Evaluation of the Antioxidant and Antimicrobial Activity of Protein Hydrolysates and Peptide Fractions Derived from Colossoma macropomum and Their Effect on Ground Beef Lipid Oxidation

The objective of this study was to obtain protein hydrolysate from the mechanically separated meat of blackfin pacu to evaluate the influence by ultrafiltration in the antioxidant and antimicrobial activities of the peptide fractions obtained and to apply in ground beef to evaluate the lipid stability. The enzymatic hydrolysis was performed using the enzyme Protamex (pH 7.0, 60°C) for 240 min. The protein hydrolysate was fractionated by ultrafiltration. Then, the antioxidant capacity of the protein hydrolysate and the peptide fractions were evaluated in vitro by the methods of 2,2’-azinobis (3-ethylbenzothiazoline sulfonic acid) radical capture, 2,2-diphenyl-1-picrylhydrazyl radical-scavenging assay, reducing power, and hydroxyl radical scavenging activity. The antimicrobial activity of the samples was evaluated by disc-diffusion against Staphylococcus aureus and Escherichia coli. After evaluation, the peptide fractions did not present higher bioactivities than that shown for the hydrolysate. The protein hydrolysate was applied to ground beef, where the substances reactive to thiobarbituric acid and color were evaluated during 7 days of storage at 4°C. Lipid oxidation was reduced up to 60.9% and there was no modification of the natural coloration. Thus, the protein hydrolysate can be used as an alternative source of antioxidant for the preservation of refrigerated meats.     

Antioxidant Properties of Protein Hydrolysate Obtained from Oyster Saccostrea cucullata (Born, 1778)

The antioxidant activities of enzymatically hydrolyzed (protease from Bacillus cereus SU12) oyster (Saccostrea cucullata) protein were studied. The hydrolysate exhibited a strong antioxidant potential in 1, 1-diphenyl-2-picrylhydrazyl (DPPH, 85.7 ± 0.37%), followed by hydrogen peroxide radical scavenging activity (81.6 ± 0.3%), hydroxyl radical scavenging activity (79.32 ± 0.6%), and reducing power assay (2.63 ± 0.2 OD at 700 nm) at a concentration of 1 mg/mL. Due to the high antioxidant potential, the hydrolysate was purified in Sephadex G-25 gel filtration chromatography. The active peptide fraction was identified by DPPH and reducing power assay. The amino acid content of the purified active peptide fraction was analyzed by high performance liquid chromatography. The active fraction contained a good quantity of both essential and nonessential amino acids. The present study revealed that oyster (S. cucullata) protein hydrolysate is a potential source for natural antioxidants.     

In Vivo Antioxidant Effects of Hydrolysate Derived from Waste Proteins of Mactra veneriformis

Mactra veneriformis has been used as seafood and traditional Chinese medicine for thousands of years in China. In the present study, hydrolysate was prepared from waste proteins of M. veneriformis by trypsin treatment. The protective effect of hydrolysate on the defense system against oxidative stress was investigated by estimating the superoxide dismutase (SOD) activity and malondialdehyde (MDA) level in normal mice. The antioxidant activity of hydrolysate was investigated in D-galactose-induced aging mice by estimating the SOD and glutathione peroxidases (GSH-px) activity and MDA level. Amino acid composition and molecular weight distribution were also determined. These investigations suggest that hydrolysate derived from by-products of M. veneriformis possess antioxidant activity and may be attributed to elevation of the activity of antioxidase and enhanced antioxidant defenses.     

Characterization of Spray-Dried Hydrolyzed Proteins from Pink Perch Meat Added with Maltodextrin and Gum Arabic

In the present study, spray-dried pink perch meat protein hydrolysate (PPH) and hydrolysate with maltodextrin and gum arabic (PPHMG) were characterized in relation to their physical, rheological, functional, antioxidant, thermal, and sensory properties. The Herschel–Bulkley model was suitable to explain the flow behavior of PPH and PPHMG emulsions. Both the samples exhibited functional and antioxidant properties. Antioxidant activities were significantly higher for PPH compared with PPHMG (p<0.05). PPH and PPHMG alone did not exhibit antimicrobial activity but enhanced the activity of chitosan against pathogens. Sensory acceptability of the samples in soup revealed that PPHMG of up to 4% was highly scored without any traces of bitterness, whereas for PPH, a concentration of up to 3% was acceptable.     

Improvement of Antioxidant Activity of Grass Carp (Ctenopharyngodon idella) Protein Hydrolysate by Washing and Membrane Removal Pretreatments and Ultrasonic Treatment

To improve the antioxidant activity of grass carp hydrolysate, washing and membrane removal pretreatments and ultrasonic treatment were applied in this study. Various pretreatment methods (washing methods and membrane removal methods) for removing the prooxidants and phospholipid in minced fish were evaluated. The effects of ultrasonic treatment prior to enzyme hydrolysis on the antioxidant activities of minced carp hydrolysate were investigated. The minced carp subjected to washing with 0.4% NaCl solution and subsequent membrane removal with CaCl₂ and citric acid solution had the lowest content of prooxidants and phospholipid remaining in the minced fish. The hydrolysate produced from washing/membrane removal pretreated carp with ultrasonic treatment for 20 min had the highest ferric reducing antioxidant power and scavenging activities against 1,1-diphenyl-2-picrylhydrazyl and 2,2?-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) radicals. This suggested that the combination approach of washing/membrane removal pretreatment and ultrasonic treatment could improve the antioxidant activity of carp hydrolysate, and this hydrolysate could be a potential antioxidant ingredient for functional foods.     

Effect of Degree of Hydrolysis and Protease Type on the Antioxidant Activity of Protein Hydrolysates From Cuttlefish (Sepia officinalis) By-Products

We have investigated the antioxidant activities of eight hydrolysates from cuttlefish by-products obtained by treatment with various gastrointestinal proteases (chymotrypsin, trypsin, and crude alkaline enzyme extracts from cuttlefish and sardinelle) and bacterial proteases (Alcalase and crude enzymes from Bacillus pumilus A1, Bacillus mojavensis A21, and Bacillus cereus BG1). The antioxidant activities of the cuttlefish by-products protein hydrolysates (CPHs) were evaluated using various in vitro antioxidant assays, such as 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity, reducing power, and total antioxidant capacity. All hydrolysates showed different degrees of hydrolysis (DH) and varying degrees of antioxidant activity. Among the different hydrolysates, cuttlefish crude enzyme hydrolysate exhibited the highest antioxidant activity, followed by sardinelle crude enzyme and Alcalase hydrolysates. Further, CPHs with different degrees of hydrolysis were prepared by treatment with proteolytic enzymes from cuttlefish, sardinelle, and B. mojavensis A21. All hydrolysates showed a greater antioxidative activity as indicated by all the methods considered. In addition, antioxidant activity in hydrolysates was positively correlated with the increase of DH. The results of this study indicated that CPHs might be a good candidate for further investigation in developing new antioxidants.     

Antioxidative Activitiy of Protein Hydrolysate from the Muscle of Common Kilka (Clupeonella cultriventris caspia) Prepared Using the Purified Trypsin from Common Kilka Intestine

Trypsin from the intestine of common kilka (Clupeonella cultriventris caspia) was purified using ammonium sulfate precipitation (30–50% saturation), Sephadex G-75, and DEAE-cellulose chromatography with the purity of 30-fold and the yield of 12%. The molecular weight of trypsin was estimated to be 23.2 kDa based on sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The trypsin had optimal activity at pH 8.0 and 60°C using N-α-benzoyl-DL-arginine-ρ-nitroanilide hydrochloride (BAPNA) as a substrate and showed high stability in the pH range of 7.0–10.0. It was stable up to 50°C. Soybean trypsin inhibitor (SBTI) and N-ρ-tosyl-L-lysine-chloromethylketone (TLCK) significantly inhibited trypsin activity (p < 0.05). Protein hydrolysate from common kilka muscle with different degrees of hydrolysis (DHs; 20, 30, and 40%) was prepared using the purified trypsin, and antioxidative activities were determined. 1,1-Diphenyl-2-picrylhydrazyl (DPPH), 2,2’-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging activities, ferric reducing antioxidant power, and ferrous chelating activity of hydrolysate increased with increasing DH up to 40% (p < 0.05). Therefore, trypsin from intestine of common kilka could be used as a processing aid for production of fish protein hydrolysate with antioxidative activity.     

Antioxidant and ACE Inhibitory Activities of Kawakawa (Euthynnus affinis) Protein Hydrolysate Produced by Skipjack Tuna Pepsin

ABSTRACT

Pepsin enzyme from skipjack tuna was extracted for the production of kawakawa (Euthynnus affinis) fish protein hydrolysate. Using ultra-fractionation, Kawakawa protein hydrolysates were separated into four different fractions, including fractioned protein hydrolysate I (FPH I) (< 1 kDa), FPH-II (1–3 kDa), FPH-III (3–10 kDa), and FPH-IV (> 10 kDa). The antioxidant activity was assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity, inhibition of linoleic acid oxidation, reducing power tests, and chelating activity of metal ions. Results indicated that FPH II fraction peptides had higher antioxidant activity in comparison with the other fractions, followed by FPH I. Further, the fractions were evaluated for angiotensin converting enzyme (ACE) inhibition, and IC₅₀ value ranged from 0.45 to 1.86 mg/ml with higher activity in FPH I (IC₅₀ 0.45). Finally, the amino acid profile of different fractions was analyzed. The fractions exhibited significant amounts of hydrophobic amino acids, which could perform as hydrogen donors, frustrate the free radicals, and inhibit the ACE. The recovered pepsin from the viscera was used to produce hydrolysates with good biological activities. Peptides lower than 3 KDa had antioxidant activity as positive controls and significant ACE activity. These are very important findings that could be used to conduct further research in a preclinical study of these peptides.     

Anti-Inflammatory,Antioxidant, and Antimicrobial Effects of Underutilized Fish Protein Hydrolysate

Antimicrobial, anti-inflammatory, and antioxidant activities of protein hydrolysates from Argentine croaker (Umbrina canosai) protein isolate (CPI) or Argentine croaker myofibrillar protein with different degrees of hydrolysis (DH: 10–20%) prepared using Alcalase or Protamex were determined. Results showed that an increase in the DH resulted in higher content of hydrophobic and aromatic amino acids (AAAs) and in a decrease in molecular weight (MW) distribution for all hydrolysates obtained. Furthermore, the enzyme and raw material used influenced the amino acid content and MW determined. Hydrolysates from CPI with a 20% DH by Alcalase had higher 2,2?-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) radical scavenging activity, metal chelation, and ferric-reducing antioxidant power (p < 0.05). All hydrolysate samples decreased the pro-inflammatory capacity. In all the evaluated microorganisms, only seven were inhibited, most being Gram-positive. Alcalase was found to exert a considerable influence on antibacterial activity. These hydrolysates are an alternative as natural antimicrobials, anti-inflammatory, and antioxidant compounds.     

大眼金枪鱼头蛋白酶解物1 ku超滤组分的抗氧化活性及其理化性质

研究大眼金枪鱼头蛋白酶解物1 ku超滤组分体外的还原力、自由基清除能力及对衰老小鼠体内抗氧化能力的影响,分析1 ku超滤组分的一般成分、氨基酸组成及分子量分布,为进一步分离纯化金枪鱼头抗氧化肽提供基础。体外结果显示,1 ku超滤组分对羟基自由基、超氧阴离子和DPPH自由基的清除活性随浓度的增加而增强,IC50分别为1.38、0.73与0.93mg/mL,还原力也随浓度的增加而增大,在浓度为12.5 mg/mL时为0.763;体内结果显示,灌胃30 mg/kg的1 ku超滤组分连续42 d,D-半乳糖致衰老小鼠肝组织的超氧化物歧化酶(SOD)活性、肝组织和血清的谷胱甘肽过氧化物酶(GSH-PX)活性显著提高(P<0.05),血清丙二醛(MDA)含量显著降低(P<0.01);理化分析结果显示,1 ku超滤组分(干基计)蛋白质含量为96.40%,脂肪0.11%,灰分4.86%,疏水性氨基酸占氨基酸总量的35.8%,活性组分分子量在1 802~2 519 u和422~922 u。     

Effect of dietary hydrolysate supplementation on growth performance,non‐specific immune response and disease resistance of olive flounder (Paralichthys olivaceus) challenged with Edwardsiella tarda

A 9‐week feeding trial was conducted to investigate the effects of dietary supplementation with protein hydrolysates on growth, innate immunity and disease resistance of olive flounder. A fishmeal (FM)‐based diet was regarded as a control, and three diets were prepared by partial replacement of FM with krill hydrolysate, shrimp hydrolysate or tilapia hydrolysate (designated as Con, KH, SH and TH, respectively). Triplicate groups of fish (24.5 ± 0.3 g) were fed one of the diets to apparent satiation twice daily for 9 weeks and then challenged by Edwarsellia tarda. Fish‐fed KH diet showed significantly (P < 0.05) higher growth performance and feed utilization compared with the Con diet. Dry matter digestibility of the diets was significantly increased by KH and TH supplementation. All the examined innate immune responses were significantly increased in fish fed KH diet. Significantly, higher respiratory burst and superoxide dismutase (SOD) activities were found in fish‐fed SH diet. Lysozyme and SOD activities were significantly increased in fish‐fed TH diet. However, no significant effect was found on fish disease resistance. This study indicates that dietary supplementation of the hydrolysates, particularly KH, can improve growth performance, feed utilization and innate immunity of olive flounder.     

Purification and Characterization of a New DPPH Radical Scavenging Peptide from Shrimp Processing By-products Hydrolysate

A peptide with high antioxidant activity was isolated and identified from shrimp processing by-products hydrolysate. The 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity was used to evaluate the antioxidant activity of fractions. The purified antioxidant peptide was identified as Ser-Val-Ala-Met-Leu-Phe-His (804.4 Da) by electrospray ionization tandem mass spectrometry. The purified peptide at 50 μg/mL showed antioxidative activity of 65.7 ± 0.9%, which was 3.18-fold higher compared with the first step separation by ion-exchange chromatography. It is possible to produce natural antioxidative peptides from shrimp processing by-products hydrolysate. The high antioxidant activity may be due to the presence of Phe-His segment at the C-terminus of the peptide.     

Effect of Enzyme Type on the Antioxidant Activities and Functional Properties of Enzymatic Hydrolysates from Sea Cucumber (Cucumaria frondosa) Viscera

ABSTRACT

Effects of various proteases were obvious on the amino acid composition, antioxidant activities, and functional properties of enzymatic hydrolysates prepared from sea cucumber viscera (SCV). Alcalase, trypsin, and flavourzyme conferred better antioxidant activities on hydrolysates and endowed them with a higher degree of hydrolysis, coming to 19.08, 32.38, and 15.94%, respectively. For functional properties, neutrase- and flavourzyme-derived hydrolysates showed better foaming properties. Amino acid composition suggested that papain, neutrase, alcalase, and bromelain granted the hydrolysate’s high nutritional quality. The results indicated that SCV hydrolysates could be used in food systems to impart desirable characteristics to products.     

Effect of size‐fractionated fish protein hydrolysate on growth and feed utilization of turbot (Scophthalmus maximus L.)

Four experimental diets were fed to turbot to examine the effect of fish hydrolysate and ultra‐filtered fish hydrolysate on growth performance, feed utilization and non‐specific immune response. Fish hydrolysate was produced by enzymatic treatment and size fractionated using ultra‐filtration (UF). The permeate (molecular weight <1000 Da) after UF and the non‐ultra‐filtered fish hydrolysate (NUF) were tested as feed ingredients. Diets UF1, UF2 contained 3.7%, 1.2% ultra‐filtered fish hydrolysate to replace fish meal protein respectively. The diets UF1, NUF were identical in composition except that the molecular weight of fish hydrolysate in the diet. Fish meal was used in the control diet. All diets were made equal in protein, lipid and energy. Each experimental diet was fed to juvenile turbot (27.87 ± 0.04 g) in triplicate for 8 weeks. Results of this study indicate that the best overall growth and feed utilization of turbot juveniles were obtained with a diet containing higher dose of the small molecular weight compounds in fish hydrolysate. Acid phosphatase, alkaline phosphatase, lysozyme and superoxide dismutase activity in serum were not affected by diet. Total antioxidant capacity was improved with increasing level of low molecule weight fish hydrolysate (UF1).     

鲢鱼鱼皮蛋白肽的制备与抗氧化活性评价

为制备抗氧化活性良好的鲢鱼鱼皮蛋白肽,采用胰蛋白酶、碱性蛋白酶、菠萝蛋白酶和木瓜蛋白酶等4种常见的商业酶对鲢鱼鱼皮进行酶解,测定酶解物的ABTS自由基清除力和Fe2+螯合力来评价其抗氧化活性,并用超滤及凝胶层析对酶解物进行分离,以期得到活性更好的酶解物分离组分。酶解后产物的抗氧化活性均有所提高,其中碱性蛋白酶酶解2 h产物活性较强。对此酶解物用截留分子量为10 k Da、5 k Da和3 k Da的中空纤维超滤膜进行超滤,得到的4个组分中,分子量越小的组分抗氧化活性越强。分子量小于3 k Da的组分经Sephadex G-15凝胶层析得到3个组分,其中分子量最大的组分活性较好,在0.51 mg/m L质量浓度下测定其ABTS自由基清除率和Fe~(2+)螯合力分别为(79.65±0.87)%和(93.40±0.20)%。该研究成果对鲢鱼鱼皮抗氧化肽的开发具有较好的指导作用。     

Antioxidant Assessment of Schizochytrium Meal Protein Enzymatic Hydrolysate and Its Potential Application

Schizochytrium meal protein (SMP) extracted from Schizochytrium meal was hydrolyzed by flavourzyme. Response surface methodology (RSM) was used to optimize the extraction conditions for the protein extraction yield from Schizochytrium meal. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (DRSA) was used to evaluate the antioxidant activity of hydrolysates. The orthogonal test was used to investigate the effects of three independent variables, namely protease dosage, hydrolysis time, and pH. The optimum conditions obtained were as follows: protease dosage of 7%, hydrolysis time of 1.5 h, pH of 6, under which, DRSA at the concentration of 2 mg/mL was 89.38%. Aspartic and glutamic acid constituted approximately 26.32% of the total amino acids, and glutamic acid was the most abundant amino acid of Schizochytrium meal protein hydrolysate (SMPH) by amino acid composition analysis, which may have contributed to the scavenging activity of SMPH. Moreover, SMPH was made into chewable tablets with suitable formula and high humidity stability. These findings indicate that Schizochytrium meal can be reused as a raw material for preparation of antioxidant peptides.     

Antioxidant Properties and Arsenic Speciation of Ultrafiltration and Nanofiltration Derived Abalone Viscera Hydrolysate Fraction

A fraction with molecular weight of 0.2–1.0 kDa was prepared from abalone viscera hydrolysate by ultrafiltration and nanofiltration. The contents of protein, carbohydrate, and fat in abalone viscera were 45.75%, 18.19%, and 13.60%, respectively. The protein in abalone viscera hydrolysate faction (AVHF) reached up to 75.73%, while carbohydrate and fat fell to 1.80% and 0.20%, respectively. The leucine (Leu) content was the highest in the AVHF. On the other hand, the hydroxyl and 2,2-diphenyl-1-picrylhydrazyl radical scavenging activities and reducing power increased with increased concentration of AVHF. The inhibiting ability of AVHF on lipid peroxidation was higher than that of ascorbic acid. The total content of K, Na, and P in AVHF was 9.78 g/100 g, and total arsenic content reached 16.23 mg/kg. The proportion of arsenobetaine and arsenocholine accounted for more than 96%, while the proportion of arsenate was 0.47%, and arsenite was not detectable. It is concluded that the AVHF from abalone viscera is a safe natural antioxidant.     

罗非鱼鱼皮酶解物及其硒螯合物的结构与抗氧化特性

为了高值化利用罗非鱼副产物资源以及为酶解物-硒螯合物的产业化提供理论依据,本实验以罗非鱼鱼皮胶原蛋白(TSC)为原料,用4种蛋白酶对其进行酶解,分别获得了酸性蛋白酶酶解物(TSCAc)、木瓜蛋白酶酶解物(TSCP)、菠萝蛋白酶酶解物(TSCB)、碱性蛋白酶酶解物(TSCAl),以Na₂SeO₃为硒源,制备并筛选出硒螯合量最高的酶解物,测定酶解物及其硒螯合物的紫外光谱、荧光光谱、红外光谱、表面疏水性,并对酶解物及其硒螯合物的抗氧化性与抗氧化稳定性进行分析。结果显示,TSCAl的水解度(DH)最高(18.2%),其与硒螯合后的硒螯合量最高(153.5 mg/g),硒螯合率最高(23.7%),小分子量(500～1 000 u)的含量最高。其与硒螯合后,紫外光谱吸收峰强度增大,荧光光谱吸收峰由波长419.8 nm红移到422.0 nm且荧光强度减弱,疏水性在螯合后降低,表明TSCAl与硒螯合生成了新的物质。推测酰胺键、氨基的氮原子、羧基的氧原子可能参与了与硒的螯合反应。研究表明,碱性蛋白酶酶解物-硒螯合物(TSCAl-Se)具有良好的抗氧化性,其在不...     

马氏珠母贝肉酶解产物的抗酒精性肝损伤作用

为探究马氏珠母贝肉酶解产物(Enzymatic hydrolysate from Pinctada martensii,EP)对酒精性肝损伤(Alcoholic liver damage,ALD)的保护作用,该研究将EP超滤分级为截留分子量>10 kD(EP-Ⅰ)、3~10 kD(EP-Ⅱ)和<3 kD(EP-Ⅲ)3个组分,检测其体外抗肝损伤活性及对ALD小鼠肝保护作用的影响。体外试验结果显示,EP-Ⅲ可显著激活体外乙醇脱氢酶(ADH)活性(P<0.01),3个超滤组分均具有一定的体外抗氧化能力且EP-Ⅲ>EP-Ⅱ>EP-Ⅰ;动物试验结果显示,与模型对照组相比,各超滤组分均能够显著降低小鼠血清中谷丙转氨酶(ALT)和谷草转氨酶(AST)活力、小鼠肝脏指数及肝脏中丙二醛(MDA)和甘油三酯(TG)含量,同时显著增强小鼠肝脏中超氧化物歧化酶(SOD)、乙醇脱氢酶(ADH)和乙醛脱氢酶(ALDH)活力,提高肝脏中的谷胱甘肽(GSH)含量。综上,马氏珠母贝肉酶解超滤组分对急性ALD具有一定的辅助保护作用,其中EP-Ⅲ的保护作用效果最佳,其机制可能与加快机体乙醇代谢和减缓乙醇对机体造成的氧化损伤相关。     

Maturation of the pancreatic and intestinal digestive functions in sea bass (Dicentrarchus labrax): effect of weaning with different protein sources

The maturation of the digestive functions in sea bass (Dicentrarchus labrax) larvae was evaluated by the enzymatic profile of pancreas and intestine brush border membranes. Sea bass larvae were weaned at day 25 with three simplified diets different by their protein nature: 100% fish meal (FP), 100% casein mixture (CP) and 50% fish meal-50% casein mixture (CFP). The casein mixture contained 35% of hydrolysate. The control group was fed live preys. The specific activity of amylase decreased with age irrespectively of the diets whereas the specific activity of trypsin was enhanced. The casein mixture reduced pancreatic secretion in amylase and trypsin. The CFP group differed from the other groups fed on compound diets, exhibiting as soon as day 32 high activities of brush border enzymes, similar to controls. This sharp increase between day 25 and 32 appeared to be crucial for larval survival. The addition of a protein hydrolysate in a weaning diet seems to facilitate this maturation process.