Extraction Of Yellow Pigment From Forsythia Flower And Its Effects On Stress Resistance Of Caenorhabditis Elegans

Food coloring is a food additives that colors foods or improves the color of foods,including artificial pigments and natural pigments.Although the chemical synthesis pigment is simple in production process and low in price,with the increase in the amount of synthetic pigment used,safety issues have gradually attracted people's attention.Natural pigments are non-toxic and harmless to the human body,and some have certain biological activities.Therefore,the development of non-toxic and harmless natural pigments has become the current market demand.Forsythia suspensa(Thunb.)Vahl is a widely used traditional Chinese medicine.It blooms in early spring.There is a long flowering period and a large amount of flowers,which a bright yellow and long-lasting color.It is a source of natural pigments with great development potential.In this paper,forsythia flower yellow pigment(FFYP)was extracted from forsythia flower to study its antioxidant activity in vitro and in vivo,and the effect of FFYP on the stress resistance of Caenorhabditis elegans.In order to develop a biologically active natural food coloring.The main contents of this article are as follow:1.Preparation and composition identification of FFYP.The optimal extraction method of FFYP was set up by single factor extraction experiment and orthogonal experiment.The results shows that the optimum extraction conditions were extracting reagent 95% ethanol,extraction temperature of 50 ?,and ratio of forsythia flower powder material to ethanol 1:100(g:m L).The main components and content of FFYP were analyzed by high performance liquid chromatography.The obtained FFYP mainly were rutin(236.1 mg/g),forsythiaside A(116.9 mg/g),rosin-?-D-glucopyranoside(82.5 mg/g),and forsythin(18.2 mg/g).2.The effect of p H,metal ion,temperature,light,oxidant and reducing on the stability of FFYP were studied.The results show that FFYP has good stability in weak acid and alkali environment;metal ions have little effect on its stability;FFYP is unstable when exposed to direct sunlight but stable under natural light or natural light,and its stability is not affected by ultraviolet light;FFYP has good stability at low temperstures,preservation rate were reduced when heated at high temperature,but the preservation rate can still be maintained above 80% after treatment at 80 °C for 1 hour;the reducing agent cannot reduce the preservation rate of FFYP,but the oxidant treatment will reduce the preservation rate of FFYP.3.Antioxidant activity of FFYP in cells and in vitro was examined.The extracellular antioxidant activity of FFYP was detected by chemical method.The results showed that FFYP has the 1,1-diphenyl-2-picrylhydrazyl(DPPH)free radical scavenging and iron ion reducing ability at a concentration of 0-0.1 mg/m L,and is concentration-dependently.The antioxidant activity of FFYP in Hep G2 cells was detected by Cellular Antioxidant Activity(CAA).The results also showed that FFYP has reactive oxygen species(ROS)scavenging ability in the cells in a dose-dependent manner.4.The effect of FFYP on the oxidative stress resistance of C.elegans was studied,and the mechanism was explored.The effects of FFYP on oxidative stress resistance of nematodes results showed that FFYP at 20,40,80 ?g/m L could increase the oxidative stress resistance of nematodes,and the effect of 40 ?g/m L was the most obvious.Detect the ROS content and antioxidant system activity of nematodes,the results showed that FFYP can significantly reduce the ROS content in nematodes,increase the activity of superoxide dismutase(SOD)and catalase(CAT),and increase the content of reduced glutathione(GSH).It indicates that FFYP enhances the resistance of nematodes to oxidative stress by increasing the activity of the antioxidant defense system in the nematode to scavenge free radicals.Using mutant nematode and q RT-PCR experiments to explore the mechanism of FFYP improve the oxidative stress resistance of nematode.The results showed that the effect of FFYP to prolongs the lifespan of wild type nematodes under oxidative stress was completely disappeared in the Skn-1 mutant nematode and attenuated in Daf-16 and Hsf-1 mutants.q RT-PCR experiments also showed that the expression levels of Daf-16,Skn-1 and Hsf-1 target genes were all increased.It is shown that the effect of FFYP on the oxidative stress resistance of nematode is through the interaction of Daf-16,Skn-1 and Hsf-1.In conclusion,the FFYP extracted in this study not only was stable but also had good antioxidant activity in vitro and in cells,also can enhance the oxidative stress resistance of C.elegans.This indicates that FFYP has ability to anti-oxidation and anti-stress,which lays a theoretical foundation for the future development and application of Forsythia.