Method for improving enzymatic hydrolysis of polyester by using a deep eutectic solvent as an additive

Abstract

This invention discloses a method for improving the enzymatic hydrolysis of polyester using a eutectic solvent as an additive, belonging to the field of surface modification technology. Specifically, the enzymatic hydrolysis method of this invention uses betaine as a hydrogen bond acceptor and polyol as a hydrogen bond donor, prepared with Tris-HCl buffer to form a mixed buffer solution, and adjusted to pH 8.5 with hydrochloric acid. Then, the refined polyester fabric is placed in the mixed buffer solution, and Novozym 51032 keratinase is added for hydrolysis. After the reaction, the fabric is washed and dried. Using this method, the stability and activity of Novozym 51032 keratinase are improved, and the release of hydrolysis products is increased by 155.17% compared to enzymatic hydrolysis without a eutectic solvent.

Description

Technical Field

[0001] This invention relates to a method for improving the enzymatic hydrolysis of polyester by using a eutectic solvent as an additive, belonging to the field of surface modification technology. Background Technology

[0002] Polyester is one of the most widely used synthetic fibers. It possesses excellent properties such as high strength, wrinkle resistance, and abrasion resistance. However, polyester has a low content of hydrophilic groups in its chain segments, resulting in poor hydrophilicity, low moisture regain, difficulty in dyeing, and a tendency to generate static electricity. This affects the comfort of wearing polyester and limits its application as a high-end material in fields such as medical devices, food packaging, and electronic components. Therefore, hydrophilic modification of polyester fibers is essential to better realize their high added value and improved wearing comfort.

[0003] Surface modification of polyester fibers can generally be achieved through techniques such as coating, surface oxidation, and surface grafting modification (e.g., chemically initiated grafting and radiation grafting) to improve the hydrophilicity of polyester fabrics. However, these methods require irritating chemical reagents or high energy input, are environmentally unfriendly and cumbersome, and have drawbacks such as time-limited modification effects and impact on fabric mechanical properties. In contrast, enzymatic methods for surface modification of polyester fabrics catalyze the hydrolysis of ester bonds on the polyester surface to generate hydrophilic groups, thereby improving the hydrophilicity of polyester. The treatment conditions are mild and environmentally friendly, and the enzymatic action is limited to the polyester surface layer, without damaging the fiber's inherent properties. However, enzymatic hydrolysis has disadvantages such as poor efficiency and long modification cycles. The main bottlenecks include enzyme activity, thermal stability, substrate accessibility, and the high crystallinity of PET.

[0004] Eutectic solvents (DES) are an alternative to ionic liquids (ILs). They possess similar properties to ionic liquids, offering advantages such as biocompatibility, safety, availability, low raw material cost, recyclability, sustainability, and biodegradability. They are used as suitable solvents, co-solvents, additives, or support agents. Research has shown that environmentally friendly eutectic solvents (DES) serve as a potential medium, exhibiting biocompatibility and providing an ideal natural environment for enzymes, preserving or even refolding protein structures. Furthermore, DES can also act as a matrix for maintaining protein activity under extreme conditions. Summary of the Invention

[0005] To address the problems of poor performance and long cycle in existing technologies that use bio-enzyme catalytic hydrolysis of polyester, this invention provides a method for improving the enzymatic hydrolysis of polyester by using a low eutectic solvent as an additive. This method not only effectively promotes enzymatic hydrolysis but also improves hydrophilicity and dyeing effect.

[0006] The purpose of this invention is to provide a method for improving the enzymatic hydrolysis of polyester using a eutectic solvent as an additive, the method comprising the following steps:

[0007] (1) Preparation of eutectic solvent

[0008] Add the hydrogen bond donor (HBD) and hydrogen bond acceptor (HBA) to a beaker and stir until clear and transparent to obtain a eutectic solvent; wherein the hydrogen bond donor is a polyol; and the hydrogen bond acceptor is one or both of choline chloride and betaine.

[0009] (2) Add the eutectic solvent prepared in step (1) to Tris-HCl buffer, sonicate to mix evenly, adjust the pH with hydrochloric acid, and prepare mixed buffer.

[0010] (3) Place the refined polyester fabric into the mixed buffer solution prepared in step (2), add Novozym 51032 keratinase for hydrolysis reaction, and after the reaction is completed, wash and dry the fabric.

[0011] In one embodiment, the polyol in step (1) is one or more of glycerol, ethylene glycol, sorbitol, and xylitol.

[0012] In one embodiment, the molar ratio of hydrogen bond acceptor to hydrogen bond donor in step (1) is 1-3:3-1; preferably 1:1-3.

[0013] In one embodiment, the temperature of stirring and dissolving in step (1) is 80 to 100°C.

[0014] In one embodiment, the Tris-HCl buffer solution in step (2) has a concentration of 5 mmol / L and a pH of 9 to 9.5.

[0015] In one embodiment, the volume fraction of the eutectic solvent in the mixed buffer solution in step (2) is 5% to 30%, preferably 20%.

[0016] In one embodiment, the ultrasound time in step (2) is 10 min, the power is 275 W, and the temperature is room temperature.

[0017] In one embodiment, the pH of the mixed buffer solution in step (2) is 7 to 9, preferably 8.5.

[0018] In one embodiment, the refined polyester fabric in step (3) refers to the polyester fabric refined in a constant temperature water bath at 80-100°C for 30 min to 1 h with a bath ratio of 1:40-60, 3-6 g / L soap flakes and 3-6 g / L anhydrous sodium carbonate added to deionized water, washed with deionized water, dried in an oven at 80-100°C to constant weight, and equilibrated for 24 h under conditions of 25±1°C and 65±2% humidity; more preferably, the bath ratio is 1:30, 5 g / L soap flakes and 4 g / L anhydrous sodium carbonate are added, refined at 98°C for 30 min, and dried at 80°C.

[0019] In one embodiment, the Novozym 51032 keratinase mentioned in step (3) has an enzyme activity of 15 KLU / g, an enzyme concentration of 6.5%, and a v / v ratio.

[0020] In one embodiment, the hydrolysis reaction bath ratio in step (3) is 1:40, the temperature is 50-70°C, and the time is 24-72h, preferably 72h.

[0021] In one embodiment, the washing in step (3) refers to washing the fabric with deionized water.

[0022] In one embodiment, the drying temperature in step (3) is 60-70°C.

[0023] A second objective of the present invention is to provide a polyester fabric obtained by the method described above.

[0024] A third objective of this invention is to provide an application of the above-described method in polyester processing.

[0025] A fourth object of the present invention is to provide an application of the above-described polyester fabric in the preparation of textiles.

[0026] The beneficial effects of this invention are:

[0027] (1) The present invention uses a low eutectic solvent (DES) as an additive to improve the enzymatic hydrolysis effect of polyester. The solvent is simple to prepare, low in cost, biodegradable, and the modification conditions are simple and environmentally friendly.

[0028] (2) In this invention, Novozym 51032 keratinase is used to catalyze the hydrolysis of ester bonds on the surface of polyester fabric, so that hydrophilic groups such as hydroxyl and carboxyl groups are generated on the surface of polyester, so that polyester can achieve the effect of hydrophilic modification while maintaining its own advantageous properties.

[0029] (3) The eutectic solvent synthesized in this invention is a hydrophilic eutectic solvent. Using the method of this invention, the activity and thermal stability of Novozym51032 keratinase are both improved.

[0030] (4) The present invention utilizes a low eutectic solvent (DES) in conjunction with keratinase to perform enzymatic hydrolysis of polyester fabrics, and the release of enzyme hydrolysis products is increased by 155.17% compared with no additives; the hydrophilic angle of activated enzymatic hydrolysis is reduced from 83.5° to 80.4°. Attached Figure Description

[0031] Figure 1 The figures show the test results of the contact angle of the polyester fabrics obtained in Example 1 and Comparative Example 1; (a) is Comparative Example 1; (b) is Example 1.

[0032] Figure 2 The images show scanning electron microscope (SEM) images of the fabrics obtained in Example 1 and Comparative Example 1; (a) is Comparative Example 1; (b) is Example 1.

[0033] Figure 3 The diagram shows the effects of different types of enzymatic hydrolysis products of polyester in Example 1 and Comparative Example 1.

[0034] Figure 4 The graph shows the K / S test results of the polyester fabrics obtained in Example 1 and Comparative Example 1 after dyeing.

[0035] Figure 5 The graph shows the effect of DES component on the release amount of enzymatic hydrolysis products in Example 5.

[0036] Figure 6 The graphs show the effects of the additives on the activity and stability of the enzyme in Example 6; (a) shows the stability results; and (b) shows the enzyme activity results. Detailed Implementation

[0037] The technical solution of the present invention will be explained and described below with reference to the embodiments of the present invention.

[0038] The polyester fabric used in the embodiments and comparative examples of this invention is: 400T semi-dull polyester filament plain weave fabric.

[0039] Novozym 51032 keratinase was purchased from Beijing Gaoruisen Technology Co., Ltd., with an enzyme activity of 15 KLU / g.

[0040] The testing method involved in this invention:

[0041] 1. Method for testing the water contact angle of fabrics

[0042] The polyester fabric to be tested was equilibrated for 24 hours at a temperature of 25±1℃ and a relative humidity of 65±2%. Three samples from different locations were cut and attached to a glass slide. The DSA 25 contact angle meter was used for testing, with deionized water as the test solution and a drop volume of 10μL. The distance between the drop needle and the surface of the polyester fabric was 10mm. The contact angle was measured and photographed. Each parallel sample was measured three times and the average value was taken.

[0043] 2. K / S value test for polyester fabrics

[0044] Prepare a dye bath with a liquor ratio of 1:100 and a methylene blue dye concentration of 0.5% owf. Add the dye to the fabric and react in a constant temperature shaker at 60℃ and 150 rpm for 2 hours. After the reaction is complete, wash with deionized water and dry at room temperature.

[0045] The K / S curves of pure and treated fabrics were plotted using a Datacolor 650 computer colorimeter (Datacolor Company, USA) under D65 light at a 10° angle. The color rendering index of each fabric sample was tested five times, and the average value was taken as the final result. The effect of DES ultrasonic pretreatment on the dyeing properties of polyester fabrics was analyzed.

[0046] 3. Quantitative analysis of hydrolysis products

[0047] TPA solutions with concentrations of 0 mg / L, 2.5 mg / L, 5 mg / L, 10 mg / L, 20 mg / L, and 40 mg / L were prepared using deionized water. The pH of these TPA solutions was adjusted to 8.0 using 0.1 mol / L NaOH solution. Using deionized water as a reference, the absorbance of the TPA solutions was measured at 240 nm using a dual-beam UV spectrophotometer. A standard curve was plotted with absorbance and TPA standard solution concentration as the ordinate and abscissa, respectively. The standard curve obtained by this invention is Y = 0.00781X + 0.00306, R... 2 =0.9994.

[0048] After inactivating the enzyme, the reaction residue was centrifuged, and the supernatant was collected. The absorbance of the reaction solution of keratinase-treated polyester was measured at a wavelength of 240 nm using a dual-beam UV spectrophotometer. Each group was tested three times. The obtained absorbance values ​​were substituted into the standard curve to obtain the TPA concentration.

[0049] 4. Qualitative analysis and testing of hydrolysis products

[0050] The Agilent 1200 series high performance liquid chromatograph (Agilent Technologies, USA) was used. The mobile phase was 1% glacial acetic acid: methanol = 35:65. The injection volume was 5 μL, the flow rate was 0.5 mL / min, the column was maintained at about 30 °C, and the detection was performed at a wavelength of 240 nm.

[0051] 5. Characterization of fabric appearance

[0052] Before testing, the samples were sputter-coated with gold, and the changes in the surface morphology of the polyester fibers were observed using a SU1510 scanning electron microscope. The accelerating voltage during the electron microscopy scanning test was 5.0 kV, and the magnification was 10.0 kV.

[0053] 6. Enzyme activity assay method

[0054] The activity of keratinase was determined using a continuous spectrophotometer. The total volume of the test reaction was 1.5 mL, including 30 μL of appropriately diluted enzyme solution, 30 μL of 50 mmol / L pNPB, and 1440 μL of 5 mmol / L TrisHCl buffer (pH 8.5). The rate of p-nitrophenol formation was recorded at a wavelength of 405 nm. Enzyme activity was defined as the amount of enzyme that catalyzes the hydrolysis of p-nitrophenylbutyrate to produce 1 μmol of p-nitrophenol per minute at 37 °C.

[0055] Example 1

[0056] A method for improving the enzymatic hydrolysis of polyester using a eutectic solvent (DES) as an additive includes the following steps:

[0057] (1) Refining treatment of polyester fabrics

[0058] Prepare a refining solution by adding 5g / L soap flakes and 4g / L sodium carbonate at a liquor ratio of 1:30. Add polyester fabric and treat at 98℃ for 30min. Wash with water and dry at 80℃. Finally, place the polyester fabric in a constant temperature and humidity chamber (25±1℃, 65±2%) for 24h to equilibrate and obtain the refined polyester fabric.

[0059] (2) Preparation of eutectic solvent

[0060] Add betaine and polyol in molar ratio to a beaker and stir at 85°C until clear and transparent to obtain a eutectic solvent;

[0061] The specific molar ratios are: betaine (Bet): ethylene glycol (EG) = 1:2; betaine: glycerol (Gly) = 1:2; betaine: sorbitol (Sor) = 1:2; betaine: xylitol (Xyl) = 1:2;

[0062] (3) Preparation of mixed buffer

[0063] Dissolve the betaine-sorbitol (BS) prepared in step (2) in Tris-HCl buffer, sonicate for 10 min to mix evenly, adjust the pH with hydrochloric acid to obtain a mixed buffer with BS concentration of 20% (v / v) and pH of 8.5.

[0064] (4) Enzymatic hydrolysis

[0065] At a liquor ratio of 1:40, the polyester fabric refined in step (1) is placed in the mixed buffer solution prepared in step (3), and Novozym 51032 keratinase is added with an enzyme concentration of 6.5% (v / v). The mixture is reacted in a constant temperature shaker at 60℃ and 150rpm for 72h. After the reaction is completed, the polyester fabric is washed with deionized water, dried at 60℃ to constant weight, and placed in a constant temperature and humidity chamber (21±1℃, 65±2%) for equilibration for 24h.

[0066] Comparative Example 1

[0067] The only difference from Example 1 is that steps (2) and (3) are omitted, and the mixed buffer in step (4) is replaced with 5 mmol / L pH 8.5 Tris-HCl buffer. That is, the polyester fabric after the refining treatment in step (1) is directly enzymatically hydrolyzed. All other parameters and conditions are the same as in Example 1.

[0068] Comparative Example 2

[0069] The only difference from Example 1 is that Novozym 51032 keratinase in step (4) is omitted, while the other parameters and conditions are the same as in Example 1.

[0070] Comparative Example 3

[0071] The only difference from Example 1 is that step (4) is changed to placing the polyester fabric refined in step (1) into the mixed buffer solution prepared in step (3), reacting at 60°C and 150 rpm for 2 hours, and washing and drying with deionized water after the reaction; adding Tris-HCl buffer solution (5 mmol / L) at a bath ratio of 1:40, and then adding Novozym 51032 keratinase with an enzyme concentration of 6.5% (v / v), and reacting in a constant temperature shaker at 60°C and 150 rpm for 72 hours. Other parameters and conditions are the same as in Example 1.

[0072] Performance testing

[0073] 1. The modified polyester fabrics obtained in Example 1 and Comparative Examples 1 and 2 were subjected to performance tests, and the test results are as follows:

[0074] Table 1. Results of enzyme hydrolysis product release from modified polyester fabrics obtained in Example 1 and Comparative Examples 1 and 2

[0075] Modified polyester fabrics Example 1 Comparative Example 1 Comparative Example 2 Product release (mg / L) 236.59 152.47 34.49

[0076] As shown in Table 1, the addition of BS significantly increased the release of enzymatic hydrolysis products from polyester, with a 155.17% increase compared to enzymatic hydrolysis without eutectic solvent. The results of Comparative Example 2 indicate that the mixed buffer containing 20% ​​BS had little effect on the polyester hydrolysis effect; the main reason for the increased hydrolysis yield was the synergistic effect of BS and the enzyme.

[0077] 2. The modified polyester fabrics obtained in Example 1 and Comparative Example 3 were subjected to performance tests, and the test results are as follows:

[0078] Table 2. Results of enzyme hydrolysis product release from modified polyester fabrics obtained in Example 1 and Comparative Example 3

[0079] Modified polyester fabrics Example 1 Comparative Example 3 Product release (mg / L) 236.59 143.64

[0080] Table 2 shows that the product release from immersing the fabric in a eutectic solvent before enzymatic hydrolysis was significantly less than that from BS-co-enzymatic hydrolysis. This indicates that the increased hydrolysis yield is due to the interaction between DES and the enzyme, and that the non-substrate swelling of DES enhances the enzymatic effect.

[0081] 3. Performance tests were conducted on the modified polyester fabrics obtained in Example 1 and Comparative Example 1.

[0082] Contact angle test results as follows Figure 1 As shown, Figure 1 (a) is Comparative Example 1, and (b) is Example 1; from Figure 1 It can be seen that the contact angle of the polyester fabric hydrolyzed alone is 83.5°, and the contact angle of the hydrophilic modified polyester fabric obtained in Example 1 is 80.4°. After treatment by the method of the present invention, the contact angle is reduced by 3.71%, and the hydrophilicity is further improved.

[0083] Figure 2 The images are scanning electron microscope images of polyester fabrics treated in Example 1 and Comparative Example 1, where Figure (a) is Comparative Example 1 and Figure (b) is Example 1. As can be seen from the figures, the fiber surface is etched after enzymatic hydrolysis in Figure (a); the fiber surface etched by the synergistic enzymatic hydrolysis with additive BS in Figure (b) is more significant, and the fiber surface becomes rough.

[0084] Figure 3 The reaction residues after enzymatic hydrolysis in Example 1 and Control Example 1 were analyzed by high performance liquid chromatography (HPLC) to determine the types of hydrolysis products of polyester after enzymatic hydrolysis. Figure 3 It can be seen that compared with enzyme treatment alone, BS synergistic enzyme treatment did not change the types of hydrolysis products. The hydrolysis products are still TPA and MHET, and the hydrolysis products are mainly TPA monomers. The significant increase in hydrolysis yield proves that the eutectic solvent as an additive can enhance the hydrolysis of polyester by Novozym 51032 keratinase.

[0085] Figure 4 The K / S test results are for the polyester fabrics obtained in Example 1 and Comparative Example 1 after dyeing. Keratinase can hydrolyze the ester bonds in the polyester molecular chain into -COOH and -OH groups, and methylene blue can combine with carboxyl groups, thus allowing for the testing of hydrolysis effectiveness. The test results show that the K / S value of the polyester fabrics hydrolyzed with BS synergistically increases significantly at wavelengths of 550-650 nm, indicating that BS helps keratinase hydrolyze polyester and improves the enzymatic hydrolysis effect.

[0086] Example 2: Effect of DES type on the amount of enzymatic hydrolysis products released

[0087] The only difference from Example 1 is that the DES solvent in step (3) of Example 1 was adjusted to BG (betaine: glycerol), BE (betaine: ethylene glycol), BS (betaine: sorbitol), and BX (betaine: xylitol), respectively. All other parameters and conditions were the same as in Example 1. The hydrophilic modified polyester fabric was subjected to performance testing, and the test results are as follows:

[0088] Table 3. Results of product release for different eutectic solvents in Example 2

[0089]

[0090] As can be seen from Table 3, the eutectic solvents synthesized using betaine as a hydrogen bond acceptor and polyol as a hydrogen bond donor can all promote the hydrolysis of polyester by keratinase, with BS showing the best effect.

[0091] Example 3 Effect of DES component molar ratio on product release amount

[0092] The only difference from Example 1 is that the molar ratio of betaine to sorbitol (Sor) in step (2) of Example 1 was adjusted to 1:1, 1:2, 1:3, 2:1, and 3:1, respectively. All other parameters and conditions were the same as in Example 1. The hydrophilic modified polyester fabric was subjected to performance testing, and the test results are as follows:

[0093] Table 4. Results of product release at different DES component molar ratios in Example 3

[0094] Mole ratio 1:1 1:2 (Example 1) 1:3 2:1 3:1 Relative yield (%) 152.12 155.17 150.73 146.84 142.39

[0095] The HBA / HBD ratio not only affects the different interactions within DES, but also the different interactions between DES components and enzymes. As shown in Table 4, the addition of BS in all five molar ratios increased the hydrolysis release by more than 140%, with 1:2 being the optimal molar ratio.

[0096] Example 4: Effect of DES concentration on product release

[0097] The only difference from Example 1 is that the DES concentration in step (3) of Example 1 was adjusted to 5%, 10%, 15%, 20%, and 25%, respectively. All other parameters and conditions were the same as in Example 1. The hydrophilic modified polyester fabric was subjected to performance testing, and the test results are as follows:

[0098] Table 5. Results of product release at different eutectic solvent concentrations in Example 3

[0099] concentration(%) 5 10 15 20 (Example 1) 25 Relative yield (%) 129.72 132.15 139.9 155.17 82.22

[0100] As can be seen from Table 5, lower DES concentrations can synergistically increase the release of hydrolyzed polyester products, while concentrations exceeding 20% ​​will inhibit enzyme hydrolysis. This may be because excessively high DES concentrations may affect enzyme conformation and cause enzyme inactivation.

[0101] Example 5: Effect of DES single component on the release amount of enzymatic hydrolysis products

[0102] To determine whether the increased hydrolysis yield was due to a single component or to DES itself, step (2) was omitted, and step (3) was adjusted to correspond to the same molar concentration of the two components in 20% BS. 4.88% betaine (B) and 15.12% sorbitol (S) were directly dissolved in Tris-HCl buffer, with all other parameters and conditions the same as in Example 1; the results were as follows: Figure 5 As shown, by comparing the aqueous solution of the components with the DES solvent, it can be seen that the increase in enzymatic hydrolysis yield is due to the three-dimensional supramolecular network formed by hydrogen bond interactions in DES, rather than the synergistic effect of the two components.

[0103] Example 6: Effect of DES on enzyme activity and stability

[0104] Novozym 51032 keratinase was placed in Tris-HCl buffer (pH 8.5) and mixed buffer (BS concentration 20% v / v) at 50–70 °C for 2 h, and then the activity of Novozym 51032 keratinase was measured. After incubation under the same conditions for 24 h, the enzyme activity was measured again to determine the stability of Novozym 51032 keratinase in different solutions.

[0105] like Figure 6As shown in Figure a, a represents the effect of BS on enzyme stability, and b represents the effect of BS on enzyme activity. Figure a shows that Novozym 51032 keratinase exhibits better stability in a mixed buffer with 20% BS (v / v) than in Tris-HCl buffer. Novozym 51032 keratinase retains 38% of its activity in the mixed buffer, while only 20% is retained in Tris-HCl buffer. Figure b shows that BS has a certain activating effect on the enzyme, increasing its activity by approximately 1.2 times.

[0106] The embodiments provided above are not intended to limit the scope of the invention, nor are the described steps intended to limit the order of execution. Any obvious modifications made to the invention by those skilled in the art based on existing common knowledge also fall within the scope of protection defined in the claims of this invention.

Claims

1. A method for improving the enzymatic hydrolysis of polyester using a eutectic solvent as an additive, characterized in that, The method includes the following steps: (1) Preparation of eutectic solvent Add the hydrogen bond donor and hydrogen bond acceptor to a beaker and stir until the mixture is clear and transparent to obtain the eutectic solvent; wherein the hydrogen bond donor is a polyol and the hydrogen bond acceptor is betaine; The polyol is sorbitol; The molar ratio of the hydrogen bond acceptor to the hydrogen bond donor is 1~3:3~1; (2) Add the eutectic solvent prepared in step (1) to the Tris-HCl buffer, sonicate to mix evenly, adjust the pH with hydrochloric acid to obtain the mixed buffer. The volume fraction of the eutectic solvent in the mixed buffer solution is 5% to 20%. (3) Place the refined polyester fabric into the mixed buffer solution prepared in step (2), add Novozym 51032 keratinase for hydrolysis reaction, and after the reaction is completed, wash and dry the fabric. The Novozym 51032 keratinase has an enzyme activity of 15 KLU / g and an enzyme concentration of 6.5%, v / v. The hydrolysis reaction bath ratio is 1:40, the temperature is 50~70℃, and the time is 24~72h.

2. The method according to claim 1, characterized in that, The pH of the mixed buffer solution described in step (2) is 7-9.

3. The polyester fabric obtained after treatment by the method of claim 1 or 2.

4. The application of the method according to claim 1 or 2 in polyester processing.

5. The application of the polyester fabric according to claim 3 in the preparation of textiles.

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