Molecular-weight determination biopolymers

A relatively common feature of many problems involving molecular weight determination of biopolymers is that of association-dissociation equilibrium. Subunit structure of enzyme proteins is well recognized (1), and methods of dissociation of subunits to obtain monomer molecular weight are widely utilized (2). A previous paper described the application of an equilibrium gel partition method to the analysis of macromolecular association in a monomer-dimer case (3). The experimental parameters in a system utilizing the Sephadex series of gel filtra-... 

The advantage of ehemical derivatives has always been their solubility potential. This ability alone lends the biomaeromolecule to more thorough chemical characterization, for example, molecular weight determination by GPC, poorly achieved with chitin and challenging with chitosan. The use of lower moleeular weight materials would alleviate the situation somewhat, but truly soluble systems especially in water would transform the evaluation of these biopolymers that is vital for biomedical applications. Coupled with the potential for new desirable properties a chanical derivative can provide, this aspect wiU increasingly attract attention. 

SEC has its place in the purification and isolation of biopolymers, e.g. insulin, and other therapeutic drugs as a module in downstream processing [8, 9]. Mass spectrometry has, however, comprehensively displaced S EC for molecular weight determination. A current field of application is in the monitoring of conformational changes and aggregation of proteins. Micro-SEC with capillary columns has foimd application in the SEC of synthetic polymers in process analysis. 

Note that for the determination of molecular weight, the charge-state distribution is not of great importance as it does not affect the m/z value of the ion involved and thus the calculated molecular weight. If the conformational state of the biopolymer is of interest, however, the distribution of charged states is a fundamental consideration and any parameter likely to change this distribution must be carefully controlled. 

The application areas for LC-MS, as will be illustrated later, are diverse, encompassing both qualitative and quantitative determinations of both high-and low-molecular-weight materials, including synthetic polymers, biopolymers, environmental pollutants, pharmaceutical compounds (drugs and their metabolites) and natural products. In essence, it is used for any compounds which are found in complex matrices for which HPLC is the separation method of choice and where the mass spectrometer provides the necessary selectivity and sensitivity to provide quantitative information and/or it provides structural information that cannot be obtained by using other detectors. 

The polarity and thermal instability of biopolymers, together with the almost exclusive formation of singly charged ions renders APCl an inappropriate ionization technique for their study. Much of the early work involving electrospray ionization, on the other hand, was connected with the analysis of this type of molecule, in particular determining the molecular weight of proteins for which it is particularly effective. 

Classical MALDI-MS requires that the material should be soluble in a suitable solvent. A suitable solvent means a solvent that is sufficiently volatile to allow it to be evaporated prior to the procedure. Further, such a solvent should dissolve both the polymer and the matrix material. Finally, an ideal solvent will allow a decent level of polymer solubility, preferably a solubility of several percentage and greater. For most synthetic polymers, these qualifications are only approximately attained. Thus, traditional MALDI-MS has not achieved its possible position as a general use modern characterization tool for synthetic polymers. By comparison, MALDI-MS is extremely useful for many biopolymers where the polymers are soluble in water. It is also useful in the identification of synthetic polymers, such as PEO where the solubility requirements are fulfilled. Thus, for PEO we have determined the molecular weight distribution of a series of compounds with the separations in ion fragment mass 44 Da corresponding to CH2-CH2 units. 

The term food colloids can be applied to all edible multi-phase systems such as foams, gels, dispersions and emulsions. Therefore, most manufactured foodstuffs can be classified as food colloids, and some natural ones also (notably milk). One of the key features of such systems is that they require the addition of a combination of surface-active molecules and thickeners for control of their texture and shelf-life. To achieve the requirements of consumers and food technologists, various combinations of proteins and polysaccharides are routinely used. The structures formed by these biopolymers in the bulk aqueous phase and at the surface of droplets and bubbles determine the long-term stability and rheological properties of food colloids. These structures are determined by the nature of the various kinds of biopolymer-biopolymer interactions, as well as by the interactions of the biopolymers with other food ingredients such as low-molecular-weight surfactants (emulsifiers). 

T Vispersions of acrylic polymer beads in rapidly polymerizable liquids are important biomaterials (I, 2). The biocompatibility and functionality of dental restoratives, dental prostheses, and surgical prostheses depend on the mechanical properties of these biopolymers as well as on their physical and chemical constitution. This investigation was part of a continuing program to determine the influence of microstructural parameters on the mechanical properties of these multiphase systems. The effects of the volume fractions of dispersed phase and matrix, molecular weight of the matrix, chain length and concentration of crosslinkers, impact modifiers, and filler were studied in terms of microstructure, hard-... 

The incompatibility phenomenon relates to both the occupation of a volume of the solution by macromolecules and the weak repulsion between unlike macromolecules. Phase separation in mixed solutions of a large number of biopolymers studied is sensitive to entropy factors given by the excluded volume of the macromolecules. Phase behaviour strongly depends, therefore, on the molecular weight and the conformation of the macromolecules. The excluded volume effect that depends on the size and shape of the macromolecules determines the phase separation threshold, water partition between the phases of WIW emulsions and biopolymer activity in mixed solution (Tolstoguzov 1986, 1991, 1992). 

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