A method for large-scale fermentation of ferritin

A technology of ferritin and fermented liquid, which is applied in the field of biomedical engineering, can solve the problems of low bacterial growth rate, low drug loading, and difficult purification of ferritin, and achieve the improvement of fermentation efficiency and entrapment effect, fermentation efficiency and Improvement of entrapment effect, improvement of fermentation efficiency and effect of entrapment effect

Active Publication Date: 2020-11-20
KUNSHAN XINYUNDA BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] In addition, due to the influence of a series of factors in the fermentation process, the purification of the prepared ferritin is difficult and expensive, resulting in problems such as low drug loading, many impurities, and poor stability.
For example, Chinese patent application CN1257931A discloses a ferritin that can be used for iron supplementation in the human body and its preparation method and application, but the growth rate of the bacteria in the fermentation process is low, and the prepared ferritin is a bacterial agent for iron supplementation, including drug loading is less effective

Method used

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  • A method for large-scale fermentation of ferritin
  • A method for large-scale fermentation of ferritin
  • A method for large-scale fermentation of ferritin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] (1) Seed culture: Take out a frozen seed from the -80°C refrigerator (Escherichia coli BW25113 containing the human HFn coding gene, which is obtained by inserting the human HFn coding gene sequence, that is, the nucleotide sequence (sequence 1) into the plasmid pSB1S Nco I / Xho I site construction, obtained by transforming Escherichia coli BW25113), thawed at room temperature, inoculated 1 mL of seeds into an Erlenmeyer flask of LB medium containing 40 mg / L streptomycin sulfate at 0.5% (v / v) ( 500mL) to obtain the seed liquid, when the OD of the seed liquid 600 At 1.6, the seed solution was inoculated in the fermenter containing the fermentation medium by the flame inoculation method at 0.5% inoculation amount.

[0071] The fermentation medium is composed of the following components: yeast peptone 14g / L, yeast extract 10g / L, sodium chloride 0.5g / L, ammonium chloride 0.5g / L, dodecahydrate disodium hydrogen phosphate 10g / L, Potassium dihydrogen phosphate 1g / L, magnesium...

Embodiment 2

[0077] (1) Seed culture: Take out a frozen seed from the -80°C refrigerator, thaw it at room temperature, and inoculate 1 mL of the seed at 3% (v / v) in the cone of LB medium containing 60 mg / L streptomycin sulfate. bottle (500mL) to get the seed solution, when the OD of the seed solution 600 At 2.2, the seed solution was inoculated in the fermenter containing the fermentation medium by the flame inoculation method at 1.5% inoculation amount.

[0078] The fermentation medium is composed of the following components: yeast peptone 18g / L, yeast extract 14g / L, sodium chloride 2g / L, ammonium chloride 2g / L, dodecahydrate disodium hydrogen phosphate 14g / L, diphosphate Potassium hydrogen 4g / L, magnesium sulfate heptahydrate 1g / L and glucose 16g / L.

[0079] (2) Fermentation culture:

[0080] Maintain the dissolved oxygen concentration (DO) of the fermentation medium at 35%, and ferment at pH 7.3 and temperature 40°C; when OD 600 At 55 o'clock, 10 g / L arabinose was added to start the ...

Embodiment 3

[0084] (1) Seed culture: Take out a frozen seed from the -80°C refrigerator, thaw it at room temperature, and inoculate 1 mL of the seed at 1% (v / v) in the cone of LB medium containing 50 mg / L streptomycin sulfate. bottle (500mL) to get the seed solution, when the OD of the seed solution 600 At 1.9, the seed liquid was inoculated in the fermenter containing the fermentation medium by flame inoculation method according to the inoculum amount of 1%.

[0085] The fermentation medium is composed of the following components: yeast peptone 20g / L, yeast extract 16g / L, sodium chloride 1.2g / L, ammonium chloride 1.2g / L, dodecahydrate disodium hydrogen phosphate 16g / L, Potassium dihydrogen phosphate 2g / L, magnesium sulfate heptahydrate 0.5g / L and glucose 16g / L.

[0086] (2) Fermentation culture:

[0087] Maintain the dissolved oxygen concentration (DO) of the fermentation medium at 32%, and ferment at pH 6.8 and temperature 38°C; when OD 600 At 48 o'clock, 5 g / L arabinose was added to...

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Abstract

The invention relates to the technical field of biomedical engineering, and particularly relates to a method for large-scale fermentation of ferritin. The method comprises the steps that a fermentation medium without microelements is used, and fermentation conditions are optimized. The method is high in fermentation efficiency and low in cost, and the prepared ferritin is good in drug loading effect and uniform and stable in quality.

Description

technical field [0001] The invention relates to the technical field of biomedical engineering, in particular to a method for large-scale fermentation of ferritin. Background technique [0002] Ferritin is a kind of natural iron storage protein that widely exists in animals, plants and microorganisms. Ferritin isolated from humans or other mammals is usually composed of two different ferritin subunits (H subunit and L subunit), with molecular masses of 21 kDa and 19 kDa, respectively. A typical ferritin structure is a cage-like structure formed by self-assembly of 24 H subunits / L subunits, with an outer diameter of 12 nm and an inner cavity structure with a diameter of 8 nm. The cage-like structure houses iron nuclei containing up to 4500 iron atoms. [0003] Since ferritin has a cage-like structure that can encapsulate drugs, remarkable stability, and small and uniform size, attempts have been made in the field to use ferritin as a drug carrier to deliver drugs, for exampl...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12P21/00C07K14/47C12R1/19
CPCC07K14/47C12N1/20
Inventor 柯天一姚德惠劳芳张泽译希伦丁凤姣王晓飞
Owner KUNSHAN XINYUNDA BIOTECH CO LTD
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