Abstract
An assay based on microbial test system (Escherichia coli) was developed to evaluate the antioxidant capacity of phytochemicals, extracts of plants, fruits, and vegetables. E. coli was cultured in Luria-Bertani (LB) medium. Quercetin was used as a standard in microbial test system (MTS) assay because it had high antioxidant activity and was easily and economically obtained. The cultivation times of E. coli entrance into mid-log phase, before and after H2O2 addition for MTS assay were investigated, respectively. The concentrations of H2O2 and tested compounds for MTS assay were also studied. The value (μ30), t = 30 min, the specific growth rate of E. coli in medium containing samples and 6.0 mM H2O2/the specific growth rate in medium containing only H2O2 was calculated to evaluate the antioxidant activity. Pure compounds (quercetin, quercitrin, hyperoside, rutin, and kaempferol-3-O-α-L-rhamnoside), fruits (apple, banana, kiwi fruit), and vegetable (tomato) were used to verify this assay, and the results were compared to three in vitro antioxidant assays (2,2´-azino-bis(3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS), 2,2-diphenyl- 1-picrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) assay). The results indicated the MTS assay based on E. coli without transferring the antioxidant genes under peroxide stress would be useful to define the action of extracts or pure compounds as antioxidants. The proposed method is useful to evaluate antioxidant activity and more biologically relevant because it accounts for some aspects of uptake and location of antioxidants within bacterial cells.
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References
Benzie IF, Strain JJ (1996) The ferric reducing ability of plasma (FRAP) as a measure of “antioxidant power”: the FRAP assay. Anal Biochem 239:70–76
Benzie IF, Strain JJ (1999) Ferric reducing/antioxidant power assay: direct measure of total antioxidant activity of biological fluids and modified version for simultaneous measurement of total antioxidant power and ascorbic acid concentration. Methods Enzymol 299:15–27
Blasa M, Angelino D, Gennari L, Ninfali P (2011) The cellular antioxidant activity in red blood cells (CAA-RBC): a new approach to bioavailability and synergy of phytochemicals and botanical extracts. Food Chem 125:685–691
Brand-Williams W, Cuvelier ME, Berset C (1995) Use of a free radical method to evaluate antioxidant activity. LWT Food Sci Technol 28:25–30
Cao GH, Alessio HM, Cutler RG (1993) Oxygen-radical absorbance capacity assay for antioxidants. Free Radic Biol Med 14:303–311
Chu YF, Sun J, Wu X, Liu RH (2002) Antioxidant and antiproliferative activities of common vegetables. J Agric Food Chem 50:6910–6916
Demple B (1991) Regulation of bacterial oxidative stress genes. Annu Rev Genet 25:315–337
Dong XX, Wang Y, Qin ZH (2009) Molecular mechanisms of excitotoxicity and their relevance to pathogenesis of neurodegenerative diseases. Acta Pharmacol Sin 30:379–387
Duduku K, Rosalam S, Rajesh N (2011) A review of the antioxidant potential of medicinal plant species. Food Bioprod Process 89:217–233
Gil MI, Tomas-Barberan FA, Hess-Pierce B, Kader AA (2002) Antioxidant capacities, phenolic compounds, carotenoids, and vitamin C contents of nectarine, peach, and plum cultivars from California. J Agric Food Chem 50:4976–4982
Guo CJ, Yang JJ, Wei JY, Li YF, Xu J, Jiang YG (2003) Antioxidant activities of peel, pulp and seed fractions of common fruits as determined by FRAP assay. Nutr Res 23:1719–1726
Lee J, Koo N, Min DB (2004) Reactive oxygen species, aging, and antioxidative nutraceuticals. Compr Rev Food Sci Food Saf 3:21–33
Leong LP, Shui G (2002) An investigation of antioxidant capacity of fruits in Singapore markets. Food Chem 76:69–75
Miller NJ, Rice-Evans CA (1997) Factors influencing the antioxidant activity determined by the ABTS+ radical cation assay. Free Radic Res 26:195–199
Miller NJ, Diplock AT, Rice-Evans CA (1995) Evaluation of the total antioxidant activity as a marker of the deterioration of apple juice on storage. J Agric Food Chem 43:1794–1801
Molyneux P (2004) The use of the stable free radical diphenylpicryl-hydrazyl (DPPH) for estimating antioxidant activity. Songklanakarin J Sci Technol 26:211–219
Musa KH, Abdullah A, Jusoh K, Subramaniam V (2011) Antioxidant activity of pink flesh guava (Psidium guajava L): effect of extraction techniques and solvents. Food Anal Methods 4:100–107
Ou BX, Hampsch-Woodill M, Prior RL (2001) Development and validation of an improved oxygen radical absorbance capacity assay using fluorescein as the fluorescent probe. J Agric Food Chem 49:4619–4626
Prevc T, Segatin N, Ulrih NP, Cigic B (2013) DPPH assay of vegetable oils and model antioxidants in protic and aprotic solvents. Talanta 109:13–19
Prior RL, Hoang H, Gu LW, Wu XL, Bacchiocca M, Howard L, Hampsch-Woodill M, Huang DJ, Ou BX, Jacob R (2003) Assays for hydrophilic and lipophilic antioxidant capacity (oxygen radical absorbance capacity (ORACFL)) of plasma and other biological and food samples. J Agric Food Chem 51:3273–3279
Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice-Evans C (1999) Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic Biol Med 26:1231–1237
Smirnova GV, Samoylova ZY, Muzyka NG, Oktyabrsky ON (2009) Influence of polyphenols on Escherichia coli resistance to oxidative stress. Free Radic Biol Med 46:759–768
Smirnova GV, Vysochina GI, Muzyka NG, Samoylova ZY (2010) Evaluation of antioxidant properties of medical plants using microbial test systems. World J Microbiol Biotechnol 26:2269–2276
Storz G, Tartaglia LA, Farr SB, Ames BN (1990) Bacterial defenses against oxidative stress. Trends Genet 6:363–371
Sun J, Chu YF, Wu X, Liu RH (2002) Antioxidant and antiproliferative activities of common fruits. J Agric Food Chem 50:7449–7454
Wang BW, Shi QS, Ouyang YS, Chen YB (2008) Progress in oxyR regulon- the bacterial antioxidant defense system—a review. Acta Microbiol Sin 48:1556–1561
Wang SS, Wang DM, Pu WJ, Li DW (2013) Phytochemical profiles, antioxidant and antimicrobial activities of three Potentilla species. BMC Complement Altern Med 13:321
Wolfe KL, Liu RH (2007) Cellular antioxidant activity (CAA), assay for assessing antioxidants, foods, and dietary supplements. J Agric Food Chem 55:8896–8907
Zhang QB, Li N, Zhou GF, Lu XL, Xu ZH, Li Z (2003) In vivo antioxidant activity of polysaccharide fraction from Porphyra haitanesis (Rhodephyta) in aging mice. Pharmacol Res 48:151–155
Zhang YJ, Wang DM, Yang LN, Zhou D, Zhang JF (2014) Purification and characterization of flavonoids from the leaves of Zanthoxylun bungeanum and correlation between their structure and antioxidant activity. Plos One 9:e105725
Zheng M, Wang X, Doan B, Lewis KA, Schneider TD, Storz G (2001a) Computation-directed identification of OxyR DNA binding sites in Escherichia coli. J Bacteriol 183:4571–4579
Zheng M, Wang X, Templeton LJ, Smulski DR, LaRossa RA, Storz G (2001b) DNA microarray-mediated transcriptional profiling of the Escherichia coli response to hydrogen peroxide. J Bacteriol 183:4562–4570
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This study was funded by the Fundamental Research Funds for the Central Universities (ZD2013010) and Special Fund for Forestry Scientific Research in the Public Interest of China (201304811).
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Dongmei Wang declares that she has no conflict of interest. Dan Zhou declares that she has no conflict of interest. Lina Yang declares that she has no conflict of interest. Zehua Liu declares that she has no conflict of interest. Yawei Zhang declares that he has no conflict of interest.
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Zhou, D., Wang, D., Yang, L. et al. A Modified and Improved Assay Based on Microbial Test System (MTS) to Evaluate Antioxidant Activity. Food Anal. Methods 9, 895–904 (2016). https://doi.org/10.1007/s12161-015-0266-8
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DOI: https://doi.org/10.1007/s12161-015-0266-8