CN108864249A - A kind of purification process of hydrophobic peptides - Google Patents
A kind of purification process of hydrophobic peptides Download PDFInfo
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- CN108864249A CN108864249A CN201810631333.2A CN201810631333A CN108864249A CN 108864249 A CN108864249 A CN 108864249A CN 201810631333 A CN201810631333 A CN 201810631333A CN 108864249 A CN108864249 A CN 108864249A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/16—Extraction; Separation; Purification by chromatography
- C07K1/20—Partition-, reverse-phase or hydrophobic interaction chromatography
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Abstract
The invention belongs to separation and purification of biomolecules technical field, in particular to a kind of purification process of hydrophobic peptides includes the following steps：Step 1, hydrophobic peptides crude product is dissolved in n,N-Dimethylformamide, is configured to the polypeptide solution of 4-40mg/L；Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；The purification condition of the high performance liquid chromatography is：Chromatographic column：C18；Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 1-2:100；Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 1-2:100；Flow velocity：1-2ml/min；The present invention can purify hydrophobic peptides, and purification process is simple, efficiently, the hydrophobic peptides purity is high obtained through this method, and and significant loss is few in purification process, save the cost, it is environmentally protective.
Description
Technical field
The invention belongs to separation and purification of biomolecules technical field, in particular to a kind of purification process of hydrophobic peptides.
Background technique
Polypeptide is a-amino acid with the compound that peptide bond links together and is formed, it is also that the intermediate of protein hydrolysis produces
Object.The compound as made of two amino acid molecular dehydrating condensations is called dipeptides, similarly analogizes also tripeptides, tetrapeptide, pentapeptide etc..
Usually the compound as made of three or three or more amino acid molecular dehydrating condensations, which can become, is polypeptide.
Hydrophobic peptides refer to the polypeptide rich in hydrophobic amino acid.Hydrophobic amino acid is that side chain has high hydrophobicity
Amino acid general name amino acid side chain hydrophobicity, be that the value for subtracting glycine hydrophobicity from the hydrophobicity of each amino acid carrys out table
Show.Hydrophobic amino acid has tyrosine, tryptophan, phenylalanine, valine, leucine, isoleucine, alanine and methionine
(methionine).Hydrophobic amino acid is in protein interior, due to its hydrophobic interaction, and in the three-level for keeping protein
It works in structure.In addition the molecule combination side of the various non-covalent bonds such as interaction between enzyme and matrix, antibody and antigen
Face plays a significant role, such as has many hydrophobic amino acids on the position with antigen binding of antibody, it participates in resisting with half
Former combination.In the configuration aspects for maintaining biomembrane, hydrophobic amino acid also has effect, such as present on red blood cell film
There are many hydrophobic amino acids for its film inner part of glycophorin, form a hydrophobic region.
Incomplete peptide, byproduct of reaction, remaining reagent etc. can be generated in the synthesis and cutting of polypeptide, it is necessary to polypeptide
It is purified.
Summary of the invention
The present invention solves the above-mentioned technical problems in the prior art, provides a kind of purification process of hydrophobic peptides.
To solve the above problems, technical scheme is as follows：
A kind of purification process of hydrophobic peptides, includes the following steps：
Step 1, hydrophobic peptides crude product is dissolved in n,N-Dimethylformamide, the polypeptide for being configured to 4-40mg/L is molten
Liquid；
Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；
The purification condition of the high performance liquid chromatography is：
Chromatographic column：C18；
Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 1-2:100；
Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 1-2:100；
Flow velocity：1-2ml/min；
Detection wavelength：220nm.
Preferably, the model of the chromatographic column：Diamonsil C18,4.6*250mm, 5um.
Preferably, the proportion of solution A is in the mobile phase：The mass ratio of trifluoracetic acid and acetonitrile is 1:100.
Preferably, the proportion of solution B is in the mobile phase：The mass ratio of trifluoracetic acid and water is 1:100.
Preferably, the flow velocity is 2ml/min.
Preferably, the hydrophobic peptides are comprising glycine, alanine, leucine, isoleucine, valine, dried meat ammonia
Acid, phenylalanine, more than any one or a few in methionine amino acid polypeptide.
Preferably, the purity of the crude product hydrophobic peptides is 87-89%.
Compared with the existing technology, advantages of the present invention is as follows,
The present invention can purify hydrophobic peptides, and purification process is simple, efficiently, the hydrophobicity obtained through this method
Purity is high, and significant loss is few in purification process, save the cost, environmentally protective.
Detailed description of the invention
Fig. 1 is the high-efficient liquid phase chromatogram of embodiment 1；
Fig. 2 is the high-efficient liquid phase chromatogram of embodiment 2；
Fig. 3 is the high-efficient liquid phase chromatogram of embodiment 3；
Fig. 4 is the high-efficient liquid phase chromatogram of embodiment 4.
Specific embodiment
The purity of hydrophobic peptides crude product employed in the following example is 87-89%.
Embodiment 1：
A kind of purification process of hydrophobic peptides, includes the following steps：
Step 1, hydrophobic peptides crude product is dissolved in n,N-Dimethylformamide, is configured to the polypeptide solution of 20mg/L；
Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；
The purification condition of the high performance liquid chromatography is：
Chromatographic column：Diamonsil C18,4.6*250mm, 5um；
Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 1:100；
Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 1:100；
Flow velocity：1ml/min；
Detection wavelength：220nm.
The hydrophobic peptides are Fmoc-Leu-Pro-Gly-OH；
High-efficient liquid phase chromatogram is as shown in Figure 1.
In the product isolated, hydrophobic peptides purity is up to 99.3%.
Embodiment 2：
A kind of purification process of hydrophobic peptides, includes the following steps：
Step 1, hydrophobic peptides crude product is dissolved in n,N-Dimethylformamide, is configured to the polypeptide solution of 4mg/L；
Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；
The purification condition of the high performance liquid chromatography is：
Chromatographic column：Diamonsil C18,4.6*250mm, 5um；
Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 1:100；
Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 1:100；
Flow velocity：2ml/min；
Detection wavelength：220nm.
The hydrophobic peptides are ILE-LEU-LYS-GLU-PRO-VAL-HIS-GLY-VAL；
High-efficient liquid phase chromatogram is as shown in Figure 2.
In the product isolated, hydrophobic peptides purity is up to 99.1%.
Embodiment 3：
A kind of purification process of hydrophobic peptides, includes the following steps：
Step 1, hydrophobic peptides crude product is dissolved in n,N-Dimethylformamide, is configured to the polypeptide solution of 40mg/L；
Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；
The purification condition of the high performance liquid chromatography is：
Chromatographic column：Diamonsil C18,4.6*250mm, 5um；
Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 2:100；
Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 2:100；
Flow velocity：2ml/min；
Detection wavelength：220nm.
The hydrophobic peptides are KAFSPEVIPMF；
High-efficient liquid phase chromatogram is as shown in Figure 3.
In the product isolated, hydrophobic peptides purity is up to 99.0%.
Embodiment 4：
A kind of purification process of hydrophobic peptides, includes the following steps：
Step 1, the molten crude product solution of hydrophobic peptides is configured to the polypeptide solution of 10mg/L in n,N-Dimethylformamide；
Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；
The purification condition of the high performance liquid chromatography is：
Chromatographic column：Diamonsil C18,4.6*250mm, 5um；
Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 1:100；
Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 1:100；
Flow velocity：1ml/min；
Detection wavelength：220nm.
The hydrophobic peptides are TPQDLNTML；
High-efficient liquid phase chromatogram is as shown in Figure 4.
In the product isolated, hydrophobic peptides purity is up to 99.2%.
It should be noted that above-described embodiment is only presently preferred embodiments of the present invention, there is no for the purpose of limiting the invention
Protection scope, the equivalent substitution or substitution made on the basis of the above all belong to the scope of protection of the present invention.
Claims (7)
1. a kind of purification process of hydrophobic peptides, which is characterized in that include the following steps：
Step 1, hydrophobic peptides crude product is dissolved in n,N-Dimethylformamide, is configured to the polypeptide solution of 4-40mg/L；
Step 2, polypeptide solution made of step 1 is subjected to high performance liquid chromatography separation purifying；
The purification condition of the high performance liquid chromatography is：
Chromatographic column：C18；
Mobile phase：Solution A：The mass ratio of trifluoracetic acid-acetonitrile solution, trifluoracetic acid and acetonitrile is 1-2:100；
Solution B：The mass ratio of trifluoracetic acid-aqueous solution, trifluoracetic acid and water is 1-2:100；
Flow velocity：1-2ml/min；
Detection wavelength：220nm.
2. the purification process of hydrophobic peptides as described in claim 1, which is characterized in that the model of the chromatographic column：
Diamonsil C18,4.6*250mm, 5um.
3. the purification process of hydrophobic peptides as described in claim 1, which is characterized in that solution A matches in the mobile phase
Than for：The mass ratio of trifluoracetic acid and acetonitrile is 1:100.
4. the purification process of hydrophobic peptides as described in claim 1, which is characterized in that solution B matches in the mobile phase
Than for：The mass ratio of trifluoracetic acid and water is 1:100.
5. the purification process of hydrophobic peptides as described in claim 1, which is characterized in that the flow velocity is 1ml/min.
6. the purification process of hydrophobic peptides as described in claim 1, which is characterized in that the hydrophobic peptides are comprising sweet
Propylhomoserin, alanine, leucine, isoleucine, valine, proline, phenylalanine, it is more than any one or a few in methionine
Amino acid polypeptide.
7. the purification process of hydrophobic peptides as described in claim 1, which is characterized in that the hydrophobic peptides crude product it is pure
Degree is 87-89%.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113092648A (en) * | 2021-04-09 | 2021-07-09 | 江苏沿海化学品检测技术服务有限公司 | Separation and purification method of amino acid |
| CN116162124A (en) * | 2023-04-21 | 2023-05-26 | 吉尔生化（上海）有限公司 | Preparation method of continuous glutamine polypeptide |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101525382A (en) * | 2009-04-21 | 2009-09-09 | 深圳市翰宇药业有限公司 | Method of purifying pramlintide |
| CN105037488A (en) * | 2015-08-25 | 2015-11-11 | 南京肽业生物科技有限公司 | Purification method of melanotan II |
| CN105906709A (en) * | 2016-05-17 | 2016-08-31 | 青岛海博瑞克生物科技有限公司 | Alaska Pollock fish skin active oligopeptides as well as synthesis method and application thereof |
| CN107698656A (en) * | 2017-05-04 | 2018-02-16 | 苏州强耀生物科技有限公司 | A kind of purification process of hydrophobic peptides raw material |
-
2018
- 2018-06-19 CN CN201810631333.2A patent/CN108864249A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101525382A (en) * | 2009-04-21 | 2009-09-09 | 深圳市翰宇药业有限公司 | Method of purifying pramlintide |
| CN105037488A (en) * | 2015-08-25 | 2015-11-11 | 南京肽业生物科技有限公司 | Purification method of melanotan II |
| CN105906709A (en) * | 2016-05-17 | 2016-08-31 | 青岛海博瑞克生物科技有限公司 | Alaska Pollock fish skin active oligopeptides as well as synthesis method and application thereof |
| CN107698656A (en) * | 2017-05-04 | 2018-02-16 | 苏州强耀生物科技有限公司 | A kind of purification process of hydrophobic peptides raw material |
Non-Patent Citations (1)
| Title |
|---|
| 王永芬、何金环主编: "《生物化学》", 31 August 2015, 中国医药科技出版社 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113092648A (en) * | 2021-04-09 | 2021-07-09 | 江苏沿海化学品检测技术服务有限公司 | Separation and purification method of amino acid |
| CN116162124A (en) * | 2023-04-21 | 2023-05-26 | 吉尔生化（上海）有限公司 | Preparation method of continuous glutamine polypeptide |
| CN116162124B (en) * | 2023-04-21 | 2023-06-30 | 吉尔生化（上海）有限公司 | Preparation method of continuous glutamine polypeptide |
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Application publication date: 20181123 |