Antioxidant Activity Of Hydrolysates Research Articles

Improving antioxidant activities of whey protein hydrolysates obtained by thermal preheat treatment of pepsin, trypsin, alcalase and flavourzyme

SummaryAntioxidant activity of whey protein concentrate (WPC) hydrolysates was evaluated. Hydrolysates were obtained by pepsin, trypsin, alcalase and flavourzyme enzymatic reaction and preheat treatment of 95 °C for 5 or 10 min. The degree of hydrolysis (DH) was determined by 2,4,6‐trinitrobenzene sulphonic acid method, and antioxidant properties were determined by three spectrophotometric methods: ferricyanide method, ferric reducing/antioxidant power assay and diphenyl‐picryl hydrazinyl radical‐scavenging activity. For all the enzymes, briefly preheat treatment (95 °C/5 min) increased DH of WPC. Alcalase hydrolysates showed the highest antioxidant activity by three methods. The changes in antioxidant activity was coincidental with the changes in DH (R2 = 0.988). Hydrolysates analysed by polyacrylamide gel electrophoresis and high performance liquid chromatography indicated that the α‐La was hydrolysed completely by pepsin, trypsin and alcalase and was resistant to flavourzyme to some extent; β‐lactoglobulin was only completely hydrolysed by trypsin and alcalase. Results indicated that antioxidant activity of hydrolysates was greatly related to the exposure of amino acid residues.

단백질 분해 효소 처리가 청국장의 항산화 활성에 미치는 영향

Alcalase, protamex 및 neutrase 효소처리가 청국장의 펩타이드 및 항산화 활성 증가에 미치는 영향을 동일한 조건으로 효소 처리한 대두분말과 비교하였다. 효소 20, 100 및 500mAU를 50℃에서 120분간 처리한 결과 청국장과 대두분말 모두에서 alcalase 및 protamex에 의한 펩타이드 생성이 neutrase보다 높게 나타났다. 항산화 활성은 청국장의 경우 alcalase 처리에 의해 DPPH, ABTS 및 hydroxyl radical 소거활성이 효소처리를 하지 않은 대조구와 비교하여 시료량에 따라 각각 37~57%, 59~106% 및 67~83%, protamex 처리에 의해 35~50%, 56~74% 및 52~75% 통계적으로 유의하게 증가하였다. 대두분말을 효소 처리한 시료에서도 유사한 결과를 보여 alcalase 처리에 의해 DPPH, ABTS 및 hydroxyl radical 소거활성이 각각 37~55%, 51~99% 및 67~83%, protamex 처리에 의해 36~44%, 44~66% 및 51~65% 증가하였다. 그러나 두 시료 모두에서 neutrase 처리에 의한 항산화 활성의 유의적 증가는 없었다. 이상의 결과는 alcalase와 protamex가 청국장에 특이적이지는 않으나 펩타이드 생성 및 항산화 활성 증가에 효율적임을 보여주고 있다.

In vitro antioxidant activity of enzymatic hydrolysates prepared from abalone (Haliotis discus hannai Ino) viscera

Pacific abalone (Haliotis discus hannai Ino) viscera, which are normally discarded as byproducts, were hydrolyzed with five commercially available proteases, including alkali protease, papain, neutral protease, pepsin, and trypsin. The hydrolysates obtained were fractionated using ultrafiltration membrane bioreactor system. The resulting hydrolysate fractions were investigated for their antioxidant activities, including the scavenging effect on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, the scavenging effect on hydroxyl radical, and the reducing power. Results showed that the hydrolysate fractions were effective antioxidants, and the antioxidant activity of the hydrolysates was related to the enzyme type employed. As byproducts, abalone viscera can be used as a potential source of antioxidant peptides.

Characterization of a novel antioxidative peptide from the sand eel Hypoptychus dybowskii

Protein derived from the sand eel, Hypoptychus dybowskii, was hydrolyzed using different proteases (alcalase, neutrase, α-chymotrypsin, papain, pepsin, and trypsin) to produce an antioxidant peptide. Antioxidant activity of hydrolysates was evaluated using DPPH radical scavenging activity. The papain hydrolysate exhibited the highest antioxidative activity compared to other hydrolysates. The free radical scavenging activity of papain hydrolysate was 77.4% at 1.0mg/ml. The peptide demonstrating the strongest antioxidative activity was isolated from the hydrolysate using consecutive chromatography. The amino acid sequences of purified peptide was identified as Ile–Val–Gly–Gly–Phe–Pro–His–Tyr–Leu (1189Da), and the EC50 value of antioxidant peptide was 22.75μM. The purified peptide exhibited an inhibitory effect against DNA oxidation induced by hydroxyl radical. The results of this study suggest that sand eel protein hydrolysate is a good source of natural antioxidants.

Antioxidant activities of pepsin hydrolysates of water‐ and salt‐soluble protein extracted from pork hams

SummaryThe objective of this study was to evaluate the antioxidant activities of pepsin‐digested water‐soluble protein (WSP) and salt‐soluble protein (SSP) extracted from pork ham. WSP and SSP were hydrolysed by pepsin for 2–10 h with 2‐h increments. The protein hydrolysates by pepsin were analysed by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis, and reducing power of the hydrolysates was measured. In addition, antioxidant activity of the hydrolysates was determined using linoleic acid emulsion system. Protein bands with molecular weight higher than 7 kDa in WSP and SSP were completely hydrolysed by pepsin after 2 h of digestion time. WSP hydrolysates (WSPH) and SSP hydrolysates (SSPH) had higher ferric reducing power than controls (WSP and SSP without pepsin digestion). Reducing powers of WSPH were higher than those of SSPH, regardless of digestion time. The oxidative activity of linoleic acid was predominantly inhibited by the addition of WSPH and SSPH, especially 0.5% protein hydrolysate. These results indicate that several functional peptides of pork protein digested by pepsin might have antioxidant activity, and thus they may be used as an antioxidant agent in muscle food system.

Antioxidant activity of hydrolysates and peptide fractions derived from porcine hemoglobin

Porcine hemoglobin hydrolysate (PHH) was prepared with 6 different proteases (flavourzyme, papain, A.S.1398, alcalase, pepsin and trypsin). There was no correlation between extent of hydrolysis and antioxidant activity (p > 0.05). The peptic hydrolysate prepared at 60min possessed the strongest antioxidant potential (67.0 ± 1.84%) among different hydrolysates, which was fractionated into 4 major types by ultrafiltration membranes with different molecular weight cut-off (MWCO), PHH-I (Mw > 10kDa), PHH-II (Mw = 5-10kDa), PHH-III (Mw = 3-5kDa) and PHH-IV (Mw < 3kDa). PHH-IV possessed higher inhibitory effects of lipid peroxidation and scavenging effects on superoxide radical compared with larger MW fractions. Four fractions possessed the scavenging effects on 1, 1-diphenyl-2-picrylhy-drazyl (DPPH) and hydroxyl radicals in the order PHH-IV > PHH-I > PHH-III > PHH-II. MW of the 2 major peptide fractions from PHH-IV was located at 2476Da (F1) and 1042Da (F2), respectively. PHH-IV could be utilized to develop physiologically functional foods or therapy drugs.

Antioxidative Activities of Hydrolysates from Duck Egg White Using Enzymatic Hydrolysis

Duck egg white (DEW) hydrolysates were prepared by five enzymes (papain, trypsin, chymotrypsin, alcalase, and flavourzyme) and their antioxidant activities investigated in this study. DEW hydrolyzed with papain (DEWHP) had the highest peptide content among the five enzymatic treatments. Besides, the peptide content of DEWHP increased when the enzyme to substrate ratio (E/S ratio) increased. It was suggested that higher E/S ratio contributed to elevate the degree of hydrolysis of DEW effectively. Similar results were also obtained by Tricine-SDS-PAGE. In addition, SDS-PAGE patterns indicated papain was the only one amongst all enzymes with the ability to hydrolyze DEW. In antioxidant properties, DEWHP showed more than 70% of inhibitory activity on linoleic acid peroxidation and superoxide anion scavenging. Moreover, the Fe 2+ chelating effect of DEWHP was greater than 90%, while no significant difference was observed in DPPH radical scavenging and reducing ability. The results of peptide contents, antioxidant activities and electrophoresis suggested that the higher the peptide content, the stronger the antioxidant activities in DEWHP. (Key Words : Duck Egg White, Hydrolysis, Antioxidant Activities)

Antioxidant peptides isolated from the marine rotifer, Brachionus rotundiformis

Protein derived from the rotifer Brachionus rotundiformis was hydrolyzed using different proteases (Alcalase, α-chymotrypsin, Neutrase, papain, pepsin and trypsin) for production of antioxidant peptide. Antioxidant activities of hydrolysates were evaluated using DPPH radical scavenging activity. Peptic hydrolysate exhibited the highest antioxidative activity compared to other hydrolysates. To identify antioxidant peptides, peptic hydrolysate was purified using consecutive chromatographic methods, and antioxidant peptides were identified to be Leu-Leu-Gly-Pro-Gly-Leu-Thr-Asn-His-Ala (1076 Da), and Asp-Leu-Gly-Leu-Gly-Leu-Pro-Gly-Ala-His (1033 Da) by Q-TOF ESI mass spectroscopy. EC 50 values of purified peptides were 189.8 and 167.7 μM, respectively. Antioxidant activities of peptides purified from the rotifer protein hydrolysate were evaluated, with results showing that peptides significantly quenched free radicals.

Functional, bioactive and antioxidative properties of hydrolysates obtained from cod ( Gadus morhua) backbones

Fish protein hydrolysates (FPH) have good and well documented functional properties. Peptides obtained from various fish protein hydrolysates have also shown bioactive and antioxidative activities. The aim of this study was to evaluate how storage and preparation of cod ( Gadus morhua) backbones influence the yield, functionality, bioactivity (CGRP and gastrin/CCK related molecules) and antioxidative properties of fish protein hydrolysates. A series of hydrolysis trials have been carried out using backbones from cod that were initially fresh or frozen and further hydrolysed for different times (10, 25, 45 and 60 min). Use of fresh raw material significantly increased yield of dry FPH, gave lighter and less yellow powders with better emulsification properties. Longer time of hydrolysis gave higher FPH yield, increased degree of hydrolysis and decreased water holding capacity of the powders. Among the hydrolysis times tested, 25 and 45 min hydrolysis demonstrated the best emulsification properties. FPH have potential to enhance product stability by preventing oxidative deterioration. The DPPH scavenging activity showed that antioxidative activity of hydrolysates could be due to the ability to scavenge lipid radicals. The ability of hydrolysates to inhibit iron induced lipid oxidation was not influenced by time of hydrolysis. This work also shows that it is possible to obtain bioactive molecules from cod backbones by protein hydrolysis. The content of bioactive peptides (gastrin/CCK- and CGRP-like peptides) could make the cod hydrolysates useful for incorporation in functional foods.

Effect of degree of hydrolysis on the antioxidant activity of loach (Misgurnus anguillicaudatus) protein hydrolysates

Abstract Loach ( Misgurnus anguillicaudatus ) proteins were hydrolysed by papain and Protamex, the antioxidant activity of loach protein hydrolysates (LPH) was investigated. The results demonstrated that extensive hydrolysis by papain and Protamex led to the browning of the hydrolysates. When the degree of hydrolysis (DH) was 23%, hydrolysates prepared by papain (HA) exhibited the strongest antioxidant activity. The maximum values of the hydroxyl, 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) radical scavenging activities and the reducing power were 56.1%, 95.5%, 2.80 mM and 1.46, respectively. The hydrolysates prepared by Protamex (HB) showed the strongest hydroxyl radical scavenging activity (55.0%) at DH 28%, DPPH radical scavenging activity (92.2%) and ABTS radical scavenging activity (2.81 mM) at DH 23%, and the reducing power (1.17) at DH 33%. At the same DH value, there were significant ( p Industrial relevance Loach has long been employed as a traditional Chinese medicine for the treatment of many kinds of diseases. From our previous work, loach was determined to be a good source of protein (accounts for approximately 17% (w/w) of the body weight). In this work, loach proteins were hydrolyzed by papain and Protamex to specific extent. The effect of DH on the antioxidant activities of hydrolysates was investigated. The results indicated that loach protein hydrolysates were potent antioxidants which were significantly affected by DH. This research is helpful for extensive development of loach product.

ANTIOXIDATIVE ACTIVITY OF PROTEIN HYDROLYSATE FROM ROUND SCAD MUSCLE USING ALCALASE AND FLAVOURZYME

Antioxidative activity of hydrolysates from round scad (Decapterus maruadsi) muscle with degrees of hydrolysis (DH ) of 20, 40 and 60%, prepared using Alcalase (HA) or Flavourzyme (HF), was determined. At the same DH, HF exhibited a higher 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity and reducing power, but a lower Fe2+ chelating ability than HA. HF from isopropanol-defatted muscle with 60% DH was extracted using different solvents, and hexane (E1), dichloromethane (E2), ethyl acetate (E3) and residual (R) fractions were obtained. Among all fractions, E2 and E3 exhibited the highest DPPH radical scavenging activity and reducing power. HF with 60% DH and E2 at 1,000 ppm exhibited antioxidant activity in linoleic oxidation and lecithin liposome systems, and the results were comparable to butylated hydroxytoluene (BHT) at 100 ppm. Therefore, type of proteinase, DH and defatting process prior to hydrolysis exerted an influence on the antioxidative activity of hydrolysates.

Purification and characterization of an antioxidant peptide obtained from tuna backbone protein by enzymatic hydrolysis

To utilize fish processing waste, tuna backbone protein was hydrolyzed using different proteases (alcalase, α-chymotrypsin, neutrase, papain, pepsin and trypsin) for production of antioxidant peptide. Antioxidant activities of hydrolysates were evaluated using lipid peroxidation inhibition assay and direct free radical scavenging activity by using electron spin resonance (ESR) spectrometer. Among hydrolysates, peptic hydrolysate exhibited the highest antioxidant activity compared to other hydrolysates. To identify antioxidant peptide, peptic hydrolysate was purified using consecutive chromatographic methods, and the antioxidant peptide was identified to be VKAGFAWTANQQLS (1519 Da) by Q-TOF ESI mass spectroscopy. The antioxidant activities of antioxidant peptide from tuna backbone protein (APTBP) was evaluated, and the results show that APTBP significantly inhibited lipid peroxidation in linoleic acid emulsion system and also quenched free radicals (DPPH, hydroxyl and superoxide) in a dose-dependent manner. Moreover, APTBP did not show any cytotoxic effect against MRC-5 and ECV304 cell lines.