📚 IB Chemistry: Chromatography Revision Guide | IB 化学：色谱考点精讲
Chromatography is a vital analytical technique in IB Chemistry used to separate, identify, and quantify the components of a mixture. It relies on the differential distribution of substances between a stationary phase and a mobile phase. Understanding the principles of chromatography, including thin-layer chromatography (TLC), gas chromatography (GC), and high-performance liquid chromatography (HPLC), is essential for success in both Paper 2 and the Internal Assessment. This revision guide covers all key concepts and common exam questions, providing balanced bilingual explanations.
色谱是 IB 化学中一种重要的分析技术，用于分离、鉴定和定量混合物中的组分。它依赖于物质在固定相和流动相之间的差异分配。理解色谱的原理，包括薄层色谱 (TLC)、气相色谱 (GC) 和高效液相色谱 (HPLC)，对于在笔试和内部评估中取得好成绩至关重要。本考点精讲覆盖所有关键概念和常见考题，提供均衡的中英双语解析。
1. What Is Chromatography? | 什么是色谱？
Chromatography is a physical separation method in which the components of a mixture are distributed between two phases: a stationary phase (which is fixed in place) and a mobile phase (which moves through or over the stationary phase). As the mobile phase carries the sample, each component interacts differently with the stationary phase, leading to different migration rates and thus separation.
色谱是一种物理分离方法，混合物中的组分在两相之间分配：固定相（固定在原位）和流动相（流过或穿过固定相）。当流动相携带样品移动时，每种组分与固定相的相互作用不同，导致迁移速率不同，从而实现分离。
The technique can be used for qualitative analysis (identifying substances by comparing their retention times or Rf values with standards) and quantitative analysis (determining the amount of each component based on peak area or spot intensity).
该技术可用于定性分析（通过比较保留时间或 Rf 值与标准品来鉴定物质）和定量分析（根据峰面积或斑点强度确定每种组分的含量）。
2. Basic Principles of Separation | 分离的基本原理
Separation in chromatography depends on the relative affinities of the analytes for the stationary and mobile phases. The more strongly a component adsorbs onto or dissolves in the stationary phase, the slower it moves. Conversely, a component that is highly soluble in the mobile phase will travel faster.
色谱分离取决于分析物对固定相和流动相的相对亲和力。一种组分吸附或溶解在固定相中的能力越强，移动越慢。相反，一种在流动相中溶解度高的组分移动得越快。
The distribution of a solute between the two phases is described by the partition coefficient (KD). In partition chromatography, separation relies on differences in solubility; in adsorption chromatography, it relies on differences in the strength of intermolecular forces with the stationary surface.
溶质在两相之间的分布由分配系数 (KD) 描述。在分配色谱中，分离依赖于溶解度的差异；在吸附色谱中，则依赖于与固定相表面分子间作用力的强弱差异。
3. Stationary and Mobile Phases | 固定相与流动相
The choice of phases determines the type of chromatography and its suitability for different samples. Common stationary phases include solid adsorbents (silica gel, alumina), liquid films coated on a solid support, or a bonded phase on a solid particle. Mobile phases can be gases, liquids, or supercritical fluids.
相的选择决定了色谱的类型及其对不同样品的适用性。常见的固定相包括固体吸附剂（硅胶、氧化铝）、涂覆在固体支持物上的液膜或键合在固体颗粒上的键合相。流动相可以是气体、液体或超临界流体。
| Chromatography Type | Mobile Phase | Stationary Phase |
|---|---|---|
| Paper / TLC | Liquid solvent | Water in paper fibers / silica gel layer |
| Column | Liquid solvent | Solid packing (silica, alumina) |
| Gas chromatography (GC) | Inert carrier gas (e.g. He, N2) | High-boiling liquid on solid support or solid adsorbent |
| HPLC | Liquid under high pressure | Small-diameter solid particles with bonded phase |
色谱类型 / 流动相 / 固定相：纸/TLC：液体溶剂 / 纸纤维中的水或硅胶层；柱色谱：液体溶剂 / 固体填料（硅胶、氧化铝）；气相色谱：惰性载气（如 He、N2） / 固体支持物上的高沸点液体或固体吸附剂；HPLC：高压液体 / 带有键合相的小直径固体颗粒。
4. Thin-Layer Chromatography (TLC) | 薄层色谱法
TLC uses a glass, plastic, or aluminium sheet coated with a thin layer of adsorbent (usually silica gel) as the stationary phase. A small spot of the sample is placed near the bottom of the plate, and the plate is placed upright in a developing chamber containing a shallow layer of solvent (mobile phase). The solvent rises by capillary action, carrying the components at different rates.
TLC 使用涂有薄层吸附剂（通常为硅胶）的玻璃板、塑料片或铝箔作为固定相。在板底部附近点上少量样品，然后将板直立放入装有浅层溶剂（流动相）的展开缸中。溶剂通过毛细作用上升，以不同速率携带组分。
Once the solvent front has migrated close to the top of the plate, the plate is removed and dried. Colourless spots can be visualised using UV light, iodine vapour, or chemical staining. The Rf value (retention factor) is calculated for each spot:
当溶剂前沿迁移到接近板顶部时，取出板并干燥。无色斑点可使用紫外光、碘蒸气或化学染色显色。计算每个斑点的 Rf 值（保留因子）：
Rf = distance travelled by component ÷ distance travelled by solvent front
Rf = 组分移动距离 ÷ 溶剂前沿移动距离
The Rf value is a dimensionless number between 0 and 1, specific to a given compound under identical stationary/mobile phase conditions, and is used for identification.
Rf 值是一个介于 0 到 1 之间的无量纲数，在相同固定相/流动相条件下对特定化合物是唯一的，可用于鉴定。
5. Paper Chromatography | 纸色谱法
Paper chromatography uses a strip of high-quality filter paper where the stationary phase is water adsorbed onto the cellulose fibres. The mobile phase is a liquid solvent, often a mixture of water and an organic solvent. It is particularly suitable for separating highly polar or water-soluble compounds, such as amino acids and sugars.
纸色谱使用高品质滤纸条，其固定相是吸附在纤维素纤维上的水。流动相是液体溶剂，通常是水与有机溶剂的混合物。它特别适合分离强极性或水溶性化合物，如氨基酸和糖类。
The procedure and analysis are similar to TLC: after development, spots can be located with a locating agent (e.g. ninhydrin for amino acids) and Rf values determined. In IB practical work, paper chromatography is often used to investigate photosynthetic pigments or amino acids in an unknown mixture.
操作和分析与 TLC 类似：展开后，可用显色剂（例如茚三酮用于氨基酸）定位斑点并测定 Rf 值。在 IB 实验活动中，纸色谱常用于研究光合色素或未知混合物中的氨基酸。
6. Column Chromatography | 柱色谱法
Column chromatography is a preparative technique used to separate larger quantities of a mixture. The stationary phase (silica gel or alumina) is packed into a vertical glass column, and the sample is placed on top. The mobile phase (eluent) is continuously poured through the column, and components are collected in separate fractions as they elute at different times.
柱色谱是一种用于分离较大样品量的制备技术。固定相（硅胶或氧化铝）填充在垂直玻璃柱中，样品置于顶部。流动相（洗脱剂）连续流过色谱柱，各组分在不同时间洗脱并分别收集馏分。
Although not always a core written topic, understanding column chromatography reinforces the principles of adsorption and partition. It is the ancestor of modern HPLC, offering students insight into how continuous elution separates components based on polarity differences.
虽然柱色谱并非 IB 笔试核心专题，但理解它有助于巩固吸附和分配原理。它是现代 HPLC 的前身，让学生了解连续洗脱如何基于极性差异分离组分。
7. Gas Chromatography (GC) | 气相色谱法
In GC, the sample is vaporised and injected into a stream of inert carrier gas (mobile phase), such as helium or nitrogen. The mixture passes through a long, coiled column containing a stationary phase, either a solid adsorbent (gas-solid chromatography) or a liquid coating on a solid support (gas-liquid chromatography).
在气相色谱中，样品被气化并注入惰性载气（流动相，如氦气或氮气）流中。混合物通过一根长而盘绕的色谱柱，柱内含有固定相，可以是固体吸附剂（气固色谱）或涂覆在固体支持物上的液体（气液色谱）。
Components are separated based on their volatility and their affinity for the stationary phase. A detector at the column outlet produces a signal that is recorded as a chromatogram. Common detectors include the flame ionisation detector (FID) and thermal conductivity detector (TCD). Each separated component appears as a peak; the time from injection to the peak maximum is the retention time (tR).
组分根据其挥发性和与固定相的亲和力被分离。柱出口处的检测器产生信号并记录为色谱图。常用检测器包括火焰离子化检测器 (FID) 和热导检测器 (TCD)。每个分离的组分以一个峰出现；从进样到峰顶点的时间为保留时间 (tR)。
8. High-Performance Liquid Chromatography (HPLC) | 高效液相色谱法
HPLC is an advanced form of column chromatography in which the mobile phase (a liquid solvent) is pumped through a column packed with very small particles under high pressure. This provides high resolution, rapid separation, and the ability to analyse thermally labile or non-volatile compounds that GC cannot handle.
HPLC 是一种高级形式的柱色谱，流动相（液体溶剂）在高压下被泵入填充有极小颗粒的色谱柱中。这提供了高分辨率、快速分离，并能分析 GC 无法处理的热不稳定或非挥发性化合物。
In reversed-phase HPLC, the stationary phase is non-polar (e.g. C18 hydrocarbon chains bonded to silica) and the mobile phase is polar (e.g. water/acetonitrile mixture). Polar analytes elute first. In normal-phase HPLC, a polar stationary phase is used with a non-polar mobile phase. A UV/Vis detector is commonly used for quantification.
在反相 HPLC 中，固定相是非极性的（例如键合在硅胶上的 C18 烃链），流动相是极性的（例如水/乙腈混合物）。极性分析物先洗脱。在正相 HPLC 中，使用极性固定相和非极性流动相。常用紫外/可见光检测器进行定量分析。
9. Interpreting Chromatograms: Rf and Retention Time | 色谱图解读：Rf 和保留时间
For planar techniques (TLC, paper), the primary identifier is the Rf value, calculated as shown earlier. By running a known standard alongside the unknown on the same plate, a match of Rf values provides a strong presumption of identity. Multiple solvent systems can improve reliability.
对于平面技术（TLC、纸色谱），主要标识符是 Rf 值，如前所示计算。通过在同一块板上同时跑已知标准品和未知物，若 Rf 值匹配，则可有力地推测为同一物质。使用多种溶剂系统可提高可靠性。
In GC and HPLC, the retention time (tR) under fixed conditions (column temperature, flow rate, mobile phase composition) is characteristic of a compound. Peaks are compared with those of known standards injected under the same conditions. The area under each peak is proportional to the amount of that component in the mixture.
在 GC 和 HPLC 中，在固定条件（柱温、流速、流动相组成）下的保留时间 (tR) 是化合物的特征。将峰与相同条件下进样的已知标准品峰进行比较。每个峰的面积与该组分在混合物中的含量成正比。
10. Factors Affecting Separation | 影响分离的因素
- Polarity: In adsorption chromatography, the more polar a substance, the more strongly it is retained on a polar stationary phase (like silica). Increasing the polarity of the mobile phase reduces retention.
- Temperature: In GC, higher column temperatures decrease retention times and can improve peak shape but may reduce resolution if too high.
- Flow rate of the mobile phase: Faster flow rates reduce separation time but can broaden peaks and reduce resolution.
- Column length and particle size: Longer columns or smaller stationary-phase particles (HPLC) increase theoretical plates, enhancing resolution but increasing backpressure.
- Partition coefficient: The thermodynamics of a compound's distribution between phases directly determines its migration speed.
- 极性：在吸附色谱中，物质极性越强，在极性固定相（如硅胶）上保留越强。提高流动相极性会减弱保留。
- 温度：在 GC 中，较高的柱温可缩短保留时间并改善峰形，但温度过高可能降低分辨率。
- 流动相流速：流速加快可缩短分离时间，但可能使峰展宽并降低分辨率。
- 柱长和粒径：较长的柱或更小的固定相颗粒（HPLC）可增加理论塔板数，提高分辨率，但会增加背压。
- 分配系数：化合物在两相间分配的热力学性质直接决定其迁移速度。
Understanding how to adjust these parameters is often tested in the context of optimising a separation, e.g., choosing a more polar solvent in TLC to improve separation of closely spaced spots.
理解如何调整这些参数常在优化分离的考题中出现，例如，在 TLC 中选择极性更强的溶剂以改善紧邻斑点的分离效果。
11. Qualitative and Quantitative Analysis | 定性与定量分析
Qualitative analysis involves identifying the components of a mixture. In TLC, an unknown spot's Rf value is compared with that of a known substance run on the same plate. In GC or HPLC, the retention time of a peak is matched against a standard's retention time. Mass spectrometry (GC-MS) can provide definitive identification by molecular mass and fragmentation pattern.
定性分析涉及鉴定混合物的各组分。在 TLC 中，将未知斑点的 Rf 值与同板上已知物质的 Rf 值进行比较。在 GC 或 HPLC 中，将峰的保留时间与标准品保留时间匹配。气相色谱-质谱联用 (GC-MS) 可通过分子质量和碎片模式提供确认性鉴定。
Quantitative analysis uses the principle that the detector response (peak area in GC/HPLC, spot intensity in TLC with a densitometer) is proportional to the concentration or mass of the analyte. A calibration curve prepared from standards of known concentration is used to determine the amount of the analyte in an unknown sample.
定量分析利用检测器响应（GC/HPLC 中的峰面积，使用光密度计的 TLC 斑点强度）与分析物的浓度或质量成正比的原理。用已知浓度标准品制备的校准曲线可用于确定未知样品中分析物的含量。
12. Applications and Limitations | 应用与局限性
Chromatography is used extensively in forensic science (drug and toxicology tests), pharmaceutical quality control (purity testing and assay of active ingredients), environmental monitoring (detecting pesticides in water), and food chemistry (analysing additives and contaminants). In the IB laboratory, TLC and column chromatography are common tools for purifying reaction products.
色谱广泛应用于法医学（药物和毒理学检测）、药品质量控制（纯度和活性成分含量测定）、环境监测（检测水中杀虫剂）和食品化学（分析添加剂和污染物）。在 IB 实验室中，TLC 和柱色谱是纯化反应产物的常用工具。
However, chromatography has limitations. Overlapping peaks (co-elution) can make quantification inaccurate. Rf values can change with temperature and solvent composition. Some detectors destroy the sample. Moreover, without a pure standard, identifying an unknown peak is tentative. These limitations highlight why hyphenated techniques (GC-MS, LC-MS) are preferred for definitive analysis.
然而，色谱也有局限性。重叠峰（共洗脱）会导致定量不准确。Rf 值可随温度和溶剂组成变化。某些检测器会破坏样品。此外，若没有纯标准品，对未知峰的鉴定只是推测性的。这些局限性说明了为何联用技术（GC-MS、LC-MS）更适于确证分析。
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