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Teosinte (Dioon Mejiae) Flour: Nutritional and Physicochemical Characterization of the Seed Flour of the Living Fossil in Honduras

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Teosinte (Dioon Mejiae) Flour: Nutritional and Physicochemical Characterization of the Seed Flour of the Living Fossil in Honduras agronomy Article Teosinte (Dioon mejiae) Flour: Nutritional and Physicochemical Characterization of the Seed Flour of the Living Fossil in Honduras José-Miguel Bastias-Montes 1,*, Laura-Elena Flores-Varela 1, Onán-Alonso Reyes-Calderón 2, Carla Vidal-San-Martín 1, Ociel Muñoz-Fariña 3 , Roberto Quevedo-León 4 and Sergio-Miguel Acuña-Nelson 1,* 1 Department of Food Engineering, Universidad del Bío-Bío, Av. Andrés Bello 720, Chillán 3780000, Chile; laura.fl[email protected] (L.-E.F.-V.); [email protected] (C.V.-S.-M.) 2 School of Biology, Universidad Nacional Autónoma de Honduras, Tegucigalpa 11101, Honduras; [email protected] 3 Institute of Food Sciences and Technology, Universidad Austral de Chile, Valdivia 5090000, Chile; [email protected] 4 Department of Aquaculture and Agrifood Resources, Universidad de Los Lagos, Osorno 5290000, Chile; [email protected] * Correspondence: [email protected] (J.-M.B.-M.); [email protected] (S.-M.A.-N.); Tel.: +56-42-2463042 (J.-M.B.-M.); +56-42-2463172 (S.-M.A.-N.) Received: 2 March 2020; Accepted: 25 March 2020; Published: 1 April 2020 Abstract: Teosinte (Dioon mejiae) is a dioecious tree native to Honduras, whose seeds are used to make flour for the preparation of traditional foods and beverages. The objective was to evaluate the nutritional and physicochemical composition of teosinte flour for the first time. Using diverse techniques, teosinte flour was found to be a high-calorie food rich in total carbohydrates and mainly composed of starch, with an amylopectin:amylose ratio of 2:1 and a concentration of resistant starch greater than 50%. Its proteins were similar to other cereals in which the essential amino acids glutamic acid, leucine, and especially lysine were the most important. Some 75% of its total dietary fiber was insoluble. The fatty acid profile was characterized by a high unsaturated fatty acid content in which oleic acid (C18:1) and linoleic acid (C18:2) predominated. As for minerals, teosinte flour had higher iron content, lower sodium concentration, and similar zinc, calcium, and phosphorus content to other cereal flours. We highlight that teosinte flour has nutrients and qualities that convert it into flour with excellent nutritional abilities and health benefits; it is also a very good industrial and technological alternative to be mixed mainly with types of flour from other sources. Keywords: Dioon mejiae; teosinte; flour; physicochemical composition; nutritional composition 1. Introduction Dioon mejiae, commonly known as teosinte, is a living fossil tree found in Honduras, originating from the Department of Olancho where the highest density of this species is located in the communities of Rio Grande and Saguay. This teosinte is a cycad, a group of plants that arose long before the division between monocots and dicotyledons used for all modern cereals used for food, so it is not related to the subspecies of Zea mays also known by the name commonly used, teosinte. The first scientific publication about this species, which named it, dates back to 1950 [1]. This work provides botanical background, information about its location, and how the natives used the seeds. We now know that, since time immemorial, the natives and criollos of this zone have obtained the seed from the female cones of this arboreal species, which was previously treated with water and Agronomy 2020, 10, 481; doi:10.3390/agronomy10040481 www.mdpi.com/journal/agronomy Agronomy 2020, 10, 481 2 of 12 quicklime, washed on repeated occasions, dried, and then ground to get flour. This cycad species flour is used to prepare typical foods such as bread, donuts, tamales, and tortillas. In addition to the seed, the starch called sago is obtained and used as a dietary supplement by indigenous communities [2]. This cycad species’ seeds have high starch contents and other components of nutritional interest that need to be characterized [3]. To date, no bibliographical information exists in which this cycad species flour is characterized or its nutritional contributions are compared with frequently used products. Based on this assumption, the objective of the present study was to establish the physicochemical and nutritional characterization of teosinte flour. 2. Materials and Methods 2.1. Raw Material Teosinte seeds (Dioon mejiae) from the community of Rio Grande in the municipality of Gualaco, Olancho, Honduras, were used. Collected seeds were treated with water and quicklime, washed on repeated occasions, dried in the shade at ambient temperature, and then ground in a hand mill to obtain flour, which was sifted in a Mesh 35 (500 µm) sieve. Finally, the flour was stored in sealed plastic bags until further analysis. 2.2. Proximate Analysis Moisture content (method 935.29), fat (method 954.02), ash (method 923.03), and proteins (method 991.20) were determined according to the AOAC [4]. The total carbohydrate content was calculated by difference [5] and the calorie content was calculated by the Atwater coefficient [6]. Analyses were repeated twice, and each analysis was undertaken in triplicate. 2.3. Determination of Free Sugars Fructose, glucose, and saccharose were determined by weighing 10 g of this cycad species flour in a 50 mL falcon tube and adding 30 mL of Milli-Q water; ultrasound was applied for 30 min, the tube was heated at 80 ◦C for 10 min, 5 mL Carrez I and 5 mL Carrez II reagent were added, and the solution was diluted to 50 mL with Milli-Q water. The solution was centrifuged at 5000 rpm for 5 min, 1 mL solution was put in an Eppendorf tube, and underwent ultracentrifugation at 12,000 rpm for 10 min. The previously mentioned sugars were determined with high-performance liquid chromatography (HPLC) equipment consisting of a gradient pump (L-6200A), differential refractometer detector (RI-71, Merck-Hitachi,), autosampler (717 plus, Waters®), column oven (Waters®), equipped with a column (Purospher STAR-NH , 250 4.6 mm, 5 µm, Merck) and a precolumn (Purospher STAR-NH , 4 4 mm, 2 × 2 × Merck), and PC chromatography software (Clarity). The separation was achieved using the method described by Doughty [7], and the injection volume was 10 µL. Acetonitrile-water was used as a mobile phase at a 75:25 ratio in an isocratic mode; the flow was adjusted to 1.2 mL/min, and the column oven temperature was set at 30 ◦C. Each compound was identified by comparing retention times with the reference standards fructose, glucose, and saccharose (Merck). 2.4. Determination of Fatty Acid Profile Lipids were extracted using the methodology proposed by Bligh and Dyer [8]. Basically, 10 mL organic phase (lower phase) was filtered with Whatman filter paper N◦ 1; the filtrate was evaporated in a rotary evaporator at 60 ◦C and then dried in an air oven at 60 ◦C for 2 h. Fatty acid methylation was performed using the procedure proposed by Hartman and Lago [9]; the fatty extract was put in a 10 mL glass tube with screw cap, 1 mL NaOH 0.5 N in methanol was added, and it was heated under reflux for 10 min in a water bath at 100 ◦C. Methylation was then performed by adding 1 mL methylation reagent (1 g NH4Cl in MeOH:H2SO4, 30:1.5) and heated under reflux for 10 min in a water bath at 100 ◦C. The fatty acid methyl esters were recovered by adding 2 mL saturated NaCl and 1 mL hexane to the methylation tubes. Finally, the tubes were centrifuged until phase separation. Agronomy 2020, 10, 481 3 of 12 Afterward, 1 mL of the upper phase (hexane) was transferred to 1.5 mL vials and these were analyzed in a gas chromatograph (GC-2010, Shimadzu) equipped with an autosampler (AOC-20s), auto-injector (AOC-20i), FID detector, and a capillary column (100 m 0.25 mm 0.20 µm, Rt-2560, Restek) using × × helium as a gas carrier. The temperature of both injector and detector (FID) was 250 ◦C. The column oven was programmed with temperature ramps from 120 to 220 ◦C. The fatty acid methyl esters were identified by comparing with the retention times of a mixture of reference standards of saturated fatty acid (C4-C24) (1000 µg/mL in hexane, analytical standard, 49453-U, Supelco), unsaturated fatty acids (C4-C24) (wt.% varied, analytical standard, 18919-1AMP, Supelco), linoleic acid methyl esters combined with cis-9 and trans-9,11 isomers, and octodecadienoic acid methyl esters cis-10 and trans-12 (05632, Sigma). 2.5. Determination of Amino Acid Profile The total amino acid profile was determined using the method described by Cohen et al. [10]. A 1 g sample was hydrolyzed with 10 mL 1% phenol in HCl 6 N in a hermetically sealed glass tube and heated at 120 ◦C for 24 h. Samples were derivatized with phenylisothiocyanate (PITC). Quantification was performed by HPLC with a detector (L-4250 UV-VIS, Merck-Hitachi), 254 nm wavelength, and a column (Hibar®RT 100-4,6 LiChrospher®100 RP-18 endcapped, 5 µm, Merck). An acetonitrile gradient acetate buffer (pH 6.4) at 30 ◦C was used as the mobile phase, and the analysis was completed in 37 min. 2.6. Minerals Iron (Fe), zinc (Zn), calcium (Ca), and sodium (Na) were determined by atomic absorption spectrophotometry (AAS) and flame emission spectrometry according to AOAC 985.3 [4]. The calcined samples were hydrated with 1 mL Milli-Q water and 1 mL concentrated hydrochloric acid for complete solubilization. After 1 h, it was filtered with Whatman filter paper N◦ 1, diluted to 50 mL with 15 MW 1 cm− Milli-Q water. The solutions were taken to the AAS (SpectrAA 55, Varian); the quantity of minerals was measured at the corresponding wavelengths with the respective hollow cathode lamps.
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