Antioxidant, antibacterial/antifungal nanostructures for medical and food packaging applications
2 P. Poni Institute of Macromolecular Chemistry, Romanian Academy, 41A Grigore GhicaVodă Alley, 700487 Iasi, Romania
Received Date: Oct 30, 2017 / Accepted Date: Nov 15, 2017 / Published Date: Nov 20, 2017
Citation: Munteanu BS, Vasile C. Antioxidant, antibacterial/antifungal nanostructures for medical and food packaging applications J Nanosci Nanomed. November-2017;1(1):15-20.
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Electrospinning is a very attractive fibers fabrication technique due to the ability to produce nanoscale materials and structures with outstanding properties. As drug delivery systems it offers nanofiber meshes with high surface/volume ratio, high porosity and high surface exposed to the release media. The biologically active substances/drugs can be encapsulated into the individual polymeric nanofibers by coaxial or monoaxial electrospinning or, by another approach by which the biologically active substances can be entrapped and attached as nanoparticles to the nanofiber mesh. As antimicrobial/antioxidant materials for biomedical applications, the electrospun formulations containing silver nanoparticles are presented. As coating materials for food packaging, electrospun nanofibers containing chitosan formulations with antibacterial/antioxidant/antifungal properties are discussed.
Electrospinning; drug delivery; coating; antibacterial; antioxidant; antifungal.
Nowadays, nanomaterials and nanoparticles are used for many applications with benefits for our current life [1-3]. One method by which nanostructures can be obtained is electrospinning, a very efficient fibers fabrication technique due to the ability to produce nanoscale materials and structures [4,5] with high porosity and high specific surface area . Also, the three-dimensional nanofibrous network allows the electrospun fibers to resemble native extracellular matrices . By choosing the base polymer they can be easily designed to have enhanced mechanical properties, biocompatibility, and cellular response, making them a good choice to be used in nanocomposites materials applicable in the medical field . The biomedical field is one of the most important application areas that utilizes the technique of electrospinning such as: tissue engineering , growth factors [10,11], cardiovascular tissue engineering , bone tissue regeneration , drug release systems , wound healing, etc. . Electrospinning can produce a macroporous scaffold comprising randomly oriented or aligned nanofibers which may incorporate a drug delivery function into the fibrous scaffold. Such electrospun nanofibrous scaffolds may provide also an optimal microenvironment for the seeded cells .
The basic electrospinning set-up is mainly comprised from four main parts: a syringe containing a polymer solution, metallic needle, power supply source, and metallic collector (with a variable design) (Figure 1a) . The high direct voltage (0 to 30 kV) is applied between the metallic collector and the syringe needle. The polymer solution is extruded through the needle tip and at the point of ejection from the needle, a polymer jet is created and extended as a result of the electric charge repulsion outrunning the solution surface tension . The polymeric solution jet flows toward the metallic collector with simultaneous evaporation of the solvent, followed by the deposition of a mat of nanofibers on the collector surface. When nanoparticles are deposited instead of nanofibres the process is usually called electrospraying .
In this review, recent drug delivery nanostructures based on electrospun nanofibers (such as eletrospun meshes containig sulfadiazine modified chitosan nanoparticles) are presented together with recent applications of electrospun formulations containing silver nanoparticles as antimicrobial/ antioxidant materials for biomedical applications. Also, recent electrospun coatings based on chitosan fomulations for antibacterial/antioxidant/ antifungal food packagings are presented.
Drug releasing systems
Compared with other dosage forms, several advantages of the use of the electrospun polymer nanofibers have been recognized. Therapeutic compounds such as lipophilic and hydrophilic drugs, proteins or antimicrobial agents  can be incorporated into the nanocarrier polymers using monoaxial or coaxial electrospinning. Electrospun scaffolds have gained an exponentially increasing popularity because of their ultrathin fiber diameter, high surface-volume ratio, high porosity and high surface exposed to the release media . Thus, the electrospun/ electrosprayed nanoporous structures provide very short diffusion length [24,25] and more rapid substance transfer  for drug release in comparison with drug-loaded films or capsules. The drug release profile can be tailored by controlling the morphology and the porosity of the nanofibers and also the composition of the fibers . Additionally, electrospun nanofibers can be coated  onto various substrates and medical devices .
Nanofibers offer an option for the treatment of skin damages as tissueengineered skin substitutes which can help skin reconstruction . In this case, the drug enclosed into the nanofibers mesh will be released by different mechanisms when the nanofibrous mesh is swollen, biodegraded  and/or absorbed by the human body. An effective wound dressing system will give a large initial burst release of the drug  which is important to stop the growth of the bacteria especially in the early stage of the wound healing process . The burst must be followed by a long term release at inhibitory level . The continued low release rate should keep the wound free from infections for days or weeks .
According to literature data, the biologically active substances/drugs can be encapsulated into the individual polymeric nanofibers by (a) coaxially  or (b) monoaxially  electrospinning of the active substance and the polymer . In another approach the biologically active substances can be (c) entrapped and attached as nanoparticles to the nanofiber mesh.
(a) Coaxial electrospinning – (Figure 1b) can be used to encapsulate drugs or biologically active substances inside the individual polymer nanofibers . In a common process, two (or more) polymer solutions are electrospun through different coaxial capillary channels (needles) (Figure 1b), resulting in a core–shell-structured composite nanofiber. The shell polymer, after the electrospinning, acts as a barrier to control the release of the loaded molecules . If the shell fluid is able to be processed by electrospinning, the core fluid can either be or not be electrospinnable. An advantage of this method is the possibility to enclose almost any drugs (especially hydrophobic ones) in the core regardless of drug–polymer interactions. Hence, drugs , proteins  and growth factors  and even genes  can be incorporated into nanofibers simply by dissolving them in the core solutions. A drawback of the coaxial electrospinning comes from the differences in the physical properties of the core and shell solutions conductivities and viscosities of the two solutions.
(b) Monoaxial electrospinning – (Figure 1a) simply encapsulates the biologically active substances/drugs within the individual nanofibers by dispersing/mixing them into the polymer solution. The obtained mixture is further electrospun through a single needle system . Using this method, electrospun nanofibers of chitosan/polyethylene oxide  and chitosan/polyurethane  were obtained, containing silver sulfadiazine with good antibacterial activity against both Gram-negative and Grampositive bacteria. In acidic medium the release of silver sulfadiazine from chitosan beads is governed by chitosan erosion  or even disintegration . The release of silver sulfadiazine from chitosan/chondroitin sulfate films at neutral pH occurred by a sustained release (over a period of days) .
Dimensions of the electrospun fibres are comparable with those of natural collagen and vary with type of electrospun material and parameters of electrospinning – Table 1 .
|Sample||Average nanofibres diameter (nm)||Reference|
|Sulfadiazine modified chitosan inner needle||32 ± 10 nm|||
|Sulfadiazine modified chitosan outer needle||30 ± 10 nm|||
|Sulfadiazine modified chitosan/chitosan mixture uniaxial||35 ± 10 nm|||
|PLA/Vitamin E/silver||140 ± 60 nm|||
|PCL/ THF:DMF (1:1)||500-900 nm|||
|PU /DMF, 3.8-12.8wt%||∼60-800nm|
|pHEMA/Monomer||315 ± 140nm|
|Vancomycin-Loaded Electrospun Rana chensinensis Skin Collagen/Poly(L-lactide) Nanofibers||500 ± 800 nm|||
|Poly(lactic acid-co-glycolic acid) (PLGA)||1000-1800 nm|||
|Core/sheath structured composite nanofibers with a core of blended salicylic acid (SA) and poly(ethylene glycol) (PEG) and a sheath of poly(lactic acid) PLA)||300-3000 nm|||
(c) Besides encapsulating the biologically active substances/drugs into the individual polymeric nanofibers by coaxially or monoaxially electrospinning, there is another approach by which the active substances can be entrapped and attached as nanoparticles to the polymeric nanofiber mesh. This can be achieved by simultaneous electrospinning the polymer and electrospraying the active substance [49,50], spraying the active substance into the electrospinning jet so that the particles containing the active substance are attached to the fiber surface prior to deposition on the mat , electrospraying the suspension containing the active substance nanoparticles onto the previously electrospun nanofiber scaffold . By monoaxial electrospinning of the mixture of polymer and active substance suspension through a single nozzle  or by coaxial electrospinning with the nanofiber polymer solution flowing through the central nozzle and of the colloidal suspension of the active substance through the outer nozzle which generates a nanofibers mesh covered with active nanoparticles, codeposited from colloidal suspension during the process of electrospinning  can be obtained.
An advantage of the nanostructures with active substances entrapped and attached as nanoparticles to a polymeric nanofiber mesh is the improved availability of the active substances to the targeted medium, in comparison with the corresponding systems with fibers containing inside the active nanoparticles [54,55]. As was previously shown, the required burst release can be easily achieved when a major part of the active nanoparticles entrapped into the nanofibrous mesh is exposed at the fiber surface .
It is known that the chitosan enhances the wound healing  favoring fibroblast attachment  and re-epithelialization  of the wound. Thus, a chitosan nanofiber mesh is a good candidate for wound healing systems also due to chitosan biocompatibility, antibacterial, and antifungal  activities. Considering these excellent properties of the chitosan, high molecular weight chitosan nanofibrous structures having attached active nanoparticles of sulfadiazine (a well-known antibacterial agent  used in the treatment of wound infections ) or sulfadiazine modified chitosan (which was found to have enhanced antibacterial properties [63-65]) were obtained  by mono-axial and coaxial electrospinning. The sulfadiazine or sulfadiazine modified chitosan nanoparticles loosely attached at the surface of the nanofibers, could provide a burst release in the first 20 min (in phosphate buffer solution of pH 6 at 37°C ) which is important to stop the possible initial infection in a wound, while the sulfadiazine or sulfadiazine modified chitosan from the nanoparticles which are better stuck (or even encapsulated) into the chitosan nanofibers were slowly released with releasing mechanism governed by the erosion/ disruption of the chitosan nanofiber mesh. Thus, the fiber forming high molecular weight chitosan  assured the formation of the nanofibrous mesh while the sulfadiazine or sulfadiazine modified chitosan both in the form of a relatively stable suspension assured the formation of the active nanoparticles attached to the chitosan nanofiber mesh.
Electrospun formulation containing silver nanoparticles as antimicrobial coatings for biomedical applications
Silver nanoparticles (AgNPs) can be used as a broad-spectrum antibacterial agent for both Gram positive and Gram negative bacteria in biomedical and food packaging applications. Because of their high reactivity originating from the large surface to volume ratio, the AgNPs can effectively eliminate bacteria and yeasts even at rather low concentrations . Furthermore, antibacterial activity of silver nanoparticles was found to be dependent on the size of silver particles, since the only nanoparticles that present a direct interaction with the bacteria preferentially have a diameter of approximately 1-10 nm . The smaller particle size provides improved antibacterial activity .
Antibacterial property of electrospun nanofibers containing AgNPs was reported by numerous studies such as polylactic acid/AgNPs fibers against Staphylococcus aureus and Escherichia coli , poly(ethylene oxide)/ AgNPs fibers intermixed with polyurethane fibers against Escherichia coli , polyacrylonitrile/ AgNPs fibers against Gram positive Bacillus cereus and Gram negative Escherichia coli micro-organisms , and Nylon-6/AgNPs nanofibers against both Gram negative Escherichia coli and Gram positive Staphylococcus aureus .
Antioxidant activity of vitamin E, a fat soluble antioxidant  was combined with antibacterial property of AgNPs in electrospun polylactic nanofibers in order to obtain multifunctional biomaterials. The polylactic acid/ AgNPs /vitamin E nanofibers inhibited growth of Escherichia coli, Listeria monocytogenes and Salmonella typhymurium up to 100%. The release rate of silver ions from the nanofibers immersed in aqueous solution was kept approximately constant even after 10 days of immersion. The polylactic acid/ AgNPs /vitamin E nanofibers had antioxidant activity and the results of the tests on fresh apple and apple juice indicated that the polylactic acid/ AgNPs /vitamin E nanofiber membrane actively reduced the polyphenol oxidase activity. These materials could find application in food industry as a potential preservative packaging for fruits and juices .
As was previously shown due to their intrinsic flexibilty, the electrospun nanofibers can be coated onto various medical devices  which require the flexibility of the coated layer. Bioactive formulations containing polyurethane and small amounts of biocompatible polymers (hydrolyzed collagen, elastin, hyaluronic acid or chondroitin sulfate, and silver nanoparticles) were coated by electrospinning onto pure polyurethane membrane in order to study the possibility of improving the antibacterial properties of the polyurethane urinary catheters. The obtained coated polyurethane membranes had good antimicrobial activity against Escherichia coli, Salmonella typhymurium, and Listeria monocytogenes . The same authors have shown that the AgNPs improved the electrospinability of the polyurethane bioactive formulations. At low content of AgNPs (less than 0.3%) the coated formulations had high cell proliferation and good biocompatibility having the advantage of adding low amounts of bioactive and biocidal components .
Antibacterial/antioxidant/antifungal electrospun coatings for food packaging
Efforts are being made to increase the storage and shell life of food by using active antimicrobial packagings. The antibacterial agents may be coated onto the packaging material [77,78]. Due to its known antibacterial activity chitosan was also used to obtain antibacterial coatings  or used to encapsulate/incorporate another antibacterial agent  in order to improve the antimicrobial activity of the coating . Various procedures have been proposed for coating the antimicrobial agent: spraying (nebulisation) , lamination , immersion, etc. [83,84].
One way to perform the coating of the active agent is the electrospinning method due to several advantages: besides the high specific surface area, the very thin thickness of the coated (deposited) layer which can be easily control by the changing of the deposition time or of the flow rate with the possibility to obtain very thin coatings which in some cases are enough to obtain the desired antibacterial effect [27,85].
Plasticizers are added to the coatings to overcome the brittleness exhibited during packaging formation  and to improve the flexibility and processability. It is known that polymer nanofibers and have higher yield strength and especially higher ductility than the corresponding bulk material [86,87] due to low nanofiber crystallinity resulting from rapid solidification of the ultrafine electrospun jets [88,87]. Thus it is expected that the nanofibrous electrospun coating layers will have the needed flexibility during the package formation/life time without the plasticizer addition.
Polyethylene films chitosan-coated by electrospinning had good antimicrobial activity against food pathogen microorganisms as Grampositive (Listeria monocytogenes) or Gram-negative (Escherichia coli, Salmonella) . The addition of vitamin E to the coatings improved the aspect, smell, pH, reaction with H2S and total number of germs for minced poultry meat packaging . Polylactic acid films coated by electrospnning with formulation containing chitosan had excellent antifungal activities against Aspergillus brasiliensis, Fusarium graminearum, Penicillium corylophilum .
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