Food Research International 119 (2019) 359–368

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Food Research International

journal homepage: www.elsevier.com/locate/foodres

Silver and gold nanoparticles based colorimetric assays for the T

determination of sugars and polyphenols in apples
⁎
Annalisa Scroccarello, Flavio Della Pelle, Lilia Neri, Paola Pittia, Dario Compagnone
Faculty of Bioscience and Technology for Food, Agriculture and Environment, University of Teramo, 64023 Teramo, Italy

A R T I C LE I N FO A B S T R A C T

Keywords: In this work, the exploitability of rapid and easy to use methods for the evaluation of the antioxidant capacity
Rapid methods (AOC) and sugars content (SC), through metal nanoparticles (MNPs) formation, has been proved and applied to
Sugars assay apples. In particular, an AgNPs-based sugar quantification assays and an AuNPs-based polyphenols antioxidant
Polyphenols assay capacity assay have been used as tools for the evaluation of apple extracts composition. Both assays are based on
Apple samples
the ability of the analytes (sugars and polyphenols) to reduce the source of metal (Au3+ and Ag+), stabilizing, at
Antioxidant capacity
Metal nanoparticles
the same time, the resulting MNPs colloidal suspensions. The AuNPs and AgNPs formation depends on the
Ag nanoparticles analyte structure and concentration, resulting in red (AuNPs) and yellow (AgNPs) colored suspensions. Both
Au nanoparticles assays require an initial mixing step, followed by MNPs formation under mild conditions (10 min, room tem-
perature or 45 °C), and the colorimetric response is easily acquired at a fixed wavelength (AgNPs: 430 nm and
AuNPs: 540 nm). The analytical performance of both assays has been proven, obtaining good reproducibility
(RSD ≤ 6%,), sensitivity (LODs ≤8.7 μmol L−1) and recoveries (91% −113.7%). The produced MNPs (AgNPs
and AuNPs) have been characterized using UV–Vis spectroscopy and TEM, and the cross-reactivity between
assays, as well as the possible endogenous interferents, studied. The assays have been tested on 42 apple samples,
and the data obtained compared with those obtained by conventional methods (i.e. FC, ABTS, and ion chro-
matography). The proposed AgNPs-sugars assay gives results comparable (R = 0.915) to those determined by
ion chromatography in terms of total sugars, and the AuNPs-polyphenols assay results able to assess the poly-
phenols antioxidant capacity, being correlated with those obtained by the ABTS method (R = 0.922). This
MNPs-based approach demonstrated to be an excellent tool for rapid and facile analysis of sugars and poly-
phenols in apple samples.

1. Introduction products. Despite, their substantial difference in terms of implication

Nowadays the accessibility of easy and fast analytical methods for
quality assurance is highly required; in fact, rapid quality control plays
a key role along the entire food production chain. In addition to con-
ventional instrumental analytical protocols, there is the need for com-
plementary screening assays, possibly environment-friendly (e.g. small
amount of solvent employed, low waste generation), easy to be used
even by unspecialized staff.
In recent years, the growing awareness of the relationship between
food and health has increased consumer concern about the composi-
tion, processing and health properties of food products. This en-
couraged the food industry to explore new food sources, technology and
formulations in order to improve and diversify the food productions
and to meet consumers' dietary needs. In this context, particular at-
tention has been reserved to sugars (SGs) and polyphenols (PCs) in food
and impact on health, both classes of compounds affect the quality and
stability of the food, their functional and sensory characteristics as well
as their technological suitability towards processing.
PCs are a heterogeneous class of compounds characterized by a
phenolic hydroxyl group(s); they are ubiquitous secondary metabolites
present in plant foods (El Gharras, 2009). These compounds have been
received exceptional attention in the last decades; in fact, PCs provide
foods an added value for their well-known health benefits, their tech-
nological role and also marketing (Della Pelle & Compagnone, 2018). In
these last years, main efforts have been devoted to assess the real PCs
health effect through in vitro and particularly in vivo studies, with
particular attention to bioavailability issues (Del Rio et al., 2013;
Pandey & Rizvi, 2009; Peluso, Miglio, Morabito, Ioannone, & Serafini,
2015; Serafini & Peluso, 2016; Zhang & Tsao, 2016). It has been proven
that long term consumption of diets rich in plant polyphenols is

⁎
Corresponding author.
E-mail address: dcompagnone@unite.it (D. Compagnone).

https://doi.org/10.1016/j.foodres.2019.02.006
Received 3 November 2018; Received in revised form 3 January 2019; Accepted 3 February 2019
Available online 06 February 2019
0963-9969/ © 2019 Elsevier Ltd. All rights reserved.
A. Scroccarello et al.
inversely associated with the risk of developing diseases linked to oxi-
dative stress (Del Rio et al., 2013; Di Mattia, Sacchetti, Mastrocola, &
Serafini, 2017; Pandey & Rizvi, 2009; Zhang et al., 2015; Zhang & Tsao,
2016), offering, thus, protection against the development of cancer,
cardiovascular diseases, diabetes, osteoporosis and neurodegenerative
diseases (Del Rio et al., 2013; Joseph, Edirisinghe, & Burton-Freeman,
2016; Pandey & Rizvi, 2009; Serafini & Peluso, 2016). Beside the bio-
logical activity, PCs play other important roles in food due to their
coloring properties, stabilizing effect towards oxidation reactions
(Shahidi & Ambigaipalan, 2015) and antimicrobial properties against
pathogenic and spoilage bacteria (Hayek, Gyawali, & Ibrahim, 2013).
However, PCs content, depending on food type, processing and storage
conditions, and end users, can account for either positive or negative
features. PCs have been shown to contribute to the definition of food
sensory characteristics, to strongly affect the rate of enzymatic
browning (Persic, Mikulic-Petkovsek, Slatnar, & Veberic, 2017), to in-
fluence the emulsification processes and the physical and chemical
stability of dispersed emulsified systems (Di Mattia et al., 2015;
Giacintucci, Di Mattia, Sacchetti, Neri, & Pittia, 2016) and to interact
with proteins. This latter phenomenon resulting from interactions of
tannins with salivary proteins is responsible for astringency perception,
formation of haze and precipitates in beverages, and inhibition of en-
zymes and reduced digestibility of dietary proteins (El Gharras, 2009;
Soares, Brandão, Mateus, & de Freitas, 2017). Moreover, PCs in some
plants can be used as maturation index due to their accumulation
during ripening (Garrido et al., 2016).
Official methods for the PCs quantification and their antioxidant
capacity (AOC) evaluation, being the latter needed due to the multiple
mechanisms of action of bioactive compounds, are not officially es-
tablished, probably because a single method cannot cover all the de-
sired functional proprieties that need to be assessed for a particular
purpose (Della Pelle & Compagnone, 2018). PCs qualitative and quan-
titative evaluation is performed classically by high-performance liquid
chromatography (HPLC) coupled with Uv-Visible (Uv-Vis), diode array
(DAD), or mass spectrometry (MS) detector (Ignat, Volf, & Popa, 2011;
Simeoni et al., 2018). Very often the PCs evaluation aim is to give an
estimation of the PCs content or an overall AOC evaluation. Thus, Folin-
Ciocalteu (FC) assay (Singleton & Rossi Jr, 1965), evaluation of the
scavenging activity against radical compounds (DPPH• and ABTS• as-
says) (Brand-Williams, Cuvelier, & Berset, 1995; Re et al., 1999), re-
duction of metal ions (FRAP and CUPRAC assays) (Apak, Güçlü,
Özyürek, & Karademir, 2004; Benzie & Strain, 1996) and competitive
methods (ORAC and TRAP) (Cao, Verdon, Wu, Wang, & Prior, 1995;
Miller, Rice-Evans, Davies, Gopinathan, & Milner, 1993), as well as
electrochemical approaches (Del Carlo et al., 2012; Della Pelle &
Compagnone, 2018; Hoyos-Arbeláez, Blandón-Naranjo, Vázquez, &
Contreras-Calderón, 2018; Hoyos-arbeláez, Vázquez, & Contreras-
calderón, 2017), are widely employed (Antolovich, Prenzler, Patsalides,
McDonald, & Robards, 2002; Ignat et al., 2011; López-Alarcón &
Denicola, 2013).
Presence and concentration of SGs can also be used as a quality and
process marker in foods. The quantity of SGs gives information on the
suitability of cultivation practices, fruit maturation at harvest, post-
harvest and overall fruit quality (Alonso-Salces et al., 2005; Beckles,
2012; Carbone, Giannini, Picchi, Lo Scalzo, & Cecchini, 2011;
Khanizadeh et al., 2008; Ma et al., 2014; Magwaza & Opara, 2015; Neri
et al., 2016; Palazzo, Facchini, & Mallardi, 2012; Wu et al., 2007).
Moreover, several reactions driven by sugars (e.g. fermentation, hy-
drolysis, caramelization, Maillard reactions, crystallization) may occur
in food processing and affect products shelf-life. In addition, SGs con-
tent is strictly related to foods sweetness, caloric content and glycemic
index or glycemic load with the latter repeatedly and independently
associated with diabetes and other chronic diseases (Dhurandhar &
Thomas, 2015; Scazzina et al., 2016).
Much effort has been devoted to developing sugars content eva-
luation methods in foods. A classical empiric method for the evaluation
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Food Research International 119 (2019) 359–368
of soluble solids related to the sugars content is based on the use of a
refractometer. It's a rudimental technique commonly used for Brix de-
gree determination in horticultural products and beverages. Nowadays,
chromatography approaches, in particular, ion chromatography (IC)
and capillary electrophoresis, coupled with different detectors are
commonly employed for the SGs qualitative and quantitative analysis
(Downes & Terry, 2010; Lacourse, 2002; Ma et al., 2014; Martínez
Montera, Rodríguez Dodero, Guillén Sánchez, & Barroso, 2004; Peters,
Levine, & Jones, 2001). Besides nuclear magnetic resonance (NMR)
(Pospíšilová, Polášek, Šafra, & Petriška, 2007) can be used for the SGs
determination, whereas near-infrared spectroscopy (NIRs) analysis can
be employed for fruits ripening evaluation during their growing/culti-
vation steps (Magwaza et al., 2012; Magwaza & Opara, 2015). More-
over, the determination of total reducing sugars can be performed by
Fehling reaction and other optical assays (Dubois, Gilles, Hamilton,
Rebers, & Smith, 1956; Quisumbing & Thomas, 1921).
The reported methods to assess PCs and SGs in foods can results
time-consuming, require several steps (extraction, clean up, running
time, titrations at high temperature, precipitations, etc.) and large
amounts of solvents, as well as expensive instrumentation. Recently
analytical nanomaterials-based approaches have become a valuable
strategy to assess a plethora of analytes using different analytical ap-
proach, allowing improvement of sensitivity and reducing the time of
analysis, and, then, resulting in sample and reagents significant vo-
lumes reduction (Blandón-Naranjo et al., 2018; Capoferri, Della Pelle,
Del Carlo, & Compagnone, 2018; Del Carlo et al., 2012; Della Pelle
et al., 2016; Della Pelle et al., 2017; Della Pelle et al., 2018; Della Pelle
& Compagnone, 2018; Della Pelle, Del Carlo, Sergi, Compagnone, &
Escarpa, 2016; Pérez-Lòpez & Merkoci, 2011; Rojas et al., 2018; Valdés,
Angela, Calzón, & Díaz-garcía, 2009).
Among nanomaterials, noble metal nanoparticles (MNPs) possess a
peculiar feature named ‘localized surface plasmon resonance’ (LSPR),
that allows the interaction with electromagnetic radiation (Jain, Huang,
El-Sayed, & El-Sayed, 2007; Mayer, Hafner, & Antigen, 2011;
Petryayeva & Krull, 2011; Willets & Van Duyne, 2007). In fact, under
the influence of electromagnetic radiation in the visible range, the
MNPs electrons of surface atoms can easily move through vacant or-
bitals generating absorption at a particular wavelength. This MNPs
property results in a typical absorption spectrum, widely exploited to
realized plasmonic-based methods and assay (Della Pelle &
Compagnone, 2018; Jain et al., 2007; Özyürek, Güngör, Baki, Güçlü, &
Apak, 2012; Palazzo et al., 2012; Petryayeva & Krull, 2011; Scarano,
Pascale, & Minunni, 2017; Vasilescu, Sharpe, & Andreescu, 2012;
Vilela, González, & Escarpa, 2012; Vilela, González, & Escarpa, 2015;
Willets & Van Duyne, 2007). MNPs-based methods turn out to be ex-
tremely modular and tunable towards specific analytes. In fact, changes
in metal source, MNPs shape, MNPs dimensions, or capping agent
employed and reaction media, lead to substantial reactivity changes
able to elicit (towards target analytes) or avoid (towards potential in-
terfering compounds) a specific class of compounds reactivity.
Recently, our group have pointed out the capacity of PCs (Della
Pelle et al., 2015; Della Pelle et al., 2015; Della Pelle et al., 2018) and
SGs (Della Pelle, Scroccarello, Scarano, & Compagnone, 2018) to form
MNPs, thanks to their reducing power and capability to support/sta-
bilize the MNPs formation.
The study here reported aims to demonstrate the suitability of two
assays based on the selective formation of different metal nanoparticles,
to assess the polyphenols antioxidant capacity and the total sugars in
apples. The reactivity of AgNPs selectively synthesized by sugars and
AuNPs produced by polyphenols has been studied and the formed MNPs
characterized. Hence, an AgNPs-based sugar quantification assays and
an AuNPs-based polyphenols antioxidant capacity assay have been
optimized and tested on 42 apple samples. The MNPs-based data were
in good agreement with the analysis of sugar content and antioxidant
capacity carried out by conventional methods. The final aim of this
work is to demonstrate the real samples applicability of these MNPs-
A. Scroccarello et al.
based assays, overcoming definitely the proof of applicability, in order
to provide to the ‘foods science community’ new effective and easy to
use tools able to asses sugars and polyphenols in apple samples.
2. Materials and methods
2.1. Reagents and stock solutions
All the chemicals were of analytical reagent grade. Epicatechin,
catechin, and phlorizin were purchased from Extrasynthese (Genay,
France). Gallic acid, chlorogenic acid, fructose, glucose, sucrose, in-
ositol, trehalose, mannitol, raffinose, sorbitol, xylitol, and xylose,
Cetyltrimethylammonium chloride (CTAC; 25% in water), hydrogen
tetracholoroaurate (HAuCl4·3H2O, 99.9%), silver nitrate
(AgNO3, > 99%), 2,2-azino-bis(3- ethylbenzothiazoline-6-sulphonic
acid) (ABTS), sodium hydroxide(NaOH), sodium carbonate (Na2CO3),
Folin & Ciocalteu's reagent, sodium phosphate mono basic mono-
hydrate ACS reagent (NaH2PO4·H2O) sodium phosphate dibasic anhy-
drous (Na2HPO4), methanol, acetonitrile and formic acid were pur-
chased from Sigma-Aldrich (St Louis, MO, USA). Polyphenol standards
stock solutions were prepared in methanol at a concentration of
1.0 × 10−2 mol L−1 and stored at −20 °C in the dark. Stock solutions of
sugar standards were prepared in water at a concentration of
1.0 × 10−2 mol L−1 and stored at +4 °C in the dark. Glucose stock
solutions were prepared 24 h before use and stored at +4 °C in the dark
to allow mutarotation. Milli-Q water (18.2 MΩ) was used for all the
experiments and stock solutions preparation.
2.2. Apparatus
Samples were shaken and centrifuged with an orbital shaker (SSL1,
Stuart equipment, Belfast, UK) and an ALC4237R refrigerated cen-
trifuge (ALC Intl., Cologno Monzese, Italy), respectively. For extracts
drying a Rotavapor LABOROTA 4000, Heidolph Instruments
(Schwabach, DE) was used. For silver nanoparticles assay the thermo-
stated mixing was carried out with a VDRL 711/CT orbital shaker from
Asal (Florence, Italy). For the gold nanoparticles assay, the reaction mix
was heated in a water bath using a 720 D thermostat digital group
(Asal, Italy). Absorbance measurements were performed using a
JENWAY 6400 spectrophotometer from Barloworld Scientific
(Staffordshire, UK,). Sugars were determined by ICS 3000 Ionic
Chromatograph Dionex (San Donato Milanese, Italy) equipped with an
ICS 3000 SP pump and an ICS 3000 ED detector. Transmission electron
microscopy measurements (TEM) were carried out using a Zeiss EM10C
transmission electron microscope. Apple samples were brought to dry
in a ScanVac CoolSafe 95–15 Pro freeze dryer (Lynge, Denmark).
2.3. Samples
Apples of two different varieties (‘Golden Delicious’ and ‘Royal
Gala’) were purchased in local markets from different batches and
seasons. The sampled batch was selected in order to randomize the
apple composition, trying to maximize the differences of color, size,
weight and ripening degree. A total of 42 samples was collected in the
year 2017 (samples from 1 to 28, Golden Delicious) and 2018 (samples
from 29 to 35, Golden Delicious; samples from 36 to 42 Royal Gala).
2.4. Preparation of extracts for polyphenols analysis
Extracts from apples collected in 2017 have been prepared as fol-
lows: apples were preliminarily peeled, the core removed by a corer and
diced (~1 cm3) and immediately freeze-dried. To this aim, aliquots of
apple cubes were preliminarily arranged in Petri dishes and frozen at
−40 °C for 24 h. Apple freeze-drying was carried out for 72 h at a
pressure of ≈ 0.5 hPa and by setting the shelves temperature at 0 °C.
After freeze-drying samples were stored at room temperature in a
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Food Research International 119 (2019) 359–368
desiccator in the dark until analysis. An aliquot of 0.5 g of freeze-dried
sample was weighted and 5 mL of acetone/water solution (70:30 v/v)
added. The mixture was homogenized with an Ultra-Turrax T18 basic
homogenizer (IKAR Werke GmbH & Co. KG, Staufen, Germany) for
1 min at 13500 rpm. Thus, the solution was centrifuged for 5 min at
3000 rpm at +4 °C. After centrifugation, the supernatant was separated
from the solid pellet and the recovered solution was filtered with a
0.45 μm MDI cellulose filters (Ambala, India). Finally, the supernatant
was dried by rotavapor and recovered with 1 mL of methanol.
Extracts from apples collected in 2018 have been prepared as
follow: apples were preliminarily peeled, the core removed by a corer
and diced (~1 cm3). An aliquot of 20 g was weighted and 100 mL of
acetone/water solution (70:30 v/v) was added. The mixture was
homogenized with Ultra-Turrax T18 basic homogenizer for 1 min at
13500 rpm and centrifuged for 5 min at 3000 rpm at +4 °C. The su-
pernatant was, then, separated from the solid pellet and filtered with an
MDI 0.25 μm PTFE filter (Ambala, India). Finally, the supernatant was
dried by rotavapor and recovered with 1 mL of methanol.
All the extracts were stored at −20 °C°C in the dark until analysis.
2.5. Preparation of extracts for sugars analysis
Extracts from apple samples were obtained according to Neri et al.
(2016) with some modification. Apples were preliminarily peeled, the
core removed by a corer and diced (~1 cm3). Each sample was
weighted (5 g) and diluted with 20 mL of distilled water. The dispersion
was finely ground and homogenized with an Ultra-Turrax T18 basic
homogenizer (IKAR Werke GmbH & Co. KG, Staufen, Germany) for
2 min. The dispersion was preliminarily shaken for 20 min at +4 °C,
then centrifuged at 5000 rpm for 10 min at +4 °C in a refrigerated
centrifuge. Then, the supernatant was filtered through a 0.45 μm nylon
filter (MDI, Ambala, India) and stored at +4 °C under dark conditions
until use.
2.6. Total polyphenols determination (Folin-Ciocalteu method)
20 μL of a properly diluted apple extract or phenolic standard was
added to 20 μL of Folin-Ciocalteu reagent and stirred for 3 min. Then
400 μL of sodium carbonate (Na2CO3, 7.5%) and deionized water were
added up to the final volume of 1000 μL. The solution was stirred for
60 min, at room temperature in the dark and the total polyphenol was
determined with a spectrophotometer by reading the absorbance at
760 nm. Gallic acid standard solutions were used to calibrate the
method.
2.7. Radical scavenging activity determination (ABTS)
The total AOC was evaluated by the ABTS method. ABTS reagent
stock solution was prepared according to Re et al. (1999). The radical
ABTS·+ solution was diluted to reach 0.70 ( ± 0.02) absorbance value
at 734 nm and directly used for the assay. An appropriate volume of
sample or standard/phenolic compounds was diluted up to the final
volume of 2000 μL with ABTS reagent; then the reaction mix was stored
in the dark, at room temperature for 5 min. Afterward, the absorbance
at 734 nm was determined and the sample or standard antioxidant
mediated ABTS·+ shutdown was compared with the respective blank/
control prepared without phenolic compounds or sample addition.
Gallic acid standard solutions were used to calibrate the method.
2.8. Ion chromatography
Chromatographic analyses of glucose, fructose, sorbitol, sucrose,
trehalose, and maltose were conducted by an ICS 3000 Ionic
Chromatograph Dionex (San Donato Milanese, Italy). The chromato-
graphic separation was performed with a CarboPac PA20 column
(3 mm × 150 mm, Dionex) equipped with a guard column (CarboPac
A. Scroccarello et al.
Fig. 1. (A) MNPs spectra obtained with fructose (50 μmol L−1), glucose (50 μmol L−1) and sorbitol (40 μmol L−1) subjected to the AgNPs-sugars assay. (B) MNPs
spectra obtained with gallic acid (10 μmol L−1), chlorogenic acid (10 μmol L−1) and epicatechin (5 μmol L−1) subjected to the AuNPs-polyphenols assay. Inset to each
figure is reported the colorimetric response obtained with increasing concentration of fructose (A) and chlorogenic acid (B), corresponding to AgNPs (A) and AuNPs
(B) colloidal solutions, respectively.
PA20, 3 mm × 30 mm, Dionex) according to Neri, Hernando, Pérez-
Munuera, Sacchetti, and Pittia (2011) with some modifications. The
following work condition was used: NaOH 5.0 × 10−2 mol L−1 as
mobile phase, a flow rate of 0.5 mL min−1, a 35 min run at a column
temperature of 30 °C, and a volume of injection of 10 μL. The sugar
detection was performed using the time/potential waveform A, as re-
commended by the Dionex technical manual. Sugars identification and
quantification were carried out using retention times and the related
sugar calibration curve, respectively.
2.9. Silver nanoparticles assay (AgNPs-sugars)
An AgNPs-based colorimetric assay (AgNPs-sugars) was used for the
total sugars quantification, according to Della Pelle, Scroccarello,
Scarano, and Compagnone (2018) with some modifications. All the
reagents of the reaction were used at room temperature. For the AgNPs
formation were used 5 μL of CTAC (1.0 × 10−3 mo L−1), 25 μL of an
AgNO3 solution (2.0 × 10−2 mol L−1), appropriate dilution of sugars
standard or sample and finally 10 μL NaOH (5.0 mol × L−1) to start the
reaction. Then, the reaction mix was brought to 500 μL with water. The
AgNPs formation was then carried out at 25 °C for 10 min under stirring
in a thermostated orbital shaker. Finally, the reaction was stopped in
ice where it was kept for 10 min. The absorbance due to the AgNPs
formed was recorded at 430 nm. All the measurements were carried out
against the blank (reaction mix without standard or sample reacted).
For each sugar standard and sample extract, a dose-response curve was
obtained by adding increasing amounts of standards/extract and
reading the absorbance at the LSPR maximum. Glucose standard solu-
tions were used to obtain a calibration curve and quantification pur-
poses.
2.10. Gold nanoparticles assay (AuNPs-polyphenols)
AuNPs based colorimetric assay (AuNPs-polyphenols) was used for
the polyphenols AOC evaluation, according to Della Pelle, Vilela, et al.
(2015), with some modifications. For the AuNPs formation 10 μL of
CTAC (8.0 × 10−4 mol L−1) as capping agent, 25 μL of HAuCl43H2O
solution (2.0 × 10−2 mol L−1) as gold source and an appropriate
amount of polyphenol standard or sample extract in a final volume of
500 μL, reached with phosphate buffer solution (PB pH 8.0;
1.0 × 10−2 mol L−1) were used. The reaction mix was stirred for 2 min
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with an orbital shaker, followed by heating for 10 min in a water bath at
45 °C; the reaction was then blocked in ice for 5 min. Finally, the ab-
sorbance of the newly formed AuNPs was recorded at 540 nm. All the
measures were conducted against a blank (reaction mix without the
reducing agent). Moreover, for each sample extract, a dose-response
curve was obtained by adding an increasing amount of extract and
reading the absorbance at the LSPR maximum. Gallic acid standard
solutions were used to obtain a calibration curve and quantification
purposes.
2.11. AgNPs-sugars and AuNPs-polyphenol assays recovery and potential
interfering compounds
A mix of the two apple samples extract of a different variety was
spiked with SGs and PCs standards in order to obtain solutions of 20, 40
and 60 μmol L−1 and 5, 10 and 10 μmol L−1, respectively. The fortified
samples were separately subjected to the AgNPs-sugars and AuNPs-
polyphenols assay and compared with the relative non-spiked samples.
Recoveries were calculated using the following eq. (1):
[(analyte concentration fortified sample − analyte concentration
unfortified sample)/analyte concentration added] × 100.
The possible interference effects of apple extract components such
as sugars, polyols, polyphenols, and organic acids were studied. To this
aim different amounts of the possible interfering compounds were
spiked on samples to evaluate the effect on the respective MNPs assay.
The concentration of the interferents tested were chosen according to
literature data (Alonso-Salces et al., 2005; Carbone et al., 2011; Hecke
et al., 2006; Khanizadeh et al., 2008; Ma et al., 2014; Neri et al., 2016;
Palazzo et al., 2012; Wu et al., 2007).
3. Results and discussion
3.1. Reactivity of sugars (AgNPs) and polyphenols (AuNPs) standards
towards metal nanoparticles formation
The aim of this study was to use colorimetric assays based on AuNPs
and AgNPs formation for the determination of SGs and polyphenolic
AOC in apples. Thus, the reactivity of standard polyphenols and sugars
was initially investigated.
An example of the visible spectra obtained, following the AgNPs-
assay (Section 2.9) and AuNPs-assay (Section 2.10) protocols are
A. Scroccarello et al. Food Research International 119 (2019) 359–368

Table 1
Analytical parameters obtained by sugars and polyphenols standards dose-response curves by using the AgNPs-sugars and AuNPs-polyphenols assays.
Linear range Linear equationa Slopea Intercepta Determination coefficient

μM y = Abs and x = μM std.dev. std.dev. R2

AgNPs-sugars assay
Glucose 10–90 y = 0.0098x + 0.0639 ± 4.2 × 10−4 ± 2.7 × 10−3 0.998
Fructose 10–90 y = 0.0101x + 0.0421 ± 5.3 × 10−4 ± 2.2 × 10−3 0.996
Sorbitol 20–80 y = 0.0100x + 0.0319 ± 4.4 × 10−4 ± 1.3 × 10−3 0.999

AuNPs-polyphenols assay
Chlorogenic acid 3–25 y = 0.0204x − 1.813 ± 1.2 × 10−3 ± 1.1 × 10−1 0.992
Epicatechin 1–7.5 y = 0.0229x − 1.012 ± 1.3 × 10−3 ± 6.1 × 10−2 0.995
Gallic acid 1–15 y = 0.0216x − 0.9237 ± 1.1 × 10−3 ± 4.6 × 10−2 0.991

a
Mean value of three dose-response curves.

Fig. 2. MNPs spectra obtained with increasing amount of apple extract subjected to the AgNPs-sugars (A) and AuNPs-polyphenols (B) assays. Inset to each figure at
the left is reported the colorimetric response corresponding to each spectrum. Apple extracts volume employed: (A) 5 (a), 10 (b), 15 (c) μL of sample 11 diluted 100-
fold; (B) 20 (a), 40 (b), 60 (c) μL of sample 11 diluted 2-fold. Inset to each figure at the right is reported a TEM micrograph corresponding to the AgNPs (A) and AuNPs
(B) formed with the respective assay.

reported in Fig.1, together with a picture of the suspensions obtained by
increasing concentrations of fructose and chlorogenic acid (insets). The
LSPR maximum (LSPRmax) obtained (employed as analytical signal) for
the AgNPs and AuNPs generated was in all the cases 430 ± 10 nm
(A430) and 540 ± 10 nm (A540), respectively, in accordance with the
literature (Della Pelle, Scroccarello, Scarano, & Compagnone, 2018;
Della Pelle, Scroccarello, Sergi, et al., 2018). Table 1 reports the ana-
lytical parameters obtained by the dose-response curves, achieved with
an increasing concentration of SGs and PCs vs. the respective MNPs
LSPRmax.
In all the cases, a reproducible (RSD ≤ 6%, n = 3) linear dose-re-
sponse range was recorded, with a good determination coefficient
(R2 ≥ 0.991). Both assays exhibited a quite low limit of detections
(LOD) of ≤8.7 μmol L−1 and ≤ 3.3 μmol L−1 for the AgNPs-sugar and
AuNPs-polyphenols assays, respectively. Moreover, the MNPs suspen-
sions result stable for 60 min at room temperature, instead if stored in
ice the stability reach the 6 h. This can be attributed to the synergistic
stabilization of analytes and capping agent resulted in MNPs formation
in mild conditions, with no aggregation, collapse, and precipitation.
These phenomena are typical drawbacks MNPs-based assays when ab-
sorbance at LSPRmax is used as analytical signal.
3.2. AgNPs-sugars and AuNPs-polyphenols assays applied to apple extracts:
MNPs characterization, recovery and interferents studies
In order to make the apple polyphenols and sugars analysis faster
and easier, the usability of rapid and effective assays, based on the
MNPs formation, have been demonstrated. Thus, to prove the exploit-
ability of the AgNPs-sugars and the AuNPs-polyphenols assays, 42 dif-
ferent apple samples (Section 2.4) and different procedures of extrac-
tion/pretreatment (2.5 and 2.6) have been carried out.
The TEM micrographs reported corresponding in Fig. 2 are related
to AgNPs (Fig. 2A, right inset) and AuNPs (Fig. 2B, left inset) formed
using directly apple extracts, following the reported procedures. Sugars
from apples were able to generate well-defined and dispersed spherical
AgNPs (ϕ < 10 nm); phenolic compounds, as well, gave rise to well-
defined and dispersed spherical AuNPs with a ϕ < 25 nm.
Fig. 2 also reports the absorbance spectra and the corresponding
colloidal suspensions obtained by increasing amounts of apple extracts,
for both the AgNPs (Fig. 2A) and the AuNPs assay (Fig. 2B). These
spectra clearly demonstrate that sugars and polyphenols in apples are
able to generate quantitatively AgNPs and AuNPs, respectively. The
apple extracts produced AgNPs and AuNPs having the maximum ab-
sorbance intensity band (LSPRmax) at 430 ± 9 nm and 540 ± 7 nm,
respectively, thus, totally in accordance with the LSPRmax obtained by
the pure compounds reported in Section 3.1. This allowed fixing the
reading at 430 (AgNPs-sugars) and 540 nm (AuNPs-polyphenols)
avoiding the run of the absorbance spectrum for each sample.
To further confirm the ability of the proposed MNP-based assays to
assess, in a quantitative way, SGs and PCs compounds, a mix of dif-
ferent extracts of apples have been spiked with different amount of
fructose and glucose in the 20–60 μmol L−1 range and with chlorogenic

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Table 2
Results of phenolic content and antioxidant properties evaluated by the AuNPs-polyphenols assay, ABTS and FC method of 42 apple extracts.
Sample AuNPS-polyphenols ABTS Folin-Ciocalteu

−1 a −1 a
(mg Kg ) RSD (%) (mg Kg ) RSD (%) (mg Kg−1)a RSD (%)

1 131.6 ± 6.0 3.0 155.8 ± 4.7 4.5 167.8 ± 5.4 3.2

2 127.8 ± 6.3 5.9 120.1 ± 7.1 4.9 117.4 ± 8.3 7.1
3 97.3 ± 9.5 7.7 92.3 ± 7.1 9.7 73.6 ± 6.0 8.1
4 130.0 ± 9.2 2.4 142.0 ± 3.4 7.1 171.2 ± 11.5 6.7
5 109.8 ± 5.0 6.0 111.4 ± 6.7 4.5 156.3 ± 10.6 6.8
6 120.0 ± 5.7 7.2 118.3 ± 8.6 4.8 138.5 ± 6.5 4.7
7 138.5 ± 6.6 4.2 132.9 ± 5.6 4.7 137.6 ± 9.6 7.0
8 94.1 ± 4.3 7.9 83.5 ± 6.6 4.6 83.3 ± 4.4 5.3
9 129.4 ± 7.0 5.1 144.5 ± 7.4 5.4 162.4 ± 1.6 1.0
10 128.8 ± 6.9 2.9 155.5 ± 4.5 5.3 177.9 ± 10.9 6.1
11 155.8 ± 7.1 6.4 174.3 ± 11.1 4.6 193.5 ± 11.6 6.0
12 110.1 ± 5.1 2.4 105.8 ± 2.5 4.6 153.9 ± 1.6 1.1
13 127.9 ± 6.2 3.1 159.1 ± 4.9 4.9 177.9 ± 11.5 6.4
14 132.6 ± 7.1 5.0 144.8 ± 7.3 5.4 170.1 ± 9.5 5.6
15 162.9 ± 23.0 3.6 201.6 ± 7.2 14.1 261.0 ± 13.7 5.2
16 172.2 ± 9.8 2.4 197.9 ± 4.7 5.7 194.4 ± 11.4 5.9
17 143.4 ± 7.8 2.9 174.4 ± 5.1 5.5 229.8 ± 6.4 2.8
18 184.2 ± 9.9 3.1 201.2 ± 6.3 5.4 227.2 ± 5.1 2.2
19 152.0 ± 12.3 3.9 161.1 ± 6.3 8.1 155.1 ± 5.1 3.3
20 164.8 ± 7.5 1.5 197.3 ± 2.9 4.6 206.0 ± 9.5 4.6
21 157.7 ± 12.7 3.4 159.8 ± 5.4 8.1 204.6 ± 9.0 4.4
22 91.0 ± 7.4 2.6 98.4 ± 2.6 8.1 78.8 ± 2.8 3.6
23 142.3 ± 8.2 1.9 162.2 ± 3.1 5.7 155.4 ± 6.4 4.1
24 144.2 ± 6.5 3.1 153.2 ± 4.7 4.5 143.1 ± 4.1 2.9
25 136.6 ± 7.3 2.4 145.7 ± 3.5 5.3 172.4 ± 12.4 7.2
26 141.9 ± 9.5 1.7 171.9 ± 2.9 6.7 184.5 ± 6.7 3.6
27 164.8 ± 9.6 1.6 165.7 ± 2.6 5.8 229.0 ± 11.8 5.1
28 108.2 ± 5.9 2.6 111.9 ± 2.9 5.5 91.4 ± 4.7 5.1
29 112.3 ± 7.1 1.1 103.5 ± 1.1 6.3 106.1 ± 6.8 6.4
30 107.6 ± 5.4 0.5 106.5 ± 0.6 5.0 103.3 ± 0.3 0.3
31 100.8 ± 6.4 0.5 87.9 ± 0.5 6.3 95.2 ± 1.7 1.8
32 134.6 ± 4.7 1.4 119.2 ± 1.7 3.5 96.0 ± 1.3 1.4
33 128.9 ± 3.3 3.8 114.7 ± 4.3 2.5 87.6 ± 2.7 3.1
34 149.6 ± 3.6 2.4 153.2 ± 3.7 9.8 107.2 ± 6.3 5.8
35 156.3 ± 2.9 1.8 157.7 ± 2.0 1.8 204.7 ± 5.3 2.6
36 170.8 ± 3.9 2.3 182.0 ± 4.2 2.3 177.7 ± 13.3 7.5
37 160.0 ± 5.8 1.2 166.4 ± 2.0 3.6 207.6 ± 7.7 3.7
38 99.1 ± 5.3 2.5 102.0 ± 2.6 5.3 90.8 ± 2.4 2.6
39 125.8 ± 3.6 0.3 118.6 ± 0.3 2.9 84.3 ± 3.5 4.1
40 127.6 ± 11.7 0.7 126.7 ± 0.9 9.2 85.0 ± 3.2 3.8
41 154.5 ± 11.3 2.0 148.3 ± 3.0 7.3 159.3 ± 5.8 3.6
42 128.9 ± 4.2 2.2 126.7 ± 2.8 3.2 82.1 ± 1.5 1.9

a
Mean value, n = 3. The results are expressed with respect to the weight of fresh apple.

acid and epicatechin in the 5–10 μmol L−1 range. Satisfactory recovery
values (from 91% to 113.7%) were obtained.
Selectivity of the assays was evaluated by testing different potential
interfering compounds. The type and concentration of the interfering
compounds was chosen according to literature data (Alonso-Salces
et al., 2005; Carbone et al., 2011; Hecke et al., 2006; Khanizadeh et al.,
2008; Ma et al., 2014; Neri et al., 2016; Palazzo et al., 2012; Wu et al.,
2007). The AuNPs-polyphenols assay was tested against different sugars
(fructose, glucose, sucrose, inositol, trehalose, mannitol, raffinose,
sorbitol, xylitol, and xylose) in a broad concentration range (from
5.0 × 10−4 mol L−1 to 1.5 mol L−1). None of the reported sugars was
able to form AuNPs in the experimental conditions used.
In addition, the possible sugars ‘perturbation’ during the AuNPs
formation was also studied using real samples. To this aim, two apple
extracts belonging to different varieties, were mixed and spiked with
chlorogenic acid to obtain a final concentration of 50 μmol L−1; the mix
was further fortified with fructose, glucose, sucrose and their mix (60%
fructose, 30% glucose, 10% sucrose) in a 5.0 × 10−4 mol L−1 to
1.5 mol L−1 concentration range. None effect, on the obtained colori-
metric response, was again observed on the AuNPs assay. The se-
lectivity of the AuNPs-based assay towards polyphenols was achieved
using the selected medium of reaction. In fact, at pH 8.0 sugars are not
ionized; ionization in strong alkaline conditions, for these compounds,
is required to reduce the Au3+ and at the same time surround/stabilize
the formed AuNPs, in the times required by this assay. This behavior is
related to the synergic action of sugars and capping agent (CTAC), that
are able to act as ‘charge carriers’, around the AuNPs, because of their
ionizable moieties. These results confirm the data reported by different
authors (Della Pelle, Scroccarello, Sergi, et al., 2018; Filippo, Serra,
Buccolieri, & Manno, 2010; Palazzo et al., 2012). It should be also
noticed that above pH 10 in the medium used for the assay, the AuNPs
are formed without the use of external reducing compounds, impairing
the possible quantification of sugars.
Ascorbic acid in principle can form MNPs due to the reducing ac-
tivity, however, both AuNPs and AgNPs based assays, have been opti-
mized in order to react, with sugars and polyphenols, respectively, and,
thus, the effect is minimal. Preliminary experiments run with AuNPs
assay have demonstrated that reactivity of ascorbic acid is about 250
times lower than polyphenols (e.g. chlorogenic acid). The content of
ascorbic acid in our sample extracts (measured on 10 random samples)
was found in each case below 50 ppm. Thus, considering concentration
and reactivity ascorbic acid does not interfere in the AuNPs based
analysis of polyphenols. The same considerations apply for AgNPs assay
for sugars since the difference in concentration of analytes is much
larger.
The AgNPs-sugars assay was tested against polyphenols.

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Table 3 3.3. Comparison with ABTS, FC and ion chromatography

Results of sugars content by the AgNPs-based assay and ion chromatography of
42 apple extracts. The quantification of the sugars and the polyphenolic AOC eva-
Sample AgNPS-sugars Ion chromatography Relative error luation was then carried out on 42 samples of apples by applying both
the MNPs-based proposed assays and the conventional methods.
(g Kg−1)a RSD (%) (g Kg−1)a RSD (%) (%)b Table 2 reports the concentration of polyphenols obtained from the
analyses performed by the different methods expressed as gallic acid
1 146.3 ± 2.4 3.5 136.5 ± 1.6 2.2 7.2
2 129.3 ± 2.7 3.5 130.2 ± 1.8 2.4 +0.7 equivalents. According to the AuNPs-based assay the polyphenolic
3 60.1 ± 1.2 0.7 77.6 ± 4.1 3.2 −22.6 content ranged from 91.9 to 230.8 mg Kg−1; good reproducibility has
4 155.4 ± 2.2 3.4 146.7 ± 1.4 2.0 +5.9 been obtained for all the samples (RSD ≤ 7.9%). Using the FC assay,
5 129.0 ± 4.1 5.4 128.7 ± 0.9 1.1 +0.3
usually used to obtain the total PCs content, data ranged from 73.5 to
6 166.9 ± 0.5 0.8 149.6 ± 1.6 2.5 +11.6
7 181.2 ± 3.4 6.1 179.5 ± 1.1 1.9 +1.0
261.0 mg Kg−1 (RSD ≤ 8.1%). The ABTS assay, related to the scaven-
8 129.2 ± 0.8 1.0 122.1 ± 3.0 3.7 +5.8 ging activity of the apple extracts, gave data from 83.5 to 201.6 mg
9 89.3 ± 4.7 4.2 85.4 ± 5.1 3.3 +4.5 Kg−1 with acceptable reproducibility (RSD ≤ 14.1).
10 122.9 ± 3.5 4.3 147.8 ± 2.9 1.0 −16.8 Table 3 reports the data obtained with the AgNPs-sugars assay and
11 87.5 ± 1.3 1.0 105.3 ± 6.4 1.7 −26.4
the ion chromatographs, being in the first case the sugar content ex-
12 141.2 ± 4.0 5.7 165.3 ± 2.4 2.6 −14.6
13 212.0 ± 3.2 6.9 198.5 ± 3.1 2.6 +6.8 pressed as glucose equivalents. The analysis carried out by the ion
14 133.9 ± 1.1 1.5 127.0 ± 2.3 3.0 +5.4 chromatography highlighted that in all the samples the sugars found in
15 149.8 ± 0.4 0.6 150.3 ± 2.2 3.3 −0.3 higher concentration are glucose and fructose (their sum 75–85%);
16 107.6 ± 6.2 6.7 97.5 ± 4.1 4.0 +10.3
lower amounts were determined for sucrose (10–20%), trehalose and
17 131.1 ± 3.3 4.3 122.6 ± 2.1 2.6 +6.9
18 140.7 ± 2.1 3.0 137.8 ± 1.5 2.0 +2.1
sorbitol (their sum 2–5%) (data not shown). In Table 3 the data have
19 182.9 ± 2.4 4.4 193.3 ± 1.7 3.2 −5.4 been expressed as the sum of the concentration of all the mono- and di-
20 189.9 ± 1.4 2.6 195.5 ± 1.5 3.0 −2.9 saccharides, each one calculated by the relative calibration curve. The
21 150.2 ± 2.5 3.7 138.2 ± 1.0 1.4 +8.6 data of the AgNPs-sugars assay and the ion chromatography ranged
22 185.8 ± 0.4 0.8 168.5 ± 1.2 2.0 +10.2
between 60.1 and 250.1 g Kg−1 and 77.6–245.3 g Kg−1, respectively.
23 119.1 ± 2.3 2.7 144.1 ± 1.4 2.0 −17.3
24 121.4 ± 3.8 4.6 148.7 ± 1.7 2.5 −18.4 The proposed AgNPs-sugars method results reproducible
25 144.0 ± 1.9 2.7 177.7 ± 1.7 3.0 −18.9 (RSD ≤ 6.9%), as well as, as expected, ion chromatography
26 161.4 ± 3.3 5.4 161.0 ± 2.5 4.0 +0.3 (RSD ≤ 4%). Overall, the obtained data are in agreement with the SGs
27 184.8 ± 0.5 0.9 179.0 ± 0.6 1.0 +3.2
and PCs content in apples reported in the literature (Alonso-Salces
28 98.5 ± 0.5 0.5 107.7 ± 2.4 2.5 −8.5
29 198.4 ± 2.3 4.7 206.4 ± 1.3 2.7 −3.9
et al., 2005; Carbone et al., 2011; Khanizadeh et al., 2008; Ma et al.,
30 173.5 ± 3.4 5.9 184.0 ± 1.7 3.2 −5.7 2014; Neri et al., 2016; Wu et al., 2007).
31 250.1 ± 1.5 3.8 245.3 ± 1.1 2.6 +2.0 Linear correlations among the methods, employed for the poly-
32 135.2 ± 4.2 5.6 149.3 ± 2.0 3.0 −9.4 phenols and sugars evaluation, are reported in Fig. 3. The AuNPs-
33 170.3 ± 2.1 3.5 168.4 ± 1.1 1.8 +1.2
polyphenols assay results well correlated with the ABTS method
34 165.4 ± 1.9 3.2 198.3 ± 1.4 2.8 −16.6
35 124.8 ± 3.7 4.6 126.8 ± 2.5 3.2 −1.6 (R = 0.922) (Fig. 3A), while a lower correlation was found with the FC
36 88.1 ± 7.7 6.8 124.6 ± 2.0 2.5 −29.3 method (R = 0.764). Hence, the AuNPs-assay gives an estimation of the
37 109.5 ± 2.7 2.9 129.5 ± 2.5 3.3 −15.4 antioxidant capacity, and this ability is related to the intrinsic me-
38 181.3 ± 1.7 3.1 195.7 ± 1.1 2.1 −7.3 chanism of formation and stabilization of the AuNPs, that results re-
39 164.4 ± 1.1 1.9 149.8 ± 1.8 2.8 +9.8
40 177.7 ± 3.6 6.4 152.4 ± 1.7 2.6 +16.6
lated to the polyphenols quantitative and qualitative (intrinsic anti-
41 108.5 ± 2.4 2.6 107.2 ± 2.0 3.2 +1.3 oxidant power) composition. The relationship of the results of the FC
42 191.8 ± 3.3 6.2 201.3 ± 0.8 1.6 −4.7 method vs. those obtained by the AuNP-based assay, as well as for the
a
ABTS assay, results in a slight data overestimation and this behavior
Mean value, n = 3. The results are expressed with respect to the weight of was already reported in the literature (Escarpa & Gonzalez, 2010;
fresh apple.
b Escarpa & González, 2001; Sanchez-Rangel, Benavides, Heredia,
Relative error calculated using the data obtained with ion chromatography
Cisneros-Zevallos, & Jacobo-Velazquez, 2013). FC is definitively non-
as the true value.
selective and in apples, the overestimation can be attributed to the
reducing sugars presents.
Chlorogenic acid, catechins and phlorizin, and their equimolar mix
Fig. 3B shows the linear correlation trend between the data obtained
have been tested, with a similar approach on the apple sample extract
by AgNPs-based assay and those by ion chromatography. The AgNPs-
(5.0 × 10−4–3.0 mmol L−1 range); also in this case, no interference
sugars assay presented a good correlation with the ion chromatographic
was found. Indeed, the ratio between sugars and antioxidant com-
data (R = 0.915), as well as a good quantitative correspondence be-
pounds in apples results strongly in favor of sugars (750/1000 fold)
tween the obtained data with a relative error between +11.6 and
resulting in high selectivity vs. polyphenols. These results confirm the
−29.3% (Table 3). Despite the different principle of the methods, the
ability of AgNPs to be formed by sugars in a selective way even in a
AgNPs-assay is able to allow a quantitative estimation of the total su-
complex matrix (Della Pelle, Scroccarello, Scarano, & Compagnone,
gars present in samples. Among the MNPs-based methods, none of them
2018).
can be used for sugars contents evaluation. Several methods, both for
Eventually, the influence of organic acids present in apples has been
polyphenols and sugars analysis need high temperatures, long times,
evaluated in both assays by adding malic acid, succinic acid, citric acid,
large volumes of sample and solvent. Moreover, many methods require
tartaric acid, oxalic acid, quinic acid, and shikimic acid. In all the cases
a seed-mediated MNPs growth that requires different steps, which
these compounds resulted not able to form MNPs or to interfere with
greatly complicates the application. For these reasons, very few are the
the assays proposed, taking into account the concentrations found in
methods easily exploitable directly for the evaluation of food con-
apples and reported in the literature (Atona & Ova, 2004; Hecke et al.,
stituents by not expert personnel. Moreover, the employed MNPs-based
2006; Samukelo et al., 2006; Wu et al., 2007).
methods require low amount of sample (from 2.5 to 60 μL of apple
extract), do not require particular solvents (apart the solvent used for
the extraction) neither the use of radicals. The equipment needed is
very simple as a photometer and the colorimetric output allows easily a

365
A. Scroccarello et al.
Fig. 3. (A) Correlation curve between the data obtained, analyzing the 42 apple extracts, with the proposed AuNPs-polyphenols assay and ABTS method
(y = 0.662x + 40.69; R = 0.922). (B) Correlation curve between the data obtained, analyzing the 42 apple extracts, with the proposed AgNPs-sugars assay and ion
chromatography (y = 0.986x − 2.17; R = 0.915).
qualitative assessment. Furthermore, the AgNPs-based sugars evalua-
tion requires 5 min vs. ion chromatography that needs around 1 h. On
the other hand, the AuNPs-based method for polyphenols evaluation
requires an analysis time of 10 min, comparable with the ABTS assay
(5 min), and is significantly faster with respect to FC (about1 h). Fur-
thermore, in term of sensitivity, robustness, and reproducibility the
proposed assays appear totally comparable with the existing methods or
result even more sensitive as for the AgNPs-based assay with respect to
ion chromatography. Finally, it should be pointed out that the whole
assays take place directly in an Eppendorf tube and that the reagents
used are stable and non-volatile. These features can allow the devel-
opment of partially automated assay kits for on-site detection.
4. Conclusions
In this study, the ability of metal nanoparticles to act as optical
nanoprobes, able to allow rapid and efficient detection of the total
sugars and polyphenol antioxidant capacity in apples has been de-
monstrated. In particular, the evaluation of SCs and polyphenols AOC
have been carried out by using the formation of AuNPs and AgNPs with
no problems of selectivity. MNPs assays reactivity has been compared
with the results obtained by conventional methods, and the AgNPs-
sugars assay gives comparable results (R = 0.915) to ion chromato-
graphy when the total sugars concentration is taken into account while
the AuNPs-polyphenols assay is able to assess the polyphenols anti-
oxidant capacity of apple extracts, with a significant correlation with
the results obtained by the ABTS method (R = 0.922). The ease of
formation of these nanoprobes (mild condition, 5–10 min) and the
simplicity of the detection (absorbance read at a fixed wavelength),
makes these probes attractive in industrial applications for quality
control. In addition, the proposed MNPs do not require radical com-
pounds, and the solvent use and waste produced are significantly lower
compared to classical methods. This approach appears, thus, promising
for rapid monitoring of AOC and SCs in food.
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