A method for extracting pea starch by the immobilized cell method

Abstract

The present application provides a method for extracting pea starch by the immobilized cell method, which relates to the technical field of pea starch extraction and includes the following steps: peeling pea grains and then grinding them into powder to obtain pea flour, adding process water to the pea flour, and stirring evenly to obtain pea milk; filtering the soybean dregs in the pea milk to obtain filtered pea milk, and homogenizing the filtered pea milk to obtain pea flour slurry; adding the compound strain gel beads to the pea flour slurry for fermentation, recovering the compound strain gel beads and the supernatant, obtaining a precipitate, and drying the precipitate to obtain pea starch; the compound strain gel beads include an immobilized carrier and a compound strain encapsulated in the immobilized carrier, the immobilized carrier is a biocompatible polysaccharide gel, and the compound strain is composed of Lactobacillus and Streptococcus thermophilus. It solves the bottleneck problems such as strain waste, insufficient purity and poor environmental protection in the prior art, and provides a high-benefit and low-pollution extraction scheme for the industrial production of pea starch.

Description

Technical Field

[0001] This application relates to the technical field of pea starch extraction, and specifically relates to a method for extracting pea starch using the immobilized cell method. Background Art

[0002] Peas are rich in high-quality protein and carotene, which can improve the body's disease resistance and recovery ability. Pea starch is a high-quality starch. Compared with other starch products, the pea starch products prepared by hot processing have a lower GI value, which can enhance the body's immune function, prevent the synthesis of high blood sugar in the human body, and reduce the incidence of chronic diseases in the human body.

[0003] In the prior art, such as the invention patent with the publication number CN115322263A, a method for extracting high-quality pea starch based on compound strain fermentation is proposed, which overcomes the problems of large water consumption, high cost, and low extraction rate in the traditional acid pulp method and water extraction method for starch extraction. However, there are still the following defects: First, directly adding free strains (Lactobacillus, Streptococcus thermophilus, and maltodextrin) results in the inability to recycle the strains, and new strains need to be added for each batch, significantly increasing the production cost. Second, the residual bacteria mixed into the starch require additional separation steps, reducing the extraction efficiency; after fermentation, the bacteria and starch are mixed, and it is difficult to completely remove the residual bacteria by centrifugal separation, resulting in a decrease in starch purity; third, maltodextrin, as an auxiliary material for compounding strains, has a relatively large molecular weight and may remain in the starch, reducing the product light transmittance and affecting the starch processing performance. Summary of the Invention

[0004] In view of the above defects or deficiencies in the prior art, this application aims to provide a method for extracting pea starch using the immobilized cell method, which solves the problems of strain waste and insufficient purity;

[0005] This method includes the following steps:

[0006] Pea grains are peeled and ground into pea powder. Process water is added to the pea powder and stirred evenly to obtain pea milk.

[0007] The soybean residue in the pea milk is filtered to obtain filtered soybean milk, and the filtered soybean milk is homogenized to obtain pea flour slurry.

[0008] Compound strain gel beads are obtained, and the compound strain gel beads are added to the pea flour slurry for fermentation and stirred evenly to obtain a fermentation broth.

[0009] The compound strain gel beads in the fermentation broth are recovered, and the supernatant is recovered to obtain a precipitate; the supernatant is used to prepare the process water for the next extraction.

[0010] The precipitate is dried to obtain pea starch.

[0011] Among them, the compound strain gel beads include an immobilization carrier and compound strains encapsulated in the immobilization carrier. The immobilization carrier is a biocompatible polysaccharide gel, and the compound strains are composed of Lactobacillus and Streptococcus thermophilus.

[0012] According to the technical solution provided by this application, the types of the compound strain gel beads include newly prepared compound strain gel beads with zero cumulative usage times and recycled compound strain gel beads with at least one cumulative usage time.

[0013] Preparing the newly prepared compound strain gel beads includes the following steps:

[0014] Take Lactobacillus powder and Streptococcus thermophilus powder, mix them according to a mass ratio of 40%-48%:40%-48% to obtain a compound powder, and the other proportioning substance is maltodextrin.

[0015] Prepare a 1.5% sodium alginate solution, heat and stir to dissolve it, and cool it to room temperature.

[0016] Add the compound powder into the sodium alginate solution, and the mass ratio of the compound powder to sodium alginate is 1:0.5 - 1:5. Stir until evenly suspended to obtain a mixed solution.

[0017] Drop the mixed solution into a 2%-5% calcium chloride solution drop by drop, and let it stand at room temperature for crosslinking to form initial gel beads.

[0018] Rinse the initial gel beads with sterile water multiple times to remove the residual calcium ions on the surface to obtain newly prepared compound strain gel beads.

[0019] According to the technical solution provided by this application, this method further includes the following steps:

[0020] When using the compound strain gel beads for fermentation each time, monitor the lactic acid concentration in the fermentation broth in real time to calculate the current lactic acid production rate during the current fermentation process.

[0021] Record the cumulative usage times of the compound strain gel beads and the corresponding current lactic acid production rate.

[0022] Use the cumulative usage times as the abscissa and the current lactic acid production rate as the ordinate to fit and generate a lactic acid production rate decay curve.

[0023] Based on the lactic acid production rate decay curve and the cumulative usage times, generate a gel bead label, and the gel bead label is used to dynamically adjust the fermentation parameters during subsequent fermentation processes; the fermentation parameters include stirring frequency, fermentation temperature, and fermentation duration.

[0024] According to the technical solution provided by the present application, generating a gel bead label based on the lactic acid production rate decay curve and the cumulative number of uses includes the following steps:

[0025] Determine the activity level according to the percentage of the current lactic acid production rate to the initial lactic acid production rate when the compound strain gel beads are first used;

[0026] Determine the loss level according to the cumulative number of uses;

[0027] Use the combination of the activity level and the loss level as the gel bead label.

[0028] According to the technical solution provided by the present application, the activity level includes high activity, medium activity, and low activity, and the loss level includes high loss, medium loss, and low loss;

[0029] After recovering the compound strain gel beads in the fermentation broth, the following steps are further included:

[0030] If the activity level of the compound strain gel beads is low activity and the loss level is high loss, then the compound strain gel beads shall be compulsorily scrapped or downgraded for use after regeneration.

[0031] According to the technical solution provided by the present application, adding the compound strain gel beads to the pea starch slurry for fermentation includes the following steps:

[0032] If the activity level of the compound strain gel beads is medium activity and the loss level is medium loss, then add the compound strain gel beads to the pea starch slurry, and dynamically supplement carbon sources or coenzyme factors according to the real-time lactic acid production rate.

[0033] According to the technical solution provided by the present application, obtaining the compound strain gel beads includes the following steps:

[0034] Obtain the target process requirements for the current extraction, and the target process requirements include the target starch purity of the starch to be extracted;

[0035] If the target starch purity is greater than the first preset purity, then select the newly prepared compound strain gel beads.

[0036] According to the technical solution provided by the present application, after obtaining the target process requirements for the current extraction, the following steps are further included:

[0037] If the target starch purity is less than or equal to the first preset purity, then select the corresponding recycled compound strain gel beads according to the target starch purity.

[0038] According to the technical solution provided by the present application, the step of selecting the corresponding recovered compound strain gel beads according to the target starch purity includes the following steps:

[0039] If the target starch purity is less than the second preset purity, select the recovered compound strain gel beads with a gel bead label of low activity - high loss or a cumulative use times of the first preset times; if the target starch purity is greater than the second preset purity and less than the first preset purity, select the recovered compound strain gel beads with a gel bead label of medium activity - medium loss and a cumulative use times less than the second preset times;

[0040] Wherein, the second preset purity is less than the first preset purity, and the first preset times is greater than the second preset times.

[0041] According to the technical solution provided by the present application, the method further includes regenerating the compound strain gel beads with a low activity level and a high loss level.

[0042] The step of regenerating the compound strain gel beads with a low activity level and a high loss level includes the following steps:

[0043] Soak the compound strain gel beads in a regenerating solution of sodium alginate, and apply high-pressure homogenization treatment to repair the surface cracks of the compound strain gel beads to obtain repaired gel beads;

[0044] Transfer the repaired gel beads to an activation solution, and perform shaking culture to restore the metabolic activity of the strains to obtain activated gel beads; the activation solution is a phosphate buffer solution containing glucose, yeast extract, and magnesium sulfate;

[0045] Detect the lactic acid production rate of the activated gel beads. If it reaches the first proportion interval of the initial lactic acid production rate, label the gel bead label of the activated gel beads as regenerated medium activity - medium loss, and limit its cumulative use times.

[0046] Compared with the prior art, the beneficial effects of the present application are as follows: By encapsulating the compound bacteria (Lactobacillus and Streptococcus thermophilus) in biocompatible polysaccharide gel beads and recovering the gel beads by physical filtration after fermentation, repeated recycling can be achieved, reducing the cost of the bacteria and avoiding the mixing of bacteria into the starch. At the same time, the homogenization treatment makes the pea slurry particles uniform, and combined with the control of the gel bead size, physical isolation of the bacteria and the substrate is achieved, reducing the residual amount of bacteria and improving the starch purity. In addition, using a gel carrier instead of maltodextrin and removing the auxiliary components (such as sodium alginate) by rinsing with sterile water can improve the light transmittance of the starch and meet the requirements of high-end food processing. Finally, the supernatant is recycled as the process water for preparing pea milk, further reducing the water consumption in the extraction process. Considering the above aspects, the bottleneck problems in the prior art such as waste of bacteria, insufficient purity and poor environmental protection are solved, providing an extraction scheme with high efficiency and low pollution for the industrial production of pea starch. BRIEF DESCRIPTION OF THE DRAWINGS

[0047] Figure 1 It is a flowchart of the steps of the method for extracting pea starch using the immobilized cell method provided by the present application. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0048] The present application will be further described in detail below with reference to the drawings and embodiments. It can be understood that the specific embodiments described herein are only used to explain the related invention, rather than limiting the invention. In addition, it should be noted that only the parts related to the invention are shown in the drawings for the sake of convenience of description.

[0049] It should be noted that, without conflict, the embodiments in the present application and the features in the embodiments can be combined with each other. The present application will be described in detail below with reference to the drawings and embodiments.

[0050] Example 1

[0051] As mentioned in the background art, in view of the problems in the prior art, the present application proposes a method for extracting pea starch using the immobilized cell method, as Figure 1 shown, which includes the following steps:

[0052] S1. Peel the pea grains and grind them into powder to obtain pea powder, add process water to the pea powder, and stir evenly to obtain pea milk.

[0053] S2. Filter the soybean residue in the pea milk to obtain filtered soybean milk, and homogenize the filtered soybean milk to obtain pea slurry.

[0054] S3. Obtain compound bacteria gel beads, and add the compound bacteria gel beads to the pea slurry for fermentation, and stir evenly to obtain a fermentation broth.

[0055] S4. Recover the compound strain gel beads in the fermentation broth and recover the supernatant to obtain a precipitate; the supernatant is used to prepare the process water for the next extraction.

[0056] S5. Dry the precipitate to obtain pea starch.

[0057] Among them, the compound strain gel beads include an immobilized carrier and a compound strain encapsulated in the immobilized carrier. The immobilized carrier is a biocompatible polysaccharide gel, and the compound strain is composed of Lactobacillus and Streptococcus thermophilus.

[0058] Further, the types of the compound strain gel beads include newly prepared compound strain gel beads with a cumulative use times of zero and recycled compound strain gel beads with a cumulative use times of at least one.

[0059] Preparing the newly prepared compound strain gel beads includes the following steps:

[0060] Take Lactobacillus powder and Streptococcus thermophilus powder, mix them according to a mass ratio of 40%-48%:40%-48% to obtain a compound powder, and other proportioning substances are maltodextrin.

[0061] Prepare a 1.5% sodium alginate solution, heat and stir to dissolve it, and cool it to room temperature.

[0062] Add the compound powder into the sodium alginate solution, and the mass ratio of the compound powder to sodium alginate is 1:0.5 - 1:5. Stir until evenly suspended to obtain a mixture.

[0063] Drop the mixture into a 2%-5% calcium chloride solution drop by drop, and let it stand at room temperature for crosslinking to form initial gel beads.

[0064] Rinse the initial gel beads with sterile water multiple times to remove the residual calcium ions on the surface to obtain newly prepared compound strain gel beads.

[0065] Specifically, the experimental data of the optimization of the preparation parameters of the newly prepared compound strain gel beads are shown in Table 1 as follows:

[0066] Table 1

[0067]

[0068] As can be seen from Table 1, the viable cell rate (92%) and the initial lactic acid production rate (1.82 mmol / (L·h)) of experimental group B (44%:44%) were higher than those of other groups (A: 88%, C: 89%), indicating that the equal proportion mixture (±44%) was the optimal ratio. Too high a proportion of bacterial powder (experimental group C, 48%:48%) led to competitive inhibition of the strains and a decrease in the metabolic rate (1.78 mmol / (L·h)). Experimental group B (1.5%) achieved a balance among mechanical strength (0.28 N / pearl), viable cell rate (92%) and metabolic rate (1.82 mmol / (L·h)). Too high a concentration (experimental group D, 2.0%) led to a decrease in porosity and limited activity of the strains (viable cell rate 85%, rate 1.65 mmol / (L·h)), while too low a concentration (experimental group E, 1.0%) resulted in insufficient mechanical strength (0.20 N / pearl) and easy breakage. The following Examples 2-6 are all different implementation manners of Example 1;

[0069] Example 2

[0070] (1) Peeling and grinding: Accurately weigh 1000 kg of pea grains, peel them with a peeling machine and then grind them to obtain 918 kg of pea flour;

[0071] (2) Preparing pea milk: Add water to the obtained 918 kg of pea flour, with the ratio of pea flour to water being 1:4, and stir evenly to obtain pea milk;

[0072] (3) Preparing compound strain gel beads for immobilized cells: The mass ratio of lactic acid bacteria powder to pea flour is 1:150. The bacterial powder contains: 50% Lactobacillus bulgaricus and 50% Streptococcus thermophilus. Add 100 ml of the lactic acid bacteria powder to 1.5% sodium alginate solution, and drip it into 200 ml of 2% calcium chloride solution to form gel beads.

[0073] (4) Removing residue: Use a gauze filter to remove soybean dregs, and repeat 2-5 times to remove soybean dregs as much as possible;

[0074] (5) Fine grinding: Further process the pea flour slurry with a homogenizer for 5 min, with a mesh number of 60 meshes.

[0075] (6) Fermentation: Add the compound strain gel beads to the slurry, stir evenly and then carry out fermentation. The fermentation temperature is 37°C and the fermentation time is 8 h. First stir for 1 h and let it stand for 3 h, then stir for 1 h and let it stand for 1 h until the fermentation ends;

[0076] (7) Separation: Filter the compound strain gel beads in the fermentation broth through a gauze filter to retain them for the next fermentation, remove the supernatant and centrifuge to collect the precipitate;

[0077] (8) Drying: Dry the precipitate with hot air at 40°C to obtain 297.3 kg of pea starch.

[0078] Example 3

[0079] (1)Peeling and grinding: Accurately weigh 1000 kg of pea grains, peel them with a peeling machine and then grind them to obtain 918 kg of pea flour;

[0080] (2)Preparing pea milk: Add water to the obtained 918 kg of pea flour, with a ratio of pea flour to water of 1:6, and stir evenly to obtain pea milk;

[0081] (3)Preparing the compound strain gel beads of immobilized cells: The mass ratio of lactic acid bacteria powder to pea flour is 1:150. The powder contains: 50% Lactobacillus bulgaricus and 50% Streptococcus thermophilus. Add 100 ml of the lactic acid bacteria powder to 1.5% polysaccharide solution, and drop it into 200 ml of 2% calcium chloride solution to form gel beads.

[0082] (4)Removing residue: Use a gauze filter to remove bean dregs, and repeat 2 - 5 times to remove as much bean dregs as possible;

[0083] (5)Fine grinding: Use a homogenizer to further process the pea flour slurry for 5 minutes, with a mesh size of 60.

[0084] (6)Fermentation: Add the compound strain gel beads of immobilized cells to the slurry, stir evenly and then carry out fermentation. The fermentation temperature is 37 °C and the fermentation time is 8 h. Stir for 1 h and let it stand for 3 h first, then stir for 1 h and let it stand for 1 h until the fermentation ends;

[0085] (7)Separation: Filter the lactic acid bacteria gel beads in the fermentation broth through a gauze filter and retain them for the next fermentation, remove the supernatant and centrifuge to collect the precipitate;

[0086] (8)Drying: Dry the precipitate with hot air at 40 °C to obtain 277.3 kg of pea starch.

[0087] Example 4

[0088] (1)Peeling and grinding: Accurately weigh 1000 kg of pea grains, peel them with a peeling machine and then grind them to obtain 918 kg of pea flour;

[0089] (2)Preparing pea milk: Add water to the obtained 918 kg of pea flour, with a ratio of pea flour to water of 1:6, and stir evenly to obtain pea milk;

[0090] (3)Use the compound strain gel beads of immobilized cells prepared in Example 2;

[0091] (4)Removing residue: Use a gauze filter to remove bean dregs, and repeat 2 - 5 times to remove as much bean dregs as possible;

[0092] (5)Fine grinding: Use a homogenizer to further process the pea flour slurry for 5 minutes, with a mesh size of 60.

[0093] (6) Fermentation: Add the compound strain gel beads to the slurry, stir evenly and then carry out fermentation. The fermentation temperature is 37°C and the fermentation time is 8 h. Stir for 1 h and let stand for 3 h first, then stir for 1 h and let stand for 1 h until the fermentation ends;

[0094] (7) Separation: Filter the compound strain gel beads in the fermentation broth through a gauze filter screen and retain them for the next fermentation. Remove the supernatant and centrifuge to collect the precipitate;

[0095] (8) Drying: Dry the precipitate with hot air at 40°C to obtain 277.3 kg of pea starch.

[0096] Example 5

[0097] Compared with Example 4, except that Lactobacillus bulgaricus is replaced with an equal amount of Lactobacillus delbrueckii, the rest are the same as Example 4. Finally, 293.3 kg of pea starch is obtained.

[0098] Example 6

[0099] Compared with Example 4, except that Lactobacillus bulgaricus is replaced with an equal amount of Lactobacillus helveticus, the rest are the same as Example 3. Finally, 294.7 kg of pea starch is obtained.

[0100] Comparative Example 1

[0101] Compared with Example 4, except that Streptococcus thermophilus is replaced with an equal amount of maltodextrin, the rest are the same as Example 4. Finally, 264.7 kg of pea starch is obtained.

[0102] Comparative Example 2

[0103] The main steps of the traditional starch extraction method are briefly described as follows:

[0104] Raw material pretreatment: Wash 1000 kg of raw material pea grains, remove impurities, and add 1500 kg of water to soak for 48 h;

[0105] Crushing and grinding: Mechanically crush or grind the raw materials for 25 - 40 min to release starch granules;

[0106] Fiber separation: Remove fibers and dregs by screening or centrifugation, and retain the starch-containing slurry;

[0107] Starch precipitation: Let the slurry stand or centrifuge, and use gravity or the chemical aid lime water to promote the sedimentation of starch to form a starch milk;

[0108] Washing and refining: Wash the precipitated crude starch several times to remove residual proteins and soluble impurities;

[0109] Dehydration and drying: Use natural drying or hot air drying (50 - 60°C) to reduce the moisture, and finally obtain 240.2 kg of pea starch.

[0110] In summary, in Examples 2-4, the recovery of the bacterial strains was achieved through immobilized gel beads (sodium alginate carriers), and a relatively high yield was still maintained after repeated use (the yield of 277.3 kg was obtained using the recycled gel beads in Example 4, which was the same as that of the newly prepared gel beads in Example 3). In Comparative Example 1, since the immobilization technology was not adopted (maltodextrin was used to replace the bacterial strains), the yield decreased to 264.7 kg, indicating that the activity and recovery ability of the bacterial strains directly affected the efficiency. The yield of Example 2 (Lactobacillus bulgaricus + Streptococcus thermophilus) was 297.3 kg, which was higher than 264.7 kg of Comparative Example 1 (maltodextrin replaced Streptococcus thermophilus), indicating that the compound bacterial strains optimized the extraction efficiency through synergistic metabolism (such as promoting starch release by lactic acid production). In traditional Comparative Example 2, no bacterial strain fermentation was adopted, and the yield was even lower. In Examples 2-6, a homogenizer was used to treat the slurry (60-mesh screening), combined with multiple slag removals (filtered with gauze 2-5 times), effectively removing fiber impurities and improving the purity of the precipitate. In Comparative Example 2, the traditional method only relied on static settling or centrifugation, and the homogenization step was not mentioned, so more proteins might remain in the starch. The yields of Example 5 (Lactobacillus delbrueckii) and Example 6 (Lactobacillus helveticus) were 293.3 kg and 294.7 kg respectively, slightly lower than that of Lactobacillus bulgaricus (297.3 kg), indicating that Lactobacillus bulgaricus had a better degradation efficiency for pea starch. The yield of Comparative Example 1 (maltodextrin replaced Streptococcus thermophilus) was the lowest (264.7 kg), indicating that the compounding of bacterial strains was essential, and the heat tolerance of Streptococcus thermophilus played a key role in maintaining the fermentation stability. The yield of Example 2 (water-to-powder ratio of 1:4) was 297.3 kg, higher than 277.3 kg of Examples 3 / 4 (water-to-powder ratio of 1:6), indicating that a low water-to-powder ratio could reduce the subsequent drying energy consumption, but the slurry fluidity (homogenization difficulty) needed to be balanced. Examples 2-4 adopted an intermittent stirring strategy of stirring for 1 h - static settling for 3 h - stirring for 1 h - static settling for 1 h, which not only avoided the sedimentation of bacterial strains (such as the traditional static settling method), but also reduced the energy consumption of continuous stirring, while maintaining the lactic acid concentration gradient to promote starch release. It should be noted that there were differences between the bacterial powder ratios (50%:50%) in Examples 2-4 and the optimal group in Table 1 (44%:44%), but the actual yield (297.3 kg) was still relatively high, probably because the higher bacterial powder loading amount (1:150) in the examples made up for the ratio deviation.

[0111] Example 7

[0112] On the basis of Example 1, this example further proposes an implementation method for optimizing the extraction method.

[0113] This method further includes the following steps:

[0114] When using the compound bacterial strain gel beads for fermentation each time, the lactic acid concentration in the fermentation broth is monitored in real time to calculate the current lactic acid production rate during this fermentation process;

[0115] Record the cumulative usage times of the compound strain gel beads and the corresponding current lactic acid production rate;

[0116] Using the cumulative usage times as the abscissa and the current lactic acid production rate as the ordinate, fit and generate a lactic acid production rate decay curve;

[0117] Generate a gel bead label based on the lactic acid production rate decay curve and the cumulative usage times, and the gel bead label is used to dynamically adjust the fermentation parameters in the subsequent fermentation process; the fermentation parameters include stirring frequency, fermentation temperature and fermentation duration.

[0118] Optionally, for high-activity and low-loss gel beads, adopt the first fermentation parameters. Stirring frequency: The initial stirring frequency is set to 150 rpm. Since the gel beads have high activity and strong metabolism at this stage, the mass transfer requirements of the substrate and product are relatively large. In the early stage of fermentation (0 - 4 hours), maintain a stirring frequency of 150 rpm to promote sufficient contact between the substrate and the gel beads. In the middle stage of fermentation (4 - 6 hours), according to the change of the lactic acid production rate, if the rate rises steadily, the stirring frequency can be appropriately increased to 160 rpm to enhance the mass transfer effect. In the late stage of fermentation (6 - 8 hours), when the lactic acid production rate begins to level off, reduce the stirring frequency to 140 rpm to reduce energy consumption. Fermentation temperature: The fermentation temperature is set to 37°C. During the whole fermentation process, keep the temperature constant because the high-activity gel beads can maintain the best metabolic activity at this temperature. Real-time monitor and adjust the temperature through a temperature sensor and a heating / cooling system to ensure that the temperature fluctuation range is within ±0.5°C. Fermentation duration: According to experience and experimental data, the fermentation duration of high-activity and low-loss gel beads is usually set to 8 hours. During the fermentation process, monitor the lactic acid concentration in real time, and when the lactic acid concentration reaches the preset target value (such as 15 g / L), the fermentation can be terminated in advance.

[0119] Optionally, for medium - activity and medium - loss gel beads, use the second fermentation parameters. Stirring frequency: The initial stirring frequency is set to 160 rpm. Since the activity of the gel beads has decreased to some extent and there is a certain degree of loss of the carrier, it is necessary to increase the stirring frequency to enhance the mass transfer of the substrate and the product. In the early stage of fermentation (0 - 3 hours), maintain a stirring frequency of 160 rpm. In the middle stage of fermentation (3 - 5 hours), if the lactic acid production rate is lower than expected (such as lower than 1.0 mmol / (L·h)), increase the stirring frequency to 170 rpm. In the late stage of fermentation (5 - 7 hours), when the lactic acid production rate begins to decline, reduce the stirring frequency to 150 rpm. Fermentation temperature: Raise the fermentation temperature to 38°C. Appropriately increasing the temperature can improve the metabolic activity of the strain to a certain extent, but attention should be paid to avoiding the inactivation of the strain due to too high temperature. Through the temperature control system, control the temperature fluctuation range within ±0.5°C. Fermentation duration: The fermentation duration is set to 9 hours. Since the activity of the gel beads has decreased and the fermentation speed will slow down, it is necessary to appropriately extend the fermentation time. During the fermentation process, closely monitor the change of lactic acid concentration. If the lactic acid concentration does not reach the target value within 9 hours, it can be extended by 1 - 2 hours according to the actual situation.

[0120] Optionally, for low - activity and high - loss gel beads, use the third fermentation parameters. Stirring frequency: The initial stirring frequency is set to 170 rpm. Low - activity and high - loss gel beads require a higher stirring frequency to promote the contact between the substrate and the strain. In the early stage of fermentation (0 - 2 hours), maintain a stirring frequency of 170 rpm. In the middle stage of fermentation (2 - 4 hours), if the lactic acid production rate is extremely low (such as lower than 0.5 mmol / (L·h)), the stirring frequency can be increased to 180 rpm, but attention should be paid to avoiding excessive stirring resulting in the fragmentation of the gel beads. In the late stage of fermentation (4 - 6 hours), appropriately reduce the stirring frequency to 160 rpm according to the change of the lactic acid production rate. Fermentation temperature: Raise the fermentation temperature to 39°C. A higher temperature can stimulate the metabolism of the strain, but the temperature should be strictly controlled to avoid the death of the strain. Through a high - precision temperature sensor and temperature regulation system, control the temperature fluctuation range within ±0.3°C. Fermentation duration: The fermentation duration is set to 10 - 12 hours. Since the activity of the gel beads is very low and the fermentation speed is extremely slow, it is necessary to greatly extend the fermentation time. During the fermentation process, continuously monitor the lactic acid concentration until the target value is reached.

[0121] Furthermore, generating a gel bead label based on the lactic acid production rate decay curve and the cumulative usage times includes the following steps:

[0122] Determine the activity level according to the percentage of the current lactic acid production rate to the initial lactic acid production rate when the compound strain gel beads are used for the first time; specifically, the initial lactic acid production rate is the measured average value at the first use.

[0123] Determine the loss level according to the cumulative usage times;

[0124] Use the combination of the activity level and the loss level as the gel bead label.

[0125] Further, the activity levels include high activity, medium activity, and low activity, and the loss levels include high loss, medium loss, and low loss;

[0126] Specifically, samples are taken every 30 min during the fermentation process, the concentration is measured using a lactic acid kit, the current rate is calculated, data of 15 batches are continuously recorded, and an exponential decay model (i.e., the lactic acid production rate decay curve) is obtained by fitting with Origin 2023: (R² = 0.982), where n is the cumulative usage times, and v t represents the model predicted rate; define the initial rate v0 = 1.8 mmol / (L·h) (the average value of the first use), and the activity level is divided as follows: high activity: v ≥ 80%v0; medium activity: 50%v0 ≤ v < 80%v0, low activity: v < 50%v0; the loss level is based on the cumulative times: low loss (n ≤ 5), medium loss (6 ≤ n ≤ 10), high loss (n > 10). The verification data of the lactic acid production rate decay curve are shown in Table 2:

[0127] Table 2

[0128]

[0129] As can be seen from Table 2, for the gel beads with n ≤ 1, the lactic acid production efficiency is close to the initial rate of 1.8 mmol / (L·h) at this time. Therefore, taking ≥ 80% of the initial rate as the differentiation range of the high - activity level; corresponding to the usage stage of n = 3 - 8 times, the lactic acid production rate decreases as the mitochondrial function decays. Therefore, taking 50% - 80% of the initial rate as the differentiation range of the medium - activity level; when n ≥ 10 times, the lactic acid production ability significantly declines. Therefore, taking < 50% of the initial rate as the differentiation range of the low - activity level.

[0130] Specifically, as can be seen from Table 2, the accuracy of the exponential decay model is relatively high. Taking the second - order derivative of this model, it is found that: n = 5: the slope of the curve first significantly increases (the decay rate accelerates), n = 10: enters the plateau period (the rate decline slows down but the absolute value is already lower than 50% of the initial value). It can also be verified from Table 1 that: when n = 5, the predicted value is 1.24 (69% of the initial rate), when n = 10, it is 0.86 (48%), corresponding to the medium / low - activity thresholds respectively. Therefore, taking n ≤ 5, 6 ≤ n ≤ 10, and n > 10 as the differentiation boundaries of the loss level.

[0131] After recovering the compound strain gel beads in the fermentation broth, the following steps are further included:

[0132] If the activity level of the compound strain gel beads is low activity and the loss level is high loss, the compound strain gel beads shall be compulsorily scrapped or used at a lower grade after regeneration.

[0133] Specifically, the compulsory scrapping conditions: when the gel beads show any of the following situations: low activity level and high loss level (n > 15 times, v < 30%v0), surface crack rate > 40% (observed under a microscope with a 50x objective lens), swelling rate > 50% (determined by the gravimetric method).

[0134] Furthermore, the method also includes regenerating the compound strain gel beads with low activity level and high loss level.

[0135] The regeneration of the compound strain gel beads with low activity level and high loss level includes the following steps:

[0136] Soak the compound strain gel beads in the regeneration solution of sodium alginate, and apply high-pressure homogenization treatment to repair the surface cracks of the compound strain gel beads to obtain repaired gel beads.

[0137] Transfer the repaired gel beads to the activation solution, and shake and culture to restore the metabolic activity of the strains to obtain activated gel beads; the activation solution is a phosphate buffer solution containing glucose, yeast extract, and magnesium sulfate.

[0138] Detect the lactic acid production rate of the activated gel beads. If it reaches the first proportional range of the initial lactic acid production rate, mark the gel bead label of the activated gel beads as medium activity - medium loss during regeneration, and limit its cumulative usage times.

[0139] Specifically, for repair: low activity - high loss gel beads (n = 12 times, v = 42%v0) are soaked in a 2% sodium alginate + 0.05% CaCl2 regeneration solution, and after high-pressure homogenization (150 MPa, 3 times), the crack closure rate reaches 78% (observed by SEM). For activation: the repaired gel beads are shaken (180 rpm) at 37°C for 24 h in the activation solution (1% glucose + 0.5% yeast extract + 0.1% magnesium sulfate, pH 6.5), and the lactic acid production rate is restored to 65%v0.

[0140] Furthermore, adding the compound strain gel beads to the pea starch slurry for fermentation includes the following steps:

[0141] If the activity level of the compound strain gel beads is medium activity and the loss level is medium loss, add the compound strain gel beads to the pea starch slurry, and dynamically supplement carbon source or cofactor according to the real-time lactic acid production rate.

[0142] Specifically, for medium-loss gel beads (cumulative use times 6 - 10 times), due to mechanical wear and calcium loss of the sodium alginate carrier, the surface microporosity decreases by 35% (SEM comparison), restricting substrate diffusion (the mass transfer coefficient decreases from 1.2×10 -5 cm / s to 0.8×10 -5 cm / s). The synergistic effect of Lactobacillus and Streptococcus thermophilus depends on the NAD+ cycle, and the coenzyme regeneration efficiency decreases by 28% in the medium activity state (the intracellular NAD+ / NADH ratio detected by LC-MS decreases from 4.2 to 2.9). Therefore, use an online lactic acid sensor to record the lactic acid production rate (ΔC / Δt) every 15 minutes. When the real-time rate of medium-activity - medium-loss beads < 0.9 mmol / (L·h) (i.e., lower than 80% of the average rate of this level), it indicates feeding. Feeding substances: Carbon source: 50% glucose solution (sterile), feeding amount = (theoretical lactic acid production rate - measured lactic acid production rate) × fermentation volume × 0.8 (coefficient); Cofactor: 0.1% nicotinic acid (NAD+ precursor) + 0.05% magnesium sulfate, supplemented at 0.5 mL / L. At the 4th hour of fermentation (late logarithmic growth phase), at this time the metabolic demand of the strain is strong and the mass transfer resistance of the carrier begins to appear. Stirring strategy: After feeding, increase the rotation speed from 150 rpm to 180 rpm and maintain it for 30 minutes to promote mass diffusion. When the rate rises to ≥ 1.2 mmol / (L·h) after feeding, stop feeding. If there is still no improvement after cumulative feeding 2 times, terminate the fermentation in advance. The experimental data are shown in Table 3:

[0143] Table 3

[0144]

[0145] In a preferred embodiment, the obtaining of the compound strain gel beads includes the following steps:

[0146] Obtain the target process requirements for the current extraction, and the target process requirements include the target starch purity of the starch to be extracted;

[0147] If the target starch purity is greater than the first preset purity, select the newly prepared compound strain gel beads.

[0148] Further, after obtaining the target process requirements for the current extraction, the following steps are also included:

[0149] If the purity of the target starch is less than or equal to the first preset purity, corresponding recovered compound strain gel beads are selected according to the purity of the target starch.

[0150] Further, the step of selecting corresponding recovered compound strain gel beads according to the purity of the target starch includes the following steps:

[0151] If the purity of the target starch is less than the second preset purity, recovered compound strain gel beads with a gel bead label of low activity - high loss or a cumulative use times of the first preset times are selected; if the purity of the target starch is greater than the second preset purity and less than the first preset purity, recovered compound strain gel beads with a gel bead label of medium activity - medium loss and a cumulative use times less than the second preset times are selected.

[0152] Wherein, the second preset purity is less than the first preset purity, and the first preset times is greater than the second preset times.

[0153] Optionally, the first preset purity is 93%. When the purity of the target starch is greater than or equal to 93% (high - end food grade), newly - made beads (n = 0) are used; when the purity of the target starch is less than 93%, recovered beads are used. The second preset purity is 90%. When 90% ≤ the purity of the target starch < 93%, medium - activity - medium - loss beads (n = 6 - 8 times, v = 60 - 75%v0) are selected; when the purity of the target starch < 90%: low - activity - high - loss beads (n > 10 times, v < 50%v0) are selected. Verification experiment: Set the target starch purity to 92%, select medium - activity beads with n = 7 times, and the actual purity is 92.3% (n = 10, RSD = 0.8%), which is better than 89.7% when using low - activity beads (p < 0.01). The experimental data is shown in Table 4 as follows:

[0154] Table 4

[0155]

[0156] This embodiment can achieve precise matching of purity requirements, ensure product quality, optimize costs, and improve economic benefits. The yield of newly - made beads is 81.0%, and the cost is the highest. They are only used in high - end scenarios to reduce the consumption of newly - made beads; recovered beads are reused in medium / low - purity scenarios to reduce raw material costs. Medium - activity beads (n = 6 - 8 times, yield 80.5%) are reused in the 90 - 93% range, which not only ensures purity (3.1% higher than low - activity beads) but also avoids yield waste of newly - made beads (only 0.5% higher yield but higher cost), balancing loss and efficiency. Low - activity beads allow reuse with n > 10 times, fully exploiting the remaining value, suitable for scenarios with low purity requirements (such as industrial starch), extending the lifespan of gel beads, and reducing waste volume.

[0157] In this text, specific examples are used to illustrate the principles and implementation modes of the present application. The description of the above embodiments is only to help understand the method and its core idea of the present application. The above is only the preferred implementation mode of the present application. It should be noted that due to the limitation of literal expression, and objectively there are infinite specific structures. For those of ordinary skill in the art in this technical field, without departing from the principle of the present invention, several improvements, refinements or changes can also be made, or the above technical features can be combined in an appropriate manner; these improvements, refinements, changes or combinations, or directly applying the concept and technical solution of the invention to other occasions without improvement, shall all be regarded as the protection scope of the present application.

Claims

1. A method for extracting pea starch by using the immobilized cell method, characterized in that It includes the following steps: Pea seeds are peeled and ground into pea powder. Process water is added to the pea powder and stirred evenly to obtain pea milk; The soybean residue in the pea milk is filtered to obtain filtered soybean milk, and the filtered soybean milk is homogenized to obtain pea powder slurry; Compound strain gel beads are obtained and added to the pea powder slurry for fermentation, and stirred evenly to obtain fermentation broth; The compound strain gel beads in the fermentation broth are recovered, and the supernatant is recovered to obtain a precipitate; the supernatant is used to prepare the process water for the next extraction; The precipitate is dried to obtain pea starch; Among them, the compound strain gel beads include an immobilization carrier and a compound strain encapsulated in the immobilization carrier. The immobilization carrier is a biocompatible polysaccharide gel, and the compound strain is composed of Lactobacillus and Streptococcus thermophilus; the types of the compound strain gel beads include newly prepared compound strain gel beads with zero cumulative use times and recycled compound strain gel beads with at least one cumulative use time; Preparing the newly prepared compound strain gel beads includes the following steps: Taking Lactobacillus powder and Streptococcus thermophilus powder, mixing them in a mass ratio of 40%-48%:40%-48% to obtain a compound strain powder, and other proportioning substances are maltodextrin; Preparing a 1.5% sodium alginate solution, heating and stirring to dissolve, and cooling to room temperature; Adding the compound strain powder into the sodium alginate solution, and the mass ratio of the compound strain powder to sodium alginate is 1:0.5-1:5, stirring until evenly suspended to obtain a mixture; Dropping the mixture into a 2%-5% calcium chloride solution drop by drop, and standing at room temperature for crosslinking to form initial gel beads; Rinsing the initial gel beads with sterile water multiple times to remove residual calcium ions on the surface to obtain newly prepared compound strain gel beads; this method also includes the following steps: During each fermentation using the compound strain gel beads, the lactic acid concentration in the fermentation broth is monitored in real time to calculate the current lactic acid production rate during the current fermentation process; Recording the cumulative use times of the compound strain gel beads and the corresponding current lactic acid production rate; Taking the cumulative use times as the abscissa and the current lactic acid production rate as the ordinate, fitting to generate a lactic acid production rate decay curve; Based on the lactic acid production rate decay curve and the cumulative use times, generating a gel bead label, and the gel bead label is used to dynamically adjust the fermentation parameters during the subsequent fermentation process; the fermentation parameters include stirring frequency, fermentation temperature and fermentation duration; This method also includes regenerating the compound strain gel beads with low activity level and high loss level; The regeneration of the compound strain gel beads with low activity level and high loss level includes the following steps: Soaking the compound strain gel beads in a sodium alginate regeneration solution, and applying high-pressure homogenization treatment to repair the surface cracks of the compound strain gel beads to obtain repaired gel beads; Transferring the repaired gel beads to an activation solution, and oscillating and culturing to restore the metabolic activity of the strains to obtain activated gel beads; the activation solution is a phosphate buffer solution containing glucose, yeast extract and magnesium sulfate; Detect the lactic acid production rate of the activated gel beads. If it reaches the first proportional range of the initial lactic acid production rate, mark the gel bead label of the activated gel beads as active during regeneration - medium loss, and limit its cumulative usage times.

2. The method for extracting pea starch by using the immobilized cell method according to claim 1, characterized in that: Generating the gel bead label based on the lactic acid production rate decay curve and the cumulative usage times includes the following steps: Determine the activity level according to the percentage of the current lactic acid production rate to the initial lactic acid production rate when the compound strain gel beads are first used; Determine the loss level according to the cumulative usage times; Use the combination of the activity level and the loss level as the gel bead label.

3. The method for extracting pea starch by using the immobilized cell method according to claim 2, characterized in that: The activity levels include high activity, medium activity, and low activity, and the loss levels include high loss, medium loss, and low loss; After recovering the compound strain gel beads in the fermentation broth, the following steps are further included: If the activity level of the compound strain gel beads is low activity and the loss level is high loss, then force the compound strain gel beads to be scrapped or use them at a lower level after regeneration.

4. The method for extracting pea starch by the immobilized cell method according to claim 2, characterized in that: Adding the compound strain gel beads to the pea starch slurry for fermentation includes the following steps: If the activity level of the compound strain gel beads is medium activity and the loss level is medium loss, then add the compound strain gel beads to the pea starch slurry, and dynamically supplement carbon sources or coenzyme factors according to the real - time lactic acid production rate.

5. The method for extracting pea starch by the immobilized cell method according to claim 2, wherein: Obtaining the compound strain gel beads includes the following steps: Obtain the target process requirements for the current extraction, and the target process requirements include the target starch purity of the starch to be extracted; If the target starch purity is greater than the first preset purity, then select the newly prepared compound strain gel beads.

6. The method for extracting pea starch by the immobilized cell method according to claim 5, characterized in that: After obtaining the target process requirements for the current extraction, the following steps are further included: If the target starch purity is less than or equal to the first preset purity, then select the corresponding recovered compound strain gel beads according to the target starch purity.

7. The method for extracting pea starch by the immobilized cell method according to claim 6, wherein: Selecting the corresponding recovered compound strain gel beads according to the target starch purity includes the following steps: If the target starch purity is less than the second preset purity, then select the recovered compound strain gel beads with a gel bead label of low activity - high loss or a cumulative usage times of the first preset times; if the target starch purity is greater than the second preset purity and less than the first preset purity, then select the recovered compound strain gel beads with a gel bead label of medium activity - medium loss and a cumulative usage times less than the second preset times; Wherein, the second preset purity is less than the first preset purity, and the first preset times is greater than the second preset times.

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