Influence of Lactobacillus plantarum-aided fermentation on the health benefits and functional role of the leaves of Ceiba pentandra

Abstract

Introduction:

Ceiba pentandra is underutilized, valued for its fiber-rich fruits, medicinal properties, and ecological significance in traditional and industrial uses. This study investigates the influence of Lactobacillus plantarum-aided fermentation on the nutritional prowess and the functional properties of Ceiba pentandra leaves.

Methods:

Dried leaves were fermented for 5 days at 0, 1, 3, and 5 days, and the findings were compared with those of the commonly consumed Corchorus olitorius leaves, serving as the reference. The nutritional and antioxidant qualities were evaluated using standard methods, while the biochemical status (α-amylase and lactate dehydrogenase (LDH)) activities were measured using precision kits. The vitamin and phytochemical statuses were qualitatively and quantitatively monitored using High-Performance Liquid Chromatographic techniques.

Results:

Fermented leaves of Ceiba pentandra were significantly higher (p < 0.05) in pH, bacterial growth, ash, and significantly lower (p < 0.05) in iron, manganese, and crude fiber compared to those of Corchorus olitorius. This fermentation significantly reduced (p < 0.05) antioxidant capacity (FRAP) by 19.11 and 13.63% in Ceiba pentandra and Corchorus olitorius leaves, respectively, which remains comparable. LDH activity significantly increased (p < 0.05) by 11.98 and 2.93% with fermentation duration in Ceiba pentandra and Corchorus olitorius leaves, respectively. Fermented leaves of Ceiba pentandra were notably rich in ginsenoside (717.824 mg/ 100 mg), catechin (129.459 mg/ 100 mg), kaempferol (112.745 mg/ 100 mg), rutin (63.263 mg/ 100 mg), stevioside (60.632 mg/ 100 mg), saponin white (52.617 mg/ 100 mg), caffeic acid (48.909 mg/ 100 mg), and resveratrol (9.037 mg/ 100 mg) in decreasing order among the sixteen bioactives detected.

Conclusion:

The fermented leaves are nutritionally rich and have health benefits, while LDH is the biochemical agent engaged by the probiotic organism.

1 Introduction

Fermentation is an ancient food preservation method based on the production of microorganisms, such as bacteria, yeasts, or fungi, that break down complex materials into simpler substances, including organic acids, alcohols, and organic esters. Fermentation, along with other conventional food processes such as cooking, smoking, and drying, helps extend the shelf life, enhance flavor, and improve the nutritive value of food. Industrial fermentation in the production of goods, such as beer and bread, was already well established in ancient Mesopotamia and Egypt thousands of years ago. This process is an easy, effective technique, as the human being does little, and most of the work is done by the microorganisms involved (Teng et al., 2021). The new changes in fermentation technology have significantly improved the means of preserving fermented food products, their safety, and the associated norms. These developments have transformed production methods, enhanced quality management, and boosted overall safety in the production of fermented products (Teng et al., 2021). Fermentation comprises a sequence of biochemical transformations that alter the chemical makeup of the substrate. This process has recently been an effective means of enhancing the concentration of bioactive constituents and improving the functional attributes of plant-derived food materials (Zhao et al., 2021).

During fermentation, lactic acid bacteria (LAB) degrade complex phenolic compounds in plant leaves into simpler forms, thereby increasing antioxidant activity depending on the type of phytochemicals generated. Antioxidants are not new solutions that protect human health at the individual level, as they can chelate harmful free radicals that contribute to the development of various diseases, including cardiovascular diseases and metabolic disorders (Saritas et al., 2024). In addition, phenolic-rich fermented extracts have demonstrated their capacity to inhibit α-amylase activity, providing evidence of a dual action that controls a surge in blood sugar levels and reduces oxidative stress (Cisneros-Yupanqui et al., 2023). The concern, particularly regarding the accessibility of environmentally friendly, non-toxic natural food sources, has been increasing and is driven by accelerated global population growth (Knorr and Augustin, 2023; Huang et al., 2026). LDH, an important enzyme in anaerobic glycolysis, plays a vital role in microbial fermentation processes, influencing metabolite production and redox balance. LDH is importantly involved in anaerobic processes, especially when rapid and excessive mobilization of energy is needed to overcome the limitations of the glycolytic process. It is also a broad-spectrum enzyme capable of metabolizing numerous substrates (Harle et al., 2020; Odutayo et al., 2020).

Although the leaves of Ceiba pentandra have nutritional potential, they are not well-known in most parts of the world (Huang et al., 2026). There is also little literature on the extent to which fermentation, especially with lactic acid bacteria, influences the biochemical composition of the leaves. Although it has been reported that fermentation positively influences the bioavailability of phytochemicals, decreases known anti-nutrients, and alters enzyme activities in plant-based foods (Samtiya et al., 2021; Knez et al., 2023), there is scarce information on the impact of these alterations on the antioxidant potential and enzyme mechanisms of the raw leaves of C. pentandra.

Although C. pentandra leaves have traditionally been used in local foods and medicine, little scientific research has been conducted on the effects of fermentation on their nutritional and health values. As established, lactic acid fermentation significantly increases the observed antioxidant capacity of plant-based materials, as reflected in enhanced free radical scavenging exhibited by fermented herbal teas, sweet potatoes, and avocado leaves. The overall increase in antioxidative capacity also helps diminish oxidative stress, which is critical in the development of chronic conditions, including heart disease (Ozturk et al., 2024). In addition, compounds with phenolic content derived from plants through fermentation have been shown to inhibit enzymes such as a-amylase, which breaks down carbohydrates, providing another possible mechanism for controlling blood sugar levels. Lactic acid bacteria fermentation of plant foods can increase the bioavailability of nutrients and minerals (Nsabimana et al., 2024; Saritas et al., 2024).

The exponential increase in the global population, projected to reach 8.6 billion by 2030 and 9.7 billion by 2050, poses a challenge for humans to expand the present spectrum of leafy vegetables to provide the necessary micronutrients and bioactives to sustain their health requirements (UN-DESA, 2017; Galan, 2025). Efforts to combat such future food insecurity were target at devising or adopting processing techniques to assist in transforming the under-utilized and wild plants including the leaves of Solestermon monostachyus and Cynthea dregie, and the seeds of Chrysophyllum albidum, Adenanthera pavonina, Hura crepitans and Carica papaya for edible purposes (Afolabi et al., 2015; Afolabi et al., 2018; Odutayo et al., 2020; Nwinyi and Hassan, 2021; Afolabi et al., 2023a; Afolabi et al., 2023b; Odutayo et al., 2023; Afolabi et al., 2025; Huang et al., 2026). The leaves of C. petandra are found in several geographical locations across the globe and have been reported to have superior phytochemical levels relative to the well-established medicinal leaves of Moringa oleifera and Cymbopogon citratus, but are less utilized than other leaves. Thus, it is proposed to possess good medicinal benefits, such as anticonvulsant, antimeasles, antistings, and antidiabetic (Te et al., 2020; Sarfo et al., 2022; Tareau et al., 2022; Asigbaase et al., 2024). The leaves of this species were selected for study due to their regular consumption among the Yoruba-speaking Okun people of Kogi State, Nigeria. Hence, requires the attention of food scientists for further investigation. This study seeks to establish a scientific basis for fermentation during the processing of C. pentandra leaves to make them safer and more edible. This study aimed to investigate the influence of L. plantarum-aided fermentation on the nutritional profile and functional properties of C. pentandra leaves, using the commonly consumed Corchorus olitorius leaves as a reference. An attempt was also made to account for the possible biochemical mechanism(s) the organism may engage to facilitate the process.

2 Materials and methods

2.1 Materials

This study engaged the following high-purity and analytical-grade chemicals and reagents: methanol, DPPH (1,1-diphenyl-2-picrylhydrazyl) powder, amylase kits, lactic acid dehydrogenase kits, ethanol, De Man Rogosa and Sharpe (MRS) agar, trichloroacetic acid (TCA), and potassium ferricyanide. All other standards for the phytochemical used were of HPLC grade and procured from Sigma-Aldrich, USA.

2.2 Methods

2.2.1 Collection and preparation of the plant leaves

The leaves of Ceiba pentandra were extensively harvested in Makutu-Isanlu town, Kogi State, Nigeria, where they were consumed as vegetable soup and locally called akuku soup, deposited and identified (voucher number: UILH/001/1844/2025) by the Herbarium unit of the Department of Plant Biology, University of Ilorin. The leaves were air-dried for 2 weeks, then blended to prepare for fermentation. Fresh leaves of the commonly consumed Corchorus olitorius were procured at the Ota market in Ogun State, Nigeria, to serve as the reference. The stalks were placed in water, their leaves removed, thoroughly washed, and then left to dry in the air for 2 weeks to remove any remaining moisture. After thorough drying, the leaves were crushed into a fine, dry powder to increase surface area and facilitate further analysis. Also, an ethical permit (CHREC/1185/2025) from the Covenant Health Research Ethics Committee, Canaanland, Ota, Nigeria, was given for the research to proceed.

2.2.2 Gathering and sub-culturing of lactic acid bacteria

The lactic acid bacteria (LAB) species (Lactobacillus plantarum) was obtained from the Molecular Biology Laboratory at Covenant University and used for this study after confirming the characteristics as non-hemolytic, catalase-negative, and Gram-positive rods. The isolate was resuscitated in MRS broth and grown in Brain Heart Infusion (BHI) broth at 37 °C under shaking conditions (200 rpm) for 4–6 h. The inoculum was prepared by centrifuging a scoop of freshly cultured LAB twice at 10,000 g x 10 min after suspending it in 2.0 mL of sterile saline solution. An aseptic introduction of a 2 mL aliquot of this inoculum, which contained roughly 2 × 10⁶ CFU/mL, was made into sterile 500 mL conical flasks that contained 300 mL of a sterilized solution made up of 3% NaCl, 3% glucose, and 20 g of dried material that had already been blended. An uninoculated flask was kept as a control, and the flasks were incubated at 37 °C for 24, 72, and 120 h. Each stage of the fermentation of the leaves, including the reference leaves, was set up in three sterilized 250 mL conical flasks placed in a holder of a Labscience Thermostat Incubator with Shaker (ZHP-100, England), rotating the content at 180 rpm until the set fermentation time (0, 3, and 5 days) for the inoculated probiotic bacteria to have one phase interaction with the content. Using colony count analysis, bacterial growth was tracked at each fermentation interval (Odutayo et al., 2023).

2.3 Procedure for the fermentation of the plant leaves

2.3.1 Sterilization of materials for use during fermentation

Hermetically sealed materials were exposed to saturated steam at 121 °C for at least 30 min under a pressure greater than 15 psi using an autoclave (model SM2080E, England).

2.3.2 Post-fermentation analysis

The plants were fermented for 5 days, with samples collected on days 0, 1, 3, and 5. After fermentation, the samples were blended and filtered using a muslin cloth to obtain a fermented solution. Both the filtrate and the solid residue were gathered and placed in sample bottles for subsequent analysis. The residue was used to analyze the proximate composition, and the recovered ash was further used to estimate the mineral composition. In contrast, the filtrate was used to analyze the effects of fermentation on antioxidant potential, phytochemical, biochemical (α-amylase and LDH activities), microbial, and vitamin profiles.

2.4 Analysis of physicochemical parameters in processed plant products

2.4.1 pH value

The hydrogen ion concentration of the extracts obtained from the leaves of both Ceiba pentandra and Corchorus olitorius was routinely measured using a pH meter (model pHep, Villafranca Padovana, Italy) as previously described (Afolabi and Oloyede, 2011).

2.4.2 Estimation of bulk-nutrient composition of the leaves

Moisture, ash, and crude fiber constituents were estimated using AOAC methods (AOAC, 2015). Proteins and carbohydrates were estimated spectrometrically as described below.

2.4.3 Procedure for protein estimation

The previously established biuret method was used to measure protein concentration (Liu and Pan, 2017). A portion of distilled water (1.0 mL) was pipetted into the well labeled blank test tubes, while 1.0 mL of standard protein solution was pipetted into the standard tubes and the corresponding test tubes to those of the sample. Each tube was filled with 4.0 mL of freshly prepared Biuret reagent after the standards and samples were prepared. The test tubes were left to stand for 30 min, and the absorbance of each solution was measured at 540 nm using a spectrophotometer (model M-26 V, serial no. 97–0183-02). Finally, their corresponding concentration (μg/ml) was extrapolated from the best line derived from a prepared standard curve (y = 0.3635x; R² = 0.7947) for Bovine Saline Albumin.

2.4.4 Procedure for carbohydrate estimation

The carbohydrate content was determined spectrophotometrically by following the Anthrone method (Ilodibia et al., 2013). Each prepared sample extract (45 μL) was mixed with distilled water (450 mL). Following this, the test tubes for the sample extract (1.0 mL) obtained from the filtrate, distilled water (1.0 mL), and a glucose solution (1.0 mL) were dispensed, serving as the sample, blank, and standard, respectively. To each of the test tubes, a freshly prepared 0.10% Anthrone reagent (5.0 mL) was later dispensed, mixed thoroughly by gently shaking, and allowed to stand in a water bath (30 °C) for 12 min. After which, the mixture was removed and cooled to room temperature. The absorbance of the samples and the standards was measured at 630 nm against the blank using a spectrophotometer. The corresponding concentration (μg/ml) of the absorbance was calculated using the equation (y = 0.0015x + 0.0129; R² = 0.9706), which was derived from the best line on a standard graph plotted from the absorbance of each standard against its corresponding concentration.

2.5 In vitro determination of antioxidant activity

Antioxidants neutralize reactive oxygen species (ROS), thereby protecting cell membrane integrity and preventing damage that can lead to various transmissible and chronic diseases.

2.5.1 DPPH assay for measuring free radical scavenging activity

The antioxidative strength of plant extracts was determined using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) method as previously described (Sakat and Juvekar, 2010). Enough methanol was added to the extract stock solution (10 mg/mL) to achieve a final volume of 1.5 mL. An aliquot of the DPPH solution (0.1 mM, 0.5 mL) was added to the diluted samples, and 0.5 mL of the DPPH solution mixed with methanol alone was used as the control. The blends were left to stand at room temperature in the dark for 30 min. The absorbance at 517 nm was then quantified using a UV–Visible spectrophotometer (model M-26 V, serial no. 97–0183-02), with methanol as the blank. Free radical scavenging activity (percentage) was calculated assuming the following (Equation. 1):

Where;

Ac = absorbance of control.

As = absorbance of the sample.

2.5.2 Determination of antioxidant activity (ferric reducing antioxidant power)

A previously described standard procedure was adopted for this analysis (Sharma et al., 2016). In test tubes, for the Ferric Reducing Antioxidant Power test, 2.5 mL of 0.2 M phosphate buffer and 2.5 mL of 1% potassium ferricyanide were mixed with extracts at different concentrations. After thoroughly mixing the solutions, they were incubated in a water bath at 50 °C for 20 min. 10% trichloroacetic acid (TCA) was added in 2.5 mL following incubation. Separately, FeCl₃ and distilled water were used to create a ferric chloride solution. The TCA-treated samples were combined with the FeCl₃ solution after a 10-min rest time. A UV–Visible spectrophotometer (Thermo Scientific Ltd., model M-26 V, serial no. 97–0183-02) was then used to measure the absorbance of the final solution at 700 nm, using ascorbic acid as the standard to assess reducing power. Distilled water served as the blank for this experiment. The equivalent concentration (μg/mL) of the absorbance obtained for the samples was extrapolated from a standard graph of ascorbic acid (R² = 0.998; y = 0.0089x + 0.006) prepared using different ascorbic acid standard values of 0, 0.2, 0.4, 0.6, and 0.8 μg/mL.

2.6 Procedures for biochemical assessments

2.6.1 Determination of α-amylase activity

The Procedure described in the manufacturer’s manual for the α-amylase kit was strictly adhered to. The α-amylase activity was evaluated using a chromogenic CNPG3 tablet dissolved in amylase buffer. The buffer solution was preheated to 37 °C to ensure complete dissolution. One substrate tablet was placed into a clean test tube or volumetric container. The full volume of the provided amylase buffer (50 mL) was added as specified in the kit manual, and the mixture was gently swirled until the tablet dissolved completely, forming the working reagent. For each test, pipette 500 μL of this working reagent into a reaction tube and add 10 μL of the sample. Mix gently and incubate the reaction mixture at 37 °C. The absorbance at 405 nm was measured immediately after sample addition, then at 30-s intervals for 2 min. The rate of increase in absorbance (ΔA/min) corresponds to the enzyme activity. To calculate α-amylase activity in units per liter (U/L), multiply ΔA/min by the factor provided by the kit, such as 3,105.

2.6.2 Procedure for the assay of lactate dehydrogenase (LDH) activity

The manufacturer’s Procedure, a Randox kit, was used to assess LDH activity in the fermented filtrate from the leaves. An aliquot of the sample (0.02 mL) was mixed with 1 mL of the provided reagent, which consisted of NADH and a substrate/buffer for the reaction. The reaction was allowed to proceed for 30 min, after which the initial absorbance was measured and subsequently monitored at 60-s intervals for 180 s at 37 °C and 340 nm. The following semi-micro precision (Equation. 2) was used to calculate the enzyme activity.

2.7 Mineral analysis

2.7.1 Sample preparation for mineral analysis

The extracted leaf sample (1.0 g) was put into a conical flask. A solution (30 mL) of nitric acid and hydrochloric acid was dispensed into the conical flask and heated in a 3:1 ratio to produce a clear solution. It was allowed to cool to ambient temperature, then filtered through a 125 mm Whatman filter paper and placed into a 50 mL container. After adding 50 mL of deionized water to the solution, the metal content of the prepared stock sample (0.02 g/ mL) was examined using Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES).

2.7.2 Procedure for the mineral analysis

ICP-OES for the Analysis of Heavy Metals and Trace Elements. Trace elements and heavy metals were analyzed using the Agilent 5,800 ICP-OES probe, following the parameters: plasma gas flow rate of 15 L/min, auxiliary gas flow rate of 0.2 L/min, instrument power of 1,450 W, and equilibration time of 15 s with a 45 s read delay. The internal calibration apparatus was calibrated using reference concentrations of heavy metals and trace elements. The argon gas instrument set up at 588.995, 766.491, 393.366, 279.553, 196.026, 238.204, 202.548, 324.754, 220.353, 257.610, and 214.439 nm wavelengths to assess sodium, potassium, calcium, magnesium, selenium, iron, zinc, copper, lead, manganese, and cadmium mineral levels of the leaves, respectively. The concentration of each mineral was automatically extrapolated from a prepared internal standard curve derived from five different concentrations: 0.1, 0.25, 0.5, 2.5, and 5.0 ppm of the appropriate standards for each element measured.

2.8 Vitamins determinations by HPLC

The selected five vitamins (A, B₂, B₉, C, and E) components of the leaves were measured using the methodology outlined by Afolabi et al. (2023b). The three separately filtered extracts obtained for each of the fermentation period were combined, mixed vigorously and assessed qualitatively and quantitatively with the aid of an Agilent HPLC 1260 Infinity II series equipped with a Quat Pump VL G711A (DEAEY01907), a column (Poroshell 120 EC C18 4um, 150×4.6 mm), Oven G7130 (DEAEQ22974), a DAD detector WR G71115A (DEAC606992), and an Auto Sampler G7129A (DEAEQ22974). The following concentrations, 2.5, 5, 10, 20, 40, and 80 mg/L of the vitamins standard, were prepared and plotted against their corresponding peak area, 434.188, 1088.03, 1551.385, 3524.667, 6198.632, and 12679.50 mAU obtained respectively, to derive a standard curve (R² = 0.9985) where the resultant absorbance generated by the samples was extrapolated to derive their concentrations. An aliquot of each of the membrane filtered extracts and the prepared standards (20 μL; 10% w/v in methanol) for vitamins analysis was injected using the auto sampler into the HPLC system with a mobile phase that consisted of a mixture of formic acid (0.1%) and absolute methanol (70:30), subjected to a steady flow rate of 1.0 mL/min, a DAD detector wavelength of 245 nm, an oven temperature of 35.0 °C, and for a run duration of 10 min to derive the chromatograph for each sample (Supplementary Figures S1–S3).

2.9 Phytochemical determinations by HPLC

The phytochemical components of the leaf extracts were measured using the methodology outlined by Afolabi et al. (2025). The extracts were assessed both qualitatively and quantitatively using an HPLC (model: Agilent LC-8518) integrated with a Biobase software (N2000 Chromatography Data System) library.

2.9.1 Sample and standard curve preparation for flavonoids analysis

1.0 g of the substance was extracted from a volumetric flask using 70% methanol (v/v). This mixture was filtered twice using Whatman filter paper no 1. After 41 h, it was then sonicated for 10 min. The resulting filtrate was diluted with the extraction solution to reach a final concentration of 1 mg/mL for HPLC measurement of flavonoids (Garg, 2021). A portion of 10 mg of quercetin dihydrate, luteolin, rutin, and catechin was weighed and transferred to a 10.0 mL volumetric flask, and the volume was adjusted with methanol to the mark to obtain a standard stock solution of 1,000 ppm. Then, 1 mL of each stock solution was diluted to 10.0 mL to prepare a working standard solution. Different concentrations of the solution (0.5, 1.0, 1.5, 2.0, 2.5 mL) from the standard were transferred into 10 mL volumetric flasks and made up to 10 mL with methanol. Standard solutions at 5, 10, 15, 20, and 25 μg/mL were used to construct the calibration curve.

2.9.2 Sample and standard curve preparation for HPLC analysis for saponins analysis

After weighing the samples, a 0.25 mg aliquot was dissolved in 25 mL of absolute methanol. The materials were filtered through a 0.45 μm membrane filter and then placed in glass containers for further analysis. A portion of 10 mg of the standard was weighed and transferred to a 10.0 mL volumetric flask, and the volume was adjusted with absolute methanol to the mark to obtain a standard stock solution of 1,000 ppm. Then 1 mL of each stock solution was taken and diluted with the same solvent to prepare a standard solution of (2, 4, 6, and 8.0 μg/mL), which was used for the calibration curve preparation.

2.9.3 Preparation of the sample and standard curve for phenolics analysis

The leaf extract (10 mg) was dissolved in the mobile phase (10 mL), stirred intermittently for 8 h, then sonicated for 30 min, and filtered through a 0.2 μm filter. Various phenolic standards (10 mg) were dissolved in 10 mL of mobile phase to prepare stock solutions at 1000 μg/mL. A series of dilutions at concentrations of 20, 30, 40, and 50 μg/mL was prepared by taking aliquots of 0.2, 0.3, 0.4, and 0.5 mL of the stock solution (1,000 μg/mL) and diluting to 10 mL with mobile phase. Each dilution (20 μL) was set up in triplicate, and the area under the curve at 270 nm was recorded after injection with a syringe. A calibration curve of mean area versus concentration was plotted to obtain a standard graph, from which the corresponding concentrations of the analyzed samples were extrapolated.

Also, the LOD and LOQ for phenolics 0.0004 and 0.001 ppb; flavonoids, saponins analysis were 0.003 ppb and 0.02 ppb; 0.005 and 0.001 ppb, respectively.

2.9.4 Chromatographic conditions for each phytochemical analysis

The phytochemical components of the leaves were identified and quantified using HPLC with a reversed-phase column; a 200 μL eluent supply pump was used for the chromatographic analysis. The system consists of a Biobase-type UV–Vis detecting device with a preset wavelength. Separation was achieved using a five μm ultra-pure silica packing material (Vertex) on a 4.6 mm × 250 mm C18 reversed-phase column under isocratic conditions. The mobile phase isocratic elution with a high-pressure gradient is carried out using a volume ratio of 30:70 (v/v) of water and acetonitrile (for saponins); acetonitrile, methanol and acetic acid (50:45:5, v/v/v) for flavonoids; and methanol and water in ratio of 50:50 (v/v) for the phenolics estimation at a flow rate of 1.0 mL/min and an injection volume of 20 μL. Each prepared solution was degassed and then passed through a membrane filter with pores as small as 0.45 μm. The UV detector was set at 203, 270, and 270 nm for saponin, phenolic compounds (including resveratrol), and flavonoids, respectively.

Meanwhile, the column was maintained at room temperature throughout the analysis. The LC ran for a total of twenty minutes. The concentration of various phytochemicals was extrapolated from an internally generated standard curve from a prepared phenolic (1,000 μg/mL) standard (Supplementary Figures S4–S12). The mean area was plotted against the concentration to construct an internal tannic acid standard solution calibration curve within the LC system, from which the concentrations were automatically extrapolated (Garg, 2021).

2.10 Statistical analysis

The results obtained on each fermentation day were statistically analyzed using the one-factor ANOVA package in MegaStat (version 10.3, release 3.2), which was integrated with Microsoft Excel (2016). The level of significance for the fermentation treatments was set at the 95% confidence level, and the pairwise t-tests package was used for Post hoc analysis to compare the means of relevant paired sample groups versus the reference leaves or control. The results were finally expressed as mean ± SD.

3 Results

3.1 Influence of fermentation technique on pH, bacterial growth, and bulk nutrients in leaves of Ceiba pentandra and Corchorus olitorius

The fermenting media progressively become more acidic in both leaves as fermentation progresses, indicating the release of acidic compounds, such as organic acids (Table 1). The leaves of C. pentandra support significantly higher (p < 0.05) bacterial growth (7.01–9.10 × 10⁶ CFU/mL) throughout fermentation than those of C. olitorius (5.81–8.00 × 10⁶ CFU/mL), indicating the possible presence of favorable compounds that can serve as substrates for the fermenting organism. This favorable growth is despite consistently lower moisture content in C. pentandra leaves than in C. olitorius. Carbohydrate contents increase with the progression of fermentation in both leaves, which is similar to the trend observed for the proteins, crude fiber, and ash contents in both leaves. Still, they are generally more pronounced in the leaves of C. olitorius (Table 1).

3.2 Influence of fermentation technique on micronutrients and heavy metals in leaves of Ceiba pentandra and Corchorus olitorius

The leaves of C. pentandra exhibited significantly lower (p < 0.05) iron (646.98 mg/L) and manganese (27.10 mg/L) contents compared to those of the C. olitorius throughout the five-day fermentation period. The values of the other minerals and vitamins in the leaves of the C. pentandra remain relatively comparable to those of C. olitorius throughout the five-day fermentation period (Table 2). The probiotic bacteria (L. plantarum) increased the amount of vitamins (Vit B₉, A, and C) with the extension of fermentation of C. olitorius leaves by 19.27, 283.95, and 10.43%, respectively, indicating the potential of the organism to synthesize these vitamins under these fermentation conditions and in the presence of certain molecules in the leaves that can serve as precursors required to synthesize these vitamins. A general increase in vitamin B₉ was observed with increased fermentation of the leaves of C. olitorius and C. pentandra, indicating the possible existence of molecule(s) that can serve as precursors for the probiotic bacteria to synthesize vitamin B₉.

A general increase in the levels of vitamins A and E occurred with the extended fermentation of the leaves of C. olitorius and C. pentandra, suggesting the presence of molecules that may serve as precursors for the probiotic bacteria to synthesize both vitamins. A general decrease in vitamins B₂ and C levels was observed with increasing fermentation of C. pentandra leaves. On the contrary, the levels of vitamins B₂ and C increased with the increasing duration of fermentation in C. olitorius, indicating the existence of possible molecules that can serve as the precursor for the probiotic bacteria to synthesize both vitamins.

3.3 Influence of fermentation technique on antioxidant prowess of the leaves of Ceiba pentandra and Corchorus olitorius

Table 3 revealed that the values (DPPH) of antioxidant quality of the leaves of C. pentandra remained relatively the same (p > 0.05) as those of the reference (C. olitorius) leaves, suggesting that it can sustain the health of consumers just as the C. olitorius leaves do to serve as a vegetable. Generally, the antioxidant (FRAP) values significantly decreased (p < 0.05) with extended fermentation time by 19.11 and 13.63% in C. pentandra and C. olitorius leaves, respectively. The antioxidant quality (FRAP) value for C. pentandra leaves was significantly higher (p < 0.05) in the first three days of the fermentation compared to reference leaves (C. olitorius). This antioxidant quality (FRAP) value decreased significantly (p < 0.05) compared with reference leaves (C. olitorius) by the end of the 5 days, suggesting a loss of a portion of its potent bioactive metabolite due to the fermentation technique (Table 3).

3.4 Biochemical enzymes deployed during the fermentation of the leaves of Ceiba pentandra and Corchorus olitorius

The influence of the fermentation techniques on the α-amylase activities in the leaves of C. pentandra was relatively the same (p > 0.05) compared to the reference (C. olitorius) leaves throughout the 5 days, suggesting a limited involvement of the enzyme in the fermentation process (Table 3). However, the influence of LDH activities was generally significantly higher (p < 0.05) in C. pentandra leaves by 11.98% during fermentation than the 2.93% noticed in C. olitorius leaves, which may account for the more rapid loss of potent bioactive metabolites observed in this study for C. pentandra leaves (Table 3).

3.5 Influence of the fermentation technique on phytochemicals detected in the leaves of Ceiba pentandra

This L. plantarum-aided fermentation technique facilitated the partial catabolism of caffeic acid, ellagic acid, gallic acid, stevioside, catechin, rutin, and an unidentified flavonoid compound (UD-Flavo2). Hence, this technique demonstrated the potential to moderate the levels of these compounds when in excess (Table 4). Conversely, the L. plantarum-aided fermentation technique facilitated the biosynthesis of resveratrol, epicatechin, saponin White, kaempferol, an unidentified phenolic compound (U-PHE1), and two unidentified flavonoid compounds (UD-Flavo1 and UD-Flavo4). The fermented leaves of C. pentandra were notably rich in ginsenoside (717.824 mg/ 100 mg), catechin (129.459 mg/ 100 mg), kaempferol (112.745 mg/ 100 mg), rutin (63.263 mg/ 100 mg), stevioside (60.632 mg/ 100 mg), saponin white (52.617 mg/ 100 mg), caffeic acid (48.909 mg/ 100 mg), and resveratrol (9.037 mg/ 100 mg) in decreasing order among the sixteen bioactives detected.

4 Discussion

4.1 Influence of fermentation technique on pH, bacterial growth, and bulk nutrients in leaves of Ceiba pentandra and Corchorus olitorius

Both C. pentandra and C. olitorius exhibited pH reductions after 5 days of fermentation by the acid-producing microbes, mainly lactic acid bacteria. C. pentandra began with a slightly acidic pH of 6.61 and further decreased thereafter to a stronger acidic pH of 4.40 on day 5, an indication that it was more slowly acidified in comparison to C. olitorius, which occurred more rapidly (Shumye Gebre et al., 2024). The higher pH values in the fermented C. pentandra leaves indicate either slower fermentation in the acidic region or the presence of buffering agents to resist the change. These results are consistent with what has been found before on raw green leafy vegetables (unfermented), which in most cases provide a pH range of 3.5–4.5, which is supposedly ideal when attempting to increase the shelf life and ensure microbial safety (Jin et al., 2025; Huang et al., 2026). This acidification affects major nutrients, suggesting the conspicuous influence of this fermentation technique on carbohydrates, crude fiber, and ash in this study (Norozi et al., 2021; Jemere et al., 2025). A minimum of 10⁶ colony-forming units per mL (CFU/mL) must be achieved for a food product to be labeled as probiotic (Zuntar et al., 2020), which falls within the probiotic concentration range recorded in this study (Table 1). Such a high probiotic dose may be appropriate to quicken their action, especially to deliver health benefits more quickly (Zuntar et al., 2020). However, it is expected that using higher probiotic doses than those applied in this study may encourage more rapid biotransformation and nutritional changes, leading to a shorter fermentation time than the 5 days used in this study. It is also appropriate to give allowance for possible growth of probiotic organism during storage of processed products to avoid harmful effects, such as systemic infection, gene transfer, excessive immune stimulation in susceptible individuals, including infants, the elderly, and immunocompromised individuals, and deleterious metabolic activities that can be attributed to their growing beyond the affordable limit (Doron and Snydman, 2015; Sarita et al., 2025). Low levels of probiotics (7.16 log cfu/ml) were reported during the processing of apples, pears, and carrots, as well as other beverages, likely due to this (Rahman et al., 2023). The probiotic organism can grow to a level that elicits a beneficial effect if administered at a low dose (Binda et al., 2020). An inoculum concentration (3%) of L. plantarum and L. acidophilus was considered appropriate for papaya juice production. However, final output can grow to as much as 10⁶–10¹⁰ cfu/mL depending on the processed products (Subhashree and Kavita, 2019; Jagelavičiūtė et al., 2025). This pilot study, which focuses on how probiotic organisms modify the nutritional phytochemical components according to their preference behavior, is useful for guiding food scientists in standardization during production. Also, the 2.35 and 2.74 mg protein/g dry wt. leaves of C. olitorius recorded for the raw leaves and the 5-day fermented leaves in this study, respectively, were comparable to 0.0207% dry wt., but generally lower than 3.8–24.0% reported by others (Choudhary et al., 1973; Osei-Owusu et al., 2023; Bonnot et al., 2025). Hence, giving credence to the overall findings of the proteins in the C. pentandra, which is the focus of this study. However, it is worth noting that vegetables are consumed primarily for micronutrients and their crude fiber, not for protein (Slavin and Lloyd, 2012).

4.2 Influence of fermentation technique on micronutrients and heavy metals in leaves of Ceiba pentandra and Corchorus olitorius

The leaves of C. pentandra reduced in iron and manganese contents relative to those of the C. olitorius throughout the five-day fermentation period. A similar reduction in mineral values, especially heavy metals, was recently reported (Millena et al., 2023). Organisms facilitating fermentation, such as Enterococcus faecium and Enterococcus faecalis, are typically implicated in consuming these minerals based on their preferences, which are influenced by the types of compounds available during the process (Odutayo et al., 2020; Odutayo et al., 2021a; Odutayo et al., 2021b; Afolabi et al., 2023a; Afolabi et al., 2023b). The probiotic bacteria (L. plantarum) exhibited the potential to synthesize vitamins (Vit B₉, A, C, and E) during the fermentation of C. olitorius leaves (Sun et al., 2019; Effiong et al., 2023). Some bacteria and fungi, such as Ashbya gossypii and Bacillus subtilis, have been implicated in vitamin synthesis (Revuelta et al., 2016; Fang et al., 2018; Wackett, 2018; Keyvan et al., 2025). The organism also exhibited the potential to catabolize vitamin B₂ and vitamin C as observed during the fermentation of C. pentandra leaves in this study, which is contrary to the synthesis of the same vitamin B₂ observed during the fermentation in C. olitorius, which suggest that the metabolism of vitamins among other metabolites may depends on the types of metabolites serviced by the leaves or other plants to serve as the precursor for the probiotic bacteria (Odutayo et al., 2021b; Afolabi et al., 2023a; Afolabi et al., 2023b; Odutayo et al., 2023; Afolabi et al., 2025).

4.3 Influence of fermentation technique on antioxidant prowess of the leaves of Ceiba pentandra and Corchorus olitorius

A high DPPH value (IC50) suggests that the materials have low antioxidant activity. DPPH assay methods operate at neutral pH, whereas FRAP is more suitable for the acidic pH conditions observed in this fermentation study (Table 3), explaining the relatively stable and redundant antioxidant activities observed in the DPPH assays of both leaves (Schlesier et al., 2002; Zhao et al., 2021). On the other hand, evaluation of antioxidant activity using the FRAP analytical technique revealed a reduction in antioxidant activity as fermentation time increased in both C. pentandra and C. olitorius leaves. Although the antioxidant prowess of the leaves of C. pentandra was relatively similar to that of the commonly consumed C. olitorius, suggesting that their bioactive components have similar health benefits (Abeyrathne et al., 2022; Effiong et al., 2024). These reductions in antioxidant prowess recorded with increasing fermentation period in both leaves are most probably induced by processes of oxidative degradation or microbial metabolism, and by antioxidants, which, in the long term, lead to a reduction in the concentration and potency of the redox-active molecule. The same trend has been reported, with prolonged fermentation or storage leading to the degradation of phenolic compounds, thereby reducing the overall antioxidant capacity (Erskine et al., 2023; Saritas et al., 2024). The antioxidant potential of both leaves decreased with fermentation duration relative to their unfermented counterparts. However, while the results focus on the effect of the fermentation process, the final antioxidant values (6.70 ± 0.28 and 6.97 ± 0.28 for C. pentandra and C. olitorius, respectively) of the leaves at the end of the 5-day fermentation indicate that their antioxidant strengths remained and were not completely eliminated, which were valuable for mopping up free radicals in both leaves. It is pertinent also to note that this antioxidant value for the leaves of C. pentandra was also comparable to that of the already established health-beneficial C. olitorius leaves. The gradual loss of antioxidant strength follows the general expectation where food processing is reported to deplete nutritional and antioxidant values through leaching of soluble compounds, especially when the processing involves water, as in blanching (Wickramasinghe et al., 2020; Maila and Tseke, 2024; Nchangwe et al., 2024; Miškec et al., 2025). The health benefits of the reference leaves (C. olitorius) were well established and reported to include anticancer, wound healing, management of cardiovascular diseases, gastric ulcer, malaria, and typhoid fever, among others (Biswas et al., 2020; Tosoc et al., 2021; Biswas et al., 2022; Dania et al., 2024; Aboulthana et al., 2026). Hence, these health-related attributes linked to the leaves of C. pentandra in this study were appropriate since the relevant nutritional and antioxidant parameters measured were comparable to those of the established reference leaves.

4.4 Biochemical enzymes deployed during the fermentation of the leaves of Ceiba pentandra and Corchorus olitorius

The leaves of C. pentandra and C. olitorius exhibited essentially equivalent α-amylase inhibitory activities, indicating that the lactic acid bacteria facilitating this process do not have a preference for the enzyme under these fermentation conditions used in this study (de Paulo Farias et al., 2021). However, it is evident that the probiotic organism actively engaged LDH to drive this fermentation for 3 days, with the process occurring in the first three days for C. olitorius and in the last three days for C. pentandra leaves (Table 1). The enzyme facilitates the conversion of pyruvate to lactate and can also metabolize numerous other phytochemicals due to its broad-spectrum nature. The fermenting organism increased LDH activity, likely facilitating the metabolism of phytochemicals in the leaves. Increased LDH activity leads to increased lactate formation, which can restore NAD⁺ and sustain glycolysis during stress, such as low oxygen circulation or high microbial activity (Piechowiak et al., 2023; Ozturk et al., 2024). α-amylase plays little part in this fermentation process since its activity was similar to that of their corresponding unfermented leaves at any of these processing stages. Hence, the enzyme was not a biochemical agent driving this process. However, this does not rule out the possibility that the same probiotic organism could engage other enzymes, including α-amylase, under different fermenting conditions and in the presence of other sources of phytochemicals. The mode of action of fermenting organisms depends on the nature of the phytochemicals available, which vary with source type and fermenting conditions (Afolabi et al., 2023a).

4.5 Influence of the fermentation technique on phytochemicals detected in the leaves of Ceiba pentandra

Catabolism is a degradation process, and partial transformation or modification of phytochemicals can occur through biosynthesis or catabolism (Judge and Dodd, 2020). This L. plantarum-aided fermentation technique may facilitate the partial catabolism of caffeic acid, ellagic acid, gallic acid, stevioside, catechin, rutin, and an unidentified flavonoid compound (UD-Flavo2). Hence, this technique demonstrated the potential to moderate the levels of these compounds when in excess (Table 4). Conversely, the L. plantarum-aided fermentation technique may facilitate the biosynthesis of resveratrol, epicatechin, saponin White, kaempferol, an unidentified phenolic compound (U-PHE1), and two unidentified flavonoid compounds (UD-Flavo1 and UD-Flavo4). It is worthy of note that an increase in concentration of these phytochemicals may also be attributed to the release of cell-wall-bound phytochemicals (DeBenedictis et al., 2023; Sun et al., 2025). Fermented leaves of C. pentandra were notably rich in ginsenoside (248.47–16.29 mg/100 mg), catechin (129.459 ± 42.221 mg/ 100 mg), kaempferol (112.745 ± 12.753 mg/ 100 mg), rutin (63.263 ± 38.011 mg/ 100 mg), stevioside (17.97–163.93 mg/ 100 mg), saponin white (9.57–124.82 mg/ 100 mg), caffeic acid (48.909 ± 4.065 mg/ 100 mg), and resveratrol (9.037 ± 2.803 mg/ 100 mg) in decreasing order among the sixteen bioactives detected (M'Be et al., 2023). These compounds are health-beneficial bioactive compounds associated with anticancer, anti-tumor, anti-inflammatory, and other health activities (Papoutsis et al., 2010; Pasinetti et al., 2011; Juan et al., 2012; Vangalapati et al., 2016; Varoni et al., 2016; Nallathamby et al., 2017). The leaves of C. pentandra contain notable levels of resveratrol, a compound commonly found in red wine and implicated in the longevity of Asian communities. The amount of resveratrol in these leaves is comparable to that level (0.1–14 mg/L) reported in red wine (Gresele et al., 2008).

In conclusion, the findings indicate that fermentation may increase antioxidant properties and enhance the nutritional value of C. pentandra leaves. The concomitant rise in the activity of lactate dehydrogenase (LDH) indicates the active role of anaerobic metabolism in the specified improvement. These findings probably show that LDH plays a role in metabolic processes. Consequently, fermentation of C. pentandra leaves is a potential solution to enrich consumers nutritionally and medicinally, which may be further improved through fermentation. Regular consumption of fermented products from C. pentandra leaves is recommended to enhance antioxidant properties and overall well-being. Therefore, more efforts are needed to integrate fermented C. pentandra leaves into traditional diets and local food practices. Further research is recommended to assess the bioaccessibility of C. pentandra leaves using an in vivo model, despite the leaves having been consumed for centuries in certain parts of the world. The C. pentandra leaves were transported over several miles from the harvesting sites to the study center, which may be a limitation. Further effort is recommended to locate the leaves in areas closer to the study center to avoid stress that could influence the outcome of this study. Additionally, further validation of the phytochemical analysis results may be needed using a more sensitive HPLC system.

References

• 1

  AbeyrathneE.NamK.HuangX.AhnD. U. (2022). Plant- and animal-based antioxidants' structure, efficacy, mechanisms, and applications: a review. Antioxidants11:1025. doi: 10.3390/antiox11051025,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 2

  AboulthanaW. M.El-FekyA. M.OmarN. I.El-SayedA. E. B. (2026). The influence of algal biostimulator and nitrogen source on the phytochemical composition and biological properties of Corchorus olitorius leaves and stems. Sci. Rep.16:8948. doi: 10.1038/s41598-026-39774-z,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 3

  AfolabiI. S.AdigunA. J.GarubaP. A.AhuekweE. F.OdutayoO. E.AdeyemiA. O. (2023a). Enterococcus faecalis-aided fermentation to facilitate edible properties and bioactive transformation of underutilized Cyathea dregei leaves. Fermentation9:707. doi: 10.3390/fermentation9080707

  • CrossRef
  • Google Scholar

• 4

  AfolabiI. S.AhuekweE. F.GarubaP. A.AdigunA. J.OdutayoO. E.AdeyemiA. O. (2023b). Enterococcus faecalis-induced biochemical transformation during fermentation of underutilized Solenostemon monostachyus leaves. Fermentation9:33. doi: 10.3390/fermentation9010033

  • CrossRef
  • Google Scholar

• 5

  AfolabiI. S.Bisi-AdeniyiT. D.AdedoyinT. R.RotimiS. O. (2015). Radiations and biodegradation techniques for detoxifying Carica papaya seed oil for effective dietary and industrial use. J. Food Sci. Technol.52, 6475–6483. doi: 10.1007/s13197-014-1698-7,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 6

  AfolabiI. S.NwachukwuI. C.EzeokeC. S.WokeR. C.AdegbiteO. A.OlawoleT. D.et al. (2018). Production of a new plant-based milk from Adenanthera pavonina seed and evaluation of its nutritional and health benefits. Front. Nutr.5:9. doi: 10.3389/fnut.2018.00009,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 7

  AfolabiI. S.OloyedeO. B. (2011). Biochemical response of sweet potato to bemul-wax coating combined with calcium chloride treatment during ambient storage. Afr. J. Biotechnol.10, 2724–2732. doi: 10.5897/AJB10.1587

  • CrossRef
  • Google Scholar

• 8

  AfolabiI. S.UgbabeA. L.NwachukwuK. A.EberechiC. E.OlaitanM. E.AhuekweE. F.et al. (2025). Bioactive interactions, physicochemical and health-promoting qualities of pro-milk extract and flour derived from processed seeds from Hura crepitans and Citrullus colocynthis. Heliyon11:e42937. doi: 10.1016/j.heliyon.2025.e42937

  • CrossRef
  • Google Scholar

• 9

  AOAC (2015). Official Methods of Analysis. J. George W. Latimer (eds.) 20th Edn Arlington: Association of Official Analytical Chemists (AOAC).

  • Google Scholar

• 10

  AsigbaaseM.AdusuD.MusahA. A.AnabaL.NsorC. A.AbugreS.et al. (2024). Ethnobotanical and ethnopharmacological survey of medicinal tree species used in the treatment of diseases by forest-fringe communities of southwestern Ghana. Heliyon10:e23645. doi: 10.1016/j.heliyon.2023.e23645,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 11

  BindaS.HillC.JohansenE.ObisD.PotB.SandersM. E.et al. (2020). Criteria to qualify microorganisms as "probiotic" in foods and dietary supplements. Front. Microbiol.11:1662. doi: 10.3389/fmicb.2020.01662,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 12

  BiswasA.DeyS.HuangS.DengY.BirhanieZ. M.ZhangJ.et al. (2022). A comprehensive review of C. Capsularis and C. olitorius: a source of nutrition, essential phytoconstituents and pharmacological activities. Antioxidants11:1358. doi: 10.3390/antiox11071358,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 13

  BiswasA.DeyS.LiD.LiuY.ZhangJ.HuangS.et al. (2020). Comparison of phytochemical profile, mineral content, and in vitro antioxidant activities of Corchorus capsularis and Corchorus olitorius leaf extracts from different populations. J. Food Qual.2020, 1–14. doi: 10.1155/2020/2931097

  • CrossRef
  • Google Scholar

• 14

  BonnotE.ClauxO.BoulkoutM.Rolland-SabatéA.Abert-VianM. (2025). Chemical and nutritional profile of Tunisian Corchorus olitorius leaves: a valuable, under-exploited leafy vegetable. Lwt232:118393. doi: 10.1016/j.lwt.2025.118393

  • CrossRef
  • Google Scholar

• 15

  ChoudharyS. B.SharmaH. K.KarmakarP. G.KumarA. A.SahaA. R.HazraP.et al. (1973). Nutritional profile of cultivated and wild jute (Corchorus) species. Aust. J. Crop. Sci.7, 1973–1982.

  • Google Scholar

• 16

  Cisneros-YupanquiM.LanteA.MihaylovaD.KrastanovA. I.RizziC. (2023). The alpha-amylase and alpha-glucosidase inhibition capacity of grape pomace: a review. Food Bioprocess Technol.16, 691–703. doi: 10.1007/s11947-022-02895-0,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 17

  DaniaO. E.DokunmuT. M.AdegboyeB. E.AdeyemiA. O.ChibuzorF. C.IwealaE. E. J. (2024). Pro-estrogenic and anti-inflammatory effects of Corchorus olitorius and Amaranthus hybridus leaves in DMBA-induced breast cancer. Phytomed. Plus4:100567. doi: 10.1016/j.phyplu.2024.100567

  • CrossRef
  • Google Scholar

• 18

  de Paulo FariasD.de AraujoF. F.Neri-NumaI. A.PastoreG. M. (2021). Antidiabetic potential of dietary polyphenols: a mechanistic review. Food Res. Int.145:110383. doi: 10.1016/j.foodres.2021.110383,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 19

  DeBenedictisJ. N.de KokT. M.van BredaS. G. (2023). Impact of processing method and storage time on phytochemical concentrations in an antioxidant-rich food mixture. Antioxidants12:1252. doi: 10.3390/antiox12061252,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 20

  DoronS.SnydmanD. R. (2015). Risk and safety of probiotics. Clin. Infect. Dis.60 Suppl 2, S129–S134. doi: 10.1093/cid/civ085,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 21

  EffiongM. E.UmeokwochiC. P.AfolabiI. S.ChineduS. N. (2023). Assessing the nutritional quality of Pleurotus ostreatus (oyster mushroom). Front. Nutr.10:1279208. doi: 10.3389/fnut.2023.1279208,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 22

  EffiongM. E.UmeokwochiC. P.AfolabiI. S.ChineduS. N. (2024). Comparative antioxidant activity and phytochemical content of five extracts of Pleurotus ostreatus (oyster mushroom). Sci. Rep.14:3794. doi: 10.1038/s41598-024-54201-x,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 23

  ErskineE.OzkanG.LuB.CapanogluE. (2023). Effects of fermentation process on the antioxidant capacity of fruit byproducts. ACS Omega8, 4543–4553. doi: 10.1021/acsomega.2c07602,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 24

  FangH.LiD.KangJ.JiangP.SunJ.ZhangD. (2018). Metabolic engineering of Escherichia coli for de novo biosynthesis of vitamin B(12). Nat. Commun.9:4917. doi: 10.1038/s41467-018-07412-6,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 25

  GalanS. (2025) World Population - Forecast about the Development 2024–2100 Statista. Available online at: https://www.statista.com/statistics/262618/forecast-about-the-development-of-the-world-population/ (Accessed October 19 2025).

  • Google Scholar

• 26

  GargP. (2021). HPLC estimation of flavanoid (quercetin) of leaves and stem extracts of Ocimum sanctum and Tinospora cordifolia. J. Phytopharmacol.10, 220–224. doi: 10.31254/phyto.2021.10401

  • CrossRef
  • Google Scholar

• 27

  GreseleP.PignatelliP.GuglielminiG.CarnevaleR.MezzasomaA. M.GhiselliA.et al. (2008). Resveratrol, at concentrations attainable with moderate wine consumption, stimulates human platelet nitric oxide production. J. Nutr.138, 1602–1608. doi: 10.1093/jn/138.9.1602,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 28

  HarleO.FalentinH.NiayJ.ValenceF.CourselaudC.ChuatV.et al. (2020). Diversity of the metabolic profiles of a broad range of lactic acid bacteria in soy juice fermentation. Food Microbiol.89:103410. doi: 10.1016/j.fm.2019.103410

  • CrossRef
  • Google Scholar

• 29

  HuangL.HuangL.JiangY.TianL.ChenJ.HuS.et al. (2026). Wild vegetables contribute to improving the diversity and sustainability of vegetable diets in China. Proc. Natl. Acad. Sci. USA123:e2524063123. doi: 10.1073/pnas.2524063123,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 30

  IlodibiaC. V.UgwuR. U.OkekeC. U.EzeabaraC. A.OkekeN. F.AkachukwuE. E.et al. (2013). Determination of proximate composition of various parts of two Dracaena specie. Int. J. Bot.10, 37–41. doi: 10.3923/ijb.2014.37.41

  • CrossRef
  • Google Scholar

• 31

  JagelavičiūtėJ.BašinskienėL.ČižeikienėD. (2025). Enhancing the nutritional value of foods through probiotics and dietary fiber from fruit and berry pomace. Fermentation11:481. doi: 10.3390/fermentation11080481

  • CrossRef
  • Google Scholar

• 32

  JemereE. M.SatheeshN.AwolS. J.TadesseE. E.KassaM. G.TeferiD. A. (2025). Influence of selected organic acid concentration on the quality and shelf life of teff injera. Discov. Food5:44. doi: 10.1007/s44187-025-00299-5

  • CrossRef
  • Google Scholar

• 33

  JinJ.den BestenH. M. W.RietjensI.Widjaja-van den EndeF. (2025). Chemical and microbiological hazards arising from new plant-based foods, including precision fermentation-produced food ingredients. Annu. Rev. Food Sci. Technol.16, 171–194. doi: 10.1146/annurev-food-111523-122059,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 34

  JuanM. E.AlfarasI.PlanasJ. M. (2012). Colorectal cancer chemoprevention by trans-resveratrol. Pharmacol. Res.65, 584–591. doi: 10.1016/j.phrs.2012.03.010,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 35

  JudgeA.DoddM. S. (2020). Metabolism. Essays Biochem.64, 607–647. doi: 10.1042/EBC20190041,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 36

  KeyvanE.AdesemoyeE.Champomier-VergesM. C.Chanseaume-BussiereE.MardonJ.Nikolovska NedelkoskaD.et al. (2025). Vitamins formed by microorganisms in fermented foods: effects on human vitamin status-a systematic narrative review. Front. Nutr.12:1653666. doi: 10.3389/fnut.2025.1653666,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 37

  KnezE.Kadac-CzapskaK.GrembeckaM. (2023). Effect of fermentation on the nutritional quality of the selected vegetables and legumes and their health effects. Life13:655. doi: 10.3390/life13030655,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 38

  KnorrD.AugustinM. A. (2023). Food systems at a watershed: unlocking the benefits of technology and ecosystem symbioses. Crit. Rev. Food Sci. Nutr.63, 5680–5697. doi: 10.1080/10408398.2021.2023092,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 39

  LiuZ.PanJ. (2017). A practical method for extending the biuret assay to protein determination of corn-based products. Food Chem.224, 289–293. doi: 10.1016/j.foodchem.2016.12.084,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 40

  MailaM. Y.TsekeP. E. (2024). Influence of blanching time on the phytochemical and nutritive value of cowpea (Vigna unguiculata L. Walp) leafy vegetable. Int. J. Food Sci.2024:9095035. doi: 10.1155/2024/9095035,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 41

  M'BeC. U.ScherJ.GaianiC.AmaniN. G.BurgainJ. (2023). Impact of processing and physicochemical parameter on Hibiscus sabdariffa calyxes biomolecules and antioxidant activity: from powder production to reconstitution. Foods12:2984. doi: 10.3390/foods12162984,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 42

  MillenaC. G.BalonzoA. R. R.RentoyJ. R.RuivivarS. S.BobilesS. C. (2023). Effect of fermentation stages on the nutritional and mineral bioavailability of cacao beans (Theobroma cacao L.). J. Food Compos. Anal.115:104886. doi: 10.1016/j.jfca.2022.104886

  • CrossRef
  • Google Scholar

• 43

  MiškecK.FrlinM.ŠolaI. (2025). Impact of different thermal processing techniques on the phytochemical composition, antioxidant capacity, and DNA-protective properties of broccoli. Appl. Sci.15:7469. doi: 10.3390/app15137469

  • CrossRef
  • Google Scholar

• 44

  NallathambyN.PhanC. W.SeowS. L.BaskaranA.LakshmananH.Abd MalekS. N.et al. (2017). A status review of the bioactive activities of tiger milk mushroom Lignosus rhinocerotis (Cooke) Ryvarden. Front. Pharmacol.8:998. doi: 10.3389/fphar.2017.00998,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 45

  NchangweC. C.KassimN.MakuleE. (2024). Effect of blanching conditions and solar drying on selected nutritional, anti-nutritional and bioactive components of formulated avocado seed powder. Afr. J. Food Agric. Nutr. Dev.24, 26608–26622. doi: 10.18697/ajfand.131.23985

  • CrossRef
  • Google Scholar

• 46

  NoroziM.RezaeiM.KazemifardM. (2021). Effect of different acid processing methodologies on the nutritional value and reduction of anti-nutrients in soybean meal. J. Food Process. Preserv.46:e16205. doi: 10.1111/jfpp.16205

  • CrossRef
  • Google Scholar

• 47

  NsabimanaS.IsmailT.LazarteC. E. (2024). Enhancing iron and zinc bioavailability in maize (Zea mays) through phytate reduction: the impact of fermentation alone and in combination with soaking and germination. Front. Nutr.11:1478155. doi: 10.3389/fnut.2024.1478155,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 48

  NwinyiO. C.HassanW. D. (2021). Production of fruit wine using a wild strain of Saccharomyces cerevisiae isolated from fresh palm wine for sustainable food security. IOP Conference Series: Earth and Environmental Science665:012073. doi: 10.1088/1755-1315/665/1/012073

  • CrossRef
  • Google Scholar

• 49

  OdutayoO. E.AdegboyeB. E.OmonigbehinE. A.OgunlanaO. O.AfolabiI. S. (2021a). Characterization of potential probiotics isolated from fermented under-utilized Chrysophyllum albidum Linn kernels using microbiological, biochemical and molecular techniques. J. Phys. Conf. Ser.2070:12036. doi: 10.1088/1742-6596/2070/1/012036

  • CrossRef
  • Google Scholar

• 50

  OdutayoO. E.AdegboyeB. E.OmonigbehinE. A.OlawoleT. D.OgunlanaO. O.AfolabiI. S. (2021b). Structural transformation and creativity induced by biological agents during fermentation of edible nuts from Terminalia catappa. Molecules26:5874. doi: 10.3390/molecules26195874,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 51

  OdutayoO. E.OmonigbehinA. E.OgunlanaO. O.AfolabiI. S. (2023). Biochemical effects of fermentation on selected phytochemicals, enzymes and antioxidant activities in the under-utilized seeds of Chrysophyllum albidum Linn and Terminalia catappa Linn. Trop. J. Nat. Prod. Res.7, 4945–4953. doi: 10.26538/tjnpr/v7i10.34

  • CrossRef
  • Google Scholar

• 52

  OdutayoO. E.OmonigbehinE. A.OlawoleT. D.OgunlanaO. O.AfolabiI. S. (2020). Fermentation enhanced biotransformation of compounds in the kernel of Chrysophyllum albidum. Molecules25:6021. doi: 10.3390/molecules25246021,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 53

  Osei-OwusuJ.KokroK. B.OforiA.ApauJ.DofuorA. K.VigbedorB. Y.et al. (2023). Evaluation of phytochemical, proximate, antioxidant, and anti-nutrient properties of Corchorus olitorius, Solanum macrocarpon and Amaranthus cruentus in Ghana. Int J Biochem Mol Biol14, 17–24.

  • Google Scholar

• 54

  OzturkT.Avila-GalvezM. A.MercierS.VallejoF.BredA.FraisseD.et al. (2024). Impact of lactic acid bacteria fermentation on (poly)phenolic profile and in vitro antioxidant and anti-inflammatory properties of herbal infusions. Antioxidants13:562. doi: 10.3390/antiox13050562,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 55

  PapoutsisA. J.LamoreS. D.WondrakG. T.SelminO. I.RomagnoloD. F. (2010). Resveratrol prevents epigenetic silencing of BRCA-1 by the aromatic hydrocarbon receptor in human breast cancer cells. J. Nutr.140, 1607–1614. doi: 10.3945/jn.110.123422,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 56

  PasinettiG. M.WangJ.MarambaudP.FerruzziM.GregorP.KnableL. A.et al. (2011). Neuroprotective and metabolic effects of resveratrol: therapeutic implications for Huntington's disease and other neurodegenerative disorders. Exp. Neurol.232, 1–6. doi: 10.1016/j.expneurol.2011.08.014,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 57

  PiechowiakT.SkoraB.BalawejderM. (2023). Effect of postharvest nicotinamide treatment on NAD(+) metabolism and redox status in strawberry fruit during storage. Phytochemistry213:113766. doi: 10.1016/j.phytochem.2023.113766,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 58

  RahmanM. S.EmonD. D.TomaM. A.NupurA. H.KarmokerP.IqbalA.et al. (2023). Recent advances in probiotication of fruit and vegetable juices. J. Adv. Vet. Anim. Res.10, 1–537. doi: 10.5455/javar.2023.j706,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 59

  RevueltaJ. L.BueyR. M.Ledesma-AmaroR.VandammeE. J. (2016). Microbial biotechnology for the synthesis of (pro)vitamins, biopigments and antioxidants: challenges and opportunities. Microb. Biotechnol.9, 564–567. doi: 10.1111/1751-7915.12379,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 60

  SakatS.JuvekarA. (2010). Comparative study of Erythrina indica lam. (Febaceae) leaves extracts for antioxidant activity. J. Young Pharm.2, 63–67. doi: 10.4103/0975-1483.62216,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 61

  SamtiyaM.AlukoR. E.PuniyaA. K.DhewaT. (2021). Enhancing micronutrients bioavailability through fermentation of plant-based foods: a concise review. Fermentation7:63. doi: 10.3390/fermentation7020063

  • CrossRef
  • Google Scholar

• 62

  SarfoA.AbotsiW. K. M.EkuadziE.WoodeE. (2022). Ceiba pentandra (L.) Gaertn hydroethanolic leaf extract exhibits anticonvulsant properties in mouse models. Phytomed. Plus2:100263. doi: 10.1016/j.phyplu.2022.100263

  • CrossRef
  • Google Scholar

• 63

  SaritaB.SamadhanD.HassanM. Z.KovalevaE. G. (2025). A comprehensive review of probiotics and human health-current prospective and applications. Front. Microbiol.15:1487641. doi: 10.3389/fmicb.2024.1487641,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 64

  SaritasS.PortocarreroA. C. M.Miranda LopezJ. M.LombardoM.KochW.RaposoA.et al. (2024). The impact of fermentation on the antioxidant activity of food products. Molecules29:3941. doi: 10.3390/molecules29163941,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 65

  SchlesierK.HarwatM.BohmV.BitschR. (2002). Assessment of antioxidant activity by using different in vitro methods. Free Radic. Res.36, 177–187. doi: 10.1080/10715760290006411,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 66

  SharmaD. C.ShuklaR.AliJ.SharmaS.BajpaiP.PathakN. (2016). Phytochemical evaluation, antioxidant assay, antibacterial activity and determination of cell viability (J774 and THP1 alpha cell lines) of P. sylvestris leaf crude and methanol purified fractions. EXCLI J.15, 85–94. doi: 10.17179/excli2015-689,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 67

  Shumye GebreT.Admassu EmireS.Okomo AlooS.ChelliahR.VijayalakshmiS.Hwan OhD. (2024). Unveiling the potential of African fermented cereal-based beverages: probiotics, functional drinks, health benefits and bioactive components. Food Res. Int.191:114656. doi: 10.1016/j.foodres.2024.114656,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 68

  SlavinJ. L.LloydB. (2012). Health benefits of fruits and vegetables. Adv. Nutr.3, 506–516. doi: 10.3945/an.112.002154,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 69

  SubhashreeS.KavitaM. S. (2019). A study on suitable non dairy food matrix for probiotic bacteria - a systematic review. Curr. Res. Nutr. Food Sci. J.7, 05–16. doi: 10.12944/crnfsj.7.1.02

  • CrossRef
  • Google Scholar

• 70

  SunL.KwakS.JinY. S. (2019). Vitamin a production by engineered Saccharomyces cerevisiae from xylose via two-phase in situ extraction. ACS Synth. Biol.8, 2131–2140. doi: 10.1021/acssynbio.9b00217,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 71

  SunS.YuY.JoY.HanJ. H.XueY.ChoM.et al. (2025). Impact of extraction techniques on phytochemical composition and bioactivity of natural product mixtures. Front. Pharmacol.16:1615338. doi: 10.3389/fphar.2025.1615338,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 72

  TareauM.-A.GreeneA.OdonneG.DavyD. (2022). Ceiba pentandra (Malvaceae) and associated species: spiritual keystone species of the neotropics. Botany100, 127–140. doi: 10.1139/cjb-2021-0099

  • CrossRef
  • Google Scholar

• 73

  TeL.SinA.JA.EtT.FsA. (2020). Phytochemical properties of Ceiba petandra (kapok tree), Moringa oleifera (Moringa) and Cymbopogon citratus (lemon grass) collected from a home garden in Igbor, Gwer east, Benue state, Nigeria. Int. J. Complement. Alt. Med.13, 62–67. doi: 10.15406/ijcam.2020.13.00494

  • CrossRef
  • Google Scholar

• 74

  TengT. S.ChinY. L.ChaiK. F.ChenW. N. (2021). Fermentation for future food systems: precision fermentation can complement the scope and applications of traditional fermentation. EMBO Rep.22:e52680. doi: 10.15252/embr.202152680,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 75

  TosocJ. P. S.NunezaO. M.SudhaT.DarwishN. H. E.MousaS. A. (2021). Anticancer effects of the Corchorus olitorius aqueous extract and its bioactive compounds on human cancer cell lines. Molecules26:6033. doi: 10.3390/molecules26196033,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 76

  UN-DESA (2017). World Population Projected to reach 9.8 Billion in 2050, and 11.2 Billion in 2100. Department of Economic and Social Affairs of the United Nations. New York, NY: Department of Economic and Social Affairs of the United Nations (UN-DESA).

  • Google Scholar

• 77

  VangalapatiM.NissyS. M.PadmajaH. (2016). Fermentation and antioxidant activity of catechins from Hibiscus sabdariffa. IJSART2, 576–578.

  • Google Scholar

• 78

  VaroniE. M.Lo FaroA. F.Sharifi-RadJ.IritiM. (2016). Anticancer molecular mechanisms of resveratrol. Front. Nutr.3:8. doi: 10.3389/fnut.2016.00008,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 79

  WackettL. P. (2018). Microbial acid fermentation products: an annotated selection of world wide web sites relevant to the topics in microbial biotechnology. Microb. Biotechnol.11, 268–269. doi: 10.1111/1751-7915.13044,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 80

  WickramasingheY. W. H.WickramasingheI.WijesekaraI. (2020). Effect of steam blanching, dehydration temperature & time, on the sensory and nutritional properties of a herbal tea developed from Moringa oleifera leaves. Int. J. Food Sci.2020:5376280. doi: 10.1155/2020/5376280,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 81

  ZhaoY. S.EweysA. S.ZhangJ. Y.ZhuY.BaiJ.DarweshO. M.et al. (2021). Fermentation affects the antioxidant activity of plant-based food material through the release and production of bioactive components. Antioxidants10:2004. doi: 10.3390/antiox10122004,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 82

  ZuntarI.PetricZ.Bursac KovacevicD.PutnikP. (2020). Safety of probiotics: functional fruit beverages and nutraceuticals. Foods9:947. doi: 10.3390/foods9070947,

  • Pubmed Abstract
  • CrossRef
  • Google Scholar