WO2004037276A1 - Methode et composition permettant d'ameliorer la fonction vasculaire - Google Patents

Methode et composition permettant d'ameliorer la fonction vasculaire Download PDF

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Publication number
WO2004037276A1
WO2004037276A1 PCT/AU2003/001403 AU0301403W WO2004037276A1 WO 2004037276 A1 WO2004037276 A1 WO 2004037276A1 AU 0301403 W AU0301403 W AU 0301403W WO 2004037276 A1 WO2004037276 A1 WO 2004037276A1
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WO
WIPO (PCT)
Prior art keywords
grape seed
seed extract
polymeric
preparing
grape
Prior art date
Application number
PCT/AU2003/001403
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English (en)
Inventor
Suzanne Roe
Original Assignee
Tarac Technologies Pty Ltd
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 Tarac Technologies Pty Ltd filed Critical Tarac Technologies Pty Ltd
Priority to AU2003273614A priority Critical patent/AU2003273614A1/en
Publication of WO2004037276A1 publication Critical patent/WO2004037276A1/fr

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/87—Vitaceae or Ampelidaceae (Vine or Grape family), e.g. wine grapes, muscadine or peppervine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system

Definitions

  • the present invention relates to a method of enhancing vascular function and to a composition for achieving the same.
  • the composition of the present invention is derived from grape seed extract (GSE).
  • vascular diseases are a scourge of western society. The treatment of vascular disease forms a significant proportion of western healthcare budgets. More importantly, vascular disease contributes to shortened life expectancy and lack of quality of life for many individuals.
  • impaired vascular health includes hypertension, lack of elasticity in vessel walls and impaired circulation in peripheral blood vessels.
  • a further, less immediately obvious, feature of impaired vascular health is the impairment of functioning of vascular endothelial cells.
  • vascular endothelium Located at the interface between blood and a blood vessel wall the vascular endothelium forms the lining of the blood vessel. The endothelium moderates many blood vessel functions and plays a critical role in the mechanics of blood flow and the regulation of coagulation.
  • Damage to the endothelium can lead to atherosclerotic plaques, the build-up of fatty materials within the walls of the arteries, and possibly to more serious cardiac conditions including angina and heart attacks.
  • Green Tea Materials rich in polyphenols, notably Green Tea, which is especially rich in catechins, have been associated with antioxidant effects. Green Tea has, in epidemiological studies, been associated with protection from both heart disease and cancer.
  • the vine grape is a preferred commercial source of polyphenol material because of the relative ease with which the material is available for extraction and the availability of significant volumes of the raw material for processing. Vine products yield polyphenols from the grape skins, grape pulp and from grape seeds. Grape seeds have been identified as having the highest concentrations of available polyphenols.
  • Polyphenols present in GSE may be present as monomers, oligomers and polymeric species. Important polyphenolic monomers are typically flavan-3-ols, for example, (-t-)-catechin, (-)-epicatechin, and the gallic acid esters thereof. Oligomeric and polymeric polyphenolic species include procyanidin compounds.
  • the present invention is concerned with compositions derived from grape seed extract for the promotion of vascular health and process for their production.
  • a method of improving vascular function comprising administering to a subject in need thereof grape seed extract.
  • compositions in accordance with the invention An improvement in vascular function can be demonstrated by monitoring the ability of vascular tissue to relax following administration of compositions in accordance with the invention.
  • Vascular function can also be monitored by the measurement of flow-mediated dilation in blood vessels.
  • a healthy artery which also has a healthy endothelium
  • blood vessels are able to relax if the blood flow increases.
  • the endothelium forms the lining of the blood vessel.
  • the endothelium moderates many blood vessel functions and plays a critical role in the mechanics of blood flow and the regulation of coagulation.
  • the ability of a healthy blood vessel to react to changes in blood flow is called flow mediated dilitation.
  • the grape seed extract is provided in the form of a powdered product.
  • the grape seed extract is added to foodstuff products to form a functional food.
  • GSE As an extract from a natural product that is a known food source GSE is likely to have high consumer acceptance. Further, it is possible to incorporate the GSE into food products in a manner that results in a final food product having attractive taste and flavour characteristics.
  • FMD decreases after a high fat meal and is related to the triglyceride (TG).
  • TG triglyceride
  • High TG levels after food ingestion are also considered a risk factor for coronary disease.
  • GSE is administered within a food product it is thus possible that the improvement in FMD resulting from the ingestion of GSE may serve to counteract the negative effects of high fat foods.
  • the grape seed extract is administered at a rate of 0.2-5g/day. More preferably still said grape seed extract contains 30-90%w/w polyphenols measured as gallic acid equivalent.
  • Polyphenols in grape seed extract may be present as oligomeric and polymeric proanthocyanidins.
  • 60-90%w/w of the total polyphenol component present is oligomeric and polymeric in form and more preferably has 2-16 monomeric units in each polymeric molecule.
  • polyphenols present in the composition of the invention contain less than 1 %w/w f lavonols.
  • a method of preparing a grape seed extract useful for improving vascular function and suitable for administration to a subject in need thereof including the steps of:-
  • the one or more purification steps may further include filtration and evaporation of solvent. DESCRIPTION OF DRAWINGS
  • Figure 1 Illustrates the relaxation effect of a composition in accordance with the invention on aortic ring preparations pre-contracted at approximate EC 50 with noradrenaline.
  • Figure 2 Illustrates the relaxation effect of a composition in accordance with the invention on aortic ring preparations pre-contracted at approximate EC 50 with 8-iso- Prostaglandin E 2 .
  • Figure 3 Illustrates the relaxation effect of a composition in accordance with the invention on mesentery bed preparations pre-contracted at approximate EC 50 with noradrenaline.
  • Figure 4 Shows a comparison of the relaxation effect of a composition in accordance with the invention on myograph preparations precontracted at approximate EC 5 o with noradrenaline.
  • the GSE product of the invention may be prepared via a range of acceptable process conditions.
  • the preparation of the compositions of the invention can be divided into two separate major processes
  • Preparation of the GSE involves the separation of grapes seeds from the remaining grape material and the extraction of the polyphenolic material from the grape seeds themselves followed by subsequent processing steps that render the extract into a form suitable for further processing into food products. Mechanical separation processes are used to remove the seeds from remaining grape material.
  • the extraction of the polyphenolic compounds of interest from the seed products may be achieved by any of a number of chemical extraction processes.
  • solvent extraction of polyphenolics into a suitable solvent such as ethanol and or water may be employed, followed by subsequent solvent removal by evaporation.
  • the extracted product may then be subjected to a spray drying or freeze-drying step to yield a powdered product.
  • the product may be supplied in liquid form.
  • the seed product may be incorporated into food products.
  • Typical food products that might incorporate the GSE formed in accordance with the invention include dairy foods such as yoghurt, milk, ice cream and other dairy products, cereals products including breads, biscuits and breakfast cereals; snack food products; fruit juices and other soft drinks, fruit products and confectionary.
  • the composition of the invention may be supplied as a more traditional form of supplement for example as a tablet or capsule or liquid tincture.
  • Products containing phenolic materials can be characterized by subjecting the materials to a number of standard tests that determine and demonstrate the physiochemical properties of the products.
  • Ferric Reducing Ability of Plasma This test measures the antioxidant capacity of (FRAP Assay) the sample. Reduction of ferric ions to ferrous ions at low pH conditions occurs, proportionate to the reducing ability of the sample. A coloured complex, ferrous-tripyridyl-triazine, is, formed and absorbance at 593 nm measured against known concentration standards.
  • FC Assay Folin-Ciocalteau Assay
  • FC reagent is an oxidizing agent containing a heteropoly phosphotungstate-molybdate complex.
  • a redox reaction occurs with the sample and a colourimetric assessment of the sample is made at 740 nm.
  • HPLC determination of Polymeric Reverse Phase HPLC was used to separate Procyanidins individual components followed by uv detection at 280nm. Quantities of gallic acid, catechin, epicatechin and epicatechin gallate were calculated by comparison with standard solutions.
  • Sub-unit Composition The proanthocyanidin subunit composition, Phloroglucinol Analysis apparent mean degree of polymerisation (mDP) and conversion yield of the samples was determined by acid catalysis in the presence of phloroglucinol.
  • Test conducted on isolated tissue are a rapid method of determining thew usefulness or otherwise of preparations and their effects on vascular tissue.
  • Tissues examined include isolated aortic ring preparations and preparations of the mesenteric vascular bed.
  • the effects of the compositions of the invention on micro- vessels (100-300 micron diameter) were determined by myograph techniques.
  • Tests were conducted using the composition of the invention together with the monomeric, oligomeric and polymeric fractions identified above.
  • Henseleit buffer pH 7.4 at room temperature gassed with carbogen (95% CO 2 :5% O 2 ) and tissue samples prepared for in vitro experimentation as described below.
  • the polymeric fraction was to a small degree insoluble in the inert physiological solvents used.
  • the compounds were made up in saline and placed on an orbital mixer for a minimum of 3 h and the solution was centrifuged for 10 min at 3000 rpm. The supernatant was used as the test solution from which serial dilutions were made.
  • Isolated segments of thoracic aorta (3 mm) were mounted under isometric conditions in organ bath chambers and the GSE were screened for ability to cause dose related endothelium-dependant relaxation in vessels precontracted at approximate EC 50 values for noradrenaline and 8- so-Prostaglandin E 2 as determined previously for WKY rats.
  • the EC 50 value is defined as the dose of compound that elicits 50% of the maximally induced effect of a compound - in this case 50% of the total relaxation by the compound after vascular tissue had been precontracted by an agonist.
  • the experimental protocol involved the following. Rats were sacrificed and the descending thoracic aorta isolated, cleared of adhering tissue and cut into rings, approximately 3 mm in length. The rings were mounted in stainless steel stirrups and place in a 15 ml organ bath chamber. The tissues were bathed in Krebs-Henseleit buffer solution comprised of the following: (mM) 113 NaCI, 4.8 KCI, 1.2 KH 2 PO , 1.2 MgSO 4 , 25 NaHCO 3 , 2.5 CaCI 2 , 11.2 glucose and ascorbic acid (0.57) in deionised MilliQ treated water. The buffer solution was continuously bubbled with carbogen and maintained at 37°C.
  • the aortic rings were allowed to equilibrate for 60 min under 4 g of resting tension before contracting with KCI to test the tissue viability.
  • the rings were pre-contracted with approximate EC 50 dose of noradrenaline or 8-/so-Prostaglandin E 2 .
  • Test GSE were prepared as described above and added cumulatively to the bath. The change in tension was monitored by a computer based data acquisition system (MP100WSW High performance data acquisition unit, BIOPAC Systems Inc.)
  • the superior mesentery artery was cannulated and flushed with heparin-saline.
  • the entire mesenteric bed, including the intestinal tract was removed and the mesenteric vascular bed was carefully stripped from the gut.
  • the mesenteric preparation was mounted in a 15 mL organ bath and continuously perfused with oxygenated Krebs- Henseleit buffer. After 30 minutes of equilibration, tissue viability was assessed by cumulative intra-luminal injection of various agonists (KCI, noradrenaline).
  • KCI noradrenaline
  • the pressure was raised by the addition of noradrenaline at approximate EC 50 in the bath perfusate.
  • Vasorelaxation responses to the composition of the invention as well as the derived fractions of the composition were determined by measuring the extent of pressure reduction. The pressure changes were monitored using MLT844 Physiological Pressure Transducer connected to a pressure amplifier (DA100C, BIOPAC Systems Inc) and a computer based data acquisition system (MP100WSW High performance data acquisition
  • the results of the tests are depicted in figure 3.
  • the preparation of the invention demonstrates the ability of the composition of the invention to relax smaller resistance arteries and to mediate the development of higher pressures in smaller arteries.
  • the results are indicative of the ability of the compositions of the invention to lower blood pressure in vivo.
  • the third arcade of mesenteric arteries of 250-350 micron diameter was cut at the distal end and transferred to the chamber of the myograph and mounted.
  • the vessels were gassed in a closed chamber with carbogen, 37°C for 45 minutes. Normalised vessels were allowed to stabilise for 15 min before testing viability with KCL (40 mM), noradrenaline and acetylcholine. Only vessels that exhibited a contraction greater than 4 mN when exposed to noradrenaline and 25% relaxation to acetylcholine (in the presence of EC 50 of noradrenaline) were used for further experimentation.
  • Myography allows the evaluation of the effect of the compounds of the invention on the contractility properties of micro-vessels.
  • the results of the tests, depicted graphically in figure 4 demonstrate the significant vaso- reactivity of the composition of the invention.
  • compositions of the invention demonstrate the ability of the compositions of the invention to induce vaso-relaxation in large blood vessels, in resistance arteries and in micro-vessels.
  • the compositions of the invention therefore clearly have the ability to lower blood pressure and/or increase peripheral circulation of blood in smaller vessels.
  • the trial was 12 weeks long and consisted of 3 four-weeks periods in a double-blind randomised crossover with control and active ingredients in 240g of yoghurt. Active ingredients consisted of 2g/day of grape seed extract (GSE) in the yoghurt. Blood samples and vascular compliance measures were taken at baseline and at the end of each period.
  • the background diet was a low polyphenol, low quercetin diet. This was achieved by restricting tea and coffee to a maximum of 2 cups per day, restricting apples to one/day and forbidding red wine and onions throughout the 12 weeks.
  • Flow mediated dilatation FMD was measured using ultrasound, vascular compliance using radial pulse analysis (HDI).
  • Brachial artery ultrasonography was carried out in patients after a 12-hour fast and after resting supine for at least 15 minutes in a quiet, temperature controlled room (21 to 25°C).
  • Endothelium-dependent post-ischemic flow-mediated dilation (FMD) and endothelium-independent glyceryl trinitrate (GTN) mediated dilation (GTNMD) were measured During the ultrasound procedure, subjects rested supine in a quiet, temperature-controlled (24°C) room. The left arm was supported comfortably in extension and supination.
  • a high-resolution 12 mHz linear array transducer connected to an Acuson Aspen System (Acuson Pty Ltd., Mountain View, CA, USA) was employed.
  • Grape seed extract alone produced an absolute 1.1 % greater dilatation compared with control (p ⁇ 0.05). GTN induced dilatation was not influenced by GSE.

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  • Health & Medical Sciences (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
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  • Engineering & Computer Science (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
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  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
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  • Alternative & Traditional Medicine (AREA)
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  • Microbiology (AREA)
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  • Heart & Thoracic Surgery (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines Containing Plant Substances (AREA)

Abstract

L'invention concerne une méthode d'amélioration de la fonction vasculaire caractérisée en ce qu'elle consiste à administrer à un sujet nécessitant ce traitement un extrait de pépins de raisin.
PCT/AU2003/001403 2002-10-23 2003-10-23 Methode et composition permettant d'ameliorer la fonction vasculaire WO2004037276A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU2003273614A AU2003273614A1 (en) 2002-10-23 2003-10-23 Method and composition for enhancing vascular function

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Application Number Priority Date Filing Date Title
AU2002095212 2002-10-23
AU200295212 2002-10-23

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WO2004037276A1 true WO2004037276A1 (fr) 2004-05-06

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0348781B1 (fr) * 1988-06-28 1992-09-30 TECNOFARMACI S.p.A. Fractions oligomériques de procyanidole, les procédés pour leur préparation et compositions pharmaceutiques les contenant
WO1998033494A1 (fr) * 1997-02-04 1998-08-06 Kosbab John V Compositions et procedes destines a la prevention et au traitement de maladies degeneratives vasculaires
JP2000279705A (ja) * 1999-03-31 2000-10-10 Shonan Koryo Kk ポリフェノール成分の抽出方法
WO2000078326A1 (fr) * 1999-06-18 2000-12-28 Dry Creek Nutrition, Inc. Procede et composition visant a prevenir ou traiter des effets physiologiques secondaires associes a la cardiopathie

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0348781B1 (fr) * 1988-06-28 1992-09-30 TECNOFARMACI S.p.A. Fractions oligomériques de procyanidole, les procédés pour leur préparation et compositions pharmaceutiques les contenant
WO1998033494A1 (fr) * 1997-02-04 1998-08-06 Kosbab John V Compositions et procedes destines a la prevention et au traitement de maladies degeneratives vasculaires
JP2000279705A (ja) * 1999-03-31 2000-10-10 Shonan Koryo Kk ポリフェノール成分の抽出方法
WO2000078326A1 (fr) * 1999-06-18 2000-12-28 Dry Creek Nutrition, Inc. Procede et composition visant a prevenir ou traiter des effets physiologiques secondaires associes a la cardiopathie

Non-Patent Citations (15)

* Cited by examiner, † Cited by third party
Title
ANN. NY ACAD. SCI., vol. 957, 2000, pages 78 - 89 *
ATHEROSCLEROSIS., vol. 142, no. 1, 1999, pages 139 - 49 *
COLLOQUES-INST. NAT. DE LA RECH. AGRONOMIQUE, vol. 69, no. POLYPHENOS 94, 1995, pages 399 - 400 *
DATABASE CA [online] KNOTEK ET AL., XP002976209, accession no. STN Database accession no. 123:306213 *
DATABASE CA [online] XP002976208, accession no. STN Database accession no. 133:309325 *
DATABASE MEDLINE [online] DELAUNAY ET AL.: "Preparative isolation of polyphenolic copmounds from Vitis vinifera by centrifugal partition chromatography", Database accession no. 12198840 *
DATABASE MEDLINE [online] FITZPATRICK ET AL.: "Vasodilating procyanidis derived from grape seeds", Database accession no. 12074963 *
DATABASE MEDLINE [online] PALMA ET AL.: "Extraction of polyphenolic compounds from grape seeds with near critical carbon dioxide", Database accession no. 10444839 *
DATABASE MEDLINE [online] YAMAKOSHI ET AL.: "Proanthocyanidin-rich extract from grape seeds attenuates the development of aortic atherosclerosis in cholesterol-fed rabbits", Database accession no. 9920515 *
DATABASE MEDLINE [online] YU ET AL.: "Study of anti-atherosclerosic effect of grape seed extract and its mechanism", Database accession no. 12600036 *
DATABASE WPI Section D13 Derwent World Patents Index; Class B04, AN 2001-074403/09 *
J. CHROMATOGR. A., vol. 849, no. 1, 1999, pages 117 - 24 *
J. CHROMATOGR. A., vol. 964, no. 1-2, July 2002 (2002-07-01), pages 123 - 8 *
KAPPAGODA ET AL., CHEMICAL INNOVATION, vol. 30, no. 9, 2000, pages 27 - 31 *
WEI SHENG YAN JIU., vol. 31, no. 4, August 2002 (2002-08-01), pages 263 - 5 *

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