Cytotoxic and antimicrobial activities of ethyl acetate extract of mangrove plant Scyphiphora hydrophyllacea C. F. Gaertn— Associated fungi

Association between mangrove plants with fungi has been explored for many biological activities. The purpose of thisstudy was to determine the antimicrobial and cytotoxic activities of endophytic fungi isolated from mangrove plantsScyphiphora hydrophyllacea C. F. Gaertn. The isolation of endophytic fungus from the mangrove has been carriedout by the direct planting method on Sabouraud Dextrose Agar medium. Eleven fungal isolates were recovered andpurified. The isolates were cultivated on the rice medium for 1 month and extracted using ethyl acetate. Eleven ethylacetate extracts were screened for cytotoxic activity by Brine Shrimp Lethality Test method and 3-(4,5-dimethylthiazol2-yl)-2,5-diphenyltetrazolium bromide assay and for antimicrobial activity toward some human pathogenic microbes.The results showed that five (45.5%) isolates of endophytic fungi had antimicrobial activity against the growth ofStaphylococcus aureus, Escherichia coli, and Candida albicans with a diameter from 9.1 to 21 mm. Furthermore, fourextracts (36.4%) were cytotoxic (cell viability < 50%) against T47D cells. Isolate ADSH2 which exhibited the highestcytotoxic activity was identified molecularly as Penicillium chrysogenum. The results of this study indicate that theethyl acetate of endophytic fungi associated with Scyphiphora hydrophyllacea has the potential to be developed asantimicrobial and anticancer compounds.

Agroindustrial biomass for xylanase production by Penicillium chrysogenum: purification, biochemical properties and hydrolysis of hemicelluloses

Background: In this work, the xylanase production by Penicillium chrysogenum F-15 strain was investigated using agroindustrial biomass as substrate. The xylanase was purified, characterized and applied in hemicellulose hydrolysis. Results: The highest xylanase production was obtained when cultivation was carried out with sugar cane bagasse as carbon source, at pH 6.0 and 20°C, under static condition for 8 d. The enzyme was purified by a sequence of ion exchange and size exclusion chromatography, presenting final specific activity of 834.2 U·mg·prot-1. T he molecular mass of the purified enzyme estimated by SDS-PAGE was 22.1 kDa. The optimum activity was at pH 6.5 and 45°C. The enzyme was stable at 40°C with half-life of 35 min, and in the pH range from 4.5 to 10.0. The activity was increased in the presence of Mg+2 and Mn+2 and reducing agents such as DTT and ßmercaptoethanol, but it was reduced by Cu+2 and Pb+2 . The xylanase presented Km of 2.3 mM and Vmax of 731.8 U·mg·prot-1 with birchwood xylan as substrate. This xylanase presented differences in its properties when it was compared to the xylanases from other P. chrysogenum strains. Conclusion: The xylanase from P. chrysogenum F-15 showed lower enzymatic activity on commercial xylan than on hemicellulose from agroindustry biomass and its biochemistry characteristics, such as stability at 40°C and pH from 4.0 to 10.0, shows the potential of this enzyme for application in food, feed, pulp and paper industries and for bioethanol production.

Two new diphenyl ether derivatives from Penicillium chrysogenum MT-12, an endophytic fungus isolated from Huperzia serrata / 中草药

Objective To investigate the chemical constituents from Penicillium chrysogenum MT-12, an endophytic fungus isolated from Huperzia serrata. Methods The compounds were isolated and purified by using various column chromatographies including silica gel, Sephadex LH-20, and semi-preparative HPLC. The structures were established using extensive spectroscopic techniques such as IR, MS, and NMR. The anti-inflammatory and AChE inhibitory activities of all the isolates were also preliminarily investigated by using in vitro models. Results Eight diphenyl ether derivatives including penicichrysogenillide A (1), penicichrysogenillide B (2), talaromyone A (3), isopenicillide (4), penicillide (5), hydroxypenicillide (6), purpactin A (7), and penicichrysogenillide C (8) were isolated from the solid fermentation cultures of P. chrysogenum MT-12. Conclusion Compounds 1 and 2 are new compounds, and this is the first report for the isolation of compound 3 from Penicillium species and compounds 4-8 from P. chrysogenum. Compounds 1 and 2 exhibited inhibitory activities anginst the nitric oxide production in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage cells with IC50 value of (72.6 ± 2.3) μmol/L and (41.2 ± 1.4) μmol/L, respectively. In contrast, all the compounds didn't exhibit inhibitory activities on AChE at the concentration of 100 μmol/L.

The secondary metabolites from a mutant 3d10-01 derived from Penicilliumchrysogenum S-3-25 by diethyl sulfate mutagenesis / 国际药学研究杂志

Objective To investigate the secondary metabolites from diethyl sulfate(DES)mutant 3d10-01 derived from Peni-cillium chrysogenum S-3-25. Methods The secondary metabolites were isolated by multiple separation techniques,such as silica gel,Sephadex LH-20 column chromatography and HPLC. The structures of the compounds were determined by ESI-MS and NMR spectroscopic data. The antitumor activity was assayed by the MTT method. HPLC-UV analysis was used to determine if compounds 1-7 were newly produced by the mutant. Results Seven secondary metabolites,including methyl 2,4-dihydroxy-3,5,6-trimethylbenzo-ate(1),6-hydroxymethyl-2,2-dimethylchromanone(2),regiolone(3),(3S,4S)-3,4-dihydro-3,4,8-trihydroxy(2H)naphthalenone (4),cerebroside C(5),cerebroside D(6)and dankasterone A(7),were isolated from the fermentation products of the mutant 3d10-01. Compounds 1,4 and 7 showed strong inhibitory effects on the five tested cell lines,but 2,3 and 6 only exhibited the proliferation of HL-60 cells to some extent. Compounds 1-4 and 7 were newly produced by the mutant 3d10-01. Conclusion Compounds 1-7 are firstly isolated from P. chrysogenum. The inhibitory effects on certain tested antitumor cell lines of compounds 1-4,6 and 7 are report-ed for the first time.

Microbial Transformation of Two Prenylated Naringenins

Microbial transformation of (±)-6-(1,1-dimethylallyl)naringenin (6-DMAN, 1) and (±)-5-(O-prenyl) naringenin-4′,7-diacetate (5-O-PN, 2) was performed by using fungi. Scale-up fermentation studies with Mucor hiemalis, Cunninghamella elegans var. elegans, and Penicillium chrysogenum led to the isolation of five microbial metabolites. Chemical structures of the metabolites were determined by spectral analyses as (±)-8-prenylnaringenin (3), (2S)-5,4′-dihydroxy-7,8-[(R)-2-(1-hydroxy-1-methylethyl)-2,3-dihydrofurano]flavanone (4), (±)-5-(O-prenyl)naringenin-4′-acetate (5), (±)-naringenin-4′-acetate (6), and (±)-naringenin (7), of which 5 was identified as a new compound.

Analysis of fungal colonization in skin surfaces of patients with atopic dermatitis / 中华皮肤科杂志

Objective To analyze the colonization of Candida, Rhodotorula, Penicillium and Aspergillus in skin surfaces of patients with atopic dermatitis, and to assess the relationship between the four common fungal allergens and severity of atopic dermatitis. Methods Fifty patients with atopic dermatitis and 20 healthy controls were enrolled. Scales were scraped from lesional and non?lesional skin of flexural extremities of the patients, as well as from normal skin of the flexural elbow of healthy controls, then were subjected to microscopic examination and culture. Scale specimens were inoculated onto Sabouraud dextrose agar medium and cultured at 25 ℃ in a constant temperature incubator. Subsequently, suspected fungal or yeast?like colonies were collected for pure culture. Finally, fungal strains were identified according to colony morphology, color, growth speed, as well as microscopic features of spores and hyphae. Results No hyphae or pseudohyphae were found in any case by microscopic examination. Candida albicans and Rhodotorula were detected in 29(58%)and 17(34%)out of the 50 patients, respectively, and in 5(25%)and 2 (10%) out of the 20 healthy controls, respectively. The detection rates of Candida albicans and Rhodotorula were significantly higher in the patients than in the controls(χ2=6.23, 4.10, respectively, both P<0.05). Of 25 patients with severe lesions, 19(76%)and 12(48%)were colonized by Candida albicans and Rhodotorula respectively;among 25 patients with moderate lesions, 10 (40%) and 5 (20%) were colonized by Candida albicans and Rhodotorula respectively. An increase was observed in the detection rates of Candida albicans and Rhodotorula in the patients with severe lesions compared with those with moderate lesions(χ2=6.65, 4.37, respectively, both P<0.05). There was no significant difference in the detection rate of Penicillium or Aspergillus between the patients and health controls. Conclusion The colonization rates of Candida albicans and Rhodotorula on skin surfaces were higher in patients with atopic dermatitis than in healthy controls, and higher in patients with severe lesions than in patients with moderate lesions, indicating that the types of colonizing fungi are associated with the health status of skin and severity of symptoms in patients with atopic dermatitis.

Yeast HXK2 gene reverts glucose regulation mutation of penicillin biosynthesis in P. chrysogenum

The mutant Penicillium chrysogenum strain dogR5, derived from strain AS-P-78, does not respond to glucose regulation of penicillin biosynthesis and β-galactosidase, and is partially deficient in D-glucose phosphorilating activity. We have transformed strain dogR5 with the (hexokinase) hxk2 gene from Saccharomyces cerevisiae. Transformants recovered glucose control of penicillin biosynthesis in different degrees, and acquired a hexokinase (fructose phosphorylating) activity absent in strains AS- P-78 and dogR5. Interestingly, they also recovered glucose regulation of β-galactosidase. On the other hand, glucokinase activity was affected in different ways in the transformants; one of which showed a lower activity than the parental dogR5, but normal glucose regulation of penicillin biosynthesis. Our results show that Penicillium chrysogenum AS-P-78 and dogR5 strains lack hexokinase, and suggest that an enzyme with glucokinase activity is involved in glucose regulation of penicillin biosynthesis and β-galactosidase, thus signaling glucose in both primary and secondary metabolism; however, catalytic and signaling activities seem to be independent.

Biodegradation of phenol in static cultures by Penicillium chrysogenum ERK1: catalytic abilities and residual phototoxicity / Biodegradación de fenol en cultivos estáticos por Penicillium chrysogenum ERK1: habilidades catalíticas y fitotoxicidad residual

A phenol-degrading fungus was isolated from crop soils. Molecular characterization (using internal transcribed spacer, translation elongation factor and beta-tubulin gene sequences) and biochemical characterization allowed to identify the fungal strain as Penicillium chrysogenum Thorn ERK1. Phenol degradation was tested at 25 °C under resting mycelium conditions at 6, 30, 60, 200, 350 and 400 mg/l of phenol as the only source of carbon and energy. The time required for complete phenol degradation increased at different initial phenol concentrations. Maximum specific degradation rate (0.89978 mg of phenol/day/mg of dry weight) was obtained at 200 mg/l. Biomass yield decreased at initial phenol concentrations above 60 mg/l. Catechol was identified as an intermediate metabolite by HPLC analysis and catechol dioxygenase activity was detected in plate assays, suggesting that phenol metabolism could occur via ortho fission of catechol. Wheat seeds were used as phototoxicity indicators of phenol degradation products. It was found that these products were not phytotoxic for wheat but highly phytotoxic for phenol. The high specific degradation rates obtained under resting mycelium conditions are considered relevant for practical applications of this fungus in soil decontamination processes.

Un aislamiento fúngico capaz de degradar fenol como única fuente de carbono y energía fue aislado de suelos agrícolas. La caracterización molecular (basada en el empleo de secuencias de espaciadores de transcriptos internos, de factores de la elongación de la traducción y del gen de la beta-tubulina) y la caracterización bioquímica permitieron identificar a esta cepa como Penicillium chrysogenum Thom ERK1. Se estudió la degradación de fenol a 25 °C en cultivos estáticos con 6, 30, 60, 200, 350 y 400 mg/l de fenol inicial. El tiempo requerido para completar la degradación de fenol aumentó al elevarse las concentraciones iniciales de dicho compuesto. La máxima tasa de degradación específica (0,89978 mg de fenol/día/mg de peso seco) se obtuvo con 200 mg/l. El rendimiento en biomasa disminuyó con concentraciones Iniciales de fenol mayores de 60 mg/l. Se identificó al catecol como intermediarlo metabolico por HPLC y se observó actividad de catecol dioxigenasa en placa, lo que sugiere que el metabolismo de degradación del fenol ocurre vía orto fisión del catecol. Se utilizaron semillas de trigo como indicadores de fitotoxicidad de los productos de degradación. Estos productos no fueron fitotóxicos para trigo, mientras que el fenol mostró una alta fitotoxicidad. La alta tasa de degradación específica obtenida en condiciones estáticas resulta de gran interés para la aplicación de este hongo en procesos de descontaminación de suelos.

Penicillin production by wild isolates of Penicillium chrysogenum in Pakistan

The present study was aimed at exploring the native wild isolates of Penicillium chrysogenum series in terms of their penicillin production potential. Apart from the standard medium, the efforts were made to utilize suitable agro-industrial wastes for the maximum yield of penicillin. Two series of P. chrysogenum were isolated from local sources and named as P. chrysogenum series UAF R1 and P. chrysogenum series UAF R2. The native series were found to possess better penicillin production potential than the already reported series of P. chrysogenum. However, P. chrysogenum series UAF R1 was found to be the best candidate for high yield of penicillin starting at 100 hour as compared to P. chrysogenum series UAF R2 which produced the highest yield of penicillin at 150 hours for a shorter period of time. Addition of Corn Steep Liquor (CSL) to the fermentation medium resulted in the production of 1.20g/L penicillin by P. chrysogenum series UAF R1 and P. chrysogenum series UAF R2. The fermentation medium in which Sugar Cane Bagasse (SCB) was replaced with CSL resulted in the highest yield of penicillin (1.92g/L) by both native series of P. chrysogenum. The penicillin production was increased by 62.5% in medium with SCB as compared to that with CSL. The penicillin yield of medium containing lactose and phenyl acetate was higher than that of control medium. Overall results revealed that P. chrysogenum series UAF R1 and P. chrysogenum series UAF R2 may be recommended for better yield of natural penicillin and this efficiency may be further enhanced by utilizing SCB as substrate in the growth medium.

Molecular Cloning and Characterization of a Glutathione S-transferase Gene Repressed by Phenylacetic Acid From Penicillium chrysogenum / 生物化学与生物物理进展

Glutathione S-transferase (GST) gene, PcgstA was cloned from the penicillin producing strain Penicillium chrysogenum,which is important for understanding the industrial fermentation process. PcgstA gene has an open-reading-frame of 840 bp in length,which is interrupted by two introns. The deduced amino acid sequence shows about 50% identity to several characterized filamentous fungi GSTs. The recombinant PcGSTA in Escherichia coli were overexpressed and purified. Enzymatic assays showed that the recombinant PcGSTA had a specific activity with 1-chloro-2, 4-dinitrobenzene of (0.159±0.031) μmol/(min· mg). It was found that the expression level of PcgstA in the penicillin producing medium supplemented with phenylacetic acid, the side chain precursor of penicillin G, was significant down regulated than that in medium without phenylacetic acid. This result suggested that PcGST may be related to phenylacetic acid metabolism in the penicillin producing strain.