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Method for in vitro screening oyster active peptides with improved memory

An in vitro screening and active peptide technology, which is applied in the screening of compounds, biochemical equipment and methods, and the determination/testing of microorganisms, etc., to achieve the effect of improving memory

Active Publication Date: 2020-07-17
INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

With the in-depth research of modern medicine, it has been found that oysters have a variety of biological activities, such as antibacterial, anti-oxidation, lowering blood pressure, anti-tumor, improving immunity, anti-aging, and cardiovascular protection. Little research has been done on memory

Method used

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  • Method for in vitro screening oyster active peptides with improved memory
  • Method for in vitro screening oyster active peptides with improved memory
  • Method for in vitro screening oyster active peptides with improved memory

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Preparation of oyster active peptide:

[0029] Take 10 parts of fresh oysters, 10g each, homogenate, add deionized water at a liquid-solid ratio of 1~3:1, stir evenly in a water bath at 45°C, add oysters and compound protease at a mass ratio of 1000:8~16 Compound protease (1.0×10 5 U / g, Suo Laibao Technology Co., Ltd.), adjust the pH to 7~8, keep it warm for 5 min, and then digest it at 45~65°C for 3~7 h. After the enzyme hydrolysis, take the enzyme The solution was heated at 95-100 °C for 10 min to inactivate the enzyme until the end of the enzymatic hydrolysis reaction, centrifuged, the supernatant was taken, and freeze-dried to obtain 10 kinds of oyster active peptides. The numbers are oyster active peptides 1-10.

[0030] The specific enzymatic hydrolysis conditions of each oyster active peptide sample are shown in Table 1.

[0031] Table 1 Enzymatic hydrolysis conditions of oyster active peptides

[0032]

Embodiment 2

[0034] In vitro screening of oyster active peptides that improve memory:

[0035] Take 10 copies of astrocytes in the logarithmic growth phase (numbered astrocytes 1-10), inoculate them in 24-well plates, inoculate density 1×10 5 cells / mL, inoculate volume 0.5 mL, and culture for 24 h, after the astrocytes grew to 80% of the orifice plate, the original astrocyte culture solution was sucked away, and the oyster active peptides (0.5mL, 10mg / mL) in Example 1 were added in turn to act on the For astrocytes, suck out the culture medium again after 24 h, add 0.5ml phosphate buffer (0.01 M, pH 7.2) to 10 astrocytes respectively, grind the astrocytes thoroughly, and keep at 4°C Centrifuge at 5000 r / min for 10 min under ambient conditions, take 0.4 mL of the supernatant, use the AchE test kit (Nanjing Jiancheng Bioengineering Research Institute, article number: A024, test method reference manual) to measure the activity of AchE, and screen out the inhibition rate of AchE The highest o...

Embodiment 3

[0040] In vitro screening of oyster active peptides that improve memory:

[0041] Take 10 copies of astrocytes in the logarithmic growth phase (numbered as astrocytes 1-10) and inoculate them in a 24-well plate with a seeding density of 1×10 6 cells / mL and an inoculum volume of 0.5 mL. Culture for 24 h, after the astrocytes grew to 90% of the orifice plate, the original astrocyte culture medium was sucked away, and the oyster active peptides (0.5mL, 10mg / mL) in Example 1 were added in turn to act on the Astrocytes, aspirate the culture medium again after 24 h, add 1.0mL phosphate buffer solution (0.01 M, pH 7.4) to 10 astrocytes respectively, fully grind the astrocytes, 4°C Centrifuge at 8,000 r / min for 15 min under ambient conditions, take 0.5 mL of the supernatant, use the AchE test kit (Nanjing Jiancheng Bioengineering Research Institute, article number: A024, refer to the instruction manual for the detection method) to measure the activity of AchE, and screen out the inhib...

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Abstract

The invention discloses a method for screening oyster bioactive peptides with effect of improving memory in vitro. The method comprises the following screening steps: inoculating astrocytes in a logarithmic phase to a 24-well plate, and culturing for 24 hours; after the astrocytes grow to 80%-90% of the 24-well plate, sucking an astrocyte culture solution, and adding to-be-screened oyster bioactive peptides to act on the astrocytes; sucking the culture solution after 24 hours again, adding phosphate buffer into the astrocytes, fully grinding the astrocytes, centrifuging to obtain supernatant, measuring acetylcholin esterase (AchE) activity with the supernatant, and screening the oyster bioactive peptide with the highest AchE inhibition rate according to the AchE activity. According to the method disclosed by the invention, the astrocytes with the widest distribution in brains of mammals are taken as a mode, and the AchE is directly and accurately extracted to be used for in-vitro activity screening of the oyster bioactive peptides.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for in vitro screening oyster active peptides capable of improving memory. Background technique [0002] With the acceleration of the pace of modern life and the increase of learning and work pressure, sub-health problems such as learning memory and cognitive function decline are becoming more and more serious. Therefore, the research and development of natural active substances that can relieve human mental stress and improve learning and memory ability Significance. [0003] Acetylcholinesterase (AchE) is a key enzyme in biological nerve transmission, mainly present in neuromuscular [0004] In the joint and cholinergic nervous system, its main physiological function is to hydrolyze the neurotransmitter in the cholinergic synaptic cleft into acetic acid and choline, and terminate the exciting stimulation. The current in vitro screening method for judging acti...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/02C12P21/06
CPCC12N2503/00C12P21/06G01N33/5038G01N33/58G01N2500/10
Inventor 王雪芹于华华邢荣娥刘松陈晓琳李荣锋李鹏程
Owner INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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