Simultaneous quantification of leukotrienes and hydroxyeicosatetraenoic acids in cell culture medium using liquid chromatography/tandem mass spectrometry

Leukotrienes (LTs) and hydroxyeicosatetraenoic acids (HETEs) are important bioactive lipid mediators that participate in various pathophysiological processes. To advance understanding of the mechanisms that regulate these mediators in physiological and pathological processes, an analytical method using liquid chromatography/tandem mass spectrometry for the simultaneous quantification of LTB₄, LTC₄, LTD₄, LTE₄, 5‐HETE_, 8‐HETE, 12‐HETE and 15‐HETE in cell culture media was developed. A Supel?‐Select HLB solid‐phase extraction cartridge was used for sample preparation. The compounds were separated on a C₁₈ column using gradient elution with acetonitrile–water–formic acid (20:80:0.1, v/v/v) and acetonitrile–formic acid (100:0.1, v/v). The calibration curves of LTB₄, LTD₄, LTE₄ and HETEs were linear in the range of 0.025–10 ng/mL, and the calibration curve of LTC₄ was linear in the range of 0.25–10 ng/mL. Validation assessment showed that the method was highly reliable with good accuracy and precision. The stability of LTs and HETEs was also investigated. Using the developed method, we measured LTs and HETEs in the culture supernatant of the human mast cell line HMC‐1. The present method could facilitate investigations of the mechanisms that regulate the production, release and signaling of LTs and HETEs. Copyright © 2014 John Wiley & Sons, Ltd.     

Theory of chemical bonds in metalloenzymes XXII: a concerted bond-switching mechanism for the oxygen–oxygen bond formation coupled with one electron transfer for water oxidation in the oxygen-evolving complex of photosystem II

ABSTRACT

QM(UB3LYP)/MM(AMBER) calculations were performed for the locations of the transition structure (TS) of the oxygen–oxygen (O–O) bond formation in the S₄ state of the oxygen-evolving complex (OEC) of photosystem II (PSII). The natural orbital (NO) analysis of the broken-symmetry (BS) solutions was also performed to elucidate the nature of the chemical bonds at TS on the basis of several chemical indices defined by the occupation numbers of NO. The computational results revealed a concerted bond switching (CBS) mechanism for the oxygen–oxygen bond formation coupled with the one-electron transfer (OET) for water oxidation in OEC of PSII. The orbital interaction between the σ-HOMO of the Mn(IV)₄–O₍₅₎ bond and the π*-LUMO of the Mn(V)₁=O₍₆₎ bond plays an important role for the concerted O–O bond formation for water oxidation in the CaMn₄O₆ cluster of OEC of PSII. One electron transfer (OET) from the π-HOMO of the Mn(V)₁=O₍₆₎ bond to the σ*-LUMO of the Mn(IV)₄–O₍₅₎ bond occurs for the formation of electron transfer diradical, where the generated anion radical [Mn(IV)₄–O₍₅₎]^-? part is relaxed to the ?Mn(III)₄?…?O₍₅₎^- structure and the cation radical [O₍₆₎=Mn(V)₁]⁺ ? part is relaxed to the ⁺O₍₆₎–Mn(IV)₁? structure because of the charge-spin separation for the electron-and hole-doped Mn–oxo bonds. Therefore, the local spins are responsible for the one-electron reductions of Mn(IV)₄->Mn(III)₄ and Mn(V)₁->Mn(IV)₁. On the other hand, the O₍₅₎^- and O₍₆₎⁺ sites generated undergo the O–O bond formation in the CaMn₄O₆ cluster. The Ca(II) ion in the cubane- skeleton of the CaMn₄O₆ cluster assists the above orbital interactions by the lowering of the orbital energy levels of π*-LUMO of Mn(V)₁=O₍₆₎ and σ*-LUMO of Mn(IV)₄–O₍₅₎, indicating an important role of its Lewis acidity. Present CBS mechanism for the O–O bond formation coupled with one electron reductions of the high-valent Mn ions is different from the conventional radical coupling (RC) and acid-base (AB) mechanisms for water oxidation in artificial and native photosynthesis systems. The proton-coupled electron transfer (PC-OET) mechanism for the O–O bond formation is also touched in relation to the CBS-OET mechanism.     

A CMOS multichannel 10-Gb/s transceiver

We describe a CMOS multichannel transceiver that transmits and receives 10 Gb/s per channel over balanced copper media. The transceiver consists of two identical 10-Gb/s modules. Each module operates off a single 1.2-V supply and has a single 5-GHz phase-locked loop to supply a reference clock to two transmitter (Tx) channels and two receiver (Rx) channels. To track the input-signal phase, the Rx channel has a clock recovery unit (CRU), which uses a phase-interpolator-based timing generator and digital loop filter. The CRU can adjust the recovered clock phase with a resolution of 1.56 ps. Two sets of two-channel transceiver units were fabricated in 0.11-/spl mu/m CMOS on a single test chip. The transceiver unit size was 1.6 mm /spl times/ 2.6 mm. The Rx sensitivity was 120-mVp-p differential with a 70-ps phase margin for a common-mode voltage ranging from 0.6 to 1.0 V. The evaluated jitter tolerance curve met the OC-192 specification.     

A new lateral MOS-gated thyristor with controlling base-current

A new lateral MOS-gated thyristor, called the Base-Current-Controlled Thyristor, is described. This device is designed so that most holes at the on-stage reach the P base through the floating P⁺ region adjacent to the P base and the on-state MOSFET. At the turn-off stage, the interruption of the hole current to the P base due to switching off the above MOSFET occurs simultaneously with the conventional turn-off operation. The concept of this device is verified experimentally by using the fabricated lateral device with the external MOSFET. This device exhibits a better trade-off relation between the on-state voltage and the turn-off time compared uith the conventional MOS-gated thyristor     

Highly reliable 200 mW AlGaAs laser diode with fundamental transverse mode

A highly reliable 200 mW AlGaAs laser diode with a fundamental transverse mode has been developed, by optimising its structure with a 0.8 mu m thick p-cladding layer, a 1200 mu m long cavity length, and a front facet coating with a low reflectivity of 2%. The maximum output power was 500 mW, and stable fundamental transverse mode operation was obtained up to 350 mW. Stable operation under 200 mW and 50 degrees C was confirmed for more than 1200 h. Optical feedback noise was below 3*10/sup -14/ Hz/sup -1/.<>     

Estimation of growth rate in unidirectionally solidified multicrystalline silicon by the growth-induced striation method

Multicrystalline silicon was grown by unidirectional solidification method using the accelerated crucible rotation technique. The application of the accelerated crucible rotation technique in unidirectional solidification method induced growth striations across the axial direction of the grown crystal. This striation pattern was observed from carbon concentration distribution, obtained by using Fourier transform infrared spectroscopy. The generated striation pattern was found to be weak and discontinuous. Some striations were absent, probably due to back melting, caused during each crucible rotation. From the growth striations and applied time period in crucible rotation, the growth rate was estimated by using Fourier transformation analysis.     

Simultaneous determination of serotonin and 5-hydroxyindole-3-acetic acid in human urine by automated precolumn derivatization and semi-microbore column liquid chromatography with fluorescence detection.

An improved high-performance liquid-chromatographic (HPLC) assay for the simultaneous determination of serotonin (5-HT) and 5-hydroxyindole-3-acetic acid (5-HIAA) in human urine is reported. Following the automated precolumn derivatization of urinary 5-HT and 5-HIAA with benzylamine, the derivatives are separated by isocratic elution on a reversed-phase C18 semi-microbore column, and are fluorometrically detected at an excitation wavelength of 345 nm and an emission wavelength of 480 nm. The detection limits of 5-HT and 5-HIAA are 7 and 5 nmol/l in urine (0.7 and 0.5 fmol/20-microl injection). The proposed automatic method permits a highly selective and sensitive determination of 5-HT and 5-HIAA in human urine without any sample purification.     

Existence of global strong solution to the micropolar fluid system in a bounded domain

In this paper we are concerned with the initial boundary value problem of the micropolar fluid system in a three dimensional bounded domain. We study the resolvent problem of the linearized equations and prove the generation of analytic semigroup and its time decay estimates. In particular, L^p–L^q type estimates are obtained. By use of the L^p–L^q estimates for the semigroup, we prove the existence theorem of global in time solution to the original nonlinear problem for small initial data. Furthermore, we study the magneto‐micropolar fluid system in the final section. Copyright © 2005 John Wiley & Sons, Ltd.