Factorial design based curcumin ethosomal nanocarriers for the skin cancer delivery: in vitro evaluation

Abstract

Melanoma is the most deadly and life-threatening form of skin cancer with progressively higher rates of incidence worldwide. The objective of the present investigation is to develop and to statistically optimize and characterize curcumin (CUR) loaded ethosomes for treatment of melanoma. A two factor, three level (3²) factorial design approach was employed for the optimization of ethosomes. The prepared ethosomes were evaluated for size, zeta potential, entrapment efficiency, in vitro skin permeation and deposition ability. The optimized ethosomal formulation was evaluated for in vitro cytotoxicity and cellular uptake studies using A375 human melanoma cells. The optimized formulation has imperfect round shaped unilamellar structures with a mean vesicle size of 247?±?5.25?nm and an entrapment efficiency of 92.24?±?0.20%. The in vitro skin permeation studies proved the superiority of ethosomes over the traditional liposomes in terms of the amount of drug permeated and deposited in skin layers. Fluorescence microscopy showed the enhanced penetration of ethosomes into the deeper layers of the skin. In vitro cytotoxicity and cellular uptake studies revealed that curcumin ethosomes have significantly improved cytotoxicity and cellular uptake in A375 human melanoma cell lines. The colony formation assay results showed that curcumin ethosomes have a superior antiproliferative effect as they effectively inhibit the clonogenic ability of A375 cells. The flow cytometry results indicate that curcumin ethosomes induce cell death in A375 cells by apoptosis mechanism. The present study provides a strong rationale and motivation for further investigation of newly developed curcumin ethosomes as a potential therapeutic strategy for melanoma treatment.     

Design and development of ethosomal transdermal drug delivery system of valsartan with preclinical assessment in Wistar albino rats

Abstract

Valsartan (VLT) is a highly selective and orally active antihypertensive drug. However, its oral administration is associated with drawbacks like low bioavailability. The objective of this study was to design and develop a transdermal delivery system for VLT using ethosomal carriers to investigate their enhanced transdermal delivery potential. VLT ethosomes were prepared by cold method. VLT ethosomes were characterized by scanning electron microscopy. The prepared ethanolic liposomes were characterized to be spherical having low polydispersity of nano-size range with good entrapment efficiency. ETC5 ethosomal suspension with 4% of phospholipon 90H and 40% of ethanol was found to have highest entrapment efficiency, i.e. 80.230?±?0.8748%. The permeation study of ethosomes was evaluated by ex vivo diffusion study through rat abdominal skin using Franz’s diffusion cells and ETC5 ethosomal suspension was found to have highest permeation with flux of 92.819?±?1.539?µg/cm²/h, when compared to the permeation profiles of drug solutions either in water or in a water–ethanol mixture. Transdermal application of ethosomal VLT on Wistar rats showed better and prolonged antihypertensive activity in comparison to orally administered VLT suspension by virtue of transdermal permeation through Wistar rat skin. Histopathological study of skin applied with ETC5 showed intercellular permeation across skin by dissolving intercellular lipids in epidermis without causing any rigorous changes in the skin cellular structure. In conclusion, ethosomes enabled the transdermal permeation of VLT, which amply proves its superiority over oral administration for antihypertensive treatment.     

Enhanced transdermal delivery of salbutamol sulfate via ethosomes

The main objective of the present work was to compare the transdermal delivery of salbutamol sulfate (SS), a hydrophilic drug used as a bronchodilator, from ethosomes and classic liposomes containing different cholesterol and dicetylphosphate concentrations. All the systems were characterized for shape, particle size, and entrapment efficiency percentage, by image analysis optical microscopy or transmission electron microscopy, laser diffraction, and ultracentrifugation, respectively. In vitro drug permeation via a synthetic semipermeable membrane or skin from newborn mice was studied in Franz diffusion cells. The selected systems were incorporated into Pluronic F 127 gels and evaluated for both drug permeation and mice skin deposition. In all systems, the presence of spherical-shaped vesicles was predominant. The vesicle size was significantly decreased (P < .05) by decreasing cholesterol concentration and increasing dicetylphosphate and ethanol concentrations. The entrapment efficiency percentage was significantly increased (P < .05) by increasing cholesterol, dicetylphosphate, and ethanol concentrations. In vitro permeation studies of the prepared gels containing the selected vesicles showed that ethosomal systems were much more efficient at delivering SS into mice skin (in terms of quantity and depth) than were liposomes or aqueous or hydroalcoholic solutions.     

Interaction of nanoparticles and cell-penetrating peptides with skin for transdermal drug delivery

Abstract

Topical or transdermal drug delivery is challenging because the skin acts as a natural and protective barrier. Therefore, several methods have been examined to increase the permeation of therapeutic molecules into and through the skin. One approach is to use the nanoparticulate delivery system. Starting with liposomes and other vesicular systems, several other types of nanosized drug carriers have been developed such as solid lipid nanoparticles, nanostructured lipid carriers, polymer-based nanoparticles and magnetic nanoparticles for dermatological applications. This review article discusses how different particulate systems can interact and penetrate into the skin barrier. In this review, the effectiveness of nanoparticles, as well as possible mode of actions of nanoparticles, is presented. In addition to nanoparticles, cell-penetrating peptide (CPP)-mediated drug delivery into the skin and the possible mechanism of CPP-derived delivery into the skin is discussed. Lastly, the effectiveness and possible mechanism of CPP-modified nanocarriers into the skin are addressed.     

Microemulsion for topical delivery of fenoprofen calcium: in vitro and in vivo evaluation

The aim of this study was to investigate microemulsion (ME) based topical delivery system for fenoprofen calcium (FPCa) to eliminate its oral gastrointestinal adverse effects. ME was prepared by the water titration method using oleic acid as oil phase, tween 80 as a surfactant and propylene glycol as a cosurfactant. Oleic acid was selected as oil phase due to its good solubilizing capacity. ME existence region was determined using pseudo-ternary phase diagrams for preparing different formulations. Six different formulations were selected with various values of oil (25–68%), water (2–3%), and the mixture of surfactant and cosurfactant (1:1) (24–67%). The selected ME formulae were characterized for optical birefringence, transmission electron microscopy (TEM), pH, % transmittance, electronic conductivity, drug content, droplet size, rheological properties and stability evaluation. In vitro release study of FPCa from ME s through the synthetic membrane and hairless rat skin were evaluated. The optimized formula ME5 consisting of 5% w/w FPCa, 60% w/w oleic acid as oil phase, 3% w/w aqueous phase, and 32% w/w of surfactant phase containing Tween 80 and propylene glycol (1:?1) showed the highest transdermal flux and highest skin permeation rate. Finally, the % inhibition of carrageenan-induced rat paw edema of the optimized formula ME5 was highly significant (p?0.001) as compared to plain gel of FPCa. In conclusion, ME is a promising technique for topical delivery of FPCa.     

Ethosomes for skin delivery of ropivacaine: preparation,characterization and ex vivo penetration properties

Abstract

Ropivacaine, a novel long-acting local anesthetic, has been proved to own superior advantage. However, Naropin® Injection, the applied form in clinic, can cause patient non-convenience. The purpose of this study was to formulate ropivacaine (RPV) in ethosomes and evaluate the potential of ethosome formulation in delivering RPV transdermally. The RPV-loaded ethosomes were prepared with thin-film dispersion technique and the formulation was characterized in terms of size, zeta potential, differential scanning calorimetry (DSC) analysis and X-ray diffraction (XRD) study. The results showed that the optimized RPV-ethosomes displayed a typical lipid bilayer structure with a narrow size distribution of 73.86?±?2.40?nm and drug loading of 8.27?±?0.37%, EE of 68.92?±?0.29%. The results of DSC and XRD study indicated that RPV was in amorphous state when encapsulated into ethosomes. Furthermore, the results of ex vivo permeation study proved that RPV-ethosomes could promote the permeability in a high-efficient, rapid way (349.0?±?11.5?μg?cm^?2 at 12?h and 178.8?±?7.1?μg?cm^?2 at 0.5?h). The outcomes of histopathology study forecasted that the interaction between ethosomes and skin could loosen the tight conjugation of corneocyte layers and weaken the permeation barrier. In conclusion, RPV-ethosomes could be a promising delivery system to encapsulate RPV and deliver RPV for transdermal administration.     

Topical delivery and in vivo antileishmanial activity of paromomycin-loaded liposomes for treatment of cutaneous leishmaniasis

The present study aimed to evaluate the potential of liposomes loaded with paromomycin (PA), an aminoglycoside antibiotic associated with poor skin penetration, for the topical treatment of cutaneous leishmaniasis (CL). Fluid liposomes were prepared and characterized for particle size, zeta potential, and drug entrapment. Permeation studies were performed with two in vitro models: intact and stripped skin. The antileishmanial activity of free and liposomal PA was evaluated in BALB/c mice infected by Leishmania (L.) major. Drug entrapment ranged from 10 to 14%, and the type of vesicle had little influence on this parameter. Particle size and polydispersity index of the vesicles composed by phosphatidylcholine (PC) and PC/cholesterol (Chol) ranged from of 516 to 362?nm and 0.7 to 0.4, respectively. PA permeation across intact skin was low, regardless of the formulation tested, while drug penetration into skin (percent of the applied dose) from PC (7.2?±?0.2%) and PC/Chol (4.8?±?0.2%) liposomes was higher than solution (1.9?±?0.1%). PA-loaded liposomes enhanced in vitro drug permeation across stripped skin and improved the in vivo antileishmanial activity in experimentally infected mice. Our findings suggest that the liposomes represent a promising alternative for the topical treatment of CL using PA.     

Enhanced plasmid DNA delivery using anionic LPDII by listeriolysin O incorporation

BACKGROUND: A major obstacle to achieving effective DNA-based therapeutics is efficient delivery of the DNA to its site of action in the cell. Upon internalization by endocytosis, the endosomal membrane represents a critical physical barrier preventing access of DNA to the cell cytosol. In order to overcome the membrane barrier and facilitate cytosolic entry, the endosomolytic bacterial protein listeriolysin O (LLO) is a potentially promising agent. METHODS: LLO was incorporated in an anionic liposome-entrapped polycation-condensed DNA delivery system (LPDII). Plasmid DNA was condensed using protamine sulfate and then complexed to anionic liposomes. LLO was incorporated into the delivery vehicle through encapsulation in anionic, pH-sensitive liposomes. Transfection levels were monitored using a model reporter plasmid encoding luciferase in P388D1 cells, a macrophage-like cell line. RESULTS: Transfection using the anionic LPDII delivery platform was enhanced through incorporation of LLO. Additionally, the net charge of the condensate, the lipid composition, and the total amount of LLO-liposomes were all capable of modulating the transfection levels of the vehicle. Importantly, in the presence of serum, transfection levels using the LLO-containing LPDII system were comparable to established cationic lipid delivery systems. CONCLUSIONS: LLO is capable of facilitating transfection using an anionic LPDII system. This anionic delivery vehicle represents the successful combination of the LPDII system for condensation of the DNA with the unique endosomolytic properties of LLO for improved transfection using plasmid DNA.     

Halobetasol propionate-loaded solid lipid nanoparticles (SLN) for skin targeting by topical delivery

The clinical use of halobetasol propionate (HP) is related to some adverse effects like irritation, pruritus and stinging. The purpose of this work was to construct HP-loaded solid lipid nanoparticles (HP-SLN) formulation with skin targeting to minimizing the adverse side effects and providing a controlled release. HP-SLN were prepared by solvent injection method and formula was optimized by the application of 3² factorial design. The nanoparticulate dispersion was evaluated for particle size and entrapment efficiency (EE). Optimized batch was characterized for differential scanning calorimetry (DSC), scanning electron microscopy, X-ray diffraction study and finally incorporated into polymeric gels of carbopol for convenient application. The nanoparticulate gels were evaluated comparatively with the commercial product with respect to ex-vivo skin permeation and deposition study on human cadaver skins and finally skin irritation study. HP-SLN showed average size between 200?nm and 84–94% EE. DSC studies revealed no drug-excipient incompatibility and amorphous dispersed of HP in SLN. Ex vivo study of HP-SLN loaded gel exhibited prolonged drug release up to 12?h where as in vitro drug deposition and skin irritation studies showed that HP-SLN formulation can avoid the systemic uptake, better accumulative uptake of the drug and nonirritant to the skin compared to marketed formulation. These results indicate that the studied HP-SLN formulation represent a promising carrier for topical delivery of HP, having controlled drug release, and potential of skin targeting with no skin irritation.     

Synthesis and characterization of ethosomal contrast agents containing iodine for computed tomography (CT) imaging applications

As a first step in the development of novel liver-specific contrast agents using ethosomes for computed tomography (CT) imaging applications, we entrapped iodine within ethosomes, which are phospholipid vesicular carriers containing relatively high alcohol concentrations, synthesized using several types of alcohol, such as methanol, ethanol, and propanol. The iodine containing ethosomes that were prepared using methanol showed the smallest vesicle size (392?nm) and the highest CT density (1107 HU). The incorporation of cholesterol into the ethosomal contrast agents improved the stability of the ethosomes but made the vesicle size large. The ethosomal contrast agents were taken up well by macrophage cells and showed no cellular toxicity. The results demonstrated that ethosomes containing iodine, as prepared in this study, have potential as contrast agents for applications in CT imaging.     

Evidence of curcumin and curcumin analogue effects in skin diseases: A narrative review

Curcumin, a natural polyphenolic and yellow pigment obtained from the spice turmeric, has strong antioxidative, anti-inflammatory, and antibacterial properties. Due to these properties, curcumin has been used as a remedy for the prevention and treatment of skin aging and disorders such as psoriasis, infection, acne, skin inflammation, and skin cancer. Curcumin has protective effects against skin damage caused by chronic ultraviolet B radiation. One of the challenges in maximizing the therapeutic potential of curcumin is its low bioavailability, limited aqueous solubility, and chemical instability. In this regard, the present review is focused on recent studies concerning the use of curcumin for the treatment of skin diseases, as well as offering new and efficient strategies to optimize its pharmacokinetic profile and increase its bioavailability.     

Preparation and Evaluation of Miconazole Nitrate-Loaded Solid Lipid Nanoparticles for Topical Delivery

The purpose of this study was to prepare miconazole nitrate (MN) loaded solid lipid nanoparticles (MN-SLN) effective for topical delivery of miconazole nitrate. Compritol 888 ATO as lipid, propylene glycol (PG) to increase drug solubility in lipid, tween 80, and glyceryl monostearate were used as the surfactants to stabilize SLN dispersion in the SLN preparation using hot homogenization method. SLN dispersions exhibited average size between 244 and 766 nm. All the dispersions had high entrapment efficiency ranging from 80% to 100%. The MN-SLN dispersion which showed good stability for a period of 1 month was selected. This MN-SLN was characterized for particle size, entrapment efficiency, and X-ray diffraction. The penetration of miconazole nitrate from the gel formulated using selected MN-SLN dispersion as into cadaver skins was evaluated ex-vivo using franz diffusion cell. The results of differential scanning calorimetry (DSC) showed that MN was dispersed in SLN in an amorphous state. The MN-SLN formulations could significantly increase the accumulative uptake of MN in skin over the marketed gel and showed a significantly enhanced skin targeting effect. These results indicate that the studied MN-SLN formulation with skin targeting may be a promising carrier for topical delivery of miconazole nitrate.     

In vivo cutaneous interferon-gamma gene delivery using novel dicationic (gemini) surfactant-plasmid complexes

BACKGROUND: Localized scleroderma (morphea and linear scleroderma) is a connective tissue disease, accompanied by excessive proliferation and deposition of collagen within the skin, inflammation, vasculopathy and a deranged immune system. Interferon gamma (IFNgamma), an inhibitor of collagen synthesis and an immunomodulator, could be a potential therapeutic agent if it could be delivered into or expressed locally in affected skin in a non-invasive manner. In this study, the feasibility of topical delivery of the IFNgamma gene and expression of IFNgamma were investigated in mice. METHODS: Novel dicationic (gemini) surfactant (spacer length n=2-16; alkyl chain m=12 or 16)-DNA complexes were formulated and characterized by circular dichroism and atomic force microscopy to select gemini analogues with the highest transfection efficiency (TE). Transfection and cellular expression of IFNgamma from the bicistronic pGTmCMV.IFN-GFP plasmid were evaluated in PAM 212 keratinocyte culture by ELISA and fluorescence microscopy. Topical delivery of plasmid using liposomal and nanoemulsion systems, based on gemini surfactant 16-3-16, was evaluated in mice by IFNgamma expression analysis. RESULTS: In vitro TE was found to be dependent on the spacer length of the gemini surfactant, with the C3 spacer showing the highest activity (both 12-3-12 and 16-3-16). Both gemini cationic liposomes and gemini nanoemulsion (3x25 microg DNA/animal) produced significantly higher levels of IFNgamma in the skin (359.4 and 607.24 pg/cm2) compared to naked DNA (135.69 pg/cm2) or a liposomal Dc-chol formulation (82.15 pg/cm2). IFNgamma expression in the lymph nodes was higher in the animals treated with gemini liposomes (422.74 pg/animal) compared to the nanoemulsion formulation (131.27 pg/animal) or the Dc-chol formulation (82pg/animal). CONCLUSIONS: The feasibility of topical delivery of pGTmCMV.IFN-GFP plasmid in mice using gemini cationic surfactant based delivery systems was demonstrated. IFNgamma expression after treatment with gemini-DNA formulations in the skin was 3-5-fold higher compared to the treatment with naked DNA (p<0.05), and 4-6-fold higher than the Dc-chol-DNA complex, indicating a significant advance in topical DNA delivery across intact skin in vivo.     

黄曲霉菌原生质体的制备和再生技术优化

黄曲霉菌的遗传转化是研究黄曲霉菌致病相关功能基因的前提和基础,而原生质体是研究和建立真菌遗传转化系统的重要工具。本文分别以黄曲霉孢子和菌丝为材料,研究不同条件下黄曲霉原生质体的形成和再生,结果表明,黄曲霉孢子在酶液浓度为纤维素酶∶蜗牛酶∶溶壁酶=1.5%∶1.5%∶1.5%,30℃酶解3 h,原生质体制备率高达97.3%,再生率达89.2%;黄曲霉菌丝在菌龄为42 h,酶液浓度为纤维素酶∶蜗牛酶∶溶壁酶=1.5%∶1.5%∶1.5%,30℃酶解1 h,可获得最高原生质体产量为2.0×10^6个/m L,再生培养基中以1 mol/L蔗糖作为渗透压稳定剂时,原生质体再生率达5.5%。故本实验条件下,黄曲霉孢子原生质体的形成和再生优于菌丝。     

伏立康唑联合光动力治疗难治性脓癣一例

脓癣是人体皮肤对亲动物性或亲土性皮肤癣菌发生的严重炎症反应,多见于儿童,感染早期常无典型临床表现,因而容易扩散传染。炎症反应明显,可快速发展为隆起性斑块或脓肿,易误诊为单纯细菌感染,延误治疗。目前尚无确切的治疗方案或共识,现报告1例特比奈芬联合伊曲康唑治疗后效果不佳、采用伏立康唑联合光动力治疗后达到完全治愈的儿童难治性脓癣病例。     

Incidence of Aspergillus flavus in roasted and raw groundnuts from supermarkets and other vendors in Pietermaritzburg,South Africa and its management using biological control agents

This study was carried out to investigate the incidence of the aflatoxin-producing fungus, Aspergillus flavus in commercially available roasted and raw groundnuts sold by various vendors. Secondly, the study evaluated the antagonistic activity of Bacillus isolates against A. flavus in vitro and in vivo. All the 42 groundnut samples collected from the various vendors were contaminated with A. flavus. There was no significant difference (p > 0.05) in the incidence of A. flavus contamination in roasted and raw groundnut samples. The results of the in vitro bioassay indicated that the Bacillus isolates inhibited the growth of A. flavus ranging from 61 to 76%. In the in vivo study using groundnut kernels, increase in incubation time and concentration of the Bacillus isolates enhanced control of A. flavus. We showed that Bacillus isolates from the phyllosphere of ryegrass has potential as biocontrol agent for the control of A. flavus in groundnuts.     

Molecular characterisation and biological control of Aspergillus flavus isolates from Saudi Arabia

Abstract

Aspergillus flavus is a phytopathogenic fungus that produces toxic compounds, aflatoxins, in infected plant tissues which harm human and animal health. In this study, 57 food/feed products were collected from 9 locations in Aseer, KSA. A total of 93 isolates were recovered from the samples and were identified as Aspergillus spp. based on their cultural and microscopic characteristics. Six isolates (Af3, Af23, Af24, Af26, Af45 and Af48) were selected and confirmed as A. flavus using polymerase chain reaction (PCR), sequencing of ITS1-5.8s-ITS2 rDNA region and phylogenetic analyses. The six sequences were deposited in the GenBank under the accession numbers of KU561932, KU561934, KU561935, KU561936, KU561937 and KU561938, respectively. Random amplification of polymorphic DNA (RAPD-PCR) of the six isolates using five primers (OPA-2, OPA-3, OPA-9, OPA-11 and OPA-15), produced polymorphic DNA bands of 12, 36, 25, 1 and 1, respectively. The band sizes ranged from 130 to 1600?bp, whereas no monomorphic bands were observed. The bio-control of the six selected A. flavus isolates using three locally isolated yeasts (Candida davisiana, Rhodotorula graminis and Exophiala dermatitidis) was assessed. On solid media, the three yeast strains inhibited all tested A. flavus isolates. The most effective yeast strain was R. graminis. In liquid media, both yeast strains C. davisiana and R. graminis inhibited the dry weights of the six A. flavus isolates. Bio-control approaches of A. flavus could help controlling the pathogen, ultimately, reduce the risk of aflatoxins in human and animal supplies and reduce the use of chemicals that affect the environment and health.     

Synthesis and biological evaluations of oleanolic acid indole derivatives as hyaluronidase inhibitors with enhanced skin permeability

Oleanolic acid (OA) is a natural cosmeceutical compound with various skin beneficial activities including inhibitory effect on hyaluronidase but the anti-hyaluronidase activity and mechanisms of action of its synthetic analogues remain unclear. Herein, a series of OA derivatives were synthesised and evaluated for their inhibitory effects on hyaluronidase. Compared to OA, an induction of fluorinated (6c) and chlorinated (6g) indole moieties led to enhanced anti-hyaluronidase activity (IC₅₀ = 80.3 vs. 9.97 and 9.57 µg/mL, respectively). Furthermore, spectroscopic and computational studies revealed that 6c and 6g can bind to hyaluronidase protein and alter its secondary structure leading to reduced enzyme activity. In addition, OA indole derivatives showed feasible skin permeability in a slightly acidic environment (pH = 6.5) and 6c exerted skin protective effect by reducing cellular reactive oxygen species in human skin keratinocytes. Findings from the current study support that OA indole derivatives are potential cosmeceuticals with anti-hyaluronidase activity.