Spectrophotometric Quantification of Phytic Acid During Embryogenesis in Bambara groundnut (Vigna subterranea L.) Through Phosphomolybdenum Complex Formation

  • Takudzwa Mandizvo University of KwaZulu-Natal, Crop Science Discipline, Private BagX01, Scottsville, 3209 Pietermaritzburg, South Africa http://orcid.org/0000-0003-2078-3041
  • Alfred Oduor Odindo University of KwaZulu-Natal, Crop Science Discipline, Private BagX01, Scottsville, 3209 Pietermaritzburg, South Africa


In relation to seed vigour and hyperosmotic stress, phytic acid has implications in imparting drought tolerance and enrichment of seed mineral reserves respectively. The present study was undertaken to determine the variation in phytic acid during seed development and physiological maturity of 4 Bambara groundnut landraces. The landraces were grown in field during 2017–2018 rain season at Ukulinga, Pietermaritzburg. The phytate content was estimated indirectly from 14-65 days after flowering (DAF) by using a spectrophotometer, evaluating the total extractable phosphorus absorbance at 720 nm. An analysis is described for the rapid determination of phosphorus in developing seeds. The colour complex (phosphomolybdenum) formed under acidic conditions absorbs maximally at 720 nm in acidic (pH<4.5) solutions. Absorbance of the chromophore when measured spectrophotometrically at 720 nm, it obeys Beer’s law over the range of 0 to 75 ppm of standard phosphorus solution. There were significant differences (P<0.001) in total extractable phosphorus at 14, 21, 28, 35, 42 and 65 DAF. The highest and lowest extractable phosphorus was recorded in G340A and Kazai respectively. Pi seed content was between 1.51 and 5.69 mgkg-1 at 14 DAF, at physiological maturity (65 DAF) Pi was recorded between 21.73 and 32.23 mgkg-1. We drew conclusions that Bambara groundnut landraces may differ in both (1) phytic acid accumulation rate and (2) phytic acid content at physiological maturity. The results reported open the possibility of a specific seed selection criterion for improving the mineral element value of Bambara groundnut through the identification of landraces with high-Pi (phytic acid).

Keywords: days after flowering, Beer's law, embryogenesis, inorganic phosphorous, Keggin ion, phytic acid


Bailly C. (2004) Active oxygen species and antioxidants in seed biology. Seed Science Research 14:93-107. DOI: 10.1079/Ssr2004159.
Bilyeu K.D., Zeng P., Coello P., Zhang Z.J., Krishnan H.B., Bailey A., Beuselinck P.R., Polacco J.C. (2008) Quantitative conversion of phytate to inorganic phosphorus in soybean seeds expressing a bacterial phytase. Plant Physiol 146:468-77. DOI: 10.1104/pp.107.113480.
Biorender. (2020) Biorender templates and illustrations, Biorender https://app.biorender.com/.
Bohn L., Josefsen L., Meyer A.S., Rasmussen S.K. (2007) Quantitative analysis of phytate globoids isolated from wheat bran and characterization of their sequential dephosphorylation by wheat phytase. J Agric Food Chem 55:7547-52. DOI: 10.1021/jf071191t.
Chibarabada T.P., Modi A.T., Mabhaudhi T. (2015) Bambara groundnut (Vigna subterranea) seed quality in response to water stress on maternal plants. Acta Agriculturae Scandinavica Section B-Soil and Plant Science 65:364-373. DOI: 10.1080/09064710.2015.1013979.
Dhole V.J., Reddy K.S. (2016) Association of phytic acid content with biotic stress tolerance in mungbean (Vigna radiata L. Wilczek). Phytoparasitica 44:261-267. DOI: 10.1007/s12600-016-0514-5.
Dijk D.v. (2007) Wageningen Evaluating Programmes for Analytical Laboratories (Wepal): A World of Experience. Communications in Soil Science and Plant Analysis 33:2457-2465. DOI: 10.1081/css-120014460.
Doria E., Galleschi L., Calucci L., Pinzino C., Pilu R., Cassani E., Nielsen E. (2009) Phytic acid prevents oxidative stress in seeds: evidence from a maize (Zea mays L.) low phytic acid mutant. J Exp Bot 60:967-78. DOI: 10.1093/jxb/ern345.
Dost K., Tokul O. (2006) Determination of phytic acid in wheat and wheat products by reverse phase high performance liquid chromatography. Analytica Chimica Acta 558:22-27. DOI: 10.1016/j.aca.2005.11.035.
Ficco D.B.M., Riefolo C., Nicastro G., De Simone V., Di Gesu A.M., Beleggia R., Platani C., Cattivelli L., De Vita P. (2009) Phytate and mineral elements concentration in a collection of Italian durum wheat cultivars. Field Crops Research 111:235-242. DOI: 10.1016/j.fcr.2008.12.010.
Iwai T., Takahashi M., Oda K., Terada Y., Yoshida K.T. (2012) Dynamic changes in the distribution of minerals in relation to phytic acid accumulation during rice seed development. Plant Physiol 160:2007-14. DOI: 10.1104/pp.112.206573.
Jennings A.C., Morton R.K. (1963) Changes in Nucleic Acids and Other Phosphorus-Containing Compounds of Developing Wheat Grain. Australian Journal of Biological Sciences 16:332-&. DOI: Doi 10.1071/Bi9630332.
John M.K. (1970) Colorimetric Determination of Phosphorus in Soil and Plant Materials with Ascorbic Acid. Soil Science 109:214-&. DOI: Doi 10.1097/00010694-197004000-00002.
Joshi-Saha A., Reddy K.S. (2015) Repeat length variation in the 5'UTR of myo-inositol monophosphatase gene is related to phytic acid content and contributes to drought tolerance in chickpea (Cicer arietinum L.). J Exp Bot 66:5683-90. DOI: 10.1093/jxb/erv156.
Karner U., Peterbauer T., Raboy V., Jones D.A., Hedley C.L., Richter A. (2004) myo-Inositol and sucrose concentrations affect the accumulation of raffinose family oligosaccharides in seeds. J Exp Bot 55:1981-7. DOI: 10.1093/jxb/erh216.
Lehrfeld J. (1994) HPLC Separation and Quantitation of Phytic Acid and Some Inositol Phosphates in Foods: Problems and Solutions. Journal of Agricultural and Food Chemistry 42:2726-2731. DOI: 10.1021/jf00048a015.
Manavalan L.P., Guttikonda S.K., Tran L.S., Nguyen H.T. (2009) Physiological and molecular approaches to improve drought resistance in soybean. Plant Cell Physiol 50:1260-76. DOI: 10.1093/pcp/pcp082.
Mandizvo T., Odindo A.O. (2019) Seed mineral reserves and vigour of Bambara groundnut (Vigna subterranea L.) landraces differing in seed coat colour. Heliyon 5:e01635. DOI: 10.1016/j.heliyon.2019.e01635.
Mangkuto R.A., Soelami F.X.N. (2017) Photometric and Colorimetric Measurements of Luminaires Using Goniometer and spectrophotometer in a Dark Chamber. Procedia Engineering 170:226-233. DOI: 10.1016/j.proeng.2017.03.054.
Martínez-Ballesta M.d.C., Egea-Gilabert C., Conesa E., Ochoa J., Vicente M.J., Franco J.A., Bañon S., Martínez J.J., Fernández J.A. (2020) The Importance of Ion Homeostasis and Nutrient Status in Seed Development and Germination. Agronomy 10. DOI: 10.3390/agronomy10040504.
McKie V.A., McCleary B.V. (2016) A Novel and Rapid Colorimetric Method for Measuring Total Phosphorus and Phytic Acid in Foods and Animal Feeds. Journal of Aoac International 99:738-743. DOI: 10.5740/jaoacint.16-0029.
Mussa S.B., Elferjani H.S., Haroun F.A., Abdelnabi F.F. (2009) Determination of available nitrate, phosphate and sulfate in soil samples. International Journal of PharmTech Research 1:598-604.
Nagul E.A., McKelvie I.D., Worsfold P., Kolev S.D. (2015) The molybdenum blue reaction for the determination of orthophosphate revisited: Opening the black box. Anal Chim Acta 890:60-82. DOI: 10.1016/j.aca.2015.07.030.
Nasri N., Kaddour R., Rabhi M., Plassard C., Lachaal M. (2011) Effect of salinity on germination, phytase activity and phytate content in lettuce seedling. Acta Physiologiae Plantarum 33:935-942. DOI: 10.1007/s11738-010-0625-4.
Ozyurek M., Guclu K., Bektasoglu B., Apak R. (2007) Spectrophotometric determination of ascorbic acid by the modified CUPRAC method with extractive separation of flavonoids-La(III) complexes. Anal Chim Acta 588:88-95. DOI: 10.1016/j.aca.2007.01.078.
Pilu R., Landoni M., Cassani E., Doria E., Nielsen E. (2005) The maize lpa241 mutation causes a remarkable variability of expression and some pleiotropic effects. Crop Science 45:2096-2105. DOI: 10.2135/cropsci2004.0651.
Pilu R., Panzeri D., Gavazzi G., Rasmussen S.K., Consonni G., Nielsen E. (2003) Phenotypic, genetic and molecular characterization of a maize low phytic acid mutant (lpa241). Theor Appl Genet 107:980-7. DOI: 10.1007/s00122-003-1316-y.
Pote D., Daniel T. (2000) Analyzing for dissolved reactive phosphorus in water samples. Methods of phosphorus analysis for soils, sediments, residuals, and waters. Southern Coop. Ser. Bull 396:91-93.
Raboy V. (2001) Seeds for a better future: 'low phytate' grains help to overcome malnutrition and reduce pollution. Trends Plant Sci 6:458-62. DOI: 10.1016/S1360-1385(01)02104-5.
Raboy V., Dickinson D.B. (1987) The timing and rate of phytic Acid accumulation in developing soybean seeds. Plant Physiol 85:841-4.
Schoenau J., O’Halloran I. (2008) Sodium bicarbonate-extractable phosphorus. Soil sampling and methods of analysis 2.
Soropa G., Nyamangara J., Nyakatawa E.Z. (2019) Nutrient status of sandy soils in smallholder areas of Zimbabwe and the need to develop site-specific fertiliser recommendations for sustainable crop intensification. South African journal of plant and soil 36:149-151.
Urbano G., Lopez-Jurado M., Aranda P., Vidal-Valverde C., Tenorio E., Porres J. (2000) The role of phytic acid in legumes: antinutrient or beneficial function? J Physiol Biochem 56:283-94. DOI: 10.1007/BF03179796.
Walker K.A. (1974) Changes in phytic acid and phytase during early development of Phaseolus vulgaris L. Planta 116:91-8. DOI: 10.1007/BF00380643.
Weber H., Borisjuk L., Wobus U. (2005) Molecular physiology of legume seed development. Annu Rev Plant Biol 56:253-79. DOI: 10.1146/annurev.arplant.56.032604.144201.
Williams S.G. (1970) The role of phytic acid in the wheat grain. Plant Physiol 45:376-81. DOI: 10.1104/pp.45.4.376.
Zhang W.H., Zhou Y.C., Dibley K.E., Tyerman S.D., Furbank R.T., Patrick J.W. (2007) Nutrient loading of developing seeds. Functional Plant Biology 34:314-331. DOI: 10.1071/Fp06271.
Zhawar V.K., Kaur N., Gupta A.K. (2011) Phytic acid and raffinose series oligosaccharides metabolism in developing chickpea seeds. Physiol Mol Biol Plants 17:355-62. DOI: 10.1007/s12298-011-0080-8.
181 Views | 180 Downloads
How to Cite
Mandizvo, T., and A. Odindo. “Spectrophotometric Quantification of Phytic Acid During Embryogenesis in Bambara Groundnut (Vigna Subterranea L.) Through Phosphomolybdenum Complex Formation”. Emirates Journal of Food and Agriculture, Vol. 32, no. 11, Jan. 2021, pp. 778-85, doi:https://doi.org/10.9755/ejfa.2020.v32.i11.2178. Accessed 28 July 2021.
Research Article