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Agilent Technologies — Technical Presentation (August 2019)
54 slides from Agilent Technologies's technical presentation, Aug 22, 2019
Slide 1: Water Injection: An Expanding Problem
Water Injection: An Expanding Problem Mark Sinnott Application Engineer August 2019 1 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 2: Water Injections in GC
Water Injections in GC How wet can you get? Page 2 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 3: Agenda
Agenda ➢ Potential problems with water injections ➢ Column bleed explanation and misconceptions ➢ Experimental study on the effects of water injections on various columns ➢ BackFlash August 22, 2019Month Page 3 ##, 200X
Slide 4: Why Inject Water?
Why Inject Water? Convenient • Aqueous samples (wastewater, drinking water, and so on) • Biological samples Sometimes Necessary Page 4 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 5: Potential Problems with Water
Potential Problems with Water Injector issues • Large expansion volume • Backflash Detector issues • Extinguish FID flame • Decrease sensitivity of ECD Page 5 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 6: Potential Problems with Water: Real and Perceived
Potential Problems with Water: Real and Perceived Solvent – Stationary phase mismatch • Poor wettability of many stationary phases by water • “Puddles” Damage to stationary phase • Change in retention times • Change in selectivity • Increase in bleed Page 6 August 22, 2019 Water Injection: An Expand
Slide 7: Water Injection: Using Nonpolar Column
Water Injection: Using Nonpolar Column 3 4 2? 1? Page 7 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 8: Water Injection: Using Nonpolar Column
Water Injection: Using Nonpolar Column 3 2 4 1 5 Page 8 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 9: Water Injection: Using Nonpolar Column
Water Injection: Using Nonpolar Column 1. ethanol (78 ºc) 2. isopropanol (82 ºc) Intuvo 9000 3 3. 1-butanol (118 ºc) Inlet: 250 °C 2 4. 1-octanol (194 ºc) FID: 275 °C 5. 1-decanol (233 ºc) Split: 25:1 Splitless: Purge flow: 50 mL/min Purge time: 0.5 min 4 Velocity: 30 cm/s 1 T prgm: 40 °C for 2 min
Slide 10: Splitless Injection
Splitless Injection Initial column temperature Hexane solvent (BP = 68-69 °C) Solvent effect Cold trapping 70 °C 50 °C 2 3 4 3 4 2 1 1 2 4 6 8 2 4 6 8 Time (min) Time (min) DB-1, 15 m x 0.25 mm id, 0.25 µm 50 °C or 70 °C for 0.5 min, to 210 °C at 20 °C/min; helium at 30 cm/s 1. n-decane 2. n-dodecan
Slide 11: Splitless Injection
Splitless Injection Polarity mismatch 4 Hexane 5 6 Methanol 1 4 2 5 6 2 3 3 1 0 1 2 3 4 5 6 1 2 3 4 5 6 Time (min) Time (min) DB-1, 15 m x 0.25 mm id, 0.25 µm 50 °C for 1 min, 50-210 °C at 20 °C/min; helium at 30 cm/s 1. 1,3-DCP 2. 3-hexanol 3. butyl acetate 4. 1-heptanol 5. 3-octanone 6. 1,2- dichl
Slide 12: Splitless Injection – Things You Can Do
Splitless Injection – Things You Can Do Polarity mismatch You could: • Change polarity of the solvent • Change the polarity of the stationary phase • Use a retention gap Page 12 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 13: Retention Gap
Retention Gap Also called a guard column Injector Detector Deactivated fused silica tubing Column Union Usually 2-10 m long and same diameter as the column (or larger if needed) Page 13 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 14: Splitless Injection
Splitless Injection 3 m x 0.25 mm id retention gap Methanol Methanol 4 No retention gap With retention gap 4 5 6 5 6 2 2 1 3 3 1 1 2 3 4 5 6 1 2 3 4 5 6 Time (min) Time (min) DB-1, 15 m x 0.25 mm id, 0.25 µm 50 °C for 1 min, 50-210 °C at 20 °C/min; helium at 30 cm/s 1. 1,3-DCP 2. 3-hexanol 3. butyl
Slide 15: What is Normal Column Bleed?
What is Normal Column Bleed? Normal background signal generated by the elution of normal degradation products of the column stationary phase Page 15 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 16: What is Bleed?
What is Bleed? The thermodynamic equilibrium process occurs to some degree in all columns. Polysiloxane backbone releases low molecular weight, cyclic fragments. It occurs at low levels in low temperature, O2-free, clean system. It is increased at higher temperatures, oxygen exposure, or chemical at
Slide 17: Bleed: Why Does It Happen?
Bleed: Why Does It Happen? “Back biting” mechanism of product formation H3C CH3 Si HO O CH3 CH3 CH3 CH3 CH3 Si O Si O Si O Si O Si Si CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH O 3 CH3 CH3 Si O Si O Si O Si O Si O Si O Si OH CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 CH3 Si O Si O Si O Si OH H3C CH
Slide 18: Low Bleed Phase Structures
Low Bleed Phase Structures CH 3 DB-5 structure DB-5ms structure O Si CH 3 CH 3 O CH 3 Si CH O 3 Si Si CH 3 CH 3 O Si CH 3 Si CH 3 CH 3 O CH 3 O Si CH 3 Si CH 3 CH 3 DB-5, HP-5 and HP-5ms DB-5ms 5% phenyl 1. Increased stability 2. Different selectivity 3. Optimized to match DB-5 Page 18 August 22, 20
Slide 19: Benefits of Low Bleed Phases
Benefits of Low Bleed Phases PAH sensitivity using DB-35ms 1. Naphthalene 2. Acenaphthylene 3. Acenaphthene Commercially Benzo[ghi]perylene 4. Fluorene available 35% phenyl column S/N = 15 5. Phenanthrene 6. Anthracene 7. Fluoranthene 8. Pyrene 9. Benz[a]anthracene 1 10. Chrysene 11. Benzo[b]fluoran
Slide 20: Low Bleed Stationary Phases
Low Bleed Stationary Phases DB-35ms versus DB-35 DB-35ms S/N = 10 * 320°C 8.00 10.00 12.00 14.00 16.00 18.00 20.00 S/N = 3 DB-35 300°C * * 5 ng decachlorobiphenyl 8.00 10.00 12.00 14.00 16.00 18.00 20.00 CLP pesticides analysis Page 20 August 22, 2019 Water Injection: An Expanding Problem Agilent Re
Slide 21: Influences on Column Bleed
Influences on Column Bleed 1.3e4 Phase type 1.2e4 Temperature 1.1e4 Column dimensions 1.0e4 9000 DB-624 30M x .53mm I.D., 3.0µm 24 pA / 260°C 8000 7000 DB-1 30m x .32mm I.D., .25µm 12 pA / 320°C 6000 0 5 10 15 20 25 Time (min.) Page 22 August 22, 2019 Water Injection: An Expanding Problem Agilent Re
Slide 22: What is a Bleed Problem?
What is a Bleed Problem? An abnormal elevated baseline at high temperature It is not: • A high baseline at low temperature • Wandering or drifting baseline at any temperature • Discrete peaks Page 23 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 23: Septum maintenance: Deconvoluted inlet septa spectrum
Septum maintenance: Deconvoluted inlet septa spectrum August 22, 2019Month Page 24 ##, 200X
Slide 24: Septum maintenance: Deconvoluted inlet septa spectrum
Septum maintenance: Deconvoluted inlet septa spectrum 10 12 14 16 18 20 Decamethyl Dodecamethyl Tetradecamethyl Hexadecamethyl Octadecamethyl Eicosamethyl cyclopentasiloxane cyclohexasiloxane cycloheptasiloxane cyclooctasiloxane cyclononasiloxane cyclodecasiloxane August 22, 2019Month Page 25 ##, 20
Slide 25: Goals of Study
Goals of Study Who cares? • Frequent inquiries • Water can cause problems • Quantitative data • Establish “guidelines” Page 26 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 26: Stationary Phases
Stationary Phases • Dimethylpolysiloxane (DB-1) • Polyethylene glycol (DB-WAX) • Cyanopropylphenyl (DB-225) • Cyclodextrin (CycloSil B) • Divinylbenzene/ethylene glycol dimethacrylate (HP-PLOT U) Page 27 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 27: Experimental Conditions
Experimental Conditions Instrument: Agilent GC with auto injector Injector: 250 °C, 1:5 split Injection volume: 1 µL Detector: FID, 300°C Carrier gas: H2 at 40 cm/s Page 28 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 28: Experimental Conditions
Experimental Conditions Oven: 130 °C, 60 °C, or 200 °C* isothermal Column dimensions: 30 m x 0.53 mm id x 1.0 µm 1,000 water injections at each temperature on each column Bleed profile after 250, 500, and 1,000 water injections Test mix after 250, 500, and 1,000 water injections *for Cyclosil B and
Slide 29: Compound Lists
Compound Lists DB-1 DB-WAX DB-225 CycloSil B HP-PLOT U 2-chlorophenol 2-octanone tetradecane dodecane methanol undecane tetradecane 2-chlorophenol (R)-linalool ethanol 2,4-dimethylaniline 1-octanol Hexadecane (S)-linalool pentane 1-undecanol methyl deconate naphthalene naphthalene ethyl ether tetrad
Slide 30: Test Mixes
Test Mixes Compound list for chromatograms HP-PLOT U Peak Compound 1 methanol 2 ethanol 3 pentane 4 ethyl ether 5 acetone Used for RI 6 hexane 7 ethyl acetate Used for k Page 31 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 31: Results DB-1
Results DB-1 Parameter Before Injection After Injection Ret. Factor (k) 14.6 14.5 Ret. Index 1 1349.88 1350.02 Ret. Index 2 1427.77 1428.16 Theor. plates 1448 1474 Bleed (pA) 12.8 11.2 Page 32 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 32: Results DB-WAX
Results DB-WAX Parameter Before Injection After Injection Ret. Factor (k) 12.6 12.6 Ret. Index 1 1149.54 1149.73 Ret. Index 2 1163.44 1163.71 Theor. plates 1277 1261 Bleed (pA) 44.8 32.1 Page 33 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 33: DB-WAX Before and After 2,000 Water Injections
DB-WAX Before and After 2,000 Water Injections Page 34 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 34: Results DB-225
Results DB-225 Parameter Before Injection After Injection Ret. Factor (k) 11.5 11.4 Ret. Index 1 1622.30 1621.26 Ret. Index 2 1711.51 1711.03 Theor. plates 1101 1110 Bleed (pA) 34.5 39.3 Page 35 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 35: Results CycloSil B
Results CycloSil B Parameter Before Injection After Injection Ret. Factor (k) 7.8 7.6 Phase Ret. Index 1306.3 1306.0 wash out Resolution 1.9 1.3 Theor. plates 2631 2025 Bleed (pA) 28.4 15.1 Page 36 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 36: Results HP-PLOT U
Results HP-PLOT U Parameter Before Injection After Injection Ret. Factor (k) 5.2 5.3 Ret. Index 538.0 540.0 Theor. plates 950 982 Bleed (pA) 74.2 35.6 Page 37 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 37: Asymmetry (Skew)
Asymmetry (Skew) Skew numbers are based on a modified Gaussian-peak model*. The numbers represent the deviation from an ideal Gaussian, that is, perfectly symmetrical, peak. The smaller the number, the better. Noticeable tailing starts at ~0.8 Obvious tailing start at ~1.2 *W.W. Yau, Anal. Chem., vo
Slide 38: Results DB-1
Results DB-1 Asymmetry Compound Before Injection After Injection Chlorophenol 0.48 0.47 Dimethyl aniline 0.47 0.46 Undecanol 0.51 0.46 Page 39 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 39: Results DB-WAX
Results DB-WAX Asymmetry Compound Before Injection After Injection Decanol 0.38 0.47 Dimethyl aniline 0.30 0.32 Dimethyl phenol 0.25 0.27 Page 40 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 40: Results DB-225
Results DB-225 Asymmetry Compound Before Injection After Injection Chlorophenol 0.46 0.47 Dimethylaniline 0.42 0.43 Undecanol 0.63 0.58 Page 41 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 41: Study Summary
Study Summary Bonded phases, no change in: For water injections on all columns with bonded stationary Polarity Selectivity isRetention observed. Efficiency For nonbonded phases, like CycloSil B, water injections can Activity wash out part of the nonbonded stationary phase – loss of resolution, reten
Slide 42: Study Conclusions
Study Conclusions For binded and cross-linked columns it is safe to inject water Nonbonded columns: Water injections can wash out stationary phase – use with caution Page 43 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 43: Water Injection is Acceptable
Water Injection is Acceptable If it is acceptable to inject water, why am I having all these problems when I inject it? Problems associated with water injections are often caused by injector-related phenomena: backflash Page 44 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 44: Typical Solvent Expansion Volumes
Typical Solvent Expansion Volumes Solvent Vapor volume (µL) of 1 µL liquid Water 1010 Methanol 450 Carbon disulfide 300 Methylene chloride 285 Acetone 245 n-Hexane 140 Page 45 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 45: Backflash
Backflash Cause • Vaporized sample expands 100 to 1000 times • Portions may leave the liner • Occurs when vapor volume > liner volume Page 46 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 46: Backflash
Backflash Sample expands to overfill injector Some sample condenses on cooler areas (bottom of septum, metal body, and so on) Page 47 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 47: Temperature Profile of a Typical Vaporization Injector vs Oven Temperature
Temperature Profile of a Typical Vaporization Injector vs Oven Temperature Bottom Injection port of Septum 10 Setpoint temperature 20 350 °C 30 Syringe 40 tip 50 60 70 80 90 Base of injection 35 °C 150 °C 300 °C port oven oven oven 50 100 150 200 250 300 350 400 Temperature in gas stream (°C) Page 4
Slide 48: Backflash
Backflash Some sample flows out of injector Lower volatility portions can condense on cooler areas Page 49 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 49: Backflash
Backflash As carrier gas flows over the Subsequent injections can condensed portion, the also dislodge condensed headspace is brought sample, as well as deposit forward more sample Sample then enters the column Page 50 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 50: Backflash
Backflash Problems • Loss of sample • Baseline interferences • “Ghost” peaks • Tailing solvent front or major component Page 51 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
Slide 51: Backflash
Backflash Minimizing • Large volume liner (possibly tapered) • Small injection volume • Low expansion solvent • Low injector temperature • High carrier gas flow rates • High head pressures (possibly pulsed) • Smaller column diameters Use vapor volume calculator! https://www.agilent.com/en/support/ga
Slide 52: Summary
Summary ➢ Water injection can be tricky ➢ Polarity miss-match ➢ For bonded columns and many PLOT columns, no negative effects of injecting water were observed ➢ Water is not recommended for Non-bonded columns and some PLOT columns, (e.g., Alumina and Molesieve, are not suitable for water injections)
Slide 53: Contact Agilent Chemistries and Supplies Technical Support
Contact Agilent Chemistries and Supplies Technical Support 1-800-227-9770 Option 3, Option 3: Option 1 for GC or GC/MS columns and supplies Option 2 for LC or LC/MS columns and supplies Option 3 for sample preparation, filtration, and QuEChERS Option 4 for spectroscopy supplies Available in the USA,
Slide 54: Questions?
Thank you! Questions? Page 55 August 22, 2019 Water Injection: An Expanding Problem Agilent Restricted
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