AU2017101408A4 - Synthetic method of Cyclo(His-Pro) - Google Patents
Synthetic method of Cyclo(His-Pro) Download PDFInfo
- Publication number
- AU2017101408A4 AU2017101408A4 AU2017101408A AU2017101408A AU2017101408A4 AU 2017101408 A4 AU2017101408 A4 AU 2017101408A4 AU 2017101408 A AU2017101408 A AU 2017101408A AU 2017101408 A AU2017101408 A AU 2017101408A AU 2017101408 A4 AU2017101408 A4 AU 2017101408A4
- Authority
- AU
- Australia
- Prior art keywords
- pro
- methyl ester
- amino group
- cyclo
- protected
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
- C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
- C07D487/04—Ortho-condensed systems
Abstract
The present invention provides a synthetic method of Cyclo(His-Pro), comprising steps of: preparing His-Pro dipeptide methyl ester with protected amino group from histidine and proline, wherein one of histidine and proline is in the form of methyl ester hydrochloride and the other is in the form of protected amino group; deprotecting and cyclizing the His-Pro dipeptide methyl ester with protected amino group to produce Cyclo(His-Pro).
Description
Synthetic Method of Cyclo(His-Pro)
Technical Field
The present invention belongs to the technical field of polypeptide synthesis technology, and particularly relates to a synthetic method of Cyclo(His-Pro).
Background of the Invention
Any discussion of the prior art throughout the specification should in no way be considered as an admission that such prior art is widely known or forms part of common general knowledge in the field.
Cyclo(His-Pro) (CHP for short, also known as His-Pro cyclodipeptide) is a class of active peptide substances, specifically is an endogenous cyclic dipeptide, and widely exists in the bodies of human, monkey, dog, cat, rat, frog, and the like. Cyclo(His-Pro) is produced in the organism by subjecting thyrotropin releasing hormone (TRH) to hydrolysis with pyroglutamic acid aminopeptidase to remove pyroglutamic acid and then to cyclization. Cyclo(His-Pro) has many biological activities, studies in recent years have shown that it plays a good role in lowering blood sugar, and is expected to be formulated to drugs and put into the market.
Chinese Patent Application No. 200510109439.9 discloses a preparing method of Pro-His cyclodipeptide, which comprises the following steps: reacting proline methyl ester and protected histidine in the presence of dicyclohexylcarbodiimide to obtain protected Pro-His dipeptide methyl ester; and removing protection in a solution of hydrogen chloride in ethyl acetate to generate Pro-His cyclodipeptide. The drawback of this method is that the protected Pro-His dipeptide methyl ester is refluxed in the solution of hydrogen chloride in ethyl acetate under a high temperature for a long period, which will cause an irreversible effect on optical purity.
It is an object of the present invention to overcome or ameliorate at least one of the disadvantages of the prior art, or to provide a useful alternative.
Disclosure of the Invention
The present invention relates to a synthetic method of preparing Cyclo(His-Pro) to overcome the defects existing in prior art.
The present invention provides a synthetic method of preparing Cyclo(His-Pro) comprising steps of: (1) preparing His-Pro dipeptide methyl ester with protected amino group from histidine and proline, wherein one of histidine and proline is in the form of methyl ester hydrochloride and the other is in the form of protected amino group; and (2) deprotecting and cyclizing His-Pro dipeptide methyl ester with protected amino group prepared in step (1) to produce Cyclo(His-Pro).
The aforementioned synthetic method, in step (2), deprotecting and cyclizing His-Pro dipeptide methyl ester with protected amino group prepared in step (1) to produce Cyclo(His-Pro) is performed in a solution of piperidine in tetrahydrofuran.
The aforementioned synthetic method, the ratio of His-Pro dipeptide methyl ester with protected amino group : tetrahydrofuran : piperidine is 1 - 4 g : 25 - 55 ml: 0.5 - 2 ml, and preferably, the ratio of His-Pro dipeptide methyl ester with protected amino group : tetrahydrofuran : piperidine is 2 g : 40 ml: 1 ml.
The aforementioned synthetic method, in step (1), His-Pro dipeptide methyl ester with protected amino group is prepared from histidine methyl ester hydrochloride and proline with protected amino group.
The aforementioned synthetic method, in step (1), proline with amino group being protected by fluorenyl methoxycarbonyl group is reacted with N-hydroxysuccinimide in the presence of Ν,Ν'-Dicyclohexylcarbodiimide to produce an active ester, and the active ester is further reacted with histidine methyl ester hydrochloride under an alkaline condition to produce His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group.
The aforementioned synthetic method, the ratio of proline with amino group being protected by fluorenyl methoxycarbonyl group : N-hydroxysuccinimide : Ν,Ν'-Dicyclohexylcarbodiimide : histidine methyl ester hydrochloride is 6-10 : 8-12 : 8-12 : 9-12, and preferably 8 : 9 : 9 : 9.6.
The aforementioned synthetic method, in step (1), His-Pro dipeptide methyl ester with protected amino group is prepared from proline methyl ester hydrochloride and histidine with protected amino group.
The aforementioned synthetic method, in step (1), histidine with amino group being protected by fluorenyl methoxycarbonyl group is reacted with N-hydroxysuccinimide in the presence of Ν,Ν'-Dicyclohexylcarbodiimide to produce an active ester, and the active ester is further reacted with proline methyl ester hydrochloride under an alkaline condition to produce His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group.
The aforementioned synthetic method, the ratio of histidine with amino group being protected by fluorenyl methoxycarbonyl group : N-hydroxysuccinimide : Ν,Ν'-Dicyclohexylcarbodiimide : proline methyl ester hydrochloride is 6-10 : 8-12 : 8-12 : 9-12, preferably 8 : 9 : 9 : 9.6.
The aforementioned synthetic method, the Cyclo(His-Pro) is L-L type Cyclo(His-Pro), L-D type Cyclo(His-Pro), D-L type Cyclo(His-Pro), or D-D type Cyclo(His-Pro).
Compared with prior art, the synthetic method of Cyclo(His-Pro) provided by the present invention has the following advantages: (1) Cyclo(His-Pro) is synthesized by using proline with protected amino group or histidine with protected amino group as starting materials, and deprotection and cyclization are performed in one step in a solution of piperidine in tetrahydrofuran, which overcomes the defect that deprotection and cyclization in previous processes must be performed in two steps; (2) Reactions are performed under an alkaline condition at room temperature, which overcomes the defect that cyclization performed at high temperature and under an acidic condition in previous processes causes change in optical purity.
Unless the context clearly requires otherwise, throughout the description and the claims, the words “comprise”, “comprising”, and the like are to be construed in an inclusive sense as opposed to an exclusive or exhaustive sense; that is to say, in the sense of “including, but not limited to”.
Brief Description of the Drawings
Figure 1 is a 'h NMR spectroscopy of L-L type Cyclo(His-Pro) according to a first embodiment of the present invention.
Figure 2 is a mass spectrum of L-L type Cyclo(His-Pro) according to a first embodiment of the present invention.
Figure 3 is a 'H NMR spectroscopy of L-L type Cyclo(His-Pro) according to a second embodiment of the present invention.
Figure 4 is a mass spectrum of L-L type Cyclo(His-Pro) according to a second embodiment of the present invention.
Specific Mode for Carrying out the Present Invention
In order to fully understand the purposes, characteristics and effects of the present invention, the present invention will be described in detail with reference to the following detailed description.
The conventional synthetic methods of Cyclo(His-Pro) have such problems that the reaction conditions are harsh, the procedures are complex, the loss in optical purity is large and the yield is low. In order to solve these problems, the present invention provides a new synthetic route, through which Cyclo(His-Pro) is produced by liquid phase synthesis. Compared with the conventional methods, the method according to the present invention has the advantages that synthesis steps are less; less amino acids are protected, only the amino group of one of the starting amino acids is protected; the reaction conditions are mild and easy to operate, there is no loss in optical purity, and the yield is very high, up to 80%.
The synthetic method according to the present invention comprises steps of: preparing His-Pro dipeptide methyl ester with protected amino group from histidine methyl ester hydrochloride and proline with protected amino group, or from proline methyl ester hydrochloride and histidine with protected amino group; and deprotecting and cyclizing His-Pro dipeptide methyl ester with protected amino group to produce Cyclo(His-Pro).
According to the first embodiment of the present invention, the synthetic method according to the present invention comprises steps of: (1) Preparing His-Pro dipeptide methyl ester with protected amino group
His-Pro dipeptide methyl ester with protected amino group is prepared from histidine methyl ester hydrochloride and proline with protected amino group. Specifically, proline with amino group being protected by fluorenyl methoxycarbonyl group is reacted with N-hydroxysuccinimide in the presence of Ν,Ν'-Dicyclohexylcarbodiimide to produce an active ester, and the active ester is further reacted with histidine methyl ester hydrochloride under an alkaline condition (for example, in the presence of NaHCC>3) to produce His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group.
Wherein, the molar ratio of proline with amino group being protected by fluorenyl methoxycarbonyl group : N-hydroxysuccinimide : Ν,Ν'-Dicyclohexylcarbodiimide : histidine methyl ester hydrochloride is 6-10 : 8-12 : 8-12 : 9-12, and preferably 8 : 9 : 9 : 9.6. (2) deprotecting and cyclizing
His-Pro dipeptide methyl ester with protected amino group prepared in step (1) is deprotected and cyclized in a solution of piperidine in tetrahydrofuran at room temperature (e.g., 25°C) to produce Cyclo(His-Pro).
Wherein, His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group : tetrahydrofuran : piperidine is 1 - 4 g : 25 - 55 ml : 0.5 - 2 ml, and preferably, the ratio of His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group : tetrahydrofuran : piperidine is 2 g : 40 ml: 1 ml.
In each step, the ingredient ratios among the substances is controlled within the above ranges, which is a result of comprehensive consideration of such factors as the synthetic method, the ingredient ratios and the like by the inventors. The described technical effect can still be achieved when the ranges are changed ±10%. However, the preferred solution is shown as above.
The Cyclo(His-Pro) synthesized by the method according to the present invention may be L-L type Cyclo(His-Pro), L-D type Cyclo(His-Pro), D-L type Cyclo(His-Pro), or D-D type Cyclo(His-Pro). L-L type Cyclo(His-Pro) is synthesized by using L-histidine methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-L-proline as starting materials; L-D type Cyclo(His-Pro) is synthesized by using L-histidine methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-D-proline as starting materials; D-L type Cyclo(His-Pro) is synthesized by using D-histidine methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-L-proline as starting materials; and D-D type Cyclo(His-Pro) is synthesized by using D-histidine methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-D-proline as starting materials. Figure 1 and Figure 2 are a 'Η NMR spectroscopy and a mass spectrum of L-L type Cyclo(His-Pro), separately.
According to the second embodiment of the present invention, the synthetic method according to the present invention comprises steps of: (1) Preparing His-Pro dipeptide methyl ester with protected amino group
His-Pro dipeptide methyl ester with protected amino group is prepared from proline methyl ester hydrochloride and histidine with protected amino group. Specifically, histidine with amino group being protected by fluorenyl methoxycarbonyl group is reacted with N-hydroxysuccinimide in the presence of Ν,Ν'-Dicyclohexylcarbodiimide to produce an active ester, and the active ester is further reacted with proline methyl ester hydrochloride under an alkaline condition to produce His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group.
Wherein, the molar ratio of histidine with amino group being protected by fluorenyl methoxycarbonyl group : N-hydroxysuccinimide : Ν,Ν'-Dicyclohexylcarbodiimide : proline methyl ester hydrochloride is 6-10 : 8-12 : 8-12 : 9-12, and preferably 8 : 9 : 9 : 9.6. (2) Deprotecting and cyclizing
His-Pro dipeptide methyl ester with protected amino group prepared in step (1) is deprotected and cyclized in a solution of piperidine in tetrahydrofuran at room temperature (e.g., 25°C) to produce Cyclo(His-Pro).
Wherein, His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group : tetrahydrofuran : piperidine is 1 - 4 g : 25 - 55 ml : 0.5 - 2 ml, and preferably, the ratio of His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group : tetrahydrofuran : piperidine is 2 g : 40 ml: 1 ml.
In each step, the ingredient ratios among the substances is controlled within the above ranges, which is a result of comprehensive consideration of such factors as the synthesis method, the ingredient ratios and the like by the inventors. The described technical effect can still be achieved when the ranges are changed ±10%. However, the preferred solution is shown as above.
The Cyclo(His-Pro) synthesized by the method according to the present invention may be L-L type Cyclo(His-Pro), L-D type Cyclo(His-Pro), D-L type Cyclo(His-Pro), or D-D type Cyclo(His-Pro). L-L type Cyclo(His-Pro) is synthesized by using L- proline methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-L- histidine as starting materials; L-D type Cyclo(His- Pro) is synthesized by using L- proline methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-D-histidine as starting materials; D-L type Cyclo(His-Pro) is synthesized by using D- proline methyl ester hydrochloride and N-fluorenylmethoxycarbonyl- L- histidine as starting materials; and D-D type Cyclo(His-Pro) is synthesized by using D- proline methyl ester hydrochloride and N-fluorenylmethoxycarbonyl-D- histidine as starting materials. Figure 3 and Figure 4 are a 'Η NMR spectroscopy and a mass spectrum of L-L type Cyclo(His-Pro), separately.
Example
Unless otherwise specified, the raw materials and instruments used in the examples below are commercially available and are conventionally used in the art. The raw materials and instruments are not defined specially, as long as they meet the experimental requirements.
Now the shorthand in the specification is interpreted as follows:
Fmoc : fluorenyl methoxycarbonyl THF : tetrahydrofuran HOSu : N-hydroxysuccinimide DCC : N,N'-Dicyclohexylcarbodiimide
His : Histidine
Pro : Proline TLC : thin-layer chromatography Example 1 L-L type Cyclo(Pro-His) 1. Fmoc-L-Pro-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, L-H-His-0CH3.2HC1 (23 g, 0.096 mol), NaHC03 (30 g, 0.36 mol), and H20 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-His-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc-Pro-His-OCH3 (31 g). 2. 20 g of Fmoc-Pro-His-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and fdtered to give a white solid product L-L type Cyclo(Pro-His) (8.2 g). The product was confirmed by mass spectrometry and 'h NMR spectroscopy.
Example 2 L-D type Cyclo(Pro-His) 1. Fmoc-L-Pro-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, D-H-His-0CH3.2HC1 (23 g, 0.096 mol), NaHC03 (30 g, 0.36 mol), and H20 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl acetate layer was combined with the reserved ethyl acetate layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-His-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc-Pro-His-OCH3 (30 g). 2. 20 g of Fmoc-Pro-His-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product L-D type Cyclo(Pro-His) (8.1 g). The product was confirmed by mass spectrometry and 'fl NMR spectroscopy.
Example 3 D-D type Cyclo(Pro-His) 1. Fmoc-D-Pro-OH (27 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, D-H-His-0CH3.2HC1 (23 g, 0.096 mol), NaHCC>3 (30 g, 0.36 mol), and H2O 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-His-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc-Pro-His-OCH3 (33 g). 2. 20 g ofFmoc-Pro-His-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product D-D type Cyclo(Pro-His) (8.4 g). The product was confirmed by mass spectrometry and 1H NMR spectroscopy.
Example 4 D-L type Cyclo(Pro-His) 1. Fmoc-D-Pro-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, L-H-His-0CH3.2HC1 (23 g, 0.096 mol), NaHC03 (30 g, 0.36 mol), and H20 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-His-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was fdtrated and evaporated to dryness to give a white foam solid product Fmoc-Pro-His-OCH3 (31 g). 2. 20 g ofFmoc-Pro-His-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product D-L type Cyclo(Pro-His) (8.3 g). The product was confirmed by mass spectrometry and *H NMR spectroscopy.
Example 5 L-L type Cyclo(His-Pro) 1. Fmoc-L-His-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, L-H-Pro-OCH3.2HCl (16 g, 0.096 mol), NaHC03 (20 g, 0.24 mol), and H2O 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-Pro-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc-His-Pro-OCH3 (33 g). 2. 20 g ofFmoc-His-Pro-OCHs was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product L-L type Cyclo(His-Pro) (8 g). The product was confirmed by mass spectrometry and 'Η NMR spectroscopy.
Example 6 L-D type Cyclo(His-Pro) 1. Fmoc-L-His-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, D-H-Pro-OCH3.2HCl (16 g, 0.096 mol), NaHC03 (20 g, 0.24 mol), and H20 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-Pro-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc- His-Pro-OCH3 (32 g). 2. 20 g of Fmoc-His-Pro-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product L-D type Cyclo(His-Pro) (8.3 g). The product was confirmed by mass spectrometry and 1H NMR spectroscopy.
Example 7 D-D type Cyclo(His-Pro) 1. Fmoc-D-His-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, D-H-Pro-OCH3.2HCl (16 g, 0.096 mol), NaHC03 (20 g, 0.24 mol), and H20 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-Pro-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc- His-Pro-OCH3 (35 g). 2. 20 g ofFmoc-His-Pro-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product D-D type Cyclo(His-Pro) (8.8 g). The product was confirmed by mass spectrometry and 'H NMR spectroscopy.
Example 8 D-L type Cyclo(His-Pro) 1. Fmoc-D-His-OH (30 g, 0.08 mol) and 500 ml of THF were added into eggplant-shaped bottle, and dissolved completely by magnetic stirring. HOSu (10.2 g, 0.09 mol), and DCC (18.3 g, 0.09 mol) were added thereto successively. After stirring over night at room temperature, a large amount of white precipitate appeared. After filtration, the filtrate was added with THF to a volume of 500 ml. Under stirring at room temperature, L-H-Pro-OCH3.2HCl (16 g, 0.096 mol), NaHC03 (20 g, 0.24 mol), and H20 150ml were added into the filtrate, respectively. After stirring over night at room temperature, TLC was showed to completely react. The filtrate then was evaporated to remove THF, added with ethyl acetate and water, stirred and allowed to separate into layers. Ethyl ester layer and water layer were separated, and the ethyl ester layer was reserved. The water layer was extracted with ethyl ester once, and the resultant ethyl ester layer was combined with the reserved ethyl ester layer. The combined ethyl ester layer was washed twice with diluted hydrochloric acid aqueous solution to remove excess H-Pro-OCH3, then washed with brine three times until the aqueous solution was neutral. The ethyl ester layer was filtrated and evaporated to dryness to give a white foam solid product Fmoc-His-Pro-OCH3 (34 g). 2. 20 g of Fmoc-His-Pro-OCH3 was dissolved in 400 ml of THF. 10 ml of piperidine was added thereto. The mixture was stirred at room temperature for 6 hours. After TLC was showed to completely react, the mixture was evaporated to remove all of the solvent. Ethyl ether was added thereto. The resultant mixture was stirred and filtered to give a white solid product D-L type Cyclo(His-Pro) (8.6 g). The product was confirmed by mass spectrometry and 'fl NMR spectroscopy.
Claims (10)
- What is claimed is:1. A synthetic method of preparing Cyclo(His-Pro), wherein the synthetic method comprises steps of: (1) preparing His-Pro dipeptide methyl ester with protected amino group from histidine and proline, wherein one of histidine and proline is in the form of methyl ester hydrochloride and the other is in the form of protected amino group; and (2) deprotecting and cyclizing His-Pro dipeptide methyl ester with protected amino group prepared in step (1) to produce Cyclo(His-Pro).
- 2. The synthetic method of claim 1, wherein, in step (2), deprotecting and cyclizing His-Pro dipeptide methyl ester with protected amino group prepared in step (1) to produce Cyclo(His-Pro) is performed in a solution of piperidine in tetrahydrofuran.
- 3. The synthetic method of claim 2, cwherein the ratio of His-Pro dipeptide methyl ester with protected amino group : tetrahydrofuran : piperidine is 1 - 4 g : 25 - 55 ml: 0.5 - 2 ml, and preferably, the ratio of His-Pro dipeptide methyl ester with protected amino group : tetrahydrofuran : piperidine is 2 g : 40 ml: 1 ml.
- 4. The synthetic method of any one of claims 1-3, wherein, in step (1), His-Pro dipeptide methyl ester with protected amino group is prepared from histidine methyl ester hydrochloride and proline with protected amino group.
- 5. The synthetic method of claim 4, wherein, in step (1), proline with amino group being protected by fluorenyl methoxycarbonyl group is reacted with N-hydroxysuccinimide in the presence of Ν,Ν'-Dicyclohexylcarbodiimide to produce an active ester, and the active ester is further reacted with histidine methyl ester hydrochloride under an alkaline condition to produce His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group.
- 6. The synthetic method of claim 5, wherein the ratio of proline with amino group being protected by fluorenyl methoxycarbonyl group : N-hydroxysuccinimide : Ν,Ν'-Dicyclohexylcarbodiimide : histidine methyl ester hydrochloride is 6-10 : 8-12 : 8-12 : 9-12, and preferably 8 : 9 : 9 : 9.6.
- 7. The synthetic method of any one of claims 1-3, wherein, in step (1), His-Pro dipeptide methyl ester with protected amino group is prepared from proline methyl ester hydrochloride and histidine with protected amino group.
- 8. The synthetic method of claim 7, wherein, in step (1), histidine with amino group being protected by fluorenyl methoxycarbonyl group is reacted with N-hydroxysuccinimide in the presence of Ν,Ν'-Dicyclohexylcarbodiimide to produce an active ester, and the active ester is further reacted with proline methyl ester hydrochloride under an alkaline condition to produce His-Pro dipeptide methyl ester with amino group being protected by fluorenyl methoxycarbonyl group.
- 9. The synthetic method of claim 8, wherein the ratio of histidine with amino group being protected by fluorenyl methoxycarbonyl group : N-hydroxysuccinimide : Ν,Ν'-Dicyclohexylcarbodiimide : proline methyl ester hydrochloride is 6-10 : 8-12 : 8-12 : 9-12, preferably 8 : 9 : 9 : 9.6.
- 10. The synthetic method of any one of claims 1-9, wherein the Cyclo(His-Pro) is L-L type Cyclo(His-Pro), L-D type Cyclo(His-Pro), D-L type Cyclo(His-Pro), or D-D type Cyclo(His-Pro).
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710004723.2A CN106674230B (en) | 2017-01-04 | 2017-01-04 | The synthetic method of pro-his cyclic dipeptide |
| CN201710004723.2 | 2017-01-04 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| AU2017101408A4 true AU2017101408A4 (en) | 2017-11-23 |
Family
ID=58848935
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| AU2017101408A Active AU2017101408A4 (en) | 2017-01-04 | 2017-10-23 | Synthetic method of Cyclo(His-Pro) |
Country Status (3)
| Country | Link |
|---|---|
| CN (1) | CN106674230B (en) |
| AU (1) | AU2017101408A4 (en) |
| WO (1) | WO2018126523A1 (en) |
Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111246859B (en) * | 2017-10-20 | 2022-05-03 | 诺麦塔制药有限公司 | Pharmaceutical composition for preventing or treating diabetes containing zinc salt, cyclodipeptide-histidine proline and antidiabetic drug as effective component |
| CN108395469A (en) * | 2018-02-01 | 2018-08-14 | 滨海吉尔多肽有限公司 | A kind of synthetic method of pseudo proline dipeptides |
| CN109438363B (en) * | 2018-10-31 | 2022-04-29 | 陕西慧康生物科技有限责任公司 | Liquid-phase high-purity large-scale synthesis method of cyclic (leucyl-arginyl) dipeptide salt |
| CN111875668B (en) * | 2020-07-29 | 2022-05-27 | 陕西慧康生物科技有限责任公司 | Synthetic method of cyclic dipeptide containing glutamine or asparagine |
| AU2022370372A1 (en) * | 2021-10-22 | 2024-05-09 | Neuren Pharmaceuticals Limited | Process for preparing bicyclic glycine-proline compounds and monocyclic glycine-proline intermediates thereof |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1500804A (en) * | 2002-11-18 | 2004-06-02 | 清华大学 | Preparing method for thread group cyclic dipeptide |
| CN1951954A (en) * | 2005-10-20 | 2007-04-25 | 海南和信堂药业有限公司 | Preparation method for pro-his cyclic dipeptide |
| CN102093470A (en) * | 2010-12-10 | 2011-06-15 | 大连伊美生物科技有限公司 | Liquid phase synthesis method of Cyclo(His-Pro) (CHP) |
| CN105294830A (en) * | 2014-06-18 | 2016-02-03 | 陈光健 | Dipeptide molecule, preparation method therefor and application thereof |
| CN105622717A (en) * | 2014-10-29 | 2016-06-01 | 四川好医生药业集团有限公司 | Six cyclic dipeptides used for wound healing |
| CN104447759B (en) * | 2014-11-24 | 2017-05-03 | 吉林农业大学 | Production method for cyclic dipeptide |
-
2017
- 2017-01-04 CN CN201710004723.2A patent/CN106674230B/en active Active
- 2017-02-28 WO PCT/CN2017/075123 patent/WO2018126523A1/en active Application Filing
- 2017-10-23 AU AU2017101408A patent/AU2017101408A4/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| WO2018126523A1 (en) | 2018-07-12 |
| CN106674230B (en) | 2019-07-02 |
| CN106674230A (en) | 2017-05-17 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2017101408A4 (en) | Synthetic method of Cyclo(His-Pro) | |
| KR101467598B1 (en) | Process for hcv protease inhibitor intermediate | |
| NO171788B (en) | ANALOGY PROCEDURE FOR THE PREPARATION OF THERAPEUTIC ACTIVE PHOSPHINIC ACID DERIVATIVES | |
| CA1188296A (en) | Tripeptides and process for preparing same | |
| EP0449079B1 (en) | Hydantoin derivative | |
| AU579835B2 (en) | The synthesis of cyclopropane amino acids and peptides | |
| CA1151154A (en) | Tripeptide amides and process for preparing same | |
| EP0541946B1 (en) | Process for diastereo selective reductive pinacol-coupling of homochiral alpha-aminoaldehydes | |
| YOSHIDA et al. | PROBESTIN, A NEW INHIBITOR OF AMINOPEPTIDASE M, PRODUCED BY STREPTOMYCES AZUREUS MH663-2F6 II. STRUCTURE DETERMINATION OF PROBESTIN | |
| EP0228625B1 (en) | Peptide derivatives inhibitors of hydroxyling enzymes, their process of preparation, the agents thereof and their use | |
| CN107098950B (en) | A kind of synthetic method of GHK or AHK tripeptides | |
| DE60301820T2 (en) | Process for the synthesis of perindopril and its pharmaceutically acceptable salts | |
| JPS6089497A (en) | Manufacture of n-alkylated dipeptide | |
| US6166217A (en) | Process for the production of alkoxycarbonyldipeptides intermediates in the synthesis of the lisinopril | |
| DE60301774T2 (en) | Process for the synthesis of (2S, 3aS, 7aS) -1 - ((S) -alanyl) -octahydro-1H-indole-2-carboxylic acid derivatives and use in the synthesis of perindopril | |
| EP0373497A2 (en) | Dipeptide derivatives with an enzyme-inhibiting activity | |
| Pettit et al. | Structural biochemistry. 25. Antineoplastic agents. 110. Synthesis of the dolastatin 3 isomer cyclo [L-Pro-L-Leu-L-Val-(R, S)-(gln) Thz-(gly) Thz] | |
| EP0410278B1 (en) | Renin inhibiting aminooligohydroxy-derivatives | |
| DE3113610A1 (en) | S-ACYLATION PRODUCTS OF A MERCAPTOACYLAMINO ACID AND A DIURETIC CONTAINING A CARBOXYL GROUP | |
| IL35005A (en) | Process for the manufacture of peptides | |
| UA71608C2 (en) | A method for producing the cyclic pentapeptide | |
| CN104557793A (en) | Synthetic method of carfilzomib intermediate and carfilzomib intermediate | |
| RU2086561C1 (en) | Method for production of nonapeptide ethyl amide | |
| US6767992B1 (en) | Method for producing L-prolyl-L-M-sarcolysyl-L-p-fluorophenylalanine and derivatives thereof | |
| CN117720617A (en) | Synthesis process of acetyl tetrapeptide-3 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| FGI | Letters patent sealed or granted (innovation patent) |