Adenosine

Adenosine C10H13N5O4

Adenosine

Applications

HPLC Method for Analysis of Nucleosides and Nucleoside Derivatives on Amaze HD Column

HPLC Method for Analysis of Nucleosides and Nucleoside Derivatives on Amaze HD Column chromatogram
Conditions of Experiment
Column:
Amaze HD
Separation Modes:
hydrogen-bonding and cation-exchange
Column Dimenstions: 3.2 x 100 mm
Mobile Phase: MeCN/MeOH from 90/10 to 50/50 with 0.1% HCOOH and 0.01% AmFm in 8 min
Detection: UV 275 nm
Sample: 0.3 mg/ml
Injection: 1 uL
Flow rate: 0.8 mL/min
Analytes
1. Thymidine
2. Uridine
3. 2′-Deoxyadenosine
4. Adenosine
5. Guanosine

Class of compounds: Aromatic base, Nucleoside
Nature of compounds: Basic, Hydrophilic, Neutral, Polar

Application description

Thymidine, uridine, deoxyadenosine, adenosine, deoxyguanosine and guanosine were separated on an Amaze HD column with LC/MS compatible method. The approach can be used to analyze nucleotides, nucleosides and their derivatives in various sample matrices. This HPLC method is robust and reproducible, and can be used for other compounds that can form hydrogen bonds with stationary phase and interact by electrostatic interaction.

HPLC Method for Analysis of Adenosine and Adenine on Amaze HD Column

HPLC Method for Analysis of Adenosine and Adenine on Amaze HD Column chromatogram
Conditions of Experiment
Column:
Amaze HD
Separation Modes:
HILIC
Column Dimenstions: 3.2 x 50 mm
Mobile Phase: MeCN/MeOH (95/5) with 0.5% HCOOH with 0.05% AmFm
Detection: UV 275 nm
Sample: 0.3 mg/ml
Injection: 1 uL
Flow rate: 1.0 mL/min
Analytes
1. Adenosine
2. Adenine

Class of compounds: Aromatic base, Nucleobase, Nucleoside
Nature of compounds: Basic, Hydrophilic, Polar

Application description

Adenosine is a purine nucleoside composed of adenine and ribose fragments. Adenine is a purine-based nucleobase. Both compounds are polar and slightly basic, with ability to form hydrogen bonding with the stationary phase. Both compounds were separated within 2 minutes with an isocratic mobile phase. Method is compatible with mass spectrometry.

Separation of Nine Nucleotides with Core-Shell Mixed-Mode Column

Separation of Nine Nucleotides with Core-Shell Mixed-Mode Column chromatogram
Conditions of Experiment
Column:
Coresep SB
Separation Modes:
reversed-phase, anion-exchange
Column Dimenstions: 4.6 x 150 mm, 5 um, 100A
Mobile Phase: 5% ACN, AmAc buffer (pH 4.5) gradient from 20 mmol to 300 mmol in 45 minutes
Detection: UV 270 nm
Sample: 0.3 mg/ml
Injection: 1 uL
Flow rate: 1.5 ml/min
Analytes
1. Cytidine
2. Guanosine monophosphate
3. Guanosine diphosphate
4. Guanosine triphosphate
5. Adenosine

Class of compounds: Amino acid, Aromatic acid, Aromatic base, Nucleoside
Nature of compounds: Acidic, Basic, Hydrophilic, Polar

Application description

Core-shell columns are able to handle higher flow-rates, while maintaining high efficiency and low back-pressure. Coresep SB was used to achieve separation of nine nucleotides. The strong embedded basic ion-pairing groups make it possible to separate compounds that don’t retain on reverse-phase columns.

Separation of Model Compounds in Reversed-Phase and Cation-Exchange Modes on Coresep 100 Mixed-Mode Column

Separation of Model Compounds in Reversed-Phase and Cation-Exchange Modes on Coresep 100 Mixed-Mode Column chromatogram
Conditions of Experiment
Column:
Coresep 100
Separation Modes:
reversed-phase and cation-exchange
Column Dimenstions: 3.2 x 100 mm, 2.7 um, 90A
Mobile Phase: ACN gradient from 10% to 65% in 5 min, buffer gradinet - Amonium formate pH 2.9 from 30 to 70 mM in 5 min
Detection: UV 270 nm, ELSD
Sample: 0.3 mg/ml
Injection: 1 uL
Flow rate: 1.2 ml/min
Analytes
1. Adenosine
2. 3,4-Difluoroaniline
3. 4-Amino-2-chloropyridine
4. 5-Aminoindole
5. 4-amino-3-chloropyridine
6. 2-Amino-5-methylthiazole
7. 4-Ethylaniline

Class of compounds: Amines, Aromatic base, Aromatic compound, Isomer, Nucleoside
Nature of compounds: Basic, Hydrophilic, Polar

Application description

Many pharmaceutical and chemical companies need a general approach for analysis of pharmaceutical ingredients and precursors. Developing universal screening procedures in chromatography can help save time in method development and utilize universal platform to analyze complex pharmaceutical mixtures,  formulation, metabolites, vitamins, etc. With over 600 reversed-phase column and thousands of mobile phase combinations chemists are facing a challenging task on what column to choose. In a lot of cases scientists need to develop several screening methods to address a complex mixture. Mixed-mode approach which combines reversed-phase, HILIC and ion-exchange mechanisms can address an issue of complex mixtures screening. When multiple columns are screened, very often in different modes of separation, multiple mobile phase needs to be prepared.  A simple and universal platform based on mixed-mode columns can be developed. This approach eliminates cumbersome multi-column multi-mobile phase efforts and streamlines method development process. Since mixed-mode columns explore at least two mechanism of interaction selectivity of the separation is drastically different. The model is based on the fact that compounds are retained by combination of reversed-phase and ion-exchange mechanism and that none of the compounds have exactly the same reversed-phase and ion-exchange properties. By exploring small difference in reversed-phase and ion-exchange properties, complex mixtures can be analyzed. The synergy effect of two mechanism allows longer retention and better selectivity than in traditional “single” mode approach. Short method with high efficiency was developed utilizing new core-shell mixed-mode column. All 7 compounds were separated within 5 minutes. Method and Coresep 100 column can be adopted in walk in labs for fast screening of new drug candidates and impurities, components of formulations and building blocks in chemical manufacturing. All methods are compatible with LC/MS and prep chromatography.

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