The Science of Resveratrol

Mark Lange, Ph. D.

Market America Director of Quality Control

Important Statement

 This presentation is on the latest research in the areas of longevity, cancer, and heart disease.

 Claims are not being made to treat disease or health conditions with dietary supplements.

 This research is in its infancy; please refrain from drawing conclusions based on preliminary data.

Berry and Fruit Extracts to be Discussed



Resveravine ® – whole grape extract containing resveratrol



BioVin ® Advanced – grape skin extract containing OPCs, anthocyanins, and resveratrol



Vita Blue ® – wild blueberry extract containing anthocyanins

Resveravine

®



A natural extract from grape combining the synergistic biological effects of trans-resveratrol and



-viniferin



A standardized extract with 20% total trans resveratrol and



-viniferin



The most concentrated source of Resveratrol

UV Fungi Copper Ozone Resveratrol is naturally produced by plants as a defense system against infection and oxidative stress due to external attacks.

Resveratrol level time

Sources of

Resveratrol

 Resveratrol is found in  Grapes (only in skin)  Wine  Grape Juice  Peanuts  Blueberries  Bilberries  Cranberries

Benefits of Resveratrol



Induces apoptosis (programmed cell death) in unhealthy cells



Promotes normal nuclear factor kappa B and cyclooxygenase-2 activity



Promotes cardiovascular health

   Normal platelet activity Normal vasorelaxation Antioxidant protection of LDL particles 

Possibly Fights Obesity and Diabetes



Promotes normal activity

of the SIRT-1 gene (involved in calorie restriction, fat mobilization, mitochondrial function, longevity)

Apoptosis

 Apoptosis is programmed cell death. The lifespan of cells have genetic limitations. The apoptotic process safely disposes of old cells and cell fragments and does not trigger inflammation.

Microscope View of Apoptosis

A. Unfertilized embryonic cell receives a biochemical signal to die.

B. Cell starts to shrink as activated enzymes cut up the cytoskeleton and DNA in the nucleus.

C. Cell prepares itself to be engulfed by macrophages.

D. Cell fragments into blobs.

Apoptosis is a Normal Process

    Rapid turnover of white blood cells in the body Remodeling of the developing fetus (fingers and toes) Formation of connections (synapses) between neurons in the brain Menstruation – sloughing of the uterine lining

Apoptosis in Cancer Cells

 Unlike normal cells, cancer cells proliferate rapidly and lose the ability to respond to cell death signals and apoptosis. Resveratrol has been found to inhibit proliferation and induce apoptosis in a number of cancer cell lines.

 Aggarwal BB, Bhardwaj A, Aggarwal RS, Seeram NP, Shishodia S, Takada Y. Role of resveratrol in prevention and therapy of cancer: preclinical and clinical studies. Anticancer Res. 2004;24(5A):2783 2840.

Importance of Apoptosis

 Defective cell cycle regulation may result in the propagation of mutations that contribute to the development of cancer. Resveratrol has been found to induce cell cycle arrest when added to cancer cells grown in culture.  Joe AK, Liu H, Suzui M, Vural ME, Xiao D, Weinstein IB. Resveratrol induces growth inhibition, S-phase arrest, apoptosis, and changes in biomarker expression in several human cancer cell lines. Clin Cancer Res. 2002;8(3):893 903.

Resveratrol

Research



Resveratrol caused significant cytotoxicity and increased apoptosis in malignant hemorrhagic tumors.



Resveratrol blocks the multi-step process of carcinogenesis at various stages: suppressing tumor initiation, promotion and progression.



Resveratrol shown to delay development and reduce the metastasizing capacity of spontaneous mammary tumors in mice due to apoptosis.

NIH Resveratrol Study



National Institutes of Health is currently sponsoring a Phase I Study of Resveratrol in Patients With Surgically Removable Colorectal Cancer

Increased frequency of apoptosis in two tumor cell lines induced by polyphenolic extracts

70 60 50 40 30 20 10 0 0,1 Caco-2 1 µg/ml 3 10 control resveratrol resveravine Poligonum cuspidatum extract 30 20 10 0 80 70 60 50 40 0,1 U937 1 µg/ml 3 10

(Japanese knotweed)

Nuclear factor-kappa B

   is a protein involved in cellular responses to stress (free radicals, UV light, and bacterial or viral attack). NF κ B plays a key role in regulating the immune response to infection.

Incorrect regulation of NF κ B has been linked to cancer, inflammatory and viral infection.

Resveratrol

and NF-kB



Resveratrol was shown to be a potent inhibitor of NF k B by blocking IkB kinase activity

 Cancer Research. 60:3477-3483, 2000. 

This study examined the anti-inflammatory effects of resveratrol on human airway epithelial cells, resveratrol inhibited NF k B

 Am J Physiol Lung Cell Mol Physiol, 2004 .



Resveratrol has been shown to promote normal NF k B activity in several studies.

  Cancer Research. 60:3477-3483, 2000.

Polish Journal of Pharmacology. 53(6):557-69, 2001.

Resveratrol

Inflammation Research

 Cyclooxygenase (COX) – of the chemical messengers (prostaglandins) that cause pain and swelling from inflammation.

Blocking this enzyme impedes the production 

Suppression of COX-2 expression by blocking the activation of MAPKs and AP-1 may represent possible molecular mechanisms responsible for previously reported anti-tumor promoting effects of resveratrol on mouse skin carcinogenesis.

 Biofactors. 21(1-4):33-39, 2004.



Study concluded in addition to other effects on gene expression, resveratrol also directly inhibited the activity of COX-2.

 J Biol Chem. 273(34):21875-21882, 1998.

Non-Steroidal Anti-Inflammatory Drugs

 Commonly prescribed for rheumatoid arthritis and osteoarthritis.

 Sept. 30, 2004: Vioxx recalled due to sudden heart attacks, strokes, and blood clots.

Resveratrol for Cardiovascular Support

 Healthy Platelet Activity 

Study reported a dose-dependent inhibition of platelet aggregation (blood clotting) by resveratrol.

 Clin Chim Acta. 235:207-219, 1995.



Rodent study demonstrated that resveratrol protects the heart and improves heart functions after diminished blood flow, affecting developed pressure and arterial flow.

 Journal of Cardiovascular Pharmacology. 35(2):263-268, 2000.



Shown to inhibit platelet aggregation both in vitro



and in vivo

International Journal of Molecular Medicine. 9:77-9, 2002.

Resveratrol Inhibits Platelet Aggregation

   Platelet aggregation is one of the first steps in the formation of a blood clot that can occlude a coronary or cerebral artery, resulting in myocardial infarction or stroke. Resveratrol has been found to inhibit platelet aggregation in vitro. Blood Cells Mol Dis. 2000;26(2):144-150. Clin Chim Acta. 1995;235(2):207-219.

Resveratrol

stimulates eNOS

 eNOS is an enzyme that catalyzes the formation of nitric oxide (NO) by vascular endothelial cells. NO is needed to maintain arterial relaxation (vasodilation), and impaired NO-dependent vasodilation is associated with increased risk of cardiovascular disease. Resveratrol has been

found to stimulate eNOS activity in cultured

endothelial cells.

 Circulation. 2002;106(13):1652-1658.

A study of mechanisms involved in vasodilation by Resveratrol

100 100 80 80 Endothelium Intact 60 60 Endothelium Removed 40 40 20 20 E-NOS Inhibitor (L-NNA) Control 0 0 1 1 5 5 10 50 50 100 100 Li etal. A Study of Mechanisms Involved in Vasodilation Induced by Resveratrol in Isolated Porcine Coronary Artery.

Physiol. Res

. 55:365-72, 2006.

Cardiovascular Support



Antioxidant protection of LDL particles



Resveratrol was shown to protect LDL by scavenging free radicals and chelating copper

 Fremont L et al. Antioxidant activity of resveratrol and alcohol-free wine polyphenols related to LDL oxidation and polyunsaturated fatty acids. 64(26):2511-21, 1999.  Belguendouz, L., et al. Resveratrol inhibits metal ion-dependent and independent peroxidation of porcine low-density lipoproteins. Biochem Pharmacol. 53(9):1347 1355, 1997.

Low-density Lipoprotein Particle (LDL)

Resveravine protects LDL from oxidation better than synthetic resveratrol, catechin and vitamin E Tlag (min) 500 400 300 200 100 0

control vitamin E catechin resveratrol resveravine

Caloric Restriction

 Caloric restriction is reducing caloric intake without malnutrition.

 Proven method for extending life span (30-50%) and postponing age-related functional decline and age-related disease.

Resveratrol Research in Extending Life Span



Restricting calorie intake extends the life spans of organisms ranging from yeast cells to lab animals.



Sirtuins enhance cell survival during times of stress, especially lack of food.



Sirtuins are regulators of aging in virtually all living organisms.



Resveratrol activates sirtuin activity without the extreme difficulty of starving oneself!

Sirtuins Help Cells Survive

 Sirtuins help rejuvenate old or dying cells by beefing up their DNA repair process and stimulating production of protective antioxidants.

 If a cell is at a point of deciding whether to live or die, Sirtuins push toward the survival mode by making the cell fix itself.

Studies Backing Resveratrol’s of Sirtuins Activation



Tests on Nothobranchius furzeri , a short-lived seasonal fish

 Resveratrol

extended the maximum life span by 59%

extended the median lifespan by 56% and  Study also showed resveratrol supplementation

age-related decline improved swimming performance and delayed motor and cognitive

 Trends in Pharmacological Sciences 26, 2005.  Current Biology, 2006. Obese mice supplemented with resveratrol

lived an average of 15% longer than obese mice not given resveratrol

 Nature, 2006.

Studies Backing Resveratrol’s of Sirtuins Activation



Saccharomyces cerevisiae (yeast), resveratrol mimics calorie restriction by stimulating Sir2, increasing DNA stability and extending lifespan by 70%.

 Howitz, K. T., et al. Small molecule activators of sirtuins extend Saccharomyces cerevisiae lifespan. Nature. 2003.



Drosophila melanogaster, resveratrol identified as being beneficial and shown to extend lifespan laboratory conditions



under normal

Bauer, J. H., et al. An accelerated assay for the identification of lifespan-extending interventions in Drosophila melanogaster. Proc Natl Acad Sci U S A. 101:12980-12985, 2004.

Resveratrol is a Superior Antioxidant



Oxidative stress is implicated in numerous disease processes



A lead resveratrol investigator, Dr. Milos Sovak commented, “ longevity but also general health.” There is no question that resveratrol is one of the best free-radical scavengers and that it has many effects whose ramifications might affect not only

Resveratrol as an Antioxidant



In vitro studies revealed that red wine extract is a potent antioxidant as evidenced by its ability to scavenge peroxyl radical in vitro.



Sato M et al. Myocardial protection with red wine extract. J Cardiovasc Pharmacol.35(2):263-8, 2000.



Olas et al demonstrated that resveratrol inhibits the production of several different reactive oxygen species (O 2 . , H 2 O 2 , singlet oxygen and organic radicals in pig blood platelets



Olas B et al. Resveratrol may reduce oxidative stress induced by platinum compounds in human plasma, blood platelets and lymphocytes. Anticancer Drugs. 16(6):659-65, 2005.

Protection against oxidative stress

120 100 80 60 40 20 0 0

Resveratrol Resveravine Polygonum cuspidatum extract

5

concentration (µg)

10

Influence of three related polyphenols on the oxidative stress of lymphocytes infected by the Epstein-Barr virus (EBV)

Resveratrol

Might Fight Obesity

 June 16 th . Google: “Martin Wabitsch endo08 webcast”  Human fat cells exposed to resveratrol or not exposed (control).

 Number of fat cells in control doubled. Number of fat cells exposed to resveratrol decreased by 50%.

Resveratrol

Might Fight Obesity

 In addition, exposure to resveratrol also reduced the secretion of substances called interleukin 6 and 8, which may be linked to the development of diabetes and clogged arteries, both thought to be obesity-related problems.

Safety of Resveratrol



Data in humans have revealed that resveratrol is safe to ingest.

 Aggarwal B et al. Role of resveratrol in prevention and therapy of cancer: preclinical and clinical studies. Anticancer Res. 24(5A):2783-840, 2004.



Human trials on Resveravine ®



Single oral doses of 1, 2.5 and 5 grams were given to 29 volunteers, and no serious adverse events were reported.

Possible Herb-Drug Interactions

 Resveratrol has been found to inhibit human platelet aggregation in vitro. Theoretically, high intakes of resveratrol (e.g., from supplements) could increase the risk of bleeding when taken with anticoagulant drugs, such as warfarin (Coumadin), and antiplatelet drugs, such as clopidogrel (Plavix), dipyridamole (Persantine), non-steroidal anti-inflamatory drugs (NSAIDs), aspirin and others.

As mentioned, Resveravine also provides



which is a dimer of Resveratrol :

-

Viniferin ,

Naturally synthesized by the plant, it is involved in the defense system against various fungi such as

Botrytis cinerea

.

It is naturally present in the grapes and wine Like Resveratrol,  -Viniferin has also very interesting biological properties: • •

anti-inflammatory antioxidant effects

BioVin

®

Advanced



BioVin Advanced is a French red wine extract, offering a full 5% of pure resveratrol



Made exclusively from French red wine grapes, naturally high in resveratrol



Contains red wine polyphenols and trans-resveratrol

BioVin

®

Advanced vs. Wine

Amount of Resveratrol in 200mg BioVin ® Advanced Amount required to be equivalent to 200 mg BioVin® Advanced 10 mg Red Wine 10 glasses (1500 ml) White Wine 40 glasses (6000 ml)

Benefits of BioVin

®

Advanced

Due to the synergistic blend of OPCs and resveratrol, BioVin Advanced offers numerous health advantages

     Cardiovascular health  Improves blood circulation   Strengthens blood vessels Decreases platelet aggregation Cellular health (as described earlier for Resveratrol)  Induces apoptosis, involvement in SIRT-1 activity, involvement in nuclear factor-kappa B and cyclooxygenase-2 activity Vision health Excellent antioxidant capabilities Skin health

VitaBlue™ Wild Blueberry Extract



Through a unique proprietary process, VitaBlue™ extract has concentrated the phytochemicals of the blueberry into an extract of unmatched purity, anthocyanin content and antioxidant potency.



Only the finest, ripest North American wild blueberries qualify to be used in this patented extract.



12.5% Anthocyanins, 40% Polyphenolics, 6500 ORAC units, including pterostilbene.

ORAC of Selected Foods 2007 U.S. Department of Agriculture Cranberry 9584 Blueberry 6552 Plum 6259 Blackberry 5347 Raspberry 4882 Strawberry 3577 Cherry 3365 Apple 3082 Pear 2941

Benefits of Blueberries

      

Blueberries have been shown to be beneficial in several applications

Provides 30 anthocyanins and pterostilbene Powerful antioxidant Inhibits cellular proliferation and induces apoptosis (programmed cell death in unhealthy cells) Promotes cardiovascular health Contains COX-2 inhibitors Promotes healthy vision

Blueberries and Cell Health



The following review summarizes current knowledge on the various molecular evidences of cancer chemoprevention by anthocyanins including 1) antioxidant activity 2) the molecular mechanisms involved in anti-carcinogenesis 3) the molecular mechanisms involved in the induction of apoptosis in tumor cells.

 Hou D et al. Potential mechanisms of cancer chemoprevention by anthocyanins. Curr Mol Med. 3(2):149-159, 2003.



In this study, blueberry exhibited potential anti-carcinogenic activity as evaluated by in vitro screening tests.

 Bomser, J., et al. In vitro anticancer activity of fruit extracts from Vaccinium species. Planta Medica. 62(3):212-216, 1996.

Hypoxia-Inducible Factor (HIF) – Why Antioxidants May Help to Prevent Cancer NORMAL CONDITIONS OXYGEN Hypoxia Nat Rev Cancer. 2003 Oct;3(10):721-32 OXYGEN

HIF Helps Cells Survive, But Free Radicals Must Be Present

Hypoxia HIF OXYGEN FREE RADICALS "HIF-1 helps an oxygen- starved cell convert sugar to energy without using oxygen and also initiates the construction of new blood vessels to bring in a fresh oxygen supply."

Cancer Cells Consume All The Oxygen Around Them

OXYGEN Hypoxia OXYGEN

But HIF Can’t Help the Cancer Cells Survive When Antioxidants Have Neutralized All The Free Radicals

.

Antioxidants Hypoxia OXYGEN FREE RADICALS HIF

And the Cancer Cells Die….

Hypoxia OXYGEN Antioxidants

Grape Extract Resveratrol Skin Eyes Cells Longevity, Obesity?, Diabetes?

Heart, Antioxidants Apoptosis, Inflammation Blueberries

Conclusions about Berry Extracts and Cell Health

   The active chemicals in selected berries have shown promise in maintaining cardiovascular, immune, and visual health, as well as lengthening life span and using fat stores more efficiently in experimental animal models.

The active chemicals are more than potent antioxidants – they interact with key enzymes and signaling pathways in cells.

Research is ongoing. More to come….