Sub-100 µm Spatial Resolution Ambient Mass Spectrometry Imaging of Rodent Brain with Laser Ablation Atmospheric Pressure Photoionization (LAAPPI) and Laser Ablation Electrospray Ionization (LAESI).

In this study, we applied a new IR laser-beam-focusing technique to enable sub-100 µm spatial resolution in laser ablation atmospheric pressure photoionization (LAAPPI) and laser ablation electrospray ionization (LAESI) mass spectrometry imaging (MSI). After optimization of operational parameters, both LAAPPI- and LAESI-MSI with a spatial resolution of 70 µm produced high-quality MS images, which allowed accurate localization of metabolites and lipids in the mouse and rat brain. Negative and positive ion LAAPPI- and LAESI-MS detected many of the same metabolites and lipids in the brain. Many compounds were also detected either by LAAPPI- or LAESI-MS, indicating that LAAPPI and LAESI are more complementary than alternative methods.

Desorption Atmospheric Pressure Photoionization Coupled with Ion Mobility-Mass Spectrometry.

Desorption atmospheric photoionization (DAPPI) is an ambient mass spectrometry (MS) technique that can be used to analyze both polar and nonpolar compounds. Here, the coupling of DAPPI with traveling wave ion mobility-mass spectrometry (TWIM-MS) and application to analysis of food, multivitamin, and pharmaceutical products is described.

Chemical profiles of birch and alder bark by ambient mass spectrometry.

Desorption atmospheric pressure photoionization (DAPPI) is an ambient mass spectrometry (MS) technique that allows the analysis of both polar and nonpolar compounds directly from the surfaces of various sample types. Here, DAPPI was used to study the chemical profiles in different parts of birch and alder tree barks. Four distinct fractions of Betula pendula (silver birch) bark were collected from three different developmental stages of the stem, after which the chemical profiles of the different tissue types were measured. Of special interest were triterpenoids, a class of important defensive substances, which are found in the bark of the silver birch. Additionally, the chemical profiles of lenticels and the surrounding surfaces in the phellem of B. pendula (silver birch), Alnus glutinosa (black alder), and Alnus incana (gray alder) were screened with DAPPI. Another ambient MS technique, laser ablation atmospheric pressure photoionization (LAAPPI), was further used for the mass spectrometry imaging of lenticels on the B. pendula phellem. All the studied birch bark fractions showed individual chemical profiles in DAPPI. The mass spectra from the young apical stem and the transition zone resembled each other more than the mature stem. Instead, the phellem was found to contain a high amount of triterpenoids in all the developmental stages of the stem. The most intense peaks in the DAPPI mass spectra of the birch bark fractions were those of betulin and lupeol. Betulinic and betulonic acid peaks were intense as well, and these compounds were detected especially in the lenticels of the tree samples. Graphical abstract.

Tissue-specific study across the stem reveals the chemistry and transcriptome dynamics of birch bark.

Tree bark is a highly specialized array of tissues that plays important roles in plant protection and development. Bark tissues develop from two lateral meristems; the phellogen (cork cambium) produces the outermost stem-environment barrier called the periderm, while the vascular cambium contributes with phloem tissues. Although bark is diverse in terms of tissues, functions and species, it remains understudied at higher resolution. We dissected the stem of silver birch (Betula pendula) into eight major tissue types, and characterized these by a combined transcriptomics and metabolomics approach. We further analyzed the varying bark types within the Betulaceae family. The two meristems had a distinct contribution to the stem transcriptomic landscape. Furthermore, inter- and intraspecies analyses illustrated the unique molecular profile of the phellem. We identified multiple tissue-specific metabolic pathways, such as the mevalonate/betulin biosynthesis pathway, that displayed differential evolution within the Betulaceae. A detailed analysis of suberin and betulin biosynthesis pathways identified a set of underlying regulators and highlighted the important role of local, small-scale gene duplication events in the evolution of metabolic pathways. This work reveals the transcriptome and metabolic diversity among bark tissues and provides insights to its development and evolution, as well as its biotechnological applications.

Solid Sampling with a Diode Laser for Portable Ambient Mass Spectrometry.

A hand-held diode laser is implemented for solid sampling in portable ambient mass spectrometry (MS). Specifically, a pseudocontinuous wave battery-powered surgical laser diode is employed for portable laser diode thermal desorption (LDTD) at 940 nm and compared with nanosecond pulsed laser ablation at 2940 nm. Postionization is achieved in both cases using atmospheric pressure photoionization (APPI). The laser ablation atmospheric pressure photoionization (LAAPPI) and LDTD-APPI mass spectra of sage leaves (Salvia officinalis) using a field-deployable quadrupole ion trap MS display many similar ion peaks, as do the mass spectra of membrane grown biofilms of Pseudomonas aeruginosa. These results indicate that LDTD-APPI method should be useful for in-field sampling of plant and microbial communities, for example, by portable ambient MS. The feasibility of many portable MS applications is facilitated by the availability of relatively low cost, portable, battery-powered diode lasers. LDTD could also be coupled with plasma- or electrospray-based ionization for the analysis of a variety of solid samples.

A Simple Method for Improving the Spatial Resolution in Infrared Laser Ablation Mass Spectrometry Imaging.

In mass spectrometry imaging of tissues, the size of structures that can be distinguished is determined by the spatial resolution of the imaging technique. Here, the spatial resolution of IR laser ablation is markedly improved by increasing the distance between the laser and the focusing lens. As the distance between the laser and the lens is increased from 1 to 18 m, the ablation spot size decreases from 440 to 44 µm. This way, only the collimated center of the divergent laser beam is directed on the focusing lens, which results in better focusing of the beam. Part of the laser energy is lost at longer distance, but this is compensated by focusing of the radiation to a smaller area on the sample surface. The long distance can also be achieved by a set of mirrors, between which the radiation travels before it is directed to the focusing lens and the sample. This method for improving the spatial resolution can be utilized in mass spectrometry imaging of tissues by techniques that utilize IR laser ablation, such as laser ablation electrospray ionization, laser ablation atmospheric pressure photoionization, and matrix-assisted laser desorption electrospray ionization. Graphical Abstract ᅟ.

Recent developments in atmospheric pressure photoionization-mass spectrometry.

Recent developments in atmospheric pressure photoionization (APPI), which is one of the three most important ionization techniques in liquid chromatography-mass spectrometry, are reviewed. The emphasis is on the practical aspects of APPI analysis, its combination with different separation techniques, novel instrumental developments - especially in gas chromatography and ambient mass spectrometry - and the applications that have appeared in 2009-2014. © 2015 Wiley Periodicals, Inc. Mass Spec Rev 36:423-449, 2017.

Profiling of Coumarins in Peucedanum palustre (L.) Moench Populations Growing in Finland.

The coumarin composition of Peucedanum palustre (L.) Moench populations growing in Finland was investigated. A total of 132 flowering P. palustre specimens from 43 locations in southern and central Finland were collected, divided into root, stem, leaf, and umbel samples, and analyzed by HPLC. HPLC coupled to high-resolution mass spectrometry was used to aid the identification of coumarins. A total of 13 coumarin-structured compounds were quantitatively analyzed from the samples. The coumarin profile of root samples was found to differ from the aerial plant parts. The main coumarins in roots were oxypeucedanin and columbianadin. In aerial parts, peulustrin isomers were the most abundant coumarin components. Umbels and leaves also contained a considerable amount of umbelliprenin, which was only found in traces in roots. Based on hierarchical cluster analysis of the coumarin profiles, some populations shared common characteristics. The most distinct property connecting certain populations was their high peulustrin content. Another notable common property between some populations was the high umbelliprenin content in aerial plant parts. Some populations were clustered together due to their low overall coumarin content.

Charge Exchange Reaction in Dopant-Assisted Atmospheric Pressure Chemical Ionization and Atmospheric Pressure Photoionization.

The efficiencies of charge exchange reaction in dopant-assisted atmospheric pressure chemical ionization (DA-APCI) and dopant-assisted atmospheric pressure photoionization (DA-APPI) mass spectrometry (MS) were compared by flow injection analysis. Fourteen individual compounds and a commercial mixture of 16 polycyclic aromatic hydrocarbons were chosen as model analytes to cover a wide range of polarities, gas-phase ionization energies, and proton affinities. Chlorobenzene was used as the dopant, and methanol/water (80/20) as the solvent. In both techniques, analytes formed the same ions (radical cations, protonated molecules, and/or fragments). However, in DA-APCI, the relative efficiency of charge exchange versus proton transfer was lower than in DA-APPI. This is suggested to be because in DA-APCI both dopant and solvent clusters can be ionized, and the formed reagent ions can react with the analytes via competing charge exchange and proton transfer reactions. In DA-APPI, on the other hand, the main reagents are dopant-derived radical cations, which favor ionization of analytes via charge exchange. The efficiency of charge exchange in both DA-APPI and DA-APCI was shown to depend heavily on the solvent flow rate, with best efficiency seen at lowest flow rates studied (0.05 and 0.1 mL/min). Both DA-APCI and DA-APPI showed the radical cation of chlorobenzene at 0.05-0.1 mL/min flow rate, but at increasing flow rate, the abundance of chlorobenzene M(+.) decreased and reagent ion populations deriving from different gas-phase chemistry were recorded. The formation of these reagent ions explains the decreasing ionization efficiency and the differences in charge exchange between the techniques. Graphical Abstract ᅟ.

Thin-Layer Chromatography/Desorption Atmospheric Pressure Photoionization Orbitrap Mass Spectrometry of Lipids.

Desorption atmospheric pressure photoionization (DAPPI) allows surface analysis in the open atmosphere and is thus an appropriate method for the direct coupling of thin-layer chromatography (TLC) and mass spectrometry (MS). Here, the capability of DAPPI-MS for ionizing and detecting lipids, namely, cholesterol, triacylglycerols, 1,2-diol diesters, wax esters, cholesteryl esters, and hydrocarbons, from TLC and high-performance thin-layer chromatography (HPTLC) plates in MS and MS2 modes was tested. Limits of detection for lipid standards separated using normal-phase (NP)-TLC and NP-HPTLC were established. TLC/DAPPI-MS was applied for lipids of vernix caseosa, a white creamy proteolipid biofilm that progressively coats the fetus during the last trimester of the pregnancy, and plant oils including caraway, parsley, safflower, and jojoba oils. Various lipids were identified by means of high resolution/accurate mass measurement of Orbitrap and comparison of the retardation factors with standards. Lipid class separation was carried out on the NP-HPTLC plates, whereas individual triacylglycerol and wax ester species were separated on the reversed-phase HPTLC plates. DAPPI-MS was found to be a simple, rapid, and efficient approach for detecting lipids separated by TLC.

Nucleophilic Aromatic Substitution Between Halogenated Benzene Dopants and Nucleophiles in Atmospheric Pressure Photoionization.

In a preceding work with dopant assisted-atmospheric pressure photoionization (DA-APPI), an abundant ion at [M + 77](+) was observed in the spectra of pyridine and quinoline with chlorobenzene dopant. This contribution aims to reveal the identity and route of formation of this species, and to systematically investigate structurally related analytes and dopants. Compounds containing N-, O-, and S-lone pairs were investigated with APPI in the presence of fluoro-, chloro-, bromo-, and iodobenzene dopants. Computational calculations on a density functional theory (DFT) level were carried out to study the reaction mechanism for pyridine and the different halobenzenes. The experimental and computational results indicated that the [M + 77](+) ion was formed by nucleophilic aromatic ipso-substitution between the halobenzene radical cation and nucleophilic analytes. The reaction was most efficient for N-heteroaromatic compounds, and it was weakened by sterical effects and enhanced by resonance stabilization. The reaction was most efficient with chloro-, bromo-, and iodobenzenes, whereas with fluorobenzene the reaction was scarcely observed. The calculated Gibbs free energies for the reaction between pyridine and the halobenzenes were shown to increase in the order I < Br < Cl < F. The reaction was found endergonic for fluorobenzene due to the strong C-F bonding, and exergonic for the other halobenzenes. For fluoro- and chlorobenzenes the reaction was shown to proceed through an intermediate state corresponding to [M + dopant](+), which was highly stable for fluorobenzene. For the bulkier bromine and iodine, this intermediate did not exist, but the halogens were shown to detach already during the approach by the nucleophile.

Desorption atmospheric pressure photoionization high-resolution mass spectrometry: a complementary approach for the chemical analysis of atmospheric aerosols.

RATIONALE: On-line chemical characterization methods of atmospheric aerosols are essential to increase our understanding of physicochemical processes in the atmosphere, and to study biosphere-atmosphere interactions. Several techniques, including aerosol mass spectrometry, are nowadays available, but they all suffer from some disadvantages. In this research, desorption atmospheric pressure photoionization high-resolution (Orbitrap) mass spectrometry (DAPPI-HRMS) is introduced as a complementary technique for the fast analysis of aerosol chemical composition without the need for sample preparation. METHODS: Atmospheric aerosols from city air were collected on a filter, desorbed in a DAPPI source with a hot stream of toluene and nitrogen, and ionized using a vacuum ultraviolet lamp at atmospheric pressure. To study the applicability of the technique for ambient aerosol analysis, several samples were collected onto filters and analyzed, with the focus being on selected organic acids. To compare the DAPPI-HRMS data with results obtained by an established method, each filter sample was divided into two equal parts, and the second half of the filter was extracted and analyzed by liquid chromatography/mass spectrometry (LC/MS). RESULTS: The DAPPI results agreed with the measured aerosol particle number. In addition to the targeted acids, the LC/MS and DAPPI-HRMS methods were found to detect different compounds, thus providing complementary information about the aerosol samples. CONCLUSIONS: DAPPI-HRMS showed several important oxidation products of terpenes, and numerous compounds were tentatively identified. Thanks to the soft ionization, high mass resolution, fast analysis, simplicity and on-line applicability, the proposed methodology has high potential in the field of atmospheric research.

Analysis of neonicotinoids from plant material by desorption atmospheric pressure photoionization-mass spectrometry.

RATIONALE: Neonicotinoids are widely used insecticides which have been shown to affect the memory and learning abilities of honey bees, and are suspected to play a part in the unexplainable, large-scale loss of honey bee colonies. Fast methods, such as ambient mass spectrometry (MS), for their analysis from a variety of matrices are necessary to control the use of forbidden products and study the spreading of insecticides in nature. METHODS: The feasibilities of two ambient MS methods, desorption electrospray ionization (DESI) and desorption atmospheric pressure photoionization (DAPPI), for the analysis of five most used neonicotinoid compounds, thiacloprid, acetamiprid, clothianidin, imidacloprid and thiamethoxam, were tested. In addition, DAPPI was used to analyze fresh rose leaves treated with commercially available thiacloprid insecticide and dried and powdered turnip rape flowers, which had been collected from a field treated with thiacloprid-containing insecticide. RESULTS: DAPPI was found to be more sensitive than DESI, with 2-11 times better signal-to-noise ratios, and limits of detection at 0.4-5.0 fmol for the standard compounds. DAPPI was able to detect thiacloprid from the rose leaves even 2.5 months after the treatment and from the turnip rape flower samples collected from a field. The analysis of plant material by DAPPI did not require extraction or other sample preparation. CONCLUSIONS: DAPPI was found to be suitable for the fast and direct qualitative analysis of thiacloprid neonicotinoid from plant samples. It shows promise as a fast tool for screening of forbidden insecticides, or studying the distribution of insecticides in nature.

The detection and mapping of the spatial distribution of insect defense compounds by desorption atmospheric pressure photoionization Orbitrap mass spectrometry.

Many insects use chemicals synthesized in exocrine glands and stored in reservoirs to protect themselves. Two chemically defended insects were used as models for the development of a new rapid analytical method based on desorption atmospheric pressure photoionization-mass spectrometry (DAPPI-MS). The distribution of defensive chemicals on the insect body surface was studied. Since these chemicals are predominantly nonpolar, DAPPI was a suitable analytical method. Repeatability of DAPPI-MS signals and effects related to non-planarity and roughness of samples were investigated using acrylic sheets uniformly covered with an analyte. After that, analytical figures of merit of the technique were determined. The spatial distribution of (E)-1-nitropentadec-1-ene, a toxic nitro compound synthesized by soldiers of the termite Prorhinotermes simplex, was investigated. Then, the spatial distribution of the unsaturated aldehydes (E)-hex-2-enal, (E)-4-oxohex-2-enal, (E)-oct-2-enal, (E,E)-deca-2,4-dienal and (E)-dec-2-enal was monitored in the stink bug Graphosoma lineatum. Chemicals present on the body surface were scanned along the median line of the insect from the head to the abdomen and vice versa, employing either the MS or MS(2) mode. In this fast and simple way, the opening of the frontal gland on the frons of termite soldiers and the position of the frontal gland reservoir, extending deep into the abdominal cavity, were localized. In the stink bug, the opening of the metathoracic scent glands (ostiole) on the ventral side of the thorax as well as the gland reservoir in the median position under the ventral surface of the anterior abdomen were detected and localized. The developed method has future prospects in routine laboratory use in life sciences.

Transmission mode desorption atmospheric pressure photoionization.

RATIONALE: Desorption atmospheric pressure photoionization (DAPPI) is an ambient mass spectrometry (MS) technique that is suitable for the direct analysis of polar and nonpolar compounds from a variety of surfaces. Conventional DAPPI uses reflection geometry, but here transmission mode (TM)-DAPPI is introduced for fast and easy analysis of liquid samples. METHODS: Stainless steel and PEEK meshes were used as sampling support in TM-DAPPI. The sample was applied either in the form of a droplet on the mesh, or by dipping the mesh in the sample solution. Physical parameters affecting the ionization efficiency were optimized for TM-DAPPI. The mesh materials were used to extract compounds from aqueous samples, which were then analyzed by TM-DAPPI. TM-DAPPI and conventional DAPPI were compared. RESULTS: In TM-DAPPI, intense signals for the analytes were achieved with less heating power, and lower nebulizer gas and dopant flow rates than optimally used in conventional DAPPI. Either due to this, or the different sample support material used, a much lower background and improved sensitivity compared to conventional DAPPI was achieved. The analytes could be extracted and concentrated from liquid samples on the mesh material used in TM-DAPPI, which was especially efficient for the nonpolar benzo[a]pyrene. This effect was utilized in the analysis of triacylglycerols from cow milk. CONCLUSIONS: While conventional DAPPI is still the method of choice for solid samples, TM-DAPPI can be utilized as a fast, easily automated method for analyzing liquid samples. The mesh materials can be utilized for extraction of low polarity compounds, such as steroid hormones or PAHs from dilute, aqueous solutions, followed by subsequent analysis by TM-DAPPI.

Feasibility of desorption atmospheric pressure photoionization and desorption electrospray ionization mass spectrometry to monitor urinary steroid metabolites during pregnancy.

Steroids have important roles in the progress of pregnancy, and their study in maternal urine is a non-invasive method to monitor the steroid metabolome and its possible abnormalities. However, the current screening techniques of choice, namely immunoassays and gas and liquid chromatography-mass spectrometry, do not offer means for the rapid and non-targeted multi-analyte studies of large sample sets. In this study, we explore the feasibility of two ambient mass spectrometry methods in steroid fingerprinting. Urine samples from pregnant women were screened by desorption electrospray ionization (DESI) and desorption atmospheric pressure photoionization (DAPPI) Orbitrap high resolution mass spectrometry (HRMS). The urine samples were processed by solid phase extraction for the DESI measurements and by enzymatic hydrolysis and liquid-liquid-extraction for DAPPI. Consequently, steroid glucuronides and sulfates were detected by negative ion mode DESI-HRMS, and free steroids by positive ion mode DAPPI-HRMS. In DESI, signals of eleven steroid metabolite ions were found to increase as the pregnancy proceeded, and in DAPPI ten steroid ions showed at least an order of magnitude increase during pregnancy. In DESI, the increase was seen for ions corresponding to C18 and C21 steroid glucuronides, while DAPPI detected increased excretion of C19 and C21 steroids. Thus both techniques show promise for the steroid marker screening in pregnancy.

Ionization of EPA contaminants in direct and dopant-assisted atmospheric pressure photoionization and atmospheric pressure laser ionization.

Seventy-seven EPA priority environmental pollutants were analyzed using gas chromatography-mass spectrometry (GC-MS) equipped with an optimized atmospheric pressure photoionization (APPI) and an atmospheric pressure laser ionization (APLI) interface with and without dopants. The analyzed compounds included e.g., polycyclic aromatic hydrocarbons (PAHs), nitro compounds, halogenated compounds, aromatic compounds with phenolic, acidic, alcohol, and amino groups, phthalate and adipatic esters, and aliphatic ethers. Toluene, anisole, chlorobenzene, and acetone were tested as dopants. The widest range of analytes was ionized using direct APPI (66/77 compounds). The introduction of dopants decreased the amount of compounds ionized in APPI (e.g., 54/77 with toluene), but in many cases the ionization efficiency increased. While in direct APPI the formation of molecular ions via photoionization was the main ionization reaction, dopant-assisted (DA) APPI promoted ionization reactions, such as charge exchange and proton transfer. Direct APLI ionized a much smaller amount of compounds than APPI (41/77 compounds), showing selectivity towards compounds with low ionization energies (IEs) and long-lived resonantly excited intermediate states. DA-APLI, however, was able to ionize a higher amount of compounds (e.g. 51/77 with toluene), as the ionization took place entirely through dopant-assisted ion/molecule reactions similar to those in DA-APPI. Best ionization efficiency in APPI and APLI (both direct and DA) was obtained for PAHs and aromatics with O- and N-functionalities, whereas nitro compounds and aliphatic ethers were the most difficult to ionize. Halogenated aromatics and esters were (mainly) ionized in APPI, but not in APLI.

Solvent jet desorption capillary photoionization-mass spectrometry.

A new ambient mass spectrometry method, solvent jet desorption capillary photoionization (DCPI), is described. The method uses a solvent jet generated by a coaxial nebulizer operated at ambient conditions with nitrogen as nebulizer gas. The solvent jet is directed onto a sample surface, from which analytes are extracted into the solvent and ejected from the surface in secondary droplets formed in collisions between the jet and the sample surface. The secondary droplets are directed into the heated capillary photoionization (CPI) device, where the droplets are vaporized and the gaseous analytes are ionized by 10 eV photons generated by a vacuum ultraviolet (VUV) krypton discharge lamp. As the CPI device is directly connected to the extended capillary inlet of the MS, high ion transfer efficiency to the vacuum of MS is achieved. The solvent jet DCPI provides several advantages: high sensitivity for nonpolar and polar compounds with limit of detection down to low fmol levels, capability of analyzing small and large molecules, and good spatial resolution (250 µm). Two ionization mechanisms are involved in DCPI: atmospheric pressure photoionization, capable of ionizing polar and nonpolar compounds, and solvent assisted inlet ionization capable of ionizing larger molecules like peptides. The feasibility of DCPI was successfully tested in the analysis of polar and nonpolar compounds in sage leaves and chili pepper.

Separation of isomeric amines with ion mobility spectrometry.

Eight selected isomeric amines were ionized using atmospheric pressure chemical ionization and atmospheric pressure photoionization producing a protonated molecule [M+H](+) for each amine. The mobility of these ions was measured by ion mobility spectrometry. The amine compound class was shown to have an important role in mobility separation of the amines. 2,4,6-collidine, N,N-dimethylaniline and N-methyl-o-toluidine with highest observed mobilities have a N-heterocyclic aromatic ring, or are tertiary or secondary amines, respectively, whereas the rest of the compounds with lower mobilities were primary amines. It is suggested that the protonated -NH2 group (-NH3(+)) interacts more with the drift gas, and therefore the primary amines have lower mobilities. The effect of the drift gas was tested by mixing argon or helium with the nitrogen drift gas. The presence of argon shifted the mobilities towards lower values, while with helium the mobility shifted towards higher values. However, in neither case did this result in better separation of the unresolved compounds.

Laser ablation atmospheric pressure photoionization mass spectrometry imaging of phytochemicals from sage leaves.

RATIONALE: Despite fast advances in ambient mass spectrometry imaging (MSI), the study of neutral and nonpolar compounds directly from biological matrices remains challenging. In this contribution, we explore the feasibility of laser ablation atmospheric pressure photoionization (LAAPPI) for MSI of phytochemicals in sage (Salvia officinalis) leaves. METHODS: Sage leaves were studied by LAAPPI-time-of-flight (TOF)-MSI without any sample preparation. Leaf mass spectra were also recorded with laser ablation electrospray ionization (LAESI) mass spectrometry and the spectra were compared with those obtained by LAAPPI. RESULTS: Direct probing of the plant tissue by LAAPPI efficiently produced ions from plant metabolites, including neutral and nonpolar terpenes that do not have polar functional groups, as well as oxygenated terpene derivatives. Monoterpenes and monoterpenoids could also be studied from sage by LAESI, but only LAAPPI was able to detect larger nonpolar compounds, such as sesquiterpenes and triterpenoid derivatives, from the leaf matrix. Alternative MSI methods for nonpolar compounds, such as desorption atmospheric pressure photoionization (DAPPI), do not achieve as good spatial resolution as LAAPPI (<400 µm). CONCLUSIONS: We show that MSI with LAAPPI is a useful tool for concurrently studying the distribution of polar and nonpolar compounds, such as phytochemicals, directly from complex biological samples, and it can provide information that is not available by other, established methods.