Analysis Combined with Specific Degradation

Methylation Analysis Combined with Specific Degradation 

Specific degradation of the carbohydrate chain has been extensively discussed in a few articles (for reviews, see Refs. 5, 7, 8,35, and 58). In the following discussion, we shall briefly present some applications that have been valuable in the sequence study of glycoprotein and glycolipid saccharides. 

Monosaccharide residues containing vicinal hydroxyl groups are oxidized by periodate, and are subsequently removed in the reduction-hydrolysis step. Therefore, the positions to which such monosaccharide residues are linked can be located by methylation analysis performed before, and after, Smith degradation. Alternatively,59 the oxidized and reduced sample is methylated, the ether hydrolyzed, and the product realkylated with CD3I or CH3CH2I. This kind of procedure can have advantages over that first described. For example, methylation before the hydrolysis step hinders the acetal protection of hydroxyl groups that can occur in acid hydrolysis.7 

An example of the use of Smith degradation for carbohydrate sequencing is a study on the sites of AcNeu-Gal-GlcNAc linkages to 

Acid-labile linkages of carbohydrates are commonly those of furano-sidic sugar residues or of deoxy monosaccharides.50,63 Consequently, neuraminic acid and fucose residues, which occur as terminal monosaccharides of protein- and lipid-linked glycans, are removed by hydrolysis with a dilute acid. The linkages formed by these two sugar residues can conveniently be differentiated by subjecting a portion of 

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V. Methylation Analysis Combined with Specific Degradation. 407... 

A number of approaches have been used to prove that a suspected transmitter resides in the presynaptic terminal of the pathway under study. These include biochemical analysis of regional concentrations of suspected transmitters, often combined with interruption of specific pathways, and microcytochemical techniques. Immunocytochemical techniques have proved very useful in localizing peptides and enzymes that synthesize or degrade nonpeptide transmitters. 

The heat from the injection port liner combined with the GC column flow causes the volatiles contained in the sample to be thermally desorbed directly onto the GC column. This reduces or eliminates interfering components of the sample matrix. An example of this additive specific extraction is shown in Figure 2-2. This technique also can be used to obtain the purity and identity of neat additive standards which are not readily soluble. By altering the injection port temperature, an analyst can extract various types of additives without thermally degrading the sample matrix. There are no limitations on the additive/polymer combinations which can be analyzed. Another advantage of this technique is that it requires only a few milligrams of sample typically 2-5 milligrams per analysis. 

As it has been shown earlier, the quantification of LCC linkages with traditional wet chemistry methods is limited mostly to relative quantification of carbohydrate sites linked to lignin. In contrast, our quantitative 2D NMR approach [21] allows for quantification of lignin sites involved in LCC linkages of different types. However, it does not provide information on the specific carbohydrate linkage sites yet. Therefore, a combination of quantitative NMR analysis and appropriate wet chemistry methods, such as a routine carbohydrate analysis, along with the methylation and DDQ oxidation degradation techniques, could be the best approach for comprehensive LCC analysis. 

Further notable developments include renewed interest in the use of laser source pyrolysis, particularly in combination with time-of-flight mass spectrometry, which offers the possibility of targeting particular chemical bonds for degradation, and the coupling of analytical pyrolysis and tandem mass spectrometry, to induce the fragmentation of specific pyrolysate ions. The latter has culminated in the construction of a number of novel instruments, including a pyrolysis ion trap mass spectrometer capable of MS analysis. 

Specifically for triazines in water, multi-residue methods incorporating SPE and LC/MS/MS will soon be available that are capable of measuring numerous parent compounds and all their relevant degradates (including the hydroxytriazines) in one analysis. Continued increases in liquid chromatography/atmospheric pressure ionization tandem mass spectrometry (LC/API-MS/MS) sensitivity will lead to methods requiring no aqueous sample preparation at all, and portions of water samples will be injected directly into the LC column. The use of SPE and GC or LC coupled with MS and MS/MS systems will also be applied routinely to the analysis of more complex sample matrices such as soil and crop and animal tissues. However, the analyte(s) must first be removed from the sample matrix, and additional research is needed to develop more efficient extraction procedures. Increased selectivity during extraction also simplifies the sample purification requirements prior to injection. Certainly, miniaturization of all aspects of the analysis (sample extraction, purification, and instrumentation) will continue, and some of this may involve SEE, subcritical and microwave extraction, sonication, others or even combinations of these techniques for the initial isolation of the analyte(s) from the bulk of the sample matrix. 

In an NMR analysis of the effects of /-irradiation induced degradation on a specific polyurethane (PU) elastomer system, Maxwell and co-workers [87] used a combination of both H and 13C NMR techniques, and correlated these with mechanical properties derived from dynamic mechanical analysis (DMA). 1H NMR was used to determine spin-echo decay curves for three samples, which consisted of a control and two samples exposed to different levels of /-irradiation in air. These results were deconvoluted into three T2 components that represented T2 values which could be attributed to an interfacial domain between hard and soft segments of the PU, the PU soft segment, and the sol... 

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