AND APPLICATION OF PARTICLE BEAM LC/MS
ON THE QUADRUPOLE
ON TRAP MASS SPECTROMETER
BRENT LAMAR KLEINTOP
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA
N PARTIAL FULFILLMENT
To Mom and Dad in appreciation for al
wish to express my sincere gratitude to my advisor
night and guidance throughout my graduate studies.
to acknowledge and thank the members of my
Winefordner, Dr. Vanecia Young,
wish to acknowledge Dr
Dr. Jim Boncella, and Dr
a valuable member of the
for many fruitful discussions,
both scientific and personal.
also wish to acknowledge those who provided funding for my research
includes the NASA Amtech Joint Agreement,
An extra thank you goes to Dr.
Thomas Behymer of the U
EPA/EMSL for his willingness to assist at any time.
have great difficulty finding the words to express the gratitude
the many special friendships
have shared while at UF
. These include fellow Yost
and Nate "Air Quaker" Yates
, as well as fellow graduate
students Kevin Kinter, Keith Palmer
and Larry "Moti Oti"
to make an extra acknowledgement of Don Eades, whom
having as my coworker on this particle beam project, and Nate Yates, for always
knowing how to change the
trap software to suit my needs.
wish them and
everyone else too numerous to mention the best of
uck in their careers.
we have shared will never be forgotten.
These were truly the
Finally and most
express my sincerest
ove and thanks to my
It is not possible to express enough thanks to my mother and father, who
have worked their entire life to provide me with whatever was necessary to help
me achieve my goals.
've never felt closer to my brother Gary whom
watched change careers,
and start (and expand) a family while
and Jarrett for not letting the family
become boring wh
Although hundreds of miles have separated me
from my family during recent years,
have never felt closer to all of them.
TABLE OF CONTENTS
S S a S S S S ii
S. . 1
The Quadrupole Io
General Theory of
Trap Mass Spectrometer
SS 5 2
on Motion in the QITMS
Principles of Operation . .
LC/MS using the QITMS .
The Particle Beam LC/MS
Overview of Organization of Dissertation
IMPORTANT PARTICLE BEAM
* S S
* S S S
Nebulizing Helium Flow Rate
. S S S
* S S S S
Desolvation Chamber Pressure
EVALUATION OF STRATEGIES TO MINIMIZE ADVERSE
EFFECTS OF RESIDUAL SOLVENT
Experimental . . . . .
Problems Caused by Residual Solvent
Strategies to Minimize Adverse Solvent Effects
Additional Stage of Momentum Separation
Elevation of rf Level . . . .
Resonant Ejection of Single Ion Species
in of Solvent 14
on Ejection Methods
*. a .a .a a .a
* a a a a a .a
* a .a .a a a a
* a a a a a a a a a
* a a
* a a a
* a a
* a a a
APPLICATION OF LC/PB/ITMS TO ANALYSIS OF PESTICIDE
MIXTURES AND COMPARISON TO OTHER LC/PB/MS
Results and Discussion
El Mass Spectra . . .
Instrument Detection Limits
Instrument Calibration Curves
Precision of Peak Areas .
Alternating EI/CI Acquisitions
Conclusions . . . .
IMPROVED QUANTITATION USING COELUTING
Benefits of Axial Modulation
Selective Storage Scans
Liquid Chromatography/Mass Spectrometry
AND FUTURE WORK
for Future Work
Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy
AND APPLICATION OF PARTICLE BEAM-LC/MS ON THE
ON TRAP MASS
Brent LaMar Kleintop
Chairperson: Richard A.
Major Department: Chemistry
This dissertation presents results from
investigations of coupling a particle
beam (PB) interface for liquid chromatography/mass spectrometery (LC/MS) with
on trap m
Unlike most other LC/MS
, PB interfaces are capable of producing
onization (El) mass spectra which may allow identification of unknown
the acknowledged sensitivity
of the QITMS
might provide lower detection limits and provide the impetus for development of
future benchtop LC/MS/MS instrumentation.
residual solvent introduced into the QITMS by the interface caused
space charging and a large degree of solvent-chemical ionization (CI).
stage of momentum
in the interface reduced the
amount of solvent which reached the interface; however
it was also necessary to
eject solvent ion
from within the trap prior to m
Two modes of ion
demonstrated to efficiently eject solvent ions.
This allowed generation of classical
spectra of several pesticides which compared favorably to both library
spectra and spectra acquired from solid
analyses (i.e. a "solvent-
free" method) of pure compounds.
The ability to perform
both isocratic and gradient elution
analyses of pesticide mixtures has been demonstrated.
The results obtained from
mits of detection and precision; however
n the relative intensities of fragment ions
at large analyte concentrations was observed
some minor differences
i. Also, a larger degree
on the ion trap system.
Quantitation using coeluting isotopically labeled internal standards (IS) has
also been investigated.
selective storage scan
A new method of rapidly performing alternating m
has been demonstrated to provide excellent
precision over four orders of magnitude
for gas chromatography/QITMS analyses.
improvements were largely
attributed to the coeluting
which improved analyte transport through the interface.
Since becoming commercially available in 1983,
the quadrupole ion trap
mass spectrometer (QITMS) has seen widespread use in both fundamental and
mass spectrometric studies.
commercially as a
benchtop mass spectrometric detector for gas chromatography (GC),
spectrometry (LC/MS) systems.
In this dissertation
, results from
focused on coupling a particle beam (PB)
nterface for LC/MS
with a QITMS wil
problems were encountered which were caused by the
solvent into the QITMS by the
strategies were investigated to minimized these adverse solvent effects and allow
environmentally significant compounds which compared favorably with
The capabilities of the LC/PB/QITMS system were then
evaluated by applying it to the LC/MS analysis of pesticide mixtures.
analytical figures of merit obtained on the LC/PB/QITMS system were compared
ft -- I *- 1 t I I a -
a CIA .-
operational modes were
improve quantitation using isotopically
labeled internal standards.
introductory chapter provides a historical perspective, as well as a
presentation of the operating principles of both the QITMS and the PB interface
and also provides an overview of the organization of this dissertation.
The Quadrueole Ion
A summary of the history of the development of the
on trap was
n 1953 by Pau
quadrupole mass filter (Paul and Steinwedel,
deas were also
proposed that same
year by Post and Heinrich (1953) and by Good (1953) at the
University of California,
spectrometer by Fischer (19
The quadrupole ion trap was first used as a
)59), who used mass-selective ion detection
using resonant absorption to produce a m
spectrum of krypton.
resonant absorption involves the application of an auxi
ary alternating current (ac)
voltage of a particular frequency across the endcap electrodes of the ion trap.
An ion undergoes resonant absorption when its fundamental frequency of motion
!FI *lk a earvAn ae *ha #ranm aanr"ri *ha
IFIAAIIIAC Crnhr Ihnhu\
su. *r Irr~ ^ ** ___
Summary of the stages of development of the QITMS.
also depicts the time periods of each operational
used during these stages of development.
First disclosure (Paul and Steinwedel)
Use as a mass spectrometer (Fischer)
of ions for rf spectroscopy
(Dehmelt and Major)
ons to an external detector
(Dawson and Whetten)
Use of QUISTOR as ion source for
quadrupole mass filter (Todd et al.)
as GC detector (Armitage
Disclosure of ion trap detector (ITDTM)
(Stafford et al.)
Ion trap mass spectrometer (ITMSTM)
(Kelley et al.)
When this occurred
, ion signal was detected by measuring the
power absorbed by the ion from the resonant excitation circuit.
Many early applications of the ion trap
involved its use with a wide variety
of gas-phase rf
The evolution of the ion trap as a mass spectrometer began in
1960s with the development of mass-selective storage
milar to the operation of the quadrupole mass
n this mode, appropriate rf and direct current (dc) potentials were applied
to the electrodes such that only a very narrow mass range of ion
Ions were detected by ejecting them from the
trap through a series of
n one of the endcap electrodes to an external detector such as an
Although this method was usefu
for analysis over a fairly
narrow mass range,
it was cumbersome for acquiring a full m
required repeating the process of mass-selective storage for each mass over the
range of interest.
as a mass-selective
filter. For th
Todd and coworkers coined the
"_yadrupole ion store" or QUISTOR for the ion trap
(Lawson and Todd,
The QUISTOR/quadrupole combination was successfully used throughout
instrumentation (Kishore and Ghosh,
1979) and operational modes (Fulford and
1978; Fulford et al.,
1980), and to study a variety of ionic processes such
as ion/molecule reactions
(Bonner et al.,
1974) and ion energetic
development of the mass-selective instability mode of operation (Stafford et al.,
Kelley et al
n this mode, ions covering a broad range of mass-to-
charge ratios (m/z) are trapped by application of an appropriate rf voltage to the
A mass spectrum
is obtained by
nearly increasing the amplitude
of the rf voltage, which causes
ons of increasing m/z to be ejected through holes
in one of the endcap electrodes and detected by an external electron multiplier.
This new scan method provides a much simpler and more rapid approach for
of mass spectra.
a single ionization
, an entire mass
spectrum can be obtained.
The development of the mass-selective instability scan was the impetu
ed to the commercial introduction of the ion trap detector (ITDTM) by Finnigan
n 1983 as a benchtop mass spectrometric detector for GC.
and sensitivity of the
1985 Finnigan MAT
The low cost
n widespread applications for routine GC
introduced the more versatile ion trap mass
STM) which allows both routine MS and more advanced MS
Interest in ion trap technology increased dramatically after the
operation of the QITMS have been the subject of several reviews (Cooks and
Nourse and Cooks,
1990; Cooks et al.,
Todd and Penman
A recent book also reviews the
and applications of the quadrupole ion trap (March and Hughes,
General Theory of Ion Motion
n the QITMS
is operated by app
cation of an rf voltage to the ring electrode
which creates a three-dimensional electric field within the
are either trapped or ejected from the trap depending upon the m/z of the ion and
the applied electric fields.
The motion of an ion in a quadrupole electric field can
be described by equation
which is also known as the Mathieu
- 2 q cos 2) u
where u represents either the radial (r) or axial (z) axis,
and a. and q, are the Mathieu stability parameters.
phase of the rf
are related to the
m ro Q2
Since ro and
= V ()
evident that the Mathieu parameter
q, is related to the rf potential applied to the ring electrode and that for a given
has a unique value of q,.
, au is related to the
applied dc potential and for a given dc potential,
has a unique a. value.
meaning the displacement of an
(the center of the trap),
ion periodically passes
or whether the displacement
which yield stable solutions to the Mathieu equation,
it will be trapped and wi
representations of stable solutions in a and q space are called stability diagrams.
The stability region which is relevant to the QITMS is shown in Figure 1
with values of a and q which
ie within the stab
ity diagram wi
The stability region is bounded by so called
=0 or 1
a complex function of a. and q, and determines
the secular frequencies
The trajectories of ion
with the same l,,
value have the same
, the amplitude of the trajectories is governed by the
a and q values of each m/z.
The trajectory of an ion in the rz plane is illustrated
n Figure 1
It has the form of a
Lissajous figure composed of two secular
frequency components with a superimposed high frequency ripple resulting from
higher order harmon
of the ion's
Principles of Operation
of the more
iar quadrupole m
A schematic of the Finnigan
STM which was
used for most of these studies is
in Figure 1-3.
ists of a hyperbolic
ring electrode and two hyperbolic endcap electrodes,
radius of the ring electrode and
where ro represents the
is the half distance between the two endcap
lines na o
0.0 q, eject = 0.908
0.40 0.80 1.20 1
ity diagram for the quadrupole
on trap plotted
with a,q parameters
ocated within the region of stability wil
trajectories and be effectively trapped.
Lissajous form of the trajectory of a trapped ion in the rz
'he figure is from a photograph of an illuminated charged
in a quadrupole
uperimposed high frequency ripple (Todd,
Electron Multiplier Detector
Schematic diagram of the Finnigan ITMSTM used throughout these
It has recently been disclosed however, that al
ion traps are actually "stretched" in the axial,
z, direction such that r
(Louris et al.,
1992) where r,
1.000 cm and zo
= 0.783 cm.
The quadrupolar trapping field of the QITMS is generated by application
of an rf voltage to the ring electrode.
, the frequency of the applied
rf signal is 1.1 MHz.
The generated field is actually not an ideal quadrupolar field,
but has contributions from higher order fields as we
Octapolar and dodecapolar
field contributions result from the "stretched" geometry of the electrode spacing.
Imperfections in the electrode surfaces and holes for injection of ionizing electrons
and ejection of ions
introduce hexapolar contributions to the trapping field.
Studying the effects of these nonlinear resonances has been the subject of many
investigations (Guidugli et al.
1992; Eades and Yost,
production of positive ions
n mass spectrometry (Duckworth et al
, ionizing electrons are created from a heated rhenium filament located
outside one of the endcap electrodes.
Electrons enter the trap by gating an
180 V for a period of time equivalent to the ionization time
(typically 1 ms) and ionize the neutral
present within the trap.
Ions are effectively
trapped in the
TMST when the values of their az and q, parameters fall within the
stability diagram (Figure 1
in the rf-only mode (az
with q, values less than q,
= 0.908 wil
be effectively trapped and stored.
pressure of a light buffer gas,
mtorr of helium.
ions and helium atoms serve to kinetically "cool" the ions, which causes the ions
to migrate toward the center of the trap.
n dramatic increases
sensitivity and resolution (Stafford et al.,
instability scan method (Stafford et aL
During the mass-selective instability
scan there is no dc potential
applied to the trap
such that all ions
possess values of q, which lie along the qz axis of the stability diagram.
inversely proportional to m/z
ions of lower m/z are located
towards the right of the q, axis.
Ions are "moved" along the q, axis by
ramping the rf potential,
*, (qz = Vfrom Equation 1-5) until they become unstable
at the stability boundary
n sequential ejection of
m/z from the trap where they are detected
by an external
of 180 psec/u.
. Mass spectra are recorded on the
TMSM using a scan rate
mass analysis, an additional axial modulation
is applied across the endcap
applied at slightly less than half the rf drive signal at approximately 6 Vp.
prior to ejection,
come into resonance with the axial modulation signal,
which increases the amplitude of the
on's trajectory in the
results in the ions being more tightly bunched upon ejection,
resolution and detection efficiency (Weber-Grabau et al.
simultaneous dc, rf,
and ac voltages.
For example, isolation of a single m/z
narrow range of m/z's
n the trap can be accomplished
by application of an
appropriate combination of rf and dc voltages such that the m/z of interest is near
the apex of the stability diagram (Figure
-1) at q,
= 0.78, az
= 0.15 (Weber-
, a "two-step"
described where higher mass ions are ejected at a working point along the $z
0 stability edge and lower mass ion
are ejected at a working point along the $z
stability edge (Gronowska et al.,
Yates et a/.
Tandem mass spectrometry (MS/MS) experiments are performed by first
mass-selecting a particular precursor ion.
supplemental resonant excitation
across the endcaps at the secular frequency
n the amplitude of its trajectory
in the z-direction.
The amplitude of the
resonant excitation voltage,
, is kept low enough to avoid losses due to
The kinetically excited precursor ion
Studies have demonstrated up to 10059
CAD efficiency and
efficiencies approximately 14 times
higher than triple quadrupole
instruments which suggests that 'the ion trap may be
the most sensitive MS/MS
experiments can be performed
by mass-selecting a
experiments up to MS12 have been successfully demonstrated
(Louris et al.
LC/MS usina the QITMS
The combination of high-performance liquid chromatography (HPLC) with
potential for analyzing a wide variety of compounds. Although HPLC does not
provide the chromatographic efficiency of capillary GC, analytes remain in the
decomposition is not a problem.
This makes HPLC-based methods an appealing
choice for the analysis of a wide variety of thermally labile, nonvolatile, and polar
compounds which are either not amenable to GC or require derivatization for GC
, it has been estimated that only about 20% of al
compounds are amenable to GC and GC/MS analyses and that HPLC-based
methods should be capable of analyzing the vast majority of the remaining 80%
(Budde et al.
The major challenge involved with using a mass
detector is the design of a suitable
spectrometer as an HPLC
nterface which can convert the high pressure
Vaporizing this quantity of liquid generates a volume of gas that must
be substantially reduced to maintain the high vacuum (typically 1
x 10.5 torr) of
represented the first successful LC/MS interfaces.
However, the moving belt was
mechanically complex and subject to memory effects, while DLI required splitting
a large portion of the LC eluent which limited the sensitivity of the technique.
, the development of a "new generation" of improved aerosol-based
(Blakley et al.,
(Willoughby and Browner,
and electrospray (Fenn etal.
broadened the applicability of LC/M
, ion traps are seeing
initial attempts to
nterface the widely used thermospray (TSP)
nterface with an
improved using a differentially pumped ion source (Bier etal.,
(ESP) interfaces have seen the greatest number of LC/ITMST applications
Berkel et aL
. Since ESP yields
multiply charged ion
, even high molecular weight biological compounds are well
within the working mass range of the
= 650 u).
Ion trap systems
TSP and ESP require the use of an ion injection system because
ions are generated within the interfaces,
external to the trap.
A number of features of the
TMST make it particularly attractive as a mass
TSP and ESP generated ions.
These include high sensitivity
ability to perform MS" experiments with high CAD efficiency,
perform kinetic measurements.
and the ability to
Since TSP and ESP are known as "soft" ionization
techniques which produce predominantly molecular ion
the capability to obtain
structural information from MS" experiments is particularly appealing.
The Particle Beam LC/MS
nterfaces based upon aerosol formation.
The primary advantage of using an
nterface is that solvent can evaporate far more rapidly from
droplets than from bulk solutions because of the
much higher surface area-to-
volume ratio (Browner et a.,
The original design was disclosed in 1984
and was named MAGIC for monodisperse aerosol generation interface for
---- 0 vww ^^^w
chromatography (Willoughby and Browner,
As variations based on MAGIC
they became simply known as particle beam
Although differences exist between various commercial PB interfaces, all
consist of a nebulizer and some type of desolvation chamber
coupled with a
separator which removes solvent vapor from the beam of largely
desolvated particles. A schematic of the prototype
n these studies
employs an additional third stage of pumping in the momentum separation region
to further reduce the amount of solvent which reaches the ion trap.
The interface works by first converting the LC
effluent into an
having a narrow range of droplet diameters.
the liquid with a high concentric flow of heliu
The nebulizer in this interface shears
m. The droplets then travel through
a heated desolvation chamber where volatile solvent is vaporized resulting in a
of solvent vapor
This mixture then passes through a beam collimator which form
nterface consists of three stages of pumping which
are separated by a series of skimmers.
Solvent and helium are pumped away
the momentum separator while the more massive and higher momentum analyte
particles are transported through a transfer probe
nto the mass spectrometer
The transfer probe was
TMST via a
transfer line probe which allowed insertion and removal without disturbing the
high vacuum of the ITMSTM
The beam of particles is "flash" vaporized when it
collides with the hyperbolic surface of the filament endcap upon
TMST analyzer cavity.
The resulting vapor then travel
toward the center of the
trap where it is ionized
with reagent ions produced from a reagent gas.
operating parameters and optimization of the interface will be described in greater
Particle beam interfaces have generated recent widespread interest largely
because of their ability to produce classical El m
spectra for a wide variety of
biological interest (Behymer et al.
Voyksner et al.,
of El spectra is advantageou
because the fragmentation patterns found
a typical goal of many LC/MS
El spectra also
can be compared to existing library spectra for the identification and confirmation
of unknown compounds.
TSP and ESP interfaces produce spectra unique to the
nterface which yield predominantly molecular weight
tandem mass spectrometry for obtaining structural
information has been reported
(Smith et al.
Van Berkel et al.
Coupling a PB interface with a QITMS would appear to be an appealing
choice for many applications employing LC/MS.
The ability of PB to generate El
spectra affords the
identification of unknown compounds,
a typical goal of many
types of analyses, while the
sensitivity of the
should also provide
all commercially available PB systems employ quadrupole
analyzers due to their ruggedness and low cost.
Although the coupling of
ions from an external ion source (Bier et al.,
1991) has been demonstrated,
rigorous studies have been performed which thoroughly evaluate the analytical
capabilities of a PB/QITMS system.
Overview of Oraanization of Dissertation
In this dissertation, results from investigations of coupling LC with a QITMS
via a PB interface are presented. A historical perspective and a review of the
basic operating principles of the QITMS and the PB interface are provided in this
The optimization of important PB operating parameters and
instrumental modifications are discussed in Chapter
Chapter 3 presents results
of several methods investigated to minimize adverse effects of residual solvent.
of the PB/ITMSTM
system to the analyses of pesticide mixtures
PB/ITMSTM system with a PB system which employs a quadruple mass analyzer.
investigations of several ion trap operational modes to
improve quantitation using
analyses are discussed
standards for both GC/ITMS
n Chapter 5.
conclusions and perspectives for
future work are presented in Chapter 6.
IMPORTANT PARTICLE BEAM
n Chapter 1
particle beam (PB)
nterfaces have attracted
environmental (Brown et al.
1990) and biological interest (Voyksner et al.,
because they typically
fragmentation patterns which can reveal structural information.
patterns of th
spectra can then be compared to reference spectra using readily
available computer library spectral searches for the identification and confirmation
of unknown compounds, a typical goal of many LC/MS analyses.
Although the important PB operating parameters are well known (Voyksner
between commercially available PB
nterfaces and MS
necessitate the need to characterize th
parameters for each system
n order to intelligently develop effective LC/MS methods.
and characterizes the important PB operating parameters for the LC/PB/ITMS
modifications were made to both the interface and the analyzer.
and the results of these modifications wil
also be discussed.
Two HPLC systems were used for these characterization studies.
NJ) 6-port manual injection valve with a 10 pL sample loop.
chromatograph fitted with a Rheodyne (Cotati,
injection valve with a
5 pL sample loop.
Solvent compositions and flow rates used are indicated in the
were performed using flow injection analysis
(FIA) of analytes.
interface was a prototype Finnigan MAT (San Jose, CA) particle
available interface in that it employs three stages of pumping in the momentum
separator region to further reduce the amount of solvent which reaches the ion
Supplemental helium was added to the third stage of momentum separation
to prevent backstreaming of pump oil from the mechanical pumps caused by low
Typical pressures in the three stages of the momentum separator
TMSTM via a V1/ probe lock which allowed insertion and removal of the interface
without disturbing the high vacuum of the mass spectrometer.
The probe tip was
- 14" away from the
on trap entrance to prevent high pressures inside
The Rulone fitting normally
n the entrance hole to electrically
isolate a probe from the endcap was removed so a larger entrance
nto the ion
trap would be present.
The mass spectrometer used in these experiments was a Finnigan MAT
(San Jose, CA)
The vacuum chamber was maintained at 100C for al
El was performed within the ion trap; detection was accomplished
with an electron multiplier set to yield
a single conversion
dynode at 0V.
No dynode voltage was used because the dynodes caused high
noise problems on the
STM at the time th
experiments were performed.
Partial pressures of residual solvent were typically
- 1 X 106 torr (uncorrected) as
measured by a Bayard-Alpert ionization gauge (Granville-Phillips,
mounted on the vacuum chamber.
Helium was added into the
chamber to produce typical operating pressures of 1
Five microscans were averaged on the
X 104 torr (uncorrected).
Sm and returned to the data system
as a single scan.
Axial modulation (Weber-Grabau et al.
1987) was employed at
530 kHz and 6Vpp during the mass-selective instability scan to improve mass
resolution and sensitivity.
Samples and Reaaents
The structures of the standards used throughout these characterization
studies are shown in Figure 2-1.
Also shown are molecular weights and the
fragmentation which yields the base peak of each compound's
El mass spectrum.
Agency (Pesticide Chemical Repository,
and rotenone were purchased from Sigma (St. Louis,
All standards were
used without further purification. HPLC g
obtained from Fisher Scientific (Pittsburgh,
;rade methanol and acetonitrile were
PA) and reagent water was obtained
from a Milli-Q water purification system.
temperature, desolvation chamber pressure, and source target temperature.
of these parameters were
investigated to determine what effects they have on
investigated with various commonly used LC mobile phases, including methanol,
, and acetonitrile/reagent water mixtures and different flow rates.
assistance of Mr. Donald Eades during these characterization studies is gratefully
+ \ _
Nebulizina Helium Flow Rate
The nebulizer used in the PB interface was a glass Meinhard (Santa Ana,
CA) nebulizer which consists of a pair of concentric tubes.
While the position of
the inner capillary tube is adjustable on other commercial interfaces (Apffel and
its position is fixed in the Meinhard nebulizer
. The nebulizer was
installed onto the desolvation chamber through a Cajon fitting which ensured
a vacuum-tight seal.
The HPLC eluant entered the nebulizer by passing through a length of 0.1
deactivated fused silica capillary column,
inserted into the
inner tube of the nebulizer
. A high flow (
of He was
through a gas port and flowed through the outer tube.
the inner capillary,
iquid flow exited
the concentric flow of He dispersed the liquid flow
aerosol consisting of a distribution of micron-sized droplets (Browner et al.
The aerosol was then swept into the desolvation chamber by the flow of He.
The flow of He into the nebulizer was controlled
pressure of the regulator on the He supply.
by varying the
The plots in Figure
varying the head
affected the signal
different mobile phase compositions.
The data points represent the average peak
areas obtained from triplicate injections of 20 ng diuron.
The peak areas were
calculated from the extracted ion profile of the base peak (m/z 72) of the El mass
spectrum of diuron.
The curves are asymmetric and show that Door sianal
- . . U tEl UptUttEl I * III..
iiu..i.i U~I~I' III I
I U U
1 1 1 120 I
Plots of peak area vs.
mobile phase compositions.
head pressure for different
IIIIII lilllllIl IlllIlllIl ii
primarily composed of low velocity drops with
arge mean diameters which are
difficult to desolvate.
Significant transport losses result from surface impact, turbulence, and
evaporation when the aerosol droplets are either too large or too small
The plots also show that, as the percentage of water in the mobile phase
increases, a higher pressure (flow) of helium is required to produce optimum
related to the surface tension and viscosity of the
gas jet causes frictional
interactions with the liquid surface.
The main forces opposing aerosol generation
are the surface tension and the viscosity of the liquid phase.
much higher surface tension and is more viscou
Since water has a
than organic solvents, more
energy (hence higher gas flow) is required to efficiently nebulize aqueous solvents
(Browner et aL
It also appears that sensitivity decreases as the percentage of water in the
mobile phase is
Even at optimum conditions for each mobile phase
the peak areas were greatest when using a pure organic mobile
This observation can be explained by considering both the nebulization
percentage of water
n the mobile phase increased.
Since water has a much
aqueous droplets is more difficult because the aerosol has less surface area and
water also has a higher heat capacity and AH- than other common LC solvents
other studies have demonstrated that the observed order
of sensitivity for different mobile phases is inversely related to the energy to heat
and vaporize the solvent (Voyksner et al.
It does not appear for this PB interface that the flow rate of the mobile
phase affects the optimum nebulizing gas flow as much as the mobile phase
shows nebulizing gas optimization curves for different
flow rates of 100% acetonitrile.
All curves indicate optimum conditions at 30-35
the best sensitivity was observed at lower mobile phase flow rates.
It was also
flow rate was
The main function of the desolvation chamber is to remove
the volatile solvent molecules from the
Solvent evaporation is
achieved by the transfer of heat from the chamber walls to the
molecules which are present are typically less volatile and remain as particles.
$ \ I
Plots of peak area vs.
mobile phase flow rates.
pressure for different
Data points represent averages of triplicate
injections of 20 ng diuron.
The desolvation chamber in this interface was an 8" long,
It was heated
by two 300 W
nto the chamber block.
The temperature of the desolvation
L) feedback temperature controller which used a thermocouple to sense
the chamber temperature.
Figure 2-4 illustrates the
effects of desolvation chamber temperature and
mobile phase composition on the signal obtained for 20 ng of diuron.
points represent the average areas obtained from triplicate FIA injections.
plots reveal that the optimum temperature of the desolvation chamber depends
on the composition of the mobile phase. Thb
the percentage of water in the mobile phase
heat capacity and AH, than organic solvents,
e optimum temperature
Since water has a larger
more energy is con
Thus, higher temperatures are needed to replace the heat lost from
adiabatic expansion due to solvent evaporation to maintain efficient desolvation.
The plots also show that signal
decreases as the percentage of water in the
mobile phase increased.
As mentioned previously,
this trend is probably a result
of both the nebulization and desolvation processes.
desolvation chamber temperature was not as critical as optimizing the flow of
While the signal obtained at non-optimum desolvation chamber
I--r 75/25 CH3CN/HO0
I I I I I1 II
30 40 50
Plots illustrating the effect of desolvation chamber temperature on
peak areas obtained for FIA of 20 ng diuron at various mobile phase
las flow rates resulted
n as much as an 80% decrease in signal
However, this was not true for all the test compounds used.
2-5 shows desolvation temperature optimization curves for diuron and carbaryl.
Each curve was normalized with respect to its largest data point to better illustrate
the loss of signal at non-optimum conditions.
Although the peak areas for diuron
not vary by more than
peak areas for
optimum desolvation chamber temperatures decreased by as much as 80% with
respect to the maximum peak area.
Since carbaryl is more volatile than diuron,
at higher temperatures carbaryl was probably being vaporized in the desolvation
chamber and pumped away in the momentum separator region of the interface.
Although of a limited test set, Figure
chamber temperature is largely compound
also illustrates that optimum desolvation
independent as both curves exhibit
maximum signal at the same temperature.
Desolvation Chamber Pressure
The pressure in the desolvation chamber is largely determined by the flow
, the pressure in the desolvation chamber is typically 200
evaporation is the
rate of heat transport from the
surrounding gas to the
of the aerosol droo
WIiW *W I V* I I Y -
as the nebulizina
I I II I I I I I I I I 1 1 II
30 40 50
I I I I I III
Normalized desolvation chamber temperature optimization curves
for diuron (m/z 72) and carbaryl (m/z 144) using a 100% acetonitrile
solvent at a flow rate of 0.3 mL/min.
environment, supplemental He was added to the desolvation chamber to enhance
England) fine metering valve through a Swagelok (Jax Valve, Jacksonville,
installed on the desolvation chamber
. A pressure gauge (Omega, Stamford,
CT) was also added to monitor the pressure of the desolvation chamber (Figure
Figure 2-6 shows the effect of adding
to the desolvation
The mobile phase for th
studies was 100% acetonitrile; the data
points represent the average peak areas obtained from either 20 ng caffeine or
100 ng carbaryl.
Optimization of the nebulization process accomplished prior to
these studies resulted in a desolvation chamber pressure of
indicate that a two-to-threefold increase in signal was obtained by
pressure in the desolvation chamber by approximately 120 torr
plateau was observed for both compounds.
at which point a
These results are consistent with
those observed by other laboratories (Browner,
interesting to note that
nterfaces of similar design do not utilize the addition of
n the desolvation chamber.
After travelling through the momentum separator
, the analyte-containing
Plots illustrating affect of adding supplemental He to the desolvation
chamber for FIA of (a) 20 ng caffeine and (b) 100 ng carbaryl.
Amount He Added (torr)
enter the ion trap where they strike the hyperbolic surface of one of the
endcap electrodes which served as the PB target.
on with the endcap
causes the particles to undergo a rapid flash vaporization.
This results in the
production of predominantly
ntact gas-phase analyte molecules.
molecules are then ionized by E
, or alternately,
introduction of a
The temperature of the PB target (endcap) plays an
signal obtained for PB analyses.
The flash vaporization process must be efficient
to prevent peak tailing,
which requires sufficiently high temperatures.
thermal degradation of some compounds can occur if too high temperatures are
Typical target temperatures used for PB analyses are between 250 and
the temperature of target endcap was maintained by radiatively
heating the entire
SM vacuum chamber
, including the ion trap and electron
multiplier, with the standard quartz heaters mounted
In order to prevent damage to the electron multiple
inside the vacuum chamber.
ier, temperatures were not
raised above 140C.
n bad sensitivity and a large degree of peak
tailing even for FIA because these
ow temperatures yielded an
n order to permit the use of typical PB target temperatures,
were made to the filament endcap electrode which served as the target.
large holes were drilled into the body of the endcap to insert two 10W cartridge
to sense the endcap temperature.
The temperature of the endcap was controlled
by an external feedback Omega (model CN-2012,
, CT) temperature
These modifications allowed the endcap to be heated to typical PB
operating temperatures of up to 290C while maintaining the electron multiplier
mounted within the vacuum chamber at normal operating temperatures of 1000C.
improvements these modifications brought
about are illustrated
are mass chromatograms
from triplicate FIA
injection of 200 ng carbaryl at endcap temperatures of 125 and 2500C.
the peaks exhibited a significant degree of peak tailing caused by
inefficient flash vaporization of the particle beam.
At higher temperatures,
shapes were dramatically
The signal intensities
improved and little evidence of peak tailing was
hown in the figure also show that there was an
increase in peak height at the higher temperature.
optimum temperature was largely compound-dependent.
Optimization curves for
the FIA of 100 ng carbaryl and 75 ng rotenone are shown
n Figure 2-8.
are normalized with respect to the maximum peak area in each data set for clarity
rotenone exhibits a maximum
at much higher target temperatures of
Rotenone is much less volatile than carbaryl,
so it is not surprising that
Il Ill' II
II II II I
I 111111 II
Target temperature optimization curves for carbaryl (m/z
indicate that the optimum target temperature is largely compound-
The carbaryl optimization curve illustrates that higher temperatures are not
optimum for all compounds.
This is because thermal decomposition of some
of the 1 44 fragment ion (base peak) occurred at 200 C and decreased
by approximately 50% at target temperatures of 290C (Figure
was observed that the total ion current signal
increased by approximately 20%
when the target temperature was increased from 200 C to 290 OC.
degree of thermal
limited the sensitivity of the m/z
fragment ion at high target temperatures.
studies demonstrate the need for
nebulizing gas flow rate, desolvation chamber temperature,
and target temperature have al
been shown to affect the
the LC/PB/ITMSTM system.
Most of th
interface parameters were found to be
largely mobile phase dependent; however,
some compound dependance was
, most notably for the source target temperature.
make it difficult to optimize interface parameters for gradient
elution analyses because the mobile phase composition will change during the
Modifications to the ion trap endcap electrode were necessary to allow
EVALUATION OF STRATEGIES TO MINIMIZE ADVERSE
EFFECTS OF RESIDUAL SOLVENT
Although ion traps are noted for their sensitivity
quadrupole mass analyzers.
Ion traps have seen limited use
because of the large amount of solvent introduced
n LC/MS systems
by most LC/MS
Excess solvent ions and neutrals typically cause space charging and
poor mass resolution and poor
overall spectral quality.
n this chapter are reported several strategies which were evaluated,
at minimizing adverse effects caused by introduction of residual solvent from a PB
included one method which reduced the
solvent which reached the trap.
The versatility of the
ST also allowed the
creation of customized scan functions which ejected unwanted solvent ion
within the ion trap prior to mass analysis, while efficiently storing analyte ions of
Although the coupling of PB with an ion trap using both direct coupling
operational modes of the ion trap which minimize space charging by ejecting
residual LC solvent ions prior to mass analysis.
determine the best way to operate the
The goal of this work was to
STM to produce quality El spectra from
introduced via LC.
El spectra obtained from isocratic LC/PB/ITMSTM
analyses of simple pesticide mixtures (
_3 components) are compared to library
El spectra and
El spectra obtained from solids probe/ITMSTM
"solvent-free" method) of pure compounds.
instrument calibration curves
and limits of detection (LOD) obtained from these isocratic LC/MS analyses on
this system are reported.
Solvent ejection studies utilized an ISCO (Lincoln,
Chromatographic separations were performed using a Hewlett-
Separations were performed on a 100
x 4.6 mm Hewlett-Packard octadecasily
, packed with 5 pm particles.
Separations were accomplished isocratically
with a mobile-phase composition of 80/20 acetonitrile/reagent water at a flow rate
of 0.3 mL/min.
The analytical column was thoroughly conditioned by pumping
a 50/50 acetonitrile/reagent water mixture through the column overnight to remove
residual impurities and column bleed.
The LC/MS interface was a prototype Finnigan MAT
of the interface are described
important PB operating parameters including desolvation chamber
temperature and nebulizing
He flow rate were optimized to provide maximum
The desolvation chamber pressure was optimized by addition of He
positioned about /4" away from the
on trap entrance to prevent high pressures
inside the trap.
. The mass spectrometer used
was a Finnigan MAT (San Jose, CA) ITMS
Fundamentals of ion trap operation,
and ion motion are described in greater detail in Chapter 1
vacuum chamber was maintained at 100C for all experiments.
As described in
, the filament endcap of the ion trap was modified to allow insertion of
10W heater cartridges which were controlled by
an external temperature
controller (Omega Engineering
Model CN 2012,
served as the particle beam target and was operated at temperatures of 250-
ionization was employed within the ion trap;
x 10 torr (uncorrected) as measured by a Bayard-Alpert ionization
gauge (Granville-Phillips, Boulder, CO) mounted on the vacuum chamber. Helium
was added into the vacuum chamber to produce typical operating pressures of
x 10" torr (uncorrected).
TMST was repeatedly scanned from 55 to 500
amu to check for formation of dimers and ion/molecule reaction products
(Weber-Grabau et al.,
at 530 kHz and 6V
during the analytical scan in al
studies to reduce the effects of space charge and
nuron were obtained from AccuStandard (New Haven, CT),
caffeine from Sigma (St. Louis,
other pesticide standards were obtained
from the U
Environmental Protection Agency (Pesticide Chemical Repository,
Research Triangle Park, NC)
All standards were used without further purification.
The structures of the standards used in these studies are shown
in Figure 3-1.
Also shown are molecular weights and the El fragmentation which yields the
peak of each compound's
, molecular weights and El
fragmentation for carbaryl,
rotenone, and diuron were shown previously in Figure
HPLC grade methanol and acetonitrile were obtained from Fisher
reagent water was obtained from
a Milli-Q water purification
Problems Caused by Residual Solvent
The major challenge encountered when interfacing LC directly with the ion
trap was minimizing adverse effects caused by the introduction of residual solvent
from the interface.
Solvent molecules (S) in the trap are ionized by El:
Space charge results when the density of ion
n the trap becomes
large enough that ion-ion interactions become significant.
This can distort the
correct mass assignments.
In the ion trap,
ions and neutrals are present within a confined space for
a period of time prior to ejection/detection.
As a result,
a large population of
and molecules can also cause undesired ion/molecule reactions to
a large degree of solvent-C
of analyte neutral
as illustrated below:
(S + H)+
(S + H)+
(M + H)+
These equations show that a solvent radical ion
(S+) can react with a neutral
a a a. S a ..1 -1 .t-a
^ -* -1
1 rl ~ 1 .
proton transfer will occur and a protonated molecule, (M+H)+
undergo little fragmentation as compared to the M'*"
ions initially formed by El.
Since the primary advantage of using PB is the ability to acquire El mass spectra,
these adverse effects of residual solvent must be minimized.
These adverse effects are illustrated in the profile mass spectrum shown
n Figure 3-2.
The mass spectrum was acquired from FIA of
-100 ng of carbary
n 100% methanol mobile phase.
The spectrum shown is an expansion of the m/z
25-150 mass range to better illustrate the resolution between adjacent masses;
the abundance of the M'
abundance of methanol
' ion at m/z 201 was
solvent ions centered
<5% relative to
m/z 33 caus
charging which resulted in poor mass resolution between adjacent low masses.
Notice that resolution
improved at higher masses.
Figure 3-2 is
provided to better illustrate the mass resolution between
ejecting trapped ion
in solvent ions being
from low to high mass (Stafford et al.
before higher mass ions during
reducing space charging when higher mass
ratio in this spectrum
a large degree of
The base peaks of the El and Cl mass spectra of carbaryl
and 145* (M+H-57)*
Although some 145" from
carbaryl (11 % relative to 144"),
the figure shows the intensity of 145+ being about
three times greater than
that of 144k
The abundant methanol
a large amount
the use of
residual solvent ions to perform solvent-CI analyses of pesticides with the same
Sm system has been demonstrated (Eades et aL,
here the large degree of solvent-CI was undesirable since the goal of this work
was to produce quality El spectra.
prior to mass analysis to minimize
This figure illustrates the
need to eiect solvent
space charging and solvent-C
good quality El m
to Minimize Adverse Solvent Effects
adverse effects of residual
solvent are illustrated
diagram of the
in Figure 3-3.
(S and A
The block diagram shows that solvent and
, respectively) elute from the
HPLC and pass through
into the ion trap where solvent and analyte ions (S+
and A+) are
formed by E
The number of solvent molecules which reach the trap could be
before the trap.
the trap could be operated such that solvent ions are ejected from
within the trap prior to m
By employing these strategies,
analyte ions of interest eventually reach the detector
E N .0-,
As mentioned previously, the
nterface used in these studies employs an
example of one strategy
at reducing the
number of solvent molecules which reach the trap.
The amount of solvent which
reaches the trap could also be reduced by employing a membrane drier
of the desolvation chamber.
In the membrane
volatile solvent diffu
transferred to the momentum separator.
The commercially available Universal
nterface from Vestec is based on this operating principle (Vestal et al.,
injection of ions created
n an external ion source.
been demonstrated using both on-axis (Louri
,1989) and off-axi
, 1989) ion sources.
from within the trap.
here at UF
for mass-selective ionization in the quadrupole ion trap to
eliminate undesired ion
of a range of mass/charge (m/z) values
(Eades et al.,
included the use of resonant
comprised of multiple frequencies.
, implementation of th
required modifications to the
ion trap operation
STM source code.
implemented with standard
In this chapter, two modes of
TMST software are evaluated for
analysis: 1) elevation of rf voltages to eject ions below a chosen m/z ratio (i.e.
impose a low mass cutoff) and 2) application of a supplemental rf signal to eject
a single solvent ion species.
Both methods were evaluated when applied either
during ionization or during an ion storage period following ionization.
of strategies which may minimize adverse solvent effects is shown below in Table
Summary of Strategies to Minimize Adverse Solvent Effects.
Remove Solvent Molecules Before the
. Membrane Drier prior to Momentum Separator
. Additional Third Stage of Momentum Separation in
External Source and Ion
Eject Solvent Ions within the Ion
Elevate rf Voltage (Impose Low Mass Cutoff)
b. During Ion Storage
Resonant Ejection of Single I
a. During Ionization Step
b. During Ion Storage
Species (Notch Filter)
Additional Staae of Momentum Separation
The same PB interface was used to determine the benefits of using the
additional third stage of momentum separation to reduce the amount of solvent
which reached the trap.
The interface was converted to a two-stage interface by
removing the third skimmer and third stage roughing pump from the interface.
Pressures in the momentum separator region were approximately 5-10 torr and
400-500 mtorr is stage one and stage two,
pressures are not
Figure 3-5 illustrates that the additional stage of pumping does reduce the
e (m/z 42) at different flow rates of 100% acetonitrile mobile
increased flow rate and that about four times
less solvent ions were formed when
three stages of pumping.
ists the ion
gauge pressures for
solvent compositions for both configurations.
obtained at a flow rate of 0.3 mL/min.
and with no
supplemental He added to the
STM vacuum chamber, or third stage of the momentum
The table indicates that the partial
0.1 0.3 0.5
Flow Rate (mL/min)
I 3-Stage 2-Stage
compositions using two and three stages
of momentum separation
Ion Gauge Pressure (torr)
1.9 X1 0i
5.8 X 10
1.9 X 104
5.7 X 106
2.8 X 104
5.7 X 104
3.2 X 104
3.5 X 10-5s
All pressures obtained at solvent flow rate of 0.3 mL/min. and no
He added to desolvation chamber
. Pressures were read directly from the ion
gauge meter with no correction factor used.
The performance of both configurations was evaluated by comparing the
El spectra obtained for FIA of 20 ng caffeine in 100% acetonitrile mobile phase.
The spectra are presented
n Figure 3-6.
spectrum obtained from a computer
shown is a library reference El
included with the
comparison with the
library spectrum with respect to both fragmentation pattern
ntensities of the m
The spectrum obtained using two
however, does not compare favorably with the library spectrum.
. .. N
-8 S C a 8
- -. .Y -LL~ .;- -r 1 r -. a a a -l a ar a~ a* *l aC a a a sa ;- ~ au C fl S
0 0) LI.
*\T I 1
SI I I I I
I 1 1 1 1
activated dissociation (CAD) to occur from collisions with background solvent
, the most abundant molecular ion peak was the (M+H)+
at m/z 195 which
arge degree of solvent-C
significant amounts of solvent and improved the performance of the LC/PB/ITMST
Elevation of rf Level
of solvent ion ejection studied
involved elevation of the rf
voltage applied to ring electrode to eject ions below a chosen m/
a low mass cutoff.
In this mode, the ion trap was operated
proportional to the rf voltage (Equation 1-5),
elevation of the rf voltage increased
the q, values
of trapped ions.
In the rf-only mode, ions with values of q, greater
than 0.908 wi
follow unstable trajectories
and be ejected from the trap
Since q, is inversely proportional to m/z
n the axial
low molecular weight LC solvents (e.g.
H20) were ejected at a
low rf potential while analyte ions of higher m/z
In this mode, the ion trap was basically operating as a high-pass mass
The effect of elevating the rf voltage upon both solvent and analyte ions is
cDCS 0 I
*n~ y) J
- tf **o -5
Ew D t
a $ iO
^.^ w m~
1(8. O S *-
E w o
.. o c ca
C O0 0)L.
o- a), 4
* ~o Ci
-._ S' wE'
+*g 0- T0
^ mo -s
u *B > *B
CO O O>4a
* U)- 0
0) 0)1 aStfl
4-' 0 -
+* O *
6D 1jOA Op
42) and the most abundant fragment ion of diuron (m/z
here that the stability diagrams are plotted in terms of actual rf and dc voltage
applied to the electrodes, rather than a,q space.
rf-only mode, the rf voltage determines the
Since the trap is operated
ow mass cutoff of the trap,
At an rf level corresponding to point A in the figure, both the solvent
and analyte ion will be trapped because this point is located within the stability
diagram of both ions.
Point B is located outside the solvent
but is still within the stability diagram of the analyte.
on stability diagram
At this rf level
be ejected while the analyte ion will remain effectively trapped.
This mode of operation was investigated when applied both during the
ionization event and afterwards
, during ion storage.
The effect of raising the rf
level (low mass cutoff) upon the ion intensity of some commonly used LC solvents
n the plots of Figure 3-8.
as the instrument's
efficiently (100%) ejected from the ion trap.
These plots were created by raising
the rf level after ionization
, during ion storage.
Although not shown,
were obtained by elevating the rf during ionization.
Ilustrates how the
intensity of the carbaryl analyte ion (144+)
was affected by elevating the rf
evel (q,) both during
onization and during ion
These plots were obtained by varying either the ionization or storage
values of q, with a constant amount of carbary
(2 ng/p4L yielding 10 ng/s
0 10 20 30 40 50
Plots of normalized solvent
evel (expressed as
low mass cutoff) during ion storage illustrating efficient ejection of
common LC solvent
I I. I I I I I
III III IIII
I lJ III II
IaI III III
\ During Ion
I I I I I I I I I
I I I I I I I I 1 I
I I I I I I I I I 50
as low mass cutoff) both during ionization and during
, signal decreased significantly with increasing ionization q,.
so does the
resulting in large initial velocities and ion losses due to quasi-unstable trajectories
These losses occur when the magnitude of an ion's
conditions for stability still exist.
Although raising an ion's
q, value during ion
storage also increases its kinetic energy,
the ions are already effectively trapped
onization value of q,
of an ion also decreases the
volume which limits the region
n which ions can be created and remain stable
The observed decreases
n intensity could also be the result of
decreased ionization cross-sections due to high electron energies at elevated rf
Preliminary studies investigating the effects of ionization rf level (q2) of n-
butylbenzene ions also showed decreased ion
(Pedder et al.
ntensities at higher values of q,
There were also some differences
n the extent
spectra obtained by ejecting solvent during
ionization and during ion storage.
shows how the ratio of 144+ (El fragment) to 145
'"C isotope of 144+)
is affected by elevating the rf
storage for carbaryl
n 100% methanol.
The rf level where the methanol solvent
Elevate rf during
Elevate rf during
III IIII I III IIIII II
SI I II II I I I I I I I I I I I I I I I I I I
) 40 50 60
Plots of ratio of 144'
(El fragment) to 145+
cutoff) both during
ion storage for carbaryl in 100% methanol.
fragment) vs. rf level
onization and during
level where methanol solvent ions are efficiently ejected from the
in the figure.
In a "pure" El spectrum,
intensity ratio would be
expected to be 9.0,
based on the natural abundance of 'C isotopes of 144'
low ratios obtained at low rf levels resulted from solvent ions causing a
degree of solvent-CI.
of solvent ions
resulting in increased 144C/145* ratios; the inability to achieve the expected ratio
for a pure El spectrum indicated that some solvent-CI still
occurred in the time
before solvent ions were ejected.
lower ratios were observed when the rf
was elevated during ion storage.
the ionization time,
This is because solvent ions were stored during
which allowed more time for solvent-CI to occur before the
solvent ions were ejected.
Resonant Election of Sinale Ion Species
The other method of solvent ion ejection studied involved resonant ejection
of a single m/z
solvent ion species by applying a notch filter (Kelley et al.
This was accompli
excitation voltage) across the endcap electrodes to kinetically excite a particular
solvent ion m/z species
such that it was ejected from the trap.
of trapped ions of a particular m/z are generally characterized by their unique
secular frequencies, wz, which are a complex function of the Mathieu parameters
az and q,, and 8z,
which relates to the frequency of the oscillation of the ions.
Application of the resonant excitation voltage at the wz of a particular solvent ion
The excited ion can then undergo CAD with residual buffer gas
and of relevance to this discussion,
the excited ion
species can also be ejected from the trap if sufficient power is absorbed such that
the amplitude of the oscillations become larger than the trap's
use of resonant ejection was investigated because it permitted ionization to be
compounds whose El mass spectra have low m/z base peaks.
As with elevating the rf amplitude, the use of a notch filter was
or ion storage.
resonant ejection of the m/z 42 ion of acetonitrile solvent both during ionization
, during ion storage.
The figure shows that m/z 42 was efficiently
100%) ejected when resonant ejection was applied during ion storage.
ions of different m/z p
signal were observed.
'ossess different secular frequencies,
no losses in analyte
This process was not nearly as efficient, however
In this case only about one-third
solvent ions were ejected.
This is because the ionization and ejection of ions
Ions were constantly being
during the ionization event; those created towards the end of the ionization period
were only resonated for a very short period of time,
and thus were not efficiently
represents the maximum amount of time an ion was resonated during ionization.
35 40 45 50 55
Resonant Excitation Voltage
Resonant Excitation Voltage
-M -h. sAJL v-tt
* I V I j
I TTTI I II I"
a) Profile mass spectra
(M+H) + solvent ions (n
illustrating resonant ejection of acetonitrile
i/z 42) during ion storage.
Resonant Excitation Voltage
35 40 45 50 55 60 65 70 75 8B
Resonant Excitation Voltage
A -A. 4'
35 40 45 50 55 60 65 70 75 80
(continued) b) Profi
acetonitrile (M+ H)+
ustrating resonant ejection of
42) during ionization.
ComParison of Solvent Ion Election Methods
The analyte signal obtained for methods which efficiently eject solvent ions
are compared in Figure 3-12.
These plots were obtained by varying the ionization
time with a constant amount of diuron (1 ng/pL yielding 5 ng/s at 0.3 mL/min.) in
100% methanol flowing through the
base peak of the El spectrum of diuron occurs at m/z 72; its
intensity is plotted
in Figure 3-1
evels were elevated such that al
below m/z 45 were
ejected for methods involving elevated rf leve
For the notch filter mode, the
resonant excitation signal was set to eject the (M+H) ion of methanol solvent
(m/z 33) for 10 ms during ion storage. As expected,
increasing the ionization time
inear increases in signal for al
space charge develops
due to too many analyte ions being trapped, at which point the plots begin to
level off. The plots also reveal that methods which involved ejection of solvent
ions during ion storage produce about two times greater signal than ejection
As discussed previously
we believe this to be because of ion
osses due to quasi-unstable trajectories
and smaller ionization volumes
values of qz during
be noted that these
effects might not be as significant for compounds whose El mass spectral base
peaks are at higher m/z values.
carbaryl show less than two times difference
n signal between these operational
was not further
mass spectra of the
I I i I I I I I I
I I I I I I
the text which
I I I I II II I I I I I I I I I I I I I I I
To obtain maximum sensitivity,
all subsequent studies ejected solvent ions
by elevating the rf level after ionization, during ion storage. Although resonant
ejection during ion storage yielded essentially the same sensitivity, elevation of the
rf amplitude ejected all solvent ion species,
as well as background
, column bleed, etc.),
below the chosen m/z.
software, ejection of more than one m/z solvent ion species by resonant excitation
would require multiple resonant ejection events.
Although not all solvent ion
10 ms for efficient ejection,
multiple resonant ejection events
would allow more time for undesired ion-molecule reactions to occur.
resonant ejection events would be necessary for LC/PB/ITMSTM analyses which
would employ gradient elution or isocratic solvent mixtures.
ADDlication to LC/PB/ITMSTM
Analyses of Simple Pesticide Mixtures
The capabilities of the LC/PB/ITMSTM
initially evaluated by
mass spectral quality,
Ionization was performed at an rf level corresponding to a low mass
ms; solvent ions
elevating the rf level to eject all ions below m/z 55.
Total ion chromatogram (TIC), mass chromatogram
mixture using LC conditions described in the text.
m/z 88 Methomyl
1B8 288 388
figure displays the total ion chromatogram
mass chromatograms of the
base peaks of the El mass spectra of each component,
and amounts of each
Structures and E
fragmentation of each component were
shown previously in Figure 2-1
than baseline separation of the compounds was achieved
ess than 6 mm.
The background-subtracted mass spectrum of 80 ng of diuron (m.w. =232
n Figure 3-14.
The spectra are quite similar with respect
to both fragmentation patterns and
relative fragment ion
abundance of the (M+1)+ ion
n the LC/PB/ITMSTM spectra indicated that only
amount of Cl occurred
, and did not significantly degrade the overall
The spectra of the other compounds
n the test mixture also
compared favorably with reference and solids probe/ITM
instrument calibration curve for diuron
STM El spectra.
is shown in Figure 3-15.
curves were constructed
by plotting the
as a function
quantitation ion for each compound was the base peak of its fu
-scan El mass
It appears from the figure that a linear calibration was obtained from
The inset of the figure is an expansion of the region from
ustrates the linearity at the low end.
The value of the correlation
spectra of diuron obtained from (a) library (Hites, 1985), (b)
robe/ITMSTM analysis, and (c) LC/PB/ITMS analysis of 80
ng diuron (background-subtracted).
60 80 100 120 140 160 180 200 220 240
- n il-
159 187 215
60 88 180 120 140 160 180 200 220 240
-^- *^k e n- rr'ia
80 100 120 140 160 180 200 220 240
SI 1 1 1 1 1 1 i I I I II i i I | | i i i I I
3 0 0 0
3 0 0 0
S0 0 0
'5y) +DZ aJV
0 0 0
The non-linearities at the ends of the calibration curve are presumably
due to particle beam transport problems at the low end (Bellar et al.
the onset of space charging caused by too many analyte ions at the high end.
Although diuron here shows linear behavior, nonlinear calibration curves, like that
obtained for linuron in Figure 3-16,
are far more prevalent
nonlinear behavior of PB has been attributed to transport losses through the
momentum separator region of the
This has been observed by several
other laboratories as a limitation of PB/LC/MS
for quantitative analyses (Doerge
The use of isotopically labelled
ntemal standards has been suggested as a possible solution to this problem and
has been the topic of several recent reports (Ho et al.,
1992, Doerge et al.
Quantitation using PB and the carrier effect will be discussed
n greater detai
concentration which reliably yielded a 3:1 signal/noise ratio for each compound's
estimated detection limits routinely obtained on this system for several carbamate
and urea-based pesticides.
are typically in the low ng range and are
comparable to LOD's
reported for PB systems which employ quadrupole mass
analyzers (Behymer et al.
The LOD reported here for
inuron is 10 times
lower than previously reported (Behymer et aL,
1990) because the base peak of
I II IIII
I I I I I II ll
instrument calibration curve from PB/LC/ITMS
ne represents region where the onset of space charging
mits for several carbamate and
n the experimental
report to eliminate interference from using
ammonium acetate in the mobile
(M+', m/z 248)
These LODs were obtained from the calibration runs and are not intended
Note that there are a number of options offered by the
STM system that could further
, higher multiplier voltages,
of a high
, including longer
dynode, and addition of mobile phase additives such as ammonium acetate to
ionization time using automatic gain control,
AGC (Stafford et al.
might lower detection limits and extend the linear dynamic range as
n this chapter
When directly coupling
nterface with an
, it was necessary to eject solvent ion
to minimize space charging and
prevent a significant degree of solvent-C
The additional third
stage of momentum separation has been demonstrated to reduce the amount of
solvent which reached the trap.
Ejection of solvent ion
prior to m
spectra and El spectra obtained by solids probe/ITM
STM analyses. Although linear
prevalent for LC/PB/ITMSTM determinations of several pesticides.
for several pesticides have been estimated to be
n the low ng range.
APPLICATION OF LC/PB/ITMS TO ANALYSIS OF PESTICIDE MIXTURES
AND COMPARISON TO OTHER LC/PB/MS
The analytical utility
of PB relies on the ability to generate classical El
compounds introduced via LC.
The vast majority of PB/LC/MS system
n use employ quadrupole mass analyzers due to their widespread availability, low
cost, and tolerance of relatively high operating pressures.
Ion traps have seen
n LC/MS systems primarily because of the large amount of solvent
introduced by most LC/MS
n the previous chapter
were investigated which minimized adverse solvent effects and allowed acquisition
favorably to reference library spectra.
Although isocratic separations are well suited for individual applications
which require simple separation strategies,
they are limited for the analysis of
Isocratic separations of multicomponent mixtures typically exhibit poor resolution
of early eluting components,
bad peak shape of late eluting components,
require the use of gradient elution,
temperature programming for GC.
n gradient elution,
the composition of the
mobile phase is changed throughout the analysis to
increase its eluent strength.
An optimized gradient affords the ability to efficiently analyze a
arge number of
components in a minimum amount of time.
This chapter will report the application of the LC/PB/ITMST system for the
analysis of a ten-component pesticide mixture.
The pesticide mixture required the
use of gradient elution to achieve adequate separation of the components.
performance of this system for gradient elution analyses wil
be compared to that
regards to detection
Finally, the capability of the ion trap to acquire both El and Cl spectra in
alternate scans over the course of an LC peak will be demonstrated.
Chromatographic separations were performed using
a 5 upL
Separations were performed on a Waters (Milford,
a I -
MA) Nova-Pak 150 mm
-I a a' .,,,,,, ....... ..:L Xt.. S-1 -1- -1- ..... ...- S
* J 11 ii __K
through the column overnight to remove residual impurities and column bleed.
The mobile phase used for these studies was an acetonitrile/0.01M ammonium
reagent water mixture at a flow rate of 0.3
LC-2600 syringe pump was used for post-
column addition of 100% acetonitrile at a flow rate of 0.1
resulting in a
total mobile phase flow rate of 0.4 mL/min.
through the interface.
The LC/MS interface was a prototype Finnigan MAT (San Jose, CA) three-
stage particle beam interface (Figure 1-4).
nterface is described
chamber temperature, pressure, and nebulizing He flow rate were optimized by
making several injections of the test mixture under different conditions to provide
compounds in thl
composition of the mobile phase changes throughout the course of the analysis.
Chapter 2 revealed that most PB parameters are largely mobile phase dependant.
in the three stages of the momentum separator were 5-10 torr,
300 mtorr, and 100 mtorr (adjusted by addition of He),
was inserted into the mass spectrometers via a /" o.d. transfer line probe through