• Mycobiology
• /
• v.48 no.4
• /
• pp.326-329
• /
• 2020

Valuable natural compounds produced by a variety of microorganisms can be used as lead molecules for development of new agrochemicals. Furthermore, high-throughput in vitro screening systems with specific modes of action can increase the probability of discovery of new fungicides. In the current study, a rapid assay tested with various microbes was developed to determine the degree of respiratory inhibition of Saccharomyces cerevisiae in two different liquid media, YG (containing a fermentable carbon source) and NFYG (containing a non-fermentable carbon source). Based on this system, we screened 100 fungal isolates that were classified into basidiomycetes, to find microbial secondary metabolites that act as respiratory inhibitors. Consequently, of the 100 fungal species tested, the culture broth of an IUM04881 isolate inhibited growth of S. cerevisiae in NFYG medium, but not in YG medium. The result is comparable to that from treatment with kresoxim-methyl used as a control, suggesting that the culture broth of IUM04881 isolate might contain active compounds showing the inhibition activity for respiratory chain. Based on the assay developed in this study and spectroscopic analysis, we isolated and identified an antifungal compound (-)-oudemansin A from culture broth of IUM04881 that is identified as Oudemansiella venosolamellata. This is the first report that (-)-oudemansin A is identified from O. venosolamellata in Korea. Taken together, the development of this assay will accelerate efforts to find and identify natural respiratory inhibitors from various microbes.

• Applied Chemistry for Engineering
• /
• v.16 no.3
• /
• pp.456-459
• /
• 2005

UV spectrometric assay for measurement of epoxide hydrolase activity was tested for efficient screening of whole cell activity of epoxide hydrolase. Epoxide hydrolase activities were determined by measuring the amount of p-nitrostyrene diol (pNSD), which was the hydrolysis product of p-nitrostyrene oxide (pNSO). Enantioselective hydrolysis of racemic pNSO using epoxide hydrolase activity of Rhodosporidium toruloides was monitored by UV spectrometric assay, and the relevant $K_m$ and $V_m$ for R. toruloides were determined as $2.457nmol/min{\cdot}mg$ and 1.078 mM, respectively.

• Journal of Microbiology and Biotechnology
• /
• v.12 no.2
• /
• pp.286-291
• /
• 2002

Histone deacetylase I (HDAC1) works as one of the components in a nucleosome remodeling (NuRD) complex that consists of several proteins, including metastasis-associated protein 1 (MTA1). Since the protein-protein interaction of HDAC1 and MTA1 would appear to be important for both the integrity and functionality of the HDAC1 complex, the interruption of the HDAC1 and MTA1 interaction may be an efficient way to regulate the biological function of the HDAC1 complex. Based on this idea, a yeast two-hybrid system was constructed with HDAC1 and MTA1 expressing vectors in the DNA binding and activation domains, respectively. To verify the efficiency of the assay system, 3,500 microbial metabolite libraries were tested using the paper disc method, and KB0699 was found to inhibit the HDAC1 and MTA1 interaction without any toxicity to the wild-type yeast. Furthermore, KB0699 blocked the interaction of HDAC1 and MTA1 in an in vitro GST pull down assay and induced morphological changes in B16/BL6 melanoma cells, indicating the interruption of the HDAC1 complex function. Accordingly, these results demonstrated that the yeast assay strain developed in this study could be a valuable tool for the isolation of a HDAC1 complex disruptor.

• Journal of the Korean Institute of Landscape Architecture
• /
• v.36 no.3
• /
• pp.21-28
• /
• 2008

The management of soil organic matter(SOM) is a key component of golf course green maintenance. As part of a major project examining the sustainable management of SOM on golf course greens, the SOM status of different age greens maintained in the same root zone composition and management were compared. Then the microbial activity, tiller number, bulk density, water content, pH, EC, and T-N in the soil were measured. In the 0${\sim}$5cm depth SOM accumulation showed no significant difference between greens. Below 5cm SOM showed a strong significance between greens and had a positive(+) correlation with year and negative(-) correlation with depth. when regression equations were used to predict SOM accumulation with year and depth, SOM below 5cm tended to increase with a rate of 0.061% . year$^{-1}$ and decrease with a rate of 0.079% . $cm^{-1}$(R2==0.841). Soil microbial activity was investigated with age and depth by using a dehydrogenase assay. Results showed a sharp drop with depth in all greens. The soil microbial activity below 5cm showed no difference between greens. The accumulated SOM below 5cm may be very resistant to decomposition in the long-term. Five years after establishment, the bulk density did not significantly change. The water content, EC, and T-N had a significant correlation with SOM. The pH decreased with the year, which may influence SOM accumulation. Organic matter accumulation was mainly affected by the pH decrase，low soil microbial activity, and high organic matter resistant to decomposition, but the effects of water content, EC, and T-N were obscure.

• Microbiology and Biotechnology Letters
• /
• v.34 no.1
• /
• pp.58-62
• /
• 2006

Rapid assay of enzyme is a primary requirement for successful application of directed evolution technology. Halo generation on a turbid plate would be a method of choice for high throughput screening of enzymes in this context. Here we report a new approach to prepare turbid plates, by controlling the crystallization of tyrosine to form needle-like particles. In the presence of tyrosine phenol-lyase (TPL), the needle-like tyrosine crystals were converted to soluble phenol rapidly than the usual rectangular tyrosine crystals. When an error-prone PCR library of Citrobacter freundii TPL was spread on the turbid plate, approximately 10% of the colonies displayed recognizable halos after 24 hours of incubation at $37^{\circ}C$. Representative positives from the turbid plates were transferred to LB-medium in 96-wellplates, cultivated overnight, and assayed for the enzyme activity with L-tyrosine as the substrate. The assay results were approximated to be proportional to the halo size on turbid plates, suggesting the screening system is directly applicable to the directed evolution of TPL. Actually, two best mutants on the turbid plates were identified to be $2{\sim}2.5$ and 1.5-fold improved in the activity.

• Journal of Korean Society of Forest Science
• /
• v.100 no.2
• /
• pp.136-141
• /
• 2011

This study was conducted to confirm the application as ingredients of cosmetics through an examination of the function for anti-cancer and anti-microbial of the fraction isolated from Pyrus ussuriensis leaves. The dried leaf of P. ussuriensis were extracted with acetone-$H_{2}O$ (6:4, v/v), concentrated and fractionated with the upper layer of acetone on a separatory funnel. Each fraction was freeze dried, then a portion of acetone soluble powder was chromatographed on a Sephadex LH-20 column using a series of aqueous methanol as eluents and also used the MIC-gel using a series of aqueous methanol as developing solvent. The isolated compounds were identified by silica-gel TLC. The growth inhibition activity was measured using the MTT assay by the mouse meltioma (B16F10) cell. The cancer cell growth inhibition rate of fractions isolated from P. ussuriensis leaf was 80%. In anti-microbial activity test, the fraction of P. ussuriensis with 0.25 mg/disc resulted in the clear zone of 1.3 cm and 2 cm for Staphylococcus aureus and S. epidermidis of gram positive bacillus, respectively. In Escherichia coli of gram negative bacillus, the fraction with 0.5 mg/disc resulted in the clear zone of 1.1 cm~1.5 cm each fraction. From these results, we confirmed that acetate fraction of P. ussuriensis has a great potential as a natural ingredients with a anti-cancer and anti-microbial source.

• Journal of Convergence for Information Technology
• /
• v.10 no.12
• /
• pp.183-190
• /
• 2020

This study evaluated the possibility of Hoangtonogak Plus extracts as a bioactive ingredients for cosmetic products. Methanol(MN) and hot-water(WN) extracts were analysed by DPPH/ABTS radical scavenging activity, FRAP value for anti-oxidant activity, MTT assay for cell viability, inhibition of NO production and iNOS protein expression for anti-inflammatory effect, paper disc diffusion method for anti-microbial activity against Staphylococcus aureus, Staphylococcus epidermidis and Escherichia coli.. The contents of total polyphenol of MN and WN extracts were 2.92±0.01 mgGAE/g and 1.67±0.02 mgGAE/g, respectively. DPPH, ABTS and FRAP values of MN extracts were higher than WN at each concentration. No significant cytotoxicity was observed in RAW264.7 cells. Furthermore, NO production of MN and WN at 1 mg/mL concentration was measured as 11.69 μM, 20.4 μM, respectively. In addition, MN extracts showed anti-microbial effect only on S. epidermidis. Also MN extracts suppressed iNOS protein level in a concentration-dependent manner. According to our results, the MN extracts demonstrated its potential as a natural source of antioxidant with anti-microbial and anti-inflammatory properties.

• Korean Journal of Environmental Agriculture
• /
• v.27 no.2
• /
• pp.139-144
• /
• 2008

A field experiment was conducted to investigate the influence of cultivars and compost on soil microbial activities and diversities in a red pepper-grown field. Compost was applied with 0, 30, and 60M/T $ha^{-1}$ in April and then red pepper seedlings of "Yong-go 4" and "Koeun" were transplanted in May 2007. Soil samples were collected in early August 2007. Measurement of microbial activities was based on a dehydrogenase assay and a fluorescein diacetate hydrolysis. Soil microbial community was characterized with Biolog $EcoPlate^{TM}$ and phospholipid fatty acid(PLFA). Red pepper cultivars did not differentiate the selected soil chemical and microbial properties. Soil pH and soil microbial community changed by amending the soil with 30 and 60 M/T $ha^{-1}$ of compost, and the soil organic matter and potassium content, and soil microbial activities increased in soils amended with 60 M/T $ha^{-1}$ of compost. Red pepper cultivar induced a little different soil chemical properties and microbial activity in soils amended with 60 M/T $ha^{-1}$ of compost even though significant differences were not found in those properties. In conclusion the effects of compost on soil chemical and microbial properties were much higher than red pepper cultivars in short-term period but the effects of red pepper cultivars should be investigated in long-term field test.

• Plant Biotechnology Reports
• /
• v.5 no.3
• /
• pp.245-254
• /
• 2011

This study describes an efficient protocol for Agrobacterium tumefaciens-mediated transformation of two subgroups of genotype AAA bananas (Musa acuminata cv. Pei Chiao and Musa acuminata cv. Gros Michel). Instead of using suspension cells, cauliflower-like bud clumps, also known as multiple bud clumps (MBC), were induced from sucker buds on MS medium containing $N^6$-Benzylaminopurine (BA), Thidiazuron (TDZ), and Paclobutrazol (PP333). Bud slices were co-cultivated with A. tumefaciens C58C1 or EHA105 that carry a plasmid containing Arabidopsis root-type ferredoxin gene (Atfd3) and a plant ferredoxin-like protein (pflp) gene, respectively. These two strains showed differences in transformation efficiency. The EHA105 strain was more sensitive in Pei Chiao, 51.3% bud slices were pflp-transformed, and 12.6% slices were Atfd3-transformed. Gros Michel was susceptible to C58C1 and the transformation efficiency is 4.4% for pflp and 13.1% for Atfd3. Additionally, gene integration of the putative pflp was confirmed by Southern blot. Resulting from the pathogen inoculation assay, we found that the pflp transgenic banana exhibited resistance to Fusarium oxysporum f. sp. cubense tropical race 4. This protocol is highly advantageous to banana cultivars that have difficulties in setting up suspension cultures for the purpose of quality improvement through genetic transformation. In addition, this protocol would save at least 6 months in obtaining explants for transformation and reduce labor for weekly subculture in embryogenic cell suspension culture systems.

• Korean Journal of Oriental Medicine
• /
• v.12 no.3 s.18
• /
• pp.79-90
• /
• 2006

This study was investigated to determine the quality monitoring of distributed materials from Glycyrrhizae Radix (26 samples), Atractylodis Rhizoma Alba (24 samples) according to storage period after $1{\sim}3$ year. We have estimated by identification, purity, loss on drying, ash, acid insoluble ash, extract content, essential oil content, assay and microbial contamination. As a result, Glycyrrhizae Radix (26 samples) were satisfied with the standard of K.P. (Korean Pharmacopoeia) and WHO's microbial contamination limit standard. In the Atractylodis Rhizoma Alba (24 samples), 2 samples were not satisfied with the standard of K.P.(Korean Pharmacopoeia) and WHO's microbial contamination limit standard. The results make practical application of the basic data for the quality control of herbal medicine in storage.