## 3 Results and Discussion
### In Vivo Antidiabetic and Toxicological Studies  
Alloxan was used to induce a hyperglycmic condition in the male rats. Alloxan is a diabetogenic chemical substance that inactivates the \(\beta\) cells of the pancreases, eliciting insulin deficiency without affecting other islet types [30]. The results of the blood glucose reduction test, presented in Figure 4, indicate that insulin-loaded MPs (Group 1) displayed a more pronounced and significant (\(p<0.05\)) blood glucose decrease, from 100 mg/dL to 48 mg/dL post-administration, compared to other samples. This is an indication that the MPs were able to provide protection to the insulin and effectively deliver it to the absorption site in vivo. A dose of 50 IU/Kg has been reported to have a similar effect to a higher dose (100 IU/Kg) with a minimum blood glucose level of 55%, and is thus taken as an effective therapeutic dose to saturate the insulin absorption sites [31]. Group 2, which received pure insulin by oral administration, showed minor hypoglycemia when compared to the group receiving  
Figure 3: Insulin release profiles at pH 7.4 (**a**) and at pH 1.2 (**b**). Formulations A1–A5: 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). Data represent mean values \(\pm\) SD (\(n=3\); \(p<0.05\)). Note: mucin and PEG are measured in grams, while insulin is measured in milliliters.  
insulin-loaded MPs, indicating that no significant absorption occurred in this group. This small hypoglycemia effect may be due to a small fraction of insulin being directly absorbed through the intestinal wall [32]. Such direct insulin absorption occurs at specific insulin receptors located in intestinal enterocytes [33] and undergoes rapid internalization by the epithelial cells through the interstitial space, ending up in the blood stream. The upper intestinal area has been reported to be the most active area of insulin absorption during fasting conditions [30]. Furthermore, no apparent blood glucose reduction was observed in the groups that received water (negative control, Group 4) or placebo (Group 5). In contrast, Group 3, which received a subcutaneous insulin injection, showed the highest glucose reduction level 2 h after administration. After this time, the glucose level increased again beyond normal levels, unlike in Group 1, where the level was maintained after reaching the highest glucose reduction. This result is an indication that subcutaneous insulin will require multiple doses to stabilize the patient within 12 h.  
Toxicological evaluations of the formulations were based on enzymes such as aspartate transaminase (AST) or serum glutamic oxaloacetic transaminase (SGOT), alanine transaminase (ALT) or serum glutamic pyruvic transaminase (SGPT), and alkaline phosphatase (ALP). These enzymes are found mainly in the liver, red blood cells, heart, kidneys, and pancreas, being useful biomarkers for the diagnosis of injured body tissues, particularly in organs such as heart and liver [34; 35]. It has been demonstrated that the more AST and ALT are released into the bloodstream, the more tissues are damaged [36]. The results obtained for our formulations showed that the quantities of these enzyme markers (i.e., SGPT, SGOT, ALP) in the serum were within the established reference values (Table 3). In addition, no significant variation from the control batch (water) was observed. Overall, these values suggest that the constituents of the insulin-loaded MPs are safe with no hepatotoxic effects.  
Figure 4: Percentage decrease of the blood glucose level in rats treated with insulin-loaded MP formulations (\(n=5\)). Groups G1–G5: G1 = received insulin-loaded MPs (formulation A5), G2 = received pure insulin solution, G3 = received subcutaneous insulin, G4 = received water, G5 = received unloaded MPs (batch A0). Data represent mean values \(\pm\) SD (\(n=3\)).