• Korean Journal of Food Science and Technology
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• v.32 no.5
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• pp.1079-1086
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• 2000

Pectinesterase inhibitor(PEI) of ripened kiwi fruit(Actinidia chinensis) was separated with affinity chromatography using CNBr-activated Sepharose 4B being covalently bound by orange pectinesterse(PE). The affinity resin strongly and selectively bound PEI, which could be eluted in high yield as a single peak by pH 9.5 without loss of inhibitory activity. The separated PEI had maintained almost inhibitory activity at $-25^{\circ}C$ and $5^{\circ}C$ during 30 days but lost it at room temperature in 4 weeks. The PEI possessed a molecular weight of 16.6 KDa, as estimated by 12.5% SDS-PAGE. PEI had optimum pH of 7.5, optimum temperature of below $10^{\circ}C$ and stability up to $70^{\circ}C$. Also, optimum inhibitory activity for PEI was obtained in 0.2 M NaCl of substrate solutions. The kind of inhibition on tomato pectinesterase was found to be noncompetitive, using citrus pectin as substrate. Fresh orange juice added with crude PEI extracts maintained almost the same cloud stability as pasteurized juice. In case of apple juice, the addition of crude PEI extracts to apple juice had decrease of L-ascorbic acid with nearly no effect on cloud loss.

• Food Science and Preservation
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• v.12 no.5
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• pp.489-495
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• 2005

Polyphenol oxidase (PPO) from Burdock (Arctium lappa L.) was purified and characterized. Purification of polyphenol oxidase was achieved by ammonium sulfate precipitation, Phenyl-sepharose CL-4B hydrophobic chromatography and Sephadex G-100 gel filtration chromatography. The molecular mass of the purified PPO was estimated to be 30 kDa by SDS polyacrylamide gel electrophoresis. In a substrate specificity, maximum activity was achieved with chlorogenic acid, followed by catechol and catechin. Whereas, there was low activity with hydroquinic acid, resorcinol or tyrosine. The optimum pH and temperature for enzyme activity were 7.0 and 35$\circC$ with catechol, respectively. The enzyme was most stable at pH 7.0 while unstable at acidic and alkaline pH. The enzyme was stable when heated to 40$\circC$. But heating at 50$\circC$ for more than 30 min caused 50% loss of activity. Ascorbic acid, L-cystein and $Cu^{2+}$ inhibited the activity of pholyphenol oxidase.

• Journal of Aquaculture
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• v.11 no.2
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• pp.271-278
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• 1998

To clarify characteristics of yolk protein of abalone, yolk protein was purified from the ovarian egg extracts of mature female Haliotis discus hannai by a gel chromatography of sepharose CL-4B. From the results of immuno-electrophoresis and Ouchterlony's diffusion test to male and female sera and ovarian egg extracts using antibodies raised against mature female and male sera and male sera and ovarian egg extrascts, it was identified that the mature female serum had female specific serum protein and its antigenecity shared with ovarian egg extracts. A single type of yolk protein was purified from ovarian egg extracts, and it was composed of two subunits. Their molecular weights were estimated to be approximately 166 KDa and 113 KDa by SDSPAGE. The antiserum against yolk proteins cross-reacted with a mature female specific serum protein and extracts of hepatopancreas of vitellogeing females, but did not reacted with extracts of hepatopancreas of mature male.

• Korean Journal of Pharmacognosy
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• v.24 no.3
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• pp.203-212
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• 1993

To find antitumor components in the hot water extract of the cultured mycelia of Ganoderma lucidum, protein-bound polysaccharides were purified and fractionated (Fr. I-V) by DEAE-cellulose ion exchange column chromatography and Sepbarose CL-4B gel filtration. When a dose of 20 mg/kg/day of each was, i.p., injected into ICR mice, the inhibition ratios against the solid form of sarcoma 180 were $64.2{\sim}75.8%$. The antitumor component was examined for immunological activity. It increased the amount of superoxide anion released by induced macrophages in peritoneal cavity to 1.8 times and the count of hemolytic plaque-forming cells (PFC) was increased to 4.4 times as compared with those of the control group. It contained 68.6% polysaccharide which consisted of mannose, glucose, galactose, fucose and xylose and 5.1% protein consisting of 17 amino acids. The contents of hexosamine were 0.78%. The molecular weight of Fr. V that showed the highest antitumor activity was $5.8{\times}10^4$ dalton by Sepharose CL-4B gel filtration. It was named lucidan.

• Journal of Ginseng Research
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• v.24 no.3
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• pp.128-133
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• 2000

A high molecular (more than 10 kDa) fraction, showing mitogenic and comitogenic activities in spleen cells of mouse, was isolated from water extract of ginseng. The crude protein substance prepared by 80% (NH$_4$)$_2$SO$_4$ precipitation from this fraction was purified and isolated by DEAE Sepharose column chromatography. Among the fractions eluted, it was found that four kinds of fractions eluted with 0 to 1 M NaCl gradient were glycoproteins, which induced proliferation of spleen cells and increased NO production in macrophages. Among them, F-2 fraction, which contained 35.9% protein,49.4% neutral sugar and 12.5% uronic acid, was found to show mitogenic activity as strong as that of LPS (lipopolysaccharide) at a concentration of 100 $\mu\textrm{g}$/ml and to remarkably stimulate NO production by murine macrophages at a concentration of 500 $\mu\textrm{g}$/ml. When F-2 is deproteinized, the mitogenic activity of F-2 was decreased significantly to 70.9% as compared with that of F-2. This results suggests that the protein moiety of F-2 may play an important role in immunomodulating activity of glycoprotein from the root of Panax ginseng.

• Korean Journal of Food Science and Technology
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• v.34 no.2
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• pp.343-345
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• 2002

The study was investigated hot-water soluble on waxy black and waxy rice flours according to gelatinization temperatures and heating time. The hot-water soluble contents of both samples were increased during heating time at $90^{\circ}C$. The waxy black and waxy rice flours solutions according to gelatinization temperatures ($80^{\circ}C,\;90^{\circ}C,\;95^{\circ}C\;and\;98^{\circ}C$) were heated for 10, 20 and 30 min. Determination of elution patterns of rice flours was used by Sepharose CL-2B column. As a result, the elution patterns were eluted the most in void volume ($V_0$) and the elution patterns of the hot-water solubles were increased according to heating time and gelatinization temperatures.

• Food Engineering Progress
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• v.13 no.4
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• pp.326-334
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• 2009

Blood coagulation and aggregation of platelet are crucial events in the pathogenesis of various ischemic diseases. The substance which can prevent blood coagulation and platelet aggregation was extracted from wood ear mushroom (Auricularia auricular-judae) and its anticoagulation activity was investigated. The dried A. auricular-judae was extracted with 0.1 N NaOH and its supernatant was further extracted with methanol and ethanol followed by $H_{2}O$. The resulting methanol soluble fraction showed significant antithrombotic activity in activated partial thromboplastin time, thrombin time, and prothrombin time assays with values of 100, 124, and 54 sec, respectively. This active substance was purified with DEAE-Sepharose CL6B and Sephacryl 400-HR and was found to be polysaccharide with the average molecular weight of over 150 kDa. This polysaccharide was xyloglucomannan of which the main component was mannose, and its anticoagulant activity was mostly mediated by inhibition of thrombin activity.

• KSBB Journal
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• v.14 no.1
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• pp.96-102
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• 1999

Marine bacterium Bacillus cereus ASK202 produced an extracellular agarase (E.C.3.2.1.81) which showed a high level of enzyme activity in the presence of agar and agarose. In the optimal culture conditions, the agarase production increased 7.7 folds compared with the one obtained from the basal medium. Agarase production reached upto 160 units/L after 24hr of cultivation in a modified marine medium at $25^{\circ}C$. The degree of purification increased 31.5 folds with 27.8% yield through freeze drying, DEAE Sepharose CL-6B and Superose 6HR 10/30 column chromatography. The molecular weight of the purified agarase was determined to be 90,000 daltons by gel-permeation filteration. Optimal temperature and pH for the enzyme activity were $40^{\circ}C$ and 7.8, respectively. The enzyme was stable up to $50^{\circ}C$ and at a broad pH range of 5.0-10.0. The $\beta$-agarase was activated by $Zn(NO_3)_2$, and was inhibited by $CuSO_4$ and $SnCl_2$. The Km and Vmax values of this enzyme for agarose as a substrate was $2.4mg/m\ell$ and 13.6 mg/m$\ell$, respectively.

• Microbiology and Biotechnology Letters
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• v.26 no.4
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• pp.297-302
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• 1998

$\beta$-Xylosidase B was produced by Escherichia coli HB101/pKMG12 carrying the xylB gene of Bacillus stearothermophilus No.236 on its recombinant plasmid. The $\beta$-xylosidase B produced was purified by ammonium sulfate fractionation, DEAE-Sepharose CL-6B, Sephacryl S-200 and Superdex 200 HR gel filtration. The purified enzyme showed the highest activity at pH 6.5 and 5$0^{\circ}C$. But, the enzyme was observed to be very sensitive to the pH and temperature of the reaction mixture. The enzyme was activated about 35% of its original activity in the presence of 1 mM of $Mn^{2+}$ but it was completely inhibited by $Ag^{+}$, $Cu^{2+}$and $Hg^{2+}$ions. In contrast with the $\beta$-xylosidase A, the B enzyme was found to have $\alpha$-arabinofuranosidase activity though the activity was fairly low compared with the $\alpha$-arabinofuranosidase produced from the arfI gene of the same Bacillus stearothermophilus. Therefore, $\beta$-xylosidase B is considered to be more suitable than $\beta$-xylosidase A at least for the biodegradation of arabinoxylan. The $K_{m}$ and V$_{max}$ values of the $\beta$-xylosidase B for o-nitrophenyl-$\alpha$-D-xylopyranoside were 6.43 mM and 1.45 $\mu$mole/min, respectively. Molecular mass of the enzyme was determind to be about 54 kDa by SDS-PAGE and 160 kDa by Superdex 200HR gel filtration, indicating that the functional $\beta$-xylosidase B was composed of three identical subunits.s.

• Microbiology and Biotechnology Letters
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• v.14 no.3
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• pp.219-223
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• 1986

In order to preparr the hybridoma cells which produce a monoclonal antibody to human fibroblast interferon(Hu IFN-$\beta$), spleen cells from BALB/cmice immunized with the purified Hu IFN-$\beta$ were fused with NS-O cells, a myeloma cell line. Forty hybrids with high titer among 1300 hybrids Isolated by an ELISA screening method were subcloned using the soft agarose cloning and limiting dilution methods, and 11 hybrids were selected. As a result of iso-typing the hybrids using the mouse monoclonal typing kit, two hybridomas were found to produce 1gG 2a type of monoclonal an-tibodies. The ascites fluid from nude mice inoculated intraperitoneally with the above hybridomas was removed and purified using a protein A-Sepharose CL-4B. Monoclonal antibody was proven to have only the heavy and light chains on SDS-polyacrylamide gel electrophoresis.