Hydrophilic High-performance Liquid Chromatography Research Articles

Simultaneous determination of cationic surfactants in disinfectants

Objectives. Cationic surfactants are one of the classes of substances most commonly used in disinfectants. The trend in recent years has been the use of mixtures of several biocides, which poses new challenges for analytical chemistry. In this study, we describe a method for simultaneous determination in the disinfectants alkyldimethylbenzylammonium chloride (ADBAC), alkyldimethyl(ethylbenzyl)ammonium chloride (ADEBAC), chlorhexidine bigluconate (CHG), and polyhexamethylene biguanide hydrochloride (PHMB). Methods. The proposed method is based on the use of reverse-phase and hydrophilic high-performance liquid chromatography with diode-array detection Results. The best separation of ADBAC, ADEBAC, and CHG was achieved using a column filled with modified spherical silica gel (5 цт , 4.6 × 250 mm) in gradient elution mode. Acetonitrile and acetate buffer with a pH of 5.4 were used as eluents at a flow rate of 1 ml/min. For the determination of PHMB in the presence of the substances under consideration, hydrophilic high performance liquid chromatography was used. The best separation was achieved on an amine phase column (5 цт , 4.6 × 250 mm) using the same eluents. To determine all the substances under consideration, a diode array detector was used. 3D chromatograms were recorded in the wavelength range from 190 to 400 nm. Conclusions. We have shown that the result of the analysis does not depend on the ratio of cationic surfactants in disinfectants. There is also no influence of N,N-bis-(3-aminopropyl)-dodecylamine (Triamine, TA) and the components most commonly used for the manufacture of disinfectants, which was confirmed by testing the method for analyzing real objects. The linearity range for ADBAC was from 0.0062 to 0.97%, for ADEBAC from 0.000726 to 0.201%, for CHG from 0.0128 to 0.111%, and for PHMB from 0.00311 to 0.0205%. The calculated relative errorfor all determined substances was about 4%.

Liquid chromatography–high-resolution electrostatic ion-trap mass spectrometric analysis of GD 3 ganglioside in dairy products

Gangliosides are thought to play a role in modifying the intestinal microflora and promoting neural development in the neonate. Milk is a good source of gangliosides, which are localised in the milkfat globule membrane. Robust quantitative analysis is required to underpin any claims made on ganglioside content in milk products. We have modified and validated our previously published method to quantify the amounts of the two predominant gangliosides classes, GD 3 and GM 3, in a range of bovine dairy products. After extraction and separation using hydrophilic high-performance liquid chromatography, an LTQ-Orbitrap™ mass spectrometer was used to extract and sum the major species of GD 3 and GM 3, generating fully resolved extracted ion current peaks for both standards and samples. The method had good repeatability (coefficient of variation 2–8%) and its results compared favourably with thin layer chromatography, a method that has been used extensively in the literature for the quantification of gangliosides in various sample matrices.

Analysis of Phospholipids in Rat Brain Using Liquid Chromatography–Mass Spectrometry

The brain is a lipid-rich organ containing complex polar lipids including phospholipids (PLs) and sphingolipids. These lipids are involved in the structure and function of cell membranes in the brain. We developed a fast and efficient liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify five different classes of PLs [Choline glycerophospholipid (consists of phosphatidyl choline and plasmenyl choline in these samples), ethanolamine glycerophospholipid (consist of phosphatidyl ethanolamine and plasmenyl ethanolamine in these samples), phosphatidyl serine, phosphatidyl inositol, and sphingomyelin] in the brain tissues of 80-day-old Wistar rats. The PLs were extracted from rat brain using chloroform/methanol/water. After separation using a hydrophilic high performance liquid chromatography column, PL-class-specific fragmentation (head group identification) with a tandem mass spectrometer in positive ion mode was utilized to measure changes in the relative concentration of the five PL classes. The advantage of this approach was its improved specificity over previously reported LC-MS methods. The method had good repeatability (coefficient of variation 3-9%, excluding phosphatidyl inositol) and recovery (92-103%) and compared well with more laborious traditional methods.

Liquid Chromatography–High‐Resolution Mass Spectrometry for Quantitative Analysis of Gangliosides

Gangliosides are a large family of glycosphingolipids that are abundant in the brain, and have been shown to affect neuronal plasticity during development, adulthood, and aging. We developed a fast, efficient, and sensitive liquid chromatography-mass spectrometry method to quantify eight different classes of gangliosides (GM(1), GM(2), GM(3), GD(3), GD(1a), GD(1b), GT(1b), GQ(1b)) in the brains of 2-day-old and 80-day-old Wistar rats. The gangliosides were extracted from rat brain using a modified Svennerholm and Fredman method. After ganglioside class separation using a hydrophilic high performance liquid chromatography (HPLC) column, the resolving power of the LTQ-Orbitrap mass spectrometer was used to extract and sum the major species of each ganglioside class, generating fully resolved extracted ion current peaks for both standards and samples. The flexibility and the specificity of this method are such that it can be applied to the analysis of other ganglioside species/classes not discussed in this paper, provided appropriate standards are available. The method had good repeatability (coefficient of variation 4.8-12.3%) and mean recoveries in the range 92-107%.

Interaction between the soluble F1 ATPase and its naturally occurring inhibitor protein. Studies using hydrophilic high-performance liquid chromatography and immunoelectron microscopy.

Binding of the isolated ATPase (F1) to its naturally occurring inhibitor protein was studied by two novel, independent techniques. High-pressure gel permeation chromatography revealed one tight binding site (Kd = 0.46 microM) for the inhibitor on F1, and a number of weak, non-specific sites. Use of an antibody directed against a non-binding region of the inhibitor protein demonstrated the formation of inhibitor/F1/immunoglobulin G complexes of 1:1:1 and 2:2:1 stoichiometry, but not of the putatively more stable cyclic 4:2:2 complexes. It was concluded that, despite the presence of three beta-subunits, only one site per F1 molecule is available for binding its inhibitor protein.