Conventional synthesis of nanoparticles chemically releases toxic byproducts so there was a sudden shift towards the synthesis of nanoparticles through biological methods as it was assumed that biosynthesized nanoparticles would not be toxic. The present study aims at comparative toxicity analysis of chemically and biologically synthesized copper oxide and silver nanoparticles. Biologically synthesized copper oxide and silver nanoparticles were subjected to antimicrobial activity against five bacterial species at different dilution rate (20%, 50%, 70% and 100%). The result obtained for this was that for silver nanoparticles synthesized chemically E.coli was susceptible at 20% and 50% dilution rate. At 70% dilution rate for copper oxide nanoparticles synthesized chemically two bacterial species i.e. E.aerogens and P.aeroginosa were highly susceptible but other three species were susceptible.
ABSTRACT
Conventional synthesis of nanoparticles chemically releases toxic byproducts so there was a sudden shift towards synthesis of nanoparticles through biological methods as it was assumed that biosynthesized nanoparticles would not be toxic. The present study aims at comparative toxicity analysis of chemically and biologically synthesized copper oxide and silver nanoparticles. Biologically synthesized (S.indica) copper oxide and silver nanoparticles were subjected to antimicrobial activity against five bacterial species i.e. E.coli, E.aerogens, S.epidermidis,P.aerogens and M.luteus at different dilution rate (20%, 50%, 70% and 100%). The result obtained for this was that for silver nanoparticles synthesized chemically E.coli was susceptible at 20% and 50% dilution rate. At 70% dilution rate for copper oxide nanoparticles synthesized chemically two bacterial species i.e. E.aerogens and P.aeroginosa were highly susceptible but other three species were susceptible. For both biologically synthesized nanoparticles it was observed that at all dilutions S.epidermidis was completely susceptible. Synthesized nanoparticles were characterized with UV-Vis spectroscopy and TEM analysis. Formation of silver (chemically) at 400nm, copper (chemically) at 380nm, copper (biologically) 395nm and silver (biologically) at 390nm plasmon absorption was revealed by UV-Visible spectroscopy respectively. Different sizes of synthesized nanoparticles in different ranges and shapes were analyzed through TEM. This analysis showed copper oxide nanoparticle of sizes between 25nm- 207nm, shapes (rod, triangular, spherical and round) synthesized chemically. Also, copper oxide nanoparticle of sizes between 12nm-172nm, shapes (rod, oval and round) synthesized from leaves of S.indica. The further analysis of silver nanoparticle through TEM showed sizes between 10nm-72nm, shape (oval) for synthesized chemically. Even TEM showed silver nanoparticle of sizes between 9nm-59nm, shape (oval, triangular) for those synthesized from leaves of S.indica (Ashoka tree). Our results confirmed that copper oxide and silver nanoparticles synthesized biologically were less toxic in comparisonto chemicallysynthesized.
INTRODUCTION
Nanotechnology can be explained as a science which deals with manipulating or creating materials at nanoscale level whose size lies in range of 1 to 100nm. Furthermore, nanotechnology can be said as study and compression of matter with dimensions between approximately 1to100 nanometers, thus enabling its use in different kind of applications for us” [National Nanotechnology Initiative (NNI) 2008]. In relation to background researches and recent current practices, related to nanotechnology, highlights issues surrounding the nanotechnology and its use like in areas- environmental remediation and even for future directions. Nanotechnology utilizes several different approaches which include starting at atomic level scale and then building structures and materials by specifically modifying or placing atom by atom using the bottom-up approach of nanotechnology.Thus, with this approach, forces of chemistry are in control and are less flexible in marking unrestrained structures like number of products for consumer and we can expect significant products in coming future. Another approach known as „top down" processing includes breaking down of something into smaller and smaller segments. This approach can be made use in understanding the different biological processes going in one"s body for example how the brain leads to transfer of sensory information. With the use of nanoscience and nanotechnology, novel researches are in process in different fields like in industrial sectors. Many areas are even making use of nanomaterials due to their beneficial properties for example in use of catalyst and as antibacterial coatings. Even nanotechology is being used as in environment technology to decrease pollution, cleaning, and treating of products that causes disruption of natural environment. Also, this technology can help in practicing site remediation. Though detailed study risks resulting from different remedies and approaches are poorly understood
Biotechnology and Nanobiotechnology are two most positive technologies of this century. Nanotechnology give us the idea of technique that helps in designing, developing and applying of such devices and materials of which least functional make up is a nanometer scale (Emerich et al, 2003;Sahoo et al, 2003). Nanobiotechnology an amalgation of biotechnology can leads to a novel development and implementing the ideas to make into useful tools for study of life. This idea draws the attention towards application of field of science such as devices molecular biology, surface science, semiconductor physics, organic chemistry, microfabricaton, etc Nanoparticles (inorganic materials) are basically the particles whose range lies at 1to100 nanometers in size. Synthesis of nanoparticles has gained importance in contemporary because of their physical and chemical properties for example nanoparticles made from silver are made use into vast range of application such as in cosmetics, wound healing therapeutics, soaps, bioremediation , electronic devices, bio imaging, (Armendariz et al, 2002; Kyriacou et al, 2004 Kim et al, 2010), antifungal and antibacterial (Rai et 2009; Sharma et al , 2009;).
Nanoparticles synthesis can be achieved by chemical method such as sol gel method, precipitation method, polyol method, hydrothermal synthesis or via biological synthesis through plants(Shankeer et al, 2004 and Ahmad et al , 2011),fungus (Vigneshwaram et al , 2007), and bacteria(Klaus et al, 1999 and Konish et al , 2007) , enzymes (Willner et al , 2006)
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Fig 1. Potential applications of NPs in differentfields
Silver nanoparticles-
Silver is a white transition metal it possess high thermal and electrical conductivity making it suitable for utilization in various electrical devices. Its nanoparticles are gaining large attention even in present days due to their biological, physical and chemical properties thus making its way towards many beneficial applications in nanobiotechnology (Rai et al , 2007). That include medical devices for disinfection purpose and water treatment, home appliances.(Bosetti et al,2002; Cho et al 2005; Gupta and Silver 1998; Jain and Pradeep,2005;Li et al,2008)
Copper oxide nanoparticles-
Copper oxides are can be applied different industrial sectors such as manufacturing processes of semiconductor devices, microelectronics, solar energy convertor, batteries, gas sensor and industrial catalyst ( Jha et al, 2010;Bar et al 2009). In spite of its various use in bulk, use of copper nanoparticles is restricted in atmospheric condition as it is prone to oxidation. Its applications include catalytic organic transformations, electrolysis and photolysis. Another property of copper i.e. its elevated temperature makes it to exist in very hot temperature and pressure, chemical reactions and various organic transformations. Copper oxide even possess antimicrobial properties thus its nanoparticles are made to be use in formation food containers, paints, plastics and textiles (Baban et al, 1998;Tomar; Garg , 2013). Humans and aquatic organisms such as ciliated protozoa, fresh water shredders are highly prone to copper nanoparticles toxicity. (Ceylan et al , 2006; Junggwon et al, 2008;Mohl et al, 2011 ; Fan et al, 2009).
Nanoparticles asAntimicrobials
Nanotechnology and nanoparticles being advantageous in nature are making its path in treating bacterial infections as nanoparticles are now days, in demand because they target bacterial cells as a substitute of antibiotics. Implantable devices require antibacterial coating on their surface so that it doesn"t create any infection in the body and promote healing of wounds. For this nanoparticles made from silver are utilized for this purpose because they possess antibacterial activity. An advantage with using nanoparticles as antibacterial is that those bacteria that are resistant to many drugs are also susceptible towards nanoparticles bactericidal activity as nanoparticle act through different unknown mechanisms to kill bacteria and development of resistant pathways through nanoparticles is difficult.
Nanoparticles and its potentialhazards
Nanotechnology is distinctive technique due to its innovative manufacturing ideas it offers. Due to high level of demand leading to more production and use of nanoscale particles there has been a growing concern over the impact of this new technology on organisms health & safety and to the environment even (Steinfeld et al, 2004; Swiss Re 2004). Several researchers have reported the potential risk of nanoparticle or materials made from nanotechnology on aquatic organisms, human health and environment (Daar et al, 2012; Zhoq et al , 2012; Ameen et al, 2013). With its use it should be remember that the assessment would be on the basis of known material and energy data and this would not allow any estimate to be made concerning products still under development.
Hazards to human
a Toxicokinetics -Fundamentally, toxicokinetics can be stated as movement of particle through which it may get into the body, how would it be circulated and gets distributed within it and how it performs its metabolic functions and finally how it got excreted. Studying this complete mechanism is important as it allows reflection of the targeted organs that could be or could not be affected, even to foretell in advance the exposure of realistic doses and to note down in what way the body would respond to nanoparticles on being exposed. Till now, only few, studies have been conducted for nanoparticles that how if any, adsorption, distribution, metabolism and excretion (ADME) happens (Kreuter and Jorg et. al, 2003).
b. In-vitro nanoparticle toxicity- Understanding of nanoparticle mechanism for toxicity observed on human cells was necessary to estimate the effect of nanoparticle on several parameters like inflammatory cytokine response, oxidative stress and apoptosis. In vitro, cell studies provide deep study of toxicity mechanism, response towards cellular defense mechanisms and possible pathogenic effects. This will help in the selection of relevant methods to know hazards it can cause. In relation to this we have noticed the role of played by particle size of nanoparticle, its movement, combining and breaking, its nature and magnitude of biochemical responses that would result in cellular fate (Lam and Chiu- Wing et al, 2004).
c. Skin, gut and other organs: As mentioned before, current managing of nanoparticles are making available of products for consumer and risk regarding on prior exposure to skin as nanoparticle can easily penetrate skin cells. The range of nanoparticles to mark its entry to the skin and cause harmful effects is not well understood. As we have a good evidence that how larger micro particles travel well into the gut that to in normal condition but similar evidence doesn"t exist that how a nanoparticle travels into the gut, thus, there is insufficient evidence that whether they act on the gut or not adversely (Derfus et al, 2004)
Environmental hazards
Many complex ecosystems which constitute fresh water, marine and terrestrial compartments including humans, there are several species that are potentially at threat of being exposed with nanoparticles at high level. Hazards related with nanoparticle exposure can create problems at an individual or population level and might lead to disruption at the structure and functioning level of organisms on the whole. Unless detailed researches are made it is difficult to say that which part of ecosystem (soils, surface, water and air) is more at risks with nanoparticles (Glor1988).
a. Effects on groundwaters and soils: A study is needed to estimate the risk related to environment and its behavior on being vulnerable to such nanoparticles in soils that includes its fauna and flora and even and groundwater. During microbe treatment while clearing of sewage with the use of nanoparticles, its toxicity needs to be maintained as particularly once sources of these to sewer had been observed (Chen et al, 1998).
bi Estimation of explosion and ignition , potential: This is the main area of concern because nanoparticles show possibility of causing hazards that needs to be studied in detail so as to estimate the explosive nature of nanoparticles. To analyze the effect of explosion and ignition potential of nanoparticles and the safety hazards associated with it in case of any miss happening. A study was performed in which nanoparticles were kept in open environment and two types of conditions were studied in first condition nanoparticles were placed near a ignition source and in second condition nanoaprticles were subjected to a ignition. A study made by the HSE (Health and Safety Executive) states that nanoparticles can create dust explosion hazard because they have large surface area and this might cause them to became flammable on getting exposed to air especially metallic nanoparticles as they easily gets oxidize (Eckhoff et al, 2004).
c Effects associated with cellular release: toxicokinetics and uptake of nanoparticles in vertebrates, invertebrates, plant and of cellular uptake, localization is not well understood yet. The effects at low doses and for longer period made to be studiedwell.
Understanding the particle physico-chemical properties and basic mechanisms of toxicity that clearly indicates appropriate endpoints for required tests used in hazard assessment is needed. Ecofriendly methods are in use for nanoparticle synthesis by researchers like using microorganisms for the synthesis of many metal nanoparticles (Klaus et al, 1999;Shahverdi et al, 2007; Konishi et al , 2007), syzygium cumini (Kumar et al, 2010), plant leaf extract of basil (Ahmad et al, 2010),onion (Saxena et al , 2010), syzygium cumini (Kumar et al, 2010). A medicinally valuable plant Saraca Indica of Family Caesalpinaceae is evergreen tree, medium in size (upto 9m in height) and has orange yellow colored flowers and that are in pattern of dense corymbs. It is geographically located in eastern and central Himalayas (Prajapati et al, 2003). The medicinal properties of this plant includes burning sensation, treatment of dyspepsia, colic, ulcers, pimples, fever, menorrhagia, anti-inflammatory, anti-diabetic, uterine tonic activity, CNS depressant, larvicidal, anthelmintic activity(Nayak et al , 2011;Bhadauria et al, 2012). Considering its importance, the present work is made to understand the antimicrobial activity against copper oxide and nanoparticles synthesized using aqueous leave extract of Saraca indica (Ashoka tree)
REVIEW OF LITERATURE
Nanotechnology can be explained as the branch of knowledge whose concern is basically with constituents and structual that explains the way in which its physical, chemical and biological phenomenon can be altered due to nanoscale size. Furthermore, it can be stated that with better understanding of this technology we can manipulate the matter at atomic, molecular and at superamolecular scale and thus creating nanoscale materials by involving its design, production and characterization and also applying them in different potential areas to have novel advances in technologies specially in field. Nanotechnology had provided a way that leads to therapy at a molecular and that can further leads to help in treatment and understanding cause of pathogenesis of disease. With this comes the limitations with conventional drugs especially with nonspecificity of drug action and thus this development of a nanomaterial system urgently so that it can be applied in diagnostic and to treat various diseases especially cancer (as it has much limitations of poor specificity, drug toxicities and sensitivity). For cancer treatment nanoparticles are being evaluated presently.They (nanostructed) is designed so as to be applied as fluorescent materials, contrast agents, drugs to target antibodies and for molecular research tools. With the advances made in this technology modifications have taken place in nanoparticulate systems like in paramagnetic nanoparticles, quantum dots, nanoshells are used for diagnosis of various ailments. Recent researches showed nanoparticles based theraphy that can help in delivery of drug at specific site and this can further help in integrating of image within nanoparticles related to contrast agents thus allowing visibility of drug delivery site to us and additionally examining of efficacy of the therapeutic agent, in vivo (Cai et al, 2007)
A paper had defined the process for silver nanoparticles synthesis by chemical method (Maribel et al, 2009). Atypical surface plasmon absorption maxima at 418-420 nm by UV-Visible spectroscopy confirmed the formation of silver nanoparticles. The better understanding of nanomaterials like bactericidal has increased the consideration of new strains of bacteria that are resistant to the strong antibiotics and thus promoting research towards the silver nanoparticles and activity of silver related compound. Nanoparticles bactericidal activity is dependent on several factors one of the most crucial factor is their size was confirmed by study carried out by Morones and its team mates(Morones et al, 2005). In a study nanoparticles of silver were tested against Escherichia coli, Staphylococcus aureus and yeast to test their antimicrobial activity. The results revealed that silver nanoparticles antimicrobial activity was because of free radical generation which was characterized by electron spin spectroscopy(Jun et al, 2014). This observation was utilized to manufacture medicinal implants with silver metallic nanoparticle coating for their antibacterial effect (Jun et al, 2014). Due to capability of nanotechnology of adjusting metals into nanosize with drastic changes in their optical, physical and chemical properties in present time because of this it is gaining large importance in present days. As pathogenic bacterial species are becoming resistant day by day there is a need to use silver nanoparticles (because its nanoparticles possess effective antimicrobial property) making it suitable for utilization in silver coated medicinal devices, nanolotions, silver based dressing and nanogels ( Rai et al, 2009).
A paper discussed the method for copper oxide nanoparticles synthesis using aqueous precipitation method (Kshirsagar, et al, 2017). Transition metal oxide based nanoparticles like nanoparticles of copper oxide which possess several industrial application such super capicitors, magnetic stirrer devices, sensors, in chemical plants. A paper stated copper oxide nanoparticles synthesis, its characterization by using thermal plasma technology and its use in antimicrobial applications (Guogang Ren et al, 2009). CuO nanoparticles suspension possesses antimicrobial activity against many drug resistant microorganisms such as Escherichia coli and a meticillinresistant Staphylococcus aureus (MRSA) having Minimum Bactericidal concentrations (MBCs) of these copper oxide nanoparticles. Nanomaterials have shown specific properties in comparison to their majority of equivalent parts and so nanotechnology has gained lot of attention from the investigators. Nanomaterials of metal oxide like CuO and ZnO have found their use at industrial level for different uses including plastics, cosmetics, textiles and paints. In a published paper, CuO, ZnO and Fe2O3nanoparticles antimicrobial activity against Gramnegative bacteria and Gram-positive bacteria was studied (Azam et al, 2017). The investigators have been attracted towards nanometer sized materials due to their unusual, interesting and unique chemical, biological and physical properties. Of all the transition metal oxides, copper oxide has gained much importance due to the properties like high critical temperature (Tc), superconductivity are much fascinating. A report highlights the process by which nanoparticle size can be controlled through unique gel combustion method to make it an effective antibacterial agent against Bacillus subtilis and Staphylococcus aureus (Ameer Azam et al, 2012). In a report the copper oxide nanoparticles antimicrobial activity was observed against Staphylococcus aureus, Micrococcus luteus, Escherichia coli, Pseudomonas aeruginosa and Klebsiella pneumoniae, fungus like Aspergillus niger, Candida albicans and Aspergillus flavus. 26mm in E.coli and 23 mm in C.albicans zone of inhibition thus showed that more inhibitory activity was in bacteria then fungus (Ramyadevi et al, 2012) .
Leaves extract of Saraca indica was used in silver nanoparticle synthesis and these nanoparticles were further carried for study of anthelmintic activity of colloidal solution as according to protocol followed by their characterization using UV-visible spectrophotometer, FTIR and TEM (Garga and Chandra, 2012). The nanoparticles shape was spherical with size up to 50 nm. Finally work concluded that aqueous extract anthelmintic activity was less than that of colloidal solution of silver nanoparticles thus indicating the concentration-dependent nature of colloidal solution and the aqueous extracts(Garga and Chandra, 2012).
In a report, the silver nanoparticle synthesis was done by biogenic method (AgNPs) with help of Saraca indica leaves extract and also used for characterization and observing their antimicrobial activity. Further focus was made on the AgNPs synthesis mechanism as the extract of plant leaves contains water soluble organic materials that reduce silver ions and causes stabilization of AgNPs. The UV-visible spectrum indicated nanoparticles formation at 412 nm and their size distribution profile by Dynamic Light Scattering (DLS) was analyzed. Transmission Electron
Microscopy (TEM) framed the AgNPs size that was in between 13-50 nm and shape of particles was spherical. X-ray Diffraction (XRD) pattern of the AgNPs exhibited 2θ values of silver nanocrystals formation (Triphati et al, 2013). Synthesized AgNPs antimicrobial activity against E. coli DH5a was studied zone of inhibition was analysis and growth curve. Moreover, biogenic AgNPs at concentration 25 μg/ml were recorded as minimal inhibitory concentration (MIC) against E. coli DH5a (Triphati et al, 2013). Being eco-friendly in nature; nanoparticles of silver can be applied in many fields for betterment and benefits of the people. In a work nanoparticles of silver were synthesized by green approach i.e. leaves extract of Saraca indica plant (Perugu et al, 2012). Biofabrication of nanoparticles from eco-friendly sources such as plants, bacteria, fungi have gained worldwide researchers attention due to low toxicity, easy method of preparations and cost effective nature of synthesis. A paper stated silver nanoparticles (AgNPs) synthesis by green method i.e. using plant sources. In which environment friendly approach was made that needs to further explore for the prospects of many plants so that they can be used in nanoparticle synthesis. The size of AgNPs nanoparticle was in between 1-100 nm. It was also found that AgNPs green synthesis can be efficiently carried out for future like in medical concerns because they possess high prospects to be used as antimicrobial agent. Thus the above study provides a detailed knowledge on synthesis of plant mediated AgNPs with specific focus on their uses, e.g. anticancer, antioxidant, antimicrobial activities (Chung et al , 2016).
In a paper, nanoparticles of silver were synthesized by using extract of Allium cepa (onion) with the help of trisoduim citrate and silver nitrate (precursor). Characterization of these nanoaprticles was done using UV-Visible absorption spectroscopy. Furthermore, toxicity analysis of silver nanoparticles was done by ToxTrak test. In this, 24hours old cultures (Bacillus subtilis) and dye (resazurin) were used to analyze percentage inhibition (PI). Percentage inhibition works on the principle of relatively measuring or analyzing increase in toxic substances with respect to increase in respiration resulting in a negative number. For comparative toxicity analysis of chemically synthesized and biologically synthesized nanoparticles percentage inhibition of both samples was calculated.As a result it was noted that the toxicity value or PI of silver nanoparticles synthesized biologically(from onion) was less (51.39%) than sliver
nanoparticles synthesized chemically (85.45%) . Thus it was clearly noted from observed result that B. subtilis was not killed by sliver nanoparticles synthesized biologically as compared to silver nanoparticles synthesized chemically (Tyagi et al, 2013). A study was performed in which nanoparticles of copper oxide were synthesized of by green synthetic approach of different sizes (Prasad et al, 2017). Leaves extract of Saraca indica made it possible to reduce copper oxide into copper ions directly and thus producing nanoparticles of different morphology and size. Nanoparticles synthesized were characterized using High-Resolution Transmission Electron Microscopy (HRTEM), Fourier Transform-Infrared spectroscopy (FT-IR), X-ray diffraction (XRD), Energy Dispersive X-ray spectroscopy (EDX), Ultraviolet- Visible spectral studies (UV- Vis) X-ray photoelectron spectroscopy (XPS) and Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM) and Energy Dispersive X-ray spectroscopy (EDX) to analyze their shapes and size. The images from SEM stated that the particles were of average diameter 40-70 nm and spherical in shape. Also the HRTEM and TEM images indicated the spherical and crystalline nature.
AIM AND OBJECTIVE
During synthesis of nanoparticles some chemicals get absorbed on their surface, causing adverse effects when applied in medical applications. This why, enormous findings by researches in synthesizing nanoparticles by green approach is constantly been done. So the aim of this work to estimate the effectiveness of synthesized nanoparticles from leaves of Saraca indica (Ashok tree) on five pathogenic bacterial species respectively and to measure the toxicity of synthesized nanoparticles(chemically and biologically).
a. Synthesis of biological and chemical nanoparticles (silver and copperoxide).
b. Purification of synthesized nanoparticles
c. Characterization of purified nanoparticles by using different characterization tools like UV-VIS spectrophotometer, Transmission Electron Microscopy (TEM), Fourier- transform infrared spectroscopy ( FTIR) etc.
d. Testing the effectiveness of chemically synthesized Silver nanoparticles on agarplates against various pathogenic species to estimate their antimicrobialactivity
e. Testing the effectiveness of biologically synthesized Silver nanoparticles on agarplates against various pathogenic species to estimate their antimicrobialactivity
f. Testing the effectiveness of chemically synthesized Copper oxide nanoparticles onagar plates against various pathogenic species to estimate their antimicrobialactivity
g. Testing the effectiveness of biologically synthesized Copper oxide nanoparticles onagar plates against various pathogenic species to estimate their antimicrobial activity
h. Estimation of toxicity effect of silver nanoparticles synthesizedchemically
i. Estimation of toxicity effect of copper oxide nanoparticles synthesizedchemically
j. Estimation of toxicity effect of silver nanoparticles synthesizedbiologically
k. Estimation of toxicity effect of copper oxide nanoparticles synthesizedbiologically
MATERIAL AND METHODS
Sample collection and preparation
All chemicals used in this experiment were of obtained from Ramkem, Fisher scientific and Thomas Baker or as otherwise indicated. For biological synthesis Saraca indica leaves were taken from botanical garden of M.I.E.T.,Meerut. Leaves extract was prepared freshly as per the required amount for the synthesis of biological nanoparticles. Sample collection of five microorganisms, namely Enterobacter aerogens, Psuedomonas aeruginosa, Staphylococcus epidermidis, Bacillus subtilis and Micrococcus luteus, were collected from microbiology division of Institute of Microbial Technology, Chandigarh, Punjab. Analytical grade Sodium hydroxide and silver nitrate of himedia were ulitilised for silver nanoparticle synthesis experiment. A stock of 1mM was prepared and stored in brown bottle to avoid bottle disintegration of silver nitrate and a stock of 38mM trisodium citrate was prepared. Sodium hydroxide and copper chloride of analytical grade were used for nanoparticles of copper synthesis chemically.
Glassware and apparatus
All glass wares (measuring cylinder, beaker,petri plates, conical flask and test tubes etc) were obtained from Borosil, India
Equipments
- Cooling centrifuge(REM)
- Magnetic stirrer
- FTIR
- TEM
- Lyophilizer
- UV (VIS)
- BOD incubator
- Autoclave
- Hot air oven
Method of preparation of nanoparticles
Synthesis of Silver nanoparticles by chemical method (CH-AgNO3NPs)
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- Quote paper
- Dr. Pankaj Kumar Tyagi (Author), 2018, Biological and Non-Biological Synthesized Nanoparticles against Bacterial Species, Munich, GRIN Verlag, https://www.hausarbeiten.de/document/1311493
