Carnosine을 이용한 흰다리새우(Litopenaeus vannamei) 사료 내 histidine 요구량

Abstract

This study was conducted to evaluate the optimum dietary histidine requirement using two molecular forms (dipeptide or crystalline amino acid) of histidine and different levels of L-carnosine in diets for Pacific white shrimp (Litopenaeus vannamei) (Exp 1 and 2). In this study, growth performance, feed utilization, diet digestibility, non-specific immunity and qPCR analysis were examined. In the Exp-1, five experimental diets were prepared. The control (Con) diet was prepared without histidine and other diets were with 0.3 and 1.2% addition of histidine in the form of crystalline AA or dipeptides (Con, C-0.3, C-1.2, D-0.3 and D-1.2). Experimental diets were kept isonitrogenous and isocaloric by decreasing alanine while increasing histidine levels. Pacific white shrimp (body weight: 0.15 g) were randomly stocked into twenty tanks (96 L) in quadruplicate groups per dietary treatment and fed one of the diets for 62 days. Final mean body weight, weight gain and specific growth rate were significantly higher in shrimp fed D-0.3 and D-1.2 than those of shrimp fed the Con diet. No significant differences were observed in feed utilization and survivals by experimental diets. Analysis of whole body composition and amino acid profile were not significantly different among all the groups. Apparent digestibility cofficiency of dry matter was significantly higher in shrimp fed C-1.2 and D-1.2 than those of shrimp fed the Con. Apparent digestibility cofficiency of protein was significantly higher in shrimp fed C-1.2, D-0.3 and D-1.2 than those of shrimp fed the Con. The relative gene expression of IGF-BP mRNA was significantly higher in shrimp fed D-1.2 than those of shrimp fed the Con. In the Exp-2, a control diet contained a minimum level of His (0.05% in diet from the dietary ingredients). Carnosine was gradually added to the control diet to contain 0, 0.26, 0.69, 1.11, 1.53, 1.95 or 2.37% histidine (designated as 0.05, 0.24, 0.40, 0.62, 0.78, 0.93 or 1.13, respectively). Experimental diets were kept isonitrogenous and isocaloric by decreasing alanine while increasing carnosine levels. Pacific white shrimp (body weight: 0.28 g) were randomly stocked into twenty-eight tanks (110 L) in quadruplicate groups per dietary treatment and fed one of the diets for 8 weeks. Final mean body weight, weight gain and specific growth rate were significantly higher in shrimp fed 0.78 and 0.93 than those of shrimp fed the Con. No significant differences were observed in feed utilization and survivals by the gradual supplementation of carnosine in diets. Phenoloxidase activity was significantly higher in shrimp fed all groups than those of shrimp fed the Con. Lysozyme activity was significantly higher in shrimp fed 0.62 and 0.78 than those of shrimp fed the Con. Anti-protease activity was significantly higher in shrimp fed 0.24, 0.4, 0.62, 0.78 and 0.93 than those of shrimp fed the Con. Total protein concentration in hemolymph was significantly higher in shrimp fed 0.78 than those of shrimp fed the Con. Muscle composition protein content of shrimp was significantly higher in shrimp fed 0.24, 0.62 and 0.78 than those of shrimp fed the Con. Prophenoloxidase (proPO) and mRNA relative gene expression of crustin was most significantly higher in shrimp fed 0.78 and 0.93 than those of shrimp fed the Con. The quadratic regression analysis indicated that the optimal dietary histidine requirement for L. vannamei is 0.79% of dry diet. In conclusion of Exp-1, the availability of histidine could be better in Pacific white shrimp when they are fed with dipeptides form rather than crystalline form. Dipeptides can be used as promising amino acid source for amino acid requirement study in Pacific white shrimp. In conclusion of Exp-2, the result from the present study indicated that dietary histidine improved growth performance and innate immunity. Based on a quadratic regression of specific growth rate, the optimal dietary histidine requirement by carnosine for Pacific white shrimp was estimated to be 0.79% in diet (2.19% of dietary protein). This information will be useful in developing amino-acid balanced diets for the intensive culture of Pacific white shrimp.

본 연구는 흰다리새우 사료의 필수아미노산 중 하나인 histidine의 형태(dipeptides or crystalline)에 따라 이용성을 비교 검토하여 정확한 필수아미노산 요구량을 조사하기 위해 총 2개의 사양 실험을 실시하였다. 실험-1 의 실험사료는 histidine이 첨가되지 않은 아미노산 mixture를 주 단백질 사료원료로 사용하였다. Histidine을 첨가하지 않은 대조구(Con)와 crystalline, dipeptides 형태로 histidine이 각각 0.3, 1.2%(C-0.3, C-1.2, D-0.3, D-1.2) 첨가된 실험구로 총 5개의 실험사료를 제작하였다. 실험새우는 초기평균 무게가 0.15 g인 흰다리새우(Pacific white shrimp, L. vannamei)를 이용하여 각각 15마리씩 총 20개의 96 L 수조에 무작위로 배치하였다. 사육실험이 종료 된 후 성장지표 측정을 위하여 무게를 측정했으며, 실험새우의 수조 당 3마리(시험구 당 12마리)의 새우를 무작위로 선별 후 간췌장을 분리하여 IGF-BP 분석을 진행하였다. 62 일간의 사양실험 결과, 성장률은 dipeptides 형태로 첨가된 모든 실험구가 대조구에 비해 유의적으로 높았다. 유전자발현 분석에서 IGF-BP 호르몬은 D-1.2실험구가 대조구에 비해 유의적으로 높게 발현되었다. 실험-2 의 실험사료는 histidine이 첨가되지 않은 아미노산 mixture를 주 단백질 사료원료로 사용하였으며, 기호성을 고려하여 오징어간분말을 최소량으로 첨가해주었다. Histidine을 첨가하지 않은 대조구(0.05)와 dipeptides 형태의 histidine을 각 0, 0.26, 0.69, 1.11, 1.53, 1.95, 2.37%(0.05, 0.24, 0.40, 0.62, 0.78, 0.93 and 1.13) 첨가된 실험구로 총 7개의 실험사료를 제작하였다. 실험새우는 초기평균 무게가 0.28 g인 흰다리새우(Pacific white shrimp, L. vannamei)를 이용하여 각각 25마리씩 총 28개의 110 L 수조에 무작위로 배치하였다. 사육실험이 종료 된 후 성장지표 측정을 위하여 무게를 측정했으며, 실험새우의 수조 당 5마리(시험구 당 20 마리)의 새우를 무작위로 선별 후 hemolymph 채혈 후, 간췌장을 분리하여 비특이적 면역과 qPCR (proPO, crustin)유전자발현 분석을 진행하였다. 8 주간의 사양실험 결과 성장률은 0.78, 0.93실험구가 대조구에 비해 유의적으로 높았다. 흰다리새우 사료 내 histidine 적정첨가 함량은 histidine의 농도와 일간성장률 간의 관계를 기준으로 하여 quadratic regression 분석을 통해 0.79% 인 것으로 나타났다. 결론적으로 흰다리새우는 crystalline AA 보다 dipeptide를 보다 효율적으로 이용하여 성장률을 높일 수 있을 것으로 판단된다. Dipeptide는 흰다리새우의 AA 요구량 연구에 있어서 crystalline AA을 대체하여 보다 더 정확한 요구량을 알아보기 위한 아미노산 원료로 사용 가능할 것으로 판단된다. Dipeptide형태인 L-carnosine을 이용하여 흰다리새우의 histidine 요구량을 연구한 결과, 흰다리새우에 있어서 배합사료 내 histidine의 적정첨가 함량은 0.79%로 판단된다. 흰다리새우의 histidine 연구는 아직 보고된바가 없어 본 연구에서 분석된 흰다리새우의 histidine 요구량은 사료 배합에 중요한 기초자료로 이용될 것으로 사료된다.