• Analytical Science and Technology
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• v.28 no.2
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• pp.132-138
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• 2015

Rosiglitazone metabolites in rat plasma were analyzed after intraperitoneal and oral administration to rats. Seven metabolites (M1-M7) were detected in rat plasma (IP and PO), and the structures were confirmed using liquid chromatography-triple time of flight (TOF) mass spectrometry; as a result, the most abundant metabolite was M5, a de-methylated rosiglitazone. Other minor in vivo metabolites were driven from monooxygenation and demethylation (M2), thiazolidinedione ring-opening (M1, M3), mono-oxygenation (M4, M7), and mono-oxygenation followed by sulfation (M6). Among them, M1 was found to be a 3-{p-[2-(N-methyl-N-2-pyridylamino)ethoxy]phenyl}-2-(methylsulfinyl)propionamide, which is a novel metabolite of rosiglitazone. There was no significant difference in the metabolic profiles resulting from the two administrations. The findings of this study provide the first comparison of circulating metabolite profiles of rosiglitazone in rat after IP and PO administration and a novel metabolite of rosiglitazone in rat plasma.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.11
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• pp.1837-1847
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• 2020

Objective: To evaluate the pancreatic differentiation potential of α-1,3-galactosyltransferase knockout (GalTKO) pig-derived bone marrow-derived mesenchymal stem cells (BM-MSCs) using epigenetic modifiers with different pancreatic induction media. Methods: The BM-MSCs have been differentiated into pancreatic β-like cells by inducing the overexpression of key transcription regulatory factors or by exposure to specific soluble inducers/small molecules. In this study, we evaluated the pancreatic differentiation of GalTKO pig-derived BM-MSCs using epigenetic modifiers, 5-azacytidine (5-Aza) and valproic acid (VPA), and two types of pancreatic induction media - advanced Dulbecco's modified Eagle's medium (ADMEM)-based and N2B27-based media. GalTKO BM-MSCs were treated with pancreatic induction media and the expression of pancreas-islets-specific markers was evaluated by real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. Morphological changes and changes in the 5'-C-phosphate-G-3' (CpG) island methylation patterns were also evaluated. Results: The expression of the pluripotent marker (POU class 5 homeobox 1 [OCT4]) was upregulated upon exposure to 5-Aza and/or VPA. GalTKO BM-MSCs showed increased expression of neurogenic differentiation 1 in the ADMEM-based (5-Aza) media, while the expression of NK6 homeobox 1 was elevated in cells induced with the N2B27-based (5-Aza) media. Moreover, the morphological transition and formation of islets-like cellular clusters were also prominent in the cells induced with the N2B27-based media with 5-Aza. The higher insulin expression revealed the augmented trans-differentiation ability of GalTKO BM-MSCs into pancreatic β-like cells in the N2B27-based media than in the ADMEM-based media. Conclusion: 5-Aza treated GalTKO BM-MSCs showed an enhanced demethylation pattern in the second CpG island of the OCT4 promoter region compared to that in the GalTKO BM-MSCs. The exposure of GalTKO pig-derived BM-MSCs to the N2B27-based microenvironment can significantly enhance their trans-differentiation ability into pancreatic β-like cells.

• Korean Journal of Food Science and Technology
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• v.18 no.5
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• pp.364-371
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• 1986

High-ester pectin was demethylated by the treatments of HCl alone and a combination of HCl and $NH_4OH$. The low-ester pectin prepared were analyzed for chemical composition and the pectin gels were evaluated for firmness by sag values and strength by puncture stress. Gels made from HCl demethylated sample showed brittle, weak and poor elastic characteristics while the $HCl-NH_4OH$ treated samples generally resulted in a smooth and elastic gels except those samples having very low content of ester group or acid amide group. Statistical analysis showed that significant correlations were found between sag values and ester content or molecular weight, and puncture stress and ester content, acid amide groups or molecular weight. The equations derived for sag, puncture stress and sag/puncture stress from chemical data could be useful for prediction of some of the physical properties of low-ester pectin gel.

• Proceedings of the Korea Technical Association of the Pulp and Paper Industry Conference
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• 1999.04b
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• pp.254-258
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• 1999

The distribution of anionic groups in the fibers, the fines, the colloidal fraction and the dissolved fraction, respectively, of thermomechanical pulp (TMP) suspensions was determined and peroxide bleaching of spruce TMP were also studied. Spruce TMP was extracted with hexane, treated with alkali, or bleached with peroxide. Suspensions made at pH 5.5 were fractionated into long fibres, large fines, small fines, a colloidal fraction and a dissolved fraction. The charge of the fractions was determined using polyelectrolyte titration. To determined the origin of the charges, the contents of fatty acids, resin acids and acidic units in hemicelluloses in the different fractions were determined by has chromatography. Extraction of TMP with hexane prior to fractionation increased the measured charge of the fibres. The removal of the wood resin probably uncovered some carboxyl groups on the fibre surfaces, or improved th e penetration of polybrene into the pores of the fibres. The charge of the fines and the colloidal fraction was lower when the wood resin had been removed. Alkaline treatment of the TMP increased the charge of the fibres and fines, mainly because of demethylation of pectins. Alkaline treatment increased the charge also of the dissolved fraction, because of the release the charge also of the dissolved fraction, because of the release of pectic acids into the water phase. Alkaline peroxide bleaching further increased the charge of fibres and the dissolved fraction, most likely because of lignin oxidation. The charge of the colloidal fraction, consisting mainly of wood resin, was only slightly affected by alkaline treatment and peroxide bleaching. The anionic groups in TMP suspensions were mainly free uronic acids in the hemicelluloses. The contribution from the fatty and resin acids was substantial only for the colloidal fraction.

• Archives of Pharmacal Research
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• v.27 no.2
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• pp.265-272
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• 2004

The pharmacokinetics of CKD-732 (6-0-4-[dimethyl-aminoethoxy)cinnamoyl]-fumagillolㆍhemioxalate) was investigated in male SD rats and beagle dogs after bolus intravenous administration. The parent compound and metabolites obtained from in vitro and in vivo samples were determined by LC/MS. The main metabolite was isolated and identified as an N-oxide form of CKD-732 by NMR and LC/MS/MS. CKD-732 was metabolized into either M11 or others by rapid hydroxylation, demethylation, and hydrolysis. The blood level following the intravenous route declined in first-order kinetics with $T_{1}$2/$\beta$ values of 0.72-0.78 h for CKD-732 and 0.92-1.09 h for M11 in rats at a dose of 7.5-30 mg/kg. In dogs, $T_{1}$2/$\beta$ values of CKD-732 and M11 were 1.54 and 1.79 h, respectively. Moreover, AUC values increased dose dependently for CKD-732 and M11 in rats and dogs. The CLtot and Vdss did not change significantly with increasing dose, indicating linear pharmacokinetic patterns. The excretion patterns through the urine, bile, and feces were also examined in the animals. The total amount excreted in urine, bile, and feces was 2.13% for CKD-732 and 1.29% for M11 in rats, and 1.58% for CKD-732 and 2.28% for M11 in dogs.

• The Plant Pathology Journal
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• v.36 no.3
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• pp.218-230
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• 2020

Net blotch of barley caused by Pyrenophora teres (Died.) Drechsler, is one of the most destructive diseases on barley in Algeria. It occurs in two forms: P. teres f. teres and P. teres f. maculata. A total of 212 isolates, obtained from 58 fields sampled in several barley growing areas, were assessed for fungicide sensitivity by target gene analysis. F129L and G137R mitochondrial cytochrome b substitution associated with quinone outside inhibitors (QoIs) resistance, and succinate dehydrogenase inhibitors (SDHIs) related mutations (B-H277, C-N75S, C-G79R, C-H134R, and C-S135R), were analyzed by pyrosequencing. In vitro sensitivity of 45 isolates, towards six fungicides belonging to three chemical groups (QoI, demethylase inhibitor, and SDHI) was tested by microtiter technique. Additionally, sensitivity towards three fungicides (azoxystrobin, fluxapyroxad, and epoxiconazole) was assessed in planta under glasshouse conditions. All tested isolates were QoI-sensitive and SDHI-sensitive, no mutation that confers resistance was identified. EC₅₀ values showed that pyraclostrobin and azoxystrobin are the most efficient fungicides in vitro, whereas fluxapyroxad displayed the best disease inhibition in planta (81% inhibition at 1/9 of the full dose). The EC₅₀ values recorded for each form of net blotch showed no significant difference in efficiency of QoI treatments and propiconazole on each form. However, in the case of fluxapyroxad, epoxiconazole and tebuconazole treatments, analysis showed significant differences in their efficiency. To our knowledge, this study is the first investigation related to mutations associated to QoI and SDHI fungicide resistance in Algerian P. teres population, as well as it is the first evaluation of the sensitivity of P. teres population towards these six fungicides.

• Journal of Environmental Analysis, Health and Toxicology
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• v.21 no.4
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• pp.243-251
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• 2018

Pharmaceuticals in wastewater effluents have been recognized as emerging pollutants threatening freshwater organisms. To extend understanding for bioaccumulation and toxicity in those organisms, information on biotransformation products (or metabolites) and their metabolic pathway are crucial. The aim of the present study is to identify and elucidate metabolites of pharmaceuticals formed in exposed organisms using suspect and nontarget screening approach using LC-HRMS. As the target pharmaceuticals, carbamazepine, ketoprofen, metoprolol, propranolol, and verapamil were selected whereas Daphnia magna and Gammarus pulex were used as test organisms. After 24h exposure, metabolites formed in the organisms were identified using LC-HRMS. The structures of metabolites were elucidated via analysis of MS/MS fragment pattern and the comparison with fragment database. As the results, a total of 10 metabolites were identified for 5 parent compounds (C253/C356 for carbamazepine, K211 for ketoprofen, M256 for metoprolol, P218/P276/P306 for propranolol, V196/V291/V441 for verapamil). Among them, the presence of C253 and V291 was confirmed using standard materials. Most of the identified metabolites were formed through oxidative reactions such as hydroxylation, N-demethylation, and dealkylation. Cysteine conjugation (phase II reaction) metabolite (C356) for carbamazepine was found in daphnia. The metabolic pathway of verapamil showed similar metabolic pathways and metabolic pathways for both species. Although the toxicological information on the identified metabolites could not be confirmed, the molecular structure information of the proposed metabolites can be used for future evaluation and prediction of toxicity.

• Journal of Plant Biotechnology
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• v.35 no.2
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• pp.133-140
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• 2008

The modification of DNA and histone plays an important role for gene expression in plant development. The objective of this research is to observe the effects of methylation on the gene expression during dedifferentiation from rice mature seeds to callus and differentiation from callus to shoots. The embryogenic callus with ability to shoot regeneration was not induced on the N6A medium supplemented with 5-azacytidine and abnormal callus with brown color was formed. When the normal rice callus was placed on the regeneration MSRA medium supplemented with 5-azacytidine, the shoot regeneration was inhibited. The results showed that 5-azacytidine, DNA demethylating agent, had negative effects on normal embryogenic callus formation and shoot regeneration. This suggested that DNA methylation of some genes was required for normal cell dedifferentiation and differentiation in tissue culture. The microarray and $GeneFishig^{TM}$ DEG screening were used to observe the gene transcript profile in callus induction and regeneration on N6A (N6 medium + 5-azaC) and MSRA (MS regeneration medium + 5-azaC). Subsets of genes were up-regulated or down-regulated in response to 5-azaC treatments. The genes related with epigenetic regulation, electron transport, nucleic acid metabolism and response to stress were up and down regulated. The different expression of some genes (germin like protein etc.) during callus induction and shoot regeneration was confirmed using RT-PCR and northern blot analysis.

• Biomolecules & Therapeutics
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• v.23 no.2
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• pp.189-194
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• 2015

P450 1A2 is responsible for the metabolism of clinically important drugs and the metabolic activation of environmental chemicals. Genetic variations of P450 1A2 can influence its ability to perform these functions, and thus, this study aimed to characterize the functional significance of three P450 1A2 allelic variants containing nonsynonymous single nucleotide polymorphisms (P450 $1A2^*8$, R456H; $^*15$, P42R; $^*16$, R377Q). Variants containing these SNPs were constructed and the recombinant enzymes were expressed and purified in Escherichia coli. Only the P42R variant displayed the typical CO-binding spectrum indicating a P450 holoenzyme with an expression level of ~ 170 nmol per liter culture, but no P450 spectra were observed for the two other variants. Western blot analysis revealed that the level of expression for the P42R variant was lower than that of the wild type, however the expression of variants R456H and R377Q was not detected. Enzyme kinetic analyses indicated that the P42R mutation in P450 1A2 resulted in significant changes in catalytic activities. The P42R variant displayed an increased catalytic turnover numbers ($k_{cat}$) in both of methoxyresorufin O-demethylation and phenacetin O-deethylation. In the case of phenacetin O-deethylation analysis, the overall catalytic efficiency ($k_{cat}/K_m$) increased up to 2.5 fold with a slight increase of its $K_m$ value. This study indicated that the substitution P42R in the N-terminal proline-rich region of P450 contributed to the improvement of catalytic activity albeit the reduction of P450 structural stability or the decrease of substrate affinity. Characterization of these polymorphisms should be carefully examined in terms of the metabolism of many clinical drugs and environmental chemicals.

• Journal of Microbiology and Biotechnology
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• v.22 no.12
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• pp.1659-1664
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• 2012

TSAHC [4'-(p-toluenesulfonylamido)-4-hydroxychalcone] is a promising antitumorigenic chalcone compound, especially against TM4SF5 (four-transmembrane L6 family member 5)-mediated hepatocarcinoma. We evaluated the potential of TSAHC to inhibit the catalytic activities of nine cytochrome P450 isoforms and of P-glycoprotein (P-gp). The abilities of TSAHC to inhibit phenacetin O-deethylation (CYP1A2), coumarin 6-hydroxylation (CYP2A6), bupropion hydroxylation (CYP2B6), amodiaquine N-deethylation (CYP2C8), diclofenac 4-hydroxylation (CYP2C9), omeprazole 5-hydroxylation (CYP2C19), dextromethorphan O-demethylation (CYP2D6), chlorzoxazone 6-hydroxylation (CYP2E1), and midazolam 1'-hydroxylation (CYP3A) were tested using human liver microsomes. The P-gp inhibitory effect of TSAHC was assessed by [$^3H$]digoxin accumulation in the LLCPK1-MDR1 cell system. TSAHC strongly inhibited CYP2C8, CYP2C9, and CYP2C19 isoform activities with $K_i$ values of 0.81, 0.076, and $3.45{\mu}M$, respectively. It also enhanced digoxin accumulation in a dose-dependent manner in the LLCPK1-MDR1 cells. These findings indicate that TSAHC has the potential to inhibit CYP2C isoforms and P-gp activities in vitro. TSAHC might be used as a nonspecific inhibitor of CYP2C isoforms based on its negligible inhibitory effect on other P450 isoforms such as CYP1A2, CYP2A6, CYP2B6, CYP2D6, CYP2E1, and CYP3A.