Abstract:Penicillium ramulosum N1 was isolated from decaying wood. This strain produces extracellular xylanases and cellulases. The highest activities of xylanases (250 U/ml) and carboxymethyl cellulose (CMCase; 6.5 U/ml) were produced when 1% barley straw was added as a carbon source. The optimum temperature and pH for xylanase activity was 55 and 3.0°C, respectively. The xylanases exhibited strong protease resistance. CMCase revealed maximum activities at pH 3.0 and in the range of 60-70°C. Filter paper activity was … Show more
“…Penicillium citrinum isolate HZN13 was observed to possess globose conidia, septate hyphae and branched conidiophore after staining with lactophenol cotton blue. Several researchers have reported the isolation of xylanase-producing isolates like Aspergillus tubingensis FDHN1 from compost pit (Adhyaru et al 2015 ), Penicillium ramulosum N1 from decaying wood (Lin et al 2015 ), Aspergillus fumigatus isolate R1 (Deshmukh et al 2016 ) and P. citrinum xym2 from garden soil (Saha and Ghosh 2014 ). Fig.…”
The present study reports the production of high-level cellulase-free xylanase from Penicillium citrinum isolate HZN13. The variability in xylanase titers was assessed under both solid-state (SSF) and submerged (SmF) fermentation. SSF was initially optimized with different agro-waste residues, among them sweet sorghum bagasse was found to be the best substrate that favored maximum xylanase production (9643 U/g). Plackett–Burman and response surface methodology employing central composite design were used to optimize the process parameters for the production of xylanase under SSF. A second-order quadratic model and response surface method revealed the optimum conditions for xylanase production (sweet sorghum bagasse 25 g/50 ml; ammonium sulphate 0.36 %; yeast extract 0.6 %; pH 4; temperature 40 °C) yielding 30,144 U/g. Analysis of variance (ANOVA) showed a high correlation coefficient (R
2 = 97.63 %). Glutaraldehyde-activated calcium-alginate-immobilized purified xylanase showed recycling stability (87 %) up to seven cycles. Immobilized purified xylanase showed enhanced thermo-stability in comparison to immobilized crude xylanase. Immobilization kinetics of crude and purified xylanase revealed an increase in K
m (12.5 and 11.11 mg/ml) and V
max (12,500 and 10,000 U/mg), respectively. Immobilized (crude) enzymatic hydrolysis of sweet sorghum bagasse released 8.1 g/g (48 h) of reducing sugars. Xylose and other oligosaccharides produced during hydrolysis were detected by High-Performance Liquid Chromatography. The biomass was characterized by Scanning Electron Microscopy, Energy Dispersive X-ray and Fourier Transformation Infrared Spectroscopy. However, this is one of the few reports on high-level cellulase-free xylanase from P. citrinum isolate using sweet sorghum bagasse.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-016-0484-9) contains supplementary material, which is available to authorized users.
“…Penicillium citrinum isolate HZN13 was observed to possess globose conidia, septate hyphae and branched conidiophore after staining with lactophenol cotton blue. Several researchers have reported the isolation of xylanase-producing isolates like Aspergillus tubingensis FDHN1 from compost pit (Adhyaru et al 2015 ), Penicillium ramulosum N1 from decaying wood (Lin et al 2015 ), Aspergillus fumigatus isolate R1 (Deshmukh et al 2016 ) and P. citrinum xym2 from garden soil (Saha and Ghosh 2014 ). Fig.…”
The present study reports the production of high-level cellulase-free xylanase from Penicillium citrinum isolate HZN13. The variability in xylanase titers was assessed under both solid-state (SSF) and submerged (SmF) fermentation. SSF was initially optimized with different agro-waste residues, among them sweet sorghum bagasse was found to be the best substrate that favored maximum xylanase production (9643 U/g). Plackett–Burman and response surface methodology employing central composite design were used to optimize the process parameters for the production of xylanase under SSF. A second-order quadratic model and response surface method revealed the optimum conditions for xylanase production (sweet sorghum bagasse 25 g/50 ml; ammonium sulphate 0.36 %; yeast extract 0.6 %; pH 4; temperature 40 °C) yielding 30,144 U/g. Analysis of variance (ANOVA) showed a high correlation coefficient (R
2 = 97.63 %). Glutaraldehyde-activated calcium-alginate-immobilized purified xylanase showed recycling stability (87 %) up to seven cycles. Immobilized purified xylanase showed enhanced thermo-stability in comparison to immobilized crude xylanase. Immobilization kinetics of crude and purified xylanase revealed an increase in K
m (12.5 and 11.11 mg/ml) and V
max (12,500 and 10,000 U/mg), respectively. Immobilized (crude) enzymatic hydrolysis of sweet sorghum bagasse released 8.1 g/g (48 h) of reducing sugars. Xylose and other oligosaccharides produced during hydrolysis were detected by High-Performance Liquid Chromatography. The biomass was characterized by Scanning Electron Microscopy, Energy Dispersive X-ray and Fourier Transformation Infrared Spectroscopy. However, this is one of the few reports on high-level cellulase-free xylanase from P. citrinum isolate using sweet sorghum bagasse.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-016-0484-9) contains supplementary material, which is available to authorized users.
“…The purified Xyl-IIb was cellulase-free with no measurable activities with CMC and only showed activity with birchwood and oat spelts xylans as the substrates. Xylanases with differing molecular mass from different organisms have been studied, such as 19 and 14 kDa from Trichoderma inhamatum (Silva et al 2015 ) and 25 kDa from Penicillium ramulosum N1 (Lin et al 2015 ). Multiple xylanase forms have also been studied in A. fumigates Z5 (Miao et al 2015 ) and Penicillium oxalicum GZ-2 (Liao et al 2015 ).…”
An extracellular thermostable xylanase (Xyl-IIb) produced by Penicillium citrinum isolate HZN13 was purified to homogeneity using DEAE-Sepharose, Sephadex G-100 and Bio-Gel P-60 chromatography with specific activity of 6272.7 U/mg and 19.6-fold purification. The purification revealed the occurrence of multiple forms of xylanases (Xyl-I, Xyl-IIa, Xyl-IIb and Xyl-III). The molecular mass of highly purified Xyl-IIb was ~31 kDa with SDS-PAGE. The enzyme was cellulase-free, thermostable (55–75 °C) and acidophilic (3.5–5.0). It was activated by Ca2+, Ba2+, DTT and β-mercaptoethanol, whereas inhibited by Hg2+, Pb2+, Ni2+ and p-CMB. Purified Xyl-IIb exhibited highest specificity toward birchwood and oat spelts xylan. Kinetics of Xyl-IIb revealed a K
m of 10 mg/ml and 16.7 mg/ml and V
max of 9523g and 15,873 U/mg with birchwood and oat spelts xylan, respectively, indicating high affinity toward birchwood xylan. The xylanase (Xyl-IIb) belongs to glycosyl hydrolase (GH) family 10 based on conserved regions. Xylanase-encoding gene (xynB) consists of 1501 bp with an open reading frame of 264 bp which was predicted to encode a protein having 87 amino acids and shared homology with endo-1,4-beta-xylanase (xynB) gene from Penicillium citrinum. Cloned xynB gene was expressed in E. coli BL21 (DE3) with xylanase activity (80 U/mg) and confirmed to be GH-10 Xyl-IIa based on molecular mass (~40 kDa). These properties of xylanase make it promising for their applications in biofuel industries.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-016-0489-4) contains supplementary material, which is available to authorized users.
“…The polymerase chain reaction (PCR) products were cloned into the pUCM-T vector and sequenced. Then, the sequence was retrieved by basic local alignment search tool (BLAST) searches on the NCBI (https://blast.ncbi.nlm.nih.gov/Blast.cgi) website, aligned using ClustalX 1.83 (Thompson et al 1997), and subjected to phylogenetic analysis using the neighborjoining method with MEGA6 (MEGA Inc., Englewood, NJ) using 1,000 bootstraps (Lin et al 2015).…”
Section: Isolation and Identification Of Laccase-producing Strain Bacmentioning
confidence: 99%
“…To determine the molecular weight of the laccase from the A4 strain, the purified enzyme was run, along with standard protein marker, using a 12% (W/V) polyacrylamide gel with an 8% stacking gel according to the method of Lin et al (2015). After electrophoresis, the gel was divided into two parts.…”
Section: Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (Smentioning
Bacillus sp. A4 exhibiting laccase production was isolated from forest soil. Its laccase secreted into a LB medium exhibited a maximum activity of 3.9 U mg -1 protein at the optimal temperature (37 °C) and pH (6.0). The purified laccase of Bacillus sp. A4 demonstrated a low molecular mass of 33 kDa, and its optimal temperature and pH were 40 °C and 4.6, respectively, when using ABTS as a substrate. The activity of the purified laccase was significantly increased in the presence of Cu 2+ , methanol, and ethanol, but it was totally inhibited by L-cysteine. The laccase production of this strain was markedly stimulated when the strain was incubated with 0.5% different lignocellulosic biomasses. The highest activity of laccase (22.6 U mg -1 protein) was obtained in using algal biomass. This new strain efficiently decreased the lignin content of lignocellulose biomasses after 9 d of incubation at 37 °C, especially lignin from grasses. Further analysis showed that, compared to that of all tested biomasses, the new strain was a more efficient decomposer of the lignin of Miscanthus, which exhibited much more lignin loss and cell wall structure destruction in a short span of time. Therefore, the potential use of this strain could be advantageous for using lignin in Miscanthus for industrial processes.
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