A rapid method for simultaneous quantification of 13 sugars and sugar alcohols in food products by UPLC-ELSD.

A novel, rapid, simultaneous analysis method for five sugars (fructose, glucose, sucrose, maltose, and lactose) and eight sugar alcohols (erythritol, xylitol, sorbitol, mannitol, inositol, maltitol, lactitol, and isomalt) was developed using UPLC-ELSD, without derivatization. The analysis conditions, including the gradient conditions, modifier concentration and column length, were optimized. Thirteen sugars and sugar alcohols were separated well and the resolution of their peaks was above 1.0. Their optimum analysis condition can be analyzed within 15min. Standard curves for sugars and sugar alcohols with concentrations of 5.0-0.1% and 2.0-0.05% are presented herein, and their correlation coefficients are found to be above 0.999 and the limit of detection (LOD) was around 0.006-0.018%. This novel analysis system can be used for foodstuffs such as candy, chewing gum, jelly, chocolate, processed chocolate products, and snacks containing 0.21-46.41% of sugars and sugar alcohols.

Simultaneous determination of ethyl carbamate and urea in Korean rice wine by ultra-performance liquid chromatography coupled with mass spectrometric detection.

In this study, a rapid method for simultaneous detection of ethyl carbamate (EC) and urea in Korean rice wine was developed. To achieve quantitative analysis of EC and urea, the conditions for Ultra-performance liquid chromatography (UPLC) separation and atmospheric-pressure chemical ionization tandem mass spectrometry (APCI-MS/MS) detection were first optimized. Under the established conditions, the detection limit, relative standard deviation and linear range were 2.83µg/L, 3.75-5.96%, and 0.01-10.0mg/L, respectively, for urea; the corresponding values were 0.17µg/L, 1.06-4.01%, and 1.0-50.0µg/L, respectively, for EC. The correlation between the contents of EC and its precursor urea was determined under specific pH (3.5 and 4.5) and temperature (4, 25, and 50°C) conditions using the developed method. As a result, EC content was increased with greater temperature and lower pH. In Korean rice wine, urea was detected 0.19-1.37mg/L and EC was detected 2.0-7.7µg/L. The method developed in this study, which has the advantages of simplified sample preparation, low detection limits, and good selectivity, was successfully applied for the rapid analysis of EC and urea.

Rapid and simple extraction of lipids from blood plasma and urine for liquid chromatography-tandem mass spectrometry.

A simple and fast lipid extraction method from human blood plasma and urine is introduced in this study. The effective lipid extraction from biological systems with a minimization of the matrix effect is important for the successful qualitative and quantitative analysis of lipids in liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). The method described here is based on the modification of the quick, easy, cheap, effective, rugged and safe (QuEChERS) extraction method, which was originally developed for pesticide residue analysis in food, for the purpose of isolating lipids from biological fluids. Applicability of QuEChERS method for lipids was evaluated by varying organic solvents for the extraction/partitioning of lipids in MgSO4/CH3COONa for the removal of water and by varying sorbents (primary secondary amines, graphitized carbon black, silica, strong anion exchange resins and C18 particles) for the dispersive solid-phase extraction (dSPE) step. This study shows that 2:1 (v/v) CHCl3/CH3OH is effective in the extraction/partitioning step and that 50mg of C18 particles (for 0.1mL plasma and 1mL of urine) are more suitable for sample cleanup for the dSPE step of the QuEChERS method. Matrix effects were calculated by comparing the recovery values of lipid standards spiked to both plasma and urine samples after extraction with those of the same standards in a neat solution using nanoflow LC-ESI-MS/MS, resulting in improved MS signals due to the decrease of the ion suppression compared to the conventional Folch method. The modified QuEChERS method was applied to lipid extracts from both human urine and plasma samples, demonstrating that it can be powerfully utilized for high-speed (<15min) preparation of lipids compared to the Folch method, with equivalent or slightly improved results in lipid identification using nLC-ESI-MS/MS.

On-line two-dimensional capillary strong anion exchange/reversed phase liquid chromatography-tandem mass spectrometry for comprehensive lipid analysis.

An on-line two-dimensional liquid chromatography method was developed for comprehensive lipid profiling by coupling strong anion exchange (SAX) and nanoflow reversed-phase liquid chromatography (nRPLC) prior to electrospray ionization-tandem mass spectrometry (2D-SAX/nRPLC-ESI-MS/MS). Lipids can be classified into four different types according to the electrical propensities of the lipids: anionic, weak anionic, neutral polar, and special lipids. In 2D-SAX/nRPLC, various lipids can be fractionated in the first dimension (SAX: 5 µm to 100 Å, 5.0 cm × 75 µm i.d.) by step elution (methanol and salt solution), followed by the molecular separation of lipids in the second dimension (RP: 3 µm to 100 Å, 7.0 cm × 75 µm i.d.) with binary gradient LC. Since the elution of lipids from SAX can be achieved with a very small volume of eluent delivered from an autosampler, it can be simply implemented with an LC-ESI-MS instrument for full automation, and the salt step elution, including the two-step injection procedure, can be used for the selective analysis of the desired lipid fraction. For nRPLC-ESI-MS/MS run in either positive or negative ion mode, a common ionization modifier (0.05% ammonium hydroxide with 5mM ammonium formate) was introduced into the binary mobile phase solutions so that 2D-LC-MS could be operated in both ion modes without changing the mobile phase solutions. The developed on-line 2D-SAX/nRPLC-ESI-MS/MS was evaluated with 22 different standard lipids for the optimization of the salt step elution and was applied to a healthy human plasma lipid extract, resulting in the identification of a total of 303 plasma lipids, including 14 different classes.

Effect of ionization modifiers on the simultaneous analysis of all classes of phospholipids by nanoflow liquid chromatography/tandem mass spectrometry in negative ion mode.

The effect of ionization modifiers added to the mobile phase of nanoflow liquid chromatography-tandem mass spectrometry (nLC-ESI-MS³) on the simultaneous analysis of all phospholipid (PL) classes in negative ion mode has been investigated. While MS analysis of most PL classes is carried out in negative ion mode, analysis of neutral polar (polar but electrically neutral) lipids like phosphatidylcholine (PC) and sphingomyelin (SM) is highly efficient in positive ion mode. Therefore, analysis of PL mixture samples often requires two separate runs in both positive and negative ion mode. In order to establish run conditions to carry out a single nLC-ESI-MS-MS for all PLs, the ionization efficiency of 13 different types of PL molecules in nLC-ESI-MS has been evaluated in negative ion mode by varying the modifiers and their concentrations. Experiments demonstrated that a mixture of 0.05% ammonium hydroxide and 1 mM ammonium formate added to the mobile phase provided effective ionization for all classes of PLs. The optimized conditions were applied to the analysis of a phospholipid mixture extracted from a human urine sample, yielding the identification of a total of 85 PL species. Analysis of the same sample with dual nLC-ESI-MS² runs in both positive and negative ion mode confirmed that nLC-ESI-MS³ with the mixed modifier run only in negative ion mode gave comparable results.

Characterization of functionalized styrene-butadiene rubber by flow field-flow fractionation/light scattering in organic solvent.

Flow field-flow fractionation (FlFFF) using an organic solvent as mobile phase has been effectively utilized for the separation and characterization of functionalized styrene-butadiene rubbers (SBR) that are polymerized and followed by coupling reaction in solution. Separation of broad molecular weight SBR was accomplished by an asymmetrical FlFFF channel in THF under field programming and the molecular weight distribution (MWD) of the SBR sample was determined by on-line measurement of light scattering. In this study, FlFFF has been utilized to characterize high-MW functionalized SBR from the low-MW non-functionalized molecules which were used for coupling reaction to produce high-MW functionalized SBRs, and to determine the coupling number of the functionalized SBRs depending on the type of the coupling reagents. The resulting MWD of the SBR samples prepared by the different coupling reagents (SnCl(4) and a polydimethylsiloxane compound) were compared.

Shotgun analysis of phospholipids from mouse liver and brain by nanoflow liquid chromatography/tandem mass spectrometry.

It was demonstrated that a shotgun approach can be utilized for the characterization of phospholipids (PLs) extracted from mouse liver and brain by using nanoflow reversed phase liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS). In this study, a dual scan method was introduced for the high throughput analysis of complex PL mixtures. Two consecutive LC-ESI-MS-MS runs were made in positive ion mode (for phosphatidylcholines (PCs) and phosphatidylethanolamines (PEs)) first followed by analysis in negative ion mode (for phosphatidylserine (PSs) and phosphatidylinositol (PIs)) using the same binary gradient elution with and without adding formic acid, respectively. The separation of the PLs was carried out using a home made pull tip capillary column (C18) with an end frit. The MS analysis of the eluted PL molecules was performed with a precursor scan followed by a data dependent MS-MS scan. The developed dual scan method was tested with the extracts of PCs and PIs mixtures from soybean, PEs from Escherichia coli, and PSs from bovine brain. It was further applied for the characterization of intact PL samples that were extracted from both mouse liver and mouse brain in the laboratory, and resulted in the identification of 90 and 80 PL species, respectively.

Nanoflow liquid chromatography-tandem mass spectrometry for the characterization of intact phosphatidylcholines from soybean, bovine brain, and liver.

Nanoflow liquid chromatography-electrospray ionization tandem-mass spectrometry (nanoLC-ESI-MS-MS) was applied for the characterization of intact phosphatidylcholine (PC) lipid molecules using a homemade reversed phase capillary column with a pulled tip for direct ESI at positive ion mode. Prior to the analytical column, a short capillary trapping column was utilized for on-line pre-concentration via microcross connection. Separation of intact phosphatidylcholines in the nanoflow LC column was carried out using a binary gradient elution method at 300 nL/min. The structures of the eluted PC components were determined by analysis of the typical fragment ions of PC molecules obtained from collision-induced dissociation (CID) after each precursor scan in mass spectrometry. In the current study, nanoflow LC-ESI-MS-MS analysis of PC molecules demonstrated the ability to obtain clear structural information, such as alkyl chain lengths and the degree of unsaturation with a protonated molecule ([M + H]+) and its characteristic fragment ions ([M + H-RCH2COOH]+, [M + H-RCH=C=O]+, and [M + H-184]+). Results from the nanoflow LC-ESI-MS experiment showed the limit of detection at 3.5 fmol for the 14:0/14:0-PC standard. This technique then was applied to intact PC extracts from soybean, bovine brain, and liver without derivatization and resulted in the identification of 28, 25, and 39 phosphatidylcholines, respectively. The LC-MS-MS method has been shown to be useful for the analysis of low concentration PC molecules in biological samples.