EFFECTS OF METHIONINE-SUPPLEMENTED TOASTED LIMA BEAN ( PHASEOLUS LUNATUS ) DIETS ON GROWTH OF OREOCHROMIS NILOTICUS

Adeparusi, E. O and Olute, B. WFisheries and Wildlife Department,Federal University of Technology,

P.M.B. 704,Akure, Nigeria

AbstractGrowth and packed cell volume of Oreochromis niloticus were analysed to determine the effect of replacing 20, 40, 60 and 80 % of menhaden fishmeal in a control diet at equi-protein level with lima bean supplemented with 1.4% methionine. Practical diets were formulated at 30% protein and 432 Kcal/100g gross energy. Fish were cultured at a mean of 24.31 0c, 7.4 and 6.15 mg/l temperature, pH and dissolved oxygen respectively and reared for 56 days. Fish fed 40% toasted lima bean diet had the highest survival rate of 100% and the best growth with specific growth rate (SGR) of 0.63, protein efficiency ratio (PER) of 0.28, protein productive value (PPV) of 30.37 and feed conversion ratio (FCR) of 2.95. Fish fed 20% inclusion of toasted lima bean diet had the least growth with 0.48 SGR, 0.21 PER, 19.13 PPV and 4.15 FCR respectively. There were no significant (P>0.05) differences in the mean weight gain, SGR and PER between fish fed the control and the 20% lima bean. Survival percentage (90%) was least on fish fed the control and those fed 80% lima bean diet.This study shows that the lima bean when toasted and supplemented with methionine can qualitatively replace 40-80% fish meal protein in the diet of Oreochromis niloticusINTRODUCTION

Fish is an important source of protein in the human diet contributing about 17% of the world animal protein supplies (Shang, 1992). Of the total wold production of fish which amounted to 112.30 million tonnes in 1995, 18.97% came from the aquaculture sector while the rest came from captured fishery (FAO, 1996). Annual rates of increase in global fish production, however, has slowed consistently over the past ten years (Watson and Blake, 1993). FAO (1996) reported a reduction in global fish production from 31.48 million tones in 1990 to 27.54 million tones in 1995, a total of 3.94 million tones reduction. Aquaculture has been proffered as the solution for bridging the gap between fish demand and production because it offers better opportunities for expansion (Watson and Blake, 1993). When considering aquaculture feeding of fish has to be taken into consideration. Fish meal remains the most widely used ingredient feed formulation because it is one of the most suitable sources of protein (Gallagher 1994; Gomes et al 1995). There are a number of factors however that discourage the use of high quantity of fish meal in feed formulation. Such factors includes: the high cost, scarcity, high production technology and the competitive use of fishmeal in animal feed and for human consumption (Gallagher, 1994, Gomes et al 1995). Out of the total fish produced in 1995, 71.17% was used for human consumption while the remaining was used for other fish based products such as fishmeal (FAO 1996). These factors have made it essential to seek alternative sources of protein in fish feed (Desilva and Gunasekera 1988).

Plant proteins are cheap and readily available, but they have some limitations that make them unsuitable for direct incorporation into animal feeds (Aletor and Fetuga 1984. Such factors include presence of anti nutritional factors that results in poor digestibility and low level of sulphur amino acids methionine and cysteine. These limitations could be successfully overcome by different methods such as heat treatment like toasting, autoclaving and cooking which destroys the heat-labile anti nutritional factor and improve digestibility or

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supplementing with methionine or cystine (Ologhobo et al 1992). Methionine is an indispensable amino acid required by terrestrial vertebrates, as well as various fish species, for normal growth and metabolic functions (Murthy and Varghese 1998). Plant proteins generally have unbalanced proportions of the essential amino acids. They have adequate level of lysine which is deficient in most cereals and other edible plant groups but are low in the sulphur amino acids methionine and cystine (NAS 1984, Ologhobo 1992). Methionine supplementation therefore has been employed as a means of improving the amino acid profile of plant proteins.

Lima bean, Phaseolus lunatus, is being investigated in this study, it is one of the under utilized legumes in Nigeria (Aletor and Aladetimi 1989). It has an amino acid profile similar to the cowpea varieties and the common beans (Ologhobo 1980, Aletor and Aladetimi 1989) but due to ignorance of its nutritional potentials, it has remained in the background. This study evaluated the effects of methionine supplemented lima bean (Phaseolus lunatus)diets on the growth and nutrient utilization of the commonly cultured Nile Tilapia, Oreochromis niloticus.

MATERIALS AND METHODS Culture ConditionsA total of 200 fingerling Nile tilapia were stocked into ten 45 L glass aquaria at the Fish Nutrition Laboratory of the Federal University of Technology, Akure, Nigeria. Water was continuously supplied from a borehole at the back of the laboratory. Water level in the tanks was maintained at a level of 30 cm through out the experimental period. 20 fingerlings were randomly stocked into each tank. There were three replicates per treatment. Fish were fed at 3% body weight with adjustments made in quantity of feed supplied every week. Feeding was done twice a day between 08.00 and 09.00Hrs and 18.00 and 18.00Hrs with the daily ration divided into two. Feeding was carried out for 8 weeks. Dissolved oxygen, water temperature and pH of the water were monitored through out the experiment.

DIET PREPARATIONToasting of beans Lima beans were put in a beaker and heated at 204oc for either 10,15,or 20 minutes. The beans were milled and included to replace 20, 40, 60 and 80% of fishmeal. The diets were designated as T2, T40, T60, T80 respectively according to the inclusion levels. All the lima bean diets were supplemented with 1.4% crystalline methionine. All feed ingredients were milled to pass through a 2mm diameter sieve, mixed together in a Hobalt mixer and pelleted using a 3.00 mm die.

Chemical & Carcass AnalysesFeed ingredients, diets and fish carcass were analysed for proximate composition. Proximate composition were determined for nitrogen, lipid, crude fibre (A. O.A.C, 1985). Fish carcass was analysed at the beginning and end of feeding trial. Energy content was determined using atomic bomb calorimeter

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Calculation of Essential Amino Acid of dietsThe essential amino acid component of each diet was calculated based on the levels present in all protein sources (fish meal, pigeon peal meal, blood meal, and groundnut-cake) as obtained from literature (NRC, 1993 and Tacon, 1990).

Biological evaluation:Biological evaluation was determined as follows:Weight gain (g) = Final weight of fish (W2) - Initial weight (W1).Average Daily Gain (g) =Weight gain /Days of feedingSpecific growth rate (SGR %day-) = 100 X (In W2 - In W1/T2-T1Where: T2-T1 represents the experimental periods in days.Protein Efficiency ratio (PER)= Weight gain (g)/Protein intake(g).Protein Productive Value (PPV) = 100 (Protein gain / Protein fed) where:Protein fed (PF)PF = Total feed consumed X % crude protein in feed.Feed efficiency ratio (FCR)= Fish weight gain (g)/Weight of feed consumed (g) Calculation of Essential Amino AcidThe essential amino acid component of each diet was calculated based on the levels present in all protein sources (fish meal, lima bean meal, blood meal, and soybean meal) as obtained from literature (NRC, 1993 and Tacon, 1990).

Statistical & Economic AnalysesData were analysed using Microsoft Excel, 1997 (Microsoft Corporation Nevada, USA) ANOVA procedure. Duncan’s multiple range test was used to determine differences among means (Zar, 1984). The prevalent prices of the component ingredients and fish were used to compute cost of producing 1kilogram feed and fish.

RESULTS AND DISCUSSIONResults are shown in Tables 2, 3, and 4. Mortality rates did not exceed two fish per diet and were probably not influenced by diets. The fish accepted the diets readily and consumed them within 15 minutes of administration. The proximate composition of the raw and toasted lima bean is shown in Table2, while the mean values is the water quality, parameters in the tanks measured weekly throughout the period of the experiment are presented in Table3. They were all within the tolerance range of the species.The best growth responses were obtained in fish fed diet T40 (40% lima bean inclusion level). This was followed by diet T60 which had 60% lima bean inclusion level. Diet T80 with 80% lima bean inclusion level was next. This was followed by diet CTR which was the control and had no lima bean. The least growth – performance was observed from diet T20 that had 20% lima bean inclusion level.The nutrient utilization parameters of fish on the different diets show that the best FCR, SGR and PER were obtained from diet T20. There was significant difference in the values of PER obtained for the five treatments at P<O.05.

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The crude protein level of all the diets showed no significant differences. Inspite of the above, diet T20 recorded the poorest growth performance. This could be due to an imbalance of plant to animal protein.When fed lima bean diet supplemented with methionine, O. niloticus performed better than when fed a diet containing about 20% fishmeal. This is in contract with the report of Abel et al (1984) who observed that mirror carp fed diets containing 50% heat treated full fat soya bean meal as a replacement for half of the fish meal attained only 60-65% of the growth obtained with the fishmeal control diet. The better performance of O. niloticus could be due to more available or better balance of essential amino acid.

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Reference:Abel, H.J., Becker K., Meske C.H.R., and Friedric W., (1984) Possibilities of using heat-treated full-fat soya beans in carp feeding Aquaculture 42.97-108.

Aletor, V.A. and Aladetimi O.O. (1989). Compositional evaluation of some cowpea varieties and some under-utilized edible legumes in Nigeria. Die Nahrung 33 (1989) 10. Pp 999-1007.

Aletor, V.A. and Fetuga, B.L. (1984). Effect of graded levels of raw lima bean (Phaseolus lunatus) on some liver enzyme activity in the rat. Nutrition Reports International March 1984. Vol. 29 No. 3.

A.O.A.C (1990): Official methods of Analysis. 15th Edn. Washington D.C. Association of Officail Analytical Chemists.

Badawi H.K anad Said M.M: A comparatively study of the blood of four Tilapia species (Pisces) Marine Biology 8,202-204 (1971).

Blaxhall (1971). The haemotological assessment of the Health of fresh water fish. J. Fish Biol. (1972) 4,593-604.

Delsilva S. Gunesekera R.M. and Atapatta O. (1989): The dietary protein requirement of young Tulip and an evaluation of the least cost dietary leve, Aquaculture 80:271.

Kelly, W. R (1979). Vet. Clinical diagnosis (2nd ed). Balliere Tindall, London.

FAO (1996): State of World Aquaculture.

Gallagher M.L., (1994). The use of soya bean meal as replacement for fish meal diets for hybrid bass (Morona saxatilis x M. chrysops). Journal of Aquaculture.

Gomes, E.F., Rema, P and Kaushik, S. (1995). Replacement of fishmeal by plant proteins in the diet of rainbow trout (Oncorhynchus Mykiss): digestibility and growth performance. Aquaculture 130, 177-186.

Murthy, H.S. and Varghese, T.J. (1998). Total Sulphur Amino Acid requirement of the Indian major carp, Labeo rohita (Hamilton). Aquaculture nutrition 1998 4; 61-65. National Academic of Science (NAS) (1984): The tropical legume resources for the future.

NRC (1983). Nutrient requirements of fish. Committee on Animal Nutrition, Board on Agric National R.C. Academy Press, Washington D.C. U.S.A 114p.

Ologhobo A.D. (1980). Biochemical and Nutritional Studies of cow pea and lima bean with particualr reference to some inherent nutritional factors. Ph. D Thesis, University of Ibadan, Ibadan, Nigeria.

Ologhobo A.D. (1992): Nutritive value of some tropical (West Africa) legumes for poultry J. Appl. Animal Res. 2:93-104.

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Shang, Y.C (1992). The role of aquaculture in the world fisheries presented at the world fisheries congress. Athens, Greece May 3-8. 1992. 30pp.

Tacon A.G.J. (1995). Fish meal replacers: Review of anti nutrients within oilseeds and pulses – A limiting factor for the aqua feed Green revolution? FAO, Rome.

Watson I. And B.F. Blake (1993). Aquaculture can answer the fisheries dilemma but how we feed the fish? In world Aquaculture (1993). (Editor Algan Cartwrigtht) PP 104-105. Stirling publication group Plc London.

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Table 1: Gross and Proximate Composition of Diets

DIETS (% Lima bean)

CTR DietT20 Diet T40 Diet T60 Diet T80

IngredientsFish meal 23.12 17.10 12.47 7.85 3.22Lima bean meal

- 12.22 23.40 33.61 45.78

Methionine - 1.4 1.4 1.4 1.4Soya bean meal

30.50 22.90 16.35 9.76 3.22

Blood meal 4.62 4.62 4.62 4.62 4.62Maize meal 30.91 28.42 27.41 26.41 25.40Vegetable oil 2.34 3.34 4.35 5.35 6.36Mineral mix 1.51 3.00 3.00 3.00 3.00Additional Vit.

2.00 2.00 2.00 2.00 2.00

Rice bran 5.00 5.00 5.00 5.00 5.00

Total 100 100 100 100 100Proximate AnalysisCrude protein 30.40 30.36 30.48 30.25 30.31Ether extract 10.05 10.25 9.82 9.97 9.98Ash 9.98 10.11 10.46 10.41 10.35

Crude fibre 5.60 6.04 5.73 6.06 5.85Moisture 2.21 2.90 2.65 3.35 2.45NFE 41.76 40.34 40.86 39.96 41.06Energy Kcal/g 436.97 432.79 431.50 427.95 432.89

Table 2: Proximate composition of raw and toasted lima bean (g\100g).

Raw LB Toasted LBCrude protein 22.82 22.61Crude lipid 2.58 3.11Crude fibre 4.11 3.61Moisture 11.20 8.31Ash 4.27 5.11Nfe 55.02 57.25

TABLE 3: WATER QUALITY PARAMETERS (MEAN WEEKLY)

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WEEKSParameters 0 1 2 3 4 5 6 7 8 x SD

Temperature (oc)

24.35 24.38 24.30 23.19 24.33 23.14 26.04 24.54 24.54 24.31 0.85

Dissolved oxygen (mg\

l)

6.38 6.72 6.06 6.16 5.60 5.46 6.90 5.44 6.64 6.15 0.56

PH 6.9 7.3 7.6 7.9 7.4 7.1 7.9 7.5 7.3 7.4 0.3

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Table 4: Growth performance and nutrient utilization of Oreochromis niloticus fed methionine supplemented Lima Bean

DIETSParameters CTR T20 T40 T60 T80 X 8D

Mean initial weight(g )

(w1)

4.92 5.04 5.47 5.72 5.16 5.26 0.328

Mean final weight (g)

(w2)

6.51 6.60 7.77 7.57 6.98 7.09 0.566

Mean wf.gain(g)

1.59bcd 1.56bcd 2.30a 1.86ab 1.82bc 1.83 0.297

Survival (%)

90 95 100 95 90 94 4.18

Av. Daily weight gain

0.028 0.028 0.041a 0.033b 0.033b 0.033 0.0053

Specific growth rate

0.50c 0.48c 0.63a 0.50c 0.55b 0.53 0.06

Protein efficiency

ratio

0.220cd 0.213cd 0.276a 0.245b 0.23bc 0.237 0.025

Protein productive

value

20.26d 19.13e 30.37a 21.87c 24.80b 23.29 4.50

Feed conversion

ratio

3.86b 4.15d 2.95e 3.77c 3.48d 3.64 0.454

Parked cell volume.

35.o 36.0 48.0 20.0 36.0 35.0 9.95

CARCASS COMPOSITION OF FISH FED LIMA BEAN DIETS

INITIAL CTR T20 T40 T60 T80Crude 58.62 64.78 64.43 67.88 65.23 66.14Protein 10.02 7.235 7.433 6.882 7.407 7.060

Moisture 9.85 9.30 9.25 8.78 9.02 8.98Ash 3.6 3.276 3.516 3.339 3.502 3.797NFE 17.88 15.40 15.37 13.12 14.84 14.07

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Hematological characteristics of Nile tilapia Oreochromis niloticusFed differently processed lima bean (Phaseolus lunatus L.) diets

Adeparusi, E. O and Ajayi, A. DFisheries and Wildlife Department,Federal University of Technology,

P.M.B. 704,Akure

ABSTRACT. The effect of substituting fishmeal protein with Lima bean on weight and some haematological indices was assessed using Oreochromis niloticus. Five diets, isonitrogenous and isocaloric, were fed in which 40% of the fish meal protein was substituted with either soaked, cooked, autoclaved or toasted lima bean protein. Fish fed the toasted lima bean diet had the highest growth while those fed the soaked bean diet had the least.The results showed significant (P<0.05) differences in packed cell volume (PCV), red blood count (RBC), haemoglobin (Hb), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC) and plasma protein (PP). Fish fed the control diet had the highest value of PCV, RBC, HB, and MCH was least in fish fed the soaked bean diet. The total white blood cell (WBC) count, lymphocyte, eosinophlis and neutrophils as well as erythrocyte sedimentation rate (ESR) were significant (P<0.05) different. Fish fed toasted lima bean diet had the highest WBC and eosinophils while those fed cooked lima bean diet had the least. Fish fed soaked lima bean had the highest value of ESR while fish fed on toasted had the least.Fish fed the control diet had the highest level of blood calcium, magnesium and phosphorus while fish fed the toasted had the least. Copper was not detected in any of the blood of fish on all diets. INTRODUCTION The demand for animal protein has gone far beyond supply as a result of the rapid human population in the developing countries. This therefore necessitates for an urgent and immediate need to increase the production of protein sources. Fish is one of the main animal protein sources and it has been cultured using different sources of feed ingredients (FAO, 1990). Fishmeal has been the main source of animal protein in fish feed. Its high cost and dwindling availability has called for the search for alternative sources of protein. Legumes have been severally tried as alternative protein sources in feed of several fish species. The main limitation to the use of legumes has been the presence of anti-nutritional factors (ANF). Such ANFs are reportedly responsible for the low digestibility of some plant proteins (Adeparusi and Jimoh, 2000, In Press). Many techniques have been used to improve the nutritional quality of legumes such as soaking in water, with or without mineral salts (Giami and Ikpimi, 1992), thermal processing (Adeparusi, 1994, Adeparusi and Balogun, 1998), germination (Bau et al, 1997), micronization (Igbasan 1997 et al) extrusion among others. Lima bean, Phaseolus lunatus is an underutilized legume grown in Nigeria and warmer areas of the World (McDonald and Law, 1984). Lima bean has amino acid profile similar to soybean and cowpea (Aletor and Aladetimi, 1989, Evans and Boulter 1974). Lima bean is used in some countries as green lunas, either frozen, canned or fresh (Martins et al, 1976). Lima bean contain some ANF that could be eliminated by some processing methods (Adeparusi, 1999). Among the several methods used in assessing protein quality in feed, blood evaluation is rarely used. Analyses of blood has been developed and well utilized in assessing the health of man and livestocks. It is therefore an important factor that could be considered in fish feed assessment. Everetth and Olusanya (1988) reported that fish blood consists of fluid with suspended cellular consistutients that circulates round the body. Svobodova, 1991 reported that ichthohaematology

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would be useful in the assessment of suitability of feeds and feed mixture, evaluation of fish conditions, determination of toxic effect of substances as well as diagnosis of disease.The Nile tilapia, Oreochromis niloticus makes up 36% of the total fish produced in Nigeria in 19995 (FAO, 1997). It is a highly favoured culture fish with worldwide acceptance. The dietary requirements of the Nile tilapia are well documented (NRC, 1983, Wilson, 1991). This fish is used in this study to assess the effect of feeding differently processed lima bean on haematological parameters.

MATERIALS AND METHODSPreparation of Meal Soaking: Lima bean were soaked in a 10% trona solution for 9 hours. The seeds were drained and The beans were dried in an oven at a temperature of 50oc for 2 hours.Normal cooking: Lima bean seeds were put in boiling water at atmospheric pressure for 1 hour drained and dried in an oven at a temperature of 50oc for 2 hours.Autoclaving: Lima beans were put in an autoclave at 121oc, 15 PSI for either 30 minutes. The cooked beans were removed and the cooking broth discarded. The beans were dried in an oven at a temperature of 50oC for 2 hours. Toasting Lima beans were put in a beaker and heated at 204o C for 10 minutes. Diet PreparationFishmeal content of the control diet was replaced with any of the processed lima bean meals on equal nitrogen basis (Table 1). All feed ingredients were milled to pass through a 2mm diameter sieve, mixed together in a Hobalt mixer and pelleted.Culture Conditions A total of 200 fingerling Nile tilapia were stocked into ten 45 L glass aquaria at the Fish Nutrition Laboratory of the Federal University of Technology, Akure, Nigeria. Water was continuously supplied from a borehole at the back of the laboratory. Water level in the tanks was maintained at a level of 30 cm through out the experimental period. 20 fingerlings were randomly stocked into each tank. There were three replicates per treatment. Fish were fed at 3% body weight with adjustments made in quantity of feed supplied every week. Feeding was done twice a day between 08.00 and 09.00Hrs and 18.00 and 19.00Hrs with the daily ration divided into two. Feeding was carried out for 8 weeks. Dissolved oxygen, water temperature and pH of the water were monitored through out the experiment.Statistical Analyses Data were analysed using Microsoft Excel, 1997 (Microsoft Corporation Nevada, USA) ANOVA procedure. Duncan’s multiple range test was used to determine differences among means. Correlation, regression and prediction equation were determined among some blood parameters (Zar, 1984).

RESULTS AND DISCUSSION Water quality parameters are presented in Table 2. All measured water quality values were within acceptable limits for culture of Nile tilapia (Wilson, 1991). Table 3 shows the weight gain and the haemogram of the fish to feeding. Fish fed dry heated diet had the highest weight gain while fish on the soaked bean diet had the least (Fig 1). Least growth of fish fed the soaked beans could be due to the presence of anti-nutrients which reduces the digestibilty and availability of nutrients (Kilgour, 1987). Higher growth of fish on processed diets shows that the tested processing methods were suitable and adequate for the removal/elimination of the anti-nutrients. Oliveiria (1991) found out that cooking increased the PER of rats fed cooked soybean. Dry heat was reported to have increased nutrient availability in pigeon pea fed to Clarias gariepinus

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(Adeparusi, 1994). Dry heat has been reported to improve the quality of soybean fed to rainbow trout (Smith, 1980). Fish on the control diet had the highest values in PCV, RBC, Hb and MCV but the least MCHC. Fish on the soaked bean diet had the least Hb and MCH. Badawi and Said (1971) listed some blood parameters of Nile tilapia, Tilapia zilli and Tilapia aureus. Changes in blood indices of fish as result of feed has been reported by Kelly, (1979); Kilgour (1987) and David and Tiggre, (1996). Ologhobo et al (1986) reported that the blood variables most consistently influenced by diet are the RBC, PCV. PP and glucose levels. Fish fed the cooked bean diet had the least values of RBC and PCV. This is agreement with the findings of Ologhobo et al (1986) who reported a lower RBC and PCV in chickens fed cooked soybean in comparison with the raw. Fish fed the dry heated bean diet had the highest mean cell haemoglobin concentration (MCHC). Although the values of PCV, Hb and RBC were higher in fish on the control diet they had the least MCHC. Table 4 shows the total and differential WBC of fish blood on the different diets. Fish on the dry heated diet had the highest WBC and eosinophil while fish fed the cooked bean diet had the highest monocyte and neutrophils. This could be due to optimal health conditions in fish on the cooked diet. The range of monocytes of Nile tialpia in this study is within the normal range. Kelly (1979) reported that monocytes comprise less than 10% of the total WBC production in animals of all species. Basophil was not recorded in fish on this study. This may be similar with what obtains in most livestocks. Kelly (1979) reported that basophil occur rarely in the blood of dog and cat and are very rare in all species of livestock. Lymphocytes were the highest proportion of the WBC in the blood of fish in this study. This is in contrast with the WBC composition of most livestock with neutrophil being the highest in dog, horse pig and cat while the highest in ruminants was eosinophils. The highest ESR was recorded in fish fed the soaked bean diet while the least value was in fish on the dry heat diet. High ESR in fish fed SLS could have due to low frictional resistance of the surrounding plasma. Low plasma protein causes reduced frictional or gravitational force on the erythrocytes thereby cau8sin it to have a high ESR (Kelly, 1979). Aletor and Egberongbe (1992) reported higher ESR in the blood of chicks fed unprocessed soybean than on those fed the processed soybean. Fish fed the dry heated beans had the highest PP but least ESR. The low ESR could have been due to the high quantity of PP in the blood which could increase the frictional force thereby reducing the velocity of the RBC. The mineral composition of the blood of fish on the diets are presented in Table 5. Fish fed cooked bean had the highest values of minerals while blood of fish on the dry heated diet had the least. Fish fed the CLS had the highest levels of calcium and magnesium. This could be due to the release of such metals usually chelated by phytic acid in the raw or soaked bean. Egberongbe (1992) reported that broiler chicken fed unprocessed soyabean have lower levels of calcium and magnesium.Table 6 shows the regression, correlation analyses and prediction equation of blood parameters of fish. There is a negative correlation between PCV and PP with a very low regression factor. There is high and positive relationship between PCV and Hb while a very weak positive relationship occurs between PCV and RBC. Relationship between RBC and Hb was positive and equally high. This shows that PCV could be used to predict the Hb content of blood while RBC will be a reliable prediction of RBC. Results from this study shows that processed lima bean could be well utilized by Nile tilapia. Fish fed on the cooked diet had growth that second highest growth while cooking reduced the RBC and Hb contents of fish. This could be due to the leaching of minerals that are required in

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the formation of blood. The highest weight gain, PP, Hb, RBC and MCHC were recorded in fish fed the dry heated beans. This shows that the processing methods used in this study were effective in reducing the ANF thereby making the nutrients available to the fish and increasing utilization.

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REFERENCESAdeparusi, E. O. (1994). Evaluation of the nutritive potential of cooked pigeon pea Cajanus cajan) meal as a plant protein source for Clarias gariepinus fingerlings. Journal of Agric. Tech, Vol. 2, No 1, 48 - 57.Adeparusi, E. O.and Balogun A. M. (1998): Evaluation of toasted pigeon pea, Cajanus cajan millisp meal as a partial replacement for fishmeal in diets of the African catfish, Clarias gariepinus Aletor, V. A. & O. O. Aladetimi (1989). Compositional evaluation of some cowpea varieties and some under utilized edible legumes in Nigeria. Die Nahrung 33 (1989) 10: 999 - 1007Badawi H. K and M. M. Said (1971) A comparative study of the blood of four Tulip species (Pisces). Marine Biology 8: 202-204.Bau, H.,C . Villaume, Nicolas J& Mejean L. (1997). Effect of germination on chemical composition, biochemical constituents &Anti-nutritional Factors of Soya bean (Glycine max) seeds. J. Sci Food Agric 73, 1-9. Delsilva S. Gunesekera R.M. and Atapatta O. (1989): The sietary protein requirement of young Tulip and an evaluation of the least cost dietary level, aquaculture 80:271.Bruno, D. W. & Prygve P. T (1996). A colour Atlas of salmonid disease. Academic press pub. 194 ppEvans M. I. & Boulter D. (1974). Composition of seed meals of yam bean (Sphenostylis stenocarpa) and lima bean (Phaseolus lunatus). Journal of Science. Food Agricultural 25: 919 - 922.Everett H. & Olusanya S. (1988). Anatomy and physiology of tropical livestock. ELBS, 1st Ed. Pg 30-38FAO (1990). Utilization of tropical food. Animal product, FAO Food and Nutrition. Paper 47. FAO, Rome, pg 8-21.Giami,S.Y and G.A . Ikpimi (1992). Effect of soaking on cooking characteristics of winged bean (Psophoearpus tetragonolobus L. DC). Nig. J. Nutr. Sci. Vol 13(1&2) 45- 49. Igbasan, F. A., Guenter, T. D., Warkerntsin and D. W. Mcandrew (1996). Protein quality of peas as influenced by location, nitrogen application and seed innoculation. Plant Foods Hum. Nutrition 49: 93 – 1Kelly, W. R. (1979). Vet. Clinical diagnosis (2nd edition) Balliere Tindall, London.Kilgour, O. F. G. (1987). Mastering nutrition. Macmillan Education Limited, London. 321 pp.MacDonald L and J. Low (1984). Fruit and Vegetables. Evans Brothers, London, 137pp.Martin J. H, H.L Warren, D.l Stamp (1976). 3rd Ed. Principles of field crop production.NRC (1983 ). Nutrient requirements of warm water fishes & shell fishes. Rev. ed. National Academy Press, Washington D. C, 102 pp.Oliveira J. E. D. (1973): Studies on the nutritive value of beans. In: nutritional aspects of common beans and other legume seeds as animal and human foods. Werner. G. Jaffe (ed.) Proc. of a meeting held in Ribeirao, Preto Pub by Archivos latinoamericanos de nutricion, Caracas - Venerzuela, 323 pp.Ologhobo, A. D. (1992). Nutritive values of some tropical (West African) legumes for poultry. Journal Appl. Animal Res. 2:93 - 104.Smith C., Wim Van Megen, Leendert Twaslfhoven and C. Hitch Cock (1980). The determination of Trypsin inhibitor levels in food stuffs. J. Sci. Food Agric, 31: 341 - 350Wilson R. P.(1991). Handbook of Nutrient requirement of finfish. Robert P. Willson (ed). CRC Press Inc, Florida 196pp.

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Svesbodora, Z, Fravda, D & Palakova, J. (1991). Unified methods of haematological examination of fish. Research Institute of Fish Culture & Hydrobiology, Vodnany, Czechoslovekia. 331pp.

Table 1: Gross and Proximate composition of lima bean diets (%) fed to O. niloticusIngredients CTR SLD CRD ALD TLDFishmeal 23.17 13.87 13.87 13.87 13.87Lima bean - 23.40 23.40 23.40 23.40Soyabean 30.50 16.35 16.35 16.35 16.35Blood meal 4.62 4.62 4.62 4.62 4.62Maize 30.91 27.41 27.41 27.41 27.41Vegetable oil 2.34 4.35 4.35 4.35 4.35Vit/min premix* 3.51 5.00 5.00 5.00 5.00Rice bran 5.00 5.00 5.00 5.00 5.00Total 100 100 100 100 100

Nutrients composition (%)Crude protein 30.40 30.28 30.22 30.22 30.27Crude fibre 5.60 6.48 6.11 6.18 5.11Ether extract 10.05 10.00 9.90 9.91 10.12Ash 9.98 10.34 10.33 11.05 10.81NFE 42.45 41.12 42.59 40.36 41.87Gross energy Kcal/100 g

439.76 433.16 437.90 428.69 437.32

Vitamin/mineral premix provided the following :Vitamins (IU or mg/kg): A, 5405.24 IU: D3, 2500 IU: E,72.2 mg: K, 4.9 mg: B12,19mg: riboflavin, 14.48mg: pyridoxine, 38 mg: thiamine, 12.2 mg: panthothenic acid, 144.57 mg: nicotinic acid, 240 mg: folic acid, 14 mg: biotin, 0.31 mg:Minerals (mg/kg)copper, 8.0: zinc,140.9 : iron,320 : iodine, 8.00: selenium, 0.16: manganese, 128:cobalt, 4.13

Table2: Mean Weekly Water Parameters.Parameters

Weeks Weeks Weeks Weeks Weeks Weeks Weeks Weeks Weeks Weeks Weeks

0 1 2 3 4 5 6 7 8 x SDTem (0c

24.35 24.38 24.30 23.19 24.33 23.14 26.04 24.54 24.54 24.31 0.35

DO2 (mg\1)

6.38 6.72 6.06 6.16 5.60 5.46 6.90 5.44 6.64 6.15 0.56

PH 6.90 7.30 7.60 7.90 7.40 7.10 7.90 7.50 7.30 7.40 0.30

Table 3: Mean Values of weight Gain and Heamatological Indices in O. niloticus Fed Differently processed Lima Bean Diets.Diets Weight

gain (g)PCV (%) RBC

(count) (x1012)T

Heamoglobin G/100ml

MCHC (%)

MCH (P.g)

MCV (FL)

Pp G/100ml

13

L-1SLD 1.995e 17.50bc 2.38cd 47.86c 28.50c 20.65d 7234.5c 1.27b

CLD 6.3b 18.50b 1.35e 47.86c 26.50d 36.85a 13846.16b

1.27b

ALD 3.31c 15.50d 2.28a 54.77b 36.50b 24.b 6637.08d 0.36dTLD 6.745a 16.50cd 2.66b 54.77b 34.50b 20.95d 6131.13e 1.57a

CRD 2.78d 40.50a 2.90a 68.59a 17.50e 23.96c 14035.54a

0.96c

X 4.24 21.70 2.31 54.77 28.70 25.41 9576.88 1.09SD 2.17 10.57 0.59 8.46 7.49 6.64 4003.39 0.46

Table 4: Differential count of fish on the different dietsDiets WBC

(count) (x103/CM3)

Lymphocyte (%)

Monocytes Neutrophils (%)

Eosinophils (%)

Basophils (%)

SLD 25.5b 67.5s 0.5c 31.5c 2.5ab 0.0CLD 18.5d 54.5e 5.5a 41.5a 0.0c 0.0ALD 22.5c 72.5b 2.5b 24.5 d 0.0 c 0.0TLD 27.5e 64.5d 0.5e 33.5b 3.5 a 0.0CRD 23.5c 24.5a 0.5e 25.5d 1.5-b 0.0 Mean with dissimilar superscripts within the same vertical colum are significantly different (P< 0.05).

Table 5: Physical property of blood of fish fed different processed lima bean Diets SLD CLD ALD TLD CRDESR milli 39.75b 27.15a 28.65c 18.85d 35.75CMean with different superscripts within the same column are significantly different (P,0.0)

Table 6: Means Of Mineral of Blood of O. niloticus fed differently processed Lima Beans.Diet Na (%) K (%) Fe (%) Ca (PPm) Mg (PPm) Cu (PPm) P (PPm)SLD 0.065b 0.085b 0.025 a 439.0 c 315.0b ND 563.5d

CLD 0.25a 0.27a 0.055 a 1305.0a 1105.0a “ 934.45a

ALD 0.075 b 0.095 b 0.025 a 607.5 b 357.85 b “ 605.0 c

TLD 0.065 b 0.12 b 0.03 a 363.5e 305.0 b “ 400.5 e

CRD 0.085 b 0.115 b 0.035a 389.0 d 321.0 b “ 704.0 b

With dissimilar superscripts within the same vertical column are significantly different (P<0.05).

Table 7:Regression and Correlation Analysis of blood parameter of fish fed on Lima bean based dietsSubject A B r Prediction equationPCV/PP 1.13 -0.004 - 0.088 Y = 1.13 – 0.004 x

14

PCV/Hb 39.91 0.69 0.87 Y = 39.91 + 0.69 xPCV/RBC 1.64 0.03 0.02 Y = 1.64 + 0.03 xRBC/Hb 31.83 9.98 0.71 Y = 31.83 + 9.98 x

%Wb Hb Pcv RBC MCH (pg)

McHC (Hb/100ml)

PPC MCV RBC RBC

Rw 49.6 2.05c 9.00d 15.00ba 6.02e 11.40 c 26.2c 60 15.00b 15.00S3H 63.12 2.73c 11.76 30.00ab 6.83e 11.55c 33.8b 59.97 30.00ab 30.00S6H 113.56 3.41bc 20ab 33.30ab 8.20cde 13.65c 28.4bc 59.95 33.30ab 33.3089H 179.55 4.77bc 22ab 36.70ab 9.29cde 15.50bc 21.5c 60.06 36.70ab 36.70C60 271.68 5.44b 23aw 28.30ab 18.17cd 20.74bc 18.5cd 60.00 28.30ab 28.30C90 112.07 4.77bc 18abc 30.00ab 14.45cde 24.06bc 18.5cd 60.05 30.00ab 30.00C120 129.59 4.09bc 17.99abc 38.30 12.45

cde30.28b 13.8e 60.07 38.30 38.30

T10 85. 5.45b 15bc 25.00b 20.41 b 34.06ab 15.4d 69.93 25.00b 25.00T15 177.78 7.50a 16bc 26.70ab 20.44 b 34.09ab 16.9 d 66.00 26.70ab 26.70T20 105.46 5.45b 22ab 36 .70 ab 21.80 b 36.33 ab 43.1a 59.95 36 .70ab 36 .70

bTmc 66.98 4.646 25 a 21.70 b 27.81 ab 46.34 ab 30.8b 60.00 21.70b 21.70X 123.126 1.516 17.977 29.245 15.079 25.268 24.264 61.453 29.245 29.245SD 65.130 2.05-

70504.679 7.042 7.157 11.751 9.087 3.338 7.042 7.042

Range

49.6-271.68

9.00-23.00

15.00-38.30

6.02-27.81

11.40-46.34

13.8-43.1

59.95.69.93

15.00-38.30

15.00-38.30

15