• 한국식품저장유통학회지
• /
• 제7권2호
• /
• pp.201-206
• /
• 2000

${\beta}$-Galactosidase was extracted and purified from strawberry. The purified ${\beta}$-Galactosidase from strawberry was investigated their physicochemical characteristics. ${\beta}$-Galactosidase was purified 25.74 fold from strawberry. The purification procedure include ammonium sulfate fraction, acetone powder treatment and gel and ion exchange chromatography. Yield of the enzyme purification was 18.11%. The purified enzyme has native molecular weight of 116,000 dalton. Vmax value and Km value of ${\beta}$-Galactosidase were 0.077 mM ONPG/ml/15mim and 1.75x10-2mM, respectively. The optimum temperature and pH of ${\beta}$-Galactosidase were 43$^{\circ}$C and pH 4.0, respectively. The ${\beta}$-Galactosidase activity was stable below 50$^{\circ}$C and at pH 4.0 to pH 6.0. Among the metal ions Ca and Mg were did not affect, whereas K, Cu and Zn show a little effect on the enzyme activity. The ${\beta}$-Galactosidase activities were inhibited by treatment with EDTA and SDS.

• Food Science and Biotechnology
• /
• 제15권6호
• /
• pp.908-913
• /
• 2006

Recombinant ${\beta}$-galactosidase from Bifidobacterium longum LL04 was expressed in Escherichia coli and partially purified by ammonium sulphate precipitation and anion-exchange chromatography (Mono-Q). The optimum temperature and pH of the partially purified enzyme were $50^{\circ}C$ and pH 7.0-8.0, respectively, when o-nitrophenyl-${\beta}$-D-galactopyranoside was used as a substrate. The enzyme was stable over the pH range of 5.0-9.0, and was active at $40^{\circ}C$ for more than 60 min at pH 7.0. The enzyme was significantly activated by $Na^+$ and $K^+$. Maximal activity was observed at the concentration of 10 mM for both $Na^+$ and $K^+$. The enzyme activity was strongly inhibited by most bivalent metal ions. The Km and Vmax on ONPG at 37 and $50^{\circ}C$ were 0.72, 167.9, and 0.507 mM, 310.9 U/mL, respectively.

• 한국식품위생안전성학회지
• /
• 제2권2호
• /
• pp.67-73
• /
• 1987

Kluyveromyces fragilis로부터 $\beta$-galactosidase의 생성조건과 조효소액의 성질 그리고 당 전이 반응에 의한 oligosaccharide 생성을 조사한 결과 다음과 같은 결과를 얻었다. \circled1 Peptone-Yeast extract 배지에 6% lactose를 첨가하였을 때 최대 효소생산을 보였다. \circled2 0.05 M potassium phosphate buffer (pH 7.0)에서 3% toluene를 첨가하여 37$^{\circ}C$ 5시간 배양하였을 때 K. fragilis로부터 효소가 최대로 추출되었다. \circled3 효소의 최적온도는 4$0^{\circ}C$이고 4$0^{\circ}C$ 이상의 열처리에서는 효소가 파괴되었으며, pH6-7에서는 상당히 안정하였다. \circled4 ONPG 기질로 사용하였을 때 Km 값은 2.5mM이었다. \circled5 당 전이 반응의 결과, 7개의 oligosaccharide가 생성되었다. 이상의 실험결과로 볼 때, 본 실험에 사용한 K. fragilis SKD 7001은 Beta-galactosidase의 생산을 위해서 이용 가치가 인정되었으며, 특히, 이 효소의 활성이 중성 pH에서 강하고 안정한 상태를 보이는 것은 시유나 원료우유의 lactose를 중성에서 해야 한다는 점을 고려할 때, 실용적 가치가 있다고 본다. 또, 이 효소의 작용과정에서 생성되는 oligosacchride는 장내 bifidus균의 생육을 촉진시키는 효과가 인정되고 있기 때문에 이용가치를 더욱 높여주는 것으로 생각된다.

• 대한미생물학회지
• /
• 제22권4호
• /
• pp.427-433
• /
• 1987

One hundred and thirty-six strains of Klebsiella pneumoniae were isolated from clinical specimen taken from pediatric patiants at 6 different hospitals and identified, characterized and investigated on the patterns of antibiotic resistance. The 136 strains showed the positive reactions in the 17 tests; Voges-Proskauer, ONPG, cirate utilization, KCN degradation, productions of lysine decarboxylase, acid and gas from glucose, utilizations of malonate, manitol, rhamnose, salicin, sucrose, raffinose, arabinose, lactose, sucrose, inositol and raffinose, but the strains showed the negative reactions in the 8 tests; production of $H_2S$, indole, arginine dehydrolase, ornithine decaraoxylase, phenylalanine deaminase, motility, methly red and gelatin liquefaction. 41 did not utilize dulcitol, and 32 did not utilize adonitol. The 36 out of them produced klebecin. The 136 strains were resistant to ampicillin, 2 to gentamicin, 14 to cephalothin, 16 to chloramphenicol, 8 to kanamycin, 13 to streptomycin, and 17 to tetracycline. Twenty strains were resistant to 2 kinds of antibiotics 5 strains to 3, 4 to 4 and 1 to 6 and 7.

• Food Science and Biotechnology
• /
• 제18권6호
• /
• pp.1342-1350
• /
• 2009

This paper investigates the production and optimization of $\beta$-galactosidase enzyme using synthetic medium by Kluyveromyces lactis NRRL Y-8279 in stirred tank bioreactor. Response surface methodology was used to investigate the effects of fermentation parameters on $\beta$-galactosidase enzyme production. Maximum specific enzyme activity of 4,622.7 U/g was obtained at the optimum levels of process variables (aeration rate 2.21 vvm, agitation speed 173.4 rpm, initial sugar concentration 33.8 g/L, incubation time 24.0 hr). The optimum temperature and pH of the $\beta$-galactosidase enzyme produced under optimized conditions were $37^{\circ}C$ and pH 7.0, respectively. The enzyme was stable over a pH range of 6.0-7.5 and a temperature range of $25-37^{\circ}C$. The $K_m$ and $V_{max}$ values for O-nitrophenol-$\beta$-D-galactopyranoside (ONPG) were 1.20 mM and $1,000\;{\mu}mol/min{\cdot}mg$ protein, respectively. The response surface methodology was found to be useful in optimizing and determining the interactions among process variables in $\beta$-galactosidase enzyme production. Hence, this study fulfills the lack of using mathematical and statistical techniques in optimizing the $\beta$-galactosidase enzyme production in stirred tank bioreactor.

• 한국식품저장유통학회지
• /
• 제7권1호
• /
• pp.1-7
• /
• 2000

To develop natural antimicrobial agents for keeping qualities of postharvested greenhouse produce the antimiocrobial actions of Polygonum cuspidatum Sieb. et Zucc. extract , which showed remarkable antimicrobial effects against microorganism causing the postharvest decay of greenhouse produce, were investigate. In the inhibitory experiment of enzymes related to energy production metabolism hexokinase activities decreased to 73% and 68% by treating with Polygonum cuspidatum Sieb. et.Zucc. extract and Eugenia caryophyllata Thumnberg extract in comparison with control, respectively. Direct visualization of microbial cells by using both transmission electron microscope and scanning electron microscope showed that microbial cell membrane was destroyed by treating with the dilute extract solution. this change of celluloar membrane permeability could be identified in the experiment that 0-nitrophenyl-${\beta}$-D-galactopyrano-side(ONPG), the artificial substrate of ${\beta}$-galactosidase, was hydrolyzed in the presence of the extract, indicating that the membrane was perturbed. The separation and identification of the most antimicrobialo substances isolated from Polygonum cuspidatum Sieb et. Zucc. extract and Eugenia caryophyllata Thunberg extract were carried out by using gas chromatography and mass spectrometry 9GC/MSD), which were identified as eugenol. As a result, the functionality of Polygonum cuspidatum Sieb. et Zucc. extract and Eugenia caryophyllata Thunberg extract as antimicrobial agents for keeping qualities of postharvested greenhouse produce may be recommended.

• 한국미생물·생명공학회지
• /
• 제13권3호
• /
• pp.185-189
• /
• 1985

L. sporogenes의 배양여액으로부터 균체외 $\beta$-galactosidase를 ammonium sulfate fractionation, Sephadex G-200 gel filtration, DEAE-Sephadex A-50 ion exchange chromatography와 hydroxyapatite adsorption chromatography등의 4단계 정제공정을 거쳐 순수하게 정제하였다. 정제효소는 347배 정제되어 비활성이 1,585 units/mg 이었으며 수율은 39.5%였다. Sephadex G-200 gel filtration에 의한 native enzyme의 분자량은 140,000이고 SDS-PAGE 에 의해서는 분자량이 72,000 한가지로 나타났으므로 L. sporogenes의 $\beta$-galactosidase는 동일한 subunit 2개로 구성된 dimer 효소이다.

• 한국축산식품학회지
• /
• 제33권5호
• /
• pp.565-571
• /
• 2013

An enzyme ${\beta}$-galactosidase or ${\beta}$-galactohydrolase [EC3.2.1.23], commonly called lactase, mediates galacto-oligosaccharide (GOS) synthesis under conditions of high substrate concentrations. Also, lactase hydrolyzes ${\beta}$($1{\rightarrow}4$) lactose into glucose and galactose, the latter is successively transferred to free lactose to make various oligosaccharides via transgalactosylation. GOS is non-digestible to human digestive enzymes and has been used as a functional prebiotics. Among the 24 lactic acid bacteria (LAB) strains used, Lactobacillus paracasei YSM0308 was selected based on its exhibition of the highest ${\beta}$-galactoside hydrolysis activity, and the crude lactase was prepared for examination of reaction conditions to affect the GOS synthesis. Lactase activity was measured with a spectrophotometer using ONPG (o-nitropheyl ${\beta}$-D-galactopyranoside) method. Lactase activity was not detected in the culture supernatant and was mostly present in the cell pellet after centrifugation. Activity of the crude lactase preparation ranges from102 to 1,053 units/mL, with the highest activity determined for L. paracasei YSM0308. Optimal conditions for GOS synthesis are as follows: concentration of whey powder, pH, temperature, and time were 30%, pH 6.5-7.0, $30^{\circ}C$, and 4 h, respectively. The final GOS concentration was 19.41% (w/v) by the crude YSM0308 lactase, which was obtained from strain YSM0308 grown in the 10% (w/v) reconstituted whey-based medium.

• 한국미생물·생명공학회지
• /
• 제17권1호
• /
• pp.35-45
• /
• 1989

Streptococcus thermophilus 510으로부터 $\beta$-galactosidase의 생성조건은 탄소원으로 0.5% lactose를 첨가한 배지에서 초기 pH7.0, 배양온도 37$^{\circ}C$, 배양기간 18시간이었다. 배양여액으로부터 $\beta$-galactosidase를 ammonium sulfate 분획, 핵산의 제거, Sephadex G-200 gel filtration 및 DEAE-Sephadex A-50 ion exchange chromatography 등의 4단계 정제과정을 거쳐 정제한 결과 18배 정제되어 단일 단백질로 분리되었다. 정제효소의 활성 최적온도는 5$0^{\circ}C$, 최적 pH는 7.0이었고, 효소활성이 Mn$^{2+}$, $K^+$과 같은 금속이온과 dithiothreitol, 2-mercaptoethanol에 의해 촉진되었고, Hg$^{2+}$, $Zn^{2+}$, Co$^{2+}$, $Ca^{2+}$, EDTA, 8-hydroxyquinoline, galactose 등에 의해 저해되었다. 효소의 분자량이 520,000, 합성기질인 ONPG에 대한 $K_{m}$ 은 1,25mM, V$_{max}$는 88.50 $\mu$mole/min.mg protein이었고, 주종 아미노산은 glutainic acid, aspartic acid, leucine 및 valine이었다.

• 한국물환경학회지
• /
• 제25권3호
• /
• pp.363-369
• /
• 2009

We developed the Eco medium for Escherichia coli and total coliforms, which was modified by Violet Red Bile (VRB) medium, and derived the standard curve of exponential phase at $OD_{410}$ by using type strains such as E. coli ATCC11303, Enterobacter cloacae KCTC2361, Klebsiella pneumoniae KCTC2241, and Citrobacter freundii KCTC2359. Also, we used total 93 samples of spring and stream water to compare the detection ability of total coliforms between the method using Eco medium and such as most probable number (MPN), and plate count methods. As a result, the qualitative analysis of E. coli and total coliforms using Eco medium contained ortho-nitrophenyl-$\beta$-galactoside (ONPG) and 4-methylumbelliferyl-$\beta$-D-glucuronide (MUG) was same as those of Korean standard methods (Colilert kit). And the colony forming unit (CFU) detected in Eco medium was similar to those of result from MPN and plate count methods. Moreover, the agreement, sensitivity, and specificity of the developed kit was more than 97.5% in comparison with Colilert kit for 350 samples. Thus, the Eco medium can be used both qualitative and quantitative analysis of E. coli and total coliforms.