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A Detail Chemistry of Coffee and Its Analysis

WRITTEN BY

Hemraj Sharma

Submitted: 20 January 2020 Reviewed: 12 February 2020 Published: 20 March 2020

DOI: 10.5772/intechopen.91725

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Coffee

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Coffee

Production and Research

Edited by Dalyse Toledo Castanheira

FROM THE EDITED VOLUME

Coffee
Edited by Dalyse Toledo Castanheira

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Abstract

This review article highlights the detailed chemistry of coffee including its components; chemical
constituents like carbohydrates, proteins, lipids, and caffeine; aromatic principles; oil and waxes; and
minerals and acids. The high extent of caffeine can be found in the coffee plants; hence, in the second
part of the study, various analytical methods are designed for the proper identification, separation,
optimization, purification, and determination of caffeine present in coffee, tea, and marketed coffee.
These analytical methods are appropriated for the separation and quantification of caffeine. The various
analytical methods include spectroscopy methods like UV, IR, and NMR spectroscopy; chromatographic
methods like paper, TLC, column, HPLC, and gas chromatography; and hyphenated techniques like LC–
MS, GC–MS, and GC–MS/MS. This article compares and contrasts the amount of caffeine by various
analytical methods.

Keywords

caffeinespectrophotometerchromatographyhyphenated techniqueselectrochemical methods

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Chapter sections

1. Introduction
Coffee consists of ripe seeds of Coffea arabica Linn., belonging to family Rubiaceae. Coffee extracted
from coffee bean is also present in crimson fruits is completely removed, and the spermoderm is
removed, occasionally. The seeds of botanical genus Coffea may be raw, roasted, whole, or ground. The
prepared drink through such coffee seeds is also called as coffee. Among 70 species of coffee, only three
are cultivated. 75% of the world’s production of coffee is provided by Coffea arabica, about 25% by
Coffea canephora, and less than 1% by Coffea liberica and others. Generally, coffee is cultivated at the
altitude of 1000–2000 [1]. It is indigenous to Ethiopia, Brazil, India, Vietnam, Mexico, Nepal Guatemala,
Indonesia, and Sri Lanka.

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2. Chemical constituents

The main constituents of coffee are caffeine, tannin, fixed oil, carbohydrates, and proteins. It contains 2–
3% caffeine, 3–5% tannins, 13% proteins, and 10–15% fixed oils. In the seeds, caffeine is present as a salt
of chlorogenic acid (CGA). Also it contains oil and wax [2].

The following sections will be discussed in detail after acceptance of this short proposal:

This article will deal on the types of carbohydrate, protein, lipids, and other chemical constituents in
detail.

This article will review on various analytical methods for the estimation of constituents present in
coffee.

Coffee is often used as antioxidants, but more importantly coffee is a good source of chromium and
magnesium that assist in controlling blood sugar by ensuring proper usage of insulin.

The main chemical ingredients in coffee beans are given below:

Caffeine
Tannin

Thiamin

Xanthine

Spermidine

Guaiacol

Citric acid

Chlorogenic acid

Acetaldehyde

Spermine

Putrescine

Scopoletin

The carbohydrate content of green and roasted coffee (Santos) was identified and measured. Green
coffee contained about 6–7% of sucrose as soluble sugars and low amount of glucose. The soluble
sugars of roasted coffee were sucrose, fructose, and glucose. The experiment was also carried out for
the isolation of holocellulose fractions of green and roasted coffee.
The holocellulose of green coffee was hydrolyzed by a novel method consisting of anhydrous sulfuric
acid and 10% potassium insoluble hydroxide, which was partially solubilized on roasting and results in
the following ratio of sugars:

1 L-arabinose/2D-galactose/2D-glucose/6D-mannose. Out of these sugars, the arabinose was easily acid-

hydrolyzed. Other coffee constituent analyzed and determined were caffeine, trigonelline, caffeic acid,
chlorogenic acid, isochlorogenic acid, and the 10 amino acids. The free amino acids disappeared in
roasting. An analytical method was developed for evaluating caffeine on chromatograms [3].

In coffee pulp, condensed tannins are the major phenolic compounds, while in the seeds, phenolic
compounds exist primarily as a family of esters formed between hydroxycinnamic acids and quinic acid,
collectively recognized as chlorogenic acids (CGA). Green coffee seeds contain up to 14% CGA, which are
present in high concentrations and have a greater influence for determining the quality of coffee and
play a vital role in the formation of the coffee flavor. The various constituents along with components of
coffee are shown in Table 1.

Constituent Components

Soluble carbohydrates Monosaccharides Fructose, glucose, galactose, arabinose (traces)

Oligosaccharides Sucrose, raffinose, stachyose

Polysaccharides Polymers of galactose, mannose, arabinose, glucose

Insoluble polysaccharides

Hemicelluloses Polymers of galactose, arabinose, mannose

Cellulose

Acids and phenols

Volatile acids

Nonvolatile aliphatic acids Citric acid, malic acid, quinic acid

Chlorogenic acids Mono-, dicaffeoyl- and feruloylquinic acid

Lignin

Lipids
Wax

Oil Main fatty acids: N Compounds

Free amino acids Main amino acids: Glu, Asp, Asp-NH2

Proteins

Caffeine Traces of theobromine and theophylline

Trigonelline

Minerals

Table 1.

Constituents along with components of coffee.

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3. Carbohydrates

Most of the carbohydrates present, such as cellulose and polysaccharides consisting of mannose,
galactose, and arabinose, are insoluble.

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4. Lipids

The lipid fraction appears to be very stable, and its composition is given below.

Linoleic acid is the predominant fatty acid, followed by palmitic acid.

Lipid composition.

Triacylglycerols.
Diterpene esters.

Diterpenes.

Triterpene esters.

Triterpenes (sterols).

Unidentified compounds.

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5. Acids

The volatile acids include formic acids and acetic acids, while nonvolatile acids include lactic, tartaric,
pyruvic, and citric acid. Minor constituents include higher fatty acids and malonic, succinic, glutaric, and
malic acids. The degradation products of citric acid are itaconic (I), citraconic (II), and mesaconic acids
(III), while fumaric and maleic acids are degraded products of malic acid:

Chlorogenic acids are the mainly rich acids of coffee.

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6. Trigonelline and nicotinic acid

Green coffee contains trigonelline (N-methylnicotinic acid) up to 0.6% and is 50% decomposed during
roasting. The degradants include nicotinic acid, pyridine, 3-methyl pyridine, nicotinic acid, methyl ester,
and other compounds.

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7. Aromatic principle

The aroma profile of coffee is composed of the following notes: sweet/caramel-like, earthy,
sulfurous/roasty, and smoky/phenolic.

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8. Minerals

Potassium is major in coffee ash (1.1%), calcium (0.2%), and magnesium (0.2%). The major anions
includes phosphate (0.2%) and sulfate (0.1%), along with traces of other elements [4].

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9. Caffeine

The best known N compound is caffeine (1,3,7-trimethylxanthine) because of its physiological effects
(stimulation of the central nervous system, increased blood circulation, and respiration). It is mildly
bitter in taste. 10% of the caffeine and about 6% of the chlorogenic acid are present in a coffee drink.
During roasting, the caffeine level in beans is decreased. Synthetic caffeine and caffeine obtained by the
decaffeination process are used by the pharmaceutical and soft drink industries. By methylation of
xanthine, synthetic caffeine is obtained which is obtained from uric acid and formamide. Medicinally,
caffeine is used as a CNS stimulant, usually combined with another therapeutic agent and in analgesic
preparations.

Theobromine acts as diuretic and smooth muscle relaxant, but not routinely used. Theophylline is used
as smooth muscle relaxant and is frequently dispensed in sustainable formulations to lower the side
effects. It is also available as aminophylline (a more soluble preparation containing theophylline with
ethylenediamine) and choline theophyllinate (theophylline and choline). The alkaloids may be isolated
from natural sources or obtained by total or partial synthesis [5].

The purine alkaloids include caffeine, theobromine, and theophylline as shown in Figure 1. They have a
limited distribution as alkaloids, but the origins are very close with those of the purine bases like
adenine and guanine, fundamental components of nucleosides, nucleotides, and the nucleic acids.
Caffeine is mainly consumed in the form of beverages like tea, coffee, and cola and is most widely
consumed and socially accepted natural stimulants. Theophylline is much more important as a drug
compound because of its muscle relaxant properties, utilized in the relief of bronchial asthma when
compared to caffeine, medicinally. The major constituent of cocoa and related chocolate products is
theobromine.

Figure 1.

Chemistry of the purine derivatives.

Out of four nitrogen atoms, two are supplied by glutamine and a third by aspartic acid. The synthesis of
the nucleotides AMP and GMP is by way of IMP and XMP, and the purine alkaloids then branch away via
XMP. The loss of phosphate via methylation generates the nucleoside 7-methylxanthosine, which is then
released from the sugar moiety. Furthermore, successive methylation on the nitrogen gives caffeine
through theobromine, while a different methylation sequence can result in the formation of
theophylline (Table 2) [6].

S.N. MethodExperiment Detection Linearity range Application Scientific outcome

Ref.no.

1 UV spectroscopy Caffeine separated from coffee using paper and TLC and was estimated
using spectroscopy Detection was done at 272 nm NA Caffeine from coffee Good
separation [7]

2. UV spectroscopy Caffeine separated from coffee using TLC and was estimated using
spectroscopy Absorbance measured at 274 nm 2–120 μg/ml Caffeine from tea powder
Good separation [8]

3. UV spectroscopy Method A: simultaneous equation method

Method B: isosbestic point method For method A: absorbance measured at 273 nm

For method B: absorbance measured at 259.5 nm 2–32 μg/ml Tablet containing caffeine and
paracetamol Determination of caffeine in mixture of tablets [9]

4. UV spectroscopy Dual wavelength method Two wavelengths of 249 and 234 nm

were selected for analysis

LOD = 0.286

LOQ = 0.863 3–18 μg/ml Tablet containing caffeine and paracetamol A new method of
determination of caffeine [10]

5. HPLC RP-HPLC comprising C18 column and 24% methanol as mobile phase UV detector at
272 nm 1–40 ppm Unroasted coffee and roasted coffee Unroasted coffee contained 0.89–2.10
(8 samples)

Roasted coffee contained 1.03–4.21 (11 samples) [11]

6. HPTLC-UV Silica gel 60F254 as stationary phase and ethyl acetate/methanol (27:3) as
mobile phase UV densitometric remission at 274 nm

LOD = 40 ng/zone

LOQ = 120 ng/zone 2–14 μg/zone Caffeine in marketed tea granules Caffeine in tea samples
was found to be 2.145%[12]

7 HPLC Zorbax eclipse XDB comprising C8 column as stationary phase and water-
tetrahydrofuran-acetonitrile as mobile phase UV detector at 273 nm

LOD = 0.07

LOQ = 0.20 0.2–100 mg/l Caffeine, theobromine, and theophylline in food, drinks, and herbal
products The recoveries range from 92.00 to 96.8% [13]

8 HPLC and biosensor method For HPLC: Shimadzu LC10A fitted with a C18 column as
stationary phase and acetonitrile and water (10:90%) as mobile phase set at a flow rate of 1 ml min−1

For biosensor: amperometricbiosensor comprising the biological sensing element, transducer,

amplification, and detector systems UV detector set at 273 nm 0.01–0.1%w/v

0.01–0.1%w/v Commercial coffee samples and cola drinks 0.033–0.072%w/v

0.030–0.076%w/v [14]

9 HPLC HPLC with solid phase extraction (SPE)

HPLC model: Waters 515, with UV detector (REX, Model pHS-25), Visi TM-1 SPE single-sample processor
(Supelco) 50 mM KH2PO4 (pH = 2)

Acetonitrile and methanol (40:8:2) was used as solvent as well as

mobile phase Caffeine was extracted from green tea, black tea, and coffee and then characterized by
melting point, λ max (UV/vis), IR absorption bands, Rf (TLC), and RT (HPLC) Crude caffeine was purified
by solid phase extraction 10–60 ppm Caffeine in tea, coffee, and soft drinks Crude black
tea, green tea, and coffee contained 7.04%, 4.88%, and 13.7% caffeine, respectively, whereas after
purification black tea, green tea, and coffee contained 3.34%, 2.24%, and 5.20% pure caffeine [15]

10. HPLC and UV UV/vis spectrophotometer

The molar decadic absorption (MDA) coefficients and transitional dipole moment of pure caffeine in
water and dichloromethane (DCM) were obtained at 272 and 274.7 nm MDA was found to be 1115 and
1010 m2 mol−1, respectively, in water and DCM

Transitional dipole moments of caffeine in water and in dichloromethane are 10.40 × 10−30 and 10.80 ×
10−30 C m, respectively 0.90–1.10% for five samples by HPLC Caffeine in coffee beans UV/vis
spectrophotometer: five independent measurements were 1.1 ± 0.01% for Bench Maji, 1.01 ± 0.04% for
Gediyo Yirga Chefe, 1.07 ± 0.02% for Tepi, and 1.19 ± 0.02% for Godere, respectively

HPLC: measurements were 1.10% for Bench Maji, 1.10% for Gediyo Yirga Chefe, 1.00% for Gomma Limu,
and 0.90% for Besema [16]

11 HPLC with DAD Stationary phase: RP-HPLC (Spherisorb ODS2 column)

Mobile phase: 0.01 M phosphate buffer of pH 4 DAD detector at 265 nm

LOD = 0.05 μg/ml 0.05–500 μg/ml Thermal degradation of caffeine in coffee of Brazil and Ivory
Coast For Brazil: green coffee (g/kg of caffeine), 12.36 ± 0.10; roasted coffee, 16.12 ± 0.05

For Ivory Coast: green coffee (g/kg of caffeine), 20.83 ± 0.22; roasted coffee, 25.55 ± 0.185 [17]

12 HPLC Stationary phase: RP-HPLC C18

Mobile phase: acetonitrile/water (8:92%) Detection at wavelength of 245 nm. Varies with
each sample Caffeine and theobromine in coffee, tea, and instant hot cocoa mixes Instant tea:
32.4–35.0 mg/cup of caffeine

Tea bag: 30.2–67.4 mg/cup, 1.0–7.8 mg/cup of caffeine

Instant hot cocoa:46.7–67.6 mg/cup of caffeine

Ground coffee: 93.0–163.5 mg/cup of caffeine [18]

13 LC–MS For LC

stationary phase: Spherisorb S5ODS2, 5 μm

Mobile phase: formic acid/methanol

For MS: ESI source with +ve mode LOD = 11.9 ng/ml

LOQ = 39.6 ng/ml 0.05–25.00 μg/mL Caffeine, trigonelline, nicotinic acid, and sucrose in
coffee Caffeine values ranged from 843.3 to 930.9 mg/100 g coffee in green and roasted Arabica coffee
samples [19]

14 Electrochemical method Voltammetric method with CH1760D electrochemical working

standard

Working electrode: lignin modified glassy carbon electrode

Auxiliary electrode: platinum coil

Reference electrode: Ag/Agcl LOD = 8.37 × 10−7

LOQ = 2.79 × 10−6 6–100 × 10−6 mol/L Caffeine content in Ethiopian coffee samples 10.78,
8.78, 6.35, 5.85 mg/g caffeine in coffee [20]

15 Electrochemical method Voltammetric method

Working electrode: pencil type graphite carbon electrode

Auxiliary electrode: platinum coil

Reference electrode: Ag/Agcl electrode LOD = 9.2 mg/L 0–500 mg/L Caffeine levels in several tea
samples Caffeine levels in several tea samples yield relative error of 1% in the concentrations
[21]

16 LC–MS/MS For LC, stationary phase: RP-HPLC C18

Mobile phase: isocratic mobile phase consisting of 0.2% formic acid in distilled water and methanol
(80:20, v/v)
For MS: spectrometer equipped with an electrospray

Ionization mode used to generate positive [M + H] + ions LLOQ = 5 ng/ml 5–5000 ng/ml
Caffeine and its three primary metabolites in rat plasma [22]

17 GC-NPDStationary phase:

capillary fused silica column

Mobile phase: carrier gas, helium (1 ml min−1) Detection was made by using nitrogen phosphorus
detector

LOD = 0.02 μg/ml

LOQ = 0.05 μg/ml 0.05–500 μg/ml Caffeine in teas, coffees, and eight beverages Caffeine in:
Nescafe coffee = 246.8 μg/ml

Coffee seed = 267.5 μg/ml

Red Bull = 297.9 μg/ml, while other samples contained less caffeine [23]

18 Infrared spectroscopy Fourier transform infrared spectroscopy (FT-IR) method The

measurement was done at 1659 cm−1 using a baseline established between 1900 and 830 cm−1

LOD = 3 mg L−1 NA Caffeine in roasted coffee samples Recovery of all samples ranges from
94.4 to 100.1% [24]

Table 2.

The various analytical methods for the determination of caffeine present in coffee.

AMP = adenosine-5′-monophosphate.

GMP = guanosine-5′-monophosphate.

IMP = inosine-5′-monophosphate.

XMP = xanthosine-5′-monophosphate.
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WRITTEN BY

Hemraj Sharma

Submitted: 20 January 2020 Reviewed: 12 February 2020 Published: 20 March 2020

REGISTER TO DOWNLOAD FOR FREE

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© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative
Commons Attribution 3.0 License, which permits unrestricted use, distribution, and reproduction in any
medium, provided the original work is properly cited.

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