Sources, Stability, Encapsulation and Application of Natural Pigments in Foods

Food Reviews International
ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/lfri20
Sources, stability, encapsulation and application of

natural pigments in foods
Slaven Jurić , Marina Jurić , Żaneta Król-Kilińska , Kristina Vlahoviček-

Kahlina , Marko Vinceković , Verica Dragović-Uzelac & Francesco Donsì
To cite this article: Slaven Jurić , Marina Jurić , Żaneta Król-Kilińska , Kristina Vlahoviček-
Kahlina , Marko Vinceković , Verica Dragović-Uzelac & Francesco Donsì (2020): Sources, stability,
encapsulation and application of natural pigments in foods, Food Reviews International, DOI:
10.1080/87559129.2020.1837862
To link to this article: https://doi.org/10.1080/87559129.2020.1837862
Published online: 28 Oct 2020.
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FOOD REVIEWS INTERNATIONAL
https://doi.org/10.1080/87559129.2020.1837862
REVIEW
Sources, stability, encapsulation and application of natural

pigments in foods
Slaven Jurića, Marina Jurićb, Żaneta Król-Kilińskac, Kristina Vlahoviček-Kahlinaa,
Marko Vinceković a, Verica Dragović-Uzelacd, and Francesco Donsìe
a
Faculty of Agriculture, Department of Chemistry, University of Zagreb, Zagreb, Croatia; bFaculty of Pharmacy and
Biochemistry, Department of Pharmacognosy, University of Zagreb, Zagreb, Croatia; cDepartment of Functional Food
Products Development, Wroclaw University of Environmental and Life Sciences, Wroclaw, Poland; dFaculty of Food
Technology and Biotechnology, Department of Food Engineering, University of Zagreb, Zagreb, Croatia; eDepartment
of Industrial Engineering, University of Salerno, Fisciano, Italy
ABSTRACT Keywords
In recent years, the replacement of synthetic colorants with natural ones has Anthocyanins; betalains;
attracted increasing consumers’ and market interest. Natural colorants include chlorophylls; carotenoids;
different groups of pigments, many of which possess also pronounced biolo stabilization; encapsulation
gical potential. This review addresses the main issues related to the use of
natural pigments in foods, starting from the sources available in nature, their
chemical properties, stabilization processes, and applications in real foods, as
discussed in the scientific literature reported in the main databases relevant to
this topic (Scopus, Web of Science, PubMed, ScienceDirect, and Google
Scholar). Notably, several natural pigments are available to cover different
needs in terms of hues and intensities, and whose use is permitted in foods
by the main regulatory agencies. However, their use is still frequently limited
by their higher price and lower stability than synthetic counterparts. This
review discusses in detail the main sources for natural pigments, focusing on
the recent trends towards those more economically favorable, such as micro
bial sources and agro-industrial residues. It also examines the most suitable
stabilization systems to protect the highly reactive and unstable molecules of
natural pigments from negative physical and chemical changes, as well as to
minimize the interactions with food systems.
Introduction
Color is one of the most important sensory attributes of food, as it contributes to providing the
consumers with a first impression of the food quality, but it is also associated with flavor, safety, and
nutritional value.[1,2] Therefore, it significantly affects the consumers’ acceptance and plays a vital role
in the market success of a product.
Pigments are the natural coloring matter, abundantly present in both animal and plant tissue. They
can also be found in leaves, fruits, vegetables, and flowers, as well as in skin, eyes, and other animal
structures and bacteria and fungi. The use of natural pigments (NPs) in food production is becoming
very popular mainly because the consumers are concerned about the health safety of synthetic food
colors. Furthermore, since some NPs might also present significant health benefits,[3] their use in food
has gathered increasing attention in the last years. Unfortunately, the use of NPs is limited by their
intrinsic molecular instability, as well as by their higher costs in comparison with synthetic pigments,
need for higher concentrations to achieve equivalent color intensity, as well as the narrower range of
hues.[4] Food products are naturally colored by four main groups of NPs: the red-blue-purple
anthocyanins, the red betalains, the green chlorophylls, and the yellow-orange-red carotenoids.[,56]
CONTACT Francesco Donsì [email protected] Department of Industrial Engineering, University of Salerno, Fisciano, Italy
© 2020 Taylor & Francis
2 S. JURIĆ ET AL.
The stability of NPs depends mainly on a combination of environmental and chemical factors such
as pH, presence of metal ions, exposure to light, high temperatures, and oxygen, but it is also
significantly influenced by the product enzymatic activities.[7] Because of the low stability in the
environmental conditions during food processing and storage, the incorporation of NPs into food
products is extremely challenging.[8]
Despite encapsulation techniques represent an effective way to protect food pigments against
deterioration,[9] significant challenges remain concerning the incorporation of encapsulated NPs
into food products.
This review article focuses on conventional and new potential sources of NPs, highlighting the
importance of their health-promoting properties, and their potential application in foods.
Furthermore, since NPs are relatively unstable molecules, this review also addresses their stabilization
and protection through encapsulation, discussing the potential applications of encapsulated NPs and
the main issues associated with their implementation in real food products. Finally, an overview of the
main applications of encapsulated pigments will be provided, with a special focus on the production of
functional foods.
Methods
The review was based on the revision of the literature regarding: a) sources of NPs available in nature,
b) NPs chemical properties and stability, c) current, novel, and prospect stabilization processes of NPs,
and d) applications of NPs in real foods. The databases for the search included: Scopus, Web of
Science, PubMed, ScienceDirect, and Google Scholar. The selected timeline was from the past 10 years,
from 2010 to 2020. The search strategy was directed towards NPs sources, stability, stabilization, and
application in foods. Approximately 7000 research papers were initially screened. Main search terms
included keywords and/or their combination: natural pigments, natural colorants, microbial pig
ments, classification, hydrophilic natural pigments, hydrophobic natural pigments, chemical struc
ture, legal aspects, regulatory approval, E-numbers, extraction, functional foods, food industry,
sources, potential sources of natural pigments, plant-, animal-, microbial- and mineral-derived
pigments, agro-industrial residue, anthocyanins, betalains, chlorophylls, carotenoids, health benefits,
health-beneficial properties, stabilization, encapsulation, wall materials, novel, application, inclusion,
implementation, sensory attributes, real foods, real food systems. The literature search included all
access types of full-text articles published in indexed peer-reviewed journals in the English language,
without restrictions about the subject area or the region of research (a global search was conducted).
The types of searched publications included final publications of research articles, articles in press,
review papers, books, and book chapters. We have specifically included information from official
websites from the main, world-renowned regulatory agencies: FAO/WHO Expert Committee,
European Food Safety Authority, The United States Food and Drug Administration. No author
names were excluded in the search. Only online articles that were not present in indexed journals,
articles published in non-English languages, short surveys, conference proceedings, conference
reviews, notes, and letters were excluded.
Classification, stability, and sources of NPs

NPs comprise a wide range of structurally diverse compounds, which, due to the polarity of the
molecule and the chemical and physical properties, can be divided into two main groups: water-
soluble (hydrophilic molecules) and lipid-soluble (hydrophobic molecules) pigments. Red-blue-
purple anthocyanins and red betalains are generally water-soluble, whereas green chlorophylls and
yellow-orange-red carotenoids are lipid-soluble.[4] Table 1 presents this classification in detail, identi
fying the main constituents for each sub-class of NPs, and indicating the main sources, the structural,
chemical and functional characteristics, as well as the extraction procedure for their isolation from
different sources.
FOOD REVIEWS INTERNATIONAL 3
Table 1. Classification of NPs, main features, sources, and extraction processes.

Chemical
Pigment/Subgroup structure Solubility/Stability Color Sources Extraction
Anthocyanins, -flavylium substituents[11] -water soluble[11] from dark blue to
located in (2-phenyl -two aromatic -the anthocyanin extracts or red[6]
vacuoles[10] benzo rings (rings A and isolated anthocyanins are
-aglycones pyrylium) B) linked by highly unstable, susceptible to
(anthocyanidins): structures a three-carbon temperature, light,
petunidin, with varying heterocyclic ring solvent,[12–14] pH, oxygen,
pelargonidin, hydroxyl and (ring C) that presence of proteins, metallic
delphinidin, methoxyl contains ions, enzymes[6]
cyanidin, oxygen[11]
peonidin,
malvidin
-glycosides[10,11]
Plum (Prunus subg. mixtures of
Prunus), different water with
berries ethanol,
(strawberry, methanol, or
blackberry, acetone[19,20]
blueberry,
cranberry),
purple grapes
(Vitis vinifera L.),
red cabbage
(Brassica oleracea
L. var. capitata
f. rubra)[15–18]
Betalains, located in -nitrogen- -water soluble[21] yellow to Ullucus precooled water
vacuoles[21] containing, -heat-sensitive violet[21] Tuberosus Caldas,[29] or aqueous
[21]
betacyanins betalamic molecules Basella rubra L.,[30] cactus pear methanol [21]
structural types: acid -stable between (Opuntia ficus-indica (L.) Mill.
betanin condenses pH 3 and 7[22] and Oxalis stricta L.),[31–35] red-
gomphrenin with imino -yellow purple pitaya (Hylocereus
amaranthin compounds betaxanthins-less polyrhizus (Weber) Britton &
ougainvillein (cyclo-DOPA stable than their Rose),[32,35–41] Amaranthus
-betaxanthins: /its glucosyl red L. species[22,41–44]
indicaxanthin derivates) or counterparts[23,24]
vulgaxanthin I[21] amino acids/ -betacyanins
derivates[21] exhibit more
peroxidase
activity than
betaxanthins[25]
-metal cations
(Fe2+, Fe3+, Sn2+,
Al3+, Cr3+, Cu2+)
accelerate
betanin
degradation[26–28]
Chlorophylls -porphyrins- -fat soluble[47] from blue- green (b)[46] -photosynthetic
-6 types identified macrocyclic -chemically green organisms
(a, b, c, d, e, f)[45] tetrapyrrole unstable (a) to (plants,
-major types: pigments[35] molecule[48] yellow- eukaryotic and
chlorophyll a with -degradation by blue-green
chlorophyll coordinated light, heat, algae) [53]
b[45,46] magnesium oxygen, acid,
in the center enzymes[49]
- hydrophobic -chlorophyll a is
segment (the less thermally
phytol) and stable than the
macrocycle chlorophyll
(hydrophilic b[50–52]
part)[24]
organic solvents [54]
(Continued)
4 S. JURIĆ ET AL.
Table 1. (Continued).
Chemical
Pigment/Subgroup structure Solubility/Stability Color Sources Extraction
Carotenoids -C40 tetraterpenes/ -lipid soluble[56] yellow, orange
-more than 700 tetraterpenoids, -sensitive in the presence of the and red[56]
identified formed from reactive oxygen species, light,
carotenoids[55] eight C5 acidic pH, high temperature[58]
- hydrocarbon isoprenoid
carotenes: units[35]
lycopene, β-
carotene, β-
cryptoxanthin
-xanthophyll
(oxygenated
derivatives):
astaxanthin,
zeaxanthin,
lutein[56,57]
-higher plants,
some
animals,[56] microorganisms,[59,60] green vegetables, tomato, orange fruits and vegetables, corn, salmon, crustaceans, red
peppers,[35,58,61,62] seaweeds[60]
ethyl lactate, ethyl acetate, 70% ethanol, hexane,[63] cold acetone,[62] dichloromethane, dimethyl and diethyl ether, octane,
limonene, and biphasic mixtures of non-polar solvents[60]
Anthocyanins, water-soluble pigments, typically located in plant cell vacuoles,[10] belong to the
group of flavonoids, which is widespread in nature, with 6400 molecules identified so far. More than
540 anthocyanins have been identified in different plant sources,[64] and they include unstable
aglycones (anthocyanidins) and more stable glycosides. 31 monomeric anthocyanidins have been
discovered, but more than 90% of those present in nature include petunidin, pelargonidin, delphini
din, cyanidin, peonidin, and malvidin.[4,65] The color of foods rich in anthocyanin pigments can vary
from dark blue (blueberry, Vaccinium sp. L) to purple (eggplant, Solanum melongena L.) and red
(strawberry, Fragaria × ananassa).[4]
Anthocyanins can be found in many sources such as roots, flowers, grain, nuts, but the most
important points of supply are vegetables and fruits.[15,16,66] Plums (Prunus subg. Prunus), different
berries (strawberry, blackberry, blueberry, cranberry), purple grapes (Vitis vinifera L.), red cabbage
(Brassica oleracea L. var. capitata f. rubra) are examples of products rich in these pigments.[15-18,67]
However, it must be remarked that the pigment concentration and composition in plant tissues depend
on several factors, such as climate,[68,69] variety,[70-76] cultivation,[74] and storage conditions.[77]
Generally, the color of anthocyanins depends on the conjugated double bonds, whereas the stability
and reactivity are strongly influenced by the number and position of hydroxyl groups, degree of
methylation -OH groups, level of glycosylation and pH level.[4] The anthocyanin’s colors are pH-
dependent. Depending on the pH and ionic strength of the aqueous environment, anthocyanins undergo
structural alterations. At pH 3 or lower, they exist as flavylium cations that are orange or red.[78]
Furthermore, stability and color of the pigments are influenced by the β-G anthocyanase (antho
cyanin-β-glucosidase), which may partially hydrolyze anthocyanins, resulting in a reduction of hue
intensity. About 80 different sugar structures may be involved in the glycosylation of flavonoid
molecules and also anthocyanins, but glucose is the most common. The main anthocyanin sugar
residue is glucose, and because of the breaking of the bond between the sugar residue and the
anthocyanidin, decolorization occurs and spontaneously degrades into colorless compounds.[79]
Since these enzymes are activated during the mechanical damage of plant cells, it is important to
inactivate them during food processing, to avoid any further degradation of the pigments. Therefore,
emerging nonthermal or mild technologies, capable of inducing the inactivation of enzymes without
the application of high temperatures for prolonged times have gathered increasing interest to ensure
the preservation of the pigments from enzymatic degradation and thermal damage. High hydrostatic
pressure,[80] high-pressure carbon dioxide,[81] pulsed electric field,[82] ohmic heating,[83] or cold
plasma[84] have all been reported to be valid alternatives to conventional thermal technologies for
enzyme inactivation.
Anthocyanins from natural sources are permitted within Europe (E163), Japan, the United States,
and many other countries to be used as food colorants in foods and beverages.[85] Joint FAO/WHO
Expert Committee on Food Additives (JECFA) concluded that anthocyanin-containing extracts had
very low toxicity with regards to the toxicological studies on mutagenicity, reproductive toxicity,
teratogenicity, acute, and short-term toxicity.[86,87] Anthocyanin colors may be used as quantum satis
in food products except in breakfast cereals where a maximum amount of 200 mg/l or mg/kg as
appropriate, (Regulation (EC) No 231/2012).[88]
Betalains are another important group of natural, water-soluble, nitrogen-containing pigments,
which are located in the cell vacuoles. About 75 different betalains have been found in nature.
Considering their chemical structure, betalains can be classified as betaxanthins (yellow-orange
color) and betacyanins (red to red-violet color).[6] Not all sources of betalains present the same
biological and antioxidant activity, mainly because of the number and position of hydroxyl/imino
groups and the aglycones glycosylation in the structure of betalains.[89]
These plant pigments are found exclusively in families of the Caryophyllales order,[90] with
exceptions in some higher-order fungi, where they replace anthocyanin pigments.[91] Examples of
the betacyanin pigments are beetroot (Beta vulgaris),[92] Amaranthus species (Amaranthus L. spp.),[44]
the prickly pear (Opuntia Mill. spp.), red pitahaya or red dragon fruit (Hylocereus polyrhizus (Weber)
Britton & Rose).[93] Several factors may affect the stability of betalains, which have to be taken into
account to ensure their properties. Stability is mainly influenced by the exposure to high temperature,
pH lower than 3 or higher than 7, light, oxygen, and in the presence of high water activity.[94-96]
Furthermore, the stability of betalains is strongly influenced by their chemical structure. The
intensity and saturation of the red color of the betalain sources depend on the ratio of betacyanins
and betaxanthins. Because of their structural groups, betacyanins exhibit better stability at high
temperature and acidic pH and are less prone to oxidation than betaxanthins, but betaxanthins
show higher stability at pH 7, and against hydrolytic enzymes. The higher stability of betacyanins
than betaxanthins may be due to the fact that some of them have a glycosylated structure, which has
a high oxidation-reduction potential.[23,24,96]
Although anthocyanins are the more widespread and more used NPs covering the red-purple color
range, betalains are more stable at high temperatures and exhibit broad pH stability, which makes
them suitable for low-acid foods, where coloring with anthocyanins usually is not possible.[56,97]
Beetroot red (E162) is a natural color authorized as a food additive in the European Union in
accordance with Annex II to Regulation (EC) No 1333/2008. Beetroot red is a natural color obtained
from the roots of natural strains of red beets (Beta vulgaris L. var. ruba). In the published literature, it is
also known as Beet Red, Betanin, or INS No. 162. Regarding the E162 natural color, EFSA concludes
that, because it is a natural dietary constituent having a long history of food consumption, it is safe to
use. Betalain exposure resulting from the use of beetroot red (E162) as a food additive is in the same
range as the exposure to the betalain from the regular diet. Currently, evaluation only refers to the
beetroot red prepared by pressing crushed beet as pressed juice or by aqueous extraction, all per the
definition of the Commission Regulation (EU) No 231/2012 and not to preparations manufactured by
solvent extraction with methanol or ethanol. The Panel concluded that, at the reported levels of use,
beetroot red (E162) does not represent a safety concern as a food additive.[98] US Food and Drug
Administration (US FDA) authorizes the use of dehydrated beets which may be safe as food colors at
concentrations consistent with a good manufacturing practice. US FDA notes that it may not be used
to color foods for which standards of identity have been promulgated under section 401 of the act
unless the use of added color is authorized by such standards.
Six different pigments of chlorophyll have been identified (a, b, c, d, e, and f),[45] with the major
types being chlorophyll a (blue-green) and chlorophyll b (yellow-green).[46] Chlorophyll molecules
generally consist of a hydrophobic segment, the phytol, and a macrocycle, the hydrophilic part.[6]
6 S. JURIĆ ET AL.
Chlorophylls are chemically unstable[48] and fat-soluble molecules,[47] which can also be degraded by
an excess of light or heat, presence of oxygen, under acidic environment, and by enzymes.[49]
Chlorophylls are significantly affected by light, and even the storage at low-temperatures might result
in their degradation.[99]
Furthermore, it has been found that chlorophyll a is less thermally stable than chlorophyll b.[50-52]
The main source of chlorophyll pigments is represented by green leafy vegetables.[100] Since chlor
ophyll is important in photosynthesis, chlorophyll-derived pigments can be found in photosynthetic
organisms (e.g. plants, eukaryotic and blue-green algae).[53]
When chlorophylls are exposed to prolonged heating, they degrade into olive-brown pheophytin,
via the replacement of Mg2+ ion with H+ in the center of the porphyrin ring of the chlorophyll
molecule. Pheophytin further degrades to pyropheophytin via decarboxylation. It should be noted that
when exposed to heat, chlorophyll a is prone to pheophytin formation than chlorophyll b, whose
higher stability may be attributed to the electron-withdrawing effect of its C-3 formyl group.
Furthermore, when chlorophylls are completely converted to pheophytins, they may degrade further
to pheophorbides. Pheophorbides may eventually degrade to some colorless compounds through
different pathways, affecting the lightness of the product.[101]
With regards to the legal aspects of the green natural colorants, European current legislation
(Regulation (EC) No 1333/2008 and its amendments, European Regulation 1169/2011) permits the
use of two natural green colorants, E140 and E141, in foods.[102] E140 comprises direct chlorophyll
derivatives, obtained through the extraction with organic solvents from natural edible plant materials.
Due to the extraction processes, the final product may contain also lipids, other pigments, and waxes,
whose presence may significantly affect pigment solubility. The E140i colorant is the direct lipid-
soluble chlorophyll derivative obtained from plant extraction, whereas the E140ii colorant (chloro
phyllins) is water-soluble, and is produced through extract saponification.[102] Because of the instabil
ity of these natural colorants, which undergo significant color change (from green to brown), the food
industry generally prefers the use of E141, which is classified in the lipid-soluble E141i, consisting of
copper chlorophylls, and the water-soluble E141ii, consisting of copper chlorophyllins. The addition
of copper to the lipid or water-soluble chlorophylls contributes to stabilizing the molecular structure,
better preserving the natural green color.[102] US FDA authorizes only the use of copper chlorophyllin
as natural green food colorants, which may be used only for coloring citrus-based dry beverage
mixes.[102]
Carotenoids are water-insoluble pigments, generally consisting of C40 tetraterpenes/tetraterpe
noids, formed by eight C5 isoprenoid units.[11] Found in some animals, microorganisms,[59] and
higher plants, carotenoids are characterized by a peculiar yellow, orange or red color.[56] Carotenoid
pigments can be divided into carotenes (containing carbon and hydrogen) and xanthophylls (oxyge
nated derivatives, hence containing carbon, oxygen, and hydrogen).[56] Lycopene, β-carotene, and β-
cryptoxanthin belong to the carotenes, while lutein, zeaxanthin, and astaxanthin to the
xanthophylls.[57] Apocarotenoids are carotenoids with a shorter carbon chain, such as bixin (annatto
pigment) and crocetin (a component of saffron).[11] The pigments are located in the plastids (sub
cellular organelles), in the chloroplasts associated mainly with proteins and in the chromoplasts
deposited as oily droplets or in crystalline form.[103] More than 700 carotenoids have been isolated
from animals, plants, fungi, and microorganisms; remarkably, among them, around 50 exhibit
provitamin A activity.[55] For example, both carotenes and xanthophylls are present in green vege
tables; lycopene, as a red-color pigment, is typical for tomatoes; astaxanthin as the pink/red-color
pigment of crustaceans; β-carotene as the orange pigmentation of fruits and vegetables[62;] capsanthin
as the red-color pigment of peppers.[61] Lycopene can be also found in red citrus and watermelon.[58]
The major sources of zeaxanthin/lutein are corn and some squash varieties,[11] while lutein can be
found also in kale, spinach, and egg yolk.[58] Papaya, tangerine, and orange are good sources of β-
cryptoxanthin, whereas trout, salmon, and crustaceans of astaxanthin.[11]
Carotenoids are, in general, extremely sensitive molecules, and may undergo a quick degradation,
especially in the presence of reactive oxygen species, or exposure to light, acidic pH and high
temperature.[58] Previous researches revealed that longer thermal processing degrades carotenoids but
makes them more bioavailable. This may be due to the consequence that heating, coupled with
agitation (such as during thermal processing), causes the disruption of the cellular matrix of the
plant material, promoting lycopene release, and may also favor the trans to cis isomerization.[104,105]
European Union approved carotenoids as food additives (as colors) with E-numbers E160 and E161
(European Union regulation (EC) No. 1333/2008 on food additives of the European Parliament and the
Council of 16 December 2008). Few carotenes, such as E160a (paprika extract, capsanthin, and
capsorubin) and E160c, are authorized as quantum satis. Canthaxanthin is not authorized in the food
categories listed in Parts D and E. The substance is in list B1 because it is used in medicinal products per
Directive 2009/35/EC of the European Parliament and the Council, 2009 (OJ L 109, 30.4.2009, p. 10).[106]
EFSA Panel on Food Additives and Nutrient Sources added to Food evaluated canthaxanthin pigment.
Canthaxanthin (E161) is a carotenoid pigment allowed as a food additive in the EU and previously
evaluated by the Joint FAO/WHO Expert Committee on Food Additives in 1974, 1987, and 1995, and the
Scientific Committee and Panels in 1983, 1987 and 1997. Both committees have established a current
acceptable daily intake of 0.03 mg/kg of body weight.[107] US FDA permits the use of canthaxanthin as
a natural food color, depending on the good manufacturing practice and amount of usage.
Potential alternative sources of NPs

As discussed above, NPs exhibit wide differences in chemical structure, solubility, stability, and various
other properties. Natural colorants can be classified by source, as plant-, animal-, microbial- and
mineral-derived. Currently, the most commonly used NPs in foods include plant-derived and
mineral-based NPs.
Plant-derived pigments, such as anthocyanins, betalains, chlorophylls, and carotenoids are extremely
common in nature, as they play an important role in plant protection and biochemical pathways.[5]
Minerals are also well-known as a source of different color hues, depending on their chemical
structure.[5] For example, of particular relevance for food applications is viridian, a chromium oxide
mineral, known for its green pigmentation.[108]
Among the less conventional NPs sources, which are emerging as important supplies of different
colors for the food industry, there are animal-derived and microbial-derived pigments.
Animal-derived pigments, such as purines, anthraquinones, pterins, and others,[1,109] are important
for the transportation of oxygen in the blood and photoprotection.[1,110] Carotenoids can be found in
association with proteins, to form carotenoproteins[111] α-crustacyanin is an example of a blue-colored
pigment present in the carapace of lobsters (Nephrops Leach, 1814), where astaxanthin is responsible
for the formation of a complex quaternary structure by binding with an apoprotein.[112] Moreover, the
blue carotenoprotein linkiacyanin can be isolated from a starfish, Linckia laevigate L.,[113] while
a violet carotenoprotein can be found in Asterias rubens L. starfish.[114] Different carotenoproteins,
together with astaxanthin and canthaxanthin, can be recovered from Moina micrura (Kurz, 1874) and
Streptocephalus dichotomus (Baird, 1860) crustaceans.
Microbial organisms are also able to produce different types of colorants. Microbial-derived NPs
have relevant economic importance, because of faster growth, better growth control, and higher
renewability than plant and animal sources,[115] as well as cheaper production, easier extraction,
and higher yields.[116] Microbial-derived NPs, such as carotenoids, quinines, and flavins, are widely
used in food production.[117] Canthaxanthin can be isolated from Bradyrhizobium spp.,[117] while
astaxanthin can be recovered from Basidiomycetous spp. yeast hosted in different animals, such as
crayfish, red shrimps, feathers of some birds, salmons, and crustaceans.[118,119] The red pigment
prodigiosin can be found in different strains of Serratia marcescens bacteria. Purple violacein can be
produced by Chromobacterium violaceum and lycopene can be isolated from different fungi, such as
Fusarium, Sporotrichioides, and Blakeslea trispora.[117]
Remarkably, while the availability of plant-derived NPs fluctuates with heat, light, and weather
conditions,[120] microbial pigments can be produced throughout the whole year from microorganisms,
8 S. JURIĆ ET AL.
with the additional advantage that many of them might grow on cheap substrates.[115,116,121] NPs
production by different types of bacteria, fungi, algae, and yeast is influenced by the available carbon
and nitrogen sources, minerals, temperature, moisture content, aeration, and pH.[120,122] Filamentous
fungi are readily available raw materials that can be used to produce food-grade pigments. With easily-
arranged large-scale cultivation of well-known strains under a controlled environment, versatile colors
can be produced, depending on the fungal chemical and pigmentation profile. Specific pigments such
as natural blue or red colorants for cochineal extract/carminic acid/carmine replacement makes them
ideal for their exploitation.[123] For example, the soil microorganism Penicillium oxalicum var.
Armeniaca CCM 8242 (a soil isolate) can produce a red color pigment (Arpink red, nowadays called
Natural red), [123] whose use in different foods as red-coloring matter raised no objection from the
Codex Alimentarius Commission (Rotterdam meeting, 2002).[124]
Microalgae, which might grow both in salt or freshwater, represent an important and renewable
source of carotenoids,[60] chlorophylls and red or blue pigments, known as phycobiliproteins.[125]
Phycocyanin is the part of phycobiliprotein complexes assembled into extremely large macromole
cular complexes called phycobilisomes. Phycobiliproteins are brilliantly colored, highly fluorescent,
water-soluble protein components. Different algal classes are known for their characteristic pigments
and features. For example, Chlorophyceae class is rich in chlorophylls a and b, carotene and xantho
phyll, and is characterized by a dark to light green coloration, similar to higher plants. Xanthophyceae
class are known for chlorophyll a, c and e, β-carotene and xanthophylls including neoxanthin and
violaxanthin. The excess of yellow xanthophylls gives the characteristic color of yellow-green.
Cryptophyceae class contain chlorophylls a and c, β-carotene, fucoxanthin, diatoxanthin, and diadi
noxanthin. Protozoa-like algae might exhibit various colors, including green, blue, brown, olive, and
red, due to the presence of chlorophylls and other secondary pigments, such as α-carotenes bilipro
teins and unique xanthophylls. Rhodophyceae class is known for the characteristic red color of the
algae, which is due to the presence of the pigments phycoerythrin and phycocyanin.[126]
Remarkably, the control of the main abiotic and biotic factors, such as temperature, light, and
nutrition, together with metabolic or genetic engineering, can be exploited to maximize the pigment
content in the microbial and microalgal cells.[117,127]
Both microalgae and microorganisms offer significant advantages not only in terms of better
growth control and renewability but also of simpler processes for the recovery of the pigments. This
is due to the easier concentration of the biomass and lower resistances to the mass transfer of the
pigments from the intracellular space to the solvent than for other types of biomass.
To date, several NPs of microbial origin, such as the colorant recovered from Monascus species, the
already cited Natural red from Penicillium oxalicum, microalgal phycocyanin from Arthrospira
platensis (Spirulina), lycopene and β-carotene from Blakeslea trispora, riboflavin from the mold fungus
Ashbya gossypii, β-carotene from the microalgae Dunaliella salina, and astaxanthin from the bacter
ium Paracoccus carotinifaciens and microalgae Haematococcus pluvialis are widely used in the food
industry.[117,128,129]
Another important source of NPs, which is on the rise for being intrinsically sustainable, is
represented by agri-food by-products and residues, whose exploitation for NPs recovery might
contribute to reducing the environmental issues associated with their management and disposal.
The seeds and surface of many fruits and vegetables, discarded during processing, may still contain
large amounts of carotenoids, often in higher concentration than in the parts retained for industrial
use.[130] For example, tomato (Solanum lycopersicum L.) peel is five times richer in lycopene than the
pulp.[131-133] Similarly, when compared with the pulp, a higher carotenoid content has been found in
avocado (Persea americana Mill.) peels (2.6 times) and seeds (2.4 times), as well as in banana (Musa
acuminate Colla) peels (5.3 times).[130] In the case of mango (Mangifera indica L.), despite the pulp is
very rich in carotenoids (45.30 µg/g),[130] significant amounts (3092 µg/g) can still be found in the
dried peel powder.[134] Remarkably, 29% of the dry weight of coffee (Coffea arabica L.) is left as a by-
product after coffee processing, including the exocarp, which is a source of anthocyanins,[135] whose
main components are cyanidin 3-rutinoside and cyanidin 3-glucoside.[136] Under optimal extraction
conditions, up to 0.145 mg cyanidin 3-glucoside/g of coffee fresh exocarp can be recovered and used as
NP in food products.[137] Apple (Malus domestica Borkh., 1803)peels, constituting up to 13% wt. of the
fresh apple,[138] can be a valuable source of cyanidin-3-galactosides (anthocyanins), with the content of
510-1275 mg/kg of dried apple peels/skin.[139] The stalks left after harvesting red beet (Beta vulgaris
subsp. vulgaris), which currently represents a potential environmental burden, are a valuable pigment
source, with up to 5.31 mg of betaxanthin and 1.28 mg of betacyanin per gram of waste.[140]
However, the exploitation of agri-food residues has to take into account the variable quality of the
raw materials, and the high costs that might be associated with their collection and transport, as well as
with the extraction, concentration and purification processes.
A more sustainable exploitation strategy of agri-food residues that has recently emerged is their use
as a substrate for the growth of pigment-bearing microorganisms, with the additional advantage that
growth conditions compatible with a non-toxic and non-pathogenic microbial source are used.[141]
For example, the fungal strains Penicillium purpurogenum and Monascus purpureus, which are
reported to grow on orange by-products, such as peels, pulp, and seeds, have been recognized as
a potential source for red and yellow pigments.[142] The corn steep liquor, a secondary stream obtained
during the corn wet milling process, can be used as a substrate for Monascus ruber, which produces
a red colorant.[143] Mung bean (Vigna radiata L.) waste flour can, instead, be used as a substrate for
carotenoid-producing Rhodotorula glutinis.[144] Rhodotorula rubra can produce a yellowish-pink
pigment, using as substrate cheese whey from cheese manufacturing and coconut water.[145] The
yeast Rhodotorula mucilaginosa was shown to be able to produce carotenoids when growing on coffee
pulp and husk.[146] Astaxanthin was produced by different yeasts, such as Yamadazyma guilliermondii,
Yarrowia lipolytica, Xanthophyllomyces dendrorhous, and Sporidiobolus salmonicolor, growing on
wheat waste.[147]
The wide availability of natural-derived pigments, together with the increasing consumers’ concern
towards the toxic effect of artificial additives in food products, has led to the rapid decline of the use of
synthetic pigments. Pigments like β-carotene, astaxanthin, and lutein, which can be recovered from
natural sources, such as fungi, bacteria, and algae, are receiving nowadays increasing attention because
they can be safely utilized in foods, pharmaceuticals, nutraceuticals, animal feed, supplements, and
cosmetics.[148] Moreover, the extraction of NPs with alternative technologies (e.g. extraction with
supercritical carbon dioxide, assisted by ultrasound, enzymatic, microwave, or high-pressure treat
ments) and using wastes and by-products as a source, present a promising approach to pursue a more
sustainable, green and economically favorable food production.[149]
Extraction of NPs
The extraction of NPs can be a challenging task, depending on the source, especially if high-quality
NPs are desired.[150,151]
Conventional extraction methods, in agitated vessels, are frequently used because of their simpli
city, low cost, and versatility. The selection of the most appropriate solvent and process temperature,
depending on the NPs properties and source structure, represent the most important parameters
regulating extraction yield and selectivity.[152] However, conventional extraction methods are not only
characterized by poor extraction efficiency, but require also large volumes of organic solvents, and may
cause thermal degradation of NPs.[153] More advanced extraction methods, by enhancing the mass
transfer of intracellular material, have been increasingly investigated to reduce the extraction time and
consumption of organic solvents, while increasing yield and quality of the extracts.[150] For example,
the use of emerging technologies, based on microwaves, ultrasound, high-pressure, or high-pressure
carbon dioxide is reported to improve the efficiency of extraction processes, significantly enhancing
the selectivity and efficient recovery of high-quality extracts from different plants and biomasses, using
green solvents.[154-159] Other emerging technologies, capable of reducing the mass-transfer resistances
by permeabilization or disintegration of cell walls and membranes, such as ultra-high pressure,[160,161]
negative pressure cavitation,[162] high voltage electrical discharges, ohmic heating and pulsed electric
10 S. JURIĆ ET AL.
fields,[60,163-166] mechano-chemical methods,[167] and high-pressure homogenization,[131,168,169]

resulted to be very efficient in promoting NPs extraction from different types of biomass, in an
environmentally friendly way, aligned with the principles of green chemistry.[159] These novel extrac
tion methods enable faster, more sustainable and efficient extraction processes, specifically suitable for
industrial scale-up.[170]
Health-promoting properties of NPs

The use of NPs in food products contributes to a clean food label declaration, due to its eco-friendly
properties and natural sources.[86,171] However, the use of NPs is of great interest to food processors
also because NPs have a strong biological activity, deriving by their key role in photosynthesis, as well
as many other biochemical pathways. The fact that, throughout history, pigments were regularly
consumed, provide evidence that human and animal body found a way to take advantage of the
physicochemical characteristics of NPs.[172] Fruit and vegetable intake are correlated with a lower risk
of cancer development, and protection against some chronic diseases,[173] and this could be, at least
partially, ascribed to the NPs present in them. Many NPs of microbial origin, are also characterized by
health benefits, because micro-organisms produce large quantities of bioactive compounds, with
antioxidants, antimicrobial, anticancer, immuno-regulatory, and anti-inflammatory properties.[117]
However, it must be pointed out that neither the consolidated uses and usage levels nor the natural
origin may ensure the lack of toxicological risks associated with the use of NPs. The main risk factors
are associated with the specific properties of the NP molecules, the applied dose, as well as the presence
of toxic compounds or traces of organic compounds not completely removed during refining (for
example, citrinin in Monascus pigment[174]). In the last years, the European Food Safety Agency
(EFSA), through its Panel on Food Additives and Nutrient Sources added to Food has published
several scientific opinions, to which the reader is addressed (https://www.efsa.europa.eu/en/panels/
faf), on the use of different NPs as food additives, where the main scientific data for assessing the safety
concerns associated with their use are reported and critically analyzed. Especially in the case of NPs
from fungal sources, the use of several fungal pigments is prohibited because they might contain
mycotoxins.[175] It is therefore important to use non-toxic and non-pathogenic strains for NP
extraction, and the accurate biosynthetic screening for toxin production of newly-discovered pigment-
producing microbes.[117]
Anthocyanins are known to be strong antioxidants with high capacity to scavenge free
radicals.[176,177] They play an important role as inhibitors for mutagenesis and carcinogenesis.[67,86]
The observed cancer-prevention action associated with the consumption of anthocyanins has been
ascribed by several authors to an anti-tumoral/inhibitory effect.[73] The intake of anthocyanins,
because of their high antioxidant capacity,[86,178] has been reported to help also in the prevention of
cardiovascular disease occurrence,[67,179] neuronal illness (cognitive function/age-related disease),
diabetes and inflammation,[180] as well as of bacterial and viral infections.[181]
Betalains are also important in the prevention of cancer [22] and oxidative-stress related
disorders,[182,183] because of their high antioxidant activity.[184] Furthermore, betalains exhibit antic
ancer properties,[185] anti-lipidemic effects,[186] and high antimicrobial activity.[187] It is important to
mention the high potential of betalains to neutralize reactive nitrogen species. Sakihama et al.
demonstrated betalains’ ability to inhibit ONOO− -dependent nitration of tyrosine.[188] Moreover,
clinical trials revealed that betalain-rich red beet juice mitigates hepatic toxicity caused by
N-nitrosodiethylamine (NDEA), carbon tetrachloride[189] and 7,12-dimethylbenz(a)anthracene
(DMBA) as a result of improved antioxidant status and enhanced expression of phase II enzyme
quinone reductase.[190] Furthermore, the researchers observed a 40% reduction in incidences of
4-nitroquinoline-l-oxide (4-NQO) initiated and 8% glycerol promoted pulmonary tumors. Oral
feeding of betanin (2.5 mg/100 mL water) significantly inhibited symptom of splenomegaly as well
as skin tumor initiated by DMBA and promoted by UV light-B.[191]
Table 2. Health benefits of carotenoids, including in vivo and in vitro investigations, research on cells, animals, and humans.
Carotenoids References
Alpha-carotene
[206–210]
decreases the risk of glaucoma occurrence
positive effect on atrophic gastritis
prevents lipid oxidation and enhances gap junction communication
decreases risk for gastric/liver/lung cancers, diabetes, cardiovascular and chronic respiratory diseases
inhibits proliferation of cancer cells
Beta-carotene
[211–223]
lowers the incidence of metabolic syndrome and prevents lipid-mediated disorders (insulin resistance) associated with lower cardiovascular disease, stroke, and cancer
decreases body mass index score, waist-to-height ratio, and subcutaneous adipose tissue content
helps in the prevention of coronary artery diseases
reduces the incidence of disease biomarkers of several chronic complications (type-2 diabetes)
highest vitamin A activity among other provitamins A carotenoids (α-carotenes and cryptoxanthins)
inhibits neuroblastoma, the most prevalent extracranial solid tumor in childhood
a potent antioxidant, functions as a lipid scavenger and a singlet oxygen quencher
possesses antioxidant properties and prevents the occurrence of fatty liver disease
reduces susceptibility to breast cancer risk in premenopausal women
reduces the risk of age-related macular degeneration
inhibits ultraviolet-induced skin cancer and oral carcinomas
moderately effective as adjuvant treatment in erythropoietic protoporphyria
Beta-cryptoxanthin
(Continued)
11
12
S. JURIĆ ET AL.
[224–247]
reduces the risk of visceral fat accumulation and high blood pressure
reduces the risk of hyperglycemia and dyslipidemia
decreases the risk of renal cell cancer development
lowers the risk of developing dyslipidemia and largely decreases body mass index
shows cardioprotective effects
decreases the risk of breast cancer development
represses lipid peroxidation and prevents inflammation caused by oxidative damage
decreases the risk of oral and pharyngeal cancer
suppresses the expression of lipopolysaccharide-inducible and/or TNFα-inducible genes
represses osteoclastic activation and prevents bone loss
decreases the risk of colon cancer
reduces lung cancer deaths in smokers
enhances the antitumoral activity and decreases the risk of stomach cancer
inhibits chemically induced lung tumorigenesis
controls bodyweight, abdominal adipose tissue weight, and waist circumference and blood glucose
improves adipocytokine profiles
modulates skin tumorigenesis
inhibits smoke-induced inflammation in human lungs
improves bone mass density
normalizes serum lipid levels
stimulates bone formation and repress bone resorption
inversely correlates with type 2 diabetes
positive effect on bone metabolism and provides anti-osteoporosis activities
decreases oxidative stress and protects against DNA oxidation damage
Lycopene
(Continued)
[248–285]
cardioprotective effects
protective role on gamma radiation at high oxygen concentrations
protects cells from ionization damage
attenuates and prevents impairment of learning ability and memory ports
anti-ulcer role
inhibits cell proliferation in human colon cancer HT-29 cells
reduces high fat-induced hepatic steatosis
reduces the development of the most common type of kidney cancer
inhibits the hepatic injury induced by alcohol
neuroprotective effects on Alzheimer disease, Huntington’s disease, and brain ischemia
prevents the reduction of the number/motility of spermatozoa and protects from testicular damage
anti-inflammatory effect
inhibits Helicobacter pylori infection
induces apoptosis of prostate cells and has a protective effect on prostate cancer
natural chemoprotective and antioxidative agent
protects the pancreatic tissues from cerulean-induced oxidative damage
inversely associated with type-2 diabetes and impaired glucose metabolism
influences cognitive function by improving the memory
antigenotoxic and antimutagenic effect on HepG2 cells
inhibits human liver cancer cells proliferation and prevents the metastatic process
inhibits the infiltration of inflammatory immunocytes into the bronchoalveolar lavage
decreases bacterial growth and improves prostatic inflammation caused by chronic bacterial prostatitis
reduces the formation of atherosclerotic plaques in the aorta and improves lipid profiles
reduces the risk of prostate, lung, leukemic and digestive tract cancers
shows chemopreventive effects in liver and ovary cells
reduces the prostate cancer risk
inhibits the proliferation of prostate cancer cells
inverses proliferation of colon cancer, lymphocytic leukemia, erythroleukemia, and Burkitt lymphoma
protective effects against DNA damage in cancerous human cells
protective effect from iron-induced oxidative damage in prostate tissue and reduces lipid peroxidation
reduces blood pressure in patients with hypertension
decreases plasma glucose and fasting insulin concentrations
protective effects against DNA damage in both normal human cells
decreases serum lipid peroxidation and LDL oxidation
delays the onset and progression of galactose cataract and reduces the incidence of selenite cataract
high antioxidant activity by quenching of singlet oxygen and the scavenging peroxyl radicals
induction of cell-cell communication and growth control
protects human lymphoid cells from singlet oxygen by binding to the surface of the cells
has a moderate curative effect on the radiation lesions and increases the survival rate
Lutein
(Continued)
13
14
S. JURIĆ ET AL.
[286–302]
antimicrobial effect and antioxidant activity
anti-inflammatory effects and cytoprotection against alcohol-induced liver injury
protects against skeletal ischemia/reperfusion injury
potential beneficial effects in diabetes-induced testicular damage
potentially can prevent inflammation-related neurodegenerative disorders
increases macular pigment optical density and visual sensitivity in early age-related macular degeneration
due to the high antioxidant activity can prevent hepatotoxicity
shows antiatherogenic properties
modulates the expression of growth- and survival-associated genes in prostate cancer cells
neuroprotective effect against transient cerebral ischemic injury
might be effective in the strengthening of immunity
antihypertensive and antioxidant effects against hypertension
reduces levels of biomarkers of cardiovascular diseases
may improve the ability to drive at night
potent neuroprotective agent
effective antiulcer agent
acts as a filter of harmful blue light in the eye and prevents the production of free radicals
Zeaxanthin
[302–311]
protects from neurodegenerative disorders
enhancer of antioxidant defense against neuronal damages
protects against cardiovascular diseases
protects against oxidative stress and has strong antioxidant activity due to the scavenging abilities
inversely associated with colorectal neoplasm
inversely associated with breast cancer risk among women
helps to stop or slow down age-related increases in human eye lens’ density
lowers the risk for age-related macular degeneration development
increases macular pigment density
acts as a filter of harmful blue light in the eye and prevents the production of free radicals
Fucoxanthin and its metabolites
(Continued)
[312–324]
stimulates levels of cytokines in cells
inhibits factors which regulate genes controlling inflammatory cytokine production
reduces blood triglyceride concentration
improves phagocytic and microbicidal capacity, decreases oxidative damage to lipids/proteins
inhibits pro-inflammatory factors
chemopreventive effect on urinary bladder cancer cells
inhibits the viability of human leukemia cells
suppresses adipocyte differentiation of murine preadipocyte cells
anti-obesity activity – reduction of white adipose tissue
shows anti-cancer effects on human colorectal adenocarcinoma cell lines
inhibits the development of endotoxin-induced uveitis
stimulates some pro-inflammatory genes
chemopreventive effects in colon carcinogenesis
inhibits the growth of human neuroblastoma cells
Canthaxanthin
[325–340]
scavenges reactive oxygen species
suppresses the release of pro-inflammatory cytokines and tumor necrosis factor-alpha
anti-atherogenic properties
increases the in vitro metabolism of aflatoxin B1
the apoptosis-inducing effect in human colon adenocarcinoma and human melanoma cells
exhibits immune-modulatory activities
reduces the size of skin papillomas
high potential in the prevention of cervical cancer
increases host resistance to pathogens
enhances gap junction communication between cells
chemoprotective agent against colon carcinogenesis
inhibits neoplastic formation
reduces the incidence of preneoplastic lesions and neoplasms
lowers the incidence of preneoplastic lesions and neoplasms in the bladder
reduces the growth of melanoma tumor and fibrosarcoma tumor cells
chemopreventive effect on mammary cancer
increases mitogen-induced lymphocyte proliferation
Astaxanthin
(Continued)
15
16
S. JURIĆ ET AL.
[335, 337, 341–349]
reduces the wall/lumen ratio of a coronary artery with fewer and straighter elastin bands in the aorta
improves insulin sensitivity
decreases renin-angiotensin system activity and systolic blood pressure
reduces serum triglycerides and increased high-density lipoprotein cholesterol and serum adiponectin
beneficial effects on human blood rheology
positive effects on a series of metabolic syndrome features
decreases blood pressure and delays the incidence of stroke
lowers hepatic metastasis
reduces the size of fibrosarcoma
reduces the incidence of preneoplastic lesions and neoplasms
lowers the incidence of preneoplastic lesions and neoplasms in the bladder
Chlorophylls are also involved in a significant number of biological functions, both in plants and
animals. Because of their bioactive properties, chlorophylls have a significant influence on human
health, as they can contribute to rebalancing the gut microbiota, and, because of their chemical
structure, exhibit antioxidant and antimicrobial properties.[192] Scientific studies revealed that the
intake of chlorophylls with foods may beneficially act on human health[193] through antioxidant,
antimutagenic,[194] and antigenotoxic activities.[195] Numerous in vivo[196] and in vitro[197] studies
revealed chlorophyll’s chemo-preventive effects in humans.[198]
Chlorophyll is structurally similar to hemoglobin and regenerates or acts as a substitution for
hemoglobin in hemoglobin deficiency conditions. In clinical conditions, such as thalassemia and
hemolytic anemia, the utilization of chlorophyll-rich juice is recommended.[199] Chlorophyll, together
with enzymes, such as superoxide dismutase, plant hormone abscisic acid or dormin, and its alkalinity
enables the development of an important anticancer function.[200]
Carotenoids represent the main class of NPs with health-beneficial properties. They are claimed to
exhibit a wide array of health-promoting benefits, such as prevention of human diseases like cancer and
age-related disorders.[201] They are also a vital dietary source of vitamin A (α-carotene, β-carotene, β-
cryptoxanthin).[202] Some of the functions of vitamin A include regulation of hormone synthesis and
improvement of immune systems, as well as growth and differentiation of skin cells.[203] Furthermore,
there are some indications that carotenoids may help in the prevention of cancer occurrence,[201]
cardiovascular diseases,[201] osteoporosis,[204] lung diseases,[205] and neurodegenerative disorders.[204]
Some notable examples from the last few decades of health benefits associated with the consumption of
carotenoids are reported in Table 2, together with a summary of the research activities that proved them.
As the interest towards NPs in the food production rises, it must be remarked that proper handling
is of paramount importance to achieve their maximum potential both in terms of food coloring
function and health-beneficial properties. Food processing often alters these functions and properties,
with the final product not resulting as expected. The inclusion of NPs in food products, while
preserving their functional features, is a crucial challenge for the food industry, and new approaches
should be employed to prevent undesirable changes both in NPs as well as in the final food products.
NPs as food colorants/additives in foods

Manufacturing and processing procedures, storage conditions, and food preparation may have
a pronounced influence on food final coloration, altering the natural color intensity and hue.[350]
Especially those operations involving the use of heat are generally responsible for the significant
alteration, degradation, or even loss of food color.[156,351] Therefore, the use of NPs in food is justified
by different purposes, such as (a) enhancing or intensifying its original color, (b) ensuring color
uniformity to improve the food appearance, or (c) providing color to otherwise uncolored food.[351]
In Table 3, the recent scientific literature (2016-2020), most relevant to the use of natural food
colorants/additives in foods, is overviewed per class of colorant, highlighting the main
applications,[350,401] as well as the stability issues associated with the inclusion in foods.[6,351] Fig. 1
helps the reader to navigate through Table 3, by identifying the main class of foods where the different
natural pigments are used.
When considering the sensory perception of food, the stability of food colorants and stored
products is one of the most important factors to be considered. Visual signs are the first ones to
reach consumers, thus influencing their food preference, acceptability, and food choice. Food indus
tries aim to provide uniform, attractive, and pleasantly colored foods to satisfy consumers’ expecta
tions, desires, and needs.[350]
There are many reports of the negative influence of some environmental factors on natural food
color stability, i.e. temperature, light, oxygen, pH, solvents, packaging materials, and storage condi
tions, all of which can influence food product attractiveness. A good example of very unstable natural
pigments used in food products is anthocyanins. When they are added to foods, they are influenced by
the food matrix (fats, proteins, moisture content, pH, etc.) and environmental factors. The
18
Table 3. Main NPs of interest to the food industry, with the indication of their major sources, products where they have been included, and a brief description of the issues associated with their use in
food, and stability.
Anthocyanins (E163; red-purple-blue; water-soluble)
Sources Application Ingredient extraction/formulation Stability Additional notes
Seeds, Cob, Details on application in bread, cakes, cookies, tortillas, beverages, soft drinks, dry mixed concentrates, candies, chewing gums, sweet jelly, confectionery, frozen dairy desserts, dairy
S. JURIĆ ET AL.
Grains, products, yogurts, sauces, baked foods, gelatin reviewed by Martins et al. (2016).[350]
FruitsBark,
HusksLeaves,
RootsTubers,
Bracts, Nuts,
Flowers
Rosella fruit Jelly and yogurt; red Highly-efficient extraction from Hibiscus Pigment retention: 87% (50°C), 61% (80° Content and antioxidant capacity of
(Hibiscus pigments,[352]beverages,[353,354]yeast,[355] sabdariffa calyces, when assisted by C) and degradation under acidic pH, aqueous infusions, as a function of the
sabdariffa L.) red pigments for food[356] ultrasounds (51.76 mg anthocyanins/g dark-incubated (pigment retention: size of suspended particles.[354]
[356]
extract). 84%, 10 days).[352] Degradation kinetics Development of a rapid and efficient
Sustainable source of natural colorants revealed that anthocyanins could be technique to monitor pigments in
with potential application in different heated to temperatures up to 70°C beverages during processing or
[356]
industrial sectors. with antioxidant contents gradually storage.[353]
decreasing.[352]Microcapsulation in
heat-treated yeast cells provides high
anthocyanin protection during storage,
if enzymes are inactivated.[355]
Skin grape (Vitis Kefir and carbonated water[357] Lyophilized anthocyanin obtained from Kefir: pigment retention of 67% during Recommendations are to store the
vinifera L.) grape skin extract.[357] storage, the half-life is of 27 days.[357] carbonated beverage in amber bottles
Carbonated beverage: light loss of when it is incorporated with
coloration after 16 days of storage.[357] anthocyanins as a natural colorant.[357]
The exposure to light of carbonated
beverage during storage resulted in
significant loss of coloration at the end
of the 16 days of storage.[357]
Onion (Allium Yogurt[358] Plant tissue powder extracted with water/ In vitro: color preserved during storage for No sensory characterization was
spp.) wastes glycerol/2-Hydroxypropyl-β- 4 weeks at 5°C and low pH (4.5). In investigated.[358]
cyclodextrin (at 40–80°C). Eco-friendly product: due to lower pH, significantly
extracts rich in polyphenols from onion lower color stability.[358]
wastes. Extracts were successfully
employed for the production of a red
colored stable fermented dairy product
of yogurt type.[358]
(Continued)
Edible flowers Yogurt[359] Coloring yogurts with yellow-orange hues Rose extract is the most suitable No significant changes in nutritional
rose (Rosa (using extracts). The extraction yields alternative to E163.[359] value, free sugars and fatty acids
spp.), (mg of anthocyanin per 100 g of petals) composition.[359]
cornflower obtained for extracts were 53% for
(Centaurea dahlia, 46% for rose, and 23% for
cyanus cornflower.[359]
L.) and dahlia
(Dahlia
pinnata Cav.)
Black carrot Colorants,[360,361]yogurt [361] Whey protein-based microcapsules were Bioactive compounds were moderately Not suitable for yogurt or jelly model
(Daucus produced with black carrot affected by the processing steps with system.[361]
carota concentrate. Encapsulation in protein the largest changes occurring early in
L.) isolate hydrogel emulsion, suitable for the process.[361]Pasteurization caused
dairy products (yogurt) to provide a higher rate of total phenolic and
different shades of pink.[361] flavonoid degradation compared to
Anthocyanins were extracted from anthocyanins.[361] Microcapsules were
black carrot with ethanol/citric acid able to protect the color by increasing
and ethanol/acetic acid.[362] its stability.[361]Extracts had the highest
degradation in alkaline pH.[360]
Colorant: stable for 25 d of storage,
when extracts obtained in ethanol/
acetic acid solution.
Microencapsulation in tetraethyl
orthosilicate did not prevent
degradation.[360]
Purple carrot Beverages, powders[363,364] Anthocyanin powder extracted from The addition of amino acids or peptide The degradation of anthocyanin was
(Daucus purple carrot. Anthocyanin powder (0.1%) improved color stability accelerated with addition of ascorbic
carota ssp. was dissolved in citric acid. Different (0.025%) with a half-life of 2–6 d.[364] acid.[365]
sativus) ratios of amino acid/peptide solutions Addition of polyphenols (0.2%)
and anthocyanin-calcium were delayed color fading. Half-life: from 2.9
combined. L-ascorbic acid powder was to 6.7 days, when combined with green
added to the anthocyanin amino acid/ tea extract.[364]
peptide mixed systems.[365]
Purple corn (Zea Beverages[366,367] Purple corn cob extraction yields with Zinc and alginate protected corn extracts Zn/alginate interacts with anthocyanin
mays L.) 60% ethanol (> 13.5 mg/g FW) is from degradation and improved from purple corn and slows its
comparable to 70% acetone (14.3 mg/ stability of color.[367] Color changed chemical degradation.[367]
g FW).[366] over 6-month storage; more yellow at

a higher temperature, turned to light
red when exposed to light.[368]
(Continued)
19
20
Purple sweet Colorants,[368–370]native antioxidants in Water extraction gave high extraction Anthocyanin content decreased and the The purple-fleshed sweet potato
potato stored fried snacks[371] yields. Extracts with good stability at color changed over a 6-month storage anthocyanins were more stable during
S. JURIĆ ET AL.
(Ipomoea pH 2.0–6.0. Color remains stable during period. Degradation process was storage than other anthocyanins
batatas L.) 30 d-storage at 20°C in the dark.[369]Co- accelerated in the presence of light and because they mainly consist of di-
[368–370]
pigmentation: color modulation by at an elevated temperature. The acylated and double glycosylated
adding to the extracts various dosages color did not change significantly forms of peonidin derivatives.[368–370]
of chlorogenic, rosmarinic acids, food- during 3-month storage.[371] Acylated
grade phenolic apple, rosemary anthocyanins – inhibited oxidative
extracts. Extracts from colored-flesh changes in lipids of the snacks.[371]
potatoes provided a pink/purple color
to snacks and increased antioxidant
activity.[370]
Barberry Colorant powder[372] Microencapsulation by spray drying with The combination of maltodextrin and Samples produced with maltodextrin and
(Berberis a combination of maltodextrins and gum arabic with core/wall ratio of 25% gelatin had highest efficiency and best
vulgaris L.) gum arabic for high encapsulation led to the higher efficiency and powder quality.[372]
efficiency.[372] provided better protection of the
anthocyanin pigments.[372]
Chokeberry Berry-based colorants,[373,374]beverages[374] Co-pigmentation: modulation of extract Aronia berry juice: addition of quercetin, The use of herbal extracts led to
(Aronia color with polyphenols, herbal extracts, sorbitol, and chlorogenic acid inhibited a significant hyperchromic effect at
melanocarpa for enhanced color stability in pH-induced loss of color.[374] much lower pigment/co-pigment
(Michx.) functional foods.[373] ratios, compared to pure
Elliott) compounds.[373]
Blackberry Colorants, healthy ingredients[375,376] Microencapsulation: maltodextrin 10DE Degradation of anthocyanins during Light had a higher impact on anthocyanin
(Rubus spp.) to produce powders by freeze- storage under light and different stability than elevated
[376]
drying. temperatures showed first order temperatures.[375]
kinetics. Co-pigmentation with rutin The observed color changes did not
and ferulic acid enhances the shelf-life correlate with the anthocyanin loss.[375]
stability.[375]
Red cabbage Colorants, food/beverage, Fresh red cabbage wastes from industrial Stable color in 48% ethanol, pH 6.3, for The stability of various anthocyanins from
(Brassica antioxidants[362,377] processing, consisting in damaged 20 weeks. Colorimetric changes due to red cabbage was related to the number
oleracea L.) leaves mixed with stubs was prepared acids, calcium, magnesium, sodium, of acylated groups and mainly affected
as hydroethanolic extract.[377] and honey, glycine, xylitol, lactose, by illumination.[362]
[377]
Nineteen food additives/ingredients maltose. The stability of the waste
were used to evaluate stability of extract is affected by illumination.
color.[377] Good stability at low pH value and as
a dry powder stored in darkness.[362]
(Continued)
Black bean coats Colorants, antidiabetes,[378,379]sports Anthocyanin-rich powders and aqueous Stable at pH 2.5 and 4°C (89.6%); half-life The selected co-pigment is permitted for
(Phaseolus beverage, aqueous extracts and extracts (water-citric acid 2%) were of 277 days.[378]All bean coat use by the food industry and its
vulgaris L.) powders[378] obtained from black bean seed coats anthocyanins encapsulated with β- concentration is in agreement with
and applied to color a sport beverage cyclodextrins to extend half-life (up to Codex Alimentarius International Food
(with 13 months), fewer differences in Standards.[379]
and without 2% b-cyclodextrin).[379] colorimetric properties (darkness, 4°
C).[379]
Black rice bran Colorants, dark purple pigments [380–382] A high content of total anthocyanins in High color stability at 60°C and pH 2.0.[381] Colorants prepared using these
(Oryza sativa bran extracts, when using Ohmic Stability increased via intermolecular techniques exhibited the highest
L.) Heating-assisted extractions.[380] co-pigmentation, though the addition concentrations of phenolics,
Phytochemicals in black colorant of glucose, phytic acid, and/or gallic carotenoids and total
[382]
powder were prepared from black acid. anthocyanins.[381]
waxy rice
bran.[381]
Blue petals Baby products[383] Anthocyanins extracted from vivid blue Anthocyanins extracted from vivid blue There is no report on the potential of
(Clitoria petals of Clitoria ternatea L. (legume petals of Clitoria ternatea L. are applications of this natural colorant for
ternatea L.) crop).[383] sensitive and quickly degrade in the coating technology.[383]
presence of light.[383]Citric acid caused
a stronger red color degradation in
comparison to tartaric acid.[383]
Black goji fruit Colorants[384] Petunidin was ~85% of total pigment. Acylation increased color retention, Black goji anthocyanins produced various
(Lycium Black goji produced vivid hues over tinctorial strength, pigment vivid hues over wide ranges of pH.[384]
[384] [384]
ruthenicum a wide pH range. stability.
Murr.)
Betalains (E162; yellow-orange, red-violet; water-soluble)
Fruits, roots Details on application in burgers, desserts, ice cream, jams, jellies, soups, sauces, sweets, drinks, dairy products, yogurts reviewed by Martins et al. (2016).[350]
[385] [365]
Beetroot Colorants, gummy candy, cow milk Red beet extract diluted with dextrin and The stability of betanin increased when Nanoencapsulation of betanin exhibited
(Beta vulgaris applied to gummy candies.[385] loaded in nanoliposomes.[385]Stability no considerable effects on overall
L.) of betalains studies in cow milk during sensory acceptance of gummy
heating at 70–90°C revealed candies.[385] Further studies are needed
degradation with increasing on degradation kinetics of natural color
temperature and time.[365] pigment in real food systems to
observe relationship between changes
in color and shelf life of food products
during processing and/or storage.[365]

(Continued)
21
22
Cactus pear Fruit juice, yogurt, soft-drink[386] Betaxanthin-rich extract obtained from Color stability: high pigment retention in Spray-drying microencapsulation
(Opuntia ficus- cactus pear.[386] the dark, for 28 d at 4°C through spray- produced powders easy to store and
indica (L.) drying microencapsulation.[386] deliver.[386]
Mill.)
S. JURIĆ ET AL.
Red pitahaya Yogurt,[387,388] gelatin gummies and Betacyanins extracted with water and Betacyanins in yogurt reduced syneresis The color acceptability of yogurt
fruits beverages, yellow pigments[389] addition of pectinase from red sustained the viability of culture, containing colorant preparation from
[387,388] [387]
(Hylocereus pitahaya. Aqueous extracts from increased the antioxidant activity. red pitahaya was similar with
polyrhizus S. pruinosus fruits were obtained.[389] Spray-drying microencapsulation with E162.[387,388]Pigments from S. pruinosus
(Weber) potato succinylated starch enhanced fruits can substitute synthetic yellow
Britton & stability during storage for 32 d in pigments in manufactured food.[389]
[388]
Rose) yogurt at pH 4.6. Stability increased
with ascorbic acid. Betaxanthins were
more stable in gelatin gummies than
beverages.[389]
Purple flowers Ice-cream betacyanins[390] Ultrasound-assisted extraction of purple Advantageous effects of freezing (−22°C) Suitable natural ice-cream colorant.[390]
(Gomphrena flowers followed by lyophilization and to maintain color throughout storage
globosa L.) incorporation in ice-cream.[390] time.[390]
Ayrampo seed Yogurt,dairy products[391] Aqueous extracts from ayrampo seeds Stability of extracts was high during heat Stability higher than red beet extracts and
extracts were used in yogurts with two fat treatment and storage, and better in comparable with synthetic
(Opuntia levels (0.1 and 3.0%).[391] yogurt than in red beet extracts. Color colorant.[391]
soehrensii retention of dyed yogurts was not
Britton and influenced by fat content.[391]
Rose)
Mature fruits Foods and beverages, powders[392] Flesh of Basella alba L. mature fruits High stability of red color at pH 3–8. Application in real food systems are
(Basella alba bearing deepviolet juice may serve as Suitable for non-transparent packaging necessary to investigate to observe
L.) natural colorant.[392] materials.[392] possible changes in color.
Fruits (Basella Fruit juice, powders [393] Manually de-seeded fruit pulp was Stability of encapsulated betalains up to Two years storage at 4°C did not hamper
rubra L.) extracted using distilled water. 2 years storage at 4°C (spray the quality of spray dried powder.[393]
[393]
Betalains rich extract was mixed with drying).
50% maltodextrin and spray dried.[393]
Purple cactus Colorants[9] Betalains extracted from purple cactus The stability of encapsulated extracts was Possible application suggested only in
fruits (Opuntia fruits and encapsulated in calcium- better at low relative humidity, while low-humidity foods.[9]
ficus-indica) alginate and in combination of calcium no protection was observed at high
alginate and bovine serum albumin.[9] relative humidity.[9]
Leaf and stem of Functional food ingredients[394] Acetonitrile/acidified water extracts were A. brasiliana exhibited higher potential as A. tenella was found to be rich in vitexin
Alternanthera prepared and characterized.[394] a colorant and bioactive compounds, and its derivatives.[394] No inclusion of
brasiliana and with high betacyanin and polyphenols extracts in foods was perfomed.
Alternanthera content (89 μg/g and 35 μg/g).[394]
tenella
(Continued)
Chlorophylls (E140/141; green; lipid-soluble)
Leaves, algae Details on application in beverages, fruit juices, pasta, dairy products, soups, sweeter preparations reviewed by Martins et al. (2016).[350]
[351]
Leaves (Centella Ingredient for food and beverages Copper-chlorophylls extracts exhibited Metal-chlorophylls extracts are very Cytotoxicity of zinc- and copper-
asiatica L.) green color; zinc-chlorophylls extract stable in bread. The color of colored chlorophyll extracts was higher than
exhibited a yellow-green color. syrups and bread did not change after those of extracts from untreated/
storage for 7 d. Use of metal- steamed leaves and synthetic
chlorophylls extracts in beverage colorant.[351]
ingredient led to increased hue value
due to their structural
rearrangement.[351]
Carotenoids (E160/161; yellow-orange-red; lipid-soluble)
Roots, flowers, Details on application in butter and margarine, cakes, milk products, and soft drinks reviewed by Martins et al. (2016).[350]
leaves, algae/
microalgae,
fungus/
yeasts, and
aquatic
animals
Red pepper Colorants[395] Inclusion of dried red pepper in meat Carotenoid degradation occurred when Studies on the storage stability of red
(Capsicum products.[395] exposed to light, heat, lower pH and in pepper carotenoids as colorants in real
annuum L.) the presence of oxygen, pro-oxidants, food products are limited.[395]
catalytic metals, oxidative enzymes,
and unsaturated lipids in food.
Inclusion of paprika in meat products
showed high stability during the shelf-
life.[395]
[396]
Tomato juice Cheese The addition of tomato juice in the Tomato juice as a stable coloring agent Sensory evaluation revealed good
(Solanum process of cheese production.[396] also increased potassium, residual firmness, crumbliness.[396]
lycopersicum scavenging activity, phenolic
L.) compounds, and physical
properties.[396]
[397,398]
Marigold flowers Colorants or supplements The addition of marigold powder to Lutein degradation was observed at high In general carotenoid retention can be
extract laying hens feeds increased egg lutein temperatures, whereas pH had a minor improved by the closed storage at
(Calendula content, yolk color and altered fatty role. Colloidal encapsulation enabled lower temperature and reduced water
[397]
officinalis L.) acid profile in the yolk. the incorporation of lutein into and cationic metal activities in absence
aqueous food and beverage of light and oxygen.[395]
products.[398]
(Continued)
23
24
S. JURIĆ ET AL.
Flower Yellow pigment powder[399] Pigments extracted from the corolla of High content of natural antioxidants.[399] Data on the implementation of isolated
(Cucurbita Cucurbita moschata using acetone and pigments into real food products are
moschata hexane.[399] necessary. [399]
Duchesne ex
Poir.; corolla)
Archaea strains Food preservatives[400] Carotenoids extracted from halophilic Nanoemulsions showed a remarkable Nano- and micro-encapsulation of
Archaea. Nano- and micro-emulsions physical stability during 7 days of carotenoids improved their functional
used to encapsulate and stabilize the storage. During the storage, a small properties.[400]
carotenoids.[400] increase of the nanoemulsion diameter
size was found. Thermodynamic
stability was observed.[400]
Figure 1. Main classes of foods where natural pigments are incorporated (from Table 3).
degradation of anthocyanins can be accelerated by the presence of some components present in the
food matrix, like ascorbic acid.[353] Citric acid is also reported to cause a stronger red color degrada
tion in comparison to tartaric acid.[383] Therefore, when considering that complex food matrices are
packed with assorted compounds, it is almost impossible to predict how natural colors will behave.
Since anthocyanins are relatively unstable and highly susceptible to degradation, naturally, the first
observable change is color loss [350,402] For example, lyophilized anthocyanins recovered from grape
skins (Vitis vinifera L.), when incorporated in kefir and carbonated beverage, exhibited a measurable
loss of coloration already after 16 days of storage. Furthermore, exposure to the light during storage
resulted in an even more pronounced color loss at the end of 16 days storage.[357] In another example,
polyphenols extracted from onion wastes (Allium spp.) and used as red colorants in yogurt, exhibited
a lower stability of the color than in vitro, likely due to the lower pH.[358,360]
Moreover, it has been highlighted that a full sensory characterization should be employed when
introducing these types of extracts into real food products,[358] due to the possible negative impact on
smell and taste. Some reports reveal that when anthocyanins from purple corn cob (Zea mays L.) are
incorporated in beverages, they exhibit a change of color due to the external factors. High tempera
tures promote color shift towards yellow hues, while light exposure promotes color shift towards
red.[368] In the case of anthocyanins, decrease and loss of color occur only after prolonged storage
(6 months) in protected storage conditions, but the degradation process is significantly accelerated in
the presence of light and at elevated temperature.[368-370] Remarkably, light has a higher impact on
anthocyanin stability than elevated temperatures, and observed color changes do not correlate with the
anthocyanin content loss.[375] The stability of various anthocyanins is related to the number of
acylated groups and this is mainly affected by illumination.[362] A good example is anthocyanins
from vivid blue petals of Clitoria ternatea L. which are very sensitive and quickly degrade in the
presence of light.
In the case of betalains, their stability depends on different factors. Betalains obtained from beetroot
(Beta vulgaris L.) in cow milk underwent degradation at high temperature as a function of exposition
time.[365] Betaxanthins from red pitahaya fruits (Hylocereus polyrhizus (Weber) Britton & Rose)
exhibited higher stability in gelatin gummies than in beverages, [389] highlighting the importance of
26 S. JURIĆ ET AL.
the food matrices. Some betalains, for example those from ayrampo seeds (Opuntia soehrensii Britton
and Rose) showed higher stability than beetroot betalains in low and high-fat yogurts. It is worth to
note that the stability of these betalains was comparable with that of synthetic colorant.[391]
Regarding the hydrophobic NPs, such as chlorophylls and carotenoids, they generally degrade
when exposed to light, heat, or at lower pH conditions. They also undergo degradation in the presence
of oxygen, pro-oxidants, catalytic metals, oxidative enzymes, and unsaturated lipids in food. Copper-
chlorophylls extracts exhibit green color while zinc-chlorophylls extracts exhibit a yellow-green color.
It was found that the cytotoxicity of zinc- and copper-chlorophyll extracts were higher than of those
present in natural extracts (Centella asiatica L. leaves) from untreated/steamed leaves or synthetic
colorant.[351] Remarkably, the data on the incorporation of natural chlorophylls in food products is
scarce and this should be investigated thoroughly.
Carotenoids, instead, are often used in food products as colorants. It is common to include
carotenoid-rich red pepper (Capsicum annuum L.) in meat products. Also, carotenoid-rich products
can be produced via animals feeding. A report suggests that the addition of marigold powder
(Calendula officinalis L.) to laying hens feed resulted in an increased egg lutein content, yolk color,
and alteration of the yolk fatty acid profile. This is an interesting aspect when considering the increase
in the color potency of a food product.[397] Generally, lutein and other carotenoids are prone to
degradation at high temperatures. Interestingly the change in pH has a minor role in lutein
degradation.[398] In general, carotenoid retention can be improved by closed storage at low tempera
ture, reduced water and cationic metal activities, and absence of light and oxygen.[395] The inclusion of
natural chlorophylls and carotenoids in food products may result in color loss and degradation during
storage. Processing and storage practices may change the nature and properties of plant matrices
resulting in variations of the chlorophylls and carotenoids release. Detailed studies on the stability of
NPs in different products following well defined and standardized protocols with well-designed model
systems must be conducted to assess the potential of hydrophobic NPs as food colorants.[395]
As clearly shown by the data reported in Table 3, NPs may quickly lose their functionality when
added to foods. Because of their chemical structures, all NPs exhibit relatively low stability after
extraction from their sources, especially when exposed to light, high temperature, pH extremes,
oxygen, mechanical stresses, reactive food ingredients, during food manufacturing, preservation,
storage, and preparation, as well as to the action of digestive enzymes during digestion,[403] hence
undergoing quick degradation. NPs stabilization to fully exploit their coloring and health-beneficial
properties can be pursued through different routes, to provide improved dispersion of NPs in different
food matrices, as well as to provide a more favorable and protective environment to slow down the
NPs degradation.
Figure 2. Main types of encapsulation microstructures: single-core, multiple-core, and matrix-type microcapsules.
Stabilization of NPs
Different routes have been investigated for NPs stabilization. Some of them are inspired to those found
in nature, for example in the stabilization of anthocyanins in flowers and fruits,[404] and are generally
based on auto-association mechanisms that exploit the interaction with other molecules to form more
stable compounds,[405,406] for example through intra- and inter-molecular co-pigmentation and metal
complexation.[407]
Especially for anthocyanins, also the lipophilization technique has been recently proposed, which
consists in the increase of molecular hydrophobicity and lipid solubility, to improve dispersibility in
lipid-based foods and to reduce the rate of degradative processes, while preserving the coloring
properties, mainly through esterification reactions.[408]
The processing methods that enable the physical separation of the labile NP molecules from the
external environment, to maintain their stability and improve their shelf life, can be generally termed
as encapsulation technology. Encapsulation is widely used for different substances in solid, liquid, or
gaseous phase (the payload), to increase their stability by providing a physical barrier against the
factors present in the external environment, which might endanger their functionalities.[150,409,410]
Besides, through encapsulation, it is also possible to enable the controlled release of the payload upon
the occurrence of triggering conditions.[411] The encapsulant material and encapsulation method
should be ideally selected based on the physicochemical properties of payload (e.g. NPs), the desired
particle size, release mechanisms, and acceptable production costs.[412] Depending on the materials
and methods used for encapsulation, microparticles of different sizes, shapes, and forms can be
obtained.[413] Encapsulation systems of interest for natural pigments can be classified into three
main types, depending on their microstructure, as shown in Fig. 2: 1. Single-core particles, character
ized by a well-identifiable shell around the core (payload); 2. Multiple-core particles, with more than
one core embedded in the shell material; 3. Matrix-type particles, with the payload materials homo
geneously dispersed in the encapsulant material. More complex structures, developed for enabling
controlled or triggered release, [413] are usually not needed for pigment encapsulation.
The most commonly used encapsulation techniques are based on (i) physical methods, such as
spray drying, lyophilization, supercritical fluid precipitation, solvent evaporation; (ii) physicochemical
methods, such as coacervation, ionic gelation, and encapsulation in liposomes; (iii) chemical methods,
including interfacial polymerization and molecular inclusion complexation.[414]
The main discriminant for the selection of the most suitable formulation for the encapsulation
system and technique depends on the solubility in water of NPs.[414] For example, water-soluble NPs,
such as anthocyanins, betalains, anthoxanthins, organic acids, polyphenols, and tannins, require that
a certain degree of chemical interaction is developed with the encapsulant material to prevent the
undesired release in the aqueous part of foods. In contrast, lipid-soluble components, such as
carotenoids, curcumin, and chlorophylls, can be encapsulated by purely physical encapsulation
processes.[414,415] It must be remarked that the encapsulated NPs are not specifically authorized by
the main regulatory bodies, which instead provide general guidelines for dealing with the intrinsic
health risk associated with the consumption of engineered particulate food ingredients.[409,416] For
example, the European Union covers the risks associated with the use of nanotechnologies in the food,
either through the general food law (Regulation (EC) No 178/2002) or through specific approval
processes, with a specific focus on manufactured materials, whose 50% of their composition exhibits
one or more dimensions in the nanoscale range (1–100 nm), through the Regulation (EU) No 1363/
2013.[416] The US FDA approach is to retain manufacturers responsible for ensuring that their
products satisfy applicable safety standards, also by engaging the appropriate government agencies
(e.g. the Emerging Technologies Interagency Policy Coordination Committee) and consulting the
appropriate guidance documents (https://www.fda.gov/science-research/nanotechnology-programs-
fda/nanotechnology-guidance-documents) regarding the regulatory status, or safety of products that
involve the application of nanotechnology.[417]
28 S. JURIĆ ET AL.
Table 4. Stabilization techniques and materials used for the protection of water and lipid-soluble NPs.
Pigment Co-pigment/Wall materials Stabilization technique Reference
WATER SOLUBLE
[418–420]
Anthocyanins Dairy proteins Complexation
[421]
Anthocyanins Pectins Complexation
[422]
Anthocyanins Whey proteins and pectins Complexation/physical entrapment
[423,424]
Anthocyanins Gum arabic Complexation
[425,426]
Anthocyanins β-cyclodextrins Molecular inclusion
[364]
Anthocyanins Green tea extracts Complexation
[427]
Anthocyanins Ferric ion Chelation
[428]
Anthocyanins Stearic acid Lipophilization
[429]
Anthocyanins Oleic acids Lipophilization
[430–433]
Anthocyanins Different fatty acids Lipophilization
[434]
Anthocyanins Montmorillonite Hybridization
[435]
Anthocyanins Methoxyl pectin Ionic gelation
[436]
Anthocyanins Sodium alginate Ionic gelation
[437]
Anthocyanins Polyethylene glycol (PEG) Ionic gelation
[438]
Anthocyanins Alginate Ionic gelation
[14]
Anthocyanins Pectin amide Ionic gelation
[14]
Anthocyanins Whey protein isolate Microemulsions
[14]
Anthocyanins Maltodextrin, pectin amide Spray drying
[439]
Anthocyanins Glycerol mono-oleate, soy Lecithin, maltodextrin, poloxamer Spray drying
338
[440]
Anthocyanins Supercritical carbon dioxide Liposomes
[441]
Betalains Sucrose Co-crystallization
[9]
Betalains Sodium alginate, sodium alginate-bovine serum Ionic gelation
[442]
Betalains Rapeseed oil, guar gum, xanthan gum Double emulsions
LIPID SOLUBLE
[443]
Carotenoids Whey protein concentrate, gum arabic Spray drying
[444]
β-carotene Wheat gluten nanoparticles, wheat gluten nanoparticle- Pickering emulsion
xanthan gum
[445]
β-carotene Maltodextrin, gum arabic, chitosan, gelatin Spray drying
[446]
β-carotene Native and hydrolyzed Pinhao starches Freeze-drying
[447]
Lycopene Gelatin, sucrose Spray drying
[448]
Lycopene Lecithin, α-tocopherol Supercritical antisolvent co-
precipitation (SAS)
[449]
Lutein Gelatin, gum arabic Coacervation
[450]
Chlorophylls Polycaprolactone Microfluidic emulsification
[451]
Chlorophylls Gum arabic, maltodextrin Spray drying
[452]
Chlorophylls Whey protein isolate Spray drying
Recent literature data on the different strategies adopted for protecting water- and lipid-soluble
NPs, with details on materials used, are presented in Table 4.
Stabilization of water-soluble pigments

Anthocyanins are water-soluble pigments that are regularly used in the food industry.[453] However,
the introduction of anthocyanins into food products is often severely challenged by their low stability
to environmental factors experienced during product manufacturing, storage, and preparation.
Anthocyanins are particularly susceptible to the presence of light, basic pH, temperatures higher
than 60-80°C, the presence of sulfite, ascorbic acid, and enzymes.[454]
Co-pigmentation of anthocyanins, through the complexation with other molecules, is often a simple
and effective approach to improve anthocyanins’ stability.[455] For example, anthocyanins from purple
carrot were stabilized by complexation with macromolecules, such as whey protein isolates and different
pectins, and tested under accelerated conditions (40°C, and in presence of ascorbic acid). A significant
reduction of the degradative processes leading to the decrease of pigment absorbance was observed
especially for whey proteins, whereas, in the case of pectins, a slight cloud color was recorded.[418]
Spontaneous complexation was reported between bovine β-lactoglobulin or α- and β-casein and malvi
din-3-O-glucoside at pH 6.3,[419,420] to enhance thermal, oxidation, and photo-stability of grape skin
extracts. A more advanced complexation process, which was applied for efficiently binding pelargonidin to
whey proteins and beet pectins, also through a thermal treatment to form anionic nanoparticles, did not
significantly improved anthocyanins preservation against degradation induced by ascorbic acid, and
exhibited physical stability only in a limited pH range.[422] The effect on pH appears to be a critical factor
in determining the efficiency and stability of anthocyanin complexation to polymers. Therefore, some
authors suggested using different types of pectins as a function of the desired pH of use.[421] The complexes
formed between polymers and anthocyanins can be ascribed to different reversible intermolecular inter
actions, including hydrophobic and hydrophilic bonding, van der Waals forces, and hydrogen bonding,
whose stability significantly depends on pH and temperature.[455]
Other biopolymers, such as gums, can also form spontaneous complexes with anthocyanins. For
example, gum arabic was efficient in stabilizing the color of anthocyanins in the presence of ascorbic
acid,[423] and upon intense thermal treatments.[424] Despite the higher cost and the specific preparation
conditions required, cyclodextrins may form stable complexes with anthocyanins, contributing to preser
ving their stability over time. In particular, the addition of β-cyclodextrins resulted in a successful strategy to
preserve the color of chokeberry juice, especially at pH 3.6, by efficiently binding to anthocyanins.[425]
However, β-cyclodextrins were observed to exhibit a decrease in anthocyanin color intensity.[426] Therefore,
to date, cyclodextrins do not appear to be the most suitable stabilizing agents for anthocyanins.[455]
Molecular complexes are reported to be spontaneously formed between anthocyanins and poly
phenols, such as vanillin, epigallocatechin gallate, green tea extract, and protocatechualdehyde.
Especially the addition of green tea polyphenols to purple carrot anthocyanin, through the develop
ment of hydrophobic interactions, significantly delayed color fading in the presence of ascorbic acid
and under accelerated aging (40°C for 7 days in the presence of light).[364] Other phenolic compounds
successfully tested as co-pigments to form complexes with anthocyanins include flavonoid C-
glycoside,[456] rutin,[457] chlorogenic acid,[458] gallic acid,[459] quercetagetin,,[460] phenolic-rich green
coffee bean extracts.[461]
Metallic ions, especially from tin, copper, iron, aluminum, magnesium, and potassium, have also been
reported to stabilize anthocyanin extracts, through metal chelation.[455,462] In general, metal chelation
complexes are extremely susceptible to the environmental conditions (e.g. pH, temperature, solvent),[462]
and may become unstable in the presence of other food ingredients.[427] However, metal chelation may play
an important role in products derived from fruit, such as juices and wines, where the addition of metal ions
may become part of product formulation for enhancing product quality.[462]
Recently, it was shown that, for the incorporation in lipid-rich matrices, the lipophilicity of anthocyanins
can be increased through the lipophilization technique, which consists in the esterification or acylation of
a lipophilic moiety, such as a fatty acid or fatty alcohol, on phenolic substrates, to obtain new molecules with
a tailored hydrophilic/lipophilic balance,[463] and increased hydrophobicity and lipid solubility.[464] In the
case of anthocyanins, lipophilization can be obtained through enzymatic reactions, forming pigments,
which, because of their increased solubility in organic solvents,[428] can be incorporated into emulsified or
lipid-based media,[465] and exhibit more stable coloring properties for pH variations, high temperature, and
the presence of oxidizing agents and other compounds.[408,429,466] The lipophilization of malvidin-
3-O-glucoside to increase its hydrophobicity was reported through the formation of stearoyl mono- and
diesters,[428] the esterification with oleic acid,[429] as well as the conjugation with fatty acids of different chain
lengths.[430] Blueberry anthocyanins were subjected to lipophilization products with the free fatty acids
present crude camellia oil, directly in the oil,[431] blackcurrant skin extract, rich in different anthocyanins,
were acylated with octanoic acid,[432] or lauric acid.[433] In general, the structural modification deriving from
lipophilization of malvidin-3-O-glucoside did not affect its antioxidant potential[429;] however, the effect on
color properties was reported to depend on the number and site of the involved lipophilization
reactions.[408]
The observation of the long-term stability of “Maya Blue”, which is obtained from the mixture of
indigo pigment and palygorskite clay, has stimulated the interest towards the development of
hybridized pigments, composed of dyes adsorbed in nanoporous solids.[467] The immobilization of
pigments in solids, such as mesoporous silica, zeolites, layered double hydroxides, and smectite, was
30 S. JURIĆ ET AL.
demonstrated to enable a certain degree of control over the photochemical and photophysical proper
ties of dyes, depending on the host-guest interactions.[467] For example, indigo blue was hybridized
with different zeolitic matrices, with the key parameters resulting to be the match between the pore
size with the dimensions of the guest molecule.[468] This approach is not always ideal for food
applications but is suitable for environmentally-friendly applications. For example, the intercalation
of anthocyanins into montmorillonite ensured that the original color was maintained at room
temperature under basic condition (pH = 11), and exhibited increased stability upon light irradiation,
because of the host-guest interaction and steric protection from the attack of atmospheric oxygen.[434]
Among the encapsulation techniques, the ionic gelation method was reported by several authors to
be specifically suitable for anthocyanins.[435-437] Ionic gelation, especially if coupled with
a solidification process, such as atomization or electrostatic spraying, or coextrusion/extrusion drip
ping, is a simple, efficient and low-cost technique that can be applied for both hydrophilic and
hydrophobic compounds,[435,469] with good encapsulation efficiency and low polydispersity.[436] It is
based on the cross-linkage of polyelectrolytes, such as chitosan, gelatin, and whey protein concentrate,
frequently used because of their high efficiency in protecting pigments during storage, with multi
valent ions, such as Ca2+, Ba2+, and Al3+.[414]
Spray drying is an encapsulation process that has been widely used for anthocyanins, because it is cost-
effective and simple, and can be adapted to different needs in terms of coloration and food compatibility, by
selecting the most appropriate biopolymer as encapsulant material, which can be of hydrophilic or
hydrophobic nature. Depending on the properties of the encapsulant material and payload compounds,
on the loading ratio, and on the atomization and drying conditions, core-shell or matrix-type microstruc
tures can be obtained. Interestingly, despite the generally high temperatures of the drying medium (usually
hot air), spray-drying is suitable also for thermolabile materials, such as anthocyanins, because of the very
short residence time in the drying chamber. In advanced spray dryers, the viscosity of the solutions to be
sprayed can be as high as 300 mPa·s, with great advantages in terms of energy savings and reduced
processing time.[470]
Interestingly, tuning of ingredients and fabrication method enables the development of micro
capsules with advanced functionality. For example, thermoresponsive microcapsules, made by an
aqueous core and a two-layer shell structure, the first consisting of glycerol mono-oleate and soy
lecithin and the second of maltodextrin and poloxamer 338, were successfully used to encapsulate
a heat-sensitive colorant (anthocyanins from black carrot), with high stability, higher color intensity in
a simulated beverage, and better stability during pasteurization than a non-encapsulated colorant.[439]
Encapsulation, comparatively investigated for ionic gelation, emulsion gelation, and spray drying, can
also significantly contribute to stabilize anthocyanins in the gastrointestinal tract in comparison with
non-encapsulated anthocyanins and regulate the release, as shown for bilberry extracts.[14]
Despite spray drying being commonly used, it presents some disadvantages, mainly related to the
thermal degradation of pigments if atomization and drying conditions are not adequately controlled.
Furthermore, there is the need for removal of large amounts of solvents, which is not desirable, when
the intended application is in liquid food products. Finally, a wide particle size distribution and large
mean size values are produced, which, coupled with a heterogeneous and diverse morphology of the
particles, might affect the bioavailability of the payload and cause the loss of finer particles in exhaust
air. Therefore, alternative encapsulation methods are intensively searched. Recently, the encapsulation
of anthocyanins was investigated in liposomes, produced in supercritical carbon dioxide (SC-CO2),
with the advantage of a sub-micrometric size and narrow size distribution, good encapsulation
efficiency in the inner aqueous compartment of liposomes, and good protection in simulated gastric
fluid, accompanied by a sustained release in simulated intestinal fluids.[440]
Betalains are water-soluble yellow, red or violet NPs with good antioxidant properties. Their
stability is strongly affected by high temperature, basic or very acidic pH, light, oxygen, and high-
water activity.[442] Betalains were successfully encapsulated and protected by ionic gelation, with
higher stability observed especially at low relative humidity.[9] Good encapsulation efficiency and
better stability of betalain were obtained by using a double emulsion (water-in-oil-in-water or w/o/w
emulsions[471]), with the betalains dispersed in the inner aqueous phase.[442] Remarkably, w/o/w
emulsion structure act as a barrier to protect the hydrophilic bioactives embedded in the inner
aqueous phase within the food product, but which enables the payload to be gradually released
when oil digestion takes place (e.g. intestinal digestion). The influence of the matrix and encapsulation
technique on the stability of betalains have been summarized in a recent systematic review.[96] The
matrix has an important influence on the stability of betalain. Polysaccharides, such as maltodextrin,
different combinations of other polysaccharides, or protein-polysaccharide complexes, show great
potential for increasing the stability of betalain. Recently, Čakarević et al. encapsulated a beetroot juice,
rich in betalain, using pumpkin protein isolates via freeze- and spray-drying techniques. This
encapsulation system enabled a gradual release of bioactive compounds during simulated
digestion.[472] The co-crystallization of betacyanins from Basella rubra L. with sucrose resulted in
better pigment handling and better retention of the pigments.[441]
Stabilization of lipid-soluble pigments

Chlorophylls are lipid-soluble NPs, which are widely distributed in green fruits and vegetables and are
characterized by brilliant green color and significant biological activity. However, their commercial-
scale application is limited because, under unfavorable conditions, degradation, and discoloration of
chlorophylls rapidly occur. A historical approach to chlorophyll stabilization consisted in the forma
tion of complexes with zinc or copper ions to form the more stable green metallo-chlorophylls.[473,474]
However, the need for concentrations of Zn+2 or Cu+2 salts above the limit concentrations established
by US FDA promoted the use of physical encapsulation for chlorophylls stabilization.[102] Spray drying
was used to enhance chlorophylls storage stability at different temperatures (4, 20, 40°C) using
different combinations of gum arabic and maltodextrins.[451] Remarkably, the use of maltodextrins
alone as wall material resulted to be optimal to obtain the highest storage stability (95-98%), because of
the minimized moisture uptake and maximized encapsulation efficiency. Researchers also reported
that chlorophyll water-solubility was increased via encapsulation into nanoscale liposomes.[475] Hsiao
et al. proposed a new polymer encapsulation technology for enhancing chlorophyll stabilization using
polycaprolactone.[450] Microfluidic emulsification enables the encapsulation of chlorophyll into
microparticles with uniformly distributed particle sizes. It was reported that chlorophylls, encapsu
lated with polycaprolactone, were protected against white light degradation, thus improving their
stability. The size of the particles can be controlled during the preparation process to ensure a high
encapsulation rate for chlorophylls, as well as convenient storage and transportation in solid-state.[450]
Whey protein isolates are promising in successful encapsulation of chlorophylls because they can
contribute to improving water solubility and biological stability of chlorophylls. Spray-dried micro
particles, obtained at a high ratio of whey protein isolate to chlorophylls, resulted in lower moisture
content, higher encapsulation efficiency, and greater chlorophyll stability. The pyrrole ring of chlor
ophyll binds to the hydrophobic residues of whey protein isolate, changing the crystalline structure of
chlorophyll, probably because of the formation of soluble complexes. These types of microparticles
may be used as a natural food additive because of good antioxidant activity and physicochemical
properties.[452]
Carotenoids, such as β-carotene, lycopene, and lutein, are another class of lipid-soluble NPs, which
are extremely susceptible to adverse environmental conditions, such as light, high temperatures, pH
extremes, and oxidation. The most common method of encapsulation of carotenoids is spray drying,
with many authors that investigated the optimal conditions, in terms of inlet temperature of drying air,
feeding rate, and the ratio between payload and wall material, to promote the retention of carotenoids
and their antioxidant capacity.[447,476,477]
β-carotene is widely used because of its high vitamin A activity and strong antioxidant capacity.
However, being extremely sensitive to oxidation, high temperature, light, and low pH,[104] it requires
proper spray-drying conditions and adequate selection of wall materials. For example, the combina
tion of chitosan, gelatin, and maltodextrin with gum arabic resulted to be effective in preserving β-
32 S. JURIĆ ET AL.
carotene during spray drying in microcapsules, which prevents its degradation during storage, with
the best performance observed for gelatin.[445] β-carotene was encapsulated also in wheat gluten
nanoparticle-xanthan gum-stabilized Pickering emulsions, with a positive effect not only on enhan
cing stability but also on β-carotene bioaccessibility.[444] Freeze-drying was used in the preparation of
β-carotene microcapsules, made of native or hydrolyzed starch, which significantly increased β-
carotene stability against ultraviolet light (especially hydrolyzed starch) concerning non-
encapsulated β-carotene.[446]
Lycopene is an acyclic carotenoid and the presence of unsaturated bonds in its molecular structure
makes it susceptible to degradation during storage, mainly in the presence of oxygen.[478]
Encapsulation through spray drying was tested to protect lycopene from chemical damage, using
gelatin and sucrose as wall materials.[447] Similarly to what was discussed for β-carotene, the wall
material composition (ratio of gelatin/sucrose and core/wall material), the feed temperature, as well as
the air inlet temperature represented the key process parameters to control encapsulation efficiency
and lycopene retention. Moreover, given the low water solubility of lycopene, its emulsification by
high-pressure homogenization prior to drying also represented a key element to consider in designing
the encapsulation process.[447] The supercritical antisolvent co-precipitation (SAS), using supercritical
CO2 as an antisolvent, enabled to obtain nano- or microparticles, containing lycopene in an α-
tocopherol and lecithin matrix, characterized by an increased encapsulation efficiency and stabiliza
tion of lycopene in comparison with non-encapsulated lycopene.[448]
Lutein, which is classified as a xanthophyll or oxygenated carotenoid, is found in high quantities in
green leafy vegetables. Lutein, besides being a natural colorant, of wide use in the food industry,
exhibits also various biologically significant activities, such as prevention or amelioration of the effects
of degenerative human diseases. However, lutein is highly sensitive to pH, light, and oxygen, and high
temperature.[479] The coacervation method was successfully used to improve the stability of lutein.[449]
Coacervation is based on liquid-liquid phase separation and requires either a single polymer or
a mixture of two oppositely charged polymers.[414] In this method, the wall material is preliminarily
Table 5. Examples of application of stabilized NPs in real food products.

Encapsulated Functional
compounds Compound donor Stabilization method Material food References
[8]
Anthocyanin Barberry (Berberis vulgaris) Spray drying Gum arabic, maltodextrin, Jelly
gelatin
[379]
Anthocyanin Black bean (Phaseolus vulgaris Molecular inclusion β-cyclodextrin Sport
L) coat beverage
[481]
Anthocyanin Grape (Vitis vinifera L.) skin Spray drying Maltodextrin Apple
puree
[482]
Anthocyanin Sour cherry (Prunus cerasus Freeze-drying Whey and soy proteins Cookies
L.) pomace extract
[483]
Betalains Barbary fig (Opuntia ficus- Ionic gelation Calcium alginate/Gelatin Gummy
indica L.) candy
[484]
Betalains Barbary fig (Opuntia ficus- Spray drying Soluble fiber [(1–3)(1-4)- Extruded
indica L.) β-D-glucan cereal
[485]
Betalains Beetroot (Beta vulgaris L.) Freeze-drying Maltodextrin Chewing
gum
[385]
Betalains Red beet (Beta vulgaris L.) Thin-film hydration- Lecithin liposome Gummy
(Betanin) extract diluted with dextrin sonication candy
technique
[486]
Carotenoids Yellow bell pepper (Capsicum Ultrasonic β-cyclodextrin Isotonic
(yellow annuum L.) homogenization, beverage
pigments) kneading
[485]
Carotenoids Saffron (Crocus sativus Freeze-drying Maltodextrin Chewing
(crocins) L.) gum
[487]
Chlorophylls Alfalfa (Medicago sativa Emulsification + Canola oil, glycerol Gummy
L.) Freeze-drying monostearate, gelatin, candy
agar
dissolved in water, and subsequently, its deposition on the surface of the finely dispersed core material
by electrostatic coagulation is triggered by the change of the environmental conditions (pH, tempera
ture or addition of salt) to induce the separation of a polymer-rich phase.[480] Lutein, encapsulated in
coacervates, presented improved stability against light, humidity, and high temperature, when the
concentration of wall material, the ratio of core to wall material, and pH were optimized.[449]
Application of stabilized NPs in real foods

The observed increasing trend, in recent years, towards NPs use as alternatives to synthetic colors in
food applications derives both from legislative actions and consumer concerns for health.[8]
Encapsulation seems to be an efficient way to protect and stabilize NPs. However, available literature
data about the application of encapsulated NPs in food products as coloring agents are still scarce, as
shown in Table 5, especially if compared with the considerable number of papers on encapsulation
methods and fabrication procedures of pigments microparticles and their characterization in vitro.
Nevertheless, a better understanding of the issues related to the inclusion of encapsulated NPs into
different food matrices, with the complexities associated to the heterogeneous and composite for
mulation and structure of food, is fundamental to enable the prediction of NPs behavior in real foods,
for what concerns both their coloring capability and eventual health-beneficial properties.
In complex food systems, the behavior of NPs is different from model systems, especially for what
concerns their stability, which is considerably affected by a faster degradation occurring due to the
physicochemical stress factors induced by food manufacturing, preservation, storage, and preparation,
by the interaction with other food ingredients, as well as the absorption at the multiple interfaces (e.g.
between aqueous and lipid or solid phases) present in the food. Moreover, diffusion of NPs in real
foods is also affected by several factors, e.g. composition (proteins, carbohydrates, fatty acids), pH,
water activity, packaging material, and the presence of trace metals. Finally, the use in foods of NPs, at
the concentration level required to develop the desired color intensity and hue, might also develop
unacceptable changes in the product organoleptic quality. For example, sour cherry (Prunus cerasus
L.) pomace extracts, encapsulated in whey and soy proteins, when incorporated in cookies, had
significant effects, well beyond the coloring effect. In fact, besides conferring a red tone, due to the
presence of a high concentration of anthocyanins, the encapsulated extracts had a positive effect also in
increasing polyphenols content and antioxidant activity, as well as storage stability of cookies, and
resulted in an intense smell and taste, which was associated with fresh cherries and elicited positive
reactions during sensory tests.[482] However, the addition of the encapsulates had a negative effect on
the shape, size, and uniformity of cookies. Protein-based particles, indeed, might induce significant
changes in the structure of cookies, mainly because proteins absorb water during cookies preparation,
resulting in cookies with increased hardness.[488] Moreover, the presence of an additional particulate
phase can increase the fragility of cookies, hindering the formation of a continuous starch network in
the dough.[489]
The selection of the encapsulant material is important both in terms of compatibility of the
encapsulated NPs with the final product and of coloring function and stability over storage time.
For example, anthocyanins, encapsulated by spray drying with wall materials based on different
combinations of maltodextrin and gum arabic, enabled to tune coloring efficiency and stability of
anthocyanins under the tested storage conditions (different temperatures and relative humidity,
illumination for 90 days), with significant improvements with respect to non-encapsulated anthocya
nins. Moreover, the jellies, where the encapsulated anthocyanins were added, exhibited better sensory
attributes and physicochemical properties, than jellies prepared with artificial coloring.[8] In general, as
shown in the other examples reported in Table 5, the major problem of the application of NPs in real
food products is their instability, due to the multiple factors discussed above. For example, in sport or
isotonic beverages, low pH values and the presence of ascorbic acids represent significant challenges to
the stability of NPs, such as anthocyanins and carotenoids, which have been faced using inclusion in β-
cyclodextrin.[379,486] Apparently, the incorporation of encapsulated NPs is less difficult in gummy
34 S. JURIĆ ET AL.
candies,[385,483,487] chewing gums,[485] and extruded cereals.[484] Encapsulation should ideally not only
contribute to overcoming any instability issue that might reduce the coloring efficiency but also
making their incorporation in foods easier. More specifically, enhancing food compatibility and,
most importantly, limiting the impact on the sensory attributes, if not ameliorating the product
organoleptic profile, are key aspects of the design of NPs’ encapsulation systems. Therefore, sensory
evaluation should always be associated with any research addressing the development of novel
formulations for NPs capsules to be used in real food products.
Conclusions and remarks

Natural pigments (NPs) respond to the increasing consumers’ demand for safer and more natural
food, with the goal, in the future, to completely replace synthetic colorants. However, to date, the
commercial development of NPs is still challenged by multiple factors, such as the low range of natural
colors approved for food use and the laborious procedure for regulatory approval of novel colorants,
the higher cost in comparison to synthetic colorants (from 5 to 20 times more expensive[5]), and the
large volumes of biomass required for extraction per unit mass of natural colorant. Moreover, NPs
usually exhibit weaker tinctorial strength than their synthetic counterparts, requiring higher dosages
to obtain the desired hues, and are generally more sensitive to environmental factors, such as light, pH,
UV, temperature, oxygen, and heat than synthetic colorants of well-established use. Finally, depending
on the source, deodorization of NPs might be needed to avoid an undesired effect in the food and,
similarly, the interaction with food components should be minimized to limit the development of off-
flavors and the alteration of the organoleptic profile.
Therefore, the discovery of novel NPs and the introduction of unusual sources is not sufficient for
their successful commercial application. More efficient production processes are needed to increase
the extraction yields and reduce the extraction times from biomass, or the introduction of more
efficient and less resource-intensive sources of NPs, such as microbial sources, which can be scaled-up
and are more readily manipulated than plants or insects.
Additionally, future research on NPs should also focus on widening the range of hues that can be
obtained, and on promoting the pigments with associated health-beneficial properties. More research
is also needed on the stabilization of natural colorants, which, to date, has been addressed through
different approaches based on molecular complexation or microencapsulation. However, only scarce
data, often contradicting the in vitro results, are currently available on the stability of NPs in real food
systems.
Declaration of interest statement

The authors declare no conflict of interest.
ORCID
Marko Vinceković http://orcid.org/0000-0001-6158-3914
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Food Reviews International

ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/lfri20

Sources, stability, encapsulation and application of

Slaven Jurić , Marina Jurić , Żaneta Król-Kilińska , Kristina Vlahoviček-

To link to this article: https://doi.org/10.1080/87559129.2020.1837862

Published online: 28 Oct 2020.

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Sources, stability, encapsulation and application of natural

Classification, stability, and sources of NPs

Table 1. Classification of NPs, main features, sources, and extraction processes.

Potential alternative sources of NPs

fields,[60,163-166] mechano-chemical methods,[167] and high-pressure homogenization,[131,168,169]

Health-promoting properties of NPs

NPs as food colorants/additives in foods

g FW).[366] over 6-month storage; more yellow at

during processing and/or storage.[365]

Stabilization of water-soluble pigments

Stabilization of lipid-soluble pigments

Table 5. Examples of application of stabilized NPs in real food products.

Application of stabilized NPs in real foods

Conclusions and remarks

Declaration of interest statement

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