Biogenic synthesized AgNPs against
Candida albicans is an opportunistic dimorphic yeast. This organism is pathogen associated to superficial and systemic infections. Actually, Candida albicans represents an emergent pathogen especially in a patient with some immunity compromises. Added to this, the use of antifungal in an indiscriminate form has increased the resistance of the existing drugs. In this aspect, the nanotechnology generates the possibility of creating new therapeutic agents. Nanoparticles are structures of 1–100 nm with special physicochemical characteristics that allow it to function as therapeutic agents or as carriers of these. Palladium, silver, and gold metallic nanoparticles and iron, titanium, zinc, and copper oxides have been used as growth inhibitors. These nanoparticles have been proved alone or in form of nanocomposites. The objective of this chapter is to describe the state of the art of the use of nanoparticles as inhibitors of the growth of Candida albicans, as well as the most relevant results regarding the mechanisms involved in this inhibition.
- metallic and oxide nanoparticles
Candidiasis is one of the most common infections worldwide. These diseases are generated by
For the treatment of invasive infection by
However, the adverse effects of antifungal therapy as nephrotoxicity, red blood cell (RBC) toxicity, and arrhythmias for amphotericin B, hepatotoxicity associated with fluconazole, cardiotoxicity and gastrointestinal (GIT) disturbances attributed to itraconazole, voriconazole exhibiting neurological and hepatic toxicities, and posaconazole shown to elevate serum aminotransferase levels and cause mild-to-moderate hepatic toxicity limit its use. These adverse effects are attributed to the action mechanism. Azoles and polyenes have ergosterol as a target from the cell membrane, and the ergosterol has a high similarity between ergosterol and human cholesterol . Azoles inhibit the ergosterol biosynthesis, specifically the lanosterol 14α-demethylase. This generates sterol precursor accumulation, resulting in a cell membrane instability and the loss of viability .
The echinocandins are more specific for fungi. This therapeutic antifungal inhibits the enzyme 1,3-b-D-glucan synthase and generates the depletion of polymer synthesis and osmotic stability loss [1, 10].
Due to the discriminated antifungal agent use,
On the other hand, the survival characteristics of
Because of this, the research for more specific antifungal therapies has been the focus of attention. In this area, the nanotechnology offers an opportunity to design and create a new pharmaceutical agent that can be specific. As pharmaceutical agent nanoparticles such as copper, silver, and palladium, among others, have been employed in order to inhibit the growth of any microorganism, in particular,
The objective of this chapter is to describe the state of the art of the use of nanoparticles as inhibitors of the growth of
Fungi form one of the largest eukaryotic kingdoms; they have a very broad function of decomposing biopolymers and different compounds in dead or alive hosts, as well as synthesizing bioactive compounds. The yeast
Candida albicanscell wall
Branched polymers of glucose with β-1,3 and β-1,6 bonds, they are called β-glucans.
Unbranched polymers of N-acetyl-D-glucosamine (GlcNAc) with β-1,4 bonds; the chitin, which contributes to the fiber insolubility; and theβ-1,3 glucan-chitin complex, which is the major constituent of the inner wall. β-1,6 glucan links the components of the inner and outer walls; thus, chitin and β-1,3 are structural polysaccharides and convey strength and shape to the cell wall [15, 17, 18].
Finally mannose polymers, mannans, covalently associated with proteins (glucomannan proteins).
The mannan of the outer cell wall is less structured but allows this fungus breakage of epithelial barriers or dysfunction of the immune system converting
This arrangement is responsible for its cell morphology and provides protection to the cell against physical, chemical, and biological (immune host system) aggression .
Likewise, the cell wall participates in both growth and morphological transitions between yeast and hyphae; this conversion from the unicellular yeast to multicellular filamentous fungus is essential for
Candida albicansvirulence factors
Virulence factors include host recognition biomolecules (adhesins) due to cell surface hydrophobicity which enhanced adherence and resistance to phagocytosis; another important factor is its ability to grow either in yeast or in hyphal form (dimorphism, especially in the presence of N-acetyl-D-glucosamine) , secreting aspartyl proteases and phospholipases .
The phenomenon of the quorum sensing (QS) has been described as a contributor to morphogenesis, playing a role in regulating the yeast to filamentous fungus; one of the conditions is the dependence on cell density associated with the regulation by QS. The QS has an important role in the ability of
Candida albicansbiofilm formation
Candida albicansresistance mechanisms
In order for
Resistance to azole antifungal has been described in three forms:  change in the affinity of cytochrome P450 sterol 14α-demethylase (gen Erg11p),  increase in the enzyme expression, and  changes in the sterol desaturase. Other alterations in an enzyme involved in the cell wall construction have been described [29, 30].
The pump efflux ejects the azoles outside the cell, without any effect inside of it. This efflux is mediated by an ATP-binding cassette (ABC) and the major facilitator superfamily. This is a complex family of genes that involve several CDR genes, including core membrane pore composed by ABC segments in the membrane; two segments are in the cytosolic side, which provide energy to pump the antibiotic outside of the cell [25, 29, 30].
It is an important highlight that this resistance could be a derivative in the resistance to other antifungals.
7. Nanotechnology as an antifungal
Nanotechnology is a new technology which manipulates the matter in a nanoscale order (1 × 10−9 m). In this order, the nanoparticles have been used as antimicrobial, as antifungal, or as carried molecules. Nanoparticles have a size from 1–100 nm on any of this dimension. In this scale, nanoparticles have particularity physicochemical characteristics . Within these characteristics, their electron configuration confers an extraordinary quantum effect. These quantum effects enhance their pharmacokinetic effect and generate nanoparticles that can be used as a protein, antibodies, or inclusive specific molecule carriers .
As an antifungal, nanoparticles show cell wall damage, an oxidative stress increase, and DNA interaction . Nevertheless, the NPs toxicity mechanisms are dependent on the nanoparticle nature, size, and shape and capping nature, among other characteristics.
This chapter reviewed some NPs characteristics such as nanoparticle nature, synthesis, size, and shape and their influence on the toxic mechanisms in
7.1. Silver nanoparticles
Silver has been used for several decades as a disinfectant. Inclusively, silver nitrate was used in burns in order to control infections. However, with the discovery of penicillin, the use of silver was slow. Nowadays, the increase of resistance to antifungal puts attention again on the use of silver as an antibiotic, though the use of silver ions implies complications in their solubility and availability .
In this sense, the silver nanoparticles (AgNPs) offer a possibility. AgNPs offer the possibility of using minimal doses and act in microorganisms which present antibiotic resistance or decrease the virulence factors.
The use of different AgNPs synthesis methods caused different effects. The general method is the salt reduction by a reducing agent. However, the addition stabilizing agent has been proved. The stabilizing or capping agent limits the NPs’ growth, avoids the agglomeration, and controls the shape, size, and superficial charge .
The NP synthesis by plant extract or secondary metabolites from microorganisms offers a new possibility. The phenolic compounds, proteins, anthocyanin, and carboxylic acid, among others, work as reducing agent and stabilizing or capping agent [35, 36].
The green AgNPs synthesis through the use of microorganisms as the use of Actinobacteria
|Inhibition test||NPs size (nm)||Synthesis method||Capped||Reference|
|18 mm (inhibition zone)||15–20||NR||Moteriya et al. 2017 |
|64 μg/mL (MIC)||12.7||Proteins||Wypij et al. 2017 |
|16.7 ± 0.25 mm (inhibition zone)||2–8||NR||El-Baz et al., 2016 |
|10 μg/mL (BIC)||40–55||Detection of functional groups as phenol, alcohol, proteins, heterocyclic amines||Muthamil et al. 2018 |
|40 μg/mL (MIC)||5–95||Proteins||Lateef et al. 2015 |
|12 mm (inhibition zone with 30 μL of solution)||18–46||Detection of functional groups as alcohol, carbonyl, amino groups||Kumar et al. 2013 |
|250 μg/mL (MIC) and 500 μg/mL (MFC)||3.77–33.22||NR||Musa et al. 2018 |
|60 μg/mL (MIC90) and 120 (MFC)||2–7||Tulsi leaf extract||NR||Khatoon et al., 2015 |
|20 mm (inhibition zone with 80 μg/20 mL)||21.65–41.05||Proteins||Oves et al. 2018 |
|75 ppm reduce 83%/150 ppm (MFC)||70||Red cabbage||Anthocyanin compounds||Ocsoy et al. 2017 |
|08 μg/mL (MIC)||9–130||NR||Ashajyothi 2016 |
|40–60 μg/mL(MIC)||4–39||Citrus lemon aqueous Juice + CTAB||Surrounded by a layer||Rahisuddin et al. 2015 |
|10.8 ± 0.8 (inhibition zone)||12–85||Clove extract||Detection of functional groups such as methoxy, alcohol, carboxilic acid, aliphatic group||Parlinska-Wojtan et al. 2017 |
|21–12 cm in combination with antifungal (inhibition zone)||10–90||Flower broth of ||NR||Padalia et al. 2015 |
|12.14 μg/mL (MIC)||47.0 ± 2.0||Protein layer||Kumar et al. 2017 |
|10.78 μg/mL (MIC)||7.6 ± 0.5|
|70% fold increase (combination with antifungal, inhibition zone)||1.836–135.78||NR||Ghiut et al. 2018 |
|50% fold increase (combination with antifungal, inhibition zone)||1.012–73.83||NR|
|14 mm(inhibition zone)||5–10||Not visible||Aazam and Zaheer 2016 |
|30 μg/mL (MIC)||4–36||Actinobacterial strain SF23||NR||Anasane et al. 2016 |
|40 μg/mL (MIC)||8–60||Actinobacteria C9||NR|
Clove extract was used in order to produce AgNPs. The high content of eugenol, β-caryophyllene, humulene, chavicol, methyl salicylate, α-ylangene, and eugenone; flavonoids such as eugenin, rhamnetin, kaempferol, and eugenitin; triterpenoids like oleanolic acid, stigmasterol, and campesterol; and several sesquiterpenes allowed the clove to act as a reducing and capping agent in AgNPs synthesis. The DLS analysis showed AgNPs size of 12 and 85 nm. The AgNPs produced to generate a complete inhibition of
In order to evaluate the AgNPs effect on biofilms, some AgNPs have been evaluated. As described, biofilms are a
The use of latex plants in the AgNPs green synthesis was proved.
Also, silver nitrate chemical reduction by sodium citrate was evaluated in intermediate and mature
The results obtained show the importance of the NPs size, being the smallest nanoparticles which show the highest inhibition . Lara et al. (2015) showed that biofilm exposed to AgNPs exhibits a few
Besides the AgNPs effect in biofilm, the AgNPs have been used alone showing the inhibition, cell wall damage, and incorporation of Ag into the cell. It was used in combination with different antifungals amphotericin b [51, 56] or nystatin and chlorhexidine digluconate, showing a synergic effect . Other molecules as cationic carboxilane have been used, proving a high
The direction in the investigation with silver nanoparticles has been driven in the green synthesis due to the low toxicity generated in the human cell and the increase of the toxicity in bacteria and yeast . Their chemical characteristics as a metallic oxidative state and superficial area, among others, allow the NPs to interact with the microorganism and inhibit their growth.
7.2. Zinc oxide nanoparticles
Due to ZnO low toxicity, ZnO has been used in a medical device such as drug carriers, antibacterial agents, and bioimaging probes, among others. For this reason, the ZnO nanoparticles (ZnONPs) have been used in
A promissory ZnONPs application area is their use as antimicrobial. Their effect has been evaluated in soil, plants, bacteria, and fungi [60, 64, 65, 66] ZnONPs different synthesis methods in ZnONPs have a big influence in the
The use of
The microwave aqueous extract from
Extracts for different plants have been proved to synthesize ZnONPs in order to evaluate the toxicity of the resulting nanoparticles in
|Inhibition test||NPs size (nm)||Synthesis method||Capped||Reference|
|24 mm (inhibition zone)||30||NR||Vijayakumar et al. 2018 |
|80 μM (MIC)/>2590 μM (MFC)||8||NR||Dobrucka et al. 2018 |
|34 ± 1.28 mm (inhibition zone with 100 g/ml)||32–40||Alkane, C=O stretching, other groups||Vijayakumar et al., 2018 |
|25 μg/mL (MIC)/50 μg/mL (MFC)||18||Proteins||Elumalai and Velmurugan 2015 |
|Strong activity||36||Protein, carbohydrates, flavonoids, tannins, mannitol||Nagarajan and Kuppusamy 2013 |
|12.5 mm (inhibition zone with 150 mg/ml)||68.64||Alkene, alcohol, ether, and alkane||Padalia et al. 2018 |
|20.3 ± 0.57 mm (inhibition zone with 200 μg/mL)/25 μg/mL (MIC) 50 μg/mL (MFC)||10–30||Proteins, alkanes, aromatic group||Elumalai et al. 2015 |
|18.01 mm (inhibition zone 100 μg/mL)||80||Alcohol, aldehyde, amine||Rajabi et al. 2018 |
|24 mm (zone inhibition)||30 nm||NR||Vijayakumar et al. 2018 |
|80 μM (MIC)/>2590 μM (MFC)||8 nm||NR||Dobrucka et al. 2018 |
Dhillon et al. 2014  used chitosan as a capping and bridge to carry on citric acid, glycerol, starch, and whey powder using the nanospray drying method. The ZnONPs produced have a size range of 93.2–402.5 nm. The smallest ZnONPs were the chitosan starch NPs. Aggregation effect was observed in the largest ZnONPs; this phenomenon could be explained by the superficial charge. However, these ZnONPs have a small effect in
Collagen was used as a reducing and capping agent. The ZnONPs produced with this method showed a size between 20 and 50 nm negatively charged. ZnONPs coated by collagen showed a MIC of 99.7 ± 0.99 μg/mL, whereas the control with zinc acetate showed an MIC of 297.9 ± 2.0 μg/mL. Biofilms exposed to increase the concentration of ZnONPs showed inhibition as a result of the ZnONPs concentration .
Gelatin was used for the ZnONPs as well. ZnONPs produced with gelatin showed a diameter of 20 nm with a negative superficial charge. The ZnONPs produced were employed to biofilm exposition. ZnONPs in a concentration of 50 μg/mL showed a thickness diminution, resulting in a weak adherence biofilm compared with the biofilm treated with fluconazole .
Also, egg albumin was used as a ZnONPs template. The synthesis method produced spherical ZnONPs with a range between 10 and 20 nm. The FITR analysis showed the interaction of egg albumin and ZnONPs. In this work, ZnONPs characterization in the culture media showed importance. The use of 45 μg/ml in Sabouraud’s dextrose (SD) nutrient media does not significantly affect ZnONPs stability, size, and integrity. The synthesized ZnONPs showed
ZnONPs were evaluated both alone and as a part of nanocomposites with polystyrene. The use of ZnONPs alone showed
Nanocomposites with ZnONPs have been produced with different materials in order to increase the growth inhibition of microorganisms [45, 46]. However, the results of the antifungal activity were contradictory; ZnONPs capped by surfactant in cotton fiber have proved the inhibition of
The principal ZnONPs toxicity mechanisms reported are (1) ion liberation, (2) interaction of ZnONPs with the cell wall, and (3) stress oxidative generation . The ion liberation and the interaction of NPs with the cell trigger the generation of oxidative stress; with that the cell lost their viability .
7.3. Copper oxide nanoparticles
Copper exhibits good characteristics such as its antimicrobial activity, chemical stability, and thermal resistance. Due to their toxicity characteristic, copper has been used by Egyptians for water disinfection. The Aztecs used copper to treat sore throats, and the Persian and Indians used it to treat eye infection and venereal ulcers .
Copper has an advantage in comparison with other materials, is cheaper than any other, and is easy to oxidize to copper oxide nanoparticles (CuONPs). The CuONPs can easily make nanocomposites with polymers, macromolecules inclusive of other metals [83, 84].
The CuONPs have recovered importance in the investigation due to their low cost and the variety of applications. CuONPs have been used in the pharmaceutical, medicine, and electronic industries, among others.
The synthesis method influences the characteristics as the size, shape, and agglomeration of the CuONPs .
CuONP electrochemical production with different solvents and reaction times produces different NPs sizes from 3 to 200 nm. The CuONPs were obtained in water-acetonitrile, with sodium hydroxide as the electrolyte. The CuONPs obtained in these conditions showed a size around 20 nm. The
CuONPs prepared by precipitation method coated with acetate were evaluated together with fluconazole. CuONPs coated with acetate were more toxic than CuONPs without coating. CuONPs in combination with fluconazole showed the complete
The use of dispersant as ethylene glycol and Tween 80 proved to be efficient in the production of CuONPs capable to the
CuONPs capped with pyrimidine derivatives have shown a size of 10 nm. The CuONPs showed interaction with DNA and antioxidant activity. However, the CuONPs showed
Nanocomposites with the incorporation of Fe  Cd and Ba CuONPs , as well as the incorporation of CuONPs into cotton fiber [95, 96], polyurethane , polyester , copolymer microgel , and open polyurethane foams with starch powder , have been proved be efficient in the
As in the other nanoparticles, the green synthesis has captured the investigators’ attention. Its methodology does not produce toxic subproduct in the nanoparticle production, and it covers the nanoparticles avoiding the agglomeration [31, 84, 87].
Extracts of garlic and ginger have been used for the oxidation of copper. Garlic produces the smallest CuONPs, with a size around 14.62–22.80 nm. However, ginger shows a big quantity of very small nanoparticles.
|Inhibition test||NPs size (nm)||Synthesis method||Capped||Reference|
|15.0 c ± 0.57735 (inhibition zone with 62 μg/mL)/1.93 ± 0.76376 (MIC)||31.7||Alginate||NR||El-Batal et al. 2018 |
|10.5 (zone inhibition)||26 ± 4||NR||Sivaraj et al. 2014 |
|22.5 mm (inhibition zone)||Nanostructures|
|Ildiz et al. 2017 |
|20 mm (inhibition zone with 50 μg/mL)||NR||NR||Vishnu et al. 2016 |
|0.15 Log10 growth reduction||22 ± 1 nm||Cellulose, chitosan, and keratin nanocomposites||Cellulose, chitosan, and keratin||Tram et al., 2017 |
Cu2+ ions realized from CuONPs are the principal toxicity mechanisms reported. However, it is not the only one. Some authors reported the CuONP accumulation in the cell wall as well as the oxidative stress increase. However, more studies are necessary in order to understand the NPs toxicity mechanisms.
7.4. Other nanoparticles
Other metal nanoparticles as gold [AuNPs, ], palladium [PdNPs, ], and selenium [SeNPs, ], among others, have been proven. The microorganisms’ elimination by all these NPs has been proved to be effective.
Chemical reduction of palladium salt has produced PdNPs in an average size of 9 nm (±3.9 nm). The PdNPs showed a significative
SeNPs were prepared by the reduction of selenium chloride inside
Schiff base ligand 2-((4,6-dihethoxypyr-imidine-2-yl)methyleneenamino)-6-methoxyphenol has been used in order to capped AuNPs and platinum nanoparticles (PtNPs). Nanoparticles synthetized presented a layer. The NPs showed a granular and spherical shape with size of 38.14 ± 4.5 and 58.64 ± 3.0 nm, respectively. Both nanoparticles showed greater inhibition than amphotericin .
Biogenic PdNP production with watery extract of
7.5. Nanoparticle characteristics and toxicity and
Candida albicansinhibition growth
The most important NPs characteristics are the size, shape, composition, superficial charge, and hydrodynamic. These characteristics directly influence their capacity to interact with the molecules or cells. Ocsoy et al. (2017)  indicate that the size is strictly related to charge density. The smallest of the NPs has the biggest charge density. This explains the big attraction between small NPs and their agglomeration. The NPs agglomeration is a phenomenon that changes their capacity to work as a caring or their toxicokinetic characteristic [33, 107]. In order to improve the distribution and decrease the agglomeration, a surfactant as Pluronic F® has been added to the synthesis process (Figure 1). Other surfactants as CTAB, SDS, cation surfactants , or PVP have been proved to be efficient .
Another synthesis methodology that improves these characteristics is the use of subproducts from plants, algae, bacteria, fungi, yeast, etc. . The high content of proteins, reducing sugars or anthocyanins, works as reducing agent; additionally, some of these molecules are adhered in the NPs surface . This addition gives different characteristics to the NPs produced by chemical reduction. The NPs obtained with biogenic methods offer homogeneous NPs with antioxidant characteristics that allow the addition of other molecules and control their toxicity characteristics.
The biogenic NPs production has proven to be efficient inhibiting the
Due to the NPs size (nanometer scale), diverse toxicity mechanisms have been proposed (Figure 2). One is the direct interaction of the nanoparticle with the cell membrane. The interaction of NPs microorganism is allowed by the NPs superficial charge but also the microorganism superficial charge. This interaction generates disruption of the cell wall and the leakage of ions and the intracellular material with the microorganism’s death [117, 118, 119].
Consequent to NPs dilution, the ion interaction with the cell is another toxicity mechanism. The ions are incorporated into the cell, and they could interact with thiol groups of proteins and enzymes leading to the inhibition of crucial biological activities . Also, the NPs internalization has been reported; this generates the interaction of NPs and ions with molecules of biological importance as DNA or enzymes [33, 120]. Inside the cell, the NPs or the ions provoke Fenton’s type reaction. Due to this, the oxidative stress increases, and the proteins, lipids, and DNA release (Figure 2).
Khan et al. (2014)  write “when CoFe2O4P NCs gained a higher energy than Eg, the electrons (e−) of CoFe2O4P NCs were promoted across the band gap to the conduction band (Ec), which creates a hole (h+) in the valence band (Ec). These e− in the EC and holes in the Ev possibly have high reducing and oxidizing powers, respectively.” These electron movements generate the NPs reactivity and the oxidative stress generation. The same phenomenon is reported by Setyawati et al. (2014)  for TiONPs. These NPs adsorbed UV light, and the adsorbed UV light generates electron excitation, creating a hole in the valence band. The electrons and holes migrate to NPs superficies where they can react with oxygen or water generating oxidative stress. The superficial e− generates the liberation of hydroxyl increasing the mitochondria membrane depolarization and liberation of cytochrome c. Added to this, the results show DNA damage . However, the oxidative stress is not the only mechanisms involved in the AgNPs toxicity; also the membrane fluidity, ergosterol content, and cellular and ultrastructure morphology are altered . The squalene monooxygenase expression in
The antibiotic resistance increase in many microorganisms has encouraged researchers to focus their efforts on the synthesis and design of multiple effective compounds in order to combat the resistance mechanism of microorganisms. NPs offer a new and effective “antibiotic” that could work against the resistance mechanism in
The biogenic NPs offer a new generation of NPs that work as a carrier. These NPs offer the possibility to drive the toxicity effect to a specific target, in such way the collateral damage could be diminished.
However, the NPs toxicity mechanisms are not completely understood. The oxidative stress is one of the described mechanisms; it is possible to investigate other mechanisms involving similarity to oxidative stress. More studies are necessary to understand the influence that nanoparticles have in the
This work was partially financed by F-PROMEP-38/REV-04. SEP-23-005 and ECOS NORD 263456.
Conflict of interest
The authors declare no conflict of interest.