阅读说明：本技术 一种提高免疫力的常温长保质期无蔗糖酸奶及其制备方法 (Normal-temperature long-shelf-life sucrose-free yoghourt capable of improving immunity and preparation method thereof ) 是由 刘冠辰 赵越 陈苏 陈丽娥 谈文诗 王健 冯玉红 欧凯 李言郡 于 2020-06-09 设计创作，主要内容包括：本发明涉及食品技术领域,公开了一种提高免疫力的常温长保质期无蔗糖酸奶及其制备方法。该提高免疫力的常温长保质期无蔗糖酸奶包括原料奶或乳粉、提高免疫力功效成分；所述提高免疫力功效成分至少包括瑞士乳杆菌和/或其突变体。本发明的酸奶由牛乳或复原乳经瑞士乳杆菌发酵而成,复配多种提高免疫力的天然功效成分,长期食用可提高人体免疫力；并且,本发明中所用的瑞士乳杆菌WHH2580在灭活状态下仍然具有增强免疫作用,因而本发明的酸奶能在常温条件下正常保存,无需冷藏,且保质期长。(The invention relates to the technical field of food, and discloses normal-temperature long-shelf-life sucrose-free yoghourt capable of improving immunity and a preparation method thereof. The normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity comprises raw milk or milk powder and functional components capable of improving the immunity; the functional component for improving immunity at least comprises lactobacillus helveticus and/or a mutant thereof. The yoghourt is prepared by fermenting cow milk or reconstituted milk with lactobacillus helveticus, is compounded with various natural functional components for improving the immunity, and can improve the immunity of a human body after being eaten for a long time; in addition, the lactobacillus helveticus WHH2580 used in the invention still has the function of enhancing immunity in an inactivated state, so that the yoghurt can be normally stored at normal temperature without refrigeration and has long shelf life.)

1. A normal-temperature long-shelf-life sucrose-free yoghourt capable of improving immunity is characterized by comprising raw milk or milk powder and functional components capable of improving immunity; the immunity-improving functional component at least comprises lactobacillus helveticus and/or a mutant thereof;

the lactobacillus helveticus is named as WHH2580, and is preserved in the common microorganism center of the china culture collection and management committee for culture collection of microorganisms in 2019, month 10 and 23, and the microorganism preservation numbers are as follows: CGMCC No.18730, and the classification name of microorganisms is Lactobacillus helveticusLactobacillus helveticus(ii) a The Lactobacillus helveticus has a 16s rRNA gene sequence shown in SEQ ID NO. 1; the mutant is obtained by carrying out mutagenesis, domestication, gene recombination or natural mutation on the lactobacillus helveticus.

2. The normal-temperature long-shelf-life sucrose-free yogurt for improving immunity of claim 1, further comprising sugar alcohol, starch, stabilizer, sweetener.

3. The normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity as claimed in claim 2, which comprises, by mass, 40% -85% of raw milk or 5-15% of milk powder, 0.001-0.020% of lactobacillus helveticus and/or mutants thereof, 0.01-10.00% of an immunity-improving effective component excluding lactobacillus helveticus and/or mutants thereof, 0.5-5.0% of sugar alcohol, 0.1-5.0% of starch, 0.01-1.00% of a stabilizer, 0.001-0.050% of a sweetener, and the balance of water; the sum of all the raw materials is 100 percent.

4. The normal-temperature long-shelf-life sucrose-free yogurt for improving immunity of claim 2 or 3, characterized in that:

the milk powder is one or more of whole milk powder, skim milk powder, whey protein powder and whole milk protein powder; and/or

The functional components for improving immunity also comprise one or more of propolis, Ganoderma extract, wall-broken Ganoderma spore powder, beta-dextran, herba Dendrobii extract, herba Sambuci Williamsii, and acerola cherry; and/or

The sugar alcohol is one or more of xylitol, maltitol, erythritol, sorbitol, isomaltitol and lactitol; and/or

The starch is one or more of physical starch, acetate starch, sodium starch octenyl succinate, hydroxypropyl distarch phosphate, phosphate distarch, acid-treated starch, oxidized hydroxypropyl starch, acetylated distarch phosphate and acetylated distarch adipate; and/or

The stabilizer is one or more of sodium carboxymethylcellulose, pectin, soybean polysaccharide, agar, sodium caseinate, propylene glycol alginate, gelatin, gellan gum, guar gum and xanthan gum; and/or

The sweetener is one or more of acesulfame potassium, aspartame, sucralose, stevioside, mogroside, isomaltulose, edmuntin and neotame.

5. A method for preparing the sucrose-free yogurt with the normal temperature and the long shelf life for improving the immunity according to any one of claims 2 to 4, which is characterized by comprising the following steps:

scheme A:

(1) homogenizing and sterilizing: preheating raw milk, homogenizing, sterilizing, and cooling to obtain sterilized cooling feed liquid;

(2) inoculating and fermenting: adding lactobacillus helveticus and/or a mutant thereof into the sterilized cooling liquid obtained in the step (1), uniformly stirring, standing at a constant temperature, and fermenting to obtain a yogurt base material;

(3) preparing glue solution: preheating water, adding a formula amount of stabilizer into the water, shearing or stirring, and cooling to obtain a glue solution;

(4) secondary burdening: cooling the yogurt base material obtained in the step (2), and then stirring and demulsifying; adding the glue solution obtained in the step (3), adding the components with the formula dosage, sugar alcohol, sweetener and starch for improving the immunity except the lactobacillus helveticus and/or the mutant thereof, mixing and stirring, and adding water to a constant volume to obtain a feed liquid;

(5) and (3) secondary homogenization: homogenizing the feed liquid obtained in the step (4);

(6) filling and sterilizing: filling, sealing, pasteurizing and cooling the homogenized material liquid in the step (5) to obtain the normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity; or (5) carrying out UHT sterilization on the homogenized material liquid, carrying out hot filling or aseptic cold filling, and sealing to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature;

scheme B:

1) preparing milk: heating water, adding the formula amount of milk powder into the water, stirring or shearing, and standing for hydration to obtain milk;

2) homogenizing and sterilizing: preheating the milk obtained in the step 1), homogenizing, sterilizing and cooling to obtain a sterilized cooling feed liquid;

3) inoculating and fermenting: adding lactobacillus helveticus and/or a mutant thereof into the sterilized cooling liquid obtained in the step 2), uniformly stirring, standing at a constant temperature, and fermenting to obtain a yogurt base material;

4) preparing glue solution: preheating water, adding a formula amount of stabilizer into the water, shearing or stirring, and cooling to obtain a glue solution;

5) secondary burdening: cooling the yogurt base material obtained in the step 3), and then stirring and demulsifying; adding the glue solution obtained in the step 4), adding the components with the formula dosage, sugar alcohol, sweetener and starch for improving the immunity except the lactobacillus helveticus and/or the mutant thereof, mixing and stirring, and adding water to a constant volume to obtain a feed liquid;

6) and (3) secondary homogenization: homogenizing the feed liquid obtained in the step 5);

7) filling and sterilizing: filling, sealing, pasteurizing and cooling the homogenized material liquid obtained in the step 6) to obtain the normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity; or carrying out UHT sterilization on the homogenized material liquid in the step 6), carrying out hot filling or aseptic cold filling, and sealing to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature.

6. The method for preparing the sucrose-free yogurt with the normal temperature and the long shelf life for improving the immunity according to claim 5, wherein the method comprises the following steps:

in the step 1), the mass of the water is 200-400 g; and/or

In the step 1), water is heated to 45-60 ℃; and/or

In the step 1), the stirring or shearing time is 15-30 min; and/or

In the step 1), standing and hydrating for 20-40 min.

7. The method for preparing the sucrose-free yogurt with the normal temperature and the long shelf life for improving the immunity according to claim 5, wherein the method comprises the following steps:

in the step (1) or the step 2), preheating to the temperature of 60-65 ℃; and/or

In the step (1) or the step 2), the homogenizing pressure is 0-8/5-30 MPa; and/or

In the step (1) or the step 2), the sterilization temperature is 90-97 ℃, and the time is 3-10 min; and/or

In the step (1) or the step 2), cooling is carried out until the temperature is 35-45 ℃.

8. The method for preparing the sucrose-free yogurt with the normal temperature and the long shelf life for improving the immunity according to claim 5, wherein the method comprises the following steps:

in the step (2) or the step (3), the fermentation temperature is 35-45 ℃ and the fermentation time is 4-16 h; and/or

In the step (2) or the step 3), the number of viable bacteria is more than or equal to 1 × 10 in the fermentation process⁸cfu/mL。

9. The method for preparing the sucrose-free yogurt with the normal temperature and the long shelf life for improving the immunity according to claim 5, wherein the method comprises the following steps:

in the step (3) or the step 4), the mass of the water is 200-400 g; and/or

In the step (3) or the step 4), water is preheated to 50-60 ℃; and/or

In the step (3) or the step (4), the shearing or stirring time is 20-40 min; and/or

In the step (3) or the step (4), the temperature is cooled to be below 30 ℃.

10. The method for preparing the sucrose-free yogurt with the normal temperature and the long shelf life for improving the immunity according to claim 5, wherein the method comprises the following steps:

in the step (4) or the step (5), cooling to the temperature below 15 ℃; and/or

In the step (4) or the step 5), the volume is adjusted to 1000mL by using water; and/or

In the step (5) or the step 6), the homogenizing pressure is 0-8/5-30 MPa.

Technical Field

The invention relates to the technical field of food, in particular to normal-temperature long-shelf-life sucrose-free yoghourt capable of improving immunity and a preparation method thereof.

Background

The season of autumn and winter is a period with more diseases every year, various epidemic diseases including influenza and the like begin to be too heavily abused, and children and old people with lower immunity are more serious areas of diseases. It is the "immunity" that is the best doctor. The measures of wearing a mask, frequently washing hands and the like can only prevent external virus invasion, and the immunity of the patient is the only strength on which the patient can rely once the patient is infected without specific medicines.

The milk is rich in nutrition, and contains multiple unique components capable of regulating immune function, such as immunoglobulin, lactoferrin and multiple amino acids. The lysozyme and lactoperoxidase contained in the dairy product can play an anti-infection role in the intestines, activate an immune system, enhance the activity of immune cells or improve the resistance of organisms to pathogenic bacteria, and protect the immune function of the bodies.

At present, a great number of documents report that probiotics can improve human immunity, for example, lactobacillus reuteri can be colonized in intestinal tracts, improve the distribution of intestinal flora, antagonize harmful bacteria, grow and improve host immunity. Meanwhile, the lactobacillus reuteri can generate 'reuterin', and can widely inhibit the growth of gram-negative bacteria, gram-positive bacteria, yeast, fungi, pathogenic protozoa and the like; the lactobacillus rhamnosus LGG can increase the secretory volume of IgA of intestinal mucosa, and the pathogenic bacteria of tissues infect epithelial cells to enhance the immunity; the Lactobacillus plantarum Lp9 and Lp91 can secrete anti-inflammatory cytokines in HT-29 cells induced by LPS and regulate immune functions. German research also shows that D-phenyllactic acid produced by fermentation of lactic acid bacteria can activate the G protein receptor HCA3 of human immune system, induce chemotaxis of neutrophil, and improve immunity. Brazil researchers found that acetate produced by lactic acid bacteria fermentation could stimulate the human GPR43-1 type interferon response against respiratory syncytial virus.

Propolis, ganoderma lucidum extract, wall-broken ganoderma lucidum spore powder, beta-glucan, dendrobium officinale extract, elderberry, acerola cherry and other food raw materials are generally considered to be rich in substances which are helpful to the immune function, such as flavone, polyphenol, ganoderic acid, fungus polysaccharide, mucopolysaccharide, anthocyanin, vitamin C and the like.

When the cow milk capable of improving the human immunity is fermented by probiotics capable of improving the human immunity, and is supplemented with an effective component capable of improving the human immunity, the three components are combined, the synergistic effect is achieved, and the effect of enhancing the immunity can be achieved well.

At present, as shown in fig. 2, the yogurt which is declared to enhance immunity in the market has the function of regulating immunity only when probiotics used are in a survival state, so that the yogurt can be stored only in a refrigeration mode, the shelf life is short (generally 15-21 days), and sugar substances such as white granulated sugar and the like are required to be added as a carbon source to maintain the activity of strains. The refrigeration storage means that the product has strict requirements on the storage, logistics and sale environments and has higher intermediate cost; the short shelf life leads to easy overdue of the product and waste; the white granulated sugar is required to be one of the intentions of obesity caused by the century, and is widely suffered by consumers and industries at present. At present, the normal-temperature long-shelf-life (3-6 months) sucrose-free yoghourt capable of improving immunity is still a market blank.

Disclosure of Invention

In order to solve the technical problems, the invention provides the sucrose-free yoghourt with the normal temperature and the long shelf life and the preparation method thereof, which can improve the immunity. The yogurt can effectively improve immunity, can be normally stored at normal temperature, does not need refrigeration, and has long shelf life.

The specific technical scheme of the invention is as follows:

a sucrose-free yogurt with long shelf life at normal temperature for improving immunity comprises raw milk or milk powder, and functional components for improving immunity; the functional component for improving immunity at least comprises lactobacillus helveticus and/or a mutant thereof;

the lactobacillus helveticus is named as WHH2580, and is preserved in the common microorganism center of the china culture collection and management committee for culture collection of microorganisms in 2019, month 10 and 23, and the microorganism preservation numbers are as follows: CGMCC No.18730, and the microorganism classification is named as Lactobacillus helveticus; the Lactobacillus helveticus has a 16s rRNA gene sequence shown in SEQ ID NO. 1; the mutant is obtained by carrying out mutagenesis, domestication, gene recombination or natural mutation on the lactobacillus helveticus.

The lactobacillus helveticus WHH2580 used in the invention is a probiotic strain screened from the yoghourt collected from the Hongshan ditch of Xinjiang nalat, and animal experiments prove that the lactobacillus helveticus strain has excellent adhesion performance, can smoothly reach the intestinal tract and is adhered to the epithelial cells of the intestinal tract to play a probiotic effect; the strain can improve in-vitro spleen lymphocyte proliferation and interleukin IL-12 secretion of spleen lymphocytes, and promote the secretion of interleukin IL-8 by human colon cancer epithelial cells HT-29; in addition, in an animal model with low immunity, the strain can also obviously improve the organ index of a mouse, promote the proliferation of splenic lymphocytes and improve the activity of NK cells. Therefore, the lactobacillus helveticus WHH2580 has a good effect of enhancing immunity. More importantly, the strain still has better immune function enhancement after inactivation. In addition, in the animal experiment process, the adverse states of death, lassitude, inappetence and the like of the experimental animal are not found, which indicates that the strain has higher safety.

Specifically, the lactobacillus helveticus WHH2580 has the following advantages:

(1) both living and non-living bodies have excellent adhesive properties: the cell adhesion performance of the strain is remarkably superior to that of a positive control strain of Lactobacillus rhamnosus LGG and a field strain LcS, and the number of single-cell adhesion living bacteria reaches 15.69 +/-5.94, and the number of single-cell adhesion non-living bacteria reaches 11.59 +/-4.02. The strain can adhere to intestinal epithelial cells to exert physiological effects. Although adhesion is not a prerequisite for the strains to possess probiotic properties, the interaction of probiotics with the intestinal mucosa is considered to be of critical importance for a number of reasons. Binding to the intestinal mucosa may prolong the time that the probiotic bacterial strain is present in the intestine. This interaction with the mucosa brings the probiotic bacteria into close contact with the intestinal immune system, giving it a better chance to modulate the immune response. It can also protect against enteric pathogens by limiting its ability to reside in the intestine.

(2) The acid-resistant and bile salt-resistant composite material has the following characteristics: under the environment of pH3.0, the survival of the strain is not obviously affected after 2h of incubation, and the viable count is reduced by 0.9 order of magnitude after 3 h; the pepsin has no obvious influence on the viability of the bacillus within 2 hours, and the viable count is reduced by 0.7 orders of magnitude within 3 hours; 3.6 orders of magnitude reduction in 0.5% bile salt system over 3 h. The food has good acid resistance and bile salt resistance, so that the food can better pass through the stomach and duodenum and smoothly reach the intestinal tract, thereby better playing the probiotic role; the good acid resistance enables the yogurt to adapt to a wider pH range in the fermentation process of preparing yogurt.

(3) In vitro cell test, it can promote proliferation of splenic lymphocyte in vitro, and the fold increase of no inducer is 1.912 + -0.087, and the fold increase of inducer is 1.533 + -0.109. Can promote spleen lymphocytes in vitro to secrete interleukin IL-12; the IL-12/IL-10 ratio was not significantly different from that of Lactobacillus rhamnosus LGG. In an in vitro cell test, the interleukin IL-8 secretion of the human colon cancer epithelial cells HT-29 can be promoted; IL-8, which stimulates secretion, is significantly higher than that of the commercial strain LGG.

(4) The compound preparation has good inhibition effect on staphylococcus aureus, salmonella paratyphi B, escherichia coli, shigella flexneri and micrococcus luteus, so that the compound preparation can inhibit pathogenic bacteria in intestinal tracts well, regulate intestinal flora and play a probiotic role.

(5) The living and non-living body have immunity enhancing effect, and the administration concentration of the living strain is 2 × 10⁸CFU/d, concentration of fire extinguishing strain taking 2 × 10⁸CFU/d can obviously improve the organ index of mice, promote the proliferation of splenic lymphocytes and improve the activity of NK cells. This can be doneThe probiotics in the yoghourt can exist in a non-living form, so that the yoghourt can have a longer shelf life at normal temperature without refrigeration.

Preferably, the yoghourt also comprises sugar alcohol, starch, a stabilizer and a sweetener.

Preferably, the yoghourt comprises 40-85% of raw milk or 5-15% of milk powder, 0.001-0.020% of lactobacillus helveticus and/or mutants thereof, 0.01-10.00% of components with the function of improving immunity except lactobacillus helveticus and/or mutants thereof, 0.5-5.0% of sugar alcohol, 0.1-5.0% of starch, 0.01-1.00% of stabilizing agent, 0.001-0.050% of sweetening agent and the balance of water in percentage by mass; the sum of all the raw materials is 100 percent.

The mass percentage is the percentage of the total mass of the yoghourt.

Preferably, the milk powder is one or more of whole milk powder, skim milk powder, whey protein powder and whole milk protein powder.

Preferably, the functional components for improving immunity further comprise one or more of propolis, ganoderma lucidum extract, wall-broken ganoderma lucidum spore powder, beta-glucan, dendrobium officinale extract, elderberry and acerola cherry.

Preferably, the sugar alcohol is one or more of xylitol, maltitol, erythritol, sorbitol, isomalt and lactitol.

Preferably, the starch is one or more of physical starch, acetate starch, sodium starch octenyl succinate, hydroxypropyl distarch phosphate, phosphate distarch, acid-treated starch, oxidized hydroxypropyl starch, acetylated distarch phosphate and acetylated distarch adipate.

Preferably, the stabilizer is one or more of sodium carboxymethylcellulose, pectin, soybean polysaccharide, agar, sodium caseinate, propylene glycol alginate, gelatin, gellan gum, guar gum and xanthan gum.

Preferably, the sweetener is one or more of acesulfame potassium, aspartame, sucralose, stevioside, mogroside, isomaltulose, edmuntin and neotame.

A method for preparing the sucrose-free yoghourt with the normal temperature and the long shelf life and the function of improving the immunity comprises the following steps:

scheme A:

(1) homogenizing and sterilizing: preheating raw milk, homogenizing, sterilizing, and cooling to obtain sterilized cooling feed liquid;

(2) inoculating and fermenting: adding lactobacillus helveticus and/or a mutant thereof into the sterilized cooling liquid obtained in the step (1), uniformly stirring, standing at a constant temperature, and fermenting to obtain a yogurt base material;

(3) preparing glue solution: preheating water, adding a formula amount of stabilizer into the water, shearing or stirring, and cooling to obtain a glue solution;

(4) secondary burdening: cooling the yogurt base material obtained in the step (2), and then stirring and demulsifying; adding the glue solution obtained in the step (3), adding the components with the formula dosage, sugar alcohol, sweetening agent and starch for improving the immunity except the lactobacillus helveticus and/or the mutant thereof, mixing and stirring, and adding water to a constant volume to obtain a feed liquid;

(5) and (3) secondary homogenization: homogenizing the feed liquid obtained in the step (4);

(6) filling and sterilizing: filling, sealing, pasteurizing and cooling the homogenized material liquid in the step (5) to obtain the normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity; or (5) carrying out UHT sterilization on the homogenized material liquid, carrying out hot filling or aseptic cold filling, and sealing to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature;

scheme B:

1) preparing milk: heating water, adding the formula amount of milk powder into the water, stirring or shearing, and standing for hydration to obtain milk;

2) homogenizing and sterilizing: preheating the milk obtained in the step 1), homogenizing, sterilizing and cooling to obtain a sterilized cooling feed liquid;

3) inoculating and fermenting: adding lactobacillus helveticus and/or a mutant thereof into the sterilized cooling liquid obtained in the step 2), uniformly stirring, standing at a constant temperature, and fermenting to obtain a yogurt base material;

4) preparing glue solution: preheating water, adding a formula amount of stabilizer into the water, shearing or stirring, and cooling to obtain a glue solution;

5) secondary burdening: cooling the yogurt base material obtained in the step 3), and then stirring and demulsifying. Adding the glue solution obtained in the step 4), adding the components with the formula dosage, sugar alcohol, sweetener and starch for improving the immunity except the lactobacillus helveticus and/or the mutant thereof, mixing and stirring, and adding water to a constant volume to obtain a feed liquid;

6) and (3) secondary homogenization: homogenizing the feed liquid obtained in the step 5);

7) filling and sterilizing: filling, sealing, pasteurizing and cooling the homogenized material liquid obtained in the step 6) to obtain the normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity; or carrying out UHT sterilization on the homogenized material liquid in the step 6), carrying out hot filling or aseptic cold filling, and sealing to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature.

The yoghourt prepared by pretreating, homogenizing, sterilizing, fermenting, re-sterilizing and filling the raw milk or milk powder and various raw and auxiliary materials can be normally stored at normal temperature without refrigeration, and the quality guarantee period can reach 150-180 days; meanwhile, the functional components are furthest kept from being damaged in the processing process, so that the functionality of the product is ensured.

Preferably, in step 1), the mass of the water is 200-400 g.

Preferably, in step 1), the water is heated to a temperature of 45-60 ℃.

Preferably, in step 1), the stirring or shearing time is 15 to 30 min.

Preferably, in the step 1), the standing hydration time is 20-40 min.

Preferably, in step (1) or step 2), the preheating is carried out to a temperature of 60 to 65 ℃.

Preferably, in the step (1) or the step 2), the homogenization pressure is 0 to 8/5 to 30 MPa.

Preferably, in the step (1) or the step 2), the sterilization temperature is 90-97 ℃ and the time is 3-10 min.

Preferably, in step (1) or step 2), the cooling is carried out to a temperature of 35 to 45 ℃.

Preferably, in the step (2) or the step 3), the fermentation temperature is 35-45 ℃ and the fermentation time is 4-16 h.

Preferably, in the step (2) or the step 3), the number of viable bacteria is more than or equal to 1 × 10 during the fermentation process⁸cfu/mL。

Preferably, in the step (3) or the step 4), the mass of the water is 200-400 g.

Preferably, in step (3) or step 4), the water is preheated to a temperature of 50-60 ℃.

Preferably, in step (3) or step 4), the shearing or stirring time is 20-40 min.

Preferably, in step (3) or step 4), the cooling is carried out to a temperature of 30 ℃ or lower.

Preferably, in step (4) or step 5), the cooling is carried out to a temperature of 15 ℃ or lower.

Preferably, in step (4) or step 5), the volume is set to 1000mL with water.

Preferably, in the step (5) or 6), the homogenization pressure is 0 to 8/5 to 30 MPa.

Compared with the prior art, the invention has the following advantages:

(1) the yoghourt is prepared by fermenting cow milk or reconstituted milk with lactobacillus helveticus WHH2580 and/or mutants thereof, is compounded with various natural functional components for improving the immunity, and can improve the immunity of a human body after being eaten for a long time;

(2) the lactobacillus helveticus WHH-2580 used in the formula of the yoghurt disclosed by the invention is a probiotic strain with an immunity-enhancing function, and is characterized in that the lactobacillus helveticus WHH-2580 still has an immunity-enhancing function in an inactivated state, so that the yoghurt disclosed by the invention can be pasteurized or UHT sterilized and has the characteristic of being preserved at normal temperature;

(3) the yoghourt is prepared by pretreating, homogenizing, sterilizing, fermenting, sterilizing and filling raw milk or milk powder and various raw and auxiliary materials, can be normally stored at normal temperature without refrigeration, and has a shelf life of up to 150-180 days; meanwhile, the functional components are furthest kept from being damaged in the processing process, so that the functionality of the product is ensured;

(4) the yoghourt disclosed by the invention is free of sucrose, can be drunk by various people in daily life, particularly people (diabetes, obesity and the like) with low autoimmunity and limited sugar intake and people who do not like cold food, has the advantages of high acceptability, high safety and no side effect, and has important efficacy value and social value.

Drawings

FIG. 1 is a flow chart of a method for preparing the sucrose-free yogurt with long shelf life at normal temperature for enhancing immunity according to the present invention;

FIG. 2 is a schematic diagram of a prior art process for preparing a yogurt with enhanced immunity;

FIG. 3 is a simplified diagram of a process for preparing the sucrose-free yogurt with long shelf life at room temperature for enhancing immunity according to the present invention;

FIG. 4 is a graph of thymus index of each group of mice in example 4;

FIG. 5 is a graph showing the results of experiments on spleen lymphocyte transformation in each group of mice in example 4;

FIG. 6 is a graph showing NK cell activities of various groups of mice in example 4;

FIG. 7 shows the morphology of Lactobacillus helveticus WHH2580 observed under a microscope;

FIG. 8 is a graph showing the effect of cell adhesion between viable, nonviable and commercial strains of Lactobacillus helveticus WHH2580, LGG and Lcs, and HT-29;

FIG. 9 is a graph showing the body weight change of BALB/c mice in each group in the animal test of example 15;

FIG. 10 is a graph showing organ ratios of BALB/c mice in each group in the animal test of example 15;

FIG. 11 is a graph showing spleen lymphocyte transformation numbers of groups of BALB/c mice in the animal test of example 15;

FIG. 12 is a graph showing NK cell activities of groups of BALB/c mice in example 15.

Detailed Description

The present invention will be further described with reference to the following examples.

General examples

A normal temperature long-shelf-life sucrose-free yoghourt for improving immunity comprises, by mass, 40% -85% of raw milk or 5-15% of milk powder, 0.001-0.020% of Lactobacillus helveticus and/or mutants thereof, 0.01-10.00% of components for improving immunity except the Lactobacillus helveticus and/or mutants thereof, 0.5-5.0% of sugar alcohol, 0.1-5.0% of starch, 0.01-1.00% of stabilizer, 0.001-0.050% of sweetener and the balance of water; the sum of all the raw materials is 100 percent.

The normal-temperature long-shelf-life sucrose-free yoghourt capable of improving the immunity is prepared through the following steps (the preparation process is shown as figure 1, and the schematic diagram is shown as figure 3):

scheme A:

(1) homogenizing and sterilizing: preheating raw milk to 60-65 ℃, and homogenizing under the homogenizing pressure of 0-8/5-30 MPa; sterilizing at 90-97 deg.C for 3-10min, and cooling to 35-45 deg.C to obtain sterilized cooling liquid;

the sterilization is the same as the conventional sterilization conditions in the field, such as pasteurization treatment;

(2) inoculating and fermenting, namely adding the Lactobacillus helveticus and/or the mutant thereof into the sterilized cooling feed liquid obtained in the step (1), uniformly stirring, placing in a constant-temperature incubator at 35-45 ℃, standing and fermenting for 4-16h, wherein the viable count is not less than 1 × 10⁸cfu/mL to obtain the yoghourt base material;

(3) preparing glue solution: weighing 200g-400g of ultrapure water, preheating to 50-60 ℃, adding the stabilizer with the formula amount into the ultrapure water, shearing or stirring for 20-40min, and cooling to below 30 ℃ to obtain glue solution;

(4) secondary burdening: cooling the yogurt base material obtained in the step (2) to below 15 ℃, and then stirring for demulsification; addition step

(3) Adding the components with the immunity improving effect except the lactobacillus helveticus and/or the mutant thereof, sugar alcohol, a sweetening agent and starch into the glue solution obtained in the step (1), mixing and stirring, and fixing the volume to 1000mL by using ultrapure water to obtain a feed liquid;

(5) and (3) secondary homogenization: homogenizing the feed liquid obtained in the step (4) at the pressure of 0-8/5-30 MPa;

(6) filling and sterilizing: filling the homogenized material liquid obtained in the step (5) into an HDPE material packaging container, sealing, and performing pasteurization and cooling to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature; or (5) carrying out UHT sterilization on the homogenized material liquid, carrying out hot filling or aseptic cold filling into a PET material packaging container, and sealing to obtain the normal-temperature sucrose-free yoghourt with long shelf life and improved immunity;

the pasteurization and UHT sterilization are the same as conventional sterilization conditions in the art.

Scheme B:

1) preparing milk: weighing 200 and 400g of ultrapure water, heating to 45-60 ℃, adding the milk powder with the formula amount into the ultrapure water, stirring or shearing for 15-30min, and standing for hydration for 20-40 min;

2) homogenizing and sterilizing: preheating the milk obtained in the step 1) to 60-65 ℃, and then homogenizing under the homogenizing pressure of 0-8/5-30 MPa; sterilizing at 90-97 deg.C for 3-10min, and cooling to 35-45 deg.C to obtain sterilized cooling liquid;

the sterilization is the same as the conventional sterilization conditions in the field, such as pasteurization treatment;

3) inoculating and fermenting, namely adding the Lactobacillus helveticus and/or the mutant thereof into the sterilized cooling feed liquid obtained in the step 2), uniformly stirring, placing in a constant-temperature incubator at 35-45 ℃, standing and fermenting for 4-16h, wherein the viable count is not less than 1 × 10⁸cfu/mL to obtain the yoghourt base material;

4) preparing glue solution: weighing 200g-400g of ultrapure water, preheating to 50-60 ℃, adding the stabilizer with the formula amount into the ultrapure water, shearing or stirring for 20-40min, and cooling to below 30 ℃ to obtain glue solution;

5) secondary burdening: cooling the yogurt base material obtained in the step 3) to below 15 ℃, and then stirring for demulsification; adding the glue solution obtained in the step 4), adding the components with the formula dosage, sugar alcohol, sweetener and starch for improving the immunity except the lactobacillus helveticus and/or the mutant thereof, mixing and stirring, and fixing the volume to 1000mL by using ultrapure water to obtain a feed liquid;

6) and (3) secondary homogenization: homogenizing the feed liquid obtained in the step 5), wherein the homogenizing pressure is 0-8/5-30 MPa;

7) filling and sterilizing: filling the homogenized material liquid obtained in the step 6) into an HDPE material packaging container, sealing, and performing pasteurization and cooling to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature; or carrying out UHT sterilization on the homogenized material liquid in the step 6), then carrying out hot filling or aseptic cold filling on the material liquid into a PET material packaging container, and sealing the container to obtain the sucrose-free yoghourt with the immunity improving function and the long shelf life at normal temperature;

the pasteurization and UHT sterilization are the same as conventional sterilization conditions in the art.