Exploring Apple Pectic Polysaccharides - Extraction, Characterization, and Biological Activities - A Comprehensive Review

International Journal of Biological Macromolecules 255 (2024) 128011
Contents lists available at ScienceDirect
International Journal of Biological Macromolecules

journal homepage: www.elsevier.com/locate/ijbiomac
Review
Exploring apple pectic polysaccharides: Extraction, characterization, and

biological activities - A comprehensive review
Neeraj Kumari a, Manoj Kumar b, *, Radha a, Nadeem Rais c, Sunil Puri a, Kanika Sharma b,
Suman Natta d, Sangram Dhumal e, Rahul D. Damale f, Sunil Kumar g, Marisennayya Senapathy h,
Sheetal Vishal Deshmukh i, T. Anitha j, T. Prabhu k, S. Shenbagavalli l, V. Balamurugan m,
Jose M. Lorenzo n, John F. Kennedy o
a
School of Biological and Environmental Sciences, Shoolini University of Biotechnology and Management Sciences, Solan 173229, India
b
Chemical and Biochemical Processing Division, ICAR-Central Institute for Research on Cotton Technology, Mumbai, 400019, India
c
Department of Pharmacy, Bhagwant University, Ajmer, Rajasthan 305004, India
d
ICAR—National Research Centre for Orchids, Pakyong 737106, India
e
Division of Horticulture, RCSM College of Agriculture, Kolhapur 416004, India
f
ICAR—National Research Centre on Pomegranate, Solapur 413255, India
g
Indian Institute of Farming Systems Research, Modipuram 250110, India
h
Department of Rural Development and Agricultural Extension, College of Agriculture, Wolaita Sodo University, Wolaita Sodo, Ethiopia
i
Bharati Vidyapeeth (Deemed to be University), Yashwantrao Mohite Institute of Management, Karad, India
j
Department of Postharvest Technology, Horticultural College and Research Institute, Tamil Nadu Agricultural University, Periyakulam 625604, India
k
Department of Spices and Plantation Crops, Horticultural College and Research Institute, Tamil Nadu Agricultural University, Periyakulam 625604, India
l
Department of Natural Resource and Management, Horticultural College and Research Institute, Tamil Nadu Agricultural University, Periyakulam 625604, India
m
Department of Agricultural Economics, Agricultural College and Research Institute, Tamil Nadu Agricultural University, Madurai, India
n
Centro Tecnológico de la Carne de Galicia, Parque Tecnológico de Galicia, Avd. Galicia n◦ 4, San Cibrao das Viñas, 32900 Ourense, Spain
o
Chembiotech Laboratories, Advanced Science and Technology Institute, Kyrewood House, Tenbury Wells, Worcs WR15 8FF, UK
A R T I C L E I N F O A B S T R A C T
Keywords: Apple (Malus domestica) is a popular and ancient fruit of the Myrtaceae family. Apple fruit is well-known for its
Apple great nutritional and phytochemical content consisted of beneficial compounds such as polyphenols, poly
Pectin saccharides, sterols, and organic acids. Polysaccharides extracted from different parts of the apple fruit, including
Bioactivities
the peel, pomace, or the whole fruit, have been extensively studied. Researchers have investigated the structural
Value-added products
characteristics of these polysaccharides, such as molecular weight, type of monosaccharide unit, type of linkage
Polysaccharides
and its position and arrangement. Besides this, functional properties and physicochemical and of apple poly
saccharides have also been studied, along with the effects of extraction procedures, storage, and processing on
cell wall polysaccharides. Various extraction techniques, including hot water extraction, enzymatic extraction,
and solvent-assisted extraction, have been studied. From the findings, it was evident that apple polysaccharides
are mainly composed of (1 → 3), (1 → 6): α-β-glycosidic linkage. Moreover, the apple polysaccharides were
demonstrated to exhibit antioxidant, hepatoprotective, anti-cancer, hypoilipidemic, and enzyme inhibitory
properties in vitro and in vivo. The potential applications of apple polysaccharides in the food, cosmetic,
pharmaceutical, nutraceutical industries have also been explored in the present review. Overall, the research on
apple polysaccharides highlights their significant potential as a source of biologically active compounds with
various health benefits and practical applications.
1. Introduction increasingly frequent extreme climatic shifts, driven by both natural

occurrences and human-induced or technological catastrophes, result
From a health standpoint, it’s clear that the planet is undergoing ing in a range of health disparities [1–6]. Therefore, nowadays, to
* Corresponding author.
E-mail address: [email protected] (M. Kumar).
https://doi.org/10.1016/j.ijbiomac.2023.128011
Received 26 August 2023; Received in revised form 6 October 2023; Accepted 8 November 2023
Available online 10 November 2023
0141-8130/© 2023 Published by Elsevier B.V.
N. Kumari et al. International Journal of Biological Macromolecules 255 (2024) 128011
promote sustainability, people are opting to incorporate natural prod major monosaccharide units of apple polysaccharides. Earlier, it has
ucts like vegetables and fruits in their diet because of their positive as been reported that the biological properties of polysaccharides are
sociation of bioactive compounds and their health benefits (i.e., strongly influenced by their monosaccharide composition and content.
reducing the incidence of cancer, chronic disorder and oxidative stress) The presence of side chains also influences the interactions of poly
[7–9]. Apple (Malus domestica Borkh., member of Myrtaceae) is the 4th saccharides with other bioactive molecules. For example, arabinan side
most devoured fresh fruit, grown in >93 countries including USA, chains in pectic polysaccharides affect their interactions with poly
China, Turkey, Poland, India, Chile, Italy, Russian Federation, Iran and phenols (phenolic acids and oligo or polymeric flavanols). Linear ara
France [10]. According to a recent report, the global annual apple binans showed higher interactions with polyphenols compared to
production stands at an impressive 70 million tons, with these apples branched arabinans, but those with covalently linked polyphenols
being consumed directly or processed into various food products such as exhibited lower interactions. It suggests that overall polysaccharide
juice, jams, jellies, beverages, cider, and candies [11,12]. Out of this structure plays a more significant role in polyphenol interactions than
total volume, value-added apple products like juice, cider, jam, and each individual component [24,25]. Additionally, polysaccharides with
dried products make up around 25–30 %, leaving behind a significant high levels of uronic acid have been found to possess increased bioac
amount of pomace biowaste, which can account for up to 25 % of the tivity due to modifications in their properties, such as improved solu
original fresh fruit weight [13]. Fortunately, this apple pomace holds bility. This suggests that polysaccharides derived from apples may have
great potential and value as it contains high concentrations of attractive significant potential as a source of biologically active compounds [26].
biomolecules such as carbohydrates, polyphenols, and triterpenes Therefore, it is important to investigate the structural characterization
[14–16]. One particularly interesting molecule present in vegetable cell and function of these polysaccharides to determine their potential health
walls is pectin, which can be effectively recovered from apple pomace benefits for humans.
and other vegetable biomass sources [17]. Apples, being a major source In recent years, there has been a growing attention towards inves
of pectin polysaccharides, are used as base in the production of ethanol, tigating the bioactivities of polysaccharides derived from various plant
animal feed, and organic fertilizers [18]. Pectin can be divided into three sources across different fields of research [27]. For example, poly
distinct regions, namely rhamnogalacturonan I (RG-I), (1 → 4)-linked saccharides extracted from apple have been studied for their hep
α-D-galacturonic acid (GalA), and rhamnogalacturonan II (RG-II) units atoprotective, antioxidant, anticancer and certain enzymatic effects
(Fig. 1). RG-I is made up of repeating disaccharide units composed of [28–32]. Furthermore, only limited research has been conducted on the
galacturonic acid and rhamnose that are linked as [→4)-α-D-GalpA-(1 various analytical methods for extracting pectin polysaccharides from
→ 2)-α-L-Rhap-(1→], as well as side chains of neutral sugar [19,20]. apples. Methods like hot-water, solvent-assisted, acidic or enzymatic
GalA is a sugar acid whereas RG-II is a complex glycan made up of a extraction techniques are some of the commonly used extraction
large number of sugar molecules linked to each other via glycosidic methods of apple polysaccharides [33–39]. In addition, from the find
bonds [21]. Furthermore, various analytical methods have determined ings of various studies it is elucidated that extraction processes including
the presence of cellulose and hemicellulose along with structural pro factors such as duration, temperature and other relevant parameters
teins in the cell wall of apple fruit. It was estimated that polysaccharide have significant impact on the monosaccharide yield, composition and
composition of fruit flesh or pomace varies with developmental stages, structural and functional properties of the polysaccharides [4–6,40,41].
genetics and growth conditions [22,23]. Moreover, it was analyzed that As aforementioned, apple polysaccharides have potential to be used for
galactose, arabinose, rhamnose, galacturonic acid and xylose are the its health benefiting activities [42]. In addition, apple polysaccharides
Fig. 1. Schematic representation of Pectin structure: includes different regions, the dominant smooth pectin region consists of a linear, partially esterified homo
galacturonan (HG) backbone, making up to 60 % of the total pectin. The hairy regions are made up of highly branched rhamnogalacturonan I (RG-I) and rham
nogalacturonan II (RG-II) domains, with smaller contributions from xylogalacturonan (XG) and apiogalacturonan (AG) (Source: [46]).
2
may be used for its value-added opportunities as food ingredient, (GC–MS) confirmed that the main monosaccharide units of the apple
functional food, cosmetics, pharmaceuticals [18,43,44]. These plant- polysaccharide are Gal, Ara, and GalA [38]. Moreover, it was reported
based polysaccharides are considered safe with no pronounced toxic that total content of uronic acids in APP was 14.4 %, indicating that
effects, hence studied for their biological activity which may be due to apple polysaccharides show potential for further development as a
the structural parameters (like molecular weight, composition of potentially significant plant extract with modified properties [50], such
monosaccharide units, glycosidic linkage) [45]. In this context, there is as bioactivities and solubility [38]. In a separate investigation, Yang
need of technological strategies to bridge the research gaps and effec et al. [32] utilized a hot-water extraction technique (80 ◦ C, 4 h) in
tively utilize apple polysaccharides in various fields, including nutra continuation with ethanol precipitation (95 % (v/v), 4 ◦ C, 24 h) to
ceuticals, food, and pharmaceuticals which are mainly present in apple isolate water-soluble apple flesh polysaccharides (AFP) and apple peel
pomace, peel or other parts of apple; generated as waste during food polysaccharides (APP) from ‘Pinklady’ apple fruit. The extraction pro
processing. However, this updated review aims to provide insights in cess resulted in APP and AFP yields of 1.4 % (w/w) and 1.7 % (w/w) of
utilizing apple waste for the extraction of polysaccharides which has the dried apple flesh and peel, respectively. To determine the mono
gained attention in the recent years and thus, address the research gaps saccharide constituents of the apple polysaccharides, the researchers
by exploring the available extraction methods and bioactivities of employed HPLC-UV, which is known for its high sensitivity and a wider
polysaccharides from apple, with the goal of valorizing apple fruit linear range, utilizing UV detection and reversed-phase C18 column. The
within a circular economy framework. Additionally, the review provides results indicated that APP is an acid heteropolysaccharide comprised of
an overview of the structural characterization, functional properties, D-galacturonic acid, D-glucuronic acid, D-ribose, D-glucose, D-xylose, D-
and potential utilization of apple polysaccharides as a functional galactose, L-rhamnose, L-arabinose, and D-mannose, in molar percent
component. By identifying research gaps and trends, this review pro ages of 13.1, 4.6, 0.8, 8.3, 4.8, 25.0, 7.3, 27.9 and 8.0 %, respectively.
vides a foundation for proposing future research in this area. On the other hand, AFP is constituted of D-galacturonic acid, D-ribose, D-
glucuronic acid, D-glucose, D-mannose, D-glucose, D-galactose, D-xylose,
2. Extraction of polysaccharide from apple and L-arabinose in mol percentages of 18.6, 1.7, 4.2, 9.0, 17.0, 9.1, and
29.7 %, respectively [32] (Table 1). Traditionally, fruit peels are
The extraction method has a strong correlation with the structural considered predominantly as source of acidic polysaccharides (i.e.,
properties of polysaccharides, consequently exerting a substantial in pectin); however, the findings of Yang et al. [32] were comparable to as
fluence on their functional and biological properties [47]. Various of APP monosaccharide profile and especially the content of galactur
extraction techniques have been investigated for the preparation of onic and glucuronic acid of AFP (22.8 %) was found to be maximum as
polysaccharides. These methods encompass both conventional ap compared to APP (17.7 %), indicating the similarity of peel and flesh
proaches like hot water, acid, or alkali extraction, and innovative apple polysaccharides. In a separate study, 8 apple pomace liquefaction
techniques such as microwave, ultrasound, enzyme, super/subcritical juices were prepared to examine the soluble cell wall component that is
water, and electric field extraction, as well as combined extraction released during enzymatic treatment (50 ◦ C, 2 h) [36]. It was revealed
methods have also been explored [48] (Fig. 2). In the case of pectin, a that the juices had high colloid values ranging from 9.7 to 19.6 g/L,
heteropolysaccharide found in apple pomace (generally, apple pomace obtained by ethanolic precipitation. Anion exchange chromatography
is discarded at end process in food industries), hot water extraction (at separated polysaccharides, revealing that 60 % of total colloids
100 ◦ C, 1 h) was used to isolate it under weakly acidic conditions (pH 7) comprised of one slightly acidic, one neutral, and one acidic polymer.
to achieve maximum yields via β-elimination reactions. From the find The results indicated that the crude polysaccharides of apple juice pre
ings, the resulting fraction was found to be rich in polysaccharide (72 dominantly consist of arabinose (14–23 mol%), galacturonic acid
%), consisting primarily of galacturonic acid (43 mol%), arabinose (23 (49–64 mol%), galactose (6–15 mol%), and traces of glucose, rhamnose
mol%), and galactose (6 mol%), which are distinguished components of and xylose [36]. In a study by Wang and Lü [51], response surface
pectic polysaccharides [34]. In addition, findings of the study revealed methodology (RSM) was utilized to improve the extraction yield of
that galacturonic acid does not take part of glycosylated polyphenols, pectic polysaccharides from apple pomace using hot-compressed water
suggesting that polysaccharides in apple pomace with hydrophobic (140 ◦ C, 5 min). The findings were consistent with previous research by
properties [34,49]. In addition, character evaluation of the poly Garna et al. [52], showing that pectic polysaccharide was made of
saccharide fraction revealed that apple pomace primarily contains Ara glucose, arabinose, xylose, rhamnose, mannose and galactose. However,
(86 %), which is responsible for the glycosidic linkages found in the yield of extracted monosaccharides varied based on the extraction
polysaccharides [34]. Moreover, polysaccharides were extracted from technique utilized, suggesting that the hot compressed water method
apple (Fuji) peel pomace by using hot water extraction method (95 % significantly impacts the composition and structural characteristics/
ethanol, 80 ◦ C, 120 min), separated by Cellulose DEAE-52 and Sephadex features of pectic polysaccharides. As an example, the lower percentage
G-150 chromatography and further characterized. The results of gel of arabinose compared to previous reports may be attributed to the
permeation chromatography revealed that these apple peel poly hydrolysis of arabinose in hairy region of pectin, which is destroyed
saccharides (APP) are heteropolysaccharides composed of fractions with during biomass hydrolysis in hot-compressed water. Large amount of
homogenous polarity. Further analysis using high-performance liquid glucose (19.85 %) suggests that some part of glucose enriched-
chromatography (HPLC) and gas chromatography–mass spectrometry amorphous cellulose or hemicellulose in apple pomace were separated
Fig. 2. Representation of different extraction methods, isolation/purification methods and structural analysis methods for apple polysaccharides.
3
Table 1
Methods of extraction of polysaccharides from apple.
N. Kumari et al.
Source/region Extraction method Extraction conditions Yield (%) Total polysaccharide/ Mw (kDa) Monosaccharide composition (%)
sugar/carbohydrate
Glu/Glc Gal Rha Ara Man Xyl Fru Fuc GlaA
content
Apple pomace (Pombal, Hot water method Aqueous acetic acid (2 100 720 g/kg _ 24 6 1 23 Tr 3 Nd Tr 43
Portugal) [34] %, v/v), 100 ◦ C, 60 min
Apple peel pomace (Fuji) Hot water method Ethanol (95 %), 80 ◦ C, 9 83 % _ 6–8 20–21 8–9 24 10–12 7–10 Nd Nd 13–18
(China) [38] 120 min
Apple peel (Pinklady) (Li- Hot water method Ethanol (80 ◦ C, 4 h) 1 – – 8 25 7 28 8 5 Nd Nd 13
Quan, China) [32]
Apple flesh (Pinklady) (Li- Hot water method Ethanol (80 ◦ C, 4 h) 1 – – 9 17 7 29 3 9 Nd Nd 18
Quan, China) [32]
Apple pomace (China) [55] Hot-water Ethanol 5 65 % – 2 20 7 32 – 6 – – 29
extraction/ethanol
precipitation
Apple pomace (China) [51] Hot compressed 140 ◦ C, 5 min – – 64 19 5 Tr 2 Tr 2 Nd Nd 48
water method/RSM
Thinned young apple Hot water extraction 95 % ethanol – 91–93 207–293 47–61 3–9 Nd Nd 26–36 3–6 Nd Nd Nd
(Qinyang Pinklady, and
Jinshiji) (China) [56]
Apple pomace (France) [40] Hot water extraction Total fraction after CC: – – 65–79 88–104 45–94 4–81 47–73 75–89 Nd Nd Nd
LA, CC:U, CC-OA or
control pre-treatment
Thinned young apple (Liquan, Hot water extraction TYAP-1, TYAP-2, TYAP- 10 96–99 479 9 23 4 78 0–7 4–7 Nd Nd 2–6
China) [28] 3/95 % ethanol
Apple pomace (Xianyang, Acid (Hydrochloric 95 % ethanol (121 ◦ C, 2 7 67 % 110 13 5 6 45 Tr 3 Nd 1 48
China) [33] acid) extraction h)
method
Apple pomace (Xianyang, Alkali (Ammonia 95 % ethanol (121 ◦ C, 2 6 75 % 89 12 10 6 7 0 7 Nd 1 51
China) [33] solution) extraction h)
4
method
Unpeeled apple (Portugal) Sequential 60 % ethanol, – – – 1–8 2–11 1 2–6 Nd 1–3 Nd Nd 71–94
[37] extraction Imidazole, Carbonate,
Citric acid
Apple pomace (China) [57] Subcritical water – 10–16 – 28 65 14–40 4–5 Tr 12 0–1 Nd Nd Nd 20–44
extraction
Apple pomace (China) [35] Solvent assisted – 11 243 Tr 7 3 – – – – Tr 47
extraction
International Journal of Biological Macromolecules 255 (2024) 128011

5
Apple pomace (F˘alticeni) Microwave assisted – 38 – 2.64 × 10 3.71 – 3.10 4.66 1.48 0.97 – – 86
[58] extraction g/mol
Apple pomace (F˘alticeni) Conventional Citric acid 38 – 2.63 × 105 6.37 – 4 7 1 1 – – 78
[58] extraction g/mol
Apple pomace (Poland) [59] Enzymatic/Acidic Endo-cellulase, endo- 10–19 331–899 4–17 g/ 4–5 g/ 0–1 g/ 3–8 g/ 0–2 g/ 1–2 g/ Nd Tr 59–74 g/
extraction xylanase/H2SO4 kDa 100 g 100 g 100 g 100 g 100 g 100 g 100 g
Apple pomace (var. Blanca de Acidic extraction 0.1 N hydrochloric acid 33–36 33 590–865 41–48 g/ 5–6 g/ Tr Tr Tr Tr Nd Nd 38–47 g/
Asturias) (Mexico) [60] mg APP/ kDa 100 g 100 g 100 g
g AP APP APP APP
Apple pulp (var. Avrolles, Solvent assisted Methanol and aqueous 71 – – 3 mg/g 28 mg/g 12 mg/ 32 9 mg/g 1 mg/g – Tr 577 mg/
Kermerrien, Jeanne Renard extraction acetone g mg/g g
and Petit Jaune) (freeze
dried) (France) [61]
Apple pomace (Shaanxi, Subcritical water 95 % ethanol, 2–14 – 7–697 16–63 8–22 0–6 1–21 0–5 3–13 Nd Tr 3–103
China) [39] extraction 100–180 ◦ C, 0–15 min mg/g mg/g mg/g mg/g mg/g mg/g mg/g
Apple Juice (Jonagold, cider – Enzymes – 28–83 % – 2–15 6–15 2–5 8–23 Nd 1–10 Nd Nd 49–77
apples) (Raw Colloids) [36]
Apple pomace (China) [53] RFAE, MWAE, CE/ Solvent pH of 2.2 (citric Pectin yield is equal to 11.24, 10.58 and 7.46 %, respectively, for RFAE, MWAE and CE
RSM acid), 88 ◦ C, and 19 min
nd = not detected, RSM = Response surface methodology, RFAE = Radio frequency assisted extraction, MWAE = Microwave assisted extraction, Tr = Traces, Glu/Glc = Glucose, Gal = Galactose, Xyl = Xylose, Rha =
Rhamnose, Man = Mannose, Fru = Fructose, Ara = Arabinose, Fuc = Fucose, GlaA = Galacturonic acid, Mw = Molecular weight, CC:LA = choline chloride: lactic acid, CC: OA = choline chloride: oxalic acid, CC: U =
choline chloride: urea, APP = Apple pomace pectin (note: all values in the table are expressed in whole number).
and linked together with pectic polysaccharides [51]. Apple poly monosaccharide composition, structural properties, rheological prop
saccharides were also extracted by acid and alkali extraction method erties and other physiological properties of the polysaccharides.
using hydrochloric acid and ammonia solution, respectively (121 ◦ C, 2
h), involving freeze-thawing (10 cycles) for protein removal. It was 3. Structural characterization
determined that extraction methods may be responsible for the poly
saccharides structure and their biological activities. From the investi Various sources such as apple peel, apple pomace or whole fruit have
gation, it was estimated that apple pomace polysaccharides extracted been used to extract polysaccharides, which have been subjected to
under acidic (ACAPP), and alkali (ALAPP) conditions have different structural analysis using various methods. Fourier-transform infrared
extraction yield, monosaccharide content (%), molecular weight, total (FTIR) spectroscopy, HPAEC-PAD, 1H and 13C NMR spectroscopy are
sugar and total polysaccharide content. It was shown that the yield of among the common techniques utilized for the structural characteriza
ALAPP (110 kDa) and ACAPP (89.6 kDa) were 6.84 % and 7.17 % and tion of apple polysaccharides (Fig. 2). In a study, polysaccharides from
total sugar contents of ALAPP and ACAPP were equal to 75.61 % and thinned-young apple varieties (Jinshiji, Pinklady and Qinyang) were
67.14 % (Table 1). From high-performance anion exchange chromato investigated by using 1H and 13C Nuclear magnetic resonance (NMR)
graph (HPAEC) analysis, glucuronic acid, fucose, rhamnose, galactose, spectroscopy. From the elucidation of the 1H and 13C NMR spectral
xylose, arabinose, glucose, fructose, galacturonic acid, mannose, man (5.50–4.90 ppm) analysis, it was suggested that the apple poly
nuronic acid and ribose were recognized as main monosaccharides of saccharides have α- and β- glycosidic linkages corresponding to the
apple pomace polysaccharide. GalA contributed the maximum mono anomeric carbon. The findings of the investigation were in line with the
saccharide compositions of the polysaccharide fraction, 51.91 % in Fourier-transform infrared (FTIR) spectroscopy outcomes. Additionally,
ALAPP and 48.99 % in ACAPP [33]. A different study was conducted to findings revealed that β-D-glucose, β-D-xylose, β-D-mannose, α-D-
evaluate the effectiveness of 3 natural deep eutectic solvents (NADES), mannose, α-D-glucose, and α-D-galactose were present, consisting of 1 →
namely choline chloride (CC): CC: oxalic acid (OA), lactic acid (LA), and 6 linked monosaccharide backbone and (1 → 3) linked glucose residues,
CC: urea (U), in a 1:2 ratio as a pre-treatment for apple pomace before as well as the presence of -COOH group. The X-ray powder diffraction
hot water extraction of pectin. The outcome showed that pretreatment (XRD) analysis validated the existence of amorphous and semi-
of CC:LA and CC:U resulted in similar total recovery yields as the control crystalline structures in young apple polysaccharides [56]. Similarly,
group. However, the CC:OA treatment led to the degradation and loss of glycosidic linkage analysis by 13C NMR spectrum revealed the occur
polysaccharides. Furthermore, CC:LA and CC:U pre-treatments at 60 ◦ C rence of (1 → 2,3)-Araf (1.3 %), (1 → 2,3,5)-Araf (3.1 %), (1 → 2,5)-Araf
and above resulted in changes in composition of cell wall mono (3.8 %), (1 → 3,5)-Araf (6.8 %), t-Araf (13.4 %) and (1 → 5)-Araf (18.8
saccharides, but CC:LA pre-treatment at 40 ◦ C for 1 h retained the %), characteristic of arabinans [62] while the occurrence of t-Xylp (0.4
original composition. The CC:LA pre-treatment allowed for the extrac %), (1 → 2)-Galp (1.6 %), t-Fucp (1.6 %), t-Galp (2.2 %), (1 → 2)-Xylp
tion of a considerable amount of high molecular weight pectin (3.8 %), (1 → 4,6)-Glcp (4.1 %) together with (1 → 4)-Glcp (14.6 mol%)
(33.1–56.1 %), equivalent to the traditional technique. On the other suggests the occurrence of xyloglucans [63]. The remaining t-Xylp (6.9
hand, CC:U pre-treatment led to saponification and affected pectin %) could be explained by the occurrence of xylogalacturonan in in apple
composition by introducing choline. The pre-treatment of apple pomace pomace [64]. In a research study, 8 different apple pomace liquefaction
with CC:LA, subsequently followed by water extraction, provides a juices were produced to study the cell wall material that was released
“green” alternative to mineral acid pectin extraction, while CC:U func after treatment with pectolytic and cellulolytic enzymes. The findings
tionalizes the pectin in pomace before hot water extraction [40]. Zheng indicated that the apple cell wall is primarily comprised of galacturonic
et al. [53] conducted a comparative study on the extraction of pectin acid, arabinose and galactose, with traces of rhamnose, glucose, and
from apple pomace using radio frequency assisted extraction (RFAE), xylose (Tables 1, 2). By separation through anion-exchange chroma
conventional extraction (CE), and microwave-assisted extraction tography, it was found that polysaccharide consists of one slightly
(MWAE) techniques. The results revealed that under optimal extraction acidic, one neutral and one acidic polymer. The structural character
conditions (pH 2.2, 80 ◦ C, 19 min), RFAE and MWAE had similar pectin ization of polysaccharides is performed through permethylation anal
yields (11.24 % and 10.58 %, respectively), both outperforming CE ysis. It was indicated that neutral polysaccharides were predominantly
(7.46 %), which required longer extraction time to achieve the highest composed of α-1,5-L-arabinans [36]. In a study, structure of galactans
yield. While the various extraction methods did not affect the structural and arabinans in 14 different apple cultivars using various methods,
properties of the extracted pectin, RFAE demonstrated its efficiency in including enzymatic cleavage and methylation analysis was analyzed.
maintaining the physicochemical properties of pectin more effectively The main objective of study was to determine specific information on
compared to MWAE and CE when operated under the same conditions. the neutral side chains of pectin. The findings revealed that all cultivars
RFAE pectin displayed the highest degree of esterification and gal had linear (1 → 4)-β linked galactans and distinct arabinans with heavily
acturonic acid content among the three extracted samples. Analytical branched branches. By using enzymatic profiling techniques, rare
techniques, such as FTIR, XRD, and 1H NMR, confirmed the presence of structural components like α-arabinop and β-arabinof residues were
characteristic functional groups naturally found in pectin for all identified (Table 2). These results indicate that neutral pectin side chains
extraction methods, with no significant differences in the main chemical in apples are highly complex [65]. Hu et al. [35] investigated the apple
structures. However, when examining the surface morphology, RFAE pectin (Red Fuji) extracted by using solvent assisted extraction/HPLC
and MWAE samples exhibited rougher textures with more cracks, method, for chain conformational studies. For the structural character
whereas CE samples had a relatively intact and smoother surface. In ization or optimization of apple pectin, FT-IR and Hydrogen nuclear
conclusion, it was suggested that RFAE holds promise as a superior magnetic resonance (1H NMR) spectroscopy was used. The results
extraction technology for obtaining pectin with higher yield and quality showed that apple pectin was mainly composed of arabinose (Ara) and
compared to MWAE and CE [53]. galactouronic acid (GalA) with molar ratios 38.2 % and 39.9 %,
Based on the findings of all studies, it was concluded that the hot respectively; in which Ara existed as (1 → 3)-, (1 → 5)-, and (1 → 2,5)-
water method is commonly employed for extracting polysaccharides due Araf and T-Araf with L-configuration. Among these, T-L-Araf (11.4 %)
to its minimal impact on the surface structure of these compounds. On and (1 → 5)-L-Araf (16.9 %) (Table 2) occupied substantial amounts of
the other hand, alternative methods, like microwave-assisted extraction, all Ara sugar residues, indicating that the RG-I region of pectin in apple
were found to affect the surface structure of polysaccharides [54]. In was extensively branched with Ara residues at terminals, in a manner
conclusion, it was proved that type of extraction procedure, extraction that is equivalent to the pectin that was recovered from peach gum in an
conditions such as reaction temperature, reaction time, pH, type of earlier study by Wei et al. [66]. In the case of GalA, the linkage between
solvent used etc., greatly affect the yield, carbohydrate/sugar content, residues occurred through an α-1,4 linked with a pyran ring in D-
5
Table 2
Structural characteristics of polysaccharides obtained from apple.
Polysaccharide Extraction, separation, Monosaccharide composition (%) Structure Structural features Reference
fraction purification method characterization
methods
Thinned-young Hot water extraction Xyl (3–6), Man (26–36), Gal (3–9), Glu FTIR and 1H and 13C (1 → 3), (1 → 6): α- [56]
apple (47–61) NMR β-D-glycosidic linkage
Apple juice Anion-Exchange and Rha (2–5), Ara (8–23), Gal (6–15), Glu – 1 → 2-linked Rhap, 1 → 2,4-branched Rhap, [36]
(Pomace) Analytical Size Exclusion (2–15), Xyl (1− 10), GalA (49–77) 1 → 5-linked Araf, 1 → 3,5-branched Araf, 1
Chromatography → 2-linked Xylp, 1 → 2,5-branched Araf, 1
→ 3-linked Galp, 1 → 4,6-branched Galp, 1
→ 4-linked Galp, 1 → 6-linked Galp, 1 → 3,4-
branched Galp, 1 → 2-linked Araf, 1 → 2,4-
branched Galp 1 → 3,6-branched Galp, 1 →
3-linked Araf
Extracellular Sequential extraction/GC/ Rha (3–4), Fuc (2–8), Ara (8–62), Xyl – Xylp-T, 4 → 6 Araf-T, Galp-T, 4-Glcp, 2-Xylp, [69]
polysaccharide Anion-exchange (6–35), Man (2–4), Gal (8–51) Glu and 3 → 6-Galp
chromatography (2–40)
Apple pulp Acidic (H2SO4) hydrolysis/ Fuc (1–2), Rha (1–2), Ara (10–19), Gal HPAEC-PAD/NMR Araf, 1 → 4-Xylp, 1 → 2-Xylp, 1 → 5-Araf, 1 [65]
High performance anion (7.0–16), Glu (34–43), Xyl (9–13), Man spectroscopy → 2,5-Araf, 1 → 2,4-Rhap, 1 → 3,5-Araf, 1
exchange chromatography (3–7), Galacturonic acid (12–15), → 2,3,5-Araf, 1 → 2-Rhap, Galp-T, 1 → 4-
Glucuronic acid (tr) Galp, Manp-T, 1 → 6-Galp, 1 → 4-Manp, 1 →
3,6-Galp, Glcp-T, 1 → 4-Glcp, 1 → 4,6-Glcp
1
Apple pomace Solvent assisted extraction/ Rha (3), Glc (Tr), Xyl + Ara (40), Gal H NMR/Ft-IR 1 → 3-L-Araf, 1 → 5-L-Araf, 1 → 2,3-L-Arap, [35]
(Red Fuji) HPLC (7), GalA (47), Fuc (Tr), HG (43), (Gal + spectroscopy 1 → 2,5-L-Araf, 1 → 2,4-L-Rhap, α-D-GalpA-
Ara + Xyl)/Rha (12), Rha/GalA (Tr), (1 → 4), 1 → 4,6-D-Galp, 1 → 2,4-D-GalAp, 1
GalA/(Fuc + Rha + Ara + Gal + Xyl + → 3,4-D-Galp, 1 → 4,6-D-Glcp, 1 → 3-D-
Glc) (Tr) Galp, 1 → 3,6-D-Glcp, 1 → 6-D-Galp
Apple pomace – Fuc (0–1), Rha (1–4), Ara (9–42), Gal GC–MS/GC-FID 1 → 2-Rhap, 1 → 2,4-Rhap, Araf-T, t-Arap, 1 [70]
(8–19), Glc (3–53), Xyl (1− 13), Man → 5-Araf, 1 → 4-Arap, 1 → 3-Araf, 1 → 2,5-
(1–4), GalA (6–44) Araf, 1 → 3,5-Araf, 1,2,3,5-Araf, Galp-T, 1
→ 4-Galp, 1 → 6-Galp, Glcp-T, 1 → 4-Glcp, 1
→ 4,6-Glcp, 1 → 4-Manp, 1 → 4,6-Manp,
Xylp-T, 1 → 2-Xylp, 1 → 4-Xylp
Apple flour Acidic extraction (Citric Pectin (14–16) XRD, 13C NMR, FTIR (1 → 5) glycosidic linkage [71]
acid/Hydrochloric acid)
Apple pomace Hot extraction method/C18 Ara (33), GalA (31), Glc (15) GC-FID/NMR (1 → 2,3)-Araf, (1 → 2,3,5)-Araf, (1 → 2,5)- [34]
cartridge solid phase Araf, (1 → 3,5)-Araf, t-Araf, (1 → 5)-Araf, t-
extraction Xylp, (1 → 2)-Galp, t-Fucp, t-Galp (2.2 %), (1
→ 2)-Xylp, (1 → 4,6)-Glcp, (1 → 4)-Glcp
HG = homogalacturonan, Fuc = Fucose, Fru = Fructose, Glc/Glu = Glucose, HPLC = High-performance liquid chromatography, GalA = Galacturonic acid, Gal =
Galactose, Rha = Rhamnose, Man = Mannose, Xyl = Xylose, GC = Gas chromatography, NMR = NMR = Nuclear magnetic resonance, FTIR = Fourier transform
infrared, GC–MS = Gas chromatography-Mass spectrometry, HPAEC-PAD=High-performance anion-exchange chromatography/pulsed amperometric detection, XRD
= X-ray Powder Diffraction, GC-FID = Gas Chromatography With Flame Ionization Detection (note: all values in the table are expressed in whole number).
configuration. The major type of linkage accounted for 35.6 % of the products. Indeed, processing effects on nutritional requirements (dietary
molar ratio. The bonding pattern of GalAp was also identified as (1 → fiber content) is often investigated by methods like drying, heating and/
2,4). Additionally, the analysis detected the presence of rhamnose (Rha) or enzymatic treatment. In food industries, the processing of fruits, such
in the form of (1 → 2, 4)-L-Rhap, which indicates that Rha was branched as canning, peeling, and blanching, may have a negative impact on their
at C4, comparable to the pectin extracted from grapefruit peel and fiber content. Moreover, the quantity of fiber and cell wall poly
Lonicera japonica Thunb reported by Wang et al. [67] and Liu et al. [68], saccharide present in the final product may also be influenced by the
respectively [35]. Based on the findings of the study, it was observed fruit’s initial maturity stage. It is worth noting that the characteristics
that in some cases, there is limited or no information available on the and composition of the raw materials have an impact on the processing
absolute configuration (L or D) and glycosidic linkage stereochemistry methods employed by fruit processing plants [72]. Research was con
(alpha or beta) of polysaccharides. However, it is important to investi ducted to examine how changes in the amount of dietary fiber in apples
gate the structural features in-depth, as they play a significant role in occur during the production of apple sauce at 5 different processing
determining the bioactivity and industrial applications of poly stages (fruit sorting, refining, cooking, sugaring and pasteurization). The
saccharides in various fields. results showed that the total fiber content decreased from 2.4 to 1.7 g/
100 g of F.W., while the soluble fiber fraction increased. The processing
4. Effect of extraction methods, processing and storage on cell led to a reduction in fiber content and a shift in the ratio of soluble to
wall polysaccharides insoluble compounds [43]. Also, during the cooking and sugaring pro
cess, insoluble polysaccharides were reduced, and prolonged heating
Apple is a commonly eaten fruit and has been extensively studied for during sugaring surprisingly resulted in the solubilization and degra
its nutritional and phytochemical content. It is an interesting source of dation of pectin. The changes in pectin were also observed to be influ
insoluble and soluble dietary fibres i.e., xyloglucans, cellulose and pectic enced by the post-maturity stage of the apples. As a result of processing
polysaccharides, respectively, both related with the cell wall of the fruit apples into applesauce, only 30 % of the dietary fiber was lost (based on
[4–6,70]. Processed apple-based food products such as sauce, are thus fresh weight), but there was a significant shift in the ratio of soluble to
advertised as healthy compared to chocolate bars or candies, often eaten insoluble fiber (from 28 % to 39 %) [43]. Similarly, in another study
in-between meals by most of the today’s generation [43]. However, effects of storage conditions of apple on enzymatic hydrolysis of cell wall
precise data is unavailable regarding the persistence of original apple polysaccharides were investigated [73]. It was found that water-
nutritional content, more precisely in whole processed apple food insoluble content decreased while water-soluble fraction content
6
remains constant in apple stored at 4 ◦ C for 30 weeks. The percentage of rose, the apparent viscosity, Mw, and GalA content of the apple pomace
polysaccharides decreased (10 %); galactose and arabinose amount polysaccharides demonstrated a comparable decreasing trend. Howev
progressively declined. There was a positive correlation between yields er, the variation of GalA content, apparent viscosity and Mw of the apple
and the enzyme hydrolysis of water-insoluble fraction, suggesting that pomace polysaccharides had a similar decreasing trend as temperature
the longer the storage duration of apple tissues, the more beneficial the increases [39].
enzymatic treatment would be. During storage, the availability of pectin Likewise, another study investigated the rheological and thermal
to polygalacturonases in apple tissues increased, leading to a higher properties of apple pomace pectin (APP) and found that the chains of
yield of pectin solubilization when hydrolyzing enzymes were used. APP disentangled quickly during high-frequency oscillation, leading to a
Additionally, the longer the storage duration, the greater the increase in pseudoplastic behavior [60]. These findings could be useful in control
pectin solubilization yield [73]. Likewise, in an investigation it was ling viscosity of polysaccharides containing salt during food processing.
found that using aqueous ozone (AO) treatment at a concentration of However, the gelling properties of pectin are influenced by several
1.4 mg L− 1 for 5 min in fresh-cut apples during cold storage can delay factors, such as molecular weight, pH, concentration of solutes, and
degradation of cell wall polysaccharides and decrease softening of the degree of methylation (DM), which includes the degree of esterification
apples. Additionally, the treatment increased the amount of water- (DE). Pectin can be classified into low-methoxyl pectin (20–40 %) that
soluble pectin fraction compared to untreated apples. These results forms gels in presence of Ca2+, and high-methoxyl pectin (60–75 %) that
indicate that AO treatment can have a significant positive impact on the forms gels in an acidic condition in presence of sugar. These classifica
structure and composition of cell wall polysaccharides in fruits and tions are important for industrial applications. In general, DM of pectin
vegetables [74]. In another study, the effect of different drying processes depends on extraction conditions (time, pH and temperature), but
on cell wall polysaccharides in apples was evaluated. Apple chips, a particularly decreases with longer duration and/or lower pH. O’Shea
popular food product in China, are produced using medium-wave et al. [75] obtained pectin through an extraction process using 1 M HCl
infrared drying and short-wave infrared drying (IR), hot air drying at room temperature for 3 and 7 h, resulting in DEs of 56.38 % and
(AD), vacuum freeze drying (FD), and instant controlled pressure drop 39.64 %, respectively. They found that extracting pectin at 90 ◦ C led to a
drying (DIC). Thus, drying causes various modifications in the cell wall lower DM than at 80 ◦ C, and pectin extracted at a pH of 1.5 had a lower
polysaccharides involving rehydration behavior, microstructure, vol DM as comparable with extracted at pH 2 [52]. Similarly, DEs from
ume expansion and so on, responsible for the texture properties of apple apple pomace extracted via HCl at 97 ◦ C and pH 2.5 were 45.98 and
chips. However, during large scale production, apple chips texture is 22.15 %, while on the other hand; by using citric acid at same conditions
unstable because of a lack of knowledge about the principles of texture were 52.51 and 50.14 %, respectively [71]. Compared to acidic
evolution, which is thought to be connected to pectic polysaccharides extraction, there are many new ways to improve the DM. Wikiera et al.
(except for the final moisture content and microstructure). Therefore, [76] showed that acid assisted extracted apple pectin had a DM of 56.1
greater understanding of the texture of apple chips and the modulation %, while commercial pectin had a DM of 66.9 %. When enzyme prep
of pectic polysaccharides should be beneficial to ensure outstanding aration Celluclast 1.5 L was used for extraction, the DM value of pectin
texture of the food products [41]. Upon thorough examination of mul was improved, ranging from 57.4 to 58.7 %. Wikiera et al. [59] deter
tiple studies, it becomes evident that the methods and conditions mined that the DM of apple pomace was 73.4 % when endo-xylanase was
employed for polysaccharide extraction play a crucial role in shaping used, 66.3 % when endo-cellulase was used, 67.5 % when both enzymes
various characteristics of these compounds, including composition, were used together and 56.1 % when acid (H2SO4) was used. Wang et al.
molecular weight, yield, as well as physiochemical, structural, and [57] used subcritical extraction procedure, with DE of the pectin ranging
rheological properties. from 83.41 to 89.69 %. Hou et al. [56] investigated the emulsion
properties of polysaccharides extracted from three varieties (Qinyang,
5. Physicochemical and functional properties Jinshiji and Pinklady) of thinned young apple and it was found that all
three polysaccharides mainly involve glucose, galactose, mannose,
The apple-derived polysaccharides exhibit various physicochemical xylose. Compared with the other two varieties, the polysaccharides
and functional characteristics. As previously stated, the properties of isolated from “Jinshiji” had better emulsifying properties whereas
these polysaccharides, including physical, chemical, and structural Qinyang has better thermal stability than others. Furthermore, cation
features, are influenced by factors such as the composition and organi concentration (Ca2+ and Na+) and pH had significant effect on rheo
zation of monosaccharides, molecular weight, chain conformation type logical properties of the emulsion of extracted polysaccharides. At pH
and location, particle size, rheological properties, and solubility [45]. In 2.0–4.0, the emulsion was stable, had uniform and small droplets than at
a study, steady-shear viscosity property in relation with chain confor high pH. The emulsion exhibited lower emulsification stability at high
mation of apple pectin (AP) in water and NaNO3 was investigated. Ac concentration of cationic ions, indicating application of apple poly
cording to the results, the consistency index (k) of AP decreased as the saccharides as emulsifying agent [56]. In another study, modified apple
concentration of NaNO3 increased, but the impact of the salt on the k polysaccharides (MAP) capped gold nanoparticles (AuNPs) were eval
value of AP (33.33–7.78) was noteworthy. Furthermore, the flow uated for its viscosity. It was found that viscosity of liquid-MAP was
behavior index (n) of AP increased from 0.24 to 0.49 with the rise in salt equal to 1.14 cP, for MAP-capped AuNPs was 1.28 cP, and for AuNPs-
concentration. Similarly, as the concentration of AP increased in pure INS was 1.46 cP. An improvement in viscosity was observed due to
water, its steady-shear viscosity also increased. AP exhibited Newtonian capping of AuNPs with MAP followed by conjugation of MAP-capped
flow characteristics at shear rates below 1 s− 1 and concentrations of AuNPs with insulin (INS) (AuNPs-INS) [77]. Apart from the rheolog
≤15 mg/mL. Overall, the research findings indicate that AP has a higher ical properties, it was proved that apple pectin polysaccharides have
molecular weight, longer side chains attached to the RG region, and less hydrophobic characters; mainly due to covalently linked polyphenols as
electrostatic repulsion, resulting in more pronounced shear thinning, a result of polyphenol oxidation reaction [34]. Moreover, according to
greater chain entanglement and a strong interaction between pectin Xiao et al. [41], drying temperature and drying methods exert signifi
chains [35]. The results of the study were similar with the investigation cant effect on the physicochemical properties including water content,
of Zhang et al. [39]; indicating that apparent viscosity of apple pomace crispness, hardness, rehydration ratio (RR), volume ratio (VR) when
polysaccharide solutions decreased significantly with increase in shear tested against apple chips; caused due to modification of cell wall
rate, thus exhibiting non-Newtonian or pseudoplastic fluid behavior. polysaccharides during drying. However, there are not many studies
The study also revealed that the rheological characteristics were closely related with the functional properties of apple polysaccharides. Besides
associated with the quantity of GalA and Mw and other physiochemical that, Rana et al. [78] and Negi and Vaidya [79] have investigated the
attributes of the polysaccharides [60]. However, as the temperature functional properties of dietary fiber (apple pomace powder) owing to
7
be rich in polysaccharides [14]. The studies revealed that apple pomace control group. These findings suggest that polysaccharides may have a
powder possesses various functional properties such as swelling capac protective effect against CCl4-induced damage. The study also examined
ity, water, and oil holding capacity, density, glucose dialysis retardation the impact of polysaccharides on enzymatic activities of serum alanine
index, foam capacity, and emulsion activity and stability. It was aminotransferase (ALT), aspartate aminotransferase (AST) and lactate
observed that the absorption of glucose increases with its concentration, dehydrogenase (LDH), which are sensitive indicators of hepatic injury.
which leads to a reduction in the rate of glucose adsorption and ulti The results showed that administration of APP and AFP reversed the
mately decreases postprandial serum glucose concentration [79]. Thus, effects of CCl4-induced liver damage and prevented the elevation of
the dietary fiber fractions present in apples have the potential to be serum AST (0.28–0.16 U/mgprot, p < 0.01), ALT (0.33–0.24 U/mgprot,
utilized as a functional ingredient in foods with high dietary fiber con p < 0.01), and LDH (4910–4230 U/mgprot, p < 0.01) in a concentration-
tent. However, the suitability of this ingredient will alter on type of food dependent manner. Additionally, administration of APP and AFP
product being developed and the overall cost-effectiveness of process. resulted in a reduction in the concentration of malondialdehyde (MDA),
an index of chain reaction of lipid peroxidation (2.32–1.81 nmol/
6. Biological activities mgprot, p < 0.01) at doses of 250 and 500 mg/kg bw. Overall, the results
of the study suggest that apple polysaccharides have a beneficial effect
Polysaccharides has attracted lot of attention in recent times due to against oxidative damage caused by CCl4 [32]. Moreover, studies have
their numerous beneficial health effects. These have been used in nu shown that apple polysaccharides can have a positive impact on the
traceutical, pharmaceutical, food and cosmetic industry for the devel metabolic functions of the liver. Chen et al. [55] shows that adminis
opment of noble products [45,80]. Different biological properties of tration of TYAP at dosage of 400 mg/kg/day and 800 mg/kg/day
apple polysaccharides have been discussed in the Sections 6.1–6.6, (orally) in HFD-induced obese mice model significantly rescued high-fat
Table 3 and Fig. 3. diet (HFD) induced hepatic metabolic impairment, ameliorated hepatic
oxidative stress and reduced body weight gain induced by HFD. In brief,
6.1. Antioxidant activity TYAP treatment inhibits the production or cause reduction in serum
levels including total cholesterol (TC) (0.70–0.53 mmol L− 1), tri
The antioxidant activities are directly related to the reducing power glycerides (TG) (0.22–0.15 mmol L− 1), low-density lipoprotein choles
of certain plant extracts [81]. A research was conducted to evaluate the terol (LDL-C) (0.08–0.03 mmol L− 1), fasting insulin level (48.9–35.50
antioxidant properties of water-soluble polysaccharides extracted from pmol/L) and decrease of HI (p < 0.05) in tested animal model. Addi
both apple peel (APP) and apple flesh (AFP) using various assays such as tionally, in palmitate-loaded HepG2 cell model, TYAP reduce mito
DPPH, HO•, and O2− as well as ferric-reducing antioxidant power chondrial reactive oxygen species (ROS), enhance palmitate-impaired
(FRAP) [32]. Findings of study shows that in-vitro scavenging effects of mitochondrial respiratory performance, and protect the cells from
APP and AFP with IC50 value equal to 18.3–85.5 % and 18.6–89.4 % for palmitate-induced insulin resistance and viability loss. According to
DPPH• assay (dose: 50–800 μg/mL); 26.8–53.3 % and 27.8–56.5 % for these results, TYAP may be able to successfully mitigate the hepatic
O2•− scavenging assay (dose: 50–1000 μg/mL); 6.8–47.8 % and metabolic problem associated with obesity by stimulating its mito
22.9–45.8 % for HO• (dose: 50–800 μg/mL) scavenging effect, respec chondrial respiratory function [55].
tively. The study found that both polysaccharide fractions derived from
apple peel and flesh exhibited a considerable ability to decrease 6.3. Anti-ulcer activity
Fe3+/K3Fe(CN)6 complex to the ferrous form (Fe2+) in a dose-dependent
manner, as determined by FRAP assay. However, there were no notable Currently, the anti-inflammatory characteristics of modified apple
differences in the scavenging capacity of APP and AFP (p > 0.05). The polysaccharides (MAP) and silver nanoparticles (AgNPs) are drawing
study’s findings suggest that the polysaccharides extracted from apple attention in the treatment of ulcerative colitis (UC) [84–88]. In UC, the
peel and flesh possess strong antioxidant properties and scavenging body releases prostaglandins and various chemotactic substances such
abilities against free radicals, which could potentially prevent or alle as interleukin-1 (IL-1), complement factors, tumour growth factor
viate oxidative damage [32]. Similarly, in another study different-water (TGF), and tumour necrosis factor (TNF). AgNPs are determined to
soluble polysaccharide (thinned-young apple polysaccharide i.e., TYAP- inhibit the expressions of these inflammatory markers [88]. Recently, a
1, TYAP-2, TYAP-3) were isolated from thinned-young apple (TYA) were study was conducted to investigate the effectiveness of using MAP
investigated for their antioxidant potential. From the findings, it was capped AgNPs loaded with mesalamine (MES) (AgNPs-MES) in treating
determined that the purified polysaccharides from TYA possess signifi UC in an acetic acid induced rat model. The researchers evaluated
cant antioxidant potential towards DPPH radical (IC50 = 2.64–4.11 mg/ several indicators such as superoxide dismutase (SOD), glutathione
mL), superoxide radical (IC50 = 2.19–4.72 mg/mL), hydroxyl radical (GSH), myeloperoxidase (MPO), tumour necrosis factor (TNF)-α and
(scavenging activity = 34.63–58.06 %) assay [28] (Table 3). Based on thiobarbituric acid reactive substances (TBARSs) to serve as markers for
the data, it is evident that apples may serve as a suitable foundation for intestinal inflammation. From the results, it was found that the animal
future research regarding the industrial application of biochemicals. model demonstrated a significant recovery rate on the 7th and 14th days
Additionally, these apples have the potential to serve as a functional when administered a higher dose of MAP capped AgNPs-MES (1–4 mg/
ingredient within the pharmaceutical and food industries. kg). Co-administration of MAP with AgNPs and MES was found to
possess better anti-ulcer properties compared to the individual treat
6.2. Hepatoprotective activity ments of MAP alone, MES alone, and AgNPs, which successfully reduced
inflammation in the intestine. The dissolution study revealed about 54 %
Liver disease is believed to be caused, at least in part, by oxidative drug release after 5 h indicating release of drug at the target colonic site.
stress that can damage biomolecules. The use of dietary antioxidants is The in-vivo biochemical and histopathological analysis supported the
theorized to decrease the likelihood of oxidative damage and help pre drug targeting effect because the combination of MAP, MES and AgNPs
vent liver disease [82,83]. Besides flavonoids, apples are recognized as provided better efficacy to treat UC as compared to their individual
good source of antioxidant polysaccharides. In a study, polysaccharide treatment. After the complete treatment regime, all groups exhibited a
fraction of apple flesh (AFP) and apple peel (APP) were tested in-vivo considerable reduction in the DAI score (p < 0.05), colonic weight,
against CCl4 induced liver damage in mice for their hepatoprotective confirming the efficacy of the combined dose of MAP, AgNPs and MES in
effect. The oral administration of APP and AFP at doses of 250 and 500 treating UC over the individual treatments [89]. In a different study,
mg/kg b.w. was found to significantly reduce liver weight and liver researchers developed MES and MAP tablets and combined them with
index (HI) (p < 0.05), especially at higher doses, compared to the probiotics to address the loss of bacteria that can occur during UC
8
Table 3
Bioactivities of apple polysaccharides.
Type of activity Source/region Organism or cell line/assay Dosage/Fraction used Key findings Reference
Antioxidant Apple peel (Shaanxi, DPPH •

50–800 μg/mL Significant scavenging activity was observed with [32]
activity North-west China) IC50 ranging from 18.3 to 88.5 %
O2•−
50–1000 μg/mL Inhibition of free radical with IC50 equal to
26.8–53.3 %
HO• 50–800 μg/mL Significant free radical scavenging capacity was
observed (6.8–47.8 %)
FRAP 50–1000 μg/mL Reduction of Fe3+/K3Fe (CN)6 complex to Fe2+
Antioxidant Apple flesh DPPH• 50–800 μg/mL Inhibition of free radical with IC50 of 18.6–89.4 % [32]
activity (Shaanxi, North- O2•− 50–1000 μg/mL Significant scavenging activity was observed with
west China) IC50 of 27.8–56.5 %
HO• 50–800 μg/mL Significant scavenging activity against HO• was
observed (22.9–45.8 %)
FRAP 50–1000 μg/mL Reduction of Fe3+/K3Fe (CN)6 complex to ferrous
form
Antioxidant Thinned young DPPH•, O2•− , HO• assay 0.25–5.0 mg/mL Significant antioxidant potential was observed. For [28]
activity apple (China) DPPH assay: IC50 = 2.64–4.11 mg/mL; HO• assay:
scavenging effect was 34.63–58.06 %; O2•− assay:
IC50 = 2.19–4.72 mg/mL
Antioxidant Apple peel DPPH•, O2•− and HO• 0.25–2.0 mg/mL Significant radical scavenging activity in a dose [38]
activity dependent manner
Antioxidant Apple pomace Liver of male Kunming mice, 50–200 mg/kg/day Increase in the expression level of CAT [33]
activity (China) weighing 20 g (411.11–543.20 U mg prot− 1) and SOD
(729.24–961.89 U mg prot− 1) and decrease in the
level of MDA (2.27–3.16 mmol mg prot− 1); more
efficient in removing excess free radicals during
exercise (p < 0.05).
Hepatoprotective Apple peel and CCl4 induced liver damage 250 and 500 mg/kg bw Apple polysaccharides cause elevation in activity [32]
activity Apple flesh mice MDA, SOD, GSH-Px, of SOD, GSH-Px, and decrease in levels of ALT, AST
(Shaanxi, North- ALT, AST and LDH and LDH
west China)
Hepatoprotective Thinned young HFD-obese mice, palmitate- Animal model: 400 mg/kg/ Reduction in impairment of hepatic metabolic, [55]
activity apple (Shaanxi loaded HepG2 cell model day and 800 mg/kg/day hepatic oxidative stress and body weight gain,
province, China) TYAP induced by HFD.
HepG2 cell line = 0-200 μg/ Protection against viability loss, palmitate-induced
mL TYAP insulin resistance, inhibited mitochondrial ROS
and improvement in mitochondrial respiratory
function impaired by palmitate in HepG2 cells
Hepatoprotective Apple peel (Shaanxi, CCl4-induced hepatotoxicity Animal model: Mice (450 mg/ Significant reduction in elevated level of serum [38]
activity China) kg bw) alanine aminotransferase, asparate
aminotransferase and lactic dehydrogenase
activities, and hepatic malondialdehyde.
Inhibition in the activity of superoxide dismutase
and glutathione peroxidase
Anti-ulcer activity Apple (Punjab, Acetic acid caused ulcerative MAP (AgNPs+MES): 1–4 mg/ Absence of disruption in epithelial cells, acute [89]
India) colitis in rat model kg inflammation, lesions indicating pathological
significance
Gastrointestinal Apple pomace B. longum and L. rhamnosus – Apple polysaccharides promote the growth of [93]
activity (China) L. rhamnosus and B. longum, indirectly; beneficial
for gastrointestinal health
Gastrointestinal Apple pomace HFD fed rats Apple polysaccharide dose = Regulation of gut microbiota composition [92]
activity (Shanxi, China) 5 % and 10 %, [wt/vol], 1 improved microbial metabolites SCFAs and
mL/100 g weight increased gut barrier dysfunction.
Reduction in chronic inflammation and gut
permeability in HFD-fed rats by autophagy
induction of goblet cells
Gastrointestinal Apple pomace (var. Human microbiota from – Stimulation of growth of Akkermansia, Prevotella, [19]
activity Perico, Regona and healthy subjects and Lachnospiraceae UCG-010, Sucinivibrio, Blautia,
Solarin) (Asturias, inflammatory bowel disease Turicibacter, Lachnospiraceae CAG-56, Eubacterium
Spain) (IBD) patients eligens, Dialister and Intestinimonas
Anti-ulcer activity Apple (Jammu and Acetic acid induced Gaur gum (5 % w/v)- Reduction in DAI score, macroscopic score as well [87]
Kashmir, India) ulcerative colitis Sprague Eudragit S 100 (8 % w/v) as colonic weight gain, peroxidation parameters
Dawley rat (250–300 g) coated MES and MAP mini (SOD, GSH, TBARs and TNF-α indicative of
tablets inflammation of the intestine
Anticancer activity Apple pomace Azoxymethane and dextran 5 % AP (w/v) Suppression in tumorigenesis, significant increase [94]
(Shaanxi, China) sodium sulfate induced in mRNA expression of macrophage M1 markers
carcinogenesis mice, Raw (iNOS, IL-23, TNF-α), upregulation in protein
264.7 expression of Myd88, TLR-4, p65 and p-p65
Anticancer activity Apple pomace Azoxymethane and dextran 10 % AP (w/v) Restoration of AOM/DSS-caused intestinal Li et al.,
(Shaanxi, China) sodium sulfate induced microbiota disorder; inhibition in regulation of 2019
carcinogenesis mice Wnt pathway in colon tissues; decrease in measure
of Fusobacterium
Anticancer activity Apples (Red Fuji) Dextran sodium sulfate- MAP (2.5 %) Prevention against colitis through down-regulation [85]
(China) induced colitis mouse, and of secretion of IL-22, and upregulation in
MCA-38 and DC2.4 cell line expression of IL-22BP
(continued on next page)
9
Table 3 (continued )
Type of activity Source/region Organism or cell line/assay Dosage/Fraction used Key findings Reference
Anticancer activity Apple (Fuji) (China) LPS stimulated CRC cells MAP = 0.01–1 mg/mL Suppression in LPS-induced migration and [106]
(HT-29 and SW620) invasiveness of CRC cells by affecting LPS/TLR4/
NF-κB pathway.
Anticancer activity Apple pomace HT29 cells Apple oligosaccharides Reduction in cell viability by inducing apoptosis [107]
(China) (0.01–1 mg/mL) and cell cycle arrest through inhibiting formation
of Cdk 2-cyclin A1 complex by decreasing Cdk 2
levels
Antifatigue activity Apple pomace Male Kunming mice, AP = 50, 100, 200 mg/kg/ Significantly reduction in biochemical parameters [33]
(China) weighing 20 g day involving BUN levels (10.23–13.87 mmol L− 1), LA
levels (3.50–5.14 mmol L− 1), and LDH levels
(2711.54–3640.11 UL− 1) which are indicators of
fatigue
DPPH = 2,2-diphenyl-1-picrylhydrazyl, SOD = Superoxide dismutase, HO• = Hydroxyl radical, FRAP = ferric reducing antioxidant capacity, O2•− = Superoxide
radical, CCl4 = Carbon tetrachloride, BUN = Blood urea nitrogen, LA = Lactic acid, MDA = Malondialdehyde, ALT = Alanine aminotransferase, GSH-Px = Glutathione
peroxidase, AST = aspartate aminotransferase, HFD=High fat diet, TYAP = Thinned young apple polysaccharide, LDH = lactate dehydrogenase, LPS = Lipopoly
saccharide, AP = Apple polysaccharide, CRC = colorectal cancer, MAP = Modified apple polysaccharide, MES = mesalamine.
Fig. 3. Representation of biological activities of the polysaccharides extracted/isolated from apple.
treatment. The tablets were designed to release the medication at the MAP with probiotics would provide a successful treatment for UC with
site of the colon, and the combination of MES and MAP provided com site-specific drug release in the colonic milieu [87].
plete protection against acetic acid (0.2–0.3 mL) induced UC in Sprague
Dawley rats of either sex. It was found that when tablet come in contact
with rat caecal content, a burst release of drug was observed (45.70 %) 6.4. Improving gastro-intestinal health
in the 6th hour which lead to 38 % drug release (p < 0.05) (i.e., between
5th h and 6th h). The effect of microbiota on the release of drug from the In the last ten years, improving human health by regulating the
polysaccharide-based matrix as well as coat can be clearly understood by microbiome has become a useful and promising technique. The micro
observing the drug release at the 10th h which was 74.80 % and at 24th biota that lives on the skin’s surface and in the mucosa are diverse and
h when it was found to be 102.20 %. The study found that rats treated numerous, and they can be used to cure illness or maintain good health
with the combination of MES and MAP mini tablets and probiotics had [90]. As a result of advancement in science and technology, scientists
significant improvements in colonic weight (p < 0.001), DAI (p < have discovered that it is possible to use dietary or non-dietary in
0.001), macroscopic score (p < 0.001), and histopathology, demon terventions to affect the makeup and metabolic traits of these microbial
strating an effective treatment for UC. Additionally, MAP was found to populations [91]. In a study, the effect of AP on gut dysbiosis in rats fed
have anti-ulcer properties, but it was concluded that co-administering with high-fat diet (HFD) for 14 weeks was investigated. The study
showed that AP treatment increased the abundance of Bacteroidetes and
10
Lactobacillus, while reducing Fusobacterium and Firmicutes. The treat of Wnt pathway in colon tissues was observed, following the treatment
ment also increased short chain fatty acids, specifically isobutyric and with 10 % (w/v) AP; suggesting that AP may prevent CACC in least part
acetic acid. Moreover, the induction of autophagy in goblet cells by AP by regulating the Wnt pathway and intestinal flora disorder [93]. It was
leads to a reduction in gut permeability, which results in a significant observed that MAP (dose = 1.25–5 %; duration = 13 weeks) influences
decrease in plasma lipopolysaccharide-binding protein (LBP) levels. the expression of MUC-1 (target in colorectal cancer) to prevent colitis-
Additionally, AP administration up-regulates Occludin while down- associated carcinogenesis in mice [38]; inhibition of growth of MCA-38
regulating inflammatory markers such as IL-1, TNF-α, CXCL-1 and cells via downregulating the expression of IL-22 and upregulating the
MCP-1, which are commonly associated with chronic inflammation and expression of IL-22BP [85].
dysbiosis-induced gut permeability [92]. Moreover, a separate research
study was conducted to explore the mechanisms by which AP manages 6.6. Anti-fatigue activity
microbiota imbalances and to assess its potential as a prebiotic. In a
study, Li et al. [93] investigated how AP affected Bifidobacterium longum Fatigue is a complex physiological condition characterized by a
and Lactobacillus rhamnosus. They administered AP to the two probiotics decrease in overall body efficiency and an inability to sustain specific
to determine if it could be used as a prebiotic in vitro. The study found physiological or predetermined exercise levels due to excessive physical
that AP did not have a direct impact on promoting the growth of pro activity [95,96]. In recent years, there has been a growing interest in the
biotics. However, quantitative polymerase chain reaction (qPCR) anal potential anti-fatigue properties of polysaccharides, as evidenced by
ysis indicated that AP treatment resulted in increased levels of numerous studies [97–100]. Recently, researchers conducted an inves
Lactobacillus and Bifidobacterium in the fecal flora of rats. Gram staining, tigation focused on polysaccharides extracted from apple pomace using
plate counting, and growth curve tests indicated that AP might consid acid and alkali extraction methods, namely ACAPP and ALAPP [33].
erably increase the diversity of fecal microbiotas. In conclusion, the These polysaccharides were tested as anti-fatigue agents through
authors discovered that AP was first metabolized followed by its utili exhaustive swimming exercise (ESE) in Male Kunming mice (wt. 20 g)
zation by E. coli and the produced metabolic by-products helped (dose (50 mg/kg/day, 100 mg/kg/day, 200 mg/kg/day). The study re
B. longum to grow [93]. Study by Calvete-Torre et al. [19] suggests that ported that mice treated with polysaccharides demonstrated increased
galactose content and (Arabinose + Galactose): Rhamnose in apple swimming duration (p < 0.05), with endurance times ranging from
pomace as well as rhamnose and galactose content in pectin, were 18.61 to 23.69 min, significantly longer than the control group’s
positively associated to the promotion of commensal microbes when endurance time of 14.67 min. Additionally, the ingestion of these
apple pomace and apple pectin were tested on human microbiota from polysaccharides had a significant impact on various biochemical pa
healthy subjects and inflammatory bowel disease patients. Overall, rameters in mice, including blood urea nitrogen (BUN), lactate dehy
tested fractions caused growth of beneficial microbes including Pre drogenase (LDH), and lactic acid (LA) levels which are indicators or
votella, Lachnospiraceae UCG-010, Akkermansia, Sucinivibrio, Turicibacter, related with fatigue [101–105]. The researchers observed that APP
Lachnospiraceae CAG-56, Blautia, Eubacterium eligens, Dialister and significantly reduced BUN levels (10.23–13.87 mmol L− 1), LA levels
Intestinimonas [19]. The findings of the above-mentioned study suggest (3.50–5.14 mmol L− 1), and LDH levels (2711.54–3640.11 UL− 1).
that AP provides another basis to understand the health benefits of apple Notably, mice in the mid and high dose groups displayed substantial
for improving intestinal microbiota that is associated with many chronic improvements in fatigue compared to the control group, and ALAPP was
diseases. found to be more effective than ACAPP, consistent with the ESE results.
This study provides promising evidence of the potential anti-fatigue
6.5. Anti-cancer activity effects of polysaccharides, particularly those extracted from apple
pomace [33]. Further study in this area may pave the way for the
Growing evidence has proved that inflammation plays significant in development of effective anti-fatigue supplements or treatments for in
occurrence and progression of cancer. According to epidemiological dividuals facing fatigue-related challenges.
research, persons with UC have a much higher chance of acquiring
colorectal cancer (CRC) than normal people. However, the mechanism 7. Value added products
underlying the progression from UC to CRC is still unclear. Numerous
clinical researches have revealed that the colon tissues of UC and CRC According to the actual projection, population growth will continue
patients exhibited higher levels of macrophage infiltration than those of until it reaches probably around 10 billion in 2050. This puts a lot of
healthy individuals. This shows that macrophages are linked to UC and pressure on the agri-food market to provide alternative sources of food
CRC, therefore it is logical to assume that controlling macrophage that should satisfy dietary needs as well as the current trend of
function would have positive impacts on preventing colitis associated consuming healthy and sustainable foods. However, the agro-food in
colorectal cancer (CACC). Sun et al. [94] conducted a study to investi dustries generate vast quantities of food waste, which can have detri
gate the impact of AP on macrophage polarization in Raw 264.7 cells mental effects on the environment due to their instability and high
and a CACC mice model induced by AOM and DSS. The findings indi moisture content. This leads to the creation of greenhouse gas emissions
cated that AP had the ability to prevent dextran sodium sulfate/azoxy and nitrogen pollution in both water and soil [108–110]. In light of this,
methane (DSS/AOM)-induced carcinogenesis and maintain a moderate Gustafsson et al. [12] conducted a novel and eco-friendly study to pro
inflammatory state in the colon. Additionally, AP shifted macrophage duce sustainable biomaterials from apple pomace. The apple pomace
polarization towards the M1 phenotype. The in-vitro study suggested used in the study contains free sugars (55.47 %) and a water-insoluble
that AP upregulated toll-like receptor (TLR)-4 signaling, triggering the fraction composed of hemicelluloses (29.42 %), cellulose (38.99 %),
transition of macrophages towards the M1 phenotype. Furthermore, the and lignin (22.94 %). The researchers employed solution casting and
TLR-4 antagonist, TAK-242 (resatorvid), suppressed the AP-induced compression molding techniques to create bio-based films and 3D ob
polarization of macrophages [94]. These results indicate that AP has jects (specifically, fiberboards), respectively. By leveraging the self-
potential to treat and prevent CRC. Moreover, in another study the binding capability of polysaccharides, the study demonstrated that the
protective effect of AP on CACC via regulation of microbial dysbiosis in naturally occurring components present in the apple pomace exhibited a
mice (duration: 15 weeks) was investigated. It was found that AP stronger plasticizing effect, resulting in the creation of more flexible and
treatment protected against CACC effectively. AP increases the micro interconnected films with significantly higher elongation (37.39 % and
biota diversity (Lactobacillus and Fusobacterium). The macrophages and 55.41 %, respectively) and tensile strength ranging from 3.02 to 5.79
number of T-cells in the colon tissue of mice was reduced, while sup MPa and an elongation of 0.93–1.56 %. In conclusion, the study’s
pression in nuclear aggregation of β-catenin and inhibition in activation findings demonstrated the potential applications of apple pomace
11
biomaterials, including their use in edible/disposable tableware or food Furthermore, polysaccharides exhibit various other beneficial proper
packaging [12]. Asforementioned earlier, maximum amount of pomace ties, including anti-ulcer, hepatoprotective, antioxidant, and microbial
is a waste generated by apple juice processing industry. Apple pomace diversity-improving effects, addressing a range of health disorders.
mainly consists of polyphenols and pectin polysaccharides. A patent has Galacturonic acid has been identified as a key element responsible for
been filed by Wang et al. [111] involving the utilization of apple pomace the observed bioactivities. It was stated that structural features,
polysaccharide in preparation of yoghurt. The invention adds in suitable composition etc. were closely associated with biological activities, thus
pomace polysaccharide in yoghurt production raw material. The poly modified polysaccharides were used to enhance their biological effects
saccharide addition improves the organoleptic quality, acidity, retention in various in vivo and in vitro studies. However, certain research gaps
ability and texture characteristics of Yoghourt. Additionally, it was still exist, including the investigation of toxicity, clinical studies, and
found out that APs are able to reduce the fermentation temperature, additional biological activities such as antidiabetic, immunomodula
process energy consumption of the yoghurt, and it can be economically tory, and anti-microbial effects, which warrant further exploration. To
beneficial. However, apple pomace is used for the preparation of variety fully unlock the value-added opportunities of apple polysaccharides,
of food products like jam, jelly, sauce, drinks, beverages, fermented particularly in functional foods, more in-depth in vivo studies are
products, ethanol, animal feed [112], but there is no direct report of necessary. Additionally, advancements in the fields of nutraceuticals,
utilizing apple pomace pectin/polysaccharides in food by-products. pharmacology, and cosmetics offer promising avenues for the utilization
Thus, there is need to investigate and develop functional foods by of apple polysaccharides in various applications. In conclusion, this re
using APs, an alternative way to fulfill the requirements of healthy diet. view serves as a foundation for the future exploration and application of
Moreover, numerous research studies have consistently demon apple polysaccharides. As the research progresses, their potential for use
strated the beneficial effects of pectin polysaccharides extracted from in diverse industries, including nutraceuticals, pharmaceuticals, and
various sources on overall health [113–115]. Pectin polysaccharides cosmetics, will undoubtedly be unlocked, paving the way for exciting
contain probiotic proteins that support proper intestinal function, aid in possibilities in the near future. Further investigation is essential to fully
toxin elimination, and protect the mucous membrane of the colon. harness the benefits of these natural compounds and propel them to
Additionally, they possess antioxidant, anticancer, antibacterial, anti- wards practical and beneficial applications.
inflammatory, and immunomodulatory properties [116–118]. A spe
cific study highlighted the potential of pectin extracted from industrially
obtained apple pomace as a gelatin substitute [118]. It was reported that Declaration of competing interest
the extracted pectin extracted by metal ion precipitation technique
exhibited a composition of 60.38 % glucose, 19.41 % mannose, 17.60 % The authors declare that they have no known competing financial
galactose, and 1.00 % rhamnose, along with a low amount of acidic interests or personal relationships that could have appeared to influence
sugar (1.61 % glucuronic acid) and a degree of methoxylation (DM) the work reported in this paper.
value of 12.5 %. The low percentage of acidic sugar relative to the total
monosaccharides suggested a highly branched structure of apple pectin Data availability
(AP). When Ca2+ ions were added, AP demonstrated remarkable gelling
ability upon cooling its heated solution to a low temperature (4 ◦ C). No data was used for the research described in the article.
However, no gel was formed at room temperature (25 ◦ C) or in the
absence of Ca2+. At a fixed pectin concentration (0.5 %, w/v), AP References
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Science and Postharvest Technology Indian Agricultural Research Institute, New
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International Journal of Biological Macromolecules 255 (2024) 128011

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International Journal of Biological Macromolecules

Exploring apple pectic polysaccharides: Extraction, characterization, and

1. Introduction increasingly frequent extreme climatic shifts, driven by both natural

International Journal of Biological Macromolecules 255 (2024) 128011

Antioxidant Apple peel (Shaanxi, DPPH •

Fig. 3. Representation of biological activities of the polysaccharides extracted/isolated from apple.

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