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Journal of Drug Delivery and Therapeutics

Open Access to Pharmaceutical and Medical Research

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Open Access Full Text Article                                                                    Research Article

Biogenic Synthesis of Copper Nanoparticles Using Hydroethanolic Flower Extract of Tabernaemontana divaricata and Assessment of Antibacterial Properties on Cotton Fabric

N. Nirmala Devi ¹*, P. Sumi ²

¹ Associate professor and Head, Department of Biochemistry and Biotechnology, Sree Narayana Guru College, K. G. Chavadi, Coimbatore.

² Assistant professor, Department of Biochemistry and Biotechnology, Sree Narayana Guru College, K. G. Chavadi, Coimbatore.

 

 

 

1. INTRODUCTION

Medicinal plants have long been recognized as valuable sources of bioactive compounds and have been extensively utilized in traditional medicine for the prevention and treatment of various diseases ^1,2. The growing demand for eco-friendly and sustainable technologies has led to increased interest in the green synthesis of nanoparticles using plant extracts. Compared with conventional physical and chemical methods, green synthesis offers several advantages, including low cost, environmental compatibility, reduced toxicity, and enhanced biocompatibility ^3,4.

Among different metallic nanoparticles, copper nanoparticles (CuNPs) have attracted considerable attention due to their remarkable antimicrobial, antioxidant, catalytic, and biomedical properties ^5,6. In addition, copper is more economical and readily available than noble metals such as silver and gold, making it suitable for large-scale industrial and medical applications ⁷. Tabernaemontana divaricata, a medicinal plant belonging to the Apocynaceae family, is rich in phytochemicals such as alkaloids, flavonoids, tannins, terpenoids, and glycosides, which can serve as reducing and stabilizing agents in nanoparticle synthesis ^8,9.

The present study focuses on the biogenic synthesis of copper nanoparticles using the hydroethanolic flower extract of T. divaricata and their characterization through UV–Visible spectroscopy, FTIR, SEM, and XRD analyses. Furthermore, the antioxidant activity and antibacterial efficacy of CuNP-treated cotton fabrics against pathogenic bacteria were evaluated to explore their potential applications in healthcare and antimicrobial textile products ^10,11.

Fig 1 : Tabernaemontana divaricata

2. METHODS AND MATERIALS

2.1 Plant Material Collection

Fresh flowers of Tabernaemontana divaricata were collected from Coimbatore, Tamil Nadu, India. The plant material was authenticated and washed thoroughly with distilled water to remove adhering impurities. The flowers were shade-dried at room temperature and pulverized into fine powder for further experimental studies ^12,13.

2.2 Preparation of Hydroethanolic Flower Extract

The dried flower powder was extracted using hydroethanol (70:30 ethanol) by maceration method. The extract was filtered through Whatman No.1 filter paper and concentrated under reduced pressure. The concentrated extract was stored at 4°C until use for nanoparticle synthesis and phytochemical analysis ^14,15.

2.3 Green Synthesis of Copper Nanoparticles

Copper nanoparticles (CuNPs) were synthesized using the hydroethanolic flower extract as a reducing and stabilizing agent. Copper nitrate solution was mixed with the extract under continuous stirring. The formation of CuNPs was visually confirmed by the color change of the reaction mixture from pale brown to bluish-green, indicating the reduction of Cu²⁺ ions into copper nanoparticles ^16,17.

2.4 Characterization of Copper Nanoparticles

The synthesized nanoparticles were characterized using different analytical techniques. UV–Visible spectroscopy was employed to monitor surface plasmon resonance and nanoparticle formation. Fourier Transform Infrared Spectroscopy (FTIR) was used to identify the functional groups involved in nanoparticle stabilization. Scanning Electron Microscopy (SEM) was performed to determine particle morphology and size, while X-Ray Diffraction (XRD) analysis confirmed the crystalline nature of the synthesized CuNPs ^18,19.

 

2.5 Phytochemical Analysis

Qualitative phytochemical screening of the hydroethanolic flower extract was carried out using standard protocols to detect carbohydrates, proteins, alkaloids, flavonoids, tannins, terpenoids, glycosides, saponins, and starch. These phytoconstituents play an important role in nanoparticle synthesis and biological activity ^20,21.

2.6 Antioxidant Assays

2.6.1 DPPH Radical Scavenging Assay

The free radical scavenging activity of the synthesized CuNPs was determined using the DPPH assay. The reduction in absorbance at 517 nm was measured and expressed as percentage inhibition ²².

2.6.2 Ferric Reducing Antioxidant Power (FRAP) Assay

The reducing power of the nanoparticles was evaluated by measuring the conversion of ferric ions (Fe³⁺) to ferrous ions (Fe²⁺), which reflects antioxidant capacity ²³.

2.6.3 Hydrogen Peroxide Scavenging Assay

Hydrogen peroxide scavenging activity was determined by assessing the ability of the synthesized nanoparticles to neutralize H₂O₂ radicals under in vitro conditions ²⁴.

2.7 Fabric Treatment

2.7.1 Scouring

Cotton fabrics were scoured using a standard detergent solution to remove impurities, oils, and waxes present on the fabric surface before nanoparticle treatment ²⁵.

2.7.2 Dip-Coating Method

The scoured cotton fabrics were immersed in varying concentrations of CuNP suspension for 24 h. The treated fabrics were washed with distilled water and dried in a hot-air oven at 37°C for 10 min to ensure proper fixation of nanoparticles onto the fabric surface ^25,26.

2.8 Wash Durability Test

The durability of the CuNP coating on cotton fabric was evaluated through repeated washing cycles using detergent solution. The antibacterial activity of the treated fabric was assessed after each wash cycle to determine coating stability ²⁶.

2.9 Antibacterial Activity

The antibacterial efficacy of untreated and CuNP-treated cotton fabrics was evaluated against Escherichia coli and Bacillus subtilis using the agar diffusion method. The inoculated Mueller–Hinton Agar plates were incubated at 37°C for 24 h, and the zones of inhibition were measured to determine antibacterial effectiveness ²⁷.

3. RESULTS AND DISCUSSION

3.1 SYNTHESIS OF COPPER NANOPARTICLES

The aqueous extract of Tabernaemontana divaricata was initially pale brown in color before the addition of the copper nitrate solution. Upon mixing with the copper nitrate solution, the reaction mixture gradually changed to a bluish-green color, indicating the formation of copper nanoparticles. This visible color change suggests the reduction of copper ions by the bioactive compounds present in the plant extract and serves as a preliminary confirmation of nanoparticle synthesis. The phytochemicals in the extract act as both reducing and stabilizing agents during the green synthesis process. thesis of Copper Nanoparticles ²⁸.

3.2 Characterization of Copper Nanoparticles 

3.2.1 UV Visible Spectral analysis

The UV–Visible spectrum of the synthesized copper nanoparticles exhibited a characteristic absorption band, which is attributed to the surface plasmon resonance (SPR) phenomenon of copper nanoparticles. The SPR absorption arises from the collective oscillation of conduction electrons on the nanoparticle surface when exposed to incident light. The position and intensity of this absorption band are influenced by the size and distribution of the nanoparticles, with changes in particle size leading to corresponding shifts in the SPR peak. Thus, the observed absorption band provides preliminary evidence for the successful formation of copper nanoparticles ²⁹

 

Figure 2: UV VISIBLE SPECTRAL ANALYSIS

3.2.2 Fourier Transform Infrared Spectroscopy (FTIR)

 

The FTIR spectrum of the synthesized nanoparticles showed a characteristic peak at 3127.36 cm⁻¹, indicating the presence of hydroxyl (–OH) functional groups ³⁰. 

 

Figure 3: Fourier Transform Infrared Spectroscopy (FTIR)

 

3.2.3 SEM Analysis (Scanning Electron Microscope)

SEM analysis revealed the successful formation of copper nanoparticles synthesized using T. divaricata extract. The micrographs showed aggregated clusters of nanoparticles with varying particle sizes, predominantly exhibiting a spherical morphology ³¹.

Figure 4: SEM Analysis(Scanning Electron Microscope)

3.2.4 X RAY Diffraction

X-ray diffraction (XRD) analysis further confirmed the successful synthesis of copper nanoparticles ³².

Figure 4: X RAY Diffraction

3.3 Phytochemical Analysis 

Preliminary phytochemical screening of the Tabernaemontana divaricata leaf extract revealed the presence of several important bioactive constituents. Qualitative analysis showed positive results for carbohydrates (Molisch test), proteins (Ninhydrin test), alkaloids (Wagner’s test), flavonoids (Lead acetate test), tannins (Ferric chloride test), terpenoids (Salkowski test), glycosides (Keller–Killiani test), saponins, and starch (Iodine test). The presence of these phytochemicals suggests that the plant is a rich source of secondary metabolites that may contribute to its reported medicinal properties, including antioxidant, antimicrobial, and anti-inflammatory activities ³³

3.4 Antioxidant Study

3.4.1 DPPH radical scavenging Assay

The antioxidant activity of the synthesized sample was evaluated using the DPPH radical scavenging assay. The results demonstrated a concentration-dependent increase in radical scavenging activity, indicating the presence of significant antioxidant potential. As the concentration increased from 200 to 1000 µg/mL, the percentage of DPPH radical inhibition also increased, suggesting an enhanced ability to donate electrons or hydrogen atoms to neutralize free radicals. This trend confirms the antioxidant properties of the sample ³⁴. 

 

3.4.2 Ferric Reducing/ Antioxidant Power Assay

The ferric reducing antioxidant power (FRAP) assay demonstrated a concentration-dependent increase in reducing activity for the sample extract. As the concentration increased from 200 to 1000 µg/mL, the reducing power of the sample increased progressively from approximately 18% to 48%, whereas the standard exhibited higher activity ranging from about 40% to 85%. The results indicate that the sample possesses significant electron-donating ability and antioxidant potential by reducing Fe³⁺ to Fe²⁺ ions. Although the antioxidant activity of the sample was lower than that of the standard, a steady increase in reducing power with increasing concentration confirms its dose-dependent antioxidant efficacy ³⁵. 

3.4.3 Hydrogen Peroxide Scavenging Assay

The hydrogen peroxide scavenging assay revealed a concentration-dependent increase in the free radical scavenging activity of the sample extract. The scavenging activity increased from approximately 4% at 200 µg/mL to 36% at 1000 µg/mL, while the standard showed higher inhibition ranging from about 9% to 47% over the same concentration range. The gradual increase in H₂O₂ scavenging activity indicates the ability of the extract to neutralize hydrogen peroxide and prevent the formation of highly reactive hydroxyl radicals. Although the activity was lower than that of the standard, the results confirm that the extract possesses notable antioxidant potential in a dose-dependent manner ³⁶.

3.5 Antibacterial Assay

The antibacterial activity of copper nanoparticle-coated cotton fabric was evaluated against Escherichia coli and Bacillus subtilis using the disc diffusion method. After incubation at 37°C for 24 h, clear zones of inhibition were observed around the treated fabric samples, indicating effective antibacterial activity. The test organisms exhibited sensitivity towards all treated samples (WT, UT, and C), while the positive control (1 mg/mL) showed the highest inhibitory effect. The results demonstrate that the copper nanoparticle-treated cotton fabric possesses significant antibacterial activity against both Gram-negative (E. coli) and Gram-positive (B. subtilis) bacteria ³⁷.

3.5.1 The wash Durability test 

The wash durability test was performed to evaluate the retention of antibacterial activity of the Tabernaemontana divaricata-mediated copper nanoparticle-coated cotton fabric after detergent washing. Even after the washing treatment, the coated fabric exhibited noticeable zones of inhibition against both Escherichia coli and Bacillus subtilis, indicating that the antibacterial coating remained effective. The results demonstrate the good durability and wash-fastness of the antimicrobial finish, confirming that the T. divaricata extract-mediated coating possesses sustained antibacterial activity against both Gram-negative and Gram-positive bacteria ³⁸.

4. CONCLUSION

In conclusion, the present study successfully demonstrated the green synthesis of copper nanoparticles using the flower extract of Tabernaemontana divaricata. The formation and stability of the nanoparticles were confirmed through UV–Visible spectroscopy, while FTIR, SEM, and XRD analyses verified their functional groups, morphology, and crystalline nature. The synthesized copper nanoparticles exhibited significant antibacterial activity against Escherichia coli and Bacillus subtilis when applied to cotton fabric. Furthermore, the coated fabric retained its antimicrobial efficacy after washing, indicating good durability. These findings suggest that the nanocomposite cotton fabric has promising applications in medical textiles and healthcare products. 

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