Preparation of a cyclic polyphenylene array for a zigzag-type carbon nanotube segment.

Preparation of cyclic polyphenylene array 2, which corresponds to a complete carbon array of a zigzag-type CNT segment with (18,0)-structure, has been established by a Diels-Alder reaction of cyclic biphenylylene-acetylene derivative 1 with tetraphenylcyclopentadienone. The reaction of 2 with excess FeCl3 realized a presumed cyclodehydrogenation reaction and elimination of the alkyl chains that were introduced as a measure to counter the low solubility problem, but this resulted in the formation of a complicated mixture that included the mass region of a presumed zigzag-type CNT segment with (18,0)-structure. The rather efficient blue emission of cyclic compounds 1 and 2 was discussed utilizing fluorescence (FL) quantum efficiencies (Φ(FL)) and lifetimes (τ(FL)) in their crystalline state along with those in dichloromethane solution. Thermal analyses of these compounds revealed their characteristic phase transition behavior. The synthesis of a novel cyclic polyphenylene array by utilizing a Diels-Alder reaction of cyclic phenylene-acetylene compounds with tetraphenylcyclopentadienone should afford an attractive pathway to a novel belt-shaped CNT segment.

Image-based quantitative determination of DNA damage signal reveals a threshold for G2 checkpoint activation in response to ionizing radiation.

BACKGROUND: Proteins involved in the DNA damage response accumulate as microscopically-visible nuclear foci on the chromatin flanking DNA double-strand breaks (DSBs). As growth of ionizing radiation (IR)-induced foci amplifies the ATM-dependent DNA damage signal, the formation of discrete foci plays a crucial role in cell cycle checkpoint activation, especially in cells exposed to lower doses of IR. However, there is no quantitative parameter for the foci which considers both the number and their size. Therefore, we have developed a novel parameter for DNA damage signal based on the image analysis of the foci and quantified the amount of the signal sufficient for G2 arrest. RESULTS: The parameter that we have developed here was designated as SOID. SOID is an abbreviation of Sum Of Integrated Density, which represents the sum of fluorescence of each focus within one nucleus. The SOID was calculated for individual nucleus as the sum of (area (total pixel numbers) of each focus) x (mean fluorescence intensity per pixel of each focus). Therefore, the SOID accounts for the number, size, and fluorescence density of IR-induced foci, and the parameter reflects the flux of DNA damage signal much more accurately than foci number. Using very low doses of X-rays, we performed a "two-way" comparison of SOID of Ser139-phosphorylated histone H2AX foci between G2-arrested cells and mitosis-progressing cells, and between mitosis-progressing cells in the presence or absence of ATM or Chk1/2 inhibitor, both of which abrogate IR-induced G2/M checkpoint. The analysis revealed that there was a threshold of DNA damage signal for G2 arrest, which was around 4000~5000 SOID. G2 cells with < 4000 SOID were neglected by G2/M checkpoint, and thus, the cells could progress to mitosis. Chromosome analysis revealed that the checkpoint-neglected and mitosis-progressing cells had approximately two chromatid breaks on average, indicating that 4000~5000 SOID was equivalent to a few DNA double strand breaks. CONCLUSIONS: We developed a novel parameter for quantitative analysis of DNA damage signal, and we determined the threshold of DNA damage signal for IR-induced G2 arrest, which was represented by 4000~5000 SOID. The present study emphasizes that not only the foci number but also the size of the foci must be taken into consideration for the proper quantification of DNA damage signal.

Constitutive expression of bergaptol O-methyltransferase in Glehnia littoralis cell cultures.

We investigated whether exogenously supplied precursors of bergapten, namely umbelliferone, psoralen and bergaptol, could be utilized to produce bergapten without elicitation in Glehnia littoralis cell suspension cultures. The levels of added psoralen and bergaptol in the medium soon decreased, and this was followed by the detection of bergapten in both culture fluid and cells. Umbelliferone was also incorporated but in this case no bergapten was produced; instead, skimmin, umbelliferone monoglucoside, was detected. To determine whether conversion of psoralen to bergapten was due to enzyme induction by precursor feeding, the transcript accumulations and enzyme activities of bergaptol O-methyltransferase (BMT, EC 2.1.1.69), which catalyzes the last step of bergapten synthesis, and of phenylalanine ammonia-lyase (PAL, EC 4.3.1.5), which catalyzes the initial step of the phenylpropanoid biosynthetic pathway and is known as a marker enzyme of elicitation, were examined. The results showed that both the expression and the activity of BMT were always detected in all cells, including control cells. Since PAL was slightly induced in the cells supplied with/without precursors, phenylethyl alcohol (PEA, a competitive inhibitor of PAL) was applied to suspension cells prior to the addition of psoralen. PAL activity was effectively inhibited by PEA at 1-5 mM concentrations. Under these conditions, PEA did not affect bergapten production by cell cultures fed with psoralen at all. These results demonstrate that BMT is constitutively expressed in G. littoralis cell cultures.

Ascorbic acid induces furanocoumarin production in organ cultures of Glehnia littoralis.

Exogenously supplied ascorbic acid (AsA) strongly induced furanocoumarin production in leaf and root cultures of GLEHNIA LITTORALIS, but not in cell suspension cultures, after 24 h of treatment. The dose dependency showed that both organ tissues responded well to AsA supplied at concentrations of 10 - 40 mM. For induction of furanocoumarin production, roots required contact with AsA for at least 6 h and productivity markedly increased after 8 h of treatment. This is the first report of the induction of furanocoumarin biosynthesis by AsA alone and of the detection of furanocoumarin biosynthesis in a root culture system.

Post-stress metabolism involves umbelliferone production in anthocyanin-producing and nonproducing cells of Glehnia littoralis suspension cultures.

The effects of yeast extract on the accumulation of transcripts of phenylalanine ammonia-lyase (PAL, EC 4.1.3.5) and chalcone synthase (CHS, EC 2.3.1.74), PAL and CHS enzyme activity and furanocoumarin and anthocyanin metabolites over a 48 h period were studied in anthocyanin-producing (Violet) and non-producing (White) cell suspension cultures of Glehnia littoralis. In the course of this period, umbelliferone, which had not been detected earlier, was detected in the culture medium of the Violet as well as White cells. In White cells, the PAL transcript accumulation and an increase in PAL activity were in good agreement with the level of umbelliferone, and was followed by the induction of bergapten. In the case of the Violet cells, the accumulation of PAL and CHS transcripts, and the increases in PAL and CHS enzyme activity as well as the anthocyanin level, all of which were highly expressed in nontreated cells, were temporarily suppressed. However, the suppression of the PAL transcript and PAL activity was not as great as that of the CHS transcript accumulation and CHS activity, in which a sharp transient increase of umbelliferone production soon after elicitation appears to be a factor.

Diurnal variations in human urinary excretion of nicotinamide catabolites: effects of stress on the metabolism of nicotinamide.

BACKGROUND: More than 500 enzymes need niacin coenzymes. Therefore, elucidation of the control mechanisms of coenzyme metabolism is fundamentally important. OBJECTIVE: NAD(+) is involved in ATP production. Because energy expenditure is generally higher during the day than at night, we investigated whether the metabolism of nicotinamide changes at various times of day and whether stress affects nicotinamide metabolism. DESIGN: Twelve women were housed in the same facility and followed the same schedule for activities of daily living for 12 d. Urinary outputs were collected during 5 specific periods to investigate diurnal variations in nicotinamide metabolism. The effects of cold exposure (physical stress), having to perform arithmetic calculations (mental stress), and dark exposure (emotional stress) on nicotinamide metabolism were investigated. RESULTS: A diurnal variation in the nicotinamide metabolites N(1)-methylnicotinamide, N(1)-methyl-2-pyridone-5-carboxamide, and N(1)-methyl-4-pyridone-3-carboxamide was observed. Of the stresses studied, cold exposure significantly increased the urinary excretory outputs of the nicotinamide metabolites. CONCLUSIONS: Diurnal variations in nicotinamide metabolism were found in these women. The biosynthesis of nicotinamide from tryptophan seemed to be increased by cold exposure.

Effects of stress on the urinary excretory pattern of niacin catabolites, the most reliable index of niacin status, in humans.

Urinary output of water-soluble vitamins has been used as an indices for vitamin nutrition. It has been pointed out that the coefficient variance of these values is high, especially for niacin catabolites. Thus, we investigated what kinds of stress affect the catabolism using female subjects. The effects of cold exposure (as a typical physical stress), calculation exercise (a typical mental stress) and dark exposure (a typical emotional stress) on the metabolism of niacin were investigated. Of the stresses, cold exposure significantly increased urinary excretory output of the niacin metabolites. The biosynthesis of nicotinamide from Trp seemed to be increased by cold exposure.