mass spectrometry with 252Cf fission fragment induced ionization

197

International Journal of Mass Spectromet*y und Zon Physics, 46 (1983) 197-200 Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands

LIQUID

CHROMATOGRAPHY/MASS

SPECTROMETRY

'=Cf

WITH

FRAGMENT

FISSION

INDUCED

IONIZATION HARTMUT

JUNGCLAS,

Kernchemie,

HARALD

DANIGEL

and LOTHAR

FB 14, Philipps-Universitat,

SCHMIDT

Hans-Meerwein-Str.,

D-355

Marburg,

FRG

ABSTRACT

252 Cf fission fragA time-of-flight desorption mass spectrometer utilizing ";iayir ;;l~~sc;"$e; y;;ic;$;;d ~~~~~a',i~n,E~a~~5~_~~~~~~s ;id :$;o:;ii;zs

.

complete mass spectrum (m/z l-1000) is obtained in approximately 1 minute. The via an spectrometer is combined with a high-performance liquid chromatograph involatile compounds in the interfacing disk samplin up to 12 fractions of effluent (= 0.5 cm3 min- 9 MeOH-H20) under rough vacuum conditions (= 1 mbar) in a vacuum-drying process. This combined LC/MS method has been applied for the quantitative analysis of antitumor drugs in human serum. INTRODUCTION The

ionization

lecular

ions

comparable Thereby liquid

from

to

all

of organic

involatile

chemical

these

compounds

the effluent

to all desorption we developed

ionization

a stepping

LC/MS All

in the

high

techniques.

urn) on high

LC/MS

quantitative

system

vacuum

analysis

of a

is

quantitative

potential

sample is

continously

(ref.5)

belt

with

as very

Technical thus

of

common

4, 51,

usually

drugs

3).

from

moving

(refs.

(f 10 kV).

already

antitumor

analysis

the

spectrometer

operated

1,2)

(ref.

high-performance

to transfer

periodically

published

of some

with

of the

Instead

voltage are

the

need

(refs.

de-

its per-

is reported

here.

ANALYSIS steps

displayed to add

to the

for

The

method

(-2

combined

is used

pressure.

quasi-mo-

techniques

be combined

desorption

induced

backings

formance

to

samples

fragment

of this

suited

yields

molecules

ionization

exchanging

thin tails

are

which

fragments

unstable

soft

disk

the fission sample

thermally 0th er

low vapor

via an interface

Cf fission

and

(HPLC)

with

252

by

and

(CIl

techniques

chromatography

organic

solids

of the

in Fig.1.

an internal

quantitative Prior standard

to extract

all compounds

biological

matrix

phase

with

mode

(e.g. isocratic

0020-7381/83/0000-0000/$03.00

LC/MS

to the HPLC (i.e.

elution 0

for pharmaceuticals is a preparation

homologous

of interest proteins).

analysis separation

or isotope

(pharmaceuticals, The

liquid

separates

labelled run

compounds

are

in order

compound)

metabolites)

chromatograph

the injected

in serum step

from

and the

in reversedin a retention

1983 Elsevier Scientific Publishing Company

198

compound

tn

add

internal

preparation

UV

absorption

HPLC

detector

interface

voc”“m-drymg

data

Fig.

1. Flowdiagram

sequence. out

of the

The effluent

splitting

into

pounds

are collected

drying

process.TweIve

twelve

fractions

disk transports trometer ionized

via

fieldfree

LC/MS tion

drift

from

a matrix data

path

the

spectrum

from

(typically

the thus

ratios.

mass

spectra

is discussed m/z

(ref.

plastic)

on a stepping a narrow

disk

At the

slit

and accelerated {he

into

COrl-

a vacuum-

in

sampling same

up to

time

the mass

data

with

m/z

repeatedly

measurement

on retention are

stored

this spec-

still pass

event-by-event

and

a

provides

by the HPLC column This

1 - 1000.

of projections

and

(-+ 10 keV1

ions

time-of-flight

All

in terms

al 1 nonvolatile

can be analysed

Information

sequence.

where

and with-

be artalysed while another is collected. -8 are of all sample molecules than 10

sample

Desorbed

50 cm),

sampling

less

continously

one perminute).

through

can

system

(2 urn aluminized

are mounted

fragments

for a given

three

a is

resul-

dimensional

sections,e.g.

reten-

51.

RESULTS

The experiments university.

foils

data

emerges

(= 1 mbar)

foils

MS

analysis.

MeOH-H20)

chamber

one sample

on m/z

K/MS

-1

successively

lock.

(-

mln

sample

therefore

of twelve

matrix

EXPERIMENTAL steem

thus fission

a vacuum

information

obtained ting

25?f

3

effluent

the samples

or sputtered,

recovered direct

sample

of the

cm

vacuum

on thin

(10s6mbar)

Utilizing

quantitative

(=0.5

a rough

matrix

TOF

investigating

an interdisciplinary Here

and Teniposide

we focus

(VM26).

on the

the

pharmacokinetics

co]laboration two

with

anti-neoplastic

of the

some

medical

agents

antitumor

clinic

Etoposide

drugs at out

IVP16-213)

199

dwt

586

Etopeside

moLwt 656

VP 16-213

The mass molecular the

spectra peaks

of the

at

chromatograph.

that

both

other was

drugs

which

serum

VM26 rated then shown

The

based

close

analysis

in fig.4. LC/MS

in

which

different

drugs

657.

Both

of the

the

were

orders

drugs

added

for

VP16/VM26

on chloroform

extraction

(compare

1 ml plain

redesolved

serum.

Each

experiments.

point

is based

Fig.

3 shows

X/MS and

(ref.

a mass

injected

The

on

showed

clearly

from

procedure

a calibration

curve,

5).

an extraction

method

ref.

(1 -100

6). VP16

extract

(MeOH:CH3CN:H20

samples

spectrum

quasi-

matrix

purpose.

The chloroform

serum

and

data

Th-e resulting

on two

dominant

separated

resulting

in 150 ul eluent

(50 pll on the chromatograph.

mixed

test

of magnitude two drugs

to

were

fraction

for these

mixed

2) display

complete

same

mixtures

three

(e.g. Fig.

we developed

to dryness,

injected

and

eluted

within

point

(20 ug) were

dependent this

with

is linear

At this man

589

pharmaceuticals, repeated

two pure

m/z

were

OH

VM 26

OH

of a serum

hu-

pg) and

was

evapo-

= 8:4:3)and

calibration analysed,

from

i.e.

sample

curve two

is in-

applying

procedure. LC/MS

Serum

ZEO

; 2M

Fig. 2. Mass spectrum of pure VP16-213 (mol .wt. 588). Dominant lines are m/z 589 and m/z 383 (fragment padophyltotoxin).

10

m/z

Fig. 3. Mass spectrum of serum sample taken 2 h after a 250 mg VP16-213 infusion. Internal standard 20 ug VM26 was added to 1.25 cm3 serum.

200

Fig. 4. Calibration curve for the antitumor drug VP16-213 in serum using 20 pg/cm3 VM26 as internal standard.

Ueight 0.05 o)l

ratio 05

(ug/Lcg) 1

2.5

5

20

t 2

1

UP16

5

in

10

20

Serum

50

loo

Cug/cm3)

DISCUSSION We conclude fraction

that

can be used

analysis.

When

in the

step.,

e.g. based

to

the

e.g. We

we are The

on m/z 116 The

alT

extraction

burg,

losses

indebted

compounds

standard

differ

and retention

in the

to

in the quantitative

retention

can imply

origins

from

ionized

compound

differences

time,

a further as in

any

LCfMS

instabi-

normalization (NH4)H2P04adcled

internal

chemical

standard behaviour,

time.

collaboration.

of the BMFT,

standard

labelled

due

to Dr. W. Achterrath, support

which

isotope

errors

are toll-ected in the same sanpling

process

(ref.51,

to Drs. K. Havemann

fruitful

which

as an internal

collection

use of an

possible

are grateful for the

financial

and

vacuum-drying

eluent. omit

crosswise

compound

lities

would

homologous

and K.H. Pfliiger, Medizinische For the

supply

FRG,

Klinik Mar-

VP16-213

and VM26

GmbH,

Neu-Isenburg,

is gratefully

acknowledged.

Bristol-Myers

Bonn,

of pure

FRG.

REFERENCES 1 2 3 4 5 6

Int.J.MassSpectrom.Ion Phys. 27,81(1976) R.D. Macfarlane and D.F. Torgerson, 0. Becker, N. Fiirstenau, F.R. Krueger, G. Wei and K. Wien, Nucl.Xstrum. Methods 139, 195 (1976) Heyden, H.R. Mar(Ed.), Soft Ionization Biological Mass Spectrometry, London (1981) and L. Schmidt, Org.Mass Spectrom. 17, 86 (1982) H. Jungclas, H. Danigel, L. Schmidt and J. Dellbrtigge, Org.m;Jss Spectrom. H. Jungclas, H. Danigel, (in press) J.J.M. Holthuis, H.M. Pinedo and W.J. van Oort, Anal.Chim.Acta 130,23(1981)