Advanced Materials Research Online: 2014-06-06 ISSN: 1662-8985, Vols. 941-944, pp 1003-1007 doi:10.4028/www.scientific.net/amr.941-944.1003 2014 Trans Tech Publications, Switzerland Extraction technology and determination of total flavonoid in Magnolia liliiflora petals Yihua Jiang 1, a, *, Xinlong Jiang 2,b and Chenggang Cai 1,c 1 Zhejiang Shuren University, Hangzhou, China 2 Institute of Food and Fermentation Engineering,Zhejiang Shuren University, Hangzhou, China a email:hzjyih828@163.com, b email:jxinl2003@163.com, c email:ccg0516@sina.com Keywords:Magnolia liliflora;total Flavonoid; Microwave-assisted Extraction; Orthogonal experiment. Abstract: Orthogonal experiment optimization of microwave-assisted extraction (MAE) conditions of total flavonoids from Magnolia liliflora petals with the factors of ethanol concentration, extraction time,microwave power,solid/liquid ratio on the yield of total flavonoids were studied. The results showed that optimum extracting conditions of total flavonoids from Magnolia liliflora were as follows: adding 1:10 g ml -1 of 40% alcohol and extracting for 5 min,microwave irradiation power 464W, under the optimal conditions, the maximal yield of flavonoids was of 2.08%. Introduction Flavonoids is exist extensively in plants.the type and content of flavonoids in the medicinal materials of flower is rich particularly.flavonoid are known to be responsible for antioxidant[1],anti-inflammatory[2], anticancer[3], hepatoprotective potential[4] and antibacterial activity[5].it Attracted widespread attention at home and abroad. Magnolia liliflora Desr is a plant of the magnolia genus magnoliaceae, with flower medicine called magnolia, used as analgesic agent, can treat headache, lumbago, rhinitis and other diseases clinically [6]. The flower also contains natural essence, industry refining can be used for the production of cosmetics [7]. Some scholars have found rich in flavonoids in buds of Magnolia liliflora [8], but not found the extraction technology and determination of total flavonoids in petals of Magnolia liliflora. In this paper,using ethanol as solvent, the microwave-assisted extraction (MAE) parameters such as extractant,microwave irradiation power, the ratio of solvents to materials, and extraction time were optimized by orthogonal experiment from Magnolia liliflora petals,with the yield of flavonoids() as the index. Determination of the content of total flavonoids was made on the basis of Magnolia petals.then the results would be the theoretical basis for the comprehensive utilization of the material. Material and Methods Material Fresh Magnolia liliflora petals were collected from our campus in November, 2013.It dried at 40 C to constant weight and were crushed using a universal grinder and sieved (60 80 mesh), and the resulting particles were stored in a desiccator until use. Rutin standard sample was purchased from chinese medicine group shanghai chemical reagent company.all other reagents were of analytical grade. All rights reserved. No part of contents of this paper may be reproduced or transmitted in any form or by any means without the written permission of Trans Tech Publications, www.ttp.net. (ID: 130.203.136.75, Pennsylvania State University, University Park, USA-13/05/16,12:28:59)
1004 Materials and Processes Technologies V Experimental Methods Extraction process Accurate weighing dry Magnolia liliflora petals powder adding a certain amount of ethanol solution microwave treatment filtering collected the supernatant constant volume,extracting liquid was obtained measuring. Product analysis Content of flavonoids in the extract was determined by spectrophotometric method, using rutin as standard sample. According to the reported method in the article[9],the regression equation C (mg)=0.6503a+0.01032 between concentration of flavonoids and absorbance was established,the linear range was 0 ~ 1.0mg,the correlation coefficient r was 0.9997. Orthogonal experiment optimization of extraction process Microwave extraction experiment optimization including ethanol concentration, solid-liquid ratio,microwave power,extraction times choice, for which several factors are parallel single factor experiment, study their influence on extraction.single factors near the optimal value should be firstly found before the orthogonal experiment so as to found the most approximately real situation. Dry samples of 0.5000g were weighed into 250 ml cone flask with plug,respectively, and added amount of extractant,microwave extraction under certain conditions, the filtrate volume was to 50mL with the corresponding extraction agent.for different ethanol concentration (20,30,40,50, 60, 70%), extraction time (1,2,3,4,5min), microwave power (144, 288, 464, 648,800W), the ratio of material to liquid (1:10,1:20,1:30,1:40,1:50 g ml -1 )and other conditions to do parallel single factor experiments.the single factor test results were shown in Fig 1.According to Fig 1,we designed the L 9 (3 4 ) program of orthogonal experiment,the factors and levels were listed in Table 1.Results were disposed by the intuitive analysis and variance analysis, significant test of flavonoids extraction factor by Duncan [10]. 20 30 40 50 60 70 Ethanol concentration/% 1 2 3 4 5 Extraction time/min 10 20 30 40 50 100 300 500 700 900 Ratio of liquid to solid/(ml g -1 ) Microwave power/w Fig.1 Effects of different factors on MAE
Advanced Materials Research Vols. 941-944 1005 Table 1 Levels and factors of orthogonal experiment design C The ratio of A Ethanol B Extraction time D Microwave Level liquid to solid concentration (%) (min) (ml g -1 power (W) ) 1 40 3 10 288 2 50 4 20 464 3 60 5 30 648 Determination of total flavonoid and recovery rate experiment Accordance with the optimum extraction conditions, were determined for 3 times, calculating the average content of flavonoids.at the same time,the recovery rate experiment were conducted by adding standard solution. Result and analysis The test results of orthogonal experiment Based on the results of single factors experiments, the extraction experiments were carried out by orthogonal experiment design according to Table 2. The intuitive analysis From Table 2 can be seen, the effect of the microwave power on the rate of extraction was the maximum.the range of data R was visible, the effect order of various factors on the extraction: microwave power > extraction time >ethanol concentration > the ratio of liquid to solid, the optimum extraction conditions were: A 1 B 3 C 1 D 2, namely ethanol concentration was 40%, extraction time was 5 min, ratio of solid to liquid was 1:10 g ml -1, microwave power was 464W. The results of orthogonal experiment about extraction time,ethanol concentration and material liquid ratio on the flavonoids had differences to the single factor experiment, which may be the interaction between various factors effect. Table 2 Orthogonal experimental program and results of extraction ( n = 3) the yield of No. A B C D flavonoids () 1 1 1 1 1 1.476 2 1 2 2 2 2.037 3 1 3 3 3 2.011 4 2 1 2 3 1.662 5 2 2 3 1 1.508 6 2 3 1 2 2.078 7 3 1 3 2 1.753 8 3 2 1 3 1.837 9 3 3 2 1 1.629 K 1 1.841 1.630 1.797 1.538 K 2 1.749 1.794 1.776 1.956 K 3 1.740 6 1.757 1.837 R 0.102 0.276 0.040 0.418
1006 Materials and Processes Technologies V The analysis of variance We can see from Table 3, microwave power was extremely significant influencing factor and the extraction time was significant influencing factor.the other factor is not significant factor.it could consider to determine, as determined by the Duncan method after checking back. Table 3 Analysis of variance source of variation SS df MS F significance A 0.0189 2 0.0094 7.9960 B 0.1153 2 0.0577 48.8059 * C 0.0024 2 0.0012 00 D 0.2786 2 0.1393 117.9252 ** SS T 0.4152 8 Note: F0.05(2, 2)=19.00,F0.01(2, 2)=99.00; * and ** significant at P=0.05 and P=0.01 respectively. The Duncan analysis[10] First were found between the level of various factors r 0.01, r 0.05 (r 0.01 =14, r 0.05 =6.09), then calculate the value of R k (standard error is 0.018, the R k = standard error *r ), then according to the calculation results,it was compared for each factor level significantly.from Table 4 can be seen, in addition to ethanol concentration and the ratio of liquid to solid,other factors have significant differences.the extraction time of 5 min is significantly to the other and microwave power of 464W is significantly to the other too. Considering the use of energy and resources, comprehensive analysis to determine the optimal extraction conditions of total flavonoids of Magnolia liliflora was A 1 B 3 C 1 D 2. It was the same to visual analysis and variance analysis. Tables 4 Compare significant difference in four levels of factors 1(A) 2(B) 3(C) 4(D) Level Y P 0.05 P 0.01 Level Y P 0.05 P 0.01 Level Y P 0.05 P 0.01 Level Y P 0.05 P 0.01 1 1.841 a A 3 6 a A 1 1.797 a A 2 1.956 a A 2 1.749 a A 2 1.794 b AB 2 1.776 b A 3 1.837 b A 3 1.740 a A 1 1.630 c B 3 1.757 b A 1 1.538 c B Determination of total flavonoid Under the optimum extraction conditions, repeated for 3 times,the measured flavonoids were: 2.082%, 2.076%, 2.081%, the average content of flavone was 2.080%, RSD=0.80% (n=3). It was the same basically to variance analysis. The Recovery rate experiment It were added to 0.2mg/ml rutin solution 1, 2, 3, 4, 5.00ml in the extract of known flavonoids respectively, determination of the content of flavonoids, calculated the recovery rate. The results in Table 5. The average recovery of measured castor leaf total flavonoids was 100.7%, RSD=1.22% (n=5). Table 5 Recovery rate experiment No. Samples containing rutin/µg measured value/µg rate of recovery/% flavonoids/µg 1 208.0 200 398.4 0.992 2 208.0 400 612.8 1.012 3 208.0 600 812.8 8 4 208.0 800 1020.0 1.015 5 208.0 1000 1215.0 7 average 100.7
Advanced Materials Research Vols. 941-944 1007 Conclusion The results of orthogonal experiment showed that ratio of liquid to solid was significant factors influencing on extraction total flavonoids from Magnolia liliflora. The optimal processing conditions were as follows: ethanol concentration 40%,the ratio of solvents to materials 10 ml g -1, microwave irradiation power 464W, and extraction 5 min, under the optimal conditions, the maximal yield of flavonoids was of 2.08%. Microwave extraction of total flavonoids from Magnolia liliflora is fast, energy saving and high extraction rate. Acknowledgments This work was supported by Elite compound build innovative carrier project(no.2012e80002), The key discipline of Zhejiang province in Colleges and Universities, China (No.YH2013S08). References [1] K.S.Gould, C.Lister, Flavonoid functions in plants. In Flavonoids: Chemistry, Biochemistry and Applications; Andersen, O.M., Markham, K.R., Eds.; CRC Press:Boca Raton, FL, USA, 2006; pp. 397 441. [2] M.F.Yam,V.Lim,I.M.Salman,et a1.hplc and anti-inflammatory studies of the flavonoid rich chloroform extract fraction of Orthosiphon stamineus leaves. Molecules2010, 15, 4452 4466. [3] T.R.Cibin, D.G.Devi, A.Abraham, Chemoprevention of skin cancer by the flavonoid fraction of Saraca asoka. Phytother. Res. 2010, 24, 666 672. [4] A.Jain, A.K.Singhai,V.K.Dixit, A comparative study of ethanol extract of leaves of Tephrosia purpurea pers and the flavonoid isolated for hepatoprotective activity. Indian J. Pharm. Sci. 2006, 68, 740 743. [5] I.Naeem,Z.Saddiqe, A.Patel, et a1. Analysis of flavonoid and antimicrobial activity of extracts of Hypericum perforatum. Asian J. Chem. 2010, 22, 3596 3600. [6] X.M.Wang,J.Weng,Z.S.Zhong.Study on the Extraction and Stability of Pigment from Magnolia liliflora Desr.[J].Northern Horticulture, 2012, (08):43-45. (in Chinese) [7] Q.Wang,H.L.ling,Determination of Total Flavonoids in Bud of Magnolia and Studies on Extraction Process[J].Research and Practice of Chinese Medicines, 2003, 17(02):32-33. (in Chinese) [8] W.Z.Wu,GC-MS Analysis of Chemical Components in Essential Oil from Flos Magnolia[J].Chinese herbal medicine, 2012, 23(9):538-541. (in Chinese) [9] L.J.Zhang, W.H.Zhang, H.J.Zhang,Determination of the total content of flavones in Qianshan Semen Cassiacs. Food Sci,2003, 24:101-103. (in Chinese) [10] R.Qian,Biostatistics [M]. Beijing: Higher Education Press, 1999.126-129. (in Chinese)
Materials and Processes Technologies V 10.4028/www.scientific.net/AMR.941-944 Extraction Technology and Determination of Total Flavonoid in Magnolia Liliiflora Petals 10.4028/www.scientific.net/AMR.941-944.1003