Abstract

Water-phospholipid (dimyristoylphosphatidylcholine) interaction was analyzed in a water-in-oil(benzene) reversed micellar system using Fourier transform infrared spectroscopy, and the effects of inhalation anesthetics (halothane, enflurane, chloroform, and carbon tetrachloride) on the interaction were studied. The O-H stretching frequency, representing water, increased from 3369 cm-1 to a steady 3430 cm-1 when the water/phospholipid mole ratio exceeded 18. The value did not quite reach the frequency of free water of 3490 cm-1 at the water/phospholipid mole ratio of 30. The O-H bending frequency of water did not appear until the water/phospholipid mole ratio exceeded 9. The P=O stretching frequency in the polar head group of unhydrated dimyristoylphosphatidylcholine was 1262 cm-1 and decreased with the addition of water, reaching a steady value of 1238 cm-1 at the water/phospholipid mole ratio of 9. However, the (CH3)3N+ stretching of the choline head, as well as the C-H stretching of the hydrocarbon tail and the C=O stretching of the ester linkage, showed little change by the addition of water. The present results suggest that the primary hydration site of dimyristoylphosphatidylcholine is the phosphate moiety, and up to 18 water molecules are restricted at the polar head group. Apparently, the choline head has a minor role in the hydration of phospholipids despite the positive electrostatic charge. Among the water molecules interacting with the phospholipid head group, about 9 water molecules are strongly bound. The water content in the micelles correlated linearly with the ratio of the absorbance band area between O-H and C=O stretching. The addition of polar anesthetics (halothane, enflurane, and chloroform) increased the O-H stretching frequency and elevated the ratio of the absorbance band area between O-H and C=O stretching, implying that the anesthetics released the structured water molecules bound at the phospholipid-water interface. The anesthetics disrupted the hydrogen bond between the phosphate moiety of the phospholipid and water. Although apolar carbon tetrachloride also released bound water molecules, the magnitude was less than that of the polar anesthetics, as expected. The anesthetics did not affect the C-H stretching or C=O stretching bands, indicating that the disordering action upon the hydrocarbon core of phospholipid membranes is minimal at low water content. These results support our view that the primary site of action of inhalation anesthetics is the membrane-water interface, releasing bound water molecules.