Talan a 276 (2024) 126254
A ailable online 12 May 2024
0039-9140/© 2024 The Au ho s. Published by Else ie B.V. This is an open access a icle unde he CC BY-NC-ND license (h p://c ea i ecommons.o g/licenses/by-
nc-nd/4.0/).
Mic oscale i a ion o ace ic acid using digi al colo ime y and pape -based
analy ical de ices
I a i Be asa e
a
,
b
,
*
, Ane Bo daga ay
a
, Rosa Ga cia-A ona
a
, Mi en Os a
a
,
William Reis de A aujo
b
, Maide Vidal
a
a
Depa men o Applied Chemis y, Uni e si y o he Basque Coun y (UPV/EHU), 20018, Donos ia/San Sebas ian, Spain
b
Po able Chemical Senso s Lab, Depa men o Analy ical Chemis y, Ins i u e o Chemis y, S a e Uni e si y o Campinas (UNICAMP), 13083-970, Campinas, S˜
ao
Paulo, B azil
ARTICLE INFO
Keywo ds:
Pape -based de ice
Mic o- i a ion
Pape mic o luidics
Colo ime y
Digi al image analysis
P ocess au oma ion
ABSTRACT
A quan i a i e me hod o acid-base i a ions in pape -based de ices (PADs) is desc ibed o analyze ace ic acid in
inega samples. In his wo k, wo di e en ypes o PADs we e de eloped: a de ice o indi idual spo es ing
and a mic o luidic de ice. Digi al colo ime y was used as he de ec ion me hod, and he images we e acqui ed
using a sma phone and a homemade box wi h LED ligh s o con olled image acquisi ion. Ti a ion cu es we e
buil wi h jus eigh poin s, using he R channel based on he g adual colo ansi ion om ed o blue o li mus, a
na u al indica o . The endpoin was accu a ely de e mined by second de i a i e calcula ions. Bo h sys ems we e
applied o i een inega samples o di e en ypes, and good concen a ion esul s we e ob ained in compa ison
o he e e ence me hod. The p oposed me hodology is simple, as , en i onmen ally iendly, and su passes he
need o calib a ion cu e cons uc ion. Mo eo e , he subjec i e endpoin iden i ica ion is elimina ed, and he
me hod was au oma ed o p o ide a high h oughpu wo k low, sui able o quali y con ol p ocesses and eal-
ime measu emen s.
1. In oduc ion
One o he mos popula adi ional me hodologies ha enables
quan i a i e analysis is olume ic analysis. As a esul , he concen a-
ion o a pa icula analy e can be measu ed using a solu ion wi h a
known concen a ion (usually used as i an ). The e a e many di e en
i a ion ypes, and hey can be ca ego ized based on he measu ing
echnique, he o de o he i an s, o he ype o esponse. The esponse
can be based on an acid-base, edox, complexome ic, o p ecipi a ion
eac ion [1]. Focusing on acid-base i a ions, he mos common
me hods employed a e manual i a ion wi h isual indica ion o he
endpoin (he ea e EP, se o co espond o equi alence poin ).
Ins umen al and au oma ed lab-based me hods using spec opho o-
me ic de ec o s and he use o pH elec odes (po en iome y) p o ide
accu a e measu es, exemp ing he analys om in e p e ing he colo
change. Howe e , o e he las decades, signi ican echnological ad-
ances ha e been made, enabling he implemen a ion o digi al colo i-
me ic analysis o ollow hese i a ions and o de ec he endpoin a
he poin -o -need [2,3].
Digi al colo ime y cons i u es a powe ul ool due o i s simplici y,
low cos , and he di e si y o imaging op ions, anging om desk op
scanne s and webcams o digi al came as and sma phones [4]. An
image can be decomposed in o di e en colo models, such as RGB, HSV
o CIELAB, and hen, colo in ensi y can be ela ed o analy e concen-
a ion [5]. Many examples can be ound whe e digi al colo ime y has
been applied. Fo ins ance, o he sc eening o disease ma ke s in se um
[6], he de e mina ion o sul onamides in wa e [7], nickel in elec oless
coa ing ba hs [8], and o al phenolic con en in ea and in usion samples
[9]. Rega ding he i a ion-based de e mina ion o o al acidi y, i has
been calcula ed using a mic opla e, ei he in wines wi h a la bed
scanne [10], o in ci ic ui s employing a sma phone [11], among
o he s. In bo h cases, a ec o is c ea ed wi h RGB alues ex ac ed om
he image, which is plo ed agains he added i an olume o ob ain a
i a ion cu e. Then, EP is calcula ed wi h i s o second de i a i es. In
ac , de i a i es a e he employed ma hema ical calcula ions o de e -
mine he in lec ion poin o sinusoidal cu es as he ones ob ained in
i a ions. Sodium chlo ide has also been de e mined by i a ion and
employing a mic opla e – sma phone sys em [12]. In his case, G
* Co esponding au ho . Depa men o Applied Chemis y, Uni e si y o he Basque Coun y (UPV/EHU), 20018, Donos ia/San Sebas ian, Spain.
E-mail add ess: [email p o ec ed] (I. Be asa e).
Con en s lis s a ailable a ScienceDi ec
Talan a
jou nal homepage: www.else ie .com/loca e/ alan a
h ps://doi.o g/10.1016/j. alan a.2024.126254
Recei ed 21 Feb ua y 2024; Recei ed in e ised o m 6 May 2024; Accep ed 11 May 2024
Talan a 276 (2024) 126254
2
channel alues we e plo ed agains added i an olume and EP was
calcula ed as he in e sec ion be ween wo s aigh lines.
Pape -based analy ical de ices (PADs) ha e also d awn a en ion as a
p omising ool in analy ical chemis y in o de o de ec and quan i y he
analy e o in e es . Pape is made o cellulose, which gi es he abili y o
anspo liquids wi hou ins umen a ion, by capilla i y [13]. The e a e
many ways o manu ac u e hese de ices, including pho oli hog aphy,
plasma ea men , wax-p in ing, ink-p in ing and lase ea men [14].
The de ec ion can also be done by se e al me hods, such as luo escence,
Raman sca e ing, elec ochemis y, o colo ime y. Because o i s lex-
ibili y and malleabili y, many di e en designs can be c ea ed, including
spo es s, mic o luidic analysis (known as
μ
PADs), and dis ance-based
measu emen s [15]. In gene al, PADs a e used in many ields,
including en i onmen al moni o ing, poin -o -ca e diagnos ics, o en-
sics, clinical analysis, and ood sa e y [16–19]. In addi ion, li e a u e
shows ha hey can be combined wi h i ime ic analyses: chela e
i a ion o Ca
2+
and Mg
2+
de e mina ion [20], edox i a ion o
alcohol con en analysis [21], acid-base i a ion o o maldehyde
quan i ica ion [22] and iodome ic i a ion o asco bic acid and
dopamine de e mina ion [23], o example. Tep ek e al. p esen ed ou
di e en sys ems including alkalini y de e mina ion by acid-base i a-
ion, ha dness quan i ica ion by complexome ic i a ion, edox i a-
ion o hiosul a e quan i ica ion and chlo ide analysis by p ecipi a ion
i a ion [24]. In hese e e ences, EP was iden i ied isually on he PADs
based on he colo change o he indica o . Scaling om
semi-quan i a i e o quan i a i e analysis can be achie ed wi h digi al
colo ime y, as i can be employed o build a calib a ion cu e. Fo
example, by he use o he R channel in ensi y [25] o Euclidean no m
[26], which can be ela ed o concen a ion.
Ace ic acid, also known as e hanoic acid o me hylca boxylic acid, is
a weak o ganic acid ha can be used in he p oduc ion o la ex emulsion
esins, syn hesis o e eph halic acid, ui inega p oduc ion, dyes o
pha maceu ical o mula ions, among o he s [27]. I is also he p ima y
o ganic componen o inega , esponsible o i s scen and bi e la o .
Vinega is de ined as a liquid p oduc ha is p oduced by alcoholic and
ace ic e men a ion o ag icul u al-o igin subs ances. I is mos ly known
o i s applica ions in he ood indus y as salad seasoning, pickling,
la o ing, o as a p ese a i e agen in bo led sauces [28].
Table 1 shows some wo ks ela ed o ace ic acid de e mina ion ound
in he li e a u e. Rega ding i a ion-based echniques, a pH ISFET cell
o wine analysis has been epo ed [29], as well as isual de ec ion o
endpoin o inega samples [30,31], and digi al colo ime y o ace ic
acid de e mina ion in wa e [32]. In ela ion o spec oscopic me hods,
UV–Vis [33] and NIR [34] ha e been used o inega analysis, and
luo escence-based de e mina ions o bac e ia [35] and Chinese wine
[36]. Finally, HPLC o inega , p obio ic and wa e ke i be e ages
[37–40], and
1
H-RMN o mul i a ia e analysis o s awbe ies [41]
ha e been epo ed. In he majo i y o he cases, analy e quan i ica ion
was ca ied ou using an analy ical cu e, ei he wi h uni a ia e cali-
b a ion using po en ial di e ence, abso bance o peak a ea, o example;
o by mul i a ia e analysis me hods.
Li e a u e shows ha ace ic acid can also be de e mined using PADs.
Acid-base i a ions ha e been ca ied ou by di e en me hods such as
spo es s [43] o
μ
PADs, ei he by s a -like designs [44] o
dis ance-based measu emen s [45]. Howe e , he majo i y o he sys-
ems a e based on quali a i e o semi-quan i a i e EP de e mina ion,
some wo ks equi e building a calib a ion line p e iously, which do no
employ he i a ion cu e i sel o acid de e mina ion. In o he cases,
images need o be aken a e each i an addi ion, which make he
me hods labo ious and signi ican ly inc eases expe imen al wo k and
da a analysis complexi y.
In his wo k, mic oscale acid-base i a ions o ace ic acid de e -
mina ion ha e been ca ied ou in wo ypes o pape -based de ices, a
spo es ing de ice and a s a -like design ollowing a mic o luidic
app oach. In he p oposed sys ems, jus an eigh -poin i a ion is
necessa y o ob ain a quan i a i e and accu a e de e mina ion o EP
based on second de i a i e calcula ions, which is achie ed using a single
image. To ou knowledge, i is he i s
μ
PAD sys em whe e EP iden i-
ica ion is comple ely objec i e and based on he i a ion cu e, wi hou
he need o isual de ec ion o p e iously building a calib a ion cu e.
A simple, low-cos , and en i onmen ally iendly me hod is p oposed,
au oma ed o as sample analyses.
2. Expe imen al
2.1. Chemicals and ma e ials
All eagen s we e o analy ical g ade and used wi hou addi ional
pu i ica ion. Li mus, ace ic acid, po assium hyd ogen ph hala e and
absolu e e hanol we e pu chased om Pan eac (Ba celona, Spain). So-
dium hyd oxide was supplied by Sigma-Ald ich (S . Louis, USA) and
Labkem (Ba celona, Spain). Phenolph halein was p o ided by Me ck
(Da ms ad , Ge many). Deionized wa e ( esis i i y ≥18 MΩ cm a
25 ◦C) was ob ained om a Milli-Q Ad an age-0.10 pu i ica ion sys em
(Millipo e, Ge many).
Solu ions o li mus 0.1%, li mus 0.166% and phenolph halein 0.1%
we e p epa ed by dissol ing he app op ia e weigh ed mass o he in-
dica o s in a small amoun o absolu e e hanol and dilu ing o ma k wi h
deionized wa e . S anda d solu ion o ace ic acid 4% was p epa ed om
96% glacial ace ic acid and NaOH 0.1 mol L
−1
was s anda dized wi h
0.056 mol L
−1
po assium hyd ogen ph hala e.
2.2. Fab ica ion o pape -based de ices
Pape -based de ices we e designed using Mic oso O ice Powe -
Poin 2016 so wa e and we e p in ed on quali a i e il e pape using a
wax p in e (Xe ox Colo Qube 8570, CT, USA). The p in ed shee s we e
placed in a he mal p ess (Maquina ec, MG, B azil) o 15 s a 100 ◦C.
Upon hea ing, wax pene a es he pape and de ines hyd ophobic ba -
ie s a ound he pa e n. The back o he p in ed de ice was co e ed
wi h anspa en ape o p e en solu ions om leaking h ough he
de ice.
Two app oaches we e used in his wo k, spo es analysis and
mic o luidic analysis, so wo PADs we e designed and ab ica ed
(Fig. 1). Spo es s we e pe o med in 5 mm diame e ci cles, pa e ned
in a 3 by 8 design in o de o analyze eigh i a ion poin s in iplica e
(Fig. 1a). Mic o luidic de ices we e ab ica ed modi ying he s a -like
design epo ed by Ka i a and Kane a [44], wi h a sample ese oi
loca ed a he cen e , eigh b anches wi h a eac ion ese oi and a
de ec ion zone a he end o each b anch (Fig. 1b).
Table 1
Summa y o analy ical me hods used o ace ic acid de e mina ion ound in he
li e a u e.
Technique Sample De e mina ion Re .
Ti a ion Wine pH ISFET cell and analy ical
cu e
[29]
Vinega Visual endpoin de ec ion [30,31]
pH-me e and analy ical
cu e
[42]
Wa e Digi al colo ime y and
i a ion cu e de i a i es
[32]
Spec oscopy Vinega UV–Vis and analy ical cu e [33]
NIR and mul i a ia e
eg ession
[34]
Bac e ia Fluo escence biosenso and
analy ical cu e
[35]
Chinese wine Fluo escen u anyl-o ganic
amewo k and analy ical
cu e
[36]
HPLC Vinega , p obio ic and
wa e ke i be e ages
UV–Vis and analy ical cu e [37–40]
1
H-RMN S awbe y Mul i a ia e analysis (PCA,
HCA and PLS-DA)
[41]
I. Be asa e e al.
Talan a 276 (2024) 126254
3
2.3. Colo ime ic acid-base i a ion
In bo h app oaches, eigh -poin acid-base i a ions we e ca ied ou .
In o de o ind he op imal measu emen condi ions, some pa ame e s
we e op imized. These pa ame e s include de ice size, indica o , i an
and sample olumes, d ying me hod and image acquisi ion ime.
Rega ding spo es s, i s , mix u es o li mus, NaOH and wa e we e
added o he spo s, and hen, sample was added. Immedia ely, PADs
we e le o d y and images we e aken a di e en imes o ind he
op imal condi ions. Fo mic o luidic analyses, inal concen a ions in
spo es s we e aken as e e ence. Fi s , a ying NaOH concen a ion
solu ions we e added o all eac ion ese oi s. Then, li mus was added
o all de ec ion zones. Finally, sample was placed in he cen e o he
μ
PAD. De ices we e le o d y and images we e aken a di e en imes
o achie e he op imal condi ion.
In bo h cases, digi al images o PADs we e acqui ed wi h a sma -
phone OnePlus No d CE ( h ee came as, 64 +2 +8 MP) in au oma ic
mode. In o de o main ain image acquisi ion condi ions cons an , a
home-made ligh box was used. The po able wooden box (dimensions:
10 cm heigh , 12 cm wid h, 15 cm leng h) was ab ica ed o elimina e
ex e nal ligh in e e ences, wi h 24 V whi e LED ligh s along he in e-
io . Bo h PADs and he sma phone we e always placed in ixed posi-
ions o a oid u he a ia ion be ween pho og aphs. RGB alues we e
ex ac ed using he ee image-edi ing so wa e GIMP ( e sion 2.10.34).
Image analysis was also ca ied ou using MATLAB R2020b e sion (The
Ma hwo ks Inc., Massachuse s, USA).
2.4. Vinega samples
Bo h o he p oposed me hods we e es ed in i een inega samples.
Fi e samples we e pu chased in local ma ke s om Campinas (S˜
ao
Paulo, B azil), including lemon, ice and alcohol inega (Cas elo Ali-
men os S.A.) and wo di e en apple inega samples (Cas elo Alimen os
S.A. and Alma omi, bo h om S˜
ao Paulo, B azil). The o he 10 samples
we e pa o an indus ial p ocess, p o ided by JR Saba e S.A. (Mu cia,
Spain), and included i e samples de i ed om alcoholic e men a ion
and ano he i e samples o whi e wine e men a ion. Comme cial
samples we e dilu ed 10 imes, as indica ed concen a ion on he label is
4%, whe eas indus ial samples, wi h a heo e ical concen a ion o
a ound 10%, we e dilu ed 25 imes.
Acid-base po en iome ic i a ion was used as he e e ence me hod.
Ace ic acid was de e mined using he 794 Basic Ti ino (Me ohm,
Swi ze land) au oma ic i a o coupled o a pH elec ode. Fi s ly, NaOH
was s anda dized wi h po assium hyd ogen ph hala e and
phenolph halein in iplica e. Then, 0.25 mL o 0.5 mL o inega sample
(depending on i s heo e ical concen a ion) and 2.0 mL o li mus 0.1%
indica o we e added o a beake . Finally, solu ions we e i a ed wi h
s anda dized NaOH solu ion un il he colo changed om ed o blue. EP
olume was used o he calcula ion o ace ic acid concen a ion. Mea-
su emen s we e pe o med in iplica e.
3. Resul s and discussion
3.1. Op imiza ion o PAD-based sensing
3.1.1. Colo channel selec ion
In his wo k, li mus was used as acid-base indica o (pK
a
6.5), which
colo change occu s be ween pH 4.5–8.3 om ed o blue, wi h pu ple
colo a neu al alues. Ace ic acid is a weak acid o which i a ion EP
lies be ween pH 6.0–9.0. Fo ha eason, li mus was chosen o e some
common indica o s such as phenolph halein, as i allows us o obse e a
b oade colo change close o he EP, p o iding be e p ecision and
sensi i i y o his weak acid-s ong base i a ion.
A syn he ic solu ion o ace ic acid 0.4%, a e a 1:10 dilu ion o he
4% s ock solu ion, was used o p ocedu e op imiza ion, and i s ly,
some p elimina y es s we e pe o med in 96-well mic opla es o selec
he op imal colo channel. Digi al images we e analyzed o ex ac ed
(R), g een (G) and blue (B) alues o each spo , which we e plo ed
agains he i an olume added o achie e a classic acid-base i a ion
cu e (Fig. S1). The h ee channels had mo e o less cons an alues
be o e EP. Then, R and G channels d as ically dec ease, while B channel
sligh ly inc eases. A e EP, R alues become cons an , while G and B
alues inc ease a bi . Fo his eason, R channel was selec ed as he colo
channel o moni o ing he i a ions, as i gi es he mos sigmoidal
cu e.
3.1.2. Op imiza ion o spo es s p ocedu e
Fab ica ion o PADs was done by wax p in ing and he mal hea ing,
and no ela ed p oblem was iden i ied du ing he expe imen s. Fi s and
o emos , he op imal indica o olume o he designed and wax-
p in ed de ice was in es iga ed. Two indica o olumes we e es ed,
5
μ
L and 7
μ
L. I was easily obse ed by naked eye ha 7
μ
L was oo
much, as d ying ime inc eased signi ican ly and some imes he d op
exceeded he wax, so 5
μ
L was chosen as op imal indica o olume.
Then, 96-well mic opla es we e used o es ima e i an olumes,
which we e scaled o PAD p opo ions. Once NaOH olumes we e ixed
as 1.0–6.0
μ
L, i a ions we e ca ied ou using h ee di e en concen-
a ions o li mus. Colo alues wi h li mus 0.05% we e oo ligh , and he
Fig. 1. Design o pape -based de ices. (a) Wax-p in ed pape de ice wi h indi idual spo s o an eigh -poin i a ion, wi h h ee ows o analysis in iplica e. (b)
Wax-p in ed
μ
PAD o mic o luidic analysis, wi h eigh iden ical b anches loca ed equidis an ly om he cen e (sampling zone). Leng hs a e indica ed in millime e s.
I. Be asa e e al.
Talan a 276 (2024) 126254
4
calcula ion o he concen a ion was mo e subjec ed o e o . In con as ,
a high concen a ion o li mus as 0.2% comp omised he i a ion, as EP
was shi ed o highe alues, and hus, he calcula ion o acidi y was
a ec ed. Wi h a concen a ion o 0.1%, bo h i a ion cu es and ace ic
acid concen a ion showed good esul s, so i was selec ed as he op imal
concen a ion.
Impo an di e ences we e obse ed be ween adding he eagen s
indi idually and adding hem oge he . When eagen s we e added
indi idually, i a ion cu es we e no homogeneous and he me hod
became un eliable. When bo h eagen s we e added oge he , epea -
abili y was imp o ed, and o ha eason, li mus and NaOH we e added
oge he . The mix u e is s able o a long ime, i can be p e iously
p epa ed, and he co ec mix u e o he eagen s is ensu ed. Mix u es o
5
μ
L li mus, 1–6
μ
L NaOH, and 1–5
μ
L wa e we e p epa ed in ials, and
11
μ
L was added o each o he eigh i a ion spo s. Then, 4
μ
L o a
s anda d o ace ic acid 0.4% was added o s a he eac ion. Thus, inal
concen a ions o each compound we e: NaOH 0.007–0.04 mol L
−1
,
li mus 0.03%, and sample 0.1%.
Two o he mos impo an ac o s o conside a e he d ying me hod
and he d ying/ eac ion ime. On he one hand, de ices we e d ied a
oom empe a u e and in he o en (Fig. 2a). PADs le a oom em-
pe a u e ook a lo o ime o d y, up o 90 min o mo e. This was caused
by mode a e o high ai humidi y, which a ies g ea ly be ween days
and seasons, impeding he s anda diza ion o he p o ocol. In con as , i
he de ices we e d ied in an o en, PADs we e comple ely d y in 30 min
maximum, so i was selec ed as d ying me hod. A empe a u e o 50 ◦C
was used o a oid he de o ma ion o he wax-based ba ie .
On he o he hand, ega ding d ying ime (Fig. 2b), i could be
isually de ec ed ha spo s changed colo in he i s minu es a e
addi ion, so images acqui ed immedia ely and a e 5 min did no gi e
good esul s. A e 10 min, good EP and concen a ion esul s we e
ob ained. Howe e , because he d ops we e no comple ely d y, pixel
selec ion o da a ex ac ion was limi ed, and hus, he shape o he
cu es was no comple ely sigmoidal. A 20 min, some spo s we e d y
and o he s we e no , so i a ion cu es showed a iable R alues wi h
high e o ba s, impac ing EP de ini ion and p o iding inaccu a e e-
sul s. A e 30 min in he o en, all spo s we e comple ely d y and image
quali y was good, which esul ed in good i a ion cu es and concen-
a ion esul s in all cases, so his ime was se as op imal.
Finally, i needs o be no ed ha he educ ion o o al eagen s and
sample olume o each spo om 15
μ
L o 10
μ
L o 8
μ
L, o example,
could be in es iga ed. I will depend on he sys em unde s udy and he
necessa y concen a ion o each eagen , bu ha way, he o al d ying
ime would be educed, accele a ing he p ocess.
3.1.3. Op imiza ion o mic o luidic analysis p ocedu e
Mic o luidic pape -based de ices (
μ
PADs) we e ab ica ed as
men ioned in he Expe imen al pa . Fi s ly, NaOH concen a ion and
olume we e op imized. Mix u es wi h di e en concen a ions o
i an we e p epa ed in small ials, conside ing he inal i an mmol
added in spo es s. Reac ion ese oi s could be illed wi h 1
μ
L in he
o iginal design [44], bu as ou design consis ed o 8 b anches ins ead o
10, op imiza ion was ca ied ou wi h 0.50, 0.75, and 1.00
μ
L o i an .
Wi h he lowes olume, inaccu a e esul s we e ob ained due o he
incomple e illing o he whole eac ion ese oi . In con as , he
highes olume sp ead h ough he channels o he de ice, and in e -
e ed wi h concen a ion calcula ions. I was de e mined ha 0.75
μ
L
(NaOH s anda ds be ween 0.02 and 0.12 mol L
−1
) was he op imal
olume o i an o be added o he eac ion ese oi s, as i a oided
bo h p oblems men ioned and o e ed good concen a ion esul s.
Rega ding li mus olume and concen a ion, i was isually de ec -
able ha 0.5
μ
L was enough o ill he de ec ion zone co ec ly. The
concen a ion was ecalcula ed acco ding o he li mus-NaOH ela ion
used in spo es s con igu a ion, so a new s ock solu ion o li mus 0.166%
was p epa ed and used.
In his
μ
PAD sys em, sample was placed in he cen e o sp ead
h ough he channels. P elimina y es s we e ca ied ou using 28, 24, 20
and 16
μ
L o sample. I was obse ed ha he wo smalles olumes we e
no enough o comple e he i a ion. By naked eye, colo s ob ained wi h
28
μ
L and 24
μ
L we e simila . Howe e , R channel alues and i a ion
cu es showed ha he mos accu a e concen a ion alues we e ob-
ained using 24
μ
L o he sample.
Once he op imal sample olume was se , eac ion ime was in es-
iga ed. Images we e acqui ed using he ligh ing box a 0, 2.5, 5 and 10
min. I was obse ed ha 10 min was he necessa y ime o comple ely
d y he de ice. Room empe a u e was selec ed as he d ying me hod
due o he size o he
μ
PAD and he small olumes used in he analysis.
3.1.4. Summa y o colo ime ic p ocedu es
Rega ding spo es s, each o he i a ion poin s was analyzed in
iplica e wi h he same de ice. Fi s ly, 11
μ
L o a eagen mix u e was
added o all spo s. The mix u e con ained 5.00
μ
L o li mus 0.1%,
a ying olumes o NaOH 0.10 mol L
−1
(1.00–6.00
μ
L), and wa e up o
11.0
μ
L. A e , 4.00
μ
L o sample 0.4% was added. Immedia ely, PADs
we e placed in he o en a 50 ◦C, and images we e aken a e 30 min.
Fo
μ
PAD analysis, 0.75
μ
L o a ying NaOH concen a ion solu ions
(0.02–0.12 mol L
−1
) we e added o all eac ion ese oi s, and hen,
0.50
μ
L o li mus 0.166% was added o all de ec ion zones. Nex , 24
μ
L o
sample 0.4% we e placed in he cen e o he
μ
PAD (sampling zone). The
de ice was d ied a oom empe a u e and images we e acqui ed a e
Fig. 2. (a) Ti a ion cu es ob ained wi h wo di e en d ying me hods. D ying a oom empe a u e ook up o 90 min, in con as wi h he o en, which ook only
30 min. (b) Ti a ion cu es ob ained om PADs d ied in he o en o di e en imes.
I. Be asa e e al.
Talan a 276 (2024) 126254
5
10 min.
3.1.5. Me hod alida ion
Once op imal analysis pa ame e s we e iden i ied, a s anda d solu-
ion o ace ic acid (0.4%) was used o me hod alida ion. In he case o
spo es s, in a-day p ecision was calcula ed using h ee PADs measu ed
on he same day (n =9), and in e -day p ecision wi h ou de ices om
h ee di e en days (n =12). In he case o
μ
PADs, i een de ices
di ided in o 5 images (3 de ices pe image) we e used o in a-day
p ecision calcula ion (n =15), and in e -day p ecision was e alua ed
using nine
μ
PADs om 3 images aken in h ee di e en days (n =9).
Tes s pe o med wi h he s anda d solu ion showed good p ecision and
accu acy esul s by bo h he p oposed me hods (Table 2), which come in
ag eemen wi h he guideline o AOAC [46].
3.2. Colo ime ic i a ion and ace ic acid calcula ion
As p e iously s a ed, images o spo s PADs we e ob ained a e 30
min in he o en, while images o
μ
PADs we e cap u ed a e 10 min a
oom empe a u e. In bo h cases, R channel alues we e used o build
i a ion cu es, ob aining sigmoidal shapes. A e ex ac ing R channel
alues, he i s and second de i a i es we e calcula ed and plo ed, as i
is he co ec way o ma hema ically calcula ing he in lec ion poin o
his ype o cu e. In his i a ion, he i s de i a i e esul s in a min-
imum alue a he endpoin , whe eas i equals ze o on he second
de i a i e.
Assuming ha he eac ion be ween he weak acid and he s ong
base is comple e, and due o he 1:1 s oichiome y o he eac ion, i can
be assu ed ha he NaOH mmol numbe equals ace ic acid mmol exac ly
a EP. Using he R colo alues and he i an olumes, expe imen al
de i a i es we e calcula ed. Then, a eg ession line was buil be ween
he maximum and minimum poin s o he second de i a i e cu e.
Subsequen ly, EP olume o mmol numbe was calcula ed o y =
0 using he slope and in e cep o he eg ession line. Finally, ace ic acid
concen a ion (%) was calcula ed.
In he case o spo es s, sample olume is a known alue (4
μ
L), so R
alues we e plo ed agains he olume o NaOH added, and ace ic acid
concen a ion was calcula ed di ec ly using ha alue. In ela ion o
mic o luidic analyses, 0.75
μ
L o di e en i an solu ions we e added
o he eac ion ese oi s, and hen, he sample was added o he cen e
o he de ice, so i could sp ead o he channels o he
μ
PAD. To build
i a ion cu es, R channel alues we e plo ed agains he added NaOH
mmol numbe . In his case, added NaOH olume was he limi ing ac o ,
so he amoun o sample ha ook pa in he i a ion is he amoun o
i an ha was ixed (0.75
μ
L). A schema ic illus a ion o he p ocedu e
o EP calcula ion is shown in Fig. 3.
3.3. Vinega sample analysis
Volume ic analysis pe o med wi h an au oma ic i a o and
po en iome ic de ec o was he e e ence me hod used o inega
sample analysis. In addi ion, he p oposed pape -based me hods, bo h
spo es s and
μ
PADs, we e employed. All indus ial and comme cial
samples we e analyzed in iplica e in all cases. Some inega ypes may
con ain small amoun s o a a ic and ci ic acid. Howe e , due o he
dilu ion applied o he samples (1:10 o 1:25), no in e e ence should be
expec ed om hose compounds. Fig. 4 shows he i a ion cu es
ob ained wi h bo h me hods o a sample (C101) selec ed as example.
Rega ding spo es s, ace ic acid concen a ions we e 4.4%–4.9% o
comme cial inega s and 10.1%–17% o indus ial samples. Mic o-
luidic measu emen s ga e concen a ions be ween 4.2% and 4.8% o
comme cial samples and 10.2%–17% o indus ial samples. P ecision
alues (RSD%) anged be ween 0.6% and 13% in spo es ing, and be-
ween 0.9% and 9.9% o
μ
PAD analysis. I needs o be no ed ha , e en
i some RSD alues we e a bi high, hey we e below 10% o he g ea
majo i y o samples. Conside ing he di e en measu ing loca ions and
ha some samples we e no measu ed as soon as hey we e opened o
p oduced, especially in he case o he indus ial samples, accep able
esul s we e ob ained wi h he p oposed PADs.
Besides, ela i e e o s ob ained by spo es s we e all below 10.5%,
wi h nine samples below 5%. E o s ob ained by mic o luidic mea-
su emen s we e simila , wi h 14% being he highes ela i e e o and
wi h en samples below 5%. In conclusion, simila esul s we e ob ained
wi h bo h me hods in compa ison o he e e ence me hod. De ailed
alues a e summa ized in Table 3.
Resul s ob ained by po en iome ic i a ion and he p oposed digi al
colo ime y i a ion on PADs we e s a is ically e alua ed. Two di e en
es s we e conduc ed. Fi s , since sample concen a ions a y in a wide
ange, a join con idence ellipse es (EJCR) o slope and in e cep was
pe o med [47,48]. In he case o spo es s, he con idence in e al o
he slope was 1.00 ±0.09, and 0.2 ±0.8 o he in e cep . Fo he
μ
PAD
app oach, he slope was 1.00 ±0.08, and in e cep was −0.1 ±0.8
(Fig. S3). Conside ing hese alues, he ideal poin (slope 1 and in e cep
0) lies wi hin hese anges in bo h PADs, so he p oposed me hods do no
di e signi ican ly om he e e ence me hod.
Then, a pai ed - es was pe o med wi h a 95% con idence le el. No
signi ican di e ences we e ound be ween e e ence and spo es s (
exp
1.80 <
c i
2.14), no be ween e e ence and mic o luidic measu emen s
(
exp
1.20 <
c i
2.14), since he calcula ed alue was below he abu-
la ed one. In conclusion, i can be assu ed ha none o he p oposed
me hods p esen s sys ema ic e o s and ha hey a e eliable me hods
o ace ic acid de e mina ion in inega samples.
Collec i ely, he p oposed mic oscale i a ion on PADs combined
wi h digi al colo ime y p esen s signi ican imp o emen s om he
G een Analy ical Chemis y poin o iew. I educes he amoun o e-
agen s consumed and was e gene a ed due o i s minia u iza ion, and
p o ides au oma ed analyses compa ed o classical i a ion, p o iding
an AGREE sco e [49] o 0.71 o spo es ing, 0.78 o he
μ
PAD
app oach and 0.49 o he classical i a ion me hod (Fig. S4). These
alues demons a e ha mic oscale i a ions pe o med on PADs a e
mo e sus ainable han he adi ional po en iome y me hod. In addi-
ion, he use o digi al colo ime y o i a ion imp o es he accu acy o
he me hod because i exemp s he analys om isually iden i ying he
colo endpoin , which may be subjec i e.
3.4. Me hod au oma ion
Me hod au oma ion in he ood indus y, speci ically o analy e
moni o ing, is c ucial o ensu ing accu acy and e iciency in he es ing
p ocess. Au oma ion educes human e o s, enhances p ecision, and
allows o a highe h oughpu o samples. In his ega d, wo image
analysis so wa e we e used and compa ed in his wo k.
On he one hand, he ee so wa e GIMP was employed. A Mic oso
Excel (Mic oso , 2016 e sion) empla e was c ea ed o simpli y and
speed up da a analysis, and jus by inse ing he R channel alues
ex ac ed manually by GIMP, he empla e makes he necessa y i s and
second de i a i e calcula ions, co esponding i a ion g aphs, and
ace ic acid concen a ion calcula ions. E en i analysis ime was
educed, he colo ex ac ion wo k becomes edious when dealing wi h a
la ge numbe o measu emen s.
As an al e na i e, a use - iendly au oma ed unc ion was designed in
MATLAB using he Image P ocessing Toolbox™. This way, manual
ex ac ion is a oided, a single so wa e is used, and a s ack o images
Table 2
Me hod alida ion esul s o he wo ypes o PADs de eloped in his wo k.
PAD ype Acidi y (%) P ecision (RSD, %) Accu acy (RE, %)
In a-day In e -day
Spo s 0.427 ±0.007 0.4–1.9 4.8 2.7–9.1
μ
PAD 0.39 ±0.02 1.0–6.4 5.4 1.4–3.8
I. Be asa e e al.
Talan a 276 (2024) 126254
6
aken in he same posi ion can be analyzed in a single un, p o iding a
high h oughpu wo k low. A schema ic illus a ion o he p ocedu e is
shown in Fig. S2. A ew inpu s need o be speci ied o un he unc ion:
image name o o ma , dilu ion ac o o he sample, and in he case o
spo es s, NaOH concen a ion. Then, all necessa y s eps om dimen-
sion selec ion o ace ic acid calcula ion a e pe o med au oma ically.
Finally, a double g aph wi h all he esul s becomes isible ( i a ion
cu e, 2
nd
de i a i e g aph, EP, and ace ic acid concen a ion in he
sample), as well as an ou pu ma ix in he wo kspace wi h he con-
cen a ion esul s.
The esul s ob ained wi h bo h me hods we e almos iden ical,
indica ing he use ulness and e ec i eness o he moni o ing o ace ic
acid concen a ion ei he in academia o indus ial p ocesses. Mo eo e ,
he p oposed PADs can be employed o pe o m o he ypes o i a ions,
as he unc ion is a ailable unde eques and i is easily adjus able o
o he eac ions jus by modi ying i an olumes o concen a ions.
3.5. Compa ison wi h o he wo ks
As was men ioned in he in oduc ion, de e mina ion o ace ic acid
using PADs has been p e iously epo ed in he li e a u e. Table 4 shows
he wo ks whe e spo es ing and
μ
PADs ha e been used o ha
pu pose.
In ela ion o spo measu emen s, he wo ks ound we e majo ly
ela ed o eaching and adap ing he pape -based me hod o expe i-
men al labo a o y cou ses. The mos used i an was NaOH, and he
Fig. 3. Schema ic illus a ion o EP calcula ion p ocedu e. Mean R alues we e ex ac ed and plo ed agains added NaOH (
μ
L o mmol, he la e in his example
igu e). Second de i a i es we e calcula ed o he cu es. A eg ession line was cons uc ed be ween he p e ious and he ollowing poin o he cu e c ossing a y
=0, and om i , EP (
μ
L o mmol) was calcula ed.
Fig. 4. Ti a ion cu es ob ained o C101 sample selec ed as example. (a) Spo es ing, iplica e analysis in he same de ice, and (b) mic o luidic measu emen s
pe o med h ee imes.
I. Be asa e e al.
Talan a 276 (2024) 126254
7
indica o s a ied om he classic phenolph halein o o he s ha o e
g adual colo changes such as jabo icaba peel ex ac o li mus, as
p oposed in his wo k. In gene al, he a iable pa ame e was he ol-
ume o i an , bu he change in ace ic acid concen a ion has also been
epo ed. PAD analysis has been ca ied ou ei he by naked eye, digi al
came a o sma phone, and EP has been de e mined based on isual
de e mina ion, bigges jumps, o he in e sec ion be ween wo lines. One
wo k p oposed a me hodology based on he 1
s
de i a i e, simila o he
one p oposed. Howe e , hese me hods a e ei he subjec i e, semi-
quan i a i e o labo ious, especially when hey a e based on images
aken a e each i an addi ion, as p oposed by Noguei a and co-
wo ke s [50], which signi ican ly inc eases expe imen al wo k and da a
analysis complexi y. The main ad an age o he p oposed spo - es ing
me hod is ha he need o cap u ing a ious images is elimina ed.
Ins ead, mul iple spo s a e used simul aneously, expedi ing he en i e
i a ion p ocess wi h a unique image.
Rega ding
μ
PADs, only wo wo ks we e ound whe e ace ic acid
de e mina ion was ca ied ou . In he wo k o Ka i a and Kane a [44], a
s a -like design wi h 10 b anches was used, i an was placed in he
eac ion ese oi s, and indica o in o he de ec ion zones, as pe o med
in his wo k. In ha s udy, a semi-quan i a i e analysis was pe o med,
and i a ion endpoin was de e mined isually based on he colo less o
pink colo change o phenolph halein indica o . In his wo k, he design
was modi ied and educed o lowe lobes in o de o in es iga e i i-
a ions can be ca ied ou quan i a i ely wi h jus wo poin s a ound EP.
As explained in p e ious sec ions, good esul s we e ob ained wi h an
8-poin i a ion, demons a ing he p ecise applicabili y o he me hod.
In he case o Dias e al. [45], ace ic acid s anda ds we e used, placed
on he
μ
PAD, and a eled dis ance was measu ed wi h a digi al calibe .
Dis ance o he colo ed channel o he
μ
PAD was ela ed o he analy e
concen a ion o build a calib a ion cu e. Then, samples we e
measu ed ollowing he same p inciple. Al hough hese app oaches a e
based on acid-base i a ion, he i a ion cu e i sel is no used o acid
de e mina ion, he colo change o phenolph halein may lead o inac-
cu a e EP de e mina ions, and he need o build a calib a ion line in-
c eases da a analysis ime. As an al e na i e, he me hod p oposed in
his wo k enables he isualiza ion o a g adual colo change, which can
be de ec ed by sma phone imaging, quan i a i e de e mina ion o EP
can be ca ied ou , and hus, ace ic acid concen a ion can be de e -
mined accu a ely based on an eigh -poin i a ion and jus one image.
As men ioned o spo es ing, as and easy- o-use me hodologies ha
upli p e iously epo ed wo ks we e de eloped.
4. Conclusions
I was demons a ed ha he p oposed me hods o mic oscale acid-
base i a ions using pape -based de ices a e eliable o he quan i i-
ca ion o ace ic acid in inega samples. Two ypes o de ices – based on
spo es s and
μ
PADs – we e de eloped and op imized o analysis,
elimina ing he subjec i e de ec ion o eac ion endpoin and he need o
build a calib a ion cu e. Ad ances om o he wo ks we e ob ained:
i a ions we e ca ied ou using jus eigh analysis poin s and one image
ob ained wi h a sma phone, a comple ely quan i a i e
μ
PAD was
Table 3
Ace ic acid concen a ion alues gi en as acidi y pe cen age and ob ained by he
e e ence me hod, spo -based PADs, and mic o luidic de ices. Rela i e e o s in
compa ison o po en iome y we e calcula ed in each case.
Sample Re e ence
me hod
Spo es s Mic o luidic analysis
C
ace ic
(%) C
ace ic
(%)
RE
(%)
Cace ic
(%)
RE
(%)
A102 10.5 ±0.2 11.3 ±
0.5
7.6 10.2 ±
0.7
−2.8
E102 10.3 ±0.3 11.2 ±
0.3
8.5 10.3 ±
1.1
0.5
A106 12.1 ±0.1 13.1 ±
0.4
8.4 10.4 ±
0.8
−13.9
D106 11.0 ±0.1 10.1 ±
1.5
−8.1 10.4 ±
0.5
−5.1
B105 9.9 ±0.1 10.1 ±
1.3
2.6 10.3 ±
0.9
5.6
E105 11.0 ±0.2 10.8 ±
0.8
−1.8 11.0 ±
0.1
0.4
F105 11.15 ±0.03 12.2 ±
0.4
9.4 10.5 ±
0.6
−6.2
B101 12.95 ±0.04 12.8 ±
0.5
−1.4 12.9 ±
0.3
−0.4
C101 14.4 ±0.1 15.0 ±
0.1
2.8 14.8 ±
1.3
2.7
F101 17.0 ±0.3 16.8 ±
1.2
−1.2 17.3 ±
0.4
1.6
Lemon
inega
4.30 ±0.01 4.4 ±0.3 3.3 4.2 ±0.3 −2.8
Rice inega 4.6 ±0.2 4.53 ±
0.05
−2.0 4.3 ±0.1 −6.7
Apple
inega (1)
4.8 ±0.1 4.9 ±0.2 3.2 4.8 ±0.1 1.1
Apple
inega (2)
4.35 ±0.04 4.8 ±0.2 10.4 4.39 ±
0.06
0.9
Alcohol
inega
4.52 ±0.05 4.8 ±0.2 4.6 4.5 ±0.2 −1.8
Table 4
Summa y o wo ks epo ed in he li e a u e o ace ic acid de e mina ion using PADs.
PAD Reagen s Colo change P ocedu e De ec ion EP V
o al
Samples P ecision and
accu acy
Re .
Spo s NaOH and jabo icaba
peel ex ac
Magen a o
g een
Va ying V
i an
Image a e each
addi ion
Sma phone 1
s
de i a i e 20
μ
L Vinega (3) RSD 2.8–4.8%
RE 4.6–7.5%
[50]
Na
2
CO
3
and
phenolph halein
Colo less o
pink
Va ying C
ace ic
One image
Digi al came a and
ligh ing box
In e sec ion o
wo lines
a
– – [51]
NaOH and
phenolph halein
Colo less o
pink
Va ying V
ace ic
Naked-eye Naked-eye 90
μ
L Foods (15) RSD 0–12.6%
RE 0.5–12.8%
[43]
NaOH and uni e sal
indica o
a
Va ying V
i an
Image a e each
addi ion
Sma phone Bigges pH jump 100
mL
Foods and
o he s
RSD 7.1–28%
RE 5.8–40%
[52]
NaOH and li mus Red o blue Va ying V
i an
One image
Sma phone and
ligh ing box
2
nd
de i a i e 110
μ
L
Vinega (15) RSD 0.6–13%
RE 1.2–10.4%
This
wo k
μ
PAD Na
2
CO
3
and
phenolph halein
Colo less o
pink
Va ying C
i an
Naked-eye Naked-eye 45
μ
L – – [44]
NaOH and
phenolph halein
Colo less o
pink
Calib a ion line Dis ance (digi al
calibe )
– 160
μ
L
Vinega (3) RSD 4.3–4.7%
RE 1.9–2.4%
[45]
NaOH and li mus Red o blue Va ying C
i an
One image
Sma phone and
ligh ing box
2
nd
de i a i e 34
μ
L Vinega (15) RSD 0.9–9.9%
RE 0.4–14%
This
wo k
a
No men ioned.
I. Be asa e e al.
Talan a 276 (2024) 126254
8
p oposed o he i s ime, he use o li mus and second de i a i es
enabled a highly accu a e endpoin calcula ion, and he me hod was
au oma ed ei he o ee image-p ocessing so wa e o o MATLAB.
This way, analysis ime and complexi y, and eagen and sample ol-
umes we e conside ably educed, minimizing was e gene a ion and
enabling a mo e en i onmen ally- iendly me hod. Bo h me hods a e
p ecise, accu a e, easy o use and as , making hem sui able o quali y
con ol p ocesses and eal- ime moni o ing.
CRediT au ho ship con ibu ion s a emen
I a i Be asa e: W i ing – e iew & edi ing, W i ing – o iginal d a ,
So wa e, Me hodology, In es iga ion, Concep ualiza ion. Ane Bo da-
ga ay: W i ing – e iew & edi ing. Rosa Ga cia-A ona: W i ing – e-
iew & edi ing. Mi en Os a: W i ing – e iew & edi ing, Supe ision,
Resou ces, P ojec adminis a ion, Concep ualiza ion. William Reis de
A aujo: W i ing – e iew & edi ing, Resou ces, Concep ualiza ion.
Maide Vidal: W i ing – e iew & edi ing, Supe ision, Resou ces,
Concep ualiza ion.
Decla a ion o compe ing in e es
The au ho s decla e ha hey ha e no known compe ing inancial
in e es s o pe sonal ela ionships ha could ha e appea ed o in luence
he wo k epo ed in his pape .
Da a a ailabili y
Da a will be made a ailable on eques .
Acknowledgemen s
This wo k was suppo ed by he Basque Go e nmen (Resea ch
G oup IT1662/22), he Uni e si y o he Basque Coun y (p e-doc o al
schola ship PIF 19/131), S˜
ao Paulo Resea ch Founda ion (FAPESP)
[G an numbe s: 2018/08782–1, and 2022/03250-7], and Na ional
Council o Scien i ic and Technological De elopmen – CNPq (G an
numbe : 310282/2022-5).
Appendix A. Supplemen a y da a
Supplemen a y da a o his a icle can be ound online a h ps://doi.
o g/10.1016/j. alan a.2024.126254.
Re e ences
[1] D.C. Ha is, Quan i a i e Chemical Analysis, se en h ed., W. H. F eeman and
Company, 2007.
[2] E. da N. Gaiao, V.L. Ma ins, W. da S. Ly a, L.F. de Almeida, E.C. da Sil a, M.C.
U. A aújo, Digi al image-based i a ions, Anal. Chim. Ac a 570 (2006) 283–290,
h ps://doi.o g/10.1016/j.aca.2006.04.048.
[3] D.S.A. Beeckman, G. Meesen, P. Van Oos eld , D. Van ompay, Digi al i a ion:
au oma ed image acquisi ion and analysis o load and g ow h o chlamydophila
psi aci, Mic osc. Res. Tech. 72 (2009) 398–402, h ps://doi.o g/10.1002/
jem .20694.
[4] S. Soa es, G.M. Fe nandes, F.R.P. Rocha, Sma phone-based digi al images in
analy ical chemis y: Why, when, and how o use, T ends Anal. Chem. 168 (2023)
117284, h ps://doi.o g/10.1016/j. ac.2023.117284.
[5] S.E. Umbaugh, Compu e Imaging: Digi al Image Analysis and P ocessing, CRC
P ess, 2005.
[6] R. Deng, X. Chao, H. Li, X. Li, Z. Yang, H.Z. Yu, Sma phone-based mic opla e
eade o high- h oughpu quan i a ion o disease ma ke s in se um, Analys 148
(2023) 735–741, h ps://doi.o g/10.1039/d2an01571d.
[7] D. Ba zallo, L. Fe e , E. Palacio, Eco- iendly sc eening me hod o sul onamides
using a 3D handheld sma phone-based luo escence de ec ion de ice and
g aphene nanopla ele -packed pipe e ip mic oex ac ion, J. En i on. Chem. Eng.
12 (2024) 111888, h ps://doi.o g/10.1016/j.jece.2024.111888.
[8] G. Albizu, M. Vidal, A. Bo daga ay, R. Ga cía-A ona, M. Os a, Semi-au oma ic
digi al image con ol o nickel concen a ion in a Ni-P elec oless acid ba h using
he hexaamminenickel blue complex, Mic ochem. J. 199 (2024) 110073, h ps://
doi.o g/10.1016/j.mic oc.2024.110073.
[9] M. Zugazua-Ganado, A. Bo daga ay, J. Ezena o, R. Ga cia-A ona, M. Os a,
Adap a ion o he Folin-Ciocal eu and Fas Blue BB spec opho ome ic me hods o
digi al image analysis o he de e mina ion o o al phenolic con en : educ ion o
eac ion ime , in e e ences and sample analysis, LWT–Food Sci. Technol. 193
(2024) 115756, h ps://doi.o g/10.1016/j.lw .2024.115756.
[10] D.A. Volme , L. Cu bani, T.A. Pa ke , J. Ga cia, L.D. Schul z, E.M. Bo ges,
De e mina ion o i a able acidi y in wine using po en iome ic, Conduc ome ic,
and Pho ome ic me hods, J. Chem. Educ. 94 (2017) 1296–1302, h ps://doi.o g/
10.1021/acs.jchemed.6b00891.
[11] L.M.A. de Oli ei a, V.B. dos San os, E.K.N. da Sil a, A.S. Lopes, H.A. Dan as-Filho,
An en i onmen - iendly spo es me hod wi h digi al imaging o he mic o-
i a ion o ci ic ui s, Talan a 206 (2020) 120219, h ps://doi.o g/10.1016/j.
alan a.2019.120219.
[12] K. Ru anako n, N. Phadungcha oen, W. Laiwa anapaisal, A. Chins iwongkul,
T. Rojana a a, Sma phone-based echnique o he de e mina ion o a i a ion
equi alence poin om an RGB linea -segmen cu e wi h an example applica ion
o minia u ized i a ion o sodium chlo ide injec ions, Talan a 233 (2021) 122602,
h ps://doi.o g/10.1016/j. alan a.2021.122602.
[13] L.A. P adela-Filho, W.B. Veloso, I.V.S. A an es, J.L.M. Gongoni, D.M. de Fa ias, D.
A.G. A aujo, T.R.L.C. Paix˜
ao, Pape -based analy ical de ices o poin -o -need
applica ions, Mic ochim. Ac a 190 (2023) 179, h ps://doi.o g/10.1007/s00604-
023-05764-5.
[14] E. No iana, T. Oze , C.S. Ca ell, J.S. Link, C. McMahon, I. Jang, C.S. Hen y,
Mic o luidic pape -based analy ical de ices: om design o applica ions, Chem.
Re . 121 (2021) 11835–11885, h ps://doi.o g/10.1021/acs.chem e .0c01335.
[15] T. Oze , C. McMahon, C.S. Hen y, Ad ances in pape -based analy ical de ices,
Annu. Re . Anal. Chem. 13 (2020) 85–109, h ps://doi.o g/10.1146/annu e -
anchem-061318-114845.
[16] L.F. de Lima, A.L. Fe ei a, I. Ranjan, R.G. Collman, W.R. de A aujo, C. de la
Fuen e-Nunez, A bac e ial cellulose-based and low-cos elec ochemical biosenso
o ul asensi i e de ec ion o SARS-CoV-2, Cell Repo s Phys. Sci. 4 (2023)
101476, h ps://doi.o g/10.1016/j.xc p.2023.101476.
[17] M. Dong, J. Wu, Z. Ma, H. Pe e z-So oka, M. Zhang, P. Komenda, N. Tang i, Y. Liu,
C. Riga o, F. Lin, Rapid and low-cos CRP measu emen by in eg a ing a pape -
based mic o luidic immunoassay wi h sma phone (CRP-Chip), Senso s 17 (2017)
684, h ps://doi.o g/10.3390/s17040684.
[18] M.S. Ka ai, L.F. de Lima, W.R. de A aujo, A disposable and low-cos lase -sc ibed
g aphene elec ochemical senso o simul aneous de ec ion o hyd oquinone,
pa ace amol and me hylpa aben, Ma e . Le . 330 (2023) 133211, h ps://doi.o g/
10.1016/j.ma le .2022.133211.
[19] M. S isa-A , K.E. Boehle, B.J. Geiss, C.S. Hen y, Highly sensi i e de ec ion o
Salmonella yphimu ium using a colo ime ic pape -based analy ical de ice
coupled wi h Immunomagne ic sepa a ion, Anal. Chem. 90 (2018) 1035–1043,
h ps://doi.o g/10.1021/acs.analchem.7b04628.
[20] S. Ka i a, T. Kane a, Chela e i a ions o Ca2+and Mg2+using mic o luidic pape -
based analy ical de ices, Anal. Chim. Ac a 924 (2016) 60–67, h ps://doi.o g/
10.1016/j.aca.2016.04.019.
[21] S.A. Noguei a, A.D. Lemes, A.C. Chagas, M.L. Viei a, M. Talha ini, P.A.O. Mo ais,
W.K.T. Col o, Redox i a ion on oldable pape -based analy ical de ices o he
isual de e mina ion o alcohol con en in whiskey samples, Talan a 194 (2019)
363–369, h ps://doi.o g/10.1016/j. alan a.2018.10.036.
[22] N. Tap ab, Y. Sameenoi, Rapid sc eening o o maldehyde in ood using pape -
based i a ion, Anal. Chim. Ac a 1069 (2019) 66–72, h ps://doi.o g/10.1016/j.
aca.2019.03.063.
[23] O. Heida y, M. Akhond, B. Hemma eenejad, A mic o luidic pape -based analy ical
de ice o iodome ic i a ion o asco bic acid and dopamine, Mic ochem. J. 182
(2022) 107886, h ps://doi.o g/10.1016/j.mic oc.2022.107886.
[24] A. Tep ek, V. Poe i A ono, W. Waiyawa , A. Limsakul, J. Shiowa ana,
A. Si ipinyanond, Semi-quan i a i e analysis by spo coun ing on o igami pape -
based de ice o endpoin de ec ion in i ime ic analysis, Mic ochem. J. 158
(2020) 105284, h ps://doi.o g/10.1016/j.mic oc.2020.105284.
[25] O.D. Oyewunmi, S.H. Sa iabadi-Tali, S. Jahanshahi-Anbuhi, Dual-modal assay ki
o he quali a i e and quan i a i e de e mina ion o he o al wa e ha dness using
a pe manen ma ke ab ica ed mic o luidic pape -based analy ical de ice,
Chemosenso s 8 (2020) 1–19, h ps://doi.o g/10.3390/chemosenso s8040097.
[26] E. Ghohes ani, J. Tashkhou ian, B. Hemma eenejad, Colo ime ic de e mina ion o
pe oxide alue in ege able oils using a pape based analy ical de ice, Food Chem.
403 (2023) 134345, h ps://doi.o g/10.1016/j. oodchem.2022.134345.
[27] F.S. Wagne , U.B. S a , Ace ic acid, Ki k-O hme Encycl, Chem. Technol. (2014)
1–21.
[28] C.W. Ho, A.M. Lazim, S. Faz y, U.K.H.H. Zaki, S.J. Lim, Va ie ies, p oduc ion,
composi ion and heal h bene i s o inega s: a e iew, Food Chem. 221 (2017)
1621–1630, h ps://doi.o g/10.1016/j. oodchem.2016.10.128.
[29] P. Gim´
enez-G´
omez, M. Gu i´
e ez-Capi ´
an, J.M. Ríos, F. Capde ila, A. Puig-Pujol,
C. Jim´
enez-Jo que a, Mic oanaly ical low sys em o he simul aneous
de e mina ion o ace ic acid and ee sul u dioxide in wines, Food Chem. 346
(2021) 128891, h ps://doi.o g/10.1016/j. oodchem.2020.128891.
[30] F. Benmeziane De adji, N. Ghanemi, L. Dje moune A koub, Compa ing
physicochemical, nu i ional and biochemical p ope ies o h ee adi ional ig
inega s p oduced in Alge ia, Ca pa hian J. Food Sci. Technol. 15 (2023) 147–158,
h ps://doi.o g/10.34302/c pj s /2023.15.2.13.
[31] K.M.J.P. Boua enin, K.A. Kouam´
e, M.E. Gueu-Kehi, N.T. Dj´
eni, K.M. Dj`
e, O ganic
p oduc ion o inega om mango and papaya, Food Sci. Nu . 9 (2021) 190–196,
h ps://doi.o g/10.1002/ sn3.1981.
[32] R.M. Ca aballo, L.M. Saleh Medina, S.G.J. Gomez, P. Vensaus, M. Hame , Tu me ic
and RGB analysis: a low-cos expe imen o eaching acid-base equilib ia a home,
I. Be asa e e al.
Talan a 276 (2024) 126254
9
J. Chem. Educ. 98 (2021) 958–965, h ps://doi.o g/10.1021/acs.
jchemed.0c01165.
[33] S. ang Supha oek, K. Ponhong, W. Si iangkhawu , K. G udpan, Employing na u al
eagen s om u me ic and lime o ace ic acid de e mina ion in inega sample,
J. Food D ug Anal. 26 (2018) 583–590, h ps://doi.o g/10.1016/j.
j da.2017.06.007.
[34] S. Kasemsum an, A. Boondaeng, S. Jung hee apanich, K. Ngowsuwan,
W. Apiwa anapiwa , P. Janchai, P. Vai hanomsa , Assessing e men a ion b o h
quali y o pineapple inega p oduc ion wi h a nea -in a ed ibe -op ic p obe
coupled wi h s abili y compe i i e adap i e eweigh ed sampling, Molecules 28
(2023) 6239, h ps://doi.o g/10.3390/molecules28176239.
[35] M. Mo mino, V. Siewe s, Y. Nygå d, De elopmen o an Haa1-based biosenso o
ace ic acid sensing in Saccha omyces ce e isiae, FEMS Yeas Res. 21 (2021)
oab049, h ps://doi.o g/10.1093/ emsy / oab049.
[36] G. Che, D. Gu, W. Yang, M. Li, Q. Pan, Tu n-on luo escence de ec ion o ace ic acid
in wine using a u anyl-o ganic amewo k, C ys . G ow h Des. 22 (2022)
1984–1990, h ps://doi.o g/10.1021/acs.cgd.2c00016.
[37] S.T. Alemneh, S.A. Emi e, M. Jekle, B. Hi zmann, E ec o e ige a ed s o age on
some physicochemical cha ac e is ics o a e -based e men ed be e age and he
iabili y o he e men ing Lac iplan ibacillus plan a um and Lac icaseibacillus
hamnosus used, J. Food P ocess. P ese . 46 (2022) e17034, h ps://doi.o g/
10.1111/j pp.17034.
[38] Z.B. Güzel-Seydim, G. S
¸a ı , Ç. G¨
okı maklı, Use o Manda in and pe simmon ui s
in wa e ke i e men a ion, Food Sci. Nu . 11 (2023) 5890–5897, h ps://doi.o g/
10.1002/ sn3.3561.
[39] A. Liu, W. Pan, S. Li, J. Li, Q. Li, L. He, S. Chen, K. Hu, X. Hu, G. Han, S. Li, J. Zhou,
F. Chen, S. Liu, Seasonal dynamics o mic obio a and physicochemical indices in
he indus ial-scale e men a ion o Sichuan Baoning inega , Food Chem. X. 16
(2022) 100452, h ps://doi.o g/10.1016/j. ochx.2022.100452.
[40] S.H. Mohamed, A.I. Salim, Y.M. Issa, A.E. Ali, De ec ion and iden i ica ion o
adul e a ion in inega samples based on e e sed-phase high-pe o mance liquid
ch oma og aphic (RP-HPLC) s a egies, ACS Food Sci. Technol. 2 (2022) 21–30,
h ps://doi.o g/10.1021/acs oodsci ech.1c00169.
[41] S. Huang, S. Ying Lim, H. Lau, W. Ni, S. Fong Yau Li, E ec o glycinebe aine on
me aboli e p o iles o cold-s o ed s awbe y e ealed by 1H NMR-based
me abolomics, Food Chem. 393 (2022) 133452, h ps://doi.o g/10.1016/j.
oodchem.2022.133452.
[42] N.S. Oli ei a, A.F. Oli ei a, A.A. Ne es, M.E.L.R. Quei oz, De elopmen o a non-
i a ion po en iome ic me hod o he de e mina ion o acidi y in inega , Talan a
256 (2023) 124261, h ps://doi.o g/10.1016/j. alan a.2023.124261.
[43] J. Huang, L. Cai, A comp ehensi e expe imen o aining unde g adua es
majo ing in nu i ion educa ion using a pape -based mic o acid-base i a ion,
J. Chem. Educ. 99 (2022) 3607–3612, h ps://doi.o g/10.1021/acs.
jchemed.2c00211.
[44] S. Ka i a, T. Kane a, Acid −base i a ions using mic o l uidic pape -based
analy ical de ices, Anal. Chem. 86 (2014) 12108–12114, 10.1021/ac5039384.
[45] L.G.A. Dias, L.C. Dua e, K.M.P. Pinhei o, N.S. Mo ei a, W.K.T. Col o, Dis ance-
based pape mic o luidics as en i onmen ally iendly pla o ms o moni o ing
acid-base i a ions, Talan a Open 7 (2023) 100216, h ps://doi.o g/10.1016/j.
alo.2023.100216.
[46] Guidelines o S anda d Me hod Pe o mance Requi emen s, 2016, h ps://doi.
o g/10.1093/9780197610145.005.006.
[47] H.C. Goicoechea, A.C. Oli ie i, Sus ained p edic ion abili y o ne analy e
p ep ocessing me hods using educed calib a ion se s. Theo e ical and
expe imen al s udy in ol ing he spec opho ome ic analysis o mul icomponen
mix u es, Analys 126 (2001) 1105–1112, h ps://doi.o g/10.1039/b100422k.
[48] A.G. Gonz´
alez, M.A. He ado , A.G. Asue o, In a-labo a o y es ing o me hod
accu acy om eco e y assays, Talan a 48 (1999) 729–736, h ps://doi.o g/
10.1016/S0039-9140(98)00271-9.
[49] F. Pena-Pe ei a, W. Wojnowski, M. Tobiszewski, Ag ee - analy ical GREEnness
me ic app oach and so wa e, Anal. Chem. 92 (2020) 10076–10082, h ps://doi.
o g/10.1021/acs.analchem.0c01887.
[50] S.A. Noguei a, L.R. Sousa, N.K.L. Sil a, P.H.F. Rod igues, W.K.T. Col o,
Moni o ing acid-base i a ions on wax p in ed pape mic ozones using a
sma phone, Mic omachines 8 (2017) 1–10, h ps://doi.o g/10.3390/mi8050139.
[51] P. Namwong, P. Ja ujam us, M. Ama a ongchai, S. Chai am, Fab ica ing simple
wax sc een-p in ing pape -based analy ical de ices o demons a e he concep o
limi ing eagen in acid-base eac ions, J. Chem. Educ. 95 (2018) 305–309, h ps://
doi.o g/10.1021/acs.jchemed.7b00410.
[52] J. Li, M.L. O’Neill, C. Pa ison, J.H.W. Zhou, J.M. I o, C.S.T. Wong, H.Z. Yu,
N. Me bouh, Mobile app o quan i y pH s ips and moni o i a ions: sma phone-
aided chemical educa ion and class oom demons a ions, J. Chem. Educ. 100
(2023) 3634–3640, h ps://doi.o g/10.1021/acs.jchemed.3c00227.
I. Be asa e e al.
