Melatonin and its derivative disrupt cancer stem-like phenotypes of lung cancer cells via AKT downregulation.

Cancer stem cells (CSCs), a small subpopulation of tumour cells, have properties of self-renewal and multipotency, which drive cancer progression and resistance to current treatments. Compounds potentially targeting CSCs have been recently developed. This study shows how melatonin, an endogenous hormone synthesised by the pineal gland, and its derivative suppress CSC-like phenotypes of human non-small cell lung cancer (NSCLC) cell lines, H460, H23, and A549. The effects of MLT and its derivative, acetyl melatonin (ACT), on CSC-like phenotypes were investigated using assays for anchorage-independent growth, three-dimensional spheroid formation, scratch wound healing ability, and CSC marker and upstream protein signalling expression. Enriched CSC spheroids were used to confirm the effect of both compounds on lung cancer cells. MLT and ACT inhibited CSC-like behaviours by suppression of colony and spheroid formation in NSCLC cell lines. Their effects on spheroid formation were confirmed in CSC-enriched H460 cells. CSC markers, CD133 and ALDH1A1, were depleted by both compounds. The behaviour and factors associated to epithelial-mesenchymal transition, as indicated by cell migration and the protein vimentin, were also decreased by MLT and ACT. Mechanistically, MLT and ACT decreased the expression of stemness proteins Oct-4, Nanog, and ß-catenin by reducing active AKT (phosphorylated AKT). Suppression of the AKT pathway was not mediated through melatonin receptors. This study demonstrates a novel role, and its underlying mechanism, for MLT and its derivative ACT in suppression of CSC-like phenotypes in NSCLC cells, indicating that they are potential candidates for lung cancer treatment.

Anthocyanin complex niosome gel accelerates oral wound healing: In vitro and clinical studies.

An anthocyanin complex (AC), composed of extracts of purple waxy corn and blue butterfly pea petals, and AC niosomes, bilayered vesicles of non-ionic surfactants, were compared in in vitro and clinical studies. Cultured fibroblasts subjected to a scratch wound were monitored for cell viability, cell migration, nuclear morphology and protein expression. Scratched cells showed accelerated wound healing activity, returning to normal 24 h after treatment with AC niosomes (0.002 mg/mL). Western blots and immunocytochemistry indicated upregulation of type I, III and IV collagens, fibronectin and laminins in AC niosome-treated scratched cells. A randomized block placebo-controlled double-blind clinical trial in 60 volunteers (18-60 years old) with oral wounds indicated that AC niosome gel accelerated wound closure, reduced pain due to the oral wounds and improved participants' quality of life more than AC gel, triamcinolone gel and placebo gel. These data are consistent with enhanced delivery of AC to fibroblasts by use of niosomes. AC niosomes activated fibroblasts within wounded regions and accelerated wound healing, indicating that AC niosomes have therapeutic potential.

Anti-Inflammatory Comparison of Melatonin and Its Bromobenzoylamide Derivatives in Lipopolysaccharide (LPS)-Induced RAW 264.7 Cells and Croton Oil-Induced Mice Ear Edema.

The pineal gland is a neuroendocrine organ that plays an important role in anti-inflammation through the hormone melatonin. The anti-inflammatory effects of melatonin and its derivatives have been reported in both in vitro and in vivo models. Our previous study reported the potent antioxidant and neuroprotective activities of bromobenzoylamide substituted melatonin. In silico analysis successfully predicted that melatonin bromobenzoylamid derivatives were protected from metabolism by CYP2A1, which is a key enzyme of the melatonin metabolism process. Therefore, the anti-inflammatory activities of melatonin and its bromobenzoylamide derivatives BBM and EBM were investigated in LPS-induced RAW 264.7 macrophages and croton oil-induced ear edema in mice. The experiments showed that BBM and EBM significantly reduced production of the inflammatory mediators interleukin-6 (IL-6), prostaglandin E2 (PGE2), and nitric oxide (NO) in a dose-dependent manner, but only slightly affected TNF-α in LPS-induced RAW 264.7 macrophages. This suggests that modifying melatonin at either the N1-position or the N-acetyl side chain affected production of NO, PGE2 and IL-6 in in vitro model. In the croton oil-induced mouse ear edema model, BBM, significantly decreased ear edema thickness at 2-4 h. It leads to conclude that bromobenzoylamide derivatives of melatonin may be one of the potential candidates for a new type of anti-inflammatory agent.

Inhibitory Effects of Erythrosine/Curcumin Derivatives/Nano-Titanium Dioxide-Mediated Photodynamic Therapy on Candida albicans.

This study focuses on the role of photosensitizers in photodynamic therapy. The photosensitizers were prepared in combinations of 110/220 µM erythrosine and/or 10/20 µM demethoxy/bisdemethoxy curcumin with/without 10% (w/w) nano-titanium dioxide. Irradiation was performed with a dental blue light in the 395-480 nm wavelength range, with a power density of 3200 mW/cm2 and yield of 72 J/cm2. The production of ROS and hydroxyl radical was investigated using an electron paramagnetic resonance spectrometer for each individual photosensitizer or in photosensitizer combinations. Subsequently, a PrestoBlue® toxicity test of the gingival fibroblast cells was performed at 6 and 24 h on the eight highest ROS-generating photosensitizers containing curcumin derivatives and erythrosine 220 µM. Finally, the antifungal ability of 22 test photosensitizers, Candida albicans (ATCC 10231), were cultured in biofilm form at 37 °C for 48 h, then the colonies were counted in colony-forming units (CFU/mL) via the drop plate technique, and then the log reduction was calculated. The results showed that at 48 h the test photosensitizers could simultaneously produce both ROS types. All test photosensitizers demonstrated no toxicity on the fibroblast cells. In total, 18 test photosensitizers were able to inhibit Candida albicans similarly to nystatin. Conclusively, 20 µM bisdemethoxy curcumin + 220 µM erythrosine + 10% (w/w) nano-titanium dioxide exerted the highest inhibitory effect on Candida albicans.

Oral Administration of Melatonin or Succinyl Melatonin Niosome Gel Benefits 5-FU-Induced Small Intestinal Mucositis Treatment in Mice.

Mucositis is one of the most adverse effects of 5-fluorouracil (5-FU) and had no standard drug for treatment. Melatonin is a neurohormone, and can ameliorate radiotherapy-induced small intestinal mucositis. Melatonin encapsulated in niosomes improved its poor bioavailability. Succinyl melatonin, a melatonin derivative, showed prolonged release compared with melatonin. This study investigated the efficacy of melatonin niosome gel (MNG) and succinyl melatonin niosome gel (SNG) in 5-FU-induced small intestinal mucositis treatment in mice. MNG and SNG with particle sizes of 293 and 270 nm were shown to have mucoadhesive potentials. The effect of a daily oral application of MNG, SNG, or fluocinolone acetonide gel (FAG, positive control) was compared to that of the normal group. The body weight, food consumption, histology, Fourier transform infrared (FTIR) spectroscopy, inflammatory cytokines (tumor necrosis factor (TNF)-α and interleukin (IL)-1ß), and malondialdehyde (MDA) in the small intestine were monitored. The results showed decreased %body weight and food consumption in all 5-FU-injected groups compared with the normal group. The MNG and SNG treatments maintained the food consumption and the normal integrity of the small intestines, as evidenced by villus length and crypt depth, similar to the observations in the normal groups. The FTIR spectra showed no change in lipids of the MNG and SNG groups compared with the normal group. Moreover, SNG could reduce IL-1ß content to a level that was not different from the level in the normal groups. Therefore, the oral application of MNG and SNG could protect against 5-FU-induced small intestinal mucositis in mice.

Topical Niosome Gel Containing an Anthocyanin Complex: a Potential Oral Wound Healing in Rats.

Anthocyanins from dietary sources showing potential benefits as anti-inflammatory in oral lesions were developed as an anthocyanin complex (AC), comprised of extracts of Zea mays (CC) and Clitoria ternatea (CT), and formulated into a niosome gel to prove its topical oral wound healing in vitro and in vivo investigations. The AC formed nano-sized clusters of crystalline-like aggregates, occurring through both intra- and inter-molecular interactions, resulting in delivery depots of anthocyanins, following encapsulation in niosomes and incorporation into a mucoadhesive gel. In vitro permeation of anthocyanins was improved by complexation and further enhanced by encapsulation in niosomes. Collagen production in human gingival fibroblasts was promoted by AC and AC niosomes, but not CC or CT. The in vivo wound healing properties of AC gel (1 and 10%), AC niosome gel (1 and 10%), fluocinolone acetonide gel, and placebo gel were investigated for incisional wounds in the buccal cavities of Wistar rats. AC gel and AC niosome gel both reduced wound sizes after 3 days. AC niosome gel (10%) gave the highest reduction in wound sizes after day 3 (compared to fluocinolone acetonide gel, p < 0.05), and resulted in 100% wound healing by day 5. Histological observations of cross-sectioned wound tissues revealed the adverse effects of fluocinolone gel and wound healing potential of AC niosome gel. Topical application of AC niosome gel exhibited an anti-inflammatory effect and promoted oral wound closure in rats, possibly due to the improved mucosal permeability and presence of delivery depots of AC in the niosome gel.

Topical Niosome Gel of Zingiber cassumunar Roxb. Extract for Anti-inflammatory Activity Enhanced Skin Permeation and Stability of Compound D.

An extract of Zingiber cassumunar Roxb. (ZC) was encapsulated in niosomes of which a topical gel was formed. (E)-4-(3',4'-dimethoxyphenyl)but-3-en-1-ol or compound D detected by a gradient HPLC was employed as the marker and its degradation determined to follow zero-order kinetics. Niosomes significantly retarded thermal-accelerated decomposition of compound D in the gel (p < 0.05) but did not change the activation energy of compound D. Niosomes enhanced in vitro permeation rate of compound D from the gel. Topical applications of ZC noisome gel gave a faster change in tail flick latency than piroxicam gel and hydrocortisone cream (p < 0.05) while there were insignificant differences in anti-inflammatory activity up to 6 h using croton oil-induced ear edema model in mice (p > 0.05). Thus, encapsulation of ZC extract in niosomes enhanced chemical stability and skin permeation with comparable topical anti-inflammatory effects to steroid and NSAID.

In vitro cytoprotective and in vivo anti-oral mucositis effects of melatonin and its derivatives.

According to our preliminary study, melatonin and its N-amide derivatives (N-(2-(1-4-bromobenzoyl-5-methoxy-1H-indol-3-yl)ethyl)acetamide (BBM) and 4-bromo-N-(2-(5-methoxy-1H-indol-3-yl)ethyl)benzamide (EBM)) inhibited the marker of acute inflammation in tests in vitro and in vivo. The anti-inflammatory agent is intended for the prevention and treatment of chemotherapy-induced toxicity. In this study aimed to evaluate the effect of melatonin and its derivatives on mechanisms related to chemotherapy-induced oral mucositis by in vitro ROS and 5-FU-induced human keratinocyte cells as well as in vivo oral mucositis model. In in vitro H2O2-induced HaCaT cells, BBM had the highest level of protection (34.57%) at a concentration 50 µM, followed by EBM (26.41%), and melatonin (7.9%). BBM also protected cells against 5-FU-induced to 37.69-27.25% at 12.5-100 µM while EBM was 36.93-29.33% and melatonin was 22.5-11.39%. In in vivo 5-FU-induced oral mucositis in mice, melatonin, BBM, and EBM gel formulations protected tissue damage from 5-FU similar to the standard compound, benzydamine. Moreover, the weight of mice and food consumption recovered more quickly in the BBM group. These findings suggested that it was possible to develop BBM and EBM as new therapeutic agents for the treatment of oral mucositis.

Melatonin inhibits cholangiocarcinoma and reduces liver injury in Opisthorchis viverrini-infected and N-nitrosodimethylamine-treated hamsters.

The human liver fluke Opisthorchis viverrini infection and N-nitrosodimethylamine (NDMA) administration induce cholangiocarcinoma (CCA) and liver injury in hamsters. Melatonin protects against liver injury and reduces the alteration of mitochondrial structure, mitochondrial membrane potential, and mitochondrial pro- and anti-apoptotic pathways in various cancer types. To investigate the chemopreventive effect of melatonin on CCA genesis and liver injury, hamsters were treated with a combination of O. viverrini infection and NDMA concurrently administered with melatonin (10 mg/kg and 50 mg/kg) for 120 days. Melatonin treatment at 50 mg/kg caused a significant reduction in liver/body weight ratios and decreased tumor volumes leading to an increase in the survival of animals. In the tumorous tissues, the high-dose melatonin reduced DNA fragmentation and mitochondrial apoptosis by inducing anti-apoptotic protein (Bcl-2) in the mitochondrial fraction and down-regulating cytochrome c, pro-apoptotic protein (Bax), and caspase-3 in tumor cytosol. Moreover, a high-dose melatonin treatment significantly increased mitochondrial antioxidant enzymes and prevented mitochondrial ultrastructure changes in the tumor. Overall, melatonin has potent chemopreventive effects in inhibiting CCA genesis and also reduces liver injury in hamster CCA, which, in part, might involve in the suppression of CCA by reducing tumor mitochondria alteration.

Corrigendum to "Curcumin-loaded nanocomplexes: Acute and chronic toxicity studies in mice and hamsters" [Toxicol. Rep. 8 (2021) 1346-1357/ S221475002100127X].

[This corrects the article DOI: 10.1016/j.toxrep.2021.06.021.].

Polyphenol and Tryptophan Contents of Purple Corn (Zea mays L.) Variety KND and Butterfly Pea (Clitoria ternatea) Aqueous Extracts: Insights into Phytochemical Profiles with Antioxidant Activities and PCA Analysis.

Plants are a rich source of phytochemical compounds with antioxidant activity. Several studies have revealed that the consumption of plant polyphenols reduces the risk of diseases. Purple corn (Zea mays L. variety KND) and butterfly pea (Clitoria ternatea; CT) were selected to be investigated as alternative natural polyphenol sources to increase the value of these plants. Phytochemical profiles and antioxidant activities of KND cob, silk, husk and CT extracts alone and in combination were investigated in this study. The results revealed that purple corn cob (C) extract had the highest tryptophan, melatonin, total anthocyanin (TAC) and delphinidin content, while the purple corn silk (S) extract showed the highest total phenolic content (TPC) and antioxidant activities. Serotonin was found only in purple corn husk (H) extract and C extract. High contents of tryptophan and sinapic acid were found in CT extract. Principal component analysis (PCA) revealed that strong antioxidant activities were strongly correlated with protocatechuic acid and p-hydroxybenzoic acid contents, moderate antioxidant activities were strongly correlated with melatonin, and low antioxidant activities were strongly correlated with sinapic acid content. Therefore, the purple corn variety KND waste cobs, silk and husks are a potentially rich source of health-promoting phytochemical compounds.

Synthesis and evaluation of 1-(substituted)-3-prop-2-ynylureas as antiangiogenic agents.

Novel urea derivatives of alkynes have been designed, synthesized, and evaluated as potential cancer therapeutics leads. The most active 1-((3-chloromethyl)phenyl)-3-prop-2-ynylurea (1) exhibited cytotoxic effect against HELA and MCF-7 cell lines with IC(50) values of 1.55 µM and 1.48 µM, respectively. Further investigation on tube formation assay in human vein umbilical cells (HUVEC) demonstrated that 1 and methyl 4-(3-(3-ethynylureido)benzyloxy) benzoate (6) possess antiangiogenic activity, with minimum effective dose of 25 nM (for 1) and 6.25 µM (for 6). The ED(50) of 1 and 6 were found to be 0.26 µM and 17.52 µM, respectively. The results from in vitro tyrosine kinase assay indicated the EGFR inhibition of 1 over other kinases (VEGFR2, FGFR1 and PDGFRß). The cytotoxicity of 1 against EGFR overexpressing cell line A431 (IC(50) 36 nM) was comparable to that of erlotinib. The binding mode of 1 from docking simulation in the EGFR active site revealed that the urea motif formed hydrogen bonding with Lys745, Thr854 and Asp855 in hydrophobic pocket of EGFR. Compound 1 is considered as a potential lead for further optimization.

Polymeric Micelles Enhance Mucosal Contact Time and Deposition of Fluocinolone Acetonide.

This study used polymeric micelles to improve quality by increasing drug solubility, extending mucosal drug retention time, enhancing mucoadhesiveness, and promoting drug permeation and deposition. Fluocinolone acetonide (FA) was loaded into polymeric micelles (FPM), which were composed of poloxamer 407 (P407), sodium polyacrylate (SPA), and polyethylene glycol 400, and their physicochemical properties were examined. Small-angle X-ray scattering (SAXS) revealed a hexagonal micellar structure at all temperatures, and the concentrations of P407 and SPA were shown to significantly affect the solubility, mucoadhesion, release, and permeation of FPMs. The proportion of P407 to PEG at a ratio of 7.5:15 with or without 0.1% w/v of SPA provided suitable FPM formulations. Moreover, the characteristics of FPMs revealed crystalline states inside the micelles, which was consistent with the morphology and nano-hexagonal structure. The results of ex vivo experiments using focal plane array (FPA)-based Fourier transform infrared (FTIR) imaging showed that the FPM with SPA penetrated quickly through the epithelium, lamina propria, and submucosa, and remained in all layers from 5-30 min following administration. In contrast, the FPM without SPA penetrated and passed through all layers. The FPM with extended mucoadhesion, improved drug-mucosal retention time, and increased FA permeation and deposition were successfully developed, and could be a promising innovation for increasing the efficiency of mouth rinses, as well as other topical pharmaceutical and dental applications.

Development of a Sericin Hydrogel to Deliver Anthocyanins from Purple Waxy Corn Cob (Zea mays L.) Extract and In Vitro Evaluation of Anti-Inflammatory Effects.

Sericin-alginate hydrogel formulations with purple waxy corn (Zea mays L.) cob extract (PWCC) for topical anti-inflammatory application are developed and evaluated. The physical properties such as viscosity, pH, and anthocyanin release are examined and in vitro anti-inflammatory activities, such as NO inhibition and IL-6, IL-1ß, TNF-α, iNOS, and COX-2 expression, are evaluated in LPS-stimulated RAW 264.7 murine macrophages. The sericin-alginate hydrogel is prepared by physical crosslinking through the ionic interaction of the polymers combined with anthocyanin from PWCC at pH 6.5. The hydrogel formulation with 2.00% w/v sericin, 0.20% w/v alginate, and 0.15% w/v PWCC (SN6) shows a suitable viscosity for topical treatment, the highest nitric oxide inhibition (79.43%), no cytotoxicity, and reduced expression of IL-6, IL-1ß, and TNF-α mediators. Moreover, the SN6 formulation displays a sustained anthocyanin release over 8-12 h, which correlates with the Korsmeyer-Peppas model. The FT-IR spectrum of SN6 confirmed interaction of the sericin polymer with anthocyanins from PWCC via H-bonding by the shifted peak of amide I and amide III. In addition, the anthocyanin is stable in sericin hydrogels under heating-cooling storage conditions. Therefore, we suggest that this hydrogel formulation has potential as an anti-inflammatory agent. The formulation will be further investigated for in vivo studies and clinical trials in the future.

Efficacy and safety of two artificial saliva-based polymers containing 0.1% pilocarpine for treatment of xerostomia: A randomized clinical pilot trial.

BACKGROUND: Topical agents are the mainstay in the treatment of xerostomia, a common complaint most frequently associated with salivary dysfunction. This study aimed to compared the efficacy and safety for xerostomia treatment of 2 artificial saliva preparations containing 0.1% pilocarpine, and, either sodium carboxymethylcellulose (SCMC), or, sodium polyacrylate (SPA). MATERIAL AND METHODS: Thirty-one xerostomia patients were randomly allocated into either a SCMC-treated group (15 patients), or, a SPA-treated group (16 patients). The formulations were taken 0.5 ml, 4 times daily for 6 weeks and double-blinded assessed before and after treatments using Xerostomia Inventory (XI) and Clinical Oral Dryness Score (CODs). Unstimulated and stimulated whole salivary flow rates were measured. RESULTS: After treatment, the SCMC-treated group had significantly lower CODs and higher unstimulated and stimulated whole salivary flow rates (p<0.001, p=0.035, and p=0.013, respectively), while the SPA-treated group showed significantly lower CODs only (p=0.004). In contrast, SCMC-treated and SPA-treated groups at the 6th week after treatments showed non-significant differences in all assessments (p>0.05, all). Some adverse events (AEs) were reported, e.g., burning tongue, dizziness and watery eyes, but no severe AEs. CONCLUSIONS: This randomized controlled pilot trial demonstrated superior efficacy of SCMC-formula over a SPA-formula after 6 weeks of xerostomia treatment. These formulations with topical pilocarpine proved safe in clinical use with minimal reported AE. Key words:Xerostomia, artificial saliva, sodium carboxymethylcellulose, sodium polyacrylate, pilocarpine.

Topical Melatonin Niosome Gel for the Treatment of 5-FU-Induced Oral Mucositis in Mice.

BACKGROUND: Oral mucositis, one of the most common complications of 5-Fluorouracil (5-FU) treatment, leads to several problems, including pain, diarrhea and malnutrition, and reduces the quality of life and subsequent treatments. Melatonin, a neurohormone with anti-inflammatory and antioxidant activities, was encapsulated in niosomes and embedded in a mucoadhesive gel formulation as a Melatonin Niosome Gel (MNG) to perform oral mucositis treatment. OBJECTIVE: This study aimed to investigate the effectiveness of MNG for the treatment of 5-FU-induced oral mucositis in mice. METHODS: Oral mucositis was induced in ICR mice by 5-FU and randomly assigned to receive daily applications of the topical oral MNG, a fluocinolone acetonide gel, a blank niosome gel, or no treatment for 5 days in comparison with a normal group. Average body weights, food consumption, and behaviors of the mice as well as microscopic histopathology, Fourier-Transform Infrared Spectroscopy (FTIR) analysis, proinflammatory cytokine levels, and oxidative stress markers of the tongues were monitored and collected after sacrifice. RESULTS: In comparison to the normal group, the average body weights of the 5-FU-MNG mice did not deviate from that of the normal group, nor was there a significant difference in the time to sleep or licking (p>0.05 for both parameters). In addition, the mice treated with MNG and fluocinolone acetonide did not show significantly different histopathological, FTIR, interleukin-1ß or malondialdehyde (MDA) results in the tongues used as the oral tissue samples. CONCLUSION: Topical MNG potentially inhibits inflammation and lipid oxidative stress in 5-FU-induced oral mucositis.

Curcumin-loaded nanocomplexes: Acute and chronic toxicity studies in mice and hamsters.

We recently developed a modified solid dispersion of curcumin-loaded nanocomplexes (CNCs) in gums which promoted the prolonged and sustained release of curcumin. However, its safety assessment has not yet been investigated. Here, acute and chronic toxicities of CNCs were assayed using mice and hamsters. CNCs were orally administered to the animals. Doses of CNCs used for acute toxicity testing were 0.1, 1.1, 11.0 g/kg body weight for mice and 0.2, 2.1 and 21.4 g/kg body weight for hamsters. Doses of CNCs for chronic toxicity testing were 0.09, 0.27, 0.8 g/kg body weight/day for mice and 0.18, 0.54 and 1.61 g/kg body weight/day for hamsters. This regimen was followed daily for 6 months. Low and medium doses of CNCs did not induce any side effects in acute and chronic toxicity tests in either animal species. However, in acute toxicity testing, the organ-weight to body-weight ratio of spleen was significantly increased in mice treated with 11 g/kg body weight along with elevated levels of some biochemical parameters. There was a significant increase in organ-weight to body-weight ratios of stomach, liver and heart in hamsters treated with 21.4 g/kg body weight, but no elevated levels of biochemical parameters. Oral LD50 of CNCs in mice and hamsters were 8.9 and 16.8 g/kg body weight (equivalent to 2.5 and 4.7 g curcumin/kg body weight), respectively. Daily CNCs high-dose treatment for 6 months significantly increased organ-weight to body-weight ratios of stomach and intestine in mice and of lung and heart in hamsters. Elevated levels of glucose, total protein, ALT, AST and globulin in mice, and increased levels of AST, but decrease in cholesterol, in hamsters were concurrently observed with inflammation in liver and lung. These abnormalities were resolved within 28 days after cessation of treatment. The no-observed-adverse-effect level of CNCs was determined at 0.27 and 0.54 g/kg body weight/day in mice and hamsters. In conclusion, toxicity of high-dose CNCs treatment was graded as very low, possibly due to the components of the nanocomplex.

Characteristic Evaluation of Gel Formulation Containing Niosomes of Melatonin or Its Derivative and Mucoadhesive Properties Using ATR-FTIR Spectroscopy.

Chitosan or polyvinyl pyrrolidone (PVP) were used in combination with hydroxypropyl methylcellulose (HPMC) and poloxamer 407 (P407) as gelling agents for oral drug delivery. The performance interaction with mucin of chitosan-composed gel (F1) and PVP-composed gel (F2) was compared using attenuated total reflectance-Fourier-transform infrared (ATR-FTIR) spectroscopy at controlled temperatures of 25 and 37 °C for 1 and 5 min. F1 containing niosome-entrapped melatonin or its derivatives was investigated for mucoadhesive interaction on mucosa by ATR-FTIR spectroscopy under the same conditions. The results showed that F1-treated mucin gave a significantly lower amide I/amide II ratio than untreated mucin and F2-treated mucin did within 1 min, suggesting improved rapid affinity between mucin and chitosan. The spectra of mucosa treated with F1 incorporating niosomes of melatonin or its derivatives showed peak shifts at C=O (amide I), N-H (amide II), and carbohydrate regions and an associated decrease in the amide I/amide II ratio and increase in the carbohydrate/amide II ratio. These results indicated electrostatic interaction and hydrogen bonding between chitosan and mucin on the mucosa. In conclusion, the molecular interaction between gels and mucin/mucosa detected at amide I and amide II of proteins and the carbohydrate region could lead to an improved mucoadhesive property of the gel on the mucosa.

Anti-inflammatory effect of photodynamic therapy using guaiazulene and red lasers on peripheral blood mononuclear cells.

INTRODUCTION: Photodynamic therapy improves oral mucositis treatment. The reactive oxygen species (ROS) generated from this reaction could contribute to an anti-inflammatory effect by suppressing inflammatory cells. OBJECTIVE: To evaluate the anti-inflammatory effect of photodynamic therapy using guaiazulene and a red laser in peripheral blood mononuclear cells (PBMCs). METHODS: Guaiazulene solutions (1, 2, 5, 25, 35, and 100 µM in 99.8 % methanol) were irradiated with red laser light (625 nm, 146.2 mW/cm2) in continuous mode at 0, 4, and 8 J/cm2 in black 96-well plates. ROS were measured using spin trapping technique with electron spin resonance (ESR) spectroscopy and fluorescence. The two highest concentrations were tested using cell viability (PrestoBlue®) and anti-inflammation (RANTES and PGE2 ELISA) assay kits. Kruskal-Wallis and Dunn Bonferroni tests were used for statistical analyses with significant differences at p-value < 0.05. RESULTS: Guaiazulene solutions between 2 and 5 µM exposed to red laser light at 4-8 J/cm2 generated significantly more singlet oxygen compared to the no guaiazulene group (p < 0.01) and reduced RANTES and PGE2 levels in TNF-α-inflamed peripheral blood mononuclear cells without affecting cell viability. CONCLUSION: Photodynamic activation of guaiazulene generated singlet oxygen and suppressed inflammatory markers in PBMCs.

Glutaryl Melatonin Niosome Gel for Topical Oral Mucositis: Anti- Inflammatory and Anticandidiasis.

BACKGROUND: Glutaryl melatonin, which is synthesized from melatonin and is a pineal glandderived neurohormone with anti-inflammatory and anti-oxidant properties, was comparatively investigated for its potential use as a topical anti-inflammatory agent. OBJECTIVE: Glutaryl melatonin, synthesized and screened for in vitro anti-candidiasis and in vitro and in vivo anti-inflammatory activities, was formulated as a niosome gel for topical oral evaluation in 5- fluorouracil-induced oral mucositis in mice. METHODS: In vitro anti-fungal activity in Candida albicans, in vitro anti-inflammatory activity in Escherichia coli liposaccharide-induced RAW cells and in vivo anti-inflammatory activity using a croton oilinduced ear edema model in ICR mice were investigated. Mucositis in mice (n= 6/group, 10-week-old mice) was induced by intraperitoneal injections of 5-fluorouracil, and the mice were subjected to a topical oral application of niosome gel containing melatonin (2% w/w) or glutaryl melatonin (2% w/w) and were compared with mice subjected to blank, fluocinolone acetonide (0.5% w/w) and control conditions. RESULTS: Glutaryl melatonin, at a 14.2 mM concentration, showed the highest fungicidal effect on C. albicans using the broth dilution method, indicating a nonsignificant difference from 1 µM of nystatin (p = 0.05). Nitric oxide, interleukin-6 and tumor necrosis factors were analyzed by ELISA. Liposaccharide-induced RAW cells were significantly reduced by glutaryl melatonin (p < 0.01). Ear edema inhibition of glutaryl melatonin was significant 1 h after application compared with that of melatonin (p = 0.03). Food consumption and body weight of the 5-fluorouracil-treated mice were significantly lower than those of the normal mice before all treatments (p < 0.05). Differences in the amount of licking behavior, which were observed in the control group for 5 min, were noticeable in the 5- fluorouracil-treated mice but not in the mice treated with the glutaryl melatonin niosome gel. CONCLUSION: Glutaryl melatonin exhibited mild anti-candidiasis and anti-inflammatory properties. The incorporation of glutaryl melatonin in a niosome gel formulation, demonstrated the potential for topical oral applications to reduce oral discomfort caused by 5-fluorouracil treatment in mice.