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The Viscosity Behaviour of PEGylated Locust Bean Gum/Rosin Ester Polymeric Nanoparticles

In this study, PEGylated locust bean gum – rosin glycerol ester polymeric nanoparticles (PEG-LBG/RE PNPs) were synthesized by using simple ultrasonic irradiation method. The nanoparticles were characterized by using Fourier-transform infrared spectroscopy (FTIR) and scanning transmission electron microscopy (STEM). The viscosity behaviors of nanoparticles were studied in different conditions (pH, sonication time, and salt). The experimental results were calculated by Huggins, Kraemer, Tanglertpaibul-Rao, and Higiro models to understand the colloidal stability, the miscibility mechanism, and coefficients of nanoparticles. The results confirmed that the homogenous distribution of nanostructure was related to sonication time (30 min) and the presence of NaOH salt. With the addition of NaOH, the nanosystem based on ionotropic gelation technique was made more homogeneous. The results made us think that nanoparticles can be a good candidate for drug delivery systems in biomedical and pharmaceutical applications.

The ultrasonic irradiation synthesis of different morphologies of nanomaterials consisted of metal/metal oxides, and polymeric materials have received considerable attention in the nanotechnology applications. The ultrasonic irradiation (20 kHz to 10 MHz) method has been employed in the preparation of the high purity, the uniform shape, and the nanosized distribution of nanomaterials. This method causes the formation of the acoustic cavitations which consist of the bubbles [10]. The growth and collapse of bubbles are related to the transfer of energy at high pressures and temperatures due to the highly reactive free radicals such as hydrogen radicals (H•) and hydroxyl radicals (OH•). Bubbles generate three zones, such as a hot spot (5000°C, 500 atm), a gas-liquid interface (300°C, 50 atm), and a bulk solution (25°C, 1 atm) [11].
When the studies in the current literature are examined, it has been found that there are very few studies on LGB based on nanoparticles [28][29][30]. It was found that no studies were performed on the locust nanostructures containing rosin gum and derivatives. In this work, the ultrasonic irradiation method was used for the preparation of novel PEGylated locust bean gum (PEG-LBG)/rosin glycerol ester (RE) polymeric nanoparticles (PNPs) at room temperature. The present research work was aimed at the colloidal stability, the viscosity behaviour, and miscibility of binary polymer blends of PEG and LBG PNPs due to the intrinsic viscosity. The intrinsic viscosity of the polymer is a significant molecular characteristic, depending on the size of the polymer chain, molecular weight, and radius of rotation of the polymer in dilute solution. The voluminosity (VE), shape factor (υ), the intrinsic viscosity [η], and Krigbaum and Wall miscibility parameter (Δb) of polymeric nanoparticles were calculated from different models such as Huggins, Kraemer, Tanglertpaibul-Rao, and Higiro [17]. The values of intrinsic viscosities were used to determine the rheological behaviour of the PEG-LBG/RE PNPs at different conditions (pH, sonication time, and salt). The homogeneous distributions of PEG with LGB had an influence on the blends ratio of PEG/LBG (1:1, 1:2), sonication time (10-70 min.), temperature (25-35°C), and salts (NaOH, KOH, CTAB). With the addition of NaOH salt, PEG-LBG/RE PNPs based on ionotropic gelation technique were made into a more homogeneous solution. The PEG-LBG/ RE PNPs were characterized to examine surface morphologies using a Fouriertransform infrared spectroscopy (FTIR) and scanning transmission electron microscopy (STEM). The aim of this study was to provide an investigation of rosin ester-based nanoparticle distributions in LGB and understand the role of polymerparticle interactions with respect to nanoparticle concentration as well to use the candidate nanocarrier for biomedical applications.

Preparation of PEG-LBG/RE polymeric nanoparticles
PEG-LBG/RE PNPs were synthesized using the ultrasonic irradiation method (Ultrasonics Vibra-Cell, probe type, amplitude %30, a frequency of 20 kHz) with different ratios of blends (PEG-LBG: 1:1, 1:2). In the procedure, two phases were prepared such as the dispersion phase and the continuous phase.
The continuous phase: 125 mg LBG was dissolved in 50 ml of distilled water (60°C for 20 min) and then added to 125 mg of PEG400 polymer solution at room temperature.
The dispersion phase: 0.01 g RE was dissolved in 0.5 ml DMSO and then 7 ml ethyl acetate was added to the solution.
7.5 ml of continuous phase and the dispersed phase was sonicated at room temperature. 42.5 ml of continuous phase was then added slowly to the blends, and the sonication procedure was continued for 30 minutes. The final solution was evaporated at room temperature for 14 hours until ethyl acetate completely evaporated in the solution. Polymer blends were also performed for different composition ratios (PEG-LBG) such as 1:1 and 1:2.

Characterization parts
The dynamics viscosities of LBG, PEG-LBG, and PEG-LBG/RE were determined by a programmable AND viscometer (SV-10, Sine-wave Vibro Viscometer, A δ D Company). PEG-LBG/RE PNPs were scanned in the dark field area with the wet STEM detector by using FEI QUANTA S50 (A copper grid, Ted Pella, support films, carbon type A, 300 meshes was utilized). STEM holder was cooled to 2°C and the pressure was set between 700 and 1300 Pa. Samples were ground with KBr powder and analyzed from 4000 to 600 cm À1 with a resolution of 4 cm À1 using 8 scans by using a PerkinElmer FTIR emission spectrometer.

Huggins
Kraemer Tanglertpaibul   Kraemer, Tanglertpaibul-Rao, and Higiro models were given comparatively in Table 2 ( Figures 1-4). In this study, we focused on the effect of nanoparticles on the morphology of immiscible polymer blends. We found that PEG-LBG/RE PNPs (1:2) were immiscible due to the mixing ratio of PEG-LBG ( Table 3). The Huggins, Kraemer, Tanglertpaibul-Rao, and Higiro plots of the intrinsic viscosities were calculated at different blend ratios, and the results showed the critical role on relation between the intrinsic viscosities and the blend ratios.  The Tanglertpaibul-Rao model and Huggins model (R 2 = 0.96-1.00) were the best models to understand the intrinsic viscosity of PEG, LBG, PEG-LBG, and PEG-LBG/RE PNPs. Behrouzian et al. [32] reported that the Tanglertpaibul and Rao model was the best model for the intrinsic viscosity determination of cress seed gum  solutions. Razavi et al. [37] reported that the best model was Tanglertpaibul and Rao model for wild sage seed gum. In this study, the intrinsic viscosity of PEG-LBG/RE PNPs in the presence of different salts (NaOH, KOH, and CTAB) was investigated (Csalt, 0.1 M; Vsalt, 2 mL; Vsolution; 50 mL) at 25°C. The effect of NaOH, KOH, and CTAB salts on the values of intrinsic viscosity of PEG-LBG/RE PNPs (1:1) was presented in Figure 5.  The pH values of the solutions (LBG, PEG-LBG, and PEG-LBG/RE PNPs) at initial pH in KOH, NaOH, and CTAB salt additions were determined: pH initial , 5.7; pH initial , 5.55; and pH initial , 5.32, respectively. In the presence of salt, the values of the intrinsic viscosity for the mixture were observed to change in two different salts such as KOH (pH final: 5.93) and NaOH (pH final : 5.72). The [η] values for PEG-LBG/ RE PNPs (1:1) did not exhibit distinctive changes in the presence of CTAB (pH final : 3.87). Jiang et al. [38] reported that the interactions between blends were dependent on the ionic strength at low salt concentration which was related to the increase of salt concentration. Consequently the addition of NaOH and KOH showed the electrostatic repulsion between charges along the backbone of the polymer blends.

Temperature and sonication time factor
The intrinsic viscosity decreased when the temperature increased, and the relation of the experimental results of PEG-LBG/RE PNPs with the temperature was shown in Figure 6.
However, when PEG-LBG/RE PNPs were sonicated, the intrinsic viscosity decreased for 30 minutes but remained constant after a period of time. These results had proven that the sonication time changed the value of viscosity and was effective on the blends (30% amp., 25°C) ( Figure 7). The viscosity of Cu-ethylene glycol (EG) nanofluids was proven to decrease with the sonication time [39]. In this study, we found a similar situation, and demonstrated that sonication time changes the viscosity, which has a role on the formation of nanoparticles.

The voluminosity, shape factor, and miscibility parameter
In this study, we investigated the relationship between the intrinsic viscosity and the surface morphology, particle size, and shape. The shape factor was calculated using the approach given as follows: (a) n < 2.5 indicates spherical shape, and (b) n > 2.5 indicates ellipsoidal particles [40]. In this study, we predicted the size and shape factor of PEG-LBG (1:1), PEG-LBG/RE PNPs (1:1), and PEG-LBG/RE PNPs (1:2) using the values of the intrinsic viscosity, associated with the shape factor, which were used to determine the change in the structure configuration. We calculated the shape and the Krigbaum and Wall (Δb) parameters of PEG-LBG/RE PNPs (1:1) using the intrinsic viscosity to determine the changes in the blends. We found that PEG-LBG/RE PNPs (1:1) had a spherical-like configuration, and the amounts of PEG had a role on the miscibility due to the interactions between the functional groups in the blends.

FTIR analysis
The FTIR spectra of pure LBG, pure RE, and PEG-LBG/RE PNPs were shown in Figure 8.
The FTIR spectrum of pure LBG showed a broad absorption peak at 3250 cm À1 (stretching of -OH group), 2952 cm À1 (stretching of -CH), 1748 cm À1 (stretching of C=O), and 1000-1100 cm À1 (stretching of C-O-H). Upadhyay et al. [41] and Chakravorty et al. [42] found the FTIR spectrum data similar. The FTIR spectrum of pure RE showed a peak at 3330 cm À1 (stretching of -OH group), 1730 cm À1 (stretching of C=O), and 1120 cm À1 (stretching of C-O-H). As we have seen from the FTIR results, we have demonstrated that the apparent OH peak of LBG disappeared and that the rosin glycerol ester is coated with surrounding PEGylated LBG.

STEM analysis
According to the STEM image of PEG-LBG/RE PNPs (160.000x and 300.000x), we can see that the interior structure of the polymeric nanoparticle is LBG with the size lower than 50 nm. We are able to tell that these particles are small agglomerates of it (Figure 9).

Conclusions
We prepared the novel PEG-LBG/RE PNPs with an average particle size of 100 nm using the ultrasonic irradiation. We dispersed the amphiphilic RE coated with PEG-LBG blends in nanosize and spherical structure. We focused on the miscibility of the blends, and shapes of the polymeric nanoparticles were calculated using the values of the intrinsic viscosity in different conditions. We estimate that PEG-LBG/RE PNPs can be used to increase the therapeutic efficacy and biocompatibility of the nanodrug in pharmaceutical and biomedical studies.