Ⅰ. Introduction
Contact lenses, which are worn directly on the cornea of the eyeball, greatly affect the eyeball depending on the material constituting them. Hydrogel lenses, which have a three-dimensional hydrophilic polymer network, began to become widely distributed with FDA approval in the 1970s. Protein-based hydrogels, including collagen, fibrin, elastin, silk, and gelatin, have been widely used in various biomedical applications. Gelatin has a series of advantages including excellent biocompatibility, solubility, degradation, and ease of synthesis methods.1-3) Also, gelatin is a type of natural hydrophilic polymer produced by the hydrolysis and denaturation of collagen at high temperatures. In this process, the three-dimensional structure of collagen is unwound and converted to gelatin. Applications in the biological field of gelatin have low thermal stability and may be limited due to the undesirable effects of various bifunctional crosslinking agents on biocompatibility. GelMA, first described by A.Van Den Bulcke et al., was synthesized by reaction of an amine group of GelMA. The double bond of the GelMA structure generated by incorporating methacrylic acid was used to manufacture GelMA hydrogels through free radical polymerization in the presence of photoinitiators.4-6) GelMA is one of the most studied versatile biopolymers to form hydrogels and is used in biomedical engineering research such as micro-patterning, fluid systems, 3D scaffolding, bioprinting using various cell types, tissue adhesives, and drug delivery. Since GelMA is versatile in terms of processing as well, it has the advantage of being hydrogelled with mechanical properties that may be adjusted by crosslinking when UV irradiated in the presence of a photoinitiator.7,8) Among the chemical crosslinking methods, the photopolymerization method is fast, simple, and inexpensive because it forms a three dimensional hydrogel network. In particular, acrylate and methacryloyl substituents are the most commonly used reaction functional groups in photopolymerization, which are reacted by a chain growth mechanism.9-12) In this study, after synthesizing GelMA, it was photopolymerized with a hydrogel lens containing Irgacure2959, which is relatively less cytotoxic among the most commonly used photoinitiators in combination with GelMA, to investigate its application as a highly functional hydrogel lens.13)
Ⅱ. Materials and methods
1. Reagents and materials
Gelatin from porcine solution (PBS), methacrylic anhydride (MA) was used to synthesize gelatin methacrylate (TRISS), and 2-hydroxyethyl methacrylate (HEMA) and 3-[tris(trimethylsiloxy)silyl]propyl methacrylate (TRISS) photoinitiator 2-Hydroxy- 4'-(2-Hydroxyethylpropiophenone (Irgacure2959) and crosslinking agent Ethylene glycol dimethacrylate (EGDMA) were used. All reagents were purchased and used from Sigma Aldrich (USA).
2. Synthesis and polymerization
1) Synthesis of gelatin methacrylate
For the experiment, GelMA was synthesized with reference to Yayun Yan et al.14) After 5 g of gelatin from porcine skin was stirred with 50 ml of PBS at 60°C until completely dissolved, 4 ml of MA was slowly added to the solution, followed by magnetic stirring at 50°C for 3 hours. After diluting by adding 200 ml of PBS, stirring was terminated at 40°C. Furthermore, for removal of salt and MA, the mixture was dialyzed for 7 days using a 14-kDa cut-off dialysis bag, and then lyophilized for 7 days to prepare GelMA.
2) Compatibility evaluation of silicone hydrogel sample including GelMA material
Based on 3-[tris(trimethylsiloxy)silyl]propyl methacrylate (TRISS), 0.50% of EGDMA, a crosslinker, and 0.50% of Irgacure2959, a photoinitiator, were added to prepare a Ref sample, and the synthesized GelMA was added in ratios of 1.00, 2.00, and 3.00%, respectively, but agitation and photopolymerization could not proceed due to the layer separation phenomenon. Fig. 1 shows the layer separation phenomenon of the solution.
3) Manufacture of hydrogel lenses including GelMA
Ref. sample was prepared by adding 0.5% of EGDMA as a crosslinker and 0.50% of Irgacure2959, a photoinitiator, based on HEMA, and synthesized GelMA was added in ratios of 1.00, 2.00, and 3.00%, respectively. The groups to which GelMA was added by ratio were named GM1, GM2, and GM3, respectively. After stirring for 1 hr using an ultrasonic stirrer, photopolymerization was performed for 120 s using a photo polymerizer. The prepared lenses for measuring the physical properties were hydrated in physiological saline for 24 hr, and then light transmittance, refractive index, water content, and contact angle were measured, respectively. Table 1 and Fig. 2 show the solution by mixing ratio and mixing ratio for each sample used in the experiment, and Fig. 3 shows the chemical structure formula of the main reagent used in this study.
4) Optimization of hydrogel lens photopolymerization conditions including GelMA
In order to find the optimal polymerization condition of the hydrogel lenses including GelMA, the polymerization time after photopolymerization of the hydrogel lenses of the group to which GelMA was added at each ratio was adjusted for 50, 60, 90, and 120 s, respectively. The polymerization state of the hydrogel lenses for each polymerization time period is shown in Fig. 4.
3. Measuring Device and Methods
In order to confirm the synthesized GelMA, the synthesis result was confirmed using an energy dispersion (EDS), (Gemini 500, Zeiss, Germany). In addition, in order to measure the physical properties of the prepared lens, the polymerized lenses were hydrated in physiological saline for 24 hr and then the light transmittance, refractive index, water content, and contact angle were measured, respectively. Spectroscopic transmittance was measured for near-ultraviolet regions (UV-A and UV-B) using Optical Transmission (Cary 60) equipment. For the measurement of refractive index, an ABBE Refractometer (NAR- 1T, Atago, Japan) equipment was used. For the measurement of the water content, an electronic scale Ohaus (PAG 214C, Ohaus, USA) was used. In order to determine the wettability, the contact angle was measured using a GMBH (Kruss, Germany) equipment by a Sessile drop method. In this study, all samples were repeatedly measured more than 5 times to increase the accuracy of the experiment, and the average values were compared and analyzed after excluding samples with large error values as a result.
Ⅲ. Results and Discussion
1. Analysis of Synthesized of GelMA
An energy dispersion spectrometer (EDS) was used to confirm the GelMA synthesis result. The EDS results are shown in Fig. 5.
2. Spectral transmittance of samples
As a result of evaluating the optical properties by measuring the near ultraviolet region (280~ 380 nm) of the manufactured lens, the UV-B region of Ref. was 34.10 to 31.90%, and the UV-A region was 90.90 to 88.30%, respectively. As the ratio of GelMA gradually increased to 1.00 to 3.00%, in the case of the average light transmittance of the GM group, the UV-B region was 24.89 to 16.56%, and the UV-A region was 74.04 to 62.76%, respectively. As the proportion of GelMA with UV protection ability gradually increases, the GelMA(GM) group has been shown to effectively block UV rays in the UV-B and UV-A regions compared to Ref. The measurement results of the near ultraviolet region for each group are shown in Fig. 6.
3. Refractive index and water content of samples
As a result of measuring the refractive index of the manufactured lens, the average refractive index of Ref. was 1.4365. As the ratio of GelMA increased, GM1 was 1.4370, GM2 was 1.4376, and GM3 was 1.4383, increasing the refractive index. Therefore, as the ratio of GelMA increased, the refractive index increased. As a result of measuring the water content of the manufactured lens, the average water content of Ref. was 39.06%. The average water content of the group to which GelMA was added by ratio was 39.07% for GM1, 39.04% for GM2, and 39.08% for GM3. In general, the increase in the refractive index of a hydrogel lens shows an inverse relationship in which the water content decreases, but the addition of GelMA in this study did not affect the inverse change in the water content due to the increase in the refractive index. The refractive index and water content measurement results for each group are shown in Fig. 7.
5. Contact angle of samples
As a result of measuring the contact angle to confirm the wettability of the manufactured lens, Ref. was found to be 70.53°. The groups in which GelMA was added by ratio were 70.43% for GM1, 70.83% for GM2, and 70.75% for GM3, showing no significant difference from the Ref. group. It is judged that there is no difference in wettability due to no change in moisture content. The measured values and images of the contact angle for each group are shown in Fig. 8 and Fig. 9, respectively.
Ⅳ. Conclusion
In this study, GelMA was synthesized using gelatin, and the synthesis result was confirmed through EDS analysis result. The synthesized gelatin methacrylate was added to basic mixture with HEMA and photopolymerized with a hydrogel lens material, thereby comparing and analyzing the properties of each. As a result of the study, the addition of GelMA gradually increased the refractive index according to the amount of addition, and the water content did not decrease even when the refractive index increased. In addition, it was shown that the UV blocking ability improved as the amount of addition increased. Therefore, GelMA added to the prepared hydrogel lens is effective in improving the refractive index of the lens and may be used in various ways as a high refractive index hydrogel lens and a medical material in view of a result that does not affect other physical properties.
