Wheat straw cultures of the brown rot fungi Gloeophyllum striatum and G. trabeum degraded 2,4-dichlorophenol and pentachorophenol. Up to 54% and 27% 14CO2, respectively, were liberated from uniformly 14C-labeled substrates within 6 weeks. Under identical conditions Trametes versicolor, a typical white rot species employed as reference, evolved up to 42% and 43% 14CO2 and expressed high activities of laccase, manganese peroxidase, and manganese-independent peroxidase. No such activity could be detected in straw or liquid cultures of Gloeophyllum. Moreover, G. striatum degraded both chlorophenols most efficiently under non-cometabolic conditions, i.e. on a defined mineral medium lacking sources of carbon, nitrogen and phosphate.

Iron-containing liquid cultures of the brown-rot basidiomycete Gloeophyllum striatum degraded 2-fluorophenol. Two simultaneously appearing degradation products, 3-fluorocatechol and catechol, were identified by gas chromatography and mass spectrometry (GC-MS). Concomitantly, fluoride was produced at approximately 50% of the amount that theoretically could be achieved upon complete dehalogenation. Defluorination was strongly inhibited in the presence of either the hydroxyl radical scavenger mannitol or superoxide dismutase, as well as in the absence of iron. The addition of the natural iron chelator oxalate caused a clear but less extensive inhibition, whereas supplementation with the artificial iron chelator nitrilotriacetic acid increased fluoride production. Extracellular 2-fluorophenol degradation was evidenced by defluorination, observed upon addition of 2-fluorophenol to cell-free culture supernatants derived from iron-containing fungal cultures. Ultrafiltered culture supernatants oxidized methanol to formaldehyde, known as a product of the reaction of methanol with hydroxyl radical. In addition, G. striatum was found to produce metabolites extractable with ethyl acetate that are capable of reducing Fe3+. GC-MS analysis of such extracts revealed the presence of several compounds. The mass spectrum of a prominent peak matched those previously reported for 2,5-dimethoxyhydroquinone and 4,5-dimethoxycatechol, fungal metabolites implicated to drive hydroxyl radical production in Gloeophyllum. Taken together, these findings further support an extracellular Fenton-type mechanism operative during halophenol degradation by G. striatum.

While degrading 2,4-dichlorophenol, two strains of Gloeophyllum striatum, a basidiomycetous fungus causing brown rot decay of wood, simultaneously produced 4-chlorocatechol and 3,5-dichlorocatechol. These metabolites were identified by comparing high-performance liquid chromatography retention times and mass spectral data with those of chemically synthesized standards. Under similar conditions, 3-hydroxyphthalic hydrazide was generated from phthalic hydrazide, a reaction assumed to indicate hydroxyl radical formation. Accordingly, during chemical degradation of 2,4-dichlorophenol by Fenton's reagent, identical metabolites were formed. Both activities, the conversion of 2,4-[U-14 C]dichlorophenol into 14 CO 2 and the generation of 3-hydroxyphthalic hydrazide, were strongly inhibited by the hydroxyl radical scavenger mannitol and in the absence of iron. These results provide new evidence in favor of a Fenton-type degradation mechanism operative in Gloeophyllum.Basidiomycetous fungi causing brown rot decay of wood play an important role in the carbon cycle. They depolymerize the cellulose component of wood while lignin remains as an amorphous brown residue. In this process, an extracellular Fentontype mechanism, providing hydroxyl radicals via Fe 2ϩ and hydrogen peroxide, has long been implicated (15). However, the potential of species like Gloeophyllum striatum and Gloeophyllum trabeum in the degradation of xenobiotics was explored only recently: two fluoroquinolone antibacterial drugs, polyethylene glycol, and two chlorophenols have been shown to be decomposed (9, 14,23,24). G. striatum produced three remarkable hydroxylated congeners of enrofloxacin, which could also be generated via Fenton's reaction (23). An extracellular 2,5-dimethoxyhydroquinone-driven Fenton reaction in G. trabeum, employed to depolymerize polyethylene glycol, represents the most recent mechanistic evidence (14).Wheat straw cultures of G. striatum catalyzed CO 2 production from the environmental pollutant 2,4-dichlorophenol (2,4-DCP). Moreover, in a defined mineral medium lacking carbon, nitrogen, and phosphate, G. striatum expressed a degradation capacity similar to that shown on wheat straw (9, 23). Here, our aim was to provide further evidence for a Fenton-type reaction in G. striatum by (i) identifying primary metabolites of 2,4-DCP formed by the fungus and in Fenton's reaction as well; (ii) testing for hydroxylation of phthalic hydrazide (PH) giving 3-hydroxyphthalic hydrazide (3-OHPH), which has been postulated to specifically indicate hydroxyl radical formation (1, 2, 19); and (iii) inhibiting the conversion of 2,4-[U-14 C]DCP to 14 CO 2 as well as hydroxylation of PH under conditions antagonizing hydroxyl radical activity. Such data are intended to help to firmly establish the capability of some higher fungi to employ a Fenton-type reaction in the degradation of xenobiotics. This reaction may even take different forms in the few genera investigated so far (13, 14). MATERIALS AND METHODSOrganisms. The source and maintenance of G. str...

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A photocleavable o-nitrobenzyl CE phosphoramidite building-block was synthesised and incorporated within oligonucleotides. After allele-specific primer extension, desalting was performed using genostrep purification plates. Release of the SNP information containing part through photocleavage created shortened molecules that are easily accessible for MALDI-TOF analysis. Additionally, incorporation of mass modified nucleosides enables flexible design of multiplex genotyping.