1237-1244 Ahmad LAJP 3705:Ahmad

Latin American Journal of Pharmacy (formerly Acta Farmacéutica Bonaerense)

Regular article Received: May 6, 2014 Revised version: July 8, 2014 Accepted: July 9, 2014

Lat. Am. J. Pharm. 33 (8): 1237-44 (2014)

Synthesis and Characterization of Graft PVA Composites for Controlled Delivery of Valsartan Muhammad SOHAIL 1, Mahmood AHMAD 1*, Muhammad U. MINHAS 1, Liaqat ALI 1, Abubakar MUNIR 1 & Ikrima KHALID 2 1 Faculty

of Pharmacy and Alternative Medicine, the Islamia University of Bahawalpur, 63100, Punjab, Pakistan 2 Faculty of Pharmaceutical Sciences, GC University, Faisalabad, Punjab, Pakistan

SUMMARY. The purpose of present study was to develop chemically cross-linked poly vinyl alcohol-copoly (acrylic acid) hydrogel (PVA-AA hydrogel) for pH-responsive and controlled delivery of valsartan. Hydrogels were synthesized using free radical polymerization technique. Polymer (PVA) was chemically cross-linked with monomer (acrylic acid) in aqueous medium. Ethylene glycol di-methacrylate (EGDMA) and benzoyl peroxide (BPO) were used as cross-linker and initiator, respectively. Drug loading was performed with valsartan as a model drug. Characterization of hydrogels was performed by SEM, FT-IR, XRD, TGA and DSC. Hydrogels were evaluated for pH responsive behavior by equilibrium swelling ratio and swelling dynamics at low and high pH. Formation of hydrogel was confirmed by FT-IR, XRD, TGA and DSC studies. Maximum swelling, drug loading and release has been observed at pH 7.4. It is concluded that highly stable PVA and AA based polymeric matrices are developed, which are highly pH-sensitive. These polymeric matrices have potential to be used as a carrier for controlled delivery of valsartan. RESUMEN. El propósito de este estudio fue desarrollar químicamente un hidrogel reticulado de polivinilo alcohol-co-poliácido acrílico (hidrogel de PVA-AA) para obtener una respuesta de valsartán sensible y controlada al pH. Los hidrogeles fueron sintetizados utilizando la técnica de polimerización por radicales libres. El polímero (PVA) fue químicamente reticulado con un monómero (ácido acrílico) en medio acuoso. Etilenglicol di-metacrilato (EGDMA) y peróxido de benzoilo (BPO) se utilizaron como agente de reticulación e iniciador, respectivamente. La carga de fármacos se realizó con valsartán como un fármaco modelo. La caracterización de los hidrogeles se realizó por SEM, FT-IR, XRD, DSC y TGA. Los hidrogeles fueron evaluados para el comportamiento de respuesta al pH por la relación de hinchamiento de equilibrio y de hinchamiento dinámico a pH bajo y alto. La formación del hidrogel se confirmó por FT-IR, XRD, TGA y los estudios de DSC. El máximo hinchamiento, carga de fármaco y liberación se observó a pH 7,4. En conclusión, se desarrollaron matrices poliméricas de PVA altamente estables basados en AA que son altamente sensible al pH. Estas matrices poliméricas pueden ser potencialmente útilies como portadores para la liberación controlada de valsartán.

INTRODUCTION Hydrogels are polymeric networks having three dimensional configurations capable of imbibing high amount of water or biological fluids 1,2. The presence of hydrophilic groups such as OH, -CONH2-, -SO3H- and -CONH- in the structure of polymer forming hydrogel make the system highly water absorbent and thus highly swellable 3,4. These groups attached with a polymer chains when come in contact with a solution, dissociates partially or completely and form strong or weak electrolyte groups within polymer chain. Gel network is increased when

these charged groups repel each other; as a result expansion of the gel network occurs 2. Hydrogel without being dissolved 5 attains different levels of hydration due to the presence of these hydrophilic groups depending upon the composition of polymer and aqueous environment 1. In physically cross-linked hydrogels, physical interactions are present in between the chains of different polymers, which cause hindrance in dissolution, while in case of chemically crosslinked gels, covalent bonds are present 6. Interest towards synthetic hydrogels is gaining attention in biomedical field due to its wide

KEY WORDS: Polyvinyl alcohol, Acrylic acid, Hydrogel, Valsartan, pH-responsive *

Author to whom correspondence should be addressed. E-mail: [email protected]

ISSN 0326 2383 (printed ed.) ISSN 2362-3853 (on line ed.)

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SOHAIL M., AHMAD M., MINHAS M.U., ALI L., MUNIR A. & KHALID I.

variety of applications. Synthetic hydrogels possess high water content, tremendous swelling behavior and good compatibility with living tissues. In the field of biomedicine, various types of hydrogels are used, depending upon their properties and applications. Poly vinyl alcohol (PVA) systems are broadly studied for their various applications 7. PVA is a water soluble and a biodegradable polymer and is used in the synthesis of hydrogels 8 has been proved to be an excellent polymer due to its compatibility and processability and is suitable for use in contact lenses, peritoneal dialysis, tissue scaffolding and as an artificial cartilage 9. Due to its noncomplex chemical structure, it proves to be one of the most non-toxic and non-carcinogenic polymer for use in biomedical and pharmaceutical field. It has excellent water retaining properties and swells well to transform into elastic nature. Therefore, it is biocompatible with body tissues. Due to these characteristics, it has been also used in drug delivery, artificial heart lining and artificial skin 10. Addition of acrylic acid (AA) to PVA imparts pH responsive properties to polymer and can be used for controlled release, sustained release and targeted delivery of various types of drugs and biologics 1,5. The carboxylic acid groups present in acrylic acid chain is readily ionizable and are very sensitive to pH. It also imparts swelling behavior of the polymer chain as carboxylic acid groups have strong affinity towards water molecules 4. Hydrogel have wide variety of applications as mentioned in literature, like bioseparation, biosensor, tissue engineering and drug delivery, etc. 11. The aim of current study was to determine the controlled release and pH responsive behavior of PVA and acrylic acid in hydrogel form and to investigate its drug delivery applications. PVA was combined with AA in different ratios

to develop hydrogel formulations by radical polymerization method. After formulation development, its stability, surface and thermal characteristics were investigated by using various techniques. The pH sensitivity and in vitro drug release performance was determined by conducting swelling and dissolution studies. MATERIALS AND METHODS Chemicals Valsartan was gifted by Werrick Pharmaceuticals (Pvt.) Ltd. Islamabad, Pakistan. Polyvinyl alcohol (PVA), acrylic acid (AA), ethylene glycol dimethacrylate (EGDMA) and benzoyl peroxide (BPO) were purchased from Sigma Aldrich, UK. Deionized distilled water was freshly prepared in our laboratory. Synthesis of PVA-co-polyAA hydrogels Hydrogel formulations in various ratios are shown in Table 1 were developed by free radical copolymerization method. This technique was previously employed by Minhas et al. 12 and was used with various modifications. A weighed amount of PVA was dissolved in water through continuous stirring at 800 rpm at 90 °C until a transparent solution was obtained. Nitrogen stream was allowed for 30 min to purge the polymer solution to remove dissolved oxygen. Benzoyl peroxide (initiator) was dissolved in a specific amount of ethanol at room temperature and acrylic acid (AA) was added after obtaining a transparent solution. The benzyl peroxide-AA mixture was added drop wise in PVA solution at room temperature with constant stirring. A specific amount of ethylene glycol diamine methacrylate (EGDMA) was added drop wise in the final solution. The resultant transparent solution was poured into glass tubes and kept in water bath at 55 °C for 4 h, 60 °C for 8 h, 65 °C for 8 h and at 70 °C for 4 h. Later on glass tubes were placed in room temperature and hydrogels

Formulation Code

Polyvinyl alcohol (g/100 g)

Acrylic acid (g/100 g)

EGDMA mol% of monomer

PAc-1 PAc-2 PAc-3 PAc-4 PAc-5 PAc-6 PAc-7 PAc-8 PAc-9

1.0 2.0 3.0 2 2 2 2 2 2

40 40 40 20 40 60 60 60 60

0.400 0.400 0.400 0.400 0.400 0.400 0.200 0.400 0.600

Table 1. Formulations of PVA-co-poly(AA) hydrogels.

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Latin American Journal of Pharmacy - 33 (8) - 2014

were cut in to 8 mm and 5 mm discs. Discs were washed with ethanol-water (70:30) to wash un-reacted contents. These discs were dried in vacuum oven at 40 °C for one week after initial drying of 24 h in laminar flow air. Swelling properties Swelling behavior and pH sensitivity measurement was performed at various pH ranges by immersing dried hydrogel discs in 0.1 M HCl, pH 1.2 and phosphate buffer solution of pH 7.4 at 37 °C. The weights of swollen discs were obtained at specific interval of time after removing surface water with blotting paper and then reimmersed the discs in respective solutions. The % swelling ratio (SR) was calculated using Eq. [1]: SR % =

Ws – Wd Wd

x 100

[1]

where Ws represents weight of swollen discs at specific time while Wd is the weight of dry hydrogel discs. Sol gel-fraction Sol and gel contents were measured to evaluate the amount of reactants consumed in the preparation of PVA-co-AA hydrogels. The soluble unreacted contents of the polymerization reaction are termed as sol contents. In order to measure the sol content, 2 mm size slabs of prepared hydrogel were cut and dried at 55 °C until constant weight is achieved. After drying, these slabs were then placed in Soxhelt apparatus and extraction was performed for twelve h in deionized boiling water. The extracted gels were removed and dried again at 55 °C until constant weight was achieved. Eqs. [2] and [3] were used to measure sole and gel fraction, respectively: Sol fraction =

Mi – Me x 100 Wd

[2]

where Mi represents the initial weight of dry gel before extraction and Me represents the weight of dry gel after extraction. Gel fraction = 100 – Sol fraction

[3]

Structure analysis (FTIR) Presence of specific functional groups in the polymer (PVA), monomer (AA) and all PVA/AA hydrogel formulations were confirmed using Fourier Transform Infrared Spectroscopy (FTIR).

Before the analysis, samples were properly milled. The analysis was made using Bruker FTIR (Tensor 27 series, Germany) by applying attenuated total reflectance (ATR) technology and Opus data collection software. FTIR spectra were obtained in the range of 4000-650 cm–1. Thermogravimetric analysis (TGA) Thermogravimetric analysis (TGA) for PVA, AA and hydrogel was performed using TA instruments Q5000 series Thermal Analysis System (TA instruments, West Sussex, UK). Samples were properly milled to powder form before the analysis. Approximately 0.5 to 5 mg sample of PVA, AA and hydrogel formulations were placed in the sample pan for analysis over a temperature range of 20 to 500 °C. Analysis was performed under a nitrogen purge and heating rate adjusted at 10 °C/min. Differential Scanning Calorimetry (DSC) TA instruments Q2000 Series Thermal Analysis system (TA Instrument West Sussex, UK) were used for the determination of glass-transition temperature (Tg) of samples. DSC analysis of PVA, AA and all formulations were performed by sealing 0.5 to 3 mg of samples in standard aluminum pan, keeping temperature range 20-500 °C at heating rate of 20 °C/min and nitrogen purging was performed at 20 mL/min. PXRD XRD pattern of samples were recorded at room temperature using Bruker D-8 powder diffractometer (Bruker Kahlsruhl, Germany). Samples in the powdered form were loaded on to the plastic sample holder and its surface was smoothened by a glass slide. Scanning of samples were performed over the range of 5-50° 2θ at a rate of 1° 2θ/min. A radiation source used was copper Kα having a wavelength of 1.542 Å and 1 mm slits. Morphological analysis (SEM) The structural morphologies of PVA/AA hydrogels were analyzed using scanning-electron microscopy (SEM). JEOL Analytical Scanning Electron Microscope (JSM-6490A, Tokyo Japan) was used for the examination of hydrogel samples. Dried discs of hydrogel were cut into optimum size particles and mounted on an aluminum stub with double adhesive tape. The stubs were coated with gold under argon atmosphere using gold sputter coater. Surface mor-

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phology of hydrogel was analyzed by taking photomicrographs randomly. Valsartan loading Valsartan was loaded as a model drug in all PVA/AA hydrogel formulations using SwellingDiffusion Method. For this purpose 0.8% drug solution was prepared in phosphate buffer of pH 7.4. As valsartan lowers the pH of solution up to 5.29 13 therefore, pH was adjusted to 7.4 again by drop wise addition of NaOH. Dried discs of 8mm were then placed in valsartan solution for 72 h at room temperature. After 72 h, the discs were dried at room temperature after washing with distilled water. At the end, loaded discs were placed in oven at 40 °C. Measurement of drug loading Measurement of valsartan loading in hydrogels were performed by extracting the weighed amount of polymer using same solvent for drug loading i.e. extraction of drug from hydrogel discs were performed using 25 mL fresh buffer solution. This process was repeated until no drug was found in solution. Calibration curve of valsartan dilutions (in buffer) were constructed to evaluate drug contents via UV-Vis-spectrophotometer (UV-1601 Shimadzu). Analysis was performed at λmax 225 nm. Dissolution properties Drug release from hydrogel at low and high pH values was evaluated to confirm the pH dependent and controlled release of valsartan from PVA/AA polymeric matrices. Dissolution studies were performed in 900 ml solutions of pH 1.2 and 7.4 in USP dissolution apparatus-II at 37 ± 0.5 °C. Samples collected at specific time points were then analyzed at 225 nm using UV–visspectrophotometer (UV-1601 Shimadzu).

Figure 1. PVA/AA Hydrogels after swelling at pH 1.2

and 7.4.

cross-linker. This fraction of hydrogel is usually soluble in water or physiological solutions 14. The sol fraction was calculated to evaluate the extant of reactants consumed. In the present study, negligible sol fraction was found and stable hydrogels were developed, consuming most of the reactants. It was observed that the gel fraction was increased with increase in AA and EGDMA ratios. Effect of different pH mediums on swelling Effect of simulated fluids with different pH on the swelling behavior of PVA-co-poly (AA) hydrogels is shown in Fig. 2. Swelling kinetics of hydrogels was evaluated in simulated buffers of pH 1.2 and 7.4. The swelling behavior of hydrogels at low and high pH can predict the behavior of drug-loaded hydrogels in different parts of gastrointestinal tract. All PVA-co-poly (AA) hydrogel formulations showed high pHsensitivity as the swelling behavior of hydrogels are highly dependent on pH of surrounding environment 15. Swelling index of hydrogels was very low at pH 1.2 which increased rapidly in

RESULTS AND DISCUSSION General properties of hydrogels Stable PVA and AA based hydrogels were prepared after successful polymerization reaction. Gels were milky white with all PVA and AA ratios. Gels were little bit soft and rubbery with over all good mechanical strength. All hydrogels had adhesive properties before drying which disappeared after drying. Discs in dried form are shown in Fig. 1. Sol-gel Fraction The sol fraction of hydrogel is composed of unreacted components like monomers and

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Figure 2. Effect different pH mediums on swelling in-

dex.


pH 7.4. The change in response to change in pH (from 1.2 to 7.4) was probable due to the presence of carboxylic groups in hydrogel structure. As carboxylic groups have hydrophilic nature, increase in pH cause deprotonation of carboxylic groups of AA to a negatively charged carboxylate ions 16,17, as a result electrostatic repulsion is produced which cause swelling of hydrogels 18. On higher pH, hydrogels interact with basic species; resulting in the restricted interaction of water molecules with acid molecules. The carboxylic groups which are non-ionized and restricted interaction between acid and water molecules result in a reduction of swelling ratio, especially at low pH (pH 1.2) 19,20. Effect of PVA and AA concentration on swelling and drug release A series of formulations were prepared with different polymer and monomer ratios. Hydrogels from PAc-1 to PAc-3 were with increasing PVA concentration, PAc-4 to PAc-6 was with increasing AA concentration. It has been observed in the developed hydrogels that by increasing PVA content in hydrogels, swelling ratio increased to some extent 21. Due to the presence of ionizable functional groups, PVA-AA hydrogels showed pH dependent behavior. PVA chains contain large number of hydroxyl groups which have made this polymer highly interactive with water. Similarly, presence of carboxylic acid groups associated with acrylic acid (AA) has made it pH responsive, which is shown by swelling behavior of PVA-AA hydrogels. These properties of hydrogel are also reported in our previously reported study 12. The behavior of hydrogels with increased ratio of PVA is due to the availability of more number of sites for cross-linking 22. As swelling and drug release are affected by same factors, so with increase of PVA ratio, drug release is also increased to some extent. Swelling rate of PVA-AA hydrogels increased as the amount of acrylic acid increased. Its seems that with increase of AA concentration, the number of positive ions (COO-) also increased within the hydrogel structure, as a result, positive osmotic pressure is developed leading the increase in swelling rate 16. Due to the presence of similar charges on the crosslinked polymeric chain, network was extended and Browny movements of chain were decreased 23. Drug release from hydrogels having higher swelling rate is also increased due to larger surface area and diffusion.

Drug Release Properties The percent release of valsartan from PVAco-poly (AA) hydrogels as a function of time is shown in Fig. 3. The concentration of valsartan released at specific time points was determined by UV-spectrophotometer. Environmental pH is an important parameter that mainly influenced the drug release from PVA-AA hydrogels. The amount of valsartan released in a specified time from PVA-AA based hydrogels, decreased with decrease in pH of the dissolution medium. As at low pH values, there is minimum electrostatic repulsion between carboxylic acid groups, present in hydrogel structure, thus swelling of gels is decreased which also results in minimum release of valsartan via diffusion. However, on high pH values, electrostatic repulsion between carboxylate groups is increased due to the presence of 0H- group, thus increasing swelling degree and ultimately release of valsartan 24,25. As can be seen in Fig. 3, the cumulative release of valsartan at pH 1.2 is less than 25% at the end of experiment (48 h), whereas at pH 7.4; more than 90% drug was released at the end of 48 h. These results indicate that due to higher swelling rates of the hydrogel at pH 7.4, providing larger surface area for diffusion and release of valsartan is facilitated. To explore PVA-AA hydrogels role as a controlled drug release system, we have evaluated the valsartan release up to 48 h. Previously, many authors 21,26,27 have investigated the role of PVA in different controlled release systems. In our experiment, we found that PVA-AA can retard valsartan release for 48 h. Cumulative percent release of valsartan is almost 25 and 95% at pH 1.2 and 7.4 respectively, as shown in Fig. 3. It has been reported in various studies that hydrogels show rapid initial release when came in contact with fluids, which normally

Figure 3. Effect different pH mediums on drug re-

lease.

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slows down after several minutes 28. The prepared PVA-AA hydrogels showed no burst release as soon as they came in contact with the release environment. The study has shown clearly that as drug carrier materials, PVA and AA combination have controlled release properties. This may be useful in designing and developing novel controlled release systems for various therapeutic agents. Structure Analysis (FTIR) FT-IR spectra of Valsartan, PVA, AA and PVA-co-poly (AA) are shown in Fig. 4. Various functional groups of drug, polymer, monomer and graft polymer can be identified and characterized for structural moieties. Valsartan spectrum showed the characteristic peaks at 2963 cm–1 for –CH aromatic, 1731 cm–1 due to >C=N and 1600 cm–1 for carbonyl groups (–C=O).

A broad absorption band appeared in PVA spectra at region of 3298.34 cm–1 that could be assigned to presence of O–H stretching vibrations. The peak at 2941.00 cm–1 indicates the presence of –CH2 groups. While peak at 1416 cm–1 is showing deformation of –OH. The FT-IR spectrum of acrylic acid (AA) revealed -CH2- stretching vibrations at 2972 cm–1, band at 1296 cm–1 shows the C–C stretching vibration, band at 1635 cm-1 is indicating C=O stretching and C–O stretching vibration of carboxylic acid is represented by band at 1173 cm1. The cross-linked network of PVA with acrylic acid showed a completely different pattern from its components. The developed cross-linked network indicates a broad high intensity peak at 1771 cm–1 that confirmed the ester linkage of carbonyl groups in cross-linking. FTIR studies confirmed the formation of new cross-linked groups and grafted copolymer of PVA and acrylic acid. Thermal Studies Before cross-linking, polymer and valsartan were evaluated with Thermogravimetric analysis and Differential calorimetry analysis. Thermograms of valsartan and drug loaded hydrogels are shown in Figs. 5-8. TGA of PVA indicates initial weight loss

Figure 5. TGA thermogram of PVA-co-AA hydrogels.

Figure 4. FTIR spectra of PVA, AA and PVA-co-AA

hydrogels.

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Figure 6. DSC thermogram of PVA-co-AA hydrogels.


Figure 7. TGA thermogram of Valsartan.

Figure 8. DSC thermogram of Valsartan.

above 80 °C and decomposition above 265 °C as shown in Fig. 5. DSC curve showed endothermic peak at 85 °C that could be assigned as Transition temperature (Tg) and a major endothermic maxima indicate decomposition of polymer at 200 °C that is melting temperature. DSC thermogram of cross-linked matrices of PVA with acrylic acid showed (Fig. 6) a completely different pattern from polymer and drug. Initial endothermic peak at 100 °C in DSC indicate water loss and endothermic maxima above 250 °C showed degradation of the polymeric network. In present study, the thermal stability of synthesized hydrogel was found to be stronger than that of individual components. Same results were reported in another study conducted by Kumari et al. 29. In a study conducted by Arndt et al. 30 blends of PVA and PAA were prepared by physical method, in which degradation of blends were occurred at 220 °C, showing the better stability of chemically crosslinked hydrogels as compared to hydrogels prepared by using physical methods. In another study conducted by Pooley et al. 4, acrylic acid based copolymers were prepared using MBA as a cross-linker, which showed thermal stability up to 200 °C. Thermal analysis of cross-linked hydrogels showed stability above 250 °C that is an adequate temperature range for stable carrier system to deliver various drugs. TGA and DSC Thermograms are shown in Figs. 7 and 8.

Powder X-ray Diffractometry (PXRD) PXRD was used to study the change in crystallinity of valsartan, shown in Fig. 9. The diffraction spectrum of valsartan showed that drug was crystalline in nature and showed various characteristic peaks. The distinct peaks at 2θ of 10.7, 19.532, 20.5, and 21.471° indicates valsartan characteristic pattern. Crystallinity of cross-linked network was also assessed by PXRD as shown in Fig. 10. The diffraction pattern of cross-linked network was different from pattern of valsartan that indicates decrease in crystallinity. In another study, conducted by Gupta et al. 31 decrease in crystallinity of polymer (PVA) is observed after the formation of hydrogel. The change in crystallinity of formulated valsartan could be assigned to distribution of valsartan in polymeric network. The diffraction pattern of loaded hydrogel showed the characteristic peaks of drug that confirms physical distribution of drug particles within the crosslinked network without chemical linkage or interaction of drug molecules with cross-linked components 32. PXRD studies explain the phenomenon of physical loading of drug in polymer matrix system and these results showed that drug has been delivered in controlled manner without changing the chemical nature of drug.

Figure 9. XRD patterns of Valsartan.

Figure 10. XRD patterns of PVA-co-AA hydrogels.

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Surface Morphology Cross-linked polymer network was studied to reveal the surface morphology of intact and cross-sectional area by scanning electron microscope. The intact and cross-sectional surface morphology of hydrogel discs was studied to observe the morphological differences. SEM micrographs indicate the porous nature of network. Hydrogel formulations with higher acrylic acid contents showed less porous while hydrogels with higher polymer ratio (PVA) showed more porous structure. Previously we have reported the similar effect of monomer contents on the porosity of cross-linked network 12. Photomicrographs of hydrogel morphology have been shown in Fig. 10. CONCLUSION PVA and acrylic acid based pH responsive co-polymer is formed exhibiting controlled release properties. New copolymer has characteristic properties and intrinsic properties of its components are modified via cross-linking. Hydrogels have good strength and are milky white in appearance. Hydrogels shows pH-responsive behavior, swelling and drug release is very low at acidic pH and high at alkaline pH. Valsartan is used as a model drug for loading and release studies. It can be concluded that PVA-AA based hydrogels would be an efficient controlled drug delivery system for various drugs. Acknowledgment. Higher Education Commission, Islamabad-Pakistan REFERENCES

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