameter. Model graphs demonstrated that SAT significantly impacts the EE% of different formulas. Fig. 3A, B show that, as the content of SAT increased, the EE% of different formulae also increased. This is consistent with the results of (Babu et al. 2011), who found the same outcomes: when the SAT was raised, the EE% was significantly raised in different formulas.

CaCl₂ change, when considered in terms of change in SAT, showed no significant effect, which indicates SAT has more effect on the PZ at all concentrations of CaCl₂; however, the effect on CaCl₂ change when examined against the change in CTN showed variability, in which a larger PS was achieved with low concentrations of CaCl₂ and higher concentrations of CTN and the lowest PS was achieved with medium concentrations of CaCl₂ and the lowest concentrations of CTN. It is evident from the model graphs displayed in Fig. 1A, C that, with increasing CaCl₂ concentration, the response surface shifts from a higher diameter value to a smaller one.

Dissolution of the PZS-HGMG formulated in a low CaCl₂ solution occurred sooner than in PZS-HGMG formulated in CaCl₂ solutions of higher concentrations, and the maximum SW ratios rose with the increase in CaCl₂ concentration at pH 1.2. The concentration of Ca²⁺ during the formulation of PZS-HGMG may improve their crosslinking density. This observation is consistent with those of (Laelorspoen et al. 2014).

It is blatantly obvious in Fig. 1G, I that the EE% was raised when the CaCl₂ content was raised due to the crosslinking reaction being favored (Ghumman et al. 2018).

As CTN concentration increases, the resulting formulation’s diameters rise (Wang et al. 2017). A thicker CTN layer may increase the size of the formulations. This trend lines up with a rise in the mean diameters of the PZS-HGMG, as shown in Fig. 1C.

The SW% index grew dramatically with rising CTN concentrations at pH 1.2. The raised concentration of CTN led to a raised SW% of different formulae (Fig. 2C). This observation is consistent with previous studies (Sinha et al. 2018).

The impacts of CTN concentration on the diameter and EE% of different formulas may explain why a reduction in CTN concentration led to a decline in EE%. The results are shown in Fig. 2C and are consistent with another study (Mi et al. 2013). By elevating the concentration of CTN in the coagulation solution, the density of the resulting membrane is increased as more SAT-CTN ionic interactions are created, resulting in a less perforated gel formation and a higher EE%; this might be because the medicine is less likely to escape from the PZS-HGMG during encapsulation.

Macroscopic evaluation of the examined rats’ gastric tissues is shown in Fig. 8. The negative control group’s gastric tissues (Fig. 9A) exhibited healthy mucosa, minimal redness, and no bleeding or regions of hyperemia. On the other hand, the injured rat groups’ (G2 and G3) showed extensive damage to the stomach mucosa (Fig. 9B, C).

Figure 9. 

The appearance of the mucosa in the stomachs of IMN-induced rats: A. Negative control group; B. Positive control group; C. Blank MC-treated group; D. Free PZS-treated group; E. PZS-HGMC-optimized formula-treated group.

When given orally (30 mg/kg), IMN caused severe bleeding sores, most of which were in the gastric glandular mucosa and very few, if any, in the antrum (Yusif et al. 2016). Most of the gastric body was discolored dark reddish from lesions, and the gastric mucosa was riddled with multiple ulcer regions of varying shapes and sizes. The gastric mucosa showed considerable hyperemia.

There were fewer stomach ulcers after administering free PZS to the rats (Fig. 9D). The lesions emerged as discrete lesions in the gastric mucosa, and the epithelial injuries were modest compared to those seen in erythematous control animals (Özgüney 2011). The incision was not observed in any of the medicated rat groups.

On the other hand, medication with a PZS-HGMC optimized formula (Fig. 9E) exhibited an almost typical stomach look, with only a little ulceration and no redness or bleeding.

During the study, the rats in the medicated groups were healthier and behaved normally. The results showed that the medication of animals with PZS-HGMC provided more protection against PUC than medication with free PZS. This could be because the medicine is released slowly over time, retaining a potent concentration of the PZS for a longer period.

The variations in the gastric tissues of the tested rat groups were confirmed through histopathological evaluation. Fig. 10 shows the histopathological analysis of various groups. The gastric histology of the control normal animals (Fig. 10A), which comprise the negative control group (G1), displays a typical glandular structure and function at the microscopic level.

Figure 10. 

Histological examination of gastric tissues from the INM-induced lesion pattern in rats: A. Normal group; B. INM-treated control group; C. Blank MC-treated group; D. Free PZS-treated group; E. PZS-HGMC-optimized formula-treated group.

Eccentric nuclei and pale eosinophilic vacuoles in the cytoplasm characterize the parietal cells that make up the top half of the stomach. On the contrary, the IMN-medicated positive control group (G2) (Fig. 10B) shows a prominent inflammatory response characterized by subepithelial edema. The heavily muscled coat was broadened and somewhat hyalinized, and the subepithelial blood arteries were clogged with plaque.

These findings demonstrated the chronic inflammation and acute stage of the lesion. The findings also showed the gastric tissue to be highly inflamed for the INM-medicated group (G3) receiving the blank MC (Fig. 10C). The gastric tissue of (G4), receiving the free PZS, exhibited moderate lesions (Fig. 10D). In (G5), receiving the PZS-HGMC optimized formula (Fig. 10E), the rats had the greatest morphological preservation. They exhibited regular epithelial and lymphatic organization, distribution, and excellent tissue morphology.