Study On Extraction And Biological Activity Of Polysaccharide From Benincasa Hispida Peels

Benincasa hispida(Thunb.)Cogn.is a kind of herbaceous plant in gourd family.It is widely used in food processing because of its high yield,low cost and rich nutrition.The outer peel of wax gourd(BP),has the effect of quenching thirst,diuresis,diarrhea and so on.BP is rich in polysaccharides and has good biological activities such as antioxidant,which is mostly discarded at present,resulting in resource waste and environmental pollution.In this paper,the polysaccharides(BPs)from wax gourd peel by using different extraction process were extracted and their physicochemical characteristics and biological activities were clarified,which provided the basis for its development in the field of medicine and health care.The main results are as follows:On the basis of single factor experiment,the technological parameters of four extraction methods including thermal reflux extraction(HRE),ultra-high pressure extraction(UPE),microwave assisted enzymatic extraction(MEE)and ultrasound assisted enzymatic extraction(UEE)were optimized,and the optimal technological conditions of HRE were obtained as follows:reflux temperature of 90?,extraction time of 120 min and liquid-solid ratio of 50 m L/g.Under these conditions,the extraction yield of polysaccharide(HBPs)by HRE was 3.50%;the optimum process conditions of UPE:pressure of 400 MPa,temperature 70?,extraction time 4 min,the extraction yield of crude polysaccharides(UPBPs)was 1.27%under these conditions.The optimal extraction conditions of MEE were as follows:liquid-solid ratio of 61.70m L/g,microwave power of 523.06 W and extraction temperature of 70.40?;under these conditions,the extraction yield of crude polysaccharides(MEBPs)was 11.38%.The optimal condition of UEE were as follows:extraction temperature of 61.70?,ultrasonic power of 415.38 W and extraction time of 32.14 min;under these conditions,the extraction yield of crude polysaccharides(UEBPs)was 14.80%.The results of process optimization showed that MEE and UEE were the best extraction processes for BPs.The MEBPs and UEBPs were separated by DEAE Sepharose CL-6B and purified by Sephadex G-150 to obtain the homogenized polysaccharide components UEBP-1,UEBP-2,MEBP-1 and MEBP-2.The physicochemical characteristics of the above crude polysaccharide and homogeneous polysaccharide were as follows:total sugar content 81.32%-89.60%,protein content 0.29%-13.24%,uronic acid content 3.70%-22.12%sulfate content0.05%-1.07%,total polyphenol content 1.25%-2.19%,and the total flavonoid content3.44%-3.78%.The purity and molecular weight(Mw)of purified polysaccharides MEBP-1,MEBP-2,UEBP-1 and UEBP-2 were determined by high performance gel chromatography(HPGPC),and they were:10050,90698,161175 and 103780 Da,of which UEBP-2 and MEBP-2 all have symmetrical narrow peaks,which proved to be uniform polysaccharides with high purity.The purified polysaccharides were subjected to acid hydrolysis,acetylation derivatization,and gas chromatography-mass spectrometry(GC-MS)method to determine the monosaccharide composition.MEBP-1 was composed of rhamnose(Rha),arabinose(Ara),xylose(Xyl)and Glucose(Glc)in a molar ratio of 0.62:0.50:1.03:0.5;MEBP-2 composed of Rha,Ara,Xyl and Glc in a molar ratio:0.47:1.20:1.04:0.18;UEBP-1 composed of Rha,galactose(Gal)and Glc in a molar ratio of 1.56:1.20:0.40;UEBP-2 was composed of Ara and Xyl in a molar ratio of 1.17:1.32.The water solubility test showed that these polysaccharides all had better water solubility.Infrared spectroscopy(IR)showed that the four homogeneous polysaccharides contained typical polysaccharide characteristic absorption peaks at 3400 and 2900 cm^-1;the COOH stretching vibration near 1616cm^-1 indicated that all four purified polysaccharides had carboxyl groups.The absorption peaks around 889 and 845 cm^-1 of the four polysaccharide components indicated the existence of?and?-glycosidic bonds;these results indicated that different processes did not change the main functional groups of polysaccharides.~1H NMR test showed that the four purified fractions all had?and?configuration glycosidic bonds,methoxyl substitution,MEBP-2 and UEBP-2 had acetyl substitution.The thermal stability test results showed that all samples showed good thermal stability,MEBP-2 and UEBP-2 had higher thermal stability,and the polysaccharides obtained by different extraction methods had different thermal stability.Circular dichroism showed that MEBP-1 and UEBP-2 had larger negative peaks,that is,stronger asymmetry.XRD analysis indicated that BPs existed in amorphous form,and the purified polysaccharides UEBP-1 and MEBP-1 had partially stable crystal structures.Congo red experiment showed that UEBP-1,MEBP-1,UEBP-2 have triple helix structure,MEBP-2 does not have triple helix structure.These results indicate that the extraction process not only significantly affects the extraction yield,but also has a significant impact on the physicochemical properties of polysaccharides.The antioxidant activity in vitro was evaluated by DPPH·,ABTS⁽¹⁰⁾·,OH·scavenging text and reducing ability text.The results showed that good scavenging ability to other radicals except OH·,and showed a significant dose-effect dependence.Moreover,the antioxidant activity of crude polysaccharide under different methods is stronger than purified polysaccharide,and the scavenging activity of UEBP-2 extracted by UEE is the best.The results of?-mylase inhibition test showed that the polysaccharide components by UEE was better than that of MEE.The bacteriostatic ability of polysaccharides was evaluated by Escherichia coli,Staphylococcus aureus and Candida albicans.The results showed that MEBP-1 and MEBP-2 had certain inhibitory ability against Escherichia coli.Considering the extraction yield and activity under different processes,UEE method is the most suitable extraction method for BPs.