Colombi Chiara, Dacarro Giacomo, Diaz Fernandez Yuri Antonio, Taglietti Angelo, Pallavicini Piersandro, Doveri Lavinia
Dipartimento di Chimica, Università degli Studi di Pavia, v. Taramelli, 12, 27100 Pavia, Italy.
Nanomaterials (Basel). 2024 Aug 11;14(16):1336. doi: 10.3390/nano14161336.
Prussian Blue nanoparticles (PBnps) are now popular in nanomedicine thanks to the FDA approval of PB. Despite the numerous papers suggesting or describing the in vivo use of PBnps, no studies have been carried out on the formation of a protein corona on the PBnp surface and its stabilizing role. In this paper, we studied qualitatively and quantitatively the corona formed by the most abundant protein of blood, human serum albumin (HSA). Cubic PBnps (41 nm side), prepared in citric acid solution at PB concentration 5 × 10 M, readily form a protein corona by redissolving ultracentrifuged PBnp pellets in HSA solutions, with C ranging from 0.025 to 7.0 mg/mL. The basic decomposition of PBnp@HSA was studied in phosphate buffer at the physiological pH value of 7.4. Increased stability with respect to uncoated PBnps was observed at all concentrations, but a minimum C value of 3.0 mg/mL was determined to obtain stability identical to that observed at serum-like HSA concentrations (35-50 mg/mL). Using a modified Lowry protocol, the quantity of firmly bound HSA in the protein corona (hard corona) was determined for all the C used in the PBnp@HSA synthesis, finding increasing quantities with increasing C. In particular, an HSA/PBnp number in the 1500-2300 range was found for C 3.0-7.0 mg/mL, largely exceeding the 180 HSA/PBnp value calculated for an HSA monolayer on a PBnp. Finally, the stabilization brought by the HSA corona allowed us to carry out pH-spectrophotometric titrations on PBnp@HSA in the 3.5-9-0 pH range, revealing a pKa value of 6.68 for the water molecules bound to the Fe centers on the PBnp surface, whose deprotonation is responsible for the blue-shift of the PBnp band from 706 nm (acidic solution) to 685 nm (basic solution).
由于普鲁士蓝（PB）获得了美国食品药品监督管理局（FDA）的批准，普鲁士蓝纳米颗粒（PBnps）如今在纳米医学领域颇受欢迎。尽管有大量论文提及或描述了PBnps的体内应用，但尚未有关于PBnp表面蛋白质冠层的形成及其稳定作用的研究。在本文中，我们对血液中最丰富的蛋白质——人血清白蛋白（HSA）形成的冠层进行了定性和定量研究。在柠檬酸溶液中以5×10 M的PB浓度制备的边长为41 nm的立方PBnps，通过将超速离心后的PBnp沉淀重新溶解于浓度范围为0.025至7.0 mg/mL的HSA溶液中，很容易形成蛋白质冠层。在生理pH值7.4的磷酸盐缓冲液中研究了PBnp@HSA的基本分解情况。在所有浓度下均观察到相对于未包覆PBnps稳定性有所提高，但确定最小C值为3.0 mg/mL时，才能获得与类似血清HSA浓度（35 - 50 mg/mL）下观察到的相同稳定性。使用改良的洛瑞法，针对PBnp@HSA合成中使用的所有C值，测定了蛋白质冠层（硬冠层）中牢固结合的HSA的量，发现其随C值增加而增加。特别是，对于C为3.0 - 7.0 mg/mL时，发现HSA/PBnp数量在1500 - 2300范围内，大大超过了PBnp上HSA单层计算得出的180 HSA/PBnp值。最后，HSA冠层带来的稳定性使我们能够在3.5 - 9.0的pH范围内对PBnp@HSA进行pH分光光度滴定，揭示了与PBnp表面Fe中心结合的水分子的pKa值为6.68，其去质子化导致PBnp谱带从706 nm（酸性溶液）蓝移至685 nm（碱性溶液）。
