CN109620951A - One kind helping pregnant composition and preparation method thereof - Google Patents

One kind helping pregnant composition and preparation method thereof Download PDF

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Publication number
CN109620951A
CN109620951A CN201811621491.6A CN201811621491A CN109620951A CN 109620951 A CN109620951 A CN 109620951A CN 201811621491 A CN201811621491 A CN 201811621491A CN 109620951 A CN109620951 A CN 109620951A
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China
Prior art keywords
parts
composition
pregnant
vitamin
raw material
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CN201811621491.6A
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Chinese (zh)
Inventor
徐红涛
谷欣
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Li Rui Guest Information Technology (beijing) Co Ltd
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Li Rui Guest Information Technology (beijing) Co Ltd
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Priority to CN201811621491.6A priority Critical patent/CN109620951A/en
Publication of CN109620951A publication Critical patent/CN109620951A/en
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    • A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
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    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/125—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives containing carbohydrate syrups; containing sugars; containing sugar alcohols; containing starch hydrolysates
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
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Abstract

The invention discloses one kind to help pregnant composition, raw material including following parts by weight: 2-6 parts of semen sojae atricolor, 5-13 parts of black-bone chicken albumen powder, 1-7 parts of pollen pini, 1-6 parts of L-carnitine, 4-9 parts and Radix Angelicae Sinensis 2-8 parts of ginger, it additionally provides the above-mentioned preparation method for helping pregnant composition and its is preparing the application in health care product and/or functional food, it is provided by the invention to help pregnant composition, it has no toxic side effect, raw material in formula interacts, prescription is scientific and reasonable, adjust tonifying five zang organs, play integrally-regulated advantage, female endocrinic disorders can effectively be adjusted, improve ovarian function, improve oocyte quality, improve female fertility.

Description

One kind helping pregnant composition and preparation method thereof
Technical field
The invention belongs to health care product and/or functional food fields, and in particular to one kind helps pregnant composition and preparation method thereof.
Background technique
Since infertility is known as female sterility caused by female factors, female sterility be a kind of disease incidence compared with High and more refractory illness more.Research data shows at present both at home and abroad: disease incidence of the infertility in child-bearing period Mr. and Mrs is about For 15%-20%, root according to the statistics of the World Health Organization: the whole world about 600-8000 ten thousand suffers from different degrees of infertility to Mr. and Mrs Disease, wherein the infertility as caused by female factors accounts for about the 50% of all infertile couples.Female fertility is in the increase at age Downward trend, the probability being pregnant in 1 year can reach 90% at 20 years old, and be only 36.4% at 40 years old or so.Chinese population base Number is big, and the opening of two child's policies in recent years, has the advanced age Mr. and Mrs of fertility demand more and more, the disease incidence of sterility is in recent years Also in rising trend.The cause of disease of atocia is varied, most directly shows as endocrine disturbance, premature ovarian failure, ovum Cell quality decline.
CN103316238A discloses a kind of Chinese medicine for treating fertility, is made of the medicinal material of following parts by weight: king is not Stay 12 parts of row, 12 parts of cortex cinnamomi, 10 parts of dark plum, 14 parts of Rhizoma Chuanxiong, 10 parts of fennel seeds, 6 parts of sweet osmanthus, 8 parts of rose, 10 parts of clearvers, 12 parts of Herba Cyperi Glomerati, 13 parts of Thesium chinese, 14 parts of amur foxtail, 12 parts of motherwort, 12 parts of root of Rumex patientia L., 12 parts of Radix Paeoniae Alba, clinical test proves, The present invention can safely and effectively treat fertility.CN106860717A disclose it is a kind of treat infertile drug, according to The raw material of parts by weight includes: 3-8 parts of fennel seeds, 5-10 parts of corydalis tuber, 3-8 parts of myrrh, 5-10 parts of Radix Angelicae Sinensis, 5-10 parts of Rhizoma Chuanxiong, cortex cinnamomi 8-15 parts, 1-3 parts of excrementum pteropi, 3-8 parts of motherwort, 3-8 parts of rhizoma cyperi, 8-15 parts of semen ziziphi spinosae, 1-3 parts of fushen, 1-3 parts of Radix Codonopsis, mountain 5-10 parts of the fruit of medicinal cornel, 3-8 parts of radix scutellariae, 1-5 parts of schizonepeta, 8-15 parts of Fructus Forsythiae, the invention is to caused by uncomfortable equal inferior healths reason It is infertile have it is effective.
In the prior art, the composition for improving and treating female acyesis, which focuses mostly on, to be changed to palace cold, inferior health the problems such as It is kind, and it is poor to the specific aim of infertility caused by oocyte quality problem, and it is infertile to oocyte quality treatment is improved at present Research it is less, CN107927790A discloses a kind of composition, including trehalose and L-carnitine, although the invention can mention High oocyte quality, but the invention has a single function, and therapeutic effect is not significant, can not effectively improve the fertility energy of women Power.
Summary of the invention
The present invention provides one kind to help pregnant composition, and the composition is safe and reliable, has no toxic side effect, and can effectively adjust women Endocrine disturbance improves ovarian function, improves oocyte quality, improves female fertility.
To achieve the goals above, the invention adopts the following technical scheme:
One kind helping pregnant composition, the raw material including following parts by weight: 2-6 parts of semen sojae atricolor, 5-13 parts of black-bone chicken albumen powder, and pollen pini 1-7 parts, 1-6 parts of L-carnitine, 4-9 parts of ginger and 2-8 parts of Radix Angelicae Sinensis.
Preferably, described to help pregnant composition, the raw material including following parts by weight: 3-6 parts of semen sojae atricolor, black-bone chicken albumen powder 6-13 Part, 2-7 parts of pollen pini, 2-6 parts of L-carnitine, 5-9 parts of ginger and 4-8 parts of Radix Angelicae Sinensis.
In some preferred embodiments, described to help pregnant composition, further include the raw material of following parts by weight: zinc-rich is rich 5-10 parts of selenium dried whole-egg, 1-5 parts of fructus lycii, 2-7 parts of Chinese yam, 2-6 parts of lotus seeds, 3-8 parts of longan, 1-5 parts of cortex cinnamomi and 5-15 parts of jujube； Preferably, the mass ratio of the cortex cinnamomi and Zinc-enriched selenium-enriched dried whole-egg is 1:3.
In some preferred embodiments, described to help pregnant composition, it further include the raw material of following parts by weight: fruit vegetable powder 8-15 parts, 1-8 parts of motherwort, 1-5 parts of trehalose, 1-6 parts of pueraria lobata, 2-8 parts of folium artemisiae argyi and 0.001-0.005 parts of vitamin；It is preferred that The mass ratio of ground, the trehalose and folium artemisiae argyi is 1:2.
In some preferred embodiments, described to help pregnant composition, it further include the raw material of following parts by weight: honey 5- 10 parts, 3-8 parts of inulin, 2-7 parts of Isin glue collagen peptide, 1-5 parts of maca and 0.0001-0.0005 parts of microelement；Preferably, institute The mass ratio for stating inulin and Isin glue collagen peptide is 1.5:1.
Preferably, the vitamin is vitamin C, vitamin E, folic acid, vitamin B1, vitamin B2, vitamin B6、 Vitamin B12, one of niacin and pantothenic acid or a variety of.
Preferably, the microelement is one of molysite, mantoquita and molybdenum salt or a variety of.
The present invention also provides a kind of preparation methods for helping pregnant composition, comprising the following steps:
(1) black-bone chicken albumen powder, pollen pini, Zinc-enriched selenium-enriched dried whole-egg, fruit vegetable powder, honey, inulin, Isin glue collagen will be removed All raw materials except peptide, vitamin and microelement are freeze-dried, are crushed into powder after the dosage mixing of composition of raw materials, Obtain mixture A；
(2) obtained mixture A is extracted with ethanol solution, is filtered after merging filtrate, is concentrated under reduced pressure to give medicinal extract B, The mass ratio of the mixture A and ethyl alcohol are 1:(4-7)；
(3) obtained medicinal extract B is obtained into mixture C after drying, crushing, sieving；
(4) mixture C that step (3) obtains is sufficiently mixed with remaining raw material is formulated, obtains described helping pregnant composition.
The condition of extraction described in step (2) are as follows: extracted 3 times at 60-70 DEG C, each extraction time is 3 hours, institute The ethanol solution stated is the ethanol water that volume fraction is 60-80%.
Sieving described in step (3) was 80 meshes.
It is of the present invention to help pregnant composition that be prepared as pulvis, tablet, pill, it can also be used as the addition of bottom side at it In his product.
The present invention also provides described, and pregnant composition to be helped to prepare the application in health care product and/or functional food.
The invention has the benefit that
(1) provided by the invention to help pregnant composition, it has no toxic side effect, the raw material in formula interacts, and prescription science is closed Reason adjusts tonifying five zang organs, plays integrally-regulated advantage, has and adjusts endocrine disturbance, improves ovarian function, improves ooplasm Amount, improves the effect of female fertility.
(2) provided by the invention to help pregnant composition that improve woman preparing for pregnancy constitution, have nourishing yin and supplementing blood, raising immune The effect of power, while the effect of interior free yl can be removed, improve skin elasticity, prevents the shape of pregnancy period cyasma and striae of pregnancy At.
Specific embodiment
In order to make the object, technical scheme and advantages of the embodiment of the invention clearer, below to technical solution of the present invention It is further described.
When embodiment provides numberical range, it should be appreciated that except non-present invention is otherwise noted, two ends of each numberical range Any one numerical value can be selected between point and two endpoints.Unless otherwise defined, all technologies used herein and section Technics has and the normally understood identical meaning of general technical staff of the technical field of the invention.
Unless otherwise specified, raw material of the present invention is ordinary commercial products.
Basic embodiment
One kind helping pregnant composition, the raw material including following parts by weight: 2-6 parts of semen sojae atricolor, 5-13 parts of black-bone chicken albumen powder, and pollen pini 1-7 parts, 1-6 parts of L-carnitine, 4-9 parts of ginger and 2-8 parts of Radix Angelicae Sinensis.
Preferably, described to help pregnant composition, the raw material including following parts by weight: 3-6 parts of semen sojae atricolor, black-bone chicken albumen powder 6-13 Part, 2-7 parts of pollen pini, 2-6 parts of L-carnitine, 5-9 parts of ginger and 4-8 parts of Radix Angelicae Sinensis.
In some preferred embodiments, described to help pregnant composition, further include the raw material of following parts by weight: zinc-rich is rich 5-10 parts of selenium dried whole-egg, 1-5 parts of fructus lycii, 2-7 parts of Chinese yam, 2-6 parts of lotus seeds, 3-8 parts of longan, 1-5 parts of cortex cinnamomi and 5-15 parts of jujube； Preferably, the mass ratio of the cortex cinnamomi and Zinc-enriched selenium-enriched dried whole-egg is 1:3.
In some preferred embodiments, described to help pregnant composition, it further include the raw material of following parts by weight: fruit vegetable powder 8-15 parts, 1-8 parts of motherwort, 1-5 parts of trehalose, 1-6 parts of pueraria lobata, 2-8 parts of folium artemisiae argyi and 0.001-0.005 parts of vitamin；It is preferred that The mass ratio of ground, the trehalose and folium artemisiae argyi is 1:2.
In some preferred embodiments, described to help pregnant composition, it further include the raw material of following parts by weight: honey 5- 10 parts, 3-8 parts of inulin, 2-7 parts of Isin glue collagen peptide, 1-5 parts of maca and 0.0001-0.0005 parts of microelement；Preferably, institute The mass ratio for stating inulin and Isin glue collagen peptide is 1.5:1.
Preferably, the vitamin is vitamin C, vitamin E, folic acid, vitamin B1, vitamin B2, vitamin B6、 Vitamin B12, one of niacin and pantothenic acid or a variety of.
Preferably, the microelement is one of molysite, mantoquita and molybdenum salt or a variety of.
It is of the present invention to help pregnant composition that be prepared as pulvis, tablet, pill, it can also be used as the addition of bottom side at it In his product.
The present invention also provides described, and pregnant composition to be helped to prepare the application in health care product and/or functional food.
Table 1-4 is the specific raw material components table of the present embodiment, wherein the "-" in table indicates that this content is not present.
Protein content is 85% in the black-bone chicken albumen powder that the present embodiment uses；The preparation method of Zinc-enriched selenium-enriched dried whole-egg Are as follows: by the egg liquid and eggshell of Se-enriched egg and zinc-rich egg, moisture content is made according to conventional technical means in the art and is lower than 4.0% dried whole-egg, wherein the Se content of Se-enriched egg is greater than 0.2mg/kg, and the Zn content of zinc-rich egg is greater than 0.822mg/ The mass ratio of kg, zinc-rich egg and Se-enriched egg is 2:1；Pollen pini is commercially available common broken masson pine pollen；Isin glue collagen peptide The content of middle protein is 97.5%；Fruit vegetable powder is grape, apple, lemon, pimento, tomato and carrot according to this field The pulvis of conventional technical means preparation, the grape, apple, lemon, pimento, tomato and the mass ratio of carrot are 0.5:1: 0.2:1:1:2；The levulan content of inulin is 90%；Vitamin is vitamin E；Microelement is ferrous gluconate.
Raw material components (parts by weight) table of 1 embodiment 1-5 of table
Raw material components (parts by weight) table of 2 embodiment 6-10 of table
Raw material components (parts by weight) table of 3 embodiment 11-15 of table
Raw material components (parts by weight) table of 4 embodiment 16-20 of table
The preparation method for helping pregnant composition of above-described embodiment 1-20, comprising the following steps:
(1)) black-bone chicken albumen powder, pollen pini, Zinc-enriched selenium-enriched dried whole-egg, fruit vegetable powder, honey, inulin, Isin glue collagen will be removed All raw materials except peptide, vitamin and microelement are freeze-dried, are crushed into powder after the dosage mixing of composition of raw materials, Obtain mixture A；
(2) ethanol water that obtained mixture A volume fraction is 66% is extracted into extraction 3 times at 65 DEG C, often It secondary 3 hours, filtered after merging filtrate, be concentrated under reduced pressure to give medicinal extract B, the mass ratio of the mixture A and ethyl alcohol is 1:5；
(3) obtained medicinal extract B is obtained into mixture C after drying, crushing, 80 meshes excessively；
(4) mixture C that step (3) obtains is sufficiently mixed with remaining raw material is formulated, crushes, crosses 80 meshes, obtain It is described to help pregnant composition.
Comparative example 1
For basic components with embodiment 4, the difference with embodiment 4 is that black-bone chicken albumen powder is free of in formula；Preparation method With embodiment 4.
Comparative example 2
For basic components with embodiment 4, the difference with embodiment 4 is that pollen pini is free of in formula；Preparation method is the same as real Apply example 4.
Comparative example 3
For basic components with embodiment 8, the difference with embodiment 8 is that Zinc-enriched selenium-enriched dried whole-egg is free of in formula；Preparation Method is the same as embodiment 8.
Comparative example 4
For basic components with embodiment 8, the difference with embodiment 8 is that cortex cinnamomi is free of in formula；Preparation method is the same as implementation Example 8.
Comparative example 5
For basic components with embodiment 15, the difference with embodiment 15 is that trehalose is free of in formula；Preparation method is same Embodiment 15.
Comparative example 6
For basic components with embodiment 15, the difference with embodiment 15 is that folium artemisiae argyi is free of in formula；Preparation method is the same as real Apply example 15.
Comparative example 7
For basic components with embodiment 19, the difference with embodiment 19 is that Isin glue collagen peptide is free of in formula；Preparation Method is the same as embodiment 19.
Influence of the experimental example 1 to egg cell
Experimental animal: 8 weeks female CD-1 mouse after birth
Mouse is randomly divided into 8 groups, every group 10, respectively control group, embodiment 1-5 group and comparative example 1-2 group.Implement Example 1-5 group gives the composition of 1-5 of the embodiment of the present invention respectively, and comparative example 1-2 group gives the comparative example 1-2 composition of preparation, According to the dosage of 20mg/kg, daily stomach-filling 1 time, control group gives the distilled water of equivalent and uses rush after successive administration 30 days Super drug row object promotees super row, collects egg cell after mouse is euthanized, and assesses the quality and quantity of egg cell, and using in vitro fertilization Its ability for forming blastaea of experimental evaluation.
Experimental result:
Influence of 5 present invention of table to egg cell
Group Egg cell degradation ratio % Blastocyst rate %
Control group 32.6 56.8
Example 1 group 17.2* 91.2*
2 groups of embodiment 17.4* 90.5*
3 groups of embodiment 16.8* 92.3*
4 groups of embodiment 14.8** 96.8**
5 groups of embodiment 15.2** 94.2**
1 group of comparative example 20.7 75.6
2 groups of comparative example 19.9 80.2
Note: compared with the control group, * P < 0.05 has significant difference；* P < 0.01 has extremely significant sex differernce.
As shown in Table 5, the embodiment of the present invention 1-5 can improve the quality of egg cell, find simultaneously, when black in composition When the mass ratio of egg white powder and pollen pini is 2:1, the quality of egg cell can be significantly improved.
Influence of the embodiment 2 to premature ovarian failure
Experimental animal: SD rat, weight 200-280g, female.
Modeling；It carries out continuous vaginal smear within 7 days before modeling, to observe the oestrous cycle of rat, selects the oestrous cycle normal 120 SD rats, rat is divided into control group (20) and model group (100) at random by weight.Control group intraperitoneal injection Physiological saline 2 times, is spaced 11 days totally；Modeling group intraperitoneal injection of cyclophosphamide solution, totally 2 times, 90mg/kg (the 1st time), 50mg/ Kg (the 2nd time) is spaced 11 days；It is daily to carry out vaginal smear, the variation of observation oestrous cycle；It is disorderly when the model group female mice oestrous cycle Disorderly, anoestrum extended, without oestrous cycle or estrus, indicated rat ovary early ageing modeling success, hormonal readiness changes conduct Auxiliary reference index.
Experimental group: blank control group, model control group, embodiment 6-10 group and comparative example 3-4 group.
Test method: choosing 80 successful rats of modeling, is randomly divided into 8 groups by weight, and every group 10, embodiment 6-10 Group gives the composition of 6-10 of the embodiment of the present invention respectively, and comparative example 3-4 group gives the comparative example 3-4 composition of preparation, and every group The dosage of rat is 0.5g/kg, and model control group and blank control group give isometric physiological saline, daily stomach-filling one It is secondary, it successive administration 30 days, in last dose next day, detects each rats in test groups serum follicular stimulating hormone (FSH) and corpus luteum generates The content of plain (LH).
Test result:
Table 6 detects the content of each group test rat blood serum FSH and LH
Group LH(mIU/ml) FSH(mIU/ml)
Blank control group 2.87±0.34 3.10±0.55
Model control group 3.15±0.50# 3.40±0.79#
6 groups of embodiment 2.65±0.23* 2.73±0.47*
7 groups of embodiment 2.74±0.38* 2.83±0.80*
8 groups of embodiment 2.15±0.26** 2.45±0.62**
9 groups of embodiment 2.69±0.43* 2.75±0.67*
10 groups of embodiment 2.29±0.32** 2.56±0.58**
3 groups of comparative example 2.95±0.36 3.16±0.71
4 groups of comparative example 2.90±0.34 3.20±0.58*
Note: compared with blank control group, #P < 0.05 has significant difference；Compared with model control group, * P < 0.05, there is significant difference；* P < 0.01 has extremely significant sex differernce.
As shown in Table 6, compared with blank control group, model control group rat blood serum LH and FSH have significant difference, table Bright model control group modeling success.Simultaneously it can be found that rat blood serum LH and FSH, table can be effectively reduced in the present invention from table 6 The bright present invention can be effectively improved and treat premature ovarian failure, restore ovarian function.When the matter of dried whole-egg Zinc-enriched selenium-enriched in composition When amount is than being 3:1, effect is preferable.
Total antioxidation acts in 3 body of experimental example
Experimental animal: Kunming mouse, female mice, weight 18-22g.
Experimental group: control group, embodiment 11-20 group and comparative example 5-8 group.
Experimental method: mouse is randomly divided into 15 groups by weight, every group 10, embodiment 11-20 group gives this hair respectively The composition of bright embodiment 11-20, comparative example 5-8 group give the comparative example 5-8 composition of preparation, and the dosage of every group of mouse is 0.1g/kg, control group give isometric physiological saline, and daily stomach-filling is primary, and after continuous gavage 30 days, every group of mouse pulls out eyeball Take blood and the execution that dislocates, 3500r/min separates serum after being centrifuged 15 minutes, is placed in that -20 DEG C of preservation are alternative, total in measurement serum Oxidation resistance (T-AOC).
Experimental result:
7 total antioxidation of table acts on (T-AOC) experimental data
Note: compared with the control group, * P < 0.05 has significant difference；* P < 0.01 has extremely significant sex differernce.
The T-AOC that can be seen that the mouse of control group from the experimental result of table 7 is 13.5U/ml, and the feeding present invention The T-AOC of the mouse for helping pregnant composition provided is 21.2-29.9U/ml, therefore, provided by the invention that pregnant composition is helped to have Oxidation resistant effect, the effect of can removing interior free yl, improve skin elasticity, prevent the shape of pregnancy period cyasma and striae of pregnancy At.
4 strengthen immunity of experimental example
Detect foundation: Ministry of Public Health's " health food is examined and assessment technique specification " (version in 2003).
Experimental animal: Kunming mouse, female, weight 18-22g.
This composition human body recommended amounts are everyone daily 50ml (0.83ml/kg), and mouse experiment dosage is arranged low, just Often, high 3 dosage levels, i.e., daily 4.15,8.30 and 24.90ml/kg (are respectively equivalent to 5,10, the 30 of human body recommended amounts Times), while negative control group is set, experiment mice is randomly divided into 2 batches, 4 groups of every batch of, every group 10 according to weight.
Respectively weigh prepared by 5.25ml, 10.50ml and 31.25ml above-described embodiment 19 and comparative example 7 helps pregnant composition, point Not plus pure water is to 50ml, is made into 0.11ml/ml, 0.21ml/ml and 0.63ml/ml strength solution respectively, stomach-filling volume is 0.2ml/10g weight, daily stomach-filling 1 time, negative control group give isometric pure water, continuous 30d.
4.1 cellular immune function
1, the mouse spleen lymphocyte transformation experiment (mtt assay) of ConA induction
It is sterile to take spleen, cell suspension is made.It is divided to two holes to be added in 24 well culture plates every a splenocyte suspension, every hole 1mL, a hole add 75 μ L ConA liquid (being equivalent to 7.5 μ g/mL), and 5%CO is set as control in another hole2, 37 DEG C of CO2It is trained in incubator Support 72h.Culture terminates preceding 4h, and every hole gently sucks supernatant 0.7mL, and RPMI1640 of the 0.7mL without calf serum is added and trains Nutrient solution, while 50 hole μ L/ MTT (5mg/mL) is added, continue to cultivate 4h.After culture, 1mL acid isopropyl alcohol is added in every hole, Ultrasonic vibration (2 seconds), is completely dissolved purple crystal.Then it is dispensed into 96 well culture plates, each hole dispenses 3 holes as parallel Sample measures OD value with enzyme-linked immunosorbent assay instrument with 570nm wavelength.ConA is not added with adding the OD value in the hole ConA to subtract The OD value in hole represents the proliferative capacity of lymphocyte, and the light that the optical density difference of test sample group is significantly higher than control group is close Difference is spent, can determine that this experimental result positive.
2, delayed allergy (DTH)
Stomach-filling the 26th day, it is used for sensitization to mouse peritoneal injection 2% sheep red blood cell (SRBC) (SRBC) 0.2mL, 4th after immune It, auris dextra uniformly smears 20 μ L of 1%DNFB solution attack, and left ear compares, and in putting to death mouse for 24 hours after injection, takes left and right identical The weighing of size auricle, the difference of weight is as swelling.The degree of DTH is indicated with swelling.The difference of test sample group is aobvious The difference for being higher than control group is write, can determine that this experimental result positive.Thymus gland and the spleen weighing for taking mouse simultaneously, calculate dirty Device/weight ratio.
4.2 humoral immune function
1, antibody-producting cell detection (Jerne improves slide method)
In the 26th day 2% (v/v) hematocrit SRBC to every mouse peritoneal injection 0.2mL of stomach-filling, 4-5 is immunized in SRBC Mouse cervical dislocation after it is put to death, sterile to take spleen, is prepared into 5 × 10 with complete medium RPMI16406A cell/mL's Splenocyte suspension.And agarose slide is made, put it into the CO of 37 DEG C, 5%21h is incubated in incubator, then every hole is added 500 μ L are continued to be incubated for 2h, be formed on every thin agar layer slide of counting with the diluted complement of complete medium (v/v, 1:10) Hemolysis plaque number, with plaque number/106Splenocyte indicates antibody-producting cell number.
2, half hemolytic value (HC50) measurement
Hematocrit SRBC is made into the cell suspension of 2% (v/v) with physiological saline, every mouse intraperitoneal injection 0.2mL is exempted from Epidemic disease.After 4d, extracts eyeball and takes blood in centrifuge tube, place about 1h, solidification blood and tube wall are removed, serum is precipitated sufficiently, 2000r/min is centrifuged 10min, collects serum.Serum SA buffer is taken to dilute 200 times.Serum 1mL after dilution is set into test tube It is interior, 10% (v/v) SRBC of 0.5mL is sequentially added, complement 1mL (is diluted with SA liquid by 1:8).Separately set the control tube of not increase serum (with the replacement of SA buffer).It sets after keeping the temperature 20min in 37 DEG C of waters bath with thermostatic control, ice bath terminates reaction.2000r/min is centrifuged 10min. Supernatant 1mL is taken, Dou Shi reagent 3mL is added, while 10% (v/v) SRBC of 0.25mL being taken to add Dou Shi reagent to 4mL, it is sufficiently mixed It is even, after placing 10min, blank is made with control tube at 540nm, measures each pipe OD value respectively.
The amount of hemolysin is with half hemolytic value (HC50) indicate, it is calculated according to the following formula:
The HC of test sample group50It is significantly higher than the HC of control group50, can determine that this experimental result positive.
4.3 monocytes/macrophages functions
1, mouse carbonic clearance is tested
Diluted india ink is injected from mouse tail vein, is calculated by every 10g weight 0.1mL.It injects to prepared Chinese ink, counts immediately When.2,10min after injection prepared Chinese ink, take 20 μ L of blood from angular vein clump respectively, exist side by side and be added into the 1%Na of 2mL2CO3Solution In, survey OD600nmSpleen blots organ surface blood stains with filter paper, weighing.
The ability of mouse carbonic clearance is indicated with phagocytic index.
K=(lgOD2min-1gOD10min)/(10min-2min)
The phagocytic index of test sample group is significantly higher than the phagocytic index of control group, can determine that this experimental result positive.
2, Turnover of Mouse Peritoneal Macrophages phagocytosis chicken red blood cell experiment (drop piece method)
2% (v/v) hematocrit SRBC0.2mL is injected to every mouse peritoneal and activates peritoneal macrophage, puts to death mouse after 5d, Intraperitoneal injection plus calf serum Hank ' s liquid 4mL/ mouse, gently rub abdomen 20 times, and 1% chicken for drawing abdominal cavity washing lotion and equivalent is red Cell mixes.1mL mixed liquor is drawn, piece is dripped, 37 DEG C of incubation 20min are rinsed, and fixed, Giemsa liquid dyeing, microscopy, counting gulps down Biting rate and phagocytic index, (phagocytic rate is that percentage shared by the macrophage of chicken red blood cell is swallowed in every 100 macrophages；It gulps down Bite the number that index is average each macrophage phagocytosis chicken red blood cell).
4.4NK cytoactive detection
1, lactic dehydrogenase (LDH) measuring method
It is sterile to take spleen, individual cells suspension is made as effector cell.Take target cell and each 100 μ L of effector cell (effect target Than 50:1), it is added in U-shaped 96 well culture plate；Target cell Spontaneous release hole adds target cell and each 100 μ L of culture solution, and target cell is most Big relief hole adds target cell and each 100 μ L of 2.5%Triton；Above-mentioned items are all provided with three multiple holes, in 37 DEG C, 5%CO2Incubator Then 96 well culture plates are centrifuged 5min with 1500r/min by middle culture 4h, the culture of 100 μ L horizontalization bottom of supernatant, 96 hole is drawn in every hole In plate, while 100 μ L of LDH matrix liquid is added, reacts 3min, the 30 μ L of HCl of 1mol/L is added in every hole, at microplate reader 490nm It measures OD value (OD).
The NK cell activity of test sample group is significantly higher than the NK cell activity of control group, that is, can determine that this experimental result It is positive.
Experimental result:
The experimental result of embodiment 19 is as shown in table 8.1 and 8.2.
Influence of 8.1 sample of table to mouse cell and humoral immune function
Note: control group is compared, P < 0.05 *, has significant difference；P < 0.01 * has extremely significant sex differernce.
By table 8.1 as it can be seen that low, middle dose group lymphopoiesis ability and auricle swelling degree weight gain are higher than control group, With significant difference (P < 0.05), lymphopoiesis ability and the auricle swelling degree weight gain of high dose group are higher than control group, With extremely significant sex differernce (P < 0.01)；The half hemolytic value of middle and high dosage group be higher than control group, have significant difference (P < 0.05)。
8.2 sample of table is to mouse monokaryon-macrophage and NK cytoactive detection result
Note: control group is compared, P < 0.05 *, has significant difference；P < 0.01 * has extremely significant sex differernce.
By table 8.2 as it can be seen that the NK cell activity of middle dose group is higher than control group, have significant difference (P < 0.05), it is high The NK cell activity of dosage group is higher than control group, has extremely significant sex differernce (P < 0.01).
The Evaluation of Immunity experimental result of comparative example 7 is as shown in table 9.1 and table 9.2.
Influence of 9.1 sample of table to mouse cell and humoral immune function
Note: control group is compared, P < 0.05 *, has significant difference.
By table 9.1 as it can be seen that lymphopoiesis ability and the auricle swelling degree weight gain of middle and high dosage group are higher than control group, With significant difference (P < 0.05).
9.2 sample of table is to mouse monokaryon-macrophage and NK cytoactive detection result
Note: P < 0.05 * indicates there is significant difference compared with the control group.
By table 9.2 as it can be seen that the NK cell activity of high dose group is higher than control group, have significant difference (P < 0.05).
Intact animal experiment is all made of in the application.In cellular immune function, humoral immune function, monocytes/macrophages In the four aspect measurements of function and NK cell activity, any two aspect test results are the positive, it is possible to determine that given the test agent tool There is strengthen immunity effect.Wherein the result of two experiments is the positive or any experiment in cellular immune function assay project Two dosage group results be the positive, determine that cellular immune function test result is positive.Two in humoral immune function measurement item The result of a experiment is two dosage group results of the positive or any experiment for the positive, determines humoral immune function test knot Fruit is positive.The result of two experiments is two agent of the positive or any experiment in mononuclear-macrophage function assay project Amount group result is the positive, determines that monokaryon-macrophage function test result is positive.NK cytoactive detection experiment one with Upper Dose Results are positive, determine that NK cell activity result is positive.
Therefore, the present embodiment 19 has the effect of strengthen immunity, meanwhile, by comparative example 7 it can be found that inulin and fish Collagen peptide synergistic effect, can significantly improve the spot number hemolysis value and NK cell activity of composition, to play better Immunization.

Claims (10)

1. one kind helps pregnant composition, which is characterized in that the raw material including following parts by weight: 2-6 parts of semen sojae atricolor, black-bone chicken albumen powder 5-13 Part, 1-7 parts of pollen pini, 1-6 parts of L-carnitine, 4-9 parts of ginger and 2-8 parts of Radix Angelicae Sinensis.
2. according to claim 1 help pregnant composition, which is characterized in that described helps pregnant composition, including following weight The raw material of part: 3-6 parts of semen sojae atricolor, 6-13 parts of black-bone chicken albumen powder, 2-7 parts of pollen pini, 2-6 parts of L-carnitine, 5-9 parts of ginger and Radix Angelicae Sinensis 4-8 parts.
3. according to claim 2 help pregnant composition, which is characterized in that further include the raw material of following parts by weight: zinc-rich is rich 5-10 parts of selenium dried whole-egg, 1-5 parts of fructus lycii, 2-7 parts of Chinese yam, 2-6 parts of lotus seeds, 3-8 parts of longan, 1-5 parts of cortex cinnamomi and 5-15 parts of jujube.
4. according to claim 3 help pregnant composition, which is characterized in that further include the raw material of following parts by weight: fruit vegetable powder 8-15 parts, 1-8 parts of motherwort, 1-5 parts of trehalose, 1-6 parts of pueraria lobata, 2-8 parts of folium artemisiae argyi and 0.001-0.005 parts of vitamin.
5. according to claim 4 help pregnant composition, which is characterized in that the vitamin be vitamin C, vitamin E, Folic acid, vitamin B1, vitamin B2, vitamin B6, vitamin B12, one of niacin and pantothenic acid or a variety of.
6. according to claim 5 help pregnant composition, which is characterized in that further include the raw material of following parts by weight: honey 5- 10 parts, 3-8 parts of inulin, 2-7 parts of Isin glue collagen peptide, 1-5 parts of maca and 0.0001-0.0005 parts of microelement.
7. according to claim 6 help pregnant composition, which is characterized in that the microelement is molysite, mantoquita and molybdenum One of salt is a variety of.
8. a kind of prepare the described in any item methods for helping pregnant composition of claim 1-7, which is characterized in that the preparation side Method, comprising the following steps:
(1) black-bone chicken albumen powder, pollen pini, Zinc-enriched selenium-enriched dried whole-egg, fruit vegetable powder, honey, inulin, Isin glue collagen peptide, dimension will be removed All raw materials except raw element and microelement are freeze-dried, are crushed into powder, obtain after the dosage mixing of composition of raw materials Mixture A；
(2) obtained mixture A is extracted with ethanol solution, is filtered after merging filtrate, is concentrated under reduced pressure to give medicinal extract B, it is described Mixture A and ethyl alcohol mass ratio be 1:(4-7)；
(3) obtained medicinal extract B is obtained into mixture C after drying, crushing, sieving；
(4) mixture C that step (3) obtains is sufficiently mixed with remaining raw material is formulated, obtains described helping pregnant composition.
9. preparation method according to claim 8, which is characterized in that the condition of extraction described in step (2) are as follows: It is extracted 3 times at 60-70 DEG C, 3 hours each extraction times, the ethanol solution is the ethanol water that volume fraction is 60-80% Solution.
10. the described in any item compositions of claim 1-7 are preparing the application in health care product and/or functional food.
CN201811621491.6A 2018-12-28 2018-12-28 One kind helping pregnant composition and preparation method thereof Pending CN109620951A (en)

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CN113730558A (en) * 2021-09-09 2021-12-03 广东圆康再生医学科技开发有限公司 Ovarian oocyte repair composition and preparation method thereof

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