• Journal of Plant Biotechnology
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• v.30 no.2
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• pp.207-214
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• 2003

This study was carried out to establish a novel bioassay system for screening of compounds affecting aromatic amino acid or anthocyanin biosynthesis through investigating a degree of sucrose-induced anthocyanin formation such as size of plant material, buffer conditions, light intensity and irradiated duration, incubation temp., etc were determined and standard procedure (suitable experimental condition) was set up as follows. The second leaf blade of white corn seeding induced by fluridone treatment were segmented into a size of 5${\times}$5 min. The segments were floated on the solution of 1% sucrose in 1.0mM MES buffer (pH6.0∼6.5) and incubated at 26$^{\circ}C$ for 2days under the continuous light condition(70∼100$\mu$mol m$^{-2}$ s$^{-1}$ ). Anthocyanin in the purpled tissues was extracted with methanol containing 1% HCl and the optical density of the clear supematants was determined at 528mm. Influences of some chemicals were tested using this system. Glyphosate, 5-enolpyruvylshikimate 3-phosphate synthase inhibitor, showed most sensitive response with I$_{50}$ value at 3.3$\mu$M. Dicyclohexylcarbodiimide(DCCD) and parachloromercuribenzenesulfonic acid(PCMBS) had a relatively strong ingibition with I50 value at 7.1$\mu$M and 10.2$\mu$M, respectively. These results show that sucrose-induced anthocyanin formation in white com leaf segment provide a very simple and rapid system for searching new compounds affecting aromatic amino acid or anteocyanin biosynthesis by screening at less than 10$\mu$M.

• Journal of the Korean Chemical Society
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• v.42 no.2
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• pp.214-219
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• 1998

Dynorphin B analogues, $[Arg^{11}, D-Ala^{12}]$dynorphin B, $[D-Ala^2, Ala^6, Arg^{11}, D-Ala^{12}]$dynorphin B, and dynorphin B (1-11) were synthesized by solid-phase method. A chloromethylated polystyrene resin cross-linked with 2% divinylbenzene was substituted with Thr in ethanol to contain 1.20 mmol Thr/g of resin. All amino groups of amino acids were protected with t-Boc group and 2,6-dichlorobenzyl and nitro groups were used to protect the side chains of Tyr and Arg, respectively. Stepwise synthetic method was applied for synthesis of the products. Dicyclohexylcarbodiimide (DCC) and 1-hydroxybenzotriazole (HOBT) were used as the coupling reagents. The crude peptides were purified by gel filteration on Sephadex LH-20 column $(2 \times 50 cm)$ using MeOH/MeCN (3/1) and then characterized with HPLC, amino acid analyzer.

• Journal of Radiopharmaceuticals and Molecular Probes
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• v.5 no.1
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• pp.18-25
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• 2019

2-Nitroimidazole derivatives have been reported to accumulate in hypoxic tissue. We prepared a novel $^{99m}Tc-MAG_3$-2-nitroimidazole and evaluated the feasibility for hypoxia imaging agent. $Bz-MAG_3$-2-nitroimidazole was synthesized by direct coupling of $Bz-MAG_3$ and 2-nitroimidazole using dicyclohexylcarbodiimide. $Bz-MAG_3$-2-nitroimidazole was labeled with $^{99m}Tc$ in the presence of tartaric acid and $SnCl_2-2H_2O$ at $100^{\circ}C$ for 30 min. And the reaction mixture was purified by $C_{18}$ Sep-pak cartridge. The labeling efficiency and the radiochemical purity were checked by ITLC-SG/acetonitrile. The tumor was grown in balb/c mice for 8~13 days after the subcutaneous injection of tumor cells, CT-26 (murine colon adenocarcinoma cell). Biodistribution study and tumor autoradiography were performed in the xenografted mice after i.v injection of 74 kBq/0.1 mL and 19 MBq/0.1 mL of $^{99m}Tc-MAG_3$-2-nitroimidazole, respectively. In vivo images of $^{99m}Tc-MAG_3$-2-nitroimidazole in tumor bearing mice were obtained 1.5 hr post injection. The labeling efficiency was $45{\pm}20%$ and the radiochemical purity after purification was over 95%. Paper electrophoresis confirmed negative charge of $^{99m}Tc-MAG_3$-2-nitroimidazole. $^{99m}Tc-MAG_3$-2-nitroimidazole was very stable at room temperature and its protein binding was 53%. The $^{99m}Tc-MAG_3$-2-nitroimidazole exhibited high uptake in the liver, stomach and intestine. In biodistribution study using tumor bearing mice, the uptakes (% ID/g) of the tumor were $0.5{\pm}0.1$, $0.4{\pm}0.0$, $0.2{\pm}0.1$ and $0.1{\pm}0.1$ at 5, 15, 30 min and 4 hrs. Tumor/muscle ratio were $1.4{\pm}0.1$, $2.2{\pm}0.83$, $3.0{\pm}0.9$, and 3.7 (n=2) for 5, 15, 30 min and 4 hrs. The uptake in hypoxic area was found higher than in non-hypoxic area of tumor tissue by autoradiography. In vivo images showed the relatively faint uptake to the hypoxic tumor region. $^{99m}Tc-MAG_3$-2-nitroimidazole was successfully synthesized and found feasible for imaging hypoxia.

• Journal of Pharmaceutical Investigation
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• v.26 no.2
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• pp.105-112
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• 1996

The surface of albumin microspheres was modified with methotrexate(MTX) by using 1,3-dicyclohexylcarbodiimide (DCC). Surface-modified albumin microspheres entrapping no MTX (SAMS), free MTX (SAMSF) and MTX-bovine serum albumin(BSA) conjugates(SAMSC) were prepared. The organ-targeting ability of free $[^3H]MTX,\;[^3H]MTX-BSA$ conjugate and the above microspheres was evaluated after i.v. administration of the preparations, equivalent to 150 nCi via the tail vein of mice. The total radioactivity in the lung increased immediately in a few minutes after i.v. injection of the microspheres, and then declined for the period of 3-4 weeks. However, the radioactivity in the liver, spleen and kidney increased slowly during the rapid decrease in radioactivity in the lung. This suggested that the microspheres could be entrapped rapidly in the lung through mechanical filtration because of their large size and slowly redistributed to the liver, spleen and kidney due to either the microspheres being degraded enough for the size to allow passage through the capillary beds of the lung and/or the release of $[^3H]MTX\;or\;[^3H]MTX-BSA$ conjugates from the microspheres. The amount of $60{sim}70%$ of the dose was targeted to the liver after the i.v. injection of SAMS, SAMSF and SAMSC, and the values of $(R_e\;^*\;_{e)liver}$ from the microspheres were $5{\sim}7$ compared to free MTX. This suggested that the liver-targeting ability from surface-modified albumin microspheres could be $5{\sim}7$ times as that of free MTX. The liver-targeted drug was accumulated in the Kupffer cells at the initial stage, thereafter the drug in the Kupffer cell was slowly transferred into the hepatocytes. The value of AUQ for liver from SAMS was higher than that from SAMSF, but much lower than that from SAMSC. This suggest that MTX bound to their surface could be eliminated slower than the entrapped free MTX, and faster than the entrapped MTX-BSA conjugates. This is consistent with the in vitro release rates order in the presence of a proteolytic enzyme. Also, surface-modified MTX was scarcely released in the absence of a proteolytic enzyme. Therefore, the surface-modified MTX nay be released (or eliminated) rapidly from SAMSC at the target site, and thereafter MTX may be released (or eliminated) slowly from the entrapped MTX-BSA conjugates in SAMSC for a long period.