Search

A new instrument that bridges the gap between gas chromatography (GC) and liquid chromatography (LC) mass spectrometry (MS) was developed. In this instrument GC-MS and electron ionization LC-MS were combined in one MS system with method based mode changing. The LC pneumatic spray formation interface to MS was mounted on top of an otherwise unused GC detector slot and was connected with a flow restriction capillary to the MS through the GC oven and into the MS transfer line, parallel to the GC capillary column. The LC output mobile phase flow is directed into a spray formation and vaporization chamber. The pneumatic spray results in fine spray droplets that are thermally vaporized at a pressure equal to or greater than ambient. A portion of the vaporized mixture is directed into the heated flow restriction capillary that connects the spray formation and vaporization chamber into the electron ionization (EI) ion source, while most of the vaporized spray is released to the atmosphere. The combined GC-MS and LC-MS system can work either with standard EI or with cold EI via interfacing the flow restriction capillary into a supersonic nozzle forming a supersonic molecular beam of a vibrationally cold sample compound. We found that EI-LC-MS with cold EI has many benefits when compared with standard EI. The EI-LC-MS interface can also serve for flow injection analysis. The performance of the combined system is demonstrated in the analysis of a few sample mixtures by both GC-MS and LC-MS analysis, sequentially without hardware adjustments.

We report the finding of doubly charged molecular ions in a range of relatively large molecules including hydrocarbons upon their electron ionization as vibrationally cold molecules in supersonic molecular beams (SMB) (also named as Cold EI). Furthermore, we also report the detection by mass spectrometry of triply charged molecular ions in large PAHs such as decacyclene and ovalene upon their cooling in SMB. We found that the relative abundance of doubly charged molecular ions strongly depends on the internal vibrational cooling. While after some vibrational cooling the fragmentation pattern became cooling independent, the relative abundance of the doubly charged molecular ions was noticeably increased upon further cooling via increasing of the cooling make-up gas flow rate. In addition, the relative abundance of the doubly charged molecular ions was strongly increased with the compounds' size, and its electron energy threshold was lower than expected. These observations indicate a new mechanism that involves two separate electron ionization processes in the same compound, most likely with the same electron but at two separate atoms (places) in large molecules, to reduce Coulombic repulsion energy that can lead to fragmentation into two singly charged ions. These findings are shedding new light on electron ionization mass spectra. Accordingly, electron ionization mass spectra are the result of three separate mechanisms with relative magnitudes that depend on the compound size: (a) single electron ionization; (b) double electron ionization; and (c) single electron ionization with subsequent internal excitation by the same ionizing electron in another place.

Pharmaceuticals require careful and precise determination of their impurities that might harm the user upon consumption. Although today, the most common technique for impurities identification is liquid chromatography-mass spectrometry (LC-MS/MS), it has several downsides due to the nature of the ionization method. Also, the analyses in many cases are targeted thus despite being present, some of the compounds will not be revealed. In this paper, we propose and show a new method for untargeted analysis and identification of impurities in active pharmaceutical ingredients (APIs). The instrument used for these analyses is a novel electron ionization (EI) LC-MS with supersonic molecular beams (SMB). The EI-LC-MS-SMB was implemented for analyses of several drug samples spiked with an impurity. The instrument provides EI mass spectra with enhanced molecular ions, named Cold EI, which increases the identification probabilities when the compound is identified with the aid of an EI library like National Institute of Standards and Technology (NIST). We analyzed ibuprofen and its impurities, and both the API and the expected impurity were identified with names and structures by the NIST library. Moreover, other unexpected impurities were found and identified proving the ability of the EI-LC-MS-SMB system for truly untargeted analysis. The results show a broad dynamic range of four orders of magnitude at the same run with a signal-to-noise ratio of over 10 000 for the API and almost uniform response.

Electrospray LC–MS, LC–MS with Cold EI and GC–MS with Cold EI (Electron Ionization (EI) of vibrationally cold molecules in supersonic molecular beams (SMB)) are compared in their sample identification capabilities. Two test mixtures were used: (a) acetaminophen, caffeine, sulfadimethoxine, terfenadine and reserpine; and (b) octafluoronaphthalene (OFN), pyrene, cholesterol and agidol 40. The major strength of EI in both LC–MS with Cold EI and GC–MS with Cold EI is its compatibility with library based sample identification that provides samples names and structures at the isomer level. GC–MS with Cold EI alleviates two major downsides of GC–MS with standard EI as it provides enhanced molecular ions that are often weak or missing in standard EI and, it significantly extends the range of compounds that are amenable for analysis. Consequently, reserpine and agidol 40 that are incompatible with standard GC–MS analyses are analyzed by GC–MS with Cold EI and yield effective library based identification. In cases involving sample compounds that are not in the library, ESI–LC–MS provides elemental formula information (when combined with high resolution MS) while GC–MS with Cold EI also provides elemental formula when TAMI software is used for the quadrupole mass accuracy improvement combined with isotope abundance analysis. We found that both LC–MS with Cold EI and GC–MS with Cold EI have broader range of small molecules amenable for analysis than ESI–LC–MS that fails to analyze relatively non-polar compounds and exhibits highly non-uniform and compound dependent ionization yields that span over four orders of magnitude. The total ion mass chromatograms S/N of ESI–LC–MS and LC–MS with Cold EI are comparable while it is better (>100 times per on-column amount) for GC–MS with Cold EI.

We report the observation of a new physical phenomenon of the addition of 2 hydrogen atoms to molecular ions thus forming [M + 2H]+ ions. We demonstrate such second hydrogen atom abstraction onto the molecular ions of pentaerythritol and trinitrotoluene (TNT). We used both gas chromatography mass spectrometry (GC-MS) with supersonic molecular beam (SMB) with methanol added into its make-up gas and electron ionization (EI) liquid chromatography mass spectrometry (LC-MS) with SMB with methanol as the LC solvent. We found that the formation of methanol clusters resulted upon EI in the formation of dominant protonated pentaerythritol ion at m/z = 137 plus about 70% relative abundance of pentaerythritol molecular ion with 2 additional hydrogen atoms at m/z = 138 which is well above the 5.7% natural C13 isotope abundance of protonated pentaerythritol. Similarly, we found an abundant protonated TNT ion at m/z = 228 and a similar abundance of TNT molecular ion with 2 additional hydrogen atoms at m/z = 229. Upon the use of deuterated methanol (CD3 OD) as the solvent, we observed an abundant m/z = 231 (M + 2D)+ of TNT with 2 deuterium atoms. We found such abundant second hydrogen atom abstraction with butylglycolate and at low abundances in dioctylphthalate, Vitamin K3, phenazine, and RDX. At this time, we are unable to report the magnitude and frequency of occurrence of this phenomenon in standard electrospray LC-MS. This observation could have important implications on the provision of elemental formula from mass spectra that are involved with protonated molecules. Accordingly, while accurate mass measurements can serve for the generation of elemental formula, their further support and improvement via isotope abundance analysis are questionable. Consequently, if a given compound can be analyzed by both GC-MS and LC-MS, its GC-MS analysis can be superior for the provision of accurate elemental formulae if its EI mass spectrum exhibits abundant molecular ions such as with GC-MS with SMB (also known as cold EI).

A new type of electron ionization LC-MS with supersonic molecular beams (EI-LC-MS with SMB) is described. This system and its operational methods are based on pneumatic spray formation of the LC liquid flow in a heated spray vaporization chamber, full sample thermal vaporization and subsequent electron ionization of vibrationally cold molecules in supersonic molecular beams. The vaporized sample compounds are transferred into a supersonic nozzle via a flow restrictor capillary. Consequently, while the pneumatic spray is formed and vaporized at above atmospheric pressure the supersonic nozzle backing pressure is about 0.15 Bar for the formation of supersonic molecular beams with vibrationally cold sample molecules without cluster formation with the solvent vapor. The sample compounds are ionized in a fly-though EI ion source as vibrationally cold molecules in the SMB, resulting in 'Cold EI' (EI of vibrationally cold molecules) mass spectra that exhibit the standard EI fragments combined with enhanced molecular ions. We evaluated the EI-LC-MS with SMB system and demonstrated its effectiveness in NIST library sample identification which is complemented with the availability of enhanced molecular ions. The EI-LC-MS with SMB system is characterized by linear response of five orders of magnitude and uniform compound independent response including for non-polar compounds. This feature improves sample quantitation that can be approximated without compound specific calibration. Cold EI, like EI, is free from ion suppression and/or enhancement effects (that plague ESI and/or APCI) which facilitate faster LC separation because full separation is not essential. The absence of ion suppression effects enables the exploration of fast flow injection MS-MS as an alternative to lengthy LC-MS analysis. These features are demonstrated in a few examples, and the analysis of the main ingredients of Cannabis on a few Cannabis flower extracts is demonstrated. Finally, the advantages of EI-LC-MS with SMB are listed and discussed.

A new type of electron ionization LC-MS with supersonic molecular beams (EI-LC-MS with SMB) is described. This system and its operational methods are based on pneumatic spray formation of the LC liquid flow in a heated spray vaporization chamber, full sample thermal vaporization and subsequent electron ionization of vibrationally cold molecules in supersonic molecular beams. The vaporized sample compounds are transferred into a supersonic nozzle via a flow restrictor capillary. Consequently, while the pneumatic spray is formed and vaporized at above atmospheric pressure the supersonic nozzle backing pressure is about 0.15 Bar for the formation of supersonic molecular beams with vibrationally cold sample molecules without cluster formation with the solvent vapor. The sample compounds are ionized in a fly-though EI ion source as vibrationally cold molecules in the SMB, resulting in 'Cold EI' (EI of vibrationally cold molecules) mass spectra that exhibit the standard EI fragments combined with enhanced molecular ions. We evaluated the EI-LC-MS with SMB system and demonstrated its effectiveness in NIST library sample identification which is complemented with the availability of enhanced molecular ions. The EI-LC-MS with SMB system is characterized by linear response of five orders of magnitude and uniform compound independent response including for non-polar compounds. This feature improves sample quantitation that can be approximated without compound specific calibration. Cold EI, like EI, is free from ion suppression and/or enhancement effects (that plague ESI and/or APCI) which facilitate faster LC separation because full separation is not essential. The absence of ion suppression effects enables the exploration of fast flow injection MS-MS as an alternative to lengthy LC-MS analysis. These features are demonstrated in a few examples, and the analysis of the main ingredients of Cannabis on a few Cannabis flower extracts is demonstrated. Finally, the advantages of EI-LC-MS with SMB are listed and discussed.