Mucor Spinosus Research Articles

Three new triterpenoids transformed from ursolic acid by Mucor spinosus AS3.3450 and their cytotoxicity

Microbial transformation of ursolic acid by Mucor spinosus AS 3.3450 was carried out in this paper. Three new compounds (1-3) were isolated and their structures were identified as 3β, 7β-dihydroxy-ursolic acid-28-ethanone (1), 3β, 7β, 21β-trihydroxy-ursolic acid (2), and 3β, 21β-dihydroxy-ursolic acid-3-O-β-D-glucopyranoside (3) based on extensive spectral analysis including HRESI-MS, 1H-NMR, 13C-NMR, HMQC and HMBC. Compared with the substrate, compound 1 had stronger cytotoxic activity against Hela cell, K562 cell and KB cell lines. Using Mucor spinosus AS 3.3450 to bio-transform ursolic acid for the first time, this study provided possible ways for reforming the chemical structure of UA and the obtained activity-enhanced bio-transformation products may lay a foundation for the development and application of lead compounds in the future.

Stereo- and regiospecific biotransformation of curcumenol by four fungal strains

Biotransformations of curcumenol (1) were performed by four fungal strains, Mucor spinosus AS 3.2450, Penicillium urticae IFFI 04015, Cunninghamella echinulata AS 3.3400, Aspergillus carbonarius IFFI 02087. Five metabolites were prepared in the biotransformation process of 1, and their structures were elucidated as 15-hydroxycurcumenol (2), 1α-hydroxycurcumenol (3), 14-hydroxycurcumenol (4), 3β-hydroxycurcumenol (5) and 12-hydroxycurcumenol (6) by spectroscopic data analysis. Among them, metabolites 2–5 are novel. All of these four fungal strains showed the ability of highly stereo- and regiospecific hydroxylation for the substrate (1), which could be used as tools for preparing the hydroxylated derivatives and in vivo metabolites of curcumenol. In addition, the inhibitory effects of substrate and obtained products on nitric oxide production in lipopolysaccaride-activated macrophages were evaluated. The substrate (1) and metabolites 2, 5, and 6 showed significant inhibitory effects.

Microbial transformation of 20(S)-protopanaxatriol by Mucor spinosus.

Microbial transformation of 20(S)-protopanaxatriol (1) by Mucor spinosus afforded two new and eight known metabolites. The structures of the metabolites were identified as 12-oxo-20(S)-protopanaxatriol (2), 23,24-en-25-methoxyl-20(S)-protopanaxatriol (3), 29-hydroxyl-20(S)-protopanaxatriol (4), 20(S),24(S)-epoxy-dammaran-3β,6α,12β,25-triol (5), 12-oxo-27-hydroxyl-20(S)-protopanaxatriol (6), 12-oxo-26-hydroxyl-20(S)-protopanaxatriol (7), 12-oxo-23β-hydroxyl-20(S)-protopanaxatriol (8), 12-oxo-11β-hydroxyl-20(S)-protopanaxatriol (9), 12-oxo-15α-hydroxyl-20(S)-protopanaxatriol (10), and 12-oxo-25,26-en-15α-hydroxyl-20(S)-protopanaxatriol (11). Among them, 3 and 11 are new compounds. The metabolites 3, 10 and 11 showed the more potent inhibitory effects against seven cell lines than the substrate.

Effects of Extraction Method on the Physicochemical and Mycological Qualities of <i>Canarium Schweinfurthii</i> Fruit Oil

The effects of improved method of extraction on the physicochemical, mycological and stability of crude Canarium Schweinfurthii fruit oil were studied. The extracted oils were then stored at 25±5oC for 24 months with samples analyzed at 6months interval for; pH, saponification value, acid value, peroxide value and iodine number. Similarly, enumeration and identification of fungi species was determined using standard mycological procedures. The results showed that crude Canarium Schweinfurthii fruit oil obtained by the improved method of extraction had better quality and stability parameters than the traditional method extracted oil. At 24months the oil quality values of; pH 5.20 – 6.61, Acid value 0.53 – 1.02 (mg of KOH/g), saponification value 151.30 – 179.52 (mg of OH/g), peroxide value 0.031 – 1.500 (meq 02/Kg) and iodine value 85.02 – 101.60 (gI2/100g). Comparatively, no significant difference (P > 0.05) was observed for pH and saponification values for the extracted oils, with values of 6.60, 178.60(mg of OH/g) and 6.62, 178.52(mg of OH/g) for traditional and improved extraction methods respectively. During storage the oils showed average fungal counts of 00.00 to 1.72 x105 and 00.00 to 8.00 x104 CFU/ml oil at 0 and 24 month for traditional and improved methods extracted oils respectively. Predominant fungal species; Aspergillus niger, Rhizopus stolonifer, Sacccaromyces cerevisiae, Mucor spinosus, Penicillium patalun, Fusarium oxysprum and Candida scotti were found associated with the stored canarium oil. Generally, Mucor spinosus (80.00%), Aspergillus niger (80.00%) and Penicillium patalum (80.00%) had the highest occurrence in traditional extracted oil. This finding suggests the need for the local processors to reevaluate the full processing method in order to retain better fungal quality and oxidative stability for Canarium schweinfurthii fruit oil.Keywords: Canarium schweinfurthii, fruit oil, Extraction, Fungal Isolates, Oxidative stability

Biotransformation of osthole by Mucor spinosus

Osthole (1) as a natural product, has the potent anti-osteoporosis activity. In the present paper, a scaled-up biotransformation of 1 by Mucor spinosus AS 3.3450 was carried out for improving its activities and water-solubility. Nine metabolites were obtained in the biotransformation process of 1, and their structures were elucidated as 4′-hydroxyl-osthole (2), osthenol (3), osthenol-7-O-β-d-glucoside (4), 5′-hydroxyl-osthole (5), 5′-hydroxyl-2′,3′-dihydro-osthole (6), 2′,3′-dihydroxyl-osthole (7), 2′-ketone-3′-hydroxyl-osthole (8), 2′-acetoxy-3′-hydroxy-osthole (9) and 2′-hydroxyl-isoothole (10), on the basis of extensive spectral data including 2D NMR and MS spectroscopy. Among them, metabolites 6 and 8 were new. The hydroxylation, dehydrogenation, demethylation and glycosylation reactions of 1 by M. spinosus AS 3.3450 were reported in the present study. And anti-osteoporosis activities of all products were also evaluated by using MC3T3-E1 cells.

A multiphasic approach for investigation of the microbial diversity and its biodegradative abilities in historical paper and parchment documents

The microbial diversity of different kinds of stains present on the surface of 14 historical documents (nine parchments and five paper letters) was evaluated through a combination of cultural and molecular methods. The samples were recovered using adhesive tape and swabs and were afterwards treated in two different ways: (1) direct inoculation on agar plates; or (2) suspension in physiological solution and plating in specific plates for the growth of bacteria and fungi. The isolated microorganisms, before identification, were selected by two different PCR-based methods – f-ITS and f-CBH, for bacteria and fungi, respectively. The f-ITS method is based on the amplification of the internal transcribed sequence between the bacterial 16S and 23S rDNA. The f-CBH method is a new molecular selection tool oriented to the fungal cellobiohydrolase gene. Both PCR selection methods produced typical profiles, which clustered the isolates in order to reduce them for subsequent sequencing identification through the amplification of the fungal 28S rRNA and the bacterial 16S rRNA genes. The cellulolytic and proteolytic abilities were screened through the use of three plate assays, the Ostazin Brilliant Red H-3B (OBR-HEC), milk agar, and gelatin agar. Massilia timonae, Lysobacter dokdonensis, and strains belonging to the genus Bacillus sp., Microbacterium sp., and Curtobacterium sp. with different fungal members such as Aspergillus fumigatus, Penicillium commune, Mucor spinosus, and Phoma herbarum (all recovered from paper) displayed both biodegradative activities. The parchment isolates with a marked proteolytic activity were Bacillus cereus, Staphylococcus epidermidis, Pseudomonas oryzihabitans, Virgibacillus sp., Micromonospora sp., and again members of the fungal genera Penicillium, Mucor, and Phoma.

Associations between Fungal Species and Water-Damaged Building Materials

Fungal growth in damp or water-damaged buildings worldwide is an increasing problem, which has adverse effects on both the occupants and the buildings. Air sampling alone in moldy buildings does not reveal the full diversity of fungal species growing on building materials. One aim of this study was to estimate the qualitative and quantitative diversity of fungi growing on damp or water-damaged building materials. Another was to determine if associations exist between the most commonly found fungal species and different types of materials. More than 5,300 surface samples were taken by means of V8 contact plates from materials with visible fungal growth. Fungal identifications and information on building material components were analyzed using multivariate statistic methods to determine associations between fungi and material components. The results confirmed that Penicillium chrysogenum and Aspergillus versicolor are the most common fungal species in water-damaged buildings. The results also showed Chaetomium spp., Acremonium spp., and Ulocladium spp. to be very common on damp building materials. Analyses show that associated mycobiotas exist on different building materials. Associations were found between (i) Acremonium spp., Penicillium chrysogenum, Stachybotrys spp., Ulocladium spp., and gypsum and wallpaper, (ii) Arthrinium phaeospermum, Aureobasidium pullulans, Cladosporium herbarum, Trichoderma spp., yeasts, and different types of wood and plywood, and (iii) Aspergillus fumigatus, Aspergillus melleus, Aspergillus niger, Aspergillus ochraceus, Chaetomium spp., Mucor racemosus, Mucor spinosus, and concrete and other floor-related materials. These results can be used to develop new and resistant building materials and relevant allergen extracts and to help focus research on relevant mycotoxins, microbial volatile organic compounds (MVOCs), and microparticles released into the indoor environment.

Microbial transformation of neoandrographolide by Mucor spinosus (AS 3.2450)

Abstract Microbial transformation of neoandrographolide ( 1 ), was performed by Mucor spinosus (AS 3.2450). Ten metabolites were obtained and identified as 14-deoxyandrographolide ( 2 ), 17,19-dihydroxy-8,13- ent -labdadien-16,15-olide ( 3 ), 3,14-dideoxyandrographolide ( 4 ), 7β-hydroxy-3,14-dideoxyandrographolide ( 5 ), 17,19-dihydroxy-7,13- ent -labdadien-16,15-olide ( 6 ), 8(17),13- ent -labdadien-16,15-olid-19-oic acid ( 7 ), 8α,17β-epoxy-3,14-dideoxyandrographolide ( 8 ), 8β,17,19-trihydroxy- ent -labd-13-en-16, 15-olide ( 9 ), phlogantholide-A ( 10 ), 19-[(β- d -glucopyranosyl)oxy]-19-oxo- ent -labda-8(17),13-dien-16,15-olide ( 11 ) by spectroscopic and chemical means. Among them, products 3 , 5 , 6 , 8 and 9 were characterized as new compounds. The inhibitory effects of compounds 1 – 11 on nitric oxide production in lipopolysaccharide-activated macrophages were evaluated and their preliminary structure–activity relationships (SAR) were discussed.

Biotransformation of 20(S)-protopanaxadiol by Mucor spinosus

Biotransformation of 20(S)-protopanaxadiol ( 1) by the fungus Mucor spinosus AS 3.3450 yielded eight metabolites ( 2– 9). On the basis of NMR and MS analyses, the metabolites were identified as 12-oxo-15α,27-dihydroxyl-20(S)-protopanaxadiol ( 2), 12-oxo-7β,11α,28-trihydroxyl-20(S)-protopanaxadiol ( 3), 12-oxo-7β,28-dihydroxyl-20(S)-protopanaxadiol ( 4), 12-oxo-15α,29-dihydroxyl-20(S)-protopanaxadiol ( 5), 12-oxo-7β,15α-dihydroxyl-20(S)-protopanaxadiol ( 6), 12-oxo-7β,11β-dihydroxyl-20(S)-protopanaxadiol ( 7), 12-oxo-15α-hydroxyl-20(S)-protopanaxadiol ( 8), and 12-oxo-7β-hydroxyl-20(S)-protopanaxadiol ( 9), respectively. Among them, 2– 5, 7, and 8 are new compounds. These results indicated that M. spinosus could catalyze the specific C-12 dehydrogenation of 20(S)-protopanaxadiol, as well hydroxylation at different positions. These biocatalytic reactions may be difficult for chemical synthesis. The biotransformed products showed weak in vitro cytotoxic activities.

Biotransformation of 20( S)-protopanaxatriol by Mucor spinosus and the cytotoxic structure activity relationships of the transformed products

Biotransformation of 20( S)-protopanaxatriol ( 1) by the fungus Mucor spinosus AS 3.3450 gave 10 metabolites ( 2– 10), of which 2– 5 were previously known. On the basis of NMR and MS analyses, structures 6– 10 were established as 12-oxo-23β-hydroxyl-20( S)-protopanaxatriol ( 6), 20 S,24 R-epoxy-dammaran-3β,6α,25-triol-12-one ( 7), 29-hydroxyl-20( S)-protopanaxatriol ( 8a), 12-oxo-11β-hydroxyl-20( S)-protopanaxatriol ( 8b), 28-hydroxyl-20( S)-protopanaxatriol ( 9) and 12-oxo-20( S)-protopanaxatriol ( 10). The biotransformation kinetics of 1 has been investigated and a possible biotransformation pathway proposed. The in vitro cytotoxic activities of metabolites against three human cancer cell lines were determined by the MTT method; compounds 8a, 9 and 10 had more potent inhibitory effects against HL-60 cell line than the substrate.

Cytotoxic biotransformed products from cinobufagin by Mucor spinosus and Aspergillus Niger

Cinobufagin ( 1) was one of important cardenolidal steroids and major components of Chan'Su, a famous traditional Chinese medicine. Bioconversion of cinobufagin by the fungi of Mucor spinosus and Aspergillus niger were investigated. Nine bioconversion products were obtained from M. spinosus and seven products from A. niger. Their structures were elucidated by high-resolution fast atom bombardment mass spectroscopy (HR-FAB-MS), extensive NMR techniques, including 1H NMR, 13C NMR, DEPT, 1H– 1H correlation spectroscopy (COSY), two-dimensional nuclear Overhauser effect correlation spectroscopy (NOESY), heteronuclear multiple quantum coherence (HMQC) and heteronuclear multiple bond coherence (HMBC). The in vitro cytotoxic activities against human hepatoma cells (HepG2, SMMC-7221 and BEL-7402) and human leukemia cells (K562, HL-60 and HEL) of all bioconversion products were determined by the MTT method, and their structure–activity relationships (SAR) were discussed.

Microbial transformation of curdione by Mucor spinosus

Twenty strains of filamentous fungi (from nine genera) and four species of bacteria were screened for their capabilities to transform curdione ( 1). Site-specific hydroxylation and epoxidation of the substrate by Mucor spinosus AS 3.3450 were reported. Five compounds were obtained, and their structures were identified as 1β,10α-epoxycurdione ( 2), 1α,10β-epoxycurdione ( 3), 2β-hydroxycurdione ( 4), 11-hydroxycurdione ( 5) and 2β,11-dihydroxycurdione ( 6), respectively, on the basis of their spectral data. Among them, products 5 and 6 are new compounds. The biotransformation kinetics was also investigated.

Microbial hydroxylation of cinobufagin by Mucor spinosus

Mucor spinosus has been employed for the biotransformation of cinobufagin (1) to afford three metabolites. On the basis of their physico-chemical data, the structures of the transformation products have been characterized as 1β-hydroxy-cinobufagin (2), 12β-hydroxy-cinobufagin (3) and 1β,12β-dihydroxy-cinobufagin (4), of which metabolites 2 and 4 are new compounds. In vitro cytotoxic activities of the biotransformation products and the substrate-cinobufagin have been assayed against four tumor cell lines of Bel 7420, BGC 823, HeLa and HL 60; they all showed cytotoxic activities.

Microbial Transformation of 20(S)-Protopanaxatriol by Mucor spinosus

Microbial biotransformation of 20(S)-protopanaxatriol (1) by the fungus Mucor spinosus (AS 3.3450) yielded four new metabolites, the structures of which were determined to be 12-oxo-15alpha-hydroxyl-20(S)-protopanaxatriol (2), 27-hydroxyl-20(S)-protopanaxatriol (3), 12-oxo-26-hydroxyl-20(S)-protopanaxatriol (4), and 12-oxo-27-hydroxyl-20(S)-protopanaxatriol (5), respectively, on the basis of their chemical and spectroscopic data. All the new metabolites, as well as the substrate, had significant cytotoxic effects on HL-60 cells (human leukemia cells).

Microbial Hydroxylation of Bufalin by Cunninghamella blakesleana and Mucor spinosus

The microbial transformation of a cytotoxic bufadienolide, bufalin (1), was carried out using two strains of filamentous fungi. Cunninghamella blakesleana catalyzed the specific 12alpha-hydroxylation of bufalin and produced 12alpha-hydroxybufalin (2) and 7beta,12alpha-dihydroxybufalin (3) as the major metabolites, together with 7beta-hydroxybufalin (4) and 12beta-hydroxybufalin (5) in low yields. Two minor products were isolated from the culture broth of Mucor spinosus and were identified as 7beta,15alpha-dihydroxybufalin (6) and 5beta,7beta-dihydroxybufalin (7), respectively. Metabolites 2, 3, 6, and 7 are new compounds, and their structures were fully characterized by NMR and MS spectroscopy.

Novel cytotoxic bufadienolides derived from bufalin by microbial hydroxylation and their structure–activity relationships

Microbial transformation was used to prepare novel cytotoxic bufadienolides. Twelve products ( 3– 14) were obtained from bufalin ( 1) by the fungus Mucor spinosus. Their structures were elucidated by high-resolution mass spectroscopy (HR-MS) and extensive NMR techniques, including 1 H NMR, 13 C NMR, DEPT, 1 H – 1 H correlation spectroscopy (COSY), two dimensional nuclear Overhauser effect correlation spectroscopy (NOESY), heteronuclear multiple quantum coherence (HMQC), and heteronuclear multiple bond coherence (HMBC). Compounds 3, 4, 9 and 11– 14 are new mono- or dihydroxylated derivatives of bufalin with novel oxyfunctionalities at C-1β, C-7β, C-11β, C-12β and C-16α positions. The in vitro cytotoxic activities against human cancer cell lines of 3– 14, together with 16 biotransformed products derived from cinobufagin ( 15– 30) were determined by the MTT method, and their structure–activity relationships (SAR) were discussed.

Characterization of fungal 17β-hydroxysteroid dehydrogenases

To promote understanding of the evolution of the steroid hormone signalling and hydroxysteroid dehydrogenases (HSDs), comparative characterization of fungal 17β-HSDs was performed. Constitutive 17β-HSD activity was determined in cytosols of the fungi: Cochliobolus lunatus, Pleospora herbarum, Fusarium lini, Trichoderma viride, Mucor spinosus, Rhizopus nigricans and Pleurotus ostreatus. The reaction equilibrium in all species except P. ostreatus was shifted towards reduction. The preferential coenzyme for reduction of androstenedione was NADPH, while for oxidation of testosterone, NAD + was preferred. The highest enzyme activities were found in the Ascomycete C. lunatus (152.4 nmol mg −1 h −1) and in the Basidiomycete P. ostreatus (69.1 nmol mg −1 h −1). No similarities on the protein and mRNA level between fungal 17β-HSDs and the purified enzyme from C. lunatus were observed. To investigate the nature of these enzymes, 17β-HSD was purified from P. ostreatus using ammonium sulphate precipitation, hydrophobic interaction chromatography, and affinity chromatography. The purified enzyme has an apparent molecular mass of ∼35 kDa and is probably a dimer as determined by gel filtration. Chemical modifications exposed Lys, His and Tyr as important for enzyme activity. Additionally, no similarities of C. lunatus and P. ostreatus enzymes were found to bacterial 3α,20β-HSD from Streptomyces hydrogenans, 3β,17β-HSD from Comamonas testosteroni and mammalian 17β-HSD types 1 and 4. The results thus suggest that there are most probably different enzymes responsible for 17β-HSD activity in filamentous fungi.

Culture analysis and external interaction models of mycelial growth

To investigate morphogenesis and in particular circularization mechanisms in young mycelia, we observe cultures of the zygomycete Mucor spinosus and develop discrete models of two-dimensional filamental branching growth. The models are based on the hypothesis that the fungus secretes a regulatory substance that diffuses into the surrounding medium and is detected by the growing hyphae. We also present a simple Markovian growth model without such a feedback, but yielding to analytical computations.

Responsiveness of basophil granulocytes of horses suffering from chronic obstructive pulmonary disease to various allergens

As basophils are the major effector cells of allergic reactions, confirmation of the allergic etiology of chronic obstructive pulmonary disease (COPD) was sought by the demonstration of a specific in vitro response of equine basophilic blood cells to some potential allergens (Aspergillus, Cladosporidium, Mucor, Penicillium, extracts of dust particles of hay and straw). The allergen induced degranulation of basophils and the histamine and protease release from basophils during incubation with the allergens were tested. By evaluating the results obtained from 14 COPD horses and eight controls it could be shown that the sensitivity of the basophils of affected horses was increased, particularly against the allergen extract of Mucor mucedo and Mucor spinosus. Further a greater percentage of COPD horses reacted positively with the Mucor allergen extract. The mitogenic stimulation of lymphocytes by PHA and by the allergen extracts used gave comparable results in affected and control horses. Thus the in vitro stimulation of basophils may be an easily to perform testing device for the identification of potential allergens involved in the pathogenesis of equine COPD.

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.