Biomedical subjects

L Elviri

Publications and source records attributed to L Elviri.

14 recordsLinked to original sources

PubMed2006

Matrix effects on accurate mass measurements of low-molecular weight compounds using liquid chromatography-electrospray-quadrupole time-of-flight mass spectrometry.

Liquid chromatography (LC) with high-resolution mass spectrometry (HRMS) represents a powerful technique for the identification and/or confirmation of small molecules, i.e. drugs, metabolites or contaminants, in different matrices. However, reliability of analyte identification by HRMS is being challenged by the uncertainty that affects the exact mass measurement. This parameter, characterized by accuracy and precision, is influenced by sample matrix and interferent compounds so that questions about how to develop and validate reliable LC-HRMS-based methods are being raised. Experimental approaches for studying the effects of various key factors influencing mass accuracy on low-molecular weight compounds (MW < 150 Da) when using a quadrupole-time-of-flight (QTOF) mass analyzer were described. Biogenic amines in human plasma were considered for the purpose and the effects of peak shape, ion abundance, resolution and data processing on accurate mass measurements of the analytes were evaluated. In addition, the influence of the matrix on the uncertainty associated with their identification and quantitation is discussed. A critical evaluation on the calculation of the limits of detection was carried out, considering the uncertainty associated with exact mass measurement of HRMS-based methods. The minimum concentration level of the analytes that was able to provide a statistical error lower than 5 ppm in terms of precision was 10 times higher than those calculated with S/N = 3, thus suggesting the importance of considering both components of exact mass measurement uncertainty in the evaluation of the limit of detection.

Calibration↗

PubMed2006

Mass spectrometry techniques for detection of ligand-dependent changes in the conformational flexibility of cellular retinol-binding protein type I localized by hydrogen/deuterium exchange.

Hydrogen/deuterium exchange, measured by electrospray ionization with orthogonal quadrupole time-of-flight mass spectrometry (ESI-Q-TOFMS) and by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS), was used as a means to probe and map differences in conformational flexibility between the ligand-free and ligand-bound forms of cellular retinol-binding protein type I. Labelled fragments were obtained by digestion of the protein with pepsin. The differences in space-resolved time courses of deuterium incorporation identified regions that exhibit a remarkably higher degree of flexibility in the apo-protein than in the holo-protein. These segments encompass residues that are thought, on the basis of structural homology of the retinol carrier with other members of the intracellular lipid-binding proteins family, to belong to the dynamic portal through which all-trans retinol can access its high-affinity, solvent-shielded, binding site.

Amino Acid Sequence↗

PubMed2005

Investigation of the separation of heterocyclic aromatic amines by reversed phase ion-pair liquid chromatography coupled with tandem mass spectrometry: the role of ion pair reagents on LC-MS/MS sensitivity.

Reversed phase ion-pair chromatography (RP-IPC) of seven heterocyclic aromatic amines encompassing quinoline (IQ, MeIQ), quinoxaline (MeIQx), pyridine (PhIP) and carboline derivatives (AalphaC, Harman, Norharman) was carried out with formate as counter ion in an aqueous eluent with acetonitrile as organic modifier. TSKgel ODS-80TS was used as the stationary phase. With the aim of acquiring a better insight into the mutual influence of ion-pair reagent and the organic modifier upon solute retention, the study was performed by using an experimental design approach able to evidencing the effect of the simultaneous variation of the two factors. A model for the chromatographic behavior of the amines is proposed that includes classical ion-pair mechanism involving formate in the case of MeIQx, PhIP, Harman and Norharman. A competitive ion-exchange mechanism was hypothesized to govern retention of quinoline compounds, whereas electrostatic interactions and hydrogen bond formation with the silanols of the stationary phase were judged to be responsible for the retention of AalphaC. Further, the chromatographic behavior of the analytes using the formic acid-ammonium formate buffer in the mobile phase was compared with that observed using acetic acid-ammonium acetate buffer. The method based on the use of RP IPC with tandem mass spectrometry when the eluent contained formate buffer at pH 2.8 exhibited higher detectability with respect to that achieved using the acetate buffer.

Acetonitriles↗

PubMed2005

Rapid assay for analyzing biogenic amines in cheese: matrix solid-phase dispersion followed by liquid chromatography-electrospray-tandem mass spectrometry.

A new rapid and sensitive method based on matrix solid-phase dispersion (MSPD) followed by liquid chromatography-electrospray-tandem mass spectrometry was devised for the determination of biogenic amines at trace levels in cheese samples. The method required 0.25 g of sample, CN-bonded silica as a dispersant sorbent, and a formic acid aqueous solution/methanol mixture as an eluting solvent. Extraction recoveries from soft cheese products were calculated in the 98 +/- 4-110 +/- 6% range. A procedure based on solid-phase extraction was also evaluated for the extraction of these compounds in cheese. Chromatographic separation was performed using a C18 column with an aqueous ammonium acetate/methanol mixture as the mobile phase under gradient conditions. The method was validated in terms of detection limits (LOD), quantitation limits (LOQ), linearity, recovery, precision, and trueness. Results in the 0.05-0.25 mg kg(-1) range were obtained for the LOD of histamine, tyramine, and beta-phenylethylamine in soft cheese samples. Linearity was established over 2 orders of magnitude. Excellent precision in terms of intra-day repeatability was calculated (RSD% < 5). The applicability of the method to the determination of biogenic amines in cheese products was demonstrated.

Biogenic Amines↗

PubMed2004

Accurate mass measurements for the confirmation of Sudan azo-dyes in hot chilli products by capillary liquid chromatography-electrospray tandem quadrupole orthogonal-acceleration time of flight mass spectrometry.

The potential of capillary liquid chromatography (microLC)-quadrupole/time-of-flight mass spectrometry (Q-TOF MS) for the confirmation of Sudan I, II, III and IV azo-dyes as contaminants in hot-chilli food products was demonstrated. Using the microLC-electrospray ionization (ESI)-Q-TOF MS technique, accurate mass measurements of Sudan dyes were performed both on standard solutions and on matrices. Precision of exact mass measurements was calculated taking into account the ion statistics according to the number of ion sampled in the measurement. Accurate mass measurements by MS/MS experiments were performed to elucidate azo-dye fragmentation patterns. Selectivity of the microLC-Q-TOF MS method was assessed by evaluating matrix suppression effects by pre-column injection of blank hot chilli tomato sauce matrices. The results were compared with those obtained on a LC-triple quadrupole-MS system. Confirmation of Sudan I present in hot chill tomato sauce samples was obtained by accurate mass measurements. In real samples trueness of exact mass measurements was estimated to be 1.6 and 4.4 ppm when calculated for hot chilli tomato sauce and hot chilli tomato with cheese sauce samples, respectively; precision was calculated around 9.5 ppm.

Azo Compounds↗

PubMed2004

Development and in-house validation of a liquid chromatography-electrospray-tandem mass spectrometry method for the simultaneous determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli products.

An accurate method based on the use of reversed-phase (RP) liquid chromatography-tandem mass spectrometry interfaced with electrospray (LC-ESI-MS/MS) was devised for the determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli food samples. A simple sample treatment procedure entailing the use of an extraction step with acetone without clean-up was developed. A C18 column with an aqueous formic acid/methanol mixture as the mobile phase was used under isocratic conditions. Mass spectral acquisition was done in positive ion mode by applying selected reaction monitoring of three fragmentation transitions per compound to provide a high degree of selectivity. The method was in-house validated in terms of detection limits (LOD), quantitation limits (LOQ), linearity, sensitivity, accuracy, recovery, and selectivity on two kinds of hot chilli sauces. Good results in the low ng/g level were obtained for LOD and LOQ of all analytes in matrices. Under both intra-day repeatability (R.S.D. between 1 and 13%) and intermediate precision (about 5-15% R.S.D. for both chilli sauce matrices) conditions, precision proved to be typical of determinations based on electrospray LC-MS and acceptable for routine monitoring purposes. Extraction recoveries for all four azo-dyes in chilli tomato sauce ranged from 92 to 103% at a spiking level of 5 microg/kg, whereas values between 72 and 97% were calculated in chilli tomato and cheese sauce at the same concentration level. The applicability of the method to the determination of Sudan azo-dyes in hot chilli products was demonstrated.

Azo Compounds↗

PubMed2003

Analysis of protein profiles of genetically modified potato tubers by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Traceability of genetically modified (GM) foods demands the development of appropriate reliable techniques in order to identify and quantify peptide or nucleic acid residues in GM plants and food products through the food chain. In this study the applicability of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) was demonstrated for the characterization of proteins of transformed and untransformed potato (Solanum Tuberosum L.) tubers. In GM tubers the expression level of the G1-1 gene, which regulates transition from dormancy to sprouting tubers, was inhibited by antisense technology. The analysis of antisense transformed lines showed that several of them exhibited a significant delay in sprouting relative to the control lines, in accordance with a decrease in the transcript level. Preliminary attempts to compare the protein patterns obtained from transformed and control lines using traditional electrophoresis were not able to reveal differences in the low-kDa range. Instead, MALDI-TOFMS applied to total peptide extract without any purification was able to distinguish spectral patterns of transformed and untransformed lines. In particular, several characteristic peaks from m/z 4373 to 4932 were detected only in the mass spectra of GM tuber samples.

Food, Genetically Modified↗

PubMed2002

Rapid method for determination of chlormequat residues in tomato products by ion-exchange liquid chromatography/electrospray tandem mass spectrometry.

A rapid method has been devised for the direct determination of chlormequat in tomato samples. No clean-up is required, and analysis uses ion-exchange liquid chromatography/tandem mass spectrometry interfaced with electrospray ionization (LC/ESI-MS/MS). A cation-exchange column was used with an aqueous ammonium acetate/acetonitrile mixture as the mobile phase under isocratic conditions. The method was validated in terms of detection limits (LOD), quantitation limits (LOQ), linearity, recovery, precision and accuracy. Good results in the low micro g kg(-1) level were obtained for the LOD and LOQ of chlormequat in tomato samples. Comparison of solvent and matrix-matched calibration curves demonstrated the absence of significant matrix effects and the feasibility of using external calibration. Linearity was established over two orders of magnitude by performing homoscedasticity and Mandel fitting statistical tests. The absence of both constant and proportional systematic errors was verified by evaluating the recovery function, demonstrating good method accuracy. Excellent precision in terms of intra-day repeatability was calculated (RSD% <3.4). Extraction recoveries from tomato products were calculated, by using a labelled internal standard (d(4)-chlormequat), to be in the 93 +/- 5-99 +/- 7% range. The applicability of the method to the determination of chlormequat residues in tomato products was demonstrated.

Calibration↗

PubMed2001

Liquid chromatography-UV determination and liquid chromatography-atmospheric pressure chemical ionization mass spectrometric characterization of sitosterol and stigmasterol in soybean oil.

A narrow-bore HPLC-UV method was developed for the analysis of two of the more abundant naturally occurring phytosterols in vegetable oils: sitosterol and stigmasterol. The method enabled detection of the compounds at a concentration of 0.42 microg/ml and quantitation at concentrations of 0.52 and 0.54 microg/ml for sitosterol and stigmasterol, respectively. An excellent linearity was determined over two orders of concentration magnitude (r2 0.999-1.000) and verified by applying the Mandel fitting test (p>0.099) and the lack-of-fit test (p>0.057) performed at the 95% confidence level. A good intra-day precision ranging from 0.15 to 1.16% was calculated at two concentration levels (2 and 100 microg/ml). The inter-day reproducibility was verified on 3 different days by performing an homoscedasticity test and analysis of variance. A solid-phase extraction method was developed on silica cartridges for the isolation of phytosterols from soybean oil providing recovery values of 101+/-9 and 106+/-7% for sitosterol and stigmasterol, respectively. Good accuracy of the method was statistically demonstrated since no matrix effect was found for both the analytes. The developed method was applied to the quantitative assay of phytosterols in a soybean oil sample (61+/-5 mg/100 g of stigmasterol and 118+/-4 mg/100 g sitosterol). The HPLC-atmospheric pressure chemical ionization MS technique enabled the identification of stigmasterol, sitosterol and campesterol in the oil sample.

Atmospheric Pressure↗

PubMed2001

Non-covalent binding of endogenous ligands to recombinant cellular retinol-binding proteins studied by mass spectrometric techniques.

Recent developments in mass spectrometry have demonstrated the capability of this technique to transfer non-covalent protein complexes, involving low and high molecular weight ligands, from a condensed state to the gas phase. In this work, electrospray mass spectrometry with a quadrupole analyzer (ES-MS) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) were used to analyze the non-covalent association between recombinant rat cellular retinol-binding protein type-I (CRBP) with its specific ligand, all-trans retinol (vitamin A), and with fatty acids. Under denaturing conditions, MALDI-TOFMS and ES-MS techniques allowed determination of the molecular weight of apo-CRBP with good accuracy (<0.01%) and to identify a protein fraction ( approximately 20%) retaining the initial methionine. By adding saturating amounts of vitamin A, ES-MS studies on the protein in the holo-form under native conditions allowed detection of retinol bound within the cavity together with water molecules, as expected from its crystal structure. ES mass spectra of CRBP in the native state were also recorded under non-denaturing conditions, with the aim to study non-covalent interactions between CRBP and non-specific ligands such as fatty acids, bound to the protein as a result of expression in various strains of E. coli grown in different media. Since ES mass spectra do not elucidate which species interact with the protein, in order to investigate the ligands possibly retained in the active site of recombinant CRBP, liquid chromatography/ES-tandem mass spectrometry was used. In particular, this technique was applied to identify and quantify fatty acids bound to CRBP. Quantitative data indicated the presence of a few fatty acids at a total concentration lower than 2% of that of the protein. Similar findings were observed for the homolog rat cellular retinol-binding protein type-II, demonstrating the high degree of purity and homogeneity of apo-CRBP preparations derived from gene expression.

Bacterial Proteins↗

PubMed2000

Spectrophotometric and coulometric detection in the high-performance liquid chromatography of flavonoids and optimization of sample treatment for the determination of quercetin in orange juice.

The capabilities of spectrophotometric and electrochemical detection techniques were investigated for the high-performance liquid chromatographic determination of flavonoids. Liquid chromatographic analyses were performed on eleven compounds belonging to three different classes of flavonoids: flavanone glycosides, flavone and flavonol aglycones. Separation of all compounds examined was carried out under reversed-phase conditions on a C18 narrow-bore column for UV detection, whereas for electrochemical detection, a C18 standard-bore column was used. UV analyses were carried out at 280 nm for flavanones and at 265 nm for flavones and flavonols, whereas controlled-potential coulometric measurements were performed using a porous graphite electrode. Analytical performances of the methods were compared in terms of linearity, limits of detection (LODs) and precision. Linearity over two orders of magnitude and LODs at low-ppm levels (0.06-1 mg/l) were demonstrated for all techniques considered. Instrumental precision in terms of relative standard deviation was found to be between 0 and 5% for the liquid chromatography (LC)-UV system and between 0.6 and 10% for the LC-electrochemical detection (ED) system. The methods developed were applied to the analysis of flavanones and flavonols in a real sample, such as an extract of orange juice. Even though quercetin glycoside is mostly present in orange juice as rutin, other different glycosides of this flavonol could be present; on this basis, the hydrolysis of all glycosides to aglycone allows one to obtain more accurate data on the flavonol concentration in orange juice. To avoid sample degradation and to increase extraction efficiency, quercetin hydrolysis was optimized using a central composite design to investigate the effects of acid concentration and hydrolysis time on extraction recovery.

Beverages↗

PubMed1999

Validation of a liquid chromatography ionspray mass spectrometry method for the analysis of flavanones, flavones and flavonols.

The application of liquid chromatography/mass spectrometry (LC/MS) with a TurboIonspray (TIS) interface was investigated as a new method for the analysis of flavonoids. Eleven compounds belonging to three different classes of flavonoids were studied: eriocitrin, neoeriocitrin, naringin, narirutin, hesperidin, neohesperidin (flavanone glycosides), quercetin, kaempferol, galangin (flavonol aglycones), chrysin, apigenin (flavone aglycones). Chromatographic separations were performed under reversed-phase conditions using a C18 narrow-bore LC column; a mixture of an aqueous solution of formic acid (pH 2.4) and acetonitrile was used as the mobile phase. Isocratic elution was operated in the case of flavanones, whereas gradient elution was used for the simultaneous separation of flavones and flavonols. The adaptability of TIS to high flow applications allows the use of LC eluent flow rates at 200 µL/min without post-column splitting. Qualitative analysis was performed in negative-ion (NI) full-scan mode, whereas response linearity, detection limits and precision of the method were studied under NI selected ion monitoring (SIM) conditions. Characterization of isomers differing in the glycosylation was found to be possible on the basis of different mass spectra. Detection limits in the low-ng range (0.08-0.4 ng) were found, about twenty-fold lower than those reported previously. The method was applied to identify and determine the content of flavonoids in an orange juice sample. Copyright 1999 John Wiley & Sons, Ltd.

Journal Article↗

PubMed1999

Liquid chromatography-electrospray mass spectrometry of beta-carotene and xanthophylls. Validation of the analytical method.

The investigation of beta-carotene and the xanthophylls beta-cryptoxanthin, lutein, zeaxanthin, canthaxanthin and astaxanthin using reversed-phase liquid chromatography-electrospray mass spectrometry interfaced with TurboIonspray (LC-TurboISP-MS) is described. Two narrow-bore C18 columns connected in series and an isocratic solvent system containing acetonitrile-methanol (0.1 M ammonium acetate)-dichloromethane at a flow-rate of 300 microl/min (without splitting) were used. Operating in the positive-ion mode over m/z 500-650, the effects on the formation of the molecular ion species or adduct ions and the MS detector response were investigated for carotenoids, varying the orifice plate voltage, the ring voltage and the ISP voltage. Both conventional ISP and TurboISP were performed; using the TurboISP-MS system, ionization efficiency increased with respect to ISP-MS, particularly at the highest temperature (500 degrees C). Good results were particularly obtained for beta-carotene, which was detectable at the low ng level, without the use of solution-phase oxidants. Using LC columns and acquiring in single-ion monitoring mode, detection limits were estimated to be in the 0.1-1 ng range; dynamic range was established between one- and two-orders of magnitude. Better sensitivity under positive-ion than negative-ion conditions was demonstrated.

Chromatography, High Pressure Liquid↗

PubMedDate not supplied

Development and validation of a method using on-line solid-phase extraction and liquid chromatography with ultraviolet detection for the determination of bisphenol A, octylphenol, and nonylphenol in groundwater.

Alkylphenols are nonbiodegradable metabolites arising from various pathways of aerobic and anaerobic degradation of the nonionic surfactants alkylphenol ethoxylates. A method based on the use of on-line solid-phase extraction (SPE) and liquid chromatography (LC) with UV detection was developed to determine bisphenol A, octylphenol, and nonylphenol at trace levels in water. The on-line coupled SPE procedure automatically enables an approximately 300-fold preconcentration of analytes, which can be further enhanced by an increase in applied sample volume. By using C18 cartridges, recoveries of >90% were obtained for all the analytes. A validation procedure was carried out with a groundwater sample to ensure the quality of the results; performance criteria such as detection limits (LODs), quantitation limits (LOQs), linearity, and precision were evaluated. LODs and LOQs in the range of 0.17-0.36 and 0.35-1.88 microg/L, respectively, were found; for all the analytes, linearity was established over 2 orders of magnitude (r2 >0.997, n = 54). The intraday repeatability values expressed as relative standard deviations were <5.3%; a nested analysis of variance was performed to verify the influence of 3 different factors, i.e., different days, extraction procedure, and LC replicate injection, on data precision.

Benzhydryl Compounds↗