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Humicola lutea 103 is a copper-tolerant fungal strain able to grow in the presence of 300 μg·mL–1Cu2+under submerged cultivation. To prevent the consequences of copper overload, microorganisms have evolved molecular mechanisms that regulate its uptake, intracellular traffic, storage, and efflux. In spite of this avoidance strategy, high heavy-metal concentrations caused distinct and widespread ultrastructural alterations in H. lutea. The mitochondria were the first and main target of the toxic action. The effect of copper on activities of the key enzymes (hexokinase, glucose-6-phosphate dehydrogenase, malate dehydrogenase, and isocitrate dehydrogenase) included in the 3 main metabolic pathways, glycolysis, pentose phosphate pathway, and tricarboxylic acid cycle, was investigated. High metal concentrations exhibited a dramatic negative effect on hexokinase, while the other 3 enzymes showed a significant and dose-dependent stimulation. On the basis of the present and previous results we concluded that the copper-induced oxidative stress plays an important role in the fungal tolerance to high Cu2+concentrations.

In the present study, we examined the adaptive response of the filamentous fungus Humicola lutea 103 to copper stress. Our findings indicate that pretreatment with low concentrations of copper ions led to the development of resistance to high concentrations of the same heavy metal. Spores subjected to 40 µg/ml Cu ++ showed higher resistance to killing by subsequent incubation in 70 or 150 µg/ml Cu ++ compared to the single treatment. Pre-exposure strategy also caused a higher amount of biomass accumulation and recovery of the fungal growth curves to the control curves shape. Moreover, this strategy exerted protective effect against the oxidative damaged proteins in H. lutea cells, which was accompanied by a corresponding accumulation of reserve carbohydrates such as trehalose and glycogen. Adaptive cell response included induction in the activity of the first antioxidative enzyme superoxide dismutase, but catalase activity was independent of copper treatment and pretreatment. Therefore, we concluded that the pre-exposure to mild copper stress induces a coordinated response that decreases the oxidative stress degree in H. lutea cells when this pre-exposure was followed by severe stress.

A cell suspension culture of devil’s claw (Harpagophytum procumbens), an African plant with high medicinal value, was induced from previously obtained callus cultures, cultivated in shake-flasks, and a purification scheme to isolate the main active constituents that accumulated in the biomass was developed. The antioxidant activities of total methanol extracts, collected fractions (phenylethanoids, terpenoids and sugars) and specific active constituents (β-OH-verbascoside, verbascoside and leucosceptoside A) were then evaluated in 2,2-diphenyl-1-picrylhydrazyl (DPPH·) and superoxide anion ( O 2 - ) radical scavenging and oxygen radical absorbance capacity (ORACFL) assays. The most active compounds were found to be β-OH-verbascoside (in DPPH· and superoxide radical scavenging assays) and leucosceptoside A (in ORAC assays). The phenylethanoid fraction may be attractive for various commercial purposes since it displayed significant antioxidant activity and it can be conveniently and economically prepared by a single-step column separation of extracts.

From 18 soil samples taken in the vicinity of the permanent Bulgarian Antarctic base “St. Kliment Ohridski” (62°38′29″S, 60°21′53″W) on Livingston Island, 109 filamentous fungi were isolated on selective media. The most widespread fungal species were members of the genera Cladosporium, Geomyces, Penicillium and Aspergillus. Other species, already recorded in Antarctic environment, were also isolated: Lecanicillium muscarium, Epicoccum nigrum and Alternaria alternata. Thirty strains demonstrating good growth were screened for antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) that play an important role in the defense of aerobic organisms against oxidative stress, by converting reactive oxygen species into nontoxic molecules. Six of them showed high enzyme activity. The tested strains produced SOD with statistically significant higher activity at 15°C than at 30°C suggesting that this enzyme is cold-active. Such SOD could be useful in medicine and cosmetics. The best producer of cold-active SOD, Aspergillus glaucus 363, cultivated in bioreactors, demonstrated optimal growth temperature at 25°C and maximum enzyme activities at 25 and 30°C for SOD and CAT, respectively. The electrophoretical analysis showed that the fungus possesses Cu/Zn-SOD.

The fungal strain Humicola lutea 103 was used as a model organism to examine the relationship between copper toxicity and oxidative stress in low eukaryotes such as filamentous fungi. Spores or submerged cultures were treated with different copper concentrations and the oxidative stress-inducing agent paraquat (PQ). Oxidative stress biomarkers such as reactive oxygen species (ROS), cyanide-resistant respiration, protein carbonyls, reserve carbohydrates, and antioxidant defence were identified in cells treated or not treated with either copper ions or PQ. Copper inhibited the growth and conidiospore formation of H. lutea 103 in a concentration-dependent manner. This treatment also resulted in increased superoxide anion radical formation. Copper stress was furthermore accompanied by transient accumulation of trehalose and glycogen, as well as increased protein carbonyl content. Compared to control cultures, copper-treated mycelia demonstrated a marked increase in the activity of protective enzymes (superoxide dismutase, catalase, and glucose-6-phosphate dehydrogenase). These increased antioxidant enzyme activities were blocked by inhibitors of protein synthesis, suggesting that de novo enzyme formation was involved. Biomarker response to the heavy metal was similar to treatment with known ROS generators such as PQ. The observed hyper-oxidative status and increased oxidative damage suggest a relationship between acute metal treatment and oxidative stress in fungal cells.

Microbiological investigations of cultural heritage may be helpful in the choice of possible intervention for the prevention of biological degradation of works of art. Fungi presented in stone, fretwork and fresco samples from different objects in Rila monastery were isolated and characterized. Pure cultures of 9 strains were obtained and tested for their ability to grow (mycelia mat radial extension) on four agar media at temperature 30°C. The best growth occurred on Sabouraud agar, beer agar and potato glucose agar.Cultivation of four strains in jar bioreactors on Sabouraud medium for assessment of growth phase, glucose consumption and protein synthesis was performed. Extensive mycelia development could even be seen at 12 h of the cultivation starting at a time when glucose was abundant in the culture medium. Maximal biomass was attained at 72 or 84 h of growth. A sharp increase in extracellular protein synthesis was noted in the early stages of growth. The data indicated that all cultures were ...

The present study aims to provide new information about the unusual location of Cu/Zn-superoxide dismutase (Cu/Zn-SOD) in lower eukaryotes such as filamentous fungi. Humicola lutea, a high producer of SOD was used as a model system. Subcellular fractions [cytosol, mitochondrial matrix, and intermembrane space (IMS)] were isolated and tested for purity using activity measurements of typical marker enzymes. Evidence, based on electrophoretic mobility, sensitivity to KCN and H(2)O(2) and immunoblot analysis supports the existence of Cu/Zn-SOD in mitochondrial IMS, and the Mn-SOD in the matrix. Enzyme activity is almost equally partitioned between both the compartments, thus suggesting that the intermembrane space could be one of the major sites of exposure to superoxide anion radicals. The mitochondrial Cu/Zn-SOD was purified and compared with the previously published cytosolic enzyme. They have identical molecular mass, cyanide- and H(2)O(2)-sensitivity, N-terminal amino acid sequence, glycosylation sites and carbohydrate composition. The H. lutea mitochondrial Cu/Zn-SOD is the first identified naturally glycosylated enzyme, isolated from IMS. These findings suggest that the same Cu/Zn-SOD exists in both the mitochondrial IMS and cytosol.

Cu/Zn-containing superoxide dismutase (Cu/Zn-SOD) from the fungal strain Humicola lutea 103 is a naturally glycosylated antioxidant enzyme which has been demonstrated to have a protective effect against myeloid Graffi tumor in hamsters and experimental influenza virus infection in mice. In this work, an improvement of enzyme production was achieved by using both an improved growing strategy and a more efficient purification protocol. The optimized fed-batch cultivation, with 7.5 mg/ml glucose fed daily after 24 h, resulted in prolonged growth and abundant mycelium production, as well as in improvement in enzyme production. A 2-fold increase in the total SOD activity and a significant increase in enzyme production were achieved (from 1.5- to 3.2-fold). Fed-batch technology contributed to a 24 h stabilization period of biosynthesis in which every stop of fermentation can be acceptable, thus making this fermentation a process of industrial interest. In addition, the improved purification procedure offers a reduction of purification steps and enhanced enzyme yield (1.6-fold). The molecular mass was proven to be 15,940 Da for the subunit and the conformational dynamics of the protein in solution was studied by electrospray ionization mass spectrometry (ESI-MS).

WOS: 000228593700009, PubMed ID: 15820508, Recent evidence shows that plant polyphenols exhibit antioxidant and radical scavenging properties. By three separate and complementary methods - DPPH assay, beta-carotene-linoleic acid assay and NBT-reduction assay it was established that a polyphenol-rich extract from the medicinal plant Geranium sanguineum L. with strong anti-influenza virus activity, possessed antioxidant and radical scavenging capacities. For comparative reasons caffeic acid and the synthetic antioxidant BHT were used. Total soluble phenolic constituents of the MeOH extract measured by Folin-Ciocalteu reagent were found as 34.60% (w/w). Further it was demonstrated that the EtOAc fraction, retaining the majority of the in vivo protective effect exhibited a strong O-2(-) scavenging activity while the n-BuOH fraction, containing the majority of the in vitro antiviral activity provoked generation of O-2(-). The O-2(-) scavenging activity of all three preparations correlated with the rate of the protective effect shown in the murine model of experimental influenza virus infection. The present results are in accordance with our intensive studies on the mode of the protective effect of the plant extract which showed positively that the protection may possibly be attributed to the combination of more than one biological activities and that the use of antioxidants might be an useful approach in the treatment of influenza infection. (c) 2005 Elsevier Inc. All rights reserved.

Investigation on the immunoprotective activity of Cu/Zn superoxide dismutase from Humicola lutea 103 AL (HLSOD) in hamsters with transplanted myeloid tumor was performed. Survivability, tumor growth and tumor transplantability were followed. The immune status of tumor-bearing animals, injected with the optimal protective HLSOD dose, was examined during 27 days after tumor transplantation by the following parameters: (i) the number, migration and phagocytic activity of peritoneal macrophages, (ii) the phagocytic activity of blood polymorphonuclear leukocytes (PMNs), (iii) the responsibility in vitro of spleen lymphocytes to T and B cell mitogens. It was established that intraperitoneal inoculation of HLSOD produced a protective effect on the development of tumors. Elongation of the latent time for tumor appearance and inhibition of the tumor growth were observed. The decreased percentage of mortality in early stage of tumor progression was established. Immunological studies on tumor-bearing hamsters (TBH) induced a tem porary immunorestoring effect on the suppressed phagocytic activities of peritoneal macrophages and blood PMNs during the first 14 days of tumor development. Moreover, HLSOD showed an expressed stimulating effect on proliferative activity in vitro of spleen B lymphocytes from healthy and TBH as well. The immunorestoring and protective effect of the enzyme was probably due to improve of the oxidant-antioxidant balance in peritoneal phagocytes. The tem porary character of the effect can be explained with the interference of immunosuppressing factors produced by tumor tissue as well as by the presence of tumor antigens, tumor cells and antigen-antibody complexes in the circulation.

Cultures of the fungal strain Humicola lutea 110 were grown in a 3-l bioreactor. Effects of dissolved oxygen concentration (DO) on cell growth, intracellular protein content and antioxidant enzyme activities (SOD and catalase) were investigated. Controlling DO from 20 to 60% lead to: (I). The lethal phase of growth was reached faster; (ii) strong reduction of the intracellular protein content, and (iii) increase of antioxidant enzyme activities. The most efficient SOD biosynthesis was achieved at the 1st maximum of activity in the culture grown under DO uncontrolled conditions.

A novel thermostable MnSOD was purified to electrophoretic homogeneity from the fungal strain Humicola lutea 110. The preparation of the pure metalloenzyme was performed using treatment with acetone followed by ion exchange and gel permeation chromatography. We found that the activity of this enzyme comprises about 80% of the total superoxide dismutase activity in the crude extract, containing two proteins: MnSOD and Cu/ZnSOD. The MnSOD has a molecular mass of approximately 76 kDa and 7200 U/mg protein specific activity. It is a tetrameric enzyme with four identical subunits of 18 860 Da each as indicated by SDS-PAGE, amino acid analysis and mass spectrometry. N-terminal sequence analysis of MnSOD from the fungal strain revealed a high degree of structural homology with enzymes from other eukaryotic sources. Physicochemical properties were determined by absorption spectroscopy and circular dichroism measurements. The UV absorption spectrum was typical for an MnSOD enzyme, but displayed an increased absorption in the 280 nm region (epsilon280 = 10.4 mM(-1). cm(-1)), attributed to aromatic amino acid residues. The CD data show that MnSOD has two negative Cotton effects at 208 and 222 nm allowing the calculation of its helical content. The ellipticity at 222 nm is 6800 deg. x m(2) x dmol(-1) and thus similar to the values reported for other MnSODs. The MnSOD from H. lutea 110 is stable over a wide range of pH (4.5-8), even in the presence of EDTA. The enzyme is thermostable at 70-75 degrees C, and more stable than MnSODs from other sources.

Conidia of Aspergillus niger 26 spores were immobilised in 3% Ca-alginate beads and some intrinsic kinetic characteristics including growth, polymethylgalacturonase (PMG) production and specific oxygen uptake rates determined. A negative correlation between the specific growth rate and specific PMG production indicated a non-growth-associated enzyme formation. Immobilisation did not change the model of PMG synthesis even in repeated replacement cultures and leads under certain conditions to an enhanced PMG production. The immobilised mycelium was concentrated at the surface layer of the alginate beads and the fungal hyphae could not be found in the central part of the gel. The dense growth of the mycelia on the outer bead surface allowed the formation of a fur-like mycelium coat, which had a large surface for oxygen uptake. This growth behaviour is a reason why the effectiveness factor increased during incubation from 0.54 to 0.93.

Humicola lutea 120-5 spores were immobilized in a mixed photo-crosslinked polyvinyl alcohol and calcium-alginate gel. Maximum enzyme synthesis was established with 1:8 (v:v) gel beads: growth medium inoculum and 48 h duration of one cycle. The free cells were very unstable in replacement fermentations. The operational stability of the immobilized system indicated the possibility of the application of Humicola lutea 120-5 in a semi-continuous process for the production of acid proteinase.

The study focuses on the morphological and physiological cell responses to oxidative stress induced by high temperature treatment in the industrially relevant fungus Aspergillus niger 26. Temperatures above 30 °C lead to growth suppression and changes in morphological characteristics: decrease in the size of hyphal elements and increase in “active length” by switching from slightly branched long filaments to a multitude of branched forms containing active cytoplasm. Transmission electron microscopy of fungal cultures heated at 40 °C demonstrated abnormal wavy septation with reduced amount of chitin (as shown by WGA-gold labelling), intrahyphal hyphae development, disintegration of mitochondria and extensive autolysis. Temperature-dependent decrease in the total intracellular protein content and a sharp increase (six to tenfold) in oxidatively damaged proteins were also demonstrated. Elevated temperatures caused a two and threefold increase in catalase and superoxide dismutase activities, respectively.

Pentachlorophenol (PCP) caused a rapid and pronounced increase in the rate of biosynthesis of the superoxide dismutase (SOD) in fungal strain Humicola lutea 110. Mn-containing SOD (Mn-SOD) was mainly responsible for modulating total cell SOD. The kinetics of SOD synthesis in the presence of PCP demonstrated the induction model of enzyme formation. This model was also supported by deinduction experiments, because the removal of the PCP was followed by a marked decrease in SOD activity. PCP also caused a moderate induction of catalase. The concentrations, which were effective in inducing the Mn-SOD, increased the cyanide-resistant respiration. It seems likely that PCP increased the rate of intracellular production of superoxide [Formula: see text]. Addition of inhibitors of transcription and translation to cultures in the presence of inducer (PCP) inhibited further accumulation of SOD activity. These data suggest that PCP, probably by the increase of [Formula: see text] content, accelerates new enzyme synthesis in fungal strain Humicola lutea 110.Key words: superoxide dismutase, superoxide, induction, pentachlorophenol, fungi, Humicola lutea.

Superoxide dismutase (SOD) was produced by the fungal strain Humicola lutea 110. Some of the factors influencing both enzyme and biomass production were studied in shake-flask cultures: glucose and nitrogen concentration, inoculum quantily, and temperature. The best results were obtained for a fermentation performed with 4% glucose, 0.152% nitrogen source, and 8% inoculum at 30°C. The time course of SOD biosynthesis showed two maxima (84.1 and 120.8 U/mg protein SOD activity respectively), which correspond to the maxima of the biomass. The dramatic increase in SOD activity during the late stationary stage indicated that the processes of O2− generation were intensified in the cells. Mn-SOD was responsible for the modulation of total SOD activity at both maxima.

The superoxide dismutase (SOD) is the first enzyme to respond against oxygen radicals and to offer the greatest response to oxidative stress. The great interest in SODs is a result of their important physiological role and therapeutic potential. The fungal strain Humicola lutea 103 has been selected as an effective producer of Cu/Zn-SOD exhibiting protective effect against influenza virus infection and myeloid Graffi tumour. The present paper s the role of control mechanisms induction and catabolite repression on SOD synthesis under copper stress conditions. Cu ions cause rapid and profound induction of enzyme activity by dose- and time-dependent manner. The deinduction experiments also demonstrate the induction model of SOD synthesis. The results suggest that SOD synthesis in the fungal strain H. lutea 103 treated with Cu ions is a subject to catabolite repression.

The effect of glucose on superoxide dismutase (SOD) activity produced in Humicola lutea 110 was determined. The relatively high glucose concentration in the medium did not repress SOD levels in the cells. Glycerol, a nonfermentable carbon source, caused a slight stimulation of SOD synthesis. Furthermore, the specific rates of enzyme production in the medium with different glucose concentration showed an insignificant difference. Cyclic AMP had no effect on SOD levels. The shift in metabolism as glucose was depleted resulted in an increase in the rate of synthesis of both isocitric dehydrogenase and SOD. Pentachlorophenol and paraquat, which cause the production of superoxide radicals, caused an increase in SOD activity. These results led us to conclude that it is superoxide ion rather than glucose that is controlling SOD levels.Key words: Humicola lutea, fungi, superoxide dismutase, intracellular enzyme, glucose effect.

Spores of fungal strain Humicola lutea 120-5 were immobilized with photo-crosslinkable prepolymer. The entrapped spores were allowed to germinate and develop in situ. The immobilized growing mycelium thus obtained was reused in batch mode for extracellular acid proteinase production. The influence of prepolymer concentration, initial inoculum gel volume, spore content, and duration of the growth cycle on the enzyme activity and mycelium growth were studied. The best yield of the acid proteinase was reached with 15 ml initial gel volume, 20% prepolymer concentration, 10 6 spores per ml gel and 56 h duration of one cycle. The optimization of these factors, which affect the stability and productivity of the immobilized system, resulted in the increase of the enzyme activity (up to 180% compared with free cells), decrease of concentration of outgrowing cells in the medium, and the realization of semicontinuous cultivation. The productive life was nearly 10–12-times longer, and the cumulative enzyme activity was 18-times higher in comparison with those of free culture. Therefore, the immobilized growing Humicola lutea 120−5 in the photo-crosslinked polymer could be used for a long-term production of an acid proteinase.

РЕЗЮМЕИсследовано влияние аргонового лазера на приживаемость, биосинтетические возможности, морфологию и стабильность алкалоидной продукции штамма Claviceps sp. Облучение не вызывает каких -либо сушественных изменений в морфологии. При воздействии аргоновым лазером не наблюдались различия в составе смеси алкалоидов, но обнаружены изменения в соотношении отдельных компонентов. По сравнению с родительским штаммом облучение способствует повышению биосинтеза алкалоида в 2.5 раза.

To extend the knowledge about the relationship between heat shock and oxidative stress in lower eukaryotes, the filamentous fungus Aspergillus niger 26 was chosen as a model system. Here, the response of A. niger cells to heat shock is reported. The temperature treatment significantly increased the levels of reactive oxygen species, superoxide anions (O2•–), and hydrogen peroxide and the rate of cyanide-resistant respiration as a marker of oxidative stress. Enhanced reactive oxygen species generation coincided with an increase in the content of oxidative damaged protein and in the accumulation of the storage carbohydrates trehalose and glycogen. Thermal survival of the A. niger cells corresponded to a significant increase in the levels of the antioxidant enzymes superoxide dismutase and catalase for all variants. These observations suggest that heat and oxidative stress have a common cellular effect.

This study was conducted to evaluate the radical scavenging capacities of extracts and preparations from a Lavandula vera MM plant cell culture with different rosmarinic acid content and to compare them with pure rosmarinic and caffeic acids as well. The methods, which were used are superoxide anion and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt radicals scavenging assays. Results showed that extracts and preparations from Lavandula vera MM possess strong radical scavengers, as the best both radical scavengers appeared to be the fractions with enriched rosmarinic acid content, obtained after ethylacetate fractioning (47.7% inhibition of superoxide radicals and 14.2 μᴍ 6-hydroxy- 2,5,7,8-tetramethylchroman-2-carboxylic acid equivalents, respectively). These data reveal the possibilities for application of these preparations as antioxidants.

Although, oxidative stress response, which protects organisms from deleterious effects of reactive oxygen species (ROS), has been extensively studied in pro- and eukaryotes, the information about filamentous fungi is fragmentary. We investigated the effect of two ROS-generating agents (paraquat, PQ, and H2O2) on cellular growth and antioxidant enzyme induction in 12 fungal species. Our results indicate that exposure of fungal spores or mycelia to PQ and H2O2 promoted oxidative stress, as evidenced by remarkable inhibition of spore germination and biomass production; stimulation of cyanide-resistant respiration; accumulation of oxidative modified proteins. Cell responses against both superoxide and peroxide stresses include enhanced expression of superoxide dismutase (SOD) and catalase, however, the extent was different: treatment with PQ increased mainly SOD, whereas exogenous H2O2 led to enhanced catalase. We also found that G6PD has a role in the mechanism of protection against superoxide and peroxide stresses. The activation of antioxidant enzyme defence was blocked by the translation inhibitor, cycloheximide, suggesting that there was de novo enzyme synthesis.

A novel Cu/Zn-containing superoxide dismutase (SOD) was isolated from the fungal strain Humicola lutea 103. Previously, a protective effect of this enzyme (HLSOD) against tumor growth and also superoxide production in Graffi tumor-bearing hamsters (TBH) were established. The aim of the present study was to investigate the effect of HLSOD on the activity of endogenous SOD and catalase in the cells from TBH during tumor progression. Our results point out that transplantation of Graffi tumor causes a significant decrease in SOD activity in the cells from liver of the hosts (from 35 to 59% compared to the control). In the tumor cells relatively low levels of SOD (about 7 U mg protein-1) were found, and Cu/ZnSOD was the main isoenzyme in total SOD activity. Tumor growth resulted in a reduction of catalase activity, which correlated with the process of tumor progression. A single dose (65 U) treatment with HLSOD caused an increase in endogenous SOD and catalase activity in healthy animals and resulted in restoration of the antioxidant ability in liver cells of the hosts at the early stage of tumor progression. The results show the possible participation of HLSOD in the host oxidant-antioxidant balance, which is probably one of the factors of its immunoprotective action established earlier.

The fungal strain Humicola lutea 103 produces a naturally glycosylated Cu/Zn SOD. To improve its yield, the effect of an increased concentration of dissolved oxygen (DO) on growth and enzyme biosynthesis by the producer, cultivated in a 3 l bioreactor, was examined. Exposure to a 20% DO level caused a 1.7-fold increase of SOD activity compared to the DO-uncontrolled culture. Maximum enzyme productivity of SOD was approximately 300 x 10(3) U (kg wet biomass)(-1). The novel enzyme was purified to electrophoretic homogeneity. The presence of Cu and Zn were confirmed by atomic absorption spectrometry. The molecular mass of H. lutea Cu/Zn SOD was calculated to be 31870 Da for the whole molecule and 15936 Da for the structural subunits. The N-terminal sequence revealed a high degree of structural homology with Cu/Zn SOD from other prokaryotic and eukaryotic sources. H. lutea Cu/Zn SOD was used in an in vivo model for the demonstration of its protective effect against myeloid Graffi tumour in hamsters. Comparative studies revealed that the enzyme (i) elongated the latent time for tumour appearance, (ii) inhibited tumour growth in the early stage of tumour progression (73-75% at day 10) and (iii) increased the mean survival time of Graffi-tumour-bearing hamsters. Moreover, the fungal Cu/Zn SOD exhibited a strong protective effect on experimental influenza virus infection in mice. The survival rate increased markedly, the time of survival rose by 5.2 d and the protective index reached 86%. The H. lutea SOD protected mice from mortality more efficiently compared to the selective antiviral drug ribavirin and to commercial bovine SOD. In conclusion, our results suggest that appropriate use of the novel fungal SOD, applied as such or in combination with selective inhibitors, could outline a promising strategy for the treatment of myeloid Graffi tumour and influenza virus infection.

Effect of immobilization on antioxidant enzyme synthesis by growing and non-growing cell culture of Aspergillus niger 26 was studied. Entrapped cells showed a greater than 1.5-fold increase in the superoxide dismutase (SOD) activity and a moderate elevation in catalase activity. The immobilization did not cause changes in the spectrum of SOD isoenzymes. The observed increase in SOD activity required de novo synthesis of this enzyme, because it was suppressed by inhibitors of the transcription and translation. The addition of various viscous substances (agar, Na-alginate and pectin) stimulated the SOD synthesis. Despite these results, it was found that the changes in SOD activity are induced in response to growth in the state of immobilization rather than to presence of alginate. Immobilized A. niger cells exhibited about a 4- to 5-fold higher level of cyanide-resistant respiration. This latter phenomenon might use as an indicator of intracellular oxy-intermediate generation in cell culture growing under stress conditions. The results are discussed relative to association between physiological stress caused by immobilization and oxidative stress.