CN115633718B - Method for improving plant lactosis enzymolysis efficiency through low-consumption physical double-field cooperation - Google Patents
Method for improving plant lactosis enzymolysis efficiency through low-consumption physical double-field cooperation Download PDFInfo
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/80—Food processing, e.g. use of renewable energies or variable speed drives in handling, conveying or stacking
- Y02P60/87—Re-use of by-products of food processing for fodder production
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Abstract
The invention discloses a method for improving the enzymolysis efficiency of plant milk by low-consumption physical double-field synergy, which comprises the steps of dissolving enzyme in water, and stirring in ice bath to obtain enzyme solution; and (3) performing low-temperature plasma treatment on the enzyme solution, then soaking and filtering the flaxseeds, grinding by a colloid mill to obtain flaxseeds homogenate, performing pretreatment on the flaxseeds homogenate by using a pulse electric field, adding the enzyme solution subjected to the CP treatment into the flaxseeds plant milk subjected to the PEF treatment for enzymolysis, inactivating the enzyme and filtering to obtain the flaxseeds plant milk with high total solid content, protein content and total phenol content and moderate viscosity. After the low-consumption physical double-field treatment, the edible quality of the flaxseed plant milk is greatly improved, the total phenol content, the protein content and the solid content are respectively improved by 66%, 17% and 12%, and the viscosity is reduced by 49%. The double-field coupling enzymolysis shows synergistic effect and has great application potential in the fields of green, clean label and high-quality plant milk processing.
Description
Technical Field
The invention relates to the technical field of functional food processing, in particular to a method for improving the enzymolysis efficiency of plant milk by low-consumption physical double-field cooperation.
Background
Enzymes are found in all naturally occurring life forms, including plants, animals, and microorganisms. Enzymes are used in the food industry primarily to alter the functional properties of products and/or to control or modify food processing. The enzyme can replace a synthetic catalyst, reduce the use of toxic reagents, generate less energy consumption, and is environment-friendly, and belongs to low-carbon and low-consumption technologies. Currently, global agricultural foods produce 25% (125 hundred million tons) of carbon emissions, the first of which is the national "carbon emissions reduction", "carbon peak" green environmental requirement, and the improvement of product functions including texture, appearance, nutritional quality, shelf life and safety by enzymes is widely accepted by the food processing industry, and therefore, the use of enzymes in the agricultural product processing industry, particularly in the food industry, is becoming increasingly attractive.
However, the enzyme is a special protein with catalytic activity and high specificity, and has very high substrate specificity, so that the complex and variable processing conditions of the industrial process and the mixing of multiple substrates further increase the instability of the enzymolysis process, thereby reducing the industrial efficiency of the enzyme, and simultaneously, the price of the enzyme per se is higher than that of a synthetic catalyst, which puts higher requirements on the improvement of the enzymolysis efficiency and the optimization of a enzymolysis process. The speed/efficiency of the enzymatic hydrolysis reaction is mainly dependent on 2 factors such as mass transfer efficiency and enzyme molecular conformation. The method has the potential of improving enzymolysis efficiency by using low-consumption, green and environment-friendly physical field treatment.
The low-temperature plasma (CP) is a fourth state of a substance, is a system composed of a large number of molecules, atoms, ions, electrons and other particles, can generate active particles with higher energy such as active oxygen and active nitrogen at lower energy consumption, and can keep high enough activity at lower temperature (less than 50 ℃), so that the conformation of enzyme molecules is influenced to different degrees, and the catalytic efficiency of the enzyme is changed. Non-heat treatment equipment and process are efficient new methods for improving the performance of plant milks. The Pulse Electric Field (PEF) is used as a low-consumption, non-thermal and novel processing technology, on one hand, high-energy particles are generated by inducing plant stress stimulation reaction, on the other hand, the food structure change is caused, for example, reversible or irreversible pore formation in cell membranes is caused by electropermeabilization effect, and further, accumulation of nutrients including proteins, phenols and the like in a food system is promoted, so that the Pulse Electric Field (PEF) can be used for processing plant raw materials, such as extraction of plant compounds, and preparation of high-quality fruit juice/plant milk and other beverages. In addition, the pretreatment of the reaction substrate by PEF has the effect of improving the contact efficiency of the enzyme and the substrate, so that the PEF has the potential of promoting the improvement of the enzymolysis efficiency. Ultrasound (US) is a non-thermal technique that has been attracting attention due to its unique cavitation effect and excellent performance, and research has shown that US treatment can increase the structural flexibility of colloidal macromolecules such as proteins, polysaccharides, etc., and has potential effects in enhancing enzyme interactions with substrates.
At present, the plasma technology has been applied to various fields and plays a positive role, such as aerospace, biology, medical treatment and the like, wherein the atmospheric pressure plasma equipment is simple and convenient, has low cost and can be widely applied to the fields of food processing and the like. Meanwhile, the pulse electric field and the ultrasonic have wide application range and low energy consumption, belong to a novel green processing technology, and are increasingly widely applied to the field of food processing at home and abroad.
Therefore, providing a method for improving the efficiency of plant lactosis by low-consumption physical double-field synergy is a technical problem to be solved by the technicians in the field.
Disclosure of Invention
In view of the above, the invention provides a method for improving the plant lactolysis efficiency by low-consumption physical double-field synergy, which utilizes the low-consumption physical double-field synergy to improve the plant lactolysis efficiency and prepares the flaxseed plant milk with high protein content and total phenol content and moderate viscosity. The method has the characteristics of simplicity and convenience in operation and high efficiency.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
a method for improving the enzymolysis efficiency of plant milk by low-consumption physical double-field cooperation comprises the following steps:
(1) Preparing an enzyme solution: weighing enzyme, adding water, mixing, and stirring in ice bath to obtain enzyme solution;
(2) Low temperature plasma treatment: sucking enzyme liquid into a clean culture dish, placing the culture dish below a nozzle of a plasma generator, treating the enzyme liquid with stable plasma, collecting the treated enzyme liquid, and sealing and preserving at 2-8deg.C for later use;
(3) Preparing flax seed homogenate: mixing microwave degummed flax seeds with water, soaking, discarding water, adding water into soaked flax seeds, circulating with colloid mill for 4-12min, and collecting plant milk;
(4) Pretreatment of flax seed homogenate by a pulsed electric field: carrying out a pulse electric field on the plant milk prepared in the step (3) to obtain a treatment fluid;
(5) Enzymolysis: adding 0.1-0.5wt% of the enzyme solution prepared in the step (1) or the step (2) into the treatment solution, inactivating enzyme after enzymolysis, and filtering to obtain the flaxseed plant milk with high protein content, high solid content and moderate viscosity.
Further, the enzyme in the step (1) is one or more of cellulase, protease, phytase, hemicellulase, saccharifying enzyme and bromelain; the mass ratio of enzyme to water is 1-5:10.
The beneficial effect of adopting above-mentioned further scheme lies in: by adopting the scheme of the invention, the solid enzyme powder can be fully dissolved, and the method is suitable for the processing requirements of low-temperature plasma equipment on samples.
Further, in the step (2), the pulse discharge voltage of the low-temperature plasma treatment is 16-20kV, the frequency is 200-2000Hz, the pulse width is 100-200ns, and the treatment time is 15-60s.
The beneficial effect of adopting above-mentioned further scheme lies in: the low-temperature plasma in the scheme of the invention generates active particles with higher energy under low energy consumption, influences the conformation of enzyme molecules and changes the enzyme activity. The cellulase is treated by low-temperature plasma for 15s, and the enzyme activity of the cellulase can be improved by 89.56 percent.
Further, the mass ratio of the microwave degummed flax seeds to the water in the step (3) is 1:5-10, and the soaking time is 0.5-3h.
The beneficial effect of adopting above-mentioned further scheme lies in: the soaking in the proportion can make the flaxseeds quickly expand and soften by absorbing water, and is beneficial to improving the pulping efficiency.
Further, the electric field strength of the pulsed electric field treatment in the step (4) is 1.3-3.7kV/cm, the pulse number is 3-9 times, the pulse width is 100-500ns, the frequency is 500-1000Hz, and the flow rate is 40-80rpm.
The beneficial effect of adopting above-mentioned further scheme lies in: the pulsed electric field is used for preprocessing the linseed homogenate, has the effect of improving the contact efficiency of enzyme and substrate, and is beneficial to assisting enzymolysis and promoting the release of total phenols in linseed plant milk.
Further, the pulsed electric field in step (4) may be replaced with ultrasound
The beneficial effect of adopting above-mentioned further scheme lies in: the pretreatment of the flaxseed homogenate by the ultrasonic wave has the effect of improving the contact efficiency of enzyme and substrate, and is beneficial to assisting enzymolysis to promote the release of protein in the flaxseed plant milk.
Further, the enzyme solution in the step (5) is added in an amount of 0.1 to 0.5wt%.
Further, the enzymolysis temperature in the step (5) is 45-55 ℃ and the enzymolysis time is 30-75min; the enzyme deactivation temperature is 90-100 ℃, and the enzyme deactivation time is 10-15min; the filtering and sieving granularity is 120-200 meshes.
The beneficial effect of adopting above-mentioned further scheme lies in: the enzyme hydrolyzes the glycosidic bond in the plant cell wall layer to promote the dissolution of the cell content, and the low-consumption physical double-field coupling enzyme method greatly improves the edible quality of the flaxseed plant milk, wherein the total phenol content, the protein content and the solid content are respectively improved by 66%, 17% and 12%, thus giving the flaxseed plant milk a higher nutritional value.
Compared with the prior art, the invention has the beneficial effects that:
1. after the low-consumption physical double-field treatment, the edible quality of the flaxseed plant milk is greatly improved, wherein the total phenol content, the protein content and the solid content are respectively improved by 66%, 17% and 12%, and the flaxseed plant milk is endowed with higher nutritive value.
2. Compared with the single conventional technology, the preparation technology of the invention has the advantages of simple and convenient preparation process, easy control of reaction process, short production period and low production cost, and belongs to the green, clean label and high-quality plant milk processing modes.
3. The invention provides a method for improving the enzymolysis efficiency of plant milk by low-consumption physical double-field coupling for the first time, which directly improves the activity of enzyme through a first physical field, directly improves the enzymolysis efficiency, shortens the reaction time, and indirectly further improves the enzymolysis effect by treating plant milk through a second physical field to promote the contact of enzyme and a substrate. The research of the invention discovers that after low-consumption physical double-field treatment, the edible quality of the flaxseed plant milk is greatly improved, wherein the total phenol content, the protein content and the solid content are respectively improved, and the viscosity is reduced by 49 percent. The data show that the double-field coupling enzymolysis shows synergistic effect and has great application potential for the fields of green, clean labels and high-quality plant milk processing.
Drawings
FIG. 1 shows the enzyme activity and the improvement rate of cellulase after various times of low-temperature plasma treatment;
FIGS. 2A and 2B show the total phenol content and viscosity of flaxseed milk prepared with the aid of cellulase at low temperature plasma, respectively;
FIGS. 3A and 3B show the solids content and protein content, respectively, of a low temperature plasma assisted bromelain for the preparation of flaxseed milk;
FIG. 4 shows the total phenol content of the pulsed electric fields of different electric field strengths after separate treatment or assisted enzymatic hydrolysis of flaxseed milk;
FIGS. 5A and 5B show the solids content of the flaxseed milk prepared with bromelain with different ultrasonic powers and different ultrasonic times, respectively;
FIGS. 6A and 6B show the protein content and the solids content of flaxseed milk produced by the low temperature plasma and ultrasonic coupled enzyme process, respectively;
FIGS. 7A and 7B show the total phenol content and viscosity of flaxseed milk prepared by low temperature plasma and pulsed electric field coupled enzyme process, respectively;
fig. 8 is a schematic diagram of the process flow of preparing flax seed milk.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Example 1 first physical field to enhance enzyme Activity
1.1. Materials and reagents
Cellulase CTS, weifang kangdian biotechnology limited; cellulase (CL) activity detection kit, AKSU043M, beijing Box manufacturing technology Co., ltd.
1.2. Main instrument and equipment
Digital display thermostatic water bath, HH-6, hongkao instrument factory in Jintan; a multifunctional enzyme-labeled instrument, spectraMAXM2, a Mei Gu molecular instrument (Shanghai) Limited, a plasma discharge device driven by a pulse power supply.
1.3. Experimental methods and results
Green physical field low-temperature plasma treatment of cellulase to improve cellulase enzyme activity
(1) Preparing an enzyme solution: 15g of cellulase is weighed and added with 50mL of water to be mixed uniformly, and the mixture is stirred for 30min in an ice bath.
(2) Low temperature plasma treatment of enzyme solution: 1mL of enzyme solution is sucked into a clean culture dish (diameter of 35 mm) and marked, and is placed below a nozzle of a plasma generator, the pulse discharge voltage is 18kV, the frequency is 1000Hz, and the pulse width is 200ns. When the plasma is stable (i.e. no obvious filiform spark exists), a timer is started, the treatment time is set to 60s, and the collected and treated cellulase is sealed and stored at the temperature of 4 ℃ for standby.
(3) Enzyme activity was determined using a box cellulase activity assay kit.
Pretreatment of enzyme solution: centrifuging the enzyme solution subjected to CP treatment at 4 ℃ for 10min, and taking the supernatant to be tested.
The measuring step comprises the following steps: the first 50. Mu.L of the reagent, the second 200. Mu.L of the reagent, the 50. Mu.L of water and the 50. Mu.L of the supernatant of the crude enzyme solution are sequentially added into the measuring tube, the crude enzyme solution in the comparison tube is replaced by the inactivated crude enzyme solution corresponding to the inactivation, and other reagents and the addition amount are kept unchanged. After fully mixing, saccharifying for 30min in a water bath at 50 ℃, immediately boiling the mixture in the water bath for 15min, (sealing to prevent water loss), and obtaining the saccharified liquid. 10. Mu.L of the saccharification liquid was aspirated, 30. Mu.L of reagent III was added, and the mixture was developed in a boiling water bath for 15min (sealed to prevent water loss) and cooled to room temperature. The standard tube saccharification was replaced with 10. Mu.L of standard diluent, and the remaining reagents and amounts added remained unchanged. 200 mu L of reaction solution is sucked into a 96-well plate, and the absorbance at 540nm is measured and marked as A measurement, A control, A standard and A blank; calculation of Δaassay=aassay-a control, Δastandard=a standard-a blank.
Cellulase (CL) activity calculation: unit definition: catalytic production of 1 μg glucose per minute per mL of liquid sample is defined as one unit of enzyme activity.
CL(U/mL)＝1000×x×V Inverse total /(V Sample ×T)＝233.33×x
Annotation: v (V) Inverse total : 0.35mL of total volume of the reaction system; v (V) Sample : adding the volume of the crude enzyme solution into the reaction system to be 0.05mL; t: saccharification time, 30min;1000: unit conversion coefficient, 1 mg/ml=1000 μg/mL.
As shown in FIG. 1, when the enzyme activity is increased and then decreased after the CP treatment is performed for different times, and the CP treatment time is 15s, the enzyme activity can be improved by 89%, and the improvement of the enzyme activity can be caused by the influence of active particles on the enzyme molecular conformation to different degrees, so that the active sites of the enzyme are exposed, and the enzyme activity is improved.
Example 2 first physical field improves the actual enzymatic efficiency of enzymes in plant milk processing
2.1. Materials and reagents
Cellulase CTS, weifang kangdian biotechnology limited; bromelain, beijing Soy Bao technology Co., ltd; dry degummed flax seeds, food grade, academy of agricultural sciences in Gansu province; sea sand, concentrated sulfuric acid, boric acid, etc. are purchased from national pharmaceutical group chemical reagent limited; kjeldahl nitrogen high efficiency catalytic tablet, fu Lin Fen all come from source leaf organisms.
2.2. Main instrument and equipment
A pulse power supply driven plasma discharge device; colloid mill, horizontal 60 type, shenzhen medical Co., ltd; digital display thermostatic water bath, HH-6, hongkao instrument factory in Jintan; electrothermal blowing drying oven, 101-1AB, tiest instruments Inc. of Tianjin; full-automatic Kai-Shi azotometer, K9860, available from the technology group Co., ltd; digital viscometer, SNB-1, shanghai precision scientific instruments Inc.
2.3. Experimental methods and results
2.3.1A method for improving enzymolysis efficiency of cellulase in plant milk processing by applying low-temperature plasma to cellulase comprises the following steps:
(1) Preparing an enzyme solution: 15g of cellulase CTS is weighed, 50mL of water is added for uniform mixing, and the mixture is stirred for 30min in an ice bath.
(2) Low temperature plasma treatment of enzyme solution: 1mL of enzyme solution is sucked into a clean culture dish (diameter of 35 mm) and marked, and is placed below a nozzle of a plasma generator, the pulse discharge voltage is 18kV, the frequency is 1000Hz, and the pulse width is 200ns. When the plasma is stable (i.e. no obvious filiform spark exists), a timer is started, the treatment time is set to 15s, and the collected and treated cellulase is sealed and stored at the temperature of 4 ℃ for standby.
(3) Preparing flax seed homogenate: soaking the degummed flax seeds for 2 hours by using microwaves in a ratio of 1:7, discarding the soaking solution, adding pure water according to the ratio of 1:7, and collecting plant milk after a colloid mill circulates for 12 minutes.
(4) The enzymolysis process comprises the following steps: adding 0.1wt% of cellulase treated by CP into the treatment solution, performing enzymolysis at 50deg.C for 30min, inactivating enzyme at 90deg.C for 15min, and filtering with 200 mesh filter cloth to obtain flaxseed plant milk.
As shown in fig. 2A and 2B, after the CP-assisted cellulase enzymes enzymatically digest the flaxseed plant milk, the application of CP was found to increase the efficiency of the reduction in the release efficiency viscosity of total phenols in the flaxseed plant milk. The complex field therefore selects CP-treated cellulases 60s.
2.3.2 method for improving the enzymolysis efficiency of bromelain in the processing of plant milk by applying low temperature plasma to bromelain, optimizing the treatment time, comprising the following steps:
(1) Preparing an enzyme solution: 1g of bromelain is weighed, added with 10mL of water and evenly mixed, and stirred for 30min in an ice bath.
(2) Low temperature plasma treatment of enzyme solution: 1.5mL of enzyme solution is sucked into a clean culture dish (diameter of 35 mm) and marked, and is placed below a nozzle of a plasma generator, and the pulse discharge voltage is 18kV, the frequency is 1kHz, and the pulse width is 200ns. When the plasma is stable (i.e. no obvious filament spark), a timer is started, the treatment time is set to 45s, and the collected and treated bromelain is sealed and stored at 4 ℃ for standby.
(3) Preparing flax seed homogenate: mixing and soaking the microwave degummed flax seeds and water for 2 hours according to the mass ratio of 1:7, discarding the soaking liquid, and collecting the plant milk after 12 minutes of colloid mill circulation.
(4) The enzymolysis process comprises the following steps: adding bromelain 0.075wt% into the treatment solution, performing enzymolysis at 50deg.C for 1 hr, inactivating enzyme at 90deg.C for 15min, and filtering with 200 mesh filter cloth to obtain flaxseed plant milk.
As shown in fig. 3A and 3B, after CP-assisted bromelain enzymatic hydrolysis of flaxseed milk, CP treatment for too long or too short a period of time was found to reduce the release efficiency of flaxseed milk proteins; and when the CP is processed for 45 or 60 seconds, the content release efficiency of the CP-assisted enzymolysis of flaxseed plant milk solids and protein is improved. Therefore, the complex field selects CP to treat bromelain for 60s to improve the enzymolysis efficiency of bromelain in the processing of plant milk.
The experimental result shows that the CP physical field can not only improve the measured enzyme activity value, but also improve the real enzymolysis effect of the CP physical field in a complex plant milk system.
Example 3 second physical field enhancement of Linseed plant dairy
3.1. Materials and reagents
Cellulase CTS, weifang kangdian biotechnology limited; bromelain, beijing Soy Bao technology Co., ltd; dry degummed flax seeds, food grade, academy of agricultural sciences in Gansu province; sea sand, concentrated sulfuric acid, boric acid, etc. are purchased from national pharmaceutical group chemical reagent limited; kjeldahl nitrogen high efficiency catalytic tablet, fu Lin Fen all come from source leaf organisms.
3.2. Main instrument and equipment
High-voltage pulse electric field generator THU-PEF4, available from Wuhan New Tianpu laboratory equipment Co., ltd; ultrasonic cell grinder, JY92-IIDN, ningbo Xinzhi biotechnology Co., ltd; colloid mill, horizontal 60 type, shenzhen medical Co., ltd; digital display thermostatic water bath, HH-6, hongkao instrument factory in Jintan; electrothermal blowing drying oven, 101-1AB, tiest instruments Inc. of Tianjin; full-automatic Kai-Shi azotometer, K9860, available from the technology group Co., ltd.
3.3. Experimental methods and results
3.3.1A method for improving the quality of flaxseed plant milk by applying a pulsed electric field to the flaxseed homogenate to optimize the electric field strength and establish a high-efficiency flaxseed lactase process, comprising the following steps:
(1) Preparing an enzyme solution: 15g of cellulase CTS is weighed, 50mL of water is added for uniform mixing, and the mixture is stirred for 30min in an ice bath.
(2) Preparing flax seed homogenate: mixing and soaking the microwave degummed flax seeds and water for 2 hours according to the mass ratio of 1:7, discarding the soaking liquid, and collecting the plant milk after 12 minutes of colloid mill circulation.
(3) Pretreatment of flax seed milk by a pulsed electric field: carrying out pulsed electric field and ultrasonic treatment on the prepared flaxseed homogenate, wherein the electric field strength of the pulsed electric field treatment is 1.8kV/cm, the pulse frequency is 3 times, the pulse width is 500 mu s, the frequency is 1kHz, and the flow speed is 40rpm, so as to obtain a treatment liquid;
(4) The enzymolysis process comprises the following steps: adding 0.1wt% of cellulase treated by CP into the treatment liquid, performing enzymolysis at 50deg.C for 30min, inactivating enzyme at 90deg.C for 15min, and filtering with 200 mesh filter cloth to obtain flaxseed plant milk.
As shown in FIG. 4, the effect of releasing total phenols of flaxseed plant milk is weak when a low-intensity pulsed electric field is applied to the flaxseed homogenate, and the content of total phenols of flaxseed plant milk is remarkably improved when the electric field intensity is more than 2.7Kv/cm, probably due to the increase of the electric field intensity when the electric field intensity is more than 2.7Kv/cm, the permeability of plant cell membranes is improved, and the release capacity of active ingredients is enhanced. And when the electric field strength is too low, it is insufficient to affect the permeability of plant cells. In conclusion, the electric field strength of the selected pulse electric field is 3.7Kv/cm, and compared with a control group, the flaxseed plant milk under the process condition contains higher total phenols.
3.3.2 method for improving the quality of flaxseed plant milk by a second physical field (ultrasound), comprising the steps of:
(1) Preparing an enzyme solution: 1g of bromelain is weighed, added with 10mL of water and evenly mixed, and stirred for 30min in an ice bath.
(2) Preparing flax seed homogenate: mixing and soaking the microwave degummed flax seeds and water for 2 hours according to the mass ratio of 1:7, discarding the soaking liquid, circulating a colloid mill for 4 minutes (12 minutes), and collecting the plant milk.
(3) Ultrasonic pretreatment of flax seed milk: carrying out ultrasonic pretreatment on the prepared linseed homogenate for 200w and 10min (10 min,200w and 400 w) to obtain a treatment liquid;
(4) The enzymolysis process comprises the following steps: adding 0.075wt% of cellulase treated or not treated by CP into the treatment liquid, performing enzymolysis at 50deg.C for 30min, inactivating enzyme at 90deg.C for 15min, and filtering with 200 mesh filter cloth. Obtaining the flaxseed plant milk.
As shown in fig. 5A and 5B, after 200w or 400w10min of the ultrasonic treatment of flaxseed milk, the solid content of flaxseed plant milk is reduced, probably because flavonoids, polysaccharides and soluble proteins are degraded due to the excessive power. By exploring the ultrasonic time, after the flaxseed plant milk is subjected to ultrasonic-assisted enzymolysis, the total solid content of the flaxseed plant milk is reduced after the ultrasonic time is 10 min; the possible reason is that the longer the ultrasonic time, the stronger the cavitation effect, the greater the loss of ammonia nitrogen and the reduction of the solid content. As can be seen from the comparison of the solid content, the flaxseed plant milk under the process conditions has higher solid content, which is improved by 5.22%, so that the final ultrasonic condition is selected to be 200w and 5min.
The experiment shows that after the plant milk is treated by using the second physical field, the enzymolysis effect can be enhanced indirectly by improving the contact efficiency of the enzyme and the substrate, and the effect of improving the quality of the plant milk is achieved.
Example 4 Dual field coupling, synergistic enhancement of Linseed plant dairy
4.1. Materials and reagents
Cellulase CTS, weifang kangdian biotechnology limited; bromelain, beijing Soy Bao technology Co., ltd; dry degummed flax seeds, food grade, academy of agricultural sciences in Gansu province; sea sand, concentrated sulfuric acid, boric acid, etc. are purchased from national pharmaceutical group chemical reagent limited; kjeldahl nitrogen high efficiency catalytic tablet, fu Lin Fen all come from source leaf organisms.
4.2. Main instrument and equipment
A pulse power supply driven plasma discharge device; high-voltage pulse electric field generator THU-PEF4, available from Wuhan New Tianpu laboratory equipment Co., ltd; ultrasonic cell grinder, JY92-IIDN, ningbo Xinzhi biotechnology Co., ltd; colloid mill, horizontal 60 type, shenzhen medical Co., ltd; digital display thermostatic water bath, HH-6, hongkao instrument factory in Jintan; electrothermal blowing drying oven, 101-1AB, tiest instruments Inc. of Tianjin; full-automatic Kai-Shi azotometer, K9860, available from the technology group Co., ltd.
4.3. Experimental methods and results
4.3.1 double field (low temperature plasma and ultrasonic) coupling, the method for synergistically improving the quality of flaxseed plant milk comprises the following steps:
(1) Preparing an enzyme solution: 1g of bromelain is weighed, added with 10mL of water and evenly mixed, and stirred for 30min in an ice bath.
(2) Low temperature plasma treatment of enzyme solution: 1.5mL of enzyme solution is sucked into a clean culture dish (diameter of 35 mm) and marked, and is placed below a nozzle of a plasma generator, and the pulse discharge voltage is 18kV, the frequency is 1000Hz and the pulse width is 200ns. When the plasma is stable (i.e. no obvious filiform spark exists), a timer is started, the treatment time is set to 60s, and the collected and treated cellulase is sealed and stored at the temperature of 4 ℃ for standby.
(3) Preparing flax seed homogenate: mixing degummed 15 times of microwave flax seeds and water according to the mass ratio of 1:7, soaking for 2 hours, discarding the soaking liquid, and recycling the plant milk by a colloid mill for 12 minutes.
(4) Ultrasonic pretreatment of flax seed milk: carrying out ultrasonic pretreatment (200 w,5 min) on the prepared flaxseed homogenate;
(5) The enzymolysis process comprises the following steps: adding bromelain 0.075wt% after CP treatment for 60s into the treatment solution, performing enzymolysis at 50deg.C for 1 hr, inactivating enzyme at 90deg.C for 15min, and filtering with 200 mesh filter cloth. Obtaining the flaxseed plant milk
As shown in fig. 6A and 6B, the effect of using the three techniques alone is limited, but the three techniques exhibit synergy when used in combination. On one hand, the release efficiency of flaxseed plant milk solids and protein is improved by low-temperature plasma application, and on the other hand, the contact of enzyme and substrate is promoted and the enzymolysis effect is further improved by ultrasonic treatment of plant milk. Finally, the flax seed milk prepared by low-consumption physical double-field treatment contains higher protein and total solid content, and the edible quality of the flax seed plant milk is greatly improved.
4.3.2 double field (low temperature plasma and pulse electric field) coupling, the method for synergistically improving the quality of flaxseed plant milk comprises the following steps:
(1) Preparing an enzyme solution: 15g of cellulase CTS is weighed, 50mL of water is added for uniform mixing, and the mixture is stirred for 30min in an ice bath.
(2) Low temperature plasma treatment of enzyme solution: 1mL of enzyme solution is sucked into a clean culture dish (diameter of 35 mm) and marked, and is placed below a nozzle of a plasma generator, the pulse discharge voltage is 18kV, the frequency is 1000Hz, and the pulse width is 200ns. When the plasma is stable (i.e. no obvious filiform spark exists), a timer is started, the treatment time is set to 15s, and the collected and treated cellulase is sealed and stored at the temperature of 4 ℃ for standby.
(3) Preparing flax seed homogenate: mixing and soaking the microwave degummed flax seeds and water for 2 hours according to the mass ratio of 1:7, discarding the soaking liquid, and collecting the plant milk after 12 minutes of colloid mill circulation.
(4) Pretreatment of flax seed milk by a pulsed electric field: carrying out pulsed electric field and ultrasonic treatment on the prepared flaxseed homogenate, wherein the electric field strength of the pulsed electric field treatment is 3.7kV/cm, the pulse frequency is 3 times, the pulse width is 500 mu s, the frequency is 1kHz, and the flow speed is 40rpm, so as to obtain a treatment liquid;
(5) The enzymolysis process comprises the following steps: adding 0.1wt% of cellulase treated by CP into the treatment liquid, performing enzymolysis at 50deg.C for 30min, inactivating enzyme at 90deg.C for 15min, and filtering with 200 mesh filter cloth to obtain flaxseed plant milk.
As shown in fig. 7, the three techniques have limited effects when used alone, but exhibit synergistic effects when the three techniques are used in combination. As shown in fig. 7A, the total phenol content of the flaxseed milk after PEF, enzymatic and CP treatments was increased by 21.07%, 12.98% and 28.26%, respectively, while the total phenol content was increased by 65.50% when the two-field complex coupled enzymatic method was used. As shown in FIG. 7B, the viscosity also shows a synergistic effect, and finally, the viscosity of the two-field compound coupling enzyme method is reduced to 77.47 mPa.s.
While embodiments of the present invention have been shown and described above, it will be understood that the above embodiments are illustrative and not to be construed as limiting the invention, and that variations, modifications, alternatives and variations may be made to the above embodiments by one of ordinary skill in the art within the scope of the invention.
Claims (3)
1. The method for improving the efficiency of the plant lactosis by the low-consumption physical double-field synergy is characterized by comprising the following steps of:
(1) Preparing an enzyme solution: weighing enzyme, adding water, mixing, and stirring in ice bath to obtain enzyme solution; the enzyme is one or more of cellulase, protease, phytase, hemicellulase, saccharifying enzyme and bromelain; the mass ratio of the enzyme to the water is 1-5:10;
(2) Low temperature plasma treatment: sucking enzyme liquid into a clean culture dish, placing the culture dish below a nozzle of a plasma generator, treating the enzyme liquid with stable plasma, collecting the treated enzyme liquid, and sealing and preserving at 2-8deg.C for later use; the pulse discharge voltage of the low-temperature plasma treatment is 16-20kV, the frequency is 200-2000Hz, the pulse width is 100-500ns, and the treatment time is 15-60s;
(3) Preparing flax seed homogenate: mixing the microwave degummed flax seeds with water, soaking, discarding the soaking solution, adding water into the soaked flax seeds, circulating for 4-12min by a colloid mill, and collecting plant milk;
(4) Pretreatment of flax seed homogenate by a pulsed electric field: performing pulsed electric field treatment on the plant milk prepared in the step (3) to obtain a treatment liquid; the electric field strength of the pulsed electric field treatment is 1.3-3.7kV/cm, the pulse times are 3-6 times, the pulse width is 100-500ns, the frequency is 500-1000Hz, and the flow velocity is 40-80rpm;
(5) Enzymolysis: adding 0.1-0.5wt% of the enzyme solution prepared in the step (2) into the treatment solution, inactivating enzyme after enzymolysis, and filtering to obtain the flaxseed plant milk with high protein content, high solid content and moderate viscosity.
2. The method for improving the efficiency of plant lactosis by low-consumption physical double-field synergy according to claim 1, wherein the mass ratio of the microwave degummed flax seeds to water in the step (3) is 1:5-10, and the soaking time is 0.5-3h.
3. The method for improving the enzymolysis efficiency of the plant milk by low-consumption physical double-field synergy according to claim 1, wherein the enzymolysis temperature in the step (5) is 45-55 ℃ and the enzymolysis time is 30-75min; the enzyme deactivation temperature is 90-100 ℃, and the enzyme deactivation time is 10-15min; the filtering and sieving granularity is 120-200 meshes.
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