CN106754835B - Cold-resistant pseudomonas extracellular protease inactivation method, raw milk and treatment method - Google Patents
Cold-resistant pseudomonas extracellular protease inactivation method, raw milk and treatment method Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/52—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from bacteria or Archaea
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C3/00—Preservation of milk or milk preparations
- A23C3/02—Preservation of milk or milk preparations by heating
Abstract
The invention discloses a method for inactivating cold-resistant pseudomonas extracellular protease, which comprises the following steps: heating the solution containing cold-resistant pseudomonas extracellular protease in a hot bath at 50-70 ℃ for 2-4min, and then heating by 800W microwave for 60-120 s. Compared with the low-temperature inactivation method, the extracellular protease inactivation method greatly shortens the treatment time, reduces the energy consumption, has the inactivation effect superior to that of the traditional method, and does not damage other nutrient substances in the solution environment of the extracellular protease.
Description
Technical Field
The invention belongs to the field of enzyme inactivation of microorganisms, and particularly relates to a method for inactivating cold-resistant pseudomonas extracellular protease. In addition, the invention also relates to a method for processing the raw milk and the raw milk obtained by processing the method.
Background
Pseudomonas is the predominant cold-resistant flora commonly found in raw milk. The cold-resistant pseudomonas can secrete heat-resistant protease in the refrigeration process of raw milk, the protease is difficult to inactivate by high-temperature heating, and the heat resistance mechanism is as follows: after high temperature treatment, the intact protease molecules in the unfolded state (inactive) refold into an active native conformation. Therefore, the enzyme can resist pasteurization (72 ℃/15s) or ultra-high temperature (120-150 ℃/0.5-8.0 s) sterilization process, and can not be completely inactivated by single heat treatment.
The heat-resistant protease causes the physical and chemical properties of the raw milk to be obviously changed mainly by degrading casein, and comprises the steps of raw milk deposition, reduction of Zeta potential and casein micelle hydration and release of polypeptide, so that the raw milk is deteriorated, and the shelf life of subsequent milk products is influenced.
The currently adopted cold-resistant bacterium protease inactivation methods comprise a high-temperature inactivation method and a low-temperature inactivation method, and the high-temperature inactivation method can destroy the nutritional ingredients of the dairy product due to high temperature; and the low-temperature inactivation method has overlong treatment time and large energy consumption. Therefore, how to rapidly inactivate the extracellular protease of cold-tolerant pseudomonas without destroying the nutritional ingredients of milk is a problem to be solved by those skilled in the art.
Disclosure of Invention
In order to solve the technical problems, the invention provides a novel method for inactivating extracellular protease of cold-resistant pseudomonas, which can avoid destroying the nutrient components of raw milk at high temperature, greatly shorten the time of low-temperature treatment and reduce the energy consumed in the inactivation process.
Specifically, in one aspect, a method for inactivating cold-resistant pseudomonas extracellular protease is provided, which comprises the following steps: heating the solution containing cold-resistant pseudomonas extracellular protease in a hot bath at 50-70 ℃ for 2-4min, and then heating by 800W microwave for 60-120 s.
Further, the microwave heating time was 90 seconds.
Further, the temperature of the heating bath was 55 ℃ and the time was 3.5 min.
Further, the heat bath is a metal bath.
In a second aspect, a method for processing raw milk is provided, which comprises the following steps: heating the raw milk at 50-70 deg.C for 2-4min, and microwave heating at 800W for 60-120 s.
In a third aspect, a raw milk is provided, which is obtained after being processed by the processing method.
Furthermore, the number of cold-resistant pseudomonas bacteria in the raw milk is less than 105cfu/ml。
The method for inactivating cold-resistant pseudomonas extracellular protease comprises the steps of heating a solution containing the cold-resistant pseudomonas extracellular protease in a hot bath at 50-70 ℃ for 2-4min to ensure that the extracellular protease is automatically degraded under the condition of being in the optimal inactivation temperature range, heating the solution for 60-120s by using microwave 800W to promote intramolecular vibration in the solution, destroy the space folding of the protease, accelerate the combination of the protease peptide chain and the autocatalytic site, promote the breakage of the peptide chain and the leaving of the degraded peptide segment from the reaction site, and further accelerate the complete inactivation of the extracellular protease.
In addition, compared with the low-temperature inactivation method, the extracellular protease inactivation method greatly shortens the treatment time, reduces the energy consumption, has an inactivation effect superior to that of the traditional method, and does not damage other nutrient substances in the solution environment of the extracellular protease.
Detailed Description
In order to more clearly illustrate the technical solution of the present invention, the technical solution of the present invention will be further illustrated with reference to the following examples.
Unlike common extracellular enzymes, thermostable proteases secreted by cold-resistant pseudomonas are difficult to completely inactivate by the existing high-temperature or low-temperature inactivation methods, and also difficult to rapidly inactivate by the conventional microwave heating method, and the conventional microwave method has low enzyme inactivation efficiency and long time, and also destroys the nutritional ingredients of cow milk like high-temperature inactivation. Therefore, the inventor develops a new method for inactivating the cold-resistant pseudomonas extracellular protease by creative work, which comprises the following steps: heating the solution containing cold-resistant pseudomonas extracellular protease in a hot bath at 50-70 ℃ for 2-4min, and then heating by 800W microwave for 60-120 s.
The method for inactivating cold-resistant pseudomonas extracellular protease comprises the steps of firstly heating a solution containing the cold-resistant pseudomonas extracellular protease at 50-70 ℃ for 2-4min to ensure that the extracellular protease is automatically degraded under the condition of being in the optimal inactivation temperature range, then heating the solution by microwave 800W for 60-120s to promote intramolecular vibration in the solution, destroy the space folding of the protease, accelerate the combination of the protease peptide chain and the autocatalytic site, promote the breakage of the peptide chain and the leaving of the degraded peptide segment from the reaction site, and further accelerate the complete inactivation of the extracellular protease. Compared with the low-temperature inactivation method, the extracellular protease inactivation method greatly shortens the treatment time, reduces the energy consumption, has the inactivation effect superior to that of the traditional method, and does not damage other nutrient substances in the solution environment of the extracellular protease.
It should be noted that in the above inactivation method, the microwave power is about 800W, such as 750W and 850W, which has a certain effect of inactivating the cold-resistant Pseudomonas extracellular protease, and preferably 800W, which is relatively more effective. On the basis of not influencing the core idea of the invention, the microwave power is reasonably adjusted and is regarded as an equivalent replacement for the technical characteristics.
Further, the microwave heating time is preferably 90 seconds. The temperature of the heating bath is preferably 55 ℃; the heating time of the heat bath is preferably 3.5 min. It can also be seen from the comparison of the examples that the inactivation effect of the cold-resistant pseudomonas extracellular protease is better under the preferable heating time and temperature.
The heat bath is that the container is placed in a heat bath medium, and the temperature heat of the heat bath medium is transferred to the substances in the container through the container, so that the purpose of heating the substances in the container is achieved. The common hot bath includes water bath, oil bath, sand bath, lead bath, etc., and in the method for inactivating cold-resistant pseudomonas extracellular protease, a metal bath is preferably used, so that the metal bath heating control is more convenient and accurate.
In another specific embodiment, a method for processing raw milk is further provided, which is characterized by comprising the following steps: heating the raw milk at 50-70 deg.C for 2-4min, and microwave heating at 800W for 60-120 s.
By adopting the method for treating the raw milk, on one hand, the nutrient substances such as vitamins, proteins and the like in the raw milk are prevented from being damaged by a high-temperature process, on the other hand, the inactivation effect on extracellular protease is superior to that of other traditional methods, and compared with the traditional methods, the method shortens the treatment time, reduces the energy consumption and is very suitable for being applied to the industrial production of the raw milk.
In another specific embodiment, raw milk obtained by the method for processing raw milk is also provided. The method for processing the raw milk has the beneficial effects, and the raw milk obtained by the method has the beneficial effects correspondingly, so the method is not repeated.
Further, the number of cold-resistant pseudomonas bacteria in the raw milk is less than 105cfu/ml. The processes of low-temperature heating and microwave heating treatment not only inactivate the extracellular protease generated by the cold-resistant pseudomonas, but also kill the cold-resistant pseudomonas, thereby ensuring that the number of the cold-resistant pseudomonas in the raw milk after treatment is less than 105cfu/ml。
In the following examples, cold-tolerant Pseudomonas strains (Pseudomonas sp), numbered 948, 1034, 1035, 1044, M73, M35, D22, D81, M73, N33, M55, J935, M97 and D72, are specific strains that produce extracellular protease in the examples, but are not involved in the inactivation method of the present invention, so that the specific strains do not affect the implementation of the present invention.
The MA solution in the following examples contains azocasein 5.0g/l, 3-morpholinopropanesulfonic acid buffer MOPS (pH 6.7)50mM, calcium chloride 1 mM. Other reagents and laboratory equipment not specifically described are commercially available.
Example 1
mu.L of the rejuvenated strain of Pseudomonas psychrotoleralis 1034, 1035, 1044, M73 and 948 was inoculated into 1mL of a 10g/L sterilized skimmed milk powder solution in an EP tube and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 55 deg.C for 3.5min, quickly transferring to microwave oven (Mei microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 90S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
Three groups of samples were each quenched with 20. mu.L of 2M trichloroacetic acidAfter centrifugation at 10000rpm at 25 ℃ for 15min, 150. mu.L of the supernatant was transferred to a 96-well cell culture plate and neutralized with 50. mu.L of 1M sodium hydroxide solution. The 96-well cell culture plate was placed in a microplate reader to measure the absorbance at 450 nm. And subtracting the light absorption value of the same blank sample (the bacterial liquid supernatant is replaced by deionized water) from the light absorption value of the reaction time of 2h to obtain a light absorption value increment (DA). The hourly increase in absorbance per ml of sample at 450nm is the protease activity (. DELTA. A h)-1mL-1)。
The protease activity of the strain is less than 0.3 delta A h-1ml-1The protease is considered to be completely inactivated.
Each test was performed twice and the average was taken. The residual enzyme activity calculation method is that the protease activity values of the control sample 2 and the test sample are divided by the protease activity value of the control sample 1 respectively to obtain the percentage of the residual enzyme activity as the respective residual enzyme activity value.
The results of the measured values of the residual enzyme activities are shown in the following table 1:
TABLE 1
As can be seen from the results in Table 1, the inactivation method treatment time for the test sample was only 5min, which is 1/2 times the treatment time for control sample 2. Moreover, for 5 strains 1034, 1035, 1044, M73 and 948, the corresponding extracellular enzymes can be inactivated by the inactivation method of the invention, and the residual activities of the proteases produced by other 4 strains except the sterilized strain 1044 are all less than those of the protease produced by the low-temperature inactivation method, which shows that the inactivation effect of the invention is far better than that of the low-temperature inactivation method.
Example 2
20. mu.L of the rejuvenated strain of Pseudomonas psychrophila M35, D22, D81, M73, N33, M55, M97 and D72 was inoculated into an EP tube containing 1mL of a 10g/L sterilized skim milk powder solution and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 55 deg.C for 3.5min, quickly transferring to microwave oven (Mei microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 90S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
The method for determining the residual enzyme activity value was the same as in example 1, and the results are shown in Table 2 below:
TABLE 2
As can be seen from the results in Table 2, the inactivation method treatment time for the test sample was only 5min, which is 1/2 times the treatment time for control sample 2. Moreover, for 8 strains M35, D22, D81, M73, N33, M55, M97 and D72, the corresponding extracellular enzymes can be inactivated by the inactivation method of the invention, and the residual activity of the protease generated by the 8 strains is far less than that of the protease generated by the low-temperature inactivation method, which shows that the inactivation effect of the invention is far better than that of the low-temperature inactivation method, and especially for the strains D22, D72, M55, N33 and the like, the inactivation effects are respectively about 7 times, 3 times, 1.7 times and 1.5 times of that of the low-temperature inactivation method.
Example 3
mu.L of the rejuvenated strain of Pseudomonas psychrotoleralis 1034, 1035, 1044 and M73 was inoculated into 1mL of sterilized skimmed milk powder solution at a concentration of 10g/L in an EP tube and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 55 deg.C for 2min, quickly transferring to microwave oven (American microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 90S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
The method for determining the residual enzyme activity value was the same as in example 1, and the results are shown in Table 3 below:
TABLE 3
As can be seen from the results in Table 3, the treatment time of the inactivation method for the test samples was shortened to 2min, and for the 4 strains 1034, 1035, 1044 and M73, the corresponding extracellular enzymes were inactivated by the inactivation method of the present invention, and the residual activities of the proteases produced by the 3 strains other than the strain 1044 were less than those produced by the low-temperature inactivation method. However, compared with the optimal condition inactivation effect of example 1, the residual activities of all the strains are greater than the enzyme activities after the treatment under the optimal inactivation condition, which indicates that the condition inactivation effect is inferior to the inactivation effect under the optimal condition.
Example 4
mu.L of the rejuvenated strain of Pseudomonas psychrophila N33 was inoculated into 1mL of sterilized skimmed milk powder solution at a concentration of 10g/L in an EP tube, and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 70 deg.C for 2min, quickly transferring to microwave oven (American microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 90S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
The method for determining the residual enzyme activity value was the same as in example 1, and the results are shown in Table 4 below:
TABLE 4
As can be seen from the results in Table 4, the residual activity of the extracellular protease produced by N33 was slightly lower than that of the low-temperature inactivation method by microwave treatment immediately after heating at 70 ℃ for 2min, but the two methods were not significantly different. Compared with the optimal condition inactivation effect of the embodiment 1, the activity of the enzyme is far higher than that of the residual enzyme treated by the optimal condition. This indicates that the condition deactivation effect is inferior to the optimal condition deactivation effect, but the condition deactivation effect is equivalent to the effect of the traditional low-temperature deactivation, and the condition deactivation effect and the traditional low-temperature deactivation effect are not obviously different.
Example 5
mu.L of the rejuvenated strain of Pseudomonas psychrophila 948 was inoculated into 1mL of sterilized skimmed milk powder solution (10 g/L) in an EP tube and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 70 deg.C for 4min, quickly transferring to microwave oven (American microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 120S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
The method for determining the residual enzyme activity value was the same as in example 1, and the results are shown in Table 5 below:
TABLE 5
As can be seen from the results in Table 5, the residual activity of the extracellular protease produced by the cold-resistant Pseudomonas 948 was slightly higher than that of the low-temperature inactivation method by the microwave treatment for 120s immediately after heating at 70 ℃ for 4min, but the two methods were not significantly different. Compared with the optimal condition inactivation effect of the embodiment 1, the activity of the enzyme is far higher than that of the residual enzyme treated by the optimal condition. This indicates that the condition deactivation effect is inferior to the optimal condition deactivation effect, but the condition deactivation effect is equivalent to the effect of the traditional low-temperature deactivation, and the condition deactivation effect and the traditional low-temperature deactivation effect are not obviously different.
Example 6
mu.L of the rejuvenated strain of Pseudomonas psychrophila 948 was inoculated into 1mL of sterilized skimmed milk powder solution (10 g/L) in an EP tube and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 70 deg.C for 4min, quickly transferring to microwave oven (American microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 120S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
The method for determining the residual enzyme activity value was the same as in example 1, and the results are shown in Table 6 below:
TABLE 6
As can be seen from the results in table 6, for extracellular protease produced by the strain 948, the residual activity is greater than the enzyme activity after the optimal inactivation condition treatment in example 1, and is equivalent to the enzyme activity after the low-temperature inactivation method treatment, which indicates that the inactivation effect under the conditions is inferior to the optimal inactivation effect, but is equivalent to the traditional low-temperature inactivation effect, and the two effects have no significant difference.
Example 7
mu.L of the rejuvenated strain of Pseudomonas psychrophila J935 was inoculated into an EP tube containing 1mL of a 10g/L sterilized skimmed milk powder solution and cultured at 6 ℃ for 8 days. The cultured bacterial suspension was centrifuged at 12000rpm at 25 ℃ for 15 min.
Control sample 1: mu.L of the supernatant was added to 100. mu.L of MA solution and reacted at 40 ℃ for 2 hours.
Control sample 2: 100 mu L of supernatant was put in a metal bath, heated at 55 ℃ for 10min, cooled at 4 ℃ for 30S, added to 100 mu L of MA solution, and reacted at 40 ℃ for 2 h.
The test sample is prepared by heating 100 μ L of supernatant in metal bath at 50 deg.C for 2min, quickly transferring to microwave oven (American microwave oven, rated voltage 220V-50Hz, 2450MHz, cavity volume 352 × 325 × 202mm (23L)), heating at 800W for 90S, cooling at 4 deg.C for 30S, adding into 100 μ L MA solution, and reacting at 40 deg.C for 2 h.
The method for determining the residual enzyme activity value was the same as in example 1, and the results are shown in Table 7 below:
TABLE 7
As can be seen from the results in Table 7, for extracellular protease produced by the strain J935, the residual activity is greater than that of the enzyme treated under the optimal inactivation condition in example 1, but is much lower than that of the enzyme treated by the traditional low-temperature inactivation method, which indicates that the inactivation effect under the condition is inferior to that of the optimal condition and superior to that of the traditional low-temperature inactivation method.
In the above examples, in order to verify the effect of the inactivation method of the present invention, the test was performed by inoculating a high-concentration bacterial solution with a skim milk powder solution, and thus a part of the protease having the activity remained in the treated sample. When the processed sample is raw milk in normal industry, the concentration of the cold-resistant pseudomonas extracellular protease in the raw milk is far less than that in the embodiment, so that the inactivation method can inactivate the extracellular protease almost completely, thereby avoiding the raw milk deterioration caused by the extracellular protease and being beneficial to prolonging the shelf life of the raw milk.
Finally, it should be noted that: the above embodiments are only used to help understanding the technical solutions and core ideas of the present invention, and are not limited thereto; while the invention has been described in detail and with reference to the foregoing embodiments, it will be understood by those skilled in the art that: it is possible to modify the solutions described in the embodiments or to substitute some or all of the technical features of the embodiments, and these modifications or substitutions also fall within the scope of the claims of the present invention.
Claims (5)
1. A method for inactivating cold-resistant pseudomonas extracellular protease is characterized by comprising the following steps: heating the solution containing cold-resistant pseudomonas extracellular protease in a hot bath at 50-70 ℃ for 2-4min, and then heating by 800W microwave for 60-120 s.
2. The method for inactivating cold-tolerant pseudomonas extracellular protease, according to claim 1, wherein the microwave heating time is 90 s.
3. The method for inactivating cold-resistant pseudomonas extracellular protease, according to claim 1, wherein the heating bath is heated at 55 ℃ for 3.5 min.
4. The method of inactivating cold-tolerant pseudomonas extracellular protease of claim 1, wherein the hot bath is a metal bath.
5. A method for processing raw milk is characterized by comprising the following steps: heating the raw milk at 50-70 deg.C for 2-4min, and microwave heating at 800W for 60-120 s.
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