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

•A pectic polysaccharide (CPE-II) was purified from the pectinase digests of citrus peels.•Administration of 10 μg/mouse of CPE-II showed a potent inhibitory effect on lung metastasis of Colon26-M3.1.•Rhamnogalacturonan II from citrus peels may have a beneficial inhibitory effect on tumor metastasis.

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.