Agilent HPLC Column Selection Guide
Agilent HPLC Column Selection Guide Agilent HPLC Column Selection Guide
Columns for Bioanalytical ChromatographyLC/MS Analysis of Angiotensin on Extend-C18Column: ZORBAX Extend-C18773700-9022.1 x 150 mm, 5 µmMobile Phase: Acidic Conditions:A: 0.1% TFA in waterB: 0.085% TFA in 80% acetonitrile (ACN)Basic Conditions:A: 10 mM NH 4 OH in waterB: 10 mM NH 4 OH in 80% ACNFlow Rate: 0.2 mL/minGradient: 15-50% B in 15 minTemperature: 35°CMS Conditions: Pos. Ion ESI- Vf 70V, Vcap 4.5 kV,N2- 35 psi, 12 L/min., 325°CSample:2.5 µL sample (50 pmol each)Angiotensin I, II, IIIAAngiotensin IMax: 10889Low pHTIC (200-1500 m/z)5.0E64.0E63.0E62.0E61.0E6AII + AIII12.05013.261AI10080604020227.1250.9340.3398.0433.0+3458.0649.5+2659.7720.3767.3844.81025.21179.11277.91326.31373.31468.8002.55.0 7.5 10 12.5Time (min)005001000m/zBAngiotensin IMax: 367225High pHTIC (200-1500 m/z)5.0E74.0E73.0E72.0E71.0E7002.5AII5.499AIAIII8.4779.6215.0 7.5 10 12.5Time (min)1008060402000433.0+3500649.0659.8+210001296.6+1m/zLC30003Reference: B.E. Boyes. Separation and Analysis of Peptides at High pHUsing RP-HPLC/ESI-MS, 4th WCBP, San Francisco, CA Jan 2000.Both small and large peptides demonstrate selectivity changes at high and low pH. At high pH, due to a change in charge, all three Angiotensins can be resolved. In addition,the spectral clarity of Angiotensin I is dramatically improved at high pH with the ammonium hydroxide mobile phase. The Extend-C18 column can be used for the analysis ofsmall peptides at high pH as well.120 Order online at www.agilent.com/chem/store
Columns for Bioanalytical ChromatographyZORBAX 300Å Extend-C18Hardware DescriptionSize (mm)Particle Size(µm) Part No.Analytical 4.6 x 250 5 770995-902Analytical 4.6 x 150 5 773995-902Rapid Resolution 4.6 x 150 3.5 763973-902Rapid Resolution 4.6 x 100 3.5 761973-902Rapid Resolution 4.6 x 50 3.5 765973-902Narrow Bore RR* 2.1 x 150 3.5 763750-902Narrow Bore RR* 2.1 x 100 3.5 761775-902Narrow Bore RR* 2.1 x 50 3.5 765750-902Guard Cartridge, 4/pk 4.6 x 12.5 5 820950-932Guard Cartridge, 4/pk 2.1 x 12.5 5 821125-932Guard Hardware Kit 820888-901Capillary Glass-lined ColumnsCapillary RR* 0.3 x 150 3.5 5065-4464Capillary RR* 0.3 x 100 3.5 5065-4465Capillary RR* 0.3 x 75 3.5 5065-4466Capillary RR* 0.3 x 50 3.5 5065-4467*RR: Rapid Resolution 3.5 µmOrder online at www.agilent.com/chem/store121
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<strong>Column</strong>s for Bioanalytical ChromatographyLC/MS Analysis of Angiotensin on Extend-C18<strong>Column</strong>: ZORBAX Extend-C18773700-9022.1 x 150 mm, 5 µmMobile Phase: Acidic Conditions:A: 0.1% TFA in waterB: 0.085% TFA in 80% acetonitrile (ACN)Basic Conditions:A: 10 mM NH 4 OH in waterB: 10 mM NH 4 OH in 80% ACNFlow Rate: 0.2 mL/minGradient: 15-50% B in 15 minTemperature: 35°CMS Conditions: Pos. Ion ESI- Vf 70V, Vcap 4.5 kV,N2- 35 psi, 12 L/min., 325°CSample:2.5 µL sample (50 pmol each)Angiotensin I, II, IIIAAngiotensin IMax: 10889Low pHTIC (200-1500 m/z)5.0E64.0E63.0E62.0E61.0E6AII + AIII12.05013.261AI10080604020227.1250.9340.3398.0433.0+3458.0649.5+2659.7720.3767.3844.81025.21179.11277.91326.31373.31468.8002.55.0 7.5 10 12.5Time (min)005001000m/zBAngiotensin IMax: 367225High pHTIC (200-1500 m/z)5.0E74.0E73.0E72.0E71.0E7002.5AII5.499AIAIII8.4779.6215.0 7.5 10 12.5Time (min)1008060402000433.0+3500649.0659.8+210001296.6+1m/zLC30003Reference: B.E. Boyes. Separation and Analysis of Peptides at High pHUsing RP-<strong>HPLC</strong>/ESI-MS, 4th WCBP, San Francisco, CA Jan 2000.Both small and large peptides demonstrate selectivity changes at high and low pH. At high pH, due to a change in charge, all three Angiotensins can be resolved. In addition,the spectral clarity of Angiotensin I is dramatically improved at high pH with the ammonium hydroxide mobile phase. The Extend-C18 column can be used for the analysis ofsmall peptides at high pH as well.120 Order online at www.agilent.com/chem/store
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