Analisis Kafein, Taurin, dan

Pemanis Buatan pada

Minuman Berenergi
• Multiple injections in a single experimental run (MISER) is a simplified
form of LC–MS that is well suited to fast analysis of the presence of a
given component within multiple samples
• We now report an investigation into the use of MISER LC–MS for the
analysis of caffeine, aspartame, taurine within soft drinks, and energy
drinks
EXPERIMENTAL
• Instrumentation
Reversed-phase high performance liquid chromatography (HPLC) experiments were
performed on an Agilent 1100 system. The Agilent stack comprised a G1312A binary
pump, a G1367A WPALS
autosampler, a G1315B diode-array detector, and a 6120 quadrupole LC–MS detector
with electrospray ionization in the positive mode. The system was controlled by
Chemstation software, with the flow
injection analysis (FIA) mode enabled.
• Preparation of Buffer Solutions
Solutions containing 2 mM ammonium formate in water (pH 3.5) and 2 Mm
ammonium formate in acetonitrile (pH 3.5): 12.6 g ammonium formate and 7.9 mL
formic acid were dissolved in 1 L of Millipore water. A 100-fold dilution of this stock
solution was performed in either pure water or a 90:10 acetonitrile–water mixture to
prepare the 2 mM solutions.
• HPLC–MS Conditions
HPLC separations were carried out on a 50 mm × 4.6 mm, 2.5-µm
XBridge Phenyl column by isocratic elution at a flow rate of 1 mL/min.
The LC eluents were 60% solvent A (2 mM ammonium formate in water,
pH 3.5) and 40% solvent B (2 mM ammonium formate in 90:10
acetonitrile– water, pH 3.5). The column and samples were maintained
at a temperature of 25 °C. The misergrams were obtained from
continuous sample injections (1 µL) every 1 min. The positive ion
electrospray ionization (ESI) parameters were as follows: skimmer, 45 V;
desolvation gas, nitrogen; temperature, 350 °C; and flow rate, 12 L/min.
The nebulizer was adjusted to 35 psig, and the fragmentor and the
capillary voltages were adjusted to 150 and 2500 V, respectively. G1969-
85000 ESI-L Low Concentration Tuning Mix (Agilent Technologies) was
used for tuning and calibration of the mass spectrometer
• Ion Suppression and Enhancement Between Target Analytes
Six sets of samples were prepared at 0.25 mg/mL. Set 1, aspartame and caffeine; set 2, caffeine;
set 3, caffeine and taurine; set 4, aspartame; set 5, aspartame and taurine; and set 6, taurine.
• All six samples were analyzed per duplicated using the standardized MISER LC–ESI(+)-MS method.
In addition, a faster MISER LC–ESI(+)-MS method was evaluated for comparison:
Column: Zorbax Stablebond SB-C18 (20 mm × 3.0 mm, 1.8 µm, Agilent);
temperature: 40 °C;
detection: selected ion monitoring (SIM) ESI-MS(+) at m/z = 195, 295, and 265 amu;
sample: 0.6-µL injection of each component solution in acetonitrile–water;
flow rate: 1 mL/min;
isocratic mobile phase: 10% eluent A: 2 mM ammonim formate in water (pH 3.5) and 90%
eluent B: 2 mM ammonium formate in 10:90 water–acetonitrile (pH 3.5).
Time between injections was 22 s.
• The extent
of ion suppression and ion enhancement effects are clearly visualized in the corresponding
misergrams.
RESULT & DISCUSSION
• Figure 3a shows the
results of such a method
development study,
clearly illustrating that
the shorter method
leads to a significant ion
suppression of caffeine
and taurine MS signal in
the presence of
aspartame, while the
longer 1-min injection
period is free from such
problems
• All samples were diluted 10 times with water, with
the exception of Red Line Xtreme and 5 Hour
Energy, which were diluted 100×. The resulting
misergrams show much higher caffeine content in
the energy drinks than in Coke and Diet Coke, with
5 Hour Energy showing a considerably higher
amount of caffeine (similar signal response as Rock
Star drinks, but the sample is 10 times more
diluted). Red Bull sugar free and Total Zero show
aspartame concentrations lower, but comparable
to Diet Coke. Rock Star shows almost no
aspartame, but checking the ingredients we found
sucralose, another artificial sweetener commonly
used in energy drink preparations. The taurine
signal response from Rock Star, Monster, and Red
Bull drinks were very similar at about 0.4 mg/mL,
but higher than AMP Boost Cherry. All of these
results are consistent with the caffeine,
aspartame, and taurine levels published in the
different literature sources
CONCLUSIONS
• The approximate levels of caffeine, aspartame, and taurine in soft
drinks and energy drinks can be easily measured by MISER LC–MS.
Considerable variation in these analyte levels is clearly visualized
among a variety of these beverages, with the energy drinks showing a
higher concentration of caffeine and taurine, and the diet soft drinks
containing a higher amount of artificial sweetener (aspartame).