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한수지 48(4), 454-458, 2015 Original Article Korean J Fish Aquat Sci 48(4),454-458,2015 저수온기 (13-15 ) 치어기참돔 (Pagrus major) 사료내비타민 C 대체제로써의감귤착즙박및발효감귤착즙박의이용가능성 이초롱 김유정 이경준 * 제주대학교해양생명과학과 Dietary Supplementation of Citrus and Fermented Citrus By-product for Juvenile Red Seabream Pagrus major at Low Water Temperature Chorong Lee, Youjeong Kim and Kyeong-Jun Lee* Department of Marine Life Sciences, Jeju National University, Jeju 63243, Korea This study compared the effects of dietary supplementation of citrus by-product (CBP) and CBP fermented with Bacillus subtilis (F-CBP) on growth performance, feed utilization, intestinal histology and innate immunity of red seabream Pagrus major with three commercial immune-boosting products. The six experimental diets were supplemented with L-ascorbyl-2-polyphosphate (LAPP; the control diet), CBP or F-CBP at a concentration of 100 mg vitamin C equivalent/kg diet or one of three commercial immune boosters. The Experimental diets were fed to triplicate groups of 17 fish (initial body weight, 116 g) for 8 weeks. The water temperature during the feeding trial was maintained at 13-15. Growth and feed utilization did not differ significantly among the six dietary treatments, nor did the phagocytic activity, superoxide dismutase or total immunoglobulin concentrations. However, myeloperoxidase activity was significantly higher in the CBP groups. For the intestinal histology, the intestine diameter, villi and enterocyte heights and number of goblet cells did not differ significantly among groups. Therefore, CBP or F-CBP can be used as a valuable eco-friendly byproduct in diets for fishes including red seabream to maintain their normal growth and health. Key words: Citrus byproduct, Fermented citrus byproduct, Immunity, Red seabream, Vitamin C 서론 C (L-ascorbic acid, AA) (Lin and Shiau, 2005). C,, (Soliman et al., 1987), C (Lim et al., 2000). (Citrus byproduct, CBP),. 75 15, 50%,. CBP C (>50 mg AA/100 g), carotenoids (500 mg carotenoid/kg), bioflavonoids pectin (Rural development administration, 2012), (Kim et al., 1999). CBP C 60%., http://dx.doi.org/10.5657/kfas.2015.0454 Korean J Fish Aquat Sci 48(4) 454-458, August 2015 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial Licens (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Received 8 June 2015; Revised 13 August 2015; Accepted 17 August 2015 *Corresponding author: Tel: +82. 64. 754. 3423 Fax: +82. 64. 756. 3493 E-mail address: kjlee@jejunu.ac.kr Copyright 2015 The Korean Society of Fisheries and Aquatic Science 454 pissn:0374-8111, eissn:2287-8815

참돔사료내감귤착즙박및발효감귤착즙박의이용가능성 455 (Khosravi et al., 2015). CBP (Lee et al., 2013) C CBP. (Ding et al., 2015). isoflavone, isoflavone (Lee et al., 2005). (Lim and Lee, 2011). C. C,,. 감귤착즙박및실험사료 재료및방법 Brown Fish (Prime grade, Chile),,. ( ). Bacillus subtilis. B. subtilis Nutrient borth (NB, Difco, USA) 30 3 1.0 10 3 CFU/mL. 2.5% B. subtilis 30 72 (fermented CBP, F- CBP). 24. C 3,400 3,000 g AA/g diet, C 100 g AA/g diet.. (Table 1) C, CBP, F- CBP, A, B C Table 1. Formulation and proximate composition of experimental diets for red seabream Pagrus major (% dry matter) Ingredient Brown fish Corn gluten Soybean LAPP CBP F-CBP Experiment diets COM-A COM-B COM-C 45.0 45.0 45.0 45.0 45.0 45.0 8.00 8.00 8.00 8.00 8.00 8.00 9.00 9.00 9.00 9.00 9.00 9.00 Wheat flour 26.0 23.1 22.7 25.9 25.5 25.7 Squid liver oil 10.0 10.0 10.0 10.0 10.0 10.0 Vitamin Mix 1 1.00 1.00 1.00 1.00 1.00 1.00 Mineral Mix 2 1.00 1.00 1.00 1.00 1.00 1.00 LAPP 3 0.0285 0.00 0.00 0.00 0.00 0.00 CBP 4 0.00 2.90 0.00 0.00 0.00 0.00 F-CBP 5 0.00 0.00 3.30 0.00 0.00 0.00 COM-A 6 0.00 0.00 0.00 0.05 0.00 0.00 COM-B 7 0.00 0.00 0.00 0.00 0.50 0.00 COM-C 8 0.00 0.00 0.00 0.00 0.00 0.30 Dietary nutrient composition (%) Protein 46.9 46.8 46.5 46.9 46.8 46.9 Lipid 14.7 14.8 14.7 14.7 14.7 14.7 Ash 9.60 9.80 9.30 9.60 9.60 9.60 Energy, MJ/kg diet 17.9 17.9 18.0 17.9 17.9 17.9 Total ascorbic acid 115 105 112 21.4 20.9 123 (mg/kg) 9 1 Vitamin Premix (g/kg of mixture): DL-tocopheryl acetate, 20.0; thiamin hydrochloride, 4.0; riboflavin, 4.4; pyridoxine hydrochloride, 4.0; niacin, 30.0; D-pantothenic acid hemicalcium salt, 14.5; myo-inositol, 40.0; D-biotin, 0.2; folic acid, 0.48; menadion, 0.2; retinyl acetate, 1.0; cholecalficerol, 0.05; cyanocobalamin, 0.01. 2 Mineral Premix (g/kg of mixture): MgSO 4.7H 2 0, 80.0; NaH 2 PO 4.2H 2 O, 370.0; KCL, 130.0; Ferric citrate, 40.0; ZnSO 4.7H 2 0, 20.0; Ca-lactate, 356.5; CuCl 2, 0.2; AlCl 3.6H 2 O, 0.15; Na 2 SeO 3, 0.01; MnSO 4. H 2 O, 2.0; CoCl 2.6H 2 O, 1.0. 3 L-ascorbic acid: Sigma Chemicals (350,000 ppm) 4 Citrus by-product: ILHAE Corporation 5 Fermented CBP with Bacillus subtilis 6 CTCbio corporation 7 Samjo life science corporation 8 Changjobio corporation 9 Total ascorbic acid (TAA) was determined by dinitrophenylhydrazine spectrophotometric method described Dabrowski and Hinterleitner (1989).

456 이초롱ㆍ김유정ㆍ이경준 6 ( LAPP, CBP, FCBP, COM-A, COM-B and COM-C ). 실험어및사육관리. 3. ( : 116 2.28 g) 18 150 L 17. 2-3 L/min. (13-15 ), 1 2 (08:00 17:00) 8. 어체측정및분석 3, 24.,,,,., 8 2-phenoxyethanol (200 ppm) 6 3. (nitrobluetetrazolium, NBT). 3, 60 (5,000 g). AOAC (1995), C Dabrowsik and Hinterleitner (1989). (Nitro-blue tetrazoium, NBT) myeloperoxidase (MPO) Kumari and Sahoo (2005). superoxide dismutase (SOD) SOD assay kit (Sigma, 19160). total immunoglobulin Siwicki and Anderson (1993). (Completely randomized design), SPSS (Version 12.0) One-way ANOVA. Tukey (HSD). (mean SD). arcsine. 결과및고찰 8,,,,,, (Table 2). C. 9 (Song et al., 2013). (13-15 )., MPO (Table 3). MPO,,,. MPO hydrogen peroxide (H 2 O 2 ) chloride anion Table 2. Growth performance of red seabream (Pagrus major, initial BW: 116 g) fed experimental diets for 8 weeks FBW 1 (g) SGR 2 (%) FCR 3 FI 4 PER 5 Survival (%) LAPP 148±2.10 0.49±0.01 1.14±0.02 39.9±2.45 1.87±0.04 98.0±3.40 CBP 152±6.99 0.49±0.08 1.17±0.18 41.7±2.87 1.86±0.30 88.2±20.9 F-CBP 152±8.16 0.48±0.08 1.30±0.30 44.2±1.73 1.71±0.35 96.1±6.79 COM-A 159±6.88 0.54±0.04 1.06±0.06 45.4±1.02 2.01±0.11 92.2±6.79 COM-B 158±0.98 0.51±0.05 1.16±0.15 45.1±1.05 1.87±0.24 98.0±3.40 COM-C 160±3.23 0.54±0.03 1.02±0.05 43.4±3.30 2.09±0.11 100±0.00 Values are mean of triplicate groups and presented as mean±sd. Values in the same column having different superscript letters are significantly different (P<0.05). 1 FBW=Final body weight 2 Specific growth rate = [(log e final body weight - log e initial body weight)/days] 100 3 Feed conversion ratio = dry feed fed/wet weight gain 4 Feed intake (g/fish) = dry feed consumed (g)/ fish 5 Protein efficiency ratio = fish weight gain (g)/ protein

참돔사료내감귤착즙박및발효감귤착즙박의이용가능성 457 Table 3. Nitro blue tetrazolium (NBT), myeloperoxidase (MPO), superoxide dismutase (SOD) and immunoglobulin (Ig) of fed red seabream Pagrus major six experimental diets for 8 weeks NBT (OD) MPO SOD Ig LAPP 0.87±0.17 1.17±0.08 ab 32.5±4.71 21.9±2.64 CBP 0.87±0.14 1.43±0.23 b 38.0±7.59 18.9±2.19 F-CBP 0.88±0.06 1.19±0.08 ab 37.7±8.07 25.4±3.50 COM-A 0.89±0.07 0.97±0.03 a 28.9±1.24 21.8±1.66 COM-B 0.88±0.06 1.14±0.18 ab 40.2±5.00 23.8±6.45 COM-C 0.87±0.04 1.16±0.10 ab 37.4±1.34 24.4±1.75 Values are mean of triplicate groups and presented as mean ± SD. Values in the same column having different superscript letters are significantly different (P<0.05). (Cl - ) hypochlorous acid, hydrogen peroxide tyrosine tyrosyl radical. hyprochlorous acid tyrosyl radical, (Kumari and Sahoo, 2005). MPO hypochlorous acid tyrosyl radical., CBP COM-A MPO, CBP, F-CBP COM-A SOD. CBP, F-CBP C... 8 (goblet cell) (Fig. 1),, CBP F-CBP LAPP (data not provided).,,, (Allen et al., 1986). probiotics B. subtilis,. C,,, Fig. 1. Changes in number of goblet cells in the intestine of red seabream Pagrus major fed the six experimental diets for 8 weeks.. C. 사사 ( ) (NRF-2011-0015925). References Allen A, Hutton DA, Leonard AJ, Pearson JP and Sellers LA. 1986. The role of mucus in the protection of the gastroduodenal mucosa. Scand J Gastroenterol 21, 71-78. AOAC. 1995. Official methods of analysis. 16 th ed. Association of Official Analytical Chemists, Arlington, Virginia, U.S.A, 1298. Dabrowski K and Hinterleitner S. 1989. Applications of a simultaneous analysis of ascorbic acid, dehydroascorbic acid and

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