Origanum majorana L. protects against neuroinflammation-mediated cognitive impairment: a phyto-pharmacological study.

BACKGROUND: Neuroinflammation and oxidative stress are critical players in the pathogenesis of numerous neurodegenerative diseases, such as Alzheimer's disease (AD) which is responsible for most cases of dementia in the elderly. With the lack of curative treatments, natural phenolics are potential candidates to delay the onset and progression of such age-related disorders due to their potent antioxidant and anti-inflammatory effects. This study aims at assessing the phytochemical characteristics of Origanum majorana L. (OM) hydroalcohol extract and its neuroprotective activities in a murine neuroinflammatory model. METHODS: OM phytochemical analysis was done by HPLC/PDA/ESI-MSn. Oxidative stress was induced in vitro by hydrogen peroxide and cell viability was measured using WST-1 assay. Swiss albino mice were injected intraperitoneally with OM extract at a dose of 100 mg/kg for 12 days and with 250 µg/kg LPS daily starting from day 6 to induce neuroinflammation. Cognitive functions were assessed by novel object recognition and Y-maze behavioral tests. Hematoxylin and eosin staining was used to assess the degree of neurodegeneration in the brain. Reactive astrogliosis and inflammation were assessed by immunohistochemistry using GFAP and COX-2 antibodies, respectively. RESULTS: OM is rich in phenolics, with rosmarinic acid and its derivatives being major constituents. OM extract and rosmarinic acid significantly protected microglial cells against oxidative stress-induced cell death (p < 0.001). OM protected against the LPS-induced alteration of recognition and spatial memory in mice (p < 0.001) and (p < 0.05), respectively. Mice that received OM extract prior to the induction of neuroinflammation showed comparable histology to control brains, with no overt neurodegeneration. Furthermore, OM pre-treatment decreased the immunohistochemistry profiler score of GFAP from positive to low positive and COX-2 from low positive to negative in the brain tissue, compared to the LPS group. CONCLUSION: These findings highlight the potential preventive effects of OM phenolics against neuroinflammation and pave the way toward drug discovery and development for neurodegenerative disorders.

Antitumor activity of Cuphea ignea extract against benzo(a)pyrene-induced lung tumorigenesis in Swiss Albino mice.

Lung cancer has one of the highest mortality rates among various types of cancer and is the most frequent cancer in the world. The incidence of lung cancer is increasing rapidly, in parallel with an increased incidence of smoking. Effective chemoprevention may be an alternative strategy to control the incidence of lung cancer. Thus, the objective of current work was to ascertain the possible preventive and therapeutic efficacies of Cuphea ignea extract in a mouse model of lung tumorigenesis and its cytotoxicity toward the A549 human lung cancer cell line. Lung tumorigenesis was induced by the oral administration of benzo(a)pyrene (50 mg/kg b.w.) twice per week to Swiss albino mice for 4 weeks. Benzo(a)pyrene-treated mice were orally administered C. ignea (300 mg/kg body weight, 5 days/week) for 2 weeks before or 9 weeks after the first benzo(a)pyrene dose, for a total of 21 weeks. At the end of the administration period, various parameters were measured in the serum and lung tissues. The results revealed that the oral administration of benzo(a)pyrene resulted in increases in relative lung weight, serum levels of tumor markers (ADA, AHH, and LDH), and the inflammatory marker NF-κB, and a decreased total antioxidant capacity compared with the control. In addition, decreased levels of enzymatic and non-enzymatic antioxidants, with a concomitant increase in lipid peroxidation, metalloproteinases (MMP-2 and MMP-12), and the angiogenic marker VEGF were detected in lung tissues. Moreover, benzo(a)pyrene administration induced the upregulation of PKCα, COX-2, and Bcl-2 expression, with the downregulation of BAX and caspase-3 expression. C. ignea treatment alleviated all alterations in these parameters, which was further confirmed by the histopathological analysis of lung tissues. The findings of the current work provide the first verification of the preventive and therapeutic potentials of C. ignea extract against benzo(a)pyrene-induced lung tumorigenesis in mice.

VUV Photodissociation Induced by a Deuterium Lamp in an Ion Trap.

Tandem mass spectrometry represents an important analytical tool to unravel molecular structures and to study the gas-phase behavior of organic molecules. Besides commonly used methods like collision-induced dissociation and electron capture or transfer dissociation, new ultraviolet light-based techniques have the potential to synergistically add to the activation methods. Here, we present a new simple, yet robust, experimental design for polychromatic activation of trapped ions using the 115-160 nm output of a commercially available deuterium lamp. The resulting continuous dissociative excitation with photons of a wide energy range from 7.7 to 10.8 eV is studied for a comprehensive set of analyte classes in both positive and negative ion modes. While being simple, affordable, compact, and of low maintenance, the new setup initiates fragmentation of most precursor ions via their known dissociation pathways. Additionally, some new fragmentation patterns were discovered. Especially, electron loss and electron capture reactions with subsequent fragmentations were observed. For oligonucleotides, peptides, carbohydrates, and organic dyes, in comparison to collision-induced dissociation, a significantly wider fragment distribution was obtained, resulting in an information increase. Since the individual photons carry enough energy to post-ionize the nascent fragments, a permanent vacuum ultraviolet light exposure inside the ion trap potentially goes along with a general increase in detection capability.

Nucleic acid and SNP detection via template-directed native chemical ligation and inductively coupled plasma mass spectrometry.

Detection of nucleic acids and single nucleotide polymorphisms (SNPs) is of pivotal importance in biology and medicine. Given that the biological effect of SNPs often is enhanced in combination with other SNPs, multiplexed SNP detection is desirable. We show proof of concept of the multiplexed detection of SNPs based on the template-directed native chemical ligation (NCL) of PNA-probes carrying a metal tag allowing detection using ICP-MS. For the detection of ssDNA oligonucleotides (30 bases), two probes, one carrying the metal tag and a second one carrying biotin for purification, are covalently ligated. The methodological limit of detection is of 29 pM with RSD of 6.7% at 50 pM (n = 5). Detection of SNPs is performed with the combination of two sets of reporter probes. The first probe set targets the SNP, and its yield is compared with a second set of probes targeting a neighboring sequence. The assay was used to simultaneously differentiate between alleles of three SNPs at 5-nM concentration.

Neuromodulatory Activity of Dietary Phenolics Derived from Corchorus olitorius L.

Dietary phenolics are known for their potent antioxidant and anti-inflammatory activities, making them promising candidates for protection against neuroinflammation and neurodegeneration. Hydroalcohol extract of Egyptian species of Corchorus olitorius L. (Co) leaves was investigated for its neuroprotective effects in a lipopolysaccharide-induced neuroinflammatory mouse model. Twenty five metabolites were characterized from the bioactive extract using high-performance liquid chromatography HPLC/PDA/HRESI/MSn , revealing 1,5-dicaffeoylquinic acid (Co11) as one of the major constituents (5.7%), which was isolated and its identity was confirmed by spectral data as first report. Co significantly protected microglia against H2 O2 -induced cytotoxicity and immunohistochemistry showed reduced expression of the astrocytic marker, glial fibrillary acidic protein, and the inflammatory marker, cyclooxygenase-2. These findings correlated with significant improvement of cognitive functions and reduction of LPS-induced neurodegeneration in Co-treated mice as revealed by histopathology. The current study shows promising effects of Co in limiting neurodegeneration and cognitive impairment caused by neuroinflammation and glial cell activation. PRACTICAL APPLICATION: Information presented here shed light on the promising effects of Corchorus olitorius (Co) for the modulation of neuroinflammatory pathways improving the neuroinflammation-related neurodegeneration and cognitive decline. This makes Co a promising candidate as a nutraceutical supplement to be used against neuroinflammation-related disorders.

Characterization of hepatoprotective metabolites from Artemisia annua and Cleome droserifolia using HPLC/PDA/ESI/MS-MS

ABSTRACT The hepatoprotective activities of two traditionally used plants, Cleome droserifolia (Forssk.) Delile, Cleomaceae, and Artemisia annua L., Asteraceae, were recently reported. However, the biologically active metabolites responsible for this activity were not identified. The aqueous extract of C. droserifolia aerial parts, and the polar fraction of A. annua leaves were screened for their antioxidant activities using the 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) assay. The in vitro viability of HepG-2 cells treated with CCl4 and the extracts were assessed by MTT assay. The effects of the extracts on the liver enzymes and the total soluble protein in CCl4-intoxicated HepG-2 cells were investigated. An HPLC/PDA/ESI/MS-MS based analysis was carried out for extract of C. droserifolia and polar fraction of A. annua. Both exhibited pronounced free radical scavenging activities (86 and 83%, respectively). Both showed a significant increase in cell viability: 86.43% for the extract of C. droserifolia and 79.32% for polar fraction of A. annua. Only the extract of C. droserifolia (39.6 ± 5.41 and 20.4 ± 6.91 IU/dl, respectively) and polar fraction of A. annua (40.8 ± 2.14 and 24.5 ± 3.11 IU/dl, respectively) restored the levels of liver enzymes (aspartate transaminase and alanine transaminase, respectively) compared to the CCl4 intoxicated group (87.5 ± 4.34 and 34.1 ± 8.12 IU/dl, respectively) and other herbal extracts. More than fifty phenolic secondary metabolites were identified in the extracts under investigation. The significant hepatoprotective activities of both extracts seemed to be strongly connected to their content of hydroxycinnamoyl quinic acids and flavonoids.

Comparison of the fragmentation behavior of DNA and LNA single strands and duplexes.

DNA and locked nucleic acid (LNA) were characterized as single strands, as well as double stranded DNA-DNA duplexes and DNA-LNA hybrids using tandem mass spectrometry with collision-induced dissociation. Additionally, ion mobility spectrometry was carried out on selected species. Oligonucleotide duplexes of different sequences-bearing mismatch positions and abasic sites of complementary DNA 15-mers-were investigated to unravel general trends in their stability in the gas phase. Single-stranded LNA oligonucleotides were also investigated with respect to their gas phase behavior and fragmentation upon collision-induced dissociation. In contrast to the collision-induced dissociation of DNA, almost no base loss was observed for LNAs. Here, backbone cleavages were the dominant dissociation pathways. This finding was further underlined by the need for higher activation energies. Base losses from the LNA strand were also absent in fragmentation experiments of the investigated DNA-LNA hybrid duplexes. While DNA-DNA duplexes dissociated easily into single stranded fragments, the high stability of DNA-LNA hybrids resulted in predominant fragmentation of the DNA part rather than the LNA, while base losses were only observed from the DNA single strand of the hybrid.

Molecules and elements for quantitative bioanalysis: The allure of using electrospray, MALDI, and ICP mass spectrometry side-by-side.

To understand biological processes, not only reliable identification, but quantification of constituents in biological processes play a pivotal role. This is especially true for the proteome: protein quantification must follow protein identification, since sometimes minute changes in abundance tell the real tale. To obtain quantitative data, many sophisticated strategies using electrospray and MALDI mass spectrometry (MS) have been developed in recent years. All of them have advantages and limitations. Several years ago, we started to work on strategies, which are principally capable to overcome some of these limits. The fundamental idea is to use elemental signals as a measure for quantities. We began by replacing the radioactive 32 P with the "cold" natural 31 P to quantify modified nucleotides and phosphorylated peptides and proteins and later used tagging strategies for quantification of proteins more generally. To do this, we introduced Inductively Coupled Plasma Mass Spectrometry (ICP-MS) into the bioanalytical workflows, allowing not only reliable and sensitive detection but also quantification based on isotope dilution absolute measurements using poly-isotopic elements. The detection capability of ICP-MS becomes particularly attractive with heavy metals. The covalently bound proteins tags developed in our group are based on the well-known DOTA chelate complex (1,4,7,10-tetraazacyclododecane-N,N',N″,N‴-tetraacetic acid) carrying ions of lanthanoides as metal core. In this review, I will outline the development of this mutual assistance between molecular and elemental mass spectrometry and discuss the scope and limitations particularly of peptide and protein quantification. The lanthanoide tags provide low detection limits, but offer multiplexing capabilities due to the number of very similar lanthanoides and their isotopes. With isotope dilution comes previously unknown accuracy. Separation techniques such as electrophoresis and HPLC were used and just slightly adapted workflows, already in use for quantification in bioanalysis. Imaging mass spectrometry (MSI) with MALDI and laser ablation ICP-MS complemented the range of application in recent years.

Synthesis and characterization of a new MeCAT reagent containing a photocleavable linker for labeling of proteins and peptides in mass spectrometric analyses.

The quantification of proteins and peptides becomes more important besides mere identification in modern life sciences. Therefore, we have developed a new reagent that adds to the known metall-coded affinity tagging strategy employed in molecular and elemental mass spectrometry containing a photocleavable linker. A synthesis route was developed that provides the new reagent in good yields. The stability of the synthesized reagents was assessed under different temperature and illumination conditions. Labeling reactions were carried out at peptide and protein level, while also the fragmentation behavior of labeled peptides was assessed. In additional experiments, the photocleavability of the new reagent was examined. Upon irradiation with ultraviolet light, the photoproducts were liberated and could be used for quantification of labeled peptides.

Complementarity of molecular and elemental mass spectrometric imaging of Gadovist™ in mouse tissues.

Drug biodistribution analyses can be considered a key issue in pharmaceutical discovery and development. Here, mass spectrometric imaging can be employed as a powerful tool to investigate distributions of drug compounds in biologically and medically relevant tissue sections. Both matrix-assisted laser desorption ionization-mass spectrometric imaging as molecular method and laser ablation inductively coupled plasma-mass spectrometric imaging as elemental detection method were applied to determine drug distributions in tissue thin sections. Several mouse organs including the heart, kidney, liver, and brain were analyzed with regard to distribution of Gadovist™, a gadolinium-based contrast agent already approved for clinical investigation. This work demonstrated the successful detection and localization of Gadovist™ in several organs. Furthermore, the results gave evidence that gadolinium-based contrast agents in general can be well analyzed by mass spectrometric imaging methods. In conclusion, the combined application of molecular and elemental mass spectrometry could complement each other and thus confirm analytical results or provide additional information.

Lipid imaging for visualizing cilastatin amelioration of cisplatin-induced nephrotoxicity.

Nephrotoxicity is a major limitation to cisplatin antitumor therapies. Cilastatin, an inhibitor of renal dehydropeptidase-I, was recently proposed as a promising nephroprotector against cisplatin toxicity, preventing apoptotic cell death. In this work, cilastatin nephroprotection was further investigated in a rat model, with a focus on its effect on 76 renal lipids altered by cisplatin, including 13 new cisplatin-altered mitochondrial cardiolipin species. Lipid imaging was performed with MALDI mass spectrometry imaging (MALDI-MSI) in kidney sections from treated rats. Cilastatin was proved to significantly diminish the lipid distribution alterations caused by cisplatin, lipid levels being almost completely recovered to those of control samples. The extent of recovery of cisplatin-altered lipids by cilastatin turned out to be relevant for discriminating direct or secondary lipid alterations driven by cisplatin. Lipid peroxidation induced by cisplatin was also shown to be reduced when cilastatin was administered. Importantly, significant groups separation was achieved during multivariate analysis of cortex and outer-medullary lipids, indicating that damaged kidney can be discerned from the nephroprotected and healthy groups and classified according to lipid distribution. Therefore, we propose MALDI-MSI as a powerful potential tool offering multimolecule detection possibilities to visualize and evaluate nephrotoxicity and nephroprotection based on lipid analysis.

Polyphenols from Tamarix nilotica: LC⁻ESI-MSn Profiling and In Vivo Antifibrotic Activity.

Tamarix nilotica (Ehrenb.) Bunge (Tamaricaceae), an indigenous plant to the Middle East region, is well-known as a medicinal plant for treating many human ailments. The current study aimed at exploring the polyphenol profile of the alcohol soluble fraction of aqueous T. nilotica extract, assessing its in vivo antifibrotic activity and the possible underlying mechanism, to unravel the impact of quantitative difference of sulphated polyphenols content on the antifibrotic activity of T. nilotca grown in two different habitats. Polyphenol profiling of T. nilotica extracts was performed using HPLC-HRESI-QTOF-MS-MS. The major polyphenol components included sulphated flavonoids, phenolic acids and free aglycones. The antifibrotic activity was evaluated through carbon tetrachloride-induced liver fibrosis in rats. Biochemical evaluations revealed that both fractions ameliorated the increased levels of hepatic aminotransferases, lipid peroxidation, hydroxyproline, α-smooth muscle actin (α-SMA), tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2) and nuclear factor kappa B (NF-κB). Moreover, both fractions reduced catalase activity (CAT) and enhanced hepatic glutathione (GSH) content. Histopathological imaging undoubtedly confirmed such results. In conclusion, the T. nilotica polyphenol-rich fraction exhibited potential antifibrotic activity in rats. Significant alterations in GSH levels were recorded based on the sulphated polyphenol metabolite content.

Comparative pharmacokinetics of trandolapril, its active metabolite, and verapamil in human plasma of Egyptian population using HPLC-MS/MS.

Trandolapril and verapamil are commonly used antihypertensive drugs. However, there is a lack of available data on the change of their pharmacokinetics in patients with liver or kidney impairment and hence the need for dose adjustment. In this article, a high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and validated for the monitoring of trandolapril, its active metabolite trandolaprilat, and verapamil in human plasma of patients with renal impairment and/or liver insufficiency. The chromatographic separation was achieved on a Gemini C18 reversed phase column using a gradient elution mode with a run time of 10 minutes. The mobile phase consisted of a mixture of methanol and 2% acetic acid. The electrospray ionization MS/MS analysis was performed in multiple reaction monitoring (MRM) mode. The assay was validated as per Food and Drug Administration (FDA) guidelines for bioanalytical method validation and proved to be suitable for the determination of therapeutic drug levels in plasma. The inter-group changes in pharmacokinetic data were compared to that of healthy volunteers. The comparison showed a significant difference in the pharmacokinetic parameters between the studied groups. The presented results exhibit the benefits of the proposed assay as a validated analytical tool for the continuous drug monitoring.

Negative nucleotide ions as sensitive probes for energy specificity in collision-induced fragmentation in mass spectrometry.

RATIONALE: The most commonly used fragmentation methods in tandem mass spectrometry (MS/MS) are collision-induced dissociation (CID) and higher energy collisional dissociation (HCD). While in CID the preselected ions in the trap are resonantly (and m/z exclusively) excited, in HCD the entire m/z range experiences the dissociative acceleration. The different excitation is reflected in different fragment distributions. METHODS: As a test-bed for particularly pronounced fragmentation specificity, here MS/MS experiments on several 4-mer oligonucleotides were conducted employing both collision methods and the results were thoroughly compared. Oligonucleotides are shown to be sensitive probes to subtle changes, especially in the negative ion mode. A detailed analysis of these differences reveals insight into the dissociation mechanics. RESULTS: The differences are represented in heat-maps, which allow for a direct visual inspection of large amounts of data. In these false colour representations the, sometimes subtle, changes in the individual dissociation product distributions become distinct. Another advantage of these graphic plots can be found in the formation of systematic patterns. These patterns reflect trends in dissociation specificity which allow for the formulation of general rules in fragmentation behavior. CONCLUSIONS: Instruments equipped with two different excitation schemes for MS/MS are today widely available. Nonetheless, direct comparisons between the individual results are scarcely made. Such comparative studies bear a powerful analytical potential to elucidate fragmentation reaction mechanism.

A new strategy for metal labeling of glycan structures in antibodies.

Quantitative analysis of complex proteins is a challenging task in modern bioanalytical chemistry. Commonly available isotope labels are still suffering from limitations and drawbacks, whereas new metal labels open numerous possibilities in mass spectrometric analyses. In this work, we have developed a new metal labeling strategy to tag glycan structures of proteins, more particularly antibodies. The oligosaccharide glycans were selectively trimmed to the last N-acetylglucosamine to which an artificial azide containing galactose residue was bound. This azide can be used for subsequent cycloaddition of an alkyne. Therefore, we developed a lanthanide-containing macrocyclic reagent to selectively connect to this azido galactose. In summary, the glycan structures of an antibody can be labeled with a metal functionality using this approach. Furthermore, the functionality of the antibodies can be fully maintained by labeling the Fc glycans instead of using labeling reagents that target amino or thiol groups. This approach enables the possibility of using elemental, besides molecular mass spectrometry, for quantitative analyses or imaging experiments of antibodies in complex biological samples. Graphical abstract Antibody labeling at sugar moieties with rare earth elements to enable application in elemental mass spectrometry.

Charge-induced geometrical reorganization of DNA oligonucleotides studied by tandem mass spectrometry and ion mobility.

Mass spectrometry is applied as a tool for the elucidation of molecular structures. This premises that gas-phase structures reflect the original geometry of the analytes, while it requires a thorough understanding and investigation of the forces controlling and affecting the gas-phase structures. However, only little is known about conformational changes of oligonucleotides in the gas phase. In this study, a series of multiply charged DNA oligonucleotides (n = 15-40) has been subjected to a comprehensive tandem mass spectrometric study to unravel transitions between different ionic gas-phase structures. The nucleobase sequence and the chain length were varied to gain insights into their influence on the geometrical oligonucleotide organization. Altogether, 23 oligonucleotides were analyzed using collision-induced fragmentation. All sequences showed comparable correlation regarding the characteristic collision energy. This value that is also a measure for stability, strongly correlates with the net charge density of the precursor ions. With decreasing charge of the oligonucleotides, an increase in the fragmentation energy was observed. At a distinct charge density, a deviation from linearity was observed for all studied species, indicating a structural reorganization. To corroborate the proposed geometrical change, collisional cross-sections of the oligonucleotides at different charge states were determined using ion mobility-mass spectrometry. The results clearly indicate that an increase in charge density and thus Coulomb repulsion results in the transition from a folded, compact form to elongated structures of the precursor ions. Our data show this structural transition to depend mainly on the charge density, whereas sequence and size do not have an influence.

Dual Internal Standards with Metals and Molecules for MALDI Imaging of Kidney Lipids.

The quest for internal standards useful in MALDI imaging studies goes on to get not only lateral distribution but also reliable relative quantitative information. We developed a method based on application of matrix and dual internal standards to allow intra- and intersample normalization of lipids intensities in kidney sections of control and cisplatin-treated Wistar rats. An inkjet printer was used to deposit a custom-prepared ink with DHB as MALDI matrix, a primary lipids-based internal standard, and a spiked lanthanide as a secondary internal standard. We applied different laser energy and varied the amounts of matrix-internal standards mixture to evaluate the normalization potential of the internal standards. Successful correction of intensity artifacts caused by instrumental drifts was possible, but not those resulting from uneven matrix application. ICP-MS absolute quantification of the lanthanide in the printed layer ensured the reproducibility of the matrix and internal standards application with RSD of 10-15%. Internal standard-normalized data allowed intrasample modification of the MALDI image to make it compatible with the optical image. Normalization to internal standards corrected a 2-fold difference in lipids intensity, which allowed a meaningful comparison of tissue lipids in control and cisplatin-treated kidneys. More importantly, normalization of lipid relative abundances based on the same adduct type (H+, Na+, and K+) for analyte and internal standard corrected for different ionization efficiencies showing a realistic signal level and enabling reliable comparison of different samples on relative quantitative basis.

Comprehensive Molecular Characterization of a Cisplatin-Specific Monoclonal Antibody.

Despite their immense and rapidly increasing importance as analytical tools or therapeutic drugs, the detailed structural features of particular monoclonal antibodies are widely unknown. Here, an antibody already in use for diagnostic purposes and for molecular dosimetry studies in cancer therapy with very high affinity and specificity for cisplatin-induced DNA modifications was studied extensively. The molecular structure and modifications as well as the antigen specificity were investigated mainly by mass spectrometry. Using nano electrospray ionization mass spectrometry, it was possible to characterize the antibody in its native state. Tandem-MS experiments not only revealed specific fragments but also gave information on the molecular structure. The detailed primary structure was further elucidated by proteolytic treatment with a selection of enzymes and high resolution tandem-MS. The data were validated by comparison with known antibody sequences. Then, the complex glycan structures bound to the antibody were characterized in all detail. The Fc-bound oligosaccharides were released enzymatically and studied by matrix-assisted laser desorption/ionization mass spectrometry. Overall 16 different major glycan structures were identified. The binding specificity of the antibody was investigated by applying synthetic single and double stranded DNA oligomers harboring distinct Pt adducts. The antibody-antigen complexes were analyzed by mass spectrometry under native conditions. The stability of the complex with double stranded DNA was also investigated.

MALDI-LTQ-Orbitrap mass spectrometry imaging for lipidomic analysis in kidney under cisplatin chemotherapy.

Imaging techniques for mapping molecular distributions in tissue sections can reveal valuable information on biomolecules involved in relevant biochemical processes. A method has been developed for comprehensive, reproducible and sensitive lipid imaging by matrix-assisted laser/desorption ionization-LTQ-Orbitrap mass spectrometry in kidney sections, showing the benefits of exact mass determination. Matrix deposition parameters for positive and negative lipid ion imaging using different matrices such as 2,5-dihydroxybenzoic acid (DHB), 9-aminoacridine (9-AA) or α-cyano-4-hydroxycinnamic acid (CHCA) have been optimized for the broadest detection and identification of renal lipids. The combination of 9-AA and DHB was found as the most suitable for negative and positive ion mode lipid imaging, respectively. Lipid mapping and related identification strategies and limitations have also been discussed. Production of 100-µm resolution images was proved to be enough for discerning lipid distribution in kidney substructures. Imaging reproducibility was assessed on parallel kidney slices with time. This method has been applied to the lipidomics analysis on kidney sections from rats treated with the antitumor drug cisplatin and compared to healthy control rats. Up to 66 different renal lipids out of 450 extracted ion images (mainly phospholipid species, in addition to sulfatides and cholesterol sulfate) have been found and identified showing a modified distribution pattern due to cisplatin-induced nephrotoxicity. These lipid species reflect either topographic, signaling or structural processes in damaged kidney and could potentially be used for nephrotoxicity assessment or as therapeutic targets. This is, to our knowledge, the first imaging lipidomics study for nephrotoxicity assessment of cisplatin chemotherapy.