q value and parent energy optimization using a low-voltage ionization approach increases resolution in linear ion trap mass spectrometry.

In this work, the isolation step in the linear ion trap was performed using different "q values" conditions at a low collision-induced dissociation (CID) energy leading to the parent ion resolution improvements, reasonably due to better ion energy distribution. According to the results, we obtained a greater resolution and mass accuracy operating in both traditional electrospray and low voltage ionization near the q value = 0.778 and with a CID energy of 10%. This effect was evaluated with low-molecular-mass compounds (skatole and arginine). The proposed optimization yielded a superior instrument performance without adding technological complexity to mass spectrometry analyses.

Anomeric discrimination and rapid analysis of underivatized lactose, maltose, and sucrose in vegetable matrices by U-HPLC-ESI-MS/MS using porous graphitic carbon.

Lactose intolerance is a common condition caused by intestinal lactase deficiency, and a lactose-free diet represents the simplest way to avoid gastrointestinal symptoms. The emerging use of dietary supplements requires analytical tools that are capable of assessing these analytes, particularly for those based on dry herbal extracts that contain lactose together with maltose and sucrose, because of cross-contamination and/or deliberate addition as excipient. Electrospray ionization mass spectrometry (ESI-MS) and MS/MS are valuable detection methods for underivatized disaccharides; however, the absence of distinctive ions and collision-induced dissociation (CID) fragmentation patterns does not allow discrimination of stereoisomers without good chromatographic resolution. We developed an ultrahigh performance liquid chromatography-ESI (U-HPLC-ESI) approach, based on porous graphitic carbon (PGC) columns, working at 5 °C to separate and detect the disaccharides in their anomeric forms as formate adducts obtained directly in-column by eluting with formate buffer/acetonitrile gradient mixtures. Using a Paul trap, we monitored the adducts [M + HCOO](-) at m/z 387 in ESI negative mode (MS(1)) as well as the CID fragment ion [M - H](-) at m/z 341 (MS(2)) and used MS(3) fragment ions at m/z 178 and 161 to confirm disaccharides identity in complex vegetable matrices. Complete resolution of lactose α- and ß-anomers, maltose α- and ß-anomers, and sucrose was obtained with R ≥ 2.0 for all peaks and selectivity α = 1.2 between α- and ß-anomers of lactose. The limits of detection were in the range of 3-7 µg/l (ppb) for the target disaccharides. Because of the rapidity and good anomeric discrimination, the described method represents an alternative tool to investigate the mutarotation phenomenon for reducing disaccharides.

Simultaneous quantification of 8-iso-prostaglandin-F(2alpha) and 11-dehydro thromboxane B(2) in human urine by liquid chromatography-tandem mass spectrometry.

Both F(2)-isoprostanes (8-iso-PGF(2alpha)), a well-known marker of oxidative stress, and thromboxanes A(2) (TXA(2)) are involved in atherosclerosis through LDL oxidation and platelet activation. Different aspects of the pathology can be described by 8-iso-PGF(2alpha) and TXA(2) so it is important to determine both their concentrations to monitor the disease progression and/or therapy effects. We developed a simple and sensitive method based on liquid chromatography-tandem mass spectrometry, using electrospray ionization in negative-ion mode, for the simultaneous measurement of the concentration of 8-iso-PGF(2alpha) and 11-dehydro thromboxane B(2) (11-DH-TXB(2)), a TXA(2) metabolite. This method was applied to analyze urine samples collected overnight from 15 atherosclerotic patients, with documented carotid artery sclerosis (CAS), and from 20 controls. The detection limit was 0.097pg/microL for 8-iso-PGF(2alpha) and 0.375pg/microL for 11-DH-TXB(2), with a linear range of 0.78-25pg/microL; the inter- and intraday imprecision was <5% for both metabolites. These analytes were higher in CAS (P<0.005 vs controls) and were positively correlated in patients but not in controls, even after adjustment for age and gender (r=0.60; P=0.032). This highly sensitive, precise, and rapid method allows for the simultaneous determination of 8-iso-PGF(2alpha) and 11-DH-TXB(2) in human urine samples in order to evaluate oxidative stress and platelet aggregation.

Unexpectedly dangerous escargot stew: oleandrin poisoning through the alimentary chain.

A female, aged 43 and a male, aged 66, experienced gastrointestinal and cardiovascular symptoms after a meal including snail stew. Twelve hours after the ingestion, they presented with nausea, vomiting, diarrhea, and cardiovascular symptoms typical of acute toxic digoxin ingestion and were hospitalized. The man's electrocardiogram was altered, and the woman's was normal. Serum digoxin levels, measured on a Roche COBAS Integra 800 with the Roche On-Line Digoxin reagent, were 1.14 and 1.00 nmol/L, respectively. Potassium levels were normal in both patients. The serum digoxin concentration decreased on the second day, and symptoms resolved on the third day with patients fully recovered (i.e., reversion to a normal sinus rhythm). Cardiac-glycoside-like intoxication symptoms follow the ingestion of leaves or flowers of Nerium oleander. The consumed snails were suspected to be responsible for the intoxication. In the homogenized snail tissue, the concentration expressed in digoxin equivalents was 0.282 nmol/g. The presence of oleandrin and oleandrigenin in the snails was confirmed by liquid chromatography-tandem mass spectrometry analysis, which was performed on a ionic-trap Finnigan LXQ instrument using an electrospray ionization interface. High-pressure liquid chromatographic separation was performed on a C18 column with a gradient of methanol/water. An extract of oleander leaves was used as reference.

Surface-activated chemical ionization and high-flow gradient chromatography to reduce matrix effect.

The new atmospheric pressure chemical ionization source, named surface-activated chemical ionization (SACI), has been used in conjunction with high-flow gradient chromatography to reduce the matrix effect. This high-flow gradient chromatography approach avoids the co-elution of analyte and biological matrix compounds that leads to a reduction in quantitation errors due to matrix effect. However, this approach cannot be employed with the classical electrospray ionization (ESI) source that usually works at low eluent flow (< 300 microL/min). SACI can work at high eluent flow (100-2000 microL/min) and can be employed in conjunction with high-flow gradient chromatography. The reduction in matrix effect in tacrolimus analysis in protein-precipitated blood samples, an important immunosuppressive agent for renal transplantation, is presented and discussed.

Surface-activated chemical ionization ion trap mass spectrometry in the analysis of drugs in dilute urine samples. Part II: analysis of morphine and other street drugs.

The new ionization method, called surface-activated chemical ionization (SACI), was employed for the analysis of fives drugs (morphine, codeine, 6-monoacetylmorphine (6-MAM), benzoylecgonine and cocaine) by ion trap mass spectrometry. The results so obtained have been compared with those achieved by using atmospheric pressure chemical ionization (APCI), no-discharge-APCI and electrospray ionization (ESI) clearly showing that SACI is the most sensible one mainly due to the high ionization efficiency and the lower chemical noise. The performance of SACI in terms of sensitivity and linearity was compared with the sensitivity and linearity obtained using APCI, no-discharge-APCI and ESI, showing that the new SACI approach gives rise to the best results. Then, SACI was used to analyze morphine, codeine, 6-MAM, benzoylecgonine and cocaine in urine samples. After the optimization of the instrumental parameters for a mixture of the standard compounds, eight urine samples were analyzed. They were strongly diluted (1 : 20 and 1 : 100) in order to prevent the chromatographic column damage due to the matrix composition. Furthermore, the diluted urine samples were directly analyzed, without pretreatment, through LC-MS and LC-MS/MS, and the obtained results are reported.

The role of different phenomena in surface-activated chemical ionization (SACI) performance.

In previous studies, the production of ions in an APCI source without any corona discharge was observed, and the intensity of the ion signals showed significant increases on placing a metallic surface at 45 degrees inside an orthogonal ion source. This method was named surface-activated chemical ionization (SACI). The present study was performed to investigate the mechanisms of ion production with or without the presence of the metallic surface, by varying instrumental parameters and the geometrical configuration. Approximate calculations show that, in the absence of corona discharge and of any additional surfaces, ions cannot be produced by collisional phenomena, because of their low kinetic energy, in the 10(-2) to 10(-3) eV range. Two alternative possibilities have been considered: the first takes into account that ions may originate by collision of neutral clusters of polar solvent molecules with the APCI source surfaces through clusterelectric effect. The second takes into account that the water dissociation constant k(w) is temperature dependent, passing from 10(-14.1669) at 20 degrees C to 10(-12.4318) at 90 degrees C. It means that the [H(+)] varies from 8.3 x 10(-8) to 6.1 x 10(-7) M going from 20 to 90 degrees C. Hence, at the high temperatures experimented in the APCI vaporizer, H(+) becomes available in solution in molar quantities analogous to those of analyte, and the protonation of the analyte itself can consequently occur. The activation of further ionization processes in the presence of the metallic surface can be reasonably attributed to interactions between gas-phase analyte molecules and solvent molecules adsorbed on the surface. Experiments performed with a thin layer of deuterated glycerol on the surface led to unequivocal results, i.e. the production of [M + D](+) ions of the analyte.

Surface-activated chemical ionization in the analysis of arginine in plasma samples.

Alterations of arginine plasma levels are involved in several disorders of amino acid metabolism such as hurtnup, argininosuccinic aciduria, histidinemia, citrullinuria, and cystinuria. In this work a new liquid ionization source, surface-activated chemical ionization (SACI), has been used to analyze arginine in human and rat plasma samples. Arginine was extracted and diluted ten times through protein precipitation. The diluted arginine samples were then analyzed using an ion-exchange chromatographic column coupled with the SACI source and an ion trap analyzer using MS(3) monitoring in order to increase the sensitivity and specificity of the analysis. The multiple-point standard additions method was used to quantify the arginine. This method was employed to eliminate the matrix effect that affects all liquid ionization sources (APCI, ESI, SACI, etc.), and also does not require use of an internal standard. High-quality results in terms of sensitivity, limit of detection, lower limit of quantitation, linearity and reproducibility, are demonstrated.

Surface-activated no-discharge atmospheric pressure chemical ionization.

A new ionization method named surface-activated chemical ionization (SACI) has been realized. In this invention a commercially available atmospheric pressure chemical ionization (APCI) chamber, employed without any corona discharge (no-discharge APCI), has been modified with the insertion of a gold surface, leading to a significant improvement in the ionization efficiency. The ionization of the sample takes place by both gas-phase and surface-activated processes. This new ionization source is able to generate ions with high molecular mass and low charge states, leading to improved sensitivity and reduced noise. The new device has been tested in the analysis of some peptides. A comparison between the performance with and without the presence of the surface, and the optimization of the operating conditions (nebulizing gas flow, sample solution flow, pH of solution, and surface area), are reported and discussed.

Determination of 17beta-estradiol in bovine plasma: development of a highly sensitive technique by ion trap gas chromatography-tandem mass spectrometry using negative chemical ionization.

A novel approach to the determination of 17beta-estradiol in bovine plasma is presented. The observed enhanced sensitivity is gained by the application of tandem mass spectrometry (MS) fragmentation to a stable, well characterized negative ion produced by chemical ionization (methane as reagent gas). A specific derivatizing reactant is employed (pentafluorobenzyl bromide), combined with bis-trimethylsilyltrifluoroacetamide, to favor the formation of a diagnostic precursor negative ion. Plasma samples are purified through a C18 solid phase extraction column and derivatized before gas chromatography-MS analysis. The accuracy and the precision of the method, tested over a set of spiked samples, were satisfactory. The limit of detection was found to be 5 pg ml(-1) and the limit of quantification was fixed at 20 pg ml(-1). The fragmentation pattern is fully explained and the method is applicable for the official analysis of bovine plasma for the detection of 17beta-estradiol according to the European criteria 256/93 and to the draft SANCO/1805/2000 rev. 3. The quantification of incurred positive samples was performed according to the proposed procedure and compared with the results obtained by standardized radio immuno assay; the estimated concentrations were significantly similar.

Analysis of peptides using partial (no discharge) atmospheric pressure chemical ionization conditions with ion trap mass spectrometry.

A novel approach, based on the use of atmospheric pressure chemical ionization ion trap mass spectrometry (APCI-ITMS) conditions, but without using corona discharge, was used to analyze peptides. The proposed method was applied to three standard peptides (bombesin, trityrosine and tyrosine-glycine-glycine) as well as peptides obtained through enzymatic digestion of two standard proteins (horse cytochrome c and horse myoglobin).

Analysis of protein ions in the range 3000-12000 Th under partial (no discharge) atmospheric pressure chemical ionization conditions using ion trap mass spectrometry.

A new approach, based on the use of atmospheric pressure chemical ionization ion trap mass spectrometry (APCI-ITMS), but without a corona discharge, was investigated for application to creating and monitoring protein ions. It must be emphasized that APCI is not usually used in protein analysis. In order to verify the applicability of the proposed method to the analysis of proteins, two standard proteins (horse cytochrome c and horse myoglobin) were analyzed. A mixture of the two proteins was also analyzed showing that this novel approach, based on the use of APCI, can be used in the analysis of protein mixtures.