Measuring fragmentation of cellulose by Thermomonospora fusca cellulase

• Published in: Enzyme and microbial technology Vol. 12; no. 5; pp. 378 - 386
• Main Authors: Walker, L.P., Wilson, D.B., Irwin, D.C.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier Inc 1990; Elsevier Science
• Subjects: Analytical biochemistry: general aspects, technics, instrumentation; Analytical, structural and metabolic biochemistry; Biological and medical sciences; cellulase; Cellulose; fragmentation; Fundamental and applied biological sciences. Psychology; soil microorganisms; thermomonospora fusca; thermophilic bacteria; thermomonospora fusca; fragmentation; Cellulose; cellulase; Enzyme; Micromonosporaceae; Theoretical study; Thermomonospora fusca; Fragmentation; Actinomycetales; Hydrolysis; Enzymatic activity; Cellulase; Microorganism culture; Bacteria; Actinomycetes; Carbohydrate; Quantitative analysis
• ISSN: 0141-0229; 1879-0909
• DOI: 10.1016/0141-0229(90)90168-P

An analytical and experimental framework is presented and used to assess the fragmentation of cellulosic particles by concentrated crude cellulase from the thermophilic soil bacterium Thermomonospora fusca. Avicel PH 102 which passed through a 150 mesh sieve and collected on a 200 mesh sieve was the substrate used in a series of hydrolysis experiments that were conducted at cellulase concentrations of 0.00, 0.05, 0.1, 0.25, and 0.50 mg ml −1. All experiments were conducted with a substrate concentration of 5%. During each experiment, samples were removed for determination of reducing sugars, protein, and particle size distribution. Reducing sugar production followed the classical nonlinear pattern with both time and enzyme concentration. An extent of binding of 78% was observed in all experiments except the one using an initial enzyme concentration of 0.50 mg ml −1, where the extent of binding was 66%. Analysis of the particle counting results showed rapid increases in the number of particles. Analysis of the volume fraction distributions revealed that a bimodal distribution was created as a result of particle fragmentation. The values of apparent mean volume decreased nearly linearly with time. The rate of change in the apparent mean particle volume with time increased linearly with increasing concentration of bound cellulase. Efforts to estimate true mean particle volume were hampered by the large variance in the measurement of apparent total volume. The results showed that fragmentation of cellulose can be qualitatively determined with a particle counter and that analysis of fragmentation can provide additional insights into the mechanisms of enzymatic hydrolysis.