• 한국균학회소식:학술대회논문집
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• 2016.05a
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• pp.19-20
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• 2016

Sesquiterpenoids are defined as $C_{15}$ compounds derived from farnesyl pyrophosphate (FPP), and their complex structures are found in the tissue of many diverse plants (Degenhardt et al. 2009). FPP's long chain length and additional double bond enables its conversion to a huge range of mono-, di-, and tri-cyclic structures. A number of cyclic sesquiterpenes with alcohol, aldehyde, and ketone derivatives have key biological and medicinal properties (Fraga 1999). Fungi, such as the wood-rotting Polyporus brumalis, are excellent sources of pharmaceutically interesting natural products such as sesquiterpenoids. In this study, we investigated the biosynthesis of P. brumalis sesquiterpenoids on modified medium. Fungal suspensions of 11 white rot species were inoculated in modified medium containing $C_6H_{12}O_6$, $C_4H_{12}N_2O_6$, $KH_2PO_4$, $MgSO_4$, and $CaCl_2$ for 20 days. Cultivation was stopped by solvent extraction via separation of the mycelium. The metabolites were identified as follows: propionic acid (1), mevalonic acid lactone (2), ${\beta}$-eudesmane (3), and ${\beta}$-eudesmol (4), respectively (Figure 1). The main peaks of ${\beta}$-eudesmane and ${\beta}$-eudesmol, which were indicative of sesquiterpene structures, were consistently detected for 5, 7, 12, and 15 days These results demonstrated the existence of terpene metabolism in the mycelium of P. brumalis. Polyporus spp. are known to generate flavor components such as methyl 2,4-dihydroxy-3,6-dimethyl benzoate; 2-hydroxy-4-methoxy-6-methyl benzoic acid; 3-hydroxy-5-methyl phenol; and 3-methoxy-2,5-dimethyl phenol in submerged cultures (Hoffmann and Esser 1978). Drimanes of sesquiterpenes were reported as metabolites from P. arcularius and shown to exhibit antimicrobial activity against Gram-positive bacteria such as Staphylococcus aureus (Fleck et al. 1996). The main metabolites of P. brumalis, ${\beta}$-Eudesmol and ${\beta}$-eudesmane, were categorized as eudesmane-type sesquiterpene structures. The eudesmane skeleton could be biosynthesized from FPP-derived IPP, and approximately 1,000 structures have been identified in plants as essential oils. The biosynthesis of eudesmol from P. brumalis may thus be an important tool for the production of useful natural compounds as presumed from its identified potent bioactivity in plants. Essential oils comprising eudesmane-type sesquiterpenoids have been previously and extensively researched (Wu et al. 2006). ${\beta}$-Eudesmol is a well-known and important eudesmane alcohol with an anticholinergic effect in the vascular endothelium (Tsuneki et al. 2005). Additionally, recent studies demonstrated that ${\beta}$-eudesmol acts as a channel blocker for nicotinic acetylcholine receptors at the neuromuscular junction, and it can inhibit angiogenesis in vitro and in vivo by blocking the mitogen-activated protein kinase (MAPK) signaling pathway (Seo et al. 2011). Variation of nutrients was conducted to determine an optimum condition for the biosynthesis of sesquiterpenes by P. brumalis. Genes encoding terpene synthases, which are crucial to the terpene synthesis pathway, generally respond to environmental factors such as pH, temperature, and available nutrients (Hoffmeister and Keller 2007, Yu and Keller 2005). Calvo et al. described the effect of major nutrients, carbon and nitrogen, on the synthesis of secondary metabolites (Calvo et al. 2002). P. brumalis did not prefer to synthesize sesquiterpenes under all growth conditions. Results of differences in metabolites observed in P. brumalis grown in PDB and modified medium highlighted the potential effect inorganic sources such as $C_4H_{12}N_2O_6$, $KH_2PO_4$, $MgSO_4$, and $CaCl_2$ on sesquiterpene synthesis. ${\beta}$-eudesmol was apparent during cultivation except for when P. brumalis was grown on $MgSO_4$-free medium. These results demonstrated that $MgSO_4$ can specifically control the biosynthesis of ${\beta}$-eudesmol. Magnesium has been reported as a cofactor that binds to sesquiterpene synthase (Agger et al. 2008). Specifically, the $Mg^{2+}$ ions bind to two conserved metal-binding motifs. These metal ions complex to the substrate pyrophosphate, thereby promoting the ionization of the leaving groups of FPP and resulting in the generation of a highly reactive allylic cation. Effect of magnesium source on the sesquiterpene biosynthesis was also identified via analysis of the concentration of total carbohydrates. Our current study offered further insight that fungal sesquiterpene biosynthesis can be controlled by nutrients. To profile the metabolites of P. brumalis, the cultures were extracted based on the growth curve. Despite metabolites produced during mycelia growth, there was difficulty in detecting significant changes in metabolite production, especially those at low concentrations. These compounds may be of interest in understanding their synthetic mechanisms in P. brumalis. The synthesis of terpene compounds began during the growth phase at day 9. Sesquiterpene synthesis occurred after growth was complete. At day 9, drimenol, farnesol, and mevalonic lactone (or mevalonic acid lactone) were identified. Mevalonic acid lactone is the precursor of the mevalonic pathway, and particularly, it is a precursor for a number of biologically important lipids, including cholesterol hormones (Buckley et al. 2002). Farnesol is the precursor of sesquiterpenoids. Drimenol compounds, bi-cyclic-sesquiterpene alcohols, can be synthesized from trans-trans farnesol via cyclization and rearrangement (Polovinka et al. 1994). They have also been identified in the basidiomycota Lentinus lepideus as secondary metabolites. After 12 days in the growth phase, ${\beta}$-elemene caryophyllene, ${\delta}$-cadiene, and eudesmane were detected with ${\beta}$-eudesmol. The data showed the synthesis of sesquiterpene hydrocarbons with bi-cyclic structures. These compounds can be synthesized from FPP by cyclization. Cyclic terpenoids are synthesized through the formation of a carbon skeleton from linear precursors by terpene cyclase, which is followed by chemical modification by oxidation, reduction, methylation, etc. Sesquiterpene cyclase is a key branch-point enzyme that catalyzes the complex intermolecular cyclization of the linear prenyl diphosphate into cyclic hydrocarbons (Toyomasu et al. 2007). After 20 days in stationary phase, the oxygenated structures eudesmol, elemol, and caryophyllene oxide were detected. Thus, after growth, sesquiterpenes were identified. Per these results, we showed that terpene metabolism in wood-rotting fungi occurs in the stationary phase. We also showed that such metabolism can be controlled by magnesium supplementation in the growth medium. In conclusion, we identified P. brumalis as a wood-rotting fungus that can produce sesquiterpenes. To mechanistically understand eudesmane-type sesquiterpene biosynthesis in P. brumalis, further research into the genes regulating the dynamics of such biosynthesis is warranted.

• Magazine of the Korean Society of Agricultural Engineers
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• v.11 no.1
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• pp.1534-1548
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• 1969

This experiment was conducted, making use of the 'NONG-RIM6' arecommended variety of rice for the year of 1968. Main purposes of the experiment are to explore possibilities of; a) ways and means of saving irringation water and, b) overcoming drought at the same time so that an increased yield in rice could be resulted in. Specifically, it was tried to determine the effects of the Rotation irrigation method combined with differentiated thickness of lining upon the growth and yield of rice. Some of the major findings are summarized in the following. 1) The different thicknesses show a significant relationship with the weight of 1,000 grains. In the case of 9cm lined plot, the grain weight is 23.5grams, the heaviest. Next in order is 3cm lined plot, 6cm lined plot, control plot, and wheat straw lined-plot. 2) In rice yield, it is found that there is a considerably moderate significant relationship with both the different thickness of lining and the number of irrigation, as shown in the table. 3) There is little or no difference among different plots in terms of a) physical and chemical properties of soil, b) quality of irrigation water, c) climatic conditions, and rainfalls. 4) It is found that there is a significant relationship between differences in the method of rotation irrigation and the number of ears per hill. The plot irrigated at an interval of 7 days shows 17.4 ears and plot irrigated at an interval of 6 days, 16.3 5) In vinyl-treated plots, it is shown that both yield and component elements are greatest in the case of the plot ith whole of $3cm/m^2$ Next in order are the plot with a hole of $2cm/m^2$ the plot with a hole of $1cm/m^2$ In the case of the plot with no hole it is found that both yield and component elements are decreased as compared to the control plot. 6) The irrigation water reqirement is measured for the actual irrigation days of 72 which are the number subtracted the days of rainfall of 30 from the total irrigation days of 102. It is found that the irrigation water requirement for the uncontrol plot is 1,590mm as compared to 876mm(44.9% saved) for the 9cm-lined plot, 959mm(39.7% saved) for the 6cm-lined plot 1,010mm(36% saved) for the 3cm-lined plot and 1,082mm(32% saved) for the wheat straw lined plot. In the case of the Rotation irrigation method it is found that the water requirement for the plot irrigated at an interval of 8 days is 538mm(65% saved), as compared to 617mm(61.6% saved) for plot irrigated at an interval of 7 day 672mm(57.7% saved) for plot irrigated at an interval of 6day, 746mm(53.0% saved) for the plot irrigated at an interval of 5 days, 890mm 44.0% saved) for the plot irrigated at an interval of 4 days, and 975mm(38.6% saved) for the plot irrigated at an interval of 3 days. 7) The rate of evapotranspiration is found 2.8 around the end of month of July, as compared to 2.6 at the begining of August 3.4 around the end of August and 2.6 at the begining of August 3.4 around the end of August and 2.6 at the begining of September. 8) It is found that the saturation quantity of 30mm per day is decreased to 20mm per day though the use of vinyl covering. 9) The husking rate shows 75 per cent which is considered better.