Effects of Conventional Processing Methods and Growing Locations on the Phenolic Content and Bioactive Compounds of Ethiopian Coffee Beans

Effects of Conventional Processing Methods and Growing Locations on the Phenolic Content and Bioactive Compounds of Ethiopian Coffee Beans | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Effects of Conventional Processing Methods and Growing Locations on the Phenolic Content and Bioactive Compounds of Ethiopian Coffee Beans Dhaba Mengesha Adula This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7194131/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 14 You are reading this latest preprint version Abstract Growing conditions, roasting, and brewing methods are among the factors and procedures that determine the bioactive compound and phenolic content of brewed coffee. This study aimed to evaluate the impact of growing locations and traditional coffee processing methods on the bioactive components and phenolic content of Ethiopian coffee. The efficient Ethiopian traditional coffee processing procedures with the commonly used medium particle size of coffee powder and widely used water type (surface water) were used for brewing by the Ethiopian traditional coffee brewing method (190 0 C of roasting, 16 min of brewing time). HPLC was used to assess the phenolic content and determine the bioactive chemicals. All five Ethiopian Coffee Arabica origins (Jimma, Sidama, Yirgachefe, Nekemte, and Hararge) had caffeine levels between 7.55 and 10.38 mg/mL. The coffee beans from Yirgachefe and Hararge had the greatest and lowest caffeine content, respectively (p < 0.05). The caffeine level of each variety varies significantly (p 0.05), at 45 mg/mL. With 36.78 mg/g of chlorogenic acid, the Hararge cultivar had the lowest levels. Trigonelline levels were also highest in the Jimma, Sidama, and Nekemte coffee varietals, with 12.88, 13.56, and 13.46 mg/mL, respectively (p > 0.05). Hararge and Yirgachefe kinds had the lowest concentrations, with respective values of 11.65 and 11.78 mg/mL (p > 0.05). TPC was considerably lower after roasting all coffee beans (p < 0.05). The following coffee beans have the lowest proportion of TPC: Jimma (24%), Sidama (26%), Nekemte (30%), Yirgachefe (23%), and Hararge (29%). On average, the reduction was a 27% factor. After roasting, the caffeine concentration of the Sidama, Nekemte, and Yirgachefe coffee beans varied significantly (p < 0.05). Jimma (83%), Sidama (79%), Nekemte (82%), Yirgachefe (81%), and Hararge (82%), among all coffee origins, had the highest significant percentage decrease in chlorogenic acid content (p < 0.05). On average, there was an 81% decrease. Furthermore, there was a substantial variation in the trigonelline content across all coffee varietals during roasting (p < 0.05). With Jimma (54%), Sidama (29%), Nekemte (45%), Yirgachefe (38%), and Hararge (34%), the average reduction was 40%. In general, the overall amount of bioactive components and phenolic contents in processed coffee is influenced by growing regions and coffee processing. Further research may be necessary to determine how coffee's phenolic content and bioactive components are affected by brewing temperature and extraction methods. Bioactive Compounds Growing Location Green Coffee Traditional Processing Roasting Introduction Coffee is becoming increasingly popular as a daily beverage and as a crop crucial to Ethiopian and international trade. In Ethiopia and around the world, coffee is one of the most commercially significant crops and one of the most culturally popular daily beverages. Its consumption is also rising these days. According to a survey by the International Coffee Organization (ICO), 4 billion cups of coffee, or 9.4 million tons, were consumed worldwide in 2016–2017. These cups are made using various brewing techniques. Coffee is composed of several chemical constituent combinations that influence its flavor and aroma. These consist of alkaloids, phenolic compounds, vitamins, minerals, carbohydrates, lipids, and nitrogenous chemicals. In addition, coffee contains a lot of health benefits and is rich in bioactive substances including trigonelline, caffeine, and chlorogenic acid [ 35 ]. Drinking coffee has been linked to a lower risk of hepatocellular carcinoma and vascular diseases [ 34 ], anti-proliferative effects on certain human cancer cells [ 14 ], and potential therapeutic benefits against Alzheimer's disease, liver disease, and diabetes[ 12 , 37 , 43 ]. The entire coffee process, from selecting coffee beans to choosing the best water for brewing to serving the coffee, affects the end product's quality and sensory attributes. To achieve a higher sensory score, coffee must have at least five distinct flavor notes in harmony [ 11 , 16 ]. The Specialty Coffee Association of America [ 8 ], criterion analysis procedure states that HPLC is used to analyze the bioactive compounds in roasted and brewed coffee. The coffee's ability to pass aspect or preliminary grading and cupping tests can be used to assess its sensory and cup quality [ 7 ]. Important steps that affect the quality and quality of the final coffee product include sorting and grading green coffee beans, roasting, grinding, and different extraction methods [ 39 ]. Depending on the desired qualities of the coffee cup, dry coffee beans are roasted by heating them to temperatures ranging from 200 to 240°C for varying amounts of time [ 13 ]. Green beans lose a lot of water, which makes them fragile. Numerous metabolic pathways, including Strecker and Maillard, produce over 1000 distinct kinds of aromatic chemicals [ 42 ]. Alcohols, aldehydes, amines, carboxylic acids, dicarbonyls, enoles, esters, furans, furanone, hydrocarbons, imidazoles, indoles, ketones, lactones, oxazoles, phenols, pyrazines, pyridines, pyrroles, quinoxalines, sulfur compounds, terpenes, and thiazoles are among the volatile substances found in roasted coffee beans [ 21 , 39 ]. Depending on the heat profile used during roasting, these chemicals may experience significant modifications. For this reason, roasting is thought to be the most crucial process in identifying the coffee bean's distinct flavor and color. The different roasting conditions have a big impact on the physical and chemical properties of roasted coffee beans. After roasting, the coffee beans are ground to maximize their surface area and regulate humidity for effective extraction. While the caffeine content does not significantly drop after roasting, 20–40% of the polysaccharides that store cell walls are broken down [ 10 , 44 ]. According to Konstantinidis [ 24 ], trigonelline's primary metabolites, nicotinic acid and N-methylpyridinium, are helpful markers of roasting level. Metabolites created by microbes may infiltrate into the beans after roasting to escape the heat process. The metabolites with the greatest ability to affect the final coffee beverage's quality are flavor-active esters. Following roasting, complex physical and chemical changes, including caramelization, are produced by the Maillard and Strecker process, which involves hundreds of biological components [ 21 ]. The Maillard reaction, which transforms sugar into color, flavor, and fragrance, is catalyzed by amino acids. Acetic and formic acids are primarily responsible for the potent scent during the initial roasting stages. As quinic acid concentration rises, the breakdown of chlorogenic acids results in astringent and bitter flavors [ 17 ]. Roasting significantly lowers trigonelline and CGA while leaving caffeine unchanged [ 20 ]. Green beans' trigonelline, carbs, and some proteins also decompose into volatile chemicals, claim [ 5 ]. Coffee beans get their distinctive dark color from melanoidin, which is a byproduct of the Maillard reactions and may help preserve the flavor components [ 27 ]. While trigonelline helps produce both acceptable and disagreeable smell molecules during roasting, caffeine accounts for more than 30% of the bitter taste [ 17 ]. Asparagine and glucose-fructose can promote the Maillard reactions, which result in the production of unwanted substances like furan and acrylamides [ 27 ]. The Strecker degradation, a component of the Maillard reaction network, contributes volatile aldehydes to the coffee's aroma spectrum, including notes of honey, sulfur, malt, and potato. Coffee origin, grinding size, and brewing time all affect the production of coffee aromas, the composition of bioactive components, and the antioxidant potential of brewed coffee. Coffee's flavor and aroma can change during the preparation process due to its biochemical makeup [ 26 ]. Ethiopian coffee is processed using very healthful methods, employing locally produced elements that are very tasty to consumers and quite beneficial after intake. The quality of Ethiopian coffee beans is therefore influenced by a variety of factors, including the growing environment, processing methods, and particularly the roasting process. Though there are some gaps in the current research on the efficient coffee processing methods and effects of coffee origin on the phenolic and bioactive compounds of brewed coffee, the coffee's quality and taste can also be greatly influenced by the coffee's origin and brewing materials. The majority of current studies, for instance, have concentrated on how coffee origin and processing methods affect the biochemical composition and cup quality of coffee beans from other nations. Furthermore, the majority of previous studies have employed rather small sample sizes; additionally, most of the existing research has used relatively small sample sizes, which makes it difficult to generalize the findings to the wider population. Ethiopian traditional coffee processing techniques and their efficacy, in addition to the growing environment, affect the physico-chemical characteristics and cup quality of the brewed coffee. Not all of the main Ethiopian coffee beans have had their bioactive components and phenolic contents thoroughly examined regarding the effects of cultivation circumstances and efficient processing methods. This study was carried out to ascertain how the bioactive composition and physicochemical characteristics of Ethiopian coffee, which is traditionally roasted and brewed, were affected by the growing environment and processing stages. This study aimed to evaluate the effects of processing procedures and growing locations on the bioactive composition, physicochemical properties, and sensory acceptability of Ethiopian brewed coffee. Materials and Methods Coffee beans Ethiopian coffee beans were gathered from the Ethiopian Coffee and Tea Authority Coffee Quality Inspection and Certification Center (ECTACQICC) in Addis Ababa, Ethiopia, representing various Jimma, Hararge, Sidama, Nekemte, and Yirgachefe origins [ 30 ]. Three kilograms of each type of coffee were gathered. After cleaning and washing the coffee beans, any damaged seeds were taken out. Standardizing the roasting and brewing process for Ethiopian traditional coffee In ten Addis Ababa sub-cities, the traditional Ethiopian coffee-processing method was surveyed. Addis Ketema, Arada, Bole, Gullele, Kirkos, Kolfe Keranio, Ledeta, Knife’s Silk Lafto, Akaky Kaliti, and Yeka were the ten sub-cities from which ten homes were chosen at random to participate in the study. The brewing process was then interviewed using questionnaires in each home. Actual measurements (weighed records) were also taken when needed. Mengesha et al [ 30 ], constructed a coffee processing flowchart based on the survey and simulated it in a lab. This consistent procedure was used to prepare all of the brewed and roasted coffee powder samples. Roasting The coffee beans were roasted in a metal roasting pan measuring 16 cm in diameter and 76 mm in thickness. Charcoal cooking required 10 to 12 minutes, whereas electric roasting took 6 to 9 minutes at 190°C. The beans were roasted and then allowed to cool at room temperature for 11 minutes before being ground using a traditional wooden mortar and pestle to a sieve size of 1.2 mm. Coffee brewing The coffee bean was roasted for 16 minutes at a maximum temperature of 190°C, which is the standard and advised level of roast [ 38 ]. The exothermic reaction was then terminated by leaving the air-cooled coffee beans overnight to allow for full taste development and CO2 degassing. The roasted coffee beans were ground into fine (2 mm), medium (4 mm), and coarse (6 mm) powder sizes using an electronic laboratory-scale coffee grinder (VTA6S, MAHLKÖNIG GmbH & Co., 2009) [ 9 ]. Following that, the ground coffee samples were placed in airtight, odor-proof polyethylene bags. Before brewing and analysis, the sample was kept at 4°C and 70% relative humidity. Coffee was made using surface water and 1:17 (w/w) ratios of coffee powder to water, as per the method described by [ 4 ]. Sample extraction for bioactive compounds To extract the total phenolic, flavonoid content, and bioactive components present in green and roasted coffee, the methodology of Iriondo-Dehond et al [ 23 ] was adopted. In summary, 3 g of ground green coffee was mixed with 20 mL of 98% methanol. Each coffee sample was mixed with twenty milliliters of 98% methanol. The samples were placed in Erlenmeyer flasks in a shaking incubator (Lucadema, Brazil) for 30 minutes at 40°C and 120 rpm, and then centrifuged for 15 minutes at 3200 rpm in an Excelsa Baby II centrifuge (Flamen, model 206-R). The supernatant's phenolic compounds were detected. Three extractions were performed. Determination of total polyphenol content (TPC) According to Hidayat et al [ 40 ], the Folin-Ciocalteu method was used to estimate the TPC. Gallic acid was used to generate the TPC measuring standard curve. Gallic acid equivalent (GAE)/g of the dry matter of coffee samples was used to express the results. One milliliter of 99% ethanol was added to standard solutions of gallic acid, which included 2, 4, 6, 8, 10, and 12 mg of gallic acid. After that, three milliliters of distilled water were added. Next came the addition of 0.1 mL of Folin-Ciocalteu reagent. Three minutes later, distilled water was added to the volumetric flasks to bring them up to the mark, and then 0.3 mL of a 2% sodium carbonate solution was added. The entire material was stored for two hours at 25°C. The gallic acid solution was then substituted with 100 mL of ethanol. The spectrophotometer's wavelength was set to 765 nm after two hours. The absorbance of each solution was measured three times, and the average was recorded after the device was initially zeroed with the control (gallic acid) solution. Following the development of a standard curve that plotted concentration (µg) against absorbance, the line equation was applied to the quantification. Ten milligrams of the green bean extract were weighed, placed in a test tube, and then completely dissolved with two milliliters of dimethyl sulfoxide (DMSO) solution to create the coffee samples. Triplicate assays were performed on each extracted sample using three 5 mL volumetric flasks. There were 0.20 mL of each extract and 3 mL of distilled water in each volumetric flask. The volumetric flasks were then filled with 100 µL of Folin–Ciocalteu reagent, and three minutes later, 0.3 mL of sodium carbonate (2%) was added. Distilled water was added to the volumetric flasks to bring them to the mark. Instead of the extract, 0.20 mL of DMSO was added to a 5 mL control volumetric flask. The mixture was homogenized and then stored for two hours at 25°C. After two hours, the absorbance of each solution was measured three times at 765 nm using a UV-Vis spectrophotometer (PerkinElmer, Model CF728YW-950, UK). The TPC was expressed as µg of GAE/g of extract using Eq. 1 . $$\:TPC=\:\frac{C*V}{M}\:\times\:\:D$$ 1 Where; C = gallic acid equivalent concentration obtained from the calibration curve (g/mL) V = volume of the extract's stock solution (mL) M = dry weight of the extract in the stock solution (g) TPC = Phenolic content expressed as (mg of GAE/g dry extract) D = dilution factor Bioactivity Analysis Caffeine, trigonelline, and chlorogenic acid extraction A significantly modified version of the [ 37 ], technique was used to extract CAF, CGA, and TRG. A 50 mL Erlenmeyer flask containing roughly 0.5 g of finely ground coffee grounds was carefully weighed. Stir on a hot plate for 20 minutes after adding 50 mL of heated (95°C) pure water. 10 µL of the extract was placed in an Agilent 1260 Infinity, Germany, high-performance liquid chromatography (HPLC) device after it had been filtered through No. 4 What-man filter paper and a 0.22 µm pore size filter. Measurement of caffeine, trigonelline, and chlorogenic acids Using a method adapted from [ 37 ], TRG, CGA, and CAF were measured concurrently using an HPLC system with a C18 column and an isocratic flow of 0.7 mL/min. A 4.6 × 250 mm column with a 5 µm particle size was used by Waters, Taunton, USA. At a flow rate of 0.7 mL/min, the following elute compounds were present in gradients with 5% acetic acid (A) and acetonitrile (B): 4% B for 0–4 minutes, 10% for 4–8 minutes, 90% for 8–12 minutes, 0% for 12–15 minutes, and 4% for 15–17 minutes. A three-minute post-run was held. TRG and CAF were measured at 272 nm, but CGA was detected at 320 nm. TRG and CAF test concentrations were 5, 10, 20, 40, 50, 100, and 150 µg/mL. The ranges for CGA, on the other hand, were 5, 10, 20, 100, 150, and 200 µg/mL. The identities of CAF, TRG, and CGA were ascertained by comparing the retention durations of the TRG (Sigma Aldrich), CGA (Acros Organics), and CAF (Fischer Scientific) standards (99%), as well as their concentrations derived from peak regions using calibration equations. Following a time of retention, the area was utilized to determine the composition of the related biological component, which was then, determined using calibration curves that plotted the area against standard concentration [ 47 ]. The limit of detection (LOD) for caffeine, trigonelline, and chlorogenic acid was 0.9, 1.02, and 1.5 µg/g, respectively and the limits of quantification (LOQ) for caffeine, trigonelline, and chlorogenic acid was 1.7, 2.3 and 3.0 µg/g, respectively. Statistical analysis The study employed two-factor factorial experimental designs with three treatment levels. Roasting and coffee-producing areas (A and B) were the chosen criteria. Five coffee origins—Jimma, Sidama, Yirgachefe, Hararge, and Nekemte coffee—were employed for brewing purposes- and green and roasting methods. At least three observations were made during the fully randomized experimental design. Using SPSS software version 26, the collected experimental findings were examined and interpreted using analysis of variance (ANOVA) at a 5% significance level. Results and discussion Standardized Ethiopian coffee brewing method Most Ethiopians drink coffee prepared by the traditional processing method, which varies slightly from home to home. Thus, this study's main goal was to simulate in the laboratory a standardized flowchart for traditional coffee processing Mengesha et al [ 30 ] using data from a qualitative survey that was carried out in each of Addis Ababa's ten sub-cities. The acrylamide levels were then measured after coffee was prepared in a lab environment using this flowchart. The Ethiopian traditional method of preparing coffee involves weighing 30 to 35 grams of coffee beans per serving, cleaning them, and washing them with clean tap water. After that, the beans were roasted in a metal roasting pan that measured an average of 16 cm in diameter and 76 mm in thickness. It took 6 to 9 minutes to roast at 190°C with electric power, and 10 to 12 minutes to cook using charcoal. A traditional wooden mortar and pestle was used to grind the beans to a sieve size of 1.2 mm after they had been roasted and allowed to cool for 11 minutes at room temperature. After that, the ground coffee was brewed in a traditional clay kettle, or boiling pot, using 25–30 grams of coffee for every 750–800 milliliters of water. The brewing procedure took 8 to 12 minutes, and the temperature ranged from 92 to 95°C. Before serving, the coffee was allowed to decant at room temperature for five minutes after boiling. The grinds are brewed three times using the same method in Ethiopian traditional coffee processing. This implies that two more brews are made using the same coffee grounds and the same amount of water following the first brew. "Abol," "Tona," and "Baraka" are the local terms for the first, second, and third rounds of coffee, respectively. Nonetheless, the survey indicates that consumers frequently favor the first and second brews. Therefore, another objective of this study was to determine the amount of acrylamide in the two serving rounds. The main way that people are exposed to acrylamide is through their diet [ 35 ]. Bread, coffee, and French fries have been repeatedly identified as major contributors to dietary acrylamide exposure in several studies that have looked at exposure among people with different characteristics [ 2 ]. The need for additional study on dietary acrylamide exposure and related health hazards is emphasized by the European Commission and the Joint FAO/WHO Expert Committee on Food Additives (European Commission, 2019; Joint FAO/WHO Expert Committee on Food Additives, 2011). The amount of acrylamide in brewed and powdered coffee from commercially processed and street-vendor sources varies significantly. Therefore, to accurately quantify the exposure to acrylamide from coffee drinking, it is imperative to follow the most commonly used processing methods. Impact of Ethiopian traditional coffee roasting on the biochemical contents The impact of roasting Ethiopian coffee on the biochemical makeup of coffee beans from various agroecological origins is documented in Table 1 . The caffeine concentration ranged from 7.55 to 10.38 mg/mL for the five Ethiopian Coffee Arabica cultivars. Yirgachefe coffee beans had the highest caffeine content, while Hararge coffee beans had the lowest (p < 0.05). All cultivars have a substantial variation in caffeine concentration (p < 0.05), except for Nekemte and Yirgachefe. As evidence for this work, several investigations have reported varying quantities of caffeine in coffee beans [ 42 ]. For instance, Mengistu et al [ 31 ] used HPLC techniques to find that the caffeine concentrations in 42 Ethiopian coffee samples ranged from 9.6 ± 0.01 to 12.3 ± 0.06 mg/mL, with an average value of 10.10 mg/mL. Other studies have shown that the average caffeine content of Arabic coffee is less than 15 mg/mL [ 22 ]. The caffeine concentration of this study is higher than that of Farah [ 15 ], investigation, which discovered that the caffeine content of various coffee varieties ranges from 9 to 25 mg/g. The highest levels of chlorogenic acid (p > 0.05) were found in the Jimma, Sidama, and Nekemte coffee varietals (45 mg/mL) in comparison to the other two. With 36.78 mg/g of chlorogenic acid, the Hararge cultivar had the lowest levels. Chlorogenic acid levels in several coffee bean varietals ranged from 4.1 to 11.3 mg/g, according to [ 15 ]. This number is greater than the one found in the current study. Trigonelline levels were also highest in the Jimma, Sidama, and Nekemte coffee varietals, with 12.88, 13.56, and 13.46 mg/mL, respectively (p > 0.05). Hararge and Yirgachefe kinds had the lowest concentrations, with respective values of 11.65 and 11.78 mg/mL (p > 0.05). Every home uses Ethiopian coffee, which is roasted and brewed distinctively. Reports on its impact on the biochemical makeup of the regional coffee types are, nevertheless, few. As seen in Table 1 , roasting significantly decreased the TPC (p < 0.05) for all coffee beans. The coffee beans from Jimma (24%), Sidama (26%), Nekemte (30%), Yirgacheffe (23%), and Hararge (29%), had the lowest percentage of TPC. On average, there was a 27% drop. The caffeine content of the Sidama, Nekemte, and Yirgacheffe coffee beans varied significantly after roasting (p < 0.05). Among all coffee varietals, the chlorogenic acid content had the largest significant percentage decrease (p < 0.05). Jimma (83%), Sidama (79%), Nekemte (82%), Yirgacheffe (81%), and Hararge (82%), were the countries with the largest percentage decreases. The average reduction was 81% overall. Furthermore, upon roasting, there were significant differences in the trigonelline content across all coffee kinds (p < 0.05). With Jimma (54%), Sidama (29%), Nekemte (45%), Yirgacheffe (38%), and Hararge (34%), the average reduction was 40%. Table 1 Effects of traditional Ethiopian coffee bean roasting on the levels of caffeine, trigonelline, chlorogenic acid, and total phenols in green and roasted Ethiopian Coffee cultivars in various agro-ecological zones. Coffee origin Coffee type Total polyphenols (mg GAE/g) Caffeine (mg/mL) Chlorogenic acid (mg/mL) Trigonelline (mg/mL) Jimma Green coffee 46.52 ± 0.82 a 8.85 ± 0.00 b 45.95 ± 0.01 a 12.88 ± 0.01 ab Roasted 35.56 ± 1.08 cd 8.84 ± 0.00 b 7.84 ± 1.08 de 5.88 ± 1.00 f Sidama Green coffee 44.31 ± 4.68 ab 8.23 ± 0.00 c 45.29 ± 0.01 a 13.56 ± 0.02 a Roasted 32.87 ± 0.05 d 8.11 ± 0.01 d 9.38 ± 0.91 d 9.60 ± 0.66 cd Nekemte Green coffee 44.55 ± 3.28 ab 10.18 ± 0.00 a 46.39 ± 0.01 a 13.46 ± 0.01 a Roasted 31.36 ± 0.19 de 9.88 ± 0.10 b 8.43 ± 0.69 de 7.33 ± 0.67 de Yirgachefe Green coffee 34.25 ± 3.83 c 10.38 ± 0.00 a 39.54 ± 0.01 b 11.78 ± 0.00 b Roasted 26.56 ± 0.91 f 9.92 ± 0.07 b 7.66 ± 0.84 ef 7.27 ± 1.19 de Hararge Green coffee 39.02 ± 2.93 bc 7.55 ± 0.00 e 36.78 ± 0.02 c 11.65 ± 0.00 b Roasted 27.86 ± 1.01 ef 7.45 ± 0.03 e 6.46 ± 0.89 f 7.64 ± 0.64 de Average Green coffee 41.73 ± 0.43 * 9.51 ± 0.66 * 42.79 ± 0.67 * 12.67 ± 0.47 * Roasted 30.84 ± 0.75 8.84 ± 0.45 7.95 ± 0.46 7.54 ± 1.34 On a dry basis, the data are presented as mean + SD (n = 3). Significant differences at p < 0.05 in mean comparison using the Duncan multiple range test are shown by mean values with different superscript letters within a column. *Uses the paired t-test to show a significant difference between green and roasted coffee beans (p < 0.05). The average CGA level in the current study ranged from 45.95 to 36.78 mg/g on a dry weight basis, while the average caffeine content ranged from 10.38 to 8.23 mg/mL. According to Farah [ 15 ], these values are often in the lower range of values previously observed for green Arabica coffee beans. Our caffeine measurements were marginally higher for both CAF and CGA than those published by [ 35 ]. Chlorogenic acid content in brewed and roasted coffee ranged from 16 to 33 percent. This is in line with the range discovered by Caprioli et al [ 28 ], who demonstrated that spent and roasted coffee still had 23–68% chlorogenic acid. 23–36% of the caffeine was still present in the wasted coffee. Retention rates for Gallic and protocatechuic acids were lower in comparison, at 4–13 and 4–24%, respectively. These findings show that the retention rates of each component differ according to its water solubility and high-temperature stability. Chlorogenic acids are the primary phenolic components found in green coffee. According to Mehari et al [ 29 ], they are esters of quinic acid and trans-hydroxycinnamic acid. It is known that these chemical groups are responsible for the color, flavor, bitterness, and astringency of coffee beverages [ 25 ]. About 6–12% of green coffee beans' dry weight is composed of chlorogenic acids [ 32 ]. When compared to the other two coffee kinds in this investigation, the Jimma, Sidama, and Nekemte varietals had the greatest levels of chlorogenic acid (p > 0.05), at 45 g/100 g. With 36.78 g/100 g of chlorogenic acid, the Hararge variety exhibited the lowest levels (Table 2 ). There is an inverse relationship between a coffee variety's chlorogenic acid concentration and beverage quality; coffee types with lower beverage quality had higher chlorogenic acid concentrations [ 31 ]. The green coffee beans gathered from nine districts in Southwest Ethiopia have a chlorogenic acid level ranging from 2.80 to 5.42 g/100 g [ 41 ]. According to Urugo et al [ 42 ], the amount of chlorogenic acids in green coffee beans gathered from northern Ethiopia ranged from 3.29 to 7.73 g/100 g. According to Farah [ 15 ], the amounts of chlorogenic acid in various coffee types ranged from 4.1 to 11.3 g/100 g. Compared to the current study, the chlorogenic acid content was lower in all of these earlier investigations. According to Awwad et al [ 6 ], chlorogenic acids exhibit potent antiviral, antidiabetic, antioxidant, and neuroprotective properties. Accordingly, the highest trigonelline levels were found in the Jimma, Sidama, and Yirgachefe coffee varietals, with 12.88, 13.56, and 13.46 mg/100 g, respectively (p > 0.05). The Nekemte and Hararge types had the lowest concentrations, measuring 11.78 and 11.65 mg/100 g, respectively (p > 0.05) (Table 2 ). The amount of trigonelline in coffee beans gathered from nine districts in Southwest Ethiopia varied significantly (from 0.80 to 1.08 g/100 g), according to [ 37 ]. Trigonelline levels in green coffee beans gathered from the country's north ranged from 0.53 to 1.27 g/100 g, according to [ 42 ]. Trigonelline levels were lower in these earlier investigations than in the current one, suggesting that it is dependent on agro-ecological variation. After caffeine, trigonelline, often referred to as N-methylpyridinium-3-carboxylate, is the second most prevalent alkaloid in coffee [ 29 ]. Gichimu et al [ 18 ] state that trigonelline is mostly found in coffee, but it is also present in barley, corn, onions, peas, soybeans, and tomatoes. Table 2 Total phenolic, flavonoid, trigonelline, chlorogenic acid, and caffeine content of Ethiopian green coffee Arabica varieties cultivated in various agro-ecological zones. Bioactive compounds Jimma coffee Sidama coffee Yirgachefe coffee Nekemte coffee Hararge coffee TPC (mg GAE/100 g) 46.52 ± 0.82 a 44.31 ± 4.68 ab 34.25 ± 3.83 c 44.55 ± 3.28 ab 39.02 ± 2.93 bc TFC (mg QE/100 g) 45.10 ± 0.36 ab 37.30 ± 0.75 bc 37.80 ± 0.55 bc 50.10 ± 0.23 a 31.50 ± 0.57 c Caffeine (mg/100 g) 8.85 ± 0.00 b 8.23 ± 0.00 c 10.38 ± 0.00 a 10.18 ± 0.00 a 7.55 ± 0.00 e Chlorogenic acid (mg/100 g) 45.95 ± 0.01 a 45.29 ± 0.01 a 39.54 ± 0.01 b 46.39 ± 0.01 a 36.78 ± 0.02 c Trigonelline (mg/100 g) 12.88 ± 0.01 ab 13.56 ± 0.02 a 13.46 ± 0.01 a 11.78 ± 0.00 b 11.65 ± 0.00 b On a dry basis, the data are presented as mean + standard deviation (n = 3). In mean comparisons using Duncan's multiple range test, mean values within a row with different superscripts indicate a significant difference at p < 0.05 using one-way analysis of variance (ANOVA). GAE stands for gallic acid equivalent, QE for quercetin equivalent, TPC for total phenolic content, and TFC for total flavonoid content. Overall, differences in the growing environment circumstances (i.e., altitude, soil type, rainfall, and other agricultural techniques) can be associated to variations in the TPC, TFC, caffeine, chlorogenic acid, and trigonelline content among coffee regions [ 4 ]. For instance, compared to Hararge and the southern regions, the southwestern, western, and northwest regions experience more rainfall overall and a longer rainy season [ 46 ]. The TPC is greatly impacted by this [ 19 ]. Climate adaptation measures and other management circumstances may counteract these environmental-driven variations in crop quality. Effects on crop quality, however, have been the subject of fewer studies [ 3 ]. For example, the volatiles 2, 3-butanedione, 2, 3-pentanedione, 2-methylbutanal, and 2, 3-dimethylpyrazine were among the significant odor-active indicators of coffee that changed as a result of these regional differences [ 3 ]. Therefore, to anticipate coffee quality using evidence-based innovations of climate adaptation, it is required to evaluate the biochemical composition of coffee beans throughout time. In addition to assessing the impact of geographic origins on biochemical composition, this study is designed to add to the database. Conclusion Geographic origin and processing technique are thought to affect coffee's quality and biological makeup. The current study's findings demonstrated the significant bioactive chemicals and phenolic contents found in coffee from Ethiopia's five main production regions: Jimma/Limu, Sidama, Hararge, Nekemte, and Yirgachefe. Additionally, it was confirmed that roasting the coffee beans decreased their bioactive chemicals. The five coffee bean cultivars have varying amounts of total phenolic and flavonoid content; the Jimma and Nekemte beans have the highest amounts. Yirgachefe coffee beans had the highest caffeine content, whereas Hararge coffee beans had the lowest. The Hararge variety of coffee had the lowest amount of chlorogenic acid, while the Jimma, Sidama, and Nekemte varieties had the greatest levels. Similarly, the coffee kinds with the greatest trigonelline concentrations were Yirgachefe, Sidama, and Jimma. The total TPC and other bioactive components have been considerably decreased after roasting all of the coffee beans. The amount of chlorogenic acid was reduced by the largest percentage after roasting. Nevertheless, roasting did not significantly alter the caffeine level of the various coffee varietals. For evidence-based climate change adaptation aimed at producing high-quality coffee, updated data on the biochemical makeup of coffee varietals throughout time can be utilized. This study proved the efficient methods and techniques of Ethiopian traditional coffee processing, starting with the collection, roasting, grinding, brewing, and services of coffee, together with the whole range of materials utilized in the operations. The overall green and roasted coffee's total phenolic and bioactive chemical content was compared with the growth area and roasting influence. Abbreviations BMI ICO HPLC TPC TRG CAF CGA Body Mass Index International Coffee Organization High Performance Liquid Chromatogram Total Phenolic Content Trigonelline Caffeine Chlorogenic Acid Declarations Consent to Publish: Not applicable, as this study does not involve any personal data, images, or identifiable information requiring individual consent. Consent to Participate: Not applicable. Ethics Declaration Not applicable. Conflicts of Interest The authors have no conflict of interest Funding Statement The present research has been supported by the Addis Ababa University and Ethiopian Agricultural Research Institute. Author Contribution Daba Mengesha Adula conceptualized and designed the research study, developed the methodology, and led the data collection from various coffee-growing regions in Ethiopia. He coordinated sample preparation and laboratory analysis for phenolic content and bioactive compounds using UV-Vis spectrophotometry and HPLC techniques. Daba performed the data analysis, interpreted the results, and wrote the original draft of the manuscript. He also oversaw the critical revision, responded to reviewer comments, and finalized the manuscript for submission. All aspects of the research—from experimental design to manuscript preparation—were conducted under his primary supervision and leadership. Acknowledgement AcknowledgementsThe author would like to express sincere gratitude to his supervisors for their continuous guidance, encouragement, and critical insights throughout the course of this research. Their mentorship has been instrumental in shaping the direction and quality of this work.Special thanks are extended to the laboratory technicians and support staff at Ethiopian Institute of Agricultural Research and and Addis Ababa University / food science and nutrition laboratory, whose technical assistance in sample preparation and analytical procedures greatly contributed to the successful completion of this study.I am also deeply thankful to my family and friends for their unwavering support, patience, and motivation during the research and writing process. Their encouragement provided strength during challenging moments and helped bring this work to fruition. Data Availability Statement Data will be made available on request. 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Valorization of coffee leaves as a potential agri-food resource: bio-active compounds, applications and future prospective. Planta. 2022;255(3):1–17. https://doi.org/10.1007/s00425-022-03846-x . Rifai L, Saleh FA. A Review on Acrylamide in Food: Occurrence, Toxicity, and Mitigation Strategies. Int J Toxicol. 2020;39(2):93–102. https://doi.org/10.1177/1091581820902405 . Rojas-González A, Figueroa-Hernández CY, González-Rios O, Suárez-Quiroz ML, González-Amaro RM, Hernández-Estrada ZJ, Rayas-Duarte P. Coffee Chlorogenic Acids Incorporation for Bioactivity Enhancement of Foods: A Review. Molecules. 2022;27(11):1–23. https://doi.org/10.3390/molecules27113400 . Sualeh A, Tolessa K, Mohammed A. (2020). Biochemical composition of green and roasted coffee beans and their association with coffee quality from different districts of southwest Ethiopia. Heliyon, 6(12). Schenker S, Heinemann C, Huber M, Pompizzi R, Perren R, Escher F. 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Exploring Arabica coffee cup quality: Correlations with green bean growing conditions, physicochemical properties, biochemical composition, and volatile aroma compounds. J Agric Food Res. 2024;18(November). https://doi.org/10.1016/j.jafr.2024.101549 . Velásquez S, Banchón C. Influence of pre-and post-harvest factors on the organoleptic and physicochemical quality of coffee: a short review. J Food Sci Technol. 2023;60(10):2526–38. https://doi.org/10.1007/s13197-022-05569-z . Wierzejska RE. Dietary supplements—for whom? The current state of knowledge about the health effects of selected supplement use. Int J Environ Res Public Health. 2021;18(17). https://doi.org/10.3390/ijerph18178897 . Worku W. (2024). Introductory Plant Physiology-Compiled lecture notes College of Agriculture School of Plant and Horticultural Sciences Compiled lecture notes Introductory Plant Physiology Compiled by . August . Worku M, Astatkie T, Boeckx P. Quality and biochemical composition of Ethiopian coffee varied with growing region and locality. J Food Compos Anal. 2023;115:105015. Santiago, W. D., Teixeira, A. R., de Andrade Santiago, J., Lopes, A. C. A., Brandao,R. M., Caetano, A. R., … Resende, M. L. V. (2020). Development and validation of chromatographic methods to quantify organic compounds in green coffee ('Coffea arabica') beans. Australian Journal of Crop Science, 14(8), 1275–1282. Additional Declarations No competing interests reported. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7194131","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":508319029,"identity":"0db8a54f-2f57-4a42-8caf-a1cb9b28fb46","order_by":0,"name":"Dhaba Mengesha Adula","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA4ElEQVRIiWNgGAWjYDCCGwxsQJKZgY+B+QCQISFDvBY2BrYEkBYeUrTwGID4hLXw3W5+9uBHjXViG/uZz69u1FjwMLAfProBnxbJO8fMDXuOpSe28eRus845BnQYT1raDXxaDG4kmEnwsB1ObGPI3WacwwbUIsFjRkBL+jfJP/+AWvjfPDPO+UeUlhwzad42oBaJHObHuW1EaJG8kVNuLNuXbtwm8cyMObcP6EhCfuG7kb7t4Ztv1rL9/MmPP+d8q5PjZz98DK8WZMAmASaJVQ4CzB9IUT0KRsEoGAUjBwAAIKhIo/B+Vy4AAAAASUVORK5CYII=","orcid":"","institution":"Ethiopian Institute of Agricultural Research (EIAR)","correspondingAuthor":true,"prefix":"","firstName":"Dhaba","middleName":"Mengesha","lastName":"Adula","suffix":""}],"badges":[],"createdAt":"2025-07-23 08:53:09","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7194131/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7194131/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":90411065,"identity":"50723937-fe66-472a-8292-7123d6af235f","added_by":"auto","created_at":"2025-09-02 12:12:46","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":822569,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7194131/v1/b6cddd05-3841-4206-a566-68aa0b91bafa.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Effects of Conventional Processing Methods and Growing Locations on the Phenolic Content and Bioactive Compounds of Ethiopian Coffee Beans","fulltext":[{"header":"Introduction","content":"\u003cp\u003eCoffee is becoming increasingly popular as a daily beverage and as a crop crucial to Ethiopian and international trade. In Ethiopia and around the world, coffee is one of the most commercially significant crops and one of the most culturally popular daily beverages. Its consumption is also rising these days. According to a survey by the International Coffee Organization (ICO), 4\u0026nbsp;billion cups of coffee, or 9.4\u0026nbsp;million tons, were consumed worldwide in 2016\u0026ndash;2017. These cups are made using various brewing techniques. Coffee is composed of several chemical constituent combinations that influence its flavor and aroma. These consist of alkaloids, phenolic compounds, vitamins, minerals, carbohydrates, lipids, and nitrogenous chemicals. In addition, coffee contains a lot of health benefits and is rich in bioactive substances including trigonelline, caffeine, and chlorogenic acid [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. Drinking coffee has been linked to a lower risk of hepatocellular carcinoma and vascular diseases [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e], anti-proliferative effects on certain human cancer cells [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e], and potential therapeutic benefits against Alzheimer's disease, liver disease, and diabetes[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e, \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe entire coffee process, from selecting coffee beans to choosing the best water for brewing to serving the coffee, affects the end product's quality and sensory attributes. To achieve a higher sensory score, coffee must have at least five distinct flavor notes in harmony [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. The Specialty Coffee Association of America [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e], criterion analysis procedure states that HPLC is used to analyze the bioactive compounds in roasted and brewed coffee. The coffee's ability to pass aspect or preliminary grading and cupping tests can be used to assess its sensory and cup quality [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Important steps that affect the quality and quality of the final coffee product include sorting and grading green coffee beans, roasting, grinding, and different extraction methods [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eDepending on the desired qualities of the coffee cup, dry coffee beans are roasted by heating them to temperatures ranging from 200 to 240\u0026deg;C for varying amounts of time [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Green beans lose a lot of water, which makes them fragile. Numerous metabolic pathways, including Strecker and Maillard, produce over 1000 distinct kinds of aromatic chemicals [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. Alcohols, aldehydes, amines, carboxylic acids, dicarbonyls, enoles, esters, furans, furanone, hydrocarbons, imidazoles, indoles, ketones, lactones, oxazoles, phenols, pyrazines, pyridines, pyrroles, quinoxalines, sulfur compounds, terpenes, and thiazoles are among the volatile substances found in roasted coffee beans [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e]. Depending on the heat profile used during roasting, these chemicals may experience significant modifications. For this reason, roasting is thought to be the most crucial process in identifying the coffee bean's distinct flavor and color. The different roasting conditions have a big impact on the physical and chemical properties of roasted coffee beans.\u003c/p\u003e\u003cp\u003eAfter roasting, the coffee beans are ground to maximize their surface area and regulate humidity for effective extraction. While the caffeine content does not significantly drop after roasting, 20\u0026ndash;40% of the polysaccharides that store cell walls are broken down [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e]. According to Konstantinidis [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e], trigonelline's primary metabolites, nicotinic acid and N-methylpyridinium, are helpful markers of roasting level. Metabolites created by microbes may infiltrate into the beans after roasting to escape the heat process. The metabolites with the greatest ability to affect the final coffee beverage's quality are flavor-active esters.\u003c/p\u003e\u003cp\u003eFollowing roasting, complex physical and chemical changes, including caramelization, are produced by the Maillard and Strecker process, which involves hundreds of biological components [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. The Maillard reaction, which transforms sugar into color, flavor, and fragrance, is catalyzed by amino acids. Acetic and formic acids are primarily responsible for the potent scent during the initial roasting stages. As quinic acid concentration rises, the breakdown of chlorogenic acids results in astringent and bitter flavors [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Roasting significantly lowers trigonelline and CGA while leaving caffeine unchanged [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Green beans' trigonelline, carbs, and some proteins also decompose into volatile chemicals, claim [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eCoffee beans get their distinctive dark color from melanoidin, which is a byproduct of the Maillard reactions and may help preserve the flavor components [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. While trigonelline helps produce both acceptable and disagreeable smell molecules during roasting, caffeine accounts for more than 30% of the bitter taste [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Asparagine and glucose-fructose can promote the Maillard reactions, which result in the production of unwanted substances like furan and acrylamides [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. The Strecker degradation, a component of the Maillard reaction network, contributes volatile aldehydes to the coffee's aroma spectrum, including notes of honey, sulfur, malt, and potato.\u003c/p\u003e\u003cp\u003eCoffee origin, grinding size, and brewing time all affect the production of coffee aromas, the composition of bioactive components, and the antioxidant potential of brewed coffee. Coffee's flavor and aroma can change during the preparation process due to its biochemical makeup [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eEthiopian coffee is processed using very healthful methods, employing locally produced elements that are very tasty to consumers and quite beneficial after intake. The quality of Ethiopian coffee beans is therefore influenced by a variety of factors, including the growing environment, processing methods, and particularly the roasting process. Though there are some gaps in the current research on the efficient coffee processing methods and effects of coffee origin on the phenolic and bioactive compounds of brewed coffee, the coffee's quality and taste can also be greatly influenced by the coffee's origin and brewing materials. The majority of current studies, for instance, have concentrated on how coffee origin and processing methods affect the biochemical composition and cup quality of coffee beans from other nations. Furthermore, the majority of previous studies have employed rather small sample sizes; additionally, most of the existing research has used relatively small sample sizes, which makes it difficult to generalize the findings to the wider population.\u003c/p\u003e\u003cp\u003eEthiopian traditional coffee processing techniques and their efficacy, in addition to the growing environment, affect the physico-chemical characteristics and cup quality of the brewed coffee. Not all of the main Ethiopian coffee beans have had their bioactive components and phenolic contents thoroughly examined regarding the effects of cultivation circumstances and efficient processing methods. This study was carried out to ascertain how the bioactive composition and physicochemical characteristics of Ethiopian coffee, which is traditionally roasted and brewed, were affected by the growing environment and processing stages. This study aimed to evaluate the effects of processing procedures and growing locations on the bioactive composition, physicochemical properties, and sensory acceptability of Ethiopian brewed coffee.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cb\u003eCoffee beans\u003c/b\u003e\u003c/p\u003e\u003cp\u003eEthiopian coffee beans were gathered from the Ethiopian Coffee and Tea Authority Coffee Quality Inspection and Certification Center (ECTACQICC) in Addis Ababa, Ethiopia, representing various Jimma, Hararge, Sidama, Nekemte, and Yirgachefe origins [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. Three kilograms of each type of coffee were gathered. After cleaning and washing the coffee beans, any damaged seeds were taken out.\u003c/p\u003e\u003cp\u003e\u003cb\u003eStandardizing the roasting and brewing process for Ethiopian traditional coffee\u003c/b\u003e\u003c/p\u003e\u003cp\u003eIn ten Addis Ababa sub-cities, the traditional Ethiopian coffee-processing method was surveyed. Addis Ketema, Arada, Bole, Gullele, Kirkos, Kolfe Keranio, Ledeta, Knife\u0026rsquo;s Silk Lafto, Akaky Kaliti, and Yeka were the ten sub-cities from which ten homes were chosen at random to participate in the study. The brewing process was then interviewed using questionnaires in each home. Actual measurements (weighed records) were also taken when needed. Mengesha et al [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e], constructed a coffee processing flowchart based on the survey and simulated it in a lab. This consistent procedure was used to prepare all of the brewed and roasted coffee powder samples.\u003c/p\u003e\u003cp\u003e\u003cb\u003eRoasting\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe coffee beans were roasted in a metal roasting pan measuring 16 cm in diameter and 76 mm in thickness. Charcoal cooking required 10 to 12 minutes, whereas electric roasting took 6 to 9 minutes at 190\u0026deg;C. The beans were roasted and then allowed to cool at room temperature for 11 minutes before being ground using a traditional wooden mortar and pestle to a sieve size of 1.2 mm.\u003c/p\u003e\u003cp\u003e\u003cb\u003eCoffee brewing\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe coffee bean was roasted for 16 minutes at a maximum temperature of 190\u0026deg;C, which is the standard and advised level of roast [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. The exothermic reaction was then terminated by leaving the air-cooled coffee beans overnight to allow for full taste development and CO2 degassing. The roasted coffee beans were ground into fine (2 mm), medium (4 mm), and coarse (6 mm) powder sizes using an electronic laboratory-scale coffee grinder (VTA6S, MAHLK\u0026Ouml;NIG GmbH \u0026amp; Co., 2009) [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Following that, the ground coffee samples were placed in airtight, odor-proof polyethylene bags. Before brewing and analysis, the sample was kept at 4\u0026deg;C and 70% relative humidity. Coffee was made using surface water and 1:17 (w/w) ratios of coffee powder to water, as per the method described by [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eSample extraction for bioactive compounds\u003c/b\u003e\u003c/p\u003e\u003cp\u003eTo extract the total phenolic, flavonoid content, and bioactive components present in green and roasted coffee, the methodology of Iriondo-Dehond et al [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e] was adopted. In summary, 3 g of ground green coffee was mixed with 20 mL of 98% methanol. Each coffee sample was mixed with twenty milliliters of 98% methanol. The samples were placed in Erlenmeyer flasks in a shaking incubator (Lucadema, Brazil) for 30 minutes at 40\u0026deg;C and 120 rpm, and then centrifuged for 15 minutes at 3200 rpm in an Excelsa Baby II centrifuge (Flamen, model 206-R). The supernatant's phenolic compounds were detected. Three extractions were performed.\u003c/p\u003e\u003cp\u003e\u003cb\u003eDetermination of total polyphenol content (TPC)\u003c/b\u003e\u003c/p\u003e\u003cp\u003eAccording to Hidayat et al [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e], the Folin-Ciocalteu method was used to estimate the TPC. Gallic acid was used to generate the TPC measuring standard curve. Gallic acid equivalent (GAE)/g of the dry matter of coffee samples was used to express the results. One milliliter of 99% ethanol was added to standard solutions of gallic acid, which included 2, 4, 6, 8, 10, and 12 mg of gallic acid. After that, three milliliters of distilled water were added. Next came the addition of 0.1 mL of Folin-Ciocalteu reagent. Three minutes later, distilled water was added to the volumetric flasks to bring them up to the mark, and then 0.3 mL of a 2% sodium carbonate solution was added. The entire material was stored for two hours at 25\u0026deg;C. The gallic acid solution was then substituted with 100 mL of ethanol. The spectrophotometer's wavelength was set to 765 nm after two hours. The absorbance of each solution was measured three times, and the average was recorded after the device was initially zeroed with the control (gallic acid) solution. Following the development of a standard curve that plotted concentration (\u0026micro;g) against absorbance, the line equation was applied to the quantification.\u003c/p\u003e\u003cp\u003eTen milligrams of the green bean extract were weighed, placed in a test tube, and then completely dissolved with two milliliters of dimethyl sulfoxide (DMSO) solution to create the coffee samples. Triplicate assays were performed on each extracted sample using three 5 mL volumetric flasks. There were 0.20 mL of each extract and 3 mL of distilled water in each volumetric flask. The volumetric flasks were then filled with 100 \u0026micro;L of Folin\u0026ndash;Ciocalteu reagent, and three minutes later, 0.3 mL of sodium carbonate (2%) was added. Distilled water was added to the volumetric flasks to bring them to the mark.\u003c/p\u003e\u003cp\u003eInstead of the extract, 0.20 mL of DMSO was added to a 5 mL control volumetric flask. The mixture was homogenized and then stored for two hours at 25\u0026deg;C. After two hours, the absorbance of each solution was measured three times at 765 nm using a UV-Vis spectrophotometer (PerkinElmer, Model CF728YW-950, UK). The TPC was expressed as \u0026micro;g of GAE/g of extract using Eq.\u0026nbsp;\u003cspan refid=\"Equ1\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003cdiv id=\"Equ1\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ1\" name=\"EquationSource\"\u003e\n$$\\:TPC=\\:\\frac{C*V}{M}\\:\\times\\:\\:D$$\u003c/div\u003e\u003cdiv class=\"EquationNumber\"\u003e1\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eWhere;\u003c/p\u003e\u003cp\u003eC\u0026thinsp;=\u0026thinsp;gallic acid equivalent concentration obtained from the calibration curve (g/mL)\u003c/p\u003e\u003cp\u003eV\u0026thinsp;=\u0026thinsp;volume of the extract's stock solution (mL)\u003c/p\u003e\u003cp\u003eM\u0026thinsp;=\u0026thinsp;dry weight of the extract in the stock solution (g)\u003c/p\u003e\u003cp\u003eTPC\u0026thinsp;=\u0026thinsp;Phenolic content expressed as (mg of GAE/g dry extract)\u003c/p\u003e\u003cp\u003eD\u0026thinsp;=\u0026thinsp;dilution factor\u003c/p\u003e\u003cp\u003e\u003cb\u003eBioactivity Analysis\u003c/b\u003e\u003c/p\u003e\u003cp\u003e\u003cb\u003eCaffeine, trigonelline, and chlorogenic acid extraction\u003c/b\u003e\u003c/p\u003e\u003cp\u003eA significantly modified version of the [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e], technique was used to extract CAF, CGA, and TRG. A 50 mL Erlenmeyer flask containing roughly 0.5 g of finely ground coffee grounds was carefully weighed. Stir on a hot plate for 20 minutes after adding 50 mL of heated (95\u0026deg;C) pure water. 10 \u0026micro;L of the extract was placed in an Agilent 1260 Infinity, Germany, high-performance liquid chromatography (HPLC) device after it had been filtered through No. 4 What-man filter paper and a 0.22 \u0026micro;m pore size filter.\u003c/p\u003e\u003cp\u003e\u003cb\u003eMeasurement of caffeine, trigonelline, and chlorogenic acids\u003c/b\u003e\u003c/p\u003e\u003cp\u003eUsing a method adapted from [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e], TRG, CGA, and CAF were measured concurrently using an HPLC system with a C18 column and an isocratic flow of 0.7 mL/min. A 4.6 \u0026times; 250 mm column with a 5 \u0026micro;m particle size was used by Waters, Taunton, USA. At a flow rate of 0.7 mL/min, the following elute compounds were present in gradients with 5% acetic acid (A) and acetonitrile (B): 4% B for 0\u0026ndash;4 minutes, 10% for 4\u0026ndash;8 minutes, 90% for 8\u0026ndash;12 minutes, 0% for 12\u0026ndash;15 minutes, and 4% for 15\u0026ndash;17 minutes. A three-minute post-run was held. TRG and CAF were measured at 272 nm, but CGA was detected at 320 nm. TRG and CAF test concentrations were 5, 10, 20, 40, 50, 100, and 150 \u0026micro;g/mL. The ranges for CGA, on the other hand, were 5, 10, 20, 100, 150, and 200 \u0026micro;g/mL. The identities of CAF, TRG, and CGA were ascertained by comparing the retention durations of the TRG (Sigma Aldrich), CGA (Acros Organics), and CAF (Fischer Scientific) standards (99%), as well as their concentrations derived from peak regions using calibration equations. Following a time of retention, the area was utilized to determine the composition of the related biological component, which was then, determined using calibration curves that plotted the area against standard concentration [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e]. The limit of detection (LOD) for caffeine, trigonelline, and chlorogenic acid was 0.9, 1.02, and 1.5 \u0026micro;g/g, respectively and the limits of quantification (LOQ) for caffeine, trigonelline, and chlorogenic acid was 1.7, 2.3 and 3.0 \u0026micro;g/g, respectively.\u003c/p\u003e\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eStatistical analysis\u003c/h2\u003e\u003cp\u003eThe study employed two-factor factorial experimental designs with three treatment levels. Roasting and coffee-producing areas (A and B) were the chosen criteria. Five coffee origins\u0026mdash;Jimma, Sidama, Yirgachefe, Hararge, and Nekemte coffee\u0026mdash;were employed for brewing purposes- and green and roasting methods. At least three observations were made during the fully randomized experimental design. Using SPSS software version 26, the collected experimental findings were examined and interpreted using analysis of variance (ANOVA) at a 5% significance level.\u003c/p\u003e\u003c/div\u003e"},{"header":"Results and discussion","content":"\u003cp\u003e\u003cb\u003eStandardized Ethiopian coffee brewing method\u003c/b\u003e\u003c/p\u003e\u003cp\u003eMost Ethiopians drink coffee prepared by the traditional processing method, which varies slightly from home to home. Thus, this study's main goal was to simulate in the laboratory a standardized flowchart for traditional coffee processing Mengesha et al [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] using data from a qualitative survey that was carried out in each of Addis Ababa's ten sub-cities. The acrylamide levels were then measured after coffee was prepared in a lab environment using this flowchart.\u003c/p\u003e\u003cp\u003eThe Ethiopian traditional method of preparing coffee involves weighing 30 to 35 grams of coffee beans per serving, cleaning them, and washing them with clean tap water. After that, the beans were roasted in a metal roasting pan that measured an average of 16 cm in diameter and 76 mm in thickness. It took 6 to 9 minutes to roast at 190\u0026deg;C with electric power, and 10 to 12 minutes to cook using charcoal. A traditional wooden mortar and pestle was used to grind the beans to a sieve size of 1.2 mm after they had been roasted and allowed to cool for 11 minutes at room temperature. After that, the ground coffee was brewed in a traditional clay kettle, or boiling pot, using 25\u0026ndash;30 grams of coffee for every 750\u0026ndash;800 milliliters of water. The brewing procedure took 8 to 12 minutes, and the temperature ranged from 92 to 95\u0026deg;C. Before serving, the coffee was allowed to decant at room temperature for five minutes after boiling. The grinds are brewed three times using the same method in Ethiopian traditional coffee processing. This implies that two more brews are made using the same coffee grounds and the same amount of water following the first brew. \"Abol,\" \"Tona,\" and \"Baraka\" are the local terms for the first, second, and third rounds of coffee, respectively. Nonetheless, the survey indicates that consumers frequently favor the first and second brews. Therefore, another objective of this study was to determine the amount of acrylamide in the two serving rounds.\u003c/p\u003e\u003cp\u003eThe main way that people are exposed to acrylamide is through their diet [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. Bread, coffee, and French fries have been repeatedly identified as major contributors to dietary acrylamide exposure in several studies that have looked at exposure among people with different characteristics [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. The need for additional study on dietary acrylamide exposure and related health hazards is emphasized by the European Commission and the Joint FAO/WHO Expert Committee on Food Additives (European Commission, 2019; Joint FAO/WHO Expert Committee on Food Additives, 2011). The amount of acrylamide in brewed and powdered coffee from commercially processed and street-vendor sources varies significantly. Therefore, to accurately quantify the exposure to acrylamide from coffee drinking, it is imperative to follow the most commonly used processing methods.\u003c/p\u003e\u003cp\u003e\u003cb\u003eImpact of Ethiopian traditional coffee roasting on the biochemical contents\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe impact of roasting Ethiopian coffee on the biochemical makeup of coffee beans from various agroecological origins is documented in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. The caffeine concentration ranged from 7.55 to 10.38 mg/mL for the five Ethiopian Coffee Arabica cultivars. Yirgachefe coffee beans had the highest caffeine content, while Hararge coffee beans had the lowest (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). All cultivars have a substantial variation in caffeine concentration (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), except for Nekemte and Yirgachefe. As evidence for this work, several investigations have reported varying quantities of caffeine in coffee beans [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. For instance, Mengistu et al [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e] used HPLC techniques to find that the caffeine concentrations in 42 Ethiopian coffee samples ranged from 9.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01 to 12.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06 mg/mL, with an average value of 10.10 mg/mL. Other studies have shown that the average caffeine content of Arabic coffee is less than 15 mg/mL [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. The caffeine concentration of this study is higher than that of Farah [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e], investigation, which discovered that the caffeine content of various coffee varieties ranges from 9 to 25 mg/g. The highest levels of chlorogenic acid (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05) were found in the Jimma, Sidama, and Nekemte coffee varietals (45 mg/mL) in comparison to the other two. With 36.78 mg/g of chlorogenic acid, the Hararge cultivar had the lowest levels. Chlorogenic acid levels in several coffee bean varietals ranged from 4.1 to 11.3 mg/g, according to [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. This number is greater than the one found in the current study. Trigonelline levels were also highest in the Jimma, Sidama, and Nekemte coffee varietals, with 12.88, 13.56, and 13.46 mg/mL, respectively (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05). Hararge and Yirgachefe kinds had the lowest concentrations, with respective values of 11.65 and 11.78 mg/mL (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e\u003cp\u003eEvery home uses Ethiopian coffee, which is roasted and brewed distinctively. Reports on its impact on the biochemical makeup of the regional coffee types are, nevertheless, few. As seen in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, roasting significantly decreased the TPC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) for all coffee beans. The coffee beans from Jimma (24%), Sidama (26%), Nekemte (30%), Yirgacheffe (23%), and Hararge (29%), had the lowest percentage of TPC. On average, there was a 27% drop. The caffeine content of the Sidama, Nekemte, and Yirgacheffe coffee beans varied significantly after roasting (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Among all coffee varietals, the chlorogenic acid content had the largest significant percentage decrease (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Jimma (83%), Sidama (79%), Nekemte (82%), Yirgacheffe (81%), and Hararge (82%), were the countries with the largest percentage decreases. The average reduction was 81% overall. Furthermore, upon roasting, there were significant differences in the trigonelline content across all coffee kinds (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). With Jimma (54%), Sidama (29%), Nekemte (45%), Yirgacheffe (38%), and Hararge (34%), the average reduction was 40%.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eEffects of traditional Ethiopian coffee bean roasting on the levels of caffeine, trigonelline, chlorogenic acid, and total phenols in green and roasted Ethiopian Coffee cultivars in various agro-ecological zones.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eCoffee origin\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eCoffee type\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eTotal polyphenols (mg GAE/g)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eCaffeine (mg/mL)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eChlorogenic acid (mg/mL)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eTrigonelline (mg/mL)\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eJimma\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGreen coffee\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e46.52\u0026thinsp;\u0026plusmn;\u0026thinsp;0.82\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8.85\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e45.95\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e12.88\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eRoasted\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e35.56\u0026thinsp;\u0026plusmn;\u0026thinsp;1.08\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7.84\u0026thinsp;\u0026plusmn;\u0026thinsp;1.08\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e5.88\u0026thinsp;\u0026plusmn;\u0026thinsp;1.00\u003csup\u003ef\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eSidama\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGreen coffee\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e44.31\u0026thinsp;\u0026plusmn;\u0026thinsp;4.68\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8.23\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e45.29\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e13.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eRoasted\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e32.87\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8.11\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e9.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.91\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e9.60\u0026thinsp;\u0026plusmn;\u0026thinsp;0.66\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eNekemte\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGreen coffee\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e44.55\u0026thinsp;\u0026plusmn;\u0026thinsp;3.28\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e10.18\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e46.39\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e13.46\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eRoasted\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e31.36\u0026thinsp;\u0026plusmn;\u0026thinsp;0.19\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e9.88\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e8.43\u0026thinsp;\u0026plusmn;\u0026thinsp;0.69\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e7.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.67\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eYirgachefe\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGreen coffee\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e34.25\u0026thinsp;\u0026plusmn;\u0026thinsp;3.83\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e10.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e39.54\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e11.78\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eRoasted\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e26.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.91\u003csup\u003ef\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e9.92\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7.66\u0026thinsp;\u0026plusmn;\u0026thinsp;0.84\u003csup\u003eef\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e7.27\u0026thinsp;\u0026plusmn;\u0026thinsp;1.19\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eHararge\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGreen coffee\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e39.02\u0026thinsp;\u0026plusmn;\u0026thinsp;2.93\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e7.55\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ee\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e36.78\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e11.65\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eRoasted\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e27.86\u0026thinsp;\u0026plusmn;\u0026thinsp;1.01\u003csup\u003eef\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e7.45\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003csup\u003ee\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e6.46\u0026thinsp;\u0026plusmn;\u0026thinsp;0.89\u003csup\u003ef\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e7.64\u0026thinsp;\u0026plusmn;\u0026thinsp;0.64\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAverage\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGreen coffee\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e41.73\u0026thinsp;\u0026plusmn;\u0026thinsp;0.43\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e9.51\u0026thinsp;\u0026plusmn;\u0026thinsp;0.66\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e42.79\u0026thinsp;\u0026plusmn;\u0026thinsp;0.67\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e12.67\u0026thinsp;\u0026plusmn;\u0026thinsp;0.47\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eRoasted\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e30.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.75\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.45\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7.95\u0026thinsp;\u0026plusmn;\u0026thinsp;0.46\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e7.54\u0026thinsp;\u0026plusmn;\u0026thinsp;1.34\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eOn a dry basis, the data are presented as mean\u0026thinsp;+\u0026thinsp;SD (n\u0026thinsp;=\u0026thinsp;3). Significant differences at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 in mean comparison using the Duncan multiple range test are shown by mean values with different superscript letters within a column. *Uses the paired t-test to show a significant difference between green and roasted coffee beans (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05).\u003c/p\u003e\u003cp\u003eThe average CGA level in the current study ranged from 45.95 to 36.78 mg/g on a dry weight basis, while the average caffeine content ranged from 10.38 to 8.23 mg/mL. According to Farah [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e], these values are often in the lower range of values previously observed for green Arabica coffee beans. Our caffeine measurements were marginally higher for both CAF and CGA than those published by [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eChlorogenic acid content in brewed and roasted coffee ranged from 16 to 33 percent. This is in line with the range discovered by Caprioli et al [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e], who demonstrated that spent and roasted coffee still had 23\u0026ndash;68% chlorogenic acid. 23\u0026ndash;36% of the caffeine was still present in the wasted coffee. Retention rates for Gallic and protocatechuic acids were lower in comparison, at 4\u0026ndash;13 and 4\u0026ndash;24%, respectively. These findings show that the retention rates of each component differ according to its water solubility and high-temperature stability.\u003c/p\u003e\u003cp\u003eChlorogenic acids are the primary phenolic components found in green coffee. According to Mehari et al [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e], they are esters of quinic acid and trans-hydroxycinnamic acid. It is known that these chemical groups are responsible for the color, flavor, bitterness, and astringency of coffee beverages [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. About 6\u0026ndash;12% of green coffee beans' dry weight is composed of chlorogenic acids [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. When compared to the other two coffee kinds in this investigation, the Jimma, Sidama, and Nekemte varietals had the greatest levels of chlorogenic acid (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05), at 45 g/100 g. With 36.78 g/100 g of chlorogenic acid, the Hararge variety exhibited the lowest levels (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). There is an inverse relationship between a coffee variety's chlorogenic acid concentration and beverage quality; coffee types with lower beverage quality had higher chlorogenic acid concentrations [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. The green coffee beans gathered from nine districts in Southwest Ethiopia have a chlorogenic acid level ranging from 2.80 to 5.42 g/100 g [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. According to Urugo et al [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e], the amount of chlorogenic acids in green coffee beans gathered from northern Ethiopia ranged from 3.29 to 7.73 g/100 g. According to Farah [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e], the amounts of chlorogenic acid in various coffee types ranged from 4.1 to 11.3 g/100 g. Compared to the current study, the chlorogenic acid content was lower in all of these earlier investigations. According to Awwad et al [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e], chlorogenic acids exhibit potent antiviral, antidiabetic, antioxidant, and neuroprotective properties.\u003c/p\u003e\u003cp\u003eAccordingly, the highest trigonelline levels were found in the Jimma, Sidama, and Yirgachefe coffee varietals, with 12.88, 13.56, and 13.46 mg/100 g, respectively (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05). The Nekemte and Hararge types had the lowest concentrations, measuring 11.78 and 11.65 mg/100 g, respectively (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05) (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). The amount of trigonelline in coffee beans gathered from nine districts in Southwest Ethiopia varied significantly (from 0.80 to 1.08 g/100 g), according to [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Trigonelline levels in green coffee beans gathered from the country's north ranged from 0.53 to 1.27 g/100 g, according to [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. Trigonelline levels were lower in these earlier investigations than in the current one, suggesting that it is dependent on agro-ecological variation. After caffeine, trigonelline, often referred to as N-methylpyridinium-3-carboxylate, is the second most prevalent alkaloid in coffee [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. Gichimu et al [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e] state that trigonelline is mostly found in coffee, but it is also present in barley, corn, onions, peas, soybeans, and tomatoes.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eTotal phenolic, flavonoid, trigonelline, chlorogenic acid, and caffeine content of Ethiopian green coffee Arabica varieties cultivated in various agro-ecological zones.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBioactive compounds\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eJimma coffee\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eSidama coffee\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eYirgachefe coffee\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eNekemte coffee\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eHararge coffee\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTPC (mg GAE/100 g)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e46.52\u0026thinsp;\u0026plusmn;\u0026thinsp;0.82\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e44.31\u0026thinsp;\u0026plusmn;\u0026thinsp;4.68\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e34.25\u0026thinsp;\u0026plusmn;\u0026thinsp;3.83\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e44.55\u0026thinsp;\u0026plusmn;\u0026thinsp;3.28\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e39.02\u0026thinsp;\u0026plusmn;\u0026thinsp;2.93\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTFC (mg QE/100 g)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e45.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.36\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e37.30\u0026thinsp;\u0026plusmn;\u0026thinsp;0.75\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e37.80\u0026thinsp;\u0026plusmn;\u0026thinsp;0.55\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e50.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.23\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e31.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.57\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eCaffeine (mg/100 g)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e8.85\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e8.23\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e10.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e10.18\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e7.55\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003ee\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eChlorogenic acid (mg/100 g)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e45.95\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e45.29\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e39.54\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e46.39\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e36.78\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTrigonelline (mg/100 g)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e12.88\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e13.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e13.46\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e11.78\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e11.65\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eOn a dry basis, the data are presented as mean\u0026thinsp;+\u0026thinsp;standard deviation (n\u0026thinsp;=\u0026thinsp;3). In mean comparisons using Duncan's multiple range test, mean values within a row with different superscripts indicate a significant difference at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 using one-way analysis of variance (ANOVA). GAE stands for gallic acid equivalent, QE for quercetin equivalent, TPC for total phenolic content, and TFC for total flavonoid content.\u003c/p\u003e\u003cp\u003eOverall, differences in the growing environment circumstances (i.e., altitude, soil type, rainfall, and other agricultural techniques) can be associated to variations in the TPC, TFC, caffeine, chlorogenic acid, and trigonelline content among coffee regions [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. For instance, compared to Hararge and the southern regions, the southwestern, western, and northwest regions experience more rainfall overall and a longer rainy season [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e]. The TPC is greatly impacted by this [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Climate adaptation measures and other management circumstances may counteract these environmental-driven variations in crop quality. Effects on crop quality, however, have been the subject of fewer studies [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eFor example, the volatiles 2, 3-butanedione, 2, 3-pentanedione, 2-methylbutanal, and 2, 3-dimethylpyrazine were among the significant odor-active indicators of coffee that changed as a result of these regional differences [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Therefore, to anticipate coffee quality using evidence-based innovations of climate adaptation, it is required to evaluate the biochemical composition of coffee beans throughout time. In addition to assessing the impact of geographic origins on biochemical composition, this study is designed to add to the database.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eGeographic origin and processing technique are thought to affect coffee's quality and biological makeup. The current study's findings demonstrated the significant bioactive chemicals and phenolic contents found in coffee from Ethiopia's five main production regions: Jimma/Limu, Sidama, Hararge, Nekemte, and Yirgachefe. Additionally, it was confirmed that roasting the coffee beans decreased their bioactive chemicals. The five coffee bean cultivars have varying amounts of total phenolic and flavonoid content; the Jimma and Nekemte beans have the highest amounts. Yirgachefe coffee beans had the highest caffeine content, whereas Hararge coffee beans had the lowest. The Hararge variety of coffee had the lowest amount of chlorogenic acid, while the Jimma, Sidama, and Nekemte varieties had the greatest levels. Similarly, the coffee kinds with the greatest trigonelline concentrations were Yirgachefe, Sidama, and Jimma. The total TPC and other bioactive components have been considerably decreased after roasting all of the coffee beans. The amount of chlorogenic acid was reduced by the largest percentage after roasting. Nevertheless, roasting did not significantly alter the caffeine level of the various coffee varietals. For evidence-based climate change adaptation aimed at producing high-quality coffee, updated data on the biochemical makeup of coffee varietals throughout time can be utilized.\u003c/p\u003e\u003cp\u003eThis study proved the efficient methods and techniques of Ethiopian traditional coffee processing, starting with the collection, roasting, grinding, brewing, and services of coffee, together with the whole range of materials utilized in the operations. The overall green and roasted coffee's total phenolic and bioactive chemical content was compared with the growth area and roasting influence.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"51%\" style=\"margin-right: calc(3%); width: 97%;\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 104px;\"\u003e\n \u003cp\u003eBMI\u003c/p\u003e\n \u003cp\u003eICO\u003c/p\u003e\n \u003cp\u003eHPLC\u003c/p\u003e\n \u003cp\u003eTPC\u003c/p\u003e\n \u003cp\u003eTRG\u003c/p\u003e\n \u003cp\u003eCAF\u003c/p\u003e\n \u003cp\u003eCGA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 255px;\"\u003e\n \u003cp\u003eBody Mass Index\u003c/p\u003e\n \u003cp\u003eInternational Coffee Organization\u003c/p\u003e\n \u003cp\u003eHigh Performance Liquid Chromatogram\u003c/p\u003e\n \u003cp\u003eTotal Phenolic Content\u003c/p\u003e\n \u003cp\u003eTrigonelline\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCaffeine\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eChlorogenic Acid\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eConsent to Publish:\u003c/strong\u003e\u003cp\u003eNot applicable, as this study does not involve any personal data, images, or identifiable information requiring individual consent.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eConsent to Participate:\u003c/strong\u003e\u003cp\u003eNot applicable.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eEthics Declaration\u003c/strong\u003e\u003cp\u003eNot applicable.\u003c/p\u003e\u003ch2\u003eConflicts of Interest\u003c/h2\u003e\u003cp\u003eThe authors have no conflict of interest\u003c/p\u003e\u003ch2\u003eFunding Statement\u003c/h2\u003e\u003cp\u003eThe present research has been supported by the Addis Ababa University and Ethiopian Agricultural Research Institute.\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eDaba Mengesha Adula conceptualized and designed the research study, developed the methodology, and led the data collection from various coffee-growing regions in Ethiopia. He coordinated sample preparation and laboratory analysis for phenolic content and bioactive compounds using UV-Vis spectrophotometry and HPLC techniques. Daba performed the data analysis, interpreted the results, and wrote the original draft of the manuscript. He also oversaw the critical revision, responded to reviewer comments, and finalized the manuscript for submission. All aspects of the research\u0026mdash;from experimental design to manuscript preparation\u0026mdash;were conducted under his primary supervision and leadership.\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eAcknowledgementsThe author would like to express sincere gratitude to his supervisors for their continuous guidance, encouragement, and critical insights throughout the course of this research. Their mentorship has been instrumental in shaping the direction and quality of this work.Special thanks are extended to the laboratory technicians and support staff at Ethiopian Institute of Agricultural Research and and Addis Ababa University / food science and nutrition laboratory, whose technical assistance in sample preparation and analytical procedures greatly contributed to the successful completion of this study.I am also deeply thankful to my family and friends for their unwavering support, patience, and motivation during the research and writing process. Their encouragement provided strength during challenging moments and helped bring this work to fruition.\u003c/p\u003e\u003ch2\u003eData Availability Statement\u003c/h2\u003e\u003cp\u003eData will be made available on request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003e(2020)., O. caffeine measurements were marginally higher for both C. and C. than those published by S. et al. (2022). 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Australian Journal of Crop Science, 14(8), 1275\u0026ndash;1282.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"discover-food","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"discoverfood","sideBox":"Learn more about [Discover Food](https://www.springer.com/44187)","snPcode":"","submissionUrl":"","title":"Discover Food","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Discover Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Bioactive Compounds, Growing Location, Green Coffee, Traditional Processing, Roasting","lastPublishedDoi":"10.21203/rs.3.rs-7194131/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7194131/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eGrowing conditions, roasting, and brewing methods are among the factors and procedures that determine the bioactive compound and phenolic content of brewed coffee. This study aimed to evaluate the impact of growing locations and traditional coffee processing methods on the bioactive components and phenolic content of Ethiopian coffee. The efficient Ethiopian traditional coffee processing procedures with the commonly used medium particle size of coffee powder and widely used water type (surface water) were used for brewing by the Ethiopian traditional coffee brewing method (190 \u003csup\u003e0\u003c/sup\u003eC of roasting, 16 min of brewing time). HPLC was used to assess the phenolic content and determine the bioactive chemicals. All five Ethiopian Coffee Arabica origins (Jimma, Sidama, Yirgachefe, Nekemte, and Hararge) had caffeine levels between 7.55 and 10.38 mg/mL. The coffee beans from Yirgachefe and Hararge had the greatest and lowest caffeine content, respectively (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). The caffeine level of each variety varies significantly (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), except for Nekemte and Yirgachefe. When compared to the other two coffee origins, the Jimma, Sidama, and Nekemte origins had the greatest levels of chlorogenic acid (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05), at 45 mg/mL. With 36.78 mg/g of chlorogenic acid, the Hararge cultivar had the lowest levels. Trigonelline levels were also highest in the Jimma, Sidama, and Nekemte coffee varietals, with 12.88, 13.56, and 13.46 mg/mL, respectively (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05). Hararge and Yirgachefe kinds had the lowest concentrations, with respective values of 11.65 and 11.78 mg/mL (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05). TPC was considerably lower after roasting all coffee beans (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). The following coffee beans have the lowest proportion of TPC: Jimma (24%), Sidama (26%), Nekemte (30%), Yirgachefe (23%), and Hararge (29%). On average, the reduction was a 27% factor. After roasting, the caffeine concentration of the Sidama, Nekemte, and Yirgachefe coffee beans varied significantly (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Jimma (83%), Sidama (79%), Nekemte (82%), Yirgachefe (81%), and Hararge (82%), among all coffee origins, had the highest significant percentage decrease in chlorogenic acid content (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). On average, there was an 81% decrease. Furthermore, there was a substantial variation in the trigonelline content across all coffee varietals during roasting (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). With Jimma (54%), Sidama (29%), Nekemte (45%), Yirgachefe (38%), and Hararge (34%), the average reduction was 40%. In general, the overall amount of bioactive components and phenolic contents in processed coffee is influenced by growing regions and coffee processing. Further research may be necessary to determine how coffee's phenolic content and bioactive components are affected by brewing temperature and extraction methods.\u003c/p\u003e","manuscriptTitle":"Effects of Conventional Processing Methods and Growing Locations on the Phenolic Content and Bioactive Compounds of Ethiopian Coffee Beans","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-02 11:56:41","doi":"10.21203/rs.3.rs-7194131/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-01-27T12:19:36+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-12-05T13:51:05+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"184979832953066153086815511743295579905","date":"2025-11-14T12:47:52+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-28T18:40:34+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"187719349154630235037278395534553443073","date":"2025-10-19T17:58:30+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-09-10T14:32:51+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"17560099615833210934394754339111024414","date":"2025-09-02T10:38:04+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"56061701992932536488324170878494030478","date":"2025-09-01T04:21:27+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"187540992477395936463839713117951443775","date":"2025-08-27T16:45:01+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-08-25T16:06:23+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-08-05T07:45:37+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-07-28T12:19:21+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-07-28T12:17:15+00:00","index":"","fulltext":""},{"type":"submitted","content":"Discover Food","date":"2025-07-23T08:37:49+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"discover-food","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"discoverfood","sideBox":"Learn more about [Discover Food](https://www.springer.com/44187)","snPcode":"","submissionUrl":"","title":"Discover Food","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Discover Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"80db6662-7659-4fc9-8bb4-435f4afdad14","owner":[],"postedDate":"September 2nd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-04-01T11:39:56+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-02 11:56:41","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7194131","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7194131","identity":"rs-7194131","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}