Technology Transition Workshop Introduction to DART MS Robert B. Cody JEOL USA, Inc.
Dec 26, 2015
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Outline
• Definition of terms
• DART operating principle
• TOF mass spectrometer overview
• The information we obtain
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High Resolution Mass Spectrometry
• We will be using exact-mass measurements to to confirm knowns and to determine elemental compositions for unknowns
• Resolving power defines how well the mass spectrometer can separate close peaks (interferences)
• The elemental composition software gives us other information for each candidate composition (e.g. unsaturation)
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Resolving Power
R = M / M
R = Resolving Power
M = m/z
M = difference in mass that can be separated
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R = M / MR = 5000m/z 500
M = Peak width at half-height = 0.1
Resolving Power Defined as: FWHM (Full width at half maximum)
0.1
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R = M / MR = 500
m/z 500 and 501 can be separated at a 10% Valley
M = 1
Resolving Power Defined as: 10% Valley Definition
501
500
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Examples for C36H74 (m/z 506.579)
R = 500 (10% valley)
Separate m/z 500 from 501
R = 5000 (10% valley)
Separate m/z 500 from 500.1
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Why the definition matters
R = 500 (10% valley) R = 500 (FWHM)
R = 5000 (FWHM)
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Mass accuracy
• millimass units (0.001) or “mmu”
ppm = 106 * (M / M)
• parts-per-million (ppm)– “Resolution” (reciprocal of resolving power)
Note: ppm is a m/z – dependent value
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Unsaturation (aka “rings and double bonds” aka “double bond equivalents”)
• Value is calculated from elemental composition• Indicates total rings, double bonds, triple bonds• Exact integer (e.g. “4.0”) or half-integer (“3.5”)
C6H6+.
D = 4.0
CH3 CH3
O H+
C3H7O+.
D = 0.5, add 0.5
CH3COO-
D = 1.5, subtract 0.5
H3O+
D = -0.5, add 0.5
Technology Transition WorkshopExamples of Even-electron ionsand Odd-electron ions
• Even-electron ions (half integer unsaturation) :Protonated molecule [M+H]+
Deprotonated molecule [M-H]-
Chloride adduct [M+Cl]-
Ammoniated molecule [M+NH4]+
Fragment F+
• Odd-electron ions (exact integer unsaturation) :Molecular radical cation M+.
Molecular radical anion M-.
Fragment F +.
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On-line Resources
• DART Users’ Google Newsgroup– http://groups.google.com/group/dart-mass-spectrometer-users?hl=en
• JEOL USA, Inc. Web Pages– http://www.jeolusa.com
• IonSense Web Page– http://www.ionsense.com
• Wikipedia article on DART– http://en.wikipedia.org/wiki/DART_ion_source
• Proton affinities, ionization energies (NIST)– http://webbook.nist.gov/chemistry/
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DART Basic Principles
See the JEOL News Article on theAccuTOF-DART product page on
www.jeolusa.com
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DART:“Direct Analysis in Real Time”
• Operational in Jan. 2003• Patent filed in April 2003• Public disclosure, Jan. 2005• Commercial product introduced March 2005• First open-air, ambient ion source for MS
1. Cody, R. B.; Laramee, J. A. “Method for atmospheric pressure ionization” US Patent Number 6,949,741 issued September 27, 2005.2. Laramee, J. A.; Cody, R. B. “Method for Atmospheric Pressure Analyte Ionization” US Patent Number 7,112,785 issued September 26, 2006.
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Prototype DART sources
Original prototype DART source (mid-2002) Second DART prototype(Early 2003)
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Why DART?
• Fast and easy way to introduce samples• Minimal sample preparation for most samples• Can tolerate “dirty” or high-concentration
samples and without contamination• Fast fingerprinting of materials
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Nothing comes without a price
• Chromatography/MS still has advantages over DART in detection limits, selectivity and sensitivity for certain samples
• Not useful for large biomolecules (no good for DNA analysis, proteins)
• DART does not ionize metals, minerals, etc.
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Penning ionization
Sample ionized directly by energy transfer from metastables (M*)
Proton transfer (positive ions)
1. He* ionizes atmospheric water
2. Ionized water clusters transfer proton to sample
Electron capture (negative ions)
1. Penning electrons rapidly thermalized
2. Oxygen captures electrons
3. O2- ionizes sample
DART Source
DART Ionization
MS APIInterface
M*
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Penning Ionization
• Metastable atoms or molecules react with analytes that posses ionization potentials less than the metastable energy,
M* + S S+. + M + electron
• The helium 23S state has 19.8 eV of internal energy and lasts up to 8 minutes in vacuum.
– Most molecules have ionization energies much lower than 19.8 eV
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Proton Transfer
• Metastable atoms react with atmospheric water to produce ionized water clusters
• Dominant reaction mechanism when helium carrier used: He(23S) energy = 19.8 eV
• Huge reaction cross section: 100 A2
He(23S) + H2O H2O+• + He(11S) + electron
H2O+• + H2O H3O+ + OH•
H3O+ + nH2O [(H2O)n+1H]+
[(H2O)nH]+ + M MH+ + nH2O
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Typical DART Low-Mass Background
0
20
40
60
80
100
Rel
. A
bu
nd
.
15 20 25 30 35 40 45 50 55
m/z
NH4+
H3O+
NO+
[(H2O)2+H]+
[(H2O)3+H]+
Normal DART Parameters
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Negative Ion Formation
• Electrons produced by direct or surface Penning ionization are rapidly thermalized
• Thermal electrons react with atmospheric oxygen and water to produce ionized clusters
• Oxygen/water cluster ions react with analyte molecules to produce analyte ions
e-* + G e- + G*
e- + O2 O2-.
O2-. + S [S-H]- + OOH.
O2-. + S S-. + O2
O2-. + S [S+O2]-.* + G [S+O2]-. + G*
Technology Transition WorkshopTypical DART Negative-IonLow-Mass Background
20 40 60 80 100
m/z
Rel
. abu
ndan
ce [H2O3]-O2-
[HCO4]-
[HCO3]-
[H2O4]-
20 40 60 80 100
m/z
Rel
. abu
ndan
ce [H2O3]-O2-
[HCO4]-
[HCO3]-
[H2O4]-
Note the absence of nitrogen oxide ions that would be produced by electrical discharge in air. NO2
- and NO3
- are problematic for detection of nitro explosives and reduce anion detection sensitivity
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Example
[M-H]-
0
50
100
Rel
. A
bu
nd
.
100 150
m/z
175.0232
0
50
100
Rel
. A
bu
nd
.
100 150
m/z
177.0410
[M+H]+
[M+H-2H2O]+
[M+H-H2O]+
Positive ions
Negative ions
Ascorbic acid, C6H8O6
127 130 133 136 139 142 145 148 151 154 157 160 163 166 169 1729
52
95
O
OH O
H O
O H
H O
Sampled directly from a melting point tube
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Notes on the AccuTOF Design and Operation
See the JEOL News Article on theAccuTOF-LC product page on
www.jeolusa.com
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Types of mass spectrometers
• Scanning: – magnetic sector, quadrupole and triple
quadrupole
• Trapped-ion: – Fourier transform, 3D ion trap, Orbitrap
– linear trap (used in triple quadupole MS)
• Time-of-flight• Hybrids
Technology Transition WorkshopDART can be fit on most mass spectrometer types
DART signals can be transient, so,
• scanning mass specs work best with selected ion monitoring or fast scanning
• Selected reaction monitoring on triple quadrupole MS is good for target compound quantitation.
• Ion traps work, but are not a good choice for quantitative analysis
• Time-of-flight is fastest MS for transient signals, and gives high-resolution data for the entire mass spectrum with no sensitivity loss.
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Time of flight principle
Heavy ions moving slowly
L’Alpe D’Huezde
Spectrometrie de Masse
Light ions moving quickly DetectorIf everyone starts at the same time and has the same kinetic energy, lighter riders will move faster
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A more realistic TOF mass spectrometer
High voltage to accelerate ions
Ion detector
Ion source:Short burst of ions
Flight tube
Kinetic Energy = qE = mv2/2
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What if ions that have the same mass have slightly different energies?
• Reflectron: make the more energetic ions travel further
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Reflectron Time of flight mass analyzer principle
MeLance
1. Fast riders miss the turn
Technology Transition WorkshopReflectron Time of flight mass analyzer principle
2. Fast riders turn around; have to travel further
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Reflectron TOF
4. Fast riders catch up, will eventually pass
Focal point
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Time-of-flight math• All ions fly with the same kinetic energy.
• Flight time is inversely proportional to the square root of the mass/charge ratio.
M
q
V
Ltof
VeqvmM u 2)(2
1
M: mass of ion [u] mu: Atom mass unit (1.6605 x 10-27 [kg/u])
v: flight speed of ion [m/s] q: charge number of ion
e: unit electric charge (1.602 x 10-19 [C]) V: Accelerating voltage [V]
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JMS-T100LC AccuTOFTM
TMP2
RP
TMP1
RP
Ion SourceIon
Transportation Analyser
Detection system
To the data collection
system
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AccuTOFTM Ion Source
TMP2
RP
TMP1
RP
Ion SourceIon
Transportation Analyser
Detection system
To the data collection
system
Technology Transition WorkshopOrthogonal ESI ion source and API interface
Nebulizer Gas
RP TMP
Desolvating Chamber
Orifice1Ring Lens
Orifice2Ion GuideDesolvating
Gas
LC Eluent
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Ion Source and Atmospheric Pressure Ionization (API) Interface
• Orthogonal ESI– Minimize contamination into API interface
• Simple API interface– Robust, few parameters, minimal maintenance
• Off-axis skimmers and ring lens, bent ion guide– Keep contamination out of high-vacuum region
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AccuTOFTM Ion Transport
TMP2
RP
TMP1
RP
Ion SourceIon
Transportation Analyser
Detection system
To the data collection
system
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Ion transport region
• Strong acceleration of ions only occurs in high-vacuum region– Minimize CID and scattering
• Quadrupole RF ion guide focuses ions to a small spot size– Spatial focus for good resolution
– “High-pass” filter (ions greater than given m/z)
• Multi-function focusing and steering lenses– Beam should be perpendicular
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AccuTOFTM Analyzer
TMP2
RP
TMP1
RP
Ion SourceIon
Transportation Analyser
Detection system
To the data collection
system
z
y
x
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AccuTOFTM Analyzer
• Two-step acceleration– Spatial focusing of ion beam
• Single reflectron– Energy focusing of ion beam in the x-direction– Minimize ion loss
• oa(Orthogonal-Acceleration)-TOF MS– Kinetic energy spread in y-direction has no effect
on resolution– The ions produced by the ESI ion source are
used efficiently.
z
y
x (reflectron)
(injection)
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Flight cycle of oa-TOF MS
• 1. Introduction of ion– Two kinds of ions are introduced at the same time.
Ion Source Low mass ionHigh mass ionMixture of both ions
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Flight cycle of oa-TOF MS
• 2. Turn on the pulser voltage– Mixture of ions at the start of flight
++++++
-
-
Ion Source
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Flight cycle of oa-TOF MS
Ion Source
• 3. Turn off the pulser voltage– continuing flight - mass separation
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Flight cycle of oa-TOF MS
Ion Source
• 4. Continuing flight – New ions are introduced in the ion acceleration part.
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Flight cycle of oa-TOF MS
Ion Source
• 5. Low mass ion reaches detector– The ion acceleration region is filled with the new ions.
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Flight cycle of oa-TOF MS
Ion Source
• 6. High mass ion reaches detector
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Flight cycle of oa-TOF MS
Ion Source
• 7. The detection of all ions is completed
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AccuTOFTM Detection system
TMP2
RP
TMP1
RP
Ion SourceIon
Transportation Analyser
Detection system
To the data collection
system
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Detector
① Micro-channel plate (MCP) 40mmφ Dual MCP
② Anode Combined with high voltage capacitor
Patent pending+
e-
in the vacuum
in the atmosphere
Dual MCP
Anode
To impedance converter
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MCP• Diameter : 40mm• Thickness : 0.6mm• I.D. of channel : 10μm• Gap of each channel : 12μm
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Data collection system for oa-TOF MS
• High time resolution– m/z 609, R=6,000 → Peak width: 3.5ns
• Continuous data collection– High duty cycle
• Real-time accumulation of mass spectrum
- Requirements -
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Data collection system for oa-TOF MS
• TDC– Super-high speed
digital stop watch
– Measures the arrival time of ions
– A premise is that there are a few ions
• Each ion arrives separately.
• Ion counting detection: signal is 0 or 1.
• Continuous Averager– A signal from the
detector is converted digital value by a high-speed ADC (Analog-to-Digital Converter).
– Spectrum can be accumulated continuously in real time.
Technology Transition WorkshopTDC (Time-to-Digital Converter)
High Voltage
Pulser
DiscriminatorAmp
Start Input
Stop Input
59us
Time-to-Digital
Converter
No. TOF [us] 1 29.4235 2 46.2890 ....
No. of Ions Detected in a Cycle
Histogram Memory
To Data System
Technology Transition WorkshopSimulation of spectrum accumulation by TDC
Output from Amplifier : Cycle 1
0
5
10
15
20
1 5 9 13 17 21 25 29 33 37 41 45
mV
Histgram memory : Cycle 1
0
1
2
3
1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46
No.
of
Ions
Technology Transition WorkshopSimulation of spectrum accumulation by TDC
Output from Amplifier : Cycle 2
0
5
10
15
20
1 5 9 13 17 21 25 29 33 37 41 45
mV
Histgram memory : Cycle 2
0
1
2
3
1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46
No.
of
Ions
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Output from Amplifier : Cycle 3
05
101520
1 5 9 13 17 21 25 29 33 37 41 45
mV
Histgram memory : Cycle 3
0
1
2
3
1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46
No.
of
Ions
Simulation of spectrum accumulation by TDC
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Output from Amplifier : Cycle 4
05
101520
1 5 9 13 17 21 25 29 33 37 41 45
mV
Histgram memory : Cycle 4
0
1
2
3
1 5 9 13 17 21 25 29 33 37 41 45
No.
of
Ions
The ion which had about two times higher intensity was detected.
It is counted only once (not twice) with TDC.
Simulation of spectrum accumulation by TDC
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Output from Amplifier : Cycle 5
0
5
10
15
201 5 9 13 17 21 25 29 33 37 41 45
mV
Histgram memory : Cycle 5
0
1
2
3
1 5 9 13 17 21 25 29 33 37 41 45
No.
of
Ions
Two ions detected in succession!
The second ion can't be counted during dead time.
Simulation of spectrum accumulation by TDC
Technology Transition WorkshopResult of spectrum accumulation by TDC
• The ratio of the peak intensity isn't correct.
• A high intense peak shifts to low mass side.
Histgram memory : Cycle 5
0
1
2
3
1 5 9 13 17 21 25 29 33 37 41 45
No.
of
Ions
model spectrum :
05
10152025
1 5 9 13 17 21 25 29 33 37 41 45
No.
of
Ions
Technology Transition WorkshopContinuous Averager
High Voltage
Pulser
Amp
59us
Intensity1528 ....
No. of Data Points on a Spectrum
(up to 256K points)
Summing Memory
To Data System
ADC
(8bit)
Timing Control Circuit
Adder
Continuous Averager
Technology Transition WorkshopSimulation of spectrum accumulation by continuous averager
Output from Amplifier : Cycle 1
0
5
10
15
20
1 5 9 13 17 21 25 29 33 37 41 45
mV
Cycle 1
01020304050
1 5 9 13
17
21
25
29
33
37
41
45
mV
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Output from Amplifier : Cycle 2
0
5
10
15
20
1 5 9 13 17 21 25 29 33 37 41 45
mV
Cycle 2
01020304050
1 5 9 13 17 21 25 29 33 37 41 45
mV
Simulation of spectrum accumulation by continuous averager
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Output from Amplifier : Cycle 3
05
101520
1 5 9 13 17 21 25 29 33 37 41 45
mV
Cycle 3
01020304050
1 5 9 13 17 21 25 29 33 37 41 45
mV
Simulation of spectrum accumulation by continuous averager
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Output from Amplifier : Cycle 4
05
101520
1 5 9 13 17 21 25 29 33 37 41 45
mV
Cycle 4
01020304050
1 5 9 13 17 21 25 29 33 37 41 45
mV
Simulation of spectrum accumulation by continuous averager
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Output from Amplifier : Cycle 5
05
101520
1 5 9 13 17 21 25 29 33 37 41 45
mV
Cycle 5
01020304050
1 5 9 13 17 21 25 29 33 37 41 45
mV
Simulation of spectrum accumulation by continuous averager
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• The ratio of the peak intensity is correct.
• There is no shift of the ion peak.
model spectrum :
05
10152025
1 5 9 13 17 21 25 29 33 37 41 45
No.
of
Ions
Cycle 5
01020304050
1 5 9 13 17 21 25 29 33 37 41 45
mV
Result of spectrum accumulation by continuous averager
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Specifications
• Mass resolution : 6,000– FWHM, Reserpine m/z 609
• Sensitivity : Reserpine 10pg S/N>10– LC-ESI [ Flow rate: 0.2mL/min ]– Mass chromatogram of m/z 609, RMS
• Mass accuracy : 5ppm RMS– With internal reference– (Typically better than that!)
Technology Transition WorkshopOnly 3 analyzer parameters are critical for routine DART analysis
TMP2
RP
TMP1
RP
Ion SourceIon
Transportation Analyser
Detection system
To the data collection
system
1 2
3
1: Orifice 1 2: “Peaks voltage”3. Multiplier V
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The 3 important parameters
• 1: Orifice 1: Typically 20V– Increase O1 to increase fragmentation
• 2: “Peaks voltage” (RF ion guide voltage)– Divide by 10 to estimate lowest detected m/z
• 3. Multiplier V: Typically 2200V to 2600V– Increase multiplier to increase signal (and
noise)
Technology Transition WorkshopInformation from the TOF mass spectrum
• Exact mass + isotope peaks: elemental composition
• Fragmentation: distinguish isomers• “Fingerprint” pattern: material ID• Ion abundance: quantitative analysis• Other experiments: H/D exchange,
derivatization, etc.
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Example: DART mass spectrum of a leaf
0
20
40
60
80
100
Rel
. A
bu
nd
.
100 150 200 250 300 350
m/z
290.174304.154
What is this?
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Elemental compositions
Elemental Composition Program
MeasuredExact Mass
Constraints
Candidate compositions
Ranked compositions
Isotope pattern matching
Technology Transition WorkshopWe have a composition. Now what?
m/z 304.1548 is C17H22NO4
CocaineScopolamine
174 184 194 204 214 224 234 244 254 264 274 28417
57
97
N
OO
O
O H
Fragments at m/z 138, 156
202 212 222 232 242 252 262 272 282 29213
56
99
N
H
H O
O
O
O
Fragments at m/z 182, 82
182
138
C10H16NO2+
C8H12NO+
156C8H14NO2
+
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Control fragmentationwith Orifice 1 and Ring Lens potentials
Nebulizer Gas
RP TMP
Desolvating Chamber
Orifice1Ring Lens
Orifice2Ion GuideDesolvating
Gas
LC Eluent
API interface change potentials to induce fragmentation
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0
20
40
60
80
100
Rel
. A
bu
nd
.
100 150 200 250 300
m/z
Fragment spectrum increase cone voltage from 20 V to 60 V
290.174
304.154156.099138.089
C8H12NO+ C8H14NO2+
Scopolamine
Atropine
C8H14N+
Scopolamine
Technology Transition WorkshopFor comparison, m/z 305.1548 fragments from a dollar bill
0
20
40
60
80
100
Rel
. A
bu
nd
.
100 150 200 250 300 350
m/z
82.065
182.118
Cocaine
C5H8N+
C10H16NO2+
C17H22NO4+
Technology Transition Workshopor…we can search for candidates from a list of target compounds.
SearchFromList Program
Components in asmokeless powder