• 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.

• The Korean Journal of Nuclear Medicine Technology
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• v.21 no.1
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• pp.39-43
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• 2017

Purpose Thyroglobulin (Tg) is a biologic marker of differentiated thyroid carcinoma (DTC), produced by normal thyroid tissue or thyroid cancer tissue. Therefore, the Tg values of DTC patients is the most specific indicator for judging whether recurrence occur or whether the remaining thyroid cancer is present. Thyroid cancer is currently the most common cancer in Korea, of which 90% is differentiated thyroid cancer. The number of patients with thyroid disease of this application also increased, and an accurate and prompt results are required. However, the incubation time of the Tg commonly takes about 24 hours in our hospital, and the result reporting time is delayed, and We could not satisfied with the requirements of clinical departments and patients. In order to fulfill these requirements, experiments were conducted by shortening the incubation time between company B's Kit currently in use and company C's Kit used in other hospitals. Through these experiments, we could perform the correlation with the original method and shortening method, and could find the optimum reaction time to satisfy the needs of the departments and the patients, and we will improve the competitiveness with the EIA examination. Materials and Methods In September 2016, we tested 65 patients company B's kit and company C's kit by three incubation ways. First method $37^{\circ}C$ shaking 2hr/2hr, Second method RT shaking 3hr/2hr, Third method 1hr/1hr shaking at $37^{\circ}C$. Fourth method RT shaking 3hr method which is the original method of Company C's Kit. Fifth method, the incubation time was shortened under room temperature shaking 2hr, Sixth method $37^{\circ}C$ shaking 2hr. And we performed and compared the correlation and coefficient of each methods. Results As a result of performing shortening method on company B currently in use, when comparing the Original method of company B kit, First method $37^{\circ}C$ shaking 2hr/2hr was less than Tg 1.0 ng/mL and the ratio of $R^2=0.5906$, above 1.0 ng/mL In the value, $R^2=0.9597$. Second method RT shaking 3hr/2hr was $R^2=0.7262$ less than value of 1.0 ng/mL, $R^2=0.9566$ above than value of 1.0 ng/mL. Third method $37^{\circ}C$ shaking 1hr/1hr was $R^2=0.7728$ less than value of 1.0 ng/mL, $R^2=0.8904$ above than value of 1.0 ng/mL. Forth, Company C's The original method, RT shaking 3hr was $R^2=0.7542$ less than value of 1.0 ng/mL, and $R^2=0.9711$ above than value of 1.0 ng/mL. Fifth method RT shaking 2hr was $R^2=0.5477$ less than value of 1.0 ng/mL, $R^2=0.9231$ above than value of 1.0 ng/mL. Sixth method $37^{\circ}C$ shaking 2hr showed $R^2=0.2848$ less than value of 1.0 ng/mL, $R^2=0.9028$ above than value of 1.0 ng/mL. Conclusion Samples with both values of 1.0 ng/mL or higher in both of the six methods showed relatively high correlation, but the correlation was relatively low less than value of 1.0 ng/mL. Especially, the $37^{\circ}C$ shaking 2hr method of company C showed a sharp fluctuation from the low concentration value of 1.0 ng/mL or less. Therefore, we are planning to continuously test the time, equipment, incubation temperature and so on for the room temperature shaking 2hr method and $37^{\circ}C$ shaking 1hr/1hr of company C which showed a relatively high correlation. After that, we can search for an appropriate shortening method through additional experiments such as recovery test, dilution test, sensitivity test, and provide more accurate and prompt results to the department of medical treatment, It is competitive with EIA test.

• Korean Journal of Agricultural Science
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• v.5 no.2
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• pp.93-114
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• 1978

The purpose of the study was to find out possible ways for increasing farm income through the sheep and Korean native goats farming, and to investigate meat productivity, wool productivity; woolskin utility, physiological characteristics and correlation between economical college animal farm of the Chungnam National University and sample farms in the suburbs of Dae jeon City were selected for feeding 20 heads of Corriedale wethers and another 20 heads Korean native kids as research materials for the periods of 5th May-26th November, 1977. The data such as growth rate, carcass, viscera weight, blood picture and plamsa components, hebage intake and economic traits were obtained and analysed. The result of the study are summarized as follows: 1. Meat production and quality 1) After 196days of feeding, the body weight of sheep and Korean native goats was increased by two times of those at the beginning of the trial, i.e. 20kg and 8kg respectively. 2) There was no significance of growth rates of sheep in housing and grazing. 3) The growth rate of Korean native goats were excellent at the mountainous areas of Gong ju-Gun where infectious diseases were not found 4) Accroding to the body measurements of 18-month-old sheep, percentages of hip height, body length, rump length, chest depth, chest width, hip width, chest girth and forearm circumference to the withers height were 103,%, 104%, 33%, 44%, 31%, 23%, 135% and 15% respectively, and those of hip height, body length, chest depth and chest girth of 8-month-old native goats to the withers height were 106%, 109%, 46% and 122,% respecitively. As a result, it was found that the percentage of hip height, body length and chest depth of Korean native goats were higher than those of sheep while that of the chest girth of goats was lower. 5) In the carcass data, 47, $52{\pm}2.27%$ of carcass percentage, $34.61{\pm}1.62%$ of lean meat, $26.07{\pm}2.51%$ of viscera, $9.75{\pm}1.4%$ of bone, and $20.95%{\pm}2.14%$ of woolskin for sheep, and $45.58{\pm}5.63%$ of carcass percentage, $27.62{\p}3.81%$ of meat, $34.86{\pm}4.16%$ of viscera, $11.66{\pm}1.83%$ of bone, $3.63{\pm}1.61%$ of skull and $9.26{\pm}2.41%$ of woolskin for native goats were obtained. 6) The contents of moisture, crude protein, crude fat and crude ash in native goat meat were much similar in both plots of housing and grazing. It was, however, known that the contents of moisture and protein were higher in grazinrg than in housing, while fat content was lower in grazing plots. 7) The weights of visceral organs shown similar tendency for both of sheep and native goats. For the weights of liver, heart, kidney and spleen, significance was not reconized among the treatments. Those of rumen, reticulum, small and large intestine were heavier in grazing than in housing, while the amount of visceral fat was heavier in housing. 2. Wool productivity and woolskin 1) The wool production of sheep for 7 months was $3.88{\pm}1.02kg$, and wool percentage, staple length, straighten length, wool growth per day and number of crimps were $9.27{\pm}1.48%$, 8. $47{\pm}1.00cm$, $10.63{\pm}0.99cm$, $0.40{\pm}0.04cm$ and $2.78{\pm}0.40$ respecitively. 2) The tensile strength and tear strength of woolskin treated by alum tanning were highest on the skin obtained from rump, i.e. $1,351kg/mm^2$ and $2,252kg/mm^2$ respectively, and they are in order of loin and shoulder. 3. Utilization and improvement of pasture. 1) The difference of herbage intake of native goats was not recognized between grazing and tethering, but the intake in the afternoon was s lightly higher than that in the morning. However the hervage intake of sheep was superior in grazing and in the afternoon. 2) The cultivation effect was lower in the native goat plots due to their cultivation abilities, in other words, the establishment rates of pasture by hoof cultivation were 60.25% in the goat plots and 77.35% in the sheep plots. 4. Correlation among economical traits. 1) The correlation between live weight of sheep and daily gain was higher. On the other hand, the correlation between other traits was not significant except that live weight, daily gain and lean meat percentage to the length of thoracic vertebrae. The live weight of native goats and meat production were highly correlated, and high correlation was also found between weights of carcass and meat. However, negative correlation was shown between viscera weight and live weight as well as daily gain. 2) The correlatoin between fleece weight of sheep and other traits such as live weight, daily gain and fleece percentage is very high at the 1% siginficant level, and this means that rapid-growth individuals can produce much fleece. 3) The correlation between the factors such as weights of live body, lean meat and viscera of sheep and body measurements, i. e. chest girth and body length was highest, and weights, of carcass and lean meat was highly correlated to chest width and depth. It will be therefore reasonable that the meat productivity estimates will have to be made on the basis of chest girth and body length. The meat production traits of native goats were highly correlated to the most of body measurement data, and the correlation coefficient between chest girth and weights of live body, carcass, lean meat and bone percentage was very high, i. e. 0.992-0.974 in particular. The correlations of meat production traits to chest depth, forearm circumference, body length were 0.759-0.911, 0.759-0.909 and 0.708-0.872 respectively. Therefore, the meat production of native goats will have to be estimated on the basis of chest data. 5. Blood picture and plasma components. 1) The number of erythrocyte and MCHC of native goats were $12.93{\times}10^6/mm^3$ and 36.14%, and those of sheep were $10.68{\times}10^6/mm^3$ and 36.26 respectively. The values of native goats were significantly higher than those of sheep. 2) The hemoglobin concentration, PVC, MCV and MCR of native goats were 10.92 g/100ml, $23.40{\mu}^3$ and 10.94 pg, and those of sheep were 11.73 g/100ml, 36.25 ml/100ml, $33.97{\mu}^3$ and 30.2 ml/100ml 8.43 pg respectively. The values of native goats were significantly lower those of sheep. 3) The number of leukocytes of native goats was significantly higher than that of sheep, that is, $11.64{\times}10^3/mm^3$ in native goats and $9.32{\times}10^3/mm^3$ in sheep. 4) In differential count of leukocyte, neutrophil was significantly high in native goats while lympocyte in sheep. On the other hand, the basophil, eosinophil and monocyte were not significant between native goats and sheep. 5) The amounts of total protein and glucose in the plasma of native goats were 6.2g/100ml and 53.6mg/100ml, and those of sheep were 5.6g/100ml and 45.7mg/100ml, which means that the values of native goats were significantly higher that those of sheep. The amount of total-lipid of native goats(127.6mg/100ml) was significantly than that of sheep(149.6mg/100ml). 6) The amount of non-protein nitrogen, cholesterol, Ca, P, K, Na and Cl were not different between native goats and sheep. 6. Economic analysis. 1) The gross revenue of a farm which fed native goats and sheep was 4,000won per head and the optimum size for feeding them in a farm as a subsidiary work is 5-10 heads. 2) Since there was no difference between housing and grazing, they can be fed in group for farm's subsidiary work. 3) They can be also fed by youths and house wives in the suburbs of cities, because labour requirement is estimated as only two hours per days for feeding 5 heads of native goats and sheep.