Sequential Injection Kinetic Flow Assay for Monitoring Glycerol in a Sugar Fermentation Process by Saccharomyces cerevisiae

A sequential injection system to monitor glycerol in a Saccharomyces cerevisiae fermentation process was developed. The method relies on the rate of formation of nicotinamide adenine dinucleotide in its reduced form (NADH, measured spectrophotometrically at 340 nm) from the reaction of glycerol with...

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Bibliographic Details
Published in:Applied biochemistry and biotechnology 2010-03, Vol.160 (6), p.1664-1673
Main Authors: Domínguez, Karina B. Hueso, Tóth, Ildikó V, Souto, M. Renata S, Mendes, Filipa, García De María, Cándido, Vasconcelos, Isabel, Rangel, António O. S. S
Format: Article
Language:English
Subjects:
Biochemistry
Biological and medical sciences
Biological Assay - methods
Biotechnology
Carbohydrates - chemistry
Chemical engineering
Chemistry
Chemistry and Materials Science
Chromatography, High Pressure Liquid
fermentation
Fermentation - drug effects
Flow Injection Analysis - methods
Fundamental and applied biological sciences. Psychology
Glucose - analysis
glycerol
Glycerol - analysis
glycerol dehydrogenase
high performance liquid chromatography
Kinetics
Methods. Procedures. Technologies
Microbial engineering. Fermentation and microbial culture technology
Monitoring systems
NAD (coenzyme)
NAD - pharmacology
Reference Standards
Saccharomyces cerevisiae
Saccharomyces cerevisiae - drug effects
Saccharomyces cerevisiae - metabolism
Solutions
spectrophotometers
sugars
Time Factors
Yeast
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Summary:A sequential injection system to monitor glycerol in a Saccharomyces cerevisiae fermentation process was developed. The method relies on the rate of formation of nicotinamide adenine dinucleotide in its reduced form (NADH, measured spectrophotometrically at 340 nm) from the reaction of glycerol with NAD⁺ cofactor, catalysed by the enzyme glycerol dehydrogenase present in solution. This procedure enables the determination of glycerol between 0.046 and 0.46 g/l, (corresponding to yeast fermentation samples with concentrations up to 50 g/l) with good repeatability (relative standard deviation for n = 10 lower than 2.2% for three different samples) at a sampling frequency of 25/h. The detection and quantification limits using a miniaturised spectrophotometer were 0.13 and 0.44 mM, respectively. Reagent consumption was of 0.45 μmol NAD⁺ and 1.8 μg enzyme per assay, and the waste production was 2.8 ml per determination. Results obtained for samples were in agreement with those obtained with a high-performance liquid chromatography method.
ISSN:0273-2289
1559-0291
DOI:10.1007/s12010-009-8675-z