Mikroanalisis Kandungan Senyawa Fenolik Total Ekstrak Biji Kedelai dengan Reagen Folin-Ciocalteu

Eriyanto Yusnawan, Joko S Utomo


One of the roles of phenolic compounds is as chemical defense system to inhibit the growth of
pathogen infection.Folin-Ciocalteu (FC) reagent is used to estimate total phenolic compounds
spectrophotometrically. Microanalysis of the phenolic compounds using FC reagent is still limited.
This study, therefore, aimed to develop a rapid method to estimate total phenolic compounds in
soybean seed extract in a 96-well microplate. Optimum kinetic reaction was achieved after 90
minutes of incubation. Response of 25 – 1.000 µg ml-1 of gallic acid was linear (r = 0.99). Hundreds
of samples could be analysed using the microplate 2 format in limited time, such as when working
with soybean germplasm collection.


Folin-Ciocalteu reagent, microanalysis, phenolic compound, soybean seed extract, 96-well microplate

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Ainsworth, E.A. and K.M. Gillespie. 2007. Estimation of total phenolic content and other oxidation substrates in plant tissues using Folin–Ciocalteu reagent. Nat. protoc. 2(4):875-877. DOI: 10.1038/nprot.2007.102.

Amarowicz, R. and R.B. Pegg. 2008. Legumes as a source of natural antioxidants. J. Lipid Sci. 110(10):865-878. DOI: 10.1002/ejlt.200800114.

Arfaoui, A., A. El Hadrami, Y. Mabrouk, B. Sifi, A. Boudabous, I. El Hadrami, F. Daayf, and M. Cherif. 2007. Treatment of chickpea with Rhizobium isolates enhances the expression of phenylpropanoid defense-related genes in response to infection by Fusarium oxysporum f. sp. ciceris. Plant Physiol. Biochem. 45(6-7): 470-479. DOI: 10.1016/j.plaphy.2007.04.004.

Attard, E. 2013. A rapid microtitre plate Folin-Ciocalteu method for the assessment of polyphenols. Cent. Eur. J. Biol. 8(1):48-53. DOI: 10.2478/s11535-012-0107-3.

Berker, K.I., F.A. Ozdemir Olgun, D. Ozyurt, B. Demirata, and R. Apak. 2013. Modified Folin–Ciocalteu antioxidant capacity assay for measuring lipophilic antioxidants. J. Agric. Food Chem. 61(20):4783-4791. DOI: 10.1021/jf400249k.

Blainski, A., G.C. Lopes, and J.C.P. De Mello. 2013. Application and analysis of the Folin-Ciocalteu method for the determination of the total phenolic content from Limonium brasiliense L. Molecules. 18(6):6852-6865. DOI: 10.3390/molecules18066852.

Boue, S.M., T.E. Cleveland, C. Carter-Wientjes, B.Y. Shih, D. Bhatnagar, J.M. McLachlan, and M.E. Burow. 2009. Phytoalexin-enriched functional foods. J. Agric. Food Chem. 57(7):2614-2622. DOI: 10.1021/jf8040403.

Cicco, N., M.T. Lanorte, M. Paraggio, M. Viggiano, and V. Lattanzio. 2009. A reproducible, rapid and inexpensive Folin–Ciocalteu micro-method in determining phenolics of plant methanol extracts. Microchem. J. 91(1):107-110. DOI: 10.1016/j.microc.2008.08.011.

Couto, C., L.R. Silva, P. Valentao, E. Velazquez, A. Peix, and P.B. Andrade. 2011. Effects induced by the nodulation with Bradyrhizobium japonicum on Glycine max (soybean) metabolism and antioxidant potential. Food Chem. 127(4): 1487-1495. DOI: 10.1016/j.foodchem.2011.01.135.

Dardanelli, M.S., F.J.F. De Cordoba, J. Estevez, R. Contreras, M.T. Cubo, M.A. Rodriguez-Carvajal, A.M. Gil-Serrano, F.J. Lopez-Baena, R. Bellogin, H. Manyani, F.J. Ollero, and M. Megias. 2012. Changes in flavonoids secreted by Phaseolus vulgaris roots in the presence of salt and the plant growth-promoting rhizobacterium Chryseobacterium balustinum. Appl. Soil Ecol. 57:31-38. DOI: 10.1016/j.apsoil.2012.01.005.

Du, G., H.Y. Zhao, Q.W. Zhang, G.H. Li, F.Q. Yang, Y. Wang, Y.C. Li, and Y.T. Wang. 2010. A rapid method for simultaneous determination of 14 phenolic compounds in Radix Puerariae using microwave-assisted extraction and ultra high performance liquid chromatography coupled with diode array detection and time-of-flight mass spectrometry. J. Chromatogr. A. 1217(5): 705-714. DOI: 10.1016/j.chroma.2009.12.017.

Everette, J.D., Q.M. Bryant, A.M. Green, Y.A. Abbey, G.W. Wangila, and R.B. Walker. 2010. Thorough study of reactivity of various compound classes toward the Folin− Ciocalteu reagent. J. Agric. Food Chem. 58(14):8139-8144. DOI: 10.1021/jf1005935.

George, S., P. Brat, P. Alter, and M.J. Amiot. 2005. Rapid determination of polyphenols and vitamin C in plant-derived products. J. Agric. Food Chem. 53(5):1370-1373. DOI: 10.1021/jf048396b.

Huang, D., B. Ou, and R.L. Prior. 2005. The chemistry behind antioxidant capacity assays. J. Agric. Food Chem. 53(6):1841-1856. DOI: 10.1021/jf030723c.

Jun-Ming, S., S. Bao-Li, H. Fen-Xia, Y. Shu-Rong, Y. Hua, and A. Kikuchi. 2011. Rapid HPLC method for determination of 12 isoflavone components in soybean seeds. Agric. Sci. China. 10(1): 70-77. DOI: 10.1016/S1671-2927(11)60308-8.

Liu, P., B. Yang, and H. Kallio. 2010. Characterization of phenolic compounds in Chinese hawthorn (Crataegus pinnatifida Bge. var. major) fruit by high performance liquid chromatography–electrospray ionization mass spectrometry. Food Chem. 121(4): 1188-1197. 10.1016/j.foodchem.2010.02.002

Lygin, A.V., S. Li, R. Vittal, J.M. Widholm, G.L. Hartman, and V.V. Lozovaya. 2009. The importance of phenolic metabolism to limit the growth of Phakopsora pachyrhizi. Phytopathol. 99: 1412-1420. DOI: 10.1094/PHYTO-99-12-1412.

Ma, Y. and P.C. Cheung. 2007. Spectrophotometric Determination of Phenolic Compounds by Enzymatic and Chemical Methods A Comparison of Structure-Activity Relationship. J. Agric. Food. Chem. 55(10): 4222-4228. DOI: 10.1021/jf070084w.

Magalhaes, L.M., F. Santos, M.A. Segundo, S. Reis, and J.L. Lima. 2010. Rapid microplate high-throughput methodology for assessment of Folin-Ciocalteu reducing capacity. Talanta. 83(2):441-447. DOI:10.1016/j.talanta.2010.09.042

Makoi, J.H. and P.A. Ndakidemi. 2007. Biological, ecological and agronomic significance of plant phenolic compounds in rhizosphere of the symbiotic legumes. Afr. J. Biotechnol. 6(12):1358-1368.

Malencic, D., M. Popovic, and J. Miladinovic. 2007. Phenolic content and antioxidant properties of soybean (Glycine max (L.) Merr.) seeds. Molecules. 12:576-581.

Mandal, S.M., D. Chakraborty, and S. Dey. 2010. Phenolic acids act as signaling molecules in plant-microbe symbioses. Plant. Signal Behav. 5(4):359-368. DOI: 10.4161/psb.5.4.10871.

Medina-Remon, A., A. Barrionuevo-Gonzalez, R. Zamora-Ros, C. Andres-Lacueva, R. Estruch, M.A. Martinez-Gonzalez, J. Diez-Espino, and R.M. Lamuela-Raventos. 2009. Rapid Folin–Ciocalteu method using microtiter 96-well plate cartridges for solid phase extraction to assess urinary total phenolic compounds, as a biomarker of total polyphenols intake. Anal. Chim. Acta. 634(1):54-60. DOI: 10.1016/j.aca.2008.12.012.

Mikulic-Petkovsek, M., A. Slatnar, R. Veberic, F. Stampar, and A. Solar. 2011. Phenolic response in green walnut husk after the infection with bacteria Xanthomonas arboricola pv. juglandis. Physiol. Mol. Plant Pathol. 76(3): 159-165. DOI: 10.1016/j.pmpp.2011.09.006.

Naczk, M. and F. Shahidi. 2004. Extraction and analysis of phenolics in food. J. Chromatogr. A. 1054(1):95-111. DOI:10.1016/j.chroma.2004.08.059.

Oh, S., J.A. Kim, H. Jeon, J.C. Park, Y.J. Koh, R. Hur, and J.S. 2008. Antifungal activity of eucalyptus-derived phenolics against postharvest pathogens of kiwifruits. Plant Pathol J. 24:322-327. DOI: 10.5423/PPJ.2008.24.3.322.

Prior, R.L., X. Wu, and K. Schaich. 2005. Standardized methods for the determination of antioxidant capacity and phenolics in foods and dietary supplements. J. Agric. Food Chem. 53(10):4290-4302. DOI: 10.1021/jf0502698.

Riedl, K.M., J.H. Lee, M. Renita, S.K. St Martin, S.J. Schwartz, and Y. Vodovotz. 2007. Isoflavone profiles, phenol content, and antioxidant activity of soybean seeds as influenced by cultivar and growing location in Ohio. J. Sci. Food Agric. 87:1197-1206. DOI: 10.1002/jsfa.2795.

Ruiz-Garcia, Y. and E. Gomez-Plaza. 2013. Elicitors: a tool for improving fruit phenolic content. Agriculture. 3(1): 33-52.

Sanchez-Rangel, J.C., J. Benavides, J.B. Heredia, L. Cisneros-Zevallos, and D.A. Jacobo-Velazquez. 2013. The Folin–Ciocalteu assay revisited: improvement of its specificity for total phenolic content determination. Anal. Methods. 5(21): 5990-5999. 10.1039/C3AY41125G.

Silva, L.R., M.J. Pereira, J. Azevedo, R. Mulas, E. Velazquez, F. Gonzalez-Andres, P. Valentao, and P.B. Andrade. 2013. Inoculation with Bradyrhizobium japonicum enhances the organic and fatty acids content of soybean (Glycine max (L.) Merrill) seeds. Food Chem. 141(4):3636-3648. DOI: 10.1016/j.foodchem.2013.06.045

Singleton, V. and J.A. Rossi. 1965. Colorimetry of total phenolics with phosphomolybdic-phosphotungstic acid reagents. Am. J. Enol. Vitic. 16(3):144-158.

Singleton, V.L., R. Orthofer, and R.M. Lamuela-raventos. 1999. Analysis of total phenols and other oxidation substrates and antioxidants by means of Folin-Ciocalteu reagent. In P. Lester (ed). Methods in Enzymology. Academic Press. p: 152-178.

Slavin, M., Z. Cheng, M. Luther, W. Kenworthy, and L.L. Yu. 2009. Antioxidant properties and phenolic, isoflavone, tocopherol and carotenoid composition of Maryland-grown soybean lines with altered fatty acid profiles. Food Chem. 114(1):20-27. DOI:10.1016/j.foodchem.2008.09.007.

Waterhouse, A.L. 2002. Determination of total phenolics. Current Protocols in Food Analytical Chemistry. John Willey and Sons. p: II.1.1-II.1.8.

Xu, B. and S.K.C. Chang. 2007. A Comparative study on phenolic profiles and antioxidant activities of legumes as affected by extraction solvents. J. Food Sci. 72(2):S159-S166. DOI: 10.1111/j.1750-3841.2006.00260.x.

Xu, B. and S.K.C. Chang. 2008a. Total phenolics, phenolic acids, isoflavones, and anthocyanins and antioxidant properties of yellow and black soybeans as affected by thermal processing. J. Agric. Food Chem. 56 (16):7165-7175. DOI: 10.1021/jf8012234.

Xu, B. and S.K.C. Chang. 2008b. Antioxidant capacity of seed coat, dehulled bean, and whole black soybeans in relation to their distributions of total phenolics, phenolic acids, anthocyanins, and isoflavones. J. Agric. Food Chem. 56(18):8365-8373. DOI: 10.1021/jf801196d.

Yusnawan, E. 2013. The effect of rhizobium inoculation on total flavonoid, phenolic contents, and antioxidant activity of soybean seeds. p. SA 478-SA 482. In: Sularso, N. A. Indriyani (eds). Proceedings National Olympiad and International Conference. Purwokerto. Muhammadiyah Purwokerto University. SA 478-SA 482.

DOI: http://dx.doi.org/10.21082/jpptp.v1n1.2017.p73-81


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