3. FOOD ChEMISTRy & bIOTEChNOLOGy 3.1. Lectures

Chem. Listy, 102, s265–s1311 (2008) Food Chemistry & Biotechnology
P06 MONITORING OF ACRyLAMIDE LEVELS
uSING LC-MS/MS: MALTS AND bEERS
VEROnIKA BARTáČKOVá, LUKአVáCLAVíK,
KATEřInA RIDDELOVá, VLADIMíR KOCOUREK and
JAnA HAJŠLOVá
Institute of Chemical Technology Prague,
Department of Food Chemistry and Analysis,
Technicka 5, Prague 6 – Dejvice, 166 28,
veronika.bartackova@vscht.cz
Introduction
Acrylamide (AA), a hazardous chemical, is classified as
a probable human carcinogen (IARC, 1994) 1 . AA is formed
during Maillard reaction in starch-rich foods. Its occurrence
in human diet was for the first time reported by Swedish
scientists in 2002 2 .
Alike many other heat processed foods prepared from
cereals, malts may contain AA, originated from sugar and
asparagin contained in barley. Besides of other factors, AA
levels depend on the temperature and the time employed for
treatment.
In our study, various types of malts, commonly used as
additives (colorants, aroma donors etc.) in bakery and/or in
brewing industry were examined for AA levels using LC-
MS/MS.
In the next phase of our experiments the transfer of this
processing contaminant into the beer was studied. The analytical
procedure used for the analysis consists of cleanup step
employing solid-phase extraction (SPE) followed by liquid
chromatography coupled to tandem mass spectrometry (LC–
MS/MS). A wide range of bottled beers obtained at the Czech
retail market was examined within this monitoring study.
Experimental
C h e m i c h a l s a n d M a t e r i a l s
AA (purity 99,5 %) was from Sigma/Aldrich/Fluka
(Switzerland). 13 C3 -AA (isotopic purity ≥ 99 %) was
purchased from CIL (USA). Magnesium sulphate was from
Sigma/Aldrich/Fluka (Switzerland). Sodium chloride was
from Penta (Czech Republic). Aluminium oxide (basic) was
from Merck (Germany). All organic solvents were of HPLC
grade quality.
Calibration standard solutions were prepared in water by
diluting stock standard solutions in concentration range
1–250 ng ml –1 with fixed amount of 13 C3 –AA –
100 ng ml –1 .
Isolute Multimode ® (6 ml, 500 mg) and Isolute EnV+ ®
(6 ml, 500 mg) SPE cartridges were purchased from IST
(U.K.).
S a m p l e s
Czech special roasted malts (4 from barley and 1 from
wheat) were employed for the study. In addition to the malt
samples, intermediates collected during processing (that was
s585
conducted under different time and temperature conditions)
were examined.
The German set of samples included 12 malts (brew,
special, caramel, wheat and rye), used in beer industry.
Thirty beer samples representing various typical brands
were purchased from the Czech retail market.
M a l t S a m p l e P r e p a r a t i o n
The key step of this method is the transfer of AA from
the primary aqueous extract into acetonitrile forced by added
salts (MgSO 4 and naCl); separation of aqueous and organic
phases is induced 3 . Most matrix interferences are then removed
from organic phase by dispersive solid phase extraction
(MgSO 4 and basic Al 2 O 3 are used for this purpose). Acetonitrile
is evaporated under a gentle stream of nitrogen and
solvent is exchanged to water.
B e e r S a m p l e P r e p a r a t i o n
After decarbonization the beer sample undergoes cleanup
with two subsequent SPE cartridges. Final extract is in
60% methanol in water (v/v solution). The methanol is evaporated
by a gentle stream of nitrogen 4 .
A n a l y s i s o f A A b y L C - M S / M S
LC-MS/MS measurements were accomplished with the
use of a separation module Alliance 2695 (Waters, USA)
interfaced to a Quattro Premier XE mass spectrometer
(Micromass, UK). Chromatographic separation was carried
out using an Atlantis T3 column (3 mm × 150 mm × 3 μm).
Isocratic elution of AA was achieved with mobile phase composed
of acetonitrile and water (3 : 97, v/v).
The mass spectrometer, equipped with pneumatically
assisted electrospray interface (ESI), was operated in a positive
ionization mode.
LOD´s and LOQ´s for malt samples were 10 µg kg –1 and
30 µg kg –1 , respectively. LOD´s and LOQ´s for beer samples
were 2 µg dm –3 and 5 µg dm –3 , respectively.
Results
AA levels in malt corresponded to the temperatures
employed for their production. In dark malts AA content was
relatively high, clearly due to advanced Maillard reaction
induced by heating process.
Table I
Czech malts
Malt AA (μg kg –1 )
Semi-finished light caramel 35
Light caramel 37
Semi-finished dark caramel 404
Dark caramel 243
Coloured 111
Czech 25
Wheat < 10