Abstract
Abstract A polysaccharide, composed of 6-O-methyl-d-glucose and d-glucose in a molar ratio of 6:4, was isolated from the watersoluble portion of the deacylated crude lipids of Mycobacterium phlei and M. tuberculosis. The polysaccharide was purified by gel filtration, ion exchange chromatography, and borate complex formation, and was shown to have a molecular weight of about 3000 and [α]d22 +160° (c, 0.1; water). Methylation analysis of the polysaccharide showed that it has a branched structure with an average chain length of 8 to 9 hexose units. The major type of glycosidic linkage is α-(1 → 4), and the branching involves position 3 of one of the 6-O-methyl-d-glucose residues. Oligosaccharides were prepared by acetolysis of the polysaccharide, and were purified by paper chromatography. Di-, tri-, tetra-, penta-, hexa-, and heptasaccharides of α-(1 → 4)-linked 6-O-methyl-d-glucose, in addition to maltose and maltotriose, were isolated. The only heterooligosaccharide found was O-α-d-glucopyranosyl-(1 → 3)-6-O-methyl-d-glucose. β-Amylase had no action on the polysaccharide, whereas α-amylase liberated about 20% of the total carbohydrate, mainly as d-glucose and maltose. Limited amylolysis yielded d-glucose, maltose, and unidentified oligosaccharides containing d-glucose. Intact lipopolysaccharide, containing 2 to 3 moles of ester-linked fatty acid, was prepared from a 70% ethanol extract of M. phlei. The lipopolysaccharide was soluble in water and in a 2:1 chloroform-methanol mixture, but was only slightly soluble in methanol or ethanol. Proton magnetic resonance and infrared and ultraviolet spectra were consistent with the results obtained by chemical analysis.
Highlights
Oligosaccharides were prepared by acetolysis of the polysaccharide, and were purified by paper chromatography
Methyl-n-glucose polysaccharide, which we have isolated from the water-soluble fraction of the deacylated lipid mixture, was fractionated on the basis of molecular size (Figs. 2 and 3), acidity (Fig. 4), and borate complex formation (Fig. 5)
The final product appeared to be homogeneous by these three criteria. Both Al. phlei and M. tuberculosis produce the same type of compound, a slight difference in their characteristics of borate complex formation was observed (Fig. 5)
Summary
Sephadex A-25 (medium), all in irregular (not bead) form, were obtained from Pharmacia. DEAE-Sephadex was converted into its borate form as follows. Sephadex with HCl and NaOH, the resin was washed on a Buchner funnel with saturated sodium tetraborate until the effluent was free of chloride ion, washed with water to remove most of the residual borate, and packed into a column and washed free of borate. Swine pancreas cr-amylase and sweet potato fl-amylase were obtained from Worthington, and were used without further purification. The amylolytic activities of the enzymes were assayed by the dinitrosalicylate method [9]. Under the conditions described by Bernfeld, a unit of c~- or fl-amylase will produce per mm at 25” 1 pmole of reducing group, calculated as maltose.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
