Determination of unbound cefamandole in rat blood by microdialysis and microbore liquid chromatography

To analyze unbound cefamandole in rat blood, a method combing microdialysis with microbore liquid chromatography has been developed. A microdialysis probe was inserted into the jugular vein/right atrium of male Sprague-Dawley rats to examine the unbound cefamandole level in the rat blood following cefamandole administration (50 mg/kg, i.v.). The dialysates were directly submitted to a liquid chromatographic system. Samples were eluted with a mobile phase containing acetonitrile-methanol-100 mM monosodium phosphate (pH 5.0; 15:20:65, v/v). The UV wavelength was set at 270 nm for monitoring the analyte. Using the retrograde method, at infusion concentrations of 1 microg/mL of cefamandole, the in vivo microdialysis recoveries were 55.44% for the rat blood (n = 6). Intra- and inter-assay accuracy and precision of the analyses were < or = 10% in the range of 0.1-10 microg/mL. Pharmacokinetic parameters were calculated from the recovery-corrected dialysate concentrations of cefamandole vs time data. The elimination half-life (t1/2,beta) was 21.6 +/- 1.6 min. The results suggest that the pharmacokinetics of unbound cefamandole in blood following cefamandole administration (50 mg/kg, i.v., n = 5) fit best to the two-compartmental model.     

A new fabrication process for a microchip electrophoresis device integrated with a three-electrode electrochemical detector

We report here a novel and simple process for the fabrication of a poly(methyl methacrylate) (PMMA)-based microchip electrophoresis device, integrated with a screen-printed three-electrode electrochemical detector that does not require a replicate mold. In this approach, a photoresist layer constitutes both an adhesion layer and side walls of 50 mum wide and 50 mum tall microfluidic channels on a screen-printed three-electrode PMMA substrate. Openings were drilled for buffer reservoirs on an additional piece of PMMA, then the final device was bonded in a PMMA/photoresist/PMMA sandwich configuration. This process is inexpensive, less time-consuming, and simpler compared with traditional fabrication methods. The combination of this PMMA-based microchip fabrication together with screen-printed electrode technology holds great promise for the mass production of a single-use micrototal analytical system. Successful determination of uric acid and L-ascorbic acid with the presented system validates its utility. In combination with a suitable electrochemical detector, this device holds much promise for the determination of other analytes in various biological samples for medical and clinical diagnosis.     

Dinitrosyl iron complexes (DNICs) [L(2)Fe(NO)2]- (L = thiolate): interconversion among {Fe(NO)2}9 DNICs, {Fe(NO)2}10 DNICs, and [2Fe-2S] clusters, and the critical role of the thiolate ligands in regulating NO release of DNICs

Dinitrosyl iron complex [(-SC(7)H(4)SN)(2)Fe(NO)(2)](-) (1) was prepared by reaction of [S(5)Fe(NO)(2)](-) and bis(2-benzothiozolyl) disulfide. In synthesis of the analogous dinitrosyl iron compounds (DNICs), the stronger electron-donating thiolates [RS](-) (R = C(6)H(4)-o-NHCOCH(3), C(4)H(3)S, C(6)H(4)NH(2), Ph), compared to [-SC(7)H(4)SN](-) of complex 1, trigger thiolate-ligand substitution to yield [(-SC(6)H(4)-o-NHCOCH(3))(2)Fe(NO)(2)](-) (2), [(-SC(4)H(3)S)(2)Fe(NO)(2)](-) (3), and [(SPh)(2)Fe(NO)(2)](-) (4), respectively. At 298 K, complexes 2 and 3 exhibit a well-resolved five-line EPR signal at g = 2.038 and 2.027, respectively, the characteristic g value of DNICs. The magnetic susceptibility fit indicates that the resonance hybrid of {Fe(+)((*)NO)(2)}(9) and {Fe(-)((+)NO)(2)}(9) in 2 is dynamic by temperature. The IR nu(NO) stretching frequencies (ranging from (1766, 1716) to (1737, 1693) cm(-)(1) (THF)) of complexes 1-4 signal the entire window of possible electronic configurations for such stable and isolable {Fe(NO)(2)}(9) [(RS)(2)Fe(NO)(2)](-). The NO-releasing ability of {Fe(NO)(2)}(9) [(RS)(2)Fe(NO)(2)](-) is finely tuned by the coordinated thiolate ligands. The less electron-donating thiolate ligands coordinated to {Fe(NO)(2)}(9) motif act as better NO-donor DNICs in the presence of NO-trapping agent [Fe(S,S-C(6)H(4))(2)](2)(2-). Interconversion between {Fe(NO)(2)}(9) [(RS)(2)Fe(NO)(2)](-) and {Fe(NO)(2)}(10) [(Ph(3)P)(2)Fe(NO)(2)] was verified in the reaction of (a) [(RS)(2)Fe(NO)(2)](-), 10 equiv of PPh(3) and sodium-biphenyl, and (b) 2 equiv of thiol, [RS](-), and [(Ph(3)P)(2)Fe(NO)(2)], respectively. The biomimetic reaction cycle, transformation between {Fe(NO)(2)}(9) [(RS)(2)Fe(NO)(2)](-) and {Fe(NO)(2)}(9) [(R'S)(2)Fe(NO)(2)](-), reversible interconversion of {Fe(NO)(2)}(9) and {Fe(NO)(2)}(10) DNICs, and degradation/reassembly of [2Fe-2S] clusters may decipher and predict the biological cycle of interconversion of {Fe(NO)(2)}(9) DNICs, {Fe(NO)(2)}(10) DNICs, and the [Fe-S] clusters in proteins.     

Self‐Assembly,Spectroscopic and Electrochemical Studies of Nickel(II)‐4,8‐Diazaundecanediamide Complex

The self‐assembly of NiCl₂·6H₂O with a diaminodiamide ligand 4,8‐diazaundecanediamide (L‐2,3,2) gave a [Ni(C₉H₂₀N₄O₂)(Cl)(H₂O)] Cl·2H₂O ( 1 ). The structure of 1 was characterized by single‐crystal X‐ray diffraction analysis. Structural data for 1 indicate that the Ni(II) is coordinated to two tertiary N atoms, two O atoms, one water and one chloride in a distorted octahedral geometry. Crystal data for 1: orthorhombic, space group P 21nb, a = 9.5796(3) Å, b = 12.3463(4) Å, c = 14.6305(5) Å, Z = 4. Through NH···Cl–Ni (H···Cl 2.42 Å, N···Cl 3.24 Å, NH···Cl 158°) and OH···Cl–Ni contacts (H···Cl 2.36 Å, O···Cl 3.08 Å, OH···Cl 143°), each cationic moiety [Ni(C₉H₂₀N₄O₂) (Cl)(H₂O)]⁺ in 1 is linked to neighboring ones, producing a charged hydrogen‐bonded 1D chainlike structure. Thermogrametric analysis of compound 1 is consistent with the crystallographic observations. The electronic absorption spectrum of Ni(L‐2,3,2)²⁺ in aqueous solution shows four absorption bands, which are assigned to the ³A_2g → ³T_2g, ³T_2g → ¹Eg, ³T_2g → ³T_1g, and ³A_2g → ³T_1g transitions of triplet‐ground state, distorted octahedral nickel(II) complex. The cyclic volammetric measurement shows that Ni²⁺ is more easily reduced than Ni(L‐2,3,2)²⁺ in aqueous solution.     

Optimization of the separation of malachite green in water by capillary electrophoresis Raman spectroscopy (CE-RS) based on the stacking and sweeping modes

A capillary electrophoresis Raman spectroscopy (CE-RS) method based on the stacking and sweeping modes are described. A non-fluorescent compound (malachite green, MG; crystal violet, CV) and a doubled Nd:YAG laser (532 nm, 300 mW) were selected as the model compound and light source, respectively. In order to carry out a quantitative and analysis of MG, a monochromator was used to collect the specific Raman line at 1616 cm⁻¹ (the N-φ and C-C stretching, corresponding to 582 nm when the wavelength of the exciting source is 532 nm). The limit of detection (LOD) for MG was 1.6 × 10⁻⁵ and 1.1 × 10⁻⁵ M, respectively, based on the CZE and MEKC modes. This could be improved to 3.4 × 10⁻⁷ and 5.3 × 10⁻⁹ M, respectively, when the stacking and sweeping modes were applied. The method was also extended to the determination of MG in an actual sample.     

Synthesis of isotopically labeled arachidonic acids to probe the reaction mechanism of prostaglandin H synthase

Prostaglandin H synthase (PGHS) catalyzes the conversion of arachidonic acid to prostaglandin G(2) in the cyclooxygenase reaction. The first step of the mechanism has been proposed to involve abstraction of the pro-S hydrogen atom from C13 to generate a pentadienyl radical spanning C11-C15. We report here the synthesis of six site-specifically deuterated arachidonic acids to investigate the structure of the radical intermediate. The preparation of these compounds was achieved using a divergent scheme that involved one advanced intermediate for all targets. The synthetic design introduced the label late in the routes and allowed the utilization of common synthetic intermediates in the preparation of various targets. Both 13(R)- and 13(S)-deuterium-labeled arachidonic acids were synthesized in high enantiomeric purity as deduced from soybean lipoxygenase assays and mass spectrometric analysis of the resulting enzymatic products. Each synthetic compound was reacted under anaerobic conditions with the wide singlet tyrosyl radical of PGHS-2 to generate a radical intermediate that was analyzed by EPR. Deuterium substitution at positions 11, 13(S), and 15 resulted in the loss of one hyperfine interaction, indicating that the protons at these positions interact with the unpaired electron. Simulation of the spectra was achieved with one set of parameters that are consistent with the assignment of a pentadienyl radical. Use of 16-[(2)H(2)]-arachidonic acid indicated that only one of the protons at C16 gives rise to a strong hyperfine interaction. The findings are discussed in the context of two proposed mechanisms for the cyclooxygenase reaction.     

Semiempirical Molecular Orbital Studies of the Acylation Step in the Lipase‐Catalyzed Ester Hydrolysis

In this study, we present the results from the semiempirical molecular orbital calculations for the acylation step in the lipase‐catalyzed ester hydrolysis. The results reveal that the lowest energy path for the formation of the tetrahedral intermediate is for the serine residue of the catalytic triad to attack the substrate, followed by coupling heavy atom movement and proton transfer. The calculations of four active site models show that the cooperation of the aspartate group and the oxyanion hole is capable of lowering the activation energy by about 16 kcalmol^?1. Our results further suggest that the lipase‐catalyzed ester hydrolysis adopts the single proton transfer mechanism.     

Application of plasma-polymerized films for isoelectric focusing of proteins in a capillary electrophoresis chip

The first use of plasma polymerization technique to modify the surface of a glass chip for capillary isoelectric focusing (cIEF) of different proteins is reported. The electrophoresis separation channel was machined in Tempax glass chips with length 70 mm, 300 microm width and 100 microm depth. Acetonitrile and hexamethyldisiloxane monomers were used for plasma polymerization. In each case 100 nm plasma polymer films were coated onto the chip surface to reduce protein wall adsorption and minimize the electroosmotic flow. Applied voltages of 1000 V, 2000 V and 3000 V were used to separate mixtures of cytochrome c (pI 9.6), hemoglobin (pI 7.0) and phycocyanin (pI 4.65). Reproducible isoelectric focusing of each pI marker protein was observed in different coated capillaries at increasing concentration 2.22-5 microg microL(-1). Modification of the glass capillary with hydrophobic HMDS plasma polymerized films enabled rapid cIEF within 3 min. The separation efficiency of cytochrome c and phycocyanin in both acrylamide and HMDS coated capillaries corresponded to a plate number of 19600 which compares favourably with capillary electrophoresis of neurotransmitters with amperometric detection.     

Kinetics of Heterogeneous Isotopic Exchange Reaction in Finite Bath

A general method to find the rate constant and particle self-diffusion coefficient is suggested for a heterogeneous isotopic exchange reaction which is controlled by surface mass reaction or controlled by a combination of surface mass reaction and intraparticle diffusion. The values of the kinetic parameter, ξ₁ (ratio of the forward surface mass reaction rate to the intraparticle diffusion rate), particle self-diffusion coefficient D and rate constant k are obtained by the proposed method for the isotopic exchange reaction systems CaCO₃(s)/Ca²⁺(aq) and CaC₂O₄(s)/Ca²⁺(aq).     

Determination of alkyltrimethylammonium surfactants in hair conditioners and fabric softeners by gas chromatography-mass spectrometry with electron-impact and chemical ionization

The commercial hair conditioners and fabric softeners were analyzed for the content of alkyltrimethylammonium compounds (ATMACs) by gas chromatography-mass spectrometry (GC-MS) with electron impact (EI) and low-pressure positive-ion chemical ionization (PICI) modes. The method involves mixed diluted samples (adjust pH to 10.0) with potassium iodide to enhance the extraction of iodide-ATMA+ ion pairs by direct liquid-liquid extraction. The iodide-ATMA+ pairs were then demethylated to their corresponding nonionic alkyldimethylamines (ADMAs) by thermal decomposition in a GC injection-port. A high abundance of ADMAs was detected at the temperature above 300 degrees C in the GC injection-port. The enhanced selectivity of quasi-molecular ion chromatograms of C12-C18-ADMA, obtained using methanol PICI-MS enables ADMAs to be identified. The accuracy and precision of the method was validated and was successfully applied to determine contents of ATMAC in commercial hair conditioners and fabric softeners. The contents of total measured ATMAC ranged from 0.4 to 6.9% for hair conditioners, and from 3.3 to 4.6% for fabric softeners.