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Poly(l -lactide) and poly(butylene succinate) immiscible blends: From electrospinning to biologically active materials
Materials Science and Engineering C 41 (2014) 119–126
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Materials Science and Engineering C journal homepage: www.elsevier.com/locate/msec
Poly(L-lactide) and poly(butylene succinate) immiscible blends: From electrospinning to biologically active materials Nikoleta Stoyanova a, Dilyana Paneva a, Rosica Mincheva b, Antoniya Toncheva a, Nevena Manolova a, Philippe Dubois b, Iliya Rashkov a,⁎ a b
Laboratory of Bioactive Polymers, Institute of Polymers, Bulgarian Academy of Sciences, Acad. G. Bonchev St, bl. 103A, BG-1113 Sofia, Bulgaria Laboratory of Polymeric and Composite Materials, Center of Innovation and Research in Materials and Polymers (CIRMAP), University of Mons — UMONS, Place du Parc 20, B-7000 Mons, Belgium
a r t i c l e
i n f o
Article history: Received 24 October 2013 Received in revised form 4 April 2014 Accepted 18 April 2014 Available online 28 April 2014 Keywords: Electrospinning Poly(butylene succinate) Poly(L-lactide) Crystallinity Mechanical properties Biological activity
a b s t r a c t For the first time the preparation of defect-free fibers from immiscible blends of high molar mass poly(lactic acid) (PLA) and poly(butylene succinate) (PBS) in the whole range of the polyester weight ratios is shown. Electrospinning using the solvent–nonsolvent approach proved most appropriate. Moreover, electrospinning revealed crucial for the obtaining of PLA/PBS materials maintaining integrity. DSC and XRD analyses attested for a plasticizing effect and for increased PLA crystallinity at PBS addition to PLA. The mechanical properties of the PLA/ PBS mats were controlled by the alignment of the fibers and changed from plastic to brittle materials upon increasing the PBS content. Drug loading and tests against pathogenic microorganisms suggested that the obtained mats can find application as antibacterial fibrous materials. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Materials from biodegradable and non-toxic biocompatible polymers which are synthesized entirely or partially from annually renewable resources have drawn great interest [1]. Their use is imposed by the need for replacement of the conventional materials derived from petroleum sources as well as in the viewpoint of environmental pollution prevention. Among all, the biocompatible and thermoplastic poly(lactide) PLA is one of the most promising biobased polymers [2], finding applications going from drug-carriers and implants to packaging and textiles. More recently, PLA-based materials have found more durable applications in automotive, communication and electronic industries [3,4]. However, drawbacks such as mechanical brittleness and low crystallization rate are often obstacles to PLA practical uses. With this respect, PBS — a partially biobased polyester, with thermal and mechanical properties comparable with those of low density polyethylene and polypropylene [5] was studied as a PLA modifier [6–14]. Because of its high flexibility (low glass-transition temperature), PBS has found applications as films, laminates, molded foams and injection-molded products in various fields, such as agriculture and packaging. The PBS relatively rapid crystallization rate and flexibility were believed to ⁎ Corresponding author. Tel./fax: +359 2 9793468/+359 2 8700309. E-mail address: [email protected] (I. Rashkov).
http://dx.doi.org/10.1016/j.msec.2014.04.043 0928-4931/© 2014 Elsevier B.V. All rights reserved.
improve PLA thermal and mechanical properties when blended and materials from PLA/PBS blends have been prepared by extrusion [6–12]. The miscibility, the phase and crystallization behavior, and the morphology of PLA/PBS blends were reported in the literature [6,15]. Interestingly, despite the improved crystallization of PLA in the presence of PBS, thermal analyses showed two distinct melting peaks over the entire composition range, thus classifying the materials as semicrystalline/semicrystalline blends. Moreover, small-angle X-ray scattering revealed that, in a blend system, the PBS component was expelled out of the interlamellar regions of PLA. Finally, for more than 40 wt.% PBS, significant crystallization-induced phase separation was observed. The thus suggested immiscibility of both polymers [15] is limiting the applications of PLA/PBS blend materials without compatibilization. A possibility to overcome the problem might be found in the electrospinning technique. Electrospinning is a cutting-edge technique that allows micro- and nanofibrous polymer materials to be easily prepared [16–22]. Because of their high surface-to-volume ratio and porous structure, the fibrous mats are well suited for applications such as filters, scaffolds for cell and tissue engineering, and carriers of bioactive substances. Electrospinning of immiscible polymer blends containing PLA is already reported in the literature [23]. The concept resulted in freezing the phase structure generated during electrospinning, thus preserving the co-continuous or interpenetrating phase morphologies within the
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fibers. Noteworthy, unlike PLA that can be easily electrospun from solutions of diverse in nature solvent systems based on chloroform or dichloromethane [24], the electrospinning of PBS solutions encounters certain difficulties and reports on the electrospinning of PBS are still scarce. The preparation of fibrous materials from PBS with a number average molar mass of ca. 75,000 g/mol by electrospinning has been reported [25]. Mixed solvent systems such as chloroform/2-chloroethanol at a ratio of 7/3 or 6/4 (w/w), dichloromethane/2-chloroethanol at a ratio of 7/3 or 6/4 (w/w) and chloroform/3-chloro-1-propanol at a ratio of 9/1 (w/w) have been used. It proved preferable to avoid the use of high boiling solvents such as 2-chloroethanol and 3-chloro-1-propanol when the polymer materials are intended to find application in the biomedical domain or simply to come into contact with human body. A short communication presenting few SEM micrographs of porous fibers prepared by electrospinning of PBS with a molar mass of 20,000 g/mol using chloroform as solvent was published in 2008 [26]. It has been found that the electrospinning of PBS with a molar mass as high as 200,000– 300,000 g/mol using chloroform as solvent is very difficult to achieve when the polymer concentration is lower than 10 wt.% or higher than 15 wt.% [27]. By using PBS with a weight average molar mass MW 3 × 105 g/mol and a chloroform/ethanol solvent system at a ratio of 75/ 25 (v/v) and polymer concentration of 11% (w/v) PBS/wollastonite mats have been electrospun [28]. DCM/trifluoroacetic acid (90/10 v/v) solvent system has been used for electrospinning of 1,6-diisocyanatohexaneextended PBS [29]. Despite the up-to-here described interest, and most probably because of the significant difficulties in performing the electrospinning, there are no literature data on the elaboration of fibrous materials from these two immiscible commercially available high molar mass PLA and PBS polymers through this method. Therefore, the present study aims at preparing defect-free fibers from immiscible blends of PLA and PBS from 100/0 to 10/90 (w/w). The role of the solvent system and the total polymer concentration on fiber morphology was studied. DSC and XRD were used to evaluate crystallinity and crystallization rate of the polyesters in the fibrous mats, and tensile tests were used to determine their mechanical properties. Drug loading and tests with pathogenic microorganisms were also performed as possible material application.
The dynamic viscosity of the spinning solutions was measured by Brookfield LVT viscometer equipped with an adaptor for small samples, a spindle, and a camera SC 4-18/13 R at 20 ± 0.1 °C; the measurements were performed in triplicate and the mean values with their standard deviations were used. For the incorporation of K5N8Q and 5Cl8HQ spinning solutions of PLA/PBS = 80/20 (w/w) in DCM/EtOH = 90/10 (w/w) at total polymer concentration of 10 wt.% were used; the concentration of K5N8Q and 5Cl8HQ was 10 or 20 wt.% in respect to the polymer weight, respectively. In the case of K5N8Q due to its poor solubility, the electrospinning was performed from a stable dispersion. The electrospinning was carried out using an electrospinning set-up consisting of a high voltage power supply (up to 30 kV); a pump [NE300 Just Infusion™ Syringe Pump (New Era Pump Systems Inc., USA)] for delivering the spinning solution at a constant rate; a syringe provided with a positively charged metal needle with inner diameter of 0.6 mm and a custom-made grounded rotating aluminum collector. The electrospinning was performed under the following conditions: feeding rate: 3 mL/h, applied voltage: 25 kV, tip-to-collector distance: 17 cm, collector rotating speed: 600 or 1900 rpm, humidity: 70% and temperature: 20 °C. The obtained mats were dried additionally under reduced pressure (320 Pa) at r.t. for 8 h. 2.3. Characterization of the electrospun materials The morphology of the fibers was examined by a scanning electron microscope (SEM). The samples were vacuum-coated with gold and observed by a Jeol JSM-5510 SEM. The fiber morphology was evaluated by applying the criteria for complex evaluation of electrospun materials as described elsewhere [30] using the ImageJ software by measuring the diameters of at least 20 fibers from each SEM micrograph. The thermal behavior of the prepared new fibrous materials was followed by differential scanning calorimetry (DSC). The analyses were performed using a PerkinElmer DSC 8500 differential scanning calorimeter in nitrogen atmosphere at a heating rate of 10 °С/min. PLA PBS (χPLA c , %) and PBS (χc , %) crystallinity degree was calculated using the following equations: h i PLA PLA PLA PLA;0 = ΔHm 100 ΔH m −ΔH cc W
χc
PLA
2.1. Reagents
χc
PBS
Poly(butylene succinate) (PBS, Bionolle 1003; Shōwa Denkō Kabushiki-gaisha — Japan; MW = 97,600 g/mol, MW /Mn = 2.56 as determined by size-exclusion chromatography using polystyrene standards); poly(L-lactide) (PLA, Ingeo™ Biopolymer 4032D, NatureWorks LLC — USA; MW = 259,000 g/mol; MW /Mn = 1.94; as determined by size-exclusion chromatography using polystyrene standards), dichloromethane (DCM, Merck); ethanol (abs. EtOH, Merck); the potassium salt of 5-nitro-8-hydroxyquinoline (K5N8Q) and 5-chloro-8-quinolinol (5chloro-8-quinolinol; 95% 5Cl8HQ; Sigma-Aldrich) were of analytical grade of purity.
where WPLA is the PLA weight fraction, and WPBS — the PBS weight fraction in the spinning solution; ΔH0m is the heat of fusion of 100% polymer crystal: ΔHPLA,0 = 93.0 J/g [31]; ΔHPBS,0 = 210.0 J/g [32]. The measurem m ments were performed in triplicate and the mean values with their standard deviations were used for interpreting the obtained results. The absence/presence of a crystalline phase in the fibrous materials was assessed by X-ray diffraction analysis (XRD). XRD spectra were recorded at r.t. using a computer controlled D8 Bruker Advance powder diffractometer with a filtered CuKα radiation source and a luminescent detector. The analyses were performed in the 2θ range from 5° to 50° with a step of 0.02° and counting time of 1 s/step. Mechanical properties were evaluated by tensile measurements performed on the fibrous scaffolds using a Zwick/Roell Z 2.5 apparatus, load cell 2 mV/V, type Xforce P, Nominal Force 2.5 kN and test Xpert II. The specimens' thickness was around 50 μm. The stretching rate was 10 mm/min, and the initial length between the clamps was 60 mm. The fibrous materials were tested along the circumferential direction of the collector on which the mat was deposited, and the mean values were calculated. The water contact angle of the mats was determined using Easy Drop DSA20E KRÜSS GmbH device at r.t. A drop of deionized water (10 μL) was deposited on the mats surface. From the images of the
;% ¼
ð1Þ
2. Experimental
2.2. Preparation of PLA/PBS fibrous materials by electrospinning Spinning solutions in DCM or DCM/EtOH [DCM/EtOH = 90/10 or 80/20 (w/w)] were prepared for PLA/PBS blends of PLA/PBS = 100/0; 90/10, 80/20, 70/30, 60/40, 50/50, 40/60, 30/70, 20/80, 10/ 90 and 0/100 (w/w). For preparation of the solutions, PLA and PBS were first dissolved separately in DCM; for the dissolution of PBS heating at 40 °C for 2 h was necessary. Then, the solutions were mixed at stirring. The total polyester concentration was 10 or 15 wt.%. In the case of the DCM/EtOH solvent systems, ethanol was added after the mixing of PLA and PBS solutions in DCM.
h i PBS PBS PBS;0 100 ; % ¼ ΔH m = ΔHm W
ð2Þ
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PLA
PLA/PBS = 90/10 (w/w)
PLA/PBS = 20/80 (w/w)
121
PLA/PBS = 80/20 (w/w)
PLA/PBS = 10/90 (w/w)
PBS
Fig. 1. Representative SEM images of PLA/PBS fibers prepared at PLA/PBS from 100/0 to 0/100 (w/w) at total polymer concentration of 10 wt.% and by using a mixed solvent system DCM/ EtOH; collector rotating speed: 600 rpm; magnification: ×1000.
droplets on the surface of the mats acquired by a digital camera and processed by a software program the average values of the water contact angles were determined based on at least 10 measurements. The samples for the water contact angle measurements were cut in the collector rotation direction and collector axis direction. 2.3.1. Microbiological tests The antibacterial activity of PLA/PBS = 80/20 (w/w) mats containing 10 wt.% K5N8Q or 20 wt.% 5Cl8HQ was monitored against the pathogenic microorganism Staphylococcus aureus (ATCC 6538 P). In order to measure the zones of inhibition, in vitro studies were performed using Tryptone glucose extract agar (DIFCO Laboratories, Detroit, USA) solid medium. The surface of the solid agar was inoculated with a suspension of S. aureus bacterial culture with a cell concentration of 1 × 105 cells/mL and on the surface of the agar in each Petri dish two mats (disks with diameters of 10 mm and weight 1.0 mg) from drug-(non)containing PLA/PBS = 80/20 (w/w) were placed. The Petri dishes were incubated for 24 h at 37 °C and subsequently the zones of inhibition around the disks were measured. The average diameters of the zones of inhibition were determined using the ImageJ software based on 15 measurements in 15 different directions for each zone.
A
3. Results and discussion 3.1. Electrospinning and fiber morphology Attempts to obtain uniform PLA/PBS fibers from both immiscible polymers were performed by electrospinning their blend solutions at 30 kV and at collector rotating speed of 600 rpm. DCM — a common solvent for both polyesters was first studied at total polymer concentration of 10 wt.% (Fig. S1). Thus, cylindrical defect-free fibers were obtained up to PLA/PBS = 70/30 (w/w). Further increase in PBS content resulted in spindle-like defects along the fibers (to 50 wt.% PBS) or beaded fibers (PBS content from 80 wt.% to 100 wt.%). Further experiments were performed at a total polymer concentration of 15 wt.% (DCM, 30 kV, 600 rpm) allowing the obtaining of defect-free fibers at 50 wt.% PBS (mean diameter of 7.3 ± 2.1 μm) (Fig. S2). However, the electrospinning process was ineffective (and even impossible at higher PBS content) due to rapid phase separation and gelation of the spinning solution. For overcoming these problems, the solvent–nonsolvent approach was attempted. It is known that adding a certain amount of a poor solvent to the spinning solution assists in the formation of fibers during electrospinning [33,34]. Thus, in the present study DCM was partially
B 500
3000
Dynamic viscosity, cP
Mean fiber diameter value, nm
3500
2500 2000 1500 1000
400
300
200
100
500 0 100/0
80/20
60/40
40/60
PLA/PBS, w/w
20/80
0/100
0 100/0 80/20 60/40 40/60 20/80 0/100
PLA/PBS, w/w
Fig. 2. Mean fiber diameter (A), and dynamic viscosity (B) for different PLA/PBS weight ratios; total polymer concentration: 10 wt.%; collector rotating speed: 600 rpm.
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Fig. 3. DSC thermograms (first heating run, A) and XRD patterns (B) at selected PLA/PBS weight ratios. DCM/EtOH; collector rotating speed: 600 rpm.
Table 1 Thermal characteristics of PLA/PBS mats as obtained from DCM/EtOH at 600 rpm. PLA/PBS
Tg PLA °C
Tcc PLA °C
ΔH cc PLA J/g⁎
Tm PLA °C
ΔH m PLA J/g⁎
χPLA %⁎⁎
Tcc PBS °C
Tm PBS °C
ΔHm PBS J/g⁎
χPBS %⁎⁎
100/0 90/10 80/20 70/30 60/40 50/50 40/60 30/70 20/80 10/90 0/100
67 58 51 53 54 52 52 52 48 48 –
90 78 70 – – – – – – – –
21.2 15.3 18.1 – – – – – – – –
167 168 166 168 168 166 168 168 167 168 –
31.3 33.6 34.4 26.3 24.2 19.2 20.4 9.2 16.2 6.5 –
10.9 21.9 21.9 22.7 25.1 29.5 35.0 30.4 50.0 70.0 –
– – – 91 89 88 89 88 88 – –
– – 114 114 114 110 114 114 112 114 116
– – 6.0 16.8 27.2 35.6 38.6 52.5 52.0 67.9 77.0
– – 14.3 26.7 32.4 29.8 30.6 35.7 31.0 36.0 36.0
⁎ The enthalpies are not recalculated taking into account the respective polyester content. ⁎⁎ Crystallinity degrees are calculated as described in the Exp. Part. ΔHccPBS is not presented since the cold crystallization overlapped PBS melting.
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fabricated using DCM/EtOH were in the range 1800–600 nm depending on the PBS content (Fig. 2A). As seen, d continuously decreased upon increasing PBS content and the thinnest defect-free fibers were obtained at 90 wt.% PBS (590 ± 140 nm). This effect might be related to the changes in the dynamic viscosity of the spinning solutions (Fig. 2B). Indeed, the dynamic viscosity decreased upon increasing PBS content — the polyester of lower molar mass (MW = 97,600). 3.2. Crystallinity and mechanical properties of the fibrous mats
Fig. 4. Stress–strain curves recorded during mechanical tests of PLA/PBS mats prepared at a collector rotating speed of 600 (A) or 1900 (B) rpm; solvent system: DCM/EtOH; the samples were cut in such a manner so that their length axis coincided with the direction of the collector rotation.
replaced by EtOH — a poor solvent for both PBS and PLA, while the total polymer concentration, the voltage and the collector rotating speed were kept constant (10 wt.%, 30 kV and 600 rpm, respectively). It was found that the addition of only 10 wt.% EtOH to DCM significantly improves the electrospinning of PLA/PBS = 50/50 (w/w) solutions. In contrast to DCM, the fibers obtained from DCM/EtOH = 90/10 (w/w) were cylindrical and defect-free. Their mean diameter was 1.1 ± 0.4 μm. Further increase of EtOH content in the spinning solution [DCM/EtOH = 80/20 (w/w)] was counterproductive as spindle-like defects appeared along the fiber axis (Fig. S2). Therefore the preparation of defect-free fibrous materials from PLA/PBS at the whole range of weight ratios was studied in the system DCM/EtOH = 90/10 (w/w) at total polymer concentration of 10 wt.% (further denoted as DCM/EtOH). SEM micrographs of the obtained fibers are presented in Fig. 1. As seen, the use of DCM/EtOH (30 kV, 600 rpm) allowed the obtaining of randomly deposited defect-free fibers from the immiscible PLA and PBS up to 80 wt.% PBS. Even at PLA/PBS = 10/90 (w/w) only few spindle-like defects appeared and the mats were predominantly composed of defect-free fibers. For neat PBS the obtained fibrous material consisted of fibers (mean diameter: 170 ± 48 nm) and bead-like defects (mean diameter: 7.8 ± 2 μm). This pattern is quite different as compared to DCM solutions, thus confirming the pertinence of the solvent–nonsolvent approach. The mean diameters of PLA/PBS fibers (d)
The as prepared fibrous mats were further characterized in terms of crystallinity and mechanical properties. Representative DSC thermograms of PLA/PBS mats depending on the PBS content are shown in Fig. 3A. Concerning the PLA component, Tg, Tcc and Tm were detected at low PBS content. This occurrence of cold crystallization is in agreement with literature data on electrospun PLA materials and might be explained by the rapid solvent evaporation hampering the polyester crystallization [35–40]. Moreover, Tg and Tcc decreased with ca. 20 °C upon increasing PBS content to 20 wt.% (Table 1). In parallel, the enthalpy of cold crystallization decreased with ca. 15%, thus suggesting some miscibility between the polyesters in the amorphous state at low PBS content. These data are in agreement with the literature on PLA/PBS materials prepared by extrusion [6,8,17], evidencing the role of PBS as plasticizer for PLA. Further increase in PBS content did not influence the PLA Tg, but led to the disappearance of Tcc, suggesting that blending with PBS most probably influenced the PLA crystallization and crystallization rate. In agreement, the melting endotherm of PLA turned bimodal and the corresponding crystallinity increased. The second Tm newly occurring at higher temperatures (max at 176 °C for PLA/PBS = 20/80, w/w) might be ascribed to the formation of crystals which are organized to a greater extent, as observed for PLA/PBS extruded blends [8]. It is to be noted that the PBS Tg was difficult to observe due to the high crystallinity and crystallization rate of the polyester. Therefore, avoiding any speculation on electrospinning induced miscibility of the PLA/PBS blends based on Tg determination is preferable. Regarding PBS, no significant difference in thermal behavior was detected between the neat and the blended fibrous mats. PBS is rapidly crystallizing polyester of high crystallinity [32], making its Tg difficult to assess. Therefore, no values were collected from the DSC analyses. The polyester melting appeared at ca. 116 °C and was directly preceded by a cold crystallization exotherm in agreement with the literature data [32]. Interestingly, Tm was not influenced by the mat composition and the corresponding enthalpy of melting increased upon increasing PBS content. The obtained results were confirmed by the performed XRD analyses. XRD data on films of PLA and PBS homopolyesters are available in the literature [41,42]. According to the literature, the PLA pattern presents diffraction peaks at 2θ = 14.8°, 16.6°, 19.0°, 22.4° and 29.2° distinctive for the α-form of optically pure PLLA (or PDLA) crystallizing in a pseudo-orthorhombic unit cell containing l03 helices [42], while PBS presents diffraction peaks at 2θ = 19.6°, 21.9°, 22.8° and 29.1° ascribed to its α-form (monoclinic system of 21 helices) [41]. As seen from Fig. 3B, the XRD patterns of PLA/PBS = 100/0 and 80/20 (w/w) mats revealed only the presence of an amorphous halo. Diffractions for crystal phase were not observed, thus confirming the hindered crystallization of the polyesters during the electrospinning process as evidenced by DSC. In the XRD pattern of PLA/PBS = 50/50 (w/w) mat a diffraction at 2θ = 22.6° was registered, and in the XRD pattern of PLA/PBS = 20/80 (w/w) mat: two diffractions were observed at 2θ = 19.6° and 22.6°, respectively. These values of 2θ are characteristic for both PLA and PBS crystal structure, and their attribution cannot be done only on the basis of the XRD data. However and taking into consideration their crystallinity degree calculated by DSC (Table 1), it can be assumed that the registered diffractions were due to the crystal structure of both PLA and PBS.
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A
B 12.5
600 rpm 1900 rpm
Tensile strength, MPa
Young's Modulus, MPa
500 400 300 200 100
80/20
50/50
7.5 5.0
0.0 100/0
20/80
*
*
50/50
20/80
2.5
80/20
PLA/PBS, w/w
PLA/PBS, w/w
C
D 600 rpm
125 100
* 75
*
50
*
25 0
100/0
80/20
60
1900 rpm
*
50/50
20/80
Crystallinity degree, %
150
Elongation, %
1900 rpm * *
10.0
0 100/0
600 rpm
PLA/600 rpm PLA/1900 rpm PBS/600 rpm PBS/1900 rpm
50 40 30 20 10 0
100/0
PBS content, wt.%
80/20
50/50
20/80
PLA/PBS, w/w
Fig. 5. Dependence of Young's modulus (A), tensile strength (B) and elongation at break (C) of PLA/PBS mats, and the crystallinity degree of PLA and PBS (D) in the fibrous materials on PBS content in the mats, prepared at collector rotation speed of 600 or 1900 rpm; solvent system: DCM/EtOH. The samples for which a break during the tests was detected are marked with an asterisk; thus, the obtained values may be considered as elongation at break.
It is worth to be mentioned that solvent-cast films from PBS and PLA/PBS lost their integrity and became very brittle on drying. In sharp contrast, PLA/PBS fibrous mats maintained integrity and could be easily handled without fragmentation. In order to determine their mechanical properties tensile tests on mats of defect-free fibers were performed. The stress–strain curves of PLA/PBS fibrous materials at different PBS contents are presented in Fig. F4A. Digital images of the samples after testing are shown as well. As seen, before reaching the ultimate tensile strength, the PLA/PBS = 100/0 (w/w) and PLA/PBS = 80/20 (w/w) mats displayed typical strain behavior of thermoplastic materials. At low stress values (in the elastoplasticity range) they rapidly reached the yield point after which necking
was observed. This necking after the yield point gave evidence for the occurrence of inelastic deformation. The subsequent increase of the stress led to hardening of the samples most probably due to the additional orientation of the fibers during specimens' elongation. Partial tearing of the samples was observed after reaching the maximum tensile stress. Further increase in PBS content (PLA/PBS = 50/50 (w/w) and PLA/PBS = 20/80 (w/w)) resulted in distinct break point, and the absence of necking, indicating quasi-ductile fracture behavior of these materials. One possible explanation of this finding might be the alignment of the fibers during stretching. However, based on the DSC and XRD data alteration in the blend morphology: a transition from dispersed structure to bicontinuous one and again to dispersed structure; alteration in the crystallinity of the blends and by interfacial adhesion [43] should not be neglected. Effects of
A 1985±400 nm
C 760±310 nm
B 953±475 nm
Fig. 6. SEM micrographs of PLA/PBS fibers prepared at total polymer concentration of 10 wt.% using DCM/EtOH in absence (A) and presence of the drug: 10 wt.% K5N8Q (B) or 20 wt.% 5Cl8HQ (C) (in respect to the total polymer weight); collector rotating speed: 1900 rpm; the collector rotation direction is indicated by a white arrow.
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S. aureus
PLA/PBS
PLA/PBS/K5N8Q
125
PLA/PBS/5Cl8HQ
Fig. 7. Digital images of the zones of inhibition against S. aureus, detected after a 24-h contact of the drug-(non)containing PLA/PBS fibrous materials with the bacterial cells. Left image — blank control.
additional orientation of the macromolecules during specimen elongation [44] are not to be excluded. To verify the hypothesis, fibrous materials were obtained at higher collector rotation speed (1900 rpm), known to increase the degree of fibers' alignment. According to theoretical simulations confirmed by experimental measurements [44], mats with higher degree of fiber alignment are characterized by improved mechanical strength, but reduced failure strain (shorter linear part of the stress–strain curves) in comparison to less aligned and randomly deposited fibers. Similar trends were observed for the PLA/PBS fibrous materials. As seen from Fig. 4B, the higher collector rotating speed affected significantly the yield point value. Furthermore, the linear part of the stress–strain curves for the samples prepared at 1900 rpm narrowed significantly upon increasing the collector rotation speed (Fig. 4B), attesting for reduced flexibility of the fibrous materials. Thus, higher degree of alignment of the fibers in the materials obtained at the higher collector rotating speed might be stated (Fig. S3A and B). At low collector rotation speed (600 rpm), the Young's modulus, the tensile strength and the elongation did not significantly depend on the PBS content and were determined to be ca. 100 MPa, 2.5 MPa and 90%, respectively (Fig. F5A, B and C). At higher collector rotation speed (1900 rpm), the detected Young's modulus and tensile strength increased by a factor of ca. 4 (350 MPa and 9.0 MPa, respectively), remaining almost independent on the PBS content. The obtained results are in correlation with the calculated higher crystallinity of PLA in PLA/PBS = 100/0 (w/w) mats prepared by applying higher collector rotating speed (Fig. 5D). Concerning the elongation (Fig. 5C), the highest value (130 ± 5%) was obtained for PLA/PBS = 80/20 (w/w). In order to explain this result, several hypotheses including polyester miscibility, blend morphology and crystallinity need more exhaustive investigations. 3.3. Drug loading and biological activity One of the potential applications of the new fibrous PLA/PBS materials is as carriers of low molecular weight bioactive substances with targeted behavior, e.g. having antibacterial and antimycotic activity [45]. In addition, the PLA/PBS mats prepared in this study were rather hydrophobic (the determined water contact angles were in the range 127– 132°). Regarding biomedical applications, the incorporation of an antibacterial agent in PLA/PBS mats would often be advisable since such materials may be a good substrate for biofilm formation. In the present study two 8-hydroxyquinoline (8HQ) derivatives were used as model drugs: the potassium salt of 5-nitro-8-hydroxyquinoline (K5N8Q) and 5-chloro-8-quinolinol (5Cl8HQ). As a polymer carrier PLA/PBS = 80/20 (w/w) mats prepared by using of DCM/EtOH as solvent and at a collector rotating speed of 1900 rpm were selected due to the lower diameter values and adequate mechanical behavior (especially regarding their elasticity behavior). SEM micrographs of the prepared fibers as well as their mean diameters values are shown in Fig. 6. The incorporation of these 8HQ derivatives led to a decrease of the mean diameter value of the fibers, which might be attributed to the
ionogenic nature of the drugs used. This decrease is in accordance with previous findings about fibrous materials prepared by electrospinning of polymer solutions containing an ionogenic low molecular weight additive [40,46,47]. The performed measurements of the water contact angle showed that the incorporation of the bioactive compounds did not alter the water contact angle value, i.e. the mats remained hydrophobic. K5N8Q and 5Cl8HQ are broad-spectrum antibacterial agents with minimum inhibitory concentration against S. aureus of 0.25 μg/mL for 5Cl8HQ [48] and 16 μg/mL for K5N8Q [49]. The antibacterial activity of the electrospun mats was assessed by performance of microbiological tests against this microorganism. The results obtained by determination of the zones of inhibition after a 24-h contact of the fibrous materials with the bacterial cells are shown in Fig. 7. A blank sample (without any fibrous material) is presented as well. As expected PLA/PBS mats did not exhibit any antibacterial activity. Well-distinguished zones of inhibition of the bacterial cells growth were detected for the drug-containing PLA/PBS = 80/20 (w/w) mats. The diameter of the zones of inhibition of PLA/PBS/K5N8Q and PLA/PBS/5Cl8HQ mats did not differ sufficiently, being 37.5 mm and 40.0 mm, respectively. The observation of zones of inhibition is evidence that the incorporated drugs in the mats retain their antibacterial activity. 4. Conclusions For the first time, immiscible blends of PLA and PBS were electrospun to obtain defect-free fibrous mats in the entire range of PLA/PBS weight ratios. Different conditions as solvent system and total polymer concentration were tested in order to determine the optimal electrospinning conditions. The fiber morphology patterns showed the pertinence of the solvent–nonsolvent approach. DSC and XRD analyses attested for the role of PBS on the crystallinity of the electrospun mats. Blending PLA with PBS decreased its Tg and suppressed its cold crystallization, suggesting for a plasticizing effect of PBS. Moreover, a second Tm for PLA appeared at higher temperatures, confirming the influence of PBS on PLA crystallization and crystallization rate. These observations were clearly confirmed by XRD. The electrospinning process revealed crucial for the obtaining of PLA/PBS blend materials maintaining integrity and allowed the preparation of fibrous materials which mechanical properties changed from plastic to brittle depending on the PBS content. The alignment of the blend fibers was controlled by the collector rotation speed and was found to increase the modulus and the tensile strength of the fibrous materials. Drug loading and tests involving pathogenic microorganisms suggested that the obtained mats can find application as antibacterial fibrous materials. Acknowledgments Financial support from the Bulgarian Science Fund (Grant DCVP 02/2/ 2009) is gratefully acknowledged. The authors thank the bilateral
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agreement between WBI/FRS-FNRS and BAS and the financial support from Wallonia and European Commission in the frame of SINOPLISSPOLYEST (Grant no. 565515-346987). N.S. acknowledges the OP-HRD Grant BG051PO001-3.3.06-006 of the European Social Fund. Appendix A. Supplementary data Supplementary data to this article can be found online at http://dx. doi.org/10.1016/j.msec.2014.04.043. References [1] G.-Q. Chen, M.K. Patel, Plastics derived from biological sources: present and future: a technical and environmental review, Chem. Rev. 112 (2012) 2082–2099. [2] J.-M. Raquez, Y. Habibi, M. Murariu, Ph. Dubois, Polylactide (PLA)-based nanocomposites, Prog. Polym. Sci. 38 (2013) 1504–1542. [3] M. Darder, P. Aranda, E. Ruiz-Hitzky, Bionanocomposites: a new concept of ecological, bioinspired, and functional hybrid materials, Adv Mater 19 (2000) 1309–1319. [4] O. Muench, Nano-alloy technology for high-performance PLA applications, Bioplastics Mag. 3 (2008) 10–11. [5] T. Fujimaki, Processability and properties of aliphatic polyesters, ‘BIONOLLE’, synthesized by polycondensation reaction, Polym. Degrad. Stab. 59 (1998) 209–214. [6] J.W. Park, S.S. Im, Phase behavior and morphology in blends of poly(L-lactic acid) and poly(butylene succinate), J. Appl. Polym. Sci. 86 (2002) 647–655. [7] A. Bhatia, R.K. Gupta, S.N. Bhattacharya, H.J. Choi, Compatibility of biodegradable poly (lactic acid) (PLA) and poly (butylene succinate) (PBS) blends for packaging application, Korea-Aust Rheol. J. 19 (2007) 125–131. [8] T. Yokohara, K. Okamoto, M. Yamaguchi, Effect of the shape of dispersed particles on the thermal and mechanical properties of biomass polymer blends composed of poly(L-lactide) and poly(butylene succinate), J. Appl. Polym. Sci. 117 (2010) 2226–2232. [9] P. Zhao, W. Liu, Q. Wu, J. Ren, Preparation, mechanical, and thermal properties of biodegradable polyesters/poly(lactic acid) blends, J. Nanomater. 287082 (2010) 1–8 (Article ID). [10] D. Wu, L. Yuan, E. Laredo, M. Zhang, W. Zhou, Interfacial properties, viscoelasticity, and thermal behaviors of poly(butylene succinate)/polylactide blend, Ind. Eng. Chem. Res. 51 (2012) 2290–2298. [11] E. Hassan, Y. Wei, H. Jiao, Y. Muhuo, Dynamic mechanical properties and thermal stability of poly(lactic acid) and poly(butylene succinate) blends composites, J. Fiber Bioeng. Inform. 6 (2013) 85–94. [12] R. Mincheva, J.-M. Raquez, V. Lison, E. Duquesne, O. Talon, Ph. Dubois, Stereocomplexes from biosourced lactide/butylene succinate-based copolymers and their role as crystallization accelerating agent, Macromol. Chem. Phys. 213 (2012) 643–653. [13] G.X. Chen, H.S. Kim, E.S. Kim, J.Y. Yoon, Compatibilization-like effect of reactive organoclay on the poly(L-lactide)/poly(butylene succinate) blends, Polymer 46 (2005) 11829–11836. [14] M. Shibata, Y. Inoue, M. Miyoshi, Mechanical properties, morphology, and crystallization behavior of blends of poly(L-lactide) with poly(butylene succinate-co-L-lactate) and poly(butylene succinate), Polymer 47 (2006) 3557–3564. [15] J.W. Park, S.S. Im, Morphological changes during heating in poly(L-lactic acid)/ poly(butylene succinate) blend systems as studied by synchrotron X-ray scattering, J. Polym. Sci. B 40 (2002) 1931–1939. [16] D. Paneva, M. Ignatova, N. Manolova, I. Rashkov, in: W.N. Chang (Ed.), Nanofibers: Fabrication, Performance, and Applications, Nova Science Publishers, Inc., New York, 2009, pp. 73–151. [17] N. Bhardwaj, S.C. Kundu, Electrospinning: a fascinating fiber fabrication technique, Biotechnol. Adv. 28 (2010) 325–347. [18] M.Z. Elsabee, H.F. Naguib, R.E. Morsi, Chitosan based nanofibers, review, Mater. Sci. Eng. C 32 (2012) 1711–1726. [19] D. Paneva, R. Mincheva, E. Yancheva, N. Manolova, O. Stoilova, Ph. Dubois, I. Rashkov, in: S.P. Davis (Ed.), Chitosan: Manufacture, Properties, and Usage, Nova Science Publishers, Inc., New York, 2011, pp. 261–320. [20] P. Raghavan, D.-H. Lim, J.-H. Ahn, C. Nah, D.C. Sherrington, H.-S. Ryu, H.-J. Ahn, Electrospun polymer nanofibers: the booming cutting edge technology, React. Funct. Polym. 72 (2013) 915–930. [21] M. Ignatova, I. Rashkov, N. Manolova, Drug-loaded electrospun materials in wounddressing applications and in local cancer treatment, Expert Opin Drug Deliv. 10 (2013) 469–483. [22] M. Ignatova, N. Manolova, I. Rashkov, Electrospun antibacterial chitosan-based fibers, Macromol. Biosci. 13 (2013) 860–872.
[23] M. Bogntizki, T. Frese, M. Steinhart, A. Greiner, J.H. Wendorff, A. Schaper, M. Hellwig, Preparation of fibers with nanoscaled morphologies: electrospinning of polymer blends, Polym. Eng. Sci. 41 (2001) 982–989. [24] L.-T. Lim, K. Cink, T. Vanyo, in: R. Auras, L.-T. Lim, S.E.M. Selke, H. Tsuji (Eds.), Poly(lactic acid): Synthesis, Structures, Properties, Processing, and Applications, John Wiley & Sons, Inc., Hoboken, New Jersey, 2010, pp. 191–215. [25] E.H. Jeong, S.S. Im, J.H. Youk, Electrospinning and structural characterization of ultrafine poly(butylene succinate) fibers, Polymer 46 (2005) 9538–9543. [26] Y. Wu, J.Y. Yu, C. Ma, Electrospun nanoporous fiber, Text. Res. J. 78 (2008) 812–815. [27] Y. Liu, J.-H. He, J.-Y. Yu, Preparation and morphology of poly(butylene succinate) nanofibers via electrospinning, Fibres Text East. Eur. 15 (2007) 30–33. [28] D. Zhang, J. Chang, Y. Zeng, Fabrication of fibrous poly(butylene succinate)/wollastonite/apatite composite scaffolds by electrospinning and biomimetic process, J. Mater. Sci. Mater. Med. 19 (2008) 443–449. [29] S. Sutthiphong, P. Pavasant, P. Supaphol, Electrospun 1,6-diisocyanatohexane-extended poly(1,4-butylene succinate) fiber mats and their potential for use as bone scaffolds, Polymer 50 (2009) 1548–1558. [30] M. Spasova, R. Mincheva, D. Paneva, N. Manolova, I. Rashkov, Perspectives on: criteria for complex evaluation of the morphology and alignment of electrospun polymer nanofibers, J. Bioact. Compat. Polym. 21 (2006) 465–479. [31] E. Fischer, H. Sterzel, G. Wegner, Investigation of the structure of solution grown crystals of lactide copolymers by means of chemical reactions, Kolloid Z. Z. Polym. 251 (1973) 980–990. [32] K. Chrissafis, K.M. Paraskevopoulos, D.N. Bikiaris, Thermal degradation mechanism of poly(ethylene succinate) and poly(butylene succinate): comparative study, Thermochim. Acta 435 (2005) 142–150. [33] J.P. Jeun, Y.M. Lim, Y.C. Nho, Study on morphology of electrospun poly(caprolactone) nanofibers, J. Ind. Eng. Chem. 11 (2005) 573–578. [34] A.G. Kanani, S.H. Bahrami, Effect of changing solvents on poly(ε-caprolactone) nanofibrous webs morphology, J. Nanomater. 724153 (2011) 1–10 (Article ID). [35] J. Zeng, X. Chen, Q. Liang, X. Xu, X. Jing, Enzymatic degradation of poly(L-lactide) and poly(ε-caprolactone) electrospun fibers, Macromol. Biosci. 4 (2004) 1118–1125. [36] R. Inai, M. Kotaki, S. Ramakrishna, Structure and properties of electrospun PLLA single nanofibres, Nanotechnology 16 (2005) 208–213. [37] H. Zhou, Th.B. Green, Y.L. Joo, The thermal effects on electrospinning of polylactic acid melts, Polymer 47 (2006) 7497–7505. [38] E. Yancheva, D. Paneva, N. Manolova, R. Mincheva, D. Danchev, Ph. Dubois, I. Rashkov, Tuning of the surface biological behavior of poly(L-lactide)-based electrospun materials by polyelectrolyte complex formation, Biomacromolecules 11 (2010) 521–532. [39] M. Spasova, N. Manolova, D. Paneva, R. Mincheva, Ph. Dubois, I. Rashkov, V. Maximova, D. Danchev, Polylactide stereocomplex-based electrospun materials possessing surface with antibacterial and hemostatic properties, Biomacromolecules 11 (2010) 151–159. [40] A. Toncheva, D. Paneva, N. Manolova, I. Rashkov, Electrospun poly(L-lactide) membranes containing a single drug or multiple drug system for antimicrobial wound dressings, Macromol. Res. 19 (2011) 1310–1319. [41] Y. Ichikawa, H. Kondo, Y. Igarashi, K. Noguchi, K. Okuyama, J. Washiyama, Crystal structures of α and β forms of poly(tetramethylene succinate), Polymer 41 (2000) 4719–4727. [42] S. Gogolewski, B. Walpoth, P. Rheiner, Polyurethane microporous membranes as pericardial substitutes, Colloid Polym. Sci. 265 (1987) 971–977. [43] J. Li, R.A. Shanks, L. Yu, Time-dependent morphology of polyethylene-polypropylene blends, Chin. J. Polym. Sci. 20 (2002) 497–508. [44] M.S. Rizvi, P. Kumar, D.S. Katti, A. Pal, Mathematical model of mechanical behavior of micro/nanofibrous materials designed for extracellular matrix substitutes, Acta Biomater. 8 (2012) 4111–4122. [45] P. Zahedi, I. Rezaeian, S.-O. Ranaei-Siadat, S.-H. Jafaria, P. Supaphol, A review on wound dressings with an emphasis on electrospun nanofibrous polymeric bandages, Polym. Adv. Technol. 21 (2010) 77–95. [46] A. Toncheva, M. Spasova, D. Paneva, N. Manolova, I. Rashkov, Drug-loaded electrospun polylactide bundles, J. Bioact. Compat. Polym. 26 (2011) 161–172. [47] A. Toncheva, D. Paneva, V. Maximova, N. Manolova, I. Rashkov, Antibacterial fluoroquinolone antibiotic-containing fibrous materials from poly(L-lactide-co-D, Llactide) prepared by electrospinning, Eur. J. Pharm. Sci. 47 (2012) 642–651. [48] P. Hongmanee, K. Rukseree, B. Buabut, B. Somsri, P. Palittapongarnpim, In vitro activities of cloxyquin (5-chloroquinolin-8-ol) against Mycobacterium tuberculosis, Antimicrob. Agents Chemother. 51 (2007) 1105–1106. [49] M.R. Jacobs, R.G. Robinson, H.J. Koornhof, Antibacterial activity of nitroxoline and sulphamethizole alone and in combination in urinary tract infections, S. Afr. Med. J. 54 (1978) 959–962
Contents lists available at ScienceDirect
Materials Science and Engineering C journal homepage: www.elsevier.com/locate/msec
Poly(L-lactide) and poly(butylene succinate) immiscible blends: From electrospinning to biologically active materials Nikoleta Stoyanova a, Dilyana Paneva a, Rosica Mincheva b, Antoniya Toncheva a, Nevena Manolova a, Philippe Dubois b, Iliya Rashkov a,⁎ a b
Laboratory of Bioactive Polymers, Institute of Polymers, Bulgarian Academy of Sciences, Acad. G. Bonchev St, bl. 103A, BG-1113 Sofia, Bulgaria Laboratory of Polymeric and Composite Materials, Center of Innovation and Research in Materials and Polymers (CIRMAP), University of Mons — UMONS, Place du Parc 20, B-7000 Mons, Belgium
a r t i c l e
i n f o
Article history: Received 24 October 2013 Received in revised form 4 April 2014 Accepted 18 April 2014 Available online 28 April 2014 Keywords: Electrospinning Poly(butylene succinate) Poly(L-lactide) Crystallinity Mechanical properties Biological activity
a b s t r a c t For the first time the preparation of defect-free fibers from immiscible blends of high molar mass poly(lactic acid) (PLA) and poly(butylene succinate) (PBS) in the whole range of the polyester weight ratios is shown. Electrospinning using the solvent–nonsolvent approach proved most appropriate. Moreover, electrospinning revealed crucial for the obtaining of PLA/PBS materials maintaining integrity. DSC and XRD analyses attested for a plasticizing effect and for increased PLA crystallinity at PBS addition to PLA. The mechanical properties of the PLA/ PBS mats were controlled by the alignment of the fibers and changed from plastic to brittle materials upon increasing the PBS content. Drug loading and tests against pathogenic microorganisms suggested that the obtained mats can find application as antibacterial fibrous materials. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Materials from biodegradable and non-toxic biocompatible polymers which are synthesized entirely or partially from annually renewable resources have drawn great interest [1]. Their use is imposed by the need for replacement of the conventional materials derived from petroleum sources as well as in the viewpoint of environmental pollution prevention. Among all, the biocompatible and thermoplastic poly(lactide) PLA is one of the most promising biobased polymers [2], finding applications going from drug-carriers and implants to packaging and textiles. More recently, PLA-based materials have found more durable applications in automotive, communication and electronic industries [3,4]. However, drawbacks such as mechanical brittleness and low crystallization rate are often obstacles to PLA practical uses. With this respect, PBS — a partially biobased polyester, with thermal and mechanical properties comparable with those of low density polyethylene and polypropylene [5] was studied as a PLA modifier [6–14]. Because of its high flexibility (low glass-transition temperature), PBS has found applications as films, laminates, molded foams and injection-molded products in various fields, such as agriculture and packaging. The PBS relatively rapid crystallization rate and flexibility were believed to ⁎ Corresponding author. Tel./fax: +359 2 9793468/+359 2 8700309. E-mail address: [email protected] (I. Rashkov).
http://dx.doi.org/10.1016/j.msec.2014.04.043 0928-4931/© 2014 Elsevier B.V. All rights reserved.
improve PLA thermal and mechanical properties when blended and materials from PLA/PBS blends have been prepared by extrusion [6–12]. The miscibility, the phase and crystallization behavior, and the morphology of PLA/PBS blends were reported in the literature [6,15]. Interestingly, despite the improved crystallization of PLA in the presence of PBS, thermal analyses showed two distinct melting peaks over the entire composition range, thus classifying the materials as semicrystalline/semicrystalline blends. Moreover, small-angle X-ray scattering revealed that, in a blend system, the PBS component was expelled out of the interlamellar regions of PLA. Finally, for more than 40 wt.% PBS, significant crystallization-induced phase separation was observed. The thus suggested immiscibility of both polymers [15] is limiting the applications of PLA/PBS blend materials without compatibilization. A possibility to overcome the problem might be found in the electrospinning technique. Electrospinning is a cutting-edge technique that allows micro- and nanofibrous polymer materials to be easily prepared [16–22]. Because of their high surface-to-volume ratio and porous structure, the fibrous mats are well suited for applications such as filters, scaffolds for cell and tissue engineering, and carriers of bioactive substances. Electrospinning of immiscible polymer blends containing PLA is already reported in the literature [23]. The concept resulted in freezing the phase structure generated during electrospinning, thus preserving the co-continuous or interpenetrating phase morphologies within the
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fibers. Noteworthy, unlike PLA that can be easily electrospun from solutions of diverse in nature solvent systems based on chloroform or dichloromethane [24], the electrospinning of PBS solutions encounters certain difficulties and reports on the electrospinning of PBS are still scarce. The preparation of fibrous materials from PBS with a number average molar mass of ca. 75,000 g/mol by electrospinning has been reported [25]. Mixed solvent systems such as chloroform/2-chloroethanol at a ratio of 7/3 or 6/4 (w/w), dichloromethane/2-chloroethanol at a ratio of 7/3 or 6/4 (w/w) and chloroform/3-chloro-1-propanol at a ratio of 9/1 (w/w) have been used. It proved preferable to avoid the use of high boiling solvents such as 2-chloroethanol and 3-chloro-1-propanol when the polymer materials are intended to find application in the biomedical domain or simply to come into contact with human body. A short communication presenting few SEM micrographs of porous fibers prepared by electrospinning of PBS with a molar mass of 20,000 g/mol using chloroform as solvent was published in 2008 [26]. It has been found that the electrospinning of PBS with a molar mass as high as 200,000– 300,000 g/mol using chloroform as solvent is very difficult to achieve when the polymer concentration is lower than 10 wt.% or higher than 15 wt.% [27]. By using PBS with a weight average molar mass MW 3 × 105 g/mol and a chloroform/ethanol solvent system at a ratio of 75/ 25 (v/v) and polymer concentration of 11% (w/v) PBS/wollastonite mats have been electrospun [28]. DCM/trifluoroacetic acid (90/10 v/v) solvent system has been used for electrospinning of 1,6-diisocyanatohexaneextended PBS [29]. Despite the up-to-here described interest, and most probably because of the significant difficulties in performing the electrospinning, there are no literature data on the elaboration of fibrous materials from these two immiscible commercially available high molar mass PLA and PBS polymers through this method. Therefore, the present study aims at preparing defect-free fibers from immiscible blends of PLA and PBS from 100/0 to 10/90 (w/w). The role of the solvent system and the total polymer concentration on fiber morphology was studied. DSC and XRD were used to evaluate crystallinity and crystallization rate of the polyesters in the fibrous mats, and tensile tests were used to determine their mechanical properties. Drug loading and tests with pathogenic microorganisms were also performed as possible material application.
The dynamic viscosity of the spinning solutions was measured by Brookfield LVT viscometer equipped with an adaptor for small samples, a spindle, and a camera SC 4-18/13 R at 20 ± 0.1 °C; the measurements were performed in triplicate and the mean values with their standard deviations were used. For the incorporation of K5N8Q and 5Cl8HQ spinning solutions of PLA/PBS = 80/20 (w/w) in DCM/EtOH = 90/10 (w/w) at total polymer concentration of 10 wt.% were used; the concentration of K5N8Q and 5Cl8HQ was 10 or 20 wt.% in respect to the polymer weight, respectively. In the case of K5N8Q due to its poor solubility, the electrospinning was performed from a stable dispersion. The electrospinning was carried out using an electrospinning set-up consisting of a high voltage power supply (up to 30 kV); a pump [NE300 Just Infusion™ Syringe Pump (New Era Pump Systems Inc., USA)] for delivering the spinning solution at a constant rate; a syringe provided with a positively charged metal needle with inner diameter of 0.6 mm and a custom-made grounded rotating aluminum collector. The electrospinning was performed under the following conditions: feeding rate: 3 mL/h, applied voltage: 25 kV, tip-to-collector distance: 17 cm, collector rotating speed: 600 or 1900 rpm, humidity: 70% and temperature: 20 °C. The obtained mats were dried additionally under reduced pressure (320 Pa) at r.t. for 8 h. 2.3. Characterization of the electrospun materials The morphology of the fibers was examined by a scanning electron microscope (SEM). The samples were vacuum-coated with gold and observed by a Jeol JSM-5510 SEM. The fiber morphology was evaluated by applying the criteria for complex evaluation of electrospun materials as described elsewhere [30] using the ImageJ software by measuring the diameters of at least 20 fibers from each SEM micrograph. The thermal behavior of the prepared new fibrous materials was followed by differential scanning calorimetry (DSC). The analyses were performed using a PerkinElmer DSC 8500 differential scanning calorimeter in nitrogen atmosphere at a heating rate of 10 °С/min. PLA PBS (χPLA c , %) and PBS (χc , %) crystallinity degree was calculated using the following equations: h i PLA PLA PLA PLA;0 = ΔHm 100 ΔH m −ΔH cc W
χc
PLA
2.1. Reagents
χc
PBS
Poly(butylene succinate) (PBS, Bionolle 1003; Shōwa Denkō Kabushiki-gaisha — Japan; MW = 97,600 g/mol, MW /Mn = 2.56 as determined by size-exclusion chromatography using polystyrene standards); poly(L-lactide) (PLA, Ingeo™ Biopolymer 4032D, NatureWorks LLC — USA; MW = 259,000 g/mol; MW /Mn = 1.94; as determined by size-exclusion chromatography using polystyrene standards), dichloromethane (DCM, Merck); ethanol (abs. EtOH, Merck); the potassium salt of 5-nitro-8-hydroxyquinoline (K5N8Q) and 5-chloro-8-quinolinol (5chloro-8-quinolinol; 95% 5Cl8HQ; Sigma-Aldrich) were of analytical grade of purity.
where WPLA is the PLA weight fraction, and WPBS — the PBS weight fraction in the spinning solution; ΔH0m is the heat of fusion of 100% polymer crystal: ΔHPLA,0 = 93.0 J/g [31]; ΔHPBS,0 = 210.0 J/g [32]. The measurem m ments were performed in triplicate and the mean values with their standard deviations were used for interpreting the obtained results. The absence/presence of a crystalline phase in the fibrous materials was assessed by X-ray diffraction analysis (XRD). XRD spectra were recorded at r.t. using a computer controlled D8 Bruker Advance powder diffractometer with a filtered CuKα radiation source and a luminescent detector. The analyses were performed in the 2θ range from 5° to 50° with a step of 0.02° and counting time of 1 s/step. Mechanical properties were evaluated by tensile measurements performed on the fibrous scaffolds using a Zwick/Roell Z 2.5 apparatus, load cell 2 mV/V, type Xforce P, Nominal Force 2.5 kN and test Xpert II. The specimens' thickness was around 50 μm. The stretching rate was 10 mm/min, and the initial length between the clamps was 60 mm. The fibrous materials were tested along the circumferential direction of the collector on which the mat was deposited, and the mean values were calculated. The water contact angle of the mats was determined using Easy Drop DSA20E KRÜSS GmbH device at r.t. A drop of deionized water (10 μL) was deposited on the mats surface. From the images of the
;% ¼
ð1Þ
2. Experimental
2.2. Preparation of PLA/PBS fibrous materials by electrospinning Spinning solutions in DCM or DCM/EtOH [DCM/EtOH = 90/10 or 80/20 (w/w)] were prepared for PLA/PBS blends of PLA/PBS = 100/0; 90/10, 80/20, 70/30, 60/40, 50/50, 40/60, 30/70, 20/80, 10/ 90 and 0/100 (w/w). For preparation of the solutions, PLA and PBS were first dissolved separately in DCM; for the dissolution of PBS heating at 40 °C for 2 h was necessary. Then, the solutions were mixed at stirring. The total polyester concentration was 10 or 15 wt.%. In the case of the DCM/EtOH solvent systems, ethanol was added after the mixing of PLA and PBS solutions in DCM.
h i PBS PBS PBS;0 100 ; % ¼ ΔH m = ΔHm W
ð2Þ
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PLA
PLA/PBS = 90/10 (w/w)
PLA/PBS = 20/80 (w/w)
121
PLA/PBS = 80/20 (w/w)
PLA/PBS = 10/90 (w/w)
PBS
Fig. 1. Representative SEM images of PLA/PBS fibers prepared at PLA/PBS from 100/0 to 0/100 (w/w) at total polymer concentration of 10 wt.% and by using a mixed solvent system DCM/ EtOH; collector rotating speed: 600 rpm; magnification: ×1000.
droplets on the surface of the mats acquired by a digital camera and processed by a software program the average values of the water contact angles were determined based on at least 10 measurements. The samples for the water contact angle measurements were cut in the collector rotation direction and collector axis direction. 2.3.1. Microbiological tests The antibacterial activity of PLA/PBS = 80/20 (w/w) mats containing 10 wt.% K5N8Q or 20 wt.% 5Cl8HQ was monitored against the pathogenic microorganism Staphylococcus aureus (ATCC 6538 P). In order to measure the zones of inhibition, in vitro studies were performed using Tryptone glucose extract agar (DIFCO Laboratories, Detroit, USA) solid medium. The surface of the solid agar was inoculated with a suspension of S. aureus bacterial culture with a cell concentration of 1 × 105 cells/mL and on the surface of the agar in each Petri dish two mats (disks with diameters of 10 mm and weight 1.0 mg) from drug-(non)containing PLA/PBS = 80/20 (w/w) were placed. The Petri dishes were incubated for 24 h at 37 °C and subsequently the zones of inhibition around the disks were measured. The average diameters of the zones of inhibition were determined using the ImageJ software based on 15 measurements in 15 different directions for each zone.
A
3. Results and discussion 3.1. Electrospinning and fiber morphology Attempts to obtain uniform PLA/PBS fibers from both immiscible polymers were performed by electrospinning their blend solutions at 30 kV and at collector rotating speed of 600 rpm. DCM — a common solvent for both polyesters was first studied at total polymer concentration of 10 wt.% (Fig. S1). Thus, cylindrical defect-free fibers were obtained up to PLA/PBS = 70/30 (w/w). Further increase in PBS content resulted in spindle-like defects along the fibers (to 50 wt.% PBS) or beaded fibers (PBS content from 80 wt.% to 100 wt.%). Further experiments were performed at a total polymer concentration of 15 wt.% (DCM, 30 kV, 600 rpm) allowing the obtaining of defect-free fibers at 50 wt.% PBS (mean diameter of 7.3 ± 2.1 μm) (Fig. S2). However, the electrospinning process was ineffective (and even impossible at higher PBS content) due to rapid phase separation and gelation of the spinning solution. For overcoming these problems, the solvent–nonsolvent approach was attempted. It is known that adding a certain amount of a poor solvent to the spinning solution assists in the formation of fibers during electrospinning [33,34]. Thus, in the present study DCM was partially
B 500
3000
Dynamic viscosity, cP
Mean fiber diameter value, nm
3500
2500 2000 1500 1000
400
300
200
100
500 0 100/0
80/20
60/40
40/60
PLA/PBS, w/w
20/80
0/100
0 100/0 80/20 60/40 40/60 20/80 0/100
PLA/PBS, w/w
Fig. 2. Mean fiber diameter (A), and dynamic viscosity (B) for different PLA/PBS weight ratios; total polymer concentration: 10 wt.%; collector rotating speed: 600 rpm.
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Fig. 3. DSC thermograms (first heating run, A) and XRD patterns (B) at selected PLA/PBS weight ratios. DCM/EtOH; collector rotating speed: 600 rpm.
Table 1 Thermal characteristics of PLA/PBS mats as obtained from DCM/EtOH at 600 rpm. PLA/PBS
Tg PLA °C
Tcc PLA °C
ΔH cc PLA J/g⁎
Tm PLA °C
ΔH m PLA J/g⁎
χPLA %⁎⁎
Tcc PBS °C
Tm PBS °C
ΔHm PBS J/g⁎
χPBS %⁎⁎
100/0 90/10 80/20 70/30 60/40 50/50 40/60 30/70 20/80 10/90 0/100
67 58 51 53 54 52 52 52 48 48 –
90 78 70 – – – – – – – –
21.2 15.3 18.1 – – – – – – – –
167 168 166 168 168 166 168 168 167 168 –
31.3 33.6 34.4 26.3 24.2 19.2 20.4 9.2 16.2 6.5 –
10.9 21.9 21.9 22.7 25.1 29.5 35.0 30.4 50.0 70.0 –
– – – 91 89 88 89 88 88 – –
– – 114 114 114 110 114 114 112 114 116
– – 6.0 16.8 27.2 35.6 38.6 52.5 52.0 67.9 77.0
– – 14.3 26.7 32.4 29.8 30.6 35.7 31.0 36.0 36.0
⁎ The enthalpies are not recalculated taking into account the respective polyester content. ⁎⁎ Crystallinity degrees are calculated as described in the Exp. Part. ΔHccPBS is not presented since the cold crystallization overlapped PBS melting.
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fabricated using DCM/EtOH were in the range 1800–600 nm depending on the PBS content (Fig. 2A). As seen, d continuously decreased upon increasing PBS content and the thinnest defect-free fibers were obtained at 90 wt.% PBS (590 ± 140 nm). This effect might be related to the changes in the dynamic viscosity of the spinning solutions (Fig. 2B). Indeed, the dynamic viscosity decreased upon increasing PBS content — the polyester of lower molar mass (MW = 97,600). 3.2. Crystallinity and mechanical properties of the fibrous mats
Fig. 4. Stress–strain curves recorded during mechanical tests of PLA/PBS mats prepared at a collector rotating speed of 600 (A) or 1900 (B) rpm; solvent system: DCM/EtOH; the samples were cut in such a manner so that their length axis coincided with the direction of the collector rotation.
replaced by EtOH — a poor solvent for both PBS and PLA, while the total polymer concentration, the voltage and the collector rotating speed were kept constant (10 wt.%, 30 kV and 600 rpm, respectively). It was found that the addition of only 10 wt.% EtOH to DCM significantly improves the electrospinning of PLA/PBS = 50/50 (w/w) solutions. In contrast to DCM, the fibers obtained from DCM/EtOH = 90/10 (w/w) were cylindrical and defect-free. Their mean diameter was 1.1 ± 0.4 μm. Further increase of EtOH content in the spinning solution [DCM/EtOH = 80/20 (w/w)] was counterproductive as spindle-like defects appeared along the fiber axis (Fig. S2). Therefore the preparation of defect-free fibrous materials from PLA/PBS at the whole range of weight ratios was studied in the system DCM/EtOH = 90/10 (w/w) at total polymer concentration of 10 wt.% (further denoted as DCM/EtOH). SEM micrographs of the obtained fibers are presented in Fig. 1. As seen, the use of DCM/EtOH (30 kV, 600 rpm) allowed the obtaining of randomly deposited defect-free fibers from the immiscible PLA and PBS up to 80 wt.% PBS. Even at PLA/PBS = 10/90 (w/w) only few spindle-like defects appeared and the mats were predominantly composed of defect-free fibers. For neat PBS the obtained fibrous material consisted of fibers (mean diameter: 170 ± 48 nm) and bead-like defects (mean diameter: 7.8 ± 2 μm). This pattern is quite different as compared to DCM solutions, thus confirming the pertinence of the solvent–nonsolvent approach. The mean diameters of PLA/PBS fibers (d)
The as prepared fibrous mats were further characterized in terms of crystallinity and mechanical properties. Representative DSC thermograms of PLA/PBS mats depending on the PBS content are shown in Fig. 3A. Concerning the PLA component, Tg, Tcc and Tm were detected at low PBS content. This occurrence of cold crystallization is in agreement with literature data on electrospun PLA materials and might be explained by the rapid solvent evaporation hampering the polyester crystallization [35–40]. Moreover, Tg and Tcc decreased with ca. 20 °C upon increasing PBS content to 20 wt.% (Table 1). In parallel, the enthalpy of cold crystallization decreased with ca. 15%, thus suggesting some miscibility between the polyesters in the amorphous state at low PBS content. These data are in agreement with the literature on PLA/PBS materials prepared by extrusion [6,8,17], evidencing the role of PBS as plasticizer for PLA. Further increase in PBS content did not influence the PLA Tg, but led to the disappearance of Tcc, suggesting that blending with PBS most probably influenced the PLA crystallization and crystallization rate. In agreement, the melting endotherm of PLA turned bimodal and the corresponding crystallinity increased. The second Tm newly occurring at higher temperatures (max at 176 °C for PLA/PBS = 20/80, w/w) might be ascribed to the formation of crystals which are organized to a greater extent, as observed for PLA/PBS extruded blends [8]. It is to be noted that the PBS Tg was difficult to observe due to the high crystallinity and crystallization rate of the polyester. Therefore, avoiding any speculation on electrospinning induced miscibility of the PLA/PBS blends based on Tg determination is preferable. Regarding PBS, no significant difference in thermal behavior was detected between the neat and the blended fibrous mats. PBS is rapidly crystallizing polyester of high crystallinity [32], making its Tg difficult to assess. Therefore, no values were collected from the DSC analyses. The polyester melting appeared at ca. 116 °C and was directly preceded by a cold crystallization exotherm in agreement with the literature data [32]. Interestingly, Tm was not influenced by the mat composition and the corresponding enthalpy of melting increased upon increasing PBS content. The obtained results were confirmed by the performed XRD analyses. XRD data on films of PLA and PBS homopolyesters are available in the literature [41,42]. According to the literature, the PLA pattern presents diffraction peaks at 2θ = 14.8°, 16.6°, 19.0°, 22.4° and 29.2° distinctive for the α-form of optically pure PLLA (or PDLA) crystallizing in a pseudo-orthorhombic unit cell containing l03 helices [42], while PBS presents diffraction peaks at 2θ = 19.6°, 21.9°, 22.8° and 29.1° ascribed to its α-form (monoclinic system of 21 helices) [41]. As seen from Fig. 3B, the XRD patterns of PLA/PBS = 100/0 and 80/20 (w/w) mats revealed only the presence of an amorphous halo. Diffractions for crystal phase were not observed, thus confirming the hindered crystallization of the polyesters during the electrospinning process as evidenced by DSC. In the XRD pattern of PLA/PBS = 50/50 (w/w) mat a diffraction at 2θ = 22.6° was registered, and in the XRD pattern of PLA/PBS = 20/80 (w/w) mat: two diffractions were observed at 2θ = 19.6° and 22.6°, respectively. These values of 2θ are characteristic for both PLA and PBS crystal structure, and their attribution cannot be done only on the basis of the XRD data. However and taking into consideration their crystallinity degree calculated by DSC (Table 1), it can be assumed that the registered diffractions were due to the crystal structure of both PLA and PBS.
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A
B 12.5
600 rpm 1900 rpm
Tensile strength, MPa
Young's Modulus, MPa
500 400 300 200 100
80/20
50/50
7.5 5.0
0.0 100/0
20/80
*
*
50/50
20/80
2.5
80/20
PLA/PBS, w/w
PLA/PBS, w/w
C
D 600 rpm
125 100
* 75
*
50
*
25 0
100/0
80/20
60
1900 rpm
*
50/50
20/80
Crystallinity degree, %
150
Elongation, %
1900 rpm * *
10.0
0 100/0
600 rpm
PLA/600 rpm PLA/1900 rpm PBS/600 rpm PBS/1900 rpm
50 40 30 20 10 0
100/0
PBS content, wt.%
80/20
50/50
20/80
PLA/PBS, w/w
Fig. 5. Dependence of Young's modulus (A), tensile strength (B) and elongation at break (C) of PLA/PBS mats, and the crystallinity degree of PLA and PBS (D) in the fibrous materials on PBS content in the mats, prepared at collector rotation speed of 600 or 1900 rpm; solvent system: DCM/EtOH. The samples for which a break during the tests was detected are marked with an asterisk; thus, the obtained values may be considered as elongation at break.
It is worth to be mentioned that solvent-cast films from PBS and PLA/PBS lost their integrity and became very brittle on drying. In sharp contrast, PLA/PBS fibrous mats maintained integrity and could be easily handled without fragmentation. In order to determine their mechanical properties tensile tests on mats of defect-free fibers were performed. The stress–strain curves of PLA/PBS fibrous materials at different PBS contents are presented in Fig. F4A. Digital images of the samples after testing are shown as well. As seen, before reaching the ultimate tensile strength, the PLA/PBS = 100/0 (w/w) and PLA/PBS = 80/20 (w/w) mats displayed typical strain behavior of thermoplastic materials. At low stress values (in the elastoplasticity range) they rapidly reached the yield point after which necking
was observed. This necking after the yield point gave evidence for the occurrence of inelastic deformation. The subsequent increase of the stress led to hardening of the samples most probably due to the additional orientation of the fibers during specimens' elongation. Partial tearing of the samples was observed after reaching the maximum tensile stress. Further increase in PBS content (PLA/PBS = 50/50 (w/w) and PLA/PBS = 20/80 (w/w)) resulted in distinct break point, and the absence of necking, indicating quasi-ductile fracture behavior of these materials. One possible explanation of this finding might be the alignment of the fibers during stretching. However, based on the DSC and XRD data alteration in the blend morphology: a transition from dispersed structure to bicontinuous one and again to dispersed structure; alteration in the crystallinity of the blends and by interfacial adhesion [43] should not be neglected. Effects of
A 1985±400 nm
C 760±310 nm
B 953±475 nm
Fig. 6. SEM micrographs of PLA/PBS fibers prepared at total polymer concentration of 10 wt.% using DCM/EtOH in absence (A) and presence of the drug: 10 wt.% K5N8Q (B) or 20 wt.% 5Cl8HQ (C) (in respect to the total polymer weight); collector rotating speed: 1900 rpm; the collector rotation direction is indicated by a white arrow.
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S. aureus
PLA/PBS
PLA/PBS/K5N8Q
125
PLA/PBS/5Cl8HQ
Fig. 7. Digital images of the zones of inhibition against S. aureus, detected after a 24-h contact of the drug-(non)containing PLA/PBS fibrous materials with the bacterial cells. Left image — blank control.
additional orientation of the macromolecules during specimen elongation [44] are not to be excluded. To verify the hypothesis, fibrous materials were obtained at higher collector rotation speed (1900 rpm), known to increase the degree of fibers' alignment. According to theoretical simulations confirmed by experimental measurements [44], mats with higher degree of fiber alignment are characterized by improved mechanical strength, but reduced failure strain (shorter linear part of the stress–strain curves) in comparison to less aligned and randomly deposited fibers. Similar trends were observed for the PLA/PBS fibrous materials. As seen from Fig. 4B, the higher collector rotating speed affected significantly the yield point value. Furthermore, the linear part of the stress–strain curves for the samples prepared at 1900 rpm narrowed significantly upon increasing the collector rotation speed (Fig. 4B), attesting for reduced flexibility of the fibrous materials. Thus, higher degree of alignment of the fibers in the materials obtained at the higher collector rotating speed might be stated (Fig. S3A and B). At low collector rotation speed (600 rpm), the Young's modulus, the tensile strength and the elongation did not significantly depend on the PBS content and were determined to be ca. 100 MPa, 2.5 MPa and 90%, respectively (Fig. F5A, B and C). At higher collector rotation speed (1900 rpm), the detected Young's modulus and tensile strength increased by a factor of ca. 4 (350 MPa and 9.0 MPa, respectively), remaining almost independent on the PBS content. The obtained results are in correlation with the calculated higher crystallinity of PLA in PLA/PBS = 100/0 (w/w) mats prepared by applying higher collector rotating speed (Fig. 5D). Concerning the elongation (Fig. 5C), the highest value (130 ± 5%) was obtained for PLA/PBS = 80/20 (w/w). In order to explain this result, several hypotheses including polyester miscibility, blend morphology and crystallinity need more exhaustive investigations. 3.3. Drug loading and biological activity One of the potential applications of the new fibrous PLA/PBS materials is as carriers of low molecular weight bioactive substances with targeted behavior, e.g. having antibacterial and antimycotic activity [45]. In addition, the PLA/PBS mats prepared in this study were rather hydrophobic (the determined water contact angles were in the range 127– 132°). Regarding biomedical applications, the incorporation of an antibacterial agent in PLA/PBS mats would often be advisable since such materials may be a good substrate for biofilm formation. In the present study two 8-hydroxyquinoline (8HQ) derivatives were used as model drugs: the potassium salt of 5-nitro-8-hydroxyquinoline (K5N8Q) and 5-chloro-8-quinolinol (5Cl8HQ). As a polymer carrier PLA/PBS = 80/20 (w/w) mats prepared by using of DCM/EtOH as solvent and at a collector rotating speed of 1900 rpm were selected due to the lower diameter values and adequate mechanical behavior (especially regarding their elasticity behavior). SEM micrographs of the prepared fibers as well as their mean diameters values are shown in Fig. 6. The incorporation of these 8HQ derivatives led to a decrease of the mean diameter value of the fibers, which might be attributed to the
ionogenic nature of the drugs used. This decrease is in accordance with previous findings about fibrous materials prepared by electrospinning of polymer solutions containing an ionogenic low molecular weight additive [40,46,47]. The performed measurements of the water contact angle showed that the incorporation of the bioactive compounds did not alter the water contact angle value, i.e. the mats remained hydrophobic. K5N8Q and 5Cl8HQ are broad-spectrum antibacterial agents with minimum inhibitory concentration against S. aureus of 0.25 μg/mL for 5Cl8HQ [48] and 16 μg/mL for K5N8Q [49]. The antibacterial activity of the electrospun mats was assessed by performance of microbiological tests against this microorganism. The results obtained by determination of the zones of inhibition after a 24-h contact of the fibrous materials with the bacterial cells are shown in Fig. 7. A blank sample (without any fibrous material) is presented as well. As expected PLA/PBS mats did not exhibit any antibacterial activity. Well-distinguished zones of inhibition of the bacterial cells growth were detected for the drug-containing PLA/PBS = 80/20 (w/w) mats. The diameter of the zones of inhibition of PLA/PBS/K5N8Q and PLA/PBS/5Cl8HQ mats did not differ sufficiently, being 37.5 mm and 40.0 mm, respectively. The observation of zones of inhibition is evidence that the incorporated drugs in the mats retain their antibacterial activity. 4. Conclusions For the first time, immiscible blends of PLA and PBS were electrospun to obtain defect-free fibrous mats in the entire range of PLA/PBS weight ratios. Different conditions as solvent system and total polymer concentration were tested in order to determine the optimal electrospinning conditions. The fiber morphology patterns showed the pertinence of the solvent–nonsolvent approach. DSC and XRD analyses attested for the role of PBS on the crystallinity of the electrospun mats. Blending PLA with PBS decreased its Tg and suppressed its cold crystallization, suggesting for a plasticizing effect of PBS. Moreover, a second Tm for PLA appeared at higher temperatures, confirming the influence of PBS on PLA crystallization and crystallization rate. These observations were clearly confirmed by XRD. The electrospinning process revealed crucial for the obtaining of PLA/PBS blend materials maintaining integrity and allowed the preparation of fibrous materials which mechanical properties changed from plastic to brittle depending on the PBS content. The alignment of the blend fibers was controlled by the collector rotation speed and was found to increase the modulus and the tensile strength of the fibrous materials. Drug loading and tests involving pathogenic microorganisms suggested that the obtained mats can find application as antibacterial fibrous materials. Acknowledgments Financial support from the Bulgarian Science Fund (Grant DCVP 02/2/ 2009) is gratefully acknowledged. The authors thank the bilateral
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agreement between WBI/FRS-FNRS and BAS and the financial support from Wallonia and European Commission in the frame of SINOPLISSPOLYEST (Grant no. 565515-346987). N.S. acknowledges the OP-HRD Grant BG051PO001-3.3.06-006 of the European Social Fund. Appendix A. Supplementary data Supplementary data to this article can be found online at http://dx. doi.org/10.1016/j.msec.2014.04.043. References [1] G.-Q. Chen, M.K. Patel, Plastics derived from biological sources: present and future: a technical and environmental review, Chem. Rev. 112 (2012) 2082–2099. [2] J.-M. Raquez, Y. Habibi, M. Murariu, Ph. Dubois, Polylactide (PLA)-based nanocomposites, Prog. Polym. Sci. 38 (2013) 1504–1542. [3] M. Darder, P. Aranda, E. Ruiz-Hitzky, Bionanocomposites: a new concept of ecological, bioinspired, and functional hybrid materials, Adv Mater 19 (2000) 1309–1319. [4] O. Muench, Nano-alloy technology for high-performance PLA applications, Bioplastics Mag. 3 (2008) 10–11. [5] T. Fujimaki, Processability and properties of aliphatic polyesters, ‘BIONOLLE’, synthesized by polycondensation reaction, Polym. Degrad. Stab. 59 (1998) 209–214. [6] J.W. Park, S.S. Im, Phase behavior and morphology in blends of poly(L-lactic acid) and poly(butylene succinate), J. Appl. Polym. Sci. 86 (2002) 647–655. [7] A. Bhatia, R.K. Gupta, S.N. Bhattacharya, H.J. Choi, Compatibility of biodegradable poly (lactic acid) (PLA) and poly (butylene succinate) (PBS) blends for packaging application, Korea-Aust Rheol. J. 19 (2007) 125–131. [8] T. Yokohara, K. Okamoto, M. Yamaguchi, Effect of the shape of dispersed particles on the thermal and mechanical properties of biomass polymer blends composed of poly(L-lactide) and poly(butylene succinate), J. Appl. Polym. Sci. 117 (2010) 2226–2232. [9] P. Zhao, W. Liu, Q. Wu, J. Ren, Preparation, mechanical, and thermal properties of biodegradable polyesters/poly(lactic acid) blends, J. Nanomater. 287082 (2010) 1–8 (Article ID). [10] D. Wu, L. Yuan, E. Laredo, M. Zhang, W. Zhou, Interfacial properties, viscoelasticity, and thermal behaviors of poly(butylene succinate)/polylactide blend, Ind. Eng. Chem. Res. 51 (2012) 2290–2298. [11] E. Hassan, Y. Wei, H. Jiao, Y. Muhuo, Dynamic mechanical properties and thermal stability of poly(lactic acid) and poly(butylene succinate) blends composites, J. Fiber Bioeng. Inform. 6 (2013) 85–94. [12] R. Mincheva, J.-M. Raquez, V. Lison, E. Duquesne, O. Talon, Ph. Dubois, Stereocomplexes from biosourced lactide/butylene succinate-based copolymers and their role as crystallization accelerating agent, Macromol. Chem. Phys. 213 (2012) 643–653. [13] G.X. Chen, H.S. Kim, E.S. Kim, J.Y. Yoon, Compatibilization-like effect of reactive organoclay on the poly(L-lactide)/poly(butylene succinate) blends, Polymer 46 (2005) 11829–11836. [14] M. Shibata, Y. Inoue, M. Miyoshi, Mechanical properties, morphology, and crystallization behavior of blends of poly(L-lactide) with poly(butylene succinate-co-L-lactate) and poly(butylene succinate), Polymer 47 (2006) 3557–3564. [15] J.W. Park, S.S. Im, Morphological changes during heating in poly(L-lactic acid)/ poly(butylene succinate) blend systems as studied by synchrotron X-ray scattering, J. Polym. Sci. B 40 (2002) 1931–1939. [16] D. Paneva, M. Ignatova, N. Manolova, I. Rashkov, in: W.N. Chang (Ed.), Nanofibers: Fabrication, Performance, and Applications, Nova Science Publishers, Inc., New York, 2009, pp. 73–151. [17] N. Bhardwaj, S.C. Kundu, Electrospinning: a fascinating fiber fabrication technique, Biotechnol. Adv. 28 (2010) 325–347. [18] M.Z. Elsabee, H.F. Naguib, R.E. Morsi, Chitosan based nanofibers, review, Mater. Sci. Eng. C 32 (2012) 1711–1726. [19] D. Paneva, R. Mincheva, E. Yancheva, N. Manolova, O. Stoilova, Ph. Dubois, I. Rashkov, in: S.P. Davis (Ed.), Chitosan: Manufacture, Properties, and Usage, Nova Science Publishers, Inc., New York, 2011, pp. 261–320. [20] P. Raghavan, D.-H. Lim, J.-H. Ahn, C. Nah, D.C. Sherrington, H.-S. Ryu, H.-J. Ahn, Electrospun polymer nanofibers: the booming cutting edge technology, React. Funct. Polym. 72 (2013) 915–930. [21] M. Ignatova, I. Rashkov, N. Manolova, Drug-loaded electrospun materials in wounddressing applications and in local cancer treatment, Expert Opin Drug Deliv. 10 (2013) 469–483. [22] M. Ignatova, N. Manolova, I. Rashkov, Electrospun antibacterial chitosan-based fibers, Macromol. Biosci. 13 (2013) 860–872.
[23] M. Bogntizki, T. Frese, M. Steinhart, A. Greiner, J.H. Wendorff, A. Schaper, M. Hellwig, Preparation of fibers with nanoscaled morphologies: electrospinning of polymer blends, Polym. Eng. Sci. 41 (2001) 982–989. [24] L.-T. Lim, K. Cink, T. Vanyo, in: R. Auras, L.-T. Lim, S.E.M. Selke, H. Tsuji (Eds.), Poly(lactic acid): Synthesis, Structures, Properties, Processing, and Applications, John Wiley & Sons, Inc., Hoboken, New Jersey, 2010, pp. 191–215. [25] E.H. Jeong, S.S. Im, J.H. Youk, Electrospinning and structural characterization of ultrafine poly(butylene succinate) fibers, Polymer 46 (2005) 9538–9543. [26] Y. Wu, J.Y. Yu, C. Ma, Electrospun nanoporous fiber, Text. Res. J. 78 (2008) 812–815. [27] Y. Liu, J.-H. He, J.-Y. Yu, Preparation and morphology of poly(butylene succinate) nanofibers via electrospinning, Fibres Text East. Eur. 15 (2007) 30–33. [28] D. Zhang, J. Chang, Y. Zeng, Fabrication of fibrous poly(butylene succinate)/wollastonite/apatite composite scaffolds by electrospinning and biomimetic process, J. Mater. Sci. Mater. Med. 19 (2008) 443–449. [29] S. Sutthiphong, P. Pavasant, P. Supaphol, Electrospun 1,6-diisocyanatohexane-extended poly(1,4-butylene succinate) fiber mats and their potential for use as bone scaffolds, Polymer 50 (2009) 1548–1558. [30] M. Spasova, R. Mincheva, D. Paneva, N. Manolova, I. Rashkov, Perspectives on: criteria for complex evaluation of the morphology and alignment of electrospun polymer nanofibers, J. Bioact. Compat. Polym. 21 (2006) 465–479. [31] E. Fischer, H. Sterzel, G. Wegner, Investigation of the structure of solution grown crystals of lactide copolymers by means of chemical reactions, Kolloid Z. Z. Polym. 251 (1973) 980–990. [32] K. Chrissafis, K.M. Paraskevopoulos, D.N. Bikiaris, Thermal degradation mechanism of poly(ethylene succinate) and poly(butylene succinate): comparative study, Thermochim. Acta 435 (2005) 142–150. [33] J.P. Jeun, Y.M. Lim, Y.C. Nho, Study on morphology of electrospun poly(caprolactone) nanofibers, J. Ind. Eng. Chem. 11 (2005) 573–578. [34] A.G. Kanani, S.H. Bahrami, Effect of changing solvents on poly(ε-caprolactone) nanofibrous webs morphology, J. Nanomater. 724153 (2011) 1–10 (Article ID). [35] J. Zeng, X. Chen, Q. Liang, X. Xu, X. Jing, Enzymatic degradation of poly(L-lactide) and poly(ε-caprolactone) electrospun fibers, Macromol. Biosci. 4 (2004) 1118–1125. [36] R. Inai, M. Kotaki, S. Ramakrishna, Structure and properties of electrospun PLLA single nanofibres, Nanotechnology 16 (2005) 208–213. [37] H. Zhou, Th.B. Green, Y.L. Joo, The thermal effects on electrospinning of polylactic acid melts, Polymer 47 (2006) 7497–7505. [38] E. Yancheva, D. Paneva, N. Manolova, R. Mincheva, D. Danchev, Ph. Dubois, I. Rashkov, Tuning of the surface biological behavior of poly(L-lactide)-based electrospun materials by polyelectrolyte complex formation, Biomacromolecules 11 (2010) 521–532. [39] M. Spasova, N. Manolova, D. Paneva, R. Mincheva, Ph. Dubois, I. Rashkov, V. Maximova, D. Danchev, Polylactide stereocomplex-based electrospun materials possessing surface with antibacterial and hemostatic properties, Biomacromolecules 11 (2010) 151–159. [40] A. Toncheva, D. Paneva, N. Manolova, I. Rashkov, Electrospun poly(L-lactide) membranes containing a single drug or multiple drug system for antimicrobial wound dressings, Macromol. Res. 19 (2011) 1310–1319. [41] Y. Ichikawa, H. Kondo, Y. Igarashi, K. Noguchi, K. Okuyama, J. Washiyama, Crystal structures of α and β forms of poly(tetramethylene succinate), Polymer 41 (2000) 4719–4727. [42] S. Gogolewski, B. Walpoth, P. Rheiner, Polyurethane microporous membranes as pericardial substitutes, Colloid Polym. Sci. 265 (1987) 971–977. [43] J. Li, R.A. Shanks, L. Yu, Time-dependent morphology of polyethylene-polypropylene blends, Chin. J. Polym. Sci. 20 (2002) 497–508. [44] M.S. Rizvi, P. Kumar, D.S. Katti, A. Pal, Mathematical model of mechanical behavior of micro/nanofibrous materials designed for extracellular matrix substitutes, Acta Biomater. 8 (2012) 4111–4122. [45] P. Zahedi, I. Rezaeian, S.-O. Ranaei-Siadat, S.-H. Jafaria, P. Supaphol, A review on wound dressings with an emphasis on electrospun nanofibrous polymeric bandages, Polym. Adv. Technol. 21 (2010) 77–95. [46] A. Toncheva, M. Spasova, D. Paneva, N. Manolova, I. Rashkov, Drug-loaded electrospun polylactide bundles, J. Bioact. Compat. Polym. 26 (2011) 161–172. [47] A. Toncheva, D. Paneva, V. Maximova, N. Manolova, I. Rashkov, Antibacterial fluoroquinolone antibiotic-containing fibrous materials from poly(L-lactide-co-D, Llactide) prepared by electrospinning, Eur. J. Pharm. Sci. 47 (2012) 642–651. [48] P. Hongmanee, K. Rukseree, B. Buabut, B. Somsri, P. Palittapongarnpim, In vitro activities of cloxyquin (5-chloroquinolin-8-ol) against Mycobacterium tuberculosis, Antimicrob. Agents Chemother. 51 (2007) 1105–1106. [49] M.R. Jacobs, R.G. Robinson, H.J. Koornhof, Antibacterial activity of nitroxoline and sulphamethizole alone and in combination in urinary tract infections, S. Afr. Med. J. 54 (1978) 959–962