Femtosecond Laser Ionization Mass Spectrometry for Online Analysis of Human Exhaled Breath.

A variety of organic compounds in human exhaled breath were measured online by mass spectrometry using the fifth (206 nm) and fourth (257 nm) harmonic emissions of a femtosecond ytterbium (Yb) laser as the ionization source. Molecular ions were enhanced significantly by means of resonance-enhanced, two-color, two-photon ionization, which was useful for discrimination of analytes against the background. The limit of detection was 0.15 ppm for acetone in air. The concentration of acetone in exhaled breath was determined for three subjects to average 0.31 ppm, which lies within the range of normal healthy subjects and is appreciably lower than the range for patients with diabetes mellitus. Many other constituents, which could be assigned to acetaldehyde, ethanol, isoprene, phenol, octane, ethyl butanoate, indole, octanol, etc., were observed in the exhaled air. Therefore, the present approach shows potential for use in the online analysis of diabetes mellitus and also for the diagnosis of various diseases, such as COVID-19 and cancers.

Machine Learning for Characterizing Biofuels Based on Femtosecond Laser Ionization Mass Spectrometry.

A sample mixture of fatty acid methyl esters (FAMEs) was measured by femtosecond laser ionization mass spectrometry (fsLIMS) using the fifth (206 nm) and fourth (257 nm) harmonic emissions of an ytterbium (Yb) laser (1030 nm). Molecular ions were observed as the major signals in this technique, providing valuable information concerning the molecular weight and the number of double bonds in the molecule. The mass spectral data were then used as explanatory variables in machine learning based on artificial intelligence (AI) to correlate with objective variables such as the cetane number, kinematic viscosity, specific gravity, a higher heating value, an iodine value, flash point, oxidative stability index, and a cloud point measured for reference biofuel samples containing various FAMEs. The properties of biofuels, i.e., the objective variables, were evaluated from the mass spectral data obtained for unknown samples. The errors in the evaluation were a few percent when the distribution of the FAMEs in the unknown biofuel sample was similar to those of the biofuels used for machine learning. As demonstrated herein, the present approach, involving a combination of fsLIMS and AI, has the potential for use in evaluating the properties of a biofuel and then in solving of environmental issues associated with global warming.

Determination of fatty acid methyl esters by two-color two-photon resonance-enhanced femtosecond ionization mass spectrometry.

A standard sample mixture containing thirty-seven fatty acid methyl esters (FAMEs) was measured by femtosecond laser ionization mass spectrometry. FAME molecules with double bonds were efficiently ionized via resonance-enhanced two-photon ionization by absorbing the first photon at 206 nm at the edge of the absorption band of the π→π* transition and subsequently ionized by absorbing the second photon at 257 nm. The intensity of the molecular radical ion was enhanced significantly using this two-color ionization scheme, which minimizes the excess energy in the ionized state, when compared with electron ionization mass spectrometry and vacuum-ultraviolet photoionization mass spectrometry. This approach was then used for the reliable identification of FAMEs contained in an actual sample of biofuel.

Trace analysis of nitrated polycyclic aromatic hydrocarbons based on two-color femtosecond laser ionization mass spectrometry.

Nitrated polycyclic aromatic hydrocarbons (nitro-PAHs) are suspected to be highly carcinogenic and mutagenic compounds that are present in the environment. Gas chromatography combined with mass spectrometry (GC-MS) is the most frequently used technique for trace analysis. The electron ionization techniques that are currently used in MS, however, typically do not result in the formation of a molecular ion, thus making the determination of these compounds more difficult. In this study, we report on the use of a compact highly-repetitive (low-pulse-energy) ultraviolet (UV) femtosecond laser as the ionization source in combination with a miniature time-of-flight mass analyzer and a time-correlated ion counting system. The UV laser pulses emitted at 343, 257, and 206 nm were produced by harmonic generations of a femtosecond Yb laser emitting at 1030 nm and were utilized for single-color multiphoton ionization. A combination of the 343-nm and 257-nm pulses was further employed to achieve two-color two-photon ionization. This technique was found to be more useful for sensitive detection and also resulted in the formation of a molecular ion. A pump-and-probe technique using these pulses was examined in a proof-of-concept study to measure the femtosecond lifetimes of the nitro-PAHs separated by GC, providing additional information for use in the characterization of the analyte. The developed technique was applied in the analysis of an authentic sample, an organic solvent extract from diesel exhaust particulates. The nitro-PAHs contained in a standard reference material (SRM1975) were determined on a two-dimensional GC-MS display, suggesting that this technique would be useful for the practical trace analysis of nitro-PAHs in environmental samples.

Determination of Barbiturates by Femtosecond Ionization Mass Spectrometry.

Barbiturates are highly susceptible to dissociation in mass spectrometry (MS) because of their long side chains combined with a nonaromatic ring consisting of several carbonyl and amine groups. As a result, they exhibit extensive α-cleavage and subsequent rearrangement, making the identification of these compounds difficult. Although a library of electron ionization MS (EIMS) is available, most barbiturates have very similar fragment patterns. Accordingly, it would be desirable to develop a technique for soft ionization, providing a molecular ion and large fragment ions as well. In this study, a molecular ion was clearly observed, in addition to large fragment ions, for a variety of barbiturates based on multiphoton ionization MS (MPIMS) using a tunable ultraviolet femtosecond laser as the ionization source (fs-LIMS). This favorable result was achieved when the optimal laser wavelength for minimizing the excess energy remaining in the ionic state was used. An examination of the photofragmentation pathways suggested that an H atom in the side chain was abstracted by an oxygen atom in the carbonyl group in the ring structure thus initiating fragmentation and subsequent rearrangement. Barbiturates that are substituted with alkyl groups (amobarbital and pentobarbital) had narrower spectral regions for optimal ionization than the other barbiturates with alkyl and alkenyl groups (butalbital and secobarbital) and more with alkyl and phenyl groups (phenobarbital). All of the barbiturates studied provided unique mass spectral patterns in fs-LIMS, which was useful for the reliable identification of these compounds in practical trace analysis.

Miniature time-of-flight mass analyzer for use in combination with a compact highly-repetitive femtosecond laser ionization source.

In most cases, a molecular ion is observed in femtosecond laser ionization mass spectrometry, which provides information concerning the molecular weight of the analyte. However, the Ti:sapphire laser currently used as the ionization source is costly and involves special skills for operation and maintenance, which prevents its practical use in many applications. In this study, we report on the development of a miniature time-of-flight mass analyzer with a flight tube length of 65 mm for use in combination with a compact highly-repetitive (120-560 kHz) femtosecond Yb laser and a time-correlated single ion counting system. The fundamental beam (1030 nm) was converted into ultraviolet beams emitting at 343, 257, and 206 nm, which was utilized as an efficient two-photon ionization source. A mass resolution of 670 was achieved for the molecular ion of chlorobenzene, the minimum time for measuring a mass spectrum being 0.1 s. This mass spectrometer was used in the on-site real-time monitoring of products appeared by the combustion of plastic, some nerve agent analogs, and an explosive in the air. The interference arising from nitrogen and oxygen in the air was suppressed, since they require nonresonant four- and three-photon ionizations, respectively. The mass spectrometer was combined with a gas chromatograph and used for the comprehensive analysis of polycyclic aromatic hydrocarbons, suggesting its potential advantage for use in the practical trace analysis of organic compounds in the environmental and forensic sciences.

Comprehensive Analysis of Analogues of Amine-Related Psychoactive Substances Using Femtosecond Laser Ionization Mass Spectrometry.

Amine-related psychoactive molecules contain N-Cα and Cα-Cß bonds, which easily dissociate to form various fragment ions in electron ionization mass spectrometry (EIMS). Therefore, observing a molecular ion and then determining the molecular weight of the analyte is difficult. In this study, we examined phenethylamine, 3,4-methylenedioxyphenethylamine, tryptamine, N-methylephedrine, and nicotine as well as analogues of amine-related psychoactive substances using EIMS and femtosecond laser ionization mass spectrometry (fs-LIMS) combined with gas chromatography for comprehensive analysis. A molecular ion was clearly observed in fs-LIMS for all of these compounds, which was in contrast to EIMS providing fragment ions dominantly (no molecular ion was observed for N-methylephedrine). This favorable result was obtained by adjusting the laser wavelength to the optimal value for two-photon ionization to minimize the excess energy remaining in the molecular ion. It therefore appears that fs-LIMS is superior to EIMS in terms of observing a molecular ion and would be potentially useful for identifying a variety of amine-related psychoactive substances, some of which are illegal and are of interest in the field of forensic science.

Deep- to near-ultraviolet Raman frequency conversion pumped by femtosecond pulses in a hollow-core waveguide.

Broadband vibrational/rotational Raman generation ranging from deep ultraviolet (DUV) to blue wavelengths is demonstrated by using molecular hydrogen in a hollow-core waveguide as a Raman-active medium pumped by a femtosecond DUV laser. We find the high-order transient stimulated Raman scattering is drastically enhanced for input beams including a circularly polarized component; a circularly polarized input beam achieves the highest conversion efficiency. Coherent vibrational anti-Stokes Raman emission is observed only for a circularly polarized pump beam, indicating that the waveguide effect also contributes to the upconversion of a DUV pulse via transient stimulated Raman scattering.

Esterification of perfluorinated carboxylic acids with bromomethyl aromatic compounds for gas chromatography combined with laser ionization mass spectrometry.

Perfluorinated carboxylic acids (PFCAs) were derivatized with two types of aromatic compounds that contained a bromomethyl group, i.e., 2-(bromomethyl)naphthalene (BMN) and benzyl bromide (BB). The conditions for derivatization were optimized in terms of reaction temperature and time and the concentration of derivatizing reagent. Using these optimal conditions, the PFCAs-MN and PFCAs-B derivatives were measured by gas chromatography (GC) combined with mass spectrometry using an ultraviolet femtosecond laser (267 nm) as the ionization source. The efficiency of derivatization for PFCAs-B was higher than that for PFCAs-MN because of the smaller size of the chromophore (benzene). The ionization efficiency of PFCAs-MN, however, was better than PFCAs-B, since a larger sized chromophore (naphthalene) and then a larger molar absorptivity was preferable for resonance-enhanced two-photon ionization. Due to superior GC separation, BB was successfully used as the derivatizing agent for the trace analysis of PFCAs, with detection limits of 6.0, 8.4, and 9.5 ng/mL for perfluoroheptanoic, perfluorooctanoic, and perfluorononanoic acids, respectively. The other bromomethyl aromatic compounds were evaluated for use as a derivatization reagent in future studies.

Femtosecond ionization mass spectrometry for chromatographic detection.

Mass spectrometry is now in widespread use for the detection of the analytes separated by chromatography. Electron ionization is the most frequently used method in mass spectrometry. However, this ionization technique sometimes suffers from extensive fragmentation of analytes, which makes identification difficult. A photoionization technique has been developed for suppressing this fragmentation and for subsequently observing a molecular ion. A variety of lasers have been employed for the sensitive and selective ionization of organic compounds. A femtosecond laser has a high peak power and is preferential for efficient ionization as well as for suppressing fragmentation, providing valuable information concerning molecular weight and chemical structure as well. In this review, we report on applications of femtosecond ionization mass spectrometry combined with gas chromatography.

Suppression of Fragmentation in Mass Spectrometry.

Suppressing fragmentation is a constant challenge in mass spectrometry because a molecular ion can readily be identified and provides information concerning the molecular weight of an analyte. Several techniques such as charge exchange chemical ionization (CECI) and vacuum ultraviolet emission ionization (VUVEI) have been developed to date for achieving this purpose. In this study, we report on the use of tunable ultraviolet (UV) and near-infrared (NIR) femtosecond (fs) lasers (35 fs) for the multiphoton ionization (MPI) of cis- and trans-4-methylcyclohexanols, the reference molecules that are currently used to examine fragmentation suppression. The results obtained here were compared with those obtained by CECI and VUVEI because they were reported as the best techniques for suppressing fragmentation. A molecular ion was strongly enhanced by carefully minimizing the excess energy in the ionic state using tunable UV and NIR fs-lasers. The ratio of the intensities for molecular and fragment ions, [M]+/[M-H2O]+, increased significantly (9.5-fold and 8.5-fold for cis- and trans-isomers, respectively, in UV fs-MPI) compared to the values obtained by CECI and VUVEI, respectively.

Single-Photon Ionization Mass Spectrometry Using a Vacuum Ultraviolet Femtosecond Laser.

The wavelength of a femtosecond Ti:sapphire laser (TS, 800 nm) was converted into the ultraviolet (UV, 200 nm) using three ß-barium borate crystals (ß-BaB2O4) for frequency doubling and subsequent mixing. The UV pulse was further converted into the vacuum ultraviolet (VUV, 185 nm) based on four-wave Raman mixing, in which a two-color pump beam consisting of the fundamental beam (800 nm) of the TS and the signal beam of an optical parametric amplifier (1200 nm) pumped by the TS was focused onto a capillary waveguide filled with hydrogen gas for molecular phase modulation and the single-color UV probe beam (200 nm) was then focused onto the waveguide for frequency modulation to generate anti-Stokes and high-order Stokes Raman sidebands at wavelengths of 185 and 218-267 nm, respectively. The efficiency of conversion from the UV (200 nm) to the VUV (185 nm) was 6%. The ionization energy was calculated for 13 amino polycyclic aromatic hydrocarbons using density functional theory, since they are associated with the development of occupational bladder cancers. The values calculated by the B3LYP/cc-pVDZ and ωB97Xd/cc-pVTZ methods were 6.24-7.14 eV (199-174 nm) and 6.41-7.35 eV (194-169 nm), respectively. A sample containing a mixture of 9-aminoanthracene, 3-aminofluoranthene, and 1-aminopyrene was separated by gas chromatography (GC), and the eluents were ionized with the VUV pulse (0.015 µJ) in mass spectrometry (MS). The analytes were observed on a two-dimensional display of GC/MS, and the detection limit obtained by single-photon ionization of 3-aminofluoranthene was 1 ng/µL.

Generation of a Nearly Monocycle Optical Pulse in the Near-Infrared Region and Its Use as an Ionization Source in Mass Spectrometry.

We report on the generation of an octave-spanning (600-1400 nm) nearly monocycle (1.1 cycle) ultrashort optical pulse (3.2 fs) in the near-infrared region by the Fourier synthesis of two pulses at 800 and 1200 nm, both of which were spectrally broadened by self-phase modulation and were compressed by chirp mirrors. The 3.2 fs pulse was converted into the ultraviolet by third harmonic generation, the pulse width being evaluated to 1.9 fs. The near-infrared pulse (3.2 fs) was employed as an ionization source in mass spectrometry, and the signal intensity was significantly increased for pentachlorobenzene, an environmental pollutant listed in the Stockholm Convention. The present data and the spectral properties obtained by quantum chemical calculations suggest that the method offers a potential advantage for the detection of Novichok, a chemical warfare agent that is thought to have been used in a terrorist attack.

Time-correlated Single Ion Counting Mass Spectrometer with Long and Short Time-of-Flight Tubes and an Evaluation of Its Performance for Use in Trace Analysis of Allergenic Substances.

An analyte molecule was ionized using a femtosecond laser as the ionization source and was measured by a twin-type time-of-flight mass spectrometer with long (42 cm) and short (6.4 cm) flight tubes. The signal was measured using an analog signal digitizer and a time-correlated single ion counting system, and performance was evaluated by comparing data obtained from both instruments. The short mass spectrometer had a mass resolution of 450 and was used in the trace analysis of allergenic substances in a fragrance.

Resonant and non-resonant femtosecond ionization mass spectrometry of organochlorine pesticides.

Thirteen organochlorine pesticides in a standard sample mixture were measured by gas chromatography combined with mass spectrometry using an ultraviolet femtosecond laser (267 nm) as the ionization source, and the observed mass spectra were compared with the corresponding spectra obtained using an electron ionization source. When an ultrashort optical pulse was used for ionization, molecular ions were typically produced which was preferential for reliably identifying the analytes. The ionization mechanism was studied based on three models constructed for resonance-enhanced two-photon ionization, non-resonant two-photon ionization, and non-resonant three-photon ionization. The optimal conditions for observing a molecular ion were investigated using data obtained for three pulse widths. The results suggest that two-photon ionization with minimum excess energy would be optimal for observing a molecular ion.

Determination of nerve agent metabolites in human urine by femtosecond laser ionization mass spectrometry using 2-(bromomethyl)naphthalene as a derivatizing reagent.

Nerve agent metabolites (NAMs) derived from alkyl methyl phosphonic acids, such as ethyl methylphosphonic acid (EMPA), isopropyl methylphosphonic acid (IMPA), and pinacolyl methylphosphonic acid (PMPA), were extracted from human urine using diethyl ether as an extractant. After exchanging the diethyl ether solvent to acetonitrile, the analytes were derivatized with 2-(bromomethyl)naphthalene (BMN). The reaction products of the BMN and NAMs, i.e., MN-EMPA, MN-IMPA, and MN-PMPA, were separated by gas chromatography (GC) and measured by mass spectrometry (MS) using a femtosecond laser emitting at 267 nm as the ionization source for resonance-enhanced two-photon ionization (RE2PI). The limits of detection (LOD) were <1 ng/mL for these analytes. The use of BMN increased the volatility of the analytes for separation by GC and also increased the ionization efficiency via the RE2PI process as the result of presence of a naphthalene functional group. A two-dimensional GC-MS display can be used for comprehensive analysis of NAMs, by-products, and impurities in the sample. Then, this approach could be used to confirm the use of chemical weapons and for forensic identification.

Determination of nitrated polycyclic aromatic hydrocarbons in particulate matter 2.5 by laser ionization mass spectrometry using an on-line chemical-reduction system.

Nitrated polycyclic aromatic hydrocarbons (NPAHs) were separated by gas chromatography (GC) and were introduced into a reactor consisting of an inner-seal Y-connector for the on-line chemical reduction of NPAH using hydrazine monohydrate as a chemical reducing agent. The reduced forms of amino polycyclic aromatic hydrocarbons (APAHs) were then determined by multiphoton ionization time-of-flight mass spectrometry (MPI/TOF-MS) using a femtosecond laser as the ionization source. Three standard samples of NPAHs, i.e., 9-nitroanthracene (9-NANT), 3-nitrofluoranthene (3-NFLU), and 1-nitropyrene (1-NPYR), were confirmed to be completely reduced to the corresponding APAHs, i.e., 9-aminoanthracene (9-AANT), 3-aminofluoranthene (3-AFLU), and 1-aminopyrene (1-APYR). The sensitivity of the method was improved by 2-8 fold by converting the NPAHs to APAHs. This analytical technique was applied to a sample extracted from PM2.5 collected in Fukuoka. The signal enhancement and background reduction by the chemical reaction resulted in the NPAHs in the actual sample being more efficiently measured, and the concentrations of 9-NANT, 3-NFLU, and 1-NPYR were determined to be 25, 305, and 217 pg m-3, respectively.

Determination of Pesticides by Gas Chromatography Combined with Mass Spectrometry Using Femtosecond Lasers Emitting at 267, 400, and 800 nm as the Ionization Source.

A standard sample mixture containing 51 pesticides was separated by gas chromatography (GC), and the constituents were identified by mass spectrometry (MS) using femtosecond lasers emitting at 267, 400, and 800 nm as the ionization source. A two-dimensional display of the GC/MS was successfully used for the determination of these compounds. A molecular ion was observed for 38 of the compounds at 267 nm and for 30 of the compounds at 800 nm, in contrast to 27 among 50 compounds when electron ionization was used. These results suggest that the ultraviolet laser is superior to the near-infrared laser for molecular weight determinations and for a more reliable analysis of these compounds. In order to study the conditions for optimal ionization, the experimental data were examined using the spectral properties (i.e., the excitation and ionization energies and absorption spectra for the neutral and ionized species) obtained by quantum chemical calculations. A few molecules remained unexplained by the currently reported rules, requiring additional rules for developing a full understanding of the femtosecond ionization process. The pesticides in the homogenized matrix obtained from kabosu ( citrus sphaerocarpa) were measured using lasers emitting at 267 and 800 nm. The pesticides were clearly separated and measured on the two-dimensional display, especially for the data measured at 267 nm, suggesting that this technique would have potential for use in the practical trace analysis of the pesticides in the environment.

Optimal Laser Wavelength for Femtosecond Ionization of Polycyclic Aromatic Hydrocarbons and Their Nitrated Compounds in Mass Spectrometry.

The ionization and fragmentation processes were examined for a standard sample mixture containing 16 polycyclic aromatic hydrocarbons (PAHs) and 3 nitro-PAHs (NPAHs) by gas chromatography combined with mass spectrometry (GC/MS) using a femtosecond laser emitting at 400, 800, or 1200 nm as the ionization source. The signal intensities of NPAHs were lower and the fragmentation more extensive compared to those values for PAHs, especially at shorter wavelengths (400 nm). These results can be explained by efficient intersystem crossing to triplet levels and the shorter excited-state lifetimes of neutral NPAHs molecules, compared to the pulse width of the laser. Fragmentation was significantly suppressed by nonresonant multiphoton ionization when a laser emitting at longer wavelengths (1200 nm) was used. This result can be explained by the absorption spectrum of the molecular ion and the excess energy remaining in the ionized state. In fact, there was no absorption band at 1200 nm for the molecular ion, and the excess energy would be minimal when a near-infrared laser is used, which suppresses the fragmentation even for NPAHs. A doubly charged molecular ion was observed for PAHs but not for NPAHs, probably owing to the higher stability of the PAH molecule, the electrons of which are more strongly bound and are more resistive to field ionization. To demonstrate the utility of this technique, the sample extracted from particulate matter 2.5 (PM2.5) emitted from a diesel engine was measured. NPAHs as well as PAHs were clearly determined at 1200 nm, at which the background signal arising from the interference was drastically suppressed.

Use of chemical conversion for determination of nitrated aromatic hydrocarbons using femtosecond ionization mass spectrometry.

A sample containing nitrated aromatic hydrocarbons (NPAHs) was injected into the sample inlet port of a gas chromatograph (GC), along with hydrazine, a reducing reagent. The analytes that eluted from the GC column were measured by mass spectrometry (MS) using an ultraviolet femtosecond laser as the ionization source. When no reducing reagent was used, large numbers of polycyclic aromatic hydrocarbons (PAHs) including NPAHs were observed in the two-dimensional GC/MS display. In contrast, when hydrazine was present, reduced forms of NPAHs, which included amino PAHs, were detected. When a palladium or platinum catalyst was placed in the GC inlet port, the compounds were further reduced to non-aromatic hydrocarbons. The present approach would be useful for studies to evaluate the chemical reaction that converts the constituents contained in exhaust emitted from a diesel engine.