Mass Determination

The mass determination of ionic species (atomic or polyatomic ions) in mass spectrometry is always a comparative measurement, which means the mass of an ionic species is determined with respect to reference masses of elements (or substances) used for mass calibration. The reference mass is thus acquired from the mass unit (m = In = 1/12) of the mass of the neutral carbon isotope (m = 1.66 X 10 kg). A mass calibration is easy to perform in solid-state mass spectrometry if the sample contains carbon (using carbon cluster ions with whole masses, as discussed above). The so-called doublet method was apphed formerly, e.g., ions and doubly charged Mg + forming a doublet at the same nominal mass number 12 were considered, where they are slightly displaced with respect to one another. The doublet method is no longer of relevance in modern inorganic mass spectrometry. Orientation in the mass spectra can be carried out via the matrix, minor and trace elements after mass calibration and by comparing the measured isotopic pattern of elements with theoretical values. 

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Fe]a B and Aebi U 1999 Moleoular mass determination by STEM and EFTEM a oritioal oomparison Micron 30 299-307... 

Ultimate resolving power (accurate mass determination) + +++... 

The techniques described thus far cope well with samples up to 10 kDa. Molecular mass determinations on peptides can be used to identify modifications occurring after the protein has been assembled according to its DNA code (post-translation), to map a protein structure, or simply to confirm the composition of a peptide. For samples with molecular masses in excess of 10 kDa, the sensitivity of FAB is quite low, and such analyses are far from routine. Two new developments have extended the scope of mass spectrometry even further to the analysis of peptides and proteins of high mass. 

Qualitatively, the spark source mass spectrum is relatively simple and easy to interpret. Most instrumentation has been designed to operate with a mass resolution Al/dM of about 1500. For example, at mass M= 60 a difference of 0.04 amu can be resolved. This is sufficient for the separation of most hydrocarbons from metals of the same nominal mass and for precise mass determinations to identify most species. Each exposure, as described earlier and shown in Figure 2, covers the mass range from Be to U, with the elemental isotopic patterns clearly resolved for positive identification. 

In carrying out a molar mass determination by freezing point depression, we must choose a solvent in which the solute is readily soluble. Usually, several such solvents are available. Of these, we tend to pick one that has the largest kf. This makes ATf large and thus reduces the percent error in the freezing point measurement From this point of view, cyclohexane or other organic solvents are better choices than water, because their kf values are larger. 

Electrospray ionization mass spectrometry (ESI-MS) is an analytical method for mass determination of ionized molecules. It is a commonly used method for soft ionization of peptides and proteins in quadmpole, ion-trap, or time-of-flight mass spectrometers. The ionization is performed by application of a high voltage to a stream of liquid emitted from a capillaty. The highly charged droplets are shrunk and the resulting peptide or protein ions are sampled and separated by the mass spectrometer. 

If it is possible to analyse end groups of a particular specimen of polymer, it may be possible to use the data to determine number average relative molar mass. If the molecules are branched the degree of branching can be measured from a combination of end group analysis and relative molar mass determination (determined by an alternative method). 

The molecular mass determined osmometrically corresponds to the formula S5O. The SO stretching vibration was observed in the infrared spectrum at 1119 cm (at -65 °G) indicating an exocyclic sulfoxide group similar to the one in SsO (see below). At -50 °G the solution of S5O may be kept for several days without decomposition which usually results in a Tyndall effect caused by a colloidal polymeric sulfuroxide which is the expected decomposition product. At 25 °G some decomposition already occurs within... 

The structure of Bosentan [30] and three of its metabolites are shown in Figure 5.45 and the product-ion spectra from the [M - - H]+ ions from these compounds in Figure 5.46. All show an ion at m/z 280 which might be assumed, simplistically, to share the same structure. Their accurate masses, determined by using a Q-ToF instrument, however, show that the ions from compounds (1)... 

Complex peptide mixmres can now be analyzed without prior purification by tandem mass spectrometry, which employs the equivalent of two mass spectrometers linked in series. The first spectrometer separates individual peptides based upon their differences in mass. By adjusting the field strength of the first magnet, a single peptide can be directed into the second mass spectrometer, where fragments are generated and their masses determined. As the sensitivity and versatility of mass spectrometry continue to increase, it is displacing Edman sequencers for the direct analysis of protein primary strucmre. 

The third described enzyme form with pH optimum about 4.7 [11, 4], we found in Fraction C - the fraction from carrot roots pulp (Fig. 2). We supposed that this form of exopolygalacturonase is relatively strongly bound on carrot cell walls and so it can be released only by higher salt concentrations. The approximative molecular mass determination on Superose 12 (Fig. 3c) showed the molecular mass about 50 000 for this form and the second, with more acidic pH optimum, form present in the fraction. The further characterization of these enzymes showed the exopolygalacturonase with pH optimum 4.7 to be identical with enzyme described sooner by Pressey and Avants [4] and exopolygalacturonase with pH optimum 3.8 to be identical with the enzyme from Fraction A. In conclusion, the exopolygalacturonase form with pH optimum 3.8 can be considered to be the main enzyme form present in carrot roots. 

Fig. 5. The approximate molecular mass determination of polygalacturonase [(0—0) - substrate 0.5% pectate, pH 4.6] and exopolygalacturonase [( — ) - substrate 1.0 pmol/ml of di(D-galactosiduronic) acid, pH 4.0] on Superose 12 column (FPLC device). Flow rate 0.5 ml/min. System 0.05 M phosphate buffer pH 7.0, 0.15 M NaCl. Standarts Ferritin (450 kDa), Katalase (240 kDa), Aldolase (158 kDa), Albumin (68 kDa), Albumin (45 kDa), Chymotrypsinogen A (25 kDa), Cytochrome C (12.5 kDa).

Another concept gaining prominence in the community is the use of identiflcation points , in which a confirmation is made when a certain number of assigned points are collected in an analysis.For example, each ion in standard MS counts for 1 point, and 1.5 points may be given for MS-MS product ions, or 2 points may be assigned for ions obtained with enough MS resuloution to achieve accurate mass determinations. 

What is the charge to mass Determine the ratio of electromagnetic tube and power supply... 

Fig. 4. Critical concentrations of polystyrene/toluene and polyacrylamide/water at 25 °C in relation to molar mass determined by viscometry and light scattering... 

Principles and Characteristics Mass spectrometry can provide the accurate mass determination in a direct measurement mode. For a properly calibrated mass spectrometer the mass accuracy should be expected to be good to at least 0.1 Da. Accurate mass measurements can be made at any resolution (resolution matters only when separating masses). For polymer/additive deformulation the nominal molecular weight of an analyte, as determined with an accuracy of 0.1 Da from the mass spectrum, is generally insufficient to characterise the sample, in view of the small mass differences in commercial additives. With the thousands of additives, it is obvious that the same nominal mass often corresponds to quite a number of possible additive types, e.g. NPG dibenzoate, Tinuvin 312, Uvistat 247, Flexricin P-1, isobutylpalmitate and fumaric acid for m = 312 Da see also Table 6.7 for m = 268 Da. Accurate mass measurements are most often made in El mode, since the sensitivity is high, and reference mass peaks are readily available (using various fluorinated reference materials). Accurate mass measurements can also be made in Cl... 

High resolution is used to determine the exact mass of an ion species in a mixture knowledge of the exact mass of an unknown substance allows its atomic composition to be established. Target analysis exact mass determination proves the presence of a particular ion species (compound) in a mixture. Mass spectrometry is perhaps the only method that can be used to find the empirical formulae of compounds that are not completely pure. 

DP-MS suffers from system saturation sample loads of a few ig are to be used. DP-ToFMS equipped with El and FI sources is a thermal separation technique for solids which allows exact mass determination (Section 6.3.3). In order to detect and characterise polymer fragments of higher molecular weight, techniques such as DCI, in which the sample is thermally desorbed by the filament on which it is directly deposited, and laser desorption... 

Applications With the current use of soft ionisation techniques in LC-MS, i.e. ESI and APCI, the application of MS/MS is almost obligatory for confirmatory purposes. However, an alternative mass-spectrometric strategy may be based on the use of oaToF-MS, which enables accurate mass determination at 5 ppm. This allows calculation of the elemental composition of an unknown analyte. In combination with retention time data, UV spectra and the isotope pattern in the mass spectrum, this should permit straightforward identification of unknown analytes. Hogenboom et al. [132] used such an approach for identification and confirmation of analytes by means of on-line SPE-LC-ESI-oaToFMS. Off-line SPE-LC-APCI-MS has been used to determine fluorescence whitening agents (FWAs) in surface waters of a Catalan industrialised area [138]. Similarly, Alonso et al. [139] used off-line SPE-LC-DAD-ISP-MS for the analysis of industrial textile waters. SPE functions here mainly as a preconcentration device. 

Wilson et al. [662-665] have described various prototype systems for total organic analysis devices. It has proved technically feasible to obtain UV, IR, NMR and MS spectra (together with atomic composition based on accurate mass determination) following RPLC separation. The fully integrated approach offers the benefit that one chromatographic run is required, thus ensuring that all of the spectrometers observe the same separation. Such multiple hyphenations might favour the analysis of complex mixtures for both confirmation of identity and structure determination (should this represent a cost-effective approach). Table 7.72 illustrates the main features of on-flow multiple LC hyphenation. 

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