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Separation of Catecholamines |
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Application Notes:
Primesep 200 separates catecholamines in less than 8 minutes. Tyrosine, dl-DOPA, dopamine, epinephrine are baseline resolved by a combination of reversed-phase, ion-exchange, and ion-exclusion mechanisms. Excellent peak shape results with a mass spec compatible mobile phase of water, acetonitrile (MeCN, ACN) and trifluoracetic acid (TFA) with UV detection at 210 nm.
Application Columns:
- Primesep® 200 is a mixed-mode HPLC column with general hydrophobic and ion-exchange properties. The Primesep 200 stationary phase combines a hydrophobic chain and a weakly acidic functional group as a cation exchanger. Acids, bases, zwitterions and neutrals can be retained and separated by Primesep 200 columns. Primesep 200 retains hydrophobic compounds by reversed-phase retention alone or a combination of reversed-phase and ion-exchange retention. Primesep 200 separates underivatized amino acids, amines, isomers and structurally related compounds by combining reversed phase and ion exchange. In this case the acidic functional group acts as an embedded ion-pairing reagent. Organic acids and diacids are separated by reversed phase and ion exclusion. Methods can be developed on Primesep columns that are fully compatible with detectors such as UV, RI, conductivity, fluorescence, evaporative light scattering (ELSD) as well as mass spectrometers (LC/MS) and preparative chromatography (prep HPLC) systems.
Application Analytes:
Application Detection:
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Application Notes:
Primesep 200 separates catecholamines in the catecholamine pathway in 10 minutes. Phenylalanine, tyrosine, DOPA, dopamine, norepinephrine, and epinephrine are baseline resolved by a combination of reversed-phase, ion-exchange, and ion-exclusion mechanisms. Excellent peak shape results with a mass spec compatible mobile phase of water, acetonitrile (MeCN, ACN) and trifluoracetic acid (TFA) with UV detection at 210 nm.
Application Columns:
- Primesep® 200 is a mixed-mode HPLC column with general hydrophobic and ion-exchange properties. The Primesep 200 stationary phase combines a hydrophobic chain and a weakly acidic functional group as a cation exchanger. Acids, bases, zwitterions and neutrals can be retained and separated by Primesep 200 columns. Primesep 200 retains hydrophobic compounds by reversed-phase retention alone or a combination of reversed-phase and ion-exchange retention. Primesep 200 separates underivatized amino acids, amines, isomers and structurally related compounds by combining reversed phase and ion exchange. In this case the acidic functional group acts as an embedded ion-pairing reagent. Organic acids and diacids are separated by reversed phase and ion exclusion. Methods can be developed on Primesep columns that are fully compatible with detectors such as UV, RI, conductivity, fluorescence, evaporative light scattering (ELSD) as well as mass spectrometers (LC/MS) and preparative chromatography (prep HPLC) systems.
Application Analytes:
Application Detection:
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Separation of Epinephrine Norepinephrine |
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Application Notes:
Primesep 200 separates the catecholamines, norepinephrine and epinephrine, less than 8 minutes. Norepinephrine and epinephrine are baseline resolved by a combination of reversed-phase, and ion-exchange mechanisms. Excellent peak shape results with a mass spec compatible mobile phase of water, acetonitrile (MeCN, ACN) and trifluoracetic acid (TFA) with UV detection at 210 nm.
Application Columns:
- Primesep® 200 is a mixed-mode HPLC column with general hydrophobic and ion-exchange properties. The Primesep 200 stationary phase combines a hydrophobic chain and a weakly acidic functional group as a cation exchanger. Acids, bases, zwitterions and neutrals can be retained and separated by Primesep 200 columns. Primesep 200 retains hydrophobic compounds by reversed-phase retention alone or a combination of reversed-phase and ion-exchange retention. Primesep 200 separates underivatized amino acids, amines, isomers and structurally related compounds by combining reversed phase and ion exchange. In this case the acidic functional group acts as an embedded ion-pairing reagent. Organic acids and diacids are separated by reversed phase and ion exclusion. Methods can be developed on Primesep columns that are fully compatible with detectors such as UV, RI, conductivity, fluorescence, evaporative light scattering (ELSD) as well as mass spectrometers (LC/MS) and preparative chromatography (prep HPLC) systems.
Application Analytes:
Application Detection:
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HPLC Separation of Compounds of Catecholamine Pathway |
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Application Notes:
The catecholamine neurotransmitters are amino acid derivatives of tyrosine. DOPA, tyrosine, phenylalanine, norepinephrine, epinephrine, and dopamine and baseline resolved on a Primesep 200 column. The HPLC separation uses a mobile phase of water, acetonitrile (MeCN, ACN), trifluoracetic acid (TFA) or ammonium formate and ultraviolet (UV) detection at 210 nm. Peak order and retention time can be changed by switching from TFA to ammonium formate in the mobile phase.
Application Columns:
- Primesep® 200 is a mixed-mode HPLC column with general hydrophobic and ion-exchange properties. The Primesep 200 stationary phase combines a hydrophobic chain and a weakly acidic functional group as a cation exchanger. Acids, bases, zwitterions and neutrals can be retained and separated by Primesep 200 columns. Primesep 200 retains hydrophobic compounds by reversed-phase retention alone or a combination of reversed-phase and ion-exchange retention. Primesep 200 separates underivatized amino acids, amines, isomers and structurally related compounds by combining reversed phase and ion exchange. In this case the acidic functional group acts as an embedded ion-pairing reagent. Organic acids and diacids are separated by reversed phase and ion exclusion. Methods can be developed on Primesep columns that are fully compatible with detectors such as UV, RI, conductivity, fluorescence, evaporative light scattering (ELSD) as well as mass spectrometers (LC/MS) and preparative chromatography (prep HPLC) systems.
Application Analytes:
Application Detection:
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HPLC Separation of Epinephrine and Epinephrine Sulfonate |
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Application Notes:
Application Columns:
- Primesep® 100 is a mixed-mode HPLC column with general hydrophobic and ion-exchange properties. The Primesep 100 stationary phase combines a hydrophobic chain and an acidic functional group as a cation exchanger. Acids, bases, zwitterions and neutrals can be retained and separated by Primesep 100 columns. Primesep 100 retains hydrophobic compounds by reversed-phase retention alone or a combination of reversed-phase and ion-exchange retention. Primesep 100 separates underivatized amino acids and amines by combining reversed-phase and ion-exchange mechanisms. In this case the acidic functional group acts as an embedded ion-pairing reagent. Organic acids are separated by reversed phase and ion exclusion. Inorganic cations can be separated by cation exchange. Methods can be developed on Primesep columns that are fully compatible with detectors such as UV, RI, conductivity, fluorescence, evaporative light scattering (ELSD) as well as mass spectrometers (LC/MS) and preparative chromatography (prep HPLC) systems.
- Primesep® SB is a mixed-mode HPLC column with general hydrophobic and ion-exchange properties. The Primesep SB stationary phase combines a hydrophobic chain and a strongly basic functional group as an anion exchanger. Acids, bases, zwitterions and neutrals can be retained and separated by Primesep SB columns. Primesep SB is ideal to retain hydrophobic compounds by reversed-phase retention due to its C18 carbon chain. Basic hydrophobic compounds are usually retained less on Primesep SB columns than on similar RP columns due to a charge repulsion effect. Primesep SB also separates bases by an ion-exclusion mechanism. Primesep SB demonstrates significant improvement in retention and selectivity for the separation of weak acid analytes by a combination of reversed phase and anion exchange. In this case the basic functional group acts as an embedded ion-pairing reagent. Primesep SB column is one of the best separation media for natural product extracts. Methods can be developed on Primesep columns that are fully compatible with detectors such as UV, RI, conductivity, fluorescence, evaporative light scattering (ELSD) as well as mass spectrometers (LC/MS) and preparative chromatography (prep HPLC) systems.
Application Analytes:
Application Detection:
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HILIC Separation of Epinephrine and Epinephrine Sulfonate |
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Application Notes:
Epinephrine (also referred to as adrenaline) is a hormone and neurotransmitter. It is a catecholamine, a sympathomimetic monoamine derived from the amino acids phenylalanine and tyrosine. This method can be used to determine and quantitate epinephrine and epinephrine sulfonate in biological fluids (urine, blood, serum) and drug formulations. Obelisc N columns are used to retain and separate epinephrine and epinephrine sulfonate in mixed-mode hydrophilic interaction chromatography. Epinephrine is retained by the combination of cation-exchange and HILIC mechanisms. Epinephrine sulfonate is retained by HILIC mechanism. Buffer concentration and pH, as well as the amount of acetonitrile, can be used to adjust retention of both compounds. Both compounds can be detected by UV, ELSD and LC/MS. Preparative separation is possible with volatile mobile phases (ammonium formate or ammonium acetate.
Application Columns:
- Obelisc N columns are the first commercially available columns with Liquid Separation Cell technology (LiSC). With multiple patents pending, LiSC technology is based on a new chemical modification of silica gel pores that creates a liquid separation cell with its own charge characteristics, ionic strength, and hydrophobic properties. Like living cells which exist in equilibrium with the outside environment, liquid separation cells exist in constant equilibrium with the mobile phase. Obelisc N, for normal phase, has very polar characteristics and works well for polar and charged analytes. In ion-exchange mode, charged analytes interact with oppositely charged groups on the stationary phase. In traditional HILIC mode, a charged or neutral polar analyte interacts with a water layer on the polar stationary phase surface. On Obelisc N the charges are greatly separated and independently accessible which results in different selectivity compared to traditional HILIC and silica columns. Mobile phase composition changes the conformation of the long hydrophilic chain. This affects the properties of the liquid separation cell and changes separation selectivity. Typical mobile phases used with Obelisc N columns are based on acetonitrile, water, and the mass spec compatible buffers ammonium formate (pH 3) and ammonium acetate (pH 5). If it is necessary to detect in low UV (<220 nm) then phosphate buffer is recommended. Obelisc N columns are compatible with mass spectrometers (LC/MS), evaporative light scattering detectors and preparative chromatography (prep HPLC) system.
Application Analytes:
Application Detection:
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Separation of Serotonin, Dopamine, and Related Compounds |
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Application Notes:
Catecholamines are chemical compounds derived from the amino acid tyrosine containing catechol and amine groups. Some of them are biogenic amines. Retention of compounds of catecholamine pathway is achieved on Obelisc N column. All polar compounds are well retained by combination of HILIC and ion-exchange mechanisms. Obelisc N columns produce very good peak shapes for all analytes. Method is very sensitive to amount of ACN, buffer and buffer pH. Retention time changes with variation of main parameters. This method can be used for quantitation of biogenic amines and related compounds (homovanillic acid, dihydroxyphenyl acetic acid, serotonin, dopamine, epinephrine, hydroxytryptophan, epinephrine and DOPA) in urine, blood and other biological fluids. Further optimization of this HPLC method can be used during screening and validation. Amines and acids can be analyzed in the same run and retained by combination of polar organic mode, cation-exchange and anion-exchange modes. Various buffers within specified pH can be employed (ammonium formate, ammonium acetate, sodium phosphate, etc.).
Application Columns:
- Obelisc N columns are the first commercially available columns with Liquid Separation Cell technology (LiSC). With multiple patents pending, LiSC technology is based on a new chemical modification of silica gel pores that creates a liquid separation cell with its own charge characteristics, ionic strength, and hydrophobic properties. Like living cells which exist in equilibrium with the outside environment, liquid separation cells exist in constant equilibrium with the mobile phase. Obelisc N, for normal phase, has very polar characteristics and works well for polar and charged analytes. In ion-exchange mode, charged analytes interact with oppositely charged groups on the stationary phase. In traditional HILIC mode, a charged or neutral polar analyte interacts with a water layer on the polar stationary phase surface. On Obelisc N the charges are greatly separated and independently accessible which results in different selectivity compared to traditional HILIC and silica columns. Mobile phase composition changes the conformation of the long hydrophilic chain. This affects the properties of the liquid separation cell and changes separation selectivity. Typical mobile phases used with Obelisc N columns are based on acetonitrile, water, and the mass spec compatible buffers ammonium formate (pH 3) and ammonium acetate (pH 5). If it is necessary to detect in low UV (<220 nm) then phosphate buffer is recommended. Obelisc N columns are compatible with mass spectrometers (LC/MS), evaporative light scattering detectors and preparative chromatography (prep HPLC) system.
Application Analytes:
Application Detection:
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HPLC Separation of Neurotransmitters and Related Drugs |
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Application Notes:
Epinephrine and norepinephrine (adrenaline and noradrenaline) are hormones and neurotransmitters. Epinephrine is synthesized from norepinephrine in a synthetic pathway shared by all catecholamines, including L-dopa, dopamine, norepinephrine, and epinephrine. Phenylephrine is used as a decongestant, available as an oral medicine or as a nasal spray. Phenylephrine is now the most common over-the-counter (OTC) decongestant. All three compounds are used in various drug compositions. Separation of epinephrine and norepinephrine is a challenging task due to polarity and close properties of two compounds. Epinephrine, norepinephrine and phenylephrine are separated in this method on Obelisc R mixed-mode HPLC columns. Method is very sensitive to variation of pH and pH adjustment can be used to achieve desired selectivity and retention time. Other catecholamines can be analyzed using this HPLC method. Method can be used as stability indicating or impurity profiling approach to analysis of neurotransmitters in drug formulation, blood, serum and urine.
Application Columns:
- Obelisc R columns are the first commercially available columns with Liquid Separation Cell technology (LiSC). With multiple patents pending, LiSC technology is based on a new chemical modification of silica gel pores that creates a liquid separation cell with its own charge characteristics, ionic strength, and hydrophobic properties. Like living cells which exist in equilibrium with the outside environment, liquid separation cells exist in constant equilibrium with the mobile phase. Obelisc R, for reversed phase, has reversed-phase character and can be used in traditional, reversed-phase type applications. Due to the presence of ionic groups and a long hydrophobic chain, Obelisc R offers additional retention and tuning that is not available with traditional reversed-phase columns. Reversed-phase separations are augmented by additional ionic and polar interactions not available on traditional reversed-phase C18 columns. Mobile phase composition changes the conformation of the long hydrophobic chain and affects the properties of the liquid separation cell and changes separation selectivity. Typical mobile phases used with Obelisc R columns are based on acetonitrile, water, and the mass spec compatible buffers ammonium formate (pH 3) and ammonium acetate (pH 5). If it is necessary to detect in low UV (<220 nm) then phosphate buffer is recommended. Obelisc R columns are compatible with mass spectrometers (LC/MS), evaporative light scattering detectors and preparative chromatography (prep HPLC) systems.
Application Analytes:
Application Detection:
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