Pure microalgae cheese and preparation method thereof

Abstract

The invention relates to pure microalgae cheese. The pure microalgae cheese comprises microalgae milk, an acidifying agent and chymosin, the microalgae milk is pure microalgae milk prepared from microalgae raw materials, wherein protein accounts for 40-60% of the dry weight of cells, grease accounts for 8-16% of the dry weight of the cells, and starch accounts for 20-30% of the dry weight of the cells. The pure microalgae milk is adopted as a raw material to prepare the cheese, exogenous starch, fat or protein is not added, a pure microalgae cheese product which is comparable with traditional cheese in flavor and sensory characteristics can be obtained, and the pure microalgae cheese product is good in processing performance and excellent in storage stability; the technical problems that traditional plant-based cheese must be supplemented with a large amount of exogenous starch, fat or protein to adjust the composition structure of raw materials, and the plant-based cheese has a large gap with animal cheese in the characteristics of taste, stretching, melting property, stability and the like are solved. According to the invention, rich high-protein nutritional products are provided for lactose intolerance, animal protein allergy and vegetarians, and the technical blank of the food type of microalgae cheese is filled. The whole microalgae enters the cheese, so that the flavor level of the cheese is enriched, and the comprehensive health-care function of the cheese product is enhanced.

Description

Technical Field

[0001] The invention relates to the technical field of microalgae food, and in particular to pure microalgae cheese and a preparation method thereof. Background Art

[0002] Cheese is a high value-added deep-processed dairy product that is deeply loved by people. Traditional original cheese is a fresh or fermented and ripened dairy product made from cow (sheep) milk, cream, and partially skimmed milk, which is fermented by bacteria and (or) rennet coagulated and the whey is discharged. Traditional cheese is a viscoelastic food, and the changes in its texture characteristics are mainly affected by the casein network structure and fat globules. In animal milk, casein is combined with micellar phosphate and exists stably as a protein aggregate of casein micelles. During the cheese production process, casein micelles can be destabilized under heat treatment, rennet, acid and other conditions to form a milk gel, which is a key step in cheese production.

[0003] However, some people are intolerant or allergic to lactose or protein in animal dairy products, which has prompted people to look for plant-based dairy products. Traditional plant-based dairy products are made from plant ingredients such as soybeans, oats, peanuts, almonds, and rice. They contain a small amount of saturated fat and no cholesterol. These plant-based dairy products provide a rich choice of protein nutrition products for lactose intolerance, animal protein allergies, and vegetarians. Plant-based cheese is a dairy product that can provide similar functions and sensory properties to animal cheese. Plant-based cheese, also known as plant-based cheese or cheese analogs, is a uniform cheese-like matrix. It is currently mainly made of a uniform mixture of vegetable oil or fat, starch, protein and water. It is made by heating to promote starch gelatinization and protein denaturation, and through reasonable ingredient ratio adjustment, it is made to have similar functions and sensory properties to traditional cheese. In order to avoid meeting the needs of lactose intolerance, animal protein allergies, and vegetarians, the fat and protein sources used in plant-based cheese try not to use fat and protein in milk, but fat and protein from plant-based sources.

[0004] When making plant-based cheese, the roles of starch, protein and fat in cheese are as follows: ① Gelatinized starch can provide the necessary structure and viscosity for plant-based cheese, helping to simulate the gelatinous texture of traditional cheese. Starch helps stabilize the water and other ingredients in the mixture, prevent separation, and give the product the right consistency. This is very important for imitating the cutting and spreading properties of real cheese. The right amount of starch can give plant-based cheese a smooth and delicate taste; ② Protein is the key component for plant-based cheese to obtain elasticity and gelatinous texture similar to traditional cheese. Through heating and other processing conditions, plant proteins (such as soy protein, pea protein or wheat gluten) can form a network structure that helps simulate the chewing feel of milk cheese. Plant protein can help evenly disperse oil, making the product more delicate and smooth. ③ Fat is one of the key factors in giving food a rich taste. It can provide a smooth texture and is a good medium for carrying flavor substances. Therefore, for plant-based cheese, fat helps to enhance the overall flavor experience. Similar to animal cheese, the fat in plant-based cheese also determines its melting behavior. The right amount of fat allows plant-based cheese to exhibit good melting properties when heated.

[0005] However, the proportion of protein, starch, vegetable oil or fat in the seeds of traditional higher plant crops, such as soybeans, oats, peanuts, almonds, rice and other plant-based ingredients, is very fixed. It is difficult to change the composition structure of protein, starch, vegetable oil or fat in these ingredients by adjusting planting conditions, regulating metabolic enzymes or environmental stress. In addition, the protein emulsification and gel function of traditional plant ingredients are weak, which makes plant-based cheese still have a large gap in taste, stretchability, melting and stability compared with traditional animal cheese. For this reason, when using these ingredients to make plant-based cheese, a large amount of exogenous fat or starch must be added to adjust the composition structure of protein, starch, vegetable oil or fat in the raw materials, so as to simulate the sensory properties similar to traditional cheese as much as possible. However, in this way, a plant-based cheese product may be mixed with ingredients from multiple plant sources, and it is impossible to make cheese with only a single plant ingredient, which cannot meet the needs of some people. On the other hand, due to the weak coagulation of these plant proteins, even if the composition structure of the raw materials is adjusted by adding exogenous fat or starch, it is still difficult to simulate the properties similar to traditional animal cheese. Summary of the invention

[0006] 1. Technical issues to be resolved

[0007] In view of the above-mentioned disadvantages and deficiencies of the prior art, the present invention provides a method for making pure microalgae cheese, which uses pure microalgae milk as a raw material to make cheese, without adding exogenous starch, fat or protein, and can obtain a pure microalgae cheese product with flavor and sensory characteristics comparable to those of traditional cheese, and has good processing performance and excellent storage stability, solving the technical problems that traditional plant-based cheese must supplement a large amount of exogenous starch, fat or protein to adjust the composition structure of the raw materials, and there are significant gaps between the characteristics of plant-based cheese such as taste, stretchability, meltability and stability and those of animal cheese.

[0008] (II) Technical solution

[0009] In the first aspect, the present invention provides a pure microalgae cheese, which comprises: microalgae milk, an acidulant and rennet; the microalgae milk is a pure microalgae milk made from a microalgae raw material with protein accounting for 40-60% of the cell dry weight, oil accounting for 8-16% of the cell dry weight, and starch accounting for 20-30% of the cell dry weight.

[0010] According to a preferred embodiment of the present invention, the acidulant is at least one of hydrochloric acid, phosphoric acid, lactic acid, acetic acid, citric acid, rice vinegar, lemon juice, lemon malic acid, sorbic acid.

[0011] According to a preferred embodiment of the present invention, the microalgae raw material is any one or several of Chlamydomonas reinhardtii, Chlorella vulgaris, Scenedesmus, Euglena gracilis, Spirulina platensis, Dunaliella salina, Nannochloropsis oculata, and any high-protein mutant strain or chlorophyll-deficient strain of any of the foregoing.

[0012] According to a preferred embodiment of the present invention, the protein content is 1.7-4.0 g / 100 ml, the fat content is 0.2-0.8 g / 100 ml, and the starch content is 0.6-1.1 g / 100 ml.

[0013] When preparing plant-based cheese, when the mass ratio of oil to starch in the raw materials is close to 1:1, the texture characteristics of the plant-based cheese are the best; while using microalgae milk prepared from a microalgae raw material with a specific nutritional structure composition, the mass ratio of the oil to the starch contained therein basically conforms to this ratio, and at the same time has a high protein content and high nutritional value.

[0014] It should be noted that the method for making microalgae milk refers to the method described in Chinese Patent Application Publication No. CN118160787A "A Microalgae Plant Milk Beverage and Its Making Method", but the hydrolysis degree of protein and starch needs to be controlled to provide certain emulsifying properties, so that the pure microalgae cheese has elasticity similar to that of traditional cheese and has a gel-like texture; the hydrolysis degree of starch should be appropriately controlled to make the microalgae cheese have a sweet taste, and can imitate the cutting characteristics and spreading characteristics of real cheese, have a smooth and delicate taste, and simulate the chewing feeling of cow's milk cheese.

[0015] In a second aspect, the present invention also provides a method for making pure microalgae cheese, which includes:

[0016] S1. Select microalgae raw materials with protein accounting for 40-60% of the cell dry weight, oil accounting for 8-16% of the cell dry weight, and starch accounting for 20-30% of the cell dry weight to make microalgae milk;

[0017] S2. Add an acidifying agent to the microalgae milk, keep it warm for acidification treatment at 80-90 °C for 10-20 min, and control the pH at the end of acidification to be 4.3-4.8; then cool it to below 40 °C, add rennet, stir well, and then let it stand to curdle;

[0018] S3. Keep it warm at 85-95 °C for 25-40 min again, then cool and let it stand to obtain a curd block. Cut the curd block, stir slowly, and remove the whey to obtain pure microalgae cheese.

[0019] According to a preferred embodiment of the present invention, in S1, the microalgae raw materials are any one or several of Chlamydomonas reinhardtii, Chlorella vulgaris, Scenedesmus, Euglena, Spirulina, Dunaliella salina, Nannochloropsis, and any high-protein mutant strain or chlorophyll-deficient strain of any of the foregoing.

[0020] Preferably, the chlorophyll-deficient microalgae are, for example, Chlorella pyrenoidosa with yellow protein or Chlorella pyrenoidosa with white protein. For example, Chlorella pyrenoidosa with yellow protein with the preservation number of CGMCC NO.40458 (see Patent CN116555040B) deposited by the applicant at the China General Microbiological Culture Collection Center can be selected; or Chlorella pyrenoidosa with white protein with the preservation number of CGMCC NO.40787, the preservation date is September 19, 2023, the taxonomic name is Chlorella pyrenoidosa, and the institution is the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, and the address is No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. These Chlorella pyrenoidosa have a high protein content, do not contain chlorophyll, can avoid the problem of browning of chlorophyll during high-temperature treatment or storage, and the chlorophyll-deficient Chlorella does not have a metallic fishy smell, nor does it produce phytol / phytadiene with a fishy smell generated by the cleavage of the chlorophyll side chain, effectively reducing the intensity of the fishy smell and improving the taste of the cheese.

[0021] According to a preferred embodiment of the present invention, in S1, the method for making microalgae milk is as follows:

[0022] Step 1: Select microalgae raw materials with protein accounting for 40-60% of the cell dry weight, oil accounting for 8-16% of the cell dry weight, and starch accounting for 20-30% of the cell dry weight;

[0023] Step 2: Adjust it into algal paste with drinking water to make the dry matter concentration of algal cells be 100 g / L - 200 g / L, and use a high-pressure homogenizer to break the cell walls of the algal cells to obtain broken-wall algal paste; after homogenization, the particles below 1 μm in the cell paste account for more than 90% of the total number of particles, and the particles below 300 nm account for more than 50% of the total number of particles.

[0024] Step 3: Dilute the broken-wall algal paste with water until the microalgae dry matter content is 20 g / L - 100 g / L, and based on the weight of the diluted algal paste, add 350 - 5000 units / kg of cellulase, 500 - 3000 units / kg of amylase, 1000 - 3000 units / kg of glucoamylase, and 350 - 1000 units / kg of protease in sequence for enzymatic hydrolysis. Control the enzymatic hydrolysis conditions to make the degree of starch hydrolysis be 25 - 40% and the degree of protein hydrolysis be 20 - 30% to obtain an enzymatic hydrolysis mixture.

[0025] Step 4: Heat the enzymatic hydrolysis mixture obtained in Step 3 to 50 - 80 °C, and transfer it into a high-pressure homogenizer again for at least one homogenization treatment. After sterilization, microalgae milk is obtained. The protein content in the microalgae milk is 1.7 - 4.0 g / 100 ml, the fat content is 0.2 - 0.8 g / 100 ml, and the starch content is 0.6 - 1.1 g / 100 ml.

[0026] By controlling an appropriate degree of starch hydrolysis, on the one hand, the starch undergoes a hydrolysis reaction to generate dextrin, oligosaccharide, oligoglycan, and a small amount of glucose or maltose to improve the food taste and increase the sweetness, etc. On the other hand, by controlling the degree of starch hydrolysis, part of the starch is retained to meet the gelatinization function of the starch in cheese preparation and promote cheese coagulation. By controlling an appropriate degree of protein hydrolysis, while ensuring the protein content, part of the protein undergoes hydrolysis to meet the requirement of the emulsifying property of microalgae milk during the production of microalgae cheese, making the prepared cheese more elastic. Control the hydrolysis degrees of protein and starch, as well as the protein, fat, and starch contents contained in the microalgae milk, so that the microalgae milk meets the various index requirements when making cheese.

[0027] The principle of the sequential enzymatic hydrolysis according to the above is as follows: First, add cellulase for enzymatic hydrolysis to hydrolyze the microalgae cell wall, reduce the roughness, and make the cheese softer; then add amylase and glucoamylase for hydrolysis to increase the sugar content and sweetness; finally, use protease for hydrolysis to obtain peptide segments with good emulsifying properties, and at the same time retain the complete structure of most proteins, making the cheese have gelation and chewiness. By hydrolyzing starch and crude fiber in advance, the inhibition of the emulsifying property of peptide segments by starch and crude fiber is reduced. Protease needs to be added last to avoid its degradation and inactivation of other enzymes.

[0028] According to a preferred embodiment of the present invention, in S2, the acidifying agent is at least one of hydrochloric acid, phosphoric acid, lactic acid, acetic acid, citric acid, rice vinegar, lemon juice, lemon malic acid, and sorbic acid.

[0029] According to a preferred embodiment of the present invention, in S2, the addition amount of rennet is 1 - 15 g of rennet / 1000 L of microalgae milk. Preferably, rennet is added to the microalgae milk that has reached the acidification end point and cooled to below 40°C. Various conventional rennets in the cheese field can be selected as rennet, such as one or more of animal rennet, plant rennet, and microbial rennet; the animal rennet is preferably chymosin and / or pepsin; the plant rennet is preferably one or more of ficin, papain, and bromelain; more preferably, it is microbial rennet.

[0030] The fungal sources of microbial rennet include Mucor miehei (which is an acidic protease, and the trade names include )), Mucor pusillus, Cryphonectria parasitica (non-allergenic, suitable for halal food), the bacterial sources of microbial rennet include Bacillus subtilis (alkaline rennet), Bacillus licheniformis; the yeast sources of microbial rennet include Kluyveromyces (no animal-derived components, meeting the needs of vegetarianism, commonly used in mozzarella cheese), Pichia pastoris. Preferably, Kluyveromyces enzyme (abbreviated as KluC rennet, non-animal-derived and low allergenicity, suitable for organic / vegetarian cheese) or Mucor or recombinant Pichia pastoris enzyme (high C / P value, reducing excessive protein hydrolysis, commonly used in the production of hard cheese) is used.

[0031] According to a preferred embodiment of the present invention, in S2, citric acid is added to the microalgae milk, and acidification treatment is carried out at a constant temperature in a water bath at 90°C for 10 min, and the pH at the acidification end point is controlled to be 4.6; then it is cooled to below 40°C, rennet is added, and after stirring evenly, it is left standing for 2 h to curdle; then in S3, it is kept at a constant temperature in a water bath at 90°C for 30 min, and then cooled and left standing to obtain a curd block. The curd block is cut, centrifuged, and the whey is removed to obtain pure microalgae cheese.

[0032] According to a preferred embodiment of the present invention, in S3, the operating method of cutting the curd block and slowly stirring is: cutting is carried out immediately after curdling is completed, slowly stirring (to discharge the whey wrapped in the curd), heating to 39°C and then keeping warm for 30 - 45 min, and the whey is removed after the curd heals.

[0033] According to a preferred embodiment of the present invention, in S3, the method for removing the whey is: filtering using a food-grade gauze, cloth bag, or cheesecloth bag, or centrifugal separation is carried out after stirring, the centrifugal force is 4000 - 5000 g, and the centrifugation treatment is carried out for 10 - 20 min.

[0034] In S2, after adding an acidifying agent to the microalgae milk, the microalgae milk is first heated to about 90 °C, and the microalgae milk is subjected to strong heat treatment to promote the acidification of the microalgae milk by the acidifying agent, so that the protein in the microalgae milk precipitates to form an acid / heat gel, and the starch gelatinizes under strong heat treatment; further, rennet is added after cooling (cooling avoids inactivation of rennet), stirred evenly, and left standing to promote curdling. In S3, strong heat treatment is carried out again to further promote coagulation to obtain curd blocks with higher strength. At this time, whey (liquid part) is separated from the curd. After removing the whey by cutting the curd, slow stirring, filtration / centrifugation, etc., the pure microalgae cheese is obtained.

[0035] The following are the descriptions of each step in the conventional cheese-making process.

[0036] Acidification: Acidification promotes the denaturation and coagulation of milk proteins into curds by lowering the pH value of the milk.

[0037] Acidification not only changes the physical properties of the cheese but also has a profound impact on its taste. The presence of acid promotes the formation of certain metabolites, such as lactic acid, which plays an important role in the development of the characteristic sour taste and other complex flavors of the cheese. In addition, acidity can inhibit the growth of harmful microorganisms while facilitating the reproduction of beneficial bacteria, which produce more flavor compounds during cheese ripening. The lower pH environment is not conducive to the growth of many pathogens and spoilage bacteria, which helps to extend the shelf life of the cheese. Moreover, proper acidification can make the cheese more stable and less likely to deteriorate during storage and transportation.

[0038] Rennet: Under normal conditions, some proteins exist in milk in the form of micelles, which carry negative charges and repel each other to remain in suspension. When an acid or an enzyme (such as rennet) is added, they can neutralize these charges, causing the micelles to aggregate together to form large aggregates, that is, "curds" are formed.

[0039] Whey separation: As the proteins coagulate, whey (liquid part) separates from the curds. This process helps to reduce the water content of the final product, thereby affecting the hardness and ripening rate of the cheese.

[0040] (III) Beneficial effects

[0041] 1. After continuous improvement and optimization of the culture conditions, microalgae have become a protein-rich food raw material and a source of high-quality protein. Currently, various microalgae have been listed as human health foods in the 21st century by the Food and Agriculture Organization of the United Nations. The present invention uses microalgae as the raw material to produce pure microalgae cheese, which is not only the development and utilization of this new food resource, enriching the types of plant-based cheese products, but also providing lactose-intolerant, animal protein-allergic, and vegetarian people with a rich choice of high-protein nutritional products, filling the technical gap in the food category of microalgae cheese. The whole microalgae is incorporated into the cheese without the need for extraction and separation of algal components, which can save production costs, reduce the residue and pollution of extractants, fully retain algal proteins, starch, fat, dietary fiber, natural antioxidants, carotenoids, vitamins, and trace elements, reduce the waste of biomass resources, enrich the flavor level of the cheese, and enhance the comprehensive health care function of the cheese product.

[0042] 2. Compared with traditional higher crops such as soybeans, oats, peanuts, almonds, rice and other plant-based food ingredients, microalgae, as single-celled organisms, have the advantages of rapid cultivation, short cultivation cycle, and the composition structure of intracellular nutrients such as proteins, starch, fatty acids / oils is very easy to be directionally regulated. For example, metabolic enzymes of certain pathways are overexpressed / silenced, and methods such as light / dark induction, medium induction, environmental stress, heterotrophic / autotrophic are used to change the composition structure of proteins, starch, vegetable oil or fat in microalgae cells, so that the composition structure of the obtained microalgae raw material nutrients exactly meets the requirements of the raw material composition for cheese making. Using the whole components of microalgae to make cheese can produce pure microalgae cheese with similar characteristics (taste, stretchability, meltability, and stability, etc.) to traditional animal cheese without supplementing a large amount of exogenous proteins, starch, fat / oils, etc. The microalgae cheese of the present invention not only has sensory characteristics comparable to traditional cow milk cheese, but also has a single and clean raw material, excellent processing performance and storage stability.

[0043] 3. The experiment also proves that microalgae protein is different from the proteins of higher plants such as beans and almonds. Microalgae protein has good self-coagulability, so rennet and the like can be used to complete coagulation better, making the produced microalgae cheese have obvious improvements in characteristics such as taste, stretchability, meltability, and stability compared with traditional plant-based cheese, and being closer to traditional animal cheese, solving the technical problems that when using beans and almonds to make plant-based cheese, it is necessary to adjust the proportion of raw material components by supplementing fat and starch, and due to the weak emulsifying and gelling properties of plant proteins, there is a large gap in characteristics such as taste, stretchability, meltability, and stability between plant-based cheese and traditional animal cheese. Description of the Drawings

[0044] Figure 1 Coagulation conditions for Comparative Example 1 and Example 1.

[0045] Figure 2 For the curdling conditions of Comparative Example 2, Comparative Example 3, and Comparative Example 4.

[0046] Figure 3 For the whey removal of the curds obtained from Comparative Example 1 and Example 1.

[0047] Figure 4 For the cheese products obtained from Comparative Example 1 and Example 1.

[0048] Figure 5 For the cheese products obtained from Comparative Example 5 and Comparative Example 6.

[0049] Figure 6 For the odor comparison results between the cow milk cheese of Comparative Example 1 and the microalgae cheese of Example 1

[0050] Figure 7 For the taste comparison results between the cow milk cheese of Comparative Example 1 and the microalgae cheese of Example 1.

[0051] Figure 8 For the comparison results of the conventional cheese indices between the cow milk cheese of Comparative Example 1 and the microalgae cheese of Example 1. Detailed Embodiments

[0052] For better explaining the present invention for easy understanding, the present invention will be described in detail below with reference to the accompanying drawings through specific embodiments. It should be noted that on the basis of conforming to the common knowledge in the art, the specific implementation conditions of each step can be appropriately expanded and combined, but all do not deviate from the scope covered by the solution of the present invention.

[0053] Example 1

[0054] The raw materials for making pure microalgae cheese in this example are: microalgae milk, edible citric acid, and microbial rennet (KluC rennet).

[0055] The method for making microalgae milk is as follows:

[0056] Step 1: Select a microalgae raw material with 50% protein, 15% oil, and 26% starch in the dry cell weight (the rest are vitamins, minerals, natural pigments, water-soluble dietary fiber, and phytosterols, etc.). This microalgae raw material is the yellow Chlorella pyrenoidosa with the preservation number CGMCC NO.40458 preserved by the applicant at the China General Microbiological Culture Collection Center.

[0057] Step 2: Adjust it into an algal slurry with drinking water to make the dry matter concentration of algal cells 120 g / L, and use a high-pressure homogenizer to break the algal cells 2 times at a pressure of 120 MPa to obtain a broken algal slurry; after homogenization, the particles below 1 μm in the cell slurry account for 90% of the total number of particles, and the particles below 300 nm account for 54% of the total number of particles.

[0058] Step 3: Dilute the broken-wall algal paste with water until the microalgae dry matter content is 60 g / L. Based on the weight of the diluted algal paste, add 1000 units / kg of cellulase, 1200 units / kg of amylase, 2000 units / kg of glucoamylase, and 800 units / kg of protease in sequence for enzymatic hydrolysis. Control the enzymatic hydrolysis conditions to make the starch hydrolysis degree 30% and the protein hydrolysis degree 20% to obtain an enzymatic hydrolysis mixture.

[0059] Step 4: Heat the enzymatic hydrolysis mixture obtained in Step 3 to 80 °C (to complete enzyme inactivation and sterilization), then transfer it to a high-pressure homogenizer and homogenize it twice at 60 MPa. After pasteurization, microalgae milk is obtained. Adjust the protein content in the microalgae milk to 1.71 g / 100 ml, the fat content to 0.64 g / 100 ml, and the starch content to 0.78 g / 100 ml.

[0060] After physical and chemical and microbiological inspections, the above microalgae milk is qualified and can be used for cheese production. The cheese production process is as follows:

[0061] Heat the microalgae milk in a 90 °C water bath. While keeping it warm, acidify it with citric acid and make the pH value at the acidification end point 4.6; cool it to 40 °C, add KluC rennet, and the addition amount is 5 g of rennet / 1000 L of microalgae milk. Stir well and evenly, let it stand for 2 h to curdle; then place it in a 90 °C water bath for 30 min, and after cooling, a curd block is formed; cut the curd and stir it slowly at a speed of 10 rpm to drain the whey wrapped in the curd. After heating to 39 °C, keep it warm for 40 min. After the curd heals, drain the whey, and then filter it with a food-grade gauze to remove the whey to obtain fresh pure microalgae cheese.

[0062] Comparative Example 1

[0063] In this comparative example, the microalgae milk in Example 1 is replaced with milk, and other steps and conditions are the same as those in Example 1. The milk is a commercially available pure milk product of a certain brand, with a protein content of 3.4 g / 100 mL (the casein accounts for about 80%), a fat content of 3.6 g / 100 mL, lactose of 4.8 g / 100 mL, and starch of 0. After physical and chemical and microbiological inspections, it is qualified and can be used for making cheese.

[0064] The cow milk cheese production process is as follows:

[0065] Heat the pure milk in a 90 °C water bath. While keeping it warm, acidify it with citric acid and make the pH value at the acidification end point 4.6; cool it to 40 °C, add rennet, stir well and evenly, let it stand for 2 h to curdle; then transfer it to a 90 °C water bath for 30 min, and after cooling, a curd block is formed; cut the curd and stir it slowly at a speed of 10 rpm to drain the whey oozed out from the curd. After heating to 39 °C, keep it warm for 40 min. After the curd heals, drain the whey, and then filter it with a food-grade gauze to remove the whey to obtain fresh cheese.

[0066] Comparative Example 2

[0067] In this comparative example, the microalgae milk in Example 1 was replaced with oat milk, and the other steps and conditions were the same as those in Example 1. Oat milk was used as the raw milk, with a protein content of 1.5 g / 100 mL (oats naturally contain gluten, but the content is low), a fat content of 3.2 g / 100 mL (added rapeseed oil), a starch (calculated as total carbohydrates) content of 1.7 g / 100 mL (composed of residual and undigested starch after enzymatic hydrolysis), and a total carbohydrate content of 8.8 g / 100 mL. After passing physical and chemical and microbiological tests, it can be used to make cheese. The production process of oat cheese was the same as that in Example 1.

[0068] Comparative Example 3

[0069] In this comparative example, the microalgae milk in Example 1 was replaced with almond milk, and the other steps and conditions were the same as those in Example 1. Almond milk was used as the raw milk, with a protein content of 1.1 g / 100 mL (almonds naturally contain a small amount of protein), a fat content of 2.9 g / 100 mL (from almonds themselves and added vegetable oil), a starch content of 0, and a total carbohydrate content of 2.7 g / 100 mL (sugar-free type, total carbohydrates include dietary fiber and possibly added thickeners). After passing physical and chemical and microbiological tests, it can be used to make cheese. The production process of almond cheese was the same as that in Example 1.

[0070] Comparative Example 4

[0071] In this comparative example, the microalgae milk in Example 1 was replaced with soy milk, and the other steps and conditions were the same as those in Example 1. Soy milk had a protein content of 3.2 g / 100 mL (natural soy protein), a fat content of 2.4 g / 100 mL (from soybeans themselves and added vegetable oil), a starch content of 0.8 g / 100 mL (from added thickeners), and a total carbohydrate content of 2.8 g / 100 mL (sugar-free type, soy oligosaccharides from soybeans themselves). After passing physical and chemical and microbiological tests, it can be used to make cheese. The production process of soy milk cheese was the same as that in Example 1.

[0072] Comparative Example 5

[0073] In this comparative example, the process of making microalgae cheese in Example 1 was changed. Using the pure microalgae milk made in Example 1 as the raw milk, microalgae cheese was made with the basic raw materials in a mass ratio of 20% microalgae milk + 15% potato starch + 15% rapeseed oil + 50% water. Potato starch was used as the starch source; rapeseed oil was used as the fat source, and all raw materials passed physical and chemical and microbiological tests.

[0074] In this comparative example, the production process of microalgae cheese is as follows: raw milk, starch source, fat source and water are fully mixed into a mixed homogenate, and the mixed homogenate is placed in a water bath at 90 °C, kept warm and stirred for 10 min, cooled and solidified to obtain plant-based cheese in the state of cheese.

[0075] Comparative Example 6

[0076] In this comparative example, the process of making microalgae cheese in Example 1 was changed. The pure microalgae milk produced in Example 1 was used as the raw milk, and microalgae cheese was made with 75% microalgae milk + 15% potato starch + 10% rapeseed oil as the basic raw materials according to the mass ratio. Potato starch was used as the starch source; rapeseed oil was used as the fat source, and all raw materials passed physical and chemical and microbiological inspections.

[0077] In this comparative example, the production process of microalgae cheese is as follows: the raw milk, starch source and fat source are fully mixed into a mixed homogenate, and the mixed homogenate is placed in a water bath at 90 °C, kept warm and stirred for 10 min, cooled and solidified to obtain plant-based cheese in the state of cheese.

[0078] The formed states of the cheeses produced in Example 1 and Comparative Examples 1-6 were compared. Among them, in Example 1, microalgae milk was used as the raw material, and cheese was made with an acidulant and rennet; in Comparative Examples 5-6, microalgae milk was used as the raw material, and cheese was made with added potato starch and rapeseed oil as the starch source and fat source respectively. In Comparative Example 1, milk was used as the raw material, and cheese was made with an acidulant and rennet; in Comparative Examples 2-4, oat milk, almond milk, soy milk, etc. were used as the raw milk respectively, and cheese was made with an acidulant and rennet in the same method as in Example 1 and Comparative Example 1.

[0079] For the forming situations of the cheeses produced in each example and comparative example, see Figures 1 - 5 as shown. As Figure 1 shown is a comparison picture of the curdling situations of cow milk cheese (left) in Comparative Example 1 and pure microalgae cheese (right) in Example 1. As Figure 2 shown, are comparison pictures of the curdling situations of cheeses made with oat milk (left), almond milk (middle), soy milk (right), etc. as the raw milk, and an acidulant and rennet in Comparative Examples 2-4. Figure 3 shown is a comparison picture after whey removal of cow milk cheese (left) in Comparative Example 1 and pure microalgae cheese (right) in Example 1. Figure 4 shown is a comparison picture of the cheese products made from cow milk cheese (left) in Comparative Example 1 and pure microalgae cheese (right) in Example 1. Figure 5 shown are comparison pictures of the finished products of microalgae cheese made with 20% microalgae milk + 15% potato starch + 15% rapeseed oil + 50% water as the basic raw materials in Comparative Example 5 and microalgae cheese made with 75% microalgae milk + 15% potato starch + 10% rapeseed oil as the basic raw materials in Comparative Example 6.

[0080] Through comparison, it can be seen that adding acidifying agents and rennet to milk and microalgae milk can both form normal curds, such as Figure 1 , while oat milk, almond milk, and soy milk in Comparative Examples 2-4 cannot achieve normal curdling during traditional cheese making, such as Figure 2 . During traditional cheese making, after the raw milk forms curds, during the process of cutting and draining whey, after centrifuging the cut curds, an obvious separation layer between whey and cheese can be seen. As Figure 3 shows, using milk as the raw milk to make cheese in Comparative Example 1 and using microalgae milk as the raw milk to make cheese in Example 1, obvious separation of whey and cheese can be seen after whey removal and centrifugation. As Figure 4 shows, the cow milk cheese in Comparative Example 1 and the microalgae cheese in Example 1 are similar in terms of appearance integrity, structural strength, tear resistance, density, and stability. As Figure 5 shows, Comparative Examples 5 and 6 use the current conventional plant-based cheese making process to make microalgae cheese. Compared with traditional cow milk cheese and the microalgae cheese prepared in Example 1, the microalgae cheese made in Comparative Examples 5-6 is inferior in terms of elasticity, structural strength, density, and tear resistance, and its storage stability is also poor.

[0081] The above experimental results show that when using microalgae milk to prepare cheese, adopting the same preparation process as cow milk cheese can produce cheese products with similar characteristics to cow milk cheese, while oat milk, almond milk, and soy milk cannot curdle normally and be made into cheese using the same preparation process as cow milk cheese; at the same time, if microalgae milk is used to make cheese according to the current plant-based cheese process, by adding external vegetable oils and starches, etc., it cannot curdle normally and produce a cheese product with a texture and characteristics similar to cow milk cheese. This may be related to the intermediate state characteristics of microalgae protein. As a microorganism, the protein of microalgae may have the characteristics of both animal and plant proteins, such as containing essential amino acids similar to animal proteins, while also having the structure of plant proteins. This may enable the formation of a curd structure similar to cow milk cheese under the action of rennet, thus being similar in texture.

[0082] It has been found that the proteins of microalgae (such as Spirulina and Chlorella) contain all 9 essential amino acids, and the proportions of lysine and leucine are close to those of bovine casein (for example, casein contains 20% essential amino acids). The proportion of hydrophobic amino acids (such as alanine and valine) reaches 30-40%, which is comparable to that of bovine casein (about 35%), endowing similar emulsifying and gelling properties. Microalgae proteins are mainly in the range of 10-150 kDa, partially overlapping with bovine casein (20-25 kDa), and are easily recognized and cleaved by chymosin. Microalgae proteins have a β-sheet and α-helix composite conformation similar to animal proteins, but contain hydrophilic domains of plant proteins, forming a "semi-flexible" network, which is conducive to curd formation. Chymosin (such as bovine chymosin or of microbial origin) targets the Phe105-Met106 site of κ-casein, and the κ-casein-like sequence in microalgae proteins (such as the Chlorella-1 protein of Chlorella contains a similar hydrophobic fragment) can be homologously cleaved to release the hydrophilic terminal peptide chain, triggering protein cross-linking. This is also an accidental discovery of the inventor's team in the process of developing food types and high-protein foods using microalgae as new food resources.

[0083] Further, the odor of the microalgae cheese in Example 1 and the bovine milk cheese in Comparative Example 1 were evaluated and compared. The scoring rules are shown in Table 1, and the results are as Figure 6 . Compared with the bovine milk cheese in Comparative Example 1, the odor of the microalgae cheese in Example 1 conforms to the characteristics of cheese, retaining the milk fragrance and umami of their respective milk sources.

[0084] Table 1: Odor sensory scoring rules

[0085] Odor Scoring Criteria Milk Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Sour Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Umami Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Mushroom Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Fruit Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Sulfur Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Putrid Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Unclean Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Maturity Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Saltiness Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Bitterness Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Odor Intensity Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10

[0086] The taste of the microalgae cheese in Example 1 and the bovine milk cheese in Comparative Example 1 were evaluated and compared. The scoring rules are shown in Table 2, and the results are as Figure 7 . Compared with the bovine milk cheese in Comparative Example 1, the taste of the microalgae cheese in Example 1 conforms to the characteristics of cheese, retaining the milk fragrance of their respective milk sources.

[0087] Table 2: Taste sensory scoring rules

[0088] Taste Scoring Criteria Milk Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Sour Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Umami Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Mushroom Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Fruit Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Sulfur Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Putrid Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Unclean Flavor Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Maturity Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Saltiness Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Bitterness Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Odor Intensity Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10 Taste Intensity Weak: 0 - 3; Medium: 4 - 7; Strong: 8 - 10

[0089] The appearance, color, melting property, oil yield and other indicators of the microalgae cheese in Example 1 and the bovine milk cheese in Comparative Example 1 were evaluated and compared. The scoring rules are shown in Table 3, and the results are as Figure 8 . Compared with the bovine milk cheese in Comparative Example 1, the microalgae cheese in Example 1 meets the conventional index requirements of current cheese foods.

[0090] Table 3: Scoring rules for other conventional indicators of cheese

[0091] Others Scoring Criteria Appearance Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10 Color and Luster Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10 Melting Property Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10 Oiliness Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10 Stability Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10 Elasticity Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10 Degree of Water Separation Average: 0 - 3; Good: 4 - 7; Excellent: 8 - 10

[0092] The examples further use Chlorella pyrenoidosa with the preservation number CGMCC NO.40787 or other high-protein microalgae raw materials with 40-60% protein, 8-16% oil, and 20-30% starch in the cell dry weight to make pure microalgae cheese. The method of Example 1 is used to prepare pure microalgae milk from the microalgae raw materials. During the preparation of microalgae milk, the cell wall breaking fineness and enzymatic hydrolysis degree of the algal solution are controlled to make the starch hydrolysis degree 25-40% and the protein hydrolysis degree 20-30%. When the protein content in the prepared pure microalgae milk is 1.7-4.0 (g / 100ml), the fat content is 0.2-0.8 (g / 100ml), and the starch content is 0.6-1.1 (g / 100ml), cheese products with texture, sensory and other characteristic indexes similar to those of the microalgae cheese in Example 1 can be finally obtained, and it is not limited to specific microalgae species. This shows that the microalgae milk preparation process of the present invention has good stability; moreover, the pure microalgae cheese prepared according to the method of the present invention is closer to traditional cow milk cheese products in terms of curd elasticity, appearance integrity, structural strength, tear resistance, density and stability compared with traditional plant-based cheeses such as soybeans and almonds. Only the selected microalgae varieties are different, and the final cheese products may show different colors, odors and tastes related to the microalgae milk sources.

[0093] Finally, it should be noted that the above examples are only used to illustrate the technical solutions of the present invention, not to limit them; although the present invention has been described in detail with reference to the foregoing examples, those of ordinary skill in the art should understand that they can still modify the technical solutions recorded in the foregoing examples, or perform equivalent replacements on some or all of the technical features; and these modifications or replacements, or when the technical features in the above examples do not conflict with each other, can be combined in the manner recorded in the examples, and these modifications, replacements or combinations do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A pure microalgae cheese, characterized in that: It contains microalgae milk, acidulant and rennet; the microalgae milk is pure microalgae milk made from microalgae raw materials in which protein accounts for 40-60% of cell dry weight, oil accounts for 8-16% of cell dry weight and starch accounts for 20-30% of cell dry weight.

2. The pure microalgae cheese according to claim 1, characterized in that: The acidulant is at least one of hydrochloric acid, phosphoric acid, lactic acid, acetic acid, citric acid, rice vinegar, lemon juice, citric malic acid, and sorbic acid; the microalgae raw material is any one or more of Chlamydomonas reinhardtii, Chlorella, Scenedesmus, Euglena, Spirulina, Dunaliella salina, Microchloropsis pseudochlorophylla, and any one of the high-yield protein mutant algae strains or chlorophyll-deficient algae strains of the foregoing.

3. The pure microalgae cheese according to claim 1, characterized in that: The raw milk used for microalgae cheese has a protein content of 1.7-4.0g / 100ml, a fat content of 0.2-0.8g / 100ml, and a starch content of 0.6-1.1g / 100ml.

4. A method for making pure microalgae cheese, characterized in that: include: S1, selecting microalgae raw materials with protein accounting for 40-60% of cell dry weight, oil accounting for 8-16% of cell dry weight, and starch accounting for 20-30% of cell dry weight to prepare microalgae milk; S2, adding an acidulant to the microalgae milk, heat-keeping and acidification treatment at 80-90°C for 10-20 minutes, and controlling the pH of the acidification end point to be 4.3-4.8; then cooling to below 40°C, adding rennet, stirring well, standing, and curdling; S3. After keeping the temperature at 85-95° C. for 25-40 minutes again, the mixture is cooled and allowed to stand to obtain curds. The curds are cut and stirred slowly to remove whey, thereby obtaining pure microalgae cheese.

5. The method for making pure microalgae cheese according to claim 4, characterized in that: In S1, the microalgae raw material is any one or more of Chlamydomonas reinhardtii, Chlorella vulgaris, Scenedesmus, Euglena, Spirulina, Dunaliella salina, Pseudo-Nanochloa, and any one of the high-yield protein mutant strains or chlorophyll-deficient strains of the foregoing.

6. The method for making pure microalgae cheese according to claim 4, characterized in that: In S1, the preparation method of microalgae milk is as follows: Step 1: Select a microalgae raw material with protein accounting for 40-60% of the cell dry weight, oil accounting for 8-16% of the cell dry weight, and starch accounting for 20-30% of the cell dry weight; Step 2: using drinking water to prepare algae slurry, so that the algae cell dry matter concentration is 100g / L-200g / L, using a high-pressure homogenizer to break the algae cells to obtain broken algae slurry; after homogenization, particles below 1μm in the cell slurry account for more than 90% of the total number of particles, and particles below 300nm account for more than 50% of the total number of particles; Step 3: adding water to dilute the broken algae slurry until the dry matter content of the microalgae is 20g / L-100g / L, adding 350-5000 units / kg of cellulase, 500-3000 units / kg of amylase and 1000-3000 units / kg of saccharifying enzyme, and 350-1000 units / kg of protease in sequence to perform enzymolysis based on the weight of the diluted algae slurry, controlling the enzymolysis conditions so that the degree of starch hydrolysis is 25-40% and the degree of protein hydrolysis is 20-30%, and obtaining an enzymolysis mixture; Step 4: Heat the enzymatic hydrolysis mixture obtained in step 3 to 50-80° C., transfer it into a high-pressure homogenizer for homogenization at least once, and sterilize it to obtain microalgae milk. The protein content of the microalgae milk is 1.7-4.0 g / 100 ml, the fat content is 0.2-0.8 g / 100 ml, and the starch content is 0.6-1.1 g / 100 ml.

7. The method for making pure microalgae cheese according to claim 4, characterized in that: In S2, the acidulant is at least one of hydrochloric acid, phosphoric acid, lactic acid, acetic acid, citric acid, rice vinegar, lemon juice, citric malic acid, and sorbic acid.

8. The method for making pure microalgae cheese according to claim 4, characterized in that: In S2, the amount of rennet added is 1-15 g rennet / 1000 L microalgae milk; the rennet is added to the microalgae milk that has reached the acidification end point and cooled to below 40° C.; the rennet is one or more of animal rennet, plant rennet and microbial rennet.

9. The method for making pure microalgae cheese according to claim 4, characterized in that: In S2, citric acid is added to the microalgae milk, and the milk is acidified in a water bath at 90°C for 10 minutes, and the pH value of the acidification end point is controlled to be 4.6; the milk is then cooled to below 40°C, rennet is added, and the milk is stirred thoroughly and allowed to stand for 2 hours to coagulate; then in S3, the milk is kept in a water bath at 90°C for 30 minutes, and then allowed to cool and stand to obtain curds, which are cut and centrifuged to remove the whey to obtain pure microalgae cheese.

10. The method for making pure microalgae cheese according to claim 4, characterized in that: In S3, the operation method of cutting the curd and slowly stirring is: cutting after the curd is completed, slowly stirring, heating to 39° C., keeping the temperature for 30-45 minutes, and removing the whey after the curd is healed; The method for removing whey is: filtering with edible gauze, cloth bag or cheese bag, or centrifugal separation after stirring, with a centrifugal force of 4000-5000g and a centrifugal time of 10-20min.

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