CN115428834A - Yoghourt with effects of relieving pressure and helping sleep as well as preparation method and application of yoghourt - Google Patents

Yoghourt with effects of relieving pressure and helping sleep as well as preparation method and application of yoghourt Download PDF

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Publication number
CN115428834A
CN115428834A CN202211274911.4A CN202211274911A CN115428834A CN 115428834 A CN115428834 A CN 115428834A CN 202211274911 A CN202211274911 A CN 202211274911A CN 115428834 A CN115428834 A CN 115428834A
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sleep
yoghourt
relieving
effects
juice
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CN115428834B (en
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方曙光
杨竹源
郭晓娟
汪欣
朱建国
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WeCare Probiotics Co Ltd
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WeCare Probiotics Co Ltd
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/13—Fermented milk preparations; Treatment using microorganisms or enzymes using additives
    • A23C9/1307—Milk products or derivatives; Fruit or vegetable juices; Sugars, sugar alcohols, sweeteners; Oligosaccharides; Organic acids or salts thereof or acidifying agents; Flavours, dyes or pigments; Inert or aerosol gases; Carbonation methods
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
    • A23C9/1234—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt characterised by using a Lactobacillus sp. other than Lactobacillus Bulgaricus, including Bificlobacterium sp.
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
    • A23C9/1238—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt using specific L. bulgaricus or S. thermophilus microorganisms; using entrapped or encapsulated yoghurt bacteria; Physical or chemical treatment of L. bulgaricus or S. thermophilus cultures; Fermentation only with L. bulgaricus or only with S. thermophilus
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/13—Fermented milk preparations; Treatment using microorganisms or enzymes using additives
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/13—Fermented milk preparations; Treatment using microorganisms or enzymes using additives
    • A23C9/1322—Inorganic compounds; Minerals, including organic salts thereof, oligo-elements; Amino-acids, peptides, protein-hydrolysates or derivatives; Nucleic acids or derivatives; Yeast extract or autolysate; Vitamins; Antibiotics; Bacteriocins
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C2240/00—Use or particular additives or ingredients
    • A23C2240/15—Use of plant extracts, including purified and isolated derivatives thereof, as ingredient in dairy products
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2400/00—Lactic or propionic acid bacteria
    • A23V2400/11—Lactobacillus
    • A23V2400/113—Acidophilus
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2400/00—Lactic or propionic acid bacteria
    • A23V2400/11—Lactobacillus
    • A23V2400/137—Delbrueckii
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2400/00—Lactic or propionic acid bacteria
    • A23V2400/21—Streptococcus, lactococcus
    • A23V2400/249—Thermophilus
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2400/00—Lactic or propionic acid bacteria
    • A23V2400/51—Bifidobacterium
    • A23V2400/533—Longum

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  • Food Science & Technology (AREA)
  • Polymers & Plastics (AREA)
  • Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Molecular Biology (AREA)
  • Nutrition Science (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Dairy Products (AREA)

Abstract

The invention provides yoghourt with functions of relieving pressure and helping sleep and a preparation method thereof. The yogurt with the effects of relieving pressure and helping sleep is prepared from the following raw materials: milk powder, whey protein powder, sodium glutamate, tea theanine, spina date seed juice, rose concentrated juice, grape juice, a sweetening agent, a stabilizing agent and water. The invention provides the yoghourt with the effects of relieving pressure and helping sleep, which has good taste, good nutritional value and health-care function and can effectively relieve the sleep disorder problem for a plurality of current sleep disorder people. The yoghourt with the effects of relieving pressure and helping sleep provided by the invention is simple in preparation process, free of special condition control and easy to produce.

Description

Yoghourt with effects of relieving pressure and helping sleep as well as preparation method and application of yoghourt
Technical Field
The invention belongs to the technical field of food processing, and relates to yoghourt with functions of relieving pressure and helping sleep, and a preparation method and application thereof
Background
In recent years, the number of cases related to sudden illness and death due to excessive fatigue has been increasing. Sufficient sleep is not only a normal physiological requirement of a human body, but also an important mode for the human body to recover the physical ability of the human body, sleep disorder often causes related symptoms such as difficulty in falling asleep, short sleep time caused by difficulty in sleep duration, and sleep quality reduction, and long-time sleep disorder not only affects physical health and mental health, but also threatens life safety even for serious people.
With the continuous development and progress of the current society, the life rhythm of people is accelerated, the life is irregular, and the insomnia phenomenon caused by various pressures of life, study, work and the like is continuously increased. At present, the insomnia phenomenon becomes an invisible 'infectious disease' which spreads among people, adults are particularly serious, and the number of the insomnia people is on an increasing trend. If the insomnia phenomenon is more serious, the work, learning efficiency and life quality of an individual are seriously influenced, and the serious people can also generate symptoms such as anxiety, even depression and the like; therefore, the sleep is taken as an important way for recovering the body and supplementing energy, and the high-quality sleep plays an extremely important role in the physical and psychological health of the human body. However, the sedative hypnotics, melatonin and other drugs used for treating insomnia at present are easy to influence the organism after long-term administration, and have certain dependence.
Gamma-aminobutyric acid (GABA), a natural functional non-protein active amino acid widely found in animals, plants and microorganisms. Gamma-aminobutyric acid is one of central nervous systems of the brain, the spinal cord and the like of mammals and an important inhibitory nerve transmission transmitter thereof, participates in brain circulation and various metabolic activities, and has a plurality of important physiological functions of soothing the nerves, resisting excitation, effectively relieving anxiety, improving memory, reducing blood pressure, treating epilepsy, relieving senile dementia and the like. In addition, GABA can obviously improve hexokinase activity in TCA circulation, thereby accelerating glucose metabolism in brain, further promoting blood supply and oxygen supply for brain, and having obvious effects of promoting sleep and improving sleep quality. Although GABA in human body can be generated by glutamate in body under the catalysis of glutamate decarboxylase with strong specificity, with the continuous increase of age and the current fast-paced life style and living state of people, the synthesized GABA content in body is continuously reduced, so that the health of body can be maintained only by exogenous supplement.
However, GABA is widely present in nature but its content is low, and thus GABA acting on the body by merely obtaining a sufficient dose by natural ingestion is far from sufficient. The gamma-aminobutyric acid synthesized by the traditional method is low in safety, so that the gamma-aminobutyric acid has great potential safety hazard when being used in food; the cost is relatively high when the plant is collected by a plant enrichment method; the microbial fermentation method is obtained by collecting the produced gamma-aminobutyric acid in the microbial growth and propagation process, has relatively high safety, and is suitable for application of food and medical-grade products.
Therefore, the method for fermenting the yoghourt by using the strain with high yield of the gamma-aminobutyric acid has great significance for human body health and has certain social and economic values for social development.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide the yoghourt with the effects of relieving pressure and helping sleep and the preparation method and the application thereof. The yogurt has the effects of reducing blood pressure, resisting fatigue, relieving nervous tension, effectively relaxing people and the like.
In order to achieve the purpose, the invention adopts the following technical scheme:
in a first aspect, the invention provides a yogurt with functions of relieving pressure and helping sleep, and the yogurt with functions of relieving pressure and helping sleep is prepared from the following raw materials: milk powder, whey protein powder, sodium glutamate, tea theanine, spina date seed juice, rose juice, grape juice, a sweetening agent, a stabilizing agent and water.
In the invention, the yoghourt with the effects of relieving pressure and helping sleep mainly comprises milk powder, white granulated sugar, whey protein powder, sodium glutamate, spina date seed juice, tea theanine, a stabilizer, rose juice and grape juice, and all the components are matched with each other to have a synergistic effect, so that the effects of relieving pressure, soothing nerves and helping sleep are further improved. The health care tea is added with natural extracts such as spina date seed juice, rose juice, grape juice and tea theanine which have the effects of calming heart, soothing nerves and relieving insomnia, wherein the tea theanine is a specific amino acid extracted from tea and has the effects of reducing blood pressure, resisting fatigue, enhancing alpha wave intensity of brain, relieving nervous tension, effectively relaxing people and the like; meanwhile, the safety of GABA synthesized by the existing chemistry has hidden trouble. Therefore, the high-yield GABA strain is used for fermentation, and the gamma-aminobutyric acid with the effects of relieving stress and assisting sleep is accumulated in the yoghourt through microbial fermentation without additionally adding the gamma-aminobutyric acid.
Preferably, the yogurt with the functions of relieving pressure and helping sleep comprises the following preparation raw materials in parts by weight: 10 to 15 portions of milk powder, 0.5 to 2 portions of whey protein powder, 0.01 to 0.2 portion of sodium glutamate, 0.01 to 0.05 portion of tea theanine, 0.5 to 1 portion of spina date seed juice, 0.03 to 0.07 portion of rose juice, 0.5 to 1 portion of grape juice, 4 to 8 portions of sweetening agent, 0.5 to 3 portions of stabilizing agent and 70 to 84 portions of water.
Specific values of the above-mentioned 10 to 15 parts may be, for example, 10 parts, 11 parts, 12 parts, 13 parts, 14 parts, 15 parts, etc.
Specific values of the above-mentioned 0.5 to 2 parts may be, for example, 0.5 part, 0.7 part, 1 part, 1.2 parts, 1.4 parts, 1.6 parts, 1.8 parts, 2 parts, etc.
Specific examples of the amount of the above-mentioned 0.01 to 0.2 parts include 0.01 parts, 0.02 parts, 0.04 parts, 0.06 parts, 0.08 parts, 0.1 parts, 0.12 parts, 0.14 parts, 0.16 parts, 0.18 parts, and 0.2 parts.
Specific values of 0.01 to 0.05 parts may be, for example, 0.01 parts, 0.02 parts, 0.03 parts, 0.04 parts, 0.05 parts, etc.
Specific values of the above-mentioned 0.5 to 1 part may be, for example, 0.5 part, 0.6 part, 0.7 part, 0.8 part, 0.9 part, 1 part, etc.
Specific values of 0.03 to 0.07 part may be, for example, 0.03 part, 0.04 part, 0.05 part, 0.06 part, 0.07 part, and the like.
Specific values of the above-mentioned 4 to 8 parts may be, for example, 4 parts, 4.5 parts, 5 parts, 5.5 parts, 6 parts, 6.5 parts, 7 parts, 7.5 parts, 8 parts, etc.
Specific values of the above-mentioned 0.5 to 3% may be, for example, 0.5 parts, 1 part, 1.5 parts, 2 parts, 2.5 parts, 3 parts, etc.
Preferably, the stabilizer is selected from any one or combination of at least two of soybean polysaccharide, pectin, sodium carboxymethylcellulose, agar, diacetyl tartaric acid ester of mono-and diglycerides, or acetylated distarch phosphate.
Preferably, the stabilizer is a combination of agar, pectin, diacetyl tartaric acid esters of mono-and diglycerides and acetylated distarch phosphate.
Preferably, the mass ratio of the agar to the pectin to the diacetyl tartaric acid ester of mono-and diglycerides to the acetylated distarch phosphate is (2-4): (1-3): (0.5-1.5).
Specific numerical values in the above (2-4) are, for example, 2, 2.2, 2.5, 2.7, 3, 3.2, 3.5, 3.7, 4 and the like.
Specific numerical values in the above (1-3) are, for example, 1, 1.2, 1.5, 1.7, 2, 2.2, 2.5, 2.7, 3 and the like.
Specific values in the above (0.5 to 1.5) are, for example, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, etc.
Preferably, the sweetener is selected from any one or a combination of at least two of white granulated sugar, sucrose, molasses, xylitol, aspartame or acesulfame potassium, preferably white granulated sugar.
In a second aspect, the present invention provides a preparation method of a yogurt with effects of relieving pressure and helping sleep as described in the first aspect, the preparation method comprises the following steps:
(1) Dissolving milk powder in water, standing and hydrating to obtain reconstituted milk; dissolving a sweetening agent and a stabilizing agent in water to obtain a colloidal solution;
(2) Mixing the reconstituted milk obtained in the step (1) with colloidal fluid, and then sequentially heating, homogenizing and sterilizing to obtain raw milk;
(3) Inoculating a fermenting agent to the raw milk obtained in the step (2), and fermenting to obtain fermented milk;
(4) And (4) stirring and demulsifying the fermented milk obtained in the step (3), and mixing with the rest raw materials to obtain the yoghourt with the effects of relieving pressure and helping sleep.
In the invention, gamma-aminobutyric acid is accumulated by fermenting the high-yield GABA strain, no gamma-aminobutyric acid is additionally added, so that the insomnia disorder people can be helped to relieve anxiety, the insomnia symptom is effectively relieved, the number of viable bacteria in the yoghourt is high, the probiotics can be helped to be supplemented to people, the nutrient absorption is promoted, and the balance of intestinal flora is favorably maintained.
Preferably, in the step (1), the temperature of the milk powder dissolved in water is 40-50 ℃, for example, 40 ℃, 42 ℃, 44 ℃, 46 ℃, 48 ℃, 50 ℃ and the like, and the time of the standing hydration is more than 30min, for example, 30min, 32min, 34min, 36min, 38min, 40min and the like.
Preferably, in the step (1), the temperature at which the sweetener and the stabilizer are dissolved in water is 60 to 70 ℃, and may be, for example, 60 ℃, 62 ℃, 64 ℃, 66 ℃, 68 ℃, 70 ℃ or the like.
Preferably, in the step (2), the heating temperature is 50 to 60 ℃, for example, 50 ℃, 52 ℃, 54 ℃, 56 ℃, 58 ℃,60 ℃ and the like.
In the step (2), the homogenization pressure is preferably 20 to 23MPa, and may be, for example, 20MPa, 20.5MPa, 21MPa, 21.5MPa, 22MPa, 22.5MPa, 23MPa, or the like.
Preferably, in step (2), the sterilization temperature is 95-97 ℃, for example 95 ℃, 95.5 ℃,96 ℃, 96.5 ℃,97 ℃ and the like, and the sterilization time is 5-20 min, for example 5min, 6min, 8min, 10min, 12min, 14min, 16min, 18min, 20min and the like.
Preferably, in step (3), the starter culture comprises Streptococcus thermophilus, lactobacillus delbrueckii subsp.
In the invention, four strains of streptococcus thermophilus, lactobacillus delbrueckii subsp bulgaricus, lactobacillus acidophilus and bifidobacterium longum are mutually matched and synergized, and the content of gamma-aminobutyric acid accumulated by fermentation can be further improved, so that the effects of relieving pressure and helping sleep of the yoghourt are further improved.
Preferably, the viable count ratio of the streptococcus thermophilus, the lactobacillus delbrueckii subsp bulgaricus, the lactobacillus acidophilus and the bifidobacterium longum is (2-4): 1-3): 0.1-2.
The "2 to 4" may be, for example, 2, 2.5, 3, 3.5, 4, or the like.
The "1 to 3" may be, for example, 1, 1.5, 2, 2.5, 3, or the like.
The "0.1 to 2" may be, for example, 0.1, 0.5, 1, 1.5, 2, or the like.
Preferably, in step (3), the amount of the starter culture inoculated is 0.02% to 0.04%, and may be, for example, 0.02%, 0.025%, 0.03%, 0.035%, 0.04%, or the like.
Preferably, in the step (3), the temperature of the inoculation is 30 to 40 ℃, and may be, for example, 30 ℃, 32 ℃, 34 ℃, 36 ℃, 38 ℃, 40 ℃ or the like.
Preferably, in step (3), the fermentation temperature is 38-44 ℃, for example 38 ℃, 39 ℃, 40 ℃, 41 ℃, 42 ℃, 43 ℃, 44 ℃ and the like, and the fermentation time is 6-8 h, for example 6h, 6.5h, 7h, 7.5h, 8h and the like.
Preferably, in step (3), the end point of the fermentation is: the texture of the fermented milk is changed from liquid state to solid curd, and the fermentation is terminated when the fermentation acidity is 65-70T (such as 65T, 66T, 67T, 68T, 69T, 70T).
In the step (4), the temperature of the stirring demulsification is 4-10 ℃, such as 4 ℃,5 ℃,6 ℃, 7 ℃,8 ℃,9 ℃,10 ℃ and the like, the rotation speed of the stirring demulsification is 500-1000 rpm, such as 500rpm, 600rpm, 700rpm, 800rpm, 900rpm, 1000rpm and the like, and the time of the stirring demulsification is 5-8 min, such as 5min, 6min, 7min, 8min and the like.
Preferably, in step (4), the temperature of the mixing is 4 to 10 ℃, for example, 4 ℃,5 ℃,6 ℃, 7 ℃,8 ℃,9 ℃,10 ℃ and the like.
Compared with the prior art, the invention has the following beneficial effects:
(1) The yoghourt with the effects of relieving pressure and helping sleep mainly comprises milk powder, white granulated sugar, whey protein powder, sodium glutamate, spina date seed juice, tea theanine, a stabilizer, rose juice and grape juice, all the components are matched with each other, the synergistic effect of relieving pressure and helping sleep is achieved, the accumulation of gamma-aminobutyric acid with the effects of relieving pressure and helping sleep in the yoghourt through microbial fermentation can be improved, and the effects of relieving pressure, soothing nerves and helping sleep (improving the sleep rate, prolonging the sleep time and shortening the sleep latency) are further improved in two aspects.
(2) The pressure-relieving sleep-aiding probiotic yogurt provided by the invention adopts a natural extract with nerve calming and pressure relieving functions, gamma-aminobutyric acid is accumulated by fermenting a high-yield GABA strain without additionally adding gamma-aminobutyric acid, so that the insomnia-handicapped people can be helped to relieve anxiety, the insomnia symptom is effectively relieved, the number of viable bacteria in the yogurt is high, the probiotics can be helped to be supplemented for people, the nutrient absorption is promoted, the balance of intestinal flora is favorably maintained, and the pressure-relieving sleep-aiding probiotic yogurt has important social value and a wide market development prospect.
(3) The streptococcus thermophilus ST36 is compounded with the Lactobacillus delbrueckii subspecies bulgaricus LB42, the Lactobacillus acidophilus LA85 and the Bifidobacterium longum BL21 to serve as a fermentation microbial inoculum, the four strains are mutually matched to realize synergistic interaction, the GABA yield is improved, and the sleep aiding effect of a yoghourt product is enhanced.
(4) The sodium glutamate and the tea theanine not only have good effects of relieving pressure and helping sleep, but also have unexpected and remarkable synergistic promotion effect on improving the yield of GABA.
(5) The milk powder, the whey protein powder, the spina date seed juice, the rose juice and the grape juice have synergistic effect in the aspects of relaxing pressure and helping sleep.
(6) In the research of the invention, the selection of the stabilizer in the yoghourt has the influence on promoting the exertion of the main functional components of the product and further influencing the sleep aiding efficacy of the product, and the stabilizer is preferably the combination of agar, pectin, diacetyl tartaric acid monoglyceride and acetylated distarch phosphate to obtain the best effect.
Detailed Description
The technical solution of the present invention is further explained by the following embodiments. It should be understood by those skilled in the art that the examples are only for the understanding of the present invention and should not be construed as the specific limitations of the present invention.
The following sources of deposits related to the species are shown below:
the Streptococcus thermophilus ST36 has the preservation unit of China general microbiological culture Collection center (CGMCC) No.20676, the preservation date of 9-18 days in 2020, and the preservation address of No. 3 Hospital No.1 on North Chen of the sunward area in Beijing.
Lactobacillus acidophilus (Lactobacillus acidophilus) LA85 is prepared by China general microbiological culture Collection center (CGMCC), with the collection number of CGMCC No.1.12735, the collection date of 2020, 07, 20 days and the collection address of Beijing Shang Yang district No.1, xilu-1, beicheng.
The Lactobacillus delbrueckii subsp. Bulgaricus LB42 is preserved in China general microbiological culture Collection center with the preservation number of CGMCC No.15751, the preservation date of 2018, 05 and 11 days, and the preservation address of No. 3 West Luo No.1 of the sunward area of Beijing.
The following Bifidobacterium longum (Bifidobacterium longum) BL21 is preserved in China general microbiological culture Collection center with preservation time of 2015, 01-27 days, preservation number of CGMCC No.10452 and preservation address of: xilu No.1 Hospital No. 3, beijing, chaoyang, north.
The following examples relate to the following sources of raw materials:
Figure BDA0003896072710000091
example 1
The embodiment provides a yoghourt with the effects of relieving pressure and helping sleep, which comprises the following components in percentage by mass: 12% of milk powder, 0.6% of whey protein powder, 0.05% of sodium glutamate, 0.04% of tea theanine, 1% of spina date seed juice, 0.05% of rose concentrated juice, 1% of grape juice, 7% of white granulated sugar, 2% of a stabilizer and the balance of purified water;
wherein the stabilizer consists of agar, pectin, diacetyl tartaric acid ester of mono-and diglycerides and acetylated distarch phosphate in a mass ratio of 3.
The yoghourt with the functions of relieving pressure and helping sleep is prepared by the following preparation method:
(1) Weighing milk powder in proportion, dissolving into warm water of about 45 deg.C, stirring thoroughly to dissolve, standing for hydrating for 30min to obtain reconstituted milk; dissolving white granulated sugar and stabilizer in pure water at 65 ℃ under stirring to obtain colloidal fluid;
(2) Homogenizing: stirring and mixing the reconstituted milk and colloidal fluid (with a volume ratio of 2:1) uniformly, heating to 55 ℃, homogenizing under 21MPa, sterilizing after homogenizing, sterilizing at 96 ℃ for 10min, and cooling to room temperature;
(3) Inoculating a starter to raw milk cooled to room temperature according to the inoculation amount of 0.04%, wherein the starter is obtained by compounding 4 strains, namely streptococcus thermophilus ST36, lactobacillus delbrueckii subsp bulgaricus LB42, lactobacillus acidophilus LA85 and bifidobacterium longum BL21, and the viable count ratio is 3; stirring uniformly after inoculation, fermenting for 7h at 41 ℃, changing the texture from liquid state to solid curd, and obtaining fermentation acidity of 68 DEG T, namely reaching the fermentation end point;
(4) Cooling fermented milk to normal temperature, stirring for demulsification (800rpm, 8 ℃,6 min), mixing with whey protein powder, sodium glutamate, theanine of tea leaves, semen Ziziphi Spinosae juice, concentrated rose juice and grape juice, aseptically filling, refrigerating overnight at 4 ℃ after filling, and aging to obtain yogurt with effects of relieving stress and improving sleep.
Example 2
The embodiment provides a yoghourt with the functions of relieving pressure and helping sleep, which comprises the following components in percentage by mass: 10% of milk powder, 2% of whey protein powder, 0.1% of sodium glutamate, 0.03% of tea theanine, 0.5% of spina date seed juice, 0.07% of rose concentrated juice, 0.5% of grape juice, 6% of white granulated sugar, 1.5% of stabilizer and the balance of purified water;
wherein the stabilizer consists of agar, pectin, diacetyl tartaric acid ester of mono-and diglycerides and acetylated distarch phosphate in a mass ratio of 4.
The yoghourt with the effects of relieving pressure and helping sleep is prepared by the following preparation method:
(1) Weighing milk powder in proportion, dissolving into warm water of about 40 deg.C, stirring thoroughly to dissolve, standing for hydration for 35min to obtain reconstituted milk; dissolving white granulated sugar and stabilizer in pure water at 70 deg.C under stirring to obtain colloid liquid;
(2) Homogenizing: stirring the hydrated reconstituted milk and the colloidal fluid, mixing uniformly, heating to 50 ℃, homogenizing under the homogenizing pressure of 23MPa, sterilizing after homogenizing, sterilizing at 97 ℃ for 12min, and cooling to room temperature;
(3) Inoculating a starter to raw milk cooled to room temperature according to the inoculation amount of 0.03%, wherein the starter is obtained by compounding 4 strains, namely streptococcus thermophilus ST36, lactobacillus delbrueckii subsp bulgaricus LB42, lactobacillus acidophilus LA85 and bifidobacterium longum BL21, and the viable count ratio of the starter is (4); stirring uniformly after inoculation, fermenting for 7h at 42 ℃, changing the texture from liquid state to solid curd, and obtaining fermentation acidity of 70 DEG T, namely reaching the fermentation end point;
(4) Cooling fermented milk to normal temperature, stirring for demulsification (500rpm, 10 ℃,8 min), mixing with whey protein powder, sodium glutamate, theanine of tea leaves, semen Ziziphi Spinosae juice, concentrated rose juice and grape juice, aseptically filling, refrigerating overnight at 4 ℃ after filling, and aging to obtain yogurt with effects of relieving stress and improving sleep.
Example 3
The embodiment provides a yoghourt with the functions of relieving pressure and helping sleep, which comprises the following components in percentage by mass: 15% of milk powder, 0.5% of whey protein powder, 0.15% of sodium glutamate, 0.05% of theanine, 1% of spina date seed juice, 0.03% of rose concentrated juice, 1% of grape juice, 5% of white granulated sugar, 3% of stabilizer and the balance of purified water;
wherein the stabilizer consists of agar, pectin, diacetyl tartaric acid ester of mono-and diglycerides and acetylated distarch phosphate in a mass ratio of 2.
The yoghourt with the effects of relieving pressure and helping sleep is prepared by the following preparation method:
(1) Weighing milk powder in proportion, dissolving into warm water of about 50 deg.C, stirring thoroughly to dissolve, standing for hydration for 32min to obtain reconstituted milk; dissolving white granulated sugar and stabilizer in pure water at 60 deg.C under stirring to obtain colloidal fluid;
(2) Homogenizing: stirring the hydrated reconstituted milk and the colloidal fluid, mixing uniformly, heating to 60 ℃, homogenizing under the homogenizing pressure of 20MPa, sterilizing after homogenizing, sterilizing at 95 ℃ for 15min, and cooling to room temperature;
(3) Inoculating a starter to raw milk cooled to room temperature according to the inoculation amount of 0.04%, wherein the starter is obtained by compounding 4 strains, namely streptococcus thermophilus ST36, lactobacillus delbrueckii subsp bulgaricus LB42, lactobacillus acidophilus LA85 and bifidobacterium longum BL21, and the viable count ratio is 2.5; stirring uniformly after inoculation, fermenting for 7 hours at 40 ℃, changing the texture from liquid state to solid curd, and obtaining fermentation acidity of 67 DEG T, namely reaching the fermentation end point;
(4) Cooling fermented milk to normal temperature, stirring for demulsification (1000rpm, 4 deg.C, 5 min), mixing with whey protein powder, sodium glutamate, tea theanine, semen Ziziphi Spinosae juice, flos Rosae Rugosae concentrated juice and succus Vitis Viniferae, aseptically packaging, refrigerating at 4 deg.C overnight after packaging, and aging to obtain yogurt with effects of relieving stress and improving sleep.
Example 4
This example provides a yogurt with effects of relieving stress and improving sleep, which is different from example 1 only in that the content of sodium glutamate is reduced to 0.008%, the content of theanine in tea leaves is increased to 0.06%, and the content of other components and the preparation method are the same as example 1.
Example 5
This example provides a yogurt with effects of relieving stress and improving sleep, which is different from example 1 only in that the content of sodium glutamate is increased to 0.25%, the content of theanine in tea leaves is reduced to 0.008%, and the content of other components and the preparation method are the same as example 1.
Example 6
The present example provides a yoghurt with effects of relieving pressure and helping sleep, which is different from example 1 only in that the content of the wild jujube seed juice is reduced to 0.02%, the content of the rose concentrated juice is increased to 2%, the content of the grape juice is reduced to 0.03%, and the content of other components and the preparation method are the same as example 1.
Example 7
This example provides a yogurt with effects of relieving stress and improving sleep, which is different from example 1 only in that the content of the wild jujube seed juice is increased to 0.4%, the content of the rose concentrated juice is decreased to 0.05%, the content of the grape juice is decreased to 1.6%, and the content of other components and the preparation method are the same as example 1.
Example 8
The present example provides a yogurt with effects of relieving pressure and helping sleep, which is different from example 1 only in that the stabilizer is composed of agar, soybean polysaccharide, sodium carboxymethyl cellulose and acetylated distarch phosphate in a mass ratio of 3.
Example 9
The present example provides a yogurt with effects of relieving pressure and promoting sleep, which is different from example 1 only in that the stabilizer is composed of soybean polysaccharide, pectin and acetylated distarch phosphate in a mass ratio of 4.
Example 10
The present example provides a yogurt with pressure-relieving and sleep-aiding effects, which is different from example 1 only in that lactobacillus delbrueckii subsp bulgaricus is not added to the leavening agent, the leavening agent is composed of streptococcus thermophilus ST36, lactobacillus acidophilus LA85 and bifidobacterium longum BL21 with a viable count ratio of 3.
Example 11
The present example provides a yogurt with pressure-relieving and sleep-aiding effects, which is different from example 1 only in that no lactobacillus acidophilus is added to the leavening agent, the leavening agent is composed of streptococcus thermophilus ST36, lactobacillus delbrueckii subsp bulgaricus LB42 and bifidobacterium longum BL21, the ratio of viable bacteria is 3.
Example 12
The present example provides a yogurt with pressure-relieving and sleep-aiding effects, which is different from example 1 only in that bifidobacterium longum is not added to the starter culture, the starter culture consists of streptococcus thermophilus ST36, lactobacillus delbrueckii subsp bulgaricus LB42 and lactobacillus acidophilus LA85, the viable count ratio is 3.
Comparative example 1
The comparison example provides a yoghurt, which is different from the yoghurt of example 1 only in that whey protein powder is not added, the content of milk powder is increased to 12.6%, and the content of other components and the preparation method are consistent with those of example 1.
Comparative example 2
This comparative example provides a yoghurt which differs from example 1 only in that, without the addition of sodium glutamate, the theanine content of the tea leaves is increased to 0.09%, the contents of other components and the preparation method are identical to example 1.
Comparative example 3
This comparative example provides a yoghurt which differs from example 1 only in that theanine is not added, the sodium glutamate content is increased to 0.09%, the other component content and the preparation method are identical to example 1.
Comparative example 4
The comparative example provides a yoghurt, which is different from example 1 only in that no spina date seed juice is added, the content of the rose concentrated juice is increased to 0.55%, the content of the grape juice is increased to 1.5%, and the content of other components and the preparation method are consistent with example 1.
Comparative example 5
The comparative example provides a yoghurt, which differs from example 1 only in that, without adding rose concentrated juice, the content of spine date seed juice is increased to 1.02%, the content of grape juice is increased to 1.03%, and the content of other components and the preparation method are the same as example 1.
Comparative example 6
The comparative example provides a yoghurt, which differs from example 1 only in that, without the addition of grape juice, the content of wild jujube juice is increased to 1.5%, the content of rose concentrated juice is increased to 0.55%, and the content of other components and the preparation method are the same as example 1.
Test example 1
GABA content test
Test samples: yoghurts provided in examples 1-3, 4-5, 10-12 and comparative examples 2-3;
and (3) testing items: after the preparation of the yoghourt is finished, tracking and measuring the GABA content change in the shelf life (21 d), and observing the stability in the shelf life;
the test method comprises the following steps: a GABA content change tracking determination method comprises the steps of determining gamma-aminobutyric acid content in yoghourt by using a high performance liquid chromatography, weighing a prepared yoghourt sample, centrifuging at 6000r/min for 10min, taking supernatant, taking out 1.5mL of supernatant by using a liquid transfer gun, adding triethylamine acetonitrile solution and PITC acetonitrile solution, standing for reaction for 1h at normal temperature after uniform oscillation, adding n-hexane, fully oscillating and shaking uniformly, standing for 10min, taking out lower layer solution after layering, diluting, and detecting in a high performance liquid chromatography;
the specific test results are shown in table 1:
TABLE 1
Figure BDA0003896072710000161
As shown in Table 1, in the shelf life (21 d) of the yogurt, the GABA content in the yogurt gradually increases (due to a series of reactions such as a small amount of free amino acids generated by proteolytic activity, the GABA content slightly increases), wherein the 21d content is the highest and is more than 0.60g/L, and compared with the 0d content, the increasing rate is about 7%. The components in the yoghourt with the functions of relieving pressure and helping sleep are matched with each other, and the strain is promoted to ferment and accumulate gamma-aminobutyric acid, so that the functions of relieving pressure and helping sleep of the yoghourt are improved.
Test example 2
Viable count test
Test samples: yoghurts with stress-relieving sleep-aiding efficacy of examples 1-12;
the test method comprises the following steps: a method for tracking and measuring the number of viable bacteria in shelf life, which is measured by a flat plate bacterial colony counting method;
the specific test results are shown in table 2:
TABLE 2
Figure BDA0003896072710000171
As shown in Table 2, the yoghourt with the functions of relieving pressure and assisting sleep has the advantages that the survival rate of viable bacteria is better in the shelf life (21 d) of the yoghourt, and the survival rate is more than 80%, so that the yoghourt with the functions of relieving pressure and assisting sleep mainly comprises milk powder, white granulated sugar, whey protein powder, sodium glutamate, spina date seed juice, tea theanine, a stabilizer, rose concentrated juice and grape juice, all the components are matched with each other, the synergism effect is realized, the GABA content can be improved, and the survival of the viable bacteria in the yoghourt can be well maintained.
Test example 3
Acidity test
Test samples: yoghurts with pressure-relieving sleep-aiding efficacy of examples 1-12;
and (3) testing items: after the yoghourt is prepared, the change of acidity and the change of flavor in the shelf life (21 d) are tracked and measured;
the specific test results are shown in table 3:
TABLE 3
Figure BDA0003896072710000181
The test results in the table 3 show that the sour milk with the functions of relieving pressure and helping sleep provided by the invention has stable acidity change in the shelf life (21 d) of the sour milk, and the acidity is 83-87 DEG T; and the flavor of the yoghourt is rich within 3-14 days of shelf life and has strong fragrance.
Test example 4
Yoghourt sleeping-aid animal experiment
Test samples: the yoghurts of examples 1-12 having pressure-relieving sleep-aiding efficacy and the yoghurts provided in comparative examples 1-6;
the test method comprises the following steps:
(1) Test animal feeding and selection
Male mice of healthy adults with a body weight of 18-20g were selected and were acclimatized for 2 weeks while controlling ambient temperature (22. + -. 2 ℃) and humidity (55%. + -. 10%), with a 12-hour light-dark cycle, and were free to ingest food and water. After adaptive feeding, mice were divided into blank and model groups.
(2) Insomnia mouse model making mould
The mouse insomnia model is made by PCPA (chlorophenylalanine) intraperitoneal injection molding method: adjusting the pH value of a 0.9% NaCl solution to be alkalescent (the pH value is 7.0-8.0), dissolving the PCPA in the adjusted alkalescent physiological saline, standing overnight, and preparing a PCPA suspension.
And (3) carrying out intraperitoneal injection on the molded mouse in time in the morning every day, wherein the injection amount is adjusted according to the weight of the mouse and is 400mg/kg, meanwhile, the blank group is continuously injected with normal saline with the same dose into the abdominal cavity for 2 days, and when the mouse disappears the day-night rhythm and constantly moves in the day and at night, the insomnia mouse model is successfully built.
(3) After the mice modeling is completed, the model group mice are divided into an insomnia model group, a GABA yoghourt intervention group (examples 1-12, comparative examples 1-6) and a positive control group, and each group of mice contains 10 mice. The GABA yoghourt intervention group is used for feeding prepared yoghourt for intragastric administration, and the GABA yoghourt intragastric administration amount is 100mg/kg; the blank group and the insomnia model group are subjected to intragastric perfusion with the same amount of physiological saline; and (3) administering diazepam liquid (sleep-aiding medicine) to the control group for intragastric administration, wherein the diazepam liquid is prepared by dissolving diazepam powder in physiological saline, and the administration amount is adjusted in time according to the weight change of the mice.
(4) The judgment standard for the sleep of the mouse is as follows: the mice lie on the flat plate on their backs, and the mice do not turn over for 1min, which means that the righting reflex of the mice disappears and the mice go to sleep. If the mouse turns 3 times within half a minute, it indicates that the mouse righting reflex is recovered and the mouse is awake. The time from the start of administration to the disappearance of the righting reflex is the sleep latency, and the time from the disappearance of the righting reflex to the restoration of the righting reflex is the sleep time of the mouse.
(5) Mouse sleep experiment: each group of mice is administered by intragastric administration for 30 days, and after the last intragastric administration for 30min, the intraabdominal cavity of each group of mice is injected with pentobarbital sodium, the injection amount is adjusted according to the surface area change of the mice, and the injection dose is 100 mu L/10g & BW. And calculating the sleeping rate of the test mouse according to the sleeping condition of the mouse.
(6) Sleep time experiments: each group of mice is administered by intragastric administration for 1 time per day for 30 days continuously, and after the last intragastric administration for 30min, each group of mice is injected with pentobarbital sodium intraperitoneally, the injection amount is adjusted according to the change of the body surface area of the mice, and the injection dose is 100 mu L/10g & BW. The length of time the mice had slept was recorded.
(7) Sleep latency experiments: each group of mice is subjected to intragastric administration for 1 time per day for 30 days continuously, and after 30min of the last intragastric administration, the intraperitoneal injection of the barbital sodium is performed on each group of mice, the injection amount is adjusted according to the surface area change of the mice, and the injection dose is 100 mu L/10g & BW. The observation recorded the sleep latency of the mice.
(8) Hypnosis test result of mice of each test group on sodium pentobarbital subthreshold dose
The specific test results are shown in table 4:
TABLE 4
Figure BDA0003896072710000201
Figure BDA0003896072710000211
The test results in table 4 show that the sleep incidence rate of the mice in the yogurt group provided by the invention is more than 70%, compared with the blank control group, the sleep effect of the mice in the GABA yogurt gavage group is obviously higher than that of the blank group, and although the effect is slightly worse than that of the control group (positive), the mice are obviously improved by taking the yogurt containing GABA.
(9) Test result of mouse pairs of test groups for prolonging sleep time of sodium pentobarbital
The specific test results are shown in table 5:
TABLE 5
Figure BDA0003896072710000212
Figure BDA0003896072710000221
As can be seen from the test results in Table 5, the sleeping time of the blank control group is 53min; the longest sleeping time of the control group mice is 123min; the time for the mice in the GABA-containing yogurt intervention group provided by the application to fall asleep reaches over 86 min; the experimental mouse can obviously prolong the sleeping time of the mouse after ingesting the yoghourt containing GABA, but the effect of the yoghourt is different from that of a control group, and the GABA yoghourt only plays a certain role in relieving the sleep disorder and can not completely achieve the effect equivalent to that of a hypnotic.
(10) Experimental results of sleep latency of mice
The specific test results are shown in table 6:
TABLE 6
Figure BDA0003896072710000222
As can be seen from the test results in Table 6, the sleep latency time of the blank control group mice is 30min; the sleep latency time of the mice in the yogurt intervention group is relatively reduced, and is below 27min, so that the mice have a certain relieving effect on shortening the sleep latency time under the intervention of the yogurt with the effects of relieving pressure and helping sleep, and the sleep duration and the sleep quality of the mice can be further improved; the sleep latency time of the control mice was relatively short, 22min. The above experimental results show that the yogurt with the effects of relieving pressure and helping sleep has obvious effects on improving the sleep rate of mice, prolonging the sleep time and shortening the sleep latency period, and the yogurt with the effects of relieving pressure and helping sleep has obvious effects on relieving sleep, improving the sleep time and helping sleep on organisms after being taken, but the yogurt with the effects of relieving pressure and helping sleep only has an effect of relieving insomnia and cannot replace the equivalent effects of medicines.
The applicant states that the present invention is illustrated by the above examples, but the present invention is not limited to the above process steps, i.e. the present invention does not mean that the present invention must rely on the above process steps to be implemented. It will be apparent to those skilled in the art that any modification of the present invention, equivalent substitutions of selected materials and additions of auxiliary components, selection of specific modes and the like, which are within the scope and disclosure of the present invention, are contemplated by the present invention.

Claims (10)

1. The yoghourt with the effects of relieving pressure and helping sleep is characterized by comprising the following preparation raw materials: milk powder, whey protein powder, sodium glutamate, tea theanine, spina date seed juice, rose juice, grape juice, a sweetening agent, a stabilizing agent and water.
2. The yoghourt with the functions of relieving pressure and helping sleep as claimed in claim 1, wherein the yoghourt with the functions of relieving pressure and helping sleep is prepared from the following raw materials in parts by weight: 10 to 15 portions of milk powder, 0.5 to 2 portions of whey protein powder, 0.01 to 0.2 portion of sodium glutamate, 0.01 to 0.05 portion of tea theanine, 0.5 to 1 portion of spina date seed juice, 0.03 to 0.07 portion of rose juice, 0.5 to 1 portion of grape juice, 4 to 8 portions of sweetening agent, 0.5 to 3 portions of stabilizing agent and 70 to 84 portions of water.
3. Yoghurt with pressure-relieving and sleep-helping effects according to claim 1 or 2, wherein the stabilizer is selected from any one or a combination of at least two of soybean polysaccharide, pectin, sodium carboxymethyl cellulose, agar, diacetyl tartaric acid esters of mono-and diglycerides, or acetylated distarch phosphate;
preferably, the stabilizer is a combination of agar, pectin, diacetyl tartaric acid esters of mono-and diglycerides and acetylated distarch phosphate.
4. Yogurt with stress-relieving and sleep-improving effects according to claim 3, wherein the mass ratio of the agar, the pectin, the diacetyl tartaric acid esters of mono-and diglycerides and the acetylated distarch phosphate is (2-4): (1-3): 0.5-1.5).
5. Yoghurt with pressure-relieving and sleep-helping effects according to any one of claims 1 to 4, wherein the sweetener is selected from any one or a combination of at least two of white granulated sugar, sucrose, molasses, xylitol, aspartame or acesulfame potassium, preferably white granulated sugar.
6. A method for preparing yoghurt with the functions of relieving pressure and helping sleep as claimed in any one of claims 1 to 5, wherein the preparation method comprises the following steps:
(1) Dissolving milk powder in water, standing and hydrating to obtain reconstituted milk; dissolving a sweetening agent and a stabilizing agent in water to obtain a colloidal solution;
(2) Mixing the reconstituted milk obtained in the step (1) with colloidal fluid, and then sequentially heating, homogenizing and sterilizing to obtain raw milk;
(3) Inoculating a fermenting agent to the raw milk obtained in the step (2), and fermenting to obtain fermented milk;
(4) And (4) stirring and demulsifying the fermented milk obtained in the step (3), and mixing with the rest of preparation raw materials to obtain the yoghourt with the effects of relieving pressure and helping sleep.
7. The preparation method according to claim 6, wherein in the step (1), the temperature for dissolving the milk powder in water is 40-50 ℃, and the time for standing hydration is more than 30min;
preferably, in the step (1), the temperature for dissolving the sweetener and the stabilizer in water is 60-70 ℃.
8. The method according to claim 6 or 7, wherein in the step (2), the heating temperature is 50 to 60 ℃;
preferably, in the step (2), the homogenizing pressure is 20-23 MPa;
preferably, in the step (2), the temperature for sterilization is 95-97 ℃, and the time for sterilization is 5-20 min.
9. The method according to any one of claims 6 to 8, wherein in the step (3), the fermentation agent comprises Streptococcus thermophilus, lactobacillus delbrueckii subsp bulgaricus, lactobacillus acidophilus and Bifidobacterium longum;
preferably, the viable count ratio of the streptococcus thermophilus, the lactobacillus delbrueckii subsp bulgaricus, the lactobacillus acidophilus and the bifidobacterium longum is (2-4) to (1-3) to (0.1-2);
preferably, in the step (3), the inoculation amount of the leavening agent is 0.02-0.04%;
preferably, in the step (3), the temperature of the inoculation is 30-40 ℃;
preferably, in the step (3), the fermentation temperature is 38-44 ℃, and the fermentation time is 6-8 h;
preferably, in step (3), the end point of the fermentation is: the texture of the fermented milk is changed from liquid state to solid curd, and the fermentation acidity is 65-70 DEG T, so that the fermentation is stopped.
10. The preparation method according to any one of claims 6 to 9, wherein in the step (4), the temperature of the stirring demulsification is 4-10 ℃, the rotating speed of the stirring demulsification is 500-1000 rpm, and the time of the stirring demulsification is 5-8 min;
preferably, in the step (4), the temperature of the mixing is 4 to 10 ℃.
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