HPLC Method for Caffeine, Theophylline, Theobromine on Primesep SB by SIELC Technologies

High Performance Liquid Chromatography (HPLC) Method for Analysis of Caffeine, Theophylline, Theobromine

Caffeine is a natural stimulant and methylxanthine alkaloid. with the molecular formula C₆H₁₀N₄O₂. Caffeine can be found in a variety of plants, including tea, coffea, cocoa, kola nuts, and guarana. Ingestion of it can increase alertness and cognitive function. It can also cause worsening anxiety, heart palpitations, and headaches.

Theophylline is a methylxanthine compound with the molecular formula C₇H₈N₄O₂. It is a caffeine metabolite and is often used in forensic analyses. Besides analytical use, it is often used in treating respiratory conditions.

Theobromine is a methylxanthine alkaloid with the molecular formula C₇H₈N₄O₂. It is a derivative of caffeine. It can be found in many foods, including chocolate and tea. Despite that, it is toxic to dogs.

Caffeine, Theophylline, Theobromine can be retained and analyzed using the Primesep SB stationary phase column. The analysis utilizes an isocratic method with a simple mobile phase consisting of water and ethanol (EtOH) with a phosphoric acid buffer. Detection is performed using UV.

Column	Primesep SB, 4.6 x 150 mm, 5 µm, 100 A, dual ended
Mobile Phase	EtOH- 10%
Buffer	Phosphoric Acid
Flow Rate	1.0 ml/min
Detection	UV 270 nm

Class of Compounds	Alkaloid
Analyzing Compounds	Caffeine, Theophylline, Theobromine

HPLC Method for Caffeine, Theobromine, Theophylline on Primesep SB by SIELC Technologies

High Performance Liquid Chromatography (HPLC) Method for Analysis of Caffeine, Theobromine, Theophylline

Caffeine is a natural stimulant and methylxanthine alkaloid. with the molecular formula C₆H₁₀N₄O₂. Caffeine can be found in a variety of plants, including tea, coffea, cocoa, kola nuts, and guarana. Ingestion of it can increase alertness and cognitive function. It can also cause worsening anxiety, heart palpitations, and headaches.

Theophylline is a methylxanthine compound with the molecular formula C₇H₈N₄O₂. It is a caffeine metabolite and is often used in forensic analyses. Besides analytical use, it is often used in treating respiratory conditions.

Theobromine is a methylxanthine alkaloid with the molecular formula C₇H₈N₄O₂. It is a derivative of caffeine. It can be found in many foods, including chocolate and tea. Despite that, it is toxic to dogs.

Caffeine, Theobromine, Theophylline can be retained and analyzed using the Primesep SB stationary phase column. The analysis utilizes an isocratic method with a simple mobile phase consisting of water and ethanol (EtOH) with a phosphoric acid buffer. Detection is performed using UV.

Column	Primesep SB, 3.2 x 100 mm, 5 µm, 100 A, dual ended
Mobile Phase	EtOH- 10%
Buffer	Phosphoric Acid
Flow Rate	0.5 ml/min
Detection	UV 270 nm

Class of Compounds	Alkaloid
Analyzing Compounds	Caffeine, Theobromine, Theophylline

For optimal results in HPLC analysis, it is recommended to measure absorbance at a wavelength that matches the absorption maximum of the compound(s) being analyzed. The UV spectrum shown can assist in selecting an appropriate wavelength for your analysis. Please note that certain mobile phases and buffers may block wavelengths below 230 nm, rendering absorbance measurement at these wavelengths ineffective. If detection below 230 nm is required, it is recommended to use acetonitrile and water as low UV-transparent mobile phases, with phosphoric acid and its salts, sulfuric acid, and TFA as buffers.
For some compounds, the UV-Vis Spectrum is affected by the pH of the mobile phase. The spectra presented here are measured with an acidic mobile phase that has a pH of 3 or lower.

HPLC Method for Theophylline, Theobromine, Caffeine on Primesep SB by SIELC Technologies

High Performance Liquid Chromatography (HPLC) Method for Analysis of Theophylline, Theobromine, Caffeine. Caffeine is a natural stimulant and methylxanthine alkaloid. with the molecular formula C₆H₁₀N₄O₂. Caffeine can be found in a variety of plants, including tea, coffea, cocoa, kola nuts, and guarana. Ingestion of it can increase alertness and cognitive function. It can also cause worsening anxiety, heart palpitations, and headaches. Theophylline is a methylxanthine compound with the molecular formula C₇H₈N₄O₂. It is a caffeine metabolite and is often used in forensic analyses. Besides analytical use, it is often used in treating respiratory conditions. Theobromine is a methylxanthine alkaloid with the molecular formula C₇H₈N₄O₂. It is a derivative of caffeine. It can be found in many foods, including chocolate and tea. Despite that, it is toxic to dogs. Theophylline, Theobromine, Caffeine can be retained and analyzed using the Primesep SB stationary phase column. The analysis utilizes an isocratic method with a simple mobile phase consisting of water and ethanol (EtOH) with a phosphoric acid buffer. Detection is performed using UV. You can find detailed UV spectra of caffeine and information about its various lambda maxima by visiting the following link. You can find detailed UV spectra of Theophylline and information about its various lambda maxima by visiting the following link. You can find detailed UV spectra of caffeine and information about its various lambda maxima by visiting the following link.

Condition

Column	Primesep SB, 3.2 x 100 mm, 5 µm, 100 A, dual ended
Mobile Phase	EtOH – 10%
Buffer	H3PO4 – 0.3%
Flow Rate	0.5 ml/min
Detection	UV, 270 nm

Description

Class of Compounds	Xanthine, Hydrophobic, Ionizable
Analyzing Compounds	Theophylline, Theobromine, Caffeine

 

Primesep B separates xanthines such as theobromine, dimethylxanthine, theophylline by a combination of reversed-phase and ion-exchange mechanisms. The anion-exchange properties of the column allow retention of hydrophilic xanthines, and the reversed-phase properties retain hydrophobic compounds. The HPLC separation uses a mobile phase of water, acetonitrile (MeCN, ACN) and either trifluoroacetic acid (TFA), perchloric acid (HClO4), or sulfuric acid (H2SO4) and UV detection.

Condition

Column	Primesep B
Mobile Phase	MeCN/H2O
Buffer	TFA , HClO4, H2SO4
Flow Rate	1.0 ml/min
Detection

 

Description

Class of Compounds	Drug, Acid, Hydrophilic, Ionizable, Hormone
Analyzing Compounds	Theobromine, 1,7 – Dimethylxanthine, Theophyline