Electrochemical Analysis of Coffee Extractions at ...

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Electrochemical Analysis of Coffee Extractions at Different Roasting

Levels Using a Carbon Nanotube Electrode

Ryotaro WADA,* Shota TAKAHASHI,* Hitoshi MUGURUMA,*† and Naomi OSAKABE,**

* Graduate School of Engineering and Science, Shibaura Institute of Technology, 3-7-5 Toyosu,

Koto, Tokyo 135-8548, Japan

** Department of Bioscience and Engineering, College of Systems Engineering and Science,

Shibaura Institute of Technology, 307 Fukasaku, Minuma, Saitama 337-8570, Japan

† To whom correspondence should be addressed.

E-mail:[email protected]

Analytical SciencesAdvance Publication by J-STAGEReceived July 31, 2020; Accepted August 31, 2020; Published online on September 11, 2020DOI: 10.2116/analsci.20N021

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Abstract

In this study, the electrochemical analysis of coffee extractions at different roasting levels by

using a carbon nanotube (CNT) electrode was reported. The roasting levels, ranging from 1

(low) to 6 (high), were determined according to the roasting time after fixing the roasting

temperature. Level 1 roasting resulted in light roasted beans and level 6 in dark roasted ones.

Based on the roasting level, the concentration of chlorogenic acids, including 3-caffeoylquinic

(3CQ), 4-caffeoylquinic (4CQ), and 5-caffeoylquinic (5CQ) acid, can be determined. Cyclic

voltammetry (CV) experiments revealed that the reduction current at +0.27 V was proportional

to the concentration of chlorogenic acids. High-performance liquid chromatography (HPLC)

revealed that the inverse correlation between the roasting level and chlorogenic acid amount.

The total amounts of chlorogenic acids in coffee extractions determined by HPLC were in

agreement with those obtained by CV using the CNT electrode at roasting levels 1–5. At level 6,

the amount of chlorogenic acids determined by the current peak was larger than that detected by

HPLC. As a result, the chlorogenic acid amount was overestimated in the CV experiment at

+0.27 V, indicating that electrochemically active materials were generated at level 6. The CV

profile showed that the reduction peak at +0.10 V increased with increasing roasting level. Thus,

the peak intensity at +0.10 V can be used to evaluate the roasting levels even if the

concentration or dilution conditions were not provided.

Keywords carbon nanotube, cyclic voltammetry, coffee infusion, roasting level, chlorogenic

acid, redox reaction.


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Introduction

Chlorogenic acid is the major component of coffee polyphenols. More than 40

compounds have been identified as chlorogenic acids, including 3-caffeoylquinic acid (3CQ),

4-caffeoylquinic acid (4CQ), and 5-caffeoylquinic acid (5CQ), whose chemical structures are

shown in Figure 1. These three acids represent over 90% of the total chlorogenic acids identified

in coffee by high-performance liquid chromatography (HPLC).1-7 The total amounts of 3CQ,

4CQ, and 5CQ in coffee extractions can be quantified by electrochemical measurement using a

carbon nanotube (CNT) electrode.7 Furthermore, we reported the quantification of coffee

polyphenols in real coffee extract (so-called a couple of coffee) such as drip, instant (soluble),

and PET-bottle coffees.7 Roasting is the most important step in the coffee processing because it

determines the taste and aroma of coffee extractions. The relationship between roasting and the

amount of chlorogenic acids determined by HPLC has been extensively studied.1-5 However,

HPLC-based methods have disadvantages such as bulky apparatus, time consumption,

requirement of trained personnel, and consumption of harmful organic solvents. The CNT

electrodes8 have traditionally been used for chlorogenic acid detection in coffee sample.9-12

Therefore, the CNT electrode for chlorogenic detection is thought to be promising in terms of

highly electrochemical functionality, such as catalytic effects and a reduction of the oxidation

potential. However, those were the standard addition methods, which were not strictly “real”

sample analysis.9-12 Furthermore, those reports did not provided the comparison between HPLC

and electrochemical method. It has not been done on electrochemical analysis about the roasting

level. Our challenge in this study is to determine the roasting level of coffee by electrochemical

analysis using the CNT electrode.


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Experimental

Chemicals

Chlorogenic acids (3CQ, 4CQ, 5CQ) and carboxymethylcellulose (CMC) were

purchased from Sigma-Aldrich (St. Louis, USA). Multi-walled CNTs (10–30 nm in diameter

and 200 µm in length) were produced by Taiyo Nippon Sanso Corporation (Tokyo, Japan).

Electrode preparation

The CNT electrode preparation was described in our previous studies.13-15 In brief,

gold evaporated onto a polyethylene terephthalate film was used. The cavity for the working

electrode was 9 mm2. The CNTs were dispersed in a 0.3% CMC aqueous solution to attain a

concentration of 0.1%. Aliquots of 5 µL of CNT-CMC solution were dropped onto the gold

surface and were dried. Then, 5 µL aliquots of 1.0% (w/v) CMC solution were dropped onto the

CNT-CMC layer and allowed to dry.

Coffee sample preparation

Green coffee arabica beans were purchased from retail stores in Japan. The base

coffee blend was composed with beans originally from Peru. A total of 200 g of green beans

were roasted in duplicates using a commercial air stream roaster operating at 210 ºC. The

roasting levels were controlled by the roasting times ranging from 5 to 15 min and were denoted

as follows: level 1 (5 min), 2 (7 min), 3 (9 min), 4 (11 min), 5 (13 min), and 6 (15 min). Level 1

roasting resulted in light roasted beans and level 6 in dark roasted ones. The roasted beans were

ground to a sieve size of ~1 mm (rough). The extraction conditions were as follows: 10 g of

ground beans was added to 100 mL of water at 90 ºC for 1 min in a paper filter.

Measurement

Electrochemical measurements were performed with an electrochemical analyser

(ALS Instruments, 701A West Lafayette, IN) using a three-electrode configuration. Reference

(Ag/AgCl, RE-1C) and counter (platinum wire) electrodes were purchased from BAS Inc.


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(Tokyo, Japan). For electrochemical measurements, citric acid buffer (50 mM, pH 6.0) was used

as the supporting electrolyte in a 5 mL vessel at 20 °C, and 0.2 mM stock chlorogenic acids

solutions were successively added to prepare samples with desired concentrations. The HPLC

system (Shimadzu LC-20AD. Kyoto, Japan) consisted of a quaternary pump, a vacuum degasser,

an automated sample injector, a column oven, a system controller, and a diode array UV

detector set at 325 nm. The separation column was a reversed-phase column (4.6 mm in

diameter, 150 mm in length, Intertsil ODS-3, Kyoto, Japan) with a 3-µm nominal particle size.

The mobile phase included mixture of methanol and 0.1% phosphate buffer. The flow rate was

1.0 mL min-1, injection volume 1.0 µL, and column temperature 40 °C.

Simulation

CV simulations were performed using the DigiElch 8 software package (Elchsoft,

Germany). Butler–Volmer kinetics were used to estimate the electron transfer parameters under

the semi-infinite one-dimensional diffusion condition. The cell IR drop was not compensated

during the measurements, and the adsorption was neglected in the simulations. The transfer

coefficient, a, was set to 0.5.

Results and Discussion

The HPLC chromatograms of the coffee extractions at different roasting levels are

shown in Figure 2. Three major peaks attributed to 3CQ, 4CQ, and 5CQ were observed in the

chromatograms. The other peaks identified in the chromatograms were smaller than the three

major peaks, indicating that 3CQ, 4CQ, and 5CQ were the major chlorogenic acids in the coffee

extractions. Moreover, the amount of the other chlorogenic acids were negligible in real coffee

extractions, which is consistent with previous reports.1-5 Therefore, the amount of chlorogenic

acids in the coffee extractions was the total amount of 3CQ, 4CQ, and 5CQ.

The CV profile of 5CQ (Figure 3A) shows the anodic and cathodic peaks at Ea = +0.293

V and +0.265 V attributed to the redox oxidation of catechol to quinone, the reaction scheme is


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shown in Figure 1. The estimated peak-to-peak potential separation (DEp) was 28 mV, and the

relationship between DEp and the electron number, n, is represented by the following equation:

DEp =59/n (mV) (1)

The calculated value of n was 2.1, which is consistent with the two-electron redox reaction of

the 5CQ catechol group (Figure 1), suggesting a reversible system. The CVs of 3CQ and 4CQ

(Figure S1) were similar to those of 5CQ. This means that the discrimination of the three

isomers due to differences in the position of ester link is not possible, but the total amounts of

3CQ, 4CQ, and 5CQ can be determined. The overlapping dot line in Figure 3A represents the

CV simulation of 5CQ using the proposed redox reaction shown in Figure 1. The determined

parameters were Eo = +0.28 V and heterogeneous electron-transfer rate constants ko = 0.2 cm s-1,

where Eo corresponds to the midpoint of Ea and Ec of 5CQ (Figure 3A), and ko is obtained by

fitting the experimental and simulated results to the Butler–Volmer equation. The value of ko

is similar with the reported value of catechol compounds.16,17 The simulated profile

adequately represented the experimental result around at the potential range of the redox peak

(+0-0.4 V). At the higher potential range (>+0.4 V), the oxidation current in the simulation is

larger than that of the experiment. It is probably attributed to the effect of adsorption.

Figures 3B-G show the CVs of coffee extractions at different roasting levels. The CV

profiles of levels 1 and 2 were similar to those of 5CQ, indicating that the coffee components at

levels 1 and 2 were mostly chlorogenic acids. The CV profiles demonstrated that higher roasting

levels resulted in wider peaks. Conversely, the reduction current at +0.10 V increased with

increasing roasting level (see arrows in Figures 3D-G).

Figure 4A shows the relationship between the roasting level and amount of chlorogenic

acids in the coffee extractions. The higher the roasting level, the lower the amount of

chlorogenic acid—this is a result of the decomposition and conversion of chlorogenic acids to

other chemicals at high roasting temperature.1-5 The concentration ratios of 3CQ, 4CQ, and

5CQ were mostly unchanged during roasting (Table S1). That is, the three chlorogenic acids


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were equally decomposed during the roasting process. The coffee extractions at level 1, using

coffee beans of different origins (Kenya, Jamaica, Colombia, and Brazil), contained similar

amounts of chlorogenic acids as those shown in Figure 1. The variation was within 8%,

indicating that different types or origins of coffee beans were used in this study. The current

ratio of ‒Ic(+0.10 V)/Ic(+0.27 V) accurately correlated with the roasting levels, as shown in

Figure 4B. This process was empirically determined. Therefore, the peak intensity at +0.10 V

can be used to evaluate the roasting level even if the concentration or dilution conditions were

not provided. The reduction peak at +0.10 V was attributed to an electrochemically active agent

with a chemical structure different from that of the chlorogenic acid generated in the roasting

process.

Figure 5 shows the plot of chlorogenic acid concentrations in real coffee extractions vs.

CV reduction currents (+0.27 V) of coffee extractions at different roasting levels and dilutions.

The dilution values were set to be in the linear range of 5CQ (0.020‒6.5 mg/dL), and the solid

line (y = ‒0.42 ‒ 3.96x) was originated from 5CQ. For example, the fitted plot indicated that the

chlorogenic acid concentration was 72 mg/dL for the coffee extraction at level 1. All plots were

close to the solid line of 5CQ, except for level 6. Thus, up to level 5, a strong correlation was

observed between the HPLC results and CV current measured using the CNT electrode. At

level 6, the chlorogenic acid concentration was overestimated by the CNT-electrode-based CV,

compared to that measured by HPLC. These results suggested that the reduction peak intensity

of coffee extractions at level 6 was originated from electrochemically active non-chlorogenic

acids, while those of level 1 to 5 from chlorogenic acids.

Conclusions

The electrochemical analysis of coffee extractions at different roasting levels using a

CNT electrode was reported. The reduction current at +0.27 V and +0.10 V can be used to


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evaluate the roasting levels of coffee beans. The current intensity at +0.27 V was proportional to

the amount of chlorogenic acids, which was correlated with the roasting levels except for the

highest level (level 6). The reduction peak at +0.10 V was also correlated with the roasting level,

and it can be used to evaluate the roasting levels even if the concentration or dilution conditions

were not provided.

Acknowledgments

We would like to thank Editage (www.editage.com) for English language editing.

Supporting Information

CV of 3CQ and 4CQ, and concentrations of individual chlorogenic acid of coffee extractions at

different roasting level by HPLC measurement. This material is available free of charge on the

Web at http://www.jsac.or.jp/analsci/.


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References

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Figure Captions

Figure 1. Molecular structures and redox reaction of chlorogenic acids.

Figure 2. HPLC chromatograms of real coffee extractions at different roasting levels.

Figure 3. (A) Experimental and simulated CVs of 5CQ using CNT electrode. Concentration was

4.9 mg/dL. (B-G) CV of coffee extractions at different roasting levels. Backgrounds are

subtracted. The diluted ratios are: (B) 41, (C) 41, (D) 14, (E) 14, (F) 11, and (G) 11. Citric acid

buffer solution (50 mM, pH 6.0) was used as electrolyte. Scan rate was 0.05 V s-1. Backgrounds

were subtracted.

Figure 4. (A) Plot of roasting levels vs. concentration of chlorogenic acid. (B) Plot of roasting

levels vs. current ratio between the potentials at +0.10 V and +0.27 V.

Figure 5. Plot of CV current vs. diluted concentrations obtained from the HPLC data in Figure 2.

The fitting line was obtained from the CV current of 5CQ.


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Figure 1

OOH

OH

OHHO O

O

OH

OH

-2H+/-2e-E°k°

R

O

O

5-Caffeoylquinic acid (5CQ)



O

OH

OHOH OH

OHO OOH

OHOH

O

OHHO O

O

OH

OH


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Figure 2

5CQ

3CQ

4CQ

1

×20×20

Time / min.

10 mV

2

Roast level

3

45

6

0 5 10 15 20


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Figure 3

-0.2 0.0 0.2 0.4 0.6 0.8 1.0

5 µA

Potential / V

Roast level1

2

3

4

5

6

Coffee infusion

Experiment

Simulation5CQ

5 µA

A

B

C

D

E

F

G


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Figure 4

1 2 3 4 5 60

20

40

60

80

0.0

0.2

0.4

0.6

0.8

1.0

Roast level

Chl

olog

enic

acid

co

ncen

tratio

n / m

g/dL

I c(+0

.10

V)/I

c(+0.

27 V

)

B

A


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Figure 5

0 1 2 3 4 5 6 7-30

-20

-10

0

y=‒0.42‒3.96x

Cur

rent

/ µA

Chlorogenic acids concentration / mg/dL

12

Roast level

3456


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Graphical Index

Roast level

1 2 3 4 5 6Low High


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