• Title/Summary/Keyword: binding activity

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Synthesis and Evaluation of $2-[^{18}F]Fluoro-A85380$, a Radioligand for ${\alpha}_4{\beta}_2$ Nicotinic Acetylcholine Receptor Imaging (${\alpha}_4{\beta}_2$ 니코틴성 아세틸콜린 수용체 영상 방사성리간드 $2-[^{18}F]fluoro-A85380$의 합성 및 평가)

  • Ryu, Eun-Kyoung;Choe, Yearn-Seong;Kim, Sang-Eun;Hwang, Sae-Hwan;Paik, Jin-Young;Choi, Yong;Lee, Kyung-Han;Kim, Byung-Tae
    • The Korean Journal of Nuclear Medicine
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    • v.36 no.4
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    • pp.261-270
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    • 2002
  • Purpose: Nicotinic acetylcholine receptors (nAChRs), which mediate excitatory neurotransmission, are known to participate in various neurophysiological functions. Severe losses of nAChRs have been noted in Alzheimer's and Parkinson's diseases. Therefore, noninvasive and quantitative imaging of nAChRs would offer a better understanding on the function of these receptors. In this study, $2-[^{18}F]fluoro-A85380\;([^{18}F]1)$, an ${\alpha}_4{\beta}_2$ nAChRs radioligand, was prepared using one HPLC purification and evaluated in mouse brain, and the results were compared with those in the literature. Materials and Methods: $[^{18}F]1$ was prepared by $[^{18}F]$fluorination of the iodo precursor followed by acidic deprotection and then purified by HPLC. Tissue distribution studies were performed in mouse brain at the indicated time points and the result was expressed as %ID/g. Inhibition studies were also carried out with pretreatment of various ligands. Results: One HPLC purification method gave the desired product in 15-20% radiochemical yield and with high specific activity ($38-55GBq/{\mu}mol$). Tissue distribution studies showed that $[^{18}F]1$ specifically labeled nAChRs in mouse brain with a high thalamus to cerebellum uptake ratio (13.8 at 90 min). Inhibition studios demonstrated selective binding of $[^{18}F]1$ to nAChRs, blocking the uptake of the $[^{18}F]1$ in nAChR-rich legions by selective ligands such as cytisine and nicotine which are well-known nAChRs agonists. Conclusion: This study demonstrated that the $[^{18}F]1$ produced by the method using one HPLC purification gave the results similar to those reported in the literature. Therefore, this synthetic method can be readily applied to the routine preparation of $[^{18}F]1$, a PET radioligand for ${\alpha}_4{\beta}_2$ nAChRs imaging.

The Antioxidant Ability and Nitrite Scavenging Ability of Plant Extracts (식물체 추출물의 항산화성 및 아질산염 소거작용)

  • Kim, Soo-Min;Cho, Young-Suk;Sung, Sam-Kyung
    • Korean Journal of Food Science and Technology
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    • v.33 no.5
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    • pp.626-632
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    • 2001
  • The plant extracted from Nameko, Gallic, Green tea, Allspice, Polygonum multiflorum, Schizandra chinensis, Armeniacae and Pine needle were utilized to investigate the effects of extracts on free radical reaction, lipid oxidation and nitrite scavenging ability. The pH of ethanol extracts showed a higher than that of hot water extracts, among of which were showed the lowest pH 3.0 in Schizandra chinensis. The important factor of lipid oxidation were $Fe^{2+}$ ion and active oxygen, in which were bound by plant extracts in case of $Fe^{2+}$ ion existed. However, the hydroxyl radical scavenging ability of extracts were lowed, compared to extracts reacted with $Fe^{2+}$ ion. Among of them, the hydroxyl radical scavenging ability of Nameko and Pine needle extracts had a lower TBARS value than those of control. The iron content of extracts were less than 2.0 mg/100 g, but the total iron content of Schizandra chinensis extracts were 6.8 mg/100 g. The ethanol extracts of pine needle were higher than those of hot water extracts on the basis of $Fe^{2+}$ ion content. The ascorbic acid content of green tea showed 14.3 mg/100 g in hot water extracts and 16.7 mg/100 g in ethanol extracts. Electron donating ability of extracts showed more than 50%, except Nameko and allspices, which were higher in ethanol extracts than those of hot water extracts. The superoxide dismutase(SOD)-like activity of green tea showed 85.3% and 63.5% in hot water and ethanol extracts, respectively. The nitrite scavenging ability of green tea was the most effective in both extracts.

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A Study on the Whitening Effect of Erigeron annuus (L.) Pers. Ethanol Extract on Melanoma Cell (B16F10) (멜라노마 세포(B16F10)에서의 개망초 추출물을 이용한 미백효과에 관한 연구)

  • Joo, Da-Hye;Yoo, Dan-Hee;Lee, Jin-Young
    • Microbiology and Biotechnology Letters
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    • v.47 no.1
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    • pp.148-157
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    • 2019
  • A 70% ethanol extract of Erigeron annuus (L.) Pers. was investigated for its whitening activity for application as a functional ingredient in cosmetic products. At the E. annuus extract concentration of $100{\mu}g/ml$, the electron-donating ability was found to be 67.83%, the tyrosinase inhibitory effect (related to skin-whitening) was 69%, the elastase inhibitory effect (related to skin-wrinkling) was 69%, and the astringent effect was 80%. The $ABTS^+$ radical-scavenging ability was 87% at the $500{\mu}g/ml$ concentration. In the cell viability test measured on melanoma cells, 96% of the cells treated with $100{\mu}g/ml$ of the extract were viable. According to the western blot results, the protein expression of the microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein (TRP)-1, and TRP-2 was decreased by 60.22%, 47.83%, 54.79%, and 67.88%, respectively, at the extract concentration of $100{\mu}g/ml$. The protein expression of phosphorylated extracellular signal regulated kinase (p-ERK) and phosphorylated cAMP response element-binding protein (p-CREB) was decreased with increasing concentrations of the extract. Reverse transcription-polymerase chain reaction of the extract showed that the mRNA expression of MITF, tyrosinase, TRP-1, and TRP-2 was decreased by 86.51%, 85.22%, 74.26%, and 66.66%, respectively, at $100{\mu}g/ml$ extract concentration. The findings suggest that the 70% ethanol extract from E. annuus (L.) Pers. has potential as a cosmeceutical ingredient with whitening effect.

Optimization and Stabilization of Automated Synthesis Systems for Reduced 68Ga-PSMA-11 Synthesis Time (68Ga-PSMA-11 합성 시간 단축을 위한 자동합성장치의 최적화 및 안정성 연구)

  • Ji hoon KANG;Sang Min SHIN;Young Si PARK;Hea Ji KIM;Hwa Youn JANG
    • Korean Journal of Clinical Laboratory Science
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    • v.56 no.2
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    • pp.147-155
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    • 2024
  • Gallium-68-prostate-specific membrane antigen-11 (68Ga-PSMA-11) is a positron emission tomography radiopharmaceutical that labels a Glu-urea-Lys-based ligand with 68Ga, binding specifically to the PSMA. It is used widely for imaging recurrent prostate cancer and metastases. On the other hand, the preparation and quality control testing of 68Ga-PSMA-11 in medical institutions takes over 60 minutes, limiting the daily capacity of 68Ge/68Ga generators. While the generator provides 1,110 MBq (30 mCi) nominally, its activity decreases over time, and the labeling yield declines irregularly. Consequently, additional preparations are needed, increasing radiation exposure for medical technicians, prolonging patient wait times, and necessitating production schedule adjustments. This study aimed to reduce the 68Ga-PSMA-11 preparation time and optimize the automated synthesis system. By shortening the reaction time between 68Ga and the PSMA-11 precursor and adjusting the number of purification steps, a faster and more cost-effective method was tested while maintaining quality. The final synthesis time was reduced from 30 to 20 minutes, meeting the standards for the HEPES content, residual solvent EtOH content, and radiochemical purity. This optimized procedure minimizes radiation exposure for medical technicians, reduces patient wait times, and maintains consistent production schedules, making it suitable for clinical application.

Effects of Motion Correction for Dynamic $[^{11}C]Raclopride$ Brain PET Data on the Evaluation of Endogenous Dopamine Release in Striatum (동적 $[^{11}C]Raclopride$ 뇌 PET의 움직임 보정이 선조체 내인성 도파민 유리 정량화에 미치는 영향)

  • Lee, Jae-Sung;Kim, Yu-Kyeong;Cho, Sang-Soo;Choe, Yearn-Seong;Kang, Eun-Joo;Lee, Dong-Soo;Chung, June-Key;Lee, Myung-Chul;Kim, Sang-Eun
    • The Korean Journal of Nuclear Medicine
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    • v.39 no.6
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    • pp.413-420
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    • 2005
  • Purpose: Neuroreceptor PET studies require 60-120 minutes to complete and head motion of the subject during the PET scan increases the uncertainty in measured activity. In this study, we investigated the effects of the data-driven head mutton correction on the evaluation of endogenous dopamine release (DAR) in the striatum during the motor task which might have caused significant head motion artifact. Materials and Methods: $[^{11}C]raclopride$ PET scans on 4 normal volunteers acquired with bolus plus constant infusion protocol were retrospectively analyzed. Following the 50 min resting period, the participants played a video game with a monetary reward for 40 min. Dynamic frames acquired during the equilibrium condition (pre-task: 30-50 min, task: 70-90 min, post-task: 110-120 min) were realigned to the first frame in pre-task condition. Intra-condition registrations between the frames were performed, and average image for each condition was created and registered to the pre-task image (inter-condition registration). Pre-task PET image was then co-registered to own MRI of each participant and transformation parameters were reapplied to the others. Volumes of interest (VOI) for dorsal putamen (PU) and caudate (CA), ventral striatum (VS), and cerebellum were defined on the MRI. Binding potential (BP) was measured and DAR was calculated as the percent change of BP during and after the task. SPM analyses on the BP parametric images were also performed to explore the regional difference in the effects of head motion on BP and DAR estimation. Results: Changes in position and orientation of the striatum during the PET scans were observed before the head motion correction. BP values at pre-task condition were not changed significantly after the intra-condition registration. However, the BP values during and after the task and DAR were significantly changed after the correction. SPM analysis also showed that the extent and significance of the BP differences were significantly changed by the head motion correction and such changes were prominent in periphery of the striatum. Conclusion: The results suggest that misalignment of MRI-based VOI and the striatum in PET images and incorrect DAR estimation due to the head motion during the PET activation study were significant, but could be remedied by the data-driven head motion correction.

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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Correlation of p53 Protein Overexpression, Gene Mutation with Prognosis in Resected Non-Small Cell Lung Cancer(NSCLC) Patients (비소세포폐암에서 p53유전자의 구조적 이상 및 단백질 발현이 예후에 미치는 영향)

  • Lee, Y.H.;Shin, D.H.;Kim, J.H.;Lim, H.Y.;Chung, K.Y.;Yang, W.I.;Kim, S.K.;Chang, J.;Roh, J.K.;Kim, S.K.;Lee, W.Y.;Kim, B.S.;Kim, B.S.
    • Tuberculosis and Respiratory Diseases
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    • v.41 no.4
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    • pp.339-353
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    • 1994
  • Background : The p53 gene codes for a DNA-binding nuclear phosphoprotein that appears to inhibit the progression of cells from the G1 to the S phase of the cell cycle. Mutations of the p53 gene are common in a wide variety of human cancers, including lung cancer. In lung cancers, point mutations of the p53 gene have been found in all histological types including approximately 45% of resected NSCLC and even more frequently in SCLC specimens. Mutant forms of the p53 protein have transforming activity and interfere with the cell-cycle regulatory function of the wild-type protein. The majority of p53 gene mutations produce proteins with altered conformation and prolonged half life; these mutant proteins accumulate in the cell nucleus and can be detected by immunohistochemical staining. But protein overexpression has been reported in the absence of mutation. p53 protein overexpression or gene mutation is reported poor prognostic factor in breast cancer, but in lung cancer, its prognostic significance is controversial. Method : We investigated the p53 abnormalities by nucleotide sequencing, polymerase chain reaction-single strand conformation polymorphism(PCR-SSCP), and immunohistochemical staining. We correlated these results with each other and survival in 75 patients with NSCLC resected with curative intent. Overexpression of the p53 protein was studied immunohistochemically in archival paraffin- embedded tumor samples using the D07(Novocastra, U.K.) antibody. Overexpression of p53 protein was defined by the nuclear staining of greater than 25% immunopositive cells in tumors. Detection of p53 gene mutation was done by PCR-SSCP and nucleotide sequencing from the exon 5-9 of p53 gene. Result: 1) Of the 75 patients, 36%(27/75) showed p53 overexpression by immunohistochemical stain. There was no survival difference between positive and negative p53 immunostaining(overall median survival of 26 months, disease free median survival of 13 months in both groups). 2) By PCR-SSCP, 27.6%(16/58) of the patients showed mobility shift. There was no significant difference in survival according to mobility shift(overall median survival of 27 in patients without mobility shift vs 20 months in patients with mobility shift, disease free median survival of 8 months vs 10 months respectively). 3) Nucleotide sequence was analysed from 29 patients, and 34.5%(10/29) had mutant p53 sequence. Patients with the presence of gene mutations showed tendency to shortened survival compared with the patients with no mutation(overall median survival of 22 vs 27 months, disease free median survival of 10 vs 20 months), but there was no statistical significance. 4) The sensitivity and specificity of immunostain based on PCR-SSCP was 67.0%, 74.0%, and that of the PCR-SSCP based on the nucleotide sequencing was 91.8%, 96.2% respectively. The concordance rate between the immunostain and PCR-SSCP was 62.5%, and the rate between the PCR-SSCP and nucleotide sequencing was 95.3%. Conclusion : In terms of detection of p53 gene mutation, PCR-SSCP was superior to immunostaining. p53 gene abnormalities either overexpression or mutation were not a significant prognostic factor in NSCLC patients resected with curative intent. However, patients with the mutated p53 gene showed the trends of early relapse.

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