- Email: [email protected]
Encapsulation of β-carotene in poly(vinylpyrrolidone) (PVP) by Electrospinning Technique
Available online at www.sciencedirect.com
ScienceDirect Procedia Engineering 170 (2017) 19 – 23
Engineering Physics International Conference, EPIC 2016
Encapsulation of β-carotene in poly(vinylpyrrolidone) (PVP) by electrospinning technique Rhyan Prayuddy Reksamunandara,b, Dhewa Edikresnhaa,b, Muhammad Miftahul Munira,b, Sophi Damayantic, and Khairurrijala,b,* a
Department of Physics Faculty of Mathematics and Natural Sciences, b Research Center for Bioscience and Biotechnology, c Department of Pharmacochemistry, School of Pharmacy, Institut Teknologi Bandung, Jalan Ganesa 10, Bandung 40132, Indonesia
Abstract β-carotene, which is the most commonly known type of carotenoids, has been proven to have antioxidant properties and to provide provitamin A activity that are important for human health. In this study, β-carotene was encapsulated into poly(vinylpyrrolidone)/PVP through electrospinning technique in order to protect β-carotene from degradation and to maintain the antioxidant properties. The precursor solution was prepared by dissolving β-carotene (1 wt.%) to various concentrations of PVP (6 wt.%, 8 wt.%, 10 wt.%, and 12 wt.%) in ethanol. SEM images showed the formation of nanoscale beaded fibers with generally increasing average diameter when the PVP concentration was increased except when the concentration of precursor solution was 12 wt.% PVP. In this case, it became too viscous and caused clogging on the needle tip and the formation of multi-jets. The average diameters of PVP/β-carotene composite nanofibers were in the range of 176 to 306 nm. The analyses of Fourier Transform Infrared (FTIR) spectra proved the presence of structural changes in PVP nanofibers after being loaded by β-carotene as a result of molecular interaction between β-carotene and PVP. This was given by a significant shift of the peak at 1440 cm-1 to lower wavenumbers of the FTIR spectrum of PVP. The β-carotene was proven to retain their antioxidant ability even after being encapsulated into the PVP nanofibers as tested by the DPPH assay method. © 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). © 2016 The Authors. Published by Elsevier Ltd. Peer-review under responsibility of the organizing committee of the Engineering Physics International Conference 2016 Keywords: β-carotene; poly(vinylpyrrolidone); electrospinning; antioxidant.
1. Introduction Carotenoids are natural pigments that are abundant in nature and available in various types of fruits, vegetables, leaves and few animals. Carotenoids provide provitamin A activity that contributes to human health since they have antioxidant properties, such as to protect cells against reactive oxygen and free radicals, to reduce the risk of cancer or other chronic diseases, to stimulate body growth, increase immune system and improve visual function [1-2]. The most commonly known of carotenoid is β-carotene [3]. β-carotene is a non-polar compound with a long chain of conjugated double bonds and unsaturated structures which cause β-carotene to be easily degraded and is highly sensitive to light and heat [4]. To stabilize β-carotene from degradation, encapsulation through electrospinning technique has been studied [5-7]. However, the number of studies on βcarotene being encapsulated by electrospinning is still rare and no detailed explanation is given regarding the effect of polymer concentration on the average diameter of composite fibers and the antioxidant properties of the β-carotene after being encapsulated. Hence, this study investigated the effect of varying the polymer concentration towards the diameter and the antioxidant properties of β-carotene.
* Corresponding author. Tel.: +62-22-250 0834; fax: +62-22-250 6452. E-mail address: [email protected]
1877-7058 © 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Peer-review under responsibility of the organizing committee of the Engineering Physics International Conference 2016
doi:10.1016/j.proeng.2017.03.004
20
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23
Electrospinning is a technique to produce the polymer fibers or their composite with high surface area to volume ratio. Electrospinning system has been designed and built to produce nanofibers with relatively narrow size distribution [8-10]. In this study, a biocompatible polymer, poly(vinylpyrrolidone) (PVP), was used as the polymer matrix with ethanol as a solvent for the precursor solution. PVP was chosen due to its acceptable level of toxicity, biocompatibility and solubility both in nonpolar and polar solvents [11]. The electrospinnability of PVP in ethanol solvent was already proven in the previous studies [12, 13]. This study reports the formation of composite nanofibers between PVP and β-carotene known for its antioxidant properties by means of electrospinning to develop an encapsulating technology for β-carotene. The morphology, the chemical compounds, and the antioxidant ability of the composite fibers will be described and discussed. 2. Method PVP (poly(vinylpyrrolidone, MW = 1300 kg/mol) and β-carotene were purchased from Sigma-Aldrich, whereas ethanol was provided by a local supplier. The polymer solution was made by mixing PVP in ethanol as the solvent in various ratios of PVP (6 wt.%, 8 wt.%, 10 wt.%, and 12 wt.%) together with 1 wt.% β-carotene. All components were then magnetically stirred for three hours at room temperature until the solution was mixed homogeneously. To obtain nanofibers mats of PVP/β-carotene, each precursor solution was inserted into a single needle syringe with inner needle diameter of 0.84 mm. Each solution was electrospun using a drum collector in electrospinning apparatus with the distance between the needle and the collector was set to be 10 cm, the flow rate used was 1 mL/h, and the applied voltage was 15 kV. The fibers were collected on an aluminum foil that was coated on the drum collector. The mats produced were kept at the temperature of 5 to 6 oC and were protected from light until being used for characterizations and antioxidant test. The effects of β-carotene loaded in variations of PVP solution to the morphology of the nanofibers and the diameter size distribution were observed using Scanning Electron Microscope (SEM) (JEOL, JCM-6000 NeoScope Benchtop) with an accelerating voltage of 15 kV. The infrared spectra of the nanofibers were analyzed by Fourier Transform Infrared (FTIR) spectrometer (Bruker Alpha, A220/D-01) to demonstrate the changes in the structure of PVP/β-carotene nanofibers. The wavenumber spectrum used for the FTIR test was within the range of 500-4000/cm-1. The antioxidant ability of the PVP/βcarotene nanofibers was tested using 1,1-diphenyl-2-picrylhydrazil (DPPH) assay method [14]. The level of antioxidant of the composite fibers can be determined by Equation (1).
% AA =
Acontrol − Asample Acontrol
(1)
×100
where Acontrol is the absorbance of the 25 ppm DPPH solution without PVP/β-carotene fibers and Asample is the absorbance of the 50 ppm DPPH solution mixed with 50 ppm PVP/β-carotene fibers in ethanol with a volume ratio of 1:1 after being incubated for 30 minutes in dark condition. The absorbance spectrum was measured using UV-Visible Spectrophotometer (Agilent, 8453) and monitored at the wavelength of 515 nm which is the peak level of the DPPH solution [14]. 3. Results and Discussion 3.1. Morphology of the composite nanofibers The morphologies of the PVP/β-carotene composite fibers are given by the SEM images with a magnification of 4000 times in Figure 1. They agreed well to those obtained from the previous studies [12, 13], but with some beads. The presence of beads was related to the solution viscosity, entanglement number of solution, and the additional substance inside the fibers [15]. Beads can be predicted to occur whenever the surface tension exceeds the charge repulsive force and viscoelastic force that support the elongation of continuous jet [16].
Fig. 1. The SEM images of the composite nanofibers of (a) 6 wt.% PVP/1 wt.% β-carotene, (b) 8 wt.% PVP/1 wt.% β-carotene, (c) 10 wt.% PVP/1 wt.% βcarotene, and (d) 12 wt.% PVP/1% β-carotene.
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23
Figure 2 gives the fibers diameter distributions of the PVP/β-carotene composite fibers for (a) 6 wt.% PVP/1 wt.% βcarotene, (b) 8 wt.% PVP/1 wt.% β-carotene, (c) 10 wt.% PVP/1 wt.% β-carotene, and (d) 12% PVP/1% β-carotene. It was found that the average diameter of fibers increases with the increase of polymer concentration. This finding can be explained as follows: a higher polymer concentration in the precursor solution will increase the chain entanglement and the amount of readily available polymer to be electrospun, hence the diameter of fiber will be larger [16]. Additionally, the increase of polymer concentration (thus lowering the wt.% of the solvent) causes the solvent to dry up faster and less time for the repulsive columbic force to stretch the fiber, the diameter of fiber will be larger [12]. However, the composite nanofibers obtained from the 12 wt.% PVP/1% β-carotene had uneven fibers diameter distribution and smaller average diameter of fibers. An explanation of this finding is as follows: the back and side parts of the liquid stream coming out from the needle at a later phase can be forced to be ejected as small side jets when the applied voltage is high enough [17]. A special case happened for the high composition of polymer (higher than 12 wt.% PVP) in which the tip blockage occurred. This is due to the rapid rate of solvent evaporation so that the liquid stream solidifies quickly while the syringe pump still pushes the liquid forward [16].
Fig. 2. The corresponding bar diagrams of fibers diameters and count rate of (a) 6 wt.% PVP/1 wt.% β-carotene, (b) 8 wt.% PVP/1 wt.% β-carotene, (c) 10 wt.% PVP/1 wt.% β-carotene, and (d) 12 wt.% PVP/1% β-carotene.
3.2. FTIR spectroscopy Figure 3 shows the FTIR spectra of β-carotene, PVP, and PVP/β-carotene. In the FTIR spectrum of β-carotene given in Fig. 3(a), the broad peak at 3411 cm-1 represents the presence of O-H stretching of the hydroxyl group, which is likely due to the interaction of β-carotene with oxygen in the air [5]. The peaks at 2929 cm-1 and 2869 cm-1 indicate the CH2 asymmetry and symmetry stretching, respectively. The peaks at 1717 cm-1 and 1366 cm-1 exhibit the presence of carbonyl groups and stretching symmetry of C-H bond group, respectively. The sharp peak at 965 cm-1 marks the deformation mode of trans-conjugate-alkenes as the specific areas of trans=CH used for identification of β-carotene [18].
Fig. 3. FTIR spectra of (a) β-carotene, (b) PVP, (c) 6 wt.% PVP/1 wt.% % β-carotene, (d) 8 wt.% PVP/1 wt.% β-carotene, (e) 10 wt.% PVP/1 wt.% β-carotene, and (f) 12 wt.% PVP/1 wt.% β-carotene.
In the FTIR spectrum of PVP given in Fig. 3(b), the broad band around 3550 - 3200 cm-1 assign as O-H stretching of the hydroxyl group. The presence of heteroatomic molecules and carbonyl groups in the pyrrolidone ring of PVP was identified by the sharp peak at 1656 cm-1 as the signature of C=O stretching and the peak at 1291 cm-1 is related to CN stretch of amide group [19, 20]. The peak at 1440 cm-1 assign as the C-H deformation of CH2 group. The FTIR spectra of PVP/β-carotene in Figs. 3(c)3(d) show a significant shift of the peak at 1440 cm-1 to lower wavenumbers due to the encapsulation of β-carotene in PVP fibers. On the other hand, the broad band (3550 - 3200) cm-1 and the peak at 1656 cm-1 did not shift significantly and the sharp peak at 965 cm-1 disappeared.
21
22
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23
3.3. Antioxidant activity Since β-carotene is already known to have antioxidant properties, this study was intended to find out the effectiveness of the antioxidant property of β-carotene after being encapsulated into the composite nanofibers mats. The antioxidant test was done four days after the production of the composite nanofibers mats. The β-carotene-encapsulated nanofibers mats were mixed with DPPH, a notorious free radical usually used to measure the antioxidant properties of any sample. DPPH is typically violet in color and becomes pale yellow when neutralized [14]. Table 1. Antioxidant activity of PVP/β-carotene composite nanofibers.
Composite nanofibers PVP 6 wt.% loaded 1 wt.% β-carotene PVP 8 wt.% loaded 1 wt.% β-carotene PVP 10 wt.% loaded 1 wt.% β-carotene PVP 12 wt.% loaded 1 wt.% β-carotene
Antioxidant activity (%) 47 + 20 47 + 30 86 + 1.2 21 + 12
Average diameter (nm) 176 + 58 278 + 55 306 + 57 177 + 121
Table 1 shows the antioxidant activities of several composite nanofibers of PVP/β-carotene with varying concentrations of PVP. The composite nanofibers mat made from a precursor solution of PVP 10 wt.% loaded by 1 wt.% β-carotene, which has the largest diameter and the uniform size distribution, gave the largest antioxidant activity. Having the largest diameter and the uniform size distribution, the composite nanofibers could well protect the β-carotene during storage hence keeping its antioxidant properties undamaged. Nevertheless, it was proven that β-carotene still retained its antioxidant activities although being encapsulated in the nanofibers for all diameters. 4. Conclusion Poly(vinylpyrrolidone)/PVP composite nanofiber mats loaded by β-carotene has successfully been produced through electrospinning. From the SEM images, the increase in PVP concentration generally increased the diameter of nanofibers except when the concentration was too high due to the appearance of multi-jets. The average diameters of PVP fibers loaded by βcarotene were ranged from 176 to 306 nm. From the FTIR characterization, it was found that the β-carotene caused a significant shift of the peak at 1440 cm-1 to lower wavenumbers of the PVP as a result of molecular interaction between β-carotene and PVP. From the antioxidant test, the β-carotene still retained its antioxidant activity although being encapsulated in the PVP nanofibers. The nanofibers composite of PVP 10 wt.% loaded 1 wt.% β-carotene gave the best antioxidant activity. Acknowledgements This research was financially supported by Directorate of Research and Community Engagement of Ministry of Research, Technology and Higher Education, the Republic of Indonesia under the University’s Excellence Research (PUPT) Grant in the fiscal years 2015-2016 and the “Riset KK ITB” Grant in the fiscal year 2015-2016. References [1] Agarwal S, Rao AV. Carotenoids and chronic diseases. Drug Metabol Drug Interact 2000;17:189-210. [2] Stahl W, Sies H. Bioactivity and protective effects of natural carotenoids. BBA – Mol Basis Dis 2005;1740:101-7. [3] McClements DJ, Decker EA, Park Y, Weiss J. Structural design principles for delivery of bioactive components in nutraceuticals and functional foods. Crit Rev Food Sci Nutr 2009;49:577-606. [4] Jyothi NV, Prasanna PM, Sakarkar SN, Prabha KS, Ramaiah PS, Srawan GY. Microencapsulation techniques, Factors influencing encapsulation efficiency. J Microencapsulation 2010;27:187-97. [5] Fernandez A, Torres-Giner S, Lagaron JM. Novel route to stabilization of bioactive antioxidants by encapsulation in electrospun fibers of zein prolamine. Food Hydrocoll 2009;23:1427-32. [6] Zômpero RHF, López-Rubio A, de Pinho SC, Lagaron JM, de la Torre LG. Hybrid encapsulation structures based on β-carotene-loaded nanoliposomes within electrospun fibers. Colloids Surf B 2015;134:475-82. [7] Peinado I, Mason M, Romano A, Biasioli F, Scampicchio M. Stability of β-carotene in polyethylene oxide electrospun nanofibers. App Surf Sci 2016;370:111-6. [8] Munir MM, Iskandar F, Khairurrijal, Okuyama K. High performance electrospinning system for fabricating highly uniform polymer nanofibers. Rev Sci Instrum 2009;80:026106. [9] Munir MM, Iskandar F, Khairurrijal, Okuyama K. A constant-current electrospinning system for production of high quality nanofibers. Rev Sci Instrum 2008;79:093904. [10] Munir MM, Suryamas AB, Iskandar F, Okuyama K. Scaling law on particle-to-fiber formation during electrospinning. Polym 2009;50:4935-43. [11] Fischer F, Bauer S. Polyvinylpyrrolidon. Chem Unserer Zeit 2009;43:376-83. [12] Vongsetskul T, Wongsomboon TCP, Rangkupan R. Effect of solvent and processing parameters on electrospun polyvinylpyrrolidone ultra-fine fibers. Chiang Mai J Sci 2015;42:436-42. [13] Miao J, Miyauchi M, Dordick JS, Linhardt RJ. Preparation and characterization of electrospun core sheath nanofibers from multi-walled carbon nanotubes and poly(vinyl pyrrolidone). J Nanosci Nanotechnol 2012;12:2387-93.
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23 [14] Suwantong O, Pankongadisak P, Deachathai S, Supaphol P. Electrospun poly(l-lactic acid) fiber mats containing crude Garcinia mangostana extracts for use as wound dressing. Polym Bull 2014;71:925-49. [15] Shenoy SL, Bates WD, Frisch HL, Wnek GE. Role of chain entanglements on fiber formation during electrospinning of polymer solutions: good solvent, non-spesific polymer-polymer interaction limit. J Polym 2005;46:3372-84. [16] Andrady AL. Science and technology of polymer nanofibers. USA: John Wiley & Sons; 2008. [17] Nuryantini AY, Edikresnha D, Munir MM, Khairurrijal, Electrospun polyvinylpyrrolidone as a carrier for leaves extracts of Anredera cordifolia (Ten.) steenis. Mater Sci Forum 2015;827:91-4. [18] Ammawath W, Man YbC. A rapid method for determination of commercial β-carotene in RBD palm olein by Fourier transform infrared spectroscopy. As J Food Agr-Ind 2010;3:443-52. [19] Laot CM, Marand E, Oyama HT. Spectroscopic characterization of molecular interdiffusion at a poly(vinyl pyrrolidone)/vinyl ester interface. J Polym 1999;40:1095-108. [20] Saroj AL, Singh RK, Chandra S. Studies on polymer electrolyte, poly(vinyl) pyrrolidone (PVP) complexed with ionic liquid: Effect of complexation on thermal stability, conductivity and relaxation behavior. Mater Sci Eng B 2013;178:231-8.
23
ScienceDirect Procedia Engineering 170 (2017) 19 – 23
Engineering Physics International Conference, EPIC 2016
Encapsulation of β-carotene in poly(vinylpyrrolidone) (PVP) by electrospinning technique Rhyan Prayuddy Reksamunandara,b, Dhewa Edikresnhaa,b, Muhammad Miftahul Munira,b, Sophi Damayantic, and Khairurrijala,b,* a
Department of Physics Faculty of Mathematics and Natural Sciences, b Research Center for Bioscience and Biotechnology, c Department of Pharmacochemistry, School of Pharmacy, Institut Teknologi Bandung, Jalan Ganesa 10, Bandung 40132, Indonesia
Abstract β-carotene, which is the most commonly known type of carotenoids, has been proven to have antioxidant properties and to provide provitamin A activity that are important for human health. In this study, β-carotene was encapsulated into poly(vinylpyrrolidone)/PVP through electrospinning technique in order to protect β-carotene from degradation and to maintain the antioxidant properties. The precursor solution was prepared by dissolving β-carotene (1 wt.%) to various concentrations of PVP (6 wt.%, 8 wt.%, 10 wt.%, and 12 wt.%) in ethanol. SEM images showed the formation of nanoscale beaded fibers with generally increasing average diameter when the PVP concentration was increased except when the concentration of precursor solution was 12 wt.% PVP. In this case, it became too viscous and caused clogging on the needle tip and the formation of multi-jets. The average diameters of PVP/β-carotene composite nanofibers were in the range of 176 to 306 nm. The analyses of Fourier Transform Infrared (FTIR) spectra proved the presence of structural changes in PVP nanofibers after being loaded by β-carotene as a result of molecular interaction between β-carotene and PVP. This was given by a significant shift of the peak at 1440 cm-1 to lower wavenumbers of the FTIR spectrum of PVP. The β-carotene was proven to retain their antioxidant ability even after being encapsulated into the PVP nanofibers as tested by the DPPH assay method. © 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). © 2016 The Authors. Published by Elsevier Ltd. Peer-review under responsibility of the organizing committee of the Engineering Physics International Conference 2016 Keywords: β-carotene; poly(vinylpyrrolidone); electrospinning; antioxidant.
1. Introduction Carotenoids are natural pigments that are abundant in nature and available in various types of fruits, vegetables, leaves and few animals. Carotenoids provide provitamin A activity that contributes to human health since they have antioxidant properties, such as to protect cells against reactive oxygen and free radicals, to reduce the risk of cancer or other chronic diseases, to stimulate body growth, increase immune system and improve visual function [1-2]. The most commonly known of carotenoid is β-carotene [3]. β-carotene is a non-polar compound with a long chain of conjugated double bonds and unsaturated structures which cause β-carotene to be easily degraded and is highly sensitive to light and heat [4]. To stabilize β-carotene from degradation, encapsulation through electrospinning technique has been studied [5-7]. However, the number of studies on βcarotene being encapsulated by electrospinning is still rare and no detailed explanation is given regarding the effect of polymer concentration on the average diameter of composite fibers and the antioxidant properties of the β-carotene after being encapsulated. Hence, this study investigated the effect of varying the polymer concentration towards the diameter and the antioxidant properties of β-carotene.
* Corresponding author. Tel.: +62-22-250 0834; fax: +62-22-250 6452. E-mail address: [email protected]
1877-7058 © 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Peer-review under responsibility of the organizing committee of the Engineering Physics International Conference 2016
doi:10.1016/j.proeng.2017.03.004
20
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23
Electrospinning is a technique to produce the polymer fibers or their composite with high surface area to volume ratio. Electrospinning system has been designed and built to produce nanofibers with relatively narrow size distribution [8-10]. In this study, a biocompatible polymer, poly(vinylpyrrolidone) (PVP), was used as the polymer matrix with ethanol as a solvent for the precursor solution. PVP was chosen due to its acceptable level of toxicity, biocompatibility and solubility both in nonpolar and polar solvents [11]. The electrospinnability of PVP in ethanol solvent was already proven in the previous studies [12, 13]. This study reports the formation of composite nanofibers between PVP and β-carotene known for its antioxidant properties by means of electrospinning to develop an encapsulating technology for β-carotene. The morphology, the chemical compounds, and the antioxidant ability of the composite fibers will be described and discussed. 2. Method PVP (poly(vinylpyrrolidone, MW = 1300 kg/mol) and β-carotene were purchased from Sigma-Aldrich, whereas ethanol was provided by a local supplier. The polymer solution was made by mixing PVP in ethanol as the solvent in various ratios of PVP (6 wt.%, 8 wt.%, 10 wt.%, and 12 wt.%) together with 1 wt.% β-carotene. All components were then magnetically stirred for three hours at room temperature until the solution was mixed homogeneously. To obtain nanofibers mats of PVP/β-carotene, each precursor solution was inserted into a single needle syringe with inner needle diameter of 0.84 mm. Each solution was electrospun using a drum collector in electrospinning apparatus with the distance between the needle and the collector was set to be 10 cm, the flow rate used was 1 mL/h, and the applied voltage was 15 kV. The fibers were collected on an aluminum foil that was coated on the drum collector. The mats produced were kept at the temperature of 5 to 6 oC and were protected from light until being used for characterizations and antioxidant test. The effects of β-carotene loaded in variations of PVP solution to the morphology of the nanofibers and the diameter size distribution were observed using Scanning Electron Microscope (SEM) (JEOL, JCM-6000 NeoScope Benchtop) with an accelerating voltage of 15 kV. The infrared spectra of the nanofibers were analyzed by Fourier Transform Infrared (FTIR) spectrometer (Bruker Alpha, A220/D-01) to demonstrate the changes in the structure of PVP/β-carotene nanofibers. The wavenumber spectrum used for the FTIR test was within the range of 500-4000/cm-1. The antioxidant ability of the PVP/βcarotene nanofibers was tested using 1,1-diphenyl-2-picrylhydrazil (DPPH) assay method [14]. The level of antioxidant of the composite fibers can be determined by Equation (1).
% AA =
Acontrol − Asample Acontrol
(1)
×100
where Acontrol is the absorbance of the 25 ppm DPPH solution without PVP/β-carotene fibers and Asample is the absorbance of the 50 ppm DPPH solution mixed with 50 ppm PVP/β-carotene fibers in ethanol with a volume ratio of 1:1 after being incubated for 30 minutes in dark condition. The absorbance spectrum was measured using UV-Visible Spectrophotometer (Agilent, 8453) and monitored at the wavelength of 515 nm which is the peak level of the DPPH solution [14]. 3. Results and Discussion 3.1. Morphology of the composite nanofibers The morphologies of the PVP/β-carotene composite fibers are given by the SEM images with a magnification of 4000 times in Figure 1. They agreed well to those obtained from the previous studies [12, 13], but with some beads. The presence of beads was related to the solution viscosity, entanglement number of solution, and the additional substance inside the fibers [15]. Beads can be predicted to occur whenever the surface tension exceeds the charge repulsive force and viscoelastic force that support the elongation of continuous jet [16].
Fig. 1. The SEM images of the composite nanofibers of (a) 6 wt.% PVP/1 wt.% β-carotene, (b) 8 wt.% PVP/1 wt.% β-carotene, (c) 10 wt.% PVP/1 wt.% βcarotene, and (d) 12 wt.% PVP/1% β-carotene.
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23
Figure 2 gives the fibers diameter distributions of the PVP/β-carotene composite fibers for (a) 6 wt.% PVP/1 wt.% βcarotene, (b) 8 wt.% PVP/1 wt.% β-carotene, (c) 10 wt.% PVP/1 wt.% β-carotene, and (d) 12% PVP/1% β-carotene. It was found that the average diameter of fibers increases with the increase of polymer concentration. This finding can be explained as follows: a higher polymer concentration in the precursor solution will increase the chain entanglement and the amount of readily available polymer to be electrospun, hence the diameter of fiber will be larger [16]. Additionally, the increase of polymer concentration (thus lowering the wt.% of the solvent) causes the solvent to dry up faster and less time for the repulsive columbic force to stretch the fiber, the diameter of fiber will be larger [12]. However, the composite nanofibers obtained from the 12 wt.% PVP/1% β-carotene had uneven fibers diameter distribution and smaller average diameter of fibers. An explanation of this finding is as follows: the back and side parts of the liquid stream coming out from the needle at a later phase can be forced to be ejected as small side jets when the applied voltage is high enough [17]. A special case happened for the high composition of polymer (higher than 12 wt.% PVP) in which the tip blockage occurred. This is due to the rapid rate of solvent evaporation so that the liquid stream solidifies quickly while the syringe pump still pushes the liquid forward [16].
Fig. 2. The corresponding bar diagrams of fibers diameters and count rate of (a) 6 wt.% PVP/1 wt.% β-carotene, (b) 8 wt.% PVP/1 wt.% β-carotene, (c) 10 wt.% PVP/1 wt.% β-carotene, and (d) 12 wt.% PVP/1% β-carotene.
3.2. FTIR spectroscopy Figure 3 shows the FTIR spectra of β-carotene, PVP, and PVP/β-carotene. In the FTIR spectrum of β-carotene given in Fig. 3(a), the broad peak at 3411 cm-1 represents the presence of O-H stretching of the hydroxyl group, which is likely due to the interaction of β-carotene with oxygen in the air [5]. The peaks at 2929 cm-1 and 2869 cm-1 indicate the CH2 asymmetry and symmetry stretching, respectively. The peaks at 1717 cm-1 and 1366 cm-1 exhibit the presence of carbonyl groups and stretching symmetry of C-H bond group, respectively. The sharp peak at 965 cm-1 marks the deformation mode of trans-conjugate-alkenes as the specific areas of trans=CH used for identification of β-carotene [18].
Fig. 3. FTIR spectra of (a) β-carotene, (b) PVP, (c) 6 wt.% PVP/1 wt.% % β-carotene, (d) 8 wt.% PVP/1 wt.% β-carotene, (e) 10 wt.% PVP/1 wt.% β-carotene, and (f) 12 wt.% PVP/1 wt.% β-carotene.
In the FTIR spectrum of PVP given in Fig. 3(b), the broad band around 3550 - 3200 cm-1 assign as O-H stretching of the hydroxyl group. The presence of heteroatomic molecules and carbonyl groups in the pyrrolidone ring of PVP was identified by the sharp peak at 1656 cm-1 as the signature of C=O stretching and the peak at 1291 cm-1 is related to CN stretch of amide group [19, 20]. The peak at 1440 cm-1 assign as the C-H deformation of CH2 group. The FTIR spectra of PVP/β-carotene in Figs. 3(c)3(d) show a significant shift of the peak at 1440 cm-1 to lower wavenumbers due to the encapsulation of β-carotene in PVP fibers. On the other hand, the broad band (3550 - 3200) cm-1 and the peak at 1656 cm-1 did not shift significantly and the sharp peak at 965 cm-1 disappeared.
21
22
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23
3.3. Antioxidant activity Since β-carotene is already known to have antioxidant properties, this study was intended to find out the effectiveness of the antioxidant property of β-carotene after being encapsulated into the composite nanofibers mats. The antioxidant test was done four days after the production of the composite nanofibers mats. The β-carotene-encapsulated nanofibers mats were mixed with DPPH, a notorious free radical usually used to measure the antioxidant properties of any sample. DPPH is typically violet in color and becomes pale yellow when neutralized [14]. Table 1. Antioxidant activity of PVP/β-carotene composite nanofibers.
Composite nanofibers PVP 6 wt.% loaded 1 wt.% β-carotene PVP 8 wt.% loaded 1 wt.% β-carotene PVP 10 wt.% loaded 1 wt.% β-carotene PVP 12 wt.% loaded 1 wt.% β-carotene
Antioxidant activity (%) 47 + 20 47 + 30 86 + 1.2 21 + 12
Average diameter (nm) 176 + 58 278 + 55 306 + 57 177 + 121
Table 1 shows the antioxidant activities of several composite nanofibers of PVP/β-carotene with varying concentrations of PVP. The composite nanofibers mat made from a precursor solution of PVP 10 wt.% loaded by 1 wt.% β-carotene, which has the largest diameter and the uniform size distribution, gave the largest antioxidant activity. Having the largest diameter and the uniform size distribution, the composite nanofibers could well protect the β-carotene during storage hence keeping its antioxidant properties undamaged. Nevertheless, it was proven that β-carotene still retained its antioxidant activities although being encapsulated in the nanofibers for all diameters. 4. Conclusion Poly(vinylpyrrolidone)/PVP composite nanofiber mats loaded by β-carotene has successfully been produced through electrospinning. From the SEM images, the increase in PVP concentration generally increased the diameter of nanofibers except when the concentration was too high due to the appearance of multi-jets. The average diameters of PVP fibers loaded by βcarotene were ranged from 176 to 306 nm. From the FTIR characterization, it was found that the β-carotene caused a significant shift of the peak at 1440 cm-1 to lower wavenumbers of the PVP as a result of molecular interaction between β-carotene and PVP. From the antioxidant test, the β-carotene still retained its antioxidant activity although being encapsulated in the PVP nanofibers. The nanofibers composite of PVP 10 wt.% loaded 1 wt.% β-carotene gave the best antioxidant activity. Acknowledgements This research was financially supported by Directorate of Research and Community Engagement of Ministry of Research, Technology and Higher Education, the Republic of Indonesia under the University’s Excellence Research (PUPT) Grant in the fiscal years 2015-2016 and the “Riset KK ITB” Grant in the fiscal year 2015-2016. References [1] Agarwal S, Rao AV. Carotenoids and chronic diseases. Drug Metabol Drug Interact 2000;17:189-210. [2] Stahl W, Sies H. Bioactivity and protective effects of natural carotenoids. BBA – Mol Basis Dis 2005;1740:101-7. [3] McClements DJ, Decker EA, Park Y, Weiss J. Structural design principles for delivery of bioactive components in nutraceuticals and functional foods. Crit Rev Food Sci Nutr 2009;49:577-606. [4] Jyothi NV, Prasanna PM, Sakarkar SN, Prabha KS, Ramaiah PS, Srawan GY. Microencapsulation techniques, Factors influencing encapsulation efficiency. J Microencapsulation 2010;27:187-97. [5] Fernandez A, Torres-Giner S, Lagaron JM. Novel route to stabilization of bioactive antioxidants by encapsulation in electrospun fibers of zein prolamine. Food Hydrocoll 2009;23:1427-32. [6] Zômpero RHF, López-Rubio A, de Pinho SC, Lagaron JM, de la Torre LG. Hybrid encapsulation structures based on β-carotene-loaded nanoliposomes within electrospun fibers. Colloids Surf B 2015;134:475-82. [7] Peinado I, Mason M, Romano A, Biasioli F, Scampicchio M. Stability of β-carotene in polyethylene oxide electrospun nanofibers. App Surf Sci 2016;370:111-6. [8] Munir MM, Iskandar F, Khairurrijal, Okuyama K. High performance electrospinning system for fabricating highly uniform polymer nanofibers. Rev Sci Instrum 2009;80:026106. [9] Munir MM, Iskandar F, Khairurrijal, Okuyama K. A constant-current electrospinning system for production of high quality nanofibers. Rev Sci Instrum 2008;79:093904. [10] Munir MM, Suryamas AB, Iskandar F, Okuyama K. Scaling law on particle-to-fiber formation during electrospinning. Polym 2009;50:4935-43. [11] Fischer F, Bauer S. Polyvinylpyrrolidon. Chem Unserer Zeit 2009;43:376-83. [12] Vongsetskul T, Wongsomboon TCP, Rangkupan R. Effect of solvent and processing parameters on electrospun polyvinylpyrrolidone ultra-fine fibers. Chiang Mai J Sci 2015;42:436-42. [13] Miao J, Miyauchi M, Dordick JS, Linhardt RJ. Preparation and characterization of electrospun core sheath nanofibers from multi-walled carbon nanotubes and poly(vinyl pyrrolidone). J Nanosci Nanotechnol 2012;12:2387-93.
Rhyan Prayuddy Reksamunandar et al. / Procedia Engineering 170 (2017) 19 – 23 [14] Suwantong O, Pankongadisak P, Deachathai S, Supaphol P. Electrospun poly(l-lactic acid) fiber mats containing crude Garcinia mangostana extracts for use as wound dressing. Polym Bull 2014;71:925-49. [15] Shenoy SL, Bates WD, Frisch HL, Wnek GE. Role of chain entanglements on fiber formation during electrospinning of polymer solutions: good solvent, non-spesific polymer-polymer interaction limit. J Polym 2005;46:3372-84. [16] Andrady AL. Science and technology of polymer nanofibers. USA: John Wiley & Sons; 2008. [17] Nuryantini AY, Edikresnha D, Munir MM, Khairurrijal, Electrospun polyvinylpyrrolidone as a carrier for leaves extracts of Anredera cordifolia (Ten.) steenis. Mater Sci Forum 2015;827:91-4. [18] Ammawath W, Man YbC. A rapid method for determination of commercial β-carotene in RBD palm olein by Fourier transform infrared spectroscopy. As J Food Agr-Ind 2010;3:443-52. [19] Laot CM, Marand E, Oyama HT. Spectroscopic characterization of molecular interdiffusion at a poly(vinyl pyrrolidone)/vinyl ester interface. J Polym 1999;40:1095-108. [20] Saroj AL, Singh RK, Chandra S. Studies on polymer electrolyte, poly(vinyl) pyrrolidone (PVP) complexed with ionic liquid: Effect of complexation on thermal stability, conductivity and relaxation behavior. Mater Sci Eng B 2013;178:231-8.
23