Chitosan-based systems aimed at local application for vaginal infections

Carbohydrate Polymers 261 (2021) 117919

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Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol

Chitosan-based systems aimed at local application for vaginal infections
Victor Hugo Sousa Araujo *, Maurício Palmeira Chaves de Souza, Gabriela Corrˆea Carvalho,
Jonatas Lobato Duarte, Marlus Chorilli **
School of Pharmaceutical Sciences, S˜
ao Paulo State University, Araraquara, SP, Brazil

A R T I C L E I N F O

A B S T R A C T

Keywords:
Chitosan
Bacterial vaginosis
Vulvovaginal candidiasis
Trichomoniasis
Vaginitis

Vaginal administration is a promising route for the local treatment of infectious vaginal diseases since it can
bypass the first-pass metabolism, drug interactions, and adverse effects. However, the commercial products
currently available for topical vulvovaginal treatment have low acceptability and do not adequately explore this
route. Mucoadhesive systems can optimize the efficacy of drugs administered by this route to increase the
retention time of the drug in the vaginal environment. Several polymers are used to develop mucoadhesive
systems, among them chitosan, a natural polymer that is highly biocompatible and technologically versatile.
Thus, the present review aimed to analyze the studies that used chitosan to develop mucoadhesive systems for

the treatment of local vaginal infections. These studies demonstrated that chitosan as a component of
mucoadhesive drug delivery systems (DDS) is a promising device for the treatment of vaginal infectious diseases,
due to the intrinsic antimicrobial activity of this biopolymer and because it does not interfere with the effec­
tiveness of the drugs used for the treatment.

1. Introduction
Alternatives to oral administration have drawn the attention of the
scientific community to the local treatment of infections. Intravaginal
administration for local effect avoids the first-pass metabolism, reduces
adverse gastrointestinal effects, and is easy to apply (De Araújo Pereira
& Bruschi, 2012; Deshpande, Rhodes, & Danish, 1992). Thus, adminis­
tration by this route is a promising alternative for the application of
contraceptive drugs and for the local treatment of infectious diseases (Da
Silva et al., 2014; De Araújo Pereira & Bruschi, 2012).
In addition to reducing adverse effects and drug interactions, a
vaginal application of therapeutic agents for the treatment of local in­
fections demonstrated, overall, comparable efficacy of oral administra­
tion
(Palmeira-de-oliveira,
Palmeira-de-oliveira,
&
Martinez-de-oliveira, 2015). However, some aspects must be consid­
ered for the development of systems for vaginal application, since this
microenvironment must be preserved, considering the pH (≈ between
4.0 and 5.0), microbiota, as well as its cyclical changes (De Araújo
Pereira & Bruschi, 2012; Srikrishna & Cardozo, 2013). Also, conven­
tional dosage forms for vaginal application are associated with
discomfort and short drug retention time, which makes it necessary to
develop mucoadhesive systems to optimize the acceptability of the

therapy and its therapeutic efficacy (dos Santos et al., 2020; Palmeir­
a-De-Oliveira et al., 2015). Mucoadhesion is a form of bioadhesion based
on mucusglycoproteins or mucous membranes (P. R. De Araújo et al.,
2019; Johal, Garg, Rath, & Goyal, 2016).
Natural polymers have been used to develop mucoadhesive vaginal
drug-delivery systems, due to their biocompatibility and ability to
remain in the vaginal mucosa, promoting the local and sustained release
of the drug (Valenta, 2005). Chitosan is one such polymer because it has
biocompatibility and antimicrobial properties, along with high adhesive
mucus power, making it a promising candidate for the development of
vaginal drug-delivery systems (Valenta, 2005). Thus, considering the
mucoadhesive and antimicrobial properties of chitosan, as well as the
advantages of vaginal application, the present study analyses the in­
fluence of this biopolymer on the development of delivery systems for
vaginal application to treat local infections.
2. General aspects of chitosan
Chitin is a structural polysaccharide present in fungi, insects, and
marine crustaceans, formed by units of (1 → 4)-2-acetamido-2-deoxyβ-D-glucan (N-acetyl D-glucosamine) (Roberts, 1992). After deacetyla­
tion of chitin, many units of acetamido become amino units; if the

* Corresponding author.
** Corresponding author.
E-mail addresses: (V.H.S. Araujo), (M. Chorilli).
/>Received 23 November 2020; Received in revised form 3 March 2021; Accepted 3 March 2021
Available online 6 March 2021
0144-8617/© 2021 Elsevier Ltd. This article is made available under the Elsevier license ( />

V.H.S. Araujo et al.

Carbohydrate Polymers 261 (2021) 117919

percentage of deacetylation is superior to 50 % of all units, this new
polymer is called chitosan. Chitosan is formed by (1 → 4)-2-acet­
amido-2-deoxy-β-D-glucan (N-acetyl D-glucosamine) and (1 →
4)-2-amino-2-deoxy-β-D-glucan (d-glucosamine) units (Fig. 1). It is a
renewable material that is extremely available in nature, and it con­
tributes to reducing environmental pollution since its main source is the
tailings from the shrimp and crab fishing industry (Campana-Filho et al.,
2007; Jung & Zhao, 2011; Robert et al., 1992; Wang, Li, & Yao, 2011).
This biopolymer has a wide application in the health sciences, as it
has properties such as biocompatibility, biodegradability, and antimi­
crobial activity, and it is. the second most available polysaccharide in
nature. (Frade et al., 2018; Nilsen-Nygaard, Strand, Vårum, Draget, &
Nordgård, 2015; Shukla, Mishra, Arotiba, & Mamba, 2013; Victorelli,
Calixto, Dos Santos Ramos, Bauab, & Chorilli, 2018; Zheng & Zhu,
2003). In other words, in addition to its diverse biological activity,
chitosan is a unique biopolymer because it is abundant, cationic,
low-toxic, non-immunogenic, and biodegradable (Felt, Buri, & Gurny,
1998; Fonseca-Santos & Chorilli, 2017).
However, before using chitosan for developing drug delivery systems
(DDS), it is essential to know some of its physicochemical properties and
how these properties impact the technological and biofunctional char­
acteristics. Much of its dispersion behavior is related to its polycationic
nature in acidic aqueous media (L. de S. Soares et al., 2019). Another
fundamental property is its degree of deacetylation (DD), which corre­
sponds to the percentage of amine groups concerning the acetamide
groups present in the chain. DD is proportional to its ability to present
low charge density: the more deacetylated the chitosan, the greater its
capacity to be protonated and the greater its ability to interact with
negatively charged surfaces (Ferreira et al., 2020). The third funda­

mental property of chitosan is its molecular mass (Mw), which indicates
the size of the polymeric chains. Low Mw is useful for the production of
systems while medium and high Mw are widely used in the production
of microsystems (Ferreira et al., 2020; Sreekumar, Goycoolea,
Moerschbacher, & Rivera-Rodriguez, 2018).
Although chitosan has been widely applied in drug dosage form, its
application depends on its dispersion (L. de S. Soares et al., 2019).
However, the chitosan dispersibility mechanism is not yet completely
clear. Research suggests that dispersibility evolves the protonation of
the groups and the chain length (De Souza et al., 2020; Ferreira et al.,
2020; L. de S. Soares et al., 2019). The DD degree can affect mostly
hydrophobicity, while molecular weight affects cytotoxicity, solubility,
and degradation (Garg, Chauhan, Nagaich, & Jain, 2019). However,
neither of these properties acts in isolation, and therefore, chitosan
performance is a conjunction of these factors (Garg et al., 2019).
Considering that chitosan is obtained by the deacetylation of chitin,
chitosan chains in the same batch do not exhibit the same degree of Mw
and DD. Deacetylation is not an accurate and complete process and it
does not occur homogeneously. The inhomogeneous deacetylation of
chitosan compromises the uniformity of structures, which in turn affects
the understanding of its impact on the properties of systems, such as
mucoadhesion, charge density, solubility, and size (Brück, Slater, &
Carney, 2010; Nwe, Furuike, & Tamura, 2010). On the other hand,
synthetic polymers have strictly known structures, with reproducibility
of synthesis and, consequently, predictable properties (Alexander, 2001;

Neuse, 2008), placing chitosan at a disadvantage in relation to synthetic
polymers. However, chitosan has other characteristics that make it a
promising polymer for the development of DDS, such as availability,
renewability, low cost, versatility, biocompatibility, and biodegrad­

ability. It is also important to highlight its versatile chemical nature,
which enables a variety of covalent and ionic modifications, making it
possible to extensively adjust the physicochemical, biological, and me­
chanical properties of chitosan-based devices (Mogocsanu, Grumezescu,
Bejenaru, & Bejenaru, 2016; Vunain, Mishra, & Mamba, 2017). Thus,
from a technological point of view, the ability to know and manipulate
the molecular characteristics of chitosan to impact its physicochemical
properties explains its wide applicability in DDS. Another important
point to be considered, besides its great technological versatility, is its
mucoadhesiveness and antimicrobial activity. Thus, chitosan is an
important device to compose DDS for the local treatment of infectious
vaginal diseases (De Lyra et al., 2007; dos Santos Ramos et al., 2019;
Mohammed, Syeda, Wasan, & Wasan, 2017).
Chitosan is able to remain adhered to mucous surfaces, thus
providing a controlled release over a long period until complete
degradation (De Souza et al., 2020). The best-accepted theory for the
adhesion between chitosan and mucin is the product of the attraction
forces resulting from hydrogen bonds, hydrophobic forces, and espe­
cially the coulombic forces that are established between the positive
charges of chitosan and the negative charges of mucin (Fig. 2), which is
negatively charged due to the presence of sialic acids and ester sulfates
(Sogias, Williams, & Khutoryanskiy, 2008).
3. Mucoadhesion and mucopenetration
The vaginal epithelium is structured in multiple layers (Hewitt,
Couse, & Korach, 2000) ordered from deepest to most superficial: the
mitotically active basal layer (“stratum basale”), the super basal layer,
and a superficial layer of flattened cornified cells, stratum corneum (SC)
(D. J. Anderson, Marathe, & Pudney, 2014). As SC is the superficial
layer, it is the principal and first point of contact with pathogens and
substances in the vagina. SC is a specialized structure that has a deposit

of glycogen and low keratinization (D. J. Anderson et al., 2014; Nilsson,
Risberg, & Heimer, 1995). It is formed by flattened, thin-layered lipid
cells, with no nuclei or organelles, slightly connected (Bragulla &
Homberger, 2009). Due to the thin lipid layer, it is possible to transfer
water to the deeper layers (epidermal epithelium) (D. J. Anderson et al.,
2014). Other substances present in the mucus composition of the SC are
mucin and syaloglycans (Bansil & Turner, 2018). The vaginal aspect
must also be considered (D. J. Anderson et al., 2014; Rampersaud,
Randis, & Ratner, 2012; Roggero, P´
erez, Bottasso, Besedovsky, & Del
Rey, 2009). The presence of the Lactobacillus sp. in the vagina can
metabolize the glycogen present in SC and turn it into lactic acid, which
reduces the pH of the SC surface (Mirmonsef et al., 2012).
The mucus can become more viscous and negatively charged in acid
conditions, which is an important factor for the adhesion of positively
charged molecules like chitosan (Bansil, Celli, Hardcastle, & Turner,
2013). Although a more viscous mucous layer can be advantageous for
adhesion, it is not interesting for the spread and penetration of the drug.
A high viscosity regime hinders the deeper drug penetration by the drug
delivery system entrapped in the mucus, and therefore, in these condi­
tions, mucopenetration can be an interesting strategy.
The mucoadhesion can also be achieved by strong interaction with
mucin. Mucin has a variable Mw range (2–20 million g/mol), and it is
one of the main components of mucus (Bansil & Turner, 2018; Bansil
et al., 2013; Sogias et al., 2008). When in a state of gelation, mucin tends
to form large aggregates due to hydrophobic interactions, hydrogen
bonding, and disulfide bonds between cysteine residues, and it can also
be negatively charged due to the presence of sialic acids and ester sul­
fates that are totally ionized at pH > 2.6. (Sogias et al., 2008).
However, electrical charge, composition, size, and shape are

important factors to be considered when developing DDS that interact

Fig. 1. Schematic representation of Chitosan.
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Carbohydrate Polymers 261 (2021) 117919

Fig. 2. General model for the interaction between chitosan and mucin.

4. Toxicity and intrinsic antimicrobial action of chitosan

with biological barriers, like the mucosa (Collado-Gonz´
alez, Gonz´
alez
Espinosa, & Goycoolea, 2019; De Souza et al., 2020; Ferreira et al.,
2020). Systems that are able to interact with the mucous layer and
remain attached to it are called mucoadhesives, and in the case of chi­
tosan, the main mechanism involves the interaction between the
opposite charge between mucus (-) and chitosan (+)(Collado-Gonz´
alez
et al., 2019).
Mucopenetration is a property that confers to the systems the ability
to permeate the mucus layer reaching the epithelium (Lai, Wang, &
Hanes, 2009). Active mucopenetration can be achieved by immobili­
zation of mucolytic enzymes on a system surface, and this strategy can
avoid the entrapment of the drug delivery in the mucus (Lai et al., 2009;
ădler, 2020). Passive mucopenetration is achieved

Taipaleenmă
aki & Sta
through the use of negatively or neutrally charged surface DDS, which
can promote penetration to the deepest mucosal layer, although in direct
contact with SC, these systems must be able to change the charge to
positive (Netsomboon & Bernkop-schnürch, 2016a, 2016b). Positively
charged surfaces have more cellular uptake than negatively charged
ăhlich, 2012).
surfaces (Fro
In this context, native and unmodified chitosan is not considered a
mucopenetrating polysaccharide, but chitosan can disrupt intercellular
junctions, increasing epithelium permeability and improving the
ădler, 2020, Vllasaliu
bioavailability of the drug (Taipaleenmă
aki & Sta
et al., 2010). However, this mechanism is not completely clear, but still,
chitosan has been proposed as a transmembrane drug delivery system
(Smith, Wood, & Dornish, 2004).
Mucopenetration and mucoadhesion have advantages and limita­
tions in drug delivery systems. An advantage of mucoadhesive DDS is
that the particles do not disrupt the mucus structure, and a limitation for
DDS intended for systemic activity is that the particles interact with
mucin and cannot reach the deeper layers. Mucopenetrative DDS, in
turn, facilitate the diffusion of the particle and allow it to reach the SC,
although it can cause temporary or permanent damage to the mucus
layer and reach the SC (Netsomboon & Bernkop-schnürch, 2016a,
2016b).
However, the systems composed of chitosan demonstrate a
mucoadhesive property, as previously mentioned, due to its cationic
character, and this property is also related to its interaction with mucin,

which is a complex phenomenon involving electrostatic interactions,
hydrogen bonds, and hydrophobic effects (Sogias et al., 2008). This
mucoadhesive property of chitosan is ideal for the local treatment of
vaginal infections, since it aims to increase the residence time of the
drug in the region, avoiding systematic action.

As previously mentioned, chitosan is widely used to develop DDS due
to its high biocompatibility, among other aspects. As pointed out by the
review of Kean and Thanou (Kean & Thanou, 2010), its biocompatibility
would vary according to the molecular modifications of this biopolymer,
making it more or less toxic. Zubareva and colleagues (Zubareva,
Shagdarova, Varlamov, Kashirina, & Svirshchevskaya, 2017) evaluated
the penetration and toxicity of chitosan and derivatives in different cell
lines (HEK-293; HaCaT; MiaPaCa-2; A431; COLO-357; RAW264. 7;
J774) in vitro. The authors observed that chitosan and its derivatives did
not cause significant cytotoxicity, except for highly quaternized de­
rivatives, which promoted interruption of the cell cycle and production
of ROS.
However, there is little information about the biocompatibility of
this biopolymer after application on mucous membranes, especially on
the vaginal mucosa, except for those present in the gastrointestinal tract
(Kean & Thanou, 2010). The lack of studies on the toxicity to vaginal
mucosa caused by polymers for local application was also pointed out by
dos Santos and collaborators (dos Santos et al., 2020), who highlighted
the need for more information related to toxicity. However, Pradines
et al. (Pradines et al., 2015) observed in ex vivo studies of porcine vaginal
mucosa that nanoparticles coated with chitosan and thiolated chitosan
did not demonstrate significant changes in histopathological analyses,
suggesting the absence of toxicity. In addition, in the study by Calvo and
collaborators (Calvo et al., 2019), films composed entirely of chitosan

demonstrated a reduction in the in vitro viability of fibroblasts up to 54
%. The authors reported that previous studies had indicated the cyto­
static potential of chitosan, which may have impacted their results
(Calvo et al., 2019; Shahabeddin et al., 1991).
In addition to its biocompatibility, the intrinsic antimicrobial activity
of chitosan is another characteristic that several studies have explored.
Chitosan is vastly applied as a natural adjuvant preservative due to its
antimicrobial action (Jennings & Bumgardner, 2016). In some studies,
chitosan is applied as a preservative agent to pharmaceutical products,
food, and beverages (Duan et al., 2019; Raafat & Sahl, 2009; Singh &
Campus, 2018)
The antibacterial effect of chitosan is more effective in gramnegative than gram-positive bacteria, with its antifungal effect being
effective against filamentous fungi and yeasts (Dutta, 2016; Kim, 2010;
Raafat & Sahl, 2009). However, the antimicrobial effect of chitosan is
dose-dependent, and in some studies it was observed that after its sep­
aration or removal from the proximity of the bacteria, some resistant
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Carbohydrate Polymers 261 (2021) 117919

bacteria could grow back, being a possible source of selection of resis­
tant bacteria (Jarry et al., 2001). Concerning the mechanism of action of
chitosan, although not clearly defined and elucidated, three hypotheses
have been considered: I) interaction with surface constituents of the cell,
II) interaction with intracellular targets, and III) antimetabolite action
(Goy, De Britto, & Assis, 2009; Raafat & Sahl, 2009).
The first mechanism is the interaction of chitosan with cellular sur­

face constituents. This ability of chitosan is considered by many re­
searchers as a membrane-disturbing compound (Chan, Mao, & Leong,
2001; Dodane, Khan, & Merwin, 1999; Fang, Chan, Mao, & Leong, 2001)
because the positively charged chitosan can disrupt the cellular mem­
brane of the bacteria (Friedman et al., 2013; Kim, 2010; Shahidi,
Arachchi, & Jeon, 1999). Polycationic chitosan can interact with the
electronegative cell surface and alter the cell permeability (Raafat &
Sahl, 2009; Shahidi et al., 1999). In a study conducted by Beck,
Yildirim-Aksoy, Shoemaker, Fuller, and Peatman (2019), a greater de­
gree of adsorption of cationic ions by chitosan at high concentrations
was observed on the cell surface. The antibacterial mechanism of chi­
tosan at high concentrations appears to be binding of the cationic-­
charged chitosan molecules onto the microbial cell surface leading to
bacteria–chitosan agglutination. At large amounts of chitosan, poly­
cationic chitosan molecules interact with the predominantly anionic cell
wall components of bacteria and link the bacteria together to form
chitosan–bacteria electrostatic complexes under acidic conditions, and
their interactions depend directly on the charge density of chitosan
(Rinaudo, 2006).
In addition, at pH > 7.0, chitosan loses its antimicrobial activity. This
can occur because deprotonation causes chitosan to lose its ability to
electrostatically bind to the negatively charged cell membrane (Atay,
2019; Sudarshan, Hoover, & Knorr, 1992). Additionally, at this pH,
chitosan is not water-soluble, and its chains interact with the cell sur­
face. Another important hypothesis is that the antibacterial activity is
related not only to the amino group and its protonation but also to other
functional groups (methyl, and OH). It plays an important role in this
complex mechanism (Kim, 2013). Chitosan at high DD is more effective
than at low DD, reinforcing the protonation hypothesis (Raafat & Sahl,
2009).

Another proposed mechanism is the binding of chitosan and DNA
(Andres, Giraud, Gerente, & Le Cloirec, 2007). Linkage to DNA leads to
the inhibition of the mRNA and protein synthesis, but before interacting
with the genetic material, chitosan needs to enter the cell. On this point,
the hypothesis is interestingly problematic, but researchers do not
believe that chitosan can penetrate the bacterial double layer and then
penetrate the nuclei of the microorganisms (Goy et al., 2009; Raafat &
Sahl, 2009). However, in another study conducted under a confocal
microscope, oligomers of chitosan were observed inside E.coli when this
bacteria was exposed to chitosan in different conditions (Fei Liu, Lin
Guan, Zhi Yang, Li, & De Yao, 2001). In this way, we believe that the Mw
of chitosan and its degree of polymerization, together with the DD de­
gree, are fundamental for this approach. Low Mw chitosan may have
greater penetrative capacity than high Mw chitosan, and therefore, the
use of chitooligosacid may be an option for this type of approach.
The third mechanism proposed is based on a known physicochemical
property of chitosan, the metal chelation. The mechanism of the com­
plex formation depends on the pH and DD degree, because in acid pH
and high DD chitosan is more efficient (Sobahi, Abdelaal, & Makki,
2014).
The chelating capacity of chitosan is important when it comes to its
antimicrobial effect (Goy et al., 2009). Chitosan can act as an antime­
tabolite, inhibiting the primary routes of bacterial nutrition, by
complexation of the metals used in bacterial metabolism, preventing its
absorption and cellular nutrition (Goy et al., 2009; Raafat & Sahl, 2009;
Sudarshan et al., 1992). On the other hand, the deposition of chitosan on
the bacterial surface may also create a physical barrier, which would
prevent the passage of nutrients, not chelated and still having an anti­
metabolite and, consequently, antimicrobial effect (Goy et al., 2009;

Raafat & Sahl, 2009; Sudarshan et al., 1992). These hypotheses are
complementary and should not occur independently. We suggest that
this is a simultaneous, complex, and cooperative mechanism resulting in
antimicrobial activity, as summarized in Fig. 3.
Palmeira-de-Oliveira (A. Palmeira-De-Oliveira et al., 2010) demon­
strated that chitosan’s antifungal activity occurs due to membrane
damage, as a result of the interaction between the protonated amino
groups and the negatively charged membrane proteins of the evaluated
candida species. This hypothesis is also discussed in the work of Albu­
querque et al. (Alburquenque et al., 2010), who observed that the
anti-candida activity of low Mw chitosan increases as the pH decreases,
which may be related to the protonation of the amino groups. Addi­
tionally, in order to assess whether the intrinsic chitosan activity varies
by its presentation, Perinelli et al. (Perinelli et al., 2018) evaluated in
vitro the activity of chitosan in suspension and nanoparticles dispersed in
HPMC gel, and observed no significant difference between the tested
groups. The hypothesis of membrane damage was also mentioned by
Tavassoli and collaborators (Tavassoli, Imani, Tajik, Moradi, & Pour­
seyed, 2012), after observing chitosan’s anti-trichomonas activity.
5. Infectious vaginal diseases
Infectious vaginal diseases are the greatest cause of demand for
medical consultations among women, with more than 70 % of adult
women having already looked for vaginal products for their treatment
(Donders, 2007; R. Palmeira-De-Oliveira et al., 2015, 2015). Although
the associated mortality rates are low, the symptoms of vaginal in­
fections have a negative impact on women’s quality of life, affecting
their sexual relationships and occupational aspects (dos Santos et al.,
2020; Karasz & Anderson, 2003; Palmeira-De-Oliveira et al., 2015). The
most common vaginal infections are caused by bacteria (bacterial vag­
inosis), fungi (vulvovaginal candidiasis), and protozoa (trichomoniasis)

(Palmeira-De-Oliveira et al., 2015).
Bacterial vaginosis (BV) is vaginitis promoted by changes in the
vaginal microenvironment, where there is an increase in the number of
anaerobes with a significant reduction in Lactobacilli, resulting in
symptoms such as vaginal malodor, increased vaginal pH, and vaginal
itching (Hay, 2017; Kenyon, Colebunders, & Crucitti, 2013). BV is the
most common vaginitis among women of childbearing age, with prev­
alence rates ranging from 20 to 60 % (Bautista et al., 2016). However,
despite the high prevalence, 50 % of BV cases are asymptomatic, leading
to treatment neglect and underreporting of cases (Hay, 2014). Despite
not having severe symptoms, BV increases the risk of other infections by
simplex virus type, Trichomonas vaginalis, Neisseria gonorrhoeae, and
Chlamydia trachomatis (Kenyon et al., 2013). Treatment options for BV
rely on oral and topical administration of antibiotic agents with a cure
rate between 80–90 % (Bradshaw et al., 2006; Coudray & Madhivanan,
2020). However, there is currently an increase in the number of recur­
rent cases of BV and species resistance to first-line treatment, making it
necessary to search for new therapeutic alternatives (Cobos, Femia, &
Vleugels, 2020; Coudray & Madhivanan, 2020).
As the second leading cause of vaginitis, Vulvovaginal candidiasis
(VVC) is also considered a global public health problem, whose inciư
dence differs among countries, from 12.1%57.3% (Gonỗalves et al.,
2016). This pathology is characterized by the infectious process caused
by the fungus of the genus Candida spp. after disorders in the vaginal
microenvironment caused by stress, hormonal variations, immunosup­
pression, diabetes, and antibiotics (Mason et al., 2012; Mtibaa et al.,
´n-Romero, Sa
´nchez-Vega, & Tay, 2003;
2017; Ruiz-S´
anchez, Caldero

Sangamithra, Verma, Sengottuvelu, & Sumathi, 2013). Among several
species of Candida spp., the most frequently isolated in CVV samples is
Candida albicans (dos Santos Ramos et al., 2016; Gonỗalves et al., 2016;
Lee, Puumala, Robbins, & Cowen, 2020; Willems, Ahmed, Liu, Xu, &
Peters, 2020). VVC causes nonspecific symptoms such as discharge,
burning, irritation, erythema, itching, pain during sex, and dryness of
the vaginal mucosa, so it can be confused with other vaginal diseases (M.
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Carbohydrate Polymers 261 (2021) 117919

Fig. 3. Mechanism of action of chitosan in bacteria, where the following processes occur: 1) electrostatic interaction between chitosan and cell membrane; 2)
alteration of membrane permeation; 3) inhibition of replication machinery; 4) DNA damage caused by DNA binding and oxidative stress; 5) metal chelation; 6) efflux
of cations. Based on the work of Chandrasekaran, Kim, & Chun, 2020.

gels, nano and microparticles, and other DDS, such as liquid crystals,
tablets, and platelets.

R. Anderson, Klink, & Cohrssen, 2004; Yano et al., 2019). Currently,
VVC treatment is performed by the oral or topical routes, with azole
agents being the most widely prescribed (Azie, Angulo, Dehn, & Sobel,
2020; Lee et al., 2020). However, in addition to a lack of treatment,
there are in the literature reports of ineffective treatments due to the
resistance of non-Candida albicans species to azoles, which can lead to
complications such as pelvic abscess, pelvic inflammatory disease,
abortion, and infertility (Gonỗalves et al., 2016; Jane, Iramiot, & Kalule,
2019; Lee et al., 2020; Marchaim, Lemanek, Bheemreddy, Kaye, &

Sobel, 2012). It is, therefore, essential to research on new therapeutic
strategies.
Trichomoniasis is a highly prevalent sexually transmitted vaginal
infection caused by the protozoan Trichomonas vaginalis (Edwards,
Burke, Smalley, & Hobbs, 2014; Mercer & Johnson, 2018). According to
WHO, there were 24,848 new cases of trichomoniasis among adults in
2008, with greater prevalence among women than men (WORLD
HEALTH ORGANIZATION, 2012). The main symptoms associated with
this infection are vulvar irritation, dysuria, and vaginal discharge. The
treatment is based on nitroimidazoles, like metronidazole and tinidazole
(Edwards et al., 2014; Rein, 2020). However, the literature has reported
an increase in drug resistance to these drugs (Dunne, Dunn, Upcroft,
O’Donoghue, & Upcroft, 2003; Kirkcaldy et al., 2012), pointing out the
importance of research on new therapies for the treatment of this
infection.
The oral therapy for the aforementioned diseases can promote pro­
nounced diverse effects, abandonment of therapy, and consequent
aggravation of the infection. It is also worth mentioning the growing
number of infections caused by etiologic agents resistant to the current
therapy (Palmeira-De-Oliveira et al., 2015). Thus, the use of topical
therapy of mucoadhesive systems is an interesting alternative, since
these systems increase the drug’s permanence time, overcoming the
limitations of conventional pharmaceutical systems (dos Santos et al.,
2020).

6.1. Gels
The definitions of gels are becoming increasingly sophisticated,
addressing the molecular (microscopic) and macroscopic points of view.
Hermans defines gels as a dispersed and coherent colloidal system of at
least two components, which in turn exhibit solid-state mechanical

properties where both the dispersion medium and the dispersed
component extend continuously throughout the system (Hermans,
1949). Gels with an aqueous dispersed phase are classified as hydrogels
(McClements, 2018).
The formation of these chitosan gels/hydrogels is explored, espe­
cially by obtaining methods in which there are necessarily attractive
interactions of formal charges (coulombic forces), hydrogen bonds, van
der Walls forces, and hydrophobic interactions, such as polyelectrolytic
complexation and ionotropic gelation, or even using chemical reactions
such as the glutaraldehyde covalent crosslinker (De Souza et al., 2020;
Ferreira et al., 2020). Although polyelectrolytic complexation and ion­
otropic gelation have been widely explored, several other methodolo­
gies are used to form these systems, making chitosan adaptable to the
reality of several research and development laboratories, which may test
chitosan as a possible drug delivery system under numerous patholog­
ical conditions.
To circumvent the side and toxic effects that conventional VVC
treatment can cause by oral administration of antifungal drugs, formu­
lations using the free drug or a delivery system dispersed in chitosanbased gels have also been studied and shown to be very promising
(Berretta et al., 2013; Campos et al., 2020; Rodero et al., 2018; Salmazi
et al., 2015). In a study, methanolic extract of Mitracarpus frigidus (Willd.
Ex Reem Schult.) was incorporated into a chitosan-based gel for the
treatment of VVC. The formulation showed pseudoplastic behavior and
became more viscous and elastic when the extract concentration was
increased (intermolecular interactions indication). The in vivo analyses
demonstrated that the formulation was better or similar to the reference
drug (clotrimazole cream 10 mg/g), reducing not only the fungal load
but also the mucosa inflammation, indicating that the developed
formulation is an interesting alternative for the treatment of VVC

6. DDS composed and coated with chitosan for the treatment of
VI
The use of chitosan to develop DDS to treat VI has been extensively
explored in the literature. Table 1 shows the developed systems, such as
5


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Carbohydrate Polymers 261 (2021) 117919

Table 1
List of studies that used chitosan for the development of DDS for the local treatment of vaginal infections.
Pathology

DDSa

Molecular Weight

Deacetylation Degree

Reference

Trichomoniasis
Trichomoniasis
Trichomoniasis
Trichomoniasis
Candidiasis
Candidiasis
Candidiasis

Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Candidiasis
Bacterial vaginosis
Bacterial vaginosis
Bacterial vaginosis
Bacterial vaginosis
Bacterial vaginosis
Bacterial vaginosis
Bacterial vaginosis

Polymeric nanoparticles
Hydrogel
Nanoparticles
Nanoparticles
Nanoparticles
Nanoparticles
Nanoemulsion

Gel
Nanoemulsion
Nanoparticles
Nanoparticles
Microcapsules
Liquid crystal system
Micro-platelets
Liquid Crystal system
Hydrogel
Tablets
Film
Nanoparticles
Hydrogel
Complexes
Gel
Nanoparticles
Nanoparticles
Nanofibers
Film
Membrane
Microspheres
Tablets

20,000 g/mol
20,000 g/mol
–
20,000 g/mol
–
LMWb
–

LMWb
–
–
LMWb
LMWb
LMWb
250,000 g/mol
LMWb
263 kDa
HMWc
230 kDa
–
< 50 kDa
LMWb
–
–
150 kDa
50− 190 kDa
310–375 kDa
185.3 kDa
50 kDa
–

–
–
–
92 %
75 %
≥ 75 %
–

–
–
–
–
–
–
85 %
–
83 %
–
80.6 %
–
90 %
–
–
75 %
97 %
75− 85%
75–85 %
83.6 %
75− 85%
77.6− 82.5%

(Malli et al., 2018)
(Malli et al., 2017)
(Elmi et al., 2020)
(Pradines et al., 2015)
(Arumugam & Rajendran, 2020)
(D. E. Araújo et al., 2020)
(de Lima et al., 2020)

(Campos et al., 2020)
(dos Santos Ramos et al., 2019)
(Amaral et al., 2019)
(Costa et al., 2019)
(Moreno et al., 2018)
(Rodero et al., 2018)
(Grisin et al., 2017)
(Salmazi et al., 2015)
(Ailincai et al., 2016)
(Fitaihi et al., 2018)
(Calvo et al., 2019)
(Arias et al., 2020)
(Perinelli et al., 2018)
(Darwesh et al., 2018)
(Berretta et al., 2013)
(Cover et al., 2012)
(Abruzzo et al., 2013)
(Zupanˇciˇc, Potrˇc, Baumgartner, Kocbek, & Kristl, 2016)
(Abilova et al., 2020)
(Tentor et al., 2020)
(Maestrelli et al., 2018)
(Paczkowska et al., 2020)

a
b
c

Drug delivery system.
Low molecular weight.
High molecular weight.

(Campos et al., 2020).
In another study, a hydrogel containing chitosan was used as a DDS
for metronidazole to treat trichomoniasis (Malli et al., 2017). The
formulation was able to control the release of metronidazole, with lower
absorption through vaginal pig mucosa in comparison with the metro­
nidazole solution, which may decrease systemic side effects. The
hydrogel presented a similar anti-T. vaginalis activity to that of the
metronidazole solution. These results show the potential of using
chitosan-based hydrogel for the treatment of trichomoniasis, combining
its features as an active pharmaceutical ingredient with its mucoadhe­
sive properties.
The results described by the mentioned studies agree with the work
carried out by Perioli and collaborators (Perioli et al., 2008) in the
previous decade. Through the development of gels for vaginal applica­
tion containing metronidazole, the authors observed that the addition of
chitosan and its derivative (5-methyl-pyrrolidinone-chitosan) to
hydroxyethylcellulose gel allowed the system to remain in the target
area without changing the vaginal pH and maintaining the microbiota.
On the other hand, the biomedical application of chitosan hydrogels
can be limited by the toxicity presented by organic crosslinkers.
Considering this aspect and looking for new alternatives, Ailincai and
collaborators (Ailincai et al., 2016) developed hydrogel containing
2-formylphenylboronic acid as a double cross-linking agent (one cova­
lent via imine formation and one physical via H bond, forming a
chemo-physical chitosan network) and an antifungal agent. The authors
obtained different degrees of crosslinking by varying the molar ratio
between amine and aldehyde, obtaining gels with elastic and rigid
characteristics, with high resistance to deformations. In addition, the
systems demonstrated anti-candida activity against C.albicans and C.

glabrata, as well as metabolic inhibition of biofilms, which the re­
searchers attributed to the already described antifungal activity of
boronic-imine compounds.

6.2. Micro and nanoparticulate systems
Chitosan can be used to obtain particles, which can be divided into
micro and nanoparticles. Microparticles are spherical particles
measuring between 1 and 1000 μm, and due to these characteristics,
these systems do not cross biological barriers, making them safe and
´llai-Szabo
´, Antal, Laki, &
pharmacokinetically predictable (Lengyel, Ka
Antal, 2019). Microparticles can be classified as microspheres composed
of a homogeneous and solid polymer matrix, and microcapsules, which
are core-shell microparticles whose core may be solid-liquid or even
have hollow spaces. Many types of biologically active molecules, indi­
vidually or simultaneously, can be inserted into a polymer matrix of
microspheres or encapsulated inside the microcapsules, and thus such
systems are considered extremely versatile (Ju & Chu, 2019). As a DDS,
microparticles have improved the efficiency of VI treatments and have
attracted researchers’ attention due to their ability to protect bioactive
drugs, proteins, and small molecules from degradation and to achieve a
controlled release rate of encapsulated drugs over hours or months,
along with easy processing and mucoadhesion (Lengyel et al., 2019;
McClements, 2018; Mishra, 2015; G. Soares et al., 2012).
Considering the properties that favor the application of micro­
systems, some studies have allied to the mucoadhesive properties of
chitosan. Maestrelli et al. (Maestrelli, Jug, Cirri, Kosalec, & Mura, 2018)
developed chitosan-alginate microspheres for the delivery of cefixime,
which demonstrated good mucoadhesive ability without interfering

with the activity of the drug. Microscale systems are also promising for
VVC treatment, as they present chitosan’s desirable characteristics like
mucoadhesiveness, low toxicity, and good biocompatibility (Costa et al.,
2019; Moreno et al., 2018). In this sense, the anti-fungal activity of
chitosan microcapsules, synthesized by electrospraying and containing
various plant extracts, was evaluated against conventionally used tablets
containing the same extracts. It was observed that in the in vitro test with
simulated vaginal fluid, the microcapsules increased the extracts’
6


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Carbohydrate Polymers 261 (2021) 117919

solubility, thus increasing their bioavailability in the vaginal environ­
ment (Moreno et al., 2018).
Nanoparticles (NPs) are one of the most widely studied DDS in recent
decades, due to advances in the areas of physics and chemistry, which
established a new area of science, "nanotechnology". NPs are defined as
particles with at least one dimension ranging between 1 and 100 nm in
diameter that can alter their physicochemical properties in comparison
with their original bulk material (Kataria et al., 2019). Research using
NPs has been carried out extensively in the most diverse fields, such as
agronomy, environmental sciences, chemistry, physics, biology, and in
the development of DDS (Pathak et al., 2019). The physicochemical
properties of a nanoparticle can change considering the biointerfaces
resulting from the interactions between NP and biological systems. This
is an unpredictability factor associated with the system and it constitutes
one of the greatest challenges for researchers aiming to develop nano­

medicines (Donahue, Acar, & Wilhelm, 2019).
Thus, considering the advantages of the local application of
mucoadhesive nanosystems, the incorporation of drugs in nano­
structured systems composed of chitosan for the treatment of BV has
been explored by some research groups. Cover and collaborators (Cover,
Lai-Yuen, Parsons, & Kumar, 2012) developed chitosan nanoparticles
for vaginal doxycycline application. The medium-sized nanoparticles of
280 nm and with 56 % ± 10 % of encapsulated drug promoted a sig­
nificant reduction in the viability of E.coli, as well as the cytotoxicity
related to the drug, indicating the potential of chitosan to increase the
biocompatibility of drugs.
Similarly, the use of chitosan nanoparticles for VVC treatment has
been shown to be a promising alternative. Ionotropic gelation synthesis
is the most broadly used method, and the murine model, mainly in fe­
male BALB/C mice, is the most suitable for in vivo evaluation of the
promising nanostructured systems for VVC treatment (Amaral et al.,
2019; D. E. Araújo et al., 2020; Arumugam & Rajendran, 2020; Costa
et al., 2019). Among these works, Arumugam and Rajendran (Arumu­
gam & Rajendran, 2020) developed chitosan nanoparticles for Callo­
phycin A delivery, and in vitro tests demonstrated that the system
optimized drug activity in all clinical isolates ((2-azole resistant; 2-azole
sensitive and 1standard strain (NCCPF). A similar result was found in in
vivo experiments, which in turn suggests the synergistic effect of chito­
san nanoparticles and the drug. Such studies corroborate the findings of
Amaral et.al. (Amaral et al., 2019), who observed that chitosan nano­
particles also increased the therapeutic efficacy of miconazole in vivo
(Amaral et al., 2019). Besides polymeric nanoparticles, the use of chi­
tosan for the coating of nanoemulsions for vaginal application was also a
matter of study, demonstrating its ability to increase the mucoadhesion
of this system and to optimize the efficacy of the incorporated drugs

(Amaral et al., 2019; D. E. Araújo et al., 2020; Arumugam & Rajendran,
2020; Costa et al., 2019; de Lima et al., 2020).
Similar to previous studies, the use of chitosan in pharmaceutical
formulations can be used to develop mucoadhesive agents to treat
trichomoniasis and as a pharmaceutically active ingredient (Elmi et al.,
2020; Malli, Bories, Bourge, Loiseau, & Bouchemal, 2018; Pradines
et al., 2015). Malli et al. showed that the possible mechanism of action of
chitosan-coated nanoparticles could be by morphological alterations,
with pits on the parasite membrane, resulting in intracellular leaking
and consequently death (Malli et al., 2018).

liquid crystals, tablets, platelets, and films.
Liquid crystals are substances that flow like a liquid but have a de­
gree of ordering between their molecules (Araujo et al., 2020; Chorilli
et al., 2009; Mezzenga et al., 2019; Victorelli, Calixto, dos Santos, Buzz´
a,
& Chorilli, 2021). Composed of amphiphilic molecules, they have a high
mucoadhesive capacity system depending on their composition, and
thus, some studies have used chitosan to increase their mucoadhesive
potential for vaginal application and, consequently, increase the effec­
tiveness against infections in this microenvironment. Curcumin is a
natural compound that has anti-candida activity. However, despite its
antimicrobial potential, its lipophilic characteristic makes it impossible
to properly explore its clinical application. Thus, considering the
anti-candida potential of curcumin and the mucoadhesion aggregated by
liquid crystals composed of chitosan, studies have used this system to
evaluate its potential against VVC (Rodero et al., 2018; Salmazi et al.,
2015). Both studies demonstrated a high mucoadhesive potential of
liquid crystalline systems, showing efficiency even greater than flu­
conazole in vivo (Rodero et al., 2018), which can be related to the in­

crease in the biomolecule’s permanence time in the vaginal
environment. On the other hand, Calvo and collaborators (Calvo et al.,
2019) suggested that the optimization of the in vitro activity of ticona­
zole chitosan films was due to the amorphous state presented by the
drug in the developed matrix.
Similarly, to increase the therapeutic efficacy of natural substances,
Paczkowska and collaborators (Paczkowska et al., 2020) developed
tablets for incorporating lyophilized extract of Chelidonium majus. In
porcine vaginal mucosa, the authors observed mucoadhesive power, and
in vitro studies demonstrated anti-S.aureus, S.epidermidis, E.faecalis, S.
pyogenes, E.coli, P.aeruginosa activity, however smaller than that of the
free extract, attributed to the release time.
Another point to be considered is the antimicrobial activity of chi­
tosan, which allows therapeutic synergism (Grisin et al., 2017). Such
synergistic activity was suggested by Grisim and collaborators (Grisin
et al., 2017), in which chitosan micro-platelets containing amphotericin
B significantly increased the drug activity, decreasing IC50 and MIC90
(4.5 and 4.8 times), which also corroborates with in vivo studies.
The combination of chitosan with other mucoadhesive agents has
also been the subject of studies for the development of DDS for vaginal
application. Abruzzo and collaborators (Abruzzo et al., 2013) evaluated
the influence of incorporating chlorhexidine digluconate on alginate and
chitosan complexes, which in turn demonstrated mucoadhesiviness with
optimization of the therapeutic effect. Similarly, in order to explore the
mucoadhesive potential of alginate and chitosan, Tentor and collabo­
rators (Tentor et al., 2020) developed a membrane for incorporating
metronidazole. In vitro studies showed that the developed system is
suitable for vaginal application, with high mucoadhesion and resistance
to vaginal fluid. In addition, the authors noted that the membrane did
not alter the drug’s effectiveness against Staphylococcus aureus and

Gardnerella vaginalis without promoting significant cytotoxic effects on
the cervix epithelial cell line. The researchers also indicated that the
combination of chitosan with other polymers increases the cyto­
compatibility of this biopolymer (Calvo et al., 2019; Shahabeddin et al.,
1991).
The mucoadhesive property of the association of chitosan and
another polymer (poly(2-ethyl-2-oxazoline)) was also evaluated in the
study of Abilova and collaborators (Abilova et al., 2020) for developing
films containing ciprofloxacin for vaginal application. By means of ex
vivo studies, the authors observed that all developed systems had
mucoadhesive potential, but mucoadhesion proved to be inversely
proportional to the increase in the proportion of poly (2-ethyl-2-­
oxazoline), suggesting that the mucoadhesiveness of these systems
depended on the presence of chitosan. The mucoadhesive potential of
chitosan was also highlighted by the study of Fitaihi and collaborators
(Fitaihi, Aleanizy, Elsamaligy, Mahmoud, & Bayomi, 2018), who
developed chitosan tablets and other mucoadhesive polymers for the
dispersion of fluconazole. The scientists demonstrated that chitosan had

6.3. Other DDS
Chitosan can interact with many polyanionic, natural, and synthetic
polymers, such as DNA, alginates, pectins, xanthan, glucosaminogly­
cans, carboxymethylcellulose and gelatin, poly(lactic-co-glycolic acid),
forming polyelectrolytic complexes. These complexes may have desir­
able properties for the development of materials (Ciro, Rojas, Alhajj,
Carabali, & Salamanca, 2020; Darwesh, Aldawsari, & Badr-Eldin, 2018).
Based on these properties, different studies have used this biopolymer to
increase the biocompatibility and mucoadhesion of other DDS, such as
7

V.H.S. Araujo et al.

Carbohydrate Polymers 261 (2021) 117919

an impact on the physical characteristics of the gel, with gels prepared
with higher concentrations of chitosan demonstrating greater
mucoadhesion.

Declaration of Competing Interest

7. General aspects and future perspectives

Acknowledgments

As observed in the reviews by Santos et al. and Palmeira-de-Oliveira
et al., the development of mucoadhesive systems for vaginal application
is a promising alternative for the treatment of infectious vaginal dis­
eases, since they can increase the residence time of the drugs in the
vaginal environment, promoting an increase in therapeutic efficacy and
greater comfort of use than the pharmaceutical products available for
this route of administration.
Few studies compared the physicochemical and biological parame­
ters of the mucoadhesive system with chitosan and the same system
without this biopolymer, which in turn compromises the clearer
assessment of the influence of chitosan on the final properties of the
developed systems. However, it was observed that the systems
composed of chitosan described in the present study had high
mucoadhesive potential in ex vivo and in vivo models. As shown in
Table 1, there is a predominance of a degree of deacetylation of ≥75 %,

indicating that such characteristic is preferable for the development of
mucoadhesive systems intended for vaginal application. It is estimated
that the use of chitosan with these characteristics corroborates previous
reports, since chitosan with a higher degree of deacetylation has greater
mucoadhesive properties (Bonferoni et al., 2006; Henriksen, Green,
Smart, Smistad, & Karlsen, 1996; Kumar, Vimal, & Kumar, 2016). The
increase in mucoadhesion is proportional to the increase in the degree of
deacetylation since this process increases the number of free and posi­
tively charged amino groups, favoring the interaction with the nega­
tively charged sialic acid of the mucous layer (Kumar et al., 2016;
´pez, & Grenha, 2012). Another important char­
Rodrigues, Dionísio, Lo
acteristic to be evaluated is the molecular weight of chitosan, for which
different studies demonstrated that high molecular weight chitosan has
greater mucoadhesive capacity due to the deeper interpenetration of the
polymer and mucus chains favored by the chain length (Khutoryanskiy,
2011; Kumar et al., 2016; Sandri et al., 2012).
In relation to the systems, a significant diversity was observed, with
emphasis on gels, micro, and nanoparticles, which reinforces the
versatility of this biopolymer. It was also observed that a significant
number of studies used other mucoadhesive polymers in order to opti­
mize this characteristic of the systems, which can offer an important
alternative to optimize the mucoadhesion promoted by chitosan.
As for its biological influence, few studies have reported references
and assessments of the anti-microbial and anti-inflammatory activity
isolated from chitosan, which in turn can offer a synergistic potential to
the therapy under study. In addition, despite its biocompatibility being
widely described, few studies have evaluated the biocompatibility of the
systems in vivo and in vitro, which may hinder their future commer­
cialization. Regarding the drug under study, a greater number of works

used these systems to deliver drugs already marketed to increase their
effectiveness and reduce the toxicity associated with them. The capacity
of these systems to disperse molecules of natural origin has also been
observed, making it possible to explore their potential for the treatment
of VI.
In general, by analyzing the studies described in the present work,
we observed that chitosan is a promising adjuvant to be used for the
development of mucoadhesive systems for vaginal application and local
treatment of VI. It is worth noting that chitosan can be associated with
other mucoadhesive polymers, making it possible to explore the
mucoadhesiveness of both. However, there is a need for studies in search
of a better understanding of the interaction of chitosan with etiologic
agents and studies that demonstrate the biocompatibility of the devel­
oped system, making them eligible for commercialization.

˜o de Aperfeicoamento de
This work was supported by the Coordenaca
Pessoal de Nível Superior – Brasil (CAPES) [Finance Code001] and
˜o de Amparo a
` Pesquisa do Estado de S˜
Fundaca
ao Paulo – Brasil
(FAPESP), numbers 2019/10261-2 and 2019/25125-7.

The authors report no declarations of interest.

References
Abilova, G. K., Kaldybekov, D. B., Irmukhametova, G. S., Kazybayeva, D. S.,
Iskakbayeva, Z. A., Kudaibergenov, S. E., et al. (2020). Chitosan/poly(2-ethyl-2oxazoline) films with ciprofloxacin for application in vaginal drug delivery.
Materials, 13(7). />Abruzzo, A., Bigucci, F., Cerchiara, T., Saladini, B., Gallucci, M. C., Cruciani, F., …

Luppi, B. (2013). Chitosan/alginate complexes for vaginal delivery of chlorhexidine
digluconate. Carbohydrate Polymers, 91(2), 651–658. />carbpol.2012.08.074
Ailincai, D., Marin, L., Morariu, S., Mares, M., Bostanaru, A. C., Pinteala, M., …
Barboiu, M. (2016). Dual crosslinked iminoboronate-chitosan hydrogels with strong
antifungal activity against Candida planktonic yeasts and biofilms. Carbohydrate
Polymers, 152, 306–316. />Alburquenque, C., Bucarey, S. A., Neira-Carrillo, A., Urza, B., Hermosilla, G., &
Tapia, C. V. (2010). Antifungal activity of low molecular weight chitosan against
clinical isolates of Candida spp. Medical Mycology, 48(8), 1018–1023. https://doi.
org/10.3109/13693786.2010.486412
Alexander, C. (2001). Synthetic polymer systems in drug delivery. Expert Opinion on
Emerging Drugs, 6(2), 345–363.
Amaral, A. C., Saavedra, P. H. V., Oliveira Souza, A. C., de Melo, M. T., Tedesco, A. C.,
Morais, P. C., … Bocca, A. L. (2019). Miconazole loaded chitosan-based
nanoparticles for local treatment of vulvovaginal candidiasis fungal infections.
Colloids and Surfaces B, Biointerfaces, 174(October 2018), 409–415. />10.1016/j.colsurfb.2018.11.048
Anderson, D. J., Marathe, J., & Pudney, J. (2014). The structure of the human vaginal
stratum corneum and its role in immune defense. American Journal of Reproductive
Immunology, 71(6), 618–623.
Anderson, M. R., Klink, K., & Cohrssen, A. (2004). Evaluation of vaginal complaints.
Jama, 291(11), 1368–1379.
Andres, Y., Giraud, L., Gerente, C., & Le Cloirec, P. (2007). Antibacterial effects of
chitosan powder: Mechanisms of action. Environmental Technology, 28(12),
1357–1363.
Araújo, D. E., de Oliveira, A. A., dos Santos Cabral, M., Costa, A. F., Silva, B. C., do Carmo
Silva, L., … Pereira, M. (2020). Investigation of thiosemicarbazide free or within
chitosan nanoparticles in a murine model of vulvovaginal candidiasis. Brazilian
Journal of Microbiology, 1–9.
Araújo, P. R. D., Maria, G., Calixto, F., Cristiane, I., Henrique, L., Zago, D. P., …
Chorilli, M. (2019). Mucoadhesive in situ gelling liquid crystalline precursor system
to improve the vaginal administration of drugs. AAPS PharmSciTech, 20(225), 1–12.

/>Araujo, V. H. S., Duarte, J. L., Carvalho, G. C., Silvestre, A. L. P., Fonseca-Santos, B.,
Marena, G. D., … Chorilli, M. (2020). Nanosystems against candidiasis: a review of
studies performed over the last two decades. Critical Reviews in Microbiology, 0(0),
1–40. />Arias, L. S., Pessan, J. P., de Souza Neto, F. N., Lima, B. H. R., de Camargo, E. R.,
Ramage, G., … Monteiro, D. R. (2020). Novel nanocarrier of miconazole based on
chitosan-coated iron oxide nanoparticles as a nanotherapy to fight Candida biofilms.
Colloids and Surfaces B, Biointerfaces, 192(February), 111080. />10.1016/j.colsurfb.2020.111080
Arumugam, G., & Rajendran, R. (2020). Callophycin A loaded chitosan and spicules
based nanocomposites as an alternative strategy to overcome vaginal candidiasis.
International Journal of Biological Macromolecules.
Atay, H. Y. (2019). Antibacterial activity of chitosan-based systems. Functional chitosan
(pp. 457–489). Springer.
Azie, N., Angulo, D., Dehn, B., & Sobel, J. D. (2020). Oral Ibrexafungerp: An
investigational agent for the treatment of vulvovaginal candidiasis. Expert Opinion on
Investigational Drugs, 1–8. />Bansil, R., & Turner, B. S. (2018). The biology of mucus: Composition, synthesis and
organization. Advanced Drug Delivery Reviews, 124, 3–15.
Bansil, R., Celli, J. P., Hardcastle, J. M., & Turner, B. S. (2013). The influence of mucus
microstructure and rheology in Helicobacter pylori infection. Frontiers in
Immunology, 4(OCT), 310. />Bautista, C. T., Wurapa, E., Sateren, W. B., Morris, S., Hollingsworth, B., & Sanchez, J. L.
(2016). Bacterial vaginosis: A synthesis of the literature on etiology, prevalence, risk
factors, and relationship with chlamydia and gonorrhea infections. Military Medical
Research. />Beck, B. H., Yildirim-Aksoy, M., Shoemaker, C. A., Fuller, S. A., & Peatman, E. (2019).
Antimicrobial activity of the biopolymer chitosan against Streptococcus iniae.
Journal of Fish Diseases, 42(3), 371–377.

8


V.H.S. Araujo et al.

Carbohydrate Polymers 261 (2021) 117919

Berretta, A. A., de Castro, P. A., Cavalheiro, A. H., Fortes, V. S., Bom, V. P.,
Nascimento, A. P., … Goldman, G. H. (2013). Evaluation of mucoadhesive gels with
propolis (EPP-AF) in preclinical treatment of candidiasis vulvovaginal infection.
Evidence-based Complementary and Alternative Medicine, 2013.
Bonferoni, M. C., Giunchedi, P., Scalia, S., Rossi, S., Sandri, G., & Caramella, C. (2006).
Chitosan gels for the vaginal delivery of lactic acid: Relevance of formulation
parameters to mucoadhesion and release mechanisms. AAPS PharmSciTech. https://
doi.org/10.1208/pt0704104
Bradshaw, C. S., Morton, A. N., Hocking, J., Garland, S. M., Morris, M. B., Moss, L. M., …
Fairley, C. K. (2006). High recurrence rates of bacterial vaginosis over the course of
12 months after oral metronidazole therapy and factors associated with recurrence.
The Journal of Infectious Diseases. />Bragulla, H. H., & Homberger, D. G. (2009). Structure and functions of keratin proteins in
simple, stratified, keratinized and cornified epithelia. Journal of Anatomy, 214(4),
516–559.
Brück, W. M., Slater, J. W., & Carney, B. F. (2010). Chitin and chitosan from marine
organisms. Chitin, chitosan, oligosaccharides and their derivatives: Biological activities
and applications (pp. 11–19). Boca Raton: Taylor & Francis.
Calvo, N. L., Svetaz, L. A., Alvarez, V. A., Quiroga, A. D., Lamas, M. C., & Leonardi, D.
(2019). Chitosan-hydroxypropyl methylcellulose tioconazole films: A promising
alternative dosage form for the treatment of vaginal candidiasis. International Journal
of Pharmaceutics, 556. Elsevier B.V.
Campana-Filho, S. P., De Britto, D., Curti, E., Abreu, F. R., Cardoso, M. B., Battisti, M. V.,
Lavall, R. L. (2007). Extraỗ
ao, estrutura e propriedades de α e β-quitina. Quimica
Nova, 30(3), 644–650.
Campos, L. M., de Oliveira Lemos, A. S., da Cruz, L. F., de Freitas Araújo, M. G., de Mello
Botti, G. C. R., Júnior, J. L. R., et al. (2020). Development and in vivo evaluation of
chitosan-gel containing Mitracarpus frigidus methanolic extract for vulvovaginal

candidiasis treatment. Biomedecine & Pharmacotherapy, 130, Article 110609.
Chan, V., Mao, H.-Q., & Leong, K. W. (2001). Chitosan-induced perturbation of
dipalmitoyl-sn-glycero-3-phosphocholine membrane bilayer. Langmuir, 17(12),
3749–3756.
Chandrasekaran, M., Kim, K. D., & Chun, S. C. (2020). Antibacterial activity of chitosan
nanoparticles: A review. Processes, 8(9), 1–21. />Chorilli, M., Prestes, P. S., Rigon, R. B., Leonardi, G. R., Chiavacci, L. A., & Scarpa, M. V.
(2009). Development of liquid-crystalline systems using silicon glycol copolymer
and polyether functional siloxane. Química Nova, 32(4), 1036–1040. https://doi.
org/10.1590/s0100-40422009000400035
Ciro, Y., Rojas, J., Alhajj, M. J., Carabali, G. A., & Salamanca, C. H. (2020). Production
and characterization of chitosan–Polyanion nanoparticles by polyelectrolyte
complexation assisted by high-intensity sonication for the modified release of
methotrexate. Pharmaceuticals, 13(1), 11.
Cobos, G. A., Femia, A., & Vleugels, R. A. (2020). Bacterial vaginosis: An update on
diagnosis and treatment. American Journal of Clinical Dermatology, 21(3), 339–353.
/>Collado-Gonz´
alez, M., Gonz´
alez Espinosa, Y., & Goycoolea, F. M. (2019). Interaction
between chitosan and mucin: Fundamentals and applications. Biomimetics, 4(2), 32.
Costa, A. F., Araujo, D. E., Cabral, M. S., Brito, I. T., De Menezes Leite, L. B., Pereira, M.,
et al. (2019). Development, characterization, and in vitro-in vivo evaluation of
polymeric nanoparticles containing miconazole and farnesol for treatment of
vulvovaginal candidiasis. Medical Mycology, 57(1), 52–62. />mmy/myx155
Coudray, M. S., & Madhivanan, P. (2020). Bacterial vaginosis—A brief synopsis of the
literature. European Journal of Obstetrics, Gynecology, and Reproductive Biology, 245,
143–148. />Cover, N. F., Lai-Yuen, S., Parsons, A. K., & Kumar, A. (2012). Synergetic effects of
doxycycline-loaded chitosan nanoparticles for improving drug delivery and efficacy.
International Journal of Nanomedicine, 7, 2411–2419. />S27328
Da Silva, P. B., Ramos, M. A. D. S., Bonif´
acio, B. V., Negri, K. M. S., Sato, M. R.,

Bauab, T. M., et al. (2014). Nanotechnological strategies for vaginal administration
of drugs - A review. Journal of Biomedical Nanotechnology, 10(9), 2218–2243.
/>Darwesh, B., Aldawsari, H. M., & Badr-Eldin, S. M. (2018). Optimized chitosan/anion
polyelectrolyte complex based inserts for vaginal delivery of fluconazole: In vitro/in
vivo evaluation. Pharmaceutics, 10(4). />pharmaceutics10040227
De Araújo Pereira, R. R., & Bruschi, M. L. (2012). Vaginal mucoadhesive drug delivery
systems. Drug Development and Industrial Pharmacy, 38(6), 643–652. />10.3109/03639045.2011.623355
de Lima, L., AS Ramos, M., Toledo, L. G., de Rodero, C. F., Hil´
ario, F., dos Santos, L. C., …
Bauab, T. M. (2020). Syngonanthus nitens (Bong.) ruhland derivatives loaded into a
lipid nanoemulsion for enhanced antifungal activity against Candida parapsilosis.
Current Pharmaceutical Design, 26(14), 1556–1565.
De Lyra, M. A. M., Soares-Sobrinho, J. L., Brasileiro, M. T., De La Roca, M. F.,
Barraza, J. A., Viana, O. D. S., et al. (2007). Sistemas matriciais hidrofớlicos e
mucoadesivos para liberaỗ
ao controlada de f
armacos. Latin American Journal of
Pharmacy.
De Souza, M. P. C., S´
abio, R. M., Ribeiro, T. D. C., Dos Santos, A. M., Meneguin, A. B., &
Chorilli, M. (2020). Highlighting the impact of chitosan on the development of
gastroretentive drug delivery systems. International Journal of Biological
Macromolecules, 159, 804–822. />Deshpande, A. A., Rhodes, C. T., & Danish, M. (1992). INTRAVAGINAL DRUG DELIVERY,
18, 1225–1279.
Dodane, V., Khan, M. A., & Merwin, J. R. (1999). Effect of chitosan on epithelial
permeability and structure. International Journal of Pharmaceutics, 182(1), 21–32.

Donahue, N. D., Acar, H., & Wilhelm, S. (2019). Concepts of nanoparticle cellular uptake,
intracellular trafficking, and kinetics in nanomedicine. Advanced Drug Delivery
Reviews, 143, 68–96.

Donders, G. G. G. (2007). Definition and classification of abnormal vaginal flora. Best
Practice & Research Clinical Obstetrics & Gynaecology. />bpobgyn.2007.01.002
dos Santos, A. M., Carvalho, S. G., Araujo, V. H. S., Carvalho, G. C., Gremi˜
ao, M. P. D., &
Chorilli, M. (2020). Recent advances in hydrogels as strategy for drug delivery
intended to vaginal infections. International Journal of Pharmaceutics. https://doi.
org/10.1016/j.ijpharm.2020.119867
dos Santos Ramos, M. A., Da Silva, P. B., De Toledo, L. G., Oda, F. B., Da Silva, I. C., Dos
Santos, L. C., et al. (2019). Intravaginal delivery of Syngonanthus nitens (Bong.)
Ruhland fraction based on a nanoemulsion system applied to vulvovaginal
candidiasis treatment. Journal of Biomedical Nanotechnology, 15(5), 1072–1089.
dos Santos Ramos, M. A., de Toledo, L. G., Fioramonti Calixto, G. M., Bonif´
acio, B. V., de
Freitas Araújo, M. G., dos Santos, L. C., … Bauab, T. M. (2016). Syngonanthus nitens
Bong. (Rhul.)-loaded nanostructured system for Vulvovaginal candidiasis treatment.
International Journal of Molecular Sciences. />Duan, C., Meng, X., Meng, J., Khan, M. I. H., Dai, L., Khan, A., … Ni, Y. (2019). Chitosan
as a preservative for fruits and vegetables: A review on chemistry and antimicrobial
properties. Journal of Bioresources and Bioproducts, 4(1), 11–21.
Dunne, R. L., Dunn, L. A., Upcroft, P., O’Donoghue, P. J., & Upcroft, J. A. (2003). Drug
resistance in the sexually transmitted protozoan Trichomonas vaginalis. Cell
Research, 13(4), 239–249. />Dutta, P. K. (2016). Chitin and chitosan for regenerative medicine. Springer.
Edwards, T., Burke, P., Smalley, H., & Hobbs, G. (2014). Trichomonas vaginalis : Clinical
relevance, pathogenicity and diagnosis. Critical Reviews in Microbiology, 1–12.
/>Elmi, T., Rahimi Esboei, B., Sadeghi, F., Zamani, Z., Didehdar, M., Fakhar, M., …
Tabatabaie, F. (2020). In vitro antiprotozoal effects of nano-chitosan on plasmodium
falciparum, Giardia lamblia and trichomonas vaginalis. Acta Parasitologica. , Article
0123456789. />Fang, N., Chan, V., Mao, H.-Q., & Leong, K. W. (2001). Interactions of phospholipid
bilayer with chitosan: Effect of molecular weight and pH. Biomacromolecules, 2(4),
1161–1168.
Fei Liu, X., Lin Guan, Y., Zhi Yang, D., Li, Z., & De Yao, K. (2001). Antibacterial action of

chitosan and carboxymethylated chitosan. Journal of Applied Polymer Science, 79(7),
1324–1335.
Felt, O., Buri, P., & Gurny, R. (1998). Chitosan: A unique polysaccharide for drug
delivery. Drug Development and Industrial Pharmacy, 24(11), 979–993.
Ferreira, L. M. B., Dos Santos, A. M., Boni, F. I., Dos Santos, K. C., Robusti, L. M. G., de
Souza, M. P. C., et al. (2020). Design of chitosan-based particle systems: A review of
the physicochemical foundations for tailored properties. Carbohydrate Polymers,
Article 116968.
Fitaihi, R. A., Aleanizy, F. S., Elsamaligy, S., Mahmoud, H. A., & Bayomi, M. A. (2018).
Role of chitosan on controlling the characteristics and antifungal activity of
bioadhesive fluconazole vaginal tablets. Journal of the Saudi Pharmaceutical Society,
26(2), 151–161. />Fonseca-Santos, B., & Chorilli, M. (2017). An overview of carboxymethyl derivatives of
chitosan: Their use as biomaterials and drug delivery systems. Materials Science and
Engineering C. />Frade, M. L., de Annunzio, S. R., Calixto, G. M. F., Victorelli, F. D., Chorilli, M., &
Fontana, C. R. (2018). Assessment of chitosan-based hydrogel and photodynamic
inactivation against propionibacterium acnes. Molecules. />molecules23020473
Friedman, A. J., Phan, J., Schairer, D. O., Champer, J., Qin, M., Pirouz, A., … Kim, J.
(2013). Antimicrobial and anti-inflammatory activity of chitosan – Alginate
nanoparticles : A targeted therapy for cutaneous pathogens. The Journal of
Investigative Dermatology, 133(5), 12311239. />Fră
ohlich, E. (2012). The role of surface charge in cellular uptake and cytotoxicity of
medical nanoparticles. International Journal of Nanomedicine, 7, 5577.
Garg, U., Chauhan, S., Nagaich, U., & Jain, N. (2019). Current advances in chitosan
nanoparticles based drug delivery and targeting. Advanced Pharmaceutical Bulletin, 9
(2), 195.
Gonỗalves, B., Ferreira, C., Alves, C. T., Henriques, M., Azeredo, J., & Silva, S. (2016).
Vulvovaginal candidiasis: Epidemiology, microbiology and risk factors. Critical
Reviews in Microbiology, 42(6), 905–927. />1040841X.2015.1091805
Goy, R. C., De Britto, D., & Assis, O. B. G. (2009). A review of the antimicrobial activity of
chitosan. Polimeros, 19(3), 241–247. />Grisin, T., Bories, C., Bombardi, M., Loiseau, P. M., Rouffiac, V., Solgadi, A., …

Bouchemal, K. (2017). Supramolecular chitosan micro-platelets synergistically
enhance anti-Candida albicans activity of amphotericin B using an
immunocompetent murine model. Pharmaceutical Research, 34(5), 1067–1082.
Hay, P. (2014). Bacterial vaginosis. Medicine (United Kingdom). />j.mpmed.2014.04.011
Hay, P. (2017). Bacterial vaginosis. F1000Research, 6, 2–6. />f1000research.11417.1
Henriksen, I., Green, K. L., Smart, J. D., Smistad, G., & Karlsen, J. (1996). Bioadhesion of
hydrated chitosans: An in vitro and in vivo study. International Journal of
Pharmaceutics. />´ HR Kruyt.
Hermans, P. H. (1949). Colloid science (pp. 483–494). New York: Ed.
Hewitt, S. C., Couse, J. F., & Korach, K. S. (2000). Estrogen receptor transcription and
transactivation Estrogen receptor knockout mice: What their phenotypes reveal
about mechanisms of estrogen action. Breast Cancer Research, 2(5), 1–8.

9


V.H.S. Araujo et al.

Carbohydrate Polymers 261 (2021) 117919

Jane, W. J., Iramiot, J. S., & Kalule, J. B. (2019). Prevalence and antifungal susceptibility
patterns of Candida Isolated on CHROMagarTMCandida at a tertiary referral
hospital, Eastern Uganda. Microbiology Research Journal International, 1–6.
Jarry, C., Chaput, C., Chenite, A., Renaud, M.–A., Buschmann, M., & Leroux, J.–C.
(2001). Effects of steam sterilization on thermogelling chitosan-based gels. Journal of
Biomedical Materials Research: An Official Journal of The Society for Biomaterials, The
Japanese Society for Biomaterials, and The Australian Society for Biomaterials and the
Korean Society for Biomaterials, 58(1), 127–135.
Jennings, J. A., & Bumgardner, J. D. (2016). Chitosan based biomaterials volume 1:
Fundamentals. Woodhead Publishing.

Johal, H. S., Garg, T., Rath, G., & Goyal, A. K. (2016). Advanced topical drug delivery
system for the management of vaginal candidiasis. Drug Delivery. />10.3109/10717544.2014.928760
Ju, X.-J., & Chu, L.-Y. (2019). Lab-on-a-chip fabrication of polymeric microparticles for
drug encapsulation and controlled release. Microfluidics for pharmaceutical
applications (pp. 217–280). Elsevier.
Jung, J., & Zhao, Y. (2011). Characteristics of deacetylation and depolymerization of $β
$-chitin from jumbo squid (Dosidicus gigas) pens. Carbohydrate Research, 346(13),
1876–1884. />Karasz, A., & Anderson, M. (2003). The vaginitis monologues: Women’s experiences of
vaginal complaints in a primary care setting. Social Science & Medicine. https://doi.
org/10.1016/S0277-9536(02)00092-8
ˇ c´
Kataria, S., Jain, M., Rastogi, A., Zivˇ
ak, M., Brestic, M., Liu, S., et al. (2019). Role of
nanoparticles on photosynthesis: Avenues and applications. Nanomaterials in plants,
algae and microorganisms (pp. 103–127). Elsevier.
Kean, T., & Thanou, M. (2010). Biodegradation, biodistribution and toxicity of chitosan.
Advanced Drug Delivery Reviews, 62(1), 3–11. />addr.2009.09.004
Kenyon, C., Colebunders, R., & Crucitti, T. (2013). The global epidemiology of bacterial
vaginosis: A systematic review. American Journal of Obstetrics and Gynecology, 209
(6), 505–523. />Khutoryanskiy, V. V. (2011). Advances in mucoadhesion and mucoadhesive polymers.
Macromolecular Bioscience. />Kim, S.-K. (2010). Chitin, chitosan, oligosaccharides and their derivatives: Biological activities
and applications. CRC Press.
Kim, S.-K. (2013). Chitin and chitosan derivatives: Advances in drug discovery and
developments. CRC press.
Kirkcaldy, R. D., Augostini, P., Asbel, L. E., Bernstein, K. T., Kerani, R. P.,
Mettenbrink, C. J., … Weinstock, H. S. (2012). Trichomonas vaginalis antimicrobial
drug resistance in 6 US cities, STD surveillance network, 2009–2010. Emerging
Infectious Diseases, 18(6), 939–943. />Kumar, A., Vimal, A., & Kumar, A. (2016). Why Chitosan? From properties to perspective
of mucosal drug delivery. International Journal of Biological Macromolecules, 91,
615–622. />Lai, S. K., Wang, Y.-Y., & Hanes, J. (2009). Mucus-penetrating nanoparticles for drug and

gene delivery to mucosal tissues. Advanced Drug Delivery Reviews, 61(2), 158–171.
/>Lee, Y., Puumala, E., Robbins, N., & Cowen, L. E. (2020). Antifungal drug resistance:
Molecular mechanisms in Candida albicans and beyond. Chemical Reviews. https://
doi.org/10.1021/acs.chemrev.0c00199. acs.chemrev.0c00199.
Lengyel, M., K´
allai-Szab´
o, N., Antal, V., Laki, A. J., & Antal, I. (2019). Microparticles,
microspheres, and microcapsules for advanced drug delivery. Scientia Pharmaceutica,
87(3), 20.
Maestrelli, F., Jug, M., Cirri, M., Kosalec, I., & Mura, P. (2018). Characterization and
microbiological evaluation of chitosan-alginate microspheres for ce fi xime vaginal
administration. Carbohydrate Polymers, 192(November 2017), 176–183. https://doi.
org/10.1016/j.carbpol.2018.03.054
Malli, S., Bories, C., Bourge, M., Loiseau, P. M., & Bouchemal, K. (2018). Surfacedependent endocytosis of poly(isobutylcyanoacrylate) nanoparticles by Trichomonas
vaginalis. International Journal of Pharmaceutics, 548(1), 276–287. />10.1016/j.ijpharm.2018.07.006
Malli, S., Bories, C., Pradines, B., Loiseau, P. M., Ponchel, G., & Bouchemal, K. (2017). In
situ forming pluronic® F127/chitosan hydrogel limits metronidazole transmucosal
absorption. European Journal of Pharmaceutics and Biopharmaceutics, 112, 143–147.
/>Marchaim, D., Lemanek, L., Bheemreddy, S., Kaye, K. S., & Sobel, J. D. (2012).
Fluconazole-resistant Candida albicans vulvovaginitis. Obstetrics and Gynecology, 120
(6), 1407–1414.
Mason, K. L., Downward, J. R. E., Mason, K. D., Falkowski, N. R., Eaton, K. A., Kao, J. Y.,
… Huffnagle, G. B. (2012). Candida albicans and bacterial microbiota interactions in
the cecum during recolonization following broad-spectrum antibiotic therapy.
Infection and Immunity, 80(10), 3371–3380.
McClements, D. J. (2018). Advances in nanoparticle and microparticle delivery systems
for increasing the dispersibility, stability, and bioactivity of phytochemicals.
Biotechnology Advances, (July) />0–1.
Mercer, F., & Johnson, P. J. (2018). Trichomonas vaginalis: Pathogenesis, symbiont
interactions, and host cell immune responses. Trends in Parasitology, 34(8), 683–693.

/>Mezzenga, R., Seddon, J. M., Drummond, C. J., Boyd, B. J., Schră
oder-Turk, G. E., &
Sagalowicz, L. (2019). Nature-inspired design and application of lipidic lyotropic
liquid crystals. Advanced Materials. />Mirmonsef, P., Gilbert, D., Veazey, R. S., Wang, J., Kendrick, S. R., & Spear, G. T. (2012).
A comparison of lower genital tract glycogen and lactic acid levels in women and
macaques: Implications for HIV and SIV susceptibility. AIDS Research and Human
Retroviruses, 28(1), 76–81.

Mishra, M. (2015). Handbook of encapsulation and controlled release. CRC press.
Mogocsanu, G. D., Grumezescu, A. M., Bejenaru, L. E., & Bejenaru, C. (2016). Natural and
synthetic polymers for drug delivery and targeting. Nanobiomaterials in drug delivery
(pp. 229–284). Elsevier.
Mohammed, M. A., Syeda, J. T. M., Wasan, K. M., & Wasan, E. K. (2017). An overview of
chitosan nanoparticles and its application in non-parenteral drug delivery.
Pharmaceutics, 9(4). />Moreno, M. A., G´
omez-Mascaraque, L. G., Arias, M., Zampini, I. C., Sayago, J. E.,
Ramos, L. L. P., … Isla, M. I. (2018). Electrosprayed chitosan microcapsules as
delivery vehicles for vaginal phytoformulations. Carbohydrate Polymers, 201,
425–437.
Mtibaa, L., Fakhfakh, N., Kallel, A., Belhadj, S., Belhaj Salah, N., Bada, N., et al. (2017).
Vulvovaginal candidiasis: Etiology, symptomatology and risk factors. Journal de
Mycologie Medicale, 27(2), 153–158. />mycmed.2017.01.003
Netsomboon, K., & Bernkop-schnürch, A. (2016a). European Journal of Pharmaceutics
and Biopharmaceutics Mucoadhesive vs. Mucopenetrating particulate drug delivery.
European Journal of Pharmaceutics and Biopharmaceutics, 98, 76–89. />10.1016/j.ejpb.2015.11.003
Netsomboon, K., & Bernkop-Schnürch, A. (2016b). Mucoadhesive vs. Mucopenetrating
particulate drug delivery. European Journal of Pharmaceutics and Biopharmaceutics,
98, 76–89.
Neuse, E. W. (2008). Synthetic polymers as drug-delivery vehicles in medicine. Metalbased Drugs, 2008.
Nilsen-Nygaard, J., Strand, S. P., Vårum, K. M., Draget, K. I., & Nordgård, C. T. (2015).

Chitosan: Gels and interfacial properties. Polymers, 7(3), 552–579.
Nilsson, K., Risberg, B., & Heimer, G. (1995). The vaginal epithelium in the
postmenopause—Cytology, histology and pH as methods of assessment. Maturitas,
21(1), 51–56.
Nwe, N., Furuike, T., & Tamura, H. (2010). Chitin and chitosan from terrestrial
organisms. Chitin, Chitosan, Oligosaccharides and Their Derivatives: Biological Activities
and Applications, 3–10.
Paczkowska, M., Chanaj-Kaczmarek, J., Romaniuk-Drapała, A., Rubi´s, B.,
Szymanowska, D., Kobus-Cisowska, J., … Cielecka-Piontek, J. (2020). Mucoadhesive
chitosan delivery system with Chelidonii Herba Lyophilized extract as a promising
strategy for vaginitis treatment. Journal of Clinical Medicine, 9(4), 1208. https://doi.
org/10.3390/jcm9041208
Palmeira-de-oliveira, R., Palmeira-de-oliveira, A., & Martinez-de-oliveira, J. (2015). New
strategies for local treatment of vaginal infections. Advanced Drug Delivery Reviews,
92, 105–122. />Palmeira-De-Oliveira, A., Ribeiro, M. P., Palmeira-De-Oliveira, R., Gaspar, C., Costa-DeOliveira, S., Correia, I. J., et al. (2010). Anti-Candida activity of a chitosan hydrogel:
Mechanism of action and cytotoxicity profile. Gynecologic and Obstetric Investigation,
70(4), 322–327. />Palmeira-De-Oliveira, R., Duarte, P., Palmeira-De-Oliveira, A., Das Neves, J.,
Amaral, M. H., Breitenfeld, L., et al. (2015). Women’s experiences, preferences and
perceptions regarding vaginal products: Results from a cross-sectional web-based
survey in Portugal. The European Journal of Contraception and Reproductive Health
Care. />Pathak, J., Ahmed, H., Singh, D. K., Pandey, A., Singh, S. P., Sinha, R. P., et al. (2019).
Recent developments in green synthesis of metal nanoparticles utilizing
cyanobacterial cell factories. Nanomaterials in plants, algae and microorganisms (pp.
237–265). Elsevier.
Perinelli, D. R., Campana, R., Skouras, A., Bonacucina, G., Cespi, M., Mastrotto, F., …
Casettari, L. (2018). Chitosan loaded into a hydrogel delivery system as a strategy to
treat vaginal Co-infection. Pharmaceutics, 10(1), 1–15. />pharmaceutics10010023
Perioli, L., Ambrogi, V., Venezia, L., Pagano, C., Ricci, M., & Rossi, C. (2008). Chitosan
and a modified chitosan as agents to improve performances of mucoadhesive vaginal
gels. Colloids and Surfaces B, Biointerfaces, 66(1), 141–145. />j.colsurfb.2008.06.005

Pradines, B., Bories, C., Vauthier, C., Ponchel, G., Loiseau, P. M., & Bouchemal, K.
(2015). Drug-free chitosan coated poly(isobutylcyanoacrylate) nanoparticles are
active against trichomonas vaginalis and non-toxic towards pig vaginal mucosa.
Pharmaceutical Research, 32(4), 1229–1236. />Raafat, D., & Sahl, H.-G. (2009). Chitosan and its antimicrobial potential–a critical
literature survey. Microbial Biotechnology, 2(2), 186–201.
Rampersaud, R., Randis, T. M., & Ratner, A. J. (2012). Microbiota of the upper and lower
genital tract. Seminars in Fetal and Neonatal Medicine, 17, 51–57.
Rein, M. F. (2020). Trichomoniasis. Hunter’s tropical medicine and emerging infectious
diseases. Elsevier. />Rinaudo, M. (2006). Chitin and chitosan: Properties and applications. Progress in Polymer
Science, 31(7), 603–632.
Robert, M. E., Eric, J. B., et al. (1992). Chitin chemistry, the.
Roberts, G. A. F. (1992). Structure of chitin and chitosan. Chitin chemistry (pp. 1–53).
London: Macmillan Education UK. />Rodero, C. F., Fioramonti Calixto, G. M., dos Santos, K. C., Sato, M. R., dos Santos
Ramos, M. A., Miro, M. S., … Chorilli, M. (2018). Curcumin-loaded liquid crystalline
systems for controlled drug release and improved treatment of vulvovaginal
candidiasis. Molecular Pharmaceutics, 15(10), 4491–4504. />acs.molpharmaceut.8b00507
Rodrigues, S., Dionísio, M., L´
opez, C. R., & Grenha, A. (2012). Biocompatibility of
chitosan carriers with application in drug delivery. Journal of Functional Biomaterials.
/>
10


V.H.S. Araujo et al.

Carbohydrate Polymers 261 (2021) 117919
Sudarshan, N. R., Hoover, D. G., & Knorr, D. (1992). Antibacterial action of chitosan.
Food Biotechnology, 6(3), 257272.
Taipaleenmă
aki, E., & Stă

adler, B. (2020). Recent advancements in using polymers for
intestinal mucoadhesion and mucopenetration. Macromolecular Bioscience, 20(3),
Article 1900342.
Tavassoli, M., Imani, A., Tajik, H., Moradi, M., & Pourseyed, S. H. (2012). Novel in vitro
efficiency of Chitosan biomolecule against Trichomonas gallinae. Iranian Journal of
Parasitology.
Tentor, F., Siccardi, G., Sacco, P., Demarchi, D., Marsich, E., Almdal, K., … Boisen, A.
(2020). Long lasting mucoadhesive membrane based on alginate and chitosan for
intravaginal drug delivery. Journal of Materials Science Materials in Medicine, 31(3).
/>Valenta, C. (2005). The use of mucoadhesive polymers in vaginal delivery. Advanced
Drug Delivery Reviews, 57(11), 1692–1712. />addr.2005.07.004
Victorelli, F. D., Calixto, G. M. F., dos Santos, K. C., Buzz´
a, H. H., & Chorilli, M. (2021).
Curcumin-loaded Polyethyleneimine and chitosan polymer-based Mucoadhesive
liquid crystalline systems as a potential platform in the treatment of cervical Cancer.
Journal of Molecular Liquids. />Victorelli, F. D., Calixto, G. M. F., Dos Santos Ramos, M. A., Bauab, T. M., & Chorilli, M.
(2018). Metronidazole-loaded polyethyleneimine and chitosan-based liquid
crystalline system for treatment of staphylococcal skin infections. Journal of
Biomedical Nanotechnology. />Vllasaliu, D., Exposito-Harris, R., Heras, A., Casettari, L., Garnett, M., Illum, L., et al.
(2010). Tight junction modulation by chitosan nanoparticles: Comparison with
chitosan solution. International Journal of Pharmaceutics, 400(1–2), 183–193.
Vunain, E., Mishra, A. K., & Mamba, B. B. (2017). Fundamentals of chitosan for
biomedical applications. Chitosan based biomaterials volume 1 (pp. 3–30). Elsevier.
Wang, C., Li, J., & Yao, F. (2011). Application of chitosan-based biomaterials in tissue
engineering. Chitosan-based hydrogels. CRC Press. />Willems, H. M. E., Ahmed, S. S., Liu, J., Xu, Z., & Peters, B. M. (2020). Vulvovaginal
candidiasis: A current understanding and burning questions. Journal of Fungi, 6(1),
27. />WORLD HEALTH ORGANIZATION. (2012). Global incidence and prevalence of selected
curable sexually transmitted infections – 2008. Switzerland: WORLD HEALTH
ORGANIZATION.
Yano, J., Sobel, J. D., Nyirjesy, P., Sobel, R., Williams, V. L., Yu, Q., … Fidel, P. L. (2019).

Current patient perspectives of vulvovaginal candidiasis: Incidence, symptoms,
management and post-treatment outcomes. BMC Women’s Health, 19(1), 48.
Zheng, L. Y., & Zhu, J. F. (2003). Study on antimicrobial activity of chitosan with
different molecular weights. Carbohydrate Polymers. />carbpol.2003.07.009
Zubareva, A., Shagdarova, B., Varlamov, V., Kashirina, E., & Svirshchevskaya, E. (2017).
Penetration and toxicity of chitosan and its derivatives. European Polymer Journal,
93, 743–749. />ˇ Potrˇc, T., Baumgartner, S., Kocbek, P., & Kristl, J. (2016). Formulation and
Zupanˇciˇc, S.,
evaluation of chitosan/polyethylene oxide nanofibers loaded with metronidazole for
local infections. European Journal of Pharmaceutical Sciences, 95, 152–160. https://
doi.org/10.1016/j.ejps.2016.10.030

Roggero, E., P´erez, A. R., Bottasso, O. A., Besedovsky, H. O., & Del Rey, A. (2009).
Neuroendocrine-immunology of experimental chagas’ disease. Annals of the New
York Academy of Sciences, 1153(1), 264–271.
Ruiz-S´
anchez, D., Calder´
on-Romero, L., S´
anchez-Vega, J. T., & Tay, J. (2003). Intestinal
candidiasis. A clinical report and comments about this opportunistic pathology.
Mycopathologia, 156(1), 9–11.
Salmazi, R., Calixto, G., Bernegossi, J., Aparecido Dos, M., Ramos, S., Bauab, T. M., et al.
(2015). A Curcumin-loaded liquid crystal precursor mucoadhesive system for the
treatment of vaginal candidiasis. International Journal of Nanomedicine. https://doi.
org/10.2147/IJN.S82385
Sandri, G., Rossi, S., Bonferoni, M. C., Ferrari, F., Mori, M., & Caramella, C. (2012). The
role of chitosan as a mucoadhesive agent in mucosal drug delivery. Journal of Drug
Delivery Science and Technology. />Sangamithra, V., Verma, R., Sengottuvelu, S., & Sumathi, R. (2013). Candida infections
of the genitourinary tract. Research Journal of Pharmacy and Technology, 6(10),
1111–1115. />Shahabeddin, L., Damour, O., Berthod, F., Rousselle, P., Saintigny, G., & Collombel, C.

(1991). Reconstructed skin from co-cultured human keratinocytes and fibroblasts on
a chitosane cross-linked collagen-GAG matrix. Journal of Materials Science Materials
in Medicine. />Shahidi, F., Arachchi, J. K. V., & Jeon, Y. J. (1999). Food applications of chitin and
chitosans. Trends in Food Science & Technology, 10(2), 37–51. />10.1016/S0924-2244(99)00017-5
Shukla, S. K., Mishra, A. K., Arotiba, O. A., & Mamba, B. B. (2013). Chitosan-based
nanomaterials: A state-of-the-art review. International Journal of Biological
Macromolecules. />Singh, A., & Campus, B. (2018). CHITIN, CHITOSAN AND THEIR PHARMACOLOGICAL
ACTIVITIES: A REVIEW Kaushal Tripathi and Anita Singh * department of
pharmaceutical sciences, bhimtal campus, kumaun university, Nainital - 263001,
uttarakhand, India. International Journal OfPharmaceutical Sciences And Research, 9
(7), 2626–2635. />Smith, J., Wood, E., & Dornish, M. (2004). Effect of chitosan on epithelial cell tight
junctions. Pharmaceutical Research, 21(1), 43–49.
Soares, G., Santos, D., Pereira, G. G., Bender, E. A., Colom´
e, M., & Guterres, S. S. (2012).
`
Desenvolvimento e Caracterizaỗ
ao de Nanopartớculas LIPIDICAS DESTINADAS A
TOPICA

APLICAầAO
DE DAPSONA. Quimica Nova, 35.

Soares, L. de S., Perim, R. B., de Alvarenga, E. S., Guimar˜
aes, L. M., Teixeira, A. V. N. C.,
Coimbra, J. S. R., et al. (2019). Insights on physicochemical aspects of chitosan
dispersion in aqueous solutions of acetic, glycolic, propionic or lactic acid.
International Journal of Biological Macromolecules. />ijbiomac.2019.01.106
Sobahi, T. R. A., Abdelaal, M. Y., & Makki, M. S. I. (2014). Chemical modification of
chitosan for metal ion removal. Arabian Journal of Chemistry, 7(5), 741–746.
Sogias, I. A., Williams, A. C., & Khutoryanskiy, V. V. (2008). Why is chitosan

mucoadhesive? Biomacromolecules, 9(7), 1837–1842. />bm800276d
Sreekumar, S., Goycoolea, F. M., Moerschbacher, B. M., & Rivera-Rodriguez, G. R.
(2018). Parameters influencing the size of chitosan-TPP nano- and microparticles.
Scientific Reports, 8(1), 4695. />Srikrishna, S., & Cardozo, L. (2013). The vagina as a route for drug delivery: A review.
International Urogynecology Journal and Pelvic Floor Dysfunction, 24(4), 537–543.
/>
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