GC-MS Characterization of Bioactive Phytochemicals in Ethanol Seed Extract of Annona Muricata Cultivated in Nigeria

GC-MS Characterization of Bioactive Phytochemicals in Ethanol Seed Extract of Annona Muricata Cultivated in Nigeria

Fatima Mutiu Odejayi, Iheanyichukwu Wopara, Akuru Owota

Department of Biochemistry, Rivers State University, Port Harcourt, Nigeria

Abstract – Annona muricata (soursop) is a medicinal plant commonly used in traditional healthcare for treating inflammatory and metabolic conditions. Although the leaves of this plant have been extensively investigated, there is comparatively little information available regarding the phytochemical profile of its seeds. The present study sought to identify the chemical components of A. muricata seed extract through gas chromatographymass spectrometry (GCMS) analysis. Finely ground dried seeds were extracted with 80% ethanol and analyzed using an Agilent 7890A5975C GCMS system. Compound identification was performed by matching mass spectra with entries in the NIST library database. The GC-MS analysis revealed the presence of key fatty acids, notably Oleic acids (9-octadecenoic acid) (32.22%), Palmitic acids (17.92%), and (9,12-octadecadienoic) Linoleic Acids with 26.05%). Oleic acids had the highest percentage. These fatty acids are recognized for their antioxidant, anti-inflammatory, and antiproliferative activities. The high proportion of unsaturated fatty acids underscores the therapeutic potential of A. muricata seeds and supports their further exploration in biological and pharmacological research.

Keywords: Annona muricata, GCMS, Phytochemicals, Antioxidant, Medicinal plants

INTRODUCTION

Medicinal plants continue to serve as an important source of therapeutic agents because of their abundant bioactive secondary metabolites. Phytochemicals obtained from plant sources have been shown to exhibit a broad spectrum of pharmacological properties, including antioxidant, anti-inflammatory, antimicrobial, and anticancer activities (Moghadamtousi et al., 2015). Consequently, the chemical profiling of plant extracts is crucial for elucidating their biological effects and potential use in pharmaceutical development.

A. muricata ,a member of the Annonaceae family, is a tropical plant commonly known as soursop. Different parts of the plant, such as the leaves, bark, fruit, and seeds, have long been used in traditional medicine for managing conditions including fever, pain, inflammation, diabetes, and cancer-related ailments (Adewole & Ojewole, 2009). Earlier investigations have indicated that A. muricata is rich in various bioactive constituents, including acetogenins, alkaloids, flavonoids, phenolic compounds, and fatty acids, all of which contribute to its therapeutic potential (Moghadamtousi et al., 2015).

Despite extensive research on the leaves of A. muricata , relatively limited information is available regarding the phytochemical composition of its seeds. Seeds are recognized as significant sources of lipid-based compounds, particularly fatty acids, which are essential for antioxidant protection, maintenance of membrane integrity, and regulation of inflammatory processes (Othman et al., 2023). Gas chromatographymass spectrometry (GCMS) is a robust analytical method widely used for the detection and quantification of volatile and semi-volatile phytochemicals, especially fatty acids and their derivatives.

Accordingly, this study was undertaken to characterize the phytochemical constituents of A. muricata seed extract using GCMS analysis, with a particular focus on identifying major bioactive fatty acids that may be responsible for the plants pharmacological effects.

Figure 1. refers to the various plant parts of Annona muricata: (A) whole plant, (B) leaves, (C) flower, (D) fruit, (E) seeds. Source:

(Coria-Téllez et al.,2022).

Figure 1. Refers to the various plant parts of Annona muricata: (A) whole plant, (B) leaves, (C) flower, (D) fruit, (E) seeds. Source:

(Coria-Téllez et al., 2022).

1. Materials and Methods

   1. Plant Material Collection and Preparation

      A. muricata seeds were obtained from the fresh fruits. The seeds were thoroughly washed, air-dried at room temperature for four weeks, and milled into fine powder.

      Five hundred grams (500 g) of the powdered seed material were exhaustively macerated in 80% ethanol for 72 hours, following standard extraction procedures (Nia et al., 2004). The extract was filtered using a Buchner funnel and Whatman No. 1 filter paper. The filtrate was concentrated under reduced pressure using a rotary evaporator (Buchi Rotavapor R-200) and stored at 4 °C until analysis.

   2. GCMS Analysis

      GCMS analysis was performed using an Agilent 7890A gas chromatograph coupled with a 5975C mass selective detector. Separation was achieved on an HP-5MS capillary column (30 m × 0.25 mm × 0.25 µm). Helium was used as the carrier gas at a constant flow rate of 1 mL/min. The injection volume was 0.5 µL with a split ratio of 10:1. The injector temperature was maintained at 250 °C, and the ion source temperature was set at 280 °C.

      The oven temperature was programmed as follows: initial temperature of 70 °C (held for 2 min), ramped to 280 °C at 15 °C/min, and held for 5 min. Mass spectra were acquired in electron impact mode at 70 eV. Compound identification was carried out by comparing the obtained mass spectra with those in the NIST mass spectral library.

2. RESULTS

   As shown in table 1, The GCMS profiling of A. muricata seed extract demonstrated a high abundance of fatty acids, this extract is dominated by oleic acid, which appears in the highest concentration (32.22%), and others such as Linoleic acid (26.05%), palmitic acid (17.92%) were identified as well.

   Table 1: Phytoconstituents component of A. muricata seed extract

   S/ N	Name of Compound	Retentio n Time (RT) (minutes )	Molecula r Formula r (Mf)	Molecula r Weight (Mw) (g/mol)	Percentag e (%)	Structure
   1	n- hexadecanoic acid	6.844	C16H32O2	256	17.92
   2	9-octadecenoic acid	7.599	C18H34O2	282	23.81
   3	9-octadecenoic acid	7.645	C16H34O2	282	32.22
   4	9,12- Octadecadieno ic acid	7.691	C18H32O2	280	26.05

3. DISCUSSION

   Gas chromatographymass spectrometry (GCMS) analysis of Annona muricata seed extract revealed a phytochemical profile predominantly composed of fatty acids and their derivatives. The chromatographic spectrum was characterized by prominent peaks corresponding to long-chain saturated and unsaturated fatty acids, confirming the lipid-rich nature of the seed matrix. Among the identified compounds, oleic acid (9-octadecenoic acid), linoleic acid (9,12-octadecadienoic acid), and n-hexadecanoic acid (palmitic acid) constituted the major components, collectively accounting for a substantial proportion of the total extract composition.

   Oleic acid (9-octadecenoic acid), representing approximately 32.22% of the extract, is a monounsaturated omega-9 fatty acid known for its antioxidant and anti-inlammatory properties. Its functional importance stems largely from its integration into cellular membranes, where it enhances membrane rigidity and reduces vulnerability to lipid peroxidation. Because oxidative stress primarily results from reactive oxygen species attacking membrane lipids, the presence of oleic acid improves resistance to oxidative damage and decreases the generation of harmful byproducts such as malondialdehyde. Additionally, oleic acid modulates inflammatory pathways by downregulating pro-inflammatory cytokines and inhibiting NF-B activation (Sales-Campos et al., 2021; Carrillo et al., 2020). Through these mechanisms, it contributes to the reduction of chronic inflammation, a key factor implicated in tissue hyperplasia and metabolic dysfunction (Wang et al., 2020; Calder, 2015). Oleic acid likely plays a principal role in the protective and inhibitory activities observed in this extract.

   Linoleic acid (9,12-octadecadienoic acid), an essential omega-6 polyunsaturated fatty acid also detected in the extract, further enhances its biological relevance. Due to its double-bond structure, linoleic acid increases membrane fluidity and supports cellular communication processes (Calder, 2020; & Duan et al., 2022). It functions as a precursor for bioactive lipid mediators that regulate immune responses and inflammation (Fritsche, 2021). Although polyunsaturated fatty acids are more susceptible to oxidation, their regulated metabolic conversion produces signaling compounds essential for inflammation resolution and tissue repair. Within complex phytochemical systems, linoleic acid can act synergistically with other fatty acids and antioxidant molecules to improve overall redox balance. Its presence therefore reinforces the extracts capacity to regulate oxidative stress and inflammatory responses, especially in pathological states associated with abnormal cell proliferation and metabolic imbalance. (Wang et al., 2020; & Zhang et al., 2021).

   Palmitic acid (n-hexadecanoic acid), a saturated fatty acid identified in substantial amounts, also contributes to the extracts functional profile. While elevated systemic palmitic acid levels are associated with metabolic disorders, its role within plant-derived matrices differs (Resh, 2020; & Jiang et al., 2023). In phytochemical contexts, palmitic acid is a structural component of phospholipids and participates in protein palmitoylation a modification that influences membrane targeting and signaling protein activity (Wang et al., 2020; & Calder, 2023). Through these mechanisms, it indirectly modulates pathways linked to cell growth and metabolic regulation. In combination with unsaturated fatty acids, it also supports membrane structural integrity and may reduce oxidative membrane fragmentation. Some studies additionally attribute mild antimicrobial and anti-inflammatory effects to palmitic acid within complex plant extracts (Zhang et al., 2021; & Kumar 2018)

   Overall, the dominance of these fatty acids in the GCMS spectrum highlights the lipid-centered bioactivity of Annona muricata seed extract. The combination of saturated, monounsaturated, and polyunsaturated fatty acids creates a balanced lipid environment capable of stabilizing cellular membranes, regulating inflammatory mediators, and maintaining redox homeostasis (Calder, 2023; Wang et al., 2020; & Duan et al., 2022). Such interactions may underlie the observed enhancement of antioxidant defense systems including superoxide dismutase, catalase, glutathione peroxidase, and reduced glutathione as well as the suppression of lipid peroxidation. Consequently, the fatty acid composition provides phytochemical support for the extracts protective and potential therapeutic properties.

   1. Synergistic Biological Effects of Major Fatty Acids

      The fatty acids identified in Annona muricata seed extract may exert synergistic pharmacological effects by collectively enhancing antioxidant defense, stabilizing cellular membranes, and modulating inflammatory pathways more effectively than when acting individually (Calder, 2015; Sales-Campos et al., 2013).

4. MEDICINAL IMPORTANCE OF ANNONA MURICATA SEED EXTRACT

   While much attention has been given to its leaves and fruit pulp, the seeds also contain significant bioactive compounds, particularly fatty acids, acetogenins, alkaloids, phenolic compounds, and other lipid-based phytoconstituents. Annona muricata seeds shows the following;

   Possesses strong antioxidant activity, attributed to its fatty acids, acetogenins, and other phytoconstituents that help reduce oxidative stress and lipid peroxidation (Moghadamtousi et al., 2015; & Valko et al., 2007).

   Exhibits potential anti-hyperlipidemic properties, as fatty acids influence lipid metabolism and inflammatory balance (Calder, 2015).

   Cardio protective effects, particularly due to the presence of monounsaturated fatty acids like oleic acid, which are associated with improved lipid profiles and reduced inflammation (Sales-Campos et al., 2013; & Calder, 2015).

5. CONCLUSION

This study successfully characterized the phytochemical constituents of A. muricata seed extract using GCMS analysis. The extract was found to be rich in bioactive fatty acids, with oleic acid constituting the highest percentage of the identified compounds. The presence of these fatty acids underscores the pharmacological relevance of A. muricata seeds and supports their potential application in the development of natural antioxidant and therapeutic agents. Further studies are recommended to explore the biological activities and mechanisms of action of these compounds.

Acknowledgement: I would like to express my gratitude to the of department biochemistry laboratory, Rivers state university, Nkpolu – Oroworukwo, Port Harcourt, Rivers State, Nigeria.

Conflicts of Interest: The author declares no conflict of interest

Author Contributions: Fatima Mutiu Odejayi- Manuscript development, Iheanyichukwu Wopara: Experimental design, Material and methodology, Akuru Owota- Review and Editing

Funding Statement: This research did not receive any specific grant from funding agencies in the public, commercial, or not-for- profit sectors.

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