Characterisation of vitamin B12 immunoaffinity columns and method development for determination of vitamin B12 in a range of foods, juices and pharmaceutical products using immunoaffinity clean-up and high performance liquid chromatography with UV detection. - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Food Additives and Contaminants Année : 2009

Characterisation of vitamin B12 immunoaffinity columns and method development for determination of vitamin B12 in a range of foods, juices and pharmaceutical products using immunoaffinity clean-up and high performance liquid chromatography with UV detection.

Eric Mackay
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Gemma Young
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Résumé

New rapid and simpler procedures using immunoaffinity columns, have been developed for determination of vitamin B12 in a range of samples including 3 different US National Institute of Standard and Technology (NIST) Reference Materials, infant formula, powdered energy drinks and bars, wheat breakfast cereal, carbonated soft drinks, fruit juices and vitamin B12 tablets. The procedures involved extraction of vitamin B12 using water or sodium acetate buffer and enzyme digestion (using pepsin or α-amylase or both) if necessary. The extract was clarified and passed through EASI-EXTRACT® Vitamin B12, an immunoaffinity column containing monoclonal antibody with high affinity and specificity to vitamin B12. Subsequently the vitamin B12 immunoaffinity column was washed with 10 ml water and the vitamin B12 was released from the immunoaffinity column with 3 ml methanol. Following evaporation the samples were reconstituted in mobile phase and analysed by HPLC-UV at 361 nm on an ACE 3AQ analytical column using a gradient elution consisting of 0.025% trifluoroacetic acid in water and acetonitrile. Analysis of 3 types of NIST Standard Reference Materials in triplicate demonstrated the results of the immunoaffinity column method were comparable to the microbiological assay results. Method repeatability data was determined for all samples analysed and ranged between 0.8-10% demonstrating the method was repeatable with complex matrices (NIST 2383) containing low level vitamin B12 (0.44 mg/100 g) as well as simpler matrices such as vitamin tablets containing high levels (2000 mg/0.849 g) of vitamin B12.

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Dates et versions

hal-00577324 , version 1 (17-03-2011)

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Elaine Catherine Marley, Eric Mackay, Gemma Young. Characterisation of vitamin B12 immunoaffinity columns and method development for determination of vitamin B12 in a range of foods, juices and pharmaceutical products using immunoaffinity clean-up and high performance liquid chromatography with UV detection.. Food Additives and Contaminants, 2009, 26 (03), pp.282-288. ⟨10.1080/02652030802429104⟩. ⟨hal-00577324⟩

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