Biotransformation of fluorene by the fungus Cunninghamella elegans.

The metabolism of fluorene, a tricyclic aromatic hydrocarbon, by Cunninghamella elegans ATCC 36112 was investigated. Approximately 69% of the [9-14C]fluorene added to cultures was metabolized within 120 h. The major ethyl acetate-soluble metabolites were 9-fluorenone (62%), 9-fluorenol, and 2-hydroxy-9-fluorenone (together, 7.0%). Similarly to bacteria, C. elegans oxidized fluorene at the C-9 position of the five-member ring to form an alcohol and the corresponding ketone. In addition, C. elegans produced the novel metabolite 2-hydroxy-9-fluorenone.     

Nitrate-nonutilizing mutants and vegetative self-incompatibility in Cunninghamella echinulata var. elegans

Cunninghamella echinulata var. elegans is a model system for the study of metabolism of toxic compounds. Unlike most other zygomycete fungi, Cunninghamella can use nitrate as a sole N source. We generated nitrate-nonutilizing (Nit-) mutants in three isolates of C. echinulata var. elegans. Mutants at the niaD and nirA- loci were unable to complement each other, suggesting that this fungus is vegetatively self-incompatible. Vegetative self-incompatibility may prove advantageous for using Cunninghamella for in situ bioremediation. © Rapid Science Ltd. 1998     

Biochemical markers in taxonomy of the genus Cunninghamella

The chemical composition of fatty acids and ubiquinones was studied in 18 strains of Cunninghamella, to establish quantitative and qualitative differences within the genus. Fatty acids analysis has shown the presence of four groups. Ubiquinone analysis, through high performance liquid chromatography (HPLC), demonstrated the existence of three different groups based on the ubiquinone type. The average percentage of fatty acids of the species Cunninghamella elegans and Cunninghamella bertholletiae, show variations in linolenic and stearic acids, suggesting the possibility of differentiation between the two species.     

Microbial hydroxylation/functionalization of terpenoid synthons derived from communic acids

Incubation of a communic acid-derived synthon with Cunninghamella elegans quantitatively affords 1 beta, 3 beta- and 7 beta- monohydroxylated derivatives.     

Stereospecific microbiological 10-O-demethylation of R-(-)-10,11-dimethoxyaporphines

The microbiological O-dealkylation of (+) and (-) 10,11-dimethoxyaporphine and the corresponding N-n-propyl analog 10,11-dimethoxy-N-n-propylnoraporphine utilizing the fungus Cunninghamella elegans (ATCC 9245) was found to proceed with regioselectivity for the 10-position, and with a high degree of substrate stereospecificity for the 6a R(-)enantiomer. Only the (R) 10-demethylated products were isolated i.e. (R) iosapocodeine and (R) N-n-propylnor-isoapocodeine. The products were confirmed by comparison with their GC/MS spectra.     

Regiospecific synthesis of isoapocodeine from 10,11-dimethoxyaporphine by using Cunninghamella elegans.

A preparative-scale regiospecific conversion of 10,11-dimethoxyaporphine to isoapocodeine was conducted with Cunninghamella elegans ATCC 9245. This biotransformation proceeded quantitatively in suspensions and was pH dependent. The influence of antioxidants on the conversion was studied. Attempts to preserve the activity of isolated C. elegans cells by a number of methods were unsuccessful.     

Regiospecific synthesis of isoapocodeine from 10,11-dimethoxyaporphine by using Cunninghamella elegans.

A preparative-scale regiospecific conversion of 10,11-dimethoxyaporphine to isoapocodeine was conducted with Cunninghamella elegans ATCC 9245. This biotransformation proceeded quantitatively in suspensions and was pH dependent. The influence of antioxidants on the conversion was studied. Attempts to preserve the activity of isolated C. elegans cells by a number of methods were unsuccessful.     

Screening and evaluation of fungal resources for loratadine metabolites

Loratadine is a selective inverse agonist of peripheral histamine H1-receptors. Microbial biotransformation gained a lot of attention for its ability to convert molecules to valuable medicinally active substances. The main objective of the present research was to investigate the ability of different fungi to biotransform the drug loratadine to its active metabolite desloratadine, because desloratadine is four times more potent, possess longer duration of action than loratadine and is effective at low doses. The screening studies were performed with selected fungi using their respective broth media and sterile incubation conditions. The drug and metabolites formed (if any) were extracted and analysed using HPLC analysis. Structural elucidation and confirmation of metabolites were by mass and proton NMR spectroscopy. Among the six fungi selected, Cunninghamella elegans, Cunninghamella echinulata and Aspergillus niger cultures showed extra peaks at 3.8, 3.6 and 4.1 min, respectively, in HPLC when compared with their controls, which indicated the formation of metabolites. The metabolites thus formed were isolated and their structures were confirmed as dihydroxy desloratadine, desethoxy loratadine and 3-hydroxy desloratadine by Cunninghamella elegans, Cunninghamella echinulata and Aspergillus niger cultures, respectively, by mass spectrometry and NMR spectroscopy. Three fungi were identified to have the ability to biotransform loratadine to its active metabolite and other different metabolites.     

Metabolism of naphthalene by Cunninghamella elegans.

Cunninghamella elegans grown on Sabouraud dextrose broth in the presence of naphthalene produced six metabolites. Each product was isolated and identified by conventional chemical techniques. The major metabolites were 1-naphthol (67.9%) and 4-hydroxy-1-tetralone (16.7%). Minor products isolated were 1,4-naphthoquinone (2.8%), 1,2-naphthoquinone (0.2%), 2-naphthol (6.3%), and trans-1,2-dihydroxy-1,2-dihydronaphthalene (5.3%). C. elegans oxidized both 1-naphthol and 1,4-naphthoquinone to 4-hydroxy-1-tetralone. The results suggest that C. elegans oxidizes naphthalene by a sequence of reactions similar to those reported for the mammalian metabolism of this hydrocarbon.     

Fungal metabolites of (E)-guggulsterone and their antibacterial and radical-scavenging activities

Fungal transformation of (E)-guggulsterone (= (17E)-pregna-4,17-diene-3,16-dione; 1) by Rhizopus stolonifer, Fusarium lini, Cunninghamella elegans, or Gibberella fujikuroi afforded ten hydroxylated metabolites (2-11; Scheme), which were fully characterized. Compounds 4-11 have not been described yet. Some of these novel hydroxylated metabolites, as well as acetylated derivatives thereof, exhibited significant antibacterial and radical-scavenging activities (Table 3).     

Biotoxic activity in the Mucorales

The toxigenicity of representatives of 15 species of Mucorales (Absidia glauca, Actinomucor elegans, Cunninghamella elegans, Helicostylum piriforme, Mortierella isabellina, Mortierella (Mucor) rammaniana, Mucor hiemalis, Mucor mucedo, Mucor spinosus, Phycomyces blakesleeanus, Rhizopus oligosporus, Rhizopus stolonifer, Syncephalastrum racemosum, Thamnidium elegans, Zygorhynchus moelleri) towards the larvae of brine shrimp (Artemia salina) and the growth of pea seedlings (Pisum sativum) and tobacco plants (Nicotiana tabacum) was evaluated. The fungi were cultivated on malt extract agar and aqueous solutions of the cultures were tested.Thamnidium elegans showed a marked toxic action towards brine shrimp (mortality: 74.1%) andPhycomyces, Actinomucor andSyncephalastrum were only weakly toxic. Length and weight of stems of pea seedlings were moderately reduced by extracts ofAbsidia, Cunninghamella, Zygorhynchus andThamnidium and to a lesser degree byMucor spinosus. Cunninghamella andMucor spinosus also inhibited the development of pea hypocotyls. The length of tobacco stems was reduced byMortierella ramanniana, Rhizopus stolonifer andCunninghamella elegans. Wilting or other toxic phenomena were never observed with both test plants. Considering the present results and data from literature it is suggested that species of Mucorales have only a weak toxigenicity.     

Microbial transformations of natural antitumor agents. 7. 14-alpha-Hydroxylation of withaferin-A by Cunninghamella elegans (NRRL 1393)

Microbial transformation experiments were conducted with the steroidlactone, withaferin-A (1a). Cunninghamella elegans (NRRL 1393) converts withaferin-A into two major metabolites, one of which has been indentified as 14alpha-hydroxywithaferin-A (1b). The metabolite is obtained in 37% yield, and its structure was determined on the basis of pmr and mass spectral evidence. The metabolite has the same level of antitumor activity as withaferin-A against the Sarcoma-180 tumor test system in mice.     

Biodegradation of aspen steryl esters and waxes by two filamentous fungi with or without other carbon sources

Aspergillus luchuensis and Cunninghamella elegans were evaluated for their growth on steryl esters and waxes, which are a major cause of pitch deposition in aspen pulping. These fungi hydrolysed aspen steryl esters and waxes into their constituent sterol and fatty acid moieties. A. luchuensis and C. elegans were also grown on steryl esters and waxes supplemented with glucose and triglycerides in order to provide a more accurate assessment of how these fungi behave on aspen wood. Both fungi consumed glucose before steryl esters and waxes while they degraded the triglycerides and steryl esters and waxes simultaneously.     

Identification of a novel metabolite in phenanthrene metabolism by the fungus Cunninghamella elegans

The metabolism of phenanthrene by the fungus Cunninghamella elegans was investigated. Kinetic experiments using [9-14C]phenanthrene showed that after 72 h, 53% of the total radioactivity was associated with a glucoside conjugate of 1-hydroxyphenanthrene (phenanthrene 1-O-beta-glucose). This metabolite was isolated by reversed-phase high-performance liquid chromatography and characterized by the application of UV absorption, 1H nuclear magnetic resonance, and mass spectral techniques. The results show that aromatic ring oxidation followed by glucosylation is a predominant pathway in the metabolism of the polycyclic aromatic hydrocarbon phenanthrene by C. elegans.     

Identification of a novel metabolite in phenanthrene metabolism by the fungus Cunninghamella elegans.

The metabolism of phenanthrene by the fungus Cunninghamella elegans was investigated. Kinetic experiments using [9-14C]phenanthrene showed that after 72 h, 53% of the total radioactivity was associated with a glucoside conjugate of 1-hydroxyphenanthrene (phenanthrene 1-O-beta-glucose). This metabolite was isolated by reversed-phase high-performance liquid chromatography and characterized by the application of UV absorption, 1H nuclear magnetic resonance, and mass spectral techniques. The results show that aromatic ring oxidation followed by glucosylation is a predominant pathway in the metabolism of the polycyclic aromatic hydrocarbon phenanthrene by C. elegans.     

Fungal transformation of fluoranthene.

The fungus Cunninghamella elegans ATCC 36112 metabolized approximately 80% of the 3-14C-labeled fluoranthene (FA) added within 72 h of incubation. C. elegans metabolized FA to trans-2,3-dihydroxy-2,3-dihydrofluoranthene (trans-2,3-dihydrodiol), 8- and 9-hydroxyfluoranthene trans-2,3-dihydrodiol, 3-fluoranthene-beta-glucopyranoside, and 3-(8-hydroxyfluoranthene)-beta-glucopyranoside. These metabolites were separated by thin-layer and reversed-phase high-performance liquid chromatography and identified by 1H nuclear magnetic resonance, UV, and mass spectral techniques. The major pathway involved hydroxylation to form a glucoside conjugate of 3-hydroxyfluoranthene and a glucoside conjugate of 3,8-dihydroxyfluoranthene which together accounted for 52% of the total ethyl acetate-soluble metabolites. C. elegans initially metabolized FA in the 2,3 position to form fluoranthene trans-2,3-dihydrodiol, which has previously been shown to be a biologically active compound in mammalian and bacterial genotoxicity tests. However, C. elegans formed predominantly glucoside conjugates of the phenolic derivatives of FA, which suggests that this fungus has the potential to detoxify FA.     

Fungal transformation of fluoranthene

The fungus Cunninghamella elegans ATCC 36112 metabolized approximately 80% of the 3-14C-labeled fluoranthene (FA) added within 72 h of incubation. C. elegans metabolized FA to trans-2,3-dihydroxy-2,3-dihydrofluoranthene (trans-2,3-dihydrodiol), 8- and 9-hydroxyfluoranthene trans-2,3-dihydrodiol, 3-fluoranthene-beta-glucopyranoside, and 3-(8-hydroxyfluoranthene)-beta-glucopyranoside. These metabolites were separated by thin-layer and reversed-phase high-performance liquid chromatography and identified by 1H nuclear magnetic resonance, UV, and mass spectral techniques. The major pathway involved hydroxylation to form a glucoside conjugate of 3-hydroxyfluoranthene and a glucoside conjugate of 3,8-dihydroxyfluoranthene which together accounted for 52% of the total ethyl acetate-soluble metabolites. C. elegans initially metabolized FA in the 2,3 position to form fluoranthene trans-2,3-dihydrodiol, which has previously been shown to be a biologically active compound in mammalian and bacterial genotoxicity tests. However, C. elegans formed predominantly glucoside conjugates of the phenolic derivatives of FA, which suggests that this fungus has the potential to detoxify FA.     

Assimilation of chlorinated alkanes by hydrocarbon-utilizing fungi.

The fatty acid compositions of two filamentous fungi (Cunninghamella elegans and Penicillium zonatum) and a yeast (Candida lipolytica) were determined after the organisms were grown on 1-chlorohexadecane or 1-chlorooctadecane. These organisms utilized the chlorinated alkanes as sole sources of carbon and energy. Analyses of the fatty acids present after growth on the chlorinated alkanes indicated that 60 to 70% of the total fatty acids in C. elegans were chlorinated. Approximately 50% of the fatty acids in C. lipolytica were also chlorinated. P. zonatum contained 20% 1-chlorohexadecanoic acid after growth on either substrate but did not incorporate C18 chlorinated fatty acids.     

Stereoselective metabolism of anthracene and phenanthrene by the fungus Cunninghamella elegans.

The fungus Cunninghamella elegans oxidized anthracene and phenanthrene to form predominately trans-dihydrodiols. The metabolites were isolated by reversed-phase high-pressure liquid chromatography for structural and conformational analyses. Comparison of the circular dichroism spectrum of the fungal trans-1,2-dihydroxy-1,2-dihydroanthracene to that formed by rat liver microsomes indicated that the major enantiomer of the trans-1,2-dihydroxy-1,2-dihydroanthracene formed by C. elegans had an S,S absolute stereochemistry, which is opposite to the predominately 1R,2R dihydrodiol formed by rat liver microsomes. C. elegans oxidized phenanthrene primarily in the 1,2-positions to form trans-1,2-dihydroxy-1,2-dihydrophenanthrene. In addition, a minor amount of trans-3,4-dihydroxy-3,4-dihydrophenanthrene was detected. Metabolism at the K-region (9,10-positions) of phenanthrene was not detected. Comparison of the circular dichroism spectra of the phenanthrene trans-1,2- and trans-3,4-dihydrodiols formed by C. elegans to those formed by mammalian enzymes indicated that each of the dihydrodiols formed by C. elegans had an S,S absolute configuration. The results indicate that there are differences in both the regio- and stereoselective metabolism of anthracene and phenanthrene between the fungus C. elegans and rat liver microsomes.     

gamma-Linolenic acid production by solid-state fermentation of Mucorales strains on cereals

Oleaginous fungi of the genus Mucorales were screened for gamma-linolenic acid (GLA) production on solid substrates containing moistened cereals. Cunninghamella elegans CCF 1318 produced the highest yields of GLA when cultivated on barley. Substrate moisture and cultivation temperature proved critical for effective GLA production. Vegetable oil supplied to the cultures improved GLA production. Rotating bottles and plastic bags were used as cultivation vessels to reproduce the conditions found in rotating drums and tray bioreactors, respectively. After 11 days of cultivation at 21 degrees C, C. elegans produced 14.2 mg of GLA per gram of dry substrate, composed of a mixture of barley, spent malt grains (SMG) and peanut oil. GLA represented 15.6% of the total fatty acids in the lipid extract.