Abstract. 1. Introduction. S. Phommanivong and S. Doolgindachbaporn. วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม 2556

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1 วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม 2556 Effects of Moringa s Leave Supplementary Diet on Growth Performances and Survival Rates of Hybrid Catfish (Clarias macrocephalus x Clarias gariepinus) S. Phommanivong and S. Doolgindachbaporn Department of fisheries, Faculty of Agriculture, Khon Kaen University, Muang, Khon Kaen, 40002, Thailand. somphone05@gmail.com Abstract The study of utilization of moringa (Moringa oleifera Lam.) leaf on growth performance and survival rate in hybrid catfish (Clarias macrocephalus x Clarias gariepinus) was conducted in completely randomize design. Diets were prepared by supplementing with moringa leaf at 0, 5, 10 and 15%, all diets contained 35 % crude protein and isocaloric diets of 3,700 kcal/kg. The feeding trial was conducted in 16 glass-tanks. Each tank had 10 fish in capacity volume of 20 L as a replicate. In this experiment, fish with initial weight of 5.02±0.04 g/fish was fed with experimental diets for 5 weeks. The result showed that growth performance was not significant differences (p>0.05) in all groups, the highest weight gain of fish nursed at 5% of moringa leave meal in diet was 51.05±0.76 g and followed by 0, 10 and 15 % of moringa leave meal in diet were 50.72±3.84, 47.82±2.46 and 44.89±7.13 g, respectively. The specific growth rates were 6.88±0.19, 6.90±0.03, 6.73±0.17 and 6.51±0.35 %/day, respectively. Average daily gains were 1.44±0.11, 1.45±0.02, 1.36±0.07 and 1.31±0.17 %/day, respectively. Total lengths were 20.40±1.59, 19.36±1.30, 21.70±1.90 and 20.94±1.06 cm, respectively. And survival rate of fish was not significant difference (P>0.05), the survival rate ranged from 93.33±11.54 to 100%. In the part of fish health were found that, hematological parameters were not significantly different (P>0.05), hepatosomatic index (HSI), vesicle somatic index (VSI) and intestinal somatic index (ISI) were similarly in all groups. Therefore, supplementing moringa leaf with 15% in diet showed no adversed effect on growth, survival rate and fish health. Keywords: Moringa oleifera, Clarias macrocephalus x Clarias gariepinus. 1. Introduction Aquaculture is one sector growing very fast of world human food production and has an annual increase every year. To sustain such a high rate of growth, matching increase in fish feed production is imperative. On the other hand, the high cost of fish feed and quality of feed its uncertain availability have led to the need to identify supplement protein sources for fish feeds. Recently, the analysis of nutritional value of wild plant materials attracted attention due to the fact that they contain significant amount of essential nutrients that can be used for both human consumption and in the formulation of animal feeds (Eromosele, 1993). In order to attain a more economically sustainable and environmentally friendly production, research interest has been directed towards the evaluation and use of unconventional protein sources, particularly from plant products such as seeds, leaves and other agricultural by-products (Olvera et al., 1988). In last 10 years, extensive studies on agronomic aspects of moringa, that is widely distributed in the tropics and having a high fresh green matter production potential have been carried out (Foidl et al., 2001). Moringa oleifera, a number of the family Moringaceae, commonly referred to as drumstick tree is fast-growing plant with great economic importance for the food and medical industry. The seeds are a rich oil and protein source and can also use for the purification of water. The leaves and pods are used as food supplements for humans and animals (Makkar and Becker, 1997). All parts of this plant have been reported to be rich source of, protein, - 1 -

2 วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม vitamin C, calcium and potassium and act as good source of natural antioxidants and enhance the self-life of fat containing foods due to the presence of various types of antioxidant compound such as ascorbic acid, flavonoids, phenolics and carotenoids. The leaves are also rich in starch, mineral, iron, vitamin A, B and C, calcium, protein and including the essential sulfur amino acid, methionine and cysteine (Foidl, et al., 2001). Makkar and Becker (1997) reported that moringa leaves contain a negligible amount of tannins, but a high level of crude saponins about 5 %, which may have antinutritional effects in animal. Solvent extraction of leaves meal eliminates these antinutrients, and the extracted residues appear to be a good source of protein for fish. Recently, a preliminary study on the evaluation of the nutritional quality of raw moringa leaf meal in tilapia revealed that a 10% in diets did not cause any adverse effect on growth performance (Yuangsoi and Charoenwattanasak, 2011), whereas a severe growth depression occurred at higher 15% inclusion levels of moringa leaves meal in diet.the current study was designed to evaluate moringa leave meal in diets for hybrid catfish (Clarias macrocephalus x Clarias gariepinus) to evaluate growth performance and survival rate. 2. Materials and methods 2.1 Experimental diets Experimental diets of feed stuffs were given in Table 1. The recipe of the dietary feed stuffs was similar to the work reported by Boonarsa (2008), which has been shown to be adequate for Green catfish. The vitamin and mineral mixtures were also similar to that of the Boonarsa (2008), except that it did not contain the same levels of dry moringa leave meal. In this experiment, threediets were prepared to contain graded levels of dry moringa leave meal in diet 5, 10, and 15 %. The test diets were formulated to contain 35% crude protein. The diets were prepared by mixing the dry ingredients with oil and water by hands and then the wet dough was placed in a grinder and extruded to pass through a 2-mm mesh sieve and finally the moist pellets were dried in a room temperature, and then stored in the fridge at 10 0 C for further uses. Table 1. The ingredients being used in feed ration for the composition of the basal diet. Ingredient Diet (0%) Diet (5%) Diet (10%) Diet (15%) Fish meal Soybean meal Fulfat Moringa Rice bran Soybean oil Wheat powder Vitamin mixture Mineral mixture Proximate composition Experimental diets were analyzed according to AOAC (1990) procedures: dry matter (105 ºC to constant weight), ash (incinerated at 550 ºC to constant weight), crude protein (N 6.25) by the Kjeldahl method

3 วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม 2556 Table 2. The proximate analysis of moringa leaves and experimental diets. Proximate composition by analysis (% dry weight on basis) Composition Moringa leaves Diet (0%) Diet (5%) Diet (10%) Diet (15%) Moisture (%) Crude protein (%) Fat (%) Fiber (%) Ash (%) NFE (%) DE (Kcal/100 g) Fish maintenance for experiment The hybrid catfish (Clarias gariepinus x Clarias macrocephalus) fingerlings were obtained from a Kam Kaenkoon commercial farm, Thailand. They were carried out in 1000 L fiber tanks upon arrival at the Department of Fisheries, Khon Kaen University. The fish were acclimatized under fiber tanks conditions for 2 weeks during which they were fed with the same diet. The feeding trial was conducted in 16 glass-tanks. Each tank had a capacity volume of 30 L as a replicate where each of them contained 20 L of water. At the beginning of the experimental period, 10 fish somewhat of a similar size mean weight (MSE), 5.011±0.111g were nursed into each tank. A provided static water system with continuous aeration was used. The fish were hand-fed to satiation for hybrid catfish, 2 times per day at 8: 00 a.m. and 05:00 p.m. Every three days, feces and excess amount of feed were removed early in the morning to maintain water quality. Fish were batch-weighed tank by tank at each week. During the week 1 weighing period, the walls of each tank were scrubbed and washed before refilling the tanks with water and then the fish were returned to the tank accordingly. The feeding trial was carried out for 5 weeks. Water temperature was maintained at temperatures of C and mean ph values from Dissolved oxygen was no less than 4.0 mg/l, and the amounts of free ammonia and nitrite were not determined. 2.4 Evaluation of growth performance parameters The collected data on live weight gained (WG), specific growth rate (SGR), feed conversion ratio (FCR), average daily gain (ADG), total length, survival rate, protein efficiency ratio (PER),hepatosomatic index(his), vesicle somatic index (VSI) and intestinal somatic index (ISI)were calculated. WG (g) = [(W t minus W i )/W i ] x 100 SGR (% /day) = (Ln final wt - Ln initial wt)/days x100 FCR = total feed intake (g)/total wet weight gained (g) ADG (g) = (final wt initial wt)/period of experiment TL (cm) = total length Survival rate (% ) = 100x (final fish number)/(initial fish number). PER (%) = g weight gained/g protein intake HIS (%) = (liver mass (g)/ body mass (g)) 100 VSI (%) = (liver mass (g)/ total inner organ mass) 100 ISI (%) = (intestine mass (g)/ body mass (g))

4 วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม Hematological profile Blood samples were obtained from the caudal vein of 5 fish from each tank. 1 ml blood sample was collected in bottle containing 0.05 ml (EDTA) as anticoagulant. Blood samples for serum analysis were collected in bottles without any anticoagulant. Serum was separated by centrifugation at 1500 rpm for 10 minutes, and kept frozen at -20 ºC for determination of total protein, aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphate (ALP) activities which were analyzed by using automated instrumentation (Veterinary diagnosis laboratory, Animal hospital, Khon Kaen University). 2.6 Statistical analysis All of the collected data were subjected to one-way analysis of variance (ANOVA) and when the differences among the groups were identified, multiple comparisons of means were made with the use of Duncan s multiple-range test (DMRT) through the use of a computer programme (SAS). 3. Results and discussion 3.1 Growth performance and feed utilization The growth performance of hybrid catfish, weight gain, specific growth rate, average daily weight gain, total length, survival rate, protein efficiency ratio and feed conversion ratio from experiment were found that groups of fish were gave with various level of moringa leave meal in diet 5%, 10% and 15% were not significantly differences (P> 0.05) when compared with control group (0%). Yuangsoi and Charoenwattanasak (2011) reported that utilization of moringa (Moringa oleifera Lam.) leaf on growth performance and protein digestibility in Tilapia (Oreochromis niloticus L.) at 0, 5, 10 and 15% for 60 days. The average weight gain, specific growth rate, feed conversion ration and feed efficiency were not significantly differences in all groups (p>0.05). Richter, et al. (2003) reported that using moringa (Moringa oleifera Lam.) as an alternative protein source for Nile tilapia (Oreochromis niloticus L.) at 10%, 20% and 30% for 7 weeks were found that Nile tilapia fed with 10% of moringa in diet was the best growth performance, but Nile tilapia was fed with moringa from 20 and 30% of moringa leave meal in diet, the growth performances were reduced. (Afuang, et al., (2003) reported that utilization of moringa (Moringa oleifera Lam.) leaf on growth performance and feed utilization in Nile tilapia (Oreochromis niloticus L.) at 13, 27 and 40% were found that Nile tilapia fed with 13% of moringa leave meal in diet was reduced growth performance. Table 3.The responses of moringa leave meal in diets on growth, feed utilization and survival rate Parameters 0% 5% 10% 15% P-value IW (g) 5.04± ± ± ± WG (g) 50.72± ± ± ± SGR (%) 6.88± ± ± ± ADG (g) 1.44± ± ± ± FCR 2.25± ± ± ± PER (%) 1.26± ± ± ± TL (cm) 20.40± ± ± ± SR (%) 96.66± ± ±

5 วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม Hepatosomatic index, vesicle somatic index and intestinal somatic index At the end of the experiment, HSI, VSI and ISI were not significantly differences (p > 0.05). HIS were 1.75 ±0.12, 1.78±0.12, 1.82±0.09 and 1.82±0.28%, VSI were 3.64±0.14, 3.70±0.11, 3.76±0.17 and 3.741±0.12% and ISI were 3.72±0.16, 3.68±0.11, 3.75±0.24 and 3.81±0.74%. Afuang, et al., (2003) reported that hepatosomatic index of Oreochromis niloticus fed with moringa leaves meal in diets showed no significant change in liver size. Likely, Yuangsoi and Charoenwattanasak (2011) reported that utilization of moringa in diet at 0, 5, 10 and 15%, at the end of experiment, the hepatosomatic index (HSI) were not significantly differences (P>0.05) in all groups of experiment compared with control. From this experiment of using various levels of moringa leave meal in diet at 5, 10 and 15% showed no affect on liver function. Table 4. Hepatosomatic index, vesicle somatic index and intestinal somatic indexof hybrid catfish. Parameters P-value HIS (%) 1.75 ± ± ± ± ISI (%) 3.72± ± ± ± VSI (%) 3.64± ± ± ± Haematological parameters of fish fed experimental diets Haemotological parameters of hybrid catfish in this study showed that there were not significant differences (p>0.05) among fish fed various levels of moringa leave meal in diets. Hematocrit of fish fed with various levels of moringa leave meal in diet at 0, 5, 10 and 15% were 27.33±0.3, 27.93±0.59, 27.97±0.22 and 28.46±0.18 (%), respectively. Total protein (TP) were 5.03±0.60, 5.05±0.60, 4.92±0.80 and 5.07±0.60 g/dl, respectively. And BT were 15.10±0.77, 15.47±1.45, 14.23±1.35 and 16.26±1.40 mg/dl, respectively. Yuangsoi and Charoenwattanasak (2011) reported that hematological parameters of Nile tilapia fed with various levels of moringa leave meal in diet at 5, 10 and 15% showed that hematocrits were not significantly differentces (p>0.05) when compared with control groups. Fish haematology is increasing importance in fish culture because of its importance in monitoring the health status of fish. Aspartate aminotransferase (AST), alkaline phosphate (ALP) and alanin aminotransferase (ALT) activities value of fish fed with various levels of moringa leave meal in diet showed that there were not significant differences (p>0.05). Aspartate aminotransferase (AST) were ±0.37, ±2.08, ±2.01 and ±1.72 U/L, respectively, alkaline phosphate (ALP) were ±0.15, ±1.63, 150±1.45 and ±5.54 U/L, respectively and alanin aminotransferase (ALT) were ±11.17, ±0.21, ±1.33 and ±0.17 U/L, respectively. Therefore attention has been focused on the changes in AST, ALT and ALP activities, which promoted gluconeogenesis from amino acid, as well as on the changes in amino transferase activities in the liver. Table 5. Hematological parameters of fish fed with experimental diets. Parameters P-value Hct (%) 27.33± ± ± ± TP (g/dl) 5.03± ± ± ± TB (mg/dl) 15.10± ± ± ± AST (U/L) ± ± ± ± ALT (U/L) ± ± ± ± ALP (U/L) ± ± ± ±

6 วารสารว จ ย ป ท 6 ฉบ บท 2 กรกฎาคม - ธ นวาคม Conclusion The results of preliminary study on nutritional quality of moringa leaves as partial feed for fish indicated that up to 15% inclusion of this material could be recommended for hybrid catfish. In view of favourable amino acid profile of moringa leaves and moringa could be considered as a potential feed component with high nutritive value for fish. However, more investigations might be done for finding feasible and simple methods for reducing the antinutritional factors for inclusion of moringa leaves in practical diet for hybrid catfish at higher levels 5. References AOAC. (Association of Official Analytical Chemists) Official Methods of Analysis, 15 th ed. Association of Official Analytical Chemists, Arlington, VA, USA. 1,298 pp. Afuang, W., Siddhuraju, P. and Becker, K Comparative nutritional evaluation of raw, methanol extracted residues and methanol extracts of moringa (Moringa oleifera Lam.) leaves on growth performance and feed utilization in Nile tilapia (Oreochromis niloticus L.). Aquaculture, 34: 1,147-1,159. Boonarsa, P Effect of optimal premix and probiotic levels on nursing of green catfish, Mystus nemurus. Master Thesis in Fisheries. Khon Kaen: The Graduate School, Khon Kaen University 2008, (in Thai). Eromosele, I.C Plant food for human nutrition. (Netherland). Agric. Sci. 43(3): Foidl, N., Makkar, H.P.S. and Becker, K The potential of Moringa oleifera for agricultural and industrial uses. In: The Miracle Tree: The Multiple Attribute of Moringa (ed. by L.J. Fuglie), pp CWS, Dakar, Senegal. Makkar, H.P.S. and Becker, K Nutritional value and antinutritional components of whole and ethanol extracted Moringa oleifera leaves. Anim. Feed Sci. Technol. 63: Makkar, H.P.S. and Becker, K Nutrient and antiquality factors in different morphological parts of Moringa oleifera tree. J. Agric. Sci. 128: Olvera-Novoa, M.A,, Martinez-palacios, C.A., Galvan, C.R. and Chavez, S.C The use of seed of leguminous plant as a partial replacement for fish meal in diets for tilapia (Oreochromis mossambicus). Aquaculture 71: Richer, N., Siddhuraju, P. and Becker, K Evaluation of nutritional quality of moringa (Moringa oleifera Lam.) leaves as an alternative protein source for Nile tilapia (Oreochromis niloticus L.). Aquaculture 217: Yuangsoi, B. and Charoenwattanasak, S Utilization of (Moringa oleifera Lam.) leaf on growth performance and protein digestibility in Tilapia (Oreochromis niloticus L.). Department of Fisheries, Faculty of Agriculture, Khon Kaen University

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