University of North Florida
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Stuart Chalk, Ph.D.
Department of Chemistry
University of North Florida
Phone: 1-904-620-1938
Fax: 1-904-620-3535
Email: schalk@unf.edu
Website: @unf

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Enzyme, galactosidase

  • CAS Registry Number: 9031-11-2

@ ChemSpider@ NIST@ PubChem

Citations 7

"Immobilized Enzyme Kinetics Analysed By Flow-through Micro-fluorimetry. Resorufin β-D-galactopyranoside As A New Fluorogenic Substrate For β-D-galactosidase"
Anal. Chim. Acta 1984 Volume 163, Issue 1 Pages 67-72
J. Hofmann and M. Sernetz

Abstract: A new method for simultaneous integral and dispersive assay of gel-immobilized enzyme activities in continuously stirred tank reactors is described. It combines a conventional, continuous-flow fluorimetric technique for measurement of total turnover rates of the reactor with micro-fluorimetric assay of the activity of individual gel particles. The high sensitivity necessary for single particles is achieved by using the cited substrate, which has the advantage of a one-step hydrolysis following Michaelis - Menten kinetics in addition to its red fluorescence emission. Results are presented for β-D-galactosidase immobilized on Sepharose 4B. By use of sieved gel fractions, effectiveness factors and Damkoehler numbers determined in individual beads can be correlated with integral turnover rates of the reactor.
Fluorescence Kinetic Immobilized enzyme Continuously stirred tank reactor

"Immunosensing With Amperometric Detection, Using Galactosidase As Label And P-aminophenyl-galactopyranoside As Substrate"
Anal. Chim. Acta 1995 Volume 304, Issue 3 Pages 353-359
Már Mássen, Zheng Liu, Tetsuya Haruyama, Eiry Kobatake, Yoshihito Ikariyama and Masuo Aizawa*

Abstract: p-Aminophenyl---galactopyranoside (PAPG) was shown to be a suitable substrate for the amperometric detection of galactosidase activity at neutral pH. The application of this amplification system for immunoassay was demonstrated. The product of the enzyme reaction, p-aminophenol (PAP), was detected at 200 mV, vs. , by flow-injection analysis (FIA), with a 50 nM detection limit. PAPG was hydrolyzed more than 2.5 times faster than p-nitrophenyl---galactopyranoside, by the enzyme. Both PAP and PAPG were stable at pH 7. The galactosidase concentration could be measured down to a concentration of 100 fM, and mouse IgG could be assayed by sandwich immunoassay down to 700 fM. PAPG was found to be a promising reagent for heterogeneous systems, like the one described, and for homogeneous assays of biological fluids.
Biological fluid Immunoassay Amperometry Enzyme Amplification reaction

"Microassay For GM1 Ganglioside β-galactosidase Activity Using High Performance Liquid Chromatography"
J. Chromatogr. B 1988 Volume 426, Issue 1 Pages 75-82
M. Naoi and M. Kondoh, T. Mutoh, T. Takahashi, T. Kojima, T. Hirooka and T. Nagatsu

Abstract: Brain tissue homogenate was incubated, at 37°C for 1 h, with GM1 ganglioside in 50 mM citric acid - 100 mM sodium phosphate buffer (pH 4.4) containing 100 mM NaCl and 0.5% of Na taurodeoxycholate. After heating at 100°C for 2 min and cooling, mobile phase, viz, 0.5 M H3BO3 - NaOH buffer (pH 8.7) was added and the mixture was centrifuged. The supernatant solution was subjected to HPLC, at 65°C, on a column (25 cm x 4.0 mm) of Shimadzu ISA-07/S2504 at 0.6 mL min-1 and with post-column derivatization by heating the eluate with 2% L-arginine - 3% H3BO3 and after cooling, fluorimetric detection at 430 nm (excitation at 320 nm). The calibration graph was rectilinear for 1 mmol of β-galactose.
Brain HPLC Fluorescence Heated reaction Post-column derivatization

"FIA For Down-stream Processing-determination Of β-galactosidase In Viscous Extraction Media"
Bioprocess Eng. 1991 Volume 7, Issue 1-2 Pages 47-52
C. Silfwerbrand-Lindh, L. Nord, L. Häggström and F. Ingman

Abstract: A flow injection analysis method for determination of the enzyme β-galactosidase produced in a fermentation process has been developed. The analytical range was 10^-1000 U/mL and the sample throughput was 40 h-1. The FIA set up was connected directly to the down-stream process and the enzyme activity in the extraction step was measured online. The viscosity of the samples from the process is high causing certain physical effects on the FIA system.
Fermentation broth Viscosity

"Assay Of β-galactosidase Activity By Flow Injection Analysis"
Biotechnol. Techniq. 1991 Volume 5, Issue 5 Pages 389-392
J. Cairó, M. Vidal, A. Villaverde, F. Valero, F. J. Lafuente and C. Solà

Abstract: A novel method to analyze β-galactosidase by Flow Injection Analysis is presented with a linear working range extended to at least 2150 U/mL, being the detection limit 25 U/mL with 55 samples per hour frequency and a RSD of 0.54% versus 2.4% obtained by manual assay. The method was tested with optimal results with samples from Escherichia coli cultures producing β-galactosidase.
Fermentation broth Spectrophotometry Buffer Heated reaction pH

"Simultaneous Online Monitoring Of Intracellular β-galactosidase Activity And Biomass Using Flow Injection Analysis In Escherichia Coli Batch Fermentations"
Biotechnol. Techniq. 1992 Volume 6, Issue 3 Pages 213-218
Valero F., Lafuente F. J., Solà C., Benito A., Vidal M., Cairó J. and Villaverde A.

Abstract: Fermentation sample was sonicated at 4°C for 45 s and subjected to flow injection analysis after mixing with phosphate buffer solution (pH 7.0) containing 2-β-mercaptoethanol. The solution was mixed with 2-nitrophenyl-β-D-galactopyranoside at 47°C before addition of 1 M Na2CO3 and 0.5% Na dodecyl sulfate and detection at 420 nm. Biomass determination was achieved offline at 550 nm. The limit of detection was 25 iu mL-1 of β-galactosidase and the calibration graph was rectilinear for 5100 iu mL-1. The coefficient of variation was 1.5%. A novel method to monitor online intracellular β-galactosidase activity and biomass simultaneously, using flow injection analysis (FIA), was developed. The automatic ultrasonic cell disruption and FIA anal. allow the processing of 10 samples/h with a wide and variable linear working range of β-galactosidase activity and biomass and a max. relative standard deviation of 1.5%. The system was optimized by monitoring biomass and intracellular β-galactosidase activity in E. coli batch fermentation
Bacteria Fermentation broth Spectrophotometry Process monitoring

"Online Determination Of Intracellular β-galactosidase Activity In Recombinant Escherichia Coli Using Flow Injection Analysis (FIA)"
J. Biotechnol. 1991 Volume 20, Issue 1 Pages 95-104
Heinrich-Andreas Kracke-Helm*, Lutz Brandes, Bernd Hitzmann, Ursula Rinas** and Karl Schügerl*

Abstract: A flow injection analysis (FIA) system was developed for the determination of cytoplasmic β-galactosidase activity in recombinant Escherichia coli. The FIA system and its application for online monitoring of β-galactosidase production during cultivation of recombinant E. coli in a 60-l airlift tower loop reactor is described. The results demonstrate that an FIA assay in conjunction with a cell disintegration step can be applied successfully for online monitoring of intracellular protein formation. The method is based on that described by Miller ('Experiments in Molecular Genetics', Cold Spring Harbour Laboratory, New York, USA, 1982). Cells were grown in a Luria broth in an airlift tower loop reactor (loc. cit.). The reactor was equipped with a sterile sampling device to provide a continuous sample flow for analysis of media compounds and by-products. Data acquisition and control of the flow injection system were carried out with the CASFA process management and control system (Frueh et al., Biotech-Forum, 1986, 3, 204) and the FERAS local data management system (Wieneke, Ph.D. Thesis, Univ. Hannover, 1989).
Bacteria Fermentation broth Reactor Process monitoring Computer