Effect of process parameters on encapsulation efficiency. Successful microencapsulation process of essential oils results in a microcapsule with both minimum surface oil content and maximum retention of the core material inside the microcapsules. In this study, the effect of wall material ratio, ultrasonication time, and core material load on the encapsulation efficiency of SGEO through the complex coacervation process was investigated. Table 2 exhibits the different microcapsule formulations and the result of encapsulation efficiency values of SGEO. Encapsulation efficiency values of the SGEO microcapsules synthesized in this study were obtained in the range of 80.06–95.28%. The conditions that provided the highest encapsulation efficiency were determined as follows: the ratio of gelatin:gum Arabic of 6:1, the core material load of 5 g, and ultrasonication time of 20 min, at which conditions encapsulation of the SGEO was carried out effectually.
Table 2
Surface oil, total oil, and encapsulation efficiency of SGEO microcapsules.
Sample | Total Oil (%) | Surface Oil (%) | Encapsulation Efficiency (%) |
12G/GA-5SGEO-5t | 11.71±0.46 | 2.33±0.10 | 80.06±1.64 |
10G/GA-5SGEO-5t | 12.21±0.27 | 2.22±0.14 | 81.81±1.46 |
8G/GA-5SGEO-5t | 13.13±0.21 | 2.07±0.05 | 84.21±0.17 |
6G/GA-5SGEO − 5t | 13.97±0.08 | 1.48±0.15 | 89.43±1.01 |
6G/GA-5SGEO-10t | 14.13±0.17 | 1.29±0.03 | 90.89±0.33 |
6G/GA-5SGEO-15t | 16.20±0.52 | 1.16±0.06 | 92.83±0.61 |
6G/GA-5SGEO-20t | 19.60±0.54 | 0.92±0.06 | 95.28±0.43 |
6G/GA-3.5SGEO-20t | 14.97±0.47 | 0.77±0.04 | 94.85±0.33 |
6G/GA-1.75SGEO-20t | 11.15±0.18 | 0.58±0.04 | 94.76±0.39 |
Data represented as Mean ± SD, n = 3. |
As shown in Table 2, the decreasing of gelatin:gum Arabic ratio had a positive effect on the encapsulation of SGEO. In the coating process, gelatin and gum Arabic were acted as a positive polyelectrolyte and a negative polyelectrolyte, respectively. The ratio of gelatin:gum Arabic affected the charge balance, causing a change in the intensity of oppositely charged ion interactions during complexation [29]. High amounts of wall materials in complex solution may cause an excess of charge, which that the formation of the complexes affected negatively.
As the ultrasonication time also increased from 5 to 20 min, the encapsulation efficiency of the microcapsules increased from 89.43–95.28%, which indicates that increasing ultrasonication time between 5 and 20 min had a positive effect on the encapsulation of SGEO. It was also observed that the shorter ultrasonic treatment time than 5 min was not sufficient to prepare a homogeneous emulsion, and the higher ultrasonic time than 20 min increased the emulsion temperature because of the high cavitation effect. The emulsion temperature may cause a decrease in the embedding rate of essential oil in polymeric materials due to the essential oil volatilization [30].
When evaluating the effect of core material load on the encapsulation efficiency, it was observed that increasing SGEO load used in the synthesis process from 1.75 g to 5.00 g positively affected the encapsulation efficiency. However, the higher oil load than 5.00 g had a negative effect on the emulsion, causing the oil droplets to not show a homogeneous mixture during ultrasonic treatment. The highest encapsulation efficiency value was recorded as 95.28% for a core material load of 5.00 g.
The optimum sample was considered as 6G/GA-5SGEO-20t microcapsules with the highest encapsulation efficiency, and the 6G/GA-5SGEO-20t microcapsules were used in the thermal, chemical, and morphological analyses in the following sections.
FTIR. The chemical structure of gum Arabic, gelatin, SGEO, and SGEO microcapsules was investigated by FTIR spectroscopy analysis and the FTIR spectra are shown in Fig. 1.
Amide groups of gelatin peptide bonds exhibit characteristic absorption spectral peaks in specific bands, which are amide I (1600–1800 cm–1), II (1470–1570 cm–1), III (1250–1350 cm–1), and A (3300–3500 cm–1) bands [31]. As shown in Fig. 1, the characteristic absorption bands of the gelatin spectrum were observed at 3289 cm-1, 1624 cm-1, 1540 cm-1, and 1238 cm-1, correspond to amide A (the vibrations of OH and NH groups), amide I (the elongation vibrations of C = O and CN groups), amide II (the vibrations of NH and CN groups) and amide III (elongation of the vibrations of NH and CN groups), respectively [32]. The FTIR spectrum of gum Arabic showed a band at 3347 cm− 1 due to the presence of hydroxyl groups. A peak that was observed with low intensity at about 2932 cm− 1 corresponds to free carboxylic groups [33]. The carboxylic groups of gum Arabic are responsible for negative charges to promote the process of coacervation [34]. In the spectrum of SGEO microcapsules, the hydroxyl and carboxyl groups shifted from 3347 cm-1 to 3300 cm-1 and from 2932 cm-1 to 2943 cm-1, respectively. These changes indicate that there were hydrogen bonding interactions in the formation of the microcapsules [32]. The other characteristic absorption bands of gum Arabic spectrum were observed at 1598 cm-1, 1364 cm-1, and 1004 cm-1, related to C = O stretching/N-H bending, C-N stretching, and C-O stretching, respectively [35].
In the FTIR spectrum of SGEO, significant vibrational bands at 3420, 2943, 1706, 1161, 965, 767, and 683 cm− 1 were observed. The weak spectral band at 3420 cm− 1 corresponds to the stretching vibrations of the OH functional group of alcohols [36]. The band with a peak point at around 2943 cm-1 could be attributed to C = C–C ring vibrations of volatile compounds and it also refers to C-H vibrations of the aromatic methoxyl, methyl and methylene groups of the side chains [37, 38]. The strong and sharp absorption peaks at 1706 cm-1 is assigned to the C = O stretching vibration peak of ketones. The peaks observed at between 1600 and 1450 cm-1 were assigned to the vibrations of C = C bonds in the aromatic compounds, while the peak at between 1000 and 650 cm-1 were related to the deformation vibrations of C–H bonds in the benzene ring [39], the stretching vibration of C–O bond in the primary and secondary alcoholic groups [37]. The characteristic bands of SGEO were also identified in the FTIR spectra of the SGEO microcapsules. This indicates that the SGEO was successfully encapsulated into the gelatin/gum Arabic structure. The peak at around 2850 cm− 1 which was related to the formation of ionic interaction between the negative carboxyl group of gum Arabic and the protonated amine group of gelatin [40] were detected in FTIR spectrums of the SGEO microcapsules, while they were not visible in the gelatin and gum Arabic spectrums. The successful encapsulation of SGEO with gelatin and gum Arabic polymer shells was supported by the FTIR spectrum of sweetgum.
Thermal properties. The thermal stability of gelatin, gum Arabic, SGEO, and SGEO microcapsules was investigated by TGA, and the differential thermogravimetric (DTG) and TGA curves of the samples are depicted in Fig. 2.
As shown in Fig. 2, SGEO presents weight loss in two stages, while gelatin, gum Arabic, and SGEO microcapsules present weight loss in three stages. In the TGA thermogram of gelatin and gum Arabic, it was observed that these two wall materials present similar decomposition behavior. Firstly, a small weight loss was detected up to a temperature of about 100°C related to the departure of adsorbed and bound water. The main stage of thermal degradation was observed in the temperature range of 230 to 450°C with a weight loss of about 60.0% for gelatin because of the thermal degradation of the peptide bonds of the gelatin molecules [41] and 59.9% for gum Arabic attributed to the degradation of polysaccharides [42].
The thermogram of SGEO presents fast thermal degradation sub-divided into two steps. While the first stage observed in the temperature range of 100 to 315°C was associated with the evaporation of the simplest volatile compounds due to their heat-sensitive structure [43], the other stage observed in the temperature range of 315 to 415.5°C corresponds to the degradation of the complex aromatic rings present in the major compounds of SGEO [44]. SGEO also completely decomposes at 415.5°C, which indicates the thermal instability of its components.
In the thermograms of the SGEO microcapsules, it was observed that the SGEO microcapsules present three different stages. The first stage was observed at temperatures up to 230°C due to the loss of adsorbed and bound water present in the samples, the volatile compounds of free SGEO on the surface of microcapsules and SGEO diffused into the pores of the microcapsules, which recorded weight losses of 19.01%. The second stage was from around 230 to 450°C with average weight loss of 64.18%. The significant mass loss is associated with the depolymerization and degradation of gelatin and gum Arabic and as a result, encapsulated SGEO was released and evaporated. The third stage was observed between 450 and 600°C because of the carbonization of all the structures of microcapsules and which recorded weight loss of 7.06%. At 600°C, the residual masses are 9.75%. These results demonstrated that the thermal resistance of the SGEO microcapsules was significantly higher than un-encapsulated SGEO and the interaction between gelatin and gum Arabic in complex coacervates was enhanced thermal stability of SGEO as a heat-sensitive compound.
XRD. In order to investigate the crystallinity of wall materials, SGEO, and the SGEO microcapsules, the X-ray diffraction spectra were recorded. From the XRD patterns (Fig. 3), gelatin and gum Arabic show a prominent broad peak at around 15–25°, indicating the amorphous nature of this biopolymer with very low crystallization [45]. The width of the XRD peak is closely related to the crystallite of the samples, and the broadened peak is usually due to the imperfect crystal structure of the samples [46]. Gelatin and gum Arabic present the characteristic broad peak at 2θ = 20.7° and 2θ = 18.7°, respectively. The microcapsules have an amorphous dispersion peak with greater intensity at about 2θ = 19.7°, which signifies the characteristic of SGEO microcapsules. The XRD spectra of SGEO microcapsules suggested that there are some changes in the original crystallization behavior of wall materials and shifting of the diffraction peak, which could be a result of emerging amide bonds between gelatin and gum Arabic [47] and hydrogen bond forces between the core material and the wall materials [30]. The XRD pattern of unencapsulated SGEO and microcapsules revealed higher crystallinity than gum Arabic and gelatin. The peak intensity of the microcapsules was higher compared to the wall materials. This phenomenon may be due to the increment in the crystallinity of the microcapsules and the homogeneous distribution of the essential oil in the biopolymer matrix [48]. SGEO exhibited a broad peak at 2θ ≈ 18.7° which could be due to organic materials adsorbed on SGEO [47]. Furthermore, the semicrystalline pattern of SGEO disappeared in the microcapsules, which confirmed the successful loading of SGEO into the microcapsules.
Morphological analysis. Figure 4 shows the SEM images of the SGEO microcapsules. As shown in Fig. 4, the SGEO microcapsules have a spherical shape with an irregular surface. The size of most microcapsules varies from about 1 µm to 20 µm. However, the microcapsules with solid walls have some cracks and small pits on their surface, which can be attributed to the drying process [40]. Elimination of water droplets on the surface of the microcapsules by oven drying may create openness in microcapsules.
Release studies. The in vitro release behavior of SGEO from the microcapsules was examined and shown in Fig. 5. The release of SGEO from the microcapsules can be divided into two phases. During the first 3 hours, there was an initial rapid release that corresponded to 72.52%, followed by a slower release up to 12 hours of 91.49% of the encapsulated SGEO amount from the microcapsules. The initial release of SGEO from the microcapsules may be relatively resulted from the part of the uncapsulated SGEO on the surface of the microcapsules, rather than the part of SGEO capsulated in the microcapsule interior.
To evaluate the mechanism of SGEO release from the microcapsules, the release data were fitted by mathematical models describing zero-order, first-order [49], Korsmeyer-Peppas [50], Higuchi [51], and Baker-Lonsdale [52] kinetic models, and the kinetic model equations are given in Table 3. In the kinetic model equations, Q and Q0 are the cumulative amount of SGEO released at time t and the initial amount of SGEO (t = 0). k0 and k1 are the zero-order and first-order release constants. Mt/M∞ is the percentage of SGEO released at time t relative to the percentage released at infinite time. n is a diffusion exponent indicating the type of release mechanism as Fickian diffusion (case I transport) (n ≤ 0.43), non-Fickian or anomalous transport (0.43 ≤ n < 0.85), or case II transport (n ≥ 0.85). kH is the Higuchi constant of dissolution and kK is the Korsmeyer-Peppas model constant, which reveals structural and geometric character of the drug release matrix.
Table 3
Parameters of the kinetic models for the analysis of the release of the sweetgum oil from the microcapsules.
Kinetic Models | Equations | | |
Zero-order | \(Q={Q}_{0}-{k}_{0}t\) | R2 | 0.8633 |
\({Q}_{0}\) | 49.0170 |
\({k}_{0}\) | 6.8256 |
First-order | \(\text{ln}\left(100-Q\right)=\text{ln}{Q}_{0}-{k}_{1}t\) | R2 | 0.9891 |
\({Q}_{0}\) | 57.3172 |
\({k}_{1}\) | 0.2939 |
Korsmeyer-Peppas Model | \(\frac{{M}_{t}}{{M}_{{\infty }}} ={k}_{K} {t}^{n}\) | R2 | 0.9938 |
kK | 52.9792 |
n | 0.2275 |
Higuchi Model | \(\frac{{M}_{t}}{{M}_{{\infty }}} ={k}_{H} {t}^{0.5}\) | R2 | 0.9507 |
kH | 18.999 |
Baker-Lonsdale Model | \(f=\frac{3}{2} \left[1-{\left(1-\frac{{M}_{t}}{{M}_{{\infty }}}\right)}^{\frac{2}{3}}\right]-\frac{{M}_{t}}{{M}_{{\infty }}}=K.t\) | R2 | 0.9342 |
K | 0.0233 |
SGEO release kinetic parameters of the microcapsules obtained by zero-order, first-order, Korsmeyer-Peppas, Higuchi, and Baker-Lonsdale kinetic models are given in Table 3. The models with higher correlation coefficients (R2) have been considered as a more appropriate model for the evaluation of the release data. Based on R2 as shown in Table 3, first-order (R2 = 0.9891), Higuchi (R2 = 0.9507), and Korsmeyer-Peppas (R2 = 0.9938) models can be chosen as an appropriate for the explaining release kinetic of SGEO. However, Korsmeyer-Peppas is the best appropriate model with the highest R2 value (R2 > 0.99) for the SGEO release from the SGEO microcapsules. The diffusion exponent in the Korsmeyer–Peppas model was found to be less than 0.43. The results suggest that the Fickian behaviour of the diffusion-controlled release process is due to the low degree of swelling and the presence of oil droplets on the surface of the microcapsules [53].
Antibacterial properties. The SGEO microcapsules were investigated for their antibacterial activity against common Gram-positive and Gram-negative pathogenic microorganisms including Staphylococcus aureus and Escherichia coli according to the standard ASTM E2149-13a [28]. The antibacterial activities were evaluated after a specific contact time and calculated by reduction in the percent of E. coli and S. aureus bacteria. As shown in Table 4, it can be noted that SGEO microcapsules are effective antibacterial materials against the two common pathogenic bacteria.
Table 4
Reduction of Escherichia coli and Staphylococcus aureus exposed to the samples after 24 h.
Sample | S. aureus Reduction % | E. coli Reduction % |
SGEO microcapsules | 99.98 | 99.99 |