Teknologi produksi fikosianin Spirulina platensis dan pemanfaatannya sebagai imunostimulan pada ikan kerapu bebek (Cromileptes altivelis)
Production technology and utilization of phycocyanin of spirulina platensis as an immunostimulant in humpback grouper (Cromileptes altivelis)
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Date
2013Author
Satyantini, Woro Hastuti
Sukenda
Harris, Enang
Bambang, Nur
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Ministry of Marine Affairs and Fisheries Republic of Indonesia has targeted the grouper production as much as 9,000 tonnes of grouper in 2011 from 7,000 tonnes in 2010. Problems arise with the more intensive cultivation of grouper that lead to failure of production is precense of disease attack. Vibriosis is fish disease that can lead to failure in the production of grouper. The use of chemotherapeutics and antibiotics for prophylaxis have been prohibited by government because of negative impact to the environment and bacteria resistant. Giving an immunostimulant from natural plants microalgae is one way that can be used to resolve the problem reducing the failure rate due to this infectious disease. Phycocyanin bioactive and water soluble polysaccharide of Spirulina may enhance biological defence activity against infectious diseases through enhancement of proliferation of erythrocyte, granulocyte-monocyte that comes from bone marrow. This study aims to produce phycocyanin from Spirulina platensis and assess its utilization as an immunostimulant to improve the immune system of grouper. This research divided into three steps of study: 1) Production technology, extraction and phycocyanin characterization of Spirulina platensis, 2) Determination of the best dose of phycocyanin administration, and 3) Term of phycocyanin administration. The first research was to determine the combination of technical nutrient and the optimum light intensity for the best population growth and phycocyanin content, and used of the best phycocyanin extraction solvent. The research divided into two steps of experiment; first experiment used Kabinawa modification (KT) and technical modification (MT) nutrients with different of light intensity (2000, 3000 and 4000 lux). The second experiment utilized different phycocyanin extraction solvent water, 0.1 M Na phosphate buffer and 1% CaCl2. Each treatment was triplicates. Data showed the best population growth was gained on day 12th in the combination of MT nutrients and 3000 lux light intensity (MT3) was 0.611 OD. The highest phycocyanin content obtained at the exponential stage of KT2 treatment at 12th but not significantly different (p>0.05) from the treatment of MT3 and KT4. The phycocyanin extract solvent of 0.1 M Na phosphate buffer resulted in yield and highest purity levels of phycocyanin compared to the other two solvents. Protein content and molecular weight of two fractions protein phycocyanin result of SDS-PAGE were 26.64%, 19.23 and 63.32 kDa, respectively. This research showed that for the mass production of Spirulina can be use the combination of technical nutrient MT with 3000 lux light intensity and 0.1 M Na phosphate buffer is the best solvent for phycocyanin extraction as an immunostimulant. The second research was to examine the effect of administration of Spirulina phycocyanin on growth, enhancement of non-specific immune responses and disease resistance of humpback grouper, Cromileptes altivelis juveniles to Vibrio alginolyticus infection. Initial body weight of grouper was 8.46±0.22 g with the density of 10 fish per container (56 L). The humpback grouper was fed with the addition of phycocyanin at 0, 150, 250, 350 and 450 mg kg-1 diet and reared for 28 days. Challenge test performed by infecting V. alginolyticus bacteria with dose 3.26x108 cfu mL-1, via intraperitoneal injection of 0.1 mL/fish. Each treatment was triplicates. The highest of final weight (Wt=14.32 g), specific growth rate (SGR=1.89%), and lowest of feed conversion ratio (FCR=1.13) was gained in fish addition at 250 mg phycocyanin kg-1 diet and significantly different (P <0.05) to the control. Total leucocytes, haemoglobin, hematocrit, phagocytic activity, lysozyme activity, serum protein, albumin and globulin increased in phycocyanin supplemented in diet. Administration of phycocyanin 250 mg kg-1 diet resulted in higher total leucocytes, haemoglobin, phagocytic activity, lysozyme activity, blood serum protein, albumin and globulin were 8.49x105 cells mm-3, 5.7 g%, 59.67%, 0.97 OD, 2.4 g dL-1,0.98 g dL-1 and 1.42 g dL-1 respectively compared to the control. The highest Relative Percent Survival (RPS) value was achieved at addition of phycocyanin 250 mg kg-1 diet was 81.83%. Dietary Spirulina phycocyanin at 250 mg kg-1 diet enhances growth, non-specific immune responses and diseases resistance in humpback grouper juvenile against V. alginolyticus infection. The third experiment was to determine different term of administration on non-specific immune responses and resistance of humpback grouper, Cromileptes altivelis juveniles to V. alginolyticus bacteria infection. Fish with initial body weight of 5.24 ± 0.48 g were reared with a density 15 fish per vessel (60 L). Fish were divided in seven groups, five groups of fish fed with the addition of phycocyanin 250 mg kg-1 diet for 7 days; 7 days intermittent; 14 days; 21 days; 28 days and the remaining group was fed the basal feed (control) for 28 days. After 28 days rearing, fish are challenged with V. alginolyticus via intraperitoneal injected with dose 4.02x108 cfu mL-1 of 0.1 mL/fish. Total erythrocytes, leukocytes, haemoglobin, haematocrit, phagocytic activity, lysozyme activity was higher in the treatment with the addition of phycoacyanin for 14 days were 12.90x105 cells mm-3, 5.84x105 cells mm-3, 4.40g%, 12.41%, 58.0%, and 1.05 OD respectively and significantly different (P <0.05) to the control. The highest RPS (Relative Percent Survival) was achieved at addition 250 mg phycocyanin kg-1 diet for 14 days consecutive was 85.71%. This result suggested that the administration of Spirulina phycocyanin at 250 mg kg-1 diet for 14 days enhancement of non-specific immune response, and disease resistance in humpback grouper C. altivelis juvenile against V. alginolyticus. In summary it can be concluded that the production of phycocyanin can be done by culturing Spirulina platensis using technical nutrient MT and the phycocyanin extract can be applied as an immunostimulant. The administration phycocyanin at 250 mg kg-1 diet increase the growth and non-specific immune response of humpback grouper juvenile. The application of phycocyanin administration at 250 mg kg-1 diet for the 14 consecutive days was effectively and efficiently increasing the production of humpback grouper juvenile.
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