Metallic nanoparticle synthesised by biological route: safer candidate for diverse applications

Abstract

Biological synthesis of nanoparticles (NPs) involves greater prospect; however, a detailed review is required for ecofriendly, faster and stable NP formulation in large scale for different commercial applications. The present article highlighted recent updates on biological route of single and bimetallic NP synthesis wherein the chemical reducing agents are eliminated and biological entities are utilised to convert metal ions to NPs. Application of the biological reducing agents ranging from bacteria to fungi and even natural plant extracts have emerged as eco‐friendly and cost‐effective routes for the synthesis of metal nanomaterials. Potential applications of such NPs, a wide range of analytical techniques used for characterisation and factors influencing the synthesis of NPs are focused. Further, elucidation of the mechanisms associated with the NP formation using microorganisms, as well as plant‐based materials are analysed which would be helpful for wide range of readers in the field of NP research for future selection and commercial implementation.

1 Introduction

Nanoparticles (NPs) draw greater attention due to their numerous applications in different fields. NPs can be broadly grouped into two categories – inorganic NPs and organic NPs. Inorganic NPs are further differentiated into noble metal NPs such as gold and silver, magnetic NPs and semiconductor NPs such as titanium oxide and zinc oxide. The organic NPs include carbon NPs. The physicochemical characteristics of NPs are because of their extremely small size, large surface to volume ratio, highly reactive surfaces, chemical compositions, shape, size, solubility and aggregation [1]. Metallic NPs are most promising and remarkable biomedical agents. Gold, silver, titanium, palladium, aluminium, zinc, carbon, iron and copper have been typically used for the synthesis of NPs. Metal NPs are of use in various catalytic and electro‐catalysis applications [2, 3], material science, physics, electronics, sensor technology [4, 5], plasmonic wave guiding material [6], environmental remediation fields [7], biology and biomedical applications such as biological levelling, disease diagnostic, drug delivery systems, antimicrobial effects and their applications in medical and pharmaceutical products (Tables 1 (a) and (b)) [8, 9, 10, 11].

Table 1a.

Practical application of gold and silver NPs synthesised through biological route

NPs types	Practical applications	References
gold NPs	• Medicinal material in treatment of tumours; as their ability to passively accumulate in tumours	[12, 13]
	• Cancer photothermal therapy; due to their unique optical and chemical properties;	[14, 15]
	• Biocompatible AuNPs for tracking and targeted delivery of anticancer drug thus improving delivery, minimising treatment durations and side effects	[16, 17, 18]
	• Targeted gene delivery; DNA labelling	[19, 20, 21]
	• In vitro assays include immunoassay, protein assay, capillary electrophoresis, time‐of‐flight secondary ion mass spectrometry and detection of cancer cells	[22, 23]
	• Antibacterial agents against various bacterial strains: for example, antimicrobial activity of vancomycin was enhanced on coating with AuNP against vancomycin resistant enterococci (VRE)	[24]
	• Antiviral properties because of blocking the viral attachment to host cell surface, e.g. anti‐HIV activity and inhibit several influenza virus	[25, 26]
	• Chemical and biological sensing	[27]
	• Bioconjugation, biomedical imaging, microscopy and photoacoustic imaging, surface‐enhanced Raman spectroscopy (SERS) applications	[28, 29, 30]
	• Catalysis applications in the reduction of aromatic nitro‐compounds, e.g. conversion of 4‐nitrophenol to 2‐amino‐phenol could be used in waste decontamination	[31]
silver NPs	• Used to treat burns, wounds and infections preservative agent in healthcare and food industries	[32]
	• Nano‐Ag used in many mechanical devices, in heat liable instruments like PCR lid, UV‐spectrophotometer and as coating materials for parts of various instruments	[33, 34, 35]
	• Research interest due to its inherent antimicrobial, anticancer, antiviral and antifungal effects as well as mechanism studies for inhibitory effect on bacteria;
	• Antiviral activity in HIV‐I, herpes simplex virus, monkey pox virus, influenza virus, respiratory syncytial virus	[36, 37]
	• Fungicidal and fungi static effects on dermatophytes Candida species, Trichophyton mentagrophytes
	• Applied as antimicrobial agents in various commercially available medical, textile, assorted electronics, household goods and consumer products	[38, 39]
	• Developed as low‐cost biosensors to detects pathogen and monitors the different stages of contaminants	[40, 41]
	• Sized controlled silver colloid NPs for antimicrobial activity against drug resistant pathogen	[42]
	• Crop protection and management of agricultural plant disease due to its antifungal effects as well as active against plasmodial pathogens;	[43]
	• Used to control plant pathogen in a safer way as compared to conventional fungicides	[44]
	• Use of antifungal activity for biostabilisation of footwear materials; 1% nanosilver solution inhibited the growth of yeast like mould and fungal strains
	• Various ecofriendly nanoproducts in commercial markets such as water purifier, bone and teeth cement, toothpastes, mouthwash, homemade products, facial cream, personal care and cosmetics such as sunscreens, acne creams, anti‐aging creams, perfumes, shampoo, soap, detergent	[45]

Table 1b.

Application of biologically synthesised nanoparticles

NPs types	Practical applications	References
iron NPs	• Anticancer agent;	[46]
	• Molecular diagnosis for biodetection;	[47, 48]
	• Used for cancer therapeutics	[49]
	• Arsenic and chromium (VI) removal ability ground water and wastewater	[50, 51]
zinc oxide NPs	• ZnO NP has selective toxicity to bacterial and minimal effects on human cells which recommended their prospective uses in agricultural and food industries	[52]
	• Bactericidal effects due to reactive oxygen species generation, cell wall attachment by electrostatic interaction, membrane disruption, loss of permeability
	• Good antibacterial activity and stability under harsh processing conditions applied for food packing and waste water treatments
	• Cosmetics and coating applications
	• Formulation as nail paint	[53]
	• Antifungal activity against Microsporum canis and Trichophyton mentagrophytes fungi causing onychomycosis	[54]
magnesium oxide NPs	• MgO NP have ability to adsorb and retain significant amount of elemental chlorine and bromine for a long time that exhibit biocidal activity against vegetative Gram‐positive bacteria, Gram‐negative bacteria and the spores;	[55, 56]
	• MgO NP possess properties of potent disinfectant
copper and copper oxide NPs	• Research interest due to strong antimicrobial agents and disinfecting properties against number of infectious organisms;	[57]
	• CuO is cheaper than silver; highly ionic nanoparticulate metal oxides; easily mixes with polymers; stable physical and chemical properties and having antimicrobial activity
	• Potential as an effective bactericide material to coat hospital equipment
platinum NPs	• Used in water electrolysis applications	[58]
	• Catalytic activity used in production of hydrogen fuel element
palladium NP	• Anticancer and biocatalytic applications	[2]
titanium dioxide NPs	• Involve in water pollution mitigation researches	[59]
	• Antibacterial activity applied as antibacterial coatings and waste water disinfection processes	[60]
	• TiO2 NPs on UV irradiation can be used to reduce disinfection time, eliminate pathogenic microorganism in food contact surfaces and enhance food safety	[61]
	• Antibacterial activity due to its photo‐activation that promotes bactericidal effects. Peroxidation of polyunsaturated phospholipid of membrane and loss of respiratory activity
	• No toxicity in dark condition	[62]
	• TiO2 NPs used in conjugation with silver as an antimicrobial agent	[63]
	• Used in cosmetics
	• Applied to filters that exhibit strong germicidal properties and remove odours
aluminium oxide and alumina NPs	• Aluminium oxide has applications in industrial and personal care products;	[64]
	• Aluminium oxide has antimicrobial properties due to generation of reactive oxygen species and high tendency to bind to the cell wall. That damage to bacterial cell wall and increased permeability	[65]
	• Alumina NPs have also growth‐inhibitory effects on E. coli due to positive charge at neutral pH. The electrostatic interaction with negatively charged E. coli cell resulted in the adhesion of NPs on the bacterial surface and reduced its growth
selenium NPs	• Utilised as potential diet supplements for selenium	[66]
	• Bacteriological reduction using E. coli protocol used for large‐scale production of protein capped Se NPs

The synthesis of metal NPs using biological system is a developing research area due to the safe method for producing NPs with specified properties and prospective application in nanomedicine. For several years, scientists have persistently explored different synthetic methods to synthesise NPs. On the contrary, the biological or green method of synthesis of NPs is easy, clean, reliable, benign, cost effective, efficient, and eco‐friendly in comparison to physical process or chemical‐mediated synthesis. The chemical synthesis is highly expensive that involves toxic solvents, high pressure, energy and high temperature conversion. Hence, biological synthesis is the greatest option to select for the synthesis of NPs. The prospect of exploiting natural resources for metal NP synthesis has become to be a competent and environmentally benign approach [67, 68]. ‘Green’ chemistry and bioprocesses for synthesis of NPs might cover the way for researchers across the globe to explore the potential of safe and easily available sources such as bacteria, actinomycetes, yeast, fungi, algae, herbs and plants in order to synthesise NPs [69].

The biosynthesis of metal NPs using microorganisms such as bacteria [70], fungi [71], actinomycetes [72], yeasts [73] and viruses [74, 75] is considered as potential biofactories due to their instinctive and natural ability to produce metal NPs either intra‐ or extra‐cellular [67, 76, 77]. Moreover, phytosynthesis that utilises parts of whole plants or plant extracts as biological factories for synthesis of metallic NPs are under exploitation and preferred as an advantageous and profitable approach [1, 68, 69, 78]. Microbe‐mediated synthesis is complex and involves multistep processes such as microbial isolation, culturing and maintenance, which cannot be feasible industrially due to its lab maintenance. In contrast, the plant‐mediated synthesis is a very rapid and cost‐effective approach and the produced NPs are more stable that can be easily scaled up for bulk production of NPs. The use of plant tissue culture techniques and optimisation of the downstream processes also makes it possible to synthesise stable and suitable metal NPs with desired size at an industrial scale. The plant‐mediated synthesis is truly a ‘green’ synthesis route in comparison to the other known methods of NP synthesis [1]. In addition, plants are known to harbour a broad range of metabolites. However, their potential is yet to be fully utilised in full throttle for synthesising metallic NPs. A schematic representation of the review is shown in Fig. 1.

Fig. 1.

Graphical representation of informational steps for green synthesis of NPs

Tables 2 and 3 summarised some of the examples of NPs synthesised through microorganisms and plant, respectively.

Table 2.

Examples of metal‐based NPs synthesised through microorganisms

Microorganisms	NP type	Size, nm	Morphology and features
bacterium
Desulfovibriode sulfuricans	Pd	—	—
Pseudomonas aeruginosa	Au	15–30	extracellular
Pseudomonas stutzeri	Ag	upto 200
Magnetotactic bacteria	magnetic (Fe3 O4)	—	—
	greigite (Fe3 S4)	—
Rhodopseudomonas capsulata	Au	50–400(at pH 4)	extracellular and intracellular
		10–20 (at pH 7)
Plectonemab oryanumUTEX 485 (Cyanobacterium)	Au	10–200	octahedral Au platelets, at the cell wall
Actinomycetes
Thermomonospora sp.	Au	8	extracellular
Rhodococcus sp.	Au	5–15	intracellular
Streptomyces sp. LK3	Ag	5	extracellular
Streptomyces sp.	Zn	65–80	extracellular, spherical
Streptomyces sp. JF741876	Ag	80–100	extracellular, spherical
Streptomyces viridogens HM10	Au	18–20	intracellular, spherical, rod
Yeast
Candida glabrata	CdS	200	intracellular
MKY3	Ag	2–5	extracellular
Schizosaccharomyces pombe	CdS	200	intracellular
Fungus
Aspergillus fumigatus	Ag	5–25	spherical, extracellular
Colletotrichum sp.	Au	20–40	spherical, triangular, extracellular
Verticillium	Ag	13–37	spherical, intracellular
	Au	20	spherical, intracellular
Fusarium oxysporum	CdS	5–20	spherical, extracellular
	Zr	3–11	spherical, extracellular
	barium titanate	4–5	spherical, extracellular
	CdSe	9–15	quantum dots, extracellular
	silica and titanium particles (SiF6 2− and TiF6 2−)	5–15	spherical, extracellular
Fusarium oxysporium	Ag	5–15	spherical, extracellular
	Au	20–40	spherical, extracellular
Neurospora intermedia	Ag	25–34	Spherical
virus
Tobacco mosaic TMV	SiO2		NP on surface
	CdS
	PbS
	Fe2 O3
M13 bacteriophase	ZnS, CdS		quantum dots, nanowires

Table 3.

Examples of metal‐based NPs synthesised through plants derivatives

Plant derivatives	NP type	Size, nm	Morphology and features
Aloe vera leaves	Ag	15.2	spherical
	Au	—	crystalline
Pelargonium graveolens leaves	Ag	16–40	crystalline
	Au	20–40	decahydral, icosahedral
Eucalyptus camaldulensis leaves	Au	5.5–7.5	crystalline
Coriandum sativum leaves	Au	6.7–57.9	triangular truncated, triangular, decahedral
Jatropha curcas latex	Ag	10–20	crystalline
Jatropha curcas seed	Ag	15–50	spherical
Carica papaya fruit	Ag	15	Cubic
Cinnamon zeylanicum Bark powder	Ag	31–40	spherical
	Pd	15–20	crystalline
Cinnamon zeylanicum leaves	Au	25	spherical, prism
Pinusdesiflora leaves	Ag	15–500	cubic
Ocimum sanctum root stem	Ag	5–10	spherical
Sorghum spp. Bran powder	Ag	10–70	—
Curcuma longa power, tuber	Ag	21–30	quasi‐spherical, triangular, rod shaped
Mangiferaindica leaves	Ag	20	spherical, triangular, hexagonal
	Au	17–20	spherical
Mukia Maderaspatna plant extract	Ag	20–50	spherical, triangle, hexagonal
	Au	20–50	spherical, triangle, hexagonal
Nicotiana tobaccum leaves	Ag	8	crystalline
Zingiberoffcinale rhizome	Ag	6–20	spherical
Piper betle leaves	Ag	3–37	spherical
Beta vulgaris SugarBeet pulp	Au	—	spherical, rod shaped, nanowires
Nyctanthesarbortristis flower extract	Au	19.8	spherical, triangular, hexagonal
Cuminum cyminum seed	Au	1–10	spherical
Pinusresinosa Bark	Pd	16–20	spherical
	Pt	6–8	Irregular
Musa paradisica peeled banana	Pd	50	crystalline irregular
Rosa indica L. petals extract	ZnO	50–100	crystalline, moderately stable
Medicago sativa (alfalfa)	Ti–Ni Alloys	1–4	fcc‐like (face‐centred cubic) geometry for smallest clusters and complex arrays for biggest
	Au	2–40	irregular, tetrahedral, hexagonal platelet, decahedral,
	Iron‐ oxide	2–10	icosahedral Crystalline

2 Microorganism‐mediated routes for NP synthesis

Microorganisms are very significant because of their enhanced genetic diversity. Various bacterial, fungal, yeast, algae and some viruses have been screened, explored and most widely used for the synthesis of silver, gold, platinum, titanium dioxide and zinc oxide etc. NPs. This route for NP synthesis is inexpensive, less cumbersome, less requirement of energy, less wastage of input, more practical control of constituent ingredients, non‐toxic and excellent yield when compared to the use of chemical and physical method in a definite time span.

Microorganisms possess a rich diversity of versatile biocatalysis and biochemically exceptional (microbial) genome, therefore they enable intracellular, as well as extracellular synthesis of different types of metallic NPs. Intracellular synthesis of silver NPs from psychrophilic (grow in very low temperature) bacteria has been reported which was due to the intracellular bacterial proteins and the chelating activity of DNA subunits [79, 80]. Extracellular synthesis of silver NPs using psychrophilic bacteria and its excellent antibacterial activity was observed by Shivaji et al. [81]. The in vivo factors such as biomolecules, enzymes and metabolic intermediate molecules decide if the microbial synthesis would be extracellular or intracellular [82]. The schematic of general mechanism for microorganism‐based NP synthesis is represented in Fig. 2.

Fig. 2.

Components of microorganism‐mediated route of NP synthesis

An immense benefit of extracellular synthesised NPs is the fact that they are native after formation. It can readily isolate and has simple downstream processing. Also, as these are externally synthesised at cell surface or at the periphery, they are applicable for various therapeutic uses, such as bioimaging, sensor integration, electronics and optoelectronics depending on their particular shape which favours them for particular application. The shapes of the NPs decide functionalities in and their roles in applications they are used in. For example, Du et al. [83] observed that the biochemical reduction of gold using aurochloric acid to the gold NPs was accomplished using DH5α of Escherichia coli. These NPs are predominantly spherical shaped along with some of triangular and hexagonal shapes and get adhered onto the cell surface. These synthesised NPs have been exploited for promising applications as a consequence of their resemblance in origin to iron‐complexed haemoglobin and other in vivo proteins.

In general, NPs synthesised at intracellular locations are smaller in size that is more desired for specific applications. However, the downstream processing approaches for intracellular NPs are very typical and the extraction procedures are tough that lead to the disadvantage of low yields. Thus, the overall processes become expensive and time consuming. Some microorganisms which have been used for the production of NPs are summarised in Table 2. The following paragraphs also describe the microorganisms used for the biosynthesis of NPs and their properties that should be inherent for the production of NPs of desired characteristics.

2.1 Bacterial‐mediated NP synthesis

Pseudomonas, Lactobacillus, E. coli, Rhodopseudomonas capsulate, Actinobacter species and Klebsiella pneumoniae are major bacterial strains that have been applied for both extracellular and intracellular synthesis of NPs. The process factors such as temperature, pH, ionic atmosphere and deviations in the physical conditions are responsible for variation in different sizes of the synthesised NPs. The smaller size range and specific shape of NPs synthesised are vital parameter for their definite application thus, a multitude of factors needs to be optimised for the specific synthesis of NPs in a particular configuration. The synthesis of NPs using Rhodopseudomonas species produces NPs at intracellular and extracellular positions but at different pH values (Table 2) [84]. The time taken for the production of NPs also varies. Some NPs had been immediately produced as soon as the culture is ready, while others required up to 2 h of incubation period and some others have taken 20–24 h [70, 85, 86].

2.2 Actinomycetes‐mediated NP synthesis

Actinomycetes were used for synthesis of different metal NPs such as gold [87, 88, 89], silver [90, 91], copper [92], manganese [93] and zinc [93]. However, their extracellular synthesis was reported on longer time period in the range of 24–120 h in comparation with the bacterial and fungal biogenic synthesis.

The thermophilic actinomycete, Thermomonospora sp., has been reported for extracellular production of gold NPs with a much improved polydispersity [72]. An attempt towards elucidating mechanism and/or conditions supporting the formation of NPs with desired features was performed by the reduction of aurochloric ions using Thermomonospora sp. biomass that had resulted in efficient synthesis of monodisperse gold NPs [94]. The reduction of metal ions and stabilisation of the gold NPs were believed to occur by an enzymatic process, as well as due to extreme biological conditions such as alkaline and slightly elevated temperature conditions used for the synthesis of gold NPs.

Based on the above hypothesis alkalotolerant Rhodococcus species has been used for intracellular synthesis of good quality monodispersed gold NPs. The researchers observed that the concentration of NPs was more on the cytoplasmic membrane than on the cell wall. They suggested that the enzymes present in the cell wall and on the cytoplasmic membrane are responsible for metal ion reduction but not due to the enzymes and proteins in the cytosol. These metal ions were not toxic to the cells which were producing them and the cells were continued to multiply even after the biosynthesis of gold NPs [95].

Golinska et al. [96] reviewed biogenic synthesis of metal NPs using Actinomycetes and proposed hypothetical mechanism of silver NPs formation. Isolated Streptomyces sp. LK3 produced extracellular enzyme NADH‐dependent nitrate reductase that was responsible for reduction of nitrate to nitrite and also reduced nitrite to nitrogenous gases. The possible mechanism was based on the electron shuttle enzymatic metal reduction process [89].

2.3 Fungal‐mediated NP synthesis

In addition to good monodispersity, NPs with well‐defined dimensions can be obtained by using fungi. Compared to bacteria, fungi could be a good source for production of NPs in large scale. Since fungi are known to secrete much higher amounts of proteins, they might have significantly higher productivity of NPs in biosynthetic approach [77]. These research groups have shown the formation of gold NPs with well‐defined dimensions and good monodispersity using fungal mycelia of Verticillium species and Fusarium oxysporum. The results have documented that the trapping of aurochloric ions on the surface of fungal cells could occur by electrostatic interaction with positively charged groups, such as lysine residues in enzymes that are present in the cell wall of the mycelia. Here the gold ions were reduced by enzymes within the cell wall leading to aggregation of metal atoms and formation of gold NPs. However, they could not find the exact mechanism of formation of gold NPs. The fungus, Aspergillus flavus also resulted in the accumulation of silver NPs on the surface of its cell wall when incubated with silver nitrate solution [97]. Ag NP biosynthesis using active and inactive cell‐free filtrates of Neurospora intermedia was conducted for elucidation of biosynthesis mechanism [98]. The role of proteins and the rate of synthesis depending on the interactions of amino acids with Ag+ were established. The inactive cell‐free filtrate established higher yield in contrast with the active cell‐free filtrate because of the higher organic contents of the inactive cells.

Various types of NPs have been synthesised using common fungi such as Fusarium, Penicillium, Aspergillus and Phoma species. Eleven isolates of Fusarium species and 18 Phoma species were screened for mycosynthesis of silver NPs. Synthesised AgNPs were distinguished by UV–vis spectroscopy which gave absorbance peak at 420 nm. The detailed characterisation was carried out using zeta potential, NPs tracking analysis, photon correlation spectroscopy, powder X‐ray diffraction (XRD), fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) analysis [99, 100].

These fungal cultures acquire some additional attributes, such as easy optimisation steps for scale‐up in the bioreactor whereas plant‐ and bacterial‐based extracts cannot. Fungi also have rapid growth rate that enables the release of sufficient concentrations of vital enzymes and proteins. These extracellular molecules facilitate faster bioreduction of metal salts and biochemically reduce metallic ions to zero‐valent NPs. Their rapid growth rate and high production capacity of extracellular enzymes and proteins eliminates the technical difficulties in the downstream process [101].

Mechanism of biosynthesis of extracellular silver NPs by Fusarium oxysporum strain was established. The synthesis occurred due to the presence of extracellular anthraquinone and NADPH‐nitrate reductase enzyme. The electron required to convert silver ions into Ag neutral (Ag⁰) was donated by both NADPH and quinine [102]. An enzymatic route using α‐NADPH dependent nitrate reductase along with phytochelatin for the formation of AgNPs was also demonstrated. The reduction of α‐NADPH to α‐NADP⁺ and the hydroxyquinoline may have acted as means of transferring the electron during the reduction of nitrate to Ag²⁺ ions converting them to Ag [103].

Biosynthesis of noble metal NPs using mycoendophytes is fewer documented. Mycoendophytes (fungal endophytes) are fungi that live within plant tissues as symbiotic relationship. They offer variety of bioactive secondary metabolites as well as extracellular enzymes suitable for NP synthesis. Golinska et al. [104] reviewed mycoendophytes as efficient candidature for bio‐NPs and certain species Epicoccum nigrum, Guignardia mangiferae, and Amylomyces rouxii have been exploited for synthesis of metal NPs. Silver NP synthesised from mycoendophytes presents stability, polydispersity, biocompatibility and remarkable antimicrobial activity against bacterial and fungal pathogens of human and plants.

2.4 Yeast‐mediated NP synthesis

The Candida glabrata and Schizosaccharomyces pombe were used for the first time in the biosynthesis of cadmium sulphide (CdS) nanocrystals (Table 2). These nanocrystals were produced using cadmium salts and are now used in quantum semiconductor crystallites. Kowshik et al. [105] improved the quantity of semiconductor CdS nanocrystals production that was achieved by using Saccharomyces pombe cells at their mid‐log phase of growth and maximal nanocrystals were obtained. The authors concluded that the formation of CdS nanocrystals was dependent on the growth phase of yeast. Various physio‐chemical parameters and culture conditions have also been standardised for the synthesis of large quantities of silver NPs by using silver‐tolerant yeast strain MKY3. The procedure for separation of these NPs has also been documented that was based on differential thawing of the samples [73]. Recently, yeast strains have been identified for their ability to produce gold NPs, whereby controlling the growth and other cellular activities resulted in controlled size and shape of the NPs [106].

3 Phytosynthesis routes for NP synthesis

Plants including inactivated plant tissue, plant extracts and living plants‐mediated biosynthesis of metal NPs are currently under exploitation and have received more attention as a suitable alternative to chemical procedures and physical methods. Plant‐mediated NP synthesis is attracting interest because of its suitability in nanomedicine‐based innovations and other multiple applications. Plants with high genetic variability contain diverse biomolecules and various coenzymes and vitamins‐based metabolic intermediates that can reduce metal ions to NPs in a single step. These methods have been conducted at room temperature and pressure conditions and have not required any stringent technical requirements. Moreover, plant materials are easily and cheaply available as compared to those of microorganisms. The plant‐based NP synthesis is easy to scale up and is environmental friendly. Whole plant extracts have also been used for synthesis of NPs [107, 108, 109]. The extract of different plant parts such as leaf, fruit, seed have showed much better results in comparison and have significant control in size and shapes of NPs (Table 3) [110, 111, 112, 113, 114, 115, 116, 117, 118].

Extracts from plants can act both as reducing and capping agents in NP synthesis. Biomolecules such as enzymes, coenzymes, proteins, amino acids, vitamins, polysaccharides and organic acids such as citrates and metabolite materials such as phenolic compounds, alkaloids, and sterols found in plant extracts serve as excellent reducing and stabilising agents for bioreduction of metal NPs. The source and nature of plant extract is the most important factor that affects the type and morphology of the synthesised NPs. This is due to the difference in biochemical concentrations in various plant extracts.

The formally applied procedure for synthesis of NPs from the plant extracts involves mixing the plant extract with metal salt solution at room temperatures. The biochemical reactions are versatile and flexible in nature and completes within few minutes and the metals are transferred from their mono or divalent oxidation states to zero‐valent states. This is actually noticed through the colour change in the culture medium vessel and shows the formation of NPs. Synthesis of gold, silver, and various other metal‐based NPs have been observed using this approach. Common plants such as Azadirachta indica (neem), Aloe vera, wild Geranium species leaf, Datura metel, Camellia sinensis (Tea leaves), Nelumbo nucifera (Lotus), Euphorbia hirta, Rhododedendron daurican (Lal buransh), Ocimum tenuiflorum (Tulsi) and Cinnamomum camphora (Camphor tree) have been used to produce NPs. The recent reports on various plant extract used for synthesis of metal NPs have been summarised in Table 3.

4 Biological synthesis of bimetallic NP

Generally, monometallic NPs have been biosynthesised for their various unique features. However, few attempts on green methods have also been described for the generation of different kinds of bimetallic NPs including Au–Ag, Au–Pd, Ti–Ni [119, 120, 121, 122, 123, 124]. Bimetallic NPs with alloy or core‐shell structures could improve the properties of their corresponding monometallic NPs. Both scientific and technological points of view the bimetallic NPs are having various novel potentials for electronic, optical, magnetic and catalytic applications [31].

The preparation of Au/Ag bimetallic NPs was investigated as a straight forward, green facile and economically viable method using degraded pueraria starch (DPS) as a matrix [31]. DPS has good water solubility and biocompatibility which acted as both a reducing agent and a capping agent for the bimetallic NPs synthesis. The research group demonstrated that these DPS‐capped Au/Ag bimetallic NPs could function as catalysts for the reduction of 4‐nitrophenol in the presence of NaBH₄ and were more effective than Au or Ag monometallic NPs.

Gopinath et al. [125] synthesised spherical silver (Ag), gold (Au) and bimetallic (Ag/Au) NPs using aqueous bark filtrate of Terminalia arjuna as simple, rapid, cost‐effective and environmentally safer biopesticide for controlling the malarial vector. The synthesised Ag, Au and Ag/Au NPs were distinguished by XRD, scanning electron microscopy‐energy‐dispersive X‐ray analysis (SEM‐EDX), FTIR and UV–vis spectrophotometer investigation for the detection of particles nature, size and shape, possible functional metabolites and absorption range, respectively. The effectiveness of the prepared Ag, Au and Ag/Au NPs was checked against third and fourth instars larvae of Anopheles stephensi, a malarial vector. The author demonstrated that Ag/Au bimetallic NPs had more significant effect compared with Au and Ag NPs treatment alone. The phyto‐mediated synthesised NPs can be a rapid, simple, cost‐effective and environmentally safer biopesticide for controlling the malarial vector.

A facile, eco‐friendly and pure ‘green route’ for synthesis of Au–Pd bimetallic NPs (∼7 nm) has been reported where no additional synthetic reagents were used as reductants or stabilisers [121]. The NPs preparation were based on simultaneous bioreduction of Au(III) and Pd(II) precursors (HAuCl₄ /PdCl₂ mixtures solution) with Cacumen platycladi leaf extract in aqueous solution. FTIR spectra analysis confirmed that the C=O and C–O groups in the plant extract played a significant role in capping the bimetallic NPs. The specific mechanisms involve in formation of bimetallic NPs are still not entirely understood.

5 Factors affecting NPs synthesis

NPs are commonly organised based on its synthesis route and their dimensionality, morphology, composition, uniformity and agglomeration. The characteristics of NPs synthesised using various routes have been found to be effective making them more powerful. The main challenges frequently encountered in the biosynthesis of NPs are to control the shape and size of the particles as well as to achieve monodispersity in solution phase. Several factors such as temperature, pH and reaction or incubation time directly influence or cause some interference in the formation of metal NPs by plant.

5.1 Effect of pH

pH can play key role in the biosynthesis of NPs. pH is an important factor in the biosynthesis and the shapes and size of NPs can be varied with the change in pH. Several researchers reported that larger NPs were formed at lower pH (2–4) compared to the higher pH. As different plant extracts and even the extracts prepared from different parts of the same plant are having different pH values, optimisation of a synthetic protocol is needed for efficient synthesis of NPs [110]. In addition, it was reported that silver NPs exhibit a lower zeta potential value (−26 mV) at strongly acidic pH compared to alkaline pH solutions indicating the higher stability and smaller size of NPs at basic pH. This was supported by the absorbance peaks in SPR spectra at different pH during their study. Andreescu et al. [126] observed a negative zeta potential of the synthesised silver NPs at different pH. The increase in pH results in an increase of the absolute value of the negative zeta potential which led to the formation of highly dispersed NPs. This phenomenon could be associated to the electrostatic repulsion at high pH or attributed to the high absolute value of the negative zeta potential.

The size of gold NPs produced by Avena sativa was highly dependent on the pH value [127]. At pH 2, large‐sized NPs (25–85 nm) were formed in a small quantity, but at pH 3–4, smaller‐sized NPs were formed in a large quantity. The researchers speculated that at low pH (pH 2), the gold NPs prefer to aggregate to form larger NPs rather than to nucleate and form new NPs. In contrast, at pH 3 and 4 more functional groups like carbonyl and hydroxyl are available for gold binding; thus a higher number of new Au(III) complexes would bind to the biomass at the same time that will nucleate separately and form NPs of relatively small size. At higher pH, the availability of a large number of functional groups facilitates a higher number of Ag(I) to bind with biomolecules and subsequently form a large number of NPs with smaller diameter. The effect of pH during synthesis of silver NPs using Cinnamom zeylanicum powder and bark extract was studied over a wider pH range (1–11) and it was concluded that the pH of the solution dropped in most of the cases after the synthesis of silver NPs. Furthermore, the large‐sized ellipsoidal silver NPs were observed at acidic pH, while at higher or alkaline pH highly dispersed, small‐sized and spherical NPs tended to form [128].

In contrast, Dwivedi and Gopal [129] revealed that silver and gold NPs prepared by using Chenopodium album leaf extract were stable in a wider range of pH (2–10) with small variation in the zeta potential values. Recently, Veerasamy et al. [130] while working on mangosteen leaf extract observed that at low pH aggregation of silver NPs was favoured over the nucleation. However, higher pH facilitated the nucleation and subsequent formation of a large number of NPs with smaller diameter.

5.2 Effect of reaction temperature

Biological synthesis of NPs has a positive correlation with an increase in temperature. The silver NPs synthesised using the Curcuma longa tuber powder extract showed the increase in surface plasmon resonance with increase in temperature which confirmed the positive correlation between the yield of the NPs and the temperature [131]. Temperature is one of the crucial factors that control the size and shape of NPs. Higher rates of gold NPs were formed at higher temperatures. In addition, nanorod and platelet‐shaped gold NPs were synthesised at higher temperatures, while spherical‐shaped NPs were formed at lower temperatures [106].

The rapid synthesis rate of silver NPs at higher temperatures was also reported by Andreescu et al. [126]. Due to an increase in temperature the rate of reaction also increased which enhanced the synthesis of NPs. Plant‐mediated synthesis of both silver and gold NPs using Tansy (Tanacetum vulgare) fruit extract exhibited an increase in the sharpness of absorption peaks with an increase in temperature from 25 to 150°C [110]. The sharpness in absorbance peak depends on the size of the synthesised NPs; with higher temperature the particle size was smaller which resulted in the sharpening of the plasmon resonance band of silver and gold NPs [129, 132].

5.3 Effect of contact or incubation time

The time duration required for completion of all steps of the reaction is defined as contact or incubation time that also affects the synthesis of NPs. Dwivedi and Gopal [129] reported an increase in the sharpness of UV absorption spectra peaks with an increase in incubation time while working with Chenopodium album leaf extract. They reported that NPs appeared within 15 min of the reaction and increased up to 2 h, but after that only slight variation occurred.

Due to the instability of NPs formed at initial stage of the reaction time, an optimum duration is always required for complete nucleation and subsequent stability of NPs. Veerasamy et al. [130] observed that the optimum time required for completion of the reaction for AgNP synthesis was 60 min during their experiment. Similarly, Ghoreishi et al. [133] also showed the requirement of an optimum reaction time for the stability of synthesised silver and gold NPs using Rosa damascene.

In case of bimetallic NPs synthesis mixing of two different aqueous metal solutions with a plant extract is required. Competitive reduction can be expected due to the presence of two metal ions with different reduction potentials in the solution. Theoretically, metal ions with greater reduction potential were reduced quicker than metal ions with lesser reduction potential. For example, during the synthesis of AuAg NPs, Au ions were reduced earlier forming the nuclei while Ag ions reduced afterwards and adsorbed on the Au particles, shaped a core‐shell structure [134]. Competitive reduction can increase the rate of reduction of metal ions. The author evaluated the reduction rate of Au ions for synthesis of monometallic Au NPs and bimetallic Au/Ag NPs. During synthesis of monometallic Au NPs, Au ions were completely reduced after 2 h while only 30 min was needed for the complete reduction of Au ions into Au NPs in case of bimetallic synthesis. A recent study by Tamuly et al. [135] discovered that the reduction process was accelerated during bimetallic synthesis. As noted, reduction of Au³⁺ during monometallic synthesis reached saturation within 3 h while during bimetallic synthesis, saturation was achieved after 90 min.

5.4 Effect of reactant concentration

The biomolecules present in plant extracts can significantly manipulate the synthesis and shape of metallic NPs. Variation in amount of sundried Cinnamomum camphora (camphor) leaf extract in the reaction medium extensively influenced the shape of the synthesised Ag and Au NPs. For instance, when increased concentrations of leaf extract were added to fixed concentration of precursor chloroauric acid, the resulting NP shape changed from triangular to spherical [136]. Chandran et al. [118] also found the change in shape of AuNPs from triangular to spherical using different volume of Aloe vera leaf extract in the reaction medium containing chloroaurate ions. The involvement of carbonyl compounds present in leaf extract was observed in shaping the particle growth.

The changes in Plectranthus amboinicus leaf extract concentration in the reaction medium modulated the Au NP size between 50 and 350 nm, as well as shapes such as spherical, triangular, hexagonal and decahedral [137]. In a study by Birla et al., enhanced reduction of silver ion with increased fungal (Fusarium oxysporum) filtrate volume was observed and 100% concentrated filtrate showed complete reduction [138]. The concentration of silver nitrate also influenced the production of Ag NPs and the suitable concentration for maximum synthesis was 1.5–1.8 mM.

6 Mechanism of NP synthesis by plant routes

Numerous studies have been mainly focused on the screening and identification of suitable plants or plant extracts for controlled synthesis of metal NPs; however, very little research has been performed to understand the actual mechanism behind the synthesis of NPs. Elucidation of the actual mechanism and biochemical pathways involved in the biosynthesis of metal NPs is necessary to progress a rational approach in this field. In recent years, several hypotheses have been proposed for NPs synthesis. However, the exact mechanism behind the plant‐mediated synthesis of metal NPs is not yet known and more detailed studies are needed.

As mentioned earlier, various plant metabolites including terpenoids, polyphenols, phenolic acids, alkaloids, saponins, steroids, tannins, sugars and proteins play a significant role in the bioreduction of metal ions forming NPs. Any plant extract has some biomolecule in its genome through which it carries out the biochemical reduction. For instance, the synthesis using geranium leaf extract and its endophytic fungus (Colletotrichum sp.) produced gold NPs of 16–40 nanometres [139] when sterilised geranium leaves and the fungus (Colletotrichum sp.) growing in the leaves were separately exposed to aqueous chloroaurate ions. The biogenic gold NPs synthesised using the fungus were essentially spherical in shape while the particles grown using the leaves exhibited a variety of shapes that included rods, flat sheets and triangles. FTIR spectroscopy of NPs showed the surface capping of gold NPs synthesised using the fungus was predominantly by proteins while terpenoids were implicated in stabilisation of the NPs synthesised using geranium leaf extract. Terpenoids are organic polymers synthesised in plants from five‐carbon isoprene units which exhibit strong antioxidant activity. Thus, it was suggested that in the synthesis of gold NPs using geranium leaves, the reducing and capping agents appeared was terpenoids and in case of fungus Colletotrichum sp., the identified component was polypeptides/enzymes.

Flavonoids are polyphenolic compounds that can actively chelate and reduce metal ions into NPs. These include flavonols, isoflavonoids, chalcones, anthocyanins, flavones and flavanones. Flavonoids contain diverse functional groups capable for NP synthesis. It has been assumed that a reactive hydrogen atom can be released during the tautomeric transformations of flavonoids from the enol‐form to the keto‐form that involves the reduction of metal ions and leads to formation of NPs. For example, in formation of silver NPs from Ag ions the transformation of flavonoids luteolin and rosmarinic acid from the enol‐ to the keto‐form of Ocimumbasilicum (sweet basil) extracts played an important role [140]. Moreover, the conversion of ketones to carboxylic acids in flavonoids was involved as internal mechanism of bioreduction.

In some cases, carbonyl groups or π‐electrons of flavonoids are able to chelate metal ions. For example, quercetin (flavonoid) has strong chelating activity due to involvement of its C3 carbonyl, C5 hydroxyls and the C3′ and C4′ position of catechol group. These molecules were involved in chelation of various metal ions such as Fe²⁺, Fe³⁺, Cu²⁺, Zn²⁺, Al³⁺, Cr³⁺, Pb²⁺ and Co²⁺ [67]. The presence of flavonoids‐based mechanisms gives possible explanations of adsorbed flavonoids on the surface of a growing NP. Furthermore, these flavonoids have a role in the initial stages of NP formation, i.e. nucleation and aggregation stage in addition to the bioreduction stage. Studies on isolated flavonoids and flavonoid glycosides also reveal the ability of the formation of metal NPs. Kasthuri et al. [141] isolated apiin (apigenin glycoside) from Lawsonia inermis (lawsonite thornless, henna) and used for production of isotropic gold and quasi‐spherical silver NP with 21–39 nm. Carbonyl group of apiin attached to the NPs was confirmed through FTIR analysis.

The formation of metal NPs can also be induced by sugars present in plant extracts. It is known that aldehyde group of monosaccharides like glucose can be used as reducing agents. Fructose which is a keto‐group containing monosaccharides can be used as antioxidant after undergoing tautomeric transformations from a ketone to an aldehyde. Again, the reducing ability of disaccharides and polysaccharides depends on the access of aldehyde and ketone group of their monosaccharide components in linear form. For example, open chain forms of maltose and lactose have reducing ability while sucrose is a non‐reducing sugar and hence notable to reduce metal ions.

Panigrahi et al. [142] described the effect of three different sugars, namely glucose, fructose and sucrose as reducing agent for the synthesis of metallic NPs such as Au, Ag, Pt and Pd. In their study, no other stabilising agent or capping agent was required to stabilise the NPs and the particles can be safely preserved in the desiccators for months. For synthesis of gold and silver NPs, when glucose was involved as reducing agent, the NPs formed were of different sizes and morphologies, whereas using fructose led to same‐sized monodispersed NPs. The silver and gold NPs of similar size may have been obtained because of the antioxidant potential of fructose. The non‐reducing sugar, i.e. sucrose was unable to reduce silver nitrate or palladium chloride into NPs while in the presence of tetrachloroauric and tetrachloroplatinic acids along with sucrose NPs were formed probably due to the acidic hydrolysis of sucrose into glucose and fructose which releases aldehyde and ketone groups. It is assumed that aldehyde group of sugar can be oxidised into a carboxyl group via the nucleophilic addition of OH⁻ which leads to the metal ions reduction. A similar mechanism was proposed for the bioreduction of gold ions using the magnolia vine extract [139].

The potential of amino acids for reduction of metal ions is found to differ with sugars. For example, lysine, cysteine, arginine, and methionine are able to bind silver ions, while lysine is capable of reducing tetrachloroauric acid to AuNPs. Valine and lysine do not have reduction or binding ability for silver NPs. All the 20 natural α‐amino acids have been tested for gold ions reduction ability and tryptophan has been found as the powerful reducing agent, whereas histidine is the strongest binding agent for Au ions [143, 144]. Amino and carboxyl groups present in main and side chains of amino acids can bind to metal ions. For example, in aspartic and glutamic acid the main carboxyl groups or in histidine the nitrogen atom of the imidazole ring was involved in NP formation. The thiol group of cysteine, thioether of methionine, hydroxyl groups of serine, threonine and tyrosine and carbonyl groups of asparagine and glutamine involved as side chains associated in binding of metal ions.

Free amino acids are used for the formation of polypeptide chain using their carboxylic and amino group that changes the metal reduction ability of individual amino acids. However, for free side chains functional groups can still contribute to the synthesis of NPs and the appropriateness of these side chain functional groups depend on the amino acid sequence of the peptide chain. Peptides are multifunctional reagents such as reducing and capping agents that can be used for the synthesis of biocompatible metal NPs under relatively mild conditions. However, it is difficult to establish sequence–reactivity relationships using biopeptides obtained from plant extract or selected by combinatorial display libraries due to their extensively changeable compositions and structures. Reports of FTIR analysis of synthesised NPs using plant extracts revealed that growing NPs are frequently found in association with different bioproteins.

Tan et al. [143] proposed the scientific basis for the rational design of peptides for the synthesis of metal nanostructures. The change in amino acid sequence of synthesised peptides can affect its capability of metal ion reduction and the author proposed a model for the peptide‐based synthesis of gold NPs in the aqueous solution. The two differently synthesised peptides; one containing those amino acids suitable for effective binding of metal ions and other peptide with amino acids acquired high metal ion reducing ability have showed lower reduction parameters than expected. This was explained in the first proposed model for the peptide synthesis of AuNPs in aqueous solution in which the reactant mixture of peptides and chloroaurate ions form peptide‐AuCl⁴⁻ complexes. These strong complexes have inhibitory effect on subsequent reduction to Au(0). Reduction facilitated by peptides can form Au atoms for nucleation and growth of nuclei into crystalline particles by addition of more Au atoms from the solutions or by fusion with other nuclei. Moreover, reaction with another synthetic peptide composed of amino acids such as leucine, phenylalanine and proline that poorly bind metal ions and is ineffective in reducing tetrachloroauric acid anions to form AuNPs. This is probably due to their inability to retain metal ions close to the reduction sites. The study concluded that protein molecules have amino acids with high potential for attracting metal ions and high reducing activity facilitated the formation of NPs. Furthermore, the amino acid sequence of a protein can significantly influence the size, morphology and amount of growing NPs. For example, a synthetic peptide GASLWWSEKL facilitated rapid synthesis of small‐sized (<10 nm) AuNPs of large amount. When the N‐ and C‐terminal amino acid were replaced in a peptide (SEKLWWGASL), it resulted in the formation of larger size (>40 nm) nanospheres and nanotriangles in shapes. This indicates that the proteins and peptides present in plant extracts play a significant role in determining the shape and size of NPs and also affect the overall yield of NPs.

Fig. 3 represents the summary of plant‐mediated NP synthesis which explains the feasibility for NP synthesis through the plant species that are biochemically rich. Various biomolecules of plant extract such as flavonoids, terpenoids, phenolic intermediates, vitamins, sugars, amino acids, protein, enzymes bring about the bioreduction of metal salt solutions. Furthermore, the synthesis can be directed towards highly selective and specific NP formation either in the form of aggregation mediated through self‐assembly or the NP stabilisation with in controlled structures. This particular ability has been very useful for application of integrated nanostructure assembly.

Fig. 3.

Factors involved in the plant‐mediated route of NP synthesis

Thus, the mechanism of metal NP synthesis in plant and plant extracts comprises of three phases which are (i) activation phase involves production of zero‐valent NP; (ii) growth phase in which the small and closer NPs spontaneously aggregate via non‐covalent interactions and (iii) process termination phase determining the final shape of the NPs. In this phase, NPs acquire the most energetically favourable conformation, the process being strongly influenced by the ability of a plant extract to stabilise metal NPs. Plants can therefore serve as readily available and faster biochemically rich source for NP synthesis.

7 Concluding remarks

Various physical, chemical, and biological methods can be used to synthesise metallic NPs. Among these methods, synthesis by biological methods is easy, safe and green processes eliminating the use of harsh, toxic and expensive chemicals, thus are environmental friendly and ecologically sound. In addition, the metal NPs synthesised using plant extracts are faster and stable in comparison with those produced by microorganisms. Currently, researchers have focused their attention in understanding the biological mechanism and enzymatic processes of NP biosynthesis, as well as the detection and characterisation of biomolecules involved in the synthesis of metallic NPs. Reduction potential of ions and reducing capacity of plant extracts depend on the presence of polyphenols, enzymes and other chelating agents present in plant and have critical effects on amount of NP production. Plant‐derived biopolymers could be important for the large‐scale production of highly stable monodispersed NPs with proper optimisation of reaction conditions. Continuous mode for production of NPs has been applied only for small‐scale production. In the chemical mode of NPs synthesis, the major production cost is associated with the cost of the metal salts, reducing agents and energy requirements while in ‘green’ route of synthesis, the main cost would be the cost of the metal precursors only since plant extracts obtained from the waste of food or agriculture industry can be used as reducing agents. Significantly associations involved in the food industry having thrust in recycling of the waste can provide financial help for NP production.