CN113767983A

CN113767983A - Human milk freeze-dried powder composition containing infant probiotic bacteria, and preparation method and application thereof

Info

Publication number: CN113767983A
Application number: CN202111064163.2A
Authority: CN
Inventors: 刘鹏; 张建瑞; 周英杰; 张勇
Original assignee: Hebei Reproductive Hospital
Current assignee: Hebei Reproductive Hospital
Priority date: 2021-09-10
Filing date: 2021-09-10
Publication date: 2021-12-10

Abstract

The invention relates to the technical field of food, in particular to a human milk freeze-dried powder composition containing infant-cadastral probiotics, a preparation method and application thereof. The three raw materials are matched and cooperated with each other, and the prepared composition can effectively promote the colonization of intestinal flora, inhibit the invasion of harmful bacteria, improve the intestinal microenvironment health of organisms and improve the immunity of the organisms.

Description

Human milk freeze-dried powder composition containing infant probiotic bacteria, and preparation method and application thereof

Technical Field

The invention relates to the technical field of food, in particular to a human milk freeze-dried powder composition containing infant probiotics, and a preparation method and application thereof.

Background

The intestinal flora is an important component of a human intestinal microecosystem, can synthesize various vitamins necessary for a human body, can participate in the metabolism of saccharides and proteins, promotes the absorption of the human body to various mineral elements, and plays a vital role in keeping the health of the human body.

The flora in the human intestinal tract is established at the beginning of life, and does not develop to be relatively stable until the age of 4 years or more, the intestinal tract flora is a key stage for customizing and establishing the intestinal tract flora in the human intestinal tract, a solid foundation can be provided for establishing the intestinal tract flora stable in the adult stage, breast milk is one of the main sources of the microbial flora in the intestinal tract of the infant and is also an important way for promoting the establishment of the intestinal tract flora and the development of immune functions of the infant, and the human milk is reported in documents, contains breast milk polysaccharide which is called Human Milk Oligosaccharides (HMOs) in fresh breast milk, has a protective effect on the infant, plays a prebiotic role and can selectively stimulate the growth of bifidobacteria and lactobacilli. However, under the influence of various objective factors, it is difficult to keep breast feeding during the establishment of intestinal flora, especially after 3 years of age, infants frequently contact with the external environment, and the change of diet and life style makes the state of intestinal flora extremely unstable during the process, which is not favorable for the normal establishment of intestinal flora.

The human intestinal flora is not invariable after being established, is closely related to the physical state, the life style and the living habit of the human body, and various adverse factors such as irregular daily work and rest, unhealthy diet, severe living environment and the like can cause the disorder of the human intestinal flora, cause different intestinal diseases and seriously endanger the life safety of people.

Disclosure of Invention

Aiming at the technical problems that the establishment of human intestinal flora is greatly influenced by external factors at the initial stage, is easily influenced by invasion of harmful bacteria, is slow to establish and has great side effect on intestinal flora in an antibiotic regulation disordered state in the prior art, the invention provides the preparation method of the human milk freeze-dried powder composition containing infant probiotic.

In order to achieve the purpose of the invention, the embodiment of the invention adopts the following technical scheme:

firstly, the embodiment of the invention provides a preparation method of a human milk lyophilized powder composition containing infant probiotic bacteria, which comprises the following steps: the preparation method takes human milk freeze-dried powder, prebiotics and infant probiotic powder as preparation raw materials, and specifically comprises the following steps:

s1: taking probiotic strains separated and purified from infant intestinal tracts, activating and culturing for 3 generations, inoculating the probiotic strains into an aseptic liquid culture medium, fermenting and culturing, centrifuging to obtain infant probiotic mud, mixing and emulsifying the obtained infant probiotic mud and a bacterial powder protective agent, freeze-drying at low temperature to remove water, crushing and sieving to obtain infant probiotic powder;

s2: taking healthy breast milk of a breast mother, carrying out pasteurization, then carrying out low-temperature freeze drying, removing water, crushing and sieving to obtain human milk freeze-dried powder;

s3: and (3) uniformly mixing the infant probiotic powder obtained in the step (S1), the human milk freeze-dried powder obtained in the step (S2) and prebiotics under an aseptic condition to obtain the human milk freeze-dried powder composition containing the infant probiotic.

Compared with the prior art, the preparation method of the freeze-dried human milk powder composition containing infant-centered probiotics disclosed by the application takes the freeze-dried human milk powder, prebiotics and probiotic powder from infant intestinal tracts as raw materials, and the probiotics in the infant intestinal tracts are more suitable for the intestinal environment of human bodies compared with probiotics from other sources, and by combining a specific preparation method, the activity and the state of the probiotics can be retained to the maximum extent, the stable colonization of the probiotics in the intestinal tracts of the human bodies is promoted, and the stable establishment of intestinal flora is facilitated; the freeze-dried human milk powder prepared by low-temperature freeze drying not only prolongs the shelf life of human milk, but also retains effective active substances in human milk to the maximum extent, and the restored freeze-dried human milk powder can provide easily absorbed nutrient substances for the growth of human bodies, further promotes the establishment of intestinal flora and maintains the stable state of the intestinal flora; the prebiotics also has an important effect on improving the gastrointestinal tract healthy microecology of organisms, can promote the permanent planting of beneficial bacteria in intestinal tracts, can also regulate and enrich the flora existing in the intestinal tracts, improve the gastrointestinal motility and the digestion and absorption capacity, improve the conditions of low peristaltic contraction amplitude of stomach tubes and esophagus and delayed gastric emptying, and reduce the occurrence of feeding intolerance.

The three raw materials are matched and cooperated with each other, and the prepared composition can effectively promote the colonization of intestinal flora, improve the intestinal microenvironment health of organisms and improve the immunity of the organisms.

The probiotic bacteria in the intestinal tract of the infant are derived from the oral cavity, the birth canal, the intestinal tract of the mother and the infant, the mother and infant flora transfer taking amniotic fluid as a passage before delivery, the vertical transfer of the birth canal flora in the delivery process, the transfer of microorganisms in the lactation process, the cross transfer of the mother and infant close contact flora and the like, and are more suitable for the intestinal tract environment of the infant and family members living together with the infant compared with other probiotic bacteria, so that the product prepared by the method is more suitable for the infant or the family members living together with the infant and can be applied to each stage of the infant or the family members after 3 weeks, the intestinal tract flora stability of the infant or the family members can be adjusted, and the health protection and driving of the infant and the family members can be realized.

In the research process of the inventor, the composition provided by the invention has an outstanding effect in regulating the colony state in the intestines and stomach of family members living together with the infant, and the symptoms of intestinal microbial disturbance are obviously improved after the family members take the human milk freeze-dried powder composition containing the infant live probiotics provided by the invention.

Preferably, the bacterial powder protective agent used in S1 is a mixture of trehalose, glycerol, skimmed milk powder, lactose and Tween-80.

Preferably, the viable count of the human milk lyophilized powder composition containing infant probiotic is more than or equal to 1 x 10⁸CFU/g。

1 x 10 is researched by the inventor⁸The CFU/g viable count content is the lowest concentration for improving the micro-environment health of the intestinal tract of an organism, improving the immunity of the organism, resisting the invasion of pathogenic bacteria, maintaining the shielding function of the intestinal tract and preventing the colonization of pathogenic bacteria under other conditions.

Preferably, the prebiotics comprise fructo-oligosaccharides and galacto-oligosaccharides, wherein the sum of the mass of the two is not less than 2/3 of the total mass of the prebiotics.

The fructo-oligosaccharide and the galacto-oligosaccharide are effective components for promoting the field planting of the bifidobacteria and the lactobacillus plantarum in the intestinal tract, can promote the absorption of the intestinal tract to nutrient components, improve the intestinal tract and promote the intestinal tract peristalsis, and have important effects on the establishment of intestinal flora and the improvement of the intestinal function.

Preferably, the low-temperature freeze drying process in the step S2 is performed in a freeze dryer, and specifically includes a pre-freezing process and a freeze-drying process, the whole low-temperature freeze drying process maintains the temperature of the hydrazine in the freeze dryer between-30 ℃ and-40 ℃, the vacuum degree is less than 30Pa, and the pre-freezing process lasts for 2.5 to 3.5 hours.

Compared with spray drying, the freeze drying has lower treatment temperature, can effectively avoid the effective components in the emulsion from being damaged due to high temperature, and the addition of the pre-freezing process can also enable the sample to be dried to be solidified into a solid state before freeze drying, thereby avoiding the loss caused by splashing of the liquid sample in the vacuum drying process and being beneficial to protecting the stable content of the effective components in the sample.

Preferably, the freeze-drying process in the freeze-drying machine is divided into 7 stages, wherein the duration of each stage in the 1 st to 6 th stages is 1.5 to 2.5 hours, the duration of the 7 th stage is 8 to 10 hours, and the temperature of the sample is controlled to be increased along with the change gradient of the freeze-drying stage in the range of-25 ℃ to 35 ℃ in the whole freeze-drying process.

The gradient rise of the sample temperature in a specific range can improve the water removal efficiency, shorten the freeze-drying time, remove the water in the sample to the maximum extent and protect the stable property of the active substances in the sample.

The invention also provides a human milk lyophilized powder composition containing infant probiotic, which is prepared by the preparation method of the human milk lyophilized powder composition containing infant probiotic, wherein the infant probiotic powder is a mixture of bacterial powders prepared from bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC 003.

Preferably, the preparation raw materials of the infant-scale probiotic-containing human milk lyophilized powder composition comprise the following components in percentage by mass: 2% -4% of infant probiotic powder, 28% -32% of human milk freeze-dried powder and 65% -70% of prebiotics; the mass ratio of the bacterial powder prepared from bifidobacterium bifidum HRHCC001, the bacterial powder prepared from bifidobacterium lactis HRHCC002 and the bacterial powder prepared from lactobacillus plantarum HRHCC003 in the probiotic bacterial powder of infant means is as follows: 0.8-1.2: 0.8-1.2: 1.6-2.4, the content of viable bacteria in the composition is more than or equal to 1 multiplied by 10⁸CFU/g. When the mass percentage, the viable count content and the mass ratio are in the preferable ranges, the obtained product has the most outstanding antibacterial performance and the optimal planting capability in intestinal tracts.

The Bifidobacterium bifidum HRHCC001 used in the invention is classified as Bifidobacterium bifidum (Bifidobacterium bifidum) and has been preserved in China general microbiological culture Collection center of China Committee for culture Collection, No.1 Hospital, No.3, located in the Shangyang area of Beijing, 6.4.2021, with the preservation number: CGMCC No. 22656;

bifidobacterium lactis HRHCC002 used in the present invention, which is classified as Bifidobacterium lactis (Bifidobacterium lactis), has been deposited in China general microbiological culture Collection center (CCTCC) No.3 of West Lu No.1 Hospital, North Cheng, the Yangxi, Beijing city, on 6.4 days of 2021, with the following deposition numbers: CGMCC No. 22657.

The Lactobacillus plantarum HRHCC003 used in the invention is classified as Lactobacillus plantarum (Lactobacillus plantarum), which has been preserved in the general microbiological culture Collection center of China microbiological culture Collection Committee No.3, West Lu No.1 Hospital, North Cheng, the Yangtze district, Beijing city, on 4 days 6 months 4 days 2021, and has the preservation number: CGMCC No. 22658.

Researches find that bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003 which are separated from intestinal tracts of healthy infants have mutual promotion and cooperative synergistic effect in the aspect of antibacterial performance, can promote the colonization of probiotics in the intestinal tracts and inhibit the invasion of harmful bacteria in the intestinal tracts, are more beneficial to the establishment of intestinal flora, maintain the stability of the intestinal flora and regulate the intestinal flora in a disordered state. The human milk lyophilized powder composition using the bifidobacterium bifidum HRHCC001, the bifidobacterium lactis HRHCC002 and the lactobacillus plantarum HRHCC003 as the infant probiotic raw materials can obviously improve the intestinal flora disorder state of family members of the infant and also has obvious improvement effect on the intestinal flora disorder state of other people.

Preferably, the composition can be prepared into dosage forms including powder, granules or tablets.

In a second aspect, the embodiment of the present invention further provides an application of the above-mentioned human milk lyophilized powder composition containing probiotics of infant formulae in the fields of infant drinks, foods and adult drinks and foods, and a baby drink, a food or an adult drink and a food for infants over 3 years old are prepared, wherein the baby food comprises a baby staple food and an auxiliary food.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

The intestinal flora in human bodies plays a vital role in keeping human health, the intestinal flora is established at the early stage of life and is developed to be relatively stable until the intestinal flora is over 4 years old, the key stage of intestinal flora customization and establishment is provided in the early stage of human intestinal flora establishment, namely the infancy stage, the intestinal flora establishment and the establishment can provide a solid foundation for the establishment of the intestinal flora stable in adulthood, breast milk is one of the main sources of the microbial flora in the intestinal tracts of infants and is an important way for promoting the establishment of the intestinal flora and the development of immune functions of the infants, and the breast milk contains breast milk polysaccharide which is called Human Milk Oligosaccharides (HMOs) and has a protective effect on the infants, plays a role of prebiotics and can selectively stimulate the growth of bifidobacteria and lactobacilli. However, under the influence of various objective factors, it is difficult to keep breast feeding during the establishment of intestinal flora in human body, especially after 3 years old, infants frequently contact with the external environment, and the state of intestinal flora is extremely unstable during the process due to the change of diet and life style, which is not beneficial to the normal establishment of intestinal flora. The human intestinal flora is not invariable after being established, is closely related to the body state, the life style and the life habit of the human body, and various adverse factors such as irregular daily work and rest, unhealthy diet, severe living environment and the like can cause the disorder of the human intestinal flora, cause different intestinal diseases and seriously endanger the life safety of people.

In order to obtain a composition capable of effectively promoting the stable establishment of intestinal flora, promoting the colonization of a native strain and regulating the stability of the intestinal flora, the application provides a preparation method of a human milk lyophilized powder composition containing infant probiotic bacteria, which comprises the following steps: the preparation method takes human milk freeze-dried powder, prebiotics and infant probiotic powder as raw materials, and specifically comprises the following steps:

s1: taking probiotic strains separated and purified from infant intestinal tracts, activating and culturing for 3 generations, inoculating the probiotic strains into an aseptic liquid culture medium, fermenting and culturing, centrifuging to obtain infant probiotic mud, mixing and emulsifying the infant probiotic mud and a bacterial powder protective agent, freeze-drying at low temperature to remove water, crushing and sieving to obtain infant probiotic powder;

s3: and (3) uniformly mixing the infant probiotic bacteria obtained in the step (S1), the human milk freeze-dried powder obtained in the step (S2) and prebiotics in an aseptic condition to obtain the human milk freeze-dried powder composition containing the infant probiotic bacteria. The human milk freeze-dried powder, the prebiotics and the probiotic powder from the intestinal tract of the infant are used as raw materials, the probiotics in the intestinal tract of the infant is more suitable for the intestinal tract environment of a human body compared with probiotics from other sources, and the activity and the state of the probiotics can be kept to the maximum extent by combining a specific preparation method, so that the stable planting of the probiotics in the intestinal tract of the human body is promoted, and the stable establishment of the intestinal flora is facilitated; the freeze-dried human milk powder prepared by low-temperature freeze drying not only prolongs the shelf life of human milk, but also retains effective active substances in human milk to the maximum extent, and the restored human milk freeze-dried powder can provide easily absorbed nutrient substances for the growth of human bodies, further promotes the establishment of intestinal flora and maintains the stable state of the intestinal flora; the prebiotics have an important function of improving the gastrointestinal tract healthy microecology of organisms, can promote the permanent planting of beneficial bacteria in intestinal tracts, can also regulate and enrich the flora existing in the intestinal tracts, improve the gastrointestinal motility and the digestion and absorption capacity, improve the conditions of low peristaltic contraction amplitude of stomach tubes and esophagus and delayed gastric emptying, and reduce the occurrence of feeding intolerance.

Exemplary of the fungal powder protective agent used in S1 is a mixture of trehalose, glycerol, skim milk powder, lactose and tween-80.

The inventor researches that the viable count of the human milk freeze-dried powder composition containing the infant probiotic is 1 multiplied by 10⁸CFU/g is the minimum content for improving the micro-environment health of the intestinal tract of the organism, improving the immunity of the organism, resisting the invasion of pathogenic bacteria, maintaining the shielding function of the intestinal tract and preventing the colonization of pathogenic bacteria under other conditions, so the viable count of the human milk freeze-dried powder composition containing the infant-like probiotics is more than or equal to 1 multiplied by 10⁸CFU/g。

Illustratively, the prebiotics used in the application comprise fructo-oligosaccharide and galacto-oligosaccharide, wherein the sum of the mass of the fructo-oligosaccharide and the galacto-oligosaccharide is not less than 2/3 of the total mass of the prebiotics, and the galacto-oligosaccharide and the fructo-oligosaccharide can be used as effective components for promoting the colonization of bifidobacteria and lactobacillus plantarum in the intestinal tract, and can also promote the absorption of the intestinal tract on nutrient components, improve the intestinal tract function, promote the intestinal tract peristalsis function and play an important role in the establishment of the intestinal flora and the improvement of the intestinal tract function.

In order to avoid splashing of a sample in the vacuum drying process, avoid loss of effective components and protect the stability of the functions and the content of the effective components to the maximum extent, the low-temperature freeze drying process in S2 is carried out in a freeze dryer, and specifically comprises a pre-freezing process and a drying process, wherein the time of the pre-freezing process is 2.5-3.5 h, the temperature of a hydrazine in the freeze dryer is maintained between-30 ℃ and-40 ℃ in the whole low-temperature freeze drying process, and the vacuum degree is less than 30 Pa.

In order to ensure the removal efficiency of water in a sample to be dried, shorten the freeze-drying time and ensure the activity of effective components, the freeze-drying process carried out in a freeze-drying machine is divided into 7 stages, wherein the duration time of each stage of the 1 st to 6 th stages is 1.5h to 2.5h, the duration time of the 7 th stage is 8 to 10h, and the temperature of the sample is controlled to be increased along with the change gradient of the freeze-drying stage in the range of-25 ℃ to 35 ℃ in the whole drying process.

The invention also provides a human milk lyophilized powder composition containing infant probiotic, which is prepared by adopting the preparation method of the human milk lyophilized powder composition containing infant probiotic, wherein the infant probiotic powder is a mixture of bacterial powders prepared from bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC 003. The baby milk freeze-dried powder composition containing the infant probiotics comprises the following preparation raw materials in percentage by mass: 2% -4% of infant probiotic powder, 28% -32% of human milk freeze-dried powder and 65% -70% of prebiotics; wherein the viable count content in the composition is not less than 1 × 10⁸CFU/g, the mass ratio of the bacterial powder prepared from bifidobacterium bifidum HRHCC001, the bacterial powder prepared from bifidobacterium lactis HRHCC002 and the bacterial powder prepared from lactobacillus plantarum HRHCC003 in the probiotic bacterial powder of the infant is as follows: 0.8-1.2: 0.8-1.2: 1.6-2.4.

the specific screening method comprises the following steps:

a1: taking fresh feces of 18-month-old healthy infants, diluting with sterile normal saline by multiple times to obtain feces 10^-4，10^-5Diluting liquid:

a2: respectively coating the diluent in an MRS solid culture medium and an MRS solid culture medium containing mupirocin lithium salt for anaerobic culture for 48-72h, respectively selecting single colonies after culture, selecting the single colonies in the MRS liquid culture medium, carrying out anaerobic culture for 18-24h at 37 ℃, after the culture medium is turbid, streaking in the same culture medium for the first time, carrying out anaerobic culture for 48-72h, and repeating for at least 3 times until colonies with uniform thallus morphology are obtained;

a3: extracting and separating the genome of the strain by using a genome extraction kit, taking the genome DNA as a template, amplifying 16s rDNA (the sequence is shown as SEQ ID NO. 1) by using bacterial universal primers 27F and 1492R, detecting a band of a PCR product at 1.5KB by electrophoresis, sequencing the PCR product, uploading the obtained sequence to NCBI, comparing the sequence with the BLAST, determining that the strain belongs to the Bifidobacterium bifidum if the similarity of the result and the Bifidobacterium bifidum is higher and is more than 97 percent, naming the strain as the Bifidobacterium bifidum HRHCC001, storing the strain in a vertical bacterial cryopreservation tube, and sending the strain to the China microorganism culture collection management committee general microorganism center of No.1 Hospital No.3 of Beijing rising Kongyu West No.1, and storing the strain with the following preservation numbers: CGMCC No. 22656.

The specific screening method comprises the following steps:

b1: taking fresh feces of Bifidobacterium bifidum HRHCC001 from the same source as the feces of the infant for 24 months, diluting with sterile normal saline by multiple times to obtain feces 10^-4，10^-5Diluting liquid:

b2: respectively coating the diluent in an MRS solid culture medium and an MRS solid culture medium containing mupirocin lithium salt for anaerobic culture for 48-72h, respectively selecting single colonies after culture, selecting the single colonies in the MRS liquid culture medium, carrying out anaerobic culture for 18-24h at 37 ℃, after the culture medium is turbid, streaking in the same culture medium for the first time, carrying out anaerobic culture for 48-72h, and repeating for at least 3 times until colonies with uniform thallus morphology are obtained;

b3: extracting and separating the genome of the strain by using a genome extraction kit, taking the genome DNA as a template, amplifying 16s rDNA (the sequence is shown as SEQ ID NO. 2) by using bacterial universal primers 27F and 1492R, detecting a band at 1.5KB of a PCR product by electrophoresis, sequencing the PCR product, uploading the obtained sequence to NCBI for BLAST comparison, determining that the strain belongs to the Bifidobacterium lactis if the similarity of the result and the Bifidobacterium lactis is higher and is more than 97 percent, storing the strain in a vertical bacterial cryopreservation tube, sending the strain to a China general microbiological culture collection center of China Committee for culture management of microorganisms, namely NO.1 and NO.3 of the west Lou of the sunward region in Beijing, and storing the strain with the following preservation number: CGMCC No. 22657.

The specific screening method comprises the following steps:

c1: taking fresh feces of Bifidobacterium bifidum HRHCC001 from the same source as the feces of the infant for 30 months, diluting with sterile normal saline by multiple times to obtain feces 10^-4，10^-5Diluting liquid:

c2: respectively coating the diluent in an MRS solid culture medium and an MRS solid culture medium containing mupirocin lithium salt for anaerobic culture for 48-72h, respectively selecting single colonies after culture, selecting the single colonies in the MRS liquid culture medium, carrying out anaerobic culture for 18-24h at 37 ℃, after the culture medium is turbid, streaking in the same culture medium for the first time, carrying out anaerobic culture for 48-72h, and repeating for at least 3 times until colonies with uniform thallus morphology are obtained;

c3: extracting and separating the genome of the strain by using a genome extraction kit, taking the genome DNA as a template, amplifying 16s rDNA (the sequence is shown as SEQ ID NO. 3) by using bacterial universal primers 27F and 1492R, then carrying out electrophoresis detection on the PCR product to detect that a band exists at the position of 1.5KB, sequencing the PCR product, uploading the obtained sequence to NCBI for BLAST comparison, and if the similarity of the result and the lactobacillus plantarum is higher and is more than 97 percent, determining that the strain belongs to the lactobacillus plantarum, namely the lactobacillus plantarum HRHCC003, storing the strain in a vertical bacterial cryopreservation tube, and sending the strain to a common microorganism center of China culture and microorganism culture collection Committee No.1 Hospital No.3 of North West Cheng of the Beijing area in Beijing for preservation, wherein the preservation numbers are as follows: CGMCC No. 22658.

According to the research, bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003 which are separated from the intestinal tract of a healthy infant have mutual promotion and cooperative synergistic effect in the aspect of antibacterial performance, can inhibit the invasion of harmful bacteria in the intestinal tract while promoting the colonization of probiotics in the intestinal tract, are more beneficial to the establishment of intestinal flora, maintain the stability of the intestinal flora and regulate the intestinal flora in a disordered state. The human milk lyophilized powder composition using the bifidobacterium bifidum HRHCC001, the bifidobacterium lactis HRHCC002 and the lactobacillus plantarum HRHCC003 as the infant probiotic raw materials can obviously improve the intestinal flora disorder state of family members of the infant and also has obvious improvement effect on the intestinal flora disorder state of other people.

The invention is further illustrated below in the following examples.

Example 1

The embodiment provides a lyophilized human milk powder composition containing infant probiotic, which is prepared from lyophilized human milk powder, prebiotics, and bacterial powder prepared from bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003 as raw materials, and the preparation method of the composition specifically comprises the following steps:

s1: respectively taking bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003, activating strains, continuously culturing for 3 generations, respectively inoculating the strains into 120L of sterilized liquid culture medium, fermenting at 37 ℃, monitoring pH value, OD value and alkali consumption, centrifugally harvesting after a harvest point, respectively obtaining bacterial sludge of corresponding probiotics, and mixing the bacterial sludge with bacterial powder protective agent consisting of trehalose, glycerol, skim milk powder, lactose and tween-80 according to the ratio of 1: 1.7, placing the mixture in a precooled freeze dryer for freeze drying, and collecting freeze-dried substances to obtain corresponding bacterium powder of the three probiotics;

s2: collecting redundant milk of healthy lactating mothers, pasteurizing in an environment of 65 ℃ for 30min after detecting the nutrient content condition by a breast milk analyzer, transferring the pasteurized human milk to a freeze drying tray for freeze drying in a freeze dryer, wherein the specific freeze drying procedure is as follows:

pre-freezing: the temperature of the cold hydrazine is-35 ℃ for 3 hours;

first-stage freeze drying: the temperature of the cold hydrazine is-35 ℃, the temperature of the sample is-25 ℃, the temperature is 2 hours, and the vacuum degree is less than 30 Pa;

second-stage freeze drying: the temperature of the cold hydrazine is-35 ℃, the temperature of the sample is-15 ℃, the temperature is 2 hours, and the vacuum degree is less than 30 Pa;

three-stage freeze-drying: the temperature of the cold hydrazine is-35 ℃, the temperature of the sample is-5 ℃, the temperature is 2 hours, and the vacuum degree is less than 30 Pa;

fourth-stage freeze-drying: the temperature of the cold hydrazine is-35 ℃, the temperature of the sample is 5 ℃, the time is 2 hours, and the vacuum degree is less than 30 Pa;

and (5) five-stage freeze-drying: the temperature of the cold hydrazine is-35 ℃, the temperature of the sample is 15 ℃, the time is 2 hours, and the vacuum degree is less than 30 Pa;

six-stage freeze-drying: the temperature of the cold hydrazine is-35 ℃, the temperature of the sample is 25 ℃, the time is 2 hours, and the vacuum degree is less than 30 Pa;

seven-stage freeze-drying: and (3) cooling hydrazine at the temperature of-35 ℃, sample temperature of 35 ℃, 9 hours, and vacuum degree of less than 30Pa, closing the freeze dryer after the freeze-drying process is finished, and crushing and sieving the residual substances to obtain the human milk freeze-dried powder.

S3: collecting bacterial powder of Bifidobacterium bifidum HRHCC001 0.5kg, bacterial powder of Bifidobacterium lactis HRHCC002 0.5kg, bacterial powder of Lactobacillus plantarum HRHCC003 1.0kg, and bacterial powder of Lactobacillus plantarum HRHCC003 30kg human milk lyophilized powder, 34kg fructo-oligosaccharide, 34kg galacto-oligosaccharide, stirring in an aseptic mixing tank for 10min, mixing well to obtain the human milk lyophilized powder mixture containing infant probiotic, and detecting the content of probiotic in the composition to 5 × 10⁸cfu/g, and hermetically packaging the obtained mixture for later use.

Example 2

This example provides 3 probiotics: bacteriostasis experiments of bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC 003;

(1): respectively mixing bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003 to obtain 4 groups of samples, wherein the 1 st group to the 3 rd group of samples are single strains of bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003 respectively, the 4 th group of samples are mixed bacteria of bifidobacterium bifidum HRHCC001+ bifidobacterium lactis HRHCC002+ lactobacillus plantarum HRHCC003, respectively inoculating the 4 groups of mixed bacteria powder into 100ml of MRS liquid culture medium, and performing static culture at 37 ℃ for 48 hours to obtain corresponding probiotic cultures;

(2): respectively adding activated Escherichia coli, salmonella and Staphylococcus aureus into 5 wt% tryptose soy broth to obtain a mixture with a concentration of 1 × 10⁶Respectively coating the prepared escherichia coli culture solution, salmonella culture solution and staphylococcus aureus culture solution in culture dishes filled with tryptic soy broth culture medium by using CFU/ml bacterial culture solution, wherein 4 groups of culture dishes are prepared for each bacterial culture solution; corresponding to the 4 probiotic cultures prepared in the step (1);

(3): the 4 groups of probiotic cultures obtained in the step (1) were respectively dropped on a circular filter paper sheet by using a capillary, and symmetrically attached to the central area of the culture dish obtained in the step (2), and incubated in an incubator at 37 ℃ for 24 hours, and the diameter of the antibacterial area in each culture dish was measured, and the results are shown in table 1:

TABLE 1 diameter of zone of inhibition obtained from probiotic bacteria inhibition experiments of different compositions

From the data in table 1, it can be seen that: the diameters of the inhibition zones of the mixed bacteria in the group 4 in the experiments of inhibiting escherichia coli, salmonella and staphylococcus aureus are all larger than the diameters of the single bacteria in the groups 1-3, which shows that the bifidobacterium bifidum HRHCC001, the bifidobacterium lactis HRHCC002 and the lactobacillus plantarum HRHCC003 can be mutually matched and promoted in the inhibition aspect, and the antibacterial performance is jointly improved, so that the invasion of harmful bacteria in intestinal tracts is effectively inhibited, the normal establishment of intestinal flora is promoted, and the health state of the intestinal flora is maintained.

Example 3

This example was carried out to test the reconstitution of the lyophilized human milk powder obtained in example 1

The raw materials of the human milk lyophilized powder prepared in step S2 in example 1 were dissolved in water according to 80%, 100%, and 120% of the original volume, the effective components in the reconstituted milk were detected by a breast milk analyzer, and the lyophilized condition of the human milk lyophilized powder was judged by comparing the content of the effective components in the original human milk, and the results are shown in table 2:

TABLE 2 distribution of main constituents in human milk and lyophilized powder reconstituted milk

Item	Raw milk	80% recovery	100% recovery	120% recovery	Unit of
						Fat	3.84	4.91	3.71	2.9	g/100g
Protein	0.94	1.13	0.91	0.67	g/100g
						Lactose	7.11	8.18	7.26	5.83	g/100g
(Energy)	63.52	79.9	61.41	51.53	kcal/100g
						Mineral substance	0.19	0.25	0.16	0.12	g/100g
Density of	88.28	109.16	87.96	74.07	g/100g
						Zinc	1.56	1.63	1.48	1.11	μg/ml
Iron	1.42	1.82	1.55	1.29	μg/ml
						Calcium carbonate	214.5	254.21	209.6	175.52	μg/ml

As can be seen from the data in table 2, the human milk lyophilization process provided by the present application does not affect the main components in human milk, can remove the original components in human milk to the maximum extent, is more suitable as a component of a composition, effectively replaces human milk, and contributes significantly to the establishment of flora in infant intestinal tracts.

Comparative example 1

The comparative example provides a probiotic-containing human milk lyophilized powder composition (non-infant body), and compared with example 1, the preparation process parameters are consistent with example 1 except that bifidobacterium bifidum powder and lactobacillus plantarum powder are purchased from jacobia bioengineering limited company and bifidobacterium lactis powder is purchased from kayashi bioengineering limited company.

Comparative example 2

Compared with the embodiment 1, except that the human milk lyophilized powder is replaced by a mixture prepared by referring to the component proportion of the human milk, and other additives, the rest preparation process parameters are consistent with those of the embodiment 1, wherein the mixture composed of the human milk lyophilized powder and the additives comprises the following components in parts by mass: 40 parts of raw milk, 29 parts of desalted whey powder, 6 parts of white granulated sugar, 1.6 parts of 3' sialyllactose, 0.5 part of phospholipid, 0.16 part of DHA (docosahexaenoic acid) algal oil powder, 0.12 part of arachidonic acid powder, 0.15 part of sialic acid, 0.03 part of nucleotide, 0.35 part of vitamin complex and 0.16 part of compound mineral.

Example of effects:

taking the composition samples prepared in example 1, comparative example 1 and comparative example 2, and respectively taking the composition samples as infant food and adult drinks to test the colonization effect on infant intestinal flora and the regulation effect on adult disordered intestinal flora, the specific experimental scheme is as follows:

infant intestinal flora field planting experiment

Selecting 21 normally-developing children over 3 years old, wherein the specific selection standard is as follows: in the same area, the medicine is not taken in the same year and a month, has no acute and chronic diseases, has good health status, does not suffer from infectious diseases such as AIDS, tuberculosis, hepatitis B and the like, accords with the declaration of Helsinki, and is signed with the informed consent of parents.

In 21 infants, one infant is the infant body screened from bifidobacterium bifidum HRHCC001, bifidobacterium lactis HRHCC002 and lactobacillus plantarum HRHCC003, the infant body is taken as an experimental group 1, the other 20 infants are equally divided into 4 groups, the 4 groups are respectively taken as an experimental group 2-an experimental group 5, the compositions prepared in the example 1, the comparative example 1 and the comparative example 2 are taken, the 21 infants are respectively fed after the compositions are dissolved in water in equal amount, the specific feeding scheme is that the composition prepared in the example 1 is taken by the experimental group 1 and the experimental group 2, the composition prepared in the comparative example 1 is taken by the experimental group 3, the composition prepared in the comparative example 2 is taken by the experimental group 4, and the experimental group 5 is a blank control group which is not taken and is taken 2 times a day and 2g each time for two weeks. The contents of bifidobacterium and lactobacillus in the intestinal tracts of the infants of each experimental group are detected by using an RT-PCR detection method before taking, after taking for 2 weeks and stopping taking for 1 week, and the average value of the data obtained by each experimental group is shown in a table 4:

the specific method for detecting the content of the probiotics by using the RT-PCR detection method comprises the following steps:

and (3) carrying out PCR on the genome by using RT-PCR to detect the content of bifidobacterium and lactobacillus in the excrement of the infant. The method for detecting the bifidobacterium system comprises the following steps of 10 mu L of Tiangen SYBR green Mix (probe), 0.6 mu L of each primer, 2 mu L of template, 6.4 mu L of water, 0.4 mu L of probe and 2 mu L of template, and the reaction conditions are as follows: keeping at 50 deg.C for 120s, pre-denaturing at 95 deg.C for 15min, at 95 deg.C for 15s, at 60 deg.C for 1min, and performing 45 cycles.

The lactobacillus detection adopts a 30 microliter system, which comprises 10 microliter SYBR green Mix, 6.8 microliter water, 0.6 microliter primers respectively, and 2 microliter templates, and the reaction conditions are as follows: keeping at 50 ℃ for 120s, pre-denaturing at 95 ℃ for 900s, pre-denaturing at 95 ℃ for 15s, pre-denaturing at 60 ℃ for 30s, pre-denaturing at 72 ℃ for 45s, performing 45 cycles at a melting temperature of 65-95 ℃, and obtaining specific primers shown in Table 3

TABLE 3 Bifidobacterium and Lactobacillus detection primer sequences

TABLE 4

As can be seen from the comparison of the data of the experimental group 2 and the experimental group 5 in table 3, the composition prepared in the example 1 of the present invention has the outstanding ability of promoting the colonization of the intestinal flora; as can be seen from the comparison of the data of the experimental group 2, the experimental group 3 and the experimental group 4, the composition prepared in the embodiment 1 of the invention is superior to the composition prepared in the comparative example 1 and the comparative example 2 in the capability of promoting the colonization of intestinal flora, and as can be seen from the comparison of the data of the experimental group 1 and the experimental group 2, the composition prepared from the probiotics screened from the body has a superior effect of promoting the intestinal flora of infants to the composition prepared from the probiotics screened from non-body.

Experiment for regulating adult intestinal flora disorder

Selecting 21 patients with disordered intestinal flora, wherein the specific selection criteria are as follows: the patients with constipation and diarrhea caused by intestinal flora disorder in the same area with similar age do not have infectious diseases such as AIDS, tuberculosis, hepatitis B and the like, accord with the declaration of Helsinki, and sign informed consent.

One of the 21 patients was selected from parents (co-life) of infants screened for Bifidobacterium bifidum HRHCC001, Bifidobacterium lactis HRHCC002 and Lactobacillus plantarum HRHCC003, and was used as experimental group a, and the remaining 20 patients were equally divided into 4 groups, which were used as experimental group b-experimental group e, respectively. An equal amount of the composition prepared in example 1, comparative example 1 and comparative example 2 was dissolved in water and administered to 21 patients, wherein the composition prepared in example 1 was administered to experimental group a and experimental group b, the composition prepared in comparative example 1 was administered to experimental group c, the composition prepared in comparative example 2 was administered to experimental group d, and the blank control group of experimental group e was not administered and administered 3 times daily for two weeks. The average results of the data obtained from each experimental group are shown in table 5, wherein the contents of bifidobacteria and lactobacilli in intestinal tracts of patients in each experimental group are tested by using the same RT-PCR detection method as in the first experiment after the patients are taken for 2 weeks and are stopped for 1 week:

TABLE 5

As can be seen from the comparison of the data in the experimental group b and the experimental group e in Table 5, the composition prepared in example 1 of the present invention is effective in regulating the disordered intestinal flora system of adults; the comparison of the data of the experimental group b, the experimental group c and the experimental group d shows that the composition prepared in the embodiment 1 has obvious improvement effect on the regulation of the intestinal flora disorder and the maintenance of the intestinal flora health on family members living together and other people living non-together, and is superior to the composition prepared in the comparative example 1 and the comparative example 2, and the comparison of the data of the experimental group a and the experimental group b shows that the composition prepared from the probiotics screened in the body of the infant of the same family has better effect on regulating the intestinal flora disorder state than the composition prepared from the probiotics screened in the body of the infant of the non-same family.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.

SEQUENCE LISTING

<110> Hospital for reproduction in Hebei province

<120> human milk freeze-dried powder composition containing infant probiotic, preparation method and application

<130> 003

<160> 3

<170> PatentIn version 3.5

<210> 1

<211> 1419

<212> DNA

<213> Bifidobacterium bifidum (Bifidobacterium bifidum)

<400> 1

ccccatgggg ggcgtcttac catgcaagtc gaacgggatc catcaagctt gcttggtggt 60

gagagtggcg aacgggtgag taatgcgtgc cgacctgccc catgctccgg aatagctcct 120

ggaaacgggt ggtaatgcca tgttccacat gatcgcatgt gattgtggga aagatttcat 180

cggcgtggga tggggtcgcg tcctatcgct tgttggtgag gtaacggctc accaaggctt 240

cgacgggtag ccggcctgag agggcgccgg ccacattggg actgagatac ggcccagact 300

cctacgggag gcagcagtgg ggaatattgc acaatgggcg caagcctgat gcagcgacgc 360

cgcgtgaggg atggaggcct tcgggttgta aacctctttt gtttgggagc aagccttcgg 420

gtgagtgtac ctttcgaata agcgccggct aactacgtgc cagcagccgc ggtaatacgt 480

agggcgcaag cgttatccgg atttattggg cgtaagggct cgtaggcggc tcgtcgcgtc 540

cggtgtgaaa gtccatcgct taacggtgga tctgcgccgg gtacgggcgg gctggagtgc 600

ggtaggggag actgaattcc cggtgtaacg gtggaatgtg tagatatcgg gagaacaccg 660

atggcgaagg caggtctctg ggccgtcact gacgctgagg agcgaaagcg tggggagcga 720

acaggattag ataccctggt agtccacgcc gtaaacggtg gacgctggat gtggggcacg 780

ttccacgtgt tccgtgtcgg agctaacgcg ttaagcgtcc cgcctggggg agtacggccg 840

caaggctaaa actcaaagaa attgacgggg gcccgcacaa gcgcggagca tgcggattaa 900

ttcgatgcaa cgcgaagaac cttacctggg cttgacatgt tcccgacgac gccagagatg 960

gcgtttccct tcggggcggg ttcacaggtg gtgcatggtc gtcgtcagct cgtgtcgtga 1020

gatgttgggt taagtcccgc aacgagcgca accctcgccc cgtgttgcca gcacgttatg 1080

gtgggaactc acgggggacc gccggggtta actcggagga aggtggggat gacgtcagat 1140

catcatgccc cttacgtcca gggcttcacg catgctacaa tggccggtac agcgggatgc 1200

gacatggcga catggagcgg atccctgaaa accggtctca gttcggatcg gagcctgcaa 1260

cccggctccg tgaaggcgga gtcgctagta atcgcggatc agcaacgccg cggtgaatgc 1320

gttcccgggc cttgtacaca ccgcccgtca agtcatgaaa gtgggcagca cccgaagccg 1380

gtggcctaac cccttgtggg atggagccgt cctaagtga 1419

<210> 2

<211> 960

<212> DNA

<213> Bifidobacterium lactis (Bifidobacterium lactis)

<400> 2

acaggcttgc ggctctacca tgcagtcgaa cgggatccct ggcagcttgc tgtcggggtg 60

agagtggcga acgggtgagt aatgcgtgac caacctgccc tgtgcaccgg aatagctcct 120

ggaaacgggt ggtaataccg gatgctccgc tccatcgcat ggtggggtgg gaaatgcttt 180

tgcggcatgg gatggggtcg cgtcctatca gcttgttggc ggggtgatgg cccaccaagg 240

cgttgacggg tagccggcct gagagggtga ccggccacat tgggactgag atacggccca 300

gactcctacg ggaggcagca gtggggaata ttgcacaatg ggcgcaagcc tgatgcagcg 360

acgccgcgtg cgggatggag gccttcgggt tgtaaaccgc ttttgttcaa gggcaaggca 420

cggtttcggc cgtgttgagt ggattgttcg aataagcacc ggctaactac gtgccagcag 480

ccgcggtaat acgtagggtg cgagcgttat ccggatttat tgggcgtaaa gggctcgtag 540

gcggttcgtc gcgtccggtg tgaaagtcca tcgcctaacg gtggatctgc gccgggtacg 600

ggcgggctgg agtgcggtag gggagactgg aattcccggt gtaacggtgg aatgtgtaga 660

tatcgggaag aacaccaatg gcgaaggcag gtctctgggc cgtcactgac gctgaggagc 720

gaaagcgtgg ggagcgaaca ggattagata ccctggtagt ccacgccgta aacggtggat 780

gctggatgtg gggccctttc cacgggtccc gtgtcggagc caacgcgtta agcatcccgc 840

ctggggagta cggccgcaag gctaaaactc aaagaaattg acgggggccc gcacaagcgg 900

cggagcatgc ggattaattc gatgcaacgc gaagaacctt acctgggctt gacatgtgcc 960

<210> 3

<211> 1113

<212> DNA

<213> Lactobacillus plantarum (Lactobacillus plantarum)

<400> 3

ggcagtggcg ggtgctaata catgcagtcg aacgaactct ggtattgatt ggtgcttgca 60

tcatgattta catttgagtg agtggcgaac tggtgagtaa cacgtgggaa acctgcccag 120

aagcggggga taacacctgg aaacagatgc taataccgca taacaacttg gaccgcatgg 180

tccgagtttg aaagatggct tcggctatca cttttggatg gtcccgcggc gtattagcta 240

gatggtgagg taacggctca ccatggcaat gatacgtagc cgacctgaga gggtaatcgg 300

ccacattggg actgagacac ggcccaaact cctacgggag gcagcagtag ggaatcttcc 360

acaatggacg aaagtctgat ggagcaacgc cgcgtgagtg aagaagggtt tcggctcgta 420

aaactctgtt gttaaagaag aacatatctg agagtaactg ttcaggtatt gacggtattt 480

aaccagaaag ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg 540

ttgtccggat ttattgggcg taaagcgagc gcaggcggtt ttttaagtct gatgtgaaag 600

ccttcggctc aaccgaagaa gtgcatcgga aactgggaaa cttgagtgca gaagaggaca 660

gtggaactcc atgtgtagcg gtgaaatgcg tagatatatg gaagaacacc agtggcgaag 720

gcggctgtct ggtctgtaac tgacgctgag gctcgaaagt atgggtagca aacaggatta 780

gataccctgg tagtccatac cgtaaacgat gaatgctaag tgttggaggg tttccgccct 840

tcagtgctgc agctaacgca ttaagcattc cgcctgggga gtacggccgc aaaggctgaa 900

actcaaagga attgacgggg gcccgcacaa gcggtggagc atgtggttta attcgaagct 960

acgcgaagaa cccttaccag gtcttgacat actatgcaaa tctaagagat tagacgtttc 1020

cctttcgggg acatgggata caggtggtgc atggttgtcg tcagctcgtg tcgtgagatt 1080

gtttgggttt aagtccccgg caaacggagc ggc 1113

Claims

1. A preparation method of a lyophilized human milk powder composition containing infant probiotics is characterized in that the composition takes lyophilized human milk powder, prebiotics and infant probiotics powder as preparation raw materials, and the preparation method specifically comprises the following steps:

s1: taking probiotic strains obtained by separation and purification from infant intestinal tracts, activating and culturing for 3 generations, inoculating the probiotic strains into an aseptic liquid culture medium, fermenting and culturing, centrifuging to obtain infant-type probiotic mud, mixing and emulsifying the infant-type probiotic mud and a bacterial powder protective agent, freeze-drying at low temperature to remove water, crushing and sieving to obtain infant-type probiotic powder;

s2: taking healthy breast milk of a breast mother, carrying out pasteurization, then carrying out low-temperature freeze drying, removing water, crushing and sieving to obtain the human milk freeze-dried powder;

s3: and uniformly mixing the infant probiotic powder, the human milk freeze-dried powder and the prebiotics under an aseptic condition to obtain the human milk freeze-dried powder composition containing the infant probiotic.

2. The method for preparing lyophilized human milk powder composition containing probiotic bacteria of infancy species as claimed in claim 1, wherein the viable count of said lyophilized human milk powder composition containing probiotic bacteria of infancy species is greater than or equal to 1 x 10⁸CFU/g。

3. The method of claim 1, wherein the prebiotic comprises fructo-oligosaccharide and galacto-oligosaccharide, and the sum of the two masses is not less than 2/3 of the total mass of the prebiotic.

4. The method for preparing a lyophilized human milk powder composition containing probiotics for infant formulas according to claim 1, wherein the low-temperature lyophilization process in S2 is performed in a lyophilizer, and specifically comprises a pre-freezing process and a lyophilization process, wherein the pre-freezing process is performed for 2.5-3.5 h, the temperature of cold hydrazine in the lyophilizer is maintained at-30 to-40 ℃ and the vacuum degree is less than 30Pa in the whole low-temperature lyophilization process.

5. The method for preparing the lyophilized human milk powder composition containing probiotic bacteria of infancy as claimed in claim 4, wherein the lyophilization process is divided into 7 stages, wherein the duration of each stage in 1-6 stages is 1.5-2.5 h, the duration of 7 stages is 8-10 h, and the sample temperature is controlled to increase with the variation gradient of the lyophilization stage in the range of-25 ℃ to 35 ℃ during the lyophilization process.

6. A lyophilized human milk powder composition containing probiotic bacteria of infancy species, which is prepared by the preparation method of the lyophilized human milk powder composition containing probiotic bacteria of infancy species of any one of claims 1 to 5.

7. The lyophilized human milk powder composition comprising probiotic bacteria of infancy according to claim 6, wherein said probiotic bacteria powder of infancy is a mixture of powders of Bifidobacterium bifidum HRHCC001, Bifidobacterium lactis HRHCC002 and Lactobacillus plantarum HRHCC 003;

the deposit number of the bifidobacterium bifidum HRHCC001 is as follows: CGMCC No. 22656;

the deposit number of the bifidobacterium lactis HRHCC002 is as follows: CGMCC No. 22657;

the lactobacillus plantarum HRHCC003 has the deposit number of: CGMCC No. 22658.

8. The lyophilized human milk powder composition containing probiotic bacteria of infancy according to claim 7, wherein the mass percentages of the raw materials for preparation in the lyophilized human milk powder composition containing probiotic bacteria of infancy are as follows: 2% -4% of infant probiotic powder, 28% -32% of human milk freeze-dried powder and 65% -70% of prebiotics; and/or

The mass ratio of the bacterial powder prepared from bifidobacterium bifidum HRHCC001, the bacterial powder prepared from bifidobacterium lactis HRHCC002 and the bacterial powder prepared from lactobacillus plantarum HRHCC003 in the infant probiotic bacterial powder is as follows: 0.8-1.2: 0.8-1.2: 1.6-2.4; and/or

The viable count of the human milk freeze-dried powder composition containing infant probiotic is more than or equal to 1 x 10⁸CFU/g。

9. The lyophilized human milk powder composition comprising probiotic bacteria of infancy species according to claim 6, wherein the dosage form of said composition comprises powder, granules or tablets.

10. Use of the lyophilized human milk powder composition containing probiotic bacteria of infancy as claimed in any one of claims 6 to 8 in the field of infant drinks, foods and adult drinks and foods, characterized in that said infant drinks, foods are for infants over 3 years old.