CN1935981A - Method for increasing acid resistance of plant lactobacillus - Google Patents
Method for increasing acid resistance of plant lactobacillus Download PDFInfo
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- CN1935981A CN1935981A CN 200610028708 CN200610028708A CN1935981A CN 1935981 A CN1935981 A CN 1935981A CN 200610028708 CN200610028708 CN 200610028708 CN 200610028708 A CN200610028708 A CN 200610028708A CN 1935981 A CN1935981 A CN 1935981A
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- acid
- plant lactobacillus
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Abstract
The invention relates to the method used to improve lactobacillus plantarum acid resistance. It adopts lactobacillus plantarum LP-Onlly, adds weak acid or weak super into fermentation medium to increase lactobacillus plantarum cell acid resistance. The invention provides operation significance method for increasing lactobacillus plantarum acid resistance. Thus it can improve product quality, prolong product shelf life.
Description
Technical field
The present invention relates to plant lactobacillus, the acid proof method of particularly a kind of raising plant lactobacillus.
Background technology
Milk-acid bacteria is a most important probiotic bacterium in the human intestinal, and its quantity and composition play keying action to keeping healthy intestinal environment and improving immunologic function.It mainly acts on and is to suppress pernicious bacteria, keep the intestinal microflora balance, protection intestinal mucosal barrier, metabolism produce organic acid, reduces enteron aisle pH, promotes intestines peristalsis, diarrhoea constipation dual regulation.
Milk-acid bacteria all can face various environmental stresss producing, preserve and being taken in the process, comprises the influence of mal-conditions such as temperature, acid, osmotic pressure, oxygen, high pressure and hunger.These factors all can influence the survival of bacterium, thereby influence its physiologically active.Though milk-acid bacteria self has stronger acid resistance, most of can the growth under the condition of pH3 just shows susceptibility under the condition below the pH3, and along with the further increase of acidity, mortality ratio can sharply rise.Therefore improving milk-acid bacteria is of great practical significance to the tolerance of environment acid fracturing power.
At present, domestic and international raising method to the acidproof ability of milk-acid bacteria mainly concentrates in the screening of acidproof bacterial strain.Employings such as river and mountain simulations human body gastric acid environment (pH=3) method screen from 104 strain human gastrointestinal tract source Bacterium lacticum and is obtained 3 strains and have acid resistance bacterial strain by force.
Summary of the invention
The purpose of this invention is to provide the acid proof method of a kind of raising plant lactobacillus, it will provide the method for a tool realistic meaning for the milk-acid bacteria acid resistance, thereby improve product quality, prolong the product shelf-life.
For solving the problems of the technologies described above, the present invention is achieved in that
The acid proof method of a kind of raising plant lactobacillus, it is characterized in that: this plant lactobacillus is a plant lactobacillus LP-Onlly, adds weak acid or salt of weak acid to improve plant lactobacillus cell acid resistance in fermention medium.
This plant lactobacillus on December 6th, 2004 at China Committee for Culture Collection of Microorganisms common micro-organisms center, be called for short: CGMCC (address: No. 13, one of Zhong Guan-cun, Haidian District, BeiJing, China city, Institute of Microorganism, Academia Sinica) preservation, and receive preservation registration number CGMCC NO 1258.Its called after of classifying: plant lactobacillus, latin name: Lactobacillusplantarum.Plant lactobacillus LP-Onlly, its 16S rRNA gene GeneBank (NCBI) sequence accession number is AY590777.
The acid proof method of described raising plant lactobacillus is characterized in that: add 20～80mmol/L acetate in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add 50mmol/L acetate in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add 10～20mmol/L propionic acid in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add the 10mmol/L propionic acid in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add 10～20mmol/L butanic acid in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add the 20mmol/L butanic acid in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add 20～80mmol/L potassium acetate in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add the 50mmol/L potassium acetate in fermention medium.
The acid proof method of described raising plant lactobacillus is characterized in that: add 2mmol/L Tripotassium Citrate or 3mmol/L Potassium tripolyphosphate in fermention medium.
Description of drawings
Fig. 1 be in the inventive method acetate to the acid proof situation synoptic diagram that influences of plant lactobacillus.
Fig. 2 be in the inventive method propionic acid to the acid proof situation synoptic diagram that influences of plant lactobacillus.
Fig. 3 be in the inventive method butanic acid to the acid proof situation synoptic diagram that influences of plant lactobacillus.
Fig. 4 be in the inventive method potassium acetate to the acid proof situation synoptic diagram that influences of plant lactobacillus.
Fig. 5 be in the inventive method Tripotassium Citrate to the acid proof situation synoptic diagram that influences of plant lactobacillus.
Fig. 6 be in the inventive method Potassium tripolyphosphate to the acid proof situation synoptic diagram that influences of plant lactobacillus.
Embodiment:
The invention provides the acid proof method of a kind of raising plant lactobacillus, main technical schemes is exactly to add weak acid or salt of weak acid to improve plant lactobacillus cell acid resistance in fermention medium.The particular case enumeration is as follows:
1. bacterial classification
Plant lactobacillus LP-Onlly (Lactobacillus plantarum LP-Onlly) is for research institute of Only Co., Ltd., Shanghai Jiantong Univ. separates from healthy babies ight soil, identifies and preserves.
2. substratum
Comprise that slant medium, seed culture medium and fermention medium are the MRS substratum.
3. cultural method
First order seed is cultivated: choose two ring bacterium to seed culture fluid from slant medium, the bottled liquid 30ml of 50ml triangle cultivates 16h for 37 ℃.Secondary seed is cultivated: with primary seed solution with ten thousand/ inoculum size insert in the seed culture fluid, the bottled liquid 30ml of 50ml triangle cultivates 16h for 37 ℃.Fermentation culture: with secondary seed solution with ten thousand/ inoculum size insert in the fermentation culture, the bottled liquid 30ml of 50ml triangle cultivates 16h for 37 ℃.
4. analytical procedure
(1) acid resistance evaluation method: the cell that will ferment with the physiological saline washed twice after in pH2,37 ℃ of following 90min, survey the cell survival rate.
(2) live bacterial count method: referring to GB/T 4789.35-2003 (the milk-acid bacteria check in the lactobacillus drink).
Embodiment 1: add acetate in fermention medium.
As shown in Figure 1: acetic acid concentration (10mmol/L) pair cell acid resistance contrast does not improve; And acetic acid concentration is when 10mmol/L is increased to 50mmol/L, and cell acid resistance contrast has just had very large raising, improves 200 times approximately at pH2,37 ℃ of following 90min cell survival rates; Further increase acetic acid concentration (being increased to 80mmol/L) cell acid resistance and obviously reduce, but contrast is also improved a lot from 50mmol/L.
Embodiment 2: add propionic acid in fermention medium.
As shown in Figure 2: the propionic acid concentration of 10mmol/L can make the cell acid resistance obtain bigger raising, contrasts and improves 90 times approximately at pH2,37 ℃ of following 90min cell survival rates; Propionic acid concentration when 10mmol/L is increased to 20mmol/L, cell acid resistance basically identical; Further increase propionic acid concentration (being increased to 50mmol/L) cell acid resistance and sharply reduce from 20mmol/L, at this moment, suitable with contrast.
Embodiment 3: add butanic acid in fermention medium.
As shown in Figure 3: the butanic acid concentration of 10mmol/L makes the cell acid resistance obtain bigger raising, and at pH2,37 ℃ of following 90min, cell survival rate contrast improves 120 times approximately; Further increase butanic acid concentration (being increased to 20mmol/L from 10mmol/L), the cell acid resistance further improves, and at pH2,37 ℃ of following 90min, cell survival rate contrast improves 200 times approximately; Increase butanic acid concentration (being increased to 50mmol/L from 20mmol/L) again, the cell acid resistance sharply reduces, and is at this moment, suitable with contrast.
Embodiment 4: add potassium acetate in fermention medium.
As shown in Figure 4: potassium acetate concentration (10mmol/L) pair cell acid resistance contrast does not improve; And acetic acid concentration is when 10mmol/L is increased to 50mmol/L, and cell acid resistance contrast has very large raising, improves 200 times approximately at pH2,37 ℃ of following 90min cell survival rates; Further increase acetic acid concentration (being increased to 80mmol/L) cell acid resistance and obviously reduce, but contrast is also improved a lot from 50mmol/L.
Embodiment 5: add Tripotassium Citrate in fermention medium.
As shown in Figure 5: the concentration of potassium citrate of 2mmol/L can make the cell acid resistance obtain bigger raising, contrasts and improves 100 times approximately at pH2,37 ℃ of following 90min cell survival rates; When concentration of potassium citrate was increased to 5mmol/L from 2mmol/L, the cell acid resistance sharply reduced; Further increase concentration of potassium citrate (being increased to 10mmol/L) from 5mmol/L, cell acid resistance basically identical, but contrast slightly improves.
Embodiment 6: add Potassium tripolyphosphate in fermention medium.
As shown in Figure 6: the tripolyphosphate potassium concn pair cell acid resistance of 1mmol/L contrast does not improve; And the tripolyphosphate potassium concn is when 1mmol/L is increased to 3mmol/L, and the cell acid resistance contrasts some raising, improves 5 times approximately at pH2,37 ℃ of following 90min cell survival rates; The tripolyphosphate potassium concn is when 3mmol/L is increased to 5mmol/L, and the cell acid resistance reduces, at this moment, with contrast quite.
The action principle of the inventive method is:
Milk-acid bacteria is acidproof to be because himself can keep the inside and outside pH gradient of cell, makes intracellular pH be unlikely to too low, but surpasses certain threshold value when inner pH drops to, and cell function will be suppressed, thereby cause necrocytosis.
As everyone knows, when cell enters stationary phase, produce σ with the closely-related gene of cell stability at the RpoS genes encoding more than 60
sInduce startup under the albumen control, therefore, cell stationary phase performance such as heat-resisting and acidproof depends primarily on σ
sConcentration, and σ
sConcentration can be by improving RpoS transcriptional level or rpos mRNA translation skill or preventing σ
sMeans such as proteolysis improve.Studies show that cAMP-CRP, UDP-glucose, EIIA, polyphosphate (polyphosphate), ppGpp, HSL, degree of ionization are the transcriptional level that NADH all can improve RpoS in 4.8～4.9 salt of weak acid (as acetate, propionic salt, benzoate etc.), Sodium Glutamate, trehalose and the born of the same parents to the transformation efficiency of NAD; And cell oozes or acid pH conversion, low temperature, growth reach certain cell density, some albumen such as Hfq, HU, DksA, RprA etc. all can promote rpos mRNA translation to height; The heat shock protein that unexpected carbon hunger, osmotic pressure raising, acid pH conversion, thermal shocking (as rising to 42 ℃ of 20min from 30 ℃) produce such as DnaK, DnaJ, GrpE all can make σ
sStable; CIpXP proteolytic enzyme in the RssB-CIpXP approach is responsible for σ
sHydrolysis, and Crl albumen can promote σ
sCore enzyme (E) combination with RNA polymerase makes σ
sAvoid combining and be hydrolyzed, thereby improve σ with RssB
sActive.Therefore, can improve σ in the cell by above-mentioned means
sConcentration, thus the cell acid resistance improved.
The inventive method is by adding weak acid (acetate, propionic acid and butanic acid) or salt of weak acid (potassium acetate, Tripotassium Citrate and Potassium tripolyphosphate) all can improve plant lactobacillus cell acid resistance to some extent in fermention medium; In addition, acetate and acetate be to plant lactobacillus acid resistance raising effect basically identical, and this shows that the acid proof influence of salt of weak acid pair cell works by its weak acid, and does not have what relation with its metal ion.The inventive method will provide the method for a tool realistic meaning for the milk-acid bacteria acid resistance, thereby improve product quality, prolong the product shelf-life.
Claims (10)
1. one kind is improved the acid proof method of plant lactobacillus, and it is characterized in that: this plant lactobacillus is a plant lactobacillus LP-Onlly, adds weak acid or salt of weak acid to improve the acid resistance of plant lactobacillus cell in fermention medium.
2. the acid proof method of raising plant lactobacillus according to claim 1 is characterized in that: add 20～80mmol/L acetate in fermention medium.
3. the acid proof method of raising plant lactobacillus according to claim 2 is characterized in that: add 50mmol/L acetate in fermention medium.
4. the acid proof method of raising plant lactobacillus according to claim 1 is characterized in that: add 10～20mmol/L propionic acid in fermention medium.
5. the acid proof method of raising plant lactobacillus according to claim 4 is characterized in that: add the 10mmol/L propionic acid in fermention medium.
6. the acid proof method of raising plant lactobacillus according to claim 1 is characterized in that: add 10～20mmol/L butanic acid in fermention medium.
7. the acid proof method of raising plant lactobacillus according to claim 6 is characterized in that: add the 20mmol/L butanic acid in fermention medium.
8. the acid proof method of raising plant lactobacillus according to claim 1 is characterized in that: add 20～80mmol/L potassium acetate in fermention medium.
9. the acid proof method of raising plant lactobacillus according to claim 8 is characterized in that: add the 50mmol/L potassium acetate in fermention medium.
10. the acid proof method of raising plant lactobacillus according to claim 1 is characterized in that: add 2mmol/L Tripotassium Citrate or 3mmol/L Potassium tripolyphosphate in fermention medium.
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|---|---|---|---|
| CN 200610028708 CN1935981A (en) | 2006-07-07 | 2006-07-07 | Method for increasing acid resistance of plant lactobacillus |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102080058A (en) * | 2010-12-09 | 2011-06-01 | 江南大学 | Lactobacillus casei and breeding method thereof |
| CN113278554A (en) * | 2021-05-24 | 2021-08-20 | 四川大学 | Method for improving acid resistance of lactic acid bacteria by using mixed bacteria biological membrane |
-
2006
- 2006-07-07 CN CN 200610028708 patent/CN1935981A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102080058A (en) * | 2010-12-09 | 2011-06-01 | 江南大学 | Lactobacillus casei and breeding method thereof |
| CN113278554A (en) * | 2021-05-24 | 2021-08-20 | 四川大学 | Method for improving acid resistance of lactic acid bacteria by using mixed bacteria biological membrane |
| CN113278554B (en) * | 2021-05-24 | 2023-05-09 | 四川大学 | Method for improving acid resistance of lactic acid bacteria by using mixed bacteria biological film |
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Open date: 20070328 |