Unlabelled

Our goal was to determine total and directly measured free 25-hydroxy vitamin D (25(OH)D) serum levels in humans with a range of 25(OH)D levels and clinical conditions associated with low and high vitamin D binding protein levels. Serum samples and clinical data were collected from 106 subjects: 62 without cirrhosis or pregnancy, 24 cirrhotic patients with albumin <2.9g/dL, and 20 pregnant women. Total 25(OH)D (LC/MS/MS) and "free" 25(OH)D (immunoassay) were measured. Total 25(OH)D was significantly lower in liver disease patients but free 25(OH)D concentrations were significantly higher in this group (p<.001). Neither total nor free 25(OH)D concentrations were significantly different in pregnant women vs. the comparator group. There were significant direct positive relationships between free 25(OH)D and total 25(OH)D concentrations for the entire dataset and for each group (p<.0001), however slopes of relationships differed in the cirrhotic group compared to pregnant women or the comparator group. In cirrhotics: y (free 25(OH)D)=2.52+0.29×X(total 25 (OH)D), r(2)=.51, p<.001; y=1.45+0.09×X; r(2)=.77, p<.0001 for pregnant women; and y=1.11+0.12×X; r(2)=.72, p<.0001 for the comparator group).

Conclusions

directly measured free 25(OH)D serum concentrations and relationships between total and free 25(OH)D vary with clinical conditions, and may differ from those predicted by indirect estimation methods. This article is part of a Special Issue entitled 'Vitamin D Workshop'.

Objective

Our goal was to compare direct quantitation of circulating free 25-hydroxyvitamin D (25(OH)D)levels to calculated free 25(OH)D levels and their relationships to intact PTH (iPTH), a biomarker of 25(OH)D effect, in humans with a range of clinical conditions.

Patients and methods

Serum samples and clinical data were collected from 155 people: 111 without cirrhosis or pregnancy (comparison group), 24 cirrhotic patients with albumin <2.9 g/dL, and 20 pregnant women (second and third trimester). Total 25(OH)D (LC/MS/MS), free 25(OH)D (immunoassay), vitamin D binding protein (DBP) (immunoassay), albumin, and iPTH (immunoassay) were measured.

Results

Total 25(OH)D, DBP, and albumin were lowest in patients with cirrhosis, but measured free 25(OH)D was highest in this group (P < .001). DBP was highest in pregnant women (P < .001), but measured free 25(OH)D did not differ from the comparison group. Calculated free 25(OH)D was positively correlated with measured free 25(OH)D (P < .0001) but explained only 13% of the variability with calculated values higher than measured. African Americans had lower DBP than other ethnic populations within all clinical groups (P < .03), and differences between measured and calculated free 25(OH)D were greatest in African Americans (P < .001). Measured free 25(OH)D was correlated with total 25(OH)D (P < .0001; r(2) = 0.51), but calculated free 25(OH)D was not. Similarly, both measured free 25(OH)D (P < .02) and total 25(OH)D (P < .05) were correlated with iPTH, but calculated free 25(OH)D was not.

Conclusions

Calculated free 25(OH)D levels varied considerably from direct measurements of free 25(OH)D with discrepancies greatest in the data for African Americans. Differences in DBP binding affinity likely contributed to estimation errors between the races. Directly measured free 25(OH)D concentrations were related to iPTH, but calculated estimates were not. Current algorithms to calculate free 25(OH)D may not be accurate. Further evaluation of directly measured free 25(OH)D levels to determine its role in research and clinical management of patients is needed.