Enhancement of chemiluminescence for vitamin B12 analysis

• Published in: Analytical biochemistry 2009-05, Vol.388 (2), p.312-316
• Main Authors: Kumar, Sagaya Selva, Chouhan, Raghuraj Singh, Thakur, Munna Singh
• Format: Article
• Language: English
• Subjects: Hydrogen Peroxide - chemistry; Hydrogen-Ion Concentration; Luminescent Measurements - instrumentation; Luminescent Measurements - methods; Luminol - chemistry; Vitamin B 12 - analysis
• ISSN: 0003-2697; 1096-0309
• DOI: 10.1016/j.ab.2009.02.029

In the current article, chemiluminescence (CL) from the vitamin B(12) and luminol reaction was studied under alkaline conditions to develop a sensitive analytical method for vitamin B(12) using the carbonate enhancement effect. The method was successfully applied to the determination of vitamin B(12) in vitamin B(12) tablets, multivitamin capsules, and vitamin B(12) injections. Experimental parameters were optimized, including luminol concentration, urea-hydrogen peroxide (urea-H(2)O(2)) concentration, effect of pH, and sequence of addition of reactants for obtaining maximum CL, which was not explored previously. The limit of detection was 5 pg/ml, and the linear range was 10 pg/ml to 1 microg/ml with a regression coefficient of R(2)=0.9998. The importance of these experimental parameters and the carbonate enhancement effect is discussed based on the knowledge of the mechanism of oxidation of luminol and decomposition of urea-H2O2 in the presence of vitamin B(12). Extraction of vitamin B(12) was carried out, and the observed recovery was 97-99.2% with a relative standard deviation in the range of 0.30-1.09%. The results obtained were compared with those of the flame atomic absorption spectrometry method.