Natural pigment extraction optimization from coffee exocarp and its use as a natural dye in French meringue

Accepted Manuscript Natural pigment extraction optimization from coffee exocarp and its use as a natural dye in French meringue Amanda Parra-Campos, Luis Eduardo Ordó ñez-Santos PII: DOI: Reference:

S0308-8146(19)30233-X https://doi.org/10.1016/j.foodchem.2019.01.158 FOCH 24253

To appear in:

Food Chemistry

Received Date: Revised Date: Accepted Date:

3 November 2018 21 January 2019 22 January 2019

Please cite this article as: Parra-Campos, A., Ordó ñez-Santos, L.E., Natural pigment extraction optimization from coffee exocarp and its use as a natural dye in French meringue, Food Chemistry (2019), doi: https://doi.org/10.1016/ j.foodchem.2019.01.158

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Natural pigment extraction optimization from coffee exocarp and its use as a natural dye in French meringue Amanda Parra-Campos and Luis Eduardo Ordóñez-Santos* Universidad Nacional de Colombia-Sede Palmira, Facultad de Ingeniería y Administración, Departamento de Ingeniería, Carrera 32 N 12-00, Palmira, Valle del Cauca, Colombia *Corresponding author. Tel.: +57 2 2868888. E-mail address: [email protected] (L.E. Ordóñez-Santos)

Abstract This study aimed to optimize the pigment extraction process for coffee exocarp and to evaluate its coloring effect on French meringue. The anthocyanins were determined with the differential pH method and the process was optimized with the response surface methodology. The solvent concentration (SC) and solid solvent ratio (SSR) had a significant effect on the anthocyanin content and surface color of the coffee cherry extracts. The optimal extraction conditions 60% SC and 25% SSR resulted in the highest concentration of anthocyanins (0.145 mg cyanidin 3-glucoside /g of coffee fresh exocarp). For the French meringue, the 3% extract concentration had the smallest total color difference (ΔE), as compared to the control, evidencing the potential of coffee exocarp dyes in place of synthetic dyes in the manufacture of French meringue. Keywords: anthocyanin; color; pH; soluble solids; response surface methodology. 1. Introduction Coffee belongs to the genus Coffea (Rubiaceae family), and its trade ranks second, after oil, in terms of dollars worldwide because the beverage obtained from coffee seeds is very popular and widely consumed (Naidu, Sulochanamma, Sampathu, & Srinivas, 2008; Esquivel & Jiménez, 2012; Heeger, Kosińska-Cagnazzo, Cantergiani, & Andlauer, 2017). According to the International Coffee Organization, production in Colombia in 2017 was 14 million 64 kg bags, with Colombia being the second largest producer in the Americas (ICO, 2018). During the processing of coffee, large quantities of by-products are generated, and exocarp accounts for 29% of the dry weight of the fruits (Murthy & Madhava Naidu, 2012, Bonilla-Hermosa, Duarte, & Schwan, 2014). The exocarp comprises the outer skin and the pulp and is rich in bioactive components such as flavonoids and anthocyanins, which are responsible for the red color seen in coffee fruits (Murthy et al., 2012, Bonilla-Hermosa 1

et al., 2014). The coffee exocarp has limited applications (Murthy et al., 2012; Heeger et al., 2017) and is usually discarded without proper treatment (Murthy et al., 2012; Bonilla-Hermosa et al., 2014). Cyanidin 3-rutinoside and cyanidin 3-glucoside have been reported as the principal anthocyanins found in coffee (Prata and Oliveira, 2007, Murthy et al., 2012), and coffee exocarp has been confirmed as a source of anthocyanins, providing an opportunity for a dye and bioactive ingredient in formulated foods (Murthy et al., 2012). Natural red dyes are scarce and highly sought after (Hurtado and Pérez, 2014). Meringue is a very popular confectionery product, made with egg white and sugar, and is the basis for the preparation of other products such as soufflés, macaroons, and angel food cake (O'Charoen, Hayakawa, Matsumoto, & Ogawa, 2014). The color in this product is an important attribute of quality valued by consumers, during its manufacture, some synthetic dyes such as Ponceau 4R, Sunset Yellow and Tartrazine are used to make them more attractive to the consumer (Pasias, Asimakopoulos, & Thomaidis, 2015). Several studies have confirmed the feasibility of using anthocyanins as an alternative to synthetic dyes, which can have negative effects on health (Kirca, Özkan & Cemeroǧlu, 2006; Aguilera-Ortíz et al., 2012; Buchweitz, Brauch, Carle, & Kammerer 2013; Assous et al., 2014; Pasqualone, Bianco, Paradiso, Summo, Gambacorta, & Caponio 2014; Adsare, Bellary, Sowbhagya, Baskaran, Prakash, & Rastogi, 2016; Gerardi et al., 2018). It has been reported that ingesting Ponceau Red 4R (E 124) at levels above 0.7 mg/kg of body weight per day can cause cognitive deficiencies (Assous et al., 2014; Amchova, Kotolova and Ruda-Kucerova , 2015). Interest in anthocyanins comes not only from their coloring effect, but also from their beneficial health properties, which result from a high antioxidant activity (Assous, Abdel-Hady and Medany, 2014). Anthocyanins have bright attractive colors and are soluble in water, facilitating their incorporation into aqueous food systems (Aguilera-Ortíz, Reza-Vargas, Madinaveitia, Valenzuela, & Baca, 2012). The growing interest in the potential of natural pigments has created a demand for efficient extraction processes (Ongkowijoyo, Luna-Vital and Gonzalez de Mejia, 2018). The response surface methodology (RSM) is a statistical tool that has been used successfully in process parameter testing, along with its interactive effects. RSM helps design mathematical models and determine optimal operating conditions (Swamy, Sangamithra, & Chandrasekar, 2014; Pedro, Granato, & Rosso, 2016).

2

With the high production of coffee, there is a need to create suitable uses for by-products, obtaining products with high added value (Bonilla-Hermosa et al., 2014). On the other hand, the use of natural colors is currently a trend because of the concern of consumers over the safety of artificial dyes (Rodriguez-Amaya, 2016). However, no literature was found that has improved the process for obtaining coffee cherry pigments or their application in food, so this study aimed to optimize the extraction process for coffee exocarp pigments and evaluate their coloring effect on French meringue. 2. Materials and methods 2.1. Materials This study used coffee cherry variety Colombia, in maturation stage III (Ramos, Sanz and Oliveros 2010), cultivated in the Municipality of Timbio Cauca at 1850 m.a.s.l, with an average temperature of 20 ° C. From a crop of 500 coffee trees, 10 trees were selected at random, and 2.5 kg of fruit were manually collected. The fruits were disinfected with a 50 ppm solution of sodium hypochlorite for 10 minutes and manually pulped to eliminate the seed and mucilage. 2.2. Determination of the physicochemical properties of the coffee exocarp The pH (Colombian Technical Standard, NTC 4592), titratable acidity (NTC 4623), soluble solids (NTC 4624) and moisture content (Darıcı and Şen, 2015) were determined. The surface color was determined using a Minolta CR-400 Colorimeter with the CIEL*a*b* coordinates; illuminant D65, 2° observer and calibration parameters Y = 89.5; x = 0.3176; and y = 0.3347. The saturation (C*), hue (h°) and total color difference (ΔE) were estimated (Salinas-Moreno, Almaguer-Vargas, Peña-Varela, & Ríos-Sánchez, 2009; Kara and Ercȩlebi, 2013). 2.3. Extraction and quantification of anthocyanins present in the coffee exocarp The exocarp of fresh coffee was mixed with 20 ml of an acidified ethanol solution at pH = 1, and refrigerated (4 ° C) for 18 hours (Murthy et al., 2012). Ethanol percentage (36-64% v/v) and amount of exocarp (15-85% w/v) were selected according to the central composite design presented in Table 1. After extraction, the samples were filtered through a filter paper (Whatman No. 1). From the filtering, 2 dilutions were prepared separately with a 10 dilution factor using a potassium chloride buffer solution pH= 1 (KCl, 0.025 M) and sodium acetate pH= 4.5 (CH3COONa, 0.4 M), which were refrigerated at 4°C for 20 minutes (Mercali, Jaeschke, Tessaro, & Marczak 2013; Liu et al., 2014; Mohideen et al., 2015). The absorbance was measured to 33 samples at a wavelength of 3

510 and 700 nm using a spectrophotometer Jenway 6320D (Cole-Parmer Ltd, Dunmow, England), and the pigment was quantified with the following equation (1): Total anthocyanin (mg / g) = (A x Mw x DF x 1000) / (ε x 1) Where, A= (A510-A700)

pH=1.0-

(A510-A700)

pH=4,5,

(1)

Mw = (molecular weight) = 449.2 g/mol for the cyanidin 3-

glucoside, DF = dilution factor (20 ml ethanol / M) * (10 ml buffer solution/1 ml of extract) * 1L / 1000 ml) M = g of coffee fresh exocarp used, l = length of the cuvette in cm, and ε = 26900, molar extinction coefficient L/mol cm for cyanidin 3-glucoside and the conversion coefficient of g to mg was 1000. 2.4. Application of the coffee exocarp pigment to French meringue The extract obtained under the optimal extraction conditions was concentrated at an anthocyanin content of 1.18 ± 0.07 mg cyanidin 3-glucoside/g of coffee fresh exocarp, in a water bath at 60 ° C (Rosero, Corral, Álvarez, Serna, & Ordoñez 2016). The French meringue was made following the methodology of Goldfarb (2016), with the ingredients: 250 g of egg white, 250 g of normal and powdered sugar, 25 g of corn starch and 0.1 ml of synthetic colorant (Ponceau 4R, E124). The egg whites were beaten to the point of snow in an electric mixer (Universal, Medellín, Colombia), incorporating the sugar slowly and continued beating for 5 more minutes, then corn starch was added, and finally the synthetic colorant, was taken to the mold and baked for 20 min at a temperature of 150 ° C in a gas oven (Zingal, Bogotá, Colombia). The color of the meringue was measured (Standard) and several concentrations were evaluated, replacing the synthetic colorant with the natural one (0, 3, 6, and 9 g/100 g of fresh dough). The process was done in triplicate. Extraction of anthocyanin present in the French meringue was determined according to Rodrigues, Fernandes, de Brito, Sousa, & Narain (2015). 0.5 g of the meringue sample was mixed with 10 ml of 46% (v / v) ethanol acidified to pH = 1, (ratio 1:20). The solution is brought to extraction with the aid of an ultrasound (25 kHz, 150 W, Branson Ultrasonics Corp. Danbury, USA) for 10 minutes at a temperature of 30 ° C and subsequently brought to centrifugation at 4000 rpm for 15 min. The quantification of the pigment in the meringue was carried out as previously described in the present study. 2.5. Statistical analysis The effect of solvent concentration (SC) and the proportion of solid solvent (SSR) on total anthocyanin content and surface color was evaluated using the response surface methodology (RSM) with a rotating central 4

composite design (RCCD), with 3 centrals, 4 axials and 4 factorial experiments were used to obtain the best combination of process variables that optimizes the extraction, and the experiments were carried out in triplicate (Table 1). The central points were established based on previous studies (Prata & Oliveira, 2007, and Murthy et al., 2012). The analysis of the data was carried out with Design Expert 11 using analysis of variance (ANOVA), regression and surface plot of responses. The model selection criteria included coefficient of determination (R2) and Lack or Fit, and the model validation was performed using a T-test between the theoretical and experiment values. The effect of the concentration of the pigment on the color changes in the product (ΔE) was evaluated with a simple randomized design; the pigment concentration (3, 6, 9% and standard) represented the evaluated treatments, with three repetitions per treatment. The analysis of the results was carried out with the Minitab statistical software (version 17) using analysis of variance (ANOVA) and multiple Tukey comparisons. 3. Results and Discussion 3.1. Physical chemical characterization of the coffee exocarp The coffee exocarp presented a pH of 4.65 ± 0.05, acidity of 0.03 ± 0.01% (malic acid), soluble solids of 11.4 ± 0.60% and 74.94 ± 2.94% moisture, values that agree with those reported by Marín-López, Arcila-Pulgarin, Montoya-Restrepo, & Oliveros-Tascón (2003) and Heeger et al. (2017). The anthocyanin content varied between 0.087 and 0.099 mg cyanidin 3-glucoside/g of coffee fresh exocarp, values that are close to those reported by Prata and Oliveira (2007). The color coordinate values (L* = 25.16 ± 0.42; a*= 5.22 ± 0.28, b* = 3.57 ± 0.12, C = 6.33 ± 0.12 and h° = 34.45 ± 2.05) were close to those reported by Carvajal-Herrera, Aristizábal-Torres, Oliveros-Tascón, & Mejía-Montoya (2011). 3.2. Effect of solvent concentration (SC) and solid solvent ratio (SSR) on the anthocyanin content and color coordinates CIEL*a*b* of the extract from coffee exocarp The anthocyanin content varied between 0.053 and 0.155 mg cyanidin 3-glucoside /g of coffee fresh exocarp (Table 1), values that are close to those reported by Prata and Oliveira (2007) and comparable with other sources of anthocyanins, such as those obtained from ornamental rice grains (Abdel-Aal, Young and Rabalski, 2006; Sompong, Siebenhandl-Ehn, Linsberger-Martin, & Berghofer, 2011) and potatoes (Lachman et al., 2009). Also, low variation was found in the color coordinates CIEL*a*b* of the extract from coffee exocarp; L* (19.67 -21.88), a* (5.59 - 6.61), b* (2.78 - 3.49), C* ( 5.93 - 7.28) and h* (25 – 31.80); the L* values were comparable to those 5

reported for other sources of anthocyanin pigments (Patil, Madhusudhan, Babu, & Raghavarao 2009; SalinasMoreno et al., 2009). According to the analysis of variance (Table 2), the 2FI model significantly explained (p <0.05) the anthocyanin

content found in the coffee exocarp extract, obtaining the following regression models: Anthocyanin content = 0.0967 + 0.0081SC - 0.0282 SSR - 0.0117 SC * SSR Where, the effect of the SC, SSR and the interaction between these two variables (SC * SSR) was significant, with a satisfactory fit to the model (lack or not significant fit) and an R2 that explained 95% of the influence of the studied factors on the variation in the anthocyanin content. Figure 1 shows the linear effect and correlation between the variables, obtaining the highest anthocyanin content with the highest SC (60% v/v) and the lowest SSR (25% w/v). The significant effect of the SSR can be attributed to changes in the concentration gradient of the solvent and the rate of diffusion of the compounds from solid to solvent (Cacace and Mazza, 2003), as a consequence of the variation in the content of the exocarp submerged in the solvent, with a lower SSR presenting a higher anthocyanin content. This behavior can also be due to an increase in the solubility of the anthocyanins as a consequence of an increase in the diffusion coefficient of the solvent in the solid matrix, favoring the desorption kinetics of the matrix compounds (Heras, Alvis and Arrazola, 2013). Similar results have been reported in other studies (Fan, Han, Gu, & Chen 2008, Celli, Ghanem and Brooks, 2015). On the other hand, it was demonstrated that a higher SC managed to extract a higher anthocyanin content, which may be attributable to the fact that the changes in the ethanol concentration modified the physical properties of the solvent, such as the density, dynamic viscosity and constant dielectric, affecting the solubility, which could influence the extraction of the compounds, and that the driving force within the particles increased as the concentration gradient of the solvent increased, causing the extraction rate to increase (Cacace and Mazza, 2003). The color coordinates L*, a* and C* satisfactorily fit to a quadratic model, with an R2 of 0.85, 0.95 and 0.99, respectively (Table 2). The SC significantly influenced the L* (clarity) and C* (intensity of color). The SSR and SC*SSR interaction significantly affected the coordinates a* (red) and C*, and the quadratic effect (CS 2 and 6

RSS2) showed significant influence on the behavior of the three color coordinates, obtaining the following regression models: L* = 20.12- 0.46SC - 0.08SSR + 0.38SC*SSR + 0.039SC2 + 0.039SSR2; a* = 6.48- 0.06SC- 0.19SSR- 0.18SC * SSR- 0.42SC2- 0.22SSR2 and C* = 7.26- 0.20SC- 0.11SSR- 0.30SC * SSR- 0.39 SC2 -0.36SSR2. The behavior of the color angle h° was affected by the SSR, adjusting to a simple linear model, with an acceptable R2 correlation of 0.61. h° = 28.51 - 0.69CS + 1.53RSS. On the other hand, the quadratic effect of the SSR was significant for b* (yellow), with an R2 of 0.78; however, the model was not significant, and the significant lack of fit meant it was not possible to predict the changes in color coordinate b* with the studied factors. Figure 2(A-D), shows the effect of the SC and SSR on the color coordinates CIEL*a*b* , where the highest SC and the lowest SSR yielded the lowest values of L* and h°, and the intermediate levels of these two variables produced a greater value of a* and C*; these results may have beer due to the fact that the equilibrium concentration in the liquid phase was reached (Cacace and Mazza, 2003). Color coordinate L* presented an inverse behavior with the anthocyanin content, indicating that the lower the L* value, the greater the presence of anthocyanins in the extract. Other studies have also reported a relationship between anthocyanin contents and color coordinates (Cevallos-Casals & Cisneros-Zevallos, 2004; Cao, Liu, & Pan, 2011; Kara et al., 2013; Su, Zhu, Xu, Ramaswamy, Lin, & Yu, 2016), including in other pigments, such as betalains (Güneşer, 2016). 3.3. Response optimization on the anthocyanin content Figure 3A shows that the extraction conditions 57.35% SC and 25.47% SSR yielded the highest anthocyanin content (0.1389 mg cyanidin 3-glucoside /g of coffee fresh exocarp), with a dark color (L* minimized), higher saturation (C maximized) and color tone (h° minimized), and satisfactory desirability of D = 0.79. However, in terms of the pigment with the highest content of anthocyanins, the optimal response (Figure 3B) was obtained at the highest SC (60%) and the lowest SSR (25%), obtaining a desirability coefficient of D = 0.90 and an anthocyanin content of 0.1449 mg cyanidin 3-glucoside /g of coffee fresh exocarp (0.578 mg cyanidin 3glucoside /g of coffee exocarp, dry basis), higher than that reported by Murthy et al. (2012) (0.240 mg/g of fresh 7

pulp, dry weight) and lower than that reported by Prata and Olivera (2007) (between 0.172 and 0.203 mg/g of fresh cherries). However, these differences may be due to different cultivation, harvest and storage conditions (Celli et al., 2015) and to the large number of variables that influenced the extraction of these compounds (Heras et al., 2013). The experiment validation was carried out by applying the optimal conditions predicted by the design, finding a value of 0.1463 ± 0.010 mg cyanidin 3-glucoside /g of coffee fresh exocarp; the differences between the theoretical and experiment values were not significant (T-test). 3.4. Application of the coffee exocarp extract to French meringue There was a decrease in pH and soluble solids and an increase in acidity, anthocyanin content and ΔE as the concentration of the natural pigment was increased (Table 3), obtaining significant differences between the treatments. The anthocyanin content at the 3% concentration was similar to that obtained in other products enriched with anthocyanins (Adsare et al., 2016). The 3% concentration had the lowest ΔE statistically, as compared to the control (Table 3). Also, there was a reduction in the color coordinates L* and C* and an increase in a*, b* and h* as the concentration of the natural dye increased (3, 6 and 9%), similar to reports from other researchers (Adsare et al., 2016; Rosero et al., 2016). The anthocyanin content was higher at the 9% concentration; however, a loss of properties (shape, texture and taste) characteristic of French meringue was also observed, which may have been attributable to the decrease in pH and increase in acidity (Table 3). It has been reported that pH is a key factor in the stabilization of anthocyanins and that a pH above 1 favors degradation, formation of non-reactive complexes and colorless forms of hemiacetal and chalcone (Kırca, Özkan, & Cemeroglu, 2006; Garzón, 2008; Buchweitz et al., 2013), limiting the application of anthocyanins as natural dyes in foods (Hurtado and Pérez, 2014, Ongkowijoyo, LunaVital and Gonzalez de Mejia, 2018). On the other hand, it has been reported that albumin foam is stable at pH= 8.6, the pH of natural egg whites, and decreases with changes in pH; however, it has been found that stability increases over time to pH= 4.8 because of the formation of small bubbles, using acids and acid salts for this purpose. However, an extremely acid pH= 1 leads to the irreversible denaturation of the protein (Lomakina and Míková, 2006), which may explain why, as the concentration of the acidified pigment was increased to pH= 1, the properties inherent to French meringue were lost in the product. In studies conducted by Pasqualone et al. (2014), changes in the physicochemical 8

properties of cookies enriched with an anthocyanin extract were reported. Microencapsulation and the study of degradation kinetics increase the stability and protection of these pigments (Arrazola, Herazo and Alvis, 2014, Assous et al., 2014; Özkan and Bilek, 2014); therefore, it is recommended that techniques that increase the potential of coffee cherry pigments as natural dyes be explored. 4. Conclusions The SC and SSR significantly influenced the anthocyanin content and surface color changes of the coffee cherry pigment extracts. The optimal extraction conditions that obtained the highest content of anthocyanins (0.1449 mg/g of fresh cherries) included an SC of 60% and SSR of 25%. In the production of French meringue, the 3% concentration of coffee cherry pigments produced a color close to that obtained with synthetic pigments; however, it was also found that, as the concentration of the natural pigment increased, the pH decreased, losing the characteristic properties of meringue. Acknowledgment This research was supported by Universidad Nacional de Colombia Sede Palmira. Conflict of interest The authors have no conflict of interest for this manuscript

References Abdel-Aal, E. S. M., Young, J. C., & Rabalski, I. (2006). Anthocyanin Composition in black, blue, pink, purple, and red. Journal of Agriculture and Food Chemistry, 54, 4696–4704. Adsare, S. R., Bellary, A. N., Sowbhagya, H. B., Baskaran, R., Prakash, M., & Rastogi, N. K. (2016). Osmotic treatment for the impregnation of anthocyanin in candies from Indian gooseberry (Emblica officinalis). Journal of Food Engineering, 175, 24-32 Aguilera-Ortíz, M., del Carmen Reza-Vargas, M., Madinaveitia, R. G. C., Valenzuela, J. A., & Baca, P. R. (2012). Antocianinas de higo como colorantes para yogur natural. Biotecnia, 14(1), 18-24. Amchova, P., Kotolova, H., & Ruda-Kucerova, J. (2015). Health safety issues of synthetic food colorants. Regulatory Toxicology and Pharmacology, 73(3), 914-922. 9

Arrazola, G., Herazo, I. and Alvis, A. (2014) ‘Microencapsulación de antocianinas de berenjena (Solanum melongena L.) mediante Secado por aspersión y evaluación de la estabilidad de su color y capacidad antioxidante’, Informacion Tecnologica, 25(3), 31–42. Assous, M. T. M., Abdel-Hady, M. M., & Medany, G. M. (2014). Evaluation of red pigment extracted from purple carrots and its utilization as antioxidant and natural food colorants. Annals of Agricultural Sciences, 59(1), 1-7. Bonilla-Hermosa, V. A., Duarte, W. F., & Schwan, R. F. (2014). Utilization of coffee by-products obtained from semi-washed process for production of value-added compounds. Bioresource technology, 166, 142-150. Buchweitz, M., Brauch, J., Carle, R., & Kammerer, D. R. (2013). Application of ferric anthocyanin chelates as natural blue food colorants in polysaccharide and gelatin based gels. Food research international, 51(1), 274282. Cacace, J. E., & Mazza, G. (2003). Optimization of extraction of anthocyanins from black currants with aqueous ethanol. Journal of Food Science, 68(1), 240-248. Cao, S. Q., Liang, L. I. U., & Pan, S. Y. (2011). Thermal degradation kinetics of anthocyanins and visual color of blood orange juice. Agricultural Sciences in China, 10(12), 1992-1997. Carvajal-Herrera, J. J., Aristizábal-Torres, I. D., Oliveros-Tascón, C. E., & Mejía-Montoya, J. W. (2011). Colorimetría del fruto de café (Coffea arabica L.) durante su desarrollo y maduración. Revista Facultad Nacional de Agronomía-Medellín, 64(2), 6229-6240. Celli, G. B., Ghanem, A., & Brooks, M. S. L. (2015). Optimization of ultrasound-assisted extraction of anthocyanins from haskap berries (Lonicera caerulea L.) using Response Surface Methodology. Ultrasonics sonochemistry, 27, 449-455. Cevallos-Casals, B. A., & Cisneros-Zevallos, L. (2004). Stability of anthocyanin-based aqueous extracts of Andean purple corn and red-fleshed sweet potato compared to synthetic and natural colorants. Food chemistry, 86(1), 69-77. Darıcı, S., & Şen, S. (2015). Experimental investigation of convective drying kinetics of kiwi under different conditions. Heat and Mass Transfer, 51(8), 1167-1176. Esquivel, P., & Jiménez, V. M. (2012). Functional properties of coffee and coffee by-products. Food Research 10

International, 46(2), 488-495. Fan, G., Han, Y., Gu, Z., & Chen, D. (2008). Optimizing conditions for anthocyanins extraction from purple sweet potato using response surface methodology (RSM). LWT-Food Science and Technology, 41(1), 155-160. Garzón, G. A. (2008). Las antocianinas como colorantes naturales y compuestos bioactivos: revisión. Acta Biológica Colombiana, 13(3), 27-36. Gerardi, C., Albano, C., Calabriso, N., Carluccio, M. A., Durante, M., Mita, G., Renna M., Serio, F., & Blando, F. (2018). Techno-functional properties of tomato puree fortified with anthocyanin pigments. Food chemistry, 240, 1184-1192. Goldfarb, W. (2016). Making a Balinese Meringue. International Journal of Gastronomy and Food Science, 4, 12-18. Güneşer, O. (2016). Pigment and color stability of beetroot betalains in cow milk during thermal treatment. Food chemistry, 196, 220-227. Heeger, A., Kosińska-Cagnazzo, A., Cantergiani, E., & Andlauer, W. (2017). Bioactives of coffee cherry pulp and its utilisation for production of Cascara beverage. Food chemistry, 221, 969-975. Heras, I., Alvis, A., & Arrazola, G. (2013). Optimización del Proceso de Extracción de Antocianinas y Evaluación de la Capacidad Antioxidante de Berenjena (Solana melonera L.). Información tecnológica, 24(5), 93-102. Hurtado, N. H., & Pérez, M. (2014). Identificación, estabilidad y actividad antioxidante de las antocianinas aisladas de la cáscara del fruto de capulí (Prunus serotina spp capuli (Cav) Mc. Vaug Cav). Información tecnológica, 25(4), 131-140. ICO

(2018)

Trade

statistics,

International

coffee

organization.

http://www.ico.org/trade_statistics.asp?section=Statistics. Accessed 10 April 2018. Kara, Ş., & Erçelebi, E. A. (2013). Thermal degradation kinetics of anthocyanins and visual colour of Urmu mulberry (Morus nigra L.). Journal of Food Engineering, 116(2), 541-547. Kırca, A., Özkan, M., & Cemeroglu, B. (2006). Stability of black carrot anthocyanins in various fruit juices and nectars. Food Chemistry, 97(4), 598-605. Lachman, J., Hamouz, K., Šulc, M., Orsák, M., Pivec, V., Hejtmánková, A., Dvorak, P., & Čepl, J. (2009). 11

Cultivar differences of total anthocyanins and anthocyanidins in red and purple-fleshed potatoes and their relation to antioxidant activity. Food Chemistry, 114(3), 836-843. Liu, J., Dong, N., Wang, Q., Li, J., Qian, G., Fan, H., & Zhao, G. (2014). Thermal degradation kinetics of anthocyanins from Chinese red radish (Raphanus sativus L.) in various juice beverages. European Food Research and Technology, 238(2), 177-184. Lomakina, K., & Mikova, K. (2006). A study of the factors affecting the foaming properties of egg white–a review. Czech Journal of Food Sciences, 24(3), 110-118. Naidu, M. M., Sulochanamma, G., Sampathu, S. R., & Srinivas, P. (2008). Studies on extraction and antioxidant potential of green coffee. Food Chemistry, 107(1), 377-384. Marín-López, S. M., Arcila-Pulgarin, J., Montoya-Restrepo, E. C., & Oliveros-Tascón, C. E. (2003). Cambios físicos y químicos durante la maduración del fruto de café coffea Arabica l var Colombia. Cenicafé, 54(3), 208225. Mercali, G. D., Jaeschke, D. P., Tessaro, I. C., & Marczak, L. D. F. (2013). Degradation kinetics of anthocyanins in acerola pulp: Comparison between ohmic and conventional heat treatment. Food Chemistry, 136(2), 853-857. Mohideen, F. W., Solval, K. M., Li, J., Zhang, J., Chouljenko, A., Chotiko, A., Prudente, A. D., Bankston, J. D., & Sathivel, S. (2015). Effect of continuous ultra-sonication on microbial counts and physico-chemical properties of blueberry (Vaccinium corymbosum) juice. LWT-Food Science and Technology, 60(1), 563-570. Murthy, P. S., Manjunatha, M. R., Sulochannama, G., & Naidu, M. M. (2012). Extraction, characterization and bioactivity of coffee anthocyanins. European Journal of Biological Sciences, 4(1), 13-19. O'Charoen, S., Hayakawa, S., Matsumoto, Y., & Ogawa, M. (2014). Effect of d‐ Psicose Used as Sucrose Replacer on the Characteristics of Meringue. Journal of food science, 79(12), E2463-E2469. Ongkowijoyo, P., Luna-Vital, D. A., & de Mejia, E. G. (2018). Extraction techniques and analysis of anthocyanins from food sources by mass spectrometry: An update. Food chemistry, 250(1), 113-126. Özkan, G., & Bilek, S. E. (2014). Microencapsulation of natural food colourants. International Journal of Nutrition and Food Sciences, 3(3), 145-156. Pasias, I. N., Asimakopoulos, A. G., & Thomaidis, N. S. (2015). Food colours for bakery products, snack foods, dry soup mixes, and seasonings. In Colour additives for foods and beverages (pp. 211-226). 12

Pasqualone, A., Bianco, A. M., Paradiso, V. M., Summo, C., Gambacorta, G., & Caponio, F. (2014). Physicochemical, sensory and volatile profiles of biscuits enriched with grape marc extract. Food Research International, 65, 385-393. Patil, G., Madhusudhan, M. C., Babu, B. R., & Raghavarao, K. S. M. S. (2009). Extraction, dealcoholization and concentration of anthocyanin from red radish. Chemical Engineering and Processing: Process Intensification, 48(1), 364-369. Pedro, A. C., Granato, D., & Rosso, N. D. (2016). Extraction of anthocyanins and polyphenols from black rice (Oryza sativa L.) by modeling and assessing their reversibility and stability. Food chemistry, 191, 12-20. Prata, E. R., & Oliveira, L. S. (2007). Fresh coffee husks as potential sources of anthocyanins. LWT-Food Science and Technology, 40(9), 1555-1560. Ramos, P., Sanz, J., & Oliveros, C. (2014). Identificación y clasificación de frutos de café en tiempo real a través de la medición de color. Cenicafé, 61(4):315-326. 2010 Rodriguez-Amaya, D. B. (2016) ‘Natural food pigments and colorants’, Current Opinion in Food Science. 7, 20–26. Rosero, G., Corral, A., Álvarez, C., Serna, L., & Ordoñez, L. (2016). Evaluación del color en yogur elaborado con extracto de residuos de uvas Isabella como colorante natural. Agronomía Colombiana, 34(1Supl), S773S775. Rodrigues, S., Fernandes, F. A., de Brito, E. S., Sousa, A. D., & Narain, N. (2015). Ultrasound extraction of phenolics and anthocyanins from jabuticaba peel. Industrial Crops and Products, 69, 400-407. Salinas-Moreno, Y., Almaguer-Vargas, G., Peña-Varela, G., & Ríos-Sánchez, R. (2009). Ácido elágico y perfil de antocianinas en frutos de frambuesa (Rubus idaeus L.) con diferente grado de maduración. Revista Chapingo. Serie horticultura, 15(1), 97-101. Sompong, R., Siebenhandl-Ehn, S., Linsberger-Martin, G., & Berghofer, E. (2011). Physicochemical and antioxidative properties of red and black rice varieties from Thailand, China and Sri Lanka. Food Chemistry, 124(1), 132-140. Su, G., Zhu, S., Xu, M., Ramaswamy, H. S., Lin, Y., & Yu, Y. (2016). Pressure degradation kinetics of anthocyanin pigment and visual color of chinese bayberry juice. International journal of food properties, 19(2), 13

443-453. Swamy, G. J., Sangamithra, A., & Chandrasekar, V. (2014). Response surface modeling and process optimization of aqueous extraction of natural pigments from Beta vulgaris using Box–Behnken design of experiments. Dyes and Pigments, 111, 64-74.

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Figure 1. Effect of the solvent concentration and solid solvent ratio on the total anthocyanins.

15

(A)

16

(B)

17

(C)

18

(D) Figure 2. Effect of the solvent concentration and solid solvent ratio on the color coordinates L* (A), a* (B), C (C) and h° (D).

19

(A)

(B) Figure 3. Optimization of the solvent concentration and solid solvent ratio on the total anthocyanins and color (A) and optimization solvent concentration and solid solvent ratio for the total anthocyanins.

20

Table 1. Total anthocyanin and CIEL*a*b* of the extract from coffee exocarp in central composite design (n=3). Variables Codified

TA

Coordinates CIEL*a*b* of the extract from coffee exocarp

Uncoded

X1

X2

SC

SSR

mg/g

L*

a*

b*

C*

h°

-1

-1

40

25

0.111 ± 0.00

21.88 ± 0.22

5.79 ± 0.28

3.17 ± 0.41

6.55 ± 0.42

27.86 ± 2.34

-1

1

40

75

0.073 ± 0.01

20.96 ± 1.67

5.92 ± 0.51

3.35 ± 0.38

6.97 ± 0.61

31.80 ± 1.65

0

0

50

50

0.099 ± 0.00

19.67 ± 0.31

6.61 ± 0.06

3.17 ± 0.25

7.22 ± 0.15

27.27 ± 1.95

0

0

50

50

0.099 ± 0.00

20.34 ± 0.6

6.41 ± 0.28

3.20 ± 0.12

7.28 ± 0.24

28.22 ± 1.76

1.414

0

64

50

0.104 ± 0.02

19.93 ± 0.39

5.59 ± 0.15

3.05 ± 0.52

6.17 ± 0.38

27.92 ± 3.22

1

1

60

75

0.07 ± 0.00

21.01 ± 1.53

5.42 ± 0.7

3.22 ± 0.41

5.93 ± 0.56

31.25 ± 5.15

0

-1.414

50

15

0.126 ± 0.01

20.84 ± 0.75

6.45 ± 0.25

2.85 ± 0.28

6.69 ± 0.34

25.00 ± 1.18

-1.414

0

36

50

0.087 ± 0.02

21.50 ± 1.36

5.76 ± 0.8

3.49 ± 0.14

6.69 ± 0.79

30.55 ± 1.62

1

-1

60

25

0.155 ± 0.03

20.40 ± 1.53

6.02 ± 0.57

3.01 ± 0.53

6.72 ± 0.73

26.64 ± 2.18

0

0

50

50

0.087 ± 0.00

20.34 ± 1.41

6.51 ± 0.61

3.00 ± 0.27

7.28 ± 0.46

26.32 ± 4.10

0

1.414

50

85

0.053 ± 0.01

20.59 ± 0.68

5.72 ± 0.48

2.78 ± 0.24

6.30 ± 0.53

28.28 ± 0.89

Values are reported as the mean ± standard deviation; SC= solvent concentration (Ethanol percent, % v/v); SSR = solid solvent ratio (% p/v); TA= Total anthocyanin (mg cyanidin 3-glucoside /g of coffee fresh exocarp)

Table 2. Analysis of variance (ANOVA) and optimization parameters

Total anthocyanin

Intercept

SC

SSR

SC*SSR

0.097

0.0227

<0.0001 ***

0.0194

* L

20.12

0.0145

b

6.48

3.19

0.2212

0.0569

SSR2

* 0.5345

0.0817

* a

SC2

Model

Lack of fit

R2

< 0.0001

0.5202

0.95

8.09

0.6287

0.85

1.70

0.4388

0.95

2.15

0.0106

0.78

4.15

0.99

0.95

*** 0.048

0.048

0.0378

*

*

*

0.0092

0.0369

0.0006

0.010

0.0032

**

*

***

*

**

0.4715

0.9128

0.2389

0.0386

0.0803

* C

7.26

C.V. %

*

0.0003

0.0038

0.0002

<0.0001

<0.0001

<0.0001

***

**

***

***

***

***

0.1338

21

h

28.51

0.1822

0.0115

0.0220

*

*

0.1229

0.61

4.66

SC= solvent concentration; SSR = solid solvent ratio

Table 3. Color and selected chemical characteristics of French meringues added of different levels of extract from coffee exocarp (n=3). pH

SS

TAC

TA ∆E

Color coordinates CIEL*a*b*

Concentration extract (%) (%)

(%)

mg/g

L*

a*

b*

C*

h°

0

6.78± 0.01a

88.55 ± 0.78b

0.001 ± 0.00d

----

80.78 ± 0.71a

8.08 ± 1.14b

15.91 ± 3.30bc

17.85 ± 3.45bc

62.86 ± 1.77b

----

3

2.87± 0.01b

91.85 ± 0.78a

0.008 ± 0.00c

0.036 ± 0.003b

83.64 ± 0.95a

6.58 ± 0.13ab

12.91 ± 0.35c

14.49 ± 0.31c

62.99 ± 0.83b

4.41± 0.38c

6

2.66± 0.01c

72.05 ± 0.16c

0.012 ± 0.00b

0.069 ± 0.013b

64.27 ± 1.05b

8.26 ± 0.05a

18.22 ± 0.22ab

20.00 ± 0.22ab

65.60 ± 0.17ab

16.67± 1.03b

9

2.45± 0.01d

53.35 ± 0.78d

0.016 ± 0.00a

0.284 ± 0.045a

59.34 ± 3.40c

9.50 ± 0.13a

22.88 ± 1.36a

24.77 ± 1.28 a

67.42 ± 1.11a

22.59± 2.84a

Average values ± standard deviation, Level of statistical significance: different letters in the sub-indices of the same column indicate differences significant according to Tukey's mean comparison (P <0.05), SS: Solubles solids. TAC: Titratable acidity (% malic acid). TA: Total anthocyanins (mg cyanidin 3-glucoside /g of baked meringue). ΔE: total color difference.

22

Highlights

1. 2. 3. 4.

Coffee exocarp is an important source of anthocyanins Concentration solvent (60%) and solid solvent ratio (25%) extract highest anthocyanin content Color coordinate L* presented an inverse behavior in anthocyanin content 3% coffee anthocyanins extract is an important color alternative in French meringue

23