The Effect of Grape Seed Extract and Vitamin C Feed Supplements Carcass Characteristics, Gut Morphology and Ileal Microflora in Broiler Chickens Exposed to Chronic Heat Stress

Document Type: Research Article


Department of Animal Science, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran


The effects of dietary supplementation of grape seed extract (GSE) and vitamin C on carcass characteristics, gut morphology and ileal microflora in broiler chickens exposed to chronic heat stress were examined. Experimental diets included a control diet (without additive), 3 levels of GSE (150, 300, 450 mg/kg of diet), and vitamin C (300 mg/kg of diet). Each of the five diets was fed to 5 replicates of 12 male Cobb-500 chicks, from 1 to 42 d of age. The birds were exposed to chronic daily heat stress of 34 ± 1 ˚C with 65-70% relative humidity for 5 hours per day from 29 to 42 d of age. Addition of GSE or vitamin C to the basal diet did not affect the percentage by weight of the edible carcass, breast, drumstick + thighs, liver, empty gizzard, or abdominal fat. Furthermore, in heat-exposed broiler chickens the relative length of duodenum, jejunum and ileum did not show significant alteration when compared to the control group before and after chronic heat stress. Dietary GSE and vitamin C caused differences in jejunum muscle layer thickness, villus height, villus width and crypt depth of birds before heat stress. Addition of the GSE or vitamin C did not affect the jejunum muscle layer thickness, villus width, crypt depth and villus: crypt ratio of broilers under heat stress (42 d). However, broilers fed a diet containing GSE had higher villus compared to the control group at 42 d (P<0.05). Adding GSE or vitamin C reduced ileal coliforms and the Escherichia coli population of broilers before heat stress condition (28 d). Under chronic heat stress conditions, broilers fed diet supplemented with GSE had lower ileal coliforms and Escherichia coli populationscompared to control birds (P<0.05). Thus, GSE can be recommended as a new herbal extract supplement to the diet of broiler chickens in order to improve jejunum villus height and decrease detrimental bacteria in the ileum of birds before and during chronic heat stress condition.


Ajakaiye J.J., Perez-Bello A. and Mollineda-Trujillo A. (2011). Impact of heat stress on egg quality in layer hens supplemented with l-ascorbic acid and dl-tocopherol acetate. Vet. Arhiv. 81, 119-132.
Akbarian A., Golian A., Kermanshahi H., Farhoosh R., Raji A.R., De Smet S. and Michiels J. (2013). Growth performance and gut health parameters of finishing broilers supplemented with plant extracts and exposed to daily increased temperature. Spanian J. Agric. Res. 11, 109-119.
Akyurek H. and Yel A. (2011). Influence of dietary thymol and carvacrol preparation and / or an organic acid blend on growth performance, digestive organs and intestinal microbiota of broiler chickens. African J. Microbiol. Res. 5, 979-984.
AOAC. (1990). Official Methods of Analysis. Vol. I. 15th Ed. Association of Official Analytical Chemists, Arlington, VA.
Arora A., Nair M.G. and Strasburg G.M. (1998). Structure-activity relationships for antioxidant activities of a series of flavonoids in a liposomal system. Free Radical Biol. Med. 24, 1355-1363.
Benzie I.F.F. and Strain J.J. (1999). Ferric reducing / antioxidant power assay: direct measure of total antioxidant activity of biological fuids and modifed version for simultaneous measurement of total antioxidant power and ascorbic acid concentration. Meth. Enzymol. 299, 15-27.
Brenes A., Viveros A., Goñi I., Centeno C., Saura-Calixto F. and Arija I. (2010). Effect of grape seed extract on growth performance, protein and polyphenol digestibilities and antioxidant activity in chickens.Spanian J. Agric. Res. 8, 326-333.
Brenes A., Viveros A., Goni I., Centeno C., Sayago-Ayerdi S.G., Arija I. and Saura-Calixto F. (2008). Effect of grape pomace concentrate and vitamin E on digestibility of polyphenols and antioxidant activity in chickens. Poult. Sci. 87, 307-316.
Brisbin J.T., Gong J. and Sharif S. (2008). Interactions between commensal bacteria and the gut-associated immune system of the chicken. Anim. Health Res. Rev. 9, 101-110.
Burkey T.E., Skjolaas A. and Minton J.E. (2009). Broad-invited review: porcine mucosal immunity of the gasterointestinal tract. J. Anim. Sci. 87, 1493- 1501.
Burkholder K.M., Thompson K.L., Einstein M.E., Applegate T.J. and Patterson J.A. (2008). Influence of stressors on normal intestinal microbiota, intestinal morphology and susceptibility to Salmonella enteritidis colonization in broilers. Poult. Sci. 87, 1734-1741.
Chamorro S., Viveros A., Centeno C., Romero C., Arija I. and Brenes A. (2013). Effects of dietary grape seed extract on growth performance, amino acid digestibility and plasma lipids and mineral content in broiler chicks. Animal. 7, 555-561.
Chedea V.S., Echim C., Braicu C., Andjelkovic M., Verhe R. and Socaciu C. (2011). Composition in polyphenols and stability of the aqueous grape seed extract from the Romanian variety “Merlot Recas”. J. Food Biochem. 35, 92-108.
CLSI. (2006). Clinical and Laboratory Standard Institute. Performance Standards for Antimicrobial Disk Susceptibility Tests; Approved Standards. Clinical and Laboratory Inst., 940, West Valley Road, Suite 1400, Wayne, Pa., 19087–1898. USA.
Cobb-Vantress. (2012). Cobb 500 TM: Broiler Performance and Nutrition Supplement Guide. Cobb-Vantress, Siloam Springs. AR.
Cox S.D., Mann C.M. and Markham J.L. (2001). Interactions between components of the essential oil of Melaleuca alternifolia. J. Appl. Microbiol. 91, 492-4977.
Cross D.E., Mcdevitt R.M., Hillman K. and Acamovic T. (2007). The effect of herbs and their associated essential oils on performance, dietary digestibility and gut microflora in chickens from 7 to 28 days of age. Br. Poult. Sci. 48, 496-506.
Deprez S., Brezielon C., Rabot S., Philippe C., Mila I., Lapierre C. and Scalbert A. (2000). Polymeric anthocyanidins are catabolized by human colonic microflora into low-molecular-weight phenolic acids. J. Nutr. 130, 2733-2738.
Duncan D.B. (1955). Multiple range and multiple F-test. Biometrics. 11, 1-42.
Etches R.J., John T.M. and Verrinder Gibbins A.M. (2008). Behavioural, physiological, neuroendocrine and molecular responses to heat stress. Pp. 49-69 in Poultry Production in Hot Climates. N.J. Daghir, Ed. Trowbridge. Cromwell Press.
Furiga A., Lonvaud-Funel A. and Badet C. (2008). In vitro study of antioxidant capacity and antibacterial activity on oral anaerobes of a grape seed extract. Food Chem. 113, 1037-1040.
García-Ruíz A., Bartolomé B., Martínez-Rodríguez A.J., Puello E., Martín-Álvarez P.J. and Moreno-Arribas M.V. (2008). Potential of phenolic compounds for controlling lactic acid bacteria growth in wine. Food Control. 19, 835-841.
Garriga C., Hunter R.R., Amat C., Planas J.M., Mitchell M.A. and Moretó M. (2006). Heat stress increases apical glucose transport in the chicken jejunum. Am. J. Physiol. Regul. Integr. Comp. Physiol. 290, 195-201.
Goni I., Brenes A., Centeno C., Viveros A., Saura-Calixto F., Rebole A., Arija I. and Estevez R. (2007). Effect of dietary grape pomace and vitamin E on growth performance, nutrient digestibility and susceptibility to meat lipid oxidation in chickens. Poult. Sci. 47, 581-591.
Goni I. and Serrano J. (2005). The intake of dietary fiber from grape seeds modifies the antioxidant status in rat scum. J. Sci. Food Agric. 85, 1877-1881.
Guban J., Korver D.R., Allison G.E. and Tannock G.W. (2006). Relationship of dietary antimicrobial drug administration with broiler performance decreased population levels of Lactobacillus salivarius and reduced bile salt deconjugation in the ileum of broiler chickens. Poult. Sci. 85, 2186-2194.
Guo F.C., Kwakkel R.P., Williams B.A., Li W.K., Li H.S., Luo J.Y., Li X.P., Wei Y.X., Yan Z.T. and Verstegen M.W. (2004a). Effects of mushroom and herb polysaccharides, as alternatives for anantibiotic, on growth performance of broilers.  Br. Poult. Sci. 45, 684-694.
GuoF.C., Williams B.A., Kwakkel R.P., Li H.S., Li X.P., Luo J.Y., Li W.K. and Verstegen M.W. (2004b). Effects of mushroom and herb polysaccharides, as alternatives foran antibiotic, on the caecal microbial ecosystem in broiler chickens. Poult. Sci. 83, 175-182.
Guo F.C., Kwakkel R.P., Williams B.A., Parmentier H.K., Li W.K., Yang Z.Q. and Verstegen M.W.A. (2004c). Effects of mushroom and herb polysaccharides on cellular and humoral immune responses of Eimeriatenella-infected chickens. Poult. Sci. 83, 1124-1132.
Hashimoto T., Kumazawa S., Nanjo F., Hara Y. and Nakayama T. (1999). Interaction of tea catechins with lipid bilayers investigated with liposome systems. Biosci. Biotechnol. Biochem. 63, 2252-2255.
Iacopini P., Baldi M., Storchic P. and Sebastiani L.(2008). Catechin, epicatechin, quercetin, rutin and resveratrol in red grape: content, in vitro antioxidant activity and interactions.J. Food Comp. Anal. 21, 589-598.
Ignat I., Radu D.G., Volf I., Pag A.I. and Popa V.I. (2013). Antioxidant and antibacterial activities of some natural polyphenols. Cellul. Chem. Technol. 47, 387-399.
Jacob J.K., Hakimuddin F., Paliyath G. and Fisher H. (2008). Antioxidant and antiproliferative activity of polyphenols in novel high-polyphenol grape lines. Food Res. Int. 41, 419-428.
Jamroz D. and Kamel C. (2002). Plant extracts enhance broiler performance. In non-ruminant nutrition: antimicrobial agents and plant extracts on immunity, health and performance. J. Anim. Sci. 80, 41-46.
Juven B.J., Kanner J., Schved F. and Weisslowicz H. (1994). Factors that interact with the antibacterial action of thyme essential oil and its active constituents. J. Appl. Bacteriol. 76, 626-631.
Khaksar V., Golian A. and Kermanshahi H. (2012).Immune response and ileal microflora in broilers fed wheat-based diet with or without enzyme Endofeed W and supplementation of thyme essential oil or probiotic PrimaLac®.African J. Biotechnol. 11, 14716-14723.
Lambert G.P. (2009). Stress-induced gastrointestinal barrier dysfunctions and its inflammatory effects. J. Anim. Sci. 87, 101-108.
Lara L.J. and Rostagno M.H. (2013). Impact of heat stress on poultry production.Animal. 3, 356-369.
Laudadio V., Dambrosio A., Normanno G., Khan R.U., Naz S., Rowghani E. and Tufarelli V. (2012). Effect of reducing dietary protein level on performance responses and some microbiological aspects of broiler chickens under summer environmental conditions. Avian Biol. Res. 5, 88-92.
Laurent C., Besancon P. and Caporiccio B. (2005). 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. 53, 5541-5548.
Leon L.R. (2007). Heat stroke and cytokines. Prog. Brain. Res. 162, 481-524.
Majo D.D., Guardia M.L., Giammanco S., Neve L.L. and Giammanco M. (2008). The antioxidant capacity of red wine in relationship with its polyphenolic constituents. Food Chem. 111, 45-49.
Mason T.L. and Wasserman B.P. (1987). Inactivation of red beet betaglucan synthase by native and oxidized phenolic compounds. Phytochemical. 26, 2197-2202.
Mountzouris K.C., Paraskevas V., Tsirtsikos P., Palamidi I., Steiner T., Schatzmayr G. and Fegeros K. (2011). Assessment of a phytogenic feed additive effect on broiler growth performance, nutrient digestibility and caecal microflora composition. Anim. Feed Sci. Technol. 168, 223-231.
Nyamambi B., Ndlovu L.R., Naik Y.S. and Kock N.D. (2007). Intestinal growth and function of broiler chicks fed shorgum based diets differing in condensed tannin levels. South African J. Anim. Sci. 37, 202-214.
Osawa R., Kuroiso K., Goto S. and Shimizu A. (2000). Isolation of tannin-degrading lactobacilli from humans and fermented foods. Appl. Environ. Microbiol. 66, 3093-3097.
Pastrana-Bonilla E., Akoh C.C., Sellappan S. and Krewer G. (2003). Phenolic content and antioxidant capacity of muscadine grapes. J. Agric. Food. Chem. 51, 4497-4503.
Pearce S. (2011). The effects of heat stress and nutritional status on metabolism and intestinal integrity in growing pigs. MS Thesis. Iowa State Univ., USA.
Perumalla A.V.S. and Hettiarachchy N.S. (2011). Green tea and grape seed extracts-potential applications in food safety and quality. Food Res. Int. 44, 827-839.
Quinteiro-Filho W.M., Gomes A.V., Pinheiro M.L., Ribeiro A., Ferraz-de-Paula V., Astolfi-Ferreira C.S., Ferreira A.J. and Palermo-Neto J. (2012). Heat stress impairs performance and induces intestinal inflammation in broiler chickens infected with Salmonella enteritidis. Avian Pathol. 41, 421-427.
Quinteiro-Filho W.M., Ribeiro A., Ferraz-de-Paula V., Pinheiro M.L., Sakai M., Sa L.R., Ferreira A.J. and Palermo-Neto J. (2010). Heat stress impairs performance parameters, induces intestinal injury and decreases macrophage activity in broiler chickens. Poult. Sci. 89, 1905-1914.
Re R., Pellegrini N., Proteggente A., Pannala A., Yang M. and Rice-Evans C. (1999). Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic. Biol. Med. 26, 1231-1237.
Sahin K., Sahin N., Kucuk O., Hayirli A. and Prasad A.S. (2009). Role of dietary zinc in heat-stressed poultry: a review. Poult. Sci. 88, 2176-2183.
Salari A., Habib Najafi M.B., Farhoosh R. and Marashi S.H. (2009). Survey of solvents extraction of grape seed extracts and assay of antimicrobial properties. Iranian Food Sci. Technol. Res.J. 4, 71-79.
Sallam K.I. (2007). Antimicrobial and antioxidant effects of sodium acetate, sodium lactate and sodium citrate in refrigerated sliced salmon. Food Control. 18, 566-575.
SAS Institute. (2001). SAS®/STAT Software, Release 8.2. SAS Institute, Inc., Cary, NC.
Sehm J., Lindermayer H., Dummer C., Treutter D. and Pfaffl M.W. (2007). The influence of polyphenol rich apple pomace or red wine pomace diet on the gut morphology in weaning piglets. J. Anim. Physiol. Anim. Nutr. 91, 289-296.
Shi J., Yu J., Pohorly J.E. and Kakuda Y. (2003).Polyphenolics in grape seeds-biochemistry and functionality.J. Med Food. 6(4), 291-299.
Shimamura T., Zhao W.H. and Hu Z. (2007). Mechanism of action and potential for use of tea catechin as an anti-infective agent. Anti-infective. Agen. Med. Chem. 6, 57-62.
Sivarooban T., Hettiarachchy N.S. and Johnson M.G. (2008). Transmission electron microscopy study of Listeria monocytogenes treated with nisin in combination with either grape seed or green tea extract. J. Food Prot. 71, 2105-2109.
Spranger I., Sun B., Mateus A.M., De-Freitas V. and Ricardo-Da Silva J.M. (2008). Chemical characterization and antioxidant activities of oligomeric and polymeric procyanidin fractions from grape seeds. Food Chem. 108, 519-532.
Tamzil M.H., Noor R.R., Hardjosworo P.S., Manalu W. and Sumantri C. (2013). Acute heat stress responses of three lines of chickens with different heat shock protein (HSP)-70 genotypes. Int. J. Poult. Sci. 12, 264-272.
Tao X. and Xin H. (2003). Temperature-humidity-velocity-index for market size broilers. Trans. ASAE. 46, 491-497.
Thomas D.V., Molan A.L. and Ravindran V. (2007). Performance and digestive tract development effects of green tea supplementation in broiler starters fed wheat-based diets. Asia. Pac. J. Clin. Nutr. 16(3), 71-76.
US EPA. (2010). United States Environmental Protection Agency. Available in: www. Epa. Gov.
Viveros A., Chamorro S., Pizarro M., Arija I., Centeno C. and Brenes A. (2011). Effects of dietary polyphenol-rich grape products on intestinal microflora and gut morphology in broiler chicks. Poult. Sci. 90, 566-578.
Ward N.C., Croft K.D., Puddey I.B. and Hodgson J.M. (2004). Supplementation with grape seed polyphenols results in increased urinary excretion of 3-hydroxy-phenylpropionic acid, an important metabolite of proanthocyanidins in humans. J. Agric. Food Chem. 52, 5545-5549.
Wei S., Morrison M. and Yu Z. (2013). Bacterial census of poultry intestinal microbiome. Poult. Sci. 92, 671-683.
Xia E.Q., Deng G.F., Guo Y.J. and Li H.B. (2010). Biological activities of polyphenols from grapes: a review. Int. J. Mol. Sci. 11, 622-646.