Nanoparticles from Microalgae and Their Biomedical Applications

Abstract

Over the years, microalgae have been a source of useful compounds mainly used as food and dietary supplements. Recently, microalgae have been used as a source of metabolites that can participate in the synthesis of several nanoparticles through inexpensive and environmentally friendly routes alternative to chemical synthesis. Notably, the occurrence of global health threats focused attention on the microalgae application in the medicinal field. In this review, we report the influence of secondary metabolites from marine and freshwater microalgae and cyanobacteria on the synthesis of nanoparticles that were applied as therapeutics. In addition, the use of isolated compounds on the surface of nanoparticles to combat diseases has also been addressed. Although studies have proven the beneficial effect of high-value bioproducts on microalgae and their potential in medicine, there is still room for understanding their exact role in the human body and translating lab-based research into clinical trials.

Full text

Citation: Sidorowicz, A.; Fais, G.;
Casula, M.; Borselli, M.; Giannaccare,
G.; Locci, A.M.; Lai, N.; Orrù, R.; Cao,
G.; Concas, A. Nanoparticles from
Microalgae and Their Biomedical
Applications. Mar. Drugs 2023,21,
352. https://doi.org/10.3390/
md21060352
Academic Editors: María
Carmen Rodríguez-Argüelles and
Noelia González-Ballesteros
Received: 29 April 2023
Revised: 3 June 2023
Accepted: 6 June 2023
Published: 7 June 2023
Copyright: © 2023 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
marine drugs
Review
Nanoparticles from Microalgae and Their Biomedical Applications
Agnieszka Sidorowicz 1,2, Giacomo Fais 1,2 , Mattia Casula 1,2, Massimiliano Borselli 3, Giuseppe Giannaccare 3,
Antonio Mario Locci 1,2, Nicola Lai 1,2 , Roberto Orrù1,2 , Giacomo Cao 1,2 and Alessandro Concas 1, 2, *
1Interdepartmental Centre of Environmental Science and Engineering (CINSA), University of Cagliari,
Via San Giorgio 12, 09124 Cagliari, Italy; a.sidorowicz@studenti.unica.it (A.S.); giacomo.fais@unica.it (G.F.);
m.casula28@studenti.unica.it (M.C.); antoniom.locci@unica.it (A.M.L.); nicola.lai@unica.it (N.L.);
roberto.orru@unica.it (R.O.); giacomo.cao@unica.it (G.C.)
2Department of Mechanical, Chemical and Materials Engineering, University of Cagliari, Via Marengo 2,
09123 Cagliari, Italy
3
Department of Ophthalmology, University Magna Grecia of Catanzaro, Viale Europa, 88100 Catanzaro, Italy;
mborselli93@gmail.com (M.B.); giuseppe.giannaccare@unicz.it (G.G.)
*Correspondence: alessandro.concas@unica.it
Abstract:
Over the years, microalgae have been a source of useful compounds mainly used as food
and dietary supplements. Recently, microalgae have been used as a source of metabolites that can
participate in the synthesis of several nanoparticles through inexpensive and environmentally friendly
routes alternative to chemical synthesis. Notably, the occurrence of global health threats focused
attention on the microalgae application in the medicinal field. In this review, we report the influence
of secondary metabolites from marine and freshwater microalgae and cyanobacteria on the synthesis
of nanoparticles that were applied as therapeutics. In addition, the use of isolated compounds on the
surface of nanoparticles to combat diseases has also been addressed. Although studies have proven
the beneficial effect of high-value bioproducts on microalgae and their potential in medicine, there is
still room for understanding their exact role in the human body and translating lab-based research
into clinical trials.
Keywords: microalgae; nanoparticles; synthesis; biomedical; anticancer; antimicrobial
1. Introduction
The increasing human population and life expectancy are causing a change in the
leading cause of death, such as heart conditions, cancer, or pulmonary diseases [
1
]. The
shift is pushing the healthcare system to find new solutions to these problems; however,
hospitals can be a source of nosocomial infections which are especially dangerous for
immunocompromised patients. Another major issue is microbial antibiotic resistance due
to antibiotics abuse which can lead to the emergence of life-threatening diseases [2].
New promising solutions are introduced by recent developments in nanotechnology
which are focused on the manipulation of matter having a characteristic size lower than
100 nm in at least one dimension. The prevailing quantum effect at such a scale can give
rise to multiple applications of the products. The prepared materials at the nanoscale might
have different features than their bulk equivalents which show the potential for obtaining
various properties even within the same element. Several methods are used to synthesize
nanoparticles (NPs) such as physical, chemical, or biological routes. Among them, great
attention is being paid to biological synthesis due to its low toxicity and biocompatibility,
which are crucial for biomedical applications.
One group of organisms used for biological synthesis is microalgae due to their rapid
increase in biomass, the independence on arable land, and the abundance of valuable
metabolites [
3
]. Moreover, microalgae can be cultivated also in the wastewater indepen-
dently from seasonal breaks which is an important economical aspect [
4
]. The bioactive
substances derived from secondary metabolism such as proteins, polysaccharides, lipids,
Mar. Drugs 2023,21, 352. https://doi.org/10.3390/md21060352 https://www.mdpi.com/journal/marinedrugs

Mar. Drugs 2023,21, 352 2 of 18
vitamins, and pigments have displayed their great potential for many applications [
5
]
The identified compounds are mainly used for their nutritional value; however, they can
participate in the synthesis of various NPs used for biomedical applications.
Although the use of metabolites in the synthesis of various NPs can result in obtaining
highly valuable materials, the mechanism behind it is still unclear [
6
,
7
]. In this review, the
state of the art related to the participation of microalgal compounds in NPs synthesis is
analyzed considering the location of the process (intra- or extracellular). Moreover, recent
biomedical applications are taken into account to show the potential of microalgae to be
applied in medicine. The recent advancements in the synthesis of NPs using microalgae
have been summarized by connecting their synthesis method with resulted performance.
Therefore, the work can provide future prospects for the optimization of the synthesis of
highly valuable materials using microalgae.
2. Biological Synthesis
2.1. Microalgal Metabolites
Microalgae are single-celled, photosynthetic organisms found in both marine and
freshwater ecosystems. The classification of these organisms is based on the properties
such as pigmentation, photosynthetic membrane organization, chemical nature of the
photosynthetic storage products, or morphological features [
6
]. The groups are polyphyletic
and highly diverse, with both procaryotic and eucaryotic organisms. The most abundant
microalgae are Cyanophyceae (blue-green algae), Bacillariophyceae (including the diatoms),
and Chlorophyceae (green algae), with 50,000 estimated existing species, out of which
30,000 species were investigated [
6
]. Microalgae produce a variety of substances including
proteins, carbohydrates, lipids, nucleic acids, vitamins, and minerals [
8
–
10
]. The cellular
content of each group varies depending on the specific strain and their physiological
reactions to biotic and/or abiotic factors such as light intensity, photoperiod, temperature,
medium composition, and growth phase [
11
,
12
]. The typical compounds reported so far
participating in the synthesis of nanoparticles are proteins, carbohydrates, and lipids.
The main mechanism involved in the biosynthesis of NPs deals with different metabo-
lites of microalgae that can reduce precursor metal ions into a zerovalent state (Figure 1).
The process involves (i) the activation phase, when the metal ion is reduced, and
nucleation occurs, followed by (ii) the growth phase, with an amalgamation of formed
unit cells into crystallites which is concluded in (iii) the termination phase, where NPs
having different shapes and sizes are thermodynamically stable [
13
]. Other factors such as
temperature, pH, or metal ion concentration could affect the synthesis process; however, the
participation of microalgae metabolites is crucial to understand the connection between the
synthesis procedure and the properties of the obtained product [
14
,
15
]. Moreover, during
microalgae cultivation sunlight and carbon dioxide can be acquired from the surroundings
while the nutrients could be converted from the wastewater to form biomass which provides
the new routes for economic sustainability [16].
2.1.1. Proteins
Proteins are an important component in the structure and metabolism of microalgae.
They are an integral part of the cellular membrane and light-harvesting complex as well
as they participate as enzymes in numerous catalytic reactions [
17
,
18
]. Several species of
microalgae are studied due to their high protein content ranging from 42–70% in some
cyanobacteria and up to 58% for Chlorella vulgaris dry weight [19,20].
The involvement of proteins in the synthesis of nanoparticles is usually investigated
using Fourier Transform Infrared Spectroscopy (FTIR) based techniques. The reduction
role of the proteins is demonstrated during the oxidation of the
−
CHO to
−
COOH group,
while NH
2
groups usually play capping functions through residual amino acids such
as cysteine, tyrosine, and tryptophan [
21
]. In the study by Chokshi et al., the spectra
between the prepared extract of Acutodesmus dimorphus and prepared Ag NPs were com-
pared, showing the role of amide linkage in the stabilization of Ag NPs by peptides and

Mar. Drugs 2023,21, 352 3 of 18
proteins [
22
]. The obtained NPs were spherical with 2–20 nm in size. Moreover, the overlap-
ping peaks between the extract and the product suggest their coating properties ensure their
stabilization and prevent agglomeration. The surface of NPs might be further modified
by sulfonated polysaccharides with proteins that provide a link between nanoparticles
and coating molecules [
21
]. Proteins also possess a strong affinity to bind to metal ions
that act as reducing agents [
21
]. Similar findings have been reported for AgCl NPs from
Chlorella vulgaris and Ti NPs from marine microalgae Phaeodactylum tricornutum [
23
,
24
].
Although the exact mechanism of the synthesis is unknown, the statistical experimen-
tal design approach has been studied by using response surface methodology for future
large-scale production.
Mar. Drugs 2023, 21, x FOR PEER REVIEW 3 of 19
Figure 1. Mechanism of NPs synthesis by microalgae.
The process involves (i) the activation phase, when the metal ion is reduced, and
nucleation occurs, followed by (ii) the growth phase, with an amalgamation of formed
unit cells into crystallites which is concluded in (iii) the termination phase, where NPs
having different shapes and sizes are thermodynamically stable [13]. Other factors such as
temperature, pH, or metal ion concentration could affect the synthesis process; however, the
participation of microalgae metabolites is crucial to understand the connection between the
synthesis procedure and the properties of the obtained product [14,15]. Moreover, during
microalgae cultivation sunlight and carbon dioxide can be acquired from the surroundings
while the nutrients could be converted from the wastewater to form biomass which provides
the new routes for economic sustainability [16].
2.1.1. Proteins
Proteins are an important component in the structure and metabolism of microalgae.
They are an integral part of the cellular membrane and light-harvesting complex as well
as they participate as enzymes in numerous catalytic reactions [17,18]. Several species of
microalgae are studied due to their high protein content ranging from 42–70% in some
cyanobacteria and up to 58% for Chlorella vulgaris dry weight [19,20].
The involvement of proteins in the synthesis of nanoparticles is usually investigated
using Fourier Transform Infrared Spectroscopy (FTIR) based techniques. The reduction
role of the proteins is demonstrated during the oxidation of the −CHO to −COOH group,
while NH2 groups usually play capping functions through residual amino acids such as
cysteine, tyrosine, and tryptophan [21]. In the study by Chokshi et al., the spectra between
the prepared extract of Acutodesmus dimorphus and prepared Ag NPs were compared,
showing the role of amide linkage in the stabilization of Ag NPs by peptides and proteins
[22]. The obtained NPs were spherical with 2–20 nm in size. Moreover, the overlapping
peaks between the extract and the product suggest their coating properties ensure their
stabilization and prevent agglomeration. The surface of NPs might be further modied by
sulfonated polysaccharides with proteins that provide a link between nanoparticles and
Figure 1. Mechanism of NPs synthesis by microalgae.
2.1.2. Carbohydrates
Similar to proteins, carbohydrates display both structural and metabolic properties.
Mono- and oligosaccharides can be found attached to proteins or lipids, forming glycopro-
teins or glycolipids, while polysaccharides are the major structural component of the cell
wall [
25
]. Moreover, glucose and starch-like energy storage products are obtained during
photosynthesis as the primary carbon-containing molecules in microalgae [
26
]. Cyanophytes
were reported to accumulate glycogen, while other species form semi-amylopectin [
27
].
Two glucose polymers, amylopectin, and amylose are starch components of Chlorophyta;
however, Rhodophyta synthesizes a carbohydrate polymer known as floridean starch [
10
,
28
].
Diatoms produce chrysolaminarin composed of
β
(1,3) and
β
(1,6) linked glucose units
which can accumulate around 7% of their total carbon content in the optimal conditions
and up to 80% under strong nutrient depletions [29,30].
Carbohydrates are rich in reducing groups, such as hydroxy and carboxy, which
can bind and reduce metal atoms, thereby acting as reducing agents. Moreover, due to
the supramolecular interactions by inter- and intra-molecular hydrogen bonding, they
can stabilize formed nanoparticles and prevent further agglomeration, acting as capping
agents [
16
,
31
]. The potential of secretory carbohydrates from C. vulgaris was tested by

Mar. Drugs 2023,21, 352 4 of 18
the removal of biomass from the culture and used for the synthesis of FeOOH NPs [
32
].
The synthesis process using carbohydrates was compared with the chemical route with
sodium hydroxide acting as a precipitating agent. The carbohydrates were reported to
be involved in the nucleation process by chelation of iron ions to prevent monotonic
nucleation and limit nuclei size, which is further controlled in the growth phase to inhibit
large particle formation. The secretory carbohydrates were described mainly for their
reducing properties rather than being capping agents. The obtained NPs were spherical
with size range 8–17 nm.
The exopolysaccharides from Botryococcus braunii and Chlorella pyrenoidosa were also
tested for the synthesis of Ag NPs [
33
]. The polysaccharides performed both reducing
and capping functions and were bound to the Ag NPs surface through carboxy and
hydroxy groups. A similar size range was reported of 5–15 nm. In a study by Jakhu
et al., Au NPs synthesized from Chlorella sp. polysaccharides were compared with Au
NPs synthesized using citrate to compare their properties [
34
]. Both products exhibited a
controlled size range; however, AuNPs from polysaccharides were stable in the pH range
2–12 while citrate-Au NPs were stable only at basic pH values. The NPs synthesized
using polysaccharides were significantly bigger than citrate with sie ranges 30–40 nm and
10–15 nm, respectively. Furthermore, citrate-Au NPs were forming agglomerates in a
30-fold lower concentration of NaCl, further proving the stabilization of the surface of Au
NPs by microalgal polysaccharides.
2.1.3. Lipids
Secondary to polysaccharide, lipids function as energy reservoirs as well as struc-
tural components of the cell membranes. In microalgae, lipids are mainly composed
of (i) neural lipids such as free fatty acids, acylglycerols, and carotenoids, and (ii) polar
lipids including phospholipids and galactolipids [
35
]. The polar lipids fraction can sig-
nificantly increase during exponential growth; however, during stationary phase when
the nutrient availability is limited under stress conditions, they can produce triacylglyc-
erols [
36
]. The fatty acid content is composed of a mixture of C16 and C18 saturated and
unsaturated fatty acids with longer carbon-chains including omega fatty acids. Saturated
fats are stored in neutral lipid bodies while unsaturated fats are connected with polar
lipids in membranes maintaining membrane fluidity under fluctuating cultivation con-
ditions [
37
,
38
]. The overall lipid fraction can represent up to 20–50% of the dry biomass,
depending on the microalgal species and cultivation conditions such as nutrient availability,
salinity, light intensity, and growth phase [
39
]. During nutrient depletion, the neutral
lipid, and polysaccharide content can increase at the expense of proteins [
40
]. Lipids re-
ceive the greatest attention for extraction followed by the production of biodiesel whereas
polyunsaturated fatty acids are used for their nutraceutical value.
In contrast to water, which is commonly used for synthesis, the involvement of
lipids requires the usage of different solvents. Kashyap et al. utilized ethanolic extract to
synthesize Ag/AgCl NPs from Chlorella sp., Lyngbya putealis,Oocystis sp., and Scenedesmus
vacuolatus [
41
]. During the optimization process, Oocystis sp. did not manage to produce
NPs while Chlorella sp. extract resulted in the synthesis of Ag/AgCl NPs of the smallest
size. The study highlighted the role of lipids and proteins along with the hydroxy group
stretching movements in the Ag/AgCl NPs formation with the size range of 10–20 nm. In a
study by Gusain et al., lipids and carbohydrates were extracted separately from Acutodesmus
obliquus and used for the synthesis of carbon dots by the microwave thermal method [
42
].
The products had a size range of 1.2–11 nm. The carbon source did not alter the fluorescence
behavior; however, the exact interactions during synthesis were not studied. The optical
properties changed with the addition of acetone which demonstrated the potential of
using different solvents for the synthesis. The role of lipids is hypothesized mainly as
capping agents.

Mar. Drugs 2023,21, 352 5 of 18
2.2. Intracellular Synthesis
Depending on the location where NPs are formed, the corresponding synthesis can
be divided into intracellular or extracellular routes. During the former one, live cultures
are exposed to the metal precursor, and charged metal ions are transported by negatively
charged sites of the cell wall [
43
]. The trapped ions undergo reduction and form NPs of
various sizes and morphologies inside the cell, which require various steps of purification
from biomass.
In a study by Li et al., Chromochloris zofingiensis culture was used to prepare Au
NPs [
44
]. The cells after synthesis were characterized showing a peak characteristic for
Au NPs in the UV-vis spectrum, which was also confirmed by transmission and scanning
electron microscopy findings. The proposed mechanism involves chelation by negatively
charged functional groups in the cell wall such as –COOH, –OH, and –OSO
3
H followed
by diffusion into the cytosol and reduction by electrons generated from photosynthetic
electron transport using enzymes. Other species with different cell wall structures were
also investigated for the synthesis mechanism. Instead, Euglena gracilis species cell wall
possesses a glycoprotein-containing pellicle that allowed metal ions to easily penetrate
the cell. On the contrary, the marine microalga Nitzschia laevis is composed of a rigid cell
wall containing amorphous hydrated porous silica frustule acting as a barrier to reduce ion
diffusion into the cells. In addition, Raman spectroscopy was explored as a tool to identify
and quantify biomass components in microalgae.
The effect of Ag/AgCl NPs synthesis on chlorophyll and lipid accumulation was
studied on freshwater microalgae Scenedesmus sp., showing a decrease of 20–35% after
120 h [
45
]. However, the cells treated with 0.5 mM AgNO
3
showed a 75.86% increase in
palmitic acid due to the stress induced by Ag/AgCl NPs. Thus, the cells were able to synthe-
size Ag/AgCl NPs and improve the quality of biodiesel production. Intracellular synthesis
was also used to obtain CdSe quantum dots from C. pyrenoidosa and S. obliquus [
46
]. First,
selenium ions were introduced to the culture to generate selenium precursors within the
photosynthetic electron transport system and after 12 h were combined with cadmium ions
to form CdSe quantum dots. The algal cells were damaged during the process probably
because of precursors reducing enzymatic activity and cell vitality. The intracellular syn-
thesis process requires proper optimization to ensure a high yield while maintaining a low
toxicity profile.
2.3. Extracellular Synthesis
Extracellular synthesis utilizes either the secreted molecules such as polysaccharides
or involves processing of the biomass to produce extract which is utilized for synthesis
of NPs [
47
]. This route is considered more convenient as NPs are easily purified from the
solution. In addition, it allows for further modification of metabolites participating in the
synthesis by varying adopted solvents, concentration, time, or pH [48,49].
The cell-free filtrate from freshwater microalgae S. obliquus culture with different nitro-
gen sources were used to extracellularly synthesize Ag NPs [
50
]. The study concentrated
on the activity of reductases, nitrogen, and sulfate, on Ag NPs synthesis depending on
the composition of the medium. The enzymes are conjugated with electron donors and
act as reducing agents. Moreover, the activity influenced not only the yield but also the
properties or the obtained Ag NPs especially their size. Consequently, their size inversely
correlated antimicrobial activity which demonstrates the importance of metabolites during
the synthesis.
The cell-free C. vulgaris culture was investigated for different factors affecting Ag NPs
synthesis including time, extract/precursor ratio, temperature, pH, precursor molarity, and
incubation conditions [
51
]. The optimal conditions were maximum incubation time (24 h),
silver nitrate/extract ratio (8:2), 37
◦
C, pH 12, 3 mM silver nitrate, and shaking. The study
shows the potential of varying multiple factors during the synthesis of NPs, which might
result in products with different properties. In a study by Shalaby et al., algal biomass
was processed to obtain an extract which provided metabolites implicated in the synthesis

Mar. Drugs 2023,21, 352 6 of 18
of iron oxide NPs [
52
]. The synthesis was perfected by varying the ratio of precursor to
extract, and the product with the highest absorbance was selected for further application.
3. Biomedical Applications of Microalgal NPs
The synthesis route free from toxic waste, as well as economical and environmentally
friendly aspects, show the great potential of NPs synthesized from marine and freshwater
microalgae for a variety of applications (Figure 2). The presence of naturally occurring
biomolecules improves their biocompatibility in comparison with other synthesis routes,
and, thus, can be used for biomedical applications. In addition, the growth parameters and
metabolites content can be easily altered to obtain a variation in the morphology of NPs for
diverse utilization.
Mar. Drugs 2023, 21, x FOR PEER REVIEW 6 of 19
2.3. Extracellular Synthesis
Extracellular synthesis utilizes either the secreted molecules such as polysaccharides
or involves processing of the biomass to produce extract which is utilized for synthesis of
NPs [47]. This route is considered more convenient as NPs are easily puried from the
solution. In addition, it allows for further modication of metabolites participating in the
synthesis by varying adopted solvents, concentration, time, or pH [48,49].
The cell-free ltrate from freshwater microalgae S. obliquus culture with dierent
nitrogen sources were used to extracellularly synthesize Ag NPs [50]. The study
concentrated on the activity of reductases, nitrogen, and sulfate, on Ag NPs synthesis
depending on the composition of the medium. The enzymes are conjugated with electron
donors and act as reducing agents. Moreover, the activity inuenced not only the yield
but also the properties or the obtained Ag NPs especially their size. Consequently, their
size inversely correlated antimicrobial activity which demonstrates the importance of
metabolites during the synthesis.
The cell-free C. vulgaris culture was investigated for dierent factors aecting Ag NPs
synthesis including time, extract/precursor ratio, temperature, pH, precursor molarity,
and incubation conditions [51]. The optimal conditions were maximum incubation time
(24 h), silver nitrate/extract ratio (8:2), 37 °C, pH 12, 3 mM silver nitrate, and shaking. The
study shows the potential of varying multiple factors during the synthesis of NPs, which
might result in products with different properties. In a study by Shalaby et al., algal biomass
was processed to obtain an extract which provided metabolites implicated in the synthesis of
iron oxide NPs [52]. The synthesis was perfected by varying the ratio of precursor to extract,
and the product with the highest absorbance was selected for further application.
3. Biomedical Applications of Microalgal NPs
The synthesis route free from toxic waste, as well as economical and environmentally
friendly aspects, show the great potential of NPs synthesized from marine and freshwater
microalgae for a variety of applications (Figure 2). The presence of naturally occurring
biomolecules improves their biocompatibility in comparison with other synthesis routes,
and, thus, can be used for biomedical applications. In addition, the growth parameters
and metabolites content can be easily altered to obtain a variation in the morphology of
NPs for diverse utilization.
Figure 2. Applications of NPs from microalgae in biomedical elds.
Figure 2. Applications of NPs from microalgae in biomedical fields.
The major drawback connected with the synthesis of NPs using a biological approach
is their possible wide size distribution or heterogenous morphology. Thus, the obtained
product might be difficult to assess in the context of molecular interactions within tissues
or organs. The reason behind the differences might be connected with the complexity of
molecules participating in the synthesis which reduce the metal ions with varying efficiency.
However, the effect can be minimized by utilizing selected classes of secondary metabolites
which can also improve the knowledge of their role in the synthesis. Currently, the lack of
understanding behind the synthesis mechanism and the long-term effects of NPs are also
limiting factors in the biological approach.
3.1. Anticancer Activity
The anticancer activity of NPs synthesized from microalgae has been extensively
investigated. The general mechanism associated with anticancer activities of NPs is related
to ROS generation (Figure 3). In a recent study by Hamida et al., Ag NPs were synthesized
using freshwater strain Coelastrella aeroterrestrica and their anticancerous activity against
four malignant cell lines was compared with chemically synthesized Ag NPs and the anti-
cancer drug 5-fluorouracil [
53
]. The results showed the highest antiproliferative activity of
microalgal Ag NPs against MCF-7, MDA, HCT-116, and HepG2 cell lines with low toxicity
against non-cancerous cell lines, compared to the other tested. The activity was attributed
to its small size, high stability, less agglomeration, and surface chemistry. However, the

Mar. Drugs 2023,21, 352 7 of 18
mechanistic pathway inside the cancer cell and the pharmacokinetic nature of Ag NPs have
yet to be explored.
Mar. Drugs 2023, 21, x FOR PEER REVIEW 7 of 19
The major drawback connected with the synthesis of NPs using a biological approach
is their possible wide size distribution or heterogenous morphology. Thus, the obtained
product might be dicult to assess in the context of molecular interactions within tissues
or organs. The reason behind the dierences might be connected with the complexity of
molecules participating in the synthesis which reduce the metal ions with varying
eciency. However, the eect can be minimized by utilizing selected classes of secondary
metabolites which can also improve the knowledge of their role in the synthesis.
Currently, the lack of understanding behind the synthesis mechanism and the long-term
eects of NPs are also limiting factors in the biological approach.
3.1. Anticancer Activity
The anticancer activity of NPs synthesized from microalgae has been extensively
investigated. The general mechanism associated with anticancer activities of NPs is
related to ROS generation (Figure 3). In a recent study by Hamida et al., Ag NPs were
synthesized using freshwater strain Coelastrella aeroterrestrica and their anticancerous
activity against four malignant cell lines was compared with chemically synthesized Ag
NPs and the anticancer drug 5-uorouracil [53]. The results showed the highest
antiproliferative activity of microalgal Ag NPs against MCF-7, MDA, HCT-116, and
HepG2 cell lines with low toxicity against non-cancerous cell lines, compared to the other
tested. The activity was aributed to its small size, high stability, less agglomeration, and
surface chemistry. However, the mechanistic pathway inside the cancer cell and the
pharmacokinetic nature of Ag NPs have yet to be explored.
Figure 3. Mechanism of NPs anticancer activity through ROS generation.
The inuence of various nanocomposites (NCs) synthesized from microalgae on
cancer cell lines was also tested. In two separate studies, S. obliquus extract was used to
conjugate Ag NPs with PtFe2O4 NPs (PtFe2O4@Ag) and GaFe2O4 NPs (GaFe2O4@Ag)
[54,55]. The cytotoxic properties of PtFe2O4@Ag NCs were related to cross-linking between
Pt and DNA to disrupt transcription and replication as well as reactive oxygen species
(ROS) generation, lipid peroxidation, and enhanced glutathione (GSH) degradation by Ag
NPs. Similarly, Ga in GaFe2O4@Ag NCs can alter iron metabolism, resulting in the
appearance of chromatin fragmentation and apoptotic bodies that induce cell apoptosis.
The prepared PtFe2O4@Ag NCs displayed a more prominent anticancerous activity than
the GaFe2O4@Ag NCs; however, it was lower than cisplatin, a gastric cancer medication
used as a control. Furthermore, MgFe2O4@Ag NCs were synthesized using an extract of
C. vulgaris showing anticancer activity through a similar apoptotic pathway [56]. The role
Figure 3. Mechanism of NPs anticancer activity through ROS generation.
The influence of various nanocomposites (NCs) synthesized from microalgae on cancer
cell lines was also tested. In two separate studies, S. obliquus extract was used to conjugate
Ag NPs with PtFe
2
O
4
NPs (PtFe
2
O
4
@Ag) and GaFe
2
O
4
NPs (GaFe
2
O
4
@
Ag) [54,55].
The
cytotoxic properties of PtFe
2
O
4
@Ag NCs were related to cross-linking between Pt and
DNA to disrupt transcription and replication as well as reactive oxygen species (ROS)
generation, lipid peroxidation, and enhanced glutathione (GSH) degradation by Ag NPs.
Similarly, Ga in GaFe
2
O
4
@Ag NCs can alter iron metabolism, resulting in the appear-
ance of chromatin fragmentation and apoptotic bodies that induce cell apoptosis. The
prepared PtFe
2
O
4
@Ag NCs displayed a more prominent anticancerous activity than the
GaFe
2
O
4
@Ag NCs; however, it was lower than cisplatin, a gastric cancer medication
used as a control. Furthermore, MgFe
2
O
4
@Ag NCs were synthesized using an extract of
C. vulgaris showing anticancer activity through a similar apoptotic pathway [56]. The role
of MgO in the NCs was attributed to the enhancement of the magnetic properties and
disruption of the cell membrane. Further studies are needed to enhance NCs activity and
describe their effects in various cancerous and non-cancerous cell lines. Other reported
NPs synthesized using microalgae tested for anticancer activity are presented in Table 1.
Microalgae can serve as a source of valuable compounds that not only take part in the
synthesis of anticancer drugs but also exhibit anticancer properties themselves. However,
their proper action requires maintaining their structure which could be damaged due
to chemical or physical factors. In a study by ˙
Inan et al., the marine microalga C. vari-
abilis and C. pyrenoidosa oil extracts were encapsulated into NPs using an electrospraying
technique [
57
]. Encapsulated oil extracts showed higher biocompatibility, while only C. vari-
abilis oil extract showed improved anticancer properties compared to the non-encapsulated
form. The activity changed in a dose-dependent manner with observed changes in the
morphology of the cells. The research on optimizing encapsulation techniques could lead
to the development of novel anticancer drugs of microalgal origin.

Mar. Drugs 2023,21, 352 8 of 18
Table 1. Anticancer activity of NPs synthesized from microalgae.
Types of NPs Microalgae
Species Used
General
Environment
Size and Morphology
of NPs
Tested Cancerous
Cell Lines Ref.
Silver NPs (Ag NPs)
Arthrospira platensis
marine, freshwater 2.23–14.68 nm, spherical A549, HCT, Hep2
and WISH [58]
Copper NPs (CuO NPs)
Arthrospira platensis
marine, freshwater 3.75–12.4 nm, spherical A549, HCT, Hep2
and WISH [58]
Silver NPs (Ag NPs)
Arthrospira platensis
marine, freshwater 30 nm, spherical A-549, MCF-7 [59]
Au/cellulose
nanocomposite Chlorella vulgaris freshwater 113–203 nm, spherical A-549 [60]
Gold NPs (Au NPs) Dunaliella salina marine 22.4 nm, spherical MCF-7 [61]
Carbon quantum dots Pectinodesmu sp. freshwater 67 nm, spherical HCC 1954, HCT 116 [62]
Silver NPs (Ag NPs) Trichodesmium
erythraeum marine 26.5 nm, cubical MCF-7, He La [63]
Silver NPs
(Ag2O/|AgO NPs) Oscillatoria sp. Freshwater 4.42–48.97 nm,
quasi-spherical CaCo-2, HeLa [64]
3.2. Biomedical Sensor
Algal-synthesized NPs have shown the potential to be utilized for the detection of
dopamine, which plays an essential role in renal, central nervous, cardiovascular, and
hormonal regulation. Moreover, it acts as a neurotransmitter in reward and movement
regulation in the brain and its abnormal concentrations can cause neurological disorders.
The study by Huang et al. utilized ethanolic extract of the marine microalga Spirulina to
synthesize Au NPs with 11–14 nm in size for dopamine detection [
65
]. The proteins and
polysaccharides from the extract participated in the synthesis as reducing and capping
agents with an abundance of –OH and –COOH groups on the Au NPs surface. The
interactions between functional groups of dopamine cause changes in Au NPs’ surface
allowing dopamine to be adsorbed, thus forming additional hydrogen, ester, and amide
bonds with other neighboring Au NPs. In that way, dopamine can act as a linkage between
Au NPs with a new enhanced plasmon resonance absorption peak changing the color of
the solution from wine red to blue-black. The set-up was tested to evaluate selectivity and
anti-interference showing specificity in the presence of various components such as amino
acids, salts, and glucose. However, when the Ca
2+
content was above 200
µ
M, it could form
complexes with the oxygen of the –COOH group causing Au NPs aggregation, which can
be thus prevented by keeping Ca
2+
concentration at lower levels. When tested in human
urine, the method was proven to be accurate and precise with a simple detection protocol.
Another compound that could be detected by NPs synthesized by microalgae is
atropine, a tropane alkaloid used to treat low heart rate (bradycardia), an overdose of
cholinergic drugs, and cholinergic poisoning, as well as to help reduce saliva, mucus, or
other secretions during surgery [
66
]. The marine microalga S. platensis biomass was used
in a process to obtain Ag NPs on their surface as a coating material with an average size
of 59 nm. The product was placed on the electrode with and without a binder to test its
properties. The results revealed high electrocatalytic activity in atropine determination with
response fluctuation by a change in pH value. The selectivity, stability, and reproducibility
test proved the potential of the material as a stable sensor. In addition, the performance was
confirmed by using atropine sulfate ampoule and water as real samples further elaborating
the high accuracy and recovery rate of the prepared electrode.
In addition to neurotransmitters or pharmaceuticals, the glucose level can also be
monitored by NPs from microalgae [
67
]. The biomass of C. vulgaris was processed using
the hydrothermal method, acid hydrolysis assisted by ultrasonics, and was followed by the
hydrothermal method to obtain carbon dots. After each hydrothermal treatment, the carbon
dots were collected to compare their synthesis method with properties. Acidic hydrolysis
was considered crucial to degrade starch and cellulose to reduce sugars to increase yield
and prevent interactions between Fe
2+
and residual –NH groups on the surface. In addition,

Mar. Drugs 2023,21, 352 9 of 18
low pH helped to prevent quenching during the fluorescence response of carbon dots. The
sensing mechanism was tested for Fe
3+
and H
2
O
2
and then applied to determine the
glucose level in blood samples based on the reaction of the glucose oxidase enzyme and the
quenching of fluorescence under optimized conditions. The results revealed high sensitivity
and selectivity of the sensor obtained with potential use in diagnostics.
3.3. Drug Delivery
Targeted drug delivery is a strategy to selectively administer pharmaceuticals into
specific areas to maximize its efficacy while avoiding side effects. In a study by Wang et al.,
microrobots were prepared for drug loading, targeted delivery, and chemo-photothermal
therapy [
68
]. First, S. platensis cells were used as a template for core–shell-structured Pd@Au
NPs synthesis by electroless deposition to act as photothermal conversion agents to allow
laser-triggered degradation to release the drug. Additionally, Fe
3
O
4
NPs were deposited on
the surface via a sol–gel process for the magnetic actuation of the material. The anticancer
drug doxorubicin was loaded on the prepared structure to allow their chemotherapeutic
effect. The obtained microrobots exhibited significant propulsion under a magnetic field
and can be structurally disassembled into individual components under laser irradiation
to be used for pH- and laser-triggered drug release. Moreover, Au and Fe
3
O
4
NPs can be
utilized as CT and MR imaging contrast agents, demonstrating the real-time monitoring
of the treatment. Considering their small size, the authors suggest oral administration for
gastrointestinal cancer therapy.
Doxorubicin-loaded microrobots were also synthesized using the marine microalga
Thalassiosira weissflogii as a template to substitute mesoporous silica NPs [
69
]. The magnetic
Fe
3
O
4
NPs were adhered to the template surface for the magnetic actuation properties.
Using an external magnetic field, the movement of microrobots can be controlled in tra-
jectories by changing the frequency, allowing microrobots to move through channels of
various diameters. The clustering behavior enables the microrobots to carry high-load to
the target position with subsequent release in a pH-sensitive manner. The microrobots
were tested on MCF-7 human breast cancer cells demonstrating their efficiency. Further
studies in vivo are required to test their application.
In addition, the marine microalga T. weissflogii was utilized in a separate study as
a template for the delivery of curcumin-loaded drugs for anticancer and antibacterial
properties [
70
]. The nanoporous architecture of T. weissflogii frustules provided a cage-
like structure for curcumin adsorption which was stabilized by interactions of functional
groups. The potential of T. weissflogii to be used for drug delivery systems shows its future
application for the treatment of a wide range of diseases.
Microalgae can act not only as a template for drug delivery but can also be a source
of valuable molecules that display therapeutic activities. Freshwater microalgae Botry-
ococcus braunii and Microcystis aeruginosa oil, rich in polyunsaturated fatty acids, were
loaded into NP electrosprayed with alginate/polyvinylidene (PVA) and tested for an-
tibacterial activity with stabilized release [
71
]. The encapsulation protected bioactive and
antioxidant properties of the oil which shows the potential for their storage and use for
commercial application.
3.4. Immunomodulatory Action
The study by Chandrarathna et al. investigated the effect of pectin, a polysaccharide
isolated from the marine strain Spirulina maxima [
72
]. The nano form of the compound
was obtained through sonication to avoid possible modifications from the addition of
chemicals. The material was tested on the mice model in comparison with non-sonicated
pectin, with an average particle size of 64.11 nm and 152.90 nm for sonicated and original
pectin, respectively. The results revealed increased weight gain in the nano-pectin-treated
mice due to the improved digestibility and availability of nutrients with the small particle
size. In addition, small particle size provided a higher surface area for microbial growth in
the intestines than the original longer pectin molecules. The mice treated with nano-pectin

Mar. Drugs 2023,21, 352 10 of 18
displayed an increased density of goblet cells in the gut barrier which blocks the access of
pathogenic microbes to the gut epithelium as well as showed higher expression of intestinal
alkaline phosphatases, which provide an anti-inflammatory effect. It was assumed that in
order to better understand the interactions between pectin and gut microbiome long-term
studies are required.
The immunomodulatory effect of nano-pectin from S. maxima was also tested in
zebrafish both
in vitro
and
in vivo
[
73
]. The low toxicity of nano-pectin was hypothesized
to be due to the morphological properties as well as the surface functionalization. In
addition, the transcriptional analysis of immune-related genes was performed to further
describe the immunomodulatory action. The results showed increased expression of
cytokines and antioxidants that participate in the innate immune response. Altogether,
nano-pectin was crucial for stress tolerance and anti-inflammatory actions on the molecular
level for disease resistance. Although the wound healing activity was not remarkable,
nano-pectin was hypothesized to engage the immune system during the process. The
studies on mice and zebrafish show the potential of nano-pectin to regulate the immune
system response in different organisms; however, further investigations are required to
understand the underlying mechanisms behind those actions.
Polysaccharides from a different marine species, Arthrospira fusiformis, were also tested
for the synthesis of Ag NPs and their immunomodulatory properties [
74
]. The prepared
material was tested
in vitro
on Pseudomonas aeruginosa as well as
in vivo
on P. aeruginosa-
infected rat models. The Ag NPs were on average around 10 nm in size with homogenous
distribution. The findings showed a strong antibacterial effect due to the breakage of the
outer membrane of P. aeruginosa, affecting cell permeability with the disruptions created
called “pits” that lead to cell lysis, as investigated by
in vitro
studies. The algal coating
was suggested to interact with human serum protein by slightly reducing its concentration
resulting in increased cellular uptake and intracellularly killing of bacteria. Moreover, Ag
NPs used as wound dressing of P. aeruginosa-infected areas enhanced the wound healing
by cytokine modulation and reducing the inflammation. The proposed mechanism involves
the liberation of Ag
+
from Ag NPs and their sequestration by H
2
S-synthesizing enzymes in
the macrophage resulting in the formation of Ag
2
S and lowering the inflammation. After
seven days of treatment, the tissue structure in rats was restored, which shows the potential
of Ag NPs from algae for future treatments which should be assessed during clinical trials.
3.5. Antibacterial Activity
Antibiotic resistance can be counted among the worst threats to human health world-
wide. Overuse of antibiotics leads to the emergence of multidrug bacterial strains, which
are difficult to target using currently available medicaments. Therefore, as an alternative,
the use of NPs has been proposed as novel antibacterial agents with superior bactericidal
activity. The NPs obtained can alter the cell wall and membrane, penetrate the cytosol,
and generate reactive oxygen species (ROS), leading to further damage to enzymes, lipids,
and DNA [
75
]. The biomolecules on the NP’s surface can enhance the antibacterial activity,
while their exact role is not well understood. The activity targets both Gram-positive and
Gram-negative bacteria including multidrug-susceptible as well as multidrug-resistant
strains [
75
]. However, the antibacterial activity can vary between different tested bacterial
strains of the same species probably due to the horizontal gene transfer [
76
]. As a result,
bacteria may acquire genome islands encoding enzymes responsible for antimicrobial
resistance to NPs. The role of NPs synthesis and morphology on their activity is presented
in Table 2. The antibacterial activity was mainly tested using Ag NPs, however, other NPs
such as Au NPs or ZnO NPs are also studied for their properties. Moreover, different
microalgal species used for the synthesis result in various morphologies of the prepared
NPs which further signifies the potential of microalgae to obtain antibacterial agents.

Mar. Drugs 2023,21, 352 11 of 18
Table 2. Antibacterial activity of NPs synthesized from microalgae.
Types of NPs Microalgae Species
Used
General
Environment
Size and Morphology
of NPs
Bacteria Species
Tested Ref.
Gold NPs (AuNPs)
Arthrospira platensis marine, freshwater 5 nm, spherical S. aureus,B. subtilis [77]
Gold NPs (AuNPs)
Neodesmus pupukensis
(MG257914) freshwater 5–34 nm, circular Pseudomonas sp.,
Serratia marcescens [78]
Silver NPs
(AgNPs)
Chlorococcum humicola
(IMMTCC-17) freshwater 2–16 nm, spherical E. coli (ATCC-1105) [79]
Silver NPs
(AgNPs)
Scenedesmus sp.
(IMMTCC-25) marine, freshwater 5–10 nm,
spherical S. cutans,E. coli [80]
Silver NPs
(AgNPs) Chlorella vulgaris sp. freshwater,
terrestrial
7 nm,
spherical S. aureus,E. coli [81]
Silver NPs
(AgNPs) Chroococcus minutus freshwater crystalline E. coli,S. aureus,
P. aeruginosa,[82]
Silver NPs
(AgNPs) Oscillatoria limnetica freshwater 3.30–17.97 nm,
spherical/anisotropic E. coli,B. cereus [83]
Silver NPs
(AgNPs) Oscillatoria princeps marine, brackish,
freshwater,
3.30–17.97 nm,
spherical
S. aureus,S. pyogenes,
E. coli,[84]
Silver NPs
(AgNPs)
Anabaena sp. 66-2,
Cylindrospermopsis sp.
USC-CRB3,Synechocystis
sp. 48-3,B. braunii,
marine, brackish,
freshwater
13–25 nm,
spherical/elongated
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]
Silver NPs
(AgNPs)
Chlorella pyrenoidosa
NCIM 2738 freshwater 8 nm, irregular
K. pneumoniae,
A. hydrophila,
Acenetobacter sp.,
S. aureus
[86]
Silver NPs
(AgNPs)
Chlorella vulgaris sp.
(C. vulgaris) freshwater 1.6–34.4 nm, spherical Staphilococcus Aureus,
Klebsiella Pneumonia [23]
Silver NPs
(AgNPs)
Neodesmus pupukensis
(MG257914) freshwater 52–179 nm, spherical
Pseudomonas
aeruginosa,E. coli,
K. Pneumoniae,
S. marcescens
[78]
Silver NPs
(AgNPs) Spirogyra varians freshwater 17.6 nm, spherical
B. cereus,P. aeruginosa
and Klebsiella,S. aureus,
L. monocytogenes,E. coli
[87]
Silver NPs
(AgNPs) Coelastrella aeroterrestrica freshwater 14.5 nm,
hexagonal
Staphylococcus aureus,
Streptococcus pyogenes,
Bacillus subtilis,
Escherichia coli,
Pseudomonas aeruginosa
[53]
Silver NPs
(AgNPs) Limnothrix sp. 37-2-1 freshwater 31.86 nm,
elongated
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]
Silver NPs
(AgNPs) Anabaena sp. 66-2 brackish 24.13 nm,
irregular
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]
Silver NPs
(AgNPs) Synechocystis sp. 48-3 marine, brackish 14.64 nm,
irregular
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]
Silver NPs
(AgNPs) Botryococcus braunii freshwater 15.67 nm,
spherical
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]

Mar. Drugs 2023,21, 352 12 of 18
Table 2. Cont.
Types of NPs Microalgae Species
Used
General
Environment
Size and Morphology
of NPs
Bacteria Species
Tested Ref.
Silver NPs
(AgNPs) Coelastrum sp. 143-1 freshwater 19.28 nm,
spherical
B. megaterium,E. coli,
B. subtilis,M. luteus,P.
aeruginosa,S. aureus
[85]
Silver NPs
(AgNPs) Limnothrix sp. 37-2-1 freshwater 25.65 nm,
spherical and elongated
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]
Silver NPs
(AgNPs) Arthrospira platensis marine, freshwater 13.85 nm,
spherical
B. megaterium,E. coli,
B. subtilis,M. luteus,
P. aeruginosa,S. aureus
[85]
Zinc oxide NPs
(ZnO)
Chlorella vulgaris sp.
(C. vulgaris) freshwater
150 nm crystalline
structure/21 nm rod-like
appearance
Staphylococcus aureus,
Enterococcus faecalis,
Escherichia coli,
Pseudomonas aeruginosa
[88]
Zinc oxide NPs
(ZnO) Arthrospira platensis marine, freshwater 30.0–55.0 nm, spherical
Bacillus subtilis,
Staphylococcus aureus,
Pseudomonas
aeruginosa,
Escherichia coli
[89]
3.6. Antifungal Activity
Emerging resistance to antifungal drugs with their limited availability becomes a
growing public health concern, leading to an increase in fungal infections. Moreover,
fungal infections can rapidly spread in hospitals, especially between immunocompromised
patients [
90
]. Therefore, increasing attention is being paid to satisfy the need to develop
new and effective antifungal agents. So far, NPs have demonstrated excellent fungicidal
activity against several fungal species, as shown in Table 3. The antifungal activity was
studied for a variety of NPs, including Au NPs, Fe
3
O
4
NPs, Ag NPs, or Co(OH)
2
nanoflakes
with various size ranges and morphologies. In addition, microalgae can contain molecules
with fungicidal activity and if they also participate in the synthesis as reducing or capping
agents, it would offer synergistic antifungal action [91].
Table 3. Antifungal activity of NPs synthesized from microalgae.
Types of NPs Microalgae
Species Used
General
Environment
Size and Morphology
of NPs
Species of
Fungi Tested Ref.
Cobalt hydroxide NPs
(Co(OH)2NMs) Arthrospira platensis marine, freshwater 3.52 nm, nanoflake
C. albicans,
C. glabrata,
C. krusei.
[92]
Cobalt oxide NPs
(Co3O4NMs) Arthrospira platensis marine, freshwater 13.28 nm, nanoflake
C. albicans,
C. glabrata,
C. krusei.
[92]
Gold NPs (AuNPs) Neodesmus pupukensis
(MG257914) freshwater 5–34 nm,
circular shape
A. niger,A. fumigatus,
A. flavus,F. solani C. albicans [78]
Gold NPs (AuNPs) Chlorella sorokiniana freshwater 20–40 nm, spherical C. tropicalis,C. glabrata,
and C. albicans [93]
Gold NPs (AuNPs) Chlorella Vulgaris freshwater 2–10 nm, spherical C. albicans [94]
Iron oxide NPs
(Fe3O4NPs) Chlorella K01 freshwater 50–100 nm, spherical
Fusarium oxysporum,
Fusarium tricinctum,
Fusarium maniliforme,
Rhizoctonia solani,and
Phythium sp.
[95]

Mar. Drugs 2023,21, 352 13 of 18
Table 3. Cont.
Types of NPs Microalgae
Species Used
General
Environment
Size and Morphology
of NPs
Species of
Fungi Tested Ref.
Silver NPs (AgNPs) Arthrospira platensis marine, freshwater 9.72 nm (before
calcination)/26.01 nm (after
calcination), oval-shaped
C. albicans,
C. glabrata,
C. krusei.
[92]
Titanium dioxide NPs
(TIO2NPs) Arthrospira platensis marine, freshwater 4.81 nm (before
calcination)/4.62 nm (after
calcination), spherical-shaped
C. albicans,
C. glabrata,
C. krusei.
[92]
Zinc oxide
NPs (ZnO) Arthrospira platensis marine, freshwater ≈30.0 to 55.0 nm, spherical C. albicans [89]
3.7. Functionalization to Reduce Toxicity
Microalgae are also a source of valuable compounds which can be further stabilized on
the NPs’ surface. The study by Torrez-Diaz et al. proposed a method to stabilize Chlorella
freshwater microalgae peptide (VECYGPNRPQF) on the Au NPs surface [
96
]. The NPs
of Au were obtained through the chemical method of citrate reduction, and then peptide
solution was added to the flask. The average product size was 15 nm in diameter and
spherical in size with high stability after removing stabilizing agents and centrifuge runs.
Peptide-functionalized Au NPs showed almost three times higher antioxidant activity than
non-functionalized Au NPs, as well as decreased marine ecosystem toxicity, which could
be linked with the increased stability of functionalized Au NPs. The effect of the peptide
on the marine organisms was hypothesized as either adaptation or usage as a nutrient in
stressful environments.
In a study by Rudi et al. Ag NPs from the commercial source were functionalized by
adding to the culture of marine microalgae S. platensis and extraction from the biomass [
97
].
The product was spherical with 8–20 nm in size and administered to rats in comparison
to PEG-Ag NPs. A greater concentration of silver was observed in the liver; however, the
presence of silver in the brain tissues confirms the ability of Ag NPs to penetrate the blood–
brain barrier. The Spirulina-functionalized Ag NPs were excreted from all organs except
the brain while non-modified PEG-Ag NPs were also present in the liver. However, in a
study by El-Deeb et al., upon treatment, the level of liver enzymes ALT and AST increased,
while the concentration of urea and albumin remained normal, suggesting the interactions
between Ag NPs mainly in the liver. The complexity of interactions between NPs and
various organs should be assessed in a long-term study to understand the underlying
molecular mechanisms.
The Se NPs synthesized using different amounts of marine microalgae S. platensis
polysaccharide extract were tested to assess their cytotoxicity against several cancer cell
lines [
98
]. Increased concentration of polysaccharides decreased sizes of the prepared Se
NPs to 20–50 nm with more homogeneous size distribution than without added polysac-
charides. Moreover, Se NPs prepared from S. platensis polysaccharides were stable for at
least three months with an almost nine-fold increase in uptake by the cells. Subsequently,
enhanced uptake improved the anti-cancer activity through apoptosis induction. In addi-
tion, the material showed selectivity between cancer and normal cell lines also displaying
the potential for cancer chemoprevention.
In a separate study, Se NPs were functionalized with different amounts of phycocyanin,
pigment purified from marine microalgae Spirulina sp., against insulinoma cells [
99
]. The
phycocyanin showed a similar tendency for the size and size distribution of Se NPs as
that of polysaccharides; however, increasing the phycocyanin content increased the shell
diameter surrounding Se NPs. Thus, the phycocyanin dosage was optimized, as smaller
NPs are up-taken easily due to their larger surface area. The functionalized Se NPs showed
protective action against intracellular ROS overproduction, mitochondria fragmentation,
and activation of enzymes leading to cell apoptosis, induced by palmitic acid. The cytopro-

Mar. Drugs 2023,21, 352 14 of 18
tective activity of functionalized Se NPs show their potential against diseases related to
pancreatic islet damage.
4. Conclusions and Future Prospectives
Undoubtedly, microalgae serve as excellent candidates for the synthesis of a variety
of NPs due to their rich content of secondary metabolites acting as capping and reducing
agents. However, the process can suffer from several limitations such as low yield, the need
for optimization of conditions, or the amount of time to complete the synthesis. The exact
mechanism involving the action of metabolites is needed to describe NPs production. The
wide range of NPs synthesized from microalgae applied in the biomedical sector shows the
potential of the metabolites to influence the physicochemical properties of NPs. Further
research is required to address the issues of kinetics, cell viability, and yield with their effect
on the properties of NPs synthesized by conventional methods and by using microalgae.
As far as the future implications are concerned, it can be assumed that the alteration of
synthesis conditions might also lead to extending the knowledge of the role of metabolites
on the obtained NPs. Various species of microalgae and precursors can be tested for their
application in the biomedical field. Moreover, diverse techniques applied in the extract
preparation, targeting certain classes of metabolites might explain their role in the synthesis.
Therefore, future studies on the synthesis of NPs using microalgae can lead not only to
the optimization of the process but also to the conceptual understanding of the connection
between synthesis, properties, and activity of NPs.
Author Contributions:
Conceptualization, A.C. and G.G.; methodology, G.F.; validation, A.S., G.F.
and M.B.; formal analysis, A.S.; investigation, A.S. and G.F.; data curation, A.S., M.C. and G.F.;
writing—original draft preparation, A.S.; writing—review and editing, A.C., R.O., A.M.L. and N.L.;
supervision, A.C. and G.C.; funding acquisition, G.C. All authors have read and agreed to the
published version of the manuscript.
Funding:
This work has been developed within the framework of the project eINS- Ecosystem of
Innovation for Next Generation Sardinia (cod. ECS 00000038) funded by the Italian Ministry for
Research and Education (MUR) under the National Recovery and Resilience Plan (PNRR)—MISSION
4 COMPONENT 2, “From research to business” INVESTMENT 1.5, “Creation and strengthening of
Ecosystems of innovation” and construction of “Territorial R&D Leaders”.
Data Availability Statement: Not applicable.
Acknowledgments:
A.S. performed her activity in the framework of the International Ph.D. in
Innovation Sciences and Technologies at the University of Cagliari, Italy.
Conflicts of Interest: The authors declare no conflict of interest.
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