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Incontri di Scienza delle

Separazioni
Il contributo della scienza delle
separazioni alle
problematiche alimentari ed ambientali
28-29 Novembre 2013
Aula Magna Vittorio Ricevuto Centro
Polifunzionale ex Facolt di Scienze
Viale Stagno dAlcontres, Universit di
Messina

Book of Abstracts

PL.01
CROMATOGRAFIA BIDIMENSIONALE COMPREHENSIVE PER LANALISI DI LIPIDI
Giovanni Dugo1,2, Paola Dugo1,3
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - Universit di Messina,

viale Annunziata, 98168 Messina, Italy

2 Chromaleont s.r.l. A start-up of the University of Messina - c/o Dipartimento di Scienze del Farmaco

e dei Prodotti per la Salute, viale Annunziata, 98168 Messina, Italy

3

Centro Integrato di Ricerca (CIR) - Universit Campus Bio-Medico, Via Alvaro del Portillo, 00128
Roma, Italy

Lo sviluppo di metodi analitici per la determinazione accurata della composizione dei lipidi
alimentari rappresenta un ramo della ricerca accademica, industriale e clinica di grande
rilevanza. Da punto di vista nutrizionale sempre stata data grande importanza allapporto
nutritivo dei lipidi nella dieta umana, considerato il ruolo fondamentale che questa classe di
sostanze ricopre sulla nostra salute. Non va inoltre trascurato il fatto che grassi e oli alimentari
sono fonte di numerose sostanze nutritive essenziali (vitamine, AGE), contribuiscono a
migliorare le propriet strutturali e organolettiche degli alimenti.
Dal punto di vista analitico la cromatografia liquida (LC) e gassosa (GC) vengono applicate
allanalisi di matrici lipidiche con ottimi risultati, spesso ricorrendo alla rivelazione con
spettrometria di massa per unaccurata delucidazione strutturale. A tal proposito lo sviluppo
delle tecniche cromatografiche bidimensionali comprehensive (GCGC, LCLC) ha contribuito
ad un ulteriore evoluzione nel campo dellanalisi dei lipidi alimentari, particolarmente complessi,
facilitando linterpretazione strutturale mediante MS, grazie alla maggiore capacit separativa
e quindi allottenimento di picchi cromatografici completamente risolti.
In questa lectio verranno illustrati i principi alla base della cromatografia bidimensionale
comprehensive sia GCxGC che LCxLC. Verranno inoltre dettagliatamente descritte le
interfaccie pi importanti con riferimento alle diverse applicazioni allanalisi dei lipidi. Tra le
matrici lipidiche studiate attraverso la tecnica GCxGC sono qui riportati esempi di separazione
e caratterizzazione di acidi grassi in lipidi di origine sia animale che vegetale, alla frazione
insaponificabile di oli e di grassi alimentari e le applicazioni LCxLC per la delucidazione del
profilo di acilgliceroli e fosfolipidi in diverse matrici lipidiche.

L.01
DEVELOPMENT OF AN ANALYTICAL PLATFORM FOR THE PROTEOMIC
CHARACTERIZATION OF OLIVE (OLEA EUROPAEA) PULP
Anna Laura Capriotti, Valentina Colapicchioni, Patrizia Foglia, Serena Stampachiacchiere
Dipartimento di Chimica - Sapienza Universit di Roma, Piazzale Aldo Moro 5, 185 Rome, Italy

The nutritional and cancer-protective properties of the oil extracted mechanically from the ripe
fruits of Olea europaea trees are attracting constantly more attention worldwide. Biological
research has focused in the past on model organisms and most of the functional genomics
studies in the field of plant sciences are still performed on model species or species that are
characterized to a great extent. However, numerous non-model plants are essential as food,
feed, or energy resource. The preparation of high-quality protein samples from non-model plant
tissues for proteomic analysis poses many challenging problems. To begin with, because of the
complexity of the matrices examined, it is difficult to obtain high quality protein extracts, and
secondly, due to the lack of sequenced genomes, the database information available is limited.
The main aim of this work was to establish an effective analytical platform for the isolation of
olive pulp proteins to provide a method for the identification of as many proteins present in these
samples as possible. More specifically, we proposed two different extraction protocols for gelfree approaches, using physiological (CHAPS buffer) and denaturing (SDS buffer) conditions
and two precipitation strategies were tested for each one. The extracted proteins were trypsin
digested, and the resulting peptide mixture analyzed by reversed-phase (RP) nanoHPLCtandem MS (MS/MS). Different protein databases, in particular Viridiplantae entries for
SwissProt and NCBI, and TAIR10, were used. The choice of different databases for MS/MS
spectra search and peptide and protein identification was made in order to provide a platform
which would allow an increase in the number of protein identifications, also by homology, thus
providing a more complete description of the proteome of non-model plants.
For recalcitrant tissues it is not possible to provide a universal and simple sample preparation
procedure. The olive pulp results showed that the SDS and CHAPS extractions, when associated
to two different precipitations, permitted the extraction of different proteins on the basis of
different solubility properties. The use of selective extraction protocols, advanced MS
techniques in combination with three databases allowed us to identify more than 1000 proteins
for the olive pulp.

L.02
THE USE OF ION MOBILITY ENABLED MASS SPECTROMETRY FOR THE
DEVELOPMENT AND CHARACTERISATION OF ROBUST ANALYTICAL METHODS
FOR TRACE RESIDUE QUANTITATION IN FOOD OF ANIMAL ORIGIN
Sergio Gallo
Waters S.p.A., Viale dell'Innovazione, 3, 20126 Milano, Italy

La mobilita' ionica in spettrometria di massa rende possibile per la prima volta la separazioe di
isobari, o in ogni caso di specie ioniche aventi lo stesso valore di m/z. Non solo e' possibile la
loro separazione, ma anche la loro selezione onde ricavarne spettri di frammentazione "ripuliti".
Viene qui mostrato quale esempio, l'analisi di derivati del fluorochinolone, in particolare
enrofloxacina e ciprofloxacina, rispettivamente in estratti di pesce e nel miele.
In questo caso cio' che varia e' il sito di protonazione, che da luogo a differenti frammenti. Si
mostrera' come e' possibile distinguere fra le varie specie isobare (che qui chiameremo
"protomeri") allo scopo di ottenere spettri di frammentazione semplificati e di facile
interpretazione.
Il metodo per LC-MS cos testato, potra' poi essere passato in routine con l'utilizzo di un semplice
spettrometro di massa a triplo quadrupolo.

L.03
TARGETED METABOLOMICS METHODS FOR THE SEPARATION AND
QUANTIFICATION OF MULTIPLE CLASSES OF PHENOLICS: APPLICATION IN FOOD
AND NUTRITIONAL STUDIES
Mattia Gasperotti, Domenico Masuero, Fulvio Mattivi, Urska Vrhovsek
Food Quality and Nutrition Department - Research and Innovation Centre - Fondazione E. Mach, via
E. Mach 2, 38010 San Michele all'Adige, Italy

Compelling evidence of the health benefits of phenolic compounds and their impact on food
quality have stimulated the development of analytical methods for the separation and
quantification of these compounds in different matrices in recent years. The complexity and
remarkable diversity of phenolics has challenged the analytical performances of separation and
detection methods in terms of resolving power, selectivity and sensitivity for the identification
and quantification of these compounds in different matrices. Targeted metabolomics is a strategy
based on the use of predefined metabolite-specific signals, such as MRM transitions, that can
be used to accurately determine the concentrations of a wide range of known metabolites.
A targeted metabolomics method [1] has been developed for the quantification >150 phenolics,
such as benzoates, phenylpropanoids, coumarins, stilbenes, dihydrochalcones, and flavonoids,
using an UPLC/MS/MS system. Reverse-phase chromatography was optimised to achieve
separation of the compounds over 15 min, reducing possible ion suppression effects and
resolving many isomeric compounds and MRM transitions were selected for accurate
quantification.
Recently a new targeted metabolomics method [2] was developed for the quantification of 23
metabolites/catabolites related to the consumption of polyphenols and associated to the
microbial digestion made by the gut microflora. The present new method was developed the
analysis of complex matrix with SPE purification before the UPLC separation. Then it was
validated and applied for human/animal biofluids or tissues.
The validated methods were applied to the analysis of fruits and wine polyphenols as well as
in nutritional studies providing a valuable tool for food quality evaluation and nutritional relevant
bioactive compounds profiling. The short duration of the analysis and the straightforward sample
preparation make the methodology suitable for high-throughput screening studies for analysis
of food as well as its application in nutritional studies to biofluids.
References:
[1] Vrhovsek U. et al., J. Agric. Food Chem. 60, 8831, (2012).
[2] Gasperotti M. et al., unpublished data

L.04
PREPARATION AND CHARACTERIZATION OF MONOLITHIC COLUMNS FOR
LIQUID CAPILLARY CHROMATOGRAPHY AND THEIR APPLICATIONS IN
PROTEOME ANALYSIS
Giuseppe Pierri, Patrizia Simone, Dorina Kotoni, Alessia Ciogli, Claudio Villani, Francesco
Gasparrini
Dipartimento di Chimica e Tecnologie del Farmaco - Sapienza Universita' di Roma, P.le A. Moro 5,
185 Roma, Italy

Lab-made capillary organic monolithic columns were prepared by polymerization of a

mixture containing monomers, cross-linkers and porogenic solvents. A mixture of monomers
consisting of 78% wt of lauryl methacrilate (LMA) and 22% wt of 1,6-hexanediol dimethacrylate
(HDDMA) was dissolved in porogenic solvents (tert-butyl alcohol and 1,4-butanediol) to obtain
the polymerization mixture. We have developed an innovative synthetic process that is based
on -radiation induced polymerization [1] inside fused silica capillaries (200 m or 250 m I.
D.) using a new tentacle type silanization agent, affording a multisite reactive surface. The
porous polymer monoliths can be obtained also by thermal polymerization using AIBN
(azobisisobutyronitrile) as radical initiator at temperature of 50 C.
The efficiency and hydrophobicity of two monolithic columns were systematically
evaluated using a set of small probe molecules (benzaldehyde, nitrobenzene, toluene,
ethylbenzene, butylbenzene) under Reversed-Phase conditions using CH3CN/H2O, 60/40 (v/
v) as mobile phase at different flow rates and at room temperature.
The relative Van Deemter curves showed that a 30-cm-long monolithic column obtained by ray can be produce ~100.000 theoretical plates per meter; (Hmin= 10.5 m at optimum
linear velocity opt= 0.50 mm/s; solute: benzaldehyde).
The performance of the monolithic column was evaluated by separation of standard mixture
of proteins (lysozyme, -lactalbumin, -lactoglobulin, carbonic anhydrase) and peak capacity
of ~800, with a 55-cm-long column and a gradient time of 240 min, was obtained.
The high efficiency and low back-pressure of these monoliths allowed to use them in the
analysis and separation of intact proteins, peptides and in high resolution techniques using
columns up to 1 meter long.
An integrated capillary high performance liquid chromatography and high-resolution mass
spectrometry (cap-LC-HRMS) approach [2] was developed, exploiting the permeability
and biocompatibility of organic monolith. Thanks to the higher observed resolution, the
monolithic capillary column was chosen for degradation studies of casein fractions extracted
from fresh and expired milk.

References:
[1] Angelini G., Ursini O., Gasparrini F., Villani C., PCT Int. Appl., WO 2008081496 A1
20080710 (2008).
[2] Pierri G., Kotoni D., Simone P., Villani C., Pepe G., Campiglia P., Dugo P., Gasparrini F.,
Journal of Chromatogr. A, 1313, 259, (2013).

L.05
SCREENING OF PESTICIDES IN WATER USING SPE ON-LINE
Stefano Maria Lucini, Monica Lusardi
Shimadzu Italia, Via G.B. Cassinis 7, 20139 Milano, Italy

Introduction
Pesticides are widely used in agricultural products to protect crops from insect infestation during
cultivation and product transport. The abuse of pesticides can cause important environmental
pollution in different areas, especially at level of lakes, rivers and aquifers. For this reason one
of the most important and attractive application is the possibility to analyze pesticides in water
without pretreatment of the sample. Shimadzu in cooperation with Arpa Asti (Italy) developed
a system for injection of water sample using a triple quadrupole system using a Solid-Phase
Exctraction (SPE) on line. We injected water samples in a LCMS-8030 Shimadzu triple
quadrupole instrument with Nexera UHPLC instrument in order to identify a mixture of 27
substances.
Methods
A standard mixture of pesticides was used for optimization of all MRM parameters. The
chromatographic separation was performed by a binary gradient of methanol and water with
0.1% of formic acid. 1mL of sample were directly injected using an SPE on-line column (YMC
Europe GMBH Guard cartridge C18 (20 x 2.1 mm)) for concentration. Shimadzu LCMS-8030
was used in MRM mode for detection of the researched compound. For construction of
calibration curve sample of water spiked with standard were used
Preliminary data
The system developed allows a simultaneous detection at the same time of a different samples
volumes by mean of SPE on-line for pre-concentration of the sample (injection of 2mL) and
Ultra-Fast analysis (50uL injection volume). The switch between the two loop is automatically
and controlled by software LabSolution. Before analysis we checked the correct volumes of
aspirations and injections of the autosampler. We developed calibration curves for 26
compounds spiked in 1mL of water.

L.06
SVILUPPO DI UN SISTEMA NANOLC-EIMS PER LA COSTRUZIONE DI UN DATABASE
DI SPETTRI DI MASSA DI COMPOSTI NON VOLATILI
Francesca Rigano1, Marina Russo1, Danilo Sciarrone1, Peter Q. Tranchida1, Luigi Mondello1,2
1

Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - Universit degli Studi
di Messina, Viale Annunziata, 98168 Messina, Italy
2

Centro Integrato di Ricerca (C.I.R.) - Universit Campus Bio-Medico, Via lvaro del Portillo, 21,
00128 Roma, Italy

Negli ultimi anni laccoppiamento della spettrometria di massa ad impatto elettronico con la
cromatografia liquida (LC-EIMS) ha accentrato particolare interesse scientifico poich
implementa lidentificazione di composti non volatili grazie al maggior numero di informazioni
contenute in uno spettro EI rispetto a quelle ottenuti mediante tecniche a pressione atmosferica.
Tale peculiare accoppiamento stato reso possibile dai notevoli progressi compiuti nellambito
della cromatografia liquida miniaturizzata che hanno consentito di ridurre significativamente la
quantit di fase liquida che viene introdotta nella sorgente ionica del rivelatore [1].
Ci anche possibile grazie alle caratteristiche delle moderne pompe da vuoto dello
spettrometro di massa che, essendo in grado di lavorare con flussi fino a 10ml/min, sono in
grado di rimuovere efficientemente il solvente proveniente dalla nano-colonna LC, mantenendo
sotto alto vuoto il sistema di ionizzazione.
Il sistema NanoLC-MS permette di estendere il campo di applicabilit della spettrometria di
massa ad impatto elettronico a molecole a polarit medio-alta e/o termolabili, senza alcuna
derivatizzazione. In particolare, una notevole attenzione si sta dedicando a molecole bioattive,
pi precisamente alle classi dei polifenoli, tocoferoli e polimetossiflavoni.
Tale progetto prevede preliminarmente la creazione di una libreria di spettri di massa dei
composti di interesse attraverso liniezione dei loro standard disponibili in commercio o di
composti puri isolati da matrici reali. Questa seconda possibilit risultata molto conveniente
grazie allutilizzo di un sistema HPLC preparativo bidimensionale che ha recentemente
permesso lisolamento di componenti non volatili (flavonoidi e polimetossiflavoni) da matrici
alimentari con rese significative.
Gli spettri ottenuti possono essere utilizzati per scopi identificativi in campioni reali e
rappresentano un importante punto di partenza per implementare le librerie di spettri di massa
gi disponibili in commercio.

Bibliografia:
[1] Cappiello A. et al. Mass Spectrometry Reviews, 20, 88, (2001)

L.07
INFLUENZA DEI FATTORI GENETICI, AMBIENTALI E TECNOLOGICI SUL
CONTENUTO E LA DISTRIBUZIONE DI ACIDI FENOLICI NEL GRANO DURO E NEI
PRODOTTI DERIVATI
Daniela Martini1,2, Federica Taddei1, Isabella Nicoletti3, Maria Grazia D'Egidio1, Danilo Corradini3
1

Consiglio per la Ricerca e la Sperimentazione in Agricoltura (CRA) - Unit di Ricerca per la

Valorizzazione Qualitativa dei Cereali, Via Cassia 176, 00191 Roma, Italy
2

Universit Campus Bio-Medico, Via Alvaro del Portillo 21, 00128 Roma, Italy

3 Consiglio Nazionale delle Ricerche (CNR) - Istituto di Metodologie Chimiche, Via Salaria km 29.300,

00015 Monterotondo Stazione (Roma), Italy

I cereali rivestono interesse crescente nel panorama scientifico grazie al loro elevato contenuto
di composti bioattivi (fibre e antiossidanti); tra questi, di particolare interesse sono gli acidi
fenolici (PAs) presenti nei cereali in 3 diverse forme: libera, coniugata e legata.
Scopo di questo studio stato mettere a punto un metodo per lestrazione e lanalisi di PAs
liberi, coniugati e legati da applicare alla valutazione dei fattori genetici (cultivar) e ambientali
(luogo e anno di coltivazione) che influenzano il contenuto di acidi fenolici nel grano duro,
cereale di particolare importanza nei Paesi dellarea mediterranea. stato inoltre valutato
leffetto dei principali processi di trasformazione del grano duro (macinazione e pastificazione)
sul contenuto e sulla distribuzione delle 3 forme di PAs.
Lestrazione degli acidi fenolici stata effettuata con il metodo di Li et al. (2008) [1] modificato
ed ottimizzato per lanalisi di PAs in matrici con diverse caratteristiche chimico-fisiche (es. crusca
e semola) [2]. Questi composti sono stati poi separati mediante RP-HPLC, utilizzando una
colonna narrow-bore associata ad un rivelatore UV a serie di fotodiodi e ad uno spettrometro
di massa con sorgente di ioni per elettrovaporizzazione (ESI-MS). Il metodo stato validato in
termini di linearit delle rette di calibrazione, ripetibilit ed accuratezza, valutata mediante analisi
del recupero.
I risultati mostrano che il genotipo, lambiente e lanno di coltivazione influenzano il contenuto
di acidi fenolici, ma non la distribuzione percentuale degli stessi nelle 3 forme. La maggiore
fonte di variabilit per rappresentata dal processo di macinazione tradizionale che comporta
una drastica riduzione del contenuto di acidi fenolici, accumulati negli scarti di lavorazione
(crusca). Al contrario, la macinazione integrale consente di preservare questi composti nella
pasta, ottenendo prodotti finiti ricchi in composti bioattivi, di maggiore interesse nutrizionale
rispetto ai prodotti raffinati.
Bibliografia:
[1] Li L. et al., J Agric Food Chem 56, 9732, (2008)
[2] Nicoletti I. et al., J Agric Food Chem In press (2013)

L.08
CARATTERIZZAZIONE DEGLI ACIDI GRASSI DEI TESSUTI COME STRUMENTO
UTILE PER VALUTARE GLI EFFETTI BENEFICI DEGLI ALIMENTI
Riccardo Gagliardi1, Deborah Pacetti1, Paolo Lucci2, Natale G. Frega1
1

Dipartimento di Scienze Agrarie, Alimentari ed Ambientali - Universit Politecnica delle Marche UNIVPM, via brecce bianche, 10, 60131 Ancona, Italy
2

School of Sciences - Department of Nutrition and Biochemistry/Food Science and Technology Pontificia Universidad Javeriana, Cr. 7 No. 43-82 Edificio51-339A, no code Bogot, Colombia

Gli effetti benefici degli alimenti funzionali o delle terapie farmacologiche possono essere
efficacemente monitorati tramite lanalisi degli acidi grassi dei tessuti. Infatti, il profilo acidico
dei tessuti negli organismi viventi fortemente influenzato dalla dieta e dalla condizione
fisiologica dello stesso organismo.
Ad esempio, lacido arachidonico, come gli acidi grassi trans e idrossilati, sono considerati
markers per i danni ossidativi e degenerativi legati a varie patologie [1]. Nel nostro gruppo di
ricerca, al fine di valutare limpatto sulla salute di pazienti ipercolesterolemici di una dieta ricca
di un olio di palma ibrido non raffinato, contenente importanti quantit di micronutrienti con
effetti cardiovascolari benefici, stato studiato il profilo degli acidi grassi negli eritrociti. In
particolare stato determinato un innovativo fattore di rischio cardiovascolare, lindice 3 [2],
che rappresentato dalla somma delle percentuali relative degli acidi eicosapentaenoico
(C20:53) e docosaesaenoico (C22:63) negli eritrociti.
In ogni caso, la procedura analitica convenzionale per effettuare lanalisi degli acidi grassi dei
tessuti prevede lestrazione dei lipidi mediante metodo Folch e la successiva transmetilazione
dellestratto lipidico. Comunque, questa procedura pu presentare molti punti critici. Ad esempio,
time consuming, richiede notevoli quantit di solventi organici, non adatta quando i tessuti
animali sono disponibili in basse quantit.
Pertanto, eseguire la reazione di transmetilazione direttamente sul tessuto non trattato (senza
effettuare lestrazione lipidica), potrebbe risultare una soluzione alternativa per superare i punti
critici precedentemente illustrati.
A tal fine, il nostro gruppo ha ottenuto risultati soddisfacenti adattando la transmetilazione con
trifluoruro di boro descritta da Harris et al. [3] per lanalisi degli acidi grassi di diversi tessuti
animali.

Bibliografia:
[1] Niki E., Biofactors 34, 171, (2008)
[2] Harris W. et al., Preventive Medicine, 39, 212, (2004)
[3] Harris W. et al., The Journal of Nutrition, 142, 1297, (2012)

L.09
UPC2 - ULTRAPERFORMANCE CONVERGENCE CHROMATOGRAPHY - UNA
NUOVA SOLUZIONE PER INCREMENTARE LA SELETTIVIT NEI LABORATORI
CROMATOGRAFICI
Andrea Perissi
Application Specialist - Waters Italia, viale dell'Innovazione 3, 20126 Milano, Italy

UltraPerformance Convergence Chromatography - UPC2 rappresenta una nuova categoria

nella scienza separativa che unisce il potenziale delle separazioni SFC Waters con la provata
tecnologia Waters UPLC. Sfrutta come fase mobile principale la CO2 in fase supercritica le
cui propriet consentono di risolvere problemi legati a separazioni complesse. Lutilizzo di un
fluido supercritico in cromatografia permette di utilizzare una fase mobile dotata di un elevato
coefficiente di diffusione, caratteristica peculiare della cromatografia in fase gassosa, unita al
potere solvente caratteristico di un liquido. Storicamente, i sistemi SFC presentavano scarsa
robustezza, dovuta allincapacit del sistema di regolare finemente la densit della CO2, con
effetti negativi sulla riproducibilit nei tempi di ritenzione, insufficiente accuratezza del sistema
di iniezione ed elevate dispersioni del sistema e da bassa sensibilit. UPC2 un sistema
olistico basato sulla tecnologia UPLC, caratterizzato da basse dispersioni e colonne
cromatografiche con un particolato inferiore ai 2m che consente di ottenere dati analitici
affidabili, sensibili e robusti. Questa tecnica, compatibile con la maggioranza dei sistemi di
rivelazione, permette di incrementare la selettivit analitica utilizzando sia fasi stazionarie tipiche
della cromatografia liquida in fase inversa, mantenendo un profilo simile di eluizione, sia fasi
stazionarie polari, tipiche della fase normale, mitigando lutilizzo di solventi organici caratterizzati
da unelevata tossicit e che comportano ingenti costi di smaltimento. Inoltre, una fine
regolazione della pressione consente allanalista, di poter utilizzare la densit della fase mobile
come parametro aggiuntivo per migliorare la separazione di miscele complesse. Ogni composto
solubile in solvente organico candidato per lanalisi in Cromatografia Convergente. Questa
tecnica risulta quindi una valida alternativa alla cromatografia liquida in fase inversa e consente
di ottenere separazioni migliori per composti chirali, isomeri, stereoisomeri e diasteroisomeri.

10

L.10
QUANTITATIVE ANALYSIS OF AROMA AND TECHNOLOGICAL MARKERS IN
COMPLEX FOOD MATRICES THROUGH MULTIPLE HEADSPACE EXTRACTION
TECHNIQUE
Luca Nicolotti, Chiara Cordero, Erica Liberto, Barbara Sgorbini, Cecilia Cagliero, Alessandra
Griglione, Patrizia Rubiolo, Carlo Bicchi
Dipartimento di Scienza e Tecnologia del Farmaco - University of Turin, Via Pietro Giuria, 9, I-10125
Turin, Italy

Quantitative profiling is a fundamental topic when the aim is the correlation of food sample
compositional characteristics to its sensory and technological profile.
Food sample volatile fraction is usually very complex since it is made up of primary and
secondary metabolites and compounds generated by thermal treatment and/or enzymatic
activity. Moreover volatiles effectively contributing to the aroma of a food are rather limited and
are sometimes present at trace level (ng/Kg).
This presentation describes the outcomes of a study focused on the performance evaluation
of Multiple Headspace Extraction (MHE) [1], an external standard quantitation technique,
particularly suitable in analyzing complex, non homogeneous solid matrices, and the advantages
it provides when combined with Two Dimensional Comprehensive Gas Chromatography Mass
Spectrometry (GCXGC-MS).
Since a correct and univocal identification of compounds is a key point for a proper quantitation,
GCXGC-MS is indeed particularly suitable to resolve the complexity underpinning complex food
samples and to provide for a detailed profiling of volatiles even when the sample matrix to be
analyzed must be reduced to comply with quantitation requirements, as happens in Multiple
Head Space Extraction Technique.
Nineteen analytes among aroma and technological markers from both hazelnut and gianduja
paste samples were investigated and results compared with data obtained through a reference
quantitation technique: Standard Addition.
Despite the good consistency observed between the two approaches [2], MHE showed a smaller
dispersion among replicates and proved to be more effective to obtain information on the matrix
effect of different samples.
References:
[1] Kolb, B, L.S. Ettre. Static Headspace-Gas Chromatography, Theory and Practice. Wiley
VCH, New York, (1997)
[2] Nicolotti L. et al, Anal. Chimica Acta 798, 115, (2013)

11

L.11
REACTION PATH EVALUATION OF HDS CATALYTIC SYSTEM IN THE O-XYLENE
ISOMERIZATION VIA GAS-CHROMATOGRAPHIC ANALYSIS
Alessandra Palella1, Giuseppe Italiano2, Loredana Romano2, Francesco Frusteri1, Lorenzo
Spadaro1
1

DIITET - CNR-ITAE, Via S. Lucia Sopra Contesse, 98126 Messina, Italy

EcoRigen R&D Division - Eurecat S.a.S, c/o Raffineria Gela. c.da Piana del Signore, 93012 Gela
(CL), Italy

In recent years, environmental quality is one of main issues of international scientific community.
In particular, it is well know that the air pollution constitutes a serious hazard for human and
environmental health since it can bring to the formation of acid rains and can cause climate
changes. These environmental implication deeply regards also the automotive sector, prompting
the development of the so-called "ultra low sulfur clean fuels" which in turn has introduced new
challenges in hydrotreating. Commercially, HDS catalysts are based on transition metal
combinations of either CoMo or NiMo, supported on alumina or silica surface. These types of
catalysts are affected by several drawbacks that limit their performance. In particular, the acidic
properties of HDS catalyst can promote undesired side reactions such as cracking,
polymerization and isomerization, depending to reaction feedstock. In this respect, an analytical
study by gaschromatography analysis has been performed in the catalytic isomerization of oxylene, aiming to ascertain kinetic mechanism and reaction path of different commercial HDS
catalyst. The kinetic study were carried out by using a GC Agilent model 7890A, analyzing the
on-line composition of reactants and products by thermo conductivity (TC) and flame ionization
(FI) detectors. Permanent gases (i.e. H2, N2, CO, CO2, and methane) were detected by using
of Molecular Sieves and Poropak-Q packed columns connected in series through an
automatized multi-way valve systems. While condensable hydrocarbon were analyzed by using
of 30m timely customized poly-ethylene-glycol capillary column (Supelcowax 10, i.d. 0.53mm,
film thickness of 2mm). As proof of the good analytic performance achieved, Figure 1 displays
the analytic chromatogram of test performed with CoMo/A2O3 catalyst. As shown, the
analytical GC configuration allows to resolve the peaks of all isomers of xylene (i.e. o-, m- and
p-xylene) and also the hydrogenated stereoisomers (i.e. cis- and trans- 1,2-dimethylcycloexane,
1,3-dimethylcycloexane, 1,4-dimethylcycloexane). The results of tests prefigure the preliminary
fast isomerization of o-xylene to m-xylene, which in turn converts to p-xylene depending to acid
character of catalyst. Then, hydrogenation, cracking and polymerization reactions became more
relevant.

12

L.12
CHROMATOGRAPHIC SEPARATION WITH AMPEROMETRIC DETECTION FOR THE
DETERMINATION OF ENVIRONMENTAL POLLUTANTS
Luca Rivoira1, Rosa M. De Carlo1, Silvano Cavalli2, Corrado Sarzanini1, Maria Concetta
Bruzzoniti1
1

Dipartimento di Chimica - Universit degli Studi di Torino, Via Pietro Giuria 5, 10125 Torino, Italy

IRSA-CNR- Sezione di Brugherio, Via del Mulino 19, 20861 Brugherio, Italy

In this work an HPLC method coupled with amperometric detection was optimized for the
determination of substances of environmental concern, such as Bentazone and Atrazine
(belonging to the herbicides class) and Carbamazepine, Phenytoin and 5-(4'hydroxyphenyl)-5phenylhydantoine, HPPH (active principles of wide used medicines) [1-2].
After the optimization of the chromatographic parameters, a preliminary characterization of the
electrochemical behavior of the compounds was carried out by cyclic voltammetry on a glassy
carbon working electrode (Ag/AgCl reference electrode). This study was necessary to
subsequently optimize the best detection mode, constant potential (DC) and/or pulsed and
integrated amperometry (PAD/IPAD) and the related waveforms. Detection conditions were
optimized as a function of eluent pH, as well.
The results obtained showed that the best sensitivities can be achieved by PAD/IPAD detection
at pH 5 by the waveform represented in the figure. Detection limits were 12.5 g/L, 6.6 g/L
and 24.0 g/L for Bentazone, HPPH and Carbamazepine, respectively. The method is linear
over two orders of magnitude. Robustness was assessed through the evaluation of intra-day
(n=13) and inter-day tests (4 days, n=52).
The method was successfully applied for the analysis of a real sample (Po River, Turin, Italy).
References:
[1] Huber R., Otto S., Rev. Environ. Contam. Toxicol., 137, 111, (1994)
[2] Rogawski M.A., Loescher W., Nat. Med., 10, 685, (2004)

13

L.13
HIGH SENSITIVITY ANALYSIS OF NITROSAMINES USING GC-MS/MS
Elena Ciceri
Thermo Fisher Scientific, Strada Rivoltana, 20090 Rodano, Italy

Nitrosamines is the common term used for compounds of the class of N-nitrosodialkylamines.
A large variety of compounds are known and described with different alkyl moieties.
The simplest N-nitrosodialkylamine with two methyl groups is the N-nitrosodimethylamine
(NDMA).
Nitrosamines are in common highly toxic compounds with high cancerogenity for humans and
animals, in higher doses leading to severe liver damage with internal bleeding. Nitrosamines
in food are mainly produced from nitrites. Nitrites are added to food as preservatives in meat
and meat products preventing the Botulinus poisoning. Antioxidant food additives like vitamin
C can prevent the formation of nitrosamines from nitrites. Another source of nitrosamines is
described by the reaction of nitrogen
oxides with alkaloids as it is reported from the drying process of the germinated malt in beer
production. As nitrosamine levels in malt and beer have been significantly reduced in the brewing
process, high analytical performance is required. In addition to the regular control of other food
products for daily consumption, malt in beer is also monitored for low levels of nitrosamines.
This presentation describes a turn-key GC-MS/MS method for routine detection and quantitation
of food borne nitrosamine compounds. The food matrix in this work has been different malt beer
products and as a final food product the commercial beer. Special focus in the method
development has been made to provide the required high sensitivity for the detection of the
nitrosamine compounds for a fast, easy to implement routine method.

14

L.14
FOOD ANALYSIS BY MEANS OF NANO-LIQUID CHROMATOGRAPHY
Anna Rocco1, Zeineb Aturki1, Laura Dugo2, Chiara Fanali2
1 Institute of Chemical Methodologies - CNR, VIA SALARIA

KM 29,300, 15 MONTEROTONDO, Italy

Centro Integrato di Ricerca (C.I.R.) - Campus Bio-Medico University, Via Alvaro del Portillo 21,
00128 Rome, Italy

The determination of food composition in order to verify its authenticity, beneficial or hazardous
effects on human health, is a very important task of analytical chemistry. Gas chromatography
(GC) and high performance liquid chromatography (HPLC) are the analytical methods so far
used for these aims. However, the use of miniaturized techniques, such as capillary zone
electrophoresis (CZE), capillary electrochromatography (CEC), capillary- and nano-LC, are
increasingly being utilized in the analytical field.
Indeed, they offer several advantages over classical techniques, for instance short analysis
time, easily coupling with MS and reduced consumption of both mobile and stationary phase,
with a consequent reduced waste and environmental pollution.
In this presentation, the applicability of nano-LC in food analysis, especially concerning the
investigation of nutraceutical compounds, is showed reporting methods, e.g., for the
determination of phytosterols in extra-virgin olive oil [1], polyphenols in tea and in bee-pollen
[2, 3], biogenic amines in wines [4] and the analysis of extracts of Aloe plants [5]. During the
development of the methods, special attention was paid to the selection of appropriate stationary
phase, to sample preparation and focusing as well as to the coupling with mass spectrometry
when employed.
Refernces:
[1] Rocco A. et al. J. Chromatogr. A 1216, 7173, (2009)
[2] Fanali C. et al., J. Chromatogr. A 1234, 38, (2012)
[3] Fanali C. et al., J. Chromatogr. A 1313, 270, (2013)
[4] Hernndez-Borges J. et al., J. Chromatogr. A 1147, 192, (2007)
[5] Fanali S. et al., J. Sep. Sc. 33, 2663, (2010)

15

L.15
NANO-LC-MS DETERMINATION OF SULFONAMIDES
EMPLOYING CORE-SHELL PARTICLES

RESIDUES

IN

MILK

Silvia Rocchi1,2, Giovanni D'Orazio1, Giorgio Cerichelli2, Salvatore Fanali1

1

Institute of Chemical Methodologies - Italian National Research Council, Via Salaria Km 29,300,
00015 Monterotondo, Italy
2

Department of Physical and Chemical Sciences - University of L'Aquila, Via Vetoio (Coppito 1),
67100 L'Aquila, Italy

Sulfonamides are a group of synthetic antimicrobial agents quite commonly used for therapeutic
and prophylactic purposes in veterinary practice. Due to their wide spectrum of activity and low
cost, they are administered to farm animals in industrial production. Additionally, acting as growth
promoting substances, they could be improperly used as excessive administration . An
inappropriate use of these antibiotics can result in the occurrence of unwanted residues in
animal-derived food-products, such as milk, involving negative repercussions on consumer
health [1].
In order to maintain their presence in foodstuffs of animal origin at safe levels, the European
Commission has established maximum residue limit of 100 g/kg for sulfa-drugs in milk, muscle,
fat, liver, kidney of all food-producing animal species [2].
In this study, a nano-LC/ESI/MS method for the simultaneous determination of eighteen
sulfonamides in milk was developed [3]. An additional confirmation of sulfonamides was done
with MS/MS experiments. The use of a miniaturized liquid chromatographic technique leads to
a series of advantages such as low consumption of mobile and stationary phase, minute sample
volume injection, reduction of chromatographic dilution and easy coupling with mass
spectrometry.
A capillary column (100 m internal diameter) packed following the slurry packing procedure
with a recent commercialized stationary phase, Kinetex C18 core-shell, was employed for the
chromatographic separation in gradient mode with a water-acetonitrile mobile phase containing
0.1 % (v/v) of formic acid. The sensitivity improvement by on-column focusing, evaluating the
appropriate sample solvent and establishing the maximum injection volume, and the use of a
rapid pretreatment consisting of acid deproteinization followed by solid-phase extraction,
allowed to obtain in milk quantitation limits in the range 8-96 g/kg and recoveries 884%.

References:
[1] Wang S. et al., Food Additives and Contaminants 23, 362, (2006)
[2] Commission of the European Communities, Off. J. Eur. Commun., Commission Regulation
(EC) No. 508/1999 of 4 March 1999, L60/16
[3] DOrazio G. et al., J. Chromatogr. A 1255, 277, (2012)

16

L.16
SVILUPPO DI METODICHE ANALITICHE AVANZATE PER
TRIACILGLICEROLI IN CAMPIONI COMPLESSI (OLI DI PESCE)

L'ANALISI

DI

Giuseppe Sullini1, Marco Beccaria1, Paola Donato2,1, Francesco Cacciola3,4, Paola Dugo1,2, Luigi
Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

viale Annunziata, 98168 Messina, Italy

2 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via lvaro del Portillo,

00128 Roma, Italy

3

Dipartimento di Scienze dell'Ambiente, della Sicurezza, del Territorio, degli Alimenti e della Salute,
University of Messina, viale F. Stagno d'Alcontres, 98166 Messina, Italy
4

Chromaleont s.r.l. - A start-up of the University of Messina, viale Annunziata, 98168 Messina, Italy

La determinazione dei triacilgliceroli (TAGs) negli oli di pesce una problematica complessa
e le tecniche analitiche basate su cromatografia liquida convenzionale si sono rivelate spesso
insufficienti per determinarne il profilo completo, data la complessit di tali campioni. Tali oli
contengono una grande variet di acidi grassi (FAs), soprattutto PUFAs (acidi grassi polinsaturi
a lunga catena). Lolio di menadi (Brevoortia tyrannus) contiene pi di 40 differenti FAs [1], da
12 a 22 atomi di carbonio (CN), da 0 a 18 doppi legami (NDB). La combinazione di questi FAs
pu portare alla formazione biosintetica di migliaia di differenti TAGs [2].
Caratteristici di questolio sono gli acidi grassi eicosapentaenoico, 20:5(3) (EPA o Ep) e
docosaesaenoico, 22:6(3) (DHA o Dh), i quali svolgono un importante ruolo preventivo in
alcune patologie [3].
In questo contributo vengono descritte metodiche analitiche avanzate per la caratterizzazione
triacilglicerolica dellolio di menadi. Lutilizzo di colonne HPLC accoppiate in serie in condizioni
di pressione ultra elevata (UHPLC) e di tecniche multidimensionali nella modalit
comprehensive hanno permesso di aumentare notevolmente lefficienza separativa rispetto
alla cromatografia liquida convenzionale.
Inoltre, grazie allaccoppiamento di queste tecniche cromatografiche con la spettrometria di
massa, sono state ottenute informazioni di tipo strutturale su questi analiti.
References:
[1] Mondello L., Tranchida P.Q., Costa R., Casilli A., Dugo P., Cotroneo A., Dugo G., J. Sep.
Sci. 26, 1467, (2003)
[2] Franois I., dos Santos Pereira A., Sandra P., J. Sep. Sci. 33, 1504, (2010)
[3] Russo G.L., Biochem. Pharmacol. 77, 937, (2009)

17

L.17
MASS SPECTROMETRIC CHARACTERIZATION OF UNKNOWN N,O SUBSTITUTED
PHENYLALANINE AND PHENYLALANINE METHYL ESTERS IN EXTRAVIRGIN
OLIVE OIL
Chiara Cavaliere, Giuseppe Caruso, Susy Piovesana, Riccardo Zenezini Chiozzi
Dipartimento di Chimica - Sapienza Universit di Roma, Piazzale Aldo Moro 5, 185 Rome, Italy

The extravirgin olive oil, differently from the other vegetable oils obtained mainly from seeds
with various treatments, is obtained from a fruit (Olea Europea L.) by solely mechanical
means. This characteristic allows extravirgin olive oil to partially keep the original chemical
composition of the fruit, in particular as concerns polar fraction, usually eliminated during other
vegetable oil preparation.
Besides the high monounsaturated fatty acid content, olive oil owes its popularity as healthy
food to the presence of minor components possessing a high biological activity, such as
antioxidant phenolics.
In a recent study on polyphenol content of extravirgin olive oil by ultra-high performance liquid
chromatography followed by high resolution mass spectrometry with a time of flight mass
analyzer and positive electrospray ionization, we revealed the presence of many intense ion
signals having an even m/z ratio, and therefore supposed to contain an odd number of nitrogen
atoms. These compound typology has never been reported in literature previously.
From the accurate mass measurements of these peaks (in most cases with accuracy below 2
ppm) and the study of their tandem mass spectra, we could infer that these substances are
substituted phenylalanine and phenylalanine methyl esters, N-hydroxy. Their general formula
is reported in the figure below, where R1 is H or methyl, R2 is H or a carbonyl (formyl, keto- or
carboxyalkyl) group, R3 is H,methyl, or a residue of phenyl ethyl alcohol, tyrosol, hydroxytyrosol
or dihydroxytyrosol.
The sample was also analyzed with a linear ion trap-Orbitrap instrument to obtain confirmatory
identification and complementary structural information.
At the moment the identification of these compounds is only tentative and needs the support
of other instrumental techniques to be unambiguous. However, from the study of the acquired
mass spectra, a coherent class scheme appears.

18

L.18
NEW TECHNOLOGIES AND NEW KNOWLEDGE: SULFOQUINOVOSYLDIGLYCERIDE
REGIOISOMERS OF Rhodobacter sphaeroides
Sara Granafei1,2, Gabriele Valenza1,2, Ilario Losito1,2, Francesco Palmisano1,2, Tommaso R.
Cataldi1,2
1

Dipartimento di Chimica - Universit di Bari, via Orabona 4, 70126 Bari, Italy

Centro Interdipartimentale SMART - Universit di Bari, Via Orabona 4, 70126 Bari, Italy

In the last 50th years the interest about a new class of glycerolipids, the
sulfoquinovosyldiglycerides (SQDGs), has grown. These sulfolipids, extracted from several
plants and bacteria, have been characterized by tandem mass spectrometry (MS/MS)[1-3] and
some chromatographic separations have been performed to resolve isobaric/isomer sulfolipids
and regioisomers.[4-6] In this work a liquid chromatography with ESI and tandem mass
spectrometry method (LC-ESI-MS/MS) using negative ion detection is described and applied
to characterize each sulfolipid extracted from the photosynthetic bacterium R. sphaeroides. A
reversed phase LC method, using a 2.7 m fused-core C18 column, was devised. The extracted
ion chromatograms (XICs) of putative SQDGs was applied and the presence of sulfolipids at
m/z 793.5 (C32:0), 817.5 (C34:2), 819.5 (C34:1), 821.5 (C34:0), 845.5 (C36:2), 847.5 (C36:1)
and 849.5 (C36:0), was ascertained. Subsequent LC-ESI-CID-MS/MS analysis of these
sulfolipids was used to establish their chemical identity, including three regioisomers at m/z
817.5 (16:1/18:1-18:1/16:1), 819.5 (16:0/18:1-18:1/16:0) and 847.5 (18:0/18:1-18:1/18:0). Very
remarkably was the separation of two isomers at m/z 845.5, never reported before, which
corresponds to an SQDG 18:1/18:1. Most likely these two chains are double-bond position
isomers, perhaps oleic (i.e., 18:1 cis-9) and cis-vaccenic (i.e., 18:1 cis-11) moieties. This
suggestion needs to be verified by GC-MS analysis of fatty acid methyl esters duly derivatized.
References:
[1] Gage D.A., Huang Z.-h., Benning C., Lipids 27, 632, (1992)
[2] Zhang X., Fhaner C.J., Ferguson-Miller S.M., Reid G.E., Int. J. Mass Spectrom. 316,
100, (2012)
[3] Naumann I.D., Kai H. Darsow, Walter C., Lange H.A., Buchholz R., Rapid Commun. Mass
Spectrom. 21, 3185, (2007)
[4] Zianni R., Bianco G., Lelario F., Losito I., Palmisano F., Cataldi T.R.I., J. Mass
Spectrom. 48, 205, (2013)
[5] Keusgen M., Curtis J.M., Thibault P., Walter J.A., Windust A., Ayer S.W., Lipids 32,
1101, (1997)
[6] Xu J., Chen D., Yan X., Chen J., Zhou C., Anal. Chim. Acta 663, 60, (2010)

19

L.19
DETAILED FOOD SAMPLE EXAMINATION: ONE STEP TARGET AND NON-TARGET
ANALYSIS OF CONTAMINANTS BY GC-HR-TOF MS
Jitka Zrostlkov1, Toms Kovalczuk1, Kamila Kalachov2, Lucie Drbov2, Jana Hajslov2, Jos
M. Sangens3
1

LECO European Technical Centre Prague, Sokolovsk 219, 190 00 Prague, Czech Republic

Institute of Chemical Technology, Technick 3, 160 00 Prague, Czech Republic

Sep Science Specialist SW Europ - LECO INSTRUMENTOS S.L., Avda. de la Industria 43, 28760
Tres Cantos, Spain

Today requirements in the analysis of food contaminants target at low ppb concentration levels
and require confident confirmation of identity. At the same time, the methodics of sample
preparation tend to be more time-efficient and universal, which results in less clean extracts
and more potential matrix interference. Therefore new technologies of GC- and LC-MS analysis
need to be implemented to meet the above requirements. New LECO multireflecting GC-TOF
MS provides mass resolution 50,000 (at m/z 219) and mass accuracy 1 ppm at data acquisition
rate up to 200 Hz. In this presentation the results from the analysis of pesticide residues and
other contaminants in heavy food matrices will be demonstrated.

20

L.20
A NOVEL FLUOROUS SOLID-PHASE EXTRACTION METHOD FOR THE TRACE
ANALYSIS OF PERLUOROALKYL ACIDS IN WATERS
Nicola Marchetti1,2, Alberto Cavazzini1, Francesco Dondi1, Aldo Lagana`3
1

Department of Chemistry and Pharmaceutical Sciences - University of Ferrara, via Luigi Borsari,
46, 44121 Ferrara, Italy
2

Laboratorio Terra&Acuqa Tech - Tecnopolo di Ferrara, , Ferrara, Italy

University of Rome "La Sapienza", P.le Aldo Moro, 6, 00185 Roma, Italy

The development of innovative techniques to address the quality of waters and preserve it is
one of the major topics that European Community is still strongly supporting across different
scientific areas. With respect to Separation Sciences, there are some subject fields that have
a key-enabling role in this process: development of new solid porous stationary phases,
fundamental research in chromatography focused on the characterization of these materials,
and the study of innovative sample preparation methods for trace analysis. The goal of the
present work is to enhance the actual potential of multiresidual analytical approaches for
determining environmental contaminants by means of improving selectivity and sensitivity.
Insights into retention mechanisms involved in the interaction between perfluoroalkyl acids
(PFAs) and perfluorinated porous solid adsorbent materials are investigated. Additionally, the
wetting behaviour of such stationary phases is studied to achieve useful information for sample
preparation purposes. The solid support is silica based, superficially derivatized with
perfluoroalkyl straight chains. This material shows unique characteristics allowing specific
selectivity mostly driven by fluoro-affinity (fluorophilicity). The influence of mobile phase
composition on the capture of PFAs on the solid adsorbent (enrichment) and their release
(clean-up and elution) is addressed. Linear tracer pulse liquid chromatography measurements
performed with a linear ion trap mass spectrometric detector are employed to investigate the
solid phase adsorption behavior for the mobile phase components. The results are gathered
and employed to develop a specific, selective and innovative fluorous-SPE (F-SPE) method,
specifically optimized, validated and applied to the trace analysis of the environmentally relevant
PFAs in waters. Direct MS detector response, internal standard calibration and mass recoveries
are evaluated under different experimental conditions.

21

L.21
EASY PESTICIDE ISOLATION & CONCENTRATION SYSTEM - IDENTIFICATION OF
300 PESTICIDES IN VARIOUS FOOD MATRICES WITHOUT SAMPLE PREPARATION
BY 2D-LC-MS/MS
Eligio Sebastiani1, M. Balci2, G. Gursu2
1

SRA INSTRUMENTS, Viale Assunta 101, 20063 Cernusco sul Naviglio Milano, Italy

JASEM LABORATUVAR Inonu Mah, Kartal Cad. No:55, Kat:2 34755 Kayisdagi-Atasehir-Istanbul,
Turkey

Analysis of pesticide residues is one of the most important tasksin food safety control. The
majority of the monitored pesticides areanalyzed by LC-MS/MS technique. But the existence
of sample matrix strongly affects the analyticalresults.
A remarkable low recovery rate caused by matrix effects (suppression) and interferences with
matrix compounds could be observed in experiments. For this reason the isolation of the sample
matrix from the target analytes is required in order to obtain high quality data.
EPICS (Easy Pesticide Isolation and Concentration System) is the first fully automated matrix
separation technique for an improved pesticide analysis using LC-MS/MS.
Pesticide residues aresimply analyzed from an acetonitrile raw extract. After centrifugation an
aliquot of the supernatantis directly injected into the LC system. Using EPICS the interfering
matrix constituents are isolated from the target compounds by means of a specific HILIC column
(Hydrophilic Interaction Liquid Chromatography). Further time-consuming clean-up steps are
not required. Automation with EPICS helps you to enhance the sample throughput and the
productivity of your laboratory.
Main advantages of Using EPICS:
- High sample throughput, enhancement of laboratory productivity
- Improvement of data quality due to matrix isolation
- Minimizing of manual sample preparation steps compared to other methods
- Environmentally compatible because of minimal chemical usage
- Simplified analytic only one injection is required to detect all pesticide residues
- High recovery rates for all pesticides, even for difficult analytes
Info http://www.jasem.biz

22

L.22
ANALISI DELLA FRAZIONE INSAPONIFICABILE DI LIPIDI PROVENIENTI DA VARI
TIPI DI LATTE MEDIANTE GC COMPREHENSIVE E SIMULTANEA RIVELAZIONE MS/
FID, IN SINERGIA CON UN SISTEMA TOF AD ALTA RISOLUZIONE PER LA
DELUCIDAZIONE STRUTTURALE
Simona Salivo1, Peter Q. Tranchida1, Paola Dugo1,2, Luigi Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute - Universit degli Studi di Messina,

Viale Annunziata, 98168 Messina, Italy

2

Centro Integrato di Ricerca (C.I.R.) - Universit Campus Bio-Medico, Via lvaro del Portillo, 21,
00128 Roma, Italy

La presente ricerca focalizzata sull'utilizzo di un metodo gascromatografico bidimensionale

comprehensive (GC GC) con doppia rivelazione, mediante lutilizzo simultaneo della
spettrometria di massa e di un rivelatore a ionizzazione di fiamma (MS/FID), per l'analisi
qualitativa e quantitativa della frazione insaponificabile dei lipidi del latte (burro di vacca, latte
di bufala, pecora e capra). La capacit dello spettrometro di massa di operare in condizioni di
alta risoluzione a tempo di volo (ToF MS) ha reso possibile la delucidazione della struttura di
alcuni componenti (in particolare gli steroli. Il set di colonne GC GC utilizzato consiste in una
prima dimensione a bassa polarit ed una seconda di media polarit, entrambe caratterizzate
da una elevata stabilit termica. L'utilizzo della doppia rilevazione ha consentito il
contemporaneo ottenimento di informazioni quali/quantitative. La complessit del fingerprint
generato dalla frazione insaponificabile giustifica pienamente l'impiego della tecnologia
bidimensionale GC. In particolare, grazie allutilizzo della cromatografia comprehensive stato
ottenuto un aumento del livello di sensibilit e la contemporanea formazione di raggruppamenti
ordinati di classi di composti nello spazio bidimensionale. Inoltre, le informazioni relative ai dati
di massa esatta ottenuti in alta risoluzione hanno reso possibile l'identificazione di alcuni
costituenti lipidici del latte, i cui spettri non erano contenuti nei database spettrali di massa
impiegati.

23

L.23
FEASIBILITY OF DIRECT-EI LC-MS FOR FOOD SAFETY APPLICATIONS:
IDENTIFICATION AND QUANTITATION OF ENVIRONMENTAL CONTAMINANTS IN
MILK POWDER.
Fabiana Capriotti1, Achille Cappiello1, Giorgio Famiglini1, Veronica Termopoli1, Pierangela
Palma1, Nick Cellar2
1

DiSTeVA - UNIVERSITA DEGLI STUDI DI URBINO, P.zza Rinascimento 6, 61029 URBINO, Italy

2 Abbott

Nutrition, 3300 Stelzer Road, 43219 Columbus, United States

Raw materials coming from developing countries are increasingly used in food industries due
to their low cost. Sometimes, these ingredients are not subjected to severe safety controls in
the countries of origin. This represents a great concern when food preparation is involved. It is
mandatory to have efficient and sensitive analytical methods to detect food contamination. We
present the feasibility of Direct-EI/LC-MS instrumentation to determine some environmental
contaminant residues in milk powder. This technique is matrix effect free and gives full EI mass
spectra of the analytes for unparallel identification. To demonstrate the feasibility, reconstituted
milk powders were spiked with an environmental contaminant standard mixture. The samples
were extracted via QuEChERS methodology. Spiked and blank sample extracts were separated
on a nanoAcquity UPLC column, 1.8m, (0.075 mm x 150 mm) with a flow rate of 400 nL/min.
The column was coupled via Direct-EI to the ion source of an Agilent 5975C MSD. Signal was
collected in full scan- and SIM-modes for qualitative and quantitative analyses. Linearity and
limit of detection were determined for method validation. Twelve compounds belonging to the
classes of phenols, estrogens, carbamoyloxime, triazine, methylcarbamate and phenoxy acids
have been considered. They are all involved in food contamination and in some cases present
difficulties in high resolution, accurate mass ESI-LC/MS identification. For example:
diethylstilbestrol returns over 900 entries with its exact mass searching by Chemspider. The
mass spectrometer was operated in SIM-mode for specificity and to maximize signal response.
Instrumental limits of detection are in the range of 0.35-5 ng that correspond to method limits
of detection ranging from 7 to 100 ppb due to a pre-concentration step during the QuEChERS
extraction. Real samples will be analyzed to confirm the usefulness of the method. In the
following figure, the Direct EI interface is shown.

24

L.24
VARIAZIONE DEL CONTENUTO IN ACIDI FENOLICI E DELL'ATTIVITA'
ANTIOSSIDANTE NEI PRODOTTI DI DECORTICAZIONE DEL FRUMENTO DURO
Federica Taddei1, Daniela Martini1,2, Isabella Nicoletti3, Danilo Corradini3, Maria Grazia D'Egidio1
1

Consiglio per la Ricerca e la Sperimentazione in Agricoltura (CRA)-Unit di Ricerca per la

Valorizzazione Qualitativa dei Cereali, via Cassia 176, 00191 Roma, Italy
2

Universit Campus Bio-Medico, via Alvaro del Portillo, 21, 00128 Roma, Italy

3 Consiglio Nazionale delle Ricerche (CNR)-Istituto di Metodologie Chimiche, Via Salaria km 29.300,

00015 Montelibretti (RM), Italy

Il consumo di alimenti integrali in questi ultimi anni in crescente aumento essendo oramai
noti, anche al consumatore, gli aspetti salutistici degli alimenti a base di cereali contenenti
frazioni cruscali ricche di composti bioattivi. Per contro, le frazioni cruscali, principalmente
costituite dagli strati pi esterni della cariosside (tegumenti), in particolare gli strati pi superficiali
dei tegumenti, possono contenere alcuni contaminanti (micotossine, metalli pesanti, pesticidi
ecc.) che abbassano la qualit igienico-sanitaria degli alimenti integrali. Il processo della
decorticazione, tradizionalmente utilizzato per svestire dalle glume fortemente adese le
cariossidi di cereali come orzo ed avena, pu essere applicato anche alle cariossidi nude al
fine sia di ridurre le eventuali sostanze contaminanti presenti in superficie sia preservare il
contenuto di composti bioattivi. In questo studio sono state eseguite sulle cariossidi di frumento
duro decorticazioni in successione di 15 secondi ciascuna, raccogliendo ad ogni tempo tutte
le frazioni (lo scarto di decorticazione e una quota di granella decorticata). Per tutte le frazioni
raccolte stata valutata la capacit antiossidante totale mediante il metodo diretto di Serpen
et al [1] ed il contenuto in acidi fenolici liberi, coniugati e legati estratti adattando la procedura
precedentemente descritta [2] alle matrici in esame. La separazione, quantificazione e
identificazione degli acidi fenolici stata eseguita mediante cromatografia liquida ad elevate
prestazioni in fase inversa (RP-HPLC) con rivelatore a serie di fotodiodi (PDA), accoppiata alla
spettrometria di massa con sorgente di ioni per elettrovaporizzazione (ESI-MS) per confermare
lidentificazione. La quantificazione stata eseguita con il metodo dello standard interno,
sottoponendo gli standard di acidi fenolici e lo standard interno alle stesse procedure di
estrazione effettuate per le tre forme degli acidi fenolici. I risultati hanno dimostrato che il
processo di decorticazione pu essere utilizzato con successo prima del processo di
macinazione tradizionale per ottenere prodotti trasformati ricchi in composti bioattivi ed allo
stesso tempo con unelevata qualit igienico-sanitaria.

Bibliografia:
[1] Serpen A. et al., J. Cereal Sci. 48 (2008) 816.
[2] Nicoletti I. et al., J. Agric. Food Chem. (2013). In pubblicazione

25

L.25
CHIRAL VOLATILE COMPOUNDS FOR THE DETERMINATION OF ORANGE HONEY
AUTHENTICITY
Antonella Verzera, Gianluca Tripodi, Concetta Condurso, Giovanna Dima
Dipartimento di Scienze Chimiche - University of Messina, Viale F.Stagno d'Alcontres, 31, 98166
Messina, Italy

Assessment of the botanical origin of the unifloral honeys is of great concern in the context of
consumer protection and quality control. Nevertheless, the methods that are currently available
are not satisfactory. In order to find alternatives to the time consuming and uncertain methods,
a new analytical approach is proposed; it is based on the enantiomeric ratio investigation of
chiral volatile constituents which are derived from the plants being visited by the bees. The
method was applied to orange honeys; firstly, the volatile fraction of orange honey and flowers
were studied by SPME-GC-MS; a large number of components were identified in orange honeys
while linalool prevailed among orange flower volatiles. The enantiomeric ratios of linalool and
its oxides were determined and analogous values between honey and flowers resulted. Even
if a wide variability in the amount of typical volatile constituents of orange honeys emerged, the
enantiomeric ratios of linalool and its oxides remained stable and thus less influenced by
production period, conditioning, packaging, storage, etc. As a result the enantiomeric distribution
of the honey volatile constituents that directly come from flowers could represent a rapid and
easy method for floral origin authenticity.

26

L.26
HPLC-MS/MS ANALYSIS OF SHIKIMATE PATHWAY POLYPHENOLS IN NICOTIANA
LANGSDORFFII: EVALUATION OF METABOLIC RESPONSE DUE TO GENETIC
MODIFICATION AND ABIOTIC STRESSES
Claudia Ancillotti, Leonardo Checchini, Lorenzo Ciofi, Sandra Furlanetto, Massimo Del Bubba
Department of Chemistry - University of Florence, Via della Lastruccia 3 - 13, 50019 Sesto Fiorentino
(Florence), Italy

Genetically transformed plants have been studied for testing their resistance towards various
kinds of abiotic stresses, including exposure to heavy metals, in order to investigate new possible
repopulation and/or remediation strategies of polluted soils [1].
Polyphenols are commonly involved in the defence strategy towards different abiotic stresses,
including those related to heavy metal exposure [2], due to both their redox and metal-chelating
properties and have been therefore investigated as possible markers of different kinds of
stresses [3]. Very recently, total polyphenols and shikimic acid have been highlighted as
interesting parameters to evaluate the effect of the exposure of wild type (WT) and GR
transformed Nicotiana langsdorffii to Cr(VI) [4]. Therefore, in this study shikimate-pathway
phenolic compounds have been determined in root and shoot of Nicotiana langsdorffii, either
WT or transformed by the insertion of GR and rolC genes, in response to the presence of Cr
(VI) in the growth medium. The different abundance of the investigated compounds seems to
be related to their position in the shikimate pathway, since the farthest from the shikimic acid
is the position of the phenolic compounds, the highest is its concentration. Gene insertion caused
a general modification in the metabolism of phenolic compounds, leading in the case of the
most expressed shikimate phenolics, to a significant increase in transformed lines. This result
seems to be particularly important for explaining the resistance of genetically modified plants
to Cr(VI) [5].

References:
[1] Zhang Y. and Liu J., J. Hazard. Mater. 189, 357, (2011)
[2] Duressa D. et al., Int. J. Plant Physiol. Biochem. 2, 38, (2010)
[3] Lavid N. et al., Planta 212, 323, (2001)
[4] Fuoco R. et al., J. Plant Physiol. 170, 668, (2013)
[5] Del Bubba M. et al., J. Hazard. Mater. 262, 394, (2013)

27

L.27
QUALITY BY DESIGN APPROACH TOWARS THE DESIGN SPACE DEFINITION IN
THE DEVELOPMENT OF A SOLVENT-MODIFIED ELECTROKINETIC CHROMATOGRAPHY
METHOD
Benedetta Pasquini, Serena Orlandini, Massimo Del Bubba, Sergio Pinzauti, Sandra Furlanetto
Dipartimento di Chimica - Universit degli studi di Firenze, via Ugo Schiff 6, 50019 Sesto Fiorentino,
Italy

Analytical methods used for the determination of active pharmaceutical ingredients and drug
products are an integral part of the Quality by Design (QbD) concept that is outlined in ICH
Guideline Q8 for pharmaceutical development [1]. The method used for analysis should meet
its intended purpose similar to the product requirements for a clinical dosage form [2]. The
application of QbD leads to the definition of Design Space (DS), a multidimensional space,
which includes any combination of the variables that have been demonstrated to provide
assurance of quality of the data produced by the method [2]. A solvent-modified micellar
electrokinetic chromatography method was developed for the simultaneous analysis of the
tricyclic antidepressant amitriptyline (AMI) and its main impurities. In a scouting phase, a suitable
pseudostationary phase was selected. Screening experimental design strategies were then
applied for investigating knowledge space. Critical process parameters were represented by
the process variables voltage, temperature, buffer concentration and pH, concentration of the
surfactant, of the cosurfactant and of the organic modifiers. The critical quality attributes (CQAs)
investigated were critical resolution values and analysis time. The DS was finally identified by
Response Surface Methodology as the multidimensional combination of variables where the
probability for the different considered CQAs to be acceptable was higher than a defined quality
level. The developed method was validated and applied to AMI tablets.
References:
[1] ICH Harmonised Tripartite Guideline. Pharmaceutical Development Q8(R2) (2009)
International Conference on Harmonisation of technical requirements for registration of
pharmaceuticals for human use.
[2] Orlandini S., Pinzauti S., Furlanetto S., Anal. Bioanal. Chem. 405, 443, (2013)

28

L.28
DETERMINAZIONE DI CONTAMINANTI ORGANICI IN MIELI SICILIANI (ITALIA)
Marcello Saitta, Maria Rita Fede, Giacomo Dugo
Dipartimento S.A.S.T.A.S - Universit degli studi di Messina, Viale F. Stagno D'alcontres, 31, 98166
Messina, Italy

Il miele negli ultimi anni ha subito una contaminazione da prodotti fitosanitari e da diverse
sostanze inquinanti, che possono residuare in esso. Lo scopo di questo lavoro stato quello
di determinare in campioni di miele, prodotti esclusivamente in Sicilia, eventuali residui di
contaminanti organici, quali IPA, PCB, pesticidi organoclorici ed organofosforici ed altri composti
(erbicidi, fungicidi, insetticidi, acaricidi) mediante gascromatografia accoppiata alla
spettrometria di massa.
Sono stati analizzati n21 campioni di mieli monofloreali e millefiori, forniti dagli apicoltori di tre
diverse province siciliane (Messina, Catania ed Agrigento). Dopo opportuno trattamento, gli
estratti sono stati sottoposti ad analisi gascromatografica con analizzatore di massa a triplo
quadrupolo (GC-MS/MS Thermo Scientific).
I valori di recupero (spike a 20 g/kg, media di 3 determinazioni) variavano da un minimo di 38
4% ad un massimo di 15224%; i valori di LOD variavano da 0.01 a 45.6 g/kg. Dai risultati
ottenuti si pu osservare che tutti i campioni analizzati contenevano residui di chlorfenvinphos
e di coumaphos (rispettivamente nel range 0.33-4.94 g/kg e 1.26-23 g/kg), insetticidi impiegati
in apicoltura.
Dai valori ritrovati si evince che il 19% dei campioni contiene residui di pesticidi oltre i limiti di
legge. Si potrebbe ipotizzare una contaminazione di fondo dei mieli, molto probabilmente
dovuta al riciclo delle cere, pratica che ciascun apicoltore compie normalmente. La cera, molto
affine nei confronti dei contaminanti organici, ha avuto nel passato una funzione di accumulo
ed ha, nel presente, una funzione di lento rilascio dei contaminanti nel miele.

29

P.01
ULTRA FAST ELUCIDATION OF FLAVONOIDS PROFILE IN BERGAMOT JUICE BY
UHPLC-IT-TOF
Eduardo M. Sommella, Giacomo Pepe, Francesco Pagano, Giovanni Ventre, Raffaella
Mastrocinque, Pietro Campiglia
Farmabiomed - Universit degli studi di Salerno, via Giovanni Paolo II 132, 84084 Fisciano (SA), Italy

In recent years flavonoids have gained considerable attention, as they are involved in a variety
of functions in plants, but especially for the large number of beneficial effects on human health,
including antioxidant, cardioprotective, anticancer, and hypolipidemic potential. Despite being
considered a waste product, the bergamot juice contains a large amount of bioactive flavonoids.
The analysis of this matrix is usually accomplished by RP-HPLC carried out on conventional
particle columns, coupled to low resolution MS instruments. In this contribution we have
developed a UHPLC-IT-TOF platform for the fast and accurate profiling of Citrus Bergamia
Flavonoids. With respect to the typical methods for the analysis of these matrices, based on
conventionl HPLC techniques, a ten fold faster separation was attained. The use of a core shell
particle column ensured high resolution, in less than 5 minutes of analysis time. Unambiguous
determination of flavonoid identity was obtained by the employment of a hybrid IT-TOF mass
spectrometer with high mass accuracy (average error 1.69 ppm). The system showed good
retention time and peak area repeatibility, with maximum RSD% values of 0.36 and 3.86,
respectively, as well as good linearity (R2 0.99). Our results show that UHPLC can be a
powerful tool for ultra fast quali/quantitative analysis of flavonoid compounds in Citrus fruit juices.

30

P.02
INDAGINE SULLA FRAZIONE ETEROCICLICA OSSIGENATA DEL SUCCO DI
LIMETTA ( CITRUS AURANTIFOLIA (CHRISTM) SWINGLE)
Selenia De Grazia1, Marina Russo1, Elisa Grasso1, Rosaria Costa1, Paola Dugo1,2, Luigi
Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

viale ss. Annunziata, 98168 Messina, Italy

2 Centro Integrato di Ricerca (C.I.R) - University Campus Bio-Medico di Roma, via Alvaro del Portillo,

21, 00128 Roma, Italy

Nellambito della produzione mondiale di succhi di Citrus, si possono distinguere quattro

principali gruppi di frutti impiegati, in ordine decrescente, per la produzione di succo: i) arance,
ii) tangerini, mandarini e clementine, iii) limoni e limette, iv) pompelmi. Questi ultimi, in particolare,
sono coltivati quasi esclusivamente per l'estrazione del succo.
La letteratura ricchissima di lavori sulle specie appartenenti al genere Citrus; si tratta infatti
di piante impiegate in tutto il mondo come materia prima per l'industria alimentare, farmaceutica
e cosmetica. Anche Citrus aurantifolia e Citrus latifolia, ovvero le specie pi diffuse di limetta,
sono oggetto di un elevato numero di pubblicazioni scientifiche, per lo pi relativamente alla
composizione chimica dellolio essenziale. In realt, ben poco o quasi nulla stato riportato,
ad oggi, sulla composizione chimica del succo ottenuto dai frutti di limetta.
Lobiettivo di questo lavoro stato dunque lo studio della frazione eterociclica ossigenata,
costituita da flavonoidi, cumarine e psoraleni, di campioni di succo di limetta ottenuto in
laboratorio. Lindagine stata effettuata mediante cromatografia liquida in fase inversa (RPHPLC) con rivelazione UV, utilizzando delle colonne prodotte con tecnologia fused-core,
impaccate con particelle di 2.7 m di diametro. Il vantaggio principale di tale tecnologia consiste
in una riduzione dei tempi di residenza dellanalita nella fase stazionaria, di conseguenza le
analisi sono pi efficienti (migliore forma del picco) e pi veloci.
I flavonoidi pi abbondanti che sono stati determinati sono: lesperidina (367,0 16,0 ppm) e
leriocitrina (148.0 7.9 ppm). Tra le 15 cumarine e furocumarine determinate, la bergamottina
(29,6 1,1 ppm), la 5-geranilossi-7-metossicumarina (16,5 0,6 ppm) e lossipeucedanina
idrato (9.9 0.5 ppm) costituivano i composti predominanti. L'alto contenuto di queste molecole,
che possiedono effetti benefici sulla salute umana, suggerisce il consumo di succo di limetta
su base giornaliera.

31

P.03
MULTIVARIATE OPTIMIZATION OF A SPME-GC-MS METHOD FOR THE
DETERMINATION OF TERPENES IN NICOTIANA LANGSDORFFII: APPLICATION TO
WILD-TYPE AND MUTANT GENOTYPES
Francisco Ardini, Marina Di Carro, Maria Luisa Abelmoschi, Emanuele Magi
Department of Chemistry and Industrial Chemistry - University of Genoa, Via Dodecaneso 31, 16146
Genoa, Italy

Terpenes are a large class of natural organic compounds, typically produced by plants in their
leaves.[1] Among them, monoterpenes (C10) represent an important class of volatiles called
volatile terpenes which play a key role in plant protection:[2] in fact, plants can react to stress
factors by inducing emissions of these volatiles, with responses strongly depending on the
stress (typology, severity, duration) and the conditions of the plant (species, physiological
status).[3] So, detecting the variations in volatile emissions of plants submitted to stresses would
facilitate the search for new sustainable methods for environmental control.[1] Because of their
importance, terpenes were included as objects of study in the Italian research project of national
interest (PRIN) Study of stress indicators in plants by new analytical methodologies, aimed
to assess possible indicators of stress-induced metabolic alterations in Nicotiana langsdorffii
genotypes.
In this work, we developed a new method based on headspace solid-phase microextraction
gas cromatography mass spectrometry (HS-SPME-GC-MS) for the determination of terpenes
in the leaves of Nicotiana langsdorffii. Five monoterpenes (-pinene, limonene, linalool, terpineol and geraniol) were chosen for the maximization of the extraction conditions and for
the quantitative determination. An experimental design (Doehlert) has been set up for the
optimization of the main experimental conditions, using a commercial sample of N.
langsdorffii as the object of study. After the assessment of the principal figures of merit, the
developed procedure was finally applied to wild-type and genetically modified specimens
subjected or not to different abiotic stresses: chemical, water and thermal stress. The obtained
results showed low levels of terpene emissions (ng g-1 level or lower) with the exception of
geraniol (hundreds of ng g-1 level), while the genetic modifications and the abiotic stresses had
different effects on the analyte concentrations.
References:
[1] Maffei M. E. et al., Nat. Prod. Rep. 28 , 1359, (2011)
[2] Loreto F. and Schnitzler J.-P., Trends Plant Sci. 15, 154, (2010)
[3] Niinemets U., Trends Plant Sci. 15, 145, (2010)

32

P.04
DETERMINAZIONE DELLA TRACCIABILITA' DI MATRICI COMPLESSE NATURALI
DI INTERESSE ALIMENTARE MEDIANTE LA GASCROMATOGRAFIA ACCOPPIATA
ALLA SPETTROMETRIA DI MASSA A RAPPORTO ISOTOPICO DEL CARBONIO (GCC-IRMS).
Luisa Schipilliti1, Ivana L. Bonaccorsi1, Paola Dugo1,2, Luigi Mondello1,2
1

Dipartimento Scienze del Farmaco e dei Prodotti per la Salute - University of Messina, Viale
Annunziata, 98168 Messina, Italy
2 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via lvaro del Portillo,

21, 00128 Rome, Italy

Lo studio dei composti aromatici della frazione volatile di differenti matrici alimentari stato
condotto per mezzo della gascromatografia accoppiata alla spettrometria di massa a rapporto
isotopico del carbonio (GC-C-IRMS), allo scopo di determinare dei parametri analitici utili per
definire la provenienza e lautenticit dei campioni in esame. Le matrici oggetto della presente
ricerca sono state: oli essenziali agrumari, alimenti aromatizzati alla frutta, rosoli agrumari e
campioni di caff tostato. Poich il frazionamento isotopico del carbonio legato alle condizioni
ambientali in cui una pianta si sviluppa e compie il suo ciclo fotosintetico, i valori relativi al
rapporto isotopico del carbonio (13C) possono essere indicativi della provenienza botanica e
geografica delle matrici. Inoltre, poich ogni sostanza naturale mostra un ben definito rapporto
isotopico dei suoi elementi costituenti, mediante la tecnica GC-C-IRMS, possibile svelare la
presenza di sostanze natural sintetiche nelle matrici alimentari, aggiunte illecitamente come
aromatizzanti. La frazione volatile stata estratta direttamente dai campioni, mediante la tecnica
della microestrazione in fase solida dello spazio di testa (HS-SPME), impiegando, in base alla
matrice in esame, il tipo di fibra pi opportuna. Il riconoscimento qualitativo dei composti volatili
stato effettuato prima dellanalisi dei rapporti isotopici, mediante gascromatografia accoppiata
alla spettrometria di massa quadrupolare (GC-MS). Infine, per confermare e completare i risultati
di questo studio, sono state condotte analisi aggiuntive della distribuzione enantiomerica dei
composti volatili chirali.

33

P.05
VALUTAZIONE DI UNA FASE STAZIONARIA A LIQUIDI IONICI DI MEDIA POLARIT
PER LANALISI DI UN OLIO ESSENZIALE DI MENTA PIPERITA
Elisa Grasso1, Carla Ragonese1, Danilo Sciarrone1, Luigi Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

Viale Annunziata, 98168 Messina, Italy

2 Centro Integrato di Ricerca (C.I.R.) - Universit Campus Bio-Medico di Rome, Via Alvaro del Portillo,

00128 Roma, Italy

Gli oli essenziali sono spesso matrici altamente complesse, in quanto la loro composizione
caratterizzata da numerosi composti, appartenenti a differenti classi chimiche.
La frazione volatile degli oli essenziali (85-99%), costituita da idrocarburi monoterpenici,
sesquiterpenici e dai loro derivati ossigenati (aldeidi, chetoni, acidi e esteri) di gran lunga
predominante rispetto a quella non volatile (1-15%).
La tecnica comunemente utilizzata per lo studio di tali matrici la gas cromatografia, con
approcci analitici che prevedono lutilizzo di colonne polari ed apolari, utilizzati sia a scopi
qualitativi che quantitativi.
Lobiettivo di questa ricerca stato quello di valutare una nuova fase stazionaria a liquidi ionici
a media polarit di recente introduzione per lanalisi GC-MS dellolio essenziale di Menta
piperita.
La valutazione di tale fase stazionaria, disponibile in commercio come SLB-IL60 (30 m 0.20
m df 0.25 mm I.D.), stata effettuata analizzando standard puri di composti generalmente
presenti negli oli essenziali.
I dati di risoluzione tra gli analiti sono stati comparati con quelli ottenuti utilizzando colonne a
media polarit ma diversa selettivit [100% poli(etileneglicole)] ed a bassa polarit (5%
difenil/95% dimetilsilossano), comunemente utilizzate per lanalisi di tali matrici.
I dati ottenuti dallanalisi GC-MS dellolio essenziale di menta piperita hanno evidenziato come
la nuova fase stazionaria a liquidi ionici sia una eccellente alternativa alle comuni fasi stazionarie
impiegate per lanalisi degli oli essenziali essendo in grado di garantire la separazione, e quindi
lidentificazione e quantificazione, di un numero superiore di analiti in una singola analisi gas
cromatografica.

34

P.06
DISPERSIVE LIQUID LIQUID MICROEXTRACTION COUPLED TO HIGH PRESSURE
LIQUID CHROMATOGRAPHY (HPLC) AND ULTRAVIOLET DIODE ARRAY
DETECTION (UV-DAD) FOR THE ANALYSIS OF PHENOLIC COMPOUNDS IN HONEY
Luca Campone1, Sonia Carabetta2, Imma Pagano1, Anna Lisa Piccinelli1, Luca Rastrelli1, Maria
Teresa Russo2
1 Dipartimento di Farmacia - Universit Degli Studi di Salerno, via Ponte Don Melillo, 84084 Fisciano,

SA, Italy
2 Dipartimento di Agraria - Universit Mediterranea di Reggio Calabria, Salita Melissari, 89124 Reggio

Calabria, Italy

A novel approach for the rapid analysis of phenolic componds in honey samples based on
dispersive liquid liquid microextraction (DLLME) is presented. Generally, the analysis of phenolic
compounds in honey involves the elimination of matrix components, mainly sugars, and the
preconcentration of analytes before the determination, which is carried out most often by HPLC
coupled with differents detector [1, 2]. Liquid-liquid extraction (LLE) with organic solvents and
solid phase extraction (SPE) are very often used to extract phenolic compounds from honey
[1, 2]. SPE on Amberlite XAD-2 followed by LLE with diethyl ether [3] is the most popular
technique applied, but the use of C18 sorbents in SPE clean-up has also been reported [4-5].
These methods are laborious time consuming and use large amounts of organic solvent. The
availability of fast, and inexpensive analytical procedures for the determination of phenolic
compounds in honey is highly demanded for the quality control, the nutraceutical research and
the authentication and characterization of botanical origin. The aim of this research was
developed a fast and inexpensive DLLME method suitable for determination of phenolic
compounds in honey. Of the main phytochemicals reported in honey, seven phenolic acids
(caffeic acid, ellagic acid, ferulic acid, p-coumaric acid, protocatechuic acid, syringic acid and
vanillic acid) and ten flavonoids (apigenin, chrysin, galangin, hesperetin, kaempferol, luteolin,
myricetin, pinobanksin, pinocembrin and quercetin) were selected as target analytes. The main
parameters affecting on DLLME efficiency, such as extraction solvent and dispersant solvent,
their volume, the matrix/water ratio, types and salt amount, water pH were carefully studied and
optimised to achieve the best extraction efficiency. HPLC-UV was selected as detection method
and HPLC-HRMS was used to characterize the compounds extracted by DLLME and to
investigate the applicability of this extraction technique to other phytochemicals of honey. After
the optimization, the developed analytical procedure was applied to the analysis of Calabrian
honey samples of different botanical origin. Moreover the analytical performance of DLLME
method were compared with the several methods most used in the analysis of phenolic
compounds in honey. The proposed method, which is demonstrated to be quick, cheap, accurate
and highly selective, was successfully applied to the analysis of tipical Italian honey.
References:
[1] Gmez-Caravaca A.M., Gmez-Romero M., Arrez-Romn D., Segura-Carretero A.,
Fernndez-Gutirrez A., J. Pharm. Biomed. Anal. 41, 1220, (2006)
[2] Pyrzynska K., Biesaga M., Trends Anal. Chem. 28, 893, (2009)
[3] Ferreres F., Toms-Barbern F.A., Soler C., Garca-Viguera C., Ortiz C., Toms-Lorente F.,
Apidologie 25, 21, (1994)
35

P.07
POLYCHLORINATED BIPHENYLS ADSORPTION ONTO ALGINATE BEADS AND
HYBRID ALGINATE MONTMORILLONITE BEADS: ISOTHERMAL AND KINETIC
STUDIES
Salvatore Barreca, Santino Orecchio, Andrea Pace
Dipartimento Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche - University, Viale delle
scienze Edif. 16, 90128 Palermo, Italy

To study the effect of montmorillonite clay in alginate gel beads for Hydrofobic Organic
coumpound (HOCs) adsorptions, we have compared the isothermal adsorption curves of
alginate gel beads (ABs) and alginate montmorillonite gel beads (MABs). In particular, calcium
alginate beads and hybrid calcium alginate montmorillonite beads were used as sorbent for
polychlorinated biphenyls (PCBs) from aqueous solutions. Adsorptions at 25 C were studied
in a batch system, following the kinetics and assessing adsorbent dose and pH effects.
Adsorption isotherms data were modeled using Langmuir, Freundlich and Chapman adsorption
models and the appropriate parameters were calculated.
About PCBs adsorption in ABs, all PCB followed a Freundlich isotherm model, while for MABs,
the absorption of tetra-,penta-, and hexa-chlorobiphenyls displayed a sigmoid-shaped isotherm
or S type. In particular, the Chapman equation showed the highest non-linear R2 values
among the three tested models.
Moreover, for MABs, the kinetic models were investigated to determine the mechanism of
adsorption showing a best fit for the pseudo-second order model (R2 from 0.998 to 0.982).

36

P.08
BENEFICI DELLAPPROCCIO OFF-LINE LC-GCGC IN COMBINAZIONE CON UNO
SPETTROMETRO DI MASSA PER LO STUDIO DI OLI ESSENZIALI DI CITRUS
Mariosimone Zoccali1, Peter Q. Tranchida1, Giovanni Dugo1,2, Luigi Mondello1,3
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

Viale Annunziata, 98168 Messina, Italy

2

Chromaleont s.r.l. A start-up of the University of Messina - Univrsity of Messina, Viale Annunziata,
98168 Messina, Italy
3 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via lvaro del Portillo,

21, 00128 Rome, Italy

La presente ricerca rivolta alla combinazione off-line della cromatografia liquida ad alte
prestazioni e la gas cromatografia multidimensionale comprehensive con uno spettrometro di
massa ad analizzatore quadrupolare (HPLC-GCGC-quadMS), per lo studio della
composizione di oli essenziali del genere Citrus. Nello specifico, stata utilizzata una colonna
di silice per la separazione dei costituenti dei campioni analizzati in due gruppi principali, gli
idrocarburi ed i composti ossigenati. In seguito, ogni frazione stata ridotta in volume, sottoposta
alla separazione GCGC, e caratterizzata tramite lutilizzo di librerie di massa commerciali e
luso interattivo degli indici di ritenzione lineari. Le frazioni ottenute e concentrate dalla preseparazione LC hanno dato origine a dei cromatogrammi bidimensionali sorprendentemente
ricchi e complessi, grazie alle caratteristiche dellaumentato potere di separazione e dell
eccellente sensibilit della tecnica GCGC. I risultati ottenuti sulla frazione dei composti
ossigenati sono particolarmente interessanti poich proprio questi costituenti caratterizzano
principalmente la fragranza/aroma dellolio essenziale, e possono anche essere utilizzati per
la valutazione della sua qualit e genuinit.

37

P.09
COMPARISON BETWEEN VOLATILES ANALYSIS OF EXTRA VIRGIN OLIVE OILS BY
FLASH GAS CHROMATOGRAPHY (FGC) ELECTRONIC NOSE AND BY SOLID
PHASE MICRO-EXTRACTION (SPME) COUPLED WITH GAS CHROMATOGRAPHY MASS SPECTROMETRY (GC-MS)
Enrico Valli1, Federica Tesini1, Sara Barbieri2, Alessandra Bendini1,2, Giuseppe Di Lecce2,
Gallina Toschi1,2

Tullia

1 Interdepartmental Centre for Industrial Agri-Food Research - University of Bologna, piazza Goidanich

60, 47521 Cesena, Italy

2 Department of Agricultural and Food Sciences - University of Bologna, piazza Goidanich 60, 47521

Cesena, Italy

The control of authenticity of extra virgin olive oil (EVOO), one of the most defrauded food in
the world, is based on many determinations of chemical and compositional parameters and,
by the law, also on sensory analysis. It is well known that the sensory descriptors of an EVOO,
both positives and negatives, are due to the presence of many specific volatile compounds,
perceived by olfactory receptors. In particular, the compounds responsible for the recognized
defects (winey, fusty, muddy, musty, rancid), and their biosynthetic or chemical origin are widely
investigated in literature. On the other hand, also the presence of positive olfactory notes
resembling herbs/fruits/vegetables can be linked to the olive varieties/quality and/or
geographical origin, being the lipoxygenase enzymatic pathway influenced by both genetic and
environmental factors. At now, the volatiles profile and/or the presence of specific volatile
markers are not indicated as legal requirements to verify the authenticity of an EVOO, in terms
of assessment to the quality category or belonging to specific quality standards (for example
monovarietal or protected designation of origin/PDO).
In this work, a set of samples sold in Italian markets as EVOOs, but different for geographical
origin, were analysed by a FGC Electronic Nose to develop a fast gas chromatographic
separation of volatile molecules by two short metal capillary columns of different polarities,
mounted in parallel and connected to separate flame ionization detectors. The two
chromatograms are elaborated to give a global olfactory fingerprint of the EVOO samples.
Moreover, to confirm the identity of selected volatile molecules in the headspace of EVOOs, a
solid phase micro-extraction (SPME) coupled to gas chromatographic separation and mass
detection (GC-MS) was used.
The research was carried out with the financial contribution of COOP Italia.

38

P.10
"ZONATION" OF TYPICAL EXTRAVIRGIN OLIVE OILS BY SELECTED MINOR
COMPONENTS AND SENSORY EVALUATION; AN ITALIAN CASE STUDY
Federica Tesini1, Sara Barbieri2, Ziad Ayyad2, Alessandra Bendini1,2, Enrico Valli1, Tullia Gallina
Toschi1,2
1 Interdepartmental Centre for Industrial Agri-Food Research - University of Bologna, piazza Goidanich

60, 47521 Cesena, Italy

2 Department of Agricultural and Food Sciences - University of Bologna, piazza Goidanich 60, 47521

Cesena, Italy

Among vegetable oils, virgin olive oil (VOO) has nutritional and sensory characteristics that to
make it unique and a basic component of the Mediterranean diet [1]. Metabolites found in the
extra virgin olive oils can be tracers for some enzymatic activity. The presence and activity of
enzymes is strictly related to genetic factors (as olive cultivar) and environmental conditions.
The characterization of minor components of extra virgin olive oil, in particular volatile
compounds and phenolic molecules, may contribute to the identification of the cultivar
environment interaction. Many of these compounds can be considered to be direct metabolites
produced by intracellular biogenic pathways.In this work different analytical approaches were
applied for the characterization of extra virgin olive oils produced from autochthon varieties of
olives from Brisighella area which has been sampled specifically to carry on a study of zoning,
by sampling oils obtained in the same product geographical district, but from different agronomic
(types of soils) and agricultural practices (as irrigation).According to the Reg. UE 61/2011, basic
quality parameters and additional determinations (free acidity, peroxide number and
spectrophotometric indices) has been carried on to be able to discriminate the quality of the
samples.The sensory evaluation of the products, conducted by a panel of fully trained judges,
and the comparison of the volatile analyses by SPME/GC-MSD and E-Nose as well as the
determination of phenolic compounds by HPLC-DAD/MSD were conducted together with the
aim to highlight the peculiarities that constitute an important element of typicality valorization.
In this way an in-depth study of the samples has been carried out in order to detect all the
specific characteristics that could contribute to discriminate between samples to underline their
typicality attributes.

Acknowledgements
The research was carried out with the financial contribution of the TERRA DI BRISIGHELLA
- CAB.

References
[1] Bendini A., Cerretani L., Carrasco-Pancorbo A., Gmez-Caravaca A., Segura-Carretero A.,
Fernndez-Gutirrez A., Lercker G., Molecules 12, 1679, (2007)

39

P.11
ESTRAZIONE DEL PARTENOLIDE CON ESTRATTORE NAVIGLIO
Eleonora Bafile1, Alessandra Di Sante1, Umberto Marini1, Manuel Sergi2
1

Sintal Dietetics - Laboratorio Chimico, zona Industriale via tevere, 18, 64020 Teramo, Italy

Facolt di Bioscienze e Tecnologie Agroalimentari ad Ambientali - Universit degli Studi di Teramo,

Via Tevere, 64020 Teramo, Italy

Nel campo degli integratori alimentari molto importante avere a disposizione delle materie
prime, estratti secchi in particolare, correttamente identificati e titolati.
Lestratto secco di partenio da tempo utilizzato nel trattamento delle infiammazioni [1,2] di
vario genere e per tale motivo in campo nutraceutico ha da tempo una elevata importanza. Il
mercato offre ad oggi, estratti secchi il cui titolo trovato spesso non corrisponde a quanto
dichiarato.
Lo scopo del nostro lavoro ottenere estratti sicuri e salubri con una alta percentuale di marker
farmacologico evitando luso delle classiche tecniche estrattive menzionate in Farmacopea ma
utilizzando lestrattore Naviglio. Lobiettivo stato raggiunto grazie ad un accurato studio sia di
parametri puramente estrattivi che dei parametri connessi con la tecnica.
Tutte le analisi quali-quantitative sono state eseguite in HPLC-DAD [3] utilizzando una colonna
Gemini C18 lavorando in condizione isocratica con fase mobile composta da acetonitrile ed
acqua.
Bibliografia:
[1]Summer H. et al., J. Pharmacy and Pharmacology 44, 737, (1992)
[2]Jain MK. and Jahgirdar D.V., Biochimica et Biophysica Acta 814, 319, (1985)
[3]Joseph ZiQi Zhou et al., J. Agric. Food Chem. 47, 1018, (1999)

40

P.12
ACCOPPIAMENTO ON-LINE DI UN SISTEMA LC-MDGC-PREP PER L'ISOLAMENTO
DI COMPOSTI PURI
Sebastiano Pant1, Danilo Sciarrone1, Peter Q. Tranchida1, Luigi Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

Viale Annunziata, 98168 Messina, Italy

2 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via Alvaro del Portillo,

21, 00128 Roma, Italy

La gas cromatografia preparativa (Prep-GC) rappresenta una valida alternativa per lisolamento
di composti, sia conosciuti che incogniti, da poter correttamente identificare.
Le colonne wide-bore (0.53 mm I.D.) sono le pi usate in prep-GC per la loro elevata sample
capacity, tuttavia, presentano una limitata efficienza che da luogo a picchi scodati e solitamente
coeluiti. Una soluzione a questi inconvenienti liniezione di basse quantit di campione,
operazione che allunga di molto il tempo necessario alla raccolta di quantit accettabbili (mg)
di composto puro per una successiva identificazione mediante NMR, GC-FTIR, spettrometria
di massa.
La cromatografia multidimenaionale heart cutting (MDGC) rappresenta, invece, la reale
soluzione al problema, permettendo la separazione di composti target, grazie allutilizzo di pi
colonne accoppiate in serie. I sistemi MDGC che utilizzano valvole di tipo Deans switch sono
stati recentemente impiegati in applicazioni di prep-GC per la raccolta, in tempi brevi, di elevate
quantit di composti (mg) aventi unabbondanza relativa compresa tra 10 al 30 %.
Tuttavia, per la raccolta di composti puri presenti in concentrazioni 10 %, questo sistema
non garantisce le stesse performance in termini temporali, a causa dell'elevato numero
d'iniezioni necessarie per l'ottenimento delle stesse quantit di composto.
A tal fine, laccoppiamento on-line di un sistema LC caratterizzato da una maggiore sample
capacity, ha reso possibile liniezione di quantit di campione pi elevate riducendo il tempo
di raccolta di composti presenti in bassa concentrazione.
Il sistema costituito da un LC munito di una colonna operante in fase normale, necessaria
per una pre-separazione del campione in famiglie chimiche, di un sistema di trasferimento delle
stesse dall LC ad un iniettore per grandi volumi di campione (LVI) e quindi di un sistema MDGCprep gi descritto in precedenza.

41

P.13
GASCROMATOGRAFIA MULTIDIMENSIONALE COMPREHENSIVE CON MODULAZIONE
A FLUSSO: SVILUPPO DI UN NUOVO APPROCCIO ANALITICO
Flavio A. Franchina1, Peter Q. Tranchida1, Paola Dugo1,2, Luigi Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

viale Annunziata, 98168 Messina, Italy

2 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via lvaro del Portillo,

00128 Rome, Italy

Lo sviluppo di modulatori a flusso in tecniche gascromatografiche multidimensionali

comprehensive (FM-GCGC) stato, ad oggi, principalmente limitato dalla generazione di
elevati flussi (> 20 mL/min) in seconda dimensione. Questa condizione, ormai pienamente
accettata sin dalle prime applicazioni in quanto necessaria per una efficiente modulazione, ha
finora frenato anche lutilizzo di rivelatori MS. Una soluzione potrebbe essere la ripartizione del
flusso in uscita dal modulatore tra il detector e uno scarico, che non risulterebbe appropriata
per analisi di campioni poco concentrati o per lanalisi in tracce di analiti target, a causa
dellinevitabile abbassamento della sensibilit.
Lo studio presentato focalizzato in particolare sulla descrizione di un nuovo modello analitico
per lo sviluppo di metodi GCGC con modulatori a flusso. Questo nuovo approccio, che permette
lutilizzo di flussi di gas inferiori ( 7-8 ml/min), senza la necessit di ripartizione del flusso,
mantenendo efficiente il processo di modulazione, consiste in una rivalutazione delle fasi di
accumulo e re-iniezione. Con lutilizzo di flussi inferiori, non solo la separazione si mantenuta
pi efficiente in seconda dimensione, ma soprattutto si sono ridotti i problemi legati
allaccoppiamento con la spettrometria di massa (cio sensibilit e capacit delle pompe rotative
di mantenere condizioni di alto vuoto). Il modello proposto infine dimostrato su fragranze e
campioni reali di origine animale.

42

P.14
SVILUPPO DI UN METODO MULTIDIMENSIONALE LCxLC PER LANALISI DI
METABOLITI IN Saccharum officinarum L.
Gabriel Mazzi Leme1,2, Francesco Cacciola3,4, Paola Donato5,1, Alberto J. Cavalheiro2, Paola
Dugo1,5, Luigi Mondello1,5
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

Viale Annunziata, 98121 Messina, Italy

2

Chemistry Institute - UNESP - Univ. Estadual Paulista, Rua Prof. Francisco Degni 55, 14800-060
Araraquara, Brazil
3

Dipartimento di Scienze dell'Ambiente, della Sicurezza, del Territorio, degli Alimenti e della Salute
- University of Messina, Viale F. Stagno d'Alcontres 31, 98166 Messina, Italy
4

Chromaleont s.r.l. A start-up of the University of Messina, c/o Dipartimento di Scienze del Farmaco
e dei Prodotti della Salute, Viale Annunziata, 98168 Messina, Italy
5 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via lvaro del Portillo,

21, 00128 Roma, Italy

Negli ultimi anni lo studio dei prodotti naturali ha ricevuto una sempre pi crescente attenzione
portando allo sviluppo di metodiche analitiche per la separazione e rilevazione simultanea di
molti costituenti in miscele complesse. Attualmente analisi non-targeted vengono applicate
allottenimento di impronte metaboliche, controllo di qualit e pi recentemente in studi di
metabolomica, dove lo sviluppo e l'applicazione di metodi analitici robusti sono sempre pi
necessari. In questo contesto, la cromatografia liquida bidimensionale comprehensive (LC
LC) appare come una tecnica promettente e il suo utilizzo diventa necessario nel caso di analisi
di campioni complessi. Questo contributo propone lo sviluppo di un metodo LCLC per l'analisi
di un estratto di Saccharum officinarum L., basato sullimpiego di una colonna ciano e di una
C18, rispettivamente in prima e seconda dimensione. Tale metodo pu essere utile per il
confronto di diverse cultivar allo scopo di migliorarne il patrimonio genetico.

43

P.15
TARGET AND UNTARGET MULTIRESIDUAL ANALYSIS OF PESTICIDES IN FOODS
USING SIMULTANEOUS SCAN-MRM GC-MS/MS ACQUISITION
Giuseppe Scollo, Monica Lusardi
Shimadzu Italia, Via G.B.Cassinis 7, 20139 Milano, Italy

This application demonstrate the capability of Shimadzu GCMS-TQ8030 triple quadrupole to

investigate for target and untarget compounds from vegetables extracts: 260 pesticides were
analyzed, acquiring the data using the simultaneous scan/MRM mode acquisition in the same
chromatographic run.
For the evaluation, analytical standards (0.001 mg/L to 0.1 mg/L) were used, as well as samples
(0.01 mg/L) extracted with the QuEChERS method, and then adding pesticides to the obtained
solution.
The Pesticides Shimadzu Database, was used to easily create an MRM acquisition table. This
data base includes 430 compounds, including name, CAS number, Linear Retention Index
(LRI), multiple MRM transitions and collision energy.
The samples were introduced by OPTIC 4 PTV, an injector that allow to use temperature ramps
up to 3600 C/minutes even for large volume injections.
Using scan/MRM mode acquisition, target pesticides were quantified by creating calibration
curves from MRM data, furthermore, the simultaneously acquisition of scan data allow the
confirmation of target compounds from the integer mass spectra and the identification of untarget
compounds using library search and LRI for a complete sample characterization.

44

P.16
MICRO FLOW UHPLC-MS/MS IN PESTICIDE ANALYSIS OF INFANT FOODS
David Baker1, Stefano Zaza2, Neil Loftus1, Simon Hird3
1

Shimadzu MSO, Trafford Wharf Road, M17 1GP Manchester, United Kingdom

Shimadzu Italia Srl, Via G.B. Cassinis, 7, 20139 Milano, Italy

The Food and Environment Research Agency, Sand Hutton, YO41 1LZ York, United Kingdom

A micro-flow UHPLC-MS method has been developed and evaluated for routine pesticide
monitoring.
In todays global marketplace, as foods are produced and distributed throughout the world, food
quality and food safety has become of increasing concern for consumers, governments and
producers. Food safety with regards to infant food is of the upmost importance; however, it is
also recognised as a challenging matrix to analyse due to the low maximum residue limit (MRL)
of 10 g/kg that is applied to all pesticides. We explore the benefits of a developed micro flowUHPLC method for routine LC-MS/MS pesticide analysis of infant foods.
Infant food extracts were supplied by the Food and Environment Agency, UK, following
established QuEChERS protocols. A total of 130 pesticides were analysed using a Shimadzu
LCMS-8040 system with micro flow UHPLC. Data was acquired in the SRM mode using
electrospray in both positive and negative ion mode.
The developed micro flow UHPLC methodology achieved the required MRL of 0.01 mg/kg for
all 130 pesticides.
Initial validation data displayed excellent linearity for all compounds, low intra- and inter-day
precision, no observed carryover, and good peak area and retention time stability over 48 hours.
Precise data was acquired with the use of low pause and dwell times (1 msec. and 3msec.
respectively)
Micro flow analysis was successfully carried out on a UHPLC system capable of both
conventional higher flow rates and lower micro flow rates.
Micro flow LC is a possible alternative to conventional flow LC if extra sensitivity is needed or
reduction in solvent consumption is required.

45

P.17
SELECTIVE EXTRACTION OF ORGANOPHOSPHATE PESTICIDES BY MEANS OF
COMPUTATIONAL DESIGNED BIOMIMETIC LIGANDS
Manuel Sergi, Valentina Lanzone, Marcello Mascini, Flavio Della Pelle, Dario Compagnone
Faculty of Bioscience and Technology for Food, Agriculture and Environment - University of Teramo,
Via C. Lerici 1, Mosciano S.Angelo, 64023 Teramo, Italy

Organophosphorus compounds have come into widespread use in agriculture, since they show
low environmental persistence; nevertheless, they exert a high acute toxicity. The principal
effect of these compounds is the inhibition of the enzyme acetylcholinesterase (AChE), which
is essential for terminating the action of the neurotransmitter acetylcholine (ACh). In pesticides
detection, sample preparation is a critical step and is a key factor in determining the success
of analysis. Solid phase extraction (SPE) is commonly used for the clean-up of complex samples.
Traditional SPE sorbents range from reverse phases to ion exchange and polymeric materials
[1-3]. These materials are not selective and can result in the co-elution of interfering compounds
with similar polarity leading to matrix effect, that can affect the reliability of the analytical method.
For these reasons, three different hexapeptides were computationally designed and tested as
selective SPE sorbent for chloropyriphos e pirimiphos-methyl. The approach of this work relies
on the design and development of artificial oligopeptides mimicking the binding site of
acetylcholinerase (AChE) [4]. The hexapeptide-pesticides complex binding scores were
compared with the SPE results. Before the SPE procedure set-up, detection conditions were
optimized with standard solutions. The extraction procedure for SPE was also optimized
considering volume loading, pH effect, washing and elution conditions and interferences. The
resulting data were presented and discussed.
References
[1] Vidal L.et al., Anal. Chim. Acta 715, 19, (2012)
[2] Toth B. et al., Top. Curr. Chem. 325, 267, (2012)
[3] Ramautar R. et al., Electrophoresis 33, 243, (2012)
[4] Mascini M. et al., Anal. Chem. 80, 9150, (2008)

46

P.18
DETERMINATION OF ALKYLPYRAZINES IN COCOA LIQUORS BY HS-SPME-GC-MS
Francisco Ardini, Marina Di Carro, Emanuele Magi
Department of Chemistry and Industrial Chemistry - University of Genoa, Via Dodecaneso 31, 16146
Genoa, Italy

The rich aroma of chocolate is due to a very complex mixture of compounds, whose composition
depends both on the cocoa bean genotype and the several processes occurring in the chocolate
production (fermentation, drying, roasting and conching).[1]
Alkylpyrazines are a very important class of substances produced by the Maillard reactions
during the chocolate roasting. For this reason they can be used as index compounds for the
roasting process [2,3] and could be one of the useful parameters for the optimization of the
whole chocolate production.
In this work, a headspace solid-phase microextraction gas chromatography mass spectrometry
(HS-SPME-GC-MS) method was developed for the study of 2,3-dimethylpyrazine, 2,5dimethylpyrazine, 2,3,5-trimethylpyrazine and tetramethylpyrazine in different cocoa liquor
samples. After some preliminary experiments, three HS-SPME parameters (equilibration time,
equilibration temperature and extraction time) were optimized by means of a Central Composite
Design. Results showed that the optimal conditions for the analysis were minimum equilibration
time and maximum equilibration temperature and extraction time. The developed procedure
was finally applied to Ecuadorian cocoa liquors roasted at different temperatures, obtaining a
relative distribution pattern of the considered alkylpyrazines in the samples.
References
[1] Afoakwa E., Chocolate science and technology, Wiley-Blackwell,Oxford, (2010)
[2] Jinap S. et al., J. Sci. Food Agr. 77, 441, (1998)
[3] Hashim L. and Chaveron H., Food Res. Int. 27, 537, (1994)

47

P.19
UTILIZZO DI UN SISTEMA LC PREPARATIVO MULTIDIMENSIONALE PER
LISOLAMENTO DI MOLECOLE BIOLOGICAMENTE ATTIVE DA MATRICI NATURALI
Marina Russo1, Veronica Inferrera1, Francesca Rigano1, Paola Dugo1,2, Luigi Mondello1,2
1 Dipartimento di Scienze del Farmaco e dei Prodotti per la Salute (SCIFAR) - University of Messina,

Viale Annunziata, 98168 Messina, Italy

2 Centro Integrato di Ricerca (C.I.R.) - University Campus Bio-Medico of Rome, Via lvaro del Portillo,

21, 00128 Rome, Italy

Lo scopo di questo lavoro stato il recupero di molecole biologicamente attive da matrici di

origine naturale mediante lutilizzo di un sistema HPLC preparativo multidimensionale (LC-LC).
Il sistema prevede limpiego di due colonne HPLC preparative, impaccate con differenti fasi
stazionarie e quindi differente meccanismo di separazione.
Un succo concentrato di bergamotto e il residuo non volatile di un olio essenziale di mandarino
estratto a freddo sono stati sottoposti ad analisi multidimensionali LC preparativa, frazionati
sulla prima colonna e, le frazioni contenenti le molecole biologicamente attive sono state raccolte
mediante un autocollettore. Le frazioni di interesse sono state sottoposte ad una successiva
separazione sulla seconda colonna HPLC preparativa, che ha garantito lisolamento del
composto puro. Il sistema automatizzato di raccolta funziona in base al segnale generato da
uno spettrometro di massa accoppiato al LC-LC preparativo: lautocollettore inizier la raccolta
in base allintensit del segnale che le molecole bioattive genereranno alla massa.
Questa strumentazione di nuova concezione ha portato allisolamento di molecole
biologicamente attive (flavonoidi e polimetossiflavoni) da un succo concentrato di bergamotto
e un residuo non volatile di un olio essenziale di mandarino estratto a freddo.
Ringraziamenti
The Project was funded by the Italian Ministry for the University and Research (MIUR) within
the National Operative Project Hi-Life Health Products from the industry of foods. Project ID:
PON0101499.

48

P.20
ACCURATE IDENTIFICATION AND PROFILING OF ELLAGITANNINS IN
STRAWBERRIES AND WOODLAND STRAWBERRIES: THE INFLUENCE OF
CULTIVAR ON THE CONCENTRATION AND COMPOSITION OF ELLAGITANNINS
Mattia Gasperotti1, Domenico Masuero1, Graziano Guella2, Luisa Palmieri1, Paolo Martinatti1,
Fulvio Mattivi1, Urska Vrhovsek1
1

Food Quality and Nutrition Department - Research and Innovation Centre - Fondazione E. Mach,
via E. Mach 2, 38010 San Michele all'Adige, Italy
2

Department of Physics - University of Trento, via Sommarive 14, 38100 Trento, Italy

Strawberries (Fragaria ananassa Duch.) are one of the most consumed berries. Although the
composition of strawberry fruit has been extensively studied, the detailed characterisation of
ellagitannins and especially the most abundant one agrimoniin has been only recently univocally
identified [1, 2]. Ellagitannins recently gained much attention in light of the experimental
evidence of their anticancer, antiproliferative, and antibacterial activities. Furthermore
agrimoniin is a known bioactive compound.
The establishment of an HPLC protocol for the HPLC separation of the ellagitannins has been
already published for the study of the ellagitannins in Rubus species [3]. The isolation and
characterisation of strawberry ellagitannins and ellagic acid derivatives, allowed us their
accurate identification and quantification of in 6 different varieties of strawberry and in 2 woodland
strawberry. The separation of 23 ellagitannins and 3 ellagic acid conjugates in the strawberry
extracts was performed in Fragaria species by HPLC-HDMS. Differences on the composition
and on the concentration of ellagitannins were observed in the ellagitannins profiling among
the cultivars [2].
Woodland strawberry was the richest in terms of absolute concentration of ellagitannins and
as consequence these data confirm that the wild species are the most interesting in term of
nutritional relevance. We observed major qualitative and quantitative differences in the amount
and profile of ellagitannins and ellagic acid conjugates in Fragaria x ananassa and Fragaria
vesca species, as well as several qualitative differences in some minor ellagitannins in the
Fragaria x ananassa cultivars. This information suggests that the ellagitannin profile could also
be interesting for characterising cultivars. Among the fruits which contain ellagitannins,
strawberries are the most widely consumed, and agrimoniin is therefore suggested to be one
of the most widely present ellagitannins in the human diet.
References:
[1] Vrhovsek U. et al., J. Agric. Food Chem. 60, 2507, (2012)
[2] Gasperotti M. et al., J. Agric. Food Chem. 61, 8597, (2013)
[3] Gasperotti M. et al., J. Agric. Food Chem. 58, 4602, (2010)

49

P.21
ENHANCED AROMA PROFILING BY GCTOF MS WITH SELECTIVE SOFT EI
IONISATION
L. McGregor1, N. Bukowski1, J. Blanch1, B. Green1, S. Smith1, Eligio Sebastiani2, L. Calamai3
1

Markes International, Gwaun Elai Medi-Science Campus, CF72 8XL Llantrisant, RCT, United
Kingdom
2

SRA INSTRUMENTS, Viale Assunta 101, 20063 Cernusco sul Naviglio Milano, Italy

CISM Mass Spectrometry Center - Universit degli Studi di Firenze, Via Ugo Schiff, 50019 Sesto
Fiorentino, Italy

Aroma profiles, such as those for wine, contain a wide variety of components at a range of
concentrations. Detection and identification of important keynote compounds with a low odour
threshold and compounds responsible for off-odours is a challenging prospect.
Gas chromatography coupled with time-of-flight mass spectrometry (GCTOF MS) is an ideal
choice for such analyses. Fast acquisition speeds, full-range spectra and low detection limits
allow trace components, including adulterants, to be identified even within the most challenging
of matrices. Novel data-mining software for the pairwise comparison of complex chromatograms
is described, allowing such minor differences to be readily distinguished.
This poster demonstrates the use of GCTOF MS for aroma profiling of wine, with revolutionary
variable-energy electron ionisation to provide enhanced molecular ions and reduced
fragmentation to aid speciation of challenging compounds.
The new technology offering variable-energy electron ionisation, and the comparative software,
will be available from January 2014. Visit Markes website to be kept up to date with the
introduction schedule and plans, or email enquiries@markes.com.

50

P.22
BRASSICA FRUTICULOSA CYR. AND B. INCANA TEN. (BRASSICACEAE) AS
MEDITERRANEAN TRADITIONAL WILD VEGETABLES : A VALUABLE SOURCE OF
BIOACTIVE COMPOUNDS
Giovanna Dima1, Antonella Verzera1, Gianluca Tripodi1, Concetta Condurso1, Salvatore Ragusa2
1

Dipartimento di Scienze Chimiche - University of Messina, Viale F.Stagno d'Alcontres, 31, 98166
Messina, Italy
2 Department of Health Sciences - University Magna Graecia, Contrada Mula, 88100 Catanzaro, Italy

Two species of Brassicaceae (Crucifer) family, used and appreciated as traditional wild
vegetables, including Brassica fruticulosa Cyr. and Brassica incana Ten., were examined as
potential source of bioactive volatile compounds. The volatile constituents released by the
chopped leaves and roots were extracted and analysed by solid-phase microextraction - gas
chromatography - mass spectrometry (SPME-GC-MS). A large number of volatile constituents
were identified: alcohols, aldehydes, esters, acids, ketones, terpenes, C13-norisoprenoides and
sulfur compounds. Volatiles included isothiocyanates with a well known anticancer activity; the
largest amount resulted in the roots with 3-butenyl isothiocyanate the most represented in both
species, of great interest also the good amount revealed in the leaves of Brassica fruticulosa
Cyr. The revaluation of these plants, a vegetable source of high antioxidant power, will be
interesting for consumer health by the production of new commercial herbal products and/or
dietary supplements of high quality and low cost.

51

Index

Abelmoschi Maria Luisa 32 (P.03)

Ancillotti Claudia 27 (L.26)
Ardini Francisco 32 (P.03), 47 (P.18)
Aturki Zeineb 15 (L.14)
Ayyad Ziad 39 (P.10)
Bafile Eleonora 40 (P.11)
Baker David 45 (P.16)
Balci M. 22 (L.21)
Barbieri Sara 38 (P.09), 39 (P.10)
Barreca Salvatore 36 (P.07)
Beccaria Marco 17 (L.16)
Bendini Alessandra 38 (P.09), 39 (P.10)
Bicchi Carlo 11 (L.10)
Blanch J. 50 (P.21)
Bonaccorsi Ivana 33 (P.04)
Bruzzoniti Maria Concetta 13 (L.12)
Bukowski N. 50 (P.21)
Cacciola Francesco 17 (L.16), 43 (P.14)
Cagliero Cecilia 11 (L.10)
Calamai L. 50 (P.21)
Campiglia Pietro 30 (P.01)
Campone Luca 35 (P.06)
Cappiello Achille 24 (L.23)
Capriotti Anna Laura 2 (L.01)
Capriotti Fabiana 24 (L.23)
Carabetta Sonia 35 (P.06)
Caruso Giuseppe 18 (L.17)
Cataldi Tommaso 19 (L.18)
Cavalheiro Alberto 43 (P.14)
Cavaliere Chiara 18 (L.17)
Cavalli Silvano 13 (L.12)
Cavazzini Alberto 21 (L.20)
Cellar Nick 24 (L.23)
Cerichelli Giorgio 16 (L.15)
Checchini Leonardo 27 (L.26)
Ciceri Elena 14 (L.13)
Ciofi Lorenzo 27 (L.26)
Ciogli Alessia 5 (L.04)
Colapicchioni Valentina 2 (L.01)
Compagnone Dario 46 (P.17)
Condurso Concetta 26 (L.25), 51 (P.22)

52

Index

Cordero Chiara 11 (L.10)

Corradini Danilo 8 (L.07), 25 (L.24)
Costa Rosaria 31 (P.02)
D'Egidio Maria Grazia 8 (L.07), 25 (L.24)
D'Orazio Giovanni 16 (L.15)
De Carlo Rosa 13 (L.12)
De Grazia Selenia 31 (P.02)
Del Bubba Massimo 27 (L.26), 28 (L.27)
Della Pelle Flavio 46 (P.17)
Di Carro Marina 32 (P.03), 47 (P.18)
Di Lecce Giuseppe 38 (P.09)
Di Sante Alessandra 40 (P.11)
Dima Giovanna 26 (L.25), 51 (P.22)
Donato Paola 17 (L.16), 43 (P.14)
Dondi Francesco 21 (L.20)
Drbov Lucie 20 (L.19)
Dugo Giacomo 29 (L.28)
Dugo Giovanni 1 (PL.01), 37 (P.08)
Dugo Laura 15 (L.14)
Dugo Paola 1 (PL.01), 17 (L.16), 23 (L.22), 31 (P.02), 33 (P.04), 42 (P.13), 43 (P.14), 48 (P.19)
Famiglini Giorgio 24 (L.23)
Fanali Chiara 15 (L.14)
Fanali Salvatore 16 (L.15)
Fede Maria Rita 29 (L.28)
Foglia Patrizia 2 (L.01)
Franchina Flavio 42 (P.13)
Frega Natale 9 (L.08)
Frusteri Francesco 12 (L.11)
Furlanetto Sandra 27 (L.26), 28 (L.27)
Gagliardi Riccardo 9 (L.08)
Gallina Toschi Tullia 38 (P.09), 39 (P.10)
Gallo Sergio 3 (L.02)
Gasparrini Francesco 5 (L.04)
Gasperotti Mattia 4 (L.03), 49 (P.20)
Granafei Sara 19 (L.18)
Grasso Elisa 31 (P.02), 34 (P.05)
Green B. 50 (P.21)
Griglione Alessandra 11 (L.10)
Guella Graziano 49 (P.20)
Gursu G. 22 (L.21)
Hajslov Jana 20 (L.19)

53

Index

Hird Simon 45 (P.16)

Inferrera Veronica 48 (P.19)
Italiano Giuseppe 12 (L.11)
Kalachov Kamila 20 (L.19)
Kotoni Dorina 5 (L.04)
Kovalczuk Toms 20 (L.19)
Lagana` Aldo 21 (L.20)
Lanzone Valentina 46 (P.17)
Liberto Erica 11 (L.10)
Loftus Neil 45 (P.16)
Losito Ilario 19 (L.18)
Lucci Paolo 9 (L.08)
Lucini Stefano Maria 6 (L.05)
Lusardi Monica 6 (L.05), 44 (P.15)
Magi Emanuele 32 (P.03), 47 (P.18)
Marchetti Nicola 21 (L.20)
Marini Umberto 40 (P.11)
Martinatti Paolo 49 (P.20)
Martini Daniela 8 (L.07), 25 (L.24)
Mascini Marcello 46 (P.17)
Mastrocinque Raffaella 30 (P.01)
Masuero Domenico 4 (L.03), 49 (P.20)
Mattivi Fulvio 4 (L.03), 49 (P.20)
Mazzi Leme Gabriel 43 (P.14)
McGregor L. 50 (P.21)
Mondello Luigi 7 (L.06), 17 (L.16), 23 (L.22), 31 (P.02), 33 (P.04), 34 (P.05), 37 (P.08), 41
(P.12), 42 (P.13), 43 (P.14), 48 (P.19)
Nicoletti Isabella 8 (L.07), 25 (L.24)
Nicolotti Luca 11 (L.10)
Orecchio Santino 36 (P.07)
Orlandini Serena 28 (L.27)
Pace Andrea 36 (P.07)
Pacetti Deborah 9 (L.08)
Pagano Francesco 30 (P.01)
Pagano Imma 35 (P.06)
Palella Alessandra 12 (L.11)
Palma Pierangela 24 (L.23)
Palmieri Luisa 49 (P.20)
Palmisano Francesco 19 (L.18)
Pant Sebastiano 41 (P.12)
Pasquini Benedetta 28 (L.27)
Pepe Giacomo 30 (P.01)
54

Index

Perissi Andrea 10 (L.09)

Piccinelli Anna Lisa 35 (P.06)
Pierri Giuseppe 5 (L.04)
Pinzauti Sergio 28 (L.27)
Piovesana Susy 18 (L.17)
Ragonese Carla 34 (P.05)
Ragusa Salvatore 51 (P.22)
Rastrelli Luca 35 (P.06)
Rigano Francesca 7 (L.06), 48 (P.19)
Rivoira Luca 13 (L.12)
Rocchi Silvia 16 (L.15)
Rocco Anna 15 (L.14)
Romano Loredana 12 (L.11)
Rubiolo Patrizia 11 (L.10)
Russo Maria Teresa 35 (P.06)
Russo Marina 7 (L.06), 31 (P.02), 48 (P.19)
Saitta Marcello 29 (L.28)
Salivo Simona 23 (L.22)
Sangens Jos 20 (L.19)
Sarzanini Corrado 13 (L.12)
Schipilliti Luisa 33 (P.04)
Sciarrone Danilo 7 (L.06), 34 (P.05), 41 (P.12)
Scollo Giuseppe 44 (P.15)
Sebastiani Eligio 22 (L.21), 50 (P.21)
Sergi Manuel 40 (P.11), 46 (P.17)
Sgorbini Barbara 11 (L.10)
Simone Patrizia 5 (L.04)
Smith S. 50 (P.21)
Sommella Eduardo 30 (P.01)
Spadaro Lorenzo 12 (L.11)
Stampachiacchiere Serena 2 (L.01)
Sullini Giuseppe 17 (L.16)
Taddei Federica 8 (L.07), 25 (L.24)
Termopoli Veronica 24 (L.23)
Tesini Federica 38 (P.09), 39 (P.10)
Tranchida Peter 42 (P.13), 7 (L.06), 23 (L.22), 37 (P.08), 41 (P.12)
Tripodi Gianluca 26 (L.25), 51 (P.22)
Valenza Gabriele 19 (L.18)
Valli Enrico 38 (P.09), 39 (P.10)
Ventre Giovanni 30 (P.01)
Verzera Antonella 26 (L.25), 51 (P.22)

55

Index

Villani Claudio 5 (L.04)

Vrhovsek Urska 4 (L.03), 49 (P.20)
Zaza Stefano 45 (P.16)
Zenezini Chiozzi Riccardo 18 (L.17)
Zoccali Mariosimone 37 (P.08)
Zrostlkov Jitka 20 (L.19)

56