Method relating detector response to sample concentration in order to perform quantitative analysis. Standard solutions of a sample to be quantitated are prepared and equal volumes chromatographed. Peak heights or peak areas are plotted versus concentrations to produce a calibration curve.
Quantitative method for the determination of the actual amount of analyte captured and retrieved during the extraction process. Analyte recovery is based on an external standard that is added to the eluant at the end of the extraction procedure.
The absorbance at one wavelength divided by the absorbance at another wavelength. Used in HPLC as a qualitative tool and to help identify hidden peaks.
In Mass Spectrometry, the amount of a given ion present in a mass spectrum. Abundance may be expressed in either absolute or relative terms.
A measure of how close a measured value is to a known true value. Accuracy is assessed by means of reference samples and percent recoveries of spiked samples. Accuracy is the goal of quantitative analysis.
Also known as MeCN or methyl cyanide.MW 41.05, Boiling Point 81.60 C, Polarity Index 5.8, Solubility in water miscible in all proportions, UV cutoff 190nm. Common solvent used in reversed phase chromatography.
A support matrix capable of covalently binding a molecule.
In adsorption chromatography, the activity refers to the relative strength of the packing surface. Activity can be controlled by adding water or some other polar modifier, which is hydrogen bonded to the active site, causing a reduction in the surface activity.
In Mass Spectrometry, an ion formed within the ion source by the interaction of two species (usually an ion and a molecule). This is often the case in chemical ionization and results in ions present at mass-tocharge ratio greater than the molecular ion of the analyte.
Refers to the measurement of the retention time adjusted for the void volume, (t' R = tR - tM (or t0)).t' R is adjusted retention time, t R is th retention time of the peak, and t M is the retention time of the unretained peak.
Refers to the retention volume adjusted for the dead volume. V'R = VR - VM, where VM is the dead volume.
The stationary phase or packing material, especially when used in liquid-solid (adsorption) chromatography. Non-bonded normal phase packing material such as silica gel or alumina
The transfer of a solute from the mobile phase onto the adsorbent. Common inorganic adsorption packings include unmodified silica, alumina, graphitized carbon, hydroxyapatite, and zirconia. Organic polymer supports such as unmodified styrene divinylbenzene are also used. Refers to the interaction between a solute and the surface of an adsorbent. The forces involved can be weak (van der Waals forces) or strong (hydrogen bonds).
Relies on the adsorption process to effect a separation. Silica gel and alumina are used most frequently in this basic type of chromatography. IUPAC Definition: Separation is based mainly on differences between the adsorption affinities of the sample components for the surface of an active solid.
Relies on the adsorption process to effect a separation. Silica gel and alumina are used most frequently in this basic type of chromatography. IUPAC Definition: Separation is based mainly on differences between the adsorption affinities of the sample components for the surface of an active solid.
A specialized type of chromatography, which is actually selective filtration. Using bonded phases which have an affinity for - or an ability to adsorb - only certain types of molecules. Molecules that are not adsorbed are eluted unretained. The retained compound can be released in a purified form at a later stage.
A high molecular weight polysaccharide that is used as a separation medium in biochromatography. It is used in bead form in gel filtration chromatography (GFC) which have aqueous mobile phases.
An organic molecule which has an open carbon chain. It has no aromaticity (ring structures).
A measured portion of sample taken for analysis.
An adsorbent commonly used in adsorption chromatography. Aluminium oxide is a porous adsorbent with a slightly basic surface, which can have advantages over silica, which has an acidic surface.
The temperature outside the chromatographic system.
Known as Amino, NH2, Amino propyl silyl, or APS and can be employed as reversed phase, normal phase, or weak anion exchange material.
The target compound(s) that needs to be extracted, analyzed and quantitated. Often referred to as the target analyte(s). A substance that is to be determined, measured or analyzed.
Refers to the internal diameter of the HPLC Column. Analytical bore columns fit in the range between semi-prep and midbore,ie 3.5mm - 5.0mm diameter
Positively charged sorbent that retains anions.
Asymmetry describes the shape of a chromatographic peak. It is assumed peaks will have a Gaussian shape and that they are symmetrical. The peak asymmetry factor is the ratio (at 10% peak height) of the distance between the peak apex and the back side of the chromatographic curve and the front side of the curve.
Peak Asymmetry (As) = B / A. Where B is the horizontal distance from the apex to the tail at 10% height and A is the horizontal distance from the apex to the front at 10% height.
The first term in the van Deemter equation.
The pressure exerted at sea level by the atmosphere. Varies with weather, but for Mass Spectrometry purposes considered to be constant at 1.013 x 10^5Pa (760 Torr).
Refers to the process of removing a gap at the head of a column by tightening the end fitting so that it moves into the column and compresses the packing material. Requires a special column end fitting.
Refers to simply reversing the direction of the column and flushing with strong solvent to elute strongly held compounds and/or remove microparticulate accumulation at the inlet frit.
Is the pressure above gravity at the inlet end of the column. It is measured in: psi, bar, atm. kg/cm², kPa or Mpa. The current IUPAC convention is MPa.
The use of heat to accelerate the removal of gases and contaminants from a vacuum chamber. In addition to heat, flow can also be used to accelerate the cleaning process.
The area (or volume) in which the component(s) is located at any given time during elution from the column. Band is usually in the column; peak is the band signal on the chromatogram.
Broadening of the elution band of a solute at increasing elution times, because separations are an equilibrium process where the solute spends a fraction of the time interacting with the support and another fraction of the time dissolved in the mobile phase. Contributing effects to band broadening are diffusion and mass-transfer effects both in the mobile phase and in and on the support material.
Refers to broadening of a chromatographic peak as it moves through the liquid chromatograph. Dilution (bandspreading) occurs at various points due to dead volume in tubing, fittings, unions and connectors, the column itself as well as the detector cell.
The tendency of a chromatographic band to broaden as it passes through the chromatogram. Also known as band broadening or band dispersion. The measure of band spreading is band width (tw) or the number of theoretical plates in the column, N.
Spectroscopic term used to define the wavelength tolerance in nanometers of a 'monochromated' beam of light. ie Bandwidth of 4nm is +/- 2nm of selected wavelength. It is normally controlled by varying the width of slits placed between the monochromator and the flow cell. In chromatography a wide bandwidth may be desirable to accomodate components of differing lambda max. Conversely it can be desirable to have a narrow band width to increase selectivity.Bandwidth is very significant with fluorescence detection.
In HPLC a unit of pressure, equal to one atmosphere; also equal to about 15 pounds per square inch (psi) or 0.1 Megapascal.
In Mass Spectrometry, the largest peak in the spectrum. This peak is assigned a relative abundance of 100 percent. The base peak is frequently a different peak than the molecular ion. For a given molecule, the ion that is the base peak is influenced by the tye of ionization and ion source parameters (such as temperature, tuning parameters and type of mass spectrometer).
A term in Mass Spectrometry for the lowest pressure that a system (or pump) can reach.
An estimate of the signal level if the peak were not present. The portion of the chromatogram recording the detector response when only the mobile phase emerges from the column.
A defined quantity of material produced in a process or series of processes so that it is expected to be homogeneous within specified limits. In the case of continuous production a batch must correspond to a defined fraction of the production, characterised by its intended homogeneity. The batch size may be defined either by a fixed quantity or the amount produced in a fixed time interval.
The sum of the interstitial volume plus the pore volume of the sorbent within the column. Roughly synonymous with the terms void volume and dead volume.
A method devloped by Bruner, Emmett and Teller, for measuring surface area by determining the adsorption/condensation of liquid nitrogen in the pores of the solid phase support. The pore volume and pore size distribution can also be determined by using this method.
A systematic error inherent in a method or caused by some artifact or idiosyncrasy of the measurment system. Temperature effects and extraction inefficiences are examples of the first kind. Blanks, contamination, mechanical losses, and calibration errors are examples of the latter kinds. Bias may be both positive and negative, and several kinds can exist concurrently so that net bias is all that can be evaluated, except under special conditions.
A mobile phase composed of two solvents, such as methanol plus water. Most mobile phases are of this type.
A column made with non-ferrous materials for use with iron sensitive biomolecules. Biocolumns made of PEEK, Titanium and glass-lined tubing are available.
Indicates that the column or component will not adsorb or deactivate biomolecules such as proteins.
Examples of biomolecules include: biopolymers such as proteins and peptides, enzymes, polysaccharides, deoxyribonucleic acid (DNA), ribonucleic acid (RNA), and lower- molecular weight, non-polymeric compounds of biogenic origin such as simple sugars, vitamins, fatty acids, simple organic acids, adenosine triphosphate, etc.