Skin wound repair peptide RL-RF10 as well as purification method and application thereof

A RL-RF10, skin trauma technology, applied in the field of biomedicine, can solve problems such as unsatisfactory activity, dysfunction, skin deformation, etc., and achieve the effects of reducing scar generation, accelerating wound healing, and promoting repair ability

Active Publication Date: 2019-12-06
KUNMING MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The second is excessive wound healing exemplified by hypertrophic scars and keloids, which can also affect appearance and cause skin deformation leading to dysfunction
Generally speaking, most of these drugs are of a single type, and have certain limitations: the former is unstable, difficult to synthesize, and unsatisfactory in activity
Generally speaking, highly active, stable, and low-cost active peptides for skin wound repair have broad market prospects, but the discovery and reports of related drug molecules are still rare

Method used

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  • Skin wound repair peptide RL-RF10 as well as purification method and application thereof
  • Skin wound repair peptide RL-RF10 as well as purification method and application thereof
  • Skin wound repair peptide RL-RF10 as well as purification method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0022] The preparation method of skin wound repairing peptide RL-RF10 comprises the following steps:

[0023] (1) Take live skin secretions of Frog yunnanensis by electrical stimulation, dissolve them in PBS, freeze-dry in vacuum, and obtain freeze-dried powder of skin secretions and store them at -80°C for later use;

[0024] (2) Dissolve the lyophilized powder in ultrapure water, then centrifuge at 12000×g for 20 minutes at 4°C, collect the supernatant, and then ultrafilter it with an ultrafilter with a molecular weight cut-off of 10 kDa;

[0025] (3) The isolated product of step (2) is subjected to high-performance liquid chromatography reversed-phase chromatography for the first time, and the active components with the function of promoting wound repair are collected and vacuum freeze-dried;

[0026] (4) Dissolve the product of step (2) in deionized water, and perform a second high performance liquid chromatography reverse phase chromatography to obtain the purified repair...

Embodiment 1

[0032] Example 1: Separation, purification and identification of novel peptide RL-RF10 for promoting skin wound repair

[0033] 1. Separation and purification

[0034] The skin secretions (dissolved in PBS) of Frog yunnanensis collected from the estuary of Yunnan were collected by electrical stimulation, vacuum freeze-dried, and stored at -80° for later use.

[0035] Step 1: Redissolve the lyophilized secretion in ultrapure water, then centrifuge at 12000×g for 20 minutes at 4°C, collect the supernatant, and then ultrafilter it with an Amicon ultrafilter to retain The molecular weight is 10 kDa (MerckMillipor, Germany).

[0036] The second step: the first high-performance liquid chromatography reversed-phase chromatography:

[0037] Take the sample dissolved in deionized water obtained in the first step, and load it on a Hypersil ODS2 5 μm column (product of Yilite, with a size of 4.6 mm × 300 mm) equilibrated with ultrapure water (containing 0.1% trifluoroacetic acid). ), ...

Embodiment 2

[0043] Example 2: Detection of the scratch repair activity of the polypeptide in HaCaT cells

[0044] Human immortalized keratinized epithelial cells were cultured in cell culture flasks with DMEM / F12 (BI, Israel) medium containing 10% fetal bovine serum and 1% double antibodies (penicillin, streptomycin, 100 U / ml) ( HaCaT). For cell scratch experiments in 24-well plates, seed 2.5 × 10 5 HaCaT cells were cultured for about 12–14 h. When the cells in each well were confluent, scratch each well with a yellow 200 μl pipette tip (Axygen, USA), and then discard the medium containing dead cells in each well. Wash each well twice with phosphate buffered saline (PBS), and finally add 500 μl of empty medium containing different concentrations of samples (10pM, 100 pM, 1 nM,) to each well without fetal bovine serum. Use a microscope (Zeiss, Germany) to take pictures every 12 hours to record the scratch healing status, and record continuously for 24 hours. We evaluated the percentage o...

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PUM

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Abstract

The invention discloses a skin wound repair peptide RL-RF10. An amino acid sequence contained in the repair peptide RL-RF10 is RFCFKGTPCG, and the invention also discloses a purification method and application of the repair peptide RL-RF10. The skin wound repair peptide RL-RF10 disclosed by the invention can be used for remarkably promoting healing of skin acute wounds and reducing scar generation, has the capacity of promoting dental ulcer repair and chronic wound healing, is one of the strongest active substances for promoting skin repair in the world, and has wide application prospects.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a skin wound repairing peptide RL-RF10 and its purification method and application. Background technique [0002] As the largest organ and physical barrier of the human body against the external environment, the skin wraps around the surface of the human body and plays an important role in protection, temperature regulation, sensation, secretion and excretion, absorption, metabolism and immunity. However, many challenges, including mechanical, thermal, chemical, and radiation stimuli, can cause skin damage and disrupt the normal function of the skin. The process of repairing skin damage is a very natural and common process, but there are many factors that can affect one or several stages of the healing process, which then lead to two severe types of wound healing. The first is chronic or non-healing wounds that cause severe physical and economic loss. burdens pa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/46C07K1/36C07K1/34C07K1/20A61K38/08A61P17/02A61K8/64A61Q19/02
CPCC07K14/463A61P17/02A61K8/64A61Q19/02A61K38/00
Inventor 杨新旺王滢唐璟王思媛李自良
Owner KUNMING MEDICAL UNIVERSITY
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