• Food Science of Animal Resources
• /
• v.38 no.4
• /
• pp.780-793
• /
• 2018

This study was conducted to compare the anti-allergic effects of a whey protein concentrate (WPC) and WPC hydrolysate. WPC hydrolysate was prepared using enzymatic digestion for 8 h with trypsin and ${\alpha}$-chymotrypsin, after which it was freeze-dried. The allergic parameters assessed in rat basophilic leukemia (RBL)-2H3 cells were degranulation and release of ${\beta}$-hexosaminidase, release of tumor necrosis factor $(TNF)-{\alpha}$, and changes in the expression of $IL-1{\beta}$, IL-4, and IL-10 by real time polymerase chain reaction (PCR). During preparation of the WPC hydrolysate, hydrolysis increased rapidly from 0 to 10 min and then gradually increased slowly from 1 h onwards, achieving a final degree of hydrolysis of 78.50%. The SDS-PAGE analysis revealed a reduction in the intensity of several protein bands in the WPC hydrolysate compared to the WPC. IgE-induced ${\beta}$-hexosaminidase release from RBL-2H3 cells was decreased to a higher degree following treatment with the hydrolysate compared to WPC treatment. W500 ($500{\mu}g/mL$ WPC) showed the least inhibition of ${\beta}$-hexosaminidase release, but there was no significant difference between W500 and W1000 ($1,000{\mu}g/mL$) (p<0.05). H1000 ($1,000{\mu}g/mL$ WPC hydrolysate) inhibited ${\beta}$-hexosaminidase release by 39%. Compared to the control, treatment with H1000 decreased $TNF-{\alpha}$ secretion to 11.87 pg/mL. The gene expression levels of IL-1${\beta}$, IL-4, and IL-13 were all significantly decreased in hydrolysate (p<0.05). In the case of $IL-1{\beta}$ and IL-4, the expression levels in W1000 treated cells were decreased by 73.67% and 65%, respectively, and that of IL-13 was decreased by 66.43% compared to the control.

• Journal of the Korean Chemical Society
• /
• v.47 no.4
• /
• pp.363-369
• /
• 2003

Selenized yeast (Se yeast) containing $0.1{\%}$(w/w) of selenium was obtained when the yeast was incubated at a selenium concentration of 1$1.14{\times}10_-3 M$ in rich medium. After washing several times, the inorganic selenium on the cell wall was confirmed with MBRT. There was no indication of inorganic selenium on the cell wall when the blue color in MBRT was stayed for 15 minutes. The selenized yeast was sonicated, then the selenium contained protein was obtained after salting out by ammonium sulfate at the concentration $80{\%}$ saturation. The seven protein bands were seperated by SDS-PAGE and the selenium concentration in protein was measured by ICP-AES. Analytical data showed that the large expressed protein band contained a relatively large amount of selenium. The proteins of the 47kDa was contained the concentrations of 69.5 ${\mu}$ Se/g of most many content. The protein (47 kDa) was seperated from PVDF membrane by tank-electroblotting. The isolated protein was hydrolyzed under acid condition and reacted with PITC. The derivatives of amino acids were analyzed by HPLC and compared with the data obtained from regular yeast. The resulting selenium-yeast was analyzed with the selenomethionine concentration of $2{\%}$ comparaed with general amino acids. The goal of this study is to analyze the selenium concentration in protein bands and measure the degree of biotransformation of selenomethionine in a specific protein.

• Applied Biological Chemistry
• /
• v.35 no.1
• /
• pp.23-29
• /
• 1992

Bromelain was purified from Korean pineapple, Ananas comosus, L. The enzyme was purified about 21 fold by DEAF-cellulose ion-exchange chromatography and gel filtration on Sephadex G-150. Purified enzyme was confirmed as active single band by polyacrylamide electrophoresis and the molecular weight was estimated to be about 22,000 by SDS-PAGE. The optimum pH and temperature were 6.0 and $60^{\circ}C$, respectively. The range of its stability to the pH and temperature were respectively 5.0 to 7.0 and below $50^{\circ}C$. It was found that $Mn^{2+}$ increased the enzyme activity, whereas $Mg^{2+}\;and\;Fe^{2+}$ decreased it abruptly. The purified enzyme was inhibited by p-chloromercuribenzoic acid, indicating that reactive SH groups are required for the enzyme activity. The reaction of the enzyme followed typical Michaelis-Menten kinetics with Km value of $5.747{\times}10^{-4}\;M\;and\;Vmax\;of\;131.58\;{\mu}g/min$ for casein. When meat was treated with the enzyme, free soluble nitrogen and amino acid nitrogen increased as enzyme concentration increased.

• Current Research on Agriculture and Life Sciences
• /
• v.14
• /
• pp.111-122
• /
• 1996

S. chibaensis J-59 produced an extracellular xylanase in a CSL medium composed of 1.5% com steep liquor, 0.1% $MgSO_4{\cdot}7H_2O$, 0.012% $CoCl_2{\cdot}6H_2O$, and 0.15% glucose containing xylan. but it did not produce in the culture medium containing xylose. The production of enzyme reached to a maximum level (0.83 uints/ml) when bacteria were cultured in 2.5 l jar fermentor for 48hrs at $30^{\circ}C$ and pH 7.0. Furthermore, S. chibaensis J-59 produced an intracellular glucose isomerase in a medium containing xylan and/or xylose. Xylanase was purified 29-fold over the culture supernatants of S. chibaensis J-59 by ammonium sulfate fractionation, chromatography on DEAE-Sephadex A-50, and gel filtration on Sephadex G-200. The purified enzyme is a monomeric enzyme with a native molecular mass of 25 kDa and a subunit molecular mass of 25 kDa. The purified enzyme requires $Mg^{2+}$ for activity, $Ca^{2+}$, $Co^{2+}$ is not an inhibitor but inhibit by $Fe^{3+}$, $Hg^{2+}$, and $Cu^{2+}$, sodium dodecyl sulfate, N-bromosuccinide. Pattern of hydrolysis demonstrated that the xylanase was an endo-splitting enzyme able to break down birchwood xylan at random giving xylobiose, xylotriose and xylotetrose as the main end products.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.46 no.4
• /
• pp.435-441
• /
• 2017

The present study was carried out to evaluate the applicability of Tenebrio molitor larvae (mealworm) as a health functional food material in order to contribute to the development of the domestic insect industry and health functional food industry. Protein hydrolysates were prepared from mealworm powder by enzymatic hydrolysis using five different proteases (alcalase, bromelain, flavourzyme, neutrase, and papain), and the hydrolysates were then tested for their antioxidant activities. Based on available amino group contents and sodium dodecyl sulphate-polyacrylamide gel electrophoresis analyses, mealworms treated with alcalase ($4,781.39{\mu}g/mL$), flavourzyme ($5,429.35{\mu}g/mL$), or neutrase ($3,155.55{\mu}g/mL$) for 24 h showed high degree of hydrolysis (HD) value, whereas HD values of bromelain ($1,800{\mu}g/mL$) and papain-treated ($1,782.61{\mu}g/mL$) mealworms were much lower. Protein hydrolysates showing high HD values were further separated into > 3 kDa and ${\leq}3kDa$ fractions by a centrifugal filter system and then lyophilized, and the production yields of the low molecular weight protein hydrolysates (${\leq}3kDa$) by alcalase, flavourzyme, and neutrase were 42.05%, 26.27%, and 30.01%, respectively. According to the RC_{50} values of the protein hydrolysates (${\leq}3kDa$) obtained from three different antioxidant analyses, all three hydrolysates showed similar antioxidant activities. Thus, alcalase hydrolysates showing the highest production yield of low molecular weight protein hydrolysates were further tested for their inhibitory effects on peroxidation of linoleic acid by measuring thiobarbituric acid values, and the results show that peroxidation of untreated linoleic acid increased dramatically during 6 days of incubation. However, pretreatment with the hydrolysates ($100{\sim}800{\mu}g/mL$) significantly inhibited linoleic acid peroxidation in a dose-dependent manner over 6 days.

• The Korean Journal of Ecology
• /
• v.16 no.4
• /
• pp.515-526
• /
• 1993

In order to study the ecotypic variation of Rohinia pseudo-acacia L. distributed in southern area of Korean peninsula, 15 local populations(Daejin, Sokcho, Kangneung, Mt. Surak, Hongcheon, Kwangneung, Namhansanseong, Chungju, Yesan, Andong, Jeonju, Dalseong, Changweon, Mokpo and Wando), located from $34^{\circ}18'N\;to\;38^{\circ}36'N$, were selected based on the latitudes and geographical distances. Seeds of these populations were collected and protein contents of seeds and their band patterns were investigated. The seed proteins of all populations were electrophoresed on SDS-polyacrylamide gel. Total number of protein bands were 35, whose molecular weights ranged from 17, 258 daltons to 142, 232 daltons. The number of bands of seed proteins was 23 in Dalseong and Hongcheon and was 32 in Daejin and Sokcho, showing an increasing tendency in the number of bands as the latitude goes high. The local populations were classified into 3 local types based on protein analysis: the middle north east coastal type(Daejin, Sokcho. Kangneung), the central type (Mt. Surak, Hongcheon, Kwangneung, Namhansanseong, Chungju) and the southern type(Yesan, Andong, Jeonju, Dalseong, Changweon, Mokpo, Wando). According to the results of cluster analysis by UPGMA based on the similarity index(c0efficient of Jaccard) of the patterns, 3 local types were subdivided further into 6 types: the middle north east coastal type(Sokcho, Kangneung), the north central type I (Mt. Surak, Hongcheon), the north central type II (Narnhansanseong, Chungju, Daejin), the north central type III (Kwangneung), the south central type (Yesan, Dalseong, Jeonju) and the southern type(Andong, Changweon, Mokpo, Dalseong, Wando). The No. 12 band of the separated seed proteins showed the highest colored density in the preparations from all the populations. The No. 11~13 and No. 23~28 bands also showed high densities. As a whole, southern type populations (Changweon, Mokpo, Wando) showed high protein contents and high colored density. Total protein contents of the seeds in each population were variable from 9. 68mg / g (Mt. Surak) to 17.30mg/g (Jeonju), showing an increasing trends toward low latitudes.

• KSBB Journal
• /
• v.19 no.1
• /
• pp.50-56
• /
• 2004

The aim of this work was to investigate the purification and characterization dixoygenases isolated from Delftia sp. JK-2, which could utilize aniline, benzoate, and p-hydroxybenoate as sole carbon and energy source. Catechol 1,2-dioxygenase (C1, 2O), catechol 2,3-dioxygenase(C2, 3O), and protocatechuate 4,5-dioxygenase(4,5-PCD) were isolated by benzoate, aniline, and p-hydroxybenzoate. In initial experiments, several characteristics of C1 ,2O, C2, 3O, and 4,5-PCD separated with ammonium sulfate precipitation, DEAE-sepharose, and Q-sepharose were investigated. Specific activity of C1 ,2O, C2, 3O, and 4,5-PCD were approximately 3.3 unit/mg, 4.7 unit/mg, and 2.0 unit/mg. C1 ,2O and C2, 3O demonstrated their enzyme activities to other substrates, catechol and 4-methylcatechol. 4,5-PCD showed the specific activity to the only substrate, protocatechuate, but the substrates(e.g., catechol, 3-methylcatechol, 4-methylcatechol, 4-chlorocatechol, 4-nitrocatechol) did not show any specific activities in this work. The optimum temperature of C1, 2O, C2, 3O, and 4,5-PCD were 30$^{\circ}C$, and the optimal pHs were approximately 8, 8, and 7, respectively. Ag$\^$+/, Hg$\^$+/, Cu$\^$2+/ showed inhibitory effect on the activity of C1, 2O and C2, 3O, but Ag$\^$+/, Hg$\^$+/, Cu$\^$2+/, Fe$\^$3+/ showed inhibitory effect on the activity of 4,5-PCD. Molecular weight of the C1, 2O, C2, 3O, and 4,5-PCD were determined to approximately 60 kDa,35 kDa, and 62 kDa by SDS-PAGE.

• Korean Journal of Food Science and Technology
• /
• v.29 no.6
• /
• pp.1248-1254
• /
• 1997

To measure antioxidative activities, the various extracts from RVS (Rhus Verniciflua Stokes) were tried out with either DPPH or thiocyanate method. Also we used the GO (Glucose Oxidase) 20 mU/mL hydroxyl radical system in mouse whole brain cell culture. Chloroform, n-hexane or ethanol were used as extract solutions which had different polarity respectively. In DPPH and thiocyanate method, the antioxidative activities of the crude ethanol extracts were stronger than other extracts. The crude ethanol extracts were fractionated 5 peaks by glass column. Among of them, antioxidative activity of peak II $(P_{II})$ was shown stronger than other fractions, a little for peak III $(P_{III})$ and peak IV $(P_{IV})$, and none for peak I $(P_I)$ and Peak V $(P_V)$. In the antioxidative effects of crude ethanol extracts (30 mg/mL), cell viabilities were evaluated $1\;{\mu}L\;(297\;{\mu}g/mL)$, $2\;{\mu}L\;(588\;{\mu}g/mL)$ of crude ethanol extracts 59%, 68% respectively. $10\;{\mu}L\;(2,727\;{\mu}g/mL)$ addition of crude ethanol extracts had 95% cell viabilities, 0.01% significant, comparing control. In addition, the compounds related to antioxidative effect of crude ethanol extract might be glycoproteins by means of SDS-PAGE. Comparison to antioxidative effects between several antioxidants (ascorbic acid, ${\alpha}-tocopherol$, catalase) $273\;{\mu}L/mL$ addition of crude ethanol extracts corresponds to $1\;{\mu}g/mL$ catalase in antioxidative effects.

• Parasites, Hosts and Diseases
• /
• v.33 no.2
• /
• pp.107-116
• /
• 1995

Two hybridoma cell lines against Cwptosporinium possum oocysts nFRl-CN911 were produced. The isotype of these 2 monoclonal antibodies (mAbs) was IgG2b (lE7.2) and and IgM (C6). Enzyme immuno-transfer blotting analysis showed that 157.2 reacted specifically to 36 kDa protein and C6 reacted to 67 and 70 kDa proteins. C. pcnlum was bound specifically to the surface region of oocysts by these mobs. No cross-reactivity was observed with tachyzoites of ToxopLosma gonnii and oocysts of Eimeria zuernii,5. bouis and E. canadensis of bovine origin. The indirect immunofluorescence assay (IIF) using mAb C6 was successful with counterstain. With the IIF using mob C6, oocysts appeared as 3 to $5{\mu}m$ spherical objects fluorescing bright apple green against a reddish dark background. The IIF using mAb C6 was agreed in specificity and sensitivity with those of a commercial diagnostic kit. These results demonstrated that the produced mAbs were specific to C. parvum and that the mAb C6 could be used for diagnosis of cryptosporidiosis.

• Journal of Microbiology and Biotechnology
• /
• v.13 no.6
• /
• pp.926-936
• /
• 2003

Overexpression of human Bcl-2 protein in recombinant Chinese hamster ovary (rCHO) cells producing humanized antibody (SH2-0.32) considerably suppressed sodium butyrate (NaBu)-induced apoptosis during batch culture by using commercially available serum-free medium, which extended the culture longevity. Due to the extended culture longevity provided by the anti-apoptotic effect of Bcl-2 overexpression, the final antibody concentration of 14C6-bcl-2 culture (Bcl-2 high producer, $23\;\mu\textrm{g}\;ml^{-1}$) was 2 times higher than that of the $SH2-0.32-{\Delta}bcl-2$ culture (cells transfected with bcl-2-deficient plasmid, $10.5\;\mu\textrm{g}\;ml^{-1}$) in the presence of NaBu. To determine the effect of NaBu/Bcl-2 overexpression on the molecular integrity of protein products, antibodies purified from 14C6-bcl-2 and $SH2-0.32-{\Delta}bcl-2$ cultures in the presence of NaBu were characterized by using various molecular assay systems. For comparison, antibody purified from the parental rCHO cell culture (SH2-0.32) in the absence of NaBu was also characterized. No significant changes in molecular weight of antibodies could be observed by SDS-PAGE. From GlycoSep-N column analysis, it was found that the core oligosaccharide structure ($GlcNAc_2Man_3GlcNAc_2$) was not affected by NaBu/Bcl-2 overexpression, while the microheterogeneity of N-linked oligosaccharide structure was slightly affected. Compared with the antibody produced in the absence of NaBu, the proportion of neutral oligosaccharides was increased from 10% (14C6-bcl-2) to 16% ($SH2-0.32-{\Delta}bcl-2$) in the presence of NaBu, which was accompanied by the reduced proportion of acidic oligosaccharides, especially of monosialylated and disialylated forms. The changes in microheterogeneous oligoformal structures of antibody in turn affected the mobility of antibody isoforms in isoelectric focusing (IEF), resulting in the occurrence of some more basic antibody isoforms produced in the presence of NaBu. However, the antigen-antibody binding properties were not changed by alteration of glycosylation pattern. The competitive enzyme-linked immunosorbent assay (ELISA) showed that the antibody produced by NaBu/Bcl-2 overexpression maintained its antigen-antibody binding properties with binding affinity of about $2.5{\times}10^9{\;}M^{-1}$. Taken together, no significant effects of NaBu/Bcl-2 overexpression on the molecular integrity of antibodies, produced by using serum-free medium, could be observed by the molecular assay systems.