Formulation And Evaluation of Biogenic Synthesis of Silver Nanoparticle Containing Diphysa Americana Extract and Its Antimicrobial Activity

Abstract

The biogenic synthesis of silver nanoparticles (AgNPs) has emerged as a sustainable and eco-friendly alternative to conventional physical and chemical methods. This research explores various biological entities, including plants, bacteria, fungi, and algae, as sources for synthesizing AgNPs. The biogenic process offers several advantages, such as low toxicity, cost-effectiveness, and enhanced biocompatibility, making it highly suitable for medical, pharmaceutical, agricultural, and environmental applications. The mechanism of AgNP biosynthesis involves the reduction of silver ions (Ag?) to metallic silver (Ag?) through bioactive compounds, followed by nanoparticle nucleation and stabilization. Using UV-Vis spectroscopy, particle size analysis, scanning electron microscopy (SEM), dynamic light scattering (DLS) particle size, and SEM image analysis, the nanoparticles were thoroughly characterized. The average particle size was determined to be in the range of less than 1000 nm Ag. Antibacterial potential of Ag nanoparticles as a function of nanoparticles concentration was tested against four different bacteria like Escherichia coli and S. aureus. The test was performed by both Disc diffusion assay and colony forming unit (CFU) estimation method. From the study, both types of nanoparticles were observed to have strong antimicrobial potential

Keywords

silver nanoparticles, biogenic synthesis, nanotechnology, green chemistry, antimicrobial, diphysa americana, antimicrobial activity

Introduction

“Nanotechnology” is the newest and one of the most promising and active areas of modern research. The technology deals with the design, synthesis, and manipulation of particles size ranging from 1–1000 nm. Within this size range, the chemical, physical, and biological properties change in the fundamental way of both individual atoms and their corresponding bulk material. This very small size increases the surface area-tovolume ratios of particles. Recent years have seen a significant increase in interest in plant extract-based nanoparticles because of their exceptional qualities and diverse range of use in catalysis, plasmonic, optoelectronics, biological sensor, water treatment, pharmaceutical applications, and agriculture and crop protection. The exponential advancement of this nascent technology has unlocked new theoretical and practical vistas, such as the creation of nanomaterials and the application of their optoelectronic and physicochemical characteristics. Many fields, including optics, mechanics, chemical and space industries, electronics, energy science, single-electron transistors, light emitters, nonlinear optical devices, photo-electrochemistry, catalysis, biomedical, cosmetics, drug and gene delivery, food and feed, and photo-electrochemicals, have seen an increase in the use of nanotechnology

Concept of nanotechnology

Nanoscience and technology are an exciting and rapidly emerging branch of science and technology which work at the atomic, molecular and macromolecular levels. Nanotechnology deals with manipulation and uses various tools and functional materials at nanoscale.

While biotechnology has revolutionized nearly every field in veterinary and animal sciences by producing new, small-scale tools and materials that are advantageous to living things, nanotechnology has opened up new avenues for applications in molecular biology. The different type’s nanomaterials are being utilized for disease diagnosis, treatment, drug delivery, animal nutrition, and animal breeding. The various types of nanoparticles are used such as metallicnanoparticles, quantum dots, carbon nanotubes, magnetic nanoparticles, fullerenes, liposomes and dendrimers.

A Nanoparticle or ultrafine particle is a particle of matter 1 to 100 nanometres (nm) in diameter. The term is sometimes used for larger particles, up to 500 nm, or fibers and tubes that are less than 100 nm in only two directions. ^? At the lowest range, metal particles smaller than 1 nm are usually called atom clusters instead.

Nanoparticles are distinguished from microparticles (1-1000 μm), "fine particles" (sized between 100 and 2500 nm), and "coarse particles" (ranging from 2500 to 10,000 nm), because their smaller size drives very different physical or chemical properties, like colloidal properties and ultrafast optical effects or electric properties.

Silver Nanoparticles

Agnps Are Among The Most Promising Items In The Nanotechnology Business Among The Different Metallic Nanoparticles. The Creation Of Reliable Procedures For Agnp Synthesis Is A Key Area Of Contemporary Nanotechnology Research. Agnps' Special Optical, Electrical, And Magnetic Properties Make Them Useful In A Variety Of Applications, Including Antibacterial, Antiviral, And Antifungal Ones; They Can Also Be Used In Composite Fibers, Biosensor Materials, Cosmetics, The Food Industry, And Electronic Components.Agnps Have Also Been Identified As Pharmaceutical And Medicinal Substances That Have Come Into Direct Contact With Human Tissue In Products Like Toothpaste, Shampoos, Detergents, Soaps, And Cosmetics. The Biomedical Utilizes Agnps Includes Their Application As Antibacterial, Antifungal, Anti-Inflammatory,Antiviral, And Anti-Diabetic Agents.Agnps Have Also Been Reported In Recent Research To Be Used In The Detection And Treatment Of Cancer As Well As As Active Or Passive Medication Carriers

APPLICATIONS

1. Antibacterial Activity

Either killing or reducing the growth of bacteria without affecting surrounding cells is known as antibacterial activity. Ag is preferred as nanoparticle for the reason that it has antibacterial property and non-toxic to human beings. AgNPs are able to overcome the resistance that has been due to antibiotics.

Antiviral Activity

In the whole world, viral infections and disease are found to be very common, so it’s very important to make antiviral agents that results in showing prominent results. AgNPs are found to be prominent in showing such results this is due to their very small size and their shape also. It has been observed that silver is found to be relatively non-toxic towards humans as well as animals and found to be effective against viruses.

Antifungal Activity

Persons having less immunity are more prone to fungal infections. To overcome the fungus related diseases, it is found that this process is found to be very tedious in nature. There are very limited numbers of antiviral drugs that are available in the market.Anti-viral drugs should be biocompatible, non-toxic as well as environmentally friendly. AgNPs are found to be prominent against many diseases that are caused due to fungi.

2. Dental materials

The oral cavity is an active ecosystem. Microorganisms, particularly pathogenic ones, frequently colonize the mouth. This increases the danger of colonization processes and contaminates dental materials and implants. Silver-based nanostructures are frequently applied to dental materials in order to alter or embed them. Typically, fluorinated diamine silver has positive effects on preventing dental cavities. AgNPs' antibacterial properties improve when their size decreases because their specific surface area increases with decreasing AgNP size. The addition of silver-based nanosystems to adhesive resins, orthodontic cements, and dental composites has been reported to be effective.

3.  Anti-inflammatory Activity

Inflammation is the state in which some part of the body becomes swollen, red, hot and sometime painful also and this may occur due to certain injury or sometime infection also. Inflammation is also found to give an immunological response that is against some foreign particles. As AgNPs are known to be their antibacterial and anti-microbial activities but their response to act as anti-inflammatory reagent are limited but they also play important role in this anti-inflammatory field.

4. Anticancer Activity

Cancer is basically an uncontrolled growth of cells in specific area in a body.  Several studies have been done to know the promising result of AgNPs. It is found to be most suitable as well as an alternative for other cancer treatments. They have ability to target specific cells or tumour at that site only by encapsulation of therapeutic agent in nanoparticle and then used as drug delivery system

AIM:

Formulation and evaluation of biogenic synthesis of silver nanoparticle containing Diphysa americana extract and its antimicrobial activity.

OBJECTIVE:

• The present study for the first time utilized extract of Diphysa americana for reduction of 1 mM silver nitrate solution to silver nanoparticles.
• The method proved to be very simple, cost-efficient, and convenient.
• Synthesis of nanoparticles was confirmed by visual detection and UV- spectral analysis in which the colourless solution gets changed to a brown-colored solution.

Further characterization was done by particle size, zeta potential and SEM analysis.

PLANT PROFILE

Diphysa Americana

Figure 1: Diphysa americana

Botanical classification

Kingdom: Plantae

Subkingdom: Tracheobionta

Division: Magnoliophyta

Class: Magnoliopssida

Sub-Class: Rosidae

Order: Fabales

Family: Faboideace

Genus: Diphysa

Species: Diphysa americana

Therapeutic uses

The leaves, stems, roots, flowers and seeds of D. americana regardless the subspecies have been used for the treatment of several illnesses including mostly malaria, a tropical endemic disease with high morbimortality. According to the ethnic differences of populations from localities, the plant is used alone or in combination with other plants or with natural substances for the preparation, especially in decoction. For the treatment, people mostly used the preparations by oral administration route.

PHARMACOLOGICAL STUDIES

•     Ant malarial activity

•     Antidiabetic and anti-lipemic effects

•     Antioxidant activity

•     Antitumor or anticancer activit

•     Anti-inflammatory activity

•     Antibacterial activity

•     Antifungal activity

•     Toxicology

ANTIBACTERIAL ACTIVITY OF AG NANOPARTICLESBY WELL DIFFUSION ASSAY

Preparation of Nutrient Agar Media

28 g of Nutrient Media was dissolved in 1 litre of distilled water. Ph of media was checked before sterilization. Media was sterilized in autoclave at 121^oc at 15 lbs pressure for 15 minutes. Nutrient media was poured into plates and placed in the laminar air flow until the agar was get solidified.

Well Diffusion Assay

The bacterial suspension of E. Coliand S. Aureuswas standardized to 10⁸ CFU/ml of bacteria and kept into the shaker. Then, 100µl of the inoculums from the broth (containing 10⁸ CFU/ml) was taken with a micropipette and then transferred to fresh and sterile solidified Agar Media Plate (Mohammadi-Sichani et al., 2012). The agar plate was inoculated by spreading the inoculums with a sterile spreader, over the entire sterile agar surface. Three wells of 6 mm were bored in the inoculated media with the help of sterile cork-borer. The wells were then formed for the inoculation of the agno3, agnps and extract (1mg/ml) solution. 100 µl of the sample was loaded. It was allowed to diffuse for about 30 minutes at room temperature and incubated for 18-24 hours at 37^o C.  Following incubation, plates were checked to see if a clear zone formed around the well, indicating that the chemicals under test had antimicrobial activity. A measurement of the zone of inhibition (ZOI) in millimeters was made.Zones were measured to a nearest millimeter using a ruler, which was held on the back of the inverted Petri plate. A black, non-reflective background was held a few inches above the Petri plate. The diameters of the zone of complete inhibition, as determined by the unassisted eye, were measured, encompassing the well's diameter.

RESULTS AND DISCUSSION

Percentage yield

Table 1: Percentage yield of extracts

Table : Organoleptic properties of Diphysa americana

Table:  Solubility study of Diphysa americana

Evaluation parameter of Silver nanoparticle

1. Color Observation

 

UV-Visible spectrophotometric analysis

Table 2: UV peak detection

Discussion

Table 3: Particle size of Silver nanoparticle

Table 4: Zeta potential

Figure 2: Antimicrobial activity against E. coli

1. 1. 2. Antimicrobial activity of Ag Nanoparticle of against S. aureus

Table 6: Antimicrobial activity of Ag Nanoparticle against S. aureus

S N	Sample name	Zone of Inhibition (mm)
1	AgNO3	8mm
2	Extract	15mm
3	Silver NPs	18mm

 

Figure 3: Antimicrobial activity against S. aureus

CONCLUSION

The conclusion drawn from the present investigations are as follows:

Ag nanoparticles were synthesized successfully by green synthesis methods (From Diphysa americanaextract), respectively. Using UV-Vis spectroscopy, particle size analysis, scanning electron microscopy (SEM), dynamic light scattering (DLS) particle size, and SEM image analysis, the nanoparticles were thoroughly characterized. The average particle size was determined to be in the range of less than 1000 nm Ag. Antibacterial potential of Ag nanoparticles as a function of nanoparticles concentration was tested against four different bacteria like Escherichia coli and S. aureus. The test was performed by both Disc diffusion assay and colony forming unit (CFU) estimation method. From the study, both types of nanoparticles were observed to have strong antimicrobial potential.

The growth study of Escherichia coliand S. aureuswas carried out in presence of different concentration of both nanoparticles to observe the effect on the growth of the bacteria in liquid media. It was observed that both the nanoparticles strongly affected the specific growth rate of E.coli and S. aureus. It was also observed that the growth rate was strongly inhibited by the presence of small concentration of nanoparticles

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