[Show abstract][Hide abstract] ABSTRACT: A simple and rapid isothermal absorptiometric assay for detection of viable microbes was studied by employing a redox color indicator of 2,6-dichlorophenolindophenol (DCPIP). Absorbance of DCPIP decreased at 600 nm due to redox reaction occurring between DCPIP and surface membrane of viable microbes, and was found to be inversely proportional to the viable cell density. The redox reaction was found with not only bacteria, but also yeast and their mixture. In the present assay, influences of light scattering and absorption caused by microbial cells and coexisting substances in a sample were excluded by time difference method. The assay required only 10 min for one incubation mixture, and highly repeatable results of three consecutive measurements were obtained by isothermal incubation during certain time at 30°C using a thermostable three-cuvettes-stir system. Thus, cell density of microbial cell suspension or growth medium was successfully determined, and practical lower detection limit for food inspection was obtained at 10(4) ∼ 10(6) cfu/mL. Single cell effects on DCPIP reduction were evaluated and compared between different species. Consequently, this assay was expected to be useful tool for rapid measurement of viable microbes as a preliminary assay of Hazard Analysis Critical Control Point (HACCP) program.
No preview · Article · Jul 2013 · Analytical Biochemistry
[Show abstract][Hide abstract] ABSTRACT: Salt-tolerant yeast Saccharomyces cerevisiae ARIF KD-003 was applied to highly sensitive and reproducible absorbance-based biochemical oxygen demand (BOD(AB-ScII)) measurement for seawater. In the previous work, we have studied the BOD(AB-ScI) method using normal Baker's yeast S. cerevisiae, and the excellent feature of the Baker's yeast as uniformly sustainable in solution could successfully be utilized. However, the BOD(AB-ScI) responses were disappeared by the existence of chloride ion as well as seawater. In the present method, uniformity in solution was also observed with S. cerevisiae ARIF KD-003, and salt-tolerance of the yeast was observed even in saturate concentration of sodium chloride. Next, characterizations of the influences of pH and incubation temperature were investigated. After optimum conditions were obtained, two calibration curves were made between 0.33 and 22 mg O2 L(-1) BOD using standard solution of glucose glutamic acid (GGA) or mixture of GGA and artificial seawater. Then, excellent reproducibility as the averages of relative standard deviation (R.S.D.(av)) in two calibration curves (nine points each) was successfully obtained at 1.10% at pure water or 1.03% at artificial seawater standard, respectively. In addition, the 3 sigma lower detection limit was calculated to be 0.07 mg O2 L(-1) BOD, and 0.11 mg O2 L(-1) BOD was experimentally detected by increase of the sample volume at 1.5-folds. The storage stability of the S. cerevisiae ARIF KD-003 was obtained at least 4 weeks.
No preview · Article · Aug 2008 · Analytica chimica acta