Yeast cell mortality related to a high-pressure shift: occurrence of cell membrane permeabilization

The shrinkage of yeast cells caused by high-pressure treatment (250 MPa, 15 min) was investigated using direct microscopic observation. A viable staining method after treatment allowed the volume variation of two populations to be distinguished: an irreversible volume decrease (about 35% of the initial volume) of pressure-inactivated cells during pressure holding time, and viable cells, which were less affected. A mass transfer was then induced during high-pressure treatment. Causes of this transfer seem to be related to a pressure-induced membrane permeabilization, allowing a subsequent leakage of internal solutes, where three ions (Na+, K+ and Ca2+), plus endogenous glycerol, were verified. This glycerol leakage was found to occur after yeast pressurization in a medium having low water activity, although the yeast was not inactivated. All these observations lead to the hypothesis that pressure-induced cell permeabilization could be the cause of yeast inactivation under pressure.     

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Amine oxidase in unicellular microorganismsMethanosarcina barkeri andTetrahymena pyriformis

A comparison has been performed of catalytic properties of unicellular microorganism amine oxidases (AO) from two new enzyme sources, the bacteriumMethanosarcina barkeri and the infusoriaTetrahymena pyriformis. It was shown that the both studied AO deaminate tyramine, serotonin, and benzylamine, but do not deaminate histamine. The AO fromMethanosarcina barkeri catalyzes deamination of all three substrates at an identical rate, while the rate of tyramine deamination under effect of AO fromTetrahymena pyriformis is one order higher than the rate of serotonin deamination, and about two orders higher than the rate of benzylamine deamination. Based on the data of the substrate-inhibitor analysis, a suggestion was made about the existence of one center for the substrate binding in the AO of the studied bacterium, while several centers in the AO of the studied infusoria.     

A rapid colorimetric assay for heparinase activity.

A rapid, sensitive, assay for enzymes that degrade heparin is described. The procedure is based on the interference of heparin with color development during the interaction of protein with the dye Coomassie brilliant blue. The loss of this property when the glycosaminoglycan is degraded by heparinase can be used to quantify activity of the enzyme in pure form, or in complex biological samples such as tissue homogenates or serum. The assay is also suitable for studying dependence of heparinase activity under conditions such as varying pH and temperature.     

Amperometric alcohol electrode with extended linearity and reduced interferences.

Conventional amperometric alcohol electrodes were constructed with oxygen- and hydrogen peroxide-base sensors and a much improved electrode was designed by placing a hydrophobic, gas-permeable membrane over the conventional hydrogen peroxide-based alcohol electrode. The immobilization of alcohol oxidase with glutaraldehyde was also studied and optimized. The upper linear ranges of the conventional and newly designed alcohol electrodes were 0.02 and 0.5% ethanol, respectively. The hydrophobic membrane of the new design eliminated the classical electrochemical interferences of hydrogen peroxide-based electrodes and the typical pH dependence of enzymatic systems.     

Procedure and Analysis of a Useful Method in Determining Mycelial Dry Weights from Agar Plates

The evaluation of growth by dry weight determination of fungus mycelium for agar plates was examined. The data obtained were statistically analyzed. This method was shown to be sufficiently accurate to be used as an investigative tool.