![Comparison of electron micrographs of plant cellulose (A) and bacterial nanocellulose (BNC) fibers (B) (reprinted with permission from [42]); electron micrograph of a cross-section of BNC (C) (Reprinted with permission from [29]).](https://www.researchgate.net/publication/349990237/figure/fig1/AS:1010724386115589@1617986880437/Comparison-of-electron-micrographs-of-plant-cellulose-A-and-bacterial-nanocellulose_Q320.jpg)
![Skin coloring 12 h after removal of BNC-DHA patches (concentration of DHA is expressed in percent), applied for 30 min (a) (reprinted with permission from [110]); Schematic representation of the HA-(BNC-R) MNs structure (b) and functioning of this innovative system: insertion of the MNs into the skin, dissolution of HA MNs and subsequent release of the bioactive molecule from the BNC membrane (c) (reprinted with permission from [119]). DHA: 1,3-dihydroxy-2-propanone, HA: hyaluronic acid, MN: microneedles, R: rutin.](https://www.researchgate.net/publication/349990237/figure/fig4/AS:1010724386131976@1617986880704/Skin-coloring-12-h-after-removal-of-BNC-DHA-patches-concentration-of-DHA-is-expressed-in_Q320.jpg)
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International Journal of
Molecular Sciences
Review
Bacterial Nanocellulose toward Green Cosmetics: Recent
Progresses and Challenges
Tânia Almeida, Armando J. D. Silvestre , Carla Vilela and Carmen S. R. Freire *
Citation: Almeida, T.; Silvestre,
A.J.D.; Vilela, C.; Freire, C.S.R.
Bacterial Nanocellulose toward Green
Cosmetics: Recent Progresses and
Challenges. Int. J. Mol. Sci. 2021,22,
2836. https://doi.org/10.3390/
ijms22062836
Academic Editor: Jeannine
M. Coburn
Received: 18 February 2021
Accepted: 9 March 2021
Published: 11 March 2021
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations.
Copyright: © 2021 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
CICECO—Aveiro Institute of Materials, Department of Chemistry, University of Aveiro,
3810-193 Aveiro, Portugal; taniaralmeida@ua.pt (T.A.); armsil@ua.pt (A.J.D.S.); cvilela@ua.pt (C.V.)
*Correspondence: cfreire@ua.pt
Abstract:
In the skin care field, bacterial nanocellulose (BNC), a versatile polysaccharide produced
by non-pathogenic acetic acid bacteria, has received increased attention as a promising candidate to
replace synthetic polymers (e.g., nylon, polyethylene, polyacrylamides) commonly used in cosmetics.
The applicability of BNC in cosmetics has been mainly investigated as a carrier of active ingredients
or as a structuring agent of cosmetic formulations. However, with the sustainability issues that are
underway in the highly innovative cosmetic industry and with the growth prospects for the market of
bio-based products, a much more prominent role is envisioned for BNC in this field. Thus, this review
provides a comprehensive overview of the most recent (last 5 years) and relevant developments
and challenges in the research of BNC applied to cosmetic, aiming at inspiring future research to go
beyond in the applicability of this exceptional biotechnological material in such a promising area.
Keywords:
bacterial nanocellulose; green cosmetics; facial mask; skin active substances; carrier;
cosmetic formulations
1. Introduction
Consumers are increasingly aware of the current issues about environmental pollution
and sustainability that the world is facing. The depletion of natural resources, waste
generation, climate changes, and water and air pollution are among the main challenges
to be tackled [
1
]. There is also an increasing concern with healthcare and lifestyle that
has changed consumer trends and is driving the demand for natural products which at
the same time are expected to be more effective and safer, especially in sectors such as
cosmetics and food [
2
,
3
]. The ambitious goals and targets for the next years set by the world
authorities for environment and sustainable development, in the scope of the 2030 Agenda
for Sustainable Development, are prompting countries to adopt more stringent environ-
mental policies [
4
]. For instance, this involves regulating the production and sales of plastic
bags and single-use plastics items, the phase-out of the intentional use of microplastics, the
labeling of harmful ingredients or even the taxes applied to the waste management, includ-
ing recycling
[5,6]
. These policies are essential to stimulate green development [
7
] that has
also been influencing how companies are evolving and becoming “greener”. In particular,
the cosmetic industry is an ever-growing sector that is among the most innovative and
science-driven industries, in which we have also been witnessing a great effort to follow
this global trend for natural, environmentally friendly and sustainable production and
products [
8
]. The environmental impact of cosmetic products lies mainly in the extensive
use of non-renewable and non-biodegradable raw materials [9]. An example of this is the
broad use of plastic microbeads as decorative or exfoliator agents or the use of parabens
as preservatives, silicones as emollients or emulsifiers, or even the inclusion of chemical
and mineral UV filters in many cosmetic products [
8
]. Thus, since most cosmetics are
used daily, the addition of these ingredients to the cosmetic formulation raises significant
pollution concerns. In addition, the manufacturing process and the packaging are two
Int. J. Mol. Sci. 2021,22, 2836. https://doi.org/10.3390/ijms22062836 https://www.mdpi.com/journal/ijms
Int. J. Mol. Sci. 2021,22, 2836 2 of 25
other important stages/steps to consider in the environmental life cycle assessment (LCA)
of a cosmetic product [8,10].
In this sense, there has been a growing investment by the cosmetic industry in develop-
ing green-tech solutions, either to replace synthetic ingredients by bio-based counterparts
or to modify the production technology or even to redesign and develop recyclable and
biodegradable cosmetic packaging materials [8,11].
In the “green” advent of the cosmetic industry, some biopolymers have gained high
popularity not only because of their biodegradable and environmentally friendly nature
but also due to their biocompatibility that categorize them as skin-friendly materials [
12
].
Biopolymers, such as proteins (e.g., collagen and wheat proteins) and polysaccharides
(e.g., cellulose, alginic acid, and hyaluronic acid), are already widely used as ingredients
in cosmetic formulations to ameliorate properties of the products (e.g., texture and con-
sistency) or boost their action on the body (e.g., hydration and wrinkles reduction) [
13
].
Furthermore, currently, biopolymers are also recognized as a viable and sustainable alter-
native to petroleum-based polymers commonly used in cosmetic packaging [11].
Cellulose is the most abundant biopolymer worldwide, which together with its re-
markable properties (e.g., hydrophilicity, extensive modification capability, and thermal
stability up to about 200
◦
C) make it an attractive source of new sustainable materials for
many industrial applications [
14
]. Cellulose is obtained mainly from plants, but it is also
produced by bacteria, fungi, algae, and marine animals [
15
]. Irrespective of its source, it is
a linear polysaccharide of
β
-D-glucopyranose units linked by
β
-1,4-glycosidic bonds [
16
].
Although plant cellulose is the most used cellulose substrate, bacterial nanocellulose (BNC,
also known as microbial cellulose, bacterial cellulose or biocellulose), is currently regarded
as a promising cellulose form for a wide range of applications, including in cosmetic [
17
,
18
],
food [18–20], pharmaceutical and biomedical [21–26], and technological fields [27–30].
BNC is a nano scale form of cellulose that is biotechnologically produced as an
exopolysaccharide by some aerobic non-pathogenic bacteria (e.g., Komagataeibacter (for-
merly Acetobacter), Agrobacterium,Aerobacter,Achromobacter,Azotobacter,Rhizobium,Sarcina,
Salmonella, and Escherichia), which combines the properties of cellulose with the features of
nanomaterials [31,32].
The applicability of BNC in cosmetics has been explored and demonstrated over
the years, especially as a support material of sheet facial masks for the skin delivery of
active substances. However, BNC has also been employed in natural scrub cosmetics
or as a structuring agent in personal care formulations [
18
,
21
]. In fact, some BNC-based
cosmetic products are already available in the market, which are many times advertised
as biocellulose and associated with ingredients that frequently place these products in
the group of “luxury” cosmetics [
33
]. However, its application is still limited despite its
recognized high-performance and unique set of properties. However, considering the
growth forecasts for the personal care market, which is expected to grow at a compound
annual growth rate (CAGR) of about 5.0% over the next five years [
34
], and the high
demand for bio-based products, it is foreseen that the research and commercial interest
on the application of BNC in green cosmetics will greatly increase in the near future. This
was also envisaged in a very recent bibliometric appraisal covering the applicability of
BNC in cosmetics in which it is shown that the number of publications and interest on this
topic have been gradually increasing in recent years [
17
]. This study also emphasizes the
reduced number of detailed bibliographic reviews on this subject that might provide an
overview of the actual status and the research trends that will foster new ideas [
17
]. Hence,
in the current review, the most recent and relevant investigation results and developments
on BNC applied to cosmetics are comprehensively systematized. From this perspective,
not only research articles but also patents will be covered, because although registered
patents not always result in commercial products, they are a good indicator of market
trends and of further directions for research and investment [
35
]. The challenges that are
hampering a faster progress of the commercial application of BNC and future perspectives
Int. J. Mol. Sci. 2021,22, 2836 3 of 25
are also highlighted, aiming to spur future research and go further in the application of
this exceptional biopolymer in such a promising area.
2. Biosynthesis and Production Methods of Bacterial Nanocellulose
Among BNC-producing bacteria, those from the Komagataeibacter genus are recognized
as the most effective [
36
]. Komagataeibacter bacteria belong to the acetic acid bacteria group
and are commonly present in sugar-rich beverages and fruit products and residues [
36
].
In particular, the species Komagataeibacter xylinus, owing to its capability in using a wide
range of sugars and the higher production rates of BNC, is considered a microbial model
to study the BNC biosynthetic pathway [
37
]; more recently, the Komagataeibacter hanseiin is
also capturing increasing interest as a model microorganism, especially because it is also
a high-yield BNC producer [
16
,
38
]. In addition to these species, others have also demon-
strated high productivities, being also recognized as efficient BNC producers, namely, the
Gluconacetobacter sacchari isolated from Kombucha tea [
39
] or the K. rhaeticus (strain PG2)
isolated from pomegranate [40].
The biosynthesis of BNC is a well-concerted and highly regulated process involving
multiple enzymes and complexes of catalytic and regulatory proteins that can be divided
in two main stages: first, the production of
β
-1,4-glucan chains, and then the assembly and
crystallization of cellulose [
41
]. Succinctly, the process initiates with the transport of the
carbon source (e.g., glucose and fructose) into the cell, where the cellulose precursor uridine
diphosphoglucose (UDPG) is formed. Then, glucose is polymerized from UDPG into
β
-1,4-
glucan chains by cellulose synthase. Finally, the cellulose chains are secreted through pores
in the cell membrane as sub-fibrils, which are further assembled into ribbons organized in
a 3D nanofiber network stabilized by hydrogen bonds [
41
,
42
]. The biosynthetic pathway
of BNC and associated mechanisms were addressed in detail in previous reviews [
37
,
41
].
More recently, Jacek et al. [
16
] reviewed the latest advances regarding the molecular aspects
behind BNC biosynthesis.
BNC is the result of an oxidative fermentation process, with the oxidation of sugars
and organic acids that can be produced either in synthetic or non-synthetic sugar and
nitrogen-rich media at temperatures ranging between 25–30
◦
C and pH 4.5–7.5 [
43
]. There
are two general methods for BNC production: static culture (in flasks or trays) and agitated
culture (in jar fermenters). The most used method at a laboratory scale is the static culture,
in which BNC is accumulated as a gelatinous membrane (hydrogel-like) (Figure 1A–C)
at the air–medium interface, which thickens with culture time [
14
,
44
]. The formation of
the floating BNC membrane helps bacteria access oxygen in the surface medium, and it is
generally accepted that it functions as a barrier to protect bacteria from radiation, natural
inhibitors, and drying [
41
,
42
]. A static fed-batch methodology may also be used by the
periodic addition of culture medium with the formation of a new BNC membrane each time,
thereby increasing the expected production [
45
,
46
]. The traditional static culture still offers
the possibility of placing a mold in the culture medium to produce a BNC membrane with
defined shapes, which is favorable for applications with a required predefined form [
20
].
Despite being widely used at the laboratory scale and by some traditional industries
(producers of nata de coco, mainly in Asia), static culture has some limitations that still
hinder its broader use at the industrial level. Specifically, it is labor-intensive, needs
high amounts of culture medium, has a long cultivation time (several days), and has low
productivity, which make it a high-cost and inefficient method [47,48].
Alternatively, in agitated culture, BNC is generated in variable shapes and sizes,
namely fibrous suspension, irregular masses, pellets, or spheres (Figure 1D–F), depending
on various factors (e.g., rotating speed, carbon source, and culture time) [
44
,
49
]. Whereas
in static culture, BNC production is limited by the restricted air surface area, in the agitated
culture, the distribution of oxygen and nutrients in liquid medium is more homogeneous,
and therefore, an improved productivity may be expected [
24
]. However, some shortcom-
ings are also associated with it. One of the major challenges is the genetic instability related
to some BNC producer strains (e.g., from Komagataeibacter genus) that under agitation
Int. J. Mol. Sci. 2021,22, 2836 4 of 25
may form spontaneous cellulose-negative (Cel
−
) mutants (viz., non-cellulose producers),
thereby lowering the overall productivity [
16
]. Another factor that may negatively influ-
ence the productivity of BNC under agitation is the non-Newtonian behavior of BNC in
the fermentation medium that will cause a significant increase in the viscosity with the
consequent reduction of homogeneity and distribution of oxygen in the medium [50,51].
Figure 1.
Bacterial nanocellulose shapes: bacterial nanocellulose (BNC) membranes produced in
static fed-batch conditions (
A
); wet BNC membrane produced in static culture in Hestrin–Schramm
(HS) medium, before purification (
B
) and after purification (
C
) (adapted with permission from [
52
]);
BNC spheres produced under agitated conditions using mannitol (
D
), glucose (
E
), and xylitol (
F
) as
carbon source (reprinted with permission from [53]).
With the increasing interest in the commercial exploitation of BNC, there has been a
significant investment in research to overcome the limitations of both static and agitated
cultures in order to achieve high production rates, reduced production costs, and/or shorter
cultivation times [
44
]. The design and development of efficient bioreactors has been one of
the explored strategies for the scale-up. In this context, agitated culture bioreactors from
the airlift [
54
] or modified airlift spherical bubble column reactor types [
55
] are among
the ones tested to produce BNC pellets, fibrous material, or sphere-like particles. The
production of BNC membranes or pellicles has also been investigated using different
bioreactor typologies, including membrane [
56
], horizontal lift [
57
], rotating disk [
58
], or
modified airlift [
59
]. Despite the advances in this field, the use of bioreactors to produce
BNC is still limited and is mostly reduced to some tests at the pilot scale. The research done
on bioreactors to produce BNC was covered in detail by previous reviews [44,47,48,60].
As in other microbial fermentation processes, the culture medium has a high con-
tribution to the overall cost of BNC production [
61
]. In addition to the commonly used
synthetic culture medium HS, other media have also been investigated and, depending on
those, the BNC production cost can be so distinct as ca.
€
60 to ca.
€
9000 per kilogram [
62
].
In this context, over the last few years, as a means to decrease the BNC production costs,
several studies have investigated and reported the use of agro-industrial by-products as a
fermentation medium with or without specific nutritional supplementation (notably with
N and P sources), showing promising results in terms of productivity when compared
Int. J. Mol. Sci. 2021,22, 2836 5 of 25
with the HS medium. Examples include the works using residues of wine and pulp in-
dustries [
63
] or dry olive mill residue water extracts [
64
] and, more recently, citrus peel
and pomace [
65
], corn steep liquor [
52
], beverage industrial waste, tomato juice [
66
], palm
date, fig, and sugarcane molasses [
67
], and cashew apple juice and soybean molasses [
68
],
indicating that this can be a promising approach to reduce the BNC production costs.
Finally, the importance of the bacterial strain and its growth characteristics in the global
process of BNC production is also well known, and the isolation of more efficient and
productive bacterial strains has deserved attention by researchers. However, with the
advent of genome sequencing, genetic engineering appears as a more promising approach
to obtain more stable and high-yield mutant strains, despite the regulatory issues that
some fields may encounter [
16
,
48
,
69
]. BNC production methods were recently addressed
in two different reviews [
70
,
71
] with discussion about several biotechnological approaches
to optimize BNC production [70].
3. Properties and Applications of Bacterial Nanocellulose
BNC is produced as a well-organized 3D network of ribbon-shaped nano- and mi-
crofibrils with extremely reduced diameter when compared with plant cellulose fibers
(Figure 2A,B) [
42
]. An electron microscopy cross-section image of a BNC membrane
(Figure 2C)
shows that BNC fibers are organized as stratified layers, bundled one after
the other as the bacteria grow at the culture medium–air interface [
29
]. Another prop-
erty of BNC that stands out over plant cellulose is its high chemical purity, being free of
lignin, hemicelluloses, and pectin, typically present in plant fibers [
14
]. This eliminates
the need for complex purification procedures, which are energy and chemically demand-
ing, with reduced yields that may affect the structure and physicochemical properties of
cellulose [72].
BNC substrate is characterized by a highly porous nanofibrillar structure, with pore
diameters typically below 10
µ
m, that results in notable permeability to gases and liquids
and a very high water-binding capacity (water content >90%) [
26
,
42
,
73
]. It is also character-
ized by a high crystallinity index (80–90%) that contributes to the high thermal stability of
BNC, which is an important characteristic when thermal sterilization is required [
36
]. BNC
shows a high degree of polymerization, typically between 4000 and 10,000 anhydro-glucose
units [
42
]. Due to its crystalline nanofibrillar structure, BNC also shows excellent mechani-
cal properties. Despite some disparity in the literature data, it is generally assumed to have
a tensile strength between 200 and 300 MPa and a Young’s modulus reaching values as
high as 15–35 GPa [36].
In addition to the remarkable physicochemical and mechanical properties, BNC
presents also high
in vivo
skin biocompatibility, as demonstrated in studies performed
with human volunteers [
74
,
75
], which is a particular attractive feature when intended to be
applied in skin-care applications.
In addition to shape, the supramolecular structure of BNC, as well as its physical
and mechanical properties can be modulated during biosynthesis by altering conditions,
such as the production method [
44
], carbon source [
76
], bacteria strain [
77
], or even by
adding additives to the culture medium [
78
], this being a great advantage over plant cellu-
lose. Therefore, these parameters should be carefully selected and controlled considering
the desired properties and intended application. In addition, as any cellulose substrate,
BNC is also prone to be modified ex situ (after purification), either by chemical treatment
(e.g., acetylation, alkylation, carboxymethylation, among others) or by the incorporation
of other materials into the BNC network [
79
,
80
]. The high surface area of BNC fibrils con-
tributes to the establishment of interactions between the abundant hydroxyl groups in the
surface of BNC and functional groups of other components, such as active low-molecular-
weight compounds [
75
,
81
], polymers [
82
–
85
], metal oxides, or metal nanoparticles [
86
–
89
].
The combination with other materials imparts novel or improved properties to the pristine
BNC, such as bioactivity (e.g., antioxidant, anti-inflammatory, and antimicrobial activi-
ties), conductive, optical, or magnetic properties or even improved mechanical properties,
Int. J. Mol. Sci. 2021,22, 2836 6 of 25
that greatly extends the potential of BNC and allows designing high-value functional
BNC-based materials with application in several fields [
79
]. The progress done in the
development of functionalized BNC-based materials has been addressed in several reviews
covering the strategies applied to enhance BNC properties that are valued in areas such as
sensors, photocatalytic nanomaterials, optoelectronic materials and devices, magnetically
responsive membranes [
79
], drug delivery [
21
,
22
], tissue engineering [
80
,
90
], wound dress-
ing [
26
], or food and food packaging [
20
]. The functionalization of BNC through chemical
modification or in combination with other polymers or active compounds has also been
a successful strategy for the exploitation of BNC in cosmetics, as will be discussed in the
next section. The properties and main applications of BNC are summarized in Figure 3.
Figure 2.
Comparison of electron micrographs of plant cellulose (
A
) and bacterial nanocellulose
(BNC) fibers (
B
) (reprinted with permission from [
42
]); electron micrograph of a cross-section of BNC
(C) (Reprinted with permission from [29]).
Figure 3. General properties and applications of bacterial nanocellulose.
Int. J. Mol. Sci. 2021,22, 2836 7 of 25
4. Application of Bacterial Nanocellulose in Cosmetics
Although there are slight variations in cosmetic definition among the three main
trading blocks of the cosmetic market, viz., Europe, USA, and Japan, a cosmetic can
be generally defined as a product used to improve the appearance of the human body
without affecting its basic structure [
91
]. More recently, another concept was introduced,
the cosmeceuticals, which are categorized as products associating features of cosmetics
and pharmaceuticals. They incorporate active substances with therapeutic benefits and
are applied to treat the most skin-relevant conditions such as photoaging, wrinkles, skin
dehydration, dark spots, or hyperpigmentation [
91
,
92
]. BNC encounters several potential
applications in cosmetics, due to its exceptional properties as referred above, mainly as
a support for skin-active substances or as a structuring agent in cosmetic formulations.
Table 1summarizes the most recent (last 5 years) and relevant developments regarding
the application of BNC or BNC-based materials in cosmetics, which are achieved in both
academic and industrial contexts. The subsequent sections will highlight the use of BNC in
cosmetics, notably as a carrier of skin active substances, enzyme immobilizer, emulsion
stabilizer, or as an alternative to microplastics, aiming to demonstrate the enormous
potential of this remarkable biopolymer as a sustainable alternative for the design of green
cosmetics (Figure 4).
Figure 4. Main applications of bacterial nanocellulose in cosmetics.
4.1. Bacterial Nanocellulose as Carrier of Skin Active Substances
BNC has attracted much attention as a delivery system because of its peculiar porous
nanostructure that favors the incorporation and release of active substances with interest
for areas such as biomedical or cosmetic [
93
,
94
]. The successful application of BNC in
the transdermal drug release of active substances has been widely demonstrated for both
hydrophobic and hydrophilic drugs [
22
], but the number of studies addressing the BNC
release of skin-active substances in cosmeceuticals is more limited. However, over the last
years, the suitability and applicability of BNC as the support material of sheet facial masks
enriched with active substances, taking advantage of its high mechanical performance,
loading capacity, and membrane-like shape, have been increasingly investigated.
Sheet facial masks are typically made of a diversity of fabrics (e.g., cotton, non-woven)
and synthetic polymers (e.g., poly (vinyl alcohol) (PVA)) and are currently one of the most
popular and fast-growing types of facial products, thus representing an important and
highly competitive niche of the cosmetic market [
95
,
96
]. The popularity of facial sheet
masks is largely due to its easy application and removal, fast use, and effective results that
make them very appealing for the nowadays consumers, compared with other mask types
(e.g., rinse-off or peel-out) [
97
]. The mechanical strength and elasticity of BNC (in the wet
Int. J. Mol. Sci. 2021,22, 2836 8 of 25
state) make it particularly resistant to handle, easy to use, and enable a perfect adhesion to
skin, enhancing the penetration of active substances into the skin. All these aspects make
BNC an exceptional bio-based material for this application and, in fact, some sheet masks
made of BNC are already commercially available, such as the CELMAT
®
(BOWIL—Biotech
Ltd, Wladyslawowo, Poland), Intense Whitening Biocellulose Mask (Elizabeth Arden, New
York, NY, USA), DHC Bio Cellulose Mask (DHC, Tokyo, Japan), Blank Expert Second
Whitening Bio-Cellulose Mask (Lancôme, Paris, France), or Second Skin Mask with Aloe
vera (CORYMER, Neerwinden, Belgium) [33], just to enumerate some examples.
In the production of BNC-based facial masks, several works have adopted a bottom–
up approach to incorporate the skin-active substances, in particular, natural compounds,
into the BNC membrane by adding them to the culture medium, thus promoting their
incorporation within the BNC fibril network during biosynthesis, which in theory turns
the scale-up easier and may reduce the costs. This was the approach used in the method
claimed in a patent by Ho et al. [
98
] to produce BNC facial masks, in which BNC was
biosynthesized in a culture medium containing a bamboo extract (0.01–100%, w/w), which
may be supplemented with additional nitrogen and carbon sources to increase the BNC
yield. After production, the BNC membrane was purified by alkaline treatment, sterilized
(ideally by hydrothermal sterilization), and dried. The presence of phenolic and carbonyl
compounds from bamboo extract was confirmed in BNC membranes after biosynthesis.
However, it is not clear if this evaluation was done before and/or after the purification
and sterilization. Furthermore, the BNC facial mask was soaked in a cosmetic emulsion
and combined with a facial scrub (particles selected from the group consisting of beads,
including jojoba esters, alginate beads, and mixtures thereof in rose powder). The bam-
boo extract (rich in flavonoids, lactones, and phenolic acids, among others) is known to
have antioxidant, antiallergic, and skin-whitening properties [
99
,
100
]. Tests with volun-
teers showed a superior performance in terms of adhesion, skin elasticity, softness, and
moisturizing effect of the obtained BNC–bamboo extract facial mask comparing with a
commercially available rayon non-woven fabric-based mask impregnated with the same
cosmetic emulsion [
98
]. A method to produce BNC using Moringa oleifera leaves powder
as the nitrogen source in the presence of a protease and ethanol was also claimed in a
recent patent [
101
]. Moreover, after BNC biosynthesis, the resulting fermentation broth,
containing enzymatically hydrolyzed proteins, flavonoids, polyphenols, and organic acids
from the M. oleifera leaves, was recovered and used to prepare a cosmetic formulation that
was embedded in the biosynthesized BNC membrane. Results in volunteers showed that
the BNC membrane with the M. oleifera fermentation broth-based formulation had a better
hydration effect than a normal membrane cloth loaded with the same cosmetic formulation.
With modern consumers demanding highly efficient skin care products, the cosmetic
industry has been investing in the development of formulations, including facial masks
with enhanced skin penetration and absorption [
102
]. In this context,
Gregoire et al. [103]
developed a patented method to prepare a composite sheet with a multilayer struc-
ture composed by BNC biosynthesized in the presence of Sphaerotilus natans-derived
microtubes (composed by a polysaccharide, commonly a deoxy sugar-modified (
β
1
→
4)-
linked glycosaminoglycan). Permeation assays using a synthetic membrane equivalent
to skin showed that the obtained BNC–microtubes composite sheet had an improved
transdermal delivery of cosmetic actives (caffein, minoxidil, 3-(2,4-dihydroxybenzyl-1-
2-(2-hydroxyethyl)-pyrrolidine-2,5-dione) and 4-(tetrahydro-pyran-4-yl)benzene-1,3-diol,
oxybenzone) compared with a BNC sheet without microtubes. This result was valid for
both water-soluble and lipid-soluble active ingredients [103].
Over the years, and in an attempt to reduce the costs of BNC production, several
works have been adopting the strategy of using food and agroforest by-products of natural
origin to biosynthesize BNC for cosmetic applications. An example of this is the method
claimed in a patent to produce BNC matrices using soybean molasses [
104
] as the nitrogen
and/or carbon sources of the BNC fermentation broth. The BNC matrix obtained with
soybean molasses substrate and immersed in a cosmetic essence with glycerin, polyethylene
Int. J. Mol. Sci. 2021,22, 2836 9 of 25
glycol, and carbomer was shown to have a high-water retention rate (98.35%) and good
moisturizing effect [
104
]. The authors claimed thereby that the soybean molasses can
be considered as a viable alternative to the standard HS medium for the production of
BNC matrices for facial masks, with economic and environmental profits. In another
patent, a method to produce BNC using expired milk or its derivatives (0.5–3.0% w/v)
as a carbon source was described [
105
]. In addition to expired milk, tea polyphenols
(0.01–0.10% w/v)
were also added to the culture medium to confer functional properties to
the BNC membrane. The use of milk and tea polyphenols resulted in an increase in the BNC
production yield when compared with glucose-based medium. Apparently, the content
of tea polyphenols was not determined in the BNC membranes after being purified by
alkaline treatment and subsequent washing until neutrality; however, the BNC membranes
demonstrated antioxidant and whitening properties [
105
], indicating that these bioactive
compounds were incorporated into BNC. The production of BNC by K. hansenii in a culture
medium containing 13% of tropical fruit by-products, known for having considerable
amounts of vitamins and minerals with recognized benefits for skin, was also reported
by Amorim et al. [
106
]. The results showed that the obtained BNC membrane presented
good mechanical performance and a high-water activity (99.13
±
0.09%). The authors also
confirmed the incorporation of ascorbic acid from the fruit residues (24.4
±
1.01 mg per
100 g cellulose) using the Tillmans method. The incorporation of this active substance into
BNC makes possible its use in cosmetic as a facial mask with hydrating and antioxidant
actions [106]; however, this was not demonstrated in the scope of the study.
The in situ incorporation of active substances into BNC by adding the potential
bioactive compounds to culture medium, although appealing in terms of scale-up, has to
take into consideration some important issues, namely, the fact that the BNC purification
process, after biosynthesis, has several washing steps with water and a treatment with an
alkaline solution, which may cause significant losses of water-soluble bioactive compounds.
Thus, to ensure the efficacy of the method, the effective incorporation of these compounds
should be carefully assessed by performing the structural characterization of the purified
BNC matrix. However, this evaluation is not always clearly shown, as was highlighted in
some of the studies/patents described above.
Alternatively, regarding the in situ incorporation approach, the pure BNC or BNC-
based composite membranes can be soaked in a solution of the active substances of interest,
which in fact is the most commonly adopted method for the production of functional
BNC materials for cosmetic applications. This was the strategy used by Silva et al. [
107
]
to investigate the potential of BNC membranes as a topical delivery system of caffein for
cellulite treatment. Scanning electron microscopy (SEM) micrographs showed that the
obtained BNC–caffein (caffein content of 8 mg cm
−2
) membranes were homogeneous with
no agglomerates’ formation. In addition, the
in vitro
permeation studies using epider-
mal membranes demonstrated that BNC–caffein membranes had lower permeation rates
than the ones obtained with conventional formulations (aqueous solution and gel) also
included in the assay. Specifically, BNC–caffein membranes presented a caffein flux of
2.55 µg cm−2h−1
and a cumulative amount permeated after 10 h of 0.39% (percentage of
applied dose), whereas the aqueous solution provided the highest flux
(7.53 µg cm−2h−1)
and percentage of caffein permeated (0.97%). As the authors point out, the complex 3D
network of BNC makes probably the diffusion pathway of caffein tortuous and might
be responsible for the slow release of caffein from BNC, which is desired for this type
of formulation whose effect is expected to persist over time. In addition, the use of this
BNC–caffein system also overcomes the lack of reproducibility of conventional gel formulas
regarding the dose of caffein that it is applied and losses caused by absorption due to
contact with clothing or other surfaces [107].
Numata et al. [
108
] have also followed a simple approach of BNC incorporation to
design a cosmetic delivery system, combining a BNC gel with poly(ethylene oxide)-b-
poly(caprolactone) (PEO-b-PCL) nanoparticles by diffusion of the nanoparticles into the gel.
To assess the effectiveness of the delivery system, PEO-b-PCL nanoparticles were loaded
Int. J. Mol. Sci. 2021,22, 2836 10 of 25
with retinol, a hydrophobic molecule, which is widely used as an anti-aging ingredient.
The releasing assays performed in acetic acid–sodium acetate buffer (pH 5.2) showed
that retinol is slowly released from PEO-b-PCL nanoparticles inside the BNC membrane;
however, the results also revealed that most of the retinol released from nanoparticles
precipitated and stayed retained in the BNC gel [
108
]. Despite the need for further studies
to overcome this problem, these findings are a good starting point to develop BNC-based
delivery systems for hydrophobic active substances that, owing to the intrinsic hydrophilic
nature of BNC, is still a challenging issue.
In the same vein, a method to produce a BNC mask with anti-inflammatory and
anti-allergic properties incorporating bee venom (rich in melittin) was claimed in a patent
by Hongli et al. [
109
]. In this study, BNC was used not only as the support of the bee
venom active components but also as emulsifier and thickener, in the form of nanofiber
filaments, of the water-soluble bee venom, replacing the traditional chemical thicken-
ers used in these formulations [
109
]. The use of BNC membranes as a carrier was also
evaluated for 1,3-dihydroxy-2-propanone (DHA), which is widely used in cosmetics as
an active ingredient in self-tanning products [
110
]. BNC membranes were impregnated
with different DHA concentrations and afterwards tested on skin. The findings clearly
indicated that after 30 min of application, the skin pigmentation increased with the DHA
concentration
(Figure 5a)
. The pigmentation of skin by DHA is based on a typical Maillard
reaction (viz., reaction of sugars with amino acids) in which DHA reacts with keratin on
the skin surface, forming pigments (melanoidins) bounded to the proteins of the stratum
corneum through lysine chains. This is a reaction limited only to this outer layer of the
skin [
111
]. Noteworthy is also the fact that the BNC–DHA patches did not leave the specific
and typical unpleasant odor of cosmetics containing DHA, which only disappears with
the skin pigmentation effect. As concluded, the method requires improvement, but it
demonstrated the potentiality of BNC as a carrier of DHA with the advantage of having
the ability of being shaped accordingly to the skin surface on which it will be applied.
Therefore, this work opens the possibility of BNC being applied in personalized patches to
mask vitiligo symptoms or in self-tanning masks that may be enriched with other active
ingredients (e.g., anti-aging compounds) [
110
]. In another study, Pacheco et al. [
96
] also
demonstrated the potential of BNC as a sustainable alternative vehicle for cosmetic actives
by investigating the incorporation of two formulations into BNC: the vegetable extract
mask (VEM) consisting of a moisturizing formulation based on Hidroviton
®
(moisturiz-
ing agent) and plant extracts (oat and rosemary) and the other composed by a propolis
extract and poly(propylene glycol) (PEG), which is identified as a propolis extract mask
(PEM). Hidroviton
®
and PEG acted as plasticizers, giving flexibility to the membranes.
SEM micrographs confirmed the incorporation of the cosmetic actives into the BNC net-
work without affecting its morphology. The release assays in phosphate-buffered saline
(PBS) were followed by infrared spectroscopy, showing that the skin-active substances
were quickly released from BNC, which is particularly relevant for a facial mask whose
application is ideal for a short period of time. The effectiveness and acceptance of BNC
membranes were evaluated through sensory tests in volunteers, which revealed high scores
in skin adhesion and mask handling because of the improved malleability given by the
incorporated formulations. The effect of the active substances on skin moisture was also
evaluated, resulting in an increased hydration effect for VEM treatment and a decrease
on skin hydration for PEM formulation after a 20 min use per day during 5 consecutive
days [
96
]. In another recent study, Amorim et al. [
112
] investigated the incorporation of
2% of aqueous propolis extract in BNC membranes simply by immersion in the extract
solution. The propolis extract is commonly used in dermatological preparations for wound
healing and treatment of burns and acne [
113
]. Thus, the aim of the authors was to develop
a material to be used as a facial mask for the treatment and healing of acne-prone skin. The
results showed that with the incorporation of propolis, the BNC crystallinity was reduced,
which directly influences the mechanical properties of the membrane, resulting in a more
elastic material with improved flexibility. This increased flexibility is favorable to a better
Int. J. Mol. Sci. 2021,22, 2836 11 of 25
adhesion and adaptation of a facial mask to skin, demonstrating therefore the potential
applicability of the obtained BNC–propolis membrane for this purpose [112].
Following the trend of BNC skin masks with improved properties and delivery of
active ingredients, researchers developed innovative bioactive BNC membranes by loading
with ionic liquids (ILs) based on phenolic acids [
114
]. Specifically, cholinium cation was
combined with anions derived from ellagic ([Chol]
2
[Ell]), caffeic ([Chol][Caf]), and gallic
acids ([Chol][Gal]), which are natural antioxidants used in cosmetics. The synthesis of
phenolic-based ILs, viz., solvents with a green connotation, aimed to improve the antioxi-
dant activity of these phenolic compounds and at the same time overcome the problems of
the low solubility and bioavailability commonly associated with phenolic compounds. The
bioactive BNC-ILs membranes showed higher re-hydration ability, which is important to
assure suitable hydration for the ILs release. The
in vitro
skin permeation assays (during
5 h), using human epidermal skin in Hanson vertical diffusion cells and PBS buffer at
37
◦
C, revealed that the bioactive BNC-ILs membranes exhibited a slow and sustained
release of the active compounds with a higher flux for BNC-[Chol][Gal] (5.42
µ
g cm
−2
h
−1
)
than for the BNC-[Chol][Caf] (4.93
µ
g cm
−2
h
−1
). The permeation of BNC-[Chol]
2
[Ell]
was not evaluated due to precipitation problems. The BNC-ILs membranes were also
non-cytotoxic and had high antioxidant and anti-inflammatory activities as desired [
114
].
Although the slow release of active compounds is not ideal for a facial mask, it may be
interesting for other dermal care applications. For instance, for membranes loaded with
the anions derived from caffeic acid, BNC-[Chol][Caf] may be used as cellulite treatment
patches, where the continuous and controlled release is required. In the same research
line, another study reported the incorporation of vitamin B-based ILs ([Chol][VITS]) in
BNC membranes [
115
]. Here, the cholinium cation was paired with anions derived from
vitamins of complex B commonly used in skincare formulations, namely, nicotinate (B3),
pantothenate (B5), and pyridoxylate (B6). The resulting membranes (dried state) had re-
duced brittleness due to the plasticizer effect of the ILs, thus avoiding the use of additional
plasticizers. Moreover, BNC-ILs membranes had an improved re-hydration ability (from
2.9 to 4.8-fold) in comparison with BNC. Dissolution tests in PBS showed a fast release of
66% of the incorporated [Chol][VITS] from the corresponding membranes in the first 5 min,
which, as authors discussed, is particularly attractive for short-term masks. Moreover, the
membranes showed to be non-cytotoxic to skin epithelial cells, reinforcing the suitability
of these membranes for skincare applications. Therefore, future studies should address
biological assays regarding the anti-aging properties of ILs such as the collagen production
or the enhancement moisturizing of the stratum corneum skin layer [115].
BNC can also be combined in different ways with bioactive macromolecules. Hyaluronic
acid (HA) is a main constituent of skin epidermis and dermis with recognized properties in
skin moisture maintenance and skin-aging process [
116
], and thus is a crucial ingredient for
cosmetics, including BNC-based masks. Examples are the methods claimed in two patents
to prepare nanocomposite facial masks of BNC and HA [
117
] and sericin–HA [
118
], which
are both prepared by addition of the active molecules to the culture medium. More recently,
an innovative patch for dermo-cosmetic applications (e.g., skin anti-aging) was developed
by Fonseca et al. [
119
] using the non-invasive technology of microneedles (MNs). For
further reading about this technology, a recent review gives a comprehensive overview
about the last developments on polysaccharides and protein-based MNs [
120
]. The system
developed by Fonseca et al. [
119
] combines the features of HA as the dissolvable MN
matrix to enable improvement of the overall appearance of the skin, with the properties of
BNC as the back layer to support the MN array and enable the release of additional active
molecules (Figure 5b,c) [
119
]. The effectiveness of the system was demonstrated with the
incorporation of the bioactive compound rutin, a natural antioxidant, in the BNC back layer
of the MNs system. Results showed that the incorporation in the MNs system did not affect
the antioxidant activity of rutin, and this activity was maintained for 6 months storage
at room temperature. Additionally, the
in vitro
skin assays also unveiled the successful
penetration of these arrays through the skin and the delivery of rutin. The safety and
Int. J. Mol. Sci. 2021,22, 2836 12 of 25
cutaneous compatibility of the MNs HA–BNC were also evaluated and demonstrated in a
preliminary
in vivo
assay performed in human volunteers, in which no significant changes
in barrier function, stratum corneum hydration, nor redness were detected [
119
]. This work
reinforces the potential of BNC as a delivery system in cosmetics.
Figure 5.
Skin coloring 12 h after removal of BNC–DHA patches (concentration of DHA is expressed
in percent), applied for 30 min (a) (reprinted with permission from [110]); Schematic representation
of the HA-(BNC-R) MNs structure (
b
) and functioning of this innovative system: insertion of the
MNs into the skin, dissolution of HA MNs and subsequent release of the bioactive molecule from the
BNC membrane (c) (reprinted with permission from [119]). DHA: 1,3-dihydroxy-2-propanone, HA:
hyaluronic acid, MN: microneedles, R: rutin.
Still in the field of facial masks, a recent innovative approach was used to improve the
delivery of cosmetic actives in a sheet (e.g., carbon cloth, cotton) facial mask by including a
battery [
121
]. The authors used a BNC carbide sheet as the positive electrode, which was
obtained by carbonization of the BNC xerogel at 1200
◦
C under nitrogen atmosphere. The
negative electrode contained at least one of these elements: magnesium, zinc, aluminum,
iron, calcium, lithium, or sodium. Since a facial mask is a disposable product, the aim was
to develop a battery using a material with low environmental impact. The authors claim
that the weak electric current flow generated by the battery allows the active ingredients to
penetrate more effectively into tissues and at the same time enable an electrical stimulation
of face muscles and lymph. The evaluation tests on volunteers confirmed that the battery-
sheet facial mask, impregnated with active ingredients (aqueous solution of carbonic and
L-ascorbic acids), had effectively increased skin moisture content when compared with a
sheet facial mask impregnated with the same active ingredients but without a battery.
The development of BNC-based materials with improved properties whether mechan-
ical, viscoelastic, or swelling behavior that favor their performance in cosmetics has also
been the aim of a few works. An example is the research describing the development of a
Int. J. Mol. Sci. 2021,22, 2836 13 of 25
nanocomposite gel composed of BNC–poly(ethylene glycol diacrylate) (PEGDA) aiming
to obtain an improved soft and flexible gel material either for cosmetic or biomedical
applications [
122
]. A series of compositions were investigated varying the ratio of PEGDA
in the BNC–PEGDA gels. Results showed that the BNC gels with 3% (w/v) and 5% (w/v) of
PEGDA were the ones with higher potential for this type of application, considering their
mechanical and viscoelastic properties. These BNC–PEGDA gels revealed to be harder (de-
formation resistance against compression) but less brittle against tension than a pure BNC
gel. The hardness prevents the change in tactile sensation after repeated finger contacts that
in the case of BNC lead to water expelling and consequent dryness of the gel. Additionally,
results also demonstrated that BNC–PEGDA gels had a similar viscoelastic behavior to that
of BNC gel, with elastic properties being more significant than viscous ones. This indicates
that the low amounts of PEGDA did not significantly affect these properties and these
composite gels could adhere to skin. Although no
in vivo
tests have been performed, this
nanocomposite is believed to have good biocompatibility, considering that both polymers
are biocompatible, what is also favorable for its use in cosmetic or medical fields [
122
], but
further studies should be performed to fully prove the applicability of these BNC–PEGDA
gels as, for instance, a support matrix for delivery of cosmetic actives.
In another study, Chunshom et al. [
123
] successfully prepared a freeze-dried BNC/PVA
nanocomposite in which BNC was used as a reinforcement material to benefit from its
mechanical and thermal properties. The blend of BNC and PVA has been the focus of
several previous works specially for biomedical applications [
124
]. However, in this study,
the innovative approach of producing a freeze-dried BNC/PVA nanocomposite makes
it attractive not only for biomedical but also for personal care cosmetics, as the dried-
state products are nowadays gaining relevance in products such as cleansing masks and
moisturizers [
123
]. One advantage of a dried-state nanocomposite is the reduced pos-
sibility of microbial contamination compared with a never-dried hydrogel counterpart
in which the water in the 3D network can favor it. Moreover, its lightweight is also an
attractive property for industrial application, since it facilitates packaging, transportation
and storage. The results revealed that BNC had a significant influence on the composite
pore size
(10 µm–100 nm),
which is likely contributing to the swelling behavior of the
nanocomposite gel. Several composites with different weight ratios of PVA:BNC were
studied. The composite gel with a ratio of 3:1 (PVA:BNC) presented an excellent swelling
behavior within 30 min either in deionized water, NaCl solution, or PBS [
123
], which is
a promising result for its application as a hydrogel in short-term cosmetic applications,
although further studies need to be conducted.
An essential issue in the manufacturing of cosmetic formulations is to assure their
quality and safety, including stability over time. Regarding BNC cosmetic masks, this
aspect is not often covered in the literature. However, a novel and simple methodological
approach that can be applied in future studies was recently proposed by Perugini et al. [
93
].
The authors investigated the suitability of two non-destructive instrumental techniques,
namely, near-infrared spectroscopy (NIR) and multiple light scattering (MLS), for the
quality assessment of BNC facial masks. In particular, the NIR technique was used to
perform a quality check by evaluating homogeneity and reproducibility among masks of the
same manufacturing batch and from different batches with regard to water and ingredients
distribution. On the other hand, MLS was applied to ascertain the stability of the cosmetic
masks and the possible interaction between the formulation and the BNC matrix, which
are important criteria to establish the shelf-life of these products [
93
]. Another aspect that
is not always included in the development of new BNC facial masks is their acceptance and
effectiveness using
in vivo
assays. In this perspective, Perugini et al. [
97
] recently provided
a scientific protocol to evaluate,
in vivo
, the efficacy of topical treatments with BNC masks
employing a methodology that combines non-invasive skin bioengineering techniques
and statistical methods. The authors evaluated the skin effect of three BNC facial masks
embedded with cosmetic formulations with different beneficial skin actions (anti-aging,
lifting, and cell renewal). The study was performed in volunteers and evaluated a broad
Int. J. Mol. Sci. 2021,22, 2836 14 of 25
range of skin effects, such as moisturization, color, viscoelastic properties, and surface
smoothness of skin, as well as the presence of wrinkles, dermal homogeneity, and stratum
corneum renewal. The results were very satisfactory for all masks, highlighting the potential
of this biopolymer to develop tailored cosmetic masks with high tolerability and efficacy,
validating at the same time a non-invasive approach to evaluate the
in vivo
effectiveness
of BNC facial masks [97].
4.2. Bacterial Nanocellulose as a Support for the Immobilization of Enzymes
In cosmetics, enzymes have been applied for many years, as is the case of some
proteolytic enzymes (bromelain and papain) used for skin peeling and smoothing, or the
superoxide dismutase (SOD) known for its capacity in removing free radicals to prevent
the associated skin aging [
125
]. However, despite their potential as skin-active substances,
enzymes encounter some problems, such as poor stability at room temperature during
prolonged storage or deactivation due to disturbance of the enzyme structure by oils
and surfactants commonly present in cosmetics [
126
]. In this context, the nano-porous
structure and high-surface area of BNC make it an attractive substrate for this application,
offering benefits in terms of the enzyme entrapment and immobilization yield as well
as a better enzyme stabilization [
127
,
128
]. In fact, the use of the BNC matrix has already
been investigated to immobilize enzymes such as laccase, which is an enzyme used in
food and paper industries [
129
,
130
], or the horseradish peroxidase, which is applied in
biotechnological and environmental applications such as the removal of contaminants
from water [
127
]. However, the use of BNC as a support matrix to immobilize enzymes in
cosmetic applications is almost unexplored. In our literature survey, only a recent study
performed by Vasconcelos et al. [
128
] describes the covalent immobilization of papain in
wet BNC membranes previously purified by alkaline treatment and oxidized via NaIO
4
.
On cellulosic supports, among the possible methods, the covalent bonding promotes the
most stable interaction with enzymes, resulting in a possible increase of the activity and
thermostability of the immobilized enzyme [
128
]. However, BNC hydroxyl groups do not
react directly with the amine groups of enzymes [
127
], and thus, a potential solution to
assure a more stable interaction is to chemically modify BNC [
128
]. In this study, covalent
bonding was achieved by imine formation between the protein amino groups and the
BNC carbonyl groups produced by an oxidative treatment. It was observed that the
percentage of recovered activity of the enzyme (relates the activity of immobilized versus
free enzyme) after immobilization on the oxidized membranes was 93.1%, confirming that
the papain remains active after immobilization. Moreover, compared with non-oxidized
BNC, a higher amount of enzyme was immobilized on the oxidized membrane because of a
better chemical interaction with the enzyme, owing to the covalent imine bond established
between the aldehyde groups (oxidation result) of BNC and the amine groups of the
amino acids structure from the enzyme, as evident by infrared spectroscopy [
128
]. The
thermogravimetric analysis showed that the oxidation reaction had lowered (about 54
◦
C),
the onset temperature (T
Onset
) of the BNC membrane, indicating a reduction in its thermal
stability. However, the T
Onset
of 262
◦
C for the oxidized membrane is still sufficiently
high for autoclaving. Finally, the oxidation reaction had also resulted in BNC membranes
with reduced mechanical resistance and higher flexibility, showing a Young’s modulus of
1.7 ±0.18 MPa
(non-oxidized BNC: 5.3
±
0.79 MPa), a tensile strength of 0.3
±
0.02 MPa
(non-oxidized BNC: 1.1
±
0.05 MPa), and an elongation at break of
31.0 ±3.55%
(non-
oxidized BNC: 26.6
±
1.28%). However, from our perspective, the mechanical resistance is
still adequate for cosmetic applications, and the increased flexibility is beneficial for the
adherence to skin. Although this study is not specifically oriented to the cosmetic domain, it
demonstrates the potential of the method to oxidize the wet BNC and to use it as a support
for the immobilization of a cosmetic enzyme, despite further proof-of-concept studies
needed to fully prove it. Therefore, given the growth prospect for the use of enzymes in
cosmetics, owing to their natural origin, consumer acceptance, and high performance, this
Int. J. Mol. Sci. 2021,22, 2836 15 of 25
must be a research field in which BNC may gain relevance in the next years in a viewpoint
of a sustainable and eco-friendly products development.
4.3. Bacterial Nanocellulose as Emulsion Stabilizer
An emulsion consists of a heterogeneous mixture of two immiscible liquids in which
droplets of one are dispersed on the other, being typically stabilized by surfactants [
131
].
Over the last few years, benefiting from advances in nanotechnology and due to irritat-
ing, toxic, and environmental problems generally associated with synthetic surfactants,
Pickering emulsions (viz., emulsions stabilized by solid particles) have gained commer-
cial relevance over the surfactant-stabilized emulsions [
132
]. In particular, the use of
biopolymeric particles instead of their synthetic counterparts is now emerging as a sus-
tainable and green alternative [
133
]. In this vein, Jia et al. [
132
] have produced individual-
ized cellulose nanofibers by the oxidation of dispersed BNC using a mixed system with
2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO), NaBr, and NaClO, aiming to get a safe,
biodegradable, and sustainable solid emulsion stabilizer. The prepared TEMPO-oxidized
BNC nanofibrils (TOBNC) showed reduced fibril size and strengthened wettability (ac-
cessed by contact angle measurements). Both features were influenced by the degree of
oxidation that had increased with the NaClO dosage. The suitable counterbalance be-
tween wettability and fibril size seems essential to achieve emulsion stability. Taking in
account the results, the authors considered that the TOBNC, prepared with 2 mmol g
–1
of NaClO, was the most effective emulsion stabilizer against creaming and coalescence,
showing a better stabilizing performance than BNC in oil–water type Pickering emulsions
(prepared using liquid paraffin) stored at room temperature over 8 months [
132
]. This
is likely due to the fact that in TOBNC, the wettability did not change much, but the
fibril size was significantly reduced, resulting then in more fibrils covering the surface
of droplets, thus creating a rigid barrier. Given the promising results, the authors fore-
see the use of this TOBNC as an emulsion stabilizer in topical, pharmaceutical, or food
formulations [
132
]. More recently, another study proposed a one-pot method to produce
TOBNC using TEMPO immobilized on silica beads and performing a simple filtration
after the oxidation reaction, instead of the centrifugation used in the previous method [
12
].
This approach ensured the total removal of reactants, which is particularly important for
skin care products. Moreover, it was also observed that the obtained TOBNC maintained
the nanofibrous structure of BNC and the water-absorbing capacity. When combined
with water and an oil–water emulsion, it showed the capacity to block the adhesion of
particulate matter on skin using a porcine skin model. By the simplicity, efficiency, and
easy scale-up, this one-pot method was shown to be a good alternative to produce TOBNC
for skincare applications at the industrial level [
12
]. Envisioning the industrial application
of BNC in cosmetic and food sectors, Martins et al. [
134
] developed a dry formulation of
BNC and carboxymethyl cellulose (CMC) using the spray-drying technique. This novel
dry formulation of BNC:CMC demonstrated to be fully dispersible in water within a few
minutes using low-energy stirring, which might have a relevant impact in production
costs. When compared with commercially available dry celluloses, the dry BNC:CMC
formulation had a better stabilizing performance, showing at a concentration of 0.50% the
capability of stabilizing low density oil-in-water emulsions against coalescence or creaming
for up to 90 days
(Figure 6),
which is a promising result for its applicability in cosmetics as
a Pickering emulsions stabilizer.
Int. J. Mol. Sci. 2021,22, 2836 16 of 25
Figure 6.
Digital photographs and optical micrographs (10
×
magnification) of 10% isohexadecane-in-water emulsions
prepared with different concentrations of BNC:CMC dry formulation (0.10%, 0.25%, and 0.50%) and with 0.50% CMC,
taken 1 day after preparation and 30 and 90 days after storage at room temperature. Black scale bars correspond to 100
µ
m
(reprinted with permission from [134]). CMC: carboxymethyl cellulose.
4.4. Bacterial Nanocellulose as an Alternative to Microplastics in Cosmetics
The environmental impact of microplastics (or plastic microbeads) (<5 mm) is nowa-
days of major concern, being the cosmetics and personal care products, one of the relevant
sources of primary microplastics (viz., manufactured microplastics) [
135
]. A broad range of
daily-use personal care and cosmetic products (e.g., shampoos, toothpastes, facial scrubs,
and soaps) contains microplastics that have either a decorative or exfoliating function.
Polyethylene is the main polymer used in microplastics production, but other synthetic
polymers are also used (e.g., polypropylene, nylon, polytetrafluoroethylene) [
136
,
137
]. The
most claimed problem of these microplastics is related with their recalcitrancy and with
the fact that they ultimately end up in the environment, and in the marine environment
in particular, and may build up in the food chain, ultimately causing adverse effects on
human health [
138
]. Thus, as in other fields, over the last few years, the awareness of
consumers and the environmental policies are driving the research and industrial players
to find environmentally friendly alternatives to microplastics. With this regard, Joon-
won [
139
] developed a patented method to prepare a BNC powder with properties that
turn it into a potential and appealing alternative to replace microplastics in cosmetic prod-
ucts (e.g., scrubs, lotions). The method includes the production of BNC sheets (thickness
of
7 to 15 mm
) in culture medium based on coconut extract and, for example, fruit juice,
followed by treatment with an alkaline solution to remove oils and impurities, a step of
air drying (60–99
◦
C), and finally, the pulverization of BNC. The authors claim that the
resulting BNC powder, with a minimum purity of 95%, should have a size between 0.1
and 1 mm and a maximum residual water content of 15%. The reduced water content is
essential for the BNC powder to have little or no water absorption ability and therefore
preserve the shape even in a liquid phase, which is important to maintain granularity.
This will be related to the complete collapse of the 3D structure of BNC during the drying
process, which prevents BNC from rehydrating. The behavior of the obtained BNC powder
was tested in purified water and compared with a conventional dried BNC powder, which
confirmed the water non-absorption property of this novel BNC powder [
139
]. Apparently,
no stability tests were performed under this study, but the properties of the BNC powder
look promising for it to be applied as a substitute of microplastics.
Int. J. Mol. Sci. 2021,22, 2836 17 of 25
Table 1. Summary of the most recent and relevant research articles and patents reporting the application of BNC for cosmetic purposes.
Active Substance/Co-Former Main Outcome Application/Potential Application Reference Year
BNC AS CARRIER OF SKIN ACTIVE SUBSTANCES
Bamboo extract added to BNC fermentation broth BNC membrane with superior performance in terms of adhesion,
skin elasticity, softness, and moisturizing effect Sheet facial mask [98]2015
Moringa oleifera leaves powder added to BNC
fermentation broth; Moringa oleifera fermentation broth
BNC membrane with embedded Moringa oleifera leaves fermentation
broth with a better hydration effect than a normal membrane cloth Sheet facial mask [101]2019
Sphaerotilus natans-derived microtubes Multilayer structure with an improved transdermal delivery
of water and lipid-soluble active substances Sheet facial mask [103]2018
Soybean molasses added to BNC fermentation broth BNC membranes with high-water retention rate (98.35%) and good
moisturizing effect Sheet facial mask [104]2018
Milk by-products and tea polyphenols added to BNC
fermentation broth
Increased BNC production yield; BNC membrane with antioxidant and
whitening properties Sheet facial mask [105]2019
Tropical fruit by-products added to BNC fermentation
broth BNC membrane with high-water activity and incorporating ascorbic
acid from the fruit by-products Sheet facial mask [106]2019
Caffein BNC—caffein topical delivery system with lower permeation rates of
caffein than conventional formulations (aqueous solution and gel);
reproducible, predictable and extended release of caffein over time Patches for cellulite treatment [107]2014
PEO-b-PCL nanoparticles encapsulating retinol BNC-based delivery system of hydrophobic molecules; slow release of
retinol from nanoparticles; retinol precipitation and retention in the
BNC gel (further studies are needed) Hydrogel for skin care [108]2015
Bee venom BNC membrane with potential anti-inflammatory and
anti-allergic properties Sheet facial mask [109]2017
DHA BNC–DHA patch applied for 30 min was effective in conferring a skin
natural tan effect Sheet facial mask [110]2018
Hidroviton®and plant extracts/PEG
and propolis extract
Effectiveness of BNC facial masks as delivery system
of active substances Sheet facial mask [96]2018
Propolis extract Improved flexibility and malleability, higher porosity of
BNC membrane Sheet facial mask [112]2020
-BNC carbide used as the positive electrode of a battery included in the
facial mask; improved penetration of active ingredients into tissues Sheet facial mask with a battery [121]2019
Cholinium-based ILs paired with anions derived from
phenolic acids (caffeic, ellagic and gallic)
BNC membrane with increased re-hydration ability; slow and
sustained release of active compounds; antioxidant and
anti-inflammatory activities Sheet facial mask [114]2019
Cholinium-based ILs paired with vitamins B anions BNC membrane with reduced brittleness, increased re-hydration
ability; fast release of active compounds Sheet facial mask [115]2020
Int. J. Mol. Sci. 2021,22, 2836 18 of 25
Table 1. Cont.
Active Substance/Co-Former Main Outcome Application/Potential Application Reference Year
HA Method to produce BNC–HA composite for facial masks preparation Sheet facial mask [117]2017
Sericin–HA Method to produce BNC–sericin–HA composite for facial
masks preparation Sheet facial mask [118]2018
HA–Rutin
Increased mechanical resistance of HA-MNs. Effective BNC controlled
release of rutin. Maintenance of rutin antioxidant activity upon MNs
system storage at room temperature for 6 weeks MNs system for skin care [119]2021
PEGDA BNC-3% and 5% PEGDA composites harder but less brittle than BNC
gel; similar viscoelastic behavior to that of BNC gel Hydrogel for facial masks [122]2017
PVA BNC–PVA composite in a freeze-dried state; reduced possibility of
contamination due to the freeze-dried nature of the BNC–PVA
composite; lightweight and good swelling rate within 30 min
Freeze-dried additive for facial masks
[123]2018
Active cosmetic formulations:
anti-aging/lifting/purifying and regenerative
Non-invasive protocol for in vivo evaluation of the effectiveness
and acceptance of BNC facial masks as delivery system of
active substances Sheet facial mask [97]2020
BNC AS A SUPPORT FOR THE IMMOBILIZATION OF ENZYMES
Papain Oxidized BNC membrane with covalently immobilized papain; a
higher amount of enzyme immobilized than in non-oxidized
membrane; 93.1% recovered activity of the enzyme after immobilization
Enzyme-based skin care [128]2020
BNC AS EMULSION STABILIZER
-TEMPO-oxidized BNC nanofibrils with reduced size; better
stabilization of oil–water emulsions interface than with BNC
(over 8 months) Emulsion stabilizer [132]2016
-
Simple and easy scalable one-pot method to obtain TEMPO-oxidized
BNC nanofibrils; the procedure assures the total removal of reactants;
maintenance of the BNC nanofibrous structure and
water-absorbing capacity.
Emulsion stabilizer [12]2019
Carboxymethyl cellulose (CMC)
Fully dispersible BNC:CMC dry formulation; better stabilizing effect of
low oil-in-water emulsions than other dry commercial available
celluloses; stabilization effect for up 90 days Emulsion stabilizer [134]2020
BNC AS AN ALTERNATIVE TO MICROPLASTICS
-BNC powder with 95% minimum purity, a size between 0.1 and 1 mm
and a maximum residual water content of 15%; shape preservation and
no-water absorption ability in liquid phase Alternative to microplastics [139]2019
Int. J. Mol. Sci. 2021,22, 2836 19 of 25
5. Conclusions and Future Perspectives
Over the last years, with the growing demand for bio-based cosmetics, the extraordi-
nary properties of bacterial nanocellulose, along with its biodegradable nature, renewable
character, and unique properties, have drawn an increasing attention by both academia
and the cosmetic industry. This review makes evident that this remarkable biopolymer
is a feasible alternative to petroleum-based counterparts for a broad range of cosmetic
applications. Benefiting from its ultrafine nano-porous structure, BNC has been extensively
explored as a carrier of skin-active substances, with several studies demonstrating its
potential and effectiveness for the loading and releasing of these substances, especially
applied as sheet facial masks. In this field, despite the advances and the well accepted
products already commercialized by top cosmetic companies, there are still some chal-
lenges needing to be deeply studied, namely the design of delivery systems of hydrophobic
molecules, which is not straightforward owing to BNC hydrophilicity. As demonstrated
in this review, the blend of amphiphilic polymers is a promising strategy. In cosmetics,
as in drug-delivery, it is desirable to have a precise delivery system tailored according
to the intended application, to reduce the loading dose of the active compound and, at
same time, guarantee the effectiveness of the treatment. Therefore, this is an aspect that
should continue to deserve attention in further investigations by including
in vitro
and/or
in vivo
assays and searching for optimized BNC-based delivery systems with efficient
releasing rates that can be modulated by physical treatments, chemical modifications, or
even association with other polymeric matrices. In this topic, the design of smart hydrogels,
namely temperature-responsive hydrogels, are gaining interest in the skin care field as
delivery systems with controlled release based on the body temperature stimulus.
BNC has also been successfully explored, although in few studies, as a sustainable
alternative to synthetic surfactants used to stabilize Pickering emulsions. However, studies
demonstrating the performance of these BNC-based emulsion stabilizers in realistic cos-
metic formulations are still missing. Therefore, considering that Pickering emulsions are
an emerging choice in topical formulations as a vehicle of skin-active substances, further
investigations should include comparative studies with different cosmetic formulations.
Moreover, the influence of the BNC-based particles in the release profile of the active
substances from the emulsion and their penetration into skin should also be addressed. In
this context, novel combinations of BNC nanofibrils with other biopolymers or the use of
BNC nanofibrils with different sizes or shapes may be investigated, aiming at obtaining
emulsions with improved properties, regarding stability, cosmetic active release, and skin
penetration, similar to what has been done with other biopolymers [133].
Over this review, it is also noteworthy that BNC also demonstrated to be a viable
alternative to other relevant cosmetic applications, such as support matrix for enzymes
immobilization or as alternative to microplastics, although these two research topics are
only in a very early stage. Regarding enzymes immobilization, additional studies are
needed with regard to the stability of the enzyme activity upon storage, but the technology
based on the immobilization of enzymes onto oxidized BNC seems promising and may be
an attractive form of using enzyme-based cosmetics. In this way, the low effectiveness and
short shelf-life, many times associated with conventional formulations, may be overcome.
Future research should also follow the consumers demand for multifunctional products
and consider the development of effective systems with multiple immobilized enzymes.
Cellulose substrates are gaining interest as an environmental-friendly alternative to
microplastics [
140
]. In this regard, a recent patent claims one method to produce BNC
powder intended to be used as microbeads in cosmetic formulations, showing thereby
that BNC may be a promising substrate for this purpose. However, this possibility is
still underexplored, and progress is expected for the next years. In this field, one of the
main challenges will be to develop simple, scalable, and cost-effective processes capable of
producing microbeads with constant and uniform diameters, allowing a close control of the
size, shape, and hardness of the microbeads to meet the desired final product properties.
Int. J. Mol. Sci. 2021,22, 2836 20 of 25
With cosmetic companies setting strict sustainability objectives, the investment in
green-tech solutions has been extended to the entire product life cycle, including packaging.
In this context, BNC may also draw attention in the future as an environmentally friendly
material for cosmetic packaging, as is already the case in the food sector. In our literature
survey, no studies were found that focused on this application. Hence, this should be a
research topic that will probably evolve in the future, especially in the segment of active
packaging. Herein, the development of BNC composites with compounds that may impart
BNC with properties highly valued in cosmetic packaging, such as the O
2
scavenging,
antimicrobial, or antioxidant activities, will be particularly attractive.
Although BNC products are already commercialized by a small number of companies,
there are still some economic constraints that are hampering its broad application and
should continue to drive future research. However, in this review, it is clear that many
studies are already considering the reduction of costs and the up-scaling to mass production
either by using agro-industrial by-products as feedstocks or by developing simple and easy
processes to be implemented at a large scale. The progress in the fermentation systems
using bioreactors and in the genetic engineering of BNC-producing bacteria should also be
crucial to achieve high production yields and cost-effective BNC, which will make it more
competitive in the cosmetic market and thereby extend its use. However, the versatility of
BNC is unquestionable, and, for the coming years, it is expected that this high-performance
biopolymer will gain even more relevance in the highly innovative cosmetic industry,
especially in green cosmetics.
Author Contributions:
Conceptualization, C.S.R.F. and C.V.; writing—original draft preparation,
T.A.; writing—review and editing, T.A., A.J.D.S., C.V. and C.S.R.F.; supervision, C.V. and C.S.R.F.;
project administration, C.S.R.F.; funding acquisition, C.S.R.F. and A.J.D.S. All authors have read and
agreed to the published version of the manuscript.
Funding:
This work was developed within the scope of the project CICECO—Aveiro Institute of
Materials (UIDB/50011/2020 & UIDP/50011/2020) financed by national funds through the Por-
tuguese Foundation for Science and Technology (FCT)/MCTES, and project Inpactus—Innovative
products and technologies from eucalyptus, Project No. 21874 funded by Portugal 2020 through
European Regional Development Fund (ERDF) in the frame of COMPETE 2020 No 246/AXIS II/2017.
FCT is also acknowledged for the research contract under Scientific Employment Stimulus to C.V.
(CEECIND/00263/2018).
Conflicts of Interest: The authors declare no conflict of interest.
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