Extraction, Purification And Microencapsulation Of Perilla Anthocyanins

Perilla frutescens,belonging to the mint family,is an annual herb plant,which is one of medicinal and edible plants promulgated firstly by the Chinese Ministry of Health.Perilla anthocyanins are water-soluble natural colorants,which confer bright color and lustre,are safe,non-toxic,and possess significant nutritional and pharmaceutical values,have great potential in food,cosmetics and medicine industries.In this study,the perilla variety with the highest anthocyanins content and strongest antioxidant activity was selected and the the best picking period was specified by analyzing the accumulation of anthocyanins in the development of different perilla varieties.The main anthocyanins of perilla leaves were identified by high performance liquid chromatography-mass spectrometry(HPLC/MS).Meanwhile,the extraction and purification methods of anthocyanins were optimized,and the stability,antioxidant acitivty and antibacterial activity of perilla anthocyanins extracts were studied.In order to further improve the stability of perilla anthocyanins,perilla anthocyanins were microencapsulated.The main results are concluded as follows:(1)Ultrasound assisted extraction(UAE)was used to extract anthocyanins and phenolic compounds from perilla leaves.Response surface methodology(RSM)was used to optimize the extraction conditions.The optimal conditions were as follows:ethanol concentration of77%,liquid-to-solid ratio of 22:1,extraction temperature of 53°C and extraction time of 54min.Under these optimal conditions,the total anthocyanins content and total phenolic content were 6.44 mg cyanidin-3-glucoside equivalent(CGE)/g and 63.11 mg gallic acid equivalent(GAE)/g,respectively.(2)The development of perilla was studied by sampling at different stages,including seedling stage,bud stage,flowering stage,fructicative stage and mature stage.It is determined that the sampling period was the bud stage.Anthocyanin content,polyphenols content and antioxidant activity of different perilla varieties in bud stage were determined,which revealed the intrinsic relationship between anthocyanin,polyphenols and antioxidant activity by the Pearson’s correlation.Structures of 4 anthocyanins extracted from perilla were identified using HPLC/MS.Anthocyanins in perilla were mainly of the type of cyanidin.Among them malonylshisoinn was the main anthocyanin accounting for 50.17-66.69%of total,followed by shisoinn,accounting for 15.75-30.71%of total.These anthocyanins were acylated by different organic acids and fatty acids.(3)The conditions for purification of perilla anthocyanins by macroporous resin were studied by static and dynamic adsorption-desorption.The results showed that XDA-8 resin was the best resin for purification on this account of the maximum adsorption rate(78.61%)and desorption rate(89.52%).The dynamic adsorption and desorption parameters were loading of 1.17 mg/mL sample solution at a flow rate of 3 BV/h,and desorption with 60%ethanol solution at a flow rate of 2 BV/h.Under the best conditions,the anthocyanins content of variety 1 was 7.52%,which was increased by 8.26 times than crude extracts.Perilla anthocyanins of 11 other varieties were purified by the above purification method,and the anthocyanins,polyphenols and antioxidant activities of 12 kinds of purified samples were determined.The correlation between anthocyanins,polyphenols and antioxidant activity was further revealed.(4)Samples I-IV with different anthocyanins content were obtained by macroporous resin purification,ethyl acetate extraction and silica gel column purification.The anthocyanins content of samples(I-IV)was 0.91%,7.52%,18.23%and 24.36%,respectively.The antioxidant activity,stability and antibacterial of different samples were determined.The ability to remove DPPH·,ABTS~+,OH·free radicals and ferric reducing ability showed that the antioxidant activity of sample II was the strongest,while the antioxidant activity of sample I was the weakest,and the antioxidant activities of I-IV samples were weaker than Vc.The kinetic data analysis during heating suggests a first-order reaction for the degradation of perilla anthocyanin with the half-life of 29.87 h,17.50 h and 14.32 h under the heating condition of 70°C,80°C and 90°C for sample I.The half-life of sample from I to IV decreased successively,indicating that the higher the anthocyanins content,the worse the stability.The effect of light on the samples II-IV was similar,and the preservation rate was84.46%,84.06%and 83.31%after 7 d treatment,but the preservation rate of the sample I was only 51.27%.The antibacterial experiment showed that anthocynins smples had a certain effect on Escherichia coli and Staphylococcus aureus,but had little effect on Bacillus subtilis,and the antibacterial activity of samples IV was the strongest,while samples I was the weakest.(5)The optimal conditions of microencapsulation of anthocyanins were obtained by single factor and response surface experiment,and analyzed the characterization and stability of microcapsule powder.The best condition of the microencapsulation was maltodextrin 75%,core-to-wall material ratio 1:7.8 and the solids content 40%.Under these conditions,the encapsulation efficiency of perilla anthocyanins was 87.61%.The microcapsules sample showed rough and irregular shape under scanning electron microscope,which was a typical freeze-dried solid.The stability test showed that the stability of the microcapsule powder was higher than that of the anthocyanin sample under dark and natural light.The preservation rate of microcapsule powder and anthocyanins was 95.73%and 87.33%under dark condition,and88.58%and 79.67%under natural light condition at 25°C after 90 d.Fluctuation range of the total color value was smaller in dark than in natural light by color measurement.Antioxidant tests showed that anthocyanins retained antioxidant activity after being microencapsulated.