Particle Beam Liquid Chromatography-mass Spectrometry Research Articles

Selenium speciation by liquid chromatography-particle beam/mass spectrometry (LC-PB/MS): application to a yeast reference material and synthetic urine

Presented here is the comprehensive speciation of organic and inorganic selenium species via liquid chromatography-particle beam/electron impact mass spectrometry (LC-PB/EIMS). Two different matrices (SELM-1 selenium-enriched yeast certified reference material and synthetic urine) were analyzed for the content of methionine and selenium species utilizing this analytical tool. An ion-paring reversed-phase chromatographic separation using a linear gradient of water and methanol (each containing 0.1% trifluoroacetic acid), at 0.9 mL min−1 with an analysis time of less than 20 minute, was employed for the selenium speciation. Electron ionization mass spectra yielded clear and simple fragmentation patterns of the chemical species eluting from the chromatographic column. Quantification by a standard addition method was accomplished for methionine (m/z = 149 Da) and selenomethionine (m/z = 196 Da) with recoveries of 93% (9% RSD, n = 4) and 97% (11% RSD, n = 4), respectively. Absolute limits of detection on the nanogram level were obtained for each of the targeted species.

Analysis of caffeic acid derivatives in echinacea extracts by liquid chromatography particle beam mass spectrometry (LC–PB/MS) employing electron impact and glow discharge ionization sources

A liquid chromatography-particle beam/mass spectrometry (LC-PB/MS) method with electron impact (EI) and glow discharge (GD) ionization sources is presented for the determination of caffeic acid derivatives in echinacea tinctures. In this work, two commercially available echinacea ethanolic extracts were used as the test samples for the separation, identification, and quantification of the caffeic acid derivatives (caffeic acid, chlorogenic acid, cichoric acid, and caftaric acid), which are suggested to have beneficial medicinal properties. Detailed evaluations of the two primary controlling parameters for EI (electron energy and source block temperature) and GD (discharge current and pressure) sources were performed to determine optimal instrument operation conditions. The mass spectra obtained from both ion sources provide clear and simple molecular fragmentation patterns for each of the target analytes. The absolute detection limits for the caffeic acid derivatives were determined to be at subnanogram levels for both the EI and GD sources. The separation of the caffeic acid derivatives in echinacea was accomplished by reversed-phase chromatography using a C(18) column and a gradient elution system of water containing 0.1% trifluoroacetic acid and methanol, with an analysis time of less than 40 min. A standard addition method was employed for the quantification of each of the caffeic acid derivatives in the tincture.

Quantitative determination of 13C-labeled and endogenous beta-carotene, lutein, and vitamin A in human plasma.

Quantitative procedures employing liquid-chromatography/particle beam-mass spectrometry (LC/PB-MS) and gas chromatography-mass spectrometry (GC-MS) were applied to the determination of the endogenous and 13C-labeled beta-carotene, lutein, and retinol in plasma of a subject who consumed kale (Brassica oleracea) that had been grown in a 13CO2-enriched atmosphere. All compounds were analyzed in the negative chemical ionization (NCI) mode using methane as the moderating reagent gas. Beta-carotene and lutein were analyzed using LC/PB-MS applying reversed-phase high-performance liquid chromatography (HPLC) separation procedures to resolve the analytes. The concentrations of the beta-carotene isotopomers in the plasma over a several-week period were determined using 2H8-beta-carotene as an internal standard. The total plasma concentrations of all trans-lutein were quantified by HPLC analysis with a photodiode array detector using beta-apo-8'-carotenal as an internal standard, and the ratio of the 13C:12C isotopomers of lutein was determined by PB-MS. The retinol isotopomers were collected from individual HPLC fractions of the plasma extract and then analyzed as the trimethylsilyl ethers by GC-MS in the NCI mode. The 13C- and 12C-retinol isotopomers were quantified using 2H4-retinol as an internal standard. These methods demonstrate the application of highly sensitive procedures employing NCI MS for the quantitative determination of carotenoids and vitamin A for the purpose of conducting metabolism studies of phytonutrients.

The use of particle beam mass spectrometry for the measurement of impurities in a nabumetone drug substance, not easily amenable to atmospheric pressure ionisation techniques.

Liquid chromatography/particle beam mass spectrometry (LC/PB-MS) was used for the structural elucidation of some impurities in nabumetone as this compound poorly ionises by atmospheric pressure ionisation (API) techniques. PB-MS was optimised for nabumetone and a sensitivity study was carried out. To obtain full scan electron ionisation spectra a minimum of 100 ng of compound on column was needed. By using 20 mg/mL solutions of nabumetone, impurities at levels of about 250 ppm mass fraction relative to nabumetone could be detected. Results were compared with LC/API-MS and previous GC/MS.

A sensitive procedure for the study of β-carotene-d8 metabolism in humans using high performance liquid chromatography–mass spectrometry

This report describes the development of a robust method of high sensitivity for studying the metabolism of β-carotene-d8 in humans using a combination of liquid chromatography/particle beam–mass spectrometry (LC/PB-MS). The utility of the LC/PB-MS method was demonstrated in a pilot study. The carotenoids were extracted from plasma into hexane and separated by reverse phase high performance liquid chromatography (HPLC) using a C-18 column. The HPLC effluent was nebulized using helium and the solvent was removed under vacuum within the dual-stage particle beam interface. The de-solvated carotenoids were ionized in the negative-ion mode (electron capture) using methane chemical ionization and detected using selected ion monitoring. The limit of detection of the method was on the order of 0.3 ng (approximately 0.6 pmol) for β-carotene. β-Carotene-d8 was quantified in the plasma over a concentration range of two orders of magnitude using β-carotene-13C40 as an internal standard. The overall coefficient of variance (CV) for determining the concentration of the analytes from 30 μl of plasma was 3.9% for β-carotene and 2.4% for β-carotene-d8.Using the LC/PB-MS method, the concentration of β-carotene-d8 was determined in the plasma of a subject who had consumed a single 5-mg dose over a 30-day period. The sensitive semiautomated procedure is capable of high sample throughput and makes large comprehensive studies feasible.

Liquid chromatography mass spectrometry with supersonic molecular beams

A new approach for liquid chromatography mass spectrometry (LC-MS) is described, based on achieving soft thermal vaporization followed by supersonic expansion and direct sample compound ionization, while in a supersonic molecular beam (SMB). The soft molecular vaporization step utilizes spray formation that is continued by fast thermal vaporization inside a channel supersonic nozzle, followed by ultrafast supercooling in a supersonic expansion. The short time (several microseconds) spent by the vaporized compound in the heated nozzle prior to its expansion cooling may result in incomplete vibrational equilibrium and thus reduced degree of dissociation. In addition, even if vibrational equilibrium at the nozzle temperature is obtained, the sample compounds have significantly reduced time for their dissociation, which is thus further minimized (kinetic consideration). As soon as the molecules expand and form a SMB, they are supercooled and any further dissociation is avoided. While in the SMB, the sample molecules can be ionized either by electron ionization as described in this paper or by hyperthermal surface ionization. The major goal of this method is to obtain high quality library searchable electron ionization mass spectra, for a broad range of thermally labile compounds, with higher sensitivity than that achievable by particle beam LC-MS. The soft thermal vaporization nozzle is described and mass spectral results with corticosterone are demonstrated. The potential advantageous features of this new method are discussed.

DROPLET ELECTROSPRAY NEBULIZATION FOR PARTICLE BEAM LC-MS

A new interface for particle beam liquid chromatography mass spectrometry (PB-LC-MS) was developed based on a droplet electrospray nebulizer (DESN). The DESN produced relatively monodisperse droplet aerosols of approximately 100 μm diameter. An efficient aerosol desolvation and transport system was also designed to optimize analyte mass transport efficiency. The DESN and desolvation system was coupled to an ion trap mass spectrometer via a three stage momentum separator (DESN-PB-LC-ITMS). The DESN provided several advantages over other nebulization techniques for PB-LC-MS. First, the resultant larger particles provided a high mass transport efficiency of analytes through the momentum separator, which resulted in a significant improvement of the sensitivity and calibration linearity of PB-LC-MS. Detection limits (3σ) ranging from 5.6 to 5.8 pg/μL were obtained for the tested compounds. Secondly, the lower dispersity of larger aerosols provided a better control of particle beam formation and significantly improved the peak shapes. The DESN method resulted in narrower and more symmetric peaks than those obtained with a thermospray nebulizer for PB-LC-ITMS. Compared with UV absorbance detection, no extra peak broadening, peak tailing, or asymmetry were observed with the DESN-PB-LC-ITMS method. Thirdly, the droplet aerosols were produced by applying a DC voltage to the electrode, which was easy to operate. In addition, the DESN could be operated well in both aqueous and organic solvents at flow rates up to 1 mL/min. The system design, optimization, and features of DESN-PB-LC-ITMS have been described in detail.

Analytical specificity and optimization of particle‐beam measurements for some biopharmaceutical applications using a hildebrand nebulizer

The present study shows how specific and selective particle-beam liquid chromatography mass spectrometry (PB-LC/MS) can be for certain biopharmaceutical applications and by which strategy maximum sensitivity can be achieved. The utility of PB-LC/MS in the analysis of different pharmaceutical drugs and endogenic substances with different molecular structures has been investigated. Using the PB interface in combination with a triple quadrupole mass spectrometer, the effects of mobile phase composition and flow rate, interface parameters---such as desolvation temperature and pressure of the nebulization gas---and the temperature of the ion source, on detection limits and sensitivity, are demonstrated. A particular substance specificity of this interface type for creatinine, naloxone and methadone could be observed after optimizing the transmission of the interface. The best sensitivity is achieved only in a narrow working range, indicating the necessity of an individual optimization of interface conditions for each substance. Linearity, sensitivity and precision of transfer rate using the EI and selected-ion monitoring modes were demonstrated. Investigation of the so-called “carrier effect” on the transfer of the selected compounds indicated that the addition of ammonium acetate as “carrier” changed the detection sensitivity in a completely different order. Copyright © 1998 John Wiley & Sons, Ltd.

Analytical specificity and optimization of particle-beam measurements for some biopharmaceutical applications using a hildebrand nebulizer

The present study shows how specific and selective particle-beam liquid chromatography mass spectrometry (PB-LC/MS) can be for certain biopharmaceutical applications and by which strategy maximum sensitivity can be achieved. The utility of PB-LC/MS in the analysis of different pharmaceutical drugs and endogenic substances with different molecular structures has been investigated. Using the PB interface in combination with a triple quadrupole mass spectrometer, the effects of mobile phase composition and flow rate, interface parameters---such as desolvation temperature and pressure of the nebulization gas---and the temperature of the ion source, on detection limits and sensitivity, are demonstrated. A particular substance specificity of this interface type for creatinine, naloxone and methadone could be observed after optimizing the transmission of the interface. The best sensitivity is achieved only in a narrow working range, indicating the necessity of an individual optimization of interface conditions for each substance. Linearity, sensitivity and precision of transfer rate using the EI and selected-ion monitoring modes were demonstrated. Investigation of the so-called “carrier effect” on the transfer of the selected compounds indicated that the addition of ammonium acetate as “carrier” changed the detection sensitivity in a completely different order. Copyright © 1998 John Wiley & Sons, Ltd.

Quantitative analysis of poly(methyl methacrylate–butyl acrylate) copolymer composition by liquid chromatography–particle beam-mass spectrometry

Current polymer formulations contain mixtures of copolymers to tailor the performance needs. Reversed-phase liquid chromatography is commonly used to separate polymers according to their chemical composition by adsorption, partition or precipitation mechanisms if retention is not influenced by molecular mass. Liquid chromatography–mass spectrometry (LC–MS) has mainly been used to identify low molecular mass polymers and additives. In this paper, we report on the use of LC–MS for the quantitative analysis of copolymer composition of several high-molecular-mass polymers by monitoring the low-mass fragments formed by thermal decomposition and electron impact ionization when using a particle beam interface. The fragment ions produced are proportional to the comonomers present and are quantitatively related to the copolymer composition. Area ratio calibration with copolymers of known composition is used to determine the composition of unknown copolymers of similar structure.

Confirmation of multiple tetracycline residues in milk and oxytetracycline in shrimp by liquid chromatography–particle beam mass spectrometry

A confirmation procedure is described for detection of residues of six tetracyclines in bovine milk, and oxytetracycline in shrimp. Residues are extracted from milk or shrimp tissue using metal chelate affinity chromatography. The extracts are desalted, further concentrated using polymeric solid-phase extraction, and chromatographed on a polymeric reversed-phase column. Analysis is by methane negative ion chemical ionization on a quadrupole mass spectrometer using a particle beam interface. Data are acquired in partial scan mode, monitoring from m/ z 378 to m/ z 480. The procedure was validated with control milk and shrimp, fortified milk (30 ng/ml) and shrimp (100 ng/g), and milk and tissue from animals treated with the drugs.

Identification of pesticides by liquid chromatography–particle beam mass spectrometry using electron ionization and chemical ionization

Liquid chromatography–mass spectrometry (LC–MS) with a particle beam (PB) interface is used to separate and identify a group of pesticides. The mass spectra obtained under the different ionization modes, electron ionization (EI) and positive and negative chemical ionization (PCI and NCI) are compared. The operating conditions under each mode, determined by studying the influence on the ion abundance of the ion source temperature of the EI mode, and the gas pressure and ion source temperature in the methane CI were optimized. EI was more sensitive than PCI and NCI and of the latter two modes, NCI gave higher responses, especially for organophosphorus compounds. When on-line solid-phase extraction–LC–PB-MS was applied to real samples, limits of detection in full scan mode were in the range of 0.5 and 10 μg l −1 for EI. The analysis of real samples by on-line solid-phase extraction–LC–PB-MS enabled EI detection of one of the pesticides studied and confirmation by PCI and NCI. The combined EI/CI information also enabled the detection of some non-target compounds.

Liquid Chromatography/Particle Beam/Mass Spectrometry Determination of Carbamates in Lettuce and Apple

A method for extraction and determination of trace amounts of Carbaryl, Baygon, Methiocarb, and Methiocarb sulfoxide in lettuce and apple samples is described. The arrangement liquid chromatography-particle beam-mass spectrometry offers the possibility of confirmation and determination of these carbamates at concentrations lower than those admitted by the European Union. Recoveries of carbamates at the 0.16–4.9 mg/kg level were between 53 and 76% with standard deviation between 6 and 12% (n = 5). Detection limits for analytes single ion monitoring mode were 5, 30, 10, and 20 ng for Methiocarb sulfoxide, Baygon, Carbaryl, and Methiocarb, respectively.

Analysis of secreted flavonoids of Cistus ladanifer L. by high-performance liquid chromatography–particle beam mass spectrometry

The analysis of secreted flavonoids of the leaf Cistus ladanifer was carried out by high-performance liquid chromatography–mass spectrometry coupled through a particle beam interface. The mass spectra were recorded using electron impact. Apigenin, 4′-methyl-apigenin, 7-methyl-apigenin, 7,4′-dimethyl-apigenin, 3-methyl-kaempferol, 3,4-dimethyl-kaempferol, 3,7-dimethyl-kaempferol and 3,7,4′-trimethyl-kaempferol were identified in the leaf resin.

Liquid chromatography-particle-beam mass spectrometric determination of β-carotene by negative-ion chemical ionization

The first method for liquid chromatography-particle-beam mass spectrometry (LC-PBMS) of β-carotene is described. It uses a reversed-phase (RP) HPLC column and methanol-dichloromethane, 90:10, as mobile phase. For comparison purposes, UV-visible detection was performed at 436 nm to determine linearity, sensitivity and precision; the absolute limit of detection of the LC-UV method was estimated to be 0.2 ng. The possible modes of ionization of β-carotene were explored for PBMS detection; the best selectivity and sensitivity were obtained by use of electron-capture negative-ion chemical ionization (NCI). When operating in NCI mode with selected ion monitoring (SIM), the detection limit was 0.3 ng for β-carotene, comparable with that of the UV detector. The relative standard deviation (RSD) of the LC-PBMS system was 2.6 % at the 10 ng level (n=10). For the determination of β-carotene in real samples absolute calibration and calibration by the method of standard addition were used for both UV and PBMS detection.

Identification of hexahydroimidazo[1,2- a]pyrazine-3,6-diones and hexahydroimidazo[1,2- a]imidazo[1,2- d]pyrazine-3,8-diones, unusual products of silica-catalyzed amino acid thermal condensation and products of their thermal decomposition using coupled high-performance liquid chromatography–particle beam mass spectrometry and gas chromatography–Fourier transform infrared spectroscopy–mass spectrometry

Sublimation of simple aliphatic amino acids (Ala, Aib, Val and Leu) at 230–250°C under reduced pressure in the presence of silica as a catalyst yields cyclic dipeptides piperazine-2,5-diones as major products. In addition, two types of unusual products have been detected. To determine their structures, we utilized a coupled HPLC–particle beam-MS and GC–Fourier transform IR–MS technique with auxiliary computer simulation of IR spectra. Based on the spectral data obtained, we identified the condensation products as substituted bicyclic and tricyclic amidines, hexahydroimidazo[1,2- a]pyrazine-3,6-diones and hexahydroimidazo[1,2- a]imidazo-[1,2- d]pyrazine-3,8-diones. Other peaks in the chromatograms has been identified as products of the amidines thermal decomposition. The general decomposition pattern includes dehydrogenation as well as cleavage of the carbon skeleton. The last process primarily affects the 6-membered pyrazine ring causing elimination of CO or HNCO, or/and the loss of α-substituents without or with α-carbon atom.

Determination of acidic chlorinated pesticides in water: Comparison of official EPA method 515.1 and liquid-solid extraction HPLC/UV and HPLC/PBMS analyses

A comparison of the official EPA method 515.1 for determination of chlorinated acidic pesticides and a modification of it is illustrated. Extraction of the analytes from water and their determination and quantitation is by gas chromatography-electron capture detection (GC-ECD), liquid chromatography-UV detection and liquid chromatography-particle beam mass spectrometry. Although HPLC-PBMS was found to be less sensitive than the GC-ECD method, it was, nevertheless, more sensitive than HPLC-UV. The modified method is simpler, quicker and allows more accurate determination of pesticides in aqueous samples.

Fast, very fast, and ultra-fast gas chromatography-mass spectrometry of thermally labile steroids, carbamates, and drugs in supersonic molecular beams

Gas chromatography-mass spectrometry (GC-MS) analyses of thermally labile compounds have been studied by using a short column fast gas chromatograph, coupled with fly-through electron ionization in supersonic molecular beams. Thirty-two compounds, which include steroids, carbamate pesticides, antibiotic drugs, and other pharmaceutical compounds, have been analyzed and the details of their GC-MS analysis are provided. The ability to analyze thermally labile compounds is discussed in relation to the speed of analysis. A new term, "speed enhancement factor" (SEF), is defined as the product of column length reduction and the carrier gas linear velocity increase, as compared with normal GC-MS conditions. Fast, very fast, and ultra-fast GC-MS are defined with a SEF in the ranges of 5-30, 30-400, and 400-4000, respectively. Trade-offs in the degree of dissociation, speed, gas chromatograph resolution, and sensitivity were studied and examined with thermally labile molecules. The experimental factors that affect the dissociation are described with emphasis on its reduction. We claim that the use of supersonic molecular beams for sampling and ionization provides the ultimate capability in the GC-MS of thermally labile compounds. The obtained 70-eV electron ionization mass spectra are shown, and an enhanced relative abundance of the molecular ion is demonstrated together with library search capability of these mass spectra, which is better than that reported with particle beam liquid chromatography-mass spectrometry. The performance of fast GC-MS in supersonic molecular beams is compared with other methods of fast GC-MS and with particle beam liquid chromatography-mass spectrometry.

Simultaneous identification of different classes of hydrocarbons and determination of nitro-polycyclic aromatic hydrocarbons by means of particle beam liquid chromatography-mass spectrometry

LC-MS with a particle beam interface was used to separate and identify aliphatic hydrocarbons, polycyclic aromatic hydrocarbons (PAHs) and their nitrated derivatives (nitro-PAHs). In particular, the behaviour of nitro-PAHs and of two nitrated biphenyls was checked in terms of instrumental response. A normal-phase chromatographic system was used, with a silica column and heptane-tetrahydrofuran as mobile phase. Electron impact and positive- and negative-ion chemical ionization (NICI) modes were used; for nitro-PAHs, the best results were obtained under NICI conditions. In the flow-injection mode, detection limits as low as pg levels were obtained for 9-nitroanthracene, 3-nitro-9-fluorenone, 1-nitropyrene, 1,8-dinitronaphthalene, 2,7-dinitrofluorene and 2,7-dinitro-9-fluorenone, whereas the low-molecular-mass 1- and 2-nitronaphthalene were not detected even at μg levels; using the analytical column, a detection limit five times higher was determined for 1-nitropyrene. For most of the nitro-PAHs considered, the dependences of the intensities on the amounts injected were found to be linear in a range of more than two orders of magnitude.

Liquid chromatography particle beam-mass spectrometry with massive cluster impact

A new liquid chromatography-mass spectrometry technique is described that utilizes a particle beam interface to transport and deposit desolvated analyte molecules onto a target surface that is bombarded by a primary beam of massive multiply charged glycerol cluster ions to generate secondary ions for mass analysis. The massive cluster ion beam is generated by electrohydrodynamic emission from a solution of 1. 5-M ammonium acetate in glycerol. In the present instrumental configuration the massive cluster ion gun is placed above the target probe and the particle beam interface is connected through a side port of the mass spectrometer. The massive cluster ion beam and particle beam are intercepted by a target surface substituted for a conventional ion volume. The target surface is positioned such that it is ∼ 45 ° to the primary cluster ion beam, the particle beam, and the mass analyzer axis. This geometric orientation represents a compromise among the performances of these three elements.The feasibility of this liquid chromatography-particle beam on-line with massive cluster impact is demonstrated by flow injections of acetylcholine chloride and gramicidin S. Spectra generated from this preliminary study indicate promise for routine liquid chromatography-mass spectrometry of polar compounds by using a robust inlet and an effective generation of secondary ions without an added matrix.