📚 Chromatography: Principles and Applications | 色谱法：原理与应用
Chromatography is an analytical technique used to separate, identify and quantify the individual components in a mixture. It has applications in forensic science, pharmaceutical analysis, environmental monitoring and food testing. All chromatographic methods depend on the distribution of substances between a mobile phase and a stationary phase.
色谱法是一种用于分离、鉴定和定量混合物中单个组分的分析技术。它广泛应用于法医学、药物分析、环境监测和食品检测。所有色谱方法都依赖于物质在流动相和固定相之间的分配。
1. Introduction to Chromatography | 色谱法简介
Chromatography is a physical separation technique in which the sample components are carried by a moving phase while interacting with a fixed phase. The different components travel at different rates, allowing them to be separated.
色谱法是一种物理分离技术，样品组分由流动相携带，同时与固定相发生作用。不同组分以不同速率移动，从而得以分离。
The technique can be used for qualitative analysis, to identify components by comparison with known standards, and for quantitative analysis, to determine how much of each component is present.
该技术可用于定性分析，通过与已知标准品比较来鉴定组分；也可用于定量分析，确定每种组分的含量。
Common types studied at A-level include thin-layer chromatography, column chromatography and gas chromatography.
A-level 阶段学习的主要类型包括薄层色谱、柱色谱和气相色谱。
2. Principles: Mobile and Stationary Phases | 原理：流动相与固定相
The mobile phase is the phase that moves over or through the stationary phase, carrying the sample along with it. It may be a liquid or a gas.
流动相是沿固定相表面或穿过固定相移动的相，携带样品前进。流动相可以是液体或气体。
The stationary phase is fixed in place and interacts with the sample components to different extents. It may be a solid or a thin film of liquid supported on an inert solid.
固定相固定在原位，与样品组分发生不同程度的相互作用。固定相可以是固体，也可以是负载在惰性固体上的薄层液体。
Separation occurs because each component has a different balance between its attraction to the stationary phase and its solubility or volatility in the mobile phase.
分离之所以发生，是因为每种组分在固定相上的吸引力与在流动相中的溶解度或挥发性之间存在不同的平衡。
Components that interact strongly with the stationary phase move more slowly, while components that spend more time in the mobile phase move faster.
与固定相作用强的组分移动较慢，而在流动相中停留时间更长的组分移动较快。
3. Thin-Layer Chromatography (TLC) | 薄层色谱 (TLC)
In thin-layer chromatography, the stationary phase is a thin layer of silica gel (SiO₂) or alumina (Al₂O₃) coated on a glass, plastic or metal sheet.
在薄层色谱中，固定相是涂覆在玻璃、塑料或金属薄片上的硅胶（SiO₂）或氧化铝（Al₂O₃）薄层。
A small amount of sample is dissolved in a volatile solvent and spotted onto a pencil-drawn baseline. The baseline must be above the level of the developing solvent.
将少量样品溶解在挥发性溶剂中，点在用铅笔画出的基线上。基线必须高于展开剂的液面。
The plate is placed in a sealed developing chamber containing a shallow layer of solvent. The solvent rises up the plate by capillary action, carrying the sample components with it.
将薄板放入含有少量溶剂的密闭展开缸中。溶剂通过毛细作用沿板上升，携带样品组分向上移动。
Separation is mainly by adsorption onto the polar stationary phase, so more polar components tend to move more slowly.
分离主要依靠组分在极性固定相上的吸附，因此极性较强的组分通常移动较慢。
After development, the plate is removed and the solvent front is marked immediately. Spots may be observed under UV light or by using a chemical locating agent.
展开完毕后取出薄板，立即标记溶剂前沿。可在紫外光下观察斑点，或使用化学显色剂显色。
4. Calculating Rf Values | Rf 值的计算
The retention factor, Rf, is used to characterise a substance in a particular solvent and stationary phase system. It is calculated using the following equation:
保留因子 Rf 用于表征物质在特定溶剂和固定相体系中的行为。其计算公式如下：
Rf = distance moved by substance / distance moved by solvent front
Both distances are measured from the baseline. The distance moved by the substance is measured to the centre of its spot, and the distance moved by the solvent front is measured to the furthest point reached by the solvent.
两个距离均从基线量起。物质移动距离量至斑点中心，溶剂前沿移动距离量至溶剂到达的最远点。
Rf is a dimensionless number with a value between 0 and 1 for components that move from the baseline.
Rf 是一个无量纲数，对于从基线移动的组分，其值在 0 到 1 之间。
le, if a spot moves 2.4 cm and the solvent front moves 6.0 cm, then Rf = 2.4 ÷ 6.0 = 0.40.
例如，若斑点移动 2.4 cm，溶剂前沿移动 6.0 cm，则 Rf = 2.4 ÷ 6.0 = 0.40。
An unknown substance can be identified by comparing its Rf value with those of known standards run on the same plate under identical conditions.
通过与在相同条件下同一薄板上运行的标准品 Rf 值比较，可以鉴定未知物质。
5. Column Chromatography | 柱色谱
Column chromatography uses a vertical glass column packed with a solid stationary phase, usually silica gel or alumina. The sample is placed carefully at the top of the column.
柱色谱使用垂直玻璃柱，填充固体固定相，通常为硅胶或氧化铝。样品小心地加在柱顶。
A liquid mobile phase, called the eluent, is passed through the column. This can be done under gravity or with applied pressure to speed up the flow.
液体流动相称为洗脱剂，通过柱体。洗脱可在重力作用下进行，也可施加压力以加快流速。
Components move down the column at different rates depending on how strongly they adsorb to the stationary phase. They leave the column at different times and can be collected as separate fractions.
各组分根据其在固定相上吸附的强弱以不同速率向下移动，在不同时间离开柱子，可分别收集为不同馏分。
Column chromatography can separate larger quantities of material than thin-layer chromatography, making it useful for preparative work.
柱色谱可比薄层色谱分离更大量的物质，因此适用于制备性分离。
6. Gas Chromatography (GC) | 气相色谱 (GC)
Gas chromatography separates volatile substances using a gaseous mobile phase. The mobile phase is an inert carrier gas, commonly helium or nitrogen.
气相色谱使用气体流动相分离挥发性物质。流动相是惰性载气，通常为氦气或氮气。
The stationary phase is a high-boiling liquid adsorbed onto an inert solid support, or a solid adsorbent. It is coated or packed inside a long coiled column placed in an oven.
固定相是吸附在惰性固体载体上的高沸点液体，或固体吸附剂。它涂覆或填充在置于柱温箱中的长螺旋柱内。
A small sample is injected through a heated injection port and is immediately vaporised. The carrier gas carries the vapour through the column.
少量样品经加热进样口注入并立即气化。载气将蒸气带入色谱柱。
Components partition between the carrier gas and the liquid stationary phase. Those with lower boiling points or weaker interactions with the stationary phase travel faster and reach the detector first.
组分在载气和液体固定相之间进行分配。沸点较低或与固定相作用较弱的组分移动较快，先到达检测器。
7. Retention Time and Peak Integration | 保留时间与峰面积积分
The retention time is the time taken for a component to travel from injection to the detector, recorded as the time at the peak maximum on a gas chromatogram.
保留时间是指组分从进样到检测器所需的时间，在气相色谱图上记录为峰最高点对应的时间。
Under fixed conditions of column type, temperature, pressure and flow rate, a given compound has a characteristic retention time. This can be used for identification.
在色谱柱类型、温度、压力和流速固定的条件下，某一化合物具有特征保留时间，可用于鉴定。
The area under each peak is proportional to the amount of that component reaching the detector. Peak areas are used for quantitative analysis, often by constructing calibration curves with known concentrations.
每个峰的面积与该到达检测器的组分量成正比。峰面积用于定量分析，通常通过已知浓度绘制标准曲线。
An internal standard may be added to improve the precision and accuracy of quantitative results.
可加入内标物以提高定量结果的精密度和准确度。
8. Gas Chromatography–Mass Spectrometry (GC-MS) | 气相色谱-质谱联用 (GC-MS)
Gas chromatography can be coupled directly to a mass spectrometer to create a GC-MS instrument. The GC separates the mixture, and the mass spectrometer identifies each separated component.
气相色谱可直接与质谱仪联用，构成 GC-MS 仪器。气相色谱分离混合物，质谱仪鉴定每个分离后的组分。
Each component leaving the GC column enters the mass spectrometer, where it is ionised and fragmented into charged pieces.
离开气相色谱柱的每个组分进入质谱仪，在其中被离子化并碎裂成带电碎片。
The mass spectrum records the mass-to-charge ratio (m/z) of these fragments. The fragmentation pattern is highly characteristic of the original molecule.
质谱记录这些碎片的质荷比 (m/z)。碎裂模式是原始分子的高度特征指纹。
The spectrum can be compared with a spectral database to identify unknown compounds quickly and with high confidence.
将质谱与谱库进行比对，可快速且高可信度地鉴定未知化合物。
9. Interpreting Chromatograms | 色谱图解析
A gas chromatogram is a plot of detector response against time. Each peak normally corresponds to one component, provided the resolution is good enough to separate them.
气相色谱图是检测器响应值对时间作图。若分离度足够好，每个峰通常对应一种组分。
The number of peaks gives the number of components detected under the chosen conditions. It does not necessarily show all components if some do not vaporise or are not detected.
峰的数量表示在所选条件下检测到的组分数目。如果某些组分不能气化或未被检测到，则色谱图不一定显示所有组分。
Retention times identify components when compared with known standards run under identical conditions. Peak areas determine the relative or absolute amounts present.
通过与相同条件下运行的标准品保留时间比较来鉴定组分。峰面积可确定存在的相对量或绝对量。
10. Limitations and Calibration | 局限性与校准
Chromatography has important limitations. Two different components can have the same or very similar retention time, causing co-elution and overlapping peaks.
色谱法有重要局限性。两种不同组分可能具有相同或非常相近的保留时间，导致共同洗脱和峰重叠。
Gas chromatography is limited to substances that are volatile and thermally
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