An internal standard in

analytical chemistry Analytical chemistry studies and uses instruments and methods to separate, identify, and quantify matter. In practice, separation, identification or quantification may constitute the entire analysis or be combined with another method. Separati ...

is a

chemical substance A chemical substance is a form of matter having constant chemical composition and characteristic properties. Some references add that chemical substance cannot be separated into its constituent elements by physical separation methods, i.e., wi ...

that is added in a constant amount to samples, the blank and

calibration In measurement technology and metrology, calibration is the comparison of measurement values delivered by a device under test with those of a calibration standard of known accuracy. Such a standard could be another measurement device of know ...

standards in a

chemical analysis Analytical chemistry studies and uses instruments and methods to separate, identify, and quantify matter. In practice, separation, identification or quantification may constitute the entire analysis or be combined with another method. Separati ...

. This substance can then be used for calibration by plotting the ratio of the

analyte An analyte, component (in clinical chemistry), or chemical species is a substance or chemical constituent that is of interest in an analytical procedure. The purest substances are referred to as analytes, such as 24 karat gold, NaCl, water, etc. ...

signal to the internal standard signal as a function of the analyte concentration of the standards. This is done to correct for the loss of analyte during sample preparation or sample inlet. The internal standard is a compound that is very similar, but not identical to the chemical species of interest in the samples, as the effects of sample preparation should, relative to the amount of each species, be the same for the signal from the internal standard as for the signal(s) from the species of interest in the ideal case. Adding known quantities of analyte(s) of interest is a distinct technique called

standard addition The method of standard addition is a type of quantitative analysis approach often used in analytical chemistry whereby the standard is added directly to the aliquots of analyzed sample. This method is used in situations where sample matrix also ...

, which is performed to correct for

matrix effect In chemical analysis, matrix refers to the components of a sample other than the analyte of interest. The matrix can have a considerable effect on the way the analysis is conducted and the quality of the results are obtained; such effects are calle ...

s. This ratio for the samples is then used to obtain their analyte concentrations from a calibration curve. The internal standard used needs to provide a signal that is similar to the analyte signal in most ways but sufficiently different so that the two signals are readily distinguishable by the instrument. For example,

deuterated Isotopic labeling (or isotopic labelling) is a technique used to track the passage of an isotope (an atom with a detectable variation in neutron count) through a reaction, metabolic pathway, or cell. The reactant is 'labeled' by replacing specific ...

chlorobenzene Chlorobenzene is an aromatic organic compound with the chemical formula C6H5Cl. This colorless, flammable liquid is a common solvent and a widely used intermediate in the manufacture of other chemicals. Uses Historical The major use of chlorob ...

(C₆D₅Cl) is an internal standard used in the analysis of volatiles on GC-MS because it is similar to

Chlorobenzene Chlorobenzene is an aromatic organic compound with the chemical formula C6H5Cl. This colorless, flammable liquid is a common solvent and a widely used intermediate in the manufacture of other chemicals. Uses Historical The major use of chlorob ...

but does not occur naturally.

Norleucine Norleucine (abbreviated as Nle) is an amino acid with the formula CH3(CH2)3CH(NH2)CO2H. A systematic name for this compound is 2-aminohexanoic acid. The compound is an isomer of the more common amino acid leucine. Like most other α-amino acids ...

is also a popular internal standard for the analysis of amino acids via GC-MS. In

NMR spectroscopy Nuclear magnetic resonance spectroscopy, most commonly known as NMR spectroscopy or magnetic resonance spectroscopy (MRS), is a spectroscopic technique to observe local magnetic fields around atomic nuclei. The sample is placed in a magnetic fiel ...

, e.g. of the nuclei ¹H, ¹³C and ²⁹Si, frequencies depend on the magnetic field, which is not the same across all experiments. Therefore, frequencies are reported as relative differences to the internal standard

tetramethylsilane Tetramethylsilane (abbreviated as TMS) is the organosilicon compound with the formula Si(CH3)4. It is the simplest tetraorganosilane. Like all silanes, the TMS framework is tetrahedral. TMS is a building block in organometallic chemistry but also ...

(TMS). This relative difference to TMS is called ''chemical shift'', and measured in parts per million. In practice, the difference between the signals of common solvents and TMS are known, and since modern instruments are capable of detecting the small quantities of protonated solvent present in commercial deuterated solvents no TMS need be added. By specifying the lock solvent to be used, modern spectrometers are able to correctly reference the sample; in effect, the solvent itself serves as the internal standard. In

chromatography In chemical analysis, chromatography is a laboratory technique for the separation of a mixture into its components. The mixture is dissolved in a fluid solvent (gas or liquid) called the ''mobile phase'', which carries it through a system (a ...

, internal standards are used to determine the concentration of other analytes by calculating

response factor Response factor, usually in chromatography In chemical analysis, chromatography is a laboratory technique for the separation of a mixture into its components. The mixture is dissolved in a fluid solvent (gas or liquid) called the ''mobile p ...

. The internal standard selected should be again similar to the analyte and have a similar retention time and similar derivitization. It must be stable and must not interfere with the sample components.

References

* Skoog, Douglas A. (1998). ''Principles of Instrumental Analysis'': Introduction, pp. 18. Harcourt Brace. * {{GoldBookRef, file=I03108, title=internal standard Analytical chemistry