Three years of experience with backflush and retention time locking on

Agilent 7000B GC-MS/MSHans Ragnar Norli

Bioforsk, Plant Health and Plant Protection Pesticide Team

The History of GC-MS Technology at Bioforsk

-Agilent 5973 MS, full scan, Targeted-Agilent 5890 GC with 30 m HP5 MSI kolonne -Pre column 2.5 m.-Gerstel PTV injector air cooled, 20 uL inj. volume-Liner: sintred-RT-lock.-Dataanalysis: ChemStation with -DRS system and AMDIS database; 196 compounds- Injection cycle: 55 min (injection to injection)

-Agilent 7000 B GC/MS Triple Quad -Agilent 7890 GC with 2X 15m HP5 MSUI -Pre column 2.5 m.-MMI inlet, air cooled, 5 uL inj. volume-Liner: 2mm DIMPLED-RT-lock-Back-flush in post run (column midpoint) -Data analysis: Masshunter; 122 compounds-Injection cycle: 30 min

20122005

2010: Introduction of backflush on GC-MS

FLOW

New Column

Column after several injections

Column after backflush

FLOW

The Idea behind Backflush Technology

Quechers of Leek

10.00 15.00 20.00 25.00 30.00 35.00 40.00

1.25e+07

Time-->

AbundanceTIC: 00501007.D\data.ms

10.00 15.00 20.00 25.00 30.00 35.00 40.00

280000TIC: 00401008.D\data.ms

10.00 15.00 20.00 25.00 30.00 35.00

8500000

TIC: 00501001.D\data.ms

10.00 15.00 20.00 25.00 30.00 35.00

280000

TIC: 00401002.D\data.ms

Leek, no back flush

ACN, injected after

Leek, with back flush

ACN, injected after

Control chart examples of Ortho-phenylphenol and Boscalid at spiking level 0.05 mg/kg

Lettuce Apple Orange

Back Flush introduced

Decreasing the Measurement UncertaintyMeasurement Uncertainty (2*CV%)

Pesticide (0.05 mg/kg) Three months before BF installation, N=49

Three months after BF installation, N=49

Change (Before-After)

Orto-phenylphenol 72 38 -34Diphenylamine 56 28 -28Pirimiphos-methyl 64 31 -33Chlorpyriphos 57 22 -35Cyprodinil 48 16 -32Prothiophos 45 20 -25Boscalid 58 23 -35Cypermethrin 79 65 -14Matrices Bean(2)

Apple (8)Lettuce (27)Wheat (8)

Rice (4)

Bean(3)Apple (13)

Lettuce (13)Wheat (5)

Rice (3)Orange (13)

-Number of analytes: 119 -Number of time segments: 19-Largest segment consist of 30 transitions (15 analytes) 3 cycles/s-RT-Lock with ditalimfos at 14.4 min-PCM:10 psi (50 psi in post run)-Column head pressure 16 psi-Backflush time 2.7 min (column hold up time x 5)-Injection cycle: 30 min

Agilent 7000 B GC-MS/MS:

BACK- FLUSH:-After last eluting compound postrun is enabled:

The pressure in the injector is lowered ->0.5 psiPCM pressure and temp in oven and injector are increased-> 50 psi, 280 °C, 280 °C

Establishing a Constant Pressure Method with Backflush in combination with RT-Lock

• For improving peak shape of early eluting peaks; include a pre column• Establish method with constant pressure P:

• Make test runs of a high standard with all pesticides and establish your chromatography with as few time segments as possible!

• Enable backflush in post run and set a pressure at your pressure controller (PCM)

• Make a run and ensure that during the run the flow in Column 2 (Column 1+ 0.04 ml/min) is always higher than Column 1

• Eventually adjust column head pressure or PCM pressure

0 5 10 15 20 250

0.5

1

1.5

2

2.5

Col 1Col 2

Run time (min)

Flow

(mL/

min

Implementing RT-Lock• Put the transitions of your Rt-lock compound into the neighboring time

segments• RT-Lock program: select the pressures at P+2 and P-2• Make runs at P+2 and P-2 and ensure that flow in Column 2 > Column 1 • Eventually adjust the P+2 or P-2 and re-inject• Run the RT-Lock setup program with your RT-Lock compound• Increase the number of time segments to improve the sensitivity

P+1 =P+10%

P

Rt

P

P-1

P-2

P+2P+1

Due to increased injector pressure, the flow in column 1 could be higher than in column 2The PCM module may also not maintain its

pressure

Due to reduced injector pressure, the flow in column 1 could be too low; constant backflush

Maintenance Routines Re-Lock withDitalimfos

Changing liner

Changing pre-column

Changing column 1

Changing column 2

Cleaning ion source

Tuning

Before starting samples

50-60 samples

50-60 samples

After 3-6 months

Once a year After 6-8 months

Every month

There is no need for venting the MS when changing column 1

Changing Columns

UltiMetal FerrulesSilTite

http://www.chem.agilent.com/Library/usermanuals/Public/G3188-90001.pdf

The UltiMetal ferrules are not leaking, simpler to install and cause no problems with snapping off the column when tightening!

ConclusionBackflush:• Protects the ion source• Decrease the measurement uncertainty for several

compounds• No need for venting the MS when changing the first

column #1• Column #2 can last twice as long as column #1• Easy change of columns with UltiMetal Ferrules• Robust system: First installed and tuned it works

without technical problems; «hassle-free»

Conclusion• Don't solve problems with traditional difficult

pesticides like captan, folpet, pyrethroids…• We still have problems with certain matrices:

– Tea -> increased retention times– Extraction at pH 1 for captan, folpet and

chlorothalonil -> reduced sensitivity• You have to invest time and effort to establish

and understand how the system work

Thanks to:

Kari Stuveseth

Anita GlosliApplication notes from Agilent: 5990-4643EN, 5990-3578EN and 5990-9317EN