Name: luminol chemiluminescence in forensics
Uploaded: 2020-03-26T07:22:29.000Z
Duration: 3 min 2 s
Channel: JoVE

通常情况下，当反应物转化为产物时，二者之间的能量差称为反应焓变，该过程可以是放热反应（释放热量），也可以是吸热反应（吸收热量）。
在化学发光反应中，反应物形成一种处于电子激发态的中间产物。该中间产物不稳定。当中间产物弛豫至基态时，其内部的电子以可见光的形式释放能量，而非以热的形式释放。
这一现象与荧光类似。回顾一下，分子中的电子通常占据基态。当它们吸收能量时（例如暴露于特定波长的光），会被激发并跃迁至更高的激发态能级。当激发态弛豫回到基态时，多余的能量以可见光的形式释放出来。
然而，区分两者非常重要。它们的区别在于分子中电子的激发方式。在化学发光中，化学反应产生的能量使电子激发&#8230;而在荧光中，电子通过直接吸收光能或其他电磁辐射而被激发。当电子弛豫回到基态时，这两种类型的发光均以发射光的形式释放能量。
让我们来看一个每种情况的例子。当将紫外光照射到一瓶汤力水上时，奎宁中的电子会吸收能量并被激发到更高的能级。当这些电子回到较低能级时，便会以蓝光的形式释放出能量。
现在考虑一个荧光棒，其外管含有草酸二苯酯和染料，内管含有过氧化氢。当你弯曲荧光棒时，内管破裂，使反应物混合。过氧化氢将草酸二苯酯氧化，生成苯酚和环状过氧化物。该环状过氧化物分解为二氧化碳，并释放能量，使染料激发至更高能级。当染料回到基态时，会释放出一个光子。此处的过程属于化学发光，因为激发染料所需的能量来自化学反应。
在本实验中，您将通过一系列反应合成3-氨基邻苯二甲酰肼，也称为鲁米诺。随后，您将使鲁米诺与固体氢氧化钾存在下的二甲基亚砜反应，以观察其化学发光现象。

鲁米诺的合成

Synthesis of Luminol

化学反应可以根据其在转变为产物时是吸收热量还是释放热量，分为吸热反应或放热反应。反应物与产物之间的能量差称为反应的焓变，通过计算产物与反应物之间的能量差（即 ΔH）来确定。若 ΔH 为正值，则该反应为吸热反应；若为负值，则该反应为放热反应。虽然化学反应传统上被视为反应物转化为产物的过程，但许多反应是分多步进行的，过程中会形成中间体。在化学发光反应中，这些中间体从高能态跃迁至基态，释放出光子，随着反应的进行，可能观察到可见光。
<h4>原子能级</h4>
丹麦物理学家尼尔斯·玻尔（Niels Bohr）提出了一个理论，即围绕原子核运动的电子只能占据某些特定的轨道或能级。当一个原子的所有电子都处于最低可能的能级时，该原子被称为处于基态。当电子占据高于基态的能级时，该原子则处于激发态。
玻尔理论中的一个关键假设是，电子在受到扰动之前会一直保持在基态。因此，电子只有在吸收能量后才会被提升到激发态。当电子从激发态回到基态时，会释放出相应的能量。这种能量通常以光子的形式释放，光子的波长与激发态和基态之间的能量差直接相关。
当电子通过吸收特定能量或波长的光而被激发至较高能级时，这种现象称为荧光。如果你将一件洗涤过的白色T恤暴露在黑光或紫外光下，就可以观察到这种效应。T恤发出的特征性辉光是由其中洗涤剂和漂白化合物的荧光引起的，这些物质使T恤在普通光线下看起来更白。 
<h4>化学发光</h4>
在化学发光反应中，电子通过吸收反应过程中释放的热量而被激发。随后，当电子弛豫回到基态时，便会释放出光。化学发光与荧光的主要区别在于，化学发光中激发电子的能量直接来源于化学反应本身。
化学发光的一个实际应用是商业荧光棒。荧光棒内含有两种分离的溶液：一种含有过氧化物，另一种含有草酸二苯酯和一种彩色染料。当两种溶液混合时（即激活荧光棒时），过氧化物与草酸二苯酯发生反应，释放出能量，使染料被激发至更高的能态。随着反应物逐渐消耗，染料返回基态并释放出光。这正是荧光棒呈现其特征颜色的原因。
<h4>鲁米诺</h4>
鲁米诺是一种具有化学发光特性的化学物质，被广泛应用于多个领域，尤其是在法医学中。鲁米诺（C8H7N3O2）在与氧化剂混合时会发出蓝光。在法医学中，鲁米诺会与血红蛋白中的铁发生反应，使法医科学家能够检测到极微量的血液痕迹。
鲁米诺通过3-硝基邻苯二甲酸与肼的脱水反应合成。该反应经加热以除去水，再加入三甘醇进一步提高温度。随后，在高pH条件下，使用连二亚硫酸钠将3-硝基邻苯二甲酰肼中的硝基还原为氨基。在碱性条件下，3-硝基邻苯二甲酰肼可溶。加入冰醋酸后，鲁米诺即沉淀析出。
在氢氧化钾作用下，鲁米诺形成二价阴离子。氢氧根阴离子使鲁米诺中连接在氮原子上的两个氢原子发生去质子化。氧气将鲁米诺氧化至其激发态。当其从激发态回到基态时，会释放出蓝白色光。
<h4>参考文献</h4>
<ol>
	<li style="font-size:17px">Kotz, J.C., Treichel Jr, P.M., Townsend, J.R. (2015). 化学与化学反应性。加利福尼亚州贝尔蒙特：Brooks/Cole, Cengage Learning。</li>
</ol>

化学反应可以根据其在转变为产物时是吸收热量还是释放热量，分为吸热反应或放热反应。

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JoVE Lab Manual

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luminol chemiluminescence in forensics

Luminol chemiluminescence shows how a chemical reaction can produce light and help with blood detection. Chemical reactions may be endothermic or exothermic, depending on whether they absorb or release heat as reactants become products. The enthalpy of a reaction is the energy difference between reactants and products, or ΔH. A positive ΔH means the reaction is endothermic. A negative ΔH means it is exothermic.

Many reactions do not happen in a single step. They can pass through intermediates along the way. In chemiluminescent reactions, these intermediates move from a high-energy state to the ground state and release a photon of light. That light can be seen as the reaction continues.

Atomic energy levels help explain why this glow happens. Niels Bohr proposed that electrons can occupy only certain orbits, or energy levels, around the nucleus. An atom is in the ground state when its electrons are in the lowest possible energy level. When an electron moves to a higher level, the atom is in an excited state. The electron must absorb energy to move up, and it releases that energy when it falls back down. This release is often a photon of light, and the light wavelength depends on the energy gap. When light causes this kind of excitation, the process is called fluorescence, such as the glow seen under a black light on a white T-shirt washed with detergent and bleach compounds.

Chemiluminescence is related to fluorescence, but the energy comes from the chemical reaction itself. In glow sticks, one solution contains peroxide and the other contains diphenyl oxalate and a color dye. When the solutions mix, the reaction produces energy that excites the dye. As the reactants are used up, the dye returns to the ground state and gives off light. This is why glow sticks have their characteristic color.

Luminol is a chemiluminescent chemical with an important forensic use. It emits blue light when mixed with an oxidizing agent. In forensic science, luminol reacts with the iron in hemoglobin, which helps detect very small traces of blood. Luminol is made by dehydrating 3-nitrophthalic acid with hydrazine. The reaction is heated to remove water, and triethylene glycol is added to raise the temperature further. Next, the nitro group in 3-nitrophthalhydrazide is reduced with sodium dithionite to form an amino group at high pH. The compound is soluble in basic conditions, and glacial acetic acid is added to precipitate the luminol. With potassium hydroxide, luminol forms a dianion because hydroxide ions remove the two hydrogens attached to the nitrogens. Oxygen gas then oxidizes luminol to its excited state. When it relaxes back to the ground state, it gives off a blue-white light.

Luminol Chemiluminescence in Forensics

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Reaction Rate, Arrhenius, and Catalysts

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Recording a Reproducible Lab Experiment

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melting points

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boiling points

Boiling Point and Vapor Pressure

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recrystallization

Purifying Crystals by Solubility

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extraction

Separating Mixtures with Solvent Extraction

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simple distillation

Separating Liquids by Boiling Point

Separating liquids by boiling point is the goal of distillation. A miscible mixture contains liquids that mix evenly, and it can be heated so that the more volatile liquid evaporates first. Volatility is a liquid’s tendency to vaporize, and boiling begins when bubbles form in the liquid phase. In a closed container, liquid and vapor can reach equilibrium, which means the rate of vaporization equals the rate of condensation. The pressure from the vapor above the liquid is called vapor pressure.

steam distillation

Steam Distillation and Low-Boiling Separation

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thin-layer chromatography

TLC Separation by Polarity

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column chromatography

Separating Mixtures by Polarity

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synthesis of luminol

Luminol chemiluminescence shows how a chemical reaction can produce light and help with blood detection. Chemical reactions may be endothermic or exothermic, depending on whether they absorb or release heat as reactants become products. The enthalpy of a reaction is the energy difference between reactants and products, or ΔH. A positive ΔH means the reaction is endothermic. A negative ΔH means it is exothermic. Many reactions do not happen in a single step. They can pass through intermediates...

esterification

Ester Formation and Everyday Fragrances

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identification of unknown aldehydes and ketones

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Carbonyl Tests for Aldehydes and Ketones

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chemistry

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Learn how luminol chemiluminescence produces blue-white light, why glow sticks glow, and how forensic scientists use luminol to detect traces of blood.

Luminol Chemiluminescence in Forensics | JoVE K12

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Luminol Chemiluminescence in Forensics

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