HPLC Separation of Hydrophobic, Hydrophilic, Basic and Acidic Compounds on Amaze TR Mixed-Mode Column
Conditions of Experiment
Column: Amaze TR
Separation Modes: RP, cation-exchange, anion-exchange
Column Dimenstions: 4.6x150 mm, 5 um, 100A
Mobile Phase: 30% ACN with 30 mM AmAc pH 5
Detection: UV 275 nm
Sample: 0.2-0.3 mg/ml
Injection: 3 uL
Flow rate: 1 ml/min
Class of compounds: Aromatic base, Aromatic compound, Catecholamine, Drug, Neurotransmitter, Organic acid, Organic compound, Preservative, Stimulant
Nature of compounds: Acidic, Basic, Hydrophilic, Hydrophobic, Neutral, Polar
Compounds: Norphenylephrine, Ephedrine, 3,4-Dihydroxybenzoic acid, Benzenesulfonic acid, Propylparaben, Propranolol
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 Neurotransmitters on Amaze HD Column
Application description

Four neurotransmitters - pseudoephedrine, norephedrine, phenylephrine and norphenylephrine - were separated in hydrogen-bonding and cation-exchange mode. Pseudoephedrine is sympathomimetic drug used either as a single ingredient or part of the composition of OTC or Rx medications. Phenylephrine is a hydrophilic basic drug which is used as decongestant in OTC medication composition, such as Sudafed, Benadryl, Advil, Nyquil, etc. Norphenylephrine is an adrenergic agent used as sympathomimetic drug. Norephedrine is naturally occurring endogenous trace amine that plays a role as neurotransmitter in the brain, and is used as decongestant in many OTC compositions as well as an appetite suppressant. All four drugs are hydrophilic and basic in nature, and have no retention on traditional reversed-phase column without an ion-pairing reagent. Method provides an efficient and robust separation for all drugs. The method can be used in analysis of neurotransmitters in biofluids and other sample matrices. The method is compatible with all detection techniques: MS, RI, UV, ELSD and CAD.

Conditions of Experiment
Column: Amaze HD
Separation Modes: hydrogen-bonding and cation-exchange
Column Dimenstions: 3.2 x 50 mm, 3 um
Mobile Phase: MeCN/MeOH (95/5) with 0.5% HCOOH and 0.05% AmFm
Detection: UV 275 nm
Sample: 0.3 mg/ml
Injection: 1 uL
Flow rate: 1.0 mL/min
Class of compounds: Amino acid, Aromatic base, Catecholamine, Drug, Neurotransmitter, Stimulant, Supplement
Nature of compounds: Basic, Hydrophilic, Polar, Zwitterionic
Compounds: Pseudoephedrine, Norepinephrine, Phenylalanine, Norphenylephrine