HPLC Separation of Hydrophilic Amines on Amaze SC Mixed-Mode Column with LC/MS Compatible Conditions
Conditions of Experiment
Column: Amaze SC
Separation Modes: reversed-phase cation-exchange
Column Dimenstions: 3.0x100 mm, 5 μm, 100A
Mobile Phase: 30% ACN with 50 mM AmFm pH 3
Detection: UV 250 nm
Sample: 0.3 mg/ml
Injection: 2 μL
Flow rate: 0.6 ml/min
Class of compounds: Amines, Aromatic base, Pyridine
Nature of compounds: Basic, Hydrophilic, Polar
Compounds: Aniline (herbicide precursor), Pyridine, 2-Aminopyridine, 4-Aminophenol, 2,6-Lutidine
Effect of Acid in Mobile Phase on Retention of Neutral, Acidic, Basic and Zwitterionic Compounds on Core-Shell Mixed-Mode Column
Application description

Seven compounds with different properties were used to study an effect of different acids in the mobile phase on retention of neutral, acidic, basic and zwitterionic compounds on core-shell mixed-mode Coresep 100 column. Retention of neutral compounds (propylparaben and benzoic acid to some extent) are not affected by change of the acid. Basic compounds (norphenylephrine, pyridine, 2-aminopyridine) respond the most to change of the strength of acidic additive. Several folds increase in retention time was observed when sulfuric acid was replaced with formic acid in the mobile phase. Effect of acid on retention time of zwitterionic amino acids was less pronounced but still provided excellent retention and peak shape for zwitterionic compounds (5-aminosalicylic acid, phenylalanine). This methods and Coresep 100 column can be used to retain and separate other acidic, basic, neutral and zwitterionic compounds without the use of ion-pairing reagent in the mobile phase. Various detection techniques can be used based on the properties of the mobile phase and analytes. Methods are compatible with mall major detection techniques 9UV, ELSD, CAD, RI, MS). Core-shell mixed-mode columns provide great combination of unique selectivity, speed and efficiency.

Conditions of Experiment
Column: Coresep 100
Separation Modes: reversed-phase, cation-exchange
Column Dimenstions: 4.6 x 150 mm, 2.7 um, 90A
Mobile Phase: 30% ACN with various acids
Detection: UV 255 nm
Sample: 0.3 mg/ml
Injection: 3 uL
Flow rate: 1 mL/min
Class of compounds: Amines, Amino acid, Aromatic acid, Aromatic base, Aromatic compound, Catecholamine, Drug, Isomer, Neurotransmitter, Organic acid, Preservative, Pyridine, Supplement
Nature of compounds: Acidic, Basic, Hydrophilic, Hydrophobic, Neutral, Polar, Zwitterionic
Compounds: 5-Aminosalicylic acid, Norphenylephrine, Phenylalanine, Pyridine, Benzoic acid, 2-Aminopyridine, Propylparaben
HPLC Method for Analysis of Pyridine and Three Isomers of Aminopyridine on Amaze HD Column
Application description

Pyridine and its derivatives are important building blocks in pharmaceutical and chemical industries. Most pyridines are hydrophilic compounds with pKa around 5.2-6. Hydrophilic nature of these compounds require the use of ion-pairing reagents that are not compatible with LC/MS. We have developed a method with good peak shape and resolution. All compounds are eluted within 6 minutes with isocratic HPLC conditions. Mobile phase allows to use mass spectrometry detection technique as well as UV, ELSD, RI and CAD.

Conditions of Experiment
Column: Amaze HD
Separation Modes: hydrogen-bonding and cation-exchange
Column Dimenstions: 3.2 x 150 mm
Mobile Phase: MeCN/MeOH (60/40) with 0.2% HCOOH and 0.25% AmFm
Detection: UV 275 nm
Sample: 0.3 mg/ml
Injection: 1 uL
Flow rate: 1.0 mL/min
Class of compounds: Amines, Aromatic base, Aromatic compound, Isomer, Pyridine
Nature of compounds: Basic, Hydrophilic, Polar
Compounds: Pyridine, 2-Aminopyridine, 3-Aminopyridine, 4-Aminopyridine