Revista Mexicana de Ingeniería Química ISSN: 1665-2738 [email protected] Universidad Autónoma Metropolitana Unidad Iztapalapa México

Corona-González, R. I.; Ramos-Ibarra, J.R.; Gutiérrez-González, P.; Pelayo-Ortiz, C.; Guatemala- Morales, G. M.; Arriola-Guevara, E. THE USE OF RESPONSE SURFACE METHODOLOGY TO EVALUATE THE FERMENTATION CONDITIONS IN THE PRODUCTION OF TEPACHE Revista Mexicana de Ingeniería Química, vol. 12, núm. 1, 2013, pp. 19-28 Universidad Autónoma Metropolitana Unidad Iztapalapa Distrito Federal, México

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Volumen 12, N´umero1, Abril 2013 ISSN 1665-2738 Vol. 12,CONTENIDO No. 1 (2013) 19-28 1

THEVolumen USE 8, OF número RESPONSE 3, 2009 / SURFACEVolume 8, number METHODOLOGY 3, 2009 TO EVALUATE THE FERMENTATION CONDITIONS IN THE PRODUCTION OF TEPACHE 1

EL USO DE LA METODOLOGIA´ DE SUPERFICIE DE RESPUESTA PARA EVALUAR213 Derivation LAS and CONDICIONESapplication of the Stefan-Maxwell DE FERMENTACI equations ON´ EN LA PRODUCCION´ DE TEPACHE (Desarrollo y aplicación de las ecuaciones de Stefan-Maxwell) 1 1 1 Stephen WhitakerR.I. Corona-Gonzalez´ , J.R. Ramos-Ibarra , P. Gutierrez-Gonz´ alez´ , 1 2 1 C. Pelayo-Ortiz , G.M. Guatemala-Morales and E. Arriola-Guevara ∗

1Departamento de Ingenier´ıaQu´ımica,CUCEI, Universidad de Guadalajara. Blvd. Marcelino Garc´ıaBarrag´an# Biotecnología / Biotechnology1421, C.P. 44430, Guadalajara, Jalisco, M´exico. 2Unidad de Tecnolog´ıaAgroalimentaria, Centro de Investigaci´ony Asistencia en Tecnolog´ıay Dise˜nodel Estado 245 Modelado de la biodegradación en biorreactores de lodos de hidrocarburos totales del petróleo de Jalisco, A.C. (CIATEJ, A.C.). Av. Normalistas # 800, Col. Colinas de la Normal, CP 44270, Guadalajara, intemperizados en suelos y sedimentos Jalisco, M´exico.

(Biodegradation Receivedmodeling of 23 sludge of August bioreactors 2012; Acceptedof total petroleum 12 of December hydrocarbons 2012 weathering in soil Abstract and sediments) Due to the artisanS.A. Medina-Moreno, nature of the production S. Huerta-Ochoa, process of tepache, C.A. Lucho-Constantino, there is no uniformity L. Aguilera-Vázquez, in fermentation conditions A. Jiménez- and raw material used. The fermentation is crucial to the final characteristics of tepache and there is a certain degree of ignorance of related González y M. Gutiérrez-Rojas organisms and the level of acceptance among consumers. In this study, tepache fermentation conditions (concentration of sugars, initial pH,259 temperatureCrecimiento, and sobrevivencia fermentation y time) adaptación were evaluated de Bifidobacterium to enable correlating infantis a condiciones the formation ácidas of products of fermentation (lactic, acetic and ethanol) with the degree of acceptance of the beverage among consumers. Sensory evaluation was measured (Growth, survival and adaptation of Bifidobacterium infantis to acidic conditions) on a 9-point hedonic scale. Results were analyzed using the response surface methodology (RSM) which showed that the fermentationL. conditions Mayorga-Reyes, for higher P. acceptanceBustamante-Camilo, were: 22oC, A. 10% Gutiérrez-Nava, (mass/volume) E. of Barranco-Florido sugars (brown sugar), y A. 72Azaola- h of fermentation and an initialEspinosa pH of 5. According to this study, to have a wider acceptance the beverage must contain about 7 g/L of ethanol, no more than 5 g/L of lactic and acetic acid, and 70 g of sucrose/L. Likewise, yeasts present in the fermentation were identified and 265 Statistical approach to optimization of ethanol fermentation by Saccharomyces cerevisiae in the it was found that Saccharomyces cerevisiae is the predominant species. Keywords:presence tepache, of fermentation, Valfor® zeolite response NaA surface methodology, hedonic scale, yeast identification.

Resumen (Optimización estadística de la fermentación etanólica de Saccharomyces cerevisiae en presencia de Debido a la naturaleza artesanal del proceso de produccion´ del tepache, no hay uniformidad en condiciones de fermentacion´ zeolita Valfor® zeolite NaA) y materia prima utilizadas. La fermentacion´ es crucial para las caracter´ısticas finales del tepache y existe cierto grado de desconocimientoG. Inei-Shizukawa, de los microorganismos H. A. Velasco-Bedrán, relacionados G. y elF. nivelGutiérrez-López de aceptacion ´and entre H. Hernández-Sánchez los consumidores. En este estudio, las condiciones de fermentacion´ (concentracion´ de azucar,´ pH inicial, temperatura y tiempo de fermentacion)´ fueron evaluadas para correlacionar los productos de la fermentacion´ (etanol, acido´ lactico´ y acetico)´ con el grado de aceptacion´ de la bebida. La Ingeniería de procesos / Process engineering evaluacion´ sensorial se realizo´ con una escala hedonica´ de 9 puntos. Los resultados fueron analizados utilizando una metodolog´ıa de superficie271 Localización de respuesta (MSR),de una laplanta cual mostrindustrial:o´ que Revisión las condiciones crítica de y fermentaciadecuaciónon,´ de para los una criterios mayor empleados aceptacion,´ en fueron: 22oC, 10% (masa/volumen)esta decisión de az ucar´ morena (piloncillo), 72 horas de fermentacion´ y un pH inicial de 5. De acuerdo con este estudio, para tener una mayor aceptacion´ la bebida debe contener alrededor de 7 g/L de etanol, no mas´ de 5 g/L de acido´ acetico´ y (Plant site selection: Critical review and adequation criteria used in this decision) lactico,´ y 70 g/L de sacarosa. Asimismo, se identificaron las especies de levaduras presentes en la fermentacion´ y se encontro´ que SaccharomycesJ.R. Medina, cerevisiae R.L. Romeroes la especie y G.A. predominante. Pérez Palabras clave: tepache, fermentacion,´ metodolog´ıa de superficie de respuesta, escala hedonica,´ identificacion´ de levaduras.

∗Corresponding author. E-mail: [email protected] Tel. 52 (33) 3817-3724; Fax: 52 (33) 13785920 ext. 27536

Publicado por la Academia Mexicana de Investigacion´ y Docencia en Ingenier´ıa Qu´ımica A.C. 19 / Corona-Gonz´alezetCorona-Gonz´alezetCorona-Gonz´alezet al. al. al.//RevistaRevistaRevista Mexicana Mexicana Mexicana de de deIIngenierngenierIngenier´ıa´ıaQ´ıaQuQu´ımicau´ımica´ımica Vol. Vol. Vol. 12, 12, 12, No. No. No. 1 1 (2013) 1 (2013) (2013) XXX-XXX XXX-XXX 19-28

41 92 evaluations (Deshpande et al., 2008). In this 41 11 Introduction Introduction 92 evaluations (Deshpande et al., 2008). In this 93 study, using a central composite design (CCD), 93 study, using a central composite design (CCD), 42 Long time ago, since pre-Hispanic era, fermented 94 we evaluated the main process variables for 42 Long time ago, since pre-Hispanic era, fermented 94 we evaluated the main process variables for 43 beverages were widely consumed in Mexico. Among 95 tepache fermentation (brown sugar concentration, 43 beverages were widely consumed in Mexico. Among 95 tepache fermentation (brown sugar concentration, 44 the most popular are tepache, pulque, tejuino, and 96 fermentation temperature, initial pH and fermentation 44 the most popular are tepache, pulque, tejuino, and 96 fermentation temperature, initial pH and fermentation 45 pozol, all considered soft drinks with low alcohol 97 time) to correlate the formation of fermentation 45 pozol, all considered soft drinks with low alcohol 97 time) to correlate the formation of fermentation 46 content (Godoy et al., 2003). Tepache is the 98 products (lactic acid, acetic acid, and ethanol) with 46 content (Godoy et al., 2003). Tepache is the 98 products (lactic acid, acetic acid, and ethanol) with 47 most popular traditional fermented drink, prepared 99 the acceptance of the beverage among consumers. 47 most popular traditional fermented drink, prepared 99 the acceptance of the beverage among consumers. 48 using traditional methods from pineapple peels, sugar 100 The results were analyzed using the RSM and the 48 using traditional methods from pineapple peels, sugar 100 The results were analyzed using the RSM and the 49 (brown sugar, a type of unrefined sugar cane), water 101 yeast in the fermentation was also identified. With 49 (brown sugar, a type of unrefined sugar cane), water 101 yeast in the fermentation was also identified. With 50 and spices (cinnamon and pepper). Fermentation 102 this study we claim to increase the knowledge of the 50 and spices (cinnamon and pepper). Fermentation 102 this study we claim to increase the knowledge of the 51 of tepache takes place in wooden barrels, at room 103 fermentation process of this type of beverages and 51 of tepache takes place in wooden barrels, at room 103 fermentation process of this type of beverages and 52 temperature, from 1 to 4 days (Alvarado et al., 2006). 104 also contribute to the discernment of the control of 52 temperature, from 1 to 4 days (Alvarado et al., 2006). 104 also contribute to the discernment of the control of 53 In distilled spirits, such as tequila and wine, ethanol 105 fermentation conditions for industrialization. 53 In distilled spirits, such as tequila and wine, ethanol 105 fermentation conditions for industrialization. 54 is desirable as the major product and total sugar 54 is desirable as the major product and total sugar 55 consumption during fermentation. In contrast, tepache 55 consumption during fermentation. In contrast, tepache 56 is a not distilled beverage, ethanol concentration 106 2 Materials and methods 56 is a not distilled beverage, ethanol concentration 106 2 Materials and methods 57 should be low and there is partial consumption of 57 should be low and there is partial consumption of 58 sugars to provide a sweet taste, and a production of 107 2.1 Preparation of tepache 58 sugars to provide a sweet taste, and a production of 107 2.1 Preparation of tepache 59 lactic, acetic and other volatile compounds that give it 59 lactic, acetic and other volatile compounds that give it 108 The fermentation was performed in closed stainless- 60 its organoleptic characteristics. Tepache fermentation 108 The fermentation was performed in closed stainless- 60 its organoleptic characteristics. Tepache fermentation 109 steel containers with 10-liter capacities. The top 61 is affected by environmental, chemical and biological 109 steel containers with 10-liter capacities. The top 61 is affected by environmental, chemical and biological 110 part of the containers had a 7-mm hole, which 62 factors, such as temperature, concentration of sugars, 110 part of the containers had a 7-mm hole, which 62 factors, such as temperature, concentration of sugars, 111 was sealed with a cotton plug to simulate semi- 63 pH, and fermentation time, among others, which 111 was sealed with a cotton plug to simulate semi- 63 pH, and fermentation time, among others, which 112 anaerobic conditions. The panela (brown sugar from 64 modify the final product characteristics (Aidoo et al., 112 anaerobic conditions. The panela (brown sugar from 64 modify the final product characteristics (Aidoo et al., 113 Pihuamo, Jalisco, Mexico) was dissolved in water by 65 2006). If the fermentation continues for more than 113 Pihuamo, Jalisco, Mexico) was dissolved in water by 65 2006). If the fermentation continues for more than 114 shaking, and subsequently 9% (w/v) of pineapple rind 66 four days, the concentration of acetic acid increases 114 shaking, and subsequently 9% (w/v) of pineapple rind 66 four days, the concentration of acetic acid increases 115 (Pineapples Ananas comosus from Diva, Veracruz, 67 and the taste is unpleasant (Moreno-Terrazas, 2005; 115 (Pineapples Ananas comosus from Diva, Veracruz, 67 and the taste is unpleasant (Moreno-Terrazas, 2005; 116 Mexico), which had previously been cut into pieces, 68 Swiers et al., 2005). 116 Mexico), which had previously been cut into pieces, 68 Swiers et al., 2005). 117 was added. The fermentation was carried out without 69 By its artisan nature, raw materials used, 117 was added. The fermentation was carried out without 69 By its artisan nature, raw materials used, 118 agitation. The initial pH of the fermentations was 70 temperature and fermentation time, the tepache 118 agitation. The initial pH of the fermentations was 70 temperature and fermentation time, the tepache 119 adjusted with NaOH or citric acid solutions. 71 process has a high degree of variation that affects 119 adjusted with NaOH or citric acid solutions. 71 process has a high degree of variation that affects 72 its fermentation; accordingly, the final characteristics 72 its fermentation; accordingly, the final characteristics 73 of the drink are different among preparations of 120 2.2 Determination of sugar and 73 of the drink are different among preparations of 120 2.2 Determination of sugar and 74 different origin (Moreno-Terrazas, 2005). Tepache 74 different origin (Moreno-Terrazas, 2005). Tepache 121 fermentation products 121 fermentation products 75 has potential for industrial scale production but it 75 has potential for industrial scale production but it 76 is necessary to know the fermentation conditions 122 Sucrose, glucose, fructose, ethanol, lactic and acetic 76 is necessary to know the fermentation conditions 122 Sucrose, glucose, fructose, ethanol, lactic and acetic 77 to generate a product of wider acceptance among 123 acids, were determined by HPLC. Sugars were 77 to generate a product of wider acceptance among 123 acids, were determined by HPLC. Sugars were 78 consumers. Furthermore, the shell of the pineapple 124 analyzed with a Bio-Rad Aminex HPX-87C (300 x 7.8 78 consumers. Furthermore, the shell of the pineapple 124 analyzed with a Bio-Rad Aminex HPX-87C (300 x 7.8 79 represents about 40% of the fruit and, since it is a 125 mm) with water, as mobile phase, at a flow rate of 0.5 79 represents about 40% of the fruit and, since it is a 125 mm) with water, as mobile phase, at a flow rate of 0.5 80 waste of industrialization, its decomposition pollute 126 mL/min and 60◦C. Acids and ethanol were quantified 80 waste of industrialization, its decomposition pollute 126 mL/min and 60◦C. Acids and ethanol were quantified 81 the environment (Moreno-Terrazas, 2005). It is likely 127 using an Alltech OA-100 column of 300 mm x 6.5 mm 81 the environment (Moreno-Terrazas, 2005). It is likely 127 using an Alltech OA-100 column of 300 mm x 6.5 mm 82 that in the process of industrialization of tepache, 128 at 60◦C and at a flow of 0.5 mL/min, with a mobile 82 that in the process of industrialization of tepache, 128 at 60◦C and at a flow of 0.5 mL/min, with a mobile 83 pineapple by-products will be totally used, giving 129 phase of 0.01N H2SO4, filtered and degasified. The 83 pineapple by-products will be totally used, giving 129 phase of 0.01N H2SO4, filtered and degasified. The 84 added value to this waste. 130 device used was integrated to a Waters 600 controller, 84 added value to this waste. 130 device used was integrated to a Waters 600 controller, 85 There are few studies about the characteristics 131 a Waters 717 plus auto injector, and a Waters 2410 RI 85 There are few studies about the characteristics 131 a Waters 717 plus auto injector, and a Waters 2410 RI 86 of tepache and the effect of fermentation conditions 132 detector. 86 of tepache and the effect of fermentation conditions 132 detector. 87 on the features of the beverage and its acceptance. 87 on the features of the beverage and its acceptance. 88 Sensory evaluation is often used to determine the 88 Sensory evaluation is often used to determine the 133 2.3 Experimental design 133 2.3 Experimental design 89 degree of acceptance of a product; similarly, the 89 degree of acceptance of a product; similarly, the 90 Response Surface Methodology (RSM) is used 134 To evaluate the fermentation conditions of the tepache 90 Response Surface Methodology (RSM) is used 134 To evaluate the fermentation conditions of the tepache 91 to optimize production with respect to sensory 135 (temperature, concentration of panela, initial pH, and 91 to optimize production with respect to sensory 135 (temperature, concentration of panela, initial pH, and

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136 178 136 fermentationfermentation time), time), a a non-replicate non-replicate central central composite composite 178 werewere determined determined using using a a P-value P-value with with a a probability probability 137 179 137 designdesign was was used. used. The The distance distance of of the the axial axial points points to to 179 valuevalue of of less less than than 0.05. 0.05. 138 the center of the design was α = 1.61 to allow the 138 the center of the design was α = ± 1.61 to allow the 139 design to rotate. Four central points± were established 139 design to rotate. Four central points were established 180 180 2.62.6 Identification Identification of of yeasts yeasts 140 140 toto provide provide a a reasonably reasonably stable stable variance variance of of the the predicted predicted 141 181 141 responseresponse (Montgomery, (Montgomery, 2010). 2010). The The range range and and levels levels of of 181 TheThe yeasts yeasts were were isolated isolated from from tepache tepache after after 72 72 h h 142 182 142 thethe researched researched variables variables are are presented presented in in Table Table 1. 1. 182 ofof fermentation. fermentation. They They were were grown grown using using GPYA GPYA 143 183 143 TheThe behavior behavior of of the the response response surface surface was was studied studied 183 mediummedium at at 30 30◦◦CC for for 48 48 h, h, to to subsequently subsequently select select the the 144 184 ff 144 withwith respect respect to to the the response response function function ( (YY)) using using a a 184 didifferenterent colonies colonies and and identify identify them them according according to to their their 145 185 145 polynomialpolynomial regression regression equation. equation. The The generalized generalized 185 morphologymorphology and and frequency frequency in in the the culture culture medium. medium. 146 186 146 responseresponse surface surface model model is is given given by by Eq. Eq. (1): (1): 186 TheThe selected selected colonies colonies were were inoculated inoculated into into tubes tubes 187 µ 187 withwith 800 800 µll of of GPY GPY broth broth to to extract extract the the DNA DNA after after 188 12 h of incubation at 30◦C and agitation at 100 22 188 12 h of incubation at 30◦C and agitation at 100 Y == ββ0 ++ββ1X1 ++ββ2X2 ++ββ3X3 ++ββ4X4 ++ββ11X1 Y 0 1X1 2X2 3X3 4X4 11X1 189 189 rpm.rpm. Yeast Yeast DNA DNA was was extracted extracted according according to to Querol Querol 22 22 22 +β22X + β33X + β44X + β12X1X2 + β13X1X3 (1) 190 +β22X22 + β33X33 + β44X44 + β12X1X2 + β13X1X3 (1) 190 etet al al.. (1992). (1992). Colonies Colonies isolated isolated were were identified identified by by +β + β + β + β 191 PCR amplification of the region spanning internal +β1414XX11XX44 + β2323XX22XX33 + β2424XX22XX44 + β3434XX33XX44 191 PCR amplification of the region spanning internal 192 192 transcribedtranscribed spacers spacers 1 1 and and 2 2 (ITS-1, (ITS-1, and and ITS-2) ITS-2) and and 147 147 Where Y is the response variable, X1, X2, X3, and 193 Where Y is the response variable, X1, X2, X3, and 193 thethe 5.8S 5.8S rRNA rRNA gene gene (5.8S-ITS (5.8S-ITS region) region) and and subsequent subsequent 148 148 X4 are the independent variables with respect to the 194 X4 are the independent variables with respect to the 194 restrictionrestriction analysis analysis according according to to and and compared compared with with 149 149 fermentation temperature, the amount of panela, the 195 fermentation temperature, the amount of panela, the 195 Esteve-ZarzosoEsteve-Zarzosoetet al al.. (1999). (1999). 150 β 150 initialinitial pH, pH, and and the the fermentation fermentation time, time, respectively, respectively, β00 151 β β β β 151 isis the the intercept intercept term, term, β11,, β22,, β33,, and and β44 areare the the linear linear 152 effects, β11, β22, β33, and β44 are the quadratic effects, 196 152 effects, β11, β22, β33, and β44 are the quadratic effects, 196 33 Results Results and and discussion discussion 153 β β β β β β 153 andand β1212,, β1313,, β1414,, β2323,, β2424,, and and β3434 areare the the interaction interaction 154 terms (Nwabueze et al., 2010). 197 154 terms (Nwabueze et al., 2010). 197 3.13.1 Conditions Conditions of of fermentation fermentation and and sensory sensory 198 198 evaluationevaluation 155 155 2.4 Sensory evaluation 2.4 Sensory evaluation 199 199 InIn the the traditional traditional process process of of preparing preparing tepache, tepache, the the 156 200 temperature is not controlled during fermentation 156 150150 untrained untrained panelists panelists evaluated evaluated batches batches of of tepache, tepache, 200 temperature is not controlled during fermentation 157 201 and it may vary from 10◦C to 35◦C throughout 157 preparedprepared according according to to the the CCD CCD combinations, combinations, with with 201 and it may vary from 10◦C to 35◦C throughout 158 202 the year, which could significantly affect the final 158 regardregard to to their their overall overall acceptance. acceptance. The The panelists panelists were were 202 the year, which could significantly affect the final 159 203 products of the drink and its acceptance. Likewise, 159 selectedselected to to participate participate based based on on their their preference preference for for 203 products of the drink and its acceptance. Likewise, 160 204 the concentration of sugars may vary because 160 fermentedfermented drinks, drinks, interest, interest, and and availability. availability. Samples Samples of of 204 the concentration of sugars may vary because 161 205 fermentation time. The fermentation time also 161 approximatelyapproximately 50 50 mL mL were were served served at at a a temperature temperature of of 205 fermentation time. The fermentation time also 162 206 depends on temperature, and at different times the final 162 4-94-9◦◦C;C; an an evaluation evaluation sheet sheet accompanied accompanied the the samples. samples. 206 depends on temperature, and at different times the final 163 207 products will have different concentrations. The pH 163 TheThe sessions sessions were were performed performed at at room room temperature temperature (24- (24- 207 products will have different concentrations. The pH 164 208 depends on the acidity of the pineapple so it could also 164 2828◦◦C).C). The The tepache tepache was was evaluated evaluated according according to to a a 9- 9- 208 depends on the acidity of the pineapple so it could also 165 = = 209 affect the formation of products in the beverage. A 165 pointpoint hedonic hedonic scale scale (1 (1 =dislikedislike extremely, extremely, 2 2 =dislikedislike 209 affect the formation of products in the beverage. A 166 = = 210 preliminary study determined the choice of all these 166 veryvery much, much, 3 3 =dislikedislike moderately, moderately, 4 4 =dislikedislike slightly, slightly, 210 preliminary study determined the choice of all these 167 = = = 211 variables. The complete experimental design is shown 167 55 = neitherneither like like nor nor dislike, dislike, 6 6 = likelike slightly, slightly, 7 7 = likelike 211 variables. The complete experimental design is shown 168 = = 212 in Table 1. 168 moderately,moderately, 8 8 = likelike very very much, much, 9 9 = likelike extremely). extremely). 212 in Table 1. 169 213 The analysis of variance of CCD showed that the 169 Finally,Finally, the the mean mean value value of of the the evaluations evaluations was was used used for for 213 The analysis of variance of CCD showed that the 170 214 overall acceptance was significantly affected by all 170 statisticalstatistical analysis analysis (Deshpande (Deshpande etet al al.,., 2008; 2008; Duarte Duarte etet 214 overall acceptance was significantly affected by all 171 215 of the studied factors (P-value < 0.05). The most 171 alal.,., 2011; 2011; Valim Valimetet al al.,., 2003). 2003). 215 of the studied factors (P-value < 0.05). The most 216 216 significantsignificant statistical statistical factors factors were were the the fermentation fermentation 217 217 time and the amount of panela used, both for the linear 172 time and the amount of panela used, both for the linear 172 2.52.5 Statistical Statistical analysis analysis 218 ffi ffi 218 coecoefficientscients and and for for the the quadratic quadratic coe coefficients.cients. The The 173 219 173 AA statistical statistical analysis analysis was was performed performed using using the the 219 analysisanalysis also also shows shows that that there there is is an an optimal optimal point point of of 174 220 174 StatgraphicsStatgraphics Centurion Centurion XV XV program program (StatPoint, (StatPoint, Inc., Inc., 220 acceptanceacceptance for for tepache tepache located located among among the the levels levels of of the the 175 221 175 2005).2005). The The adjusted adjusted response response surface surface model model was was 221 factorsfactors or or variables variables considered considered for for fermentation; fermentation; this this is is 176 222 176 statisticallystatistically evaluated evaluated using using the the ANOVA ANOVA F F statistic, statistic, 222 suggestedsuggested by by the the fact fact that that three three quadratic quadratic terms terms in in the the 177 ff 223 177 andand the the significant significant e effectsects of of the the dependent dependent variables variables 223 modelmodel were were statistically statistically significant. significant.

www.rmiq.orgwww.rmiq.orgwww.rmiq.org 2133 Corona-Gonz´alezetCorona-Gonz´alezet al./ al.Revista/ Revista Mexicana Mexicana deI dengenierIngenier´ıaQ´ıauQ´ımicau ´ımica Vol. Vol. 12, 12, No. No. 1 (2013) 1 (2013) XXX-XXX 19-28 Corona-Gonz´alezet al./ Revista Mexicana deIngenier ´ıaQu ´ımica Vol. 12, No. 1 (2013) XXX-XXX

Table 1. Matrix and summary of the results of the central composite design Table 1. Matrix and summary of the results of the central composite design Run Temp. Conc. Initial Time Acceptancea Ethanol Lactic Acetic Sucrose Glucose Fructose Run Temp. Conc. Initial Time Acceptancea Ethanol Lactic Acetic Sucrose Glucose Fructose #(◦C) (%) pH (h) (g/L) acid acid (g/L) (g/L) (g/L) #(◦C) (%) pH (h) (g/L) acid acid (g/L) (g/L) (g/L) X1 X2 X3 X4 (g/L) (g/L) X1 X2 X3 X4 (g/L) (g/L) 1 25 5 4 48 6.57 2.06 1.12 ND 32.53 ND 2.70 1 25 5 4 48 6.57 2.06 1.12 ND 32.53 ND 2.70 2 35 5 4 48 3.93 3.57 14.12 4.94 10.71 10.75 10.53 2 35 5 4 48 3.93 3.57 14.12 4.94 10.71 10.75 10.53 3 25 15 4 48 7.47 7.23 2.98 ND 92.12 16.57 15.61 3 25 15 4 48 7.47 7.23 2.98 ND 92.12 16.57 15.61 4 35 15 4 48 6.87 6.05 18.45 6.18 87.29 21.78 21.36 4 35 15 4 48 6.87 6.05 18.45 6.18 87.29 21.78 21.36 5 25 5 6 48 5.93 ND 2.53 ND 35.46 1.28 4.42 5 25 5 6 48 5.93 ND 2.53 ND 35.46 1.28 4.42 6 35 5 6 48 5.17 1.72 9.97 3.22 39.58 ND ND 6 35 5 6 48 5.17 1.72 9.97 3.22 39.58 ND ND 7 25 15 6 48 6.13 4.42 6.61 ND 101.87 18.09 20.28 7 25 15 6 48 6.13 4.42 6.61 ND 101.87 18.09 20.28 8 35 15 6 48 6.37 6.64 20.88 5.28 88.87 11.03 12.80 8 35 15 6 48 6.37 6.64 20.88 5.28 88.87 11.03 12.80 9 25 5 4 96 4.18 16.06 17.39 6.01 ND 0.26 3.70 9 25 5 4 96 4.18 16.06 17.39 6.01 ND 0.26 3.70 10 35 5 4 96 3.18 22.17 25.05 4.49 1.85 0.81 1.38 10 35 5 4 96 3.18 22.17 25.05 4.49 1.85 0.81 1.38 11 25 15 4 96 7.18 27.31 33.92 6.50 5.24 30.46 45.52 11 25 15 4 96 7.18 27.31 33.92 6.50 5.24 30.46 45.52 12 35 15 4 96 3.93 52.72 21.64 1.12 3.96 9.51 23.50 12 35 15 4 96 3.93 52.72 21.64 1.12 3.96 9.51 23.50 224 13 25 5 6 96 3.73 12.27 20.10 6.31 ND 1.36 6.76 224 13 25 5 6 96 3.73 12.27 20.10 6.31 ND 1.36 6.76 14 35 5 6 96 3.27 18.26 16.34 3.03 2.07 0.89 1.02 14 35 5 6 96 3.27 18.26 16.34 3.03 2.07 0.89 1.02 15 25 15 6 96 4.12 22.09 15.06 15.23 6.99 29.87 44.05 15 25 15 6 96 4.12 22.09 15.06 15.23 6.99 29.87 44.05 16 35 15 6 96 5.40 30.25 30.24 6.36 0.73 4.13 26.52 16 35 15 6 96 5.40 30.25 30.24 6.36 0.73 4.13 26.52 17 22.0 10 5 72 7.73 7.30 5.25 4.16 67.63 4.48 1.09 17 22.0 10 5 72 7.73 7.30 5.25 4.16 67.63 4.48 1.09 18 38.0 10 5 72 6.87 5.71 8.45 5.59 57.20 12.87 8.86 18 38.0 10 5 72 6.87 5.71 8.45 5.59 57.20 12.87 8.86 19 30 2.0 5 72 2.80 9.85 6.04 4.37 0.49 ND 1.50 19 30 2.0 5 72 2.80 9.85 6.04 4.37 0.49 ND 1.50 20 30 18.0 5 72 6.47 11.05 24.35 7.28 61.34 12.19 30.41 20 30 18.0 5 72 6.47 11.05 24.35 7.28 61.34 12.19 30.41 21 30 10 3.4 72 6.00 21.59 9.44 ND 2.72 15.73 27.26 21 30 10 3.4 72 6.00 21.59 9.44 ND 2.72 15.73 27.26 22 30 10 6.6 72 4.50 16.36 7.79 6.35 7.28 16.15 20.83 22 30 10 6.6 72 4.50 16.36 7.79 6.35 7.28 16.15 20.83 23 30 10 5 33.4 6.93 1.18 6.49 2.77 73.86 7.44 3.76 23 30 10 5 33.4 6.93 1.18 6.49 2.77 73.86 7.44 3.76 24 30 10 5 110.6 2.67 27.56 25.23 10.30 ND 0.80 5.52 24 30 10 5 110.6 2.67 27.56 25.23 10.30 ND 0.80 5.52 25 30 10 5 72 7.40 7.00 17.73 3.58 18.69 20.06 26.33 25 30 10 5 72 7.40 7.00 17.73 3.58 18.69 20.06 26.33 26 30 10 5 72 7.23 3.80 15.99 5.53 29.93 19.73 18.08 26 30 10 5 72 7.23 3.80 15.99 5.53 29.93 19.73 18.08 27 30 10 5 72 5.80 11.95 14.39 ND 10.91 18.29 26.76 27 30 10 5 72 5.80 11.95 14.39 ND 10.91 18.29 26.76 28 30 10 5 72 6.63 9.56 10.65 ND 8.90 24.97 31.38 28 30 10 5 72 6.63 9.56 10.65 ND 8.90 24.97 31.38 aMean values of 30 evaluations; ND: Not detected aMean values of 30 evaluations; ND: Not detected

225 Using the results of the experiments and the statistical 237 developed to predict the sensory scores of the tepache 225 Using the results of the experiments and the statistical 237 developed to predict the sensory scores of the tepache 226 analysis, a second-order polynomial equation was 238 produced under different fermentation conditions was 226 analysis, a second-order polynomial equation was 238 produced under different fermentation conditions was 227 obtained (Equation 2) for the estimated value of 239 acceptable, considering that the response variable is 227 obtained (Equation 2) for the estimated value of 239 acceptable, considering that the response variable is 228 acceptance (Y) as a function of the temperature (X1, 240 a hedonic measurement, which may often display 228 acceptance (Y) as a function of the temperature (X1, 240 a hedonic measurement, which may often display 229 ◦C), the concentration of panela (X2, %), the initial 241 considerable variations (Valim et al., 2003). 229 ◦C), the concentration of panela (X2, %), the initial 241 considerable variations (Valim et al., 2003). 230 pH (X3), and the fermentation time (X4, h). 230 pH (X3), and the fermentation time (X4, h). 242 The quadratic model (Equation 2) for overall 242 The quadratic model (Equation 2) for overall 243 acceptance was used to generate the response Y = 4.97493 0.567992X1 + 0.728927X2 + 1.73046X3 243 acceptance was used to generate the response Y = 4.97493 − 0.567992X1 + 0.728927X2 + 1.73046X3 244 − 2 surface graphs. Fig. 1 illustrates the influence of the + . − + . . 244 surface graphs. Fig. 1 illustrates the influence of the 0 11907X4 0 097375X1X3 0 0282162X22 245 +0.11907X4 + 0.097375X1X3 − 0.0282162X2 fermentation time and the amount of panela on the 2 − 2 2 245 fermentation time and the amount of panela on the 0.465171X 2 0.00110431X 2 246 overall acceptance of the tepache. It can be observed . 3 . 4 246 overall acceptance of the tepache. It can be observed −0.465171X3 − 0.00110431X4 overall acceptance of the tepache. It can be observed − − (2) 247 by the curving of the graph (Fig. 1a) that both, the 2 (2) 247 by the curving of the graph (Fig. 1a) that both, the 231 The value of the correlation coefficient (R2) was 248 linear and the quadratic coefficients of these variables, 231 The value of the correlation coefficient (R ) was 248 linear and the quadratic coefficients of these variables, 232 0.826; this value was considered sufficiently high. 249 affect the overall acceptance. In addition, the optimal 232 0.826; this value was considered2 sufficiently high. 249 affect the overall acceptance. In addition, the optimal 233 Some authors indicate that an R2 value of at least 250 levels of each variable can be determined: between 40 233 Some authors indicate that an R value of at least 250 levels of each variable can be determined: between 40 234 0.80 is sufficient to explain the variability of a 251 h and 60 h for the fermentation time, and between 10% 234 0.80 is sufficient to explain the variability of a 251 h and 60 h for the fermentation time, and between 10% 235 model, and to explain the variance of sensory data 252 and 15% for the amount of panela (Fig. 1a). To help 235 model, and to explain the variance of sensory data 252 and 15% for the amount of panela (Fig. 1a). To help 236 (Prasad and Nath, 2002). Therefore, the model 253 visualize the shape of the response surface (Fig. 1b), 236 (Prasad and Nath, 2002). Therefore, the model 253 visualize the shape of the response surface (Fig. 1b), 224 www.rmiq.org 4 www.rmiq.org Corona-Gonz´alezet al./ Revista Mexicana deIngenier ´ıaQu ´ımica Vol. 12, No. 1 (2013) XXX-XXX Corona-Gonz´alezetCorona-Gonz´alezet al. al./ Revista/ Revista Mexicana Mexicana de deIngenierIngenier´ıaQ´ıauQ´ımicau ´ımica Vol. Vol. 12, 12, No. No. 1 (2013) 1 (2013) XXX-XXX 19-28

254 289 initial pH of 4, and 54 h of fermentation.

254 289 290 initialMicrobiologically, pH of 4, and 54 h oflow fermentation. values of pH in the 290291 fermentationMicrobiologically, of beverages low are values considered of pH to in be the a 291292 fermentationselective and of favorable beverages factor. are considered Most bacteria, to be with a 292293 selectivethe exception and favorable of acetic factor. and lactic Most bacteria, bacteria, prefer with a 293294 theneutral exception to slightly of acetic alkaline andpH lactic and bacteria, do not prefer develop a 294295 neutralat the low to slightly pH of must. alkaline However, pH and fordo not yeast, develop a pH 295296 atrange the of low between pH of 3 must. and 6 However, is more favorable for yeast, for a their pH 296297 rangegrowth of and between fermentation 3 and 6 activity. is more favorable The fermentation for their 255 Fig 1 297298 growthtemperature and fermentation of wine varies activity. widely The forfermentation temperatures 255256 ff Fig.Fig 1 1. E ect of time and amount of panela on the 298 299 temperaturethat may range of wine from varies 10◦C to widely 30◦C. for The temperatures advantages 256257 Fig. 1. Effect of time and amount of panela on the overall acceptance of tepache, (a) Response surface, 299300 that may range from 10◦C to 30◦C. The advantages 257 of low-temperature fermentation are the development 258 overall(b) surface acceptance contours. of tepache, (a) Response surface,

300 301 ofof low-temperature a more fruity and fermentation fresh character are the of the development drink, the 258 (b) surface contours.

301 302 offormation a more fruityand lower and losses fresh character of ethanol, of in the addition drink, to the a

302 303 formationreduction inand the lower risk oflosses contamination of ethanol, by in additionbacteria and, to a

303304 reductionconsequently, in the a risklower of risk contamination of producing by volatile bacteria acids. and,

304305 consequently, a lower risk of producing volatile acids./ Finally, wines must contain between 120 to 250 g L 305 / 306 Finally,of sugar wines because must at contain concentrations between greater 120 to than 250 g300L

306307 ofg/L, sugar the osmotic because pressure at concentrations may have greater a negative than eff 300ect

307308 gon/L, the the yeast, osmotic which pressure may may decrease have yeast a negative growth eff andect

308309 on the yeast, which may decrease yeast growth and fermentation activity (Benda, 1984).

259 309310 fermentation activity (Benda, 1984). Fig 2 Our results coincide with those found by other

259260 ff 310 Fig.Fig 2 2. E ect of pH and fermentation temperature 311 authorsOur resultsfor other coincide similar with fermented those found beverages; by other for

260261 Fig. 2. Effect of pH and fermentation temperature on the overall acceptance of tepache.(a) Response 311312 authorsinstance, for the other optimal similar values fermented for the fermentation beverages; for of

261 262 onsurface, the overall (b) surface acceptance contours. of tepache.(a) Response 312313 instance, the optimal values for the fermentation of mango wine were 22.53◦C and a pH of 3.8 (Suder et

262 surface, (b) surface contours. 313 314 mangoal., 2009), wine and were the 22.53 optimal◦C and fermentation a pH of 3.8 temperature (Suder et

263 the contours of the response surface are graphed. 314315 alof., Jabuticaba 2009), and (Myrciaria the optimal cauliflora fermentation) was 20 temperature◦C (Duarte 4

263264 theIn the contours contour of graph, the response the constant surface response are graphed. lines 315316 ofet Jabuticaba al., 2011). (Myrciaria Moreover, cauliflora the) conditions was 20◦C (Duarte of pH, 4

264 316317 et al., 2011). Moreover, the conditions of pH, 265 Inare the graphed contour in graph, the x1, thex2 constantplane. response Each contour lines temperature, and even the concentration of sugar that

265266 arecorresponds graphed to in a the particularx1, x2 heightplane. of Each the response contour 317318 temperature,was predicted and to be even optimal the concentration by the model, of were sugar similar that

266267 corresponds to a particular height of the response 318319 wasto those predicted found to in be the optimal preparation by the model,of wine. were The similar pH of surface (Montgomery, 2010). In Fig. 1b, it can

267 319320 to those found in the preparation of wine. The pH of 268 surfacebe observed (Montgomery, that the maximum 2010). overall In Fig. acceptance 1b, it can is grapes must vary between 3.0 and 3.9.

320 268269 befound observed at the center that the of themaximum circle, at overall 54 h of acceptance fermentation is 321 grapesResults must obtained vary between in this 3.0 study and are3.9. innovative since

269270 foundand with at the 13% centerpanela of the. circle, at 54 h of fermentation 321322 sugarResults contents, obtained to be in containedthis study are in the innovative drink for since its

270 and with 13% panela. 322323 sugar contents, to be contained in the drink for its 271 Figure 2a illustrates the response surface between acceptance, was unknown (both ends, few, or too

323324 acceptance, was unknown (both ends, few, or too 271272 theFigure fermentation 2a illustrates temperature the response andsurface the pH. between It can many sugars are unpleasant). Similarly, as to the

324 272273 thebe observed fermentation that temperature the overall acceptance and the pH. decreased It can 325 manyfermentation sugars time are unpleasant). concerns, tepache Similarly, producers as to quite the

273 325326 fermentationoften use as far time as concerns,96 h; however, tepache we producersfound that quite large 274 bewith observed low temperature that the overall and high acceptance pH of the decreased drink;

326327 often use as far as 96 h; however, we found that large 274275 withmoreover, low fortemperature high temperature and high and pHlow pH,of the the overall drink; fermentation times are not acceptable to consumers.

5 327 275276 moreover,acceptance for decreased high temperature even more. and low This pH, behavior the overall can fermentation times are not acceptable to consumers. 5 276277 acceptancebe explained decreased in terms even of more. the linear This ebehaviorffect of can the 328 3.2 Products formation of fermentation and 277278 be explained in terms of the linear effectff of the fermentation temperature and the quadratic e ect of 328 3.2 Products formation of fermentation and 329 sensory evaluation 278279 fermentationthe pH on the temperature overall acceptance and the of quadratic the tepache. effect The of 329 sensory evaluation 279 280 thecontour pH on graph the overall (Fig. acceptance2b) shows of that the the tepache. maximum The 330 In Table 1 is shown the results of the sensory analysis, 280 281 contouracceptance graph is found (Fig. between 2b) shows 22◦C that and the 25◦C, maximum with an 330331 IninTable addition 1 is to shown the final the concentration results of the of sensory sugars, analysis, ethanol 281 282 acceptanceinitial pH of is 3.5 found to 5.0. between 22◦C and 25◦C, with an 331332 inand addition acids. to the More final than concentration half ofthe of sugars, combinations ethanol 282 283 initialRSM pH ofisa 3.5 procedure to 5.0. that allows a general idea of 332333 andgot meanacids. values More greater than half than of 5 the (neither combinations like nor 283284 theRSM optimal is a conditions procedure tothat be allows rapidly a general and effi ideaciently of 333334 gotdislike); mean among values those, greater five than obtained 5 (neither values like greater nor 284285 theobtained optimal (Duarte conditionset al., to 2011). be rapidly The and levels effi ofciently each 334335 dislike);than 7 (like among moderately). those, five The obtained best score values was greater 7.73 285286 obtainedfactor estimated (Duarte byet the al., statistical 2011). The model levels (Equation of each 2) 335336 thana value 7 (like close moderately). to “like very The much” best scoreon the was hedonic 7.73 286287 factorfor the estimated optimization by the of statistical the overall model acceptance (Equation of the 2) 336337 ascale. value The close fermentation to “like very conditions much” onfor the this hedonic tepache 287288 fortepache the optimization were a temperature of the overall of 22◦C, acceptance 13% panela of, the an 337338 scale.were 10% Thepanela fermentation, an initial conditions pH of 5.0, for a fermentationthis tepache 288 tepache were a temperature of 22◦C, 13% panela, an 338 were 10% panela, an initial pH of 5.0, a fermentation www.rmiq.org 5 www.rmiq.orgwww.rmiq.org 235 Corona-Gonz´alezetCorona-Gonz´alezetCorona-Gonz´alezet al. al. al.//RevistaRevista/ Revista Mexicana Mexicana Mexicana de deI deIngenierngenierIngenier´ıa´ıaQQ´ıauuQ´ımica´ımicau ´ımica Vol. Vol. Vol. 12, 12, 12, No. No. No. 1 1 (2013) (2013) 1 (2013) XXX-XXX XXX-XXX 19-28

/ 339 Corona-Gonz´alezet al. Revista Mexicana deIngenier374 ´ıaQu ´ımica Vol. 12, No. 1 (2013) XXX-XXX 339 temperaturetemperature of of 22 22◦◦C,C, and and a a fermentation fermentation time time of of 72 72 374 fermentativefermentative production production of of ethanol, ethanol, are are temperature, temperature, 340 375 340 h.h. It It can can be be observed observed (Table (Table 1) 1) that, that, in in general, general, 375 initialinitial pH pH and and fermentation fermentation time. time. Although Although statistical statistical 339341 374376 ff 341 temperaturethethe concentrations concentrations of 22◦C, of of and lactic lactic a fermentation acid acid and and ethanol ethanol time of were were 72 376 fermentativeanalysisanalysis of of this this production study study showed showed of ethanol, no no significant significantare temperature, e effectect 340342 375377 342 h.greatergreater It can with with be prolonged prolonged observed fermentation fermentation (Table 1) that, times times in (96 (96 general, h). h). In In 377 initialofof temperature temperature pH and fermentation on on ethanol ethanol production production time. Although in in the the statistical tepache, tepache, 341343 376378 343 thecontrast,contrast, concentrations over over short short of fermentation fermentation lactic acid periods periods and ethanol (48 (48 h), h), were the the 378 analysiswewe found found of that that this time, time, study the the showed initial initial no pH, pH, significant and and additional additional effect 342344 377379 ff 344 greaterconcentrationsconcentrations with prolonged of of these these fermentation products, products, mainly mainly times of of (96 ethanol, ethanol, h). In 379 ofbrownbrown temperature sugar sugar concentration, concentration, on ethanol production did did a affectect in the the the response tepache,response 343345 378380 345 contrast,werewere lower. lower. over In In shortmost most treatments, treatments,fermentation the the periods acetic acetic acid acid (48 content contenth), the 380 weofof this this found variable. variable. that time, Problems Problems the initial caused caused pH, by by and the the additional retention retention 344346 / 379381 346 concentrationsdiddid not not exceed exceed of 5 5 gthese g/L,L, regardless regardless products, of of mainly the the fermentation fermentation of ethanol, 381 brownoror loss loss sugar of of alcohol alcohol concentration, and and aromatic aromatic did aff substances substancesect the response at at low low 345347 380382 347 weretime.time. lower. The The amount amountIn most treatments,of of residual residual the sugars sugars acetic in in acid the the tepachecontent tepache 382 ofandand this high high variable. temperatures, temperatures, Problems respectively, respectively, caused by can can the also also retention occur, occur, 346348 ff 381383 ff 348 didwaswas not also also exceed di differenterent 5 g for for/L, each each regardless treatment. treatment. of the However, However, fermentation there there 383 oraaffectingecting loss of the the alcohol optimal optimal and temperature temperature aromatic for forsubstances the the growth growth atlow and and 347349 382384 349 time.waswas a a The tendency tendency amount for for of thethe residual nearly nearly sugars total total in consumption, consumption, the tepache 384 andphysiologicalphysiological high temperatures, activity activity of of respectively, the the yeast yeast (Benda,can (Benda, also 1984); occur,1984); 348350 383385 ff 350 wasofof sucrose sucrose also diff aterent at 96 96 for h h ofeach of fermentation, fermentation, treatment. However, regardless regardless there of of 385 afurthermore,furthermore,ffecting the optimal the the lactic lactic temperature acid acid production production for the was was growth a affectedected and 349351 384386 351 wasthethe initial initial a tendency amount amount for of of thepanelapanela nearly.. Intotal In the the consumption, majority majority of of 386 physiologicalbyby the the fermentation fermentation activity temperature, temperature, of the yeast the the initial initial (Benda, amount amount 1984); of of 350352 / 385387 < . 352 oftreatments,treatments, sucroseat the the 96 presence presence h of fermentation, of of fructose fructose and and regardless/oror glucose glucose of 387 furthermore,brownbrown sugar sugar and andthe the lacticthe fermentation fermentation acid production time time ( (PP was< 00 a.05)ff05)ected but but 351353 386388 > . 353 thewaswas initial found, found, amount even even with with of prolonged prolongedpanela. Infermentation fermentation the majority times. times. of 388 bynotnot the by by fermentation the the initial initial pH pH temperature, ( (PP > 00.05).05). the Figureinitial Figure amount 3b 3b shows shows of 352354 387389 < . 354 treatments,TheThe concentrations concentrations the presence measured measured of fructose for for the the and treatment treatment/or glucose with with 389 brownthethe response response sugar and surface surface the fermentation for for lactic lactic acid; acid; time note( noteP that0 that05) only only but 353355 = = 388390 ffi > . 355 wasthethe greatest greatest found, evenacceptance acceptance with prolonged (T (T= 2222◦◦C,C, fermentationpanelapanela= 10%,10%, times. pH pH 390 notthethe bylinear linear the coe coe initialfficientscients pH ( ofP of percentage percentage0 05). Figure of of brown brown 3b shows sugar sugar 354356 = = / 389391 ff 356 The= 5,5, andconcentrations and time time= 7272 h) h) measured were were (g (g/L):L): for 7.30 7.30 the of oftreatment ethanol, ethanol, with5.25 5.25 391 theandand response the the fermentation fermentation surface temperature temperaturefor lactic acid; a affectect note the the thatresponse, response, only 355357 390392 357 theofof lacticlactic greatest acid, acid, acceptance 4.16 4.16 of of (Tacetic acetic= 22 acid, acid,◦C, panela 67.63 67.63= of of10%, sucrose, sucrose, pH 392 theandand linear that that the the coe concentrationffi concentrationcients of percentage of of lactic lactic of acid acid brown is is directly directly sugar 356358 391393 358 =4.484.485, and of of glucose, glucose, time = 72 and and h) 1.09 1.09 were fructose. fructose. (g/L): 7.30 of ethanol, 5.25 393 andproportionalproportional the fermentation to to the the increase increase temperature the the percentage percentage affect the response,of of brown brown 357359 392394 359 of lacticFigureFigure acid, 3 3 shows shows 4.16 of graphs graphs acetic of of acid, response response 67.63 surfaces surfaces of sucrose, for for 394 andsugarsugar that and and the temperature. temperature. concentration Figure Figure of lactic 3c 3c shows shows acid is the the directly linear linear 358360 393395 ff 360 4.48productionproduction of glucose, of of ethanol, ethanol, and 1.09 lactic lacticfructose. acid acid and and acetic acetic acid acid 395 proportionaleeffectect of of time time to and and the fermentation fermentation increase the temperature temperature percentage of (the (the brown only only 359361 394396 361 inin tepache. tepache.Figure 3 Alcoholicshows Alcoholic graphs fermentation fermentation of response is is the thesurfaces result result for of of 396 sugarsignificantsignificant and factors)temperature. factors) on on the the productionFigure production 3c shows of of acetic acetic the acid. acid. linear At At 360362 395397 362 productionmanymany interactions; interactions; of ethanol, not not lacticonly only depends depends acid and on on acetic the the strain strain acid 397 eshortershorterffect of fermentation fermentation time and fermentation times times and and lower lower temperature temperatures, temperatures, (the only the the 361363 396398 363 inbutbut tepache. also also on onphysicochemical Alcoholicphysicochemical fermentation environmental environmental is the result factors, factors, of 398 significantamountamount of of thisfactors) this acid acid on is is reduced; reduced;the production likewise, likewise, of acetic as as reported reported acid. Atby by 362364 397399 364 manysugars,sugars, interactions; acidity, acidity, temperature temperature not only and and depends others. others. on The The the ethanol ethanol strain 399 shorterSuderSuder etet fermentation al al.. (2009), (2009), times acetic acetic and acid acid lower concentration concentration temperatures, shows shows the 363365 398400 365 butcontentcontent also can can on impact impactphysicochemical on on the the perception perception environmental of of the the “body”, factors,“body”, 400 amountvariationvariation of with with this temperature.acid temperature. is reduced; likewise, as reported by 364366 399401 366 sugars,thethe viscosity viscosity acidity, and, and, temperature to to a a lesser lesser and extent, extent, others. on on the the The sweetness, sweetness, ethanol 401 SuderWithWithet al the. the (2009), data data acetic obtained obtained acid concentration from from the the sensory sensory shows 365367 400402 367 contentacidity,acidity, aroma,can aroma, impact flavor flavor on intensity intensity the perception and and textural textural of the properties properties “body”, 402 variationevaluationsevaluations with and and temperature. the the final final concentration concentration of of ethanol, ethanol, 366368 401403 368 the(Suder(Suder viscosityetet al al.,., and, 2009). 2009). to a lesser Figure Figure extent, 3a 3a shows shows on the the the sweetness, quadratic quadratic 403 acids,acids,With and and the sugars, sugars, data obtained multivariate multivariate from analysis analysis the sensory was was 367369 ff ff 402404 369 acidity,eeffectect of of aroma, pH pH and and flavor the the intensitylinear linear e effect andect of of textural brown brown properties sugar sugar on on 404 evaluationsperformedperformed to to and correlate correlate the final the the concentration concentrations concentrations of of ofethanol, these these 368370 403405 370 (Suderthethe amount amountet al of of., ethanol2009). ethanol production; production;Figure 3a shows at at pH pH values the values quadratic near near 5, 5, 405 acids,componentscomponents and in in sugars, the the tepache tepache multivariate with with the the analysis degree degree of of was the the 369371 404406 371 eethanolethanolffect of levels levels pH and are are lowerthe lower linear than than eat atff higherect higher of pHbrown pH values values sugar below below on 406 performeddrink’sdrink’s acceptance. acceptance. to correlate Table Table the concentrations 3 3 shows shows the the of Pearson Pearson these

370372 405407 ffi 372 the5.5. amountRatman Ratman ofetet ethanol al al.. (2003, (2003, production; 2005) 2005) using using at pH a values a CCD CCD near found found 5, 407 componentsproduct-momentproduct-moment in the correlation correlation tepache coewith coefficients thecients degree (r) (r) for for of each each the

371373 ff 406408 373 ethanolthatthat the the levels most most areimportant important lower than physical physical at higher factors, factors, pH values which which below a affectect 408 drink’scompound,compound, acceptance. with with overall overall Table acceptance acceptance 3 shows of of the the the tepache. tepache. Pearson

372 5. Ratman et al. (2003, 2005) using a CCD found 407 product-moment correlation coefficients (r) for each

373 that the most important physical factors, which affect 408 compound, with overall acceptance of the tepache.

409 409 410 Fig 3 410 Fig.FigFig. 3 3. 3. Response Response surface surface for for production production of of (a) (a) Ethanol, Ethanol, (b) (b) Lactic Lactic acid, acid, and and (c) (c) Acetic Acetic acid. acid. 409 410 Fig.Fig 3 3. Response surface for production of (a) Ethanol, (b) Lactic acid, and (c) Acetic acid.

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6 Corona-Gonz´alezetCorona-Gonz´alezet al. al.//RevistaRevista Mexicana Mexicana de deIIngenierngenier´ıa´ıaQQuu´ımica´ımica Vol. Vol. 12, 12, No. No. 1 1 (2013) (2013) XXX-XXX XXX-XXX Corona-Gonz´alezet al./ Revista Mexicana deIngenier ´ıaQu ´ımica Vol. 12, No. 1 (2013) XXX-XXX

Corona-Gonz´alezet al./ Revista Mexicana deIngenier ´ıaQu ´ımica Vol. 12, No. 1 (2013) 19-28 TableTable 2. 2. Correlation Correlation for for acceptance acceptance AceticAcetic acid acidTable Lactic Lactic 2. Correlation acid acid Ethanol Ethanol for acceptance Fructose Fructose Glucose Glucose Sucrose Sucrose 411411 rrAcetic -0.2487 -0.2487 acid Lactic -0.0004 -0.0004 acid Ethanol -0.3643 -0.3643 Fructose 0.4105 0.4105 Glucose 0.5240 0.5240 Sucrose 0.5624 0.5624 411 P-value 0.0672 0.9975 0.0063 0.0019 0.0000 0.0000 P-valuer -0.2487 0.0672 -0.0004 0.9975 -0.3643 0.0063 0.4105 0.0019 0.5240 0.0000 0.5624 0.0000 P-value 0.0672 0.9975 0.0063 0.0019 0.0000 0.0000

412412 TheThe range range of of these these correlation correlation coe coeffifficients,cients, ranging ranging 454454 productsproducts of of fermentation fermentation is is very very high, high, leading leading to to a a 413413 fromfrom 11 to to ++1,1, measures measures the the strength strength of of the the 455455 decreasedecrease in in the the acceptance acceptance of of the the tepache tepache after after long long 412 The range−− of these correlation coefficients, ranging 454 products of fermentation is very high, leading to a 414414 linearlinear relationship relationship between between variables. variables. A A correlation correlation 456456 fermentationfermentation times times ( (>> 7272 h). h). 413 from 1 to +1, measures the strength of the 455 decrease in the acceptance of the tepache after long 415 415 ofof ++11− means means that that there there is is a a perfect perfect direct direct linear linear 457 414 linear relationship between variables. A correlation 456457 fermentationTheThe chemical chemical times ( composition> composition72 h). of of the the tepache tepache was was 416 416 relationshiprelationship (positive) (positive) between between the the two two variables, variables, 458 415 458 aaffffectedected by by the the fermentation fermentation conditions, conditions, given given that that of +1 means that there is a perfect direct linear 457 417 The chemical composition of the tepache was 417 whilewhile a a correlation correlation of of 11 means means that that there there is is a a perfect perfect 459 416 459 manymany biosynthetic biosynthetic routes routes adopted adopted by by yeasts yeasts and and relationship (positive)−− between the two variables, 458 418 affected by the fermentation conditions, given that 418 inverseinverse linear linear relationship relationship (negative) (negative) between between the the two two 460 417 460 bacteriabacteria are are related related to to the the formation formation of of the the aroma aroma while a correlation of 1 means that there is a perfect 459 419 many biosynthetic routes adopted by yeasts and 419 variables.variables. Finally, Finally, a a− correlation correlation of of 0 0 is is interpreted interpreted 461 418 461 andand flavor flavor and and are are a affffectedected by by various various factors, factors, inverse linear relationship (negative) between the two 460 420 bacteria are related to the formation of the aroma 420 asas the the absence absence of of a a linear linear relationship relationship between between two two 462 419 462 suchsuch as as the the composition, composition, the the pH pH of of the the medium, medium, variables. Finally, a correlation of 0 is interpreted 461 421 and flavor and are affected by various factors, 421 variablesvariables (Lincoln, (Lincoln, 1985). 1985). 463 420 as the absence of a linear relationship between two 463 concentrationconcentration of of sugar, sugar, dissolved dissolved oxygen oxygen and and the the 462 such as the composition, the pH of the medium, 464 421422422 variablesIt can (Lincoln, be observed 1985). that the sugars exerted a great 464 prevailingprevailing temperature temperature of of the the fermentation fermentation (Swiers (Swiers It can be observed that the sugars exerted a great 463 concentration of sugar, dissolved oxygen and the 465 423423 dealdeal of of influence influence in in their their linear linear relationship relationship with with 465 etet al al.,., 2005; 2005; Estela-Escalante Estela-Escalante etet al al.,., 2012; 2012; T Tellez-´ellez-´ 422 It can be observed that the sugars exerted a great 464 prevailing temperature of the fermentation (Swiers 466 424424 acceptance.acceptance. The The table table also also shows shows the the P-value P-value that that 466 MoraMora etet al al.,., 2012). 2012). Other Other authors authors have have already already 423 deal of influence in their linear relationship with 465 et al., 2005; Estela-Escalante et al., 2012; Tellez-´ 467 425425 establishesestablishes the the statistical statistical significance significance of of the the estimated estimated 467 reportedreported (Duarte (Duarte etet al al.,., 2011; 2011; Swiers Swiers etet al al.,., 2005) 2005) 424 acceptance. The table also shows the P-value that 466 Mora et al., 2012). Other authors have already 468 426426 correlations.correlations. P-values P-values below below 0.05 0.05 indicate indicate correlations correlations 468 thatthat the the fermentation fermentation temperature, temperature, the the initial initial sugar sugar 425 establishes the statistical significance of the estimated 467 reported (Duarte et al., 2011; Swiers et al., 2005) 469 ff 427427 thatthat are are significantly significantly di difffferenterent from from zero, zero, with with a a 469 concentration,concentration, and and the the pH pH significantly significantly a affectect the the 426 correlations. P-values below 0.05 indicate correlations 468 that the fermentation temperature, the initial sugar 470 ff 428428 confidenceconfidence level level of of 95.0%. 95.0%. The The ethanol, ethanol, sucrose, sucrose, 470 chemicalchemical composition composition of of di differenterent alcoholic alcoholic drinks. drinks. 427 that are significantly different from zero, with a 469 concentration, and the pH significantly affect the 429 429 glucose,glucose, and and fructose fructose have have shown shown some some correlation correlation 471471 The temperature affffects the yeast and as a result 428 confidence level of 95.0%. The ethanol, sucrose, 470 chemicalThe temperature composition a ofects different the yeast alcoholic and asdrinks. a result 430 430 withwith overall overall acceptance acceptance of of the the tepache. tepache. These These results results 472472 considerablyconsiderably a affffectsects the the course course of of the the fermentation. fermentation. 429 glucose, and fructose have shown some correlation 471 The temperature affects the yeast and as a result 431 431 showshow the the type type of of relationship relationship between between the the final final 473473 TheThe death death of of yeast yeast at at high high temperatures temperatures causes causes 430 with overall acceptance of the tepache. These results 472 considerably affects the course of the fermentation. 432 432 concentrationconcentration of of sugar sugar and and ethanol ethanol on on the the acceptance acceptance 474474 thethe fermentation fermentation to to stop, stop, which which is is accompanied accompanied 431 show the type of relationship between the final 473 The death of yeast at high temperatures causes 433 433 ofof the the tepache. tepache. In In some some drinks, drinks, the the sensation sensation of of 475475 byby the the danger danger of of contamination contamination by by thermophilic thermophilic 432 concentration of sugar and ethanol on the acceptance 474 the fermentation to stop, which is accompanied 434 434 acidityacidity is is perceived perceived less less frequently frequently when when the the products products 476476 microorganisms.microorganisms. However, However, low low temperatures temperatures may may 433 of the tepache. In some drinks, the sensation of 475 by the danger of contamination by thermophilic 435 435 areare sweeter. sweeter. For For this this reason, reason, some some producers producers of of 477477 produceproduce problems problems in in delaying delaying the the beginning beginning of of 434 acidity is perceived less frequently when the products 476 microorganisms. However, low temperatures may 436 436 tepachetepache often often compensate compensate for for the the high high degree degree of of 478478 fermentation.fermentation. In In addition, addition, to to the the optimal optimal temperature temperature 435 are sweeter. For this reason, some producers of 477 produce problems in delaying the beginning of 437 437 fermentationfermentation with with the the addition addition of of sugar, sugar, with with the the goal goal 479479 forfor the the growth growth and and physiological physiological activity activity of of the the yeast, yeast, 436 tepache often compensate for the high degree of 478 fermentation. In addition, to the optimal temperature 438 438 ofof decreasing decreasing the the sensation sensation of of acidity, acidity, given given their their 480480 problemsproblems caused caused by by the the retention retention or or loss loss of of alcohol alcohol 437 fermentation with the addition of sugar, with the goal 479 for the growth and physiological activity of the yeast, 439 439 extensiveextensive experience experience in in balancing balancing the the flavor flavor of of the the 481481 andand aromatic aromatic substances substances at at low low and and high high temperatures, temperatures, 438 of decreasing the sensation of acidity, given their 480 problems caused by the retention or loss of alcohol 440 440 drinkdrink (Moreno-Terrazas, (Moreno-Terrazas, 2005). 2005). However, However, the the ethanol ethanol 482482 respectively,respectively, can can also also occur occur (Benda, (Benda, 1984). 1984). 439 extensive experience in balancing the flavor of the 481 and aromatic substances at low and high temperatures, 441441 content may impact the perception of the “body”, the content may impact the perception of the “body”, the 483 440 drink (Moreno-Terrazas, 2005). However, the ethanol 482483 respectively,TheThe pH pH influencescan influences also occur the the growth(Benda, growth of of 1984). microorganisms microorganisms 442442 viscosity,viscosity, and and to to a a lesser lesser degree degree the the sweetness, sweetness, acidity, acidity, 441 content may impact the perception of the “body”, the 484484 andand their their metabolism, metabolism, which which is is frequently frequently modified modified 483 443443 aroma,aroma, flavor flavor intensity, intensity, and and textural textural properties properties (Suder (Suder The pH influences the growth of microorganisms 442 viscosity, and to a lesser degree the sweetness, acidity, 485485 asas a a result result of of a a change change in in pH pH of of up up to to 1-1.5 1-1.5 (Scragg, (Scragg, 484 444444 etet al al.,., 2009). 2009). and their metabolism, which is frequently modified 443 aroma, flavor intensity, and textural properties (Suder 486486 2009).2009). Some Some studies studies mention mention that that the the growth growth of of 485 as a result of a change in pH of up to 1-1.5 (Scragg, 444445445 et al.,TheseThese 2009). results results can can be be explain explain the the linear linear and and 487487 aceticacetic bacteria bacteria during during fermentation fermentation is is correlated correlated with with 486 2009). Some studies mention that the growth of 446446 quadraticquadratic e effffectect of of time time on on the the overall overall acceptance acceptance 488488 thethe initial initial pH pH of of the the medium. medium. When When the the pH pH of of 445 These results can be explain the linear and 487 acetic bacteria during fermentation is correlated with 447447 ofof the the tepache, tepache, given given that that in in the the initial initial stages stages of of 489489 thethe must must is is relatively relatively high, high, the the amount amount of of acetic acetic 446 quadratic effect of time on the overall acceptance 488 the initial pH of the medium. When the pH of 448448 fermentationfermentation ( (<< 4848 h), h), the the sugar sugar remains remains practically practically 490490 bacteriabacteria is is greater greater at at the the end end of of fermentation, fermentation, which which 447 of the tepache, given that in the initial stages of 489 the must is relatively high, the amount of acetic 449449 unconsumedunconsumed and and the the compounds compounds (lactic (lactic acid acid and and 491491 isis an an indication indication that that these these bacteria bacteria may may grow grow during during 448 fermentation (< 48 h), the sugar remains practically 490 bacteria is greater at the end of fermentation, which 450450 ethanol)ethanol) that that provide provide the the tepache tepache with with better better sensory sensory 492492 alcoholicalcoholic fermentation fermentation (Escalante (Escalante etet al al.,., 2004). 2004). The The 449 unconsumed and the compounds (lactic acid and 491 is an indication that these bacteria may grow during 451451 characteristicscharacteristics are are not not yet yet formed. formed. However, However, with with 493493 presencepresence of of acetic acetic bacteria bacteria leads leads to to souring, souring, brown brown 450 ethanol) that provide the tepache with better sensory 492 alcoholic fermentation (Escalante et al., 2004). The 452452 prolongedprolonged fermentation fermentation times, times, the the sugars sugars are are almost almost 494494 discoloration,discoloration, a a bittersweet bittersweet taste, taste, and and turbidity turbidity of of 451 characteristics are not yet formed. However, with 493 presence of acetic bacteria leads to souring, brown 453453 entirelyentirely consumed, consumed, and and the the concentration concentration of of the the 495495 fermentedfermented drinks drinks due due to to the the production production of of acetic acetic acid acid 452 prolonged fermentation times, the sugars are almost 494 discoloration, a bittersweet taste, and turbidity of 453 entirely consumed, and the concentration of the 495 fermented drinks due to the production of acetic acid www.rmiq.orgwww.rmiq.org 77 www.rmiq.org 7

www.rmiq.org 25 Corona-Gonz´alezetCorona-Gonz´alezetCorona-Gonz´alezet al. al. al.//RevistaRevista/ Revista Mexicana Mexicana Mexicana de deI deIngenierngenierIngenier´ıa´ıaQQ´ıauuQ´ımica´ımicau ´ımica Vol. Vol. Vol. 12, 12, 12, No. No. No. 1 1 (2013) (2013) 1 (2013) XXX-XXX XXX-XXX 19-28

496496 (Manca(Manca etet al al.,., 2006). 2006). Concentrations Concentrations of of 0.3-1.1 0.3-1.1 g g//LL 546546 experienceexperience a a decrease decrease in in their their sensitivity sensitivity to to ethanol, ethanol, 497497 maymay become become undesirable, undesirable, depending depending on on the the type type of of 547547 providingproviding a a significant significant contribution contribution to to the the taste taste of of the the 498498 drinkdrink (Swiers (Swiers etet al al.,., 2005; 2005; Flores-Ramirez Flores-Ramirez etet al al.,., 548548 winewine (Romano (Romanoetet al al.,., 2006). 2006).

499499 2005).2005). 549549 YeastsYeasts of of the the genera genera CandidaCandida andand HanseniasporaHanseniaspora 500500 InformationInformation obtained obtained in in this this study study is is important important 550550 havehave also also been been found found in in other other natural natural fermentations. fermentations. 501501 becausebecause it it was was previously previously expected expected that that a a higher higher 551551 ItIt was was reported reported the the presence presence of of these these genera genera in in 502502 alcoholalcohol content content and and lactic lactic acid acid would would have have a a greater greater 552552 fermentationsfermentations of of pineapple pineapple juice; juice; even even the the species species 503503 acceptanceacceptance among among consumers consumers and and did did not. not. On On the the 553553 C.C. apicola apicola waswas identified, identified, although although only only occasionally occasionally 504504 otherother hand, hand, it it was was found found that that for for a a fermentation fermentation time time 554554 andand at at a a very very low low concentrations concentrations during during fermentation fermentation 505505 longerlonger than than 72 72 h, h, the the acceptance acceptance of of the the beverage beverage is is 555555 (Chanprasartsuk(Chanprasartsuk etet al al.,., 2010). 2010). However, However, the the genus genus 506506 reducedreduced since since the the lower lower concentration concentration of of sugars sugars and and 556556 HanseniasporaHanseniaspora waswas one one of of the the most most frequently frequently 507507 higherhigher contents contents of of alcohol alcohol and and acetic acetic acid. acid. 557557 foundfound genres genres during during the the entire entire fermentation fermentation process process 558558 ofof pineapple pineapple juice. juice. Moreover, Moreover, although although yeasts yeasts 559559 ofof the the genus genus CryptococcusCryptococcus werewere not not found found during during 508508 3.33.3 Identification Identification of of yeast yeast 560560 fermentation,fermentation, they they were were isolated isolated and and identified identified from from

561561 the pineapple rind (Chanprasartsuk et al., 2010); the 509509 AA total total of of 33 33 colonies colonies were were obtained, obtained, from from which which DNA DNA the pineapple rind (Chanprasartsuk et al., 2010); the 562562 presence of this yeast may be associated with the 510510 extractionextraction was was performed. performed. SaccharomycesSaccharomyces cerevisiae cerevisiae presence of this yeast may be associated with the 563563 pineapple rind used. 511511 waswas the the predominant predominant species species (70% (70% of of the the total total of of pineapple rind used. 564564 However, S. cerevisiae was the only species 512512 thethe identified identified strains) strains) and and was was found found practically practically in in However, S. cerevisiae was the only species 565565 isolated in different samples it was found at least one 513513 allall of of the the analyzed analyzed samples; samples; for for some some samples, samples, it it isolated in different samples it was found at least one 566566 different species. The variation in the species found 514514 waswas the the only only species species that that was was isolated. isolated. The The other other different species. The variation in the species found 567567 could be attributed to the fermentation conditions, the 515515 speciesspecies were were as as follows: follows: CandidaCandida apicola apicola (9%),(9%), could be attributed to the fermentation conditions, the 568568 fermentation stage in which the sample was taken, 516516 CryptococcusCryptococcus skinneri skinneri (9%),(9%), HanseniasporaHanseniaspora (6%),(6%), fermentation stage in which the sample was taken, 569569 and/or the natural flora of the skins of pineapple and 517517 andand SaccharomycesSaccharomyces sp.sp. (6%). (6%). The The species species of of the the and/or the natural flora of the skins of pineapple and 570570 brown sugar concentration. 518518 generageneraHanseniasporaHanseniasporawerewere not not di difffferentiatederentiated because because brown sugar concentration. 519519 withwith these these enzymes, enzymes, it it is is not not possible possible to to do do so so and and 520520 wouldwould require require further further digestion digestion with with an an additional additional 571 521521 enzymeenzyme ( (DdeDde I).I). In In studies studies performed performed on on di difffferenterent 571 ConclusionConclusion 522522 fermentedfermented fruit fruit products, products, it it has has been been reported reported that that the the 572572 With the response surface methodology (RSM) it was 523523 speciesspecies S.S. cerevisiae cerevisiae isis predominant predominant during during the the final final With the response surface methodology (RSM) it was 573573 possible to correlate the fermentation products with 524524 stagesstages of of fermentation. fermentation. It It has has also also been been mentioned mentioned that that possible to correlate the fermentation products with 574574 the degree of acceptance of the beverage, and to 525525 inin not not inoculated inoculated fermentations, fermentations, the the final final product product is is the degree of acceptance of the beverage, and to 575575 determine the optimal conditions for fermentation of 526526 thethe result result of of the the combined combined activity activity of of various various strains strains determine the optimal conditions for fermentation of 576576 tepache for a wider acceptance among consumers. 527527 ofof yeast yeast that that grow grow more more or or less less successively successively over over tepache for a wider acceptance among consumers. 577577 With the graphs of contours, it is possible to 528528 thethe course course of of the the fermentation fermentation (Gonz (Gonzalez-Hern´alez-Hern´ andez´andez´ With the graphs of contours, it is possible to 578578 predict the fermentation conditions to achieve that 529529 etet al al.,., 2012). 2012). The The first first stages stages of of fermentation fermentation predict the fermentation conditions to achieve that 579579 desired levels of ethanol, acetic acid, lactic acid 530530 areare dominated dominated by by non- non-SaccharomycesSaccharomyces yeast,yeast, whose whose desired levels of ethanol, acetic acid, lactic acid 580580 and residual sugars. The optimum conditions for 531531 growthgrowth rapidly rapidly declines declines due due to to its its sensitivity sensitivity to to and residual sugars. The optimum conditions for 581581 fermentation of tepache, predicted by quadratic model, 532532 ethanol;ethanol; subsequently, subsequently, the the S.S. cerevisiae cerevisiae strains,strains, which which fermentation of tepache, predicted by quadratic model, 582582 were: temperature 22◦C, brown sugar 13%, an initial 533533 areare more more tolerant tolerant to to ethanol, ethanol, begin begin to to dominate dominate and and were: temperature 22◦C, brown sugar 13%, an initial 583583 pH of 4, and 54 h of fermentation. The yeast 534534 areare the the yeast yeast strains strains that that complete complete the the wine wine production production pH of 4, and 54 h of fermentation. The yeast 584584 species identified in the difffferent treatments were: 535535 processprocess (Garde (Garde and and Anc Anc´ın,´ın, 2006). 2006). In In the the production production species identified in the di erent treatments were: 585585 Candida apicola (9%), Cryptococcus skinneri (9%), 536536 ofof wine, wine, the the non- non-SaccharomycesSaccharomyces yeastyeast present present in in Candida apicola (9%), Cryptococcus skinneri (9%), 586586 Hanseniaspora (6%), Saccharomyces sp. (6%) and 537537 grapegrape juice, juice, such such as as HanseniasporaHanseniaspora ((KloeckeraKloeckera)) Hanseniaspora (6%), Saccharomyces sp. (6%) and 587587 Saccharomyces cerevisiae. The latter, about 70%, was 538538 andand CandidaCandida,, begin begin to to experience experience a a decrease decrease in in Saccharomyces cerevisiae. The latter, about 70%, was 588588 the predominant species at the end of fermentation. 539539 theirtheir population population by by half half during during fermentation fermentation when when the predominant species at the end of fermentation. 540540 thethe ethanol ethanol produced produced by by S.S. cerevisiae cerevisiae exceedsexceeds 5- 5- 541541 7%.7%. The The production production of of ethanol ethanol by by S. S. cerevisiae cerevisiae 589589 AcknowledgmentsAcknowledgments 542542 isis the the factor factor that that most most greatly greatly a affffectsects the the growth growth 543543 ofof non- non-SaccharomycesSaccharomyces yeasts.yeasts. However, However, when when the the 590590 TheThe authors authors wish wish to to thank thank the the University University of of 544544 fermentationfermentation is is performed performed at at temperatures temperatures below below 591591 Guadalajara,Guadalajara, the the State State Research Research and and Assistance Assistance in in 545545 15-2015-20◦◦C,C, the the species species HanseniasporaHanseniaspora andand CandidaCandida 592592 TechnologyTechnology and and Design Design Center Center of of Jalisco, Jalisco, A.C. A.C.

2688 www.rmiq.orgwww.rmiq.orgwww.rmiq.org Corona-Gonz´alezetCorona-Gonz´alezetCorona-Gonz´alezet al. al./ al./RevistaRevista/ Revista Mexicana Mexicana Mexicana de deIngenierI dengenierIngenier´ıa´ıaQQ´ıauuQ´ımica´ımicau ´ımica Vol. Vol. Vol. 12, 12, 12, No. No. No. 1 1 (2013) (2013) 1 (2013) XXX-XXX XXX-XXX 19-28

593593 (Centro(Centro de de Investigaci Investigacion´on´ y y Asistencia Asistencia en en Tecnolog Tecnolog´ıa´ıa y y 637637 Estela-Escalante,Estela-Escalante, W., W., Rychtera, Rychtera, M., M., Melzoch, Melzoch, K. K. and and 594594 DiseDiseno˜no˜ del del Estado Estado de de Jalisco, Jalisco, A. A. C.), C.), and and the the National National 638638 Hatta-Sakoda,Hatta-Sakoda, B. B. (2012). (2012). E Effffectect of of aeration aeration 595595 CouncilCouncil on on Science Science and and Technology, Technology, CONACYT CONACYT 639639 onon the the fermentative fermentative activity activity of ofSaccharomycesSaccharomyces 596596 (Consejo(Consejo Nacional Nacional de de Ciencia Ciencia y y Tecnolog Tecnolog´ıa).´ıa). 640640 cerevisiaecerevisiae culturedcultured in in apple apple juice. juice. RevistaRevista 641641 MexicanaMexicana de de Ingenier´ıaQu´ımica11 Ingenier´ıaQu´ımica11,, 211-226. 211-226.

642642 Esteve-Zarzoso,Esteve-Zarzoso, B., B., Belloch, Belloch, C., C., Uruburu, Uruburu, F. F. and and 597597 ReferencesReferences 643643 Querol,Querol, A. A. (1999). (1999). Identification Identification of of yeast yeast 644644 byby RFLP RFLP analysis analysis of of the the 5.8S 5.8S rRNA rRNA gene gene 598598 Aidoo,Aidoo, K., K., Rob, Rob, M. M. and and Sarkar, Sarkar, P. P. (2006). (2006). 645645 andand the the two two ribosomal ribosomal internal internal transcribed transcribed 599599 OccurrenceOccurrence and and function function of of yeasts yeasts in in Asian Asian 646646 spacers.spacers. InternationalInternational Journal Journal of of Systematic Systematic 600600 indigenousindigenous fermented fermented foods. foods. FEMSFEMS Yeast Yeast 647647 BacteriologyBacteriology 49 49,, 329-337. 329-337. 601601 ResearchResearch 6 6,, 30-39. 30-39.

648648 Flores-RamFlores-Ram´ırez,´ırez, N., N., Mart Mart´ınez-Peniche,´ınez-Peniche, R. R. A., A., 602602 Alvarado,Alvarado, C., C., Garc Garc´ıa,´ıa, B., B., Martin, Martin, S. S. and and Regalado, Regalado, 649649 FernFernandez-Escart´andez-Escart´ ´ın,´ın, E., E., Gallegos-P Gallegos-Perez,´erez,´ J. J. L., L., 603603 C.C. (2006). (2006). Food-associated Food-associated lactic lactic acid acid bacteria bacteria 650650 DD´ıaz-Cervantes,´ıaz-Cervantes, M. M. I I and and V Vasquez-Garc´asquez-Garc´ ´ıa,´ıa, S. S. 604604 withwith antimicrobial antimicrobial potential potential from from traditional traditional 651651 R.R. (2005). (2005). Physical Physical and and chemical chemical properties properties in in 605605 MexicanMexican foods. foods. RevistaRevista Latinoamericana Latinoamericana de de 652652 redred wines: wines: traditional traditional vinification vinification and and carbonic carbonic 606606 Microbiolog´ıa48Microbiolog´ıa48,, 260-268. 260-268. 653653 macerationmaceration with with two twoSaccharomycesSaccharomyces cerevisiae cerevisiae 654654 strains.strains. RevistaRevista Mexicana Mexicana de de Ingenier´ıa Ingenier´ıa 607607 Benda,Benda, I. I. (1984). (1984). Wine Wine and and Brandy. Brandy. In: In: PrescottPrescott 655655 Qu´ımica4Qu´ımica4,, 289-297. 289-297. 608608 &&Dunn’sDunn’s industrial industrial microbiology microbiology,, (4th (4th edn) edn) 609609 (G.(G. Reed Reed ed.), ed.), Pp Pp 293-402. 293-402. Avi Avi publishing publishing 656656 Garde,Garde, T. T. and and Anc Anc´ın,´ın, C. C. (2006). (2006). Contribution Contribution of of wild wild 610610 company,company, Connecticut. Connecticut. USA. USA. 657657 yeastsyeasts to to the the formation formation of of volatile volatile compounds compounds in in 658658 inoculatedinoculated wine wine fermentations. fermentations.EuropeanEuropean Food Food 611611 Chanprasartsuk,Chanprasartsuk, O., O., Prakitchaiwattana, Prakitchaiwattana, C., C., 659659 ResearchResearch and and Technology Technology 222 222,, 15-25. 15-25. 612612 Sanguandeekul,Sanguandeekul, R. R. and and Fleet, Fleet, G. G. (2010). (2010).

613613 AutochthonousAutochthonous yeasts yeasts associated associated with with mature mature 660660 Godoy,Godoy, A., A., Herrera, Herrera, T. T. and and Ulloa, Ulloa, M. M. (2003). (2003). M´asM´as 614614 pineapplepineapple fruits, fruits, freshly freshly crushed crushed juice juice and and their their 661661 all´adelall´adel pulque pulque y y el el tepache tepache.. UNAM, UNAM, M Mexico.´exico.´ 615615 fermentsferments and and the the chemical chemical changes changes during during 616616 naturalnatural fermentation. fermentation. BioresourceBioresource Technology Technology 662662 GonzGonzalez-Hern´alez-Hern´ andez,´andez,´ J. J. C., C., P Perez,´erez,´ E., E., Dami Damian,´an,´ R. R. 617617 101101,, 7500-7509. 7500-7509. 663663 M.M. and and Ch Chavez,´avez,´ M. M. C. C. (2012). (2012). Isolation, Isolation, 664664 molecularmolecular and and fermentative fermentative characterization characterization of of a a

618618 Deshpande,Deshpande, R., R., Chinnan, Chinnan, M. M. and and Mc Mc Watters, Watters, 665665 yeastyeast used used in in ethanol ethanol production production during during mezcal mezcal 619619 K.K. (2008). (2008). Optimization Optimization of of a a chocolate- chocolate- 666666 elaboration.elaboration. RevistaRevista Mexicana Mexicana de de Ingenier´ıa Ingenier´ıa 620620 flavored,flavored, peanut-soy peanut-soy beverage beverage using using response response 667667 Qu´ımica11Qu´ımica11,, 389-400. 389-400. 621621 surfacesurface methodology methodology (RSM) (RSM) as as applied applied to to 668668 Lincoln,Lincoln, L. L. (1985). (1985). Regresi Regresion´on´ y y correlaci correlacion.´on.´ In: In: 622622 consumerconsumer acceptability acceptability data. data.LWTLWT Food Food Science Science 669669 Introducci´onaIntroducci´ona la la estad´ıstica estad´ıstica,, (L. (L. Lincoln Lincoln ed.), ed.), 623623 TechnologyTechnology 4 4,, 1485-1492. 1485-1492. 670670 PpPp 399-424. 399-424. CECSA, CECSA, M Mexico.´exico.´

624624 Duarte,Duarte, W. W. F., F., Amorim, Amorim, J. J. C., C., de de Assis Assis Lago, Lago, L., L., Dias, Dias, 671671 Manca,Manca, M., M., Strasser, Strasser, A. A. and and Arena, Arena, M. M. (2006). (2006). 625625 D.D. R, R, and and Schwan, Schwan, R. R. (2011). (2011). Optimization Optimization of of 672672 BebidasBebidas fermentadas. fermentadas. In: In:Microbiolog´ıadeMicrobiolog´ıade los los 626626 FermentationFermentation Conditions Conditions for for Production Production of of the the 673673 alimentosalimentos,, (M. (M. Torres, Torres, A. A. Castillo, Castillo, eds.), eds.), Pp Pp 627627 JabuticabaJabuticaba (Myrciariacauliflora) (Myrciariacauliflora) Spirit Spirit Using Using 674674 371-392.371-392. Universidad Universidad de de Guadalajara, Guadalajara, M Mexico.´exico.´ 628628 thethe Response Response Surface Surface Methodology. Methodology.JournalJournal of of

629629 FoodFood Science Science 76 76,, C782-C790. C782-C790. 675675 Montgomery,Montgomery, D. D. (2010). (2010). Dise˜noDise˜no y y An´alisis An´alisis de de 676676 ExperimentosExperimentos.. Limusa Limusa Wiley, Wiley, M Mexico.´exico.´ 630630 Escalante,Escalante, A., A., Rodr Rodr´ıguez,´ıguez, M., M., Mart Mart´ınez,´ınez, A., A., L Lopez-´opez-´ 631631 MunguMungu´ıa,´ıa, A., A., Bol Bol´ıvar,´ıvar, F. F. and and Gosset, Gosset, G. G. 677677 Moreno-Terrazas,Moreno-Terrazas, R. R. D. D. (2005). (2005). Determinaci´onDeterminaci´on 632632 (2004).(2004). Characterization Characterization of of bacterial bacterial diversity diversity 678678 dede las las caracter´ısticas caracter´ısticas microbiol´ogicas, microbiol´ogicas, 633633 inin pulque, pulque, a a traditional traditional Mexican Mexican alcoholic alcoholic 679679 bioqu´ımicas,bioqu´ımicas, fisicoqu´ımicas fisicoqu´ımicas y y sensoriales sensoriales 634634 fermentedfermented beverage, beverage, as as determined determined by by 16S 16S 680680 parapara la la estandarizaci´on estandarizaci´on del del proceso proceso de de 635635 rDNArDNA analysis. analysis. FEMSFEMS Microbiology Microbiology Letters Letters 681681 lala elaboraci´on elaboraci´on del del tepache tepache.. [PhD [PhD 636636 235235,, 273-279. 273-279. 682682 thesis].thesis]. Iztapalapa, Iztapalapa, M Mexico:´exico:´ Universidad Universidad

www.rmiq.orgwww.rmiq.orgwww.rmiq.org 2799 Corona-Gonz´alezetCorona-Gonz´alezetCorona-Gonz´alezet al. al.// al.RevistaRevista/ Revista Mexicana Mexicana Mexicana de deIIngenier dengenierIngenier´ıa´ıaQQ´ıauuQ´ımica´ımicau ´ımica Vol. Vol. Vol. 12, 12, 12, No. No. No. 1 1 (2013) (2013) 1 (2013) XXX-XXX XXX-XXX 19-28

683683 AutAutonoma´onoma´ Metropolitana. Metropolitana. Available Available 715715 Romano,Romano, P., P., Capece, Capece, A. A. and and Jespersen, Jespersen, L. L. (2006). (2006). 684684 from:from: http: http:////tesiuami.izt.uam.mxtesiuami.izt.uam.mx//uamuam//aspuamaspuam// 716716 TaxonomicTaxonomic and and ecological ecological diversity diversity of of food food 685685 presentatesis.php?recnopresentatesis.php?recno==11992&docs11992&docs==UAMIUAMI 717717 andand beverage beverage yeasts. yeasts. In: In: YeastsYeasts in in Food Food and and 686686 11992.PDF.11992.PDF. Accessed Accessed 19 19 August August 2011. 2011. 718718 BeveragesBeverages,, (A. (A. Querol, Querol, G. G. Fleet Fleet eds.), eds.), Springer, Springer, 719719 NewNew York. York. USA. USA. 687687 Nwabueze,Nwabueze, T. T. U. U. (2010). (2010). Review Review article: article: Basic Basic 720 688688 stepssteps in in adapting adapting response response surface surface methodology methodology 720 Scragg,Scragg, A. A. (2009). (2009). Biotecnolog´ıaBiotecnolog´ıa para para ingenieros: ingenieros: 721 689689 asas mathematical mathematical modeling modeling for for bioprocess bioprocess 721 SistemasSistemas biol´ogicosen biol´ogicosen procesos procesos tecnol´ogicos tecnol´ogicos.. 722 690690 optimizationoptimization in in the the food food systems. systems. InternationalInternational 722 Limusa,Limusa, M Mexico.´exico.´ 691691 JournalJournal of of Food Food Science Science Technology Technology 45 45,, 1768- 1768- 723723 StatPointStatPoint Tecnology, Tecnology, Inc Inc (2005). (2005). Statgraphics Statgraphics 692692 1776.1776. 724724 CenturionCenturion (Data (Data Analysis Analysis Software Software System), System), v. v. 725725 XV,XV, Warrenton, Warrenton, VA VA (www.statgraphics.com). (www.statgraphics.com). 693693 Prasad,Prasad, K. K. and and Nath, Nath, N. N. (2002). (2002). Comparison Comparison of of 694694 sugarcanesugarcane juice juice based based beverage beverage optimization optimization 726726 Suder,Suder, Y., Y., Prakasam, Prakasam, R. R. S. S. and and Reddy, Reddy, O. O. V. V. 695695 usingusing response response surface surface methodology methodology with with Fuzzy Fuzzy 727727 S.S. (2009). (2009). Optimization Optimization of of fermentation fermentation 696696 method.method. Sugar Sugar Technology Technology 4, 4, 109-115. 109-115. 728728 conditionsconditions for for mango mango (Mangiferaindica (Mangiferaindica L.) L.) 729729 winewine production production by by employing employing response response 697697 Querol,Querol, A., A., Barrio, Barrio, E., E., Huerta, Huerta, T. T. and and Ram Ramon,´on,´ 730730 surface.surface. InternationalInternational Journal Journal of of Food Food Science Science 698698 D.D. (1992). (1992). Molecular Molecular monitoring monitoring of of wine wine 731731 TechnologyTechnology 44 44,, 2320-2327 2320-2327 699699 fermentationsfermentations conducted conducted by by active active dry dry yeast yeast 700700 strains.strains. AppliedApplied Environmental Environmental Microbiology Microbiology 732732 Swiers,Swiers, J., J., Bartowsky, Bartowsky, E., E., Henschke, Henschke, P. P. and and Pretorius, Pretorius, 701701 5858,, 2948-2953. 2948-2953. 733733 I.I. (2005). (2005). Yeast Yeast and and bacterial bacterial modulation modulation of of 734734 winewine aroma aroma and and flavor. flavor. AustralianAustralian Journal Journal of of 702702 Ratman,Ratman, B., B., Narasimha, Narasimha, M., M., Damodar, Damodar, M., M., Subba, Subba, 735735 GrapeGrape Wine Wine Research Research 11 11,, 139-173. 139-173. 703703 S.S. and and Ayyanna, Ayyanna, C. C. (2003). (2003). Optimization Optimization of of 704 704 fermentationfermentation conditions conditions for for the the production production of of 736736 TTellez-Mora,´ellez-Mora,´ P., P., Peraza-Luna, Peraza-Luna, F. F. A., A., Feria- Feria- 705 705 ethanolethanol from from sago sago starch starch using using response response surface surface 737737 Velasco,Velasco, A. A. and and Andrade-Gonz Andrade-Gonzalez,´alez,´ I. I. (2012). (2012). 706 706 methodology.methodology. WorldWorld Journal Journal of of Microbiology Microbiology 738738 OptimizationOptimization of of fermentation fermentation process process for for 707 707 andand Biotechnology Biotechnology 19 19,, 523-526. 523-526. 739739 tequilatequila production production using using response response surface surface 740740 methodologymethodology (RSM). (RSM). RevistaRevista Mexicana Mexicana de de 708708 Ratman,Ratman, B., B., Subba, Subba, S., S., Damodar, Damodar, M., M., Narasimha, Narasimha, 741741 Ingenier´ıaQu´ımica11Ingenier´ıaQu´ımica11,, 163-176. 163-176. 709709 M.M. and and Ayyanna, Ayyanna, C. C. (2005). (2005). Optimization Optimization 710710 ofof medium medium constituents constituents and and fermentation fermentation 742742 Valim,Valim, M. M. F., F., Rossi, Rossi, E. E. A., A., Silva, Silva, R. R. S. S. and and Borsato, Borsato, 711711 conditionsconditions for for the the production production of of ethanol ethanol 743743 D.D. (2003). (2003). Sensory Sensory acceptance acceptance of of a a functional functional 712712 fromfrom palmyra palmyra jaggery jaggery using using response response surface surface 744744 beveragebeverage based based on on orange orange juice juice and and soy soy milk. milk. 713713 methodology.methodology. WorldWorld Journal Journal of of Microbiology Microbiology 745745 BrazilianBrazilian Journal Journal of of Food Food Technology Technology 6 6,, 153- 153- 714714 andand Biotechnology Biotechnology 21 21,, 399-404. 399-404. 746746 156.156.

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