Nephelometric and turbidometric assays of cellulase activity

Extensively ball-milled cellulose fibers were used as natural substrate for the determination of cellulase activity. This physical treatment breaks the large cellulose fibers to small but insoluble particles yielding a substrate accessible for complete enzymatic breakdown. The parameters studied to estimate the activity of cellulases were (a) the decrease in optical density of ball-milled suspension of fibers and (b) simultaneous measurement of liberated sugars during hydrolysis. A good correlation was found between the initial rate of reaction and the amount of sugar released at given times.     

Comparison of two lignin-degrading fungi:Phlebia radiata andPhanerochaete chrysosporium

Summary The ligninolytic enzymes ofPhlebia radiata were produced in static conditions earlier developed forPhanerochaete chrysosporium. The production pattern of lignin peroxidases resembled that ofP. chrysosporium. The extracellular proteins ofPhlebia radiata were separated by isoelectric focusing. Four proteins with acidic isoelectric points (4.15) were detected by peroxidase staining. The peroxidases ofP. radiata reacted with antibodies produced against a peroxidase ofPhanerochaete chrysosporium and vice versa. Thus the lignin peroxidases of the two fungi have major similarities despite slight differences in their isoelectric points and molecular weights. Veratryl alcohol was produced by both fungi and degraded to veratraldehyde, two lactones and a quinone by the ligninolytic cultures.     

A new appraisal of the endoglucanases of the fungus Trichoderma reesei.

The properties and enzymic activity of endoglucanases (EC 3.2.1.4) of the fungus Trichoderma reesei were studied by means of immunological methods and by using polyglycosidic substrates. Endoglucanases exist in the culture liquid as a series of immunologically related components. The most active endoglucanase component has an Mr of 43 000 and pI value of 4.0. The most abundant components have a value of pI about 5.0, an Mr of 56 000-67 000 and specific activity only one-fifth of that of the pI-4.0 component. During purification and storage the endoglucanases are spontaneously modified; the relative proportion of components having greater Mr values, more alkaline pI values and lower specific activities is increased. The hexose content of the endoglucanase components is 2-7%. Endoglucanases hydrolyse soluble beta-1,4 glycans. The enzymes described here differ from endoglucanase preparations described previously in not showing activity towards insoluble substrates. The role of endoglucanases in wood hydrolysis is consequently limited to the stage where wood constituents are already in soluble form.     

Formation of Fusarium metabolites in barley grain

Summary The influence of moisture content and temperature during storage of grain on the formation of Fusarium metabolites was studied. Naturally and artificially contaminated barley grain samples were stored at 15%, 25%, and 30% moisture contents, and at temperatures of + 5°C + 25°C, and + 30°C. Time of storage varied between one week and six months and the occurrence of Fusarium species and metabolites was analysed. The only Fusarium metabolite detected was zearalenone. The extent of Fusarium contamination decreased during storage whilst the concentration of zearalenone increased. To avoid the danger of mycotoxicoses, grains must be dryed immediately after harvest and then stored at a low temperature.     

Differential response of cycling and noncycling cells to inducers of DNA synthesis and mitosis

The objective of this study was to determine whether cells in G(0) phase are functionally distinct from those in G(1) with regard to their ability to respond to the inducers of DNA synthesis and to retard the cell cycle traverse of the G(2) component after fusion. Synchronized populations of HeLa cells in G(1) and human diploid fibroblasts in G(1) and G(0) phases were separately fused using UV-inactivated Sendai virus with HeLa cells prelabeled with [(3)H]ThdR and synchronized in S or G(2) phases. The kinetics of initiation of DNA synthesis in the nuclei of G(0) and G(1) cells residing in G(0)/S and G(1)/S dikaryons, respectively, were studied as a function of time after fusion. In the G(0)/G(2) and G(1)/G(2) fusions, the rate of entry into mitosis of the heterophasic binucleate cells was monitored in the presence of Colcemid. The effects of protein synthesis inhibition in the G(1) cells, and the UV irradiation of G(0) cells before fusion, on the rate of entry of the G(2) component into mitosis were also studied. The results of this study indicate that DNA synthesis can be induced in G(0)nuclei after fusion between G(0)- and S-phase cells, but G(0) nuclei are much slower than G(1) nuclei in responding to the inducers of DNA synthesis because the chromatin of G(0) cells is more condensed than it is in G(1) cells. A more interesting observation resulting from this study is that G(0) cells is more condensed than it is in G(1) cells. A more interesting observation resulting from this study is that G(0) cells differ from G(1) cells with regard to their effects on the cell cycle progression of the G(2) nucleus into mitosis. This difference between G(0) and G(1) cells appears to depend on certain factors, probably nonhistone proteins, present in G(1) cells but absent in G(0) cells. These factors can be induced in G(0) cells by UV irradiation and inhibited in G(1) cells by cycloheximide treatment.     

Exercise training in childhood-onset systemic lupus erythematosus: a controlled randomized trial

Introduction

Exercise training has emerged as a promising therapeutic strategy to counteract physical dysfunction in adult systemic lupus erythematosus. However, no longitudinal studies have evaluated the effects of an exercise training program in childhood-onset systemic lupus erythematosus (C-SLE) patients. The objective was to evaluate the safety and the efficacy of a supervised aerobic training program in improving the cardiorespiratory capacity in C-SLE patients.

Methods

Nineteen physically inactive C-SLE patients were randomly assigned into two groups: trained (TR, n = 10, supervised moderate-intensity aerobic exercise program) and non-trained (NT, n = 9). Gender-, body mass index (BMI)- and age-matched healthy children were recruited as controls (C, n = 10) for baseline (PRE) measurements only. C-SLE patients were assessed at PRE and after 12 weeks of training (POST). Main measurements included exercise tolerance and cardiorespiratory measurements in response to a maximal exercise (that is, peak VO₂, chronotropic reserve (CR), and the heart rate recovery (ΔHRR) (that is, the difference between HR at peak exercise and at both the first (ΔHRR1) and second (ΔHRR2) minutes of recovery after exercise).

Results

The C-SLE NT patients did not present changes in any of the cardiorespiratory parameters at POST (P > 0.05). In contrast, the exercise training program was effective in promoting significant increases in time-to-exhaustion (P = 0.01; ES = 1.07), peak speed (P = 0.01; ES = 1.08), peak VO₂ (P = 0.04; ES = 0.86), CR (P = 0.06; ES = 0.83), and in ΔHRR1 and ΔHRR2 (P = 0.003; ES = 1.29 and P = 0.0008; ES = 1.36, respectively) in the C-SLE TR when compared with the NT group. Moreover, cardiorespiratory parameters were comparable between C-SLE TR patients and C subjects after the exercise training intervention, as evidenced by the ANOVA analysis (P > 0.05, TR vs. C). SLEDAI-2K scores remained stable throughout the study.

Conclusion

A 3-month aerobic exercise training was safe and capable of ameliorating the cardiorespiratory capacity and the autonomic function in C-SLE patients.

Trial registration

NCT01515163.     

Survival Rate and Enzyme Activities ofLactobacillus acidophilusFollowing Frozen Storage

The ability of two strains of Lactobacillus acidophilus, CRL 640 and CRL 800, to survive and retain their biological activities under frozen storage was determined. Freezing and thawing, as well as frozen storage, damaged the cell membrane, rendering the microorganisms sensitive to sodium chloride and bile salts. Both lactic acid production and proteolytic activity were depressed after 21 days at -20 degreesC, whereas beta-galactosidase activity per cell unit was increased. Cell injury was partially overcome after repair in a salt-rich medium. Copyright 1998 Academic Press.     

Oxidation of milled wood lignin with laccase, tyrosinase and horseradish peroxidase

In this paper the oxidation of milled wood lignin (MWL), catalysed by three enzymes, i.e. laccase, tyrosinase and horseradish peroxidase (HRP) was studied. The oxidation was followed by measuring the consumption of O₂ during laccase and tyrosinase treatment and of H₂O₂ during HRP treatment. Both laccase and HRP were found to oxidise lignin effectively, whereas the effect of tyrosinase was negligible. The changes in MWL molecular-weight distributions caused in the reactions were analysed by gel permeation chromatography. Both laccase and HRP treatments were found to polymerise MWL. Peroxidase treatment was found to decrease the amount of phenolic hydroxyls in MWL, whereas no such effect could be detected in the laccase-treated sample. Both laccase and HRP treatments were, however, found to increase the amount of conjugated structures in MWL. The formation of phenoxy radicals during the treatments was studied by electron paramagnetic resonance spectroscopy. Phenoxy radicals were detected in both laccase and HRP-treated samples. The amount of the formed phenoxy radicals was found to be essentially constant during the detected time (i.e. 20–120 min after the addition of enzyme).     

TNF-α is associated with loss of lean body mass only in already cachectic COPD patients

Background

Systemic inflammation may contribute to cachexia in patients with chronic obstructive pulmonary disease (COPD). In this longitudinal study we assessed the association between circulating C-reactive protein (CRP), tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6 levels and subsequent loss of fat free mass and fat mass in more than 400 COPD patients over three years.

Methods

The patients, aged 40–76, GOLD stage II-IV, were enrolled in 2006/07, and followed annually. Fat free mass and fat mass indexes (FFMI & FMI) were calculated using bioelectrical impedance, and CRP, TNF-α, IL-1ß, and IL-6 were measured using enzyme immunoassays. Associations with mean change in FFMI and FMI of the four inflammatory plasma markers, sex, age, smoking, FEV₁, inhaled steroids, arterial hypoxemia, and Charlson comorbidity score were analyzed with linear mixed models.

Results

At baseline, only CRP was significantly (but weakly) associated with FFMI (r = 0.18, p < 0.01) and FMI (r = 0.27, p < 0.01). Univariately, higher age, lower FEV₁, and use of beta2-agonists were the only significant predictors of decline in FFMI, whereas smoking, hypoxemia, Charlson score, and use of inhaled steroids predicted increased loss in FMI. Multivariately, high levels of TNF-α (but not CRP, IL-1ß or IL-6) significantly predicted loss of FFMI, however only in patients with established cachexia at entry.

Conclusion

This study does not support the hypothesis that systemic inflammation is the cause of accelerated loss of fat free mass in COPD patients, but suggests a role for TNF-α in already cachectic COPD patients.