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BY 4.0 license Open Access Published by De Gruyter Open Access October 30, 2023

Determination of caffeine content in coffee drinks prepared in some coffee shops in the local market in Jeddah City, Saudi Arabia

  • Saad M. Al-Baqami , Torki A. AlZughabi , Mohammad A. Almostady , Nourah A. Alotaibi , Amal Almostadi , Tahreer M. AlRaddadi and Saleh O. Bahaffi EMAIL logo
From the journal Open Chemistry

Abstract

The current study was aimed to develop a low cost HPLC method for determination of caffeine content in coffee samples in some coffee shops in Jeddah local market. Caffeine was extracted with water and separated through ZORBAX Eclipse XDB C-18 column using methanol/water (40:60) as mobile phase. The average peak response time for caffeine was 2.747 min with a relative standard deviation of 1.061. The proposed assay was able to detect caffeine in the concentration range from 5 to 45 ppm with limits of detection (LOD = 3σ/b) and quantification (LOQ = 10σ/b) of 1.086 and 3.619 ppm using two-channel UV detector set at 274 and 264 nm with correlation coefficient (R 2) of 0.9996, respectively. The assay was adopted for the determination caffeine content in 30 coffee samples collected from eight brand coffee shops and from local market. Each sample was injected in triplicate (n = 3) and the average and confidence limit of the mean were calculated. The concentration of caffeine in coffee brand shops varied from 513.82 ± 12.91 to 8080.05 ± 3.06 ppm, whereas in beans and instant coffee its content was diverse from 360.18 ± 86.72 to 1306.98 ± 27.31 ppm. These results were validated using Student’s t-test at 95% confidence level.

1 Introduction

Coffee is one of the greatest common beverages in the world [1,2]. The most widely cultivated species of coffee are Coffea arabica and Coffea robusta, which supply almost all of the world’s consumption [2]. The world production of green coffee in 2021 was 175.35 million (60 kg/bags) and more than 17.89 million tons of coffee beans in Saudi Arabia has been imported with a value of $127.18 million [3]. Nowadays, there is a growth in coffee shops and high consumption in coffee drink in Saudi Arabia; however, there is great alarm on the impact of having coffee for the public health [4,5]. Caffeine (1,3,7-trimethylxanthine) is an alkaloid that occurs naturally in the leaves, seeds, fruit, tea, coffee, cocoa, kola trees, and more than 60 other plants [6]. Caffeine drug is used to provide a boost of energy or a feeling of heightened alertness and is often used to stay awake longer [79]. A pleasant stimulant feeling is obtained by consuming low dose of caffine [10].

Caffeine consumption at high levels have been a concern in various disorders such as increase of gastric-acid secretion, kidney malfunction, heart disease (cardiac arrhythmia), and disturbances of the central nervous system such as seizures and delirium [11,12]. On the other hand, extraordinary consumption of caffeine leads to rhabdomyolysis which damages muscle fiber and enter the blood stream, increase the incidence of bladder and stomach cancer, increases blood pressure, aggravates diabetes, and damages the lining of the stomach [12,13]. Thus, there is a great request for precise determination of caffeine compounds in soft drinks and energy drinks for assurance of food safety, and quality control is mandatory.

Recently, numerous techniques have been reported for trace determination of caffeine in various types and drinks of coffee samples including voltammetry [14], NIR spectroscopy [15], UV/visible spectrometry [16], chromatographic methods [17], gas chromatography (GC)-FID [18], GC-NP detector [19], GC-mass spectrometry [20], and high performance liquid chromatography (HPLC) [2123].

In today’s era, there is a increase in coffee shops and high consumption in coffee drink in Saudi Arabia; therefore, the current study was designed to (i) precisely assess a low cost HPLC method for caffeine trace determination using HPLC in coffee water extract, (ii) sorting of 30 commercial coffee samples available in the various commercial coffee drink brands available in the local coffee shops in Jeddah city, KSA with respect to caffeine in the coffee extract in boiled water, and finally (iii) studying the relationship between the average content of caffeine in coffee samples with the prices in Saudi Arabian market.

2 Experimental

2.1 Materials and methods

Caffeine anhydrous ≥99.0% HPLC grade was purchased from Fluke Chem. Division, San Diego, CA, USA. Methanol for HPLC ≥99.9% grade was provided from Sigma-Aldrich Chemicals Private Limited, Bangalore, India. All measurements were performed in triplicate. The sample was filtered through Whatman No. 1 filter paper, and then again filtered through Millipore 0.45 µm before injection in the HPLC instrument.

2.2 Instrumentation

HPLC analysis was performed using UFLC Shimadzu at King Fahad Medical Research Center equipped with control processing unit 20A, solvent delivery pump LC 20 AD, degassing unit 20As, photodiode array detector, pump mode: low pressure gradient LC 20 DA at a flow rate of 1.0 mL/min. pressure max 15 MPa. UV–VIS detector SPD-20A equipped with deuterium lamp D2, wavelength length 1 at 274 nm, and wave length 2 at 264 nm were used. Autosampler SIL20AC sample Rack 1.5 mL 70 vials, oven CTO-20A, temperature adjusted at 25°C, and temperature control cell SPD-20A were also used. A ZORBAX Eclipse XDB C-18 analytical column of 4.6 × 150 mm 5 μm (Agilent 5301 Stevens Creek Blvd. Santa Clara, CA, USA) was used at a flow rate of 1.0 mL/min. Milli-Q plus system (Millipore, Bedford, MA, USA) water system was used for providing ultra-pure water for the preparation of the solutions.

2.3 Sampling

Thirty coffee samples were collected from the local market of Jeddah city and classified as follows: eight different brand shops (24 sample), and six samples Harare, Lagmti, Kholani, Katamano, Edition signature blend, and Cofiq Prive were purchased from Al Rashid store for dates and coffee, Abdullah Alsulymani Street, Jeddah, KSA. Coffee brands were taken in cups, cooled, and filtered before injection, coffee powder was prepared in boiling water as traditional Saudi coffee. The other three fast preparing coffee brands were purchased from local market. All samples are filtered through Whatman No. 1 and then again filtered using Millipore 0.45 µm filter paper before injection in the HPLC Autosampler, except Expresso coffee that was diluted ten times before injection. All measurements were injected in triplicate in an autosampler with 5.0 µL injected in HPLC injection volume.

2.4 Recommended standard solution preparation

Caffeine stock standard solution of 1,000 µg/mL was prepared by dissolving 0.10 g of caffeine standard (Sigma-Aldrich) in 80 mL deionized water and sonicated for 10 min. The solution was then transferred to 100 mL volumetric flask and the flask was filled up to the mark by deionized water. Working standard solutions of 5, 10, 15, 20, 25, 30, 35, 40, 45 µg/mL were prepared by serial dilution of the stock solution with deionized water. A blank solution was prepared as same as the standard without caffeine in it. Each concentration was injected in triplicate and an external calibration curve of the average peak areas (n = 3) versus concentration of the standards was plotted. The results are presented in the calibration plot (Figure 1) for standard preparations of standard caffeine.

Figure 1 
                  Calibration curve for caffeine determination.
Figure 1

Calibration curve for caffeine determination.

3 Results and discussion

The calibration graph in Figure 1 shows a strong positive correlation between the instrumental signal and the standard caffeine concentrations. Ten different concentrations from 5 to 45 µg/mL including the blank were precisely analyzed, which represent the available caffeine in the coffee product. The calibration graph was plotted using the caffeine concentration in the standard against the detector response (peak area). The calibration plot of various caffeine concentrations was found to be linear with excellent correlation coefficient (R 2 = 0.9996). The limits of detection (LOD) and quantification (LOQ) were calculated based on the standard deviation of the response (Sy) of the curve and the slope of the calibration curve (S) at levels approximating the LOD according to the formula: LOD = 3.3σ/b and LOQ = 10σ/b, where σ is the standard deviation of the blank and b is the slope of the calibration plot. The standard deviation of the response was determined based on the standard deviation of y-intercepts of regression lines. The calculated values of LOD and LOQ were found to be equal to 1.086 and 3.619 µg/mL, respectively, with a sensitivity factor of 15,870. Table 1 summarizes the regression data and statistical parameters for the designed calibration plot of caffeine (Figure 2).

Table 1

Regression data and statistical parameters for caffeine calibration curve

Correlation coefficient (R 2) Slope (S) Y-intercept Linear range (ppm)
0.9996 15,870 768.29 5–45
Figure 2 
               Chromatogram of 5 ppm caffeine at 274 nm.
Figure 2

Chromatogram of 5 ppm caffeine at 274 nm.

The data of average concentrations (μg/g) of caffeine in coffee brand shops are summarized in Table 2 and the representative chromatograms are illustrated in Figures 3 and 4. On the other hand, caffeine concentration (µg/g) in coffee beans and instant coffee (μg/g) is summarized in Table 3. The distribution pattern of caffeine concentration (µg/g) in various coffee beans and instant coffee is illustrated in Figure 5. Caffeine contents in the collected samples varied widely according to brands’ type and the selected coffee shops in the current study. Overall, based on the caffeine content, the results can be categorized into two categories and can be arranged as follows:

Table 2

Caffeine concentration (ppm) in coffee brand shops

Number Coffee shop Concentration (ppm)
Black & V 60 Americano Espresso
1 LA Cima 937.61 ± 4.16 1030.26 ± 9.55 7701.54 ± 6.99
2 Brew 92 971.52 ± 21.18 854.49 ± 7.13 5801.12 ± 11.79
3 Cloud 9 956.54 ± 14.01 1093.71 ± 9.99 4852.15 ± 23.31
4 Starbucks 513.82 ± 12.91 819.67 ± 1.39 2488.87 ± 7.12
5 Ratio 1081.71 ± 4.20 1319.43 ± 4.78 8080.05 ± 3.06
6 Costa 725.16 ± 3.26 485.56 ± 4.50 1366.29 ± 5.16
7 Kims 821.69 ± 21.89 557.16 ± 7.38 5647.89 ± 1.68
8 Dose 723.23 ± 2.42 516.54 ± 6.46 839.686 ± 20.12
Figure 3 
               Chromatogram obtained for one of the injections of one coffee sample.
Figure 3

Chromatogram obtained for one of the injections of one coffee sample.

Figure 4 
               Caffeine concentration in coffee drinks in different brand shops.
Figure 4

Caffeine concentration in coffee drinks in different brand shops.

Table 3

Caffeine concentration in coffee beans and instant coffee

No Coffee brand Concentration (ppm)
1 Harare 1264.20 ± 13.21
2 Lagamti 1306.98 ± 27.31
3 Kholani 895.37 ± 12.18
4 Catamona 360.18 ± 86.72
5 Edition sig. blend 583.61 ± 28.00
6 Cofique Prive 530.77 ± 33.68
Figure 5 
               Caffeine concentration (ppm) in coffee beans and instant coffee.
Figure 5

Caffeine concentration (ppm) in coffee beans and instant coffee.

The first category of selected coffee shops:

LA Cima Espresso > Americano > Black & V60

Cloud Espresso > Americano > Black & V60

Starbucks Espresso > Americano > Black & V60

Ratio Espresso > Americano > Black & V60

The second category of coffee brands:

Brew 92 Espresso > Black & V60 > Americano

Costa Espresso > Black & V60 > Americano

Kims Espresso > Black & V60 > Americano

Dose Espresso > Black & V60 > Americano

The concentration of caffeine was set from high to low Lagmti > Harari > Kholani, whereas in instant coffee, the caffeine concentration is highest in the Edition signature blend, followed by the Confique Prive and Catamona.

4 Conclusion and future perspectives

In summary, the current study reported the levels of caffeine in various commercial products of coffee beans, instant coffee, and coffee shop drinks and provides data on the spectrum of caffeine intake levels by human. Caffeine in coffee can enrich a diet with microelements that have antioxidant activity. The level of caffeine content in eight different brand shops varied, where high levels were found in the espresso in all coffee brand shops while Americano and Black & V60 share the arrangement. In the traditional coffee, Lagmti has the highest followed by Harari and Kholani, whereas in instant coffee, the Edition signature blend has the highest concentration of caffeine followed by Confique Prive and Catamona the last. Thus, there is a need for more research that would also take into account the content of caffeine in the water used to prepare infusions, as well as their bioavailability for humans. Further research and exploration are merited for the possible physical dependence potential of low doses of caffeine such as those concentrations found in decaffeinated coffee. A detailed study will be performed to fully assign other microelements and their relation with caffeine in various coffee brands.

Acknowledgements

The authors would like to thank King Fahad Medical Research Center, Main Laboratory and Forensic Laboratory for chromatographic analysis. The authors also thank Prof. Dr Mohammad El-Shahawi for his assistance and guidance. They would also like to thank AFAQ Speciality Coffee and Trad Basonbul (development consultant) for their assistance and guidance.

  1. Funding information: No funding was received for conducting this study.

  2. Author contributions: Saad M. Al–Baqami: formal analysis and writing – first draft; Torki A. AlZughaibi: project co-supervisor and writing – review and editing; Mohammad A. Almostady, Nourah A. AlOtaibi, and Amal A. Almostadi: formal analysis and data procuration. Tahreer M. Alraddadi: formal analysis, validation, and data assignment. Saleh O. Bahaffi: project administration, supervision, and writing – original draft.

  3. Conflict of interest: No conflict of interest among the authors and all authors agreed to the contents in the text.

  4. Ethical approval: This article does not contain any studies with human participants or animals performed by any of the authors.

  5. Data availability statement: The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding authors. The raw data supporting the conclusion of this article will be made available by the authors without undue reservation.

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Received: 2023-07-06
Revised: 2023-09-07
Accepted: 2023-09-15
Published Online: 2023-10-30

© 2023 the author(s), published by De Gruyter

This work is licensed under the Creative Commons Attribution 4.0 International License.

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