CN108070021A - It can be assembled into the method for the small-molecular peptides and assembling structure high-sequential nanofiber of high-sequential nanofiber - Google Patents
It can be assembled into the method for the small-molecular peptides and assembling structure high-sequential nanofiber of high-sequential nanofiber Download PDFInfo
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- CN108070021A CN108070021A CN201711446630.1A CN201711446630A CN108070021A CN 108070021 A CN108070021 A CN 108070021A CN 201711446630 A CN201711446630 A CN 201711446630A CN 108070021 A CN108070021 A CN 108070021A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
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- D—TEXTILES; PAPER
- D01—NATURAL OR MAN-MADE THREADS OR FIBRES; SPINNING
- D01F—CHEMICAL FEATURES IN THE MANUFACTURE OF ARTIFICIAL FILAMENTS, THREADS, FIBRES, BRISTLES OR RIBBONS; APPARATUS SPECIALLY ADAPTED FOR THE MANUFACTURE OF CARBON FILAMENTS
- D01F4/00—Monocomponent artificial filaments or the like of proteins; Manufacture thereof
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Abstract
A kind of method of small-molecular peptides for caning be assembled into high-sequential nanofiber and assembling structure high-sequential nanofiber, is related to technical field of biological materials.The small-molecular peptides for caning be assembled into high-sequential nanofiber are mainly made of hydrophobic alkyl chain, β-pleated sheet amino acid, hydrophilic amino acid and N Amino End Groups sealing end；Preparation method is synthesized using the polypeptide solid-state reaction method of FMOC Preservation tactics; small-molecular peptides segment is extended successively on vector resin from C-terminal to N-terminal; reaction condition is simple, mild; obtained small-molecular peptides have it is amphipathic, can be assembled into the high-sequential nanofiber with good biocompatibility.The assemble method of high-sequential nanofiber is that above-mentioned small-molecular peptides are dissolved in water to concentration for 0.05wt% 12wt%, the pH of aqueous solution is adjusted to 78, it is placed at room temperature for, reaction condition is simple, mild, and obtained high-sequential nanofiber has good biocompatibility.
Description
Technical field
The present invention relates to technical field of biological materials, and more particularly to a kind of it can be assembled into the small of high-sequential nanofiber
The method of molecular peptide and assembling structure high-sequential nanofiber.
Background technology
In nanometer technology, structure high-sequential nanostructured has a wide range of applications value.Recent study shows height
Spend ordered nano-structure has special biological function in biological field.Jiang Xing spaces seminar just pointed out in 2008：Stem cell
The functional cells such as neuron, muscle cell, vascular endothelial cell can be formed according to certain geometric direction differentiation, these work(
The arrangement of energy property cell within the organization has directionality, therefore functional cell can be formed specifically according to certain orientation growth
Tissue.When the chemical constitution and mechanical property of cell culture substrate have directionality, the direction of growth of functional cell has
Selectivity, therefore it is Tissue Engineering Study to be grown according to a certain direction come control function cell by the design at culture medium bottom
An important topic.ChungCY seminars sequentially reported 2007 and Stupp seminars in 2010：In brain, the heart
Also the extracellular fibril of substantial amounts of high-sequential arrangement can be found in the vitals such as dirty, bone and spinal cord, they are to respective
The performance of function plays important function, but their forming process is wrapped in mystery.In order to effectively annotate ordered structure
How peculiar biological effect builds the ordered nano-structure with biocompatibility into matter of utmost importance.
Current own research and development goes out the skill method of a variety of orderly nano-arrays of structure, including the Static Spinning of polymer
Silk technology and vacuum deposition technique etc..The electrostatic spinning technique of polymer be by polymer solution or melt under electrostatic interaction
It carries out injection stretching and obtains the spinning process of micro nano-scale fiber, the severe reaction conditions of this method are, it is necessary to high temperature and pressure.
And evaporating deposition technique is using the physical processes such as thermal evaporation or glow discharge, arc discharge, and required apply is deposited in substrate surface
The technology of layer, this technology needs high pressure, and the biocompatibility of product is poor, is unfavorable for the application in biological direction.
It is simple, mild therefore, it is necessary to build reaction condition, there is the ordered fiber of good biocompatibility.
The content of the invention
It is an object of the invention to provide a kind of small-molecular peptides for caning be assembled into high-sequential nanofiber and its preparation sides
Method, reaction condition is simple, mild, and obtained small-molecular peptides have amphipathic, can be assembled into the height with good biocompatibility
Spend ordered nano-fibers.
It is an object of the invention to provide a kind of high-sequential nanofiber and its assemble method, reaction condition is simple, temperature
There is good biocompatibility with, obtained high-sequential nanofiber.
The present invention is solved its technical problem and is realized using following technical scheme.
The present invention proposes a kind of small-molecular peptides for caning be assembled into high-sequential nanofiber, and small-molecular peptides sequence is successively by dredging
Water alkyl chain, beta sheet amino acid, hydrophilic amino acid and N- Amino End Groups sealing end are formed.
Further, in present pre-ferred embodiments, hydrophobic alkyl chain includes stearic acid, palmitic acid, myristic acid, the moon
One kind in cinnamic acid.
Further, in present pre-ferred embodiments, beta sheet amino acid is the hydrophobic amino acid with beta sheet, is had
Body includes one or more of alanine, valine, glycine, phenylalanine.
Further, in present pre-ferred embodiments, hydrophilic amino acid include glutamic acid, one kind in lysine or
It is several.
Further, in present pre-ferred embodiments, the structural formula of small-molecular peptides is C15H31CO-AAAAAKK-CONH2
Or C15H31CO-VVVVKKK-CONH2, wherein, A is alanine, and V is valine, and K is lysine.
The present invention proposes a kind of preparation method of the above-mentioned small-molecular peptides for caning be assembled into high-sequential nanofiber, is to adopt
It is synthesized with the polypeptide solid-state reaction method of FMOC Preservation tactics, vector resin is Rink Amide-AM Resin resins, by small molecule
Peptide segment extends successively on vector resin from C-terminal to N-terminal.
Further, in present pre-ferred embodiments, comprise the following steps：
S1, Rink Amide-AM Resin resins are placed in Solid-phase synthesis peptides column, use n,N-Dimethylformamide
Washing empties solvent, then is swollen with n,N-Dimethylformamide, empties solvent；
S2, piperidines/n,N-Dimethylformamide solution, stirring, with N, N- dimethyl formyls are added in into synthesis in solid state column
Amine washs, and empties solvent；
S3, beta sheet amino acid, benzotriazole-N, N, N are added in into synthesis in solid state column ', N '-tetramethylurea fluorine phosphorus
Hydrochlorate, 1- hydroxy benzo triazoles and n,N-diisopropylethylamine/n,N-Dimethylformamide solution, stirring；
Whether S4, beta sheet amino acid of taking a sample to check have been connected on resin；
If S5, beta sheet amino acid have been connected on resin, repeat step S2-S4, until connect on resin corresponding β-
Fold amino acid and quantity；
S6, piperidines/n,N-Dimethylformamide solution, stirring, with N, N- dimethyl formyls are added in into synthesis in solid state column
Amine washs, and empties solvent；
S7, hydrophilic amino acid, benzotriazole-N, N, N are added in into synthesis in solid state column ', N '-tetramethylurea fluorine phosphorus
Hydrochlorate, 1- hydroxy benzo triazoles and n,N-diisopropylethylamine/n,N-Dimethylformamide solution stir 1.5-4h；
Whether S8, hydrophilic amino acid of taking a sample to check have been connected on resin；
If S9, hydrophilic amino acid have been connected on resin, above-mentioned S6-S8 is repeated, until having met corresponding parent on resin
Aqueous amino acid and quantity；
S10, hydrophobic alkyl chain, benzotriazole-N, N, N are added in into synthesis in solid state column ', N '-tetramethylurea fluorophosphoric acid
Salt, 1- hydroxy benzo triazoles and n,N-diisopropylethylamine/n,N-Dimethylformamide solution, stirring；
Whether S11, hydrophobic alkyl chain of taking a sample to check have been connected on resin；
If S12, hydrophobic alkyl chain have been connected on resin, washed respectively with n,N-Dimethylformamide, methanol and dichloromethane
Resin is washed, after normal-temperature vacuum drying, obtains dry resin；
S13, trifluoroacetic acid/water/tri isopropyl silane mixing liquid, stirring at normal temperature are added in into dry resin, collection is cut
Fall liquid, then concentrated by rotary evaporation to thick liquid is added dropwise in cold ether and precipitates, supernatant, air drying are removed after centrifugation.
Further, in present pre-ferred embodiments, check that the method whether group has been connected on resin is：Take tree
Fat is put into ebuillition of heated 3-5 minutes in the methanol solution of ninhydrin and tests color, if non-discolouring, shows that group has been connected on resin
On.
The present invention proposes a kind of assemble method of high-sequential nanofiber, is that above-mentioned small-molecular peptides are dissolved in water
In to concentration be 0.05wt%-12wt%, the pH of aqueous solution is adjusted to 7-8, is placed at room temperature for.
The present invention proposes a kind of high-sequential nanofiber, is the assembling side using above-mentioned high-sequential nanofiber
Method obtains.
The small-molecular peptides for caning be assembled into high-sequential nanofiber and assembling structure high-sequential of the embodiment of the present invention are received
The advantageous effect of method of rice fiber is：The small-molecular peptides sequence for caning be assembled into high-sequential nanofiber of the embodiment of the present invention
It is made of successively hydrophobic alkyl chain, beta sheet amino acid, hydrophilic amino acid and N- Amino End Groups sealing end；Preparation method is to use
The polypeptide solid-state reaction method synthesis of FMOC Preservation tactics, vector resin is Rink Amide-AM Resin resins, by small-molecular peptides
Segment extends successively on vector resin from C-terminal to N-terminal, and reaction condition is simple, mild, and obtained small-molecular peptides have amphiphilic
Property, it can be assembled into the high-sequential nanofiber with good biocompatibility.The high-sequential Nanowire of the embodiment of the present invention
The assemble method of dimension is that above-mentioned small-molecular peptides are dissolved in water to concentration for 0.05wt%-12wt%, by the pH of aqueous solution
7-8 is adjusted to, is placed at room temperature for, reaction condition is simple, mild, and obtained high-sequential nanofiber has good biocompatibility.
Description of the drawings
It in order to illustrate the technical solution of the embodiments of the present invention more clearly, below will be to needed in the embodiment attached
Figure is briefly described, it should be understood that the following drawings illustrates only certain embodiments of the present invention, therefore is not construed as pair
The restriction of scope, for those of ordinary skill in the art, without creative efforts, can also be according to this
A little attached drawings obtain other relevant attached drawings.
Fig. 1 is the structure chart for the small-molecular peptides that the embodiment of the present invention 1 provides；
Fig. 2 is the mass spectrogram for the small-molecular peptides that the embodiment of the present invention 1 provides；
Fig. 3 is the SEM figures for the high-sequential nanofiber that the embodiment of the present invention 2 provides；
Fig. 4 is the petrographic microscope figure for the high-sequential nanofiber that the embodiment of the present invention 3 provides；
Fig. 5 is the structure chart for the small-molecular peptides that the embodiment of the present invention 4 provides；
Fig. 6 is the mass spectrogram for the small-molecular peptides that the embodiment of the present invention 4 provides；
Fig. 7 is the SEM figures for the high-sequential nanofiber that the embodiment of the present invention 5 provides.
Specific embodiment
It, below will be in the embodiment of the present invention to make the purpose, technical scheme and advantage of the embodiment of the present invention clearer
Technical solution be clearly and completely described.The person that is not specified actual conditions in embodiment, builds according to normal condition or manufacturer
The condition of view carries out.Reagents or instruments used without specified manufacturer is the conventional production that can be obtained by commercially available purchase
Product.
The small-molecular peptides for caning be assembled into high-sequential nanofiber to the embodiment of the present invention and assembling structure height below
The method of ordered nano-fibers is specifically described.
The embodiment of the present invention provides a kind of small-molecular peptides for caning be assembled into high-sequential nanofiber, while good water solubility,
Biocompatibility is excellent.Small-molecular peptides sequence is successively by hydrophobic alkyl chain, beta sheet amino acid, hydrophilic amino acid and N- ends ammonia
Base sealing end is formed, and sequence general formula is hydrophobic alkyl chain+beta sheet amino acid+hydrophilic amino acid-CONH2.Wherein, hydrophobic alkane
Base chain includes one kind in stearic acid (C18), palmitic acid (C16), myristic acid (C14), lauric acid (C12), hydrophobic alkyl chain
General structure beBeta sheet amino acid be the hydrophobic amino acid with β-pleated sheet, specifically include alanine (A),
One or more of valine (V), glycine (G), phenylalanine (F)；Hydrophilic amino acid includes glutamic acid (E), relies ammonia
One or more of sour (K).
In the present embodiment, the structural formula of small-molecular peptides can be C15H31CO-AAAAAKK-CONH2Or C15H31CO-
VVVVKKK-CONH2, wherein, A is alanine, and V is valine, and K is lysine.
The embodiment of the present invention provides a kind of preparation side of the above-mentioned small-molecular peptides for caning be assembled into high-sequential nanofiber
Method is synthesized using the polypeptide solid-state reaction method of FMOC Preservation tactics, and vector resin is Rink Amide-AM Resin resins (tree
Fat degree of substitution 0.6-0.65mmol/g), small-molecular peptides segment is extended successively on vector resin from C-terminal to N-terminal, it is specific to synthesize
Step is as follows：
S1, the Rink Amide-AM Resin resins for weighing 0.5-1.5g are placed in Solid-phase synthesis peptides column, with N, N-
Dimethylformamide (DMF) washs at least three times, empties solvent, then is swollen 0.5- with the n,N-Dimethylformamide of 15-40ml
After 2h, solvent is emptied.
The FMOC groups that alpha-amido is protected in S2, removing Rink Amide-AM Resin resins, specifically to synthesis in solid state
It is 15%-40% that volume ratio is added in column：Piperidines (Piperiding)/n,N-Dimethylformamide solution of 60%-85%, is stirred
After mixing 20-60min, with N, N- dimethylformamides wash at least three times, empty solvent.
S3, into synthesis in solid state column add in 2-4 times of molar equivalent beta sheet amino acid (such as Fmoc-Ala-OH,
Fmoc-Lys (Boc)-OH), benzotriazole-N, N, N ', N '-tetramethylurea fluorophosphate (HBTU), three nitrogen of 1- hydroxy benzos
Azoles (HOBT) and volume ratio are 5%-20%：The N of 80%-95%, N- diisopropylethylamine (DIEA)/N, N- dimethyl formyls
Amine aqueous solution is slowly stirred 1.5-4h.
Whether S4, beta sheet amino acid of taking a sample to check have been connected on resin, are specifically taken a small amount of resin, are put into ninhydrin
Methanol solution (0.01mg/ml) in ebuillition of heated test color within 3-5 minutes, if non-discolouring, show that beta sheet amino acid has been connected on
On resin；
If S5, beta sheet amino acid have been connected on resin, repeat step S2-S4, until connect on resin corresponding β-
Fold amino acid and quantity；
The FMOC groups that alpha-amido is protected in S6, removing Rink Amide-AM Resin resins, specifically to synthesis in solid state
It is 15%-40% that volume ratio is added in column：The piperidines of 60%-85%/n,N-Dimethylformamide solution stirs 20-60min
Afterwards, washed at least three times with n,N-Dimethylformamide, empty solvent；
S7, hydrophilic amino acid, the HBTU that 1.5-4 times of molar equivalent is added in into synthesis in solid state column, 2-3 times mole is worked as
The HOBT and volume ratio of amount are 5%-20%：The n,N-diisopropylethylamine of 80%-95%/n,N-Dimethylformamide solution,
Stir 1.5-4h；
Whether S8, hydrophilic amino acid of taking a sample to check have been connected on resin, are specifically taken a small amount of resin, are put into ninhydrin
Methanol solution (0.01mg/ml) in ebuillition of heated test color within 3-5 minutes, if non-discolouring, show that hydrophilic radical has been connected on tree
On fat；
If S9, hydrophilic amino acid have been connected on resin, above-mentioned S6-S8 is repeated, until having met corresponding parent on resin
Aqueous amino acid and quantity；
S10, hydrophobic alkyl chain, HBTU, HOBT and DIEA/ that 2.5-4 times of molar equivalent is added in into synthesis in solid state column
DMF solution, stirring；
Whether S11, hydrophobic alkyl chain of taking a sample to check have been connected on resin, are specifically taken a small amount of resin, are put into ninhydrin
Methanol solution (0.01mg/ml) in ebuillition of heated test color within 3-5 minutes, if non-discolouring, show that hydrophobic alkyl chain has been connected on tree
On fat；
If S12, hydrophobic alkyl chain have been connected on resin, respectively with n,N-Dimethylformamide, methanol and dichloromethane
(DCM) resin is washed, after normal-temperature vacuum drying, obtains dry resin；
S13, cut polypeptide, it is 94%-96% that volume ratio is added in specifically in the dry resin into synthesis in solid state column：
2%-3%：Trifluoroacetic acid (TFA)/H of 2%-3%2O/ tri isopropyl silanes (TIS) mixing liquid, stirring at normal temperature 1.5-4h,
Collection cuts liquid, and then concentrated by rotary evaporation to thick liquid is added dropwise in cold ether and precipitates, supernatant is removed after centrifugation, often
Dry more than the 12h of temperature, obtains small-molecular peptides.
The embodiment of the present invention provides a kind of assemble method of high-sequential nanofiber, comprises the following steps：
The small-molecular peptides for caning be assembled into high-sequential nanofiber are dissolved in the water, form concentration as 0.05wt%-
The aqueous solution of 12wt%；
The pH of aqueous solution is adjusted to 7-8, obtains assembly；
Assembly is placed at room temperature for a period of time, obtains high-sequential nanofiber.
In above process, above-mentioned small-molecular peptides can be dissolved in the water, forms concentration as 0.05wt%-
The aqueous solution of 0.1wt% (low concentration), then the pH of aqueous solution is adjusted to 7-8, assembly is obtained, finally obtains high-sequential Nanowire
Dimension.Above-mentioned small-molecular peptides can also be dissolved in the water, form the aqueous solution that concentration is 7wt%-12wt% (high concentration), then
The pH of aqueous solution is adjusted to 7-8, in 70 DEG C of -90 DEG C of heating 1-2h, assembly is obtained, finally obtains high-sequential arranged in parallel and receive
Rice fiber.
The embodiment of the present invention provides a kind of high-sequential nanofiber, is using above-mentioned high-sequential nanofiber
Assemble method obtains.
The feature and performance of the present invention are described in further detail with reference to embodiments.
Each raw material sources approach in following embodiment：Rink Amide-AM Resin resin (resin degree of substitution
0.625mmol/g), alanine (Fmoc-Ala-OH), alpha-amido and the side group that a-amino acid is protected by 9-fluorenylmethyloxycarbonyl
Amino protected respectively by 9-fluorenylmethyloxycarbonyl and tert-butyl carbonyl lysine (Fmoc-Lys (Boc)-OH), benzotriazole
- N, N, N ', N '-tetramethylurea fluorophosphate (HBTU) and 1- hydroxy benzo triazoles (HOBT) buy in gill it is biochemical (on
Sea) Co., Ltd.
Palmitic acid, piperidines (Piperiding), ninhydrin, N,N-dimethylformamide (DMF), methanol, dichloromethane
(DCM) buy in Chinese medicines group.
Trifluoroacetic acid (TFA), N, N- diisopropylethylamine (DIEA) are bought in Aladdin (aladdin).
Tri isopropyl silane (TIS) is bought in Sa En chemical technologies (Shanghai) Co., Ltd..
Embodiment 1
The present embodiment provides a kind of small-molecular peptides, structural formula C15H31CO-AAAAAKK-CONH2, specific synthesis step is such as
Under：
1) 1g resins are weighed to be placed in Solid-phase synthesis peptides column, is washed three times with DMF, solvent is emptied, then with 20ml's
Solvent is emptied after DMF swellings 1h.
2) the FMOC groups that alpha-amido is protected in RinkAmide-AMResin resins are removed.It is added in into synthesis in solid state column
20%Piperiding/DMF (V/V) after stirring 30min, is washed three times with DMF, empties solvent.
3) add in Fmoc-Lys (Boc)-OH's, HBTU, HOBT and 2mlDIEA of 3 times of molar equivalents into synthesis column
DMF solution 20ml, is slowly stirred 2h.
4) a small amount of resin in taking 3) is put into ebuillition of heated 3-5 minutes in the methanol solution (0.01mg/ml) of ninhydrin
Color is tested, if non-discolouring, shows that amino acid has been connected on resin.
5) repeat it is above-mentioned 2), 3), 4) step, until having connect corresponding amino acid quantity.
6) the FMOC groups that alpha-amido is protected in Fmoc-Ala-OH are removed.20% is added in into synthesis in solid state column
Piperiding/DMF (V/V) after stirring 30min, is washed three times with DMF, empties solvent.
7) Fmoc-Lys (Boc)-OH, HBTU of 2.4 times of molar equivalents of 2 times of molar equivalents is added in into synthesis column,
The DMF solution 20ml of HOBT and 2mlDIEA, is slowly stirred 2h.
8) a small amount of resin in taking 7) is put into ebuillition of heated 3-5 minutes in the methanol solution (0.01mg/ml) of ninhydrin
Color is tested, if non-discolouring, shows that amino acid has been connected on resin.
9) repeat it is above-mentioned 6), 7), 8) step, until having met corresponding amino acid quantity
10) DMF solution of the palmitic acid of 3 times of molar equivalents, HBTU, HOBT and 2mlDIEA is added in into synthesis column
20ml is slowly stirred 8h.
11) a small amount of resin in taking 10) is put into the methanol solution (0.01mg/ml) of ninhydrin ebuillition of heated 3-5 points
Clock tests color, if non-discolouring, shows that palmitic acid has been connected on resin.
12) respectively with DMF, methanol and DCM washing resins, after normal-temperature vacuum drying, the dry resin obtained is for use.
13) cut polypeptide.The TFA/H of 30ml is added in into the dry resin synthesis column 12) obtained2O/TIS (V/V/V=
95%/2.5%/2.5%) mixing liquid, stirring at normal temperature 2h, collection cuts liquid concentrated by rotary evaporation to thick liquid, then by it
It is added dropwise in cold ether and precipitates, supernatant is removed after centrifugation, air drying 12h obtains small-molecular peptides.
Small-molecular peptides obtained by the present embodiment are detected, Fig. 1 is the structure chart of the small-molecular peptides obtained by the present embodiment；
Fig. 2 is the mass spectrogram of the small-molecular peptides obtained by the present embodiment.The result shows that：The actual molecular weight and theoretical molecular weight of small-molecular peptides
It is consistent, small-molecular peptides C15H31CO-AAAAAKK-CONH2。
Embodiment 2
The present embodiment provides a kind of high-sequential nanofibers, are obtained in accordance with the following methods：
By small-molecular peptides C made from embodiment 115H31CO-AAAAAKK-CONH2Concentration is dissolved in water to as 0.1wt%,
Obtain aqueous solution.
Aqueous solution pH is adjusted to 7.4, obtains assembly.
Assembly is being placed at room temperature for a period of time, is obtaining high-sequential nanofiber.
High-sequential nanofiber obtained by the present embodiment is detected, Fig. 3 is the high-sequential obtained by the present embodiment
The SEM figures of nanofiber.The result shows that：Small-molecular peptides C15H31CO-AAAAAKK-CONH2Really high-sequential can be assembled into
Nanofiber.
Embodiment 3
The present embodiment provides a kind of nanofibers with orientation, are obtained in accordance with the following methods：
By small-molecular peptides C made from embodiment 115H31CO-AAAAAKK-CONH2Concentration is dissolved in water to as 10wt%,
Obtain aqueous solution.
Aqueous solution pH is adjusted to 7.4,70 DEG C of -90 DEG C of heating 1-2h and obtains assembly.
Assembly is being placed at room temperature for a period of time, is obtaining the nanofiber with orientation.
The nanofiber obtained by the present embodiment with orientation is detected, Fig. 4 is the high-sequential obtained by the present embodiment
The petrographic microscope figure of nanofiber.The result shows that：Small-molecular peptides C15H31CO-AAAAAKK-CONH2Really tool can be assembled into
There is a nanofiber of orientation, and the small-molecular peptides C from side illustration15H31CO-AAAAAKK-CONH2The nanofiber being assembled into
With property arranged in parallel.
Embodiment 4
The present embodiment provides a kind of small-molecular peptides, structural formula C15H31CO-VVVVKKK-CONH2, specific synthesis step is such as
Under：
1) 1g resins are weighed to be placed in Solid-phase synthesis peptides column, is washed three times with DMF, solvent is emptied, then with 20ml's
Solvent is emptied after DMF swellings 1h.
2) the FMOC groups that alpha-amido is protected in RinkAmide-AMResin resins are removed, are added in into synthesis in solid state column
20%Piperiding/DMF (V/V) after stirring 30min, is washed three times with DMF, empties solvent.
3) Fmoc-Lys (Boc)-OH of 3 times of molar equivalents is added in into synthesis column, HBTU, HOBT and 2mlDIEA's
DMF solution 20ml, is slowly stirred 2h.
4) a small amount of resin in taking 3) is put into ebuillition of heated 3-5 minutes in the methanol solution (0.01mg/ml) of ninhydrin
Color is tested, if non-discolouring, shows that amino acid has been connected on resin.
5) repeat it is above-mentioned 2), 3), 4) step, until having connect corresponding amino acid quantity.
6) the FMOC groups that alpha-amido is protected in Fmoc-Val-OH are removed.20% is added in into synthesis in solid state column
Piperiding/DMF (V/V) after stirring 30min, is washed three times with DMF, empties solvent.
7) to synthesis column in add in 2 times of molar equivalents Fmoc-Lys (Boc)-OH, the HBTU of 2.4 times of molar equivalents,
The DMF solution 20ml of HOBT and 2mlDIEA, is slowly stirred 2h.
8) a small amount of resin in taking 7) is put into ebuillition of heated 3-5 minutes in the methanol solution (0.01mg/ml) of ninhydrin
Color is tested, if non-discolouring, shows that amino acid has been connected on resin.
9) repeat it is above-mentioned 6), 7), 8) step, until having met corresponding amino acid quantity
10) palmitic acid of 3 times of molar equivalents, the DMF solution of HBTU, HOBT and 2mlDIEA are added in into synthesis column
20ml is slowly stirred 8h.
11) a small amount of resin in taking 10) is put into the methanol solution (0.01mg/ml) of ninhydrin ebuillition of heated 3-5 points
Clock tests color, if non-discolouring, shows that palmitic acid has been connected on resin.
12) resin is washed with DMF, methanol and DCM respectively, after normal-temperature vacuum drying, the dry resin obtained is for use.
13) cut polypeptide, the TFA/H of 30ml is added in into the dry resin synthesis column 12) obtained2O/TIS (V/V/V=
95%/2.5%/2.5%) mixing liquid, stirring at normal temperature 2h, collection cuts liquid concentrated by rotary evaporation to thick liquid, then by it
It is added dropwise in cold ether and precipitates, supernatant is removed after centrifugation, air drying 12h obtains small-molecular peptides.
Small-molecular peptides obtained by the present embodiment are detected, Fig. 5 is the structure chart of the small-molecular peptides obtained by the present embodiment；
Fig. 6 is the mass spectrogram of the small-molecular peptides obtained by the present embodiment.The result shows that：The actual molecular weight and theoretical molecular weight of small-molecular peptides
It is consistent, small-molecular peptides C15H31CO-VVVVKKK-CONH2。
Embodiment 5
The present embodiment provides a kind of high-sequential nanofibers, are obtained in accordance with the following methods：
By small-molecular peptides C made from embodiment 315H31CO-VVVVKKK-CONH2Concentration is dissolved in water to as 0.1wt%,
Obtain aqueous solution.
Aqueous solution pH is adjusted to 7.4, obtains assembly.
Assembly is being placed at room temperature for a period of time, is obtaining high-sequential nanofiber.
High-sequential nanofiber obtained by the present embodiment is detected, Fig. 7 is the high-sequential obtained by the present embodiment
The SEM figures of nanofiber.The result shows that：Small-molecular peptides C15H31CO-AAAAAKK-CONH2Really high-sequential can be assembled into
Nanofiber.
In conclusion the small-molecular peptides for caning be assembled into high-sequential nanofiber of the embodiment of the present invention and its preparation side
Method, reaction condition is simple, mild, and obtained small-molecular peptides have amphipathic, can be assembled into the height with good biocompatibility
Spend ordered nano-fibers.The high-sequential nanofiber and its assemble method of the embodiment of the present invention, reaction condition is simple, mild,
Obtained high-sequential nanofiber has good biocompatibility.
Embodiments described above is part of the embodiment of the present invention, instead of all the embodiments.The reality of the present invention
The detailed description for applying example is not intended to limit the scope of claimed invention, but is merely representative of the selected implementation of the present invention
Example.Based on the embodiments of the present invention, those of ordinary skill in the art are obtained without creative efforts
Every other embodiment, belongs to the scope of protection of the invention.
Claims (10)
1. a kind of small-molecular peptides for caning be assembled into high-sequential nanofiber, which is characterized in that the small-molecular peptides sequence is successively
It is made of hydrophobic alkyl chain, beta sheet amino acid, hydrophilic amino acid and N- Amino End Groups sealing end.
2. the small-molecular peptides according to claim 1 for caning be assembled into high-sequential nanofiber, which is characterized in that described to dredge
Water alkyl chain includes one kind in stearic acid, palmitic acid, myristic acid, lauric acid.
3. the small-molecular peptides according to claim 1 for caning be assembled into high-sequential nanofiber, which is characterized in that the β-
Folding amino acid is the hydrophobic amino acid with beta sheet, is specifically included in alanine, valine, glycine, phenylalanine
It is one or more of.
4. the small-molecular peptides according to claim 1 for caning be assembled into high-sequential nanofiber, which is characterized in that the parent
Aqueous amino acid includes one or more of glutamic acid, lysine.
5. the small-molecular peptides according to claim 1 for caning be assembled into high-sequential nanofiber, which is characterized in that described small
The structural formula of molecular peptide is C15H31CO-AAAAAKK-CONH2Or C15H31CO-VVVVKKK-CONH2, wherein, A is alanine, V
For valine, K is lysine.
6. a kind of preparation method of the small-molecular peptides as described in claim 1 for caning be assembled into high-sequential nanofiber, special
Sign is, is to be synthesized using the polypeptide solid-state reaction method of FMOC Preservation tactics, vector resin is Rink Amide-AM Resin trees
Fat extends small-molecular peptides segment successively on the vector resin from C-terminal to N-terminal.
7. the preparation method of the small-molecular peptides according to claim 6 for caning be assembled into high-sequential nanofiber, feature
It is, comprises the following steps：
S1, Rink Amide-AM Resin resins are placed in Solid-phase synthesis peptides column, are washed with n,N-Dimethylformamide,
Solvent is emptied, then is swollen with n,N-Dimethylformamide, empties solvent；
S2, piperidines/n,N-Dimethylformamide solution, stirring, with N, N- dimethyl formyls are added in into the synthesis in solid state column
Amine swelling washing, empties solvent；
S3, beta sheet amino acid, benzotriazole-N, N, N are added in into the synthesis in solid state column ', N '-tetramethylurea fluorophosphoric acid
Salt, 1- hydroxy benzo triazoles and n,N-diisopropylethylamine/n,N-Dimethylformamide solution, stirring；
Whether S4, beta sheet amino acid of taking a sample to check have been connected on resin；
If S5, beta sheet amino acid have been connected on resin, step S2-S4 is repeated, until having connect corresponding beta sheet on resin
Amino acid and quantity；
S6, piperidines/n,N-Dimethylformamide solution, stirring, with N, N- dimethyl formyls are added in into the synthesis in solid state column
Amine washs, and empties solvent；
S7, hydrophilic amino acid, benzotriazole-N, N, N are added in into the synthesis in solid state column ', N '-tetramethylurea fluorophosphoric acid
Salt, 1- hydroxy benzo triazoles and n,N-diisopropylethylamine/n,N-Dimethylformamide solution stir 1.5-4h；
Whether S8, hydrophilic amino acid of taking a sample to check have been connected on resin；
If S9, hydrophilic amino acid have been connected on resin, above-mentioned S6-S8 is repeated, until having connect corresponding hydrophily on resin
Amino acid and quantity；
S10, hydrophobic alkyl chain, benzotriazole-N, N, N are added in into the synthesis in solid state column ', N '-tetramethylurea fluorophosphoric acid
Salt, 1- hydroxy benzo triazoles and n,N-diisopropylethylamine/n,N-Dimethylformamide solution, stirring；
Whether S11, hydrophobic alkyl chain of taking a sample to check have been connected on resin；
If S12, hydrophobic alkyl chain have been connected on resin, tree is washed with n,N-Dimethylformamide, methanol and dichloromethane respectively
Fat after normal-temperature vacuum drying, obtains dry resin；
S13, trifluoroacetic acid/water/tri isopropyl silane mixing liquid, stirring at normal temperature are added in into the dry resin, collection is cut
Fall liquid, then concentrated by rotary evaporation to thick liquid is added dropwise in cold ether and precipitates, supernatant, air drying are removed after centrifugation.
8. the preparation method of the small-molecular peptides according to claim 7 for caning be assembled into high-sequential nanofiber, feature
It is, checks that the method whether group has been connected on resin is：Resin is taken, is put into ebuillition of heated in the methanol solution of ninhydrin
It tests color within 3-5 minutes, if non-discolouring, shows that group has been connected on resin.
9. a kind of assemble method of high-sequential nanofiber, which is characterized in that it is by such as any one of claim 1 to 5
The small-molecular peptides are dissolved in water to concentration as 0.05wt%-12wt%, and the pH of aqueous solution is adjusted to 7-8, is placed at room temperature for.
10. a kind of high-sequential nanofiber, which is characterized in that it is using high-sequential nanometer as claimed in claim 9
The assemble method of fiber obtains.
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