Introduction to Mass Spectrometry

Navnath Jaybhaye
Manager- Product development & Application

BioAnalytical Technologies (I) Pvt. Ltd.

1

Agenda

Introduction Overview of MS Ionization Techniques Mass Analyzers Quadrupole Operations Applications Of LCMS

Analytical Assays used in Pharmaceutical Industry Labs for New Chemical Entities
Method
HPLC
(UV &Fluorescence)

1990
75% 12% 3% 10%

1998
50-60% 3% 40-50% 10%

2000
20% 2% 60-75% 10%

2006
2% 0 98% 0
3

GC/MS LC/MS/MS Immunoassay

(ELISA/FPIA etc.)

Mass Spectrometers
Separate and measures ions based on their mass-tocharge (m/z) ratio. Operate under high vacuum (keeps ions from bumping into gas molecules) Key specifications are resolution, mass measurement accuracy, and sensitivity. Several kinds exist: for analysis, quadrupole, time-offlight (TOF) and ion traps are most used.

MS vs. MS/MS
GC HPLC CE Inlet Ionize Mass Analyze Detect

Separation

MS
Mass Analyze MS1

Identification

Inlet

Ionize

Fragment Collision Cell

Mass Analyze MS2

Detect

MS/MS

Mass Spectrometry
+CH 3 +COH

CH3COCH3

CH3+COCH3

CH3C+OCH3

+COCH 3

Sample Inlet

Ionization Fragmentation & Adsorption (Dissociation) of Excess Energy

Mass Analysis

Detection

Components of Tandem Mass Spectrometer

Ionization Source Mass Spectrometer ESI APPI APCI MALDI Quatrupole Magnetic Sector Collision Cell Argon Xenon Mass Detector Spectrometer Quatrupole Magnetic Sector Time-of-flight

MS1

Collision cell

MS2

Sample introduction
Ion Souce
Transforms sample molecules to ions Soft ionization
Places positive or negative charge on the analyte without significantly fragmenting the analyte M+1 ion (or M-1 ion) No need to volatilize Down to fmol detection limits

Atmospheric Pressure Ionization (API)

Electrospray MALDI APCI APPI
8

The Macabre History of Electrospray

The Abb Nollet experimented with electrified liquids in the 18th century ! He observed that when a person was connected to a high-voltage generator he/she would not bleed normally after cutting ...blood sprayed from the wound !

F. Lemire, LCGC Europe LC-MS Supplement, December 2001, p29-35

9

How Sample is Introduced

Sample Inlet Ion Spray Inlet

Ion Spray Heater

Ion Spray Probe

Heater

The sample is held on the surface of a capillary tube, a fine wire, or a small cup.
10

The Electrospray Phenomenon

J. Zelene, Phys. Rev., 10, 1-6 (1917)

11

Ionization Source

12

Ionization Source
Spraying Needle Sample Cone
Orifice = 400m

Vacuum Isolation Valve

13

Ionization Methods.
Gas Phase Ionization Electron Impact (EI) Chemical Ionization (CI) Spray Ionization / Atmospheric Pressure Ionization (API)
Electrospray (ESI) / API Atmospheric Pressure Chemical Ionization (APCI) Atmospheric Pressure photo Ionization (APPI)

Desorption Ionization

14

II. Spray Ionization/API

The compound of interest in a volatile buffer mobile phase, is passed through heated, narrow bore tubing directly into the ion source of a mass spectrometer. The solution is vaporized in the tubing, and analyte ions desorb into the gas phase and pass into the mass analyzer. Electrospray (ESI) / API Atmospheric Pressure Chemical Ionization (APCI) Atmospheric Pressure photo Ionization (APPI)
15

Electrospray Ionization(ESI)

16

Ion Sources make ions from sample molecules

Electrospray ionization:
Pressure = 1 atm Inner tube diam. = 100 um Sample Inlet Nozzle (Lower Voltage) Partial vacuum

N2
+ + + + ++ ++ + + ++ ++ + + ++ + ++ + ++ + ++ + ++ + ++ + ++

MH+
+ + + + + + + + + +

Sample in solution N2 gas

MH2+ MH3+

High voltage applied to metal sheath (~4 kV)

Charged droplets 17

TURBOIONSPRAY (TIS)

18

API 4000 Turbo Ion Spray

TM

19

ESI Spectrum of Trypsinogen (MW 23983)

1599.8

M + 15 H+ M + 14 H+

M + 16 H+
1499.9 1714.1

M + 13 H+

1411.9

1845.9 1999.6 2181.6

m/z

Mass-to-charge ratio
20

APCI

21

APPI

22

MALDI: Matrix Assisted Laser Desorption Ionization

Sample plate h

Laser

MH+

1. Sample is mixed with matrix (X) and dried on plate. 2. Laser flash ionizes matrix molecules. 3. Sample molecules (M) are ionized by proton transfer: XH+ + M MH+ + X.

+/- 20 kV

Grid (0 V)
23

The mass spectrum shows the results

MALDI TOF spectrum of IgG
Relative Abundance
40000

MH+

30000

(M+2H)2+
20000

10000

(M+3H)3+
0

50000

100000

150000

200000

Mass (m/z)
24

Differential vacuum in MS
OR IQ1

DP-FP
RNG

Collisional Focussing
QO

Q1 Analyzer
IQ2

Collision Cell (N2)

RO2 IQ3

Q3 Analyzer
DET

ST

RO1

RO3

DF

8 x 10-3 torr 1 torr

1x10-5 torr

1-5 x 10 torr

-4

1-2 x 10-5 torr

Turbo

Roughing pump

25

Significance of vacuum
A vacuum is necessary to permit ions to reach the detector. It reduces or eliminates the chances of ion collisions with mass analyzer. The major reason for maintaining high vacuum is to increase the mean-free path of ions. Increases sensitivity and resolution of Mass Spectrum.

26

Types Of Analyzers
Quadrupole Ion Trap Time-of-flight

27

Analytical Quadrupole

28

Schematic Diagram of Triple quad Instrument

29

Quadrupole Theory
Pre-filter Quadrupole Mass Filter
Stable Trajectory

Unstable Trajectories

Only ions with the correct m/z values have stable trajectories within an RF/DC Quadrupole field. Ions with unstable trajectories collide with the rods, or the walls of the vacuum chamber, and are neutralised.
30

1. Quadrupole

31

Quadrupole: Pros & Cons

Advantages Relatively small and low-cost systems Low-energy collision-induced dissociation (CID) MS/MS Triple quadrupole and hybrid mass spectrometers. Limitations Limited resolution Peak height vs. mass response must be 'tuned'. Not well suited for pulsed ionization methods Applications Majority of benchtop GC/MS and LC/MS systems Triple quadrupole MS/MS systems quadrupole hybrid MS/MS systems
32

Tandem Quadrupole
MS1 Collision cell MS2

33

Components of Tandem Mass Spectrometer

Ionization Source Mass Spectrometer ESI APPI APCI MALDI Quatrupole Magnetic Sector Collision Cell Argon Xenon Mass Detector Spectrometer Quatrupole Magnetic Sector Time-of-flight

MS1

Collision cell

MS2

34

Operation Modes
Product Ion Scanning
Analyzes all products of a single precursor

Precursor Ion Scanning

Analyzes all precursors of a single charged product

Neutral Loss Scanning

Analyzes all precursors of a single uncharged product

Multiple Reaction Monitoring

Analyzes for specific precursors producing specific products.
35

Full Scan Acquisition Mode

MS1

Collision cell

MS2

MS1

Collision Cell

MS2

Scanning

Rf only, pass all masses

SCANNING MODE: The first quadrupole mass analyzer is Scanning over a mass range. The collision cell and the second quadrupole mass analyzer allow all ions to pass to the detector.

36

Full Scan Acquisition Mode

Mass Spectrum: Progesterone

[M+H]+
100
315.1
CH3 O CH3 CH3

%
316.1

0 200

220

240

260

280

300

320

340

360

380

m/z 400

37

MS1

Collision cell

MS2

Product ion scanning

Argon gas

Products Precursor

Static (m/z 315.1)

Scanning

The first quadrupole mass analyzer is fixed at the mass-to-charge ratio (m/z) of the precursor ion to be interrogated while the second quadrupole is Scanning over a user-defined mass range.

38

Product Ion Spectrum: Progesterone

Product ion scanning

100
CH3

CH3

CH3

315.1

% 0 300
97.0

Precursor ion
305
CH2

316.1

Mass Spectrum from MS1

m/z 330

310
O CH3

315

320

325

100 % 0

109.0

CH2

H3C CH3

Product ions
100 125 150 175

Product ion spectrum from MS2

m/z 200 225 250 275 300 325
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Precursor ion scanning

Precursor Ion Scan

MS1

Collision cell

MS2

Argon gas

Product Precursors

Scanning Static The first quadrupole mass analyzer is Scanning a mass range while the second quadrupole is fixed, or Static, at the mass-to-charge ratio (m/z) of a product ion known to be common to the analytes in a mixture. 40

Acylcarnitines
R=0 to 18 carbon alkyl chain.

Derivatization and Fragmentation

H CH COOH

Precursor ion scanning

RCOO (CH3)3N CH2 CH

Butylation
RCOO (CH3)3N CH2 CH H CH - RCOOH -(CH3)3N -C4H8 COOC4H8

CID

All compounds of this type fragment to produce the 85 ion.

[ CH2

CH (m/z 85)

CH

COOH

]+
41

Normal Acylcarnitine Profile

Precursor ion scanning
100

d3-free carnitine

d3-C16 carnitine

C2 carnitine
%

d3-C3 carnitine C16 carnitine d3-C8 carnitine

0 225 250 275 300 325 350 375 400 425 450 475 500
42

m/z

MS1

Collision cell

MS2

Neutral loss scanning

Argon gas
Products Precursors

Scanning (M)

Scanning (M-102)

In a neutral loss scan the two quadrupole mass filters are Scanning synchronously at a user-defined offset. The neutral loss is known to be common to the analytes in a mixture. 43

Neutral and Acidic Amino Acids Derivatization and Fragmentation

O R OH NH2

(Generic)
O R NH2 O CH3

HO

+
CH3

HCl

Neutral or Acidic AA
O R NH3
+

Butanol

Amino acid butyl ester

O

Fragmentation R
O CH3
NH2
+

+
HO

CH3

Neutral or Acidic AA

Fragment

Butyl formate Neutral loss of 102Da 44

Normal Amino Acid Profile

Neutral loss scanning
Pro
100

Deuterated internal standards for quantification

d3-Leu d5-Phe
%

d6-Tyr

d4-Ala Ser Gly d8-Val d3-Met

Glu

0 140 16 0 180 200 220 240 2 60

m /z 280 45

Multiple Reaction Monitoring

MS1

Collision cell

MS2

Argon gas
Product(s)

Precursor(s)

Static (m/z 315.1)

Static (m/z 109.0)

Both the first and second quadrupole mass analyzers are held Static at the mass-to-charge ratios (m/z) of the precursor ion and the most 46 intense product ion, respectively.

Collision induced dissociation

Argon gas
O CH3 CH3
H3C CH2 CH2 O CH3

CH3
O

CH3

Precursor ion

Product ions

In the collision cell, the TRANSLATIONAL ENERGY of the ions is converted to INTERNAL ENERGY.
Collision conditions (FRAGMENTATION) is controlled by altering: The collision energy (speed of the ions as they enter the cell) Number of collisions undertaken (collision gas pressure)
47

2. Quadrupole Ion Trap

In an Ion trap the ions are trapped in a radio-frequency quadrupole field. The ions are then ejected detected as the radio frequency field is scanned. Ions are dynamically stored in a three-dimensional quadrupole ion storage device. The RF and DC potentials can be scanned to eject successive mass-to-charge ratios from the trap into the detector.

48

QTRAP Linear ion trap

Ion accumulation

N2 CAD Gas

Exit lens

Q0

Q1

Q2

Q3

ion selection

Fragmentation

linear ion trap 3x10-5 Torr

49

3-D Ion Trap Schematics

Heated quartz capillary

MS Scan Product Ion Scan MSn

Ion Trap ID ~ 10 mm
50

Q-Ion Trap- Pros & Cons

Benefits High sensitivity Multi-stage MS Limitations Poor quantitation. Subject to space charge effects and ion molecule reactions. Collision energy not well-defined in CID MS/MS. Applications Compact mass analyzer

51

3. Time-of-Flight (TOF)
Time of flight mass spectrometer measures the massdependent time It takes ions of different masses to move from the ion source to the detector. Ions are either formed by a pulsed ionization method (usually MALDI), or various kinds of rapid electric field switching are used as a 'gate' to release the ions from the ion source in a very short time.

52

TOF - Pros & Cons

Benefits Fastest MS analyzer Well suited for pulsed ionization methods High ion transmission Highest practical mass range of all MS analyzers Limitations Requires pulsed ionization method or ion beam switching Fast digitizers used in TOF can have limited dynamic range Limited precursor-ion selectivity for most MS/MS experiments Applications MALDI systems Very fast GC/MS systems

53

DETECTORS
Mass analysis is the separation of bunches or streams of ions according to their individual mass-to-charge (m/z) ratio The mass analyzer sorts the ions according to m/z and the detector records the abundance of each m/z.

54

Detectors are eyes of the Instrument

Once the ion passes through the mass analyzer it is then detected by the ion detector, The final element of the mass spectrometer.
55

METHODS OF ION DETECTION

The detector generates a signal from incident ions by two ways:
1. Inducing current generated by a moving charge 2. Generating secondary electrons, which are further amplified.

56

Current generated by a moving charge.

Flow of electrons in the wire is detected as an electric current which can be amplified and recorded. The more ions arriving, the greater the current. Variation in the magnetic field, changes the flow of ion stream to the detector which produce a current proportional to the number of ions arriving. Timing mechanisms, which integrate those signals with the scanning voltages, allow the instrument to report which m/z strikes the detector
57

Current generated by secondary electrons

Detector operates by producing a signal current from incident ions by generating secondary electrons, which are further amplified Key part of such type of detectors is a dynode. Dynode is electron-multiplying electrode.
Incoming Ion

Secondary Electrons

58

Contd

Secondary electrons Electrode surface(dynode)

Dynodes For Amplification

Process of Secondary electron emission.

Electrons accelerated, and strike the surface of electrode (dynode) Energy deposited by the incident electrons result in re- emission
59

CHARECTERISTICS
Certain of these
characteristics are common, like :

high sensitivity linear, quantitative response Some detectors are

designed for specific functions or applications.
60

Electron Multipliers contd

Electron multiplier is made up of a series of dynodes maintained at ever increasing potentials. Typical amplification or current gain of an electron multiplier is one million.

61

Electron Multipliers contd

Two major modifications: Multiple-dynode type Continuous-dynode type

62

1. Multiple-dynode type EM
Incident ions

Cathode

Anode

Incoming ion reaches the first dynode Ejects several other electrons by secondary emission. This process repeated at each succeeding dynode having
a higher potential than the preceding dynode.

When it arrives at the anode, the electron flow is

significantly amplified
63

2. Continuous Electron Multiplier

resistive-material inner coating

Contains a glass pipe with a coating on its inner surface The electron flow moves along the pipe, reflecting from
the inner wall and progressively gaining electrons

The electrical field accelerating the flow is formed by

the high voltage applied across the two ends
64

Features of Electron Multipliers

Advantages:
Very

high current gain Sensitive Fast

Disadvantage:
Short lifetime Requires good vacuum to operate

Electron multipliers are widely used in Quadrupole and

Ion trap Instruments.
65

Channel Electron Multiplier

A modified continuous
dynode electron multiplier

Comprised of "the channel,"

a hollow, cornucopia-shaped tube made of semi conductive glass.

66

Channel Electron Multiplier contd..

IONS

The primary incoming ions passes through the inlet and strikes the
surface of the CEM The collision energy eject an electron from CEM wall Ejected electrons accelerated into interior of CEM Trigger secondary emission and the process continues to produce an output electron
67

RECORDER
Analogue signal is produced by the detector. Analogue Digital Converter sends the output to the computer.

Detector

ADC

Recorder

68

IDENTIFICATION

CHARACTERIZATION

Single MS Specificity
Precursor Ion Neutral Loss Enh.Multi Charged

High Sensitivity Full Scan MSMS MS3 Capabilities

Plug & Play Sources

QUANTITATION
SIM or MRM

TurboIonspray Heated Nebulizer (APCI) Nanospray Integrated Syringe pump Built-in divert valve

AUTOMATION
Info. Dep. Acquisition
MetID
69

Where are Mass Spectrometer Used

Biotechnology: the analysis of Proteins, Peptides, Oligonucleotides Pharmaceutical: Drug Discovery, Combinatorial Chemistry, Pharmacokinetics, Drug Metabolism Environmental: PAHs, PCBs, water quality. Food contamination Clinical: neonatal Screening, Hemoglobin analysis, drug testing Geological: Oil composition
70

How can Mass Spectrometry help Biochemists

Accurate molecular weight measurements: sample confirmation, to determine the purity of a sample, to verify amino acid substitutions, to detect post-transnational modifications, to calculate the number of disulfide bridges Reaction monitoring:to monitor enzyme reactions, chemical modification, protein digestion Amino acid sequencing:sequence confirmation, de novo characterization of peptides, identification of proteins by database searching with a sequence tag from a proteolytic fragment Oligonucleotide sequencing:the characterization or quality control of Oligonucleotides Protein structure:protein folding monitored by H/D exchange, proteinligand complex formation under physiological conditions, macromolecular structure determination
71

Application Example 1

LC-MS/MS
Selectivity and Sensitivity comparisons Application Area: Environmental

72

LC/MS or LC/MS/MS? Selectivity & Sensitivity

Quantitation of 3 pesticides in a surface water extract Simazine; Atrazine and Metabenzthiazuron Chromatographic and Mass Spec conditions: Ionization technique : APCI
73

Product Ion Spectra

MSMS Product Ion Scan= N2
NO2 NO2 NO2 NO2

Q1 fixed, =

CAD Collision

Q3 scanning

MH+

Simazine (202-132)

Metabenzthiazuron (222 MH+ 165)

MH+

Atrazine (216-174)
74

Simazine in surface water extract (50 pg injected on column)

API 150EXTM LC/MS System
(SIM MODE; m/z 202)

API 2000TM LC/MS/MS System

(MRM MODE; m/z 202-174)

75

Application Example 2

LC-MS/MS
Impurity profiling

76

Impurity/Degradation Product Profiling

Experimental Conditions:
4 Commercially available OTC Melatonin Tablet preparations LC Column: 4.6 x 50 mm Chromolith SpeedRod RP-18 Targeted analysis using MRM as survey for known impurities and EMS for profiling in IDA followed by EPI Confirmation of impurity/degradation product by MS/MS using EPI

77

Impurity/Degradation Product Profiling

Multiple MRMs (1) EMS (2) Enhanced Resolution

IDA Criteria Level 1

For known impurities, targeted MRM analysis can be used. At the same time an EMS survey scan can search for unknown impurities for unknown impurities, IDA triggered MS/MS and MS3 information can be collected to aid in identification.

Dependent Scan (s) Dependent Scan (s) Dependent Scan (s) Dependent Scan (s) Add to Exclusion List

IDA Criteria Level 2

Second Dependent Second Dependent Second Dependent Scan (s) Scan (s) Scan (s)

78

Impurity/Degradation Product Profiling

Targeted MRM Known Impurities
H 3C O

OH

H N

oxidation Impurities O 249 m/z

N H

HO

H N OH

di-oxidation Impurity O 265 m/z

N H

H 3C O

EMS Unknown Impurities

79

Impurity/Degradation Product Profiling

+ E P I (4 7 7 .0 0 ) C h a rg e (+ 2 ) C E (3 0 ): E xp 3 , 5 .5 6 8 m in fro m S a m p le 1 (S tre s s e d 1 0 n g /u L ) o f S tre s s ... 100% 95% 90% 85% 80% 75% 70% 65% 60% 55% 50% 45% 40% 35% 30% 25% 20% 15% 10% 5% 60 80 100 120 140 160 180 200 2 2 8 .0 1 6 0 .0 2 3 3 .0 2 6 7 .1 2 7 3 .0 220 240 260 280 300 m /z , a m u 3 2 9 .0 320 340 360 380 400 4 0 5 .2 4 1 8 .3 420 440 460 480 500 1 8 6 .0 2 0 2 .9 2 4 5 .0 M a x. 1 .6 e 5 c p s .

EPI Melatonin-Formaldehyde Impurity 477 m/z

H 3CO N N O H 3CO N H
4 7 7 .2

H N O

245

80

Impurity/Degradation Product Profiling

X IC o f + E M S : E x p 1 , 2 4 9 .0 a m u f r o m S a m p le 9 ( M e la to n in T a b le t (1 0 0 n g /u L ) I D A E M S _ E R _ E P I w i. .. 8 .3 e 6 8 .0 e 6 7 .5 e 6 7 .0 e 6 6 .5 e 6 6 .0 e 6 5 .5 e 6 5 .0 e 6 4 .5 e 6 4 .0 e 6 3 .5 e 6 3 .0 e 6 2 .5 e 6 2 .0 e 6 6 .9 1 1 .5 e 6 1 .0 e 6 5 .0 e 5 0 .0 1 2 3 4 T im e , m in 5 6 7 8 9 3 .8 1 7 .2 1

software can help with data analysis through sample and control comparison for degradation products
2 .8 6

M a x . 8 .3 e 6 c p s .

XICs of peaks present in Sample, but not Control. MS/MS information collected through IDA

81

Specificity of Detection for LC

UV chromophore
all compounds with a chromophore responding at the selected wavelength will interfere

MS molecular mass
interference from isobaric compounds chemical noise

MS/MS molecular mass and structural information

interference from structural isomers only
82

HPLC-UV Analysis of Sirolimus in Whole Blood

1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. Wash all glassware in methanol x2 and tert-butyl methyl ether (TBME) x2. Place 50L of internal standard (in methanol) into each screw-cap glass tube. Add 200L Sirolimus calibrator (5x concentrated in methanol) or 200L methanol for patient samples. Add 1.0mL blank whole blood to calibrators or 1.0mL patient whole blood. Add 2.0mL 0.1M ammonium carbonate buffer. Mix thoroughly. Add 7.0mL TBME and extract for 15min. Transfer upper layer to clean tube and re-extract lower layer with 7.0mL TBME. Combine TBME extracts and evaporate to dryness. Redissolve residue in 5.0mL ethanol and evaporate to dryness. Redissolve residue in 1.0mL ethanol, transfer to Eppendorf tube and evaporate to dryness. Redissolve residue in 100L 85% methanol. Inject 80L (equivalent to 800L whole blood) and analyse using two 4.6mm x 250mm C18 columns connected in series (30min run time).
83

Sirolimus: HPLC - UV Example

84

Immunosuppressant Sample Preparation LC-MS/MS Analysis

Whole Blood (10L - 40L) Add ZnSO4 Soln. Add 2 volumes MeCN with IS, Seal & Vortex Mix

Centrifuge, Inject 5 - 20L

85

Full Scan Acquisition Mode

Sirolimus: MS Spectrum
[M+NH4]+
100
821.5

822.5

[M+Na]+

[M+Li]+ [M+H]+
810.5

826.5 827.5

[M+K]+

0 790

795

800

805

810

815

820

825

830

835

840

845

m/z 850

86

Sirolimus:
Single ion monitoring (MS)

LC-MS (SIM) vs LC-UV

100

30g / L HPLC-MS SIR m/z 821

%

1.5 min

HPLC-UV

87

Full Scan Acquisition Mode

Sirolimus: MS Spectrum
[M+NH4]+
100
821.5

822.5

[M+Na]+

[M+Li]+ [M+H]+
810.5

826.5 827.5

[M+K]+

0 790

795

800

805

810

815

820

825

830

835

840

845

m/z 850

88

Product ion scanning

MS1

Collision Cell

Ar (2.5 3.0e-3mBar) MS2

Products Precursor

Static (m/z 821.5)

Scanning

The first quadrupole mass analyzer is fixed, or Static, at the mass-to-charge ratio (m/z) of the precursor ion to be interrogated while the second quadrupole is Scanning over a user-defined mass range.
89

NH4+

Product ion scanning

100

821.5

Mass spectrum from MS1

822.5

%
826.5 827.5 810.5

0 790

795

800

805

810

815

820

825

830

835

840

845

m/z 850

100

768

Product ion spectrum from MS2

%
576 548 558 718 750 786 821

0 200

250

300

350

400

450

500

550

600

650

700

750

800

850

m/z 900

90

Multiple Reaction Monitoring

MS1

Collision Cell

Ar (2.5 3.0e-3mBar) MS2

Precursor(s)

Product(s)

Static (m/z 821.5)

Static (m/z 768.5)

MS/MS : Compound-Specific Monitoring

91

Multiple Reaction Monitoring

Sirolimus LC-MS(SIM) vs LC-MS/MS (MRM)

100 100

3g / L
% %

30g / L

SIR m/z 821

0 100

0 100

MRM m/z 821>768

0.50

1.00

1.50

Time 0

0.50

1.00

1.50

Time

92

Questions please?...

93