microscopic scenarios of the molecular processes involved.
Important mathematical relationships are given to aid users with their practical applications and theoretical studies of the dozens of separation modes performed in the three most common platforms: capillaries, microchips, and slabs. The two most important layouts of these three platforms are also described and compared: the conventional open layout, with an inlet and an outlet for the samples, and the toroidal or closed loop layout, where samples run in a quasi-continuous circulating mode until the desired resolution is obtained.
Motivation to write this book came from twenty years of teaching both undergraduate and graduate students, as well as from stimulating collaborations with so many bright and talented colleagues, to all of whom I am deeply grateful.
The author is thankful to Dr C. Heller, Professor J. H. Z. dos Santos, Professor C. L. Petzhold, Professor A. Manz, Professor Bingcheng Lin, Professor B. Gaš, Professor J. W. Jorgenson, and Professor H-P. Grieneisen for their helpful discussions and suggestions. Additionally, the author is grateful to E. J. M. Bradley for proofreading the manuscript. Finally, it must be mentioned that these people should not be blamed for the errors the author insisted on keeping in this book!
January, 2020
Tarso B. Ledur Kist
Porto Alegre
Introduction
Toroidal electrokinetic separation techniques are based on separation tracks with toroidal layouts. These techniques can produce analytical and preparative separations with unprecedented high resolutions and peak capacities. Runs are performed in a closed loop with a quasi-continuous circulating mode of migration until the desired resolutions are achieved. They are different to the commonly used open layouts, where runs are always limited in either space (with an inlet and an outlet) or time (with a start and a finish line) and either electroosmosis or a pressure driven counter-flow must be applied to increase the resolving power. Toroidal layouts allow much more freedom in the use of the operating conditions, as will be described and compared in the following chapters.
Electrokinetic phenomena is a class of phenomena that includes electrophoresis, electroosmosis, streaming current and potential, surface conductivity, dielectric dispersion, and electroacoustics. The phenomena occur in liquid solutions (some of them can also occur in gels) containing electrolytes, and are intimately related to the theory of electromagnetism and classical fluid dynamics. However, they are distinct from electrochemistry related phenomena as they focus on the transport of charged and uncharged entities instead of on the chemical reactions, which involve the movement of electrical charges between electrodes and electrolytes.
The application of static or alternating electric fields to liquid solutions promotes the analytical and preparative separation of charged and uncharged entities, ranging from mono-atomic to macroscopic particles. This has led to the development of dozens of techniques that are indistinctly called electrophoretic, electromigration, electromigrative, electrodriven, or electrokinetic separation techniques. Luckily, all of them have the same acronym: ESTs! However, the term electrophoretic seems to be too specific, as it only refers to one of the electrokinetic phenomena. The term electromigration became commonly used in microelectronics to denote the phenomenon of atom displacement in solid conductors caused by the flow of electrons (the term “displacement of atoms by the electron wind” is commonly used in this field). Therefore, this word does not appropriately describe the separation techniques discussed here. The terms electrodriven and electromigrative are, perhaps, too broad to be used to refer to these separation techniques. In conclusion, the term electrokinetic separation techniques seems to be the most precise name for these techniques, and will hereafter be taken as their official name, as it undoubtedly specifies the set of phenomena used in the separations.
The categorization of the dozens of electrokinetic separation techniques (ESTs) is highly necessary as they have important theoretical and practical differences. Consequently, the categorization of these techniques into three levels is proposed as it produces a simple and practical nomenclature. This categorization begins with the first level: the layout of the separation path, which can be either open (O) or toroidal (T) in shape (as previously described).
The second level of categorization is the platform where the ESTs are performed. Capillary (C), microchip (M) and slab (S) platforms are the most common. Flexible fused-silica microtubes, popularly known as capillaries, are already widely used to achieve high separation efficiencies in the open layout and are also starting to be used in the toroidal layout. Cylindrical capillaries are the most frequently used, but square and rectangular capillaries are also available. In the case of microchips, microchannels are etched onto slides of polymeric materials, glasses, fused silica (amorphous), and quartz (crystalline). They show great promise for the development of novel, multifunctional microstructures. The use of 3D printing allows an even larger number of techniques to be performed on this platform, both for basic research and an uncountable number of applications. The slab platform is normally made of a slab of gel, cellulose acetate, nylon membrane, or another porous substance. These macroscopic supports always have an anti-convective effect that prevents the sample from spreading due to convection. In addition, they usually play additional, well established, roles in the separation mechanisms.
The third and lowest level of categorization is the separation mode. This refers to the underlying molecular mechanism used to promote the separation of the analytes of interest, both among themselves and from any undesired sample interferant. The modes include affinity electrophoresis (AE), electrochromatography (EC), end-labeled free-solution electrophoresis (ELFSE), free-solution electrophoresis (FSE), isoelectric focusing (IEF), isotachophoresis (ITP), microemulsion electrokinetic chromatography (MEEKC), micellar electrokinetic chromatography (MEKC), and sieving electrophoresis (SE), to mention only a few. It is interesting to note that this nomenclature is currently normally used for the separation modes, as a result of the recommendations made by IUPAC.
When combining these three levels of categorization to accurately name, produce an acrynom, and describe an EST, the following intuitive rules are proposed: (1) the first word represents the first level of categorization; either open (O) or toroidal (T). (2) The second word specifies the platform: capillary (C), microchip (M), or slab (S). (3) The last word specifies the separation mode. A summary of the ESTs covered in this book is given in alphabetical order in the following table using this proposed nomenclature.
| Layout | Platform | Separation mode |
| (1) Open | (1) Capillary | (1) Affinity electrophoresis |
| (2) Toroidal | (2) Microchip | (2) Electrochromatography |
| (3) Slab | (3) End-labeled free-solution electrophoresis | |
| (4) Free-solution electrophoresis | ||
| (5) Isoelectric focusing | ||
| (6) Isotachophoresis | ||
| (7) Microemulsion electrokinetic chromatography | ||
| (8) Micellar electrokinetic chromatography | ||
| (9) Sieving electrophoresis |
The
