GC/MS/MS on the quadrupole ion mass spectrometer software development and the examination of the effects of ion population

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Title:
GC/MS/MS on the quadrupole ion mass spectrometer software development and the examination of the effects of ion population
Physical Description:
xvii, 230 leaves : ill. ; 29 cm.
Language:
English
Creator:
Griffin, Timothy P., 1963-
Publication Date:

Subjects

Subjects / Keywords:
Quadrupoles   ( lcsh )
Mass spectrometry   ( lcsh )
Trapped ions   ( lcsh )
Genre:
bibliography   ( marcgt )
theses   ( marcgt )
non-fiction   ( marcgt )

Notes

Thesis:
Thesis (Ph. D.)--University of Florida, 1995.
Bibliography:
Includes bibliographical references (leaves 225-228).
Statement of Responsibility:
by Timothy P. Griffin.
General Note:
Typescript.
General Note:
Vita.

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Source Institution:
University of Florida
Rights Management:
All applicable rights reserved by the source institution and holding location.
Resource Identifier:
aleph - 002058434
notis - AKP6484
oclc - 34424534
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AA00003193:00001

Full Text










GC/MS/MS ON THE QUADRUPLE ION TRAP MASS SPECTROMETER
SOFTWARE DEVELOPMENT AND
THE EXAMINATION OF THE EFFECTS OF ION POPULATION















By


TIMOTHY P


. GRIFFIN


A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
nr 'r~rin r~ vM-n~ ru 1 C'r-MnI i\/












ACKNOWLEDGMENTS


There have been many peop


n my


ife who have made this possible.


First,


would like to thank my family


, in particular my mom and dad,


for their


encouragement and support throughout the time that


have spent here.


people would listen to us,


world would be a perfect place,


a whole lot


cleaner. A special thank you goes to Sara Griffin.


We have had our problems


and we are


ust now getting on with our lives, but


couldn't have gone thi


without her belief in me.


also would like to thank the many people that


years that have helped me


have met over the


to grow and develop into the great and humble


person that


am today.


n particular


would


ike to thank Linda Doubenmeir


for her support during the times


at Floyd Brown and Associates.


surprising to me to this day that we made it through without


of our minds.


right out of school.


osing too much


Thanks also goes to Floyd Brown for believing in a young punk


offered me a chance to turn into a man and


world from a different point of view.


would especially like to thank Rick Yost.


He gave me the freedom and


support not only to develop into a true scientist but also a more self-confident







over the


years,


I lift my 32oz Coke to them all.


Imagine


grad school without


caffeine

I


and sugar--I don't think it could be done.

am very indebted to Nate Yates who not only got me started on the


software but also has been a true


and caring friend during the these last four


years


The canoe trips were a blast.


All of the canoeing,


swimming,


camping


and of course drinking made for a great stress relief.


n Gainesville would


be hard to


Stephenson's


imagine without the


house.


BBQs and dinners at Jim (Nerd Boy) and Tracy


What a great way to relieve stress and the frustrations of


scrapings and leavings.


will never forget the afternoons


hitting and watching the sights walk by


with Don Eades and Brad Coopersmith


they were truly some


awesome sights.


also would


ke to thank Shannan (I spelled it correctly) Carlson for her


friendship over the past months, even if


he didn't acknowledge me on her


poster.


We have had a lot of fun


, and spent a lot of money


'll never forget


the Abby.


It was great meeting another misunderstood person


n the world.


would also like to acknowledge Matt Booth for all the discussion


along the way.


and help


also have enjoyed seeing how much Coke he could drink


a day.


A very special and heartfelt thank you goes to Emily Carter


. During the


time that


have known her


have truly felt alive.


These last months would


have been unbearable had it not been for her caring and understanding.


From
















TABLE OF CONTENTS


ACKNOWLEDGMENTS


i*
. . . .II


LIST OF TABLES


S S S S S S Uvii


LIST OF FIGURES ... .. .. ..


ABSTRACT


CHAPTERS


INTRODUCTION
Background
Specific Aims


S C S * S
S U S S S S U S


Function/Description of the
Theory of Operation .


* Trap .

Trap


* .1 S *


S S U S S S 55


Fundamental Parameters
Stability Diagram
Tandem Mass Spectrometry .
RF/DC Isolation . . .


* S S S S
* C S C S


Overview of Dissertation

SOFTWARE DESCRIPTION


* S S S S C S S S S S S S S C S

* 5 5 S S I S S S S C S S U S S


* S S S S C S S S U S S

* S S S C S S C C I S 5 0


introduction


Overview of Software


Down Menu


Spectrum Display


Parameter Button


Scan Editor


S . .


.. .. .. .. 38


Clipboard Editor
SAP Table Editor


Scan


* S C S S S S P
* I S A S S S C S


Table Editor


Custom Calibration


RF Calibration


n~








Auxi


ary Board Description


Auxiliary Board Editor
GC Acquisition
Procedure Language ..
Procedure Words


Example Procedure Program
Summary . . . . .


s . . . . . .


RAMPED DC VOLTAGE.
Experimental .. ..
Instrumentation


Automated Acquisition Procedure
Results and Discussion


Stability Boundary
Figures of Merit for I
Isolation Efficiencies


olation
* a a .


. a a a a a a a a S S .
* a a a a a a a a a a a a a a .


Summary


4 MASS-SELECTIVE AUTOMATIC GAIN CONTROL


introduction


Set Ionization


Time


* a S S a
* a a a a


Variable Ionization


Time (AGC)


Mass-selective Automatic Gain Control


Experimental
Instrumentation
Timing Diagram
Compounds
Results and Discussion


* a a a a a 4 a a a a a a a . a a a a a a a a
* a a a a a a. a a S . a a a .a a a a a a a a .


Summary


STABILITY DIAGRAM BOUNDARIES


Introduction
Background
Objectives..
Experimental


* a a a a a a a a a a a a a a a a a a a S S S 5 a a a a S S
*. a.. a. a a. a a. S. a. a. a. a. a. a a. a .a .a a. a . .S a
*. a . .a. .a. .a. .a. .a. .a. .a. .a a . .S. .S S .* S


Instrumentation
Scan Function


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 a a a a a a a a a a a a a a


Mapping of Entire Stability Diagram
Detailed Examination of Boundaries


Isolation Efficiencies
Results and Discussion .
A in..n...s-. rn44rn O4..k


* a a a a a S S S a
a a a a a a S S S S S S *


tI, ',n r#r u







EFFECTS OF


ON POPULATION ON


SECULAR FREQUENCY


Introduction


Background
Objectives......
Experimental
Scan Function


Acqu


ition of Frequency Profile


Ion Populations . .
Compounds
Results and Discussion


Frequency Comparison


Changes in az


Changes
Summary


n qz


S. 4 4 S S S S *


Values
Values


7 CONCLUSIONS AND FUTURE WORK


Conclusions


220


Gatorware


Fundamental Studies


Future Work


Gatorware


Fundamental Studies


REFERENCE LIST


BIOGRAPHICAL SKETCH














LIST OF TABLES


Table


Auxiliary board memory map


List of control words (Assembly Language)


. . . . 66


Scan

Scan


Table flags for procedure

Table values for procedure


. . . 79


. . 80


SAP Table values for procedure


Experimental setup of previous studies.


S SS *a *aa aS 1d13


Experimental overview


. . 146


Experimental parameters to obtain a frequency profile


pace
e













LIST OF FIGURES


Figure


pag


Cross sectional view of the quadrupole ion trap mass


spectrometer. The
endcap electrodes.


on trap


is composed of one ring and two


The RF and DC voltages are applied to the


ring electrode while the auxiliary frequency is applied to the


endcap electrodes.


The filament creates electrons to form


within the


on trap wh


e the detector is used to detect ion


acquire a mass spectrum.


Block diagram depicting the communication between the


TS-40


and personal computer.


The communication


can be seen as


being composed of two parts.


The first part is the acquisition


and storage of the mass spectrum by the scan and acquisition
processor (SAP) board while the second is the display and


processing of the data by the personal computer (PC).


Very


limited real-time control of the ion trap


is allowed the user.


a a 9


Typical scan function for a QITMS.


The mass spectrum is


acquired as the RF trapping voltage is ramped to larger values


and the ion


become unstable and are ejected from the QITMS.


Also note that the ions are ejected


n order of


increasing m/z.


. 1


Graphical representation of the stability region of the QITM


The overlap of the


z and r stable regions (cross hatched)


depicts the region of stability.
different m/z.


The solid dots denote ion


Scan functions depicting the tandem mass spectrometry on a


QITMS.


Notice that the scan function uses two-step


isolation to


store a single m/z.


n addition, the MS/MS stage is performed


by collision-induced dissociation (CID).







Graphical representation of two-step isolation.


ejection of the


Notice that the


ow m/z and high m/z are performed at different


position


in the stability diagram.


The m/z of interest also must


be placed in close proximity to the stability boundaries.


Main screen of the
page is the central


TS-40 software and Gatorware.


The exec


location to reach all sub routines of the


software.


To reach the


page followed by IT


S-SC


TS-Scan
;AN. .


is typed from this
a* . a p p a a a U


The main screen of Gatorware called


TS-Scan.


This is the


primary screen for controlling the QITMS.


includes a real-time


display of the mass spectrum and nine of the most commonly
adjusted parameters . . . . . . .


. 32


The option


pull down menu.


The menu allows the user to


control


the automatic calculation and downloading of the scan


function to the SAP


n addition


, the display and real-time


acquisition can be set up through this menu.


Menu control to download the firmware to the SAP


. The menu


also enables the user to send the scan function to the QITMS to


control the operation of the trap.


The scan table clipboard allows the user to control the order of


the tables


in the scan function.


Tables


also can be added and


deleted from the scan function.


The SAP table editor allows the user to directly control the lines


that the microprocessor uses to operate.


most flexibility; however,
electronics is necessary.


This editor allows the


thorough knowledge of the QITMS


The scan table editor is a simplified editor.


The editor requires


the user to know the fundamental parameters of the QITMS


however


, no knowledge of the electronic is necessary.


The tools pu
instrument.


down menu enables


The calibration


the user to calibrate the


includes the RF and DC voltages


along with the auxiliary board.


<"G>







The mass calibration is the calibration of the RF voltage. The
calibration can be performed with either the standard calibration


compound (PFTBA) or the user can


addition


the u


ser can use


nput the m


the custom scan functions to


to use.


calibrate the QITM


2-10.


Calibration editor for the DC voltage.


The calibration is


performed by attaching a voltmeter to J3 of the power board.


The respon,
determined.


e of the DC voltage supply to the set DAC is then
The slope and offset are then entered into the


editor.


2-11


Automatic gain control editor allows the user to use custom


scan function


AGC calculation


for AGC.


The parameters that are used


can be altered


in this editor.


for further details and an example using the editor.


n the


See Chapter 4


Block diagram of the auxiliary board used with Gatorware. T
board communicates directly with the microprocessor on the
SAP which allows different frequencies to be used during a
scan


function.


2-13.


Block diagram depicting the communication of the 80186


microprocessor with memory and external devices.


uses PCS lines to control which device is being communicated


with.


The auxiliary board uses the PCS


2 line.


2-14.


The auxiliary board editor lets tt
apply to the endcap electrodes.


changed during any table


ie user control the frequency to
The frequency can be


n a scan function.


The acquisition editor lets the user have the ability to acquire


data during a GC run.


The autosampler and GC along with the


QITMS editors can be reached from this screen.


The QITMS editor controls the acquisition of the data during a


GC run.


data acquired through this method are centroid.


Scan functions made


in ITS-Scan can be used during a GC run


set the background mass to 11


and type in the


SE Filename.


asses


The 80186








Oscilloscope traces of ramped RF and DC voltages.


The upper


trace is proportional to the RF voltage measured at the DAC out


(197mV/div corresponding to 32Vop actual RF) on the Analog
-a ~. a I -.- aa ... *


Board.


ower trace


the DC voltage (50 V/div) measured


*


at J3 of the Power Board.


The time scale is 0.1


ms/div.


Scan function depicting the qz, az values used for the
experiments. The az value for high mass ejection (Table 2) was


first examined


, followed by that for


ow mass ejection (Table 3).


Experimental determination of the stability boundary at qz=0.71


for single-step and ramped DC voltages.


The theoretical


stability diagram boundary is depicted by the solid line.


The approach to the stability diagram boundary of ions at a


range


n m/z: (a) Theoretical approach and (b) approach with


overshoot of the DC voltage.


Oscilloscope traces for changing the DC voltage on an ITS-40


from 0 to 130V
voltage change
desired value.


. The desired voltage is reached by one


V.


The overshoot can be as


arge


arge as 15% of the


Simulation showing the trajectories for m/z 100 at q


=0.710


while (a) az=0.00 and (b) az


excursion near the pz


=-0.235.


boundary.


Notice the
a . C .


arger


Plot of the efficiencies
m/z 57 and (b) m/z 5E


for isolation of m/z 56 and ejection of (a)
5 of 1-octene using ramped and single-


step DC voltage methods.


S108


Oscilloscope traces for changing the DC voltage on an


from 0 to 130 V.


TMS


The desired voltage is reached by (a) one


large voltage change and (b) two steps of voltage change with
an initial change of 90 V. . . . . . . .


The response


of the


on trap mass spectrometer for set


ionization times.


arbitrary


The space charge region is placed in an


location.


The response of the


on trap mass


spectrometer with AGC.


B_ --_ *_ ._ t -- -1---- --..---- -- .. .. ....


I


I


LI ,r


t







AGC scan function depicting the RF voltage and ion intensity


detected.


Notice the two section


of the scan function


and analytical scan.


Chromatogram showing the effect of AGC.


prescan


Furan was bled


on trap vacuum manifold at a constant rate (manifold


pressure


was


= 4x10-torr) during the entire run.


injected onto the GC column in the splitless


the complete loss of the furan


Dimethyl disulfide


on signal as dimethyl disulfide


Notice


elutes due to a large decrease in the ionization time.


MSAGC scan function depicting the RF voltage and


detected.


Two-step


intensity


solution is used during the prescan to


store and detect only the


on of


interest.


. 1


Structure of the compound (molecular weight M+4) used for the


experiments on co-eluting peaks.


sites for additional isotopic
(M+8). . . . .


abeling


The asterisks (*) denote the


n the


internal standard


Calibration curves for m/z


and (b) MSAGC.


199 (internal standard) for (a) AGC


Point A coresponds to 525 ng of the


compound and its internal standard.


Point B corresponds to


525 ng of the compound and 4 ng of the internal standard.


Average of triplicate


injections are shown


corresponding to +1 standard deviation.


with error bars


Mass spectra of 525 ng of compound with 4.0 ng of the


co-eluting


internal standard.


The mass spectra are for (a) AGC


and (b) MSAGC.


S 6 4 6 4 5 5 9 6 9 6 6 4 S 6 9 S S 6 6


Experimentally determined stab


hown by the dashed line


ity diagram [32].


Th


is the theoretical diagram.


e diagram
The


experimental diagram was determined using Ne+ ionized for the
different times shown.


Scan function used for determining the stability boundaries.
The only changes during the experiments occur during table 3.


Diagram depicting the slices of the stability diagram examined.


The smaller dashed


indicate the region where high


radel um44n ran 'nvr' Wfl fl,'ntt $If -sC%1 *, ,.


mode.







Experimentally determined stability diagram for Ar+


helium.


with no


ne denotes the theoretical stability


boundaries.


Blown-up sections (a) upper apex (b)bottom


The solid line denotes the theoretical boundary.


eft of Figure 5-4


Single,
helium.


lice of the stability diagram for Ar+


lice shown


non-linear resonances are


; depicted
indicated.


=0.70 without


n the inset.


The major


Topographical map showing the entire stability diagram for Ar+


without helium.


The map is for 0.2 ms ionization time.


solid line is the theoretical location of the stability boundaries.


Topographical map showing the entire stability diagram for Ar+


without helium.


line is the theoretical


The map is for 10m


onization time.


location of the stability boundaries.


solid


Location of the stability boundary for different mass-to-charges


atqg,


=0.700.


dotted


ne denotes the theoretical position of


the boundary.


. a S 4 a S 5 5 S


Change


n the az value for different ionization times


The changes
times. The d


are measured with respect to 0.5 m


otted


(qz=0.700).
onization


ne denotes the step size that was used for


these experiments.


Location of the stability boundary for different mass-to-charges


at qz=0.850.
the boundary.


Change
The char


times.


The dotted


ne denotes the theoretical position of


in the az value for different ionization times


iges


=0.850).


are measured with respect to 0.5 ms ionization


The dotted lines


denotes the step size used for these


experiments.


5-13.


Isolation efficiencies for different ion populations and mass-to-


charges.


solution efficiency is defined in equation 3-1.


Three ion profiles for different ion populations. Tht
f IrarI ,'s,, +1n U Li fl+ : S.,. .a It--. I "\ -A -


e ion probe
n ti -ai







The method used to determine the secular frequency.


frequency profile shown is for the C04


butanol (m/z 56) at q


corresponding to


z=0.205 and az=O.C


fragment ion from
10. The middle point


20% above the minimum value was said to be


the secular frequency.


Scan function u


ed to obtain the frequency profiles.


The only


section that was changed during the experiments was the
resonant excitation section.


Frequency dependence of the butanol ion (m/z 56) without


helium buffer gas.


The solid line denotes the theoretical values


Frequency dependence of the furan ion (m/z 68) without helium


buffer gas.


The solid


ne denotes the theoretical values.


Frequency dependence of the thiophene


helium buffer gas.


on (m/z 84) without


The solid line denotes the theoretical values.


The effect of ion population on the secular frequency


. The


secular frequencies for (a) q


depicted for a range of


=0.30 and (b) qz


on populations.


=0.70 are


The change of frequency versus set q


are (a) C4H8


, (b) C4H40


z values.


, and (c) C4H4S+ for


*he ion probes
on populations


of 30,000 counts.


The effect of different storage parameters,


az values for C4H8


e. q, valu


(


+ from butanol (m/z 56) without helium.


Aaz was determined by the


3n the
The


secular frequency [equation 6-4].


.205


6-10.


The effect of different storage parameters,


, q, values,


az values for C4H40+ from furan (m/z 68) without helium.


on the
The


Aaz was determined by the secular frequency [equation 6-4].


6-11


.207


The effect of different storage parameters, i.e., qz values, on the
az values for C4H4S+ from thiophene (m/z 84) without helium.
The Aaz was determined by the secular frequency
[equation 6-4]. . . . . . . . . .


The deviation of the experimentally determined qz value from
*.- I -| i_ .1 !.. _- ? .^ /-- I_ rn1j~ 2. -*-- _-.L. -- r- ^r ^







The deviation of the experimentally determined qz value from


theory for the furan ion (m/z 68) without helium.


denotes the theoretical response
the iterative approach. .


6-14.


The solid


which was calculated using


21


The deviation of the experimentally determined qz value from
theory for the thiophene ion (m/z 84) without helium. The so
line denotes the theoretical response which was calculated
using the iterative approach.


)


The change in qz versus the set qz. The
C4H8+, (b) C4H40 and (c) C4H4S+ for
30,000 counts. .. . .. .. .. .


Sion probes


are (a)


on population


6-13.












GC/MS/MS ON THE QUADRUPLE ION TRAP MASS SPECTROMETER:
SOFTWARE DEVELOPMENT AND


THE EXAMINATION OF THE EFFECTS OF


TIMOTHY P


ON POPULATION


. GRIFFIN


December 1995


Chairman


Richard A.


Yost


Major Department


Chemistry


Since tandem mass spectrometry (M


S/MS) was first developed as an


analytical technique

analytical problems.


it has become

Currently, tan(


the method of choice to solve many


dem mass spectrometry is performed


primarily on triple quadrupole mass spectrometers.


However, the


quadrupole


ion trap mass spectrometer promises to be the


next generation mass


spectrometer.


The high sensitivity of the


ion trap makes it ideal for multiple


stages of tandem mass spectrometry (MSn).


Tandem mass spectrometry


performed


n the ion trap by applying an


external frequency to the endcap electrodes.


When the external frequency


equals the frequency of ion motion,


increase.


the ion


This increase causes the ion


buffer gas and fragment.


Theoretically


absorb energy and their orbits


to undergo energetic collisions with


ion frequency can be calculated;







This change of ion frequency makes it difficult to automate the process of


MS/MS


on the quadruple ion trap.


This dissertation presents studies of the


effect of ion population upon the trapping parameters to help to predict


accurately the ion frequency.


n addition to the frequency studies,


a method


has been developed to automatically control the number of ions of a given


mass-to-charge ratio in the ion trap,


called mass-selective automatic gain


control


, to help alleviate the frequency


hifts


this should make it easier to


predict the ion frequency for MS/MS.

Ion traps that are currently commercially available are designed to be


routine analytical


instruments; therefore, user control of the trapping


parameters is not available.


The operation of the ion trap,


as opposed to other


mass


spectrometers, is software-intensive, meaning that most changes in


operation of the ion trap can be performed by software changes.


fundamental studies on the ion trap,


To perform


software was written to allow complete


control


of the trapping parameters.


n addition


, a digital frequency synthesizer


along with its control software was developed which allows the study of ion

frequency.













CHAPTER 1
INTRODUCTION


Background


Currently


tandem m


commonly on trip


spectrometry (M


quadrupole m


S/MS) is performed most


spectrometers [1].


high sensitivity


and high MS/MS


efficiency of the quadrupole ion trap mass


(QITMS) [2] make it ideal for multiple stages of tandem m


n addition


pectrometer

spectrometry


, the QITMS is software-intensive (rather than hardware-


intensive) which allows the user to change its operation quickly and easily.


QITMS


is still in the early stages of development and many fundamental


question


still need to be answered


instrument to hold promise


as the


, though its inherent capabilities show thi


next generation tandem mass spectrometer


Recent capab


resolution [4],


cities


and ion


of the


QITMS


njection[5]


include mass range extension[3],


.Th


features of the


high


on trap show that it is


a highly versatile


instrument.


However


, many of the


above-mentioned features


are only available on research grade instruments,


designed to allow flexibility


which enables the


researcher to develop techniques


that use


the ion trap to its


fullest potential (limited by the hardware).


To take full advantage of this









properties of the ion trap.


Most commercially available ion trap mass


spectrometers,


on the other hand


, were designed for routine analysis which


limits flexibility of the

straightforward. For


instrument, but makes their operation easy and relatively


example, most commercial ion trap mass spectrometers


are designed for routine gas chromatography mass spectrometry (GC/MS);


however


, only research grade ion trap mass spectrometers perform MS/MS


experiments.


Within the past few month


a new generation of commercial


instruments have been


introduced which promise routine GC/M


S/MS


on the


QITMS.


The manufacturers use different techniques


implement GC/MS/MS


and there is not yet consensus


by the scientific community as to which method


or methods give the best results.


The compact size of the QITMS


In addition


makes it


deal for a benchtop instrument.


, the ion trap has a higher sensitivity then beam


triple quadrupole) [2],


n part to high storage efficiency.


instruments (e.g.,


ons can be


trapped and detected for seconds or even minutes in the ion trap with higher


transmission rates than on the triple quadrupole mass spectrometer.


There are


limitations


however


to the number of ions that can be stored in the


on trap as


columbic interaction


between stored ion


in the confined space become


significant.


columbic


nteractions


, often referred to as "space charge,"


cause degradation of performance (e.g.,


loss of mass resolution).









challenge.


During a chromatographic run,


number of


ons that are formed


and stored can vary by orders of magnitude


as compounds elute from the


column


The efficiency of MS/MS


is dependent on the


number of ions in the


ion trap.


it is impossible to determine a single set of operating


parameters to use during an entire GC/MS/MS


these limitations previous


Techniques to overcome


ly have been performed in this laboratory [6].


aim of this work was to design methods to adjust automatically the MS/MS


parameters during a GC run to make GC/MS/MS

analytical method.


in the QITMS a routine


Specific Aim


To develop techniques to automate GC/MS/MS


n an ion trap mass


spectrometer, it was necessary to write new


as to


instrument control


implement several hardware modification


software,


This software enables


researcher to control all of the function


of the


ion trap


including some


previously available even with research grad


on trap mass spectrometers.


was with this software that the


effects of i


on population upon the operation of


ion trap were examined,


as wel


as the


new methods for GC/M


S/MS


automation.


need to develop a more powerful


research grade instrument was









not allow sufficient control of the


ion trap.


The ITMS u


an imbedded 8086


microprocessor coupled with an 80286 PC to control the


operation of the


QITMS.


The 8086 and the 80286 PC have limited memory


therefore, the


design severely limits the usefulness


can be very difficult to use


of the ion trap.


and does not always function


n addition


as expected,


software


in part


because there is not enough memory to permit easy-to-use software to be


written


A more advanced version of the


TMS


software called Ion Catcher


Mass


Spectrometer (ICMS) software was written


n this


ab by Nathan A.


Yates[7]


. Although


CMS was a marked


improvement over the original


TMS


software


it was sti


limited.


We have developed a new research grade instrument based upon a


commercially availabi


second generation GC/MS


instrument (Finnigan MAT


ITS-40).


TS-40 software is designed for routine


GC/MS


and allows very


limited control of the


ion trap.


modifications which were made were


designed to allow the user fu


control of the


instrument.


The source code that


allowed the modifications to be made to the


TS-40 was furnished by Finnigan


MAT


. Although most modifications to the


TS-40 were software, some


hardware changes were necessary.


new control software for the


TS-40 is


called Gatorware.


Procedures to automate MS/MS


on the


ion trap were developed









ion formation time


was also developed based upon automatic gain control


(AGC) first developed by Finnigan MAT[8]


. The


improvement


n mass isolation


is necessary to perform efficient MS/MS,


while varying the ion formation time


controls the


number of ion


produced and stored


n the ion trap.


The effects


of the numbers of ion


stored during operation of the


ion trap were


investigated in detai


Function/Description of the Ion


Trap


A cross sectional view of the


ion trap is shown


n Figure 1


.The


ion trap


consists of two endcap electrodes and a ring electrode.

all electrodes are machined to a hyperbolic shape. A ri


voltage and a direct current (DC) voltage are applied to the


The inner surfaces of


radio frequency (RF)


ring electrode.


The two voltages coupled with the hyperbolic shape of the electrodes create a

quadrupolar trapping field which allows ions to be stored within the ion trap.


n addition to the RF and DC voltages an auxiliary frequency can be applied to


the endcap electrodes which enables the


as ion excitation for MS/M


from electron


examination of the ion frequencies as


filament provides electrons to form ions


onization (El) of neutral compounds within the QITMS, while the


detector is an electron multiplier used to detect the ions as they are


from the


ected


ion trap.













0)
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0. ~cn.

0-0)23





g Ca




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rL
I : I ]=



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U)


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cutc ,
t a S.
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sc 0n C 0


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.0 C-
OE ^


na a
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E 4c 0.


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cn r


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I


Y U
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m
























































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-U









to the ion trap are controlled by a 80186 microprocessor


on the


TS-40


which


ocated on the scan and acquisition processor (SAP) board.


controlling the voltages applied to the ion trap,


data acquired from the detector.


n addition to


the SAP also processes the


The data can then be sent to the PC and


downloaded to the hard drive or displayed on the screen.


All post-acquisition


routines


are performed on the PC.


The SAP controls the voltages by


sequentially stepping through a


ist of


instructions or tables


known as scan


tables.


The entire


ist of tab


called a scan list (or scan function).


A diagram of a typical El scan function is


shown in Figure 1-3.


The top


trace


the amplitude of the RF trapping voltage applied to the ring electrode


while the bottom trace is the detected ion signal.


SAP sets the RF voltage to zero for a set time.


n table 1


n table


(Figure 1-3) the


, the SAP applies


correct RF value to the ring electrode and the electrons are gated


nto the ion


trap to form ions.


Table 3 requires the ramping of the RF voltage.


During the


table


, the electron multiplier,


ocated behind an endcap electrode,


enabled.


This is accomplished by raising the voltage applied to the detector to give a


gain of 105


voltages typically run at


-1100 to


-1600 V.


When the detector is


not active the voltage is lowered to a value less then the operating voltage to


conserve the life of the detector.


are then ejected from the ion trap


order of increasing m/z and detected by the electron multiplier


. A mass
















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Onk



,r)



C- r
v E



fl~ C
E~ -,
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CU oq

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eeo







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oww
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S












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- S~


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tc









Theory of Operation


Fundamental Parameters


Three fundamental parameters of the quadruple ion trap are important to


understanding how the ion trap operates.


The fundamental parameters are qz


and f


and are discussed


ength


n Quadrupole Storaae Mass


Spectrometry [9].


The parameter q


is related to the RF voltage by


2
m (r+


Where V is equal to the RF voltage (0-peak),


(1.60219x101' C),


mol-1)-1


2 2


e is the charge on an electron


m is the mass of the ion ((u)xlO'kg mol'1)(6.02205x1023


, zo is the distance (m) from center of trap to the endcap electrode,


is the distance (m) from the center of the trap to the ring electrode, and Q is


2xf where f is the frequency of the RF voltage (in Hz).


The az parameter is


related to the DC voltage (U) by


-8eU


2
m(r+ o


2)Q2'









n addition, if the


a, and q,


are known B


which will be related to the


frequency later


can be calculated from the equation


(2 + z)


(4 +z)


(6 +flZ)


-...etc


(1-3)


-...etc


Which can be simplified (for qz<0.80) to


-1)q


+7)q


z+58a

-1)s(a


+29)q

-4)(a,


64(a


Stability Diaqram


Different combination


of az and qz


cause ions of particular m


ass-to-charge


(m/z) ratios to be stored within the


ion trap.


A plot of the regions of ion


stab


vs. values of azqz,


called a stab


ity diagram,


is shown


in Figure


plot shows region


of stability


n the


z direction


, between the endcaps,


-1)3~a




















No

SOC




Wa,


coz
*-

Sow
Co v
4- a


CC[)
00
.-



Z'w
.- 4-



UJ )
-C-C


Cc
P.C


(A
(A

0r *

C E
0

a3)


(U
4- 4























































c'I 0 c'J









in the ion trap where


z stable


and r stab


region


overlap (shown in Figure 1-4


by the


cross-hatched region).


n other words


are stable and can be


stored within the ion trap when combination


correspond to az, qz points that lie within the


ons that are stored within the


of RF and DC voltages

cross hatched region.


on trap move with a particular frequency of


motion


secular frequency, which is a function of the ions m/z,


as wel


the az and qz values.


frequency that the ions move within the ion trap is


related to B


by the


equation


2


Where wo


is the


on secular frequency (Hz) of motion


n the


z direction and Q


the frequency (Hz) of the RF voltage applied to the ring electrode.


Notice


n equation 1 that q


nversely proportional to the m/z of the ion.


For a given RF voltage,


This is


ons


hown graphically by the


with lower m/z values are at higher q


circles


values.


n Figure 1-4; the relative size of the


dots denotes the relative m/z of the


A m


pectrum is acquired


n the


mass-selective instability scan [10] mode when the RF voltage is ramped whi


DC voltage is held at zero


the ions become unstable, and thu


are ejected


from the


ion trap in order of increasing m/z.









Tandem Mass


Spectrometry


The method most commonly used for fragmenting ions to perform tandem


mass spectrometry is collision


nduced dissociation (CID).


On the ion trap,


MS/MS


includes five steps (warmup,


ionization


solution


, CID, and detection),


a seen


n the scan function shown


n Figure 1


The warmup step (table 1


Figure 1-5) gives the SAP time to reset and


warmup the electronics.


onization step (table


, Figure 1-5) is the same


as for the normal El scan function (Figure 1-3).


During the ionization step


electrons are gated


nto the


on trap.


The electrons col


de with the sample


eluting from the GC causing


ons


to be formed.


The RF voltage then traps and


stores ion


that fall within the stability region in Figure 1-4.


To fragment only


the ion of


interest it is necessary to store ion


of a single m/z


n the ion trap


(called isolation) before fragmentation can be done.


The isolation step (table


, Figure 1-5)


As is


ejects al


hown


but ion


of the m/z of


in table 4 (Figure 1-5),


after


interest from the ion trap.


of a single m/z are stored


within the ion trap an auxiliary frequency is applied to the endcap electrodes.

When the auxiliary frequency corresponds to the secular frequency of the


isolated ion


increase causes


they absorb energy and the size of their orbits increase.


the ion


This


to undergo energetic collisions with the constant


pressure of helium buffer aas that is


n the ion trao.


As the ions collide with


I


*















(U
-Cu)

(U)




C)
*) -

Or
c~a


"4'
E


SC)
Cu -


COL
5U)
(U
v)o



ESC
O)
cO 4
C o



CU0

Q.
.C ,
* -
UO
c 6
-


*Cc

Cc
o -
CC


Cw
C Q
O
W e
































CU





CI

0

CO
CU





~a




C

CO
0d
o0
-


0J)
Cc
'-C
cO
00


O -

OC


CO









sequentially ejected from the ion trap,


using the mass selective


instability scan


to acquire a m


pectrum (table


5, Figure 1-5).


RF/DC


solution


To distinguish which of the detected ion


from those ions which were formed during ionization,


are fragment ions formed by CID


imperative to isolate


a single m/z


n the


on trap before CID occurs.


solution is performed


number of ways


including apex [11]


two-step [1


,13],


white noise[14]


broad


band[15],


, SWIFT[16]


filtered noi


e field[17]


and forward-reverse scan


s[18].


All of the methods except two-step and apex use auxiliary frequencies applied


to the endcap electrodes.


The use


of frequencies can make the techniques


difficult and problematic to use when sample concentration varies (see Effect


of Ion Population,


below)


Therefore, two-step isolation was used primarily for


the work presented


n this dissertation.


Two-step isolation is


hown graphically


n Figure 1-6.


The ions are first


formed and stored at a low qz and az


are then set so that the ion of


= 0 (point A).


interest


The RF and DC voltages


ust within the stability diagram at


point B.


At point B the higher m/z are no longer stable within the ion trap and


are ejected.


After the high masses are


ected from the ion trap the RF and


DC voltages are changed until the


on of interest


ies within the stability

















"tl.


-N
4-



4-
C)U





(Un

4-'
4 .-
C~U



a-

'A,,
4-Cx,



CU -
r(


I-n!

EE
-Co
O)cn,





















































('U 0 c'J tJ* CD









voltages are then lowered to correspond to point A in Figure 1-6.


experiments


Further


including CID can then be performed on the single m/z ion


stored within the ion trap.


Two-step isolation requires exact knowledge of the


location of the stability


diagram boundaries


to use


the correct RF and DC voltages.


Any movement of


the stab

addition


ity diagram boundaries can affect the efficiency of isolation.


, the efficiency of isolation might vary depending upon the compound.


The reason for the variations are not understood.


problems


demonstrate the need to


improve the


olation techniques.


Overview of Dissertation


This dissertation focuses


on two areas.


software to control the ion trap mass


the use


the development of the


spectrometer (Chapter 2).


of the software to examine ways of automating GC/MS/MS


The second is


n an ion


trap mass spectrometer (Chapters 3-6).


Chapter


2 provides an overview of the software that was written to


control


TS-40 ion trap mass spectrometer


. This dissertation includes a


detailed description of the control software and the functions that are made


available by the software.


A description of the automatic acquisition routine,


along with examples,


is also included


n this chapter.







25
detailed examination of the effect of the method upon isolation efficiencies is

also examined.

Chapter 4 discusses a method to continually adjust the ion formation time


to control the number of ion


of a particular m/z that are created and stored.


The method is used to determine the linear dynamic range of a co-eluting drug


and its isotopically


abeled i


internal standard.


Chapter 5 examines the effect that stored ion population has upon the


operation of the ion trap.


The position of the stability diagram boundaries for


different stored ion population


is examined.


The effect of different mass-to-


charges also is examined.

Chapter 6 examines the effect that the stored ion population has upon the


effective az,q parameters.


The frequency of the ions


at different qz values is


examined for different ion populations.

secular frequencies is also studied in


The effect of mass-to-charge upon


close detail.


Chapter 7 presents the conclusions and suggestions for future work.


chapter is a summary of the work presented in this dissertation.


Possibilities


for future studies also are suggested.













CHAPTER


SOFTWARE DESCRIPTION


Introduction


The quadruple ion trap mass


spectrometer (QITMS) is a very versatile


instrument, primarily due to its software-intensive nature.


Properly designed


software enables the function of the QITMS to be varied by a simple


command.


This has allowed many facets of the QITMS


to be investigated and


operating techniques to be developed.


This dissertation will describe software


and hardware, named Gatorware, that was developed to control a

commercially available QITMS (ITS-40 by Finnigan MAT).


TS-40 was designed


as a routine GC/MS instrument and as such


the user has very limited control of its function.


Gatorware was designed to


allow the user complete control of the


research grade


TS-40's


operation.


instrument was developed from one


In this way a


designed for routine


use.


The goal of Gatorware was to make an

techniques which make GC/MS/MS on


instrument to allow the development of


an QITMS a routine analytical too


While Gatorware was designed for


ease


understanding of the


fundamentals of the QITMS


is necessary.


An option also was


included that









ectronics and in particular how the


TS-40 operates.


Gatorware is


, in many


ways,


based on the


on Catcher software (ICMS) [16] developed in our


laboratory by Nathan A.


Yates.


CMS, however, is for use


with the


TMS (see


Chapter 1) and thus does not provide


the versatility that is available with


Gatorware.


The source code for the


TS-40 which enabled the development of


Gatorware was furnished by Finnigan MAT.


This chapter of the dissertation gives a detailed


ook at Gatorware.


included is a walk through of al


the features that are


incorporated


n the


software.


A simple


explanation of how the microprocessor that controls the


TS-40 operates also is included.


automation of the


New procedure words which permit


experiments are explained and examples given.


Overview of Software


Gatorware is a DOS-based system which can be entered by typing





at the DOS prompt (in the correct directory).


The first screen seen


upon entering the software is the Exec Page (Figure


The Exec Page is


the primary screen of the software and allows access to lower level routines.


For example, the screen


in Figure


1 shows a listing of the user programs that


are available.


The primary routine


of Gatorware is called


TS-Scan.


This


dissertation only describes those functions not available on the unmodified


TS-40


unless otherwise noted.






















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The major display of


TS-Scan (Figure


2-2) allows the user to adjust the


SAP and thu


the function of the QITMS.


The software is mouse


driven,


although textual strings can be entered through the keyboard,


which facilities


the use of the software.


The primary screen (Figure


major areas: the pull down menu,


the mass spectrum,


-2) consists of three

and inputs for nine


major parameters.


n addition


, the multiplier


filament, and RF voltages can be


enabled by a click of the mouse button.


parameters and see,


The ability to adjust the trapping


n real-time, how the mass spectrum is affected enables


the user to smoothly and quickly fine tune the operation of the QITM


Down Menu


The pu


down menu allows the user to the set up the


instrument and


software before acquiring a mass spectrum. I

(Figure 2-3) the user can decide if the data wi


For example, under options


be displayed in profile or


centroid mode. Prof

DAC steps and thus,


ile shows al


of the data points that correspond to the RF


values between integer mass units (u) are seen.


Centroid


converts the profile data to


nteger u values so that lines corresponding only to


integer u are


in the spectrum.


Acquiring


n centroid mode


limiting and can


cause


experimental errors.


For example,


if the peak corresponding to a m/:


interest does not lie


n the correct


location the centroid routines can split the


























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~-u)


o i
ci:SP

a>
("(
c,)
VTi t
o n


* A


ti)
(4-a



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00 3







































































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N
01
0


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C





V
AC-O


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OW


o c
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34










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m A Ea F
ra.
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Therefore, routines were written to acquire both profile and centroid data to


ASCII files. To

selected in the


set up the acquisition file,


menu.


"Options


This option allows the


Real Time Acquisition" is


user to select centroid or profile


data, enter the mass range,


and the name


of the file to write.


After the file


set up by the


menu option,


Acquire button at the bottom center of the main


screen can be highlighted.


This button tells the software to write the data to


the disk after every mass spectrum is taken.


The data that are collected


n this


manner can be manipulated


n any spreadsheet program that accepts ASC


files.


A program,


Gatorwork


also


was written to manipulate the


ASCI


data


files acquired in Gatorware.


Many of the


function


that are made available


n Gatorware required


extensive changes to the microprocessor control software on the SAP. The

modified software is called GATORware, as opposed to Gatorware. Before


TS-Scan can be used


GATORware must be downloaded


downloaded upon entrance of Gatorware.


, it is automatically


It is often necessary,


however, to


download GATORware while


is selected from the menu (Fik


n ITS-Scan; to do so "Control I Load GATORware"

3ure 2-4).


Spectrum Display


The middle section of the


can main screen


hows the currently


















0
.4
1..
0)






CA


Cc'



DO
E*



i-a,
0


.4-E

Ca


0)0
4-



aC0



QC

0o

CC





o r





~cn



























z
0
Co)
C
r

C
I
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S:
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: 04 0

0 000
e i-I .


Ps Ir It I
I rln lri


.34d

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.ao-
0.0.0
flc~









feature also is available by clicking on the normalize button (N).


The values


100% of the displayed scale along with the total ion intensity (TIC or RIC) are


hown at the top of the display.


The ionization time and currently used scan


function also are shown.


In addition


, the x-axis scale can be adjusted by using


the buttons located to the bottom


be selected by dragging the mouse


,ft of the spectrum or the mass range can

over the required range on the x-axis.


Parameter Button


Located below the mass


pectrum display are


nputs for nine of the


most frequently adjusted parameters.

table are shown and can be adjusted t


The values that are used for the current


n real-time, the effect that the


parameter changes have upon the mass spectrum.


In addition


six of the table


flags can be toggled from this display.


The currently active table number also


is displayed along with arrows to change the active table.


jumped to by clicking on the table number


Any table can be


. Clicking on the displays the


Scan


Table Editor to allow control


of al


of the QITMS's


operations (see Scan


Table Editor below).


Scan Editor


The operation of the QITMS can be altered by changing the scan









reaction time.


n order for the operator to have complete control of the


operation of the QITMS


a Scan Editor was designed.


The editor consists of


three major parts.


The first is the Clipboard Editor which enables the


modification of the entire scan function.


The second


the SAP Table Editor


which modifies


the SAP control


ines.


Scan


Table Editor is the third


section.


This editor is a simplified method for control


ng the SAP control


nes.


Clipboard Editor


sting of the scan function that is being edited can be shown by


selecting "TableI Scan Clip"


. The Scan


Table Clipboard (Figure 2-5) shows a


listing of the tables that compose the scan function,


a simple El scan function


n this case (Figure 2-5).


n addition


the tables can be erased


inserted


their order changed.


These abilities by themselves are useful; however, the


flexibility of the software is further enhanced by the addition of table editors


that allow each table to be


individually modified.


SAP Table Editor


The SAP table editor (Figure


2-6) allows the user complete control of the


SAP


The editor can be entered by clicking on the SAP Table box


n the Scan


Table Editor


. The editor enables the user to enter the values that the SAP






























Figure 2-5.


The scan table clipboard allows the user to control


the order of


the tables in the scan function.
deleted from the scan function.


Tables also can be added and




















Scan
Scan ile :

Scan


Table


DEmO .SCN


tt
t Saved


Tables:
Tab les:


Table List


SAUE

INSERT


First Table ----
Prelonize (Uarn up)
Ionize
Post Ionize (Cool)
Preacquire (Multiplier Warnup)
cqu ire
Reset


< This


ERASE


Clipboard


TbiN l# 1


hilrn~





























Figure 2-6.


The SAP table editor allows the user to directly control the lines


that the microprocessor uses to operate.


most flexibility; however
electronics is necessary


This editor allows the


thorough knowledge of the QITMS




























TABLE EDITOR


1 First Table
Table Mass.....
Scan Flags.....
Start RF DAC...


DAC Step Size.
#Steps of Data
Data Address..
#Steps No Data
UUait Loops...


User 8.
User 1.


* *S*~*** *5* *


* ** St**tSt~


* *S.tt*.......... *0*
* S **S* S 55*S***** S


User


User 3.
User 4.


User


* ** *5~~SS*S S S 55*t*
* S S *** . S* *S~*C
* .. USS****** S S S S


User 6.


38.81*


I 8a
I 01











|I 6



8


0 CALC SAP*SCAN


SAP









control the RF voltage to apply to the ring electrode.


While this allows the


most flexibility


n the operation of the QITM


it requires that the user have a


thorough knowledge of how the electronics operate, making the editor difficult


and time-consuming for most experiments.


Gatorware, the Scan


Scan


To help facilitate the use of


Table Editor was written.


Table Editor


Scan


Table Editor (Figure 2-7) has


nputs for fundamental trap


parameters such as az and qz,


values.


and automatically calculates the SAP control


The editor can be entered by clicking on the "E" located on the bottom


left of the main screen of


TS-Scan.


table mass also are available.


nputs for table time, start and end q


n addition


, the user can select table types, e.g.,


acquisition,


ionization


or AGC.


The table mass and qz or az values are used


to calculate the RF or DC voltage (equation 1


, Chapter 1) to apply to the


ring electrode, after which the correct digital-to-analog converter (DAC) values


are calculated and sent to the SAP


n this way, the user can use


the same


scan function for examining ions of different m/z


necessary is to the table mass.


instrument while retaining the


the only change that is


The scan table editor facilitates the use of the


versatility that is necessary for many


experiments.


To use


this editor


, knowledge of the fundamental trap





























Figure 2-7.


The scan table editor


is a simplified editor.


The editor requires


the user to know the fundamental parameters of the QITMS


however


, no knowledge


of the electronic is necessary.





















Scan


Table


Editor


-I-


FEirst


Table


J Acquire Flag
n Iult Flag
n Ion Flag
Tr igger Flag
D TTLs
5 Wait
[3 DC Flag

5 FAGC Editor
n Aux Editor


Table Mass


Tline


Start q
End q
Mass Cutoff
Acquire Range
Start az
End az


I 38.80
493
0.988i

8.80

O.OO- O
|e 9.6896 1
| 9.60996


n SAP Table


Em-


CALC SCAN-SAP


SL- Table [-"


Table


E l


~k~m~a


U-]









Custom Calibration


In order for the correct RF and DC voltages to be applied to the ring


electrode, the voltages must be calibrated.


TS-40 only allows for very


mited methods for calibration of the RF voltage; no means are available to


calibrate the DC voltage.


The RF calibration was modified to enable the user


more control of the calibration.


n addition


DC calibration was added to


Gatorware to ensure the correct values


are applied to the ring electrode.


RF Calibration


The QITMS calibration routines were modified to allow the user to select


the scan function to use for calibration.


TS-40 only allows calibration


using perfluortributylamine (PFTBA) with a normal El or CI scan function.


this reason custom calibration routines were written for Gatorware.


These


routines


are selected by the "Tools


Mass Calibration" pu


down menu (Figure


2-8).


The Mass Calibration Box (Figure 2-9) has options that can be selected


using the mouse and includes user masses and the ability to use a custom


scan function.


The user masses option furnishes the user with the ability to


calibrate with compounds other than PFTBA.


The ability to use custom scan


functions enables different techniques,


other then GC


to be used for the


introduction of samples


into the QITMS.


This function is very helpful for


















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E


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c'J





































04'4"'
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Figure 2-9.


The mass calibration


the calibration of the RF voltage.


calibration can be performed with either the standard calibration
compound (PFTBA) or the user can input the masses to use. Ir


addition


the user can use the custom scan functions to calibrate


the QITMS.
























Mass


Cal vibration


Calibrate


FC-43


n User


Masses


O Use


Custom


Scan


Function


1 Display

(I


Current


Ca l ibrat ion


Cance


Curve
C o'.l/


Dk









DC Calibration


n order to apply the DC voltage to the ring electrode, the value to enter


nto the DC DAC must be calculated.


calibration of the DC voltage is


performed by selecting "Too


Setup DC Voltage"


n the pull down menu


(Figure 2-8).


A voltmeter is then attached to J3 on the power board of the


The calibration slope and intercept of the DC DAC value versus the


measured DC voltage output is determined.


The slope and intercept are then


entered into the software (Figure


Automatic Gain Contro


Editor


Automatic gain control


developed by Finnigan MAT


(AGC) and automatic reaction control (ARC),

[6] are methods to control the ion population


formed and stored


n the QITMS.


AGC for El


and ARC for CI


are methods to


automatically adjust the ionization time to keep the number of stored ion


of the space charge region.


A detailed description can be found in chapter 4


of this dissertation.


The Scan


Table Editor has a


ub-editor for AGC and ARC which is


selected by clicking on the AGC Editor Box (Figure


(Figure


The AGC Editor


11) allows new methods of AGC and ARC to be developed and


nntimi7Air


nhsntor A. fnr vnoimnntQ a icinri tho A(^l ra tr




























Figure


Calibration editor for the DC voltage.


The calibration is


performed by attaching a voltmeter to J3 of the power board.
The response of the DC voltage supply to the set DAC is then


determined.


The slope and offset are then entered into the


editor.























DC

Set


Slope
Offset


DAC


(U/DAC)
(Uolts)


UALUE


-8.82888
8.014088


Ok


Cancel






























Figure 2-11.


Automatic gain control


editor allows the user to use custom scan


functions for AGC.


calculations
further detail


The parameters that are used


can be altered in this editor


and an example u


n the AGC


. See Chapter 4 for


ng the editor.

























AGC
l AGC Flag


Editor


Target/Scaling


O EI
flcx


AGC Flag
ARC Flag


Ion Time


. Ion


Time


Rxn Tine


I e!
8


Ik~rml









Auxiliary Board


The ability to select the frequency and amplitude of the auxiliary

frequency to apply to the endcap electrodes permits a frequency profile of the


ions to be performed.


The auxiliary board allows the auxiliary frequency to be


set equal to the secular frequency of motion.


When th


two frequencies are


equal,


the orbits of the ions increase.


The increased orbit can cause CID or


ion ejection to occur


. When the energy absorbed by the ion


increases


the ion


orbit within the boundaries of the electrodes


, more energetic collisions


with the


helium buffer gas occur causing fragmentation.


When the orbit of the ions


increase beyond the boundaries of the electrodes ion ejection occurs.


cases, CID and ejection,


n both


the ion intensity decreases when the auxiliary


frequency equals to the secular frequency of the ions.


It is important to be


able to control the amplitude of the auxiliary frequency to effect CID.


f the


amplitude is too


arge,


the orbits of the ion


become so large that the ions


strike an electrode.


Therefore, the amplitude must be


increase the orbits while small


arge enough to


enough to prevent ion ejection.


board (designed and built by Nathan Yates,


Scott Quarmby,


An auxiliary


and myself) and


control software (designed and written by Nathan Yates and myself) were

designed to allow the user either to perform CID or the acquisition of
fran tm i ,,n / ra",f lao









Auxiliary Board Description


A diagram depicting the auxiliary board is shown


board can be seen as being composed of three parts.


n Figure


The first is the


decoding which allows the microprocessor on the SAP to communicate with


the auxiliary board.


The second is the frequency synthesizer that applies the


frequency to the endcap electrodes.


The third is the


n which lets external


devices trigger the QITM


The communication of the


frequency board to the microprocessor


located on the SAP is performed as is shown


n Figure


The 80186


microprocessor uses a 16 bit bus to communicate with memory and external


devices.


A detailed description of the operation of the 80186 microprocessor


can be found in Microsystem Components Handbook--Microprocessors


Volume 1


1986[19].


The architecture of the microprocessor uses peripheral


chip select (PCS) lines to let the external devices know if they should read the


data on the bu


Up to seven peripheral devices can be controlled by the


microprocessor (PCS 0


The PCS


correspond to specific address


blocks.


For example, (with the base address equal to 8000h) when an


address between


807Ch-81 FFh


written to or read from


the PCS 2


activated.


n this way the microprocessor can communicate with a specific


IAvirnA tn nArfnrm an nnwratinn


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4-4


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Table


Auxiliary Board Memory Map.


Address Description
(Hex)
00 DDS1 Phase Increment A (PIRA) bits 0-7 (LSB)
02 DDS1 PIRA bits 8-15
04 DDS1 PIRA bits 16-23
06 DDS1 PIRA bits 24-31 (MSB)
08 DDS1 Phase Increment B (PIRA) bits 0-7 (LSB)
OA DDS1 PIRB bits 8-15
0C DDS1 PIRB bits 16-23
OE DDS1 PIRB bits 24-31 (MSB)
10 DDS Synchronous Mode Control (SMC)
12 DDS1 Reserved
14 DDS1 Asynchronous Mode Control (AMC)
16 DDS1 Reserved
18 DDS1 Accumulator Reset Register (ARR)
1A DDS Reserved
1C DDS1 Asynchronous Hop Clock (AHC)
1E DDS1 Reserved
20 DDS2 PIRA bits 0-7 (LSB)
22 DDS2 PIRA bits 8-15
24 DDS2 PIRA bits 16-23
26 DDS2 PIRA bits 24-31 (MSB)
28 DDS2 PIRB bits 0-7 (LSB)
2A DDS2 PIRB bits 8-15
V fl n fllwflf L..n i-'. 4 r. an









Table


-- continued.


Address Description
(Hex)
32 DDS2 Reserved
34 DDS2 AMC
36 DDS2 Reserved
38 DDS2 ARR
3A DDS2 Reserved
3C DDS2 AHC
3E DDS2 Reserved
40 DDS Control Port (16-bit register)
44 DDS1 & DDS2 HOPCLK (synchronous)
48 DDS1 & DDS2 PMCLK (synchronous)
4C DDS1 & DDS2 EXTMUXCLK (synchronous)
50 Amplitude Control DACA
54 Amplitude Control DACB
58 Balun DACCS
5C BALUN DACST
60 No Connection (N/C) Spare
64 N/C Spare
68 TTL Out Strobe
6C N/C Spare
70 N/C Spare
74 N/C Spare
78 N/C Spare
ae a a .n a







65
assembly program that controls the microprocessor uses a structured variable


list so that the control words correspond to the correct memory locations.


listing of the control words


is shown


Table


It is important to note that


the control words are written


n the same order as the memory map and extra


padding is used to line up the control words to the correct memory locations.

The Qualcomm Q2334 dual direct digital synthesizer was the chip used


to synthesize the frequencies.

independent frequencies. Th<


frequency


The Q2334 has the ability to output two


e starting phase also can be selected for each


A direct digital synthesizer (DDS) works on the basis that a sine


wave can be generated by accumulating a predetermined phase


increment.


This phase increment value can be calculated based on the following equation.


(2-1)


where FG


the generated frequency


is the clock frequency


A@4 is the


phase


ncrement value, and N is the number of bits in the phase accumulator.


A detailed description of the Q2334 can be found


available from Qualcomm[20]


n the


Technical Data Sheet


. The amplitude of the frequency


is controlled by


a multiplying digital-to-analog converter (DAC) that is controlled by the


software.


The frequency is coupled to th


endcap electrodes


by a balun


circuit provided by Finnigan MAT.










Table


List of control word (Assembly Language).


Variable Size

PI 1 A 0 word

PI 1 A 1 word
I- -r -I

PI 1 A 2 word

PI 1 A 3 word

PI 1 B 0 word

PI 1 B 1 word

PI 1 B 2 word

PI 1 B 3 word

SMC 1 word

AMC 1 word

ARR 1 word

_A HOP CLOCK1 word

PI 2 A 0 word
I -I -r
PI 2 A 1 word

PI 2 A 2 word

PI 2 A 3 word

PI 2 B 0 word
-- -r
PI 2 B 1 word
1- -
PI 2 B 2 word

PI 2 B 3 word
SMC 2 word

AMC 2 word


ARR 2 word









Table 2-2 -- continued.


Variable


Size


S HOP CLOCK word
PM CLOCK word
EXTMUX CLOCK word
DDS AMPL A word
DDS AMPL B word
BAL DACCS word
BAL DACST word
ADC STROBE word
ADC CHIP SELECT word
TTL OUT word
TTL IN word









user


nputs; if the values


are equal the


QITMS operates as normal.


If the


values are not equal the QITMS will hold at the


current RF and DC voltages


they are equal.


In this manner the


QITMS wi


idle at the


given RF and DC


voltages until external devices tel


it to start scanning.


This can be used to


synchronize the


QITMS's


mass spectrometer


. The


operation with another device, e.g.,

software was written so that the TTL


a time-of-flight


n is compared


to the input values at the


beginning of the scan tables,


, before any


changes are made for that tab


Auxiliary Board Editor


The Auxiliary Board Editor is


hown


in Figure


14 and can be accessed


by clicking on the Aux Editor button in the Scan


Table Editor


. The


editor has


inputs for control


ng the frequency


amplitude, and starting ph


for one


auxiliary frequency


frequency can be adjusted from approximately 50kHz


.1MHz.


limit for the


frequency is primarily from the


balun circuit that is


used to coup


the frequency synthesizer to the endcap electrodes.


synthesized frequency,


however


cannot be more then half of the


clock speed


used to drive the


be varied from 0 to


Q2334 (4MH


10 Vo_.


clock speed


starting ph


in this case).


amplitude can


can be adjusted by


increments


of 45


0. Th


ese


controls allow the


maximum flexibility that could be designed






























Figure 2-14.


The auxiliary board editor lets the user control the frequency to


apply to the endcap electrodes.


during any table in a


The frequency can be changed


can function.


























Aux


Board


Editor


Board


Outputs:


U Aux Freq


Frequency


Start


Phase


Amp itude


(Hz)
e (deg)
(nU)


i Aux Ampl


Amplitude


(mU)


I 51
a
a

a
I---"

I---5


~cr~irm~









written to use both channels of the frequency synthesizer (see


Table


and 2-2)


memory limitations


TS-Scan (


,64kb for FORTH) permit the use


of only one channel.


An amplitude control,


however


is available for a second


device, such as an arbitrary waveform generator


GC Acquisition


acquisition control


the Exec Page.


for the


TS-40 can be entered by typing


The acquisition editor is shown


n Figure





The software


lets the user name


the file to which to write the data (DATA FILE).


In addition


the user can set up the autosampler (AS METH),


the gas chromatograph (GC


METH),


and the


QITMS


(ACQ METH).


The editor for the QITMS is


shown in


Figure


The values that are normally able to be adjusted are shown on


the left hand side of the editor.


The software was modified to permit the use of


scan function


built


TS-Scan.


nput


SE filename


the name of the


scan function that was built and saved to disk (in the same directory as the


method file).


When background mass


is set to eleven (11),


the microprocessor


will use the custom scan function


all other times


use the scan function


normally used on the


TS-40.


n this manner


, both types of scan functions can


be used and a comparison of the custom scan function can be made those


normally available.


The control buttons to enable Auto ion control (AGC or

























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Procedure Lanauaae


Many experiments that are performed on the QITMS


chromatographic run


are not simple gas


The experiments are used to determine the effect of a


specific parameter upon the function of the QITMS (for example, the


determination of the optimum qz to use


for ionization).


The optimization


employs a series of experiments that sequentially changes the qz value and


takes a mass spectrum after each change.


n this manner


, a plot of ion


intensity vs.


can be used to determine the optimum q


value.


While the


experiments can be performed by hand,


the ability to automatically perform


these experiments save time and enables a more detailed look at the


parameter under investigation.


The unmodified ITS-40 software does not have


any method to obtain data other then centroid acquired during a GC run.


Therefore, routines were added to the procedure


language to permit


acquisition of both profile and centroid data.


n addition


methods to automate


experiments also were


incorporated.


The procedure routines


make available


of the routines


that are


incorporated


TS-Scan.


The scan functions are


first made in


TS-Scan


the procedure routines


are then run to automatically


change


specific values


in the scan function and acquire the mass spectra.


Procedure Words











Start of a PIF-PTHEN or PIF-PELSE-PTHEN conditional structure.


Example


(if we make PEAK-INTENSITY an integer)


FORTH
PEAK-INTENSITY 2000 PROCEDURE PIF
FORTH


NCREASE-IONIZATION-TIME


PROCEDURE
PELSE
FORTH
GET-NEXT-SCAN
PROCEDURE
PTHEN


Comment


If the statement


true (i.e.,


PEAK-INTENSITY


< 2000) we


increase the


onization time


, otherwise we get the next scan.


The PIF


statement does not have to have the PELSE statement however, the
PTHEN statement tells when the argument is over and must be present.


Any compare function


(e.g.,


=) can be used.


PELSE
See PIF.

PTHEN
See PIF.


SCAN-ENTER
Enters the scan editor for the procedure language.


SCAN-EXIT
Turns off all control lines to the QITMS and exits the procedure.



Starting position of a procedure loop operation.


Example:


(SCN-COUNTER


the number of times


eft to loop )


FORTH 10 PROCEDURE





FORTH
SCN-COUNTER
PROCEDURE












See the example for .


n order to change the appropriate values


n the scan function


the user


must know the correct names of the table flags (Table 2-3),


the table scan


values (Table 2-4),


and the table SAP values (Table 2-5).


Any of these values


can be modified during a procedure by using FORTH commands. In order to

change the variable the value is put on the stack before the variable. It is


subsequently stored by either D->


-> for double or single precession


numbers


, respectively.


important to note that FORTH u


only integer


arithmetic and thus


, no decimal points are used.


Example Procedure Proarams


An example procedure program (with comments) is listed below.


example procedure is meant to show how to turn on the


instrument and set up


and load a custom scan function from the computer's


hard drive.


After the


scan function is loaded from the disk the values


can be changed as desired.


The scan function can then be used for the subsequent acquisitions.


included is an example of opening an ASCI


can be opened at one time (numbered 0,


Also


acquisition file; up to three files


GATORware also can be


- a


.1 A. A.nr I,.. t%, L,.,, flA L ...a. .f ~t


I' 'rrr


rr


n ,r,,,




















Table 2-3.


Scan


Table Flags for Procedure.


ION-FLAG Ionize during this table
PROTECT-FLAG Turn multiplier on
DC-FLAG DC voltage enable not used for ramped DC voltage
TRIGGER-FLAG Trigger enable

ACQUIRE-FLAG Acquire during this table only
TTL1-FLAG Spare TTL 1 pulse

TTL2-FLAG Spare TTL 2 pulse
TTL3-FLAG Spare TTL 3 pulse

CALC-AGC-FLAG Calc AGC values AGC scan table
EI-AGC-FLAG AGC ionization table variable time
SCAN-TBL-FLAG Tells which editor to use (SAP or SCAN)
TABLE-CALC-FLAG Enables calculations from Scan Table to SAP Table

EXT-FREQ-FLAG Auxiliary frequency board enable
EXT-AMPL-FLAG Auxiliary amplitude control enable


TTL-IN-FLAG


Turn on


TTL inputs


---- --
-------





















Scan


Table Values for Procedure.


TABLE-MASS Table mass in u 10
TABLE-START-Q Start q, value
TABLE-END-Q End q, value
TABLE-START-AZ Start a, value
TABLE-END-AZ End a, value
TABLE-SCAN-RANGE Range to acquire over
TABLE-MAX-AGC-TIME Max time for AGC
TABLE-AGC-TIME Time used in AGC prescan
TABLE-FREQUENCY Auxiliary frequency
TABLE-AMPLITUDE Auxiliary amplitude


TABLE-PHASE


Auxiliary phase


TABLE-AMPL1 Auxiliary amplitude control


TABLE-TTL-IN


n value


-- --




















Table 2-5.


SAP Table Values for Procedure.


TABLE-SCAN-FLAGS Value of the scan flags
TABLE-START-DAC Start RF DAC value for table
TABLE-DATA-STEPS Number RF DAC acquire steps
TABLE-NO-DATA-STEPS Number RF DAC non acquire steps
TABLE-DAC-STEP-SIZE Size of RF DAC steps
TABLE-DATA-ADDRESS Memory address for acquisition (on SAP)
TABLE-WAIT-LOOPS Padding to equal Table Time
TABLE-USERO Used for multiple functions
TABLE-USER1 Used for multiple functions
TABLE-USER2 Used for multiple functions
TABLE-USER3 Used for multiple functions
TABLE-USER4 Used for multiple functions
TABLE-USER5 Used for multiple functions
TABLE-USER6 Used for multiple functions









return


, exiting the screen,


typing


, the name of the procedure, and the


return key again.


n the following example the code is written between the solid lin


isted by screens.


Comments within the code are denoted by the back slash


, anything following the back slash is ignored when the procedure is


Any text written between the


ines can be entered


nto the software and


the procedure will run as described.


Preceding each screen listing further


explanations of the code are given to explain the purpose of each command.


The comments between each screen listing are not to be entered


nto the


software, they only are included here to help further the understanding of the

code.


The first screen


used to turn on the QITMS.


The voltages and gases


(e.g.,


for CI) are stabilized.


When it is not necessary to turn on a particular


item a back slash before the command wil


disable the function.


Screen


SCAN-ENTER


\ enter


TS-Scan


FORTH


TOGGLE-RF
TOGGLE-MULT
TOGGLE-FIL
TOGGLE-CG


\ turn on RF voltage
\ turn on multiplier
\ turn on filament
\ open cal gas vial


TOGGLE-CI


\open C


gas line


SELECT-CENT-MODE
10000 MSEC-WAIT


\ display centroid data
\let everything stabilize for 10 seconds


PROCEDURE













change the flow of the example program the two values being compared (1


and 2) must be altered.


The commands


as written


only perform the


statements following the PIF without performing the commands following the


PELSE statement.


The second new function is the downloading of


GATORware to the SAP


. The scan function must first be created in


The scan function can then be loaded


nto memory from the PC's


TS-Scan.


hard drive


with the third procedure function.


Screen


\ Example of if-else-then statement


\ Example of


n the procedure


language.


oading the firmware and a scan function from disk.


FORTH 1 2 PROCEDURE


\ put values


to compare on stack


\ compare values


FORTH


LOAD-FIRMWARE


\ if 1


< 2 load gator to SAP


PROCEDURE


PELSE


\ otherwise do


FORTH


"DEMO"
LOAD-SCAN-FILE


PROCEDURE
PTHEN


\ name of scan function file
\read from disk and load to SAP


The third screen


hows how to open an acquisition file.


number, mass range, and acquire type must all be


nitialized.


The file


Three data files


can be opened at one time, numbered 0


The mass range is set by storing


the starting and ending values,


in u 10.


For example,


750.


is equal to 75.0 u.


The type of data (i.e.


profile or centroid) must also be set by putting a true


I I.A.


41 a r nn i nn +k ann r, nnran r -A,- (l ~ ,%l-. anV.-. A .' la -aa aaa &a a .


qeJ


A rin ,,,,,,,,,,r


I


~ n nc~l A CI