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Chapter 8
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Actinomycetes asNanofactories: Synthesis
andTherapeutic Applications
SompreetiPaul, SindhooraLakshmi, T.Amala, DakshithaAkula,
MrudulaRao, PiyushMohapatra, andAswaniThekkangil
Abstract In recent times, the use of nanomaterials has gained prominence in
the elds of agriculture, biological science, electronics, and medicine owing to
their numerous applications. Their unique characteristics, such as high surface/
volume ratio, specic structures, and dimensions that are similar to those of
biomolecules, provide special features for biomedical and therapeutic applications. It is well known that algae, bacteria, and fungi can take metals from their
surroundings and transform them into elemental nanoparticles, which can then
be accumulated or released. Actinomycetes, as green nanofactories, have been
regarded as an emerging eld for research to produce low-cost, high-quality
approaches for producing sustainable useful products with numerous applications. The distinctiveness of biologically active actinomycetes has drawn the
attention of experts all around the world to better understand their possibility as
effective nanofactories. As a dominant microbial phylum, actinomycetes are
helpful for the breakdown and transformation of organic and inorganic substrates and can also synthesize economically viable proteins, enzymes, and antibiotics. Antibiotics produced by these organisms have been utilized in almost
every medical intervention and are now known to facilitate the production of
nanoparticles with preferred surface and size attributes. Actinomycetes-derived
S. Paul · S. Lakshmi · D. Akula · M. Rao · A. Thekkangil (*)
Department of Biotechnology, School of Applied Science, REVA University Bangalore,
Bangalore, Karnataka, India
e-mail: aswani.t@reva.edu.in
T. Amala
Moulana College of Paramedical Sciences, Malappuram, Kerala, India
P. Mohapatra
Amity Institute of Biotechnology, Amity University, Noida, Uttar Pradesh, India
Ltd. 2024
S. Bose et al. (eds.), Concepts in Pharmaceutical Biotechnology and Drug
Development, Interdisciplinary Biotechnological Advances,
https://doi.org/10.1007/978-981-97-1148-2_8
139© The Author(s), under exclusive license to Springer Nature Singapore Pte
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nanoparticles have also been extensively studied for a wide range of applications
related to biomedical sciences. This chapter provides an overview of the synthesis of nanomaterials from actinomycetes, characterization techniques, and various biomedical and therapeutic applications. In addition, the challenges
pertaining to toxicity studies associated with the use and synthesis of biogenic
metal nanoparticles are discussed.
Keywords Nanomaterials · Actinomycetes · Nano factories · Therapeutic
application
S. Paul et al.
8.1 Introduction
Nanoparticles (NPs) are small particles that typically range in size from
one- hundred nm. These particles can exist naturally and can be synthesized
using different modes. They are similar to certain things in nature, such as
DNAand proteins (Khan etal. 2019). They are distinct in their sizes and surface areas.
NPs, owing to their unique characteristics, have a diverse array of applications in
multiple elds. Their exceptional properties, such as distinctive optical, magnetic,
and catalytic properties, along with enhanced electrical conductivity, increased
reactivity, and larger surface area, make them invaluable for a wide range of applications (Khan etal. 2019). Like in the eld of biomedicine, NPs are used to enhance
drug delivery. Their minuscule size allows them to penetrate biological systems and
reach cells or tissues effectively. Medications encapsulated within nanoparticles
exhibit improved solubility, stability, and controlled release, leading to more efcient treatments with reduced side effects (Modena etal. 2019). NPs have applications in food packaging to extend the shelf life and enhance safety. They are also
used in agriculture to optimize the management of fertilizers, pesticides, and growth
chemicals, minimize environmental impacts, and maximize crop yield (Khan etal.
2019). NPs are indispensable for environmental purication. They can be harnessed
to combat air pollution, facilitate waste management, and purify water by removing
impurities. Their use provides effective and long-lasting solutions to environmental
problems.
NPs can be synthesized from diverse resources like metals (gold, platinum,
and silver), metal oxides (like iron oxide and titanium dioxide), semiconductors
(such as tiny particles called quantum dots), plastics, carbon-based stuff (such
as graphene and carbon tubes), and biological sources like plants and microbes
(Jamkhande etal. 2019). There are different approaches to NP synthesis, such
as chemical synthesis, physical synthesis, and green synthesis (from living
things). The characteristics of NPs can also be regulated using different
approaches.
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141
8.2 Green Nanosynthesis: ASustainable Method
NPs can be synthesized in two ways: Biological and nonbiological. Nonbiological
techniques require unusual response conditions, signicant energy inputs, and precarious ingredients. An eco-friendly, clean, nontoxic, and economical process is
promised through the use of biological systems as nanofactories. Green synthesis
has gained attention because it produces size-controlled, resilient, and dispersed
nanomaterials with desired physicochemical qualities by reducing the ionic concentration through biological processes (Ying etal. 2022). Moreover, green synthesis
methods are progressively replacing physical and chemical methods to forestall
various setbacks such as excessive energy consumption, emissions of harmful and
toxic substances, and the prerequisite of complex equipment and synthesis
conditions.
Green synthesis makes use of organic resources and environmentally desirable
components, such as microbial cultures and plant parts (Yosri etal. 2021). Metal
nanoparticle synthesis utilizing microorganisms, specically actinomycetes, has
been attempted and succeeded in several cases. Several Streptomycetes species, for
instance, are capable of synthesizing both AgNPs and AuNPs (Ying etal. 2022). For
the conversion of nanoparticles, the proteins and polyphenols found in green sources
may serve as reducing agents. The advantages of green synthesis, such as environmentally sound, economical, and stable materials, emphasize the importance of
green nanoparticles (GNPs). However, there are some limitations such as increased
reaction time, reduced product quality, and difculties in raw material extraction.
8.3 Top-Down andBottom-Up Approaches forNP Synthesis
Two basic methods have been proposed for the synthesis of metal nanoparticles:
top-down and bottom-up (Fig.8.1). Top-down methodology controls the process of
manufacturing NPs by breaking down, mechanically grinding, or slicing bulk
Fig. 8.1 Basic approaches for synthesis of nanoparticle
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material into nanoscale particles, whereas the bottom-up approach synthesizes NPs
by assembling atoms at the nanoscale (Abid etal. 2022). The physical generation of
nanoparticles uses a top-down approach, whereas biological and chemical syntheses use a bottom-up approach. However, unlike physiochemical synthesis, biogenic
synthesis uses a range of bioactive metabolites and enzymes from microbial and
plant sources to catalyze the breakdown of metal substances to produce the correct
amount of NPs under ambient conditions. This is because metal ions are reduced
considerably more quickly using natural agents (Prakasham 2012).
S. Paul et al.
8.4 Actinomycetes asNanofactories
Actinomycetes, which are G
a high G+C (Guanine + Cytosine) content, distributed extensively in both terrestrial and marine environments (Thekkangil and Suchithra 2020; Thekkangil etal.
2021). These ubiquitous organisms are well-known for their ability to produce a
wide range of distinct secondary metabolites with a wide range of applications
(Aswani etal. 2019; Kumari etal. 2020). Owing to this potential, studies on the
bioprospecting of actinomycetes for various purposes have garnered signicant
interest. The production of nanoparticles by actinomycetes is a rapidly emerging
eld, with various species such as Streptomyces, Nocardia, Rhodococcus, and
Thermomonospora being known to produce nanometals (Sandhu and Goel 2023).
These organisms are capable of generating different types of nanoparticles, including silver, gold, iron, and copper, which possess various biological activities.
+ve
(gram-positive) fungi-like lamentous bacteria with
8.4.1 Extracellular andIntracellular Actinomycetes-Mediated
Biosynthesis ofNPs
Actinomycetes are known to adapt to extreme environments and tolerate a high
concentration of metals. They also have the capability to degrade inorganic elements into NPs. As part of their microbial defense strategy for cellular detoxication, these organisms are able to utilize metal ions from the surrounding environment,
which further convert the ions into elemental metal form by the process of enzymatic reduction (Kumari etal. 2020). Based on the site of manifestation, NP production can be categorized as extracellular or intracellular (Table8.1).
During extracellular synthesis, metal ions are reduced to their elemental form by
microbial enzymes and proteins. During the process of intracellular NP synthesis,
positively charged metal ions initially attract to the negatively charged carboxyl
groups in the microbial cell wall via electrostatic force of attraction. This makes it
possible for the metal ions to inltrate through the cellular membrane and undergo
further metallic reduction through intracellular proteins/enzymes within the cell
8 Actinomycetes asNanofactories: Synthesis andTherapeutic Applications
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Table 8.1 Examples of extracellular and intracellular actinomycetes-mediated nanoparticles
EC/
NPs Actinomycetes Sources
Copper Actinomycetes sp. Endophytes
IC References
EC/ICRajivgandhi etal. (2022)
(Palntaa)
Streptomyces sp. Soil EC/ICRajivgandhi etal. (2022)
Gold Rhodococcus sp. Water IC Salem etal. (2020)
S. viridogens HM10 Soil IC Rasool and Hemalatha (2017)
Silver S. coelicolor Soil – Chackaravarthi etal. (2023)
S. antimycoticus
L-1
Endophytes
(Plantb)
EC Salem etal. (2020)
Nocardiopsis sp. Water EC Chackaravarthi etal. (2023
Rhodococcus sp. Water IC Manimaran and Kannabiran
(2017)
S. albidoavus CNP10 Water EC/ICRasool and Hemalatha (2017)
143
S. aureofaciens MTCC
Water EC Golinska etal. (2014)
365
S. pseudogriseolus
Endophytes EC Fouda etal. (2020)
Acv-11
S. hygroscopicus
Water EC Rasool and Hemalatha (2017)
BDUS49
S. zaomyceticus
Oc-5
Endophytes
(Plantc)
IC Fouda etal. (2020)
S. rochei Water EC Chackaravarthi etal. (2023)
S. capillispiralis
Ca-1
Streptomyces sp.
VITBT7
Streptomyces sp.
Endophytes
EC Fouda etal. (2020)
(Plantd)
Soil EC Manimaran and Kannabiran
(2017)
Water EC Rajivgandhi etal. (2022)
VITPK1
Zinc Streptomyces sp. Soil EC/ICManimaran and Kannabiran
(2017)
Streptomyces sp. Soil/water EC Golinska etal. (2014)
S. coeruleorubidus Endophytes
EC Chackaravarthi etal. (2023)
(marine)
NP nanoparticle, EC extracellular, IC intracellular
a
medicinal plant Lonicera japonica; S Streptomyces, R Rhodococcus, N Nocardiopsis
b
Mentha longifolia L.
c
Oxalis corniculata L.
d
Convolvulus arvensis L.& Oxalis corniculata L.
(Venkateswaran etal. 2022). Several reports on both extracellular and intracellular
synthesis have been postulated, and in every hypothesis, the major enzymes included
are oxidoreductases such as sulte reductase, nitrate reductase, and cellular transporters (Koul etal. 2021). The intracellular process entails the transfer of metal ions
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inside the microbial cell with the help of transporter enzymes, whereas in the extracellular method, enzymatically reduced ions accumulate on the cell surface.
Actinomycetes-mediated intracellular NP production involves the reduction of
metal ions at the mycelia and cytoplasmic membrane (Kumari etal. 2020). The
synthesis of AgNPs is a typical example of an intracellular method for synthesizing
metal NPs. In one of the proposed mechanisms for AgNP synthesis by intracellular
mechanism, rst Ag+ ions got trapped on the surface of actinomycete cells through
electrostatic interactions with negatively charged carboxylate groups present in the
mycelial cell wall and are further reduced to metallic AgNPs in the nuclei by the
reductase enzyme. Extracellular synthesis in actinomycetes mostly occurs within
the nitrogen cycle via enzymatic reduction of nitrates to nitrites. It is commonly
recognized that the biogenic synthesis of AgNPs is signicantly inuenced by
NADH-dependent nitrate reductase (Mabrouk etal. 2021). The most acceptable
mechanism for AgNP synthesis is the bioreduction of Ag+ ions to metallic Ag by
nitrate reductase. This enzymatic metal-reduction reaction occurs in various electron shuttle chains (Koul etal. 2021). A related process for AuNPs is the reduction
of Au ions from aqueous AuCl4. It appears that electron transfer from NADH initiates the bioreduction of gold ions, which is catalyzed by NADH-dependent nitrate
reductase as a carrier for electrons. Au ions are further reduced to electrons, converted to gold, and subsequently to gold NPs (Kumari etal. 2020).
S. Paul et al.
Mechanisms ofNanoparticle Biosynthesis
The reduction of metal ions to their corresponding nanoparticles is facilitated by a
few secondary metabolites as well as by intracellular and extracellular enzymes
(Fig.8.2). The process can be divided into three phases. In the rst phase, metallic
ions are either absorbed within microbial cell membranes via cationic membranetransport systems or collected on the cell surface through electrostatic interactions
with the negatively charged cell wall. In the second phase, metal ions (M+) are
reduced to zero-valent metals (M°), catalyzed by oxidoreductase enzymes, such as
nitrate reductase. Which further results in the conversion to monovalent- or divalentmetal ions from zero valent states. Ultimately, the resultant NPs coalesced into a
variety of morphological forms, such as cubes, ovals, triangles, hexagons, and
spheres. The third step involves the stabilization of the nanoparticles via capping
agents to inhibit development and agglomeration while also regulating their size
and form (Messaoudi and Bendahou 2020).
8.4.2 Bioprocess ofNPs Using Actinomycetes
Bioprocesses can be classied into the downstream, fermentation, and upstream subcategories. Downstream processing includes inoculum preparation and medium formulation. Actinomycetes can be isolated from various sources, such as soil, water, and
8 Actinomycetes asNanofactories: Synthesis andTherapeutic Applications
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145
Fig. 8.2 Proposed nanoparticle synthesis mechanism: Figure illustrating the processes involved in
extracellular and intracellular synthesis of MtNPs. In extracellular NP synthesis, Metal ions (M+)
are adsorbed on the cell wall and further reduced to metals (M°) by extracellular oxidoreductase
enzyme. In the intracellular synthesis of MtNPs, M+ is transferred into the cytoplasm of the cell
and subsequently reduced by metabolic processes involving various oxidoreductase enzymes.
plant parts, or revived/activated from stock culture. Multiple media options exist for
the growth of actinomycetes, including Starch Casein Agar, Bennett Agar, International
Streptomycetes Project Mediums, and Chitin Agar (Salem etal. 2020). The cultivation
conditions must be optimized for optimal NP yield, including the appropriate culture
medium, pH, temperature, and agitation. Following a 3–4day incubation period at
28°C with shaking, mycelia were removed from the culture broth by ltration and
subsequently wash with sterile distilled water (Kumari etal. 2020). Both the supernatant and biomass were tested for NP synthesis, and it was observed that microorganisms were capable of synthesizing NPs extracellularly or intracellularly.
Extracellular synthesis involves mixing the cell-free supernatant with a metal
precursor salt solution, whereas intracellular synthesis involves mixing the washed
biomass with a metal precursor salt solution. The color formation after incubation
indicated the synthesis of nanoparticles. Centrifugation was then used to recover
and collect the precipitated nanoparticles as bottom pellets. If the nanoparticles are
synthesized inside the cell, an additional step of cell disruption is necessary, which
can be executed through repeated cycles of ultrasonication to release the NPs from
the cell for further recovery (Moholkar et al. 2021). Following recovery,
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