3.1. Thermal Degradation Kinetics

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where C is the concentration of CP (µg/mL), t is the incubation time (min), k is the degradation rate constant (min−1 ), and n is the order of the reaction where n = 0 is zero order, and n = 1 is first order. The order of drug degradation was determined using graphical methods. The remaining concentration of CP and the natural logarithm remaining CP were plotted versus time for prediction of zero order and first order degradations, respectively. The results are shown in Figure 2a,b. The linear regression was added to determine the correlation coefficient (r 2 ). As presented in Table 1, the obtained r 2 values from the first-order reaction plots were close to 1, suggesting that the thermal degradation of CP followed the first-order kinetics. The kinetic parameters obtained from fitting the first-order kinetics model are shown in Table 2. The results demonstrate that the elevated temperature could significantly cause an increase in the degradation rate of CP. The results also confirm that CP in CP-F possessed significantly higher stability than in CP-P and CP-W (p < 0.05). 

The acceleration effect of temperature on the rate of chemical reactions is generally described by the Arrhenius equation [29], which is the relationship between the rate constant and temperature, as shown in Equation (7):

where k is the reaction rate constant of first-order kinetic (min−1 ), A is the frequency factor, Ea is the activation energy (cal mol−1 ), R is the gas constant (1.987 cal mol−1K−1 ), and T is the absolute temperature in degrees, Kelvin. The determination of the Arrhenius parameter is based on a plot of the natural logarithm of k against the reciprocal of absolute temperature (1/T). Estimation of the appropriate rate or rate constant for CP degradation is an important step in predicting the stability of CP in each formulation. From the results, the Arrhenius plots provide a good description of CP degradation, as is evident from the linearity (r 2 = 0.99) of the plots as seen in Figure 2c for all formulations. The Ea value for CP degradation in each formulation was calculated according to the Arrhenius plots. It was found that the Ea value of CP in CP-F was higher than that in CP-P and CP-W, with values of 33.06 ± 0.83, 17.01 ± 0.69, and 11.87 ± 0.49 kcal/mol, respectively. The results suggested that the activation energy for CP degradation in the nanofibrous film is approximately two times higher than CP in the polymer solution and three times higher than CP in the water solution. These results show the high potential of the nanofibrous film for the protection of CP from thermal degradation. 

3.2. Degradation Kinetics of CP by UV Light

The evaluation of the photostability of the drugs and the formulations is an essential issue for formulation development. Tooth-whitening agents such as hydrogen peroxide and CP are photosensitive agents [30,31]; therefore, their formulated products may degrade during manufacturing and storage. In the present study, the photostability tests of the formulations were carried out under UV light. As shown in Figure 3a, after the samples were exposed to UV light for 1 h, CP-F showed a higher proportion of CP remaining than CP-P and CP-W. The CP content of CP-P and CP-W significantly decreased when compared to the initial measurement (p < 0.05), whereas that of CP-F was not significantly different from the initial measurement. The CP remaining of all formulations showed a signifificant difference from the initial measurement (p < 0.05) after 4 h exposure to UV light. 

The degradation profiles of CP under UV light exposure are shown in Figure 2. Plotting the data according to the first-order degradation, the linear relationship was obtained as shown in Figure 3b, with the r 2 close to 1, as shown in Table 3. From these results, the reaction rate constant of CP in CP-F was shown to be significantly lower than that in CP-W and CP-P. The results demonstrate that nanofibrous film prevented the degradation of CP from UV light. It is also considered that the solid dosage form has UV protection properties significantly greater than the liquid, e.g., solution, dosage form. 

3.3. Long-Term Stability of CP-F

According to World Health Organization [32], the recommendation of the testing condition for the long-term stability of products was 25 ± 2 °C/60 ± 5% RH or 30 ± 2 °C/75 ± 5% RH for a minimum period of 12 or 6 months, respectively. In this present study, the long-term storage was conducted at an average temperature of 25 ± 2 °C for 12 months. To compare the effects of temperature and humidity, a high temperature of 45 ◦C was used. A humidity of 75% was selected from climatic zones IV and compared with the low humidity of 30%. Thus, the storage conditions of 25 ◦C/30% RH, 25 ◦C/75% RH, and 45 ◦C/30% RH were used for 12 months. The changes in physicochemical properties, i.e., color, morphology, internal structure, molecular interaction, mechanical properties, and mucoadhesive properties were investigated. The gradual changes in CP content in CP-F kept at the three conditions were also determined. 

3.4. Color Changes after Long-Term Storage 

The color parameters of CP-F investigated by the colorimetric measurements are shown in Table 4. Initially, CP-F was white as a high value of L* was obtained. For the degree of green−red, CP-F was achromatic as the* value was close to 0, and for the degree of blue−yellow, CP-F was slightly blue as a negative b* value was presented. The differences between the L*, a*, and b* values of CP-F stored at 25 ◦C/30% RH were not signifificant, indicating that the color of CP-F kept at 25 ◦C/30% RH for 12 months was not changed. The L* value of CP-F stored at 45 ◦C/30% RH was the lowest compared to other conditions, indicating a signifificant decrease in the lightness of the samples. CP-F kept at 25 ◦C/75% RH and 45 ◦C/30% RH showed a high negative a* value and a high positive b* value, indicating the green and yellow of this sample were changed. It has been reported that the nanofibrous films containing PVA can change into yellow or brown after thermal decomposition [33]. Therefore, the color change of CP-F was probably due to the decomposition of PVA which was present in the films. Among various storage conditions, no signifificant differences in the ∆E values were detected. As the ∆E value was lower than 3, the color change was difficult to distinguish by human eye perception [34].

3.5. Morphology Changes after Long-Term Storage 

The SEM images of surface morphology and the diameter of the nanofibers in the nanofibrous films before and after keeping at the test storage conditions are presented in Figure 4. Initially, CP-F exhibited a smooth fibrous structure with a diameter range in nanosize, without any undesirable parts. After storage at 25 ◦C/30% RH for 12 months, CP-F showed slight defects of a straight line. However, no signifificant difference in average diameter was observed. The average diameters of 237 ± 57 and 267 ± 72 nm were found for CP-F at initial measurement and after storage at 25 ◦C/30% RH for 12 months, respectively.

After storage at 25 ◦C/75% RH for 12 months, CP-F exhibited different morphology from the initial measurement, and the fibrous structure was changed. Nanofibers fused and the structure of the fibers in nanosize was almost absent. The nanofibers could not keep the original structure. These phenomena also happened with CP-F after storage at 45 ◦C/30% RH for 12 months. This CP-F showed the merging of the nanofibers. It was found that the remaining fibers showed discontinued and tear line structure. From these results, we considered that high temperature and high humidity are the important factors that affect the morphology of CP-F. It has been reported that high temperatures can destroy the PVA-based nanofibrous film [35]. In the present study, the main composition of the nanofibers of CP-F was PVA and PVP and the results revealed that CP-F, after being subjected to high temperatures, became brittle, and the structure of the nanosized fibers was absent.

3.6. Internal Structure Changes after Long-Term Storage

The XRD patterns of intact CP and CP-F before and after storage are displayed in Figure 5. Intact CP exhibited sharp identical peaks at 14◦, 23◦, and 28◦, indicating that the internal structure of CP was a crystalline form. The crystalline peaks of CP were absent in the XRD pattern of the freshly prepared CP-F. The disappearance of the CP crystalline peaks indicated that the drug was well incorporated in the nanofibrous film by electrospinning technique, and CP was converted from a crystalline state to an amorphous state. This halo pattern was also found in CP-F after storage at 25 ◦C/30% RH for 12 months, indicating that CP recrystallization did not occur during a long-term period of storage in this condition.

However, the XRD patterns of the stored CP-F under 25 ◦C/75% RH and 45 ◦C/30% RH showed high identical crystalline peaks of CP at 22◦ and 25◦. Moreover, the XRD peaks at 46◦, 49◦, and 50◦of CP-F after storage at 45 ◦C/30% RH were of higher intensity than that of CP-F storage at 25 ◦C/75% RH. These peaks are possibly related to urea as they resemble the peak patterns of urea powder. Generally, CP dissociated into hydrogen peroxide and urea [36]. The degradation products of hydrogen peroxide are oxygen and water [37] and these products may be lost during storage. Urea was the degradation product that remained in the formulation. The amorphous urea had partially recrystallized as the crystalline form under the storage conditions; hence some crystalline peaks of urea were visible. Recrystallization of drugs and polymers during storage can occur [38]. Storage conditions such as temperature and humidity could trigger the molecular mobility of the drug, which might accelerate the recrystallization of the amorphous drug [39,40]. Moreover, the rearrangement of the amorphous state to the crystalline phase can be accompanied by the thermo-oxidation process in the solid state [41]. The results of the present study suggest that high temperature and high humidity accelerated the degradation of CP and increased drug recrystallization. 

3.7. Thermal Behavior Changes after Long-Term Storage 

The thermal behavior of CP-F before and after storage in the different conditions characterized by DSC is shown in Figure 6. It was found that the DSC thermogram of intact CP displayed a sharp endothermic peak at 92 ◦C. Two broad endothermic curves of the blank nanofibrous film were observed at 68 and 213 ◦C. The DSC thermogram of CP-F showed two broad peaks similar to the blank, however, the peaks were slightly shifted. The first endothermic broad peak of CP-F appeared at approximately 87 ◦C and another broad peak appeared at about 194 ◦C. This might be due to an interaction between CP and the excipients in the nanofibrous film. In addition, the absence of the melting CP peak in peak in the CP-F thermogram suggested that CP was dispersed into the nanofibrous film as an amorphous form.

After long-term storage at 25 ◦C/30% RH, the DSC thermogram of CP-F looked similar to that of the initial state of CP-F, with no melting peaks representing the crystalline characteristics of CP or polymers observed. The results imply that the amorphous CP remained stable in the nanofibrous film. However, the endothermic peak at 118 ◦C was observed in the CP-F stored at 25 ◦C/75% RH. It has been reported that the polymer type and storage condition has a strong impact on solid-state properties [42]. PVA and PVP are hydrophilic polymers and often hygroscopic: these polymers can absorb a high amount of moisture from the environment [43–45]. Hence, it is feasible that PVA and PVP present in CP-F would absorb water from the high humidity of 75% RH. In the meantime, the degradation of CP yields hydrogen peroxide and urea, which can further break down into water and ammonia [37]. Therefore, the endothermic peak of the CP-F thermogram that appeared at 118 ◦C might represent the water dehydration peak of the film after the sorption of the water from the degradation of CP, and the high humidity of the storage container during storage time.

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