AB Sciex API 5000 LC/MS/MS Installation instructions

Hardware Guide
API 5000™System
D1000092206 B
May 2010

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Document Number: D1000092206 B 3
Contents
Chapter 1 System Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
Principles of the System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
Applications for the System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
Features of the Instrument. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
How the Instrument Works. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
Parts of the Instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
I/O Panel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
Gas and Vacuum Panel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
Control Panel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Source Exhaust System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Vacuum Chamber . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Shutting Down and Turning on the System . . . . . . . . . . . . . . . . . . . 11
Source/Gas Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Turbo V™ Ion Source Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Compound Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Related Documentation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Technical Support. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Chapter 2 API 5000™Instrument Parameters. . . . . . . . . . . . . . . . . 15

Contents
4 Document Number: D1000092206 B

Document Number: D1000092206 B 5
1
System Overview
The API 5000™ LC/MS/MS system includes a triple quadrupole mass
spectrometer, a Turbo V™ ion source, a computer, and the Analyst®software.
Figure 1-1 API 5000™ system and bench
Principles of the System
This section describes features and applications of the API 5000™ system.
Applications for the System
The API 5000 system is designed for quantitative analysis of small molecules.
This application involves measurement of specific molecular weight compounds,
usually a drug or metabolite in urine or other body fluid, and their resulting
WARNING! If you need to move the system, contact an FSE to assist you. Risk of
personal injury or instrument damage
Note: Before you operate the instrument, make sure you have read the Safety Practices
guide.
Bench
Mass Spectrometer
Line Adjustment
Transformers
Roughing Pumps

System Overview
6 Document Number: D1000092206 B
fragment ions for determining the exact quantity of the compound in the sample
of interest. Quantitation is performed using a standard curve of mass
spectrometer signal intensity for various known concentrations of the compound.
The signal in the test sample is compared to the standard curve to determine the
concentration.
The typical scan type used for this application in the API 5000 system, as well as
all triple quadrupole instruments, is MRM (Multiple Reaction Monitoring).
Features of the Instrument
The API 5000 LC/MS/MS system combines all of the features of the API 4000™
LC/MS/MS system with the additional benefits of improved sensitivity and
enhanced signal-to-noise ratio. It allows comprehensive analysis of
biopharmaceutical compounds and it has the specificity needed for new drug
development. It has the following features:
•A wide diameter intake orifice which improves sensitivity by flowing more
ions into the vacuum chamber.
•A QJet®ion guide that prefocuses the ions before they enter the vacuum
chamber for higher sensitivity and signal-to-noise ratio.
•Improved Q2 lenses reduce CAD gas leakage into the vacuum chamber
and improve sensitivity and signal-to-noise ratio.
•Higher quadrupole operating frequency, which improves ion transmission
and increases sensitivity, especially for low ion masses.
•Higher sensitivity in all RF/DC scan modes.
•Scanning in a mass range of m/z 5–1250 in quadrupole mode.
•Enhanced resolution at reduced scan speeds.
For pharmaceutical and pharmacokinetic samples, MS/MS has the sensitivity and
specificity needed to analyze hundreds of samples per day without requiring
extensive sample preparation.
How the Instrument Works
Mass spectrometry measures the mass-to-charge ratio of ions to identify
unknown compounds, to quantify known compounds, and to provide information
about the structural and chemical properties of molecules. The API 5000 system
has a series of quadrupole filters that transmit ions according to their m/z (mass-
to-charge ratios). The first quadrupole in this series is the QJet ion guide located
between the orifice plate and Q0. The QJet ion guide does not filter ions, but
focuses them before they enter Q0. By prefocusing the larger ion flux created by
the wider orifice, the QJet ion guide increases instrument sensitivity and
improves the signal-to-noise ratio. In Q0 the ions are again focused before
passing into Q1.
Q1 is a filtering quadrupole that sorts the ions before they enter Q2. Q2 is a
collision cell in which ions can be broken into fragments by collisions with gas
molecules. This technique allows users to design experiments that measure the
m/z of product ions to determine the composition of the parent ions. After
passing through Q2 the ions enter Q3 for additional filtering, and then enter the
detector. In the detector, the ions create a current that is converted into a voltage
pulse. The voltage pulses leaving the detector are directly proportional to the
quantity of ions entering the detector. The instrument monitors these voltage

Hardware Guide
Document Number: D1000092206 B 7
pulses and converts the information into a signal. The signal represents the ion
intensity for a particular m/z and the instrument displays this formation as a mass
spectrum. For more information, see Figure 1-2 Ion optics path.
Figure 1-2 Ion optics path
The API 5000 system uses the Turbo V™ ion source to produce ions from liquid
samples. The Turbo V source can use either the TIS (TurboIonSpray®) probe or the
APCI (atmospheric pressure chemical ionization) probe. The instrument is
configured to perform complex MS/MS analysis, but for less rigorous analytical
requirements it can perform single MS scans.
Parts of the Instrument
This section contains diagrams of the instrument and general descriptions of its
components.
Figure 1-3 Front view
Curtain Gas™ flow
Detector
Exit
Interface Region Analyzer Region
Curtain
Plate Orifice IQ1
ST3
Stubbies Stubbies
Deflector
Q0
ST1
Q1 Q2
ST2
IQ3IQ2 Q3
QJet®Ion Guide
Source Exhaust Vacuum Chamber Detector

System Overview
8 Document Number: D1000092206 B
Figure 1-4 Back view
I/O Panel
The input/output panel on the right-hand side of the chassis has a socket for the
external AC power supply, the IEEE-488 (GPIB) connections to the computer, the
AUX I/O to the peripherals, and a sources connector for optional sources.
Figure 1-5 I/O panel
Gas and Vacuum Panel
The vacuum line to the external roughing pumps is connected through the gas
and vacuum panel through the port labeled backing pump. The panel also
houses the gas supply connections and the external connection for the source
exhaust pump.
B BA A
RF FEEDBACK RF FEEDBACK
I/O Panel
Gas and Vacuum Panel
Control
Panel
AUX I/0
IEEE-488
207-242V
50-60 Hz 10A
Sources

Hardware Guide
Document Number: D1000092206 B 9
Figure 1-6 Gas and vacuum panel
Control Panel
The control panel is located on the right side of the instrument. It encloses the
power distribution module.
The control panel contains the main power switch, four fuses, and two
Indicator/Reset buttons. The Vacuum Status button (green) and the Fault button
(red) indicate the vacuum status on the system controller board. When triggered
together, simultaneously pushed and released, the buttons reset the two
microprocessors on the system controller. This has an effect similar to restarting
the computer.
CAUTION! Resetting the instrument using these buttons interrupts any scans and
results in the loss of any unsaved files and data.
When the operational vacuum conditions are satisfied and the instrument is in
analysis mode, the Vacuum Status button is illuminated and the Fault button is
extinguished. The Fault button flashes when a vacuum fault is detected. In Pump-
Down mode, the buttons indicate the status of the Pump-Down sequence.
BACKING PUMP
CURTAIN GAS SUPPLY
MAX 60 PSIG
GAS 1 / GAS 2
MAX 105 PSIG
EXHAUST SUPPLY
MIN 55 PSIG
MAX 60 PSIG
EXHAUST WASTE OUT

System Overview
10 Document Number: D1000092206 B
Figure 1-7 Control panel
Source Exhaust System
The source exhaust system is a safety feature that isolates the sample vapors and
exhaust products from the laboratory environment. The source requires that the
source exhaust system is properly connected and functioning. The exhaust pump
draws the solvent vapors from the enclosed source housing and delivers them to
a trap at the rear of the instrument chassis. It is recommended that these vapors
be passed through this trap and then vented to a fume hood or outside port.
F4
F3
F2
F1
POWER
ON
OFF
VACUUM
STATUS
FAULT RESET
WARNING
(see Note
below)
Spare Output B
Heated Nebulizer (APCI)
Card Cage Blower
Spare Output A
.
I
O

Hardware Guide
Document Number: D1000092206 B 11
Figure 1-8 Source exhaust system
A filtered nitrogen or zero air gas supply (oil free) is delivered to the source
exhaust pump at pressures indicated on the gas panel at the back of the
instrument. The previous figure shows the exhaust supply connection points for
the API 4000™ and API 5000™ systems.
Vacuum Chamber
The vacuum chamber houses the mass filter rail, including the ion optics, the
quadrupole rod sets, the collision cell, and the ion detector.
Shutting Down and Turning on the System
Use the following procedures if you need to shut down or turn on the system.
To shut down the system
1. In the Analyst®software, complete or stop any ongoing scans.
2. Turn off the sample flow to the instrument.
WARNING! Hazardous Materials: Take all necessary precautions to ensure the safe
disposal of the source exhaust gas. Follow all applicable local regulations.
BACKING PUMP
CURTAIN GAS SUPPLY
MAX 60 PSIG
GAS 1 / GAS 2
MAX105 PSIG
EXHAUST SUPPLY
MIN 55 PSIG
MAX 60 PSIG
EXHAUST WASTE OUT
Ion Source
Pressure Switch
Exhaust Supply
Gas Panel
Exhaust Waste Out
Venturi Pump
Front Bulkhead Assembly
B BA A
RFFEEDBACK RFFEEDBACK

System Overview
12 Document Number: D1000092206 B
CAUTION! Potential Instrument Damage: Shut off the sample flow before you
shut down the instrument.
3. In the Analyst software, deactivate the hardware profile and then close the
application software.
4. Stop the Analyst®Service. (See the Software Reference Guide for the
Analyst software.)
CAUTION! Leave the roughing pumps running for a minimum of 15 minutes
after turning off the instrument’s main power switch. When the main power
switch is turned off, the turbo pumps continue to rotate without power for a
few minutes and continue to provide vacuum to the vacuum chamber. If, during
this time, the roughing pumps are turned off, the pressure in the vacuum line
between the roughing pumps and the turbo pumps increases. The increase in
back pressure can create an incorrect load on the turbo pumps’ bearings and
can cause a catastrophic failure of the turbo pumps.
CAUTION! If the instrument is to be shut down for any length of time, we
recommend that the vacuum chamber be vented to prevent exhaust from the
roughing pump being sucked back into the vacuum chamber. To vent the
vacuum chamber, follow steps 5 to 7.
CAUTION! If the vacuum chamber is not going to be vented while the
instrument is shut down, we recommend the roughing pump remain turned on
to prevent the pump exhaust from being sucked back into the vacuum chamber.
If you do not want to vent the vacuum chamber, skip steps 6 and 7.
5. Turn off the main power to the instrument. As you face the instrument
where the instrument name is visible and with the source to your left, the
switch is located on the bulkhead at the back right corner of the chassis.
6. After fifteen minutes, turn off the roughing pumps. The power switch is
located beside the power supply input attachment on the roughing
pump.
7. Let the vacuum chamber vent naturally through the orifice for ten minutes
to allow the instrument to reach atmospheric pressure.
8. Unplug the mains power cord to the instrument from the rear bulkhead
near the main power switch for the instrument.
To turn on the system
Certain conditions outside the direct control of the instrument firmware must be
satisfied before the turbo pumps will be initiated. The Curtain Gas™ supply must
be turned on at the source, and the roughing pump must be turned on manually.
Note: The roughing pump has its own power toggle switch and must be turned
off manually. The roughing pump is not controlled remotely by the system
controller.

Hardware Guide
Document Number: D1000092206 B 13
Interlocks (pressure switches) monitored by the firmware detect if the Curtain Gas
supply and the roughing pump are switched on. If the interlocks are not satisfied,
the turbo pumps are not initiated.
CAUTION! If the ion source is removed, the system electronics will be disabled,
interrupting any data acquisition tasks. The turbo pump and the vacuum system will not
be affected.
1. Turn on the roughing pump, if it was turned off. The power switch is
located beside the power supply input attachment on the roughing
pump.
2. Make sure that all gas supplies are flowing correctly to the instrument.
3. Plug the mains power cord into the bulkhead.
4. Turn on the main power switch.
5. Make sure that the GPIB (general purpose interface bus) cable is
connected to both the instrument and the computer.
6. Turn on the computer, if it was turned off, and then start the Analyst®
software.
Source/Gas Parameters
The parameters in Table 1-1 are optimized for different LC conditions involving
flow rate. For more information about the parameters, see the Analyst®software
Help system.
Note: The roughing pump has its own power toggle switch and must be turned
on manually. The roughing pump is not controlled remotely by the system
controller.
Note: We recommend that you run the instrument with the Curtain Gas™flow set to at
least 20 to maintain good instrument performance.
Table 1-1 Source/Gas Parameters
Parameter Value
Curtain Gas (CUR) 20
IonSpray Voltage (IS) 5000
Temperature (TEM) 700
Ion Source Gas (GS1) 60
Ion Source Gas 2 (GS2) 60

System Overview
14 Document Number: D1000092206 B
Turbo V™ Ion Source Settings
Table 1-2 shows the recommended starting values for the X- and Y-axis
parameters. For more information, see the Turbo V™ Ion Source Operator’s
Guide.
Compound Parameters
In general, you can use the preset values for most of the parameters in Table 1-3.
For more information about the parameters, see the Analyst®software Help
system.
Related Documentation
The guides and tutorials for the instrument and the Analyst software are installed
automatically with the software and are available from the Start menu: All
Programs > AB SCIEX. A complete list of the available documentation can be
found in the online Help. To view the Analyst software Help, press F1.
Technical Support
AB SCIEX and its representatives maintain a staff of fully-trained service and
technical specialists located throughout the world. They can answer questions
about the instrument or any technical issues that may arise. For more information,
visit the web site at http://www.absciex.com.
Table 1-2 Vertical and Horizontal Parameters
Parameter Value
X-axis 5
Y-a xis 5
Table 1-3 Compound Parameters
Parameter Value
CAD Gas Use the preset value and optimize for your
compound.
DP (Declustering Potential) Use the preset value and optimize for your
compound.
EP (Entrance Potential) Use the preset value.
CXP (Collision Cell Exit
Potential)
Use the preset value and optimize for your
compound.
CE (Collision Energy) Use the preset value and optimize for your
compound.
IE1 (Ion Energy 1) — For
experienced instrument
operators only
Do not adjust.
IE3 (Ion Energy 3) — For
experienced instrument
operators only
Do not adjust.

Document Number: D1000092206 B 15
2
API 5000™Instrument Parameters
The following table contains generic parameters for the API 5000™ instrument.
The first number under each scan type is the preset value; the range of numbers
is the accessible range for each parameter.
Table 2-1 API 5000™Instrument Parameters
Parameter
ID Access ID
Positive ion mode Negative ion mode
Q1 Q3 MS/MS Q1 Q3 MS/MS
CUR CUR 10 10 10 10 10 10
10 to 50 10 to 50 10 to 50 10 to 50 10 to 50 10 to 50
CAD CAD 0 1 4 0 1 4
n/a 0 to 12 0 to 10 n/a 0 to 12 0 to 12
IS(1)(2) IS(1)(2) 5500 5500 5500 –4500 –4500 –4500
0 to 5500 0 to 5500 0 to
5500
–4500 to
0
–4500 to
0
–4500 to
0
NC(3) NC(3) 333–3–3–3
0 to 5 0 to 5 0 to 5 –5 to 0 –5 to 0 –5 to 0
TEM(2)(3) TEM(2)(3) 000000
0 to 750 0 to 750 0 to 750 0 to 750 0 to 750 0 to 750
OR DP 120 120 120 –100 –100 –100
(DP = OR) 0 to 400 0 to 400 0 to 400 –400 to
0
–400 to
0
–400 to 0
Q0 EP 10 10 10 –10 –10 –10
(EP = –Q0) 15 to 2 15 to 2 15 to 2 –15 to
–2
–15 to
–2
–15 to –2
IQ1 IQ1 Q0 + (–1) Q0 + (–1) Q0 +
(–1)
Q0 + 1 Q0 + 1 Q0+1
(IQ1 = Q0 + offset) –0.5 to –2 –0.5 to –2 –0.5 to
–2
0.5 to 2 0.5 to 2 0.5 to 2
ST ST Q0 + (–7) Q0 + (–7) Q0 +
(–7)
Q0 + 7 Q0 + 7 Q0 + 7
(ST = Q0 + offset) –8 to –4 –8 to –4 –8 to –4 4 to 8 4 to 8 4 to 8
RO1 IE1 1 n/a 1 –1 n/a –1
(IE1 = Q0 – RO1) 0.5 to 2 0.5 to 2 –2 to
–0.5
–2 to
–0.5
RO1 RO1 n/a Q0 + (–2) n/a n/a Q0 + 2 n/a
(IE1 = Q0 + offset) –0.5 to –2 0.5 to 2
(1)IonSpray™ ion source (2)TurboIonSpray®ion source (3)Heated Nebulizer (4) 1=ON
and 0=OFF (5)DuoSpray™ ion source, 1=TIS, and 2=HN

API 5000™Instrument Parameters
16 Document Number: D1000092206 B
IQ2 IQ2 Q0+
(–20)
Q0+ (–20) Q0+
(–20)
Q0 + 20 Q0 + 20 Q0 + 20
(IQ2 = Q0 + offset) –100 to
–8
n/a n/a 100 to 8 n/a n/a
RO2 RO2 –100 –20 n/a 100 20 n/a
–200 to
200
–145 to
–2
–200 to
200
2 to 145
RO2 CE n/a n/a 30 n/a n/a –30
(CE = Q0 – RO2) 5 to 130 –130 to
–5
ST3 ST3 –120 n/a n/a 150 n/a n/a
–200 to
200
–200 to
200
ST3 CXP n/a 20 15 n/a –20 –15
(CXP = RO2 – ST3) 0 to 55 0 to 55 –55 to 0 –55 to 0
RO3 RO3 –150 n/a n/a 100 n/a n/a
–200 to
200
–200 to
200
RO3 IE3 n/a 2 2 n/a –1.5 –1.5
(IE3 = RO2 – RO3) –0.5 to 5 –0.5 to 5 –5 to 0 –5 to 0
DF DF –200 –200 –200 200 200 200
–400 to 0 –400 to 0 –400 to
0
0 to 400 0 to 400 0 to 400
CEM CEM 2000 2000 2000 2000 2000 2000
500 to
3297
500 to
3297
500 to
3297
500 to
3297
500 to
3297
500 to
3297
GS1 GS1 20 20 20 15 15 20
0 to 90 0 to 90 0 to 90 0 to 90 0 to 90 0 to 90
GS2 GS2 0 0 0 0 0 0
0 to 90 0 to 90 0 to 90 0 to 90 0 to 90 0 to 90
ihe(4) ihe 1 1 1 1 1 1
0 or 1 0 or 1 0 or 1 0 or 1 0 or 1 0 or 1
IHT IHT 40 40 40 40 40 40
10 to 250 10 to 250 10 to
250
10 to
250
10 to
250
10 to 250
svp(5) svp 1 1 1 1 1 1
1 or 2 1 or 2 1 or 2 1 or 2 1 or 2 1 or 2
Table 2-1 API 5000™Instrument Parameters (cont’d)
Parameter
ID Access ID
Positive ion mode Negative ion mode
Q1 Q3 MS/MS Q1 Q3 MS/MS
(1)IonSpray™ ion source (2)TurboIonSpray®ion source (3)Heated Nebulizer (4) 1=ON
and 0=OFF (5)DuoSpray™ ion source, 1=TIS, and 2=HN