Controlled Release of Retinyl Acetate from β-Cyclodextrin

Article pubs.acs.org/JAFC

Controlled Release of Retinyl Acetate from β‑Cyclodextrin Functionalized Poly(vinyl alcohol) Electrospun Nanofibers Solomon M. Lemma,†,‡,∥ Matteo Scampicchio,† Peter J. Mahon,‡ Igor Sbarski,§ James Wang,§,∥ and Peter Kingshott*,†,§ †

Faculty of Science and Technology, Free University of Bolzano, Piazza Università 5, 39100 Bolzano, Italy Department of Chemistry and Biotechnology, Swinburne University of Technology, Hawthorn, VIC 3122, Australia § Department of Mechanical and Product Design Engineering, Swinburne University of Technology, Hawthorn, VIC 3122, Australia ∥ Industrial Research Institute Swinburne (IRIS), Swinburne University of Technology, Hawthorn, VIC 3122, Australia ‡

ABSTRACT: Retinyl acetate (RA) was effectively incorporated into electrospun nanofibers of poly(vinyl alcohol) (PVA) containing β-cyclodextrin (β-CD) in order to form inclusion complexes for encapsulation to prolong shelf life and thermal stability. The physical and thermal properties of encapsulated RA were determined by scanning electron microscopy (SEM), Xray diffraction (XRD), and differential scanning calorimetry (DSC). The nanofibers of PVA/RA and PVA/RA/β-CD exhibited bead free average fiber diameters of 264 ± 61 and 223 ± 49 nm, respectively. The surface chemistry of the functional nanofibers was investigated by X-ray photoelectron spectroscopy (XPS). Thermogravimetric analysis (TGA) demonstrated different thermal stabilities between the bioactive and the polymer, with and without β-CD. Square-wave voltammogram peak current changes were used to follow the release kinetics of RA from the nanofibers. Results indicate that RA coated inside PVA/β-CD nanofibers was protected against oxidation much better than RA in PVA nanofibers and should extend the shelf life. In addition, RA encapsulated in the PVA/β-CD had better thermal stability than PVA nanofibers. KEYWORDS: electrospinning, retinyl acetate, nanofiber, cyclodextrin, poly(vinyl alcohol) (PVA), X-ray photoelectron spectroscopy (XPS)

■

units.10 The nature of guest outer surfaces and host inner cavities of β-CD provides a unique feature enabling inclusion complex formation with encapsulated ingredients into a cavity of suitable and stable geometrical size.11,12 The complex formed in the inner cavities of β-CD has been investigated widely for protection of the core component with bioactives,13 enzymes,14 essential oils, and flavors to improve storage conditions and extend shelf-life.15,16 These studies confirmed that the thermal stability and shelf life were improved due to the efficiency of the core, specifically using β-CD for inclusion complexation.17,18 Recently, electrospinning has drawn great interest for the production of micro- and nanofibers with unique physicochemical properties, such as large surface area to volume ratio and multiple surface functionalities, which lead to wide food processing applications.19 An important number of applications have reported the use of electrospun nanofibers in food encapsulation and packaging,20,21 sensor modification for bioactives,22 and juice clarification and filtration.23 Consequently, research into the encapsulation of bioactives has gained attention to develop adequate delivery matrixes and product formulations. Particularly, β-CD-IC and aqueous soluble poly(vinyl alcohol) (PVA) polymeric functionalized nanofibers have been investigated and reported for the application of encapsulation due to its efficient and effective

INTRODUCTION Vitamin A and its derivatives are found naturally in food substances, used as food supplements and as an antioxidant in the food and pharmaceutical industries. A number of studies have been conducted to understand the tremendous importance of vitamin A and its derivatives for food processing and biological activities.1,2 Among the many derivatives, retinyl acetate (RA) is one of the most commonly used in food and nutraceutical products.3,4 However, retinyl acetate is very sensitive to oxidation, light, and high temperature processing.5,6 In order to overcome these problems, strategies involving preserving materials to maintain stability and natural characteristics for a long shelf life are an area of interest. Several encapsulation techniques have been studied to increase the stability of retinyl acetate against these potential degrading factors.7 Among the most dominant, spray drying and extrusions have been used in a wide range of encapsulation processes that use different mechanisms.6,7 A number of factors can impact the efficiency of encapsulation using these methods that limits their use in a wider range of applications. They are mainly related to high processing temperatures and shear force changes.8,9 Encapsulating vitamin A (RA) into cyclodextrins offers a number of potential advantages for delivery, besides overcoming the expensive cost-in-use and complexity of these technologies, including protecting the organoleptic characteristics of vitamins based on molecular selectivity.9 Cyclodextrins (CDs) are one of the most important cyclic oligosaccharides comprising different numbers of D-glucose units, with β-cyclodextrin (β-CD) having seven glucopyranose © XXXX American Chemical Society

Received: January 8, 2015 Revised: March 16, 2015 Accepted: March 16, 2015

A

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry Table 1. Viscosity, Electrical Conductivity of Solutions, and Average Fiber Diameters of Electrospun Nanofibers solutions

% PVAa (w/v)

PVA PVA/β-CD PVA/RA PVA/β-CD/RA

12 10 10 10

% β-CDb (w/w)

% RA (w/w)

30 30

10 10

viscosityc (Pa·s) 0.62 0.78 0.94 0.87

± ± ± ±

0.04x 0.06y 0.16z 0.08y

conductivityc (mS/cm) 1.76 1.76 1.52 1.52

± ± ± ±

0.02x 0.07x 0.09y 0.04y

AFDc (nm) 266 282 264 223

± ± ± ±

59x 70y 61w 49z

With respect to solvent (water). bWith respect to polymer (PVA). cMean ± SD. The mean values with the same letter in a column are not significantly different (P > 0.05) at 95% confidence interval.

a

The viscosity of the solutions was measured with an AR2000 Advanced Rheometer (4 cm, 2° cone, TA Instruments, Delaware, USA) at 23 °C with a shear rate from 1 up to 1000 s−1. The morphology and the fiber diameter of the electrospun nanofibers were analyzed by a field-emission scanning electron microscope (FE-SEM, Zeiss Supra 40 VP, Oberkochen, Germany). The nanofiber samples were coated with 15 nm of Au (EMITECH K975X) prior to SEM imaging. The average fiber diameter (AFD) for the samples was calculated by analysis of 100 fibers from the SEM images. The X-ray diffraction (XRD) patterns of powder PVA, β-CD, RA, and nanofiber membranes were obtained with a Bruker D8 Advance XRD with Cu Kα radiation at a scanning rate of 1°/min over a 2θ range of 5−40°. Thermal properties of the samples were investigated by using a differential scanning calorimeter (DSC) and a thermogravimetric analyzer (TGA) (DSC-TGA-2960 SDT V3.0F, TA Instruments Inc., Delaware, USA). For DSC analyses, the sealed 5 mg samples were initially equilibrated at 10 °C and then heated to 300 °C at a rate of 10 °C/min using nitrogen as a purge gas. TGA measurements were performed for electrospun nanofibers after 1 day of storage. The TGA data for 10 mg samples were recorded from room temperature to 600 °C at a heating rate of 10 °C/min, under a nitrogen atmosphere. X-ray photoelectron spectroscopy (XPS) (Kratos Axis Nova Analytical Ltd., Manchester, U.K.) was used to determine the surface chemistry of the nanofiber membranes. A monochromated Al Kα X-ray source was used at a power of 150 W. Elements were identified from survey spectra (0−1200 eV) with a detector pass energy of 160 eV. The atomic concentrations of the detected elements were calculated using integral peak intensities and the sensitivity factors supplied by the manufacturer. Electrochemical Measurements. Electrochemical experiments were performed with a CH Instruments Model CH610 Electrochemical Analyzer. The electrochemical cell consisted of a threeelectrode system: a glassy carbon (GC) working electrode (BASi MF2012), a platinum wire (BASi MW-4130) auxiliary electrode, and a Ag/AgNO3 reference electrode filled with 100 mM TBAP−acetonitrile (BASi MF-2062). The working electrode was cleaned with a polishing cloth (Buehler, Lake Bluff, IL, 60044, USA) impregnated with a 0.05 μm alumina slurry until a mirror-like surface was obtained. The polished electrodes were sonicated in acetonitrile to remove any alumina. Sample Preparation. The nanofibers of PVA/RA and PVA/RA/ β-CD-IC were measured frequently to determine the remaining retinyl acetate against the storage time. These nanofiber membranes were stored at room temperature in the laboratory for a certain period of time prior to analyses. Sample solutions of each type of nanofiber were prepared by soaking 40 mg of nanofibers in ethanol with sonication for 30 min at 25 °C to release the retinyl acetate into the solution. The release rates of retinyl acetate from the nanofibers were monitored by square-wave voltammetry (SWV) using 0.1 M tetrabutyl ammonium hexafluorophosphate (TBAPF6) in acetonitrile at 25 °C. The SWV conditions were as follows: step potential, 4 mV; amplitude, 25 mV; frequency, 15 Hz. The amount of retinyl acetate present in the nanofibers was monitored using a scanning potential range from 0 to 0.9 V. To perform the kinetic analysis, the signal obtained from each voltammogram was baseline subtracted and the value of the peak current (Ip) with the corresponding storage day was used for the subsequent analyses. The exact concentration of the released retinyl acetate was obtained using a calibration graph of retinyl acetate standard. The released amount was determined according to the

protection for active food components in the core from thermal, light, and other storage condition constraints.16,18 In this study, PVA and β-CD were chosen as the base material and RA as a core bioactive. The nanofiber encapsulation based on PVA has been extensively reported as a protecting coat for food materials.16,18,24 The aim of this study was to use the electrospinning technique in order to develop a PVA/RA/β-CD nanofiber functional matrix that provides support of thermal stability and long shelf life of RA. The RA was incorporated separately into the PVA nanofibers for assessing the relation between the inclusion complex and the variation of the retained core. Furthermore, the RA in the nanofibers demonstrated the effect and influence on the morphology of the fibers and also determined release kinetics as a function of fiber material composition under room storage conditions.

■

MATERIALS AND METHODS

Materials. Poly(vinyl alcohol) (PVA) (Mw: 85 000−124 000, 99+ % hydrolyzed), retinyl acetate (RA), β-cyclodextrin (β-CD), and tetrabutyl ammonium hexafluorophosphate were purchased from Sigma-Aldrich (Castle-Hill, NSW 1765, Australia). Ethanol (100% proof) and acetonitrile (99.8%) were obtained from Chem-Supply (Gillman, SA 5013, Australia). All of the reagents used were of analytical grade and used as received. Milli-Q water was used throughout the experiments. Preparation of the Solutions. The powdered PVA was dissolved in Milli-Q water at 75 °C using a magnetic stirrer (1 krpm) for 2 h to make up a 12% (w/v) solution.12 In the same method, 10% (w/v) PVA solution was prepared containing a different concentration of retinyl acetate and β-CD inclusion complex. To this solution, 30% (w/ w, with respect to polymer PVA) of β-CD was added at 75 °C with constant stirring for 10 min. Subsequently, the solution was cooled to 25 °C with stirring and 10% (w/w, respective to PVA) of retinyl acetate powder was added to make the PVA/retinyl acetate/β-CD solution. The solutions were stirred for a further 1 h at 25 °C prior to electrospinning. Homogeneous and clear solutions were obtained for PVA and PVA/β-CD compositions. In contrast, these solutions turned turbid by addition of retinyl acetate. Finally, the prepared solution of PVA and composition with retinyl acetate and β-CD were electrospun separately at room temperature. The composition and rheological properties of the solutions are summarized in Table 1. Electrospinning. The prepared solutions were placed in a 5 mL syringe connected with a stainless steel needle having an inner diameter of 0.5 mm. The solution in the syringe was pumped at 200 μL/h horizontally (model: KDS Legato 111, KD Scientific, Holliston, MA, USA), and the solutions were electrospun with an applied voltage of 18 kV using a homemade high voltage power supply. Then, the nanofibers were deposited on a grounded stationary metal collector covered with aluminum foil at 20 cm from the syringe nozzle to the tip. The electrospinning experiments were carried out at room temperature in an enclosed Plexiglas box. Measurements and Characterization. Ionic conductivities of the polymer solutions were measured using a conductivity meter (METTLER TOLED: S230, Zurich, Switzerland) that was calibrated with 1413 μS/cm standard conductivity solution at room temperature. B

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry

Figure 1. SEM images and fiber diameter distributions of the electrospun nanofibers obtained from (a) PVA, (b) PVA/β-CD, (c) PVA/RA, and (d) PVA/β-CD/RA. The insets show the high magnification images. The fiber diameter distributions of each are shown on the right. decrease of peak current with respect to storage time. The release (%) is expressed by the equation

Release (%) = 100 ×

of the PVA and PVA/β-CD nanofibers; instead, the diameters of the nanofibers decreased slightly (Figure 1). Similar findings were observed in the range between 200 and 360 nm in other studies when PVA and PVA/β-CD were used for encapsulation of food flavors.13,18 For comparison, both the PVA and PVA/βCD nanofibers were produced in the same concentration solutions as fibers which have RA. In the electrospinning, uniform nanofibers were obtained and no huge variation was encountered along the electrospun fibers in the whole sample. The representative SEM images and the fiber diameter distribution along with the average fiber diameter of PVA, PVA/β-CD-IC, PVA/RA, and PVA/RA/β-CD-IC nanofibers are depicted in Figure 1. The SEM images of electrospun nanofibers were obtained from homogeneous solutions of 12% (w/v) PVA in Milli-Q water and 30% (w/ w) β-CD and 10% (w/w) RA with respect to PVA. Uniform nanofibers were obtained from all the solutions of the mixture. In the case of PVA/RA/β-CD-IC, nanofibers were mostly uniform and had smaller diameters. The fibers were also obtained without any aggregates of retinyl acetate on their surfaces, implying that RA was uniformly incorporated within the fibers. The result showed that incorporation of RA in the electrospun PVA and PVA/β-CD fibers had little effect upon the morphology. The average fiber diameter with the RA had smaller diameters compared to controls (Table 1 and Figure 1). The X-ray diffraction pattern measurements were carried out in order to elucidate the form of crystalline aggregates in the nanofibers and the crystalline nature of the pure powder PVA samples. The nanofibers with samples of PVA, RA, and β-CD

Ip(Day k) − Ip(Day k + t ) Ip(Day k)

where Ip(Day k) is the peak current for an earlier storage time compared to the peak current Ip(Day k + t) with longer storage time. Statistical Analysis. All data were expressed as mean ± standard deviation. Statistical significance by analysis of variation (ANOVA) was used to determine the difference among samples’ viscosity, conductivity, and AFD, P < 0.05.

■

RESULTS AND DISCUSSION Measurements and Crystalline Structure. The PVA solutions containing retinyl acetate and retinyl acetate/β-CDIC were electrospun from the aqueous solution mixture. The conductivity and shear viscosity of the PVA and PVA/β-CD with and without retinyl acetate solutions are shown in Table 1. The addition of 10% (w/w) RA to the PVA and PVA/β-CD solutions significantly affects the viscosity and the conductivity of the solutions, as shown in Table 1. The viscosity of PVA/βCD has a higher value compared to the PVA. In the same manner, the viscosity of PVA/RA has the highest value compared to all the rest of the samples. In general, the conductivities of all the sample solutions are significantly different in the presence of RA (P < 0.05) and the mean fiber diameters also are significantly different as the compositions of the polymer solutions vary, as shown in Table 1. The results showed that the addition of RA did not change the morphology C

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry

the PVA nanofiber matrix. In these spectra, the existence of RA is not apparent in the nanofibers of PVA/RA/β-CD and this implies that the RA incorporates in the form of RA/β-CD-ICs in the PVA nanofiber matrix. The existence of RA in the nanofibers of PVA/RA and PVA/RA/β-CD-IC affected the crystallization behavior of the nanofibers. This may need further study as to the reasons why RA has a greater tendency to alter the crystal structure of the final products. The absence of the peak for RA in the PVA/RA/β-CD nanofibers confirmed that the RA dispersed homogeneously in the PVA matrix and is deprived of the formation of crystal aggregates. Thermal Analyses. The thermal stability was determined by thermogravimetric analysis in a temperature range of 25 °C up to 600 °C, as shown in Figure 3 for the TGA and DTGA curves. Thermogravimetric analysis revealed the thermal effects of PVA nanofibers with the presence of β-CD and RA at different concentrations. Figure 3a shows TGA curves of the PVA powder and the PVA nanofibers, with and without the composite β-CD and RA. All the samples exhibited three main weight loss steps. The first weight loss from 5 to 10%, which covers the temperature range of about 25−230 °C, corresponds to weakly physisorbed water (indicated in the inset of Figure 3a), while the loss in the range 230−400 °C is due to the decomposition of the PVA side chains with the decomposition of the main chain occurring at about 400−600 °C. A great deal of weight loss is observed in the range of the second stage of thermal decomposition, as shown in Figure 3, of the TGA and first order differential of the thermograms (DTGA). The powdered PVA shows a small weight loss in the first stage as compared with the nanofibers of PVA. For the electrospun nanofibers with β-CD, better thermal stability is observed than for the other samples (Figure 3b). This is evidence that β-CD improves the thermal stability of the encapsulate.18,26 The retinyl acetate weight loss was exhibited in four different stages. The first weight loss up to 100 °C corresponds to the evaporation of the adsorbed moisture. The second stage has 15% weight loss in the range 200−300 °C. The next 15% weight loss was observed in the range 300−430 °C. Moreover, the predominant and further weight loss was exhibited in the fourth stage in the range 430−600 °C. It is noticed that greater than 80% of the loss is observed after 300 °C. According to the

were subjected to X-ray powder diffractometry at RT. The XRD patterns obtained are depicted in Figure 2. The β-CD has

Figure 2. XRD spectra of received samples and electrospun nanofibers.

guest molecules which are localized separately from each other that affect the formation of crystals.25 A wide spectral peak of the RA centered at 2θ = 13° suggests a semicrystalline structure. The diffractogram peak at 2θ = 19.5° for PVA powder and PVA nanofibers largely reveals their similarity. The PVA nanofibers preserve the semicrystalline nature of the powder after electrospinning except for a small additional peak at 2θ = 29.5°. These observed XRD diffraction peak results agree well with the literature. The reflection peak for electrospun PVA/β-CD and PVA/RA was detected at 2θ = 21.5° and shifted to the lower shoulder peak, which indicates the slight change in crystallinity. In addition, the PVA/RA/βCD fibers had broad peaks at 2θ = 19.5° and 2θ = 21.5°, which is evidence of the existence of the introduced RA and β-CD in

Figure 3. (a) TGA thermograms for the PVA and RA powders and the electrospun fibers and (b) first order derivatives of the TGA thermograms for powdered PVA and RA and the electrospun fibers. D

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry

∼10 nm of the outer surface of the samples. XPS wide energy surveys of PVA and PVA/β-CD nanofibers show different atomic concentrations of oxygen and carbon, namely, 65.6%/ 33.8% and 48.2%/49.0%, respectively (Table 2 and Figure 5).

TGA finding (Figure 3a), the RA in PVA/β-CD nanofibers has more thermal stability than the RA incorporated into PVA nanofibers. These results are consistent with results in other studies,13,26 which include reports on how incorporating cyclodextrin inclusion improved the stability of the capsulate. Differential scanning calorimetry (DSC) is used to identify if the guest molecules are incorporated inside the CD cavities.25,27 The semicrystalline polymer of PVA has both the removal of its moisture and melting temperature (Tm), as depicted in Figure 4. Due to the introduction of RA, the

Table 2. Surface Atomic Composition Measured by XPS total atomic composition (%) material

C

O

PVA PVA/β-CD PVA/RA PVA/RA/β-CD

65.6 48.2 42.7 41.7

33.9 49.0 54.6 56.2

N

Na

1.7 0.8 2.0

0.5 1.1 2.0 0.2

The atomic composition identified of PVA nanofibers contains exclusively carbon, oxygen, and sodium. The electrospun PVA/ β-CD nanofibers were analyzed to study the difference of compositions in detail (Figure 5b). In addition, the surface chemistry study by XPS investigated the variation of the surface composition with the presence of β-CD and retinyl acetate (Table 2). The XPS spectra of the PVA/RA and PVA/RA/βCD nanofiber composites show four distinct peaks, corresponding to Na, N, C, and O, as shown in Figure 5c and d. Particularly, in the nanofibers of PVA/RA and PVA/RA/β-CD, increased concentrations of oxygen and slight variations of nitrogen were observed due to the introduction of retinyl acetate into the nanofibers (Figure 5c and d). The potential of using β-CD was assessed by observing its tendency to preserve oxygen against some common oxidation of the retinyl acetate. The electrospun nanofibers of PVA were used as a control for comparing the variation due to the introduction of functional groups from the RA and β-CD inclusion complex. The result displayed a high oxygen atomic concentration, which is in agreement with the previous finding that the β-CD inclusion complex has a critical role to reduce loss of oxygen mainly in the form of oxidation that reduces the shelf life of the samples.34,35 These results confirmed the formation of an inclusion complex and its preserving capacity to prolong the shelf life of the products. Monitoring Release of Retinyl Acetate. The quantity and release kinetics of RA in electrospun nanofibers of PVA/ RA and PVA/β-CD/RA were determined using the squarewave voltammogram peak current response against storage time under room storage conditions. The peak signal response of the retinyl acetate corresponds to the concentration of RA found in the nanofibers using the calibration graph of standard retinyl acetate (inset of Figure 7). The subsequent controlled release kinetics from the nanofibers. A known amount of electrospun PVA/RA and PVA/β-CD/ RA nanofibers was soaked in ethanol and sonicated to release the retinyl acetate. Subsequently, 600 μL of this solution was added into 0.1 M TBAPF6 in acetonitrile solution in a 10 mL electrochemical cell. The SWV signal response revealed the amount of entrapped RA in the nanofibers according to the storage time, as shown in Figure 6. As can be seen from Figures 6 and 7, the concentration of RA in the nanofibers PVA/RA/βCD and PVA/RA decreased with respect to increased storage time. Furthermore, the release rate was faster in both the PVA/ RA/β-CD and PVA/RA nanofibers during the first 3 weeks as compared to the remaining times. Generally, the peak current signal suggested that the release rate of PVA/RA/β-CD-IC was slower than PVA/RA within the same storage conditions. This slower release rate for the PVA/RA/β-CD-IC nanofibers can be

Figure 4. DSC thermograms of powders and electrospun nanofibers.

thermal behavior of PVA is expected to affect the composition interaction. Figure 4 shows the DSC curves of the powdered PVA and the electrospun nanofibers. The moisture content of the RA powder was removed at 105 °C, and the melting point at 160.6 °C was observed. The sharp melting point peak of βCD appeared at 142.5 °C. The powder of PVA had moisture removed at 84 °C, whereas it occurred at 87 °C for the electrospun PVA nanofibers. The melting temperature of PVA powder and electrospun PVA nanofibers appeared at 221 and 225.5 °C, respectively. This result agrees with the literature value.28 However, the melting point of PVA nanofibers increased by 4.5 °C, which may be due to the processing of the fibers. Generally, the melting point of the nanofibers is exhibited at the same temperature around 221 °C. The melting points of RA and β-CD powder were not observed in the nanofibers. This absence of the melting point suggested that RA incorporated inside the cavity and β-CD distributed through the nanofiber matrix for formation of inclusion complexation. Several studies have elucidated the β-CD unique molecule delivery nature that can be used for vitamin A encapsulation to increase the thermal stability and control release.29,30 For instance, the retinyl acetate without encapsulation starts to lose its stability earlier than 150 °C.31 However, the stability of encapsulated retinyl acetate is improved in the range 230−300 °C without the presence of high chemical deterioration.32,33 XPS Characterization. X-ray photoelectron spectroscopy (XPS) was used to investigate chemical changes at the surface of the electrospun nanofibers and identify the atomic concentrations of the PVA, PVA/β-CD, PVA/RA, and PVA/ RA/β-CD nanofibers. Accordingly, XPS survey spectra of the nanofibers were used to identify the surface elements present in E

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry

Figure 5. XPS spectrum of the (A) PVA, (B) PVA/β-CD, (C) nanofiber encapsulated retinyl acetate in PVA/RA, and (D) PVA/β-CD/RA.

Figure 6. SWV of 40 mg of nanofiber membrane in 2 mL of ethanol. 600 μL aliquots were removed from each sample for analysis. (A) Fibers of PVA/RA and (B) fibers of PVA/RA/β-CD after storage time of a = day 1, b = day 8, c = day 23, d = day 35, e = day 46, f = day 54, g = day 61, h = day 80, and i = day 94. Experimental conditions: supporting electrolyte acetonitrile with 0.1 M TBAPF6 with a glassy carbon electrode; scan from 0.0 to 0.9 V using an amplitude of 25 mV, step of 4 mV, and frequency of 15 Hz.

attributed to the effective host−guest complex formation between the retinyl acetate and the β-CD. Since it is known that RA is highly unstable and sensitive to oxidation and light when it is stored without a coating matrix,

studies carried out with RA shelf life tests aided in the formation of free-radicals, confirming that it has a short storage time due to subsequent loss of activity.36,37 In contrast, RA encapsulated into 2-hydroxypropyl-β-CDs was stable for 3 F

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry

Figure 7. Releasing kinetics monitored by square-wave voltammograms of retinyl acetate in the nanofibers of (A) PVA/RA and (B) PVA/RA/β-CD. Inset: the calibration graph of retinyl acetate and graph equation used to calculate the corresponding concentration signal of retinyl acetate in the electrochemical cell released from nanofibers of PVA/RA and PVA/β-CD/RA. Experimental conditions same as those for Figure 6.

months of storage at room temperature.38 In addition, RA encapsulated through nanoencapsulation into a polymer poly(ethylene glycol)−4-methoxycinnamoylphthaloylchitosan (PCPLC) increased its stability by 99% from free RA.5 Similarly, the electrochemical detection results in Figure 7 showed these encapsulations protected RA to extend the shelf life. These results demonstrate that electrospinning can be used as a method to fabricate nanofibrous structures for encapsulating bioactive retinyl acetate to prolong shelf life and thermal stability. This was achieved by encapsulating retinyl acetate inside the cavity of β-CD within the fibers to improve stabilities over longer time periods. Thus, electrospun nanofibers could be suitable for encapsulating food materials effectively and efficiently for prolonged shelf life and higher temperature stability and may be applicable in other areas such as the pharmaceutical and cosmetics industries, for edible food packaging, encapsulating antioxidants, food colors, flavors, and drugs.

■

(2) Webster, R. D. Voltammetry of the liposoluble vitamins (A, D, E and K) in organic solvents. Chem. Rec. 2012, 12 (1), 188−200. (3) Loveday, S. M.; Singh, H. Recent advances in technologies for vitamin A protection in foods. Trends Food Sci. Technol. 2008, 19 (12), 657−668. (4) Taepaiboon, P.; Rungsardthong, U.; Supaphol, P. Vitamin-loaded electrospun cellulose acetate nanofiber mats as transdermal and dermal therapeutic agents of vitamin A acid and vitamin E. Eur. J. Pharm. Biopharm. 2007, 67 (2), 387−397. (5) Arayachukeat, S.; Wanichwecharungruang, S. P.; Tree-Udom, T. Retinyl acetate-loaded nanoparticles: dermal penetration and release of the retinyl acetate. Int. J. Pharm. 2011, 404 (1−2), 281−288. (6) Gonnet, M.; Lethuaut, L.; Boury, F. New trends in encapsulation of liposoluble vitamins. J. Controlled Release 2010, 146 (3), 276−290. (7) Champagne, C. P.; Patrick, F. Microencapsulation for the improved delivery of bioactive compounds into foods. Curr. Opin. Biotechnol. 2007, 18 (2), 184−190. (8) Swetank, Y. H.; Karthik, P.; Anandharamakrishnan, C. Effect of whey protein isolate and β-cyclodextrin wall systems on stability of microencapsulated vanillin by spray−freeze drying method. J. Food Chem. 2014, 174, 16−24. (9) Gouin, S. Microencapsulation: industrial appraisal of existing technologies and trends. Trends Food Sci. Technol. 2004, 15 (7), 330− 347. (10) Del Valle, E. M. M. Cyclodextrins and their uses: a review. Process Biochem. 2004, 39 (9), 1033−1046. (11) Bazhban, M.; Nouri, M.; Mokhtari, J. Electrospinning of cyclodextrin functionalized chitosan/PVA nanofibers as a drug delivery system. Chin. J. Polym. Sci. 2013, 31 (10), 1343−1351. (12) Uyar, T.; Hacaloglu, J.; Besenbacher, F. Electrospun polystyrene fibers containing high temperature stable volatile fragrance/flavor facilitated by cyclodextrin inclusion complexes. React. Funct. Polym. 2009, 69 (3), 145−150. (13) Kayaci, F.; Uyar, T. Electrospun zein nanofibers incorporating cyclodextrins. Carbohydr. Polym. 2012, 90 (1), 558−568. (14) Moreno, I.; González-González, V.; Romero-García, J. Control release of lactate dehydrogenase encapsulated in poly (vinyl alcohol) nanofibers via electrospinning. Eur. Polym. J. 2011, 47 (6), 1264−1272.

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Phone: +61 (0)3 9214 5033. Funding

The authors acknowledge financial support received from Free University of Bozen-Bolzano, Italy, and Swinburne University of Technology, Australia. Notes

The authors declare no competing financial interest.

■

REFERENCES

(1) Gentili, A.; Caretti, F.; Bellante, S.; Ventura, S.; Canepari, S.; Curini, R. Comprehensive profiling of carotenoids and fat-soluble vitamins in milk from different animal species by LC-DAD-MS/MS hyphenation. J. Agric. Food Chem. 2013, 61 (8), 1628−1639. G

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Article

Journal of Agricultural and Food Chemistry

(35) Pinho, E.; Grootveld, M.; Soares, G.; Henriques, M. Cyclodextrins as encapsulation agents for plant bioactive compounds. Carbohydr. Polym. 2014, 101, 121−135. (36) Afzal, M.; Walton, J. C. Oxidation of retinyl acetate and analogues by nitric oxide and nitrogen dioxide. Bioorg. Med. Chem. Lett. 1996, 6 (19), 2329−2332. (37) Lemma, S. M.; Scampicchio, M.; Bulbarello, A.; Mason, M.; Schweikert, L. Concerted determination of the hydrogen atom and electron transfer capacity of lipid soluble reducing agents. Electroanalysis 2014, 26 (7), 1582−1587. (38) Lin, H. S.; Chean, C. S.; Ng, Y. Y.; Chan, S. Y.; Ho, P. C. 2hydroxypropyl-beta-cyclodextrin increases aqueous solubility and photostability of all-trans-retinoic acid. J. Clin. Pharm. Ther. 2000, 25 (4), 265−269.

(15) Kayaci, F.; Uyar, T. Encapsulation of vanillin/cyclodextrin inclusion complex in electrospun polyvinyl alcohol (PVA) nanowebs: Prolonged shelf-life and high temperature stability of vanillin. Food Chem. 2012, 133 (3), 641−649. (16) Marques, H. M. C. A review on cyclodextrin encapsulation of essential oils and volatiles. Flavour Fragrance J. 2010, 25 (5), 313−326. (17) Astray, G.; Gonzalez-Barreiro, C.; Mejuto, J. C.; Rial-Otero, R.; Simal-Gándara, J. A review on the use of cyclodextrins in foods. Food Hydrocolloids 2009, 23 (7), 1631−1640. (18) Kayaci, F.; Ertas, Y.; Uyar, T. Enhanced thermal stability of eugenol by cyclodextrin inclusion complex encapsulated in electrospun polymeric nanofibers. J. Agric. Food Chem. 2013, 61 (34), 8156−8165. (19) Kriegel, C.; Arecchi, A.; Arrechi, A.; Kit, K.; McClements, D. J.; Weiss, J. Fabrication, functionalization, and application of electrospun biopolymer nanofibers. Crit. Rev. Food Sci. Nutr. 2008, 48 (8), 775−97. (20) Fabra, M. J.; Lopez-Rubio, A.; Lagaron, J. M. High barrier polyhydroxyalcanoate food packaging film by means of nanostructured electrospun interlayers of zein. Food Hydrocolloids 2013, 32 (1), 106− 114. (21) Mascheroni, E.; Fuenmayor, C. A.; Cosio, M. S.; Di Silvestro, G.; Piergiovanni, L.; Mannino, S.; Schiraldi, A. Encapsulation of volatiles in nanofibrous polysaccharide membranes for humiditytriggered release. Carbohydr. Polym. 2013, 98 (1), 17−25. (22) Li, F.; Scampicchio, M.; Mannino, S. Carbon nanotubeadsorbed electrospun nanofibrous membranes as coating for electrochemical sensors for sulfhydryl compounds. Electroanalysis 2011, 23 (8), 1773−1775. (23) Fuenmayor, C. A.; Lemma, S. M.; Mannino, S.; Mimmo, T.; Scampicchio, M. Filtration of apple juice by nylon nanofibrous membranes. J. Food Eng. 2014, 122, 110−116. (24) Li, X.; Kanjwal, M. A.; Lin, L.; Chronakis, I. S. Electrospun polyvinyl-alcohol nanofibers as oral fast-dissolving delivery system of caffeine and riboflavin. Colloids Surf., B 2013, 103, 182−188. (25) Celebioglu, A.; Uyar, T. Electrospinning of polymer-free nanofibers from cyclodextrin inclusion complexes. Langmuir 2011, 27 (10), 6218−6226. (26) Song, L. X.; Xu, P. A comparative study on the thermal decomposition behaviors between beta-cyclodextrin and its inclusion complexes of organic amines. J. Phys. Chem. A 2008, 112 (45), 11341− 11348. (27) Nuchuchua, O.; Saesoo, S.; Sramala, I.; Puttipipatkhachorn, S.; Soottitantawat, A.; Ruktanonchai, U. Physicochemical investigation and molecular modeling of cyclodextrin complexation mechanism with eugenol. Food Res. Int. 2009, 42 (8), 1178−1185. (28) Ding, B.; Kim, H.-Y.; Lee, S.-C.; Shao, C.-L.; Lee, D.-R.; Park, S.-J.; Choi, K.-J. Preparation and characterization of a nanoscale poly(vinyl alcohol) fiber aggregate produced by an electrospinning method. J. Polym. Sci., Part B: Polym. Phys. 2002, 40 (13), 1261−1268. (29) Ghouchi-Eskandar, N.; Simovic, S.; Prestidge, C. A. Solid-state nanoparticle coated emulsions for encapsulation and improving the chemical stability of all-trans-retinol. Int. J. Pharm. 2012, 423 (2), 384−391. (30) Mathapa, B. G.; Paunov, V. N. Cyclodextrin stabilised emulsions and cyclodextrinosomes. Phys. Chem. Chem. Phys. 2013, 15 (41), 17903−17914. (31) Palmieri, G. F.; Wehrlé, P.; Duportail, G.; Stamm, A. Inclusion complexation of vitamin A palmitate with β-cyclodextrin in aqueous solution. Drug Dev. Ind. Pharm. 1992, 18 (19), 2117−2121. (32) Carlotti, M. E.; Rossatto, V.; Gallarate, M. Vitamin A and vitamin A palmitate stability over time and under UVA and UVB radiation. Int. J. Pharm. 2002, 240 (1), 85−94. (33) Kim, K.; Kim, D. High-performance liquid chromatography separation characteristics of molecular-imprinted poly (methacrylic acid) microparticles prepared by suspension polymerization. J. Appl. Polym. Sci. 2005, 96 (1), 200−212. (34) Krishnaswamy, K.; Orsat, V.; Thangavel, K. Synthesis and characterization of nano-encapsulated catechin by molecular inclusion with beta-cyclodextrin. J. Food Eng. 2012, 111 (2), 255−264. H

DOI: 10.1021/acs.jafc.5b00103 J. Agric. Food Chem. XXXX, XXX, XXX−XXX