There is some evidence for a nutritional interaction between vitamin D and vitamin K status. We have recently reported that serum percentage undercarboxylated osteocalcin (%ucOC; a marker of vitamin K status) was inversely correlated with serum 25-hydroxyvitamin D (25(OH)D) concentration (reflective of vitamin D status) in healthy Danish girls (aged 11–12 years), in line with a similar relationship reported in elderly women. While the causal nature of the relationship between vitamin D status and serum %ucOC has been tested in studies of elderly women, it has not been investigated in children. The objective of the present study was to test the hypothesis that improving vitamin D status significantly lowers serum %ucOC. Serum samples from sixty-seven healthy Danish girls (aged 11–12 years), who participated in a 12-month double-blind, placebo-controlled, vitamin D3 intervention trial were used for the present study. These girls were a subset of subjects which began and finished the intervention during wintertime, thus avoiding the influence of seasonality on vitamin D status. A total of thirty-three and thirty-four of the girls had been randomised to treatment with 10 μg vitamin D3 per d and placebo, respectively, for 12 months. Total osteocalcin and the fraction of ucOC in serum (via enzyme-immunoassay) as well as serum 25(OH)D (via HPLC) were assessed at baseline and end-point. Vitamin D3 supplementation significantly increased serum 25(OH)D (21·6 %; P < 0·002) but had no effect on serum %ucOC (P>0·8). In conclusion, the findings of the present intervention study in young girls suggest that vitamin D supplementation does not affect serum %ucOC, a marker of vitamin K status.

Vitamin K contributes to bone health, probably through its role as cofactor in the carboxylation of osteocalcin. Intervention studies in adults have demonstrated that markedly higher osteocalcin carboxylation is obtained by intakes of vitamin K well above the current recommended dietary intake. However, the relationship between increased vitamin K2 intake and enhanced osteocalcin carboxylation has never been shown in healthy children. The objective was to study the effect of 45 μg menaquinone-7 (MK-7; one of the vitamin K2 species) on the circulating levels of undercarboxylated osteocalcin (ucOC) and carboxylated osteocalcin (cOC) in healthy prepubertal children. We hypothesised that MK-7 supplementation will reduce the ucOC:cOC ratio (UCR), indicating an improved vitamin K status. The present study is a double-blind randomised placebo-controlled trial examining the effect of 8 weeks MK-7 supplementation on the carboxylation of osteocalcin in healthy children (n 55). Serum levels of ucOC, cOC and MK-7 were measured at baseline and after 8 weeks, together with bone markers and coagulation parameters. The UCR was used as an indicator of vitamin K status. In the MK-7-supplemented group (n 28), the circulating concentration of inactive ucOC reduced and the UCR improved whereas the concentration of MK-7 increased. Within the placebo group, ucOC, cOC, UCR and MK-7 did not significantly change over time. In both groups, bone markers and coagulation parameters remained constant over time. These findings demonstrate that in healthy, prepubertal children, modest supplementation with MK-7 increases circulating concentrations of MK-7 and increases osteocalcin carboxylation.

Vitamin K acts as a cofactor during the γ-carboxylation of vitamin K-dependent proteins. Undercarboxylated osteocalcin (ucOC) is a suggested biomarker of vitamin K status. The +2255 polymorphism of the vitamin K epoxide reductase gene (VKORC1) was shown to be associated with the recycling rate of the active form of vitamin K. We investigated the association between dietary vitamin K intake and serum ucOC and hypothesized that this association might vary by VKORC1 genotype. ucOC and total intact osteocalcin (iOC) concentrations were quantified using specific ELISA tests in serum samples of 548 male and female participants (aged 18–81 years) of the Bavarian Food Consumption Survey II. ucOC was expressed relative to iOC (ucOC/iOC ratio). Dietary intake of vitamin K (phylloquinone and menaquinones) was estimated from three 24 h dietary recalls using previously published food composition data. The association between dietary vitamin K intake and ucOC/iOC ratio was analysed using linear and non-linear regression models. Median intakes of phylloquinone/menaquinones were 83·4/37·6 μg/d in men and 79·6/29·8 μg/d in women, respectively. As expected, vitamin K intake was significantly inversely associated with the ucOC/iOC ratio. The ucOC/iOC ratio differed significantly across variants of the +2255 polymorphism in the VKORC1 gene. Stratification by VKORC1+2255 genotype revealed that only in carriers of the GG genotype (39 % of all participants) did the ucOC/iOC ratio significantly decrease with increasing intake of vitamin K. Thus, the results show that the inverse association between dietary vitamin K intake and serum ucOC depends on a functionally relevant allelic variant of the VKORC1 gene.

In adult bone, vitamin K contributes to bone health, probably through its role as co-factor in the carboxylation of osteocalcin. In children, the significance of vitamin K in bone-mass acquisition is less well known. The objective of this longitudinal study was to determine whether biochemical indicators of vitamin K status are related to (gains in) bone mineral content (BMC) and markers of bone metabolism in peripubertal children. In 307 healthy children (mean age 11·2 years), BMC of the total body, lumbar spine and femoral neck was determined at baseline and 2 years later. Vitamin K status (ratio of undercarboxylated (ucOC) to carboxylated (cOC) fractions of osteocalcin; UCR) was also measured at both time points. Markers of bone metabolism, sex steroids, vitamin D status and growth hormones were measured at baseline only. Large variations in the levels of the UCR were found at both time-points, indicating a substantial interindividual difference in vitamin K status. Improvement of vitamin K status over 2 years (n 281 children) was associated with a marked increase in total body BMC (r − 49·1, P < 0·001). The UCR was associated with pubertal stage, markers of bone metabolism, sex hormones and vitamin D status. A better vitamin K status was associated with more pronounced increase in bone mass in healthy peripubertal children. In order to determine the significance of these findings for childhood bone health, additional paediatric studies are needed.

Recent cross-sectional data suggest that better vitamin K status in young girls (aged 3–16 years) is associated with decreased bone turnover, even though it is not associated with bone mineral content (BMC). The objective of the present study was to investigate the relationship between serum percentage of undercarboxylated osteocalcin (%ucOC), as an index of vitamin K status, and BMC and biochemical indices of bone turnover in peri-pubertal Danish girls. This peri-pubertal stage is a dynamic period of bone development, and as such, may represent an important window of opportunity for vitamin K status to modulate childhood bone health. Serum %ucOC and serum 25-hydroxyvitamin D (25 (OH) D) were measured at baseline in a study of 223 healthy girls aged 11–12 years. Urinary pyridinium crosslinks of collagen and serum total osteocalcin as markers of bone resorption and formation, respectively, as well as BMC (total body and lumbar spine) were also measured. Serum %ucOC (median 21·9 %) was not associated with markers of bone resorption or with total osteocalcin. Serum %ucOC was inversely correlated with serum 25 (OH) D (r − 0·143; P < 0·05). Serum %ucOC was negatively associated with BMC of the total body (β − 0·045; P < 0·001) and lumbar spine (β − 0·055; P < 0·05), after adjustment for potential confounders including vitamin D status. Better vitamin K status was associated with increased BMC, but not bone turnover, in healthy peri-pubertal Danish girls. There is a need for well-designed, randomized phylloquinone supplementation trials in children and adolescents to confirm epidemiological findings of an association between vitamin K status and bone health.

Low phylloquinone (vitamin K1) intakes have been associated with low bone mineral density in older adults. Phylloquinone intakes and serum undercarboxylated osteocalcin (ucOC) levels were assessed in ninety-seven apparently healthy, free-living Irish women aged 50–75 years. Phylloquinone intakes were estimated using a detailed dietary history, which measured habitual food intakes from a typical 14 d period, and recently published food composition data for phylloquinone. Fasting serum ucOC was measured using an enzyme immunoassay. The median daily intake of phylloquinone in the group from all sources was 108·8 mg and from food sources only was 106·6 mg, indicating that approximately 99% of the phylloquinone came from food. Vegetables and vegetable dishes contributed 67% of the total phylloquinone intake, but further analysis showed that broccoli, cabbage and lettuce were the primary sources, making a total contribution of 44 %. Twenty per cent of the women had a phylloquinone intake below the UK recommendation of 1μg/kg body weight per day and 34% failed to meet the US Adequate Intake value of 90mg/day. Mean serum ucOC levels in the women were 6·2 (SD 1·7) ng/ml and were predicted by phylloquinone intake (β— 22·20, generated from log-transformed phylloquinone intake data; P=4 0·04). On the basis of comparisons with both UK recommendations and US A equate Intakes for phylloquinone, the habitual intakes of phylloquinone in a high proportion of Irish postmenopausal women may not be adequate.

The vitamin K concentration in the circulation and the availability of vitamin K to bone may be affected by factors influencing lipoprotein metabolism, such as apoE genotype. The relationships between markers of vitamin K status, bone mineral content and apoE genotype were studied in healthy older men and women aged 60–83 years, 177 from Shenyang, China, and 132 from Cambridge, UK. Fasting plasma was analysed for vitamin K1, triacylglycerol, total osteocalcin, undercarboxylated osteocalcin (ucOC) and apoE genotype. Hip bone mineral content was measured using dual-energy X-ray absorptiometry. Subjects were grouped according to apoE genotype as E2/3, E3/3 and [E3/4+E4/4]. The mean plasma vitamin K1 concentration of the three genotype groups was significantly higher and the percentage ucOC was lower in the Chinese than in the British subjects (P<0·01). A higher vitamin K1 concentration was found in subjects with [E3/4+E4/4] than those with either E2/3 or E3/3 in Cambridge (32·2 (SE 14·6) %, P=0·03; 24·6 (SE 10·7) %, P=0·02). Similar trends were observed although were not statistically significant in Shenyang (26·5 (18·9) %, P=0·16; 23·1 (13·0) %, P=0·08). Subjects with [E3/4+E4/4] had a lower percentage ucOC (total osteocalcin adjusted) than did those with either E2/3 or E3/3 in Shenyang (65·1 (27·2) %, P=0·02; 49·6 (19·9) %, P=0·01 respectively) but not in Cambridge. This study demonstrates that a superior vitamin K status is associated with the apoE4 genotype in healthy older individuals from China and the UK.

To explore whether differences in vitamin K nutrition might, at least in part, underlie differences in fracture incidence between Asian and European populations, the vitamin K status of older individuals in Shenyang, China (eighty-six men, ninety-two women) and in Cambridge, UK (sixty-seven men, sixty-seven women) was compared. Dietary information was collected by food questionnaire in Shenyang and food diary in Cambridge and used to estimate the intake and sources of phylloquinone. Fasting blood was analysed for phylloquinone, triacylglycerol, total osteocalcin (tOC) and undercarboxylated osteocalcin (expressed as percentage of tOC; %ucOC). The mean intakes of green leafy vegetables were 127 (SD 90) g/d in Shenyang and 39 (SD 48) g/d in Cambridge. The estimated phylloquinone intakes (geometric means) were 247 (95% CI 226, 270) μg/d in Shenyang and 103 (95% CI 94, 112) μg/d in Cambridge. Plasma phylloquinone concentrations (geometric means) were significantly higher in the Shenyang subjects (2·17 (95% CI 1·95, 2·42) nmol/l) than in the Cambridge subjects (0·69 (95% CI 0·63, 0·76) nmol/l; P<0·001). Plasma phylloquinone concentration was positively related to phylloquinone intake in both the Shenyang (coefficient 0·17 (SE 0·08); P=0·03) and Cambridge subjects (coefficient 0·29 (SE 0·10); P=0·005). tOC concentration and %ucOC (after adjusting for tOC) were significantly lower in the Shenyang than in the Cambridge subjects (tOC 25·2 (SE 4·2) % and %ucOC 68·5 (SE 10·0) % lower respectively; P<0·001). After adjusting for tOC and triacylglycerol, %ucOC was negatively related to plasma phylloquinone concentration in both the Shenyang (coefficient −0·41 (SE 0·11); P=0·0003) and Cambridge subjects (coefficient −0·17 (SE 0·07); P=0·02). The present study demonstrates that older individuals in northern China have a better vitamin K status compared with their British counterparts in Cambridge, UK.