Velvet mushroom strong-flavor peptide as well as preparation and application thereof

Abstract

The invention relates to the technical field of protein active peptides, in particular to a pilose antler mushroom strong-flavor peptide as well as preparation and application thereof. The amino acid sequence of the velvet antler mushroom strong taste peptide provided by the invention is selected from one of the sequences shown as SEQ ID NO.1 and SEQ ID NO.2. The pilose antler mushroom strong-flavor peptide provided by the invention is obtained by separating and purifying pilose antler mushroom, and sensory detection results show that the two pilose antler mushroom strong-flavor peptides have good strong flavor and salty and delicious taste enhancing effects; the pilose antler mushroom strong taste peptide provided by the invention is a small molecule peptide and is easy to absorb by a human body; the extraction method of the velvet antler mushroom strong-flavor peptide is simple to operate and free of any pollution, and the prepared velvet antler mushroom strong-flavor peptide is outstanding in strong flavor and is a high-quality flavor base material.

Description

Technical Field

[0001] The invention relates to the technical field of protein active peptides, in particular to a velvet antler mushroom strong flavor peptide and the preparation and application thereof. Background Art

[0002] Sweet, sour, bitter, salty and umami are the five traditional basic tastes. In the 1990s, Japanese scholars found that water extracts of garlic and onions can significantly increase the richness and continuity of soup bases. They then separated and identified the key components as γ-glutamyl peptides, and called such substances that can cause taste impact, continuity and complexity as Kokumi substances. Kokumi is a compound word of "Koku" plus "mi" ("mi" means taste in Japanese). Koku consists of three elements: complexity, fullness and continuity. It is an overall feeling that includes taste, aroma and texture.

[0003] Kokumi is the sixth flavor after the traditional five basic flavors. Kokumi is different from the other flavors. Kokumi itself has no taste, or only a very light taste, but adding it to other matrices (such as chicken soup) can significantly enhance the intensity of other flavors (such as salty, fresh). For example, beef, mushrooms, onions, cheese and other ingredients are rich in rich flavor ingredients, which can make the taste fuller, chewier and more mouthfeeling.

[0004] With the continuous deepening of Kokumi research, more and more Koku substances have been discovered, most of which are separated and purified from natural food raw materials. At present, Koku substances are mainly divided into two categories: peptides and non-peptides. Peptide Kokumi substances mainly include amino acids and their derivatives, oligopeptides, and proteins. The research on non-peptide Kokumi substances is relatively small, and the current research hotspots are mainly focused on oligopeptide Kokumi substances.

[0005] As a rare edible and medicinal mushroom, Lyophyllum decastes is rich in nutrition, high in protein and low in fat. It has the effects of anti-tumor, reducing three highs, regulating immunity, protecting the liver, etc. There are few reports on the research of flavor peptides. However, actual measurements show that dried Lyophyllum decastes contains more protein than other varieties. Therefore, the exploration of the rich flavor peptides of Lyophyllum decastes is still needed by technicians in this field. Summary of the invention

[0006] In order to solve the above problems, the purpose of the present invention is to provide a kind of thick flavor peptide of velvet antler mushroom and its preparation and application. The amino acid sequence of the thick flavor peptide of velvet antler mushroom provided by the present invention is selected as one of the sequences shown in SEQ ID NO.1 and SEQ ID NO.2. The thick flavor peptides of velvet antler mushroom provided by the present invention are all obtained by separation and purification from velvet antler mushroom. Using sensory detection, the results show that the two thick flavor peptides of velvet antler mushroom have good thick flavor and salty and fresh taste enhancement effects; the thick flavor peptides of velvet antler mushroom provided by the present invention are all small molecule peptides, which are easily absorbed by the human body; the extraction method of the thick flavor peptide of velvet antler mushroom provided by the present invention is simple to operate, without any pollution, and the obtained edible fungus thick flavor peptide has a prominent thick flavor, which is a high-quality flavor base.

[0007] The purpose of the present invention can be achieved by the following technical solutions:

[0008] The first object of the present invention is to provide a velvet antler mushroom thick flavor peptide, the amino acid sequence of which is selected from one of the sequences shown in SEQ ID NO.1 and SEQ ID NO.2.

[0009] The second object of the present invention is to provide a method for preparing a thick flavor peptide of velvet antler mushroom, wherein the preparation method is selected from one of the following methods:

[0010] (1) obtained by separation and extraction of Pleurotus eryngii;

[0011] (2) Artificial synthesis.

[0012] In one embodiment of the present invention, the specific steps of separating and extracting Pleurotus eryngii are as follows:

[0013] (S1) dissolving the dry antler powder and mixing it to obtain an enzymatic hydrolysis raw material solution;

[0014] (S2) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S1) to perform a first enzymatic hydrolysis treatment; then adding flavor protease to perform a second enzymatic hydrolysis treatment to obtain an enzymatic hydrolysis solution;

[0015] (S3) The enzymatic hydrolysate prepared in step (S2) is centrifuged, and the supernatant is post-treated to prepare Pleurotus eryngii thick flavor peptide.

[0016] In one embodiment of the present invention, in step (S1), the particle size of the dry velvet antler powder is 0.076-2.54 mm, the dry velvet antler powder is dissolved in water, and the mass ratio of the dry velvet antler powder to water is 1:10-100.

[0017] In one embodiment of the present invention, in step (S2), the mass ratio of the flavor protease to the pectinase is 2:1 to 1:2;

[0018] The mass ratio of the sum of the added amounts of flavor protease and pectinase to the enzymatic hydrolysis raw material liquid is 1:100-1000.

[0019] In one embodiment of the present invention, in step (S2), during the first enzymatic hydrolysis process, the pH is 2-3, the temperature is 40-60° C., and the time is 1-5 hours.

[0020] In one embodiment of the present invention, in step (S2), during the second enzymatic hydrolysis process, the pH is 5 to 7, the temperature is 40 to 60° C., and the time is 1 to 5 hours.

[0021] In one embodiment of the present invention, in step (S3), the rotation speed is 5000 to 11000 rpm / min, and the time is 10 to 20 min;

[0022] The post-treatment is to carry out concentration and drying treatment in sequence.

[0023] In one embodiment of the present invention, the supernatant is concentrated by passing through an ultrafiltration membrane;

[0024] The drying is selected from freeze drying, vacuum drying or spray drying.

[0025] The third object of the present invention is to provide an application of a velvet antler mushroom rich flavor peptide in the preparation of a food seasoning, wherein the food seasoning is a food additive for increasing the rich flavor of food.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] (1) The kokumi peptides provided by the present invention are obtained by separation and purification from kokumi mushroom. The results of sensory testing show that the two kokumi peptides have good kokumi and salty and fresh taste enhancing effects.

[0028] (2) The velvet antler mushroom rich flavor peptides provided by the present invention are all small molecule peptides and are easily absorbed by the human body.

[0029] (3) The extraction method of the velvet antler mushroom rich flavor peptide provided by the present invention is simple to operate, has no pollution, and the obtained edible fungus rich flavor peptide has a prominent rich flavor and is a high-quality flavor base material. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is the chromatogram of membrane separation components with molecular weight less than 1000Da after Sephadex G-15 gel chromatography separation;

[0031] Figure 2 This is the sensory radar chart obtained by sensory evaluation and analysis of P1, P2, and P3 after ultrafiltration;

[0032] Figure 3This is the sensory radar chart obtained after sensory evaluation and analysis of F1, F2, F3, F4, and F5;

[0033] Figure 4 is the total ion current diagram of liquid chromatography-mass spectrometry (LC-MS / MS) of the F3 component;

[0034] Figure 5 This is the secondary mass spectrum of the thick flavor peptide LDRLE from velvet antler mushroom;

[0035] Figure 6 This is the secondary mass spectrum of the thick flavor peptide FDLHR from Pilosa velutipes;

[0036] Figure 7 The sensory radar chart obtained by sensory evaluation analysis of the product prepared in Comparative Example 1;

[0037] Figure 8 This is a sensory radar chart obtained by performing sensory evaluation analysis on the product prepared in Comparative Example 2;

[0038] Fig. 9 This is a sensory radar chart obtained by performing sensory evaluation analysis on the product prepared in Comparative Example 3;

[0039] Fig.10 This is a sensory radar chart obtained by performing sensory evaluation analysis on the product prepared in Comparative Example 4. DETAILED DESCRIPTION

[0040] In the specification of the present invention and the following examples, unless otherwise specified, the velvet antler mushroom kochi peptide refers to the velvet antler mushroom kochi peptide obtained by, for example, but not limited to, the extraction method described herein. Those skilled in the art will appreciate that the velvet antler mushroom kochi peptide of the present invention can be obtained by conventional methods or the extraction method described in the specification, or can be obtained by artificial synthesis according to the amino acid sequence provided in the specification.

[0041] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0042] The following examples use conventional instruments and equipment in the art. The experimental methods in the following examples that do not specify specific conditions are usually carried out under conventional conditions or under conditions recommended by the manufacturer. The various raw materials used in the following examples are conventional commercial products unless otherwise specified, and their specifications are conventional specifications in the art. In the specification of the present invention and the following examples, unless otherwise specified, "%" means weight percentage, and ratio means weight ratio; "GSH" means "glutathione"; pectinase is purchased from Bioss (item number: 04837344); flavor protease is purchased from Bioss (item number is 04835107).

[0043] Example 1

[0044] This embodiment provides a method for preparing a velvet mushroom thick flavor peptide, which specifically comprises the following steps:

[0045] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0046] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0047] (S3) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 2.5 using a 10 mol / L hydrochloric acid solution, and then performing a first enzymatic hydrolysis treatment (50° C., 2 h). After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to fully inactivate the enzymatic activity of the pectinase;

[0048] (S4) After step (S3), flavor protease is added, and the pH of the enzymatic hydrolysis raw material solution is adjusted to 6 using a 10 mol / L sodium hydroxide solution, and then a second enzymatic hydrolysis treatment (50° C., 3 h) is performed. After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to obtain an enzymatic hydrolysis solution;

[0049] The amount of pectinase added is 0.3% of the amount of dry velvet antler mushroom powder;

[0050] The amount of flavor protease added is 0.2% of the amount of dry velvet antler mushroom powder;

[0051] (S5) The enzymatic hydrolysate prepared in step (S4) is centrifuged at 8000 rpm / min for 10 min to collect the supernatant; the supernatant is then centrifuged through an ultrafiltration membrane (25°C, ultrafiltration separation is performed at 25°C using ultrafiltration membranes with molecular weight cutoffs of 1000 Da and 3000 Da, respectively), and components with molecular weights of <1000 Da (denoted as "P1"), 1000-3000 Da (denoted as "P2"), and >3000 Da (denoted as "P3") are collected and freeze-dried, and then the components are redissolved in ultrapure water, purified by a gel chromatography column, and eluted with ultrapure water (elution conditions are: sample volume 20 mL, flow rate 1.2 mL / min, ultraviolet detector sensitivity 1.0, detection wavelength 220 nm) to obtain chromatography components, which are freeze-dried to obtain a product (and stored at -20°C for future use).

[0052] When sample P1 was eluted, the collection tube was updated every 10 minutes, and five chromatographic components were obtained (represented by F1, F2, F3, F4 and F5, respectively, and the chromatogram is shown in FIG. Figure 1 shown).

[0053] Performance Analysis:

[0054] 1) Analyze the taste characteristics of ultrafiltration fractions P1-P3 and chromatography fractions F1-F5 using artificial sensory evaluation method

[0055] The sensory evaluation method uses quantitative analysis (QDA) to determine the taste intensity of each component obtained by membrane separation and gel separation. The specific operation is as follows: Take the freeze-dried component samples of P1-P3 and F1-F5 respectively to prepare a 5mg / mL solution, and use the scoring method to score the sour, sweet, bitter, salty, fresh and strong taste, with a full score of 5 points. There are 8 sensory evaluators (4 men and 4 women, aged 25 to 30 years old); before the sensory analysis, 1% sucrose solution is used as a sweetness standard, 0.35% sodium amide solution is used as a saltiness standard, 0.08% quinine solution is used as a bitterness standard, 0.35% sodium glutamate solution is used as a savory standard, 0.08% citric acid solution is used as a sourness standard, and chicken soup solution with 5mmol / L GSH added is used as a strong taste standard. Train the members of the sensory analysis team. Sip from 9:00am to 12:00am every day for five days to train their taste senses. The test was conducted at room temperature, with each sample tested three times at different times, and the final result was the average of the three tests.

[0056] The taste intensity of P1-P3 and F1-F5 is shown in Figure 2 and Figure 3 .

[0057] from Figure 2 and Figure 3 It can be seen that the membrane separation fraction P1 and the chromatography fraction F3 have the strongest kokumi taste.

[0058] The results show that there is a certain correlation between the molecular weight of peptides and the Kokumi taste. For example, Maillard reaction peptides of 300 to 3000 Da have a strong Kokumi effect, and most of the γ-glutamyl peptides with Kokumi effect that have been studied more are dipeptides or tripeptides with a molecular weight of less than 500 Da. In recent years, it has also been found that the molecular weight of polypeptides with Kokumi effect is concentrated below 1000 Da.

[0059] 2) Identification of the peptide sequence structure of component F3

[0060] Sample preparation: Dissolve the F3 component sample in pure water, mix it on a vortex mixer to fully dissolve it, then centrifuge it and take the supernatant for later use.

[0061] The LC-MS / MS assay conditions are as follows:

[0062] Liquid phase conditions: packing: Reprosil-Pur 120C18-AQ 3μm analytical column (100μm id×180mm); injection volume 20μL; flow rate 600nL / min; two eluents were used as mobile phases for gradient elution, eluent A was 0.1% formic acid aqueous solution, and eluent B was 0.1% formic acid aqueous solution and 80% acetonitrile aqueous solution; elution conditions were as follows: 0-3min: 4% B; 3-73min: 15% B; 73-93min: 25% B; 93-109min: 40% B; 109-110min: 95% B; 110-120min: 95% B.

[0063] Mass spectrometry conditions: The full scan range of the mass spectrometry was 100-1500 m / z, the primary mass spectrometry resolution was set to 70,000, AGC was 3e6, and MaximumIT was 100 ms; the parent ion with an ion intensity of 20 in the full scan was selected and fragmented using the high-energy collision fragmentation (HCD) method for secondary mass spectrometry detection. The secondary mass spectrometry resolution was set to Resolution: 17,500, AGC was 1e5, MaximumIT was 50 ms, and the peptide fragmentation collision energy was set to 28 to generate the raw data (.raw) of mass spectrometry detection. The total ion current is shown in the figure. Figure 4 shown.

[0064] After LC-MS / MS separated and identified component F3, the library was searched according to the identification results, and the amino acid sequence of the F3 separated component was identified by secondary mass spectrometry. After analysis, the final peptide sequences were LDRLE (peptide 1: SEQ ID NO.1) and FDLHR (peptide 2: SEQ ID NO.2). The results are shown in Figure 5 , Figure 6 .

[0065] In summary, two kokumi peptides of fraction F3 separated by gel chromatography were obtained, and their polypeptide sequences were LDRLE and FDLHR.

[0066] The velvet antler mushroom thick flavor peptide in this embodiment has an obvious flavor enhancement effect, can be used in the food field, and can be added to seasonings as a base material and an auxiliary material to enrich the taste of food.

[0067] Example 2

[0068] This embodiment provides a method for preparing a velvet mushroom thick flavor peptide, which specifically comprises the following steps:

[0069] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0070] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0071] (S3) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 3 using a 10 mol / L hydrochloric acid solution, and then performing a first enzymatic hydrolysis treatment (60° C., 1 h), and performing an enzyme inactivation treatment (90° C. water bath for 5 min) after the enzymatic hydrolysis is completed to fully lose the enzymatic activity of the pectinase;

[0072] (S4) After step (S3), flavor protease is added, and the pH of the enzymatic hydrolysis raw material solution is adjusted to 5 using a 10 mol / L sodium hydroxide solution, and then a second enzymatic hydrolysis treatment (40° C., 5 h) is performed. After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to obtain an enzymatic hydrolysis solution;

[0073] The amount of pectinase added is 0.3% of the amount of dry velvet antler mushroom powder;

[0074] The amount of flavor protease added is 0.15% of the amount of dry velvet antler mushroom powder;

[0075] (S5) The enzymatic hydrolyzate prepared in step (S4) is centrifuged at 5000 rpm / min for 20 min to collect the supernatant; the supernatant is then centrifuged through an ultrafiltration membrane (25°C, ultrafiltration separation is performed at 25°C using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da), and components with a molecular weight of less than 1000 Da are collected and freeze-dried to obtain pilose antler mushroom thick flavor peptide.

[0076] Example 3

[0077] This embodiment provides a method for preparing a velvet mushroom thick flavor peptide, which specifically comprises the following steps:

[0078] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0079] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0080] (S3) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 2 using a 10 mol / L hydrochloric acid solution, and then performing a first enzymatic hydrolysis treatment (60° C., 1 h), and performing an enzyme inactivation treatment (90° C. water bath for 5 min) after the enzymatic hydrolysis is completed to fully inactivate the enzymatic activity of the pectinase;

[0081] (S4) After step (S3), flavor protease is added, and the pH of the enzymatic hydrolysis raw material solution is adjusted to 7 using a 10 mol / L sodium hydroxide solution, and then a second enzymatic hydrolysis treatment (40° C., 5 h) is performed. After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to obtain an enzymatic hydrolysis solution;

[0082] The amount of pectinase added is 0.1% of the amount of dry velvet antler mushroom powder;

[0083] The amount of flavor protease added is 0.2% of the amount of dry velvet antler mushroom powder;

[0084] (S5) The enzymatic hydrolyzate prepared in step (S4) is centrifuged at 11000 rpm / min for 10 min to collect the supernatant; the supernatant is then centrifuged through an ultrafiltration membrane (25°C, ultrafiltration separation is performed at 25°C using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da), and components with a molecular weight of less than 1000 Da are collected and freeze-dried to obtain pilose antler mushroom thick flavor peptide.

[0085] The performance of the velvet antler mushroom thick flavor peptides prepared in Examples 2 and 3 is comparable to that of the velvet antler mushroom thick flavor peptides prepared in Example 1.

[0086] Comparative Example 1 (pectinase treatment only)

[0087] This comparative example provides a method for preparing a thick flavor peptide of velvet antler mushroom, which specifically comprises the following steps:

[0088] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0089] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0090] (S3) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 2.5 using a 10 mol / L hydrochloric acid solution, and then performing a first enzymatic hydrolysis treatment (50° C., 2 h). After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to fully lose the enzymatic activity of the pectinase, thereby obtaining an enzymatic hydrolysis solution;

[0091] The amount of pectinase added is 0.3% of the amount of dry velvet antler mushroom powder;

[0092] (S4) the enzymatic hydrolysate prepared in step (S3) was centrifuged at 8000 rpm / min for 10 min, and the supernatant was collected; the supernatant was then centrifuged through an ultrafiltration membrane (25° C., ultrafiltration separation was performed at 25° C. using ultrafiltration membranes with molecular weight cutoffs of 1000 Da and 3000 Da, respectively), and components with molecular weights of <1000 Da, 1000-3000 Da, and >3000 Da were collected for freeze-drying (the sensory radar diagram is as shown in FIG. 1 ). Figure 7 shown).

[0093] pass Figure 7 It can be found that only using pectinase enzymatic hydrolysis cannot enzymatic hydrolyze the velvet antler mushroom into small molecule peptides, so Kokumi peptide cannot be produced, but its sensory results still have salty, fresh and sweet tastes, which may be due to the free amino acids in the velvet antler mushroom.

[0094] Comparative Example 2 (flavor protease treatment only)

[0095] This comparative example provides a method for preparing a thick flavor peptide of velvet antler mushroom, which specifically comprises the following steps:

[0096] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0097] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0098] (S3) adding flavor protease to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 6 using a 10 mol / L sodium hydroxide solution, and then performing a first enzymatic hydrolysis treatment (50° C., 3 h), and after the enzymatic hydrolysis is completed, performing an enzyme inactivation treatment (90° C. water bath for 5 min) to fully inactivate the enzymatic activity of the pectinase, thereby obtaining an enzymatic hydrolysis solution;

[0099] The amount of flavor protease added is 0.2% of the amount of dry velvet antler mushroom powder;

[0100] (S4) The enzymatic hydrolysate prepared in step (S3) was centrifuged at 8000 rpm / min for 10 min, and the supernatant was collected; the supernatant was then centrifuged through an ultrafiltration membrane (25° C., ultrafiltration separation was performed using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da at 25° C.), and components with a molecular weight of less than 1000 Da were collected for freeze-drying (the sensory radar diagram of which is as shown in FIG. 1 ). Figure 8 shown).

[0101] pass Figure 8 It can be found that the velvet antler mushroom protein cannot be melted out by only using flavor protease treatment, the protease cannot react with the velvet antler mushroom protein, and the Kokumi peptide cannot be prepared.

[0102] Comparative Example 3 (excessive addition of pectinase)

[0103] This comparative example provides a method for preparing a thick flavor peptide of velvet antler mushroom, which specifically comprises the following steps:

[0104] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0105] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0106] (S3) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 2.5 using a 10 mol / L sodium hydroxide solution, and then performing a first enzymatic hydrolysis treatment (50° C., 2 h). After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to fully inactivate the enzymatic activity of the pectinase;

[0107] (S4) After step (S3), flavor protease is added, and the pH of the enzymatic hydrolysis raw material solution is adjusted to 6 using a 10 mol / L sodium hydroxide solution, and then a second enzymatic hydrolysis treatment (50° C., 3 h) is performed. After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to obtain an enzymatic hydrolysis solution;

[0108] The amount of pectinase added is 0.5% of the amount of dry velvet antler mushroom powder;

[0109] The amount of flavor protease added is 0.2% of the amount of dry velvet antler mushroom powder;

[0110] (S5) The enzymatic hydrolysate prepared in step (S4) was centrifuged at 8000 rpm / min for 10 min, and the supernatant was collected; the supernatant was then centrifuged through an ultrafiltration membrane (25° C., ultrafiltration separation was performed at 25° C. using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da), and the components with a molecular weight of less than 1000 Da were collected for freeze-drying (the sensory radar diagram of which is as shown in FIG. 1 ). Fig. 9 shown).

[0111] pass Fig. 9 It can be found that when pectinase is added in excess, it may cause the concentration of other components in the solution (such as polysaccharides, nucleic acids, etc.) to increase, which may subsequently affect the enzymatic hydrolysis of flavor proteases. At the same time, excess pectinase may also interact with proteins.

[0112] Comparative Example 4 (too much flavor protease added)

[0113] This comparative example provides a method for preparing a thick flavor peptide of velvet antler mushroom, which specifically comprises the following steps:

[0114] (S1) using a grinder to grind commercially available dried velvet antler mushroom, and passing the mixture through a 200-mesh sieve to obtain dried velvet antler mushroom powder;

[0115] (S2) mixing the dry pilose antler powder prepared in step (S1) with water (the material-liquid ratio is 1:10) to obtain an enzymatic hydrolysis raw material liquid;

[0116] (S3) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S2), and adjusting the pH of the enzymatic hydrolysis raw material solution to 2.5 using a 10 mol / L sodium hydroxide solution, and then performing a first enzymatic hydrolysis treatment (50° C., 2 h). After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to fully inactivate the enzymatic activity of the pectinase;

[0117] (S4) After step (S3), flavor protease is added, and the pH of the enzymatic hydrolysis raw material solution is adjusted to 6 using a 10 mol / L sodium hydroxide solution, and then a second enzymatic hydrolysis treatment (50° C., 3 h) is performed. After the enzymatic hydrolysis is completed, an enzyme inactivation treatment (90° C. water bath for 5 min) is performed to obtain an enzymatic hydrolysis solution;

[0118] The amount of pectinase added is 0.3% of the amount of dry velvet antler mushroom powder;

[0119] The amount of flavor protease added is 0.8% of the amount of dry velvet antler mushroom powder;

[0120] (S5) The enzymatic hydrolysate prepared in step (S4) was centrifuged at 8000 rpm / min for 10 min, and the supernatant was collected; the supernatant was then centrifuged through an ultrafiltration membrane (25° C., ultrafiltration separation was performed at 25° C. using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da), and the components with a molecular weight of less than 1000 Da were collected for freeze-drying (the sensory radar diagram of which is as shown in FIG. 1 ). Fig.10 shown).

[0121] pass Fig.10It can be found that when an excessive amount of flavor protease is added, the peptides may be overly hydrolyzed, resulting in the re-fragmentation of the produced peptides, thereby reducing the types of target peptides and the content of target peptides.

[0122] Through Examples 1 to 3 and Comparative Examples 1 to 4, it can be found that in the preparation method of a pilose antler mushroom thick flavor peptide of the present invention, the synergistic cooperation of pectinase and flavor protease (including dosage and processing conditions) is required to obtain the desired pilose antler mushroom thick flavor peptide product.

[0123] The above description of the embodiments is to facilitate the understanding and use of the invention by those skilled in the art. It is obvious that those skilled in the art can easily make various modifications to these embodiments and apply the general principles described herein to other embodiments without creative work. Therefore, the present invention is not limited to the above embodiments, and improvements and modifications made by those skilled in the art based on the explanation of the present invention without departing from the scope of the present invention should be within the scope of protection of the present invention.

Claims

1. A kind of thick flavor peptide of velvet antler mushroom, It is characterized in that The amino acid sequence is selected from one of the sequences shown in SEQ ID NO.1 and SEQ ID NO.

2.

2. A method for preparing the velvet antler mushroom rich flavor peptide as claimed in claim 1, It is characterized in that The preparation method is selected from one of the following methods: (1) obtained by separation and extraction of Pleurotus eryngii; (2) Artificial synthesis.

3. The method for preparing the thick flavor peptide of Pleurotus eryngii according to claim 2, It is characterized in that The specific steps of separation and extraction of Pterygium amara are as follows: (S1) dissolving the dry antler powder and mixing it to obtain an enzymatic hydrolysis raw material solution; (S2) adding pectinase to the enzymatic hydrolysis raw material solution prepared in step (S1) to perform a first enzymatic hydrolysis treatment; then adding flavor protease to perform a second enzymatic hydrolysis treatment to obtain an enzymatic hydrolysis solution; (S3) The enzymatic hydrolysate prepared in step (S2) is centrifuged, and the supernatant is post-treated to prepare Pleurotus eryngii thick flavor peptide.

4. The method for preparing the thick flavor peptide of Pleurotus velutipes according to claim 3, It is characterized in that In step (S1), the particle size of the dry velvet antler powder is 0.076-2.54 mm, the dry velvet antler powder is dissolved in water, and the mass ratio of the dry velvet antler powder to water is 1:10-100.

5. The method for preparing the thick flavor peptide of Pleurotus eryngii according to claim 3, It is characterized in that In step (S2), the mass ratio of the flavor protease to the pectinase is 2:1 to 1:2; The mass ratio of the sum of the added amounts of flavor protease and pectinase to the enzymatic hydrolysis raw material liquid is 1:100-1000.

6. The method for preparing the thick flavor peptide of Pleurotus velutipes according to claim 3, It is characterized in that In step (S2), during the first enzymatic hydrolysis process, the pH is 2-3, the temperature is 40-60° C., and the time is 1-5 hours.

7. The method for preparing the thick flavor peptide of Pleurotus eryngii according to claim 3, It is characterized in that In step (S2), during the second enzymatic hydrolysis process, the pH is 5 to 7, the temperature is 40 to 60° C., and the time is 1 to 5 hours.

8. The method for preparing the thick flavor peptide of Pleurotus eryngii according to claim 3, It is characterized in that In step (S3), during the centrifugal treatment, the rotation speed is 5000-11000 rpm / min, and the time is 10-20 min; The post-treatment is sequentially performing concentration and drying.

9. The method for preparing the thick flavor peptide of Pleurotus velutipes according to claim 8, It is characterized in that The supernatant is concentrated by passing through an ultrafiltration membrane; The drying is selected from freeze drying, vacuum drying or spray drying.

10. Use of the velvet antler mushroom thick flavor peptide as claimed in claim 1 in preparing food seasoning.

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