Utilization of Moringa oleifera Lam.

leaves as an Alternative Protein Source of Soybean Meal in the Practical Diet of Chanos chanos (Forsskal) Fingerlings
Jessel Y. Tuazon and Graceous Von Yip Thesis Advisers: Dr. Lourdes V. Alvarez and Mr. Melchor M. Tayamen, OIC-NFRDI

ABSTRACT
Soybean meal, a widely used replacement to fishmeal, has been utilized as a protein source in the practical diet of Chanos chanos (Forsskal) fingerlings. However, in the Philippines, soybean meal production is scarce which forced the aquaculture industry to import the meal, thus, increasing its cost. The study was conducted to utilize the leaves of Moringa oleifera Lam. as an alternative protein source of soybean meal in the practical diet of Chanos chanos (Forsskal) fingerlings. Diets were formulated with 0%, 20%, 40%, 60%, 80% and 100% (-C, D1, D2, D3, D4, and +C, respectively) replacement of Moringa leaf meal on soybean meal. A forty-two day feeding trial was carried out using six 300L aquaria, integrated in a recirculatory system and compartmentalized into three (for triplicate set-up) with 30 fingerlings (1.84g; 55.43mm-standard length; 45.14mm-fork length and 7.08mm-body depth) each. Daily fish ration was calculated at 5% of their metabolic body weight per day. Body weight, standard length, fork length and body depth were measured weekly. Results revealed that the negative control (-C), still has potential superior effect on the growth performance of Chanos chanos (Forsskal) fingerlings obtaining the highest mean values in terms of body weight, standard length, fork length and body depth followed by D1 and D2 having the second and third highest mean values. Subsequently, the calculated values for Feed Conversion Ratio (FCR), Protein Efficiency Ratio (PER), Gross Feed Conversion Efficiency (GFCE) and Incidence of Cost showed that 20% and 40% inclusion of Moringa leaf meal can still be an effective alternative protein source on soybean meal. Keywords: Moringa oleifera Lam., practical diet, growth performance, protein utilization

INTRODUCTION
Aquaculture is the fastest growing sector of human food production with farmed seafoods which currently accounts for about 33% of all fish, crustaceans and molluscs consumed worldwide. The expansion of aquaculture production has been accompanied by rapid growth of industrially-manufactured aquafeeds. Regardless, the aquaculture industry must limit or eliminate their reliance on fishmeal, on which many aquafeeds are largely based, due to the increase of its price in the market over the past few years. (Richter et al., 2003). Aquaculturists have found conventional plant protein sources, such as soybean meal, which are effective in replacing fishmeal in the aquafeed. In the Philippines, however, soybean production is inadequate forcing the aquaculture industry to

import soybean meals making their costs too high (Fasakin et al., 1999). Moringa oleifera Lam., a member of the family Moringaceae, is a perennial softwood tree that is widely available in the tropics and subtropics and is advocated for its nutritional and medicinal properties (Becker and Makkar, 1999; Foidl et al., 2001). In recent times, its leaves have been promoted as an outstanding indigenous source of highly digestible protein, calcium, iron, Vitamin C, and carotenoid (Foidl et al., 2001). However, there is less information regarding the utilization of Moringa leaves in fish feeds. Therefore, the study was carried out to utilize the leaves of Moringa oleifera Lam. as an alternative protein source of soybean meal in the practical diet of Chanos chanos (Forsskal) fingerlings.

Utilization of Moringa oleifera Lam. Leaves as an Alternative Protein Source of Soybean Meal in the Practical Diet of

METHODOLOGY Preparation of Experimental System The study was conducted in six glass aquaria with 300L capacity, carried out in the Macrobrachium center at the Bureau of Fisheries and Aquatic Resources-National Freshwater Fisheries Technology Center, CLSU Compound, Science City of Muoz, Nueva Ecija. The site was provided with two fluorescent lamps to maintain a 12hr light: 12hr dark photoperiod of the fingerlings (Satyaranayana, 2009). Each of the aquaria was subdivided into three equal compartments for the triplicate set-ups using opaque plastic divisions. Twenty five circular spaces (2mm in diameter) were drilled in the plastic divisions to allow water to flow into the three compartments. Every aquaria were labeled with -C, D1, D2, D3, D4, and +C respectively and were filled with 270L of dechlorinated water. The aquaria were integrated into a recirculatory system. Diet Formulation Approximately 50kg of fresh Moringa oleifera Lam. leaves were collected along the vicinity of BFAR-NFFTC and Philippine Carabao Center (PCC), washed with tap water and were air-dried using hapa nets until they were crispy to touch, while retaining their greenish coloration. Dried leaves were manually pulverized using bare hands and kept in a Ziploc bag before feed formulation and analysis. The practical diet of Chanos chanos (Forsskal) fingerlings include soybean meal, rice bran, copra meal, corn bran and vegetable oil, as proposed by BFAR-NFFTC. Prior to feed formulation, the crude protein of Moringa leaf meal and the other ingredients were determined. The composition of the practical diet which contained no Moringa leaf meal served as the negative control (-C). The positive control (+C) was composed of the 100% replacement of Moringa leaf meal on soybean meal in the diet. Four experimental diets were formulated with Moringa leaf meal replacing soybean meal at inclusion levels of 20%, 40%, 60% and 80% and were designated as D1, D2, D3 and D4, respectively. (Table 1). Proximate composition, except crude ash, of the formulated diets was carried out using standard procedure. Crude Protein was determined by Kjeldahl Method and crude moisture by oven drying (AOAC, 1990). Crude fiber and crude lipid were analyzed in the Bureau of Animal Industry.

Experimental set-up and fish A group of 540 Chanos chanos (Forsskal) fingerlings reared at Bureau of Fisheries and Aquatic Resources-National integrated Fisheries and Technology Center were transferred to the experimental system, and kept in a rectangular tank for one-hour acclimation. The fingerlings (1.84g; 55.43mm-standard length; 45.14mm-fork length and 7.08mm-body depth) were randomly TABLE 1: Feed Formulation of the Control and Experimental Diets using Multiple Ingredient Method
(-)C Ingredients (in grams) Soybean meal Moringa leaf meal Rice bran Copra meal Corn bran Vegetable oil 0% 95 0 1 1 1 2 D1 20%* 76 19 1 1 1 2 D2 40%* 57 38 1 1 1 2 D3 60%* 38 57 1 1 1 2 D4 80%* 19 76 1 1 1 2 (+)C 100%* 0 95 1 1 1 2

*% inclusion of Moringa leaf meal replacing soybean meal

divided into eighteen groups of thirty fish, each of which was stocked and assigned to the six feeding treatments as labeled in each of the aquaria. Water quality was monitored during the experiment and temperature and pH were maintained at 29C and 7.7, respectively. The fish were fed with 5% of their metabolic body weight per day (four times feeding maintenance). Every week, fifteen randomly chosen fingerlings for every replicate were weighed individually to adjust feeding level for the subsequent week. Their standard length, fork length and body depth were also obtained. No feed was given before and during the sampling period and proper sedation technique using Tricaine methylsulfonate (MS222) was observed. Determination of Growth and Nutrient Utilization and Economic Evaluation The weekly weights recorded and feed supplied were used to compute the growth and nutrient utilization parameters following the method of Oliva Teles and Goncalves (2001). water hyacinth meal was put as processing costs. The economic evaluations of the diets were calculated from the method of New (1989)

Utilization of Moringa oleifera Lam. Leaves as an Alternative Protein Source of Soybean Meal in the Practical Diet of

Specific Growth Rate = Log wf Log wi x 100/t;

weight gain (g);

Food Conversion Ratio = Feed intake (g) / Fish Gross Feed Conversion Efficiency (%) = 1/Feed
Conversion rate x 100;

TABLE 3. Proximate composition of experimental diets of Chanos chanos (Forsskal) fingerlings (% based on dry matter base)

Protein Efficiency Ratio = Mean weight gain (g) /

Mean protein intake Incident of Cost = Cost of Feed (pesos)/Mean weight gain of fish produced (g)

Statistical Analyses Data were statistically processed for One-way Analysis of Variance (ANOVA) and Duncans Multiple Range Test (DMRT) using Statistical Packages for Social Sciences (SPSS) and Statistical Analysis Software (SAS) to find out the significant differences among the control groups and experimental groups. RESULTS AND DISCUSSIONS Composition of feed ingredients and formulated diets The crude protein contents of the ingredients including the soybean meal, Moringa leaf meal, rice bran, corn bran and copra meal are summarized in Table 2. The crude protein content of soybean meal is 43.5%, being the highest among the other ingredients followed by Moringa leaf meal (26.34%), copra meal (16.7%), rice bran (11%) and corn bran (6.8%). TABLE 2: Crude Protein of Feed Ingredients
Ingredients Soybean Meal Moringa leaf meal Rice Bran Copra Meal Corn Bran Crude Protein (%) 43.5 26.34 11 16.7 6.8

Values are means of triplicate determinations

The proximate composition, except for crude ash, of the diets formulated is presented in Table 3. The negative control (-C) has the highest value for crude protein (41.67%) and crude moisture (14.31%), as compared to the other diets which have shown decreasing values. Inversely, the positive control (+C) has the highest value for crude lipid (9.5%) and crude fiber (5.7%) as compared to the other diets which have shown decreasing values. The proximate analysis is dependent on the ratio between soybean meal and Moringa leaf meal. As the inclusion level of Moringa leaf meal increases, crude protein and crude moisture decreases and, inversely, the crude lipid and crude fiber decreases. (Tacon, 1995; Kaushik, 2004) Survival of the Fingerlings No feed-related mortality is observed all throughout the experimental period. However, certain mortalities are generally caused by mishandling during the sampling period and over-exposure to MS222. Tricaine methylsulfonate (MS222) is a compound developed by Sandoz and is a widely used anaesthesia for different fish species (Palmer and Mesinger, 2004). However, a formidable list of physiological consequences of use of MS222 have been documented, including many internal physiological changes which are greatly caused by over-exposure and chemical shock (Ross and Geddez, 1979)

Values are means of triplicate determinations

Generally, dry aquafeed ingredients are classified as energy sources- essentially of plant origin and contain less than 20% crude protein and protein sources- made of either plant or animal material and contain 20% crude protein or more. Soybean meal and Moringa leaf meal are then classified as protein sources and rice bran, copra meal and corn bran are considered as energy sources (National Agriculture Extension Research and Liaison Services, 2002).

Utilization of Moringa oleifera Lam. Leaves as an Alternative Protein Source of Soybean Meal in the Practical Diet of

a b c d e f

a ab ab bc c d

a ab b b c d

a b bc bc bc c

Values are mean of triplicate determinations standard deviations Values for each experiment in the each legend followed by different superscripts are significantly different (P 0.05)

FIGURE 3: Growth parameters of Chanos chanos (Forsskal) fingerlings subjected to experimental diets for 42 days Growth Parameters Figure 3 presents the average weight gain, average increase in standard length, fork length and body depth of Chanos chanos (Forsskal) fingerlings fed with the formulated diets for 42 days. It is observed that -C has superior effects on Chanos chanos (Forsskal) fingerling since it has the highest mean values in terms of body weight, standard length, fork length and body depth, followed by D1 and D2. Analysis of Variance (ANOVA) showed significant differences among the different diets in terms of the growth parameters (P<0.05). Moreover, multiple comparisons also showed significant differences among the different diets. The growth of the fingerlings is primarily dependent on the crude protein content of the formulated diets (Craig, 2004).

TABLE 4. Growth performance and protein utilization of the Chanos chanos (Forsskal) fingerlings fed with experimental diets. (-)C SGR(% day) FCR GFCE (%) PER 2.030.490a 1.000.075a 102.577.019a 32.513.196a D1 1.840.468ab 1.090.089ab 94.747.938b 30.373.010b D2 1.580.417ab 1.010.117a 103.229.018a 35.403.437c D3 1.480.405b 1.090.115ab 95.958.567b 37.283.803d D4 1.180.272c 1.360.117b 76.666.878c 34.143.543e (+)C 1.030.230d 1.530.138c 68.517.515d 34.093.186e

Values are mean of triplicate determinations standard deviations Values for each experiment in the same row followed by different superscripts are significantly different (P0.05)

Growth Performance and Protein Utilization Parameters The growth performance and protein utilization of the fingerlings per diet is summarized in Table 4. For specific growth rate (SGR), the negative control (-C) has the highest

value of 2.03% while the positive control (+C) obtained the lowest value of 1.03%. For feed conversion ratio (FCR), the negative control (-C) obtained the lowest value of 1.0g while the positive control (+C) obtained the highest value of 1.53g. Moreover, FCR value fluctuated among D1 (1.09g), D2 (1.01g) and D3 (1.09g). For Gross

Utilization of Moringa oleifera Lam. Leaves as an Alternative Protein Source of Soybean Meal in the Practical Diet of

Feed Conversion Efficiency (GFCE), the highest value is observed in D2 (103.22%) while the lowest value is observed in the positive control (68.51%). Lastly, for protein efficiency ratio (PER), highest value was obtained by D3 (37.28) and the lowest value was obtained by D2 (30.37%). Analysis of Variance (ANOVA) showed that no significant differences among C, D1 and D2 were observed (P>0.05). Moreover, multiple comparisons showed no significant differences among C, D1 and D2 in terms of SGR, FCR and GFCE. This implies that fingerlings will have a significant growth when fed with diets which contain 0%, 20% or 40% inclusion of Moringa leaf meal. Incidence Cost TABLE 5. Incidence of cost of formulated feeds Diet Incidence Cost (pesos) (-)C 21.25 D1 15.21 D2 10.73 D3 7.06 D4 3.41 (+)C 0.87 Table 5 presents the incidence cost of the formulated feeds. Results show that increasing the inclusion of Moringa leaf meal decreases the cost of feeds and with the positive effects on the growth performance and protein utilization, D1 and D2 are more economical to -C. CONCLUSIONS AND RECOMMENDATIONS The results of the study on the utilization of Moringa oleifera Lam. leaves as an alternative protein source indicate that 20% and 40% replacement of this material on soybean meal in the practical diet of Chanos chanos (Forsskal) fingerlings can be recommended for the favorable growth of the fish and their economic value compared to that of the practical diet. The researchers recommend to test the effectiveness of Moringa leaf meal on the growth of Chanos chanos (Forsskal) fingerlings in extensive and semi-intensive culture methods as means of comparison, increase the duration of the study to determine if usage of Moringa leaf meal would have adverse effects over time, include amino acid analysis and tests for anti-nutrient factors to

determine the real potential of Moringa leaf meal in aquafeed production and future studies must be done to test the effectiveness of Moringa leaf meal on other life stages of Chanos chanos. LITERATURES CITED Acda, S., Masilungan, H.G. and Moog, B. 2010.

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