Development of a fast and simple gas chromatographic protocol based on the combined use of alkyl chloroformate and solid phase microextraction for the assay of polyamines in human urine.

Polyamines are aliphatic amines with low molecular weight that are widely recognized as one of the most important cancer biomarkers for early diagnosis and treatment. The goal of the work herein presented is the development of a rapid and simple method for the quantification of free polyamines (i.e., putrescine, cadaverine, spermidine, spermine) and N-monoacetylated polyamines (i.e., N1-Acetylspermidine, N8-Acetylspermidine, and N1-Acetylspermine) in human urine. A preliminary derivatization with propyl chloroformate combined with the use of solid phase microextraction (SPME) allowed for an easy and automatable protocol involving minimal sample handling and no consumption of organic solvents. The affinity of the analytes toward five commercial SPME coatings was evaluated in univariate mode, and the best result in terms of analyte extraction was achieved using the divinylbenzene/carboxen/polydimethylsiloxane fiber. The variables affecting the performance of SPME analysis were optimized by the multivariate approach of experimental design and, in particular, using a central composite design (CCD). The optimal working conditions in terms of response values are the following: extraction temperature 40 °C, extraction time of 15 min and no addition of NaCl. Analyses were carried out by gas chromatography-triple quadrupole mass spectrometry (GC-QqQ-MS) in selected reaction monitoring (SRM) acquisition mode. The developed method was validated according to the guidelines issued by the Food and Drug Administration (FDA). The satisfactory performances reached in terms of linearity, sensitivity (LOQs between 0.01 and 0.1 µg/mL), matrix effect (68-121%), accuracy, and precision (inter-day values between -24% and +16% and in the range 3.3-28.4%, respectively) make the proposed protocol suitable to be adopted for quantification of these important biomarkers in urine samples.

A solid-phase microextraction-gas chromatographic approach combined with triple quadrupole mass spectrometry for the assay of carbamate pesticides in water samples.

A simple and sensitive method was developed for the quantification of five carbamate pesticides in water samples using solid phase microextraction (SPME) combined with gas chromatography-triple quadrupole mass spectrometry (GC-QqQ-MS). The performance of five SPME fibers was tested in univariate mode whereas the other variables affecting the efficiency of SPME analysis were optimized by the multivariate approach of design of experiment (DoE) and, in particular, a central composite design (CCD) was applied. The optimum working conditions in terms of response values were achieved by performing analysis with polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber in immersion mode for 45min at room temperature with addition of NaCl (10%). The multivariate chemometric approach was also used to explore the chromatographic behavior of the carbamates and to evaluate the importance of each variable investigated. An overall appraisement of results shows that the factor which gave a statistically significant effect on the response was only the injection temperature. Identification and quantification of carbamates was performed by using a gas chromatography-triple quadrupole mass spectrometry (GC-QqQ-MS) system in multiple reaction monitoring (MRM) acquisition. Since the choice of internal standard represented a crucial step in the development of method to achieve good reproducibility and robustness for the entire analytical protocol, three compounds (2,3,5-trimethacarb, 4-bromo-3,5-dimethylphenyl-n-methylcarbamate (BDMC) and carbaryl-d7) were evaluated as internal standards. Both precision and accuracy of the proposed protocol tested at concentration of 0.08, 5 and 3 µg l⁻¹ offered values ranging from 70.8% and 115.7% (except for carbaryl at 3 µg l⁻¹) and from 1.0% and 9.0% for accuracy and precision, respectively. Moreover, LOD and LOQ values ranging from 0.04 to 1.7 ng l⁻¹ and from 0.64 to 2.9 ng l⁻¹, respectively, can be considered very satisfactory.

Sarcosine as a marker in prostate cancer progression: a rapid and simple method for its quantification in human urine by solid-phase microextraction-gas chromatography-triple quadrupole mass spectrometry.

Sarcosine is an amino acid derivative of N-methylglycine and is involved in the amino acid metabolism and methylation processes that are enriched during prostate cancer progression. It could also serve as a new target to be measured during therapeutic interventions and help in the identification of aggressive tumors for radical treatment. In this study, we present a new urine test that can help early diagnosis of prostate cancer. The method for the quantification of sarcosine in urine consists of a solid-phase microextraction (SPME) step followed by gas chromatography-triple quadrupole mass spectrometry analysis. We used a preliminary derivatization step with ethyl chloroformate/ethanol and the corresponding ester was then extracted by SPME in immersion mode. Several fibers were evaluated and the optimization of the parameters affecting the SPME process was carried out using an experimental design. The optimal values were 20 min extraction time, 10% NaCl, and 270°C using a divinylbenzene/Carboxen/polydimethylsiloxane fiber. The triple quadrupole analyzer acquired data in selected reaction monitoring mode, allowing us to obtain reconstructed chromatograms with well-defined chromatographic peaks. The accuracy and precision of this method were evaluated at concentrations of 70, 250, and 800 ng/ml and were found to be acceptable. Very satisfactory values (0.10 and 0.16 ng/ml, respectively) were also achieved for the limit of detection and the limit of quantification. The proposed protocol represents a rapid, simple, selective, and sensitive tool to quantify sarcosine in urine samples for prostate cancer diagnosis and for a screening test.

Tandem mass spectrometry in food safety assessment: the determination of phthalates in olive oil.

A gas chromatography-tandem mass spectrometry (GC-MS/MS) method for the detection of six phthalates in olive oil was developed. A gel permeation chromatography (GPC) clean-up step with cyclohexane:dichoromethane 7:3 as mobile phase was employed to remove the high-molecular mass species present in oil. Two ionization methodologies, i.e. electron (EI) and isobutane-chemical ionization (CI), were compared, in MS/MS mode, to achieve better analytical performances. An overall evaluation of all analytical parameters shows that the EI-MS/MS approach provides satisfactory results and is to be preferred to CI-MS/MS, at least in the case of the examined analytes. The observed accuracies, ranging from 71.7% to 112.2%, and the RSD values less than 9.7%, confirm the effectiveness of the proposed method in the assay of phthalate content in such a complex matrix as olive oil. The proposed protocol for the identification and assay of phthalates in olive oil might be of interest for the implementation of the QS (quality assurance scheme) for residue monitoring in food safety assessment.

A metabolomic approach to the evaluation of the origin of extra virgin olive oil: a convenient statistical treatment of mass spectrometric analytical data.

The selection of suitable markers from the secondary metabolism of lipoxygenase, in experimental olive oils produced from drupes harvested in different areas of the Italian Calabria region and of Tunisia, allows an easy discrimination between each cluster of samples. The origin of the foodstuff can be ascertained even when the distances between the production zones are very close to each other as in Calabria. Olive oils produced from irrigated and nonirrigated farms in Tunisia were also clearly distinguishable. The markers were detected by chemical ionization mass spectrometry with an ion trap gas chromatography-mass spectrometry apparatus. The quantitative data of Calabrian olive oil samples were subjected to linear discriminant analysis, whereas the Tunisian data were treated by means of other two statistical tools, i.e., the Kruskal-Wallis test and the Wald-Wolfowitz test.