A method for preparing seaweed umami peptide by dynamic high pressure microfluidization homogenization assisted enzymatic hydrolysis

Abstract

This invention discloses a method for preparing seaweed umami peptides through dynamic high-pressure microfluidic homogenization-assisted enzymatic hydrolysis, belonging to the field of umami peptide preparation technology. The method includes: soaking dried seaweed, deodorizing it, and washing it clean to obtain pretreated seaweed; pulping and homogenizing it to obtain an initial seaweed homogenate; enzymatic hydrolysis to obtain a seaweed hydrolysate; mixing it with xylose and cysteine, and performing a Maillard reaction to obtain a seaweed Maillard solution; filtering to obtain a sample solution; ultrafiltration using ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa, and <1kDa; and spray drying to obtain seaweed umami peptides. This invention provides a method for preparing umami peptides that is energy-efficient, low-cost, highly efficient, and environmentally friendly, resulting in higher quality umami peptides.

Description

Technical Field

[0001] This invention belongs to the field of umami peptide preparation technology, and particularly relates to a method for preparing seaweed umami peptides by dynamic high-pressure microfluidic homogenization-assisted enzymatic hydrolysis. Background Technology

[0002] With social and economic development, healthy eating has become a major trend, and people are increasingly choosing natural seasonings. Umami peptides are a type of natural seasoning characterized by safety, lack of pollution, nutritional value, health benefits, high temperature resistance, high stability, and good solubility. They not only bring delicious flavor to food but also reduce bitterness, playing a significant auxiliary role. Furthermore, they can meet the low-salt dietary needs of certain groups (such as those with hypertension), thus being widely used in food as a food additive and flavor enhancer. Umami often exhibits a synergistic effect with other flavor compounds; therefore, methods such as enzymatic hydrolysis are used to prepare complex flavor compounds for food enhancement. Currently, umami peptides are mainly obtained from seafood and then processed into flavor enhancers to improve the umami flavor of food and satisfy people's taste preferences.

[0003] Enzymatic hydrolysis is a method that uses proteases to hydrolyze protein peptide bonds to obtain polypeptides. Because proteases have the ability to specifically recognize and cleave amino acid sites, they offer the potential for controlled preparation of polypeptides with specific structural characteristics. Furthermore, protease reactions are mild, derived from natural and safe sources, and are easily terminated, making them suitable for large-scale application and representing the most promising route for preparing umami peptides. However, current research on the preparation technology of umami peptides from seafood is limited. Traditional enzymatic hydrolysis techniques for seaweed products mostly employ cellulose hydrolysis, a biological method that is time-consuming, energy-intensive, and generally ineffective. Therefore, physical hydrolysis techniques are becoming a more scientific and rational trend. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention proposes a method for preparing seaweed umami peptides through dynamic high-pressure microfluidic homogenization-assisted enzymatic hydrolysis. This method provides a umami peptide preparation method that is low in energy consumption, low in cost, high in preparation efficiency, low in environmental pollution, and produces umami peptides of better quality.

[0005] To achieve the above objectives, this invention provides a method for preparing seaweed umami peptides by dynamic high-pressure microfluidic homogenization-assisted enzymatic hydrolysis, comprising the following steps:

[0006] (1) Soak dried seaweed to remove the fishy smell, wash it clean with water, and obtain pre-treated seaweed;

[0007] (2) The pretreated seaweed obtained in step (1) is pulped, homogenized, and the initial seaweed homogenate is obtained.

[0008] (3) The initial seaweed homogenate obtained in step (2) was adjusted to pH 7-8 and enzymatically hydrolyzed to obtain seaweed hydrolysate.

[0009] (4) The seaweed hydrolysate obtained in step (3) is adjusted to pH 7-11, mixed with xylose and cysteine, and subjected to Maillard reaction to obtain seaweed Maillard solution.

[0010] (5) The seaweed Maillard solution obtained in step (4) was filtered to obtain a sample solution. It was then ultrafiltered using ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa, and spray-dried to obtain seaweed umami peptides.

[0011] Preferably, the dried seaweed in step (1) is one or more of dried kelp, dried laver, dried wakame, dried sea lettuce, or dried agaric.

[0012] Preferably, the soaking in step (1) is carried out in water for 2 to 4 hours; the deodorization in step (1) is carried out by soaking in an acetic acid solution with a mass concentration of 4% to 6% for 20 to 40 minutes.

[0013] Preferably, the pulping process in step (2) involves adding water at 15 to 20 times the weight of the pretreated seaweed and pulping for 5 to 10 minutes.

[0014] Preferably, the homogenization in step (2) is dynamic high-pressure microjet homogenization, the pressure of which is 10000-15000 PSI, and the homogenization is repeated once; the particle size of the seaweed in the initial seaweed homogenate in step (2) is <1μm.

[0015] Preferably, the enzymatic hydrolysis in step (3) uses flavor protease, the amount of flavor protease added is 4wt% to 6wt% of the pretreated seaweed, the enzymatic hydrolysis time is 3 to 4 hours, and the enzymatic hydrolysis temperature is 50 to 60°C.

[0016] Preferably, the amount of xylose used in step (4) is 12wt% to 16wt% of the pretreated seaweed mass; the amount of cysteine ​​used in step (4) is 10wt% to 14wt% of the pretreated seaweed mass; the temperature of the Maillard reaction in step (4) is 100 to 120°C; and the time of the Maillard reaction is 100 to 150 min.

[0017] Preferably, the filtration in step (5) includes filtration using four layers of gauze and filtration using a 0.45μm organic filter membrane.

[0018] Preferably, in step (5), the air intake volume of the spray drying is 65%, the temperature of the spray drying is 190°C, and the feed rate of the spray drying is 35%.

[0019] This invention provides a method for preparing seaweed umami peptides to obtain seaweed umami peptides.

[0020] Compared with the prior art, the present invention has the following advantages and technical effects:

[0021] (1) This invention utilizes dynamic high-pressure micro-jet homogenization technology to break down and dissolve seaweed cells, achieving nanoscale particle size and making the internal proteins more easily degraded by enzymes, thus significantly improving enzymatic hydrolysis efficiency and eliminating the need for the traditional first step of cellulase hydrolysis. Dynamic high-pressure micro-jet homogenization technology is a highly efficient, stable, and safe physical cell wall breaking technology for nanoscale material processing. It is a high-pressure processing technology based on ultra-high pressure theory, fluid mechanics theory, and impact theory. It utilizes the airflow generated by pressure to subject the material to shearing, collision, vortexing, and cavitation, resulting in material homogenization, ultra-micronization, and microemulsification. Dynamic high-pressure micro-jet homogenization technology is a new homogenization method that can be operated continuously, simultaneously achieving a series of operations such as material delivery, mixing, ultra-micronization, pressurization, and homogenization in a short time. Under the impetus of instantaneous pressure, the flowing mixture is subjected to a series of combined forces such as strong impact, high-frequency shearing, high-speed oscillation, and cavitation explosion within the interaction cavity, achieving shearing, collision, or agglomeration effects, thereby achieving the purpose of pulverization and achieving nanoscale particle size. Seaweed cell walls are rich in vitamins, and dynamic high-pressure microfluidic homogenization technology can be used to optimize the cell wall breaking effect during enzymatic hydrolysis, thereby increasing the dissolution rate of internal proteins and significantly improving the efficiency of subsequent enzymatic hydrolysis. Combined with proteolytic hydrolysis and Maillard reaction, seaweed umami peptides can be extracted and prepared, thus providing a certain theoretical basis and value for the development of natural seasonings.

[0022] (2) The umami peptide products prepared by the process provided by the present invention have better quality, flavor and taste. Traditional methods for preparing umami peptides from seafood usually require a long enzymatic hydrolysis time, while dynamic high pressure microfluidic homogenization technology can significantly shorten the enzymatic hydrolysis time and accelerate the cell wall breaking speed of seaweed, which can not only improve production efficiency, but also save time and cost.

[0023] (3) The process provided by the present invention does not require the addition of any chemical reagents or solvents, which reduces environmental pollution. At the same time, the energy consumption required by the technology is relatively low, with an energy consumption of 1908±356.4kJ / h, a soluble nitrogen release rate of up to 85.03±0.47%, and a material loss rate of 1.48±0.02%, which helps to reduce production costs and meets the requirements of sustainable development. Attached Figure Description

[0024] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0025] Figure 1 Radar graph showing the sensory evaluation of the kelp umami peptides prepared in Example 1;

[0026] Figure 2 Radar graph showing the sensory evaluation of the laver umami peptides prepared in Example 2;

[0027] Figure 3 Radar graph for sensory evaluation of the wakame umami peptides prepared in Example 3;

[0028] Figure 4 Radar graph for sensory evaluation of the sea lettuce umami peptides prepared in Example 4;

[0029] Figure 5 Radar graph for sensory evaluation of the agaric umami peptides prepared in Example 5;

[0030] Figure 6 This is a flowchart illustrating the preparation process of the seaweed umami peptide of the present invention. Detailed Implementation

[0031] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0032] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0033] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0034] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0035] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0036] Example 1

[0037] (1) Soak dried kelp in water for 3 hours to rehydrate, then soak in acetic acid solution with a mass concentration of 5% for 30 minutes to remove the fishy smell, and wash it clean to obtain pre-treated kelp.

[0038] (2) Add 40g of pretreated kelp to 800mL of purified water, and beat it for 10min with a pulping machine. Set the dynamic high pressure micro-jet homogenizer pressure to 15000PSI for homogenization and cycle once to obtain an initial kelp homogenate with a particle size of <1μm.

[0039] (3) The initial homogenate of kelp was adjusted to pH 8, 2.4g of flavor protease with an enzyme activity of 20000U / g was added, and the mixture was enzymatically hydrolyzed at 60℃ for 3.5h. The enzyme was then inactivated by placing it in boiling water for 10min to obtain kelp enzymatic hydrolysate.

[0040] (4) Adjust the pH of the kelp enzymatic hydrolysate to 11, mix it with 6.4g xylose and 5.6g cysteine, and perform a Maillard reaction at 120℃ for 150min to obtain kelp Maillard solution;

[0041] (5) The kelp Maillard solution was cooled to room temperature and filtered with four layers of gauze. Then, the supernatant was filtered with a 0.45 μm organic filter membrane to obtain the sample solution. Ultrafiltration was performed using ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa. The solution was then spray-dried (the inlet air volume of the spray dryer was 65%, the temperature was 190℃, and the feed rate was 35%) to obtain kelp umami peptides of three components. The obtained peptides were packaged at room temperature using a heat shrink film packaging machine.

[0042] Example 2

[0043] (1) Soak dried seaweed in water for 3 hours to rehydrate it, then soak it in a 5% acetic acid solution for 30 minutes to remove the fishy smell, and wash it clean to obtain pretreated seaweed.

[0044] (2) Add 40g of pretreated laver to 680mL of purified water, beat with a pulper for 5min, set the dynamic high pressure micro-jet homogenizer pressure to 12000PSI for homogenization, and cycle once to obtain laver initial homogenate with laver particle size <1μm.

[0045] (3) The initial homogenate of seaweed was adjusted to pH 7, 1.6g of flavor protease with an enzyme activity of 20000U / g was added, and the seaweed was enzymatically hydrolyzed at 50℃ for 3h. The enzyme was then inactivated by placing it in boiling water for 10min to obtain seaweed enzymatic hydrolysate.

[0046] (4) Adjust the pH of the seaweed enzymatic hydrolysate to 8, mix it with 4.8g xylose and 4.8g cysteine, and perform a Maillard reaction at 100℃ for 120min to obtain seaweed Maillard solution;

[0047] (5) The Maillard solution of seaweed was cooled to room temperature and filtered with four layers of gauze. Then, the supernatant was filtered with a 0.45 μm organic filter membrane to obtain the sample solution. Ultrafiltration was performed using ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa. The solution was then spray-dried (the air volume of the spray dryer was 65%, the temperature was 190℃, and the feed rate was 35%) to obtain three components of seaweed umami peptides. The obtained peptides were packaged at room temperature using a heat shrink film packaging machine.

[0048] Example 3

[0049] (1) Soak dried wakame seaweed in water for 3 hours to rehydrate, then soak it in a 5% acetic acid solution for 30 minutes to remove the fishy smell, and wash it clean to obtain pretreated wakame seaweed.

[0050] (2) Add 40g of pretreated wakame to 720mL of purified water, and beat the wakame in a pulper for 8min. Set the dynamic high-pressure micro-jet homogenizer to 14000PSI for homogenization and cycle once to obtain an initial wakame homogenate with a particle size of <1μm.

[0051] (3) The initial homogenate of wakame seaweed was adjusted to pH 7.5, 2g of flavor protease with an enzyme activity of 20000U / g was added, and the mixture was enzymatically hydrolyzed at 55℃ for 3h. The enzyme was then inactivated by placing it in boiling water for 10min to obtain the wakame seaweed hydrolysate.

[0052] (4) Adjust the pH of the wakame enzymatic hydrolysate to 9, mix it with 5.6g xylose and 4.8g cysteine, and perform a Maillard reaction at 110℃ for 110min to obtain the wakame Maillard solution.

[0053] (5) The Maillard solution of wakame seaweed was cooled to room temperature and filtered with four layers of gauze. Then, the supernatant was filtered with a 0.45 μm organic filter membrane to obtain the sample solution. The sample solution was ultrafiltered with ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa. The sample solution was spray dried (the air volume of the spray dryer was 65%, the temperature was 190℃, and the feed rate was 35%) to obtain three components of wakame seaweed umami peptides. The obtained peptides were packaged at room temperature using a heat shrink film packaging machine.

[0054] Example 4

[0055] (1) Soak dried sea lettuce in water for 3 hours to rehydrate, then soak in acetic acid solution with a mass concentration of 5% for 30 minutes to remove the fishy smell, and wash it clean to obtain pretreated sea lettuce.

[0056] (2) 40g of sea cabbage was added to 760mL of purified water and pulped for 10min using a pulping machine. The pressure of the dynamic high-pressure micro-jet homogenizer was set to 15000PSI for homogenization and circulated once to obtain an initial homogenized solution of sea cabbage with a particle size of <1μm.

[0057] (3) The initial homogenate of sea lettuce was adjusted to pH 8, 2g of flavor protease with an enzyme activity of 20000U / g was added, and the sea lettuce was enzymatically hydrolyzed at 60℃ for 3h. The enzyme was then inactivated by placing it in boiling water for 10min to obtain sea lettuce enzymatic hydrolysate.

[0058] (4) Adjust the pH of the sea cabbage enzymatic hydrolysate to 10, mix it with 6g xylose and 5.6g cysteine, and perform a Maillard reaction at 120℃ for 100min to obtain the sea cabbage Maillard solution.

[0059] (5) The Maillard solution of sea lettuce was cooled to room temperature and filtered with four layers of gauze. Then, the supernatant was filtered with a 0.45 μm organic filter membrane to obtain the sample solution. Ultrafiltration was performed using ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa. The solution was then spray-dried (the inlet air volume of the spray dryer was 65%, the temperature was 190℃, and the feed rate was 35%) to obtain three components of sea lettuce umami peptides. The obtained peptides were packaged at room temperature using a heat shrink film packaging machine.

[0060] Example 5

[0061] (1) Soak dried agar-agar in water for 3 hours to rehydrate, then soak in acetic acid solution with a mass concentration of 5% for 30 minutes to remove the fishy smell, and wash it clean to obtain pretreated agar-agar.

[0062] (2) Add 40g of pretreated agar to 600mL of purified water, and beat with a pulper for 5min. Set the dynamic high pressure micro-jet homogenizer pressure to 10000PSI for homogenization and cycle once to obtain an initial agar homogenate with a particle size of <1μm.

[0063] (3) The initial homogenate of agar-agar was adjusted to pH 7, 1.6g of flavor protease with an enzyme activity of 20000U / g was added, and the mixture was enzymatically hydrolyzed at 50℃ for 3h. The enzyme was then inactivated by placing it in boiling water for 10min to obtain agar-agar hydrolysate.

[0064] (4) Adjust the pH of the agar-agar enzymatic hydrolysate to 7.5, mix it with 4.8g xylose and 4g cysteine, and perform a Maillard reaction at 100℃ for 100min to obtain agar-agar Maillard solution;

[0065] (5) The Maillard solution of agar-agar was cooled to room temperature and filtered with four layers of gauze. Then, the supernatant was filtered with a 0.45 μm organic filter membrane to obtain the sample solution. The sample solution was ultrafiltered with ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa. The sample solution was spray dried (the air volume of the spray dryer was 65%, the temperature was 190℃, and the feed rate was 35%) to obtain the three components of agar-agar umami peptides. The obtained peptides were packaged at room temperature using a heat shrink film packaging machine.

[0066] Comparative Example 1

[0067] Compared with Example 1, only step (2) is different. Step (2) is changed to: 40g of pretreated kelp is added to 800mL of purified water, treated with a colloid mill for 12min, and homogenized with a dynamic high pressure microjet homogenizer pressure of 26106PSI to obtain an initial kelp homogenate with a kelp particle size of <1μm.

[0068] Comparative Example 2

[0069] Compared with Example 1, only step (2) is different. Step (2) is changed to: 40g of pretreated laver is added to 680mL of purified water, treated with a colloid mill for 8min, and homogenized with a dynamic high pressure microjet homogenizer pressure of 23206PSI to obtain an initial laver homogenate with a laver particle size of <1μm.

[0070] Comparative Example 3

[0071] Compared with Example 1, only step (2) is different. Step (2) is changed to: 40g of pretreated wakame seaweed is added to 720mL of purified water, treated with a colloid mill for 9min, and homogenized with a dynamic high-pressure micro-jet homogenizer pressure of 24656PSI to obtain an initial wakame seaweed homogenate with a particle size of <1μm.

[0072] Comparative Example 4

[0073] Compared with Example 1, only step (2) is different. Step (2) is changed to: 40g of pretreated sea lettuce is added to 760mL of purified water, treated with a colloid mill for 10min, and homogenized with a dynamic high pressure micro-jet homogenizer pressure of 26106PSI to obtain an initial homogenized sea lettuce slurry with a particle size of <1μm.

[0074] Comparative Example 5

[0075] Compared with Example 1, only step (2) is different. Step (2) is changed to: 40g of pretreated agar-agar is added to 600mL of purified water, treated with a colloid mill for 5min, and homogenized with a dynamic high pressure micro-jet homogenizer pressure of 21755PSI to obtain an initial homogenized agar-agar slurry with a particle size of <1μm.

[0076] Comparative Example 6

[0077] Compared with Example 1, only step (3) is different. Step (3) is changed to: the initial homogenate of kelp is adjusted to pH 4, 1.6g of cellulase with an enzyme activity of 50000U / g is added, and the mixture is placed in a constant temperature water bath shaker at 55℃ for 3h for enzymatic hydrolysis. The mixture is then placed in boiling water for 10min to inactivate the enzyme and obtain kelp enzymatic hydrolysate.

[0078] Comparative Example 7

[0079] Compared with Example 2, only step (3) is different. Step (3) is changed to: the initial homogenized laver liquid is adjusted to pH 5, 0.8g of cellulase with an enzyme activity of 50000U / g is added, and the mixture is placed in a constant temperature water bath shaker at 50℃ for 2.5h for enzymatic hydrolysis. The mixture is then placed in boiling water for 10min to inactivate the enzyme and obtain laver enzymatic hydrolysate.

[0080] Comparative Example 8

[0081] Compared with Example 3, only step (3) is different. Step (3) is changed to: the initial homogenate of wakame seaweed is adjusted to pH 4.5, 1.2g of cellulase with an enzyme activity of 50000U / g is added, and the mixture is placed in a constant temperature water bath shaker at 55℃ for 2h for enzymatic hydrolysis. The mixture is then placed in boiling water for 10min to inactivate the enzyme and obtain the wakame seaweed enzymatic hydrolysate.

[0082] Comparative Example 9

[0083] Compared with Example 4, only step (3) is different. Step (3) is changed to: the initial homogenate of sea lettuce is adjusted to pH 4, 1.6g of cellulase with an enzyme activity of 50000U / g is added, and the mixture is placed in a constant temperature water bath shaker at 55℃ for 3.5h for enzymatic hydrolysis. The mixture is then placed in boiling water for 10min to inactivate the enzyme and obtain sea lettuce enzymatic hydrolysate.

[0084] Comparative Example 10

[0085] Compared with Example 5, only step (3) is different. Step (3) is changed to: the initial homogenate of agar-agar is adjusted to pH 5, 0.8g of cellulase with an enzyme activity of 50000U / g is added, and the mixture is placed in a constant temperature water bath shaker at 45℃ for 2h for enzymatic hydrolysis. The mixture is then placed in boiling water for 10min to inactivate the enzyme and obtain agar-agar enzymatic hydrolysate.

[0086] Comparative Example 11

[0087] Compared with Example 1, only step (2) is different. Step (2) is changed to: pre-treat 40g of kelp, crush it with a high-speed pulverizer, add 800mL of pure water, use a heat-collecting constant temperature magnetic stirrer, and cook at 50℃ for 4h to obtain the initial kelp homogenate.

[0088] Comparative Example 12

[0089] Compared with Example 2, only step (2) is different. Step (2) is changed to: pre-treat 40g of laver, crush it with a high-speed grinder, add 680mL of pure water, use a heat-collecting constant temperature magnetic stirrer, and cook at 40℃ for 3.5h to obtain the initial laver homogenate.

[0090] Comparative Example 13

[0091] Compared with Example 3, only step (2) is different. Step (2) is changed to: pre-treat 40g of wakame seaweed, crush it with a high-speed grinder, add 720mL of purified water, and cook it at 45℃ for 3h using a heat-collecting constant temperature magnetic stirrer to obtain the initial homogenized wakame seaweed slurry.

[0092] Comparative Example 14

[0093] Compared with Example 4, only step (2) is different. Step (2) is changed to: pre-treat 40g of sea lettuce, crush it with a high-speed grinder, add 760mL of pure water, use a heat-collecting constant temperature magnetic stirrer, and cook at 50℃ for 3h to obtain the initial homogenate of sea lettuce.

[0094] Comparative Example 15

[0095] Compared with Example 5, only step (2) is different. Step (2) is changed to: pre-treat 40g of agar-agar, crush it with a high-speed grinder, add 600mL of pure water, and use a heat-collecting constant temperature magnetic stirrer to cook it at 40℃ for 3h to obtain the initial homogenate of agar-agar.

[0096] Comparative Example 16

[0097] Compared with Example 1, the difference is that: 40g of pretreated kelp was crushed by a high-speed pulverizer, 1.6g of cellulase with an enzyme activity of 50000U / g was added, and the kelp was enzymatically hydrolyzed at 55℃ for 3h, and then placed in boiling water for 10min to inactivate the enzyme, thus obtaining kelp enzymatic hydrolysate.

[0098] Comparative Example 17

[0099] Compared with Example 2, the difference is that: 40g of pretreated laver was crushed in a high-speed grinder, 0.8g of cellulase with an enzyme activity of 50000U / g was added, and the laver was enzymatically hydrolyzed at 50℃ for 2.5h. The enzyme was then inactivated by placing it in boiling water for 10min to obtain laver enzymatic hydrolysate.

[0100] Comparative Example 18

[0101] Compared with Example 3, the difference is that: 40g of pretreated wakame seaweed was crushed in a high-speed grinder, 1.2g of cellulase with an enzyme activity of 50000U / g was added, and the seaweed was enzymatically hydrolyzed at 55℃ for 2h, and then placed in boiling water for 10min to inactivate the enzyme, thus obtaining wakame seaweed enzymatic hydrolysate.

[0102] Comparative Example 19

[0103] Compared with Example 4, the difference is that: 40g of pretreated sea lettuce was crushed in a high-speed grinder, 1.6g of cellulase with an enzyme activity of 50000U / g was added, and the sea lettuce was enzymatically hydrolyzed at 55℃ for 3.5h, and then placed in boiling water for 10min to inactivate the enzyme, thus obtaining sea lettuce enzymatic hydrolysate.

[0104] Comparative Example 20

[0105] Compared with Example 5, the difference is that: 40g of pretreated agar-agar was pulverized in a high-speed pulverizer, 0.8g of cellulase with an enzyme activity of 50000U / g was added, and the mixture was enzymatically hydrolyzed at 45℃ for 2h, and then placed in boiling water for 10min to inactivate the enzyme, thus obtaining agar-agar enzymatic hydrolysate.

[0106] Comparative Example 21

[0107] Compared with Example 1, the difference is that the pre-treated kelp was pulverized using a high-speed pulverizer and then pulverized using a CWF-300s ultrafine pulverizer for 0.17 hours to obtain an initial kelp homogenate.

[0108] Comparative Example 22

[0109] Compared with Example 2, the difference is that the pre-treated seaweed was pulverized using a high-speed pulverizer and then pulverized using a CWF-300s ultrafine pulverizer for 0.17 hours to obtain the initial seaweed homogenate.

[0110] Comparative Example 23

[0111] Compared with Example 3, the difference is that the pre-treated wakame seaweed was pulverized using a high-speed pulverizer and then pulverized using a CWF-300s ultrafine pulverizer for 0.17 hours to obtain an initial homogenized wakame seaweed slurry.

[0112] Comparative Example 24

[0113] Compared with Example 4, the difference is that the pretreated sea lettuce was pulverized using a high-speed pulverizer and then pulverized using a CWF-300s ultrafine pulverizer for 0.17 hours to obtain the initial homogenized sea lettuce slurry.

[0114] Comparative Example 25

[0115] Compared with Example 5, the difference is that the pretreated agar-agar was pulverized using a high-speed pulverizer and then pulverized using a CWF-300s ultrafine pulverizer for 0.17 hours to obtain an initial homogenized agar-agar slurry.

[0116] Experimental Example 1

[0117] The energy consumption ratios of umami peptides from kelp, nori, wakame, sea lettuce, and agar-agar prepared by different methods in Comparative Examples 1-5 and Comparative Examples 1-25 are shown in Table 1.

[0118] Table 1 Energy consumption ratio

[0119]

[0120] As shown in Table 1, the energy consumption of Examples 1-5 (dynamic high-pressure microjet homogenization) was the lowest at 1908±356.4 kJ / h, and its soluble nitrogen release rate was as high as 85.03±0.47%. In contrast, the energy consumption of Comparative Examples 1-5 (colloidal mill-high-pressure microjet homogenization) was 2268±486 kJ / h, which was higher than that of the dynamic high-pressure microjet treatment. Moreover, its soluble nitrogen release rate was 82.87±0.83 kJ / h, which was lower than that of the dynamic high-pressure microjet treatment. However, there was no significant difference in the material loss rate between the two. The energy consumption of Comparative Examples 6-10 (dynamic high-pressure microjet homogenization + cellulosic hydrolysis) was the highest, reaching 19732±5222.4 kJ / h. The working time was also extended by nearly 4 hours, resulting in increased energy consumption. Therefore, considering the above, the dynamic high-pressure microjet homogenization combined with the cellulosic hydrolysis process can omit the cellulosic hydrolysis step and is superior to the colloid mill-high-pressure microjet homogenization process.

[0121] In terms of working time, Examples 1-5 (dynamic high-pressure micro-jet homogenization) and Comparative Examples 21-25 (ultra-fine pulverization) are superior to Comparative Examples 1-5, 6-10, 11-15, and 16-20. Although Comparative Example 21-25 (ultra-fine pulverization) has the shortest working time, its material loss rate is the highest at 10.48±0.26%. Comparative Examples 11-15 have lower energy consumption than Comparative Examples 16-20, but their soluble nitrogen release rate is the lowest at 56.57±1.21%.

[0122] Experiment Example 2

[0123] The particle size of different preparation methods in Examples 1-5, Comparative Examples 1-5, and Comparative Examples 21-25 was investigated, and the results are shown in Table 2.

[0124] Table 2 Particle Size Statistics

[0125]

[0126] As shown in Table 2, ultrafine grinding can reduce the seaweed particle size to 10–40 μm, colloid mill-high pressure microjet homogenization treatment can reduce the seaweed particle size to 300–600 nm, and dynamic high pressure microjet homogenization treatment can reduce the seaweed particle size to 300–500 nm, thus greatly improving the seaweed cell wall breaking efficiency.

[0127] Experimental Example 3

[0128] Sensory evaluation of the seaweed umami peptides prepared in Examples 1-5 was conducted as follows:

[0129] Sensory evaluation was conducted on six aspects: color, aroma, bitterness, astringency, saltiness, umami, and seaweed-like odor. A trained panel of 10 members (5 women and 5 men, aged 20–25 years) from Sichuan Agricultural University was tested. The presented samples were evaluated using a sensory rating scale (see Table 3), with scores of 8–10 (Excellent), 6–8 (Very Good), 4–6 (Good), 2–4 (Average), and 0–2 (Poor). The final scores for each item (color, aroma, bitterness, astringency, saltiness, umami, and seaweed-like odor) were calculated as the average of the ten scores. Before conducting sensory evaluation, sensory evaluation standards were prepared: standards for bitterness, astringency, saltiness, umami, and seaweed flavor were prepared using caffeine, sodium chloride, monosodium glutamate, and seaweed extract, respectively, at concentrations of 0.08%, 0.3%, 0.5%, and 0.35%. Standards at these concentrations were rated 5 points, with scores of 0-5 indicating a flavor intensity lower than the standards, and 5-10 indicating a flavor intensity higher than the standards. Sensory personnel were trained to familiarize themselves with the rating method, the terminology for each attribute, and the sensory characteristics of seaweed umami peptides. Each sample was repeated three times by the entire group. Results are as follows: Figures 1-5 As shown.

[0130] Table 3 Sensory Standard Rating Scale

[0131]

[0132]

[0133] Therefore, this invention utilizes dynamic high-pressure microfluidic homogenization technology to break down and dissolve seaweed cells, achieving nanoscale particle sizes and making the internal proteins more easily degraded by enzymes. This significantly improves enzymatic hydrolysis efficiency, eliminating the need for the traditional first-step cellulase hydrolysis process. The umami peptides prepared through initial acetic acid soaking for deodorization and subsequent Maillard secondary deodorization have better quality, flavor, and texture. No chemical reagents or solvents are required, reducing environmental pollution. Furthermore, this technology requires relatively low energy consumption (1908±356.4 kJ / h), with a soluble nitrogen release rate as high as 85.03±0.47% and a material loss rate of 1.48±0.02%, helping to reduce production costs and meeting the requirements of sustainable development.

[0134] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for preparing seaweed umami peptides by dynamic high-pressure microfluidic homogenization-assisted enzymatic hydrolysis, characterized in that, Includes the following steps: (1) Soak dried seaweed to remove the fishy smell, wash it clean with water to obtain pre-treated seaweed; (2) The pretreated seaweed obtained in step (1) is pulped, homogenized, and the initial seaweed homogenate is obtained. (3) The initial seaweed homogenate obtained in step (2) was adjusted to pH 7-8 and subjected to enzymatic hydrolysis to obtain seaweed hydrolysate; (4) The seaweed hydrolysate obtained in step (3) is adjusted to pH 7-11, mixed with xylose and cysteine, and subjected to Maillard reaction to obtain seaweed Maillard solution; (5) The seaweed Maillard solution obtained in step (4) was filtered to obtain a sample solution. It was then ultrafiltered using ultrafiltration membranes with molecular weights >3kDa, 3k~1kDa and <1kDa, and spray-dried to obtain seaweed umami peptides. The homogenization in step (2) is dynamic high-pressure microjets homogenization; the pressure of the homogenization is 10000~15000 PSI, and the cycle is repeated once; The enzyme used in step (3) for enzymatic hydrolysis is flavor protease.

2. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The dried seaweed mentioned in step (1) is one or more of dried kelp, dried laver, dried wakame, dried sea lettuce, or dried agaric.

3. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The soaking in step (1) is carried out in water for 2-4 hours; the deodorization in step (1) is carried out by soaking in an acetic acid solution with a mass concentration of 4%-6% for 20-40 minutes.

4. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The pulping process described in step (2) involves adding water at 15 to 20 times the weight of the pretreated seaweed and pulping for 5 to 10 minutes.

5. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The particle size of the seaweed in the initial homogenate of seaweed in step (2) is <1μm.

6. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The amount of flavor protease added in step (3) is 4wt%~6wt% of the pretreated seaweed mass, the enzymatic hydrolysis time is 3~4h, and the enzymatic hydrolysis temperature is 50~60℃.

7. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The amount of xylose used in step (4) is 12wt%~16wt% of the pretreated seaweed mass; the amount of cysteine ​​used in step (4) is 10wt%~14wt% of the pretreated seaweed mass; the temperature of the Maillard reaction in step (4) is 100~120℃, and the time of the Maillard reaction is 100~150min.

8. The method for preparing seaweed umami peptides according to claim 1, characterized in that, The filtration in step (5) includes filtration using four layers of gauze and filtration using a 0.45μm organic filter membrane.

9. The method for preparing seaweed umami peptides according to claim 1, characterized in that, In step (5), the air intake volume of the spray drying is 65%, the temperature of the spray drying is 190°C, and the feed rate of the spray drying is 35%.

10. The seaweed umami peptide prepared by the method for preparing seaweed umami peptides according to any one of claims 1 to 9.

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