• Korean Journal of Food Science and Technology
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• v.43 no.6
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• pp.754-759
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• 2011

Zinc protoporphyrin (ZnPP) is produced endogenously during heme metabolism and treated in cells as a heme oxygenase inhibitor. In the present study, the effects of ZnPP on the color response of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, a commonly-used method for analyzing cell viability, were investigated. ZnPP induced rapid decolorizaion of MTT formazan under light; the degradation rates were 10- and 20- folds faster in the presence of 5 and $10{\mu}M$ ZnPP, respectively. Methylene blue (MB), another type of photosensitizer, also accelerated degradation of formazan under light. Butylated hyroxytoluene did not inhibit ZnPP- or MB-induced formazan degradation. The color degradation of formazan dye was signficantly delayed in the presence of N-acetylcysteine or ${\beta}$-carotene. The present results suggest that certain photosensitizing compounds may affect the color and stability of MTT formazan, which should be carefully considered when conducting the MTT assay.

• Korean Journal of Food Science and Technology
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• v.50 no.1
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• pp.1-7
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• 2018

The MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay is commonly used for analyzing the cell viability. In this study, effects of various solvents, different lights, and zinc protoporphyrin (ZnPP) on the chemical behavior of MTT formazan were investigated. The color response of MTT formazan in NaOH was highly pronounced; the absorbance of MTT formazan in 0.1 N NaOH at 550 nm was >2-fold higher than that in water, dimethyl sulfoxide (DMSO), methanol, and ethanol. MTT formazan in DMSO and NaOH (>0.1 N) was relatively stable under fluorescent and UV light at 365 nm; its rapid degradation was induced under UV light at 254 nm in all solvents. ZnPP degraded MTT formazan under light in a time- and concentration-dependent manner; MTT formazan in 0.1 N NaOH was the most sensitive to ZnPP, followed by DMSO. These results suggest that NaOH and DMSO might be suitable media for MTT formazan for monitoring photosensitizing properties.

• Applied Biological Chemistry
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• v.36 no.5
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• pp.364-369
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• 1993

Triton X-100 enhances to a marked extent the analytical sensitivity of the nitrobluetetrazolium (NBT) reduction method for the assay of superoxide$(O^-_{\.{^.2}})$ production. In the present work, it was attempted to elucidate the physicochemical nature of this Triton X-100 effect, focusing on not only the surfactant-caused stabilization of the water-insoluble formazan colloid but also the kinetic competition between the $NBT-O^-_{\.{^.2}}$ reaction and the autodisproportionation of concommitantly occuring in aqueous media. The measurements of formazan and $H_2O_2$ formed in a number of reaction systems, as prepared by vortex-mixing potassium superoxide dissolved in an aprotic solvent with aqueous solutions of NBT, revealed that Triton X-100 exerts its effect both through preventing formazan colloid from aggregation and thereby increasing the formazan absorbance and through suppressing the autodisproportionation reaction of $O^-_{\.{^.2}}$. It also turned out that the relative shares of the colloid stabilization effect and the kinetic effect in the contribution to the sensitivity amplification of the NBT method are dependent upon the reaction conditions, particulary the molar concentration ratio of NBT to $O^-_{\.{^.2}}$ in the reaction systems.

• Microbiology and Biotechnology Letters
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• v.25 no.3
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• pp.335-337
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• 1997

In this study, we show a convenient MTT assay for detect the susceptibility of yeast-like form of Trichosporon beigelii against antifungal agents. This assay was developed based on mitocondrial respiration by determining reduction of 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) to formazan. Cells of T beigelii are seeded into 96-well microtiter plates, and antifungal agents, amphotericin B, magainin and CA-ME hybrid peptide were added with various concentrations. After 24 hr incubation, MTT was added, then incubations were continued for 4 hr. Formazan formation was quantified photometrically after extraction of the formazan with acid sodium dodesyl sulfate (SDS). From this assay, we could obtained MICs of antifungal agents against T. beigelii. The presented method can easily be used as an effective methods to assess the antiftingal action of various agents on yeasts with minimal amounts of antifungal agents.

• Bulletin of the Korean Chemical Society
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• v.35 no.7
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• pp.1933-1938
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• 2014

Despite increasing importance of in vitro cell-based assays for the assessment of nanoparticles (NPs) cytotoxicity, their suitability for the assessment of NPs toxicity is still in doubt. Here, limitations of widely used cell viability assay protocol (i.e., MTT asssay) for the cytotoxicity assessment of P25 $TiO_2$ NPs were carefully examined and an alternative toxicity assessment method to overcome these limitations was proposed, where the artifacts caused by extracellularly formed formazan and light scattered by agglomerated NPs were minimized by measuring only the intracellular formazan via image cytometric methods.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.31 no.6
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• pp.920-926
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• 1998

To investigate the effects of yellow loess on the microbial community after applying into C. polykrikoides as a red tide centrol method during decomposition process, we conducted this study using microcosm experiments, which consisted of sediment collected from Jinhae and Masan bay. The composition, number of bacteria and respiratory electron transport system activity (ETSA) were analyzed. The number of heterotrophic bacteria examined in the samples of both stations reached maximum value within 12 hrs with $10^7$ cells/dry g, independent with the yellow loess applied. In addition, a differenee in the variation of heterotrophic bacterial composition was not observed by adding the yellow loess, and Vibrio spp. always appeared during the culture periods, However, in day 8 culture, the sulfate reducing bacteria was $3.8\times10^7$ cells/dry g in Masan bay and $5.5\times10^6$ cells/dry g in Jinhae bay samples without yellow loess, and these were 120, 350 fold-and 160, 420 fold-increased when yellow loess was added (1 : 1, 1 : 2). The average ETSA was 6.8$\~$7.6 $\mu$g formazan $h^{-1}$ dry $g^{-1}$ independently with yellow loess in aerobic condition for both samples, but activity was decreased by addition of yellow loess in anaerobic. Thus the addition of yellow loess to marine sediment seems to have an effect to inhibit the anaerobic decomposition process and growth of sulfate reducing bacteria which lead to the bad condition of marine environments.

• Korean Journal of Food Science and Technology
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• v.33 no.3
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• pp.378-383
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• 2001

Formation of superoxide anion $O_2\bar{{\bullet}}$ in the autoxidation of L-ascorbic acid (AsA) in the presence of heavy metal ions were determined. The generation of $O_2\bar{{\bullet}}$ was studied by using superoxide dismutase (SOD) in aqueous and buffer solution, and using nitro bule tetrazolium (NBT) in methanol solution. The remaining amount of AsA was significantly higher in the presence of SOD than in its absence. It suggested that SOD stabilizes AsA in aqueous and buffer solution because of scavenging $O_2\bar{{\bullet}}$ formed during the autoxidation reaction of AsA in the presence of heavy metal ions. NBT has an absorption maximum at about 560 nm in methanol solution. The absorbance at 560 nm increased during the oxidation of AsA, suggested the formation of $O_2\bar{{\bullet}}$in methanol solution. Thus, the formation of $O_2\bar{{\bullet}}$ was confirmed during the autoxidation of AsA not only in aqueous solution but also in methanol solution in the presence heavy metal ions.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.40 no.3
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• pp.307-311
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• 2014

2,3,5-Triphenyltetrazolium chloride (TTC) is used as a redox indicator in culture media. It is colorless in the oxidized form and is reduced to formazan, an insoluble pigment, by dehydrogenases in actively growing microbial cells. The aim of this study was to assess by microbial test of the pathogenic microorganisms using TTC reduction. The pathogenic microorganisms were reduced in medium by dehydrogenase to produce insoluble red formazan. We observed that the optimization method of TTC allowed more than 12 h incubation in 0.04% concentration. Also, the growth of microorganisms with media was increased formazan production. We confirmed that microorganisms were quickly observed to grow colonies cultured red color without affecting the growth of microorganisms. It is suggested that the microbial test using TTC can provide better and quicker test method in cosmetics development.

• The Korean Journal of Zoology
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• v.15 no.3
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• pp.101-104
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• 1972

A simple and economical method for separation of lactate and malate dehydrogenase isozymes is described in detail. The method is based on cellulose acetate strip electrophoretic separation of the isozymes, tetrazolium reduction to purple formazan. Resolution is as good as in the experiment using expensive equipments.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.4
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• pp.494-498
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• 2008

MTT (3-(4, 5-dimethyl thiazol-2-yl)-2, 5-diphenyl tetrazolium bromide) reduction assay is a method that validates the viability of an active cell. Dehydrogenase in mitochondria converts yellow colored insoluble tetrazolium salt to purple colored water-soluble formazan. Sperm also have mitochondria in the midpiece, therefore sperm viability could be evaluated by MTT reduction assay. Several studies have already demonstrated the capability of application of the MTT reduction assay to sperm of several species in Hepes-BSA buffer. Because most liquid semen was diluted in extender like BTS (Beltsville Thawing Solution), Modena or Androhep when it is used or transferred, semen needed another dilution in Hepes-BSA buffer to assess sperm viability. In this study, we evaluated boar sperm viability especially in BTS extended semen and compared the efficiency of this test with eosin-nigrosin staining. We used the fresh BTS extended semen from a local A.I center. Semen sample was diluted to $3.0{\times}10^7$ sperms/ml in BTS. The rates of formazan production were measured in 96-well microtiter plates immediately and 1h after incubation at $17^{\circ}C$ using a spectrophotometer at wave length 560 nm. Simultaneously, split samples of the same semen were tested, using eosin-nigrosin staining to compare the efficiency of the MTT assay of sperm viability in BTS. The correlation between the results of these tests was calculated using Student-t test and ANOVA. The results revealed a strong correlation between the results of MTT reduction rate and the results that were simultaneously determined by eosin-nigrosin staining at 1 h. In conclusion, the MTT reduction test was an effective and simple method to validate sperm viability and it could be used as a simple tool to evaluate sperm viability in the local A.I center and laboratory.