• Journal of Microbiology and Biotechnology
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• 제19권9호
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• pp.987-996
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• 2009

Aspergillus niger GH1 previously isolated and identified by our group as a wild tannase producer was grown under solid-state (SSC) and submerged culture (SmC) conditions to select the enzyme production system. For tannase purification, extracellular tannase was produced under SSC using polyurethane foam as the inert support. Tannase was purified to apparent homogeneity by ultrafiltration, anion-exchange chromatography, and gel filtration that led to a purified enzyme with a specific activity of 238.14 IU/mg protein with a final yield of 0.3% and a purification fold of 46. Three bands were found on the SDS-PAG with molecular masses of 50, 75, and 100 kDa. PI of 3.5 and 7.1% N-glycosylation were noted. Temperature and pH optima were 600e and 6.0 [methyl 3,4,5-trihydroxybenzoate (MTB) as substrate], respectively. Tannase was found with a $K_M$ value of $0.41{\times}10^{-4}M$ and the value of $V_{max}$ was $11.03{\mu}$moL/min at $60^{\circ}C$ for MTB. Effects of several metal salts, solvents, surfactants, and typical enzyme inhibitors on tannase activity were evaluated to establish the novelty of the enzyme. Finally, the tannase from A. niger GH1 was significantly inhibited by PMSF (phenylmethylsulfonyl fluoride), and therefore, it is possible to consider the presence of a serine or cysteine residue in the catalytic site.

• Journal of Microbiology and Biotechnology
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• 제21권9호
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• pp.960-967
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• 2011

Tannin acyl hydrolase, also known as tannase, is an enzyme with important applications in the food, feed, pharmaceutical, and chemical industries. However, despite a growing interest in the catalytic properties of tannase, its practical use is very limited owing to high production costs. Several studies have already demonstrated the advantages of solid-state fermentation (SSF) for the production of fungal tannase, yet the optimal conditions for enzyme production strongly depend on the microbial strain utilized. Therefore, the aim of this study was to improve the tannase production by a locally isolated A. niger strain in an SSF system. The SSF was carried out in packed-bed bioreactors using polyurethane foam as an inert support impregnated with defined culture media. The process parameters influencing the enzyme production were identified using a Plackett-Burman design, where the substrate concentration, initial pH, and incubation temperature were determined as the most significant. These parameters were then further optimized using a Box-Behnken design. The maximum tannase production was obtained with a high tannic acid concentration (50 g/l), relatively low incubation temperature ($30^{\circ}C$), and unique low initial pH (4.0). The statistical strategy aided in increasing the enzyme activity nearly 1.97-fold, from 4,030 to 7,955 U/l. Consequently, these findings can lead to the development of a fermentation system that is able to produce large amounts of tannase in economical, compact, and scalable reactors.

• Journal of Microbiology and Biotechnology
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• 제21권12호
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• pp.1312-1321
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• 2011

Enzyme extracts of cellulase [filter paper cellulase (FPase) and carboxymethyl cellulase (CMCase)], chitinase, and chitosanase produced by Aspergillus niger NRRL-567 were evaluated. The interactive effects of initial moisture and different inducers for FP cellulase and CMCase production were optimized using response surface methodology. Higher enzyme activities [FPase $79.24{\pm}4.22$ IU/gram fermented substrate (gfs) and CMCase $124.04{\pm}7.78$ IU/gfs] were achieved after 48 h fermentation in solid-state medium containing apple pomace supplemented with rice husk [1% (w/w)] under optimized conditions [pH 4.5, moisture 55% (v/w), and inducers veratryl alcohol (2 mM/kg), copper sulfate (1.5 mM/kg), and lactose 2% (w/w)] (p<0.05). Koji fermentation in trays was carried out and higher enzyme activities (FPase $96.67{\pm}4.18$ IU/gfs and CMCase $146.50{\pm}11.92$ IU/gfs) were achieved. The nonspecific chitinase and chitosanase activities of cellulase enzyme extract were analyzed using chitin and chitosan substrates with different physicochemical characteristics, such as degree of deacetylation, molecular weight, and viscosity. Higher chitinase and chitosanase activities of $70.28{\pm}3.34$ IU/gfs and $60.18{\pm}3.82$ to $64.20{\pm}4.12$ IU/gfs, respectively, were achieved. Moreover, the enzyme was stable and retained 92-94% activity even after one month. Cellulase enzyme extract obtained from A. niger with chitinolytic and chitosanolytic activities could be potentially used for making low-molecular-weight chitin and chitosan oligomers, having promising applications in biomedicine, pharmaceuticals, food, and agricultural industries, and in biocontrol formulations.

• Korean Chemical Engineering Research
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• 제52권3호
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• pp.302-306
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• 2014

목질계 분해효소 활성 증대를 위해 밀짚을 이용한 고체발효에서 주요 발효인자의 최적화를 수행하였다. Trichoderma reesei와 Aspergillus niger를 이용한 혼합배양에서 고체발효에 주요한 영향을 미친다고 알려진 배양온도, pH, 수분함량과 고체기질 크기를 순차적 최적화를 진행하였다. 실험에 적용 된 인자 모두 목질계 분해효소 활성에 유의한 효과를 주었으며, 발효온도 $40^{\circ}C$, pH 7, 수분함량 75%와 고체기질 크기 0.25~0.5 mm가 목질계 분해효소 생산을 위한 최적 조건임을 알 수 있었다. 최적조건 하에서 밀짚을 이용한 고체발효를 수행하였을 때, 효소활성 기준 cellulase 10.3 IU, endoglucanase 100.3 IU, ${\beta}$-glucosidase 22.9 IU와 xylanase 2261.7 IU/g dry material을 배양 96시간에 확인할 수 있었다. 본 결과는 기존 효소활성 대비 각각 72.6, 48.7, 55.2와 51.9% 증가한 수치로 혼합배양과 순차적 최적화를 적용하여 효과적인 목질계 분해효소 활성 증대가 가능함을 확인하였다.

• 한국연초학회지
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• 제25권1호
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• pp.70-79
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• 2003

$\beta$ -Glucosidase-catalysed synthesis of glucosides with aromatic alcohols and monoterpene alcohols as accepters and cellobiose as a donor in the presence of various commercial $\beta$ -glucosidases were described. $\beta$ -Glucosidases from Aspergillus niger spp,. Trichoderma spp., Penicillium sup. and bitter almond have been shown to catalyze synthesis of $\beta$ -glucosides of benzyl alcohol, 2-hydroxybenzyl alcohol, 4-hydroxybenzyl alcohol, 2-phenylethyl alcohol, geraniol and citronellol in the presence of cellobiose as sugar donor. Among enzyme preparations tested, each $\beta$ -glucosides prepared from Aspergillus niger were isolated in the pure state by Diaion HP-20 and silica gel column chromatography. The products were identified as $\beta$ -glucosyl products of benzyl alcohol, 2-hydroxyhenzyl alcohol, 4-hydroxybenzyl alcohol, 2-phenyl ethyl alcohol, geraniol and citronellol by spectrometry (UV, IR, $^1$H-NMR, $^{13}$ C-NMR) and enzymatic hydrolysis with $\beta$ - glucosidase. Monoterpene alcohols with a sterically hindered hydroxyl group, such as linalool, $\ell$-menthol and $\alpha$-terpineol were not used as acceptors in transglycosylation reaction.

• 한국응용과학기술학회지
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• 제26권1호
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• pp.51-59
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• 2009

Natural Artemisia extraction was extracted from Artemisia component using diethyl ether as a solvent, and we tested various pharmacetical and chemical characteristics of this extract. Characteristic experiments to use natural Artemisia extract tested antimicrobial experiment using microbe in pharmacetical material, and tested dye experiment using fiber in chemical material. From the result of characteristics experiment, some conclusions are obtained as follow. From the result of extraction experiment, it obtained about 10.4%-Artemisia extraction ratio as semi-solid state, and after dried in freezing from Artemisia extract of semi-solid state, it obtained about 10%-Artemisia extraction ratio as solid state of dark blue-green color. From result of antimicrobial experiment of Artemisia extract, number of staphylococcus aureus (ATCC-01) and aspergillus niger (ATCC-02) in microbe decreased more and more according to time passage. This phenomenon showed that Artemisia extract influences to antimicrobial effect. From the result of dye experiment of Artemisia extract, it appeared in direction of dark blue-green color after dyed to use cotton and silk with fiber to control in pH 7.5. Specially the result which confirmed dye of fiber with optical electron microscope (OEM), we could know that it appears darker silk than cotton.

• 한국응용과학기술학회지
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• 제26권3호
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• pp.341-349
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• 2009

Natural grape extraction was extracted from grape component using diethyl ether as a solvent, and we tested various pharmaceutical and chemical characteristics of this extract. Characteristic experiments to use natural grape extract tested antimicrobial experiment using microbe in pharmaceutical material, and tested dye experiment using fiber in chemical material. From the result of characteristics experiment, some conclusions are obtained as follow. From the result of characteristics experiment, it obtained about 7.5%-grape extraction ratio as semi-solid state, and after dried in freezing from grape extract of semi-solid state, it obtained about 10%-grape extraction ratio as solid state of dark purple color. From result of antimicrobial experiment of grape extract, number of staphylococcus aureus (KCMC-01) and aspergillus niger (KCMC-02) in microbe decreased more and more according to time passage. This phenomenon showed that grape extract influences to antimicrobial effect. From the result of dye experiment of grape extract, it appeared in direction of dark purple color after dyed to use cotton and silk with fiber to control in pH 7.5. Specially the result which confirmed dye of fiber with optical electron microscope(OEM), we could know that it appears darker silk than cotton.

• 보존과학연구
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• 통권22호
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• pp.5-25
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• 2001

The germicidal and insecticidal properties of volatile components extracted from star anise(Illicium verum Hooker filius) and clove (Eugenia caryophyllata THUNBERG)were evaluated against five microorganisms and three insects for the purpose of developing biocidal active substances from medicinal plants. The volatile components of star anise and clove showed strong antimicrobial effect against Aspergillus niger, Penicillium funiculosum, Mucor hiemalis, Trichoderma viride, and Aureobasidium pullulans. The extracts of each medicine also showed insecticidal effects against Sitophilusoryzae L., Lyctus linearis GOZE, and Reticulitermes spertus kyushuensis Morimoto. Fumigant toxicities to adult insects were determined. In the case of fumiganttoxicity, the extract of star anise showed 100% mortality against R. spertus, S.oryzae, and L. linearis at rates of $2.5\mu\ell$, $50\mu\ell$, $250\mu\ell$/filter paper, respectively but showed no killing effects by clove. The volatile components of star anise and clove were investigated by means of GC/MS. The main constitute, anethole among 20components from star anise and eugenol among 9 components from clove were identified. The mixture of star anise and clove as the volume ratio of 2 : 1 showed higher properties for antimicrobial and insecticidal effect than each volatile component. A. niger was inhibited by the mixture(125ml/$m^3$) for up to 10 days of exposure. Also, from the result of observing state change of organic materials by volatile extracts of star anise and clove, volatile extracts effects have no effect on natural organic materials of organic cultural properties and can be used as biological control agent. As research contents as above, the insecticidal and germicidal agents from star anise and clove and the mixture of them were more efficient and high level to prevent biological damage for conservation of organic cultural properties. So they may be used in new development of biologicalinsecticidal and germicidal agents for conservation of cultural properties.

• Journal of Microbiology and Biotechnology
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• 제25권1호
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• pp.33-43
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• 2015

GAM-1 and GAM-2, two themostable glucoamylases from Aspergillus niger B-30, possess different molecular masses, glycosylation, and thermal stability. In the present study, the effects of additives on the thermal inactivation of GAM-1 and GAM-2 were investigated. The half-lives of GAM-1 and GAM-2 at 70℃ were 45 and 216 min, respectively. Data obtained from fluorescence spectroscopy, circular dichroism spectroscopy, UV absorption spectroscopy, and dynamic light scattering demonstrated that during the thermal inactivation progress, combined with the loss of the helical structure and a majority of the tertiary structure, tryptophan residues were partially exposed and further led to glucoamylases aggregating. The thermal stability of GAM-1 and GAM-2 was largely improved in the presence of sorbitol and trehalose. Results from spectroscopy and Native-PAGE confirmed that sorbitol and trehalose maintained the native state of glucoamylases and prevented their thermal aggregation. The loss of hydrophobic bonding and helical structure was responsible for the decrease of glucoamylase activity. Additionally, sorbitol and trehalose significantly increased the substrate affinity and catalytic efficiency of the two glucoamylases. Our results display an insight into the thermal inactivation of glucoamylases and provide an important base for industrial applications of the thermally stable glucoamylases.

• 대한화학회지
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• 제65권2호
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• pp.93-105
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• 2021

Mononuclear Cu(II), Ni(II), Co(II), Mn(II), Zn(II), Fe(III), Ru(III), and UO2(II) complexes of 2-hydroxy-4-(p-tolyldiazenyl)benzylidene)-2-(p-tolylamino)acetohydrazide (H2L) were prepared by direct method. The ligand and its complexes were isolated in solid state and characterized by analytical techniques such as elemental and thermal analyses, molar conductance, magnetic susceptibility measurements and spectroscopic techniques such as UV-Visible, IR, 1H-NMR and 13C-NMR. The spectral data indicated that the ligand acted as neutral/monobasic bidentate or monobasic/dibasic tridentate ligand bonded to the metal ions through the oxygen atom of ketonic or enolic carbonyl group, azomethine nitrogen atom and deprotonated/protonated phenolic oxygen atom forming either tetragonally distorted octahedral or octahedral. Antimicrobial activities of the ligand and its complexes were evaluated against Escherichia coli, Bacillus subtilis and Aspergillus niger by well diffusion method. The results of antifungal activity showed that the Fe(III) complex (10) exhibited higher antifungal against Aspergillus niger than the other complexes. However, the results of antibacterial activity revealed that Cu(II) complex (4) is the most active against Escherichia coli while the Cu(II) complex (5) and Fe(III) complex (10) exhibited higher antibacterial effect on Bacillus subtilis than the other complexes.