An in vitro evaluation of Genipin-crosslinked and Hypericum perforatum incorporated novel membranes for skin tissue engineering applications

Incorporating medicinal plant extracts in membranes have a great potential as scaffolds for tissue engineering applications or vehicles for delivering therapeutic agents. Herein, Hypericum perforatum oil (0.25, 0.50, % vol/vol) loaded membranes were developed with Polyvinyl alcohol and chitosan polymer, where Genipin works as a chemical crosslinker to obtain a wound dressing material with acceptable characterization properties. Chemical groups, surface morphology, water uptake capacity, water vapor permeability rate, hydrophilicity, and mechanical properties of membranes were thoroughly investigated. Increasing oil concentration had a significant effect on the water uptake, surface morphology. and water vapor permeability rate of the membranes. Cytocompatibility of the membrane was also investigated with mouse embryonic fibroblasts (MEF) by 3-(4,5-dimethylthiazoyl-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay for direct and indirect cell culture studies. SEM was used to investigate the cell morphology on the membranes. The MTT assay findings prove that Genipin crosslinked H. perforatum oil loaded scaffolds are highly biocompatible and enhance the adhesion and proliferation of MEF cells. In addition to this, the genotoxicity test was performed to show DNA fragmentation. Results showed that the H. perforatum oil loaded polyvinyl alcohol-chitosan membrane presents suitable properties for potential skin tissue engineering applications.     

Development and in vivo response of hydroxyapatite/whitlockite from chicken bones as bone substitute using a chitosan membrane for guided bone regeneration

Biomaterials that meet the requirements to stimulate bone tissue formation play a vital role in orthopedics and dentistry. In this work, chitosan and a biphasic, non-cytotoxic material hydroxyapatite/whitlockite were obtained from natural sources, which are available as organic waste. The osteogenic activity was assessed using a rabbit model animal with a chitosan barrier membrane in combination with a bone-filling graft substitute composed of hydroxyapatite/whitlockite. FT-IR results showed the typical absorption bands of the chitosan and hydroxyapatite. Moreover, the X-ray diffraction pattern revealed a typical hexagonal phase of hydroxyapatite and rhombohedral structures related to whitlockite. Masson's trichrome stain showed an early formation of extracellular matrix mineralized, in accord with the surface morphology of a cortical mature bond observed by Scanning Electron Microscopy. The immunocytochemistry results showed a significant increase of positive immunoreactive cells to osteonectin in the treated defects in comparison with the control defects 6 and 8 weeks postoperatively. Overall, the results confirm that the use of this low-cost and versatile biomaterial as a barrier membrane and a bone substitute graft are useful for bone tissue engineering.     

Formulation of sage essential oil (Salvia officinalis,L.) monoterpenes into chitosan hydrogels and permeation study with GC-MS analysis

This study deals with the formulation of natural drugs into hydrogels. For the first time, compounds from the sage essential oil were formulated into chitosan hydrogels. A sample preparation procedure for hydrophobic volatile analytes present in a hydrophilic water matrix along with an analytical method based on the gas chromatography coupled with the mass spectrometry (GC-MS) was developed and applied for the evaluation of the identity and quantity of essential oil components in the hydrogels and saline samples. The experimental results revealed that the chitosan hydrogels are suitable for the formulation of sage essential oil. The monoterpene release can be effectively controlled by both chitosan and caffeine concentration in the hydrogels. Permeation experiment, based on a hydrogel with the optimized composition [3.5% (w/w) sage essential oil, 2.0% (w/w) caffeine, 2.5% (w/w) chitosan and 0.1% (w/w) Tween-80] in donor compartment, saline solution in acceptor compartment, and semi-permeable cellophane membrane, demonstrated the useful permeation selectivity. Here, (according to lipophilicity) an enhanced permeation of the bicyclic monoterpenes with antiflogistic and antiseptic properties (eucalyptol, camphor and borneol) and, at the same time, suppressed permeation of toxic thujone (not exceeding its permitted applicable concentration) was observed. These properties highlight the pharmaceutical importance of the developed chitosan hydrogel formulating sage essential oil in the dermal applications.     

Synthesis,characterization, and antimicrobial activity of chitosan–zinc oxide/polyaniline composites

Organic–inorganic materials of chitosan–zinc oxide/polyaniline (CS–ZnO/PANI) composite were prepared via precipitation with a polymerization method and characterized by FT-IR, XRD, EDXS and TEM analysis, thereby providing evidence of composite formation. The size of the prepared CS–ZnO/PANI composite was found to be 100–200 nm, thereby rendering the morphology suitable for biomedical applications. Antibacterial activities of chitosan–ZnO (CS–ZnO), polyaniline (PANI) and CS–ZnO/PANI composites were determined against Gram-positive bacterium, Staphylococcus aureus (S. aureus), and Gram-negative bacterium, Pseudomonas aeruginosa (P. aeruginosa) and were tested in-vitro at 5–50 μg/mL. Results showed that CS–ZnO/PANI composite had broad-spectrum antibacterial activity that was greatly enhanced in comparison with CS–ZnO. In addition, CS–ZnO/PANI composite has tested fungal strains of Candida albicans (C. albicans) and relatively higher activities were observed than the known antibiotics. Finally, the antimicrobial activity of CS–ZnO/PANI composite against established biofilms was also examined and resulted in more than 95% inhibition in biofilm formation.     

Combination of heat treatment and chitosan coating to improve postharvest quality of wolfberry (Lycium barbarum)

The synergistic application of hot water dip at 42 °C for 30 min and 1% chitosan coating on differentiation in postharvest quality traits, microstructure as well as microbiological evolution of wolfberry fruits was investigated. Fresh wolfberry fruits were stored at 2 ± 0.5 °C and 90% relative humidity (RH) for 28 days. Results indicated the combination of prestorage heat treatment and chitosan coating maintained higher levels of ascorbic acid, total phenolic contents and antioxidant capacity as well as lower decay, compared with untreated wolfberry fruits. The possible mechanism was that the heat treatment almost sealed open stomata to limit the sites of pathogen penetration into fruits independently, followed by the biofilm formed by chitosan which controlled secondary infections as well as slowed changes in fruit respiration and metabolic activity in wolfberries. The synergistically treated fruit also exhibited a higher acceptability obtained by sensory analysis after cold storage. In this sense, the integrated application of heat treatment and chitosan coating could be regarded as an effective strategy to extend storage life and maintain the postharvest quality of wolfberry fruits.     

Controlled release of fluorouracil (5-FU) from chitosan-co-poly(ethylene glycol)/ poly(glycerol sebacate)-co-poly(ethylene glycol)-coated iron oxide

A blend of polyglycerol sebacate-poly ethylene glygol/chitosan-poly ethylene glycol-coated iron oxide (PGS-PEG/CS-PEG@Fe₃O₄) nanoparticles for 5FU delivery was prepared by reverse ultrasonic emulsification method. To enhance polymers’ solubility, PEG was grafted. Chemical characterization was performed through Fourier transformed infrared and proton nuclear magnetic resonance spectra. In vitro assay revealed that release profile of 5FU-loaded PGS-PEG/CS-PEG@Fe₃O₄ is sustained. Moreover, cytotoxicity was analyzed on HT29 cell line at the presence of external magnetic field using the lactate dehydrogenase and Alamar Blue. Results illustrate that (PGS-PEG/CS-PEG@Fe₃O₄) is promising to use as a carrier for 5FU anticancer agent with sustained tailored release.     

Precise quantification of the antibacterial activity of chitosan by NB medium neutralizer

In the present research,nutrient broth(NB)medium was identified to be able to neutralize the antibac-terial activity of chitosan and its derivatives.Therefore,an improved test method independent of NB medium was proposed to precisely quantify the antibacterial effectiveness and efficiency of chitosan.The minimum bactericidal concentration(MBC)of chitosan was 60 μg mL-1 against S.aureus and E.coli,and 0.01％(w/v)chitosan could kill 100％of bacteria within 3 min.From another point of view,the neutralizing efficiency of NB could be tripled by adding 25 g L-1 of sodium chloride.Then the neutraliz-ing mechanism of NB medium was ascribed to flocculation between chitosan and protein.Adding extra sodium chloride could significantly reduce the size of floccules,and smaller floccules would lose the ability of binding with bacteria directly,showing higher neutralizing rate on the macro scale.     

Chitosan coatings enriched with active shrimp waste for shrimp preservation

An active coating solution composed of chitosan (Ch) and a shrimp protein-lipid concentrate (PCc), both obtained from Litopenaeus vannamei processing wastes, was applied to preserve shrimp during chilled storage. The addition of PCc increased the antioxidant capacity of the Ch coating, yielding a lower-viscosity mixture which, however, was viscous enough to adhere to the shrimp while maintaining its activity. The shrimp storage trial showed that the Ch coatings, especially when enriched with PCc (Ch-PCc), delayed microbial growth, mainly by extending the lag phase. Pseudomonas spp. appeared to be the predominant bacteria in the microbiota. H₂S-producing organisms and luminescent colonies were especially sensitive to this active coating, with inhibition greater than 4 and 2 log cycle respectively, while the growth of lactic acid bacteria was not favored. The Ch-PCc coating delayed the onset of melanosis and did not confer any sensorially detectable color, taste or odor. It therefore shows promise as a means to improve the quality of shrimp during cold storage.     

Removal of chlorophenols from groundwater by chitosan sorption

The equilibrium and kinetics of chlorophenol (CP) sorption by chitosan, poly D-glucosamine, were studied under simulated groundwater conditions. Lower temperature, from 25 degrees C to 15 degrees C and then 5 degrees C, markedly decreased the adsorption rates by a factor of 30-53% and 7-22%. Comparison between two types of chitosan, flakes and highly swollen beads, demonstrated that the maximum pentachlorophenol (PCP) uptake capacities in Langmuir and Freundlich models depend on the specific surface area of the particle. Low temperature (5 degrees C) significantly increased the PCP uptake capacity in comparison to higher temperatures (15 degrees C and 25 degrees C). PCP uptake capacity was halved at pH levels higher than 6.5, and NaCl concentrations greater than 1% blocked PCP sorption almost completely. Of five kinds of chlorophenols, i.e. 2,4,6-trichlorophenol (2,4,6-TCP), 3,4-dichlorophenol (3,4-DCP), 2,3-dichlorophenol (2,3-DCP), 2,6-dichlorophenol (2,6-DCP), 3-monochlorophenol (3-MCP), TCP had the maximum sorption efficiency on flake-type chitosan, followed by DCPs, and finally MCP (the three kinds of DCP, with the same elemental compositions, achieved similar sorption performances).