Synthesis of vitamin D3 analogues with A-ring modifications to directly measure vitamin D levels in biological samples

C-3-substituted 25-hydroxyvitamin D-3 analogues were synthesized as tools to directly measure levels of vitamin D in biological samples. The strategy involves vinyloxycarbonylation of the 3 beta-hydroxy group and formation of a carbamate bond with a hydroxyl or amino group at the end of the alkyl chain. Biotinylated conjugates of synthesized derivatives were generated to be linked with vitamin D binding protein (DBP). The spacer group present in the alkyl chain is important in the binding of antibodies to the analogue-DBP complex. When compared to 25-hydroxyvitamin D-3-DBP, the binding of some antibodies to the analogue-DBP complex of the 25-hydroxyvitamin D-3 derivative 10 that posses an 8-aminoctyl alkyl chain is significantly reduced, but this analogue displaced [26,27-H-3]-25-hydroxyvitamin D-3 from DBP. In contrast, the 8-hydroxyoctyl alkyl chain analogue 9 showed less displacement. (C) 2013 Elsevier Ltd. All rights reserved.