Structural elucidation of anti-metastatic rhamnogalacturonan II from the pectinase digest of citrus peels (Citrus unshiu)

The aim of this study was to characterize a polysaccharide found in citrus peels with an anti-metastatic property. CPE-II was purified by the pectinase digestion of citrus peels. During in vivo lung metastasis of Colon26-M3.1, administration of 10 μg of CPE-II per mouse showed 81.3% inhibition of metastasis. CPE-II consists of 15 different monosaccharides and 22 different glycosyl linkages, characteristic of rhamnogalacturonan II (RG-II). The primary structure was elucidated based on sugar composition, methylation analysis, oligosaccharide analysis, and sequencing using GC, GC–MS, LC–MS, and ESI–MS/MS analyses. Sequential degradation using partial acid hydrolysis indicated that CPE-II contained Rhap-(1 → 5)-Kdo, Araf-(1 → 5)-Dha, an AceA-containing nonasaccharide, and an uronic acid-rich oligosaccharide in addition to an α-(1 → 4)-galacturono-oligosaccharide main chain. The molecular weight of CPE-II was observed to decrease from 9 to 5 kDa at a pH value of <2.0, as observed by HPSEC. Thus, we propose that the anti-metastatic CPE-II is primarily present as an RG-II dimer.