• Title/Summary/Keyword: agar diffusion assay

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Development of an Agar Diffusion Method to Measure Elastase Inhibition Activity Using Elastin-Congo Red

  • Jung Kyung-Hwan;Kim Hyun-Joo
    • Journal of Microbiology and Biotechnology
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    • v.16 no.8
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    • pp.1320-1324
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    • 2006
  • The pancreatic and neutrophil elastases are associated with several illnesses including lung and vascular diseases, various cancers, and pancreatitis. The development of a potent and specific inhibitor to the elastases could lead to new therapies. In this study, an agar diffusion method was modified to include a substrate-dye conjugate (Elastin-Congo red) as a substrate of elastase and an indicator of elastase inhibitory activity. The Elastin-Congo red agar plates consisted of 0.1 % Elastin-Congo red and 2.5% agar. The elastase and elastase inhibitors were simultaneously loaded into wells, ultimately resulting in halo formations in which the halo diameter decreased as the concentration of elastase inhibitor increased. The concentration of elastase inhibitor in the samples, therefore, was inversely proportional to the halo diameters. This simplified method provided an excellent correlation with the standard microplate technique, which uses a chromogenic substrate. The concentration of elastase inhibitor obtained from the culture supernatant of a recombinant elastase inhibitor produced by the yeast Pichia pastoris was easily determined. This study has established a simple modified and inexpensive agar diffusion method that is potentially useful for the identification, quantification, and screening of new elastase inhibitors.

Antimicrobial Effects of Propolis against Oral Microorganisms (프로폴리스의 구강구취균에 대한 항균성)

  • Kim, Sang-A;Chung, Hyun-Jung
    • Korean Journal of Food Science and Technology
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    • v.45 no.3
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    • pp.370-375
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    • 2013
  • Propolis is a resinous mixture found in the tree buds, sap flows, and other botanical sources, which is used by honey bees in the construction of their hives. Antimicrobial effects of propolis were evaluated against Streptococcus mutans KCTC 3065, S. sobrinus KCTC 3308, S. sobrinus KCTC 5134, and Porphyromonas gingivalis KCTC 5352 by an agar diffusion assay. Sensitivity of these microorganisms to propolis was evaluated in broth containing different concentrations of propolis at $37^{\circ}C$, followed by observation using transmission electron microscopy (TEM). Propolis inhibited all oral microorganisms tested at the minimum inhibitory concentration (MIC) of $0.14mg/{\mu}L$ in the agar diffusion assay. Treatment with 0.06 and $0.22mg/{\mu}L$ of propolis had a bactericidal effect in a concentration- and treatment time-dependent manner against the tested microorganisms. TEM of propolis-treated S. mutans KCTC 3065 and P. gingivalis KCTC 5352 revealed structural damage of the cell membrane. The activity of propolis was affected by heat and pH treatment. The results indicate that propolis shows antibacterial activity against oral microorganisms and that it has potential for future applications in the food industry.

Antimicrobial Effect of Caesalpinia sappan L. Extract on Foodborne Bacteria (식중독 세균에 대한 소목 추출물의 항균 활성)

  • Kim, Yi-Seul;Shim, Hye-Mee;Kim, Kwang-Yup
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.45 no.7
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    • pp.1026-1034
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    • 2016
  • To develop a natural antimicrobial agent, we investigated the antimicrobial activities of 13 species of edible herbal plant extracts against major Gram-positive foodborne bacteria. Among the 13 screened edible herbal plants, Caesalpinia sappan L. showed the highest antimicrobial activity. In the paper disc agar diffusion assay, Caesalpinia sappan L. extracts had strong antibacterial activities against most Gram-positive bacteria but did not have antibacterial activities against most Gram-negative bacteria. Minimum inhibitory concentrations of the ethanol extract were 0.06 mg/mL against Clostridium difficile and Listeria monocytogenes and 0.03 mg/mL against Staphylococcus aureus. Their inhibitory activities were not reduced by heat treatment or pH adjustment against C. difficile, L. monocytogenes, and S. aureus. Antimicrobial activities were higher in ethanol extract than in distilled water extract. These results support the potential use of Caesalpinia sappan L. ethanol extract as an antimicrobial agent or functional food components against Gram-positive bacteria.

Screening of New Bioactive Materials from Microbial Extracts of Soil Microorganism (I) Antimicrobial Activity from 200 Sampled Using Microdilution Assay

  • Jung, Sang-Oun;Kim, Joon;Chang, Il-Moo;Ryu, Jae-Chun
    • Archives of Pharmacal Research
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    • v.21 no.3
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    • pp.278-285
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    • 1998
  • The microdilution assay recommended by NCCLS (National Committee for Clinical Laboratory Standards) is one of the standardized methods of antibiotic susceptibility test. This method has been widely used clinically to obtain MIC values of antibiotics on pathogenic microorganisms. It is more convenient, rapid and simple to test many samples than other test methods such as agar diffusion assay and broth macrodilution assay. The screening of antimicrobial agents from microbial extracts is too laborious in its process. Therefore, a number of screening methods having more simple procedure have been developed. In our laboratory, we applied microdilution assay for screening the antimicrobial agents. This assay showed dose-response results and was more sensitive than disc diffusion assay in our system. We tested 200 samples of microbial extracts originated from 100 microbial strains and selected several samples as potential candidates. In this report, we show that the microdilution assay is more convenient method in screeing of antibiotic susceptibility than those previously reported.

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Antibacterial Activities of Essential Oil from Zanthoxylum schinifolium Against Food-Borne Pathogens (산초 정유성분의 식중독균에 대한 항균 활성)

  • Jang, Mi-Ran;Seo, Ji-Eun;Lee, Je-Hyuk;Kim, Gun-Hee
    • Korean journal of food and cookery science
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    • v.26 no.2
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    • pp.206-213
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    • 2010
  • In this study, the antibacterial activities of essential oil from Zanthoxylum schinifolium against four Gram-positive bacteria and six Gram-negative bacteria were investigated. The antibacterial activity of the oils was determined using the agar-well diffusion assay, MIC (minimum inhibitory concentration) and MBC (minimum bactericidal concentration). In particular, essential oil from Z. schinifolium showed higher antibacterial activity against Gram-positive bacteria than against Gram-negative bacteria. Essential oil from Z. schinifolium displayed large inhibition zones especially against Bacillus cereus (31 mm). At concentrations between 0 and $20\;{\mu}g/mL$ the oils showed an antibacterial effect against both Gram-negative and Gram-positive bacteria. The minimum inhibitory concentration (MIC) values against nine bacteria ranged from 1.25 to $5\;{\mu}g/mL$. The minimum bactericidal concentration (MBC) values against eight bacterial ranged from 2.5 to $20\;{\mu}g/mL$, except Shigella sonnei. Furthermore, our finding on the antibacterial activities of essential oils from Zanthoxylum schinifolium validated the use of this plant for medical purposes.

Antibacterial Action against Food-Borne Pathogens by the Volatile Flavor of Essential Oil from Chrysanthemum morifolium Flower (국화 꽃 휘발성 향기성분의 식중독균에 대한 항균 작용)

  • Jang, Mi-Ran;Seo, Ji-Eun;Lee, Je-Hyuk;Chung, Mi-Sook;Kim, Gun-Hee
    • The Korean Journal of Food And Nutrition
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    • v.23 no.2
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    • pp.154-161
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    • 2010
  • The aim of this study was to investigate antibacterial activities of essential oil from C. morifolium against four Grampositive bacteria and six Gram-negative bacteria. The antibacterial activity of the oils was determined by agar-well diffusion assay, minimum inhibitory concentration(MIC), and minimum bactericidal concentration(MBC). Essential oil of C. morifolium had a large inhibition zones especially against Salmonella enterica(21 mm) and Bacillus cereus(19 mm). Essential oil of C. morifolium generally showed higher antibacterial activity against Gram-positive bacteria than Gram-negative bacteria. MIC of essential oil from C. morifolium was 5 ${\mu}g/m{\ell}$ against ten food-borne pathogens. MBC values were determined to be from 5 to 20 ${\mu}g/m{\ell}$ against eight bacteria except Salmonella choleraesuis and Listeria monocytogenes. Therefore, the essential oil of C. morifolium and its components have a potent antibacterial activity against food-borne pathogens, and is expected to be used as a novel food preservative.

In Vitro Antioxidant Activity Profiles of ${\beta}$-Glucans Isolated from Yeast Saccharomyces cerevisiae and Mutant Saccharomyces cerevisiae IS2

  • Song, Hee-Sun;Moon, Ki-Young
    • Food Science and Biotechnology
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    • v.15 no.3
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    • pp.437-440
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    • 2006
  • To explore the possible usefulness of ${\beta}$-glucans as natural antioxidants, the antioxidant profiles of ${\beta}$-glucan, extracted from Saccharomyces cerevisiae KCTC 7911, and water soluble and insoluble mutant ${\beta}$-glucan, isolated from yeast mutant S. cerevisiae IS2, were examined by five different in vitro evaluation methods: lipid peroxidation value (POV), nitric oxide (NO), 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, reducing power, and ${\beta}$-carotene diffusion assay. The antioxidant activities of all ${\beta}$-glucans evaluated in POV test were comparable to or better than that of the known antioxidant, vitamin C. Remarkably, the ${\beta}$-glucan and water insoluble mutant ${\beta}$-glucan possessed 2.5-fold more potent activity than vitamin C at a dosage of 2 mg. Although vitamin C showed 100-fold greater activity than all ${\beta}$-glucans in NO and DPPH tests for measuring the radical scavenging capacity, all ${\beta}$-glucans revealed higher radical scavenging activity than the known radical scavenger, N-acetyl-L-cysteine (NAC), in DPPH test. The water insoluble mutant ${\beta}$-glucan had 2.6- and 5-fold greater antioxidative activity than water soluble ${\beta}$-glucan in NO and DPPH tests, respectively, showing that all ${\beta}$-glucans were able to scavenge radicals such as NO or DPPH. While all ${\beta}$-glucans revealed lower antioxidant profiles than vitamin C in both reducing power activity and ${\beta}$-carotene agar diffusion assay, the ${\beta}$-glucan and water insoluble mutant ${\beta}$-glucan did show a marginal reducing power activity as well as a considerable ${\beta}$-carotene agar diffusion activity. These results confirmed the potential usefulness of these ${\beta}$-glucans as natural antioxidants.

Purification of Therapeutic Serums of Snake Anti-Venom with Caprylic Acid

  • Norouznejad, Nilofar;Zolfagharian, Hossein;Babaie, Mahdi;Ghobeh, Maryam
    • Journal of Pharmacopuncture
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    • v.25 no.2
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    • pp.114-120
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    • 2022
  • Objectives: Antivenom serums have been used extensively for over a century and are the only effective treatment option for snake bites and other dangerous animal envenomations. In therapeutic serum centers, a wide range of antivenoms is made from animal serum, mainly equine and sheep, that are immunized with single or multiple venoms. This work aimed to use caprylic acid (CA) to purify therapeutic snake antivenom. Methods: Plasma was obtained from equine immunized with a mixture of venoms. Immunized plasma was obtained by precipitation of different concentrations (2-5%) of CA. This methodology was compared to that based on ammonium sulfate (AS) precipitation. Sediment plasma proteins were purified by ion-exchange chromatography. Protein assay, SDSPAGE, and agar gel diffusion were performed. Results: The total protein precipitation with AS was higher than precipitation with CA, but the best results were obtained when CA was added to the plasma until a final CA concentration of 5% was reached. Chromatography and electrophoresis indicated a stronger band for the 5% CA, and the gel diffusion assay showed antigen-antibody interaction in the purified serum. Conclusion: The use of CA compared to the routine method for purifying hyperimmune serums is a practical and cost-effective method for preparing and producing therapeutic serums. It constitutes a potentially valuable technology for alleviating the critical shortage of antivenom in Iran.

Detection of tobacco mosaic virus from "Kimchi" (김치에서의 활성 TMV 검출)

  • 박은경;김정화;이영근
    • Journal of the Korean Society of Tobacco Science
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    • v.5 no.1
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    • pp.43-47
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    • 1983
  • Tobacco mosaic virus (TMV) was detected from Kimchi by biological and serological assay 5. Kimchi samples three month after cooked were collected, and were inoculated on N. tabacum var. Burley 21 and NC 95. Out of 33 samples, 6 showed typical symptoms induced by TMV, local necrotic lesions on Burley 21 and mosaic on NC 95. All saps from tobacco leaves showed the mosaic symptom reacted positively against TMV antiserum by agar gel double diffusion test. Based on the results, the Kimchi is considered as one of the important inoculum sources in Korea.

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Anti-bacterial effect of fusion formulation of Coptis rhizoma and Pelargonium sidoides on the growth of bronchial diseases bacteria (황련과 Pelargonium sidoides 복합제제의 호흡기 감염 세균에 대한 항균 효과)

  • Lee, Jong Rok;Min, Byung-Gu;Park, Chung A;Kim, Sang Chan;Park, Sook Jahr
    • Herbal Formula Science
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    • v.25 no.4
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    • pp.449-456
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    • 2017
  • Objective : Coptis rhizoma is traditional herb in Korean medicine, and Pelargonium sidoides extract has been used for relief of acute bronchitis in Western medicine. The present study examined the antibacterial effect of fusion formulation of Coptis rhizoma extract and Pelargonium sidoides extract against bronchial diseases bacteria. Methods : Test sample, fusion formulation of Korean and Western medicine, was prepared by mixing Coptis rhizoma extract and Pelargonium sidoides extract at a ratio of 1:2 (w/w). Antimicrobial properties of test sample were determined by agar diffusion assay and minimum inhibitory concentration (MIC) against bronchus diseases bacteria. Results : In agar diffusion assay, the highest amount of test sample (4 mg/disk) exhibited antibacterial activity against all microorganisms tested. Test sample showed the high activity for S. aureus (19.5 mm), C. diptheriae (16.5 mm), A. fumigatus (19.3 mm), F. nucleatum (22.7 mm) and Mycobactrium sp. (17.3 mm), whereas it showed a low activity for K. pneumonia (9.7 mm). The MIC value was determined as $250{\mu}g/m{\ell}$ against C. diptheriae. Test sample showed better growth inhibitory effects against S. aureus and A. fumigatus with the MIC valus of $125{\mu}g/m{\ell}$. Conclusion : These results suggest the possibility of application to chronic respiratory diseases of fusion formulation of Korean and Western medicine, which was prepared with Coptis rhizoma extract and Pelargonium sidoides extract.