Hydrolysis of polysaccharides

Much of the information on acid hydrolysis of polysaccharides is in the form of unpublished, empirical observations, although inferences (such as, that during the acid hydrolysis of guaran, the D-galactopyranose residues are readily removed, whereas D-mannose is set free somewhat more slowly ) may be found in the literature. 

The hydrolysis of cellulose was reported by Calvert in 1855. A more systematic investigation was reported by Girard in 1875. Since then, many investigations have been made (particularly of the kinetics), and several reviews have appeared.  

Early work on the hydrolysis of starch was reviewed by Rolfe and Defren, who made the first, systematic investigation (following the report by Kirchoffi in 1811 that acid hydrolysis of starch yields a sugarlike sub- 

The percentage of products formed after a particular degree of hydrolysis can be examined. Theoretical models have been proposed for the random hydrolysis of polysaccharides. When the theoretical (calculated) 

Additional evidence includes the fact that the initial rate of hydrolysis of the cyclodextrins is much lower than the rate of hydrolysis of the corresponding malto-oligosaccharides. Whereas the hydrolysis of maltose and other disaccharides follows normal, first-order kinetics, the rate constant for the hydrolysis of amylose increases with time, until it becomes the same as that of maltose, perhaps because of an increase in non-reducing ends brought about by slow, internal cleavages. There is a decrease in the hydrolysis rate-constant of cellodextrins with an increase in 

Many authors have commented upon the problems associated with the hydrolytic step, which has been considered to be the main source of loss in carbohydrate analysis.19 The fact that, in determining uronic acids, differing results were obtained, depending on whether hydrolysis or methanolysis was employed, led others20 to conclude that 

Certain polysaccharides are normally hydrolyzed with mineral acid, usually sulfuric acid, either by direct refluxing with dilute acid or by preliminary dissolution in concentrated acid. Typical procedures have been described, and the associated problems discussed.22,23 Although prior solution of the polysaccharide in 72% sulfuric acid is a standard procedure,24 it has been shown that part of the carbohydrate may become sulfated, leading to erroneous results.23 When noncrystalline polysaccharides are being hydrolyzed, the treatment with 72% acid may be slightly modified.26 In special situations, oxidative hydrolysis, for example, of carrageenan, may be achieved by using sulfuric acid in the presence of bromine.27 

Nitric acid is less commonly used for hydrolysis, but, in combination with urea, has been recommended for polysaccharides containing uronic acid residues28 and it has been used in a study of apple pectin.29 

Trifluoroacetic acid is volatile, and thus readily removed. This acid was used by Albersheim and coworkers for the hydrolysis of plant cell-walls,39 and has since been employed for cell walls,40-43 plant mucilages,44 blood-group oligosaccharides,45 peptidogalactoman-nans,46 heparin,47 and disaccharides in blood and urine.48,49 It has also been suggested as an alternative to 6 M hydrochloric acid in the determination of amino sugars,50 and for the hydrolysis of polyalcohols produced by periodate oxidation of polysaccharides.503 Lee 

Hough and coworkers50c compared the use of hydrochloric, sulfuric, and trifluoroacetic acids for hydrolysis, and confirmed that hydrochloric acid gives lower recoveries than the other two acids. Also, they recommended that hydrolyses be conducted in an atmosphere of nitrogen. 

---

For the most part, low molecular weight carbohydrates of commerce are made by depolymerization via enzyme- or acid catalyzed hydrolysis of polysaccharides. Only sucrose and, to a very much lesser extent, lactose, both disaccharides, are commercial low molecular weight carbohydrates not made in this way. 

For many of the organic materials in seawater, some form of chemical pretreatment is necessary before analysis is possible. The obvious cases are the hydrolysis of polysaccharides and proteins before the analysis for monomeric constituents, and the formation of volatile derivatives to permit analysis by gas chromatography. These methods will be discussed further in Chap. 9. 

Analysis of the rates of decomposition of monosaccharides by 2 M TFA has been accomplished in two studies by continuing hydrolysis of polysaccharides after most of the monosaccharides had been liberated. Albersheim and coworkers studied the hydrolysis of pinto-bean hypo-cotyl cell-walls (10 mg/mL) for up to 6 h at 121°, although the monosaccharides, except glucose and mannose, had been liberated after 1 h. After 6 h, >50% of the xylose and arabinose and >25% of the galactose, rham-nose, and fucose had been decomposed however, as most of the monosaccharides are liberated within 1 h, these rates of decomposition are tolerable. 

In an attempt to make this account as complete as possible, certain related aspects, such as the hydrolysis of polysaccharides, are briefly discussed, but, in general, the only references cited are to those papers that have also mentioned use of gas-liquid chromatography. These ancillary sections must, therefore, not be considered exhaustive in treatment, but rather to be representative. 

In the literature, there are several examples of partial hydrolysis of polysaccharides or glycoconjugates containing 2-amino-2-deoxyhex-ose residues. One example is the hydrolysis of carboxyl-reduced, N deacetylated chondroitin, with the formation of 4-0-(2-amino-2-deoxy-/3-D-galactopyranosyl)-D-glucose.51 Another is the isolation of 6-0-(2-amino-2-deoxy-/8-D-galactopyranosyl)-D-galactose after N-de-acetylation and partial hydrolysis of the pentasaccharide (19) obtained... 

However, the data available suggest that the results obtainable from mass spectrometry will considerably facilitate the structural analysis of oligosaccharides and thus permit direct investigation of the products of partial hydrolysis of polysaccharides on the oligomer level. 

Wohl4 was the first to refer to the formation of higher saccharides by the action of acids on monosaccharides as reversion. This process, the acid-catalyzed formation of glycosidic bonds between sugar residues, constitutes the most rudimentary form of condensation polymerization. Reversion of oligosaccharides has also been observed. 0 The relationships between the hydrolysis of polysaccharides and the reversion of mono- and oligo-saccha-rides is illustrated for amylose in Fig. 1. 

Chapters 17 through 21 deal with carbohydrate-enzyme systems. Hehre presents some new ideas on the action of amylases. Kabat presents some new immunochemical studies on the carbohydrate moiety of certain water-soluble blood-group substances and their precursor antigens. Hassid reviews the role of sugar phosphates in the biosynthesis of complex saccharides. Pazur and co-workers present information obtained by isotopic techniques on the nature of enzyme-substrate complexes in the hydrolysis of polysaccharides. Gabriel presents a common mechanism for the production of 6-deoxyhexoses. An intermediate nucleoside-5 -(6-deoxyhexose-4-ulose pyrophosphate) is formed in each of the syntheses. 

With the heterogeneous hydrolysis of polysaccharides like cellulose, these general considerations are valid, too, of course, but the rate of cleavage is slowed down by one or two orders of magnitude by the limited accessibility of the acetalic O atoms. The rate of reaction depends largely on the physical structure of the original samples and on the state... 

Hydrolysis of Polysaccharides with Trifluoroacetic Acid and its Application to Rapid Wood and Pulp Analysis... 

For the total hydrolysis of polysaccharides, trifluoroacetic acid (TFA) has important advantages over sulfuric acid. The reaction time is short and there is no need for conventional neutralization, as TFA is volatile and can be removed by evaporation. Several methods have been developed, depending on the substance to be hydrolyzed. Soluble saccharides (e.g., polyoses) can be hydrolyzed with diluted TFA, while cellulose, pulp, and wood need treatments with concentrated TFA in homogeneous solution. The presence of lignin impedes the hydrolysis of polysaccharides thus, especially for wood samples, an intensive treatment with TFA is necessary, and correction values have to be considered. Several application examples show that the hydrolysis with TFA enables a rapid quantitative determination of the composition of polysaccharides, pulps, and woods. 

We applied TFA to hydrolysis of polysaccharides under various conditions (7,8,9). Many tests have shown that some troubles known from sulfuric acid treatment can be eliminated using TFA. In addition, the tests resulted in a wide applicability and flexibility of the method, and it can be easily adapted to many polysaccharides. 

TFA has been used for about three years in our laboratory, and it has become our exclusive reagent for the total hydrolysis of polysaccharides. Moreover, dilute TFA is applied to partial hydrolysis of lignin-polysaccharide complexes (13). 

---