CN108174919A - A kind of leben and preparation method thereof - Google Patents
A kind of leben and preparation method thereof Download PDFInfo
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- CN108174919A CN108174919A CN201711422480.0A CN201711422480A CN108174919A CN 108174919 A CN108174919 A CN 108174919A CN 201711422480 A CN201711422480 A CN 201711422480A CN 108174919 A CN108174919 A CN 108174919A
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/13—Fermented milk preparations; Treatment using microorganisms or enzymes using additives
- A23C9/1307—Milk products or derivatives; Fruit or vegetable juices; Sugars, sugar alcohols, sweeteners; Oligosaccharides; Organic acids or salts thereof or acidifying agents; Flavours, dyes or pigments; Inert or aerosol gases; Carbonation methods
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Abstract
A kind of leben disclosed by the invention, raw material include Paenibacillus polymyxa, skimmed milk, sweetener and/or acid, disclosed herein as well is the methods for preparing the leben, the Paenibacillus polymyxa that deposit number is CGMCC No.10062 is inoculated in skimmed milk including (a) and is fermented, acidified milk is obtained after fermentation;(b) it blends to form blending liquid using sweetener and/or acid, by acidified milk mixing made from blending liquid and step (a), homogeneous, sterilizing obtains leben.The advantage of the invention is that, the leben that the method for the present invention obtains, it being capable of 1 half-life period of GLP in prolonged human body, enhance the insulin secretion of glucose induction, adjust blood sugar concentration, compared with the bacterial strain of other normal fermentation skimmed milks, the acidified milk prepared by Paenibacillus polymyxa CGMCC No.10062 is more notable to the inhibitory activity of DPP IV, and stability is preferable.
Description
Technical field
The invention belongs to dairy products technical field, specially a kind of leben and preparation method thereof.
Background technology
Leben of the prior art refers to the beverage containing milk, would generally contain in this leben a large amount of
Sugar so the patient of hyperglycemia cannot drink, thus restricts the developing in leben market and functional abundant, existing
There is not the leben that can quote diabetes patient to appear on the market in market, this, which is that those skilled in the art are urgently to be resolved hurrily, asks
Topic.
Invention content
To solve the defects of leben eurytopicity of the prior art is not strong, the present invention provides a kind of acidified milk drinks
Method of material and preparation method thereof, the purpose of realization is prosthodontic treatment functionality leben blank in the market, is enriched
The functionality of leben is allowed to be suitble to wider audience.
To achieve these goals, the present invention provides following technical scheme:A kind of leben, raw material include mostly viscous
Series bacillus, skimmed milk, sweetener and/or acid, the deposit number of the Paenibacillus polymyxa is CGMCC
No.10062, Classification And Nomenclature be Paenibacillus polymyxa, preservation date be on November 26th, 2014, the culture presevation
In China General Microbiological culture presevation administrative center (China General Microbiological Culture
Collection Center, CGMCC), address:Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Chinese Academy of Sciences microorganism
Research institute, postcode:100101, Paenibacillus polymyxa is in the final concentration of 2x10 of leben6~2x107cfu/mL。。
Further, it is 9%~13% that the sugariness of the leben, which is converted into sweetness of cane sugar,.
Further, the titratable acidity of the leben is 65 ° of T~75 ° T.
Further, the leben is to inhibiting rate >=75% of dipeptidyl peptidase-IV.
Further, the skimmed milk includes skimmed milk powder and water, skimmed milk powder account for the mass percent 5% of skimmed milk~
10%.
The invention also discloses the methods for preparing above-mentioned leben, include the following steps:
(a) Paenibacillus polymyxa that deposit number is CGMCC No.10062 is inoculated in skimmed milk and fermented,
Acidified milk is obtained after fermentation;
(b) it blends to form blending liquid using sweetener and/or acid, by acidified milk made from blending liquid and step (a)
Mixing, homogeneous, sterilizing obtain leben.
Further, the temperature fermented in the step (a) is 25 DEG C~35 DEG C, and the time of fermentation is 3d~7d.
Further, in step (a), the fermentation is shaken cultivation, and hunting speed is 125rpm~220rpm.
Further, in step (b), the mixed volume ratio of the acidified milk and blending liquid is 1:1~1:4.
Further, the pressure of homogeneous is 15~30MPa in step (b).
Further, the temperature to sterilize in step (b) is 95 DEG C~125 DEG C, and the time of sterilizing is 5min~20min.
The present invention is using above-mentioned technical proposal, including following advantageous effect:The leben obtained through the method for the present invention,
Can GLP-1 half-life period in prolonged human body, enhance the insulin secretion of glucose induction, blood sugar concentration adjusted, with the prior art
It compares, the preparation method in above-mentioned technical proposal, for the first time using Paenibacillus polymyxa CGMCC No.10062, is made with skimmed milk
It ferments for culture medium, is prepared with dipeptidyl peptidase-IV inhibitory activity leben, is taken off with other normal fermentations
The bacterial strain of fat breast is compared, suppression of the acidified milk prepared by Paenibacillus polymyxa CGMCC No.10062 to dipeptidyl peptidase-IV
System activity is more notable, and stability is preferable.
In addition, the preparation method in above-mentioned technical proposal greatlies simplify production stage, production cost is saved, is dropped simultaneously
The pollution risk brought in low noncontinuity operation;Fermentation medium derives from a wealth of sources, is of low cost, is natural used by preparation
Safety, while Material Cost is reduced, improves the safety of food.
Specific embodiment
The present invention is described in further detail below by specific embodiment.It is not specified in the following example specific
The experimental method of condition selects according to conventional methods and conditions or according to product manual.If the reagent used in embodiment
Not plus illustrate, be analytical reagents, buy from Chinese medicines group.Other test apparatuses, reagent, strain, if do not illustrated,
Can directly it be bought by commercial sources.
Embodiment one:A kind of leben provided by the invention, the raw material of the beverage include Paenibacillus polymyxa,
Skimmed milk, sweetener and/or acid, the deposit number of the Paenibacillus polymyxa is CGMCC No.10062, mostly viscous class
Bacillus is in the final concentration of 2x10 of leben6cfu/mL。
It is 9% that the sugariness of the leben, which is converted into sweetness of cane sugar,.The sweetener is the sweet taste of this field routine
It is one or more in agent, preferably xylitol, Aspartame, honey element and acesulfame potassium.
The titratable acidity of the leben is 65 ° of T, and acid is the acid of this field routine, preferably lemon
It is one or more in lemon acid, malic acid, tartaric acid, lactic acid and acetic acid.
The leben is to inhibiting rate >=75% of dipeptidyl peptidase-IV.
The skimmed milk includes skimmed milk powder and water, and skimmed milk powder accounts for the mass percent 5% of skimmed milk.
The method for preparing above-mentioned leben, includes the following steps:
(a) Paenibacillus polymyxa that deposit number is CGMCC No.10062 is inoculated in skimmed milk and fermented,
Acidified milk is obtained after fermentation;
(b) it blends to form blending liquid using sweetener and/or acid, by acidified milk made from blending liquid and step (a)
Mixing, homogeneous, sterilizing obtain leben.
It is described to ferment for shaken cultivation, hunting speed 125rpm in step (a).Suddenly in (b), the acidified milk is with hooking
The mixed volume ratio for converting liquid is 1:1.The pressure of homogeneous is 15MPa in step (b).The temperature to sterilize in step (b) is 95 DEG C,
The time of sterilizing is 5min.
Embodiment two:A kind of leben provided by the invention, the raw material of the beverage include Paenibacillus polymyxa,
Skimmed milk, sweetener and/or acid, the deposit number of the Paenibacillus polymyxa is CGMCC No.10062, mostly viscous class
Bacillus is in the final concentration of 2x10 of leben7cfu/mL。
It is 13% that the sugariness of the leben, which is converted into sweetness of cane sugar,.The sweetener is the sweet taste of this field routine
It is one or more in agent, preferably xylitol, Aspartame, honey element and acesulfame potassium.
The titratable acidity of the leben is 75 ° of T, and acid is the acid of this field routine, preferably lemon
It is one or more in lemon acid, malic acid, tartaric acid, lactic acid and acetic acid.
The leben is to inhibiting rate >=75% of dipeptidyl peptidase-IV.
The skimmed milk includes skimmed milk powder and water, and skimmed milk powder accounts for the mass percent 10% of skimmed milk.
The method for preparing above-mentioned leben, includes the following steps:
(a) Paenibacillus polymyxa that deposit number is CGMCC No.10062 is inoculated in skimmed milk and fermented,
Acidified milk is obtained after fermentation;
(b) it blends to form blending liquid using sweetener and/or acid, by acidified milk made from blending liquid and step (a)
Mixing, homogeneous, sterilizing obtain leben.
It is described to ferment for shaken cultivation, hunting speed 220rpm in step (a).Suddenly in (b), the acidified milk is with hooking
The mixed volume ratio for converting liquid is 1:4.The pressure of homogeneous is 30MPa in step (b).The temperature to sterilize in step (b) is 125
DEG C, the time of sterilizing is 20min.
Embodiment three:A kind of leben provided by the invention, the raw material of the beverage include Paenibacillus polymyxa,
Skimmed milk, sweetener and/or acid, the deposit number of the Paenibacillus polymyxa is CGMCC No.10062, mostly viscous class
Bacillus is in the final concentration of 6x10 of leben6cfu/mL。
It is 11% that the sugariness of the leben, which is converted into sweetness of cane sugar,.The sweetener is the sweet taste of this field routine
It is one or more in agent, preferably xylitol, Aspartame, honey element and acesulfame potassium.
The titratable acidity of the leben is 80 ° of T, and acid is the acid of this field routine, preferably lemon
It is one or more in lemon acid, malic acid, tartaric acid, lactic acid and acetic acid.
The leben is to inhibiting rate >=75% of dipeptidyl peptidase-IV.
The skimmed milk includes skimmed milk powder and water, and skimmed milk powder accounts for the mass percent 7% of skimmed milk.
The method for preparing above-mentioned leben, includes the following steps:
(a) Paenibacillus polymyxa that deposit number is CGMCC No.10062 is inoculated in skimmed milk and fermented,
Acidified milk is obtained after fermentation;
(b) it blends to form blending liquid using sweetener and/or acid, by acidified milk made from blending liquid and step (a)
Mixing, homogeneous, sterilizing obtain leben.
It is described to ferment for shaken cultivation, hunting speed 185rpm in step (a).Suddenly in (b), the acidified milk is with hooking
The mixed volume ratio for converting liquid is 1:3.The pressure of homogeneous is 20MPa in step (b).The temperature to sterilize in step (b) is 110
DEG C, the time of sterilizing is 10min.
Through the above-mentioned leben prepared, to inhibiting rate >=75% of dipeptidyl peptidase-IV, II types can be helped sugared
Urinate patient's hypoglycemic.
The leben that each embodiment is prepared can be used following methods and obtain liquid to be detected:DPP-IV is pressed down
Preparation is dissolved in a concentration of 100mmol/L, in the Tris-HCl buffer solutions of pH 8.0, at room temperature, 10,000g centrifugation 10min,
Take supernatant, you can for the detection of dipeptidyl peptidase-IV inhibitory activity contained in leben.
The screening model of dipeptidyl peptidase-iv inhibitor:25 μ L samples to be tested is taken to be managed in 1.0mL EP (Eppendorf),
50 μ L are subsequently added into, a concentration of 10mU/mL dipeptidyl peptidase-IVs (Sigma, the U.S.), mixing, 37 DEG C incubate 15min, add
Glycyl-prolyl-p-nitrophenyl amine hydrochlorate (Sigma, the U.S.) containing 0.8mmol/L in 25 μ L, mixing, 37 DEG C anti-
Answer 30min, be eventually adding 100 μ L, 1mol/L, pH 4.0 sodium acetate buffer terminate reaction.4 DEG C, 10,000rpm from
Heart 2min takes 200 μ L supernatants in 96 hole microwell plates (Greiner bio-one, Germany), Spectra Max M5 multifunctional enzyme marks
Instrument (Molecular Devices, the U.S.) measures absorbance value under 405nm wavelength.Sample to be tested is to dipeptidyl peptidase-IV
Inhibitory activity, as shown by the following formula:
Wherein:Negative control group substitutes sample to be tested for blank skimmed milk;
Negative blank group:Three (hydroxymethyl) aminomethane hydrochloride buffer (Tris-HCl, the pH of concentration 100mmol/L
=8.0) dipeptidyl peptidase-IV in negative control group is substituted;
Sample blank group substitutes the dipeptidyl peptidase-IV in sample to be tested group for Tris-HCl.
By the inoculum concentration in embodiment 1, skimmed milk concentration, the speed that fermentation time, fermentation are vibrated is adjusted one by one, is obtained
The leben product that following set of distinct methods are prepared was obtained, acidified milk is drunk obtained by each group measured using the above method
Material is as shown in table 1 to the inhibition of dipeptidyl peptidase-IV.
1. distinct methods of table prepare the inhibition of gained leben
It can be obtained from the result shown in table 1, by inoculum concentration in the preparation method for preparing the leben product,
Skimmed milk concentration, cultivation temperature, when the speed that fermentation time, fermentation are vibrated is adjusted to except preferred scope, mostly viscous class buds
The spore bacillus CGMCC No.10062 skimmed milks that can still ferment generate dipeptidyl peptidase-iv inhibitor, but the dipeptides of product
Base peptase-IV inhibitions are decreased obviously.
Parallel test may be repeated several times using the above method, finally obtain currently preferred fermentation parameter range it
When outer, Paenibacillus polymyxa CGMCC No.10062 fermentation skimmed milks, the inhibition effect of obtained dipeptidyl peptidase-iv inhibitor
Fruit is decreased obviously.For example, when inoculum concentration is very few, culture propagation speed is slow so that the dipeptidyl peptidase-IV finally obtained inhibits
The activity of agent is less than preferred inoculum concentration range;When inoculum concentration is excessive, strain individual is because of living space and battalion after a certain period of time
Foster limitation and dead or growth arrest, are unfavorable for the accumulation of metabolin, but also the dipeptidyl peptidase-IV finally obtained inhibits
The activity of agent is less than preferred inoculum concentration range.In another example when fermentation temperature is less than preferred scope value, strain metabolism is slow,
The activity of dipeptidyl peptidase-iv inhibitor can be caused to decrease.Further for example, when the hunting speed of fermentation is less than preferred scope,
The dissolved oxygen amount of strain fermentation is inadequate, and growth and metabolism are restricted, and leads to the activity of dipeptidyl peptidase-iv inhibitor to reduce.
The leben that the present invention obtains is to the measure of dipeptidyl peptidase-IV inhibiting rate:It will in the diluted method of sesquialter
Testing sample solution is diluted, and obtains the sample to be tested group of various concentration, and various concentration is measured using above-mentioned detection method
Sample to be tested is to the inhibiting rate of dipeptidyl peptidase-IV, according to the linear relationship that concentration and dipeptidyl peptidase-IV inhibiting rate are formed,
Inhibiting rate of the sample to dipeptidyl peptidase-IV is calculated.It is detected into after parallel test for several times, obtains the acidified milk drink of the present invention
Expect inhibiting rate >=75% to DPP-IV.
Effect example 1:Product taste is tested with fancy grade
Leben product A, B, C prepared by above-described embodiment 1-3 is taken to be dissolved in warm water, is obtained a concentration of
The leben solution of 100mg/mL as experimental subjects, carries out the taste tests of product.50 people of test number.It tastes
Mode:It is tasted by the way of blank marking;Respectively to color and luster, flavor, the mouth of above-mentioned leben product A, B, C
Sense, nutrition item are individually given a mark, and each single item full marks are 25 points, calculate average mark and its total score, statistical result are recorded in table 2.
Meanwhile the opinion provided according to the whole fancy grade to product, count the hobby number to each single product, statistical result record
In table 3.
2. product taste tests result data statistical form of table
Leben product | A | B | C |
Color and luster | 23 | 22 | 24 |
Flavor | 23 | 24 | 21 |
Mouthfeel | 21 | 22 | 23 |
Nutrition | 24 | 22 | 24 |
Total score | 91 | 90 | 92 |
3. product fancy grade test result data statistical form of table
Fancy grade | A | B | C |
Like | 44 | 38 | 47 |
Well | 4 | 7 | 2 |
Generally | 2 | 4 | 1 |
It does not like | — | 1 | — |
It can be seen that in general from product taste tests and fancy grade statistical result, pass through technical solution of the present invention
In method obtained by the leben with dipeptidyl peptidase-IV inhibitory activity in terms of product special flavour, mouthfeel, nutrition
It can be received by most of consumer.
Effect example 1:The stability of dipeptidyl peptidase-iv inhibitor inhibition under refrigerated condition
After product A, B, C prepared by embodiment 1-3 are respectively charged into hermetic bag, be placed in (8 DEG C) of refrigerated condition preserve 0,
10th, it is taken out after 20 and 30d, measures 503nhibiting concentration IC of each sample to dipeptidyl peptidase-IV respectively50Value, as a result such as 4 institute of table
Show.
The stability of product inhibition under 4. refrigerated condition of table
As shown in Table 4, the dipeptidyl peptidase-iv inhibitor product of all tests is after (8 DEG C) preservation 30d of refrigeration, to two
Peptidyl peptidase-IV inhibitory activity is maintained at same level, i.e. stability is preferable.
Effect example 2:
1 the method for reference implementation example, compares by Paenibacillus polymyxa CGMCC No.10062, Lactobacillus casei
(L.casei) ATCC 393 (buying from ATCC), lactobacillus bulgaricus (L.bulgaricus) LB340 are (by Denis section public affairs
Department provide), streptococcus thermophilus (S.thermophilus) ST-BODY-3 (being provided by Hansen Corp. of section) prepare leben
To the inhibition of dipeptidyl peptidase-IV, concrete operations are as follows:
1st, materials and methods
(a) preparation of seed (fermenting microbe):
The preparation of Lactobacillus casei and lactobacillus bulgaricus seed:By Lactobacillus casei ATCC 393 and bulgarian milk
The freeze-dried powder of bacillus LB340 is dissolved respectively with a small amount of sterile distilled water, and each personal oese takes a ring to line MRS solid cultures
On base (Merck, Germany), 37 DEG C of Anaerobic culturels take out for 24 hours, with oese picking single bacterium colony be put into 1mL MRS liquid (Merck,
Germany), bacterium colony is dispersed in fluid nutrient medium with vortex oscillator, 37 DEG C of Anaerobic culturels take out for 24 hours, with 2% (v/
V) inoculum concentration is inoculated in 50mL MRS liquid, and after 37 DEG C of cultures for 24 hours, culture 9,000rpm is centrifuged 10 minutes, is discarded supernatant, bacterium
After body washs 2 times with sterile distilled water, suspended with the sterile distilled water of former volume of culture, obtain the seed of corresponding fermentation.
The preparation of streptococcus thermophilus seed:The freeze-dried powder of streptococcus thermophilus ST-BODY-3 is molten with a small amount of sterile distilled water
Solution takes a ring to line on M17 solid mediums (Merck, Germany) with oese, and 40 DEG C of Anaerobic culturels take out for 24 hours, with inoculation
Ring picking single bacterium colony is put into 1mL M17 liquid (Merck, Germany), and bacterium colony is dispersed in liquid training with vortex oscillator
It supports in base, 40 DEG C of Anaerobic culturels take out for 24 hours, and 50mL M17 liquid is inoculated in 2% (v/v) inoculum concentration, after 40 DEG C of cultures for 24 hours,
9,000 rpm of culture centrifuges 10min, discards supernatant, after thalline washs 2 times with sterile distilled water, with the nothing of former volume of culture
Bacterium distills aqueous suspension, obtains the seed of fermentation.
(b) preparation of skimmed milk:With embodiment 1.
(c) prepared by sample to be tested:With embodiment 1.
2nd, the preparation of leben
By each bacterial strain with final concentration of 8.0x106Cfu/ml inoculum concentrations aseptic inoculation is trained respectively in 8% (w/w) skimmed milk
Support (37 DEG C of Anaerobic culturels of lactobacillus bulgaricus and Lactobacillus casei, 40 DEG C of Anaerobic culturels of streptococcus thermophilus, mostly viscous class gemma bar
30 DEG C of bacterium, 220rpm shaken cultivations) 4d, obtain corresponding leben.
3rd, the measure of dipeptidyl peptidase-IV inhibitory activity
Leben prepared by above-mentioned different strains is to the 503nhibiting concentration IC of dipeptidyl peptidase-IV50As shown in table 5:
Leben prepared by 5 different strains of table is to the 503nhibiting concentration IC of dipeptidyl peptidase-IV50
As shown in Table 5, the product of other conventional skim-milk fermentation strain fermentation skimmed milks does not have dipeptidyl peptidase-IV
Inhibitory activity, and the inhibition of leben prepared by Paenibacillus polymyxa CGMCC No.10062 to dipeptidyl peptidase-IV
Active highly significant.
The foregoing is only a preferred embodiment of the present invention, is not intended to restrict the invention, for the skill of this field
For art personnel, the invention may be variously modified and varied.All within the spirits and principles of the present invention, that is made any repaiies
Change, equivalent replacement, improvement etc., should all be included in the protection scope of the present invention.
Claims (11)
1. a kind of leben, which is characterized in that the raw material of the beverage include Paenibacillus polymyxa, skimmed milk, sweet taste
Agent and/or acid, the deposit number of the Paenibacillus polymyxa is CGMCC No.10062, and Paenibacillus polymyxa is being sent out
The final concentration of 2x10 of kefir milk beverage6~2x107cfu/mL。
2. leben according to claim 1, which is characterized in that the sugariness of the leben is converted into sucrose
Sugariness is 9%~13%.
3. leben according to claim 1, which is characterized in that the titratable acidity of the leben is 65 ° of T
~75 ° of T.
4. leben according to claim 1, which is characterized in that the leben is to dipeptidyl peptidase-IV
Inhibiting rate >=75%.
5. leben according to claim 1, which is characterized in that the skimmed milk includes skimmed milk powder and water, takes off
Fat milk powder accounts for the mass percent 5%~10% of skimmed milk.
A kind of 6. method for preparing any one leben of claim 1-5, which is characterized in that this method is included such as
Lower step:
(a) Paenibacillus polymyxa that deposit number is CGMCC No.10062 is inoculated in skimmed milk and fermented, fermented
After obtain acidified milk;
(b) it blends to form blending liquid using sweetener and/or acid, by acidified milk mixing made from blending liquid and step (a),
Homogeneous, sterilizing, obtains leben.
7. the method according to claim 6 for preparing leben, which is characterized in that ferment in the step (a)
Temperature is 25 DEG C~35 DEG C, and the time of fermentation is 3d~7d.
8. the method according to claim 6 for preparing leben, which is characterized in that in step (a), the fermentation is
Shaken cultivation, hunting speed are 125rpm~220rpm.
9. the method according to claim 6 for preparing leben, which is characterized in that in step (b), the acidified milk
Mixed volume ratio with blending liquid is 1:1~1:4.
10. the method according to claim 6 for preparing leben, which is characterized in that the pressure of homogeneous in step (b)
For 15~30MPa.
11. the method according to claim 6 for preparing leben, which is characterized in that the temperature to sterilize in step (b)
It it is 95 DEG C~125 DEG C, the time of sterilizing is 5min~20min.
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