Molecular mass, determination distribution

The observation of molecular size or polydispersity and the subsequent determination of relative molecular mass, (MJ or molecular mass (weight) distribution (MWD), is the most common analytical application of SEC. The goal of these types of experiments is to either observe the solvated size of one or more molecular species or to observe the distribution of sizes present in a mixture... 

The chains of the produced polymer are lengthened by combination, but not by disproportionation. This affects the molecular mass distribution but the differences are not very large, differing by a factor of 2 at most. Due to the inaccuracies in molecular mass determinations, it is almost impossible to make estimates of the relation between termination and disproportionation from the distributions. Even labelling of the initiator and determination of the average number of its fragments in a macromolecule (one for disproportionation and two for combination) is usually unsafe because of transfer. 

Relative molecular mass (RMM) distributions for components of biochemical and polymer systems can be determined with a 10% accuracy using standards. With biochemical materials, where both simple and macro-molecules may be present in an electrolyte solution, desalting is commonly employed to isolate the macromolecules. Inorganic salts and small molecules are eluted well after such materials as peptides, proteins, enzymes and viruses. Desalting is most efficient if gels with relatively small pores are used, the process being more rapid than dialysis. Dilute solutions of macromolecules can be concentrated and isolated by adding dry gel beads to absorb the solvent and low RMM solutes. 

When the averaged molecular masses determined by CGE are compared with those given by the suppliers (measured via SEC), molecular masses determined by SEC are always lower. When the CGE values are compared with results from MALDI-TOF measurements a closer correlation could be found. This may be an indication that in SEC polymer adsorption on the matrix was the reason for higher elution volumes, resulting in lower molecular masses. Of course, polydispersity is also affected by adsorption. Consequently CGE is a fast and reliable method for characterization of the molecular masses of polyelectrolytes and their distribution. 

The determination of polymer molecular mass and distribution by MALDI-MS requires not only accurate mass measurement but also quantitative measurement of ions. The equations used for molecular mass determination and polydispersity (PD) calculation are ... 

The analysis of a broad-polydispersity polymer (PD > 1.2) can be carried out by combining GPC or size-exclusion chromatography (SEC) with MALDI-MS [161-172]. In this approach, a wide-polydispersity polymer is first separated by GPC and fractions at a defined time interval are collected. The time interval is properly chosen so that the individual fraction would contain only a narrow-polydispersity (PD < 1.2) polymer, which can then be analyzed by MALDI-MS for accurate molecular mass determination. The molecular mass information generated from the MALDl analysis of aU individual fractions can be used to convert the time domain in the GPC chromatogram into a mass domain. The polymer distribution can be determined from this chromatogram. 

Polymer characterization usually requires a combination of several analytical tools such as NMR and GPC. Today, a number of analytical techniques exist that can provide molecular mass, structure, and composition information, and MALDI-MS is now emerging as a powerful method for polymer characterization. Some demonstrated advantages of the technique include the ability to determine average molecular mass and distributions without the need of polymer standards and with high speed, precision, and accuracy, to analyze polymer mixtures with minimiun sample work-up, and to provide structural and compositional information via... 

In particular, the ensemble of a new phase during phase separation proceeding in a multi-component system belongs to such systems. In this case, the turbidity spectrum method provides the determination of the mass-volume new-phase particle concentration, as well as the degree of phase transition this can be used to construct the molecular-mass polymer distribution function (the method of spectroturbidimetric titration for polymer solutions—subsection 3.2.3) and for a phase analysis (identification) of the pheise separation type in polymer. systems (paragraph 3.6.2.5, sections 6.2 and 6.4). 

Flash desorption of oligomers relies on kinetic competition between evaporation and thermal decomposition. Molecular ion signals from the MS have been used to determine the average molecular masses and distribution of oligomers, whereas expert systems can be used to establish mechanisms for the thermal degradation of polymers, e.g. to determine the relationships between polymer structure and the corresponding Py-MS spectra. 

The broad pore size distribution of Sepharose makes it well apt for the analysis of broad molecular mass distributions of large molecules. One example is given by the method for determination of MWD of clinical dextran suggested in the Nordic Pharmacopea (Nilsson and Nilsson, 1974). Because Superose 6 has the same type of pore size distributions as Sepharose 6, many analytical applications performed earlier on Sepharose have been transformed to Superose in order to decrease analysis time. However, Sepharose is suitable as a first try out when no information about the composition of the sample, in terms of size, is available. 

Fig. 4. Molecular mass distribution of Fraction A purified on Concanavalin A -cellulose on Superose 12 column. Buffer - 0.05 M phosphate, pH 7.0, 0.15 M NaCl, fraction size 0.5 ml/min. Exopolygalacturonase activity determined with penta(D-galactosiduronic) acid pH 5.0 and pH 3.8 (0—0)-...

The corresponding liquid-phase chemistry can be used to promote ion formation by appropriate choice of solvent and pH, salt addition to form M.Na+ or M.NH4+, and postcolumn addition of reagents. The primary applications of ESI-MS are in the biopolymer field. The phenomenon of routine multiple charging is exclusive to electrospray, which makes it a very valuable technique in the fine chemical and biochemical field, because mass spectrometers can analyse high-molecular-mass samples without any need to extend their mass range, and without any loss of sensitivity. However, with ESI, molecules are not always produced with a distribution of charge states [137], Nevertheless, this phenomenon somehow complicates the determination of the true mass of the unknown. With conventional low-resolution mass spectrometers, the true mass of the macromolecule is determined by an indirect and iterative computational method. 

SEC-GC-FID, according to Figure 7.40, has been used to carry out the simultaneous determination of the polymer average molecular masses and molar mass distribution and the concentration of additives [984]. The effluent was split and adsorbed on PTV packing material before GC analysis. The choice of PTV... 

A measure of the breadth of the molecular mass distribution is given by the ratios of molecular mass averages. The most commonly used ratio Mw/Mn — H, is called the polydispersity index. Wiegand and Kohler discuss the determination of molecular masses (weights) and their distributions in Chapter 6. 

Since it may be assumed that part of poly(3HB) synthesized is degraded during accumulation, that the equilibrium determines the content of poly(3HB), the molecular mass, and molecular mass distribution, a detailed analysis of the regulation of the poly(3HB) cycle will be useful for a better understanding as well as optimization of industrial production of poly(3HB). 

An analytical ultracentrifugation method for determining the molecular mass, diffusion coefficient, and/or state of oligomerization of a macromolecule by conducting sedimentation conditions to establish an equilibrium distribution of the macromolecule from the meniscus to the bottom of the observation cell. 

Selected entries from Methods in Enzymology [vol, page(s)] Association constant determination, 259, 444-445 buoyant mass determination, 259, 432-433, 438, 441, 443, 444 cell handling, 259, 436-437 centerpiece selection, 259, 433-434, 436 centrifuge operation, 259, 437-438 concentration distribution, 259, 431 equilibration time, estimation, 259, 438-439 molecular weight calculation, 259, 431-432, 444 nonlinear least-squares analysis of primary data, 259, 449-451 oligomerization state of proteins [determination, 259, 439-441, 443 heterogeneous association, 259, 447-448 reversibility of association, 259, 445-447] optical systems, 259, 434-435 protein denaturants, 259, 439-440 retroviral protease, analysis, 241, 123-124 sample preparation, 259, 435-436 second virial coefficient [determination, 259, 443, 448-449 nonideality contribution, 259, 448-449] sensitivity, 259, 427 stoichiometry of reaction, determination, 259, 444-445 terms and symbols, 259, 429-431 thermodynamic parameter determination, 259, 427, 443-444, 449-451. 

A polymer, even in its pure state, corresponds to a distribution of macromolecules with different masses. Exclusion chromatography can determine the distribution in molecular weights and the most probable mass and the mean mass. For this type of application, a calibration curve is made using macromolecules of known masses by plotting the retention times (or volumes) on the abscissa and the logarithms of the molecular masses on the ordinate. As can be seen in Fig. 7.5, a linear relationship is obtained. Using this graph, an approximate mass of the unknown can be determined by use of the retention time (volume). This assumes that the mass and the molecular volumes are directly related. 

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