Conversion of Agilent EI GC/MSD Systems To Hydrogen Carrier Gas Agilent Technologies 1
Introduction: Converting From He To H 2 Many GC/MS users are considering changing from helium to hydrogen carrier gas due to price/availability problems with helium. This talk describes the steps recommended for converting EI GC/MS methods. It is important to recognize the differences with using hydrogen carrier. Time should be allotted for adapting the method, optimization, and resolving potential problems. Areas that will need attention include: choice of supply of H 2 Remove/bypass carrier gas filter (may be re-installed later if needed) GC/MSD hardware changes choosing new chromatographic conditions a 2-5 fold reduction in signal-to-noise ratio changes in spectra and abundance ratios for some compounds activity and reactivity with some analytes Page 2
Introduction: Converting From He To H 2 Methods that will generally require less optimization include analytes that: are durable compounds are at higher concentrations are analyzed with split injections are derivatized Methods that will generally require more optimization include analytes that: are fragile compounds are at trace concentrations Allow time for necessary updates to SOPs and validation. Page 3
4 H 2 Safety
Letter From Agilent Safety Engineer. Describes H 2 Safety Features of Agilent GCs 5
Designed for Reliability H 2 Safety Safety Shutdown When gas pressure set points are not met, the valve and heater are shut off to prevent explosion Flow Limiting Frit If valve fails in open position, inlet frit limits the flow Oven ON/OFF Sequence Fan purges the oven before turning on heater to remove any collected H 2 Explosion Test GC designed to contain parts in case of explosion Page 6
Safety First: Read This Before You Start G3170-90010 Page 7
Source of H 2 Carrier and Plumbing 8
Source of Hydrogen: Use A Hydrogen Generator Higher initial expense than cylinders, much lower cost over time Very clean H 2, >99.9999% available More consistant purity Safety considerations H 2 is only generated at needed pressure (like 40 psig) Flow is limited (like 250 ml/min) Auto-shutdown if setpoint pressure cannot be maintained Minimal stored gas (like 50 ml at 40 psig) of H 2 at any one time Make sure to buy a good one with a low spec for water and oxygen 9
10 This is an example of the type of H 2 generator to use. Look for generators with the > 99.9999% specification and low individual specs on water and oxygen.
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Plumbing The Instrument Chromatographic quality stainless steel tubing is often recommended for H 2 plumbing and is probably the best choice if available. Users may have to follow local codes or internal company guidelines. We have also used new 1/8 th copper that has been cleaned for GC use. Dirty tubing will cause huge contamination problems, as H 2 appears to carry dirt out of metal more than He does. Don t use really old copper tubing, as it becomes brittle and can break. Note that MSD leak checks will not always find big outgoing leaks. Leak check when complete with electronic leak detector. When plumbing a H 2 generator, start out with no traps. Only add traps if needed. Make sure the water and oxygen levels are low enough. 12 Confidentiality Label September 6, 2012
Split Vent and Septum Purge Vent Should Be Connected to Exhaust 13
MS Components Needed Before Conversion: Magnet and Drawout Lens 14
Check The Magnet In The 5975 If the magnet in the 5975 does NOT have a serial number along the left edge (as viewed through the source window), it is suitable for use with Helium, but not Hydrogen. You should contact your Agilent service engineer to change it to a Helium and Hydrogen compatible magnet before converting the system to H 2 carrier. Note: all 5973 magnets are compatible with both He and H2 15
Replace The Standard 3 mm Drawout Lens With The Optional 6 mm When Using H 2 Carrier G2589-20045 6 mm Drawout Plate, Inert Standard 3 mm Drawout Plate, Inert 16 Confidentiality Label September 6, 2012
Choosing A Column And Conditions 17 Confidentiality Label September 6, 2012
MSD Pumping Capacity For H 2 Is Less Than For He This limits column choice Complete info on H 2 pumping specs are not yet available These are approximate values for the 5975. Max flow of H 2 to maintain reasonable source pressure: Performance turbo: 2 ml/min Standard turbo: 1 ml/min Diffusion pump: 0.75 ml/min Pressure pulsing: turbos <= 3 ml/min, diffusion <= 2.5 ml/min It is very helpful to have an ion gauge on the MS to monitor the vacuum vs column flow. Try to avoid flows that produce pressures higher than 5 x 10-5 torr. You can get useful data above this pressure, but performance starts to degrade rapidly 18
Guidelines For Choosing Column Dimensions And Flows For Method Converted From He To H 2 Determine max flow of H 2 into MS that will give source pressure of 5 x 10-5 torr or less source pressure. This is your max column flow. Choose column dimensions at initial oven temp of method to give: A flow <= max column flow for vacuum pump A flow >= min column flow for efficiency An inlet pressure of at least 5 psig Keeping a temperature ramp of the same number of C/void time will give similar elution order. Use the method translator for this. These are only approximate guidelines. Sometimes you have to deviate from them 19
Van Deemter Curves If the flow of He or H 2 is too low, you loose efficiency MUCH FASTER than if it is too high. This is why operating at or above minimum linear velocity is important. Use 35 cm/sec as minimum for H 2 20
Column Choice When in doubt (or a hurry), use: Use 20m x 180 um column and set constant flow at 0.3-0.5 ml/min. Try to get a linear velocity of at least 35 cm/sec Try to use the same phase and phase ratio to get similar elution order. Smaller bore columns have lower capacity. You may have overloading on high level cals or samples You may need to use pulsed injection to load sample into small bore column with low flow rate. The flow range for your H 2 setup is limited on the low side by the flow to get 35 cm/sec and on the high side by the vacuum pump capacity 21
Using The Flow Calculator. Set Linear Velocity to 35 For H 2 Carrier. If InletPressure Too Low, Raise Flow Inlet pressure is too low Set inlet pressure to 5 psig 22
Example: 30m x 0.25 mm id x 0.25 um Checkout Column With Perf Turbo Meets flow requirements for pump and column efficiency with 2 x speed gain 23
Example: 20m x 0.18 mm id x 0.18 um Column With Perf Turbo and Std Turbo Meets flow requirements for std turbo pump. Flow here is chosen to give a 2.5 x speed gain. 24
Status Of Calculators For Column Selection HP FlowCalc 2.0 and HP Method Translator used in earlier slides DO NOT WORK on Win 7 Newer versions of these can also be found in: GC/GCMS Instrument Utilities Kit. This CD ships with every Agilent GC, GC/MS, and autosampler. It also has manuals for GC and a firmware updating tool. 25
Initial Startup With Hydrogen 26
Initial Problems With Switching GC/MS Methods to H 2 High background that looks like hydrocarbons Reduced signal to noise (worse MDL) Significant tailing for many compounds Page 27
Switch to H 2. Early problem- High Background You will see this throughout work with H 2. It will get much smaller with time and use. It used to takes days or weeks for this to fall to acceptable levels. Agilent has a new conditioning protocol to clean up the background much faster (see later slide). Page 28
Tox Checkout Sample Abundance Time--> 2000000 1800000 1600000 1400000 1200000 1000000 800000 600000 400000 200000 0 Immediately after conversion to H 2, peakshapes on TIC are poor and tailing badly. 20m x 0.18mm id x 0.18 um DB-5MSUI 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 12.00 13.00 Hydrogen Abundance 3000000 2800000 2600000 2400000 2200000 2000000 1800000 1600000 1400000 1200000 1000000 800000 600000 400000 200000 0 Time--> 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 Helium Page 29
GC/MS Toxicology Checkout Mixture 5190-0471 1 Amphetamine 2 Phentermine 3 Methamphetamine 4 Nicotine 5 Methylenedioxyamphetamine(MDA) 6 Methylenedioxymethamphetamine(MDMA) 7 Methylenedioxyethylamphetamine 8 Meperidine 9 Phencyclidine 10 Methadone 11 Cocaine 12 SKF-525a (RTL Compound) 13 Oxazepam 9 14 Codeine 5 ng/ul each compound 17 15 Lorazepam 16 Diazepam 17 Hydrocodone 18 Tetrahydrocannabinol 19 Oxycodone 20 Temazepam 21 Flunitrazepam 22 Diacetylmorphine 23 Nitrazepam 24 Clonazepam 25 Alprazolam 26 Verapamil 27 Strychnine 28 Trazodone 1 2 3 4 6 7 8 10 12 11 14 16 15 18 21 20 19 22 23 24 25 26 27 28 5 13 2 4 6 8 10 12 14 16 18 30
Tailing Peaks: There is More Going On Here Than Simple Tailing Cocaine Abundance 7.20 7.40 7.60 7.80 8.00 8.20 8.40 8.60 8.80 9.00 Peak Spectrum Abundance Tail Spectrum 24000 Average of 8.018 to 8.036 min.: ToxCKO_H2_4.D\data.ms Average of 8.048 to 8.085 min.: ToxCKO_H2_4.D\data.ms 82 91 22000 182 14000 20000 18000 12000 16000 10000 14000 Cocaine 12000 8000 Not Cocaine 42 10000 6000 78 105 8000 41 6000 4000 64 55 4000 68 2000 115 303 128 167 2000 122 152 167 198 272 141 154 191 207221 135 242 215 265 281 300 0 230244 259 288 327341 0 314327341 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340 m/z--> m/z--> Page 31
Main Ions Of Cocaine Do Not Tail. Tail Made Up of Other Ions Tail is different than peak. May be formed from catalytically shredded main component. This problem is reduced with time Cocaine Tail is not cocaine ions Page 32
Minimizing The Time For Chromatography and Background To Stabilize Have a good source of clean hydrogen like a >99.9999% purity hydrogen generator. Keep H 2 flow limited to value suitable for your pumping system Use 6 mm drawout lens After setup, purging and pumpdown: Set the source to max temp for your source (check to see what your s is) Reduce the EMV to <= 800V Leave the FILAMENT ON overnight. This cleans up background rapidly. Peakshape will be much better and background will be much lower in the morning. Lower the source temp to method value, retune, and run some samples It is a good idea to have an extra set of filaments on hand in case one burns out. This hasn t been a problem, but a good idea. Page 33
H 2 Method With 6 mm Drawout And Overnite Cleaned Source. Peakshape Much Better and Background is Lower. Abundance 2600000 2400000 2200000 2000000 1800000 1600000 1400000 1200000 1000000 800000 600000 400000 200000 Time--> 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 12.00 13.00 14.00 Page 34
Performance Expectations Signal-to-noise ratio is usually worse by about 2-5 x. This obviously varies from compound to compound While most spectra remain same, there are always exceptions. Users should check the reference spectra for important targets to make sure they have not changed Same comment for target/qualifier ratios Some compounds may disappear at low levels. Examples include: some nitro and oxygen containing compounds (alcohols, aldehydes, ketones) like those found in flavor samples 35
Other Considerations: H 2 is not an inert gas, and therefore inertness problems will still exist or be worse. Some things to consider: Use the lowest inlet temp that works (to reduce reactions with H 2 ) Use pulsed injection, especially with small bore columns Consider using an MMI in cold splitless mode for fragile compounds Using a deactivated S/SL weldments might help (but don t ever scrub it) PN:G3452-60570 (bottom) PN:G3452-60586 (top) Avoid using methylene chloride as a solvent (especially wet). At higher inlet temps (like 300C), HCl is formed and causes problems. It can remove the gold from the seal and corrode the weldment. If DCM must be used, use lowest inlet temp and maybe a deactivated S/SL weldment (or MMI) Also avoid carbon dissulfide as a solvent Use liners with a taper at bottom to minimize sample contact with gold seal Use Agilent Ultra Inert Liners. 36
Hydrogen Carrier Summary 1. Get a GOOD hydrogen generator and plumb it properly 2. Pick a column that meets the flow needs of method and MSD pump 3. Do magnet update (if necessary) and install 6 mm drawout lens 4. After purging and pumpdown, set source to max temp and quad to 200C. Leave filaments on all night. 5. Reset source temp, retune, and run samples. Run practice samples for a few days to let system stabilize