• Archives of Pharmacal Research
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• 제13권4호
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• pp.330-337
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• 1990

The crude polysaccharide from Panax ginseng prepared by hot water extration and precipiation with ethanol was further fractionated into neutral and acidic fractions by DEAE- cellulose ion exchange chromatography. The chemical compositions were 85.0% carbohydrorate and 15.0% protein for the neutral fraction, and 28.4% carbohydrate, 10.0% protein and 29.0% uronic acid for the acidic fraction. The acidic fraction was more effective in increasing of the ratio of spleen to body weight, the number of antibody secreting cells to SRBC and phagocytic activity of reticuloendothelial system, as well as antitumor activity against the solid form of sarcoma 180 in ICR mice than the neutral fraction. All polysaccharide fractions were mitogenic to cultured spleen cells of C57BL/6 mice. However, FA was different from FN in the co-mitogenicities with lectin mitogens. Both crude and acidic fractions potentiated remarkably the mitogenic activity of PHA-P or LPS in dose-dependent manner but neutral fraction enhanced only that of LPS. Three polysaccharide fractions had no effect on that of Con A. These results suggest that the acidic fraction may stimulate B and Td cells as well as macrophages while the neutral fraction may simulate only B cells and macropages.

• Mycobiology
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• 제36권1호
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• pp.50-54
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• 2008

This study examined the chemical composition of A. blasiliensis and the chemical structural properties of an immuno-stimulating polysaccharide. The amino acids, free sugars, and organic acids by HPLC and fatty acids by GC were analyzed. The immuno-stimulating substance from A. blasiliensis was extracted with hot water and purified by ethanol precipitation. It underwent ion exchange chromatography on DEAE-cellulose and gel filtration on Toyopearl HW 65F. Through GP-HPLC, the substance was found to be homogeneous. Its chemical structure was determined by $^{13}C-NMR$. Fatty acids, organic acids, and sugar alcohol composition consisted exclusively of linoleic acid, fumaric acid and mannitol, respectively. The amino acids were mainly glutamic acid, glycine, and arginine. By $^{13}C-NMR$ analysis, the immuno-stimulating substance was identified as ${\beta}-(1{\rightarrow}3)\;(1{\rightarrow}6)$-glucan, composed of a backbone with $(1{\rightarrow}3)$-linked D-glucopyranosyl residues branching a $(1{\rightarrow}6)$-linked D-glucopyranosyl residue. The ${\beta}$-glucan from A. blasiliensis showed pronounced immuno-stimulating activity on the antibody-production ability of B-lymphocytes by the hemolytic suspension assay. In these results, A. blasiliensis was estimated to have potent pharmacological properties and potential nutritional values.

• 한국미생물·생명공학회지
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• 제19권3호
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• pp.253-258
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• 1991

Aspergillus ficuum이 생산하는 exoniulinase가 CM-Sephadex, DEAE-Sepharose 6B, 및 HPLC gel filtration을 통해 단백질 mg당 약 2,800U의 specific activity로 정제되었다. 이 효소는 native 상태에서 약 83,000+1,000의 분자량을 나타냈으며 당을 함유하고 있었다. 이 효소는 최적 pH가 4.4-4.7이었고, 55'C에서 8시간 노출 후에도 그 활성을 95 유지하였다. 이 효소의 I/S ratio는 약 0.35이고 전형적인 non-speific Beta-fructofuranosidase의 특성을 나타내었으며 raffinose와 stachyose를 분해할 수 있었다.

• 한국미생물·생명공학회지
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• 제23권4호
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• pp.446-453
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• 1995

$\alpha $-Arabinofuranosidase was produced by E. coli HB101 haboring the recombinant plasmid pKMG11 which contained the arfI gene of Bacillus stearothermophilus. The maximum production of the enzyme was observed when E. coli HB101 cells were grown at 37$\circ$C for 20 hours in the medium containing 0.5% arabinose, 1.0% tryptone, 0.5% yeast extract, and 1% NaCl. The $\ALPHA $-arabinofuranosidase produced was purified to homogeneity using a combination of 20-50% ammonium sulfate precipitation, DEAE-Sepharose CL-6B ion exchange column chromatography and Sepharose 6B-100 gel filtration. The purified enzyme was most active at 55$\circ$C and pH 6.5. The K$_{m}$ and V$_{max}$ values of the enzyme on $\rho $-nitrophenyl-$\alpha $-arabinofuranoside was determined to be 2.99 mM and 0.43 $\mu $mole/min (319.74 $\mu $mole/min/mg), respectively. The pI value was 4.5. The molecular weight of the native protein was estimated to be 289 kDa. The SDS-polyacrylamide gel clectrophoresis analysis suggested that the functional protein was a trimer of the 108 kDa identical subunits. The N-terminal amino acid sequence of the a-arabinofuranosidase was identified as X-Ser-Thr-Ala-Pro-Arg( \ulcorner )-Ala-Thr-Met-Val-Ile-Asp-X-Ala-Phe.

• 한국미생물·생명공학회지
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• 제24권4호
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• pp.472-477
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• 1996

Natural human epidermal growth factor (nhEGF) was purified from pregnant human urine by benzoic acid adsorption, DEAE-Sepharose ion exchange, and immunoaffinity chromatography. The purified nhEGF was further separated into four fractions using Bondapak C$_{18}$ HPLC system. Following characterization by Western blot analysis and double immu- nodiffusion, we found that each fraction corresponds to four derivatives of the nhEGF. For biological analysis of nhEGF, we optimized the labeling time and serum concentration for the incorporatioin of 5-bromo-2'-deoxy uridine (BrdU), a non-radioactive alternative for [$^{3}$H]-thymidine uptake, into NIH 3T3 cells. The DNA synthesis of NIH 3T3 cells was gradually increased at the nhEGF concentrations between 0.1 - 10 ng/ml in the Dulbecco's Modified Eagles Medium (DMEM) containing 0.2% Fetal calf serum (FCS). When we assayed the biological activity of four fractions, the activity of the second fraction was superior to that of the others. Based on the results from the HPLC analysis spiked with recombinant human epidermal growth factor (rhEGF) and amino acid sequencing, we concluded that the second fraction was nhEGF and the other three fractions were the derivatives of nhEGF. In addition, the proportion of nhEGF was approximately 46% is compared with that of the other three derivatives.

• 한국수산과학회지
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• 제40권4호
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• pp.193-200
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• 2007

Ion-exchange chromatography and ultra-filtration permeation were used to extract carnosine from the eel Anguilla japonica. In an investigation of its antioxidant properties, the eel carnosine prevented lipid peroxidation in linoleic acid systems, scavenged free radicals, and exhibited superoxide dismutase-like activity. These activities increased as the carnosine concentration increased. The nitrite scavenging effects (NSEs) of commercial carnosine and the eel carnosine were measured at various acidic pHs (1.2, 3.0, and 4.2). For both types of carnosine, the maximum NSE was observed at pH 1.2. At this pH, the NSE of the eel carnosine was 65.3%. Both types of carnosine were effective at maintaining reasonably good color of ground beef patties over 5 days of storage at $4^{\circ}C$ and inhibited metmyoglobin formation as well as lipid peroxidation. These data suggest that the eel carnosine might be useful as a "natural" antioxidant in commercial production and storage of muscle foodstuffs.

• 한국수산과학회지
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• 제44권6호
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• pp.630-634
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• 2011

Enzymatic hydrolysate of porphyran from Porphyra yezoensis was prepared by treatment with ${\beta}$-agarase. The hydrolysate was fractioned into molecular sizes of <3, 3-30, and 30-300 kDa using an ultrafiltration membrane. The membrane fractions were further separated into neutral and anionic fractions using Dowex $1{\times}8$ ion exchange chromatography. After hydrolysis of porphyran with ${\beta}$-agarase, 23.2% of the starting porphyran was recovered as a neutral fraction of low-molecular weight (<3 kDa), and 28.9% remained as an enzyme-resistant anionic fraction of high molecular weight (>300 kDa). Desulfation of porphyran and $^{13}C$-NMR analysis of the anionic fraction of low molecular weight (<3 kDa) showed that the anionic fraction has a backbone consisting of 3-linked ${\beta}$-D-galactose units alternating with either 4-linked a-L-galactose 6-sulfate or 3, 6-anhydro-a-L-galactose units. These results indicate that porphryan is a copolymer of two moieties, about 25% of which are composed of neoagarose moieties and 75% as anionic moieties.

• 한국식품영양과학회지
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• 제27권1호
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• pp.38-45
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• 1998

To extract insoluble proteins and to improve funtional properties of abolished proteins, an phytase producing Aspergillus sp. SM-15 was isolated from soil. The enzyme was purified and its enzymological characteristics were investigated. Phytase production reached to maximum when the wheat bran medium containing 1% mannose, 1% yeast extract, 1% (NH4)2HPO4 and 0.2% calcium chloride was cultured for 4 days. Phytase was purified 17.1 fold and specific activity was 244.32unit/mg by a sequencial process of ammonium sulfate fraction, ion exchange chromatography and gel filtrations Pruified enzyme was confirmed as a single band by the polyacrylamide gel electro-phoresis. The molecular weight of phytase was estimated to be 46,000. The optimum pH and temperature for the phytase activity were 5.5 and 5$0^{\circ}C$. The enzyme is stable in pH 4.5~5.5, 6$0^{\circ}C$. The activity of purified enzyme was inhibited by Hg2+ whereas activited by Pb2+ and Fe2+. The activity of phytase was inhibited by the treatment with iodine. The result indicate the possible involvement of histidine at active site. Km and Vmax of the puridied phytase were 37.037mM/L and 159.87umol/min, respectively.

• 한국식품영양과학회지
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• 제28권1호
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• pp.9-15
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• 1999

Taurine contents in alcoholic and non alcoholic beverages, milk and milk products, sugars and condiments were determined for 83 food items commonly used by Koreans. Taurine concentrations of food samples were analyzed by using an automated amino acid analyzer(Biochrom 20, Pharmacia LKB) based on ion exchange chromatography. Taurine was not detected in most carbonated beverages, but found in a variety of fruit and vegetable beverages(canned) in the range of 0.13 ~1.43mg taurine/ 100g wt, and in instant coffee or tea beverages(canned) in the range of 0.10~0.41mg taurine/100g wt. The traditional Korean turbid rice wine(takju) sample contained the highest level of taurine (2.29mg/100g wt) among alcoholic beverages tested, which was followed by wine(0.59~0.88mg tau rine/100g wt) and beer(0.53~0.73mg taurine/100g wt). Ordinary milk samples contained 1.05~1.40 mg taurine/100g wt, and a variety of taurine supplemented infant formulas manufactured by Korean companies included 17.3~25.4mg taurine/100g wt. Taurine was not detected in most sugars and sweetners except a couple of chocolate samples(2.13~2.18mg taurine/100g wt). Exceptionally high level of taurine was found in a curry powder sample(16.9mg taurine/100g wt), while the rest of commonly used condiments contained less than 2.90mg taurine/100g wt.

• BMB Reports
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• 제38권3호
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• pp.294-299
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• 2005

In this report, we describe the high-yield secretory expression of the recombinant human anti-HBsAg Fab fragment from Pichia pastoris that was achieved by co-integration of the genes encoding the heavy and light chains (both under the control of alcohol oxidase promoter) into the genome of the yeast cells. The fed-batch fermentations were carried out in a 5 L scale. Both chains of the Fab were successfully expressed upon methanol induction. The absorbance ($OD_{600}$) of the broth can reach 350~500 at the end of fed-batch phase. After the induction, the expression level of the recombinant Fab (soluble) reached 420~458 mg/L. The recombinant Fab fragment was purified from the crude culture supernatant by ion exchange chromatography and the purity of the recombinant Fab fragment was over 95%. The affinity activities of the crude fermentation supernatant and the purified Fab were analyzed by indirect ELISA, which showed that the purified recombinant Fab fragment had high affinity activity with hepatitis B surface antigen.