Autolysis of krill protein from North Pacific krill Euphausia pacifica during protein recovery via isoelectric solubilization/precipitation

Autolysis of North Pacific krill protein in acidic and alkaline solubilization during protein recovery via isoelectric solubilization/precipitation was investigated. Pronounced auto-degradation of myosin heavy chain and actin from krill protein was observed at alkaline pH values (pH 9.0–12.0), with maximum autolytic activity at about pH 12.0. Meanwhile, proteolytic activity in krill protein at pH 12.0 was observed at low temperatures, suggesting a possible cause for the autolysis of krill protein during the protein recovery process. Three major proteinases were detected by zymographic analysis of myofibrillar protein, with approximate molecular weights of 26 kDa (KP-1), 18 kDa (KP-2) and 17 kDa (KP-3). KP-3 was active over a pH range from pH 5.0–12.0, suggesting that it may be responsible for the autolysis of krill protein during alkaline-induced protein recovery. Further study on substrate specificity and inhibitory specificity of KP-3 showed that KP-3 is serine type proteinase.     

Preparation,Gel Electrophoresis Analysis,and Nutritional Evaluation of a Functional Krill Protein Concentrate with Low Fluoride Level from Antarctic krill (Euphausia superba)

A functional krill protein concentrate (FKPC) with low level of fluoride was prepared from frozen Antarctic krill (Euphausia superba) using isoelectric precipitation and phosphoric acid treatment. Variations in proteins and nutritional properties of the FKPC were analyzed. Gel electrophoresis analysis indicated that some variations in protein composition were introduced by autolysis and pH shift during the process of isoelectric precipitation and fluoride removal. The FKPC was composed of 81.01% ± 0.45% crude protein (dry basis) and 16.78% ± 0.26% total lipid (dry basis), exhibited excellent amino-acid composition in all the essential amino acids, exceeding the requirements of the FAO/WHO/UNU for food protein, and was rich in omega-3 polyunsaturated fatty acids, especially eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids present in phospholipids. The fluoride level of FKPC was 79.79 ± 2.10 μg/g (dry basis), which met the USFDA requirements. The surimi yield reached 43.74%, and the mean recovery rate of protein and fat was 52.68% and 41.44%, respectively. The nutritional value, safe level of fluoride, and high yield rate provide evidence that FKPC has a promising future in the food industry as a new aquatic protein and lipid resource.     

Extracting Protein from Antarctic Krill (Euphausia superba)

ABSTRACT

The isoelectric solubilization/precipitation (ISP) process was applied to Antarctic krill protein extraction. The solubility of Antarctic krill proteins and their fluoride content were studied between pH 1.00–13.75. The previous krill protein extraction conditions using ISP were improved. Excess fluoride in Antarctic krill was removed using a water-washing method according to its solubility. To save water, a multistage countercurrent system was used to optimize the ISP method. The results showed that the solubility of Antarctic krill proteins reached a minimum at pH 4.5 and a maximum at pH 13.5, while the concentration of dissolved fluoride was relatively unchanged (16.78–20.50 mg/L), with pH 2.00–13.50. Water-washing 4 times could reduce fluoride 98.14 ± 0.01% from krill with about a 5.14 ± 0.21% protein loss. The reduction allows krill protein to meet the Chinese National Standard (GB 2762-2005) for fluoride. Meanwhile, the four-stage countercurrent system achieved almost the same high fluoride removal rate (98.58 ± 0.01%) using only 25% of the water.     

Amino acids composition and protein quality evaluation of marine species and meals for feed formulations in cephalopods

The amino acid composition and protein levels of three species of cephalopods (Octopus vulgaris, Loligo gahi and Todarodes sagittatus), the natural diets of common octopus (O. vulgaris) and different kinds of meals were determined in order to optimise the content of these nutrients in artificial feeds. Arginine, leucine and lysine were the most abundant essential amino acids in cephalopods, while glutamate and aspartate represented the main non-essential amino acids. Arginine and leucine were the limiting amino acid in most samples, with maximum Chemical Score values for mussel (79–98 %), squid (84 %) and crustaceans (65–91 %); medium for fish (41–70 %); and minimum for meals (29–64 %). Mussel, squid, crustaceans and fish showed a high essential amino acid index according to Oser (OI: 88–99 %) suggesting a suitable amino acid balance. The protein from animal meals (fish and krill) covered all the essential amino acids except arginine and lysine in fish meal. The vegetable meal presented the worst amino acid balance (OI: 74–89 %) with several deficiencies in essential amino acids, including arginine, threonine, lysine and methionine. Supplementation with arginine or leucine and protein complementation of crustaceans and bivalves with fish or animal meal are proposed as alternatives for improving the performance of protein in feed for cephalopods.     

Physicochemical and Functional Properties of Protein Recovered from Fish Waste

ABSTRACT

This work aimed to evaluate the physicochemical and functional properties of proteins recovered from whitemouth croaker (Micropogonias furnieri) and Argentine anchovy (Engraulis anchoita) residues obtained by the process of acid solubilization using HCl at pH = 2 and alkaline solubilization using NaOH at pH = 12 and isoelectric precipitation (pH = 5.5). The proximate composition and functional properties were determined. The solubilization and water holding capacity (WHC) of the recovered protein were studied in the pH range of 3, 5, 7, 9, and 11. The high protein content and lowest lipid reduction presented by the protein recovered from the whitemouth croaker residue and obtained by the acid solubilizing process was 97.1 and 98.4%, respectively. The maximum solubilized protein was at pH 3 and 11, and the minimum solubilized protein was at pH 5 for both species. The greater ability to retain oil was observed for the alkaline solubilization process (8.3 and 7.3). The lower WHC was at pH 5. The acid and alkali residue concentrates of the whitemouth croaker presented higher WHC at pH 3 and 11. These proteins can be used in preparing edible films and have been used to produce bioactive compounds for supplements or functional ingredients.     

正交实验优化南极磷虾蛋白肽的纳滤脱盐工艺

为建立南极磷虾(Euphausia superba)蛋白肽纳滤脱盐工艺,以脱盐率和蛋白损失率为评价指标,通过单因素实验和正交实验对影响南极磷虾蛋白肽纳滤脱盐效果的主要因素(蛋白肽浓度、压力、循环次数)进行优化。结果显示,采用纳滤技术对南极磷虾蛋白肽进行脱盐处理的最佳工艺条件：蛋白肽浓度为3%、纳滤压力为1.2 MPa、循环次数3次,在该条件下,南极磷虾蛋白肽的脱盐率达到(86.35±2.11)%、蛋白损失率为(9.10±0.35)%。采用优化工艺获得的南极磷虾蛋白肽的盐分含量为(1.14±0.12)%,蛋白质含量为(92.73±2.29)%,相对分子质量主要分布于3000 Da以下,氨基酸组成合理且符合联合国粮农组织/世界卫生组织规定的标准。研究将为高品质南极磷虾蛋白肽产品开发提供技术支撑。     

Fish Protein Isolates Recovered from Silver Carp (Hypophthalmichthys molitrix) By-Products Using Alkaline pH Solubilization and Precipitation

ABSTRACT

Fish protein isolates (FPI) were produced from silver carp processing by-products by means of the isoelectric solubilization-precipitation method, and the quality was evaluated. The alkali-aided process led to significantly more lipid reduction (p < 0.05) compared with the acid-aided process. Similar protein types were extracted for the acid and alkali solubilization, and most protein types can be recovered from the raw materials, as observed in the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The FPI was higher in total amino acids and essential amino acids (EAAs) compared with the original by-products (p < 0.05). The alkali-aided process yielded a higher nutritional and color quality than the acidic process. Furthermore, we found that alkali-aided protein had higher oxidative stability than the acid-aided protein. Our study indicates that high-quality fish proteins can be efficiently recovered from low-value silver carp processing by-products using isoelectric solubilization/precipitation, particularly the alkali-aided process, and subsequently be used in value-added human foods.     

Nutritional Characteristics of a Peruvian Anchovy (Engraulis ringens) Protein Concentrate

ABSTRACT

The aim of the present study is to present a new anchovy protein concentrate and compare it to other similar fishery products. A 76.4% protein concentrate was prepared from fresh Peruvian anchoveta (Engraulis ringens) (APC) by means of acid treatment with 1% citric acid, solubilization with 1% sodium chloride, and isoelectric precipitation processes. Remaining fat was 5.2%, and 0.51 A_w indicates good stability. When APC was rehydrated for 10 minutes, it had a 3.5 initial weight increase. It provided 17.8% protein of very high nutritional quality, with only histidine content deficient for children 1–2 years old. Fat content in APC offered 33.4% EPA+DHA. No changes in the essential/non-essential amino acids (EAA/non-EAA), polyunsaturated/saturated fatty acids (PUFA/SFA), or w6/w3 ratios were observed as a result of the process in the raw material. Because of the high sodium content, a 25 g portion is recommended to fulfill nutritional FAO recommendations. APC can be considered as an important source of calcium (30% adults and 35% children requirements), phosphorous (30% adults and 52% children requirements), and iron content (approximately 20% adults and children needs). APC is an important high-quality protein, providing w-3 fatty acids, calcium, phosphorous, and iron.     

The Fermentation of Antarctic Krill Juice by a Variety of Microorganisms

In order to produce a fermented seasoning of Antarctic krill juice with pleasant taste and unique flavor, the Antarctic krill juice was fermented by application of Aspergillus oryzae, Streptococcus thermophilus, Saccharomyces cerevisiae, and Bacillus licheniformis, respectively. Organic acids and the composition of amino acids in these fermented products were analyzed, as well as the sensory evaluation and taste components analysis. It was found that all four kinds of fermented liquor of Antarctic krill had better flavor, higher amino acid content, unique organic acid content, and higher antioxidant activity. The best flavor and the most abundant nutrition were in the Antarctic krill juice fermented by S. cerevisiae. The amino acid content was up to 1,368.30 mg/100 mL, the scavenging activity of 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals was 71.308%, and there was a variety of amino acids in this fermented liquor, which contained all eight of the essential amino acids not manufactured by the human body.     

冻融循环对南极磷虾虾肉糜滋味成分的影响

以南极磷虾虾(Euphausia superba)肉糜为原料,通过测定南极磷虾虾肉糜在反复冷冻和解冻过程中ATP及其关联产物、游离氨基酸等的变化,分析冻融循环对虾肉糜滋味成分的影响。结果显示,随着冻融循环次数的增加,南极磷虾虾肉糜的ATP及其关联产物降解加剧,呈鲜味的核苷酸含量逐渐减少,呈苦味的核苷酸含量逐渐增加。1次冻融对应的鲜味苦味比为3.32;2次及以上的冻融循环会造成鲜苦比的显著降低(P<0.05),同时K值迅速增大,表明鲜度发生大幅下降;3次冻融循环后,K值显著增大(P<0.05);4次冻融对应的K值为(86.88±3.58)%,南极磷虾虾肉糜已不新鲜。冻融循环次数的增加导致游离氨基酸总含量增加,鲜味氨基酸和甜味氨基酸比例逐渐减小,苦味氨基酸的比例逐渐增大,导致虾肉糜的滋味进一步劣化。研究表明,冻融循环次数控制在2次以内,可较好地保持南极磷虾虾肉糜的滋味。     

南极磷虾粉营养成分的分析与比较

对两种不同来源的南极磷虾粉营养成分进行了分析和评价,并与进口鱼粉的营养成分进行了比较。结果显示,上海开创远洋渔业有限公司开利轮产南极磷虾粉(磷虾粉S)粗蛋白含量为62.89%,高于辽宁远洋渔业有限公司安兴海轮产南极磷虾粉(磷虾粉D)和进口鱼粉的粗蛋白含量,两种磷虾粉粗蛋白质含量分别达到一级品和二级品指标。磷虾粉S、磷虾粉D和进口鱼粉18种氨基酸总量分别为66.36%、57.19%、67.24%,鲜味氨基酸含量较高分别为24.65%、20.69%和25.25%,8种必需氨基酸含量分别为31.43%、21.74%、25.62%,且磷虾粉S必需氨基酸含量明显高于磷虾粉D和鱼粉,磷虾粉S、磷虾粉D和进口鱼粉三种样品必需氨基酸指数(EAAI)分别为57.83、49.89、57.07;磷虾粉S、磷虾粉D和进口鱼粉三种样品脂肪酸中,多不饱和脂肪酸含量较高分别为43.28%、40.25%、46.58%,且磷虾粉S的EPA与DHA的总量(36.14%)高于磷虾粉D(31.30%)及鱼粉(27.65%)。分析结果表明,南极磷虾粉蛋白含量高,且氨基酸组成符合FAO/WHO推荐的蛋白质理想模式,含有丰富的多不饱和脂肪酸,营养价值较高,具有较大的开发利用前景。     

南极磷虾主要过敏原的分离纯化及性质研究

为了探究南极磷虾致敏问题,从南极磷虾中筛选、鉴定、分离纯化其主要过敏原,并对过敏原的性质进行研究,实验通过缓冲盐溶液提取南极磷虾蛋白;采用SDS-PAGE和Western blot(WB)筛选南极磷虾过敏原;使用高效液相色谱-串联质谱鉴定过敏蛋白;通过等电点沉淀、硫酸铵盐析、阴离子交换柱层析分离纯化过敏原;采用SDS-...     

Intestinal absorption of amino acids in the Pacific bluefin tuna (Thunnus orientalis): in vitro lysine–arginine interaction using the everted intestine system

The interaction between lysine (Lys) and arginine (Arg) in the proximal intestinal region of Pacific bluefin tuna (Thunnus orientalis) was evaluated using the everted intestine method. This in vitro intestinal system has been shown to be an effective tool for studying the nutrient absorption without the need to handle the tuna fish in marine cages as needed for digestibility and amino acid (AA) absorption. We used a factorial design with two sets of variables: low and high Lys concentration (10 and 75 mM) and four different Arg concentrations (3, 10, 20, and 30 mM). Both amino acids were dissolved in marine Ringer solution with a basal amino acidic composition consisting of a tryptone solution (9 mg mL^?1). No interaction was observed between the absorption of Lys and Arg during the first 10 min of the experiment when low concentration of Lys and Arg was used in the hydrolyzate solution. However, there seemed to be a positive effect on Lys absorption when both amino acids were at high concentrations (30 and 75 mM, respectively). This type of studies will led us to test different formulations and/or additives to better understand the efficiency of AA supplementation as an alternative to in situ studies that are difficult to follow to design with the Pacific Bluefin Tuna.     

Effects of varying protein level in practical diets on survival,growth, feed utilization and body composition of juvenile tench (Tinca tinca L.)

The aim of this study was to evaluate the effects of practical diets with different protein content on survival, growth, feed utilization and body composition of juvenile tench (Tinca tinca). A 90-day experiment was conducted with 6-month-old juveniles (34.35 mm total length, 0.411 g weight). Six practical diets differing in the protein level were tested: 40, 44, 48, 52, 56 or 60 %. Survival rates ranged from 96.7 to 100 %. The 52 % protein enabled the highest growth (55.49 mm total length, 2.11 g weight, 1.80 % day^?1 specific growth rate) and the lowest feed conversion ratio (1.61) without significant differences (P > 0.05) from the 48 %. Protein productive value ranged from 15.64 to 22.01. The percentages of fish with visible deformities ranged from 1.1 to 4.4 %. The relationship among amino acid profiles of the diets, growth of juveniles, body composition and amino acid requirements of other fish species is discussed. Second-order polynomial regression analysis showed that the optimum dietary protein requirement for maximum growth of juvenile tench may be 52.7 %.     

南极磷虾粉作为鲽形目鱼饲料蛋白源的营养价值评价

分析商品南极磷虾粉、自制酶解磷虾粉的主要营养成分(氨基酸和脂肪酸)组成,同时采用生化方法比较了5种鲽形目鱼肉蛋白的氨基酸组成。以5种鲽形目鱼肉蛋白为参比蛋白,利用必需氨基酸指数、氨基酸比值系数分和关联度分析法评价南极磷虾粉作为鲽形目鱼饲料蛋白源的营养价值,同时对2种磷虾粉的氟含量进行了分析。结果显示,鲽形目鱼肉(干样)中含有16种常见氨基酸,其中,7种为人体必需氨基酸,4种为呈味氨基酸(总量分别为28.52%–38.03%、25.26%–33.56%);5种鲽形目鱼氨基酸组成均符合 FAO/WHO 的理想模式。南极磷虾粉和酶解磷虾粉的粗蛋白分别为60.84%和68.60%,粗脂肪分别为12.08%和10.79%,达到了规定鱼粉的一级品甚至特级品的指标。酶解处理后蛋白含量显著升高(P<0.05),脂肪含量显著下降(P<0.05),灰分含量无差异(P>0.05)。从必需氨基酸指数(EAAI)来看,EAAI 均大于0.95,磷虾粉的必需氨基酸与鲽形目鱼的必需氨基酸拟合度较高。从氨基酸比值系数分(SRC)和关联度分析法来看,2种磷虾粉对于鲽形目鱼的必需氨基酸平衡性较好,相关系数与鱼粉相差不大。氟含量方面,酶解后磷虾粉的氟含量[(331.21±6.70) mg/kg]显著降低(P<0.05),低于欧盟标准(350 mg/kg)。综合来看,2种磷虾粉氨基酸平衡性较好、营养价值较高,是优质蛋白源。     

Acoustic characterization of biological backscatterings in the Kuroshio-Oyashio inter-frontal zone and subarctic waters of the western North Pacific in spring

The acoustic characteristics of biological backscattering in the western North Pacific were studied to verify expert knowledge on species composition in the echosigns. The survey was conducted in the Kuroshio-Oyashio inter-frontal zone and the subarctic waters in April 2003. The species composition of backscatterings was identified by using midwater trawl, Isaacs-Kidd midwater trawl and Bongo net. The differences of mean volume backscattering strengths between 120 and 38 kHz (ΔMVBS₁₂₀₋₃₈) were calculated for the backscatterings. Six types of backscatterings were classified based on the results of net samples and the ΔMVBS₁₂₀₋₃₈. The ΔMVBS₁₂₀₋₃₈ of each group was as follows: copepods 13.7–17.3 dB, krill 11.6–15.3 dB, Japanese anchovy −1.6 to 1.1 dB, a myctophid, Diaphus theta −0.8 dB, sand lance 6.1 dB and larvae and juveniles of pelagic and mesopelagic fish −9.6 to −4.0 dB. The results suggest that biological backscatterings in the Kuroshio-Oyashio inter-frontal zone and subarctic waters of the western North Pacific in spring can be characterized by using ΔMVBS.     

The proteolytic digestive activity and growth during ontogeny of Parachromis dovii larvae (Pisces: Cichlidae) using two feeding protocols

The proteolytic digestive activity and growth of Parachromis dovii larvae during the ontogeny were evaluated in a recirculation system using two feeding strategies during a 28-day period. Larvae were reared using two feeding protocols (three replicates each): (A) Artemia nauplii (at satiation), fed from exogenous feeding [8 days after hatching (DAH)] until 15 DAH followed by nauplii substitution by formulated feed (20 % day^?1) until 20 DAH and then formulated feed until 28 DAH; (B) formulated feed (100 % BW daily) from exogenous feeding until 28 DAH. Levels of acid (pepsin type) and alkaline digestive proteases as well as growth and survival of larvae were measured along the feeding period. Survival was high and similar between treatments: 98.9 ± 0.0 for Artemia, 97.3 ± 0.0 % for formulated feed. The specific growth rate for length and weight was higher in larvae fed with Artemia nauplii than in larvae reared with formulated feed: 3.4 ± 0.1 versus 1.8 ± 0.1 % day^?1 for body length (P = 0.009) and 12.2 ± 0.1 versus 6.5 ± 0.3 % day^?1 for body weight (P = 0.002). The acid and alkaline proteolytic activity was detected, in both treatments, from the beginning of the experiment, at 8 DAH. The total enzymatic activity (U larva^?1) for acid and alkaline proteases was higher in larvae reared with Artemia after 12 DAH, whereas the specific enzymatic activity was similar for both enzyme types in the two treatments. The results suggest that P. dovii larvae were capable to digest formulated diets from the beginning of exogenous feeding and that they could be reared with formulated feeds. However, the formulated feed used should be nutritionally improved because of the poor growth obtained in this research.     

Partial characterization of the digestive enzymes of Pacific bluefin tuna <Emphasis Type="Italic">Thunnus orientalis</Emphasis> under culture conditions

The digestive enzyme activities of Pacific bluefin tuna Thunnus orientalis were evaluated for specific activity and characterized for pH and temperature optima in crude extracts of stomach, caecal mass, and proximal, middle and distal intestine. A higher level of alkaline proteolytic activity was detected in the caecal mass than in the proximal intestine. Total alkaline proteases, trypsin, chymotrypsin and leucine aminopeptidase (LAP) were tested. The temperature and pH analyses showed that proteolytic activity as well as lipase were maximal in the alkaline range, with a maximum at pH 9.0 and at temperatures between 35 and 60°C, except for the pepsin, which showed maximum activity at the same temperatures but in the acid range (pH 3.0). The α-amylase activity showed a broader range in activity, both for pH and temperature, with higher activity over the alkaline pH values and higher temperature. The lipase activity seems to be nondependent on bile salts under our assay conditions, resulting in a significant activity reduction in the presence of bile salts. This knowledge will allow the development of a gastrointestinal model (everted intestine) where food or feed will be hydrolysed with the fish’s own enzymes, a project that is being undertaken in our laboratory as a contribution to the development of novel diets for tuna fish.     

Production of Biscuit from Chinese Sturgeon Fish Fillet Powder (Acipeneser sinensis): A Snack Food for Children

ABSTRACT

Malnutrition in children is a current global burden. A common snack for children is biscuits prepared from cereals. However, these types of biscuits have low protein content. Therefore, producing healthier biscuits enriched with essential amino acids (EAAs) and other micronutrients is of great interest to researchers and manufacturers. In this study, biscuits produced from low gluten wheat flour fortified with sturgeon fillet powder (SFP) in various mixing ratios (100:0, 95:5, 93:7, and 90:10, respectively) were investigated. The results revealed that the nutritional and proximate composition of biscuits were significantly (P ≤ 0.05) improved: moisture (6.77 ± 0.06–7.65 ± 0.06), protein (13.23 ± 0.35–17.12 ± 0.02), fat (15.57 ± 0.18–15.85 ± 0.25), ash (0.84 ± 0.07–0.92 ± 0.06), spread ratio (14.0 ± 0.92–14.33 ± 0.21), and hardness (2,057.90 ± 28.33–3,089.02 ± 51.53). Total amino acids were 6.91, 9.34, 10.32, and 10.58; and EAAs were 2.43, 3.97, 4.48, and 4.22 g/100 g for 0%, 5%, 7%, and 10% SFP, respectively. Leucine was the major amino acid with 5% and 10% SFP, while methionine was the major amino acid with 7% SFP. Fungal counts were not observed, and quality indices were below the maximum acceptable limits in the products. Biscuits supplemented with 7% SFP showed the highest sensory evaluation score. Overall, the inclusion of 7% SFP could produce protein enriched cereal-based fortified biscuits with acceptable sensory and quality attributes and ultimately could insure effective fish utilization and solve malnutrition problems.     

Functional Properties of White Shrimp (Litopenaeus vannamei) By-Products Protein Recovered by Isoelectric Solubilization/Precipitation

The processing of fish in general entails quantities of by-products, such as exoskeleton and cephalothorax of shrimp, which may reach 70%. Protein recovered from by-products has been investigated in recent years for its ability to offer more nutritious foods and improved functional properties. The objective of the present study was the use of the pH shifting process to recover proteins from solubilization and precipitation that are present in the by-products (heads and shells) of white shrimp (Litopenaeus vannamei). The proximal composition of the by-products presented 58.5% protein (d.b). The isoelectric point used to precipitate the protein was defined at pH 4. The protein concentrate presented 73.1% of protein. The yield of the protein concentrate mass resulted in 47.8% (w.b). The greatest solubility found for the protein concentrate was 51.3% at pH 8. The oil and water retention capacity resulted in values of 8.5 and 2.5 mL/g of protein, respectively, and the highest emulsifying capacity was found at pH 8. The by-products and concentrate amino acids profiles confirmed the presence of 16 amino acids. In general, the shrimp protein concentrate obtained showed the potential to be included as an ingredient in food formulations.