recycling

Article
Estimation of Phenolic and Flavonoid Compounds
and Antioxidant Activity of Spent Coffee and Black
Tea (Processing) Waste for Potential Recovery and
Reuse in Sudan
Samar A. Abdeltaif 1 , Khitma A. SirElkhatim 1 and Amro B. Hassan 1,2, * ID

1 Environment and Natural Resource and Desertification Research Institute (ENDRI),

National Center for Research, P.O. Box 6096, 11111, Khartoum, Sudan; [email protected] (S.A.A.);
[email protected] (K.A.S.)
2 Institute of Geography, Faculty of Science, University of Bern, Hallerstrasse 12, 3012 Bern, Switzerland
* Correspondence: [email protected] or [email protected];
Tel.: +41-316-313-856 or +24-911-492-8606




Received: 3 April 2018; Accepted: 11 June 2018; Published: 12 June 2018


Abstract: This study aimed to evaluate the antioxidant power associated with spent coffee and black
tea processing waste. Ethanolic extracts from the samples were prepared in order to determine the
quantities/concentrations of the phenolic and flavonoid compounds, polyphenols, and associated
levels of antioxidant activity. The results showed that both the spent coffee and black tea waste
had high amounts of phenolic compounds and high antioxidant activity rates. The total phenolic
and flavonoids content was found to be significantly higher (p < 0.05) in the spent black tea than
in the spent coffee. The total phenolic content was found to be 152.8 and 97.87 mg of gallic acid
equivalent/g, while the total amount of flavonoids was found to be 47.40 and 34.32 mg catechin/g in
spent black tea and coffee, respectively. However, the spent coffee had a significantly higher (p < 0.05)
antioxidant activity than that detected in the spent black tea (57.83%). Consequently, the results
revealed that the waste residue of spent coffee and black tea may be considered as natural sources of
bioactive compounds and that there may be potential for recycling these waste products, which could
be applied in different industries to further develop functional foods.

Keywords: coffee; black tea; waste; phenolic; flavonoids; antioxidants activity

1. Introduction
Recently, wasted food has received more consideration in both academic and social interest groups
in developed and developing countries. The interest and concern stem from the annual increase in the
amount of food waste that is expected to reach about 126 Metric tons in the next few years [1]. Such a
notable increase in the rate of waste production has a negative impact on the environmental, economic,
and social sectors. In response to these observations, several policies and actionable recommendations
have focused on recycling and the production of natural bioactive compounds from food waste as
potential solutions [2]. Generally, the waste that is produced from the food manufacturing processes
is considered as a viable potential source of natural bioactive compounds with a high antioxidant
power. These compounds, which include phenolic compounds and other phytochemical compounds,
have high-value products and their recovery may be economically attractive [3–5]. Moreover, they have
nutritional and health benefits for humans. Besides these benefits, antioxidants have important
contributions to the food industry. They are capable of preventing the propagation reaction during
the oxidation process, which results in maintaining the quality and shelf-life of food products during

Recycling 2018, 3, 27; doi:10.3390/recycling3020027 www.mdpi.com/journal/recycling

Recycling 2018, 3, 27 2 of 9

handling and storage [6,7]. Furthermore, these compounds can be reused as natural and functional
additives, since they are natural, inexpensive, and available in enormous amounts [8–10].
In recent times, the use of antioxidants extracted from natural, rather than artificial sources,
in food manufacturing, has increased as a result of several factors, including health and functional
factors. Moreover, additional consideration is being paid to classifying natural compounds as well as
their economic application as effective antioxidants in functional food [9,11].
Coffee and tea are considered to be the richest sources of the bioactive compounds, which contain
an appreciable amount of antioxidants, flavonoids, and phenolic compounds. Consequently,
their processing waste may contain a significant quantity of natural antioxidants [12,13]. It has
been determined that the waste from coffee and black tea residue, particularly from the coffee industry,
has experienced annual increases of an estimated 6 million tons internationally [14]. Technologies
have been developed to recycle coffee and black tea waste by converting this sizable quantity of waste
products into new sources of more desirable natural products. Subsequently, there is a need determine
the potential feasibility and utility of using recycled by-products from spent coffee and black tea waste,
as well as determining the extent of their application in different food processes, so as to manufacture
functional foods. In particular, an estimation of the antioxidant capacity of spent coffee and black tea
waste is required so as to support the exploration of their potential use as an alternative source of
artificial antioxidants in the food industry.
Normally, in Sudan, coffee and black tea are used daily after being prepared in a domestic way,
especially by the traditional sellers. Through the survey, we found that the majority of the sellers
use the same types of coffee and black tea. Unfortunately, there has been no real data regarding the
amount of coffee and tea waste that has been produced by these sellers in Sudan. An accumulation
of this waste poses a problem to the environment, as it is prone to microbial spoilage, and therefore,
there is a need to upgrade the recycling system in order to evaluate the potential for recycling the
by-products from these wastes, to find out the possibilities of their use in different food formulations
or the manufacture of health-promoting products or functional foods in Sudan. Therefore, this study
has evaluated the capacity of phenolic and flavonoids compounds and determined the antioxidant
activity in spent coffee and black tea waste. The assessments were then compared to the findings that
were based on raw coffee and black tea.

2. Materials and Methods

2.1. Collection and Preparation of Coffee and Black Tea Samples

The coffee and black tea samples were purchased from the local market in Khartoum. The waste
from the coffee and black tea was prepared according to the domestic method for processing.
Subsequently, 50 g of black tea leaves was brewed in 1000 mL of hot water (100 ◦ C) for five minutes,
followed by filtering and drying processes in an air-dried oven at 45 ◦ C for 48 h. The coffee was
roasted, following the domestic methods for preparing coffee at a household level. The spent coffee
was prepared by adding 50 g of roasted coffee to 100 mL of water, and it was boiled for 10 min. In order
to obtain the dried spent coffee, the same filtration and drying methods that were used to obtain the
spent black tea were followed.

2.2. Preparation of Extracts

To prepare the extract, the milled sample was mixed with ethanol at a ratio of 1:25 (w/v), and it
was then shaken at ambient temperature for 24 h. The mixture was then filtered using a filter paper
(Whatman No. 1, Double ring, China). The residue was washed with ethanol and the collected extract
was dried under vacuum using a rotary evaporator and kept dry for further analysis. The extracts
were reconstituted by pure ethanol directly before the analysis of the total phenolic content and
total flavonoids.
Recycling 2018, 3, 27 3 of 9

2.3. Determination of Total Phenolic Content

The total phenolic content of the samples was determined by the Folin-Ciocalteu’s reagent
method [15], with slight modification. An aliquot (20 µL) of a dried extract ethanolic solution (1:10 w/v)
was added to 1.58 mL of water and 100 µL of the Folin-Ciocalteu reagent. After five minutes, 300 µL of
the 10% sodium carbonate solution was added to the mixture and was carefully agitated for 10 min.
The mixture was allowed to stand in the dark for two hours at 20 ◦ C. The absorption was measured
at 765 nm using ultraviolet (UV)-spectrophotometer, Jenway, UK. Different concentration of gallic
acid dissolved in pure ethanol were used to prepare the calibration curve (R2 = 0.9672). The total
phenolic content was expressed as milligrams of gallic acid equivalent per gram of dried samples
(mg GAE/g DW).

2.4. Determination of Total Polyphenols

The polyphenols of each sample were estimated using Prussian blue assay, as described by Price
and Butler [16]. About 60 mg of the ground sample was extracted, with 3 mL methanol in a 50 mL
conical flask, and then poured into a filter paper. The tube was quickly rinsed with an additional
3 mL methanol and the content was poured once into the filter paper. The filtrate was diluted to
50 mL with distilled water and was mixed with 3 mL 0.1 M FeCL3 in 0.1 N-HCL for three minutes,
followed by a timed addition of 3 mL 0.008 M K3 Fe(CN)6 . The mixture was allowed to stand for
10 min. The absorption was read at 720 nm using spectrophotometer.

2.5. Determination of Total Flavonoids Content

The total flavonoids content (TFC) of the extracts was measured according to the colourimetric
assay of Kim et al. [17]. One milliliter of the ethanolic extract (1:10 w/v) was added to a 300 µL sodium
nitrite solution (5%), followed by 300 µL aluminium chloride (5%). The mixtures were incubated
at room temperature for five minutes, and then 2 mL of 1 mol/L sodium hydroxide was added.
Immediately, the volume of the reaction mixture was made to 10 mL with distilled water and was then
thoroughly vortexed. The absorbance of the mixture was determined at 510 nm. A calibration curve
was prepared from the different concentrations of catechin (R2 = 0.974). The total flavonoid content
was reported as milligrams of catechin equivalents, per gram dry weight sample (mg CE/g DW).

2.6. Determination of Antioxidant Activity

The scavenging activity of the diphenyl-2-picrylhydrazyl (DPPH) radicals of extract was measured
according to the method that was reported by Chang et al. [18], which was cited in Shyur et al. [19].
Assays were performed in 3 mL reaction mixtures containing 2.0 mL of 0.1 mM DPPH-ethanol
solution, 0.9 mL of 50 mM Tris-HCl buffer (pH 7.4), and 0.1 mL of deionized water or sample extract.
The absorbance was measured at 517 nm. The capacity to scavenge for the DPPH radical was calculated
using the following equation:

Scavenging of DPPH% = [(Absorbance control − Absorbance sample)/Absorbance control] × 100

2.7. Statistical Analysis

All of the data were conducted using a completely randomized block design with at least three
replicates. The data were analyzed using a two-way analysis of variance (ANOVA) and the significant
differences were calculated (p < 0.05) the using least significant difference (LSD).
Recycling 2018, 3, 27 4 of 9

3. Results and Discussion

3.1. Phenolic Compounds of Raw and Spent Coffee and Black Tea
Figure 1 describes the total phenolic content of coffee (raw, roasted, and spent) and black tea
(raw and
Recycling spent
2018, tea).PEER
3, x FOR It was clearly
REVIEW observed that both the coffee and black tea contained a reasonable 4 of 9
amount of phenolic compounds, however, the total phenolic compound in black tea was significantly
(p
(p << 0.05)
0.05) higher
higher than
than that
that in
in coffee,
coffee, which
which waswas found
found toto be
be 104.3
104.3 and
and 187.50
187.50 mgmg GAE
GAE /g/g in
in raw
raw coffee
coffee
and
and black
black tea,
tea, respectively.
respectively. Similarly,
Similarly, thethe spent
spent coffee
coffee contained
contained less
less of
of the
the total
total phenolic
phenolic content
content
(97.87
(97.87 mgmg GAE/g)
GAE/g)than thanthat
thatin
inthe
thespent
spentblack
blacktea
tea (152.87
(152.87 mg
mg GAE/g).
GAE/g).The Theanalysis
analysisof ofvariance
variance showed
showed
that there was no significant (p < 0.05) difference between the total phenolic
that there was no significant (p < 0.05) difference between the total phenolic in the raw and in the rawspent
and coffee,
spent
coffee,
however, however, it wassignificantly
it was found found significantly
(p < 0.05)(pless
< 0.05)
in theless in the
spent spent
black tea black
comparedtea compared
with the rawwith the
black
raw black tea. Previous studies showed that both the coffee and black tea contained
tea. Previous studies showed that both the coffee and black tea contained a high amount of the phenolic a high amount
of the phenolic
compound. compound.
However, However,
the capacity of the capacity
these of these
compounds compounds
depended depended
on several on several
factors, factors,
including the
including
varieties and the extraction
varieties and extractionGeremu
procedures. procedures. Geremu
et al. [20] et al.that
reported [20]a reported
significant that a significant
variation in the
variation
content ofin thethe content
phenolic of the phenolic
compound in coffeecompound
cherry pulps in was
coffee cherry between
observed pulps was observed
different between
varieties that
different varieties
were extracted bythat were extracted
different solvents. by differentto
According solvents. According
the previous worktoofthe previous
Zuorro andwork of Zuorro
Lavecchia [21],
and Lavecchia
the spent coffee[21], the spent
grounds coffeecollected
that were groundsfromthat were
coffeecollected from coffee
bars or recovered bars
from or recovered
coffee capsules from
were
coffee capsules were investigated as a potential source of phenolic
investigated as a potential source of phenolic compounds and energy. They demonstrated compounds and energy.thatThey
the
demonstrated that the total
total phenolic compound ofphenolic compound
coffee ranged of coffee
between 17.75 ranged
and 21.56between 17.75 and 21.56 mg GAE/g.
mg GAE/g.

raw roasted spent

250

200 a

b
Total phenolic

150
mg GAE/g

c
100 c

d
50

0
Coffee Tea

Figure 1.
Figure Phenolic content
1. Phenolic content of
of spent
spent coffee
coffee and
and black
black tea
tea waste.
waste. Data
Data represent
represent the mean ±
the mean standard
± standard
deviation (SD) (n = 3). a–d Means with different letters are significantly different (p ≤ 0.05) as assessed
deviation (SD) (n = 3). Means with different letters are significantly different (p ≤ 0.05) as assessed
a-d

by least
by least significant
significant difference
difference (LSD).
(LSD). GAE—gallic
GAE—gallic acid
acid equivalent.
equivalent.

Figure 2 shows the content of total polyphenol in raw and spent coffee and black tea. As shown
in Figure 2, the total
total polyphenol
polyphenol content
content ofof the
the raw,
raw, roasted,
roasted, and
and spent
spent coffee
coffee was
was found
found to
to be
be 43.93,
43.93,
37.82 and 23.79 mg/g,
mg/g,respectively.
respectively. The
The analysis
analysis ofof variance
variance indicated
indicated that
that the
the total
total polyphenol
polyphenol was
significantly (p << 0.05)
0.05) varied
varied in
in the
theraw,
raw, roasted,
roasted, and
and spent
spent coffee,
coffee, which
which might
might have
have been
been influenced
influenced
by heat treatment. In In contrast,
contrast, the
the total
total polyphenol
polyphenol content
content of
of the
the raw
raw and
and spent
spent black
black tea
tea was
considerably (p << 0.05)
0.05) the same, and it was found to have been 40.35 and 42.52 mg/g in the raw and
42.52 mg/g in the raw
spent black tea, respectively.
Recycling 2018, 3, 272018, 3, x FOR PEER REVIEW
Recycling 5 of 9 5 of 9

raw roasted spent

50
a
ab
b
40 c

Total poly phenol 30

mg/g d
20

10

0
Coffee Tea

Total 2.polyphenols
Figure 2. Figure (mg/g)
Total polyphenols (mg/g) content
content of of spent
spent coffeecoffee and
and black teablack tea waste.
waste. Data representData represent the
the mean
a-da–d
mean ± SD (n = 3).
± SD (n = 3). Means with different letters are significantly different (p ≤ 0.05) as assessed
Means with different letters are significantly different (p ≤ 0.05) as assessed by LSD.
by LSD.
Flavonoids have been considered as the most common, important, and widely distributed single
group of phenols that are present in plants with highly effective antioxidants [22]. Flavonoids have
inhibitedhave
Flavonoids metal-initiated lipid oxidation
been considered as theby most
forming complexesimportant,
common, with metal ions
and[23]. The content
widely of
distributed single
the total flavonoids of the raw and spent coffee and black tea is seen in Figure 3. The total flavonoid
group of phenols that are present in plants with highly effective antioxidants [22]. Flavonoids have
content of coffee and black tea was determined as a form of catechin equivalents. It was cleared that
inhibited the
metal-initiated lipid oxidation by forming complexes with metal ions [23]. The content of
values of total flavonoids were varied between raw, roasted, and spent coffee. It was found to be
the total flavonoids of the
52.07, 30.65, and 34.32 raw
(mgand spent coffee
catechin/g), and black
respectively, as seentea
in is seen3.in
Figure TheFigure
analysis3. of
The total flavonoid
variance
showed that raw coffee contained significantly (p < 0.05) higher flavonoids than
content of coffee and black tea was determined as a form of catechin equivalents. It was cleared thatthe roasted and spent
the valuescoffee, however, there was no significant difference between roasted and spent coffee that was
of total flavonoids were varied between raw, roasted, and spent coffee. It was found to
observed. On the other hand, both the black tea and spent black tea contained a similar amount of
be 52.07, 30.65, and 34.32
total flavonoids, (mg
which wascatechin/g),
found to haverespectively,
been 49.57 and as seen
47.40 (mgin Figure 3.respectively.
catechin/g), The analysis In theof variance
showed thatstudyraw coffee
that was contained
conducted significantly
to compare the content(p of <
the0.05) highercapacities
antioxidants flavonoids thantypes
of different the roasted
of and
raw however,
spent coffee, and spent tea [24], the
there was results showed that difference
no significant both the rawbetween
and spent tea contained
roasted anda spent
high amount
coffee that was
observed.ofOn total flavonoids, however, it was found to be less in the spent tea compared with the tea. In general,
the other hand, both the black tea and spent black tea contained a similar amount of
the variation between our findings and the other results might have been because of the extraction
total flavonoids,
methods,which was
as it was foundthat
reported to the
have been 49.57
flavonoid andwas
extraction 47.40 (mg by
affected catechin/g),
the extraction respectively.
method, In the
study thattemperature
was conducted(above 40to°C),
compare the content
and extraction time. of the antioxidants capacities of different types of
raw and spent tea [24], the results showed that both the raw and spent tea contained a high amount of
total flavonoids, however, it was found to be less in the spent tea compared with the tea. In general,
the variation between our findings and the other results might have been because of the extraction
methods, as it was reported that the flavonoid extraction was affected by the extraction method,
temperature (above
Recycling 2018, 3,40 ◦ C),
x FOR and
PEER extraction time.
REVIEW 6 of 9

60 raw roasted spent

a
ab b
50
Total flavonoids

40
c
mg CE/g

c
30

20

10

0
Coffee Tea

Figure 3. Flavonoids content

Figure 3. Flavonoids of spent
content coffee
of spent coffeeand
and black teawaste.
black tea waste.DataData represent
represent the± SD
the mean mean ± SD (n = 3).
(n = 3).
a–c Meansa-cwith
Means with different
different letters
letters areare significantly different
significantly different ≤ 0.05)
(p ≤(p0.05) as assessed by LSD. by LSD.
as assessed

3.2. Antioxidant Activity of Raw and Spent Coffee and Black Tea
In this study, the DPPH scavenging method was used to assess the antioxidant activity of the
samples. The scavenging model of the DPPH radical was widely used as a method for assessing the
antioxidant activity in a period that was relatively short compared with the other methods. Figure 4
displays the antioxidant activity of coffee (raw, roasted, and spent) and black tea (raw and spent), in
terms of DPPH scavenging. As shown in Figure 4, the antioxidant activity rate was found to have
 10

0
Coffee Tea
Recycling 2018, 3, 27 6 of 9

Figure 3. Flavonoids content of spent coffee and black tea waste. Data represent the mean ± SD (n = 3).
a-c Means with different letters are significantly different (p ≤ 0.05) as assessed by LSD.
3.2. Antioxidant Activity of Raw and Spent Coffee and Black Tea
In
3.2.this study, the
Antioxidant DPPH
Activity scavenging
of Raw method
and Spent Coffee and was
Black used
Tea to assess the antioxidant activity of the
samples. In Thethis study, the DPPH scavenging method was usedwidely
scavenging model of the DPPH radical was used
to assess the as a methodactivity
antioxidant for assessing
of the the
antioxidant
samples.activity in a period
The scavenging modelthat
of was relatively
the DPPH short
radical wascompared
widely usedwithas a the other
method formethods.
assessing Figure
the 4
displays the antioxidant
antioxidant activity in aactivity of coffee
period that (raw, roasted,
was relatively and spent)
short compared withand blackmethods.
the other tea (rawFigure
and spent),
4
in terms of DPPH
displays scavenging.
the antioxidant As shown
activity of coffeein(raw,
Figure 4, theand
roasted, antioxidant
spent) and activity
black tearate
(rawwasand found
spent), to
in have
terms of DPPH scavenging. As shown in Figure 4, the antioxidant activity rate
been significantly (p < 0.05) higher in the raw, followed by spent, and then roasted coffee 76.79%, was found to have
been
72.93%, andsignificantly (p < 0.05) higher
59.37%, respectively. in the raw, followed
The degradation by spent, and
of the antioxidant then roasted
activity of coffee coffee
might76.79%,
have been
72.93%, and 59.37%, respectively. The degradation of the antioxidant
because of the degradation of the native antioxidants and the formation of new ones duringactivity of coffee might havecoffee
been because of the degradation of the native antioxidants and the formation of new ones during
roasting [25]. For black tea, it was observed that the antioxidant activity of the raw and spent was
coffee roasting [25]. For black tea, it was observed that the antioxidant activity of the raw and spent
significantly (p < 0.05) the same. It ranged between 57.83% and 59.27%, however, it was significantly
was significantly (p < 0.05) the same. It ranged between 57.83% and 59.27%, however, it was
lowersignificantly
than the rate of than
lower the antioxidant
the rate of theactivity in coffee.
antioxidant activityMore or less,
in coffee. Moretheortrend that
less, the wasthat
trend observed
was in
the DPPH results was the same when comparing the total phenolic compound
observed in the DPPH results was the same when comparing the total phenolic compound and total and total flavonoids
content. Thus, content.
flavonoids fundamentally, the DPPH scavenging
Thus, fundamentally, ability might
the DPPH scavenging have
ability depended
might on the on
have depended amount
the of
total amount
phenolic of compound
total phenolic and total flavonoids
compound and total content
flavonoids in content
the coffee and
in the black
coffee teablack
and extracts.
tea extracts.

raw roasted spent

90
a
75 b
Antioxidant activity

c c
60 c

45
%

30

15

0
Coffee Tea

Figure 4. Antioxidant activity of spent coffee and black tea waste. Data represent the mean ± SD
(n = 3). a–c Means with different letters are significantly different (p ≤ 0.05) as assessed by LSD.

The results were in agreement with the statement that was reported by Lee et al. [20], who stated
that the extracts that were enriched with flavonoids or phenolic, showed a much higher DPPH
scavenging ability than those of the other extracts. Moreover, Romdhane et al. [26] found that
the antioxidant activity of the coffee residual ranged between 64.57% and 52.83%. Similarly,
Farhoosh et al. [27] reported that the old black tea leaves and black tea waste had a high antioxidant
activities rate.
The correlation coefficient (R2 ) values between the antioxidant parameters in the raw and spent
coffee and black tea are shown in Table 1. From the table, it was clearly observed that there was an
appositively high correlation between the antioxidant parameters in the raw and spent coffee and
black tea. The correlation values in the parameters between the raw and spent coffee were found to be
0.992 and 0.997 between the raw and spent black tea. On the other hand, the correlation between the
coffee and black tea were found to be 0.9078 in the raw samples, and 0.857 in the spent samples.
Recycling 2018, 3, 27 7 of 9

Table 1. Correlation coefficient (R2 ) between the antioxidants parameters in the raw and spent coffee
and black tea.

Raw Coffee Spent coffee Raw Tea Spent Tea

Raw coffee 1
Spent coffee 0.9922 1
Raw tea 0.9078 0.8491 1
Spent tea 0.9141 0.857 0.997 1

In general, the obtained results revealed that the waste from the processed coffee and black
tea contained appreciable amounts of the phenolic compound, as well as a high antioxidant activity
rate. The highly positive correlation in the antioxidant parameters between the coffee and black
tea waste revealed that they had the potentiality to be an alternative natural resource for the
bioactive compounds.
According to Ghafoor et al. [28], the phenolic compounds from natural resources were
recommended as natural food additives and they were considered to be more suitable for application
in food products compared with artificial compounds that contained antioxidant properties. Moreover,
the addition of phenolic in food was also reported for its nutritional and health benefits [29].
Furthermore, a positive correlation between the antioxidant power and total phenolic content indicated
that the phenolic compound could be one of the main contributors to the antioxidant capacities of this
waste [30,31].

4. Conclusions
In conclusion, the values of the phenolic compounds in the spent coffee and black tea were
high, indicating that they could be inexpensive and readily available resources of natural bioactive
compounds for use in the food industries. Therefore, their wastes could be considered as a good source
of bioactive compounds and could be, potentially, applied as a natural antioxidant for developing
functional foods. Furthermore, both the spent coffee and black tea may be considered as a low-cost
resource for the recovery of phytochemicals, which may have a significant outlook as pharmaceuticals,
cosmetic constituents, and food additives, for the industry in Sudan. However, an evaluation of the
quality and quantity of the bioactive compounds in coffee and black tea waste is required. In addition,
further research, such as the application of different extraction procedures and extraction solvents, will
be needed in order to optimize the extraction condition at the industry level. Moreover, the bioactivity,
bioavailability, and toxicology of coffee and black tea waste phytochemicals need to be carefully
evaluated by in vitro and in vivo studies before considering the likelihood of the reuse of this waste in
food industries in Sudan.

Author Contributions: Authors S.A.A. and K.A.S. performed the practical experiments. Author A.B.H. conceived
and designed the experiments, analyzed the data, supervised the study and, wrote the manuscript.
Acknowledgments: This study was carried out with the support of the National Center for Research, Sudan (Food
Waste Recycling Project).
Conflicts of Interest: The authors declare no conflict of interest.

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