\[c = \frac{A_{280}}{\varepsilon_{280}\, l},\qquad \varepsilon_{280} \approx 5500\,n_\mathrm{W} + 1490\,n_\mathrm{Y} + 125\,n_\mathrm{SS}\ \ \mathrm{L\,mol^{-1}\,cm^{-1}}\]
含义Meaning
蛋白质在 280 nm 的吸收主要来自色氨酸和酪氨酸（及二硫键），由序列可估算摩尔吸光系数（Pace 法）。
Protein absorbance at 280 nm comes mainly from Trp and Tyr (and disulfides); ε can be estimated from sequence (Pace method).
符号说明Symbols
| 符号Symbol | 含义 Meaning | 单位 Unit |
|---|---|---|
| \(n_\mathrm{W}, n_\mathrm{Y}\) | 色氨酸、酪氨酸残基数number of Trp and Tyr residues | 1 |
| \(n_\mathrm{SS}\) | 二硫键（胱氨酸）数number of disulfides | 1 |
| \(l\) | 光程path length | cm |
推导Derivation
朗伯–比尔定律 \(A = \varepsilon c l\)。
Beer–Lambert \(A = \varepsilon c l\).
在变性条件下各发色团近似独立吸收，ε 为各残基贡献之和。
Under denaturing conditions chromophores absorb independently; ε is the sum of contributions.
适用条件与局限Assumptions & limitations
核酸在 260 nm 强吸收，会干扰（A₂₆₀/A₂₈₀ > 0.6 提示核酸污染）。
Nucleic acids absorb strongly at 260 nm; A₂₆₀/A₂₈₀ > 0.6 signals contamination.
无 Trp/Tyr 的肽不适用；折叠态 ε 与估算值可有约 5% 差异。
Not for peptides lacking Trp/Tyr; folded-state ε can differ by ~5 %.