Fluorescence Resonance Energy Transfer Between Acridine Orange and Rhodamine B and Analytical Application on Determination of Vitamin B₁₂

Abstract

A new method to determine Vitamin B₁₂ by applying energy transfer fluorescence quenching of Acridine orange (AO)–Rhodamine B (RB) has been studied and established. It was found that the effective energy transfer could occur between AO and RB at λ_ex/λ_em=470/575 nm in the dodecyl benzene sodium sulfonate (DBS) and Na₂HPO₄–citric buffer (pH=7.0). The fluorescence intensity of RB increased sharply. Vitamin B₁₂ diminished the fluorescence intensity of RB depending on the isochromacticity between them. The detection limit of the method was up to 1.5×10⁻⁷ mol L⁻¹. The range of determining concentration of Vitamin B₁₂ was 2.5×10⁻⁷∼8.0×10⁻⁶ mol L⁻¹. The relative standard deviation was 1.93% for six parallel determinations of Vitamin B₁₂. The recoveries of Vitamin B₁₂ were 97∼101%. The method was applied to the determination of Vitamin B₁₂ in injection.

Keywords:

• Acridine orange
• rhodamine B
• energy transfer
• fluorescence quenching
• Vitamin B₁₂

The Natural Science Foundations of China and Hebei Province, and a Visiting Scholar Foundation of the Key Laboratory at Hebei University supported this work. Thank you for the help of Professors Yang and Liu.