A method for preparing microalgae plant yogurt using acid-producing and aroma-producing bacteria

Abstract

The present invention relates to the technical field of food processing, and in particular to a method for preparing microalgae plant yogurt using acid-producing and aroma-producing bacteria, which comprises: S1, selecting microalgae raw materials: using microalgae with a protein content of ≥35% in algae cells as raw materials; S2, cell disruption: using a high-pressure homogenizer to disrupt algae cells to obtain broken wall algae pulp; S3, blending and sterilization: adding sugar, thickener and pure water, adjusting the protein concentration of the broken wall algae pulp to obtain blended algae pulp, sterilization treatment; S4, fermentation: inoculating mixed strains, fermenting in two stages, including: first inoculating Clostridium butyricum, Clostridium cohnii and yeast for a predetermined fermentation time, and then inoculating lactic acid bacteria for fermentation; S5, subpackaging and refrigeration: subpackaging the fermented algae pulp into a packaging container, placing it at 0-8°C for refrigeration and post-ripening, and obtaining microalgae plant yogurt. The present invention metabolizes most of the algae smell substances through clostridium and yeast fermentation, produces more abundant fragrance substances than fermentation with lactic acid bacteria alone, and obtains fragrant and delicious microalgae yogurt. The microalgae plant yogurt of the present invention can meet the needs of people who are allergic to milk and pursue plant-based foods.

Description

Technical Field

[0001] The invention belongs to the technical field of food processing, and in particular relates to a method for preparing microalgae yogurt by utilizing acid- and aroma-producing bacteria. Background Art

[0002] Traditional yogurt is mainly made from animal milk such as cow's milk and has a history of thousands of years of consumption. Yogurt is rich in nutrition and has a unique flavor, and is very popular among people. Although lactic acid bacteria fermentation consumes part of the lactose and casein is also degraded to a certain extent, there are still some people who are intolerant or allergic to yogurt. Plant protein products can solve the needs of this group of people for protein supplementation. In addition, the production process of plant-based foods pollutes and affects the environment and consumes much less resources than traditional animal husbandry production. Some vegetarians and environmentalists will refuse or reduce the consumption of animal products, while hoping not to affect nutritional intake and good food enjoyment. These people need corresponding animal protein substitutes, and various plant protein products are very popular with them.

[0003] As for yogurt substitutes, there are some plant yogurt products on the market, mainly soy milk, almond yogurt and coconut yogurt. These plant yogurt products have various disadvantages. First, their flavors are not as strong as yogurt. Second, they have some tastes that consumers don't like (including smell and taste). For example, soy milk has a distinct smell of stinky bean juice, almond yogurt has a dizzy and unpleasant odor, and coconut yogurt has a corrupted taste. The reasons for the shortcomings of plant yogurt flavor are related to the raw materials and the fermentation process. The strains of traditional fermented yogurt are separated from traditional fermented yogurt. These lactic acid bacteria can produce good flavors for the chemical components of milk and will not produce bad flavors. However, if these lactic acid bacteria are fermented on plant raw materials, they can not only fail to metabolize the special odor in the plant raw materials, but also fail to produce rich flavor components like fermented milk raw materials. At the same time, for higher plants such as peanuts, nuts (such as almonds), and coconuts, the ratio of protein, oil, and starch content is relatively fixed. For example, peanuts contain 24-35g / 100g of protein and 40-50g / 100g of fat or oil; almond kernels contain 28g / 100g of protein and 55-61g / 100g of fat (oil); coconut meat contains 3.3-4g / 100g, and the oil content is quite high, about 33.5-35g / 100g. It is difficult to find a balance between high protein and low oil when making dairy products with these plant raw materials, and it is difficult for higher plants themselves to adjust the composition ratio of protein, oil, and starch in cells through culture conditions, which may not meet the needs of people who pursue a high-protein, low-fat diet.

[0004] As a single-cell plant, microalgae has the advantages of a wide variety, short growth cycle, rapid cultivation, and the ability to easily adjust the composition ratio of protein, oil and starch in cells through culture conditions. Therefore, the production of protein drinks using microalgae as raw materials can easily take into account high protein and low fat or make targeted adjustments according to the needs of specific groups of people. Compared with animal raw materials or higher plant raw materials, the production of microalgae biomass also occupies less natural resources. Not only that, microalgae also contain nutrients such as Ω-3 polyunsaturated fatty acids, carotenoids (such as astaxanthin, lutein, fucoxanthin, etc.), and multiple vitamins. If whole algae biomass is used to make food, microalgae biomass can be fully utilized and consumers can be provided with algae polysaccharides, dietary fiber and other ingredients that are beneficial to human health. As more and more microalgae varieties are included in the food list, downstream foods or nutritional supplements using microalgae as raw materials are gradually becoming known to people.

[0005] However, at present, most of the development and application of microalgae food raw materials are limited to extracting algal oil DHA, carotenoids, protein, etc. from microalgae cells and using them as additives in beverages. There is rarely any development based on whole algae applications, and no yogurt products based on whole algae raw materials have been seen. Summary of the invention

[0006] 1. Technical issues to be resolved

[0007] In view of the above-mentioned shortcomings and deficiencies of the prior art, the present invention provides a method for preparing microalgae plant yogurt by using acid-producing and aroma-producing bacteria. The method mainly uses high-protein-yielding microalgae as raw materials to make yogurt from whole algae. On the one hand, the microalgae have richer nutritional components than soybeans, almond kernels and coconuts, which can make the yogurt have higher nutritional value. On the other hand, most of the algae odor substances can be metabolized through the fermentation of clostridium and yeast, producing more abundant aroma substances than fermentation with lactic acid bacteria alone, making the prepared microalgae yogurt more fragrant and delicious, thereby overcoming the technical problems of the poor taste of existing plant yogurt.

[0008] (II) Technical solution

[0009] In a first aspect, the present invention provides a method for preparing microalgae plant yogurt using acid- and aroma-producing bacteria, comprising:

[0010] S1. Select microalgae raw materials: use microalgae with a protein content of ≥35% in algae cells as raw materials;

[0011] S2. Cell disruption: using a high-pressure homogenizer to disrupt algae cells to obtain broken algae slurry;

[0012] S3, preparation and sterilization: adding sugar, thickener and purified water, adjusting the protein concentration of the broken algae slurry to obtain the prepared algae slurry, and sterilizing;

[0013] S4, fermentation: inoculating mixed bacteria, and fermenting in two stages, including: first inoculating Clostridium butyricum, Clostridium cohnii and yeast for a predetermined fermentation time, and then inoculating lactic acid bacteria for fermentation;

[0014] S5. Packaging and refrigeration: Pack the fermented algae pulp into packaging containers, place them in a refrigerator at 0-8°C and ripen them to obtain microalgae plant yogurt.

[0015] According to a preferred embodiment of the present invention, in S1, the raw material is Chlorella pyrenoidosa.

[0016] According to a preferred embodiment of the present invention, in S2, the microalgae raw material is processed by a high-pressure homogenizer in the form of an algae cell suspension, and the concentration of the algae cell suspension during the treatment is 50-200 g / L (algae dry matter), the homogenization pressure is 100-130 MPa, and the homogenization is circulated 1-3 times until the particles below 1 μm in the broken algae slurry 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.

[0017] Among them, if the raw material is dry algae powder, water is added to adjust the concentration of the algae cell suspension to 50-200g / L (algae dry matter); if the raw material is algae liquid, it is concentrated or water is added to adjust the concentration of the algae cell suspension to 50-200g / L (algae dry matter).

[0018] According to a preferred embodiment of the present invention, in S3, 40-80 g / L of glucose, fructose syrup or starch syrup (based on the volume of the prepared algae slurry, syrup is calculated as glucose), 5-20 g / L of carrageenan, gellan gum, acetyl distarch phosphate or hydroxypropyl distarch phosphate (based on the volume of the prepared algae slurry) and pure water are added to the broken algae slurry and mixed so that the protein content in the prepared algae slurry is ≥30 g / L.

[0019] According to a preferred embodiment of the present invention, in S3, the sterilization treatment adopts any one of ultra-high temperature instantaneous sterilization, high temperature sterilization and pasteurization. The ultra-high temperature instantaneous sterilization condition is 135-150℃ for 2-10s; the high temperature sterilization condition is 90-115℃ for 1-10min; the pasteurization condition is 70-90℃ for 10-20min.

[0020] According to a preferred embodiment of the present invention, in S4, a mixed strain of Clostridium butyricum, Clostridium cohnii and Saccharomyces cerevisiae is first inoculated and fermented under anaerobic conditions at 30-34°C for 24-32 hours, and then Lactobacillus bulgaricus and Streptococcus thermophilus are inoculated and fermented anaerobically at 40-45°C for 4-8 hours.

[0021] According to a preferred embodiment of the present invention, in S4, Clostridium butyricum is selected from the strain ATCC19398, Clostridium cohnii is selected from the strain ATCC8527, the yeast is selected from Saccharomyces cerevisiae CICC33068, and the lactic acid bacteria is selected from a mixture of Lactobacillus delbrueckii subspecies bulgaricus ATCC11842 and Streptococcus thermophilus ATCC19258.

[0022] First, prepare seed solution with Clostridium butyricum and Clostridium cohnii, and inoculate in the form of seed solution. When the concentration of seed solution is (1~10)×10 7 cfu / mL, the inoculation volume was 0.5% (v / v); yeast can be inoculated in the form of seed liquid or dry powder. When the seed liquid concentration is (1-10)×10 7 cfu / mL, the inoculation volume is 1% (v / v); Lactobacillus bulgaricus and Streptococcus thermophilus can be inoculated in the form of seed liquid or dry powder. When the seed liquid concentration is (1-5)×10 8 During the inoculation process, the above inoculation amount can be referred to and converted according to the seed solution concentration or the number of live bacteria per unit gram weight of dry powder to obtain an amount equivalent to the above inoculation amount.

[0023] According to a preferred embodiment of the present invention, in S5, the yogurt is packaged into small packages of 80-120 mL or large packages of 1000 mL in a workshop with a cleanliness level of 4 or above (GB50073-2013), and after packaging, the product is stored at 0-8°C for 24 hours to complete post-ripening.

[0024] After the yogurt is fermented, it is placed in a low temperature environment for a period of time. This helps to slow down or stop the activity of lactic acid bacteria and prevent the yogurt from being over-acidified, thereby controlling the acidity of the final product within an appropriate range. At the same time, the protein in the yogurt is further coagulated, making the texture more delicate and smooth, while increasing the viscosity and improving the overall eating experience. In addition, in the post-ripening stage, the carboxylic acids and alcohols in the yogurt further react into esters and develop in balance with other flavor substances, making the flavor of the yogurt more aromatic, soft, and mellow, reducing the pungent sour taste.

[0025] In a second aspect, the present invention provides a microalgae plant yogurt prepared by using acid- and aroma-producing bacteria, which is prepared by the method of any of the above embodiments.

[0026] (III) Beneficial effects

[0027] The main technical effects of the present invention are:

[0028] (1) The present invention uses high-protein microalgae as the raw material for producing plant yogurt, and utilizes the characteristics of microalgae such as short growth cycle, rapid cultivation, industrial production, and easy adjustment of the nutritional composition structure of microalgae to produce a high-protein plant milk beverage product, thereby making up for the problems that the raw materials of existing plant milk beverages such as peanut milk, nut milk, oat milk, and rice milk require seasonal production and the protein / fat ratio is difficult to adjust.

[0029] (2) The present invention uses high-protein-yielding microalgae as processing raw materials and uses whole algae to make yogurt drinks. The technical effect is that it not only has a high protein content, but also contains rich carotenoid components (including but not limited to: lutein for protecting eyesight, antioxidant astaxanthin, fucoxanthin for promoting lipid metabolism and protecting the liver, etc.), polyunsaturated fatty acids, vitamins, polysaccharides, dietary fiber and trace elements that are not available in most other commercially available plant protein products, making the product more nutritionally comprehensive.

[0030] (3) The present invention inoculates mixed strains of Clostridium butyricum, Clostridium cohnii and yeast into algae pulp for fermentation. On the one hand, the undesirable flavor components such as algae smell in the algae pulp are metabolized, and butyric acid, caproic acid, ethanol and other ester-forming flavoring base substances are produced, and aromatic substances such as ethyl butyrate and ethyl caproate are further produced. After that, lactic acid bacteria are inoculated for fermentation, and organic acids such as lactic acid are produced, which can form esters with alcohol substances produced by yeast fermentation, such as ethyl lactate. On the other hand, lactic acid bacteria fermentation reduces the pH value of the fermentation liquid, promotes protein coagulation, and forms a coagulable yogurt tissue state, making the yogurt texture more delicate and smooth, increasing viscosity, and improving the overall eating experience.

[0031] (4) The present invention produces ethanol through the yeast fermentation process, which has the conditions to produce flavoring ingredients such as ethyl butyrate, ethyl caproate and ethyl lactate. These esters can give yogurt a particularly strong aroma; in addition, yeast can also produce some ester substances to make the product fragrant. This is an advantage compared to traditional yogurt and plant yogurt available on the market. DETAILED DESCRIPTION

[0032] In order to better explain the present invention and facilitate understanding, the present invention is described in detail below through specific implementation methods.

[0033] The present invention discloses a method for preparing microalgae plant yogurt by using acid- and aroma-producing bacteria, comprising the following steps:

[0034] (1) Selecting microalgae raw materials: The microalgae raw materials are high-protein-producing microalgae, and the protein content in the algae cells is ≥ 35%. Preferably, the microalgae is Chlorella pyrenoidosa.

[0035] (2) Cell disruption: Add water to adjust the microalgae cell concentration to 50-200 g / L on a dry weight basis, and use a high-pressure homogenizer to disrupt the cells to obtain algae slurry.

[0036] The degree of cell disruption is preferably such that particles below 1 μm in the broken algae slurry account for more than 90% of the total number of particles, and particles below 300 nm account for more than 50% of the total number of particles.

[0037] If the initial raw material is dry algae powder, water is added to adjust the concentration of the algae cell suspension to 50-200 g / L (algae dry matter); if the initial raw material is algae liquid, it is concentrated or water is added to adjust the concentration of the algae cell suspension to 50-200 g / L (algae dry matter). Preferably, the algae cell concentration is adjusted to 100 g / L and homogenized at a pressure of 100-130 MPa.

[0038] (3) Preparation: Add 40-80 g / L of glucose, fructose syrup or starch syrup (based on the prepared volume, calculated as glucose), 5-20 g / L of carrageenan, gellan gum, acetyl distarch phosphate or hydroxypropyl distarch phosphate (based on the prepared volume) and purified water to the broken algae slurry and mix well, so that the protein content in the prepared algae slurry is ≥30 g / L.

[0039] Preferably, 80 g / L of fructose syrup and 10 g / L of hydroxypropyl distarch phosphate as thickener are added, purified water is added and mixed with the broken algae pulp, and the concentration of the algae pulp is controlled to make its protein content ≥ 30 g / L. After being prepared according to the above method and conditions, the final drink has the most moderate sweetness and taste, high protein content, good smoothness and gel properties.

[0040] (4) Sterilization: The sterilization method is ultra-high temperature instantaneous sterilization, high temperature sterilization or pasteurization. The conditions for ultra-high temperature instantaneous sterilization are 135-150℃ for 2-10 seconds; the conditions for high temperature sterilization are 90-115℃ for 1-10 minutes; the conditions for pasteurization are 70-90℃ for 10-20 minutes.

[0041] (5) Fermentation: The fermentation is carried out in two steps, first with Clostridium butyricum, Clostridium cohnii and yeast, and then with lactic acid bacteria. Clostridium butyricum is selected from the strain ATCC19398, Clostridium cohnii is selected from the strain ATCC8527, yeast is selected from Saccharomyces cerevisiae CICC33068, and lactic acid bacteria are selected from Lactobacillus delbrueckii subspecies bulgaricus ATCC11842 and Streptococcus thermophilus ATCC19258. Specifically, a mixed strain of Clostridium butyricum, Clostridium cohnii and Saccharomyces cerevisiae is first inoculated and fermented under anaerobic conditions at 30-34°C for 24-32 hours, and then Lactobacillus bulgaricus and Streptococcus thermophilus are inoculated and fermented anaerobically at 40-45°C for 4-8 hours.

[0042] In the first fermentation step, Clostridium butyricum and Clostridium cohnii are inoculated at 0.5% (v / v) respectively, and yeast is inoculated at 1% (v / v); in the second fermentation step, the inoculation amount of lactic acid bacteria is 1% (v / v). The inoculation amount can be referred to the following method or equivalent conversion:

[0043] First, prepare seed solution with Clostridium butyricum and Clostridium cohnii, and inoculate in the form of seed solution. When the concentration of seed solution is (1~10)×10 7 cfu / mL, the inoculation volume was 0.5% (v / v); yeast can be inoculated in the form of seed liquid or dry powder. When the seed liquid concentration is (1-10)×10 7 cfu / mL, the inoculation volume is 1% (v / v); Lactobacillus bulgaricus and Streptococcus thermophilus can be inoculated in the form of seed liquid or dry powder. When the seed liquid concentration is (1-5)×10 8 cfu / mL, and the inoculation amount was 1% (v / v).

[0044] (6) Packaging and refrigeration: After fermentation is completed, the fermentation liquid is packaged in a workshop with a cleanliness level of 4 or above (GB50073-2013) in small packages of 80-120 mL or large packages of 1000 mL. After packaging, the product is stored at 0-8℃ for 24 hours. During this period, the yogurt is ripened.

[0045] The present invention is described below with reference to specific embodiments.

[0046] Example 1

[0047] This embodiment uses Chlorella pyrenoidosa as the raw material of microalgae plant yogurt, and the steps are as follows:

[0048] (1) Through heterotrophic fermentation culture, 100 L of fermentation liquid with an algae dry matter concentration of 100 g / L and a cell protein content of 60% (w / w, dry matter) was obtained. The algae cells were separated by a centrifuge to obtain algae cells with a moisture content of about 75%, i.e. 40 kg of algae mud (containing 10 kg of algae dry matter).

[0049] (2) Place algae mud in a 250L storage tank, add drinking purified water to prepare an algae cell suspension, the total volume of the suspension is 100L, the algae cell dry matter concentration is 100g / L, and homogenize it three times with a high-pressure homogenizer at 120MPa to obtain a broken algae slurry. In the broken algae slurry, particles below 1μm account for 95%, and particles below 300nm account for 58%.

[0050] (3) Place the broken algae pulp in a 200L mixing tank, add 32kg of fructose syrup (sugar content 50%) and 2kg of hydroxypropyl distarch phosphate (first dissolved in 50L of water), and finally add water to a total volume of 200L. Stir and mix while heating, and control the temperature at 50±2°C until all substances are evenly dispersed or dissolved. Finally, adjust the pH value to 7.0±0.2 with 2mol / L sodium hydroxide solution to obtain the prepared algae pulp. The protein content of the prepared algae pulp is 30g / L.

[0051] (4) The prepared algae slurry is transported into a shell-and-tube heat exchanger or a plate heat exchanger for sterilization. The flow rate and temperature are adjusted to control the material temperature to quickly reach 115°C and maintain it for 1 minute. It is then quickly cooled to 32±2°C and transported to a sterilized fermentation tank.

[0052] (5) In the fermentation tank, the temperature was controlled at 32°C, and the pre-cultured strains were inoculated at a concentration of 2×10 7 cfu / mL of Clostridium butyricum and strain concentrations were 2.5×10 7 cfu / mL of Clostridium cohnii seed culture solution, inoculated with a pre-cultured strain at a concentration of 3×10 7 cfu / mL yeast seed culture solution was fermented anaerobicly without ventilation or stirring for 24 h; the fermentation solution temperature was adjusted to 43°C, and the pre-cultured strains were inoculated at a concentration of 2.0×10 8 cfu / mL of Lactobacillus bulgaricus and strain concentrations at 2.0×10 8 cfu / mL of thermophilic Streptococcus seed liquid, ferment for 8h, and end the fermentation process.

[0053] (6) In a workshop with a cleanliness level of Class 4 or above that complies with the GB50073-2013 standard, the mixture is packaged into 1000 mL beverage bottles and stored at 4° C. for 24 hours to obtain a microalgae plant yogurt product.

[0054] Example 2

[0055] The difference between this example and Example 1 is that the cell protein content of the raw material Chlorella vulgaris used for preparation is 35% (w / w, dry matter basis), and the other steps and conditions are the same as those of Example 1. The protein content of the prepared slurry obtained in step (3) is 17.5 g / L.

[0056] Example 3

[0057] The difference between this embodiment and embodiment 1 is that the conditions for breaking cells by a high pressure homogenizer in step (2) are different. In this embodiment, the cells are homogenized twice at 100 MPa to obtain a broken algae slurry. The particles below 1 μm account for 90% of the broken algae slurry, and the particles below 300 nm account for 51%. The other steps and conditions are the same as those in embodiment 1.

[0058] Example 4

[0059] The difference between this embodiment and embodiment 1 is that the sterilization of the prepared algae slurry in step (4) is carried out under the condition of 80°C and 15 minutes of pasteurization. The other steps and conditions are the same as those in embodiment 1.

[0060] Example 5

[0061] The difference between this embodiment and embodiment 1 is that the fermentation conditions in step (5) are adjusted to allow Clostridium butyricum, Clostridium cohnii and yeast to ferment for 32 hours, and the fermentation liquid temperature is adjusted to 43°C; then Lactobacillus bulgaricus and Streptococcus thermophilus are inoculated and fermented for 4 hours. The other steps and conditions are the same as those in embodiment 1.

[0062] Example 6

[0063] The difference between this embodiment and embodiment 1 is that the inoculation amount of the fermentation bacteria in step (5) is different: the inoculation amount of Clostridium butyricum is 1.8×10 7 cfu / mL of seed solution was inoculated at 0.56% (v / v), and Clostridium cohnii was inoculated at 2.5×10 7 cfu / mL seed solution was inoculated at 0.5% (v / v), and the pre-cultured strain concentration was inoculated at 1.2% (v / v) at a concentration of 2.5×10 7 cfu / mL yeast seed culture was fermented anaerobicly for 24 h without ventilation or stirring; the fermentation liquid temperature was adjusted to 41°C, and the pre-cultured strain was inoculated at a concentration of 2.5×10 8 cfu / mL of Lactobacillus bulgaricus and 1% (v / v) inoculation volume were inoculated with the pre-cultured strain at a concentration of 2.5×10 8 cfu / mL of thermophilic Streptococcus seed solution, fermented for 7.5 hours, and then the fermentation process was terminated. Other steps and conditions were the same as those in Example 1.

[0064] Comparative Example 1

[0065] The difference between this comparative example and Example 1 is that the pressure of the high-pressure homogenizer for disrupting cells in step (2) is 60 MPa and the homogenization frequency is 1 time, and the rest is the same as Example 1. The other steps and conditions are the same as Example 1.

[0066] Comparative Example 2

[0067] The difference between this comparative example and Example 1 is that the first stage of fermentation is omitted in the fermentation stage of step (5), and only lactic acid bacteria fermentation is retained, and the rest is the same as Example 1. The other steps and conditions are the same as Example 1.

[0068] Comparative Example 3

[0069] The difference between this comparative example and Example 1 is that in step (5), Clostridium butyricum and Clostridium cohnii were not inoculated in the first stage of fermentation, but only yeast was inoculated. The second stage was the same as Example 1, and the other steps and conditions were the same as Example 1.

[0070] Quality characterization of microalgae plant yogurt products:

[0071] (1) The main flavor substances of the microalgae plant yogurt prepared in the above examples were detected. The test results are shown in Table 1.

[0072] Table 1: Comparison of the content of main flavor substances in yogurt (normalized %)

[0073]

[0074]

[0075] Note: “-” in the table means no detection; Examples 1-6 are represented by the abbreviations of Examples 1-6, and Comparative Examples 1-3 are represented by the abbreviations of Comparative Examples 1-3.

[0076] The gel properties were tested using a TA.XT21 texture analyzer with a probe model of P0.5, a probe descent speed of 2 mm / s, a total descent depth of 10 mm, and the force when the gel was broken was the gel strength (N).

[0077] The acidity was determined using the method of GB 5009.239-2016.

[0078] The volatile flavor substances were analyzed by solid phase microextraction-gas chromatography-mass spectrometry, and the results were expressed as content (%) by peak area normalization method.

[0079] (2) The flavor characteristics of the microalgae plant yogurt prepared in each of the above examples were tasted and sensory evaluation was performed. The evaluation method is as follows. The evaluation results are shown in Table 2.

[0080] Sensory evaluation was performed by 15 sensory evaluators on the same batch of samples of the embodiments. The sensory evaluation indexes were smooth taste, algae smell and aroma.

[0081] Scoring rules: Smooth taste 0-9 points, representing a very rough taste to a smooth and refreshing taste. The higher the score, the smoother it is. 0-3 points are very rough, and 7-9 points are very smooth.

[0082] The algae smell score is 0-9, representing a strong algae smell to no algae smell. 0-3 means a very strong algae smell, 4-6 means a distinct algae smell, and 7-9 means a light or no algae smell.

[0083] The aroma score is 0-9 points. The aroma of yogurt is mainly close to the aroma of liquor. The score ranges from light aroma without obvious aroma to strong aroma. 0-3 points means light aroma without obvious aroma, 4-6 points means obvious aroma, and 7-9 points means strong wine aroma.

[0084] Group Real 1 Real 2 Reality 3 Reality 4 Reality 5 Actual 6 1 2 3 Smooth taste 9 9 7.3 9 9 9 4.5 7.6 9 Algae smell 9 8 8 9 9 9 7.5 4.3 5.3 aromatic 9 8.7 8 9 8.7 9 6.5 5.3 6.4

[0085] Note: Examples 1-6 are represented by the abbreviations Examples 1-6, and Comparative Examples 1-3 are represented by the abbreviations Comparative Examples 1-3.

[0086] In Table 1, hexanol, 1-octen-3-ol, β-ionone, and hexanal are representative components of bad flavor substances in microalgae food. Hexanol is the main source of grassy taste, 1-octen-3-ol is the source of earthy smell, hexanal is the source of fishy smell, and β-ionone is the source of woody taste. By detecting the content of these bad flavor substances, the elimination of bad flavor of algae can be monitored. The lower their content, the less strange taste of microalgae yogurt drink, and the greater the acceptability.

[0087] 3-Hydroxy-2-butanone, 2,3-pentanedione, 2,3-butanedione and acetaldehyde are representative of the aroma components of yogurt and are typical products of fermentation by thermophilic Streptococcus and bulgaricus Lactobacillus; the higher their content, the stronger the milk-like aroma of the yogurt drink.

[0088] Ethyl acetate, ethyl butyrate and ethyl caproate have strong fruity and wine aromas, are the products of esterification of acetic acid, butyric acid, caproic acid and ethanol, and are the result of the reaction of acid-producing bacteria and Saccharomyces cerevisiae metabolites. Lactic acid is a metabolite of lactic acid bacteria. In traditional yogurt, lactic acid and its esters (such as isobutyl lactate) exist to a certain extent, which is the characteristic flavor component of traditional yogurt; but since there is little ethanol in the products produced only by lactic acid bacteria fermentation, fatty acid ethyl esters are also rarely detected. The fermentation process of the present invention is based on the use of butyric acid bacteria (clostridium butyricum) and caproic acid bacteria (clostridium cohnii), and then yeast fermentation is carried out to produce acid and ethanol in combination, which creates material conditions for the formation of strong-flavor esters similar to the aroma of white wine, such as ethyl butyrate, ethyl caproate, ethyl acetate, ethyl lactate, etc.

[0089] Compared with Example 1, the pressure of the high-pressure homogenizer used in Comparative Example 1 is not enough, the cell disruption effect is poor, so the cell content is not fully released, the protein coagulation effect of the product is poor, the effect of decomposing the substances with algae smell is relatively poor, and there is still a certain algae smell. Because the particles are large, the yogurt graininess is more obvious, and the smooth taste is also poor. Comparative Example 2 lacks the first stage fermentation process. On the one hand, the decomposition of the cell bad flavor substances is very poor, and the algae smell is retained more, showing that the microalgae yogurt has a significant algae smell; on the other hand, due to the lack of Clostridium and yeast fermentation, the production of butyric acid, caproic acid, and acetic acid in the product is greatly reduced, accompanied by low or undetectable corresponding ester content, so the product lacks fruity and wine aromas. Although Comparative Example 3 is also fermented in two stages, no butyrate- and caproate-producing Clostridium is added in the first stage of fermentation, and only yeast fermentation is used, resulting in very low or even undetectable contents of ethyl acetate, ethyl butyrate, and ethyl caproate in the yogurt. Although the content of ethyl lactate has not decreased, the aroma of ethyl lactate is not as prominent as that of ethyl butyrate and ethyl caproate. Therefore, the product has lower fruity and wine aromas and a stronger alcohol taste.

[0090] Comparing Example 1 with Example 3, it can be found that when the protein content in the microalgae cells reaches 60%, the homogenization effect can be improved by increasing the homogenization pressure or increasing the number of homogenization times; and the better the homogenization effect, the better the yogurt gel state is when the aroma-producing bacteria participate in the fermentation, the smoother the taste, the moderate acidity, the higher the degree of elimination of the algae smell, and the final product has not only the normal aroma of yogurt, but also a strong wine aroma. It can be seen that for the same microalgae raw material, if the homogenization effect does not reach the optimal state, it will affect the release of cell contents, the removal of algae smell and the quality of the finished product.

[0091] Comparing Example 1 with Example 2, it can be seen that microalgae with a protein content of 35% can also produce microalgae yogurt of good quality, but because the protein content of the microalgae raw material is much lower, the nutrition of the yogurt is worse and the gel properties of the yogurt are also worse.

[0092] Comparing Example 1 with Example 4, the high temperature sterilization method and pasteurization method for the raw materials have no significant effect on the quality of the yogurt. Both methods can kill most of the microorganisms in the raw materials and in the production process. Since the inoculation amount of Clostridium, yeast and lactic acid bacteria in the fermentation is sufficient to form a dominant fermentation of working bacteria, the fermentation produces acid quickly, the yogurt is at a high acidity of about pH 4.0, and the miscellaneous bacteria are inhibited. Since the yogurt is refrigerated at low temperature and sold in the cold chain, the common food sterilization conditions are basically applicable.

[0093] Comparing Example 1 with Example 5, Example 5 prolongs the first stage fermentation time and shortens the lactic acid fermentation time. Finally, the content of butyric acid and caproic acid in the yogurt product increases, butyrate and caproate increase, and the typical metabolites of lactic acid bacteria, 3-hydroxy-2-butanone, 2,3-pentanedione, and 2,3-butanedione, also decrease, and the wine aroma is more prominent.

[0094] Comparing Example 1 with Example 6, it can be known that when the bacterial concentration in the fermentation bacteria seed culture solution decreases, the volume ratio of inoculation can be appropriately increased or the fermentation time can be extended. When the bacterial concentration in the fermentation bacteria seed culture solution increases, the volume ratio of inoculation can be appropriately reduced or the fermentation time can be shortened, and the various indicators of the final product are relatively close. In addition, the fermentation temperature also affects the reproduction speed of the fermentation bacteria and the speed of fermentation. Generally, the higher the fermentation temperature is within the suitable temperature range of the fermentation bacteria, the faster the fermentation speed is. Therefore, in actual production, the fermentation conditions, including the initial bacterial inoculum amount, the fermentation time, the fermentation temperature, etc., can be reasonably adjusted within the scope recorded in this application according to parameters such as ambient temperature, the concentration of each component in the prepared slurry, and the expected quality of the product.

[0095] In addition, the test results in Table 1 are basically consistent with the sensory evaluation in Table 2.

[0096] In summary, the present invention provides a method for preparing microalgae plant yogurt using acid- and aroma-producing bacteria. The prepared microalgae yogurt has a high protein content and can be used as a substitute for yogurt, providing an ideal protein drink for people who are allergic to milk, vegetarians, and those who pursue environmental protection, enriching food shelves and providing consumers with more diverse choices of protein supplement drinks.

[0097] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing microalgae plant yogurt using acid-producing and aroma-producing bacteria, characterized in that: include: S1. Select microalgae raw materials: use microalgae with a protein content of ≥35% in algae cells as raw materials; S2. Cell disruption: using a high-pressure homogenizer to disrupt algae cells to obtain broken algae slurry; S3, preparation and sterilization: adding sugar, thickener and purified water, adjusting the protein concentration of the broken algae slurry to obtain the prepared algae slurry, and sterilizing; S4, fermentation: inoculating mixed bacteria, fermenting in two stages, including: first inoculating a mixed bacteria of Clostridium butyricum, Clostridium cohnii and Saccharomyces cerevisiae for 24-32 hours under anaerobic conditions at 30-34°C, and then inoculating Lactobacillus bulgaricus and Streptococcus thermophilus for 4-8 hours under anaerobic conditions at 40-45°C; S5. Packaging and refrigeration: Pack the fermented algae pulp into packaging containers, place them in a refrigerator at 0-8°C and ripen them to obtain microalgae plant yogurt.

2. The method according to claim 1, characterized in that In S1, the raw material is Chlorella pyrenoidosa.

3. The method according to claim 1, characterized in that In S2, the microalgae raw material is processed by a high-pressure homogenizer in the form of an algae cell suspension, and the concentration of the algae cell suspension during the treatment is 50-200 g / L, the homogenization pressure is 100-130 MPa, and the homogenization is circulated 1-3 times until the particles below 1 μm in the broken algae slurry 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.

4. The method according to claim 1, characterized in that In S3, the sugar is glucose, fructose syrup or starch syrup, and the amount of glucose, fructose syrup or starch syrup converted into glucose is 40-80 g / L based on the volume of the prepared algae slurry; The thickener is carrageenan, gellan gum, acetylated distarch phosphate or hydroxypropyl distarch phosphate, and the amount of the thickener added is 5-20 g / L based on the volume of the prepared algae slurry; The amount of pure water added is controlled to the amount required to make the protein content in the prepared algae slurry ≥30g / L.

5. The method according to claim 1, characterized in that In S3, the sterilization treatment adopts any one of ultra-high temperature instantaneous sterilization, high temperature sterilization and pasteurization; the conditions for ultra-high temperature instantaneous sterilization are 135-150℃ for 2-10s; the conditions for high temperature sterilization are 90-115℃ for 1-10min; the conditions for pasteurization are 70-90℃ for 10-20min.

6. The method according to claim 1, characterized in that In S4, Clostridium butyricum is selected from the strain ATCC19398, Clostridium cohnii is selected from the strain ATCC8527, the yeast is selected from Saccharomyces cerevisiae CICC33068, and the lactic acid bacteria is selected from a mixture of Lactobacillus delbrueckii subsp. bulgaricus ATCC11842 and Streptococcus thermophilus ATCC19258.

7. The method according to claim 6, characterized in that In S4, the inoculum amount is: Clostridium butyricum and Clostridium cohnii were inoculated in the form of seed solution. When the concentration of seed solution was (1-10)×10 7 cfu / mL, the inoculation volume was 0.5% (v / v); the yeast was inoculated in the form of seed liquid or dry powder, and when the seed liquid concentration was (1-10)×10 7 cfu / mL, the inoculation volume was 1% (v / v); Lactobacillus bulgaricus and Streptococcus thermophilus were inoculated in the form of seed liquid or dry powder. When the seed liquid concentration was (1-5)×10 8 cfu / mL, the inoculation volume was 1% (v / v). During the inoculation process, the above-mentioned inoculation volume was used as a reference and the inoculation was performed based on the seed solution concentration or the number of live bacteria per gram of dry powder.

8. A microalgae plant yogurt prepared by using acid- and aroma-producing bacteria, which is prepared by the method described in any one of claims 1 to 7.

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