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Gelatin composition, and preparation method and application thereof

A composition and gelatin technology, applied in the field of medicine, can solve problems such as poor hemostatic effect, and achieve the effects of strong functionality, low antigenicity and allergenicity, and enhanced elasticity

Active Publication Date: 2013-08-28
李肯
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

As an enzyme preparation, MTG is easy to live at room temperature, so it must be stored at low temperature. However, during the operation, it will be taken out of the low temperature immediately and used directly. Ordinary MTG cannot fully exert its activity under low temperature conditions, resulting in poor hemostatic effect.

Method used

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  • Gelatin composition, and preparation method and application thereof
  • Gelatin composition, and preparation method and application thereof
  • Gelatin composition, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1 genetic engineering method prepares microbial transglutaminase (MTG)

[0034] The wild MTG strain gene is shown in sequence 1 (SEQ ID NO.1), and the MTG strain gene used in the present invention is shown in sequence 2 (SEQ ID NO.2). G at position 73 is changed to T, and T at position 750 is changed to g. The wild MTG amino acid sequence is shown in sequence 3 (SEQ ID NO.3), the MTG amino acid sequence used in the present invention is shown in sequence 4 (SEQ ID NO.4), the 25th Gly mutation is Cys, and the 250th Ser mutation for Arg. Design and commission a biotechnology company (such as Beijing Quanshijin Biotechnology Co., Ltd., Shanghai Bioengineering Company) to synthesize the MTG strain gene of the sequence shown in SEQ ID NO.2, and then clone it into the EcoRV of the Escherichia coli expression vector pET-32 site, and then transfer the expression vector to Escherichia coli BL21 to form engineering bacteria. The genetically engineered bacteria were c...

Embodiment 2

[0035] The fine purification of embodiment 2 microbial transglutaminase MTG

[0036] The supernatant obtained in Example 1 was filtered with a 0.22 μm filter membrane, and the filtrate was collected. Through a strong cation exchange column, the ion exchange column is GE's prepacked HiPrep16 / 10SP XL column, the flow rate is 2ml / min, and the sample is equilibrated with 50mM pH6.0 phosphate buffer before passing through the ion exchange column. The elution is 0-1M sodium chloride linear gradient elution, the elution peak is MTG at 100mM sodium chloride, the peak is collected, and then passed through the gel filtration column HiLoad16 / 60Superdex, the flow rate is 1ml / min, and 50mM pH6.0 Phosphate buffer eluted and the peaks were collected.

[0037] The purified MTG was tested for enzyme activity at different temperatures, and the experimental results were as follows: figure 1 as shown, figure 1 It shows that the optimal reaction temperature of the microbial transglutaminase of ...

Embodiment 3

[0039] The fermentation of embodiment 3 microbial transglutaminase MTG

[0040] Strain: Streptoverticillium mobaraense obtained by mutagenesis screening, the result obtained by sequencing is identical to the sequence of SEQ ID NO.1.

[0041] Mutagenesis method: Add 10ml of cold sterile seed medium to the eggplant bottle slant medium (Gaoshi No. 1 medium), scrape the surface mycelia fully with an inoculation needle, and pour it into a bottle containing 20-30 glass beads medium, 30°C, 200r / min shaking culture for 1h, so that the spores are in the state of germination. The operation is carried out under red light or dark conditions, and the ultraviolet light is turned on 0.5h in advance to stabilize the light source. Take 5 ml of the spore suspension diluted to 10 times and place it in a sterilized petri dish with a diameter of 9 cm and a magnetic stirrer for ultraviolet mutagenesis. The mutagenesis conditions are as follows: the power of the ultraviolet lamp is 15W, the irradi...

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Abstract

The invention provides a gelatin composition. The gelatin composition comprises hydroxylated gelatin, microbial glutamine transaminase and protein glutaminase, wherein the hydroxylated gelatin is 20-60%proline-hydroxylated gelatin. The gelatin composition has an extremely-good bleeding stopping effect at a temperature in a range of 4-10DEG C. The invention also discloses a preparation method of the gelatin composition and an application of the gelatin composition.

Description

technical field [0001] The invention belongs to the technical field of medicine, and relates to a gelatin composition, which includes hydroxylated gelatin, microbial transglutaminase and protein glutaminase, and the composition can be used as a hemostatic agent, and is used for prepackaging of hard gelatin capsules preparation. Background technique [0002] Gelatin has been widely used in the preparation of capsules and hemostasis in the prior art, such as gelatin composition (Chinese patent application number: 98811268.X), soft gelatin capsule preparation (Chinese patent application number: 200780010679.0), Surgiflo of Johnson & Johnson Medical Equipment Co., Ltd. Fluid gelatin, chitosan gelatin (Chinese patent application number: 200310121182.X), modified gelatin (Chinese patent application number: 2009102444834.6) and other gelatins for hemostasis, such as Chinese patents 200980131973.6, 200780051215.4, 201110365186.7, etc.; US patent 6,652,840 , 8,133,484, 6,194,138, et...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L15/32A61L15/38A61K47/42C12P21/00
Inventor 易正芳裴正培丛晓楠吕方刘明耀
Owner 李肯
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