Pullulans fractionation

Fig. 2 HPSEC profiles of three RG-II fractions purified from red wine (a), apple juice (b) and tomato juice (c) on Shodex OHpak KB columns. Elution times and Mw (kDa) of the pullulan standards are shown.

The molecular-weight distribution of a sample of a glucan, pul-lulan, isolated from cultures of the fungus Pullularia pullulans grown in sucrose solutions,"6 was determined by Granath and Kvist69 by the method developed by these workers for use with dextrans. Here, too, the column was calibrated with dextran fractions, as the shape of the pullulan molecule was believed to be sufficiently similar to that of a dextran to permit use of the same correlation of Kav with molecular weight for both polysaccharides. 

Oguchi and coworkers examined the decarboxylation behavior of -aminosalicylic acid (PAS) as a freeze-dried solid under thermal stress conditions (80°C) in the presence of the excipients pullulan (a linear polysaccharide which can not form inclusion complexes with PAS) and a-cyclodextrin (39). The solid-state stability was shown to correlate with the fraction of amorphous PAS. Increasing relative amounts of pullulan resulted in higher fractions of amorphous PAS. Rapid freezing (liquid nitrogen) was shown to result in a greater relative amount of amorphous drug, as expected. 

FIGURE 2 Degradation of pullulan (MW 200,000) in replicate anaerobic enrichment cultures of marine bacteria from anoxic sediments. Open circles show total pullulan concentrations remaining in the medium at each time point. The molecular weight distribution of the pullulan is shown by the stacked bars white >10,000 Da, stripes 5000 Da, black <1200 Da. Note that the lower molecular weight fraction progressively accumulated between 50 and 64 h. [Data from Arnosti et al. (1994).]... 

After fractionation with cetylpyridinium chloride, the acidic heteropolysaccharide of Aureobasidium (Pullularia) pullulans contains D-mannose, D-galactose, D-glucose, and D-glucuronic acid in the ratios of 43 35 10 12. It has a main chain of (1— 6)-linked a-D-mannopyrano-syl units substituted as in structure158 38. 

Having established that the fractionation was satisfactory for the individual pullulan and dextran samples in 0.2 M NaNOa, the lightscattering and viscosity results are viewed as a whole. The universal calibration plots for the pullulans and dextrans were superimposable. 

The validity of the power law, eq 2, was established in all cases. For atactic polystyrene (a-PS) and polyethylene oxide (PEO) a was found to be 1.5, and rather remarkably, irrespective of solvent quality, in the few instances when the solvent was varied. A value of 1.7 for a has just been reported (9) for the good solvent bromobenzene (not used by us) from a different source. For the bacterial polysaccharide pullulan, which happened to be obtainable in comparatively sharp fractions, a was 1.8 in what is believed to be good solvent (iO). 

An a-amylase found in the culture filtrate of a strain of Thermoactinomyces vulgaris was purified by ammonium sulphate fractionation, and ion-exchange chromatographies.The purified enzyme showed a single band on disc gel electrophoresis. The pH optimum, temperature optimum, and isoelectric point were pH 5.0, 70 C, and pi 5.2, respectively. The a-amylase was stabilized by Ca and was found to hydrolyse pullulan to panose. Therefore, the hydrolytic pattern of this enzyme is different from those of pullulanase and isopullulanase. 

Pullulan (Fig. 13.10) is one of the few neutral water-soluble microbial polysaccharides that can be obtained in large quantities by fermentation [50, 51]. Pullulan bios3uithesis is accomplished through mediation of sugar nucleotide-lipid carrier intermediates associated with the cell membrane fraction [52], It is an extracellular, unbranched homopolysaccharide consisting of maltotriose and maltotetraose units with both a-(l — 6) and a-(l — 4) linkages,... 

Duval, C. Le Cerf, D. Picton, L. Muller, G. Aggregation of amphiphilic pullulan derivatives evidenced by on-line flow field flow fractionation/multi-angle laser light scattering. J. Chromatogr. B, 2001, 753, 115 122. 

An a-amylase preparation from Thermoactinomyces vulgaris has been used to hydrolyse pullulan to panose. The digest was applied to a carbon-Celite column and eluted with a linear gradient of propan-l-ol from 0 to 5%. From the trisaccharide fractions eluted, panose was prepared in about 70% yield. The production of amylase activity by Thermomonospora curvata has been investigated. ... 

A strain of Bacillus cereus synthesizes a pullulanase and a -amylase, which together convert starch into maltose in high yield. The homogeneous pullulanase was obtained by fractional precipitation, adsorption onto starch and Celite, and gel filtration. The purified enzyme (pH optimum 6.0—6.5, mol. wt. 1.10 0.20 X 10 ) released maltose, maltotriose, and maltotetraose from -limit dextrin and maltotriose from pullulan, but it did not release amylose-like substances from amylopectin. The enzymic activity was inhibited by mercuric chloride and 4-chloromercuribenzoate, although the activity in the latter instance could be restored by cysteine. 

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