CN102796196A - Polyester-connected deuterohaemin-beta-Ala-His-Thr-Val-Glu-Lys (DhHP-6) and preparation method thereof - Google Patents
Polyester-connected deuterohaemin-beta-Ala-His-Thr-Val-Glu-Lys (DhHP-6) and preparation method thereof Download PDFInfo
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Abstract
The invention belongs to the technical field of biology, and particularly relates to a polyester-connected deuterohaemin-beta-Ala-His-Thr-Val-Glu-Lys (DhHP-6) and a preparation method thereof. The method comprises the following steps of: preparing a polyester-NHS (N-Hydroxysuccinimide) ester under a water-free and oxygen-free condition, mixing with deuterohaemin, and reacting by taking DMSO (Dimethyl Sulfoxide) as a solvent; and after the reaction, precipitating a polymer out of water, centrifuging, collecting a precipitate, purifying and drying to obtain the polyester-connected DhHP-6. A deuterohaemin hexapeptide used in the method belongs to a small molecular peptide, secondary and ternary structures do not exist, and the change of a steric configuration caused in a reacting process does not cause inactivation of enzymes; most of immobilized enzymes are naked outside a carrier, so that contact of enzymes with a substrate is facilitated; polyglycolic acid of a carboxyl end group and a polylactic acid or polylactic acid hydroxyacetic acid copolymer are taken as immobilized materials; covalent coupling immobilized enzymes have the advantage of convenience in recovering and repeatedly utilizing; and the activity of immobilized enzymes is equivalent to 92.2 percent that of naked enzymes, and is basically kept.
Description
Technical field
The invention belongs to biological technical field, be specifically related to ferric iron secondary heme six peptides of a kind of polyester connection and preparation method thereof.
Background technology
Px is present in plant (horseradish, soybean peel), mikrobe (whiterot fungi) and the animal body (POD in the milk) usually, is prothetic group with the protoheme, has multiple biological function, mainly through catalysis H
2O
2Other materials are produced oxygenizement.Px has been widely used in aspects such as food-processing, medical science detection, lignin degradation, agricultural byproducts storage, biocatalysis and WWT at present.
Natural px has extraction and processing difficulties usually, poor stability, and shortcomings such as application cost height have limited its application.Secondary heme six peptides (deuterohaemin-β-Ala-His-Thr-Val-Glu-Lys; DhHP-6) be to be foundation with little enzyme and natural ascorbate peroxidase enzymatic structure; A kind of small peptide that contains deuterohemin of design is a kind of mimics of peroxidase preferably; Have with the identical active site of px, having confirmed has certain curative effect to cataract, type-II diabetes, myocardial ischemia-reperfusion injury etc.; This small peptide analogue enztme molecular weight can develop into anti-oxidation medicine efficiently less than 3000; The same with similar px have a very strong catalytic activity; But its enzyme stability has then improved a plurality of orders of magnitude because of its simple structure; Because the relative simple production of its structure is with low cost, make it possibly become one of important substitute of present px simultaneously.
The covalent coupling immobilized enzyme is that carrier and enzyme interconnect through chemical bond, same data by MoM and MEI, and covalent linkage coupling method immobilized enzyme is the strongest a kind of method of bonding force between enzyme and the carrier.Normally amino residual on the enzyme surface is carried out chemical reactions such as diazotization, alkylation, amination and curing with some functional group on the carrier, enzyme is fixed on the carrier.The immobilized enzyme major part of this method is exposed beyond carrier, thus help the contact of enzyme-to-substrate, but because the employed solvent of immobilization, the influence of reaction conditions may be to changing the sterie configuration of enzyme, and then cause the inactivation of enzyme.We employed DhHP-6 belongs to small-molecule peptide, does not have two, a tertiary structure, that is: the change of sterie configuration can not cause the inactivation of enzyme, so we select DhHP-6 is fixed on the polyester molecule.
POLYACTIC ACID, PGTA, POLYACTIC ACID-polyester molecules such as PGTA multipolymer are the synthetic Biodegradable material, and in order to improve the application performance of DhHP-6, the method that we adopt the covalency idol to connect is fixed on DhHP-6 on the above-mentioned polyester molecule.
Vigor through the enzyme after the enzymic activity test discovery immobilization is equivalent to 92.0% of naked enzyme activity, and vigor keeps basically.
Summary of the invention
One of the object of the invention is secondary heme six peptides that provide a kind of polyester with excellent biological compatibility and biological degradability to be connected; It is that (relative molecular mass of above-mentioned materials is 5000～50000g/mol as polyester connection material with biodegradable PGTA (PGA), POLYACTIC ACID (PLA) or polylactic acid-glycolic guanidine-acetic acid multipolymer (PLGA); MWD 1.2～2.0); Adopt the N-hydroxy-succinamide active ester method; The end carboxyl of PLGA, PGA or PLA is formed amido linkage with the amino reaction of the end on the DhHP-6 Methionin, thereby DhHP-6 is incorporated on PLGA, PGA or the PLA.
Ferric iron secondary heme six peptides that the polyester that relates among the present invention connects have suc as formula the general structure shown in the I:
POLYACTIC ACID (m=34～347 wherein; N=0) or PGTA (m=0; N=43～431) or polylactic-co-glycolic acid multipolymer (m=3～312; N=4～387) relative molecular mass is 5000～50000g/mol, and ferrous secondary heme six peptides of trivalent of an amino are removed in-R representative, have suc as formula the constitutional features shown in the II;
As preferred embodiment; The molecular weight of POLYACTIC ACID, polyhydroxylactic acid or polylactic acid-glycolic guanidine-acetic acid multipolymer that ferric iron secondary heme six peptides that above-mentioned polyester connects are used is 5000～50000g/mol; In the multipolymer lactic acid account for lactic acid and oxyacetic acid segment total mole number and 10～90%; In the preferred embodiment; In the multipolymer lactic acid account for lactic acid and oxyacetic acid segment total mole number and 40～60%, in the most preferred embodiment, in the multipolymer lactic acid account for lactic acid and oxyacetic acid segment total mole number and 50%.
Be used to prepare the method for ferric iron secondary heme six peptides that polyester recited above connects, concrete steps are following:
(1) with POLYACTIC ACID, PGTA or polylactic acid-glycolic guanidine-acetic acid multipolymer; NSC 57182 (DCC); N-hydroxy-succinamide (NHS) joins in the anhydrous methylene chloride solution according to the mixed of mol ratio 1:2:2 then, and the concentration range of polyester is 20mg/ml～200mg/ml in the control solution; 20～39 ℃ of temperature of reaction system, stirring reaction 0.5～24h; Reaction finishes to remove by-product of dicyclohexylurea (DCU) with the oil phase filtering with microporous membrane of 0.45 μ m and obtains settled solution;
(2) the gained settled solution is settled out in cold diethyl ether (20 ℃), abandoning supernatant after the centrifugal treating, the thing that settles out dissolves with anhydrous methylene chloride; Cold diethyl ether settles out, and repeats above-mentionedly to settle out, centrifugal, dissolution process 3～5 times;
(3) with the thing vacuum-drying 0.5～24h under 10～35 ℃ of conditions that settles out after the last spinning of step (2), obtain POLYACTIC ACID-NHS, PGTA-NHS or polylactic acid-glycolic guanidine-acetic acid-NHS Acibenzolar;
(4) POLYACTIC ACID-NHS, PGTA-NHS or the polylactic acid-glycolic guanidine-acetic acid-NHS Acibenzolar with 0.4～4g joins in DMSO 99.8MIN. (DMSO) solution of 5～500ml; Secondary heme six peptides that add 1～200 μ L triethylamine and 0.1g again, 10～30 ℃ of reaction 0.5～24h;
(5) reaction settles out polymkeric substance in water after finishing, and is centrifugal, collects the thing that settles out, and again with washing 3～5 times, vacuum-drying 0.5～24h under 10～35 ℃ of conditions promptly obtains ferric iron secondary heme six peptides that polyester connects at last.
Enzyme testing method alive used among the present invention is Wo Xintongfa:
Substrate:
1.5ml concentration is 16.2mg/ml phenol and 1.5ml concentration is phosphate buffer solution (PBS) or the DMSO solution of 0.5mg/ml 4-AAP;
1.5ml, concentration is 1mM H
2O
2PBS (pH=7.0) solution or DMSO solution;
Enzyme liquid: secondary heme six peptides that 0.1ml secondary heme six peptides are connected with polyester.
Under the kinetic measurement pattern of UV-2450, setting sweep time is 30～120s, and the setting absorbing wavelength is 510nm.The vigor of the slope calculating enzyme through fitting a straight line.
A unit vigor is defined as: 25 ℃, during PH=7.0, changing 0.01 with 510nm place absorbancy in the per second is 1 peroxidase activity unit (U).Calculation formula is:
Wherein, △ A510/t is is the fitting formula slope, and this formula provides after by the UV match automatically; The quality (g) of secondary heme six peptides that secondary heme six peptides that W is taken by weighing when being preparation enzyme liquid to be measured or polyester connect, V
TBe the secondary heme six peptide solution TVs (ml) of secondary heme six peptides or polyester connection, V
SThe secondary heme six peptide solution volumes (ml) that secondary heme six peptides that use during for mensuration or polyester connect.
The invention has the advantages that
The immobilized enzyme major part of this method is exposed beyond carrier, so help the contact of enzyme-to-substrate, employed DhHP-6 belongs to small-molecule peptide, does not have two, a tertiary structure, and the change of the sterie configuration that causes in the reaction process can not cause the inactivation of enzyme;
Select for use POLYACTIC ACID, PGTA or the polylactic-co-glycolic acid multipolymer of carboxy blocking to belong to synthesized polymer material; Adopt the mode of covalent coupling immobilized enzyme that secondary heme six peptides are fixed on the polyester molecule; Adopt the form of immobilized enzyme, purpose is to be convenient to reclaim and recycling; The carrier polyester molecule belongs to " natural circulation type fully " Biodegradable material, and the discarded back of material only need simply be buried processing and got final product, and can not cause the pollution of environment.The vigor of the enzyme after the immobilization is equivalent to 92.2% of naked enzyme activity, and vigor keeps basically.
Description of drawings
Ultraviolet concentration-absorbancy the typical curve of Fig. 1: DhHp-6 in DMSO solution; As can be seen from the figure the ultraviolet absorptivity of the DMSO solution of secondary heme six peptides changes with the proportional example of concentration, and this typical curve is used for the coupling efficiency of secondary heme six peptides of POLYACTIC ACID, PGTA or the connection of polylactic acid-glycolic guanidine-acetic acid multipolymer is carried out quantitative analysis;
Fig. 2: be in 200～800nm scope, secondary heme six peptide DMSO solution to be composed scanning entirely, can see that secondary heme six peptides have maximum ultraviolet absorption peak at the 390.50nm place, be the ultraviolet charateristic avsorption band of secondary heme six peptides;
Fig. 3: the DMSO solution that is secondary heme six peptides that in 200～800nm scope, PLGA connected is composed scanning entirely, can see that secondary heme six peptides that polyester connects have the ultraviolet absorption peak of maximum at 392.00nm place; Comparison diagram 2 can find out that red shift takes place in the ultraviolet maximum absorption peak position of secondary heme six peptides, explains that the PLGA molecule has been connected on secondary heme six peptide molecules.
Fig. 4: be the infrared absorpting light spectra of secondary heme six peptides of polyester among the embodiment 1 (PLGA) connection, wherein curve a is PLGA-DhHP-6, and b is pure PLGA, in curve a, and 1650cm
-1There is the carbonyl absorption peak of tangible DhHP-6 amido linkage at the place, among the curve b, does not then detect this peak, explains that DhHP-6 has been coupled on the PLGA;
Fig. 5: the differential scanning calorimetric curve (DSC) that is polyester among the embodiment 1 (PLGA);
Fig. 6: the DSC that is secondary heme six peptides (PLGA-DhHP-6) that polylactic acid-glycolic base lactic acid connects among the embodiment 1;
Fig. 7: be polylactic acid-glycolic base lactic acid and secondary heme six peptide blended DSC among the embodiment 1;
Can find out that through Fig. 5,6, the contrast of 7 DSC spectrogram the melt temperature of PLGA is 55.50 ℃; The melt temperature of secondary heme six peptides (PLGA-DhHP-6) that polyester connects is 61.39 ℃; The melt temperature of secondary heme six peptides of polyester physical mixed is 54.20 ℃, and being linked on secondary heme six peptide molecules of polyester molecule (PLGA) success is described.
Fig. 8: be that the intermediate product PLGA-NHS that polylactic acid-glycolic guanidine-acetic acid link coupled secondary heme six peptides prepare in the process among the embodiment 1 carries out nmr analysis.PLGA-NHS's
1H NMR, test is with deuterochloroform (CDCl
3) be solvent, 2.621 places are the methylene peak of N-hydroxy-succinamide among the figure, 1.545～1.603 is the methyl peak of lactic acid among the PLGA.According to the PLGA molecular weight is 10000, and the block ratio is 50: 50, and the molecular weight of a structural unit is 128, one PLGA
10000Molecule approximately contains 78 methyl, A in theory
δ=2.621/ A
δ=1.545～1.603=4/78 * 3=1/58.5 (A is an integral area).In fact A
δ=2.621/ A
δ=1.545 ~ 1.603=1/55.85, coupling efficiency is 113%, that is, average 1.13 NHS of coupling on each PLGA molecule, coupling efficiency are greater than 100%, and this is because the MWD broad (being about 1.5) or the systematic error of PLGA material cause.Lines not should be grey, should be black, revise!
Fig. 9: be the active maintenance situation of secondary heme six peptides before and after POLYACTIC ACID among the embodiment 2-PGTA connects, as shown in the figure.The kinetics fit equation of naked enzyme is: y=2.13804+0.005x; The kinetics fit equation of PLGA-DhHP-6 is: y=0.03372+0.00376x.According to calculating of formula 3.1 at DMSO:H
2O=1: the vigor of DhHP-6 is in 1 the mixed solvent: 1.51 * 10
4U/ (mgs), the vigor of PLGA-DhHP-6 is: 1.39 * 10
4U/ (mgs)---the amount (0.027mg) that is converted to naked enzyme by 0.25mgPLGA-DhHP-6 is calculated.That is: the vigor of the enzyme after the immobilization is equivalent to 92.0% of naked enzyme activity, and vigor keeps basically.
Embodiment
In order further to understand the present invention, below in conjunction with instance the preferred embodiments of the invention are described, but should be appreciated that these just restriction in order to further specify characteristics of the present invention and advantage rather than patent of the present invention to be required is described.
Used POLYACTIC ACID in this patent, PGTA, the polylactic acid-glycolic guanidine-acetic acid is available from Shandong medical device research institute;
Used secondary heme six peptide DhHP-6 are provided by life science institute of Jilin University, and its preparation method sees Chinese patent for details: 200810050307.7, and publication number: CN101220084A.
The preparation of PLGA-NHS Acibenzolar, (molecular weight is 10000Da to 0.5g PLGA, and wherein lactic acid-polyhydroxylactic acid mol ratio is 50:50; M=31～38, n=26～39), 0.02mg DCC; 0.012g (PLGA:DCC:NHS mol ratio ≈ 1:2:2 is dissolved in the anhydrous methylene dichloride of 10ml NHS, and 20 ℃ of reactions of magnetic agitation are spent the night; Reaction finishes the oil phase filtering with microporous membrane of back with 0.45 μ m, removes by product DCU.Clear liquid settles out in cold diethyl ether (20 ℃), abandoning supernatant after the centrifugal treating, the multiplexing anhydrous methylene chloride dissolving of the thing that settles out; In cold diethyl ether, settle out again; Three times so repeatedly, the thing room temperature that settles out after spinning vacuum-drying 4h obtains PLGA-NHS Acibenzolar 0.46g;
The preparation of secondary heme six peptides that polyester (PLGA) connects, the PLGA-NHS Acibenzolar 0.5g that gets according to embodiment 1 preparation joins among the 5ml DMSO, adds 10 μ L triethylamines and 0.12g DhHP-6 room temperature reaction 8h again.Settle out polymkeric substance in water in reaction end back, centrifugal, collects the thing that settles out, and again with washing 3 times, room temperature vacuum-drying 10h had both got PLGA coupling DhHP-60.56g, and enzyme activity is 1.39 * 10
4U/ (mgs) compares with secondary heme and to have kept 92.0% vigor.
The preparation of PGA-NHS Acibenzolar, (molecular weight is 10000Da to 0.5g PGA, m=0; N=74～105); 0.015g DCC, 0.012g NHS (PGA:DCC:NHS mol ratio ≈ 1:2:2) is dissolved in the anhydrous methylene dichloride of 10ml, and 25 ℃ of reactions of magnetic agitation are spent the night; Reaction finishes the oil phase filtering with microporous membrane of back with 0.45 μ m, removes by product DCU.Clear liquid settles out in cold diethyl ether, abandoning supernatant after the centrifugal treating, and the multiplexing methylene dichloride dissolving of the thing that settles out settles out in cold diethyl ether again, and three times so repeatedly, the thing room temperature that settles out after spinning drying under vacuum overnight obtains PGA-NHS Acibenzolar 0.5g.
The preparation of secondary heme six peptides that polyester (PGA) connects, the PGA-NHS Acibenzolar 0.5g that gets according to embodiment 3 preparations joins among the 5ml DMSO, adds 8 μ L triethylamines and 0.12g DhHP-6 room temperature reaction 10h again.Settle out polymkeric substance in water in reaction end back, centrifugal, collects the thing that settles out, and again with washing 3 times, room temperature vacuum-drying 10h had both got PGA coupling DhHP-60.49g, and enzyme activity is 1.37 * 10
4U/ (mgs) compares with secondary heme and to have kept 90.7% vigor.
The preparation of PLA-NHS Acibenzolar, 0.5g PLA (molecular weight 10000Da, m=55～83; N=0); 0.015g DCC, 0.012g NHS (PLA:DCC:NHS mol ratio ≈ 1:2:2) is dissolved in the anhydrous methylene dichloride of 10ml, 25 ℃ of reactions of magnetic agitation 9h; Reaction finishes the oil phase filtering with microporous membrane of back with 0.45 μ m, removes by product DCU.Clear liquid settles out in cold diethyl ether, abandoning supernatant after the centrifugal treating, and the multiplexing methylene dichloride dissolving of the thing that settles out settles out in cold diethyl ether again, and three times so repeatedly, the thing room temperature that settles out after spinning drying under vacuum overnight obtains PLA-NHS Acibenzolar 0.48g.
The preparation of secondary heme six peptides that polyester (PLA) connects, the PLA-NHS Acibenzolar 0.5g that gets according to embodiment 1 preparation joins among the 5ml DMSO, adds 10 μ L triethylamines again and 0.12g DhHP-6 room temperature reaction spends the night.Settle out polymkeric substance in water in reaction end back, centrifugal, collects the thing that settles out, and again with washing 3 times, room temperature vacuum-drying 10h had both got PLA coupling DhHP-60.53g, and enzyme activity is 1.36 * 10
4U/ (mgs) compares with secondary heme and to have kept 89.8% vigor.
Embodiment 7:
The preparation of PLGA-NHS Acibenzolar, (molecular weight is 5000Da to 0.5g PLGA, and wherein lactic acid-polyhydroxylactic acid mol ratio is 10: 90; M=2～5, n=37～42), 0.04mg DCC; 0.023g (PLGA:DCC:NHS mol ratio ≈ 1:2:2 is dissolved in the anhydrous methylene dichloride of 10ml NHS, and 20 ℃ of reactions of magnetic agitation are spent the night; Reaction finishes the oil phase filtering with microporous membrane of back with 0.45 μ m, removes by product DCU.Clear liquid settles out in cold diethyl ether (20 ℃), abandoning supernatant after the centrifugal treating, the multiplexing anhydrous methylene chloride dissolving of the thing that settles out; In cold diethyl ether, settle out again; Three times so repeatedly, the thing room temperature that settles out after spinning vacuum-drying 4h obtains PLGA-NHS Acibenzolar 0.45g;
The preparation of secondary heme six peptides that polyester (PLGA) connects, the PLGA-NHS Acibenzolar 0.5g that gets according to embodiment 7 preparations joins among the 5ml DMSO, adds 5 μ L triethylamines and 0.12g DhHP-6 room temperature reaction 4h again.Settle out polymkeric substance in water in reaction end back, centrifugal, collects the thing that settles out, and again with washing 3 times, room temperature vacuum-drying 10h had both got PLGA coupling DhHP-60.51g, and enzyme activity is 1.35 * 10
4U/ (mgs) compares with secondary heme and to have kept 89.4% vigor.
Embodiment 9:
The preparation of PLGA-NHS Acibenzolar, (molecular weight is 50000Da to 0.5g PLGA, and wherein lactic acid-polyhydroxylactic acid mol ratio is 90: 10; M=295～330, n=22～66), 0.004mg DCC; 0.002g (PLGA:DCC:NHS mol ratio ≈ 1:2:2 is dissolved in the anhydrous methylene dichloride of 10ml NHS, and 20 ℃ of reactions of magnetic agitation are spent the night; Reaction finishes the oil phase filtering with microporous membrane of back with 0.45 μ m, removes by product DCU.Clear liquid settles out in cold diethyl ether (20 ℃), abandoning supernatant after the centrifugal treating, the multiplexing anhydrous methylene chloride dissolving of the thing that settles out; In cold diethyl ether, settle out again; Three times so repeatedly, the thing room temperature that settles out after spinning vacuum-drying 4h obtains PLGA-NHS Acibenzolar 0.48g;
The preparation of secondary heme six peptides that polyester (PLGA) connects, the PLGA-NHS Acibenzolar 0.5g that gets according to embodiment 9 preparations joins among the 5ml DMSO, adds 10 μ L triethylamines and 0.12g DhHP-6 room temperature reaction 8h again.Settle out polymkeric substance in water in reaction end back, centrifugal, collects the thing that settles out, and again with washing 3 times, room temperature vacuum-drying 10h had both got PLGA coupling DhHP-60.55g, and enzyme activity is 1.36 * 10
4U/ (mgs) compares with secondary heme and to have kept 90.1% vigor.
Claims (6)
1. ferric iron secondary heme six peptides that connect of a polyester, its structural formula is shown in (I):
Wherein the relative molecular mass of POLYACTIC ACID, PGTA or polylactic acid-glycolic guanidine-acetic acid multipolymer is 5000～50000g/mol, and ferrous secondary heme six peptides of trivalent of an amino are removed in-R representative, its structural formula shown in (II),
2. ferric iron secondary heme six peptides that a kind of polyester as claimed in claim 1 connects is characterized in that: m=34～347, represent secondary heme six peptides that POLYACTIC ACID connects during n=0.
3. ferric iron secondary heme six peptides that a kind of polyester as claimed in claim 1 connects is characterized in that: m=0, secondary heme six peptides that PGTA connects o'clock are represented in n=43～431.
4. ferric iron secondary heme six peptides that a kind of polyester as claimed in claim 1 connects is characterized in that: secondary heme six peptides that polylactic acid-glycolic guanidine-acetic acid multipolymer connects o'clock are represented in m=3～312, n=4～387.
5. ferric iron secondary heme six peptides that a kind of polyester as claimed in claim 1 connects is characterized in that: in the polylactic acid-glycolic guanidine-acetic acid multipolymer lactic acid account for lactic acid and oxyacetic acid segment total mole number and 10～90%.
6. the ferric iron secondary heme six peptide preparing methods that connect of the described a kind of polyester of claim 1, concrete steps are following:
(1) with POLYACTIC ACID, PGTA or polylactic acid-glycolic guanidine-acetic acid multipolymer; NSC 57182; N-hydroxy-succinamide NHS joins in the anhydrous methylene chloride solution according to the mixed of mol ratio 1:2:2 then, and the concentration range of polyester is 20mg/ml～200mg/ml in the control solution; 20～39 ℃ of temperature of reaction system, stirring reaction 0.5～24h; Reaction finishes to remove by-product of dicyclohexylurea with the oil phase filtering with microporous membrane of 0.45 μ m and obtains settled solution;
(2) the gained settled solution is settled out in cold diethyl ether, abandoning supernatant after the centrifugal treating, the thing that settles out dissolves with anhydrous methylene chloride; Cold diethyl ether settles out, and repeats above-mentionedly to settle out, centrifugal, dissolution process 3～5 times;
(3) with the thing vacuum-drying 0.5～24h under 10～35 ℃ of conditions that settles out after the last spinning of step (2), obtain POLYACTIC ACID-NHS, PGTA-NHS or polylactic acid-glycolic guanidine-acetic acid-NHS Acibenzolar;
(4) POLYACTIC ACID-NHS, PGTA-NHS or the polylactic acid-glycolic guanidine-acetic acid-NHS Acibenzolar with 0.4～4g joins in the dimethyl sulphoxide solution of 5～500ml; Secondary heme six peptides that add 1～200 μ L triethylamine and 0.1g again, 10～30 ℃ of reaction 0.5～24h;
(5) reaction settles out polymkeric substance in water after finishing, and is centrifugal, collects the thing that settles out, and again with washing 3～5 times, vacuum-drying 0.5～24h under 10～35 ℃ of conditions promptly obtains ferric iron secondary heme six peptides that polyester connects at last.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103965287A (en) * | 2014-05-07 | 2014-08-06 | 吉林大学 | Deuterohemin-beta-Ala-His-Lys(DhHP-3), and preparation method and application thereof |
| CN105137099A (en) * | 2015-09-08 | 2015-12-09 | 长春理工大学 | Method for detecting total cholesterol by utilization of DeuteroheminAlaHisThrValGluLys |
| CN107312065A (en) * | 2017-07-05 | 2017-11-03 | 吉林大学 | The application of ferriporphyrin and its derivative short peptide compound and ferriporphyrin and its derivative short peptide compound |
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| CN101220084A (en) * | 2008-01-25 | 2008-07-16 | 吉林大学 | Polyglycol modifying ferric iron deuterohemin short peptide compound and production method thereof |
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| CN101220084A (en) * | 2008-01-25 | 2008-07-16 | 吉林大学 | Polyglycol modifying ferric iron deuterohemin short peptide compound and production method thereof |
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| GUAN S. 等: "A deuterohemin peptide extends lifespan and increases stress resistance in Caenorhabditis elegans", 《FREE RADICAL RESEARCH》 * |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103965287A (en) * | 2014-05-07 | 2014-08-06 | 吉林大学 | Deuterohemin-beta-Ala-His-Lys(DhHP-3), and preparation method and application thereof |
| CN105137099A (en) * | 2015-09-08 | 2015-12-09 | 长春理工大学 | Method for detecting total cholesterol by utilization of DeuteroheminAlaHisThrValGluLys |
| CN107312065A (en) * | 2017-07-05 | 2017-11-03 | 吉林大学 | The application of ferriporphyrin and its derivative short peptide compound and ferriporphyrin and its derivative short peptide compound |
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