Gelatin-chlorogenic acid composite antifreeze agent as well as preparation method and application thereof
A composite antifreeze agent was prepared by covalently combining gelatin and chlorogenic acid using an alkaline method. This solved the problem of fish paste quality degradation caused by traditional antifreeze agents, and achieved efficient, safe, and environmentally friendly fish paste freezing protection, which is suitable for the freezing and preservation of aquatic products.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-01-06
- Publication Date
- 2026-04-10
AI Technical Summary
Traditional commercial antifreeze agents cause a decline in the quality of fish paste during freezing and do not meet consumers' demands for low sweetness and health. Therefore, finding new, efficient, healthy, and green antifreeze agents has become an important issue.
A stable covalent complex of gelatin and chlorogenic acid was formed by covalent bonding through an alkaline method to prepare a gelatin-chlorogenic acid composite antifreeze agent. The gelatin-chlorogenic acid covalent complex was formed by dialysis and freeze-drying processes, which significantly improved the antifreeze properties and safety.
Gelatin-chlorogenic acid composite antifreeze agent significantly improves antifreeze efficacy and safety during the freezing process of surimi, reduces environmental pollution, improves the quality changes of surimi during frozen storage, and enhances the protection efficiency of frozen fish.
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Figure CN121817257A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the technical field of frozen product preservation, and particularly relates to a gelatin-chlorogenic acid composite antifreeze and a preparation method and application thereof. BACKGROUND
[0002] Frozen storage is an important method for preserving surimi, but improper freezing method can cause denaturation of fish muscle fiber protein, thereby causing quality decline of surimi. Frozen surimi is affected by various factors such as freezing time, freezing temperature, freezing and thawing cycle, etc., resulting in quality decline of surimi in storage and other processing processes. Adding an antifreeze is considered to be one of the most effective methods for preventing denaturation of water proteins during freezing.
[0003] The most commonly used traditional commercial antifreeze is a compound of 4% sucrose and 4% sorbitol, which leads to excessive sweetness and excessive calorie intake, which does not meet the requirements of consumers for low sweetness and low calories and natural health of food in recent years. Therefore, finding a new type of efficient, healthy and green antifreeze has become a big problem in the storage and transportation process of surimi products, which has important economic benefits and practical significance. SUMMARY
[0004] The purpose of the present application is to solve the problems of the prior art, and to provide a gelatin-chlorogenic acid composite antifreeze and a preparation method and application thereof. The following technical solutions are used: In a first aspect, the present application provides a preparation method of a gelatin-chlorogenic acid composite antifreeze, comprising the following steps: dissolving gelatin in water, then adjusting pH to 8-12 with NaOH solution, and stirring uniformly to obtain a gelatin solution; dissolving chlorogenic acid in water to obtain a chlorogenic acid solution; mixing the gelatin solution and the chlorogenic acid solution, adjusting pH to 8-12 with NaOH solution, and stirring uniformly to obtain a gelatin-chlorogenic acid composite solution; dialyzing the gelatin-chlorogenic acid composite solution, and freeze-drying to obtain the gelatin-chlorogenic acid composite antifreeze.
[0005] The present application provides a method for preparing a composite antifreeze from gelatin and chlorogenic acid. The method uses gelatin as raw material and covalently combines chlorogenic acid by alkali method. The product is dialyzed and freeze-dried to obtain a gelatin-chlorogenic acid covalent compound, which is a high-efficiency antifreeze. The antifreeze efficiency and safety are significantly improved, and the raw material used is green and environmentally friendly, reducing environmental pollution.
[0006] The gelatin and chlorogenic acid are combined by covalent binding to form stable amide bonds, the high bond energy of the covalent bonds makes the structure of the combined product stable, effectively avoids the loss of chlorogenic acid, does not damage the molecular structure of the gelatin, realizes the combination of the active groups of chlorogenic acid, and makes the covalent product have the gelation, film forming and other characteristics of gelatin and the antioxidant activity of chlorogenic acid, realizes the functional complementation and synergistic effect, and the anti-freezing effect of the covalent combined product of gelatin-chlorogenic acid is significantly improved.
[0007] As a further preferred embodiment, the concentration of the gelatin solution is 76.6 mM-19.2 mM; and the concentration of the chlorogenic acid solution is 0.4 mM-19.2 mM.
[0008] As a further preferred embodiment, the specific steps of the dialysis are as follows: The gelatin-chlorogenic acid complex solution is placed in a dialysis bag, and then dialysis is performed in an ice bath.
[0009] As a further preferred embodiment, the dialysis bag is a dialysis bag with a molecular weight cut-off of 500 Da-2000 Da.
[0010] As a further preferred embodiment, the temperature of the ice bath is 2℃-8℃.
[0011] As a further preferred embodiment, the method further comprises the following steps before the freeze-drying: The gelatin-chlorogenic acid complex solution after dialysis is first placed in a-45℃~-85℃ refrigerator for pre-freezing for 2 h-10 h, and then is placed in a freeze dryer for freeze-drying after forming a solid.
[0012] As a further preferred embodiment, the conditions of the freeze-drying are as follows: The material layer thickness is 1 mm-6 mm, the freezing temperature is-40℃~-80℃, the drying pressure is 5 Pa-20 Pa, the baffle temperature is 30℃-60℃, and the drying time is 24 h-60 h.
[0013] In a second aspect, the application provides a gelatin-chlorogenic acid complex antifreezing agent prepared by the above preparation method.
[0014] In a third aspect, the application provides the use of the above gelatin-chlorogenic acid complex antifreezing agent in the frozen storage of aquatic products.
[0015] As a further preferred embodiment, the aquatic products include surimi.
[0016] The application has the following beneficial effects: (1) The gelatin-chlorogenic acid composite antifreeze agent is prepared by alkali covalent cross-linking, the process raw material cost is low, and the whole preparation process is simple and easy to control;The raw materials of gelatin and chlorogenic acid are abundant in source, easy to obtain, low in cost, green and environmentally friendly, and the environmental pollution is reduced; (2) The composite antifreeze agent prepared by the preparation method of the application is prepared by taking gelatin and chlorogenic acid as raw materials, the covalent modification of gelatin and chlorogenic acid changes the hydrophilic property of gelatin, improves the surface hydrophilicity, and the chlorogenic acid is a kind of polyphenol compound with multiple pharmacological activities, has antioxidant effect, and the prepared composite antifreeze agent has significantly enhanced antifreeze performance, and can be applied to various frozen systems as antifreeze. BRIEF DESCRIPTION OF DRAWINGS
[0017] In order to more clearly illustrate the technical solutions in the embodiments of the application, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the application, and all other drawings obtained by those skilled in the art without creative labor are within the scope of protection of the application.
[0018] Figure 1 The myofibrillar protein content of the fish paste after the antifreeze agent prepared in example 1-3 and comparative example 1-2 is used for the cold frozen freeze-thaw cycle of the fish paste of silver carp; Figure 2 The turbidity of the fish paste after the antifreeze agent prepared in example 1-3 and comparative example 1-2 is used for the cold frozen freeze-thaw cycle of the fish paste of silver carp; Figure 3 The Ca 2+ ATPase activity of the fish paste after the antifreeze agent prepared in example 1-3 and comparative example 1-2 is used for the cold frozen freeze-thaw cycle of the fish paste of silver carp; Figure 4 The total sulfhydryl content of the fish paste after the antifreeze agent prepared in example 1-3 and comparative example 1-2 is used for the cold frozen freeze-thaw cycle of the fish paste of silver carp; Figure 5 The active sulfhydryl content of the fish paste after the antifreeze agent prepared in example 1-3 and comparative example 1-2 is used for the cold frozen freeze-thaw cycle of the fish paste of silver carp. DETAILED DESCRIPTION
[0019] The technical solutions in the embodiments of the application will be described clearly and completely below with reference to the drawings in the embodiments of the application. Obviously, the described embodiments are only some embodiments of the application, not all embodiments. Based on the embodiments in the application, all other embodiments obtained by those skilled in the art without creative labor are within the scope of protection of the application.
[0020] Example 1 A preparation method of a gelatin-chlorogenic acid composite antifreezing agent, which specifically comprises the following steps: (1) 0.25 g of gelatin is added to deionized water to prepare a gelatin solution with a concentration of 19.2 mM, then 0.1 M NaOH is used to adjust the pH to 9, and finally it is placed on a room temperature magnetic stirrer for constant speed (120 rpm) stirring for 24 h; (2) A 0.4 mM chlorogenic acid solution is prepared according to the method in step (1) (chlorogenic acid is added to deionized water), without stirring for 24 h, it is directly mixed with the gelatin solution, 0.1 M NaOH is used to adjust the pH to 9, and it is continuously stirred at a constant speed (120 rpm) on a magnetic stirrer exposed to air at room temperature for 24 h, and a gelatin-chlorogenic acid composite solution is obtained by crosslinking; (3) The gelatin-chlorogenic acid composite solution in step (2) is dialyzed in a dialysis bag with a molecular weight cut-off of 500 Da, deionized water is used, and it is placed in a 4°C refrigerator ice bath for 48 h, and the deionized water is replaced every 6 h to remove free chlorogenic acid, and a covalent solution is obtained, wherein the volume ratio of the composite solution to pure water is 1:3 v / v; (4) The covalent solution in step (3) is taken in a box-shaped plastic container, and pre-frozen for 4 h in a -80°C refrigerator, until it becomes a solid, wherein the amount of the covalent solution in the plastic container satisfies the following condition: no more than 1 / 3 of the volume of the container; the solid composite after the first stage of pre-freezing is subjected to freeze-drying, wherein the freeze-drying conditions are: material layer thickness of 4 mm, freezing temperature of -60°C, vacuum degree of 10 Pa during drying, baffle temperature of 45°C, and drying time of 48 h, and it is taken out after 2 days, to obtain a gelatin-chlorogenic acid covalent composite product.
[0021] The gelatin-chlorogenic acid covalent composite product prepared above is used for white carp surimi freezing, and the effect test is carried out, and the specific process is as follows: (1) The composite product prepared above is taken, and 1% of the composite product by weight of white carp surimi is added, to obtain white carp surimi with added antifreezing agent; (2) The white carp surimi treated with the antifreezing agent in step (1) is subjected to frozen storage, and the frozen storage condition is: -20°C. The white carp surimi is subjected to freeze-thaw cycle, and the freeze-thaw cycle condition is: frozen in a -20°C refrigerator for 10 d, and thawed in a 4°C refrigerator for 12 h, for a total of 4 cycles; (3) The myofibrillar protein content, turbidity, Ca 2+ ATPase activity, total sulfhydryl content, and active sulfhydryl content of the surimi after freeze-thaw cycle in step (2) are monitored.
[0022] Example 2 A preparation method of a gelatin-chlorogenic acid composite antifreezing agent, which specifically comprises the following steps: (1) 1 g gelatin was added into deionized water to prepare a gelatin solution with a concentration of 76.6 mM, then the pH was adjusted to 9 using 0.1 M NaOH, and finally the solution was stirred at a constant speed (120 rpm) on a magnetic stirrer at room temperature for 24 h; (2) A 19.2 mM chlorogenic acid solution was prepared according to the method in step (1) (chlorogenic acid was added into deionized water), without stirring for 24 h, and directly mixed with the gelatin solution. The pH was adjusted to 9 using 0.1 M NaOH, and the mixture was continuously stirred at a constant speed (120 rpm) on a magnetic stirrer at room temperature for 24 h under exposure to air to obtain a gelatin-chlorogenic acid complex solution; (3) The gelatin-chlorogenic acid complex solution in step (2) was dialyzed using a dialysis bag with a molecular weight cut-off of 500 Da, deionized water was used, and the dialysis was performed in a refrigerator at 4 °C for 48 h, with the deionized water being replaced every 6 h to remove free chlorogenic acid, to obtain a covalent solution, wherein the volume ratio of the complex solution to pure water was 1:3 v / v.
[0023] (4) The covalent solution in step (3) was pre-frozen in a box-shaped plastic container in a refrigerator at -80 °C for 4 h until it became solid, wherein the amount of the complex solution in the plastic container met the following condition: not more than 1 / 3 of the volume of the container. The solid complex after the first stage of pre-freezing was subjected to freeze-drying, wherein the freeze-drying conditions were as follows: the thickness of the material layer was 4 mm, the freezing temperature was -60 °C, the vacuum degree during drying was 10 Pa, the temperature of the baffle was 45 °C, and the drying time was 48 h. After 2 days, the product was taken out, and a gelatin-chlorogenic acid covalent complex product was obtained.
[0024] The gelatin-chlorogenic acid covalent complex product prepared above was used for the freezing of silver carp surimi, and the effect was tested, with the specific process being as follows: (1) The complex product prepared above was added to white carp surimi at a weight ratio of 1% to obtain white carp surimi with an antifreeze agent added.
[0025] (2) The white carp surimi treated with the antifreeze agent in step (1) was subjected to frozen storage, and the frozen storage conditions were as follows: -20 °C. The white carp surimi was subjected to freeze-thaw cycles, and the freeze-thaw cycle conditions were as follows: frozen in a refrigerator at -20 °C for 10 d, thawed in a refrigerator at 4 °C for 12 h, and a total of 4 cycles.
[0026] (3) The myofibrillar protein content, turbidity, Ca 2+ ATPase activity, total sulfhydryl content, and active sulfhydryl content of the surimi after the freeze-thaw cycles in step (2) were monitored.
[0027] Example 3 A preparation method of a gelatin-chlorogenic acid complex antifreeze agent, which specifically comprises the following steps: (1) Add 0.25 g of gelatin to deionized water to prepare a gelatin solution with a concentration of 19.2 mM. Then, use 0.1 M NaOH to adjust the pH to 9. Finally, place it on a room temperature magnetic stirrer and stir at a constant speed (120 rmp) for 24 h. (2) Prepare a 0.4 mM chlorogenic acid solution according to the method in step (1), without stirring for 24 h (add chlorogenic acid to deionized water), and mix it directly with the gelatin solution. Adjust the pH to 9 with 0.1 M NaOH, and stir continuously at a constant speed (120 rpm) for 24 h at room temperature and exposed to air to obtain a gelatin-chlorogenic acid composite solution. (3) Dialyze the mixture in step (2) using a dialysis bag with a molecular weight cutoff of 1000 Da, using deionized water, and in a 4°C refrigerator ice bath for 48 h. Replace the deionized water every 6 h to remove free chlorogenic acid and obtain a covalent solution, wherein the volume ratio of the composite liquid to the pure water is 1:3 v / v.
[0028] (4) Take the covalent solution from step (3) into a box-type plastic container and pre-freeze it in a -80℃ refrigerator for 4 hours until it becomes solid. The amount of covalent solution in the plastic container meets the following conditions: it does not exceed 1 / 3 of the container volume. Take the solid composite after the first stage of pre-freezing and freeze-dry it. The freeze-drying conditions are: material layer thickness 4 mm, cold sink temperature -60℃, vacuum degree 10 Pa during drying, partition temperature 45℃, drying time 48 hours, and take it out after 2 days to obtain the gelatin-chlorogenic acid covalent composite product.
[0029] The gelatin-chlorogenic acid covalent complex product prepared above was used for freezing silver carp surimi, and the effect was tested. The specific process is as follows: (1) Take the complex product prepared above and add 1% of the weight of silver carp surimi to the complex to obtain silver carp surimi with added antifreeze.
[0030] (2) The silver carp surimi treated with the antifreeze agent in step (1) is frozen and stored under the following conditions: -20℃. The silver carp surimi is subjected to freeze-thaw cycles under the following conditions: frozen at -20℃ for 10 days and thawed at 4℃ for 12 hours, for a total of 4 cycles.
[0031] (3) Monitor the myofibrillar protein content, turbidity, and Ca content of the fish surimi after freeze-thaw cycles in step (2). 2+ ATPase activity, total thiol content, and active thiol content.
[0032] Comparative Example 1 A method for preparing a gelatin antifreeze agent, specifically comprising the following steps: (1) Add 0.25 g of gelatin to deionized water to prepare a gelatin solution with a concentration of 19.2 mM. Then place the gelatin solution on a magnetic stirrer at room temperature and stir at a constant speed (120 rmp) for 24 h. (2) Take the gelatin solution from step (1) and place it in a box-type plastic container. Pre-freeze it in a -80℃ freezer for 4 hours until it becomes solid. The amount of gelatin solution in the plastic container should meet the following conditions: not exceeding 1 / 3 of the container volume. Take the solid material after the first stage of pre-freezing and freeze-dry it. The freeze-drying conditions are: material layer thickness 4 mm, cold sink temperature -60℃, vacuum degree 10 Pa during drying, partition temperature 45℃, and drying time 48 hours. Take it out after 2 days to obtain the gelatin antifreeze product.
[0033] The gelatin antifreeze product prepared above was used for freezing silver carp surimi, and the effect was tested. The specific process is as follows: (1) Take the gelatin antifreeze product prepared above and add 1% of the weight of silver carp surimi to the complex to obtain silver carp surimi with added antifreeze; (2) The silver carp surimi treated with the antifreeze agent described in step (5) is frozen and stored under the following conditions: -20℃. The silver carp surimi is subjected to freeze-thaw cycles under the following conditions: frozen at -20℃ for 10 days and thawed at 4℃ for 12 hours, for a total of 4 cycles. (3) Monitor the myofibrillar protein content, turbidity, and Ca content of the fish surimi after freeze-thaw cycles in step (6). 2+ ATPase activity, total thiol content, and active thiol content.
[0034] Comparative Example 2 A method for preparing a chlorogenic acid antifreeze agent, specifically comprising the following steps: (1) Add 7.1 mg of chlorogenic acid to deionized water to prepare a chlorogenic acid solution with a concentration of 0.4 mM. Then place the chlorogenic acid solution on a room temperature magnetic stirrer and stir at a constant speed (120 rmp) for 24 h.
[0035] (2) Take the chlorogenic acid solution from step (1) and place it in a box-type plastic container. Pre-freeze it in a -80℃ refrigerator for 4 hours until it becomes solid. The amount of solution in the plastic container should meet the following conditions: not more than 1 / 3 of the container volume. Take the solid material after the first stage of pre-freezing and freeze-dry it. The freeze-drying conditions are: material layer thickness 4 mm, cold sink temperature -60℃, vacuum degree 10 Pa during drying, partition temperature 45℃, drying time 48 hours, and take it out after 2 days to obtain the chlorogenic acid antifreeze product.
[0036] The chlorogenic acid antifreeze product prepared above was used for freezing silver carp surimi, and the effect was tested. The specific process is as follows: (1) Take the chlorogenic acid antifreeze product prepared above and add 1% of the weight of silver carp surimi to the complex to obtain silver carp surimi with added antifreeze.
[0037] (2) The silver carp surimi treated with the antifreeze agent described in step (1) is frozen and stored under the following conditions: -20℃. The silver carp surimi is subjected to freeze-thaw cycles under the following conditions: frozen at -20℃ for 10 days and thawed at 4℃ for 12 hours, for a total of 4 cycles. (3) Monitor the myofibrillar protein content, turbidity, and Ca content of the fish surimi after freeze-thaw cycles in step (2). 2+ ATPase activity, total thiol content, and active thiol content.
[0038] The antifreeze properties of the products from Comparative Examples 1-3 and 1-2 were obtained by adding different products to silver carp surimi in proportion, and conducting several freeze-thaw experiments to test the myofibrillar protein content, turbidity, and Ca content. 2+ ATPase activity, total sulfhydryl content, and active sulfhydryl content were tested using methods referenced from the 2019 Master's thesis at Shanghai Ocean University, "Screening and Mechanism of Action Study of Novel Antifreeze Agents for Frozen Tilapia Surimi." The results are as follows: Figures 1-5 As shown. By Figures 1-5 As can be seen from Comparative Examples 1-3 and 1-2, with the increase of freeze-thaw cycles, the present invention can effectively alleviate the problem of reduced surimi quality during freeze-thaw cycles and effectively improve surimi quality. Example 1: The gelatin-chlorogenic acid alkaline covalent bonding method of the present invention significantly improves the antifreeze efficacy and safety of the resulting covalent, and its raw materials are green and environmentally friendly, reducing environmental pollution. The concentrations of gelatin and chlorogenic acid are key factors affecting their alkaline covalent bonding. The gelatin concentration determines the abundance and distribution of amino reaction sites, while the chlorogenic acid concentration regulates the supply of carboxyl sites and oxidative polymerization side reactions. The ratio of their concentrations directly affects the covalent bonding rate and product structure. A suitable concentration can maximize the covalent bonding efficiency and concentrate the molecular weight distribution of the product.
[0039] The core of the alkaline covalent modification of gelatin and chlorogenic acid lies in the dehydration condensation of free amino groups in gelatin and carboxyl groups in chlorogenic acid under alkaline conditions to form amide bonds. This process is accompanied by side reactions such as the self-oxidative polymerization of chlorogenic acid and intermolecular cross-linking of gelatin. The dialysis bag plays a crucial regulatory role in the cross-linking reaction, enabling dynamic control of system purification, small molecule removal, and reaction equilibrium. Its molecular weight cutoff is the core regulatory variable in this cross-linking process and a key factor affecting experimental results. Therefore, selecting an appropriate molecular weight cutoff can significantly improve the covalent binding rate of gelatin and chlorogenic acid, resulting in a more concentrated molecular weight distribution of the covalent product, reducing the residue of small molecule peptides, maximizing the utilization rate of chlorogenic acid, and effectively preventing the dialysis loss of free chlorogenic acid.
[0040] In summary, the gelatin-chlorogenic acid covalent complex prepared by the method provided in this invention exhibits excellent antifreeze properties, effectively increases the protein content retention of surimi, and significantly improves the quality changes of surimi during frozen storage, thereby greatly enhancing the protection efficiency of frozen fish. The prepared antifreeze agent can be widely and extensively applied to the antifreeze protection of frozen surimi.
[0041] The embodiments of this application have been described above with reference to the accompanying drawings. Specific examples have been used to illustrate the principles and implementation methods of this application. The description of the above embodiments is only for the purpose of helping to understand the core ideas of this application. However, this application is not limited to the specific embodiments described above. The specific embodiments described above are merely illustrative and not restrictive. Those skilled in the art can make many other forms under the guidance of this application without departing from the spirit and scope of the claims, and all of these forms are within the protection scope of this application.
Claims
1. A method for preparing a gelatin-chlorogenic acid composite antifreeze agent, characterized in that, Includes the following steps: Dissolve gelatin in water, then adjust the pH to 8-12 with NaOH solution, stir well to obtain gelatin solution; Chlorogenic acid is dissolved in water to obtain a chlorogenic acid solution; The gelatin solution and the chlorogenic acid solution were mixed, and the pH was adjusted to 8-12 with NaOH solution. The mixture was stirred until homogeneous to obtain a gelatin-chlorogenic acid composite solution. The gelatin-chlorogenic acid composite solution was dialyzed and freeze-dried to obtain the gelatin-chlorogenic acid composite antifreeze agent.
2. The preparation method according to claim 1, characterized in that, The concentration of the gelatin solution is 76.6 mM to 19.2 mM; the concentration of the chlorogenic acid solution is 0.4 mM to 19.2 mM.
3. The preparation method according to claim 1, characterized in that, The specific steps of dialysis are as follows: Place the gelatin-chlorogenic acid composite solution in a dialysis bag and then dialyze in an ice bath.
4. The preparation method according to claim 3, characterized in that, The dialysis bag is a dialysis bag with a molecular weight cutoff of 500 Da-2000 Da.
5. The preparation method according to claim 4, characterized in that, The temperature during the ice bath is 2℃~8℃.
6. The preparation method according to claim 1, characterized in that, The freeze-drying process also includes the following steps: The gelatin-chlorogenic acid composite solution after dialysis was first pre-frozen in a refrigerator at -45℃ to -85℃ for 2 h to 10 h to form a solid state, and then placed in a freeze dryer for freeze drying.
7. The preparation method according to claim 6, characterized in that, The freeze-drying conditions are as follows: The material layer thickness is 1 mm to 6 mm, the cold sink temperature is -40℃ to -80℃, the drying pressure is 5 Pa to 20 Pa, the partition temperature is 30℃ to 60℃, and the drying time is 24 h to 60 h.
8. A gelatin-chlorogenic acid composite antifreeze agent, characterized in that, It is prepared by the preparation method according to any one of claims 1-7.
9. The application of the gelatin-chlorogenic acid composite antifreeze agent according to claim 8 in the frozen preservation of aquatic products.
10. The application according to claim 9, characterized in that, The aquatic products include fish paste.