CN116333045A - Preserved meat flavor peptide and preparation method and application thereof - Google Patents
Preserved meat flavor peptide and preparation method and application thereof Download PDFInfo
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Images
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L27/00—Spices; Flavouring agents or condiments; Artificial sweetening agents; Table salts; Dietetic salt substitutes; Preparation or treatment thereof
- A23L27/10—Natural spices, flavouring agents or condiments; Extracts thereof
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/18—Peptides; Protein hydrolysates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Nutrition Science (AREA)
- Engineering & Computer Science (AREA)
- Polymers & Plastics (AREA)
- Food Science & Technology (AREA)
- Organic Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Medicinal Chemistry (AREA)
- Mycology (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- Genetics & Genomics (AREA)
- General Health & Medical Sciences (AREA)
- Seasonings (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention discloses a bacon flavor peptide, the amino acid sequence of which is shown in SEQ ID NO: 1. The preparation method comprises the following steps: firstly extracting water-soluble components from preserved meat, separating ultrafiltration components with a cut-off molecular weight of less than 3000Da by ultrafiltration, and then further separating and purifying the ultrafiltration components by an RP-HPLC method to obtain the flavor peptide of the preserved meat. The flavor peptide prepared by the method has the function of enhancing the delicate flavor, the fresh flavor and the thick flavor of food, can be applied to food or seasonings, and enriches the mouthfeel of the food and the seasonings. The two polypeptides provided by the invention have higher ACE inhibition rate, and can be applied to the preparation of blood pressure-reducing health-care food.
Description
Technical Field
The invention belongs to the technical field of biology, and particularly relates to a bacon flavor peptide, a preparation method and application thereof.
Background
The peptide is a small molecular bioactive substance, is a product of protein after hydrolysis, and is named as flavor peptide. The taste peptide is a kind of small molecular peptide capable of producing different tastes of sour, sweet, bitter, salty, fresh and the like, and is usually prepared by means of alcoholysis, microbial fermentation or chemical synthesis. The taste characteristics are often related to amino acid composition and amino acid sequence, and different taste peptides can generate different taste sensations and taste intensities; when being used together with the flavoring agent and the sour agent, the flavoring agent can improve the original flavor, enhance the flavor and the thick flavor of food, improve the flavor intensity of the food and increase the nutrition of the food. Besides the flavor development effect, the flavor development peptide is taken as a small molecular peptide, the absorption and utilization efficiency of human body is higher than that of macromolecular protein and free amino acid, the nutrition characteristic is excellent, the problem that the flavor formed by a single flavoring base material is not thick enough is solved, and the food quality is improved, so that the flavor development peptide is taken as a flavor development base material or auxiliary material to be applied to the food field, and has a great market prospect.
The traditional preserved meat is mainly popular in Sichuan, hunan, guangdong and the like, has the unique smoking flavor which is deeply favored by markets in the national range, has the praise of 'one-pot meat for hundreds of feet', has mellow and delicious taste, is fat but not greasy, has unique flavor, and is rich in nutrient substances such as fat, protein, amino acid and the like. However, the research on cured meat at the present stage mainly focuses on the technology and the problem of nitrite exceeding reduction, and is rarely reported on flavor peptide separated from the cured meat.
Disclosure of Invention
The invention aims to provide a bacon flavor peptide, a preparation method and application thereof, wherein the bacon flavor peptide can be added into food or seasonings, so that the mouthfeel of the food and the seasonings is enriched, and the bacon flavor peptide is aromatic and delicious.
In order to achieve the above purpose, the first technical scheme adopted by the invention is as follows:
a bacon flavor peptide, the amino acid sequence of which is shown in SEQ ID NO: 1.
Preferably, the bacon taste peptide is extracted from bacon.
The second technical scheme adopted by the invention is as follows:
the preparation method of the bacon flavor peptide comprises the following steps:
peeling preserved meat, superfine pulverizing, and extracting crude peptide powder by adopting a phosphate method;
separating ultrafiltration components with molecular weight cutoff less than 3000Da from the crude peptide powder by ultrafiltration;
carrying out chromatography on the ultrafiltration component by adopting a gel chromatography method; and
and separating and purifying the components obtained after chromatography.
The third technical scheme adopted by the invention is as follows:
the application of bacon taste peptide in food.
The fourth technical scheme adopted by the invention is as follows:
a food seasoning characterized by comprising the bacon-flavor peptide of the first aspect.
The taste peptide can also be synthesized by chemical methods using amino acids as raw materials, as is well known in the art.
Meanwhile, the taste peptide has higher ACE inhibitory activity and has a certain blood pressure reducing function.
Therefore, the bacon taste peptide can be used for preparing blood pressure-reducing health-care food.
The preserved meat flavor peptide provided by the invention has a low flavor threshold value, is lower than a flavor standard threshold value, has a good flavor effect, and can obviously improve the salty taste, flavor and concentration of food and coordinate the mouthfeel.
Drawings
FIG. 1 is a chromatographic chart of an ultrafiltration component with a molecular weight below 3000Da after Sephadex G-15 gel chromatographic separation;
FIG. 2 shows the results of taste dilution analysis of ultrafiltration fraction P and chromatographic fractions P1, P2, P3;
FIG. 3 is an ultra performance liquid chromatography separation profile of chromatographic fraction P2;
FIG. 4 is a secondary mass spectrum of the sequence EVHEEVH;
fig. 5 shows ACE inhibitory activity before and after digestion of the taste peptide.
Detailed Description
The present invention will be further described in detail with reference to the following detailed description and the accompanying drawings, in order to make the objects, technical solutions and advantages of the present invention more apparent. It should be understood that the description is only illustrative and is not intended to limit the scope of the invention. In addition, in the following description, descriptions of well-known structures and techniques are omitted so as not to unnecessarily obscure the present invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
The first embodiment of the invention provides a bacon flavor peptide, the amino acid sequence of which is shown in SEQ ID NO: 1.
The taste peptide separated from the preserved meat is extracted from the preserved meat, in particular from the traditional Sichuan style preserved meat.
The second embodiment of the invention provides a preparation method of flavor peptide separated from preserved meat, comprising the following steps:
step 1: peeling the preserved meat and then carrying out superfine grinding;
step 2: adding phosphate buffer solution into the crushed preserved meat, homogenizing for 10 min in ice bath state 8000 r/min, and centrifuging; adding 3 times volume of absolute ethyl alcohol into the supernatant, and standing for 12 hours at 4 ℃; centrifuging, collecting supernatant, suction filtering, rotary evaporating, lyophilizing to obtain crude peptide powder of conventional Chuan style bacon, and storing at-20deg.C;
step 3: separating ultrafiltration component P with molecular weight cutoff less than 3000Da from the crude peptide powder by ultrafiltration;
step 4: the ultrafiltration component P is prepared into a solution with the concentration of 20mg/ml, and the SephadexG-15 gel chromatography is adopted for chromatography.
Step 5: separating and purifying the ultrafiltration component by RP-HPLC method to obtain the flavor peptide with the amino acid sequence as follows from preserved meat, wherein the flavor and the concentration are the components with the strongest sense of taste (artificial sense evaluation) after chromatography:
Glu-Val-His-Glu-Glu-Val-His(EVHEEVH)。
in a preferred embodiment, the centrifugation conditions in step 2 are 4℃and 12000 Xg for 20 min; the rotary evaporation temperature is 35-38 ℃.
In a preferred embodiment, the method of SephadexG-15 gel chromatography in the step 4 is that the elution flow rate is 1ml/min, the detection wavelength is 220nm, and the loading amount is 4ml.
In the following embodiments of the present invention, the ultrafiltration fraction is chromatographed by Sephadex G-15 gel chromatography, and the fraction having the strongest flavor and thickness (artificial sensory evaluation) is selected and further separated by RP-HPLC.
Wherein the artificial sensory evaluation adopts a method of Taste Dilution Analysis (TDA) to evaluate the dilution factor, namely the TD value of each sample, and the larger the TD value is, the stronger the taste characteristic of the sample is. The TD values were determined as follows: the components to be measured are dissolved in purified water with equal proportion according to the following ratio of 1:1 (volume ratio) was diluted and each dilution level was evaluated using a three-point assay until the taste was not tasted at a certain dilution factor. The dilution factor is the taste dilution factor (TD) of the solution, and the higher the TD value is, the stronger the taste of the component is. Sensory evaluation was completed by 8 sensory evaluators (4 men and 4 women, ages 25 to 30 years), and the sensory evaluators were trained prior to the experiment.
Example 1
A method for extracting, separating and purifying taste peptide separated from preserved meat comprises the following specific steps:
step (1): superfine grinding the peeled 500g Chuan-type traditional preserved meat;
step (2): adding 2L phosphate buffer solution into crushed preserved meat, homogenizing for 10 min in ice bath state 8000 r/min, and centrifuging at 12000 Xg at 4deg.C for 20 min; adding 3 times of absolute ethyl alcohol into the supernatant, and standing for 12 hours at 4 ℃; centrifuging under the same condition again, collecting supernatant, suction filtering, rotary evaporating at 36 deg.C, lyophilizing the concentrated solution to obtain crude peptide powder of conventional Chuan style preserved meat, and storing at-20deg.C;
step (3): separating ultrafiltration component P with molecular weight cutoff less than 3000Da from the crude peptide powder by ultrafiltration, freeze drying the obtained component P, and storing at-20deg.C;
step (4): preparing the ultrafiltration component into a solution with the concentration of 20mg/ml, carrying out chromatography by adopting SephadexG-15 gel chromatography, and taking double distilled water as eluent, wherein the chromatography method is that the elution flow rate is 1ml/min, the detection wavelength is 220nm, and the loading amount is 4ml. The chromatographic results are shown in FIG. 1, the abscissa represents the elution time (min), and the ordinate represents the detection wavelength of 220nm.
As shown in figure 1, 3 separated components obtained in the chromatographic collection and elution process are marked with P1, P2 and P3 in sequence, and are freeze-dried and stored at the temperature of minus 20 ℃ for standby.
Step (5): the P-component and the P1, P2, and P3-components obtained after chromatography were subjected to artificial sensory evaluation by flavor dilution analysis (TDA), the results are shown in fig. 2, the abscissa represents each component, and the ordinate represents the flavor intensity dilution factor of each component.
As shown in FIG. 2, the TD values of fraction P and chromatographic fraction P2 were highest, and the taste sensation was strongest in the same range.
The results of evaluation of the artificial sensory taste characteristics of the ultrafiltration component P and the chromatographic components P1, P2, and P3 are shown in Table 1. As can be seen from Table 1, the descriptions of the flavors of the chromatographic component P2 and the component P are the closest, the fermentation flavor is obvious, and the taste is intense.
Table 1 sensory evaluation results
By combining the graphs with Table 1, it can be considered that the taste of the peptide of the P2 fraction is the most intense and most similar to that of the ultrafiltration fraction P, and therefore the P2 fraction was selected for further RP-HPLC separation and purification.
Step (6): the P2 component is further separated and purified by adopting an RP-HPLC method. The column used was Spurisil C18 (5 μm, 250X 4.6 mm) and the RP-HPLC separation conditions were: isocratic elution, mobile phase A solution is deionized water containing 0.05% TFA (90% -65%); mobile phase B was acetonitrile (10% -35%) with 0.05% TFA. The flow rate is 1mL/min, the sample injection volume is 10ul, the sample concentration is 10mg/mL, the column temperature is 25 ℃, and the elution peak is detected at 214 nm.
The RP-HPLC separation spectrum of the chromatographic fraction P2 is shown in FIG. 3. The abscissa in the figure shows the elution time (min), and the ordinate shows the abundance at the detection wavelength of 214 nm.
As shown in FIG. 3, 6 absorption peaks were collected during the elution process, which were designated as F1, F2, F3, F4, F5 and F6 in this order, and each fraction was freeze-dried and stored at-20 ℃.
Step (7): amino acid sequence analysis: analyzing the amino acid sequence of the component F4 by utilizing electrospray quadrupole time-of-flight mass spectrometry, and identifying that the amino acid sequence of the taste peptide is:
Glu-Val-His-Glu-Glu-Val-His(EVHEEVH)。
FIG. 4 is a secondary mass spectrum of the component. The following table shows the taste peptide information:
TABLE 2 taste peptide information
Example 2
(1) Taste threshold and character of taste peptides
Before the experiment, the panelists were trained on the reference standards of umami, sweet, salty, bitter and sour taste, respectively, monosodium glutamate solution (0.35%, w/v), sucrose solution (1%, w/v), sodium chloride (0.35%, w/v), L-isoleucine solution (0.25%, w/v) and citric acid solution (0.08%, w/v) to achieve more accurate sensory evaluation results.
The taste peptide obtained in example 1 was prepared as a 1% strength synthetic peptide solution according to the principle of the triangle test method, at 1:1, each dilution level solution is gradually diluted by adopting 3-point measurement and is evaluated until the dilution level solution is just distinguished from the other two blank controls (purified water), the concentration of the sample at the moment is the taste threshold value of the sample, and the stimulation threshold value of each evaluation group adopts the average value of the results of each evaluator. The results are shown in Table 3. At the same time, panelists are also asked to verbally describe the tastes they experience, including umami, sweet, sour, salty, and bitter.
TABLE 3 taste Properties and threshold values of aqueous solutions of synthetic peptides
As shown in the table, the sequence (EVHEEVH) has a low taste threshold of 0.22mg/mL and a taste effect better than that of the sodium glutamate solution of 3.5 mg/mL which is lower than that of the umami standard. And exhibits good taste sensation and coordinated mouthfeel, possibly salty peptides.
(2) Enhancement properties of peptide-salt solutions
The salty taste of 0.35% salt solution is used as a sensory standard, the score is 5 minutes, the taste peptide obtained in example 1, namely, the taste peptide with the concentration of 2mg/mL in 0.35% salt solution is added into the salt solution, the salty taste is scored, the standard salt solution is 5 minutes, the synergistic effect of the synthetic peptide and the salt is inspected, and the final result is averaged. The results are shown in Table 4.
TABLE 4 enhancement Properties of peptide-salt solutions
From the results in Table 4, it is clear that the salty taste of the solution can be improved and the flavor can be enhanced by the synergistic effect of the taste peptide EVHEEVH and salt, so that the taste is more coordinated.
(3) Flavor enhancing effect of peptides
The flavor enhancement effect of the sample is examined by scoring the flavor of 0.35% monosodium glutamate solution as a sensory standard, wherein the score is 5 minutes, the flavor development peptide obtained in the example 1, namely, the flavor development peptide concentration in 0.35% monosodium glutamate solution is 2mg/mL, is added into the monosodium glutamate solution, and the final result is averaged. The results are shown in Table 5.
Table 5 umami taste enhancing effect of peptides
From Table 5, it is clear that the sequence EVHEEVH has a strong flavor enhancing effect.
Example 4 hypotensive function test.
An activity assay for ACE inhibitory peptides comprising the steps of:
the in vitro ACE inhibitory activity was determined using modified High Performance Liquid Chromatography (HPLC). A sample solution of the flavor peptide of example 1 of 40 uL and 20 uL of ACE enzyme solution (0.1U/mL, dissolved in 0.1 mol/L borate buffer containing 0.3 mol/L NaCl at pH 8.3) were added to a centrifuge tube, vortexed, mixed well, placed in a 37℃water bath for 5 min, 40 uL of 5 mmol/L HHT solution (dissolved in 0.1 mol/L borate buffer containing 0.3 mol/L NaCl at pH 8.3) was added, and placed in a 37℃water bath for 30 min, and 125 uL of 1 mol/L HCl solution was added to stop the reaction, as a sample set. Meanwhile, the same volume of borate buffer solution is used as a control group to replace the sample solution, and the peak area of the content of the hippuric acid is measured by HPLC. HPLC chromatographic conditions: sepax GP-C18 column (4.6 mmx250 mm,5 um,120A): column temperature 25 ℃; mobile phase a:0.05% TFA-H 2 O, mobile phase B:0.05% tfa-acetonitrile, mobile phase a: b=75%: 25%; flow rate: 1 mL/min; detection wavelength: 228 nm; sample injection amount: 20 uL; analysis time: 20min. The results are shown in FIG. 5.
Wherein A is 1 Peak area for hippuric acid measured in the reaction in the presence of both the taste peptide sample and ACE enzyme of example 1: a is that 2 Peak area of hippuric acid measured without inhibitor in the reaction.
As a result, as shown in FIG. 5, the taste peptide of example 1 had a high ACE inhibition rate, the blood pressure lowering inhibition rate before digestion of the taste peptide was 56.24%, and the inhibition rate after digestion was 58.17%.
The above description is not intended to limit the invention, nor is the invention limited to the examples described above. Variations, modifications, additions, or substitutions will occur to those skilled in the art and are therefore within the spirit and scope of the invention.
Claims (6)
1. A bacon flavor peptide, which is characterized in that the amino acid sequence is shown as SEQ ID NO: 1.
2. The bacon-flavor peptide of claim 1, wherein the bacon-flavor peptide is extracted from bacon.
3. A method for preparing a bacon-flavor peptide according to claim 1 or 2, comprising the steps of:
peeling preserved meat, superfine pulverizing, and extracting crude peptide powder by adopting a phosphate method;
separating ultrafiltration components with molecular weight cutoff less than 3000Da from the crude peptide powder by ultrafiltration;
carrying out chromatography on the ultrafiltration component by adopting a gel chromatography method; and
and separating and purifying the components obtained after chromatography.
4. Use of the bacon-flavor peptide of claim 1 or 2 in food products.
5. Use of the bacon taste peptide according to claim 1 or 2 for preparing a blood pressure lowering health food.
6. A food seasoning comprising the bacon-flavor peptide of claim 1 or 2.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102603870A (en) * | 2012-04-10 | 2012-07-25 | 上海海洋大学 | Taste peptide as well as preparation method and application thereof |
| CN105901534A (en) * | 2016-04-28 | 2016-08-31 | 成都大学 | Low-nitrite non-smoked preserved meat product and processing method thereof |
| CN106819930A (en) * | 2017-02-14 | 2017-06-13 | 成都大学 | A kind of pickle cured meat product and its processing method with ferment local-flavor |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102603870A (en) * | 2012-04-10 | 2012-07-25 | 上海海洋大学 | Taste peptide as well as preparation method and application thereof |
| CN105901534A (en) * | 2016-04-28 | 2016-08-31 | 成都大学 | Low-nitrite non-smoked preserved meat product and processing method thereof |
| CN106819930A (en) * | 2017-02-14 | 2017-06-13 | 成都大学 | A kind of pickle cured meat product and its processing method with ferment local-flavor |
Non-Patent Citations (2)
| Title |
|---|
| C. BAUCHART: "Small peptides (<5 kDa) found in ready-to-eat beef meat", MEAT SCIENCE, vol. 74, pages 658 * |
| 张崟: "微粉碎骨粉对火腿肠食用品质的影响", 成都大学学报, vol. 36, no. 2, pages 148 - 151 * |
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