Practical HPLC method development

Snyder, Lloyd R.; Glajch, Joseph L.; Kirkland, J. J.; Abbott, Richard

doi:10.1016/s0003-2670(00)80235-4

Cited by 116 publications

(247 citation statements)

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“…In LC, the early mixed-mode RP/ion-exchange phases have been developed predominantly based on organic polymer because of their good pH stability and the relative ease of introducing cation exchange sites [45]. However, these purely organic polymeric phases generally have low plate counts compared to silica based stationary phases [46] and equilibration upon a change in solvent can be slow [46,47]. More recently, several examples of silica based mixed-mode RP/ion-exchange phases have also been reported [23-30,48-50].…”

Section: Introductionmentioning

confidence: 99%

Novel ultra stable silica-based stationary phases for reversed phase liquid chromatography—Study of a hydrophobically assisted weak acid cation exchange phase

Zhang¹,

Carr²

2011

Journal of Chromatography A

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A mixed-mode reversed-phase/weak cation exchange (RP/WCX) phase has been developed by introducing a small amount of carboxylate functionality into a hydrophobic hyper-crosslinked (HC) platform. This silica based HC-platform was designed to form an extensive polystyrene network completely confined to the particle's surface. The fully connected polymer network prevents the loss of bonded phase, which leads to superior hydrolytic stability of the new phase when compared to conventional silica based phases. Compared to previously introduced HC phases the added carboxylic groups impart a new weak cation exchange selectivity to the base hydrophobic HC platform. The phase thus prepared shows a mixed-mode retention mechanism, allowing for both neutral organic compounds and bases of a wide polarity range to be simultaneously separated on the same phase under the same conditions. In addition, the new phase offers the flexibility that gradients in organic modifier, pH or ionic competitors can be used to affect the separation of a wide range of solutes. Moreover, the inherent weak acid cation exchange groups allow formic and acetic acid buffers to be used as eluents thereby avoiding the mass spectrometric ionization suppression problems concomitant to the use of non-volatile additives such as strong amine modifiers (e.g. triethylamine) or salts (e.g. NaCl) to elute basic solutes from the strong cation exchange phase which was previously developed in this lab. The use of the new phase for achieving strong retention of rather hydrophilic neurotransmitters and drugs of abuse without the need for ion pairing agents is demonstrated.

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Section: Introductionmentioning

confidence: 99%

Novel ultra stable silica-based stationary phases for reversed phase liquid chromatography—Study of a hydrophobically assisted weak acid cation exchange phase

Zhang¹,

Carr²

2011

Journal of Chromatography A

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“…It is well recognized that conducting liquid chromatography (LC) at elevated temperatures (typically 30–80°C) results in beneficial effects, such as improvements in peak shape, selectivity, and resolution 5,6. Furthermore, elevated temperatures are sometimes required for the successful elution of very hydrophobic proteins 7.…”

Section: Introductionmentioning

confidence: 99%

Quantitative Improvements in Peptide Recovery at Elevated Chromatographic Temperatures from Microcapillary Liquid Chromatography−Mass Spectrometry Analyses of Brain Using Selected Reaction Monitoring

et al. 2010

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Elevated chromatographic temperatures are well recognized to provide beneficial analytical effects. Previously, we demonstrated that elevated chromatographic temperature enhances the identification of hydrophobic peptides prepared from enriched membrane samples. Here, we quantitatively assess and compare the recovery of peptide analytes from both simple and complex tryptic peptide matrices using the SRM mass spectrometry. Our study demonstrates that elevated chromatographic temperature results in significant improvements in the magnitude of peptide recovery for both hydrophilic and hydrophobic peptides from both simple and complex peptide matrices. Importantly, the analytical benefits for quantitative measurements in whole mouse brain matrix are demonstrated, suggesting broad utility in the proteomic analyses of complex mammalian tissues. Any improvement in peptide recovery from chromatographic separations translates directly to the apparent sensitivity of downstream mass analysis in μLC-MS/MS based proteomic applications. Therefore, the incorporation of elevated chromatographic temperatures should result in significant improvements in peptide quantification as well as detection and identification.

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“…The addition of modifiers (TFA and acetic acid) was also studied since these have substantial effects on selectivity and efficiency [13]. However, we found no significant improvements of the HPLC separation or peak shape of (R,R)-Fen.…”

Section: Resultsmentioning

confidence: 81%

HPLC–electrospray mass spectrometric assay for the determination of (R,R)-fenoterol in rat plasma

Siluk

Kim

Cole

et al. 2008

Journal of Pharmaceutical and Biomedical Analysis

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A fast and specific liquid chromatography-mass spectrometry method for the determination of (R,R)-fenoterol ((R,R)-Fen) in rat plasma has been developed and validated. (R,R)-Fen was extracted from 125 µl of plasma using solid phase extraction and analyzed on Atlantis HILIC Silica 3 µm column. The mobile phase was composed of acetonitrile:ammonium acetate (pH 4.1; 20 mM) (85:15, v/v), at a flow rate of 0.2 ml/min. The lower limit of detection (LLOD) was 2 ng/ml . The procedure was validated and applied to the analysis of plasma samples from rats previously administered (R,R)-Fen in an intravenous bolus.

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Practical HPLC method development

Cited by 116 publications

References 0 publications

Novel ultra stable silica-based stationary phases for reversed phase liquid chromatography—Study of a hydrophobically assisted weak acid cation exchange phase

Novel ultra stable silica-based stationary phases for reversed phase liquid chromatography—Study of a hydrophobically assisted weak acid cation exchange phase

Quantitative Improvements in Peptide Recovery at Elevated Chromatographic Temperatures from Microcapillary Liquid Chromatography−Mass Spectrometry Analyses of Brain Using Selected Reaction Monitoring

HPLC–electrospray mass spectrometric assay for the determination of (R,R)-fenoterol in rat plasma

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