| Hazelnut is an important nut with high nutritional and economic value.Hazelnut is one of the main cash crops in the world.There are 25 species of Corylus in the world,and they are widely distributed and cultivated all over the world.In recent years,with the improvement of living standards,the demand for hazelnut products is increasing year by year,and the hazelnut processing by-products such as hazelnut shell,skin and green leaf cover are also increasing year by year,resulting in a waste of resources.In recent years,scientific research has found that polysaccharides are important functional components of these by-products,which have many biological activities such as antioxidation,hypolipidemia and so on.In this paper,the extraction technology,structural characteristics and anti-tumor activity of polysaccharides from hazelnut shell and hazelnut green leaf cover were studied.The main results are as follows:1)The polysaccharides of hazelnut shell and hazelnut green leaf cover were extracted by ultrasound-assisted hot water method.Single factor experiments were used to explore the effects of material-liquid ratio,ultrasonic time,ultrasonic power and ultrasonic temperature on the extraction rate of polysaccharides from hazelnut processing by-products.On the basis of single-factor experiments,taking the extraction rate of polysaccharides as the evaluation index,the Box-Behnken experiment was designed.Through the analysis of variance of the regression model,the order of the effects of each factor on the extraction rate of hazelnut shell polysaccharides and hazelnut green leaf cover polysaccharides was as follows: time > temperature > ratio of material to liquid > power;ratio of material to liquid > time > temperature > power.Through the response surface optimization experiment,the optimum extraction conditions of hazelnut chitosan were obtained as follows: material-to-liquid ratio 1:10 g/m L,ultrasonic time 52 min,ultrasonic power 327 W and ultrasonic temperature 69℃,the extraction rate of polysaccharide was 5.35 mg/g,and the optimum extraction conditions of hazelnut green leaf cover polysaccharides were as follows: material-to-liquid ratio 1:8 g/m L,ultrasonic time 39 min,ultrasonic power 423 W and ultrasonic temperature 72℃,and the extraction rate of polysaccharide was 35.78 mg/g.2)The polysaccharides of hazelnut shell and hazelnut green leaf cover obtained by ultrasonic extraction were decolorized by macroporous resin and deproteinized by Sevage to obtain polysaccharides HSPs(Hazelnut shell polysaccharides)and HGPs(Hazelnut green leaf cover polysaccharides).The two polysaccharides were detected by scanning electron microscope,gel permeation chromatography,high performance liquid chromatography,iodine-potassium iodide analysis,Congo red reaction,X-ray diffraction and thermal stability analysis.The results are as follows: under scanning electron microscope,HSPs is thin and curly,while HGPs is thicker flake,and many small pores can be seen at 500 x.According to the determination of molecular weight,it was found that the components of the two polysaccharides were relatively uniform,and that of HSPs was more uniform.Monosaccharide composition analysis showed that both polysaccharides were composed of11 monosaccharides.HSPs was mainly composed of D-arabinose and HGPs was mainly composed of D-galacturonic acid.The results of iodine-potassium iodide analysis and Congo red analysis showed that both HSPs and HGPs did not contain starch and contained longer side chains and more branches,and neither of them belonged to triple helix configuration polysaccharides.Thermal stability analysis showed that the thermal stability of HGPs was higher than that of HSPs.Through X-ray diffraction,it was found that the polysaccharides of the two by-products of hazelnut processing were semi-crystalline polymers,and the crystallinity of HSPs was better.3)The antitumor activities of the two polysaccharides were analyzed by CCK-8cytotoxicity test,scratch healing test,apoptosis detection by TUNEL method,cell cycle and intracellular reactive oxygen species level by flow cytometry.The results are as follows:cytotoxicity test showed that both HSPs and HGPs could inhibit the proliferation of human cervical cancer cell He La and human colon cancer cell SW480.Among them,HGPs had better anti-tumor effect.The cell scratch healing experiment showed that the cell scratch healing rate decreased with the increase of polysaccharide dose,and HGPs could inhibit cell migration more than HSPs.The experimental results of apoptosis were consistent with those of CCK-8,which showed that HGPs had better anti-tumor effect.Cell cycle analysis showed that He La cells treated with HSPs blocked the cell cycle at G2 M phase and HGPs blocked the growth cycle at intermitotic phase.When SW480 cells were treated with HSPs,the cell cycle was blocked in G0/G1 phase,and the growth cycle was blocked in S phase by HGPs.The changes of intracellular reactive oxygen species content showed that SW480 cells treated with hazelnut shell and hazelnut green leaf cover polysaccharides could promote the accumulation of ROS in SW480 cells,resulting in apoptosis.To sum up,the optimum technological parameters of ultrasonic-assisted hot water extraction of polysaccharides from hazelnut shell and hazelnut green leaf cover were determined in this paper.The two polysaccharides belong to homogeneous and stable polysaccharides and have good anti-tumor activity.This study provides a theoretical basis for the research,development and application of hazelnut shell polysaccharides and hazelnut green leaf cover polysaccharides. |
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