Deuterium-labeled phylloquinone has tissue-specific conversion to menaquinone-4 among Fischer 344 male rats

Abstract

Phylloquinone (PK) is converted into menaquinone-4 (MK-4) via side chain removal-addition. Stable isotope use is an effective approach to identify the tissue location of this conversion, which is currently unknown. Following a 14-d PK-deficient diet, male Fischer 344 rats (8 mo; n = 15) were fed 1.6 mg deuterium-labeled PK (L-PK) per kg diet for 0 (control), 1 d (PK-1d), and 7 d (PK-7d). Both L-PK and deuterium-labeled MK-4 (L-MK-4) were detected in tissues in PK-1d and PK-7d, although the results varied. Whereas some tissues had an overall increase in MK-4 in response to L-PK, total brain, testes, and fat MK-4 concentrations did not. In contrast, L-MK-4 concentrations increased in all 3 tissues. The deuterium label was found only on the L-MK-4 naphthoquinone ring, confirming the need for side chain removal for the formation of MK-4. Labeled menadione (MD) was detected in urine and serum in PK-1d and PK-7d, confirming its role as an intermediate. A Caco-2 cell monolayer model was used to study the role of the enterocytes in the conversion process. Neither MK-4 nor MD was detected in Caco-2 cells treated with PK. However, when Caco-2 cells were treated with MD, MK-4 was formed. Similarly, MK-4 was formed in response to MD-treated 293T kidney cells, but not HuH7 liver cells. These data demonstrate that MK-4 is the predominant form of vitamin K in multiple tissues, but there appears to be a tissue-specific regulation for the conversion of PK to MK-4.

FIGURE 1

Total (A) and unconjugated (B) MD concentrations in urine of untreated male Fischer 344 rats and those fed 1.6mg L-PK /kg diet for 1 or 7d. Urine total MD concentrations include both conjugated and unconjugated forms. Values are means ± SD, n = 5. Labeled means for a variable without a common letter differ, P < 0.017 (Wilcoxon rank sum test). C, control; L-PK, deuterium-labeled phylloquinone; MD, menadione; PK-1d, fed 1 d of 1.6 mg L-PK/kg diet; PK-7d, fed 7 d of 1.6 mg L-PK/kg diet.