Research Update:Oligomeric Proanthocyanidins,OPCs:Introduction and Its Benefit Applications.
 seminal trace...Oligomeric Proanthocyanidins:OPCs,glycosidic anthocyanidins,monomeric flavan-3-ol (catechins) and flavan-3,4-diol units,antioxidants,free radical scavengers
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 Phytochemical info of Grape Seed Extract and Grape Skin Extract:
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Definition: from Grape Seed Extract and Grape Skin Extract are majorly composed of .
Chemical information disclosed as following table:
Research Update:Oligomeric Proanthocyanidins,OPCs:
 Quantitative analysis of polymeric procyanidins (Tannins) from grape (Vitis vinifera) seeds by reverse phase high-performance liquid chromatography.:
J Agric Food Chem. 2001; 49(1):26-31 (ISSN: 0021-8561).Peng Z; Hayasaka Y; Iland PG; Sefton M; H??j P; Waters EJ.Cooperative Research Centre for Viticulture, P.O. Box 154, Glen Osmond, South Australia 5064, Australia. zpeng@awri.adelaide.edu.au
A reverse phase C(18) HPLC method with potential for high automated throughput has been developed for the quantitative analysis of polymeric procyanidins (tannins) in grape seed extracts. Chromatography gave rise to 13 distinct UV-absorbing peaks with good baseline separation. The UV-absorbing peak eluting last is distinct and therefore easily quantified. Biochemical analyses including ultrafiltration, protein precipitation, and Sephadex LH20 chromatography combined with electrospray mass spectrometric analyses establish that this peak predominantly contains polymeric procyanidins. The polymers, which appear to be galloylated to various degrees and seem to fragment in a characteristic manner during electrospray mass spectrometry, are well separated from catechins and procyanidin oligomers of up to 4 units. The recovery of polymeric grape seed tannins with this HPLC method was 86%, which is similar to the 89% recovery achieved with commercial quebracho tannins. The concentration of tannins in seeds from ripe Vitis vinifera cv. Shiraz grapes ranged from 1360 to 2830 mg/kg of berries.
Antioxidant activity of a proanthocyanidin-rich extract from grape seed in whey protein isolate stabilized algae oil-in-water emulsions.:
J Agric Food Chem. 2004; 52(16):5272-6 (ISSN: 0021-8561).Hu M; McClements DJ; Decker EA.Department of Food Science, University of Massachusetts, Amherst, Massachusetts 01003, USA.
Algae oil-in-water emulsions stabilized with 0.2% whey protein isolate (WPI) at pH 3.0 and 7.0 were chosen to evaluate antioxidant activity of a proanthocyanidin-rich extract from grape seed. In this emulsion system, (+)-catechin and ascorbic acid (620 microM) were found to be prooxidative at pH 3.0 and ineffective at pH 7.0. Grape seed extract was not able to effectively inhibit both lipid hydroperoxides and propanal formation when added to the emulsion at 124 microM. However, increasing the concentration of the grape seed extract to 620 microM resulted in inhibition of both lipid hydroperoxide and propanal formation at pH 3.0 and 7.0. None of the antioxidants tested had any effect on the physical stability of the WPI-stabilized emulsion. The superior antioxidant activity of the grape seed extract is likely due to the presence of oligomeric procyanidins which are better antioxidants compared to their monomeric counterparts.
Grape seed extract suppresses lipid peroxidation and reduces hypoxic ischemic brain injury in neonatal rats.:
Brain Res Bull. 2005; 66(2):120-7 (ISSN: 0361-9230)Feng Y; Liu YM; Fratkins JD; LeBlanc MH.Department of Pediatrics, University of Mississippi Medical Center, 2500 North State Street, Jackson, MS 39216, USA.
Oxygen radicals play a crucial role in brain injury. Grape seed extract is a potent anti-oxidant. Does grape seed extract reduce brain injury in the rat pup? Seven-day-old rat pups had the right carotid arteries permanently ligated followed by 2.5 h of hypoxia (8% oxygen). Grape seed extract, 50 mg/kg, or vehicle was administered by i.p. 5 min prior to hypoxia and 4 h after reoxygenation and twice daily for 1 day. Brain damage was evaluated by weight deficit of the right hemisphere at 22 days following hypoxia and by histopathology. Grape seed extract reduced brain weight loss from 20.0+/-4.4% S.E.M. in vehicle pups (n=21) to 3.1+/-1.6% in treated pups (n=20, P<0.01). Grape seed extract improved the histopathologic brain score in cortex, hippocampus and thalamus (P<0.05 versus vehicle). Concentrations of brain 8-isoprostaglandin F2alpha and thiobarbituric acid reacting substances significantly increased due to hypoxic ischemia. Grape seed extract reduced this increase. Treatment with grape seed extract suppresses lipid peroxidation and reduces hypoxic ischemic brain injury in neonatal rat.
Rejuvenation of antioxidant system in central nervous system of aged rats by grape seed extract.:
Neurosci Lett. 2005; 383(3):295-300 (ISSN: 0304-3940).Balu M; Sangeetha P; Haripriya D; Panneerselvam C.Department of Medical Biochemistry, Dr. ALM Post Graduate Institute of Basic Medical Sciences, University of Madras, Taramani Campus, Chennai 600 113, India.
Oxidative stress is considered as a major risk factor that contributes to age-related increase in lipid peroxidation and declined antioxidants in the central nervous system during aging. Grape seed extract, one of the bioflavonoid, is widely used for its medicinal properties. In the present study, we evaluated the role of grape seed extract on lipid peroxidation and antioxidant status in discrete regions of the central nervous system of young and aged rats. Male albino rats of Wistar strain were divided into four groups: Group I-control young rats, Group II-young rats treated with grape seed extract (100 mg/kg body weight) for 30 days, Group III-aged control rats and Group IV-aged rats supplemented with grape seed extract (100 mg/kg body weight) for 30 days. Age-associated increase in lipid peroxidation was observed in the spinal cord, cerebral cortex, striatum and the hippocampus regions of aged rats (Group III). Activities of antioxidant enzymes like superoxide dismutase, catalase, glutathione peroxidase and levels of non-enzymic antioxidants like reduced glutathione, Vitamin C and Vitamin E were found to be significantly decreased in all the brain regions studied in aged rats when compared to young rats. However, normalized lipid peroxidation and antioxidant defenses were reported in the grape seed extract-supplemented aged rats. These findings demonstrated that grape seed extract enhanced the antioxidant status and decreased the incidence of free radical-induced lipid peroxidation in the central nervous system of aged rats.
The effect of grape-seed extract on 24 h energy intake in humans.:
Eur J Clin Nutr. 2004; 58(4):667-73 (ISSN: 0954-3007).Vogels N; Nijs IM; Westerterp-Plantenga MS.Maastricht University, Human Biology, Maastricht, The Netherlands. N.Vogels@HB.Unimaas.NL
OBJECTIVE: Since grape-seed extract has been shown to stimulate lipolysis in vitro and reduce food intake in rats, we assessed the efficacy of grape-seed extract with respect to energy intake (EI) and satiety. DESIGN: In a randomized, placebo-controlled, double-blind, cross-over study, 51 subjects (age 18-65 y, body mass index 22-30 kg/m2) ate an ad libitum lunch and dinner twice in the University Restaurant for 3 days. Standard breakfasts and snacks were provided. Supplements were taken 30-60 min prior to each meal.
RESULTS: In the total study population, no difference in 24 h EI was found between the grape-seed extract and placebo. However, in the subgroup of subjects (n=23) with an energy requirement > or =the median of 7.5 MJ/day, EI was reduced by 4% (DeltaEI 352.1 kJ/24 h, P=0.05) after grape-seed extract compared to placebo treatment. Meanwhile, there were no significant differences in macronutrient composition, attitude towards eating, satiety, mood or tolerance.
CONCLUSIONS: Grape seed reduced 24 h EI, with on average 4% in subjects who had an energy requirement > or =7.5 MJ/day, without further effects on satiety, mood or tolerance. These findings suggest that grape seed could be effective in reducing 24 h EI in normal to overweight dietary unrestrained subjects, and could, therefore, play a significant role in body-weight management.
Age-related oxidative protein damages in central nervous system of rats: modulatory role of grape seed extract.:
Int J Dev Neurosci. 2005; 23(6):501-7 (ISSN: 0736-5748).Balu M; Sangeetha P; Murali G; Panneerselvam C.Department of Medical Biochemistry, Dr. ALM Post Graduate Institute of Basic Medical Sciences, University of Madras, Taramani Campus, Chennai 600113, India.
Oxidative stress has been shown to play a major role in aging and in neurodegenerative disorders. Protein modification is one of the important consequences of oxidative stress. In the present study, we evaluated the role of grape seed extract on memory, reactive oxygen species production, protein carbonyls (PCO), and thiol status in discrete regions of central nervous system of young and aged rats. Male albino rats of Wistar strain were divided into four groups: Group I--control young rats, Group II--young rats treated with grape seed extract (100 mg/kg BW) for 30 days, Group III--aged control rats and Group IV-aged rats supplemented with grape seed extract (100 mg/kg BW) for 30 days. Memory loss was observed in the aged rats. Age associated increase in reactive oxygen species production and protein oxidation was observed in the spinal cord; cerebral cortex, striatum and the hippocampus regions of aged rats (Group III). The levels of total thiol, non-protein thiol, protein thiols were found to be significantly decreased in spinal cord and all the brain regions studied in aged rats when compared to young rats. Supplementation of aged rats with grape seed extract showed increased memory performance and declined reactive oxygen species production, decreased protein carbonyl levels and improved thiol levels. These findings demonstrated that grape seed extract enhanced the antioxidant status and decreased the incidence of free radical induced protein oxidation in aged rats thereby protecting the central nervous system from the reactive oxygen species.
The quality specification of grape seed extract:
Zhongguo Zhong Yao Za Zhi. 2005; 30(18):1406-8 (ISSN: 1001-5302).Shao YD; Gao WY; Su YF; Xiao PG.Tianjin Jianfeng Natural Product Co., Ltd., Tianjin 300402, China.
This paper reiviewed the current situation of quality control of grape seed extract in domestic and international market. Considering the fact that there is no national or industrial technical specifications established for the extract product, the authors suggested that two sets of quality specifications should be established for the grape seed extract. The two sets of specifications are: the high purity grape seed extract should contain polyphenol NLT 95%, monomer NLT 10%; and the grape seed extract with ordinary quality should have a procyanidolic value NLT 95, and monomer NLT 6%.
Modulatory role of grape seed extract on age-related oxidative DNA damage in central nervous system of rats.:
Brain Res Bull. 2006; 68(6):469-73 (ISSN: 0361-9230).Balu M; Sangeetha P; Murali G; Panneerselvam C.Department of Medical Biochemistry, Dr. ALM Post Graduate Institute of Basic Medical Sciences, University of Madras, Taramani Campus, Chennai 600 113, India.
Aging is the accumulation of diverse deleterious changes in the cells and tissues leading to increased risk of diseases. Oxidative stress is considered as a major risk factor and contributes to age related increase in DNA oxidation and DNA protein cross-links in central nervous system during aging. In the present study, we have evaluated the salubrious role of grape seed extract on accumulation of oxidative DNA damage products such as 8-OHdG and DNA protein cross-links in aged rats. Male albino rats of Wistar strain were divided into four groups: Group I, young control rats; Group II, young rats treated with grape seed extract (100mg/kgb.wt.) for 30 days; Group III, aged control rats; Group IV, aged rats supplemented with grape seed extract (100mg/kgb.wt.) for 30 days. Our results, thus, revealed that grape seed extract has inhibiting effect on the accumulation of age-related oxidative DNA damages in spinal cord and in various brain regions such as cerebral cortex, striatum and hippocampus.
Grape seed extract prevents gentamicin-induced nephrotoxicity and genotoxicity in bone marrow cells of mice.:
Basic Clin Pharmacol Toxicol. 2006; 99(3):230-6 (ISSN: 1742-7835).El-Ashmawy IM; El-Nahas AF; Salama OM.Department of Pharmacology, Faculty of Veterinary Medicine, Alexandria University, Alexandria, Egypt.
The protection conferred by grape seed extract against gentamicin-induced nephrotoxicity and bone marrow chromosomal aberrations have been evaluated in adult Swiss albino mice. The activity of reduced glutathione peroxidase (GSH peroxidase), the levels of glutathione (GSH) and lipid peroxidation as malondialdehyde (MDA) in the kidneys homogenates, serum urea and creatinine were measured, and in addition the changes in kidney histology and bone marrow chromosomes were investigated. Gentamicin (80 mg/kg b.wt. intraperitoneally for 2 weeks) induced kidney damage as indicated from a pronounced changes in kidney histology, a significant increase in serum urea and creatinine and MDA content in the kidney homogenate. While the activity of the antioxidant enzyme GSH peroxidase and the level of GSH were significantly decreased. Gentamicin induced genotoxicity indicated by increased the number of aberrant cells and different types of structural chromosomal aberrations (fragment, deletion and ring chromosome) and showed no effect on mitotic activity of the cell. Pretreatment with grape seed extract (7 days) and simultaneously (14 days) with gentamicin significantly protected the kidney tissue by ameliorating its antioxidant activity. Moreover, grape seed extract significantly protected bone marrow chromosomes from gentamicin induced genotoxicity by reducing the total number of aberrant cells, and different types of structural chromosomal aberrations. It could be concluded that grape seed extract acts as a potent antioxidant prevented kidney damage and genotoxicity of bone marrow cells.
Grape and grape seed extract capacities at protecting LDL against oxidation generated by Cu2+, AAPH or SIN-1 and at decreasing superoxide THP-1 cell production. A comparison to other extracts or compounds.:
Free Radic Res. 2003; 37(5):573-84 (ISSN: 1071-5762).Shafiee M; Carbonneau MA; Urban N; Descomps B; Leger CL.Laboratoire de Nutrition Humaine et Ath??rogenese, UPRES-EA 2033, Facult?? de M??decine, Universit?? de Montpellier I, F-34060 Montpellier, France.
A large body of evidence supports the key role of oxidized low-density lipoprotein in atherosclerosis. The aim of this study was to compare the capacity of natural polyphenols (PP) from Vitis vinifera and Olea europea at protecting LDL against oxidation brought about by Cu2+, oxygen-centered radical-generating AAPH, or peroxynitrite-generating SIN-1 in vitro systems, or at impairing superoxide production in promonocyte cells (THP-1) conveniently differentiated into adherent macrophages. PP were either from the whole grape (fraction A) containing mainly procyanidins, (epi)-catechin and anthocyanins, or from grape seed extracts (fractions B and C) consisting of tannins and procyanidin oligomers with a higher content in B than in C, or from a grape skin extract (fraction D) consisting mainly of anthocyanins, or from a hydrosoluble olive mill wastewater PP extract (fraction E) containing hydroxytyrosol and oleuropein. Chlorogenic acid (F) and catechin (G) were taken as archetypes of PP preventing oxidation partly as copper scavenger and as radical scavenger only, respectively. All grape fractions were efficient towards Cu2+ system (equally or more efficient than F), whereas they were rather poorly efficient towards AAPH and SIN-1 (less efficient than G but as efficient as F). Among the PP fractions, B was the most effective at protecting LDL in the SIN-1 system and at impairing THP-1 superoxide production. Taken together, these data suggest that the PP fraction from grape seed rich in procyanidins achieves the best compromise between the direct and indirect (i.e. cell-mediated) types of action in protecting LDL against oxidation, strengthening the need for improving the knowledge of its bioavailability in humans.
Neuroprotective effects of grape seed extract on neuronal injury by inhibiting DNA damage in the gerbil hippocampus after transient forebrain ischemia.:
Life Sci. 2004; 75(16):1989-2001 (ISSN: 0024-3205).Hwang IK; Yoo KY; Kim DS; Jeong YK; Kim JD; Shin HK; Lim SS; Yoo ID; Kang TC; Kim DW; Moon WK; Won MH.Department of Anatomy, College of Medicine, Hallym University, Chunchon, 200-702, South Korea.
Grape seed extract (GSE) possess cardioprotective abilities by functioning as in vivo antioxidants and by virtue of their ability to directly scavenge ROS including hydroxyl and peroxyl radicals. In the present study, we investigated the neuroprotective effects of grape seed extract (GSE) in the gerbil hippocampus after 5 min transient forebrain ischemia. Neuronal cell density in GSE-treated ischemic animals was significantly increased as compared with vehicle-treated ischemic animals 4 days after ischemic insult. In the GSE-treated groups, about 60% of pyramidal cells of the sham-operated group were stained with cresyl violet 4 days after ischemic insult. In this study, we found that GSE had neuroprotective effects on neuronal injury by inhibiting DNA damage in the CA1 region after ischemia. In vehicle-treated groups, 8-hydroxy-2'-deoxyguanosine (8-OHdG) immunoreactivity was significantly changed time-dependently, whereas the immunoreactivity in the GSE-treated group was similar to the sham-operated group. In addition, we confirmed that astrocytes and microglia did not show significant activation in the CA1 region 4 days after ischemia-reperfusion, because many CA1 pyramidal cells were not damaged. Therefore, these results suggest that GSE can protect ischemic neuronal damage by inhibiting DNA damage after transient forebrain ischemia.
Grape seed and skin extracts inhibit platelet function and release of reactive oxygen intermediates.:
J Cardiovasc Pharmacol. 2005; 46(4):445-51 (ISSN: 0160-2446).Vitseva O; Varghese S; Chakrabarti S; Folts JD; Freedman JE.Whitaker Cardiovascular Institute and Evans Department of Medicine, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
Red wine and purple grape juice contain polymeric flavonoids with antioxidant properties believed to be protective against cardiovascular events but the alcohol and sugar content of these beverages has curtailed their medicinal use. Acute cardiac events are also associated with enhanced inflammation and thrombosis. In this study, the extracts from grape skins or seeds were examined for their anti-inflammatory properties and effect on platelet release of reactive oxygen intermediates. Incubation of platelets with seed or skin extract led to a decrease in platelet aggregation from 68.8+/-19.8% to 45+/-3.6% for seeds and to 27+/-7.2% for skin, respectively (P<0.05). Platelet incubation with grape skin or seed extracts led to a marked decrease in superoxide release from 73+/-6.2 to 2+/-3.4 for grape seeds and to 0.33+/-0.57 for grape skin (chemilum. units; P<0.05) as well as a significant increase in radical-scavenging activity, decrease in reactive oxygen species release by confocal microscopy, and enhanced platelet NO was measured using an NO-sensitive microelectrode. These effects were dose dependent for both grape extracts. Coincubation with seeds and skins led to additive inhibition of platelet aggregation, enhanced NO release, and prevented superoxide production. Incubation with seed or skin extracts led to an immediate attenuation of release of the inflammatory mediator, soluble CD40 ligand. Thus, the extracts from purple grape skins and seeds inhibit platelet function and platelet-dependent inflammatory responses at pharmacologically relevant concentrations. These findings suggest potentially beneficial platelet-dependent antithrombotic and anti-inflammatory properties of purple grape-derived flavonoids.
Polyphenolics in grape seeds-biochemistry and functionality.:
J Med Food. 2003; 6(4):291-9 (ISSN: 1096-620X)Shi J; Yu J; Pohorly JE; Kakuda Y.Food Research Center, Agriculture and Agri-Food Canada, Guelph, Canada. shij@agr.gc.ca
Grape seeds are waste products of the winery and grape juice industry. These seeds contain lipid, protein, carbohydrates, and 5-8% polyphenols depending on the variety. Polyphenols in grape seeds are mainly flavonoids, including gallic acid, the monomeric flavan-3-ols catechin, epicatechin, gallocatechin, epigallocatechin, and epicatechin 3-O-gallate, and procyanidin dimers, trimers, and more highly polymerized procyanidins. Grape seed extract is known as a powerful antioxidant that protects the body from premature aging, disease, and decay. Grape seeds contains mainly phenols such as proanthocyanidins (oligomeric proanthocyanidins). Scientific studies have shown that the antioxidant power of proanthocyanidins is 20 times greater than vitamin E and 50 times greater than vitamin C. Extensive research suggests that grape seed extract is beneficial in many areas of health because of its antioxidant effect to bond with collagen, promoting youthful skin, cell health, elasticity, and flexibility. Other studies have shown that proanthocyanidins help to protect the body from sun damage, to improve vision, to improve flexibility in joints, arteries, and body tissues such as the heart, and to improve blood circulation by strengthening capillaries, arteries, and veins. The most abundant phenolic compounds isolated from grape seed are catechins, epicatechin, procyanidin, and some dimers and trimers.
Effects of exposure to grape-seed polyphenols and vitamin C on lipid eroxidation in vivo.:
Asia Pac J Clin Nutr. 2004; 13(Suppl):S76 (ISSN: 0964-7058).Ward NC; Hodgson JM; Puddey IB; Croft KD.School of Medicine & Pharmacology (RPH), University of Western Australia, Perth WA, Australia.
Introduction - Oxidative stress has been implicated in a number of disease processes. There is evidence suggesting that vitamin C, a major water-soluble antioxidant, may reduce oxidative stress. The effects of dietary polyphenols, water-soluble compounds with potent antioxidant activity in vitro, on oxidative stress are unclear. Objectives - The objectives of this study were to investigate the effect of supplementation with grape-seed polyphenols on oxidative stress, and to compare any effects to those of vitamin C. Design- Following a 3-week washout, participants were randomised to receive (i) 500mg/day vitamin C + matched placebo (n = 19), (ii) 1000mg /day polyphenols + matched placebo (n= 16), (iii) 500mg/day vitamin C + 1000mg/day polyphenols (n = 16), or (iv) matched placebos (n = 18). Plasma and urinary F(2)-isoprostanes and oxidised low-density lipoproteins were analysed as markers of oxidative damage. Outcomes - Supplementation with grape-seed polyphenols resulted in a significant increase in urinary excretion of specific phenolic acids (3-hydroxyphenylproprionic acid), but did not alter F(2)-isoprostane concentrations or oxidised low-density lipoproteins. The phenolic acid metabolites, markers of exposure to grape-seed polyphenols, were not related to changes in markers of oxidative stress. Plasma vitamin C levels increased significantly following supplementation. Plasma F(2)-isoprostane concentrations fell following supplementation with vitamin C (p=0.056). There was no change in urinary F(2)-isoprostane concentrations or oxidised low-density lipoproteins. There was no relationship between increases in plasma vitamin C and changes in markers of oxidative stress. Conclusions - These results support the suggestion that supplementation with vitamin C may reduce in vivo lipid peroxidation. However, supplementation with grape-seed polyphenols and exposure to phenolic acid metabolites had no effect on in vivo lipid peroxidation.
Supplementation with grape seed polyphenols results in increased urinary excretion of 3-hydroxyphenylpropionic Acid, an important metabolite of proanthocyanidins in humans.:
J Agric Food Chem. 2004; 52(17):5545-9 (ISSN: 0021-8561).Ward NC; Croft KD; Puddey IB; Hodgson JM.School of Medicine and Pharmacology, University of Western Australia, and Western Australian Institute for Medical Research, Perth, Australia. nward@cyllene.uwa.edu.au
Grape seed extract provides a concentrated source of polyphenols, most of which are proanthocyanidins. Polymeric proanthocyanidins are poorly absorbed in the small intestine of humans, and exposure may result from metabolism to phenolic acids by colonic bacteria. Any biological effects of proanthocyanidins may be due to the phenolic acid metabolites. Several phenolic acids have been identified as proanthocyanidin metabolites, but these may be derived from a range of other dietary sources. The aim of this study was to determine if 24-h urinary excretion of specific phenolic acids increased significantly and consistently following regular supplementation with grape seed extract. In a randomized, double-blind placebo-controlled trial, 69 volunteers received grape seed extract (1000 mg/day total polyphenols) or placebo for 6 weeks. Supplementation with grape seed polyphenols resulted in a consistent increase in the excretion of 3-hydroxyphenylpropionic acid (3-HPP, P < 0.001) and 4-O-methylgallic acid (P < 0.001) and a less consistent increase in the excretion of 3-hydroxyphenylacetic acid (P = 0.002). The observed increase in 3-HPP is in line with the suggestion that this compound is a major phenolic acid breakdown product of proanthocyanidin metabolism in vivo.
Total phenolics and antioxidant activities of fenugreek, green tea, black tea, grape seed, ginger, rosemary, gotu kola, and ginkgo extracts, vitamin E, and tert-butylhydroquinone.:
J Agric Food Chem. 2004; 52(16):5183-6 (ISSN: 0021-8561).Rababah TM; Hettiarachchy NS; Horax R.Department of Food Science, University of Arkansas, 2650 North Young Avenue, Fayetteville, Arkansas 72704, USA.
The total phenolics and antioxidant activities of fenugreek, green tea, black tea, grape seed, ginger, rosemary, gotu kola, and ginkgo extracts, vitamin E, and tert-butylhydroquinone, were determined. Grape seed and green tea were analyzed for their phenolic constituents using high-performance liquid chromatography. The total phenolics of the plant extracts, determined by the Folin-Ciocalteu method, ranged from 24.8 to 92.5 mg of chlorogenic acid equivalent/g dry material. The antioxidant activities of methanolic extracts determined by conjugated diene measurement of methyl linoleate were 3.4-86.3%. The antioxidant activity of the extracts using chicken fat by an oxidative stability instrument (4.6-10.2 h of induction time) followed a similar trend in antioxidant activity as determined by the Folin-Ciocalteu method. Seven phenolics in grape seed and green tea extracts were identified that ranged from 15.38 to 1158.49 and 18.3 to 1087.02 mg/100 g of extract, respectively. Plant extracts such as green tea and grape seed extracts can be used to retard lipid oxidation in a variety of food products.
Grape seed extract prevents gentamicin-induced nephrotoxicity and genotoxicity in bone marrow cells of mice.:
Basic Clin Pharmacol Toxicol. 2006; 99(3):230-6 (ISSN: 1742-7835).El-Ashmawy IM; El-Nahas AF; Salama OM.Department of Pharmacology, Faculty of Veterinary Medicine, Alexandria University, Alexandria, Egypt.
The protection conferred by grape seed extract against gentamicin-induced nephrotoxicity and bone marrow chromosomal aberrations have been evaluated in adult Swiss albino mice. The activity of reduced glutathione peroxidase (GSH peroxidase), the levels of glutathione (GSH) and lipid peroxidation as malondialdehyde (MDA) in the kidneys homogenates, serum urea and creatinine were measured, and in addition the changes in kidney histology and bone marrow chromosomes were investigated. Gentamicin (80 mg/kg b.wt. intraperitoneally for 2 weeks) induced kidney damage as indicated from a pronounced changes in kidney histology, a significant increase in serum urea and creatinine and MDA content in the kidney homogenate. While the activity of the antioxidant enzyme GSH peroxidase and the level of GSH were significantly decreased. Gentamicin induced genotoxicity indicated by increased the number of aberrant cells and different types of structural chromosomal aberrations (fragment, deletion and ring chromosome) and showed no effect on mitotic activity of the cell. Pretreatment with grape seed extract (7 days) and simultaneously (14 days) with gentamicin significantly protected the kidney tissue by ameliorating its antioxidant activity. Moreover, grape seed extract significantly protected bone marrow chromosomes from gentamicin induced genotoxicity by reducing the total number of aberrant cells, and different types of structural chromosomal aberrations. It could be concluded that grape seed extract acts as a potent antioxidant prevented kidney damage and genotoxicity of bone marrow cells.
Ingestion of IH636 grape seed proanthocyanidin extract to prevent selenite-induced oxidative stress in experimental cataract.:
J Cataract Refract Surg. 2006; 32(6):1041-5 (ISSN: 0886-3350)Durukan AH; Evereklioglu C; Hurmeric V; Kerimoglu H; Erdurman C; Bayraktar MZ; Mumcuoglu T.Department of Ophthalmology, G??lhane Military Medical Academy, Ankara, Turkey. ahakand@yahoo.com
PURPOSE: To investigate whether dietary supplementation with IH636 grape seed proanthocyanidin extract (GSPE) prevents selenite-induced cataract. SETTING: Department of Ophthalmology, Gulhane Military Medical Academy, Ankara, Turkey. METHODS: Thirty Spraque-Dawley rat litters were put randomly into 3 groups. In group 1 (n = 10), sodium selenite (30 nmol/g body weight) was injected subcutaneously on postpartum day 10. In group 2 (n = 10), sodium selenite (30 nmol/g body weight) was injected on postpartum day 10 and oral GSPE (100 mg/kg body weight) was given for 1 week after sodium selenite injection. Only subcutaneous saline was injected in group 3 (control, n = 10). The development of cataract was assessed for 3 weeks, and its density was graded and photographed with a slitlamp. Removed rat lenses were analyzed for glutathione (GSH) and malondialdehyde (MDA). RESULTS: All of the rats in group 1 had cataract between stage 6 and stage 3. In group 2, only 5 of 10 eyes had cataract between stage 3 and stage 2 and no cataract occurred in the remaining 5 rats. The difference between mean cataract stages in group 1 and group 2 was significant (P<.05). The mean GSH level in group 1 was significantly lower than in group 2 and controls (P<.05). The mean MDA level in group 1 was significantly higher than in group 2 and controls (P<.05). CONCLUSIONS: IH636 grape seed proanthocyanidin extract effectively suppressed cataract formation in rats. Routine consumption of grape seed proanthocyanidin extract in the form of food or dietary supplement may offer a prophylactic measure against onset and progression of cataract.
Adaptogenic and nootropic activities of aqueous extract of Vitis vinifera (grape seed): an experimental study in rat model.:
BMC Complement Altern Med. 2005; 5:1 (ISSN: 1472-6882)Sreemantula S; Nammi S; Kolanukonda R; Koppula S; Boini KM.Pharmacology Division, Department of Pharmaceutical Sciences Andhra University, Visakhapatnam-530 003, Andhra Pradesh, India. satyanarayana_sreemantula@rediffmail.com
BACKGROUND: The aerial parts of Vitis vinifera (common grape or European grape) have been widely used in Ayurveda to treat a variety of common and stress related disorders. In the present investigation, the seed extract of V. vinifera was evaluated for antistress activity in normal and stress induced rats. Furthermore, the extract was studied for nootropic activity in rats and in-vitro antioxidant potential to correlate its antistress activity.
METHODS: For the evaluation of antistress activity, groups of rats (n = 6) were subjected to forced swim stress one hour after daily treatment of V. vinifera extract. Urinary vanillylmandelic acid (VMA) and ascorbic acid were selected as non-invasive biomarkers to assess the antistress activity. The 24 h urinary excretion of vanillylmandelic acid (VMA) and ascorbic acid were determined by spectrophotometric methods in all groups under normal and stressed conditions. The nootropic activity of the extract as determined from acquisition, retention and retrieval in rats was studied by conditioned avoidance response using Cook's pole climbing apparatus. The in vitro antioxidant activity was determined based on the ability of V. vinifera to scavenge hydroxyl radicals.
RESULTS: Daily administration of V. vinifera at doses of 100, 200 and 300 mg/kg body weight one hour prior to induction of stress inhibited the stress induced urinary biochemical changes in a dose dependent manner. However, no change in the urinary excretion of VMA and ascorbic acid was observed in normal animals at all the doses studied. The cognition, as determined by the acquisition, retention and recovery in rats was observed to be dose dependent. The extract also produced significant inhibition of hydroxyl radicals in comparison to ascorbic acid in a dose dependent manner.
CONCLUSION: The present study provides scientific support for the antistress (adaptogenic), antioxidant and nootropic activities of V. vinifera seed extract and substantiate the traditional claims for the usage of grape fruits and seeds in stress induced disorders.
Quality assessment of commercial dietary antioxidant products from Vitis vinifera L. grape seeds.:
Nutr Cancer. 2005; 53(2):244-54 (ISSN: 0163-5581)Monagas M; Hern??ndez-Ledesma B; Garrido I; Mart??n-Alvarez PJ; G??mez-Cordov??s C; Bartolom?? B.Instituto de Fermentaciones Industriales (CSIC), Madrid, Spain.
Phenolic preparations from Vitis vinifera L. grape seeds are products commonly used in the formulation of dietary antioxidant supplements. In this article, we used a methodology (the oxygen radical absorbance capacity, ORAC) to evaluate the in vitro antioxidant capacity of commercial dietary grape seed products and studied the relationship of the antioxidant capacity with the phenolic composition of these products. The ORAC value of the different brands of commercial products studied (n = 16) varied from 2.71 to 26.4 micromol Trolox equivalents/mg (approximately equal to 10-fold difference). For four of these products, the batch-to-batch ORAC variation, expressed as a coefficient of variation of the mean, was 10.5% (n = 6), 13.1% (n = 3), 19.4% (n = 4), and 7.8% (n = 4). Analysis of monomeric and oligomeric flavan-3-ols by liquid chromatography-diode array detection (LC-DAD)/electrospray-mass spectrometry and procyanidins by thiolysis-LC-DAD also revealed large differences among the commercial grape seed products. Moreover, the ORAC value could be fitted to a regression model using variables from contents of individual phenolic compounds and procyanidins. The product-to-product and batch-to-batch variation in ORAC values and flavan-3-ol composition found among the commercial products studied demonstrated that they are poorly standardized, resulting in inconsistent composition and biological activity.
Antigenotoxic effect of grape seed procyanidin extract in Fao cells submitted to oxidative stress.:
J Agric Food Chem. 2004; 52(5):1083-7 (ISSN: 0021-8561).Ll??piz N; Puiggr??s F; C??spedes E; Arola L; Ard??vol A; Blad?? C; Salvad?? MJ.Departament de Bioqu??mica i Biotecnolog??a, Unitat d'Enologia del Centre de Refer??ncia en Tecnologia dels Aliments de la Generalitat de Catalunya, Universitat Rovira i Virgili, Imperial T??rraco 1, 43005 Tarragona, Spain.
The protective effects of grape seed procyanidin extract on the repair of H(2)O(2)-induced DNA lesions were tested using Fao cells. Cells were exposed to 600 microM H(2)O(2) for 3 or 21 h. A procyanidin extract from grape seed (PE) was incubated or preincubated (1 h) during the exposure to H(2)O(2). The ability of procyanidins to protect against the genotoxicity of H(2)O(2) was compared with those of the monomeric flavanols (+)-catechin and (-)-epicatechin and the flavonol quercetin. After treatment, DNA damage was monitored using alkaline single-cell gel electrophoresis (the comet assay) (Aherne, S. A.; O'Brien, N. M. Nutr. Cancer 1999, 34, 160-166). At the end of the experiment, PE significantly decreased the damage caused by H(2)O(2). The results also showed that quercetin was the most effective of the flavonoids tested, which is consistent with its powerful antioxidant character. The results indicate that procyanidins are more effective than the corresponding individual monomers, catechin and epicatechin, at preventing DNA lesions in hepatocytes and that this protection is higher after preincubation than after co-incubation.
Defatted milled grape seed protects adriamycin-treated hepatocytes against oxidative damage.:
Eur J Nutr. 2006; 45(5):251-8 (ISSN: 1436-6207).Valls-Belles V; Torres MC; Mu??iz P; Beltran S; Martinez-Alvarez JR; Codo??er-Franch P.Depto. de Pediatr??a, Ginecolog??a y Obstetricia Facultad de Medicina, Universidad de Valencia, Valencia, Spain.
Defatted milled grape seed (DMGS) is a wine by-product obtained from the oil extraction of the grape seed that contains different types of phenolic compounds. The present study was designed to evaluate the possible protective effect of DMGS on toxicity induced by adriamycin (ADR) in isolated rat hepatocytes. The study was carried out by examining the results of lactate dehydrogenase (LDH) release to estimate cytotoxicity; the thiobarbituric acid reactant substances (TBARS) and carbonyl group levels were measured as biomarkers of oxidative stress and ATP and GSH levels as estimation of intracellular effect. The results showed that DMGS extract protects the cellular membrane from oxidative damage and consequently prevents protein and lipid oxidation. The levels of ATP and GSH changes for the ADR toxicity were restored to control value in the presence of DMGS extract. The experimental results suggest that this wine by-product may be used to decrease oxidative stress.
Isolation and structure identification of grape seed polyphenols and its effects on oxidative damage to cellular DNA:
Yao Xue Xue Bao. 2004; 39(11):869-75 (ISSN: 0513-4870).Fan PH; Lou HX.College of Pharmacy, Shandong University, Jinan 250012, China.
AIM: To isolate polyphenols from grape seeds and to evaluate their antioxidant effects. METHODS: Pure compounds were isolated by using Diaion HP20, Toyopearl HW40 chromatography repeatedly, as well as semi-preparative RP-HPLC, from ethyl acetate extract of grape seeds. IR, MS, NMR, CD, X-Ray crystal diffraction spectral analysis were used to identify the structures. The antioxidant effects of different type of structures were screened by reducing power and DPPH (alpha,alpha-diphenyl-beta-picrylhydrazyl) free radical scavenging tests. Then, SCGE (single cell gel-electrophoresis) technique was used to investigate the effects of these potent antioxidant phytochemicals on cellular DNA oxidative damage with mice spleen cells, damage was induced by H2O2. RESULTS: Eleven compounds were obtained including 3 novel structures, viniferones A, B and C. Proanthocyanidin B4, catecin, epicatechin and gallic acid showed strong antioxidant power, and at lower concentration (10 micromol x L(-1), 25 micromol x L(-1)), they can prevent cellular DNA damage, while 150 micromol x L(-1) catechin induced damage by itself. CONCLUSION: Viniferones A, B and C were reported for the first time. That polyphenols investigated were shown to be good cellular DNA oxidative damage-preventing phytochemicals at lower concentration, could be used to explain the nutrient effect of grape seed polyphenols at certain degree. At the same time, higher concentration of polyphenols can induce oxidative damage, suggesting that dose is one factor to determine the nutrient effects.
Protective effects of grape seed proanthocyanidins against oxidative stress induced by lipopolysaccharides of periodontopathogens.:
J Periodontol. 2006; 77(8):1371-9 (ISSN: 0022-3492).Houde V; Grenier D; Chandad F.Research Group in Oral Ecology, Faculty of Dentistry, Laval University, Qu??bec City, QC, Canada.
BACKGROUND: During phagocytosis or stimulation with bacterial components, macrophages activate various cell processes, including the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS), which are critical for successful defense against invading organisms. Increased levels of ROS/RNS create oxidative stress that results in tissue and bone destruction. Grape seed proanthocyanidins have been reported to possess a wide range of biologic properties against oxidative stress. In the present study, we investigated the effects of a grape seed proanthocyanidin extract (GSE) and commercial polyphenols on the production of ROS and RNS and on the protein expression of inducible nitric oxide synthase (iNOS) by murine macrophages stimulated with lipopolysaccharides (LPS) of periodontopathogens.
METHODS: Macrophages (RAW 264.7) were treated with non-toxic concentrations of either GSE or commercial polyphenols (gallic acid [GA] and [-]-epigallocatechin-3-gallate [EGCG]) and stimulated with LPS of Actinobacillus actinomycetemcomitans or Fusobacterium nucleatum, and iNOS expression was evaluated by immunoblotting. Nitric oxide (NO) production was quantified using the colorimetric Griess assay, whereas ROS production was measured with the fluorescent 123-dihydrorhodamine dye.
RESULTS: GSE strongly decreased NO and ROS production and iNOS expression by LPS-stimulated macrophages. GA also revealed a strong inhibitory effect on NO production without affecting iNOS expression but slightly increasing ROS production. EGCG showed an inhibitory effect on NO and ROS production and on iNOS expression by macrophages.
CONCLUSION: Our findings demonstrate that proanthocyanidins have potent antioxidant properties and should be considered a potential agent in the prevention of periodontal diseases.
Chemoprevention by grape seed extract and genistein in carcinogen-induced mammary cancer in rats is diet dependent.:
J Nutr. 2004; 134(12 Suppl):3445S-3452S (ISSN: 0022-3166)Kim H; Hall P; Smith M; Kirk M; Prasain JK; Barnes S; Grubbs C.Departments of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, AL 35294, USA. helenkim@uab.edu
Many popular dietary supplements are enriched in polyphenols such as the soy isoflavones, tea catechins, and resveratrol (from grape skins), each of which has been shown to have chemopreventive activity in cellular models of cancer. The proanthocyanidins, which are oligomers of the catechins, are enriched in grape seeds and form the basis of the dietary supplement grape seed extract (GSE). Evidence suggests that the proanthocyanidins may be metabolized to the monomeric catechins. This study was carried out to determine whether GSE added to rodent diets protected against carcinogen-induced mammary tumorigenesis in rats and whether this was affected by the composition of the whole diet. Female rats were begun on 5%, 1.25%, or 0% (control) GSE-supplemented diets at age 35 d. At age 50 d they were administered 7,12-dimethylbenz[a]anthracene (DMBA) in sesame oil at 80 mg/kg body weight. They were weighed and monitored weekly for tumor development until 120 d after DMBA administration. Administration of GSE in AIN-76A diet did not show any protective activity of GSE against DMBA-induced breast cancer. However, administration of GSE in a laboratory dry food diet (Teklad 4% rodent diet) resulted in a 50% reduction in tumor multiplicity. In similar experiments, genistein administered in AIN-76A diet also failed to show chemopreventive activity against the carcinogen N-methyl-N-nitrosourea; however, when administered at the same dose in the Teklad 4% rodent diet, genistein exhibited significant chemopreventive activity (44-61%). These results demonstrate that GSE is chemopreventive in an animal model of breast cancer; moreover, the diet dependency of the chemopreventive activity for both GSE and genistein suggests that whether or not a compound is chemopreventive may depend on the diet in which the agent is administered.
The combination of vitamin C and grape-seed polyphenols increases blood pressure: a randomized, double-blind, placebo-controlled trial.:
J Hypertens. 2005; 23(2):427-34 (ISSN: 0263-6352)Ward NC; Hodgson JM; Croft KD; Burke V; Beilin LJ; Puddey IB.School of Medicine and Pharmacology, University of Western Australia and West Australian Institute for Medical Research (WAIMR), Perth, Australia. nward@cyllene.uwa.edu.au
BACKGROUND: There is growing evidence that oxidative stress contributes to the pathogenesis of hypertension and endothelial dysfunction. Thus, dietary antioxidants may beneficially influence blood pressure (BP) and endothelial function by reducing oxidative stress.
OBJECTIVE: To determine if vitamin C and polyphenols, alone or in combination, can lower BP, improve endothelial function and reduce oxidative stress in hypertensive individuals.
DESIGN: A total of 69 treated hypertensive individuals with a mean 24-h ambulatory systolic blood pressure > or = 125 mmHg participated in a randomized, double-blind, placebo-controlled, factorial trial. Following a 3-week washout, participants received 500 mg/day vitamin C, 1000 mg/day grape-seed polyphenols, both vitamin C and polyphenols, or neither for 6 weeks. At baseline and post-intervention, 24-h ambulatory BP, ultrasound-assessed endothelium-dependent and -independent vasodilation of the brachial artery, and markers of oxidative damage, (plasma and urinary F2-isoprostanes, oxidized low-density lipoproteins and plasma tocopherols), were measured.
RESULTS: A significant interaction between grape-seed and vitamin C treatments for effects on BP was observed. Vitamin C alone reduced systolic BP versus placebo (-1.8 +/- 0.8 mmHg, P = 0.03), while polyphenols did not (-1.3 +/- 0.8 mmHg, P = 0.12). However, treatment with the combination of vitamin C and polyphenols increased systolic BP (4.8 +/- 0.9 mmHg versus placebo; 6.6 +/- 0.8 mmHg versus vitamin C; 6.1 +/- 0.9 mmHg versus polyphenols mmHg, each P < 0.0001) and diastolic BP (2.7 +/- 0.6 mmHg, P < 0.0001 versus placebo; 1.5 +/- 0.6 mmHg, P = 0.016 versus vitamin C; 3.2 +/- 0.7 mmHg, P < 0.0001 versus polyphenols). Endothelium-dependent and -independent vasodilation, and markers of oxidative damage were not significantly altered.
CONCLUSION: Although the mechanism remains to be elucidated, these results suggest caution for hypertensive subjects taking supplements containing combinations of vitamin C and polyphenols.
Composition and cellular localization of tannins in grape seeds during maturation.:
J Agric Food Chem. 2003; 51(27):8051-4 (ISSN: 0021-8561)Geny L; Saucier C; Bracco S; Daviaud F; Glories Y.UMR OEnologie-Amp??lologie 1219, Facult?? d'oenologie-INRA, Universit?? Victor S??galen Bordeaux 2, 33405 Talence, France. laurence.geny@oenologie.u-bordeaux2.fr
Cell walls were isolated from seeds of grape berries (Vitis vinifera L.), and proanthocyanidin composition was determined over the course of ripening for different levels of vine water status. During the ripening period the tannins from the cell walls were always more polymerized than those from the inner part of the cell. At maturity this difference becomes more significant compared to v??raison, due to a significant increase in the mean degree of polymerization of the cell wall tannins. The tannin composition was typical of grape seed tannins and was quite similar in the two cell fractions studied, but the epicatechin gallate proportion was significantly higher in the cell wall fraction. There were no significant effects of water deficit on composition and polymerization of seed tannins.
Effect of a standardized grape seed extract on low-density lipoprotein susceptibility to oxidation in heavy smokers.:
Metabolism. 2003; 52(10):1250-7 (ISSN: 0026-0495)Vigna GB; Costantini F; Aldini G; Carini M; Catapano A; Schena F; Tangerini A; Zanca R; Bombardelli E; Morazzoni P; Mezzetti A; Fellin R; Maffei Facino R.Department of Clinical and Experimental Medicine, Section of Internal Medicine II, University of Ferrara, Ferrara, Italy.
The aim of our study was to evaluate the effect of a standardized formulation of a polyphenolic extract of grapes (Leucoselect-Phytosome [LP]) on low-density lipoprotein (LDL) susceptibility to oxidation in a group of heavy smokers. A randomized, double-blind, crossover study was undertaken in 24 healthy male heavy smokers, aged > or = 50 years. Enrolled subjects were given 2 capsules twice daily for 4 weeks (phase 1). Each capsule contained 75 mg of a grape procyanidin extracts and soy-phosphatidlcholine or placebo consisiting of 75 mg lactose and soy-phosphatidlcholine. A wash out period of 3 weeks was then followed by 4 weeks of the opposite treatment (phase 2). Blood samples were taken at baseline and at the end of each phase and assayed for plasma lipids and LDL susceptibility to oxidation. Compliance was good, and no adverse effects were recorded. Subjects did not show significant modification of total cholesterol (TC), triglycerides (TG), high-density lipoprotein-cholesterol (HDL-C) and LDL-C during LP treatment. Among oxidative indices, thiobarbituric acid reactive substances (TBARS) concentration was significantly reduced in subjects taking LP (-14.7% +/- 21.1% v +5.0% +/- 18.1%, P <.01), and the lag phase prolonged (+15.4% +/- 24.4% v -0.1% +/- 16.0%, P <.05) compared with placebo and basal values. The antioxidant potential of grape seed extract polyphenols may prove effective in a model of oxidative stress (smoking); however more investigational data are needed before use in wider clinical settings.
Inhibitory effects of grape seed extract on lipases.:
Nutrition. 2003; 19(10):876-9 (ISSN: 0899-9007).Moreno DA; Ilic N; Poulev A; Brasaemle DL; Fried SK; Raskin I.Biotech Center, Cook College, Rutgers--The State University of New Jersey, New Brunswick, New Jersey 08901-8520, USA. moreno@aesop.rutgers.edu
OBJECTIVE: The aim of the present study was to assess the effects of grape seed extract (GSE) on the fat-metabolizing enzymes pancreatic lipase, lipoprotein lipase, and hormone-sensitive lipase in vitro and evaluate its potential application as a treatment for obesity.
METHODS: Crushed grape seeds were extracted in ethanol, and the extract was assayed for the measurement of inhibitory effects on pancreatic lipase and lipoprotein lipase activities and on lipolysis of 3T3-L1 adipocytes.
RESULTS: The GSE rich in bioactive phytochemicals showed inhibitory activity on the fat-metabolizing enzymes pancreatic lipase and lipoprotein lipase, thus suggesting that GSE might be useful as a treatment to limit dietary fat absorption and the accumulation of fat in adipose tissue. The observed reduction in intracellular lipolytic activity of cultured 3T3-L1 adipocytes may reduce the levels of circulating free fatty acids that have been linked to insulin resistance in obese patients.
CONCLUSION: The GSE rich in compounds that inhibit lipases may provide a safe, natural, and cost-effective weight control treatment.
Protective effects of oligomers of grape seed polyphenols against beta-amyloid-induced oxidative cell death.:
Ann N Y Acad Sci. 2004; 1030:317-29 (ISSN: 0077-8923).Li MH; Jang JH; Sun B; Surh YJ.Research Institute of Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Shinlim-dong, Kwanak-ku, Seoul 151-742, South Korea.
beta-Amyloid (Abeta) is considered to be responsible for the formation of senile plaques that accumulate in the brains of patients with Alzheimer's disease (AD). There is compelling evidence supporting the notion that Abeta-induced cytotoxicity is mediated through the generation of reactive oxygen species (ROS). Recently, considerable attention has been focused on a wide array of non-vitamin antioxidants present in edible plants that are able to scavenge ROS, thereby protecting against oxidative damage. In this study, we have investigated the possible protective effects of formulated polyphenol oligomers (Oligonol) derived from grape seed extracts on Abeta-induced oxidative cell death. Rat pheochromocytoma (PC12) cells treated with Abeta exhibited increased accumulation of intracellular ROS and underwent apoptosis, as determined by positive in situ terminal end labeling, decreased mitochondrial membrane potential, and the cleavage of poly(ADP-ribose)polymerase. Oligonal attenuated Abeta-induced cytotoxicity, apoptotic features, intracellular ROS accumulation, and lipid peroxidation and increased the cellular glutathione pool. Moreover, Abeta transiently induced the activation of nuclear factor kappaB in PC12 cells, which was suppressed by pretreatment with Oligonol.
Effect of grape seed extract on puromycin-aminonucleoside-induced nephrosis in rats.:
Pediatr Nephrol. 2003; 18(9):872-7 (ISSN: 0931-041X)Mattoo TK; Kovacevic L.Division of Pediatric Nephrology, Children's Hospital of Michigan, 3901 Beaubien Boulevard, Detroit, MI 48201, USA. tmattoo@med.wayne.edu
Recent studies indicate that an excessive production of oxidants plays an important role in the pathogenesis of glomerular disease. Grape seed extract (GSE) is a potent antioxidant, and the aim of this pilot study was to evaluate its effect on puromycin-aminonucleoside (PAN)-induced nephrosis in rats. Fifty Sprague-Dawley rats were divided into five groups. Groups 1 and 2 rats received water from day 0 to day 30. Rats in groups 3, 4, and 5 received GSE at 10 mg/100g of body weight (BW), which was started on day 0, 6, and 3 of the experiment, respectively. In group 5 animals the GSE dose was increased (40 mg/100g BW) on day 9. Intraperitoneal dextrose (group 1) or PAN 15 mg/100g BW (groups 2-5) was administered on day 3. Urine and blood specimens were collected at regular intervals, and the comparison between the various groups was made by analysis of variance (ANOVA). Rats in all study groups (groups 3-5) showed a decrease in urine protein and serum cholesterol and triglyceride levels, which was statistically significant in group 3 animals. No significant changes were noted in serum albumin and creatinine levels. In conclusion, GSE administration decreases urine protein excretion and serum cholesterol and triglyceride levels in rats with PAN-induced nephrosis.
Anti-thrombotic effect of proanthocyanidin, a purified ingredient of grape seed.:
Thromb Res. 2005; 115(1-2):115-21 (ISSN: 0049-3848)
INTRODUCTION: Moderate and regular consumption of wine reduces the risk of acute coronary thrombotic events. The mechanism of the anti-thrombotic effect of wine is not clear. Extract or purified ingredients of grapes have not yet been studied for anti-thrombotic effect.
MATERIALS AND METHODS: Anti-thrombotic effect of proanthocyanidin, a highly purified ingredient of grape seed, was assessed by a shear-induced thrombosis test in vitro and by a laser-induced thrombosis test in the mouse carotid artery, in vivo. RESULTS AND CONCLUSIONS: Intravenously (20 mg/kg body weight, BW) or orally (2 x 200 mg/kg BW) administered proanthocyanidin significantly inhibited the laser-irradiation induced thrombus formation in the carotid artery (both P=0.01). Subsequent to oral administration of proanthocyanidin, in vitro platelet reactivity to shear stress has been inhibited. The latter suggests that the in vivo anti-thrombotic effect of proanthocyanidin may be due to a direct inhibitory effect on platelets.
The absorption, metabolism and excretion of flavan-3-ols and procyanidins following the ingestion of a grape seed extract by rats.:
Br J Nutr. 2005; 94(2):170-81 (ISSN: 0007-1145)Tsang C; Auger C; Mullen W; Bornet A; Rouanet JM; Crozier A; Teissedre PL.Plant Products & Human Nutrition Group, Graham Kerr Building, Division of Biochemistry & Molecular Biology, Institute of Biomedical & Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
Rats were fed a grape seed extract (GSE) containing (+)-catechin, (-)-epicatechin and dimers, trimers, tetramers and polymeric procyanidins. Liver, kidney, brain and gastrointestinal (GI) tract together with plasma, urine and faeces were collected over a 24 h period and their flavan-3-ol content was analysed by HPLC with tandem mass spectrometry and diode array detection. Small amounts of the GSE flavan-3-ols moved out of the stomach and into the duodenum/jejunum, and to a greater extent the ileum 1 h after ingestion, and into the caecum after 2 h with relatively small amounts being detected in the colon after 3 h. The GI tract contained the parent GSE flavan-3-ols and procyanidins with only trace amounts of metabolites and there were no indications that proanthocyanidins were depolymerised in the GI tract releasing monomeric flavan-3-ols. Plasma contained exclusively catechin glucuronides and methylated glucuronide metabolites which were also detected in the liver and kidneys. These metabolites were also present in urine together with sulphated metabolites and low amounts of the procyanidin dimers B1, B2, B3 and B4 as well as the trimer C2 and an unknown GSE trimer. The amounts of (+)-catechin and (-)-epicatechin metabolites excreted in urine relative to the quantity of the monomers ingested were 27 and 36 %, respectively, after 24 h. This is similar to the levels of urinary excretion reported to occur by other investigators after feeding (-)-epicatechin to rats and provides further, albeit indirect, evidence that the procyanidin oligomers in the GSE were not depolymerised to monomers to any extent after ingestion. No convincing analytical data were obtained for the presence of flavan-3-ol metabolites in the brain.
Grape seed extract prevents H(2)O(2)-induced chromosomal damage in human lymphoblastoid cells.:
Biol Pharm Bull. 2004; 27(9):1459-61 (ISSN: 0918-6158)
We examined the effects of grape seed extract (GSE) on chromosomal damage in two ways; induction on its own and prevention against treatment of reactive oxygen species (ROS). Chromosomal damage was evaluated by cytokinesis-block micronucleus assay (CBMN) in a human lymphoblastoid cell line, WIL2-NS cells. The GSE was composed of 89% proanthocyanidin with a degree of polymerization ranging from 2 to 15. GSE did not induce chromosomal damage in WIL2-NS cells at GSE concentrations up to 5 mg/l. In contrast, pretreatment with GSE dose-dependently prevented H(2)O(2)-induced chromosomal damage at an effective dose of 0.3 to 1 mg/l. A similar preventive effect of GSE was not detected in tert-butyl hydroperoxide-induced damage even at 5 mg/l. In a cell free system, GSE (<5 mg/l) directly scavenged H(2)O(2), but produced slight amounts of H(2)O(2) at higher concentrations (>50 mg/l). These results suggest that GSE is not genotoxic, but rather has an antigenotoxic effect against H(2)O(2) via direct scavenging action of H(2)O(2).
Mechanical evaluation of effect of grape seed proanthocyanidins extract on debilitated mandibles in rats.:
Dent Mater J. 2004; 23(2):67-74 (ISSN: 0287-4547)
Grape seed proanthocyanidins extract (GSPE), whose principal ingredient is proanthocyanidins, shows many activities such as cholesterol lowering effects, antioxidant effects, anti-tumor effects, cardioprotective effects, and protection against ultraviolet rays. However, reports of the effects of GSPE on bone are rare. We performed a mechanical analysis of the effect of GSPE on the interior structure of rat mandibular bone in the growth period, using three-dimensional peripheral quantitative computed tomography (pQCT). A low-calcium/high-calcium diet with supplementary GSPE was compared to a low-calcium/high-calcium diet in rats with debilitated mandibular bones. The group who received added GSPE showed a significant increase in cortical bone density, cross-sectional area, and trabecular bone mineral content (p<0.05). A significant increase was also seen in the results of a non-invasive stress strain index (SSI) (p<0.01) in the added GSPE. Our findings suggest that GSPE can increase bone quality and bone strength of rat mandibles in the growth period.
Amelioration of doxorubicin-induced myocardial oxidative stress and immunosuppression by grape seed proanthocyanidins in tumour-bearing mice.:
J Pharm Pharmacol. 2005; 57(8):1043-52 (ISSN: 0022-3573)Zhang XY; Li WG; Wu YJ; Gao MT.Department of Physiology, College of Basic Medicine, Lanzhou University, Lanzhou 730 000, People's Republic of China. zhangxiaoyuly@yahoo.com
We have investigated the protective effects of grape seed proanthocyanidins on doxorubicin-induced toxicity in tumour-bearing mice. The intraperitoneal administration of doxorubicin (2 mg kg(-1) every other day, cumulative dosage for 18 mg kg(-1)) significantly inhibited the growth of sarcoma 180, and induced myocardial oxidative stress with decreased superoxide dismutase and glutathione peroxidase activity while increasing malondialdehyde formation in the heart or serum. Doxorubicin-induced myocardial oxidative stress also reduced lactate dehydrogenase and creatine kinase activity in the heart and elevated their levels in the serum. Doxorubicin also affected immune functions of tumour-bearing mice with significantly decreased interleukin-2 (IL-2) and interferon-gamma (INF-gamma) production, and slightly decreased natural killer (NK) cell cytotoxicity, lymphocyte proliferation and CD4+/CD8+ ratio. It markedly increased the percentages of cytotoxic T cells (CD3+CD8+), helper T cells (CD3+CD4+), IL-2R+CD4+, and IL-2R+ cells as compared with untreated tumour-bearing mice. The intragastric administration of proanthocyanidin (200 mg kg(-1) daily) significantly inhibited tumour growth, and increased NK cell cytotoxicity, lymphocyte proliferation, CD4+/CD8+ ratio, IL-2 and INF-gamma production. Moreover, proanthocyanidin strongly enhanced the anti-tumour effect of doxorubicin and the above immune responses, and completely eliminated myocardial oxidative stress induced by doxorubicin. In conclusion, intragastric administration of proanthocyanidin could enhance the anti-tumour activity of doxorubicin and ameliorate doxorubicin-induced myocardial oxidative stress and immunosuppression in tumour-bearing mice.
Protective effect of grape seed polyphenols against high glucose-induced oxidative stress.:
Biosci Biotechnol Biochem. 2006; 70(9):2104-11 (ISSN: 0916-8451)Fujii H; Yokozawa T; Kim YA; Tohda C; Nonaka G.Institute of Natural Medicine, University of Toyama.
We aimed to clarify whether grape seed polyphenols (GSPs) are candidates therapeutic agents against diabetes mellitus, and to determine what degree of GSP oligomerization has the most potent efficacy. We studied the protective effects of various molecular weight GSPs (monomer, oligomer, polymer, and oligonol) on high glucose-induced cytotoxicity. In the present study, a high concentration of glucose (30 mM) induced cytotoxicity and oxidative stress (reactive oxygen species and nitric oxide) in cultured LLC-PK1 cells, but treatment with GSPs, especially oligomer GSPs, had potent protective effects against high glucose-induced oxidative stress. In addition, high glucose induced nuclear translocation of nuclear factor-kappa B, and increased expression of cyclooxygenase-2, inducible nitric oxide synthase, and bax, but GSP treatment inhibited them. These results indicate that GSPs have protective effects against high glucose-induced cytotoxicity, and among them, oligomer GSPs have more potent effects than other GSPs (monomer, polymer, and oligonol) on high glucose-induced renal cell damage.
Effects of vitamin C and grape-seed polyphenols on blood pressure in treated hypertensive individuals: results of a randomised double blind, placebo-controlled trial.:
Asia Pac J Clin Nutr. 2003; 12 Suppl:S18 (ISSN: 0964-7058)Ward NC; Hodgson JM; Croft KD; Clarke MW; Burke V; Beilin LJ; Puddey IB.School of Medicine and Pharmacology, University of Western Australia, West Australian Institute for Medical Research (WAIMR), Perth, Australia.
Background - Oxidative stress may contribute to the pathogenesis of hypertension and endothelial dysfunction via increased production of free radicals in the arterial wall. Objective - To investigate the effect of water-soluble antioxidants, vitamin C and polyphenols, on blood pressure (BP), endothelial function and oxidative stress in hypertensive individuals. Methods - 69 treated hypertensive individuals with a mean 24hr ambulatory systolic BP >=125 mmHg were involved in a randomised, double blind, placebo-controlled factorial trial. Following a 3-week washout, participants received either 500 mg/d vitamin C, 1000 mg/d grape-seed polyphenols, both vitamin C and polyphenols, or neither, for 6-weeks. At baseline and post-intervention, 24hr ambulatory BP, ultrasound assessed endothelium dependent and independent vasodilation of the brachial artery, and markers of oxidative damage, including plasma and urinary isoprostanes, oxidised low density lipoproteins and plasma tocopherols, were measured. Results - A significant interaction was observed, therefore results could not be analysed for main effects. In comparison to placebo, vitamin C lowered systolic BP (-1.8 +/- 0.8 mmHg, P=0.03), polyphenols did not significantly alter BP, but the combination of vitamin C and polyphenols significantly increased systolic (4.8+/- 0.9mmHg, P<0.0001), and diastolic (2.7+/- 0.6 mmHg, P<0.0001) BP. Endothelium-dependent and independent vasodilation, and markers of oxidative damage were not significantly altered. Conclusion - The combination of vitamin C and polyphenols significantly increased BP, but the mechanism remains to be elucidated.
Procyanidin B1 is detected in human serum after intake of proanthocyanidin-rich grape seed extract.:
Biosci Biotechnol Biochem. 2003; 67(5):1140-3 (ISSN: 0916-8451)
To confirm the absorption of proanthocyanidin (PA) into the human body, four healthy adults were administered 2.0 g of PA-rich grape seed extract (GSE). Blood were drawn before intake and 2 h after intake. Through the enzymatic treatment of sulfatase and beta-glucuronidase, blood samples were analyzed by HPLC coupled with mass-spectrometry (LC/MS). Procyanidin B1 [epicatechin-(4beta-->8)-catechin] was detected in human serum 2 h after intake. Its concentration was 10.6 +/- 2.5 nmol/l.
Grape seed proanthocyanidins induce pro-oxidant toxicity in cardiomyocytes.:
Cardiovasc Toxicol. 2003; 3(4):331-9 (ISSN: 1530-7905)Shao ZH; Vanden Hoek TL; Xie J; Wojcik K; Chan KC; Li CQ; Hamann K; Qin Y; Schumacker PT; Becker LB; Yuan CS.Tang Center for Herbal Medicine Research, Emergency Resuscitation Research Center, and Department of Medicine, Pritzker School of Medicine, University of Chicago, Chicago, IL, USA.
Grape seed proanthocyanidin extract (GSPE), a polyphenolic compound with antioxidant properties, may protect against cardiac ischemia and reperfusion injury. However, its potential toxicity at higher doses is unknown. The authors tested the effects of GSPE on reactive oxygen species (ROS) generation, cell survival, lactate dehydrogenase (LDH) release, and caspase- 3 activity using chick cardiomyocytes incubated with GSPE at 5, 10, 50, 100, or 500 micrograms/mL in medium for 8 h. Exposure to increasing concentrations of GSPE (100 or 500 micrograms/mL) resulted in an increase in ROS generation and cell death as measured by propidium iodide uptake and LDH release. Caspase-3 activity was significantly increased fourfold in cells exposed to GSPE 500 micrograms/ mL compared to controls; this was abolished by the selective caspase-3 inhibitor Ac-Asp-Gln-Thr-Asp-H (50 microM), which also significantly reduced the cell death resulting from GSPE (500 micrograms/mL). The antioxidant N-acetylcysteine (NAC, 100 microM) reduced cell death induced by GSPE (500 micrograms/mL) but failed to attenuate caspase-3 activation. Collectively, the authors conclude that higher doses of GSPE could cause apoptotic cell injury via effector caspase-3 activation and subsequent induction of ROS generation. Consumers may take higher doses of dietary supplements in the belief that natural herbs have no major side effects. This study demonstrates that dosages of GSPE should be optimized to avoid potential harmful pro-oxidant effects.
Dietary grape-seed proanthocyanidin inhibition of ultraviolet B-induced immune suppression is associated with induction of IL-12.:
Carcinogenesis. 2006; 27(1):95-102 (ISSN: 0143-3334)Sharma SD; Katiyar SK.Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL, USA and Birmingham VA Medical Center, Birmingham, AL 35294, USA.
We have shown previously that dietary grape seed proanthocyanidins (GSPs) inhibit UVB-induced photocarcinogenesis in mice. As UVB-induced immune suppression has been implicated in the development of skin cancer risk, we investigated whether dietary GSPs can modulate the effects of UVB on the immune system. We found that the UVB-induced (180 mJ/cm2) ear swelling response (inflammatory reaction) was significantly lower in mice fed with a GSP-supplemented (0.5 and 1.0%, w/w) diet than mice fed with the standard AIN76A diet. Dietary GSPs markedly inhibited UVB-induced (180 mJ/cm2) suppression of contact hypersensitivity responses in a local model of immunosuppression but had only moderate inhibitory effect in a systemic model of immunosuppression. Dietary GSPs reduced the UVB-induced increase in immunosuppressive cytokine interleukin (IL)-10 in skin and draining lymph nodes compared with mice that did not receive GSPs. In contrast, GSPs enhanced the production of immunostimulatory cytokine IL-12 in the draining lymph nodes. Intraperitoneal injection of GSPs-fed mice with a neutralizing anti-IL-12 antibody abrogated the protective effects of the GSPs against UVB-induced suppression of the contact hypersensitivity response. These data indicate for the first time that GSPs modulate UVB-induced immunosuppression and suggest that this may be one of the possible mechanisms by which they prevent photocarcinogenesis in mice.
Metabolism of grape seed polyphenol in the rat.:
J Agric Food Chem. 2003; 51(24):7215-25 (ISSN: 0021-8561)
The metabolism of grape seed polyphenol (GSP) has been investigated in rats by high-performance liquid chromatography analysis of the serum and urinary concentrations of the GSP metabolites (+)-catechin (CT), (-)-epicatechin (EC), 3'-O-methyl-(+)-catechin, and 3'-O-methyl-(-)-epicatechin. The serum concentration of these four metabolites reached a maximum 3 h after the oral administration of GSP. The urinary excretion of these GSP metabolites accounted for 0.254% (w/w) of the administered dose of GSP (1.0 g/kg), and the majority of these metabolites were excreted within 25 h of oral administration. The serum concentration and urinary excretion of these metabolites were also compared after the oral administration of different GSP monomers (gallic acid, CT, and EC), normal GSP, and the high molecular weight components of GSP (GSPH). No metabolites were detected in the serum of rats given GSPH. The urinary percentage excretion of the GSP metabolites derived from the respective monomers (CT or EC) did not vary with the administration of different substances (CT or EC, GSP, or GSPH). Taken together, these results suggest that only the monomers of GSP are absorbed and metabolized.
Grape seed proanthocyanidins extract promotes bone formation in rat's mandibular condyle.:
Eur J Oral Sci. 2005; 113(1):47-52 (ISSN: 0909-8836)
We studied the effect of dietary supplementation with grape seed proanthocyanidins extract (GSPE) 3 mg added in 100 g high-calcium diet with a calcium content of 1697 mg 100 g(-1) on mandibular condyle bone debility, which was induced by a low-calcium diet. Forty Wistar male rats, 5 week old, were randomly divided into control (Co), low-calcium diet (LC), low-calcium/high-calcium diet (LCH), and low-calcium/high-calcium with supplementary GSPE diet (LCHG) groups for 6 wk. Bone formation of the mandibular condyle was measured using peripheral quantitative computed tomography (pQCT). Significant differences were not seen among the four groups for body weight, measured weekly. The LCHG group scored significantly higher in cortical bone density, total bone cross-sectional area, cortical bone cross-sectional area, cortical bone mineral content, total bone density, total bone mineral content, and in the stress-strain index to the reference axis x when compared with the LCH group. We concluded that a high-calcium diet combined with GSPE supplementation is more effective in reversing mandibular condyle bone debility in rats than is a low-calcium diet, standard diet, or high-calcium diet alone.
Grape seed extract inhibits EGF-induced and constitutively active mitogenic signaling but activates JNK in human prostate carcinoma DU145 cells: possible role in antiproliferation and apoptosis.:
Oncogene. 2003; 22(9):1302-16 (ISSN: 0950-9232)Tyagi A; Agarwal R; Agarwal C.Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver 80262, USA.
A loss of functional androgen receptor and an enhanced expression of growth factor receptors and associated ligands are causal genetic events in prostate cancer (PCA) progression. These genetic alterations lead to an epigenetic mechanism where a feedback autocrine loop between membrane receptor and ligand (e.g. EGFR-TGFalpha) results in a constitutive activation of MAPK-Elk1-AP1-mediated mitogenic signaling in human PCA at an advanced and androgen-independent stage. We rationalized that inhibiting these epigenetic events could be useful in controlling advanced PCA growth. Recently, we found that grape seed extract (GSE), a dietary supplement rich in flavonoid procyanidins, inhibits advanced and androgen-independent human PCA DU145 cell growth in culture and nude mice. Here, we performed detailed mechanistic studies to define the effect of GSE on EGFR-Shc-MAPK-Elk1-AP1-mediated mitogenic signaling in DU145 cells. Pretreatment of serum-starved cells with GSE resulted in 70% to almost complete inhibition of EGF-induced EGFR activation and 50% to complete inhibition of Shc activation, which corroborated with a comparable decrease in EGF-induced Shc binding to EGFR. Conversely, EGF-induced ERK1/2 phosphorylation was inhibited only by lower doses of GSE; in fact, higher doses showed an increase. Additional studies showed that GSE alone causes a dose- and time-dependent increase in ERK1/2 phosphorylation in starved DU145 cells that is inhibited by an MEK1 inhibitor PD98059. Independent of this increase in ERK1/2 phosphorylation, GSE showed a strong inhibition of ERK1/2 kinase activity to Elk1 in both cellular and cell-free systems. GSE treatment of cells also inhibited both EGF-induced and constitutively active Elk1 phosphorylation and AP1 activation. GSE treatment also showed DNA synthesis inhibition in starved and EGF-stimulated cells as well as loss of cell viability and apoptotic death that was further increased by adding MEK1 inhibitor. Since GSE strongly induced apoptosis independent of its affect on an increase in phospho-ERK1/2, we hypothesized that apoptotic effect of GSE could be by other mechanism(s) including its effect on stress-associated MAPK, the JNK. Indeed, GSE-treated cells showed a strong and sustained increase in phospho-JNK1/JNK2 levels, JNK activity and phospho-cJun levels. An inhibition of GSE-induced JNK activation by a novel JNK inhibitor SP600125 resulted in a significant reversal of GSE-induced apoptotic death suggesting the involvement of JNK activation by GSE in its apoptosis response. Together, these results suggest that anticancer effects of GSE in PCA be mediated via impairment of EGFR-ERK1/2-Elk1-AP1-mediated mitogenic signaling and activation of JNK causing growth inhibition and apoptosis, respectively.
Ethanol and polyphenolic free wine matrix stimulate the differentiation of human intestinal Caco-2 cells. Influence of their association with a procyanidin-rich grape seed extract.:
J Agric Food Chem. 2005; 53(14):5541-8 (ISSN: 0021-8561)Laurent C; Besan??on P; Caporiccio B.Equipe Nutrition et Aliments EA 3762, Universit?? Montpellier II, 34095 Montpellier, France. cbabot@yahoo.com
The effect of daily contact with ethanol on Caco-2 cell differentiation was investigated. Pure ethanol (1%) and a polyphenolic free wine matrix (polyphenol-free wine containing 1% ethanol) associated or not with a procyanidin-rich grape seed extract (GSE) were added to Caco-2 cells from confluency for 2 h a day after successive incubation in salivary, gastric, and pancreatic media. Treatment with 1% ethanol did not appear to be cytotoxic to cells, but it also stimulated Caco-2 cell differentiation, particularly in the first days following confluency, and this effect was more marked when associated with polyphenolic free wine matrix constituents. This activation resulted in an increase in microvillar density, organization, and elongation (+70%) and was associated with strong stimulation of sucrase-isomaltase (+780%) and a concomitant regular increase in cell protein content (+50-88%). While the presence of GSE in alcoholic solutions did not modify the morphological pattern observed in cells subjected to ethanol and polyphenolic free wine matrix alone, it had a clear reducing effect on their microvillus elongation (-30%). However, these stimulating effects of ethanol on morphological differentiation were attenuated from day 10 postconfluency, which could suggest cell cytoprotection against ethanol. These are the first results in support of the notion that moderate concentration of ethanol may stimulate the differentiation of Caco-2 cells, particularly when integrated with a polyphenolic free wine matrix.
Pre- and post-mortem use of grape seed extract in dark poultry meat to inhibit development of thiobarbituric acid reactive substances.:
J Agric Food Chem. 2003; 51(6):1602-7 (ISSN: 0021-8561)Lau DW; King AJ.Department of Food Science and Animal Science, University of California, Davis 95616, USA.
Diets containing grape seed extract (GSE)-control, GSE [low GSE, low GSE + methionine, high GSE, and high GSE + methionine], or alpha-tocopherol-were fed to broiler chicks to estimate the antioxidative activity of GSE in processed meat. GSE was detrimental to the growth of chicks, and methionine did not reverse the detrimental effect. GSE with 85.4 g of gallic acid equiv/100 g (GAE 85.4) was added to ground dark turkey meat to obtain treatments with no GSE, 1.0% GSE, and 2.0% GSE and then processed as unsalted or salted and unheated or heated. Processed treatments were analyzed for thiobarbituric acid reactive substances (TBARS) and percent expressible moisture (%EM). GSE at 1.0 and 2.0% decreased TBARS values nearly 10-fold as compared to the control. GSE (1.0%) had a %EM value significantly greater than that of the control. GAE 85.4 decreased TBARS values more than GAE 88.9.
Grape seed procyanidins prevent oxidative injury by modulating the expression of antioxidant enzyme systems.:
J Agric Food Chem. 2005; 53(15):6080-6 (ISSN: 0021-8561)Puiggros F; Ll??piz N; Ard??vol A; Blad?? C; Arola L; Salvad?? MJ.Departament de Bioqu??mica i Biotecnologia, Unitat d'Enologia del Centre de Refer??ncia en Tecnologia dels Aliments de la Generalitat de Catalunya, Universitat Rovira i Virgili, 43007 Tarragona, Spain.
In the present paper, we report the effect of a grape seed procyanidin extract (GSPE) on antioxidant enzyme systems (AOEs). Gene expression was tested using the hepatocarcinoma cell line HepG2 by exposing it to several GSPE doses between 0 and 100 mg/L for 24 h. We evaluated mRNA expression and enzyme activity levels using real time RT-PCR and spectrophotometry. The results suggested a transcriptional GSPE regulation of glutathione related enzymes caused by an increase both in mRNA and in enzyme activity levels overall at 15 mg/L. We also assessed the GSPE effect on AOEs in cells submitted to oxidative stress. Under oxidative conditions (1 mM H(2)O(2), 1 h), we found a decrease in GSH content and an increase in MDA, and we suggested a posttranslational regulation of GPx/GR mRNAs and a transcriptional enhancement of GST mRNA. The GSPE pretreatment (15 mg/L, 23 h) before HepG2 submission to H(2)O(2) (1 mM, 1 h) showed an increase of the mRNA of GPx/GR with respect to the H(2)O(2) group, whereas the GSH content was similar to the control group. However, the GPx/GR enzyme activities were not increased. We hypothesize that GSPE probably improves the cellular redox status via glutathione synthesis pathways instead of regulation of the GPx and/or GR activities protecting against oxidative damage.
Fractionation of grape seed extract and identification of gallic acid as one of the major active constituents causing growth inhibition and apoptotic death of DU145 human prostate carcinoma cells.:
Carcinogenesis. 2006; 27(7):1445-53 (ISSN: 0143-3334).Veluri R; Singh RP; Liu Z; Thompson JA; Agarwal R; Agarwal C.Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver, CO 80262, USA.
The anti-cancer efficacy of grape seed extract (GSE) against prostate cancer (PCA) via its anti-proliferative, pro-apoptotic and anti-angiogenic activities in both cell culture and animal models have recently been described by us. GSE is a complex mixture containing gallic acid (GA), catechin (C), epicatechin (EC) and several oligomers (procyanidins) of C and/or EC, some of which are esterified to GA. To determine which components are most active against PCA, an ethyl acetate extract of GSE was separated by reverse-phase high-performance liquid chromatography (HPLC) into three fractions. Fraction 1 was far more effective than others in causing growth inhibition and apoptotic death of human PCA DU145 cells. Of the components in this fraction, GA showed a very strong dose- and time-dependent growth inhibition and apoptotic death of DU145 cells, but C and procyanidins B1 (EC-C dimer), B2 (EC-EC dimer) and B3 (C-C dimer) were nearly ineffective. Mechanistic studies demonstrated a strong caspase-9, caspase-3 and poly (ADP-ribose) polymerase (PARP) cleavages by GA in DU145 cells. Procyanidin oligomers eluting in HPLC Fractions 2 and 3 were obtained in larger quantities by separating GSE into eight fractions (I-VIII) on a gel filtration column. All fractions were analyzed by HPLC-UV and negative-ion electrospray mass spectrometry. Fractions I-III contained the active compound GA and inactive components C, EC, B1 and B2. Fraction IV contained other dimers and a dimer-GA ester and was also less active than GSE in DU145 cells. Fractions V-VIII, however, caused significant growth inhibition and apoptosis with the highest activity present in the later fractions that contained procyanidin trimers and GA esters of dimers and trimers. Together, these observations identify GA as one of the major active constituents in GSE. Several procyanidins, however, and especially the gallate esters of dimers and trimers also may be efficacious against PCA and merit further investigation.
Age associated changes in erythrocyte membrane surface charge: Modulatory role of grape seed proanthocyanidins.:
Exp Gerontol. 2005; 40(10):820-8 (ISSN: 0531-5565)Sangeetha P; Balu M; Haripriya D; Panneerselvam C.Department of Medical Biochemistry, Dr AL Mudaliar Post Graduate Institute of Basic Medical Sciences, University of Madras, Taramani Campus, Chennai 600 113, India.
Aging, a multifactorial process of enormous complexity is characterized by impairment of physio-chemical and biological aspects of cellular functions. It is closely associated with increased free radical production, which situation ultimately leads to devastation of normal cell function and membrane integrity. The present study was aimed to determine the effect of proanthocyanidins rich grape seed extract (GSP) on membrane surface charge density in erythrocytes during animal age associated oxidative stress. GSP (100 mg/day/kg body weight) was administered orally for 15 and 30 days to young and aged rats. Significant decrease in surface charge levels with concomitant increase in protein carbonyls and decrease in glycoprotein, antioxidants status was noted in erythrocytes of aged rats when compared with young rat erythrocytes. Duration dependent supplementation of GSP increased the erythrocyte surface charge density to near normalcy in aged rats. Decrease in protein carbonyls level and increase in glycoproteins as well as antioxidant status was observed in aged rat erythrocytes on GSP treatment. Thus, from our results, we conclude that GSP is an effective anti-aging drug in preventing the oxidative stress associated loss of membrane surface charge, which thereby maintains the erythrocyte membrane integrity and functions in elderly.
In vivo antioxidant activity of procyanidin-rich extracts from grape seed and pine (Pinus maritima) bark in rats.:
Int J Vitam Nutr Res. 2006; 76(1):22-7 (ISSN: 0300-9831).Busserolles J; Gueux E; Balasi??ska B; Piriou Y; Rock E; Rayssiguier Y; Mazur A.Centre de Recherche en Nutrition Humaine d'Auvergne, Unit?? des Maladies M??taboliques et Micronutriments, INRA, Theix, Saint-Gen??s-Champanelle, France.
BACKGROUND: In vitro evidence exists for the potential antioxidant benefits of procyanidin-rich extracts, but in vivo studies are scarce. We have evaluated the effects of selected procyanidin-rich extracts on oxidative stress in rats in condition of prolonged consumption of these compounds and also after single administration i.e. in postprandial conditions.
METHODS: Rats were fed for 8 weeks with diets supplemented with either a grape seed extract (GE), a pine bark extract (PE), or a high-degree polymerized pine bark extract (HPE). An additional study was performed in order to assess the postprandial effect of these extracts on plasma antioxidant capacity. The ferric-reducing antioxidant power (FRAP) and thiobarbituric acid-reactive substances (TBARS) were determined in plasma. For lipid peroxidation study of heart tissue, homogenates were prepared and TBARS were measured after lipid peroxidation induced by FeSO4-ascorbate.
RESULTS: After 8 weeks of dietary treatment, total antioxidant capacity in plasma was significantly higher in the GE and PE groups as compared with the other two groups. Plasma TBARS concentrations and heart susceptibility to peroxidation were not significantly different between the groups. In the postprandial state, by comparing plasma antioxidant capacity 2 hours after ingestion of the different procyanidin-rich extracts (500 mg/kg body weight), we observed that FRAP values were higher in the procyanidin-rich extracts groups as compared with the control group. Moreover, plasma FRAP concentration was significantly higher in the GE group as compared with the other groups.
CONCLUSION: The results of the present experiment constitute positive evidence for an in vivo antioxidant effect at the plasma level of procyanidin-containing plant extracts.
90-day oral toxicity study of a grape seed extract (IH636) in rats.:
J Agric Food Chem. 2002; 50(7):2180-92 (ISSN: 0021-8561).Wren AF; Cleary M; Frantz C; Melton S; Norris L.The Wren Group, 900 South Meadows Parkway, Suite 2013, Reno, Nevada 89511, USA.
To assess the safety of grape seed extract with less than 5.5% catechin monomers (IH636), 4 groups of male and female Sprague-Dawley rats were provided grape seed extract in the diet at levels of 0 (control), 0.5, 1.0, or 2.0% for a period of 90 days. All animals survived the duration of the study, and no significant changes in clinical signs, hematological parameters, organ weights, ophthalmology evaluations, or histopathological findings were observed. A significant increase in food consumption was observed in male and female rats provided the grape seed extract diets compared to that of the control rats, especially in male rats consuming 2.0% grape seed extract. This effect was not accompanied by increases in body weight gains. Grape seed extract appeared to increase the insoluble fraction of the diet. Male rats in the high-dose group exhibited decreased serum iron levels and decreased serum iron/total iron binding capacity ratio compared to those of the controls, although all values were within historical ranges for Sprague-Dawley rats. In conclusion, administration of the grape seed extract IH636 to male and female Sprague-Dawley rats in the feed at levels of 0.5, 1.0, or 2.0% for 90 days did not induce any significant toxicological effects.
Polymeric grape-seed procyanidins, but not monomeric catechins and oligomeric procyanidins, impair degranulation and membrane ruffling in RBL-2H3 cells.:
Bioorg Med Chem. 2006; 14(3):641-9 (ISSN: 0968-0896)
Grape-seed proanthocyanidins (GSPs) are catechin polymers that are predicted to form helices in their global minimum-energy conformation and to have a mean degree of polymerization of seven (mDP = 7). The highly polymerized GSP-H fraction (mDP = 10) was found to impair degranulation in RBL-2H3 cells after stimulation with an antigen (Ag) and treatment with the Ca-ATPase inhibitor thapsigargin (Tg). In addition, GSP-H affected actin cytoskeleton and inhibited membrane ruffling in these cells, resulting in the suppression of exocytosis. By contrast, monomeric epicatechin, the dimeric procyanidins PA-1, PA-2, and PB-2, and the oligomerized GSP-L (mDP = 3) had no effect on membrane ruffling and degranulation. These findings indicate that the molecular size and length of GSP-H are needed for the inhibition of membrane ruffling and degranulation in RBL-2H3 mast-cell lines.
Grape Seed Extract vs. Colon Cancer:
Oct. 19, 2006 -- Grape seed extract may pack a punch against colon cancer, early lab tests show.
But the researchers -- who work at the University of Colorado -- arent making any promises yet.
"The value of this preclinical study is that it shows grape seed extract can attack cancer, and how it works, but much more investigation will be needed before these chemicals can be tested as a human cancer treatment and preventive," says Rajesh Agarwal, PhD, one of the researchers.
"With these results, we are not suggesting that people run out and buy and use grape seed extract," Agarwal says in a news release from the American Association for Cancer Research. "That could be dangerous since so little is known about doses and side effects."
Testing the Extract:
In their study, the researchers began by testing various doses of grape seed extract on human colon cancer cells in test tubes.
The cancer cells treated with grape seed extract were more likely to halt their normal growth cycle and die, compared to those not treated with the extract.
Cancer cells that got the biggest doses of grape seed extract for the longest time were the most likely to halt their growth cycle.
Those results were "encouraging," the researchers write.
So the scientists took their experiment one step further.
They injected human colon cancer tumors under the skin of mice, then funneled grape seed extract into the mices mouths through a tube.
The tumors grew more slowly in those mice, compared to the tumors in mice that didnt get grape seed extract.
The mice that got grape seed extract didnt gain weight, change their diets, or show other side effects during the eight-week study.
Agarwal and his colleagues write that antioxidants called proanthocyanidins may be responsible for grape seed extracts effects.
SOURCES: Kaur, M. Clinical Cancer Research, Oct. 15, 2006; vol 12: pp 6194-6202. News release, American Association for Cancer Research.
Grape Seed Extract for Blood Pressure?:
March 27, 2006 -- Grape seed extract might help control blood pressure, a new study shows.
The news comes from scientists at the University of California at Davis. They studied 24 adults with metabolic syndrome, a cluster of abnormalities that raises the risk of diabetes and heart disease.
Most participants had prehypertension; the others had normal blood pressure. Prehypertension falls between normal and high blood pressure.
Participants were split into three groups. One group got a fake (placebo) treatment. Another group took 150 milligrams per day of grape seed extract. The third group took 300 milligrams daily of the same grape seed extract.
Four weeks later, blood pressure had dropped for those taking grape seed extract. Blood pressure didn't change in the placebo group.
The findings were presented in Atlanta at the American Chemical Society's National Meeting & Exposition.
Blood Pressure Effects
The two extract doses were "equally efficient in lowering the blood pressure," researcher C. Tissa Kappagoda, MD, PhD, tells WebMD. Kappagoda specializes in cardiovascular medicine at the University of California at Davis.
Systolic blood pressure (the top number in a blood pressure reading) dropped 12 points, on average. Diastolic blood pressure (the bottom number in a blood pressure reading) dropped an average of 8 points, Kappagoda says.
The blood pressure changes were seen "across the board," not just in people with prehypertension, says Kappagoda. He notes that his team had already done preliminary work on people with normal blood pressure and found no dangerous drop in blood pressure with grape seed extract.
Kappagoda and colleagues had previously found that grape seed extract might help dilate blood vessels. "Because of that, we thought it had potential as a blood-pressure-lowering agent," Kappagoda says.
In Kappagoda's latest study, LDL "bad" cholesterol levels also fell in people taking the higher extract dose, but not in those taking the lower dose.
Talk to Doctor First
Grape seed extract isn't marketed with any health claims, Kappagoda points out.
He says people who are thinking about taking grape seed extract should consult their doctors. Kappagoda also says he "wouldn't want to recommend" that anyone stop taking medicine for high blood pressure in favor of grape seed extract.
"But on the other hand, if their physicians have said, 'You know, you're kind of borderline,' and so on, then there may be some place for this after consulting their physician," Kappagoda says, pointing out that doctors usually recommend lifestyle change -- not prescription drugs -- to treat prehypertension.
The study was funded by the extract's maker, Polyphenolics Inc.
SOURCES: American Chemical Society National Meeting & Exposition, Atlanta, March 26-30, 2006. Tissa Kappagoda, MD, PhD, professor of internal medicine (cardiovascular medicine), University of California, Davis. News release, University of California, Davis. News release, American Chemical Society.
Grape Seed Extract Inhibits Colorectal Cancer Cell Growth:
NEW YORK (Reuters Health) Oct 30 - Grape seed extract (GSE) inhibits the growth of human colorectal carcinoma cells in vitro and in animal models, according to a report in the October 15th issue of Clinical Cancer Research.
GSE has previously been shown to reduce the incidence of carcinogen-induced mammary tumors and skin tumors in animal models and to inhibit the growth of various human cancer cells in vitro and in vivo, the authors explain.
Dr. Chapla Agarwal and colleagues from the University of Colorado at Denver investigated the anticancer activity of GSE against human colon carcinoma cells in cell culture and xenograft studies.
GSE dose-dependently and time-dependently inhibited growth and induced apoptotic cell death in HT29 and LoVo human colorectal cancer cell lines, the researchers report.
GSE treatment was associated with cell cycle arrest (predominantly in G1 phase) and increases in both cytoplasmic and nuclear levels of Cip1/p21 protein, the results indicate. "Growth inhibitory and apoptotic effects of GSE against colorectal cancer could be mediated via an up-regulation of Cip1/p21," Dr. Agarwal and colleagues surmise.
Feeding GSE to athymic nude mice inhibited HT29 colon carcinoma tumor xenograft growth without causing toxicity, the researchers note, and tumor samples from these animals confirmed the antiproliferative and pro-apoptotic effects of GSE.
"GSE may be an effective chemopreventive agent against colorectal cancer," the team concludes. "A dose-dependent in vivo study with GSE is needed in the future that would provide additional information regarding the lowest effective as well as highest nontoxic doses of GSE."
Clin Cancer Res 2006;12:6194-6202.
Grape Seed Extract Has Modest Blood Pressure-Lowering Effect:By Martha Kerr
NEW YORK (Reuters Health) Apr 04 - Polyphenolic compounds derived from grape seed appear to lower blood pressure in patients with metabolic syndrome, investigators reported last week at the 231st national meeting of the American Chemical Society in Atlanta.
The study, conducted by Dr. C. Tissa Kappagoda and co-investigators at the University of California, Davis, involved 24 men and women who met the criteria for metabolic syndrome set by the Adult Treatment Panel of the National Cholesterol Education Program. "They weren't chosen for high blood pressure, per se," Dr. Kappagoda told Reuters Health.
The subjects were randomized into one of three treatment arms: one group received 150 mg grape seed extract daily, a second received 300 mg daily and a third received placebo.
Parameters of the metabolic syndrome were measured at baseline and at the end of the 4-week study period. These included blood glucose, plasma insulin and insulin resistance, serum lipids and oxidized low-density lipoprotein cholesterol. Blood pressure was measured using an ambulatory blood pressure monitoring device over a 12-hour period. Platelet aggregation and activation were also evaluated at the beginning and end of the study.
Baseline blood pressures ranged from 120-130 mm Hg systolic and below 80 mm Hg diastolic. After four weeks of treatment, subjects receiving grape seed extract had a drop in systolic blood pressure of approximately 12 mm Hg and in diastolic blood pressure of approximately 9 mm Hg, Dr.
Kappagoda told Reuters Health.
Perhaps more significant was the change in oxidized LDL, he added. "The higher the oxidized LDL at baseline, the greater the drop in blood pressure," he said.
Polyphenolic compounds, derived from gallic acid in grape seed extract, cause a release in nitric oxide, which in turn results in an endothelium-dependent relaxation, Dr. Kappagoda explained. "These compounds are found in brightly colored fruits and vegetables," he pointed out. While the effects were statistically significant, "we are not suggesting this is a treatment for hypertension."
The UC Davis team is continuing their investigation, studying the effects of grape seed extract in patients specifically selected for prehypertension. Those data are expected to be available this summer.
The study was funded by Polyphenolics, Inc. of Madera, California.
Phlebotonics for venous insufficiency:MJ Martinez
Background:Chronic venous insufficiency (CVI) is a common condition caused by inadequate blood flow through the veins, usually in the lower limbs. It can result in considerable discomfort with symptoms such as pain, itchiness and tiredness in the legs. Sufferers may also experience swelling and ulcers. Phlebotonics are a class of drugs that are often used to treat CVI.
Objectives:To assess the efficacy of oral or topical phlebotonics.
Search strategy:We searched the Cochrane Peripheral Vascular Diseases Group trials register (April 2005), the Cochrane Central Register of Controlled Trials ( Issue 2, 2005), MEDLINE (January 1966 to April 2005), EMBASE (January 1980 to April 2005) and reference lists of articles. We also contacted pharmaceutical companies.
Selection criteria:Randomised, double blind, placebo 每 controlled trials (RCTs) assessing the efficacy of rutosides, hidrosmine, diosmine, calcium dobesilate, chromocarbe, centella asiatica, disodium flavodate, french maritime pine bark extract, grape seed extract and aminaftone in CVI patients at any stage of the disease.
Data collection and analysis:Two reviewers independently extracted data and assessed trial quality. The effects of treatment were estimated by relative risk (RR) or by standardised mean differences (SMD) by applying a random effects statistical model. Sensitivity analyses were also performed.
Main results:Fifty 每 nine RCTs of oral phlebotonics were included, but only 44 trials involving 4413 participants contained quantifiable data for the efficacy analysis: 23 of rutosides, ten of hidrosmine and diosmine, six of calcium dobesilate, two of centella asiatica, one of french maritime pine bark extract, one of aminaftone and one of grape seed extract. No studies evaluating topical phlebotonics, chromocarbe, naftazone or disodium flavodate fulfilled the inclusion criteria.
Outcomes included oedema, venous ulcers, trophic disorders, subjective symptoms (pain, cramps, restless legs, itching, heaviness, swelling and paraesthesias), global assessment measures and side effects. The results of many variables were heterogeneous. Phlebotonics showed some global benefit (i.e. oedema reduction) (relative risk 0.72, 95% confidence interval 0.65 to 0.81). The benefit for the remaining CVI signs and symptoms must be evaluated by phlebotonic group. There were no quantifiable data on quality of life.
Authors' conclusions:There is not enough evidence to globally support the efficacy of phlebotonics for chronic venous insufficiency. There is a suggestion of some efficacy of phlebotonics on oedema but this is of uncertain clinical relevance. Due to the limitations of current evidence, there is a need for further randomised, controlled clinical trials with greater attention paid to methodological quality.
Flavonoid-Rich Grapeseed Extracts: A New Approach In High Cardiovascular Risk Patients?:P. Kar; D. Laight; K. M. Shaw; M. H. Cummings
Summary and Introduction
The management of traditional risk factors such as hypertension and dyslipidaemia has been successful in reducing the development of cardiovascular disease. However, this has not resulted in the amelioration of complications; prompting attention to be focused on novel markers of vascular risk such as endothelial dysfunction (a determinant of vascular tone), vascular inflammation, oxidative stress and insulin resistance. With an ever-growing interest in plant-derived products, agents that could have a beneficial effect on this complex web of pathophysiology have thus been a major area of research and interest. Flavonoids have been a major focus of attention since the days of the French paradox and the presence of high quantity of flavonoids in grapeseed extracts has prompted research looking at its effects on novel markers of vascular risk.
This review briefly summarises mechanisms implicated in the development of vascular disease and then focuses upon the potential role of the antioxidant properties of flavonoid-rich grapeseed extracts in the reversal of these processes.
Introduction
In the last few decades, a number of metabolic factors that are implicated in the pathogenesis of cardiovascular disease have been identified. They include dyslipidaemia, hypertension, altered glucose tolerance and hyperinsulinaemia.
However, while the management of these traditional vascular risk factors such as hypertension and dyslipidaemia has been successful in reducing the development or progression of cardiovascular disease, it has not totally ameliorated this complication.
Novel markers of vascular risk that have received much attention include endothelial dysfunction (a determinant of vascular tone), vascular inflammation, oxidative stress and insulin resistance. Recent reviews have focused upon the links between vascular tone and inflammation, oxidative stress, insulin sensitivity and the aetiology and pathogenesis of high cardiovascular risk-linked pathologies such as diabetes. With an ever-growing interest in plant-derived products, agents that could have a beneficial effect on this complex web of pathophysiology have thus been a major area of research and interest. This review briefly summarises mechanisms implicated in the development of vascular disease and then focuses upon the potential role of plant-derived flavonoids, especially flavonoid-rich grapeseed extracts, in the reversal of these processes.
The 'Complex' Linkage
Stern proposed the Common Soil hypothesis in 1995 (linking atherosclerosis, inflammation and the metabolic syndrome) and there has been an abundance of evidence linking these conditions and the complex pathophysiology underlying the metabolic syndrome.
The vascular endothelium plays a key role in the regulation of vascular homeostasis and increasing evidence suggests that alterations in endothelial function contribute to the pathogenesis and clinical expression of cardiovascular disease.[2] Endothelial cells regulate vascular homeostasis by producing factors that act locally in the vessel wall and lumen, and a key endothelial product is nitric oxide (NO). Impaired NO production, increase in peroxynitrite formation, a pro-inflammatory milieu, and endothelial damage are some of the mechanisms leading to impaired vasodilatory capacity and endothelial dysfunction. In 1999, Duffy et al. revealed the importance of endothelial functional integrity in atherosclerosis and coronary ischaemia.[3] Atherosclerosis was seen to be dependent upon metabolic vasodilation via tonic release of endothelial NO and vasodilator prostanoids.[4]
Abnormal endothelial function is however often portrayed as a simple reduction in NO-mediated endothelium-dependent vasodilator responses, when this is just one feature of a more complex pattern of changes. Indeed, as vasodilator function decreases, there is increased vasoconstriction due to excess endothelin-1 (ET-1) synthesis[5,6] and other vasoconstricting agents. These alterations in vascular tone are coincident with a spectrum of pro-inflammatory and pro-thrombotic vascular changes. Endothelial dysfunction is now considered important both as a target and mediator in high cardiovascular risk patients such as type 2 diabetes.[7,8]
Oxidative stress has a deleterious effect upon endothelial function and is a major culprit in the complex web of factors propagating the syndrome of insulin resistance and type 2 diabetes mellitus (DM). Unsurprisingly, this has prompted a great deal of research and investigations looking at the effects of antioxidants on endothelial function, because of the therapeutic modality of reducing oxidative stress.
Atherosclerosis and the Role of Oxidative Stress upon Endothelial Function
Atherosclerosis is a chronic inflammatory disease that develops in lesion-prone regions of medium-sized arteries. As is well known, they may be present and clinically silent for decades before becoming active and producing clinical events such as cardiac death and cerebrovascular events.
Atherosclerosis can be taken as an example of a process for which there is substantial evidence of the effect of oxidative stress. Hypercholesterolaemia is universally accepted as a major risk factor for atherosclerosis. However, at any given concentration of plasma cholesterol, there is still great variability in the occurrence of cardiovascular events. One of the major breakthroughs in atherogenesis research has been the realisation that oxidative modification of low-density lipoprotein (LDL) might be a crucially important step in the development of the atherosclerotic plaque.[10,11] Native LDL causes the formation of foam cells from monocyte-derived macrophages in early atherosclerotic lesions only after the modification of LDL by various chemical reactions such as oxidation. As oxidation of LDL is primarily a free radical-mediated process that is inhibited by antioxidants, antioxidant depletion might be a risk factor for cardiovascular disease. Inactivation of endothelium-derived vasodilator NO is caused by vascular superoxide anion.Nitric oxide combines with superoxide intravascularly to create peroxynitrite, a cytotoxic molecule, which then oxidises LDL.
Evidence for LDL oxidation in vivo is now well established. In immunocytochemical studies, antibodies against oxidised LDL stain atherosclerotic lesions but not normal arterial tissue. In young survivors of myocardial infarction (MI), an association has been demonstrated between increased susceptibility of LDL to oxidation and the degree of coronary atherosclerosis, whereas the presence of ceroid, a product of lipid peroxidation, has been shown in advanced atherosclerotic plaques.[15]
Apart from the atherogenic consequences of LDL oxidation, it is increasingly recognised that reactive oxygen and nitrogen species directly interact with signalling mechanisms in the arterial wall to regulate vascular function.[16]
The effects of antioxidants on these processes are complex but provide alternative mechanisms by which antioxidant supplementation might ameliorate vascular pathology, for instance by improving endothelial function.
Evidence suggests that the balance between pro-oxidant properties of peroxynitrite formed from NO and antioxidant properties of plasma and NO determines the progression of pathology in atherosclerosis.
The Role of Inflammation in the Metabolic Syndrome
In recent years inflammation has also been found to play a key role in the causation of atherosclerotic changes and C-reactive protein (CRP) has been deemed as an important biological marker of inflammation that has been linked to cardiovascular disease.[19] Profound interest has been generated in inflammation being a mediator in insulin resistance and furthermore in the pathogenesis and complications of type 2 diabetes.[20]
Current evidence suggests that oxidative stress, insulin resistance, endothelial dysfunction (in the peripheral arterial bed) and more recently inflammation might be acting synergistically, to promote the development of the metabolic syndrome and subsequent pathology.[21,22] Chronic subclinical inflammation is widely being recognised as a key component in the aetiology of macrovascular disease and possibly even the metabolic syndrome and type 2 diabetes [Insulin Resistance & Atherosclerosis (IRAS) study].[23] A rise in inflammatory markers (CRP, fibrinogen and white cell count) in conjunction with insulin resistance was demonstrated in the IRAS study. Similarly Ridker et al.[24] have shown a strong association between elevated CRP and atherosclerosis and also elevation in CRP and interleukin (IL)-6 levels being predictive of development of diabetes.[25] Clapp et al.[26] showed that inflammation can cause endothelial dysfunction, reduce vascular NO bioavailability and increase oxidative stress, which may alternatively suggest inflammation as the initiating factor. In this study, basal and stimulated endothelial NO bioavailability were measured, while systemic effects were determined by measuring cytokine response, total antioxidant status and urinary protein excretion.
The Importance of Nuclear Factor-百B
Nuclear factor-百B (NF-百B) is an inducible eukaryotic transcription factor of the rel family, which is critical in regulating transcription of specific genes, most of which, are involved in the immune, inflammatory responses and infection and induction of various agents including IL-1 and IL-6, and adhesion molecules. Research has shown an increase in NF-百B activation leads to a proinflammatory state (27) giving rise to suggestions that a decrease in this parameter might positively affect the pathophysiology of the metabolic syndrome. Examples of this are borne out by the effect of antioxidant and anti-inflammatory properties of glitazones,[28] while high-dose aspirin has also been said to have a similar effect on glucose metabolism in type 2 diabetes.[29] NF-百B acts as a transcription factor for various proteins including VCAM-1 and ELAM-1, which are responsible for monocyte adhesion to endothelium[30] - an initial trigger in the formation of atheroma.
Ochnoflavone, a naturally occurring bioflavonoid is thought to inhibit NF-百B due to its inhibition of lipopolysaccharide-induced NO formation - which may be the basis for its anti-inflammatory effects. Recent research has also suggested flavonoid-rich products such as apple extracts[32] and green tea[33] to downregulate NK-百B signalling, thereby possibly reducing the pro-inflammatory state.
Flavonoids
In 1992, examination of WHO epidemiological data showed an intriguing anomaly in Toulouse, France where subjects, in spite of high saturated fat consumption, comparable cholesterol and similar risk factors showed considerably lower incidence of death from coronary heart disease compared with other countries such as the United States and the United Kingdom. This apparent discrepancy, also known as the 'French Paradox',[34] triggered a specific scrutiny for an explanation of this phenomenon. Review of the epidemiological data suggested that alcohol consumption, especially red wine, may have conferred superior protection compared with other beverages. This suggested that the beneficial effects of red wine were, at least in part, due to components other than alcohol. Although the specific mechanism behind the French paradox has not been identified, further studies, both in vivo and in vitro, showed flavonoid components in red wine to have antioxidant properties possibly contributing to cardiovascular benefits.[35-38]
While some studies suggest that flavonoid intake was not associated with reduced CHD, two other prospective trials suggested a lower risk of MIs. A total of eight cohort studies found lower CHD mortality with total or specific flavonoid intake[40,44-49] but one large cohort study of 38,445 women found a non- significant inverse association between flavonoid intake and CHD mortality.[41] However, a recent meta-analysis indicate a significant protective association between flavonoid intake and risk of CHD mortality, RR = 0.81 (CI 0.71-0.92).[50] One of the authors of the studies[51] that did not show any association described a high background consumption of milk consumption that might have contributed to the null finding, as milk intake can potentially prevent intestinal absorption of flavonoids.[52] Interestingly, most studies, apart from one[53] showed no association for stroke risks,[46,47] although it is likely that these studies did not have sufficient power to study strokes or indeed their various subtypes.
Chemistry of Flavonoids
Revival of interest in traditional medicine coupled with rapid advances in pharmacognosy has led to a greater understanding of the biochemistry and pharmacology of flavonoids.Flavonoids are a subgroup of a class of compounds known as polyphenols and are derivatives of 2-phenyl-1-benzopyran-4-one with varied chemical structures present in fruits, vegetables, nuts and seeds. They are polyphenolic compounds possessing 15 carbon atoms; two benzene rings joined by a linear 3-carbon chain (Figure 1).
Over 4000 flavonoids have been identified and they are divided into several groups according to their chemical structure. The six major subgroups are: chalcones, flavonols (quercetin and kaempherol), flavanone (the catechins), flavones (apigenin), anthocyanins and isoflavonoids (genistein).
Flavonoids, Oxidative Stress and Inflammation
Flavonoid intake has been inversely related with coronary heart disease in the Zutphen Elderly Study, the Seven Countries Study and a cohort study in Finland(Figure 2). Subsequent research has shown the flavonoid component in wine to inhibit oxidation of human LDL[35] and also inhibit platelet aggregation and adhesion. Cishek et al. showed red wine to cause endothelium-dependent relaxation (EDR).[56] Subsequent experiments indicated that flavonoid monomers failed to show this effect. However, oligomeric procyanidins produced a dose-dependent EDR.
Pathophysiological factors involved in the metabolic syndrome, atherosclerosis and type 2 diabetes and the potential action of flavonoid-rich grapeseed extracts (GSE).
Quercetin, a flavonoid prominent in onions and apples, has been epidemiologically linked with protection from coronary artery disease and cancer. It has also been shown to inhibit monocyte adhesion to endothelial cells,[57] which is believed to be the first step in the process of atherosclerosis.
Chocolate derived from the plant Theobroma cacao, rich in flavonoids, have shown improved endothelium-dependent flow-mediated dilation (FMD).Flavonoids in elderberry have been shown to confer significant protective effects against oxidative insult and thus have important implications upon preserving endothelial cell function and thereby preventing the initiation of endothelial cells associated with vascular disease.[59] In vitro studies have shown flavonoid-rich chocolates to inhibit lipoxygenase pathways, which give rise to proinflammatory leukotrienes.[60,61] In addition to this, some have shown chocolate procyanidins can modulate a variety of other cytokines such as IL-5, tumour necrosis factor-alpha, tumour growth factor-beta - reducing their inflammatory effects.[62-65] A recent review by Yoon and Back elucidates the anti-inflammatory effects of polyphenols highlighting the potential role for dietary polyphenols to confer health benefits in reducing inflammation.[66]
'Mechanism of Action' of Flavonoids
The mechanism underlying the effects of flavonoids/procyandins on the endothelium has yet to be defined. One of the suggested mechanisms of flavonoid action has been that it creates a 'pseudo-laminar shear stress response', which counters the endothelial dysfunction.[67]
Laminar shear stress (LSS) is the frictional force generated by blood flowing over the endothelium and in the context of normal endothelial function, this is one of the important regulatory factors as it induces vasodilation through NO and prostacyclin synthesis while inhibiting vasoconstriction by suppressing endothelin-1 production.[68,69]
LSS also has been shown to alter expression of a spectrum of genes. Gene array studies have demonstrated that physiological levels of LSS suppress the mRNA levels of genes associated with vascular dysfunction and increase the expression of protective genes.[70,71]
Grapes and Grapeseed Extract:
The medicinal and nutritional value of grapes (Vitis vinifera) has been heralded for thousands of years. Egyptians consumed this fruit at least 6000 years ago, and several ancient Greek philosophers praised the healing power of grapes - usually in the form of wine. European folk healers developed an ointment from the sap of grapevines to cure skin and eye diseases. Unripe grapes were used to treat sore throats and dried grapes (raisins) were used to heal consumption, constipation, and thirst. The ripe, sweet grapes were used to treat a range of health problems including cancer, cholera, smallpox, nausea, eye infections, and skin, kidney, and liver diseases.
A general composition of the grape ( Table 1 ) consists of 2-6% stems, 5-12% skins, 80-90% juice and 0-5% seeds. Chemically, one of the important constituents are phenolic substances (frequently called polyphenols) which embrace many classes of compounds ranging from phenolic acids to simple and complex flavonoids. Grapeseeds, although they make up a small percentage of the weight of grapes, contain two-thirds of the extractable phenols.[72] The seeds are highest in phenol content and may contain up to 5-8% phenols by weight[73] and are essentially all flavonoids. They are also referred to as monomeric flavan-3-ols, which when joined together is known as oligomeric procyanidins. The procyanidins have been the subject of intensive research - mostly looking at its antioxidant role and its effect on the vascular endothelium.
The rich presence of flavonoid components in grape juice/grapeseed extract (GSE) further prompted research in cardiovascular disease ( Table 2 ). Initially, reduced platelet aggregation was noted with GSE in canine coronary arteries.[33] Suppression of lipid peroxidation has also been shown in neonatal rats. Rejuvenation of antioxidant system in central nervous system of rats, by GSE, has also been shown.
Further research showed reduced LDL oxidation in human high cardiovascular risk patients reduced human platelet aggregation, enhanced NO release and improved human endothelial function.Post-prandial oxidative stress was also seen to be reduced in models of oxidative stress (smoking).[77] Flow-mediated vasodilation has also seen to be improved after GSE administration.[78] In this context, it is important to remember that although there is no existing data relating impaired FMD to cardiac events in subjects without coronary disease, those patients with coronary disease who have very impaired FMD have more events.[79] Subjects with impaired FMD are also more likely to have coronary disease on angiography.[80] Statins improve mortality and one mechanism may be via their improvement of FMD.[81]
The University of California demonstrated that constituents present in GSE relaxed isolated blood vessels from rabbits by a pathway in which NO production is implicated, while flavonoid-rich extract from GSE showed attenuation of development of aortic atherosclerosis in cholesterol-fed rabbits.[83]
Endothelium-dependent vasorelaxing activity in an aortic ring model is increased by incubation with grape products and these changes appear to be mediated by the NO-cGMP pathway. Interestingly, in vitro tests using a cupric-ion-mediated LDL + VLDL oxidation model showed a synergistic effect of GSE with both Vitamin C and Vitamin E. Research by Peng et al. also indicated a decrease in arterial pressure in spontaneously hypertensive rats - probably via an antioxidant mechanism.[85]
Grapeseed extract has also been shown to be a more potent scavenger of oxygen-free radicals (an oxidative stressor) than other common antioxidants such as Vitamin C and E. Uses of grapeseed-derived procyanidins (flavonoids with oligomeric structure) have also been shown to have an anti-hyperglycaemic effect on streptozotocin-induced diabetic rats. This was significantly increased if accompanied by a low insulin dose, which may partially have been due to the insulinomimetic activity of procyanidins on insulin-sensitive cell lines.[88] Research from
Vinson et al. in 2001 used GSE in 17 subjects (nine normal and eight hypercholesterolaemic). Lipid profile and plasma antioxidant capacity were seen to be improved in the high-cholesterol subjects without a concomitant improvement in the healthy individuals.[89] Recent experimental trials have also suggested that proanthocyanidin-rich extract has protective effects against ischaemia-reperfusion-induced renal damage associated with oxidative stress.[90]
Other potential benefits of GSE have been centred on its effect on neoplasia and there have been several encouraging trials undertaken suggesting a beneficial effect.Tebib et al.reported an interesting finding in 1994 whereby faecal excretion of cholesterol by rats fed GSE was approximately twofold higher compared with the control group and could be attributed to a reduction or inhibition of intestinal cholesterol absorption.
Side-effects and Safety Profile of Grapeseed Extract
It is important to keep in mind with any potential treatment the possible side effects it brings with it. Fortunately, in vivo studies have not reported any major side effects and there are no known scientific reports of interactions between grapeseed and conventional medications either.
Grapeseed extract has been used as a dietary supplement for a number of years - both in Europe and the USA, and at present holds GRAS (Generally regarded as safe) status - as assessed by the FDA. 'Recommended dosage' has varied from 100 to 300 mg/day while pregnant and breast-feeding women are advised to avoid GSE.
Conclusion
Cardiovascular disease is one of the most important public health issues in the modern world - accountable for a sizeable portion of morbidity and mortality. An increasing population coupled with reduced exercise levels and dietary indiscretions have combined to increase the development of this pathology many fold. The recent past has seen an enormous advancement in the understanding of mechanisms underlying this condition. The links between oxidative stress, metabolic syndrome and inflammation are being increasingly unravelled and there is a great interest in therapies that influence the function of endothelial cells, which are key regulatory cells in the vessel wall. The identification of naturally occurring flavonoids that can modulate the production of NO in body cells is an exciting prospect.
The antioxidant and vascular protective aspects of flavonoid-rich products such as GSE, when combined with the potential hypolipidaemic and anti-platelet effects make a strong case for its potential in preventing and treating diseases associated with endothelial injury, oxidative damage and inflammation; chief among which are type 2 DM and atherosclerotic vascular disease. GSE are also an attractive proposition due to their wide availability and safety profile.
To date, there have been no studies examining the effects of GSE upon novel cardiovascular risk factors in high-risk patient groups. Given this observation and the emerging evidence that GSE may have an effect upon reducing oxidative stress, the use of GSE in such patients may demonstrate concomitant improvements in insulin resistance, endothelial function, inflammation in high-risk patient groups and ultimately cardiovascular outcome.
Reprint Address
Dr Partha Kar, 8, Winstanley Road, Nursling, Southampton SO16, 0TF, UK, Tel.: 07738173099. Email: partha.kar@porthosp.nhs.uk
Topical Vitamins, Minerals and Botanical Ingredients As Modulators of Environmental and Chronological Skin Damage:A. Chiu, A.B. Kimball, Department of Dermatology, Stanford University School of Medicine, Stanford, CA
Abstract and Introduction
Abstract
Ageing skin is characterized by fine lines, wrinkles, lentigines, dyspigmentation and increased coarseness. Topical preparations alleged to combat these changes abound in the over-the-counter market. Some of the most popular ingredients used in these products are vitamins, minerals and botanical extracts. Proposed mechanisms for antiageing effects on skin range from antioxidant properties to improved collagen synthesis or protection from collagen breakdown. Despite the media attention and consumer popularity that these ingredients have generated, there have been few scientific studies to support these claims. In this report, we review recent published studies on the most common of these ingredients for the topical photoprotection and the treatment of ageing skin.
Introduction
Environmental or exogenous factors such as ultraviolet (UV) radiation, wind and smoke contribute to the extrinsic ageing of skin. This type of ageing, combined with intrinsic, or chronological ageing, results in the degeneration of the skin barrier, the development of rhytides, discoloration and possible malignant degeneration, among other changes.[1,2] Cosmetic changes associated with ageing, especially on the face, are particularly concerning to a patient population which wishes to remain looking youthful. As the demand for products that reduce the cosmetic effects of ageing continues to grow, healthcare professionals have a responsibility to educate themselves and to become informed about the scientific basis and established data, if any, behind these products. Clearly, topical, over-the-counter products alleged to benefit ageing skin are immensely popular among patients. These products account for annual sales of over 2 billion dollars in the U.S.A. alone.[3] Although topical medications such as tretinoin have been demonstrated in the scientific literature to reduce the signs of ageing,[4,5] patients often seek over-the-counter antiageing products due to the market availability, comparably cheaper prices, and the lack of the physician bottleneck.
Recently, consumer and media attention has focused specifically on products utilizing 'natural' ingredients such as vitamins, minerals and botanical extracts. These ingredients have the appeal of appearing wholesome and 'organic'. Although scientific evidence shows that some of these ingredients do have possible in vitro antiageing activity, the question remains whether it is possible to deliver adequate doses to the skin in vivo. and to produce either histological or clinical improvement of wrinkles, lentigines, coarseness, pigmentary changes, dryness and other characteristics of ageing skin. This article reviews recent published studies on the most common of these 'natural ingredients' and summarizes their proposed effects on ageing skin.
Methods
The Medline electronic database was searched from 1996 to 2002 using the following keywords: vitamins, minerals, botanical extracts, tea extracts, herbal, vitamin E, alpha-tocopherol, vitamin C, ascorbic acid, retinol, retinal, nicotinamide, niacinamide, coenzyme Q10, Ginkgo, grape seed extract, lemon, lavender, ginseng, rosemary, green tea, soybean, genistein, seaweed, peppermint, algae, cucumber, aloe vera, wheat protein, witch hazel and panthenol, cross-matched with the key words ageing/aging and skin. The bibliographies of these papers were then scanned for relevant references which form the basis of this report.
Because the topic is broad, this review is limited to recent reports, as well as older studies which utilized human subjects, established background information for recent reports, or demonstrated a particularly important research finding.
Vitamin E (alpha-Tocopherol)
Vitamin E is one of the better established ingredients in over-the-counter treatments of skin ageing. Vitamin E, or 汐-tocopherol, is a lipid-soluble antioxidant which plays key roles in protecting cellular membranes from lipid peroxidation by free radicals.[6,7] These free radicals contribute significantly to the environmental, or exogenous ageing of skin, especially UV-mediated damage.[8] Although the human skin possesses various intrinsic defence systems which help to mitigate these types of oxidative damage, both excessive and chronic exposure to free radicals can deplete the body's defence. Free radicals cause the disruption of normal biomolecules such as lipids, proteins and nucleic acids, as well as deplete the body's own endogenous antioxidants.[9] Antioxidants, such as 汐-tocopherol, modulate this damage by scavenging free radicals and lipid peroxyl radicals.[10]
Interestingly, vitamin E is distributed in a gradient fashion in the stratum corneum of healthy skin, with the highest levels in the deepest layers and the lowest levels closest to the surface.In a recent review of antioxidants in the skin, Thiele et al. concluded that 汐-tocopherol is the major antioxidant in the human epidermis, and that its depletion is an early and sensitive marker of environmental oxidative damage.[11] Vitamin E is available over-the-counter in various forms, most commonly as 汐-tocopherol or tocopherol acetate. However, as with other vitamins and antioxidants applied topically, success of delivery and clinical efficacy are separate and crucial elements of an effective antiageing treatment. This was demonstrated in a human study in which the acetate form of tocopherol showed no evidence of conversion to the biologically active form, 汐-tocopherol, despite adequate absorption into the skin.[12] Further, a recent study on the metabolic conversion of 汐-tocopherol acetate into 汐-tocopherol in skin demonstrated that permeation and metabolism of 汐-tocopherol acetate was highly dependent on the delivery system, re-emphasizing the importance of formulation in cosmetic preparations.
In a rare hemiface trial with human subjects, 5% vitamin E was better than a vehicle control in reducing rhytides, skin roughness, length of facial lines and depth of wrinkles as measured by optical profilometry.Gehring et al. demonstrated that the topical application of vitamin E increased stratum corneum hydration and enhanced water-binding capacity.[7] In various in vitro experiments by Ricciarelli et al. 汐-tocopherol reduced the age-dependent increase of collagenase expression by inhibiting protein kinase C activity.[15]
The protective effects of vitamin E against photoageing have been demonstrated in various animal and in vitro skin models. In hairless mice, topical vitamin E has been shown to decrease the severity of UV-induced skin wrinkling by 75% and significantly to reduce erythema and oedema.[16,17] Similar photoprotective effects were evident in experiments on pig skin.[18] A study using a topical vitamin E analogue, Trolox, showed inhibition of UVB-induced intracellular peroxide generation in human keratinocytes.[19] However, results from various studies conflict over whether vitamin E applied after UV exposure can mitigate UV-induced erythema and oedema. Although some animal studies support the benefit of vitamin E by diminishing UV-related responses of the skin, a recent trial using human subjects failed to demonstrate benefits when vitamin E was applied after UV exposure.[20] Hence, although vitamin E has thus far yielded promising results for the treatment of both intrinsic and extrinsic ageing, very few of these studies have used human subjects.
Vitamin C
Vitamin C has gained mass popularity in the last few years as an over-the-counter ingredient for the treatment of photoageing. l-ascorbic acid, the biologically active form, has well-established roles in the human body as an antioxidant and as a cofactor for collagen synthesis.[21,22] Ascorbate participates in the hydroxylation of procollagen, and studies show that it may also stimulate collagen synthesis directly by activating its transcription and stabilizing procollagen mRNA.[21,23] An in vitro study showed collagen biosynthesis to be inversely related to the age of donor human fibroblasts, but that stimulation of collagen synthesis by ascorbic acid was independent of fibroblast age.[24] As an antioxidant, ascorbic acid is the body's main water-soluble, nonenzymatic electron scavenger, enabling it to function efficiently in aqueous compartments.[22] In addition to these roles in the maintenance of skin health, ascorbic acid also helps to regenerate the oxidized forms of 汐-tocopherol.[25,26] These properties of ascorbic acid are the basis for the antiageing claims made by the skincare industry. However, because it is water-soluble, topical preparations of vitamin C have been difficult to stabilize in solution and to formulate in a manner that adequately penetrates the skin.[22]
In one of the very few published reports on the effects of topical 'natural' skincare products in a placebo-controlled clinical trial, Traikovich found that topical ascorbic acid in the form of Cellex-C applied for 3 months showed a 73.7% improvement in optical profilometry image analysis.[27] This study compared a randomized hemiface application of ascorbic acid with a vehicle control, and demonstrated significant improvements in clinical, subjective and photographic assessments of wrinkling, roughness, rhytides, laxity and sallowness.
The physiological mechanisms affected by vitamin C are likely to include the increased production of collagen as well as the decreased production of matrix metalloproteinase, an enzyme which enhances dermal collagen degradation.[28] In a randomized, blinded human study, 5% vitamin C was applied to one forearm, while placebo was applied to the other for 6 months. Skin biopsies showed increased mRNA levels of collagen I, collagen III and tissue inhibitor of matrix metalloproteinase 1 on the side treated with vitamin C. Interestingly, the results were most remarkable in subjects with lower dietary intake of the vitamin. However, in this same study, mRNA levels of elastin, fibrillin and tissue inhibitor of matrix metalloproteinase 2 were not significantly affected.[29]
Alster and West propose that ascorbic acid may also have some anti-inflammatory effects, as suggested by its ability to reduce post-CO2 laser-induced erythema, a clinical indicator of dermal inflammation caused by laser injury.[30] Further, its role as an antioxidant gives ascorbate similar photoprotecting properties as vitamin E by neutralizing UVB-generated free radicals. Darr et al. demonstrated in pigs the reduction of both UVB-induced erythema and histological sunburn cells by pretreatment with 10% ascorbic acid.[31] Ascorbate may also have similar protective effects against UVA, as the investigators also showed a reduction of sunburn cells in UVA-exposed porcine skin presensitized with psoralen.[31] Various other models using human skin have demonstrated similar effects either with ascorbate alone, or in combination with vitamin E.[32,33] These results have been also corroborated in a small study with 10 human subjects.[34]
In the hairless mouse, both topical vitamin E and vitamin C reduced UVB-induced skin wrinkling. However, UVA-induced sagging was unaffected.Beyond its well-known antioxidant effects, Catani et al. suggest that vitamin C may also have cytoprotective effects by inhibiting activator protein-1, a gene transcription activator induced by UV irradiation which results in keratinocyte cell death.[35] Further, Takashima et al. found that vitamin C as ascorbyl phosphate may have some efficacy in the lightening of skin dyspigmentation.[36]
Studies show that vitamin C clearly has numerous roles in the maintenance of skin health, and may prove to be an important mediator of cutaneous ageing. However, inconsistencies in product stability and difficulties in the adequate penetration of skin limit the topical efficacy of many, if not most, of the skincare products containing vitamin C. The most common forms of vitamin C found in cosmetic products are l-ascorbic acid, and its ester form ascorbyl palmitate. Topically applied l-ascorbic acid increased ascorbate levels in pig skin up to 25 times.[31] However, ascorbic acid has been notoriously difficult to stabilize, as it rapidly oxidizes in solution. More recently, stable and hydrophobic solutions of ascorbic acid have been developed consisting of the molecule in its nonionized form at a low pH.[37] Ascorbyl palmitate, which is the fat-soluble form of ascorbic acid, is not as well established in its stability and penetration, but was found to be 30 times more effective than ascorbic acid when acting as a tumour inhibitor in mouse skin.
In summary, although vitamin C appears to be a promising cosmetic ingredient, larger studies with human subjects need to be done before the role of vitamin C in skin ageing can be firmly established, and a stable, skin-penetrating compound can be documented.
Vitamin A As Retinol
Retinoids function in the normal maintenance of epidermal differentiation and growth.Various retinoid receptors, when bound with retinoid ligands, act as important transcription factors in the human cell.[41] Deficiency of dietary vitamin A results in generalized xerosis, hyperkeratosis and squamous metaplasia of mucous membranes.[42] In the treatment of photoageing, numerous large-scale, double-blinded, placebo-controlled trials have shown the efficacy of topical tretinoin.[4,43,44] However, as tretinoin remains a prescription drug, over-the-counter products mainly utilize the vitamin A derivatives retinol and retinyl palmitate. These forms are not biologically active until enzymatic conversion to the principal active metabolite, retinoic acid.[45] Whether the skin has adequate levels of these conversion enzymes to make a clinically apparent difference has yet to be proven. Many of the studies discussed here are in vitro and do not address this issue of possible enzyme saturation directly. There are surprisingly few published studies focused on the over-the-counter retinoids, considering their widespread use. Although it is well established that UV irradiation decreases both retinol and retinyl ester levels in the epidermis, whether these decreases can be overcome by topical application is not understood.[46] Further, how the photoinactivation of vitamin A in human skin affects the topical supplementation of vitamin A is also not well addressed in the current literature.[47]
Researchers suggest that cutaneous ageing results from the interplay of extrinsic damage by UV radiation, intrinsic increases in collagen-degrading matrix metalloproteinases, and decreased collagen synthesis.[48] It is believed that retinol plays important roles in countering these mechanisms.[49,50] A study utilizing human skin samples found that topical 1% retinol inhibits the increase in metalloproteinases and stimulates collagen synthesis in both aged, sun-protected skin and in photoaged skin. Varani et al. also used tissue specimens to show that retinol may be able to restimulate fibroblast growth potential, which seems to decrease with increasing age.[51] Further, Kang et al. showed that topical retinol does increase epidermal thickness in human skin, but it was 20 times less potent than topical retinoic acid.[52] Evidence for the effectiveness and biological activity of the ester form of vitamin A, retinyl palmitate, is sparse, and no studies address clinically relevant findings in skin ageing. Further, in mice, retinol stores in the epidermis are partially UVB resistant, while retinyl esters were not.
In an open study comparing the efficacy of vitamin E cream and 0.075% retinol, 14 volunteers followed a complex regimen involving the application of a vitamin E cream for a month, followed by retinol cream for 2 months, followed by vitamin E cream for another month. Samples of the stratum corneum during retinol treatment were shown to be less sensitive to sodium lauryl sulphate by corneosurfometry bioassay. UV squamometry tests and optical profilometry of UV-induced shallow wrinkling also demonstrated retinol to be more effective than vitamin E.[53]
Although some studies suggest the possible efficacy of retinol in improving the cosmetic appearance of ageing skin, over-the-counter products usually contain lower levels of retinol than used in these studies. Ultimately, because of variable retinol concentrations in cosmetic products and the lack of blinded clinical trials, the results of these studies should be interpreted carefully.
Other Vitamin-Based Ingredients (Nicotinamide and Coenzyme Q10)
Among the newer vitamin-based ingredients promoted by the cosmetic industry are nicotinamide (niacinamide) and coenzyme Q10 (ubiquinone). Studies on nicotinamide, a derivative of niacin, have mainly focused on its anti-inflammatory and antiacne vulgaris actions.[54,55] Researchers believe anti-inflammatory effects may ultimately improve skin appearance by reducing leucocyte peroxidase systems that may lead to localized tissue damage. Nicotinamide is a B vitamin and an integral part of the coenzymes nicotinamide adenine dinucleotide (NAD) and NAD phosphate. In a double-blinded study of 76 patients with acne vulgaris, 4% nicotinamide gel and 1% clindamycin gel were comparable in efficacy.[54] Tanno et al. showed that nicotinamide might have some utility in the treatment of chronological ageing by decreasing transepidermal water loss.[56] Further, sphingolipids, such as ceramides, are a major component of the stratum corneum,[57] and this in vitro experiment also preliminarily suggests that nicotinamide increases the activity of the rate-limiting enzyme involved in sphingolipid synthesis, as well as the mRNA levels of sphingolipids. When the investigators incubated keratinocytes with nicotinamide, increased levels of free fatty acids, cholesterol and ceramides were found in the stratum corneum.[56]
Published studies focusing on the possible antiageing effects of the mitochondrial electron transfer protein coenzyme Q10 are rare. Coenzyme Q10, otherwise known as ubiquinone, is an endogenous cellular antioxidant present in almost all the tissues of the body, including the skin.[10,25] It is believed that ubiquinone may have roles in both the extrinsic and chronological processes of ageing. Various authors have reported that the tissue levels of coenzyme Q10 decrease with age.[58,59] Hoppe et al. demonstrated this age-related reduction specifically in
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