CN103772484A - Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof - Google Patents

Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof Download PDF

Info

Publication number
CN103772484A
CN103772484A CN201310750012.1A CN201310750012A CN103772484A CN 103772484 A CN103772484 A CN 103772484A CN 201310750012 A CN201310750012 A CN 201310750012A CN 103772484 A CN103772484 A CN 103772484A
Authority
CN
China
Prior art keywords
dipeptides
ace
dipeptide
hmg
activity
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.)
Granted
Application number
CN201310750012.1A
Other languages
Chinese (zh)
Other versions
CN103772484B (en
Inventor
王伟
王楠
张玉
杨玉英
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Zhejiang Academy of Agricultural Sciences
Original Assignee
Zhejiang Academy of Agricultural Sciences
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Zhejiang Academy of Agricultural Sciences filed Critical Zhejiang Academy of Agricultural Sciences
Priority to CN201310750012.1A priority Critical patent/CN103772484B/en
Publication of CN103772484A publication Critical patent/CN103772484A/en
Application granted granted Critical
Publication of CN103772484B publication Critical patent/CN103772484B/en
Expired - Fee Related legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Landscapes

  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

The invention belongs to the field of biotechnology, and particularly relates to a dipeptide which can be bonded with angiotensin converting enzyme, inhibit the activity thereof, and also can inhibit the activity of 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase. Specifically, the invention discloses a dipeptide SD with dual functions of lowering blood pressure and blood fat. The amino acid sequence of the dipeptide SD is as follows: Ser-Asp. The invention also provides an application of the dipeptide SD in preparation of ACE (ASCII-compatible encoding) inhibitory peptide and/or HMG-CoA reductase inhibitory peptide.

Description

There is hypotensive and the bifunctional dipeptides SD of reducing blood-fat and uses thereof
Technical field
The invention belongs to biological technical field, particularly an energy is combined with Zinc metallopeptidase Zace1, suppresses its activity, also can suppress the active dipeptides of 3-hydroxy-3-methylglutaric acid list acyl coenzyme A (HMG-CoA) reductase enzyme.
Background technology
Zinc metallopeptidase Zace1 (Angiotensin converting enzyme, ACE, EC3.4.15.1, in document, former name has kininaseIl, dipeptidyl carboxypeptidase I etc.) be a kind of two carboxypeptidases, to cause a hypertensive key enzyme, it changes into angiotensin II by hydrolytic action hypertensinⅠ, and meanwhile, ACE also can passivation bradykinin, these two kinds of effects all can cause vasoconstriction, thereby cause hypertension.Therefore ACE is considered to cause a hypertensive important factor.Find after deliberation angiotensin converting enzyme inhibitor (ACEI), can reach hypotensive effect by the activity that suppresses ACE.ACE inhibitor is widely used in the diseases such as treatment is cardiovascular, hypertension, heart failure, renal failure.ACE inhibitor is found at first from snake venom, it is found that subsequently the ace inhibitory peptide extracting from foodstuff raw material, as gelatin, casein, fish, Fructus Fici natural gum, α-zein etc. all can be used as the raw material of preparing ace inhibitory peptide.
3-hydroxy-3-methylglutaric acid list acyl coenzyme A (HMG-CoA) reductase enzyme is the key enzyme that in body, catalysis 3-hydroxy-3-methylglutaric acid list acyl coenzyme A (HMG-CoA) generates dihydroxymethyl valeric acid (MVA), this step is the rate-limiting step of synthetic cholesterol in body, is also the target spot of current topmost hyperlipidemia clinical medicine.HMG-CoA reductase inhibitor is one of blood fat reducing function composition and drug screening Main Means.
Hypotensive medicine common are:
1, diuretic(s): represent that medicine has that hydrogen chlorine bites that throat, phenalgin butterfly are suck, in spiral shell the tenth of the twelve Earthly Branches purport etc.
2, sheet receptor-blocking agent: represent that medicine has Pu Cailuoer, metoprolol, atenolol USP 23, nadolol etc.
3, calcium channel blocker: represent that medicine has nifedipine, amlodipine, felodipine, nitrendipine, Lacidipine (62 etc.
4, angiotensin converting enzyme inhibitor: represent that medicine has captopril, Zinadril Briem, lisinopril, enalapril, Yipingshu etc.
5, a mono-receptor-blocking agent: represent that medicine has croak throat, spy to draw and mile tremnbles etc.
6, hypertensin 11 receptor antagonist: represent that medicine has losartan, picks Sha Tan, irbesartan, Candesartan, Irb, telmisartan etc.
The medicine of reducing blood-fat has:
1, fibrate: this type of medicine has fenofibrate, gemfibrozil, bezafibrate etc.Fibrate drug reducing blood lipid is strong, rapid-action, and the effect of triglyceride reducing is stronger than the effect of decreasing cholesterol.
2, trishydroxymethyl glutaryl-CoA-reductase inhibitors: this type of medicine has lovastatin, simvastatin, general Liprevil etc.This type of medicine is take decreasing cholesterol as main, and effect for reducing fat is strong, rapid-action.
3, nicotinic acid class: in this type of medicine, Acipimox is more conventional, the effect that reduces serum levels of triglyceride is stronger than reducing cholesterol.
4, polyunsaturated fatty acid class: comprise various plant seed oils.As rubber seed oil, seed of Radix Oenotherae erythrosepalae, the oil of Silymarin seed and the preparation of ocean fish.This class medicine has reducing blood-fat and reduces the effect of blood viscosity, but effect is gentleer.
5, Pantethine: be the derivative of coenzyme A, have the effect that reduces serum cholesterol, triglyceride and high density lipoprotein increasing-cholesterol.
6, propylene glycol alginate sodium sulfate (PPS): be to scoop up the heparitin marine drug of thing as raw material take marine alga.There is remarkable reduction blood viscosity, vasodilation and reduction blood fat, the effect of rising HSD level.Be mainly used in the control of ischemic cardio cerebrovascular diseases.
7, other blood lipid-lowering medicines: can make serum levels of triglyceride (TG) significantly reduce as ginkgo class (taponin) experimental results show that.
Existing medicine report except Chinese medicine, has hypotensive rarely found with medicine reducing blood lipid simultaneously.Because the target difference of these two kinds of medicine effects.
Summary of the invention
The technical problem to be solved in the present invention is to provide one and has hypotensive and the bifunctional dipeptides SD of reducing blood-fat and uses thereof.
In order to solve the problems of the technologies described above, the invention provides a kind of dipeptides SD, the aminoacid sequence of this dipeptides SD is: Ser-Asp.
The present invention also provides the purposes of above-mentioned dipeptides SD simultaneously: as ace inhibitory peptide and HMG-CoA reductase enzyme inhibiting peptide.
Dipeptides SD of the present invention can be by entrusting synthetic acquisition of the biochemical (Shanghai) Co., Ltd. of gill.
The dipeptides SD that the present invention reports all has for ACE and two targets of HMG-CoA reductase enzyme the activity of inhibition, has the hypotensive and bifunctional characteristic of reducing blood-fat thereby show simultaneously.
Every detection method related in the present invention is specific as follows:
1, ACE suppresses active detection method:
ACE is at 37 ℃, and under the condition that pH value is 8.3, the stand-in Hippuryl-L-Histidyl-L-Leucine (HHL) of catalytic decomposition angiotensin I produces urobenzoic acid (HA), and this material has charateristic avsorption band at ultraviolet 225nm place; In the time adding ACE inhibitor, ACE is suppressed the catalyticing decomposition action of HHL, and the growing amount of urobenzoic acid reduces, and by HPLC method, the variation of measuring the amount that adds the inhibitor front and back urobenzoic acid that generates can calculate the size that suppresses active.
Reaction system is: add respectively successively the ACE of 20 μ L0.1U/mL, 50 μ L ace inhibitory peptides (being SD dipeptides) to bathe 5min 37 ℃ of temperature, then add the HHL substrate of 10 μ L5mM to start the catalyzed reaction of ACE, after 37 ℃ of shaking bath 30min, add the HCl termination reaction of 250 μ L1.0moL/L, system solution is crossed and is carried out RP-HPLC after 0.45 μ m filter membrane and detect the content of analyzing urobenzoic acid (HA).Above-mentioned similarity condition, in the borate buffer of 50 μ L0.1moL/L, (containing NaCl, the pH=8.3 of 0.3moL/L) replaces ACE inhibitor as blank reaction system.
Note: above-mentioned ACE, HHL substrate are all that the borate buffer (containing NaCl, the pH=8.3 of 0.3moL/L) take 0.1moL/L is solvent.
Ace inhibitory peptide (SD dipeptides), is dissolved in different concns in the borate buffer of 0.1moL/L in (containing NaCl, the pH=8.3 of 0.3moL/L) and obtains.
RP-HPLC detects: solvent I is 0.05%(V/V) trifluoroacetic acid (TFA) and triethylamine 0.05%(v/v) (TTA) be dissolved in deionized water, the chromatographically pure second eyeball that solvent II is 100%.The ratio of solvent I and solvent II is 70%:30%(volume ratio), flow velocity is 0.5mL/min, and detection wavelength is 225nm, and detecting column temperature is 30 ℃.
ACE suppresses active and calculates according to following formula:
I%=（A-B）/A×100%
A: the peak area of the urobenzoic acid while not adding small peptide inhibitor;
B: the peak area of the urobenzoic acid while adding small peptide inhibitor;
ACE:1U unit definition is, under standard detection condition, at 37 ℃, catalytic substrate (Hippuryl-L-Histidyl-L-Leucine, HHL) in the 1min time, produces the amount of 1 μ M ACE that urobenzoic acid consumes., be the activity unit of ACE.
2, the external detection method of reducing blood-fat peptide activity:
Chromatographic condition
c1 8chromatographic column (5 μ m, 4.6mm × 250mm).Moving phase is: V (K 2hPO 4-KH 2pO 4): V (methyl alcohol)=85:15, pH7.2, isocratic elution, flow velocity 1mL/min; Detect wavelength 337nm; Sample size 20 μ L; 25 ℃ of column temperatures.
Reaction system experimental procedure
In reaction, the add-on of various components and order are as table 1, and 37 ℃ of water-baths of temperature of reaction, add 200 μ L0.5mol/L NaOH solution termination reactions, by the concentration of NADPH in above-mentioned chromatographic condition measure sample after having reacted.Reaction times is determined according to enzyme control group time gradient.
Table 1 reaction composition composition
Figure BDA0000449539920000031
Figure BDA0000449539920000041
Remarks explanation: the potassium phosphate buffer that reaction buffer is 0.1mol/L, the concentration of HMGR solution is 6.56 μg/mL, HMG-CoA strength of solution is 5.23mmol/mL, NADPH strength of solution is 1mg/mL.
The calculating of experimental result
Figure BDA0000449539920000042
Add after inhibitor, the activity of HMG-CoA reductase enzyme is suppressed, and the amount of substrate reactions reduces.Therefore, can utilize the variation of HPLC assaying reaction front and back NADPH amount to evaluate the inhibiting rate of inhibitor to HMG-CoA reductase enzyme.Calculation formula is:
R=(S inhibitor-S contrast)/(S blank-S contrast) × 100
In formula, R is inhibiting rate (%); S blank, S contrastand S inhibitorbe respectively the peak area (mAU.min) of NADPH in blank group, enzyme control group and inhibitor group.
Advantage of the present invention and positively effect:
(1) this dipeptides SD can suppress the activity of Zinc metallopeptidase Zace1.
According to aminoacid sequence of the present invention, can entrust the biochemical (Shanghai) Co., Ltd. of gill synthetic, thereby obtain ace inhibitory peptide of the present invention (or referred to as dipeptides SD).
(2) ace inhibitory peptide of the present invention (or referred to as dipeptides SD) has HMG-CoA reductase active simultaneously.
Usage and the consumption of ace inhibitory peptide of the present invention and HMG-CoA reductase enzyme inhibiting peptide (or referred to as dipeptides SD) are as follows:
Dipeptides of the present invention is oral type, and consumption is oral, each 1.0g, every day 2～3 times.
Embodiment
Embodiment 1,
1), the ACE of dipeptides SD under the concentration of 1.0mg/mL suppresses active:
Chromatographic condition: solvent I is that the triethylamine (TTA) of 0.05% trifluoroacetic acid (TFA) and 0.05% is dissolved in deionized water (being the trifluoroacetic acid that contains 0.5mL in every liter of solvent I and the triethylamine of 0.5mL), the chromatographically pure second eyeball that solvent II is 100%.The ratio of solvent I and solvent II is 70%:30%(volume ratio), ultimate3000 wears peace liquid chromatograph, and chromatographic column is waters Symmetry C 185 μ m4.6 × 250mm, flow velocity is 0.5mL/min, sample size 10 μ L, detection wavelength is 225nm, detecting column temperature is 30 ℃.
Detection method: by this dipeptides SD obtaining by chemical synthesis, carry out activity and detect (detection method is the same).Now SD concentration is 1.0mg/mL.
Result: it is 75.10% that the ACE of dipeptides SD in the time of 1.0mg/mL suppresses activity.
2), the HMG-CoA reductase active of dipeptides SD under the concentration of 1.0mg/mL:
Chromatographic condition: c1 8chromatographic column (5 μ m, 4.6mm × 250mm).Moving phase is: V (K 2hPO 4-KH 2pO 4): V (methyl alcohol)=85:15, pH7.2, isocratic elution, flow velocity 1mL/min; Detect wavelength 337nm; Sample size 20 μ L; 25 ℃ of column temperatures.
Detection method: by this dipeptides SD obtaining by chemical synthesis, carry out activity and detect (detection method is the same).Now SD concentration is 1.0mg/mL.
Result: the HMG-CoA reductase active of dipeptides SD in the time of 1.0mg/mL is 40.03%.
Embodiment 2,
1), the ACE of dipeptides SD under the concentration of 2.0mg/mL suppresses active:
Chromatographic condition: solvent I is that the triethylamine (TTA) of 0.05% trifluoroacetic acid (TFA) and 0.05% is dissolved in deionized water; Solvent II is 100% chromatographically pure second eyeball.The ratio of solvent I and solvent II is 70%:30%, and ultimate3000 wears peace liquid chromatograph, and chromatographic column is waters Symmetry C 185 μ m4.6 × 250mm, flow velocity is 0.5mL/min, sample size 10 μ L, detection wavelength is 225nm, detecting column temperature is 30 ℃.
Detection method: by this dipeptides SD obtaining by chemical synthesis, carry out activity and detect (detection method is the same).Now SD concentration is 2.0mg/mL.
Result: it is 88.54% that the ACE of dipeptides SD in the time of 2.0mg/mL suppresses activity.
2), the HMG-CoA reductase active of dipeptides SD under the concentration of 2.0mg/mL:
Chromatographic condition: c1 8chromatographic column (5 μ m, 4.6mm × 250mm).Moving phase is: V (K 2hPO 4-KH 2pO 4): V (methyl alcohol)=85:15, pH7.2, isocratic elution, flow velocity 1mL/min; Detect wavelength 337nm; Sample size 20 μ L; Column temperature 25.℃
Detection method: by this dipeptides SD obtaining by chemical synthesis, carry out activity and detect (detection method is the same).Now SD concentration is 2.0mg/mL.
Result: the HMG-CoA reductase active of dipeptides SD in the time of 2.0mg/mL is 66.74%.
By inhibition concentration and activity data in embodiment 1 and embodiment 2, activity and the concentration amount effect relationship of this dipeptides SD are described, this dipeptides SD has ACE inhibition activity simultaneously and HMG-CoA reductase enzyme suppresses to have no report, belongs to the new difunctional peptide that ACE suppresses active and HMG-CoA reductase active that has concurrently.
Finally, it is also to be noted that, what more than enumerate is only several specific embodiments of the present invention.Obviously, the invention is not restricted to above embodiment, can also have many distortion, such as separating obtained SD structure and the derivatize structure thereof of different proteins source degraded.All distortion that those of ordinary skill in the art can directly derive or associate from content disclosed by the invention, all should think protection scope of the present invention.
<110> Zhejiang Academy of Agricultural Science
 
<120> has hypotensive and the bifunctional dipeptides SD of reducing blood-fat and uses thereof
 
<160>?1
 
<210>?1
<211>?2
<212>?PRT
<213> artificial sequence
 
<220>
<223> dipeptides SD
 
<400>?1
Ser?Asp
 

Claims (2)

1. there is the hypotensive and bifunctional dipeptides SD of reducing blood-fat, it is characterized in that: the aminoacid sequence of described dipeptides SD is: Ser-Asp.
2. as claimed in claim 1 have the hypotensive and bifunctional dipeptides SD of reducing blood-fat in the application of preparing in ace inhibitory peptide and/or HMG-CoA reductase enzyme inhibiting peptide.
CN201310750012.1A 2013-12-30 2013-12-30 Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof Expired - Fee Related CN103772484B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201310750012.1A CN103772484B (en) 2013-12-30 2013-12-30 Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201310750012.1A CN103772484B (en) 2013-12-30 2013-12-30 Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof

Publications (2)

Publication Number Publication Date
CN103772484A true CN103772484A (en) 2014-05-07
CN103772484B CN103772484B (en) 2015-05-20

Family

ID=50565297

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201310750012.1A Expired - Fee Related CN103772484B (en) 2013-12-30 2013-12-30 Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof

Country Status (1)

Country Link
CN (1) CN103772484B (en)

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101007841A (en) * 2007-01-29 2007-08-01 浙江大学 Method for separating and purifying ACE inhibition peptide from rice draff and active peptide obtained therefor

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101007841A (en) * 2007-01-29 2007-08-01 浙江大学 Method for separating and purifying ACE inhibition peptide from rice draff and active peptide obtained therefor

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
PETER SOMMER ET AL.: "Proteolysis of Peptide Dendrimers", 《CHEMBIOCHEM》, vol. 10, no. 9, 15 June 2009 (2009-06-15), pages 1527 - 1536 *
肖如武: "蓝蛤蛋白源鲜味肽的制备及分离研究", 《中国优秀硕士学位论文全文数据库》, 31 March 2011 (2011-03-31), pages 1 - 2 *

Also Published As

Publication number Publication date
CN103772484B (en) 2015-05-20

Similar Documents

Publication Publication Date Title
CN103242430A (en) Angiotensin-converting enzyme inhibitory peptide, and preparation method and application thereof
US20110171690A1 (en) Angiotensin converting enzyme inhibitory peptide
CN102573885B (en) Composition having lipolysis-promoting effect
CN102399262B (en) Tripeptides with angiotensin converting enzyme inhibition activity and their use and composition
CN108892710A (en) Asparagus is depressured peptide extract and asparagus Antihypertensive Peptides and its application
CN108484723A (en) The inhibiting peptide of tonin and its preparation method and application in Enteromorpha source
Zhao et al. Exploration, sequence optimization and mechanism analysis of novel xanthine oxidase inhibitory peptide from Ostrea rivularis Gould
CN103755782A (en) Dipeptide ST with double functions of lowering blood pressure and lowering blood fat and application thereof
CN105175500B (en) The active peptides and application thereof prepared using high performance liquid chromatography
CN105200108A (en) Preparation method of hippocampus ACE (angiotension converting enzyme) inhibitory peptide
CN103755781B (en) There is hypotensive and the bifunctional dipeptides GD of reducing blood-fat and uses thereof
CN103739666B (en) Dipeptide QE with dual functions of lowering blood pressure and lowering blood fat and application thereof
CN102643889B (en) Antihypertensive active rapeseed peptide and preparation method and application thereof
CN110467653B (en) Polypeptide for inhibiting angiotensin converting enzyme activity and application thereof
CN105330721A (en) ACE inhibiting peptide and application thereof
CN103736077B (en) Dipeptide ES with double functions of lowering blood pressure and lowering blood fat and application thereof
CN103755783A (en) Dipeptide QD with double functions of lowering blood pressure and lowering blood fat and application thereof
CN103772484B (en) Dipeptide SD with dual functions of lowering blood pressure and blood fat and application thereof
CN103739665B (en) Dipeptide TE with dual functions of lowering blood pressure and lowering blood fat and application thereof
CN103736076B (en) Dipeptide DL with double functions of lowering blood pressure and lowering blood fat and application thereof
CN110396123A (en) Duck protein sources have ACE inhibitory activity p277 SYVP and purposes
Sultana et al. A dipeptide YY derived from royal jelly proteins inhibits renin activity
CN110498836A (en) 6 peptide QYPVEK and purposes with ACE and HMG-CoA reductase inhibitory activity
CN110386960A (en) Duck protein sources have the p277 SPAF and purposes of ACE and HMG-CoA reductase inhibitory activity
CN103012182B (en) N-acyl-sphingosine compound, preparation method and application of N-acyl neuro sphingosine compound

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C14 Grant of patent or utility model
GR01 Patent grant
CF01 Termination of patent right due to non-payment of annual fee
CF01 Termination of patent right due to non-payment of annual fee

Granted publication date: 20150520