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Lichen: from genome to ecosystem in a changing world (3I – P4) Submission ID: IAL0175-00001 EFFECT OF SUGAR ON SECONDARY METABOLISM IN CULTURED LICHEN MYCOBIONT OF CLADONIA RAMULOSA (3I – P5) Submission ID: IAL0180-00001 3I-P Hara K. 1 , Usuniwa Y. 1 , Komine M. 1 , Yamamoto Y. 1 1 Graduate School of Bioresource Sciences, Akita Prefectural University, Akita, Japan Lichens produce peculiar secondary metabolites known as lichen substances. However, cultured lichen mycobionts tend not to produce lichen substances. To investigate the influence of sugar on the secondary metabolism in lichenized fungi, 125 strains of mycobiont were cultured on malt–yeast extract medium (MY, control), MY medium with 20% sucrose (S20) and Lilly–Bernett medium (LB). After three or six months, lichen substances were produced on S20 medium by four species, while only one species Cladonia ramulosa produced lecanoric acid (LA) after one-month culture on LB. Addition of sugar alcohols on LB medium decreased LA levels. The LA level was elevated on LB with fructose, suggesting that polyketide biosynthetic pathway was regulated by sugars or was involved in sugar metabolisms. Two partial cDNAs of C. ramulosa polyketide synthases (CrPKS1 and CrPKS2) were cloned and used for RT-PCR analyses. The levels of CrPKS1 transcript began to accumulate and reached a maximum level at 12 days after transferring to LB medium with fructose, prior to LA production at 16 days. It is hypothesized that CrPKS1 products synthesize lecanoric acids and that fructose plays a key role in control of symbiotic/non-symbiotic metabolism in C. ramulosa mycobionts. HETEROLOGOUS EXPRESSION OF POLYKETIDE SYNTHASE GENES OF LICHEN CLADONIA METACORALLIFERA Kim J. 1 , Yu N. H. 1 , Jeong M. H. 1 , Hur J. 1 1 Korean Lichen Research Institute, Sunchon National University, Sunchon, Korea Lichens produce unique polyketide secondary metabolites including depsides, depsidones, dibenzofurans, and depsones. The biosynthesis of these compounds is governed by polyketide synthase (PKS), but the mechanism via which they are produced has remained unclear until now. Heterologous expression in a surrogate host provides an alternative approach for functional analysis of lichen polyketide biosynthesis. Cultured mycobiont of Cladonia metacorallifera producing a large amount various polyketide were used to isolate and characterize polyketide synthase genes. The CmPKSs showed greatest homology with uncharacterized genes from filamentous fungi and composed exclusive clades in reducing and non-reducing PKSs. We construct subclones using spliced full length cDNA of CmPKS1 and CmPKS35 for stable expression in the filamentous fungus Aspergillus nidulans. There are 10% efficiency of single copy inserted transforments about CmPKS1 and CmPKS35. We are expecting the new polyketide product by LC-MS and HPLC analysis. 134
The 7 th International Association for Lichenology Symposium 2012 (3I – P6) Submission ID: IAL0186-00001 ANTIOXIDANT CAPACITY OF PSEUDOEVERNIA FURFURACEA EXTRACT Ćilerdžić J. 1 , Stajić M. 1 , Vukojević J. 1 1 Institute of Botany, University of Belgrade, Faculty of Biology, Belgrade, Serbia Evaluation of the antioxidant capacity of ethanol extract of Pseudoevernia furfuraceae (L) Zopf. was the aim of this study. P. furfuraceae originated from Tara Mountain (Serbia) was used for the study. 5.5 g of air dried sample was extracted by stirring with 30-fold higher volume of 70% ethanol at 30 º C for 72 hours and filtering through Whatman No. 4 filter paper. The obtained ethanol extract was evaporated at 40 º C to dryness, redissolved in 96% ethanol to a final concentration of 20 mg/ml. Antioxidant capacity was determined spectrophotometrically by methanol solution of DPPH at 517 nm. Antioxidant activity of the tested extract concentrations were studied in comparison to commercial antioxidant, BHA. The concentrations of phenol and flavonoid compounds in the extract were measured spectrophotometrically by Folin-Ciocalteu reagent at 740 nm and ethanol/ aluminium nitrate/potassium acetate mixture at 415 nm, respectivelly. Tests were carried out by various extract dilutions, in triplicate. Six extract concentrations ranged from 10 mg/ml to 0.312 mg/ml, obtained by double disolutions, caused decrease of DPPH free radical-scavenging in the range from 90.04% to 21.39%. Defined EC 50 value was 1.85 mg/ml. The tested extract concentrations had strong antioxidant activity compared to BHA one (92.61%). The amounts of total phenol compounds were ranged from 61.53 µg/ml (10 mg/ml) to 5.540 µg/ ml (1.25 mg/ml), while at lower extract concentrations these compounds were missed. The total flavonoids content was significantly higher, ranged from 240.13 μg/ml (10 mg/ml) to 62.97 μg/ml (0.625 mg/ml), while at the concentration of 0.312 mg/ml their presence was not noted. Although, antioxidant activity had some value at the concentration of 0.312 mg/ml, it could be concluded that other compounds were responsible for it. Scavenging effect directly correlates with the different flavonoid content (r 2 = 0.9641) and less with phenol compounds content (r 2 = 0.6826). (3I – P7) Submission ID: IAL0236-00001 ANTIFUNGAL ACTIVITY AGAINST PLANT PATHOGENIC FUNGI FROM CRUDE EXTRACT OF USNEA PULVINULATA Pengproh R. 1 , Papong K. 1 , Sangdee A. 1 , Chantiratikul P. 2 1 Biology, Mahasarakham University, Mahasarakham, Thailand 2 Chemistry, Mahasarakham University, Mahasarakham, Thailand Usnea pulvinulata contains various secondary metabolites such as usnic acid, diffractaic acid, menegazziaic acid, norstictic acid, protocetraric acid, salazinic acid and ursoric acid. This study tested the inhibition of crude extracts of U. pulvinulata from absolute water, acetone, ethanol and methanol against 6 plants pathogenic fungi such as Colletotrichum gloeosporiodes (banana anthracnose, cultivated banana anthracnose, chili anthracnose, Dendrobium anthracnose and mango anthracnose), Colletotrichum capsici (chili anthracnose and papaya anthracnose), Curvularia lunata (dirty panicle disease of rice), Diplodia sp. (leaf blight of Dendrobium), Fusarium monilifore (bakanea disease of rice) and Pestalotiopsis guepinii (leaf blight of Guava). The crude extract of U. pulvinulata from acetone showed the highest inhibition against all six plants pathogenic fungi. Ethanol and methanol crude extracts can be antifungal against plant pathogen fungi. The crude extract from absolute water, however, did not inhibit the tested fungi. 135 3I-P
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The 7 th IAL Symposium 2012 Lichens
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The 7 th International Association
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Welcome Message The 7 th Internatio
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Report The 7 th International Assoc
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IAL COUNCIL (2008-2012): The 7 th I
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