FTIR studies of vitamin D2-model membrane interactions.

Vitamin D2 (ergocalciferol) is a lipid soluble vitamin which does not occur naturally. It is obtained by irradiation of the provitamin ergosterol from yea@ which have ergosterol as their sterol component. It plays an important role in calcium and phosphorus metabolism, in immune function and cell differentiation [ 11 and recently it has been proposed as a membrane antioxidant [2]. Thm have been no studies on the interaction of vitamin D2 with model and native membranes. In the present study interaction of high concentration of vitamin Dz (12 mol % ) with dipalmitoylphosphatidylcholiie (DPPC) model membranes has been investigated by Fourier Transform Infrared Spectroscopy in the C-H stretching region. Ergocalciferol and DPPC were purchased from Sigma ( St. Louis, Mo). Phospholipid multilamellar liposomes were prepared according to the procedure reported in Severcan et al 131. Infrared spectra were obtained using a NICOLET 510 FT-IR spectrometer and setting was reported in Severcan et al.[3]. Temperature was regulated by a Unicam Specac digital temperature contmller unit. The infrared spectra of DPPC liposomes shows three bands in the C-H region. The strong bands appear around 2920 cm-' and 2850 cm-' correspond to the CH, antisymmetric and symmetric stretchig modes of acyl chains of the phospholipid, respectively. The minor band at around 2956 cm-l are due to the asymmetric stretching mode of terminal CH3 groups. Fig. 1 A shows the temperature dependence of the frequency of the CH2 antisymmetric stretching modes of DPPC liposomes in the absence and presence of 12 mol % vitamin D2 With the inclusion of vitamin D2 the main phase transition is completely diminished and phase transition curve remarkably broadens which implies the loss of cooperativity between the lipid chains of DPPC. An abrupt decrease in the fkquency is observed especially in the liquid crystalline phase which indicates a decrease in the number of gauche conformers (ordering) [4-61. Temperature dependence of the bandwidths of the CH, antisymmetric stretching modes of DPPC multibilayers in the absence and in the psence of 12 mol % vitamin D2 at 0.75 x peak height position are shown in Fig. 1 B. The bandwidth monitors the freedom of motion of the absorbing group i.e. amplitudes and rates of motion within its immediate environment [4]. The bandwidths decrease with the addition of vitamin D, implying that vitamin D, stabilizes phospholipid membranes by decreasing the mobility of the acyl chains. The band at 2956 cm'l provides a monitor of the center of bilayer [5]. Temperature dependence of the frequencies of the CH3 asymmetric stretching mode of DPPC liposomes in the absence and presence of vitamin D2 are depicted in Fig. 2., the fresuency considerably decreases with the inclusion of vitamin D, indicating that vitamin Dz decreases the dynamics of the deep interior of the bilay er. In conclusion the addition of high concentration of vitamin 4 in the DPPC liposomes alters the shape of phase transition profile with a significant brodening of the phase transition curve. It increases the order and decreases the fluidity of the DPPC ' 0 DPPC+IPMd % W t D2 c 2924 t * I '3 1 2922