On the occurrence of cellulolytic activity in the digestive gland of some marine carnivorous molluscs

• 1.1. The occurrence of a complete cellulolytic system has been demonstrated in the digestive gland of 8 Mediterranean species of carnivorous molluscs belonging to Cephalopoda and Gastropoda.
• 2.2. Although no specific method for assaying the activity of component C₁ is available because of the unspecified action of this biocatalyst whose presence is required for the hydrolysis of highly ordered cellulose by the component C_x, an indirect indication of the contribution given by the component C₁ to the overall cellulolytic activity was obtained using an assay procedure which also permitted the evaluation of the contribution of the third component of the enzyme system, i.e. β-glucosidase.
• 3.3. Results are discussed and a probable physiological significance of such a system is suggested.

     

A semi-micro quantitative assay for cellulolytic activity in microorganisms

A semi-micro quantitative assay for microbial cellulytic activity has been developed. Cellulose-azure substrate was incorporated into the upper of two agar layers in a small tube. Dye was released from this substrate as a result of the action of cellulolytic organisms. The amount of dye released was determined densitometrically, and is a measure of cellulolytic activity. Standardisation of inoculum size and calibration of the system with known amounts of cellulase allowed expression of such activity in terms of units of cellulase per mg of inoculum protein. The assay is simple, quick and relatively inexpensive. It is suitable for use in screening programmers and may be applicable to other enzymes using dye—substrate complexes.     

Simple cultural test for relative cellulolytic activity of fungi

A simple method is described for determining the relative cellulolytic activity of fungi. Opaque columns of an agar medium containing a partially crystalline cellulose preparation were inoculated with the fungi. Depth of the clear zone that developed beneath the growing cultures provided a visual measure of cellulolytic activity on a continuous, cumulative basis. Depth of clearing (DC) was determined for 25 species of fungi differing widely in cellulolytic activity, and compared by correlation analysis with results of three other methods for measuring cellulolytic activity. Relatively high coefficients of correlation (greater than 0.6) were obtained between DC and weight loss of cotton sliver, loss in tensile strength of cotton duck, and carboxymethyl cellulase activity in culture filtrates. In comparison with conventional assay procedures, the clearing method offered several advantages: (i) results were at least as well correlated with the capacity to utilize native cellulose as a substrate; (ii) the method measured activity of growing cultures rather than culture filtrates, thus involving less risk of losses due to product inhibition, binding, or denaturation of enzymes; (iii) repeated measurements were made on the same experimental set up, so that errors due to arbitrarily selected times of harvest were avoided conveniently; and (iv) the method required less working time and very simple equipment, making it convenient for large-scale screening tests.     

Endowing non-cellulolytic microorganisms with cellulolytic activity aiming for consolidated bioprocessing

With the exhaustion of fossil fuels and with the environmental issues they pose, utilization of abundant lignocellulosic biomass as a feedstock for biofuels and bio-based chemicals has recently become an attractive option. Lignocellulosic biomass is primarily composed of cellulose, hemicellulose, and lignin and has a very rigid and complex structure. It is accordingly much more expensive to process than starchy grains because of the need for extensive pretreatment and relatively large amounts of cellulases for efficient hydrolysis. Efficient and cost-effective methods for the production of biofuels and chemicals from lignocellulose are required. A consolidated bioprocess (CBP), which integrates all biological steps consisting of enzyme production, saccharification, and fermentation, is considered a promising strategy for reducing production costs.     

Comparison of HPLC and colorimetric methods for measuring cellulolytic activity

Summary The sugars released during a standard filter paper assay were quantified by colorimetric (Dinitrosalicylic acid and Nelson and Somogyi) and HPLC methods. It was found that the composition of the sugars present in the hydrolysates greatly influenced the final filter paper units of activity obtained. In a -glucosidase deficient cellulase system the filter paper activity reported using colorimetric methods could be enhanced by adding supplemental amounts of -glucosidase. However, HPLC methods of quantitating filter paper activity were not affected provided the accumulated cellobiose did not inhibit the cellulase enzymes. Glucose oxidase methods of assaying -glucosidase activity were also influenced by wood decomposition products present in the culture filtrates. HPLC methods of quantifying hydrolysates gave the most representative values of the hydrolytic potential of the cellulase enzyme complex.     

Mechanoptical transducer for quantifying activity in small insect muscles

Contraction strength, frequency, rate, relaxation rate, as well as transient and static muscle lengths were quantified for onion fly (Delia antiqua) oviducal muscle to demonstrate the application of a newly developed, sensitive, rapidly responding, stable and linear mechanoptical transducer system. Contractile patterns were also differentiated for the complex array of muscle segments in the oviduct. Computer-assisted analysis of analogue records showed that within-train contraction strength varied inversely as a function of contraction frequency in a train. Also, a tonic component, assumed to be a function of contraction frequency and the viscoelastic properties of the tissue, was superimposed on trains of phasic, longitudinal contractions. The transducer system described in this report provides opportunities to quantify contraction phenomena occurring at intervals approaching 1 ms in small (nom. 1 mm) tissue samples with resolution in the order of 1 μg of force and 10 μm of displacement.     

Characterization of the cellulolytic activity of a Bacillus isolate

A group I Bacillus strain, DLG, was isolated and characterized as being most closely related to Bacillus subtilis. When grown on any of a variety of sugars, the culture supernatant of this isolate was found to possess cellulolytic activity, as demonstrated by degradation of trinitrophenyl-carboxymethyl cellulose. Growth in medium containing cellobiose or glucose resulted in the greatest production of cellulolytic activity. The cellulolytic activity was not produced until the stationary phase of growth, and the addition of glucose or cellobiose to a culture in this phase had no apparent effect on enzyme production. Fractionation of the culture supernatant showed that the molecular weight of the enzymatic activity was less than 100,000. Maximum cellulolytic activity in assays was observed at pH 4.8 and at 58C, although maximum thermal stability of the activity. Kinetic experiments suggested that more than one enzyme was acting upon trinitrophenyl-carboxymethyl cellulose. Exocellular protein produced by this Bacillus isolate showed roughly one-fifth the cellulolytic activity displayed by Trichoderma reesei C30 on noncrystalline, cellulosic substrates. In contrast to T. reesei cellulase, the Bacillus enzymatic activity showed no ability to degrade crystalline forms of cellulose, nor was cellobiase activity detectable.     

Cellulose degradation and cellulase activity of five cellulolytic fungi

Biodegradation of pure cellulose powder, bagasse and wheatstraw by five cellulolytic fungi,Aspergillus niger, Chaetomium globosum, Scopulariopsis brevicaulis, Trichoderma koningii andTrichothecium roseum, was studied in solid culture conditions. Minimum degradation was with pure cellulose. Bagasse and wheatstraw were the most suitable for growth and activity of cellulolytic fungi. All fungi contained cellulase activity.

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Résumé On a étudié la biodégradation de la poudre de cellulose pure, de la bagasse et de la paille de froment chez cinq moisissures cellulolytiques,Aspergillus niger, Chaetomium globosum, Scopulariopsis brevicaulis, Trichoderma koningii etTrichothecium roseum, en condition de culture sur milieu solide. La dégradation minimum a lieu avec la cellulose pure. La bagasse et la paille de froment offrent la meilleure croissance et la meilleure activité cellulolytique fungale. Toutes les moisissures exhibent l'activité de la cellulase.

     

Antibiotic inhibition of pectolytic and cellulolytic enzyme activity in twoFusarium species

Among all antibiotics tested, amoxycillin (500 ppm) completely inhibited the polygalacturonase and pectinmethylgalacturonase enzyme activity inF. oxysporum; none of the antibiotics did so inF. moniliforme. No antibiotic completely inhibited the cellulase activity in both test organisms, however, amoxycillin was better than other antibiotics in inhibiting the cellulase activity in both the organisms.     

Stability of AtVSP in the insect digestive canal determines its defensive capability

We have previously demonstrated that Arabidopsis vegetative storage protein (AtVSP) is an acid phosphatase that has anti-insect activity in in vitro feeding assays [Liu et al., 2005. Plant Physiology 139, 1545-1556]. To investigate the functionality of AtVSP in planta as an anti-insect defense protein, we produced AtVSP-overexpressing as well as AtVSP-silenced transgenic Arabidopsis lines, and evaluated impact on the polyphagous American grasshopper Schistocerca americana. Grasshoppers showed no significant difference in weight gain and growth rate when feeding on wild type, overexpressing, or silenced lines, respectively. In addition, AtVSP protein was undetectable in either the midgut or frass of grasshoppers reared on transgenic plants suggesting that AtVSP was unable to withstand proteolytic degradation. To determine the stability of the AtVSP protein in grasshopper digestive canal, midgut extracts from various nymphal stages were incubated with bacterially expressed AtVSP for different periods of time. AtVSP was hydrolyzed rapidly by grasshopper midgut extract, in stark contrast with its fate when incubated with cowpea bruchid midgut extract. Multiple proteases have been detected in the midgut of grasshoppers, which may play important roles in determining the insect response to AtVSP. Results indicate that stability of an anti-insect protein in insect guts is a crucial property integral to the defense protein.     

Determination of the cellulolytic activity ofMicromycetes by an express method

An express method of testing the cellulolytic activity ofTrichoderma viride in media with cellulose-containing substrates by the rate of glucose production has been elaborated. The optimum temperature selections and duration of sample incubation (1 h, 50°C) have been described, and the temperature coefficient for the scale-down of the enzyme complex from 50 to 30°C (0.77) has been determined.     

Supplementation with skim milk enhances the cellulolytic activity of fungi

Summary Addition of skim milk powder to Reese medium increased cellulolytic activity of T. reesei. Exoglucanase (filter paper) activity increased by 6.6, 5.3 and 2.2 folds when estimated on 5,10 and 15th day respectively in presence of skim milk (0.2%) as compared to its control without supplements. The endoglucanase (CMCase) activity improved in the same pattern. The xylanase activity increased by 2.3 fold when estimated on 5th day and maintained the improvement upto 10th day. The -glucosidase activity remained unaltered. The cellulolytic activities of a few other fungal cultures improved in the same manner in the presence of skim milk.