A sensitive method for quantitative analysis of phospholipid molecular species by high-performance liquid chromatography

A simple and sensitive method was developed for quantitative analysis of phospholipid molecular species. Diacylglycerols were prepared from phospholipids by phospholipase C treatment and converted to the corresponding dinitrobenzoyl derivatives, which could be sensitively detected at 254 nm. The derivatives of 21 molecular species were resolved by high-performance liquid chromatography with an octadecylsilyl reversed-phase column. All the derivatives had the same peak area per mol, and peak areas were proportional to the amounts of the derivatives. Quantification was carried out at the picomole level.     

Potential Distribution and Ionic Concentration at the Bean Root Surface of the Growing Tip and Lateral Root Emerging Points

The electrical potential distribution has been measured preciselyaround the root surface of the bean sprout Vigna mungo (L) Hepper.A large negative potential well was found at the growth portionof the root tip. Also, in the matured region of the root, wefound a negative potential well at an unspecified position inspite of the fact that nothing was detected on the smooth surface.A lateral root emerge was found to have initiated after 15–20hours just at the position corresponding to the potential well.With the expectation that these potentials can be elucidatedbased on the transport of ions which are released or absorbedby the root as a result of cell activity, we precisely measuredthe concentrations of major ion species (K⁺, H⁺, and Cl^–)around the root. The theoretical potential distribution curvesobtained by putting all the concentration data into the Henderson'sEquation for a liquid junction (diffusion) potential coincidedwell with the experimental curves. (Received October 24, 1994; Accepted March 24, 1995)     

Differential pulse polarographic determination of plasma menadione

A differential pulse polarographic assay for plasma vitamin K₃ (menadione) has been developed. Details of the assay are (i) lipid-soluble material is extracted from plasma into ether by the method of Bjornsson et al. [(1978) Thromb. Haemostas.2, 466–473]; (ii) ether is evaporated under nitrogen and the residue is dissolved in the supporting electrolyte, methanol: 0.2 m borate buffer (9:1), pH 6.8; (iii) current height is measured at ?0.32 V vs SCE on the differential pulse polarogram. The lower sensitivity limit of this technique after addition of standard vitamin K₃ to plasma is 0.3 μm; the calibration curve is linear from 0.6 through 10 μm. Two patients treated with a single dose of menadiol sodium diphosphate, 20 mg/M² i.m., achieved measurable plasma vitamin K₃ levels at 0.5 to 1.0 h ranging between 0.5 (0.08 μg/ml) and 2 μm (0.3 μg/ml).     

Quantitative and specific detection of a trichloroethylene-degrading methanotroph, Methylocystis sp. strain M, by a most probable number-polymerase chain reaction method

We developed a rapid and specific enumeration method for a trichloroethylene-degrading methanotroph, Methylocystis sp. strain M, based on a most probable number-polymerase chain reaction method for monitoring the bacterium at bioremediation sites. The primers designed for the mmoC gene of the soluble methane monooxygenase gene cluster were specific to strain M. Recovery of the cells with a membrane filter enabled us to detect strain M in trichloroethylene-contaminated groundwater. We used the enumeration method to monitor the number of strain M cells in effluent from soil columns supplied with trichloroethylene-contaminated groundwater. The number of strain M cells in the effluent depended on the amount of the strain M inoculated and the number of cells measured by the most probable number-polymerase chain reaction method was correlated with that measured by a culture method. The detection limit for strain M in effluent detected by MPN-PCR method was 4 to 8 x 10(2) cells/ml.     

Daily Rhythms of P‐glycoprotein Expression in Mice

Recent studies have shown the gene expression of several transporters to be circadian rhythmic. However, it remains to be elucidated whether the expression of P‐glycoprotein, which is involved in the transport of many medications, undergoes 24 h rhythmicity. To address this issue, we investigated daily profiles of P‐glycoprotein mRNA and protein levels in peripheral mouse tissues. In the liver and intestine, but not in the kidney, Abcb1a mRNA expression showed clear 24 h rhythmicity. On the other hand, Abcb1b and Abcb4, the other P‐glycoprotein genes, did not exhibit significant rhythmic expression in the studied tissues. In the intestine, levels of whole P‐glycoprotein also exhibited a daily rhythm, with a peak occurring in the latter half of the light phase and a trough at the onset of the light phase. Consistent with the day‐night change of P‐glycoprotein level, the ex vivo accumulation of digoxin, an Abcb1a P‐glycoprotein substrate, into the intestinal segments at the onset of dark phase was significantly lower than it was at the onset of the light phase. Thus, Abcb1a P‐glycoprotein expression, and apparently its function, are 24 h rhythmic at least in mouse intestine tissue. This circadian variation might be involved in various chronopharmacological phenomena.     

Polarographic studies of the oxidation of vitamin K5 in aqueous solution.

Electrochemical behavior of 4-amino-2-methyl-1-naphthol hydrochloride (VK₅) has been studied by de and ac polarography in order to determine the applicability of these methods to the determination of VK₅ in aqueous solution. VK₅ exhibits two well-defined anodic waves on dc polarograms, one of which is due to the oxidation of VK₅ to the corresponding quinoimine and the other, to the formation of mercurous chloride on a dropping mercury electrode. Complete preremoval of dissolved oxygen makes the former wave reproducible, and its limiting current is found to be proportional to the VK₅ concentration ranging from 1.0 × 10^?5 to 1.0 × 10^?3m. On ac polarograms, electrooxidation of VK₅ also gives rise to a distinct peak whose height is proportional to the VK₅ concentration in the range of 5.0 × 10^?7 to 1.0 × 10^?5m. These facts can be applied to the microdetermination of VK₅ present in concentrations as low as the 0.1-ppm range.     

Species richness of Protozoa in Japanese lakes

The protozoan fauna and species richness in the pelagic zone of 15 Japanese lakes were investigated in 1996 using polyurethane foam (PF) substrates. The most common species were flagellates, such as Cryptomonas erosa, Oikomonas termo, and Pleuromonas jaculans. Cinetochilum margaritaceum and Actinophrys sol were the most common species of the Ciliata and Sarcodina, respectively. The similarity of species occurrence was calculated from presence/absence data, but this revealed no clear trend with respect to the influence of lake properties such as trophic state, surface area, or mean depth. The occurrence pattern of Protozoa was most similar in L. Chuzenji and L. Biwa (north basin), two oligomesotrophic natural lakes. Log species richness was positively correlated with log total phosphorus (r = 0.54, P < 0.05) and negatively with log mean depth (r = −0.58, P < 0.05). The diversity index (Margalef's formula), highly correlated with the total species number (r = 0.85, P < 0.01), was negatively correlated with log lake area (r = −0.71, P < 0.01). The logarithm of Phytomastigophora number was positively correlated with log total nitrogen (r = 0.53, P < 0.05), and the logarithm of Ciliata number was negatively correlated with log lake area (r = −0.55, P < 0.05). The species richness of Protozoa on PF substrates was determined by both the nutrient status of the lake and the distance from the location of the suspended PF substrate to the lake bottom or shore. Received: September 25, 1999 / Accepted: January 6, 2000     

A 3D-Video-Based Computerized Analysis of Social and Sexual Interactions in Rats

A large number of studies have analyzed social and sexual interactions between rodents in relation to neural activity. Computerized video analysis has been successfully used to detect numerous behaviors quickly and objectively; however, to date only 2D video recording has been used, which cannot determine the 3D locations of animals and encounters difficulties in tracking animals when they are overlapping, e.g., when mounting. To overcome these limitations, we developed a novel 3D video analysis system for examining social and sexual interactions in rats. A 3D image was reconstructed by integrating images captured by multiple depth cameras at different viewpoints. The 3D positions of body parts of the rats were then estimated by fitting skeleton models of the rats to the 3D images using a physics-based fitting algorithm, and various behaviors were recognized based on the spatio-temporal patterns of the 3D movements of the body parts. Comparisons between the data collected by the 3D system and those by visual inspection indicated that this system could precisely estimate the 3D positions of body parts for 2 rats during social and sexual interactions with few manual interventions, and could compute the traces of the 2 animals even during mounting. We then analyzed the effects of AM-251 (a cannabinoid CB1 receptor antagonist) on male rat sexual behavior, and found that AM-251 decreased movements and trunk height before sexual behavior, but increased the duration of head-head contact during sexual behavior. These results demonstrate that the use of this 3D system in behavioral studies could open the door to new approaches for investigating the neuroscience of social and sexual behavior.     

Studies on the Metabolism of Aspergilli

Studies were made on the effect of water level of culture medium on the mycelial compositions and enzyme production in Aspergillus sojae K. S. The mold was grown on the media of various water levels made of powder of defatted soybean and wheat granule. The mycelia grown on the medium of low water level produced more protease and α-amylase, consumed more oxygen, formed less ammonia, and were richer in 2 n H₂SO₄-soluble glycogen, 60% H₂SO₄-soluble carbohydrates, protein and RNA per mg dry weight than the mycelia grown on the medium of high water level. Chromatographic analyses were carried out for nucleotides, sugar phosphates and free carbohydrates in cold TCA-soluble fraction of the mycelia.