Study on the Polysaccharide Extracted from Bergamot (Citrus medica L. var. sarcodactylis) Residue by Two Methods and Its Immunomodulatory Function in RAW264.7 Cells

In this paper, bergamot residue after extracting essential oils and flavonoids was used as raw material to extract polysaccharide by CPE (Continuous phase-transition extraction) and HWE (Hot water extraction) methods, respectively. Based on the single factor experiments, the response surface methodology was employed to obtain the optimal extraction conditions of polysaccharide using extraction rate as the index and the RAW264.7 macrophage was used to evaluate the immunological activity of the extracted polysaccharide. Therein, MTT assay, neutral red phagocytic assay and Griess method were used to detect the effects of the bergamot polysaccharide on the growth, phagocytic activity and NO release of RAW264.7, respectively. The results showed that the optimum extraction conditions of CPE method were as follows:The extraction temperature of 99℃, the extraction time of 4.5 h, the extraction flow rate of 66 L/h. The corresponding extraction rate and yield were 16.09%±0.12% and 21.31%±0.13%, respectively. For HWE method, the optimal conditions were:The extraction temperature of 95℃, the extraction time of 6.0 h, the liquid material ratio of 63 mL/g. The resulted extraction rate and yield were 15.43%±0.21% and 20.22%±0.16%, respectively. Compared with HWE method, CPE method increased the extraction rate by 4.28% and the yield by 5.39% (P<0.05) of bergamot polysaccharide and shorten the extraction time by 1/4. Bergamot polysaccharide had no significant toxic effect on RAW264.7 macrophages concentrations under 500 μg/mL, and could promote the secretion of NO and enhance the phagocytic activity. To summarize, the CPE method is suitable for the extraction of bergamot polysaccharide, and is superior to the traditional HWE method. Furthermore, the bergamot polysaccharide has the potential to develop immune health foods and medicines.