Revista Cienfica, FCV-LUZ / Vol. XXXV Recibido: 22/08/2025 Aceptado: 08/11/2025 Publicado: 19/12/2025 UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico 1 of 8 Revista Cienfica, FCV-LUZ / Vol. XXXVI https://doi.org/10.52973/rcfcv-e361779 UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico Comparave phytochemical composion and bioacvies of local variety of Prunus domesca L. from Northeastern Türkiye Composición fitoquímica comparava y bioacvidades de la variedad local de Prunus domesca L. del noreste de Turquía Vesile Duzguner 1,2 * , Altug Kucukgul 2 ¹Ardahan University, Faculty of Health Sciences, Ardahan, Türkiye ²Mustafa Kemal University, Faculty of Veterinary Sciences, Hatay, Türkiye *Correspondence author: vesileduzguner@ardahan.edu.tr ABSTRACT The Cancur plum (Prunus domesca L.), a local variety grown in Northeastern Türkiye, represents a valuable source of bioacve compounds with potenal funconal food applicaons. This study aimed to comprehensively evaluate the phytochemical composion and funconal properes of Cancur plum fruits collected from two regions with contrasng climac condions: Posof (microclimate, 1260 m) and Çıldır (connental climate, 1585 m). Anoxidant potenal was assessed using mulple approaches, including TAC, DPPH, glutathione, SOD, and catalase assays, while the quanes of total phenolics, flavonoids, and anthocyanins were also determined. The characterizaon of individual phenolic and flavonoid molecules was performed via LC-MS/MS, volale constuents were profiled through GC- MS, and free sugars were quanfied using HPLC. Comparave evaluaon showed that the Çıldır vinegars exhibited greater DPPH radical scavenging capacity and elevated catalase acvity, whereas the Posof vinegars were richer in total phenolics and glutathione. Despite these differences, both groups displayed a similar overall anoxidant capacity, though mediated by disnct biochemical mechanisms. LC-MS/MS profiling highlighted shikimic, chlorogenic, and p-coumaric acids as the predominant phenolics, with run and hesperidin occurring in higher amounts in the Çıldır sample. In terms of aroma-acve compounds, acec acid and acetoin dominated in the Çıldır vinegars, while ethanol and furfural were more pronounced in the Posof samples.Glucose and fructose were idenfied as the primary sugars, with minor sucrose detected only in Çıldır fruits. Microclimac differences strongly shape the biochemical and funconal profiles of Cancur plum, highlighng its value as a source of anoxidants, flavor compounds, and funconal food ingredients. Key words: Cancur plum; Prunus domesca L.; anoxidant; phenolic profile; volale compounds RESUMEN La ciruela Cancur (Prunus domesca L.), una variedad local culvada en el noreste de Turquía, es una valiosa fuente de compuestos bioacvos con posibles aplicaciones en alimentos funcionales. Este estudio tuvo como objevo evaluar exhausvamente la composición fitoquímica y las propiedades funcionales de los frutos de la ciruela Cancur recolectados en dos regiones con condiciones climácas contrastantes: Posof (microclima, 1260 m) y Çıldır (clima connental, 1585 m). El potencial anoxidante se evaluó mediante diversos enfoques, incluyendo los ensayos de TAC, DPPH, glutaón, SOD y catalasa, mientras que también se determinaron las candades de fenoles totales, flavonoides y antocianinas. Los compuestos fenólicos y flavonoides se idenficaron mediante LC-MS/MS, los perfiles de voláles se determinaron mediante GC-MS y el contenido de azúcares libres se analizó mediante HPLC. Los resultados revelaron que las muestras de Çıldır mostraron una acvidad de depuración de DPPH y niveles de catalasa significavamente mayores, mientras que las muestras de Posof presentaron un mayor contenido de fenoles totales y concentraciones de glutaón. Ambas muestras mostraron una capacidad anoxidante total comparable, aunque a través de diferentes vías bioquímicas. El análisis LC-MS/MS reveló que los ácidos shikímico, clorogénico y p-cumárico eran los fenólicos predominantes en la muestra de Çıldır, mientras que la runa y la hesperidina fueron más abundantes. El perfil GC-MS destacó el ácido acéco y la acetoína como los principales voláles en la muestra de Çıldır, mientras que el etanol y el furfural predominaron en la muestra de Posof. La glucosa y la fructosa se idenficaron como los azúcares primarios, mientras que solo se detectaron pequeñas candades de sacarosa en las frutas de Çıldır. Las diferencias microclimácas influyen fuertemente en los perfiles bioquímicos y funcionales de la ciruela Cancur, resaltando su valor como fuente de anoxidantes, compuestos de sabor e ingredientes alimentarios funcionales. Palabras clave: Ciruela cancur; Prunus domesca L.; anoxidante; perfil fenólico; compuestos voláles

Comparave Phytochemistry and Bioacvity of Local Prunus domesca L. / Duzguner and Kucukgul UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico INTRODUCTION Increased fruit and vegetable consumpon has been linked to a reduced incidence of degenerave diseases due to the high anoxidant potenal of these plant foods [1] Epidemiological studies indicate that consumpon of fruits and vegetables rich in phenolic compounds provides significant benefits in prevenng deaths from cardiovascular and cerebrovascular diseases and some cancers [2]. One of the most effecve ways to increase anoxidant intake is to increase the frequency of consumpon of fruits and vegetables rich in polyphenols [3] Plum (Prunus spp.) is a stone fruit belonging to the Rosaceae family and is disnguished from other Prunus species (peaches, cherries, etc.) by its morphological characteriscs. Despite their low-calorie content, plums have high nutrional value; they contain carbohydrates such as sucrose, glucose, and fructose; organic acids such as citric and malic acids; fiber (pecn); tannins; aromac compounds; and various enzymes [4]. They are also parcularly rich in vitamins A, C, E, B1, and B2, as well as minerals such as potassium, phosphorus, calcium, and magnesium. Because they are high in potassium and low in sodium/potassium, consuming plums appears to be parally beneficial for paents with hypertension. [5]. Plums contain high levels of anthocyanins and other phenolic compounds. These bioacve compounds exhibit higher anoxidant properes compared to oranges, apples, and strawberries. In vitro studies suggest that the high anradical acvity detected in plum extracts is due to the phenolic compounds contained in the fruit. [6]. Besides their health benefits, plums also have potenal uses in sustainable livestock feeding and waste reducon. The rising cost of animal feed and the limited availability of natural resources have made it necessary to explore alternave feed sources. In this regard, wastes and by-products from the fruit and vegetable processing industry offer significant potenal, as they can help reduce environmental waste and support nutrient recycling. Plum fruits and their processing residues, rich in carbohydrates, fiber, minerals, and anoxidant compounds, can serve as an energy source and funconal addive in ruminant diets [6]. Previous studies have shown that adding fruit residues to animal feed at appropriate levels can improve digesbility, lower greenhouse gas emissions, and enhance anoxidant capacity. Therefore, plum-derived materials and other fruit and vegetable by-products may represent valuable feed alternaves for sustainable and environmentally friendly livestock producon. Building on these consideraons, evaluang the biochemical composion and funconal properes of local plum variees may provide valuable insights into their potenal use in both human nutrion and sustainable agricultural pracces [7]. This study was designed to invesgate the potenal of Caucasian plum species nave to northeastern Türkiye. This variety, known locally as ‘cancur,’ is of parcular interest due to its unique biochemical properes and potenal health benefits. It thrives in diverse climates, parcularly in Ardahan’s Çıldır (Öncül village; elevaon: 1793 m) and Posof (Türkgözü; elevaon: 1276 m) [8]. This study aims to comprehensively evaluate the anoxidant capacity, total phenolic and flavonoid content, phenolic and volale components of the Cancur plum (Prunus domesca L.) grown in two different climates in two regions. It was thought that a comprehensive analysis of these species grown in two different climates in the same region would be useful in revealing their potenal as a funconal food. The study’s findings, which are significant for scienfic and industrial applicaons, will enlighten the field and pave the way for the development of innovave funconal foods, offering new possibilies for public health. MATERIALS AND METHODS Collecon of fruit samples Samples of Cancur plum (Prunus domesca L.) fruit were collected from Turkgozu village of Posof/Ardahan (Türkiye) (Altude: 1260 m, 41° 27’ 22’’N, 42° 12’ 56’’E) and from Oncul village of Kurtkale locality of Çıldır/Ardahan (Altude: 1585 m, 41°14’52.9838’’N ‘’ N, 43°8’2.8842’’E ‘’ E). The collected samples were washed and cleaned of dust and soil residues and used in analyses. Posof, one of the districts where plums are harvested, has a microclimate with rainy winters and hot summers. The district of Çıldır, on the other hand, has a connental climate, with harsh, cold winters and warm, rainy summers. Extracon of plums Fresh fruits (CP: for Çıldır plum, PP: for Posof plum) were pied and dried at 40–45°C to constant weight, then ground to obtain a fracon of ≤ 0.5 mm. Aſter weighing approximately 10.0 grams (g) (Meler Toledo, Switzerland), 200–250 mL of HPLC- grade methanol was added to a Soxhlet flask and extracted for six hours (h). The resulng extract was filtered through a 0.22 µm PTFE filter and transferred to amber vials for analysis. It was used in the analysis of anoxidant and bioacvity parameters. Total anoxidant capacity and DPPH analyses A commercial spectrophotometric kit (TAC; Rel Assay Diagnoscs) was used to determine the total anoxidant level. For this purpose, ABTS soluon was incubated with metmyoglobin (peroxidase) and hydrogen peroxide (H₂O₂) to form the ABTS⁺• radical caon, which is blue-green and exhibits maximum absorpon at 660 nm. Measurements were performed spectrophotometrically (Shimadzu Co., Japan) in triplicate, and the results were expressed in Trolox Equivalent (TE). Trolox was used as the reference standard in creang the calibraon curve. Analyses were performed in triplicate, and mean values were used. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging acvity was determined based on the method developed by Blois [9]. Before the analysis, a fresh stock soluon was prepared by dissolving the DPPH radical in methanol at a concentraon of 1 mM. Aſter stock soluon preparaon, 2 mL of DPPH soluon and 0.1 mL of the plum extract were mixed, and the mixture was kept in the dark for 30 min. Aſter the incubaon, the acvity levels were measured using a UV-Vis spectrophotometer (Shimadzu Co., Japan) at 517 nm. Methanol was used as a blank, and 0.1 mL of methanol was added instead of vinegar in the control group. Analyses were performed in triplicate, and mean values were used. 2 of 8

Revista Cienfica, FCV-LUZ / Vol. XXXVI UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico Determinaon of glutathione The glutathione (GSH) determinaon was carried out using its ability to reduce 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB). During this reacon, the thiol groups in GSH interact with DTNB, generang 2-nitro-5-mercaptobenzoic acid, which gives a yellow coloraon. The absorbance of this chromogenic compound was then measured, and GSH levels were expressed as mmol/L [10]. All measurements were conducted in triplicate, and the average values were considered for evaluaon. Determinaon of anoxidant enzyme acvies Superoxide dismutase (SOD) acvity was determined using an ELISA kit that measures the enzyme’s ability to reduce superoxide anions generated in a xanthine/xanthine oxidase (XOD) system. In this assay, SOD acvity was quanfied based on its capacity to inhibit or reduce the formaon of a chromogen, allowing for measurement of SOD levels. Catalase (CAT) acvity was assessed by measuring the amount of H₂O₂ remaining aſter CAT catalysis. The measurements performed by using an ELISA kit and analyses were performed in triplicate, and mean values were used. Total phenolic content and total flavonoid content Total phenolic content was determined using the Folin- Ciocalteu method [11]. 200 μL of extract, 1000 μL of Folin- Ciocalteu, and 800 μL (7.5 %) Na 2 CO 3 were added to a glass tube, and the mixture was incubated for 2 h at room temperature. Aſter that, absorbance was measured at 750 nm against a 50 % ethanol-water mixture in a spectrophotometer (Shimadzu Co., Japan). The total phenolic content of the samples was determined as milligrams per 100 g using a gallic acid standard. Analyses were performed in triplicate, and mean values were used. The total flavonoid content was determined according to the method described by Queer-Deleu et al. [12]. One milliliter of the extract was mixed with 1 milliliter of 2 % AlCl 3 and incubated for 1 h at room temperature in the dark. The absorbance of the samples was determined using a spectrophotometer at 415 nm and calculated in milligrams per 100 g based on the calibraon curve prepared using roune methods. Analyses were performed in triplicate, and mean values were used. Total anthocyanin content The total anthocyanin content was determined using the pH differenal method [13]. The absorbances of the samples incubated in 0.025 M KCl buffer [pH 1.0] and 0.4 M CH 3 COONa buffer were measured at 520 and 770 nm by a spectrophotometer. Analyses were performed in triplicate, and mean values were used. Phenolic profile screening with LC-MS/MS Methanol extracts of plum samples were obtained using a simple extracon method. 10 g of parally dried and ground plant samples were shaken with 50 mL of methanol at 100 rpm for 24 h. The solvent in the resulng mixture was removed using a rotary evaporator (SciLogex RE100-Pro). The extracts were frozen and lyophilized for 48 h and stored at +4 °C (Arçelik, Türkiye) unl analysis. For analysis, 50 mg of the dry extract was sonicated for 5 min in 1 mL of water-methanol (50:50), filtered through a 0.25 µm PETF filter, and diluted 1:10 with the same solvent mixture before being transferred to vials. Samples prepared for secondary metabolite analyses were filtered using a PTFE filter (Isolab) with a pore diameter of 0.45 µm and transferred to capped glass vials before analysis. No diluon was applied to the samples. Qualitave and quantave analyses of secondary metabolites containing phenolic and flavonoid compounds were performed using liquid chromatography (Spark Holland) and tandem mass spectrometry (AB SCIEX 4000 QTRAP). Chromatographic separaon was provided by a C18-type column (Inertsil ODS-3V, 250 mm × 4.6 mm, 5 µm). 0.1 % (v/v) formic acid soluon (phase A) and methanol (phase B) were used as mobile phases. The injecon volume was 10 µL, the flow rate was 0.700 mL/min, and the column oven temperature was 30 °C. The chromatographic analysis me was determined as 20 min. Volale components screening with GC-MS Analysis of volale components was carried out using a Gas chromatography and mass spectrometry (GC–MS; QP 2010 Ultra, Shimadzu, Japan) system equipped with a Headspace (HS) unit. A TRB-5MS capillary column (60 m length, 0.25 mm inner diameter, 0.25 µm film thickness) was used in GC analysis. Helium (He) was selected as the carrier gas, and the pressure was set at 164.9 kPa. The column oven was inially kept at 40 °C, then increased to 320 °C at 12 °C/min and held at this temperature for 2 min. The injecon temperature was 230 °C, the injecon mode was set as “split”, and the split rao was set as 70:1. The total flow rate was 113.8 mL/min, the column flow rate was 1.56 mL/min, and the linear velocity was 31.8 cm/s. Under headspace condions, the valve oven temperature was set at 120 °C, the transfer line temperature at 120 °C, and the sample plate temperature at 120°C. The sample plate equilibration time was set at 3 min, and the sample equilibration time at 2 min. The mixer was turned off, the mix level was set at 5, the mix time was set at 5 min, and the stabilization time was set at 0.5 min. Pressurization was set at 10 PSIG for 2 min, the loop fill pressure was set at 5 PSIG, the fill time was set at 2 min, and the injection time was set at 3 min. Free sugar analysis Fresh plum fruits collected from two different regions (Posof and Çıldır) were dried in an oven at 40 °C (CTO-10AS VP, Shimadzu, Japan) for three d. A 10 g sample of the dried fruits was weighed, cut into small pieces, and then covered with ~50 mL of dislled water. The samples were extracted at room temperature on a magnec srrer for 24 h. The extracts were filtered, and the filtered soluons were transferred to glass petri dishes and dried at 40 °C for another two d. 100 mg of completely dried samples were weighed and dissolved in 1 mL of dislled water, followed by 10 min of sonicaon and 10 min of vortexing (VELP ZX3, Italy). The soluons were centrifuged (Elektromag M4808 PR, Türkiye) at 25155 g for 15 min. The supernatant was diluted with an equal volume of dislled water and subjected to HPLC analysis. Analyses were performed using a Shimadzu LC-20AT HPLC system equipped with a SIL-20A HT autosampler, SPD-M20A 3 of 8

Comparave Phytochemistry and Bioacvity of Local Prunus domesca L. / Duzguner and Kucukgul UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico diode array detector (DAD), CTO-10AS VP column oven, and LC Soluon data analysis soſtware. Separaon was performed using a SilUR NH₂ HPLC column (250 × 4.6 mm, 5 µm), and a mixture of acetonitrile and water (80:20, v/v) was used as the mobile phase. The injecon volume was 20 µL, the flow rate was 1.0 mL/min, the column temperature was 30 °C, and the analysis me was 35 min. Stascal analysis All experiments were conducted in triplicate, and results were expressed as mean ± standard deviaon (SD). Stascal analysis was performed using IBM SPSS Stascs version 25.0. RESULTS AND DISCUSSION Posof (PP) and Çıldır (CP) samples of Cancur plum were compared in terms of anoxidant capacity (TABLE I). Total anoxidant capacity values were found to be 1513 ± 256 µmol trolox equiv./L in PP and 1590 ± 137 µmol trolox equiv./L in the CP sample. While DPPH radical scavenging acvity was 46.98 ± 0.01 % in PP, it was 58.69 ± 0.02 % in the CP sample and was found to be stascally significantly higher (P < 0.001). SOD acvies, one of the enzymac anoxidants, were detected at similar levels in both samples. Glutathione level was 2.20 ± 0.02 in PP, while it was lower in CP (1.680 ± 0.005; P < 0.001). CAT acvity was significantly higher in the CP sample (P < 0.001). TABLE I Anoxidant Levels of Cancur Plum PP CP Total Anoxidant Capacity (µmol trolox equiv./L) 1513 ± 256 1590 ± 137 DPPH (%inhibion) 46.98 ± 0.01 58.69 ± 0.02 * Superoxide Dismutase 5.45 ± 0.04 5.39 ± 0.04 Glutathione 2.20 ± 0.02 1.680 ± 0.005 * Catalase 2.48 ± 0.22 5.19 ± 0.22 * *P<0.001 (PP: Posof Plum vs CP: Çıldır Plum) Plums, especially the Cancur variety, are rich fruits for not only nutrion but also in flavonoids, anthocyanins, carotenes and polyphenolic acids [7]. These compounds affected the high anoxidant capacity of plums, with the CP sample exhibing superior DPPH radical scavenging capacity and CAT acvity. On the other hand, the PP sample comes forward in terms of total phenolics and glutathione levels. This suggests that CP primarily prevents oxidave stress through enzymac mechanisms, while PP does so through phenolic compounds and glutathione. The similar total anoxidant capacity of both plum samples suggests that they exhibit the same protecve acvity through different biochemical pathways and thus provide a range of health benefits to consumers. As seen in TABLE II, when phenolic compounds and other bioacve parameters were evaluated, the total anthocyanin amount was determined as 56.44 ± 2.33 mg/L cyanidin-3- glucoside in the PP sample and 52.21 ± 1.36 mg/L in the CP sample. The total phenolic substance amount was found to be 860.33 ± 14.16 mg/L gallic acid equivalent in the PP sample, which was stascally significantly higher than the CP sample (788.00 ± 2.51 mg/L; P < 0.05). On the other hand, the total flavonoid amount was found to be 3.57 ± 0.017 mg/L in the CP sample, which was significantly higher than the PP sample (P < 0.05). TABLE II Different Bioacvity Levels of Cancur Plum PP CP Total Antocyanine (Siyanidin-3-glikozit) 56.44 ± 2.33 52.21 ± 1.36 Total Phenolic (mg/L gallic acid) 860.33 ± 14.16 788.00 ± 2.51 ** Total Flavonoid (mg/L) 3.47 ± 0.008 3.57 ± 0.017 ** ** p<0.05 (PP: Posof Plum vs CP: Çıldır Plum) Flavonoid content was significantly higher in CP, while anthocyanin levels were similar in the two samples. Rop et al. [5] also suggested that regional plums had higher phenolic, anoxidant, and mineral contents than common variees in a study of 12 plum variees grown in the same region. These differences are believed to be due to genec variaon and adaptaon to microclimac condions. Similarly, Rupasinghe et al. [6] reported a relavely high correlaon between total phenolic and anoxidant acvity in European genotypes. The findings confirm the phenolic content- anoxidant acvity relaonship revealed in the literature and show that Cancur plum variees have funconal food potenal for human health and can be valuable resources for breeding new variees. When the volale compound profiles were examined, it was seen that the dominant compounds in the CP sample were acec acid (48.89 %), 2-butanone-3-hydroxy (acetoin; 19.51 %), and 1,1’-bibicyclo (2.2.2) octyl-4-carboxylic acid (22.54 %). Addionally, ethanol, hexanoic acid, and nonanal were detected at lower levels (TABLE III). In the PP sample, the highest concentraons were ethanol (51.98 %) and 3,3-dimethyl-2-phenyl-2-(1-oxo-1,2,3,4- tetrahydronaphthalen-2-yl) azirane (29.88 %). 4 of 8

Revista Cienfica, FCV-LUZ / Vol. XXXVI UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico Addionally, acec acid was detected at 10.04 %, furfural at 3.86 %, and nonanal at 1.44 %. Other volale compounds (e.g., diacetyl, pentanal, hexanal, benzene, acetaldehyde) were recorded at lower percentages (TABLE III). Volale metabolites are generated both as part of fruit ripening and because of ssue disrupon, with some arising from endogenous metabolic pathways and others triggered by cellular injury. Their composion and abundance are strongly influenced by genec background, environmental condions, culvaon pracces, maturity stage, and postharvest storage [14]. In evaluang the volales responsible for fruit aroma, it becomes clear that disnct chemical classes make crical contribuons to the percepon of flavor and the unique sensory properes of each fruit [15]. The analysis of volale constuents in Cancur plum vinegars from the Çıldır (CP) and Posof (PP) regions demonstrated that geographical origin markedly influences their flavor aributes. In the CP samples, acec acid emerged as the dominant compound, imparng the typical sharp and sour character associated with vinegar. Elevated acetoin levels further contributed buery–creamy notes [14], enhancing mouthfeel smoothness. Conversely, ethanol was the most abundant volale in the PP samples, arising from fermentaon and contribung significantly to flavor balance and aromac complexity [16]. TABLE III Volale Compounds of Cancur Plum Samples Aromac Compounds Ret. Time Area Area % CP 1,1’-bibicyclo(2.2.2)octyl-4-carboxylic acid 3.582 2013638 22.54 Ethanol (CAS) Ethyl alcohol 4.030 386400 4.33 Acec acid 6.750 4366910 48.89 2-Butanone, 3-hydroxy- (CAS) Acetoin 8.641 1742699 19.51 Hexanoic acid (CAS) n-Hexanoic acid 16.819 252052 2.82 Nonanal 20.311 171019 1.91 PP 3,3-Dimethyl-2-phenyl-2-(1-oxo-1,2,3,4-tetrahydronaphthalen-2-YL) Azirane 3.616 19270443 29,88 Ethanol (CAS) Ethyl alcohol 4.083 33520578 51,98 2,3-Butanedione (CAS) Diacetyl 5.162 291829 0,45 Butanal, 3-methyl- 6.295 104663 0,16 Pentanal (CAS) n-Pentanal 7.234 49652 0,08 Hexanal (CAS) n-Hexanal 10.261 150869 0,23 Acec acid 10.581 6475089 10,04 2-Furancarboxaldehyde (CAS) Furfural 11.439 2489100 3,86 2-Butanone, 3-hydroxy- (CAS) Acetoin 11.720 879230 1,36 N Heptanal 13.855 77151 0,12 Octanal 17.954 96153 0,15 Benzeneacetaldehyde 19.754 151188 0,23 Nonanal (CAS) n-Nonanal 22.044 930641 1,44 (PP: Posof Plum vs CP: Çıldır Plum) Addional compounds such as diacetyl, furfural, and several aldehydes enriched the PP profile with fruity and floral nuances [13], leading to a more intricate aromac composion. In contrast, the CP sample displayed a narrower aldehyde spectrum, represented mainly by nonanal (1.91 %), which pointed to a simpler aroma profile. Collecvely, these findings emphasize that climac and environmental factors specific to each culvaon site are decisive in shaping the volale composion, thereby conferring disnct sensory characteriscs to Cancur plums depending on their region of origin. The free sugar contents of the Posof and Çıldır samples of the Cancur plum were determined by HPLC (TABLE IV). The most dominant sugar in both samples was glucose, which was 18.88 mg/mL (37.77 g/100 g; 58.01% area rao) in CP and 19.02 mg/ mL (38.05 g/100 g; 61.20 %) in PP. Fructose levels were found to be 13.04 mg/mL (26.09 g/100 g; 41.17 %) in CP and 11.83 mg/mL (23.67 g/100 g; 38.80 %) in PP. However, sucrose was detected at a low level only in the CP sample and was not detected in the PP sample. Other sugar species (turanose, maltose) were not measured in either sample. Sugar content, type, and rao play important roles in determining fruit flavor, and their regulaon is influenced by various factors, including transcripon factors (TFs), epigenec modificaons, phytohormones, and environmental condions [17]. The results of the study indicate that glucose and fructose are the dominant components in the free sugar composion of Cancur plum. Glucose (37.77 g/100 g) and fructose (26.09 g/100 5 of 8

Comparave Phytochemistry and Bioacvity of Local Prunus domesca L. / Duzguner and Kucukgul UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico g) were prominent in the CP, while similar levels of glucose (38.05 g/100 g) but lower fructose (23.67 g/100 g) were found in the PP. This suggests that samples from both regions have similar glucose profiles in terms of sweetness percepon, but the difference in fructose levels may contribute to the more intense fruity taste in the Çıldır samples. Sucrose was detected at low levels (1.14 g/100 g) only in CP samples and was not found in PP samples. This difference may be related to the breakdown of sucrose into glucose and fructose by the invertase enzyme during ripening. Indeed, it has been previously reported that sucrose decreases, and glucose and fructose raos increase during ripening in various fruits [17],[18]. Therefore, it is likely that sucrose was completely degraded in PP. Sugar composion in fruits varies depending on environmental condions, genotype, and ripening stage. Ecological factors, parcularly temperature and light intensity, have a direct impact on carbohydrate metabolism [18], 19]. In this context, the different microclimac characteriscs of the Çıldır and Posof regions may be one of the main reasons for the differences in fructose levels. TABLE IV Free Sugar Levels of Cancur plum by HPLC Sugar CP Concentraon (mg/mL) CP g/100 g CP Area % PP Concentraon (mg/mL) PP g/100 g PP Area % Fructose 13.0471 26.09 41.17 11.8360 23.67 38.80 Glucose 18.8869 37.77 58.01 19.0266 38.05 61.20 Saccarose 0.5679 1.14 0.82 ND ND ND Turanose ND ND ND d ND ND ND Maltose ND ND ND d ND ND ND CP: Çıldır Plum, PP: Posof Plum, ND: not detected As shown in TABLE V, LC-MS/MS analyses revealed that the phenolic compounds in the PP and CP samples of Cancur plum exhibited both common and disnct characteriscs. The most dominant compound in both samples was shikimic acid, which was found to be higher in the PP (392.98 µg/g) than in the CP (355.26 µg/g). Chlorogenic acid, p-coumaric acid, and vanillin were also other significant compounds detected in high amounts in both samples. The amount of p-coumaric acid was higher in the PP than in the CP. It is noteworthy that some phenolic compounds were found only in certain samples. For example, protocatechuic acid was measured only in the CP at 10.94 µg/g, while in PP, it was below the LOQ. Similarly, hesperen was detected only in the CP. Caffeic acid was detected in both samples but was found at a higher level in the PP (12.43 µg/g) than in the CP (6.26 µg/g). TABLE V Phenolic and Flavonoid Compounds of Cancur Plum Analytes Retenon Time R 2 LOQ e (ng/mL) LOD e (ng/mL) Quanficaon (µg/G extract) CP PP Shikimic acid 1.367 0.9985 18.59 7.17 355,26 392,98 Gallic acid 3.808 0.9986 13.17 3.16 N.D. d N.D. d Protocatechuic acid 5.554 0.9993 18.59 7.17 10,94 <LOQ Catechin 6.888 0.9992 7.50 1.71 N.D. d N.D. d Chlorogenic acid 7.396 0.9986 25.90 11.59 225,97 132,57 Hydroxybenzaldehyde 7.767 0.9992 12.87 4.97 N.D. d N.D. d Vanillic acid 7.826 0.9981 724.21 89.04 N.D. d N.D. d Caffeic acid 7.838 0.9985 24.16 6.92 6,26 12,43 Syringic acid 8.396 0.9958 857.34 358.50 N.D. d N.D. d Caffein 8.399 0.9995 15.50 6.81 N.D. d N.D. d Vanillin 8.428 0.9989 40.54 14.59 34.55 38,20 p-coumaric acid 9.410 0.9984 17.54 3.53 156,18 172,16 Salicylic Acid 9.746 0.9983 82.96 47.67 10,33 39,09 Taxifolin 9.768 0.9999 23.51 11.03 N.D. d N.D. d Polydan 9.772 0.9987 1.84 1.15 N.D. d N.D. d 6 of 8

Revista Cienfica, FCV-LUZ / Vol. XXXVI UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico Resveratrol 9.775 0.9989 13.56 4.58 N.D. d N.D. d Trans-ferulic acid 10.096 0.9995 11.53 6.12 N.D. d N.D. d Sinapic acid 10.244 0.9991 4.97 1.94 N.D. d N.D. d Scutellarin 11.054 0.9989 4.00 3.13 N.D. d N.D. d o-coumaric acid 11.506 0.9995 8.00 4.02 N.D. d N.D. d Coumarin 11.510 0.9986 20.70 6.01 N.D. d N.D. d Protocatechuic ethyl ester 11.573 0.9987 24.92 14.56 N.D. d N.D. d Run 11.649 0.9995 240.67 59.56 38,35 25,42 Isoquercitrin 11.650 0.9989 11.27 9.94 21,07 10,48 Hesperidin 12.271 0.9979 17.68 4.14 23,96 <LOQ Quercen-3-xyloside 12.402 0.9992 69.41 18.71 0,45 N.D. d Kaempferol-3-glucoside 13.072 0.9996 4.52 1.16 0,53 N.D. d Fisen 13.344 0.9985 44.37 10.90 1,52 N.D. d Baicalin 13.544 0.9996 3.10 0.53 0,50 N.D. d Trans-cinnamic acid 14.325 0.9980 22.03 11.19 N.D. d N.D. d Quercen 14.841 0.9986 16.91 4.66 5,57 N.D. d Naringenin 14.994 0.9957 0.46 1.37 N.D. d N.D. d Morin 15.681 0.9984 0.53 0.13 0,03 N.D. d Hesperen 15.684 0.9981 0.65 0.30 2,34 N.D. d Kaempferol 16.377 0.9984 5.40 1.87 N.D. d N.D. d Baicalein 17.150 0.9991 0.96 0.60 N.D. d N.D. d Biochanin A 17.926 0.9993 0.73 0.15 0,04 N.D. d Luteolin 17.967 0.9988 21.45 12.05 N.D. d N.D. d Chrysin 18.002 0.9997 0.13 0.07 N.D. d N.D. d CP: Çıldır Plum, PP: Posof Plum, d : not detected When flavonoid derivaves were examined, it was observed that the amount of run was higher in the CP (38.35 µg/g) than in the PP (25.42 µg/g). Isoquercitrin was also found to be significantly higher in the CP than in the PP. Hesperidin was detected at 23.96 µg/g in the CP, while in the PP, it was below the LOQ. In contrast, some flavonoid compounds, such as quercen, fisen, and baicalin, were detected in low amounts only in the PP. The phenolic composion exhibited clear regional differences between the samples. Shikimic acid represented the most abundant constuent in both vinegars, with notably higher levels in the PP. Chlorogenic acid, p-coumaric acid, and vanillin were also present at substanal concentraons in both regions. Among these, p-coumaric acid accumulated more strongly in Posof, whereas run and hesperidin were more enriched in Çıldır. Caffeic acid was idenfied in both samples but predominated in Posof, while protocatechuic acid and hesperen appeared exclusively in Çıldır. Regarding flavonoid derivaves, run, isoquercitrin, and hesperidin were detected in both samples, although their relave proporons varied considerably. In agreement with these observaons, Çelik et al. [20] reported that chlorogenic and caffeic acids were the most prominent phenolics in three different plum variees, alongside run, gallic acid, and vanillic acid. They aributed such composional differences to environmental and genec influences. The authors also highlighted that flavonol glycosides tend to accumulate in the peel due to light exposure and that climac condions exert a major role in phenolic biosynthesis. Overall, the present findings indicate that ecological factors shape the phenolic profile of Cancur plums, thereby influencing their biofunconal potenal depending on the region of culvaon. In addion to these composional and ecological aspects, the potenal ulizaon of plum-derived materials in livestock systems represents another dimension of their biofunconal value. Plum processing residues, such as pulp and kernels, contain valuable nutrients including protein, fat, and fiber, which may allow their use as feed ingredients in ruminant diets. When properly processed to eliminate cyanogenic compounds, these by-products can contribute to nutrient recycling and sustainable livestock producon. Studies have also indicated that limited inclusion of fruit and vegetable residues in sheep diets can improve digesbility and anoxidant status while reducing methane emissions. However, further studies are needed to evaluate the specific effects of plum-based materials on animal performance and safety [7 ,[21]. 7 of 8

Comparave Phytochemistry and Bioacvity of Local Prunus domesca L. / Duzguner and Kucukgul UNIVERSIDAD DEL ZULIA Serbiluz Sistema de Servicios Bibliotecarios y de Información Biblioteca Digital Repositorio Académico CONCLUSION Marked variaons were idenfied in the anoxidant capacity, phenolic composion, volale profile, and sugar content of Cancur plum vinegars derived from the Çıldır and Posof regions. The Çıldır samples appeared to migate oxidave stress predominantly through enzymac pathways, whereas the Posof samples relied more heavily on phenolic constuents and glutathione. Disncons in aroma-acve volales and carbohydrate composion further highlighted the influence of regional microclimates on the biofunconal and sensory characteriscs of the product. Collecvely, these results underscore the relevance of Cancur plum as a promising funconal food and highlight its potenal ulity as a raw material for innovave applicaons in the food industry. Conflict of interests The authors of this study declare that there is no conflict of interest with the publicaon of this manuscript. BIBLIOGRAPHICS REFERENCES [1] Oz AT, Kaas E. Phytochemicals in Fruits and Vegetables. In: Waisundara V, Shiomi N, (editors). Superfood Funct. Food Process. Ul. InTech. [Internet]. 2017; 8:175-184. doi: hps://doi.org/qg56 [2] Hashim MS, Lincy S, Remya V, Teena M, Anila L. Effect of polyphenolic compounds from Coriandrum savum on H 2 O 2 -induced oxidave stress in human lymphocytes. Food Chem. [Internet]. 2005; 92(4):653-660. doi: hps:// doi.org/bpk94r [3] Lima GPP, Vianello F, Corrêa CR, da Silva Campos RA, Borguini MG. Polyphenols in Fruits and Vegetables and Its Effect on Human Health. Food Nutr. Sci. [Internet]. 2014; 5(11):1065-1082. doi:hps://doi.org/qg57 [4] Kristl J, Slekovec M, Tojnko S, Unuk T. Extractable anoxidants and non-extractable phenolics in the total anoxidant acvity of selected plum culvars (Prunus domesca L.): Evoluon during on-tree ripening. Food Chem. [Internet]. 2011; 125(1):29-34. doi: hps://doi. org/dcs68v [5] Rop O, Jurikova T, Mlcek J, Kramarova D, Sengee Z. Anoxidant acvity and selected nutrional values of plums (Prunus domesca L.) typical of the White Carpathian Mountains. Sci. Horc. [Internet]. 2009; 122(4):545-549. doi: hps://doi.org/ct4gcb [6] Rupasinghe HPV, Jayasankar S, Lay W. Variaon in total phenolics and anoxidant capacity among European plum genotypes. Sci. Horc. [Internet]. 2006; 108(3):243- 246. doi: hps://doi.org/fxhfnd [7] Gonzáles-García E, Marina ML, Concepción García M. Plum (Prunus domesca L.) by-Product as a New and Cheap Source of Bioacve Pepdes: Extracon Method and Peptes Characterizaon. J. Funct. Foods. [Internet]. 2014; 11:428-437. doi: hps://doi.org/f6t5vc [8] Murathan ZT, Arslan M, Erbil N. Analyzing Biological Properes of Some Plum Genotypes Grown in Turkey. Int. J. Fruit Sci. [Internet]. 2020; 20(3):S1729-S1740. doi:hps://doi.org/qg58 [9] Blois MS. Anoxidant determinaon by the use of a stable free radical. Nature. [Internet]. 1958; 181:1199- 1200. doi: hps://doi.org/bdq98t [10] Sedlak J, Lindsay RH. Esmaon of total, protein-bound, and nonprotein sulydryl groups in ssue with Ellman’s reagent. Anal. Biochem. [Internet]. 1968; 25(1):192-205. doi:hps://doi.org/csbsfm [11] Spanos GA, Wrolstad RE. Phenolics of Apple, Pear, and White Grape Juices and Their Changes with Processing and Storage - a Review. J. Agric Food Chem. [Internet]. 1992; 40(9):1478-1487. doi: hps://doi.org/b27dmg [12] Queer-Deleu C, Gressier B, Vasseur J, Dine T, Brunet C, Luyckx M, Cazin M, Cazin JC, Bailleul F, Tron F. Phenolic compounds and anoxidant acvies of buckwheat (Fagopyrum esculentum Moench) hulls and flour. J. Ethnopharmacol. [Internet]. 2000; 72(1-2):35-42. doi: hps://doi.org/ſt249d [13] Garzón GA, Wrolstad RE. The stability of pelargonidin- based anthocyanins at varying water acvity. Food Chem. [Internet]. 2001; 75(2):185-196. doi: hps://doi. org/bdgkq5 [14] Gonçalves B, Oliveira I, Bacelar E, Morais MC, Aires A, Cosme F, Ventura-Cardoso J, Anjos R, Pinto T. Aromas and Flavours of Fruits. In: Vilela A, editor. Generaon of Aromas and Flavours. Rijeka: IntechOpen. 2018; 2:9-31. hps://doi.org/gpx6qr [15] Sánchez-Rodríguez L, Ali NS, Cano-Lamadrid M, Noguera- Araga L, Lipan L, Carbonell-Barrachina ÁA, Sendra E. - Flavors and Aromas. In: Yahia EM, editor. Postharvest Physiology and Biochemistry of Fruits and Vegetables. Chapter 18. Cambridge, Inglaterra: Woodhead Publishing; 2019. p. 385-404. doi: hps://doi.org/gjvq89 [16] Aragüez I, Valpuesta-Fernández V. Metabolic engineering of aroma components in fruits. Biotechnol. J. [Internet]. 2013; 8(10):1144-1158. doi:hps://doi.org/f2dv49 [17] Du M, Zhu Y, Nan H, Zhou Y, Pan X. Regulaon of sugar metabolism in fruits. Sci. Horc. [Internet]. 2024; 326:112712. doi: hps://doi.org/g8rk6p [18] Pashaei M, Hassanpour H. Phenolic, amino acids, and fay acids profiles and the nutrional properes in the fresh and dried fruits of black rosehip (Rosa pimpinellifolia L.). Sci. Rep. [Internet]. 2024; 14(1):19665. doi: hps://doi. org/qg6b [19] Kaas E, Kosar M, Türemis N, Baser KHC. Analysis of sugars, organic acids and vitamin C contents of blackberry genotypes from Turkey. Food Chem. [Internet]. 2006; 97(4):732-736. doi: hps://doi.org/c8jjfp [20] Celik F, Gundogdu M, Alp S, Muradoglu F, Ercisli S, Gecer MK, Canan I. Determinaon of Phenolic Compounds, Anoxidant Capacity and Organic Acids Contents of Prunus domesca L., Prunus cerasifera Ehrh. and Prunus spinosa L. Fruits by HPLC. Acta Chromatogr. [Internet]. 2017; 29(4):507-510. doi: hps://doi.org/qg6c [21] Sahoo A, Sarkar S, Lal B, Kumawat P, Sharma S, De K. Ulizaon of Fruit and Vegetables Waste as an Alternave Feed Resource for Sustainable and Eco-Friendly Sheep Farming. Waste Manag. [Internet]. 2021; 128:232-242. doi: hps://doi.org/gj3gs 8 of 8