New Wool Ensures More Accurate Semivolatiles Analyses

By Scott Grossman, Innovations Chemist

• Lower detection limits for 2,4-dinitrophenol with new wool for semivolatiles.
• Improve data accuracy with complete sample transfer.
• Extend column lifetime by reducing matrix contamination.

Ensuring success for semivolatiles analysis is dependent on several issues. The column is an important factor; however, even with a high performance column, other parameters can significantly impact final results. Liner choice is one of the most important noncolumn factors that affects end data and to take full advantage of the column's performance the analytes need to reach the column without degradation or discrimination. How much of the sample is transferred, how reproducible the transfer is, and how representative the data will be of the original sample are all influenced by injection port factors. In this article, we will demonstrate how liners with Semivolatiles Wool—a new wool, specifically designed for semivolatiles analysis—can be used to optimize the injection port for more complete sample transfer, increased accuracy, and lower detection limits.

For semivolatiles analysis, many analysts operate their inlets in the splitless mode using a single gooseneck liner to protect the vaporized sample cloud and prevent analyte breakdown during the relatively long dwell time in the hot injection port. Wool can be used in the liner to promote sample vaporization, minimize molecular weight discrimination, improve chromatography, and extend column lifetime by preventing matrix contaminants from entering the column. However, wool must be chosen with care as some types may have active sites, which can interact with target analytes and negatively affect both peak shape and response. For example, some active phenolic compounds, such as 2,4-dinitrophenol (DNP), can be completely adsorbed and not detected at all, depending on the wool that is used.

Restek's new Semivolatiles Wool was developed specifically for semivolatiles analysis. Liners with Semivolatiles Wool were compared to MS Certified Liners packed with wool, and the Semivolatiles Wool liners were found to have greater response for 2,4-dinitrophenol, allowing it to be detected at much lower levels (Figure 1). Semivolatiles Wool liners also demonstrated greater inertness through consistent results in initial analyses, i.e., very little priming was observed (Table I). While inertness was initially tested using a flame ionization detector (FID), results for mass spectrometry (MS) were also excellent (Figure 2). More than twice the required response factor for 2,4-dinitrophenol was routinely achieved when analyzing just 1ng on-column by MS (Figure 3).

An inert sample pathway allows complete sample transfer and using Semivolatiles Wool liners is key to ensuring accurate, reproducible semivolatiles data when using a wool-packed liner. This new wool outperforms analogous MS Certified Liners and can help extend column lifetime and lower detection limits for active semivolatile compounds.

Figure 1  Active compounds, such as 2,4-dinitrophenol, can be seen at lower levels with Restek's Semivolatiles Wool liners.

Peaks 1. 3-Nitroaniline 2. 2,4-Dinitrophenol 3. Pentachlorophenol 4. Phenanthrene GC_EV01032 Column Rxi®-5Sil MS, 30 m, 0.25 mm ID, 0.25 µm (cat.# 13623) Sample Phenols/Anilines/Pesticides Test Mix (cat.# 35245) Conc.: 10 ng on-column Injection Inj. Vol.: 1.0 µL splitless (hold 0.75 min.) Liner: Gooseneck Splitless (4mm) w/Semivolatiles Wool (cat.# 20798-231.1) Inj. Temp.: 250 °C Oven Oven Temp: 50 °C (hold 3.5 min.) to 180 °C at 35 °C/min. (hold 3.5 min.) to 300 °C at 20 °C/min. (hold 0.5 min.) Carrier Gas He, constant flow Flow Rate: 1.2 mL/min. Detector FID @ 330 °C Notes FID: Hydrogen: 30mL/min. Air: 400mL/min. Nitrogen: 25mL/min. 10ng 2,4-dinitrophenol (DNP) on-column, single gooseneck liner with wool, FID analysis with phenanthrene internal standard.

Table I  Higher, more consistent responses with Restek's Semivolatiles Wool allow lower detection limits for active compounds.
Restek's Liner with Semivolatiles Wool
Replicate	2,4-DNP Area	Phenanthrene Area	Response Factor
1	25.1	125.9	0.199
2	26.2	125	0.210
3	29.4	135.9	0.216
4	30.7	140	0.219
		Average RF =	0.211
		STD =	0.009
		%RSD =	4.19
Competitor's MS Certified Liner with Wool
Replicate	2,4-DNP Area	Phenanthrene Area	Response Factor
1	7.4	162.3	0.046
2	11.8	165	0.072
3	13.5	165	0.082
4	14.8	165.6	0.089
		Average RF =	0.072
		STD =	0.019
		%RSD =	26.51

Figure 2  The inertness of Semivolatiles Wool liners, coupled with the inertness and resolution of Rxi® technology, results in excellent peak shape, resolution, and response for semivolatiles.

Peaks 1. 1,4-Dioxane 2. N-Nitrosodimethylamine 3. Pyridine 4. 2-Fluorophenol (SS) 5. Phenol-d6 (SS) 6. Phenol 7. Aniline 8. Bis(2-chloroethyl) ether 9. 2-Chlorophenol 10. 1,3-Dichlorobenzene 11. 1,4-dichlorobenzene-d4 (IS) 12. 1,4-Dichlorobenzene 13. Benzyl alcohol 14. 1,2-Dichlorobenzene 15. 2-Methylphenol 16. Bis(2-chloroisopropyl) ether 17. 4-Methylphenol/3-Methylphenol 18. N-Nitrosodi-N-propylamine 19. Hexachloroethane 20. Nitrobenzene-d5 (SS) 21. Nitrobenzene 22. Isophorone 23. 2-Nitrophenol 24. 2,4-Dimethylphenol 25. Benzoic acid 26. Bis(2-chloroethoxy)methane 27. 2,4-Dichlorophenol 28. 1,2,4-Trichlorobenzene 29. Naphthalene-d8 (SS) 30. Naphthalene 31. 4-Chloroaniline 32. Hexachlorobutadiene 33. 4-Chloro-3-methylphenol 34. 2-Methylnaphthalene 35. 1-Methylnaphthalene 36. Hexachlorocyclopentadiene 37. 2,4,6-Trichlorophenol 38. 2,4,5-Trichlorophenol 39. 2-Fluorobiphenyl (SS) 40. 2-Chloronaphthalene 41. 2-Nitroaniline 42. 1,4-Dinitrobenzene 43. Dimethyl phthalate 44. 1,3-Dinitrobenzene 45. 2,6-Dinitrotoluene 46. 1,2-Dinitrobenzene Peaks 47. Acenaphthylene 48. 3-Nitroaniline 49. Acenaphthene-d10 (IS) 50. Acenaphthene 51. 2,4-Dinitrophenol 52. 4-Nitrophenol 53. 2,4-Dinitrotoluene 54. Dibenzofuran 55. 2,3,5,6-Tetrachlorophenol 56. 2,3,4,6-Tetrachlorophenol 57. Diethyl phthalate 58. 4-Chlorophenyl phenyl ether 59. Fluorene 60. 4-Nitroaniline 61. 4,6-Dinitro-2-methylphenol 62. N-Nitrosodiphenylamine (Diphenylamine) 63. 1,2-Diphenylhydrazine (as Azobenzene) 64. 2,4,6-Tribromophenol (SS) 65. 4-Bromophenyl phenyl ether 66. Hexachlorobenzene 67. Pentachlorophenol 68. Phenanthrene-d10 (IS) 69. Phenanthrene 70. Anthracene 71. Carbazole 72. di-n-Butyl phthalate 73. Fluoranthene 74. Benzidine 75. Pyrene-d10 (SS) 76. Pyrene 77. p-Terphenyl-d14 (SS) 78. 3,3'-Dimethylbenzidine 79. Butyl benzyl phthalate 80. Bis(2-ethylhexyl) adipate 81. 3,3'-Dichlorobenzidine 82. Benz[a]anthracene 83. Bis(2-ethylhexyl)phthalate 84. Chrysene-d12 (IS) 85. Chrysene 86. Di-n-octyl phthalate 87. Benzo[b]fluoranthene 88. Benzo[k]fluoranthene 89. Benzo[a]pyrene 90. Perylene-d12 (IS) 91. Dibenz[a,h]anthracene 92. Indeno[1,2,3-cd]pyrene 93. Benzo[ghi]perylene C = Toluene GC_EV01129 Column Rxi®-5Sil MS, 30 m, 0.25 mm ID, 0.25 µm (cat.# 13623) Sample 8270 MegaMix® (cat.# 31850) Benzoic acid (cat.# 31879) 8270 Benzidines Mix (cat.# 31852) Acid Surrogate Mix (4/89 SOW) (cat.# 31025) Revised B/N Surrogate Mix (cat.# 31887) 1,4-dioxane (cat.# 31853) SV Internal Standard Mix (cat.# 31206) Diluent: methylene chloride Conc.: 10 ng on-column Injection Inj. Vol.: 1.0 µL pulsed splitless (hold 0.25 min.) Liner: Gooseneck Splitless (4mm) w/Application-Specific Wool (cat.# 20798-231.1) Inj. Temp.: 250 °C Pulse Pressure: 25 psi (172.4kPa) Pulse Time: 0.3 min. Purge Flow: 60 mL/min. Oven Oven Temp: 40 °C (hold 1 min.) to 280 °C at 25 °C/min. to 320 °C at 5 °C/min. (hold 1 min.) Carrier Gas He, constant flow Flow Rate: 1.2 mL/min. Detector MS Mode: Scan Transfer Line Temp.: 280 °C Analyzer Type: Quadrupole Source Temp.: 250 °C Tune Type: DFTPP Ionization Mode: EI Scan Range: 35-550 amu Instrument Agilent 7890A GC & 5975C MSD

Figure 3  Improve low level response for active compounds—more than twice the required RF for 2,4-DNP at just 1ng on column. (Extracted ion chromatogram)

Peaks 1. 2,4-Dinitrophenol GC_EV01031 Column Rxi-5Sil MS, 30 m, 0.25 mm ID, 0.25 µm (cat.# 13623) Sample 8270 MegaMix® (cat.# 31850) Conc.: 1 ng on-column Injection Inj. Vol.: 1.0 µL splitless (hold 0.2 min.) Liner: Siltek® Gooseneck Splitless (4mm) w/ Semivolatiles Wool (cat.# 22406-231.5) Inj. Temp.: 250 °C Oven Oven Temp: 35 °C (hold 1 min.) to 280 °C at 25 °C/min. to 310 °C at 5 °C/min. (hold 1 min.) Carrier Gas He, constant flow Flow Rate: 1.2 mL/min. Detector MS Mode: Scan Transfer Line Temp.: 280 °C Ionization Mode: EI Scan Range: 35-500 amu

To order the new Semivolatiles Wool in prepacked liners, add the corresponding suffix number to the liner catalog number.

qty.	IP Deactivated Liner  with Semivolatiles Wool
each	-231.1 (addl. cost)
5-pk.	-231.5 (addl. cost)
25-pk.	-231.25 (addl. cost)

By Amanda Rigdon, Pharmaceutical Innovations Chemist and Rick Lake, Pharmaceutical Market Development Manager