We investigated the discoloration of a highly concentrated monoclonal antibody (mAbZ) in sodium acetate (NaAc) and histidine/lysine (His/Lys) buffer after exposure to visible light. The color change of the mAbZ formulation was significantly more intense in NaAc buffer and developed a characteristic absorbance with a λ_max of ca. 450 nm. We characterized this photo-chemically generated chromophore by comparison with visible light photo-degradation of a concentrated solution of a model compound for protein Trp residues, N-acetyl-l-tryptophan amide (NATA). The photo-degradation of NATA generated a chromophoric product with a λ_max of ca. 450 nm and UV–vis spectroscopic properties identical to those of the product generated from mAbZ. This product was isolated and analyzed by high-performance liquid chromatography tandem mass spectrometry (HPLC–MS/MS) and ¹H, ¹³C, and ¹H–¹³C heteronuclear single-quantum correlation NMR spectroscopy. MS/MS analysis reveals a product characterized by the loss of 33 Da from NATA, referred to as NATA-33. Together, the NMR data suggest that this product may be N-(2,4-dihydrocyclopenta[b]indol-2-yl)acetamide (structure P3a) or a tautomer (P3b–d).

中文翻译：

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高浓度制剂中单克隆抗体的可见光降解：通过与 N-乙酰基-l-色氨酸酰胺的光降解比较来表征色氨酸衍生的发色光产物

我们研究了高浓度单克隆抗体 (mAbZ) 在乙酸钠 (NaAc) 和组氨酸/赖氨酸 (His/Lys) 缓冲液中暴露于可见光后的变色情况。mAbZ 制剂的颜色变化在 NaAc 缓冲液中明显更强烈，并产生了特征吸光度，λ _max为 ca。450 纳米。我们通过与蛋白质色氨酸残基模型化合物N-乙酰-l-色氨酸酰胺 (NATA)的浓缩溶液的可见光光降解比较来表征这种光化学产生的生色团。NATA 的光降解产生具有 λ _{max的发色产物}约 450 nm 和 UV-vis 光谱特性与 mAbZ 产生的产品相同。通过高效液相色谱串联质谱 (HPLC-MS/MS) 和¹ H、¹³ C 和¹ H- ¹³ C 异核单量子相关核磁共振波谱分离和分析该产物。MS/MS 分析揭示了一种产物，其特征是从 NATA 中丢失 33 Da，称为 NATA-33。总之，NMR 数据表明该产物可能是N- (2,4-dihydrocyclopenta[ b ]indol-2-yl)acetamide（结构P3a）或互变异构体（P3b – d）。