Patricia Isabel Ramos

Research Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn (Zea Mays L., IPB Var 6) Tea Patricia Isabel K. Ramos RND1, Diorella Marie M. Tria MSc1,2 1 and Clarissa B. Juanico RND, PhD With the increasing cases of non-communicable diseases and mineral deficiencies in the Philippines, people are shifting towards healthier food options. Consumption of products rich in antioxidants and minerals is one way to address the rising prevalence of these diseases. This study aimed to develop a tea beverage made from nixtamalized corn kernels (IPB Var 6) and determine its antioxidant activity (AA), calcium content and general acceptability. Results showed that there was no significant difference between the antioxidant activity of the tea made from nixtamalized (88.49 ± 0.25 AA%) and nonnixtamalized (87.31 ± 1.42 AA%) corn tea. However, there was a significant difference on the calcium content, wherein the nixtamalized corn tea exhibited higher content (263.33 ± 4.91 mg/100g). Further, there was no significant difference observed on the general sensorial acceptability of the products. Hence, the results suggest that nixtamalization process increased the calcium content of the corn tea without altering its functional property and acceptability. Keywords: nixtamalization; corn; tea; antioxidant activity; calcium; non-communicable diseases INTRODUCTION consumption of tea beverages even among coffeedrinking nations as consumers are shifting towards healthier food products and tea is one of the foods associated with wellness. Tea is one of the most widely consumed beverages in the world and its reported estimated revenue in the Philippines amounts to an equivalent of USD 79.4 million in year 2019. This beverage market is expected to grow annually by 6.7% (Statista, 2019). The country has a strong coffee culture but the growing appreciation and intake of tea by Filipinos through the years may be a good opportunity to explore the potential nutritional benefit of tea consumption and expound on its potential nutritional and functional values. Many varieties of tea using various plant parts or fruit prepare tea drinks by steeping these in boiling water and are more appropriately called herbal infusions or tisanes (Sanlier, et al., 2018). All teas and tisanes will be referred to as tea for the purpose of this study. One functional property of teas includes having antioxidants. The antioxidants in tea may prevent NCDs because of its action on oxidative stress which is why drinking tea is beneficial. More than half of annual deaths in the Philippines are caused by non-communicable diseases (NCDs), accounting for an estimated 67% of the reported cases on cardiovascular diseases, cancers, respiratory diseases, diabetes, chronic kidney diseases, and others (WHO, 2018). NCDs have many modifiable risk factors, and interventions are needed to alleviate such diseases. Diet and obesity are two of the important determinants of NCDs (Hyseni et al., 2017) and these contribute more to the global burden of malnutrition than tobacco, alcohol and physical inactivity combined. Among the important dietary changes in children and adults that are associated with obesity as a risk to NCD is the consumption of unhealthy beverages especially those high in sugar content. However, the extent to which the beverage intake can contribute to obesity in Filipinos is unknown (Gubat et al., 2015). More people are now interested on the market of natural beverages that may offer promising effects to health improvement. One example is the increasing 1 2 Institute of Human Nutrition and Food, College of Human Ecology University of the Philippines Los Baños, College, Laguna, Philippines Science Education Institute, Department of Science and Technology, Bicutan, Taguig, Philippines Corresponding Author: Patricia Isabel K. Ramos /- JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 | 1 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn In this research, nixtamalized corn (Zea mays L., IPB Var 6) was used in developing tea. Since corn and corn products were underutilized in the country despite its affordability, high production, and its recognized significant nutritional value from researches (Salazar et al., 2016), some of the corn production go to waste. Further, corn consumption is limited to areas (e.g. the Visayas) where it is a staple food and part of the dietary pattern. For the rest of the country, corn is generally prepared as a snack food. This may be due to lack of awareness on the nutritional benefits of corn as well as limited knowledge on preparation of corn products (Cinco, 2016). Nixtamalization, a process commonly employed for corn, will be used in this study for its recognized effects on enhancing the nutritional properties of corn, most especially on the calcium content (Valderrama-Bravo et al., 2010). The results of 8th National Nutrition Survey (NNS) by Food and Nutrition Research Institute (FNRI) showed that the proportion of household meeting the estimated average requirement for the various nutrients ranges from 25-35%, except calcium (15%) and iron (9%). Department of Science and Technology (DOST) recommends calcium-rich diets for Filipinos. The same survey results also reported the alarming increase in overweight and obesity which are considered risk factors of NCD cases in the country (FNRI-DOST, 2016). There is very limited data, to date, on nixtamalized corn-based beverages, specifically tea. This study aimed to determine the antioxidant activity, calcium content and general acceptability of nixtamalized tea made from corn (Zea Mays, L., IPB Var 6). This study also intends to promote corn consumption as a beverage for its nutritional value and consequently encourage increased production and development of corn products by the local food industry, particularly its potential in tea manufacturing. METHODS Corn Nixtamalization IPB-bred corn kernels (Var 6) used in this study were obtained from the Institute of Plant Breeding (IPB) in Los Baños, Laguna, Philippines. The kernels were sundried to 14% moisture prior to tea processing. Calcium Carbonate used for nixtamalization was obtained from the Institute of Chemistry (IC), in University of the Philippines Los Baños, Laguna (UPLB). The nixtamalization process starts with cooking the kernels with alkaline solution at 90 °C for 30 minutes with 1% (w/w) calcium carbonate (CaCO3) salt. A rice cooker was used to nixtamalize the corn kernels to maintain the constant temperature. The ratio of corn to water is 1:2. Distilled water was used to prevent addition of other minerals that may be present in hard water. The cooked kernels were steeped for 16 hours at room temperature (Santiago-Ramos et al., 2018; RuizGutierrez et al., 2011; Rodriguez-Mendez, 2013). Figure 1. Tea processing of corn kernels (Zea Mays, L., IPB Var 6) used in the study: (1.1) after dehydration [Left – nixtamalized; Right – non-nixtamalized], (1.2) after roasting [Left – nixtamalized; Right – non-nixtamalized], (1.3) after grinding [Left – nixtamalized; Right – non-nixtamalized], and (1.4) after steeping – final product 92 | JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn Processing of Corn Tea Beverage Sensory Evaluation The nixtamalized corn kernels as well as the washed non-nixtamalized corn kernels were dehydrated at 45 °C for 24 hours. The dehydrated, nixtamalized and non-nixtamalized corn kernels were both oven-roasted at 150 °C for 180 minutes (Xiong et al., 2018). After roasting, the samples were ground using mortar and pestle, and were packaged in individual teabags at 10 grams per teabag. The samples were subjected to chemical analysis. For sensory evaluation, the teabags were steeped in freshly boiled water at 90-95 °C for 4-5 minutes. One teabag was steeped in 150 mL of water. In evaluating the general acceptability of the nixtamalized and non-nixtamalized corn tea, a 9point hedonic scale was used. Ten laboratory panelists were selected based on their knowledge and experience on conducting sensory evaluation of foods to evaluate the quality attributes of the corn tea. The two samples were served simultaneously to each panelist at room temperature (25 °C). Panelists were instructed to rinse their mouths with distilled water prior to tasting the tea samples to cleanse their palate (Watts et al., 1989). The overall liking, appearance liking, aroma liking, taste liking, and aftertaste liking were assessed. Participants' confidentiality of research information was assured in this study. All participants described in this research gave their written informed consent about their willingness to participate. They were also assured about confidentiality and anonymity regarding their participation in the study. Antioxidant Activity Analysis The antioxidant activity (AA) of nixtamalized and non-nixtamalized corn were determined using 1diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay (Zhang, 2017) in triplicates. One mL of aliquot was placed in a test tube and diluted with 4 mL distilled water. One mL of newly prepared 1mM DPPH methanolic solution was also added. The solution was left in the dark for 20 minutes. The absorbance was read at 517nm against a blank. Lower absorbance at 517 nm represents the higher DPPH scavenging activity. The % DPPH scavenging activity (a) and percent difference (b) were calculated using the formulas below: DPPH scavenging activity (%) = Difference (%) = Absorbance of control Absorbance of sample x Absorbance of control 100 (a) mean AA% nixtamalized mean AA% nonnixtamalized x 100 (mean AA% nixtamalized+mean AA% nonnixtamalized) ÷2 (b) Statistical Analysis Descriptive statistics was generated for nixtamalized and non-nixtamalized corn tea for the antioxidant activity, calcium content and general acceptability. Independent t-test was used in comparing the antioxidant activity and calcium content of the two samples wherein assumptions for equal variances was tested. The general acceptability of the two tea samples was compared using Wilcoxon Signed Rank Test. Values were expressed as mean ± standard deviation and differences were measured at 5% level of significance (p<0.05). Calcium Content Analysis RESULTS AND DISCUSSION Calcium content (CC) (mg/100g) analysis was done in triplicates for both samples. Wet digestion using furnace was employed instead of ashing since the samples were in liquid form (Purificacion et al., 2018). The samples were digested using 1:3 Nitric acid-Hydrochloric acid. The calcium content was measured using Flame Atomic Absorption Spectroscopy (FAAS). Percent difference (c) was calculated using the formula below: Difference (%) = mean CC nixtamalized mean CC nonnixtamalized (mean CC nixtamalized + mean CC nonnixtamalized) ÷2 x 100 Nixtamalization has been incorporated as a primary step in producing several corn-based products to soften the pericarp and endosperm for ease of processing (Mora-Rochin, 2010). Studies show that nixtamalization can change structural (Santiago-Ramos, 2018; Ruiz-Gutierrez, 2011; Gomez et al., 1989) and enhance proximate and functional (Atienza et al., 2019) properties of corn. The nixtamalization process is also acknowledged to have significant effects on some of the nutritional JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 | 3 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn properties of corn (Rosales et al., 2016; Morales & Zepeda, 2017), however limited data is available for its effect when developed into tea beverage. The study conducted by Vernarelli and Lambert (2013) concluded that hot tea consumption was inversely associated with obesity. Their findings supported the potential of hot tea consumption as a regulator of body weight and total body fat. The decrease in BMI, waist circumferences, and inflammatory markers promotes lesser risk to the development of NCDs. Another study about tea consumption had also concluded a significant negative association with the prevalence of central obesity and cholesterol level. Their findings highlighted the benefits of polyphenols found in tea as a health-promoting habit to lessen the risk to chronic diseases (Grosso et al., 2015). This study aimed to determine the antioxidant capacity, calcium content and general acceptability of tea made from corn to determine the efficiency of nixtamalization process for the development of a potential health-promoting corn-beverage product. Results were compared to the non-nixtamalized corn tea samples using the same variety. (p-value: <0.0000) from each other at 5% level of significance. The percent difference of the calcium content levels of the samples was also computed (Table 1). From the analysis, it can be concluded that the nixtamalization process significantly increased the calcium content of the corn tea. However, it should be noted that the nixtamalization process used in this study did not include washing of the corn kernels after steeping, but were directly dried, as opposed to the common nixtamalization methods from other studies. Data results of the components of raw, dry IPB Var 6 corn (Table 2) for comparison and reference, however, were adapted from the study of Purificacion et al. (2018) which is a limitation of this study. Over the last decades, researches about exogenous antioxidants, the type of antioxidants people get from their diet, have been increasing. The interest on its potential benefits toward human health has drawn great attention especially since various literatures had successfully linked dietary antioxidants to the prevention of some of the leading causes of mortality such as non-communicable and chronic diseases (Grosso, 2018; Frei, 2004; Blomhoff, 2005). The result of the DPPH scavenging capacity of the processed tea samples showed insignificant difference from each other however, antioxidant levels were higher when compared from raw dry corn data (Table 2) of the same variety which was adapted from the study made by Purificacion et al. (2018). The increase in the antioxidant activity between the raw and the processed corn can be attributed to roasting the kernels at 150 ºC since thermal processing increases the antioxidant activity (Liu et al., 2002) of food products. Thermal processing promotes a progressive polymerization of phenolic compounds to form brown-coloured macromolecular products that may lead to the formation of stable intermediates Antioxidant Activity and Calcium Content DPPH assay was used to measure the radical scavenging capacity of the tea samples. Results showed that the mean DPPH scavenging activity of the nixtamalized corn tea is slightly higher than the non-nixtamalized tea however, there was no significant difference (p-value: 0.2275) at 5% level of significance. The computed percent difference of the AA% levels of the samples is presented on Table 1. The calcium content of the samples was determined in which showed that the mean calcium content of the nixtamalized corn tea was evidently higher than the non-nixtamalized tea. Additionally, the amount of calcium in the tea samples were significantly different Table 1. Antioxidant Activity (DPPH %) and Calcium Content (mg/100g) of the nixtamalized and non-nixtamalized corn tea beverage Analysis DPPH, % Calcium, mg 100 g-1 Nixtamalized Corn Tea 1 88.49 ± 0.25a 263.33 ± 4.91 b Non-Nixtamalized Corn Tea 1 87.31 ± 1.42a c 46.50 ± 1.72 1 Difference 1.34% 139.97% Values are represented as mean ± SD of triplicate measurements. Means with the same letters per analysis are not significantly different (p<0.05). 04 | JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn Table 2. Proximate, amino acid, starch, minerals and phytochemical contents of raw, dry IPB Var 6 corn (adapted from Purificacion et al., 2018 as partial data based on tabulated results of components for different IPB-bred varieties: IPB Var 6, 8, 9, 11 and 13) Component Proximate Composition 1 Moisture, % Ash, % Crude Fat, % Crude Protein, % Crude Fiber, % Total Carbohydrates, % Amino Acid Content2 Lysine, % Tryptophan, % Starch Profile 1 Total Starch, % Amylose, % Amylopectin, % Mineral content1 Iron, mg 100 g-1 Zinc, mg 100 g-1 Potassium, mg 100 g-1 Calcium, mg 100 g-1 Magnesium, mg 100 g-1 Phytochemical content1 DPPH, % TPCa, g GAE 100 g-1 TFCb, mg CE 100 g-1 TCCc, mg 100 g-1 Value 5.52 ± 0.04 1.56 ± 0.25 4.66 ±- ± 0.29 4.63 ±- ± 0.69 0.67 ± 0.04 0.06 ±- ±- ±- ± 0.95 3.72 ± 0.01 2.37 ±- ±- ±- ±- ± 2.71 4.18 ± 0.91 5.29 ± 0.72 0.73 ± 0.21 1 Values are represented as mean of dry corn ± SD of triplicate measurements. Values are represented as mean of dry corn ± SD of duplicate measurements. a TPC=Total Phenolic Content b TFC=Total Flavonoid Content c TCC=Total Carotenoids Content 2 which can exhibit strong antioxidant activity (Manzoco et al., 1998). Several studies (Oboh et al., 2010; Woo et al., 2018) reported similar findings wherein the antioxidant capacity of the corn significantly increased with higher roasting temperature and longer time. The reduction of natural antioxidants in foods are generally expected after thermal processing, however, some heatinduced processes can improve its functional properties (Al-juhaimi et al., 2018). The result of the antioxidant levels of the tea samples showed that the nixtamalization process in this study does not alter the antioxidant activity of the corn. The study by Purificacion et al. (2018) observed that the calcium content of IPB-bred varieties ranged from- mg 100 g-1. The same study also showed (Table 2) the calcium content of raw and dry IPB Var 6 corn. As seen in Table 1, the calcium content of the non-nixtamalized corn sample was lower than that of the raw corn (see Table 2). The loss of calcium content from the non-nixtamalized corn sample can be attributed to the cooking process that the raw corn had undergone (e.g. dehydration and roasting) (Prasanthi, 2017). The latter result is similar with previous studies (Santiago-Ramos et al., 2018; Atienza et al., 2019; Bressani et al., 2002; Carrera et al., 2012) wherein the calcium content of nixtamalized corn product was observed to significantly increase. This can be attributed to: (a) cooking and steeping time of nixtamalization and (b) physical state of the kernels (Morales & Zepeda, JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 | 5 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn 2017; Fernandez-Muñoz et al., 2004). Cooking of corn using the alkaline solution softens the pericarp due to hydration which then causes the protein denaturation. The denaturation affects the partial gelatinization of starch which leads to the occurrence of the interaction of starch with calcium (SantiagoRamos et al., 2018). During cooking, the kernels boiled in the alkaline solution results in removal of starch granules altering the inner endosperm (Bressani et al., 2002). It should be noted that the nixtamalization process used in this study did not include washing of the corn kernels after steeping since the removal of the pericarp was not necessary for tea processing, thus kernels were subjected directly to drying. Thus, this also contributed to the higher calcium content of the corn tea. The diffusion of calcium in corn kernels was observed to be fastest on its outer most component, pericarp, followed by germ and then the endosperm (Morales & Zepeda, 2017; Fernandez-Muñoz et al., 2004). A study about the mechanism of calcium uptake in corn kernels during nixtamalization showed that the calcium absorption in the pericarp occurred for short steeping times (Valderrama-Bravo et al., 2010). Longer hours of steeping means calcium ions can be more diffused through the inner most parts of the kernels for a higher increase in calcium content. Nixtamalized Corn Tea General Acceptability The general acceptability of the tea samples was assessed based on the degree of likeness of its appearance, aroma, flavour, aftertaste and overall liking. The 10 panelists evaluated the sensory attributes of the corn tea and were asked to assess the overall acceptability of the beverage. Figure 2 shows the difference between the nixtamalized and non-nixtamalized corn tea based on the intensities of the previously mentioned attributes. The compared samples were not significantly different in its appearance (p-value: 0.8717), aroma (p-value: 0.5212), flavour (p-value: 0.8686) and aftertaste (pvalue: 0.7503). The attribute of appearance was found to have the highest accumulated acceptance score while aftertaste was the least. Statistical analysis also showed that there was no significant difference between the overall liking (p-value: 0.5324) of the respondents for the nixtamalized and non-nixtamalized corn tea at 5% level of significance. The result of the sensory assessment between the nixtamalized and non-nixtamalized corn tea beverages showed no distinction regarding the various attributes and overall rating. The appearance of the tea, which is the highest scoring attribute, is Non-nixtamalized Corn Tea Overall Liking- Aftertaste Flavour Appearance Aroma Figure 2. Plot comparing the intensities of the different sensory attributes of the nixtamalized and non-nixtamalized corn tea beverage (quality scoring). 6 | JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn highly related to the colour. A study about the consumer acceptability of teas had concluded that the “yellowness”, as a characteristic of the traditional tea, is one of the drivers for consumers to like the tea products (Lee et al., 2009). Traditional teas usually obtain the colour depending on the degree of oxidation. A longer oxidation process produces more pigments which produce the yellow to black colour of tea (Sai et al., 2011). However, the tea beverage in this study that was subjected to oven-roasting did not undergo the same process as any traditional tea. Thus, the colour of the tea may be attributed to the presence of sugars, amino acids, and other organic acids in the corn kernels. The condensation of amino groups on protein, peptides and amino acids creates Maillard reaction. The reaction can lead to compositional and structural changes to the sugars, amino acids and other organic acids of the corn tea and thus, have a potential implication on food color (Lund & Ray, 2017). The low score accumulated from the aftertaste of the tea samples can be explained through the hint of bitterness and astringency which are reasonably present in particular beverages such as tea. High intensity of bitter taste and astringency is perceived to be the driver for the potential consumers to dislike most of the tea products (Lee et al., 2009). Some studies which focused on chemical composition and sensory evaluation of tea stated that the bitterness and astringency of the tea were correlated with the contents of flavonols (catechins) and other phenolic compounds (Adnan et al., 2013; Xu et al., 2018). The IPB-bred corn (IPB Var 6) used in this study evidently contains phenolic compounds, flavonoids and amino acids (Table 2). Besides catechins and caffeine, some amino acids such as alanine or arginine also contribute to the bitterness of tea infusion (Sai et al., 2011). The corn tea made in the study is naturally caffeine-free thus, flavonols, phenolic content and amino acids from the corn (IPB Var 6) which were subjected to heat might have contributed to the bitterness and astringency directly affecting its aftertaste. CONCLUSION Nixtamalization process can increase the amount of some functional and nutrient content of corn and corn products as based on previous studies. Hence, this study utilized nixtamalized IPB var 6 in developing corn tea. The corn tea samples were analyzed for its antioxidant activity and nutrient component. The DPHH results showed no significant increase in the antioxidant activity of the corn tea with nixtamalization, however, the FAAS result showed a significant increase in the calcium content. And the general sensorial acceptability of the nixtamalized corn showed that the overall liking rate of the tea samples have no significant difference. The similarities of the tea samples in terms of their sensory attributes suggested that the nixtamalization process does not make any distinctive changes on the attributes of a traditional or regular corn tea thus, may increase the chance of consumer acceptance especially by those who are already tea drinkers. Corn-based beverage in the form of tea is a strategic way of increasing the utilization of corn nationwide in view of its benefits and lower raw material cost for the food industry. It can be concluded that the nixtamalization process can enhance the potential functional property from the corn tea consumption without altering overall acceptability. ACKNOWLEDGMENTS The authors would like to acknowledge the Bureau of Agricultural Research of the Department of Agriculture (DA-BAR) for funding this study. The authors would also like to express their gratitude to the Institute of Human Nutrition and Food, University of the Philippines Los Baños (IHNF-UPLB) for providing administrative support and the facilities for the experiment and sensory analysis. JNDAP January-December 2020 Vol. 34 Nos. 1 & 2 | 7 Ramos et. al. / Antioxidant Activity, Calcium Content and General Acceptability of Nixtamalized Corn REFERENCES 1. 2. 3. 4. 5. 6. Adnan M, Ahmad A, Ahmed A, Khalid N, Hayat I, and Ahmed I. 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