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Article

Content of Phytomelatonin in Acorns (Quercus sp.) in Its Possible Use as a Phytogenic in Animal Nutrition

by
Soundouss Kaabi
1,2,
Brahim El Bouzdoudi
2,
Mohammed L’bachir El Kbiach
2,
Antonio Cano
1,
Josefa Hernández-Ruiz
1 and
Marino B. Arnao
1,*
1
Phytohormones Laboratory, Department of Plant Biology, Plant Physiology Area, Faculty of Biology, University of Murcia, 30100 Murcia, Spain
2
Plant Biotechnology Team, Faculty of Sciences, Abdelmalek Essaâdi University, Tetouan 93002, Morocco
*
Author to whom correspondence should be addressed.
Processes 2025, 13(7), 2202; https://doi.org/10.3390/pr13072202
Submission received: 30 May 2025 / Revised: 30 June 2025 / Accepted: 7 July 2025 / Published: 9 July 2025
(This article belongs to the Special Issue Advanced Natural Product Technology: Extraction, Separation, and Analysis)
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Versions Notes

Abstract

Phytogenics are functional compounds with a growing interest in animal nutrition. These plant-derived compounds are often used to improve health and behavioral aspects in livestock, and used as antipathogenic agents. Melatonin, an indolic hormonal compound, has been studied as an interesting phytogenic in animal nutrition. This study analyzes the possibilities of acorn-fed flour as a phytomelatonin contributor and its beneficial roles for health. The fruits of two varieties of acorns (Quercus suber var. Maamora and var. Bouhachem), recollected in two different regions of Morocco, have been studied according to their eco-physiological origin. The content in phytomelatonin was analyzed using a solid extractive method and determined by liquid chromatography with fluorescence detection. The results show great morphological differences between the two varieties, and also significant differences in their phytomelatonin content. It is concluded that acorn-fed flour can be an interesting raw material as a phytomelatonin contributor for the functionality of certain feeds and animals. More specific studies using phytomelatonin-rich plants as feed have been proposed to implement specific functionalities in livestock.

1. Introduction

Phytogenics, also known as plant-based feed additives, have gained significant attention in animal nutrition due to their natural origin and potential health benefits. These bioactive compounds are derived from herbs, spices, seeds, and other plants, and are used to improve animal performance, health, and product quality. One of the main advantages of phytogenics is their ability to enhance feed intake and digestibility [1,2]. Many plant extracts contain essential oils, flavonoids, and phenolic compounds that stimulate appetite and improve gut health. This can lead to better growth rates and feed conversion efficiency in livestock [3]. Phytogenics also possess antimicrobial properties, which can help reduce the reliance on antibiotics in animal production [2,4]. By modulating the gut microbiota, these plant compounds can suppress pathogenic bacteria and promote beneficial microorganisms, leading to improved immune responses [5,6,7]. Furthermore, phytogenics have antioxidant effects, helping animals combat oxidative stress caused by environmental factors or intensive production systems. This can result in improved overall health, reduced disease incidence, and better reproductive performance [1,8,9,10,11,12].
In addition to health benefits, phytogenics can influence the quality of animal products. For example, meat, milk, and eggs from animals fed with plant-based additives often show enhanced flavor, shelf life, and nutritional profile, including increased levels of beneficial fatty acids and antioxidants. The use of phytogenics is also aligned with consumer preferences for natural and organic products. As a result, many producers are incorporating these plant extracts into their feeding strategies to meet market demands and improve sustainability [13,14,15,16,17,18,19].
However, the effectiveness of phytogenics depends on factors such as plant source, extraction method, dosage, and animal species. Proper formulation and standardization is essential to ensure consistent results. Overall, phytogenics represent a promising alternative to synthetic additives and antibiotics in animal nutrition. Their multifaceted benefits contribute to healthier animals, improved product quality, and more sustainable production systems [1,5,7,15,20,21,22,23].
Phytomelatonin is a naturally occurring plant compound that is structurally similar to the hormone melatonin found in animals and humans [24,25]. It is synthesized by various plant species, including fruits, vegetables, grains, and seeds. As a plant-derived molecule, phytomelatonin has garnered increasing interest for its potential health benefits and its application as a nutraceutical in food and as a phytogenic in feed [26,27,28]. Melatonin in animals and humans is primarily known for regulating circadian rhythms and sleep–wake cycles and for possessing antioxidant properties [29,30]. Similarly, phytomelatonin can influence these biological processes when consumed through diet, acting as a natural supplement to support health [31,32,33]. One of the key roles of phytomelatonin as a nutraceutical is its potent antioxidant activity. It scavenges free radicals and reduces oxidative stress, which is linked to aging and numerous chronic diseases such as cardiovascular diseases, neurodegenerative disorders, and cancer. By neutralizing reactive oxygen species, phytomelatonin helps protect cells and tissues from damage [34,35,36,37,38,39,40].
In plants, phytomelatonin plays multiple roles, such as influencing seed germination, root and shoot development, and photosynthesis improvement; enhancing root elongation; improving flowering time and responses to environmental stresses like drought, salinity, and extreme temperatures, mainly by oxidative damage; and also improving defense against pathogen attacks, thereby increasing crop yields. It acts as an antioxidant, protecting plant cells from oxidative stress by scavenging free radicals and as a phytohormone regulator. This protective function enhances plant resilience and productivity under adverse conditions [41,42,43,44,45]. The discovery of phytomelatonin presence in edible plants has opened new avenues for its application in animal nutrition. As a phytogenic additive, meaning derived from plants, phytomelatonin offers several potential benefits when incorporated into animal diets. Its natural origin aligns with the growing demand for sustainable and organic feeding practices [1,46,47].
One of the primary roles of phytomelatonin in animal nutrition is its antioxidant capacity. Oxidative stress is a common challenge in intensive animal farming, leading to cellular damage, reduced immune function, and decreased productivity. Supplementing animal diets with phytomelatonin can help mitigate oxidative stress by neutralizing reactive oxygen species (ROS), thereby improving overall health and performance [1,3]. Moreover, melatonin/phytomelatonin exhibits anti-inflammatory properties that can support immune health in livestock [48]. By modulating inflammatory responses, it may reduce the incidence of disease and improve recovery rates. This attribute makes it particularly valuable during stressful periods such as transportation, weaning, or environmental changes [2,9,20,49,50].
Research studies have demonstrated that dietary supplementation with melatonin/phytomelatonin can enhance reproductive performance in animals [51,52,53,54,55]. For example, improved sperm quality in males and better conception rates have been observed in some species following phytomelatonin administration. These effects are attributed to its influence on hormonal regulation and cellular protection mechanisms [54,56]. Additionally, phytomelatonin has been linked to improved gut health by maintaining intestinal integrity and reducing inflammation within the gastrointestinal tract. A healthy gut is essential for optimal nutrient absorption and overall growth performance [2,57,58,59,60,61]. In terms of growth promotion, some studies suggest that phytomelatonin may positively influence feed efficiency and weight gain by reducing metabolic stress and supporting anabolic processes. Its role as a natural bioactive compound makes it an attractive alternative to synthetic growth promoters [62,63,64]. For example, the use of melatonin implants in slow-growing chickens improves the color, water activity, and lipid anti-oxidation of meat, which could improve preservation and extend shelf life [65,66]. Also, melatonin treatment improves intestinal development in sucking piglets, affecting neural development, barrier integrity, nutrient absorption, and microbiota [60].
Furthermore, the use of phytomelatonin aligns with consumer preferences for natural additives over synthetic chemicals or antibiotics. Its inclusion in feed formulations can contribute to more sustainable livestock production systems with minimal environmental impact. Despite these promising benefits, the research on optimal dosages, long-term effects, and species-specific responses continues. It is essential to establish standardized guidelines for incorporating phytomelatonin into different animal diets to maximize efficacy while ensuring safety [67].
By all, phytomelatonin represents a promising plant-derived bioactive compound with multifaceted roles in enhancing animal health and productivity. As a phytogenic additive, it offers antioxidant, anti-inflammatory, reproductive, and growth-promoting benefits that align with modern sustainable agriculture practices. Ongoing research will further elucidate its potential applications across various livestock species and production systems.
Acorn-fed flour is used in feed, especially, for example, in animal feed for pigs. Feeding pigs with acorn has several reasons related to its nutritional benefits, its tradition, and its impact on meat quality. Acorns are rich in complex carbohydrates, healthy fats, proteins, vitamins, and essential minerals. Acorn meal takes advantage of these nutrients to supplement the pigs’ diet. The intake of acorns during fattening contributes to a more intense flavor, a softer fat infiltration, and a favorable lipid profile, rich in monounsaturated fatty acids such as oleic. This results in high-quality hams and sausages, highly appreciated in gourmet markets. Acorn meal provides an important energy source for pigs, especially in extensive systems where feedstuffs may be scarce or expensive. Using acorn-flour in feed can be an efficient way to take advantage of locally available natural resources, reducing costs and promoting sustainable practices. Also, the presence of natural antioxidants in acorns can help reduce oxidative stress in animals, improving their welfare and potentially their productive performance. Feeding with acorns positively influences the organoleptic characteristics of the pig meat, such as taste, aroma, and texture, which increases its commercial value. In many regions, especially in Spain, feeding with acorns is an ancestral practice that is part of the traditional Iberian pig breeding system. This method has been maintained for its cultural value and for the quality of the final product. In summary, feeding pigs acorn-fed meal combines cultural, nutritional, and economic aspects, as well as contributing to high-quality meat products that are highly valued by both producers and consumers [68,69,70,71,72,73].
However, the exact percentage of acorn-fed meal used in feed can vary depending on the type of feed and the production target. Generally, in the feeding of Iberian pigs in “Montanera” (pastureland practice), acorn-fed flour can represent approximately between 10% and 30% of the composition of the feed. In some cases, especially in diets designed to maximize the quality of Iberian ham, it can be used in percentages close to 20–25%. It is important to note that acorn-fed flour is highly valued for its flavors and nutritional properties, but its use in feed is regulated and adapted to ensure a balanced diet [70,74,75].
In the case of poultry, such as chickens or turkeys, acorn-fed flour is not so common in their usual diet, since their diet is usually based on cereals, vegetable proteins, and other specific ingredients for their growth and health. However, in some special feeding programs or in poultry diets of organic or high-quality production, acorn-fed meal may be included in small quantities. In general, the percentage of acorn meal in poultry feed is quite low, typically less than 5%, and in many cases even lower, so as not to disturb the nutritional balance and avoid digestive problems. The idea is to take advantage of its nutritional and aromatic properties without overloading the diet [69,76,77,78]. An important aspect to consider when feeding animals with acorn-flour is its tannin content, which can be toxic to some species. Generally, the tannin content is reduced with specific treatments of the flours such as slurring, boiling, drying, and fermenting [79,80].
The primary aim of this study was to determine the content of phytomelatonin in acorn samples, in the pericarp and seed, from two different ecotypes of the same variety. Our secondary aim will be to assess the feasibility of incorporating these plant matrices into animal diets, considering physiological requirements and the potential for synergistic interaction with other bioactive elements naturally present in acorns. We suggest that the fruits of Quercus suber represent a significant natural source of phytomelatonin and could be valorized as innovative phytogenic ingredients in animal nutrition. We also hypothesize that their phytomelatonin profile varies according to ecotype and environmental conditions, which could have an impact on their functional potential.

2. Materials and Methods

2.1. Chemicals

All the reagents and solvents (ethyl acetate, acetonitrile, and methanol, HPLC gradient grade) used were provided by Sigma-Aldrich Co. (Madrid, Spain). Melatonin (99%) was provided by Acros Organics Co. (Geel, Belgium). Ultra-pure water from Milli-Q system (Milli-Q Corp, Merck, Darmstadt, Germany) was utilized.

2.2. Plant Material

Over two consecutive years (2022/2023 and 2023/2024), fruits of the cork oak (Quercus suber L.) were harvested randomly at full maturity and in the same season (December and January) in two distinct regions of Morocco (plain and mountain), each characterized by specific ecotypes (Table 1).
The collection was made for two consecutive years (2022/2023 and 2023/2024) and in several localizations (Table 1). A total of 90 mature Quercus suber L. acorns were collected in each study region (Maamora and Bouhachem) over two consecutive years (2022/23 and 2023/2024). To ensure biological and environmental representativeness, the acorns were randomly sampled in 3 separate collection campaigns of 30 acorns each, all within the same season (December–January). Botanical identification of the acorns was carried out with the assistance of a forestry expert, Mr. M. Kaabi, affiliated with the High Commission for Water, Forests, and the Fight Against Desertification, Regional Directorate of Chefchaouen (Morocco).
Afterwards, the acorns were transported to the laboratory. Each acorn was subjected to a rigorous visual inspection. Material showing visible signs of deterioration or infestation was discarded. After sorting, the acorn organs (pericarp and seed) were separated and subjected to oven-drying protocol (50 °C) until the dry weight was stabilized, thus preserving their natural composition as much as possible. Once dried, the seeds were ground using a Microtron-MB550 at 8000 rpm (Kinematica AG, NY, USA), then sieved through a stainless steel sieve to obtain particles with a diameter of less than 0.2 mm. The powder obtained was kept in the dark at room temperature to prevent any alteration in its physicochemical properties.

2.3. Climatological Conditions

Climatological data recorded over the two consecutive years highlighted clear environmental contrasts between the two studied regions (Maamora (plain) and Bouhachem (mountain)). These differences are essential for understanding the eco-physiological responses of Quercus suber acorns to regional climatic conditions. In terms of thermal regime, Maamora consistently exhibited higher mean temperatures (Tmean ranging from 17.93 °C to 18.94 °C) compared with Bouhachem (Tmean from 14.09 °C to 14.92 °C). Notably, Tmin values in Bouhachem were considerably lower (down to 0.4 °C in 2021 and 0.5 °C in 2023), reflecting the mountainous and cooler profile of this region. In contrast, Maamora displayed milder winter temperatures, with Tmin never dropping below 6.5 °C. Regarding precipitation (P), Bouhachem received substantially more rainfall, particularly in 2021 (Pmax = 354.4 mm; Pmean = 82.13 mm), emphasizing its humid and orographic influence. By comparison, Maamora experienced lower and more variable precipitation, with Pmean values decreasing drastically in 2023 (18.96 mm), confirming a trend of increasing aridity over time. Minimum precipitation values were close to zero at both sites across all years, confirming the existence of dry spells typical of the Mediterranean climate.

2.4. Phytomelatonin Extraction

The samples for the phytomelatonin analysis were prepared according to our methodology [81,82]. In brief, 4 mL of ethyl acetate was added to 0.2 g of dry material from the different acorn samples in PVPP tubes. Each sample was repeated three times. After overnight (15 h) in a rotating agitator at 20 °C in darkness, the samples were centrifuged for 10 min at 7500 rpm using a Sorvall RC 5B Plus centrifuge (DuPont, Wilmington, DE, USA) and then decanted into a new tube. Next, the solvent was evaporated to dryness under vacuum using a SpeedVac (ThermoSavant SPD111V, Thermo-Fisher Sci, Waltham, MA, USA). The final residue was resuspended in 1 mL of acetonitrile using an agitator vortex to ensure homogeneity before being filtered (PTFE filter 0.22 μm). Finally, the phytomelatonin content of the samples was analyzed by LC-FLUO.

2.5. Phytomelatonin Analysis by Liquid Chromatography with Fluorimetric Detection (LC-FLUO)

Phytomelatonin was quantified using a high-performance liquid chromatograph (HPLC). An online degasser, quaternary pump, autosampler, thermo-stated column, and a Phenomenex-Luna ODS2 S5 (150 × 4.6 mm) column were all included in the Jasco model 2000 HPLC (Jasco Co., Tokyo, Japan). The instrument was connected to a Jasco FP-2020-Plus fluorescence detector (λexcitation = 280 nm, λemission = 350 nm). The isocratic mobile phase consisted of acidified water (85%) and methanol (15%), at a flow rate of 0.5 mL/min and a column temperature of 36 °C.
The retention times compared with standard melatonin (tR = 11.2 min) and an in-line fluorescence analysis of the molecule’s excitation and emission spectra using the Jasco ChromNav 2.0 Spectra Manager software were applied to identify melatonin in our samples. Additionally, an Agilent LC-chromatograph (Agilent Technologies, Santa Clara, CA, USA) connected to a 6550 Q-TOF mass spectrometer (LC/QTOF-MS) (Agilent Technologies, Santa Clara, CA, USA) to the exact melatonin/phytomelatonin identification in the samples was applied [81,82].

2.6. Statistical Analysis

Statistical analyses were performed using R programming language with the support of RStudio software (v.2024.12.1+563; Posit Software, PBC, Boston, MA, USA). All data were processed and analyzed within this environment. The statistical significance was considered at p < 0.05.

3. Results and Discussion

3.1. Acorn Morphological Characteristics

Oak fruits, known as acorns, are single-seeded endosperm-less nuts containing an etiolated embryo. Acorns of different Quercus species display a morphological diversity influenced by phylogenetic and ecological factors. There is considerable variability not only between species but also between ecotypes. Numerous studies have been carried out to establish correlations between morphological characteristics (shape and size) and ecological factors such as edaphology, climatic variables, water availability, and vegetation type. In addition, acorn size is positively correlated with the length of the development period and with rainfall [73].
The morphometric analysis revealed significant differences between the two ecotypes (Figure 1) (Table 2). No significant morphological differences (p > 0.05) were observed between the two consecutive years. However, substantial ecotypic variation was recorded. The sweet acorn ecotype (QM) showed nearly twice the fresh and dry weight of the bitter acorn (QB), in both 2022/2023 and 2023/2024. For instance, in 2022/2023, the fresh weight of QM reached 11.55 g compared with 6.15 g for QB, while the dry weight was 7.63 g versus 2.46 g, respectively. Similar trends were maintained in 2023/2024 (QM: 11.04 g fresh; 6.96 g dry vs. QB: 4.50 g fresh; 3.45 g dry). Additionally, QM ecotype acorns were consistently larger than QB ones in length, width, and thickness. For example, in 2022/2023, QM acorns measured 46.36 mm in length and 19.87 mm in width, while QB acorns were significantly smaller (30.97 mm length; 16.12 mm width).

3.2. Distribution of Acorn Masses: Pericarp and Seed

The analysis of acorn mass partitioning between pericarp and seed across two harvest seasons (2022/2023 and 2023/2024) revealed significant differences between the two Quercus suber ecotypes: sweet acorns (QM) and bitter acorns (QB) (Table 3). During both seasons, seeds represented the major fraction of total acorn mass, ranging from 78.14% to 84.18%, while the pericarp accounted for 15.82% to 21.85%. QM acorns exhibited a significantly higher proportion than QB acorns (p < 0.05). Specifically, in 2023/2024, the QM ecotype showed the highest pericarp ratio (21.85%), while the lowest value was recorded in QB acorns from 2022/2023 (15.82%). Inversely, QB acorns had a significantly higher seed proportion, especially in 2022/2023 (84.18%).
Fresh and dry weight measures supported these findings. In both years, fresh pericarp weight (FPW) and dry pericarp weight (DPW) were significantly greater in the QM ecotype compared with the QB one. For example, FPW reached 2.37 g in QM vs. 0.76 g in QB in 2023/2024. Similarly, fresh seed weight (FSW) and dry seed weight (DSW) were markedly higher in QM acorns, confirming the overall larger size and mass of QM acorns.

3.3. Phytomelatonin Content in Acorns

LC-FLUO analysis showed a peak corresponding to melatonin/phytomelatonin detected at 11.2 min (Figure 2). A standard melatonin calibration curve was also presented (Figure 3). Under the same conditions. The presence of phytomelatonin in the samples of acorns was detected at 11.2 min (Figure 4) and checked from the respective excitation and emission spectra [21].
The content of phytomelatonin of the different acorns studied is shown in Table 4. In the 2022/2023 season, the highest phytomelatonin content was observed in the seed of the sweet acorn (9.94 ng/g DW), followed by the pericarp of the same ecotype in 2023/2024 (6.67 ng/g DW). Successively, lower concentrations were recorded in the seed of the QB acorns, with 4.88 ng/g DW in 2022/2023 and 4.37 ng/g DW in 2023/2024. The pericarp of the QB acorns showed an inverse trend, with phytomelatonin contents of 3.09 ng/g DW in 2023/2024 and 1.86 ng/g DW in 2022/2023. Notably, the lowest level detected was in the pericarp of the QM acorns (1.01 ng/g DW) in 2022/2023, which was comparable to the QB acorn pericarp (1.86 ng/g DW) from the same year. Considering global organs, the total phytomelatonin contents by each ecotype pointed to a higher acorn phytomelatonin content in sweet seeds (QM) than in bitter seeds (QB) in the two-year period studied (Table 4).

3.4. Relationship Between Phytomelatonin Contents and Morpho-Physiological Parameters

Pearson correlation coefficients were calculated for 13 variables (Figure 5). This analysis highlights the statistical relationships between morphological and weight parameters, as well as phytomelatonin content in the acorn. This matrix was constructed from data collected over two consecutive years, comparing two ecologically contrasting ecotypes (QM and QB).
The results reveal a strong positive correlation between the morphological parameters of the acorns (L, W, T) and their fresh and dry weights (FW, DW), suggesting morpho-weight coherence between these variables and indicating that biomass is proportional to morphometric dimensional acorn structures. The very high correlations between fresh (FSW and FPW) and dry (DSW and DPW) weights indicate proportional growth between the different acorn fractions.
With regard to phytomelatonin, pericarp content (MP) showed a marked negative correlation with the majority of morphological and weight variables, suggesting that phytomelatonin content in the pericarp may be inversely related to tissue growth.
Conversely, phytomelatonin in seed (MS) showed a slightly positive correlation with several parameters, but without reaching high levels of significance, which may indicate a physiological association between phytomelatonin accumulation and seed biomass. This could reflect a distinct function of phytomelatonin according to fruit compartment, with the seed playing the role of metabolic reserve while the pericarp may be more exposed to environmental variations.
Considering the absolute values of phytomelatonin content, acorn-seeds have relevant values between 7.46 and 10.95 ng/g (Table 4). Compared with other seeds, we find average values of 39 ng/g in almonds, 5.8 ng/g in common beans, 3.5 ng/g in walnuts, 2.3 ng/g in peanuts, and 3.8 ng/g in lupin seeds; all of these values are higher than that of common cereal flours, which usually contain between 0.5 and 2.0 ng/g of phytomelatonin [14,26,37].

4. Conclusions

This study highlights marked morphological, weight, and phytomelatonin content differences between two Quercus suber ecotypes from contrasting environments, consistently observed over two consecutive years, remarking the following:
•
The Maamora ecotype (QM) was distinguished by larger heavier acorns, and a higher proportion of pericarp than Bouhachem (QB) ecotype, confirming the influence of ecological factors on morphology and mass distribution.
•
Phytomelatonin content also varied significantly between ecotypes and between fruit compartments (seed vs. pericarp), with overall higher levels in seeds, particularly those of the QM ecotype.
•
Correlation analysis revealed a high degree of consistency between morphological dimensions and the fresh and dry masses of acorn organs, reflecting proportional growth between length, width, thickness, and biomass. In contrast, phytomelatonin content in the pericarp was negatively correlated with most morphometric and weight traits, suggesting an inverse regulatory mechanism linked to tissue development. Conversely, phytomelatonin content in seeds showed a slightly positive correlation with some variables, possibly reflecting an accumulation associated with the reserve functions of embryonic tissue.
These results underline the importance of an integrative approach combining morphometry, secondary metabolite chemistry, and statistical analysis to better understand intra-specific functional diversity in acorns.
With regard to the use of phytogenics, such as melatonin or extracts rich in it (phytomelatonin) [83], there are interesting studies on the benefits of melatonin in different livestock species [46]. The possible use of acorn flour as a source of functional phytomelatonin could be very interesting for supplying the use of synthetic melatonin, mainly in pigs, where many studies have indicated the beneficial use of melatonin in piglets as an immunomodulator and gastrointestinal protector [60,61]. Also in gilts, melatonin treatments improved estrus duration and embryo survival and alleviated the effects of heat stress on gilts during the summer [84]. In sows, melatonin supplementation during gestation could improve maternal–placental–fetal redox status and reproductive performance by ameliorating placental antioxidant status, inflammatory response, and mitochondrial dysfunction [85]. In sows at the late gestational stage, regarding the reproductive performance, melatonin treatments potentially increased the litter size and birth survival rate and significantly increased the birth weight as well as the weaning weight and survival rate of piglets compared with the controls.
To conclude, we can point out that the use of phytogenics in livestock farming promises relevant advances to replace at least gradually the large use of antibiotics and other drugs, and melatonin/phytomelatonin is also presented as a phytogenic in research with promising applications.

Author Contributions

Conceptualization, M.B.A., B.E.B. and M.L.E.K.; methodology, J.H.-R., A.C. and S.K.; software, A.C. and J.H.-R.; validation, M.B.A.; formal analysis, S.K.; investigation, S.K.; data curation, S.K., A.C. and J.H.-R.; writing—original draft preparation, S.K. and M.B.A.; writing—review and editing, S.K., A.C., J.H.-R. and M.B.A.; visualization, S.K., J.H.-R. and A.C.; supervision, J.H.-R., M.B.A. and A.C.; project administration, A.C. and M.B.A. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Data Availability Statement

The data presented in this study are available on request from the corresponding author.

Conflicts of Interest

The authors declare no conflicts of interest.

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Figure 1. Morphological comparison of acorns from the two Quercus suber ecotypes: (A) Maamora sweet acorn (QM) and (B) Bouhachem bitter acorn (QB). On graph paper, thick markings represent 1 cm.
Figure 1. Morphological comparison of acorns from the two Quercus suber ecotypes: (A) Maamora sweet acorn (QM) and (B) Bouhachem bitter acorn (QB). On graph paper, thick markings represent 1 cm.
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Figure 2. Representative chromatogram of standard melatonin measured by HPLC-FLUO (λexc = 280 nm and λemi = 350 nm).
Figure 2. Representative chromatogram of standard melatonin measured by HPLC-FLUO (λexc = 280 nm and λemi = 350 nm).
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Figure 3. Calibration curve for standard melatonin.
Figure 3. Calibration curve for standard melatonin.
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Figure 4. Representative chromatogram of a Quercus suber sample measured by LC-FLUO (λexc = 280 nm and λemi = 350 nm). The melatonin peak at the retention time of 11.2 min is marked.
Figure 4. Representative chromatogram of a Quercus suber sample measured by LC-FLUO (λexc = 280 nm and λemi = 350 nm). The melatonin peak at the retention time of 11.2 min is marked.
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Figure 5. Correlation matrix between morphological parameters (L—length, W—width, T—thickness), weight parameters (FW—fresh acorn weight, DW—dry acorn weight, FSW—fresh seed weight, DSW—dry seed weight, FPW—fresh pericarp weight, DPW—dry pericarp weight). Pericarp (PP) and seed (PS) percentages, as well as phytomelatonin content in seeds (MS) and pericarps (MP) of Quercus suber acorns from two contrasting ecotypes (QM and QB) in two consecutive years (2022/2023 and 2023/2024).
Figure 5. Correlation matrix between morphological parameters (L—length, W—width, T—thickness), weight parameters (FW—fresh acorn weight, DW—dry acorn weight, FSW—fresh seed weight, DSW—dry seed weight, FPW—fresh pericarp weight, DPW—dry pericarp weight). Pericarp (PP) and seed (PS) percentages, as well as phytomelatonin content in seeds (MS) and pericarps (MP) of Quercus suber acorns from two contrasting ecotypes (QM and QB) in two consecutive years (2022/2023 and 2023/2024).
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Table 1. Geographic origin, morphology, and taste of the studied cork oak ecotypes.
Table 1. Geographic origin, morphology, and taste of the studied cork oak ecotypes.
Plant SpeciesQuercus suber L.
ParameterEcotype 1 (Maamora)Ecotype 2 (Bouhachem)
GPS coordinates34°6′44.01′′ N35°11′59.4168′′ N
6°37′51.1896′′ W5°21′7.6644′′ W
Ecosystem typePlainMountain
RegionMaamora Forest
(Rabat-Salé-Kénitra, Morocco)		Bouhachem Forest
(Tanger-Tetouan-Al Hoceïma, Morocco)
Approximate altitudeLow, <200 mHigh, >600 m)
Acorn sizeLargeSmall
Acorn tasteSweetBitter
Traditional useEdible human foodForage use
Table 2. Morphological traits of acorns from two ecotypes (sweet and bitter): Maamora (QM) and Bouhachem (QB).
Table 2. Morphological traits of acorns from two ecotypes (sweet and bitter): Maamora (QM) and Bouhachem (QB).
Years2022/20232023/2024
EcotypesQMQBQMQB
Length (mm)46.36 ± 0.61 a30.97 ± 0.66 b46.12 ± 0.81 a30.18 ± 0.41 b
Width (mm)19.87 ± 0.21 a16.12 ± 0.62 b18.70 ± 0.30 a15.25 ± 0.17 b
Thickness (mm)19.23 ± 0.23 a16.02 ± 0.62 b18.50 ± 0.30 a15.11 ± 0.16 b
Fresh weight (g)11.55 ± 0.33 a6.15 ± 1.10 b11.04 ± 0.49 a4.50 ± 0.15 b
Dry weight (g)7.63 ± 0.23 a2.46 ± 0.10 b6.96 ± 0.29 a3.45 ± 0.39 b
Different lowercase letters (a and b) in the same row indicate statistically significant differences between groups (p < 0.05), according to ANOVA followed by Tukey’s HSD post hoc test.
Table 3. Seed and pericarp mass distribution in sweet and bitter acorns of cork oak.
Table 3. Seed and pericarp mass distribution in sweet and bitter acorns of cork oak.
Years2022/20232023/2024
EcotypesQMQBQMQB
% Pericarp19.48 ± 0.38 b15.82 ± 0.28 d21.85 ± 0.47 a17.28 ± 0.40 c
% Seed80.52 ± 0.38 c84.18 ± 0.28 a78.14 ± 0.47 d82.73 ± 0.40 b
FPW (g)2.19 ± 0.06 a0.65 ± 0.02 b2.37 ± 0.11 a0.76 ± 0.02 b
FSW (g)9.36 ± 0.28 a5.49 ± 1.10 bc8.66 ± 0.40 ab3.75 ± 0.13 c
DPW (g)1.64 ± 0.04 a0.46 ± 0.02 b1.56 ± 0.06 a0.95 ± 0.38 ab
DSW (g)5.99 ± 0.19 a1.99 ± 0.09 b5.40 ± 0.24 a2.50 ± 0.08 b
FPW—fresh pericarp weight. DPW—dry pericarp weight. FSW—fresh seed weight. DSW—dry seed weight. Distinct lower-case letters (a, b, c and d) on the same line indicate statistically significant differences between groups (p < 0.05).
Table 4. Phytomelatonin content in acorn organs of two ecotypes in two consecutive years.
Table 4. Phytomelatonin content in acorn organs of two ecotypes in two consecutive years.
YearsEcotypeOrganPhytomelatonin
(ng/g DW)		Total Phytomelatonin Content (ng/g DW)
2022/2023QMPericarp1.01 ± 0.04 e10.95 ± 0.28 a
Seed9.94 ± 0.24 a
QBPericarp1.86 ± 0.31 e6.74 ± 0.44 b
Seed4.88 ± 0.13 c
2023/2024QMPericarp6.67 ± 0.43 b7.98 ± 0.51 b
Seed1.31 ± 0.08 e
QBPericarp3.09 ± 0.33 d7.46 ± 0.39 b
Seed4.37 ± 0.06 c
Data are represented by the mean (n = 3) ± SD. Distinct lower-case letters indicate statistically significant differences between groups (p < 0.05).
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MDPI and ACS Style

Kaabi, S.; El Bouzdoudi, B.; Kbiach, M.L.E.; Cano, A.; Hernández-Ruiz, J.; Arnao, M.B. Content of Phytomelatonin in Acorns (Quercus sp.) in Its Possible Use as a Phytogenic in Animal Nutrition. Processes 2025, 13, 2202. https://doi.org/10.3390/pr13072202

AMA Style

Kaabi S, El Bouzdoudi B, Kbiach MLE, Cano A, Hernández-Ruiz J, Arnao MB. Content of Phytomelatonin in Acorns (Quercus sp.) in Its Possible Use as a Phytogenic in Animal Nutrition. Processes. 2025; 13(7):2202. https://doi.org/10.3390/pr13072202

Chicago/Turabian Style

Kaabi, Soundouss, Brahim El Bouzdoudi, Mohammed L’bachir El Kbiach, Antonio Cano, Josefa Hernández-Ruiz, and Marino B. Arnao. 2025. "Content of Phytomelatonin in Acorns (Quercus sp.) in Its Possible Use as a Phytogenic in Animal Nutrition" Processes 13, no. 7: 2202. https://doi.org/10.3390/pr13072202

APA Style

Kaabi, S., El Bouzdoudi, B., Kbiach, M. L. E., Cano, A., Hernández-Ruiz, J., & Arnao, M. B. (2025). Content of Phytomelatonin in Acorns (Quercus sp.) in Its Possible Use as a Phytogenic in Animal Nutrition. Processes, 13(7), 2202. https://doi.org/10.3390/pr13072202

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