7000A Quadrupole GC/MS/MS

• Why GC/MS/MS?

• MS/MS Process and General Benefits

• General Examples of MS/MS Data

• 7000A Hardware description

• 7000A Software description

• Performance Specification

• Performance Data from the 7000A

• Summary

� High Sensitivity

� Fast MRM Speed

� MassHunter Software

� Agilent Reliability

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Why a Quadrupole GC/MS/MS System?

• MS/MS allows for the accurate quantitation of targetcompounds even in high chemical background samples

– MS/MS selectivity means less sample prep

– Sample prep must meet requirements of the GC inlet and column

• Quadrupole MS/MS has better precision and linearity than ion trap MS/MS

• MS/MS provides lower S/N in complex matrices than single quadrupole scan or SIM

• Newer regulations in some markets specify the analytical power of GC/MS/MS

GC/MS Triple Quad (QQQ) for GC/MS/MS

DetectorQuad 1Mass Analysis

Collision Gas (Ar, N2, He)

Quad 2Collision Cell

Ion Source•Ionize Quad 3

Mass Analysis

Carrier Gas (He, H2 )

Mean Free Path

Long LongShort

Collisions No NoYes

MS MS

Multiple Reaction Monitoring (MRM)

170 210 250 290

210

222

268 280165

Quad Mass Filter (Q2)Quad Mass Filter (Q1) Collision Cell

Spectrum with backgroundions (from EI)

Q1 lets onlytarget ion 210 pass through

190 210

210

Collision cell breaks ion 210 apart

150 170 190 210

210158

191

Q2 monitors onlycharacteristic fragments 158 and 191 from ion 210 for quant and qual.

160

158

190

191

no chemical background

MS/MS Eliminates Interferences

analyte

Product 1Product 3

Product 2

Single Quad MSselectivity proportional to

spectral resolutionno selectivity against ions

with same m/z

Triple Quad MSPrecursor selectivity same as MS buthigh probability that at least one of the

product ions will be a unique dissociation product of the precursor only

AND not the interferenceinterference

analyte

interference

unit mass resolution

MS/MS Ensures Lowest Detection Limits

EI: many ions from the source

Q1 SIM isolate precursor before CID

chemical noise eliminated

Product 1Product 3

Product 2

CID +Q2 SIM

Lower m/z Product Ions measured against zero chemical noise

Eliminates the “Invisible” Interferences of SIM

EI-SIMUnit mass resolution

But what happens when a much more intense ion

of a multi-carbon compound has an ion 1 m/z lower?

EI-SIMUnit mass resolution

filters the intense ion that is 1 m/z lower, BUT NOT

the isotope peak from that intensity ion—this

can be a common interference in very ‘dirty’

samples

Isotope ion not removed by SIM

analyte ion

MS/MS eliminates this interference

Removed by SIM

Note: in complex matrices, this isotope interference creates incorrect SIM ratios

and SIM reports with false negatives

103

20 x 103

MS/MS Succeeds Where MS Fails

GC/MS Triple Quad MRM

GC/MS Single Quad SIM

S/N: 116:1 RMS

100 fg HCB

MS SIM

MS/MS MRM A chromatographer’s dream: single peak on flat baseline

Interfering matrix peaks = chemical noise

As Matrix Increases - MS/MS is More Valuable100 fg HCB in Clean Matrix 300 fg HCB in Diesel

Single

MS:

SIM 28

3.8MS

/MS:

283.8

:213.9

S/N=6:1 RMS

S/N=86:1 RMSS/N=37:1 RMS

S/N=26:1 RMS

SIM about equal to MS/MS in clean matrix

MS/MS 15x better than SIM in complex matrix

SIM target

m/z 181

RMS S/N 30 : 1

m/z 219 -> 147

RMS S/N 222 : 1

*

SIM (5973 Single Quad) vs MRM (7000A Triple Quad)a-HCH 3.2 pg injected

Lindanes: MRM Quant Transition 219 147

3.2 pg injected

1.6 pg injected

0.4 pg injected

0.2 pg injected

Page 12

Agilent Innovations in Triple Quad MS

The best technologies from the industry leading 5975C Series GC/MSD

– Heated monolithic gold plated quartz quadrupole

– Proven reliable high performance ion source design

– Low Noise Triple-Axis Detector

…and the 6400 Series LC/MS/MS

– Linear acceleration enhanced hexapole collision cell

– MassHunter software

Inert, high temp source (same as 5975C)

Triple-Axis Detector(as same 5975)

Gold plated quartz, hyperbolic quadrupole(same as 5975C)

Split Flow Turbo

New high performance hexapole collision cell (proprietary design)

High Performance from the Source to the Detector

Inert, high temp source(same as 5975C)

Triple-Axis Detector(as same 5975)

Gold plated quartz, hyperbolic quadrupole

(same as 5975C)

Split Flow Turbo

New high performance hexapole collision cell(proprietary design)

Click on hyperlinked

descriptions for more

details

Hinged access to source and detector

Proven Inert Source Performance

• Several thousands sources in current use

• Solid, inert materials, not a coating that can be damaged by cleaning

• Stay-Clean design extends maintenance intervals with dirty samples

• Source tune parameters in the Autotune file

• Dual filament design reduces maintenance intervals

Partition between source and analyzer compartments

Proprietary Heated Quartz Quadrupole

• High 200C temperature operation results in greater reliability with “dirty” samples

– MS/MS methods typically have more “dirty” samples and benefit even more from the higher temp quad analyzer

• Virtually eliminates the need to clean quads as a maintenance issue

• More stable tunes and methods over a longer period of time in real world sample environments

Analyzer Details

Hinges

Agilent 7000A (QHQ) Design

DetectorQuad 1

Collision Gas (N2 )

Hexapole Collision Cell

Ion Source Quad 2

The hexapole field has excellent transmission effic iency for precursor and product ions

Why a Hexapole: Comparison of Transmission Characteristics

0

0.2

0.4

0.6

0.8

1

1.2

0 500 1000 1500

Transmission

m/z

Mass Range Transmission

Quadrupole Hexapole Octopole

GC/MSm/z=1050

Quadrupoles are the best mass filters (analyzers)Hexapoles and octapole are the best transmission devices

High Performance Collision Cell Design

• Linear acceleration design is optimized for high speed performance

• MRM speed to 500 MRMs/sec allows determination of more compounds per ion group

• High sensitivity with wide mass bandwidth eliminates the need to “tune on your compound” for optimum sensitivity

• Patent pending “Helium Quenching” chemical noise reduction increases S/N

Quadrupole post-filter

Quadrupole pre-filter

Support cradle

(Cell housing and 2 rods removed for clarity)

Why Helium Quenching?Collision Cell Process: Typical Description

Collision Cell

Precursor Ions In

Product Ions Outcollision induced dissociation

1 ml/min N2Collision Gas

Source Detector

Quad Analyzer Quad Analyzer

Why Helium Quenching?Collision Cell Process: Full Description for GC

Collision Cell

Precursor Ions InHe* +

Product Ions Out + He*

In GC/MS, neutral noise is buried in much higher chemical noise.

In GC/MS/MS, chemical noise is greatly reduced so neutral noise is a critical source of noise.

collision induced dissociation

A high population of highly energetic helium metastables are produced in an EI source; since metastable helium is not charged, it can pass through mass analyzer field into the collision cell and through to the HED-EM

1 ml/min N2Collision Gas

At the detector, metastable helium generates neutral noise.

Source Detector

Quad Analyzer Quad Analyzer

Agilent Collision Cell Process with Quench Gas

Collision Cell

Product Ions Out

Precursor Ions InHe* +

collision induced dissociation

1 ml/min N2Collision Gas

Transmission of metastable helium to the detector eliminated = ultra-low neutral noise.

Source Detector

Quad Analyzer Quad Analyzer

He Buffer Gas

He* + He → 2 He + heat

Benefit from “Helium Quench”

Noise Decreased

Helium Quench Gas ON

Helium Quench Gas OFF

tested with hexachlorobenzene

SameSignal

SameSignal

S/N 341:1

S/N 1294:1

High Performance Triple-Axis Detector

• Ultra low neutrals noise

• Long life and high linearity

• “Gain Normalization” corrects tune file for detector aging to allow repeatable long term method sensitivity

• Same design as used in the 5975C GC/MSD

hyperbolic quartztransmission

quadrupoleanalyzer

triple channelelectron multiplier

high energy dynode

steering rod

ion beam

X

Y

OffsetZ

shield for secondary particles

triple channelelectron multiplier

high energy dynode

steering rod

ion beam

XX

YY

OffsetZ

OffsetZ

shield for secondary particles

Technically Superior Design

• Maximum Signal

– Inert MSD source with novel entrance lens

– Highly efficiency, quartz, hyperbolic quadrupole analyzers

– Wide mass bandwidth collision cell

– Increase ion transmission through to the HED-EM

• Minimum Noise– Chemical noise eliminated by MS/MS processes

– Neutral noise eliminated by helium quench gas and Triple-Axis Detector configuration (both unique to Agilent)

Page 26

Smashing Barriers: Agilent 7000A Sensitivity

272:222 Transition180:1 RMS Signal-to-Noise

100fg OFN

Agilent proprietary collision cell innovation led t o this new level of triple quadrupole GC/MS sensitivity

Pesticide Analysis Most Popular Application using GCQQQ

1. Pesticides in Traditional Chinese Medicine (TCM)

2. Pesticides in Tobacco

Page 27

* more pesticides application in “Pesticide Workflo w” talk in the afternoon .

Excellent Result for Qualification in Real Sample - Cyanophos in TCM matrix, NOT solvent

•“ Inert ” Qualifier Ion Ratio0.1ppb(LOD)-1000ppb

Page 28

Excellent Result for Quantitation in Real Sample - Cyanophos in TCM matrix, NOT solvent

R^2=0.99940.2ppb(LOQ)-100ppb(9 Points)

•“ Stable ” Quant AccuracyEven Up to 1000ppb (~104 linear Range)

Page 29

Build up your confidence in Quantitation Results - 1ppb Bromopropylate in TCM matrix, 10 times lower t han Uniform Limit*

RSD = 5.12%

*10 ppb is the uniform limit in Japan Positive List

Page 30

1ppb Cyanophos in TCM Matrix, 6 injection

RSD = 4.83%

Page 31

SIM vs MS/MS Comparison: Chlorpyrifos Not Confirmed in SIM Due to Matrix Interferences!

Chlorpyrifos (28 ppb) Easily Detected and Quantitated by GC/MS/MS – Incurred Carrot

Coun

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MassHunter MS Workstation Software

• Modern software interpretation of the proven industry standard GC/MS Chemstation platform

• Single software platform for all Agilent MS Systems

– LC/SQ , LC/QQQ, LC/TOF, LC/QTOF

– GC/SQ, GC/QQQ

– ICP/MS

Triple Quad GC/MS Software Modules

Acquisition

Analysis

Excel Reporting

Instrument Control UI/Navigation

Autotune

• Proprietary program tunes the source, mass analyzer, and detector for (as applicable):

– Ion transmission

– Mass axis calibration

– Mass resolution

– Detector gain vs Voltage

• Autotune settings are saved with the method for repeatable method performance

• Manual Tune override is available

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Designed for Performance and Reliability

Making femtogram level sensitivity and high speed M RM accessible to a wide range of users

– Leading sensitivity: 100fg of OFN at 100:1 RMS S/N

– High performance MRM with 500 MRMs /sec speed

– New proprietary hexapole collision cell technology

– Reliable, heated gold plated hyperbolic quartz quadrupoles

– Agilent 7890 GC with Capillary Flow technology

– MassHunter Software

Agilent 7000A Triple Quadrupole GC/MSGold Plated Hyperbolic Quartz Quadrupole

Relative Performance of Different MS Modes

• General Summary of Benefits

• Scan versus DRS example

• SIM versus MS/MS example

• Conclusions from ASMS 2008 Poster

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General Statement of the Relative Benefits of Deconvolution and MS/MS

• Amount of improvement depends on nature of the separation and the complexity and intensity of the matrix peaks

– Benefit increases as more coelutions are observed

– Benefit increases as the ratio of the analyte/matrix decreases (intense ions from the matrix peaks bury the analyte ions)

• For MS-DRS, detection limits in matrix are as much as 10x to 50x lower than standard (SIM or scan) methods

• For MS/MS, detection limits in matrix are as much as 100x to 1000x lower than standard (SIM or scan) methods– MS/MS gains in performance are more consistent for more analytes

Report-Spinach Example 1C

All of these compounds were missed due to chemical noise, but AMDIS detected and confirmed all five by NIST05.

Hit number of the top 100 hits from 163,000+ compounds

DRS: Detect the Undetected

DRS A.03 format

Iprodione in Tobacco - SIM could not confirm

Although the plot at m/z 314 might correspond to iprodione, the response at all of the confirmatory ions was over whelmed by the matrix (no distinct peak seen)

Unknown Tobacco Sample – Iprodione 76 ppb

For MS/MS, the quant and qual ions produced cleaned peaks on a simple, flat baseline; ratio of these MRM transitions confirmed iprodione.

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ASMS 2008 Poster Conclusions

Pesticides spiked into neat orange oil (no cleanup)

Poster Title: Bridging the Performance Gap Between GC/MS and GC/MS/MS with Deconvolution TechnologyAuthors: M. Churley, T. Sheehan, P. Zavitsanos, M S zelewski

•MS-DRS lowered LOQ by X10 to X20 for most pesticides but was overwhelmed by intense matrix in some parts of the separation

•MS/MS lowered LOQ by 30x to 100x

•MS/MS consistently detected pesticides in this oil matrix at 20 pg on-column and had sub-pg LOQ for many pesticides