National High Magnetic Field Laboratory at Florida State University

FT-ICR

Mass Spectrometry Advantages

Every molecule has mass!

Mass Defect as a Chromophore

Isotopes: Tracing/Quantitation

Attomoles

Interpretation (One Peak/Cpd)

(m/z)max(m/z)minm/z

Peak Capacity = m50%

(m/z)max - (m/z)min

m50%

Separation Method

Maximum # of Components

MaximumPeak Capacity

HP-TLC 6 25

Isocratic LC 12 100

Gradient LC 17 200

HPLC 37 1,000

CE 37 1,000

Open Tubular GC 37 1,000

ESI FT-ICR MS 525 200,000

B

h B= q B m=

BNMR or EMR ICR

Differential Amplifier

FT

100 150 200 250Frequency (kHz)

7+

8+

10+

11+

12+

9+

600 1000 1400 1800

12+

11+

10+

9+

8+

7+

m/z

0

80 240 400Time (ms)

ImageCurrent

BovineUbiquitin

10721071

mz

A= + B

2

Capillary tip

Taylor Cone

HV

Counter Electrode

Electrospray Ionization

Adapted from: Enke, C. Anal. Chem. 1997, 69, 4885-4893.

Heated Metal Capillary

Sleno et al.

178.10178.06178.04 178.08m/z

178.07233

178.08043

178.08489FT-ICR (IRMPD)

QqTOF (CID)

178.06107

C8H8N3O2+

m/z 178.06110

+ 13C1

12C713C1H9N4O+

m/z 178.08044

N

N

NHHO

O

N

N

NH

OH2N

C8H10N4O+•

m/z 178.08491

+N

O

C

N

H2N

N

C

CH2

C7H8N5O+

m/z 178.07234

N

N OH2N

N

NH2N

HO

N

NHO

N

N

m/z 220

-0.04

-0.03

-0.02

-0.01

0

0.01

0.02

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34

1H

2H13C 14N 15N

16O

31P32S 34S

12C

AtomicMass Defects

(Dalton)

0

2

4

6

8

10

521.0 521.1 521.2 521.3 521.4 521.5

(1 mDa Bins)

CcHhNnSsOo

S. KimRodgers

Mass

0

2

4

6

8

10

521.10 521.105 521.110 521.115 521.120

(1 mDa Bins) S. KimRodgers

Mass

CcHhNnSsOo

0

1

2

3

4

5

6

7

521.100 521.105 521.110 521.115 521.120

(0.5 mDa Bins)

Mass

S. KimRodgers

CcHhNnSsOo

0

1

2

3

4

521.110 521.112 521.114 521.116 521.118 521.120

(0.1 mDa Bins)

CcHhNnSsO

o

Mass

+/- 100 ppb =1 Composition per bin!

Negative-Ion APPI FT-ICR MS

-2 -1 0 1 2

100

90

80

70

60

50

40

30

20

10

Mass Accuracy vs. Relative Abundance (12,449 Assigned Masses)

Mass Accuracy (ppm)

Re

lati

ve

Ab

un

dan

ce

S. American Crude Oil

± 100 ppb

229225221

C17H21+

C16H33+

C16H17O+

C15H13S+

Measured Theoretical225.07326 225.07325 225.12733 225.12739 225.16375 225.16378 225.25769 225.25768

m/z 250200150 300

C15H13S+

C16H17O+

C17H21+

C16H33+

Raw Diesel Feedstock 1L Septum Injection

RodgersAndersenWhiteHendrickson

C8H7N

Isomers

N- H+

H N

+ H+

NH

+N

900800700600500400300200

~

-900 -800 -700 -600 -500 -400 -300 -200m/z

17,000+ Compositionally Distinct Components Resolved by High Resolution 9.4 Tesla Electrospray FT-ICR MS

6,118 resolved components

11,127 resolved components

0

Positive Ion ESI MSNegative Ion ESI MS

HugheyHendricksonRodgersQian

m/z 900800700600500400300

11,000 Peaks250 < m/z < 900

m/z492.475492.400492.325492.250

Expansionat m/z 492C3 SH4

Positive ESI FT-ICR MSS. Amer. Heavy Crude Oil

m2 – m1 = 3.4 mDa

HugheyHendricksonRodgersQian

m/z431.4431.3431.2431.1431.0

8 σ

m/z700640580520460400340280

*

63 Spectral Peaks above 8 σ62 Spectral Peaks Assigned* Not assigned

APPI FT-ICR MS

Incr

ea

sin

g

DB

E

Ke

nd

ric

k M

ass

De

fect

Incr

ea

sin

g

DB

E

0.1

0.2

0.3

0.4

0.5

0.0

0.1

0.2

0.3

0.0

0.4

0.5

300 400 500 600 700 800 900Nominal Kendrick Mass0.00 0.03

0.00 0.07

Crude Oil B

Asphaltenes Deposit B

m/z650550450350250

Canola Oil

Olive Oil

Soybean Oil

Negative-Ion ESI FT-ICR MS

WuRodgers

m/z285284283282281280279278277

C18:3

C18:2

C18:1

C18:0

C18:2

C18:1

C18:0

C18:3

C18:2

C18:1C18:0

Canola Oil

Olive Oil

Soybean Oil

Fatty Acids

WuRodgers

m/z890885880875

Canola Oil

Olive Oil

Soybean Oil

TriacylglycerolsC54:6

C54:5

C54:8

C54:7WuRodgers

m/z455.5455.4455.3455.2455.1

C30H47O3-

C29H43O4-

C25H43O7-

C24H39O8-

C27H35O6-

C23H35O9-

C26H31O7-

C22H31O10-

C22H32O8P-

C25H27O8-

C21H28O9P-

Negative-Ion ESI of Olive Oil

WuRodgers

0

4

8

12

16

0

4

8

12

O2

Canola Oil

Olive Oil

O3 O4 O6O5 O7 O9O8 O10

0

20

40

60

S NP

Soybean Oil

Acidic Species Detected by Negative-Ion ESI16

WuRodgers

m/z

435430425420415410405400

C27H45O2-

(δ-tocopherol)

C28H47O2-

(β, γ-tocopherol)

m/z429.4429.3429.2429.1

C29H49O2-

(α-tocopherol)

Tocopherols in Soybean Oil

O

R1

OH

R2

R3

Negative-Ion ESIWuRodgers

m/z415.6415.5415.4415.3415.2415.1

C28H47O2- (β, γ-tocopherol)

Canola Oil

Olive Oil

Soybean Oil

C19H27O10- C20H31O9

-

WuRodgers

m/z657.62657.58657.54657.50

C41H70O4P+

C41H69O6+

C45H69O3+

C42H73O5+

C46H73O2+

C43H77O4+

C47H77O+

Positive-Ion ESI FT-ICR MS

Soybean Oil

WuRodgers

m/z610605600

Canola Oil

Olive Oil

Soybean Oil

Diacylglycerols

C36:3C36:2

C36:1

C36:4

C36:5

WuRodgers

274

278

282

20

40

60

80

100

PureOlive

PureSoybean

C18:3

Fatty Acids

Relative Abundance %

C18:2

WuRodgers

876

880

884

20

40

60

80

100

PureOlive Oil

PureSoybean Oil

Triacylglycerols

C54:7

C54:5C54:6

Relative Abundance %

WuRodgers

NHO

OH

OOOH

OH

OH

OH

OOH

NHAc

OH

O

OH

OH

O

OH

OH

OH

ONHAc

OH

OH COOHOH

OH O

O

OO

GM1a

GM1b

NHO

OH

O

O

OOH

OH

OH

OOH

NHAc

OH

O

OH

OH

O

OH

OH

OHONHAc

OH

OH COOHOH

OH

O

OO

OH

HexNAc = N-Acetylhexosamine

NeuAc = N-Acetylneuraminic Acid

(Sialic Acid)

Hex = Hexose

200 400 800 1000 1200 1400600

Ceramide

Ceramide’ [M+2H] 2+

’’

’

’ )M-( ’)M-(

) M-(

’)M-( M-

’)M-(

)M-(

m/z

’(fr

om

0,2)

oo o o

O

OH

HNO

[M+2H]2+

’

’

’)M-(

’)M-(

)M-(’)M-(

M-( )’

200 ms IRMPD of GM1

100 ms IRMPD of GM1

HNO

OH

O

o o oo

200 400 800 1000 1200 1400600

[M-2H]2-

Cer

amid

e’

Cer

amid

eCer

amid

e’’

~1.5~

’

Sp

hin

go

sin

e-N

’ ’)M

-(

)M

-(

’)M

-(

’

’)M

-(

’)M

-(

M-

)’M

-(

m/z

oo o o

HNO

OH

OHNO

OH

O

o o oo’(

fro

m

0

,2)

’

ECD of GM1

Sugar

Lipid

M-x

Unknown

Amino Acid

0

0.2

0.4

0.6

0.8

1

0 500 1000 1500

Neutral Mass

and/or

HNO

OH

O

MassDefect

Sialic Acid

Hexose

HexNAcHN

O

OH

O

Hydrocarbon

Peptides fromDigest andStandards

McFarland

Atmospheric Pressure PhotoionizationFT-ICR MS

http://www.chem.agilent.com

Inlet

Nebulizing GasNebulizer

Vaporizer

Drying Gas

Capillary

UV Lamp

h To FT-ICR MS

M

+ H+

- e-

[M+H]+

M+•

Nominal Mass

    Odd Even

N0, N2, N4, … [M+H]+ M+•

N1, N3, N5, … M+• [M+H]+

Nitrogen RuleMcLafferty, Interpretation of Mass Spectra, 1993

APPI

[C30H41N1S1 + H]+

[C32H37N113C1]+ •

[C33H37N1 + H]+

m/z

448.310448.305448.300448.295448.290

4.5 mDa 3.4 mDa

Why Ultrahigh Mass Resolving Power?

South American Crude Oil3.4 mDa C3 versus SH4

4.5 mDa 13C versus CH

PurcellRodgers

[M+H]+

M+•

0

Ion Energy

Ion Trapping Period

Upper Mass LimitNumber of Ions

Mass Resolving Power

Scan Speed (LC/MS)

Highest Non-Coalesced Mass

0

7 T

7 T 9.4 T9.4 T

21 T

B (tesla)2121

B (tesla)

14.5 T

14.5 T

21 T

(2004)

(1995)(1985)

#2764 IT: 119.142 ST: 6.31 uS: 1 CS: 2 AMW: 1569.67 NL: 6.55E4F: FTMS + p ESI Full m s [ 0.00-6144.00]

0 500 1000 1500 2000 2500 3000 3500 4000 4500 5000 5500 6000

m /z

0

5

10

15

20

25

30

35

40

45

50

55

60

65

70

75

80

85

90

95

100

Relat

ive In

tensity

68.7004390.1465

772.3228 1192.9299 1624.7856 2084.4141 2499.5969 3175.2803 3667.2979 4082.1877 4361.0347 4825.6611 5262.9180 5699.0908

Glu fib 500K FT_041001162804 #1 RT: 0.00 AV: 1 NL: 2.98E5T: FTMS + p ESI Full ms [ 400.00-2000.00]

783 784 785 786 787 788 789 790m/z

0

5

10

15

20

25

30

35

40

45

50

55

60

65

70

75

80

85

90

95

100

Re

lativ

e A

bu

nd

an

ce

0 2 4 6

Time (s)

783 784 785 786 787 788 789 790m/z

Glu-Fib, 1 pmol/μL

785.842

786.343

786.845

HendricksonMackayQuinn

m/m50% = 914,000

External Calibration

Mass Accuracy: 50 ppb

Nanomate LTQ Broadband FT ICR MS

857 858m/z

Absorption

Magnitude Ubiquitin

BeuBlakneyHendricksonQuinn

[M+10H]10+

0

Ion Energy

Ion Trapping Period

Upper Mass LimitNumber of Ions

Mass Resolving Power

Scan Speed (LC/MS)

Highest Non-Coalesced Mass

0

7 T

7 T 9.4 T9.4 T

B (tesla)21

B (tesla)

14.5 T (2004)

(1995)(1985)

$1.5 M

$0.5 M$0.2 M

$?? M

NHMFLKBSIPNNL

21 T

(2009)

NSF

Florida State U.

NIH ExxonMobil

STINT (Sweden)

Schlumberger-Doll

Dow

Support for ICR at NHMFL

Fluor Canada

Saudi Aramco

ThermoElectronOil Phase DBR

Baker Petrolite

UnoCal

KBSI (Korea)

0

100

200

300

400

500

600

700

1976 1980 1984 1988 1992 20001996 2004

WorldwideFT-ICR MS Systems

NHMFL ICRProgram Starts