CN114158715B - Rose Huang Huangyou bolete delicious peptide and preparation method thereof - Google Patents
Rose Huang Huangyou bolete delicious peptide and preparation method thereof Download PDFInfo
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- 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
- A23L27/11—Natural spices, flavouring agents or condiments; Extracts thereof obtained by solvent extraction
-
- 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)
- Engineering & Computer Science (AREA)
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- Polymers & Plastics (AREA)
- Seasonings (AREA)
Abstract
The invention discloses a delicious peptide of a fudge Huang Huangyou bolete and a preparation method thereof, wherein fresh fudge Huang Huangyou bolete is treated by a hot processing mode, gel chromatography is carried out, reverse high performance liquid chromatography is carried out, and after purification, the delicious peptide is identified by nano liter liquid phase mass spectrum tandem mass spectrum, and the prepared delicious peptide has the effects of delicious taste and enhancing freshness and salty taste, effectively improves the richness of the flavor of a seasoning, can blend or enhance the flavor of a flavor food, and is an ideal green and healthy food additive and seasoning.
Description
Technical Field
The invention belongs to the technical field of food additives and preparation methods thereof, and particularly relates to a fumosoroseus Huang Huangyou umami peptide and a preparation method thereof.
Background
The taste sensation can be initially assessed for important nutrients and potentially toxic substances. Subsequent to sour, sweet, bitter, salty, umami is recognized as a fifth basic taste. The flavor is derived from free amino acid, nucleotide, organic acid, organic base, peptide, etc., and can make the taste of food softer, more harmonious and more mellow. The umami peptide can be extracted from natural food or synthesized from amino acids, and has effects of harmonizing or enhancing flavor, and reducing bitter and astringent taste. The molecular weight of the food is about 150-3000Da, and the food is widely distributed in various foods, such as vegetables, seafood, meat, cheese and the like. With the emphasis of people on natural flavor formulations, the extraction and utilization of flavor components from natural sources can effectively relieve the negative emotion of consumers on food additives, and is beneficial to developing products with more special flavors and values.
The edible fungi have unique delicate flavor, are high-protein low-fat food, can be matched with various food materials, ensure that the whole flavor is more delicious and mellow, and are high-quality potential raw materials for preparing the delicate flavor peptide. The Yunnan wild fungus has rich resources and various varieties, is a non-abundant delicious, and the fresh and delicious rose Huang Huangyou bolete is favored by consumers because of high nutrition and unique flavor. However, current research on wild bacteria focuses on their origin identification and medicinal value, and few reports on taste components are available. Fewer studies were performed on the fumosoroseus Huang Huangyou bolete. Most of the preparation of the umami peptide adopts methods of enzymolysis, thermal processing, fermentation and the like, and the enzymolysis and fermentation technology can change the taste of the raw materials, compared with the prior art, the thermal processing technology retains the original taste of the raw materials. And in view of safety, the wild bolete also needs to be eaten after being cooked. Therefore, the hot processing technology is selected to prepare the fumosoroseus Huang Huangyou delicious peptide, natural safety is ensured, and meanwhile, the delicious taste is improved, and the method has positive influence on preparing the wild fungus seasoning.
Disclosure of Invention
The invention provides a fumosoroseus Huang Huangyou bolete delicate flavor peptide and a preparation method thereof, wherein the delicate flavor peptide is delicious and mellow in taste and natural in flavor, has a certain effect of enhancing freshness and salty taste, can replace salt and monosodium glutamate seasonings in daily life, has a low delicate flavor threshold, and is an ideal natural seasoning and a delicious enhancer.
The invention is realized by the following technical scheme:
the preparation method of the fumosoroseus Huang Huangyou bolete umami peptide comprises the following steps:
(1) Treatment of Boletus fumosoroseus Huang Huangyou: selecting fresh Huang Huangyou bolete of uniform size without opening umbrella, cleaning, draining, weighing, cutting, adding ultrapure water in proportion for homogenizing for later use;
(2) Preparing hydrolysate freeze-dried powder: boiling the homogenate at high temperature to obtain a fumosoroseus hydrolysate Huang Huangyou, cooling to room temperature, filtering with gauze, centrifuging the obtained filtrate, and taking supernatant for freeze-drying;
(3) Gel chromatography separation: the freeze-dried powder of the hydrolysis liquid is redissolved by ultrapure water, gel chromatography analysis is carried out, sephadex G-15 separation and ultrapure water elution are carried out, so as to obtain a separated component, freeze-drying is carried out, the most fresh component is evaluated in a sense, the most fresh component is subjected to reverse liquid chromatography purification and then freeze-drying, and the most fresh component is the liver fungus umami peptide of the rose Huang Huangyou in a sense.
In the step (1), the mass volume ratio g of the rose Huang Huangyou bolete to the ultrapure water is 1:4 (w/v).
And (3) high-temperature cooking conditions in the step (2): the temperature is 90-100 ℃ and the duration is 100-120 min; the 8 layers of gauze were filtered and centrifuged at 5000rpm for 20min at 4 ℃.
The chromatographic conditions of the gel in the step (3) are as follows: the concentration of the hydrolysate freeze-dried powder is 20mg/mL, the flow rate is 1.0mL/min, the loading amount is 1mL, and the detection wavelength is 220nm.
The reverse high performance liquid chromatography purification conditions in the step (3) are as follows: the chromatographic column AQ-C18 (10×250nm,5 μm) has a column temperature of 30deg.C, a loading amount of 50μl, a flow rate of 1.0mL/min, a detection wavelength of 214nm, and a running time of 25min, wherein the eluent is obtained by mixing A with B in a volume ratio of A to B of (90:10), A is an ultrapure water solution of trifluoroacetic acid with a volume fraction of 0.05%, and B is an acetonitrile solution of trifluoroacetic acid with a volume fraction of 0.05%.
Adopting Nano-LC-MS/MS to identify amino acid sequence of the most fresh liquid phase component, wherein the MS conditions are as follows: ESI+ mode, data dependent scanning mode, full scan acquisition (m/z 100-1500) in an orbitrap with resolution 70000 (AGC 3e 6) resulted in the identification of four umami peptides with sequences GSAQA, ISKEK, LPKERLYV and VENQQLAL, i.e., gly-Ser-Ala-Gln-Ala, ile-Ser-Lys-Glu-Lys, leu-Pro-Lys-Glu-Arg-Leu-Tyr-Val, val-Glu-Asn-Gln-Leu-Ala-Leu.
The invention takes natural wild bolete as a base material, utilizes a high-temperature boiling method and a freeze-drying technology to ensure food safety and fully lock the delicate flavor components of the bolete, adopts a series of separation and purification means, gradually screens out the most delicate flavor components according to sensory evaluation, and finally obtains the fudge Huang Huangyou bolete delicate flavor peptide which has delicate flavor and delicate flavor enhancing effect and has the delicate flavor threshold range of 0.20mmol/L-1.39mmol/L.
The rose Huang Huangyou bolete delicious peptide has mellow and natural delicious taste and high strength, can generate delicious taste and enhance the delicious taste, has obvious effect of enhancing the delicious taste and the salty taste of chicken soup, can reduce the use of salt and sodium glutamate, and can be widely applied to preparing the wild fungus seasonings for enhancing the delicious taste and reducing the salt.
Drawings
FIG. 1 is a chromatographic chart after Sephadex G-15 gel chromatographic separation;
FIG. 2 is a flavor profile of gel components F1 and F2;
FIG. 3 is a RP-HPLC separation spectrum of gel fraction F2;
FIG. 4 is a flavor profile of liquid phase components F2-1, F2-2 and F2-3;
FIG. 5 is a mass spectrum of the F2-2 component (A: GSAQA, B: ISKEK, C: LPKERLYV, D: VENQQLAL);
FIG. 6 is the effect of synthetic peptides on the taste of real chicken soup.
Detailed Description
The present invention is further illustrated below with reference to specific examples, which are not intended to limit the scope of the invention in any way.
Example 1
A preparation method of the fumosoroseus Huang Huangyou bolete delicious peptide comprises the following specific steps:
(1) Treatment of Boletus fumosoroseus Huang Huangyou: selecting fresh bolete Huang Huangyou with uniform size without opening an umbrella, cleaning, draining, weighing, cutting, adding ultrapure water into the bolete to be homogenized for later use according to the mass volume ratio g of the bolete Huang Huangyou to the ultrapure water of 1:4;
(2) Preparing hydrolysate freeze-dried powder: the homogenate is boiled at high temperature to obtain the rhodo Huang Huangyou bolete hydrolysate, and the high-temperature boiling conditions are as follows: the temperature is 90-100 ℃ and the duration is 100-120 min; cooling to room temperature, filtering with 8 layers of gauze, centrifuging at 5000rpm for 20min at 4deg.C, collecting supernatant, and lyophilizing;
(3) Gel chromatography separation:
dissolving the freeze-dried powder of the rhodozyma hydrolysate of the rose Huang Huangyou into ultrapure water to prepare a loading solution with the concentration of 20mg/mL, filtering the loading solution by a 0.45 mu m water phase filter membrane, loading 1mL of loading solution into a Sephadex G-15 gel filtration chromatographic column, eluting the ultrapure water at the flow rate of 1.0mL/min at the temperature of 25+/-2 ℃, recording the ultraviolet absorption at 220nm, collecting, combining and freeze-drying the same fraction for sensory evaluation, separating the two components by gel chromatography, and freeze-drying the two components to obtain F1 and F2, and storing the two components in a refrigerator at the temperature of-80 ℃;
(4) Sensory evaluation
The sensory analysis laboratory at a temperature of 23.+ -. 2 ℃ was run and the panel consisted of 10 (5 men and 5 women, ages 23-30) screened and trained panels that did not have any olfactory and gustatory defects, and two hours prior to evaluation, had no contact with any pungent odor, the basic taste reference solution was set as follows: sweet taste (1% sucrose solution by mass); salty taste (mass fraction 0.35% sodium chloride solution); bitter taste (0.08% caffeine solution by mass); umami taste (mass fraction 0.35% monosodium glutamate solution); sour taste (mass fraction 0.08% citric acid solution), taste intensity expressed in 10 minutes, sensory intensity of reference solution uniformly 5 minutes, panelists randomly tasting component solution reconstituted with deionized water, giving corresponding fractions of taste attribute, taking average value as final sensory evaluation result and identifying with radar chart; after each sample is tasted, rinsing with 60mL deionized water for 3 times, resting for 1-2min, and then performing subsequent tasting;
FIG. 2 shows that the component selected for the most intense umami taste in the GFC fraction is F2, also with a characteristic bolete-like taste, whereby the F2 component is selected for continued purification;
(5) Reverse high performance liquid chromatography purification:
dissolving F2 component freeze-dried powder into 10mg/mL solution with ultrapure water, filtering with 0.22 μm water phase filter membrane, mixing eluent A and B with volume ratio A of B=90:10 on C18 high performance liquid chromatographic column (AQ-C18×250mm,5 μm), wherein A is 0.05% trifluoroacetic acid ultrapure water solution, B is 0.05% trifluoroacetic acid acetonitrile solution, setting column temperature at 30deg.C, loading sample amount of 50 μl, flow rate of 1.0mL/min, detection wavelength of 214nm, running for 25min, collecting components F2-1, F2-2 and F2-3 respectively, concentrating, freeze drying;
sensory evaluation was performed according to the method of step (4), and the most prominent umami taste of the F2-2 component in FIGS. 3 and 4 was namely freeze-dried powder of the umami peptide of Boletus roseus.
Adopting Nano-LC-MS/MS to identify amino acid sequence of the most fresh liquid phase component, and adopting Nano-liter liquid phase chromatographyThe analytical column was a C18 reverse phase analytical column (150 μm i.d.× 150mm,packed with Acclaim PepMap RPLC C18,1.9 μm,
) The mobile phase A is a mixed solution of 99.9% of water by volume fraction and 0.1% of formic acid by volume fraction, the mobile phase B is a mixed solution of 80% of acetonitrile by volume fraction and 0.1% of formic acid by volume fraction, and the liquid phase elution gradient is as follows: 0-2min,4% -8% B; 2-45 min, 8-28% B; 45-55 min, 28-40% B; 55-56 min, 40-95% B; 56-66 min,95% B, the balance being water; the flow rate of the mobile phase is 600nL/min;
the MS conditions were: ESI+ mode, data dependent scanning mode, full scan acquisition (m/z 100-1500) is performed in an orbitrap with resolution 70000 (AGC 3e 6).
After resolution, the peptide sequence was analyzed to be GSAQA, ISKEK, LPKERLYV and VENQQLAL, i.e., the amino acid sequence was: gly-Ser-Ala-Gln-Ala, ile-Ser-Lys-Glu-Lys, leu-Pro-Lys-Glu-Arg-Leu-Tyr-Val, val-Glu-Asn-Gln-Gln-Leu-Ala-Leu, see FIG. 5 for details.
In order to verify that the umami peptide shows a freshness effect, the identified polypeptide sequence is entrusted to the Shanghai Jier biochemical limited company to be synthesized by adopting an Fmoc solid phase method, and a sensory evaluation is used as a standard, and an evaluation group carries out taste description analysis on a synthetic peptide solution; then, according to the taste dilution analysis (TAD), 1mg/mL of the synthetic peptide solution was gradually diluted in a ratio of 1:1 (v/v) until the panelist clearly distinguished the taste difference between a certain dilution concentration and two blanks (deionized water), and the average of the individual threshold values of the panel was taken as the freshness threshold value of the polypeptide.
The taste profile and freshness threshold of the liver-of-rose Huang Huangyou bolete umami peptide solution are shown in table 1 below:
TABLE 1
| Umami peptide | Fresh threshold (mmol/L) | Description of taste |
| GSAQA | 1.39 | Obvious sweet taste and slightly fresh |
| ISKEK | 0.75 | Fresh taste, sweet taste and slightly astringent taste |
| LPKERLYV | 0.25 | Moderate umami, salty and sweet, slightly sour |
| VENQQLAL | 0.20 | Intense umami, salty and sweet, sour |
Constructing a synthetic peptide and real chicken soup model, cleaning and cutting real chicken soup (200 g chicken breast, adding 1L of water and 3g of ginger, keeping micro boiling for 2 hours, cooling to room temperature, filtering out meat blocks, adding 5g of salt into the chicken soup, and fixing the volume to 1L) to obtain mother liquor, preparing 1mg/mL synthetic peptide and 3.5mg/mL monosodium glutamate solution (MSG), and measuring the delicate flavor, salty taste and sweet taste of each solution, wherein the sensory intensity of the chicken soup taste is designated as 5 minutes, and the evaluation mode is consistent with the sensory evaluation standard; as shown in fig. 6, in real chicken soup, 4 peptide fragments in the umami peptide can well improve the umami taste, the salty taste and the sweet taste, wherein GSAQA and ISKEK show better salty taste enhancing ability than lpkenyv and VENQQLAL; the VENQQLAL has the strongest flavor and freshness enhancing ability.
Claims (1)
1. Use of a umami peptide isolated from bolete fumosoroseum of fumosoroseum Huang Huangyou in a flavouring, characterized in that the umami peptide consists of an amino acid sequence such as: gly-Ser-Ala-Gln-Ala, ile-Ser-Lys-Glu-Lys, leu-Pro-Lys-Glu-Arg-Leu-Tyr-Val and Val-Glu-Asn-Gln-Gln-Leu-Ala-Leu.
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| CN202111487724.XA CN114158715B (en) | 2021-12-07 | 2021-12-07 | Rose Huang Huangyou bolete delicious peptide and preparation method thereof |
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| CN202111487724.XA CN114158715B (en) | 2021-12-07 | 2021-12-07 | Rose Huang Huangyou bolete delicious peptide and preparation method thereof |
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| CN114158715A CN114158715A (en) | 2022-03-11 |
| CN114158715B true CN114158715B (en) | 2023-06-23 |
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