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Probiotics compound for treating inflammatory bowel disease

A technology of inflammatory bowel disease and probiotics, applied in the field of probiotic drugs, can solve the problem of low utilization rate of EcN, achieve the effect of improving colonic inflammation and promoting expression

Active Publication Date: 2019-05-17
CHENGDU MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, after the microencapsulated EcN was left standing for 1 hour in an environment with a pH value of 2.5, only one-thousandth of the bacteria could survive; if it was placed in gastric acid with lower pH and containing pepsin, There will be fewer bacteria that can survive, and the utilization rate of EcN is extremely low

Method used

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  • Probiotics compound for treating inflammatory bowel disease
  • Probiotics compound for treating inflammatory bowel disease
  • Probiotics compound for treating inflammatory bowel disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] Embodiment 1 The preparation method of EcN complex of the present invention

[0066] 1. Cell activation

[0067] Mix 0.5 mL of EcN and 0.5 mL of 50% glycerol in a 1.5 mL centrifuge tube and store at -80°C. When cultivating bacteria, streak the plate on LB solid medium, cultivate overnight in a 37°C incubator, then pick colonies and inoculate them in LB liquid medium, shake overnight at 37°C and 220rpm / min, and then press the ratio of 1:100 To expand the culture, place it on a shaker at 37°C and 220rpm.

[0068] 2. Chitosan-sodium alginate coating

[0069] Select 20 mL of EcN bacterial solution in the logarithmic growth phase, centrifuge at 4000 rpm at 4 °C for 5 min in a refrigerated centrifuge, discard the supernatant, wash twice with 5 mL of PBS, centrifuge at 4000 rpm for 3 min, discard the supernatant, and resuspend the bacteria in 4 mL of PBS.

[0070] (1) Chitosan encapsulation: Add 0.5mL 1mg / ml chitosan solution to the resuspension, shake up and down for 10s, ...

Embodiment 2

[0077] EcN living bacteria growth logarithmic phase judgment before embodiment 2 wrapping

[0078] Before wrapping, ECN was inoculated in LB medium, cultured on a shaker at 37°C and 220r / min, and the OD value of the bacterial solution was detected at different times, and the bacteria in the logarithmic growth phase were selected for wrapping. The vertical axis in the figure represents the OD value, The abscissa is the time of cultivation, and the specific steps are as follows:

[0079] 1) Pick a single colony from the plate and inoculate it in LB liquid medium, and culture it in a shaker at 37°C and 220r / min until OD 600nm The value is around 0.6.

[0080] 2) Transplantation of EcN

[0081] Take 15 25ml test tubes, add 6mL LB liquid medium to each tube, and cultivate the bacterial solution to OD at a ratio of 1:200 600nm After the value was about 0.6, they were respectively inoculated into 14 test tubes, and 1 test tube was not inoculated, which was used as a negative contr...

experiment example 1ZE

[0087] Experimental example 1 ZETA potential detection

[0088] Use a ξ potential detector to detect the charge of each layer in the process of layer-by-layer wrapping 0, 1, 2, 3, 4, and 5, take the wrapped bacterial solution, and after dilution, use a ξ potential detector to detect the charge of each layer in the layer-by-layer wrapping process. The charge of the layer, according to the positive and negative charge ratio of each layer, judges its wrapping situation, and draws it into a graph.

[0089] like image 3 As shown, during the encapsulation process, positively charged chitosan will combine with ECN, and the negative charge on the surface of the bacterium will decrease, and a layer will be wrapped. When negatively charged sodium alginate is added, the negative charge on the surface of the bacterium will There is an increase, and the second layer is wrapped, and the fluctuation of the zeta potential represents the number of wrapped layers.

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Abstract

The invention provides a probiotics compound for treating inflammatory bowel disease. The probiotics compound is obtained in a manner that escherichia coli Nissle 1917(EcN) is alternately wrapped withchitosan and sodium alginate, and then calcium ions are cross-linked with the sodium alginate located at the outermost layer; the total number of wrapping layers is the even number larger than 2. Bymeans of the probiotics compound for treating inflammatory bowel disease, the activity of the EcN is kept while gastric acid and mechanical friction can be resisted, and the anti-inflammation functionof the EcN in the digestive tract is developed; the defect that the EcN is easily inactivated by the entogastric environment is overcome, a preparing method is simple, and the probiotics compound hasquite-excellent application value.

Description

technical field [0001] The invention relates to the field of probiotic drugs, in particular to a probiotic compound for treating inflammatory bowel disease. Background technique [0002] Escherichia coli Nissle 1917 (EcN) is a kind of probiotics derived from the human intestinal tract. It is widely used to prevent infectious diarrhea, inflammatory bowel disease and prevent the colonization of pathogenic bacteria in the digestive tract of newborns; its use is mainly oral. [0003] In the treatment of intestinal diseases, the activity of orally administered EcN has been greatly reduced when it reaches the intestinal lesions because it is easily eroded by gastric acid and squeezed and rubbed by the digestive tract, and its therapeutic effect is very limited. [0004] Protective microcapsules composed of sodium alginate and chitosan have been reported. The microcapsules are usually coated with bacteria by a layer of sodium alginate, cross-linked calcium ions, and then coated wit...

Claims

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Application Information

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IPC IPC(8): A61K9/50A61K35/74A61K47/36A61P1/00
CPCY02A50/30
Inventor 宋海星罗晓明
Owner CHENGDU MEDICAL COLLEGE
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