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The basidiomycetous fungus Nematoloma frowardii produced manganese peroxidase (MnP) as the predominant ligninolytic enzyme during solid-state fermentation (SSF) of wheat straw. The purified enzyme had a molecular mass of 50 kDa and an isoelectric point of 3.2. In addition to MnP, low levels of laccase and lignin peroxidase were detected. Synthetic 14 C-ring-labelled lignin ( 14 C-DHP) was efficiently degraded during SSF. Approximately 75% of the initial radioactivity was released as 14 CO 2 , while only 6% was associated with the residual straw material, including the well-developed fungal biomass. On the basis of this finding we concluded that at least partial extracellular mineralization of lignin may have occurred. This conclusion was supported by the fact that we detected high levels of organic acids in the fermented straw (the maximum concentrations in the water phases of the straw cultures were 45 mM malate, 3.5 mM fumarate, and 10 mM oxalate), which rendered MnP effective and therefore made partial direct mineralization of lignin possible. Experiments performed in a cell-free system, which simulated the conditions in the straw cultures, revealed that MnP in fact converted part of the 14 C-DHP to 14 CO 2 (which accounted for up to 8% of the initial radioactivity added) and 14 C-labelled water-soluble products (which accounted for 43% of the initial radioactivity) in the presence of natural levels of organic acids (30 mM malate, 5 mM fumarate).

Crude and purified manganese peroxidase from the white-rot fungi Nematoloma frowardii and Phlebia radiata catalyzed the partial depolymerization of a [14C-ring]labelled synthetic lignin into water-soluble fragments (30–50%). The in vitro depolymerization of the 14C-labelled lignin was accompanied by a release of 14CO2 ranging from 4 to 6%. Small quantities of the thiol mediator glutathione stimulated the depolymerization of lignin resulting in a mineralization and solubilization of up to 10 and 64%, respectively. Most of the water-soluble substances formed had molecular masses around 0.7 kDa, although a higher-molecular mass fraction was also detectable (>2 kDa). Photometric assays using 2,2′-azinobis(3-ethylbenzothiazolinesulphonate) as an indicator demonstrated that high levels of Mn(III), which were very probably responsible for the depolymerization and mineralization of the 14C-labelled lignin, were adjusted within the first 24 h of incubation. The manganese peroxidase catalyzed depolymerization process was not necessarily dependent on H2O2; also in the absence of the H2O2-generating system glucose/glucose oxidase, effective solubilization and mineralization of lignin dehydrogenation polymerizate occurred, due to the in part superoxide dismutase sensitive, ‘oxidase-like’ activity of MnP which probably produces radical species and peroxides from malonate.

The degradation of the nitroaromatic pollutant 2,4,6-trinitrotoluene (TNT) by the manganese-dependent peroxidase (MnP) of the white-rot fungus Phlebia radiata and the main reduction products formed were investigated. In the presence of small amounts of reduced glutathione (10 mM), a concentrated cell-free preparation of MnP from P. radiata exhibiting an activity of 36 nkat/ml (36 nmol Mn(II) oxidized per sec and per ml) transformed 10 mg/l of TNT within three days. The same preparation was capable of completely transforming the reduced derivatives of TNT. When present at 10 mg/l, the aminodinitrotoluenes were transformed in less than two days and the diaminonitrotoluenes in less than three hours. Experiments with 14C-U-ring labeled TNT and 2-amino-4,6-dinitrotoluene showed that these compounds were mineralized by 22% and 76%, respectively, within 5 days. Higher concentrations of reduced glutathione (50 mM) led to a severe inhibition of the degradation process. It is concluded that Phlebia radiata is a good candidate for the biodegradation of TNT as well as its reduction metabolites.

A manganese peroxidase preparation from the white-rot fungus Nematoloma frowardii was found to be capable of releasing up to 17% 14CO2 from 14C-labelled synthetic humic substances. The latter were prepared from [U-14C]catechol by spontaneous oxidative polymerization or laccase-catalysed polymerization. The ex-tent of humic substance mineralization was considerably enhanced in the presence of the thiol mediator glutathione (up to 50%). Besides the evolution of 14CO2, the treatment of humic substances with Mn peroxidase resulted in the formation of lower-molecular-mass products. Analysis of residual radioactivity by gel-permeation chromatography demonstrated that the predominant molecular masses of the initial humic substances ranged between 2 kDa and 6 kDa; after treatment with Mn peroxidase, they were reduced to 0.5–2 kDa. The extracellular depolymerization and mineralization of humic substances by the Mn peroxidase system may play an important role in humus turnover of habitats that are rich in basidiomycetous fungi.

Preparations of extracellular manganese peroxidase from the white-rot fungus Nematoloma frowardii and the litter decaying fungus Stropharia rugosoannulata converted rapidly the main intermediates of the explosive 2,4,-trinitrotoluene--the aminonitrotoluenes. In a cell-free system, 2-amino-4,6-dinitrotoluene, 4-amino-2,6-dinitrotoluene and 2,6-diamino-4-nitrotoluene were degraded without formation of identifiable metabolites. Radioactive experiments using a complex mixture of uniform ring-labeled 14C-TNT reduction products demonstrated the partial direct mineralization of these compounds by manganese peroxidase. The extent of aminonitrotoluene conversion as well as the release of 14CO2 from TNT reduction products were considerably enhanced in the presence of thiols like reduced glutathione or the amino acid L-cystein, which probably act as secondary mediators.

The main manganese peroxidase isoenzyme MnP2 of the South American white-rot fungus Nematoloma frowardii b19 was purified to homogeneity using anion-exchange chromatography (Mono Q) and preparative isoelectric focusing. The purified enzyme has a molecular mass of 44 kDa and a pI of 3.2.

A manganese peroxidase preparation of the white-rot fungus Nematoloma frowardii mineralized uniformly ring-labeled C-2-amino-4,6-dinitrotoluene directly to CO . In the cell-free system, a release of 52% CO was demonstrated in the presence of reduced glutathione as secondary thiol mediator. This is the first description of the direct enzymatic combustion of an aromatic compound by an extracellular enzyme of the ligninolytic system.

The Penicillium strain Bi 7/2 utilized phenol, catechol, resorcinol, hydroquinone, pyrogallol, hydroxyhydroquinone, phloroglucinol, m- and p-cresol, orcinol, 4-methylcatechol. 4-methoxy-phenol, 4-aminophenol, benzyl alcohol, benzoic acid, 2-, 3- and 4-hydroxybenzoic acid, anthranilic acid, protocatechuic acid and gallic acid as sole sources of carbon and energy. The central metabolites catechol, protocatechuic acid and hydroxyquinone could be determined by HPLC with diode-array detection. Pathways for the degradation of aromatic substances were proposed.

Summary Anthers of Digitalis lanata were cultivated on a modified Murashige-Skoog medium with 25 pmol NAA, 1 µmol kinetin, and 0.6 mol sucrose L-1. Up to 33 % of the anthers formed calli, most of which contained haploid cells. If the calli were grown on a nutrient medium with 5µmol 2,4D and 1µmol kinetin L-1 the haploid cells disappeared rapidly. In suspension cultures derived from the callus, haploid cells were detectable for longer periods of time, and suspension cultures containing haploid cells were also obtained from apparently diploid callus. Diploid plants were regenerated from callus via somatic embryos by successive cultivation on a nutrient medium with 0.05 µmol NAA and 5 µmol BA L-1, and a nutrient medium without growth regulators. The cardenolide contents of the regenerated plants showed greater variability than those of the clone plants they were derived from.

Isotope enrichment of some elements is required in support of the Rare Isotope Accelerator (RIA) in order to obtain the beam intensities, source efficiencies and/or source lifetime required by RIA. The economics of using Atomic Vapor Laser Isotope Separation (AVLIS) technology as well as ElectroMagnetic (EM) separation technology has been evaluated. It is concluded that such an AVLIS would be about 10 times less expensive than a facility based on electromagnetic separation - $17 M versus $170 M. In addition, the AVLIS facility footprint would be about 10 times smaller, and operations would require about 4 years (including 2 years of startup) versus about 11 years for an EM facility.

The direct involvement of manganese peroxidase (MnP) in the mineralization of natural and xenobiotic compounds was evaluated. A broad spectrum of aromatic substances were partially mineralized by the MnP system of the white rot fungus Nematoloma frowardii . The cell-free MnP system partially converted several aromatic compounds, including [U- 14 C]pentachlorophenol ([U- 14 C]PCP), [U- 14 C]catechol, [U- 14 C]tyrosine, [U- 14 C]tryptophan, [4,5,9,10- 14 C]pyrene, and [ring U- 14 C]2-amino-4,6-dinitrotoluene ([ 14 C]2-AmDNT), to 14 CO 2 . Mineralization was dependent on the ratio of MnP activity to concentration of reduced glutathione (thiol-mediated oxidation), a finding which was demonstrated by using [ 14 C]2-AmDNT as an example. At [ 14 C]2-AmDNT concentrations ranging from 2 to 120 μM, the amount of released 14 CO 2 was directly proportional to the concentration of [ 14 C]2-AmDNT. The formation of highly polar products was also observed with [ 14 C]2-AmDNT and [U- 14 C]PCP; these products were probably low-molecular-weight carboxylic acids. Among the aliphatic compounds tested, glyoxalate was mineralized to the greatest extent. Eighty-six percent of the 14 COOH-glyoxalate and 9% of the 14 CHO-glyoxalate were converted to 14 CO 2 , indicating that decarboxylation reactions may be the final step in MnP-catalyzed mineralization. The extracellular enzymatic combustion catalyzed by MnP could represent an important pathway for the formation of carbon dioxide from recalcitrant xenobiotic compounds and may also have general significance in the overall biodegradation of resistant natural macromolecules, such as lignins and humic substances.

Within a screening program, 91 fungal strains belonging to 32 genera of different ecological and taxonomic groups (wood- and litter-decaying basidiomycetes, saprophytic micromycetes) were tested for their ability to metabolize and mineralize 2,4,6-trinitrotoluene (TNT). All these strains metabolized TNT rapidly by forming monoaminodinitrotoluenes (AmDNT). Micromycetes produced higher amounts of AmDNT than did wood- and litter-decaying basidiomycetes. A significant mineralization of [14C]TNT was only observed for certain wood- and litter-decaying basidiomycetes. The most active strains, Clitocybula dusenii TMb12 and Stropharia rugosa-annulata DSM11372 mineralized 42% and 36% respectively of the initial added [14C]TNT (100 microM corresponding to 4.75 microCi/l) to 14CO2 within 64 days. Micromycetes (deuteromycetes, ascomycetes, zygomycetes) proved to be unable to mineralize [14C]TNT significantly.