Purification and characterisation of antioxidant and nitric oxide inhibitory peptides from Tilapia (Oreochromis niloticus) protein hydrolysate

Nitric oxide (NO)‐inhibitory and antioxidative activities of tilapia hydrolysates were prepared using ultrasound pretreatment at 70 W for 30 and 45 min, respectively, followed by Flavourzyme hydrolysis for 1 h. Both hydrolysates were fractionated using size exclusion chromatography on Sephadex G‐25 column and purified by RP‐HPLC. The amino acid sequence of the most potent and purified fractions was determined using LC/MS/MS. The antioxidant peptide (KAFAVIDQDKSGFIEEDELKLFLQNFSAGARAGDSDGDGKIGVDEFAALVK, MW: 6334.49 KDa) and NO‐inhibitory peptide (AFAVIDQDKSGFIEEDELKLFLQNFSAGARAGDSDGDGKIGVDEFAALVK, MW: 6309.49 Da) produced no cytotoxicity in RAW264.7 macrophage cell lines at the concentration of 100 mg mL^?1. The purified peptides at the concentration 100 μg mL^?1 possessed antioxidative and NO‐inhibitory activities 83.0 ± 1.1% and 40.9 ± 0.2%, respectively, which were about 100 times those of their counterpart crude hydrolysates.     

Feasibility of fishmeal replacement by shrimp head silage protein hydrolysate in Nile tilapia (Oreochromis niloticus L) diets

This study provides information on the use of shrimp head silage protein hydrolysate (SPH) as an alternative protein source for tilapia feeding. Six diets (28% protein, 12% lipid) were prepared where fishmeal protein was replaced at levels of 0 (control), 10, 15, 20, 25 and 30% with the hydrolysate. The diets were supplied to Nile tilapia fry (338 mg initial weight) stocked in plastic recirculating 20 l tanks (10 animals per tank), with three replicates per treatment. After an 8 week experimental period, fish fed the diets containing 10 and 15% SPH showed significantly better performance in terms of final body weight, weight gain (%), mean daily weight gain (mg day^?1), specific growth ratio and feed conversion ratio than those fed the control diet (fishmeal as protein source) and higher‐SPH diets. It is concluded that shrimp head hydrolysate is a promising alternative protein source for tilapia feeding, improving growth ratio at dietary inclusion levels as high as 15%. In addition, the diets with added shrimp silage protein were well accepted by the fish, which avidly consumed the feed during the experiment. © 2002 Society of Chemical Industry     

Antiphotoaging effect and purification of an antioxidant peptide from tilapia (Oreochromis niloticus) gelatin peptides

In this study, the effect of tilapia gelatin peptides (TGP) on UV-induced damages to mice skin was evaluated. The antioxidant indicators, lipid, collagen and glycosaminoglycan of mice skin were determined and histological changes of the collagen were depicted. The results showed TGP could alleviate the UV-induced abnormal changes of antioxidant indicators, and the protective effect was in dose-dependent manners. TGP could protect skin lipid and collagen from the UV radiation damages, and the change of glycosaminoglycan was also recovered significantly. The action mechanisms of TGP mainly involved the antioxidant properties and the repairing to endogenous collagen synthesis. Therefore, the key antioxidant peptide was further purified from TGP. Finally, one antioxidant peptide was identified and the amino acid sequence was Leu-Ser-Gly-Tyr-Gly-Pro (592.26 Da). The IC₅₀ value of this peptide on hydroxyl radical scavenging activity was 22.47 μg/mL. TGP could be a novel antiphotoaging agent from natural resources.     

罗非鱼鱼肉及组分蛋白酶解产物的抗氧化特性

采用胃蛋白酶、中性蛋白酶、Alcalase2.4L碱性蛋白酶分别酶解罗非鱼肉蛋白,研究比较不同蛋白酶在1、2、4、6、8h对罗非鱼肉蛋白的酶解效果及产物的抗氧化活性,结果表明:3种酶的酶解产物都具有一定的抗氧化活性,中性蛋白酶酶解产物的水解度和蛋白回收率最高,胃蛋白酶酶解产物对DPPH自由基和羟自由基的清除效果最好;综合各项指标选择中性蛋白酶对罗非鱼肉的组分蛋白(肌原纤维蛋白、肌浆蛋白、基质蛋白)进行4h酶解,研究比较不同组分蛋白的酶解效果及产物的抗氧化活性,结果表明:组分蛋白中基质蛋白酶解产物的蛋白回收率最高,为89.8%,肌原纤维蛋白酶解产物的水解度最高,为10.1%,肌浆蛋白酶解产物的抗氧化活性最好,羟自由基和DPPH自由基的IC50值分别为12.352mg/m L和3.554mg/m L。      

Preparation of reactive oxygen scavenging peptides from tilapia (Oreochromis niloticus) skin gelatin: optimization using response surface methodology

Abstract: Gelatin extracted from tilapia skin was hydrolyzed with Properase E. Response surface methodology (RSM) was applied to optimize the hydrolysis condition (temperature [T], enzyme‐to‐substrate ratio [E/S], pH and reaction time [t]), to obtain the hydrolysate with the highest hydroxyl radical (?OH) scavenging activity. The optimum conditions obtained were T of 44.2 °C, E/S of 2.2%, pH of 9.2, and t of 3.4 h. The predicted ?OH scavenging activity of the hydrolysate under the optimum conditions was 60.7%, and the actually experimental scavenging activity was 60.8%. The hydrolysate was fractionated by ultrafiltration, and 4 fractions were collected. The fraction TSGH4 (MW < 2000 Da) showed the strongest ?OH scavenging activity with the highest yield. Furthermore, reactive oxygen species (ROS) scavenging activities of TSGH4 with different concentrations were investigated in 5 model systems, including superoxide anion radical (?O₂), ?OH, hydrogen peroxide (H₂O₂), peroxynitrite (ONOO^?), and nitric oxide (NO?), compared with reduced glutathione (GSH). The results showed that TSGH4 significantly scavenged these ROS, and could be used as a functional ingredient in medicine and food industries.     

Selenium fractionation from plasma, muscle and liver of Nile tilapia (Oreochromis niloticus)

This paper presents the results obtained from selenium fractionation in plasma, muscle and liver samples of Nile tilapia’s (Oreochromis niloticus) after protein separation. The plasma, muscle and liver proteome was obtained by 2D-PAGE, and selenium in protein spots was qualitatively and quantitatively determined by synchrotron radiation X-ray fluorescence and graphite furnace atomic absorption spectrometry (GFAAS). The fluorescence spectra indicated the presence of selenium in three protein spots of plasma, two of muscle and one of liver. Selenium was found to be distributed mainly in proteins with a molar mass smaller than 57.0 kDa and with pI in the range of 5.9–9.6, with one exception in the plasma sample, which presented protein with a molar mass of 60.0 kDa. After acid mineralization of the protein spots, a GFAAS determination of the concentration of selenium bound to these proteins indicated a range of 1.35–6.82 mg per g of protein.     

Antioxidant and nitric oxide inhibitory activities of tilapia (Oreochromis niloticus) protein hydrolysate: effect of ultrasonic pretreatment and ultrasonic‐assisted enzymatic hydrolysis

Ultrasound was incorporated to processing of fish protein hydrolysate to facilitate homogenate pretreatment and enzymatic hydrolysis of tilapia (Oreochromis niloticus) muscle protein. Their effects on Flavourzyme hydrolysis and biological activities of the tilapia hydrolysate were examined. The ultrasound‐assisted hydrolysis caused reduction in degree of hydrolysis ranging from 23% to 35% relative to that of the conventional process. The 70 W ultrasound‐assisted hydrolysis process increased DPPH radical‐scavenging activity and reducing power of tilapia hydrolysate prepared from the non‐pretreatment homogenate by 33% and 45%, respectively. All hydrolysates have no cytotoxicity on RAW264.7 cell lines at the maximum concentration of 20 mg protein mL^?1. The 70 W ultrasound pretreatment at 30 and 45 min combined with conventional hydrolysis is the suitable condition for producing tilapia hydrolysate with nitric oxide inhibitory and antioxidative activities on RAW264.7 cell lines, respectively. As a result, ultrasound could be applied to enzymatic protein hydrolysis either as pretreatment or during the hydrolysis.     

利用罗非鱼卵黄原蛋白建立雌激素污染的检测方法

目的以尼罗罗非鱼为实验对象,以卵黄原蛋白(Vg)为生物标志物开发环境雌激素的生物检测技术。方法采用凝胶过滤与离子交换层析相结合的方法从17β-雌二醇诱导后的罗非鱼血浆中分离纯化Vg,对纯化的蛋白进行鉴定后制备多克隆抗血清,建立Vg的酶联免疫吸附测定实验(ELISA),并用于样品测定。结果建立的ELISA工作范围为15.6~1000 ng/m L,组内与组间差异分别为6.85%与6.79%,能有效检测100、500、1000μg/L双酚A对雄鱼血浆Vg的诱导效应。结论本研究建立的Vg ELISA具有较高的敏感度、特异性与精确度,可为水体环境雌激素污染检测提供重要工具。     

Nutritional quality of spray dried protein hydrolysate from Black Tilapia (Oreochromis mossambicus)

The nutritional quality of spray-dried protein hydrolysate from black tilapia, a fresh water fish, was evaluated. Hydrolysed protein from Oreochromis mossambicus was spray-dried at two different temperatures of 150 °C/76 °C (inlet/outlet temp) and 180 °C/90 °C. Proximate analyses revealed that the dried hydrolysates consisted of 37.7–49.6% protein, 2.6–2.8% fat, 1.6–4.0% moisture and 8.6–8.7% ash. The higher drying temperature used was found to significantly decrease the contents of all amino acids analysed. Nevertheless, the protein quality of both dried hydrolysates was found to be high, with in vitro digestibilities of 88.4 and 92% and protein digestibility corrected amino acid scores of 0.34 and 0.82, respectively. In addition, the predicted protein efficiency ratios of the dried hydrolysates were calculated to be 2.97 and 2.53.     

Original article: Optimisation of extraction conditions and characteristics of skin gelatin from Nile tilapia (Oreochromis niloticus)

Response surface methodology was used to optimise gelatin extraction conditions from the skin of Nile tilapia (Oreochromis niloticus), and characteristics of the gelatin were determined. Concentration of NaOH (%, X₁), alkaline treatment time (h, X₂), concentration of HCl (‰, X₃) and acid treatment time (min, X₄) were chosen for independent variables. Dependent variable was yield of gelatin (%, Y). Optimal conditions were X₁ = 3.2 (%), X₂ = 2.3 (h), X₃ = 0.7 (‰) and X₄ = 84 (min), and predicted value of response optimal conditions was Y = 20.4%. Actual value was 19.3% by verification experiments under optimal conditions. Crude protein content of the tilapia skin gelatin was 88.5%. The content of imino acids including proline and hydroxyproline in the gelatin was 185 residues per 1000 total amino acid residues. Its gel strength was 260 g. The gelling and melting points were 18.0 and 22.4 °C, respectively.     

罗非鱼皮明胶酶解物及其模拟消化产物的抗氧化活性

以罗非鱼皮为原料提取罗非鱼皮明胶,选用风味蛋白酶和胰蛋白酶制备罗非鱼皮明胶酶解物,采用ABTS自由基、DPPH自由基、羟基自由基及亚油酸过氧化体系,初步评价罗非鱼皮明胶酶解物的抗氧化活性,再通过模拟体外胃肠道消化实验,结合分子质量分布测定,进一步考察罗非鱼皮明胶酶解物的抗氧化活性。结果显示,在酶解过程中,风味蛋白酶及胰蛋白酶酶解的水解度逐渐升高,在3 h时达到最高,分别达到5.8%和25.36%。在酶解60 min时其TCA可溶性肽得率最高,风味蛋白酶、胰蛋白酶酶解物分别达56.82%和54.44%。通过比较半抑制浓度（IC₅0）,确定了酶解60 min时风味蛋白酶酶解物的清除DPPH自由基及抑制亚油酸过氧化能力较胰蛋白酶酶解物强。模拟体外胃肠道消化后,酶解物羟基自由基清除活性均显著提高（p<0.05）,亚油酸脂质过氧化活性明显降低,消化前后样品分子量分布范围均主要集中于3000⁵000 Da,消化后风味蛋白酶及胰蛋白酶酶解物3000⁵000 Da组分的含量分别提高了45%及13%。以上研究结果表明,罗非鱼皮明胶酶解后制备的明胶水解物具有一定的抗氧化能力,具有潜在的开发价值。      

Effect of starvation and diet on the gel forming ability of tilapia (Oreochromis niloticus)

To elucidate the factors affecting the variation in the gel forming ability of fish muscle within a species, the effect of starvation and diet on the proximate composition and gelling property of tilapia muscle was examined. Starvation resulted in a decrease in protein and lipid contents coupled with an increase in water content. Protein depletion, mainly in the myofibrillar fraction, in the starved fish caused a lowering of the gel forming ability of their muscle. The myosin-degrading activity at 65°C was not affected by starvation for 56 days. Tilapia (Oreochromis niloticus L) fed on either fishmeal or krillmeal did not differ in their proximate composition. Phytoplankton-fed fish had a slightly higher lipid content and had the greatest gel forming ability. There was no significant difference in the gel strength between the fish fed fishmeal and the fish fed krill or 58 days. Compared with the phytoplankton, however, prolonged feeding of fishmeal seemed to increase the myosin-degrading activity of the muscle.     

Manipulation of fatty acid composition of Nile tilapia (Oreochromis niloticus) fillets with flaxseed oil

Nile tilapia (Oreochromis niloticus) is one of the most consumed species among freshwater fish reared in Brazil. However, studies show low levels of n‐3 fatty acids in freshwater fish reared in captivity in comparison with those reared in their natural habitats. The Nile tilapia used in this study were raised in captivity for a period of 5 months and fed varying amounts (0, 1.25, 2.5, 3.75 and 5%) of flaxseed oil as a substitute for sunflower oil (control). No significant differences (P > 0.05) in total lipid (TL) content were found between fillets of tilapia fed the different diets. TL analysis of fatty acid methyl esters by capillary gas chromatography revealed a total of 50 components common to all treatments studied. The major fatty acids present were linoleic acid (18:2n‐6), oleic acid (18:1n‐9) and palmitic acid (16:0). All treatments led to significant % increases in α‐linolenic acid (18:3n‐3), eicosapentaenoic acid (20:5n‐3) and docosahexaenoic acid (22:6n‐3). Increases in both total n‐3 fatty acids and total polyunsaturated fatty acids were observed concomitantly with decreases in total n‐6 fatty acids, resulting in increases in n‐3/n‐6 ratio, with increasing level of flaxseed oil in the feed. Thus feed supplementation with flaxseed oil contributed greatly to raising the nutritional lipid value of Nile tilapia fillets. Copyright © 2007 Society of Chemical Industry