难溶性药物的胶束给药系统(MDDS)在其赋形剂的安全性方面比替代制剂具有很大的优势,储存稳定性,简单的生产。一个典型的例子是甘氨胆酸盐(GC)和卵磷脂的混合胶束,人体血液中的两种内源性物质。限制MDDS使用的是注入后转换的复杂性。特别是,由于MDDS在注射后部分或完全分解,药物必须安全地转移到血液中的内源性载体上,例如人血清白蛋白(HSA)。如果这次转移受到损害,该药物可能会沉淀-在任何情况下都需要排除这一过程。本文的关键问题是,在MDDS溶解的时刻和位点,GC的高局部浓度是否会暂时饱和HSA结合位点,因此,危及药物快速转移。为了解决这个问题,我们使用了一种新方法,这是HSA中单一色氨酸的时间分辨荧光光谱,Trp-214,以表征GC和药物替代品苯胺基萘磺酸酯(ANS)与HSA的竞争性结合。Trp-214的时间分辨荧光相对于已建立的解决该问题的方法显示出重要的优势。几十年来,ANS一直是研究白蛋白结合的标准“模型药物”,考虑到它与含萘的酸性药物的结构相似性,以及它被许多药物(可能与相同位点结合)从HSA中取代的事实。我们的复杂全局拟合使用了平均寿命表现类似于单个寿命的临界近似,但由此产生的误差被发现是适度的,结果提供了令人信服的解释,乍一看,违反直觉的行为。因此,基本上与文献一致,我们观察到两种类型的位点结合ANS在HSA:3型A,相当外围,和2个B型,可能更多的中央网站。后者通过Förster共振能量转移(FRET)淬灭Trp-214,每个ANS的速率常数约为0.4ns-1。添加毫摩尔浓度的GC将ANS从A位点置换而不是从B位点置换。在不完全ANS饱和时,这导致GC诱导的ANS从A转移到更有FRET活性的B位点。这导致明显的悖论,即ANS从HSA的部分位移增加了其对Trp-214的猝灭作用。最重要的结论是,(类ANS)药物不能从B型位点移位,因此,在溶解胶束附近,GC的竞争性结合不会损害药物向这些位点的转移。第二个结论是,对于高于CMC(9mM)的未结合GC,ANS在HSA和GC胶束之间平衡,但强烈偏爱HSA上的游离位点。这意味着,一旦暴露于HSA,即使持续存在的胶束也容易失去它们的货物。对于共享此属性的所有MDDS,涉及它们作为纳米载体的靶向药物递送方法将是毫无意义的。
Micellar drug delivery systems (MDDS) for the intravenous administration of poorly soluble drugs have great advantages over alternative formulations in terms of the safety of their excipients, storage stability, and straightforward production. A classic example is mixed micelles of glycocholate (GC) and lecithin, both endogenous substances in human blood. What limits the use of MDDS is the complexity of the transitions after injection. In particular, as the MDDS disintegrate partially or completely after injection, the drug has to be transferred safely to endogenous carriers in the blood, such as human serum albumin (HSA). If this transfer is compromised, the drug might precipitate─a process that needs to be excluded under all circumstances. The key question of this paper is whether the high local concentration of GC at the moment and site of MDDS dissolution might transiently saturate HSA binding sites and, hence, endanger quick drug transfer. To address this question, we have used a new approach, which is time-resolved fluorescence spectroscopy of the single tryptophan in HSA, Trp-214, to characterize the competitive binding of GC and the drug substitute anilinonaphthalenesulfonate (ANS) to HSA. Time-resolved fluorescence of Trp-214 showed important advantages over established methods for tackling this problem. ANS has been the standard \"model drug\" to study albumin binding for decades, given its structural similarity to the class of naphthalene-containing acidic drugs and the fact that it is displaced from HSA by numerous drugs (which presumably bind to the same sites). Our complex global fit uses the critical approximation that the average lifetimes behave similarly to a single lifetime, but the resulting errors are found to be moderate and the results provide a convincing explanation of the, at first glance, counterintuitive behavior. Accordingly, and largely in line with the literature, we observed two types of sites binding ANS at HSA: 3 type A, rather peripheral, and 2 type B, likely more central sites. The latter quench Trp-214 by Förster Resonance Energy Transfer (FRET) with a rate constant of ≈0.4 ns-1 per ANS. Adding millimolar concentrations of GC displaces ANS from the A sites but not from B sites. At incomplete ANS saturation, this causes a GC-induced translocation of ANS from A to the more FRET-active B sites. This leads to the apparent paradox that the partial displacement of ANS from HSA increases its quenching effect on Trp-214. The most important conclusion is that (ANS-like) drugs cannot be displaced from the type-B sites, and consequently, drug transfer to these sites is not impaired by competitive binding of GC in the vicinity of a dissolving micelle. The second conclusion is that for unbound GC above the CMC (9 mM), ANS equilibrates between HSA and GC micelles but with a strong preference for free sites on HSA. That means that even persisting micelles would lose their cargo readily once exposed to HSA. For all MDDS sharing this property, targeted drug delivery approaches involving them as the nanocarrier would be pointless.