Method for extracting purified high ferro myoglobins from cardiac muscle

A metmyoglobin and myoglobin technology, which is applied in hemoglobin/myoglobin, peptide preparation methods, chemical instruments and methods, etc., can solve problems that are not suitable for industrial production, and achieve low cost, simple process, and high efficiency. The effect of economic value

Inactive Publication Date: 2006-07-12
NANJING NORMAL UNIVERSITY
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  • Abstract
  • Description
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  • Application Information

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  • Method for extracting purified high ferro myoglobins from cardiac muscle
  • Method for extracting purified high ferro myoglobins from cardiac muscle
  • Method for extracting purified high ferro myoglobins from cardiac muscle

Examples

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Embodiment 1

[0028] Embodiment 1: the preparation of metmyoglobin

[0029] Material

[0030] Fresh pig heart, anion exchange chromatography material (DEAE-Sepharose Fast Flow, Pharmacia), gel filtration chromatography material (Sephacryl S-200HR, Amersham-Pharmacia), horse cardiac myoglobin standard substance (Sigma, purity > 90 %), Coomassie brilliant blue kit, and other reagents were of analytical grade. Myoglobin extraction buffer: 10mM Tris-HCl, 1mM EDTA, pH8.0; Equilibration buffer A: 10mM Tris-HCl, 1mM EDTA, 10mM NaCl, pH8.5; Equilibration buffer B: 100mM Tris-HCl, 100mM NaCl , pH7.5. Main instruments and reagents

[0031] DS-1 high-speed tissue masher, FJ-200 high-speed dispersion homogenizer, JB-2 constant temperature magnetic stirrer, PHS-3C precision pH meter, BioLogic LP chromatography system, Gel Doc2000 imaging system, JeDa801 gel analysis Software, Spectrumlab54 UV-visible spectrophotometer, LGJ1.5 freeze dryer, LGR10-4.2 refrigerated high-speed centrifuge, Eppendorf 5415...

Embodiment 2

[0041] Embodiment 2: Qualitative and quantitative of metmyoglobin

[0042] Carry out SDS-PAGE electrophoresis test with the crude extraction sample, dialysate sample, ion exchange chromatography sample and gel filtration treatment sample obtained in (1)-(4) step of implementing 1, the result is as follows image 3 shown. After electrophoresis, the gel was photographed with the Gel Doc2000 imaging system, and the band scanning quantitative analysis was carried out by JeDa801 gel analysis software. The results are as follows: Figure 4 .

[0043] From image 3 and Figure 4 It can be seen from the analysis that there are many types of proteins contained in the crude extraction sample, and the proteins cannot be separated well ( Figure 4 A); after 70%-100% ammonium sulfate segmental salting out and dialysis, roughly nine bands can be seen, namely mainly composed of nine kinds of proteins, and gel scanning after electrophoresis shows that in the extract solution after dialysi...

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Abstract

The invention relates to a protein extracting purify craft which is a method for extracting and purifying ferrihemoglobin. It dose first treatment to the fresh animal's myocardium, obtains the vegetable protein course extracting matter by ejection; the course matter uses sulfuric acid diammonium salt out and the dialyzing treatment with trapping molecular weight 14000 to obtain the dialysis example; the dialysis example uses DEAE-Sepharose Fast Flow anion exchange color printing to collect the brunneus eluent; it collects the eluent and uses Sephacryl S-200HR gel filtration chromatography to collect the brunneus eluent which is corresponding to the clean peek to obtain the ferrihemoglobin.

Description

technical field [0001] The invention relates to a protein extraction and purification process, in particular to a method for extracting and purifying metmyoglobin from myocardium. Background technique [0002] Myoglobin is an important heme protein in the cytoplasm of vertebrate skeletal muscle and myocardial tissue. Myoglobin has the property of reversibly binding oxygen. Its main function is to transport and store oxygen in muscle cells. Metmyoglobin (MetMb) is its oxidized form. It can be converted into deoxymyoglobin and oxymyoglobin under the action of certain enzymes in animals, which is important for the completion of normal respiratory process. Physiological effect, if this process is obstructed, it will affect the normal progress of the breathing process. Purified metmyoglobin can be used in the study of animal meat color stability, etc., and can also be used as a substrate for the detection of human Keshan disease. Therefore, the purified metmyoglobin can be used...

Claims

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Application Information

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IPC IPC(8): C07K1/14C07K14/805
Inventor 金邦荃汤祥明刘兴余
Owner NANJING NORMAL UNIVERSITY
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