Chunqi Zhao

Bioethanol production from hydrolysates of inulin and the tuber meal of Jerusalem artichoke by Saccharomyces sp. W0.

It has been confirmed that Saccharomyces sp. W0 can produce high concentration of ethanol. However, this yeast strain cannot secrete inulinase. Therefore, in this study, inulin was hydrolyzed into reducing sugar by the recombinant inulinase produced by Pichia pastoris X-33/pPICZaA-INU1. It was found that 38.2U of the recombinant inulinase per gram of inulin was suitable for the inulin hydrolysis and ethanol production by Saccharomyces sp. W0 and the fermentation period was 120 h. At the end of the fermentation, over 14.6 ml of ethanol per 100ml of the fermented medium was produced, the ethanol productivity was over 0.384 g of ethanol/g of inulin and over 98.8% of total sugar was utilized. When the Saccharomyces sp. W0 was grown in the mixture of 4.0% hydrolysate of soybean meal and 20.0% of the hydrolysate of inulin for 120 h, over 14.9 ml of ethanol per 100ml of the fermented medium was yielded, the ethanol productivity was over 0.393 g of ethanol/g of inulin and 98.9% of total sugar was used by the yeast strain. When Saccharomyces sp. W0 carrying the same inulinase gene was grown in the medium containing 50 g of the tuber meal of Jerusalem artichoke per 100ml for 144 h, over 12.1+/-0.35%ml of ethanol per 100ml of the fermented medium was yielded, the ethanol productivity was 0.319+/-0.9 g of ethanol/g of sugar and 3.7% (w/v) of total sugar and 0.5% (w/v) of reducing sugar were left in the fermented media.

Structural characterization and antioxidant properties of an exopolysaccharide produced by the mangrove endophytic fungus Aspergillus sp. Y16

A homogeneous exopolysaccharide, designated As1-1, was obtained from the culture medium of the mangrove endophytic fungus Aspergillus sp. Y16 and purified by anion-exchange and gel-permeation chromatography. Results of chemical and spectroscopic analyses, including one- and two-dimensional nuclear magnetic resonance (1D and 2D NMR) spectroscopy showed that As1-1 was mainly composed of mannose with small amounts of galactose, and that its molecular weight was about 15 kDa. The backbone of As1-1 mainly consists of (1→2)-linked α-d-mannopyranose units, substituted at C-6 by the (1→6)-linked α-d-mannopyranose, (1→)-linked β-d-galactofuranose and (1→)-linked β-d-mannopyranose units. As1-1 possessed good in vitro antioxidant activity as evaluated by scavenging assays involving 1,1-diphenyl-2-picrylhydrazyl (DPPH) and superoxide radicals. The investigation demonstrated that As1-1 is an exopolysaccharide different from those of other marine microorganisms, and could be a potential antioxidant and food supplement.

Chemical characteristic of an anticoagulant-active sulfated polysaccharide from Enteromorpha clathrata

A sulfated polysaccharide FEP from marine green alga Enteromorpha clathrata was extracted with hot water and further purified by ion-exchange and size-exclusion chromatography. Results of chemical and spectroscopic analyses showed that FEP was a high arabinose-containing sulfated polysaccharide with sulfate ester of 31.0%, and its average molecular weight was about 511kDa. The backbone of FEP was mainly composed of (1→4)-linked β-L-arabinopyranose residues with partially sulfate groups at the C-3 position. In vitro anticoagulant assay indicated that FEP effectively prolonged the activated partial thromboplastin time and thrombin time. The investigation demonstrated that FEP was a novel sulfated polysaccharide with different chemical characteristics from other sulfated polysaccharides from marine algae, and could be a potential source of anticoagulant.

Preparation, structure and anticoagulant activity of a low molecular weight fraction produced by mild acid hydrolysis of sulfated rhamnan from Monostroma latissimum.

A low molecular weight fraction, designated LMWP, was prepared by mild acid hydrolysis of sulfated rhamnan from Monostroma latissimum and purified by anion-exchange and gel-permeation chromatography. Chemical and spectroscopic analyses showed that LMWP was mainly composed of rhamnose, and its molecular weight was about 33.6 kDa. The backbone of LMWP consists of 1,3-linked α-L-rhamnose units with partially sulfate groups at the C-2 position. Approximately 25% of 1,3-linked α-L-rhamnose units is substituted at C-2 by sulfated or non-sulfated 1,3-linked α-L-rhamnose and 1,2-linked α-L-rhamnose units. LMWP effectively prolonged clotting time as evaluated by the activated partial thromboplastin time assay and was a potent thrombin inhibitor mediated by heparin cofactor II. The investigation demonstrated that LMWP is a novel sulfated polysaccharide with anticoagulant activity.

Structural elucidation of an extracellular polysaccharide produced by the marine fungus Aspergillus versicolor

A homogenous extracellular polysaccharide, designated AWP, was isolated from the fermented liquid of the marine fungus Aspergillus versicolor from the coral Cladiella sp. and purified by anion-exchange and size-exclusion chromatography (SEC). Chemical and spectroscopic analyses, including one- and two-dimensional nuclear magnetic resonance (1D and 2D NMR) spectroscopy showed that AWP consisted of glucose and mannose in a molar ratio of 8.6:1.0, and its average molecular weight was estimated to be 500kDa. AWP is a slightly branched extracellular polysaccharide. The backbone of AWP is mainly composed of (1→6)-linked α-D-glucopyranose residues, slightly branched by single α-d-mannopyranose units attached to the main chain at C-3 positions of the glucan backbone. The investigation demonstrated that AWP is a novel extracellular polysaccharide different from those of other marine microorganisms.

Galactomannan with novel structure produced by the coral endophytic fungus Aspergillus ochraceus

The homogeneous extracellular polysaccharide, AW1, was obtained from the fermented broth of the fungus Aspergillus ochraceus derived from coral Dichotella gemmacea. AW1 was a galactomannan with a molar ratio of mannose and galactose of 2.16:1.00 and a molecular weight of about 29.0kDa. The structure of AW1 was investigated by chemical and spectroscopic methods, including methylation analysis, one- and two-dimensional nuclear magnetic resonance (1D, 2D NMR) and electrospray mass spectrometry with collision-induced dissociation (ES-CID MS/MS) spectroscopic analyses. The results showed that the backbone of AW1 consisted of (1⟶2)-linked α-d-mannopyranose residues. The mannopyranose residues in the backbone were substituted at C-6 by the (1⟶)-linked α-d-mannopyranose units and (1⟶5)-linked β-d-galactofuranose oligosaccharides with different degrees of polymerization. The investigation demonstrated that AW1 was a novel galactomannan with different structural characteristics from other fungal galactomannans, and could be a potential resource of the (1⟶5)-linked β-d-galactofuranose oligosaccharides.

Preparation and structural elucidation of a glucomannogalactan from marine fungus Penicillium commune.

The coral-associated fungus Penicillium commune produces an extracellular polysaccharide, FP2-1, when grown in potato dextrose-agar medium. FP2-1 was isolated from the fermented broth using anion-exchange and size-exclusion chromatography, and its structure was elucidated by chemical and spectroscopic analyses, including detailed nuclear magnetic resonance spectroscopy. The results showed that FP2-1 was a glucomannogalactan with a molar ratio of galactose, mannose and glucose of 3.9:1.9:1.0. Structure of FP2-1 may be represented, at an average, as a backbone of (1→2)-linked α-mannopyranose with the every second residue substituted at position 6 by a pentasaccharide branch. The branches consist of four (1→6)-linked β-galactofuranose residues with terminal α-glucopyranose residue attached to the last galactofuranose residue at position 2. FP2-1 was a novel galactofuranose-containing extracellular polysaccharide differing from previously described extracellular polysaccharides.

Sequence analysis of the sulfated rhamno-oligosaccharides derived from a sulfated rhamnan

Three sulfated rhamno-oligosaccharides, designated O1, O2 and O3, were obtained by mild acid hydrolysis of the sulfated rhamnan and purified by gel-permeation chromatography. On the basis of one- and two-dimensional nuclear magnetic resonance (1D, 2D NMR) spectroscopic analyses, the oligosaccharide O1 was characterized to be α-L-Rhap-(2SO4)-(1→3)-α-L-Rhap. The fragmentation pattern of the homogeneous disaccharide in the product ion spectra was recognized by negative-ion electrospray tandem mass spectrometry with collision-induced dissociation (ES-CID MS/MS). With the principles established, the sequences of the oligosaccharides O2 and O3 were deduced to be α-L-Rhap-(2SO4)-(1→3)-α-L-Rhap-(1→3)-α-L-Rhap, and α-L-Rhap-(2SO4)-(1→3)-α-L-Rhap-(1→3)-α-L-Rhap-(1→3)-α-L-Rhap (2SO(4)), respectively. The investigation demonstrated that the sulfated rhamnan-derived oligosaccharides were novel sulfated oligosaccharides different from those of other polysaccharides-degraded from algae, and it could be possible to determine the sequence of the sulfated rhamno-oligosaccharides directly from the glycosidic cleavage fragmentation in the product ion spectra.

Chemical characteristics and anticoagulant activities of two sulfated polysaccharides from Enteromorpha linza (Chlorophyta)

Two sulfated polysaccharides, designated MP and SP, were extracted from the marine green alga Enteromorpha linza using hot water and then purified using ion-exchange and size-exclusion chromatography. The anticoagulant activities of MP and SP were examined by determination of their activated partial thromboplastin time (APTT), thrombin time (TT) and prothrombin time (PT) using human plasma. Results showed that MP and SP were composed of abundant rhamnose with small amounts of xylose and glucuronic acid, whereas SP also contained a small amount of galactose. Approximate molecular weights of MP and SP were 535 and 502 kDa, respectively. As compared with SP, MP had higher contents of sulfate ester (19.0%) and uronic acid (14.9%). The MP mainly consisted of (1→4)-linked rhamnose residues with partially sulfated groups at the C-3 position, and small amounts of (1→3, 4)-linked rhamnose, (1→2, 4)-linked rhamnose, (1→4)-linked glucuronic acid and (1→4)-linked xylose residues. The SP contained abundant (1→4)-linked rhamnose with minor amounts of (1→3)-linked rhamnose, (1→3, 4)-linked rhamnose, (1→2, 4)-linked rhamnose, (1→4)-linked glucuronic acid, (1→4)-linked xylose, and (1→3)-linked galactose residues. The sulfate groups were mainly located at C-3 of (1→4)-linked rhamnose residues. Both MP and SP, in particular the former, effectively prolonged APTT and TT. This work demonstrates that MP and SP have unique structural characteristics distinct from those of other sulfated polysaccharides from Enteromorpha. The MP is a potential source of anticoagulant, and the difference in anticoagulant activities of the two sulfated polysaccharides is directly linked to the discrepancy of their chemical features.