Glossary Of HPLC-LC Separation Terms – T

The listing should be helpful to those just starting in HPLC but it also can serve as a refresher for long-time users in the field.

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T

Tailing: The phenomenon in which a normal Gaussian peak has an asymmetry factor greater than 1. The peak will have an extended trailing edge. Tailing is caused by packing sites that have both a stronger-than-normal retention for the solute and slower desorption kinetics. A typical example of a tailing phenomenon would be the strong adsorption of amines on the residual silanol groups of a low-coverage reversed-phase packing at intermediate pH values. Tailing also can result from injecting an excessive mass or sample, badly packed columns, excessive extracolumn volume, poor fittings, excessive detector volume, and slow detector response. See Figure 1.

Tailing factor: U.S. Pharmacopeia measure of peak asymmetry defined as the ratio of the peak width at 5% of the apex to twofold the distance from the apex to the 5% height on the short time side of the peak. Greater than unity for tailed peaks. See also asymmetry factor.

Temperature programming: Changing column temperature as a function of time during the separation. Rarely used in HPLC; if so, usually in a stepwise manner.

Ternary mobile phase: Mobile phase that is a mixture of three solvents or buffers.

Theoretical plate (N): A concept described by Martin and Synge. Relates chromatographic separation to the theory of distillation. Length of column relating to this concept is called height equivalent to a theoretical plate. See also HETP. Plates are calculated as N = 16(V R/w b)2 = 16 (t R/w b)2, where V R is the retention volume, w b is the width at the peak base, and t R is the retention time. See also N.

Thermally tuned tandem column chromatography: A form of LC in which two columns with distinctly different selectivities are placed in tandem and operated at two temperatures to optimize the resolution or analysis speed. Both columns use a common eluent, and the entire sample passes through both columns and is detected with a single detector. It is not a two-dimensional technique because each sample component provides only one peak.

Titania: An uncommon adsorbent used in adsorption chromatography.

t m See migration time.

t M Holdup time.

Tortuosity or tortuosity factor: A packed-column property that controls the inhibition of longitudinal diffusion of the solute as it diffuses along the column axis. The B term in the van Deemter equation is proportional to the tortuosity. See also B term, γ, and molecular diffusion term.

Total mobile-phase volume (V t ): The total volume of mobile phase in an SEC column. Also known as totally included volume. Same as V M .

Total permeation volume (V p ): The retention volume of an SEC packing in which all molecules smaller than the smallest pore will be eluted. In other words, all molecules totally permeate all of the pores at V p and are eluted as a single peak. Same as V M .

Total porosity (´ T ): Ratio of the total volume of mobile phase in the column to the total column volume; ´ = V M/V c = ´e + ´i (1 - ´e); where V M is the mobile-phase volume, V c is the column volume, ´e is the interstitial porosity, and ´i is the intraparticle porosity.

Totally porous packing: The stationary phase is a porous matrix, and solutes penetrate the porous matrix to interact with the stationary phase.t R See retention time.

t R 9See adjusted retention time and retention time.

Trace enrichment: Technique in which trace amounts of compounds are retained on an HPLC or precolumn packing out of a weak mobile phase or solution and then are eluted by adding a stronger mobile phase in a concentrated form. The technique has been applied most successfully in the concentration of trace amounts of hydrophobic compounds such as poly-nuclear aromatic hydrocarbons from water using a reversed-phase packing. A strong solvent such as acetonitrile will elute the enriched compounds.

Triethylamine: A very common additive used to block silanol groups in reversed-phase chromatography when separating basic analytes.

Trifluoroacetic acid: A very common additive in reversed-phase chromatography for peptides and proteins.

Tryptic digestion: A method for selectively and reproducibly dissecting peptide chains of proteins to yield a characteristic pattern of smaller units that enables analysis of the parent protein by gradient elution reversed-phase LC.

Turbulence: The state in which fluid velocity fluctuates randomly at a point. See also Reynolds number and turbulent flow.

Turbulent flow: A form of fluid motion in which the flow ceases to be smooth and steady and becomes chaotic and fluctuates with time. It is characterized by a pressure drop significantly higher than what would be extrapolated from the laminar region to achieve the same volumetric flow rate.

Turbulent flow chromatography: Chromatography performed at very high linear velocities with large particles under conditions using high Reynolds numbers. At these conditions, the H versus ν curves show a decrease in H as ν increases. See Figure 2.

t w See bandwidth.

Two-dimensional chromatography: A procedure in which part or all of the separated sample components are subjected to additional separation steps. It can be performed by conducting a particular fraction eluted from the first column into a second column or system that has a different separation characteristic. It includes techniques such as two-dimensional TLC using two eluent systems in which the second eluent is applied after rotating the plate through 90°. It also includes LC followed by GC and one LC mode followed by a different mode such as reversed-phase chromatography followed by SEC. See also multidimensional chromatography.

Type A silica: Silica gel formed by gelling soluble silicates. Generally has higher acidity, higher surface area and porosity, more trace metals, and poorer high-pH stability than Type B silicas.

Type B silica: See sol gel.t 0 See void time.

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