2 Advanced Electrospinning Setups and Special Fibre and ... · PDF fileapparatus is basically...

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Claudio Migliaresi, Giuseppe Alberto Ruffo, Fabio Zomer Volpato and Dario Zeni

2.1 Introduction

Electrospinning is a powerful technique for the production of fibres with small diameters, in the range from micrometres down to tens of nanometres. The traditional apparatus is basically composed of a needle fed by a polymer melt or solution and that is connected to a high voltage generator, and by a grounded target fixed at a certain distance from the needle. Usually, the needle and the target are positioned vertically with the needle set above the target. In the former configuration some problems can be caused by the gravitational force, which allows drops to fall from the needle causing defects on the mat. This problem can be avoided either by placing the target above the needle or changing the configuration to a horizontal disposition.

By changing the process parameters, such as polymeric solution composition, needle-target distance and applied voltage, a non-woven network composed of randomly oriented fibres of controlled diameter is obtained. However, the technique presents some limitations. One of the main problems is the limited throughput, intrinsically derived from the production of a single nanofibre. A technical issue, on the other hand, is the clogging of the needle, causing the interruption of the process. Both these problems are particularly relevant in industrial applications, where a continuous and fast process is needed.

Numerous solutions have been studied for electrospinning, combining solvents and polymers with different characteristics, and new apparatuses have been developed, modifying either the spinneret/feeding system or the target, in order to improve process, mesh morphology or fibre properties. Mats can be thus modified and adapted to the specific purpose they are intended for. Moreover, also the features of the single fibres can be tailored to obtain the best performance of the network produced.

2 Advanced Electrospinning Setups and Special Fibre and Mesh Morphologies

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Electrospinning for Advanced Biomedical Applications and Therapies

Typical electrospinning mats are composed of solid fibres, with a narrow distribution of diameter, with a smooth surface and a circular cross-section. However, setup parameters can be adjusted to different fibre morphologies, such as porous, hollow, ribbon-like or wrinkled cross-section, multi-modal diameter distribution, as well as in contemporaneous with beads, so producing mats with characteristics that could be exploited for specific biomedical uses. In the next sections, these special electrospinning setups are described.

2.2 Advanced Electrospinning Setups

The basic electrospinning apparatus, as described previously, is probably the most widely used setup due to its simplicity and popularity. Despite this fact, it is worth emphasising that this configuration presents various limitations that several research groups tried to overcome through the modification of the apparatus design and adding further elements to the basic setup. The present section will describe modified electrospinning ‘architectures’ used to produce fibres with more complex morphologies and ‘non-conventional’ mats.

2.2.1 Enhanced Electrospinning Throughput

One of the main issues to overcome is the limited throughput obtainable by the use of a single needle. For industrial applications in the biotechnological field, despite the commonly high value-added of products, the low production rate (in the order of tenths of grams per hour) of the basic electrospinning apparatus represents a severe restraint [1].

The basic idea to overcome these limits is to multiply the number of Taylor cones from which fibre jets are emitted and finally collected on a target. As will be discussed in the following section, the devised apparatuses cover a really wide spectrum of technical solutions and, in some cases, a marked example of ingenuity.

2.2.1.1 Multi-needle Electrospinning

One of the most logic developments, starting from the basic design, has been the increase of the number of syringes (multi-needle electrospinning) thus consistently used to enhance the throughput. This design, already conceived by Formhals in 1934, has been more recently re-adopted as a possible solution to the problem [2]. Although an increase in the fibre deposition rate has been reported, it is important to stress that to reach a throughput comparable to other industrial spinning techniques

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Advanced Electrospinning Setups and Special Fibre and Mesh Morphologies

it should be necessary to employ a large number of syringes or spinning heads. Furthermore, this solution would lead to a dramatic increase in the probability to encounter clogging problems of syringes [3]. Another disadvantage is related to the electric field interferences generated by the numerous spinning heads: in fact, several problems of non-uniform fibre deposition have been reported due to the mutual repulsion generated by the Coulombic forces of the ejected thread [4, 5]. Although this drawback has been minimised by modifying the setup design [6] and using a collector with an opposite charge (for the syringes) instead of the traditional target connected to ground, the solution still presents many complexities for industrial utilisation [7].

2.2.1.2 Needleless Electrospinning with Confined Feed

In order to avoid the use of capillaries and needles and their inherently related limits, several authors have devised new forms of spinnerets that are usually called ‘needleless’ electrospinning. These can be broadly divided into two general groups: needleless electrospinning with a ‘confined feeding system’ and with an ‘unconfined feeding system’. To the first group belong setups that have a reservoir for the polymer solution that is subsequently injected into an enclosed nozzle or spinning head (when the nozzle is not the reservoir itself). In the unconfined feed system, the Taylor cones are formed instead, on a free liquid surface and no nozzles are needed.

One of the first approaches developed has been to employ a flat spinneret as an alternative to syringes. In this case the spinning head, which is made of either a metallic [3, 8] or an insulating [9] material, is a disk with several nozzles disposed in a regular pattern on the lower circular surface facing the collector. Authors reported an increase in the polymer throughput compared to single needle solution and an improvement in the fibre deposition in contrast to multi-needle electrospinning, resulting from the reduced mutual fibre repulsion and of the generated electric field between the spinneret and the collector, as indicated by the finite element method analysis.

A different spinneret design consisted of a polyethylene tube with a porous surface acting as a nozzle with hundreds of micrometric holes [10] (Figure 2.1). To maximise the ‘working’ surface, authors used the lateral surface of the cylinder as a spinning head while the collector, a wire mesh connected to ground, has been consequently disposed as a coaxial cage to the polymeric nozzle. The spinning tube was filled with the polymer solution that was forced through the pores of the nozzle by means of pressured air; at the same time the electric potential was applied inserting an electrode in the polymer solution. The parametric study carried out with different polymer concentrations showed that the frequency distribution of fibre diameters obtained with this setup compared to the single needle electrospinning had a broader dispersion but a similar mean value. Although this system allowed a great enhancement of the

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production rate (about 250 times), it was not specified if was subjected to clogging and how to obviate the problem.

Figure 2.1 Apparatus scheme (a) and porous structure (b) of the nozzle-tube surface viewed at the scanning electron microscope (SEM). Reproduced with

permission from O.O. Dosunmu, G.G. Chase, W. Kataphinan and D.H. Reneker, Nanotechnology, 2006, 17, 1123. ©2006, IOP Publishing Ltd [10]

In order to overcome some limitations of the basic electrospinning apparatus, researchers have tried to maximise the efficiency of the electric field and also to couple it with other forms of energy to increase the productivity or to ameliorate fibre features. This is the example of the so-called rotary jet-spinning (Figure 2.2), where the electrostatic forces are supplemented with centrifugal forces that enable a reduction in the electric field necessary to induce the jet formation, a better fibre morphology and an increase in the throughput [11]. The apparatus consisted of a motor with controllable rotation speed that, through a shaft, put in motion a solution reservoir provided with two sidewall orifices. By regulating the speed of revolution, the orifice diameter and the other basic parameters of the electrospinning process (polymer solution characteristics, electric field and reservoir-collector distance), authors were able to control morphology and fibre diameter of the different polymers employed. In particular, by increasing the rotational speed a reduction of beads was observed, also with emulsions and suspensions that usually have a scarce spinnability. Furthermore the centrifugal forces, apart from enhancing the fibre stretching, allowed the tailoring of the fibre alignment, a characteristic that authors demonstrated to be useful for cell alignment in specific tissue engineering applications.

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Figure 2.2 Rotary-jet spinning scheme (a) and its functioning principles (b). Reproduced with permission from M.R. Badrossama, H.A. McIlwee, J.A. Goss

and K.K Parker, Nano Letters, 2010, 10, 2257. ©2010, American Chemical Society [11]

A further alternative solution to the use of syringes has been devised which utilises as a spinneret a multichannel microfluidic apparatus [12]. The main advantages of this approach lie in the simple prototyping and ease of fabrication of the component thanks to polydimethyl siloxane-based micromoulding technology. Practically, the polymer solution is injected at one end of the microfluidic device (where it is charged by an electrode) and then divides through the various arrays of channels until it is ejected toward the collector at the other extremity of the spinning head. In the literature that deals with this kind of device, however, no study on the possible interaction among the jets has been carried out, nor was there mention of any problem related to non-uniform fibre deposition. It is therefore difficult to assess now whether the technique has promising perspective or not and consequently further studies are needed.

2.2.1.3 Needleless Electrospinning with Unconfined Feed

The previously mentioned examples of setups, modified to increase the electrospinning throughput, present a large number of small holes that act as nozzles. These, as well as syringes, can undergo clogging and thus working conditions have to be carefully controlled during the whole process. To avoid the complexity of this kind of apparatuses, a completely different set of configurations had been conceived. When, several years ago [13], it has been discovered that it was possible to electrospin a polymeric solution from a free liquid surface, many research groups have undertaken this approach to implement the technique. As mentioned before, this kind of apparatus design can be classified as needleless electrospinning with ‘unconfined feed system’.

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As stated, the breakthrough in this direction has been made following the observation that jets are emitted even from free or nearly flat liquid surfaces [13]. In fact, the current theory of electrohydrodynamics implies that some form of concentration of the electric charges has to take place in order to overcome the surface tension and generate the spinning jet. In the case of a flat conductive solution it has been demonstrated that a perturbation on the free liquid surface is sufficient to create the critical field strength concentration that induces the jet formation from the wave crests formed [14]. The observation of such phenomenon has prompted the arrangement of new experiments on the self-organisation and the inter-jet distance of multi-jet electrospinning from near one-dimensional free surface of polymer solution (Figure 2.3) [14]. Furthermore, the equations drawn from this study seem to generalise the theory for capillary electrospinning firstly laid by Taylor. Although under strongly simplified assumptions, with few parameters and dimensionless analysis authors were able to give a comprehensive description of the electrospinning jetting based on the classic dispersion law. The concepts extended to a wider scientific panorama (especially wave interaction on two-dimensional liquid surface, spinning solution thickness and viscosity) might represent a powerful tool for future developments of the technique.

Figure 2.3 Setup scheme for the electrospinning experiments from a free liquid surface (a) [where (2) is the solution container, (3) high voltage source, (4)

grounded collector, (6) metallic plates that replace the needles and (7) polymeric jets], and self-organisation of the inter-jet distance as function of the electric field (b and c). Reproduced with permission from D. Lukas, A. Sarkar and P. Pokorny,

Journal of Applied Physics, 2008, 103, 084309. ©2008, American Institute of Physics [14]

Another typical example of needleless setup from a free liquid surface was developed by a bottom-up design where the spinning head was made of a thin layer of polymer solution over a ferromagnetic fluid. Once the latter had been excited with a magnetic

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field, many vertical spikes were generated, and under a normal electric field these tips, perturbing the free surface of the polymer solution, acted as critical sites from which a corresponding number of jets were ejected [15]. The throughput was quantified as 12 times higher than a comparable multi-needle system thanks to the relatively high number of jets per area (26 cm-2). Although the device configuration is quite complex, so limiting its practical use, this needleless electrospinning showed how a high production rate could be achieved avoiding the drawbacks implicit in multi-needle systems.

Apparatuses implementing a plate edge as electric field concentrator for the spinning appeared to be more feasible for practical uses. In this kind of horizontal setup, the polymeric solution is dripped from a reservoir directly onto a metal plate that act as the spinneret and that is tilted at a certain angle in respect to the perpendicular of the collector to facilitate the solution flow [16] (Figure 2.4a). In this way, the polymer solution, slowly flowing by gravity, reaches the edge of the plate where it forms the Taylor cone due to the local electric field concentration generated by the metallic edge. The advantages of this configuration lie in its intrinsic simplicity with many jets starting from the plate edge without the possibility to incur in the clog of the spinning head. However, with this setup, the spinning solution flow parameters strongly affect the quality of the collected fibres.

Figure 2.4 Scheme of the single plate (a) and multi-plate (b) electrospinning. Reproduced with permission from N.M. Thoppey, J.R. Bochinski, L.I. Clarke and

R.E. Gorga, Polymer, 2010, 51, 4928. ©2010, Elsevier [16]

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In order to further increase the jet number and hence the throughput, a multi-plate configuration has also been tested (Figure 2.4b). The new system was called ‘waterfall geometry’ since each plate, apart from being a spinneret acted also as reservoir for the plate below by dripping the polymer solution from one plate to the next. Indeed, authors noticed that, due to the larger flow rate needed to supply the many plates, the resulting spinning process was intermittent and, more importantly, the production yield was lower than that for a single plate.

The concept of a polymeric solution dripping from a reservoir to the spinning head was also at the base of a recently proposed design [17]. In this case however, the solution was electrospun from a horizontal, slowly rotating cylinder and not from a fixed plate. The idea of a rotating cylinder was conceived to avoid the solution from dropping away and consequently to lower the waste of material. Adjusting the various parameters (cylinder rotational speed, applied voltage, gap distance and solution parameters), the fibre fabrication rate was enhanced by 24-45 times, if compared to single needle electrospinning. Nevertheless, the introduction of a rotating device seems a worthless complication in contrast to the simplicity of the previously described setup.

An alternative design, with a horizontal rotating cylinder partially immersed in the polymer solution (exploiting a different operational process) led to probably one of the most successful designs in the recent past for electrospinning setups (Figure 2.5a). In proximity of the revolution apex, the electric field between the cylinder and a collector above was sufficiently high to generate many jets from the thin layer of solution remaining on the cylinder surface (Figure 2.5b). Since its invention [13], this setup has demonstrated to have great potentiality for the spinning of various polymers [18, 19] and in the scale up for mass fibre production at industrial level. At the present, it remains one of the few electrospinning apparatus that has been commercialised on a relatively large scale, with the brand name of ‘nanospider’.

Another design having some similarities to the ‘nanospider’, used as a spinneret, was a metallic rotating cone disposed vertically over the collector [1]. The feeding system simply consisted of a reservoir from which the polymer solution was dripped directly over the lateral surface of the cone. Once the lower edge of the cone was reached, the thin layer of solution started to divide and to form a large number of jets. With this system, however, the gravitational and mostly centrifugal forces deriving from the cone rotation had a double action on the fluid: in fact, they helped to spread the solution and also favoured the spinning so much that authors reported the possibility of spinning nanometric fibres by just increasing the rotational speed of the cone after turning off the electrical field. This means that the speed of revolution could be used as a further parameter to control the fibre morphology and size or to facilitate the spinning of polymeric solution, which would otherwise be difficult to obtain. The rate at which the nanofibres could be electrospun was in the order of the grams per

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minute (the maximum reported is 12.5 g/min for a cone with a base diameter of 40 mm). Interestingly, within the reported range, the fibre diameter had a low sensitivity toward throughput variation and actually a decrease in the fibre size distribution for higher production rate was observed.

Figure 2.5 (a) ‘Nanospider’ scheme [polypropylene (PP) is the supporting material] and (b) an image of the working system. Reproduced with permission from O.

Jirsak, P. Sysel, F. Sanetrnik, J. Hruza and J. Chaloupek, Journal of Nanomaterials, 2010, 2010, 1. ©2010, Hindawi Publishing Corporation [19]

Recently, a type of electrospinning setup has been reported in the literature, which has some features of both the unconfined and the confined needleless feeding system. Made primarily of a conical wire coil used as spinning head [20], it can be considered a confined setup since the polymer solution is contained inside the conical volume (lying upside-down) designed by coiled metal wire used as spinning electrode. At the same time, however, the polymer solution that slowly flows through the wire gaps forms a thin film on the outer surface of the coil and from there, in a similar manner as described for the previous examples, the Taylor cones form and hence the fibres are ejected. This configuration displayed an increased throughput in comparison to the basic needle setup (around one order of magnitude) with a similar or even lower diameter for the electrospun fibres. Nevertheless, it also had some drawbacks and in

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particular the need for a high voltage to initiate the process (45 kV) and the necessity to ‘adjust’ the wire gaps to the desired surface tension and rheology of the polymer solution (or vice versa).

A final example of electrospinning from a free liquid solution is bubble-electrospinning [21]. The design is as simple as it is ingenious: it consists simply of a reservoir of polymer solution where bubbles are generated by means of a gas insufflation. The solution is charged by dipping an electrode in it and, once the bubbles grow sufficiently large, they ‘explode’ and generate many jets that can be collected on the target. Obviously, the solution needs a sufficiently high surface tension to generate the bubbles and, usually, the collected fibres have a large diameter and a relatively short length due to the intrinsic intermittency of the process. In spite of that, authors reported a throughput that can reach several grams per hour just working with a single bubble [22]. Furthermore, this kind of experiment, thanks to its unusual process of fibre generation, may give a new glimpse into the electrospinning phenomenon from a thin layer of free liquid surface.

2.2.1.4 Charge Injection Method

The so-called electrospinning by the charge injection method is so peculiar that it merits a separate section for its description. This technique has several advantages and, as will be shown, a high throughput is the most attractive among these.

The process is substantially different from the other apparatuses in the sense that no Taylor cones are formed during the fibre formation and the electrical charge operates more on splitting the emitted jet instead of accelerating and stretching it. It is not possible to give here the theoretical details of the process (see Salem [23] for an exhaustive description); however, from a phenomenological point of view, this is clearly reminiscent of the Rayleigh-type Coulombic rupture. This phenomenon occurs when a liquid column is charged by very high electric field to the point that the electric stresses overcome the surface tension causing the ramification of the fluid in many branches (Figure 2.6b). For an electrospinning process this event is usually competitive with the whipping that is rarely observed in the charge injection method.

As stated, the ramification of the ejected solution is at the base of the technique since it induces a dramatic increase of the produced fibres and hence the throughput can be extremely high (some millilitres of solution per second!). The yield is also fostered by the commonly employed apparatus architecture [24] (Figure 2.6a). Indeed this is made of a grounded orifice (‘blunt electrode’); above that lies an emitter electrode that, also thanks to its sharpened geometry, creates a particularly high electric field. When the fluid flows around the emitter, it is charged and then ejected through the

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orifice by means of a forced system. At this point, the fluid starts the ramification due to the high charge density accumulated and is finally collected on a target. As can be clearly understood, the velocity of the fluid stream depends only on the applied pressure and therefore it can be extremely high; at the same time, however, due to the close proximity of the emitter electrode and the grounded orifice, it is possible to use only insulating or weakly conductive fluids. Another drawback of this design is the low control over the fibre dimension and size distribution. In fact, the electrospun mat is usually composed of microfibres interconnected by a branched network of nanometric fibres.

Figure 2.6 (a) Charge injection method setup scheme and (b) jet formation by ramification. Reproduced with permission from A.J. Kelly, Journal of Aerosol

Science, 1994, 25, 1159. ©1994, Elsevier [24]

2.2.2 Setup Modification for Special Purposes

2.2.2.1 Patterned Deposition of Fibres

Despite the fact that a large effort has been addressed to the enhancement of the throughput, other unusual electrospinning setups have been devised for special purposes. One of the first studied and most common investigations concerned the

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production of orderly-patterned fibre depositions instead of the chaotic woven/non-woven mat. A large portion of the section on ‘mesh morphologies’ will be devoted to this topic; in particular, that section will dwell on the methodologies devised to align fibres by means of a particular collector design and arrangement. Here, only a special apparatus, named near-field electrospinning (NFES), will be considered since it implies the modification of the spinning head and consequently it matches the subject discussed in this section.

The major characteristic of NFES apparatus is a spinneret-collector distance that ranges from a few millimetres to hundreds of microns [25]. This proximity between the spinneret and the collector can cause electric breakdown or fibre splattering on the target due to incomplete solvent evaporation. Consequently, the polymer solution has to be properly adjusted. Electric breakdown, instead, is the consequence of the high electric field generated on the spinneret tip (often a sharpened tip dipped in the solution is used instead of a syringe) and hence the voltage has to be limited to few kV or less. At the same time, however, the fibre jet always lies in the stable region and no bending instabilities are observed. This can be used to deposit a straight fibre along an established pattern [26], provided that the self-expelling of nanofibres (the mutual repulsion of the deposited fibre due to their even charge) is limited by keeping the spinning head in continuous relative motion to the collector. The technique can thus be useful for various applications ranging from sensors to electronic devices, but it is worth underlining that the limited amount of material used at a time is a severe restriction to its implementation.

2.2.2.2 Solvent-free Electrospinning

An NFES-like setup has also been adapted for the spinning of a polymer swollen in supercritical carbon dioxide [27]. This complex technique was developed to spin a polymer from very small droplets before solvent evaporation. In fact, carbon dioxide acted as a solvent and, due to the peculiar working environment, the polymer could be maintained indefinitely in a swollen state. At the same time, however, the results reported in the literature showed that the spun fibre had a remarkably large diameter (hundreds of microns) and, at the present, could form just a short wire from the spinneret tip to the collector.

The utilisation of solvents (except water) in industrial processes is usually unwanted due to several complications: the cost of solvents (sometimes relevant), the environmental restrictions (due to their toxicity and inflammability), their recovery, storage and dispose after use. In most cases, for example for biomedical uses, solvents must be completely removed from the end product. Moreover, important industrial polymers (such as PP or polyethylene) are extremely difficult to process from solution.

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To solve this limitation, in the 1980s electrospinning from melt polymers was investigated [28] but since then a few other research groups have brought further developments to the technique. In fact, the equipment required is relatively expensive, needs a precise temperature-control system and does not systematically produce fibres of nanometric diameter. In fact, due to the high melt viscosity, the diameters of electrospun fibres are usually micrometric and in the few studies where nanofibres were obtained, the key factors affecting the results remained elusive [29]. To reduce the fibre diameter, some strategies have been devised, such as use of viscosity-reducing additives [30] or vacuum chambers [31]. Despite these limitations some interesting applications can be devised, such as the electrospinning of a fibre mat directly on to cells [32].

2.2.2.3 Mats of Multiple Polymers

Electrospinning is, per se, a versatile technique and consequently it has been quite easy to conceive setups able to combine different polymers. One of the simplest ways to obtain hybrid electrospun materials is to use a multi-needle setup (as previously described) to spin different polymers from different syringes [33] (co-electrospinning). Obviously the same difficulties affecting multi-needle design are still present in this technique; however, since the throughput is not a primary concern in this case, interesting final properties can be obtained if the fibre non-uniform deposition is taken under control [34].

Bi-component single fibres have been obtained by merging side-by-side, two spinnerets fed by separated reservoirs [35, 36]. In spite of the fact that electrospinning of polymer blends can be achieved by spinning a mixture of polymers or polymer solutions through a single needle, this setup avoids some limitations related to the physics of polymeric blends. Indeed, with a single spinneret, a homogenous, thermodynamically stable mix of the polymers is required and, in the case of solution electrospinning, the interactions between the polymer and the solvent of the opposing pair become crucial. With two separate spinnerets, the polymer solutions come in contact only at the end of the spinning head, allowing for the formation of blend fibres that are otherwise difficult to produce.

A similar concept is that of the coaxial electrospinning, where two concentric needles are used (Figure 2.7a). In this way, it is possible to simultaneously spin fibres with different outer shell-inner core materials [38] (Figure 2.7b). The system needs an optimal assembling precision to avoid non-concentric syringes and hence fibre deformations. At the same time, however, it allows for a certain degree of versatility in fibre morphologies by simply ‘playing’ with the surface tension and rheology of the two immiscible solutions (see next sections). Furthermore the inner electrospun

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solution does not necessarily need to be polymeric, it can simply be a liquid that finally forms a cavity. The hollow fibres so produced promise to be a versatile drug delivery system as will be better described in the following sections [37].

Figure 2.7 (a) Working principles of coaxial electrospinning and (b) particular of the Taylor cone displaying the core-shell system.

PEG = polyethylene glycol, DMF = dimethylformamide, PCL = polycaprolactone. (a) Reproduced with permission from H. Jiang, Y. Hu, Y. Li, P. Zhao, K. Zhu and

W. Chen, Journal of Controlled Release, 2005, 108, 237. ©2005, Elsevier [37] (b) Reproduced with permission from J.E. Díaz, A. Fernández-Nieves, A. Barrero,

M. Márquez and I.G. Loscertales, Journal of Physics: Conference Series, 2008, 127, 012008. ©2008, IOP Publishing Ltd [38]

2.3 Mesh Morphologies

While electrospinning is an outstanding production method for the manufacture of nano- and micro-fibres with defined diameter, controlling the overall porosity has remained elusive, particularly for large values of this parameter. The porosity dimension is still an obstacle limiting the application of electrospun scaffolds in tissue engineering since it does not allow cell migration through the small pores [39], and prevents vascular ingrowth [40] and nutrient diffusion, which limits the thickness of the scaffolds and reduces cell proliferation and metabolism. Conventional electrospun

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meshes are usually composed of nanometric fibres that are densely packed, thus reducing the pore dimension. Although the conventional electrospun meshes have high porosity and good interconnectivity, the reduced pore size is a limitation for cell ingrowth.

Porosity, pore size and distribution, fibre size and alignment constitute, together with the intrinsic properties of the materials are the main features responsible for the crosstalk between scaffold and cells. It has been also demonstrated that micro- and nano-structures as well as fibre alignment affect cellular behaviour such as adhesion migration and cytoskeletal organisation [41].

During the evolution of the electrospinning technique, authors developed methods to control the fibre deposition. Studies have focused on the modification of the electrical parameters of the system either by the application of auxiliary electrodes and adaptation of the target geometries.

Three types of electrostatic forces mainly control the deposition of the electrospun fibres: the external electric field applied on the polymer solution; the target geometry; and any adjacent charged fibres. When a conventional target, a stationary continuous conductive plate is used, the electrostatic forces will have no preferential direction and the mesh will present a random disposition of fibres. Thus, the manipulation of the electric field and the use of dynamic collectors are the most common methodologies of getting controlled deposition of fibre assemblies. Usually, the two methods are combined to achieve greater order in the fibre deposition.

Normally, the manipulation of the electric field occurs by the application of external electrodes or changing the configuration of the stationary target. Altering the electric field, it is possible to better control the fibre deposition either for jet constriction or fibre alignment. The field vectors of a few different electrospinning setups are presented in Figure 2.8, where (a) represents the conventional electrospinning, (b) the application of auxiliary ring electrodes and (c) parallel electrodes used as target. Deiztel and co-workers [42] applied a positively charged, auxilliary ring electrodes (Figure 2.8b) below the tip of the syringe needle in order to create a pulling force down to the target and constrict the jet to reduce the mesh diameter. Li and co-workers [43] demonstrated that two electrodes placed in parallel collected aligned fibres in the gap between the electrodes. They presented the electrical field vectors for this configuration (Figure 2.8c), which exert a pulling force towards the target and by the balance of electrostatic forces are able to align the fibres in the gap.

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Figure 2.8 Calculated electric field strength vectors in the region between the needle and the target. The arrows denote the direction of the electrostatic field lines at (a) the conventional electrospinning, (b) the presence of auxiliary ring

electrodes and (c) parallel electrodes are used as target [conductive silicon (Si)]. (a) and (b) reproduced with permission from J.M. Deitzel, J.D. Kleinmeyer, J.K. Hirvonen and N.C. Beck Tan, Polymer, 2001, 42, 8163. ©2001, Elsevier [42]. (c) reproduced with permission in part from D. Li, Y. Wang and Y. Xia, Nano

Letters, 2003, 3, 1167. ©2003, American Chemical Society [43]

As previously mentioned, the modification of the target geometry is an alternative method that modifies the electrical field of the electrospinning system. In contrast to the conventional stationary continuous target where the fibres are deposited randomly and densely packed, here we present some of the modified targets and 3D deposition techniques for controlled fibre deposition and for controlled scaffolds porosity in three dimensions, respectively, with their common advantages and disadvantages.

2.3.1 Rotating Mandrel

Being the most common derivation of the conventional target system, the rotating mandrel (Figure 2.9a) allows the even collection of the fibres with a uniform deposition and controlled porosity. The deposition of a well aligned fibre is possible as soon as the tangential speed of the mandrel reaches the threshold of the polymer jet speed

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[41, 44], and has been exploited to produce either aligned and random meshes as well as to manufacture tubular scaffolds for vascular grafts. Moreover, aligned and random fibre layers can be alternated in order to design the mechanical properties of the mats. Nevertheless, this system does not allow the formation of thick polymer meshes due to the insulator effect caused by the first layers of fibres.

2.3.2 Rotating Tube Collector with Knife-edge Electrodes

This geometry was developed to overcome the limitation of grounded rotating targets where the deposited polymer acts as an insulating barrier for the mesh growing in thickness. Applying negative charged auxiliary knife-edge electrodes below the insulating rotating mandrel (Figure 2.9b) authors have demonstrated the ability to produce highly aligned electrospun meshes with different orientation angles [45]. Positioning the electrodes below the mandrel, it is possible to avoid the insulation effect of the deposited fibres and maintain the electrical field in order to deposit thicker layers of ordered nanofibres. An additional feature is the ability to tilt the needle and the electrodes (Figure 2.9c) to deposit fibres with a different orientation and manufacture multi-layered scaffolds with tailored mechanical properties.

2.3.3 Disk Collector

Another approach to positioning and aligning individual fibres was proposed by Theron and co-workers [46], by using a grounded sharped-edge disk as a collector (Figure 2.9e). The knife-edge concentrates the electrical field allowing the deposition of most of the electrospun fibres in a continuous manner on the rotating disk. With this deposition, fibres retained substantial electrical charge before reaching the target, this leading to repulsion resulting in a larger distance between two aligned fibres. This distance may vary depending on the repulsive forces related to the residual charges and fibre diameter. Although this geometry is suitable for highly aligned fibres and allows the continuous production by removing the fibres during the wheel rotation, there are some disadvantages such as the small area of fibre deposition and loss of alignment as the mesh grows in the 3D direction.

2.3.4 Parallel Electrodes

Grounded parallel electrodes have been firstly proposed by Li and co-workers [43], who showed the possibility to uniaxially align fibres by using two electrodes positioned parallel to each other (Figure 2.9f). The alignment of the fibres among the electrodes has been correlated to the equilibrium of the electrostatic forces developed

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by the split of electric field on the targets and by the charged fibre on the surface of the targets [43, 47]. This geometry allowed the production of highly aligned fibres and their easy transfer to another substrate. The distance between the electrodes may vary from microns to several centimetres, depending on the applied electrical field. Nevertheless, there are some limitations to be considered, such as difficulties in producing aligned thick meshes due to the insulator effect of the deposited fibres and the necessity of depositing fibres with a minimum diameter of 150 nm in order to support the mechanical stress caused by their weight.

2.3.5 Patterned Electrodes

The patterned electrodes setup is a derivation of the parallel electrodes geometry. In fact, the physical aspects that govern the deposition are the same [47]. Nevertheless, in this configuration more electrodes are used in order to have multi-layered mats with controlled hierarchical structure. Controlling the location and configuration of the paired electrodes (Figure 2.9g), the fibres’ spatial orientation and position can be determined since there is the uniaxial direct fibre deposition in the electrode’s gap. Moreover, controlling the scheme of high voltage application, it is possible to deposit multi-layered mats with different orientation, which is of important interest for mechanical properties. Nevertheless, analogously to the parallel electrodes, there are some limitations to the process such as minimum thickness of the fibres and maximum distance between electrodes.

2.3.6 Rotating Wire Mandrel

Taking advantage of the potential of the rotating drum and parallel electrodes to align the electrospun fibres, authors have combined the two geometries to develop a new target [48, 49]. This target is composed of several equidistant grounded wires in order to form a cylinder (Figure 2.9h). In this configuration the electrical field is similar to the parallel electrodes due to the rotation of the cylinder, which deposit highly aligned fibres. Nonetheless, the limitation of short length deposition of a few centimetres is overcome by the continuous rotation of the target. However, difficulties with detachment of the fibres is always a concern in this configuratio

2.3.7 Parallel Rings

A pair of grounded rings placed equidistantly from the capillary tip yielded the deposition of aligned nanofibres between the rings (Figure 2.9i). Despite the fact that this target geometry allowed the production of highly aligned fibres, the suspended

41


fibres are difficult to characterise and post-process without disturbing the fibre alignment.

2.3.8 Microfibre Assisted Rotating Collector

An innovated target system has recently been developed in order to produce nanofibre-coated microfibres [50, 51]. In this method, the nanofibre deposition is assisted by a previously produced microfibre, which is moved slowly towards a small collector that rotates around it (Figure 2.9j). The rotation of the collector forces the nanofibres to twist around the microfibre, thus creating the coating. This modification allows the formation of aligned and continuous threads of either single or multi polymer compositions that can be further interlaced to form a woven scaffold or applied as produced. A highly porous (up to 97%) nanofibre-coated microfibre scaffold can be obtained by the application of this target geometry [51].

2.3.9 Liquid Bath Collector

Unlike most of the solid targets applied for electrospinning, some authors have presented an alternative method for collecting the electrospun fibres in a liquid bath [52, 53] (Figure 2.9k). In this geometry the polymer solution is electrospun into a liquid bath, which may contain different solvents depending on the purpose of the deposition. A solid grounded electrode is positioned inside the liquid in order to direct the electric field towards the liquid, which inhibits the fibre deposition on the solid surface. The authors have used this geometry for two main purposes: first, as a method to avoid the deposition on a solid surface, in order to collect single nanofibres or yarns [52], and second, in order to induce phase separation of the fibres described next [53].

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Figure 2.9 Schematic diagram of various electrospinning targets. (a) Rotating mandrel; (b and c) rotating tube collector with knife-edge electrodes; (d) rotating frozen mandrel; (e) disk collector; (f) parallel electrodes; (g) patterned electrodes;

(h) rotating wire mandrel; (i) parallel rings; (j) microfibre assisted rotating collector and (k) liquid bath collector. PLA = polylactic acid (a, b, c, e, f, g, h, i and k) Reproduced with permission from W.E. Teo and S. Ramakrishna,

Nanotechnology, 2006, 17, R89. ©2006, IOP Publishing Ltd [54]. (d) Reproduced with permission from M. Simonet, O.D. Schneider, P.

Neuenschwander and W.J. Stark, Polymer Engineering & Science Engineering, 2007, 47, 2020. ©2007, John Wiley and Sons [55].

(j) Reproduced with permission from A. Thorvaldsson, H. Stenhamre, P. Gatenholm and P. Walkenström, Biomacromolecules, 2008, 9, 1044. ©2008,

American Chemical Society [51]

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2.3.10 Rotating Frozen Mandrel

The low temperature deposition, or cryogenic electrospinning, applies the rotating frozen mandrel as previously presented, where the cylindrical target is kept at temperatures below -30 °C in order to allow ice crystals to form into the meshes along with the polymer deposition. This is a recent refinement of the electrospinning process that aspires to increase the porosity of the meshes as well as enlarge pore dimensions. First presented by Simonet and co-workers [55] (Figure 2.9d), the technique is based on the condensation of water vapour from the environmental humidity on the target surface while the polymer is deposited. The low temperature of the target freezes the condensed moisture inhibiting the polymer deposition in certain areas, thus forming an open 3D mesh with large void spaces, as can be seen in Figure 2.10. In fact, Simonet has shown the contrast between conventional and the low temperature collection of fibres, where there is a significant increase of mesh porosity and mean fibre distances. The dimension, distribution and amount of porosity are directly related to the humidity in the surrounding environment. He achieved 95% of mesh porosity when deposition occurred at 85% of relative humidity.

Figure 2.10 (a) and (c) Side view (cross sections; SEM images) of poly(lactic-co-glycolic acid) (PLGA) meshes electrospun at 30% relative humidity and room

temperature; (b) and (d) using a dry ice cooled collection drum. Reproduced with permission from M. Simonet, O.D. Schneider, P.

Neuenschwander and W.J. Stark, Polymer Engineering & Science Engineering, 2007, 47, 2020. ©2007, John Wiley and Sons [55]

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The authors have correlated the condensation of surrounding moisture at the target’s surface to the temperature gradient formed between the environment and the cooled cylinder. As a consequence, the air becomes colder, passing the dew point. Once saturated, water precipitates from the humid air forming the condensed water droplets, thus, forming ice crystals from the contact with the cold cylinder [55, 56].

The biological evaluation of networks manufactured by this method has been addressed by Leong and co-workers [56] and Schneider and co-workers [57]. The analyses have shown the improved cell infiltration and proliferation using scaffolds produced by low temperature deposition. Leong has shown that 3T3/NIH fibroblasts penetrated the scaffold up to 50 μm in depth after seven days of culture in contrast to the conventional electrospun meshes that did not allow any cell ingrowth.

2.3.11 Addition/leaching of Salts and Gas Foaming Methods

Methods based on porogen leaching have received much attention in recent years because of their efficacy in controlling the porosity of the scaffolds produced by the simple variation of the amount and size of leachable particles, such as sodium chloride. Nevertheless, the scaffolds usually have a lack of pore interconnectivity. In order to solve this limitation, authors have introduced leachable porogens able to produce gas while dissolution occurs [58, 59], such as ammonium bicarbonate, thus allowing the interconnection of the pores.

Taking advantage of the salt leaching technique for production of 3D scaffolds for biomedical applications [60, 61], authors were able to significantly enhance the porosity of electrospun networks by combining the two methods [59, 62, 63]. In this new modification of the electrospinning process, the deposition of salts among the electrospun mesh is performed by a simple sieving technique [63] or a more sophisticated method where the porogens are delivered from a concentric needle mounted externally to the needle that delivers the polymer solution [62].

The application of porogens along with electrospinning has been shown to enhance the porosity and thickness of the mats, nevertheless some issues must to be taken into account when applying this technique. The amount of porogens has to be properly controlled to avoid an excessive content, which would significantly reduce the bonding among the fibres, thus causing the delamination of the mesh [62]. A second concern is related to the collapse of the electrospun structure. Polymers with low mechanical properties and/or the application of large sized porogens may cause the collapse of the mesh after porogen removal. Several authors have suggested overcoming the structure collapse by crosslinking the mesh before the salt leaching [59, 63].

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Biological evaluation of scaffolds manufactured with this technique has been reported in literature. Nam and co-workers [62] reported that CFK2 cell line has migrated for 4 mm into a 6 mm scaffold, covering approximately 70% of the scaffold cross-section.

Although there are several reports in the literature, there is still need for improvement of this combined technique since they do not cover all the aspects that influence the physical and biological properties of the mesh.

2.3.12 Selective Dissolution of Fibres

Selective dissolution of fibres is a method based on the co-electrospinning setup in which, as previously presented, there is the deposition of two distinct polymers contemporaneously, usually with a rotating mandrel as target. In this technique one of the selected polymers, also called the sacrificial polymer, is soluble in aqueous solution and allows the dissolution of the fibres after deposition, so increasing porosity and pore size in the meshes. It is an interesting method since it allows the production of either aligned or random meshes. The porosity can be easily controlled by the ratio and diameter of sacrificial fibres present in the mesh. Nevertheless, it is necessary to correctly design the system in order to avoid the collapse of the structure after removal of the fibres. As with the salt leaching technique, crosslinking the polymer mesh may overcome the collapse problems. Baker and co-workers [64] and Whited and co-workers [65] have applied the electrospinning of sacrificial fibres in order to increase pore dimensions to enhance cell proliferation and ingrowth on polycaprolactone (PCL) and poly-L-lactide (PLLA) scaffolds, respectively. In fact, Baker showed (Figure 2.11) the application of the technique to aligned fibres of PCL and polyethylene oxide (PEO), where the average pore area triplicated when 60% of PEO fibres were used. This result was confirmed by in vitro analysis, which demonstrated the ability of cells to migrate up to 100 μm deep into the scaffold.

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Figure 2.11 Composite fibrous scaffolds can be formed with individual fibres of distinct polymer composition. The removal of one sacrificial fibre population

increases scaffold porosity. (a) Fluorescently-labelled PCL (red) and PEO (green) fibres showed pronounced alignment and interspersion. (b) Submersion of

scaffolds in an aqueous solution removed the PEO component but left the PCL fibres intact. SEM images of (c) as-spun and (d) post-submersion composite

scaffolds revealed increases in pore size with the removal of sacrificial PEO fibres. Scale bars: 50 μm. Reproduced with permission from B.M. Baker, A.O. Gee, R.B. Metter, A.S. Nathan, R.A. Marklein, J.A. Burdick and R.L. Mauck, Biomaterials,

2008, 29, 2348. ©2008, Elsevier [64]

2.3.13 Bimodal Distribution of Fibre Diameters

While most researchers have focused on the deposition of unimodal fibre distributions both on the nanoscale or the microscale for the production of two-dimensional (2D) and 3D scaffolds, others decided to apply a different approach by depositing a combination of two or more fibre populations [66-69]. Either applying the co-electrospinning technique or simple multi-step deposition, this approach allows the production of highly porous mesh morphologies with large pore diameters. Depending on the applications, continuous or discontinuous mesh morphology can be produced by selecting the deposition methods such as co-electrospinning or multi-step deposition, respectively. Authors have proposed this method in order to better mimic the architecture of the extracellular matrix, which is composed either by nano- and micro-metric fibres [70], and consequently to enhance cell infiltration.

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An additional feature of these scaffolds is the improved mechanical properties since the thicker fibres can support extra load.

Several authors have carried out biological evaluation of scaffolds with multimodal fibre distribution and the system has been proven to enhance cell behaviour in terms of cell infiltration, proliferation and activation. Santos and co-workers [68] demonstrated the ability of a scaffold with a biomodal fibre distribution, 160 μm and 400 nm fibre diameters, to provide structural and organisational stability to endothelial cells (EC). They also observed a marked angiogenic potential due their ability to form extensive networks of capillary-like structures in such scaffolds.

2.3.14 Combined Solid Free Form (SFF) and Electrospinning Process

Rapid prototype techniques, such as SFF, have been applied for the manufacture of tissue engineering scaffolds for the past decade. The SFF technique is based on the deposition of a polymer melt or solution on a computer-based platform. The system has three or four degrees of freedom, allowing the manufacture of 3D structures by the deposition of successive layers of polymer strands with a precise control of the scaffold characteristics. Despite the great control of the scaffold features such as pore size, porosity and pore distribution, the technique has limitations related to the biological interactions. In contrast to electrospinning, SFF scaffolds have large pores and fibre diameters, smooth fibre surface and outstanding mechanical properties [71, 72], which affect material-cell interactions. Cells seeded on SFF scaffolds tend to respond as if they were on a 2D surface due to the large strand diameter, in the order of several hundred μm [73, 74], and the smooth surface and distance between strands can inhibit cell attachment and proliferation/migration, respectively [75].

To vanquish the limitations, authors have combined the SFF process to electrospinning [71, 73, 74, 76], allowing the contemporaneous deposition of polymer micrometric strands along with electrospun nanometric fibres (Figure 2.12). As with the bimodal distribution of fibre diameters, the combination of the processes aims to achieve 3D scaffolds that mimic the natural porosity of human tissues and overcome the limitations of the single techniques in terms of the scaffold’s porosity, cell infiltration, adhesion and proliferation.

Authors have shown the improvement of 3D SFF/electrospun scaffolds in supporting cell attachment and proliferation of various cells, such as chondrocytes and mesenchymal stem cells (MSC), when compared to conventional electrospun networks and SFF scaffolds [71, 72]. The morphology of the scaffolds’ manufacture with this technique can be observed in Figure 2.12.

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Figure 2.12 Schematic representation of the process combining SFF and electrospinning and a SEM image of the scaffold morphology. Reproduced with permission from G.H. Kim, J.G. Son, S.A. Park and W.D. Kim, Macromolecular Rapid Communications, 2008, 29, 1577. ©2008, Wiley-VCH Verlag GmbH &

Co. KGaA [71]

2.4 Fibre Morphologies

One of the most interesting aspects of electrospinning is the possibility of producing mats with very high surface-to-volume ratio. Polymeric fibres with a diameter in the range of hundreds or tens of nanometres can have a specific surface area in the order of 1-100 m2/g [77]. This value is found in smooth fibres, but even higher values (30-65% more than traditional electrospun fibres [78]) can be obtained if a sub-micrometric or nanometric surface texture or bulk structure is formed. Typical examples are the production of fibres with a corrugated surface (and not-circular cross-section) or of fibres with high porosity. These features strongly influence the fibre characteristics in

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relation to their biomedical applications. Surface porosity, for example, is fundamental when considering a cell’s interaction with electrospun scaffolds.

In this research area, several studies have been conducted, dealing with the cellular behaviour when they are in contact with a nanoscale environment [79], with enhancement (through the control of micro and nano roughness) of the activity of osteoblasts [80], EC [81], MSC [82], fibroblasts [83] and other cell types.

Another key issue in tissue engineering is the entrapment/release of bioactive molecules. For this purpose, the production of coaxial or hollow fibres is of great interest. This research topic is connected to the release of drugs in a controlled way and to the chemical pattern of surfaces, which guide cell adhesion, proliferation and differentiation. The continuous development of new drugs, in particular the protein-based ones, requires the parallel development of proper releasing systems; among these, biodegradable polymeric fibres and particles are the most promising [84, 85]. Integration of different techniques in scaffold production is very important [86]. The possibility of integrating scaffold production with the controlled release of drugs or of growth factors shows tremendous potential in tissue engineering [87-89].

It is worth mentioning that the phenomena described here for electrospinning are also interesting for the production of micro-particles using electrospraying. Indeed, the two processes are strongly connected by the balance between viscous forces and surface tension of the polymeric solution. Changing the process parameters and the solution properties, it is possible to produce fibres, beads or both. A low polymer concentration (meaning a low viscosity), for example, leads to the production of beads, but an increase of this parameter causes the formation of a beads-on-fibres structure; a further increase leads to the formation of fibres [90]. Thus, the processes analysed for fibres can also be applicable to beads, such as the production of multi-layer microparticles with a co-axial setup or the formation of pores on a bead surface [91, 92].

2.4.1 Wrinkled and Ribbon-like Fibres

After the ejection from the needle, fibres dry due to solvent evaporation. Fibre cross-sectional geometry is not always circular; depending on the material used, several geometries can be observed, such as flat ribbon-like or wrinkled-surface fibres [93, 94] (Figure 2.13). Solvent evaporation from the surface leads to the formation of a thin sheath of solid polymer, which surrounds a softer core (still rich in solvent). The core shrinks during solidification, causing a circumferential compressive stress in the glassy skin. If the compressive strength reaches a critical value, which depends on geometrical factors and on the elastic properties of the fibre, it causes a buckling

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of the solid outer skin, resulting in a wrinkled surface and in a cross-section with a non-constant radius.

Figure 2.13 Different morphologies (fibres and cross-section) obtained with electrospinning. Reproduced with permission from L. Wang, C-L. Pai, M.C. Boyce and G.C. Rutledge, Applied Physics Letters, 2009, 94, 151916. ©2009, American

Institute of Physics [77]

The phenomenon is characterised by a specific wave number (k), defined as the number of local radius maxima along the perimeter of the fibre cross section [77]. Minimising the energy of the system and approximating the core Poisson’s ratio (ν) to a value close to 0.5 (which is a good approximation for the core of solidifying fibres), a critical value for the wave number can be calculated as:

��

��

� � ��

� � ��

�� (2.1)

where c refers to the core, s refers to the sheath, E is the Young’s modulus, a is the radius of the core and t is the thickness of the sheath. Equation 2.1 describes the wave

51


number, which minimises the total energy of the system. If the core has a low relative stiffness, meaning low values of Ec/Es and a/t, its deformation requires little energy and the wave number is small, with the minimum being k = 2, corresponding to the ribbon-like cross-section (Figure 2.13a). On the contrary, if more energy is necessary to deform the core, a high wave number results, corresponding to fibres with several wrinkles on their surface (Figure 2.13b and 2.13c). The parameter t, thickness of the polymer skin, is influenced by the volatility of the solvent, thus allowing the control of the wave number by means of a proper choice of the solvent used. For a given polymeric solution (polymer and solvent), all the parameters in Equation 2.1 are fixed except for the core radius a, thus the wave number is proportional to the fibre diameter. Bigger fibres have a higher wave number, while the dimension of wrinkles (crest to crest distance) is independent of the radius. Interestingly, in a mat composed of beads and fibres, solvent evaporation is almost uniform, leading to similar values of t, but beads have higher a values, resulting in higher wave number (Figure 2.13d) [77].

The collapse of the fibre in a ribbon-like shape (k = 2) causes a redistribution of the electrical charges, which migrate toward the edges of the flat fibres, causing a repulsive force that favours the collapse. Moreover, the collapse is enhanced by attractive forces between opposite sides of the forming skin that come in contact. The final result is a flat fibre, which can possibly present two tubes as edges, depending on charge repulsion and on the skin stiffness [93]. As collapse occurs at randomly distributed sites and on different radial directions, flat fibres appear as if they were twisted

2.4.2 Porous Fibres

As stated previously, porous electrospun fibres have attracted increasing interest in tissue engineering. The formation of pores can be localised only on the surface or involve the entire cross-section of fibres, depending on the physical phenomena occurring during the electrospinning process. Porous fibres are interesting for a wide variety of applications. High specific surfaces are interesting in filtration applications and in entrapment of bioactive molecules, such as enzymes. Furthermore, the presence of pores can enhance the capture of nanoparticles and strongly influence the encapsulation and release of drugs. Fibre porosity is also of fundamental importance in the production of electrospun scaffolds for tissue engineering. Cell behaviour is strongly influenced by the surrounding environment, thus the control of the topographical features of the surface they are in touch with guides their adhesion, migration, proliferation and differentiation.

Sub-micrometric structures can be obtained in polymeric materials using a variety of techniques. In particular, self-assembling processes are of great interest due to the high control of features down to the nanometric range and to the possibility of

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performing simple one-step processes, which form the fibre and the substructure at the same time. In electrospinning, porous fibres have been observed in several experiments, using diverse polymers, solvents and process parameters. Despite the fact that the formation of pores have been widely studied in films and bulk polymers, the complexity of the electrospinning process (due to fast fibre formation, electric charge distribution, stretching and small dimension of fibres) leads to a number of problems in determining the physical phenomena responsible for the final structures obtained. Mainly two processes have been found to be responsible for the formation of porous electrospun fibres: the Breath Figures process and phase separation [78, 95-97].

The Breath Figures process indicates the pattern formed by water droplets condensing from atmospheric humidity on a solid or liquid surface which is cooler than the surrounding atmosphere. During the drying of a polymeric solution, evaporation of solvent causes a cooling of the surface. Depending on solvent properties, such as enthalpy of vaporisation and vapour pressure, the cooling can be sufficient to promote the condensation of water droplets, which arrange themselves on the surface and partially sink in the solution, acting as a template. The solution surrounds the droplets, guided principally by surface tension, convective motion, interfacial and kinetic parameters [98]. Finally, droplets evaporate, after solution drying, leaving an array of pores on the polymer. This process has been studied mainly using polymeric films to obtain an honeycomb-like regular array of pores, disposed both in mono- or multi-layers, but it can occur also during fibre formation (Figure 2.14). Breath Figures formation requires the presence of a certain level of humidity in the atmosphere and the use of a volatile solvent (for the surface cooling) with a low water solubility (so condensing water droplets do not mix with the solution). The pores features depend on the characteristics of the polymer (chemistry and molecular weight), of the solvent (vapour pressure, enthalpy of evaporation, surface tension) and of the solution (viscosity, hydrophilicity, thermal properties, viscosity). Electrospinning is a very fast process, since drying of nanofibres takes milliseconds to occur, so it is difficult to understand if the breath figures templating process can occur in such a short time.

Phase separation can be exploited to obtain porous fibres in several ways. First of all, it should be considered that phase separation could involve different systems in which an immiscibility area is present in the thermodynamic phase diagram. The solution composition, initially under conditions of miscibility, passes through the binodal curve during the process, thus entering in to the metastable region where phase separation occurs by nucleation and growth. Alternatively, it can pass this region fast enough to avoid nucleation of the new phase and, through the spinodal curve, enter in to the unstable region in which spinodal decomposition occurs. The morphology of the phases depends on which one of these two areas the solution ends in. Nucleation and growth leads to the formation of a continuous phase in which the second phase grows in isolated zones, while spinodal decomposition leads to two

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interconnected phases. The previously mentioned phase separation phenomena lead to the formation of two phases in a polymeric solution: one that is solvent-rich and the other that is solvent-poor. After fibre drying (solvent evaporation), the solvent-rich regions form the pores.

Figure 2.14 Porous fibres obtained electrospinning poly-DL-lactide acid (PDLLA) in chloroform at 70% relative humidity

A mathematical model of the possible morphologies that could be obtained depending on the balance between solvent evaporation rate and thermodynamic driven phase separation was proposed by Dayal and Kyu [99]. Initial polymer concentration, solvent diffusivity and temperature were varied in this study. If solvent evaporation is fast enough, no phase separation occurs. Instead, a solid skin develops, resulting in a skin-core structure in which the inner part of the fibre is rich in solvent. With the proper conditions, a hollow fibre can be obtained (Figure 2.15a). With a slower evaporation rate and starting from a concentration, which is in the unstable region, the system can be driven in the stable one-phase region, resulting in a monolithic fibre (Figure 2.15b). Reducing the evaporation rate and accelerating the kinetics of phase separation (lowering the temperature), the core has enough time to phase separate, even though a solid skin still forms (Figure 2.15c). If the evaporation rate is further decreased, phase separation occurs in the whole fibre, leading to the formation of a fibre, which is porous throughout (Figure 2.15d).

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Figure 2.15 Fibre morphologies obtained starting from the points a–d within the phase diagram of the polymer-solvent system. Darker regions correspond to lower polymer concentration. Φ is the polymer concentration, T is the temperature, Tc

is the critical temperature. Reproduced with permission from P. Dayal and T. Kyu, Journal of Applied Physics, 2006, 100, 043512. ©2006, American Institute of

Physics [99]

If a one-polymer solution is considered, phase separation can be caused if the concentration of the solution increases above the miscibility limit. This could happen due to solvent evaporation or due to penetration of a non-solvent in the solution. During electrospinning, evaporative cooling can cause the condensation of water on the surface and its penetration into the solution. If the solvent is water-soluble and the polymer is not, the miscibility limit of the polymeric solution can be passed, due to the formation of a mixed-solvent of low quality, or, in other words, to the solvent extraction by the non-solvent. When the non-solvent is adsorbed from the atmosphere, this process is called vapour-induced phase separation [100].

The non-solvent can also be introduced into the polymeric solution as a liquid. A polymeric solution composed of a polymer (PLLA), a volatile solvent (dichloromethane, DCM) and a non-volatile non-solvent (butanol, BuOH) was used by Qi and co-workers [101]. During electrospinning, DCM evaporates rapidly, leaving a fibre composed mainly of PLLA and BuOH. Phase separation occurs due to the non-solubility of the polymer in BuOH. After fibre deposition, the non-solvent evaporates, leaving a porous structure (Figure 2.16). Interestingly, depending on the relative difference

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in evaporation rate of the two liquids, both interconnected or isolated pores were observed, suggesting spinodal decomposition or nucleation and growth, respectively, to be responsible for the porous structure.

Figure 2.16 Porous fibres from a solution of PLLA in DCM/BuOH (60/40). (b) is an higher magnification of a portion of (a). Reproduced with permission from Z.

Qi, H. Yu, Y. Chen and M. Zhu, Materials Letters, 2009, 63, 415. ©2009, Elsevier [101]

A particular non-solvent used to promote phase separation is carbon dioxide (CO2). Precipitation with a compressed fluid antisolvent is a technique that promotes phase separation by spraying a polymeric solution in a CO2 bath, resulting in short hollow or porous fibre [102]. Carbon dioxide is a non-solvent for most polymers but can be mixed with most organic solvents. Its addition in a polymeric solution causes the creation of a mixed solvent of low quality and also dilates the polymer solvent, which increases the free-volume difference, resulting in phase separation. Near-critical or supercritical CO2 diffuses and extracts the solvent faster than CO2. Liu and co-workers [103] have obtained porous and hollow fibres using a high-pressure electrospinning apparatus, in which the production of fibres was performed in presence of pressurised CO2. During the process a solid skin on the fibre is rapidly formed due to solvent evaporation, while CO2 penetrates into the inner-trapped material causing phase separation. Initially, the non-solvent rich regions (in which pores originate after solvent evaporation and depressurisation) nucleate and grow, but the fast penetration of further CO2 can lead to an increase in its concentration, causing spinodal decomposition. Polymer-poor regions can develop to a limited extension, forming a porous fibre with a solid skin, or can coalesce to the limit of forming a

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hollow fibre. Coalescence of these regions is enhanced by the fibre stretching and diameter reduction and also depends on polymer concentration and CO2 pressure.

If the phase diagram exhibits an upper critical solution temperature (UCST) in the range of process parameters, a temperature decrease can lead to a thermally induced phase separation (TIPS). In the case of electrospinning, surface cooling due to solvent evaporation can lower the temperature to values below the UCTS, thus causing phase separation. The presence of pores only on the surface of fibres can be related to a cooling effect which is limited to the surface [100]. A different structure has been obtained inducing TIPS by immersing the collector of electrospun fibres in liquid nitrogen [104]. Phase separation also occurs in the bulk of the fibre, guided by the fast cooling, and the solvent is removed by successive heating to room temperature, resulting in fibres that are porous throughout.

A different technique involving phase separation is based on a system composed of two immiscible polymers and a common solvent. The solution can be electrospun as in traditional electrospinning, but with solvent evaporation the two immiscible polymers form two co-continuous phases. After complete fibre drying, one phase can be removed through selective dissolution. Before the removal, the other phase can be stabilised, usually by crosslinking [105, 106].

2.4.3 Coaxial Fibres

Electrospinning has been studied extensively as a technique for the production of mats as drug delivery devices. Several drugs can be embedded in the polymeric fibres and the release is controlled by their interaction with the polymeric matrix. Polymer degradation is not a concern, since the polymers, e.g., PLA and PCL, degrade in a much longer time than the one required for the release. Antibiotics, proteins and drugs have been tested for sustained release from electrospun fibres. However, the distribution inside the fibres and the kinetics of release are not perfectly controllable. Moreover, proteins and drugs have to be mixed with the polymeric solution to spin, and often the solvent used can damage them or reduce their bioactivity. This problem can be solved with the use of a coaxial setup, as previously described. Drugs are (often) dissolved in an aqueous solution, which preserves their activity, while the polymer solution is obtained using traditional solvents. The two solutions are electrospun passing through two coaxial needles, thus forming fibres composed of an inner core containing the drug and of an outer polymeric sheath. As mentioned before, the drug does not contact organic solvents, preserving its conformation and bioactivity. The outer sheath forms a barrier, which protects the core and avoids rapid dissolution, controlling the release by means of diffusion or desorption mechanisms. Varying the process parameters, it is possible to control the thickness of the sheath and the

57


diameter of the core, obtaining a precise control over the amount of drug inside the fibres, as well as on the release rate. Furthermore, the coaxial system avoids the burst release often observed with other techniques. In monoaxial electrospinning the drug can segregate near the fibre surface, resulting in a fast and uncontrolled initial release. In coaxial electrospinning, on the contrary, the drug is present only in the core, so burst release is strongly reduced. Also, the increased number of variables allows a better control over the release kinetic [107].

Srikar and co-workers [108] compared dye diffusion from polymethylmethacrylate and PCL nanofibres, finding that the release occurs due to two processes: desorption of the drug from the polymer and its diffusion into the release medium. Different materials can behave differently, determining which one of these processes is the limiting one. Desorption depends on the nanoporosity of the material (if it is present, as in PCL), sensitivity to polymer concentration and molecular weight variations, and on the desorption enthalpy, which in turns depends on the intermolecular forces between the drug and the polymer in which it is embedded [108, 109]. Interestingly, not all the drug can be released, but only the portion that desorbs from nanopores, until polymer degradation occurs.

An advantage of coaxial electrospinning is the possibility of producing fibres composed of materials that are difficult, if not impossible, to spin with the traditional apparatus. These materials can be used to produce the core, while more spinnable polymers are used for the outer sheath. The outside sheath material makes a fibre that traps and guides the inner core, and its selective dissolution leaves a mat composed only of the core material. Dissolution can be preceded by a process that stabilises the inner core, such as crosslinking. Diaz and co-workers [38] have used an alginate as the core and PEO as the sheath material. The deposited fibres were immersed in an aqueous calcium chloride solution, so that calcium ions can diffuse through the sheath, while it dissolves, and crosslink the alginate. Washing in water removed the residual PEO, resulting in a mat composed of alginate nanofibres. A similar process has been used by Wang and co-workers [110] to produce silk fibres. In this case, the silk core was stabilised by annealing in high humidity to promote conformational change of silk into an insoluble phase, before the PEO shell was dissolved in water.

2.4.4 Hollow Fibres

As mentioned previously, hollow fibres have been observed to form due to phase separation in particular conditions, such as the coalescence of polymer-poor regions or when a rapid formation of a skin layer occurs. However, coaxial electrospinning is the technique most used to produce hollow fibres, through selective dissolution of the core material. The production of this kind of fibres is interesting in the field

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of tissue engineering for drug release applications, filtration and for the production of scaffolds which mimic natural tissues such as muscle, nerves and the vascular system [111-113]. Immersion of the mats in a liquid, which is a solvent for the core material and a non-solvent for the sheath, results in the removal of the inner part of fibres, thus producing micro-tubes. Fibres composed of a shell made of PCL and of a PEO core, for example, are transformed into hollow micro-tubes after washing the mats in water [114]. Ou and co-workers [113] used a coaxial electrospinning setup and a rotating mandrel to produce a film composed of super aligned parallel fibres, composed of a polyethylene glycol (PEG)-core and a PLLA-sheath. The dissolution of the core in water leads to the formation of membranes composed of epitaxial-like packed hollow fibres (Figure 2.17).

Figure 2.17 (a) Cross-sectional view of electrospun PLLA hollow-fibres. In the white circle in (b), fibres connected together are shown. Reproduced with

permission from K-L. Ou, J-C. Lu, Y-C. Shu, W-C. Tseng, J-C. Yang, S-Y. Lee and C-C. Chen, European Polymer Journal, 2011, 47, 882. ©2011, Elsevier [113]

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As indicated by the previously mentioned examples, PEG is widely used in this technique because of its water solubility, low cytotoxicity and fast clearance in the human body. Nevertheless, other polymers can be used and removed in different ways, for example heat degradation [38]. Another variation is the use of a non-polymeric core. If the sheath material has sufficient mechanical strength, a simple liquid can form the core, which is eliminated simply by diffusion through the shell. As found in traditional electrospinning, if viscous forces are too low, surface tension leads to the formation of beads instead of fibres. The same phenomenon can occur in the core of coaxial fibres, leading to a shell fibre containing a sequence of beads. This process was described by Diaz and co-workers [38] in the coaxial electrospinning of polyisobutylene as the sheath material and water as a template for the core.

2.4.5 Emulsion Electrospinning

The coaxial electrospinning apparatus is not the only one that can be used to produce fibres with a polymer as the outer sheath containing an inner core of a different material. Emulsion electrospinning is another technique that allows a drug to be embedded in polymeric fibres. Several molecules have been embedded in electrospun fibres, especially proteins [115, 116]. The basic idea is to produce (usually) a water-in-oil emulsion, composed of a polymer dissolved in an organic solvent, as the continuous oil phase, and of a water phase, composed of sub-micrometric drops dispersed in the polymeric solution. The emulsion can be stabilised by the addition of a surfactant. The drug to be released is dissolved in the aqueous phase; this protects it from the organic solvent, which could damage it. The solution is than electrospun, resulting, after the evaporation of the solvent, in polymeric fibres containing the drug. Depending on the process parameters, the stretching of the fibre and the applied electric field can elongate the water phase droplets, leading to coalescence and to the formation of a core fibre structure [117]. As in the coaxial setup, the main advantages are the protection of the drug from organic solvents and the hindering of the burst release. In addition, the apparatus is simpler, needing only one needle, as in traditional electrospinning. However, the presence of surfactants can be a problem due to cytotoxicity.

2.5 Final Remarks

Over the years various electrospinning setups have been developed due to the vast interest in the process as well as the desire to control it and to fabricate different assemblies. Problems related to industrial production and fibre quality of electrospun mats have been addressed in order to increase the process throughput as well as advanced setups for controlled fibre deposition. Further modifications have taken

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place on the target system and on the manipulation of the electric field, which have proven to be successful in the fabrication of several fibrous assemblies.

The application of electrospun fibres in tissue engineering has been constantly rising due to the wide range of structures that the process is able to produce. The influence of micro- and nano-structures on cellular behaviour such as attachment, migration and cytoskeletal organisation has been extensively studied. Several limitations related to the biomedical applications, presented in the process, have been overcome over the years by the modification of electrospinning configuration and by the ingenious approaches that have been developed. Another key issue in tissue engineering, to which electrospun fibres have been applied, is the entrapment/release of bioactive molecules. For this purpose, the production of coaxial or hollow fibres is of great interest since molecules can be entrapped in the core of the fibres and in this way these particular fibres can control the drug release or guide the cell’s adhesion, proliferation and differentiation.

The potential of electrospun fibres are still not completely explored. The latest developments in drug delivery and cell guiding seem to be the short-term future for this manufacturing method. Nevertheless, authors are constantly ingeniously devising new modifications to control single fibre morphologies.

References

1. B. Lu, Y. Wang, Y. Liu, H. Duan, J. Zhou, Z. Zhang, Y. Wang, X. Li, W. Wang, W. Lan and E. Xie, Small, 2010, 6, 15, 1612.

2. D. Fang, B.S. Hsiao and B. Chu, Polymer Preprints, 2003, 44, 2, 59.

3. F-L. Zhou, R-H. Gong and I. Porat, Journal of Applied Polymer Science, 2010, 115, 5, 2591.

4. S.A. Theron, A.L. Yarin, E. Zussman and E. Kroll, Polymer, 2005, 46, 9, 2889.

5. A. Varesano, R.A. Carletto and G. Mazzuchetti, Journal of Materials Processing Technology, 2009, 209, 11, 5178.

6. G. Kim, Y-S. Cho and W.D. Kim, European Polymer Journal, 2006, 42, 9, 2031.

7. Y. Yamashita, F. Ko, H. Miyake and A. Higashiyama, Sen’i Gakkaishi, 2008, 64, 1, 24.

61


8. F-L. Zhou, R-H. Gong and I. Porat, Polymer Engineering & Science, 2009, 49, 12, 2475.

9. A. Kumar, M. Wei, C. Barry, J. Chen and J. Mead, Macromolecular Materials and Engineering, 2010, 295, 8, 701.

10. O.O. Dosunmu, G.G. Chase, W. Kataphinan and D.H. Reneker, Nanotechnology, 2006, 17, 4, 1123.

11. M.R. Badrossamay, H.A. McIlwee, J.A. Goss and K.K. Parker, Nano Letters, 2010, 10, 6, 2257.

12. Y. Srivastava, M. Marquez and T. Thorsen, Journal of Applied Polymer Science, 2007, 106, 5, 3171.

13. O. Jirsak, F. Sanetrnik, D. Lukas, V. Kotek, L. Martinova and J. Chaloupek, inventors; Technicka Universita V Liberci, assignee; US Patent 7585437B2, 2009.

14. D. Lukas, A. Sarkar and P. Pokorny, Journal of Applied Physics, 2008, 103, 8, 084309.

15. A. Yarin and E. Zussman, Polymer, 2004, 45, 9, 2977.

16. N.M. Thoppey, J.R. Bochinski, L.I. Clarke and R.E. Gorga, Polymer, 2010, 51, 21, 4928.

17. S. Tang, Y. Zeng and X. Wang, Polymer Engineering & Science, 2010, 50, 11, 2252.

18. F. Cengiz, T.A. Dao and O. Jirsak, Polymer Engineering & Science, 2010, 50, 5, 936.

19. O. Jirsak, P. Sysel, F. Sanetrnik, J. Hruza and J. Chaloupek, Journal of Nanomaterials, 2010, 2010, 1.

20. X. Wang, H. Niu, T. Lin and X. Wang, Polymer Engineering & Science, 2009, 49, 8, 1582.

21. Y. Liu, J-H. He and J-Y. Yu, Journal of Physics: Conference Series, 2008, 96, 012001.

22. Y. Liu, L. Dong, J. Fan, R. Wang and J-Y. Yu, Journal of Applied Polymer Science, 2011, 120, 1, 592.

62


23. D.R. Salem in Nanofibers and Nanotechnology in Textiles, P.J. Brown and K. Stevens (eds)., CRC Press, Boca Raton, FL, USA, 2007, p.1.

24. A.J. Kelly, Journal of Aerosol Science, 1994, 25, 6, 1159.

25. D. Sun, C. Chang, S. Li and L. Lin, Nano Letters, 2006, 6, 4, 839.

26. C. Chang, K. Limkrailassiri and L. Lin, Applied Physics Letters, 2008, 93, 12, 123111.

27. N. Levit and G. Tepper, The Journal of Supercritical Fluids, 2004, 31, 3, 329.

28. L. Larrondo and R.S.J. Manley, Journal of Polymer Science. Part B. Polymer Physics, 1981, 19, 6, 909.

29. Y. Liu, X. Wang, H. Li, H. Yan and W. Yang, Society of Plastics Engineers Plastics Research Online, 2010, 10.1002/spepro.003055.

30. P.D. Dalton, D. Grafahrend, K. Klinkhammer, D. Klee and M. Möller, Polymer, 2007, 48, 23, 6823.

31. R. Rangkupan and D.H. Reneker, Journal of Metals, Materials and Minerals, 2003, 12, 2, 81.

32. P.D. Dalton, K. Klinkhammer, J. Salber, D. Klee and M. Möller, Biomacromolecules, 2006, 7, 3, 686.

33. J.W. Shin, H.J. Shin, S.J. Heo, Y.J. Lee, Y.M. Hwang, D.H. Kim, J.H. Kim and J.W. Shin in Proceedings of the 3rd Kuala Lumpur International Conference on Biomedical Engineering (IFMBE), Kuala Lumpur, Malaysia, 2006, p.692.

34. W. Bonani, D. Maniglio, A. Motta, W. Tan and C. Migliaresi, Journal of Biomedical Materials Research Part B: Applied Biomaterials, 2011, 96, 2, 276.

35. T. Lin, H. Wang and X. Wang, Advanced Materials, 2005, 17, 22, 2699.

36. P. Gupta and G.L. Wilkes, Polymer, 2003, 44, 20, 6353.

37. H. Jiang, Y. Hu, Y. Li, P. Zhao, K. Zhu and W. Chen, Journal of Controlled Release, 2005, 108, 2-3, 237.

38. J.E. Díaz, A. Fernández-Nieves, A. Barrero, M. Márquez and I.G. Loscertales, Journal of Physics: Conference Series, 2008, 127, 1, 012008.

63


39. J.J. Stankus, J. Guan, K. Fujimoto and W.R. Wagner, Biomaterials, 2006, 27, 5, 735.

40. J.H. Brauker, V.E. Carr-Brendel, L.A. Martinson, J. Crudele, W.D. Johnston and R.C. Johnson, Journal of Biomedical Materials Research, 1995, 29, 12, 1517.

41. F.Z. Volpato, S.L.F. Ramos, A. Motta and C. Migliaresi, Journal of Bioactive and Compatible Polymers, 2011, 26, 1, 35.

42. J. Deitzel, J.D. Kleinmeyer, J.K. Hirvonen and N.C. Beck Tan, Polymer, 2001, 42, 19, 8163.

43. D. Li, Y. Wang and Y. Xia, Nano Letters, 2003, 3, 8, 1167.

44. Z-M. Huang, Y-Z. Zhang, M. Kotaki and S. Ramakrishna, Composites Science and Technology, 2003, 63, 15, 2223.

45. W.E. Teo, M. Kotaki, X.M. Mo and S. Ramakrishna, Nanotechnology, 2005, 16, 6, 918.

46. A. Theron, E. Zussman and A.L. Yarin, Nanotechnology, 2001, 12, 3, 384.

47. D. Li, Y. Wang and Y. Xia, Advanced Materials, 2004, 16, 4, 361.

48. P. Katta, M. Alessandro, R.D. Ramsier and G.G. Chase, Nano Letters, 2004, 4, 11, 2215.

49. J.K. Hong and S.V. Madihally, Tissue Engineering, 2011, 17, 2, 125.

50. A. Thorvaldsson, J. Engström, P. Gatenholm and P. Walkenström, Journal of Applied Polymer Science, 2010, 118, 1, 511.

51. A. Thorvaldsson, H. Stenhamre, P. Gatenholm and P. Walkenström, Biomacromolecules, 2008, 9, 3, 1044.

52. E. Smit, U. Büttner and R.D. Sanderson, Polymer, 2005, 46, 8, 2419.

53. M-S. Khil, S.R. Bhattarai, H-Y. Kim, S-Z. Kim and K-H. Lee, Journal of Biomedical Materials Research. Part B, Applied Biomaterials, 2005, 72, 1, 117.

54. W.E. Teo and S. Ramakrishna, Nanotechnology, 2006, 17, 14, R89.

64


55. M. Simonet, O.D. Schneider, P. Neuenschwander and W.J. Stark, Polymer Engineering & Science Engineering, 2007, 47, 12, 2020.

56. M.F. Leong, M.Z. Rasheed, T.C. Lim and K.S. Chian, Journal of Biomedical Materials Research Part A, 2009, 91, 1, 231.

57. O.D. Schneider, F. Weber, T.J. Brunner, S. Loher, M. Ehrbar, P.R. Schmidlin and W.J. Stark, Acta Biomaterialia, 2009, 5, 5, 1775.

58. Y.S. Nam, J.J. Yoon and T.G. Park, Journal of Biomedical Materials Research, 2000, 53, 1, 1.

59. Y.H. Lee, J.H. Lee, I-G. An, C. Kim, D.S. Lee, Y.K. Lee and J-D. Nam, Biomaterials, 2005, 26, 16, 3165.

60. Y. Wang, E. Bella, C.S.D. Lee, C. Migliaresi, L. Pelcastre, Z. Schwartz, B.D. Boyan and A. Motta, Biomaterials, 2010, 31, 17, 4672.

61. Q. Hou, D.W. Grijpma and J. Feijen, Biomaterials, 2003, 24, 11, 1937.

62. J. Nam, Y. Huang, S. Agarwal and J. Lannutti, Tissue Engineering, 2007, 13, 9, 2249.

63. T.G. Kim, H.J. Chung and T.G. Park, Acta Biomaterialia, 2008, 4, 6, 1611.

64. B.M. Baker, A.O. Gee, R.B. Metter, A.S. Nathan, R.A. Marklein, J.A. Burdick and R.L. Mauck, Biomaterials, 2008, 29, 15, 2348.

65. B.M. Whited, J.R. Whitney, M.C. Hofmann, Y. Xu and M.N. Rylander, Biomaterials, 2011, 32, 9, 2294.

66. R. Gentsch, B. Boysen, A. Lankenau, and H. G. Börner, Macromolecular Rapid Communications, 2010, 31, 1, 59.

67. S. Soliman, S. Pagliari, A. Rinaldi, G. Forte, R. Fiaccavento, F. Pagliari, O. Franzese, M. Minieri, P. Di Nardo, S. Licoccia and E. Traversa, Acta Biomaterialia, 2010, 6, 4, 1227.

68. M.I. Santos, K. Tuzlakoglu, S. Fuchs, M.E. Gomes, K. Peters, R.E. Unger, E. Piskin, R.L. Reis and C.J. Kirkpatrick, Biomaterials, 2008, 29, 32, 4306.

69. K. Tuzlakoglu, N. Bolgen, A.J. Salgado, M.E. Gomes, E. Piskin and R.L. Reis, Journal of Materials Science: Materials in Medicine, 2005, 16, 12, 1099.

65


70. F. Rosso, A. Giordano, M. Barbarisi and A. Barbarisi, Journal of Cellular Physiology, 2004, 199, 2, 174.

71. G. Kim, J. Son, S. Park and W. Kim, Macromolecular Rapid Communications, 2008, 29, 19, 1577.

72. S.H. Ahn, Y.H. Koh and G.H. Kim, Journal of Micromechanics and Microengineering, 2010, 20, 6, 065015.

73. H. Yoon and G. Kim, Journal of Pharmaceutical Sciences, 2011, 100, 2, 424.

74. P. Tayalia, C.R. Mendonca, T. Baldacchini, D.J. Mooney and E. Mazur, Advanced Materials, 2008, 20, 23, 4494.

75. E. Sachlos and J.T. Czernuszka, European Cells & Materials, 2003, 5, 29.

76. C. Mota, D. Puppi, D. Dinucci, C. Errico, P. Bártolo and F. Chiellini, Materials, 2011, 4, 3, 527.

77. L. Wang, C.-L. Pai, M.C. Boyce and G.C. Rutledge, Applied Physics Letters, 2009, 94, 15, 151916.

78. C.L. Casper, J.S. Stephens, N.G. Tassi, D.B. Chase and J.F. Rabolt, Macromolecules, 2004, 37, 2, 573.

79. A.S.G. Curtis, N. Gadegaard, M.J. Dalby, M.O. Riehle, C.D.W. Wilkinson and G. Aitchison, IEEE Transactions in Nanobioscience, 2004, 3, 1, 61.

80. B.D. Boyan, S. Lossdörfer, L. Wang, G. Zhao, C.H. Lohmann, D.L. Cochran and Z. Schwartz, European Cells & Materials, 2003, 6, 22.

81. T-W. Chung, D-Z. Liu, S-Y. Wang and S-S. Wang, Biomaterials, 2003, 24, 25, 4655.

82. L. Moroni, R. Licht, J. de Boer, J.R. de Wijn and C.A. van Blitterswijk, Biomaterials, 2006, 27, 28, 4911.

83. Y.B. Truong, V. Glattauer, G. Lang, K. Hands, I.L. Kyratzis, J.A. Werkmeister and J.A.M. Ramshaw, Biomedical Materials, 2010, 5, 2, 25005.

84. K.Y. Lee and S.H. Yuk, Progress in Polymer Science, 2007, 32, 7, 669.

85. S.L. Tao and T.A. Desai, Advanced Drug Delivery Reviews, 2003, 55, 3, 315.

66


86. L. Moroni, J.R. de Wijn and C.A. van Blitterswijk, Journal of Biomaterials Science. Polymer Edition, 2008, 19, 5, 543.

87. V. Luginbuehl, L. Meinel, H.P. Merkle and B. Gander, European Journal of Pharmaceutics and Biopharmaceutics, 2004, 58, 2, 197.

88. S.M. Willerth and S.E. Sakiyama-Elbert, Advanced Drug Delivery Reviews, 2007, 59, 4-5, 325.

89. M. Biondi, F. Ungaro, F. Quaglia and P.A. Netti, Advanced Drug Delivery Reviews, 2008, 60, 2, 229.

90. M.M. Munir, A.B. Suryamas, F. Iskandar and K. Okuyama, Polymer, 2009, 50, 20, 4935.

91. Y. Wu and R.L. Clark, Journal of Colloid and Interface Science, 2007, 310, 2, 529.

92. A. Jaworek and A.T. Sobczyk, Journal of Electrostatics, 2008, 66, 3-4, 197.

93. S. Koombhongse, W. Liu and D.H. Reneker, Journal of Polymer Science Part B: Polymer Physics, 2001, 39, 21, 2598.

94. Q.Z. Yu, Y. Li, M. Wang and H.Z. Chen, Chinese Chemical Letters, 2008, 19, 2, 223.

95. C.J. Luo, M. Nangrejo and M. Edirisinghe, Polymer, 2010, 51, 7, 1654.

96. J. Zheng, A. He, J. Li, J. Xu and C.C. Han, Polymer, 2006, 47, 20, 7095.

97. M.F. Leong, K.S. Chian, P.S. Mhaisalkar, W.F. Ong and B.D. Ratner, Journal of Biomedical Materials Research. Part A, 2009, 89, 4, 1040.

98. E. Servoli, G.A. Ruffo and C. Migliaresi, Polymer, 2010, 51, 11, 2337.

99. P. Dayal and T. Kyu, Journal of Applied Physics, 2006, 100, 4, 043512.

100. S. Megelski, J.S. Stephens, D.B. Chase and J.F. Rabolt, Macromolecules, 2002, 35, 22, 8456.

101. Z. Qi, H. Yu, Y. Chen and M. Zhu, Materials Letters, 2009, 63, 3-4, 415.

102. G. Luna-Bárcenas, S.K. Kanakia, I.C. Sanchez and K.P. Johnston, Polymer, 1995, 36, 16, 3173.

67


103. J. Liu, Z. Shen, S-H. Lee, M. Marquez and M.A. McHugh, The Journal of Supercritical Fluids, 2010, 53, 1-3, 142.

104. J.T. McCann, M. Marquez and Y. Xia, Journal of the American Chemical Society, 2006, 128, 5, 1436.

105. Y. You, J.H. Youk, S.W. Lee, B-M. Min, S.J. Lee and W.H. Park, Materials Letters, 2006, 60, 6, 757.

106. W.S. Lyoo, J.H. Youk, S.W. Lee and W.H. Park, Materials Letters, 2005, 59, 28, 3558.

107. S. Chakraborty, I-C. Liao, A. Adler and K.W. Leong, Advanced Drug Delivery Reviews, 2009, 61, 12, 1043.

108. R. Srikar, A.L. Yarin, C.M. Megaridis, A.V. Bazilevsky and E. Kelley, Langmuir, 2008, 24, 3, 965.

109. M. Gandhi, R. Srikar, A.L. Yarin, C.M. Megaridis and R.A. Gemeinhart, Molecular Pharmaceutics, 2009, 6, 2, 641.

110. M. Wang, J.H. Yu, D.L. Kaplan and G.C. Rutledge, Macromolecules, 2006, 39, 3, 1102.

111. Y. Srivastava, I. Loscertales, M. Marquez and T. Thorsen, Microfluidics and Nanofluidics, 2008, 4, 3, 245.

112. L. Moroni, R. Schotel, J. Sohier, J.R. de Wijn and C.A. van Blitterswijk, Biomaterials, 2006, 27, 35, 5918.

113. K-L. Ou, J-C. Lu, C-S. Chen, L-H. Lin, Y-C. Shu, W-C. Tseng, J-C. Yang, S-Y. Lee and C-C. Chen, European Polymer Journal, 2011, 47, 5, 882.

114. Y. Dror, W. Salalha, R. Avrahami, E. Zussman, A.L. Yarin, R. Dersch, A. Greiner and J.H. Wendorff, Small, 2007, 3, 6, 1064.

115. X. Li, Y. Su, X. Zhou and X. Mo, Colloids and Surfaces B: Biointerfaces, 2009, 69, 2, 221.

116. Y. Yang, X. Li, M. Qi, S. Zhou and J. Weng, European Journal of Pharmaceutics and Biopharmaceutics, 2008, 69, 1, 106.

117. X. Li, Y. Su, S. Liu, L. Tan, X. Mo and S. Ramakrishna, Colloids and Surfaces B: Biointerfaces, 2010, 75, 2, 418.

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