Homologous double-phase reconstituted agaricus bisporus umami composite seasoning particles and a preparation method thereof

Abstract

The application discloses a homologous double-phase reconstructed agaricus bisporus umami composite seasoning particle and a preparation method thereof. The same batch of agaricus bisporus fruiting bodies is divided into umami extraction components and carrier preparation components; the umami extraction components are subjected to low-oxygen pulse electric field treatment and continuous phased enzymolysis to obtain umami extraction liquid, and the umami extraction liquid is subjected to low-temperature reduced-pressure concentration to form a first umami loading liquid with low solid content and a second umami coating liquid with high solid content; the carrier preparation components are subjected to low-temperature drying, ultrafine pulverization and wet agglomeration to form porous cores, and the remaining first umami loading liquid is subjected to pulse negative pressure infiltration to enter the internal pores of the porous cores to form umami internal loading phases, and the second umami coating liquid is subjected to fluidized bed spraying to form umami external coating phases. The method is beneficial to improving the loading efficiency and particle dispersibility of umami extracts and reducing the static sedimentation rate.

Description

Technical Field

[0001] This invention relates to the field of edible fungi processing technology, and in particular to a homologous biphase reconstructed Agaricus bisporus umami compound seasoning granule and its preparation method. Background Technology

[0002] Button mushroom ( Agaricus bisporus Button mushrooms, with their delicious flavor and rich content of amino acids, nucleotides, polysaccharides, and dietary fiber, are an important edible fungus. However, the fruiting bodies of button mushrooms have a high water content and vigorous post-harvest respiration and metabolism, making them prone to opening, browning, and spoilage. During fresh sales, canning, salting, and drying processes, byproducts such as substandard mushrooms, stems, and mushroom fragments are also generated. Although these byproducts do not meet the appearance requirements for fresh sales or canning, they still retain rich flavor and nutritional components, making them valuable for further processing. The umami flavor of button mushrooms mainly comes from flavor amino acids such as glutamic acid and aspartic acid, as well as flavor nucleotides such as 5′-guanylic acid. Flavor amino acids and 5′-flavor nucleotides have a synergistic flavor-enhancing effect, and small molecule peptides, organic acids, and soluble sugars also contribute to the flavor characteristics of mushroom seasonings. Therefore, how to release and retain different types of flavor components is a problem that needs to be solved in the preparation of high-quality mushroom seasonings.

[0003] The preparation of existing edible fungi seasoning products typically involves processes such as drying and pulverizing, hot water or enzymatic extraction, mixing of excipients, and granulation. Related food processing technologies also involve using pulsed electric fields to promote the release of intracellular components from raw materials, using vacuum to allow liquid components to enter porous matrices, and surface coating of particles. Some existing technologies combine vacuum wetting of porous matrices with subsequent surface coating; however, the treated objects are mainly starch-puffed matrices or exogenous adsorbent carriers, and the loaded components are mainly oils or exogenous flavorings. They do not involve preparing umami extracts and mushroom-derived porous cores separately from the same batch of button mushrooms, nor do they involve grading homologous umami extracts according to different solid content and using them separately for internal loading and external coating of porous cores.

[0004] CN108354160B discloses a mushroom salt seasoning made from enzymatic hydrolysate of button mushrooms. The method involves washing and slicing fresh button mushrooms, blanching them in boiling water, mixing the blanched mushrooms with the blanching liquid to form a homogenate, and then performing enzymatic hydrolysis using a two-step process. The hydrolysate is obtained by centrifugation and filtration, and then microwave-dried at 100-120°C and ground into powder to obtain button mushroom enzymatic hydrolysate. Subsequently, the button mushroom enzymatic hydrolysate is mixed with salt, maltodextrin, and starch, and then subjected to rotary granulation and fluidized bed drying to produce the mushroom salt seasoning. However, the high temperature treatment in this method may cause loss of characteristic aroma components and may affect the color and natural flavor of the enzymatic hydrolysate. Furthermore, the water-soluble umami components and insoluble solid components in the same batch of button mushroom raw materials remain separated, failing to achieve synergistic utilization of the water-soluble umami components and the mushroom-derived carrier components.

[0005] For example, CN103584073B discloses a method for preparing edible fungus flavored food, which involves mixing edible fungus slices, strips, or cubes with a flavor impregnation liquid and using a combination of vacuum impregnation and atmospheric pressure impregnation to allow the flavor components to penetrate the edible fungus tissue. This method targets edible fungi that retain their natural tissue morphology and is mainly used to introduce exogenous flavoring components. It does not involve the formation of porous cores from the aggregation of mushroom ultrafine powder, nor does it address the coordination between internal particle loading and external coating.

[0006] Therefore, existing technologies still need to address the issues of insufficient utilization of the water-soluble umami components and insoluble solid components of Agaricus bisporus, as well as the insufficient uniformity of loading and particle stability when umami extracts are simply mixed with mushroom powder. There is an urgent need to develop a method for preparing seasoning particles that can perform graded processing and structural reconstruction of homologous Agaricus bisporus components. Summary of the Invention

[0007] To address the shortcomings of existing technologies, the technical problem this invention aims to solve is to provide homologous two-phase reconstructed Agaricus bisporus umami compound seasoning particles and their preparation method, thereby resolving the issues of insufficient utilization of the water-soluble and insoluble solid components of Agaricus bisporus, and uneven loading and insufficient particle stability caused by simple mixing of umami extracts and powders in existing technologies. To solve the above technical problems, this invention provides the following technical solution: The first aspect of this application relates to a method for preparing homologous biphase reconstructed mushroom umami compound seasoning particles, comprising the following steps: Step S1: Clean, cut and mix the same batch of button mushroom fruiting bodies, and divide the mixed button mushroom fruiting bodies into umami extract components and carrier preparation components according to dry weight. Step S2: Mix the dry weight of the umami extract with the weight of water at a ratio of 1:3 to 1:8 to form a mushroom broth. Before pulsed electric field treatment, introduce nitrogen gas into the closed treatment system connected to the mushroom broth for displacement until the oxygen concentration sensor in the gas phase space of the closed treatment system detects that the oxygen volume content is stable at 2% to 5%, and maintain the oxygen volume content during the pulsed electric field treatment. Perform pulsed electric field treatment on the mushroom broth so that the energy input per unit mass of mushroom broth is 40 to 80 kJ / kg, and control the temperature of the mushroom broth after treatment to not exceed 35 ℃. Step S3: In the same enzymatic hydrolysis container, without solid-liquid separation and enzyme inactivation between adjacent enzymatic hydrolysis stages, the mushroom liquid treated with pulsed electric field is sequentially subjected to cell wall hydrolysis, neutral protease hydrolysis, flavor protease hydrolysis and nucleic acid hydrolysis. After all enzymatic hydrolysis is completed, enzyme inactivation and solid-liquid separation are performed to obtain a umami extract containing flavor amino acids, peptides with a molecular weight not higher than 3 kDa and 5′-flavor nucleotides. Step S4: Under the condition that the temperature is not higher than 45 ℃, the umami extract is concentrated under reduced pressure to a solid content of 8% to 12%, a portion of which is taken as the first umami loading liquid, and the remaining portion is further concentrated under reduced pressure to a solid content of 25% to 35% as the second umami coating liquid. Step S5: The carrier preparation components are subjected to low-temperature drying and ultra-fine pulverization to obtain Agaricus bisporus fruiting body ultra-fine powder. 5% to 15% of the total mass of the first umami loading liquid is used as the agglomeration liquid for wet agglomeration, and pre-dried to a moisture content of no more than 8% to form a porous core with open internal channels. The porosity of the porous core is 30% to 50%, and the liquid absorption per unit dry mass is 0.8 to 1.4 g / g. Step S6: Add the remaining first umami loading liquid to the porous core at a mass ratio of (3-5):1 (dry basis) of (3-5):1. Perform 2-4 pulsed immersion cycles of "vacuuming-restoring normal pressure" under a gauge pressure of -0.08 to -0.06 MPa to allow the remaining first umami loading liquid to enter the internal channels of the porous core. After immersion, remove and recover the unabsorbed flowable liquid on the surface of the porous core. Then, perform a first-stage low-temperature drying to allow the umami extract solids introduced during the wet agglomeration process in Step S5 and the umami extract solids that entered the internal channels during the pulsed negative pressure immersion process to form an internal umami support phase. Step S7: Place the porous core containing the umami inner carrier phase in a fluidized state, and spray the second umami coating liquid onto the outer surface of the porous core. Perform a second stage of low-temperature drying under the condition that the material temperature is not higher than 50 ℃ to form an umami outer coating phase, and obtain button mushroom umami compound seasoning particles including the porous core, the umami inner carrier phase distributed in the internal channels of the porous core, and the umami outer coating phase distributed on the outer surface of the porous core.

[0008] Furthermore, the dry basis mass ratio of the umami extract component to the carrier preparation component is (1-3):1; steps S1 to S3 are all carried out in a closed processing space, and the oxygen volume content in the closed processing space is maintained at 2.5% by means of an oxygen concentration sensor and nitrogen flow rate adjustment.

[0009] Furthermore, the pulsed electric field treatment employs a bipolar square wave with an electric field strength of 15–25 kV / cm and a pulse width of 1–20 μs. Each material unit receives a cumulative number of pulses of 20–200. Based on the measured conductivity of the mushroom slurry, the pulse frequency and slurry flow rate are adjusted to ensure that the combination of electric field strength, pulse width, and cumulative pulse count satisfies the requirement that the energy input per unit mass of mushroom slurry is 40–80 kJ / kg, and that the temperature of the mushroom slurry after pulsed electric field treatment does not exceed 35 °C.

[0010] Further, the cell wall hydrolytic enzyme includes at least one of β-glucanase and cellulase, with β-glucanase added at 120 U per gram of umami extract raw material (dry basis) and cellulase added at 60 U per gram of umami extract raw material (dry basis); the neutral protease added at 1000 U per gram of umami extract raw material (dry basis) and the flavor protease added at 500 U per gram of umami extract raw material (dry basis); the nucleic acid hydrolytic enzyme includes at least one of nuclease P1 and 5′-phosphodiesterase, with nuclease P1 added at 80 U per gram of umami extract raw material (dry basis) and 5′-phosphodiesterase added at 80 U per gram of umami extract raw material (dry basis).

[0011] Furthermore, the pH, temperature, and time for each enzymatic hydrolysis stage are selected from any complete combination of the following parameters: The first parameter combination is as follows: cell wall hydrolase hydrolysis at pH 5.5, temperature 40 ℃, and time 0.5 h; neutral protease hydrolysis at pH 6.5, temperature 42 ℃, and time 0.5 h; flavor protease hydrolysis at pH 6.0, temperature 40 ℃, and time 0.5 h; and nucleic acid hydrolase hydrolysis at pH 5.0, temperature 40 ℃, and time 0.5 h. The second parameter combination is as follows: cell wall hydrolase hydrolysis at pH 5.5, temperature 45 ℃, and time 1 h; neutral protease hydrolysis at pH 6.8, temperature 48 ℃, and time 1 h; flavor protease hydrolysis at pH 6.2, temperature 45 ℃, and time 1 h; and nucleic acid hydrolase hydrolysis at pH 5.5, temperature 45 ℃, and time 0.75 h. The third parameter combination is as follows: the enzymatic hydrolysis of cell wall hydrolysates is performed at pH 5.5, temperature 50 ℃, and time 2 h; the enzymatic hydrolysis of neutral protease is performed at pH 7.2, temperature 52 ℃, and time 2 h; the enzymatic hydrolysis of flavor protease is performed at pH 7.0, temperature 50 ℃, and time 2 h; and the enzymatic hydrolysis of nucleic acid hydrolysates is performed at pH 6.5, temperature 50 ℃, and time 2 h.

[0012] Furthermore, the D of the Agaricus bisporus fruiting body ultrafine powder 50 The diameter is 20–35 μm; the D of the porous core is... 50 The size ranges from 180 to 320 μm.

[0013] Furthermore, the mass ratio of the solid umami extract in the first umami loading liquid to that in the second umami coating liquid is 1:(0.5~2), wherein the first umami loading liquid used for wet agglomeration in step S5 accounts for 5%~15% of the total mass of the first umami loading liquid.

[0014] Furthermore, in step S6, the vacuum is maintained for 3-8 minutes each time, and the pressure is restored to normal for 3-8 minutes. During the soaking process, the mixture is slowly turned over at 20-40 rpm. After soaking, the excess liquid on the surface is removed and the mixture is dried at a temperature not higher than 45 ℃ until the moisture content is not higher than 8%. In step S7, the moisture content of the mushroom umami compound seasoning granules obtained is not higher than 6%.

[0015] A second aspect of this application relates to a homologous two-phase reconstructed Agaricus bisporus umami composite seasoning particle, comprising a porous core formed by the agglomeration of Agaricus bisporus fruiting body ultrafine powder, an umami inner carrier phase distributed in the internal channels of the porous core, and an umami outer coating phase distributed on the outer surface of the porous core, wherein both the umami inner carrier phase and the umami outer coating phase are formed from Agaricus bisporus umami extract; the dry basis mass ratio of the Agaricus bisporus umami extract to the Agaricus bisporus fruiting body ultrafine powder constituting the porous core is (0.5-2):1, and the dry basis mass ratio of the umami inner carrier phase to the umami outer coating phase is 1:(0.5-2); the D of the composite seasoning particle is... 50 The particle size is 180–400 μm, and the content of free glutamic acid is not less than 8.0 mg / g and the content of 5′-guanylic acid is not less than 0.40 mg / g, based on the dry basis of the product.

[0016] Furthermore, the moisture content of the button mushroom umami compound seasoning granules is not higher than 6%, and the water activity is not higher than 0.40.

[0017] Compared with the prior art, the present invention has the following beneficial effects: (1) In this invention, the same batch of Agaricus bisporus fruiting bodies are divided into a flavor extraction component and a carrier preparation component according to their dry weight. The flavor extraction component is subjected to low-oxygen pulsed electric field treatment and continuous staged enzymatic hydrolysis to obtain a flavor extract. The carrier preparation component is subjected to low-temperature drying and ultra-fine pulverization to obtain Agaricus bisporus fruiting body ultrafine powder. The two are then recombined to form composite flavoring particles. This enables the carrier preparation component to form a mushroom-derived porous core, and the water-soluble flavor extract obtained from the flavor extraction component is reloaded onto the porous core, improving the synergistic utilization of the carrier preparation component and the flavor extract in the same batch of raw materials, and reducing the dependence on exogenous carriers such as maltodextrin and starch. The hypoxic pulsed electric field treatment first improves the permeability of the mushroom tissue, followed by continuous cell wall hydrolysis, protein hydrolysis and nucleic acid hydrolysis in the same container. This promotes the release of soluble components such as glutamic acid, aspartic acid, peptides with a molecular weight not exceeding 3 kDa and 5′-flavor nucleotides. The hypoxic environment also helps to reduce the adverse effects of oxidation on flavor components during the treatment process, thereby improving the yield of soluble solids in the umami extract and the retention level of flavor components.

[0018] (2) In this invention, the low-solids umami loading liquid is first introduced into the internal channels of the mushroom ultrafine powder porous core under negative pressure, and then the high-solids umami coating liquid is sprayed onto the outer surface of the porous core to form a composite particle composed of a porous core, an inner umami carrier phase and an outer umami coating phase. This is different from the simple mixing or one-time surface spraying of umami extract and carrier powder. It is beneficial to improve the loading efficiency of umami extract, reduce the excessive concentration of umami extract on the particle surface, improve the dispersion performance of composite seasoning particles in water, and reduce the sedimentation rate after standing. The resulting product has a high umami extract loading, good umami coordination and low water content and water activity. Attached Figure Description

[0019] Figure 1 This is a flowchart of the method for preparing the Homologous Two-Phase Reconstructed Agaricus bisporus Umami Compound Seasoning Granules of the present invention. Detailed Implementation

[0020] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions in the embodiments of this invention will be described more clearly and completely below. Obviously, the described embodiments are only some embodiments of this invention, and not all embodiments. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0021] The term "same batch of button mushroom fruiting bodies" in this invention refers to button mushroom fruiting bodies originating from the same batch of raw materials, harvested at the same time, and stored under the same conditions, and thoroughly mixed before grouping. As raw material control conditions, the moisture content of the raw materials is 88%–93%, and they are processed within 48 hours of harvesting, or stored at 2–8°C for no more than 24 hours.

[0022] The "umami internal support phase" of this invention includes umami extract solids introduced and distributed within the porous core body by a portion of the first umami loading liquid during wet agglomeration, and umami extract solids formed by the remaining first umami loading liquid entering the internal channels of the porous core body during pulsed negative pressure wetting and then drying. The "umami external coating phase" refers to the umami extract solids distributed on the outer surface of the porous core body after the second umami coating liquid is fluidized bed sprayed and dried.

[0023] The term "open internal pores" in this invention refers to the presence of pores within the porous core that can communicate with the external liquid; the porosity measured by the true density-apparent density method is 30% to 50%, and the liquid absorption per unit dry mass measured according to the method described in the instruction manual is 0.8 to 1.4 g / g.

[0024] The "low-oxygen condition" referred to in this invention means maintaining the gaseous oxygen volume content at 2% to 5% by introducing nitrogen gas into the sealed processing space of the corresponding processing equipment; the preferred embodiment is to control it at 2.5%. The gaseous oxygen volume content is continuously detected by an oxygen concentration sensor and is regulated in a closed loop by a nitrogen flow regulating valve and an exhaust valve.

[0025] In this embodiment of the invention, the pulsed electric field treatment employs a continuous processing chamber, and the pulse waveform is a bipolar square wave. The energy input per unit mass is calculated as the ratio of the voltage-current integral value of the pulsed power supply within the processing time to the mass of the processed material, and is verified using the measured value from the equipment's online energy meter. The electric field strength, pulse width, cumulative pulse count, liquid conductivity, and processing flow rate are mutually matched, with energy input per unit mass and post-processing temperature as the main control indicators. These parameters can be freely combined within their respective ranges. Representative combinations of pulsed electric field parameters, as well as the pulsed electric field equipment and liquid parameters, are shown in Tables 1 and 2 below.

[0026] Table 1 Representative parameter combinations of pulsed electric field Table 2 Pulse Electric Field Equipment and Feed Parameters The β-glucanase, cellulase, neutral protease, flavor protease, nuclease P1, and 5′-phosphodiesterase used in this invention are all food-grade commercial enzyme preparations. Different commercial enzyme preparations should be retested under uniform activity assay conditions, and the amount added should be calculated based on the actual measured enzyme activity. Specific parameters are shown in Table 3.

[0027] Table 3. Enzyme sources, specifications, and unified enzyme activity definition In the wet agglomeration process of this invention, the bed material quantity is 3–8 kg, the apparent fluidization velocity is 0.6–1.0 m / s, the atomization pressure is 0.15–0.25 MPa, the spray rate is 3–6 g / (min·kg bed material), the nozzle outlet height from the static bed surface is 150–250 mm, and the material temperature is 35–45°C. By controlling the fluidization velocity, atomized droplet size, and spray rate, the agglomeration liquid connects the ultrafine powder particles in the form of liquid bridges, while avoiding over-wetting that causes particle densification, thereby forming a porous core with open pores.

[0028] The dry basis mass of the porous nucleus referred to in this invention refers to the mass of dry matter actually contained in the porous nucleus after wet agglomeration, pre-drying and sieving in step S5, which enters step S6. It includes the dry basis mass of the mushroom fruiting body ultrafine powder and the mass of umami extract solids introduced by the first umami loading liquid during the wet agglomeration process, but does not include the water content.

[0029] Example 1: This embodiment provides a method for preparing homologous biphase reconstructed mushroom umami compound seasoning granules.

[0030] Step S1, Raw Material Grouping: Take 100 kg of fresh button mushroom fruiting bodies from the same batch, wash, cut, and mix them evenly. The moisture content of the raw materials is 90.2%, and they should be put into processing within 18 hours after harvesting. After determining the moisture content of the raw materials, they are divided into umami extraction components and carrier preparation components according to a dry basis mass ratio of 2:1. During the cutting, mixing, and grouping of the raw materials, nitrogen gas is continuously introduced into the processing space to maintain an oxygen volume content of 2.5%.

[0031] Step S2, Low-Oxygen Pulsed Electric Field Treatment: Five times the dry weight of deionized water was added to the umami extract component, and the mixture was homogenized at low temperature to obtain a button mushroom substrate solution. The solution was then transported to a continuous pulsed electric field treatment chamber with an electrode spacing of 10 mm. A voltage of 20 kV was applied, the pulse waveform was a bipolar square wave, and the pulse frequency was 100 Hz. The measured conductivity of the solution was 0.15 mS / cm, and the treatment flow rate was 30 L / h. Nitrogen gas was continuously introduced into the sealed treatment space to maintain an oxygen volume content of 2.5%. The electric field strength was 20 kV / cm, the pulse width was 10 μs, and each material unit received a total of 100 pulses. The measured energy input per unit mass of the equipment was 60 kJ / kg, and the temperature of the treated solution was 32 ℃.

[0032] Step S3, Continuous Staged Enzymatic Hydrolysis: The following treatments are performed sequentially within the same sealed enzymatic hydrolysis tank, without solid-liquid separation or enzyme inactivation between adjacent stages: Adjust the pH of the feed solution to 5.5, add β-glucanase and cellulase at 120 U and 60 U per gram of dry raw material, respectively, and hydrolyze at 45 °C for 60 min; adjust the pH to 6.8, add neutral protease at 1000 U / g dry raw material, and hydrolyze at 48 °C for 60 min; adjust the pH to 6.2, add flavor protease at 500 U / g dry raw material, and hydrolyze at 45 °C for 60 min; adjust the pH to 5.5, add nuclease P1 at 80 U / g dry raw material, and hydrolyze at 45 °C for 45 min. The oxygen volume content in the gas phase space of the enzymatic hydrolysis tank is maintained at 2.5%. After all enzymatic hydrolysis was completed, the enzymes were inactivated by maintaining the temperature at 90 °C for 10 min. Then, the mixture was centrifuged at 5000 × g for 15 min and filtered through a 100 μm filter to obtain the umami extract.

[0033] Step S4, Fractional Concentration of Umami Extract: The umami extract obtained in Step S3 is concentrated under reduced pressure at a temperature not exceeding 45 ℃ and a gauge pressure of -0.08 MPa until the solid content is 10%. A portion of this is collected as the first umami loading solution. The remaining umami extract is further concentrated under reduced pressure until the solid content is 30%, which is used as the second umami coating solution. The mass ratio of umami extract solids in the first umami loading solution to that in the second umami coating solution is controlled to be 1:1.

[0034] Step S5, Porous Core Preparation: The carrier preparation components are vacuum dried at 45 ℃ and a gauge pressure of -0.08 MPa until the moisture content is no higher than 8%, and then ultra-finely pulverized at a pulverization temperature no higher than 25 ℃ to D. 50The particle size is 25 μm. Wet agglomeration is performed using a top-spray fluidized bed granulation device. The fluidized bed includes an airflow distribution plate at the bottom of the processing chamber, atomizing nozzles above the bed, and a temperature sensor for detecting the material temperature. The atomizing nozzles spray liquid downwards along the axial direction of the processing chamber. 5.0 kg of Agaricus bisporus fruiting body ultrafine powder is added to the fluidized bed for each batch. The nozzle orifice diameter is 1.0 mm, and the nozzle outlet is 200 mm above the surface of the stationary bed.

[0035] Start the fluidizing blower and control the apparent fluidizing velocity at 0.6–1.0 m / s and the inlet air temperature at 48–52℃ to ensure full fluidization of the ultrafine powder. After the material temperature stabilizes at 38–42℃, use 10% of the total first flavor loading liquid as the agglomeration liquid and perform top-spray wet agglomeration under the conditions of an atomization pressure of 0.15–0.25 MPa and a spray rate of 3–6 g / (min·kg bed material). During the spraying process, adjust the fluidizing velocity according to the bed pressure difference and particle fluidization state to maintain the material temperature at 38–45℃ and avoid the formation of obvious wet clumps or equipment wall adhesion.

[0036] After the agglomeration liquid is sprayed, continue fluidized drying for 15–25 min until the particle moisture content is no higher than 8%. The resulting particles are passed through a 40-mesh sieve to remove large particles and through a 100-mesh sieve to remove fine powder. Collect the particles that pass through the 40-mesh sieve but are retained on the 100-mesh sieve to obtain D. 50 The porous core has a diameter of 220 μm. The porosity of the porous core was measured to be 42.6% ± 2.1%, and the liquid absorption per unit dry weight was 1.16 ± 0.08 g / g.

[0037] Step S6, Negative Pressure Internal Loading: Place the porous core obtained in Step S5 into a vacuum impregnation device equipped with a slow-turning mechanism. Add the remaining first umami loading liquid at a dry basis mass ratio of 3.1:1 (remaining first umami loading liquid to porous core). Slowly turn the core at 30 rpm to uniformly wet the porous core. Perform three pulsed negative pressure impregnation cycles: each time, evacuate to a gauge pressure of -0.07 MPa and maintain for 5 min, then restore to normal pressure and maintain for 5 min. After impregnation, stop turning, allow to stand and drain for 2 min, recover the residual liquid on the surface and include it in the material balance; then dry at 45 ℃ until the moisture content is no higher than 8%, so that the umami extract solids introduced by wet agglomeration and negative pressure impregnation together constitute the umami internal support phase.

[0038] Step S7, Fluidized Bed External Coating: The porous core containing the umami-containing internal carrier phase is placed in a fluidized bed apparatus, and the second umami coating liquid is sprayed onto the outer surface of the porous core. The inlet air temperature is 50-55 ℃, and the material temperature does not exceed 45 ℃; after spraying, continue fluidized drying until the product moisture content is 5.5%. The resulting product D 50The sample size was 252 μm, the water activity was 0.36, the free glutamic acid content was 12.7 mg / g, and the 5′-guanosine monophosphate content was 0.71 mg / g, all on a dry basis.

[0039] Example 2: The difference between this embodiment and Embodiment 1 is that, in step S1, the dry basis mass ratio of the umami extraction component to the carrier preparation component is 1:1; the cutting and mixing are carried out in an atmosphere with an oxygen volume content of 2.5%.

[0040] In step S2, the mass ratio of the umami extract component to water is 1:3; the electric field strength of the pulsed electric field treatment is 15 kV / cm, the pulse width is 10 μs, the number of pulses is 100, and the energy input per unit mass of mushroom liquid is 40 kJ / kg.

[0041] In step S3, the cell wall hydrolase hydrolysis was performed at a temperature of 40 °C for 0.5 h; the neutral protease hydrolysis was performed at a pH of 6.5, a temperature of 42 °C for 0.5 h; the flavor protease hydrolysis was performed at a pH of 6.0, a temperature of 40 °C for 0.5 h; and the nucleic acid hydrolase hydrolysis was performed at a pH of 5.0, a temperature of 40 °C for 0.5 h.

[0042] In step S4, the solid content of the first umami loading liquid is 8%, the solid content of the second umami coating liquid is 25%, and the mass ratio of the solid content of the umami extract contained in the two liquids is 1:0.5.

[0043] In step S5, the carrier preparation component is dried at 50 °C and then ultra-finely pulverized to D at a pulverization temperature of 25 °C. 50 20 μm; wet agglomeration forms D 50 It is a porous nucleus with a diameter of 180 μm.

[0044] In step S6, the ratio of the remaining first umami loading liquid to the dry basis of the porous core is 4.35:1, the gauge pressure under negative pressure is -0.06 MPa, and a pulsed wetting process of "vacuuming and restoring normal pressure" is adopted twice.

[0045] In step S7, the ratio of the total solid amount of umami extract in the first umami-supporting liquid and the second umami-coating liquid to the dry weight of the button mushroom fruiting body ultrafine powder is approximately 0.60:1. Based on material balance, the actual dry weight ratio of the solid amount of umami extract to the ultrafine powder in the final product is approximately 0.55:1, and the measured dry weight ratio of the umami inner supporting phase to the umami outer coating phase is approximately 1:0.52; the resulting composite seasoning granules D 50 The sample size was 185 μm, the moisture content was 5.7%, and the water activity was 0.35. All other steps and process parameters were the same as in Example 1.

[0046] Example 3: The difference between this embodiment and Embodiment 1 is that, in step S1, the dry basis mass ratio of the umami extraction component to the carrier preparation component is 3:1; the cutting and mixing are carried out in an atmosphere with an oxygen volume content of 2.5%.

[0047] In step S2, the mass ratio of the umami extract component to water is 1:8; the electric field strength of the pulsed electric field treatment is 25 kV / cm, the pulse width is 10 μs, the number of pulses is 100, and the energy input per unit mass of mushroom liquid is 80 kJ / kg.

[0048] In step S3, the cell wall hydrolase hydrolysis was performed at a temperature of 50 °C for 2 h; the neutral protease hydrolysis was performed at a pH of 7.2, a temperature of 52 °C for 2 h; the flavor protease hydrolysis was performed at a pH of 7.0, a temperature of 50 °C for 2 h; and the nucleic acid hydrolase hydrolysis was performed at a pH of 6.5, a temperature of 50 °C for 2 h.

[0049] In step S4, the solid content of the first umami loading liquid is 12%, the solid content of the second umami coating liquid is 35%, and the mass ratio of the solid content of the umami extract contained in the two liquids is 1:2.

[0050] In step S5, the carrier preparation component is dried at 50 °C and then ultra-finely pulverized to D at a pulverization temperature of 25 °C. 50 35 μm; wet agglomeration forms D 50 It is a porous nucleus with a diameter of 320 μm.

[0051] In step S6, the ratio of the remaining first umami loading liquid to the dry basis of the porous core is 4.60:1, the gauge pressure under negative pressure is -0.08 MPa, and a pulse-type impregnation process of "vacuuming and restoring normal pressure" is adopted four times.

[0052] In step S7, the ratio of the total solid amount of umami extract in the first umami-supporting liquid and the second umami-coating liquid to the dry basis mass of the button mushroom fruiting body ultrafine powder is approximately 2:1. Based on material balance, the actual dry basis mass ratio of the umami extract solids to the ultrafine powder in the final product is approximately 1.82:1, and the measured dry basis mass ratio of the umami inner support phase to the umami outer coating phase is approximately 1:1.94; the resulting composite seasoning granules D 50 The sample size was 386 μm, the moisture content was 5.8%, and the water activity was 0.37. All other steps and process parameters were the same as in Example 1.

[0053] Example 4: The difference between this embodiment and Embodiment 1 is that in step S2, the electric field strength of the pulse electric field treatment is 20 kV / cm, the pulse width is 1 μs, and the cumulative number of pulses is 200; the conductivity of the liquid is 0.50 mS / cm, the pulse frequency is 500Hz, the processing flow rate is 20 L / h, and the measured unit mass energy input of the equipment is 40 kJ / kg.

[0054] In step S3, only β-glucanase was used as the cell wall hydrolase; 5′-phosphodiesterase was used as the nucleic acid hydrolase; the order of addition of neutral protease and flavor protease, as well as the remaining enzymatic hydrolysis conditions, were the same as in Example 1. All other steps and process parameters were the same as in Example 1.

[0055] Example 5: The difference between this embodiment and Embodiment 1 is that in step S2, the electric field strength of the pulse electric field treatment is 20 kV / cm, the pulse width is 20 μs, and the cumulative number of pulses is 20; the conductivity of the liquid is 0.50 mS / cm, the pulse frequency is 50 Hz, the processing flow rate is 20 L / h, and the measured unit mass energy input of the equipment is 80 kJ / kg.

[0056] In step S3, only cellulase is used as the cell wall hydrolase; nuclease P1 is used as the nucleic acid hydrolase; the order of addition of neutral protease and flavor protease and the remaining enzymatic hydrolysis conditions are the same as in Example 1. The remaining steps and process parameters are the same as in Example 1.

[0057] Comparative Example 1: This comparative example was used to investigate the effect of pulsed electric field pretreatment on the extraction of umami components. The difference between Comparative Example 1 and Example 1 is that pulsed electric field treatment was not applied to the umami extract components; instead, the mushroom substrate was stirred for the same amount of time under low oxygen conditions. Subsequent staged enzymatic hydrolysis, solid-liquid separation, porous core preparation, negative pressure internal loading, and fluidized bed external coating were all the same as in Example 1.

[0058] Comparative Example 2: This comparative example was used to investigate the effect of direct mixing of umami extract powder and Agaricus bisporus fruiting body ultrafine powder on product performance. Umami extract was prepared according to steps S1 to S3 of Example 1, and then vacuum concentrated and vacuum dried at 45 °C and a gauge pressure of -0.08 MPa to obtain Agaricus bisporus umami extract powder. D was prepared according to the method of Example 1. 50 The product is an ultrafine powder of 25 μm Agaricus bisporus fruiting bodies. Following the same dry weight ratio of umami extract solids to ultrafine powder as in Example 1, the umami extract powder and ultrafine powder were dry-mixed, then wet-granulated with an appropriate amount of deionized water, and dried at 45 °C to ensure the final product moisture content did not exceed 5.5%. This comparative example did not involve negative pressure internal loading or fluidized bed external coating.

[0059] Comparative Example 3: This comparative example is used to examine the differences between a one-time fluidized bed spraying and the two-stage loading method of this invention. The umami extract and porous core were prepared according to the method of Example 1. All the umami extract was concentrated to a solids content of 25%, without distinguishing between the first umami loading liquid and the second umami coating liquid, and without negative pressure internal wetting. All the concentrated liquid was sprayed onto the outer surface of the porous core in a single application and dried under the fluidized bed conditions of Example 1 to obtain surface-sprayed Agaricus bisporus seasoning granules. The theoretical amount of umami extract solids added to the concentrated liquid remained consistent with that of Example 1.

[0060] Comparative Example 4: This comparative example was used to investigate the effect of a low-oxygen atmosphere. Except for steps S1 to S3 where nitrogen was not introduced and the oxygen volume content in the processing space was maintained at 20.9% ± 0.5% of the air level, the other pulsed electric field, enzymatic hydrolysis, concentration, agglomeration, negative pressure internal loading, and fluidized bed external coating conditions were the same as in Example 1.

[0061] Detection method: 1. Solid content of umami extract; Weigh approximately 10 g of the umami extract and place it in a pre-weighed weighing dish. Dry at 105℃ until constant weight. The solid content is calculated using the following formula: Solid content = mass of solids after drying ÷ mass of umami extract × 100%.

[0062] 2. Yield of soluble solids; The total mass and solids content of the umami extract were determined, and the total mass of soluble solids in the umami extract was calculated. The yield was calculated according to the following formula: Soluble solids yield = Dry weight of soluble solids in umami extract ÷ Dry weight of umami extract raw material × 100%.

[0063] 3. Amino acid nitrogen; The formaldehyde value was determined using a dual-indicator titration method.

[0064] Take an appropriate amount of umami extract and centrifuge at 8000×g for 10 min. Take the supernatant as the sample solution. Accurately transfer two equal volumes of sample solution, 10.00 mL each, into two 250 mL Erlenmeyer flasks, and add 50 mL of carbon dioxide-free water to each.

[0065] Add 2-3 drops of neutral red indicator solution to the first sample, and titrate with 0.1000 mol / L sodium hydroxide standard titration solution until the solution changes from red to amber or yellow-orange and the color remains unchanged for 30 s. Record the volume of sodium hydroxide standard titration solution consumed as V1.

[0066] Add 20.00 mL of neutral formaldehyde solution and 3 drops of thymolphthalein indicator to the second sample, shake well, let stand for 1 min, and then titrate with 0.1000 mol / L sodium hydroxide standard titration solution until the solution turns light blue and remains so for 30 s. Record the volume of sodium hydroxide standard titration solution consumed as V2.

[0067] Simultaneously perform a reagent blank test: Take 50 mL of carbon dioxide-free water, add 20.00 mL of neutral formaldehyde solution and 3 drops of thymolphthalein indicator solution, titrate to a light blue color using the same method, and record the volume of sodium hydroxide standard titration solution consumed as V0.

[0068] Each sample was measured independently three times, and the average value was taken as the result. When the umami extract was dark in color, a pH meter could be used to help determine the endpoint. The pH corresponding to the neutral red endpoint was approximately 7.0, and the pH corresponding to the thymolphthalein endpoint was approximately 9.2.

[0069] Based on the volume of sodium hydroxide standard solution consumed in the two titrations and the sample volume, the amino acid nitrogen content is calculated in g / 100 mL using the following formula: ; In the formula: X represents the amino acid nitrogen content in the umami extract, in g / 100 mL; V2 represents the volume of sodium hydroxide standard titration solution consumed after titration with neutral formaldehyde and thymolphthalein, in mL; V1 represents the volume of sodium hydroxide standard titration solution consumed with neutral red as an indicator, in mL; V0 represents the volume of sodium hydroxide standard titration solution consumed in the reagent blank, in mL; c represents the actual concentration of the sodium hydroxide standard titration solution, in mol / L; 0.014 is the nitrogen mass equivalent to 1.00 mL, 1 mol / L sodium hydroxide standard titration solution, in g; F is the dilution factor of the sample solution; V S The value indicates the volume of the umami extract used for the test; in this test, it was 10.00 mL.

[0070] The amino acid nitrogen content (g / 100 g solids) was subsequently calculated by comparing the amino acid nitrogen content of the umami extract with the mass of solids contained in 100 mL of the umami extract.

[0071] 4. Product load capacity and material balance; The following data were recorded: dry weight of the Agaricus bisporus fruiting body ultrafine powder, loss of ultrafine powder due to agglomeration and sieving, feed weight of umami extract solids in the first and second umami loading liquids, recovery of surface residual liquid, loss due to adhesion to equipment walls, and dry weight of the final product. The actual retained dry weight of ultrafine powder = dry weight of ultrafine powder feed - confirmed loss of ultrafine powder and agglomerates; the actual retained dry weight of umami extract solids in the final product = dry weight of the final product - actual retained dry weight of ultrafine powder. Cross-verification was performed using the recovery rates of glutamic acid and 5′-guanylic acid.

[0072] Loading weight = Mass of umami extract solids actually retained in the final product ÷ Dry weight of the final product × 100%.

[0073] 5. Load efficiency; Record the total mass of umami extract solids added to the first umami loading liquid and the second umami coating liquid. The sum of the two is the total mass of umami extract solids added. Although the surface residue recovered in step S6 can be recycled, it does not enter the final product of this batch. Therefore, it is included in the loss of umami extract in this batch and is not deducted from the total mass of umami extract solids added.

[0074] Load efficiency = (actual mass of umami extract solids retained in the final product) ÷ (total mass of umami extract solids fed into the feed) × 100%. Table 4 provides material balance data for representative batches.

[0075] Table 4 Material Balance of Representative Batches 6. Determination of glutamic acid and aspartic acid; In this invention, the contents of glutamic acid and aspartic acid were determined by pre-column derivatization high-performance liquid chromatography using phthalaldehyde. The analytes were free glutamic acid and free aspartic acid, and the sample was not acid-hydrolyzed. The umami extract was taken, and an equal volume of 10% sulfosalicylic acid solution was added. After mixing, the mixture was allowed to stand at 4 °C for 15 min, and then centrifuged at 12000 × g for 10 min. The supernatant was collected, and the pH was adjusted if necessary, diluted with ultrapure water, and then filtered through a 0.45 μm filter membrane. 50 μL of the sample solution was taken, and 100 μL of borate buffer and 50 μL of phthalaldehyde derivatizing reagent were added. After mixing, the mixture was reacted in the dark for 1 min, and the sample was injected within 2 min after derivatization. The chromatographic conditions were as follows: the column was a Nova-Pack RP-C18 column (150 mm × 3.9 mm, particle size 4 μm); the column temperature was 35 ℃; mobile phase A was a mixture of 10 mmol / L sodium phosphate buffer, methanol, and tetrahydrofuran at a volume ratio of 91:8:1, with the pH of the sodium phosphate buffer being 7.3; mobile phase B was a mixture of methanol and 10 mmol / L sodium phosphate buffer at a volume ratio of 80:20; gradient elution was used at a flow rate of 1.0 mL / min; the excitation wavelength of the fluorescence detector was 335 nm, and the emission wavelength was 440 nm; the injection volume was 20 μL.

[0076] The gradient elution program was as follows: 0–6 min, mobile phase A was 100%; at 17 min, mobile phase A decreased to 85%; at 25 min, it decreased to 80%; at 33 min, it decreased to 70%; at 45 min, it decreased to 60%; at 61 min, it decreased to 20%; at 67–70 min, it decreased to 0%; at 71 min, it was restored to 100% and equilibrated to 80 min. The retention times of glutamic acid and aspartic acid standards were used for qualitative analysis, and the content was calculated using the external standard method. The results were expressed as the mass of glutamic acid or aspartic acid per gram of umami extract solids.

[0077] 7.5′-Guanyu acid content determination; In this invention, the 5′-guanylic acid content was determined by high performance liquid chromatography. An appropriate amount of umami extract was taken, diluted with ultrapure water and brought to a final volume. After centrifugation, the supernatant was collected and filtered through a 0.45 μm aqueous filter membrane as the test solution.

[0078] The chromatographic conditions were as follows: a SunFire C18 column (250 mm × 4.6 mm, particle size 5 μm); a mobile phase consisting of an aqueous solution containing 10 mmol / L potassium dihydrogen phosphate and 5 mmol / L sodium heptanesulfonate, adjusted to pH 3.0 with phosphoric acid; isocratic elution at a flow rate of 0.8 mL / min; a photodiode array detector at a detection wavelength of 250 nm and a column temperature of 30℃; an injection volume of 10 μL; and disodium 5′-guanylate as the standard, with all results converted to 5′-guanylic acid. Qualitative analysis was performed using the retention time and UV absorption spectrum of the 5′-guanylic acid standard. The 5′-guanylic acid content was calculated using the external standard method, and the results were expressed as the mass of 5′-guanylic acid per gram of umami extract solids.

[0079] 8. Determination of the relative peak area of ​​peptides not exceeding 3 kDa; In this invention, the molecular weight distribution of peptides was determined using gel permeation high-performance liquid chromatography (GPC). The chromatographic column was a TSK gel G2000 SWXL column (300 mm × 7.8 mm); the mobile phase was a mixture of acetonitrile, water, and trifluoroacetic acid in a volume ratio of 45:55:0.1; the flow rate was 0.5 mL / min; the column temperature was 30 ℃; the UV detection wavelength was 220 nm; and the injection volume was 10 μL. A calibration curve was established using peptide standards with known molecular weights to correlate retention time with the logarithm of molecular weight. The relative peak area percentage (ratio) of peptides with a molecular weight not exceeding 3 kDa was calculated based on the proportion of peak area of ​​peptides with a molecular weight not exceeding 3 kDa to the total peak area of ​​all peptides.

[0080] 9. Particle size, moisture content, and water activity; D of Agaricus bisporus fruiting body ultrafine powder, porous granules and compound seasoning granules 50 The particle size distribution was determined using a laser particle size analyzer in dry dispersion mode at a dispersion pressure of 0.20 MPa. Each sample was measured in triplicate, and the porous cores and finished particles were verified by sieving. Moisture content was determined by drying at 105 °C to constant weight; water activity was determined using a water activity meter at 25 °C.

[0081] 10. Porosity and liquid absorption of porous cores; The true density of the porous core was determined using the gas displacement method, and the apparent particle density was determined using the known volume displacement method. Porosity = (1 - apparent particle density ÷ true density) × 100%. For liquid absorption measurement, 1.000 g of the porous core was weighed, soaked in sufficient deionized water for 10 min, passed through a 100-mesh sieve, and drained for 2 min before weighing. Liquid absorption per unit dry basis mass = (mass after absorption - mass before absorption) ÷ dry basis mass before absorption. Each group was measured independently three times.

[0082] 11. Solubility and dispersibility; Weigh 5.0 g of the sample and add it to 100 mL of deionized water at 60 °C. Stir at 200 rpm. Record the time required for the system to become uniformly dispersed without any obvious floating clumps after the sample is added; this time is taken as the dispersion time. After stopping stirring, let it stand for 10 min, collect the bottom sediment, and dry it to constant weight. The sedimentation rate is calculated using the following formula: Sedimentation rate = dry weight of sediment ÷ dry weight of sample × 100%.

[0083] Because the product contains insoluble mushroom dietary fiber, the evaluation criteria were "dispersion time" and "sedimentation rate," and complete dissolution was not used as the evaluation standard.

[0084] 12. Ratio of umami-containing inner phase to umami-coating outer phase; Accurately weigh 0.5 g of whole button mushroom umami compound seasoning particles, add 10.0 mL of 10 mmol / L phosphate buffer (pH 6.8) pre-cooled at 4 ℃, and gently elute by inverting at 60 rpm for 30 s. Immediately after elution, centrifuge at 4 ℃ and 5000×g for 1 min, and use the supernatant as the umami outer coating phase assay solution. Thoroughly grind and break up the particles after surface elution, add 20.0 mL of ultrapure water, and sonicate at 40 ℃ for 30 min. Centrifuge and use the supernatant as the umami inner carrier phase assay solution.

[0085] The contents of glutamic acid and 5′-guanylic acid in the umami outer phase and umami inner phase assay solutions were determined using the methods described above. Simultaneously, the unit content of glutamic acid and 5′-guanylic acid in the solids of the first umami loading solution and the second umami coating solution was determined. Based on the total mass of the markers in each assay solution and the unit content of the markers in the corresponding loading or coating solution solids, the solid mass of the umami inner phase and the umami outer phase were calculated, and their dry basis mass ratio was determined.

[0086] Surface elution conditions were pre-validated using particles containing only the umami inner phase and particles containing only the umami outer phase. Particles containing only the umami inner phase were prepared according to the porous core preparation and pulsed negative pressure wetting method described in the examples. Specifically, a first umami loading liquid was used for wet agglomeration and internal wetting, and after the first stage of low-temperature drying, fluidized bed spraying of the second umami coating liquid was not performed. Particles containing only the umami outer phase used ultrafine powder of *Agaricus bisporus* fruiting bodies to prepare porous cores. During wet agglomeration, an equal mass of deionized water was used instead of the first umami loading liquid, and pulsed negative pressure wetting was not performed. After pre-drying, the second umami coating liquid was sprayed onto the outer surface of the porous core at the same spray solids ratio as the test compound seasoning particles and then dried. Blank porous cores were wet-agglomerated using deionized water, without wetting with the first umami loading liquid or spraying with the second umami coating liquid. At least three independent batches of each of the above three types of validation samples were prepared.

[0087] During the preparation of validation samples, the mass concentrations of glutamic acid and 5′-guanylic acid in the first umami loading solution and the second umami coating solution were measured, respectively. The theoretical loading of markers from the umami inner phase particles alone was calculated by subtracting the measured mass of the corresponding markers in the recovery residue, equipment wall deposits, and equipment washing solution from the total mass of markers added to the first umami loading solution. Similarly, the theoretical loading of markers from the umami outer coating particles alone was calculated by subtracting the measured mass of the corresponding markers in the corresponding spraying residue, equipment wall deposits, and equipment washing solution from the total mass of markers added to the second umami coating solution. Blank porous nuclei were analyzed using the same method, and the corresponding background values ​​were subtracted from the marker measurement results of each eluent or extract.

[0088] For particles containing only the umami-containing outer coating phase, surface elution was performed. The surface elution recovery rate of the outer coating phase was calculated as "the mass of the marker in the surface eluent ÷ the theoretical loading of the marker × 100%". For particles containing only the umami-containing inner carrier phase, the same surface elution was performed, and the release rate of the inner carrier phase was calculated as "the mass of the marker in the surface eluent ÷ the theoretical loading of the marker × 100%". The elution time, buffer volume, and inversion speed were adjusted to ensure that the surface elution recovery rates of both glutamic acid and 5′-guanosine monophosphate outer coating phases were not less than 85%, and the release rate of the inner carrier phase during the surface elution process was not higher than 10%.

[0089] After surface elution, the remaining particles are thoroughly ground and internally extracted. The total recovery rate of fractionated extraction is calculated as "the sum of the mass of the marker in the surface eluent and the internal extract ÷ the theoretical loading of the marker in the corresponding validation sample × 100%". The total recovery rate of fractionated extraction for glutamic acid and 5′-guanylic acid should be 90%–110%. The relative deviation of the mass ratio of the umami inner phase to the umami outer phase obtained by the glutamic acid marker method and the 5′-guanylic acid marker method should be calculated by dividing the absolute value of the difference between the two determinations by the average value of the two determinations, and should not exceed 15%. The same sample should be measured in parallel at least 3 times, and the relative standard deviation of the repeated determinations should not exceed 8%. If the above limits are exceeded, the surface elution time, buffer volume, inversion speed, or internal extraction conditions should be re-determined or validated. The average mass ratio of the inner and outer phases obtained by the glutamic acid marker method and the 5′-guanylic acid marker method is used as the detection result of the ratio of the umami inner phase to the umami outer phase, and is listed in Table 5.

[0090] Table 5. Validation Results of the Internal and External Phase Grading Detection Methodology 13. Determination of the content of free glutamic acid and 5′-guanylic acid in the finished product; Weigh the finished granules, grind them thoroughly, add ultrapure water at a ratio of 10 mL extract / g dry basis, extract by ultrasonication at 40 ℃ for 30 min, centrifuge at 12000×g for 10 min, and collect the supernatant; determine the free glutamic acid and 5′-guanosine monophosphate according to the methods in items 6 and 7 above, and express the results in mg / g dry basis.

[0091] 14. Sensory evaluation test of umami flavor in umami compound seasoning granules; The evaluation was conducted by a panel of 20 healthy, sensory-sharp, and trained food and related professionals (half male and half female). Three batches of each example and comparative example were prepared independently, and each independent batch underwent two sensory evaluations, with an interval of at least 24 hours between evaluations. Each evaluation was performed by the aforementioned 20 evaluators. 0.5 g of sample was weighed, added to drinking water and diluted to 25.00 mL, and sodium chloride was added to achieve a final mass-volume concentration of 0.30 g / 100 mL in the dispersion. After thorough dispersion, the sample was dispensed into capped sensory evaluation cups with random three-digit codes. Evaluators were unaware of the corresponding example or comparative example for each sample; the order of sample presentation was randomly determined, and unblinding was performed after all sensory evaluations and original scoring records were completed. The temperature of the samples was controlled at 55±2 ℃ when submitted to the evaluators, and evaluation began within 2 minutes of dispensing. Evaluators rinsed their mouths with warm water between adjacent samples, with an interval of at least 1 minute before evaluating the next sample.

[0092] The overall umami quality is evaluated using a 9-point scale, where 1 point indicates virtually no umami, 3 points indicates weak umami, 5 points indicates moderate umami, 7 points indicates a distinct umami flavor that is well-balanced with the mushroom flavor, and 9 points indicates a strong umami flavor that is well-balanced and lasts for a long time.

[0093] For each independently prepared batch, the average of two evaluations by each evaluator was first calculated, followed by the average of the scores from all 20 evaluators. The final result was expressed as the mean ± standard deviation of the average scores from the three independently prepared batches. Within-group correlation coefficients were used to assess the consistency among evaluators; a correlation coefficient of at least 0.75 was considered acceptable for the evaluation team. Repeated measures ANOVA was used to compare scores from different samples, and Tukey's method was used to analyze differences between groups. A p-value < 0.05 was considered statistically significant.

[0094] Table 6. Results of umami extraction and flavor component testing in different embodiments and comparative examples. As shown in Table 6, the soluble solids yield of Example 1 under the intermediate parameter combination was 42.6%, the amino acid nitrogen was 1.62 g / 100 g solids, and the contents of glutamic acid, aspartic acid and 5′-GMP were 25.4 mg / g solids, 10.8 mg / g solids and 1.42 mg / g solids, respectively. The relative peak area ratio of molecules not higher than 3 kDa was 58.4%, indicating that hypoxia pulsed electric field treatment and continuous staged enzymatic hydrolysis can jointly obtain a umami extract containing flavor amino acids, peptides with a molecular weight not higher than 3 kDa and flavor nucleotides.

[0095] Compared to Comparative Example 1 without pulsed electric field treatment, Example 1 showed improvements in all umami extraction indicators, indicating that under the same low-oxygen atmosphere and subsequent enzymatic hydrolysis conditions, pulsed electric field pretreatment is beneficial for promoting the release of soluble components. Compared to Comparative Example 4 under normoxic pulsed electric field treatment, Example 1 showed increases in soluble solids yield, amino acid nitrogen, glutamic acid, aspartic acid, 5′-GMP, and the relative peak area ratio of peptides not exceeding 3 kDa, respectively, by approximately 9.5%, 13.3%, 11.4%, 12.5%, 13.6%, and 8.8%.

[0096] Examples 2 and 3 employed low-value and high-value combinations of the main parameters, respectively. While the extraction indices of Example 2 were lower than those of Example 1, it still yielded a soluble solids yield of 34.8% and an umami extract containing glutamic acid, aspartic acid, 5′-GMP, and small peptides, indicating the feasibility of the defined low-value combination. Example 3 showed higher levels of soluble solids yield, amino acid nitrogen, glutamic acid, 5′-GMP, and the proportion of peptides not exceeding 3 kDa compared to other examples, indicating that increasing the pulsed electric field energy input and extending the staged enzymatic hydrolysis time could further promote the release of umami components; however, the processing time and energy consumption also increased accordingly.

[0097] Example 4 used β-glucanase and 5′-phosphodiesterase, achieving a 5′-GMP content of 1.50 mg / g solids. This demonstrates that even with β-glucanase and 5′-phosphodiesterase combined with appropriate pulsed electric field parameters, a umami extract containing 5′-GMP can still be obtained, indicating the feasibility of this combination. Example 5 used cellulase and nuclease P1, combined with appropriate pulsed electric field parameters, also successfully obtaining a umami extract containing glutamic acid, aspartic acid, 5′-GMP, and small peptides, demonstrating the feasibility of the corresponding enzyme combinations used in Examples 4 and 5.

[0098] The umami extraction stages of Comparative Examples 2 and 3 were essentially the same as those of Example 1; therefore, the indicators listed in Table 6 are generally similar to those of Example 1. This result indicates that the differences in particle performance between Example 1 and Comparative Examples 2 and 3 in Table 7 mainly stem from the subsequent negative pressure internal loading and fluidized bed external coating methods, rather than from significant differences in the composition of the initial umami extract. In summary, the results in Table 6 demonstrate that Example 1 achieves a good overall balance between umami component release effect, processing time, and energy consumption.

[0099] Table 7 Performance test results of compound flavoring granules in different embodiments and comparative examples Note: The values ​​marked with * in Comparative Example 2 represent the solids retention rate, not the loading efficiency of the umami extract into the carrier pores.

[0100] Table 8 Results of porous core structure and marker content in finished product As shown in Table 7, the loading amount of Example 1 was 40.2%, the loading efficiency was 92.6%, and the dry basis mass ratio of the umami inner phase to the umami outer phase was 1:0.98. The measured D... 50 The particle size was 252 μm, the moisture content was 5.4%, and the water activity was 0.36; the dispersion time was 39 s, the sedimentation rate was 6.5%, and the umami score was 8.4. Referring to Table 8, the porous core of Example 1 had a porosity of 42.6%, a liquid absorption per unit dry basis of 1.16 g / g, and the finished product contained 12.7 mg / g free glutamic acid and 0.71 mg / g dry basis, respectively. This indicates that the combination of intermediate parameters achieved a good overall balance between loading, dispersion, and sensory quality.

[0101] Example 2 used a relatively low amount of umami extract, a low proportion of the coating phase, and a small porous core particle size, resulting in a loading of 35.4%, lower than Example 1. However, the dispersion time was shortened to 32 s and the sedimentation rate decreased to 5.6%, indicating that under the experimental conditions, reducing the proportion of the coating phase and decreasing the particle size is beneficial to improving the dispersion rate of the particles. Example 3 used a higher amount of umami extract and a higher proportion of the coating phase, achieving a loading of 64.5%, but D 50 With the particle size increased to 386 μm, the dispersion time and sedimentation rate increased to 53 s and 8.2%, respectively. These results indicate that increasing the loading has a certain impact on the umami flavor of the product, but excessively high particle size and surface coating will correspondingly reduce the dispersion speed. Therefore, the parameter combination in Example 1 exhibits a better overall balance between loading, dispersibility, sedimentation rate, and umami score.

[0102] The loading efficiency of Examples 4 and 5 is higher than 91%, the ratio of inner phase to outer phase is close to 1:1, and the particle performance is similar to that of Example 1, indicating that the back-end biphase reconstruction process still has good applicability when the front-end pulse parameters and enzyme combination change.

[0103] Comparative Example 2 involved direct mixing and granulation of umami extract powder and ultrafine powder. Although the solids retention rate was 86.3%, no distinguishable internal and external phases were formed. The dispersion time was 73 s, and the sedimentation rate was 15.4%, which were approximately 87.2% longer and 136.9% higher than those in Example 1, respectively. The umami score also decreased to 7.1. This indicates that simple mixing of umami extract and powder may lead to uneven distribution of umami substances and particle agglomeration, failing to achieve the dispersion and sensory effects brought about by the two-phase spatial distribution of this invention.

[0104] Comparative Example 3 underwent a single surface coating process, with a 0:1 ratio of umami inner carrier phase to umami outer coating phase, a loading efficiency of 83.2%, a dispersion time of 59 s, and a sedimentation rate of 10.8%. Compared to Comparative Example 3, Example 1 showed a 9.4 percentage point increase in loading efficiency, a reduction of approximately 33.9% in dispersion time, and a decrease in sedimentation rate of approximately 39.8%. This indicates that firstly, negative pressure internal wetting with a low-solids first umami loading liquid followed by external coating with a high-solids second umami coating liquid can reduce excessive surface enrichment and material loss during the drying process, and allow for a more reasonable spatial distribution of umami components between the internal pores and the outer surface of the particles.

[0105] Comparative Example 1, lacking pulsed electric field pretreatment, exhibited lower umami extraction and particle performance indicators compared to Example 1. In contrast, Comparative Example 4, under normal oxygen conditions while retaining pulsed electric field treatment, still showed lower soluble solids yield, flavor component content, loading efficiency, and umami score compared to Example 1, illustrating that a low-oxygen atmosphere may be beneficial in reducing oxidation losses during processing.

[0106] While the present invention has been disclosed above, its scope of protection is not limited thereto. Those skilled in the art can make various changes and modifications without departing from the spirit and scope of this disclosure, and all such changes and modifications will fall within the scope of protection of this invention.

Claims

1. A method for preparing homologous two-phase reconstructed Agaricus bisporus umami compound seasoning granules, characterized in that, Includes the following steps: Step S1: Clean, cut and mix the same batch of button mushroom fruiting bodies, and divide the mixed button mushroom fruiting bodies into umami extract components and carrier preparation components according to dry weight. Step S2: Mix the dry weight of the umami extract with the weight of water at a ratio of 1:3 to 1:8 to form a mushroom broth. Before pulsed electric field treatment, introduce nitrogen gas into the closed treatment system connected to the mushroom broth for displacement until the oxygen concentration sensor in the gas phase space of the closed treatment system detects that the oxygen volume content is stable at 2% to 5%, and maintain the oxygen volume content during the pulsed electric field treatment. Perform pulsed electric field treatment on the mushroom broth so that the energy input per unit mass of mushroom broth is 40 to 80 kJ / kg, and control the temperature of the mushroom broth after treatment to not exceed 35 ℃. Step S3: In the same enzymatic hydrolysis container, without solid-liquid separation and enzyme inactivation between adjacent enzymatic hydrolysis stages, the mushroom liquid treated with pulsed electric field is sequentially subjected to cell wall hydrolysis, neutral protease hydrolysis, flavor protease hydrolysis and nucleic acid hydrolysis. After all enzymatic hydrolysis is completed, enzyme inactivation and solid-liquid separation are performed to obtain a umami extract containing flavor amino acids, peptides with a molecular weight not higher than 3 kDa and 5′-flavor nucleotides. Step S4: Under the condition that the temperature is not higher than 45 ℃, the umami extract is concentrated under reduced pressure to a solid content of 8% to 12%, a portion of which is taken as the first umami loading liquid, and the remaining portion is further concentrated under reduced pressure to a solid content of 25% to 35% as the second umami coating liquid. Step S5: The carrier preparation components are subjected to low-temperature drying and ultra-fine pulverization to obtain Agaricus bisporus fruiting body ultra-fine powder. 5% to 15% of the total mass of the first umami loading liquid is used as the agglomeration liquid for wet agglomeration, and pre-dried to a moisture content of no more than 8% to form a porous core with open internal channels. The porosity of the porous core is 30% to 50%, and the liquid absorption per unit dry mass is 0.8 to 1.4 g / g. Step S6: Add the remaining first umami loading liquid to the porous core at a ratio of (3-5) : 1 (dry basis mass of the remaining first umami loading liquid to the porous core). Perform 2-4 pulsed impregnation cycles of "vacuuming-restoring normal pressure" under a gauge pressure of -0.08 to -0.06 MPa to allow the remaining first umami loading liquid to enter the internal channels of the porous core. After impregnation, remove and recover the unabsorbed flowable liquid on the surface of the porous core. Then, perform a first-stage low-temperature drying to allow the umami extract solids introduced during the wet agglomeration process in step S5 and the umami extract solids that entered the internal channels during the pulsed negative pressure impregnation process to form an internal umami carrying phase. Step S7: Place the porous core containing the umami inner carrier phase in a fluidized state, and spray the second umami coating liquid onto the outer surface of the porous core. Perform a second stage of low-temperature drying under the condition that the material temperature is not higher than 50 ℃ to form an umami outer coating phase, and obtain button mushroom umami compound seasoning particles including the porous core, the umami inner carrier phase distributed in the internal channels of the porous core, and the umami outer coating phase distributed on the outer surface of the porous core.

2. The preparation method according to claim 1, characterized in that, The dry basis mass ratio of the umami extract component to the carrier preparation component is (1-3):1; steps S1 to S3 are all carried out in a closed processing space, and the oxygen volume content in the closed processing space is maintained at 2.5% by means of an oxygen concentration sensor and nitrogen flow rate adjustment.

3. The preparation method according to claim 1, characterized in that, The pulsed electric field treatment uses a bipolar square wave with an electric field strength of 15–25 kV / cm and a pulse width of 1–20 μs. Each material unit receives a total of 20–200 pulses. Based on the measured conductivity of the mushroom slurry, the pulse frequency and slurry flow rate are adjusted to ensure that the combination of electric field strength, pulse width, and cumulative pulse count satisfies the requirement that the energy input per unit mass of mushroom slurry is 40–80 kJ / kg, and that the temperature of the mushroom slurry after pulsed electric field treatment does not exceed 35 °C.

4. The preparation method according to claim 1, characterized in that, The cell wall hydrolytic enzyme includes at least one of β-glucanase and cellulase, with β-glucanase added at 120 U per gram of umami extract raw material (dry basis) and cellulase added at 60 U per gram of umami extract raw material (dry basis); the neutral protease added at 1000 U per gram of umami extract raw material (dry basis) and the flavor protease added at 500 U per gram of umami extract raw material (dry basis); the nucleic acid hydrolytic enzyme includes at least one of nuclease P1 and 5′-phosphodiesterase, with nuclease P1 added at 80 U per gram of umami extract raw material (dry basis) and 5′-phosphodiesterase added at 80 U per gram of umami extract raw material (dry basis).

5. The preparation method according to claim 4, characterized in that, The pH, temperature, and time for each enzymatic hydrolysis stage are selected from any of the following complete parameter combinations: The first parameter combination is as follows: cell wall hydrolase hydrolysis at pH 5.5, temperature 40 ℃, and time 0.5 h; neutral protease hydrolysis at pH 6.5, temperature 42 ℃, and time 0.5 h; flavor protease hydrolysis at pH 6.0, temperature 40 ℃, and time 0.5 h; and nucleic acid hydrolase hydrolysis at pH 5.0, temperature 40 ℃, and time 0.5 h. The second parameter combination is as follows: cell wall hydrolase hydrolysis at pH 5.5, temperature 45 ℃, and time 1 h; neutral protease hydrolysis at pH 6.8, temperature 48 ℃, and time 1 h; flavor protease hydrolysis at pH 6.2, temperature 45 ℃, and time 1 h; and nucleic acid hydrolase hydrolysis at pH 5.5, temperature 45 ℃, and time 0.75 h. The third parameter combination is as follows: the enzymatic hydrolysis of cell wall hydrolysates is performed at pH 5.5, temperature 50 ℃, and time 2 h; the enzymatic hydrolysis of neutral protease is performed at pH 7.2, temperature 52 ℃, and time 2 h; the enzymatic hydrolysis of flavor protease is performed at pH 7.0, temperature 50 ℃, and time 2 h; and the enzymatic hydrolysis of nucleic acid hydrolysates is performed at pH 6.5, temperature 50 ℃, and time 2 h.

6. The preparation method according to claim 1, characterized in that, The D of the mushroom fruiting body ultrafine powder 50 The diameter is 20–35 μm; the D of the porous core is... 50 The size ranges from 180 to 320 μm.

7. The preparation method according to claim 1, characterized in that, The mass ratio of the solid umami extract in the first umami loading liquid to that in the second umami coating liquid is 1:(0.5~2), wherein the first umami loading liquid used for wet agglomeration in step S5 accounts for 5%~15% of the total mass of the first umami loading liquid.

8. The preparation method according to claim 1 or 7, characterized in that, In step S6, vacuum is maintained for 3-8 minutes each time, and normal pressure is restored and maintained for 3-8 minutes. During the soaking process, the mixture is slowly turned over at 20-40 rpm. After soaking, the excess liquid on the surface is removed and the mixture is dried at a temperature not higher than 45 ℃ until the moisture content is not higher than 8%. In step S7, the moisture content of the mushroom umami compound seasoning granules obtained is not higher than 6%.

9. A homologous dual-phase reconstructed Agaricus bisporus umami compound seasoning granule, characterized in that, The compound seasoning particles comprise a porous core formed by the agglomeration of ultrafine powder from *Agaricus bisporus* fruiting bodies, an internal umami-carrying phase distributed within the pores of the porous core, and an external umami-coating phase distributed on the outer surface of the porous core. Both the internal and external umami-carrying phases are formed from *Agaricus bisporus* umami extract. The dry-basis mass ratio of the *Agaricus bisporus* umami extract to the ultrafine powder constituting the porous core is (0.5–2):1, and the dry-basis mass ratio of the internal umami-carrying phase to the external umami-coating phase is 1:(0.5–2). 50 The particle size is 180–400 μm, and the content of free glutamic acid is not less than 8.0 mg / g and the content of 5′-guanylic acid is not less than 0.40 mg / g, based on the dry basis of the product.

10. The button mushroom umami compound seasoning granules according to claim 9, characterized in that, The moisture content of the button mushroom umami compound seasoning granules is not higher than 6%, and the water activity is not higher than 0.40.

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