Characterization and quantification of molybdenum blue production in Enterobacter cloacae Strain 48 using 12 molybdophosphate as the reference compound
M.Y.A. Shukor, N.A. Shamaan*, M.A. Syed, C.H. Lee and M.I.A. Karim1
Department of Biochemistry and Microbiology, Faculty of Science and Environmental Studies,
Universiti Putra Malaysia 43400 UPM Serdang, Selangor, Malaysia
1Department of Biotechnology Faculty, of Food Science and Biotechnology, Universiti Putra Malaysia,
43400 UPM Serdang, Selangor, Malaysia.
Received 2 June 2000 /Accepted 14 December 2000
Abstract.
The absorption spectrum between 400-980 nm of molybdenum blue produced by Enterobacter cloacae strain 48, originally isolated from Chengkau, Malaysia, cultured in static condition in media containing 10 mM molybdate and 2.9 mM phosphate at 30oC, and reduced with 2.5% (w/v) ascorbic acid, was compared with that produced chemically using 12 molybdophosphate and 20 molybdodiphosphate. The absorption spectra of the three products were found to be similar with a shoulder peak at 700-720 nm and a major peak between 860-870 nm. The absorbance values of the spectra increased with increasing fermentation times of E. cloacae Strain 48, indicating that formation of molybdenum blue was dependent on fermentation time. Based on the maximum absorption peak at 865 rim, this wavelength was taken for measurement of molybdenum blue produced in subsequent experiments. A standard curve for the reduction of 12 molybdophosphate by ascorbic acid was constructed by measurement of absorbance at 865 rim. The plot obtained was linear and reproducible (R2, 0.9855; coefficient of variance, < 5%). The extinction coefficient of molybdenum blue at 865 nm was calculated to be 16.7 mM-1cm-1 Production of molybdenum blue in E. cloacae Strain 48 coincided with the drop in pH where molybdenum blue concentration increased after the pH dropped to pH 5.7. The time course development of molybdenum blue colour followed saturation kinetics with a plateau after 150 min incubation time. Reduction of 12 molybdophosphate chemically also followed saturation kinetics with 2.5% (w/v) ascorbic acid achieving maximum reduction after 1 h incubation. The formation of molybdenum blue was inhibited at high phosphate concentration, both chemically and by fermentation. Measurement of absorbance of molybdenum blue at 865 nm was found to be useful in the study of molybdenum reduction in E. cloacae Strain 48.
Keywords: Molybdenum blue, 12 molybdophosphate, absorbance measurement at 865 run, Enterobacter cloacae Strain 48
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