Determination of the anticancer activity of standardized extract of Centella asiatica (ECa 233) on cell growth and metastatic behavior in oral cancer cells.

Background and purpose: The anticancer drugs used for oral cancer treatment present many disadvantages, such as low solubility, low permeability, and poor bioavailability. However, the anticancer activity of ECa 233 has not been widely studied. Therefore, the anticancer activity of ECa 233 was investigated in this study. Experimental approach: MTT assay was carried out to determine cell viability. Characterizations of cell apoptosis were monitored using DAPI and FDA staining and Hoechst 33258 and AO staining. Confirmation of the apoptosis-induced KON cells was done using annexin V-FITC staining, and ROS generation was determined by DCFDA staining. Cell death and the cell cycle arrest activity of ECa 233 were demonstrated by a flow cytometer. The anti-migration and anti-invasion properties of ECa 233 were examined. The anti-proliferative of ECa 233 was investigated. Cellular uptake of ECa 233 was measured by TEER values. The pharmacokinetics of ECa 233 were estimated using the pkCSM web server. Findings/Results: ECa 233 decreased the KON cell viability. Morphological analysis showed the KON cells' loss of cell stability and structure, disorganized nucleus and cytoplasm, and induced cell death. ECa 233 acted as a cell cycle arrest in the G0/G1 phase and reduced the migration and invasion ability in KON cells. TEER values significantly increased in KON cells, which decreased cell colony and multicellular spheroid formations. The pharmacokinetic profiles of the main components are of interest for future usage. Conclusion and implication: ECa 233 can be used as an alternative therapy as well as a medicinal plant selected for sensitizing oral cancer cells to chemotherapy.

Unveiling the Molecular Dynamics, Anticancer Activity, and Stability of Spearmint Oil Nanoemulsions with Triglycerides.

Although spearmint oil (SMO) has various pharmacological properties, especially for cancer treatment, its low water solubility results in poor bioavailability. This limits its application as a medicine. One possible solution is to the use of SMO in the form of nanoemulsion, which has already been shown to have anticancer effects. However, the mechanism of SMO nanoemulsion formation remains unclear. The objective of this study was to use molecular dynamics (MD) for clarifying the formation of SMO nanoemulsion with triglycerides (trilaurin, tripalmitin, and triolein) and Cremophor RH40 (PCO40). Nanoemulsions with different SMO:triglyceride ratios and triglyceride types were prepared and analyzed for anticancer activity, droplet size, droplet morphology, and stability. Despite switching the type of carrier oil, SMO nanoemulsions retained strong anticancer effects. A ratio of 80SMO:20triglycerides produced the smallest droplets (<100 nm) and exhibited excellent physical stability after a temperature cycling test. MD simulations showed that polyoxyethylenes of PCO40 are located at the water interface, stabilizing the emulsion structure in an egglike layer. Droplet size correlated with triglyceride concentration, which was consistent with the experimental findings. Decreasing triglyceride content, except for the 90SMO:10triglyceride ratio, led to a decrease in droplet sizes. Hydrogen bond analysis identified interactions between triglyceride-PCO40 and carvone-PCO40. Geometry analysis showed PCO40 had an "L-like" shape, which maximizes the hydrophilic interfaces. These findings highlight the value of MD simulations in understanding the formation mechanism of SMO and triglyceride nanoemulsions. In addition, it might also be beneficial to use MD simulations before the experiment to select the potential composition for nanoemulsions, especially essential oil nanoemulsions.

Nanoemulsion-Based Orodispersible Film Formulation of Guava Leaf Oil for Inhibition of Oral Cancer Cells.

Among natural sources, guava leaf oil (GLO) has emerged as a potential anticancer agent. However, its limited water solubility poses a significant challenge for its use. Oil-in-water nanoemulsions are used to address the limitation of water solubility of GLO prior to its incorporation into orodipersible films. Nanoemulsions containing GLO:virgin coconut oil (VCO) at a ratio of 50:50 to 70:30 presented a small droplet size of approximately 50 nm and a relatively low zeta potential. GLO:VCO at a ratio of 70:30 was selected for incorporation into sodium alginate film at various concentrations ranging from 1% to 30% w/w. Tensile strength and elongation at break relied on the concentration of nanoemulsions as well as the internal structure of films. Fourier transform infrared spectroscopy revealed that GLO was compatible with sodium alginate. Film containing 2% w/w of nanoemulsions (2G_ODF) exhibited effective in vitro antioral cancer activity, with an IC50 of 62.49 ± 6.22 mg/mL; furthermore, its anticancer activity showed no significant difference after storage at 25 °C for 1 year. Moreover, 2G_ODF at IC60 arrested colony formation and cell invasion. There is also evidence that cell death occurred via apoptosis, as indicated by nuclear fragmentation and positive Annexin-V staining. These findings highlight the potential of orodispersible films containing GLO nanoemulsions as a prospective oral anticancer agent.

Sustained release gel (polymer-free) of itraconazole-loaded microemulsion for oral candidiasis treatment: time-kill kinetics and cellular uptake.

Itraconazole (ICZ) was prepared in a self-microemulsifying (SM) gel. This gel was intended for use in the oral mucosa, where low volume and flow of saliva result in limited solubility and absorption of drugs that are poorly water-soluble. The drug-loaded gel formulation (ICZ-SM) was selected as a clear solution in the ternary phase diagram to improve the solubility of ICZ. Seven ratios (S1-S7) were prepared by mixing polyoxyl 35 castor oils (P35), a medium chain with a blend of mono-, di-, and triglycerides (MCT), and water. Phase separation of large-sized emulsions by countering with artificial saliva were observed in dilution tests for the formulation contained MCT, P35, and water at the ratios of 70:20:10 (S1), 10:80:10 (S3), and 20:60:20 (S4). Formulations in the ratios of 15:50:35 (S5) and 19:43:38 (S6) produced strong ICZ-SM gels, as shown by rheology tests, whereas the formulations at the ratios of 30:60:10 (S2) and 10:43:47 (S7) exhibited no elasticity. A model of zero-order kinetic (S5) and first-order kinetic (S6) were found to best fit the release kinetics of ICZ from the gels. Time-killing assays revealed that S5 and S6 required less time compared with S2 and the ICZ solution. Furthermore, S5 exhibited the highest increase in cell uptake compared with S2, S6, and the ICZ solution. These findings suggest that the ICZ-SM gel was a free polymer capable of delivering an ICZ for the treatment of oral candidiasis, and that ICZ-SM gels applied locally exhibit enhanced absorption into cells.

New Approach for Preparing Solid Lipid Nanoparticles with Volatile Oil-Loaded Quercetin Using the Phase-Inversion Temperature Method.

Quercetin (QCT), a natural flavonoid, is of research interest owing to its pharmacological properties. However, its pharmacokinetic limitations could hinder its widespread therapeutic use. Nanocarriers, especially solid lipid nanoparticles (SLNs), might overcome this constraint. This study aimed to investigate QCT-loaded SLNs prepared via a new approach using a volatile oil. The phase-inversion temperature method was used to incorporate rosemary oil (RMO) into SLNs prepared using solid lipids possessing different chemical structures. Among the solid lipids used in the formulations, trilaurin (TLR) exhibited the smallest particle size and good stability after a temperature cycling test. SLNs prepared with a ratio of RMO to TLR of 1:3 could load QCT with an entrapment efficiency of >60% and drug loading of ~2% w/w. The smallest particle size was achieved using the polyoxyethylene-hydrogenated castor oil RH40, and the particle size depended on the concentration. The drug-release profile of QCT_TLR exhibited prolonged biphasic release for >24 h. QCT_TLR was a safe formulation, as indicated by a cell viability percentage of >75% at <2% v/v. In a computer simulation, the system with RMO enabled smaller sized SLNs than those without RMO. This new discovery shows great promise for producing SLNs via the phase-inversion temperature method with incorporation of volatile oil, particularly for delivering compounds with limited water solubility.

Preparation and Characterization of Nifedipine-Loaded Dry Medium Internal-Phase Emulsions (Dry MIPEs) to Improve Cellular Permeability.

A nifedipine (NP) dry emulsion was fabricated by the adsorption of medium internal-phase emulsions (MIPEs). Simple homogenizers were first used to mix conventional liquid MIPEs, and then a microfluidizer was used to reduce the resulting emulsions' droplet sizes. The dry MIPEs (solid) were produced by adsorbing the emulsions onto solid carriers with a high surface area. The dry MIPEs were diluted in a simulated gastric fluid under gentle agitation to form emulsions. The diluted dry MIPEs were divided into three groups based on an NP content of 0.3%, 0.5%, and 0.7%, with sizes of 5026-5404 nm, 2583-3233 nm, and 1318-1618 nm in diameter, respectively. Powder X-ray diffraction (PXRD) measurements and differential scanning calorimetry (DSC) were used to characterize the physical properties of the dry MIPEs. The samples contained 0.5% or 0.7% drug, 2-4% surfactant, and 8-16% oil (5RH2/8, 7RH2/8, and 7RH4/16) and showed the characteristic peak for NP. No NP peak was observed in formulations with 0.3% NP and any oil-phase content (3RH2/8, 3RH4/16, and 3RH8/32). The formulations with 0.5% drug, 4-8% surfactant, 16-32% oil (5RH4/16 and 5RH8/32) and those with 0.7% drug, 8% surfactant, and 32% oil (7RH8/32) also did not show the peak for NP. These findings demonstrated that microfluidization improved the solubility of NP in the formulations. The subsequent drug dissolution results were consistent with the DSC thermogram and PXRD pattern results. 3RH2/8, 3RH4/16, 3RH8/32, 5RH4/16, 5RH8/32, and 7RH8/32 were completely dissolved and showed higher dissolved NP amounts than 5RH2/8, 7RH2/8, 7RH4/16, and NP powder. The lowest mean dissolution time was for 7RH8/32 (13.31 ± 0.87 min). Caco-2 cells were used to determine drug uptake, and 7RH8/32 showed the maximum intracellular uptake (10.89%). After storage under accelerated and normal conditions (3 and 6 months), the selected formulations remained stable. The developed formulations can be used to improve NP solubility and absorption.

Impact of Fixed Oil on Ostwald Ripening of Anti-Oral Cancer Nanoemulsions Loaded with Amomum kravanh Essential Oil.

Recently, essential oil from Amomum kravanh (AMO) was reported to exert anti-oral cancer effects. Although it was more effective after being loaded into nanoemulsions, AMO without an Ostwald ripening inhibitor was unable to form stable nanoemulsions because of the Ostwald ripening phenomenon. In this study, we examined the influence of Ostwald ripening inhibitors, such as fixed oils and polyethylene glycol 4000 (PEG 4000), on nanoemulsion properties prepared by a phase inversion temperature method. Several fixed oils, including virgin coconut oil (VCO), palm oil (PMO), olive oil (OLO), and PEG 4000, were evaluated, and their Ostwald ripening inhibitory effects were compared. The results suggest that the type and ratio of AMO:fixed oils influence the formation and characteristics of nanoemulsions. PEG 4000 was unable to produce nanoemulsions; however, stable nanoemulsions with small droplet sizes were observed in preparations containing OLO and VCO at an AMO:fixed oil ratio of 80:20, which may be the result of specific molecular interactions among the components. Using an MTT assay, we demonstrated that the AMO:OLO (80:20) nanoemulsion produced the most significant cytotoxic effect on oral cancer cells with a percentage of 99.68 ± 0.56%. Furthermore, the AMO:OLO 80:20 nanoemulsion inhibits metastasis and induces oral cancer cell death through the intrinsic apoptosis pathway. In conclusion, AMO nanoemulsion with anti-oral cancer activity was successfully produced by varying the amount and type of fixed oils. In the future, this discovery may lead to the development of stable nanoemulsions employing additional volatile oils.

Monolaurin-Loaded Gel-Like Microemulsion for Oropharyngeal Candidiasis Treatment: Structural Characterisation and In Vitro Antifungal Property.

Recently, monolaurin (ML) has received great interest due to its possible use as an alternative antifungal. However, the limited water solubility of ML is still a major obstacle to its formulation and application. Gel-like microemulsions are one of the promising carriers for low-water-solubility substances due to both the advantages of gels and microemulsions and may be applied for ML. In this study, ML was incorporated into gel-like microemulsions and evaluated for its physicochemical and antifungal properties. The results indicated that the properties of gel-like microemulsion changed after the incorporation of ML, suggesting that ML can induce the transition of internal structure. When simulating the oral cavity environment, changes in the microstructure were observed and depended on the times of dilution. The lamellar structure was formed at 1.5-2 times dilution. However, this structure was disrupted after dilution five times or more. The structural change following dilution was associated with the release profiles. After contacting the formulations with the medium, ML was promptly released, with the majority of ML being released within 2 h. Regarding the antifungal assay, the ML-loaded gel-like microemulsions decreased the survival of Candida albicans within 3 h, although ML was immediately released, suggesting that the ML-loaded in oil droplets required time to permeate through the fungal cell wall. Additionally, the gel-like microemulsions possessed acceptable stability after the temperature cycling test. Therefore, gel-like microemulsions can be a possible carrier for ML loading, and ML-loaded gel-like microemulsions may be applied as an alternative antifungal preparation in the future. Graphical abstract.

Improvement in Solubility and Absorption of Nifedipine Using Solid Solution: Correlations between Surface Free Energy and Drug Dissolution.

Ternary solid solutions composed of nifedipine (NDP), amino methacrylate copolymer (AMCP), and polysorbate (PS) 20, 60, or 65 were prepared using a solvent evaporation method. The dissolution profiles of NDP were used to study the effect of the addition of polysorbate based on hydrophilic properties. A solid solution of NDP and AMCP was recently developed; however, the dissolution of NDP was <70%. In the present study, polysorbate was added to improve the dissolution of the drug by altering its hydrophilicity. The suitable formulation contained NDP and AMCP at a ratio of 1:4 and polysorbate at a concentration of 0.1%, 0.3%, or 0.6%. Differential scanning calorimetry and powder X-ray diffraction were used to examine the solid solutions. No peak representing crystalline NDP was observed in any solid solution samples, suggesting that the drug was molecularly dispersed in AMCP. The NDP dissolution from NDP powder and solid solution without PS were 16.82% and 58.19%, respectively. The highest dissolution of NDP of approximately 95.25% was noted at 120 min for the formulation containing 0.6% PS20. Linear correlations were observed between the surface free energy and percentages of dissolved NDP (R2 = 0.7115-0.9315). Cellular uptake across Caco-2 was selected to determine the drug permeability. The percentages of cellular uptake from the NDP powder, solid solution without and with PS20 were 0.25%, 3.60%, and 7.27%, respectively.

Enhancing oral absorption of poorly water-soluble herb (Kaempferia parviflora) extract using self-nanoemulsifying formulation.

Kaempferia parviflora, a medicinal herb, treats hypertension and promotes longevity with good health and well-being. Its bioactive component is poorly soluble in water, resulting in poor absorption. This study aimed to enhance the bioavailability of K. parviflora dichloromethane (KPD) extract using a self-nanoemulsifying drug delivery system (SNEDDS). KPD was dissolved in diethylene glycol monoethyl, polyoxyl-35 castor oil and caprylic/capric glyceride, and clear yellow SNEDDS solution was obtained. The methoxyflavone markers were used for content and dissolution analysis. Solid SNEDDS was prepared by stepwise mixing of KPD using a mortar and pestle (1:1 ratio) with five solid carriers: Aerosil® 200, Florite® RE, Neusilin® US2 (NEUS), Fujicalin®, and Neusilin® UFL2. The USP apparatus II with simulated gastric fluid USP (SGF without pepsin, pH 1.2) was used in order to perform the in vitro dissolution. The methoxyflavones dissolution at 60 min from KPD, SEDDS, and SNEDDS/NEUS were approximately 16, 92, and 73%, respectively. The pharmacokinetic profiles of methoxyflavones for oral administration were studied using Wistar rats; the areas under the curve of SNEDDS/NEUS (1.77-fold) and SNEDDS (5.38-fold) were significantly higher than that of KPD. The developed formulations showed good stability after storage for 6 months under accelerated and normal conditions.

Design and characterisation of electrospun shellac-polyvinylpyrrolidone blended micro/nanofibres loaded with monolaurin for application in wound healing.

Due to their ultrafine network structures, electrospun nanofibres have been potentially used for wound application. In order to develop a desired wound dressing material, shellac (SHL) was blended with polyvinyl pyrrolidone (PVP). Monolaurin (ML), which is a natural antimicrobial lipid, was incorporated into the SHL-PVP blended fibres to prevent delayed wound healing resulting from microbial infection. A full factorial design with three replicated centre points was employed in order to determine the main and interaction effects of various factors including SHL ratio in SHL-PVP blended solution, ML content and applied voltage on the multiple responses such as morphology, surface wettability, absorbency and mechanical properties. According to the results, an increase in the PVP content could lead to a significant increase in tensile strength and elongation. In addition, the presence of PVP contributed to an improvement in the drug loading capacity and dissolution rate. The fabricated fibres loaded with ML exhibited an excellent activity against Staphylococcus aureus and Candida albicans, and also provided an enhanced ability in the cell adhesion. Therefore, SHL-PVP blended fibres loaded with ML might be effectively used for application in wound healing.

Fabrication and characterization of spearmint oil loaded nanoemulsions as cytotoxic agents against oral cancer cell.

Spearmint oil (SMO), a commonly used essential oil for oral care products, possesses various interesting functions, especially for anticancer property. However, the application of SMO for cancer treatment is limited due to water insoluble. In the present study, nanoemulsions, which have been widely accepted as dosage forms for poorly water-soluble drugs, were selected as candidate carriers for SMO to inhibit oral cancer cell. The nanoemulsions were fabricated using phase inversion temperature method. The factors affecting formation and properties of nanoemulsions including type and amount of surfactants, oil loading and ratio of SMO to virgin coconut oil (VCO) were investigated. Among the surfactants used, the nanoemulsions containing polyoxyethylene castor oil derivatives (Kolliphor®EL; PCO35, Cremophor®RH40; PCO40, Eumulgin®CO60; PCO60) and polyoxyethylene sorbitan fatty acid esters (PSF80) showed 100% creaming after temperature cycling test indicating excellent physical stability while those containing PCO40 demonstrated more transparency and better physical stability. With an increasing amount of PCO40, the droplet size tended to decrease and was in the nano-size range (<1000 nm) after increasing to more than 5% (w/w). SMO-VCO loading also influenced on the droplet size. At 5% (w/w) PCO40, the maximum SMO-VCO loading of 25% (w/w) to attain nanoemulsions was observed. Moreover, the composition of oils had an impact on size of emulsions. The transparent nanoemulsions were only prepared in the range of SMO-VCO from 40:60 to 80:20, suggesting the optimum ratio of SMO to surfactant and the composition of oils were the critical factors for formation of nanoemulsions. NMR study disclosed that the interaction between PCO40 with both VCO and SMO should be a possible stabilization mechanism. Furthermore, the SMO-VCO nanoemulsions exhibited significant cytotoxic effect against oral carcinoma (KON) cell line using MTT assay. The finding, therefore, revealed the good feasibility of SMO-VCO nanoemulsions as novel carriers for treating of oral cancer.

Improved dissolution of Kaempferia parviflora extract for oral administration by preparing solid dispersion via solvent evaporation.

Kaempferia parviflora, a plant in the family Zingiberaceae, has been used in Thai traditional medicines for treating hypertension and promoting longevity with good health and well-being. However, its limited aqueous solubility and low dissolution restrict its bioavailability. The aim of the study was therefore to improve the dissolution rate of K. parviflora extracted with dichloromethane (KPD) by solid dispersions. Different water-soluble polymers were applied to improve dissolution of KPD. The solid dispersions in different ratios were prepared by solvent evaporation method. Only hydroxypropyl methylcellulose (HPMC) and polyvinyl alcohol-polyethylene glycol grafted copolymer (PVA-co-PEG) could be used to produce homogeneous, powdered solid dispersions. Physical characterization by scanning electron microscopy, hot stage microscopy, differential scanning calorimetry and powder X-ray diffractometry, in comparison with corresponding physical mixtures, showed the changes in solid state during the formation of solid dispersions. Dissolution of a selected marker, 5,7,4'-trimethoxyflavone (TMF), from KPD/HPMC and KPD/PVA-co-PEG solid dispersions was significantly improved, compared with pure KPD. The dissolution enhancement by solid dispersion was influenced by both type and content of polymers. The stability of KPD/HPMC and KPD/PVA-co-PEG solid dispersions was also good after 6-month storage in both long-term and accelerated conditions. These results identified that the KPD/HPMC and KPD/PVA-co-PEG solid dispersions were an effective new approach for pharmaceutical application of K. parviflora.