• Archives of Pharmacal Research
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• v.19 no.2
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• pp.126-131
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• 1996

The effects of various synthetic benzimidazole derivatives on gastric H^+/K^+$ATPase activity in vitro were examined. The results showed that the effects of substituents on the benzimidazole ring were not significant. However, replacement of sulfoxide connecting two ring systems to sulfide resulted in a completely inactive compound in vitro, suggesting the essential role of sulfoxide group in the inhibition. In addition, compounds with 5 or 6-membered oxacyclic substituents attached to the pyridine ring displayed the most effective inhibitory activity. Among these derivatives, AU-47 was the most potent, and detailed mechanistic studies with the compound were carried out. AU-47 inhibited gastricH^+/K^+$ATPase in a concentration and time dependent manner with 50% inhibition at $6\muM$. The presence of sulfhydryl reducing agents or substrate analogue protected H^+/K^+$ATPase from the inactivation. The inhibition by AU-47 was potentiated by acid pretreatment of the compound, suggesting the structural conversion of AU-47 into a more active intermediate which was favored in acidic condition. Consistent with in vitro results, AU-47 inhibited in vivo gastric acid secretion. The results suggest that AU47 is a relevant candidate for the development of new antiulcer agent.

• Biomolecules & Therapeutics
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• v.2 no.3
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• pp.236-243
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• 1994

A splenocyte culture system supplemented with liver microsomes was developed to detect immunotoxic chemicals which require metabolic activation using cyclophosphamide as a positive standard. When liver microsomes were added to splenocyte cultures isolated from female B6C3Fl mice, the proliferation of splenocytes by lipopolysaccharide (LPS) was increased and the proliferation by concanavalin A (Con A) was decreased. However, when compared with each corresponding control, cyclophophamide was successfully activated to metabolites capable of suppressing Iymphoproliferative responses. This suppression was clearly dependent upon the amounts of microsomes added and/or the concentration of cyclophosphamide exposed. In these cultures, the proliferation of splenocytes was suppressed when the cells were exposed to cyclophosphamide on the day of culture initiation. On the other hand, microsome was responsible for the increase in LPS mitogenicity and NADPH was responsible for the decrease in Con A mitogenicity. Finally, our present culture system was compared with the hepatocyte-splenocyte coculture system which we had developed earlier. We found that the hepatocyte-splenocyte coculture was better able to activate cyclophosphamide to metabolites capable of suppressing the antibody response to sheep erythrocytes. Although our present culture system was relatively poor to activate cyclophosphamide in cultures for antibody response, it will be useful as a simple screening method to detect suppression of certain in vitro immunotoxic parameters like LPS mitogenicity by chemicals which require metabolism.

• Toxicological Research
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• v.25 no.1
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• pp.9-15
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• 2009

Historically, research in toxicology has utilized non-human mammalian species, particularly rats and mice, to study in vivo the effects of toxic exposure on physiology and behavior. However, ethical considerations and the overwhelming increase in the number of chemicals to be screened has led to a shift away from in vivo work. The decline in in vivo experimentation has been accompanied by an increase in alternative methods for detecting and predicting detrimental effects: in vitro experimentation and in silico modeling. Yet, these new methodologies can not replace the need for in vivo work on animal physiology and behavior. The development of new, non-mammalian model systems shows great promise in restoring our ability to use behavioral endpoints in toxicological testing. Of these systems, the zebrafish, Danio rerio, is the model organism for which we are accumulating enough knowledge in vivo, in vitro, and in silico to enable us to develop a comprehensive, high-throughput toxicology screening system.

• Journal of Microbiology and Biotechnology
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• v.9 no.6
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• pp.881-884
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• 1999

The gene encoding the RNA-dependent RNA polymerase of the hepatitis C virus was cloned and expressed with a C-terminal hexahistidine tag. The protein was purified from Escherichia coli to near homogeneity and characterized in vitro. When the 21 amino acids from the C-terminus of the protein were deleted, an inclusion body was not formed and a better purification yield was achieved. However, the activity of the purified enzyme decreased compared to that of the full length protein. The purified enzyme did exhibit ribonucleotide-incorporation activity on an in vitro transcribed RNA containing the 3' end of the HCV genome. It also possessed ribonucleotide incorporation activity, to a lesser extent, on in vitro transcribed foreign RNA templates when RNA or DNA primers were present. The activity was higher with DNA primers than with RNA primers. Accordingly, this assay system will facilitate the screening of inhibitors for hepatitis C virus replication.

• Journal of Forest and Environmental Science
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• v.27 no.2
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• pp.61-72
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• 2011

Micropropagation is an alternative mean of propagation that can be employed in mass multiplication of plants in relatively shorter time. Recent modern techniques of propagation have been developed which could facilitate large scale production of true-to-type plants and for the improvement of the species using genetic engineering techniques in the next century. An overview on the in vitro propagation via meristem culture, regeneration via organogenesis and somatic embryogenesis is presented. The usefulness of the plants in commercial industry as well as propagation techniques, screening for various useful characteristics and the influence of different cultural conditions in the multiplication, rooting and acclimatization phases on the growth of tissue cultured plant discussed.

• Journal of the Korean Society of Food Science and Nutrition
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• v.40 no.7
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• pp.1053-1062
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• 2011

Reactive oxygen species (ROS), including singlet oxygen (${O_2}^1$), superoxide anion radical ($O_2{\cdot}^-$), hydroxyl radical ($HO{\cdot}$), peroxyl radical ($ROO{\cdot}$), hydrogen peroxide ($H_2O_2$), and hypochlorous (HOCl), are generated as byproducts of normal cellular metabolism. ROS induce damage to many biological molecules, such as lipids, proteins, carbohydrates, and DNA. It is widely believed that some degenerative diseases caused by ROS can be prevented by the high intake of fruits and vegetables due to their antioxidant activities. Recently, research on natural antioxidants has become increasingly active in various fields. Several assays have been developed to measure the total antioxidant capacity of antioxidants in fruits and vegetables in vitro. These assays include those for DPPH radical scavenging activity, SOD-like activity, total polyphenol content, oxygen radical absorbance capacity, reducing power, trolox equivalent antioxidant capacity (ABTS assay), single-cell gel electrophoresis (comet assay), and a cellular antioxidant activity assay. Because different antioxidant compounds may act through different mechanisms in vitro, no single assay can fully evaluate the total antioxidant capacity of foods. Due to the complexity of the composition of foods, it is important to be able to measure antioxidant activity using biologically relevant assays. In this review, recently used assays were selected for extended discussion, including a comparison of the advantages and disadvantages of each assay and their application to fruits and vegetables.

• The Plant Pathology Journal
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• v.15 no.1
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• pp.68-71
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• 1999

To illustrate the action mechanism of pencycuron on Rhizoctonia solani, two experiments were conducted including the comparison of amino acids of $\beta$-tubulin between R-C (sensitive isolate) and Rh-131 (non-sensitive isolate), and the inhibitory effect of pencycuron on tubulin assembly in vitro. Both $\beta$-tubulin genes of R-C and Rh-131 proved to have 1,582 nucleotides encoding a protein of 445 amino acids, showing 98% homology in amino acid sequences between them. It was found that codons at 103, 236, and 267 for lysine (AGG), valine (GTC) and isoleucine (ATT) in R-C were replaced by codons for methionine (ATG), isoleucine (ATT) and methionine (ATG) in Rh-131, respectively. No inhibitory effect of pencycuron on the tubulin assembly was observed. It suggests that pencycuron may have no direct inhibitory effects on the assembly of tubulin at least in vitro.

• The Korean Journal of Food And Nutrition
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• v.15 no.4
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• pp.307-313
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• 2002

The primary objective of these studies was to screen the materials showing inhibitions of HMG-CoA reductase in vitro. The secondary objective was to determine the effect of garlic, lovastatin and copper on cholesterol concentrations in plasma, liver and breast tissues in pullets. The degree of inhibition of the selective samples on HMG-CoA reductase activity was determined in vitro. The inhibition ratios of water soluble garlic extracts, lovastatin (methanol extracts) and copper to HMG-CoA reductase activity were 51.3%, 87.5%, and 82.0%, respectively. Control diet (basal diet) and experimental diets, garlic powder (3% in diet), lovastatin (300mg/Kg of diet) and copper (200mg/Kg of diet) were fed to pullets in order to investigate the changes of cholesterol concentration in plasma and tissues. Total cholesterol, HDL- and LDL-cholesterol in blood plasma were significantly reduced in pullets fed diet containing 3% garlic powder. However, copper significantly increased total cholesterol compared to control and lovastatin did not affect plasma cholesterol concentration. Total cholesterol and triglyceride of liver and breast tissues in pullets were not affected by adding the cholesterol-lowering materials to diets. The data suggests that it is not easy for HMG-CoA reductase inhibitors to reduce cholesterol levels in body due to complication of cholesterol metabolism. However, garlic administration can lower the levels of plasma cholesterol in pullets.

• Animal cells and systems
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• v.13 no.3
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• pp.283-288
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• 2009

The ubiquitous plasma membrane $Na^+/H^+$ exchanger 1 (NHE1) is rapidly activated in response to various extracellular stimuli and maintains normal cytoplasmic pH. Yeast two-hybrid screening was used in order to identify proteins interacting with NHE1 using its cytoplasmic domain as a bait from HeLa cDNA library. One of the interacting cDNA clones was human Lactate dehydrogenase B (LDHB). In vitro translated LDHB was pulled down together with GST-NHE1.cd protein in the GST pull down assay, confirming the interaction in vitro. LDHB antibody immunoprecipitated endogenous LDHB together with NHE1 from H9c2 cells, validating cellular interaction between NHE1 and LDHB. Subsequent analysis revealed that the overexpression of LDHB increased intracellular PH, implying opening of the NHE1 transporter. Moreover, overexpression of LDHB activated caspase 3 and induced cell death, consistent with the expected phenotype of hyper-activation of NHE1. Collectively, our data indicate that LDHB modulates NHE1 activity via physical interaction.

• Archives of Pharmacal Research
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• v.19 no.4
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• pp.321-325
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• 1996

Aiming at the development of anticancer agents by modification of phenolic benzo[c]phenanthridine alkaloid, additional hydroxyl group was put on C10 position of fagaridine (1) by a biomimetic synthetic procedure to afford 10-hydroxyfagaridine (12). All of the synthetic intermediates were also screened in vitro antitumor activities against five different cell lines as well as 12. Among them the representative cytotoxic results are shown as follows; P-quinone (11) $[ED_50;(A549=0.22; {\mu}g/ml)$, $(HCT;15=0.21 {\mu}g/ml)$, fagaridine (1) $(HCT;15=0.41 {\mu}g/ml)$, olefin (6) $(HCT; 15=0.06 {\mu}g/ml)$, acetal (7) $(SKMEL-2=0.07 {\mu}g/ml)$, dihydrofagaridne (10) $(A549=0.38 {\mu}g/ml)$, 10-hydroxyfagaridine (12) $(A 549=0.45{\mu}g/mi)$. From these observation three main remarks can be drawn; (i) the iminium part of benzo[c]phenanthridine is not essential for showing acitvities, (ii) the additional hydroxyl group did not contribute to enhance the cytotoxicity, (iii) the 3-arylisoquinolin-1(2H)-one derivatives were found to display significant in vitro antitumor activity.