Photo-physical characterizations and evaluation of in-vitro antioxidant, anti-inflammatory and antidiabetic potentials of green synthesized ackee (Blighia sapida) selenium nano-particles

Abstract

Background

Green synthesized nanoparticles have recently gained significant medicinal applications and oftentimes outperform their green sources. Selenium is of fundamental importance to human health, stemming from its distinctive physicochemical properties, such as antioxidant activity, inhibition of Lipid peroxidation, stabilization of membrane proteins, maintenance of membrane fluidity and modulation of cell signaling. Though reports have shown some therapeutic potential of Ackee plant parts such as antioxidant, anti-inflammatory, antimicrobial, neuroprotective, very few scientific proofs still exist in support of these effects.

Methods

This study synthesized selenium nanoparticles (Se-NPs) from crude methanolic extracts of Ackee leaves (AKL) and Ackee arils (AKA), examined the photo-physical characteristics of the Se-NPs and determined the in-vitro antioxidant, antidiabetic, and anti-inflammatory potentials of AKL, AKA, and their Se-NPs using established protocols.

Results

In both leaves and arils Se-NPs: UV spectroscopy revealed a qualitative absorbance at 310 nm; FTIR indicated multiple vibrations around 4000 cm⁻¹- 400 cm⁻¹; SEM images of 5 µm principally showed consistent size distribution of amorphous and granular shape at a magnification of 10,000X; while EDS spectra strongly confirm the presence of atomic Se compound at 30 kV. Various antioxidant activities assays carried out showed a range of approximately 4 to 60 times higher activities of the AKL, AKA, and Se-NPs than Ascorbic acid—the standard drug used. Furthermore, appreciable activities of more than 50% were obtained for alpha-amylase and alpha-glucosidase inhibitory activities, along with highly significant activities of haemoglobin glycosylation, glucose uptake, membrane stabilization, anti-arthritic, anti-haemolysis activities, when AKL, AKA, and Se-NPs were compared with standard drugs.

Conclusion

Encouraging the development and utilization of AKL, AKA, and Se-NPs will provide tremendous therapeutic efficacy and bioavailability approaches towards the management of diabetes, inflammation, and other oxidative stress-related diseases.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12906-024-04694-w.

Background

Advancements in the field of nanotechnology and its applications to medicines, pharmaceuticals and industry have revolutionized the twentieth century. Nanotechnology is the study of extremely small structures, they are minuscule structures typically in the range of 1 to 100 nm [1, 2]. These infinitesimal entities, with their unique physicochemical properties, have opened doors to a myriad of groundbreaking applications in medicine (such as: targeted drug delivery; tissue engineering and regenerative medicine; cancer treatment and diagnosis; implantable devices—pacemakers, prosthetics), in food and agriculture (food safety and quality control; precision agriculture and crop monitoring; nano-scale fertilizers and pesticides), in biotechnology (synthetic biology and bioengineering; gene editing and gene therapy; biomimetic materials and systems) and environment (air pollution control and monitoring; water purification and treatment; soil remediation and conservation) [2–5]. Recent research in the field of nanomedicine and nanotechnology has unveiled remarkable advancements with profound implications for human health and the environment [3, 4]. Thus, nanotechnology is the treatment of individual atoms, molecules, or compounds into structures to produce materials and devices with special properties [2, 5]. Nanoparticles, encompassing a diverse array of materials such as metals, metal oxides, polymers, and lipids, are engineered to serve as versatile carriers, delivery vehicles, and diagnostic tools. They hold the potential to revolutionize drug delivery, enhance imaging techniques, and facilitate targeted therapies, ushering in an era of precision medicine [6, 7].

In an era increasingly characterized by sustainability and eco-consciousness, the concept of green synthesis has gained significant traction [8, 9]. Green synthesis involves the fabrication of nanoparticles using environmentally friendly processes and natural resources [10, 11]. Presently green synthesis has shown to offers a more versatile, eco-friendly, biocompatible, sustainable and effective approaches, which have led to the discovery of novel nanoparticles that have higher stability, improved properties, better shelf life and reduced environmental impact, when compared to physical or chemical synthesis [12, 13]. It has provided an excellent option for various medical, pharmaceutical, industrial and materials science leading to innovative applications [14].

One fascinating facet of this approach involves harnessing microelements such as zinc (Zn), manganese (Mn), and selenium (Se) to create nanoparticles with enhanced biocompatibility [9, 15]. Among these microelements, selenium dioxide (SeO₂) nanoparticles have garnered particular attention for their potential therapeutic applications in human, stemming from their distinctive physicochemical properties. Selenium exhibits its biological function by acting as an antioxidant and catalyst in the production of active thyroid hormone and is needed for the proper functioning of the immune system [8, 16]. Selenium also plays vital role as a key micronutrient in mitigating the development of virulence and inhibiting HIV progression to AIDS. It is also highly required for sperm motility and may reduce the risk of miscarriage [17]. Selenium deficiencies have been equivocally linked to adverse mood states and associated with lots of oxidative stress-related diseases like diabetes, inflammation, and cardiovascular diseases [16]. In addition, an elevated selenium intake is presumed to be associated with reduced cancer risk, and ongoing research has been geared towards establishing that low or diminishing selenium status in some parts of the world (notably in some European countries) may be associated with this health concern [16]. Concurrently, the paradigm of green synthesis has risen to prominence as a suitable approach to nanoparticle fabrication, harnessing the inherent therapeutic capabilities of natural resources from plants [18, 19].

The use of plants in traditional medicine has generated a lot of attention and anxiety due to their potency and safety margin. Nevertheless, local herbs are still widely in use in their various forms: such as the roots, stems, leaves and seeds, around the world. This is mostly related to the presence of several biologically active chemical compounds with diverse beneficial and favourable therapeutic effects in some disease conditions (e.g. diabetes, hypertension, inflammation, etc.) and generally, for optimal good health [20]. Currently, many drugs used in modern medicine applications are sourced from whole plant or their extracts. For several decades, plant extracts were employed for traditional medicinal motives, basically, for therapy in indigenous medicine and herbage, however, they are at present widely used in modern medicine [21].

Ackee (Blighia sapida) is a popularly known indigenous tree crop of West Africa and Southern America, as these environments are naturally made up of subtropical and tropical vegetation [22, 23]. Various parts of the Ackee plant, that is, leaves, bark, roots, arils, and seeds, are used in traditional medicine for the treatment of fever, malaria, internal haemorrhage, dysentery, yellow fever, inflammation, and diabetes. Its leaves and arils, rich in phytochemical diversity, have long been harnessed for their therapeutic properties, including antimicrobial and antioxidant effects [24]. Ackee leaves contain a diverse range of bioactive phytochemicals, including flavonoids, alkaloids, and polyphenols [25, 26] Ackee arils are yellowish to cream-coloured fleshy, edible part of the ackee fruit, they are rich in essential nutrients, vitamins, and antioxidants, making them valuable in traditional medicine and culinary applications. Ripe arils are often eaten fresh, fried, dried, roasted, or made into soup in some parts of West Africa [27]. Therefore, the diverse phytochemicals found in Ackee leaves and arils offer promise for novel therapeutic interventions. In our previous studies, the presence of numerous essential phytochemicals at appreciable levels in Ackee leaves and arils have been well established [24]. Furthermore, green synthesized nanoparticles have also gained tremendous medicinal application and they have been shown to outperform their green sources. However, the precise intrinsic biological potentials of Ackee leaves and arils, and their green-synthesized selenium nanoparticles (Se-NPs) remain to be elucidated.

Though, selenium may have been attributed to have narrow toxicity margins, Se-NPs particles possess the opposite, by having wide therapeutic window and reduced toxicity, as mortalities caused by toxicity associated with Se were minimized by the use of Se-NPs up to four times in rodent models [28, 29]. Furthermore, in pathological conditions such as diabetes, cancer, inflammatory disorders, Se-NPs have showed more bioavailability and biological activity when compared to Se compounds [28, 30]. Antioxidant effects of SeNPs when compared with Se-methyl, selenocysteine and selenomethionine and selenite, were reported to increase the activity of selenoenzymes such as Glutathione Peroxidase family (GPXs) and Thioredoxin Reductase (TR), with equal efficacy and less toxicity [29, 31].

In the context of green synthesis, Ackee's contribution to the fabrication of Se-NPs thus represents a convergence of traditional wisdom and cutting-edge nanotechnology [26]. This research therefore embarks on an in-vitro comprehensive assessment of the intrinsic biological potential of Se-NPs synthesized using Ackee-derived methanolic extracts. This includes characterizing their physicochemical attributes, elucidating their impact on a spectrum of cellular metabolic activities, and unveiling their potential therapeutic applications. These encompass glucose uptake modulation, anti-glycation properties, membrane stabilization, anti-arthritic effects, anti-haemolysis potential, α-amylase inhibition, and α-glucosidase inhibition. The holistic approach undertaken in this study aims to provide insights into the adaptability and versatility of these entities in diverse health contexts, ultimately contributing to the advancement of nanomedicine and sustainable nanotechnological practices.

To comprehensively assess the biological potential and effects of green-synthesized selenium nanoparticles (Se-NPs) using Ackee leaves and arils, comparative analyses were conducted. Ackee plant leaves and arils, known for their diverse phytochemical compositions, serve as vital control groups in the experiments. Additionally, pharmaceutical drugs with established effects, including Gentamicin (GT), Aspirin (Asp), Metformin (Met), Diclofenac (Dic), Butylated hydroxyanisole (BHA), and Acarbose, (Acb) were incorporated as reference substances. These drugs, each chosen for their specific pharmacological properties, offer crucial reference points for evaluating the impact of green-synthesized Se-NPs on various cellular activities and metabolic processes. By contrasting the Se-NPs with the well-characterized drug references and the Ackee plant extracts, this study provides a comprehensive understanding of the potential therapeutic applications and mechanisms of these nanomaterials, thereby contributing to the advancement of biomedical research. It is anticipated that our findings will pivot further research in elucidating other significant physiological purposes these Se-NPs could offer, when compared with other established pharmaceuticals.

Materials and methods

Chemicals and reagents

Methanol, diethyl ether, sodium selenite, zinc acetate dehydrate, sodium hydroxide pellet, potassium permanganate, sodium hydrogen phosphate, sodium dihydrogen phosphate, sodium chloride, potassium hydrogen phosphate, potassium ferrycyanide, dextrose, sodium citrate, citric acid were acquired from Sigma Aldrich Chemie GmBH and Sigma-Aldrich Co. All other reagents used were of analytical grade and glass distilled water was used.

Collection of O⁻ hemoglobin blood group and research ethical approval

Whole cell hemoglobin O⁻ blood group employed in some of the biological in-vitro assays in this research work, were previously collected from healthy volunteers deposited at Federal University Oye Ekiti, University Clinic Blood Bank. The consent for use has been given by the donors for clinical and experimental purposes and ethical approval was given by our University Research Committee for its collection and use. All the experimental methods used were approved by the Research Ethics Committee for animal experimentation of the Faculty of Science, FUOYE with approval number: FUOYEFSC 201122 – REC2024/015. Our work does not require any direct patient trial; hence there is no Clinical Trial Number for this research as in-vitro assays were only conducted throughout the course of this research work.

Sample collection and preparation

Mature fresh leaves and arils of Ackee (Blighia sapida) were collected from a local farm plantation in Idofin area of Oye- Ekiti (Latitude 7° 53′ 21.91'' N and Longitude 5° 20′ 41.35'' E). The plant samples were authenticated at the publicly available herbarium at Forestry Research Institute of Nigeria, Forestry Hill, Jericho, Private Mail Bag 5054, Ibadan Oyo State, Nigeria by Adeyemo A. and Egunjobi A.J. with authentication / deposition number FHI 114046. The leaves and arils were thoroughly washed under a running tap and weighed.

Methanolic extraction of ackee leaves and arils

2.8 kg of fresh Ackee leaves was weighed in a beaker using a high precision scale; it was then smoothly blended in a Kenwood Food Processor (MultiPro Compact + FDM313SS), with 6 L of absolute methanol to give a smooth paste. For the extraction of Ackee arils, 2 L of diethyl ether was initially added to 17 kg of Ackee arils in a Kenwood food processor and blended to fine paste. The paste was then transferred into a muslin cloth and content therein pressed out to facilitate removal of the fat contents present in the arils. To the defatted arils, 2 L of methanol was then added and blended again in a Kenwood food processor. This was then carefully transferred into a muslin cloth and the content was subsequently pressed out in order to get the extract. The extracts were then separately clarified using a table top BIOBASED High Speed centrifuge (BKC-TH16R BKC-TH20R) at a speed of 4000 rpm, the clear supernatants were then reduced to 1/3 of its original volume using a BIOBASED Rotary Evaporator (RE 100-Pro) at 50 °C. The concentrated extracts were then freeze-dried to powder using a BIOBASED Freeze Drier (BK FD18PT). The dried powders were kept in airtight containers and used for subsequent analyses.

Synthesis of selenium nanoparticles from ackee leaves and arils methanolic extracts

12 g of leaves and arils extracts were separately resolubilized in 300 mL of distilled water, with each final concentration of 40 mg/mL. These solutions were clarified by centrifuging for 3 min at 14,000 rpm. Subsequently, the supernatants obtained were then stored in a conical flask for further use. The Se-NPs were synthesized using the clarified Ackee (Blighia sapida) methanolic leaves and arils extracts. Briefly, a 10 mM selenium solution was prepared by adding 0.263 g of sodium selenite (Na₂SeO₃) in 100 mL of distilled water. This was added separately to 25 mL of leaves and arils methanolic extracts in a Hisense Microwave Oven (H20MOMBS4HGUK) at 800 Watts fixed power and 4 min exposure time. The leaves and arils synthesized Se-Nps (Se-NP_L and Se-NP_A) were then stored in 50 mL Falcon tubes, and these were used for further analysis [32, 33].

Photo-physical characterization of synthesized selenium nanoparticles

UV–visible spectroscopy analysis

The optical transmission/absorption spectra of Se-NPs dispersed in water were recorded using a BIOBASED UV–VIS spectrophotometer. Distilled water was used as a blank in the measurement. 3 mL of synthesized Se-NPs were placed in a quartz cuvette with a 1 cm path length and inserted in a UV–Visible spectrophotometer in the wavelength range of 190 nm to 360 nm [34].

Fourier transform infrared (FTIR) spectroscopy analysis

The chemical composition of the synthesized Se-NPs was studied by using a PerkinElmer Spectrum™ 3 FT-IR spectrometer (L1280138). To characterize biomolecules that were attached to Se-NPs after the green synthesis process, FTIR spectra of purified Se-NPs aqueous solution was acquired. The spectra of each sample were collected at a resolution of 4 cm⁻¹ and 64 interferogram scans in the range of 400–4000 cm⁻¹. Several modes of vibrations were recognized and assigned in order to identify various functional groups that are present in the purified Se-NPs solution [35].

Scanning electron microscopy (SEM) analysis

SEM was carried out to know the shape, surface morphology and size of the Se-NPs using Zeiss SEM (Detector VPSE G4, Variable Pressure). Briefly, Se-NPs suspension was air dried before loading them to sample holders. Later, Se-NPs were coated with gold using a sputter coater in a vacuum and thereafter, images were taken at 30 kV using different magnifications [36].

Energy-dispersive X-ray spectroscopy analysis (EDS)

EDS was carried out to analyze the elemental chemical composition of synthesized Se-NPs. The Se-NPs were dissolved in absolute ethanol and one drop of the suspension was placed on a sample loading grid, evenly dried and elemental/atomic analysis was performed [37].

Antioxidant assays

The antioxidant potentials of Ackee leaves (AKL) and arils (AKA) and their Se-NPs (Se-NPL and Se-NPA), were measure following their mechanism of action as they relate to: Total Antioxidant Capacity (TAC); Free Radical Scavenging (FRS); and Electron Transfer (ET) assays.

Total antioxidant capacity (TAC) assay

TAC measures the overall antioxidant capacity of sample. TAC was measured in a mixture containing 0.6 mL of sample and appropriate volumes of 0.6 M sulfuric acid, 28 mM sodium phosphate, and 4 mM ammonium molybdate. This mixture was incubated at 95 °C for 90 min. After cooling to room temperature, absorbance of the solution (A_S) was recorded at 695 nm against a reagent blank. As a reference, the total antioxidant capacity of Ascorbic acid (A_C) was also determined [38]. TAC was subsequently calculated and expressed as % activity as follows:

$$\%\text{TAC activity}=({\mathrm{A}}_{\mathrm{S}}/{\text{A}}_{\text{C}})\text{X}100$$ 1

Radical Scavenging assays

These assays measure the ability of antioxidants to scavenge and neutralize free radicals. Assays measured include:

• (i)Hydroxyl radical (OH) scavenging ability
  A mixture was prepared by combining 0.25 mL of extract with 2.25 mL of sodium phosphate buffer (150 mM, pH 7.4), and 0.25 mL of FeSO4–EDTA (10 mM). Subsequently, 0.25 mL of H2O2 was added to the mixture. This was incubated for 30 min and absorbance of the solution (AS) was determined at 520 nm. Methanol was employed as the blank (AO). As a reference standard, ascorbic acid was used [39]. The OH scavenging activity was subsequently calculated and expressed as:

  $$\%\mathrm{OH}\mathrm{scavenging}\mathrm{activity}=[({\mathrm{A}}_{\mathrm{O}}-{\mathrm{A}}_{\mathrm{S}})/{\mathrm{A}}_{\mathrm{O}}]\times 100$$ 2
• (ii)Superoxide anion (SA) scavenging ability
  This assay consisted of a mixture of 0.5 mL of extract and 2.25 mL of Tris–HCl buffer (50 mmol/L, pH 8.2). Incubation of mixture was done at 25 °C for 20 min, subsequently; 0.2 mL of 25 mmol/L pre-heated pyrogallol at 25 °C was introduced into the mixture. The reaction was allowed to proceed for 4 min and was terminated by adding 0.05 mL of 8 mol/L HCl solution. The resulting mixture was then subjected to centrifugation at 1000 rpm for 15 min, and absorbance of the samples (AS) and control (AO) solutions were individually taken at 325 nm [40]. The SA scavenging activity was subsequently calculated and expressed as:

  $$\%\mathrm{SA}\mathrm{scavenging}\mathrm{activity}=\{[{\mathrm{A}}_{\mathrm{O}}-({\mathrm{A}}_{\mathrm{S}}-{\mathrm{A}}_{\mathrm{O}})]/{\mathrm{A}}_{\mathrm{O}}\}\times 100$$ 3
• (iii)1, 1-diphenyl–2 picrylhydrazyl (DPPH) scavenging ability
  The ability of the extracts to decolorize DPPH radical was accessed using the method described by Moniruzzaman et al. [41]. The reaction mixture consisted of 2.9 mL of extract and 0.1 mL of 0.1 mmol/L DPPH. Subsequently, the different solutions were incubated in darkness at 25 °C for 30 min, after which, the absorbance of the solutions were measured at 517 nm. The absorbance of extract samples (AS), blank (only methanol; AB) and of only DPPH (AC). L-ascorbic acid was used as a standard [41]. The DPPH scavenging activity was subsequently calculated and expressed as:

  $$\%\mathrm{DPPH}\mathrm{scavenging}\mathrm{activity}=\{1-[({\mathrm{A}}_{\mathrm{S}}-{\mathrm{A}}_{\mathrm{B}})/({\mathrm{A}}_{\mathrm{C}}-{\mathrm{A}}_{\mathrm{B}})]\}\times 100$$ 4
• (iv)Nitric oxide (NO) scavenging ability

  This was determined in a mixture containing equal volumes of extract and 0.025 M sodium nitroprusside (SNP). This was subjected to 5 h incubation at 29 °C, 0.5 mL of the mixture was extracted after the incubation period and diluted with an equal volume of Greiss reagent, which comprises of 1% sulphanilamide, 2% phosphoric acid, and 0.1% naphthylene diamine dihydrochloride dissolved in water. The absorbance of the resulting solution was measured at 546 nm. The absorbance of extract sample (AS), while the absorbance of control that contains no extract (AC). L-ascorbic acid was used as a standard [42]. The NO scavenging activity was subsequently calculated and expressed as: % NO Scavenging activity following in (Eq. 1).

Electron transfer assays

These assays measure the ability of antioxidants to transfer electrons to reduce oxidized compounds. Assays measured include:

• (i)Ferric reducing antioxidant power (FRAP)

  The reaction mixture consists of 125 μL extract, 1.25 mL of 200 mM sodium phosphate buffer and 1.25 mL of 1% potassium ferricyanide. The resulting solution was incubated at 50 °C for duration of 20 min. Afterwards, 1.25 mL of 10% trichloroacetic acid was incorporated, and the solution was centrifuged at 650 g for 10 min. The supernatant was carefully collected and subsequently, an equal volume of double-distilled water and 1 mL of 0.1% ferric chloride was added. Absorbance of the sample (AS) solution was recorded at 700 nm against a blank and L-ascorbic acid (AC) served as the standard for reference [43, 44]. The FRAP activity was subsequently calculated and expressed as % FRAP activity following in (Eq. 1).

• (ii)2, 2-azinobis-3-ethylbenzo-thiazoline-6-sulfonate (ABTS) scavenging ability

  A mixture was prepared by combining 7 mM ABTS with an equal volume of 2.45 mM K2SO2. This mixture was allowed to stand in the dark for 12–16 h at room temperature. The ABTS solution was diluted with saline phosphate-buffer (pH 7.4) until an absorbance of approximately ± 0.01 was achieved at 734 nm. Subsequently, 3 mL of the ABTS solution was separately introduced to 20 µL of a standard (ascorbic acid) and each sample. The resulting mixture was then incubated in darkness for a duration of 2 h and absorbance of solutions of ascorbic acid (AO) and sample (AS) were recorded at 734 nm against a blank [45, 46]. The ABTS scavenging activity was subsequently calculated following in (Eq. 2).

Intrinsic biological assays (Anti-inflammatory and Diabetes biomarkers)

Anti-inflammatory biomarkers

• (i)Membrane stabilization
  Fresh whole human O− blood group collected from healthy volunteers deposited at Federal University Oye Ekiti, Clinic Blood Bank, with ethical approval by the University Research Committee, was obtained and mixed with an equal volume of sterilized Alsever solution containing 2% dextrose, 0.8% sodium citrate, 0.05% citric acid, and 0.42% sodium chloride in water. The mixture was centrifuged at 3000 rpm for 10 min, and packed cells were obtained. The blood packed cells were then reconstituted to form a 10% v/v suspension with isosaline. Subsequently, 1 mL of blood cell suspension was added to equal volume of extracts and standard drug at various concentrations (100 µg, 200 µg and 500 µg). The assay mixture was incubated at 37 °C for 30 min and later centrifuged. The absorbance of the samples (AS) and control (has no extract; AO) solutions were individually measured at 560 nm. Aspirin was used as standard [47]. The percent of membrane stabilization was subsequently calculated and expressed as:

  $$\%\mathrm{Membrane}\mathrm{stabilization}=[100-({\mathrm{A}}_{\mathrm{S}}/{\mathrm{A}}_{\mathrm{O}})]\times 100$$ 5
• (ii)Anti-arthritic activity
  The reaction mixture consisted of 0.2 mL egg albumin, 2.8 mL of saline phosphate-buffer (0.1 M, pH 6.4), and 2 mL extract at various concentrations (100 µg, 200 µg and 500 µg). The mixture was pre-incubated at 37 °C for 15 min and subsequently incubated at 70 °C for 5 min. After cooling, the absorbance of the samples (AS) and control (has no extract; AO) solutions were individually quantified at 660 nm using a UV–Visible Spectrophotometer (model 1720, Shanghai yoke instrument Co., Ltd). Diclofenac sodium was employed as a reference drug [48]. The percent of inhibition of protein denaturation that translates to the anti-arthritic activities of the extracts were subsequently calculated and expressed as:

  $$\%\mathrm{Anti}-\mathrm{arthritic}\mathrm{activity}=[({\mathrm{A}}_{\mathrm{S}}/{\mathrm{A}}_{\mathrm{O}})-1]\times 100$$ 6
• (iii)Anti-Haemolysis activity
  Human blood samples (O−) were meticulously collected from healthy volunteers and subsequently aliquoted into EDTA (ethylene diamine tetra acetic acid) bottles. To isolate the red blood cells (RBCs) from the sample, the blood samples were subjected to centrifugation at 3000 rpm for 10 min, effectively removing the plasma, platelets, and buffy coat. The resulting RBCs were then subjected to two washes with cold PBS (0.1 M, pH 7.4), following which was utilized for the hemolysis inhibition assay, conducted in accordance with the protocol outlined by McCaughey et al., [49]. The assay mixture consisted of 2 mL of the RBC suspension, various concentrations of extracts (100 µg, 200 µg and 500 µg), 0.1 mL H2O2 and 1 mL methanol. The mixture was incubated at room temperature with continuous shaking for 2 h 30 min on a Stuart orbital shaker (SSL1) operating at 100 rpm. This was followed by centrifugation at 2000 rpm for 10 min. The absorbance of supernatant of the samples (AS) and control (has no extract; AO) solutions were individually measured at 540 nm. Butylated hydroxyanisole (BHA) was used as a reference standard, being a high antioxidant compound with great potential to prevent hemolysis [49]. The percent of anti-haemolysis activities of the extracts were subsequently calculated and expressed as:

  $$\%\mathrm{Anti}-\mathrm{haemolysis}\mathrm{activity}=({\mathrm{A}}_{\mathrm{S}}/{\mathrm{A}}_{\mathrm{O}})\times 100$$ 7
• (iv)Haemoglobin Glycosylation
  The assay mixture consisted of varied concentrations of extract (100 µg, 200 µg and 500 µg), 1 mL of glucose (2% in phosphate buffer—0.01 M, pH 6.9) and 1 mL of haemoglobin (0.6% in phosphate buffer—0.01 M, pH 6.9). The resulting reaction mixture was incubated under dark conditions at room temperature for 72 h, and absorbance of the samples (AS) and control (has no extract; AO) solutions were individually measured at 520 nm. Gentamicin was used as standard [50]. The percent increase in glycosylation of the haemoglobin was subsequently calculated and expressed as:

  $$\%\mathrm{Haemoglobin}\mathrm{glycosylation}=[({\mathrm{A}}_{\mathrm{S}}-{\mathrm{A}}_{\mathrm{O}})/{\mathrm{A}}_{\mathrm{S}}]\times 100$$ 8

Antidiabetics biomarkers

• (i)Glucose uptake capacity by yeast cells
  This was evaluated in a mixture containing appropriate volume of extracts (100 µg, 200 µg and 500 µg), and 1 mL glucose solution (25 mM). This was incubated for 10 min at 37 °C. Subsequently, 100 μL of 10% v/v yeast suspension was added to each mixture, this was vortexed The PerkinElmer Spectrum™ 3 FT-IR spectrometer (L1280138) and further incubated for 60 min at 37 °C. The mixture was further subjected to centrifugation for 5 min at 3800 rpm, and absorbance of the samples (AS) and control (has no extract; AO) solutions were individually read at 520 nm. Metformin was used as standard [51]. The percent increase in glucose uptake capacity by yeast cells was subsequently calculated and expressed as:

  $$\%\mathrm{Glucose}\mathrm{uptake}\mathrm{capacity}\mathrm{activity}=[({\mathrm{A}}_{\mathrm{O}}-{\mathrm{A}}_{\mathrm{S}})/{\mathrm{A}}_{\mathrm{O}}]\times 100$$ 9
• (ii)Αlpha-amylase inhibition

  The assay mixture consisted of varying concentrations of extracts (100 µg, 200 µg and 500 µg), and sodium phosphate buffer (0.02 M, pH 6.9) which contained 0.5 mg/mL α-amylase solution. These were incubated at 25 °C for 10 min and 500 mL of 1% starch solution in sodium phosphate buffer (0.02 M, pH 6.9) was subsequently added. The mixtures were incubated at 25 °C for 10 min and the reaction was halted by the addition of 1.0 mL of dinitrosalicyclic acid color reagent. The resulting mixture was later subjected to incubation in a boiling water bath for 5 min. After cooling, absorbance of the samples (AS) and control (has no extract; AO) solutions were individually read at 504 nm and acarbose was used as standard [52].

  The percent of alpha-amylase inhibition of the extracts were subsequently calculated and 396 expressed as

  $$\%\mathrm{alpha}-\mathrm{amylase}\mathrm{inhibition}\mathrm{activity}=[({\mathrm{A}}_{\mathrm{O}}-{\mathrm{A}}_{\mathrm{S}})/{\mathrm{A}}_{\mathrm{O}}]\times 100$$ 10
• (iii)Αlpha-glucosidase inhibition

  This was evaluated using the method described by Ademiluyi et al. [53]. The reaction mixtures contained 60 μL of varying concentrations of extracts (100 µg, 200 µg and 500 µg), and 50 μL phosphate buffer (0.1 M, pH 6.8) containing 0.2 U/mL α-glucosidase solution. The mixtures were incubated at 37 °C for 20 min. Thereafter, 50 μL of 5 mM p-nitrophenyl-α-D-glucopyranoside (PNPG) solution in phosphate buffer (0.1 M, pH 6.8) was added and the mixtures were incubated at 37 ºC for 20 min. The reaction was stopped by the addition of 160 μL of 0.2 M Na2CO3. The absorbance of the samples (AS) and control (has no extract; AO) solutions were individually taken at 405 nm and acarbose was used as standard [54].

  The percent of alpha-glucosidase inhibition of the extracts were subsequently calculated and expressed as % alpha-glucosidase inhibition activity following (Eq. 6).

Statistical analysis

The analysis was done using Graph Pad Prism (V.5.0). All results were expressed as mean ± standard deviation (SD). One-way Analysis of Variance (ANOVA) followed by student-paired T-test was done and differences between groups were regarded as significant at P ≤ 0.05.

Results

Photo-physical characterization of synthesized selenium nanoparticles (Se-NPs)

UV–Visible spectroscopy characterization of se-NPs

The leaves and arils synthesized Se-Nps (Se-NP_L and Se-NP_A) were as presented in Supplementary Fig. 1a, 1b and 1c, which represent 10 mM Na₂SeO₃ solution and Ackee leaves (AKL); 10 mM Na₂SeO₃ solution and arils (AKA); Se-NP_L and Se-NP_A, respectively. The UV–Visible spectrum revealed a very sharp peak absorbance at 310 nm in both AKL and AKA (Fig. 1). This wavelength represents where selenium synthesized nanoparticles absorb maximally.

Fig. 1.

UV- Visible spectrum of Ackee (a) leaves and (b) arils Se-NPs

Fourier transform infrared (FTIR) spectroscopy characterization of se-NPs

FTIR spectra of Se-NP_L and Se-NP_A were as presented in Fig. 2. Multiple intensity peaks at 4000 cm⁻¹- 400 cm⁻¹ were found in both the leaves and arils Se-NPs spectra. Of utmost importance of these peaks were OH-group of 3348.8 cm⁻¹ and 3333.6 cm⁻¹; amide vibration of 1640 cm⁻¹ and 1649.6 cm⁻¹; C-H bending in alkynes of 1358.9 cm⁻¹ and 1395 cm⁻¹; C-H bending of polysaccharides of 1049.9 cm⁻¹ and 1100.3 cm⁻¹; C–C stretching vibration of 813.48 cm⁻¹and 919 cm⁻¹; C-X stretching in alkyl halide of 789.25 cm⁻¹ and 774.1 cm⁻¹; C-N–C bending in amine of 543.9 cm⁻¹ and 498.46 cm⁻¹ and Se-O stretching vibration of 774.1 cm⁻¹ and 1049.9 cm⁻¹ for the AKL and AKA respectively. All these confirm the existence of reducing groups in the Se-Nps.

Fig. 2.

FTIR of Ackee (a) leaves and (b) arils Se-NPs

Scanning electron microscopy (SEM) characterization of se-NPs

The Morphologies of Se-NPs; Ackee leaves (Se-NPL) and arils (Se-NPA) were confirmed by SEM analysis and were found to be amorphous granular in shape with notable spikes (Fig. 3). SEM images of 1 µm and 3 µm taken at a magnification of 25,000X (Fig. 3a). and 10,000X (Fig. 3b) for Se-NPL and Se-NPA, respectively. Other images taking at lower magnification are represented in Supplementary Fig. 2a and 2b for Se-NPL and Se-NPA, respectively.

Fig. 3.

SEM image of Ackee (a) leaves and (b) arils Se-NPs

Energy-dispersive x-ray spectroscopy (EDS) characterization of se-NPs

The EDS showed the existence of two strong Se peaks taken at 30 kV, 10000X magnification, 36.93 takeoff, 60 live times, 7.68-amp time and 128.5 resolution of the EDS spectrum. This confirms the presence of elemental and atomic Se to be 70.19% and 86.17% in Se-NPL (Fig. 4a) and 67.74% and 84.75% in Se-NPA (Fig. 4b) respectively.

Fig. 4.

EDS spectra of Ackee (a) leaves and (b) arils Se-NPs

Antioxidant potentials of crude methanolic (AKL and AKA) and nano synthesized Se-NPs (Se-NPL and Se-NPA) of ackee leaves and arils extracts

The results obtained for Total Antioxidant Capacity (TAC) as presented in Table 1 showed that Se-NPL (10 mg/mL) had 0.7 times lesser activity compared to the crude leaves (AKL; 10 mg/mL) methanolic extract and its activity compared to 5.75 times of Vitamin C (200 mg/mL), a potent standard antioxidant drug. Moreover, the Se-NPA (10 mg/mL) gave 1.12 times higher activity in comparison with crude Ackee arils (AKA; 10 mg/mL) methanolic extract and its activity compared to 4.8 times of Vitamin C (200 mg/mL) antioxidant potential.

Table 1.

Antioxidant potentials of crude methanolic and Se-NPs Ackee leaves and arils extracts

Results in red indicate lower activity; green indicates higher activity while blue indicates no significant differences, when AKA, AKA are compared with their Se-NPs (Se-APL and Se-APA)

Values represent averages of means (n = 3). Results were considered statistically significant at P ≤ 0.05

Where: Se-NPL = Nano synthesized Ackee leaf; Se-NPA = Nano synthesized Ackee arils; AKL = crude Ackee leaf; AKA = crude Ackee arils, and Ascorbic acid = Standard drug

TAC Total Antioxidant Capacity, OH Hydroxyl Radical Scavenging, FRAP Ferric Reducing Antioxidant Power, ABTS 2,2'-Azino-Bis-3-Ethylbenzothiazoline-6-Sulfonic acid, DPPH 2, 2-Diphenyl-1-Picrylhydrazyl, NO  Nitric oxide scavenging, SARS Superoxide Anion Radical Scavenging

The values obtained for hydroxyl radical scavenging property indicated that Se-NPL (10 mg/mL) had 1.27 times greater activity when compared with the AKL (10 mg/mL) methanolic extract and its activity compared to 0.96 times of Vitamin C (200 mg/mL) antioxidant potential. Furthermore, Se-NPA (10 mg/mL) gave 0.92 times activity in comparison with AKA (10 mg/mL) methanolic extract and its activity compared to 0.89 times of vitamin C (200 mg/mL) antioxidant potential (Table 1).

The values obtained for superoxide anion radical scavenging antioxidant property indicated that the Se-NPL (10 mg/mL) had 1.05 times greater activity when compared with AKL (10 mg/mL), while its activity was 12.92 times when compared to vitamin C (200 mg/mL) antioxidant potential. In addition, the synthesized Se-NPA (10 mg/mL) gave 1.89 times lower activity compared to AKA (10 mg/mL), while its activity was 5 times higher when compared to vitamin C (200 mg/mL) antioxidant potentials.

The results obtained for DPPH antioxidant property as displayed in Table 1 showed that the Se-AKL (10 mg/mL) and AKL (10 mg/mL) had similar activity, while Se-NPL has 18 times lesser activity when compared to vitamin C (200 mg/mL) antioxidant activity. Likewise, the Se-NPA (10 mg/mL) and AKA (10 mg/mL) had similar activity, while Se-NPA has 166 times lesser activity when compared to vitamin C (200 mg/mL) antioxidant activity.

The nitric oxide scavenging activity as also presented in Table 1 showed that the Se-NPL (10 mg/mL) had 0.43 times lesser activity compared to AKL (10 mg/mL), while its activity was 22.76 times when compared to that of vitamin C (200 mg/mL) antioxidant potential. Moreover, the Se-NPA (10 mg/mL) had 2.7 times less activity in comparison with AKA (10 mg/mL), while its activity was 17.25 times when compared to vitamin C (200 mg/mL) antioxidant property.

The results obtained for ferric reducing antioxidant power (FRAP) as shown in Table 1 indicated that Se-NPL (10 mg/mL) and AKL (10 mg/mL) methanolic extract had relatively similar activity and Se-NPL activity compared to 2 times of vitamin C (200 mg/mL) antioxidant potential. Likewise, Se-NPA (10 mg/mL) and AKA (10 mg/mL) methanolic extract had similar activity, and Se-NPA activity was compared to 1.31 times of Vitamin C (200 mg/mL) antioxidant potentials.

The values obtained for ABTS antioxidant evaluation indicated that Se-NPL (10 mg/mL) gave 1.8 times greater activity in comparison with AKL (10 mg/mL) and its activity was 5.7 times when compared to vitamin C (200 mg/mL) antioxidant potential. In like manner, the synthesized Se-NPA (10 mg/mL) gave 2.3 times greater activity when compared with the AKA (10 mg/mL) and its activity was 21.87 times when compared to that of vitamin C (200 mg/mL) antioxidant potentials (Table 1).

Anti-inflammatory and antidiabetic potentials of crude methanolic (AKL and AKA) and nano synthesized se-NPs (Se-NPL and Se-NPA) of ackee leaves and arils extracts

The potential health advantages of several components derived from the Ackee plant [particularly, the leaves (AKL), arils (AKA), and their cognate Selenium-NPs (Se-NPL and Se-NPA)] were investigated by assessing various biological anti-inflammatory activities: membrane stabilization, anti-arthritic potential, anti-haemolysis, and haemoglobin glycosylation; and antidiabetic activities: glucose uptake, α-amylase inhibition, and α-glucosidase inhibition. Their actions were compared to the therapeutic efficacy of standard medications, particularly in the context of diabetes and inflammation. Various amounts of substances were quantified at levels of 100, 200, and 500 µg/mL. The activities of extracts derived from AKL and AKA exhibit an increase in response to varying concentrations, indicating a dose-dependent relationship. These effects are observed to be more pronounced at greater concentrations of the extracts. Summarized data of AKL, AKA, Se-NPL and Se-NPA are as presented in Supplementary Table 1..

Anti-inflammatory potentials

The percent membrane stabilizing activity of AKL, AKA, Se-NPL and Se-NPA and Aspirin (STD-ASP) used as standard drug are as presented in Fig. 5a. It showed an overall increase in membrane stabilizing activities for all the samples as the concentration increases. The membrane stabilizing activity of AKL was observed to be the highest across all administered doses. The membrane stabilizing activity of AKA ranks second, following Se-NPL and Se-NPA, though the membrane stabilizing activity of STD-ASP was significant enough, the activity recorded for the four samples was higher.

Fig. 5.

The percent of: a membrane stabilization; b anti-anthritic activity; c anti-haemolysis; d hemoglobin glycosylation, of crude methanolic (AKL and AKA); and synthesized Se-Nps (Se-NPL and Se-NPA)

Figure 5b showed the result obtained for the assessment of the degree of hyaluronidase inhibition, an enzyme responsible for the degradation of hyaluronic acid, a significant constituent of joint cartilage that serves as an indicator of the efficacy of anti-arthritic properties. This depicts the relationship between concentration and anti-arthritic efficacy for AKL, AKA, Se-NPL and Se-NPA, demonstrating that as the concentration increases, and so does the effectiveness in managing arthritis. Though the anti-arthritic activity of Diclofenac (STD-DIC); used as standard drug was significantly higher at all concentrations studied when compared to the four samples, Se-NPL demonstrates the most pronounced anti-arthritic efficacy across all concentrations. The order of anti-arthritic activity thus follows: Se-NPL > AKL > AKA > Se-NPA.

The anti-haemolysis activities of AKL, AKA, Se-NPL and Se-NPA are as presented in Fig. 5c. The evaluation of anti-haemolysis activity involves quantifying the extent to which red blood cell disintegration is inhibited and expressed as a percentage. This illustrates that the anti-haemolysis activity of all the four samples and Butylated hydroxyanisole (STD-BHA); used as standard drug were significant to inhibit haemolysis. In all concentrations studied, STD-BHA demonstrated higher inhibition, however, the order of anti-haemolysis activity follows: Se-NPL > Se-NPA > AKA > AKL.

The percent of hemoglobin glycosylation of the AKL, AKA, Se-NPL and Se-NPA and Gentamicin (STD-GT) used as standard drug are as presented in Fig. 5d. The figure indicated that the lowest level of hemoglobin glycosylation was exhibited by STD-GT; while following descending order the hemoglobin glycosylation activity was found to be Se-NPL > Se-NPA AKL > AKA. The results therefore reflect the efficacy of the different extracts and Se-NPs in promoting the binding of glucose to the hemoglobin, even higher than the standard drug STD-GT at all tested concentrations.

Antidiabetic potentials

Figure 6a illustrates the relationship between glucose uptake capacities with the varying concentrations of extracts (AKL, AKA), Se-NPs (Se-NPL, Se-NPA) and Metformin (STD-MET) used as standard drug. Although, the glucose absorption rate increases with increasing concentration for all samples, AKL had the maximum glucose absorption across all concentrations. The glucose absorption capacity of AKA is however second only to Se-NPL. The STD-Met exhibits the lowest absorption of glucose.

Fig. 6.

The percent of: a glucose uptake capacity; b α-amylase; c α-glucosidase of crude methanolic (AKL and AKA); and synthesized Se-Nps (Se-NPL and Se-NPA)

The inhibitory effects of AKL, AKA, Se-NPL and Se-NPA on alpha-amylase and alpha-glucosidase activities are as presented in Fig. 6a and b, respectively. It indicated that more pronounced inhibition was obtained as doses of the samples and Acarbose (STD-ACB); used as standard drug, increases. In all concentrations studied, STD-ACB showed higher inhibitory effects on these two enzymes, however, the order of anti-diabetic activity follows: AKL > AKA > Se-NPL > Se-NPA.

Discussion

The antioxidant potentials of Ackee leaves (AKL) and arils (AKA) extracts with their synthesized Se-NPs (Se-NPL and Se-NPA) were assessed through their total antioxidant capacity, hydroxyl radical scavenging property, ferric reducing antioxidant property, ABTS radical scavenging property, DPPH radical scavenging property, nitric oxide radical scavenging property and superoxide anion radical scavenging property. The extracts and Se-NPs displayed excellent antioxidant potentials in all the evaluated parameters with the maximum activity found in the nitric oxide radicals scavenging and ABTS properties, respectively. These antioxidant properties exhibited in AKL and AKA could be attributed to the presence of several phytochemical compounds such as alkaloids and phenolics [55]. This report agrees with the previous studies where Ackee leaves and arils extracts demonstrated excellent antioxidant properties [24, 27].

With the aid of UV–visible spectroscopy, which is a potent analytical instrument, nanoparticles can be classified according to their absorption spectra [56]. Intensity of the produced nanoparticles' absorbance at various wavelengths is displayed in the spectra. Peaks and valleys in the spectrum reveal details on the dimensions, morphology, and makeup of the nanoparticles. These revealed the characteristics of the selenium synthesized nanoparticles (Se-NPs) of Ackee leaves and arils extracts. These characteristics can be used to optimize the properties of these nanoparticles for a variety of applications, such as biomedical, environmental, and industrial uses [34]. The spectra of Se-NPs made from Ackee leaves and arils exhibited a broad absorption band with a peak at 310 nm. This agrees with previous research works that have also reported 310 nm as the wavelength for the detection of selenium nano-synthesized particles [57, 58]. Furthermore, reports of Kokila et al. [59] and Anu et al. [60] who have prepared Se-NPs from Diospyros montana leaf and Allium sativum extracts, respectively by green synthesis approach and observed UV–visible absorption maximum at 310 nm. Likewise, Fesharaki et al. [61] have observed UV–visible maximum at 280 and 310 nm for Klebsiella pneumoniae mediated bio-synthesized Se-NPs. On the other hand, Shah et al. [62] have synthesized polyvinyl alcohol-stabilized Se-NPs by wet chemical method involving a reaction of acetone and noticed UV–visible maximum at 310 nm. The variations in the composition and characteristics of the plant components employed in the synthesis may be the cause of the disparities in spectra between the nanoparticles made from Ackee leaves and arils. The decrease and stabilization of the nanoparticles may be influenced by the presence of various phytochemicals and metabolites in Ackee leaves and arils. This clearly showed that sodium selenite was successfully converted into Se-NPs by reduction action of Ackee leaves and arils methanolic extracts.

FTIR is also a potent application that aids in the identification of functional groups present in a sample and offers details on chemical bonding, molecular vibrations, and structural characteristics. FTIR spectroscopic analysis was performed to further confirm the feasible role of Ackee leaves and arils extracts in the bio-synthesis of Se-NPs. FTIR allows the determination of the functional groups that exist on the surface of nanoparticles by measuring the vibrational frequencies of chemical bonds. The FTIR spectra of Ackee leaves and arils Se-NPs reflected useful details on the functional groups present in the produced nanoparticles and aids in identifying their structural characteristics and chemical make-up based on their distinctive peaks [63]. The molecular data obtained assists to establish structural/conformational changes of the self-assembled functional groups on the surface of nanoparticles [64]. The result of FT-IR analysis of synthesized Se-NPs has shown in Fig. 2a and b, showed broad peaks at 3348.8 cm⁻¹ and 333.6 cm⁻¹ corresponds to O–H stretch alcohols and phenols. This suggested that selenium has interacted with the hydroxyl group from leaves and arils extracts of Ackee plant through hydrogen bonding and facilitated biosynthesis of Se-NPs. Likewise, prominent peaks at 1649.5 cm⁻¹ and 1640.4 cm⁻¹ which are amide vibrations which show the interaction of proteins of leaves and arils extracts of Ackee plants with selenium through the amine groups. The intense peaks at 1049.7 cm⁻¹ and 774.1 cm⁻¹ in arils and leaves extracts of Ackee plants represent the characteristics Se-O stretching vibration and accomplishment of the successful biosynthesis of Se-NPs as reported by [65]. Peaks at 1050 cm⁻¹, 1001.3 cm⁻¹ which are R-O-R ether and 1660 cm⁻¹ H–N–H, are all indicative of carbohydrate and protein characters respectively. These are consistent with all these previous investigations [64, 65] that demonstrate the presence of several functional groups. The various peaks confirm the role of different phytochemicals in facilitating the biosynthesis of Se-NPs by reduction and increasing the stabilization of nanoparticles.

The SEM images demonstrated the amorphous and granular shape and largely consistent size distribution of the Se-NPs. The arils appear to contain nanoparticles that are slightly larger in size than the leaves. Ahmeda et al. [66] reported a similar outcome, whereby synthesized nanoparticles manufactured using Ziziphora clinopodioides Lam. exhibited a homogeneous size distribution and spherical shape.

The Energy-Dispersive X-ray Spectroscopy (EDS) technique examines the elemental makeup of materials, including nanoparticles. The matching peaks in the spectra can also be used to estimate the relative concentrations of these nanoparticles. The EDS spectra of Ackee leaves and arils Se-NPs showed that Selenium (Se) was present, the existence of two strong Se peaks taken at 30 kV, 10000X magnification, 36.93 takeoff, 60 live time, 7.68 amp time and 128.5 resolution of the EDS spectrum also confirm the presence of elemental and atomic Se to be 70.19% and 86.17% for the Ackee leaf (AKL; Fig. 2a), and 67.74% and 84.75% respectively, for Ackee arils (AKA; Fig. 2b), respectively. The Se-NPs are present in relatively high concentrations, as evidenced by the prominent peak that corresponds to Se. These nanoparticles have been found to have distinctive physicochemical features that may affect their biological interactions, these findings have implications for the possible health impacts of ingesting Ackee plants and their synthesized Se-NPs [36, 37].

The intrinsic biological properties of crude methanolic and synthesized Se-NPs extracts of Ackee leaves and arils were evaluated to provide crucial insights their health implications. These investigations facilitate the assessment of the therapeutic efficacy of these substances in the context of diabetes, inflammation, and cellular health as opined by Iravani and Varma [67, 68]. The identification of hemoglobin glycosylation across many categories represents a significant discovery. Glycosylation is utilized as a means to quantify the quantity of glucose that is attached to the protein hemoglobin within red blood cells. The process of hemoglobin glycosylation leads to an elevation in mean blood glucose levels [50]. To comprehend the quantitative ramifications of glucose binding to hemoglobin within erythrocytes, it is imperative to employ this particular methodology. It’s believed that Glycosylation plays significant roles in the aging and protection of red blood cells from oxidative damage, which consequently contributes to their longevity by facilitating prolonged red blood cells interaction with sialic acid [69, 70]. As red blood cells age, they lose their sialic acid coating making them susceptible to destruction by spleen. The life span of red blood cell is usually around 120 days, however, the presence of sialic acid on the surface of the cells can extend their life span of red blood cells by up to 50% (170 days) [69, 70]. It is shown that the presence of sialic acid makes the red blood cells more hydrophilic, which allows them to circulate more easily in the blood stream [69, 70]. Se-NPs and Ackee extracts exhibited higher incidence of hemoglobin glycosylation more than the standard drug Gentamicin (Fig. 5a). This shows clearly the physiological roles consumption of these Ackee plants parts could have been conferring on the consumers and this should be strongly pointed out and advocating more consumption should be made.

The membrane stabilizing function of a plant extract suggests its potential for protecting cell membranes. Cell membranes are composed of lipids and proteins, serving many functions such as nutrient and waste transportation, as well as cellular signaling. Cell death can occur as a result of impaired cell membranes [71]. Diseases characterized by cell membrane damage, such as inflammation, cancer, and Alzheimer's disease, could potentially be addressed through the utilization of innovative drugs or therapies employing these plant extracts, as indicated by the data obtained in this study. Based on the findings, the extracts derived from Ackee leaves have the most potent characteristics in terms of stabilizing cell membranes (Fig. 6a). This discovery implies potential applications in the protection of cell membranes, which play a crucial role in numerous biological processes [71, 72].

The capacity of a plant extract to absorb glucose is a metric used to evaluate its effectiveness in enhancing cellular glucose uptake. Cells derive energy from the molecule glucose. Impaired glucose uptake is a characteristic feature of Type II diabetes [73]. The data obtained underscores the potential utilization of plant extracts for the development of pharmaceutical interventions or treatments for Type II diabetes. Hence, the utilization of Ackee plant extracts has the potential to contribute to the development of a novel oral diabetes treatment aimed at assisting individuals with Type II diabetes in managing their blood glucose levels as results obtained for glucose uptake of the Ackee extracts and Se-NPs were far higher than metformin. Another study has demonstrated that extracts from Ackee leaves exhibit a notable capacity for glucose absorption [74, 75]. This implies that the utilization of these extracts may have the potential to enhance cellular glucose uptake, a critical aspect in the management of Type II diabetes.

The observed anti-arthritic properties of a botanical extract suggest its potential for safeguarding joint health. Arthritis is a physiological condition characterised by joint inflammation and associated suffering. Arthritis is typified by the degradation of joint cartilage [76, 77]. This study showcases plant extracts that have the potential to be utilized in the development of pharmaceutical interventions or therapeutic approaches for individuals suffering from arthritis. The utilization of Ackee leaves and arils extracts holds potential for the development of a topical cream or oral medication aimed at alleviating joint inflammation and pain in individuals suffering from arthritis.

The anti-haemolysis activity of a plant extract refers to its ability to provide protection to red blood cells. Haemolysis may arise due to various factors, including infections, toxic substances, and pharmaceutical agents. The occurrence of damage to red blood cells can result in the development of anemia, a medical disorder characterized by a deficiency of healthy red blood cells that are responsible for the transportation of oxygen to various organs inside the body. Novel drugs or therapies utilizing plant extracts may be employed in the treatment of malaria, sickle cell anemia, and drug-induced anemia. The observed features of the Ackee extracts and Se-NPs indicate their potential efficacy in mitigating arthritis and preventing haemolysis, hence offering protective effects on both joints and red blood cells. This implies that the utilization of these botanical extracts may hold potential in the development of medications or therapies targeting arthritis and anemia.

Persistent hyperglycemia and disturbed protein, lipid, and carbohydrate metabolism are hallmarks of diabetes mellitus. For the treatment of diabetes, inhibitors of the essential carbohydrate metabolism enzymes; alpha-amylase and alpha-glucosidase are used. They facilitate the hydrolysis of starch molecules, resulting in the production of glucose. The inhibition of these enzymes result in delayed carbohydrate digestion and glucose absorption [54]. Therapeutic drugs such as acarbose, miglitol, and voglibose used in management of diabetes are α-amylase and α-glucosidase inhibitors [78, 79]. Thus, the alpha-amylase and alpha-glucosidase inhibitory actions of the Ackee extracts and Se-NPs are of important therapeutic value. Hence, suggesting their potential anti-diabetic capabilities and could be exploited in the development of pharmaceutical interventions for the treatment of diabetes.

Consequently, Ackee extracts (AKL and AKA) and their Se-NPs (Se-NPL and Se-NPA) demonstrate potential in many health indices. They showed enhanced haemoglobin glycosylation, membrane integrity, glucose absorption, and they exhibited possible anti-arthritic properties. This suggests that they may have the ability to enhance glucose metabolism and alleviate symptoms associated with arthritis. The observed relationship between dosage and outcome provides evidence for the therapeutic capabilities of these samples. Thus, our obtained data provide evidence of the possible health benefits of AKL, AKA, Se-NPL and Se-NPA, and their efficacy in managing various health conditions [80, 81].

By conducting a comparative analysis of the constituents with standard conventional medications, most importantly, Gentamicin (GT), Aspirin (Asp), Metformin (Met), Diclofenac (Dic), Butylated hydroxyanisole (BHA), and Acarbose (Acb), it becomes evident that certain compounds and concentrations exhibit comparable or superior biological efficacy, notably in terms of their anti-haemolysis, haemoglobin glycosylation, membrane-stabilizing effects and anti-arthritic potentials. The utilization of natural chemicals derived from the Ackee plant parts, thus has the potential to serve as viable alternatives or supplementary adjuncts for the treatment of inflammatory, cellular membrane illnesses, diabetes and hemolytic disorders, and could also enhance glucose metabolisms in general [80].

In most cases, it was observed that Se-NPL exhibited greater biological activity as compared to Se-NPA. It therefore showed that Se-NPL exhibited notable enhancements in many biological parameters, including haemoglobin glycosylation, membrane stabilization, glucose uptake, anti-arthritic potential, anti-haemolysis, α-amylase inhibition, and α-glucosidase inhibition, as compared to Se-NPA. This does suggest that Se-NPL may possess promising biological efficacy. Interestingly, both AKL and AKA were recently characterized by Ibraheem et al. [24], several alkaloids, phenolics and saponin compounds were found to be present. Hence, it’s suggested that synergistic interactions between these bioactive compounds in both AKL, AKA and their Se-NPs could have contributed significantly to the observed antioxidative, antidiabetic and anti-inflammatory activities.

Conclusion

This study demonstrates that the components of the crude methanolic and Selenium nano synthesized particles (Se-NPs) of Ackee leaves (AKL) and arils (AKA) have the potential to confer health advantages. This study investigated the photo-physical characteristics of the Se-NPs, the in-vitro antioxidant activities of AKL, AKA and Se-NPs, and their biological activities; haemoglobin glycosylation, membrane stability, glucose absorption, anti-arthritic potential, anti-haemolysis, α-amylase inhibition, and α-glucosidase inhibition. These activities demonstrate a correlation that depends on three varying concentrations, whereby extracts exhibit larger advantages at higher concentrations. FTIR spectra showed the presence of different functional groups in the Se-NPs, and these might be of significant medicinal importance. In future we proposed the use of Dynamic Light Scattering (DLS) so as to analyse the size distribution of the Se-NPs as different sizes were synthesized as illustrated in Supplementary Fig. 2. The use of these Se-NPs in management of illness could potentially eliminate the challenges associated with the use of chemically synthesized agents that often have contraindications when taken. Since the Se-NPs have good cellular biocompatibility and act as potential membrane protectors, they could therefore be useful in management of illness associated to cellular damages. Overall, our findings indicated that natural ingredients from Ackee plants could have the potential to be utilized as alternative or supplementary therapeutic options for diabetes, arthritis and hemolytic disorders. The wide therapeutic window and reduced toxicity Se-NPs could be attributed to their potentials in membrane stability and antioxidant properties [29, 30]. The biological mechanism employed by Se-NPs among others which may include: reactive oxygen species (ROS) scavenging ability; reducing oxidative stress and lipid peroxidation that helps in maintain membrane integrity; can be incorporated into the membrane, which thus increase membrane fluidity and reduce rigidity that influences lipid raft formation, increasing resistance to stress [30, 31]. Se-NPs can also interact with membrane proteins, maintaining their structure and function, consequently, preventing denaturation, reducing changes in osmotic potential due to nanoparticle concentration [31].Se-NPs can also decrease water permeability through the membrane, reducing osmotic stress and modulate aquaporin activity by interacting with aquaporins, regulating water transport and maintaining osmotic balance [29–31].

Data obtained by our in-vitro experimentation may not comprehensively capture the intricate interactions that occur within living organisms. Thus, additional research, encompassing in-vivo and clinical studies are important to ascertain and understand the mechanisms through which these constituents can mitigate and manage diverse health conditions. It will also be imperative that very thorough cytotoxicity assays are carried out in order to fully establish the safety and/ or any potential side effects of these Se-NPs and the AKL and AKA methanolic extracts could have on human cellular physiology. Since Se-NPs are well tolerable to cell membrane it will be therefore imperative to evaluate the tolerable concentration that wouldn’t induce cytotoxicity. We believe the concentration used in this research (10 mg/ml), may not induce cytotoxicity. However, by conducting the pharmacokinetics (PK) and pharmacodynamics (PD) studies will ensure the understanding of the bio-distribution of Se-NPs as regards their absorption, distribution, metabolism and excretion in the body, ensuring them reaching the intended target sites. PK and PD will also assist in evaluating the efficacy and safety of the Se-NPs on biological systems, revealing their therapeutic potentials and probable toxicity; optimizing dosing regimens by informed dosage selection; minimizing adverse effects while maximizing therapeutic benefits; consequently, ensuring their effective use in future medicinal and nutritional applications.

Authors’ contributions

OI conceptualized, designed, and supervised the study, and edited the manuscript, OHO contributed to data analysis and writing of the manuscript, OMO and EOA contributed equally to data collection and analysis and writing of the manuscript. All the authors reviewed the final version of the manuscript and agreed to publication.

Funding

This research did not receive any fund, grant or support from any organization.

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

Whole cell hemoglobin O- blood group employed in some of the biological in-vitro assays in this research work, were previously collected from healthy volunteers deposited at Federal University Oye Ekiti, University Clinic Blood Bank. The consent for use has been given by the donors for clinical and experimental purposes and ethical approval was giving by our University Research Committee for its collection and use. The experimental methods used were approved by the Research Ethics Committee for animal experimentation of the Faculty of Science, FUOYE with approval number: FUOYEFSC 201122 – REC2024/015.Our work does not require any direct patient trial; hence there is no Clinical Trial Number for this research as in-vitro assays were only conducted throughout the course of this research work.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.