Received: 20 January 2020 Revised: 4 May 2020 Accepted: 4 May 2020

DOI: 10.1002/cem.3251

SPECIAL ISSUE - REVIEW

PhotoMetrix and colorimetric image analysis using

smartphones

Fernanda Carla Böck1 | Gilson Augusto Helfer2 | Adilson Ben da Costa2 |

~o1,3
Morgana Bazzan Dessuy1 | Marco Flôres Ferra

1
Instituto de Química, Universidade
Federal do Rio Grande do Sul, Av. Bento
Abstract
Gonçalves, Porto Alegre, RS, 9500, Brazil The steady advances in technology employed in smartphones, coupled with
2
Universidade de Santa Cruz do Sul,, Av. the high availability and the ease access to these devices, increased the interest
Independência, Santa Cruz do Sul, RS,
in applying it to analytical purposes. The main goal is to develop simple, fast,
2293, Brazil
3 and low-cost analytical methods, generating immediate results. Different ana-
Instituto Nacional de Ciência e
Tecnologia-Bioanalítca (INCT- lytical strategies using smartphones have been proposed to identify or deter-
Bioanalítica), Cidade Universitária, mine different analytes in a wide range of matrices. Some of them use the
Zeferino Vaz s/n, Campinas, SP, Brazil
smartphone to capture the images, to process them, and to provide the analyti-
Correspondence cal results. Others use this device just for image acquisition. In this context,
Fernanda Carla Böck, Instituto de this review aimed to relate previous works that used smartphones to acquire
Química, Universidade Federal do Rio
Grande do Sul, Av. Bento Gonçalves, images for analytical purposes, with a special focus on PhotoMetrix applica-
Porto Alegre 9500, RS, Brazil. tion, which is a tool for chemical analysis. This application allows the decom-
Email: fernanda.c.bock@gmail.com
position of digital images acquired by smartphone cameras and to process
them within the same device, allowing in situ chemical analysis.

KEYWORDS
colorimetry, digital image analysis, PhotoMetrix®, smartphone

1 | INTRODUCTION

Smartphones are increasingly common in people's daily lives. It is estimated that about 5.1 billion people use a mobile
device, which is equivalent to about 67% of the world's population.1 The constant technological advances in mobile
phones, along with the high availability of these devices, allow them to be exploited for functions beyond traditional
communication. Several articles report the increasing advance using smartphones in various areas, such as food analy-
sis, biosensors, and medical, using different forms of detection (fluorescent, chemiluminescent, colorimetric, and
electrochemical).2–10
This review aimed to relate previous works that used smartphones to acquire images for analytical purposes, with a
special focus on PhotoMetrix application. Figure 1 presents researches based on this principle, and they are organized
according to year of publication and the processing method within the smartphone or outside the smartphone. We con-
sidered that processing data in the smartphone means that the capture of images, decomposition, processing, and gen-
eration of results were carried out by the smartphone itself, using specific applications. From this research,
125 references were compiled, from which 59 were processed embedded in the smartphone and 66 externally. Figure 1
does not show works that used PhotoMetrix application (a total of 17), which will be presented in more details below.
In Figure 1, we can see that one of the pioneers in the use of smartphones was Martinez et al.11, who used images
taken by the camera's device to quantify glucose and protein in artificial urine samples. In the following year, Yao Lu

Journal of Chemometrics. 2020;e3251. wileyonlinelibrary.com/journal/cem © 2020 John Wiley & Sons, Ltd. 1 of 19
https://doi.org/10.1002/cem.3251
2 of 19 BÖCK ET AL.

FIGURE 1 Timeline of works using image analysis by smartphone. Source: Scopus April 2020

et al.12 presented a methodology based on smartphone imaging for immunoassays, and David Breslauer et al.13 demon-
strate automated bacillus counting via image analysis. From 2011, the use of these devices begins to be more explored,
and the first applications for image processing on the phone itself were reported.14–17
By evaluating the works that used smartphones for image acquisition and performed the external processing of the
acquired data (Figure 1), we can see that these devices are explored in several areas, such as the detection of diseases
and markers of diseases causing in the medical field, immunoassay,18–27 quality control of the environment,28–39 foren-
sic analysis,40–42 control and quality of beverages and food,15,43–64 quality evaluation of pharmaceutical products,54,65,66
fuel quality control,67,68 pigments,69 chemical education, kinetic studies, and chemical receptors.70–77
In Figure 1, it is possible to see that the number of works that use smartphone for acquisition, processing, and
results generation by the device is relatively small than the works that use the device for image acquisition and perform
external processing. As previously mentioned, the works that used smartphones for the whole process are from several
areas, such as colony counting,78 sensors, biosensors, immunoassays and clinical test,79–85 quality control of environ-
mental parameters (soil, air, and water),86–90 beverage, and food analysis. 91–99
It is important to highlight that although there is a considerable amount of work that develops every procedure on
the smartphone, many mobile applications are specific to certain samples or analytes, which does not make using these
applications accessible. 100–115
In this context, the PhotoMetrix® application was developed in order to provide a free tool that can be used in vari-
ous analysis, with different analytes and samples, as well as enabling univariate and multivariate analysis in the same
application. The application was initially developed for the Android, IOS, and Windows operating systems; however, in
its evolution, the focus was directed to the Android platform, as it is the most popular and easiest to maintain system.
Therefore, the tool is available for free download, although with the restriction of the multivariate calibration tool for
IOS and Windows systems, as it is an analysis tool recently included.
BÖCK ET AL. 3 of 19

2 | PHOTOMETRIX®

The PhotoMetrix application was developed in 2015 with the purpose of offering a tool for chemical analysis from the
decomposition of digital images acquired by smartphone cameras and processed within the same device, allowing its
use for in situ analysis. Figure 2 shows the evolution of the application since its development, when in its first version
allowed only univariate analysis (calibration) and multivariate analysis (principal component analysis [PCA])116.
In the second developed version, denominated PhotoMetrix PRO®, registered in INPI (Instituto Nacional da
Propriedade Industrial),117 two tools were included in multivariate mode, hierarchical cluster analysis (HCA), and par-
tial least squares (PLS). The third version, PhotoMetrix UVC®, has been adapted for external camera use in image
acquisition. The first version was discontinued, and the other two versions are still available and freely accessible.
The application allows to perform image decomposition in two ways, one based on the red (R), green (G), and blue
(B) primary color addition model, where the histogram based on the RGB model is performed. Another way is from the
independent decomposition of RGB channels and RGB derived color models such as hue, saturation, and value (HSV);
hue, saturation, and lightness (HSL); and hue, saturation, and intensity (HSI).118 Hue determines the color as described
by wavelength, for example, the distinction between red and yellow. Saturation is the amount of the color that is pre-
sent, for instance, the distinction between red and pink. Lightness, intensity, or value is the amount of light, the distinc-
tion between a dark red and light red or between dark gray and light grey.119 Figure 3 shows the application interfaces
of PhotoMetrix application. Figure 3A presents the application's initial screen, where there are analysis options (univar-
iate and multivariate), settings, and application information. Figure 3B shows the settings interface, where the user can
set parameters such as number of samples, region of interest size, whether to use smartphone flash, graphics edit set-
tings, and options for saving data. By selecting the “univariate analysis” option at Figure 3A, the next step is to define
the decomposition form of the images (Figure 3C), which has the options “Multiple channel” and “Vector RGB.” After
selecting the segregation form, the univariate analysis interface is presented (Figure 3D), where it is possible to build
the calibration curves, sampling, and save the results.
Figure 4 presents the options to perform multivariate analysis (Figure 4A). In the multivariate module (Figure 4B),
three analysis options are possible: PCA, PLS, and HCA. When selecting the PCA option, a new screen is displayed
(Figure 4C), where it is possible to select between options for “sampling,” “re-processing,” viewing “saved results,” and
“help.” To perform the sampling, some parameters must be set (Figure 4F), such as number of samples, method for
decomposing images, and adding a name to the model after which images can be acquired; the data reprocessing is per-
formed on the same screen (Figure 4F). To build the PLS model, Figure 4D is presented, with the “calibration,”
“reprocessing,” “sampling,” and “saved results” options, as well as for PCA, the analysis parameters are set on
Figure 4F. To perform the HCA, the application leads directly to Figure 4E to select the color model that will be used in
the decomposition.
To highlight how important the exploitation of applications on mobile devices is and should be further explored,
Figure 5 presents a map of uses until the February 2020 period of the PhotoMetrix application. We can see in the
upper-left corner of Figure 5 that this application presented 13 535 uses until this period, showing the importance of
developing this type of tool in science, enabling teachers to insert new methodologies in classrooms, and providing an
alternative to schools that do not have infrastructure conditions for higher cost laboratory equipment.

F I G U R E 2 PhotoMetrix
application evolution timeline
4 of 19 BÖCK ET AL.

F I G U R E 3 PhotoMetrix application interfaces: (A) home screen, (B) settings screen, (C) choice of color model for univariate analysis,
and (D) interface for univariate analysis

F I G U R E 4 PhotoMetrix module for multivariate analysis: (A) home screen, (B) multivariate analysis options (PCA, PLS, and HCA),
(C) PCA module, (D) PCA and PLS processing options, (E) PLS module, and (F) HCA module

Table 1 presents the published works that used PhotoMetrix® in their different versions, demonstrating which sam-
ple and analyte were evaluated, the analysis tool used, and some analytical features. In addition, after Table 1, a short
summary about the development of each work is discussed.
BÖCK ET AL. 5 of 19

FIGURE 5 Map of uses of the PhotoMetrix application around the world, up to February 2020

In the work developed by Grasel et al.120, the PhotoMetrix® was used to separate six varieties of commercial tannin
extracts, using PCA as an exploration tool. The samples studied in this work were black wattle, quebracho, tara, chest-
nut, myrobalan, and valonea. The authors used five different lots for each tannin species, resulting in a total of 30 sam-
ples. The extracts were arranged in a polyethylene bottle, compressed, with the purpose of capturing images with the
same parameter for all samples. The images were captured with an apparatus made from a cardboard box, with a hole
for the phone camera, and the illumination was performed with an ultraviolet lamp (25 W). The PCA was built through
the H, S, V, L, and I color variables. The results obtained in the digital image analysis agreed with the results obtained
by infrared and ultraviolet evaluated in previous work by the authors, thus showing the ability of color separation using
an easily accessible, inexpensive tool like a camera, and an application from a smartphone.
PhotoMetrix® was also used by de Oliveira,121 with the purpose of identifying cocaine and quantifying some adulter-
ants (levamisole, procaine, and caffeine). The tests were performed by colorimetric reactions, and the results obtained
by the app were compared with those obtained by spectrophotometry in the visible region of the spectrum. For the
acquisition of images, samples were arranged in a black box, with a fixed LED strip containing nine white light emitters
in it. The analysis was performed using the univariate module present in the app. The results obtained through the
analysis of images for quantification of caffeine and levamisole were not satisfactory, although they presented good
results in the build of calibration curves; in the quantification, they showed to suffer interference from the presence of
cocaine, which could not be overcome even using chemometric tools. For the determination of cocaine and procaine
quantification, the results obtained by the app were similar to those obtained by spectroscopy, thus showing the feasi-
bility of using a simple tool, such as the smartphone, to perform preliminary analysis, including in situ analysis.
A spot test was developed by Soares et al.122 for indirect determination of iodine value in biodiesel by using digital
images. The analysis is based on the iodine consumption during the halogenation reaction of unsaturated compounds
in biodiesel. The procedure shown in Figure 6 was based on adding iodine (24 μl) in a biodiesel sample (40 μl), mixing
it and then transferring 5 μl of this mixture to a paper, so a starch solution was added for complexation. Images were
taken for univariate analysis using PhotoMetrix®. The papers containing the mixtures were placed on a white sheet,
positioned in an environment with controlled lighting provided by two tube lamps. The results obtained with digital
TABLE 1 Summary of works using PhotoMetrix
6 of 19

Detection target Analysis Analytical features Reference

RSD
R2 LOD LOQ CV (%) (%) RMSEC RMSEP

Tannins PCA - - - - - - - Grasel et al.120

Levamisole, procaine, and Calibration curve 0.99811a/0.99607b/0.99663c/0.9796d 49a/2.47b/1.51c/43.10d mg - 1.10a/1.46b - - - de Oliveira121

−1
caffeine in cocaine (univariate L
analysis)

Iodine in biodiesel Calibration curve 0.980 8 g I2/100 g biodiesel - 4.9 - - - Soares et al.122
(univariate
analysis)

Acid-base titrations acetic acid Calibration curve 0.99 - - <8 - - - Nogueira

in vinegar samples (univariate et al.123
analysis)

Ethanol in sugar cane spirit Calibration curve 0.9963* 0.19% (v/v) 0.62% (v/v) <4 <0.5 0.1073 0.0798 Böck et al.124
(PLS)

Thermal stability of raw milk Calibration curve 0.9917* - - ≤5 - 0.261 0.0980 Helfer et al.125
(PLS)

Total dissolved solid content in Calibration curve 0.9907* - - - - 0.1483 - Bento and de
mango (PLS) Sousa
Dantas126

Detection of wax content in PCA calibration 0.9095 2.14% - - <3 - - Yulia et al.127
gorengan curve
(univariate
analysis)

Lactase enzyme in Calibration curve 0.986 0.0006 mg 0.002 mg - 3-13 - - Dionizio

pharmaceutical formulation (univariate et al.128
analysis)

Total dissolved iron and Calibration curve 0.9999e/ 0.9999f/0.9999g/0.9999h 0.15e/0.029f/0.72g/h mg L−1 0.50e/0.097f/2.40g/h mg L−1 - <10e/ - - Lumbaque
hydrogen peroxide (PLS) <7f/ et al.129
g
determination in water <8 /
<5h

Quantifying proteins in human Calibration curve 0.984i/0.980j/0.989k 0.6i/0.7j/0.7k μg 1.9i/2.0j/2.4k μg - - - - de Jesus

serum (univariate et al.130
analysis)

Determination of methanol in Calibration curve 0.990l/0.994m 10 mg kg−1 l/m - 4.8l/m - - - Soares et al.131
biodiesel (univariate
analysis)
BÖCK ET AL.
TABLE 1 (Continued)

Detection target Analysis Analytical features Reference

BÖCK ET AL.

RSD
R2 LOD LOQ CV (%) (%) RMSEC RMSEP
−1
Determination of methanol in Calibration curve 0.964 90 mg kg - 3.7 - - - Soares and
biodiesel (univariate Rocha132
analysis)

Chromium speciation in Calibration curve 0.993 0.6 mg kg−1 2 mg kg−1 - 7.5 - - Costa et al.133
leather (univariate
analysis)

Determination of fluoride and Calibration curve 0.998n/0.999o 0.05n/0.10o mg L−1 0.10n/0.019o mg L−1 <1n/o - - - Pappis
phosphorus in water (univariate et al.134
analysis)

Quantification of biodiesel in Calibration curve 0.998 1.0% (v/v) - 0.9 - - - Soares et al.135
diesel blends (univariate
analysis)

Brazilian banknotes PCA - - - - - - - Vittorazzi

et al.136

Abbreviations: CV, coefficient of variation; LOD, limit of detection; LOQ, limit of quantification; PCA, principal component analysis; PLS, partial least squares; RMSEC, Root Mean Squared
Error Calibration; RMSEP, root mean squared error of prediction; RSD, Relative Standard Desviation.
a
Cocaine.
b
Procaine.
c
Levamisole.
d
Caffeine.
e
Total dissolved iron.
f
Iron speciation.
g
Hydrogen peroxide (low-range calibration).
h
Hydrogen peroxide (high-range calibration).
i
Albumin.
j
IgG.
k
Carbonic anhydrase.
l
600 μl of sample extract.
m
150 μl of sample extract.
n
Fluoride.
o
Phosphorus.
* 2
R cal.
7 of 19
8 of 19 BÖCK ET AL.

F I G U R E 6 Schematic diagram of the colorimetric spot test. (A) Reaction of biodiesel with iodine; (B) mixing and subsequent reaction
time; (C) transfer of 5 μl of the mixture to the paper; (D) addition of 5 μl of starch (1% m/v); and (E) acquisition of the RGB value by
employing the software PhotoMetrix®. Reproduced with permissions from Gonzalez and Woods118; Elsevier

F I G U R E 7 Illustration of the method of acquisition of the digital images by the smartphone. (A) Front image of the system, (B) side
image, and (C) higher image. Reproduced with permissions from Grasel et al.120; Springer

image detection agreed with those of the reference procedure (Friedman Titration), showing that there is not matrix
effect. It must be highlighted the very small volumes of sample and other solutions needed for the analysis.
The work developed by Nogueira et al.123 performed the monitoring of acid-base titration through univariate cali-
bration from digital images using the PhotoMetrix app. The authors used jaboticaba peel extract as an indicator and
performed the titration on wax printed paper microzones. First, buffer solutions from pH 2 to pH 12 were used to con-
struct the analytical curve, then three vinegar samples were evaluated for acetic acid quantification. Images were cap-
tured in ambient light with controlled capture distances. The results obtained with the digital imaging method were
compared with those from conventional titration method (reference method), and the difference between them was less
than 8%. By applying the Student t test, it was possible to prove that there were no significant differences between the
two methodologies for a 95% confidence level.
Böck et al.124 performed ethanol determination in cachaça using digital image analysis, obtained through a
smartphone and evaluated them using PhotoMetrix®, in multivariate mode through the PLS calibration model. The
authors used an apparatus with brightness controlled by two LED strips with six lamps each, resulting in an illumina-
tion of 9 W/m2. Figure 7 shows the illustration of this apparatus. The obtained results by the proposed methodology
(using the mobile phone) were compared with those from UV-VIS, which was used as the reference method. The
RMSEP obtained by the model was 0.0677%, considered satisfactory since the concentration range of ethanol in cachaça
is between 38% and 48%. Moreover, through the application of statistical tests, it was possible to verify that there was
no significant difference between the results from the proposed and reference methodologies.
An alternative methodology for interpreting the thermal stability of raw milk by colorimetric reaction with alizarol
was proposed by Helfer et al.125 The methodology (see Figure 8) was developed by building a PLS calibration model
BÖCK ET AL. 9 of 19

F I G U R E 8 Procedure used
for pH determination by mobile
device. Reproduced with
permissions from de Oliveira121;
Springer

with pH variation from 3 to 12; the solutions were packed in a black box with brightness controlled by a 6 W LED lamp,
where the images were captured to build a calibration model and sample prediction. The obtained results by the colori-
metric method using PhotoMetrix® were compared with the reference method (potentiometric). Raw milk samples
were obtained from seven local producers and evaluated. The results obtained by the colorimetric method using the
application showed no significant statistical differences in relation to the results obtained by the reference method.
Bento and Dantas126 proposed the use of PLS from digital images to determine maturation of pink mango samples
using the PhotoMetrix app to build the calibration model. Mango samples from different maturation stages were pur-
chased from the local market to build the model. Firstly, the fruit was cross-sectioned to extract part of the pulp to
quantify total dissolved solids, by reference method, for later use of these values in the calibration model. The images
were acquired from the outside of the fruit (peel), with standard capture distance and regularity of illumination. The
results obtained by the calibration model presented a good coefficient of determination at low RMSEC, showing the
potential of the application of this tool in the evaluation of fruit maturity, such as mango, because it is an easily accessi-
ble, low cost, and nondestructive methodology.
Yulia et al.127 developed a methodology for detecting wax in Gorengan samples. The test was conducted primarily
by evaluating the best conditions to perform the experiment; the optimization was based on the correlation coefficient
evaluated by the PhotoMetrix application in univariate mode, where the objective was to achieve the highest correlation
and best stability of the test color. After optimizing the reagent conditions, the authors used PCA to discriminate sam-
ples with different concentrations of added wax to cooking oil. The built-in PCA proved to be very efficient as it was
able to discriminate samples into three different groups, namely, low, medium, and high Gorengan wax addition index.
A methodology for quantification of the lactase enzyme present in pharmaceutical formulations using digital images
taken by a mobile phone camera and processed by the PhotoMetrix® was proposed by Dionizio et al.128 For lactase
quantification, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was used; images were captured
at a controlled distance, and cell flash was used. The app univariate mode was explored, where channel B was chosen
for quantification. Figure 9 shows the fragmented chromatogram and calibration curve for lactase standard. Six com-
mercial lactase samples were analyzed and quantified, and a recovery test was carried out to verify the accuracy of the

F I G U R E 9 Fragmented chromatograms of
different mass (0–0.38 mg) of standard lactase
(90 kDa) on gel electrophoresis along with
molecular weight marker proteins and
calibration curve for lactase. Reproduced with
permissions from Böck et al.124; BrJAC
10 of 19 BÖCK ET AL.

methodology. The results obtained by the recovery tests ranged from 78% to 104%, demonstrating the potential of the
proposed methodology for the quantification of lactase via SDS-PAGE through digital image analysis.
Lumbaque et al.129 proposed the determination of dissolved iron and hydrogen peroxide during solar photo-Fenton
process through digital image analysis. Iron determination was performed by two methodologies, one employing
1,10-phenantroline and the other with hydroquinone, and the determination of H2O2 was performed with ammonium
metavanadate. Both were performed using the PhotoMetrix app in multivariate mode (PLS), and the results were com-
pared with UV-VIS. The evaluated solutions were arranged in a 96-well plate, where 340 μl of the solutions were added.
The plate was placed in a box, and the images were acquired at constant distance and using the mobile flash as illumi-
nation. The results obtained by the PhotoMetrix app were very similar to those obtained by the reference methodology
(UV-vis), showing the ability of smartphones as alternatives to perform on-site analysis, with less waste generation, con-
sidering the low sample volume required for analysis.
PhotoMetrix® has also been used as an alternative in quantifying protein at microgram levels in human serum sam-
ples. Jesus et al.130 used SDS-PAGE for separation and quantification of three proteins. After separation, the gel was
photographed, and calibration curves were constructed for each protein. The polyacrylamide gel plate was placed inside
an acrylic box, where the images were captured. Recovery tests were performed to evaluate the trueness of the results.
The use of smartphones for protein quantification proved to be efficient, presenting good features such as precision and
accuracy, as well as detection and quantification limits, showing potential for a low-cost alternative for protein quantifi-
cation in different samples.
A methodology for the determination of methanol in biodiesel by digital images was proposed by Soares et al.131,
and the procedure was based on the colorimetric oxidation reaction of methanol to formaldehyde and subsequent reac-
tion with Schiff reagent. Methanol quantification was performed using the PhotoMetrix® in the univariate module, and
images were acquired with illumination and controlled distance, directly from the 1.5-ml polypropylene microtube,
where the reaction was performed. Recovery tests were performed, and the results found by both the recovery test and
the comparison with the reference methodology indicated that the methodology has potential for application in metha-
nol quantification in biodiesel.
Soares and Rocha132 also proposed a microdistillation methodology followed by methanol determination in biodie-
sel and quantification by digital image analysis. For this purpose, the authors set up a methanol determination system
based on the evaporation of the component from the biodiesel matrix. After evaporation, methanol was absorbed in the

F I G U R E 1 0 (A) Analytical
curve in the range of
0.05–0.75 mg L-1; (B) colors
related with standards used in
analytical curve. Reproduced
with permissions from
Lumbaque et al.129; UNESP
BÖCK ET AL. 11 of 19

F I G U R E 1 1 (A) Standard solutions used to create the calibration curve (left) and the calibration plot (right) for fluoride determination
with the PhotoMetrix® application [amount of shade × fluorine concentration (mg L−1)]; (B) standard solutions used to create the
calibration curve (left) and the calibration plot (right) for phosphorus determination using the PhotoMetrix® application [amount of
hue × phosphorus concentration (mg L−1)]. Reproduced with permissions from de Jesus JR et al.130; BrJAC

upper part of the system containing an acidified potassium permanganate solution, causing methanol oxidation by per-
manganate, thus being able to quantify the analyte through the discoloration of the permanganate. The authors used a
1.5-ml microtube, containing the permanganate solution, fixed in the upper part of Falcon® tube, containing the sam-
ple. The images were captured from the microtubes, at a fixed distance and lighting control; the images were processed
using the PhotoMetrix® in the univariate module. The results found by the proposed methodology based on micro-
distillation, followed by quantification by analysis of digital images, presented similar results to those obtained by refer-
ence methodologies, showing a low-cost alternative regarding equipment and reagent consumption for methanol
quantification in biodiesel.
PhotoMetrix® was also used by Costa et al.133 for chromium speciation in leather samples in an experiment con-
ducted during a chemistry class with undergraduate students. Seven leather samples, four bovine tanned with chro-
mium salt, and three ovine tanned with vegetal tannin were evaluated. The quantification reaction was performed with
12 of 19 BÖCK ET AL.

DPC chromogenic reagent, and the results obtained with the application were compared with UV-vis. For the acquisi-
tion of images, a wooden box was used in order to offer a homogeneous environment for all image acquisitions.
Figure 10 shows the calibration curve obtained at the PhotoMetrix® and the standard solutions used for it. The results
obtained by the app and the reference methodology showed no significant differences between them, demonstrating
the potential of applying the methodology in the quantification of chromium (IV) in leather samples. It also offers a
low-cost alternative to studying chemistry in leather classes.
Pappis et al.134 proposed a methodology for fluoride and phosphorus determination in natural and treated water
samples. The determinations were based on colorimetric reactions, evaluated through digital image analysis using the
PhotoMetrix app in the univariate module, and compared to reference methodology (spectrophotometer). The images
were acquired in a controlled lighting environment with a LED lamp (6 W); Figure 11 shows the calibration curve
obtained at the PhotoMetrix® and the standard solutions used for (A) for fluoride determination and (B) Phosphorus
determination. The results obtained by the proposed methodology showed an accuracy between 98% and 100.09% for
fluoride determination and between 100% and 100.4% for phosphorus determination, showing no differences from ref-
erence methodology, thus providing an alternative for in situ-based water analysis with immediate results.
A methodology for the determination of biodiesel in diesel blends by digital imaging was proposed by Soares
et al.135 The determination was based on the colorimetric reaction of violet complex formation between Fe (III) and
hydroxamate ion, generated by the reaction of characteristic alkyl esters of biodiesel with hydroxylamine and subse-
quent quantification by digital image analysis. For the analysis, it used a mobile phone for image acquisition and Photo-
Metrix® for processing, and a box with LED lamp was used in order to provide controlled environment. Recovery tests
were performed, and the obtained results were compared with the reference methodology, indicating no significant dif-
ferences among both methodologies. The analytical characteristics of the proposed method were compared with other
methodologies available in literature, which presented similar results. A great advantage over other methods is related
to low reagent consumption and residue generation (1 ml), which makes this methodology extremely attractive when
compared with others.
Vittorazzi et al.136 proposed a method for classifying Brazilian banknotes from analysis by digital images using a
smartphone. The objective of the work was to identify the authenticity of Brazilian banknotes of R$ 10, 20, 50, and
100 using the PhotoMetrix application. For the development of the work, the authors used authentic (n = 4) banknotes

F I G U R E 1 2 Possibilities of
image acquisition using
PhotoMetrix application
BÖCK ET AL. 13 of 19

obtained from the local bank and counterfeit (n = 12), made by the author himself, produced in three different com-
mercial printers, in addition to counterfeit banknotes provided by the Technical-Scientific Police of the State of Espírito
Santo. First, an experimental planning was carried out with the R$ 50.00 banknote to define parameters such as the best
region for image acquisition (holographic strip, tactile region in the lower left and upper right numbers, microprints on
the effigy, lower left number, animal figure, watermark, and puzzle), size of the region of interest (ROI) (16 × 16;
32 × 32; 64 × 64), focal length (8, 12, 15, and 20 cm), and brightness control (laboratory with controlled lighting envi-
ronment and in the field). After the optimization, the analysis of the banknotes of the other values R$ (100, 20, and 10)
was carried out considering the criteria that had previously presented the best results. Finally, the smartphone model
was varied to observe its influence on the results. For the analysis, the authors used PCA with channels R, G, B, H, S,
V, L, and I, where it was possible to visualize the classification between authentic and counterfeit/simulated samples,
showing a simple alternative, easy to perform, and low cost to exploit adulteration in banknotes, in addition to being a
nondestructive proposal; it is fast being a viable alternative for analyzing authenticity of banknotes in loco, because
with the results obtained in the field, it was possible to demonstrate that the brightness is not a determining factor the
data obtained, providing greater practicality and speed in obtaining the results.
Through the researches that used PhotoMetrix® early presented, it is possible to visualize the versatility of this appli-
cation, because it was used in different analysis, presenting significant results. It is possible to carry out direct
analysis,126,136 that is, without performing sample preparation, analysis of solid120 and liquid samples,131–133,135 with
the use of chambers to store the samples,121,124,125,129,134 or without lighting control accessories,127 in addition to all-
owing the sample to be conditioned in different ways to capture images such as paper122,123 and SDS-PAGE128,130 for
example.
Figure 12 shows four different ways of acquiring images related to the previously presented articles that used Photo-
Metrix app in their analysis, where in some works, chambers were made to capture images, and in others, the image
acquisition was performed without light control. Images were also captured from SDS–PAGE gel, in addition to samples
packed in paper microzones. These examples demonstrate the versatility of the analysis of digital images for several
kinds of matrices. Moreover, when compared with UV-vis, it presents several advantages: (i) low-cost; (ii) it is more
accessible and widespread; and (iii) as already discussed, and it allows direct analyzes of liquid and solid samples, with-
out any sample preparation.
Besides the listed advantages over UV-VIS, PhotoMetrix® also presents some advantage over other free applications
available for download: the possibility of carrying out analyzes such as Univariate and Multivariate Calibration, PCA,
and HCA on the mobile device itself, without the need to export the data for further analysis, in addition to offering
tools that are still available and which were not explored in this review, such as the use of external cameras, enabling
the acquisition of standard images, regardless of the camera quality of the smartphone, what can make the method
more robust.

3 | CONCLUSIONS AND FUTURE PERSPECTIVES

Analyses based on light spectrum have been explored for a long time, but the use of smartphones with detectors is very
recent, especially when compared with the use of spectrophotometer, for example. Smartphones are increasingly being
explored as analytical tools, and in this sense, this work showed different ways to explore this device, and it highlighted
the relevance of developing tools that enable not only data acquisition but also processing and generating the result on
the device itself. According to what was presented herein, it is clear that PhotoMetrix application fit very well to those
aspects.
Future objectives for analysis using smartphones are related to the inclusion of videos evaluation, thus enabling
kinetic studies. The implementation of machine learning allows the reduction of errors, thus leading external analysis
without brightness control.

A C K N O WL E D G M E N T S
The authors are grateful to Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPQ) and Instituto
Nacional de Ciência e Tecnologia (INCT)—Bioanalítica. This study was financed in part by the Coordenaç~ao de
Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES)—Finance Code 001.
14 of 19 BÖCK ET AL.

ORCID
Fernanda Carla Böck https://orcid.org/0000-0003-4791-2544
Adilson Ben da Costa https://orcid.org/0000-0003-3009-7459
~
Marco Flôres Ferrao https://orcid.org/0000-0002-3332-0540

R EF E RE N C E S
1. Valente J. EBC - Empresa Brasil de Comunicaç~ao. EBC. http://www.ebc.com.br/. Published 2019.
2. Roda A, Michelini E, Zangheri M, Di Fusco M, Calabria D, Simoni P. Smartphone-based biosensors: a critical review and perspectives.
TrAC - Trends Anal Chem. 2016;79:317-325. https://doi.org/10.1016/j.trac.2015.10.019
3. Li F, Bao Y, Wang D, Wang W, Niu L. Smartphones for sensing. Sci Bull. 2016;61(3):190-201. https://doi.org/10.1007/s11434-015-0954-1
4. Huang X, Xu D, Chen J, et al. Smartphone-based analytical biosensors. Analyst. 2018;143(22):5339-5351. https://doi.org/10.1039/
c8an01269e
5. Ding X, Mauk MG, Yin K, Kadimisetty K, Liu C. Interfacing pathogen detection with smartphones for point-of-care applications. Anal
Chem. 2019;91(1):655-672. https://doi.org/10.1021/acs.analchem.8b04973
6. Grudpan K, Kolev SD, Lapanantnopakhun S, McKelvie ID, Wongwilai W. Applications of everyday IT and communications devices in
modern analytical chemistry: a review. Talanta. 2015;136:84-94. https://doi.org/10.1016/j.talanta.2014.12.042
7. Rezazadeh M, Seidi S, Lid M, Pedersen-Bjergaard S, Yamini Y. The modern role of smartphones in analytical chemistry. TrAC - Trends
Anal Chem. 2019;118:548-555. https://doi.org/10.1016/j.trac.2019.06.019
8. McCracken KE, Yoon JY. Recent approaches for optical smartphone sensing in resource-limited settings: a brief review. Anal Methods.
2016;8(36):6591-6601. https://doi.org/10.1039/c6ay01575a
9. Yang K, Peretz-Soroka H, Liu Y, Lin F. Novel developments in mobile sensing based on the integration of microfluidic devices and
smartphones. Lab Chip. 2016;16(6):943-958. https://doi.org/10.1039/c5lc01524c
10. Rateni G, Dario P, Cavallo F. Smartphone-based food diagnostic technologies: a review. Sensors (Switzerland). 2017;17(6):1–22. https://
doi.org/10.3390/s17061453
11. Martinez AW, Phillips ST, Carrilho E, Thomas SW, Sindi H, Whitesides GM. Simple telemedicine for developing regions: camera
phones and paper-based microfluidic devices for real-time, off-site diagnosis. Anal Chem. 2008;80(10):3699-3707. https://doi.org/10.
1021/ac800112r
12. Lu Y, Shi W, Qin J, Lin B. Low cost, portable detection of gold nanoparticle-labeled microfluidic immunoassay with camera cell phone.
Electrophoresis. 2009;30(4):579-582. https://doi.org/10.1002/elps.200800586
13. Breslauer DN, Maamari RN, Switz NA, Lam WA, Fletcher DA. Mobile phone based clinical microscopy for global health applications.
PLoS One. 2009;4(7):1-7. https://doi.org/10.1371/journal.pone.0006320
14. Zhu H, Mavandadi S, Coskun AF, Yaglidere O, Ozcan A. Optofluidic fluorescent imaging cytometry on a cell phone. Anal Chem. 2011;
83(17):6641-6647. https://doi.org/10.1021/ac201587a
15. Zhu H, Yaglidere O, Su TW, Tseng D, Ozcan A. Cost-effective and compact wide-field fluorescent imaging on a cell-phone. Lab Chip.
2011;11(2):315-322. https://doi.org/10.1039/c0lc00358a
16. Delaney JL, Hogan CF, Tian J, Shen W. Electrogenerated chemiluminescence detection in paper-based microfluidic sensors. Anal
Chem. 2011;83(4):1300-1306. https://doi.org/10.1021/ac102392t
17. Canning J, Lau A, Naqshbandi M, Petermann I, Crossley MJ. Measurement of fluorescence in a rhodamine-123 doped self-assembled
“giant” mesostructured silica sphere using a smartphone as optical hardware. Sensors. 2011;11(7):7055-7062. https://doi.org/10.3390/
s110707055
18. Veigas B, Jacob JM, Costa MN, et al. Gold on paper-paper platform for Au-nanoprobe TB detection. Lab Chip. 2012;12(22):4802-4808.
https://doi.org/10.1039/c2lc40739f
19. Zhu H, Sikora U, Ozcan A. Quantum dot enabled detection of Escherichia coli using a cell-phone. Analyst. 2012;137(11):2541-2544.
https://doi.org/10.1039/c2an35071h
20. Shen L, Hagen JA, Papautsky I. Point-of-care colorimetric detection with a smartphone. Lab Chip. 2012;12(21):4240-4243. https://doi.
org/10.1039/c2lc40741h
21. Petryayeva E, Algar WR. Proteolytic assays on quantum-dot-modified paper substrates using simple optical readout platforms. Anal
Chem. 2013;85(18):8817-8825. https://doi.org/10.1021/ac4020066
22. Park TS, Li W, McCracken KE, Yoon J-Y. Smartphone quantifies Salmonella from paper microfluidics. Lab Chip. 2013;13(24):4832-
4840. https://doi.org/10.1039/c3lc50976a
23. Roda A, Michelini E, Cevenini L, Calabria D, Calabretta MM, Simoni P. Integrating biochemiluminescence detection on smartphones:
mobile chemistry platform for point-of-need analysis. Anal Chem. 2014;86(15):7299-7304. https://doi.org/10.1021/ac502137s
24. Long KD, Yu H, Cunningham BT. Smartphone instrument for portable enzyme-linked immunosorbent assays. Biomed Opt Express.
2014;5(11):1–15. https://doi.org/10.1364/boe.5.003792
25. Christodouleas DC, Nemiroski A, Kumar AA, Whitesides GM. Broadly available imaging devices enable high-quality low-cost photom-
etry. Anal Chem. 2015;87(18):9170-9178. https://doi.org/10.1021/acs.analchem.5b01612
BÖCK ET AL. 15 of 19

26. Oliveira KA, Damasceno D, De Oliveira CR, Da Silveira LA, De Oliveira AE, Coltro WKT. Dengue diagnosis on laser printed
microzones using smartphone-based detection and multivariate image analysis. Anal Methods. 2016;8(35):6506-6511. https://doi.org/10.
1039/c6ay01262k
27. Ravazzi CG, Franco MDOK, Vieira MCR, Suarez WT. Smartphone application for captopril determination in dosage forms and syn-
thetic urine employing digital imaging. Talanta. 2018;189:339-344. https://doi.org/10.1016/j.talanta.2018.07.015
28. Choodum A, Kanatharana P, Wongniramaikul W, Nic DN. Using the iPhone as a device for a rapid quantitative analysis of trinitrotolu-
ene in soil. Talanta. 2013;115:143-149. https://doi.org/10.1016/j.talanta.2013.04.037
29. Wang H, Li YJ, Wei JF, Xu JR, Wang YH, Zheng GX. Paper-based three-dimensional microfluidic device for monitoring of heavy
metals with a camera cell phone. Anal Bioanal Chem. 2014;406(12):2799-2807. https://doi.org/10.1007/s00216-014-7715-x
30. Wang Y, Li Y, Bao X, et al. A smartphone-based colorimetric reader coupled with a remote server for rapid on-site catechols analysis.
Talanta. 2016;160:194-204. https://doi.org/10.1016/j.talanta.2016.07.012
31. McCracken KE, Tat T, Paz V, Yoon JY. Smartphone-based fluorescence detection of bisphenol A from water samples. RSC Adv. 2017;7
(15):9237-9243. https://doi.org/10.1039/c6ra27726h
32. Lima MJA, Nascimento CF, Rocha FRP. Feasible photometric measurements in liquid-liquid extraction by exploiting smartphone-
based digital images. Anal Methods. 2017;9(14):2220-2225. https://doi.org/10.1039/c7ay00388a
33. Salcedo ARM, Sevilla FB. Colorimetric determination of mercury vapor using smartphone camera-based imaging. Instrum Sci Technol.
2018;46(4):450-462. https://doi.org/10.1080/10739149.2017.1395745
34. Wang H, Sun Y, Li H, Yue W, Kang Q, Shen D. A smartphone-based ratiometric resonance light scattering device for field analysis of
Pb2+ in river water samples and immunoassay of alpha fetoprotein using PbS nanoparticles as signal tag. Sens Actuators B. 2018;271
(May):358-366. https://doi.org/10.1016/j.snb.2018.05.103
35. Moerkens R, Brenard N, Bosmans L, et al. Protocol for semi-automatic identification of whiteflies Bemisia tabaci and Trialeurodes
vaporariorum on yellow sticky traps. J Appl Entomol. 2019;143(6):652-658. https://doi.org/10.1111/jen.12630
36. Carretero-Peña S, Blázquez LC, Pinilla-Gil E. Estimation of PM10 levels and sources in air quality networks by digital analysis of
smartphone camera images taken from samples deposited on filters. Sensors (Switzerland). 2019;19(21):1–19. https://doi.org/10.3390/
s19214791
37. Sankar K, Lenisha D, Janaki G, et al. Digital image-based quantification of chlorpyrifos in water samples using a lipase embedded
paper based device. Talanta. 2020;208:120408. https://doi.org/10.1016/j.talanta.2019.120408
38. de Oliveira LMA, dos Santos VB, da Silva EKN, Lopes AS, Dantas-Filho HA. An environment-friendly spot test method with digital
imaging for the micro-titration of citric fruits. Talanta. 2020;206(May 2019):120219. https://doi.org/10.1016/j.talanta.2019.120219
39. Zarghampour F, Yamini Y, Baharfar M, Javadian G, Faraji M. On-chip electromembrane extraction followed by sensitive digital image-
based colorimetry for determination of trace amounts of Cr (vi). Anal Methods. 2020;12(4):483-490. https://doi.org/10.1039/c9ay02328c
40. Thanakiatkrai P, Yaodam A, Kitpipit T. Age estimation of bloodstains using smartphones and digital image analysis. Forensic Sci Int.
2013;233(1–3):288-297. https://doi.org/10.1016/j.forsciint.2013.09.027
41. Valderrama L, Valderrama P. Nondestructive identification of blue pen inks for documentoscopy purpose using iPhone and digital
image analysis including an approach for interval confidence estimation in PLS-DA models validation. Chemom Intel Lab Syst. 2016;
156:188-195. https://doi.org/10.1016/j.chemolab.2016.06.009
42. da Silva GO, de Araujo WR, Paix~ao TRLC. Portable and low-cost colorimetric office paper-based device for phenacetin detection in sei-
zed cocaine samples. Talanta. 2018;176(August 2017):674-678. https://doi.org/10.1016/j.talanta.2017.08.082
43. Thongprajukaew K, Choodum A, Sa-E B, Hayee U. Smart phone: a popular device supports amylase activity assay in fisheries research.
Food Chem. 2014;163:87-91. https://doi.org/10.1016/j.foodchem.2014.04.080
44. San Park T, Baynes C, Cho SI, Yoon JY. Paper microfluidics for red wine tasting. RSC Adv. 2014;4(46):24356-24362. https://doi.org/10.
1039/c4ra01471e
45. Maravillas MJ, Mendenilla AM, Merino AF, Quinto E. Development of a method through digital image analysis for analyzing poly-
aniline based sensors for total volatile basic nitrogen (TVBN) as fish freshness indicators. J Chem Eng. 2018;18(2):7-12.
46. Moonrungsee N, Peamaroon N, Boonmee A, Suwancharoen S, Jakmunee J. Evaluation of tyrosinase inhibitory activity in Salak
(Salacca zalacca) extracts using the digital image-based colorimetric method. Chem Pap. 2018;72(11):2729-2736. https://doi.org/10.
1007/s11696-018-0528-1
47. Acevedo MSMSF, Lima MJA, Nascimento CF, Rocha FRP. A green and cost-effective procedure for determination of anionic surfac-
tants in milk with liquid-liquid microextraction and smartphone-based photometric detection. Microchem J. 2018;143(June):259-263.
https://doi.org/10.1016/j.microc.2018.08.002
48. Lee K, Baek S, Kim D, Seo J. A freshness indicator for monitoring chicken-breast spoilage using a Tyvek® sheet and RGB color analysis.
Food Packag Shelf Life. 2019;19:40-46. https://doi.org/10.1016/j.fpsl.2018.11.016
49. Granica M, Tymecki Ł. Analytical aspects of smart (phone) fluorometric measurements. Talanta. 2019;197(October 2018):319-325.
https://doi.org/10.1016/j.talanta.2019.01.032
50. León-Roque N, Aguilar-Tuesta S, Quispe-Neyra J, Mamani-Navarro W, Alfaro-Cruz S, Condezo-Hoyos L. A green analytical assay for
the quantitation of the total saponins in quinoa (Chenopodium quinoa Willd.) based on macro lens-coupled smartphone. Talanta.
2019;204(March):576-585. https://doi.org/10.1016/j.talanta.2019.06.014
16 of 19 BÖCK ET AL.

51. dos Santos VB, da Silva EKN, de Oliveira LMA, Suarez WT. Low cost in situ digital image method, based on spot testing and
smartphone images, for determination of ascorbic acid in Brazilian Amazon native and exotic fruits. Food Chem. 2019;285(February):
340-346. https://doi.org/10.1016/j.foodchem.2019.01.167
52. Peng B, Zhou J, Xu J, et al. A smartphone-based colorimetry after dispersive liquid–liquid microextraction for rapid quantification of
calcium in water and food samples. Microchem J. 2019;149(July):104072. https://doi.org/10.1016/j.microc.2019.104072
53. Kong L, Gan Y, Liang T, et al. A novel smartphone-based CD-spectrometer for high sensitive and cost-effective colorimetric detection
of ascorbic acid. Anal Chim Acta. 2020;1093:150-159. https://doi.org/10.1016/j.aca.2019.09.071
54. Lamarca RS, de Lima Gomes PCF. A low cost method for carbamazepine, ciprofloxacin and norfloxacin determination in pharmaceuti-
cal formulations based on spot-test and smartphone images. Microchem J. 2020;152(October 2019):104297. https://doi.org/10.1016/j.
microc.2019.104297
55. Yu L, Shi ZZ, Fang C, Zhang YY, Liu YS, Li CM. Disposable lateral flow-through strip for smartphone-camera to quantitatively detect
alkaline phosphatase activity in milk. Biosens Bioelectron. 2015;69:307-315. https://doi.org/10.1016/j.bios.2015.02.035
56. de Lima CM, Fernandes DDS, Pereira GE, de Gomes A, de Araújo MCU, Diniz PHGD. Digital image-based tracing of geographic ori-
gin, winemaker, and grape type for red wine authentication. Food Chem. 2020;312(December 2019):126060. https://doi.org/10.1016/j.
foodchem.2019.126060
57. Phonchai A, Rattana S, Thongprajukaew K. A portable sol-gel urea colorimetric method for the determination of urea in feedstuffs.
Food Chem. 2020;319(March 2019):126545. https://doi.org/10.1016/j.foodchem.2020.126545
58. Monošík R, Bezerra dos Santos V, Angnes L. A simple paper-strip colorimetric method utilizing dehydrogenase enzymes for analysis of
food components. Anal Methods. 2015;7(19):8177-8184. https://doi.org/10.1039/c5ay01556a
59. Bueno L, Meloni GN, Reddy SM, Paix~ao TRLC. Use of plastic-based analytical device, smartphone and chemometric tools to discrimi-
nate amines. RSC Adv. 2015;5(26):20148-20154. https://doi.org/10.1039/c5ra01822f
60. Silva Neto GF, Fonseca A, Braga JWB. Classification of mineral waters based on digital images acquired by smartphones. Quim Nova.
2016;39(7):876-881. https://doi.org/10.5935/0100-4042.20160088
61. Botelho BG, Dantas KCF, Sena MM. Determination of allura red dye in hard candies by using digital images obtained with a mobile
phone and N-PLS. Chemom Intel Lab Syst. 2017;167(April):44-49. https://doi.org/10.1016/j.chemolab.2017.05.004
62. Pérez-Bernal JL, Villar-Navarro M, Morales ML, Ubeda C, Callejón RM. The smartphone as an economical and reliable tool for moni-
toring the browning process in sparkling wine. Comput Electron Agric. 2017;141:248-254. https://doi.org/10.1016/j.compag.2017.07.029
63. de Oliveira Krambeck Franco M, Suarez WT, dos Santos VB. Digital image method smartphone-based for furfural determination in
sugarcane spirits. Food Anal Methods. 2017;10(2):508-515. https://doi.org/10.1007/s12161-016-0605-4
64. de Oliveira Krambeck Franco M, Suarez WT, Maia MV, dos Santos VB. Smartphone application for methanol determination in sugar
cane spirits employing digital image-based method. Food Anal Methods. 2017;10(6):2102-2109. https://doi.org/10.1007/s12161-016-
0777-y
65. Coutinho MS, Morais CLM, Neves ACO, Menezes FG, Lima KMG. Colorimetric determination of ascorbic acid based on its interfering
effect in the enzymatic analysis of glucose: an approach using smartphone image analysis. J Braz Chem Soc. 2017;28(12):2500-2505.
https://doi.org/10.21577/0103-5053.20170086
66. Lamarca RS, Luchiari NDC, Bonjorno AF, Passaretti Filho J, Cardoso AA, De Lima Gomes PCF. Determination of formaldehyde in
cosmetic products using gas-diffusion microextraction coupled with a smartphone reader. Anal Methods. 2019;11(29):3697-3705.
https://doi.org/10.1039/c9ay00720b
67. Ribeiro FCP, Oliveira AS, Araújo A, et al. Detection oxidative degradation in lubricating oil under storage conditions using digital
images and chemometrics. Microchem J. 2019;147(October 2018):622-627. https://doi.org/10.1016/j.microc.2019.03.087
68. Jo~ao AF, Squissato AL, Fernandes GM, Cardoso RM, Batista AD, Muñoz RAA. Iron (III) determination in bioethanol fuel using a
smartphone-based device. Microchem J. 2019;146:1134-1139. https://doi.org/10.1016/j.microc.2019.02.053
69. Safarik I, Baldikova E, Prochazkova J, Pospiskova K. Smartphone-based image analysis for evaluation of magnetic textile solid phase
extraction of colored compounds. Heliyon. 2019;5(12):e02995. https://doi.org/10.1016/j.heliyon.2019.e02995
70. Salles MO, Meloni GN, De Araujo WR, Paix~ao TRLC. Explosive colorimetric discrimination using a smartphone, paper device and
chemometrical approach. Anal Methods. 2014;6(7):2047-2052. https://doi.org/10.1039/c3ay41727a
71. Koesdjojo MT, Pengpumkiat S, Wu Y, et al. Cost effective paper-based colorimetric microfluidic devices and mobile phone camera
readers for the classroom. J Chem Educ. 2015;92(4):737-741. https://doi.org/10.1021/ed500401d
72. De Morais CDLM, Silva SRB, Vieira DS, Lima KMG. Integrating a smartphone and molecular modeling for determining the binding
constant and stoichiometry ratio of the Iron (II)-Phenanthroline complex: an activity for analytical and physical chemistry laboratories.
J Chem Educ. 2016;93(10):1760-1765. https://doi.org/10.1021/acs.jchemed.6b00112
73. Gee CT, Kehoe E, Pomerantz WCK, Penn RL. Quantifying protein concentrations using smartphone colorimetry: a new method for an
established test. J Chem Educ. 2017;94(7):941-945. https://doi.org/10.1021/acs.jchemed.6b00676
74. Abaoag CJ, Alforque R, Ordoño JB, Quinto E. Non-invasive detection of human body liquor intake based on optical biosensor. In:
MATEC Web of Conferences. Vol 156. ; 2018. https://doi.org/10.1051/matecconf/201815605005
75. Kumar A, Sahoo PR, Arora P, Kumar S. A light controlled, sensitive, selective and portable spiropyran based receptor for mercury ions
in aqueous solution. J Photochem Photobiol a Chem. 2019;384(July):112061. https://doi.org/10.1016/j.jphotochem.2019.112061
76. Resque IS, dos Santos VB, Suarez WT. An environmentally friendly analytical approach based on spot test and digital image to evaluate
the conformity of bleaching products. Chem Pap. 2019;73(7):1659-1668. https://doi.org/10.1007/s11696-019-00717-w
BÖCK ET AL. 17 of 19

77. Bogucki R, Greggila M, Mallory P, et al. A 3D-printable dual beam spectrophotometer with multiplatform smartphone adaptor. J Chem
Educ. 2019;96(7):1527-1531. https://doi.org/10.1021/acs.jchemed.8b00870
78. Austerjost J, Marquard D, Raddatz L, et al. A smart device application for the automated determination of E. coli colonies on agar
plates. Eng Life Sci. 2017;17(8):959-966. https://doi.org/10.1002/elsc.201700056
79. Gallegos D, Long KD, Yu H, et al. Label-free biodetection using a smartphone. Lab Chip. 2013;13(11):2124-2132. https://doi.org/10.
1039/c3lc40991k
80. Delaney JL, Doeven EH, Harsant AJ, Hogan CF. Use of a mobile phone for potentiostatic control with low cost paper-based micro-
fluidic sensors. Anal Chim Acta. 2013;790:56-60. https://doi.org/10.1016/j.aca.2013.06.005
81. Lee S, Kim G, Moon J. Performance improvement of the one-dot lateral flow immunoassay for aflatoxin B1 by using a smartphone-
based reading system. Sensors (Switzerland). 2013;13(4):5109-5116. https://doi.org/10.3390/s130405109
82. Junrat J, Saikaew KR, Eua-Anant N, Pattaramanon R. Implementation and evaluation of SDS-PAGE image analysis on a mobile app.
2014 Int Comput Sci Eng Conf ICSEC 2014. 2014:346–349. doi:https://doi.org/10.1109/ICSEC.2014.6978220
83. Koh A, Kang D, Xue Y, et al. A soft, wearable microfluidic device for the capture, storage, and colorimetric sensing of sweat. Sci Transl
Med. 2016;8(366):1-13. https://doi.org/10.1126/scitranslmed.aaf2593
84. Cevenini L, Calabretta MM, Lopreside A, et al. Exploiting NanoLuc luciferase for smartphone-based bioluminescence cell biosensor for
(anti)-inflammatory activity and toxicity. Anal Bioanal Chem. 2016;408(30):8859-8868. https://doi.org/10.1007/s00216-016-0062-3
85. Aydın HB, Cheema JA, Ammanath G, et al. Pixelated colorimetric nucleic acid assay. Talanta. 2020;209(June 2019):120581. https://doi.
org/10.1016/j.talanta.2019.120581
86. Gómez-Robledo L, López-Ruiz N, Melgosa M, Palma AJ, Capitán-Vallvey LF, Sánchez-Marañón M. Using the mobile phone as munsell
soil-colour sensor: an experiment under controlled illumination conditions. Comput Electron Agric. 2013;99:200-208. https://doi.org/10.
1016/j.compag.2013.10.002
87. Moonrungsee N, Pencharee S, Jakmunee J. Colorimetric analyzer based on mobile phone camera for determination of available phos-
phorus in soil. Talanta. 2015;136:204-209. https://doi.org/10.1016/j.talanta.2015.01.024
88. Sun H, Jia Y, Dong H, Fan L, Zheng J. Multiplex quantification of metals in airborne particulate matter via smartphone and paper-
based microfluidics. Anal Chim Acta. 2018;1044:110-118. https://doi.org/10.1016/j.aca.2018.07.053
89. Klllç V, Alankus G, Horzum N, Mutlu AY, Bayram A, Solmaz ME. Single-image-referenced colorimetric water quality detection using
a smartphone. ACS Omega. 2018;3(5):5531-5536. https://doi.org/10.1021/acsomega.8b00625
90. Firdaus ML, Aprian A, Meileza N, et al. Smartphone coupled with a paper-based colorimetric device for sensitive and portable mercury
ion sensing. Chem. 2019;7(25):1-9. https://doi.org/10.3390/chemosensors7020025
91. Ludwig SKJ, Zhu H, Phillips S, et al. Cellphone-based detection platform for rbST biomarker analysis in milk extracts using a micro-
sphere fluorescence immunoassay. Anal Bioanal Chem. 2014;406(27):6857-6866. https://doi.org/10.1007/s00216-014-7984-4
92. Masawat P, Harfield A, Namwong A. An iPhone-based digital image colorimeter for detecting tetracycline in milk. Food Chem. 2015;
184:23-29. https://doi.org/10.1016/j.foodchem.2015.03.089
93. Fang J, Qiu X, Wan Z, et al. A sensing smartphone and its portable accessory for on-site rapid biochemical detection of marine toxins.
Anal Methods. 2016;8(38):6895-6902. https://doi.org/10.1039/c6ay01384h
94. Cheng Y-S, Wang S-C, Liu Y-H, Peng B-R. An intelligent noninvasive taste detection app for watermelons. In: Proceedings - 2017 5th
international conference on applied computing and information technology, 2017 4th international conference on computational
science/intelligence and applied informatics and 2017 1st international conference on big data, Cloud Compu ; 2017:90–94. https://doi.
org/10.1109/ACIT-CSII-BCD.2017.74
95. Liu Z, Zhang Y, Xu S, et al. A 3D printed smartphone optosensing platform for point-of-need food safety inspection. Anal Chim Acta.
2017;966:81-89. https://doi.org/10.1016/j.aca.2017.02.022
96. Lin B, Yu Y, Cao Y, et al. Point-of-care testing for streptomycin based on aptamer recognizing and digital image colorimetry by
smartphone. Biosens Bioelectron. 2018;100(378):482-489. https://doi.org/10.1016/j.bios.2017.09.028

97. Capoferri D, Alvarez-Diduk R, Del Carlo M, Compagnone D, Merkoçi A. Electrochromic molecular imprinting sensor for visual and
smartphone-based detections. Anal Chem. 2018;90(9):5850-5856. https://doi.org/10.1021/acs.analchem.8b00389
98. Shahvar A, Saraji M, Gordan H, Shamsaei D. Combination of paper-based thin film microextraction with smartphone-based sensing
for sulfite assay in food samples. Talanta. 2019;197(January):578-583. https://doi.org/10.1016/j.talanta.2019.01.071
99. Marinho OR, Lima MJA, Rocha FRP, Reis BF, Kamogawa MY. A greener, fast, and cost-effective smartphone-based digital image pro-
cedure for quantification of ethanol in distilled beverages. Microchem J. 2019;147:437-443. https://doi.org/10.1016/j.microc.2019.03.054
100. Mudanyali O, Dimitrov S, Sikora U, Padmanabhan S, Navruz I, Ozcan A. Integrated rapid-diagnostic-test reader platform on a
cellphone. Lab Chip. 2012;12(15):2678-2686. https://doi.org/10.1039/c2lc40235a
101. Coskun AF, Wong J, Khodadadi D, Nagi R, Tey A, Ozcan A. A personalized food allergen testing platform on a cellphone. Lab Chip.
2013;13(4):636-640. https://doi.org/10.1039/c2lc41152k
102. You DJ, Park TS, Yoon JY. Cell-phone-based measurement of TSH using Mie scatter optimized lateral flow assays. Biosens Bioelectron.
2013;40(1):180-185. https://doi.org/10.1016/j.bios.2012.07.014
103. Oncescu V, O'Dell D, Erickson D. Smartphone based health accessory for colorimetric detection of biomarkers in sweat and saliva. Lab
Chip. 2013;13(16):3232-3238. https://doi.org/10.1039/c3lc50431j
104. Lopez-Ruiz N, Curto VF, Erenas MM, et al. Smartphone-based simultaneous pH and nitrite colorimetric determination for paper
microfluidic devices. Anal Chem. 2014;86(19):9554-9562. https://doi.org/10.1021/ac5019205
18 of 19 BÖCK ET AL.

105. Liang PS, Park TS, Yoon JY. Rapid and reagentless detection of microbial contamination within meat utilizing a smartphone-based bio-
sensor. Sci Rep. 2014;4(1):4-11. https://doi.org/10.1038/srep05953
106. Wei Q, Nagi R, Sadeghi K, et al. Detection and spatial mapping of mercury contamination in water samples using a smart-phone. ACS
Nano. 2014;8(2):1121-1129. https://doi.org/10.1021/nn406571t
107. Lee S, Oncescu V, Mancuso M, Mehta S, Erickson D. A smartphone platform for the quantification of vitamin D levels. Lab Chip. 2014;
14(8):1437-1442. https://doi.org/10.1039/c3lc51375k
108. Oncescu V, Mancuso M, Erickson D. Cholesterol testing on a smartphone. Lab Chip. 2014;14(4):759-763. https://doi.org/10.1039/
c3lc51194d
109. Chathura Priyankara HA, Withanage DK. Computer assisted plant identification system for Android. MERCon 2015 - Moratuwa Eng
Res Conf. 2015:148–153. doi:https://doi.org/10.1109/MERCon.2015.7112336
110. Park TS, Yoon J-Y. Smartphone detection of Escherichia coli from field water samples on paper microfluidics. IEEE Sens J. 2015;15(3):
1902-1907. https://doi.org/10.1109/JSEN.2014.2367039
111. Levin S, Krishnan S, Rajkumar S, Halery N, Balkunde P. Monitoring of fluoride in water samples using a smartphone. Sci Total Envi-
ron. 2016;551–552:101-107. https://doi.org/10.1016/j.scitotenv.2016.01.156
112. Tonmoy TH, Hanif MA, Rahman HA, Khandaker N, Hossain I. Error reduction in arsenic detection through color spectrum analysis.
19th Int Conf Comput Inf Technol ICCIT 2016. 2016:343–350. doi:https://doi.org/10.1109/ICCITECHN.2016.7860221
113. Adi K, Pujiyanto S, Dwi Nurhayati O, Pamungkas A. Beef quality identification using thresholding method and decision tree classifica-
tion based on android smartphone. J Food Qual. 2017;2017:1-10. https://doi.org/10.1155/2017/1674718
114. Nurhayati OD. Omega-3 chicken egg detection system using a mobile-based image processing segmentation method. Eighth Int Conf
Graph Image Process (ICGIP 2016). 2017;10225(February 2017):1022516. https://doi.org/10.1117/12.2266088
115. Snow JW, Ceylan Koydemir H, Karinca DK, Liang K, Tseng D, Ozcan A. Rapid imaging, detection, and quantification of Nosema
ceranae spores in honey bees using mobile phone-based fluorescence microscopy. Lab Chip. 2019;19(5):789-797. https://doi.org/10.
1039/c8lc01342j
116. Helfer GA, Magnus VS, Böck FC, Teichmann A, Ferr~ao MF, Da Costa AB. PhotoMetrix: an application for univariate calibration and
principal components analysis using colorimetry on mobile devices. J Braz Chem Soc. 2017;28(2). doi:https://doi.org/10.5935/0103-
5053.20160182
117. Costa AB da, Helfer GA. PHOTOMETRIX PRO Certificado de Registro de Programas de Computador Processo no: BR 51 2018
000567-5. 2018:1.
118. Gonzalez RC, Woods RE. Digital Image Processing. 3rd ed. New Jersey: Pearson Prentice Hall; 2008.
119. Russ JC. The Image Processing Handbook. 6th ed. Boca Raton: CRC Press; 2011.
120. Grasel FS, Ferr~ao MF, Helfer GA, Costa AB. Principal component analysis of commercial tannin extracts using digital images on
mobile devices. J Braz Chem Soc. 2016;27(12):2372-2377. https://doi.org/10.5935/0103-5053.20160135
121. de Oliveira PAC. Medidas colorimétricas com smartphones para identificaç~ao de amostras de cocaína e quantificaç~ao de alguns
adulterantes. Programa Pós Grad Em Química - UnB. 2017;01. http://repositorio.unb.br/handle/10482/25272
122. Soares S, Lima MJA, Rocha FRP. A spot test for iodine value determination in biodiesel based on digital images exploiting a
smartphone. Microchem J. 2017;133:195-199. https://doi.org/10.1016/j.microc.2017.03.029
123. Nogueira SA, Sousa LR, Silva NKL, Rodrigues PHF, Coltro WKT. Monitoring acid–base titrations on wax printed paper microzones
using a smartphone. Micromachines. 2017;139(5):1-10. https://doi.org/10.3390/mi8050139
124. Böck FC, Helfer GA, da Costa AB, Dessuy MB, Ferr~ao MF. Rapid determination of ethanol in sugarcane spirit using partial least
squares regression embedded in smartphone. Food Anal Methods. 2018;11(7):1951–1957. https://doi.org/10.1007/s12161-018-1167-4
125. Helfer GA, Tischer B, Filoda PF, et al. A new tool for interpretation of thermal stability of raw milk by means of the alizarol test using
a PLS model on a mobile device. Food Anal Methods. 2018;11(7):2022-2028. https://doi.org/10.1007/s12161-018-1190-5
126. Bento IX, de Sousa Dantas, A.N. Utilizaç~ao Do Smartphone Para Análise Rápida Do Brix De Mangas Utilizando O App Photometrix.
In: III CONAPESC. ; 2017.
127. Yulia WH, Fatoni A. Qualitative analysis method of detection of wax content in gorengan using smartphone. J Pharm Sci Community.
2018;15(1):7-15. https://doi.org/10.24071/jpsc.1511052
128. Dionizio BS, Babos DV, de Souza DHFs, Pereira-Filho ER. Evaluation of the quality of formulations containing lactase (β-galactosidase)
employing gel electrophoresis and cell phone. Brazilian J Anal Chem. 2019;6(24):1–9. https://doi.org/10.30744/brjac.2179-3425.ar-13-
2019
129. Lumbaque EC, da Silva BA, Böck FC, Helfer GA, Ferr~ao MF, Sirtori C. Total dissolved iron and hydrogen peroxide determination using
the PhotoMetrixPRO application: a portable colorimetric analysis tool for controlling important conditions in the solar photo-Fenton
process J Hazard Mater Junho 2019. https://doi.org/10.1016/j.jhazmat.2019.06.017378, 120740.
130. de Jesus JR, Guimar~aes IC, Arruda MAZ. Quantifying proteins at microgram levels integrating gel electrophoresis and smartphone
technology. J Proteomics. 2019;198(September 2018):45-49. https://doi.org/10.1016/j.jprot.2018.12.002
131. Soares S, Torres KG, Pimentel EL, Martelli PB, Rocha FRP. A novel spot test based on digital images for determination of methanol in
biodiesel. Talanta. 2019;195:229-235. https://doi.org/10.1016/j.talanta.2018.11.028
132. Soares S, Rocha FRP. A simple and low-cost approach for microdistillation: application to methanol determination in biodiesel exploi-
ting smartphone-based digital images. Talanta. 2019;199:285-289. https://doi.org/10.1016/j.talanta.2019.02.064
BÖCK ET AL. 19 of 19

133. Costa V, Neiva A, Pereira-Filho E. Chromium speciation in leather samples: an experiment using digital images, mobile phones and
environmental concepts. Eclética Química J. 2019;44(1):62-74. https://doi.org/10.26850/1678-4618eqj.v44.1
134. Pappis C, Librelotto M, Baumann L, et al. Point-of-use determination of fluoride and phosphorus in water through a smartphone using
the PhotoMetrix ® App. Brazilian J Anal Chem. 2019;6(25):1-9. https://doi.org/10.30744/brjac.2179-3425.TN-25-2019
135. Soares S, Nunes LC, Melchert WR, Rocha FRP. Spot test exploiting smartphone-based digital images for determination of biodiesel in
diesel blends. Microchem J. 2020;152(September 2019):104273. https://doi.org/10.1016/j.microc.2019.104273
136. Vittorazzi B V., Costa RA, Coelho LM, et al. Classifying brazilian banknotes (R$) using smartphone image analysis. Quim Nova. 2020;X
(00):1–8. https://doi.org/10.21577/0100-4042.20170508
137. Mohan PJ, Gupta SD. Intelligent image analysis for retrieval of leaf chlorophyll content of rice from digital images of smartphone under
natural light. Photosynthetica. 2019;57(2):388-398. https://doi.org/10.32615/ps.2019.046
138. Cerrato-alvarez M, Frutos-puerto S, Miro-Rodríguez C, Pinilla-Gil E. Measurement of tropospheric ozone by digital image analysis of
indigotrisulfonate-impregnated passive sampling pads using a smartphone camera. Microchem J. 2020;154:104535. https://doi.org/10.
1016/j.microc.2019.104535
139. Li C, Adhikari R, Yao Y, et al. Measuring plant growth characteristics using smartphone based image analysis technique in controlled
environment agriculture. Comput Electron Agric. 2020;168(October 2019):105123. https://doi.org/10.1016/j.compag.2019.105123

How to cite this article: Böck FC, Helfer GA, AB da Costa, Dessuy MB, Ferr~ao MF. PhotoMetrix and
colorimetric image analysis using smartphones. Journal of Chemometrics. 2020;e3251. https://doi.org/10.1002/
cem.3251