Temperature‐programmed capillary high‐performance liquid chromatography with atmospheric pressure chemical ionization mass spectrometry for analysis of fatty acid methyl esters

A new capillary high‐performance liquid chromatography method with atmospheric pressure chemical ionization mass spectrometry was developed for the analysis of fatty acid methyl esters and long‐chain alcohols. The chromatographic separation was achieved using a Zorbax SB‐C18 HPLC column (0.3 × 150 mm, 3.5 μm) with a mobile phase composed of acetonitrile and formic acid and delivered isocratically at a flow rate of 10 μL/min. The column temperature was programmed simply, using a common column oven. Good reproducibility of the temperature profile and retention times were achieved. The temperature programming during the isocratic high‐performance liquid chromatography run had a similar effect as a solvent gradient; it reduced retention times of later eluting analytes and improved their detection limits. Two atmospheric pressure chemical ionization sources of the mass spectrometry detector were compared: an enclosed conventional ion source and an in‐house made ion source with a glass microchip nebulizer. The enclosed source provided better detectability of saturated fatty acid methyl esters and made it possible to determine the double bond positions using acetonitrile‐related adducts, while the open chip‐based source provided better analytical figures of merit for unsaturated fatty acid methyl esters. Temperature‐programmed capillary high‐performance liquid chromatography is a promising method for analyzing neutral lipids in lipidomics and other applications.     

Synthesis of a Monophosphoryl Derivative of Escherichia coli Lipid A and Its Efficient Coupling to a Tumor‐Associated Carbohydrate Antigen

Monophosphoryl lipid A is a safe and potent immunostimulant and vaccine adjuvant, which is potentially useful for the development of effective carbohydrate‐based conjugate vaccines. This paper presents a convergent and efficient synthesis of a monophosphoryl derivative of E. coli lipid A that has an alkyne functionality at the reducing end, which is suitable for coupling with various molecules. The coupling of this derivative to an N‐modified analogue of tumor‐associated antigen GM3 through click chemistry is also presented.     

Site‐Specific Investigation of the Steady‐State Kinetics and Dynamics of the Multistep Binding of Bile Acid Molecules to a Lipid Carrier Protein

The investigation of multi‐site ligand–protein binding and multi‐step mechanisms is highly demanding. In this work, advanced NMR methodologies such as 2D ¹H–¹⁵N line‐shape analysis, which allows a reliable investigation of ligand binding occurring on micro‐ to millisecond timescales, have been extended to model a two‐step binding mechanism. The molecular recognition and complex uptake mechanism of two bile salt molecules by lipid carriers is an interesting example that shows that protein dynamics has the potential to modulate the macromolecule–ligand encounter. Kinetic analysis supports a conformational selection model as the initial recognition process in which the dynamics observed in the apo form is essential for ligand uptake, leading to conformations with improved access to the binding cavity. Subsequent multi‐step events could be modelled, for several residues, with a two‐step binding mechanism. The protein in the ligand‐bound state still exhibits a conformational rearrangement that occurs on a very slow timescale, as observed for other proteins of the family. A global mechanism suggesting how bile acids access the macromolecular cavity is thus proposed.     

General Synthesis and Physicochemical Characterisation of a Series of Peptide‐Mimic Lysine‐Based Amino‐Functionalised Lipids

A series of novel malonic acid diamides (second generation) with two long hydrophobic alkyl chains and an alkaline polar head group was synthesised and characterised as a new class of amino‐functionalised lipids. These peptide‐mimic lipids are suitable for polynucleotide transfer. The lipids bear a novel backbone consisting of a lysine unit and a malonic acid unit. Six different head‐group structures, which vary in size and number of amino groups that can be protonated, were attached to the backbone structure. Furthermore, different alkyl chains were used to build the lipophilic part (namely tetradecyl, hexadecyl, and oleyl). Phase transitions of the new compounds in aqueous dispersions at pH 10 were analysed and discussed in terms of head group and alkyl chain variations. The shape and size of the formed aggregates of selected lipid dispersions were investigated by dynamic light scattering and transmission electron microscopy.