Mass calibration compound

Any mass spectrometer requires mass calibration before use. However, the procedures to perform it properly and the number of calibration points needed may largely differ between different types of mass analyzers. Typically, several peaks of well-known m/z values evenly distributed over the mass range of interest are necessary. These are supplied from a well-known mass calibration compound or mass reference compound. Calibration is then performed by recording a mass spectrum of the calibration compound and subsequent correlation of experimental m/z values to the mass reference list. Usually, this conversion of the mass reference list to a calibration is accomplished by the mass spectrometer s data system. Thereby, the mass spectrum is recalibrated by interpolation of the m/z scale between the assigned calibration peaks to obtain the best match. The mass calibration obtained may then be stored in a calibration file and used for future measurements without the presence of a calibration compound. This procedure is termed external mass calibration. 

Note The numerous ionization methods and mass analyzers in use have created a demand for a large number of calibration compounds to suit their specific needs. Therefore, mass calibration compounds will occasionally be addressed later in the chapters on ionization methods. It is also not possible to specify a general level of mass accuracy with external calibration. Depending on the type of mass analyzer and on the frequency of recalibration, mass accuracy can be as high as 1 mmu or as low as 0.5 u. 

Mass Calibration (time-of-flight) A means of determining m/z values from their times of detection relative to initiation of acquisition of a mass spectrum. Most commonly this is accomplished using a computer-based data system and a calibration file obtained from a mass spectrum of a compound that produces ions whose m/z values are known. 

If high-resolution measurements are performed in order to assign elemental compositions, internal mass calibration is almost always required. The calibration compound can be introduced from a second inlet system or be mixed with the analyte before the analysis. Mixing calibration compounds with the analyte requires some operational skills in order not to suppress the analyte by the reference or vice versa. Therefore, a separate inlet to introduce the calibration compound is advantageous. This can be achieved by introducing volatile standards such as PFK from a reference inlet system in electron ionization, by use of a dual-target probe in fast atom bombardment, or by use of a second sprayer in electrospray ionization. 

Brinded, K.A. Tiller, P.R. Lane, S.J. Triton X-100 As a Reference Compound for Ammonia High-Resolution CI-MS and as a Tuning and Calibration Compound for Thermospray. Rapid Commun. Mass Spectrom. 1993, 7, 1059-1061. 

Mass Calibration The process by which the mass analyzer is calibrated such that a measured and displayed m/z is accurate. Well-characterized calibration compounds are utilized, and measured m/z values for these compounds are compared to theoretical m/z values. Calibrants commonly used include various polymeric species (such as polypropylene glyol, or PPGs poly tyrosine (poly-t)) or fluorinated species (perfluorokerosene or PFK) but can be any compound or mixture (Nal/KI) of compounds properly characterized for MS. 

Lock Mass Similar to internal calibration. The lock mass compound is monitored during analysis of the unknown, and the mass calibration is adjusted based on the comparison of the measured m/z and the theoretical m/z for the lock mass compound. If multiple lock mass compounds are used across the m/z range, the process effectively becomes internal calibration. Lock mass compound(s) can be introduced into the LC-MS source via a tee into the LC flow or sheath liquid inlet or dedicated sprayer. 

The LTQ-Orbitrap has resolution and mass accuracy performance close to that of the LTQ-FTICR. As shown in Table 5.3 (column 4), LTQ-Orbitrap accurate mass measurements, using external calibration, for a set of 30 pharmaceutical compounds resulted in less than 2.3 ppm error. The data were acquired with a 4-min, 1-mL/min-flow-rate, positive-mode LC-ESI-MS method where all measurements were performed within 5h from mass calibration. Mass accuracies below 2-3 ppm, and often below 1 ppm, can be routinely achieved in both the positive- and negative-ion mode (Table 5.3, columns 4 and 5). The long-term mass stability of the LTQ-Orbitrap is not as consistent as observed for the LTQ-FTICR-MS, and the Orbitrap requires more frequent mass calibration however, mass calibration is a routine procedure that can be accomplished within 5-10 min. Figure 5.7 displays a 70-h (external calibration) mass accuracy plot for three negative ions collected with a LTQ-Orbitrap where the observed accuracy is 2.5 ppm or better with little mass drift for each ion. Overall, for routine accurate mass measurements on the Orbitrap, once-a-week calibration (for the desired polarity) is required however, considering the ease of the process, more frequent external calibration is not a burden. 

This sort of correction should be helpful for improving specific mass measurements for GC/FTMS analyses where it is possible to add a calibrant compound to the mixture, or where some component is known to be present. Unlike the method where a static... 

Using a horizontal 4.7 Tesla cryoshimmed supercon magnet with a bore diameter of 15 cm, we have obtained two-parameter mass calibrations with absolute mass accuracies of better than 1.5 ppm without, and better than 0.4 ppm with an internal calibrant over a mass range from 18 to 502 amu (10). Absolute accuracies with an internal calibrant are slightly better because the calibrant and the unknown compound are measured under identical physical conditions, in particular with the exact same number of ions in the cell, resulting in identical space-charge shifts (2). 

Caroli and his co-workers [14] published multielement speciation results including data about Mn speciation. These data regarded samples from 60 mothers (approximately day 30 after delivery) living in different areas of Italy. The investigation was based on SEC fractionation using a mass calibrated column and total concentration of Mn was found to be around 3 p,g l-1. They found 28 percent of Mn in the void fraction (>2000 kDa), 30 percent bound to the nonprotein fraction, (relatively LMW compounds), and the remaining Mn was found at low levels in the other fractions. 

M. Moini, B.L. Jones, R.M. Rogers, L. Jiang, Sodium trifiuoroacetate as a tune/calibration compound for positive- and negative-ion ESI-MS in the mass range of 100-4000Da, J. Am. Soc. Mass Spectrom., 9 (1998) 977. 

Photolytically degraded PBO was re-examined by GC-EIMS at a higher mass spectrometer resolution (5000) with PFK in the mass spectrometer source as an internal mass calibrant. This procedure permits the accurate mass measurement of molecular and key fragment ions and hence the determination of their elemental compositions and fragmentation pathways. This information, along with the molecular weight information (GC-CIMS) and the low-resolution GC-EIMS spectrum, allows a structure to be proposed for each of the degradation compounds observed. 

The determination of differences in isotopic ratios requires very precise measurements. The combustion step for the sample preparation is usually carried out immediately prior to the injection into the MS. There exist instruments which associate in line a gas chromatograph, a tubular combustion oven, containing copper oxide heated to 800 °C, and a low-resolution MS equipped with several Faraday detectors, each collecting the signal corresponding to a specific mass. A calibration compound is co-injected with the product to be studied. 

Although the determination of the metal or metalloid is relatively straightforward, the complete identification of the ligand(s) presents some challenge. A systematic approach is required if the ligands are to be completely characterised. As a first step after the separation of the constituents, the column should be calibrated with compounds of known molecular mass. Calibration kits are obtainable from most manufacturers, however, if... 

The mass spectrum of calibration compound, FC-43, as shown in Figure 15.15, is well-known to mass spectrometry practitioners because it is seen frequently during... 

The most common use of batch inlets is to introduce a controlled flow of compounds for calibrating the mass scale. Frequently used calibration compounds include perfluorotributylamine (FC-43, heptacosa) and perfluorokerosene (PFK) both are effective with electron ionization (El) but give limited responses in chemical ionization (Cl). If used for calibration in Cl, or to provide lock masses (Section 3.1.1), the concentration of the reagent gas must be reduced. This reducation somewhat compromises the effectiveness of the Cl process. 

A low-resolution mass measurement is a simple procedure and can be performed with most of the mass spectrometric systems discussed in Chapter 3. The instrument is set up at a resolving power (RP) above 1000. At this low resolving power, the molecular ions of most organic compounds that differ by a unit mass are well separated. The mass spectrometer is first mass-calibrated with an external calibration procedure. During the calibration scan, the computer stores the peak centroid (the center of gravity) time and the area of each peak. The mass of the ion is related exponentially to the peak centroid time ... 

A MALDI-FT-MS-based automated system for rapid screening of a large array of compounds is also available [109]. In this approach, the library compounds were mixed with a suitable matrix and were deposited on an auto-indexed multiple-sample disk. The matrix-sample mixture was ionized by irradiation with UV nitrogen laser (337 nm), and mass calibrant ions were generated by electron ionization of perfluorotributyl amine. 

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