Nanoencapsulation of styrax liquidus (Liquidambar orientalis) with different polymers

Objective: The aim of this thesis was to synthesize and characterize zein and poly-ε-caprolactone (PCL) nanoparticles encapsulated with styrax liquidus, examine their bioavailability, assess their anti-ulcerogenic effect, and evaluate their sensory consumability. Material and Methods: After the preprocessing of raw styrax liquidus balsam to make it suitable for nanoencapsulation studies, the thesis work was conducted in five stages: (I) determination of suitable production conditions for styrax liquidus-loaded zein and PCL nanoparticles using simple and flash nano-precipitation methods, (II) optimization of process conditions for zein nanoparticles, (III) morphological, physical, and chemical characterizations, as well as analyses in static in vitro digestion environments, for zein and PCL nanoparticles produced at the optimum point, (IV) determination of the anti-ulcer effects of zein and PCL nanoparticles produced at the optimum point, and (V) sensory evaluation of the optimum zein nanoparticles. Styrax liquidus-loaded zein and PCL nanoparticles were synthesized using simple nano-precipitation and flash nano-precipitation methods. The particle size, transmittance, polydispersity index (PDI), zeta potential values, encapsulation efficiency, and release rates in the static in vitro digestion environments (oral and gastric) of zein nanoparticles were analyzed and the optimum production conditions were determined. Different ratios of styrax liquidus were studied in the production of PCL nanoparticles and optimum productions were identified at appropriate styrax liquidus ratio for each methods. Physical (color, moisture, water activity, water solubility, bulk-tapped density, flowability), chemical (total phenolic content, antioxidant capacity, trans-cinnamic acid content), in vitro digestion release of styrax liquidus (oral, gastric, and intestinal environments), and biological activity (anti-ulcerative effect) analyses were performed for optimum zein and PCL nanoparticles. The anti-cancer effect of raw styrax liquidus was investigated on human prostate cancer cells (PC3 and LNCaP). Styrax liquidus -loaded zein nanoparticles produced under optimum conditions were also evaluated for sensory characteristics. Results: The particle sizes of styrax liquidus-loaded zein nanoparticles obtained using the simple nano-precipitation (SN) and flash nano-precipitation (FN) methods were measured as 140.31 to 427.40 nm and 151.00 to 233.95 nm, respectively, showing a narrow size distribution (PDI: 0.1-0.2%). The highest encapsulation efficiency and product yield in the production of zein nanoparticles were achieved with the FN method, with values of 95.81% and 98.90%, respectively. The release percentages of styrax liquidus in the static in vitro oral environment for zein nanoparticles produced by the SN method (average 7.41% and lowest 1.22%) were lower than those produced by the FN method (average 11% and lowest 2.65%). The release percentages in the static in vitro gastric environment for zein nanoparticles produced by the FN method (average 69% and highest 90.58%) were higher than those produced by the SN method (average 50% and highest 94.77%). It was observed that colloidal zein nanoparticles maintained their stability, and those produced by the FN method were more stable compared to those produced by the SN method. Sensory analysis indicated that the optimum powders produced by the FN method were found to be more consumable in terms of color, agglomeration, and styrax liquidus flavor intensity. The FN method was deemed more suitable for the production of stable styrax liquidus-loaded zein nanoparticles. Styrax liquidus-loaded PCL nanoparticles were produced using the simple nano-precipitation (SN) and flash nano-precipitation (FN) methods under different production conditions, resulting in maximum styrax liquidus-loaded nanoparticle powders at rates of 2% and 3%, respectively. At high styrax liquidus ratios, both methods experienced a dramatic decrease in encapsulation efficiency and product yield. The sizes of PCL nanoparticles produced by the SN method varied between 258.67 nm and 510.00 nm, while those produced by the FN method ranged from 225.88 nm to 389.98 nm. In the static in vitro digestion analysis of PCL nanoparticles produced by the SN method, styrax liquidus releases ranged from a minimum of 1.59% in the oral environment to a maximum of 69.03% in the gastric environment. In contrast, the releases of nanoparticles produced by the FN method were higher, ranging from a minimum of 2.88% in the oral environment to a maximum of 75.00% in the gastric environment. The FN method was found to be more advantageous for the production of styrax liquidus-loaded PCL nanoparticles. The in vitro gastric release profiles over time were investigated for optimum zein and PCL nanoparticles to select the suitable material and method for styrax liquidus nanoencapsulation. The lowest burst release of 20% in the first 15 min and a long-term sustained release profile was obtained from PCL carrier material by FN method. styrax liquidus-loaded zein and PCL nanoparticles produced by the FN method showed similar physical properties, while their chemical characteristics and in vitro digestion release profiles were influenced by the method and material type, resulting in variations. The anti-ulcerative effects of styrax liquidus and the obtained optimum nanoparticles were examined, and SN-PCL nanoparticles exhibited the highest urease enzyme inhibition. The anti-carcinogenic effect of raw styrax liquidus was studied on PC3 and LNCaP human prostate cancer cell lines at different doses. The IC50 values were determined to be 198.79 μg/mL and 59.89 μg/mL after 48 hours of treatment, respectively. Conclusion: Within the scope of the thesis study, the nanoencapsulation of styrax liquidus, possessing high bioactive properties, was successfully achieved using zein and PCL materials through the BN and FN methods. Loading styrax liquidus into zein and PCL carrier nanoparticles and obtaining stable nanoparticles were accomplished in a single step and rapidly using low-cost simple and flash nano-precipitation methods. The in vitro digestion analysis of the powdered nanoparticles loaded with styrax liquidus obtained at the optimal point revealed low release properties in the oral environment and high release properties in the stomach and intestinal environments. This, coupled with the anticipation that their small sizes would facilitate easy cellular uptake, is expected to enhance the bioavailability of styrax liquidus. Consequently, it is predicted that this will enhance the health effects of styrax liquidus, which has anti-ulcer and anti-cancer effects. The conversion of styrax liquidus, which is challenging for oral consumption, into powdered zein nanoparticles was concluded to make it sensorially acceptable for direct consumption. Keywords: Simple/flash nanoprecipitation, Styrax liquidus, Nanoparticles, Antiulcerogenic-anticarcinogenic effect.