High-performance Liquid chromatography-MS Research Articles

Single-cell mass spectrometry studies of drug metabolism heterogeneity and primary resistance to gefitinib in non-small cell lung cancer cells

Patients with epidermal growth factor receptor (EGFR) wild-type non-small cell lung cancer (NSCLC) often show primary resistance to gefitinib therapy. It is thus necessary to study the metabolism of gefitinib in NSCLC cells to comprehensively reveal the reasons for the primary resistance of tumors. Herein, we develop a platform for studying drug metabolism heterogeneity based on single-cell mass spectrometry (sDMH-scMS) by integrating living-cell electrolaunching ionization MS (ILCEI-MS) and high-performance liquid chromatography-MS (HPLC-MS) analysis, and the primary resistance of NSCLC cells to gefitinib was studied using this platform. The ILCEI-MS analysis showed that approximately 11.9% of NSCLC single cells contained the gefitinib metabolite M11; HPLC-MS detection diluted the intensity of M11 in subpopulations and concealed the heterogeneity of drug metabolism in tumor single cells. The intensity of gefitinib in EGFR wild-type A549 cells was markedly lower than in mutant PC9 cells, and the intensity of gefitinib metabolites was significantly higher than in PC9 cells, suggesting that the primary resistance of NSCLC cells is related to gefitinib metabolism. Moreover, the combination of gefitinib and the drug-metabolizing enzyme inhibitor α-naphthoflavone was shown to overcome the primary resistance of the NSCLC cells. Overall, the results of this study are expected to be applicable for clinical drug resistance diagnosis and treatment at the single-cell level.

A study of the oxidation mechanism of the organic pigment indigo in grottoes murals by ozone under dark conditions

In this paper, the organic pigments indigo and isatin were detected by high performance liquid chromatography (HPLC) in The Mural of Four Buddhas which is in Cave 3 from the ancient Chinese Tiantishan grottoes (Ming Dynasty, East Slope). By analysing the preservation conditions of the mural and the environmental conditions of the place where the Tiantishan grottoes are located, we speculated that the isatin detected in this mural was mainly produced by the oxidative decomposition of indigo by ozone (O3), rather than by photodegradation of indigo. We have used theoretical calculation software Gaussian09 (G09) and Amsterdam Density Functional (ADF) module in the Amsterdam Modeling Suite (AMS) software to simulate the reaction mechanism of O3 oxidation of indigo, and the end products of the oxidation of the natural plant dye indigo by O3 were identified as isatin, C16H10N2O3 and C8H6NO3 using HPLC, fluorescence spectroscopy and high performance liquid chromatography-mass spectrometry(HPLC–MS). This finding confirmed the accuracy of the mechanism of indigo fading by O3 oxidation. These findings provided a theoretical basis for subsequent research into the derivation of natural organic dyes in the face of increasing O3 pollution and for better protection of valuable historical and cultural heritage such as ancient Chinese grottoes murals.

New Approach to the Detection of Short-Lived Radical Intermediates.

We report a new general method for trapping short-lived radicals, based on a homolytic substitution reaction SH2′. This departure from conventional radical trapping by addition or radical–radical cross-coupling results in high sensitivity, detailed structural information, and general applicability of the new approach. The radical traps in this method are terminal alkenes possessing a nitroxide leaving group (e.g., allyl-TEMPO derivatives). The trapping process thus yields stable products which can be stored and subsequently analyzed by mass spectrometry (MS) supported by well-established techniques such as isotope exchange, tandem MS, and high-performance liquid chromatography-MS. The new method was applied to a range of model radical reactions in both liquid and gas phases including a photoredox-catalyzed thiol–ene reaction and alkene ozonolysis. An unprecedented range of radical intermediates was observed in complex reaction mixtures, offering new mechanistic insights. Gas-phase radicals can be detected at concentrations relevant to atmospheric chemistry.

Natural aspirin of some members of the Rosaceae and Salicaceae families

White willow Salix alba L., a member of the family Salicaceae is characterized by the presence of phenolic compounds and their glycosides (1.5–11.0 %), such as salicin, salicortin, salireposide, picein, triandrin, syringin, tremulacin, flavonoids (1–4 %) namely luteolin, apigenin, quercetin, quercimeritrin, cosmosin, diosmetin, tannins (8–20 %) i.e. catechins, anthocyanidins, halotanines of organic acids, ascorbic acid. Preparations of white willow bark are used as an antipyretic, anti-inflammatory, diaphoretic, analgesic, diuretic, and astringent. The rhizomes with roots of the commonly known dropwort or fern-leaf dropwort are Filipendula vulgaris Moench., member of the family Rosaceae are characterized by phenologicoside gaulterin, the hydrolysis of which produces salicylic aldehyde, tannins (up to 33 %), flavonoids, ascorbic acid, polysaccharides, starch. The herbs and flowers are used as anti-inflammatory, anti-rheumatic, vasodilator, gastric, and antiulcer agents. The aim of the work is to determine the composition of the white willow bark Salix alba L. tincture, a member of the willow family Salicaceae, using gas-liquid chromatography (GLC) and HPLC and to determine the composition of the tincture of the dropwort root Filipendula vulgaris Moench. family Rosaceae. Materials and methods. Raw materials (underground organs) of the dropwort Filipendula vulgaris Moench. and bark of the white willow Salix alba L., which were harvested in July 2020 at the ZSMU research site. Tinctures were prepared according to the production recipe (1:5) (extractant was 70 % ethanol), investigated by the gas chromatograph Agilent 7890B with mass spectrometric detector 5977B. The mass spectra library NIST 14 was used to identify the components. White willow bark tincture (70 % ethyl alcohol extractant) was used to determine the phenolic compounds content and salicylic acid derivatives. The study was performed by HPLC (high-performance liquid chromatography) and HPLC MS. Results. Qualitative and quantitative analysis of the components of Salix alba L. willow bark tincture was performed by gas-liquid chromatography, 39 components of different classes of biologically active compounds have been identified, among which the 10 components are predominated in quantitative content. The main one was Salicin (39.53 %). The GC analysis of the tincture of the dropwort roots, Filipendula vulgaris Moench., identified 34 components of different groups of biologically active compounds, dominated by 11 components, the main is Ethyl.alpha.-d-glucopyranoside (34.55 %). HPLC analysis confirmed the presence of phenolic compounds (flavonoids such as apigenin, and quercetin) and salicylic acid derivatives (glycoside salicin) in the tincture of white willow bark (Salix alba L.). Conclusions. By GC analysis the 39 components in tincture of white willow bark Salix alba L. were identified, 10 components were predominated, the main one was Salicin (39.53 %). The 34 components in the tincture of the dropwort roots, Filipendula vulgaris Moench., were identified, dominated by 11 components, the main was Ethyl.alpha.-d-glucopyranoside (34.55 %). The presence of phenolic compounds (flavonoids were apigenin and quercetin) and salicylic acid derivatives (glycoside salicin) was confirmed by HPLC MS in the tincture of white willow bark (Salix alba L.).

Discovery and Identification of Arsenolipids Using a Precursor-Finder Strategy and Data-Independent Mass Spectrometry.

Arsenolipids are a class of lipid-soluble arsenic species. They are present in seafoods and show high potentials of cytotoxicity and neurotoxicity. Hindered by traditional low-throughput analytical techniques, the characterization of arsenolipids is far from complete. Here, we report on a sensitive and high-throughput screening method for arsenolipids in krill oil, tuna fillets, hairtail heads, and kelp. We demonstrate the detection and identification of 23 arsenolipids, including novel arsenic-containing fatty acids (AsFAs), hydroxylated AsFAs, arsenic-containing hydrocarbons (AsHCs), hydroxylated AsHCs, thiolated trimethylarsinic acids, and arsenic-containing lysophosphatidylcholines not previously reported. The new method incorporated precursor ion scan (PIS) into data-independent acquisition. High-performance liquid chromatography (HPLC) electrospray ionization quadrupole time-of-flight mass spectrometry (ESI–qToF–MS) was used to perform the sequential window acquisition of all theoretical spectra (SWATH). Comprehensive HPLC–MS and MS/MS data were further processed using a fragment-guided chromatographic computational program Precursorfinder developed here. Precursorfinder achieved efficient peak-picking, retention time comparison, hierarchical clustering, and wavelet coherence calculations to assemble fragment features with their target precursors. The identification of arsenolipids was supported by coeluting the HPLC–MS peaks detected with the characteristic fragments of arsenolipids. Method validation using available arsenic standards and the successful identification of previously unknown arsenolipids in seafood samples demonstrated the applicability of the method for environmental research.

Electrostatic Spray Ionization-Mass Spectrometry for Direct and Fast Wine Characterization.

Due to the globally existed and economically motivated adulteration including mislabeling and/or blending, fast wine characterization is important in wine industry. Herein, we developed an electrostatic spray ionization-mass spectrometry (ESTASI-MS)-based method to distinguish wines. Wine samples were directly analyzed by ESTASI-MS without any pretreatment. Microdroplets of wine were deposited on a plastic plate for analysis. The collection of each mass spectrometric datum can be accomplished in 1–2 min without any need of pretreatment to the sample, followed by principle component analysis to discriminate wines with different labels and vintages. Long-term storage of wine was simulated and characterized by utilizing the method. High-performance liquid chromatography-MS was further applied to identify the distinctive compounds in wines to indicate their difference. We found that the method can offer a strategy for quick wine analysis, which is of practical value in wine industry for wine classification and aging control.

Determination of vortioxetine in human serum and saliva samples by HPLC–DAD and HPLC–MS

Vortioxetine is a new drug against major depressive disorder with high affinity for a range of different serotonergic targets in the central nervous system. Therapeutic drug monitoring is an important tool for the clinical management of patients receiving a pharmacotherapy, particularly in psychiatry. For this reason, determination of drug concentration in biological fluids is important for a rational dosage of drugs. Rapid and reliable analytical assays are also required to detect and identify drugs of toxicological importance. For analysis of vortioxetine by high-performance liquid chromatography (HPLC), no procedures for its determination in saliva have been reported and there are only a few ones for its determination in serum. A sensitive and selective highperformance liquid chromatography with diode array detector (HPLC-DAD) or mass spectrometer (HPLC-MS) method was developed for the fast quantification of vortioxetine in human saliva and serum. The determination was performed on a Synergi Polar RP column in isocratic mode under the optimal mobile phase containing 70% methanol, 20% acetate buffer at pH 3.5, 10% double distilled water, and 0.025 M L−1 diethylamine.

Inhibitory Effect of Citalopram on the Pharmacokinetics of Carvedilol in Rats and in vitro Models

Background/Aims: The aim of this study was to investigate the drug-drug interaction between carvedilol and citalopram based on carvedilol metabolism in vitro and his pharmacokinetics (PKs) in vivo after the oral administration of the single drug and both drugs, and reveal citalopram effects on the PKs of carvedilol. Methods: Each rat was cannulated on the femoral vein, prior to being connected to BASi Culex ABC®. Carvedilol was orally administrated in rats (3.57 mg/kg body weight [b.w.]) in the absence of citalopram or after a pre-treatment with multiple oral doses of citalopram (1.42 mg/kg b.w.). Plasma concentrations of carvedilol were determined using high-performance liquid chromatography-MS at the designated time points after drug administration, and the main PK parameters were calculated by noncompartmental analysis. In addition, effects of citalopram on the metabolic rate of carvedilol were investigated using rat-pooled liver microsome incubation systems. Results: During co-administration, significant increases of the area under the plasma concentration-time curve as well as of the peak plasma concentration were observed. The rat-pooled liver microsome incubation experiment indicated that citalopram could decrease the metabolic rate of carvedilol. Conclusion: Citalopram co-administration led to a significant alteration of carvedilol's PK profile in rats; it also demonstrated, in vitro, these effects could be explained by the existence of a drug-drug interaction mediated by CYP2D6 inhibition.

Identification of Impurities in 5-Aminolevulinic Acid by Two-Dimensional Column-Switching Liquid Chromatography Coupled with Linear Ion Trap Mass Spectrometry

Identification of impurities in 5-aminolevulinic acid (ALA) by mass spectrometry is difficult, because MS-incompatible mobile phases, such as phosphate buffers or ion-pair reagents, need to be used to separate the major component from impurities. In this study, the unknown impurities in ALA have been identified by two-dimensional (2D) column-switching high-performance liquid chromatography (HPLC) coupled with linear ion trap mass spectrometry (LIT MS). The first-dimensional analytical column was a Gemini C18 (150 mm × 4.6 mm, 5 μm) with a non-volatile salt mobile phase at a flow rate of 1.0 mL min−1, and the second-dimensional analytical column was a ZORBAX SB C8 (150 mm × 4.6 mm, 3.5 μm) with a volatile salt mobile phase at a flow rate of 1.0 mL min−1. The detection wavelength was 205 nm. Mass spectra were acquired with an ESI source, in both positive and negative ion modes. Six impurities were identified by their MS2 and MS3 fragments, and the mass fragmentation patterns and structural assignments of these impurities were studied. The results obtained by the two-dimensional column-switching method were further compared with those of the conventional one-dimensional normal-phase HPLC–MS using an amide column and an MS-compatible mobile phase for separation. The two-dimensional column-switching method described herein proved to be advantageous in terms of the number of impurities identified. The column-switching and online demineralization technique made the mobile phase conditions compatible with mass spectrometry. Thus, the method solves the problem of incompatibility between non-volatile salt mobile phases and mass spectrometry, making it worthy of popularization and application in impurity identification.

Anti-helminthic activity of Momordica charantia L. against Fasciola hepatica eggs after twelve days of incubation in vitro

Fasciolosis, a parasitic disease caused by the trematode Fasciola hepatica underreported is expanding both in human and animal population, throughout the world. The constant use of synthetic drugs to treat this condition has led to the natural selection of resistant strains of the parasite. Hence, there is a growing focus on the potential anti-helminthic properties of medicinal plants and phytopharmaceuticals. The current study assessed the potential anti-fasciolicide action of Momordica charantia leaf extracts and fractions on the eggs of F. hepatica parasites. The lyophilized crude extract (CE) of M. charantia leaves and its sub-fractions, obtained from liquid–liquid partitioning with organic solvents, were analysed by High Performance Liquid Chromatography (HPLC), suspended in 1% DMSO and used in in vitro tests. Quadruplicates of 50F. hepatica eggs were incubated at 23°C with M. charantia leaf CE in different concentrations. After 12days no larvae were formed in eggs incubated with CE concentrations above 12.5mg/mL. Eggs incubated with CE sub-fractions at concentrations of 1000, 100, 10, 1, 0.1, 0.01μg/mL affected embryonic development, with n-butanol presenting the strongest inhibition of miracidia formation. In contrast, on the 12th day, 90% of the miracidia hatched in the control experiments using 0.03% DMSO whereas embryogenesis was completely abolished with any concentration of albendazole sulphoxide ABZ(SO). Chemical analysis of the CE and sub-fractions revealed a prominent presence of flavonoids. HPLC–MS confirmed Quercetin to be one of the main flavonoids present in the CE and the n-butanol subfraction. This is the first study to analyse the potential anti-fasciolicide action of M. charantia leaf CE and subfractions.

A new approach to live reaction monitoring using active flow technology in ultra-high-speed HPLC with mass spectral detection.

A new type of chromatography column referred to as a parallel segmented flow (PSF) column enables ultra-high-speed high-performance liquid chromatography-MS to be undertaken. This occurs because the separation efficiency obtained on PSF columns has been shown in prior studies to be superior to conventional columns, and the flow stream is split radially inside the outlet end fitting of the column, rather than in an axial post-column flow stream split. As a result, the flow through the column can be five times higher than the flow through the MS. In this work, the degradation of amino acids in dilute nitric acid was used to illustrate the process. Separations were obtained in less than 12 s, although the reinjection process was initiated 6 s after the previous injection. The degradation rate constant of tryptophan, in the presence of tyrosine and phenylalanine, was determined.

Simple correction improving long-term reproducibility of HPLC-MS.

Chromatographic peak areas in long series of high-performance liquid chromatography-MS experiments often vary, which decrease reproducibility and may cause bias in the results. It was found that the sensitivity of various components change differently; in our case, variability is in the order of 20-40%, and it is most likely due to changing conditions in electrospray ionization (ESI). The most often used peak area correction methods do not take this effect into account. The change in peak areas can be well described by a polynomial function; we found that a fourth-order polynomial is most often suitable. We suggest a simple correction algorithm based on polynomial fitting. When the experiments were inherently well reproducible, this correction improved reproducibility from 12% to 3% (on average for various components). When random errors were larger, this improvement was less significant (15% to 12% in nano-ESI) but nevertheless essential in order to avoid possible bias in the results.

High-Throughput and Comprehensive Lipidomic Analysis Using Ultrahigh-Performance Supercritical Fluid Chromatography-Mass Spectrometry.

New analytical approach for high-throughput and comprehensive lipidomic analysis of biological samples using ultrahigh-performance supercritical fluid chromatography (UHPSFC) with electrospray ionization-mass spectrometry (ESI-MS) is presented in this work as an alternative approach to established shotgun MS or high-performance liquid chromatography-MS. The lipid class separation is performed by UHPSFC method based on 1.7 μm particle-bridged ethylene hybrid silica column with a gradient of methanol-water-ammonium acetate mixture as a modifier. All parameters of UHPSFC conditions are carefully optimized and their influence on the chromatographic behavior of lipids is discussed. The final UHPSFC/ESI-MS method enables a fast separation of 30 nonpolar and polar lipid classes within 6 min analysis covering 6 main lipid categories including fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, sterols, and prenols. Individual lipid species within lipid classes are identified based on positive and negative-ion full-scan and tandem mass spectra measured with high mass accuracy and high resolving power. Developed UHPSFC/ESI-MS method is applied for the analysis of porcine brain extract as a complex lipidomic sample, where 24 lipid classes containing 436 lipid species are identified. The method is validated for the quantitative analysis of lipid species in biological tissues using internal standards for each lipid class. This high-throughput, comprehensive and accurate UHPSFC/ESI-MS method is suitable for the lipidomic analysis of large sample sets in the clinical research.

Quantitative Profiling of Volatile and Phenolic Substances in the Wine Vernaccia di Serrapetrona by Development of an HS-SPME-GC-FID/MS Method and HPLC-MS

A headspace solid-phase microextraction coupled to gas chromatography with mass spectrometry and flame ionization detection (HS-SPME-GC-MS/FID) method has been developed for quantifying the main volatiles of the Italian sparkling red wine Vernaccia di Serrapetrona, leading to establish that the divinylbenzene/carboxen/polydimethylsiloxane-coated (gray) fiber, an equilibration time of 15 min, and an extraction time of 15 min at a temperature of 35 °C are the conditions representing the best compromise in terms of sensitivity and time expense, to carry out the analyses. Among the volatiles quantified, 2-phenylethanol, 3-methylbutyl acetate, and ethyl esters levels resulted to be above their odor thresholds, thus being probably responsible for the aroma of this wine with their positive attributes. Phenolic compounds, namely gallic acid, p-coumaric acid, caffeic acid, (+)-catechin, and (+)-epicatechin, were also quantified using high-performance liquid chromatography-MS, thus obtaining an overall characterization of molecules fundamental for both the sensory and the healthy characteristics of the wine. The total phenolic content was found to be in the range of 19.25–61.67 mg l−1 with the most abundant compound being gallic acid (7.44–25.78 mg l−1). Main differences in volatile and phenolic compounds between samples were discussed and analyzed by chemometric techniques as principal component analysis.

The Pseudomonas type III effector HopQ1 activates cytokinin signaling and interferes with plant innate immunity

We characterized the molecular function of the Pseudomonas syringae pv. tomato DC3000 (Pto) effector HopQ1. In silico studies suggest that HopQ1 might possess nucleoside hydrolase activity based on the presence of a characteristic aspartate motif. Transgenic Arabidopsis lines expressing HopQ1 or HopQ1 aspartate mutant variants were characterized with respect to flagellin triggered immunity, phenotype and changes in phytohormone content by high-performance liquid chromatography-MS (HPLC-MS). We found that HopQ1, but not its aspartate mutants, suppressed all tested immunity marker assays. Suppression of immunity was the result of a lack of the flagellin receptor FLS2, whose gene expression was abolished by HopQ1 in a promoter-dependent manner. Furthermore, HopQ1 induced cytokinin signaling in Arabidopsis and the elevation in cytokinin signaling appears to be responsible for the attenuation of FLS2 expression. We conclude that HopQ1 can activate cytokinin signaling and that moderate activation of cytokinin signaling leads to suppression of FLS2 accumulation and thus defense signaling.

In vitro bioavailability of total selenium and selenium species from seafood

In vitro bioavailability of total selenium and selenium species from different raw seafood has been assessed by using a simulated gastric and intestinal digestion/dialysis method. Inductively coupled plasma-mass spectrometry (ICP-MS) was used to assess total selenium contents after a microwave assisted acid digestion, and also to quantify total selenium in the dialyzable and non-dialyzable fractions. Selenium speciation in the dialyzates was assessed by high performance liquid chromatography (HPLC) coupled with ICP-MS detection. Major Se species (selenium methionine and oxidized selenium methionine) from dialyzate were identified and characterized by HPLC coupled to mass spectrometry (HPLC–MS). Selenocystine was detected at low concentrations while Se-(Methyl)selenocysteine and inorganic selenium species (selenite and selenate) were not detected in the dialyzate. Low bioavailability percentages for total selenium (6.69±3.39 and 5.45±2.44% for fish and mollusk samples, respectively) were obtained. Similar bioavailability percentages was achieved for total selenium as a sum of selenium species (selenocystine plus oxidized selenium methionine and selenium methionine, mainly). HPLC–MS data confirmed SeMet oxidation during the in vitro procedure.

Chiral separation of bavachinin in Fructus Psoraleae and rat plasma by liquid chromatography using permethylated-β-CD as a chiral selector

A simple, sensitive and selective method of high-performance liquid chromatography (HPLC) has been successfully developed for separation of bavachinin enantiomers in Fructus Psoraleae and rat plasma. The separation and detection conditions of HPLC were optimized. Chiral bavachinin were separated with the mobile phase of methanol and water (70:30, v/v) at a flow rate of 1.0mL/min. The linear ranges were in the range of 20–1000μg/mL. The detection limits were tested as 4ng/mL and 6ng/mL for (+)-bavachinin and (−)-bavachinin, respectively. The method has been applied to analyze chiral bavachinin in rat plasma. HPLC–MS method was used to test the accuracy.

Different Analytical Methods of Para-Phenylenediamine Based Hair Dye

Many cases of toxicity and mortality either due to accidental or deliberate ingestion of hair dyes were reported in Egypt, Sudan, Israel, Morocco, Saudi Arabia, India and Tunisia. In the field of forensic analytical toxicology, the identification and quantification of chemicals are essential in deaths related to intoxication. The aim of the present study is to conduct chemico-analytical characterization of para-phenylenediamine (PPD) based hair dye of (Black stone hair dye, SHD) currently used for dying hear in Egypt, to confirm the identification of its components, that may be potentially toxic on oral or dermal exposure, this study uses comparison between different analytical methods. The applied analytical methods were inductively coupled plasma/MS (ICP/MS), witch used to determine the metal components of the dye, high performance liquid chromatography/MS (HPLC/MS) and gas chromatography/mass spectroscopy (GC/MS) were used for qualitative and quantitative analysis of PPD in the dye. A derivatization method was applied by transforming PPD (aromatic amines) into corresponding imine derivatives. This compound was prepared by condensation of 4-methylbenzeneamine with benzaldehyed to give N-benzylidene-4-methylbenzene-amine, which is a very stable internal standard. ICP/MS analysis revealed that most metal components of hair dye are Be, Na, Al, K, Ca, Sc, Ti, V, Cr, Mn, Fe, Cu, and Zr, where HPLC/MS analysis revealed that PPD is the main ingredient of SHD. Derivatization is necessary for accurate determination of PPD in hair dye by GC/MS method and this results could not be obtained by direct injection of dye (without derivatization).

Analysis of the Nitrogen Content of Distillate Cut Gas Oils and Treated Heavy Gas Oils Using Normal Phase HPLC, Fraction Collection and Petroleomic FT-ICR MS Data

The determination of the nitrogen content of petroleum products is important because nitrogen compounds decrease product quality and can be difficult to remove through processes such as hydrotreating. In recent years, Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) has been used to study the nitrogen species present in petroleum samples (petroleomics), with the goal of identifying hydrotreatment resistant species. While open column separations of petroleum samples are common, there has been little work done that employs a high-performance liquid chromatography (HPLC) separation prior to FT-ICR MS analysis. In this work, distillate cut and treated gas oils are separated on a commercially available dinitrophenyl “DNAP” column and the fractions are collected offline and analyzed by FT-ICR MS. HPLC separations on the “DNAP” column are shown, along with separations on a custom synthesized HPLC column (HC-Tol). Four peak regions are identified on the “DNAP” chromatograms, and the nitrogen species identified in each fraction using positive and negative electrospray ionization are discussed. It was found that pyridines are present in the first three fractions, while pyrroles are present in fractions 2 and 3. Acids are isolated in fraction 4. Alkylation of nitrogen species decreases their level of retention on the “DNAP” column, and the HPLC chromatograms can be used to qualitatively compare nitrogen content between samples. A comparison of HPLC fraction data to MS analysis of unfractionated gas oils found that fraction data are not adequate for quantitative comparisons of carbon number and double bond equivalents between samples.

Chemical Composition and Biological Activities of Polar Extracts and Essential Oil of Rose‐scented Geranium, Pelargonium graveolens

Pelargonium graveolens (Geraniaceae) was characterized with respect to its chemical composition, antioxidant potential and antimicrobial activities. This is the first investigation focusing on the comparison of both essential oil and polar extracts from this species. The chemical composition of the essential oil of the aerial parts of P. graveolens was analyzed by gas chromatography/mass spectrometry. The main constituents of the oil were found to be β-citronellol (21.9%), citronellyl formate (13.2%), geraniol (11.1%), 10-epi-γ-eudesmol (7.9%), geranyl formate (6.2%) and (l)-linalool (5.6%). Nine flavonoids were identified by high-performance liquid chromatography-MS in leaf and flower extracts. Kaempferol 3-O-rhamnoside-glucoside, isorhamnetin aglycone, quercetin 3-O-glucoside, kaempferol 3,7-di-O-glucoside, quercetin 3-O-pentose and kaempferol 3-O-glucoside, quercetin 3-O-rhamnoside-glucoside, quercetin 3-O-pentoside-glucoside, myrisetin 3-O-glucoside-rhamnoside flavonoids were detected in methanolic and aqueous extracts, respectively. The total flavonoids ranged between 29.9 and 78.2 mg QE/g in flower water and methanol extracts, respectively, and 22.5 and 71.2 mg QE/g dry weight in leaf water and methanol extracts, respectively. The highest antioxidant activities using two methods of free radical scavenging capacities were obtained with the essential oil (9.16 mM of Trolox and 2.68 µg/ml). All P. graveolens essential oil and polar extracts were active against at least one bacterium.