Kinetics of hypericin association with low-density lipoproteins

Steady-state and time-resolved fluorescence spectroscopy have been used for the study of the incorporation kinetics of hypericin (Hyp) into low-density lipoproteins (LDL). Biphasic kinetics of Hyp association with LDL was observed when solutions of Hyp and LDL were mixed at various concentration ratios. The rapid phase of Hyp incorporation is completed within seconds, while the slow phase lasts several minutes. The relative contributions of the individual phases show that a higher amount of Hyp molecules (65%) are incorporated into LDL in the second phase. The kinetics of the incorporation of Hyp into LDL particles preloaded with Hyp (Hyp/LDL=25:1) was also investigated. The decreased intensity of Hyp fluorescence is a sign of the formation of Hyp aggregates after penetration of additional Hyp molecules into Hyp/LDL=25:1 complex. The time dependence of Hyp fluorescence was measured after mixing the complex Hyp/LDL =200:1 with appropriate amounts of free LDL molecules. For each final Hyp/LDL ratio, an increase in the intensity and lifetime of Hyp fluorescence was observed, suggesting a monomerization of Hyp aggregates. The half-time of Hyp transfer from Hyp/LDL complex to LDL particles is similar to the half-time of the slow phase of Hyp incorporation into free LDL particles.