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Jul 28, 2017 - Effect of turmeric (Curcuma longa) dietary supplementation on growth performance, immune response, and antioxidant status in broiler ...

Alexandria Journal of Veterinary Sciences AJVS. Vol. 54 (2): 19-28. July 2017 DOI: 10.5455/ajvs.272232 Immunity, Antioxidant Status, and Performance of Broiler Chickens Fed Turmeric (Curcuma Longa) Rhizome Powder Amal H. Osman1*, Ali H. El-Far1*, Kadry M. Sadek1, Ismail I. Abo-Ghanema2, Mervat A. Abdel-Latif3 1Department

of Biochemistry, Faculty of Veterinary Medicine, Damanhour University, Damanhour, 22511, Egypt Department, Faculty of Veterinary Medicine, Damanhour University, Damanhour, 22511, Egypt 3Department of Nutrition and Veterinary Clinical Nutrition, Faculty of Veterinary Medicine, Damanhour University, Damanhour, 22511, Egypt 2 Physiology

ABSTRACT Key words: Broiler; Turmeric; Immunity; Antioxidant; Performance; Muscle malondialdehyde

Correspondence to: [email protected]

Effect of turmeric (Curcuma longa) dietary supplementation on growth performance, immune response, and antioxidant status in broiler chickens was evaluated by the determination of growth measurements, serum total protein, albumin, globulin, alanine aminotransferase (ALT), creatinine, total cholesterol, triacylglycerol (TAG), serum immunoglobulins (IgA, IgM, and IgG), interferon-γ (INF-γ), and interleukin-10 (IL-10). Also, the malondialdehyde (MDA), glutathione reduced (GSH), total superoxide dismutase (T.SOD) and glutathione S-transferase (GST) levels in the thigh and breast muscles were evaluated. One hundred and twenty one-day-old Cobb chicks were allocated into four equal groups that kept as a control group that supplemented with the basal diet, Turmeric I (0.5% of basal diet), Turmeric II (1.0% of basal diet) and Turmeric III (1.5% of basal diet) groups. Turmeric significantly increased the serum total protein, globulin, immunoglobulins, INF-γ, IL-10, muscle GSH, T.SOD, and GST. Whereas, serum total cholesterol, TAG, and muscle MDA levels were significantly decreased. The obtained results stated that Turmeric I (0.5%) gives the best feed conversion ratio with less feed intake as compared with the other turmeric-treated and control groups with enhancement of immunity and antioxidant activities. Also, birds in Turmeric I group had a significant decline in breast and thigh muscles MDA that produce a meat product of high quality and health benefits for human consumption.

1. INTRODUCTION Antibiotic growth promoters have been an integral part of the poultry feed industry. Instead, because of the antibiotic resistance in human, the herbs, spices, and their derivatives have antimicrobial, antioxidant and regulation of the gut flora in poultry production. Herbs stimulate body metabolism, improve digestion, have bactericidal, immunostimulant, and improved the productivity of poultry (Sapra and Mehta, 1990). Therefore, they used nowadays as antibiotic alternatives (Alloui et al., 2014). Plant active principles are an important task for developing therapeutic agents. Herbal components are extracted from different parts like leaves, bark, roots, flowers and seeds (Rajashree et al., 2012).

Application of medicinal plant as alternative feed additives is of important values on the performance and the antioxidant status of birds (Garcia et al., 2007; Ayoub et al., 2011; El-Far et al., 2016). The Curcuma longa L. (Family: Zingiberaceae) named turmeric is a perennial herb. Turmeric has received attention as a component of foods for its disease-preventing ability (Kelloff et al., 1996) mainly the inflammatory diseases and has a powerful antioxidant potential (Ammon and Wahl, 2007; Asai et al., 2014). The active ingredients found in turmeric are curcumin, desmethoxycurcumin, bisdemethoxycurcumin, and tetrahydrocurcuminoids (Osawa et al., 2014). Therefore, the effects of turmeric on broiler chickens were done by 19

Osman et al. 2017. AJVS 54(2): 19-28

was 2 min and diluted samples of 1 μl were injected automatically using auto-sampler AS3000 coupled with GC in the splitless mode. The ion source and quadrupole temperatures were set at 200 and 150°C, respectively (Barakat et al., 2016). The mass spectra of the identified components were determined by comparison to NIST MS/MS database. 2.4. Serum parameters At the 3rd and 5th weeks, blood sample was collected from all groups from the wing vein, left to coagulate at room temperature and centrifuged at 3000 rpm for 5 min. The clear sera were subjected to determination of the serum total protein, albumin, alanine aminotransferase (ALT, EC, creatinine, total cholesterol, and TAG following the instructions enclosed in the manufactured kits produced by Biodiagnostic Company, Egypt. Also, serum globulin levels were calculated by subtraction of albumin value from the total protein value of the same sample (Coles, 1986). 2.5. ELISA assays The serum levels of immunoglobulin A (IgA), immunoglobulin G (IgG), immunoglobulin M (IgM), Interferon-γ (INF-γ), and interleukin-10 (IL-10) were determined by ELISA kit (Elabscience Co, Wuhan, China). 2.6. Preparation of muscle tissue homogenate At the end of the experimental period, the birds of control and turmeric-treated groups (n= 15) were sacrificed under anesthesia with an intramuscular injection of sodium pentobarbital (50 mg/kg BW), and then muscle samples from left breast and left thigh of each bird were immediately eviscerated and immersed in ice-cold saline 0.9%. They were homogenized using a motor-driven Teflon and glass PotterElvehjem homogenizer in 0.1 M Tris-HCl buffer of pH 7.4 containing 5 mM β-mercaptoethanol (1:4 w/v). The homogenates were centrifuged at 105,000 ×g for 60 min at 4°C; the supernatants were divided into aliquots then stored at -20°C for further evaluation of oxidative stress and antioxidant parameters. 2.7. Determination of oxidative stress parameters 2.7.1. Determination of lipid peroxidation Malondialdehyde was analyzed after the incubation of supernatants with thiobarbituric acid at 95°C for 30 min (pH 3.6) resulting thiobarbituric acidreactive substances of pink color. MDA levels were assessed at 532 nm as nmol MDA/mg protein (Ohkawa et al., 1979).

determination of growth performance parameters, serum total protein, albumin, globulin, ALT, creatinine, total cholesterol, triacylglycerol (TAG), IgA, IgG, IgM, interferon-γ (INF-γ), and interleukin10 (IL-10). Also, the antioxidant potential of turmeric was monitored by determination of the levels of malondialdehyde (MDA), glutathione reduced (GSH), total superoxide dismutase (T.SOD), and glutathione S-transferase (GST) in the breast and thigh muscles. 2. MATERIAL AND METHODS

2.1. Birds, accommodation, and management The present study is affirmed by the Ethics of Animal Experiments Committee, Damanhour University, Egypt. Whereas, one hundred and twenty of Cobb broiler chicks, one-day-old, were floor reared, incubated and randomly allocated into four equal groups at the first week of age. Each group was subdivided into three replicates (10 birds per replicate). The housing of chicks was done in a clean well-ventilated room. The room temperature was scheduled according to age by electric heaters. Furthermore, the birds were vaccinated by Hitchner IB (7th day), Gumboro (14th day) and Gumboro and Clone (21st day) by eye drop.

2.2.Diet and experimental design The chicks were nourished on the feeding program from the 1st to 21st days on the starter and from the 22nd to 35th days on grower diets. The control diet composition was represented in Table 1 and analyzed according to AOAC (2005). The diet was formulated to meet the requirements of NRC (1994). Turmeric was washed, ground, and properly mixed with the basal ration by the concentrations of 0.5% in Turmeric I, 1.0% in Turmeric II and 1.5% in Turmeric III groups, while control one was fed a basal diet. All birds were accessed to water and feed ad libitum. 2.3. Gas chromatography–mass spectrometry (GC-MS) analysis The fine powder of turmeric powder was extracted by methanol by a dilution of 1: 3 (w: v). 10 µl of turmeric methanolic extract was injected in Trace GC Ultra-ISQ mass spectrometer with a direct capillary column TG–5MS (30 m×0.25 mm×0.25 µm). The column oven temperature was started 60°C and then increased by 5°C/min to reach 280°C. The injector and detector (MS transfer line) temperatures were kept at 250°C. Helium flow rate of 1 ml/min was used as carrier gas for 37.83 min. The solvent delay 20

Osman et al. 2017. AJVS 54(2): 19-28

2.7.2. Determination of reduced glutathione levels Reduced glutathione assay was depended on the reductive cleavage of 5, 5′-dithiobis (2-nitrobenzoic acid) (DTNB) by sulfhydryl group and advancement of a yellow color (Sedlak and Lindsay, 1968). The absorbance was recorded at 412 nm as µmol GSH/mg protein. 2.7.3. Determination of the Total superoxide dismutase activity The reduction of nitro blue tetrazolium with NADH-mediated by phenazine methosulfate (PMS) under aerobic condition was inhibited by superoxide dismutase. The radical being generated in the reoxidation of reduced PMS. This assay was resolved at 560 nm and represented as U/mg protein (Nishikimi et al., 1972). 2.7.4. Determination of the glutathione Stransferase activity The activity of GST was perceived following the protocol of Vessey and Boyer (1984). This test depended on checking the rate of enzyme-catalyzed conjugation of the 1-chloro-2,4-dinitrobenzene (CDNB) with GSH. GST activity was measured as the

increase in absorbance at 340 nm and signified as U/mg protein. 2.7.5. Determination of tissue protein Protein concentrations in muscle homogenates were evaluated using the bovine serum albumin as standard following the method of Bradford (1976). 2.8. Performance parameters The basal diets of both starter and grower phases were set up as per the of National Research Council Nutrient Requirements for Broiler Chickens (NRC, 1994). The final body weight, feed intake (FI), feed conversion ratio (FCR) (Lambert et al., 1936) and protein efficiency ratio (McDonald et al., 1987) were determined throughout the whole experimental period. 2.9. Statistical Analysis The obtained data were analyzed by one-way analysis of variance (ANOVA), with Duncan's multiple range tests for significant between means using SPSS software package v.20 (IBM SPSS. 20 ®, Michigan, USA). The data of ELISA assays and antioxidant status were analyzed by One-way ANOVA, Tukey’s multiple range tests by GraphPad Prism 5 (GraphPad Software, San Diego, CA, USA). All declarations of significance depended on p 0.05) among all groups at the 3 rd and 5th weeks. The data obtained in Figure 2B revealed that the serum IgG levels were significantly increased (P0.05) in Turmeric I in relation to control at the 3 rd week. Serum IgG levels were non-significantly increased (P>0.05) in Turmeric I, Turmeric II and Turmeric III in relation to control at the 5th week. The data obtained in Figure 2C revealed that the serum IgM levels were significantly increased (P