Tanning salt-enzyme mixed solution as well as preparation method and application thereof

Abstract

The invention belongs to the field of leather animal skin unhairing, and discloses a leather-making salt-enzyme mixed solution as well as a preparation method and application thereof. The preparation method comprises the following steps: (1) adding neutral protease, alpha-amylase, peptidase, protease and glucanase into water to prepare a mixed enzyme solution; and (2) adding a salt substance into the mixed enzyme solution prepared in the step (1), and uniformly mixing to prepare the tanning salt-enzyme mixed solution. The tanning salt-enzyme mixed solution is applied to animal skin unhairing, collagen fiber bundle splitting and / or softening, the efficiency and effect of enzyme unhairing are improved, hydrolysis of skin grain surface collagen is reduced, and grain surface looseness is reduced; non-collagen components such as proteoglycan, elastic fiber and impure protein in the skin tissue are thoroughly hydrolyzed, so that the dispersity of collagen fiber bundles is improved.

Description

Technical Field

[0001] The invention belongs to the field of leather-making animal skin dehairing, and particularly relates to a leather-making salt-enzyme mixed solution, a preparation method and application thereof. Background Art

[0002] The traditional ash-alkali system for dehairing and splitting collagen fiber bundles will produce a series of water, soil and air pollutants such as ammonia nitrogen, suspended solids, hydrogen sulfide, and ammonia gas due to its reaction process of destroying hair and chemical hydrolysis. The irrational discharge of these harmful substances seriously endangers the surrounding water environment, soil environment and atmospheric environment, and thus endangers people's health; secondly, due to the low efficiency of the traditional ash-alkali system in the process of dehairing and splitting collagen fiber bundles, large water consumption, and multiple and cumbersome unit operations, the cost of human, material and natural resource input in the production process will increase, which is not conducive to the sustainable development of leather production.

[0003] While enzymatic dehairing technology is currently available, issues such as incomplete dehairing, loose grain, and severe grain hydrolysis have limited the use of enzymes and enzyme technologies in leather dehairing. Separating collagen fiber bundles requires the addition of carbohydrate enzymes, while softening requires pancreatic enzymes. These factors increase unit operation and process costs, highlighting the immaturity of enzyme technology. Therefore, thoroughly addressing the pollution issues inherent in traditional leather-making processes, improving the existing technical solutions in enzymatic leather-making processes, and achieving cleaner leather production are pressing challenges. Summary of the Invention

[0004] In view of the problems and shortcomings in the prior art, the present invention aims to provide a tanning salt-enzyme mixed solution and a preparation method and application thereof.

[0005] In order to achieve the purpose of the present invention, the technical solution adopted by the present invention is as follows:

[0006] The first aspect of the present invention provides a method for preparing a tanning salt-enzyme mixed solution, comprising the following steps:

[0007] (1) adding neutral protease, α-amylase, peptidase, protease and glucanase into water to prepare a mixed enzyme solution;

[0008] (2) adding salt substances to the mixed enzyme solution prepared in step (1), mixing them evenly, and obtaining a tanning salt-enzyme mixed solution.

[0009] Preferably, in step (1), the usage ratio of neutral protease, α-amylase, peptidase, protease, glucanase and water is (2-3.5) g: (1.5-3) g: (0.1-0.4) g: (0.05-0.3) g: (0.03-0.1) g: 1 L.

[0010] Preferably, the neutral protease CAS No.: 9068-59-1, Product No.: D832687, α-amylase CAS No.: 9000-90-2, Product No.: A834632, peptidase CAS No.: 9031-96-3, Product No.: P984275, protease CAS No.: 39450-01-6, Product No.: P6157, glucanase CAS No.: 9025-70-1, Product No.: D822632 in step (1), the neutral protease, α-amylase, peptidase, protease and glucanase are all purchased from Shanghai MacLean Biochemical Technology Co., Ltd.

[0011] Preferably, in step (2), the ratio of the amount of the salt substance to the mixed enzyme solution is (2.5-12) g:1 L, and the salt substance is an inorganic salt, an ionic liquid or a deep eutectic solvent.

[0012] More preferably, the inorganic salt is one or more of Na2SO2, KCl, MgCl2, NH4SO4, NH4Cl, ZnCl2, ZnSO4, Na2HPO4, NaCl, CaCl2, CH3COONa, Na2MoO4, BaCl2, and LiBr; the ionic liquid is one or more of imidazole ionic liquids, quaternary ammonium ionic liquids, pyridine ionic liquids, and quaternary phosphonium ionic liquids; and the deep eutectic solvent is one or more of choline chloride-urea, choline chloride-carboxylic acid, choline chloride-polyol, and zinc chloride-urea.

[0013] Further preferably, the pyrazole ionic liquid is one or more of 1-allyl-3-methylimidazolium chloride ([AMIM][Cl]), 1-butyl-3-methylimidazolium chloride ([BMIM][Cl]), and 1-butyl-3-methylimidazolium tetrafluoroborate ([BMIM][BF4]).

[0014] Preferably, the mixing process in step (2) is carried out by stirring evenly at room temperature.

[0015] The second aspect of the present invention provides a tanning salt-enzyme mixed solution prepared by the method described in any one of the first aspects.

[0016] The third aspect of the present invention provides a use of the tanning salt-enzyme mixed solution described in the second aspect in dehairing, splitting collagen fiber bundles and / or softening animal skins.

[0017] Preferably, the method for dehairing animal skins is as follows: the degreased and desalted animal skins soaked in water are mixed with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), and stirred for 2-6 hours at a temperature of 25-35°C and a rotation speed of 15-60 r / min.

[0018] Preferably, the method for splitting collagen fiber bundles is: mixing the dehaired animal skin with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), and stirring at a temperature of 25-35° C. and a rotation speed of 15-60 r / min for 1-24 hours.

[0019] Preferably, the method for dehairing / splitting collagen fiber bundles of animal skins is: mixing the degreased and desalted soaked animal skins with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), and stirring for 2-24 hours at a temperature of 25-35°C and a rotation speed of 15-60r / min.

[0020] Preferably, the method for dehairing / splitting collagen fiber bundles / softening animal skins is as follows: mixing the degreased and desalted soaked animal skins with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), stirring at a temperature of 25-40°C and a rotation speed of 15-60 r / min for 6-30 hours.

[0021] Compared with the prior art, the present invention has the following advantages:

[0022] (1) The tanning salt-enzyme mixed solution of the present invention promotes the salt-dissolving effect of proteins due to the electrostatic interaction between appropriate amounts of different types of ions and enzymes, thereby increasing the solubility (dispersibility) of various enzymes and improving the catalytic activity (vitality) of the enzymes. At the same time, due to the presence of salt ions, the osmotic pressure of the salt-enzyme solution system is increased, promoting the permeability of the enzyme into the skin tissue. Also, due to the electrostatic attraction of ions, the binding ability of the enzyme to the substrate is improved, thereby improving the catalytic efficiency of the enzyme on the substrate (proteoglycans, mucopolysaccharides, elastin, etc. in animal skins).

[0023] (2) The tanning salt-enzyme mixed solution of the present invention improves the efficiency and effect of enzymatic dehairing, reduces the hydrolysis of collagen on the leather grain surface, and reduces grain relaxation; non-collagen components such as proteoglycans, elastic fibers, and miscellaneous proteins in the leather tissue are completely hydrolyzed, thereby improving the dispersibility of collagen fiber bundles.

[0024] (3) The tanning salt-enzyme mixed solution of the present invention is used in dehairing, splitting collagen fiber bundles and / or softening of animal skins to improve the tanning production efficiency.

[0025] (4) The tanning salt-enzyme mixed solution of the present invention does not involve alkaline or acidic solutions during the application process, has no harmful gas emissions, has a hair-preserving process, has a low COD content in the water, uses less water (reduced by about 80%), and has simple process operations, which greatly reduces environmental pollution, reduces tanning costs and resource consumption, and improves the mechanical properties and thermal stability of leather. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 Zeta potential diagram of salt-enzyme mixture corresponding to different salt concentrations;

[0027] Figure 2 The contact angle diagrams of different types of salt-enzyme mixed solutions on cowhide;

[0028] Figure 3 This is a comparison chart of the effects before and after depilation of raw cowhide in Example 37;

[0029] Figure 4 This is a comparison chart of the effects before and after depilation of raw cowhide in Example 62;

[0030] Figure 5 The enzyme tracing diagrams of Comparative Example 1, Example 37 and Example 62 after 6 hours of depilation, wherein Figures A, B and C correspond to Comparative Example 1, Example 37 and Example 62, respectively;

[0031] Figure 6 EVG staining images of cross-sections of skin tissue after separation of collagen fiber bundles from raw cowhide, Comparative Example 2, and Example 73, where images A, B, and C correspond to raw cowhide, Comparative Example 2, and Example 73, respectively;

[0032] Figure 7 This is an EVG staining image of a cross section of skin tissue after separation of collagen fiber bundles in Example 98;

[0033] Figure 8 These are scanning electron micrographs of cross-sections of crust leather obtained after dehairing / collagen fiber bundle splitting / softening in Comparative Example 3, Comparative Example 4, Example 146, and Example 171, wherein Figures A, B, C, and D correspond to Comparative Example 3, Comparative Example 4, Example 146, and Example 171, respectively;

[0034] Figure 9 This is a graph showing the tensile strength of the crust leather obtained after dehairing / collagen fiber bundle splitting / softening in Comparative Example 3, Comparative Example 4, Examples 146 to 159, and Example 171;

[0035] Figure 10 This is a diagram of the wet heat shrinkage temperature of the crust leather obtained after dehairing / splitting the collagen fiber bundles / softening of Comparative Example 3, Comparative Example 4, Examples 146 to 159, and Example 171. DETAILED DESCRIPTION

[0036] In order to make the purpose, technical solutions and advantages of the present invention more clear and distinct, the present invention is further described in detail below through examples. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0037] Example 1

[0038] A tanning salt-enzyme mixed solution, the preparation method of which is as follows:

[0039] (1) Weigh 0.27 g of neutral protease, 0.22 g of α-amylase, 0.03 g of peptidase, 0.02 g of protease and 0.006 g of glucanase respectively, and add them to 100 mL of water to prepare a mixed enzyme solution, wherein the neutral protease CAS number: 9068-59-1, product number: D832687, α-amylase CAS number: 9000-90-2, product number: A834632, peptidase CAS number: 9031-96-3, product number: P984275, protease CAS number: 39450-01-6, product number: P6157, glucanase CAS number: 9025-70-1, product number: D822632, the neutral protease, α-amylase, peptidase, protease and glucanase were purchased from Shanghai MacLean Biochemical Technology Co., Ltd.;

[0040] (2) Weighing 0.8 g of a salt substance and adding it to the mixed enzyme solution of step (1), stirring it thoroughly at room temperature (about 25° C.) to completely dissolve the salt substance, thereby preparing a salt-enzyme mixed solution; wherein the salt substance is KCl.

[0041] Example 2 to Example 14

[0042] The contents of Examples 2 to 14 are substantially the same as those of Example 1, except that different salts are used in step (2). The salts used in step (2) of Examples 2 to 14 are: Na2SO2, MgCl2, NH4SO4, NH4Cl, Na2HPO4, NaCl, CaCl2, CH3COONa, Na2MoO4, NaHCO3, Na2SiO3, BaCl2, and LiBr, respectively.

[0043] Example 15 to Example 25

[0044] The contents of Examples 15 to 25 are substantially the same as those of Example 1, except that the amount of salt added in step (2) is different. The amounts of salt added in step (2) of Examples 15 to 25 are 0.05 g, 0.1 g, 0.15 g, 0.2 g, 0.4 g, 0.6 g, 0.7 g, 0.9 g, 1.2 g, 1.6 g, and 2 g, respectively.

[0045] Example 26

[0046] The contents are basically the same as those of Example 1, except that the salt substance used in step (2) is different. Example 26 replaces KCl in step (2) with 1-allyl-3-methylimidazolium chloride ([AMIM][Cl]).

[0047] Example 27 to Example 36

[0048] The contents of Examples 27 to 36 are substantially the same as those of Example 26, except that the amount of salt added in step (2) is different. The amounts of salt added in step (2) of Examples 27 to 36 are 0.05 g, 0.1 g, 0.15 g, 0.2 g, 0.4 g, 0.6 g, 0.9 g, 1.2 g, 1.6 g, and 2 g, respectively.

[0049] Performance test of different concentrations of tanning salt-enzyme mixed solution:

[0050] (1) Dispersion test

[0051] The KCl-enzyme mixtures with different KCl concentrations prepared in Examples 1 and 15 to 25 were subjected to Zeta potential tests, and the [AMIM][Cl]-enzyme mixtures with different [AMIM][Cl] concentrations prepared in Examples 26 to 36 were subjected to Zeta potential tests. The test results are shown in FIG. Figure 1 The higher the absolute value of the zeta potential of the salt-enzyme solution, the more stable the solution is and the less likely it is to flocculate and precipitate.

[0052] from Figure 1 It can be seen that with increasing salt concentration, the absolute value of the potential of the salt-enzyme solution first increases and then decreases. The absolute value of the potential is maximum when the KCl concentration is approximately 0.5%, and the absolute value of the potential is maximum when the [AMIm]Cl concentration is approximately 0.6%. Compared with the zeta potential of the KCl-enzyme solution, after the zeta potential of the [AMIm]Cl-enzyme solution reaches its maximum value, the zeta potential is always higher than that of the KCl-enzyme solution. This shows that when the [AMIm]Cl content is high, the [AMIm]Cl-enzyme solution still has high stability. High enzyme solution stability, that is, high enzyme dispersibility, increases the contact probability between the enzyme and the substrate (proteoglycans, mucopolysaccharides, and elastin in the skin), thereby improving the catalytic efficiency of the enzyme. Therefore, the appropriate addition of KCl and [AMIm]Cl is beneficial to increase the charge on the enzyme surface, enhance the interaction between protein molecules and water molecules, increase the solubility of the enzyme in the solution, and thus reduce the enzyme particle size, increase the enzyme's dispersibility, and make the enzyme solution more stable, as manifested by an increase in the absolute value of the zeta potential. On the contrary, higher concentrations of salts destroy the charge structure on the enzyme surface, capture more water molecules inside the enzyme, and cause the enzyme to flocculate and precipitate, which is manifested as a decrease in the absolute value of the Zeta potential.

[0053] (2) Penetration testing

[0054] The KCl-enzyme mixed solution prepared in Example 1 and the [AMIM][Cl]-enzyme mixed solution prepared in Example 26 were selected to perform contact angle tests on cowhide. At the same time, in order to compare the effects of the salt-enzyme mixed solutions of Example 1 and Example 16, the mixed enzyme prepared in step (1) of the example was used for effect comparison, which was recorded as Mixed enzyme solution.

[0055] The contact angle test results are as follows Figure 2 As shown in Figure 2(A), it can be seen that in the first minute, the contact angle of the salt-enzyme mixed solution on the cowhide is greater than that of the enzyme solution. After two minutes, the contact angle of the salt-enzyme mixed solution on the cowhide surface decreases rapidly, especially the [AMIm]Cl-enzyme solution. After 46 minutes, the solution has basically penetrated into the leather. In addition, the KCl-enzyme solution has basically penetrated into the leather at 54 minutes, while the mixed enzyme solution has not completely penetrated into the leather at 60 minutes. At the same time, it can be seen in Figure 2(B) that the diameter of the salt-enzyme mixed solution droplets is larger than that of the mixed enzyme solution droplets, and the wettability is stronger. This is mainly because after the salt is added to the enzyme solution, the osmotic pressure of the solution increases, providing the driving force for the enzyme solution to penetrate into the skin. At the same time, the salt ions in the enzyme solution increase the charge on the surface of the skin tissue, increase the interaction between water molecules and the skin tissue, and thus improve the wettability between the enzyme solution and the skin tissue. Therefore, the appropriate addition of a certain concentration of salt is beneficial to improving the permeability of the enzyme solution, reducing the excessive hydrolysis of the grain surface by the enzyme attached to the skin surface, and increasing the enzyme's hydrolysis of proteoglycans and miscellaneous proteins in the skin, thereby achieving the purpose of dispersing collagen fiber bundles and softening.

[0056] Example 37

[0057] A method for depilating raw hides comprises the following steps: weighing 30 g of degreased, desalted and soaked raw cow hides, adding 90 g of the KCl-enzyme mixed solution prepared in Example 1, and performing depilation for 6 h at a temperature of 32° C. and a rotation speed of 50 r / min using a rotary drum for stirring. The depilation operation is completed.

[0058] Example 38 to Example 50

[0059] The contents of Examples 38 to 50 are substantially the same as those of Example 37, except that the salt-enzyme mixed solutions prepared in Examples 2 to 14 are used for dehairing the raw hides.

[0060] Example 51 to Example 61

[0061] The contents of Examples 51 to 61 are substantially the same as those of Example 37, except that the salt-enzyme mixed solutions prepared in Examples 15 to 25 are used for dehairing the raw hides.

[0062] Example 62

[0063] The content of Example 62 is basically the same as that of Example 37, except that the salt-enzyme mixed solution prepared in Example 26 is used to dehair the raw hide.

[0064] Example 63 to Example 72

[0065] The contents of Examples 63 to 72 are substantially the same as those of Example 37, except that the salt-enzyme mixed solutions prepared in Examples 27 to 36 are used for dehairing the raw hides.

[0066] Example 73

[0067] A method for separating collagen fiber bundles from bare skin, the specific process is as follows:

[0068] Weigh 30 g of the depilated bare skin, add 90 g of the KCl-enzyme mixed solution prepared in Example 1, stir the mixture at 32° C. and 50 r / min to split the collagen fiber bundles for 18 h. This completes the operation of splitting the collagen fiber bundles of the bare skin.

[0069] Example 74 to Example 86

[0070] The contents of Examples 74 to 86 are basically the same as those of Example 73, except that the salt-enzyme mixed solutions prepared in Examples 2 to 14 were used to separate the collagen fiber bundles of bare skin.

[0071] Example 87 to Example 97

[0072] The contents of Examples 87 to 97 are basically the same as those of Example 73, except that the salt-enzyme mixed solutions prepared in Examples 15 to 25 were used to separate the collagen fiber bundles of bare skin.

[0073] Example 98

[0074] The content of Example 98 is basically the same as that of Example 73, except that the salt-enzyme mixed solution prepared in Example 26 is used to separate the collagen fiber bundles of bare skin.

[0075] Example 99 to Example 109

[0076] The contents of Examples 99 to 109 are basically the same as those of Example 73, except that the salt-enzyme mixed solutions prepared in Examples 27 to 36 were used to separate the collagen fiber bundles of bare skin.

[0077] Example 110

[0078] A method for dehairing / splitting collagen fiber bundles of raw hides, comprising the following steps: weighing 30 g of degreased, desalted, and soaked raw cow hide, adding 90 g of the KCl-enzyme mixed solution prepared in Example 1, and performing dehairing / splitting of the collagen fiber bundles for 24 hours at a temperature of 32° C. and a speed of 50 r / min using a rotary drum for stirring. This completes the dehairing / splitting of the collagen fiber bundles of the raw hides.

[0079] Example 111 to Example 123

[0080] The contents of Examples 111 to 123 are substantially the same as those of Example 110, except that the salt-enzyme mixed solutions prepared in Examples 2 to 14 are used to perform raw hide dehairing / collagen fiber bundle splitting.

[0081] Example 124 to Example 134

[0082] The contents of Examples 124 to 134 are substantially the same as those of Example 110, except that the salt-enzyme mixed solutions prepared in Examples 15 to 25 are used to dehair the raw skin / split the collagen fiber bundles.

[0083] Example 135

[0084] The content of Example 135 is basically the same as that of Example 110, except that the salt-enzyme mixed solution prepared in Example 26 is used to dehair the raw skin / split the collagen fiber bundles.

[0085] Example 136 to Example 145

[0086] The contents of Examples 136 to 145 are substantially the same as those of Example 110, except that the salt-enzyme mixed solutions prepared in Examples 27 to 36 are used to perform raw hide dehairing / collagen fiber bundle splitting.

[0087] Example 146

[0088] A method for dehairing / splitting collagen fiber bundles / softening raw hides comprises the following steps: weighing 30 g of degreased, desalted, and soaked raw cow hides, adding 90 g of the KCl-enzyme mixed solution prepared in Example 1, and dehairing / splitting collagen fiber bundles for 30 hours at a temperature of 32° C. and a speed of 50 r / min using a rotary drum for stirring. The dehairing / splitting collagen fiber bundles / softening of the raw hides is completed.

[0089] Example 147 to Example 159

[0090] The contents of Examples 147 to 159 are basically the same as those of Example 146, except that the salt-enzyme mixed solutions prepared in Examples 2 to 14 are used respectively for dehairing / splitting collagen fiber bundles / softening the raw hide.

[0091] Example 160 to Example 170

[0092] The contents of Examples 160 to 170 are basically the same as those of Example 146, except that the salt-enzyme mixed solutions prepared in Examples 15 to 25 are used to dehair / splitting collagen fiber bundles / softening the hide, respectively.

[0093] Example 171

[0094] The content of Example 171 is basically the same as that of Example 146, except that the salt-enzyme mixed solution prepared in Example 26 is used to dehair / splitting collagen fiber bundles / softening the raw hide.

[0095] Example 172 to Example 181

[0096] The contents of Examples 172 to 181 are basically the same as those of Example 146, except that the salt-enzyme mixed solutions prepared in Examples 27 to 36 are used to dehair / splitting collagen fiber bundles / softening the hide, respectively.

[0097] The application performance test was carried out using the tanning salt-enzyme mixed solution prepared in Examples 1 to 36 of the present invention as an example, and the application performance of the tanning salt-enzyme mixed solution in raw hide dehairing, bare hide collagen fiber bundle splitting, raw hide dehairing / splitting collagen fiber bundles / softening was tested. At the same time, the application effect was analyzed through comparative experiments.

[0098] Comparative Example 1: Depilation is performed using the mixed enzyme alone. Specifically, the following steps are performed:

[0099] Comparative Example 1

[0100] A method for depilating raw hides is substantially the same as that of Example 37, except that the mixed enzyme solution prepared in step (1) of Example 1 is used for depilation.

[0101] Comparative Example 2 uses the traditional ash-alkali method to separate the collagen fiber bundles. The specific steps of Comparative Example 2 are as follows:

[0102] Comparative Example 2

[0103] A method for separating collagen fiber bundles from bare hides, comprising the following steps: weighing the dehaired hide, and reliming the hide with 5 wt% calcium hydroxide (based on the hide weight) and 200 wt% water (based on the hide weight) at 25°C with shaking (50 rpm) for 24 hours. Deliming the hide with 200 wt% water and 3 wt% ammonium sulfate (based on the hide weight after liming) at 32°C with shaking (100 rpm) for 60 minutes. The hide is then washed with 200 wt% water for 30 minutes. The hide is then separated into collagen fiber bundles using a conventional ash-alkali method.

[0104] Comparative Example 3 uses traditional methods to remove hair from the raw hide, separate collagen fiber bundles, and soften the raw hide. Specifically, Comparative Example 3 is as follows:

[0105] Comparative Example 3

[0106] A method of dehairing / splitting collagen fiber bundles / softening raw hides, wherein the specific process is as follows:

[0107] Take 15g of fresh cowhide, use 100wt% water, 2.8wt% sodium sulfide and 5wt% calcium hydroxide (based on the weight of wet cowhide) to destroy the hair at 25℃ for 3h. Then, weigh the cowhide, shake at 25℃ (50r / min) for 24h with 5wt% calcium hydroxide and 200wt% water based on the weight of the cowhide. Deliming is done by shaking at 32℃ (100r / min) for 60min with 200wt% water and 3wt% ammonium sulfate (based on the weight of the leather after liming). Wash the bare skin with 200wt% water for 30min. Soften with 200wt% water and 0.06wt% neutral protease at 32℃ (100r / min) for 60min and store at 4℃ to obtain bare skin that has been dehaired / split collagen fiber bundles / softened using traditional methods.

[0108] Comparative Example 4: Using the mixed enzyme alone to perform raw hide dehairing / collagen fiber splitting / softening. Specifically, Comparative Example 4 is as follows:

[0109] Comparative Example 4

[0110] A method for dehairing / splitting collagen fiber bundles / softening raw hides is basically the same as Example 146, except that: the mixed enzyme solution prepared in step (1) of Example 1 is used to dehair / splitting collagen fiber bundles / softening raw hides.

[0111] Application performance testing

[0112] (1) Enzyme tracer test

[0113] The cowhide samples of comparative example 1, example 37 and example 62 after depilation for 6 hours were selected for enzyme tracing test. The specific process of the enzyme tracing was as follows: the mixed enzyme used for depilation was labeled with FITC and used for the salt-enzyme process depilation operation. The cowhide samples with different operations were cut into slices with a thickness of 8 μm by ice cutting, and the slices were moved to a glass slide. A digital slice scanning system was used to observe the penetration and distribution of the mixed enzyme in the cowhide after depilation at an excitation wavelength of 494 nm and an emission wavelength of 527 nm.

[0114] The cowhide of Example 37 and Example 62 before and after depilation for 6 hours Figure 3 and Figure 4 ,Depend on Figure 3 It can be seen that the hair on the cowhide can be removed by using KCl-enzyme mixed solution. Figure 4It can be seen that the [AMIM][Cl]-enzyme mixed solution can be used to remove hair from cowhide. After dehairing, the surface of the leather is smooth, without damage, fine hair, hair roots or hair piles.

[0115] The results of enzyme tracer test are as follows Figure 5 As shown, Figures A, B, and C correspond to the test results of Example 1, Example 37, and Example 62, respectively. Figure 5 It can be seen that when using mixed enzyme hair removal alone, the hair follicle outline is the same as the actual outline, which is relatively clear, and proteoglycans are still present, indicating that the hair roots and hair in the hair follicles have not been removed ( Figure 5 (A)) When the hair is removed with a salt / enzyme system, the hair follicles are deformed and disappear, the hair root structure is gone, and the proteoglycans in the hair follicles are completely hydrolyzed ( Figure 5 (B) and Figure 5 (C)) This indicates that the salt enzyme system hydrolyzes the polysaccharides in the hair follicles quickly, accelerating the rate of hair removal, removing the hair roots, and completing hair removal.

[0116] (2) EVG staining test

[0117] The cross-section of the skin tissue after separating the collagen fiber bundles from the raw cowhide, comparative example 2, example 73 and example 98 bare skin was selected for EVG staining test. The results are as follows Figure 6 and Figure 7 As shown, the blue-black filamentous fibers are elastic fibers, the yellow area is proteoglycan, and the orange-red and pink areas are collagen fibers. Figure 6 A, B, and C correspond to the test results of raw cowhide, comparative example 2, and embodiment 73, respectively. Figure 7 is the test result corresponding to Example 98, Figure 6 It can be seen that after the traditional ash-alkali method is used to separate the collagen fiber bundles, the proteoglycans in the skin group still exist. However, by using the process of Example 73, the proteoglycans in the skin tissue are completely hydrolyzed, which is conducive to the dispersion of the collagen fiber bundles. Figure 7 It can be seen that by adopting the process of Example 98, the proteoglycans and elastic fibers in the skin tissue are completely hydrolyzed, and this process has a good effect of splitting the collagen fiber bundles.

[0118] (3) Scanning electron microscope test

[0119] The leather obtained after dehairing / splitting collagen fiber bundles / softening in Comparative Example 3, Comparative Example 4, Example 146 and Example 171 was subjected to scanning electron microscopy testing. The results are as follows: Figure 8 As shown, Figure 8 A, B, C, and D correspond to the test results of comparative example 3, comparative example 4, embodiment 146, and embodiment 171, respectively. Figure 8 (A) is a traditional craft. Figure 8 (B) is a mixed enzyme process, Figure 8(C) is the KCl-mixed enzyme method, Figure 8 (D) is the [AMIM][Cl]-mixed enzyme method, which is used to observe the dispersion of collagen fiber bundles. Figure 8 In (A) and 8(B), it can be seen that the collagen fiber bundles are thick and the collagen microfibrils are cohesive and not dispersed. Figure 8 (C) and 8 (D) show small, fluffy collagen fiber bundles, with the collagen microfibrils widely dispersed and not sticking together. Therefore, the salt-mixed enzyme process is superior to the traditional and mixed enzyme methods in resolving collagen fiber bundles.

[0120] (4) Thermal and mechanical properties testing

[0121] The crust leather obtained after dehairing / splitting of collagen fiber bundles / softening of Comparative Example 3, Comparative Example 4, Examples 146 to 159, and Example 171 was selected for tensile strength and wet heat shrinkage temperature tests, and the measurements were performed according to the methods described in "GBT 4689.5-1984-Determination of tensile strength and elongation of leather" and "GBT4689.8-1984-Determination of shrinkage temperature of leather", respectively.

[0122] The experimental results are as follows Figure 9 and 10 As shown, comparative example 3 is recorded as Control, comparative example 4 is recorded as Mixed enzyme, Figure 9 It can be seen that the tensile strength of the crust leather prepared by the salt-enzyme system dehairing / collagen fiber splitting / softening bare leather is better than or close to the tensile strength of the traditional process, and better than the tensile strength of the enzyme process. Among them, the tensile strength of the crust leather prepared by the [AMIm]Cl-enzyme method is the highest. Figure 10 It can be seen that the wet heat shrinkage temperature of the crust leather prepared by salt-enzyme system dehairing / splitting collagen fiber bundles / softening bare leather is not much different from that of the traditional process and enzyme process, among which the wet heat shrinkage temperature of the crust leather prepared by [AMIm]Cl-enzyme method is the highest.

Claims

1. A method for preparing a tanning salt-enzyme mixed solution, characterized in that: The following steps are involved: (1) adding neutral protease, α-amylase, peptidase, protease and glucanase into water to prepare a mixed enzyme solution; (2) adding salt substances to the mixed enzyme solution prepared in step (1), mixing them evenly, and obtaining a tanning salt-enzyme mixed solution.

2. The preparation method according to claim 1, characterized in that In step (1), the usage ratio of neutral protease, α-amylase, peptidase, protease, glucanase and water is (2-3.5) g: (1.5-3) g: (0.1-0.4) g: (0.05-0.3) g: (0.03-0.1) g: 1 L.

3. The preparation method according to claim 2, characterized in that In step (2), the ratio of the salt substance to the mixed enzyme solution is (2.5-12) g:1 L, and the salt substance is an inorganic salt, an ionic liquid or a deep eutectic solvent.

4. The preparation method according to claim 3, characterized in that The inorganic salt is one or more of Na2SO2, KCl, MgCl2, NH4SO4, NH4Cl, ZnCl2, ZnSO4, Na2HPO4, NaCl, CaCl2, CH3COONa, Na2MoO4, NaHCO3, Na2SiO3, BaCl2, and LiBr; the ionic liquid is one or more of imidazole ionic liquids, quaternary ammonium ionic liquids, pyridine ionic liquids, quaternary phosphonium ionic liquids, and functionalized ionic liquids; the deep eutectic solvent is one or more of choline chloride-urea, choline chloride-carboxylic acid, choline chloride-polyol, and zinc chloride-urea.

5. A tanning salt-enzyme mixed solution prepared by the method according to any one of claims 1 to 4.

6. Use of the tanning salt-enzyme mixed solution according to claim 5 in dehairing, splitting collagen fiber bundles and / or softening animal skins.

7. The use according to claim 6, characterized in that The animal skin dehairing method comprises: mixing the animal skin with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), stirring at a temperature of 25-35° C. for 2-6 hours.

8. The use according to claim 6, characterized in that The method for splitting the collagen fiber bundles is as follows: mixing the dehaired animal skin with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), stirring at a temperature of 25-35° C. for 1-24 hours.

9. The use according to claim 6, characterized in that The method for dehairing / splitting collagen fiber bundles from animal skin comprises: mixing the animal skin with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), stirring at a temperature of 25-35° C. for 2-24 hours.

10. The use according to claim 6, characterized in that The method for dehairing / splitting collagen fiber bundles / softening animal skins comprises: mixing the animal skins with a tanning salt-enzyme mixed solution in a mass ratio of 1:(3-5), stirring at a temperature of 25-40° C. for 6-30 hours.

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