Petit J M, Maftah A, Ratinaud M H, Julien R
Institut de Biotechnologie, Faculté des Sciences, Limoges, France.
Eur J Biochem. 1992 Oct 1;209(1):267-73. doi: 10.1111/j.1432-1033.1992.tb17285.x.
The acridine orange derivative, 10N-nonyl acridine orange, is an appropriate marker of the inner mitochondrial membrane in whole cells. We use membrane model systems to demonstrate that 10N-nonyl acridine orange binds to negatively charged phospholipids (cardiolipin, phosphatidylinositol and phosphatidylserine). The stoichiometry has been found to be 2 mol 10N-nonyl acridine orange/mol cardiolipin and 1 mol dye/mol phosphatidylserine or phosphatidylinositol, while, with zwitterionic phospholipids, significant binding could not be detected. The affinity constants were 2 x 10(6) M-1 for cardiolipin-10N-nonyl-acridine-orange association and only 7 x 10(4) M-1 for that of phosphatidylserine and phosphatidylinositol association. The high affinity of the dye for cardiolipin may be explained by two essential interactions; firstly an electrostatic interaction between the quaternary ammonium of nonyl acridine orange and the ionized phosphate residues of cardiolipin and secondly, hydrophobic interactions between adjacent chromophores. A linear relationship was demonstrated between the cardiolipin content of model membranes and the incorporated dye. Consequently, a convenient and rapid method for cardiolipin quantification in membranes was established and applied to the cardiolipin-containing organelle, the mitochondrion.
吖啶橙衍生物10N-壬基吖啶橙是完整细胞内线粒体膜的一种合适标记物。我们使用膜模型系统证明10N-壬基吖啶橙与带负电荷的磷脂（心磷脂、磷脂酰肌醇和磷脂酰丝氨酸）结合。已发现化学计量比为2摩尔10N-壬基吖啶橙/摩尔心磷脂以及1摩尔染料/摩尔磷脂酰丝氨酸或磷脂酰肌醇，而对于两性离子磷脂，未检测到明显结合。心磷脂-10N-壬基吖啶橙缔合的亲和常数为2×10⁶ M⁻¹，磷脂酰丝氨酸和磷脂酰肌醇缔合的亲和常数仅为7×10⁴ M⁻¹。该染料对心磷脂的高亲和力可通过两种基本相互作用来解释；首先是壬基吖啶橙的季铵与心磷脂的离子化磷酸残基之间的静电相互作用，其次是相邻发色团之间的疏水相互作用。模型膜的心磷脂含量与掺入的染料之间呈现线性关系。因此，建立了一种简便快速的膜中心磷脂定量方法，并应用于含心磷脂的细胞器——线粒体。
