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Factors affecting HPLC analysis of vitamin K in serum, food and dietary supplements
Author(s) -
Craft Neal E,
Park Heasoon
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.996.18
Subject(s) - chromatography , chemistry , high performance liquid chromatography , hexane , vitamin , elution , sample preparation , biochemistry
Vitamin K (VK) is one of 4 fat‐soluble vitamins essential for humans. It is most well‐known for its role in blood clotting. Recently, VK was shown to be involved in bone growth. It presents a difficult challenge to measure in blood and food due to its low level and lack of a specific chromaphore. Aim To examined the effects of sample preparation, HPLC separation and detection on the measurement of K1, K2, and MK‐7 in blood and dietary sources. Normal‐phase (NPLC) and reversed‐phase (RPLC) methods were tested. In RPLC, detection was compared using UV and post‐column reduction with fluorescence (PCFL). Reduction modes included: Pt, Zn, and coulometric. Results NPLC required gradient elution, failed to resolve all components, and was not amenable to PCFL detection. PCFL was >10x more sensitive than UV detection. Zn and Pt reduction were equally effective but Zn was more practical. Food and supplements were extracted with DMSO then partitioned into hexane. Blood serum was precipitated with alcohol and extracted with hexane, then loaded on Si SPE cartridges. The recovery of K1 or K2 spiked into samples was nearly 100%. Acceptable method performance in serum samples was verified through the KEQAS program.