Differential Mobility Spectrometry with the AB SCIEX SelexION™ Technology for 5500 Systems A New Dimension of Selectivity Mauro Aiello, Senior Product Manager
2 © 2011 AB SCIEX
AB SCIEX SelexION™ Technology
Differential Mobility Spectrometry with the SelexION™ Technology
for 5500 Systems delivers a new dimension of selectivity and
performance for any application requiring the separation of isobaric
species, isolation of challenging co-eluting contaminants and reduction
of high background noise.
3 © 2011 AB SCIEX
AB SCIEX SelexION™ Technology
• Robust hardware designed for ease of use
•Few minutes to install/remove
•No tools required
•No need to break vacuum
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QTRAP® 5500 System Ion Path with SelexION™ Technology
SelexION™ Technology ion path components
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How does SelexION™ Technology separate Ions?
Separation waveform (SV):
Radially displaces ions towards
one or the other electrode,
depending upon high and low
field mobility characteristics
Compensation voltage (COV):
Restores the trajectory for a
given ion to allow them to
transmit through the DMS device
and enter the mass
spectrometer
SV COV
To
MS Gas
flow
•Differential Mobility Spectrometry (DMS) is the term used for planar geometry
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Advantages of Planar DMS geometry
Short residence times
Rapid voltage changes for MRM operation
– MRM cycle times of 25 msec, (20 msec pause time)
– Fast LC support
Transparent Mode
– Allows all ions to be transmitted by turning off voltages
Minimal diffusion losses
Homogeneous electric fields within the DMS analyzer.
– Improved resolution at high voltages
Uniform conditions for the addition of chemical modifiers
– More on this to come…
Also:
– Scheduled MRM™ support
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Modified Transport Gases Improve Separations
•Liquid modifiers can be added to the curtain gas flow
•Improves separations
•More options for separation in difficult cases
11 compounds: methylhistamine, minoxidil, ephedrine, norfentanyl,
acyclovir, clenbuterol, tramadol, quinoxifen, pamaquin, fendiline,
buscopan.
1.0
0.8
0.6
0.4
0.2
0.0
-60 -40 -20 0
1.0
0.8
0.6
0.4
0.2
0.0
Norm
aliz
ed S
ignal
-60 -40 -20 0
CV (V)
Nitrogen Transport Gas
118 Td
7x improved separation
1.5% IPA
8 © 2011 AB SCIEX
Separations are Chemical in Nature
High field
Declustering
Mobility Increases
Low Field
Clustering-
Mobility Decreases
+
Krylov et al., Int. J. Mass Spectrom., 2009, 285, 149-156
High field
Declustering
Mobility Increases
Low Field
Clustering-
Mobility Decreases
Dynamic Cluster/Decluster Model
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Effects of Different Modifiers Gas phase LC?
1.0
0.8
0.6
0.4
0.2
0.0
Norm
aliz
ed S
ignal
-50 -40 -30 -20 -10 0
CV (V)
Isopropanol Separation
1.0
0.8
0.6
0.4
0.2
0.0
Norm
aliz
ed S
ignal
-50 -40 -30 -20 -10 0
CV (V)
Acetone Separation
1.0
0.8
0.6
0.4
0.2
0.0
Norm
aliz
ed S
ignal
-50 -40 -30 -20 -10 0
CV (V)
Acetonitrile Separation
Red: Ephedrine
Black: Acyclovir
Green: Norfentanyl
Blue: Clenbuterol
Purple: Imipramine
Brown: Diazepam
Pink: Quinoxifen
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Clenbuterol analysis from urine samples
LC-MSMS analysis of clenbuterol from urine samples is known to exhibit high degree of interferences in all major MRM transitions monitored.
These interferences also vary greatly in terms of complexity and intensity levels between subject samples.
Here we investigated the use of DMS to increase the selectivity of the LC-MSMS analysis
11 © 2011 AB SCIEX
Clenbuterol Spiked in Human Urine (dil. 1:1 prior to analysis) QTRAP® 5500 vs 5500 with SelexION™ Technology
MRM
276-203
MRM
276-168
MRM
276-132 QTRAP
5500
SelexION
Technology
on 5500
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Eliminating High Chemical Noise with SelexION™ Technology
Pentoxifylline ~20 times LOQ Improvement
10 pg/uL
QTRAP® 5500 With SelexION
QTRAP® 5500
13 © 2011 AB SCIEX
Eliminating High Chemical Noise with SelexION™ Technology
Pentoxifylline ~20 times LOQ Improvement
1 pg/uL
QTRAP® 5500 With SelexION
QTRAP® 5500
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Measuring Large Peptides BNP +6 Charge State
For some large endogenous peptides that don’t fragment well, monitoring intact by SIM can provide good sensitivity but is prone to interferences
DMS can significantly reduce the presence of a co-eluting nearly isobaric matrix peak, to improve detection limits
3.2 3.4 3.6 3.8
Time, min
0
2e4
4e4
Inte
nsity
Blank – DMS off
Blank – DMS on
128 pg/mL
640 pg/mL
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N
NNH
N
NN
O
OH
N
NN
OH
OHO
Regular QTRAP® 5500
With SelexION™ technology
MRM
70-43
MRM
158-70
MRM
128-70
1.0 2.0 3.0 Time, min
1.0 2.0 3.0 Time, min
1.705
Triazole other pesticide degradants in Grapefruit
Grape fruit extract fortified with conazole degradant products at 0.01mg/kg. Samples kindly supplied by R. Schoening, Bayer CropScience, Monheim, Germany, & J. Jasak,
Technical University, Institute of Food Chemistry, Dresden, Germany
Triazole acetic acid Triazole lactic acid Triazole
1.0 2.0 3.0 Time, min
1.714
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Peak area Buprenorphine, CV% = 6.8%
Peak area Clenbuterol, CV% = 5.6%
IS corrected Peak area Buprenorphine, CV% = 1.7%
IS corrected Peak area Clenbuterol, CV% = 1.6%
QTRAP® 5500 System with SelexION™ Technology
Robustness for Demanding Applications in regulated labs
1000 plasma samples injected over 66 hours
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Separation of Isobaric Compounds (m/z 309)
1.0
0.8
0.6
0.4
0.2
0.0
No
rma
lize
d S
ign
al
-50 -40 -30 -20 -10 0
CV (V)
Bestatin
Warfarin Phenylbutazone
Quinoxyfen
Benoxinate
SelexION™ Technology Applications
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Separation of Isomers - Pseudoephedrine/Ephedrine
Pseudoephedrine Ephedrine
•Separation of pseudoephedrine and ephedrine with
SelexION™
•Indistinguishable by MS or MS/MS
SelexION™ Technology Applications
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Steroid Interference Removal with SelexION™ Technology
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Experiment Conditions
5 subjects each of male and female serum samples obtained from a customer laboratory
STD was prepared in pooled children under age 7 serum samples.
Two sample preparation methods:
– LLE with 90/10 hexane/ ethyl acetate ( 200 uL serum; 1 mL solvent; recon 150 uL with 50/50 MeOH/H2O)
– PPT with ACN (200 uL serum; 600 ACN; dry down and recon 150 uL with 50/50 MeOH/H2O)
Column: Kinetex 2.6 u 50x2.1
mm; run time: 7 min; flow rate
0.3 mL/min
MPA: Water+0.1% FA;
MPB: MeOH+0.1% FA
ESI: positive
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Testosterone Female #3 Serum Sample: LLE
DMS OFF/
289/109 DMS OFF/
289/97
DMS ON/
289/109 DMS ON/
289/97
Interference
Ion ratio~21 Peak of interested Interference
Peak of interested
Ion Ratio=1.28
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Testosterone Female #3 Serum Sample: PPT
DMS OFF/
289/109
DMS OFF/
289/97
DMS ON/
289/109
DMS ON/
289/97
Peak of interested Peak of interested
@ RT 3.37 min
23 © 2011 AB SCIEX
STD Curve in Serum Sample with DMS on (PPT)
4 female
subjects
Sample Neat (n=28)
Serum Sample
PPT (n=25)
Serum Sample
LLE (n=25)
Average Ion Ratio
(97/109) 1.23 1.28 1.24
STDEV 0.04 0.07 0.04
%CV 3.44 5.47 3.12
Accuracy 96- 105%
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Steroids panel: androstenedione at RT=8.14 min / protein precipitation sample preparation
DMS OFF MRM 287/109
DMS OFF MRM 287/97
Peak of interest
Peak of interest
Peak of interest
Peak of interest
DMS ON MRM 287/109
DMS ON MRM 287/97
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DMS as separation technique when LC is not an option…
Infusion based analysis
Surface Sampling
– LESA
Direct desorption technique
– DESI
– LDTD
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Advion LESA (Liquid Extraction Surface Analysis)
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Propranolol Glucuronide Structural
Isomers (acyl glucuronide)
Aromatic Glucuronide
Aliphatic Glucuronide
Isobaric metabolites
Same fragments and dominant MRM transition!
Direct Tissue Profiling using the Advion LESA instrument
DMS separation with 1.5% ACN modifier
DMS resolves propranolol and the 2
glucuronide structural isomers!
CV = -17.4 V
CV = -10.4 V
28 © 2011 AB SCIEX
LDTD with SelexION™ Technology
LDTD on 5500 with DMS
22X background
reduction
2576 cps
J Wu et al., Anal. Chem., 2007, 79, 4657-4665.
2.5 pg Clozapine in protein precipitated plasma
LDTD on 5500
5576 cps
Hesham Ghobarah, Yves Leblanc, Michael Jarvis, Adrian Taylor, and Pierre Picard
Thermal desorption/APCI
Low
STD
Matrix
BLANK
30 © 2011 AB SCIEX
Summary
An added dimension of selectivity through differential mobility spectrometry with the AB SCIEX SelexION™ Technology on the QTRAP® 5500 system and the Triple Quad™ 5500 system
Improvement in data quality and LOQs for assays with interferences
Robust and reliable for use in a regulated environment
Improved throughput – Less need for chromatographic separations and sample prep
– Can be coupled to non-LC sample introduction
Combined with MRM3 on the QTRAP 5500 system, SelexION Technology gives several options
– High sensitivity in MRM mode
– High selectivity for analytes that fragment well with MRM3
– High selectivity for analytes that fragment poorly with DMS-MS or DMS-MS/MS with the SelexION Technology
– Separation for isobaric analytes with common fragments
31 © 2011 AB SCIEX
Thank You
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