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. 2019 Feb 6;28(4):975–982. doi: 10.1007/s10068-019-00563-2

Effect of nixtamalization processes on mitigation of acrylamide formation in tortilla chips

Alfonso Topete-Betancourt 1, Juan de Dios Figueroa Cárdenas 1,✉, Adriana Lizbeth Rodríguez-Lino 2, Elvira Ríos-Leal 3, Eduardo Morales-Sánchez 4, Héctor Eduardo Martínez-Flores 5
PMCID: PMC6595089  PMID: 31275697

Abstract

Acrylamide can be generated from food components during tortilla chips frying. Thus, the aim of this research was to study different nixtamalization processes as traditional (TNP) with lime [Ca(OH)2], ecological (ENP) with CaCO3, classic nixtamalization (CNP) that uses wood ash and extrusion (EXT) with no Ca+2 source on mitigating the acrylamide formation in deep-fat frying tortilla chips. Acrylamide quantification was done through HPLC–UV. Lower acrylamide content in tortilla chips was for CNP with 46.3 µg/kg, followed by TNP with 55.0 µg/kg, ENP with 694.6 µg/kg and EXP with 1443.4 µg/kg. Differences in acrylamide values among samples can be related to effect of cations (Ca2+, Mg2+, Fe2+, Zn2+, Na+ and K+) present in wood ashes, lime and salts used as raw materials. Correlation of (r = 0.85; p <0.0005) was observed in color of tortilla chips, moisture, texture, blisters, and oil with acrylamide. Nixtamalization process is an effective and inexpensive strategy for acrylamide mitigation.

Keywords: Acrylamide, Nixtamalization, Tortilla chips, Mono- and di-cations

Introduction

Swedish researchers, found the formation of high levels of acrylamide (CH2=CH–CONH2) in some foodstuffs heated at high temperatures, especially in carbohydrate rich foods. This fact has caused worldwide concern because acrylamide has been classified as probably carcinogenic in humans (Tareke et al., 2002). Acrylamide can be formed in food by heat treatment with a range of about 120 to 185 °C, as result of the Maillard browning reaction (Mottram et al., 2002; Stadler et al., 2002). Laboratory studies revealed a temperature–time dependence of acrylamide formation in food. Acrylamide was not found in unheated control or boiled foods (Tareke et al., 2002).

There is a considerable interest in the determination of acrylamide in different samples and the principal effort is in the accuracy assessment from the method during sample preparation. Furthermore, Kim et al. (2011) mentioned the procedures of acrylamide detection in foods which must be reliable, economic and rapid. Oracz et al. (2011) mentioned that due to the high-priced reagents required for the detection of acrylamide and the complicated sample preparation, there are different procedures for acrylamide quantification in food. Hu et al. (2015) compared standard versus rapid detection methods of acrylamide, and concluded that rapid methods still need more enhancement to make it accurate, sensitive, repeatable and reproducible. Wang et al. (2013) reported that high-performance liquid chromatography-ultraviolet (HPLC–UV) represents, a cost-effective method that can be adequate to check the acrylamide content in food products.

Several strategies have been reported to reduce acrylamide level in heat processed foods, among them is the use of aspartic acid, Ca2+ to block the formation of Schiff base of asparagine (Gökmen and Senyuva, 2007a; Kalita and Jayanty, 2013), antioxidants (Salazar et al., 2014), hydrocolloids (Zeng et al., 2011), phospholipids, lecithin, and amino acids (Granvogl and Schieberle, 2006; Zamora et al., 2011), among other additives as effective mitigation agents. Also, the effect of several chemical compounds have been evaluated as agents in the nixtamalization process for reducing the formation of acrylamide on thermally processed foods, specially baked tortilla and frying tortilla chips (Salazar et al., 2014). In addition to the mentioned compounds, the use of chemical additives in foods such as mono, di- and tri-valent cations Na+, K+, Fe2+, Mg2+, Zn2+, Ba2+, V2+, Fe3+, and Al3+ have been investigated as an easy, and inexpensive approach for reducing acrylamide in foods (Elder et al., 2005; Tomoda et al., 2004). However, not all of these additives can be widely applied due to the fact that some of them are not recognized as GRAS, others are expensive and impractical for applying at an industrial scale, and finally, others need to be added in higher levels than authorized to obtain significant decreases in acrylamide (Zamora et al., 2011). The Ca2+ content in corn grains increases during the nixtamalization process and its presence affects directly the reduction of acrylamide content generated during the frying of foods prepared from nixtamalized corn (Salazar et al., 2014). They found that acrylamide content was mainly affected by the ash, calcium, soluble fiber in flour, as well as pH values.

In this context, the basis of nixtamalization processes that use wood ashes, lime and Ca2+ salts, may have the potential to mitigate the acrylamide levels in nixtamalized products.

Tortilla chips are the second place of preferred snacks, especially by children, only behind potato products (Salazar et al., 2014). The high health risks involved in the consumption of acrylamide that is present in those products, require the development of alternatives aimed for reducing the levels of this compound in foods that are consumed by a large segment of the population. Thus, the aim of this research was to study the effect of lime [Ca(OH)2], calcium salts (CaCO3), wood ash cations, as well as, extrusion processes on acrylamide formation in deep-frying tortilla chips.

Materials and methods

Raw materials

Commercial hard endosperm white maize was obtained in a local market in Queretaro, Mexico. It was stored at 4 °C until needed for processing. Lime [Ca(OH)2] and calcium carbonate (CaCO3) salts of 97–99% purity food grade were used. Wood ash used was of oak trees (Quercus) from Oaxaca, Mexico.

Nixtamalization processes

The Traditional nixtamalization process (TNP) reported by Campechano et al. (2012) and the Ecological nixtamalization process (ENP) process patented by Figueroa-Cárdenas et al. (2011) were used in which lime was replaced with calcium salts (calcium carbonate) to obtain whole grain flour and the Classic nixtamalization process (CNP) reported by Mariscal-Moreno et al. (2015) was used for the preparation of masa for elaborating the tortilla chips. The fresh masa was used for elaborating tortillas and those were processed into tortilla chips as indicated below.

Extrusion process (EXP)

Extruded masa without lime was prepared using the following the procedure reported by Martínez-Flores et al. (2002).

Tortilla chips preparation

The tortilla chips were prepared from nixtamalized dough (masa) with coarse particle size, the masa was kneaded with water at 27 °C to obtain the proper consistency to make the tortilla chips. The masa was pressed in a flat tortilla circle using rolls of tortilla machine (Casa González, México, D.F.) with resting time of 2 min. The tortilla was baked on an IR oven at 260 °C for 45 s (15 s, 15 s, 15 s, by each side), followed by 3 min resting time. The tortilla chip was then fried at 180 °C for 42 s and was cooled for 5 min at room temperature, after this time, from each treatment, the sampling was performed randomly with 20 g samples; this represents approximately 10 tortilla chips to evaluate the physicochemical properties.

Tortilla chip features

Color determination in tortilla chip samples

Color changes were determined using a colorimeter MiniScan XE, model 45/0-L (Hunter Associates Laboratory, 11491 Sunset Hill Rd., Reston, Va., U.S.A.). Total color differences (DE) were calculated from the determined CIELAB L* a* b* DE = [(ΔL*)2 + (Δa*)2 + (Δb*)2]½; where L* = brightness or lightness (100 = perfect white, to 0 = black); a* = greenness/redness [negative (green) to positive (red)]; b* = yellowness/blueness [negative (blue) to positive (yellow)]; ΔL*, Δa*, and Δb* = absolute differences of the values between the reference tile (white porcelain) and sample values; ΔE = total difference between reference and sample color. The reference values (calibration) were: L* = 92.10, a* = -0.87 and b* = 0.7.

Physical properties

Tortilla chip thickness was evaluated in 10 tortilla ship samples for each treatment with a digital caliper (model CD-6, Mitutoyo, Kanagawa, Japan). The blister distribution, dimensions as well as areas were performed by using the Image software (National Institutes of Health); and texture analyses were performed by the use of texture analyzer (TA-XT2 Texture Technologies Corporation, Stable Micro Systems, Godalming, U.K) with a TA-52 probe (2 mm in diameter). Each one of the samples was punctured at 0.5 mm/s to a depth of 2 mm. The test was repeated 10 times for each treatment.

Chemical analyses

Determination of fat in tortilla chips was performed by using the approved AACCI (2010) method 30-10.01 with acid hydrolysis; moisture in tortilla chips was determined using the AACCI (2010) method 44-19.01, and the pH with method 943.02 AACCI (2010).

Acrylamide analyses

Acrylamide was evaluated using HPLC–UV with the method reported by Barber et al. (2001), with some modifications. The equipment and materials were a liquid chromatograph Varian 9010 with a UV-201 nm detector 9050 and Columna: Zorbax Eclipse Plus C18 de 4.6 × 150 m × 5 µ (Agilent, Santa Clara, CA, USA). An ultrasonicator Branson 2510 filter of Nylon of 0.22 µm, and Merck acrylamide standards were sodium pentan sulfonate (C5H11NaO3S5), hexane for HPLC from Tecsiquim (Toluca, Edo de Mex, Mexico). The chromatographic conditions were UV detector UV-210 nm, mobil phase: C5H11NaO3S5 mM/H2O:MeOH (99:1)2O5) with a flux: 0.5 mL/min, injection volume of 50 µL.

Three grams of each sample were weighed by triplicate and sonicated with 10 mL of hexane and the hexane was decanted and dried by evaporating the solution. The dried sample was again sonicated with 20 mL of water, filtered with paper and with nylon membrane filters of 0.22 µm, from that solution, a sample of 50 µL was injected to the HPLC chromatograph. The identification was done with retention time using an acrylamide calibration curve (Fig. 1). The data was reported as average of three replications from each sample.

Fig. 1.

Fig. 1

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Typical chromatogram of HPLC-UV and the calibration curve

Mineral analyses

The mineral content (Ca2+, Fe2+, Mg2+, Zn2+, K+ and Na+) was determined in duplicate with the method 40-75.01 using inductively coupled plasma spectroscopy (ICP) (AACC International, 2010).

Statistical Analyses

All evaluations were performed under a completely randomized design using the General Linear Model. The experiments were replicated three times and the average values were reported, the simple Pearson’s correlations were classified as significant and highly significant at p ≤ 0.05 and p ≤ 0.001, respectively. The mean comparison in the analyses of variance were carried out using Tukey’s test with a Statistical Analysis System (SAS Version 9.2, SAS Institute Inc. Cary, NC, USA).

Results and discussion

Effect of the pH, and moisture in the concentrations of acrylamide

Physicochemical properties of the tortilla chip from different nixtamalization processes (TNP, CNP, ENP and EXP), are reported in Table 1. Different values between the pH before and after washing the cooked maize from cooking liquor (nejayote) were obtained. The higher pH was obtained for TNP with the use of calcium hydroxide at 3% with values of 12.1 and 12.4, respectively. This condition promoted alkaline solution and complete solubility of the pericarp that allowed for the diffusion of water and calcium cations in cooked kernels. Due to this, the moisture of tortilla chips from TNP was of 2.5%. This value was similar for tortilla chips from CNP with a moisture of 2.2% but with pH of 7.5–8.6 before and after the washing, respectively. The classic nixtamalization (CNP) that used of wood ash present more di- and mono-valent cations (Table 2) that promoted lightly alkaline solution when these cations were present in water and led to a little pericarp solubility. The pH in the ENP was lightly acid with a range of 5.6–6.2, before and after washing (Table 1). A low pH in food would inhibit acrylamide formation by blocking the nucleophilic addition of asparagine with carbonyl compound and the formation of Schiff base (Pedrechi et al., 2005). However, in the case of the ENP, this effect did not show clearly, although, the concentration of acrylamide was 694.6 µg/kg, so that in the reaction of Maillard, the reactivity of reducing sugar and the amino group was not influenced by the pH as expected, although it was the same for CNP and TNP, where a high pH that represents the optimal conditions for the acrylamide formation, it was not as expected. This may be due to the concentration of different minerals in the tortilla chip during deep-frying (Table 2) (Martins et al., 2001; Weisshaar, 2004). Regarding the extrusion process in acrylamide formation, it had the highest contribution with values of 1443.4 µg/kg, explained in part for the drastic shear conditions of the process that increase the starch gelatinization and release more soluble sugar when compared with the other nixtamalization processes. Besides, the pH was neutral in this process due to the use of only water without any calcium salt, so under these conditions increased the acrylamide level, and these results were agree with that showed by Mulla et al. (2011). On the other hand, Mestdagh et al. (2008) indicated that the addition of different types of organic acids, to obtain a value of pH lower than 5, reduce the formation of acrylamide when compared with the neutral pH 7. However, the results presented in this study show a variation of pH from 6.2 to 12.3 from different processes nixtamalization, suggesting that the most important effect on the acrylamide formation is the amount and type of ions present in the food instead of the pH.

Table 1.

Physicochemical properties of tortilla chips

Process pH0 pHf Moisture (%) Color (ΔE) Texture (N) Blister area (%) Oil (%) Acrylamide (µg/kg)
Classic (Ash) 7.5 ± 0.1b 8.6 ± 0.1b 2.2 ± 0.1c 37.5 ± 0.0c 4.1 ± 0.1b 52.8 ± 1.1a 21.7 ± 0.1a 46.3 ± 0.7c
Traditional (Lime) 12.1 ± 0.1a 12.3 ± 0.1a 2.5 ± 0.1b 36.4 ± 1.0c 4.7 ± 0.5b 31.9 ± 0.6b 18.9 ± 0.1c 55.0 ± 1.5c
Ecological (Carbonate) 5.6 ± 0.1d 6.2 ± 0.1d 1.8 ± 0.0d 39.8 ± 0.6b 3.9 ± 1.0b 50.0 ± 2.0a 21.0 ± 0.3b 694.6 ± 8.3b
Extrusion (only water) 7.0 ± 0.0c 7.0 ± 0.0c 5.3 ± 0.0a 41.7 ± 0.2a 7.9 ± 0.6a 15.1 ± 2.8c 16.2 ± 0.2d 1443.4 ± 4.3a
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Means ± SD (n = 2) followed by the same letter in the same column are not significantly different at p < 0.05

Table 2.

Mineral content (mg/100 g) in tortilla chips, using different nixtamalization processes

Process Ca2+ Fe2+ Zn2+ K+ Mg2+ Na+
Classic (ash) 170.4 ± 1.1c 5.5 ± 0.2ab 13.9 ± 0.0a 262.8 ± 0.1a 113.6 ± 0.0c 179.3 ± 0.7a
Traditional (lime) 276.5 ± 2.1a 5.4 ± 0.2b 7.4 ± 0.1c 241.2 ± 0.1b 112.5 ± 0.2d 153.1 ± 0.4b
Ecological (CaCO3) 186.9 ± 1.8b 5.6 ± 0.1a 12.0 ± 0.2b 196.2 ± 0.0d 118.0 ± 0.0a 138.9 ± 0.1c
Extrusion (only water) 22.0 ± 0.2d 4.0 ± 0.3c 2.2 ± 0.1d 222.4 ± 0.3c 111.7 ± 0.0b 70.8 ± 0.5d
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Means ± SD (n = 2) followed by the same letter in the same column are not significantly different at p < 0.05

Relations of color, texture, and blister with the concentration of acrylamide in tortilla chip of different nixtamalization processes

Table 1 shows the color of tortilla chips from CNP, ENP, TNP and EXP, where color for tortilla chips for CNP and TNP was similar with a value of ΔE 37.5–36.4, respectively, but were different from tortilla chips from ENP with ΔE 39.8 and tortilla chips from EXP with ΔE 41.7. The highest change in color corresponded to tortilla chips prepared from EXP, the differences in color of tortilla chips from the above mentioned processes, may be due to the fact that, during deep-fat frying, the browning reaction takes place when amino acids and sugars are available for the Maillard reaction and this promotes the production of different substances that induce distinct taste and the darkness. Figure 2, shows tortilla chip images that were used to make different physicochemical tests. In the present research there was a highly significant correlation (r = 0.85; p < 0.0005), between tortilla chip color and acrylamide (Table 3). These results confirm the findings of different researchers who reported that color due to browning has a relationship with acrylamide content (Gökmen and Senyuva, 2006; Pedrechi et al., 2005; Ramonaitytè et al., 2009; Taubert et al., 2004).

Fig. 2.

Fig. 2

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Tortilla chips from different nixtamalization processes: (A) Traditional, (B) classic, (C) ecological and (D) extrusion

Table 3.

Correlation coefficients among the mineral content in tortilla chips, acrylamide and color

Parameter Ca2+ Fe2+ Zn2+ K+ Mg2+ Na+
Acrylamide − 0.87** − 0.84** − 0.47ns − 0.60* 0.79* − 0.95**
Color − 0.84** − 0.67* − 0.22ns − 0.63* 0.88** − 0.82**
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*, **Significant and highly significant at p ≤ 0.05 and ≤ 0.001, respectively

ns not significant

Regarding other quality parameters such as texture (fracture force (N)) in tortilla chips from distinct nixtamalization processes, there was a correlation (r = 0.90; p < 0.0001) with acrylamide where tortilla chips (EXT) with high level of acrylamide and poor quality presented higher texture with 7.9 N, followed by tortilla chips (CNP), tortilla chips (TNP) and tortilla chips (ENP) with 4.1 N, 4.7 N and 3.9 N, respectively. With respect to the blister and oil in tortilla chips, there were negative relationships of acrylamide with correlation coefficients of r = − 0.82; p < 0.0001 and r = − 0.85; p < 0.0004, respectively. This means that tortilla chips quality correlated negatively with acrylamide content and the effect seems to depend on the nixtamalization process.

Effect of nixtamalization process on acrylamide formation of tortilla chips

The levels of acrylamide from different nixtamalization processes are shown in Table 1.

Tortilla chips from CNP that uses ashes, showed the lower concentration in the formation of acrylamide during the frying step with 46.3 µg/kg, followed by tortilla chips of TNP that uses lime [Ca(OH)2] and tortilla chips from ENP that uses calcium carbonate with 55.0 µg/kg and 694.6 µg/kg, respectively. Some researchers have indicated that di- and mono-valent cations reduce acrylamide formation during frying of snacks (Elder et al., 2005; Gökmen and Senyuva, 2007b; Lindsay and Jang, 2005). Wu et al. (2016) reported that at high concentration of cations (Ca2+, Mg2+, Cu2+, Fe2+, Zn2+, Na+ and K+) similar to that found in tortilla chips of CNP (Table 2), decreases the decomposition rate of reducing sugars; these cations inhibit the degradation of glucosamine into methylglyoxal, that is a precursor from the acrylamide. The increase of Fe2+, Zn2+, K+, Mg2+ and Na+ in tortilla chips from TNP and ENP compared to tortilla chips from EXT was due to those reagents. Lime [Ca(OH)2] and calcium carbonate (CaCO3) used were food grade and they have traces of these minerals (Table 2). Similar trends in the mineral profile of tortillas was reported by Mariscal-Moreno et al. (2015) who studied the mineral content of raw material used in nixtamalization processes, and reported that the wood ash had di- and mono- valent cations, and lime of TNP, presented twice of cations of calcium (Ca2+) when compared with the CNP, having low presence of di-(Mg2+; Zn2+, Fe2+, Fe3+), and mono-valent (K+, Na+) cations. Lindsay and Jang, (2005) reported that treatment of potatoes slices with Fe3+ ions reduce acrylamide formation by 70% and Ca2+ ions reduced acrylamide formation by 59%, therefore, mono and divalent cations present in the different nixtamalization processes showed a positive effect in the mitigation of acrylamide formation in respect to the extrusion process, which had higher acrylamide values (two to three fold). Regarding acrylamide levels obtained from different nixtamalization processes, in the present study, the values were similar to those found in the literature. Friedman (2003) reported 38 to 416 µg/kg acrylamide in corn chips. Salazar et al. (2014) reported from 800 to 900 µg/kg acrylamide. Delgado et al. (2014) reported from 688 to 741 µg/kg acrylamide. Salazar et al. (2012) reported from 1141 to 1333 µg/kg acrylamide in tortilla chips. The nixtamalization processes that were used as raw materials—lime (traditional nixtamalization), calcium salts (ecological nixtamalization) and ashes (classic nixtamalization)—during corn cooking promoted different levels of acrylamide in the deep fat frying at 180 °C of tortilla chips depending on the kind of ions in the raw material, used for the nixtamalization.

The lower acrylamide content in tortilla chips is for classical nixtamalization process due to the positive effect of mono- and di-valent cations (Ca2+, Mg2+, Fe2+, Zn2+, Na+ and K+) present in the ashes used in the process. This nixtamalization process could be effective for an acrylamide mitigation strategy, because it represents a simple and inexpensive way to reduce acrylamide content in tortilla chips. The traditional nixtamalization (with lime) and ecological nixtamalization (Ca2+ salts) also promoted significant reduction of acrylamide in tortilla chips due the effect of the calcium (Ca2+) cations that inhibit the Maillard reaction precursors of acrylamide formation as indicated by the tortilla chip color.

It can be concluded that using different mineral sources during nixtamalization (Classic, Traditional and Ecological) could be an effective strategy in the decreasing of acrylamide in tortilla chips. Classic nixtamalization was the most effective method for acrylamide mitigation in tortilla chips due to the presence of mono- and di-valent cations (Ca2+, Mg2+, Fe2+, Zn2+, Na+ and K+), followed by the traditional and ecological nixtamalization processes.

In this study the specific cations present in tortilla chip were more important, than pH values and moisture content, it was confirmed by the presence of high-levels of acrylamide in tortilla chips from the extrusion process.

Acknowledgements

Alfonso Topete Betancourt thanks the CONACYT for the Ph.D. scholarship.

Footnotes

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Contributor Information

Alfonso Topete-Betancourt, Email: alfonso.topete@cinvestav.mx.

Juan de Dios Figueroa Cárdenas, Phone: +52-442-2119915, Email: jfigueroa@cinvestav.mx.

Adriana Lizbeth Rodríguez-Lino, Email: lizbeth2510@gmail.com.

Elvira Ríos-Leal, Email: erios@cinvestav.mx.

Eduardo Morales-Sánchez, Email: eduardo.morales.sanchez@yahoo.com.mx.

Héctor Eduardo Martínez-Flores, Email: hedu65@hotmail.com.

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