We showed here in our graded-dose examination that serum Gla-OC levels increased significantly with intakes of 900 μg/day or more and serum ucOC levels and the UGR decreased significantly with intakes of 600 μg/day or more. Therefore, MK-4 supplementation at 600 μg/day after a week’s initial intake at 300 μg/day improved vitamin K status in terms of bone health, especially on the basis of serum ucOC level as a marker of vitamin K deficiency and the UGR as a sensitive marker of γ-carboxylation of osteocalcin.
Dietary vitamin K intake tends to be low in the British population  and to be less than the adequate intake in the DRIs in North America . Currently, the Japanese DRIs is 60 μg/day in females, and 75 μg/day in males, aged 18 to 29 years . However, in 2015 the Japanese DRIs will be revised upward to 150 μg/day for both males and females aged 18 to 29 years . The average dietary vitamin K intake in Japanese people in their 20s (201 μg/day)  satisfies the current and proposed future DRIs. In contrast, the dietary vitamin K intakes of some of our subjects were very much lower than the DRIs and the Japanese average dietary vitamin K intake. Notably, also, plasma phylloquinone and menaquinone-7 levels decreased during the study period, possibly reflecting decreases in dietary vitamin K intake on Days 15, 29, and 36, because dietary vitamin K consists mainly of phylloquinone (leafy green vegetables) and menaquinone-7 (fermented foods). The serum ucOC levels at baseline (Table 2) indicated that our subjects tended to have vitamin K deficiency, and an additional MK-4 intake of 600 μg/day or more was needed to reduce serum ucOC levels to close to the cut-off value (4.5 ng/mL) . Depending on dietary habits, therefore, intakes of vitamin K–rich food, or vitamin K supplementation, exceeding the DRIs may have been needed to improve bone health in our intervention study. Vitamin K status at baseline might affect γ-carboxylation of osteocalcin in response to supplementary vitamin K ; this could be one reason why our effective dose was lower than that in the report by Takeuchi et al.
. Those authors found that supplementation with MK-4 at 1500 μg/day for 4 weeks significantly improved vitamin K status in healthy males, but supplementation with MK-4 at 500 or 1000 μg/day did not produce a clear difference compared with placebo.
Our study had some limitations. Because it was a graded-dose study, each preceding, lower, dose may have affected the subsequent serum ucOC and Gla-OC levels and the UGR. Furthermore, because the assessment items for dietary vitamin K intake were limited to nine vitamin K–rich foods, the minimum dietary vitamin K intake was 0 and the dietary vitamin K intake tended to decrease during the study period. To make the dietary assessment more closely reflect the entire dietary vitamin K intake, in future we will have to select more sophisticated methods: for example, we may need to estimate each participant’s dietary intake of not only vitamin K but also other nutritional components [9, 10]. We will also need to investigate the lowest effective dose by measuring other bone metabolism parameters in a randomized, double-blind, placebo-controlled study that considers the dietary habits and genotypes of the subjects .
In conclusion, in this graded-dose study, we showed that supplementation with MK-4 at 600 μg/day or more is likely to be important in terms of vitamin K requirements for bone health. These relatively low MK-4 dosages (below 1500 μg/day) may improve bone formation and reduce bone fracture risk.
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