• KSBB Journal
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• v.15 no.6
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• pp.635-643
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• 2000

To compare the antioxidative activities of fish skin and bovine skin gelatin hydrolysate, gelatin hydrolysates from Alaska pollack and bovine skin were prepared by various enzymatic hydrolysis methods (1st step, Alcalase; 2nd step, pronase E; 3rd step, collagenase) using a continuous three-step membrane reactor. The molecular weight distributions of the 1st, 2nd and 3rd step hydrolysates were 7∼10 kDa, 2∼5 kDa and 0.7∼0.9 kDa, respectively. The antioxidative activity of fish skin gelatin hydrolysate was stronger than that of bovine skin gelatin hydrolysate, and in particular, both of 2nd step hydrolysates showed more antioxidative activity than hydrolysates of any other step. The optimum antioxidative activity concentration of the 2nd step hydeolysates of fish and boving skin were 1% (w/w) in a linoleic acid water-alcohol emulsion. In cultured cells exposed to t-butyl hydroperoxide (t-BHP), the 2nd step hydrolysate of fish skin gelatin delayed cell death most. These results suggest that the antioxidative activity of fish skin gelatin hydrolysate is higher than that of bovine skin gelatin hydrolysate because of their different amino acid contents.

• Applied Biological Chemistry
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• v.37 no.2
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• pp.85-93
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• 1994

The fish skin gelatin hydrolysates were produced using a continuous two-stage membrane (MWCO 10,000, MWCO 5,000) reactor, and molecular weights, amino acids and functional properties of the hydrolysates were investigated. The major molecular weights distribution of the major fractions were $8{\sim}10\;KDa$ and $4.5{\sim}6.5\;KDa$ in the 1st-step hydrolysates, $2{\sim}6\;KDa$ and $0.5{\sim}2\;KDa$ in the 2nd-step hydrolysates. Among the amino acids in the hydrolysates, glycine, proline, serine, alanine, hydroxyproline, glutamic acid and aspartic acid having sweet taste were responsible for $68{\sim}72%$ of the total amino acids. But valine, methionine, isoleucine, leucine, phenylalanine and histidine having a bitter taste were only $23{\sim}25%$ Taste evaluations show that the gelatin hydrolysates have a brothy and sweet taste, 2nd-step hydrolysate have more a favorable taste than 1st-step hydrolysate. The hydrolysates were completely soluble and clear over the entire pH range. Moisture sorption at intermediate water activities of the 2nd-step hydrolysate was much higher than the unmodified fish skin gelatin, but foaming and emulsification properties were poor. Buffer capacity of the 2nd-step hydrolysate was higher than the fish skin gelatin and 1st-step hydrolysate, while viscosities of the hydrolysates were lower than the fish skin gelatin.

• Fisheries and Aquatic Sciences
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• v.14 no.1
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• pp.1-10
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• 2011

This study was conducted to obtain hydrolysates with potent antioxidative activity from rockfish skin gelatin. Gelatin was extracted under high temperature/high pressure using a two-step enzymatic hydrolysis with commercial enzymes such as Alcalase, Flavourzyme, Neutrase, and Protamex. The second rockfish-skin gelatin hydrolysate (SRSGH) was prepared by further incubating the first gelatin hydrolysate (FRSGH), which had been hydrolyzed with Alcalase for 1-h (FRSGH-A1), with Flavourzyme for 2-h (SRSGH-F2). The second gelatin hydrolysate showed higher antioxidative activity of 3.72 as measured by a Metrohm Rancimat and superior angiotensin I-converting enzyme (ACE) inhibiting activity of 0.82 mg/mL. Compared with the gelatin, the relative proportion in SRSGH-F2 was markedly decreased in the 100-kDa peak, whereas it was increased in that less than 100-kDa. The amino acid composition of SRSGH-F2 was rich in glycine (25.9%), proline (10.8%), alanine (9.1%), and glutamic acid (9.1%). In contrast, it was poor in cystine (not detected), methionine (1.6%), tyrosine (0.4%), hydroxylysine (0.9%), and histidine (0.9%). In recent years, demand for natural functional foods has been increasing, and SRSGH-F2 can be used as a functional food ingredient in the food industries. However, further detailed studies on SRSGH-F2 with regard to its antioxidant activity in vivo and the various antioxidant mechanisms are needed.

• Journal of the Korean Society of Food Science and Nutrition
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• v.21 no.4
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• pp.398-406
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• 1992

In order to develop a new flavourant using the fish skin gelatin, the proteolytic renditions for the gelatin hydrolysate of the alkali (B-type) and Alcalase (E-type) pretreated flounder (Limanda aspera) skin gelatin were investigated, and some physical properties, molecular weight and amino acid compositions of the hydrolysates were, also, compared with each other. The proteolytic conditions of the gelatins (B-type and E-type) by trypsin were as follows : reaction temperature, 55$^{\circ}C$ : pH, 9.0 : enzyme concentration, 0.1% : re-action time, 4hrs for B-type and 1 hr for E-type. The degrees of hydrolysis of the B-type and E-type gelatin un-der the renditions stated above were 63% and 82%, respectively. The rnajor molecular weights of the hydrolysates were 15,000 dalton for B-type and 12,400 dalton for E-type. Among the amino acids in the hydrolysates, glycine, alanine, proline, hydroxyproline and serine having a sweet taste were responsible for 57% of the total amino acid. But valine, leucine, phenylalanine, tyrosine, methionine, arginine and histidine having a bitter taste were only 18%.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.29 no.2
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• pp.246-255
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• 1996

To develop a natural antioxidative peptide, the gelatin was extracted from fish (Yellowfin sole) skin by hot $water(50^{\circ}C)$ extraction method and hydrolyzed with Alcalase, pronase and collagenase through a continuous 3-step membrane reactor. Each step enzymatic hydrolysates were determined the antioxidative activity and their synergistic effects, compared with $\alpha-tocopherol$ and butylated hydroxytoluene (BHT). Also, we tried to investigate the antioxidative disposition of peptide which was successfully separated by gel filtration, ion-exchange chromatography, and HPIC in cultured rat hepatocytes intoxicated with tert-butyl hydroperoxide (TBHP). Second step enzymatic hydrolysate (SSEH) among all hydrolysates and $\alpha-tocoperol$ was showed the strongest antioxidative activity. The optimum concentration of antioxidative activity for SSEH was $1\%(w/w)$ in linoleic acid. The synergistic effects were increased in using the hydrolysate with tocopherol and BHT. In the presence of the peptide isolated from SSEH, supplemented hepatocytes exposed to TBHP showed that delayed cell killing and decreased significantly the lipid peroxidation, compared with hepatocytes not cultured with isolated peptide.

• Fisheries and Aquatic Sciences
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• v.14 no.3
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• pp.168-173
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• 2011

The purpose of this study was to obtain a fraction with high antioxidative activity from second rockfish gelatin hydrolysates (SRSGHs), which were hydrolyzed with Alcalase and Flavourzyme through ultrafiltration membranes with serial digestions for 1 and 2 h, respectively, and to investigate the feasibility of this fraction as a potential functional food ingredient. Among various fractions that were ultrafiltered from the SRSGH with four types of membrane (1, 5, 10, and 30 kDa), the SRSGH-III fraction, which permeated the 10 kDa membrane but not the 5 kDa membrane, showed the highest antioxidant activity (protection factor=5.13) and angiotensin-I-converting enzyme-inhibiting activity ($IC_{50}$=0.82 mg/mL). These results suggest that the SRSGH-III fraction from the SRSGH can be used as a functional food ingredient. However, further studies examining its antioxidant activity in vivo as well as the different antioxidant mechanisms are needed.

• Applied Biological Chemistry
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• v.37 no.2
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• pp.130-141
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• 1994

A continuous two-stage membrane (1st-SCMR, MWCO 10,000; 2nd-SCMR, MWCO 5,000) reactor was developed and optimized for the production of fish skin gelatin hydrolysate with different molecular size distribution profiles using trypsin and pronase E. The optimum operating conditions in the 1st-step membrane reactor using trypsin were: temperature, $55^{\circ}C$ ; pH 9.0; enzyme concentration, 0.1 mg/ml; flux, 6.14 ml/min; reaction volume, 600 ml; and the ratio of substrate to trypsin, 100 (w/w). After operating for 1 hr under the above conditions, 79% of total amount of initial gelatin was hydrolysed. In the 2nd-step using pronase E under optimum operating conditions[temperature, $50^{\circ}C$ ; pH 8.0; enzyme concentration, 0.3 mg/ml; flux, 6.14 ml/min; reaction volume, 600 ml; and the ratio of substrate to pronase E, 33 (w/w)], the 1st-step hydrolysate was hydrolysed above 80%. Total enzyme leakages in the 1st-step and 2nd-step membrane reactors were about 11.5% at $55^{\circ}C$ for 5hrs and 9.0% at $50^{\circ}C$ for 4 hrs, respectively. However, there was no apparent correlation between enzyme leakage and substrate hydrolysis. The membrane has a significant effect on activity lose of trypsin and pronase E activity for 1 hr of the membrane reactors operation. The loss of initial activity of enzymes were 34% and 18% in the 1st-step and 2nd-step membrane reactor, whereas were 23% and 10% after operating time 3 hr in the 1st-step and 2nd-step membrane reactor lacking the membrane, respectively. The productivities of 1st-step and 2nd-step membrane reactor for 8 times of volume replacement were 334 mg and 250 mg per mg enzyme, respectively.

• Applied Chemistry for Engineering
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• v.5 no.4
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• pp.681-697
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• 1994

The enzymatic hydrolysate of gelatin extracted from fish skin was fractionated and recycled through the membrane reactor according to the molecular weight for the purpose of using as functional material. In addition, the enzymatic hydrolysis conditions of gelatin, enzyme stability by membrane and mechanical shear, and effect on the long-term operational stability of the recycle membrane reactor were investigated. Using the pH-drop technique, Alcalase, pronase E and collagenase were identified as the most suitable enzymes for the hydrolysis of fish skin gelatin. The optimum hydrolysis conditions in the 1st-step membrane reactor(1st-SMR) by Alcalase were enzyme concentration 0.2mg/ml, substrate-to-enzyme ratio(S/E) 50(w/w), $50^{\circ}C$, pH 8.0, reaction volume 600ml and flow rate 6.14ml/min. In the 2nd-SMR by pronase E were enzyme concentration 0.3mg/ml, S/E 33(w/w), $50^{\circ}C$, pH 8.0, reaction volume 600ml and flow rate 6.14ml/min. In the case of 3rd-SMR, enzyme concentration 0.1mg/ml, S/E 100(w/w), $37^{\circ}C$, pH 7.5, reaction volume 600ml and flow rate 10ml/min. Decreased enzyme activities by mechanical shear and membrane were 30% and 15% in the 1st-SMR, were 14% and 5% in the 2nd-SMR, and 18% and 8% in the 3rd-SMR, respectively. Under the optimum conditions, the degree of hydrolysis in the 1st, 2nd and 3rd-SMR were 3.5%(Kjeldahl method, 87%), 3.1%(77%) and 2.7%(70%), respectively. The productivity of hydrolysate in the continuous three-step membrane reactor was 430mg per enzyme(mg) for 10 times of volume replacements.