Revista Mexicana Ciencias Agrícolas   volume 12   number 3   April 01 - May 15, 2021

DOI: https://doi.org/10.29312/remexca.v12i3.2599

Investigation note

Morphometric analysis of flour, starch and protein of Tropaeolum
tuberosum Ruiz and Pavón

Deysi Guevara-Freire1§

Diana Coello-Fiallos2

1Faculty of Agricultural Sciences-Campus Querochaca-Technical University of Ambato. Cevallos-Quero Highway, Tungurahua, Ecuador. ZC. 180602. Tel. 59 32580191. 2Group of Materials Science and Technology-Faculty of Civil and Mechanical Engineering-Huachi Chico Campus-Technical University of Ambato. Av. Los Chasquis and Río Payamino, Ambato, Ecuador. CP. 180207. Tel. 59 30328411. (jc.coello@uta.edu.ec).

§Corresponding author: da.guevara@uta.edu.ec.

Abstract

Mashua (Tropaeolum tuberosum Ruiz and Pavón) is an edible tuber and was domesticated in the central Region of the Andes; it has little importance compared to other Andean crops, but it has great potential to be exploited in the industry. For this reason, the objective of this study was to determine the morphological characteristics of the starch, flour and freeze-dried protein of four mashua genotypes grown in the herbarium of the Faculty of Agricultural Sciences in 2018. Using the scanning electron microscope (MEB), the results showed flour granules with spherical, oval shapes with a diameter greater than and less than 6 µm and 5 µm in the yellow genotype (MA); 9.4 µm and 8.6 µm in the white genotype (MB), 8.3 µm and 6.9 in the yellow green genotype (MVA), 9.9 µm and 9.1 µm in the poza rondador genotype (MPR). Starch granules have sizes of 7.1 µm and 6.1 µm (MA), 18 µm and 16.3 µm (MB), 8.3 µm and 7.3 µm (MVA), 7 µm and 6.5 µm (MPR) with spherical, oval and spherical-truncated shapes. While the micrographs of the freeze-dried protein were observed sheets with sizes of 130, 221, 224 and 237 µm (MVA, MA, MB and MPR, respectively). The oval and spherical shape are highlighted in the flour and starch granules and the flattened polygonal shape in the protein, while the largest were the MB and MPR genotypes (starch and flour) and in the case of the MPR protein.

Keywords: Andean, granule shape, size.

Reception date: February 2021

Acceptance date: March 2021

In pre-Columbian times the Andean peoples domesticated about 70 crops in Ecuador, nine species have been studied and one tuber named mashua stands out for its nutritional and medicinal qualities. Tropaeolum tuberosum Ruiz & Pavón is the scientific name for this species, which is taxonomically located in the family Tropaeolaceae, genus Tropaeolum (Lim, 2016).

The genetic diversity of the tuber is varied but less compared to the oca (Oxalis tuberosa) and the olluco (Ullucus tuberosus) (Pissard et al., 2008), between genotypes are differentiated by their color, shape and flavor (Flores et al., 2003). The tuber has an important nutritional composition, with high levels of ascorbic acid (vitamin C), riboflavin (vitamin B2), thiamine (vitamin B1) (National Research Council, 1989), high protein content even higher than potato, oca and olluco (Manrique et al., 2013), fiber (Flores et al., 2003) and high carotene content (Barrera et al., 2004).

The uses of mashua are varied, it stands out in culinary preparations (Barrera et al., 2004) and traditional medicine for its bioactive compounds beneficial for health (Campos et al., 2006). Andean farmers usually consume the entire plant and especially the tuber (Flores et al., 2003) from which they obtain by-products such as flour and starch. From the first processing of mashua as flour, a spicy product is obtained, with a strong-smell and a yellowish appearance, which is classified as unsatisfactory (Grau et al., 2003). Ramallo (1999), studied the composition of mashua flour, reporting 8% protein comparable to cornmeal and high energy content 416 kcal 100 g-1, suggesting it as pig food.

On the other hand, starch was noted for its nutritional properties (Grau et al., 2003), creating great interest in researchers. Mashua contains 41.35% (BS) starch in the tuber, with 27.44% (BS) amylose (Valcárcel-Yamani et al., 2013). Mashua starch has smaller granules compared to tubers such as oca and ulluco, it is composed of 2-3 heterogeneous grains (Surco, 2004). Its shape can become ovoid, spherical and truncated with sizes of 4.39 and 16.29 µm in length and 4.07 and 13.09 µm diameter (Valcárcel-Yamani et al., 2013).

According to Valcárcel-Yamani et al. (2013) the starch of Andean tubers such as oca, ulluco and mashua are considered of importance in the food industry for the production of products of easy cooking, high viscosity and low stability to refrigeration, but unlike oca and ulluco mashua stands out for its high digestibility when possessing small granules.

The shape and size of mashua starch granules has been discussed by several researchers; however, recent research on the morphology of mashua flour and protein is not yet known, the objective of this research was to characterize morphometrically the starch, flour and protein of four vegetable samples of mashua and look for its differences and similarities with other similar crops.

Plant material

The tubers of four mashua genotypes (T. tuberosum) were collected from the germplasm bank of the Faculty of Agricultural Sciences of the Technical University of Ambato. Genotypes were cultivated and harvested in 2018; from four mashua accessions from different geographical origins of Ecuador: the yellow genotype (MA) of the province of Tungurahua, yellow green (MVA) and poza rondador (MPR) in the province of Chimborazo and white (MB) of Cotopaxi province, these correspond to vulgar names acquired by local sources of peasants (Valle-Parra et al., 2018).

The tubers were selected for their uniform size and no bruises. Mashua starch extraction. Starch extraction was carried out four days after harvest, using the methods described by Torruco-Uco and Betancur-Ancona (2007), with slight modifications.

The tubers were washed and brushed to remove all impurities (dust, leaves, roots) and then cut into cubes of approximately 3 cm on each side. The cubes were crushed in a blender with distilled water in a ratio of 1:2 (p/v) for 3 min. The resulting suspension was filtered with a muslin cloth to remove the fiber and the filtrate was left to settle at 4 oC for 4 hours in a precipitation vessel. The starch was separated from the supernatant by centrifugation at 3 000 rpm for 5 min (Melian, 2010) in a Sorvall ST 8 (X-150 Swinging Bucket Rotor) centrifuge, the supernatant was discarded.

The starch obtained was re-suspended in water and left in sedimentation, then it was filtered as described above. This procedure was repeated 3-4 times with distilled water and one last time it was filtered through a cotton layer until obtaining a white starch and a floating translucent. The clean starch was dried on a stove (Modell 100-800) for 24 hours at 37 oC, ground into a mortar and sieved until obtaining a homogeneous powder. The starch was stored in plastic bottles with a hermetic lid closure and stored at -20 oC.

Scanning electron microscopy. The structural morphology of starch, flour and protein of mashua was studied using a scanning electron microscope (MEB) (model Tescan Vega3) with an increase of up to 100.000X, the images were made working at 20.0 keV in high vacuum. The samples were prepared and fixed on double-sided pieces of carbon tape and coated with gold using an SPI-Module cathodic spray coating device.

For data tabulation, random measurements of the major or longitudinal diameter (DM), minor or transverse diameter (Dm) and length (L) of starch granule, flour and protein sheets micrographs were performed. Statistical analysis. For statistical analysis, the InfoStat version 2008 program was used, using descriptive statistics, with Tukey test to determine significant differences between averages. Figure 1 details the morphology of starch granules and freeze-dried samples of flour and protein from four mashua genotypes.

In the first column there are micrographs of flour highlighting the oval and spherical shape in the MB, MA and MPR genotypes while in the MVA genotype predominates the oval shape, in addition in all flour samples it can see the presence of particulate matter and the presence of sheets that could correspond to protein. All flour samples have smooth appearance surfaces such as potato starch granules (Solanum tuberosum) (Zhou et al., 2013), with some cracks similar to maca (Zhang et al., 2017) except in MA flour granules where bumps and small grooves similar to corngranules are observed (Singh et al., 2003). The size of the MB and MVA granules do not differ significantly otherwise from the MA and MPR genotypes (Table 1).

Table 1. Size and shape of starch granules, flour and protein sheets of four mashua genotypes.

a-b= different superscripts in the same row indicate significant differences (p> 0.05). Values in brackets indicate the maximum and minimum values observed.

MA flour granules have average diameters of 6.4 µm (DM) and 5 µm (Dm), the MB genotype reports 9.4 µm (DM) and 8.6 µm (Dm), the MVA genotype has sizes of 8.3 µm (DM) and 6.9 µm (Dm) and MPR flour granules report 9.9 µm (DM) and 9.1 µm (Dm). The micrographs of the starch granules are shown in the second column in Figure 1. The starch granules of the four genotypes have a morphology where oval and spherical structures predominate and with less participation the spherical-truncated forms.

These similarities are exposed in the four genotypes, results consistent with other studies, such as yellow mashua granule micrographs reported by Valcárcel-Yamani et al. (2013) and other tubers such as makal (Xanthosoma yucatanensis) (Torruco-Uco and Betancur-Ancona, 2007) and arrowroot (Maranta arundinacea) (Peroni et al., 2006).

The size of starch granules is similar between MA, MVA and MPR genotypes, while the MB genotype differs by having larger granules (Table 1). The larger diameters of the granules have averages from 18 µm for the MB genotype, 7.1 µm in MA, 8.3 µm in MVA and 7 µm in MPR and diameters less than 6.1, 16.3, 7.3 and 6.5 µm respectively.

These values are similar to the size of granules reported in yellow mashua genotypes with 4.39 and 16.29 µm (DM) and 4.07 and 13.09 µm (Dm) (Valcárcel-Yamani et al., 2013). Also, other similar values have been reported such as maca (Lepidium meyenii Walpers) (9 µm), cassava (Manihot sculpted) (12.9 µm) and makal (12.4 µm) (Charles et al., 2005; Hernández-Medina et al., 2008; Torruco-Uco and Betancur-Ancona, 2007; Zhang et al., 2017).

Figure 1. Electronic micrographs (SEM) from left to right: flour, starch and mashua protein genotype MB (A-A2); MA (B-B2); MVA (C-C2); MPR (D-D2) with 100000 X amplification.

Differences in granule size and shape can be attributed to several factors, one of them agriculture, researchers expose that Andean crops over time have undergone changes in domestication and reproduction suggesting an influence on the morphological characteristics and size of the starch granule (King and Gershoff, 1987; Flores et al., 2003); however, with modern advances today it is believed that the size of the granule could be related to agricultural practices (Torres et al., 2011).

Certain studies attribute the results to the biological origin, biochemistry of chloroplast and plant physiology (Singh et al., 2003). While, other authors argue that the morphology and size of starch granules depend on the genetics of the plant, in addition to the membranous structures and physical characteristics of the plastids (Lindeboom et al., 2004).

The last column reports protein micrographs of the four mashua genotypes (Figure 1). The shape of the protein is peculiar as they are not spherical or oval but laminar structures with brittle surface and asymmetrical polygonal shape. These sheets have different average lengths ranging from 130, 221, 224 and 237 µm (MVA, MA, MB and MPR, respectively) (Table 1), its shape resembles the beach pea albumin (Lathyrus maritimus L.) with plate topography and smooth surface (Chavan et al., 2001) and the bean albumin (Phaseolus vulgaris L.) Great Northern variety in the shape of rod (Sathe and Salunkhe, 1981). Past research has indicated that the topographical characteristics of proteins could intervene in the physicochemical and functional properties of starch (Chavan et al., 2001), but being at low levels would have no negative effects according as exposed by Hoover (2001).

Conclusions

The study of starch morphology showed oval and spherical shapes similar to flour granules, but different from the freeze-dried proteins with laminar structures with asymmetric polygonal shape, regarding the size of the starch and flour granules, it was observed that the MB and MPR genotypes were the ones that stood out in the largest size respectively, while the protein sheets had significant differences being the largest MPR genotype.

Acknowledgments

The authors wish to express their deep appreciation to the engineer Cristian Pérez coordinator to the Materials Laboratory of the Faculty of Civil and Mechanical for her valuable support with the use of the scanning electron microscope.

Cited literature

Torres, F. G.; Troncoso, O. P.; Díaz, D. A. and Amaya, E.; 2011. Morphological and thermal characterization of native starches from Andean crops. Starch-Starke. 63(6):381-389. https://doi.org/10.1002/star.201000155.

Barrera, V.; Espinosa, P.; Tapia, C.; Monteros, A. y Valverde, F. 2004. Caracterización de las raíces y los tubérculos andinos en la ecoregión andina del Ecuador (Capítulo 1). In: Barrera, V.; Tapia, C. y Monteros, A. (Ed.). Raíces y tubérculos andinos: alternativas para la conservación y uso sostenible en el Ecuador. Serie: conservación y uso de la biodiversidad de raíces y tubérculos andinos: una década de investigación para el desarrollo (1993-2003). Instituto Nacional Autónomo de Investigaciones Agropecuarias- Centro Internacional de la Papa, Agencia Suiza para el Desarrollo y la Cooperación. Quito, Ecuador-Lima, Perú. No. 4. 3-30 pp.

Campos, D.; Noratto, G.; Chirinos, R.; Arbizu, C.; Roca, W. and Luis, C. Z. 2006. Antioxidant capacity and secondary metabolites in four species of Andean tuber crops: native potato (Solanum sp.), mashua (Tropaeolum tuberosum Ruiz & Pavón), Oca (Oxalis tuberosa Molina) and ulluco (Ullucus tuberosus Caldas). J. Sci. Food Agric. 86(2):1481-1488. https://doi.org/10.1002/jsfa.

Charles, A. L.; Chang, Y. H.; Ko, W. C.; Sriroth, K. and Huang, T. C. 2005. Influence of amylopectin structure and amylose content on the gelling properties of five cultivars of cassava starches. J. Agric. Food Chem. 53(7):2717-2725. https://doi.org/10.1021/ jf048376+.

Chavan, U. D.; McKenzie, D. B. and Shahidi, F. 2001. Protein classification of beach pea (Lathyrus maritimus L.). Food Chem. 75(2):145-153. https://doi.org/10.1016/S03088146(01)00122-4.

Flores, H. E.; Walker, T. S.; Guimarães, R. L.; Bais, H. P. and Vivanco, J. M. 2003. Andean root and tuber crops: Underground rainbows. HortScience. 38(2):161-167. https://doi.org/ 10.21273/hortsci.38.2.161.

Grau, A., Ortega Dueñas, R., Nieto Cabrera, C., y Hermann, M. 2003. Mashua Tropaeolum tuberosum Ruíz & Pav. Promoting the conservation and use of underutilized and neglected crops.1ra Ed. Lima-Peru y Roma-Italia. International Potato Center/International Plant Genetic Resources Institute. 54 p.  https://www.bioversityinternational.org/fileadmin/-migrated/uploads/tx-news/Mashua-Tropaeolum-tuberosum-Ru%c3%adz-amp-Pav.-880.pdf.

Hernández-Medina, M.; Torruco-Uco, J. G.; Chel-Guerrero, L. and Betancur-Ancona, D. 2008. Caracterización fisicoquímica de almidones de tubérculos cultivados en Yucatán, México. Ciencia y Tecnología Alimentos Campinas. 28(3):718-726.

Hoover, R. 2001. Composition, molecular structure, and physicochemical properties of tuber and root starches: a review. Carbohydrate Polymers. 45(3):253-267.

King, S. R. and Gershoff, S. N. 1987. Nutritional evaluation of three underexploited andean tubers:Oxalis tuberosa (Oxalidaceae), Ullucus tuberosus (Basellaceae), and Tropaeolum tuberosum (Tropaeolaceae). Economic Botany. 41(4):503-511. https://doi.org/10.1007/ BF02908144.

Lim, T. K. 2016. Edible medicinal and non medicinal plants. Dordrecht, Alemania. Springer. 12:94-102. https://doi.org/10.1007/978-94-017-9511-1.

Lindeboom, N.; Chang, P. R. and Tyler, R. T. 2004. Analytical, biochemical and physicochemical aspects of starch granule size, with emphasis on small granule starches: a review. Starch-Stärke. 56(34):89-99. https://doi.org/10.1002/star.200300218.

Manrique, I.; Arbizu, C.; Vivanco, F.; Gonzales, R.; Ramírez, C.; Chávez, O. and Ellis, D. 2013. Tropaeolum tuberosum Ruíz & Pav. Colección de germoplasma de mashua conservada en el Centro Internacional de la Papa (CIP). Centro internacional de la Papa. 1ra. Ed. Lima, Perú. 122 p.

Melian, D. 2010. Ensayo comparativo de dos metodologías de extracción de almidón de papa usando muestras de diez variedades nativas de Chiloé y dos Daniela Elizabeth Melian Subiabre. Universidad Austral de Chile.

National Research Council. 1989. Lost Crops of the Incas: Little-Known Plants of the Andes with Promise for Worldwide Cultivation. In: National Academy Press. 1ra. ed. Washington, DC. 66-73 p. https://doi.org/10.17226/1398.

Peroni, F. H. G.; Rocha, T. S. and Franco, C. M. L. 2006. Some structural and physicochemical characteristics of tuber and root starches. Food Sci.Technol. Inter. 12(6):505-513. https://doi.org/10.1177/1082013206073045.

Pissard, A.; Arbizu, C.; Ghislain, M. and Bertin, P. 2008. Influence of geographical provenance on the genetic structure and diversity of the vegetatively propagated andean tuber crop, Mashua (Tropaeolum tuberosum), highlighted by intersimple sequence repeat markers and multivariate analysis methods. Inter. J. Plant Sci. 169(9):1248-1260. https://doi.org/10.1086/591979.

Ramallo, R. 1999. Diseño de una planta procesadora de harina de isaño como base de la alimentación porcina. Universidad Privada Boliviana. Cochabamba, Bolivia.

Sathe, S. K. and Salunkhe, D. K. 1981. Functional properties of the great northern bean (Phaseolus vulgaris L.) proteins: emulsion, foaming, viscosity, and gelation properties. J. Food Sci. 46(1):71-81. https://doi.org/10.1111/j.1365-2621.1981.tb14533.x.

Singh, N.; Singh, J.; Kaur, L.; Singh, N. and Singh, B. 2003. Morphological, thermal and rheological properties of starches from different botanical sources. Food Chem. 81(2):219-231. https://doi.org/10.1016/S0308-8146(02)00416-8.

Surco, F. 2004. Caracterización de almidones aislados de tuberculos andinos: mashua (Tropaeolum tuberosum), Oca (Oxalis tuberosa), Olluco (Ullucus tuberosus) para su aplicación tecnológica. 1-55 pp. http://cybertesis.unmsm.edu.pe/handle/cybertesis/2588.

Torruco-Uco, J. and Betancur-Ancona, D. 2007. Physicochemical and functional properties of makal (Xanthosoma yucatanensis) starch. Food Chem. 101(4):1319-1326. https://doi.org/10.1016/j.foodchem.2006.03.047.

Valcárcel-Yamani, B.; Rondán-Sanabria, G. G. and Finardi-Filho, F. 2013. The physical, chemical and functional characterization of starches from andean tubers: Oca (Oxalis tuberosa molina), olluco (Ullucus tuberosus caldas) and mashua (Tropaeolum tuberosum Ruiz & Pavón). Brazilian J. Pharmaceutical Sci. 49(3):453-464. https://doi.org/10.1590/S1984-82502013000300007.

Valle-Parra, M.; Pomboza-Tamaquiza, P.; Buenaño-Sánchez, M.; Guevara-Freire, D.; Chasi-Vizuete, P.; Vásquez, C. and Pérez-Salinas, M. 2018. Morphology, phenology, nutrients and yield of six accessions of Tropaeolum tuberosum Ruiz y Pav (Mashua). Trop. Subtrop. Agroecosystems. 21(1):131-139.

Zhang, L.; Li, G.; Wang, S.; Yao, W. and Zhu, F. 2017. Physicochemical properties of maca starch. Food Chem. 218(1):56-63. https://doi.org/10.1016/j.foodchem.2016.08.123.

Zhou, Q.; Shi, W.; Meng, X. and Liu, Y. 2013. Studies on the morphological, crystalline, thermal properties of an underutilized starch from yam Dioscoreae zingiberensis C. H. Wright. Starch/Staerke. 65(1-2):123-133. https://doi.org/10.1002/star.201200058.