Mass spectrometry combined with mass spectrometry

Mass spectrometry combined with mass spectrometry

International Elsevier Journal Scientific Mass Spectrometry and Ion Physics, 45 (1982) of Publishing MASS SPECTROMETRY F.W. COMBINED I.J. McL...

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International Elsevier

Journal

Scientific

Mass Spectrometry and Ion Physics, 45 (1982)

of

Publishing

MASS SPECTROMETRY F.W.

COMBINED

I.J.

McLAFFERTY,

COHEN,

P.O.

Chemistry

Company,

DANIS,

WITH

AMSTER,

G.H.

Dept.,

Amsterdam

MASS SPECTROMETRY M.A.

BALDWIN,

C.J.

KRUPPA,

Cornell

323

323-329 in The Netherlands

- Printed

M.P.

PROCTOR,

University,

BARBALAS,

and

Ithaca,

F.

M.T.

CHENG,

S.L.

TURECEK

NY 14853

USA

ABSTRACT Tandem mass spectrometry (MS/MS) is a rapidly growing field of high promise for structure determination and quantitative analysis because of its sensitivity, specificity and speed. Rapid progress has been made in instrumentation, including the triple quadrupole and tandem double-focusing instruments, automated with on-line computers. Probably the most valuable application to date is the quantitative analysis of targeted compounds at trace levels with high specificity. However, the extra information available from MS/MS appears to be of particular value for the structure determination of large molecules, such as those best ionized by fast atom bombardment.

INTRODUCTION Tandem

can

individual enhance

the

For

the

sequent

former,

be the

gives

of

laboratories For

those

spectrum

from

MS-I

to

GC/MS

from

its

mass

dissociation

by

MS-I.

This

principal

compounds

of

in

areas

of

mixtures

ions

which

and

can

Spectrometry"

by

be

mass

from

used

of

components

for

MS-I

can

of contains

the

can

separation

either

which

so

These

other

their

description 1)

sub-

spectrometry.

leading

in

structural

quantitative

a score

these

MS techniques

and

specific

(ref.

of

mass.

ions

its

all

with

conventional

produced

up-to-date

than

as

a characteristic

primary

for

LC/MS, spectrometry)

species by

the

or

and mass

separated

other of

by more

analysis. the

field descrip-

research

field. structure

sample separated

0020-7381/82/0000-0000/$02.75

most

a detailed

Mass

or

ionized

spectrum

areas

the

the the

of

ions

component in

molecular

from of

mass this

"Tandem

in

(or

data

for

analysis

the is

dissociation

subject

molecular

molecular as

by

interested to

of

produces

of

MS/MS

a second

separated

spectrometry

analogous

mixture

a secondary

referred

utilizes

spectral

spectrum

(chromatography

compound

the

Readers

tions

mass

is

sample

separated

characterization

are

MS/MS method

the

mixture;

MS-II

mass mass

identification

MS/MS

a specific

then

of

1-4)

elucidation.

specific in

primary

quantitative

a separation

Thus,

(ref.

secondary

the

performance

structure

combine

(MS/MS)

measure in

application,

molecular

of

to peaks

analytical

that

spectrometry

(MS-II)

selector of

mass

determination, adduct)

ion

matrix

in

molecular 0 1982

from fast ions Elsevier

impurities atom then

be or

used

bombardment); provides

Scientific

to

anomolous the structural

Publishing

Company

separate peaks

the (such

secondary information.

324

Alternatively, (EI)

the

can

fragment

fragmentation

be

separated

ions

can

in then

spectrum

MS-I, be

and

used

such

secondary

to

as

mass

provide

more

from

electron

spectra

of

detailed

ionization the

separated

information

of

their

structures.

APPLICATIONS

OF MS/MS

Identification

of

Although than

for

components

present

commercial

chromatographic/MS

nesses

are

dentification obvious

one

peak

method

of

many

per

for

components.

for

identification.

valuable by collisionally

to

spectra,

matching

which

their CAD

by

Targeted

analysis

and

are

many

targeted

or

be

or

MS-I

and

sample ticularly

the

5),

of

LC/MS

spectra also

particu-

spectra

for be

this

have

producing

are

mass

can

i-

MS ionization

pro-

qualitatively

CAD

and

this

an without

spectra

are

the

the

produces

find

mass

for

(ref. Mun

and

component

secondary

ions

similar structural

identified

a data

by

base

a PBM search

of

algorithm

laboratory.

to

by

the

can

be

in

complex

both

MS-I

seconds

is

simple

complex

By for

MS-II

can

MS/MS. and

be

primary

from

GC/MS/MS mixtures.

and

ion's (mass

the

chosen,

A key require introduction

sample

fragment

be

selected.

changers

a particular

losses

which

can

usually

automatic

for

knowing

Continuous

containing

neutral

sensitivity

these

separations

fractionation while

very

mixtures.

and

possible,

specific

and

selective

for

Compounds

corresponding

helpful, for

speed,

chromatographic

latter.

Even

promising

for

10T3

samples the

selectivity,

of

conditions

speed;

corresponding

MS-II). probe

peaks is

liquid with

indicated

MS-II

the

references;

compounds ionization

compared

gaseous

interpreting

I.K.

GC/MS

(CAD)

fragment

advantages

sought,

MS/MS

advantageous

and

valuable

mass

of

minutes,

can

Staedeli

EI

in

For

analysis

the.optimum

advantage

been

normal

weakfor

obtained.

to

per

MS/MS

and

favorable

chromatography

peak

or

collected

of

compounds

helpful molecular

be

mixture,

dissociation

against

W.

has

In

expensive

strengths are

difficult

ion

Further,

have

compound MS/MS

the

is

unknown

more

their

can

usually one

CAD spectrum

spectra

implemented

is

least

activated

Unknown

information.

an

is

which

resolution,

it

at

some

duced

in

sufficient

while

produce

larly

-,lOOO

With

ways cases

sensitivities

components

component, can

ions

EI

many

MS/MS

other In

subpicogram

advantage.

which

in

complementary.

technique,

an

instruments

instruments,

surprisingly

particular

of

mixture

are

substructure CAD spectrum

differences

in

between

direct-introduction

LC/MS/MS

appear

to

be

par-

325 GC/MS/MS An

interesting

case

deacetylmetipranolol, metabolite

of

bolite

the

using

could

not

give

Although

MS/MS

the

peak

at

m/r

Ihe

not

266

for

314

to instrument,

267.

higher

additional obviously

was

314 -b 267. than

of

5c

or

3 orders

of

magnitude

at

lo-I1

structure

MS/MS

can

should

sen-

but

a base

the

in m/z

in

complex

to

lo-I1

triple

is

also of

the the

a linear

yielded

at

m/z

quadrupole

can from instrument

reaction

moni-

3081266

over

and

a range

a signal:

of

noise

coefficients

be

the

MS/MS

m/z

with

runs

the

are

produced

response

concentration,

mixture, suf-

by using

"multiple

a

inter-

grams,

the

processes

gives

sample

peaks

the

control

308

reducing

the

achieved

152

studies

Repeat

m/z

C3H6.6~ of

was

Computer

showed

Using

this

of

ratio

variation

be

portant

molecules ions

lationship,

elucidation provide

fragment

the

from ion

spectrum

of by

of

CAD spectra.

primary

mass

of

primary to

the

spectrum this

ion, the

second

fragment mass

and

and

is indicates ion.

ions, elemental

their

those

composition

if

in

the

the

particular whose

of

can

mapping,

found that

Of

im-

molecular

structures

For also

similarly

"Mapping"

indicates

compositions

information,

and

amounts.

spectrum

secondary

the

up

subnonomole

mass

structure

biological

elemental

of

add

molecular

their

primary

composition

in

EI

of larger

their

a larger

pairs

for

only normal

fragmentation

plementary

dimension

valuable

available

while

fragment

a second

unusually

from

characterized

formed

556,

corresponding

between

g.

drug. complexity

better.

Molecular

which

of

energies

these

curve

the

tris-trifluoroacetyl m/z

CAD of

improved

whose

alternating

than

3:l

were

CF3COOH.

A calibration

energy

Quantitation

For

used,

at

sufficiently the

a sufficient

the

peak

components

energies

of

of

MHf

loss

other

collision

advantageous. mode

low

study.

of

not of

of

meta-

detection

+CH2CH(OCOCF3)CH2N(COCF3)CH(CH3)2.

GC/MS/MS

collision

was

because

(CH4)

RNH-CH(CD3)2,

elimination

toring"

better

metabolic

Variation

at

of

using

which

GC provided

a small

this

ion

elimination

capillary

of active

of

multiple

grams,

rearrangement from

derivative,

and

more

the

with

low-level

ionization

MS/MS

noise")

the

hexadeuterio

useful;

quantitation the

sensitive

with

corresponding

limits

the

gives

from

IO-IO

sufficiently

metabolite

peak

setisitivity

of

GC/MS/MS

("chemical

ficient

level

Quantitation

GC/MS

below

Chemical

the 308,

m/z

ference

is

also

quadrupole

unique

low

metipranolol.

study

mixture,

of

a triple

the

capillary

limits

enhancement.

derivative

or

crucial

was

sample

sitivity

involves

drug

column

detection

for

the

6)

'-blocking

packed

sensitive

of

(ref.

(CH,),-phenyl(OH)OCH2CHOHCH2NHCH(CH3)2,

be an

important

secondary

first

ion

can

value

are

com-

masses of

re-

and the

CAD be

elemental

molecular

ion;

326

thus

the

structures

structure are

of

used

those

of

the

to

by

of

that

interpretation

of

electron

the

of through

ion

the

from

5~

-Pregnan-3

unknown

(ref.7).

weight

of

formula

mass

and

exact

and

the

had

evidence

of

proved

very

mass

spectra

and

delineating

the

the

EI

showed

spectrum

and

the

CAD The

tunately

reference

different

5~,3~-, the

strated the

the

m/z

84

peak

hydrogen

of

High

resolution C23H30N20

as

the

unknown

an

MS, and

chemical the

matched

that

of

Djerassi

technically

ionization,

Fortunately

m/z

m/z

information

STIRS

demoof

spectrum this

coworkers drug

of

reciprocal (ref.

9).

attracted overdose

established indicated

234

233,

of

deaths. the

below

molecular in

the for-

CAD spectrum

u-methylfentanyl,

and

and

5u,3u-, of

reference

product

in

to

C rings,

from

implicated

com-

characteristically

the

and

of

C2IH3402,

corresponding and

CAD spectrum

the

give

secondary

ions molecule,

B,

show

the

not

pos-

assignments

stereoisomers the

for by

narcotic

structural

A,

contribution

expected

compound, illicit

a peak

the

the

even been

CAD

5-positions.

give

Self-

could

substituent

of

molecular

However,

odd-electron

structure

I3C

il-

saturated

have

from

that

possible of

the

not

before,

Similarly,

demonstrated This

White.

publicity

this four

be

a molecular

the

would

and

to

5a,36-isomer.

structure

rearrangement

China

for

the

the

the

wide

of the

5;1,30-.

was

not

Comparison

corrected

this

does

corresponds

for

and

CH3COCH$H2+;

former

using

of

total

fragment

the

indicated

the 3-

often

a hypothetical

substituents.

pair

these

fragment

spectrum

prepared

the

e-

is

will

as

indicates

course

confirming at

ions,

indicates

Information

complementary

CAD spectra

unknown, that

formula

of

CI6H260

been

for

C5H80;

fragmentations

5ir.,3ti-, from

helpful

and

ZO-keto

have

used

8)

stereochemistry.

the

spectrum

latter.

never

the

for

corresponding

peak

(ref.

of

frag-

spectra

approaches

been

this

which

stereochemistry

CI5H2502

pOSitiOnS

and

spectra,

the

spectrum this

System

3-hydroxy

reference compounds

unequivocal

of

Retrieval

the

has

of

the

compounds.

mass

measurement

available,

primary CAD

These

steroid

examination

and

against if

mass

Computer

skeleton

sible

This

as

large

of

energy, is

developed

their

fentanye

ionization

internal none

useful,

For

ions

omitting

a quantitative

already

in

molecule. and

is

If be

CAD spectrum

the

fragment

ion's

those

molecules.

steroid

electron

Interpretive

steroid

of to

ion

the

often to

total

spectrum,

sought.

represented the

F -ol-20-one.

C2IH34O2.

matching

of

spectrum

Its

can

of

peices

applications

318,

Training

spectra

examination primary

by

spectra

the

primary

an

of be

similar

substructural

possible

lustrated

CAD mass

such

of

can

indicate

CAD mass

independent spectrum

qualitatively

ionization

lucidation

of The

processes,

structure,

are

can

CAD spectra

energy

reference

rules

together

structures.

low

its

a matching

mentation

The their

formed

characteristic

fragments

molecule.

elucidate

peaks

to

these

bold.

327

CH=CH2

3

However, EI

in

mass

contrast

spectrum

indicates

mary

the

latter

of

the

by

indicate

the

ions abundant

the

the

structures

one

is

ring,

the

Of

which

is

hydrogen,

by

the the

mass

of

first

forms

respectively. should

and

CBHBN+,

completing peak

be

Actually,

ions.

are

CAD spectra

146

pri-

peaks

products

in

m/z

such

The

their

value of

of

CBH50t

and

com-

CHsN=CHCHz,

ionized

particular

confirmed

and

do a molecular

consistent

CAD spectrum

the rule

finding

highest

of

electron"

of

odd-electron

spectrum,

CAD spectrum

"even

C13H16NOt.

to

more

primary

shown.

assignment

piperidine

complementary correspond

EI

the

CHBCH=C=O

these

peaks

dissociation

and

would

containing

in

The

further

example,

of

important

probability

CIBHIgN2+, loss

these

the

For

the

ions. low

from

ions.

producing

even-electron

structure

even-electron

rearrangement

as

above,

a relatively

C16H23N20f,

probability,

relatively

unknown

ions

fragment

reactions

low

be

fragment

represent two

However,

steroid only

will

even-electron spectrum

of

there

pairs

the

the

represent

that

plementary

to

of

with

the

IJ/Z

by

new

110

peak.

High

molecular One

weight

of

application ization these

the

most

to

higher

techniques spectra

compounds

or

no

from

such of

fragment such

such

high

as

as as

ion mass

fast

atom

formed

(M + Na)+

information. ions

to

Lhat

can the

(M + K)+.

weights

above

2000

Fortunately, distinguish

mass

spectrometry

made

possible

(FAB)

(ref.

bombardment

from

and

of

compounds

peaks

those

molecular

areas

weight

anomalous

such

peak,

of

research

molecular

contain

determination, adduct

compounds exciting

be

In

matrix

addition,

daltons

ion

species

ion-

However, structure

or

molecular FAB

provide can

its

for

the

often

CAD spectra molecular

10).

misleading

glycerol

is

be such

ion

spectra little

obtained as

(M + H)+

328

from

adduct

or

characterize

matrix

the

peaks, of

Instrumentation. structed 2.3

at

tesla

10

measurement

of

molecules.

This

precursor Human

Their

CAD

spectra

losses CAD

of

Na

spectrum

to

losses

were

of

found

observation

did

not

amino the

gastrin

ion

Vitamin

abundance is

the

the

ther,

the

lar,

directly

in the

others, ion.

evidence

axial abundant

cobalamin

assignment

that

m/z

from

the

axial 114

chain peak for

is

spectrum

show shows

chain

--m/z 1329°~ethylcobalamin

the 1329

132

CAD

spectrum

to

can is

and

be

have

large

are

a mass

of 971

similar.

with

ex-

m/z

peak of

rela-

the

the

for

loss

cyano.

Fur-

surprisingly losses

is

peaks

of

(HPO3). 57,

these

the

times

that

1270

of

IJ/Z

mass

time

3-10

loss

80

for of

and

the

the

and

confirmed

closely

high

sequential

(sugar),

IJ/~

most

higher,

a very

the

11).

clearly

to

through

and

indicated in

(ref.

is

of

sections but

and

was ions

while

cross

with

1329 corresponds --m/z of IJ/Z 1355, 1329,

+ H),

FAB

from

products,

peaks

CAD spectra

CAD

412

shows

these

results

cholecystokinin

peptide

cleavages

of

the

(M + H)+

their

substantially

no

encouraging

IJ/Z

1404,

and

corresponding

small

cyanocobalmin

first

their Its

that

indicating

the

change The

the

2134.

molecule,

respectively,

1387,

CsI of

spectrum

and

Similar

CAD

12%,

to

(M + Na)+,

peaks

these

mass;

of

the

of

collectible

1368,

H)+,

Particularly

and

spectrum

but

ionization

backbone,

1355,

CAD fragmentation

most

not

FAB.

of

Surprisingly, 2.7%

peaks

to

(dimethylbenzimidazole group

1.6%.

FAB

these

(M + H)+

strong

2.3%

1341,

sample

of

26,

The

of

the

increasing

chain,

1329,

abundances of

with

a FAB 2118,

N-terminus.

for

six

""80%

peaks

sequence

sections

up the

of

with

(M t

spectrum

and

peaks

losses

sequence

bradykinins.

are

side

m/z

FAB

yields yields

B-12.

at

posure

as

three

dissociation

represent

centered

from

C-

10,000

possible

2096,

to

eight

the

CAD cross

seriously

the

peaks

1355

and

shows

both

CAD spectra

Met-Asp-Phe-NH2 from

peak

cm

ions.

IJ/~

intense FAB

spaced

section,

correspond

to

a 60

transmission

produces at

of

up

ion

makes

product

the

ions

sequential

CAD cross

less

to

from

the

regularly 8966)

region

these giving

(M + H)+ acids

decrease

metastable

of

the

contrast

that

tripeptide

tive

of

In

angiotensin,

the

MI

K.

for

peptides,

2096

or

of

(m/z

con-

addition

retain

give

showing

mass

that two

FAB

to

spectrometer the

heptadecapeptide

high

latter

to

collectible

the

indicate the

analysis

high

This

in

with

mass

MS-II

yielding

peaks

(M + K)+,

in

mass

+ ions

unusually

information

11).

potential by

Cs35134

(15-Met).

three

(ref.

improved

the

produced

an

ions

gastrin

only

and

shows '

been

allows

of

fragmentation

ions

accelerating

a CAD spectrum

CS35I34

provide

double-focusing

has

which

' ions csn+lln MS-I analysis

15000.

with

12)

KV ion

to these

tandem

(ref.

magnet

using

efficiency. m/z

The

Cornell MS-I

daltons

and

structures

simiof

146

The

final

-CH(CH3)CH2NH-, at

reference

m/z

914. CAD

The spectra;

329 ACKNOWLEDGMENT

Army

We thank

the

Research

Office,

National Durham

Institutes (Grant

of

Health

G29-79-C-0046)

(Grant for

GM-16609)

and

generous

financial

the

support.

REFERENCES

9 10 11 12

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