Conclusion.

The results clearly show that although UAE provides a higher and better quality yield the benefits are highly time dependent. Although the extraction yields of caffeine (6-8%) and 5-CQA (0.8-0.9%) are relatively low for both species, if they are put into context with the annual SCG waste, 0.63 billion kg of caffeine and 71.92 million kg of 5-CQA are wasted each year.

These figures along with the antioxidant scavenging and electron transfer abilities provide substantial evidence that SCGs are indeed a feasible source of natural antioxidants as well as caffeine.

Acknowledgments

This work was funded by the Robert Gordon University, Aberdeen. Another vote of thanks is extended to Costa® and Nandos Chicken Land for the donation of the SCGs. I would also like to thank the lab staff and Professor Kong for their time and their help throughout.

N.B If you would like to read the full paper please contact me via the email address listed below.

Gareth fenn

1201058@rgu.ac.ukGarethfenn@hotmail.co.uk

Determination of the polyphenols and caffeine contents recovered from spent coffee grounds and their antioxidant capacities.

GARETH FENNBSc (Hons) Project1201058

An introduction to coffee.

Thought to have originated in a province of Ethiopia, coffee is a member of the Rubiaceae family under the genus Coffeae. According to the integrated taxonomical information system there are 97 genus listed under the Rubiaceae family of which, only the Coffeae genus has true economical value. Coffea Arabica and Coffea Canephora hold the most value with C. Canephora commonly being referred to as Robusta coffee. Robusta is the generic term used to describe a mix of C. Canephora sub species and is known to have a lower cup quality and therefore value compared to C. Arabica.

Producing the bean.

Coffeae plants produce small purple/yellow fruits which contain the coffee bean. Two methods are commonly employed to extract the bean from the coffee cherry, the dry method and the wet method. The dry method involves sun drying the cherries then the removing of the husk and parchment using de-hulling machines. The wet method uses rollers to compress the cherry squeezing the pulp out without damaging the bean. The pulped cherries are fermented to breakdown the mucilage layer. The resulting products are then dried and de hulled to remove the parchment. The beans from both methods are then bagged relating to size with premium beans being polished and the coffee Silver skin (SS) removed. (See figure 1 below)

Figure 1. Components of a coffee cherry

As well as showing species dependance, Figure 3 clearly shows that the extraction aid used has a major impact on the levels of caffeine and 5-CQA as well as the antioxidant activity of the polyphenols.

TE/100µg (FRAP)

caffeine (µg/ml)

5-CQA (µg/ml)

GAE/100mg (FC)

EC50 (µg/ml) (DPPH)

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

Arabica (UAE 150 min) Robusta (UAE 150 min) Robusta (UAE 15min)

Rat

io o

f UA

E/M

S

Figure 3. Standardised ratio of UAE/MS showing a graphical representation of all factors between all three data sets

Although it cannot be directly seen from the above graph, UAE provides an extract with higher antioxidant scavenging and electron transfer abilities than those extracts produced by MS. Figure 3 does however show that the benefits of UAE are highly time dependant, this can be seen by compairing the two robusta sets. Set one was was assisted by UAE for 150mis with set two only being exposed for 15mins. It can be seen that the shorter exposure time not only increased the radical scavenging ability of the antioxidents reducing the EC50 it also increased the reducing power as well as the extraction yeild of caffeine and 5-CQA. It should however be noted that the total polyphenol count for set two was lower.

The average extraction yeild of caffeine and 5-CQA obtained were 6-8% and 0.8-0.9% respectively.

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Determining the caffeine and 5-CQA levels

HPLC-DAD was used to determine the levels of caffeine and 5-CQA within the SCGs. The parameters set were adapted from literature with the following buffers used, 10mM Citric acid (Solvent A) and 50:50 Methanol:Acetonitrile (Solvent B).

For the determination of caffeine, the samples were dissolved in methanol with the following conditions. The flow rate was set at 1.5mL/min with a solvent ratio of 70:30 (A:B). An Agilent C18 5µm column was used for separation and an absorbance wavelength of 233nm selected. The caffeine concentration within the SCG samples were determined via the method of standard addition.

The conditions set for the determination of 5-CQA were as follows; a flow rate of 0.5mL/min was chosen with a solvent ratio of 85:15 (A:B). The column used for the determination of 5-CQA was a Phenomenex C18 3µm column at 325nm.

For the determination of 5-CQA, the samples were dissolved in the mobile phase in a ratio of 85:15 this was in order to reduce the solvent front interfering with the determination. A 3µm column was also used to help aid the separation of the CGAs. This was deemed necessary due to the high levels of similarity within the chemical structures of the different CGAs. The levels of 5-CQA were determined from a valid calibration curve.

Results and conclusions

The results shown on page 6 (figure 3) are given as a ratio of the values obtained via UAE/MS. This method of displaying the data was chosen to standardise any errors caused between days.

The results give an indication that the levels of caffeine and 5-CQA as well as the antioxidant activity of the polyphenols are dependent on the species of SCG. The results, in agreement with literature, show that Robusta SCGs contain a higher percentage of caffeine and 5-CGA as well as exhibiting higher antioxidant activities.

Coffee Waste

The resulting processing waste takes many forms including the coffee SS, husk and pulp The commercial waste produced are known as spent coffee grounds (SCG). SCG are simply the soiled ground bean waste produced from preparing a coffee beverage. The International coffee organisation’s (ICO) world consumption report estimated that in 2014 over 8.9 billion Kg of green coffee bean (GCB) were consumed, generating the equivalent waste in the form of spent coffee grounds (SCG). If we include the waste in the production sector the total mass of coffee waste increases to 16.35 billion Kg.

Health benefits of Coffee

Like most plants the polyphenols and antioxidants, which are naturally produced, are known to be beneficial to the health of the consumer. The seeds and fruits of these plants are known to contain a high percentage of these compounds, hence any beverage of food utilizing these seeds would also be beneficial. This is the reasoning behind the health benefits associated with coffee. Two of the main beneficial compounds found within coffee are: caffeine and a group of polyphenols collectively named Chlorogenic acids (CGAs).

Caffeine

In moderate doses, caffeine has been shown to reduce the risk of cardiovascular diseases and helped prevent type 2 diabetes as well as Parkinson’s. Caffeine’s role as an adenosine antagonist is thought to be the pathway through which it helps prevent neurological degeneration and diseases such as Alzheimer’s, Huntington’s, Parkinson’s and schizophrenia. Nevertheless, due to its metal chelating abilities, caffeine has been shown to increase the risk of developing osteoporosis. However, this effect is commonly counteracted by the addition of milk or cream to the beverage by increasing the calcium intake of the individual.

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Chlorogenic Acids (CGAs)

CGAs are the collective name given to esters of trans cinnamic and quinic acids. These acids have documented antioxidant properties and consist of three sub groups, caffeoylquinic acids (CQAs), feruloylquinic acids (FQA) and dicaffeoylquinic acids (diCQAs). 5-CQA is thought to be the most abundant CGA within coffee (see figure 2 below)

Figure 2. Chemical structures of CQAs1, FQAs2 and diCQAs3

Research suggests that the antioxidant and the radical scavenging properties of the CGAs found in coffee play a role in preventing degenerate brain diseases as well as possessing anti-carcinogenic properties. These scavenging properties differ between each CGA with some being better Oxygen scavengers and others possessing higher metal chelation abilities.

If we consider the brewing procedure associated with coffee, it is logical to assume that the 8.9 billion Kg of SCG produced per annum would still contain these highly beneficial compounds. It was the purpose of this paper to determine the levels of caffeine and CGAs within the SCG and assess their resulting antioxidant capacities.

Extraction.

Before extraction the two samples of SCG, Costa Robusta and Nandos Arabica, were freeze dried to obtain a dry mass. 1g samples of the two SCGs were then exposed to an optimized solvent extraction procedure with some modifications. The extraction procedure was modified using to two different extraction aids. All extractions were repeated in triplicate.

The first set of extractions (3 Robusta and 3 Arabica) were aided by magnetic stirring (MS) for 150 mins with the second set (3 Robusta and 3 Arabica) exposed to an ultrasonic assisted extraction (UAE) for 150 mins. The third set of extractions were carried out on the Robusta SCG only in closed container conditions. For this extraction a comparison between MS for 150min and UAE for 15min was investigated.

The samples were then centrifuged, the solvent evaporated then freeze dried to obtain a dry extract. This extract was used to determine the polyphenol caffeine and 5-CQA levels within the SCG and the resulting antioxidant capacities.

Determining the antioxidant activities

The total polyphenol levels within the extracts were determined using the Folin-Ciocalteau (FC) assay. This assay relies on the electron transfer abilities of the extract requiring the oxidation of the yellow molybdenum (VI) complex to form the blue molybdenum (V) complex.

The antioxidant capacities were assessed via the DPPH* assay and the Ferric Reducing Antioxidant Power (FRAP) assay. The radical scavenging activity of the antioxidants was determined by its ability to convert the purple DPPH* to yellow DPPHH with the results expressed as EC50. This is the concentration required to reduce the absorbance and hence the concentration of DPPH* molecule by 50 %.

The FRAP assay determines the antioxidants electron transfer ability by assessing its ability to reduce the yellow Fe(III) complex to the blue Fe(II) complex

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