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1 J. of Aquaculture Vol. 9(4) : 54-60, 006 µ Journal of Aquaculture Korean Aquaculture Society e» e 9ƒ y d e w Ÿ,, ½, * w w, w l Apparent Protein and Phosphorus Digestibilities of Nine Different Dietary Protein Sources and Their Effects on Growth of Juvenile Olive flounder, Paralichthys olivaceus Gwangyeol Yoo, Se-Min Choi, Kang-Woong Kim and Sungchul C. Bai* Department of Aquaculture/feeds and foods nutrition research center, Pukyong National University, Busan , Korea Aquafeed Research Center, East Sea Fisheries Research Institute, National Fisheries Research and Development Institute, Pohang 79-80, Korea A 6-week feeding trial was conducted to determine apparent protein and phosphorus digestibilities in order to evaluate nine different dietary protein sources in growing olive flounder, Paralichthys olivaceus. Nine diets containing fish meal analog (BAIFA-M TM ), white fish meal (WFM), flounder muscle powder (FMP), squid liver powder (SLP), leather meal (LM), soybean meal (SM), corn gluten meal (CGM), poultry by-product (PBP) or egg albumin(ea) were prepared by mixing a basal diet (BD) with one of nine test ingredients at the ratio of 7 to 3. Apparent protein digestibilities of FMP, SLP, WFM, SM, CGM, LM, PBP, BAIFA-M TM, and EA were 94%, 9%, 86%, 8%, 75%, 7%, 7%, 7%, and 30%, respectively. Apparent phosphorus digestibilities of FMP, SLP, BAIFA-M TM, LM, WFM, PBP, CGM, EA and SM were 77%, 7%, 65%, 55%, 54%, 50%, 0%, 0%, and 7%, respectively. Weight gain of fish fed FMP (33 a ) was significantly higher than those of fish fed the other diets, and those of fish fed basal diet (30 b ), SLP (305 b ) and WFM (308 b ) diet were significantly higher than those of fish fed SM (74 c ), BAIFA-M TM (68 cd ), PBP (6 de ), LM (5 e ), CGM (54 e ) and EA (8 f ). Based on the results of apparent protein digestibilities, apparent phosphorus digestibilities and weight gain, SLP, SM and BAIFA-M TM could be one of good protein source to replace fish meal in flounder diets. Keywords: Digestibility, Protein, Phosphorus, Protein sources, Olive flounder ü ƒ 40~70% w w. x w» w v œ v w w œ w w w. w ùkü,» j» w œ w(kaushik and Cowey, 99). e p x¾ ü ƒ. ù x œ, ƒ, ƒ w w *Corresponding author: scbai@pknu.ac.kr 54 w ƒ š (Kim et al. 000; Choi et al., 004; Cho et al., 005a; Cho et al., 005b). w w ü dw eš, w yw Bioavailibility w ¾ w. tsƒ y dw ƒ» wù., y d,», w ƒe w v ƒ. w, y ³w w w. e» e y d e w dwš,

2 y d mw w w» w. x x w e e û m w, w w w 000 L x y w w, x x x j» w» w œ w. z, s³ 5.0Û0.06 g (meanûsd) e 60 L vp ƒ 0 0 x (» + 9ƒ x ) ƒ x 3 ew w. œ 3~5%(») z œ w. ƒ w œ w m e w ƒ sykƒ wš, x» 9~4 o C x» w, 3Û. L/min w, 6 x w. x x w»(basal diet, BD) Table w,» e(casein), p(gelatin) š e (Flounder muscle powder), ky p(dextrin) wš, w. w, 9ƒ x w Table ùkü. 9ƒ x» 70% 9ƒ 30% y ww. x 9ƒ BAIFA-M TM (t), (White fish meal, WFM), e (Flounder muscle powder, FMP), (Squid liver powder, SLP), (Leather meal, LM), e» e 9ƒ sƒ 55 Table. Composition of the basal diet Ingredients g/kg Casein 330 Gelatin 50.0 Flounder meal 3.0 Dextrin 76 Squid liver oil Corn oil Vit. mix Min. mix Attractant 6.0 CaCl 4.0 NaH PO Cr O Proximate analysis Moisture (%) 0. Crude protein (%, DM) 50.4 Crude fat (%, DM).5 Ash (%, DM) 6.3 Phosphrous (%, DM) 0.57 Baker Commodities Inc., L.A., USA. Prepared by lyophilized muscle from adult fish. 3 Ewha Oil Company, Busan, Korea. 4 Contains (as mg/kg in diets): Ascorbic acid, 300; dl-calcium pantothenate, 50; Choline bitatrate, 3000; Inositol, 50; Menadione, 6; Niacin, 50; Pyridoxine HCl, 5; Riboflavin, 30; Thiamine mononitrate, 5; dl-α-tocopherol acetate, 0; Retinyl acetate, 6; Biotin,.5; Folic acid, 5.4; B, Contains (as mg/kg in diets): NaCl, 437.4; MgSO 4 7H O ; ZnSO 4 7H O, 6.4; Fe-Citrate, 99; MnSO 4, 0.06; FeSO 4, ; CuSO 4, ; Calcium iodate, ; MgO, ; NaSeO 3, (Soybean meal, SM), g l(corn gluten meal, CGM), ƒ (Poultry by-product PBP), š ù (Egg albumin). Cr O 3 0.5% ƒ z, mm r» xw w, x Table 3 ùkü. j» sieve š e z, w -0 o C þ w w, Table. Proximate composition of dietary protein sources (% of dry matter) Ingredients Moisture Protein Lipid Ash Phosphorus BAIFA-M TM White fish meal Flounder muscle powder Soybean meal Corn gluten meal Leather meal Poultry by-product Squid liver powder Egg albumin BAIFA-M TM = fish meal replacer. Prepared in this laboratory by proper composition. Mixture of the following ingredients: blood meal, squid liver powder, meat and bone meal, leather meal, feather meal, poultry by-products, Attractant. Prepared by lyophilized muscle from adult flounder.

3 56 Ÿ,, ½, Table 3. Chemical composition of reference and test diet (%, dry matter) Diets Moisture Protein Lipid Ash Phosphorus BD BAIFAD WFMD FMPD SMD CGMD LMD PBPD SLPD EAD Pooled SEM BD = basal diet, BAIFAD = BD (70%) + BAIFA-M TM (fish meal replacer, 30%), WFMD = BD (70%) + WFM (white fish meal, 30%), FMPD = BD (70%) + FMP (flounder muscle powder, 30%), SLPD = BD (70%) + SLP (squid liver powder, 30%), LMD = BD (70%) + LM (leather meal, 30%), SMD = BD (70%) + SM (soybean meal, 30%), CGMD = BD (70%) + CGM (corn gluten meal, 30%), PBPD = BD (70%) + PBP (poultry by-product, 30%), EAD = BD (70%) + EA (egg albumin, 30%). Pooled standard error of mean. œ 3.87Û0.6%(») z(08:00, 6:00) œ w. d d w, dw» w 4 k z MS- (00 ppm) g dw. x z, (percent weight gain, %), (specific growth rate, %/day), yz(protein efficiency ratio), z(feed efficiency, %), (hepatosomatic index), (condition factor) (survival rate, %) w. w ƒ 3 dw.» d w. Wt. gain (%) = (final wt. - initial wt.) 00 / initial wt. Feed efficiency (%) = (wet weight gain / dry feed intake) 00 Specific growth rate (%/day) = (loge final wt. - loge initial wt.) / days Protein efficiency ratio = (wet weight gain / protein intake) Hepatosomatic index = (liver weight / body weight) 00 Condition factor = [fish wt. (g) / fish length (cm)3] 00 x ƒ 5 w w w, AOAC (Association of Official Analytical Chemists, 000) ƒ (35 o C, ), kjeldahl (NÜ 6.5), z zy w. v w z, soxtec system 046 (Tacator AB, Sweden) w soxhlet w. x z, wì x w x xw» ¾ 4 g. x ƒ 3 w z z» w x x xwš micro-hematocrit (Brown, 980) w xmjp(hematocrit, PCV) dw, Drabkin s w cyan-methemoglobin (Sigma Chemical, St. Louis MO; total hemoglobin procedure No. 55) x (hemoglobin, Hb) dw. x w y Cr O 3 w ICP (Inductively Coupled Plasma, Perkinelmer ICP-OES 000DV, ) w w. y d x z, ƒ x w ù šw e g y d w. ƒ x œ 0 6 w, œ z É» w. ú 8 e w. 7 w. z k z w ¾ -0 C o w. y w» w» x y Maynard and Loosli (969) œ w w, ƒ y Sugiura et al. (998) w œ w. Apparent digestibility coefficients (ADCs) = 00 - [{nutrient in feces (%) Cr O 3 in diet (%)}Ý {nutrient in diet (%) Cr O 3 in feces (%)} 00] ADCs of the test ingredient = (nutrient in test diet ADCs of test diet nutrient in basal diet ADCs of basal diet) / (0.3 nutrient in test ingredient)

4 m m Computer Program Statistix 3. (Analytical Software, St. Paul MN. USA) (ANOVA test) w (LSD: Least Significant Difference) s³ (P<0.05) w. sƒ 6 w e Table 4 ùkü., z, e x ƒ x w ùkû(p<0.05). š,» x (BD), x (WFMD), x (SLPD) x (SMD), BAIFA x (BAIFAD), ƒ x (PBPD), g l x (CGMD), x (LMD), ù x (EAD) ùkû(p<0.05).,» x (BD), x (WFMD), x (SLPD) ƒ ùkù, ù x (EAD)ƒ x w ƒ û ùkû(p<0.05). yz x (SLPD)ƒ x w ùkû, w» (BD), x (WFMD), e x (FMPD)ƒ x (SMD), BAIFA x (BAIFAD), ƒ x (PBPD), g l x (CGMD), x (LMD), ù x (EAD) ùkû(p<0.05).,» x (BD), x (WFMD), e e» e 9ƒ sƒ 57 Table 4. Effects of the basal diet and nine test diets on growth performances in juvenile olive flounder Table 5. Apparent digestibilities of protein and phosphorus of nine different protein sources Digestibility(%) 4 Ingredients Protein Phosphorus BAIFA-M TM 7 d 65 b WFM 86 b 54 c FMP 94 a 77 a SLP 9 a 7 a LM 7 c 55 c SM 8 b 7 d CGM 75 c 0 d PBP 7 c 50 c EA 30 e 0 d Pooled SEM Values are means from triplicate groups, values in the same column not sharing a common superscript are significantly different (P<0.05). BAIFA-M TM =fish meal replacer, WFM=white fish meal, FMP=flounder muscle powder, SLP=squid liver powder, LM=leather meal, SM=soybean meal, CGM=corn gluten meal, PBP=poultry byproduct, EA=egg albumin 3 Pooled standard error of mean. 4 Apparent digestibility values were calculated as previously described by Sugiura et al. (998). x (FMPD) ƒ ùkù. z e x (FMPD)ƒ x w ƒ ùkû, ù x (EAD) x w ƒ û ùk û(p<0.05). e x (FMPD), x (SLPD), x (WFMD),» (BD)ƒ w, w ü Diets Pooled BD BAIFAD WFMD FMPD SLPD LMD SMD CGMD PBPD EAD SEM 3 WG (%) 4 30 b 68 cd 308 b 33 a 305 b 5 e 74 c 54 e 6 de 8 f 7.30 FE (%) 5 0 b 90.0 c 05 b a 03 b 83.0 e 9.0 c 85.0 de 88.0 cd 6.0 f.53 SGR b 3.0 c 3.35 b 3.43 a 3.33 b.99 e 3.4 c 3.0 e 3.06 cd.46 e 0.05 PER 7.94 b.64 de.88 bc.94 b.7 a.57 de.83 c.55 e.64 cd 0.97 e 0.06 CF PCV Hb(g/dl) Values are means from triplicate groups, values in the same row not sharing a common superscript are significantly different (P<0.05). BD = basal diet, BAIFAD = BD (70%) + BAIFA-M TM (fish meal replacer, 30%), WFMD = BD (70%) + WFM (white fish meal, 30%), FMPD = BD (70%) + FMP (flounder muscle powder, 30%), SLPD = BD (70%) + SLP (squid liver powder, 30%), LMD = BD (70%) + LM (leather meal, 30%), SMD = BD (70%) + SM (soybean meal, 30%), CGMD = BD (70%) + CGM (corn gluten meal, 30%), PBPD = BD (70%) + PBP (poultry by-product, 30%), EAD = BD (70%) + EA (egg albumin, 30%). 3 Pooled standard error of mean. 4 Weight gain (%) : (final wt. - initial wt.) 00 / initial wt. 5 Feed efficiency (%) : (wet weight gain / dry feed intake) 00 6 Specific growth rate : (loge final wt. - loge initial wt.) / days. 7 Protein efficiency ratio : wet weight gain / protein intake 8 Condition factor : [fish wt. (g) / fish length (cm)3] PCV = Hematocrit. 0 Hb (g/dl) = Hemoglobin.

5 58 Ÿ,, ½, w yz y x ƒ ùkù w. š ù x w wš û., xmjp, x x ƒ ùkù. ysƒ 9ƒ w y Table 5 ù kü. y e 94%, 9%, 86%, 8%, g l 75%, 7%, ƒ 7%, BAIFA-M TM 7%, ù 30% ùkû. y e 77%, 7%, BAIFA-M TM 65%, 55%, 54%, ƒ 50%, g l 0%, ù 0%, 7% ùkû. š y w sƒ w ƒ w, ky p w y d Cr O 3 yww dw š (Maynard and Loosli, 969; Cho et al., 99; Cho et al., 994). w, y sƒ» sƒwš w 7:3 yww œ w š. w sƒwš w w w w» w, p» e, p, p wš. w w» k x š. w, w w y»y» ƒwù, yw w ƒ š (Teshima et al., 993, Berge et al., 999, Olsen et al., 006). e y sƒƒ» e (Flounder muscle powder, FMP) w y sƒ w. FMP w 85.4%, zw 3.7% û ùkû, y ƒƒ 94%, 77% ùkû» e x ƒw. w v y x v ƒw w(choi, 00). (Squid liver powder, SLP) w 45% w w û, y FMP w x w w ùkû(p<0.05). w, SLP zw û w p, w v EPA&DHAƒ tw ƒ z w w q(park, 000). š SLP w 0.7% w û ùkû, y w ùkû. SLP e ƒ w, v w ƒ ùkû(choi, 00). w»y w ƒ, š. (White fish meal, WFM) w 7.7%, y 86% ùkû. w 3.3% ƒ, 54% ùkû» y»k ƒ d. û ƒ, œ, w ³x v w ü w ƒ (Murai and Nose., 98), (Cho et al., 974),»(Belal and Assem., 995), e(kim et al., 000; Choi et al., 004), v(lim et al., 004). (Soybean meal, SM) y 8% w Chinook salmon (NRC, 993), v(choi et al., 004) ù kû, (NRC, 993),»(NRC, 993), Blue tilapia (NRC, 993), Red drum (Gaylord and Gatlin, 996) û ùkû. w, ky yw û y û ùkù» (Rychly and Spannhof, 979; Sullivan, 993). x y 7% û ùkû, w w»k xk vpk (Ketola and Harland, 993) vpk ƒwj z phytase ƒ w» (NRC, 993). g l(corn gluten meal, CGM) y 75% v w(choi, 00), y 0% SM ƒ û ùkû. (Leather meal, LM), ƒ (Poultry by-product, PBP) y 7%, y ƒƒ 55%, 50% ùkû. v y w, LM w, PBP 5% ùkû. t BAIFA-M y û TM, ƒ, w 7% û ùkû. w, y 65% ùkû» y ƒw

6 y w w yz ƒ, z ƒœ w q. ù(egg albumin) w, w. w, e ù y 30% û ùkù w q., sƒ x mw, w w ƒ w q, t BAIFA-M TM y ƒš», yzƒ z mw y w w ƒ ƒ, w y ƒš» Phytase ƒ y w w mw ƒ. ü ƒ š e y d mw w kw. x 9ƒ BAIFA-M TM (t), (White fish meal, WFM), e (Flounder muscle powder, FMP), (Squid liver powder, SLP), (Leather meal, LM), (Soybean meal, SM), g l(corn gluten meal, CGM), ƒ (Poultry by-product PBP), š ù(egg albumin). x, ƒ y FMP, 94%; SLP, 9%; WFM, 86%; SM, 8%; CGM, 75%; LM, 7%; PBP, 7%; BAIFA-M TM, 7%; EA, 30% ùkû. ƒ y FMP, 77%; SLP, 7%; BAIFA-M TM, 65%; LM, 55%; WFM, 54%; PBP, 50%; CGM, 0%; EA, 0%; SM, 7% ùkû. w p w,. w ww w w x ¾ tw. šx AOAC (Association of Official Analytical Chemists), 000. Cunniff, P. (Ed.), Official Methods of Analysis of the Association of Official Analytical Chemists, 6th edition. Association of Official Analytical Chemists, Inc., Arlington, VA. e» e 9ƒ sƒ 59 Belal, I. E. H. and H. Assem, 995. Substitution of soybean meal and oil for fish meal in practical diets fed to channel catfish, Ictalurus punctatus: effects on body composition. Aquaculture Research, 6, Berge, G. M., B. Grisdale-Helland and S. J. Helland, 999. Soy protein concentrate in diets for Atlantic halibut Hippoglossus hippoglossus. Aquaculture, 78, Brown, B. A., 980. Routine hematology procedures. (in) B. A. Brown (ed.), Hematology: Principles and Procedures. Lea and Febiger, Philadelphia, pp. 7. Cho, C. Y., H. S. Bayray, and S. J. Slinger, 974. Partial replacement of herring meal with soybean meal and other changes in a diet for rainbow trout (Salmo gairdneri). J. Fish. Res. Board Can. 3, Cho, C. Y., J. D. Hynes, K. R. Wood, and H. K. Yoshida, 99. Quantitation of fish culture wastes by biological (nutritional) and chemical (limnological) methods; the development of high nutrient dence (HND) diets. (in) Cowey, C. B. and C. Y. Cho (eds.), Nutritional Strategies and Aquaculture Waste. Proceeding of the First International Symposium on Nutritional Strategies in Management of Aquaculture Waste, University of Guelph, Ontario, Canada, pp Cho, C. Y., J. D. Hynes, K. R. Wood, and H. K. Yoshida, 994. Development of high nutrient dence, low pollution diet and prediction of aquaculture waste using biological approaches. Aquaculture, 4, Cho, S. H., S. M. Lee, S. M. Lee, B. H. Park, I. S. Park, C. Y. Choi, B. H. Min, S. B. Hur and J. Y. Jo, 005a. Effect of partial replacement of fish meal with squid liver meal TM in the diet on growth and body composition of juvenile olive flounder (Paralichthys olivaceus) during winter season. J. Fish. Sci. Technol, 8() Cho, S. H., S. M. Lee, B. H. Park, I. S. Park, C. Y. Choi, S. M. Lee, B. H. Min, S. B. Hur and Y. S. Kim, 005b. Effect of partial dietary substitution of meat meal for fish meal on the growth and body composition of the juvenile olive flounder Paralichthys olivaceus. J. Fish. Sci. Technol, 8(3), Choi, S. M., 00. Apparent protein and phosphorus digestibilities of dietary protein sources in juvenile Korean rockfish, Sebastes schlegeli. Master course dissertation, Pukyong National University, Busan, Korea. Choi, S. M., S. R. Lim, K. W. Kim, X. J. Wang, S. C. Bai, and I. S. Shin, 004. Effects of dietary dehulled soybean meal as a fish meal replacer in fingerling and growing olive flounder, paralichthys olivaceus. Aquaculture research 35, Gaylord, T. G. and D. M. Gatlin III, 996. Determination of digestibility coefficients of various feedstuffs for red drum Sciaenops ocellatus. Aquaculture, 39, Kaushik, S. J. and C. B. Cowey, 99. Ammoniogenesis and dietary factors affecting nitrogen excretion. (in) Cowey, C. B. and C. Y. Cho (eds.), Nutritional Strategies and Aquaculture Waste, University of Guelph, Ontario, Canada, pp Ketola, H. G., and B. F. Harland, 993. Influence of phosphorus in rainbow trout diets on phosphorus discharge in effluent water. Transation Americna Fisheries Society,, 0 6.

7 60 Ÿ,, ½, Kim, Y. S., B. S. Kim, T. S. Moon and S. M. Lee, 000. Utilization of deffated soybean meal as a substitute for fish meal in the diet of juvenile flounder (Paralichthys olivaceus). J. Korean Fish. Soc, 33(5), Lim S. R., S. M. Choi, X. J. Wang, K. W. Kim, I. S. Shin, T. S. Min and S. C. Bai, 004. Effects of dehulled soybean meal as a fish meal replacer in diets for fingerling and growing Korean rockfish Sebastes schlegeli. Aquaculture, 3, Maynard, L. A. and J. K. Loosli, 969. Animal nutrition, 6th ed. McGraw-Hill, New York, St Louis, pp. 63. Murai, T., H. Ogata and T. Nose, 98. Methionine coated with various materials supplemented to soybean meal diet for fingerling carp Cyprinus carpio and channel catfish Ictalurus punctatus. Bulletin of the Japanese Society of Scientific Fisheries, 4, National Research Council (NRC), 993. Nutrient Requirements of Fish. National Acad. Press, Washington DC. Olsen R. E., J. Suontana, E. Langmyhr, H. Mundheim, E. Ringo, W. Melle, M. K. Malde and G. I. Hemre, 006. The replacement of fish meal with Antarctic krill, Euphausia superba in diets for Atlantic salmon, Salmo salar. Aquaculture Nutrition,, Park, S. H., 000. Studies to develop low pollute diet for juvenile olive flounder, Paralichthys olivaceus reared in the recirculating system. Master course dissertation, Pukyong National University, Busan, Korea. Rychly, J. and L. Spannhof, 979. Nitrogen balance in trout. I. Digesitibility of diets containing varying levels of protein and carbohydrate. Aquaculture, 6, Sugiura, S. H., F. M. Dong, C. K. Rathbone, and R. W. Hardy, 998. Apparent protein digestibility and mineral availabilities in various feed ingredients for salmonid feeds. Aquaculture, 59, Sullivan, J. A., 993. Apparent digestibility coefficients for common feedstuff in diets for original-cross hybrid striped bass Morone saxatilis Morone chrysops. M.S. Thesis, Louisiana State University, Baton Rouge, LA, 49 pp. Teshima, S. I., A. Kanazawa, S. Koshio, S. Itoh, 993. L-ascorbyl- phosphate-mg as vitamin C source for the Japanese flounder (Paralichthys olivaceus). Fish nutrition in practice., Institut national dela recherche agronomique, Paris (France), š : :