J. Adv. Educ. Sci. Humanit. (July - December 2023) 1(2): 7-12 https://doi.org/10.5281/zenodo.14602136 ISSN: XXXX-XXXX ORIGINAL ARTICLE  Mario A. García marioifal@gmail.com Received: 22 March 2023 / Accepted: 19 June 2023 / Published online: 31 July 2023 © The Author(s) 2023 Yailén García-Mirabal 1 · Oscar Ros 1 · Mario A. García 2 Abstract A spirit was developed from sorghum to eval- uate its potential application in whisky production. An acid hydrolysis process was employed at a pressure of 1.3 atm for four hours at 124 °C to extract nutrients from the grain. The resulting culture medium, maintained at 30 °C, was fil- tered and neutralized before adding Saccharomyces cerevisi- ae yeast for alcoholic fermentation. The spirit was obtained by distilling the fermented medium. The alcohol content, acidity, and pH of the sorghum spirits and the sorghum and corn mixtures were determined. The results showed that the sorghum flour’s particle size significantly influenced alcohol yield, with the lowest yield associated with larger particle sizes (1000-2000 µm). The alcohol yield of the spirits ob- tained from flour mixtures showed no significant differences. However, the 50% sorghum and corn flour mixture tended to increase this parameter compared to the other spirits. Sen- sory evaluators rated the spirits made with a 1:1 mixture of sorghum and corn flour as “Acceptable,” suggesting its po- tential as a raw material for whisky production. Keywords spirits, sorghum, corn, sensory evaluation, whisky. Resumen Se desarrolló un aguardiente a partir de sorgo con el objetivo de evaluar su posible aplicación en la elabo- ración de whisky. Para la extracción de nutrientes del grano se empleó un proceso de hidrólisis ácida a presión de1,3 atm durante cuatro horas a 124 °C. El medio de cultivo resultan- te, mantenido a 30 °C, fue filtrado y neutralizado previo a la adición de la levadura Saccharomyces cerevisiae para su fermentación alcohólica. El aguardiente se obtuvo median- te destilación del medio fermentado. Se determinó el grado alcohólico, acidez y pH de los aguardientes de sorgo y de las mezclas de sorgo y maíz. Los resultados mostraron que el tamaño de las partículas de la harina de sorgo tuvo una influencia significativa en el rendimiento de alcohol, siendo el menor rendimiento asociado al tamaño de partículas más grande (1000-2000 µm). El rendimiento alcohólico de los aguardientes obtenidos a partir de mezclas de harinas no pre- sentó diferencias significativas, aunque la mezcla de harina de sorgo y maíz al 50 % mostró una tendencia a incrementar este parámetro en comparación con los demás aguardientes. El aguardiente elaborado con una mezcla 1:1 de harina de sorgo y harina de maíz fue clasificado como “Aceptable” por los evaluadores sensoriales, lo que sugiere su potencial como materia prima en la producción de whisky. Palabras clave aguardiente, sorgo, maíz, evaluación senso- rial, whisky. How to cite García-Mirabal, Y., Ros, O., & García, M. A. (2023). Development and evaluation of sorghum-based spirits with potential for whisky production. Journal of Advances Education, Sciences and Humanities, 1(2), 7-12. https://doi.org/10.5281/zenodo.14602136 1 Instituto de Farmacia y Alimentos, Universidad de La Habana, La Habana, Cuba. 2 Facultad de Ciencias Matemáticas, Físicas y Químicas, Universidad Técnica de Manabí, Portoviejo, Ecuador. Development and evaluation of sorghum-based spirits with potential for whisky production Desarrollo y evaluación de aguardientes a base de sorgo con potencial para la producción de whisky

J. Adv. Educ. Sci. Humanit. (July - December 2023) 1(2): 7-12 8 Introduction Cereals are staple foods in many people’s diets, especially in developing countries. Their price in the global market has increased due to the use of these crops for biofuel produc- tion, primarily bioethanol (Prasad & Ingle, 2019). This pro- cess uses the starches and sugars present in cereals, which are fermented and distilled to obtain alcohol. This phenome- non has created direct competition between food production and biofuels, negatively affecting food prices (Prasad & In- gle, 2019). Barley, an essential ingredient in producing whisky, beer, and other malted products, has not been immune to this effect. This cereal is a significant source of sugars, prote- ins, dextrins, minerals, vitamins, and other compounds that determine its high nutritional value and the quality of the derived products. It is frequently used as an “adjunct” in pro- duction processes to reduce costs and increase profitability (Lukinac et al., 2022). The increase in malt prices directly impacts the cost of products made from this raw material, such as whisky. This distilled spirit, made from cereals such as barley, wheat, rye, and corn, is obtained through fermentation, distillation, and aging in oak barrels. Due to the high cost of barrels and the long time required for traditional aging, accelerated aging techniques have been implemented (Kelly et al., 2023). The- se techniques aim to optimize the contact between the spirit and the oak and enhance the extraction of compounds from the wood, thus improving the process (Krüger et al., 2022). The alcohol content of whisky varies between 40 and 62% (Ablin, 2012) and is known as the “water of life” (Tapia, 2008). To reduce production costs, sorghum emerges as a viable alternative significantly cheaper than barley. This cereal, whose main form of carbohydrate storage is starch, has an average content of 73.8%, mainly composed of amylopec- tin (70-80%) and a smaller proportion of amylose (20-30%). Genetic and environmental factors influence these propor- tions, giving sorghum different characteristics depending on its variety (Kang et al., 2022). Nutritionally, sorghum protein is deficient in lysine and threonine while containing exces- sive levels of leucine and low levels of methionine (Paoletti et al., 2022). Since the malting of sorghum does not efficiently activa- te its enzymes, alternative methods, such as acid hydrolysis under pressure, must be used to extract its nutrients. This process, which uses high temperatures, pressure, and acid over a controlled time, breaks down the starch and other compounds in the grain, yielding a suitable medium for fer- mentation (Feyera, 2021). In this context, this study aimed to develop a spirit from sorghum grain, using acid hydrolysis under pressure as a method for nutrient extraction, to apply it in whisky production. Methodology The production of the spirit involved the use of sorghum variety ISIAP-DORADO, cultivated on lands of the Nice- to Pérez Cooperative (Sancti Spíritus), and the yeast Sac- charomyces cerevisiae. The sorghum grains were polished to remove impurities using a polisher (PNO 12-013) and a vacuum cleaner (PNO 12-009). They were then ground in a Pulvex 200 mill, obtaining flours classified as acceptable (250-315 μm), medium (250-1000 μm), and coarse (1000- 2000 μm). Culture media were prepared with 266 g/L of sorghum flour for each particle size, weighed on a Sartorius 31005 technical balance (PNO 11.08.065.01.98), and acidified with H₂SO₄ 1 mol/L to pH=1. Three media were also formulated by combining sorghum and corn flour in 75%-25%, 50%- 50%, and 100% corn proportions. All media were prepared in duplicate and hydrolyzed in a Shenan autoclave at 1.3 atm and 124°C for 4 hours. After hydrolysis, they were neutrali- zed with NaOH 1 mol/L to pH=5. The activation of S. cerevisiae required 2.5 g of yeast hy- drated in 100 mL of distilled water with 10% sucrose, resting for 30 minutes. Each fermentation medium received 30 mL of activated yeast solution, incubated at 30 °C for 72 hours, followed by sedimentation at 4 °C for 24 hours. To distill the spirit, 500 mL of each fermented medium was taken and subjected to distillation at 100 °C. Fractions of 20 mL were collected until 100 mL per sample was com- pleted. The alcohol content was analyzed by pycnometer ac- cording to NC 623 (2008), relating relative density at 20 °C to alcohol percentage by volume and weight. The sensory evaluation used a five-point ordinal scale, according to NC-ISO 4121 (2005), rating sensory attributes with trained panelists. Samples (Old Premiers whisky, spirit, and Chanceler whisky) were randomly assigned codes, and averages and ranges were calculated (rejectable: 1-1.5; ex- cellent: 4.6-5). Quantitative data were analyzed using Statistica softwa- re (version 7, 2004, StatSoft, Inc., Tulsa, USA), calculating means, standard deviations, and analysis of variance (ANO- VA). The Duncan multiple range test was applied with signi- ficance p ≤ 0.05 (López-Planes, 1994).

J. Adv. Educ. Sci. Humanit. (July - December 2023) 1(2): 7-12 9 Results and discussion Table 1 shows the pH and acidity values of sorghum spi- rit with different particle sizes and varying concentrations of sorghum flour mixed with corn flour. Table 1. pH and acidity values of the spirits Spirit pH Acidity (% acetic acid) Treatment I (Fine) 4.36 (0.03) 9.5 (0.5) Treatment II (Medium) 4.23 (0.09) 8.3 (0.6) Treatment III (Coarse) 4.1 (0.05) 7.8 (0.3) Treatment IV (75% Sf-25% Cf) 4.37 (0.03) 10.3 (0.4) Treatment V (50% Sf-50% Cf) 4.58 (0.02) 11.3 (0.6) Treatment VI (100% Cf) 4.48 (0.07) 10.5 (0.8) Fine: 250 – 315 μm; Medium: 250 – 1000 μm; Coarse: 1000 – 2000 μm. Sf: sorghum flour; Cf: corn flour. Mean (Standard deviation); n = 2. No significant differences were observed in the pH values among the analyzed spirits, reflecting the homogeneity in the neutralization levels applied to the different fermenta- tion medium formulations. This result is consistent with the typical behavior of hydrolytic fermentation reactions, which generally cause slight decreases in pH compared to initial values. The obtained pH values fall within the characteristic range for freshly distilled spirits. Similarly, the acidity values (% acetic acid) showed no significant variations and aligned with the acidity levels commonly reported in fresh or newly distilled spirits (Barnes et al., 2022). Table 2 shows the results for alcohol production when var- ying the substrate particle sizes while maintaining a cons- tant concentration. The best spirit yields (expressed as mL of ethanol/g of sorghum) were recorded with particle sizes between 250-315 µm (fine), which reached the maximum value and showed significant differences compared to parti- cles of 1000-2000 µm (coarse) but not compared to particles Table 2. Alcohol content and yields of spirits obtained from sorghum flours with different particle sizes Particle size (µm) Fraction (20 mL) Alcohol content (%) Yield (mL of ethanol/g of sorghum) 250 - 315 µm (Fine) 1 29 (1) a 0.17 (0.01) a 2 10.5 (0.7) d 3 3.3 (0.8) gh 4 2.4 (0.1) hi 5 0.65 (0.07) k 250 - 1000 µm (Medium) 1 26 (0.3) b 0.15 (0.01) ab 2 7.4 (0.3) e 3 4 (1) g 4 1.96 (0.22) hj 5 0.97 (0.09) jk 1000 - 2000 µm (Coarse) 1 23.3 (0.3) c 0.135 (0.001) b 2 5.95 (0.35) f 3 4.15 (0.07) g 4 1.97 (0.04) jk 5 0.9 (0.1) jk Mean (Standard deviation); n = 2. Different letters between means of the same indicator differ significantly at p ≤ 0.05. 1000 µm (medium). This behavior was attributed to the fact that smaller particles allow for greater extraction of es- sential nutrients, such as starch, and improve flour recovery, optimizing the effectiveness of the hydrolytic treatment un- der acid hydrolysis at pressure. The flours with particle sizes of 250-315 µm (fine) and 250-1000 µm (medium) were mixed in a 1:1 ratio to substi-

J. Adv. Educ. Sci. Humanit. (July - December 2023) 1(2): 7-12 10 Table 3. Alcohol content and yields of spirits obtained from mixtures of sorghum and corn flour Substrate Fraction (20 mL) Alcohol content (%) Yield (mL of ethanol/g of sorghum) Treatment IV (75% Sf-25% Cf) 1 29,5 (0,7) b 0,17 (0,01) a 2 10,5 (0,8) d 3 2,65 (0,07) fg 4 2,4 (0,1) g 5 0,77 (0,03) h Treatment V (50% Sf-50% Cf) 1 31,1 (0,2) a 0,185 (0,003) a 2 11,5 (0,1) c 3 3,4 (0,6) e 4 2,48 (0,03) g 5 0,9 (0,1) h Treatment VI (100% Cf) 1 29,8 (0,3) b 0,178 (0,001) a 2 10,85 (0,07) cd 3 3,55 (0,07) e 4 2,25 (0,07) g 5 0,95 (0,07) h Sf: sorghum flour; Cf: corn flour. Mean (Standard deviation); n= 2. Different letters between means of the same indicator indicate significant differences for p ≤ 0.05. tute for corn flour in spirits, as no significant differences in in ethanol production were observed between them. Table 3 reports the alcohol content values obtained from hydrolyzed preparations of mixtures of sorghum and corn flours at different concentrations. Since no significant differences were observed in the al- cohol yield between the spirits obtained from 100% corn flour and the 75% sorghum flour and 25% corn flour mix- ture, the feasibility of implementing spirits production for whisky production using mixtures with sorghum flour was considered. Although the alcohol yield did not show significant va- riations, the spirits tended to increase with a 50% sorghum flour and 50% corn flour mixture, possibly due to the nutrient balance achieved in this proportion. Alcohol yield could be influenced by genetic, environmental factors, and cultivation practices; moreover, processing operations, such as poli- shing, could impact the grain properties and, consequently, this property. The sensory evaluation of the spirits from Treatment V, a 1:1 mixture of sorghum flour and corn flour, was conduc- ted according to Walsh (2007), who suggested that sensory evaluation is a quantitative science in which numerical data is collected to determine specific relationships between pro- duct characteristics and human perception. This spirit was selected due to its tendency for higher alcohol yield. Table 4 presents the scores obtained for each beverage evaluated by the judges. Table 4. Evaluation of the sensory quality of the beverages evaluated Sample (Code) Parameter Score Sensory rating Old Premiers (534) Mode 4 Very good Minimum 2 Acceptable Maximum 4 Very good Average 3.2 Good Spirits (124) Mode 2 Acceptable Minimum 1 Rejectable Maximum 3 Good Average 1.9 Acceptable Chanceler (234) Mode 5 Excellent Minimum 1 Rejectable Maximum 5 Excellent Average 3.2 Good

J. Adv. Educ. Sci. Humanit. (July - December 2023) 1(2): 7-12 11 Conclusions The particle size of the sorghum flour significantly in- fluenced the alcoholic yield of the spirits, with the lowest yield observed in larger particle sizes. Although no sig- nificant differences were detected in the alcoholic yields of spirits obtained from flour mixtures, the equal combi- nation of sorghum flour and corn flour showed a trend toward increased yields compared to the other formula- tions. The spirits produced with this balanced mixture were classified as “Acceptable”, highlighting the potential to continue developing this product by optimizing pro- duction processes to improve its quality and sensory ac- ceptance. The alcoholic yield and sensory characteristics support the use of sorghum flour as a viable component for diversifying and expanding whisky production. References Ablin, A. (2012). El mercado del whisky. Alimentos argenti- nos MinAgri, 1-21. https://alimentosargentinos.magyp. gob.ar/HomeAlimentos/AyB/bebidas/productos/Whis- ky_2012_11Nov.pdf Barnes, Q., Vial, J., Thiébaut, D., De Saint, C., Steyer, D., Contamin, M.A., Papaiconomou, N., & Fernandez, X. (2022). Characterization of Flavor Compounds in Dis- tilled Spirits: Developing a Versatile Analytical Method Suitable for Micro-Distilleries. Foods, 11(21), 3358. https://doi.org/10.3390/foods11213358 Feyera, M. (2021). Overview of Malting and Fermentation Role in Sorghum Flour, Primarily for Antinutrient Re- duction. Journal of Human Nutrition and Food Science, 9(1), 1138. https://www.jscimedcentral.com/jounal-ar- ticle-pdfd/Journal-of-Human-Nutrition-and-Food-Sci- ence/nutrition-9-1138.pdf Kang, X., Zhu, W., Xu, T., Sui, J., Gao, W., Liu, Z., Jing, H., Cui, B., Qiao, X., & Abd El-Aty, A.M. (2022). Charac- terization of starch structures isolated from the grains of waxy, sweet, and hybrid sorghum (Sorghum bicolor L. Moench). Frontiers in Nutrition, 9, 1052285. https:// doi.org/10.3389/fnut.2022.1052285 Kelly, T.J., O’Connor, C., & Kilcawley, K.N. (2023). Sources of Volatile Aromatic Congeners in Whiskey. Beverages, 9(3), 64. https://doi.org/10.3390/beverages9030064 Krüger, R.T., Alberti, A., & Nogueira, A. (2022). Current Technologies to Accelerate the Aging Process of Alco- holic Beverages: A Review. Beverages, 8(4), 65. https:// doi.org/10.3390/beverages8040065 Lukinac, J., & Jukić, M. (2022). Barley in the Production of Cereal-Based Products. Plants (Basel), 11(24), 3519. https://doi.org/10.3390/plants11243519 NC 623 (2008). Determinación de alcohol por picnometría. Cuba. NC-ISO 4121 (2005). Análisis sensorial. Directrices para la utilización de escalas de respuestas cuantitativas. Cuba. Paoletti, A., Fakiha, A., Tul-Noor, Z., Pencharz, P.B., Levesque, C.L., Ball, R.O., Kong, D., Elango, R., & Courtney-Martin, G. (2022). Bioavailable Lysine As- sessed Using the Indicator Amino Acid Oxidation Method in Healthy Young Males is High when Sor- ghum is Cooked by a Moist Cooking Method. Journal of Nutrition, 152(3), 770-778. https://doi.org/10.1093/ jn/nxab410. Prasad, S., & Ingle, A.P. (2019). Impacts of sustainable bio- fuels production from biomass. Eds. Mahendra Rai, Avinash P. Ingle, Sustainable Bioenergy, Elsevier. https://doi.org/10.1016/B978-0-12-817654-2.00012-5. Tapia, A. (2008). Whisky. El agua de la vida. www.shannon- sleon.com/archivos/Whisky_ El_agua_de_la_vida.pdf. Walsh, C. (2007). Consumer responses to low-salt food products. Eds. David Kilcast, Fiona Angus, In Wood- head Publishing Series in Food Science, Technology and Nutrition, Reducing Salt in Foods, Woodhead Pub- lishing. https://doi.org/10.1533/9781845693046.1.124 Conflicts of interest The authors declare that they have no conflicts of interest. Author contributions Conceptualization: Yailén García-Mirabal, Oscar Ros. Data curation: Yailén García-Mirabal, Mario A. García. Formal analysis: Yailén García-Mirabal. Research: Yailén García-Mirabal, Oscar Ros, Mario A. García. Methodology: Oscar Ros, Mario A. García. Software: Mario A. García. Supervision: Oscar Ros. Validation: Oscar Ros. Visualiza- tion: Yailén García-Mirabal, Mario A. García. Writing the original draft: Yailén García-Mirabal, Oscar Ros, Mario A. García. Writing, review and editing: Yailén García-Mira- bal, Mario A. García. Data availability statement The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. Statement on the use of AI The authors acknowledge the use of generative AI and

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