Use of Gold Nanoparticles to Investigate the Drug Embedding and

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Use of Gold Nanoparticles to Investigate the Drug Embedding and Releasing Performance in Biodegradable Poly(Glycerol Sebacate) Yi-Kong Hsieh, Chia-Teng Chang, I-Hsin Jen, Fan-Chih Pu, Shu-Huei Shen, Dehui Wan, and Jane Wang ACS Appl. Nano Mater., Just Accepted Manuscript • DOI: 10.1021/acsanm.8b00723 • Publication Date (Web): 01 Aug 2018 Downloaded from http://pubs.acs.org on August 6, 2018

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ACS Applied Nano Materials

Use of Gold Nanoparticles to Investigate the Drug Embedding and Releasing Performance in Biodegradable Poly(Glycerol Sebacate) Yi-Kong Hsieh , Chia-Teng Chang, I-Hsin Jen, Fan-Chih Pu, Shu-Huei Shen§,⊥, Dehui Wan, and Jane Wang*

Department of Chemical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan Institute of Biomedical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan §Department of Radiology, Taipie Veterans General Hospital, Taipei 11217, Taiwan ⊥School of Medicine, National Yang-Ming University, Taipei 11221, Taiwan Keywords: poly(glycerol sebacate), gold nanoparticles, biodegradable polymer

ABSTRACT Poly(glycerol sebacate) (PGS) is a soft elastomer with excellent biocompatibility, and has great potential in the application of soft tissue engineering and drug delivery. However, there is still limited knowledge about the drug embedding capabilities of PGS and the drug releasing performance as PGS naturally degrades in vivo. In this work, PGS containing various amounts of gold nanoparticles (AuNPs) are prepared to simulate the embedment of nanomedicine in PGS, and to reveal the changes of AuNPs concentration and distribution in PGS

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before and after enzymatic degradation. The results show that AuNPs up to 15.9 µg are released from degraded PGS films after 28 days of enzyme degradation, along with mass loss up to 40.6% and around swelling of 5%. It is also found that there is an inversely proportional relationship between the AuNPs embedding concentration and the AuNPs release rate. The results obtained by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) reveal that not only is PGS degraded via surface erosion but lateral diffusion of AuNPs presents a diffusion coefficient of 1.126×10-14 cm2s-1 during the enzymatic degradation. With the quantitative characterization of AuNPs release rate from PGS composite materials, it is possible to apply PGS in embedding drug molecules for long-term drug delivery.

Keywords: poly(glycerol sebacate); gold nanoparticles; biodegradable polymer; laser ablation inductively coupled plasma mass spectrometry; diffusion coefficient

1. Introduction Gold nanoparticles (AuNPs) had been widely applied in biotechnology over the past two decade due to their unique physiochemical prosperities 1. They have been proven particularly useful in imaging diseases 2-8, delivering therapeutic agents 3, 9-10, and enhancing radiotherapy 11-14. As the surface of these particles are highly susceptible to a wide range of chemical modifications, functionalized AuNPs have been especially promising in the field of nanomedicine delivery, including protein, RNA, DNA, and numerous drugs 15-19. Although intravenous injections and oral ingestions remain two of the most common drug delivery methods, a multitude of drug carrying and delivering methods have emerged over the past two decades. Depending on delivery pathways and vehicles, delivery methods include transdermal, nasal, respiratory, and ocular drug delivery by employing novel vehicles such as liposomes 20, micelles 21, nanoparticulates 22, and biodegradable polymeric systems 23. Besides from instantaneous drug delivery, timed and controlled released of drug molecules are also of high interests for diseases that require long-duration treatment methods, and is particularly promising for chronic disease patients. Amongst the various drug delivery methods, drug delivery systems made from drug-embedded biodegradable polymers are considered very useful in local and long-term drug release, especially for drug delivery through the blood brain barrier (BBB) or high precision dosage control. In order to maintain high functioning controlled release


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ability in polymeric systems, the characterization of drug diffusion and the biodegradation of the polymeric system are the key to designing suitable long-term polymeric delivery systems 24. In the work of Niwa et. al, nano-spherical poly(lactic-co-glycolic acid) (PLGA)-embedded drugs were first synthesized and the release rate of drugs from biodegradable polymer was characterized 25. More work on the discussion between drug diffusion and polymer degradation was also done later on multiple polymeric systems, including PLGA, polylactic acid (PLA), polyvinyl alcohol (PVA), polycaprolactone (PCL), and polyethylene glycol (PEG) 26-33, all of which show high anticipation in applying biocompatible and biodegradable polymers toward the field of controlled release. However, most of the attempts 34-40 in the past two decades in creating drug-carrying polymeric devices had suffered from high drug diffusion rates through polymeric systems, resulting in unexpected and uncontrolled release of drug molecules. The drug release rates are also observed to be even less predictable during polymer degradation 37. As a new class of biodegradable polymer developed in the early 2000s, poly(glycerol sebacate) (PGS) is a glycerol-based network biodegradable elastomer 41. It is highly elastic and transparent due to its network polymeric properties. It has been proven especially applicable toward soft tissue engineering, as it degrades faster than common biodegradable polymer but slower than hydrogels 42-43. With the high biodegradability and low immunogenic nature during degradation, PGS may be suitable for an even wider range of applications besides tissue engineering. Although there are some studies that showed implants formed from PGS conjugated with 5-fluorouracil-1-acetic acid (5-FU) applicable toward anti-cancer drug release in vivo 44-45, the results focused only on the drug releasing performance of PGS without investigating drug distribution within PGS. Irina et al. investigated the mechanism and the physical changes of PGS during enzymatic degradation, but the PGS sample contained no drug or other additives 46. Since most drug molecules are organic molecules, it is very difficult to distinguish them from the polymeric systems they embedded in. Thus most of the existing works focused on characterize drug release rates after the drug molecules have left the polymeric systems 37. In this work, AuNPs are chosen as the embedded drugs for two reasons: 1. It has been demonstrated to be very useful in carrying a wide variety of drugs regardless of hydrophilicity 47. 2. It enables the observation of drug molecule movements through the polymeric systems using LA-ICP-MS, making it possible to estimate the diffusion coefficient of the drugs in PGS. In order to evaluate the applicability of PGS as a drug releasing polymeric implant in depth, the release profile of the nanoparticles while the polymer degraded are studied through a series of quantitative and analytical methods. 2. Experimental 2.1 PGS pre-polymer synthesis PGS pre-polymer was synthesized according to previously published methods 41. All materials were purchased from Sigma Aldrich. Briefly, an equimolar mixture of glycerol and sebacic acid was placed in a two-neck round bottom flask and melted at 130 ℃ under nitrogen for 2 h. The pre-polymer was formed through dehydration reaction under low pressure at 130 ℃ for 24 hrs. The prepolymer was cooled to room temperature and diluted with 99% ethanol for later use. 2.2 AuNPs synthesis


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AuNPs were prepared through sodium-citrate-mediated reduction of HAuCl4 according to previously published methods 48. Briefly, 50 mL of 1 mM aqueous HAuCl4 was heated until boiling with vigorous stirring. Then 5 mL of 38.8 mM sodium citrate was added to the boiling solution and continuously heated for 15 min. After the observation of color changed from paleyellow to wine-red, the solution was cooled to room temperature. To assist with nanoparticle suspension, 55 mg of polyvinylpyrrolidone (MW=55,000) was dissolved in the solution for 3 hrs, and then centrifuged to obtain AuNPs. The AuNPs were then resuspended in 1.5, 3, 7.5 and 15 mL ethanol to obtain AuNPs solution at 4.69, 2.35, 0.94 and 0.47 mM, respectively. 2.3 Preparation of PGS doped with AuNPs (PGS-AuNPs) Different concentrations of AuNPs ethanol solutions were mixed with PGS pre-polymer in the ratio of 1:10 (v/v). The mixed PGS/AuNPs solutions were dropped into a PDMS mold and evaporated simultaneously to remove the ethanol. Then the mold containing PGS/AuNPs was put into a vacuum oven at 160 and cured for 12 hrs. The weight ratios of AuNPs in PGS are suggested to be 3.0 × 10-2, 1.5 × 10-2, 6.0 × 10-3 and 3.0 × 10-3 wt% (AuNPs: PGS). 2.4 In vitro enzymatic degradation of PGS-AuNPs PGS-AuNPs composite materials were punched with a 13-mm biopsy punch and weighted before use. The weights of punched PGS-AuNPs were around 200 mg, and the thickness and diameter were about 1.5 and 10 mm, respectively. Type II lipase from porcine pancreas (100-500 units/mg) was diluted to 8 % (v/v) using phosphate buffered saline (PBS) solution. PGS-AuNPs were immersed in 4 mL of the lipase and incubated at 37 ℃. Enzyme solutions were renewed every two days. The recovered enzyme solutions were conserved at 4 ℃ for future analyses. PGS-AuNPs samples were soaked in distilled water for 30 min before the application of fresh enzyme solutions. Upon completion of degradation, samples were soaked in distilled water for 8 hours each. The samples were weighted and dried for 24 hours at 100 ℃. The dried samples were weighted, and weight loss and swelling ratio were obtained. 2.5 ICP-MS measurement The concentrations of released AuNPs were measured by inductively coupled plasma mass spectrometry (ICP-MS). Before the ICP-MS measurement, 27 µL of 30% hydrochloric acid was added to 0.5 mL of withdrawn lipase solution, followed by the addition of 50 µL of 65% nitric acid. The mixed solution was further diluted with distilled water to 5 mL, and then spiked with 10 µL containing 2 ppm of 205Tl as an internal standard.197Au was analyzed using a quadrupole ICP mass spectrometer (7500a, Agilent, USA). An operating RF power of 1.5 kW was applied and the plasma gas, auxiliary gas, carrier gas and makeup gas flow rates were 15, 2, 1 and 0.1 Lmin-1, respectively. 2.6 LA-ICP-MS measurement Fresh and degraded PGS-AuNPs composite materials were examined by laser ablation ICP-MS (LA-ICP-MS). Laser ablation system (UP-193, Electro Scientific Industries, USA) equipped with an ArF gas laser (wavelength: 193 nm) was applied to ablate solid sample. During the ablation, laser beam (diameter: 150 µm; laser energy: 4.0 J cm-2) was used to vaporize the sample in a selected area and introduced into the ICP-MS system, using Ar as the carrier gas (1.0


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L min-1). Three analyses were performed in this work: surface mapping, cross-section mapping, and depth profiling with detailed parameters in Table 1, and illustrations of three analyses are shown in supporting information Figure S1. For depth profiling analysis, the laser beam remains stationary for 10, 15, 20 and 30 sec each, which is also known as dwell time of laser beams. Elemental maps and integration of a defined area were produced using the Matrix Laboratory (MATLAB) software package (MathWorks Software, USA). A detailed analyses of the resolution of the three methods are summarized in Table S1. Table 1. Operating conditions for surface mapping, cross-section mapping and depth profiling of the LA-ICP-MS system Modes of Operation Surface Mapping Cross-Section Mapping Depth Profiling Ablation Mode Linear Scan Linear Scan Stationary Ablation Beam Size (µm) 150 150 150 Firing Frequency (Hz) 10 5 10 -1 Scan Speed (µm s ) 100 50 0 Laser Energy (J cm-2) 4.0 4.0 4.0 Scan Region 4.2 × 7.3 1.5 × 3.0 0.15 × 0.15 (mm × mm) MS Acquisition Rate 1 1 10 (Hz) 2.7 FE-SEM Image Fresh and degraded PGS-AuNPs composite materials were investigated by scanning electron microscope (SEM). Field emission scanning electron microscope (SU 8010, Hitachi, Japan) was used to measure the surface morphology of PGS. Samples were sputter-coating with palladium for 90s. The interested images were taken by using 10 kV accelerating voltage. 2.8 Surface topography measurement The depth of craters on PGS-AuNPs composite materials ablated by laser were measured through surface topography profilometer (Dektak 150, Veeco, USA) at 2000 µm of scan length, 50 µm s-1 of speed and 10 mg of stylus force. 3. Results and discussion 3.1 The Synthesis and Degradation of PGS-AuNPs Composite Materials To characterize the diffusion of drug molecules within the polymeric system as well as the drug release rate, AuNPs were chosen. The choices of AuNPs were especially advantageous for their capacity in carrying both a multitude of drugs 49-51 and their analyzability under LA-ICP-MS in solid matrices. In this work, 13 nm-sized AuNPs were embedded in PGS, a network polymer, to form PGS-AuNPs composite materials at four different concentrations: 3.0 × 10-3 (PGS-Au03), 6.0 × 10-3 (PGS-Au06), 1.5 × 10-2 (PGS-Au15) and 3.0 × 10-2 wt% (PGS-Au30). As shown in Figure 1 (a), the usually transparent PGS films transformed into burgundy colored films ranging from pale pink to dark purple. In order to investigate the diffusion coefficient of AuNPs in degrading PGS polymeric matrices, the theoretical model of self-diffusion from one thin film toward both sides was applied 52. To map the change of AuNPs in polymer matrices, depth


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profiling of LA-ICP-MS was employed. The x-axis of Figure 1 (b) was converted from time into depth of crater through the analysis of correlation between laser dwell time and crater depth in Figure S2. Since depth profiling etches composite materials around 0.46 µm at each ablation, the concentrated layer of AuNPs was intentionally placed near the surface for efficient observation of particle movements. In Figure 1 (b), the initial distribution of the AuNPs in each concentration was found most densely distributed between 2-6 µm from the surface.

(a)

(b)

Figure 1. (a) AuNPs composite materials at concentration of 3.0 × 10-3, 6.0 × 10-3, 1.5 × 10-2 ,and 3.0 × 10-2 wt% (from left to right). All nanoparticles are 13 nm in size. (b)AuNPs distribution of the four AuNPs-PGS materials obtained via LA-ICP-MS. Spatial resolution in zdirection of sample: 0.46 µm between points. PGS-Au03, PGS-Au06, PGS-Au15 and PGS-Au30 composite materials were soaked in lipase solution to facilitate the hydrolysis of the PGS matrix. Enzyme solution was renewed every two days. The withdrawn solutions were analyzed by ICP-MS to measure the concentration of the AuNPs released from the composite materials during the two-day period. The degraded PGS-AuNPs composite materials were collected every 7 days and analyzed via LA-ICP-MS and SEM for the distribution and concentration of AuNPs remained in the composite materials after degradation. The profiles of mass loss as well as swelling were shown in Figure 2. It is revealed through Figure 2 (a) that the weight loss was linearly proportional to


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the degradation duration, and the overall weight losses were about 30 to 40 % after 28 days of enzyme treatment. The high linearity between degradation time and weight loss directly indicated that surface erosion is the dominating degradation mechanism of the composite material. The dominance of surface erosion was further confirmed by the consistent swelling indices over the 28 days of degradation observed in Figure 2 (b), where the swelling indices increased by one to two percent during the full degradation period in all four of the concentrations. It is also worth noting that between the four composite materials with various AuNPs concentration, there were very little difference in swelling indices, but the degradation rate of the four materials were significantly different.

Figure 2. (a)Mass changes and (b) swelling index of PGS films containing different concentrations of AuNPs during the enzymatic degradation. 3.2 Analysis of Gold Nanoparticle Distribution in PGS-AuNPs Composite Material via LA-ICPMS and SEM Through the degradation of AuNPs composite materials via lipase, the samples with higher AuNPs concentration in PGS films were found to have degraded slower than those with lower concentration, while the swelling indices barely changed. In order to further understand this phenomenon, a series of analyses were conducted for the measurement of the AuNPs release profiles over the course of the degradation. To observe the changes in AuNPs concentration distributed on the surface of the degraded composite materials, surface mapping was conducted through LA-ICP-MS. Although LA-ICP-MS analyses have been proven an effective method in mapping the distribution of nanometals in animal studies, very few works have been done on the analyses of polymeric material. With the strong three dimensional mapping power of LA-ICPMS, it is possible to scan the PGS-AuNPs composite scaffold in both the x-y plane, as well as in the z-axis for a clear understanding of the distribution of AuNPs in the PGS matrix. With the laser ablation system, the surfaces of the composite materials containing PGS-Au30 are physically scanned (Figure S1 (a)). The ionized signals of residual AuNPs in the composite materials over 28 days are shown in Figure 3. The distribution and concentrations of AuNPs in the first 5 to 10 µm of the surface was clearly mapped through the scan, and was tracked throughout the 28 days of degradation. In Figure 3 (a) and (b), the Au concentrations on PGS surfaces were slightly decreased after 7 and 14 days of enzyme degradation, and after 21 and 28 days of degradation, the concentrations dropped even more significantly (Figure 3 (c) and (d)). It


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suggests that AuNPs were released from degraded PGS films vastly between 14 and 21 days of enzyme degradation. This was also confirmed through the analyses of released AuNPs in the withdrawn lipase solution using ICP-MS.

(a)

(b)

(c)

(d)

(e)

Figure 3. Mapping of the ion intensities of PGS-Au30 after (a) 0, (b) 7, (c) 14, (d) 21 and (e) 28 days of enzyme degradation. Spatial resolution in x and y-direction of sample: 100 and 150 µm between points. The cross-sections of PGS-Au30 over 0, 7, 14, 21 and 28 days of degradation were analyzed under linear scan mode with laser beam size of 150 µm, scan speeds of 50 µm s-1 and firing frequencies at 5 Hz (Figure 4 (a-e)). Further details on laser ablation parameters are listed in Table 1. It is clearly shown through Figure 4 that there was a concentrated layer of AuNPs in the bottom of the PGS-AuNPs composite materials in all samples, which decreased in concentration over the course of the degradation. The signals of concentrated layer in Figure 4 were integrated to estimate the AuNPs distribution in PGS-AuNPs. There were 2.7×106, 7.4×105, 5.4×105, 1.8×105 and 1.7×105 counts in the area of 0.5 mm × 3 mm in (width × length) in Figure 4 (a-e), respectively. The ratios of integrated area (concentrated layer) to the total signals were 68%, 71%, 73%, 70% and 67%. In the work of Matusch et al. and Barst et al., the mapping method through LA-ICP-MS is concluded to present an effective spatial resolution of approximately 100 µm 53, 54. Through similar calculations, the spatial resolution in Fig. 4 was approximately 50 µm in x-direction and 150 µm in y-direction in this work.


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(a)

(c)

(b)

(d)

(e)

Figure 4. LA-ICP-MS analysis of cross-section of PGS films containing PGS-Au30 after (a) 0, (b)7, (c) 14, (d) 21 and (e)28 days of degradation. Spatial resolution in y and z-direction of sample: 50 and 150 µm between points. To further investigate the AuNPs distributions in the bottom of the PGS films, stationary ablation mode with a laser beam size of 150 µm and shooting frequencies at 5 Hz was employed. Details of the parameters are shown in Table 1 and Figure S1 (c). The change of AuNPs distribution in the bottom 12 µm of PGS-Au30 over 28 days of degradation is compiled in Figure 5. The x-axis of Figure 5 was converted from time into depth of crater through the equation obtained in Figure S2 and aligned according to fit the analyzed surface of the scaffold. As the main degradation mechanism of the composite material was identified as surface erosion, the change in the distribution over time observed in Figure 5 hints at AuNPs diffusing through the bulk of PGS polymer. The results in Figure 5 indicated that there was a layer of AuNPs accumulated at about 2.39 µm from the bottom of the composite material before the degradation began. The peak of AuNPs was observed to have slowly migrated away from the bottom of the scaffold over the 28 days of degradation. As indicated in Figure 5 with red arrows, the peak moved 0.4 µm at 14 days of degradation, and then 0.79 and 1.59 µm at 21 and 28 days of degradation. It is indicated that AuNPs embedded in PGS became mobile after 14 days of degradation, while the height of the Au signal peaks decreased steadily. It is worth noting that the distribution of AuNPs in PGS films became apparently broader upon 21 days of degradation. The integrated counts in depth from 0 to 6 µm after 0, 7, 14, 21 and 28 days of degradation were 4.0 × 106, 3.8 × 106, 3.0 × 106, 2.0 × 106, and 1.3 × 106, respectively. The total counts were 1.6 × 107, 1.5 × 107, 1.2 × 107, 9.1 × 106 and 8.9 × 106 in samples after 0, 7, 14, 21 and 28 days of


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degradation, and the ratios of integrated counts in depth from 0 to 6 µm to the total signals were 25%, 25%, 25%, 22% and 15%. The results of integrated counts in Figure 5 indicated a decay in AuNPs concentrated layer by 60% after 28 days of degradation, which is significantly faster than the 30.8 % weight loss in Figure 2 (a). The diffusion condition of the AuNPs in PGS matrix were further analyzed using the self-diffusion model from one thin film toward both sides as shown in Eq (1). ∗

∗

∗

Eq (1)

When fitting the diffusion pattern over 28 days, it is found that the diffusion coefficient approaches 1.126×10-14 cm2 s-1 for AuNPs diffusing toward the membrane-water boundary. Although drug diffusion through biodegradable polymeric materials has long been observed, a precise measurement of the diffusion coefficient swelled state was never reported 53. The measured diffusion coefficient in this work certainly helps with the prediction of drug encapsulation ability of PGS polymer. The detailed information and simulation results are showed in Table S2.

Figure 5. The depth profile of Au signals obtained by LA-ICP-MS in PGS-Au30 after 0, 7, 14, 21 and 28 days of degradation. Spatial resolution in z-direction of sample: 0.46 µm between points. The degradation mechanism of the composite material is further confirmed through the SEM images in Figure 6, where both the surface and cross-sectional morphology of PGS-Au30 were observed under SEM over 28 days of degradation. Comparing between Day 0 of degradation and the Day 7, 14, 21, and 28 samples, the morphology of the composite material is observed to have become rough on the surface. It is worth noting that although the surface has become coarse upon degradation, the morphology of the cross-sections remains much of the


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same, with the exception of the porous surface layer (red circle). These results further confirmed that surface erosion is the dominating degradation mechanism for the composite material.

Figure 6. SEM images of the surfaces and cross-sections PGS-Au30 degraded over 28 days 3.3 The concentrations of AuNPs released from degraded PGS films As fresh lipase solution was applied every two days, the removed lipase solution was analyzed using ICP-MS, and cumulative amount of AuNPs released over the course of degradation was monitored via ICP-MS and presented in Figure 7. The results revealed that even though the mass loss was consistent throughout the 28 days of degradation, the concentration of AuNPs released during the first 14 days was very low. The release rate significantly increased from Day 16. This increase in release rate of AuNPs on Day 16 was consistent across all concentrations of AuNPs in the composite material, but the overall release rate is directly correlated to the concentration. This increase on 16 days of degradation may have been contributed by the mobilized AuNPs in the thoroughly swelled PGS film, which also corresponds to the higher correlation in the diffusion model shown in Table S2 in the suppling information.


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Figure 7. The release concentrations of AuNPs from PGS films during the enzymatic degradation 3.4 The proposed degradation rate, mechanism and released AuNPs doses Table 2 compares between the weight losses of PGS corresponding to the AuNPs release after 28 days of degradation. It was clear that under the same degradation condition, the higher the embedded AuNPs concentration, the lower the degradation rate. The higher the degradation rate, the higher the AuNPs release rate. However, the AuNPs release rate did not match with degradation rate linearly. Generally, the release rate of AuNPs was higher than the degradation rate of the overall composite materials, with the exception that AuNPs release rate was lower than the degradation rate of PGS-Au30. In summary, there was an inversely proportional relationship found between the AuNPs concentration and AuNPs release rate. It was previous reported by Li et al. that when PGS are mixed with fumed silica, the strand density may be greatly increase and the rate of degradation reduced 56. Similarly, the addition of AuNPs may have enhanced the endurance of PGS against enzymatic degradation, subsequently retaining the nanoparticles longer within the composite material. The low AuNPs release rate for the highly concentrated composite material also points to a possible threshold in nanoparticle embedding concentration toward reducing the particle release rate. As AuNPs are known to be easily modified for carrying various types of drug molecules, it is chosen in this work as a model molecule for drug release rate characterization. It is anticipated that as drug molecules of various hydrophilicity are attached on the AuNPs, the diffusion of drug molecules and the degradation properties of the polymeric composite materials will change accordingly. Also, AuNPs were intentionally embedded in a concentrated band near the bottom of the polymeric matrix for ease of observation through LA-ICP-MS and analysis for diffusion coefficient. However, as is proven through this work that AuNPs are easily monitored


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through analytical methods, precise control over the rate and concentration of drug release may be achieved by designing specific gradients of drug molecule distribution for long-term and steady release of drug molecules. The design of proper drug distribution may assist in solving problems from unpredictable drug release of homogeneous polymer-drug composites previously reported by others 37. Table 2. Comparison between weight losses of PGS corresponding to the AuNPs releases. (n=3) Weight (%)

Loss AuNPs released (µg)

Estimated mass of Percent AuNPs in PGS (µg) AuNPs released (%)

PGS-Au30

30.8 ± 0.2

15.9 ± 1.2

61.4 ± 0.7

25.9 ± 0.2

PGS-Au15

31.6 ± 0.3

12.2 ± 1.3

30.7 ± 0.4

39.7 ± 0.2

PGS-Au06

34.4 ± 0.2

9.3 ± 0.4

12.3 ± 0.5

75.6 ± 0.4

PGS-Au03

40.6 ± 0.3

4.0 ± 0.1

6.1 ± 0.1

65.6 ± 0.1

Conclusions In the present study, PGS-AuNPs composite materials were synthesized and the release profile of AuNPs from PGS biodegradable polymer was evaluated by employing ICP-MS and LA-ICPMS. In order to characterize the diffusion behavior of the AuNPs through the self-diffusion model from one thin film toward both sides, AuNPs were embedded in the bottom of the polymer films. The distribution of AuNPs within the polymer matrices were observed through surface mapping and depth profiling analyses. By tracking the concentration of the AuNPs released into the enzyme solution via ICP-MS, the concentrations of AuNPs released from degrading composite materials were observed to increase rapidly after 16 days of degradation. SEM images and the linear mass loss of the composite materials over 28 days of degradation also indicated that surface erosion was the dominated mechanism during enzymatic degradation, and a thin layer of degraded PGS film was clearly observed. The physical changes of PGS-AuNPs, including mass loss and swelling index, were also investigated, further proving that the composite material degraded mainly via surface erosion. Overall, it was found that AuNPs


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diffuse through polymer at a rate higher than PGS polymer degradation, which coincided with the drug diffusion rate observed in other biodegradable polymer. Through the methods described in this work, the diffusion coefficient of AuNPs in PGS biodegradable polymer was successfully measured. It is also found that by adding AuNPs to network polymers like PGS, the degradation rate of the overall composite material may be modified. Especially with the long term release capability of drug molecules attached on AuNPs via embedment in biodegradable polymers, local drug release dosage may be precisely predicted and controlled. Since the diffusion behavior was characterized, steady release of drug molecules may be achieved by designing concentration gradients based on the drug diffusion rate in clinical applications. ASSOCIATED CONTENT Supporting Information. The Supporting Information is available free of charge. The illustrations of PGS-AuNPs composite material analysis via LA-ICP-MS; The correlation between laser dwell time and crater depth on PGS-Au composites; Spatial resolution of LA-ICPMS methods; Diffusion condition of the AuNPs in PGS films using the self-diffusion model (Supporting Information for Publication.docx) AUTHOR INFORMATION Corresponding Author * Jane Wang; E-mail address: [email protected]; postal address: NO.101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan Notes


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The authors declare no competing financial interest. ACKNOWLEDGMENT Funding for this work was provided by Ministry of Science and Technology (Taiwan) under contract MOST 105-2218-E-007-008, 106-3114-E-039-001, 106-3114-E-007-009, and MOST 107-2923-E-033 -001. The postdoctoral fellowship of Y. K. Hsieh was funded by Ministry of Science and Technology (MOST 106-2811-E-007-047) and is gratefully acknowledged. We also thank the Instrument Center at National Tsing Hua University (Taiwan) for ICP-MS and LAICP-MS support. ABBREVIATIONS PGS, poly(glycerol sebacate); AuNPs, gold nanoparticles; LA-ICP-MS, laser ablation inductively coupled plasma mass spectrometry; SEM, scanning electron microscope; PLGA, poly(lactic-co-glycolic acid); PLA, polylactic acid; PVA, polyvinyl alcohol; PCL, polycaprolactone; PEG, polyethylene glycol; 5-FU, 5-fluorouracil-1-acetic acid; PGS-AuNPs, PGS doped with AuNPs; PBS, Phosphate buffered saline; ICP-MS, inductively coupled plasma mass spectrometry; REFERENCES 1. Daniel, M.-C.; Astruc, D., Gold Nanoparticles: Assembly, Supramolecular Chemistry, Quantum-Size-Related Properties, and Applications toward Biology, Catalysis, and Nanotechnology. Chemical Reviews 2004, 104, 293-346. 2. Wang, H.; Huff, T. B.; Zweifel, D. A.; He, W.; Low, P. S.; Wei, A.; Cheng, J.-X., In Vitro and In Vivo Two-Photon Luminescence Imaging of Single Gold Nanorods. Proceedings of the National Academy of Sciences of the United States of America 2005, 102, 15752-15756. 3. Zavaleta, C. L.; Smith, B. R.; Walton, I.; Doering, W.; Davis, G.; Shojaei, B.; Natan, M. J.; Gambhir, S. S., Multiplexed Imaging of Surface Enhanced Raman Scattering Nanotags in Living Mice Using Noninvasive Raman Spectroscopy. Proceedings of the National Academy of Sciences 2009, 106, 13511-13516.


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