Optimized extraction of phospholipids and lysophospholipids for nanoflow liquid chromatography-electrospray ionization-tandem mass spectrometry

The efficiencies of four different methods for the extraction of phospholipids (PLs) and lysophospholipids (LPLs) from human plasma samples were examined by comparing extraction recovery values using nanoflow liquid chromatography-electrospray ionization-mass spectrometry (nLC-ESI-MS). For recovery measurements, six PL and six LPL standards of different head groups were spiked into a human plasma sample, and the peak areas of each individual species after extraction were measured from the chromatograms of the nLC-ESI-MS runs. Recovery was calculated by comparing the peak area of an extracted standard species with that of the same species' spike after extraction of the same plasma sample. For lipid extraction, four different extraction methods were examined: three based on the Folch method with different organic solvents such as CHCl(3), methyl-tert-butyl ether (MTBE), and MTBE/CH(3)OH, and one relatively fast method involving CH(3)OH only. Evaluations of recovery showed that the modified Folch method with MTBE/CH(3)OH proposed in this study was effective for extracting most PL and LPL standards. Then, the four extraction methods were compared with the identified numbers of plasma PLs and LPLs, of which molecular structures can be confirmed by data-dependent, collision-induced dissociation experiments during nLC-ESI-MS-MS. These results demonstrated that the proposed method yielded the identification of 54 LPLs and 66 PLs from a plasma sample, which was the highest identification rate among the four methods.     

Structural and thermodynamic investigations on the aggregation and folding of acylphosphatase by molecular dynamics simulations and solvation free energy analysis

Protein engineering method to study the mutation effects on muscle acylphosphatase (AcP) has been actively applied to describe kinetics and thermodynamics associated with AcP aggregation as well as folding processes. Despite the extensive mutation experiments, the molecular origin and the structural motifs for aggregation and folding kinetics as well as thermodynamics of AcP have not been rationalized at the atomic resolution. To this end, we have investigated the mutation effects on the structures and thermodynamics for the aggregation and folding of AcP by using the combination of fully atomistic, explicit-water molecular dynamics simulations, and three-dimensional reference interaction site model theory. The results indicate that the A30G mutant with the fastest experimental aggregation rate displays considerably decreased α1-helical contents as well as disrupted hydrophobic core compared to the wild-type AcP. Increased solvation free energy as well as hydrophobicity upon A30G mutation is achieved due to the dehydration of hydrophilic side chains in the disrupted α1-helix region of A30G. In contrast, the Y91Q mutant with the slowest aggregation rate shows a non-native H-bonding network spanning the mutation site to hydrophobic core and α1-helix region, which rigidifies the native state protein conformation with the enhanced α1-helicity. Furthermore, Y91Q exhibits decreased solvation free energy and hydrophobicity compared to wild type due to more exposed and solvated hydrophilic side chains in the α1-region. On the other hand, the experimentally observed slower folding rates in both mutants are accompanied by decreased helicity in α2-helix upon mutation. We here provide the atomic-level structures and thermodynamic quantities of AcP mutants and rationalize the structural origin for the changes that occur upon introduction of those mutations along the AcP aggregation and folding processes.     

A complete determination of Rabinowitsch polynomials

Let m be a positive integer and fm(x) be a polynomial of the form fm(x)=x²+x−m. We call a polynomial fm(x) a Rabinowitsch polynomial if for and consecutive integers x=x₀,x₀+1,…,x₀+s−1, |fm(x)| is either 1 or prime. In this paper, we show that there are exactly 14 Rabinowitsch polynomials fm(x).     

Ultrasound-assisted copper deposition on a polymer membrane and application for methanol steam reforming

Copper particles were electrolessly deposited on a palladium aerosol activated polymer membrane in the presence of ultrasound. An application of ultrasound introduced a faster deposition (220 μg min^?1 in deposition rate) and finer copper particles (9 nm in crystallite size) than those (11 and 41 μg min^?1; 27 and 32 nm) in the absence of ultrasound (i.e. respectively 20 and 45 °C in bath temperature with mechanical agitation). A better performance of methanol steam reforming (0.59 in mean conversion during 5 h operation; 1.3 and 1.6 times respectively higher than those from 20 to 45 °C cases) at a 300 °C reaction temperature was materialized for the ultrasound application, probably due to a finer (i.e. a more textured) copper particle deposition on a polymer membrane.     

Standing Waves for Nonlinear Schrödinger Equations with a General Nonlinearity: One and Two Dimensional Cases

For N = 1,2, we consider singularly perturbed elliptic equations ?²Δ u ? V(x) u + f(u)= 0, u(x)> 0 on R ^N , lim_|x|→∞ u(x)= 0. For small ? > 0, we show the existence of a localized bound state solution concentrating at an isolated component of positive local minimum of V under conditions on f we believe to be almost optimal; when N ≥ 3, it was shown in Byeon and Jeanjean (2007 Byeon , J. , Oshita , Y. ( 2004 ). Existence of multi-bump standing waves with a critical frequency for nonlinear Schrödinger equations . Comm. PDE 29 : 1877 – 1904 . [Google Scholar]).     

Quantification and Metabolite Identification of Sulfasalazine in Mouse Brain and Plasma Using Quadrupole-Time-of-Flight Mass Spectrometry

Sulfasalazine (SAS), an anti-inflammatory drug with potent cysteine/glutamate antiporter system xc-(SXC) inhibition has recently shown beneficial effects in brain-related diseases. Despite many reports related to central nervous system (CNS) effect of SAS, pharmacokinetics (PK) and metabolite identification studies in the brain for SAS were quite limited. The aim of this study was to investigate the pharmacokinetics and metabolite identification of SAS and their distributions in mouse brain. Using in vivo brain exposure studies (neuro PK), the PK parameters of SAS was calculated for plasma as well as brain following intravenous and oral administration at 10 mg/kg and 50 mg/kg in mouse, respectively. In addition, in vivo metabolite identification (MetID) studies of SAS in plasma and brain were also conducted. The concentration of SAS in brain was much lower than that in plasma and only 1.26% of SAS was detected in mouse brain when compared to the SAS concentration in plasma (brain to plasma ratio (%): 1.26). In the MetID study, sulfapyridine (SP), hydroxy-sulfapyridine (SP-OH), and N-acetyl sulfapyridine (Ac-SP) were identified in plasma, whereas only SP and Ac-SP were identified as significant metabolites in brain. As a conclusion, our results suggest that the metabolites of SAS such as SP and Ac-SP might be responsible for the pharmacological effect in brain, not the SAS itself.     

Liquid chromatography-high resolution mass spectrometric method for the quantification of monomethyl auristatin E (MMAE) and its preclinical pharmacokinetics

MMAE is a potent antimitotic drug used as payload of an antibody-drug conjugate which shows potent activity in preclinical and clinical studies against a range of lymphomas, leukemia and solid tumors. Liquid chromatography-high resolution mass spectrometric method was developed for the quantification of MMAE and its preclinical pharmacokinetics. The method consisted of protein precipitation using acetonitrile (ACN) for sample preparation and liquid chromatography – quadrupole – time-of-flight – tandem mass spectrometry (LC-qTOF-MS/MS) analysis in the positive ion mode. A quadratic regression (weighted 1/concentration²), with an equation y = ax² + bx + c, was used to fit calibration curves over the concentration range of 1.01–2,200 ng/mL for MMAE. The qualification run met the acceptance criteria of ±25% accuracy and precision values for QC samples. Recovery was 42.84%. The dilution integrity was determined for 5-fold dilution and the accuracy and precision ranged within ±25%. The stability results indicated that MMAE was stable for the following conditions: short-term (4 h), long-term (4 weeks), freeze/thaw (3 cycles) and post-preparative stability (12 h). This qualified method was successfully applied to a pharmacokinetic study of MMAE in rat as a preclinical animal model. The PK results suggest that MMAE has moderate CL and low BA.Also, these results would be helpful in having a comprehensive understanding of the PK characteristics of MMAE and developing ADC in future.     

Symmetry and monotonicity of least energy solutions

We give a simple proof of the fact that for a large class of quasilinear elliptic equations and systems the solutions that minimize the corresponding energy in the set of all solutions are radially symmetric. We require just continuous nonlinearities and no cooperative conditions for systems. Thus, in particular, our results cannot be obtained by using the moving planes method. In the case of scalar equations, we also prove that any least energy solution has a constant sign and is monotone with respect to the radial variable. Our proofs rely on results in Brothers and Ziemer (J Reine Angew Math 384:153–179, 1988) and Mariş (Arch Ration Mech Anal, 192:311–330, 2009) and answer questions from Brézis and Lieb (Comm Math Phys 96:97–113, 1984) and Lions (Ann Inst H Poincaré Anal Non Linéaire 1:223–283, 1984).