• Journal of Applied Biological Chemistry
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• v.59 no.3
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• pp.265-271
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• 2016

Recently, liver damage contributes to big percentage of the morbidity and mortality rates worldwide. Excessive intake of alcohol is one of the major causes of liver injury. When liver injury is repeated and becomes chronic, it leads to development of fibrosis and cirrhosis. In the liver, TGF-${\beta}$ is a profibrogenic cytokine, which participates in various critical events cause liver fibrosis. Seahorse (Hippocampus abdominalis) is a common traditional Chinese medicine and has been widely used for centuries. Seahorse has been known to have a variety of bioactivities, such as anti-oxidant, anti-fatigue, and anti-tumor. Peptide is one of the main compounds of seahorse. In this study, we isolated enzymatic hydrolysate from seahorse H. abdominalis by alcalase hydrolysis and investigated the effect of the hydrolysate on liver injury. In the present in vitro studies, the hydrolysate increases cell viability of Chang cells and protects Huh7 cells from ethanol toxicity. In addition, the hydrolysate inhibits TGF-${\beta}$-induced responses. In vivo studies show that the pretreatment of hydrolysate reduces alcohol-induced increases of serum Glutamic oxaloacetic acid transaminase and Glutamic pyruvate transaminase activities and increases liver weight and body weight. These results suggest that seahorse may have a hepatoprotective effect.

• Applied Chemistry for Engineering
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• v.26 no.1
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• pp.111-115
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• 2015

Two new analogs of bestatin were prepared from $\small{D}$-leucine and $\small{D}$-valine in a stereoselective and efficient way. An aminopeptidase inhibitor bestatin shows significant biological effects on immunomodulation and is marketed for the treatment of acute myelocytic leukemia. The key intermediates, trans-oxazolidine methyl esters 2a and 2b, were obtained with more than 20 to 1 stereoselectivity in a one-pot procedure by the three cascade reactions between N-hydroxymethyl protected ${\alpha}$-amino aldehydes (4a and 4b) and phenylsulfonylnitromethane ($PhSO_2CH_2NO_2$) and the following in-situ ozonolysis. Basic hydrolysis of 2a and 2b, and then the peptide coupling with $\small{L}$-Leu-OMe produced the protected derivatives of two new bestatin analogs, 3a and 3b, respectively. The new isobutyl and isopropyl analogs of bestatin (1a and 1b) were produced in overall 51% and 38% yields, respectively, with high stereoselectivity from the corresponding protected ${\alpha}$-amino aldehydes 4 in a six-step process.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.42 no.5
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• pp.417-425
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• 2009

The potential utility of fish scales to the functional food industry has been investigated due to its antioxidant and antihypertensive characteristics. In this study, we report on the reactive oxygen species (ROS) scavenging and angiotensin I converting enzyme (ACE) inhibitory activities of gelatin hydrolysates processed from Branchiostegus japonicus scales, which are also high in protein content (about 46.1%). We prepared the enzymatic gelatin hydrolysates with four proteases (${\alpha}$-chymotrypsin, Alcalase, Neutrase and trypsin) from B. japonicus scale gelatin, which was prepared according to different reaction times, substrate/enzyme ratios and substrate concentrations. The enzymatic hydrolytic degrees of the gelatin increased time-dependently up to 6 hrs, while the Alcalase gelatin hydrolysates showed the highest hydrolysis degrees compared to the others. Furthermore, gelatin hydrolysates of Neutrase and ${\alpha}$-chymotrypsin showed the highest DPPH radical and $H_2O_2$ scavenging activities ($IC_{50}$ value; 9.18 mg/mL and 9.74 mg/mL), respectively. However, the activities were not significant (P<0.05). We also observed that the four gelatin hydrolysates significantly increased ACE inhibitory activities from approximately 20% to 60% (P<0.05), Among them, the Alcalase gelatin hydrolysates showed the higher ACE inhibitory activity ($IC_{50}$ value; 0.73 mg/mL) compared to the others. These results suggest that the enzymatic gelatin hydrolysates prepared from B. japonicus scales may possess a potentially useful function as an ACE inhibitory agent. As such, the utility of B. japonicus scales should be given due consideration for application in the functional food industry.

• Journal of Microbiology and Biotechnology
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• v.21 no.8
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• pp.861-868
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• 2011

A xylanase gene, xyn7c, was cloned from Paenibacillus sp. 12-11, an alkalophilic strain isolated from the alkaline wastewater sludge of a paper mill, and expressed in Escherichia coli. The full-length gene consists of 1,296 bp and encodes a mature protein of 400 residues (excluding the putative signal peptide) that belongs to the glycoside hydrolase family 10. The optimal pH of the purified recombinant XYN7C was found to be 8.0, and the enzyme had good pH adaptability at 6.5-8.5 and stability over a broad pH range of 5.0-11.0. XYN7C exhibited maximum activity at $55^{\circ}C$ and was thermostable at $50^{\circ}C$ and below. Using wheat arabinoxylan as the substrate, XYN7C had a high specific activity of 1,886 U/mg, and the apparent $K_m$ and $V_{max}$ values were 1.18 mg/ml and 1,961 ${\mu}mol$/mg/min, respectively. XYN7C also had substrate specificity towards various xylans, and was highly resistant to neutral proteases. The main hydrolysis products of xylans were xylose and xylobiose. These properties make XYN7C a promising candidate to be used in biobleaching, baking, and cotton scouring processes.

• Journal of Applied Biological Chemistry
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• v.59 no.4
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• pp.285-288
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• 2016

Proteolytic enzymes perform hydrolysis of the peptide bonds in the protein and most commonly use in the industry. Pseudoxanthomonas sp. WD12 and WD32 were previously isolated as protease producers from a rotten wood sample. Here, we report the secreted proteolytic enzymes. The optimum enzyme reaction temperature for the secreted crude enzyme from the strain WD12 and WD32 were $50^{\circ}C$ at pH 9.0 and $45^{\circ}C$ at pH 8.0, respectively. The enzyme activities of both strains were increased by addition of KCl, NaCl, $CaCl_2$ or $MnSO_4$, and decreased by addition of $AgNO_3$, $CuSO_4$, $FeCl_3$ or $AlCl_3$. Secreted enzymes of both strains were most strongly inhibited by addition of $FeCl_3$ or $CuSO_4$. Taken together these results, WD12 could be a candidate strain of industrial alkaline protease production.

• Microbiology and Biotechnology Letters
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• v.30 no.1
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• pp.1-7
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• 2002

Phytase from Escherichia coli WC7 was purified from cell extracts and its molecular mass was estimated to be 45 kDa by SDS-PAGE. Its optimum temperature and pH for phytate hydrolysis was 6$0^{\circ}C$ and pH 5.0, respectively. The enzyme was stable up to 6$0^{\circ}C$ and over broad pH range (pH 2-12). The enzyme had higher affinity for sodium phytate than p-nitrophenylphosphate (pNPP). That is, the apparent Km value for sodium phytate and pNPP were $0.15\pm$0.02 mM and 2.82$\pm$0.05 mM, respectively. The gene encoding the phytase was cloned in E. coli XL1-Blue. Sequence analysis showed an open reading frame of 1241 Up encoding a signal peptide (22 aa) and a mature enzyme (410 aa). WC7 phytase was expressed up to 17.5 U/ml in the transformed E. coli XL1-Blue/pUEP, which was 23-fold higher than the activity from wild strain.

• Food Science of Animal Resources
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• v.29 no.6
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• pp.659-667
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• 2009

Casein is considered to be the main source of protein in milk; therefore, many studies have been conducted to identify casein-derived bioactive peptides and their physiological effects. Casein is inactive within the parent protein but can be liberated by various proteases and enzymatic hydrolysis during microbial fermentation and gastrointestinal digestion. Once absorbed, casein exhibits different bioavailabilities in the body. Specifically, casein-derived peptides function as angiotensin converting enzyme (ACE) inhibitor in the cardiovascular system; thus, they are expected to reduce and prevent hypertension. Additionally, casein-derived peptides behave as opioid-like peptides in the nervous system, which impacts relaxation. These peptides are also expected to modulate various aspects of immune functions. Finally, caseinophosphopeptide (CPP) and glycomacropeptide (GMP) may exhibit a number of nutritional effects such as the absorption of calcium, iron or zinc. Many studies have been conducted to evaluate casein-derived peptides due to their multifunctional properties and the results of these studies have contributed to the development of a wide variety of functional dairy products. The purpose of this paper was to review the generation of bioactive peptides, their absorption and metabolism, and their specific bioactive effects.

• Food Science of Animal Resources
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• v.30 no.2
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• pp.286-290
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• 2010

Angiotensin-converting enzyme (ACE) inhibitory activity and production optimization of ovotransferrin hydrolysates were studied. Ovotransferrin was hydrolyzed by several enzymes (protamex, alcalase, trypsin, pepsin, neutrase, and flavorzyme) and acid (0.03 N HCl). Ovotransferrin hydrolysate reduced ACE activity by 60.2%, 55.8%, and 42.6% when treated with trypsin, acid, and pepsin, respectively. Trypsin was selected for production of peptide having maximum AC inhibitory effect, which was greatest with 7 h hydrolysis. Central composite design determined that optimum composition of ACE inhibitory substances using substrate concentration of 20-35%, temperature of $35-55^{\circ}C$, and pH of 6.0-8.0. The optimum composition was 1% trypsin, substrate concentration of 26.32%, $51.29^{\circ}C$, and pH 6.32. Under this conditions, a maximum ACE inhibitory effect of 69.1% was evident, similar to the predicted value.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.25 no.3
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• pp.229-235
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• 1992

Fish protein hydrolysates(FPH) prepared from defatted mackerel meal by proteases such as complex enzymes, bromelain, alcalase, $\alpha-chymotrypsin,$ trypsin, papain and pepsin were tested for inhibitory activity against angiotensin-I converting enzyme(ACE). Among proteases tested, the hydrolysates obtained from the treatment of complex enzymes or bromelain showed relatively higher activity. ACE inhibitory activity of the hydrolysates increased until hydrolysis of 8 hrs, and was stable by heat treatment for 20min at $100^{\circ}C.$ From the profiles of fractionation of the hydrolysates with Bio-gel P-2, the most active fraction had about MW 1,450 and it's amino acid was abundant in Asp, Glu, Lys, Leu, Val and Ala. $IC_{50}\;(amounts\;of\;inhibitors\;needed\;for\;50\%\;inhibition)$ of the active fraction of the hydrolysates obtained from the treatment of complex enzyme and bromelain was 90 and $130 {\mu}g,$ respectively.

• Journal of Animal Science and Technology
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• v.45 no.2
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• pp.319-326
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• 2003

Angiotensin-I converting enzyme(ACE)inhibitor was isolated from beef by-products. The beef by- product hydrolysates prepared with various proteases were tested for the inhibitory effects against ACE. The proteases used were proteinase A from bakers yeast, protease type ⅩIII fungal and thermolysin. The maximum inhibitory effect was observed after hydrolysis for 12hrs(beef heart) and 24hrs(beef spleen), respectively. After gel filtration, IC50 value was 0.37mg/ml in beef heart and 1.84mg/ml in beef spleen. After RP-HPLC, the IC50 value of peak 1, peak 2, peak 3 and peak-4 were 0.28mg/ml, 0.26mg/ml, 0.25mg/ml and 0.35mg/ml, respectively. In the results of amino acid composition of peak 1, peak 2, peak 3 and peak 4, it was observed that peak 1 was consisted mainly of glycine and methionine, peak 2 was proline, cystine and methionine, peak 3 was proline and peak 4 was alanine, methionine and leucine. In conclusion, beef heart hydrolysate treated with thermolysin+ proteinase A was shown to have the highest inhibitory effect for 12hrs incubation at 37$^{\circ}C$.