Positive Electrospray Ionization Mass Spectrometry Research Articles

Transfer of alkaloids from Delphinium to Castilleja via root parasitism. Norditerpenoid alkaloid analysis by electrospray mass spectrometry

Norditerpenoid alkaloids which are present in the roots of Delphinium occidentale are assimilated into its root parasite, Castilleja sulphurea. The alkaloids are found in all plant parts of the Castilleja (roots, stems, leaves, flowers, seeds) and in the flower nectar. The major root alkaloids (browniine, delcosine, deltaline and dictyocarpine; analyzed by positive ion electrospray mass spectrometry, ESMSH +) were transmitted into the Castilleja. Little or none of the highly mammalian toxic alkaloids (such as methyllycaconitine) were found in D. occidentale. ESMSH + also was used to identify alkaloids of D. barbeyi, where browniine, delcosine and a small amount of methyllycaconitine (roots only) were found. ESMSH + provided a useful method for analysis of norditerpenoid alkaloids. Their presence as assimilates in a root parasite plant could provide a defense against herbivory.

Analysis of terfenadine in human plasma using microbore high-performance liquid chromatography–electrospray ionisation mass spectrometry

An assay based on combined microbore high-performance liquid chromatography–positive ion electrospray ionisation mass spectrometry with selected ion recording has been developed for the measurement of the antihistamine drug terfenadine in human plasma. A deuterated analogue of terfenadine was synthesised for use as an internal standard and extraction of terfenadine was carried out on C 18 solid phase extraction columns. The limit of detection of terfenadine in plasma is 0.1 ng/ml and the intra-assay coefficient of variation at 1 ng/ml is 10.1%. Plasma concentrations of terfenadine measured in six normal subjects following a 120 mg oral dose are reported.

Quantitative determination of BMS186716, a thiol compound, in dog plasma by high-performance liquid chromatography-positive ion electrospray mass spectrometry after formation of the methyl acrylate adduct.

As it is extremely unstable in blood, the thiol compound BMS186716 was stabilized by the addition of methyl acrylate (MA) to blood samples. The blood samples were then kept in ice for 10-15 min for completion of the Michael addition reaction to occur between the thiol group of BMS186716 and MA, after which the plasma was separated by centrifugation under refrigeration. For sample analysis, the standard and quality control samples were prepared by spiking blank plasma with the BMS186716-MA adduct. After addition of the internal standard, BMS 188035-MA, each sample was acidified with HCI and then extracted with methyl tert.-butyl ether. Each reconstituted extract was injected into a high-performance liquid chromatography-positive ion electrospray ionization mass spectrometric system. The electrospray condition was chosen to enhance the [M+NH4]+ signal at the expense of the [M+H]+ signal. Monitoring the [M+NH4]+ signal, a lower limit of quantitation of 2.5 ng/ml was achieved, with 0.5 ml plasma. We have thus shown that a sulfhydryl compound (BMS186716) in blood can successfully be stabilized by reacting it with MA and that the adduct produced is adequately stable in blood and plasma to allow the development of a rugged quantitative bioanalytical method.

A New Derivatization Reaction for the Identification and Determination of Organic Acids by Positive-ion Electrospray Mass Spectrometry

A new derivatization reaction was developed for the characterization of organic acids by electrospray ionization mass spectrometry in the highly sensitive positive-ion mode, using N,N′-dicyclohexylcarbodimide and N,N′-dimethylethylenediamine. Suitability of the reaction was demonstrated on different types of organic acids. When selected-ion monitoring was applied, the detection limit was at the low femtomol level. © 1997 John Wiley & Sons, Ltd.

Organotin halides and their hydrolysed species studied in solution by positive and negative ion electrospray mass spectrometry

Hydrolysis products of the organotin compounds R 3SnCl (R = Ph, Me or Bu n), Ph 3SnBr and Ph 2SnCl 2 have been investigated by electrospray mass spectrometry. Both positive and negative ion modes were employed to obtain information on the range of charged species produced in aqueous acetonitrile solution. The positive ion electrospray spectra of the triorganotin species yielded aggregates [(R 3Sn) n (OH) n-1 ] +, alongside R 3Sn + and solvated cations, including [R 3Sn(NCCH 3) +]. Addition of pyridine (py) to hydrolysed Ph 3SnCl afforded predominantly [Ph 3Sn(py)] +. In the negative ion spectra a number of species could be identified. Solutions of Ph 3SnX (X = Cl or Br) yielded [Ph 3SnX 2] −, [Ph 3SnX(OH)] −, [Ph 3SnO] − and [Ph 3SnOSnPh 3X] −, whilst those of Ph 2SnCl 2 showed chiefly [Ph 2SnCl 3] −. Me 3SnCl yielded [Me 3SnCl 2] − accompanied by [Me 3SnOSnMe 3(OH)] −, whereas Bu 3 nSnCl gave [Bu 3 nSnCl 2] − and [Bu 3 nSnCl(OH)] −. Addition of Cl −, Br − and I − to a hydrolysed solution of Bu 3 nSnCl allows identification of all six mixed dihalostannate complexes [Bu 3 nSnXY] − (X, Y = Cl, Br or I).

An electrospray mass spectrometric study of organomercury(II) and mercuric interactions with peptides involving cysteinyl ligands

Positive ion electrospray mass spectrometry (MS) has been used to investigate the interaction of Hg2+, [MeHg]+ and [PhHg]+ cations with cysteine (Cys), glutathione (GSH) and a 27-residue polypeptide (Pp) containing one cysteinyl sulfur located at the N-terminus. MSIMS experiments showed that organomercury adduction occurs primarily at the sulfhydryl group, with some evidence for isomeric species in which the organomercury cation is bound to either an amino or a carboxylic group. Following Ellman modification of GSH and Pp, the maximum number of adducted organomercury cations was reduced by 2 and 1, respectively, indicating a 2:1 and a 1:1 interaction between [RHg]+ and the cysteinyl sulfur. Unlike [PhHg]+, [MeHg]+ showed an almost exclusive affinity for the cysteinyl sulfur of GSH and Pp, Both Cys and GSH reacted with Hg2+ to form polynuclear species. Collisional activation mass spectra of the [2Cys + Hg -H]+ ion indicated that the Hg(II) was most probably bridged between the two cysteinyl residues, although not necessarily via a conventional -S-Hg-S- linkage.

Microbore liquid chromatography–electrospray mass spectrometry of selected synthetic pyrethroid insecticides

The pyrethroid insecticides kadethrin, α-cypermethrin, flucythrinate and SSI-116 have been studied by positive-ion electrospray mass spectrometry (+ESMS) in the presence of ammonium acetate and formic acid. Ammoniated molecule base peak ions [M + NH4]+ were observed for all the insecticides studied at low electrospray source sampling cone voltages. The effect of increasing the cone voltage (40–120 V) and its influence on the extent of fragmentation experienced by each insecticide were studied. A number of these key fragment ions found in +ESMS spectra of α-cypermethrin have been examined by MS–MS under low-energy collisional activation (CA) conditions. On-line microbore reversed-phase liquid chromatographic separations were performed on mixed pyrethroid standards. The eluates were analysed by +ESMS to establish the lower limits of detection using full-scan and selective-ion recording (SIR) modes. Limits of detection (signal-to-noise ratio better than 3:1) for each component of the pyrethroid mixture, injected on column, were in the range 120–300 pg (0.30–0.77 pmol) using full-scan mode and 12–60 pg (0.03–0.15 pmol) by SIR.

Determination of lipid ester ozonides and core aldehydes by high-performance liquid chromatography with on-line mass spectrometry

Unsaturated triacylglycerols (TG) and choline (PC) and ethanolamine (PE) phosphatides of known structure were subjected to ozonization and reduction with triphenylphosphine to yield the corresponding lipid ester core aldehydes. Mono- and di-C 9 aldehyde palmitoylglycerols were prepared from oleoyldipalmitoyl and oleoyllinoleoylpalmitoyl glycerols, respectively, while egg yolk PC and PE provided the mono-C 5 and mono-C 9 aldehydes of palmitoyl-and stearoyl glycerophospholipids. The aldehydes were isolated in the free form and as the dinitrophenylhydrazone (DNPH) derivatives by thin-layer chromatography (TLC). The intermediate ozonides, free aldehydes and hydrazones were identified by reversed phase high performance liquid chromatography (HPLC) with on-line negative ion thermospray and normal phase HPLC with on-line positive ion electrospray mass spectrometry (LC-MS). The synthetic aldehydes were used as carriers during isolation from natural sources and as reference compounds in quantitative analyses

Investigation of complexes between some glycopeptide antibiotics and bacterial cell‐wall analogues by electrospray‐ and capillary zone electrophoresis/electrospray‐mass spectrometry

Complexation in aqueous solutions between vancomycin, ristocetin A, teicoplanin and two bacterial cell-wall analogues, Ac2-L-Lys-D-Ala-D-Ala (tripeptide) and Ac-D-Ala-D-Ala (dipeptide) has been examined by positive-ion electrospray mass spectrometry (ES-MS) and capillary zone electrophoresis (CZE)/ES-MS. The ES-MS data demonstrate that as well as complexes between monomeric antibiotics and either peptide, the investigated solutions contained complexes ranging from the simple homodimer of the antibiotic to a more complex association giving ions of the type [2(antibiotic) + 3(tripeptide) + 3H]3+. The same data also demonstrate that the homodimers of the investigated antibiotics are significantly suppressed in the presence of the tripeptide. The use of CZE/ES-MS made it evident that the complexes between the antibiotic and the tripeptide were present in the solution prior to their introduction into the ion source. The two sets of data are compared with existing UV difference spectroscopy and nuclear magnetic resonance data on complexation and dimerization.

Effect of Solution Ionic Strength on Analyte Charge State Distributions in Positive and Negative Ion Electrospray Mass Spectrometry

The variability of ESMS charge state distributions was investigated for a number of analytes when the ionic strength of their solutions was altered over a wide range. Solution ionic strength was adjusted by adding varying amount of electrolytes (NH 4 OAc, CsCl or (n-C 4 H 9 ) 4 N + Cl - ) to protein solutions of myoglobin and lysozyme. Measured solution QH values were found to be nearly constant over the entire range of CsCl and (n-C 4 H 9 ) 4 N + Cl - addition, and a slight increase in QH was observed upon NH 4 OAc addition. Analyte peak intensities in positive ion ESMS reproducibly decreased as the solution ionic strength increased. However, analyte charge state distributions remained quite constant desite 4 orders of magnitude variation in electrolyte concentration

Effects of Anions on the Positive Ion Electrospray Ionization Mass Spectra of Peptides and Proteins

Positive ion electrospray ionization mass spectra of polypeptides are usually obtained from solutions that are acidified and therefore contain relatively high concentrations of anions. The present study describes an investigation of the effects of these ubiquitous anions on the positive ion electrospray ionization mass spectra of peptides and proteins. Certain anionic species in the spray solutions were observed to cause a marked decrease in the net average charge of peptide and protein ions in the mass spectra compared to the average charge measured in the absence of these anions. This charge neutralization effect was found to depend solely on the nature of the anionic species and was independent of the source of the anion (acid or salt), with the propensity for neutralization following the order: CCl3COO- > CF3COO- > CH3COO- approximately Cl-. A mechanism for the observed charge reduction effect is proposed that involves two steps. The first step occurs in solution, where an anion pairs with a positively charged basic group on the peptide. The second step occurs during the process of desolvation or in the gas phase, where the ion pair dissociates to yield the neutral acid and the peptide with reduced charge state. The different propensities for charge neutralization of the different anionic species is presumed to reflect the avidity of the anion-peptide interaction. These findings demonstrate that any attempt to correlate the distribution of charge states observed on proteins in the gas phase (by positive ion electrospray ionization mass spectrometry) with the net charge residing on the protein in solution will require that the described anion effect be taken into account. In addition, it appears that some control over the distribution of charge states on peptides and protein ions can be exercised by an appropriate choice of anion in the electrospray solution.

Positive and negative ion electrospray mass spectrometric studies of some amino acids and glutathione, and their interactions with alkali metal ions and methylmercury(II)

Positive and negative ion electrospray mass spectrometry (ESMS) has been used to investigate solutions of some simple amino acids (glycine, histidine, methionine and cysteine) and the small peptide glutathione. Negative ion ESMS detected the simple anions [AA-H −] in each case (AA=amino acid) and often the dimeric species [(AA) 2-H] − were also observed. In the positive ion mode ESMS allowed the observation of the protonated cluster ions [H(AA) x ] + ( x=1–4). Some interactions between alkali metal ions and the amino acids were observed, but in general the peaks in the ES mass spectra were weak and the alkali metal complexation could not compete with protonation. In contrast, methylmercury(II) reacted strongly with the amino acids and glutathione. In addition to species of 1:1 stoichiometry, all the amino acids gave peaks in their ES mass spectra corresponding to the ions [(MeHg) 2(AA-H)] +, while glutathione gave a clear indication of the formation of a 3:1 species.

Identification of molecular species of glycerophospholipids and sphingomyelin using electrospray mass spectrometry.

This paper describes the use of positive and negative ion electrospray mass spectrometry (MS) and MS/MS (tandem mass spectrometry) to identify glycerophospholipid and ceramide headgroups and their alkyl, alkenyl and acyl constituents. Molecular ion adducts were the primary products formed by positive ionization, occurring as [M+H]+, [M+Na]+, [M+K]+, [M+formate]+, or [M+acetate]+, depending upon the class of glycerophospholipid and the presence or absence of these ionization-promoting species. Similar (negatively charged) ions corresponding to the loss of the groups listed above were formed in negative ion MS. Positive ion electrospray MS/MS provides information on the nature of the headgroup, with the formation of an ion corresponding to the headgroup itself, or the loss of the headgroup from the molecular ion H+ or Na+ adduct. Acyl constituents are identified during negative ion MS/MS from the formation of their RCOO- ions. The nature of alkyl or alkenyl substituents in glycerophosphoethanolamine (PE) molecular species can be identified from residual ions following the loss of ethanolamine plus loss of the acyl moiety in the sn-2 position, and cyclization of a phosphate oxygen with C-2 of glycerol. In glycerophosphoinositol (PI) species, it appears that an RCO- ion is formed during negative ion MS/MS, possibly to steric interference from the bulky phosphoinositol headgroup that prevents cyclization (and subsequent stabilization) of the ion described for PE species. Positive and negative ion electrospray MS spectra for molecular species of commercial preparations of PE, PI, phosphatidylserine (PS), glycerophosphocholine (PC) and sphingomyelin (SM) produced similar profiles. For phospholipids occurring as Na+ adducts, concentrations above ca. 1 ng/microliter produced significant quantities of both [M+H]+ and [M+Na]+ ions for those molecular species present in the largest quantities, complicating interpretation of the spectra. Complete profiles of molecular species were obtained from as little as 10 picograms of material. Major components of PE were identified from 0.1 picogram total lipid. Using single ion monitoring of the Na+ adduct of beta-acetyl-gamma-O-hexadecyl L-alpha-phosphatidylcholine, 10 femtograms of material was detected. A mixture of 1 nanogram each of PE, PI, PS, and PC was readily resolved into individual molecular species, with little apparent loss of resolution or preferential ionization. Electrospray MS did not provide information on the position (sn-1 or sn-2) of fatty acids, and was not capable of differentiating in all instances between alkyl-acyl and alkenyl-acyl substituents without prior separation of these lipid subclasses.(ABSTRACT TRUNCATED AT 400 WORDS)

Electrospray mass spectrometry for characterization of lipid a fromEnterobacter agglomerans

Negative ion and positive ion electrospray mass spectrometry have been employed to characterize the lipid A mixture produced by hydrolysis of lipopolysaccharides from Enterobacter agglomerans, a Gram-negative bacterium commonly found in field cotton. Neutral monophosphoryl lipid A molecules form stable anions via deprotonation, but adduct formation via cation attachment occurs in low yield. Dephosphorylated lipid A molecules, on the other hand, readily form adducts with Na+, whereas deprotonation occurs in low yield. The mass spectra of lipid A produced by E. agglomerans reveal the presence of lipid A ions which differ in the nature of attached fatty acid side chains. At least two heptaacyl forms of lipid A are present, one of which has a structure which appears to be the same as the structure of heptaacyl lipid A produced by Salmonella minnesota. The second structure differs only by the nature of the side chain at position 3' of the disaccharide backbone where a hydroxymyristoyloxymyristoyl group replaces the myristoyloxymyristoyl substituent. Collisionally activated dissociations prior to mass analysis enable the identification of fragment ions which can be distinguished from at least eight intact deprotonated molecules present in crude lipid A.

Through the skin barrier with NDELA: Bronaugh, R. L., Congdon, E. R. & Scheuplein, R. J. (1981). The effect of cosmetic vehicles on the penetration of N-nitrosodiethanolamine through excised human skin. J. invest. Derm. 76, 94

A rapid, sensitive, accurate and specific ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC–MS/MS) method for the detection of N-nitrosodiethanolamine (NDELA), a highly toxic contaminant in cosmetic raw materials and products was developed and validated. Systematized sample preparation steps were developed according to product types. Various SPE cartridges and columns were examined to establish the condition of SPE and chromatographic separation for NDELA. Sample cleanup steps consisting of solvent and liquid–liquid extraction tailored to the various sample matrix types were established prior to mixed mode SPE (Bond Elut AccuCAT). Chromatographic separation was achieved within 7 min on a porous graphitic carbon (PGC) column using a gradient elution with the mobile phase of 1 mM ammonium acetate containing 0.1% acetic acid and methanol. NDELA was monitored using an electrospray positive ionization mass spectrometry in the multiple reaction monitoring (MRM) mode (m/z 134.9>103.7(quantifier) and 73.7(qualifier ion)) with d8-NDELA (m/z 143.1>111.0) as internal standard. The standard curves were linear over the concentration range of 1–100 ng/mL with a correlation coefficient higher than 0.99. The limit of detection (LOD) and the limit of quantification (LOQ) was 10 and 20 μg/kg, respectively (0.5 and 1 ng/mL in standard solution). The intra- and inter-day precisions were estimated to be below 11.1% and accuracies were within the range of 90.8–115.8%. The validated method was successfully applied to the analysis of real samples including raw materials, skin care, make-up, shampoos and hair products.