Convergence of Free Energy Profile of Coumarin in Lipid Bilayer

Atomistic molecular dynamics (MD) simulations of druglike molecules embedded in lipid bilayers are of considerable interest as models for drug penetration and positioning in biological membranes. Here we analyze partitioning of coumarin in dioleoylphosphatidylcholine (DOPC) bilayer, based on both multiple, unbiased 3 μs MD simulations (total length) and free energy profiles along the bilayer normal calculated by biased MD simulations (∼7 μs in total). The convergences in time of free energy profiles calculated by both umbrella sampling and z-constraint techniques are thoroughly analyzed. Two sets of starting structures are also considered, one from unbiased MD simulation and the other from “pulling” coumarin along the bilayer normal. The structures obtained by pulling simulation contain water defects on the lipid bilayer surface, while those acquired from unbiased simulation have no membrane defects. The free energy profiles converge more rapidly when starting frames from unbiased simulations are used. In addition, z-constraint simulation leads to more rapid convergence than umbrella sampling, due to quicker relaxation of membrane defects. Furthermore, we show that the choice of RESP, PRODRG, or Mulliken charges considerably affects the resulting free energy profile of our model drug along the bilayer normal. We recommend using z-constraint biased MD simulations based on starting geometries acquired from unbiased MD simulations for efficient calculation of convergent free energy profiles of druglike molecules along bilayer normals. The calculation of free energy profile should start with an unbiased simulation, though the polar molecules might need a slow pulling afterward. Results obtained with the recommended simulation protocol agree well with available experimental data for two coumarin derivatives

An Expedient Synthesis of CMF-019: (S)-5-Methyl-3-{1-(pentan-3-yl)-2- (thiophen-2-ylmethyl)-1H-benzo[d]imidazole-5-carboxamido}hexanoic Acid, a Potent Apelin Receptor (APJ) Agonist.

BACKGROUND: Apelin receptor (APJ) is a G protein-coupled receptor (GPCR) activated by the endogenous peptide apelin. The apelin-APJ system has emerged as an important regulator of cardiovascular homeostasis. Recently, a potent benzimidazole-derived apelin peptidomimetic, CMF-019, was patented but without a comprehensive description of its synthesis and a complete spectroscopic characterization of the intermediates. OBJECTIVE: Here, a detailed preparation of CMF-019 through a modified and improved synthetic pathway is described. METHOD: In particular, the benzimidazole ring in 7 was tailored by the condensation of methyl 3- amino-4-(pentan-3-ylamino)benzoate (4) with (thiophene-2-yl)acetimidate salt 6. Saponification of 7 and the subsequent condensation of the free acid 8 with the corresponding enantiopure β-amino acid methyl ester generated methyl (S)-5-methyl-3-{1-(pentan-3-yl)-2-(thiophen-2-ylmethyl)-1Hbenzo[ d]imidazole-5-carboxamido}hexanoate (9). Hydrolysis of the latter with KOH in THF/water, followed by HPLC-purification, afforded the desired product, CMF-019 (potassium salt) 10. RESULTS & CONCLUSION: The approach reported herein enables preparation of 10 at a total yield of 12% over seven linear steps. Additionally, it does not require applying expensive designated microwave reactors and high-pressure hydrogenators. Thus, the elaborate synthesis provides a latent availability of potent agonist 10 for further exploring the physiologically essential apelin-APJ system

Locating Intercalants Within Lipid Bilayers Using Fluorescence Quenching by Bromophospholipids and Iodophospholipids

In previous work, we have been able to determine the depth of intercalated molecules within the lipid bilayer using the solvent polarity sensitivity of three spectroscopic techniques: the 13C NMR chemical shift (δ); the fluorescence emission wavelength (λem), and the ESR β-H splitting constants (aβ-H). In the present paper, we use the quenching by a heavy atom (Br or I), situated at a known location along a phospholipid chain, as a probe of the location of a fluorescent moiety. We have synthesized various phospholipids with bromine (or iodine) atoms substituted at various locations along the lipid chain. The latter halolipids were intercalated in turn with various fluorophores into DMPC liposomes, biomembranes and erythrocyte ghosts. The most effective fluorescence quenching occurs when the heavy atom location corresponds to that of the fluorophore. The results show that generally speaking the fluorophore intercalates the same depth independent of which lipid bilayer is used. KBr (or KI) is the most effective quencher when the fluorophore resides in or at the aqueous phase. Presumably because of iodine\u27s larger radius and spin coupling constant, the iodine analogs are far less discriminating in the depth range it quenches

NMR-Based Molecular Ruler for Determining the Depth of Intercalants Within the Lipid Bilayer. Part V: A Comparison of Liposomes, Bioliposomes and Erythrocyte Ghosts

Afri et al., 2014a, Afri et al., 2014b have recently reported their mapping of DMPC liposomes using 13C NMR in conjunction with a wide range of difunctional intercalants: n-ketoesters, n-ketoacids and n-ketophosphatidylcholines. The present study initiates a comparable study of bioliposomes and erythrocyte ghosts. This required the 13C NMR characterization of these systems for the first time, and further involved a determination of the signals of three doubly 13C-labeled intercalants, in particular, n-ketophosphatidylcholines where n = 4, 8 and 12. This study reveals that DMPC liposomes, bioliposomes and erythrocyte ghosts, with all their structural differences, are not radically different from the perspective of polarity gradient. Any differences observed reflect the additives often naturally present in these lipid systems

Li-O-2 cells with LiBr as an electrolyte and a redox mediator

After many years of successful and disappointing results, the field of Li-O-2 research seems to have reached an equilibrium state. The extensive knowledge that has accrued through advanced analytical studies enables us to delineate the weaknesses of the Li-O-2 battery. It is now clear that the instability of the cell components toward extreme conditions existing during cell operation leads to early cell failure as well. One serious challenge is the high oxidation potential applied during the charge process. Redox-mediators may reduce the over-potential and, therefore, improve the efficiency and cyclability of Li-O-2 cells. Their use in Li-O-2 cells is mandatory. We have previously shown that LiI can indeed behave in such a manner; however, it also promotes the formation of side products during cell operation. We have, therefore, embarked on a comprehensive study of lithium halide salts as electrolytes for use in Li-O-2 cells. We examine herein the effect of other components in the cell, such as solvents and contaminants, on the lithium halide salt activity. Based on the electrochemical behavior and the identity of the final cell products under various conditions, we can glean substantial information regarding the detailed operation mechanisms for each specific case. We have concluded that low concentration of LiBr in diglyme solution can improve the cell performance with fewer side effects than LiI. With LiBr, only the desired Li2O2 is formed during discharge. During charge, the bromine redox couple (Br-/Br-3(-)) can reduce the oxidation potential to only 3.5 V. Higher efficiency and better cyclability of cells containing LiBr demonstrate that the electrolyte solution is the key to a successful Li-O-2 battery

NMR-Based Molecular Ruler for Determining the Depth of Intercalants Within the Lipid Bilayer. Part IV: Studies on Ketophospholipids

In our companion paper, we described the preparation and intercalation of two homologous series of dicarbonyl compounds, methyl n-oxooctadecanoates and the corresponding n-oxooctadecanoic acids (n = 4–16), into DMPC liposomes. 13C NMR chemical shift of the various carbonyls was analyzed using an ET(30) solvent polarity–chemical shift correlation table and the corresponding calculated penetration depth (in Å). An iterative best fit analysis of the data points revealed an exponential correlation between ET(30) micropolarity and the penetration depth (in Å) into the liposomal bilayer. However, this study is still incomplete, since the plot lacks data points in the important area of moderately polarity, i.e., in the ET(30) range of 51–45.5 kcal/mol. To correct this lacuna, a family of ketophospholipids was prepared in which the above n-oxooctadecanoic acids were attached to the sn-2 position of a phosphatidylcholine with a palmitic acid chain at sn-1. To assist in assignment and detection several derivatives were prepared 13C-enriched in both carbonyls. The various homologs were intercalated into DMPC liposomes and give points specifically in the missing area of the previous polarity–penetration correlation graph. Interestingly, the calculated exponential relationship of the complete graph was essentially the same as that calculated in the companion paper based on the methyl n-oxooctadecanoates and the corresponding n-oxooctadecanoic acids alone. The polarity at the midplane of such DMPC systems is ca. 33 kcal/mol and is not expected to change very much if we extend the lipid chains. This paper concludes with a chemical ruler that maps the changing polarity experienced by an intercalant as it penetrates the liposomal bilayer

On the Challenge of Electrolyte Solutions for Li–Air Batteries: Monitoring Oxygen Reduction and Related Reactions in Polyether Solutions by Spectroscopy and EQCM

Polyether solvents are considered interesting and important candidates for Li–O₂ battery systems. Discharge of Li–O₂ battery systems forms Li oxides. Their mechanism of formation is complex. The stability of most relevant polar aprotic solvents toward these Li oxides is questionable. Specially high surface area carbon electrodes were developed for the present work. In this study, several spectroscopic tools and in situ measurements using electrochemical quartz crystal microbalance (EQCM) were employed to explore the discharge–charge processes and related side reactions in Li–O₂ battery systems containing electrolyte solutions based on triglyme/lithium bis­(trifluoromethanesulfonyl)­imide (LiTFSI) electrolyte solutions. The systematic mechanism of lithium oxides formation was monitored. A combination of Fourier transform infrared (FTIR), NMR, and matrix-assisted laser desorption/ionization (MALDI) measurements in conjunction with electrochemical studies demonstrated the intrinsic instability and incompatibility of polyether solvents for Li–air batteries

NMR-Based Molecular Ruler for Determining the Depth of Intercalants Within the Lipid Bilayer: Part III: Studies on Keto Esters and Acids

The development of “molecular rulers” would allow one to quantitatively locate the penetration depth of intercalants within lipid bilayers. To this end, an attempt was made to correlate the 13C NMR chemical shift of polarizable “reporter” carbons (e.g., carbonyls) of intercalants within DMPC liposomal bilayers – with the polarity it experiences, and with its Angstrom distance from the interface. This requires families of molecules with two “reporter carbons” separated by a known distance, residing at various depths/polarities within the bilayer. For this purpose, two homologous series of dicarbonyl compounds, methyl n-oxooctadecanoates and the corresponding n-oxooctadecanoic acids (n = 4–16), were synthesized. To assist in assignment and detection several homologs in each system were prepared 13C-enriched in both carbonyls. Within each family, the number of carbons and functional groups remains the same, with the only difference being the location of the second ketone carbonyl along the fatty acid chain. Surprisingly, the head groups within each family are not anchored near the lipid–water interface, nor are they even all located at the same depth. Nevertheless, using an iterative best fit analysis of the data points enables one to obtain an exponential curve. The latter gives substantial insight into the correlation between polarity (measured in terms of the Reichardt polarity parameter, ET(30)) and penetration depth into the liposomal bilayer. Still missing from this curve are data points in the moderate polarity range