ENCAPSULATION OF HEMOGLOBIN IN A BICONTINUOUS CUBIC PHASE LIPID

The effects of encapsulating bovine hemoglobin (BHb) in the bicontinuo us cubic phase formed by monooleoylglycerol and water was investigated with Fourier transform infrared (FTIR) spectroscopy and X-ray diffrac tion. Cubic phase was formed in the presence of 1-10 wt% BHb. Studies using X-ray diffraction reveal that at 0.5-2.5 wt% BHb, the cubic phas e structure is characterized by the double diamond lattice (Pn3m). At 2.5-5 wt% BHb, coexistence of two cubic phase structures, Pn3m and the gyroid lattice (Ia3d), was observed while at BHb concentrations highe r than 5 wt% the gyroid structure persists. FTIR shows there is an inc rease in intensity of the free upsilon C = 0 (1745 cm(-1)) and a corre sponding decrease in the intensity of the hydrogen bonded upsilon C = 0 (1720 cm(-1)) as the BHb concentration is increased. The upsilon C-O -CO peak shifts from 1183 cm(-1) to 1181 cm(-1) as the concentration o f BHb is raised from 2.5 to 10 wt% indicating BHb may induce subtle ch anges in the interfacial region of cubic phase monoolein. The bandwidt h of the upsilon(as)CH(2) stretch (2926 cm(-1)) increased in the prese nce of 5 wt% BI-IT, compared to samples with 2.5 or 10 wt% BHb. The in crease in frequency of the upsilon(s)CH(2) stretch (2854 cm(-1)) induc ed by increasing temperature 20 to 60 degrees C was dampened when BHb was present compared to samples heated in isotonic buffer. Analysis of the amide I band at 1650 cm(-1) showed that the secondary structure o f BHb is not affected by encapsulation in monoolein. In vitro release studies showed that 45% of the entrapped BHb was released after 144 h at 37 degrees C. The porous nature of bulk cubic phase was further dem onstrated by diffusion of K2Fe(CN)(6) and conversion of 73% of the oxy hemoglobin to methemoglobin after 1 h. These results suggest that the cubic phase may be useful for encapsulation of Hb as a red cell substi tute and for the encapsulation and delivery of other bioactive agents.