Chemical composition and antioxidant activity of Coccoloba cowellii Britton

Abstract

The qualitative and quantitative chemical composition (total phenolics, tannins and flavonoids contents), as well as the antioxidant activity of an ethanolic extract of the leaves of Coccoloba cowellii (Polygonaceae) was determined in this study. The extract was prepared by dynamic ultrasound-assisted extraction using firstly petroleum ether:ethyl ether (1:1, v/v) and therefore ethanol:water (8:2, v/v). Quantitative chemical composition (total phenolics, tannins and flavonoids content) was determined using spectroscopic techniques, while the antioxidant activity was determined by means of the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) and ferric reducing power tests. The ethanolic extract showed significant antioxidant activity in both assays, with values of 34,01 % ± 4,03 % in the DPPH and reduced Fe 3+ ferricyanide complex to the ferrous form (Fe 2+), similar to the ascorbic acid used as standard in both assays. The antioxidant activity of the extract is consistent with the total phenolic, tannins and flavonoid contents of the extract.

Full text

Rev. Cubana Quím.
Vol.32, no.2, enero-abril, 2019, págs. 185-198, e-ISSN: 2224-5421
http://ojs.uo.edu.cu/index.php/cq
Composición químicay actividad antioxidante de Coccoloba
cowellii Britton
Chemical composition and antioxidant activity of Coccoloba
cowellii Britton
Lic. Daniel Méndez-Rodríguez I, Dr. C Enrique Molina-Pérez I, Dr. C Iraida
Spengler-Salabarria II, Dr. C Julio César Escalona-Arranz III, Dr. C Paul Cos IV
daniel.mendez@reduc.edu.cu; jcea@uo.edu.cu
I Universidad de Camagüey, Cuba; II Universidad de la Habana, Cuba; III Universidad
Oriente, Cuba; IV Laboratory for Microbiology, Parasitology and Hygiene, Faculty of
Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp,
Universiteitsplein 1, 2610 Antwerp, Belgium.
Recibido: 14 de noviembre de 2018 Aprobado: 11 de febrero de 2019
Resumen
En este trabajo se determinó la composición química cualitativa y cuantitativa (contenido de
compuestos fenólicos, taninos y flavonoides totales), así como la actividad antioxidante in vitro
de un extracto etanólico de las hojas de Coccoloba cowelli (Polygonaceae). Para ello se realizó
una extracción dinámica asistida por ultrasonido con éter de petróleo:éter etílico (1:1, v/v) y
luego con etanol:agua (8:2, v/v). En los extractos se cuantificaron los compuestos fenólicos,
taninos y flavonoides totales mediante técnicas espectrofotométricas. La actividad antioxidante
fue determinada mediante los ensayos del radical 1,1-difenil-2-picrilhidracil (DPPH) y del poder
reductor. El extracto etanólico mostró actividad antioxidante significativa en ambos ensayos,
con valores de 34,01 % ± 4,03 % en el DPPH y reduciendo el complejo Fe3+ ferricianuro a su
forma ferrosa, los que fueron comparables al del ácido ascórbico usado como estándar. La
actividad antioxidante es consistente con el contenido de compuestos fenólicos, taninos y
flavonoides totales del extracto.
Palabras clave: fenoles totales, taninos, flavonoides, actividad antioxidante, extracción
dinámica asistida por ultrasonido.
Abstract
The qualitative and quantitative chemical composition (total phenolics, tannins and flavonoids
contents), as well as the antioxidant activity of an ethanolic extract of the leaves of Coccoloba
cowellii (Polygonaceae) was determined in this study. The extract was prepared by dynamic
ultrasound-assisted extraction using firstly petroleum ether:ethyl ether (1:1, v/v) and therefore
ethanol:water (8:2, v/v). Quantitative chemical composition (total phenolics, tannins and
flavonoids content) was determined using spectroscopic techniques, while the antioxidant
activity was determined by means of the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) and ferric
reducing power tests. The ethanolic extract showed significant antioxidant activity in both
assays, with values of 34,01 % ± 4,03 % in the DPPH and reduced Fe3+ ferricyanide complex to
the ferrous form (Fe2+), similar to the ascorbic acid used as standard in both assays. The
antioxidant activity of the extract is consistent with the total phenolic, tannins and flavonoid
contents of the extract.

Lic. Daniel Méndez-Rodríguez, Dr. C Enrique Molina-Pérez, Dr. C Iraida Spengler-Salabarria,
Dr. C Julio César Escalona-Arranz, Dr. C Paul Cos
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Keywords: total phenolics, tannins, flavonoids, antioxidant activity, dynamic ultrasound-
assisted extraction
Introducción
The consumption of herbal medicines is increasing over the world, primarily due to its
benefits. The World Health Organization estimates, that nearly 80 % of the world’s
population in developing countries relies primarily on herbal medicine to satisfy their
healthcare needs [1]. Besides, in recent years, compounds derived from medicinal and
aromatics plants have been commercially exploited by the pharma, food, flavor,
fragrance, dyeing and pesticide industries.
Among the various biological activities reported for plants, the antioxidant stands out,
related to the need to have protectors against oxidative stress. Reactive oxygen species
(ROS) and reactive nitrogen species (RNS) are continuously generated under
physiological conditions and are involved in the growth, differentiation, progression and
death of cells. Low concentrations of these species are beneficial and even
indispensable for processes such as intracellular signaling and defense against
microorganisms [2]. However, when the body's natural antioxidant defenses are
overcome by excessive production of ROS/RNS, the so-called "oxidative stress" occurs.
In this state, cellular and extracellular macromolecules (lipids, proteins and nucleic
acids) can suffer oxidative damage, causing harm to different tissues.
Because in certain occasions the innate defenses are not enough against a severe or
continuous oxidative stress, it is necessary to supply certain amounts of exogenous
antioxidants to maintain an adequate value of them in order to balance ROS/RNS levels.
Because of this, there has been an increase in interest regarding natural antioxidants,
such as polyphenols, present in medicinal and edible plants, which could help in the
prevention of oxidative stress.
The Polygonaceae are a family of plants belonging to the order Caryophyllales, and
comprise approximately 1 200 species distributed in approximately 48 genera. The
largest genera are Eriogonum (240 species), Rumex (200 species), Coccoloba (120
species), Persicaria (100 species) and Calligonum (80 species). The family is present all
over the world, but it is more diverse in the Northern Temperate Zone.
The genus Coccoloba comprises approximately 120-150 shrubs and trees, mostly
perennialsflowering plants, of which more than 25 occurs in Cuba. It is native to the

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tropical and subtropical regions of America, in South America, the Caribbean and
Central America, with two species that extend to Florida.
Biological assays revealed antifungic activity in C. acrosticoides [3] and C. dugandiana
[4], antibacterial in C. crosticoides [3] and C. Cozumelensis [5] and alelochemical in C.
uvifera [6]. In relation to the chemical profile of the genus Coccoloba, the species until
now investigated are represented by flavonoids [4, 7], terpenoids [3, 8], benzenoids [4]
and carboxylic acids and esters [9].
C. cowellii, a critically endangered plant, endemic of serpentinitic savannas of
Camagüey province, is locallyknown as "Uverillo" and "Moco de guanajo”. In the
scientific literature, there are no publications related to the phytochemistry and
biological activity of this specie. Taking into account endemism of C. cowellii and
biological activities and chemical composition reported in allied species, this work is
aimed to determine and quantify the phenolics content of the leaves of C. cowellii and to
evaluate its antioxidant properties in vitro.
Materials and methods
Plant Material
Leaves of C. cowellii (fig. 1) were collected in april 2018 near to Albaisa, in the
municipality of Camagüey (Lat 21.43615, Lon -77.83253). Plant was taxonomically
identified by the curator of “Julián Acuña Galé” herbarium at the University of
Camagüey (HIPC), where a voucher specimen was deposited (number 12057).
Plantmaterial after cleaned was dried on the shadow at room temperature until constant
weight. Afterward, it was milled in a blade mill and stored in the dark at a dry place
until further use.
Fig. 1. Young Coccoloba
cowelliishrub

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Chemical reagents
Folin-Ciocalteu, DPPH (1,1-diphenyl-2-picryl-hydrazyl), ferric chloride, gelatin and
quercetin reagents were purchased from Sigma(USA). Tannic acid was from BDH
Laboratory reagents (England). All other used reagents were purchased from UNI-
CHEM Chemical Reagents (China). All solvents used for extraction and
chromatography were of analytical purity grade.
Extracts preparation
A dynamic ultrasound-assisted extraction device, based on the method suggested by
Xungang et al. [10] with some modifications was constructed for the extraction. Five
grams of plant material were filled into extraction cell (9 x 2 cm, i.d.). The column was
prepared by adding a small plug of cotton, plant material, and suitable amount of
cottonas shown in fig. 2, to form an extraction column. The column was set in an
ultrasonic cleaner from Scientz, model SB-3200STD (China), vibration frequency at 40
kHz, 80 % power, with inlet connected to a peristaltic pump by a tube, and outlet to a
flask. Dynamic ultrasound-assisted extraction was carried out by continuously feeding
extraction solvent into the column by the pump and assisting by ultrasonic wave, where
a flow rate of 3 mL/min was used, and temperature was set at 40 ºC by adjusting the
regulation of ultrasonic bath. The plant material was firstly defatted with 1 mL of ethyl
ether: petroleum ether (1:1, v/v) for 1 min and dynamically extracted for 35 min. Then a
second dynamic extraction was performed using aqueous ethanol (8:2, v/v) for 35 min
to obtain the ethanolic extract. After the procedure, the fraction of the effluent (≈100
mL) was collected into 150 mL of volumetric flask.
Fig. 2. Schematic diagram of dynamic ultrasound-assisted extraction (1, Solvent; 2, Peristaltic
pump; 3, Extraction cell (9×2 cm, i.d.); 4, Ultrasonic bath; 5, Collection flask; 6,
Cotton; 7, Plant material)

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Qualitative phytochemical studies
The ethanolic extract was subjected to qualitative phytochemical analysis. Presence of
various phytochemicals such as phenols, steroids, glycosides, saponins, flavonoids,
terpenoids, alkaloids and reducing sugars were determined by various phytochemical
tests [11].
Quantification of Total Phenolic content (TPC)
The total phenol concentration of the sample was determined using the Folin-Ciocalteu
method described by Makkar [12]. Briefly, 50 μL of ethanolic extract in 950 μL of
methanol were mixed with 1,0 mL of distilled water and 0,5 mL of Folin-Ciocaleu
reagent (1N) in a test tube. After vortexed the tube and allowed to stand for 5 min at
room temperature, 2,5 mL of 5 % aqueous sodium carbonate were added, the tube was
vortexed again and incubated for 40 min in a dark place. The absorbance was measured
at 725 nm using a UV/Vis spectrophotometer from Rayleigh, model UV-1601 (China).
A standard curve using tannic acid (TA) was created with 5 points. Total phenolic
concentration was expressed as tannic acid equivalent in mg per gram of defatted
extract (mg TA/g extract). The methanol solution was used as a blank. All assays were
carried out in triplicate.
Quantification of Total Tannin Content (TTC)
The gelatin reagent for the precipitation of tannins was prepared according to Velázquez
[13], mixing in a volumetric flask 5 mL of the 10 % gelatin solution and 10 mL of 10 %
NaCl solution in 1 % HCl. The mixture was stirred well and adjusted to 50 mL with
distilled water, then allowed to stand for 30 minutes to allow the insoluble solids to
settle, filtered and the filtered solution was used as a reagent for the analysis of total
tannins. After that, 500 μL of the ethanolic extract was mixed with 500 μL of the gelatin
reagent and incubated for 30 minutes in a water bath at 37 °C, then centrifuged at 5 000
rpm for 10 min using a centrifuge from Yingtai Instrument, model TG16 (China), and
500 μL of the supernatant was taken for the determination of total phenols by the
method of Folin-Ciocalteu as previously described [12]. The total content of tannins
was calculated using the following formula:
Tannins (g) = Total phenolics (g) − Non-tannin phenolics (g). (1)
Quantification of Total Flavonoid Content (TFC)
The amount of total flavonoids in the extracts was measured spectrophotometrically as
previously reported [14]. Briefly, 500 μL of the extract were mixed with 1,50 mL

Lic. Daniel Méndez-Rodríguez, Dr. C Enrique Molina-Pérez, Dr. C Iraida Spengler-Salabarria,
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of 95 % ethanol, 0,10 mL of 10 % aluminum chloride (AlCl3.6H2O), 0,10 mL of sodium
acetate (NaC2H3O2) (1 M) and 2,80 mL of distilled water. After incubation for 40 min,
absorbance was measured at 415 nm using a UV/Vis spectrophotometer from Rayleigh,
model UV-1601 (China). To calculate the concentration of flavonoids, a calibration
curve was prepared using quercetin (QE) as standard. The flavonoid concentration is
expressed as quercetin equivalents in mg per gram of defatted extract (mg QE/g
extract). All assays were carried out in triplicate.
Evaluation of antioxidant activity
Free radical-scavenging ability by the use of a stable DPPH radical
The DPPH radical-scavenging activity was determined using the method described by
Blois [15] with slightly modifications. Briefly, 5 mL of a 0,1 mM DPPH solution was
mixed with sample solutions at different concentrations (10, 20, 30, 40, 50 µg/mL). A
control (Abs Control) containing methanol and DPPH solution was also prepared. The
mixture of methanol, DPPH and ascorbic acid served as positive control. All solutions
obtained were then incubated for 20 min at room temperature. The radical scavenging
capacity was evaluated by measuring the decrease of absorbance at 517 nm using a
UV/Vis spectrophotometer from Rayleigh, model UV-1601 (China). The percentage of
inhibition of samples was calculated from obtained absorbances by the equation:
% inhibition = ((Abs control-Abs test)/Abs control) × 100(2)
Then, a curve was constructed by plotting percentage of inhibition against concentration
in µg/mL. All assays were carried out in triplicate.
TLC-DPPH test
In order to relate specific compounds with the antioxidant activity, the assay previously
described was develop on a Thin Layer Chromatography plate (TLC). The stable 2,2-
diphenyl-1-picrylhydrazyl radical (DPPH) has an absorption maximum at 517 nm,
which decreases upon reduction through reaction with a radical scavenger. The
corresponding color change can thus be observed in a TLC bioassay [16]. The TLC
plate with the sample was developed with the elution solvent (chloroform:ethyl
acetate:methanol 2:3:1, v/v/v) and then dried. It was then sprayed with a solution
of 0,2 % (DPPH) in methanol. The plate was examined in daylight after 30 min. Active
(free-radical scavenging) compounds appear as yellow-white spots against a purple
background. To compare, another two plates were developed with the same solvent,

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dried and revealed with 254 nm UV light and sulfuric acid-vanillin reagent,
respectively.
Reducing power ability
The reducing power of plant extracts can be detected based on the ability to reduce
ferric ions in the reaction mixture to ferrous ions. The ferricyanide reagent is first
incubated in phosphate buffer at pH 6.6 with antioxidants, and the reduction product,
ferrocyanide, combines with the later added Fe3+ to produce Prussian blue, which is
detected at 700 nm. The assay is based in the following chemical reactions:
Fe(CN)63- + ArOH Fe(CN)64- + ArO• + H+ (3)
Fe(CN)64- + Fe3+ + K+ KFe[Fe(CN)6] (4)
The reduction will favor the production of thecolored complex where the intensity is
dependent on the concentration of reductants. Reducing power was determined using
the method described by Oyaizu [17] with slightly modifications. Briefly, sample
solutions at different concentrations (10, 20, 30, 40, 50 µg/mL) were mixed with 2,5 mL
of 0,2 M phosphate buffer (pH 6,6) and 2,5 mL of 1 % potassium ferricyanide and
incubated at 50 ºC for 20 min. Then, 2.5 mL of 10% trichloroacetic acid were added,
and the tubes were centrifuged at 10,000 rpm for 10 min using a centrifuge from Yingtai
Instrument, model TG16 (China). Then, 2.5 mL of the upper layer were mixed with 2,5
mL of distilled water and 0,5 mL of 0,1 % ferric chloride, and the absorbance of the
reaction mixtures was measured at 700 nm using a UV/Vis spectrophotometer from
Rayleigh, model UV-1601 (China). Ascorbic acid was used as a positive control. All
assays were carried out in triplicate.
Statistical analysis
Statistical analysis was carried out with GraphPad Prism 7 software (Graph Pad
Software, Inc., USA). The data, in the different experiments, were checked for
normality through the Kolmogorov-Smirnov test. Results were expressed as means ±
standard deviation of three replicates.
Results and discussion
Preliminary phytochemical tests for the leaves extract of Coccoloba cowellii
The qualitative analysis of the ethanol extract is showed in table 1. The main
phytochemical constituents of the extract were the phenolic compounds, represented by
tannins and flavonoids, and in a minor degree, reducing sugars. The phenolics

Lic. Daniel Méndez-Rodríguez, Dr. C Enrique Molina-Pérez, Dr. C Iraida Spengler-Salabarria,
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compounds are present in the polar extracts of other species of Coccolobagenus,
specifically tannins (both condensed and hydrolysables) and flavonoids glucosides of
quercetin, catechins and myricetin [3, 18].
TABLE 1. PHYTOCHEMICAL SCREENING OF THE ETHANOLIC
EXTRACT OF COCCOLOBA COWELLII
Chemical constituents
Results of chemical reaction
Alkaloids
-
Triterpenes and steroids
-
Coumarins
-
Phenols and tannins
+++
Free aminoacids
-
Cardiac glycosides
-
Flavonoids
++
Quinones
-
Reducing sugars
+
Saponins
-
Precipitate or coloration: very abundant, +++; abundant, ++; middle, +; not detected, - (the
number of positive signs indicated the intensity of the reactions).
Phenolic, flavonoid and tannin contents
The total phenolic content of the ethanolic extract, calculated from the calibration curve
(Y = 0,018 03*X + 0,013 8; R2 = 0,999) was 264,77 ± 5,47 tannic acid equivalents/g,
the total tannin content (Y = 0,018 03*X + 0,013 8; R2 = 0,999) was 148,02 ± 2,63
tannic acid equivalents/g and the total flavonoid content (Y = 0,006 806*X + 0,474 7;
R2 =0,983) was 177,04 ± 1,08 quercetin equivalents/g (table 2). These results were
comparable to those obtained for the acetone extract of C. uviferaleaves [19], and
demonstrate the high content of this type of secondary metabolites in the evaluated
extract. No other scientific reports regarding the metabolites concentration of other
species for this genus were found in the consulted literature.
Phenolic compounds have redox properties, which allow them to act as antioxidants
[20]. As their free radical scavenging ability is facilitated by their hydroxyl groups, the
total phenolic concentration could be used as a basis for rapid screening of antioxidant
activity. Antioxidant activity of flavonoids, including flavones, flavanols and condensed
tannins, is related to the presence of free OH groups, especially 3-OH. Plant flavonoids
have antioxidant activity in vitro and also act as antioxidants in vivo [21].

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TABLE 2. TOTAL PHENOLICS, TANNINS AND FLAVONOIDS
CONTENT OF ETHANOLIC EXTRACT OF C. COWELLIILEAVES
Total phenolics contenta
264,77 ± 5,47
Total tannins contenta
148,02 ± 2,63
Total flavonoids contentb
177,04 ± 1,08
amg tannic acid equivalent (TAE)/g DW.
bmg quercetin equivalent (QE)/g DW.
Values are mean ±standard deviation of three replicates.
Antioxidant activity
Several methods have been developed for the assessment of antioxidant capacity.
Because many active species and reaction mechanisms are involved in oxidative stress
processes, no simple universal method can be applied for an accurate and quantitative
measurement of antioxidant capacity. Generally, in these methods, a radical is generated
and the antioxidant capability of a sample against the radical is evaluated. In the present
study, the antioxidant activity of C. cowellii ethanolic leaf extract was determined using
the DPPH radical scavenging and the reducing power assay. In the present study, the
antioxidant activity of C. cowellii ethanolic leaf extract was determined using the DPPH
radical scavenging, alone and combined with TLC plates, and the reducing power assay.
DPPH free radical scavenging assay
The radical DPPH has the advantage of being unaffected by certain side reactions, such
as metal-ion chelation and enzyme inhibition, brought about by various additives. A
freshly prepared DPPH solution exhibits a deep purple color with a maximum
absorption at 517nm [22]. This purple color generally fades/disappears when an
antioxidant is present in the medium. Thus, antioxidant molecules can quench DPPH
free radicals (i.e., by providing hydrogen atoms or by electron donation, conceivably via
a free-radical attack on the DPPH molecule) and convert them to a colorless/bleached
product, DPPH-H (i.e., 2,2-diphenyl-1-hydrazine, or a substituted analogous hydrazine),
resulting in a decrease in absorbance at 517nm.
It has been found that ascorbic acid, tocopherol and polyhydroxy aromatic compounds
reduce and decolorize DPPH by their hydrogen donating ability [22]. It appears that C.
cowellii possesses hydrogen donating abilities and can act as an antioxidant through this
pathway.DPPH scavenging activity of C. cowellii ethanolic extract was 34,01 % at a
concentration of 50 µg/mL, while that of the control, ascorbic acid, was 29,35 % (fig. 4,
a).These results were also comparable to those obtained for C. uvifera [23], and showed

Lic. Daniel Méndez-Rodríguez, Dr. C Enrique Molina-Pérez, Dr. C Iraida Spengler-Salabarria,
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that the extract exhibited an antioxidant potential comparable to the antioxidant capacity
of commonly used ascorbic acid.
TLC-DPPH Test
The results showed the presence of radical scavengers in the ethanolic extract as a
yellow-white zone over a purple background (fig. 3, c). The same compounds can be
visualized using 254 nm UV light (fig. 3, a) and the sulfuric acid-vanillin reagent (fig. 3,
b). Taking into account the chromatographic behavior of the extract and the color
developed with the sulfuric acid-vanillin reagent, the presence of proanthocyanidins or
condensed tannins would be possible. Those images were taken from the one of the ten
different solvent systems (data not showed) that displayed a better TLC resolution.
Further analyses are necessary to corroborate this hypothesis.
Fig. 3. TLC plates of the ethanolic extract. a, exposed to 254 nm
UV light; b, sprayed with sulfuric acid-vanillin reagent; c,
sprayed with 0,2 % DPPH solution in methanol
Ferric-reducing power assay
The reducing power of a compound is related to its electron transfer ability and may
serve as a significant indicator of its potential antioxidant activity. In this assay, the
yellow color of the test solution changes to green and blue depending on the reducing
power of test specimen. Greater absorbance at 700 nm indicates higher reducing power.
In the concentration range investigated, the extract demonstrated reducing power that
increased linearly with concentration. Significant changes in absorbance at 700 nm were
observed (0,43–0,63) with increasing concentrations of extract (10–50 µg/mL) (fig. 4,
b). The reducing power of the C. cowellii leaves might be due to its hydrogen-donating
ability.
The high phenolic and flavonoid content is possibly responsible for the antioxidant
activity of thisextract. Flavonoids are highly effective scavengers of most oxidizing
molecules, including singlet oxygen, and various other free radicals implicated in
a
b
c

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several diseases [24]. Flavonoids suppress reactive oxygen formation, chelate trace
elements involved in free-radical production, scavenge reactive species and up-regulate
and protect antioxidant defenses [25]. Similarly, phenolics compounds confer oxidative
stress tolerance on plants. Crude extracts of fruits, herbs, vegetables, cereals and other
plant materials rich in phenolics are increasingly being used by the food industry for
their antioxidative properties and health benefits.
Fig. 4. (a) Free radical scavenging activity and (b) ferrous reducing capacity of C. cowelli leaves
ethanolic extract. Ascorbic acid was included as a positive control. Each value is the mean ±
standard deviation of three replicates
Conclusions
In vitro antioxidant activities of C. cowelliiare consistent with the presence of
phenolic compounds, like flavonoids and tannins, and the results are similar to other
species of the Coccoloba genus. Results suggested that C. cowellii is a potential
source of antioxidant agents, possibly due to the presence of phenolic compounds.
Therefore, it may be considered a viable specimen for vegetal biotechnology
techniques of in vitro propagation. Nevertheless, more definitive phytochemical
analysis must be required to isolate and to characterize plant metabolites that show
the pharmacological effects. These results constitute the first report of the
phytochemical composition and potential antioxidant activity of C. cowellii.
Acknowledgements
Authors are grateful to Isidro Méndez and Andrys Martínez from “Centro de Estudio de
Manejo Ambiental” of Faculty of Applied Sciences of the University of Camagüey for

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Dr. C Julio César Escalona-Arranz, Dr. C Paul Cos
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support in plant collection and, to Humberto Morris and Orlando Abreu for suggestions
in manuscript writing.
This study was supported in part by the Belgian Development Cooperation through
VLIR-UOS (Flemish Interuniversity Council-University Cooperation for Development)
in the context of the TEAM Project “Installing a center of excellence in the Central-
Eastern region of Cuba to enhance production and research on bioactive plants” with the
University of Camagüey and the Institutional University Cooperation Program with
University of Oriente, especially by means of the P-3 project “Biopharmaceutical
Products from Natural Sources in the Development of Biotechnology”.
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