Method for preparing microalgae plant-based yogurt by means of using acid- and aroma-producing microbes

Abstract

A method for preparing microalgae plant-based yogurt by means of using acid- and aroma-producing microbes. The method comprises: S1, selection of microalgae raw materials: using microalgae with a protein content of ≥35% in algae cells as raw materials; S2, cell disruption: disrupting the algae cells by means of using a high-pressure homogenizer to obtain a wall-broken algal slurry; S3, blending and sterilization: adding sugar, a thickening agent and purified water to the wall-broken algal slurry, adjusting the protein concentration of the wall-broken algal slurry to obtain a blended algal slurry, and performing sterilization treatment; S4, fermentation: inoculating a mixed strain for two-stage fermentation, comprising: first inoculating Clostridium butyricum, Clostridium kluyveri and yeast for a predetermined period of time, and then inoculating lactic acid bacteria for fermentation; and S5, sub-packaging and refrigeration: sub-packaging the fermented algal slurry into a packaging container, placing same at 0°C-8°C for refrigeration and post-ripening, and thus obtaining the microalgae plant-based yogurt. By means of fermentation with clostridia and yeast, most of algal off-flavor substances are metabolized, and a richer variety of aromatic substances are produced compared with fermentation with lactic acid bacteria alone, resulting in an aromatic and palatable microalgae-based yogurt that meets the needs of populations with milk allergies and those pursuing plant-based foods.

Description

Method for preparing microalgae yogurt by using acid and aroma producing bacteria TECHNICAL FIELD

[0001] The present application belongs to the technical field of food processing, and particularly relates to a method for preparing microalgae yogurt by using acid and aroma producing bacteria. BACKGROUND

[0002] Traditional yogurt is mainly made of animal milk such as cow milk, and has a history of thousands of years. Yogurt is rich in nutrients and has a unique flavor, and is very popular among people. Although lactic acid bacteria fermentation consumes part of lactose and casein is degraded to some extent, some people still have intolerance or allergic reactions to sour cow milk, and plant protein products can meet the protein supplement needs of these people. In addition, the production process of plant food causes much less pollution and impact on the environment and resource consumption than traditional animal husbandry. Some vegetarians and environmentalists will refuse or reduce the consumption of animal products, while still wanting to maintain nutritional intake and good food enjoyment. These people need corresponding animal protein substitutes, and various plant protein products are very popular among them.

[0003] As for yogurt substitutes, some plant yogurt products have appeared on the market, mainly including sour soy milk, almond yogurt and coconut yogurt. These plant yogurt products all have various shortcomings. First, their flavors are not as rich as sour cow milk, and second, they have some flavors (including odor and taste) that consumers do not like. For example, sour soy milk has a distinct stinky soybean odor, almond yogurt has a nauseating unpleasant odor, and coconut yogurt has a rancid taste. The reasons for the flavor shortcomings of plant yogurt are related to the raw materials and the fermentation process. The strains for traditional fermented yogurt are isolated from traditional fermented sour cow milk, and these lactic acid bacteria fermentation can produce good flavor for the chemical composition of cow milk and will not produce undesirable flavor. However, if these lactic acid bacteria are used to ferment plant raw materials, they cannot metabolize the special odor of plant raw materials, nor can they produce rich aroma components like fermenting cow milk raw materials. Meanwhile, for plants such as peanuts, nuts (such as almonds, etc.), and coconuts, the ratio of protein, oil, and starch content is relatively fixed. For example, peanuts contain 24-35 g / 100 g of protein and 40-50 g / 100 g of fat or oil; almond kernels contain 28 g / 100 g of protein and 55-61 g / 100 g of fat (oil); coconut meat contains 3.3-4 g / 100 g of protein and a relatively high oil content of about 33.5-35 g / 100 g. It is difficult to balance high protein and low oil when making dairy products from such plant raw materials, and it is also difficult to adjust the composition ratio of protein, oil, and starch in cells through cultivation conditions for higher plants, which may not meet the needs of people who pursue high-protein and low-fat diets.

[0004] As a kind of unicellular plant, microalgae has the advantages of various species, short growth cycle, rapid cultivation, easy adjustment of the composition ratio of proteins, oils and starches in cells by culture conditions, etc., so that the production of protein drinks using microalgae as raw materials can easily balance high protein and low fat or be adjusted according to the needs of specific groups, and compared with animal raw materials or higher plant raw materials, the production of microalgae biomass occupies less natural resources. Moreover, microalgae also contains omega-3 polyunsaturated fatty acids, carotenoids (such as astaxanthin, lutein, fucoxanthin, etc.), various vitamins and other nutrients. If the whole algal biomass is used to manufacture food, the algal polysaccharides, dietary fiber and other ingredients beneficial to human health can also be fully utilized and provided to consumers. With more and more microalgae varieties being listed in the food directory, downstream foods or nutritional supplements using microalgae as raw materials are gradually known by people.

[0005] However, at present, most of the development and application of microalgae food raw materials are limited to extracting algal oil DHA, carotenoids, proteins and other substances from microalgae cells and using them as additives in drinks, and there are few developments based on whole algae applications, and no yogurt products based on whole algae raw materials have been seen. SUMMARY

[0006] (One) technical problems to be solved

[0007] In view of the above shortcomings and deficiencies of the prior art, the present application provides a method for preparing microalgae plant yogurt by using acid-producing and aroma-producing bacteria, which mainly uses microalgae with high protein yield as raw material to prepare yogurt from whole algae. On the one hand, microalgae has more nutritional ingredients than soybeans, almonds and coconuts, so that the yogurt has higher nutritional value. On the other hand, through the fermentation of clostridium and yeast, most of the algae fishy substances can be metabolized, and more aroma substances can be produced than only using lactic acid bacteria fermentation, so that the prepared microalgae yogurt is more fragrant and delicious, thereby overcoming the technical problems of poor taste of existing plant yogurt.

[0008] (Two) technical solutions

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

[0010] S1, selecting microalgae raw material: using microalgae with protein content ≥ 35% in algal cells as raw material;

[0011] S2, cell disruption: breaking algal cells by using a high-pressure homogenizer to obtain broken cell algal slurry;

[0012] S3, blending and sterilization: adding sugar, thickening agent and pure water, adjusting the protein concentration of the broken cell algal slurry to obtain blended algal slurry, and sterilizing;

[0013] S4, fermentation: inoculate mixed bacteria, two-stage fermentation, including: first inoculate Clostridium butyricum, Clostridium coccoides and yeast fermentation for a predetermined time, then inoculate lactic acid bacteria fermentation;

[0014] S5, cold storage: the fermented algae slurry is divided into packaging containers, stored at 0-8℃ and post-ripened, and the microalgae plant yogurt is obtained.

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

[0016] According to the preferred embodiment of the present application, in S2, the microalgae raw material is treated by a high-pressure homogenizer in the form of an algal cell suspension, and the concentration of the algal cell suspension is 50-200g / L (algal dry matter) during treatment, the homogenization pressure is 100-130MPa, and the homogenization is repeated 1-3 times until the particles below 1μm account for more than 90% of the total number of particles and the particles below 300nm account for more than 50% of the total number of particles in the broken algal slurry.

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

[0018] According to the preferred embodiment of the present application, in S3, glucose, fructose syrup or starch syrup 40-80g / L (calculated by the volume of the prepared algal slurry, the syrup is calculated by glucose), carrageenan, gellan gum, acetyl di-starch phosphate or hydroxypropyl di-starch phosphate 5-20g / L (calculated by the volume of the prepared algal slurry), and pure water are added and mixed with the broken algal slurry to make the protein content in the prepared algal slurry ≥30g / L.

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

[0020] According to the preferred embodiment of the present application, in S4, first inoculate Clostridium butyricum, Clostridium coccoides and Saccharomyces cerevisiae mixed bacteria to ferment at 30-34℃ under anaerobic conditions for 24-32h, and then inoculate Lactobacillus bulgaricus and Streptococcus thermophilus to ferment at 40-45℃ under anaerobic conditions for 4-8h.

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

[0022] First, seed liquid is prepared by Clostridium butyricum and Clostridium coslifer, and inoculation is performed in the form of seed liquid, when the concentration of seed liquid is (1-10)×10 7 cfu / mL, the inoculation amount is 0.5% (v / v) respectively; yeast can be inoculated in the form of seed liquid or dry powder, when the concentration of seed liquid is (1-10)×10 7 cfu / mL, the inoculation amount is 1% (v / v); Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus can be inoculated in the form of seed liquid or dry powder, when the concentration of seed liquid is (1-5)×10 8 cfu / mL, the inoculation amount is 1% (v / v) respectively. During inoculation, the above-mentioned inoculation amount can be referred to, and the equivalent amount of the above-mentioned inoculation amount can be obtained by conversion according to the number of viable bacteria per unit gram weight of dry powder or the concentration of seed liquid.

[0023] According to the preferred embodiment of the present application, in S5, the yogurt is divided into small packages of 80-120 mL or large packages of 1000 mL in a workshop with cleanliness of level 4 or above (GB50073-2013), and the product is preserved at 0-8℃ for 24 h after post-ripening.

[0024] Post-ripening is that the yogurt is placed at low temperature for a period of time after fermentation is completed, which helps to slow down or stop the activity of lactic acid bacteria, prevent the yogurt from over-acidification, so as to control the acidity of the final product within a suitable range, and further coagulate the protein in the yogurt, so that the texture becomes more delicate and smooth, and the viscosity is increased, and the overall eating experience is improved. In addition, in the post-ripening stage, carboxylic acid and alcohol in the yogurt further react into ester and develop other flavor substances, so that the flavor of the yogurt is more aromatic, soft and mellow, and the pungent sour taste is reduced.

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

[0026] (Three) beneficial effects

[0027] The main technical effect of the present application is:

[0028] (1) The present application uses microalgae with high protein content as the raw material for the production of plant yogurt, taking advantage of the short growth cycle, rapid cultivation, industrial production, and easy adjustment of the nutritional composition of microalgae, to produce high-protein plant milk beverage products, which makes up for the problems of seasonal production and difficulty in adjusting the protein / oil ratio of existing plant milk beverage raw materials such as peanut milk, nut milk, oat milk, and rice milk.

[0029] (2) The present application uses microalgae with high protein content as the raw material for the production of plant yogurt, taking advantage of the short growth cycle, rapid cultivation, industrial production, and easy adjustment of the nutritional composition of microalgae, to produce high-protein plant milk beverage products, which makes up for the problems of seasonal production and difficulty in adjusting the protein / oil ratio of existing plant milk beverage raw materials such as peanut milk, nut milk, oat milk, and rice milk.

[0030] (3) The present application inoculates the algae slurry with mixed bacteria such as Clostridium butyricum, Clostridium coccoides, and yeast, and ferments it. On the one hand, it metabolizes the undesirable flavors such as algae smell in the algae slurry, and produces ester-based flavoring substances such as butyric acid, caproic acid, and ethanol. Further, it produces aromatic substances such as butyl butyrate and ethyl caproate. Then, it inoculates the fermented liquid with lactic acid bacteria, produces organic acids such as lactic acid, and esterifies with the alcohol produced by yeast fermentation, such as ethyl lactate. On the other hand, lactic acid bacteria fermentation reduces the pH of the fermentation liquid, promotes protein coagulation, forms a coagulated yogurt texture, makes the yogurt texture more delicate and smooth, increases the viscosity, and improves the overall eating experience.

[0031] (4) The present application produces ethanol through yeast fermentation, provides the conditions for the production of ester-based flavoring substances such as butyl butyrate, ethyl caproate, and ethyl lactate, which can impart a particularly rich aroma to the yogurt. In addition, yeast can also produce some ester substances, making the product have a fragrance. This is an advantage compared to traditional sour milk and existing plant yogurt on the market. DETAILED DESCRIPTION

[0032] In order to better explain the present application, the following specific embodiments are used to describe the present application in detail.

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

[0034] (1) Selecting microalgae raw material: the microalgae raw material is high-protein microalgae, and the protein content in the algal cells is ≥ 35%. Preferably, the microalgae is Chlorella pyrenoidosa.

[0035] (2) Cell disruption: water is added to adjust the concentration of microalgae cells to 50-200 g / L (dry weight) and a high-pressure homogenizer is used for disruption to obtain algae slurry.

[0036] The degree of cell disruption is preferably achieved as follows: the percentage of particles below 1 μm in the broken cell algae slurry is more than 90% of the total number of particles, and the percentage of particles below 300 nm is 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 (dry matter of algae); 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 (dry matter of algae). Preferably, the concentration of the algae cells is adjusted to 100 g / L, and homogenization is performed at a pressure of 100-130 MPa.

[0038] (3) Formulation: 40-80 g / L (based on the volume of the completed formulation, calculated based on glucose) of glucose, high fructose syrup or starch syrup, 5-20 g / L (based on the volume of the completed formulation) of carrageenan, gellan gum, acetyl di-starch phosphate or hydroxypropyl di-starch phosphate, and purified water are added and mixed with the broken cell algae slurry to make the protein content of the completed formulation algae slurry ≥ 30 g / L.

[0039] Preferably, 80 g / L of high fructose syrup is added as the sugar, 10 g / L of hydroxypropyl di-starch phosphate is added as the thickening agent, and purified water is added and mixed with the broken cell algae slurry to control the concentration of the algae slurry so that the protein content is ≥ 30 g / L. After formulation according to the foregoing method and conditions, the final beverage has a moderate sweetness and mouthfeel, a high protein content, and good smoothness and gelation.

[0040] (4) Sterilization: one of the following methods is used for sterilization: ultra-high temperature instant sterilization, high temperature sterilization or pasteurization. The conditions for ultra-high temperature instant sterilization are 135-150 °C for 2-10 seconds; the conditions for high temperature sterilization are 90-115 °C for 1-10 minutes; and the conditions for pasteurization are 70-90 °C for 10-20 minutes.

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

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

[0043] First, seed liquid is prepared from Clostridium butyricum and Clostridium coccoides, and the seed liquid is inoculated when the concentration of the seed liquid is (1-10) x 10 7 cfu / mL, and the inoculation amount is 0.5% (v / v); yeast can be inoculated in the form of seed liquid or dry powder, and the inoculation amount is 1% (v / v) when the concentration of the seed liquid is (1-10) x 10 7 cfu / mL; Lactobacillus bulgaricus and Streptococcus thermophilus can be inoculated in the form of seed liquid or dry powder, and the inoculation amount is 1% (v / v) when the concentration of the seed liquid is (1-5) x 10 8 cfu / mL.

[0044] (6) Cold storage: after fermentation, the fermentation liquid is stored in a clean room with a cleanliness of level 4 (GB50073-2013) for 24 hours at 0-8°C, during which the yogurt is post-ripened.

[0045] The present application is described below in combination with specific examples.

[0046] Example 1

[0047] In this example, Chlorella pyrenoidosa is used as the raw material for microalgae yogurt, and the steps are as follows:

[0048] (1) 100 L of fermentation liquid with an algal dry matter concentration of 100 g / L and a cell protein content of 60% (w / w, based on dry matter) is obtained by heterotrophic fermentation culture, and the algal cells are separated by a centrifuge to obtain algal cells with a water content of about 75%, i.e., 40 kg of algal slurry (containing 10 Kg of algal dry matter).

[0049] (2) The algal slurry is placed in a 250 L storage tank, and drinking purified water is added to prepare an algal cell suspension with a total volume of 100 L and an algal cell dry matter concentration of 100 g / L. The suspension is homogenized by a high-pressure homogenizer for 3 times at 120 MPa to obtain broken algal slurry. In the broken algal slurry, particles below 1 μm account for 95%, and particles below 300 nm account for 58%.

[0050] (3) Put the broken cell algae paste into a 200L preparation tank, add fructo-oligosaccharide 32kg (sugar content 50%), hydroxypropyl distarch phosphate 2kg (first dissolved in 50L water), and finally add water to a total volume of 200L. Mix while heating and stirring, control the temperature at 50±2℃, until all materials 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 paste. The protein content of the prepared algae paste is 30g / L.

[0051] (4) Transport the prepared algae paste into a tubular heat exchanger or plate heat exchanger for sterilization. Adjust the flow rate and temperature, control the material temperature to quickly reach 115℃ and maintain for 1min, and quickly cool to 32±2℃ and transport to the sterilized fermentation tank.

[0052] (5) In the fermentation tank, control the temperature at 32℃, inoculate the seed culture of Clostridium butyricum with a strain concentration of 2×10 7 cfu / mL at an inoculation amount of 0.5% (v / v) and the seed culture of Clostridium coccoides with a strain concentration of 2.5×10 7 cfu / mL at an inoculation amount of 1% (v / v), and the seed culture of yeast with a strain concentration of 3×10 7 cfu / mL at an inoculation amount of 1% (v / v) in an anaerobic manner without aeration and stirring for 24h; adjust the fermentation liquid temperature to 43℃, inoculate the seed culture of Lactobacillus bulgaricus with a strain concentration of 2.0×10 8 cfu / mL at an inoculation amount of 1% (v / v) and the seed culture of Streptococcus thermophilus with a strain concentration of 2.0×10 8 cfu / mL at an inoculation amount of 1% (v / v), and ferment for 8h to end the fermentation process.

[0053] (6) In a workshop with a cleanliness of 4 or above according to the GB50073-2013 standard, dispense into 1000mL beverage bottles, and store at 4℃ for 24h to obtain the 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 pyrenoidosa 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 17.5g / L.

[0056] Example 3

[0057] The difference between this example and Example 1 is that the condition of breaking cells by high pressure homogenizer in step (2) is different. In this example, the cell slurry is homogenized twice at 100 MPa. The cell slurry contains 90% of particles below 1 μm and 51% of particles below 300 nm. Other steps and conditions are the same as in Example 1.

[0058] Example 4

[0059] The difference between this example and Example 1 is that the sterilization condition of the algae slurry in step (4) is different. In this example, the algae slurry is sterilized by the Busch method at 80°C for 15 min. Other steps and conditions are the same as in Example 1.

[0060] Example 5

[0061] The difference between this example and Example 1 is that the fermentation conditions in step (5) are adjusted. In this example, Clostridium butyricum and Clostridium coccoides, and yeast are fermented for 32 h, and the temperature of the fermentation broth is adjusted to 43°C. Then Lactobacillus bulgaricus and Streptococcus thermophilus are inoculated and fermented for 4 h. Other steps and conditions are the same as in Example 1.

[0062] Example 6

[0063] The difference between this example and Example 1 is that the inoculation amount of the fermentation strains in step (5) is different. In this example, Clostridium butyricum is inoculated at 0.56% (v / v) from a seed liquid with a strain concentration of 1.8 x 1010cfu / mL, Clostridium coccoides is inoculated at 0.5% (v / v) from a seed liquid with a strain concentration of 2.5 x 1010cfu / mL, yeast is inoculated at 1.2% (v / v) from a seed liquid with a strain concentration of 2.5 x 1010cfu / mL, Lactobacillus bulgaricus is inoculated at 1% (v / v) from a seed liquid with a strain concentration of 2.5 x 1010cfu / mL, and Streptococcus thermophilus is inoculated at 1% (v / v) from a seed liquid with a strain concentration of 2.5 x 1010cfu / mL. Other steps and conditions are the same as in Example 1. 7 7 7 8 8

[0064] Comparative Example 1

[0065] The difference between this comparative example and Example 1 is that the pressure of the high pressure homogenizer in step (2) is 60 MPa and the number of homogenization is 1. Other steps and conditions are the same as in Example 1.

[0066] Comparative Example 2

[0067] ​​​​​The difference between the present comparative example and Example 1 is that the fermentation stage in step (5) omits the first stage fermentation and only retains the lactic acid bacteria fermentation, and the other steps and conditions are the same as Example 1.

[0068] Comparative Example 3

[0069] The difference between the present comparative example and Example 1 is that the first stage fermentation bacteria in step (5) is not inoculated with Clostridium butyricum and Clostridium cosl, but only inoculated with yeast. The second stage is the same as Example 1, and the other steps and conditions are the same as Example 1.

[0070] Quality characterization of microalgae plant yogurt products:

[0071] (1) The main flavor substances of the microalgae plant yogurts prepared in each of the above examples were detected, and the detection results are shown in Table 1.

[0072] Table 1: Comparison of main flavor substance contents of yogurts (normalized method %)

[0073] Note: "-" in the table means not detected; Examples 1-6 are represented by the abbreviations 1-6, and Comparative Examples 1-3 are represented by the abbreviations 1-3.

[0074] Gel properties were detected by a TA.XT21 texture analyzer, the probe type was P0.5, the probe lowering speed was 2 mm / s, the total lowering depth was 10 mm, and the force at which the gel broke was the gel strength (N).

[0075] The determination of acidity was determined by the method of GB 5009.239-2016.

[0076] The analysis of volatile flavor substances was carried out by solid phase microextraction-gas chromatography-mass spectrometry, and the results were expressed as content (%) by peak area normalization method.

[0077] (2) The flavor properties of the microalgae plant yogurts prepared in each of the above examples were tasted and the sensory evaluation was counted, and the evaluation method is as follows, and the evaluation results are shown in Table 2.

[0078] The sensory evaluation was descriptive testing by 15 sensory evaluators on the same batch of example samples. The indicators of sensory evaluation are smooth mouthfeel, algae smell and aroma.

[0079] Scoring rules: smooth mouthfeel 0-9 points, representing from very rough mouthfeel to smooth and smooth mouthfeel, the higher the score, the smoother, 0-3 points for very rough, 7-9 points for very smooth;

[0080] Algal taste score 0-9, representing from heavy algal taste to no algal taste, 0-3 is very heavy algal taste, 4-6 is obvious algal taste, 7-9 is light algal taste or no algal taste;

[0081] Fragrance score 0-9, the fragrance of yogurt is mainly close to liquor fragrance, the score represents from light smell to strong fragrance, 0-3 is light smell without obvious fragrance, 4-6 is obvious fragrance, 7-9 is strong liquor fragrance.

[0082] Note: Examples 1-6 are represented by abbreviations 1-6, and Comparative Examples 1-3 are represented by abbreviations 1-3.

[0083] In Table 1, hexanol, 1-octen-3-ol, beta-ionone and hexanal are representative components of undesirable flavor substances of microalgae food, hexanol is the main source of grassy flavor, 1-octen-3-ol is the source of earthy flavor, hexanal is the source of fishy flavor, and beta-ionone is the source of woody flavor. By detecting the content of these several undesirable flavor substances, the elimination of undesirable flavor of algae can be monitored, and the lower the content, the less the strange taste of microalgae yogurt drink, and the greater the acceptability.

[0084] 3-hydroxy-2-butanone, 2,3-pentanedione, 2,3-butanedione and acetaldehyde are representative components of yogurt aroma, and are typical products of fermentation by Streptococcus thermophilus and Lactobacillus bulgaricus; the higher the content, the stronger the yogurt drink similar to milk aroma.

[0085] Ethyl acetate, ethyl butyrate and ethyl hexanoate have strong fruit and liquor aroma, and are the products of esterification of acetic acid, butyric acid and hexanoic acid with ethanol, which are the results of metabolic products of acid-producing bacteria and Saccharomyces cerevisiae. Lactic acid is a metabolic product of lactic acid bacteria. In traditional sour milk, lactic acid and its esters (such as isobutyl lactate) exist to some extent, which are characteristic flavor components of traditional sour milk; however, since there is little ethanol produced by lactic acid bacteria fermentation, there is also little detection of fatty acid ethyl esters. The fermentation process of the present application uses butyric acid bacteria (Clostridium butyricum) and hexanoic acid bacteria (Clostridium carboxidivorans) as the basis, and then performs yeast fermentation, which combines acid production and ethanol production, creating material conditions for the formation of strong esters such as butyl acetate, ethyl hexanoate, ethyl acetate and ethyl lactate, which are similar to liquor aroma.

[0086] Compared with Example 1, the pressure used in Comparative Example 1 is not enough for the high-pressure homogenizer treatment, and the cell disruption effect is poor, so the cell content is not fully released, the protein coagulation effect of the product is poor, and the decomposition of the algae-like odor substance is relatively poor, and there is still some algae-like odor. Because the particles are large, the yogurt has a strong grainy taste, and the smooth mouthfeel is also poor. Comparative Example 2 lacks the first-stage fermentation process. On the one hand, the decomposition of cell odor substances is poor, and the algae-like odor is retained, which makes the microalgae yogurt have a strong algae-like odor. On the other hand, because there is no clostridium and yeast fermentation, the production of butyric acid, caproic acid, and acetic acid in the product is greatly reduced, and the corresponding ester content is low or not detected, so the product lacks fruit aroma and wine aroma. Although Comparative Example 3 is also a two-stage fermentation, the first-stage fermentation does not add butyric acid and caproic acid-producing clostridium, and only yeast fermentation is used, resulting in a very low or undetectable content of ethyl lactate, ethyl butyrate, and ethyl caproate in the yogurt. Although the content of ethyl lactate does not decrease, the aroma of ethyl lactate is not as prominent as that of ethyl butyrate and ethyl caproate, so the product has low fruit aroma and wine aroma and a strong alcohol smell.

[0087] Comparing Example 1 and Example 3, it can be found that when the protein content in microalgae cells reaches 60%, the homogenization effect can be improved by increasing the homogenization pressure or the number of homogenization. The better the homogenization effect, the better the yogurt gel state, the smoother the mouthfeel, the moderate acidity, and the higher the degree of elimination of algae-like odor. The final product not only has the normal aroma of yogurt, but also has a strong wine aroma. Therefore, for the same microalgae raw material, if the homogenization effect is not optimal, it will affect the release of cell contents, the removal of algae-like odor, and the quality of the final product.

[0088] Comparing Example 1 and Example 2, it can be found that microalgae yogurt with a protein content of 35% can also be produced with good quality. However, because the protein content of the microalgae raw material is much lower, the yogurt has poorer nutrition and poorer gel properties.

[0089] Comparing Example 1 and Example 4, the use of high-temperature sterilization and pasteurization methods for the raw material does not have a significant impact on the quality of the yogurt. Both methods can kill most of the microorganisms in the raw material and during production. Because 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, and the yogurt is in a high-acid state at pH 4.0, which inhibits the growth of other bacteria. Because the yogurt is stored in a cold environment and sold in a cold chain, common food sterilization conditions can be used.

[0090] Compared with Example 1, Example 5 prolongs the first-stage fermentation time, shortens the lactic acid fermentation time, increases the content of butyric acid and hexanoic acid in the final yogurt product, increases butyrate and hexanoate, reduces 3-hydroxy-2-butanone, 2,3-pentanedione and 2,3-butanedione which are typical metabolic products of lactic acid bacteria, and makes the wine aroma more prominent.

[0091] Compared with Example 1, Example 6 can appropriately increase the inoculation volume ratio or prolong the fermentation time when the concentration of the fermentation bacteria seed culture solution is reduced, and can appropriately reduce the inoculation volume ratio or shorten the fermentation time when the concentration of the fermentation bacteria seed culture solution is increased, and the 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, and generally, the higher the fermentation temperature within the suitable temperature range of the fermentation bacteria, the faster the fermentation speed. Therefore, in actual production, the fermentation conditions, including the initial bacteria inoculation amount, the fermentation time length and the fermentation temperature, can be reasonably adjusted within the range described in the application according to the environmental temperature, the concentration of each component in the slurry, the expected quality of the product and other parameters.

[0092] In addition, the detection results in Table 1 are basically consistent with the sensory evaluation in Table 2.

[0093] In summary, the present application provides a method for preparing microalgae plant yogurt by using acid-producing and aroma-producing bacteria, and the microalgae yogurt prepared by the method has high protein content and can be used as a substitute for sour milk, providing ideal protein drinks for people with milk allergy, vegetarians and people pursuing environmental protection, enriching the food shelf and providing consumers with more diversified choices of protein supplement drinks.

[0094] Finally, it should be noted that: the above examples are only used to illustrate the technical solutions of the present application, and not to limit them; although the present application has been described in detail with reference to the foregoing examples, those skilled in the art should understand that they can still modify the technical solutions described in the foregoing examples, or make equivalent replacement for part or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present application.

Claims

1. A method for preparing a microalgal plant yogurt using an acid- and aroma-producing bacterium, characterized by, The method comprises the following steps: S1, selecting microalgae raw material: using microalgae with protein content in algal cells ≥ 35% as raw material; S2, cell disruption: using a high-pressure homogenizer to break the algal cells to obtain broken cell slurry; S3, blending and sterilization: adding sugar, thickening agent and pure water, adjusting the protein concentration of the broken cell slurry to obtain blended slurry, and sterilizing; S4, fermentation: inoculating mixed bacteria, and carrying out two-stage fermentation, including: inoculating Clostridium butyricum, Clostridium coccoides and yeast for a predetermined time, and then inoculating lactic acid bacteria for fermentation; S5, cold storage and packaging: packaging the fermented algal slurry into packaging containers, and placing the containers in a 0-8℃ cold storage for post-ripening, thereby obtaining microalgae plant yogurt.

2. The method of claim 1, wherein, In S1, the raw material is Chlorella pyrenoidosa.

3. The method of claim 1, wherein, In S2, the microalgae raw material is in the form of algal cell suspension, and the concentration of the algal cell suspension is 50-200g / L during the treatment by the high-pressure homogenizer, the homogenization pressure is 100-130MPa, and the homogenization is repeated 1-3 times until the particles below 1μm account for more than 90% of the total number of particles and the particles below 300nm account for more than 50% of the total number of particles in the broken cell slurry.

4. The method of claim 1, wherein, In S3, the sugar is glucose, fructose glucose syrup or starch syrup, and the addition amount of glucose, fructose glucose syrup or starch syrup converted into glucose is 40-80g / L based on the volume of the blended slurry after blending; The thickening agent is carrageenan, gellan gum, acetylated distarch phosphate or hydroxypropyl distarch phosphate, and the addition amount of the thickening agent is 5-20g / L based on the volume of the blended slurry after blending; The addition amount of pure water is controlled to the amount required to make the protein content in the blended slurry ≥ 30g / L.

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

6. The method of claim 1, wherein, In S4, the mixed bacteria of Clostridium butyricum, Clostridium coccoides and Saccharomyces cerevisiae are inoculated for anaerobic fermentation at 30-34℃ for 24-32h, and then the mixed bacteria of Lactobacillus bulgaricus and Streptococcus thermophilus are inoculated for anaerobic fermentation at 40-45℃ for 4-8h.

7. The method according to claim 1 or 6, characterized in that, In S4, the strain ATCC19398 is selected for Clostridium butyricum, the strain ATCC8527 is selected for Clostridium coccoides, the Saccharomyces cerevisiae CICC33068 is selected for yeast, and the mixed bacteria of Lactobacillus delbrueckii subsp. bulgaricus ATCC11842 and Streptococcus thermophilus ATCC19258 are selected for lactic acid bacteria.

8. The method of claim 7, wherein, In S4, the inoculation amount of the strains is as follows: Clostridium butyricum and Clostridium coccoides are inoculated in the form of seed liquid, when the concentration of the seed liquid is (1-10)×10 7 cfu / mL, the inoculation amount is 0.5% (v / v) respectively; the yeast can be inoculated in the form of seed liquid or dry powder, when the concentration of the seed liquid is (1-10)×10 7 cfu / mL, the inoculation amount is 1% (v / v); Lactobacillus bulgaricus and Streptococcus thermophilus can be inoculated in the form of seed liquid or dry powder, when the concentration of the seed liquid is (1-5)×10 8 cfu / mL, the inoculation amount is 1% (v / v) respectively; during the inoculation, the inoculation amount is converted according to the equivalent amount of the viable bacteria number of the seed liquid concentration or the unit gram weight of the dry powder according to the aforementioned inoculation amount.

9. The method of claim 1, wherein, In S5, the yogurt is packaged into small packages of 80-120mL or large packages of 1000mL in a workshop with cleanliness of level 4 or above (GB50073-2013), and the packaged product is stored at 0-8℃ for 24h to complete post-ripening.

10. A microalgae plant yogurt prepared by using acid-producing and aroma-producing bacteria, which is prepared by the method of any one of claims 1-9.

Images