• Title/Summary/Keyword: Growth hormones

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THE EFFECT OF THE GENISTEIN ON THE PROLIFERATION OF HT1080 AND EXPRESSION OF MEMBRANE TYPE 1-MATRIX METALLOPROTEINASE (MT1-MMP) mRNA (Genistein이 사람 섬유육종 세포주 증식 및 Membrane Type 1-Matrix Metalloproteinase (MT1-MMP) mRNA 발현에 미치는 영향)

  • Kang, Jin-Han;Myoung, Hoon;Kim, Myung-Jin
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.27 no.4
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    • pp.314-320
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    • 2001
  • Matrix metalloproteinases have long been viewed as ideal candidates for proteinases that enables tumor cells to permeated basement membrane defenses and invade surrounding tissue. There is growing evidence that the MMPs have an expanded role, as they are important for the creation and maintenance of a microenvironment that facilitates growth and angiogenesis of tumors at primary and metastatic sites. MT-MMPs are not secreted but instead remaining attached to cell surfaces. Although not all of the MT-MMPs are fully characterized, MT-MMPs have important role in localizing and activating secreted MMPs. The MMP genes are transcriptionally responsive to a wide variety of oncogene, growth factors, cytokine, and hormones. Currently, a number of MMP inhibitors are being developed and some have reached clinical trials as anti-metastatic or anti-cancer therapies. MT1-MMP is involved in the activation of proMMP-2. MT1-MMP is significant not only as a tumor marker but as a new target for chemotherapy against cancer. The purpose of this study was to evaluate the effects of protein kinase C inhibitor(genistein) on the proliferation of HT1080 and expression of MT1-MMP mRNA. Human fibrosarcoma cell line HT1080 was cultured and divided 2 groups. The experimental group was treated with $100{\mu}M$ genistein and incubated 12h, 24h for $[3^H]-thymidine$ uptake assay and northern hybridization individually. And the control group was treated with same amount of PBS for the above procedures. $[3^H]-thymidine$ incorporation was measured with ${\beta}$ ray detector. And RT-PCR and northern blotting for MT1-MMP mRNA was performed. The results were as follows 1. $[3^H]-thymidine$ uptake was reduced in experimental group with statistical significance. 2. MT1-MMP mRNA expression was significantly reduced in experimental group. These results showed that protein kinase C inhibitor (genistein) inhibited proliferation of HT1080 and almost completely blocked transcription of MT1-MMP mRNA. So, it is possible to use the protein kinase inhibitor (genistein) as anti-metastatic and anti-proliferative agent.

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Milk Conjugated Linoleic Acid (CLA) Profile and Metabolic Responses of Dairy Cows Fed with High-temperature-micro-time (HTMT) Treated Diets Containing High Quantity Extruded Soybean (ESB)

  • Lee, H.G.;Hong, Z.S.;Wang, J.H.;Xu, C.X.;Jin, Y.C.;Kim, T.K.;Kim, Y.J.;Song, M.K.;Choi, Yun.-Jaei
    • Asian-Australasian Journal of Animal Sciences
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    • v.22 no.11
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    • pp.1504-1512
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    • 2009
  • A feeding trial was conducted to examine the effect of high-temperature-micro-time (HTMT) processing of diets containing extruded soybean (ESB) in high quantity on milk fat production, metabolic responses, and the formation of conjugated linoleic acid (CLA) and trans-vaccenic acid (TVA). Twenty-one multiparous Holstein cows in mid-lactation were blocked according to milk yield in the previous lactation. Cows within each block were randomly assigned to either normal concentrate or HTMT treated diets containing ESB (7.5% HTMT-ESB and 15% HTMT-ESB). It was hypothesized that the HTMT-ESB would affect the undegradable fatty acids in the rumen and, thus, would modify the fatty acid profile of milk fat. Both 7.5% and 15% HTMT-ESB did not affect milk yield, fat, protein, lactose and solid-not-fat (SNF), but the proportion of cis-9, trans-11 CLA in milk fat was significantly increased by these treatments. Content of TVA in milk fat was not affected by HTMT-ESB. The HTMT-ESB influenced the fatty acid profile in milk fat, but there was little difference between 7.5% and 15% of supplementation. HTMT-ESB feeding significantly decreased the concentration of plasma insulin and glucose, while plasma growth hormone (GH), triglyceride (TG), non-esterified fatty acid (NEFA) and HDLcholesterol were increased by 7.5% and 15% ESB-HTMT supplementation in comparison to the control group (p<0.05). However, no significant difference was observed in plasma LDL-cholesterol, insulin like growth factor (IGF)-1, T3, T4, and leptin concentrations among treatments (p>0.05). The present results showed that cis-9, trans-11 CLA production was increased by HTMT treatment of dietary ESB without reduction of milk fat, and the unchanged milk fat and yield was assumed to be associated with the constant level of thyroid hormones, leptin, and IGF-1.

Analysis of gibberellic acid from fruits using HPLC/UV-vis (HPLC/UV-vis을 이용한 과일류 중의 지베렐린 산 분석)

  • Ma, Kyung Na;Cho, Hyun-Woo;Myung, Seung-Woon
    • Analytical Science and Technology
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    • v.26 no.1
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    • pp.19-26
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    • 2013
  • Gibberllic acid ($GA_3$) is one of gibberellins (GAs) that are a class of plant growth hormones that exert profound and diverse effects on plant growth and development. $GA_3$ is essentially non-UV absorbing and is difficult to assay by UV-detector. For effective extraction of gibberellic acid from fruits by using liquid-liquid extraction, optimized pH and extraction solvent were established. The selective and sensitive derivative of $GA_3$ for HPLC/UV-vis was derivatized using phenacyl bromide, and the experimental factors, including reaction time, reaction temperature and amount of derivatizing reagent and base were investigated for the effective synthesis. The derivatized $GA_3$ with phenacyl bromide was effectively analyzed by HPLC/UV-vis. The structure of derivatized $GA_3$ was confirmed by HPLC/ESI-MS. For apple, LOD and LOQ were 0.008 mg/kg and 0.027 mg/kg, respectively. For pear, LOD and LOQ were 0.003 mg/kg, 0.012 mg/kg, respectively. The established method can be applied to more effective analysis of $GA_3$ from plant and food.

Effects of Thyroid Hormones on Settlement, Survival and Growth in Olive Flounder, Paralichthys olivaceus Larvae (Thyroid hormone 처리에 의한 넙치, Paralichthys olivaceus 자어의 착저, 생존 및 성장)

  • Bang In Chul;Kim Yoon;Kim Kyung-Kil;Kim Dong Soo
    • Journal of Aquaculture
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    • v.8 no.2
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    • pp.133-140
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    • 1995
  • Olive flounder (Paralichthys olivaceus) larvae of pre-metamorphic and early metamorphic stage were treated with different concentrations of triiodothyroxine $(T_3)\;and\; thyroxine\;(T_4)$ by immersion (0.02, 0.04, 0.08 and 0.16 ppm) for 11 days. Although $T_3$ was more potent than $T_4,\;both\;T_3\;and\;T_4$ accelerated the settlement time of fish larvae. Duration upto complete settlement in hormone-treated groups was significantly shorter than that in controls. Survival of hormone-treated groups at the pre-metamorphic stage decreased as hormone concentrations increased, however groups treated at the early metamorphic stage were not differ from control. Growth rates of groups treated with high concentration of hormone were slightly lower than that of control except 0.02ppm $T_4$ treated group.

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Effects of Weekly Administration of Implant-type Recombinant Porcine Somatotropin on the Performance and Carcass Characteristics of Finishing Pigs

  • Kim, Y.H.;Jung, H.J.;Park, J.C.;Kwon, O.S.;Chung, C.S.;Ko, Y.D.;Moon, H.K.
    • Asian-Australasian Journal of Animal Sciences
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    • v.18 no.4
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    • pp.557-561
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    • 2005
  • The present study was undertaken to investigate the effects of weekly administration of implant type recombinant porcine somatotropin (rpST) on the performance and carcass characteristics in finishing pigs. A total of 120 crossbred (Landrace${\times}$Yorkshire${\times}$Duroc) pigs were employed for 11 weeks in a growth trial in experiment. A rpST designed to implant every 7 d was used. Forty pigs, each weighing 75 kg, were allocated into three rpST treatments; control (CONT), implant of rpST from 75 kg (TRT1) or 90 kg (TRT2) of body weight. The CONT pig and pigs in TRT2 from 75 kg to 90 kg were treated without rpST but with placebo. In rpST-treated pigs, each 100 mg and 125 mg of the equivalent rpST was implanted from live weight of 75 kg to 90 kg and from 90 kg to market weight, respectively. Half of the pigs from each treatment were marketed at live weight of 110 kg and the rest at 130 kg. All pigs were allowed ad libitum access to a commercial feed containing 0.94% and 0.88% of lysine from 75 to 110 kg, 110 to 130 kg of body weights, respectively. rpST had no effect on daily gain, while feed efficiency was improved by 7 to 13% (p<0.05) in the rpST-treated groups compared with the CONT. Compared with the CONT, backfat thickness was decreased by 12% (p<0.05) in TRT1 at 110 kg of market weight, and by 23 to 32% (p<0.05) in the rpST-treated groups at 130 kg of market weight, respectively. Lean muscle rate tended to be higher in TRT1 at both 110 kg and 130 kg of market weight, and carcass fat percentage in the rpST-treated groups was decreased by 33 to 46% (p<0.05) compared with the CONT.

Analysis of Medroxyprogesterone Acetate in Meats (유통 식육 중 성장호르몬제 초산메드록시프로게스테론의 분석)

  • Lee, Yoon-Ae;Shim, Jee-Youn;Lee, Ryun-Kyung;Kim, Seung-Hwan;Oh, Hyun-Sook;Cho, Tae-Yong
    • Journal of Food Hygiene and Safety
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    • v.25 no.3
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    • pp.226-231
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    • 2010
  • We aimed at the monitoring medroxyprogesterone acetate (MPA) residue amount in meats and confirmed the safety of its residue in meats. Optimized condition for analytical and instrumental methods was obtained by method validation. The limit of detection (LOD) and limit of quantification (LOQ) were validated at 1.5 and 5.0 ug/kg, respectively. The calibration curve showed good linearity ($r^2$ = 0.9968) within the concentration range of 5.0~50.0 ug/kg. We selected progesterone-d9 for internal standard, The recoveries in fortified meat ranged from 67.5 to 109.56% at the 3 spiking levels. As the regulation of MPA analysis method used by LC-MS/MS on other products have established. We selected 3 species of farm stock products (cattle, pig, chicken) and purchased at the markets of seven major cities. The total 196 of meat including 46 of domestic beef, 43 of import beef, 60 of domestic pork, 12 of import pork and 35 of domestic chicken. No residue of synthetic growth hormones were detected in cattle, pig and chicken samples tested.

Influences of LH, FSH, EGF and Cysteine on In Vitro Canine Oocyte Maturation (개 난자의 체외성숙에 대한 LH, FSH, EGF 및 Cysteine의 효과)

  • Song, Hye-Jin;Kang, Eun-Ju;Ock, Sun-A;Jeon, Byeong-Gyun;Rho, Gyu-Jin;Choe, Sang-Yong
    • Reproductive and Developmental Biology
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    • v.31 no.3
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    • pp.169-174
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    • 2007
  • Despite many efforts to improving canine in vitro maturation(IVM), the efficiency is still low compared to that of other mammalian species. The present study investigated the effects of gonadotropin, epidermal growth factor and cysteine supplementation on in vitro maturation of canine oocytes. Cumulus-oocyte complexes (COCs) were matured in basic medium (TCM-199 containing 10% FBS, 0.11mg/ml sodium pyruvate supplemented with or without $10{\mu}g/ml$ FSH, 10ng/ml EGF and 0.57mM cysteine) for 72 hr at $38.5^{\circ}C$ in humidified atmosphere of 5% $CO_2$ in air. After culture, oocytes were stained with Hoechst 33342 $(10{\mu}g/ml)$ for 30 min at $4^{\circ}C$, and assessed their nuclear status (GV: germinal vesicle, GVBD: germinal vesicle break down, MI: metaphasse I, MII: metaphase II, UK: unknown stage). No differences were observed in GV, MI and MII rate except GVBD rate between with and without gonadotropins addition respectively (6.7%, vs. 17.2%). Supplementation of 10ng/ml of EGF in hormones added IVM medium resulted in significantly (p<0.05) higher MII rate (4.54% vs.7.06%). Although there are no significantly difference, total of MI and MII rates were increased by adding cysteine. In conclusion, the present study indicates that supplementation of $10{\mu}g/ml$ LH and FSH, 10ng/ml EGF and 0.57mM cysteine in canine IVM medium show a positive influence on the progression of maturation to MII at 72 hr.

Characterization of Somatolactin cDNA from Rock Bream (Oplegnathus fasciatus) (돌돔(Oplegnathus fasciatus) somatolactin cDNA의 분석)

  • 강현실;여인규;이제희
    • Journal of Life Science
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    • v.13 no.6
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    • pp.805-813
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    • 2003
  • cDNA encoding somatolactin (SL) was obtained by RT-PCR from pituitary glands of rock bream (Oplegnathus fasciatus). The full length cDNA of rock bream somatolactin (rbSL) is 1636 bp long. It contains a 696 bp open reading frame encoding a signal peptide of 24 amino acids (an) and a mature protein of 207 aa. rbSL has seven cysteine residues$(Cys^{5},\; Cys^{15},\; Cys^{42},\; Cys^{65},\; Cys^{181},\; Cys^{198}\; $and $Cys^{206})$ and two potential N-glycosylation sites at positions $Asn^{121}$and $Asn^{153}$. The rbSL shares 61.1∼92.6% amino acid sequence similarities and 63∼92.6% nucleotide sequence identities with other teleost SLs, except for goldfish and channel catfish SL. Amino acid sequence alignment revealed that rbSL has four conserved domains $(A_{SL},\; B_{SL},\; C_{SL}and\; D_{SL})$ common to all SLs. Out of these domains, $(A_{SL},\; B_{SL},\; C_{SL}and\; D_{SL})$, are also conserved in all teleost growth hormones and prolactins. The cDNA of rbSL has been cloned into pET expression vector in order to produce recombinant rbSL in E. coli BL2l(DE3) cells. The recombinant protein showed a molecular weight of 27 kDa in SDS-PAGE.

Study of Rat Mammary Epithelial Stem Cells In Vivo and In Vitro (생체 및 시험관에서 유선 상피 모세포의 분리와 동정)

  • Nam Deuk Kim;Kee-Joo Paik
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.24 no.3
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    • pp.470-486
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    • 1995
  • Mammary epithelial cells contain a subpopulation of cells with a large proliferativ potential which are responsible for the maintenance of glandular cellularity and are the progenitor cells of mammary cancer. These clonogens give rise to multicellular clonal alveolar or ductal units(AU or DU) on transplantation and hormonal stimulation. To isolate putative mammary clonogens, enzymatically monodispersed rat mammary epithelial cells from organoid cultures and from intact glands are sorted by flow cytometry according to their affinity for FITC labeled peanut lectin(PNA) and PE labeled anti-Thy-1.1 antibody(Thy-1.1) into four subpopulations : cells negative to both PNA and Thy-1.1(B-), PNA+cells, Thy-1.1+cells, and cells positive to both reagents(B+). The in vivo transplantation assays indicate that the clonogenic fractions of PNA+cells from out-growths of organoids in primary cultures for three days in complete hormone medium(CHM) are significantly higher than those of cells from other subpopulations derived from cultrues or from intact glands. Extracellular matrix(ECM) is a complex of several proteins that regulated cell function ; its role in cell growth and differentiation and tissue-specific gene expression. It can act as a positive as well as a negative regulator of cellular differentiation depending on the cell type and the genes studied. Regulation by ECM is closely interrelated with the action of other regulators of cellular function, such as growth factors and hormones. Matrigel supports the growth and development of several different multicellular colonies from mammary organoids and from monodispersed epithelial cells in culture. Several types of colonies are observed including stellate colonies, duct-like structures, two- and three-dimensional web structures, squamous organoids, and lobulo-duct colonies. Organoids have the greatest proliferative potential and formation of multi-cellular structures. Phase contrast micrographs demonstrate extensive intracellular lipid accumulation within the web structures and some of duct-like colonies. At the immunocytochemical and electron micrograph level, casein proteins are predominantly localized near the apical surface of the cells or in the lumen of duct-like or lobulo-duct colonies. Squamous colonies are comprised of several layers of squamous epithelium surrounding keratin pearls as is typical fo squamous metaplasia(SM). All-trans retinoic acid(RA) inhibits the growth of SM. The frequency of lobulo-ductal colony formation increased with the augmentation of RA concentration in these culture conditions. The current study models could provide powerful tools not only for understanding cell growth and differentiation of epithelial cells, but also for the isolation and characterization of mammary clonogenic stem cells.

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Sesquiterpenoids Bioconversion Analysis by Wood Rot Fungi

  • Lee, Su-Yeon;Ryu, Sun-Hwa;Choi, In-Gyu;Kim, Myungkil
    • 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.

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