A mild, reproducible procedure has been developed for the isolation of D-apiose-containing polysaccharides from the cell wall of Lemna minor. The procedure is based on the extraction of the cell walls with 0.5% ammonium oxalate. conditions used at 22° have no known degradative effect on polysaccharides. On a dry weight basis, the polysaccharides extracted with ammonium oxalate made up 14% of the material designated cell walls and contained 20% of the D-apiose orig- inally present in the cell walls. The cell walls, as isolated, contained 83% of the D-apiose present in L. minor. After extraction with ammonium oxalate, purified polysaccharides were obtained by DEAE-Sephadex column chromatography and by fractional precipitation with sodium chloride. With these procedures, the material extracted at 22° could be separated into at least five polysaccharides. On a dry weight basis, two of these polysaccharides made up more than 50% of the material extracted at 22o. There was a direct relationship between the D-apiose content of the polysaccharides and their solubility in sodium chloride solutions;​ those of highest D-apiose content were most soluble. All of the polysaccharides David A. Hart isolated appeared to be of one general type, namely galac- turonans to which were attached sidechains containing D-apiose. The D-apiose content of the apiogalacturonans varied from 7.9 to 38.1%. The content of esterified D-galacturonic acid resi- dues in all apiogalacturonans was low, being in the range 1.0-3.5%. Hydrolysis of a representative apiogalacturonan with dilute acid resulted in the complete removal of the D-apiose with little or no degradation of the galacturonan portion. Treatment of polysaccharide fractions with pectinase established that those of high D-apiose content and soluble in 1.0 M-sodium chloride were not degraded whereas those of low D-apiose content and insoluble in 1.0 M-sodium chloride were extensively degraded. When the D-apiose was removed from a typical pectinase-resistent polysaccharide, the remainder of the polysaccharide was readily degraded by this enzyme. Periodate oxidation of representative polysaccharide fractions and apiogalacturonans and determination of formaldehyde released, showed that about 50% of the D-apiose molecules were substituted at either the 3- or the 3'-position.

The apiogalacturonans, as the sodium salts, were par- tially degraded when heated under mildly acidic conditions. The extent of the hydrolysis under these conditions was approximately equal to the percent D-apiose of the aplogalac- turonans. The same two degradation products were obtained from all of the apiogalacturonans. These were D-apiose and a disaccharide of D-apiose named apibiose. The residues from the degradation, the galacturonans, were not characterized. Periodate oxidation of apibiose and crystalline apibiose phenylosotriazole and determination of formaldehyde released, showed that the position of the linkage between the two D-apiose molecules was 1→3'. Proton magnetic resonance spec- trometry and molecular rotational data suggested that the linkage had the β configuration. Methylation analysis of apiblose and apibiose phenylosotriazole indicated that the non-reducing terminal D-apiose molecule had the D-apio-D- furanose configuration. The configuration at C-3 of the reducing terminal D-apiose molecule was not determined. Therefore the disaccharide is O-β-D-apio-D-furanosyl-(1-3') - D-apiose. When attached as sidechains to the a-(1—4) - galacturonan, the disaccharide is (2 and/or 3)-0-[0-8-D- apio-D-furanosyl-(1—3')-(α or β)-D-apio-(D or L)-furanosyl]- galacturonan.