Microsoft Word - 46 BIOTROPIA NO. 24, 2005 : 46 - 53 EFFECT OF ANTANAN (CENTELLA ASIATICA) AND VITAMIN C ON THE BURSA OF FABRICIUS, LIVER MALONALDEHIDE AND PERFORMANCE OF HEAT-STRESSED BROILERS ENGKUS KUSNADI1, REVIANY WlDJAJAKUSUMA2, TOHA SUTARDI3, PENI.S.HARDJOSWORO4 and ARIFIEN HABIBIE5 'Department of Animal Production.Faculty of Animal Husbandry, Andalas University(Unand), Padang, Indonesia 2Department of Physiology and Pharmacology,Faculty of Veterinary Medicine, Bogor Agriculture University(IPB), Bogor, Indonesia 3Department of Animal Nutrition & Feed Science, Faculty of Animal Husbandry, Bogor Agriculture Universirty(IPB), Bogor, Indonesia 4Department of Animal Production, Faculty of Animal Husbandry, Bogor Agriculture University (IPB), Bogor, Indonesia 1 Deputy Assistant for Agroindustry-Jakarta, Coordinating Ministry for Economic Affairs, Jakarta, Indonesia ABSTRACT High environmental temperatures may cause heat stress in poultry. This may increase water consumption, decrease feed consumption and in rum, decrease productivity level. In addition, high temperature contributes to oxidative stress, a condition where oxidant activity (free radicals) exceeds antioxidant activity. In our research, antanan (Centelta asiatica) and vitamin C were utilized as anti heat-stress agents for heat-stressed broilers. We used 120 male broilers 2 - 6 weeks old, kept at 31.98 ± 1.94 °C during the day and 27.36 ± 1.31 °C at night. The data collected were analyzed with a completely randomized factorial design of 2 x 3 (2 levels of vitamin C, 3 levels of antanan at 4 replications) and continued with the contrast-orthogonal test when significantly different. The results indicate that the treatments of 5 and 10% of antanan with or without 500 ppm of vitamin C and vitamin C alone significantly (P<0.05) decreased the heterophil/lymphocyte (H/L) ratio and liver malonaldchydc (MDA). These treatments, however, significantly (P<0.05) increased the bursa of Fabricius weight, feed consumption and body weight gain. It could be concluded that basal ration administered with 5% antanan and 500 ppm vitamin C could effectively prevent broilers from heat stress. The results support the conclusion that a basal ration supplemented with 5% antanan and 500 ppm of vitamin C or their combinations, effectivelly reduces heat stress in broilers. Key words : Heat stress/ Centella asiatica I Vitamin C INTRODUCTION High environmental temperatures may result in the accumulation of body heat load so that the body suffers from heat stress. As one of the homeothermic species, poultry could maintain their body temperature relatively constant by increasing respiration rate and water consumption and/or decreasing feed consumption. As a result, their growth rate and productivity will decrease. 46 BIOTROPIA NO. 24, 2005 May and Lott (2001) showed that body weight gain of 3 to 7-week-old male broilers raised at a temperature of 30°C was 1869 g significantly lower than for those raised at 22°C with body weight gains of 2422 g and feed conversion decreased from 3.28 to 2.54. The lower performances of broilers raised at high temperatures may have occurred as a result of lowered secretion of thyroid hormones (Geraert et al. 1996), decreased blood hemoglobin and hematocrit levels (Yahave/a/. 1997), or increased excretion of some minerals (Belay et al. 1992) and some amino acids (Tabiri et al. 2001). In addition, heat stress may also cause oxidative stress in the body and develop abundant free radicals, promoting the occurrence of peroxidation of membrane lipids and hence attacking DNA and protein membranes (Rahman 2003). Takahashi and Akiba (1999) indicated that feeding oxidized lipids to broilers significantly decreased feed consumption, body weight gain, plasma vitamin C, and plasma a-tocopherol. In fact, the results were followed by an increase in plasma malonal-dehyde (MDA) and blood heterophyl/lymphocytes (H/L) ratios as biological indices of stress in avian species. Antanan/pegagan (Centella asiatica (L.) Urban), one of the medicinal plants containing active materials such as asiatic acid, asiaticoside, and madecasic, is readily available and evidently eliminates stress in rats (Kumar and Gupta 2003). Shukla et al. (1999) reported that placing asiaticoside on rats wound increases curability and accelerates enzymatic and non- enzymatic antioxidant activities of new-growing tissues. In addition, vitamin C reportedly eliminated cold stress (Sahin and Sahin 2002) and heat stress in poultry (Puthpongsiriporn et al. 2001) and showed synergism with some active materials contained in antanan (Bonte et al. 1994). With this in mind, we have examined the effect of antanan (Centella asiatica) and vitamin C on the bursa of Fabricius, liver malonaldehide and performance in heat-stressed broilers. MATERIALS AND METHODS This research used 2 to 6-week-old male broilers placed in several pens located in an open poultry house. Each pen was flitted with a 40-Watt lamp and a zinc-plate backing functioning as a heat reflector. Temperature and relative humidity measurements obtained at noon and afternoon were 31.98 ± 1.28°C and 78.82 ± 5.43%, respectively. Temperature and relative humidity measurements at night and early morning were 27.36 ± 0.88°C and 86.23 ± 3.93%, respectively. The levels of 500 ppm vitamin C and 10% antanan (all plant parts) of ration used were established during preliminary trials. Vitamin C was dissolved in drinking water and served in the morning, two hours after the broilers received their last waterfeeding. 47 Effect of antanan (Centella asiatica) and vitamin C ‐ Engkus Kusnadi et al.  One‐hundred‐and‐twenty‐two‐week‐old male broilers were randomly allocated into 24 pens,  5  broilers  each.  Antanan  (5%  and  10%)  was  mixed  with  other  ingredients  to  make  three  different  rations  as  follows:  1)  The  control  ration  contained  the  calory  as  metabolizable  energy  (ME)  3245.02  kcal/kg  and  20.84%  crude  protein  ,  2)  A5  =  5%  antanan  contained  ME  3222.95 kcal/kg and crude protein 20.91%, and 3) A10 = 10% antanan contained ME 3202.87  kcal/kg  and  crude  protein  20.99%.  The  ration  formulation  and  nutrient  composition  of  treatments are presented in Table 1.                  The broilers were subjected to six treatments, 20 broilers each, as follows:  1) K (Control)/ration neither contained antanan nor vitamin C.  2) A5/ration was supplemented with 5% antanan  3) AlO/ration was supplemented with 10% antanan  4) C, drinkwater contained 500 ppm vitamin C  5) A5C, combination of A5 and C, and  6) A10C, combination of A10 and C  48  BIOTROPIA NO. 24, 2005 Variable measurements: 1) Relative bursa of Fabricius weight taken from 4-week-old broilers, by weighing the organ and divided by body weight (Puvadolpirod and Thaxton 2000). 2) Heterophyl/lymphocyte ratio was taken from 4-week-old broilers, by hemo-cytometer method. Blood was diluted 1:101 in a red blood cell pipette with Nat and Herrick diluent. The total leucocyte count includes heterophils, lymphocytes, monocytes, basophils, and eosinophils is divided by the number of lymphocytes. 3) Liver malonaldehyde (MDA) taken from 6-week old broilers, by measuring the thiobarbituric acid (TEA) value using Tarladgis method (Apriyantono et al. 1989). Destilate from liver sample with pH: 1.5, is added to the TBA reagents, covered, mixed, and incubated in boiling water bath for 35 minutes. After cooling, the absorbance of filtrate (D) was determined at 528 nm wave length. The TBARs values = 7.8 D were expressed as mg/kg of malonaldehyde per kg of tissue. 4) Feed consumption, body weight gain, and feed conversion are measured for 4 weeks (from 2 to 6-week-old broilers). Feed consumption was determined by weighing the given ration minus the leftover. Body weight gain was measured by weighing the final body weight at 6 weeks old minus the body weight at 2 weeks old. Feed conversion was measured by dividing feed consumption with body weight gain. Statistical analysis The collected data were analyzed using a completely randomized factorial design (CRD) 2 x 3 (2 levels of vitamin C i.e. 0 and 500 ppm and 3 levels of antanan i.e. 0, 5 and 10% of rations at 4 replications), and where applicable, continued with orthogonal contrast test according to Steel and Torrie (1980). RESULTS AND DISCUSSION The results of the effect of antanan and vitamin C administration on bursa of Fabricius weight, H/L ratio and liver MDA contents are presented in Table 2. The data for feed consumption (FC), body weight gain (BWG), and feed conversion are presented in Table 3. From Table 2, the average relative of 4 - week bursa of Fabricius weight for controls (K) is significantly lower than those treated with A5, A10, C, A5C, or A10C. The bursa of Fabricius weight is similar for all treatments from A5 to A10C. 49 Effect of antanan (Centella asiatica) and vitamin C ‐ Engkus Kusnadi et al.  These results suggest that antanan, vitamin C, or their combinations increase burs of Fabricius  weight of broilers that suffered heat stress.  The ability of antanan to stimulate lymphoid gland weight was reported i stressed rats  by Sharma et al. (1996). Antanan contains phenol compounds th< potentially can prevent  peroxidation of lipid membranes including T‐ and E lymphocyte membranes. T‐lymphocytes  produce cellular immunities while E lymphocytes produce humoral immunities produced by the  bursa of Fabricius in bir species. Phenol compounds of tea extracts were potentially capable of  stimulatin the production of  lymphoid cells  in rats (Murtini et al. 2003). On the other hanc  vitamin C acts, as a water‐soluble antioxidant capable of protecting lymphocyte from suffering  heat  stress  (Puthpongsiriporn  et  al.  2001).  As  a  result,  the  number  o  circulating  lymphocytes  increased so that the H/L ratio decreased.  Table 2 shows a relationship between the increase of the bursa of Fabriciu weight and  decrease in H/L ratio because the bursa of Fabricius is a lymphoid orgai producing lymphocytes.  Thus, the smaller the bursa of Fabricius size, the fewe lymphocytes will be produced, and in  turn, the higher the H/L ratio will be. Increase of the relative lymphoid organ weight occurred as a  result of antanan feeding whicl was demonstrated by Sharma et al. (1996), while increase of the  relative bursa o Fabricius weight as a result of supplementing vitamin C to broilers was reported  b] Anim et al. (2000).                                  Heat stress generating oxidative stress may increase MDA content as a result of lipid  peroxidation, especially for unsaturated fatty acids of membrane cells. Feeding  50    BIOTROPIA NO. 24, 2005 antioxidants, i.e. antanan and vitamin C, significantly decreases liver MDA content. Besides of being able to relieve free radicals by releasing an electron and a proton (a hydrogen ion), phenol compounds are also able to provide a chelating effect in such a way that phenols bind to transition ions. Unbound metals might increase free radicals (Pietta 2000). In addition, phenol compounds are characterized by flavonoids that are able to decrease fluidity of cell membranes so that it may decrease diffusion of free radicals and MDA contents. This is also true for vitamin C, a water-soluble antioxidant with 2 hydroxyl groups at €2 and €3 that are readily oxidized (Sediaoetama 1987).                  In addition, it is unmistakable that feeding antanan and vitamin C increases feed consumption and body weight gain of broilers over four weeks (2-6 weeks of age) but not for feed conversion (Table 3). This agrees with the reports of Anim et al. 2000) for broilers and Sharma and Sharma (2002) for rats suffering from stress. Antanan contains antioxidants such as phenol compounds that are capable of eliminating oxidative stress processes (Blokhina 2000), as is apparent from the decrease of H/L ratios, liver MDA levels and increase of bursa of Fabricius weights reported here.  CONCLUSIONS Feeding antanan or vitamin C or the combination of antanan and vitamin C increases bursa of Fabricius weight, feed consumption, and body weight gain but decreases H/L ratio and liver MDA levels. The combination of 5% antanan and 51   Effect of antanan (Centella asiatica) and vitamin C — Engkus Kusnadi et al. vitamin C tends to increase feed consumption and body weight gain and , therefore this treatment tends to be very effective in alleviating heat stress in broilers. 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