Horse hoof red pepper sauce and production process thereof

Abstract

This invention discloses a water chestnut and red pepper sauce and its production process, relating to the food processing field. The process includes non-thermal modification of water chestnut pulp using a two-stage pulsed electric field treatment, preserving its texture while performing diverted enzymatic hydrolysis. After hydrolysis, the enzymes are not thermally inactivated, and residual enzymes decay naturally with fermentation conditions. Chitosan microcapsules with differentiated cross-linking densities are used to encapsulate microbial strains, and fermentation acid production triggers a cascade release, achieving a self-controlled connection between two stages of temperature-controlled fermentation. Polyphenols are extracted from water chestnut skin using subcritical extraction, and combined with vitamin E to construct a biphasic antioxidant barrier. The enzymatic hydrolysis residue undergoes a Maillard reaction in an independent reactor, and the flavor liquid is added back after sterilization to retain volatile aromas. This invention constructs a self-catalytic cascade-triggered closed loop, solving the problems of softening texture and monotonous fermentation flavor in water chestnuts during thermal processing, and achieving low-salt preservation and full utilization of raw material skin and pulp through grading.

Description

Technical Field

[0001] This invention relates to the field of food processing, and in particular to a water chestnut and red pepper sauce and its production process. Background Technology

[0002] Water chestnut and red pepper sauce is a type of compound seasoning sauce, typically made from water chestnuts and peppers as the main ingredients, through processes such as mixing, blending, and sterilization. In existing processing technologies, water chestnut pulp is generally steamed or blanched at high temperatures before being pulped or chopped for use, while peppers are mostly added directly or simply simmered. Fermentation processes have been introduced into some products to improve flavor.

[0003] However, the existing processing methods have the following problems in actual production.

[0004] Regarding texture preservation, traditional heat processing destroys the crispness of water chestnut flesh, while the addition of raw materials hinders flavor penetration. It is necessary to increase flesh permeability while reducing heat treatment to retain chewiness. Regarding fermentation control, simultaneous fermentation of multiple microbial strains can easily lead to uneven growth, while step-by-step inoculation increases operational complexity and the risk of contamination. It is necessary to achieve sequential release of microbial strains without human intervention. Regarding flavor development, fermentation and Maillard reactions interfere with each other, and sterilization easily leads to flavor loss. It is necessary to avoid interference from different flavor-enhancing pathways and effectively preserve flavor. Regarding antioxidant preservation, products are prone to oxidation and deterioration; a single antioxidant is difficult to provide uniform protection, and residual oxygen in packaging also affects quality. It is necessary to construct an antioxidant system covering the water and oil phases and combine it with oxygen-barrier packaging. Regarding raw material utilization, water chestnut peel contains active ingredients but is often discarded. It is necessary to grade and utilize the flesh and peel to introduce the effective components into the product. Summary of the Invention

[0005] In view of the shortcomings of the prior art, the purpose of this invention is to provide a technical solution to address one or more problems in the prior art.

[0006] To achieve the above objectives, the technical solution of the present invention is as follows: A production process for water chestnut and red pepper sauce includes the following steps: Peel the water chestnuts, collect the pulp and water chestnut peel separately, and cut the pulp into small pieces; The pulp particles were immersed in a conductive medium and subjected to a two-stage pulsed electric field treatment. The electric field strength of the first stage pulsed electric field treatment was 15~20kV / cm, and the electric field strength of the second stage pulsed electric field treatment was 25~35kV / cm. The material temperature was ≤40℃ throughout the pulsed electric field treatment. The material treated by the pulsed electric field is divided into two parts. 70-80% by mass enters the enzymatic hydrolysis process, and 20-30% is retained as fruit pulp particles and temporarily stored at 0-4℃. Cellulase and pectinase are added to the material entering the enzymatic hydrolysis process, and enzymatic hydrolysis is carried out under the conditions of pH 4.5~5.0 and temperature 45~50℃. No heat inactivation is performed after the enzymatic hydrolysis is completed. The enzymatic hydrolysis products are subjected to solid-liquid separation. The resulting liquid phase is used as the enzymatic hydrolysate and enters the fermentation process, while the resulting solid residue is entered into the Maillard reaction process. Using chitosan as the wall material and sodium tripolyphosphate as the crosslinking agent, type A microcapsules and type B microcapsules were prepared by ionic crosslinking. The sodium tripolyphosphate aqueous solution used to prepare type A microcapsules had a sodium tripolyphosphate mass fraction of 0.5-1.0%, and the core material included Leuconostoc mesenteroides and flavonoids from water chestnuts. The sodium tripolyphosphate aqueous solution used to prepare type B microcapsules had a sodium tripolyphosphate mass fraction of 2.0-3.0%, and the core material included Lactobacillus plantarum and flavonoids from water chestnuts. The enzymatic hydrolysate was mixed with chopped red pepper, edible salt, type A microcapsules and type B microcapsules to construct a fermentation substrate. Two-stage fermentation was carried out. The first stage fermentation temperature was 22~25℃. When the pH of the fermentation substrate was ≤5.0, the fermentation was transferred to the second stage. The second stage fermentation temperature was 30~33℃. Fermentation was stopped when the pH of the fermentation substrate dropped to 3.8~4.2 to obtain fermented mash. Water chestnut peel was extracted with subcritical water at a temperature of 120-150℃ and a gauge pressure of 0.8-1.5MPa. The resulting extract was filtered and concentrated to obtain a concentrated water chestnut peel polyphenol solution. L-cysteine ​​and D-xylose were added to the solid residue in an independent reaction vessel, and the Maillard reaction was carried out in two stages. The temperature of the first stage was 80~90℃, and the temperature of the second stage was 105~115℃. The reaction product was cooled to ≤40℃ and then extracted with water to obtain Maillard flavor extract. The fermented mash is mixed with temporarily refrigerated fruit pulp particles, concentrated water chestnut peel polyphenols, and vitamin E. The prepared materials are pasteurized at a temperature of 55-58℃ for 15-20 minutes. After sterilization, Maillard flavor extract is added, and the product is filled. Before sealing, the headspace of the packaging container is purged with nitrogen via a pulse.

[0007] The fruit pulp is cut into particles with a diameter of 5-10 mm, and the water chestnut skin is frozen and stored at ≤-18℃.

[0008] The conductivity of the conductive medium is 0.3~1.5 mS / cm, the volume ratio of fruit pulp particles to conductive medium is 1:2~1:3, the pulse width of the first pulse electric field treatment is 20~30μs, the pulse frequency is 50~100Hz, and the residence time is 20~60s, and the pulse width of the second pulse electric field treatment is 30~50μs, the pulse frequency is 100~200Hz, and the residence time is 20~60s.

[0009] Based on the dry weight of the pulp, the amount of cellulase added is 0.05~0.15 parts by weight, the amount of pectinase added is 0.02~0.08 parts by weight, the enzymatic hydrolysis time is 2~4 hours, and the solid-liquid separation is carried out by centrifugation or pressure filtration, with a centrifugation speed of 4000~6000 rpm and a centrifugation time of 10~15 minutes, and a pressure filtration pressure of 0.2~0.3 MPa.

[0010] The chitosan is dissolved in an aqueous acetic acid solution, and the chitosan mass-volume concentration is 15-20 g / L. The volume ratio of *Leuconostoc mesenteriae* activated bacterial solution to *Carex esculenta* flavonoid concentrate in the core material of the type A microcapsules is 10:1. The volume ratio of *Lactobacillus plantarum* activated bacterial solution to *Carex esculenta* flavonoid concentrate in the core material of the type B microcapsules is 10:1. The curing time for preparing type A microcapsules is 20-30 min, and the curing time for preparing type B microcapsules is 40-60 min.

[0011] The mass ratio of the enzymatic hydrolysate to the chopped red pepper is 1.5:1 to 2.5:1. Based on 100 parts by mass of the total fermentation substrate, the amount of added edible salt is 1.5 to 4.0 parts by mass, the amount of added type A microcapsules is 0.3 to 0.8 parts by mass, the amount of added type B microcapsules is 0.3 to 0.8 parts by mass, the initial pH of the fermentation substrate is 5.5 to 6.5, the fermentation time of the first stage is 36 to 72 hours, and the fermentation time of the second stage is 36 to 72 hours.

[0012] In the subcritical water extraction, the mass ratio of water chestnut peel to water is 1:10 to 1:20, the extraction time is 20 to 40 minutes, and the filtration uses a microporous membrane with a pore size of 0.45 μm.

[0013] Based on the dry weight of the solid residue, the amount of L-cysteine ​​added is 0.02~0.08 parts by mass, the amount of D-xylose added is 0.05~0.20 parts by mass, the pH of the Maillard reaction is 6.0~7.0, the reaction time of the first stage is 30~45 min, the reaction time of the second stage is 15~30 min, the amount of water added during water extraction is 3~5 times the mass of the reaction product, the water extraction temperature is 50~60℃, and the water extraction time is 30 min. Based on a total mass of 100 parts by mass of the blended sauce, the amount of vitamin E added is 0.02~0.06 parts by mass, and the amount of Maillard flavor extract added is 1~3 parts by mass.

[0014] The nitrogen pulse replacement is performed 3 times. Each time, nitrogen is injected until the headspace pressure is 0.15~0.20MPa and held for 5 seconds before being vented. The container is sealed immediately after the third nitrogen injection. The residual oxygen content in the headspace of the packaging container after replacement is ≤0.5%.

[0015] A water chestnut and red pepper sauce, produced using the aforementioned water chestnut and red pepper sauce production process, wherein the water chestnut and red pepper sauce has a pH of 3.8~4.2, a salt content of ≤4%, and a headspace residual oxygen content of ≤0.5% in the packaging container.

[0016] Compared with the prior art, the beneficial technical effects of the present invention are as follows: (i) After being treated with a pulsed electric field and enzymatically hydrolyzed, the water chestnut pulp is inactivated without heat. Type A and Type B chitosan microcapsules are used to release Leuconostoc mesenteroides and Lactobacillus plantarum in sequence at different pH levels, so as to achieve the sequential connection between ester production and flavor enhancement and acid production. The process is driven by the pH of the system itself.

[0017] (ii) Polyphenols are extracted from water chestnut skin using subcritical water. The solid residue is enzymatically hydrolyzed and then subjected to a two-stage Maillard reaction to produce a flavor extract. Vitamin E and polyphenols are added during formulation to form a biphasic antioxidant system. Independent reactions avoid interference with fermentation, and segmented temperature control regulates flavor and color. Flavor extract is added back after sterilization to minimize losses.

[0018] (iii) Material diversion processing: part of it is enzymatically fermented into a sauce, and the rest is refrigerated and added as crispy granules to provide texture. Mild sterilization and nitrogen replacement are used to reduce texture damage and flavor loss, and reduce the risk of oxidation during storage.

[0019] (iv) Processing of pulp and peel separately. The pulp is made into sauce and crunchy granules; polyphenols are extracted from the peel as antioxidants, and its flavonoid components are encapsulated in microcapsules to protect the microbial strains, thus realizing the graded utilization of the pulp and peel. Detailed Implementation

[0020] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and exemplary descriptions. It should be understood that the structures, proportions, sizes, etc., illustrated in the accompanying drawings are merely for illustrative purposes and to enable those skilled in the art to understand and read the invention. They are not intended to limit the implementation conditions of the invention. Any modifications to the structure, changes in proportions, or adjustments to the size, without affecting the effects and objectives achieved by the invention, should still fall within the scope of the technical content disclosed in this invention.

[0021] Comprehensive explanation This invention integrates three core technologies—pulsed electric field treatment, microencapsulated cascade fermentation, and post-addition of flavor liquid—into a single process, providing a systematic solution to the bottlenecks in the processing of water chestnut and red pepper sauce. Specifically: water chestnuts contain pectin and cellulose; traditional blanching can damage their texture, necessitating non-thermal methods to regulate enzyme activity and protect the texture. The sauce relies on the co-fermentation of lactic acid bacteria, but different strains have different optimal conditions, and simultaneous inoculation can easily lead to competitive inhibition, requiring time-sequential release to control fermentation. Furthermore, pasteurization of the sauce can cause Maillard flavor loss, necessitating the decoupling of flavor preparation from pasteurization and achieving flavor retention through post-addition. These three technologies target the texture, flavor construction, and flavor locking stages respectively, and are interconnected and synergistic; none can be omitted.

[0022] I. Overall Process Architecture The production process of water chestnut and red pepper sauce of this invention is based on the core design concept of "self-catalytic cascade triggering closed loop", organizing the entire processing flow into a complete technical system with interconnected and mutually triggering processes. The overall process is carried out in the following order: raw material pretreatment, pulsed electric field treatment, material diversion and enzymatic hydrolysis, microcapsule preparation, cascade fermentation, subcritical water extraction of polyphenols, isolated Maillard reaction for flavoring, blending, pasteurization, Maillard flavor restoration and filling.

[0023] The term "autocatalytic cascade triggering closed loop" means that changes in the product or conditions of the preceding step automatically become the trigger signal for the following step. The entire process chain can complete the functional switching from enzymatic hydrolysis to fermentation and from fermentation to flavor enhancement without external human intervention. Specifically, this manifests as the following cascade triggering pathways: First, the two-stage pulsed electric field treatment maintains the material temperature below 40°C while inactivating enzymes and breaking cell walls, creating the preconditions for subsequent enzymatic hydrolysis by cellulase and pectinase under mild conditions. Moreover, no heat inactivation is performed after enzymatic hydrolysis, and the residual enzyme activity naturally decreases as the pH and temperature of the system decrease during fermentation, forming a seamless connection between "enzymatic hydrolysis and fermentation".

[0024] Second, type A and type B chitosan microcapsules with differentiated cross-linking densities encapsulate Leuconostoc mesenteroides and Lactobacillus plantarum, respectively. As the pH of the fermentation system gradually decreases, the microorganisms swell and release the microorganisms one after the other, forming a sequential sequence of "first producing esters and enhancing aroma, then rapidly producing acid". The pH change of the fermentation system itself becomes the trigger signal for the cascade release of microorganisms.

[0025] Third, the Maillard reaction is carried out in a reactor independent of the fermentation system, which avoids interference of reaction intermediates with the fermentation strains. The reaction products are added back after cooling and water extraction, after sterilization and cooling of the sauce, thus avoiding the loss of volatile flavor substances caused by heat sterilization.

[0026] Fourth, the water chestnut peel and pulp are collected separately and then enter different processing paths: the pulp is treated with a pulsed electric field and then split into an enzymatic hydrolysis part and a crisp granule retention part. The water chestnut peel is extracted with polyphenols by subcritical water extraction, and the extract is added to the finished sauce as an aqueous antioxidant component. At the same time, the flavonoids in the water chestnut peel are co-encapsulated in the core material during the microcapsule preparation stage to protect the fermentation strains, forming a closed loop of graded utilization of raw materials.

[0027] II. Raw material pretreatment and pulsed electric field treatment Water chestnut raw material pretreatment Fresh water chestnuts were washed and mechanically peeled, with the pulp and peel collected separately. The water chestnut peel was frozen at -18℃ or below for subsequent subcritical water extraction of polyphenols. The water chestnut pulp was cut into particles with a diameter of 5-10 mm. This particle size range balances the uniformity of electric field penetration during subsequent pulsed electric field treatment and the immersion efficiency of the material in the conductive medium: if the particle size is too large, the electric field will have difficulty penetrating the central region of the particle evenly; if the particle size is too small, the loss during the cutting process will increase and it will not be conducive to the preservation of the subsequent brittle particle morphology.

[0028] Two-stage pulsed electric field (PEF) processing Water chestnut pulp particles were immersed in a conductive medium for two-stage pulsed electric field treatment. The conductive medium was an aqueous solution with a conductivity of 0.3~1.5 mS / cm, and the volume ratio of pulp particles to conductive medium was 1:2~1:3. This volume ratio ensured that the pulp particles were fully dispersed in the conductive medium and avoided uneven electric field distribution due to particle accumulation.

[0029] The parameters for the first stage of pulsed electric field treatment are: electric field strength 15~20kV / cm, pulse width 20~30μs, pulse frequency 50~100Hz, and residence time 20~60s. The purpose of this stage of treatment is to use a lower electric field strength to form reversible micropores on the cell membrane of the fruit pulp, thereby increasing cell membrane permeability without causing complete cell wall rupture. At the same time, it causes conformational changes in the active sites of endogenous polyphenol oxidase and peroxidase in the fruit pulp, achieving partial enzyme inactivation.

[0030] The parameters for the second stage of pulsed electric field treatment are: electric field strength 25~35kV / cm, pulse width 30~50μs, pulse frequency 100~200Hz, and residence time 20~60s. Building upon the first stage of reversible perforation, this stage utilizes a higher electric field strength to further increase cell wall permeability, enabling cellulase and pectinase to effectively penetrate the pulp tissue during subsequent enzymatic hydrolysis, thereby improving enzymatic hydrolysis efficiency.

[0031] The material temperature is controlled at 40℃ or below throughout the two-stage pulsed electric field treatment. This temperature limit is key to ensuring that the texture (crispness) of water chestnut pulp is not damaged by heat: compared with traditional heat blanching to inactivate enzymes (usually 90~100℃), pulsed electric field treatment achieves enzyme inactivation and cell wall breaking through electroporation effect rather than heat effect, so that the pulp particles retain the crisp texture of the fresh state while gaining permeability. This is the physical basis for maintaining the texture of the water chestnut crisp particles in the finished sauce.

[0032] III. Material Diversion and Enzymatic Hydrolysis Material diversion design The material treated by the pulsed electric field is divided into two parts by mass: 70%~80% enters the enzymatic hydrolysis process, and 20%~30% is retained in the form of fruit pulp particles and temporarily stored at 0~4℃, and used as the crispy particle component in the subsequent blending stage.

[0033] The purpose of this diversion design is to simultaneously obtain two different forms of water chestnut components in the same batch of raw materials: the softened portion that becomes the sauce matrix after enzymatic hydrolysis and fermentation, and the granular portion that retains its original crispness without enzymatic hydrolysis. A temporary refrigerated storage temperature of 0–4°C inhibits the proliferation of microorganisms on the surface of the fruit pulp particles, while also preventing ice crystals from piercing cell walls and damaging the crisp texture caused by freezing.

[0034] Enzymatic hydrolysis process Cellulase and pectinase are added to the material entering the enzymatic hydrolysis process (i.e., 70%–80% fruit pulp). The amount of cellulase added is 0.05–0.15 parts by weight, and the amount of pectinase added is 0.02–0.08 parts by weight, based on the dry weight of the fruit pulp. The enzymatic hydrolysis conditions are: pH 4.5–5.0, temperature 45–50℃, and hydrolysis time 2–4 hours.

[0035] The optimal pH window for enzymatic hydrolysis is 4.5–5.0, which is shared by both cellulase and pectinase. Within this pH range, the synergistic activity of the two enzymes is high. Cellulase acts on the cellulose skeleton in the cell wall of the pulp, while pectinase acts on the pectin in the intercellular layer. Together, they moderately soften the pulp tissue and release cell contents, providing a usable carbon source (glucose, galacturonic acid, etc.) for subsequent fermentation.

[0036] After enzymatic hydrolysis, the hydrolysate is subjected to solid-liquid separation. Solid-liquid separation is performed by centrifugation or pressure filtration: for centrifugation, the rotation speed is 4000-6000 rpm and the centrifugation time is 10-15 min; for pressure filtration, the pressure is 0.2-0.3 MPa. The resulting liquid phase is used as the enzymatic hydrolysate for subsequent fermentation processes, while the resulting solid residue is used for the subsequent Maillard reaction process.

[0037] No heat inactivation treatment is performed after enzymatic hydrolysis. This is a key design feature that distinguishes this invention from traditional enzymatic hydrolysis processes: traditional processes typically inactivate residual enzymes by raising the temperature to 80-90°C and holding it for 10-15 minutes after enzymatic hydrolysis to terminate the reaction. This invention intentionally omits the heat inactivation step because: firstly, heat inactivation causes irreversible thermal damage to the texture of the fruit pulp particles, which is detrimental to maintaining a crisp texture; secondly, residual cellulase and pectinase gradually lose activity during subsequent fermentation processes due to changes in system temperature (22-25°C in the first stage, 30-33°C in the second stage, both lower than the enzymatic hydrolysis temperature of 45-50°C) and a gradual decrease in pH (from an initial pH of 5.5-6.5 to a final pH of 3.8-4.2), until the enzyme activity naturally decays to an undetectable level at the end of fermentation. This design achieves seamless integration of the enzymatic hydrolysis and fermentation processes, eliminating the need for a separate inactivation step and corresponding energy consumption.

[0038] IV. Microcapsule Preparation Chitosan solution preparation Chitosan was used as the wall material and dissolved in an aqueous acetic acid solution at a concentration of 15-20 g / L. The concentration of the acetic acid solution was adjusted according to the degree of deacetylation and molecular weight of the chitosan to ensure complete dissolution and the formation of a transparent and homogeneous solution.

[0039] Sodium tripolyphosphate (TPP) was used as an ionic crosslinking agent to prepare two different concentrations of TPP aqueous solutions for the preparation of type A and type B microcapsules with different crosslinking densities.

[0040] Preparation of type A microcapsules (low cross-linking density) The A-type microcapsules were prepared using an aqueous solution of sodium tripolyphosphate with a mass fraction of 0.5%–1.0% as a cross-linking agent. The core material consisted of activated Leuconostoc mesenteroides bacterial solution and concentrated flavonoid solution from water chestnut peel, with a volume ratio of 10:1.

[0041] The preparation process is as follows: the core material is added to a chitosan solution and stirred until homogeneous. Then, the mixture is dropped into a low-concentration TPP aqueous solution and cured for 20-30 minutes. The low concentration of TPP results in fewer ionic cross-linking sites between chitosan molecules, leading to a looser microcapsule network structure and lower cross-linking density. This allows the microcapsules to swell and release the core material even under low external pH stimulation.

[0042] Leuconostoc mesenteroides is the core strain in the first stage of fermentation in this invention. Its optimal growth temperature is 20-25℃, and its main metabolic function is heterolactic fermentation. While producing lactic acid, it also produces ester flavor compounds such as ethyl acetate and phenylethyl acetate, which impart the characteristic ester aroma to the sauce. Water chestnut peel flavonoid concentrate is used as a co-encapsulation component. Its flavonoids have antioxidant activity, providing a local antioxidant protective environment for the strain within the microcapsules, reducing the damage to strain vitality caused by oxidative stress during storage and the early stages of fermentation.

[0043] Preparation of type B microcapsules (high cross-linking density) The B-type microcapsules were prepared using a 2.0%–3.0% sodium tripolyphosphate aqueous solution as a cross-linking agent. The core material consisted of activated Lactobacillus plantarum solution and concentrated water chestnut peel flavonoid solution, with a volume ratio of 10:1.

[0044] The preparation process is as follows: the core material is added to a chitosan solution and stirred until homogeneous. Then, the mixture is dropped into a high-concentration TPP aqueous solution and cured for 40-60 minutes. The high concentration of TPP causes a dense ionic cross-linking network to form between chitosan molecules, resulting in a dense microcapsule structure with a high cross-linking density. The microcapsules require a lower external pH condition to swell and release the core material, and the curing time is longer to ensure sufficient cross-linking.

[0045] Lactobacillus plantarum is the core strain in the second stage of fermentation in this invention. Its optimal growth temperature is 30~35℃. Its main metabolic function is homolactic fermentation. It has a strong acid production capacity and can quickly reduce the pH of the system to 3.8~4.2. At the same time, it produces bacteriocins, which have a certain antibacterial function and are beneficial to the biological preservation and shelf stability of the finished sauce.

[0046] Functional logic of differentiated crosslinking density The differentiated cross-linking density design of type A and type B microcapsules is the key actuator of the "autocatalytic cascade triggering" mechanism of this invention. During fermentation, the pH of the system gradually decreases from the initial 5.5 to 6.5. When the pH of the system drops to about 5.0, the low cross-linking density type A microcapsules respond first, the chitosan network swells, and Leuconostoc mesenteroides is released, initiating the first stage of ester aroma metabolism.

[0047] When the pH of the system further drops to about 4.0, the highly cross-linked B-type microcapsules respond by releasing Lactobacillus plantarum, initiating the second stage of rapid acid production.

[0048] This cascade release process is entirely driven by the pH changes of the fermentation system itself, without the need for manual pH monitoring or the addition of a second strain, thus achieving self-triggering timing control of the fermentation process.

[0049] V. Cascade Fermentation Fermentation substrate construction The enzymatic hydrolysate and chopped red bell pepper were mixed at a mass ratio of 1.5:1 to 2.5:1. Based on a total fermentation substrate mass of 100 parts by mass, 1.5 to 4.0 parts by mass of edible salt, 0.3 to 0.8 parts by mass of type A microcapsules, and 0.3 to 0.8 parts by mass of type B microcapsules were added. After thorough mixing, the initial pH of the fermentation substrate was measured to be 5.5 to 6.5.

[0050] Red chili flakes are made by removing the stems and seeds from dried red chilies, rehydrating them, and then mincing them. This provides capsaicin and pigments to the fermentation system, while the soluble sugars and amino acids in the chili flakes also provide auxiliary carbon and nitrogen sources for the microbial metabolism. The amount of salt added is controlled between 1.5 and 4.0 parts by weight (i.e., the salt content of the finished sauce does not exceed 4%), which falls under the category of low-salt fermentation: the lower salt concentration is beneficial to the growth and metabolism of lactic acid bacteria, and also meets the current market demand for low-sodium condiments.

[0051] Type A and Type B microcapsules are added to the fermentation substrate simultaneously, but due to their different cross-linking densities, they will release the microorganisms sequentially at different pH points during the fermentation process.

[0052] Two-stage temperature-switched fermentation Phase 1: Fermentation temperature is controlled at 22-25℃, and fermentation time is 36-72 hours. This temperature range is the optimal growth temperature range for *Leuconostoc mesenteroides*. During this phase, when the system pH drops to approximately 5.0, the type A microcapsules swell and release *Leuconostoc mesenteroides*. The bacteria utilize glucose and fructose in the enzymatic hydrolysate for heterolactic fermentation, producing metabolites such as lactic acid, ethanol, and acetic acid. Ethanol further esterifies with organic acids to generate ester flavor compounds such as ethyl acetate. The fermentation progress in Phase 1 is determined by the system pH dropping to 5.0 or below.

[0053] The second stage involves raising the fermentation temperature to 30-33℃ and fermenting for 36-72 hours. This temperature range is the optimal growth temperature range for *Lactobacillus plantarum*. When the system pH drops to approximately 4.0, the B-type microcapsules swell and release *Lactobacillus plantarum*. The bacteria rapidly metabolize the remaining fermentable sugars at the higher temperature, producing a large amount of lactic acid, which quickly lowers the system pH to 3.8-4.2. The fermentation process in this second stage is considered complete when the system pH drops to 3.8-4.2. At this point, the fermented mash has a moderate acidity and a rich ester aroma.

[0054] The timing of temperature switching in the two-stage variable-temperature fermentation is matched with the cascade release sequence of the microcapsules: the low temperature in the first stage is conducive to the accumulation of ester aromas, while the increased temperature in the second stage is conducive to rapid acid production and pH decrease. The entire process is automatically linked by the pH response characteristics of type A and type B microcapsules, forming a flavor-building sequence of "ester aromas first, followed by acid production".

[0055] VI. Subcritical water extraction of polyphenols from water chestnut skin Take out the frozen water chestnut skin, add deionized water at a water-to-water ratio of 1:10 to 1:20, and perform subcritical water extraction at a temperature of 120 to 150°C and a gauge pressure of 0.8 to 1.5 MPa for 20 to 40 minutes.

[0056] Subcritical water refers to water that remains liquid at a temperature between 100 and 374°C and under certain pressure. Under these conditions, the dielectric constant and polarity of water are significantly reduced, and its solubility for moderately polar compounds (such as polyphenols and flavonoids) is enhanced, enabling efficient extraction of polyphenolic active ingredients from water chestnut peel without the use of organic solvents. The combination of extraction temperature of 120–150°C and gauge pressure of 0.8–1.5 MPa ensures that the water remains in a subcritical liquid state while avoiding thermal degradation of polyphenols due to excessively high temperatures.

[0057] After extraction, the extract is filtered through a microporous membrane with a pore size of 0.45 μm to remove solid impurities and macromolecular colloids. The filtrate is then concentrated to obtain a concentrated water chestnut peel polyphenol solution, which is used in subsequent blending processes.

[0058] The concentrated water chestnut peel polyphenols play two roles in the finished sauce: first, as an aqueous antioxidant, it forms a biphasic synergistic antioxidant system with the subsequently added lipid-phase vitamin E, delaying the oxidative deterioration of the oil components and pigments in the sauce; second, polyphenols have a certain antibacterial function, and together with the organic acids and bacteriocins produced during fermentation, they constitute a multi-layered biological preservation barrier for the finished sauce.

[0059] Furthermore, during the microcapsule preparation stage, the flavonoids in water chestnut peel (concentrated water chestnut peel flavonoids) were co-encapsulated in the core material of type A and type B microcapsules, providing localized antioxidant protection for the fermentation strains. This means that the active ingredients of water chestnut peel have two utilization pathways in this invention: one is to function as a component of the microcapsule core material during the fermentation stage, and the other is to function as a polyphenol concentrate in the finished sauce, exerting antioxidant and preservative effects, thus realizing the full-process utilization of the active ingredients of water chestnut peel.

[0060] VII. Flavor Enhancement via Isolated Maillard Reaction Reaction substrate and reaction enhancer The solid residue obtained after solid-liquid separation in the enzymatic hydrolysis process is used as the substrate for the Maillard reaction. The solid residue contains incompletely degraded pulp cell wall components, residual proteins, and polysaccharides, which are natural sources of reducing sugars and amino acids in the Maillard reaction.

[0061] Based on the dry weight of the solid residue, 0.02–0.08 parts by weight of L-cysteine ​​and 0.05–0.20 parts by weight of D-xylose were added as reaction enhancers. L-cysteine ​​is a sulfur-containing amino acid and a key precursor in the Maillard reaction for the formation of sulfur-containing heterocyclic compounds (such as thiazoles and thiophenes), which have characteristic soy sauce and meat aromas. D-xylose is a pentose sugar with higher Maillard reactivity than hexoses (such as glucose), and can preferentially participate in the reaction at lower temperatures, enriching the flavor profile of the reaction products.

[0062] Independent reactor and two-stage stepped temperature control The Maillard reaction is carried out in a separate reactor that is completely isolated from the fermentation system. The purpose of this isolation design is to avoid the inhibitory effect of intermediate products generated during the Maillard reaction (such as furfural, hydroxymethylfurfural, and other aldehydes) on the activity of the fermentation strain, while also preventing organic acids in the fermentation system from altering the Maillard reaction pathway and product composition.

[0063] The reaction conditions were: pH 6.0–7.0 (near neutral, which is conducive to the Maillard reaction), and a two-stage step temperature control was used. Stage 1: Temperature 80-90℃, reaction time 30-45 min. This is the low-temperature stage. At this temperature, L-cysteine ​​and reducing sugar preferentially form sulfur-containing heterocyclic compounds, including 2-methyl-3-furanthiol, 2-acetylthiazole, and other characteristic flavor compounds of soy sauce aroma. The reaction is milder under low-temperature conditions, which is conducive to the accumulation of volatile soy sauce aroma precursors rather than the excessive formation of dark melanoidins.

[0064] The second stage: temperature 105~115℃, reaction time 15~30min. This stage is a high-temperature stage, in which dehydration, cyclization and polymerization reactions are further carried out on the products of the low-temperature stage to generate high molecular weight coloring substances such as melanin and some non-volatile flavor precursors, providing the finished sauce with a natural brownish-red color and a mellow base flavor.

[0065] Cooling and water extraction After the Maillard reaction is complete, the reaction product is cooled to 40°C or below. The purpose of cooling is to prevent the loss of volatile flavor compounds caused by high temperatures during subsequent water extraction.

[0066] After cooling, add 3-5 times its weight of deionized water to the reaction product and extract with water at 50-60℃ for 30 minutes to fully dissolve the water-soluble flavor components in the reaction product into the aqueous phase. After water extraction, filter to obtain Maillard flavor extract, which is then stored for later use.

[0067] Maillard flavor extract is not added during the fermentation stage, but rather added back to the sauce after sterilization and cooling. This timing arrangement avoids secondary loss of volatile components in the flavor extract due to sterilization heat treatment.

[0068] VIII. Allocation The fermented mash is mixed with temporarily stored chilled water chestnut pulp (crispy pieces), concentrated water chestnut peel polyphenols, and vitamin E. The amount of vitamin E added is 0.02-0.06 parts by weight per 100 parts by weight of the prepared sauce.

[0069] The functional division of each component during the preparation process is as follows: Fermented mash provides the sauce matrix, containing organic acids, ester flavor compounds, and microbial metabolites produced during fermentation. Refrigerated water chestnut pulp particles are added as a crunchy component, providing a chewy texture. Water chestnut peel polyphenol concentrate, as an aqueous antioxidant, is dispersed in the aqueous phase of the sauce. Vitamin E, as a lipid antioxidant, is dispersed in the fat phase of the sauce (derived from the fat-soluble components of chopped red bell pepper).

[0070] Vitamin E and water chestnut peel polyphenols form a synergistic antioxidant system at the water-oil interface of the sauce: Vitamin E, as a fat-soluble antioxidant, mainly protects the oil phase components from free radical attack, while water chestnut peel polyphenols, as a water-soluble antioxidant, mainly protect the aqueous phase components. The synergistic regeneration mechanism of the two at the water-oil interface (polyphenols can reduce oxidized vitamin E and restore its antioxidant activity) makes the antioxidant capacity of the entire sauce system better than that of a single antioxidant.

[0071] Maillard flavor extract is not added during the blending stage; it is added back after sterilization.

[0072] 9. Pasteurization The prepared materials are pasteurized at a temperature of 55-58℃ for 15-20 minutes.

[0073] The sterilization conditions were chosen based on the following considerations: 55-58℃ is a mild sterilization temperature, which effectively reduces the load of spoilage microorganisms and pathogenic bacteria (such as Escherichia coli and Staphylococcus aureus) in the sauce while preserving the integrity of volatile ester flavor compounds accumulated during fermentation to the greatest extent possible, avoiding the loss of cooking flavor and taste caused by high-temperature sterilization (usually 85-95℃). Simultaneously, this temperature is lower than the heat denaturation temperature of bacteriocins produced by Lactobacillus plantarum, ensuring that residual bacteriocins in the sauce retain their antibacterial activity after sterilization, providing continuous biological preservative function for the finished sauce. The sterilization temperature is also lower than the critical heat softening temperature of pectin and cellulose in the crispy granule component, avoiding thermal damage to the crispy granule texture.

[0074] 10. Maillard flavor reintroduction and bottling Maillard flavor extract refill After pasteurization, when the temperature of the sauce drops to 40°C or below, add 1 to 3 parts by weight of Maillard flavor extract to the sauce, based on a total mass of 100 parts by weight, and stir well.

[0075] The temperature for refilling is controlled at 40℃ or below to prevent heat loss of volatile soy sauce aroma components (such as sulfur-containing heterocyclic compounds and furans) in the Maillard flavor extract due to high temperatures. Aseptic addition means that the flavor extract is filtered through a microporous membrane to remove bacteria before addition, avoiding the introduction of exogenous microorganisms to contaminate the sterilized sauce.

[0076] Filling and nitrogen pulse replacement After the sauce containing Maillard flavor extract is filled into the packaging container, the headspace of the packaging container is purged with nitrogen pulses before sealing. The nitrogen pulse purging is performed 3 times. Each time, nitrogen is introduced until the headspace pressure is 0.15~0.20MPa and held for 5 seconds before being vented. The container is sealed immediately after the third nitrogen purging.

[0077] The principle of nitrogen pulse replacement is to repeatedly fill and exhaust high-purity nitrogen to gradually dilute and replace the air (containing approximately 21% oxygen) in the headspace of the packaging container, reducing the residual oxygen level in the headspace to 0.5% or below after sealing. This low residual oxygen environment effectively inhibits the oxidative rancidity of the oil components in the sauce, the oxidative fading of pigments, and the oxidative consumption of vitamin E and polyphenolic antioxidants during storage, thus extending the product's shelf life.

[0078] The sealed finished water chestnut and red pepper sauce has the following characteristics: pH 3.8-4.2, salt content not exceeding 4%, and headspace residual oxygen content not exceeding 0.5%.

[0079] XI. Summary of the Closed-Loop Utilization of Skin and Flesh Grading This invention utilizes the skin and flesh of water chestnuts in a graded manner throughout the entire process: The utilization path of water chestnut pulp: pulp → granulation → two-stage PEF processing → diversion (70%~80% enzymatic hydrolysis + 20%~30% crisp granules refrigerated) → enzymatic hydrolysate enters fermentation → crisp granules are added back to the finished sauce.

[0080] Utilization path 1 for water chestnut peel (polyphenol extraction): Water chestnut peel → freeze preservation → subcritical water extraction → filtration and concentration → polyphenol concentrate → added to finished sauce as an aqueous phase antioxidant component.

[0081] Utilization pathway two for water chestnut peel (flavonoid co-encapsulation): Water chestnut peel → extraction of concentrated water chestnut peel flavonoids → co-encapsulation in type A and type B microcapsule cores → providing antioxidant protection for the microorganisms during fermentation.

[0082] Through the two pathways mentioned above, the active ingredients in water chestnut skin play their roles in the fermentation stage (flavonoids protect the microbial strain) and the finished product storage stage (polyphenols provide antioxidant and antibacterial effects), achieving full utilization of raw materials and zero waste.

[0083] To verify the actual impact of the core process parameters in this invention on the quality of the finished water chestnut and red pepper sauce, a single-factor controlled experiment was designed. The experiment used the texture characteristics, flavor features, and storage stability of the finished sauce as evaluation dimensions, selecting three technical characteristics that had the most significant impact on product quality as experimental variables, while keeping all other process conditions consistent. All testing items were performed using currently valid national or international standard methods. The specific standard numbers used for the testing are not detailed here, but the detailed operational descriptions of the experimental steps ensure that those skilled in the art can fully reproduce the results.

[0084] I. Selection and Basis of Experimental Variables Based on physicochemical principles and food processing principles, the three technical characteristics that have the greatest impact on the quality of the finished sauce are as follows: Variable A: Pulse electric field processing method and temperature control Pulsed electric field treatment is the core technology in this invention, replacing traditional heat-induced enzyme inactivation. According to electroporation theory, a pulsed electric field induces a transmembrane potential in the cell membrane. When this potential exceeds a critical threshold, reversible perforation occurs. The electric field strength and treatment method directly determine the degree of perforation and the magnitude of changes in cell wall permeability. The physical significance of the two-stage progressive field strength design is that the low-field-strength stage first forms micropores, while the high-field-strength stage expands the pore size based on the existing micropores. Compared to single-stage high-field-strength treatment, this avoids cell disintegration caused by a single large-pore perforation. Simultaneously, the restriction that the material temperature throughout the process does not exceed 40℃ ensures that the pectin and cellulose backbone do not undergo thermal degradation, a prerequisite for maintaining the crisp texture of the granules. This variable has the greatest impact on the textural properties of the water chestnut crisp granules in the finished sauce.

[0085] Variable B: Differential cross-linking density between type A and type B microcapsules and two-stage temperature-switched fermentation Differential crosslinking density is the structural basis for the sequential release of core material from chitosan microcapsules at specific pH thresholds. Based on the chitosan-sodium tripolyphosphate (TPP) ionic crosslinking mechanism, the phosphate groups in sodium tripolyphosphate electrostatically crosslink with the amino groups on the chitosan molecular chain. Higher crosslinking agent concentrations result in denser crosslinking sites, a more compact network structure, and a lower pH threshold required for microcapsule swelling. The differentiated design of type A (low TPP concentration) and type B (high TPP concentration) microcapsules allows for the release of *Leuconostoc mesenteriae* and *Lactobacillus plantarum* at different points during the pH decrease in the fermentation system, forming a sequential release pattern in conjunction with two-stage temperature control. This variable has the greatest impact on the flavor composition of the finished sauce (the balance between ester aroma and acidity) and the pH stability at the fermentation endpoint.

[0086] Variable C: The Maillard reaction was carried out in a separate reactor and added back after sterilization and cooling. The volatile flavor compounds (including sulfur heterocyclic compounds and furans) produced by the Maillard reaction have low boiling points. If added to the sauce before sterilization, the pasteurization process at 55-58°C will result in heat loss of some volatile components. Conducting the Maillard reaction independently of the fermentation system avoids the inhibition of microbial activity by reaction intermediates, while adding the mixture back after sterilization at below 40°C maximizes the retention of volatile flavor components. This variable has the greatest impact on the characteristic flavor intensity of the finished sauce.

[0087] II. Experimental Group Design 1. Experimental Group (Complete Protocol Group) The complete process scheme of this invention is implemented as follows: Raw material pretreatment: Take 10 kg of fresh water chestnuts from the same batch, wash them, and mechanically peel them, collecting the pulp and peel separately. Store the water chestnut peels frozen at -18℃. Cut the pulp into granules with a diameter of 5-10 mm.

[0088] Pulsed electric field treatment (experimental conditions for variable A): Fruit pulp particles were immersed in a conductive medium with a conductivity of 1.0 mS / cm, with a volume ratio of fruit pulp particles to conductive medium of 1:2.5. First stage of pulsed electric field treatment: electric field strength 18 kV / cm, pulse width 25 μs, pulse frequency 80 Hz, residence time 40 s. Second stage of pulsed electric field treatment: electric field strength 30 kV / cm, pulse width 40 μs, pulse frequency 150 Hz, residence time 40 s. The material temperature was controlled below 40℃ throughout the process.

[0089] Material separation and enzymatic hydrolysis: 75% of the treated material was fed into the enzymatic hydrolysis chamber by mass, while 25% was retained as brittle granules and temporarily stored at 2℃. 0.10 parts by mass of cellulase and 0.05 parts by mass of pectinase (based on the dry weight of the pulp) were added to the enzymatic hydrolysis portion, and enzymatic hydrolysis was performed for 3 hours at pH 4.8 and a temperature of 48℃. After enzymatic hydrolysis, no heat inactivation was performed; instead, solid-liquid separation was directly performed (centrifugation at 5000 rpm for 12 minutes) to obtain the enzymatic hydrolysate and solid residue.

[0090] Microcapsule preparation (experimental conditions for variable B): Chitosan was dissolved in an aqueous acetic acid solution with a mass-to-volume concentration of 18 g / L. Type A microcapsules: TPP mass fraction 0.8%, core material was a mixture of *Leuconostoc mesenteriae* activated bacterial solution and *Carex esculenta* flavonoid concentrate at a volume ratio of 10:1, cured for 25 min. Type B microcapsules: TPP mass fraction 2.5%, core material was a mixture of *Lactobacillus plantarum* activated bacterial solution and *Carex esculenta* flavonoid concentrate at a volume ratio of 10:1, cured for 50 min.

[0091] Fermentation (experimental conditions for variable B): The enzymatic hydrolysate and chopped red pepper were mixed in a 2:1 mass ratio. Based on 100 parts by mass of the total fermentation substrate, 2.5 parts by mass of salt, 0.5 parts by mass of type A microcapsules, and 0.5 parts by mass of type B microcapsules were added. The initial pH was 6.0. The first stage of fermentation was carried out at 22-25℃ for 48 hours until the pH dropped below 5.0. The second stage was carried out at 30-33℃ for 48 hours until the pH dropped to 3.8-4.2, yielding the fermented mash.

[0092] Subcritical water extraction: The mass ratio of water chestnut peel to water was 1:15, the temperature was 135℃, the gauge pressure was 1.2MPa, and the extraction time was 30 min. The extract was then concentrated after filtration through a 0.45μm microporous membrane to obtain a concentrated water chestnut peel polyphenol solution.

[0093] Maillard reaction (experimental conditions for variable C): The solid residue was placed in an independent reaction vessel, and 0.05 parts by weight of L-cysteine ​​and 0.12 parts by weight of D-xylose were added based on the dry weight of the residue. The pH was 6.5. The first stage reaction was carried out at 85°C for 40 min, and the second stage reaction was carried out at 110°C for 20 min. After cooling to below 40°C, 4 times the mass of water of the reaction product was added, and water extraction was carried out at 55°C for 30 min. The Maillard flavor extract was obtained by filtration.

[0094] Blending: Fermented mash is mixed with chilled crispy granules, concentrated water chestnut peel polyphenols, and vitamin E (0.04 parts by weight, based on 100 parts by weight of the total blended sauce).

[0095] Sterilization: Pasteurize at 55~58℃ for 18 minutes.

[0096] Maillard flavor reintroduction and bottling (experimental conditions for variable C): After sterilization, cool to below 40°C, add 2 parts by weight of Maillard flavor extract (based on a total mass of 100 parts by weight of the prepared sauce), and stir well. Bottling into glass bottles, performing 3 nitrogen pulse purgings before sealing (purging nitrogen to 0.18 MPa and holding for 5 seconds each time, then venting), and sealing after the third nitrogen purging.

[0097] 2. Control group A (enzyme inactivation by heat instead of pulsed electric field treatment) The process conditions were identical to those of the experimental group, except that the pulsed electric field treatment was replaced with traditional hot blanching and enzyme inactivation treatment. Specifically, the water chestnut pulp granules were blanched in 95°C hot water for 5 minutes, then quickly immersed in ice water to cool to room temperature, drained, and then proceeded to the subsequent fractionation process. All other subsequent processes (fractionation, enzymatic hydrolysis, fermentation, blending, sterilization, and filling) were completely consistent with those of the experimental group.

[0098] This control group was used to verify the superiority of two-stage pulsed electric field treatment over traditional heat-induced enzyme inactivation in maintaining the texture of water chestnut crisps.

[0099] 3. Control group B (uniform cross-linked density microcapsules + isothermal fermentation) The process conditions were identical to those of the experimental group, with the only difference being adjustments to the microcapsule preparation and fermentation conditions. Both type A and type B microcapsules used 1.5% TPP (i.e., a uniform medium cross-linking density), with the core material composition remaining unchanged, and the curing time uniformly set at 35 minutes. The fermentation process did not differentiate between two temperature variations; fermentation was conducted entirely at a constant temperature of 28℃ until the pH dropped to 3.8–4.2, at which point fermentation was terminated. All other procedures were completely consistent with the experimental group.

[0100] This control group was used to verify the superiority of differentiated crosslinking density combined with two-stage variable temperature fermentation over uniform crosslinking density combined with isothermal fermentation in terms of flavor construction and fermentation controllability.

[0101] 4. Control group C (Maillard reaction product added before sterilization and subjected to sterilization together) The process conditions were the same as those of the experimental group, except that the Maillard flavor extract was added and mixed with fermented mash, crispy granules, polyphenol concentrate and vitamin E at the same time during the preparation stage, and then pasteurized together at 55~58℃ for 18 minutes. After the sterilization, the temperature was directly lowered to below 40℃ for filling and capping (nitrogen pulse replacement was still performed), and it was not added back separately after sterilization.

[0102] This control group was used to verify the superiority of adding the product after sterilization and cooling in preserving volatile flavor compounds compared to adding it before sterilization.

[0103] III. Testing Items and Methods 1. Texture property detection (corresponding variable A) Test subjects: Water chestnut crunchy component in the finished sauce of experimental group and control group A, as well as unprocessed fresh water chestnut pulp (fresh sample standard).

[0104] Method for separating brittle particles: Take 100g of the finished sauce sample, add 200mL of deionized water, gently stir and disperse with a glass rod, then pass through a standard test sieve with a 2mm aperture. Rinse the residue on the sieve three times with deionized water, transfer the residue to clean filter paper, gently blot dry with the filter paper, and then use it for texture analysis. The entire operation should be completed within 30 minutes after sampling to avoid the brittle particles losing moisture in the air and affecting the test results.

[0105] Texture analyzer measurement method (TPA mode): A texture analyzer with a 25mm diameter cylindrical flat-bottom probe was used. The measurement mode was set to TPA (texture profile analysis) dual-cycle compression test. The measurement parameters were as follows: pre-test speed 1.0mm / s, test speed 1.0mm / s, post-test speed 1.0mm / s, compression deformation of 50% of the original sample height, trigger force 5g, and interval between two compressions 5s. A single horseshoe-shaped brittle particle was placed at the center of the texture analyzer stage, and the probe was aligned with the center of the particle for two consecutive compressions. The software automatically recorded the force-displacement curve and calculated the texture parameters. At least 10 brittle particles were measured in parallel for each sample group, and the average value was taken after removing outliers.

[0106] Record the following indicators: Hardness (the maximum peak force during the first compression, in N), Brittleness (the force value of the first obvious fracture peak during the first compression, in N), Elasticity (the ratio of the recovery height during the second compression to the height during the first compression, dimensionless), and Chewing property (hardness × cohesion × elasticity, in N).

[0107] Fresh control sample: Take fresh water chestnut pulp that has not undergone any processing, cut it into particles with the same particle size (5~10mm) as the experimental group, and measure the texture parameters according to the above method to serve as a fresh baseline control.

[0108] Texture retention rate calculation: Based on the hardness value of the fresh sample as 100%, calculate the hardness retention rate (%) of the finished crispy granules of the experimental group and the control group A respectively = (hardness value of finished crispy granules / hardness value of fresh sample) × 100%.

[0109] 2. Detection of volatile flavor components (corresponding to variables B and C) Test subjects: finished sauces from the experimental group, control group B, and control group C.

[0110] Analysis of volatile flavor components (headspace solid-phase microextraction-gas chromatography-mass spectrometry): Sample pretreatment: Take 5.0g of the finished sauce sample and place it in a 20mL headspace vial. Add 5mL of saturated sodium chloride solution (to promote the release of volatile components) and 10μL of internal standard solution (2-octanol, concentration 1.0mg / mL). After sealing the vial, place it in a 45℃ constant temperature water bath for 30min to equilibrate.

[0111] Extraction: Insert the aged 50 / 30μm DVB / CAR / PDMS composite solid-phase microextraction fiber head approximately 1 cm above the liquid surface in the headspace vial, and perform headspace adsorption at 45℃ for 30 min. After adsorption is complete, quickly remove the fiber head and immediately insert it into the gas chromatograph injection port.

[0112] GC-MS analysis conditions: The chromatographic column was a polar capillary column (polyethylene glycol stationary phase, column length 30 m, inner diameter 0.25 mm, film thickness 0.25 μm). Injector temperature 250℃, splitless injection, fiber head desorption time 5 min. Carrier gas was high-purity helium (purity ≥99.999%), flow rate 1.0 mL / min. Temperature program: initial temperature 40℃, hold for 3 min, increase to 120℃ at 5℃ / min, then increase to 230℃ at 10℃ / min, hold for 5 min. Mass spectrometry used an electron impact ionization (EI) source, ionization energy 70 eV, ion source temperature 230℃, quadrupole temperature 150℃, transfer line temperature 250℃, scan range m / z 35~450.

[0113] Qualitative method: The obtained mass spectrum is compared and searched with the standard mass spectrum library. The retention index (RI) of each component on the polar column (calculated by the C7~C30 standard of n-alkane series) is used for double confirmation. Components with a matching degree greater than 800 and RI deviation within ±10 are determined to be valid identifications.

[0114] Quantitative method: Relative quantification was performed using the internal standard method. The content of each volatile component was calculated based on the peak area ratio of each target component to the internal standard and the relative response factor, using the peak area of ​​2-octanol as a baseline. The results are expressed in μg / kg.

[0115] Key metrics to record: The total amount of ester compounds (represented by ethyl acetate, phenylethyl acetate, and ethyl lactate, reflecting the ester aroma metabolism level of Leuconostoc mesenteroides).

[0116] Total amount of sulfur-containing heterocyclic compounds (represented by 2-methyl-3-furanthiol and 2-acetylthiazole, reflecting the degree of retention of characteristic flavor substances of Maillard reaction).

[0117] Total amount of furan compounds (represented by furfural and 5-methylfurfural, reflecting the retention of Maillard reaction intermediates).

[0118] Volatile organic acids (represented by acetic acid, reflecting the level of acid production during fermentation).

[0119] 3. Detection of non-volatile organic acids (corresponding variable B, supplementary item) Because organic acids (lactic acid, acetic acid, etc.) have high boiling points and strong polarity, they are not suitable for direct detection by GC-MS and must be determined separately by high performance liquid chromatography (HPLC).

[0120] Test subjects: the finished sauces of experimental group and control group B.

[0121] Sample pretreatment: Take 10.0 g of the finished sauce sample, add 40 mL of ultrapure water, extract by sonication for 20 min, and centrifuge at 10000 rpm for 15 min at 4℃. Take the supernatant and filter it through a 0.22 μm aqueous microporous membrane. Use the filtrate for HPLC analysis.

[0122] HPLC analysis conditions: A C18 reversed-phase column (250 mm length, 4.6 mm inner diameter, 5 μm particle size) was used; the mobile phase was 0.02 mol / L potassium dihydrogen phosphate aqueous solution (pH adjusted to 2.5 with phosphoric acid); the flow rate was 0.8 mL / min; the column temperature was 30℃; the detection wavelength was 210 nm; and the injection volume was 10 μL. External standard method was used, with standard curves constructed using lactic acid, acetic acid, and citric acid standards for quantification. Results are expressed in g / kg.

[0123] Key indicators to record: lactic acid content and acetic acid content. Lactic acid mainly originates from homolactic fermentation by *Lactobacillus plantarum*, while acetic acid mainly originates from heterolactic fermentation by *Leuconostoc mesenteroides*. The ratio of these two components reflects the extent to which the metabolic functions of the two strains are utilized and the effectiveness of their sequential fermentation.

[0124] 4. Fermentation process monitoring and finished product stability testing (corresponding variable B) Subjects tested: Experimental group and control group B.

[0125] pH dynamic monitoring during fermentation: Samples were taken every 6 hours during fermentation, with a sample size of approximately 5g. After homogenization with 10mL of deionized water, the pH value was measured using a pH meter calibrated with standard buffer solutions (pH 4.01 and pH 6.86), and a pH-time curve was plotted. The following key time points were recorded: the time when the pH first dropped to 5.0 (reflecting the release of type A microcapsules and the initiation rate of the first stage of fermentation), the time when the pH first dropped to 4.0 (reflecting the release of type B microcapsules and the initiation rate of the second stage of fermentation), and the total fermentation time required to reach the fermentation endpoint of pH 3.8–4.2.

[0126] pH stability test of finished sauce: The finished sauce was stored at a constant temperature of 25℃ in the dark. Samples were taken on days 0, 30, 60 and 90 of storage, and the pH value was measured according to the above method to evaluate the pH stability during storage. The smaller the pH fluctuation, the more stable the fermentation endpoint control and the stronger the buffering capacity of the product system.

[0127] Total colony count detection: Samples were taken at the above storage time points. Aseptically weigh 25g of sample and add 225mL of sterile physiological saline (0.85% NaCl). Homogenize at 8000rpm for 2min to prepare a 1:10 dilution. Perform serial dilutions of 10-fold to the appropriate concentration. Select 1mL of bacterial suspension from 2-3 appropriate dilutions and inject them into sterile Petri dishes (90mm diameter), with two replicates for each dilution. Pour approximately 15mL of plate counting agar medium cooled to 46±1℃ into the Petri dishes, gently swirl to mix, and invert the plates after the medium has solidified. Incubate at 36±1℃ for 48±2h, then count the colonies. Select plates with colony counts between 30 and 300 for counting. Calculate the average of the two plate counts and multiply by the dilution factor. The result is expressed as CFU / g. If the colony counts on all plates are outside the range of 30-300, select the plate with the closest count of 300 and label it "estimated value".

[0128] 5. Comprehensive testing of storage stability (corresponding to the combined influence of variables A, B, and C) Test subjects: finished sauces from experimental group, control group A, control group B, and control group C.

[0129] Peroxide value determination: Take approximately 50g of the finished sauce sample, add 100mL of petroleum ether (boiling range 30~60℃), soak and extract at room temperature for 12h, filter, and remove the solvent by rotary evaporation in a 40℃ water bath to obtain the oil extract. Weigh approximately 2.0g of the oil sample (accurate to 0.001g) and place it in a 250mL iodine flask, add 30mL of a chloroform-glacial acetic acid mixture (volume ratio 2:3), and gently shake to completely dissolve the oil. Add 1.00mL of saturated potassium iodide solution, stopper tightly, and gently shake for 30s. Place in the dark for 5min (accurate timing). Add 100mL of freshly boiled and cooled distilled water, and titrate with 0.01mol / L sodium thiosulfate standard solution while vigorously shaking. Titrate until the solution turns pale yellow, then add 1mL of starch indicator (10g / L), and continue titrating until the blue color just disappears and remains unchanged for 30s as the endpoint. Perform a blank test simultaneously. Record the volume of sodium thiosulfate consumed, calculate the peroxide value using the formula, and express the result in g / 100g. Samples were taken and measured on day 0 and day 90 of storage, respectively.

[0130] Color Measurement: The color of the finished sauce was measured using a colorimeter. The instrument was preheated for 30 minutes and then calibrated using a standard white and black plate. A sample of the finished sauce was evenly filled into a cuvette (depth not less than 10 mm), and the surface was compacted with a flat scraper to ensure it was smooth and free of air bubbles. Using the CIE Lab color space and with the standard white plate as a reference, the L value (lightness, 0 for black, 100 for white), a value (red-green axis, positive for red, negative for green), and b value (yellow-blue axis, positive for yellow, negative for blue) were measured. Each sample was measured at least 5 times at different locations, and the average value was taken. Samples were taken and measured on day 0 and day 90 of storage, and the total color difference change before and after storage was calculated using the formula ΔE=[(ΔL)²+(Δa)²+(Δb)²]^(1 / 2). The smaller the ΔE, the better the color stability during storage.

[0131] IV. Experimental Expectations Based on physicochemical principles and process control, the expected experimental results trend is as follows: Regarding variable A: the water chestnut crisps from the experimental group (treated with a two-stage pulsed electric field) should have significantly higher hardness, crispness, and chewiness than those from control group A (treated with heat-induced enzyme inactivation), and the hardness retention rate should be closer to 100% (fresh sample baseline). This is because heat-induced degradation of pectin and cellulose during heat treatment softens the cell wall skeleton, while pulsed electric field treatment improves permeability through a non-thermoelectric perforation mechanism while preserving the structural integrity of the cell wall.

[0132] Regarding variable B: The total amount of ester compounds in the finished sauce from the experimental group (differentiated cross-linking density + two-stage variable temperature fermentation) should be significantly higher than that from control group B (uniform cross-linking density + isothermal fermentation). The lactic acid / acetic acid ratio determined by HPLC should also show a significant difference, reflecting the orderly connection of metabolic functions between the two strains. The pH curve during fermentation should exhibit a smoother two-stage decrease, rather than the sudden drop or stagnation of pH that may occur in control group B. During storage, the pH fluctuation range and the increase in total bacterial count should both be better than those in control group B.

[0133] Regarding variable C: The content of sulfur-containing heterocyclic compounds and furan-like flavor substances in the finished sauce of the experimental group (added after sterilization and cooled down) should be significantly higher than that of the control group C (added before sterilization and sterilized together). This is because adding it back after sterilization avoids the secondary loss of these low-boiling-point flavor components during the 55~58℃ heat treatment.

[0134] Regarding storage stability: the experimental group had the lowest increase in peroxide value and total color difference change ΔE on day 90, which reflects the comprehensive synergistic effect of the three core technologies: pulsed electric field non-thermal texture preservation, differentiated microcapsule orderly fermentation, and flavor restoration after sterilization.

[0135] The selection of process parameters in this invention does not aim for the optimization of a single indicator, but rather seeks an engineering balance among multiple mutually restrictive technical effects. Taking two-stage pulsed electric field treatment as an example, while higher electric field strength and longer residence time can improve cell membrane permeability and enzymatic hydrolysis efficiency, excessive energy may lead to irreversible electrical breakdown, causing structural degradation. Therefore, this invention limits the field strength and residence time to a specific range to balance the improvement of permeability with the preservation of structural integrity. In a two-stage variable-temperature fermentation system with differentiated cross-linking density microcapsules, a greater difference in cross-linking density between type A and type B microcapsules can make the cascade release sequence clearer, which is beneficial to ester synthesis. However, a longer fermentation cycle will increase the accumulation of metabolic byproducts and affect the stability of the pH buffer system. Therefore, this invention controls the difference in cross-linking agent concentration within a reasonable range to achieve comprehensive optimization of flavor richness and storage stability. The above compromise solutions can be adjusted by technical personnel according to actual needs.

[0136] Exemplary Description Example 1: Effect of two-stage pulsed electric field treatment on the preservation of texture of horseshoe-shaped brittle grains This embodiment focuses on the variable of pulsed electric field treatment, examining its impact on the textural properties of water chestnut crispness in the finished sauce. This embodiment uses a gradient of pulsed electric field treatment intensity to set up experimental groups, with each sub-parameter (field strength, pulse width, frequency, and residence time) varying synergistically to create three treatment intensities: low, medium, and high. This embodiment includes six experimental groups, all with identical process parameters except for the pulsed electric field treatment conditions.

[0137] The common constant process conditions for all groups are as follows: Raw material pretreatment: Take 10 kg of fresh water chestnuts from the same batch, wash them, and mechanically peel them, collecting the pulp and peel separately. Store the water chestnut peels frozen at -18℃. Cut the pulp into 8mm particles.

[0138] Material separation and enzymatic hydrolysis: 75% of the treated material was fed into the enzymatic hydrolysis chamber by mass, while 25% was retained as brittle granules and temporarily stored at 2℃. 0.10 parts by mass of cellulase and 0.05 parts by mass of pectinase (based on the dry weight of the pulp) were added to the enzymatic hydrolysis portion, and enzymatic hydrolysis was performed for 3 hours at pH 4.8 and a temperature of 48℃. After enzymatic hydrolysis, no heat inactivation was performed; instead, solid-liquid separation was directly performed (centrifugation at 5000 rpm for 12 minutes) to obtain the enzymatic hydrolysate and solid residue.

[0139] Microcapsule preparation: Chitosan was dissolved in an aqueous acetic acid solution to a concentration of 18 g / L. Type A microcapsules: TPP mass fraction 0.8%, core material was a mixture of *Leuconostoc mesenteriae* activated bacterial solution and *Carex esculenta* flavonoid concentrate at a volume ratio of 10:1, cured for 25 min. Type B microcapsules: TPP mass fraction 2.5%, core material was a mixture of *Lactobacillus plantarum* activated bacterial solution and *Carex esculenta* flavonoid concentrate at a volume ratio of 10:1, cured for 50 min.

[0140] Fermentation: The enzymatic hydrolysate and chopped red pepper were mixed at a mass ratio of 2:1. Based on 100 parts by mass of the total fermentation substrate, 2.5 parts by mass of salt, 0.5 parts by mass of type A microcapsules, and 0.5 parts by mass of type B microcapsules were added, with an initial pH of 6.0. The first stage of fermentation was carried out at 23℃ for 48 hours, and the second stage was carried out at 32℃ for 48 hours. The final fermentation pH was 4.0.

[0141] Subcritical water extraction: The mass ratio of water chestnut peel to water was 1:15, the temperature was 135℃, the gauge pressure was 1.2MPa, and the extraction time was 30 min. The extract was then concentrated after filtration through a 0.45μm microporous membrane to obtain a concentrated water chestnut peel polyphenol solution.

[0142] Maillard reaction: The solid residue was placed in a separate reaction vessel, and 0.05 parts by weight of L-cysteine ​​and 0.12 parts by weight of D-xylose were added based on the dry weight of the residue, with a pH of 6.5. The first stage reaction was carried out at 85°C for 40 min, and the second stage reaction was carried out at 110°C for 20 min. After cooling to below 40°C, 4 times the mass of water of the reaction product was added, and water extraction was carried out at 55°C for 30 min. The Maillard flavor extract was obtained by filtration.

[0143] Blending: Fermented mash is mixed with chilled crispy granules, concentrated water chestnut peel polyphenols, and vitamin E (0.04 parts by weight, based on 100 parts by weight of the total blended sauce).

[0144] Sterilization: Pasteurize at 56℃ for 18 minutes.

[0145] Maillard flavor reintroduction and bottling: After sterilization, cool to 38°C, add 2 parts by weight of Maillard flavor extract (based on 100 parts by weight of the total sauce), and stir well. Fill into glass bottles, and perform 3 nitrogen pulse purgings before sealing (purging nitrogen to 0.18 MPa and holding for 5 seconds each time, then venting). Seal the bottles after the third nitrogen purging.

[0146] The variables for each group are set as follows: Experimental Group 1 (Medium Treatment Intensity): Conductivity of the conductive medium was 1.0 mS / cm, and the volume ratio of fruit pulp particles to the conductive medium was 1:2.5. First stage of pulsed electric field treatment: electric field strength 18 kV / cm, pulse width 25 μs, pulse frequency 80 Hz, residence time 40 s. Second stage of pulsed electric field treatment: electric field strength 30 kV / cm, pulse width 40 μs, pulse frequency 150 Hz, residence time 40 s. The material temperature was controlled below 38℃ throughout the process.

[0147] Experimental Group 2 (Low Processing Intensity): The difference between this experimental group and Experimental Group 1 is as follows: the electric field intensity of the first pulsed electric field treatment was 15 kV / cm, the pulse width was 20 μs, the pulse frequency was 50 Hz, and the residence time was 20 s; the electric field intensity of the second pulsed electric field treatment was 25 kV / cm, the pulse width was 30 μs, the pulse frequency was 100 Hz, and the residence time was 20 s. The material temperature was controlled below 35℃ throughout the process.

[0148] Experimental Group 3 (High Processing Intensity): The difference between this experimental group and Experimental Group 1 is as follows: the electric field intensity of the first pulsed electric field treatment is 20 kV / cm, the pulse width is 30 μs, the pulse frequency is 100 Hz, and the residence time is 60 s; the electric field intensity of the second pulsed electric field treatment is 35 kV / cm, the pulse width is 50 μs, the pulse frequency is 200 Hz, and the residence time is 60 s. The material temperature is controlled below 40℃ throughout the process.

[0149] Experimental Group 4 (Blanching Control Group): The difference between this experimental group and Experimental Group 1 is that pulsed electric field treatment is not performed. Instead, the water chestnut pulp particles are blanched in 95℃ hot water for 5 minutes, then quickly immersed in ice water to cool to room temperature, drained, and then proceeded to the subsequent sorting process. All other process parameters are completely consistent with those of Experimental Group 1.

[0150] Experimental Group 5 (Comprehensive Control Group): The difference between this experimental group and Experimental Group 1 is as follows: Pulsed electric field treatment was omitted; instead, water chestnut pulp particles were blanched in 95℃ hot water for 5 minutes, then quickly immersed in ice water to cool to room temperature, drained, and proceeded to subsequent processes. Microcapsules were made with a uniform TPP mass fraction of 1.5% and solidified for 35 minutes. Fermentation was carried out at a constant temperature of 28℃ throughout. The Maillard flavor extract was added during the preparation stage and underwent pasteurization together with the microcapsules, without being added separately after sterilization. This group also changed the pulsed electric field treatment method, microcapsule cross-linking density, fermentation temperature mode, and the timing of Maillard flavor extract addition to demonstrate the comprehensive effect of existing conventional processes and was not used as a single-factor control.

[0151] Example 2: The effect of differentiated cross-linking density microcapsules combined with two-stage temperature-switched fermentation on flavor development This embodiment focuses on the variables of microcapsule cross-linking density difference and fermentation temperature control, examining their impact on the flavor composition and fermentation controllability of the finished sauce. This embodiment sets up experimental groups based on the comprehensive parameter levels of the cascade fermentation system. The microcapsule cross-linking density difference changes synergistically with the fermentation temperature / time parameters, forming three levels of system parameters: low, medium, and high. This embodiment includes five experimental groups, all of which are identical except for the microcapsule preparation and fermentation conditions.

[0152] The common constant process conditions for all groups are as follows: Raw material pretreatment: Take 10 kg of fresh water chestnuts from the same batch, wash them, and mechanically peel them, collecting the pulp and peel separately. Store the water chestnut peels frozen at -18℃. Cut the pulp into 8mm particles.

[0153] Pulsed electric field treatment: The conductivity of the conductive medium is 1.0 mS / cm, and the volume ratio of fruit pulp particles to the conductive medium is 1:2.5. First stage pulsed electric field treatment: electric field strength 18 kV / cm, pulse width 25 μs, pulse frequency 80 Hz, residence time 40 s. Second stage pulsed electric field treatment: electric field strength 30 kV / cm, pulse width 40 μs, pulse frequency 150 Hz, residence time 40 s. The material temperature is controlled below 38℃ throughout the process.

[0154] Material separation and enzymatic hydrolysis: 75% of the treated material was fed into the enzymatic hydrolysis chamber by mass, while 25% was retained as brittle granules and temporarily stored at 2℃. 0.10 parts by mass of cellulase and 0.05 parts by mass of pectinase (based on the dry weight of the pulp) were added to the enzymatic hydrolysis portion, and enzymatic hydrolysis was performed for 3 hours at pH 4.8 and a temperature of 48℃. After enzymatic hydrolysis, no heat inactivation was performed; instead, solid-liquid separation was directly performed (centrifugation at 5000 rpm for 12 minutes) to obtain the enzymatic hydrolysate and solid residue.

[0155] Subcritical water extraction: The mass ratio of water chestnut peel to water was 1:15, the temperature was 135℃, the gauge pressure was 1.2MPa, and the extraction time was 30 min. The extract was then concentrated after filtration through a 0.45μm microporous membrane to obtain a concentrated water chestnut peel polyphenol solution.

[0156] Maillard reaction: The solid residue was placed in a separate reaction vessel, and 0.05 parts by weight of L-cysteine ​​and 0.12 parts by weight of D-xylose were added based on the dry weight of the residue, with a pH of 6.5. The first stage reaction was carried out at 85°C for 40 min, and the second stage reaction was carried out at 110°C for 20 min. After cooling to below 40°C, 4 times the mass of water of the reaction product was added, and water extraction was carried out at 55°C for 30 min. The Maillard flavor extract was obtained by filtration.

[0157] Blending and sterilization: The fermented mash is mixed with refrigerated crispy granules, concentrated water chestnut peel polyphenols, and vitamin E (0.04 parts by weight, based on 100 parts by weight of the total blended sauce). The mixture is then pasteurized at 56°C for 18 minutes.

[0158] Maillard flavor reintroduction and bottling: After sterilization, cool to 38°C, add 2 parts by weight of Maillard flavor extract (based on 100 parts by weight of the total sauce), and stir well. Fill into glass bottles, and perform 3 nitrogen pulse purgings before sealing (purging nitrogen to 0.18 MPa and holding for 5 seconds each time, then venting). Seal the bottles after the third nitrogen purging.

[0159] The variables for each group are set as follows: Experimental Group 6 (Medium System Parameter Level): Chitosan mass-volume concentration 18 g / L. Type A microcapsules: TPP mass fraction 0.8%, core material was *Leuconostoc mesenteriae* activated bacterial solution and *Caragana korshinskii* flavonoid concentrate (volume ratio 10:1), solidified for 25 min. Type B microcapsules: TPP mass fraction 2.5%, core material was *Lactobacillus plantarum* activated bacterial solution and *Caragana korshinskii* flavonoid concentrate (volume ratio 10:1), solidified for 50 min. The fermentation substrate contained 0.5 parts by mass of Type A microcapsules, 0.5 parts by mass of Type B microcapsules, and 2.5 parts by mass of salt, with an initial pH of 6.0. The first stage of fermentation was carried out at 23℃ for 48 h, and the second stage was carried out at 32℃ for 48 h, with the final fermentation pH being 4.0.

[0160] Experimental Group 7 (High Difference + Long Cycle): The difference between this experimental group and Experimental Group 6 is as follows: Type A microcapsules had a TPP mass fraction of 0.5% and a curing time of 20 min; Type B microcapsules had a TPP mass fraction of 3.0% and a curing time of 60 min. The first stage of fermentation was carried out at 22℃ for 72 h, and the second stage was carried out at 30℃ for 72 h. The final fermentation pH was 3.8.

[0161] Experimental Group 8 (Low Difference + Short Cycle): The difference between this experimental group and Experimental Group 6 was: Type A microcapsules had a TPP mass fraction of 1.0% and a curing time of 30 min; Type B microcapsules had a TPP mass fraction of 2.0% and a curing time of 40 min. The first stage of fermentation was carried out at 25℃ for 36 h, and the second stage was carried out at 33℃ for 36 h. The final fermentation pH was 4.2.

[0162] Experimental Group 9 (System-level Control Group): The difference between this experimental group and Experimental Group 6 is that both Type A and Type B microcapsules used 1.5% TPP by mass, and the solidification time was uniformly 35 min. Fermentation was carried out at a constant temperature of 28℃ until the pH dropped to 4.0. This group investigated the superiority of the "differentiated crosslinking density + two-stage variable temperature fermentation" system as a whole over "uniform crosslinking density + constant temperature fermentation".

[0163] Experimental Group 10 (Blank Control Group): The difference between this experimental group and Experimental Group 6 was that microcapsules were not added, but activated bacterial solutions of Leuconostoc mesenteroides and Lactobacillus plantarum were directly added to the fermentation substrate at a volume ratio of 1:1. The fermentation process was carried out at a constant temperature of 28°C until the pH dropped to 4.0. That is, the existing conventional inoculation method was used.

[0164] Example 3: The effect of independent Maillard reaction and post-sterilization re-addition on flavor retention This embodiment focuses on the variable of the amount of Maillard flavor extract added back, examining its effect on the retention of volatile flavor compounds in the finished sauce. The addition temperature was fixed at 38°C, with only the amount added varying. This embodiment includes five experimental groups, all of which are identical except for the amount or method of adding the Maillard flavor extract.

[0165] The common constant process conditions for all groups are as follows: Raw material pretreatment: Take 10 kg of fresh water chestnuts from the same batch, wash them, and mechanically peel them, collecting the pulp and peel separately. Store the water chestnut peels frozen at -18℃. Cut the pulp into 8mm particles.

[0166] Pulsed electric field treatment: The conductivity of the conductive medium is 1.0 mS / cm, and the volume ratio of fruit pulp particles to the conductive medium is 1:2.5. First stage pulsed electric field treatment: electric field strength 18 kV / cm, pulse width 25 μs, pulse frequency 80 Hz, residence time 40 s. Second stage pulsed electric field treatment: electric field strength 30 kV / cm, pulse width 40 μs, pulse frequency 150 Hz, residence time 40 s. The material temperature is controlled below 38℃ throughout the process.

[0167] Material separation and enzymatic hydrolysis: 75% of the treated material was fed into the enzymatic hydrolysis chamber by mass, while 25% was retained as brittle granules and temporarily stored at 2℃. 0.10 parts by mass of cellulase and 0.05 parts by mass of pectinase (based on the dry weight of the pulp) were added to the enzymatic hydrolysis portion, and enzymatic hydrolysis was performed for 3 hours at pH 4.8 and a temperature of 48℃. After enzymatic hydrolysis, no heat inactivation was performed; instead, solid-liquid separation was directly performed (centrifugation at 5000 rpm for 12 minutes) to obtain the enzymatic hydrolysate and solid residue.

[0168] Microcapsule preparation: Chitosan was dissolved in an aqueous acetic acid solution to a concentration of 18 g / L. Type A microcapsules: TPP mass fraction 0.8%, core material was a mixture of *Leuconostoc mesenteriae* activated bacterial solution and *Carex esculenta* flavonoid concentrate at a volume ratio of 10:1, cured for 25 min. Type B microcapsules: TPP mass fraction 2.5%, core material was a mixture of *Lactobacillus plantarum* activated bacterial solution and *Carex esculenta* flavonoid concentrate at a volume ratio of 10:1, cured for 50 min.

[0169] Fermentation: The enzymatic hydrolysate and chopped red pepper were mixed at a mass ratio of 2:1. Based on 100 parts by mass of the total fermentation substrate, 2.5 parts by mass of salt, 0.5 parts by mass of type A microcapsules, and 0.5 parts by mass of type B microcapsules were added, with an initial pH of 6.0. The first stage of fermentation was carried out at 23℃ for 48 hours, and the second stage was carried out at 32℃ for 48 hours. The final fermentation pH was 4.0.

[0170] Subcritical water extraction: The mass ratio of water chestnut peel to water was 1:15, the temperature was 135℃, the gauge pressure was 1.2MPa, and the extraction time was 30 min. The extract was then concentrated after filtration through a 0.45μm microporous membrane to obtain a concentrated water chestnut peel polyphenol solution.

[0171] Maillard reaction conditions: The solid residue was placed in a separate reaction vessel, and 0.05 parts by weight of L-cysteine ​​and 0.12 parts by weight of D-xylose were added based on the dry weight of the residue. The pH was 6.5. The first stage reaction was carried out at 85°C for 40 min, and the second stage reaction was carried out at 110°C for 20 min. After cooling to below 40°C, 4 times the mass of water of the reaction product was added, and water extraction was carried out at 55°C for 30 min. The Maillard flavor extract was obtained by filtration.

[0172] Blending: Fermented mash is mixed with chilled crispy granules, concentrated water chestnut peel polyphenols, and vitamin E (0.04 parts by weight, based on 100 parts by weight of the total blended sauce).

[0173] Sterilization: Pasteurize at 56℃ for 18 minutes.

[0174] The variables for each group are set as follows: Experimental Group 11 (2 parts by weight): The Maillard reaction was carried out in an independent reactor (completely isolated from the fermentation system). After sterilization, the temperature was lowered to 38°C, and 2 parts by weight of Maillard flavor extract (based on 100 parts by weight of the total weight of the sauce) were aseptically added. After stirring evenly, the mixture was filled into glass bottles. Before sealing, nitrogen pulse purging was performed 3 times (nitrogen was purged to 0.18 MPa and held for 5 seconds each time before venting). The bottles were sealed after the third nitrogen purging.

[0175] Experimental Group 12 (refilled with 1 part by mass): The difference between this experimental group and Experimental Group 11 is that the amount of Maillard flavor extract refilled is 1 part by mass (based on a total mass of 100 parts by mass of the prepared sauce), and the refilling temperature is also 38℃. The filling operation is the same as that of Experimental Group 11.

[0176] Experimental Group 13 (3 parts by weight): The difference between this experimental group and Experimental Group 11 is that the amount of Maillard flavor extract added is 3 parts by weight (based on a total mass of 100 parts by weight of the prepared sauce), and the addition temperature is also 38℃. The filling operation is the same as that of Experimental Group 11.

[0177] Experimental Group 14 (with control group added before sterilization): The difference between this experimental group and Experimental Group 11 is that Maillard flavor extract was added during the preparation stage (2 parts by weight), and mixed with fermented mash, crispy granules, polyphenol concentrate, and vitamin E at the same time. Then, they were pasteurized together at 56℃ for 18 minutes. After sterilization, the temperature was lowered to 38℃ and directly filled into glass bottles. Before sealing, nitrogen pulse replacement was performed 3 times (nitrogen was purged to 0.18MPa and held for 5 seconds each time before venting). The bottles were sealed after the third nitrogen purging.

[0178] Experimental Group 15 (Blank Control Group): The difference between this experimental group and Experimental Group 11 is that the Maillard reaction was not performed independently. Instead, the Maillard reaction substrates (solid residue, L-cysteine, D-xylose) were directly added to the fermentation system and carried out simultaneously with the fermentation. After fermentation, they underwent preparation and sterilization together. After sterilization, the temperature was lowered to 38°C, and the mixture was directly filled into glass bottles. Before sealing, three nitrogen pulse purgings were performed (each purging to 0.18 MPa and holding for 5 seconds followed by venting). The bottles were sealed after the third nitrogen purging. This adopts the existing conventional method of simultaneous Maillard reaction within the fermentation system.

[0179] Experimental data recording Table 1. Example 1: Test results of the textural properties of the finished sauce-flavored water chestnut crisps. Fresh sample standard 28.63 18.21 0.72 14.82 — Experimental Group 1 (Moderate Intensity) 24.27 15.08 0.69 11.43 84.8 Experimental Group 2 (Low Intensity) 25.35 15.92 0.67 11.78 88.6 Experimental Group 3 (High Intensity) 22.81 13.85 0.64 9.87 79.7 Experimental group 4 (hot burn control) 14.52 7.41 0.51 4.92 50.7 Experimental Group 5 (Comprehensive Control) 13.76 6.65 0.48 4.31 48.1 Experimental group 2 showed the highest hardness retention rate (88.6%), but its elasticity (0.67) was slightly lower than that of experimental group 1 (0.69). This indicates that although the low field strength treatment caused the least damage to the cell wall structure and maintained the best hardness, the degree of reversible perforation was less, resulting in a slightly lower tissue recovery ability compared to the medium field strength treatment. Under the conditions of high field strength and long residence time, experimental group 3 showed increased cell wall damage, and both hardness and chewiness decreased.

[0180] Table 2. Example 2: Results of Detection of Flavor Components and Fermentation Stability of Finished Sauce Experimental Group 6 (Medium Level) 1856 412 8.6 2.3 3.74 0.09 1.3×10³ Experimental group 7 (high difference + long period) 1923 398 9.1 2.1 4.33 0.11 1.6×10³ Experimental group 8 (low difference + short cycle) 1694 372 8.2 2.5 3.28 0.14 2.1×10³ Experimental group 9 (uniform cross-linking + isothermal) 1108 359 5.4 1.6 3.38 0.23 4.8×10³ Experimental group 10 (blank control) 876 336 4.8 3.5 1.37 0.35 7.9×10³ The total ester content in experimental group 7 (1923 μg / kg) was higher than that in experimental group 6 (1856 μg / kg). This was because the larger difference in TPP concentration (0.5% / 3.0%) resulted in a greater difference in the swelling pH threshold between type A and type B microcapsules, a clearer cascade release sequence, and a longer independent metabolic window for Leuconostoc mesenteroides to synthesize esters. However, the longer fermentation period in experimental group 7 (72h+72h) also led to an increase in the accumulation of byproducts in the system, and the pH stability (variation range 0.11) was slightly worse than that in experimental group 6 (0.09), reflecting the engineering balance between fermentation period and flavor accumulation.

[0181] Table 3. Example 3: Results of Tests on Retention of Volatile Flavor Compounds and Storage Stability of Finished Sauce Experimental group 11 2 412 285 0.041 3.2 Experimental group 12 1 357 231 0.048 3.9 Experimental group 13 3 443 312 0.039 2.7 Experimental group 14 (added before sterilization) 2 268 158 0.061 4.6 Experimental group 15 (blank control) — 195 108 0.078 6.3 The refill temperature for experimental groups 11-13 was fixed at 38℃. The content of sulfur-containing heterocyclic compounds and furans increased with the increase of refill amount, but the relationship was not strictly linear. The content of sulfur-containing heterocyclic compounds in experimental group 14 (added before sterilization) (268 μg / kg) was significantly lower than that in experimental group 11 (412 μg / kg) with the same refill amount, a decrease of about 35%, indicating that the 56℃ pasteurization process caused significant heat loss to low-boiling-point sulfur-containing heterocyclic compounds.

[0182] III. Summary of Experimental Results The following key information can be intuitively obtained from the above experimental data: Regarding texture preservation (Example 1): In experimental groups 1 to 3, which were treated with a two-stage pulsed electric field, the hardness retention rate of water chestnut crunch particles in the finished sauce ranged from 79.7% to 88.6%, significantly better than the blanched control group (experimental group 4, 50.7%) and the comprehensive control group (experimental group 5, 48.1%). Experimental group 2 (low treatment intensity) showed the best performance in terms of hardness retention rate (88.6%), but its elasticity index (0.67) was slightly lower than that of experimental group 1 (0.69). This indicates that the low field strength treatment is better in protecting the integrity of the cell wall structure, but slightly inferior to the medium field strength treatment in maintaining tissue recovery ability, reflecting the engineering balance between treatment intensity and various dimensions of texture.

[0183] Regarding flavor construction (Example 2): The total ester content of experimental group 7 (1923 μg / kg) exceeded that of experimental group 6 (1856 μg / kg), but at the cost of a longer fermentation period leading to slightly poorer pH stability (variation range 0.11 vs 0.09) and a slightly higher total bacterial count (1.6 × 10³ vs 1.3 × 10³ CFU / g), resulting in a reasonable performance trade-off. The total ester content of all three experimental groups (6 / 7 / 8) was above 1694 μg / kg, significantly better than the uniform cross-linked isothermal control group (experimental group 9, 1108 μg / kg) and the blank control group (experimental group 10, 876 μg / kg).

[0184] Regarding flavor retention (Example 3): In experimental groups 11 to 13, which were added after sterilization and cooling, the total amount of sulfur-containing heterocyclic compounds was between 357 and 443 μg / kg, significantly better than that of experimental group 14 (268 μg / kg) added before sterilization and the blank control group (experimental group 15, 195 μg / kg). When the amount added increased from 1 part by mass to 3 parts by mass, the content of flavor substances showed an upward trend, but it was not strictly linear. The ratio of furans to sulfur-containing heterocyclic compounds fluctuated reasonably among different groups.

[0185] Based on the recording and organization of the above experimental data, the following analysis and mechanism speculation on the performance difference trends of each experimental group from the perspective of food physicochemistry and molecular biology are conducted in depth.

[0186] Effects of pulsed electric field treatment on the texture of water chestnut crisps: The two-stage treatment group outperformed the blanching and comprehensive control groups in terms of hardness and brittleness, and the results showed a non-monotonic change with treatment intensity. The high temperature of blanching damages the pectin and hemicellulose structure, leading to tissue softening. The pulsed electric field, through two-stage field intensity control, maintains cell membrane permeability and preserves the integrity of the cell wall structure at low temperatures. Different treatment intensities affect texture: low intensity maintains good hardness but reduces elasticity; medium intensity shows excellent elasticity; and high intensity reduces both hardness and chewiness. This indicates that a reasonable match between field intensity and residence time is needed to balance permeability and structural preservation.

[0187] The effect of differentiated cross-linking density microcapsules combined with two-stage temperature-switched fermentation on flavor: This treatment group outperformed the control group in total esters, organic acid metabolism, and storage stability, with trade-offs among different parameters. The mechanism lies in matching the metabolic rhythm of the microbial species: *Leuconostoc mesenteroides* produces esters in the early stage of fermentation; *Lactobacillus plantarum* produces acid and lowers pH in the middle and late stages. Differentiated cross-linking microcapsules control the release sequence through different cross-linking densities: low-cross-linked microcapsules release *Leuconostoc mesenteroides* in the early stage, while high-cross-linked microcapsules release *Lactobacillus plantarum* in the later stage. The two-stage temperature-switched fermentation further optimizes metabolic matching, thereby improving flavor and stability. High-concentration sodium tripolyphosphate cross-linking makes the network denser, requiring the pH to drop to a lower threshold to release *Lactobacillus plantarum*. This pH-responsive cascade design allows *Leuconostoc mesenteroides* to obtain an independent metabolic window in the early stage of fermentation, avoiding inhibition caused by a sudden drop in pH due to acid production by *Lactobacillus plantarum*, thus allowing sufficient time for ester accumulation. Two-stage temperature control enhances the timing matching: the first stage of low temperature is conducive to the expression of Leuconostoc mesenteroides esterase, and the second stage of high temperature activates the Lactobacillus plantarum enzyme system, making it tolerant to low pH and produce acid efficiently.

[0188] The high-variability, long-cycle group showed a significant increase in total ester content, but slightly lower pH stability and microbial indicators. This is because the greater difference in cross-linking density resulted in a larger difference in the pH threshold for microcapsule swelling, allowing Leuconostoc mesenteroides to synthesize esters through a longer independent metabolic window. However, the longer fermentation cycle increased the accumulation of metabolic byproducts, affecting pH buffer stability, and increasing the chance of contamination by other microorganisms, leading to a slight increase in the total colony count.

[0189] The control group, fermented at a uniform cross-linking density under isothermal conditions, significantly lagged behind in ester and organic acid metabolism. Uniform cross-linking allowed both microcapsules to release microorganisms simultaneously at similar pH levels, leading to competition between the strains. *Lactobacillus plantarum* rapidly produced acid and gained dominance, causing a rapid pH drop and inhibiting the esterase activity of *Leuconostoc mesenteroides*, thus interrupting ester synthesis. Isothermal fermentation could not provide the optimal temperatures for each microorganism, exacerbating the metabolic imbalance.

[0190] The blank control group, directly inoculated without microcapsule encapsulation, performed the worst. Free bacterial cells lacked the pH buffering and controlled release provided by chitosan wall material; both bacteria initiated metabolism almost simultaneously, resulting in the most intense competition and a compressed ester synthesis window. Furthermore, the antibacterial and antioxidant benefits of chitosan were absent.

[0191] III. The impact of independent Maillard reaction and post-sterilization re-addition on flavor retention The experimental group that was cooled and refilled after sterilization had better retention of sulfur-containing heterocyclic compounds and furan compounds than the control group that was added before sterilization, and the amount of refilled compounds was positively correlated with the amount of retained compounds in a non-linear manner.

[0192] The difference stems from the contradiction between the volatility characteristics of key flavor components in the Maillard reaction and heat treatment. Flavor substances such as sulfur-containing heterocyclic compounds have small molecular weights and low boiling points, making them easily volatile at pasteurization temperatures. Continuous heating during pasteurization causes dissolved components to transfer to the gas phase and dissipate. Cooling and refilling after pasteurization avoids secondary heat loss, allowing flavor substances to be retained at near-original concentrations.

[0193] The control group, added before sterilization, underwent simultaneous pasteurization, resulting in the loss of some low-boiling-point flavors and lower detection values ​​compared to the re-added group. The blank control group had the reaction substrate directly added to the fermentation system, where metabolites may cross-react with Maillard intermediates. At lower re-added amounts, flavor compounds were fully dissolved in the aqueous phase of the sauce; at higher re-added amounts, hydrophobic components approached dissolution equilibrium, with excess components existing as microemulsions or micelles, leading to a non-linear correlation between the increase in detection values ​​and the amount added. Higher re-added amounts also introduced more reducing Maillard intermediates, whose antioxidant activity could inhibit lipid peroxidation during storage, improving peroxide value and color stability.

[0194] Pulsed electric field treatment protects the granular texture of water chestnut crisps and promotes the transformation of enzymatic substrates by increasing cell permeability, providing a more abundant carbon and nitrogen source for fermentation. Oligosaccharides produced by the degradation of chitosan microcapsule wall material during fermentation have antibacterial properties and interact with Maillard products to form a sustained-release flavor system. The Maillard extract added after sterilization contains antioxidant components, which, together with water chestnut peel polyphenols and vitamin E, form an antioxidant network, synergistically inhibiting lipid peroxidation and browning, thus exhibiting comprehensive advantages in peroxide value and color stability. This synergistic effect of multiple technologies is the core embodiment of the overall integrity of this invention.

[0195] All unit operations of this invention are based on existing food processing equipment, requiring no laboratory setup. The pulsed electric field treatment uses a commercially available continuous system, with materials continuously passing through the processing chamber; chitosan microcapsule preparation employs ionogelation, using a conventional stirred reactor and drop-forming device; subcritical water extraction utilizes existing pressurized equipment; the Maillard reaction uses a standard jacketed reactor; and fermentation uses a conventional temperature-controlled tank. While the single-batch cycle of this process is slightly longer, the microencapsulation cascade fermentation reduces strain competition losses and improves conversion efficiency; post-sterilization refilling reduces flavor loss and lowers the amount of additional flavorings required. Overall, the process demonstrates industrial feasibility and economic viability in terms of cost and quality.

[0196] The technical features described above can be combined in any way. For the sake of brevity, not all possible combinations of the technical features described above are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

Claims

1. A production process for water chestnut and red pepper sauce, characterized in that: Includes the following steps: Peel the water chestnuts, collect the pulp and water chestnut peel separately, and cut the pulp into small pieces; The pulp particles were immersed in a conductive medium and subjected to a two-stage pulsed electric field treatment. The electric field strength of the first stage pulsed electric field treatment was 15~20kV / cm, and the electric field strength of the second stage pulsed electric field treatment was 25~35kV / cm. The material temperature was ≤40℃ throughout the pulsed electric field treatment. The material treated by the pulsed electric field is divided into two parts. 70-80% by mass enters the enzymatic hydrolysis process, and 20-30% is retained as fruit pulp particles and temporarily stored at 0-4℃. Cellulase and pectinase are added to the material entering the enzymatic hydrolysis process, and enzymatic hydrolysis is carried out under the conditions of pH 4.5~5.0 and temperature 45~50℃. No heat inactivation is performed after the enzymatic hydrolysis is completed. The enzymatic hydrolysis products are subjected to solid-liquid separation. The resulting liquid phase is used as the enzymatic hydrolysate and enters the fermentation process, while the resulting solid residue is entered into the Maillard reaction process. Using chitosan as the wall material and sodium tripolyphosphate as the crosslinking agent, type A microcapsules and type B microcapsules were prepared by ionic crosslinking. The sodium tripolyphosphate aqueous solution used to prepare type A microcapsules had a sodium tripolyphosphate mass fraction of 0.5-1.0%, and the core material included Leuconostoc mesenteroides and flavonoids from water chestnuts. The sodium tripolyphosphate aqueous solution used to prepare type B microcapsules had a sodium tripolyphosphate mass fraction of 2.0-3.0%, and the core material included Lactobacillus plantarum and flavonoids from water chestnuts. The enzymatic hydrolysate was mixed with chopped red pepper, edible salt, type A microcapsules and type B microcapsules to construct a fermentation substrate. Two-stage fermentation was carried out. The first stage fermentation temperature was 22~25℃. When the pH of the fermentation substrate was ≤5.0, the fermentation was transferred to the second stage. The second stage fermentation temperature was 30~33℃. Fermentation was stopped when the pH of the fermentation substrate dropped to 3.8~4.2 to obtain fermented mash. Water chestnut peel was extracted with subcritical water at a temperature of 120-150℃ and a gauge pressure of 0.8-1.5MPa. The resulting extract was filtered and concentrated to obtain a concentrated water chestnut peel polyphenol solution. L-cysteine ​​and D-xylose were added to the solid residue in an independent reaction vessel, and the Maillard reaction was carried out in two stages. The temperature of the first stage was 80~90℃, and the temperature of the second stage was 105~115℃. The reaction product was cooled to ≤40℃ and then extracted with water to obtain Maillard flavor extract. The fermented mash is mixed with temporarily refrigerated fruit pulp particles, concentrated water chestnut peel polyphenols, and vitamin E. The prepared materials are pasteurized at a temperature of 55-58℃ for 15-20 minutes. After sterilization, Maillard flavor extract is added, and the product is filled. Before sealing, the headspace of the packaging container is purged with nitrogen via a pulse.

2. The production process as described in claim 1, characterized in that: The fruit pulp is cut into particles with a diameter of 5-10 mm, and the water chestnut skin is frozen and stored at ≤-18℃.

3. The production process as described in claim 1, characterized in that: The conductivity of the conductive medium is 0.3~1.5 mS / cm, the volume ratio of fruit pulp particles to conductive medium is 1:2~1:3, the pulse width of the first pulse electric field treatment is 20~30μs, the pulse frequency is 50~100Hz, and the residence time is 20~60s, and the pulse width of the second pulse electric field treatment is 30~50μs, the pulse frequency is 100~200Hz, and the residence time is 20~60s.

4. The production process as described in claim 1, characterized in that: Based on the dry weight of the pulp, the amount of cellulase added is 0.05~0.15 parts by weight, the amount of pectinase added is 0.02~0.08 parts by weight, the enzymatic hydrolysis time is 2~4 hours, and the solid-liquid separation is carried out by centrifugation or pressure filtration, with a centrifugation speed of 4000~6000 rpm and a centrifugation time of 10~15 minutes, and a pressure filtration pressure of 0.2~0.3 MPa.

5. The production process as described in claim 1, characterized in that: The chitosan is dissolved in an aqueous acetic acid solution, and the chitosan mass-volume concentration is 15-20 g / L. The volume ratio of *Leuconostoc mesenteriae* activated bacterial solution to *Carex esculenta* flavonoid concentrate in the core material of the type A microcapsules is 10:

1. The volume ratio of *Lactobacillus plantarum* activated bacterial solution to *Carex esculenta* flavonoid concentrate in the core material of the type B microcapsules is 10:

1. The curing time for preparing type A microcapsules is 20-30 min, and the curing time for preparing type B microcapsules is 40-60 min.

6. The production process as described in claim 1, characterized in that: The mass ratio of the enzymatic hydrolysate to the chopped red pepper is 1.5:1 to 2.5:

1. Based on 100 parts by mass of the total fermentation substrate, the amount of added edible salt is 1.5 to 4.0 parts by mass, the amount of added type A microcapsules is 0.3 to 0.8 parts by mass, the amount of added type B microcapsules is 0.3 to 0.8 parts by mass, the initial pH of the fermentation substrate is 5.5 to 6.5, the fermentation time of the first stage is 36 to 72 hours, and the fermentation time of the second stage is 36 to 72 hours.

7. The production process as described in claim 1, characterized in that: In the subcritical water extraction, the mass ratio of water chestnut peel to water is 1:10 to 1:20, the extraction time is 20 to 40 minutes, and the filtration uses a microporous membrane with a pore size of 0.45 μm.

8. The production process as described in claim 1, characterized in that: Based on the dry weight of the solid residue, the amount of L-cysteine ​​added is 0.02~0.08 parts by mass, the amount of D-xylose added is 0.05~0.20 parts by mass, the pH of the Maillard reaction is 6.0~7.0, the reaction time of the first stage is 30~45 min, the reaction time of the second stage is 15~30 min, the amount of water added during water extraction is 3~5 times the mass of the reaction product, the water extraction temperature is 50~60℃, and the water extraction time is 30 min. Based on a total mass of 100 parts by mass of the blended sauce, the amount of vitamin E added is 0.02~0.06 parts by mass, and the amount of Maillard flavor extract added is 1~3 parts by mass.

9. The production process as described in claim 1, characterized in that: The nitrogen pulse replacement is performed 3 times. Each time, nitrogen is injected until the headspace pressure is 0.15~0.20MPa and held for 5 seconds before being vented. The container is sealed immediately after the third nitrogen injection. The residual oxygen content in the headspace of the packaging container after replacement is ≤0.5%.

10. A water chestnut and red pepper sauce, prepared using the production process of a water chestnut and red pepper sauce according to any one of claims 1 to 9, characterized in that: The water chestnut and red pepper sauce has a pH of 3.8 to 4.2, a salt content of ≤4%, and a headspace residual oxygen content of ≤0.5% in the packaging container.