CyTA - Journal of Food

ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/tcyt20

Simultaneous consumption of green and black

tea infusions from Cnidoscolus aconitifolius leaves
with metformin treatment improves the health
outcome in type II diabetic rats

Al-Mansour Mahammad, Florian Amel Tekou, Cerile Ypolyte Woumbo,

Michel Peguy Kentsop, Vanessa Djuine & Dieudonné Kuate

To cite this article: Al-Mansour Mahammad, Florian Amel Tekou, Cerile Ypolyte Woumbo,
Michel Peguy Kentsop, Vanessa Djuine & Dieudonné Kuate (2023) Simultaneous consumption
of green and black tea infusions from Cnidoscolus aconitifolius leaves with metformin treatment
improves the health outcome in type II diabetic rats, CyTA - Journal of Food, 21:1, 386-393, DOI:
10.1080/19476337.2023.2208193

To link to this article: https://doi.org/10.1080/19476337.2023.2208193

© 2023 The Author(s). Published with

license by Taylor & Francis Group, LLC.

Published online: 10 May 2023.

Submit your article to this journal

View related articles

View Crossmark data

Full Terms & Conditions of access and use can be found at

https://www.tandfonline.com/action/journalInformation?journalCode=tcyt20
CYTA – JOURNAL OF FOOD
2023, VOL. 21, NO. 1, 386–393
https://doi.org/10.1080/19476337.2023.2208193

Simultaneous consumption of green and black tea infusions from Cnidoscolus

aconitifolius leaves with metformin treatment improves the health outcome in
type II diabetic rats
Al-Mansour Mahammad, Florian Amel Tekou, Cerile Ypolyte Woumbo, Michel Peguy Kentsop, Vanessa Djuine
and Dieudonné Kuate
Faculty of Sciences, Department of Biochemistry, Laboratory of Biochemistry of Medicinal Plants, Food Sciences and Nutrition, University of
Dschang, Dschang, Cameroon

ABSTRACT ARTICLE HISTORY

The efficacy of concomitant administration of metformin and Cnidoscolus aconitifolius (Chaya) teas Received 11 November 2022
in type 2 diabetic rat models was evaluated. Diabetic rats were treated for 28 days with metformin Accepted 25 April 2023
and infusions of green (GTC) and black (BTC) chaya teas. Total phenol and flavonoid contents, as the KEYWORDS
antioxidant capacities of tea infusions, were assessed. Fasting blood glucose (FBG), lipid profile and Cnidoscolus aconitifolius; tea;
markers of hepatic and renal functions were assessed. GTC exhibited the highest phenolic and antioxidant; hypoglycemic;
flavonoid contents, as the best antioxidant activities. Daily co-administration of GTC and BTC with hypolipidemic; type 2
metformin significantly (p < .05) lowered the FBG, triglyceride, total cholesterol, LDL cholesterol, ALT, diabetes
AST, creatinine and urea, as well as a significant increase in the serum protein. The combination of
Chaya tea infusions with metformin treatment performed better than metformin taken alone in
regulating fasting blood glucose in diabetic rats, with the best results observed in the group
receiving GTC infusion.

1. Introduction including reduction of circulating lipids and increased antiox­

Type 2 diabetes mellitus is the most common form of dia­
betes mellitus and is largely the result of overweight and
sedentary lifestyle (Potier, 2014). More than 95% of diabetic
patients have type 2 diabetes (World Health Organisation
[WHO], 2022). The International Diabetes Federation (IDF)
estimated its global prevalence at 10.5% in 2021. This pre­
valence could rise to 11.3% in 2030 (International Diabetes
Federation [IDF], 2021). Estimated at 24 million in 2021, the
number of diabetic patients in sub-Saharan Africa will
increase to 55 million in 2045, an increase of 134%
(FID, 2021). Many strategies have been put in place to man­
age this disease, including oral antidiabetic drugs and low-
calorie diets. Low-calorie diets are slow to produce desired
results, while drugs, in addition to their side effects, are
expensive (Kuate et al., 2015). Furthermore, epidemiological
studies and clinical trials have shown that dietary vegetable
consumption is correlated with a reduced risk of developing
chronic diseases such as cancer, type 2 diabetes and others
(Woumbo et al., 2017). In the same line, several studies have
been carried out on Cnidoscolus aconitifolius, an
Euphorbiaceae used as a food and medicinal plant (Iyke
et al., 2018). It is rich in nutritional compounds and minerals
and contains dietary fiber, vitamins A and C, flavonoids and
polyphenols (Perez et al., 2019). Roy et al. (2016) also demon­
strated its in vitro inhibitory activity on α-glucosidase and α-
amylase enzymes and antidiabetic potential in type 2 diabetic
Wistar rats induced by streptozotocin. Studies have demon­
strated beneficial effects of the C. aconitifolius drinks,
idant activity (Guevara-Cruz et al., 2021). Previous studies have
established that drinks made from C. aconitifolius only had
moderate activity and could not therefore be taken alone for
an effective management of type 2 diabetes mellitus, but no
previous work has evaluated the synergistic activity of
C. aconitifolius drinks taken in combination with a reference
oral antidiabetic drug. The present work was aimed at evalu­
ating the efficacy of the concomitant administration of met­
formin and Chaya teas in type 2 diabetic rats.
2. Materials and methods
2.1. Plant material
The leaves of C. aconitifolius, were harvested in Dschang (5°
27′ north, 10° 04′ east), West region of Cameroon, identified at
the National Herbarium of Yaoundé under the voucher N°
6748/HNC. The leaves were transported to the Laboratory of
Biochemistry of Medicinal Plants, Food Sciences and Nutrition,
Department of Biochemistry of the University of Dschang
where they were sorted, washed and divided into several
portions to obtain the different teas (green and black).
2.2. Tea production
For obtaining green tea, fresh leaves were first withered for
18 h. After withering, they were subjected to a high tem­
perature (70°C) for 1 h. Dried leaves were ground using an
electric grinder (Gaon, 800 W, 5 cycles of 1 min each at full

CONTACT Cerile Ypolyte Woumbo woumbocerile@yahoo.fr Faculty of Sciences, Department of Biochemistry, Laboratory of Biochemistry of Medicinal
Plants, Food Sciences and Nutrition, University of Dschang, Dschang, Cameroon
© 2023 The Author(s). Published with license by Taylor & Francis Group, LLC.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly cited. The terms on which this article has been published allow the posting of the Accepted
Manuscript in a repository by the author(s) or with their consent.

power), sieved (50 μm mesh sieve) and stored in plastic bags
for further use. To obtain black tea, the leaves were withered
for 18 h, rolled so as to break the cells they contain and thus
facilitate oxidation. They were then allowed to rest for 2
h before they were air-dried using an air oven (70°C for 1
h). The dried leaves were treated as per the same protocol
described for green tea production.
Then, 2 g of tea powders (“green” or “black”) was respec­
tively infused (4 min) in 100 mL of boiling water. The green
tea yielded a pale yellow to orange brew, while black tea will
yielded a darker colored tea ranging from amber to dark
brown tones.
2.3. Determination of phenolic composition and
antioxidant properties
2.3.1. Total phenol content
The total phenol content (TPC) of green and black tea infu­
sions of C. aconitifolius was determined using the Folin–
Ciocalteu method as described by Dohou et al. (2003).
Distilled water (1.39 mL) and 0.2 mL of a tenfold diluted
Folin–Ciocalteu’s reagent were added to 0.01 mL of tea infu­
sion (2 mg/mL) and left for 3 min, 0.4 mL of 20% Na2CO3 was
added and the mixture vortexed and incubated for 20 min at
40°C using a water bath, then the absorbance was read
against a blank at 760 nm using a BIOMATE spectrophot­
ometer. Quantification of total phenols was made according
to a linear calibration curve (y = 0.044x) produced by
a standard (gallic acid) at different concentrations under the
same conditions as the sample.
2.3.2. Flavonoid content
The total flavonoid content (TFC) of green and black tea
infusions was determined according to the method described
by Woumbo et al. (2022). Distilled water (1.4 mL) and 0.03 mL
of a 5% sodium nitrite (NaNO2) solution were added to 0.1 mL
of tea infusion (2 mg/mL). Then, 0.2 mL of a 10% aluminum
trichloride (AlCl3) solution was added. After resting for 5 min,
0.2 mL of a 10% sodium hydroxide solution (NaOH) and 0.24
mL of distilled water were, respectively, added. The mixture
was vortexed before the absorbance was read against a blank
at 510 nm using a spectrophotometer. Quantification of fla­
vonoids was made according to a linear calibration curve (y =
0.319x) produced by a reference standard (catechin).
2.3.3. Antioxidant activity
● 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical
scavenging activity
The protocol established by Mensor et al. (2001) was used to
assess the scavenging activity of the teas. Briefly, 100 μl of
tea infusion was added to 900 μl of DPPH reagent (0.3 mM
DPPH (2,2-Diphenyl-1-picrylhydrazyl) dissolved in methanol).
After 30 min of incubation at room temperature, the absor­
bance was read at 517 nm against a blank; butylated hydro­
xytoluene (BHT) was used as a standard.
● Ferrous ion reducing capacity (FRAP)
The ability of the tea to reduce the ferric iron was assayed
according to the protocol established by Oyaizu (1986).
Briefly, 100 μL of tea infusion was added to 500 μL of
a phosphate buffer solution (0.2 M; pH 6.6), followed by
500 μL of aqueous solution of potassium hexacyanoferrate.
After 30 min of incubation at 50°C in a water bath, 500 μL of
CYTA - JOURNAL OF FOOD 387
trichloroacetic acid solution (10% v/v) was added.
Absorbance was read at 700 nm against a blank; Vitamin
C was used as the standard.
2.4. Induction of obesity and diabetes in experimental
animals
Three-week-old Wistar rats were obtained from the animal
facility of the department of Biochemistry of the University
of Dschang and maintained according to the OECD (2008)
guidelines. Obesity was induced using a high-fat high-
sucrose diet over a period of 12 weeks. The test groups
and the positive control (untreated diabetic rats, T+)
received a diet containing corn meal (52.9%), fish meal
(20%), beef tallow (17.6%), bones (1%), vitamins (0.5%), salt
(1%) and sucrose (7%), while the negative control (healthy
rats, T-) received a staple diet consisting of corn flour
(77.8%), fish meal (20%), bone (0.1%), palm olein (1%), vita­
mins (0.1%), salt (1%). Animals fed on hypercaloric diet had
free access to a 2% sucrose solution and those with a body
mass index (BMI) ≥0.68 g/cm2 were considered obese
(Novelli et al., 2007). These obese animals received a single
intraperitoneal dose of 50 mg/kg of streptozotocin solution
(dissolved in citrate buffer; 0.1 M, pH 4.5) with the aim of
inducing diabetes. Two days after injection of streptozoto­
cin, animals with a fasting blood glucose level above 2 g/L
were considered diabetic (Temiz, 2021).
The treatment lasted 28 days, and the animals were
grouped according to the treatments received as follows:
MET (metformin), MET + GTC (metformin + green tea), MET
+ BTC (metformin + black tea), T- (healthy control) and T+
(untreated diabetic rats). T- and T+ received water. Each
group was made up of six rats, individually housed, which
had free access to food and water. The treatments were
administered once daily using an esophageal tube. The
various “teas” of C. acotinifolius were administered at
a dose of 6 mL/kg, while metformin was administered at
250 mg/kg. All experiments were performed in accordance
with the regulations of the Ethics and Welfare Committee for
Experimental Animals.
2.5. Biological parameters: blood sugar and serum
lipids, transaminases, total protein, urea and creatinine
Animal blood glucose concentrations were measured, after
overnight fasting (8 h) on 5–10 μL of blood obtained from
the tail tip after, using a portable glucometer (Accu-Chek)
every week during the treatment according to the standard
protocols described by Trinder (1969).
The lipid profile was determined by colorimetric methods
using Sigma Diagnostic kits. The triglycerides were deter­
mined according to the method of Fossati and Principe
(1982). Total cholesterol was determined as per the protocol
established by Huang et al. (1997) using SIGMA Diagnostics’
EZ HDLTM cholesterol reagent, while LDL cholesterol was
estimated using the formula established by Friedewald et al.
(1972).
Colorimetric methods (Randox kits) were also used to
determine the serum level of alanine aminotransferase and
aspartate aminotransferase. The standard protocol described
by Gornall et al. (1949) was used to determine total protein
as well as Bartels et al. (1972) for creatinine and Marshall
(1913) for urea.
388 A.-M. MAHAMMAD ET AL.

2.6. Statistical analysis
Statistical analysis was performed using XLSTAT 2016 soft­
ware. Results were expressed as mean ± standard deviation.
A one-way analysis of variance (ANOVA) with Tukey’s test
was used for the statistical analysis of the mean difference
between the groups. The differences were considered sig­
nificant at p < .05.
3. Results and discussion
3.1. Total phenol and flavonoid contents and
antioxidant capacity
3.1.1. Total phenols and flavonoid contents
Figures 1 and 2, respectively, show the total phenol and
flavonoid contents of GTC and BTC. Green tea (GTC) pre­
sented the highest phenol content (140.13 mg EAG/g). The
same observation was made with the total flavonoid content
where GTC had 5.82 mg CE/g and BTC had 5.22 mg CE/g.
Such observations can be explained by the methods of tea
production. Hot water used in tea production should have
contributed to breaking down the membranes, permitting
a better diffusion of bioactive compounds from the biologi­
cal matrix to the solvent, thus explaining the high content
obtained with the different teas in general (Woumbo et al.,
2021). However, low TPC and TFC from BTC compared to
145
b gen atoms to DPPH (O. Ojo et al., 2014). We also observed
Total phenolic content (mg
GTC could be due to the implication of these molecules in
the enzymatic browning occurring during the fermentation
step or to other non-enzymatic processes like the microbial
activity (Zhao et al., 2019). These results corroborate with
those of Veljković et al. (2013) who reported high phenols
and flavonoids content in green tea infusions of herbs and
fruits consumed in Serbia.
3.1.2. Antioxidant capacity
Figure 3 shows the DDPH radical scavenging of the tea
infusions at different concentrations compared to vitamin
C. GTC exhibited the highest antioxidant activity compared
to black tea at concentrations ranging from 50 to 200 µg/mL.
The FRAP assay has been reported to serve as
a meaningful indicator of a plant extract antioxidant activity
(A. Ojo et al., 2019). Figure 4 presents the variations of the
iron reducing power of GTC and BTC at different concentra­
tions compared to butylhydroxytoluene (BHT). It also came
out that GTC exhibited the best reducing power.
Figure 5 presents the hydroxyl radical inhibitory activities
of the different C. aconitifolius tea infusions at different con­
centrations compared to that of BHT. The highest scavenging
ability was observed with GTC at a concentration of 200
µg/mL.
For all the tests used to assess the antioxidant capacity of
the teas, GTC exhibited the highest activity. This may be
associated with its high content of bioactive compounds
such as phenols and flavonoids, which may be able to
neutralize the free radical by transferring electrons or hydro­

the same trends regarding reducing power and phenolic

140
content; this difference could be explained by the presence
of phenolic compounds in our different samples as shown
GAE/g)

135 by Katalinic et al. (2006).

The ferrous reducing capacity could also be explained by
the presence of electron donor compounds which lead to the
a
130 reduction of Fe3+ to Fe2+. This would be due to their respec­
tive contents of total polyphenols and total flavonoids.
Indeed, many studies attribute the reducing power to pheno­
125
lic compounds (Fabri et al., 2009; H. Li & Wang, 2009). This
property may provide protection to affected organs through
120 the ability of the plant extract to scavenge harmful metabo­
GTC BTC lites released during disease states (Ajiboye et al., 2016).

Figure 1. Total phenol content (TPT) of C. aconitifolius tea infusions.

GTC: Green tea, BTC: Black tea. a and b: Bands with different letters are significantly 3.2. Evolution of the fasting blood glucose
different at p < .05.
Table 1 presents the changes in the fasting blood glucose
(FBG) concentration in the different groups throughout the
6 treatment period. A significant reduction of the FBG was
Total flavonoid content (mg CE/g)

b
observed in all groups, except untreated diabetic rats (T+)
5.8
which experienced a continuous increase in their FBG during
5.6 the experiment. The concomitant intake of metformin and the
different teas of C. aconitifolius significantly decreased the FBG
5.4 of the diabetic rats than treatment with metformin alone.
a
That reduction can be attributed to the presence of phenolic
5.2
compounds in our different samples since polyphenols are
5
able to reduce glycaemia by many mechanisms including an
increase in insulin sensitivity. R. Li et al. (2006) postulated that
4.8 green tea catechins could ameliorate insulin resistance by
GTC BTC acting as ligands for peroxisome proliferator-activated recep­
tors with a dual alpha/gamma agonist effect. It was also
Figure 2. Total flavonoid (TFT) content of C. aconitifolius tea infusions.
GTC: Green tea, BTC: Black tea. a and b: Bands with different letters are significantly
reported that green tea phytochemicals can prevent intestinal
different at p < .05. glucose absorption by inhibiting the sodium-dependent
 CYTA - JOURNAL OF FOOD 389

100

Percentage of inhibition (%)

90
80
70
60
50 GTC
40
BTC
30
20 Vit C
10
0
12.5 25 50 100 200
Concentration (µg/mL)
Figure 3. Evolution of the antiradical activity of C. aconitifolius tea infusions at different concentrations compared to vitamin C. GTC: Green tea, BTC: Black tea,
Vit C: Vitamin C.

0.12
Absorbance (700 nm)

0.10

0.08

0.06 BHT
GTC
0.04
BTC

0.02

0.00
12.5 25 50 100 200
Concentration (µg/mL)

Figure 4. Evolution of the reducing power of C. aconitifolius tea infusions compared to BHT.

0.250
Optical density (560 nm)

0.200

0.150
BHT

0.100 GTC

BTC
0.050

0.000
12.5 25 50 100 200

Concentration (µg/mL)

Figure 5. Changes in the hydroxyl radical inhibitory capacity of tea infusion at different concentrations compared to that of BHT.

Table 1. Changes in the FBG (g/L) during the treatment.

Groups Day 0 Day 7 Day 14 Day 21 Day28 Difference
T- 0.81 ± 0.4ɑ 0.91 ± 0.3ɑ 0.86 ± 0.2ɑ 0.83 ± 0.3ɑ 0.90 ± 0.2ɑ 0.09a
T+ 3.62 ± 0.4c 3.88 ± 0.2d 4.06 ± 0.4e 4.86 ± 0.4d 5.1 ± 0.7d 1.48b
Met 3.37 ± 0.5b 3.09 ± 0.4c 2.61 ± 0.6d 2.10 ± 0.5e 1.65 ± 0.32c −1.72c
Met + GTC 3.68 ± 0.5b 2.93 ± 0.5b 2.31 ± 0.3b 1.74 ± 0.4b 1.29 ± 0.38b −2.39d
Met + BTC 3.81 ± 0.7b 3.01 ± 0.3bc 2.43 ± 0.6c 1.77 ± 0.5b 1.33 ± 0.28b −2.48d
T-: Healthy control; T+: Untreated diabetics; MET: Diabetics treated with metformin; Met +GTC: Diabetics treated with metformin and green
tea; Met +BTC: Diabetics treated with metformin and black tea. Difference = Day 28 – Day 0. a, b and c: In the same column, values with
different letters differ significantly (p < 0.05).
390 A.-M. MAHAMMAD ET AL.

glucose transporter in rabbit intestinal epithelial cells (Toolsee present in the leaves of C. aconitifolius (Oluwatobi et al.,
et al., 2013). Combining this effect with the hypoglycaemic 2020). In addition, the increase of HDL cholesterol concen­
activity of metformin explains the better results obtained with trations in animals receiving tea infusions could be related
the combined treatment over the sole use of metformin. to their polyphenolic content and antioxidant capacity as
stated by Asante et al. (2016) who showed that phenolic
compounds have the ability to increase serum HDL choles­
terol levels. The drop in the LDL level in the animals receiv­
3.3. Effect of treatments on the lipid profile
ing chaya “teas” could be explained by the presence (in
Figure 6a–d shows the serum levels of triglycerides, total high quantities) of compounds with a strong hypocholes­
cholesterol, HDL-cholesterol and LDL-cholesterol, respec­ terolemic and hypolipidemic power, in particular phenolic
tively, of the animals after 28 days of treatment. The posi­ compounds. The drop in LDL levels is in agreement with
tive control group (untreated diabetics) showed those of Guevara-Cruz et al. (2021) who showed that chaya
a significant increase in triglyceride, total cholesterol and leaves lead to a drop in LDL levels in subjects with
LDL cholesterol levels compared to the healthy control dyslipidemia.
group. Diabetes mellitus in adults is often associated with
hyperlipidemia, either due to inadequate secretion or insu­
lin resistance. It generally results in an increase in the
3.4. Effect of treatments on serum transaminases and
activity of lipases, which is responsible for disorders of
total proteins
lipid metabolism. It is well known that hyperlipidemia is
accepted as a risk factor for cardiovascular disorders (Garg The liver is the central organ involved in metabolic pro­
& Grundy, 1990). cesses in the body and is affected by the complications of
In this study, the treatment of animals by daily adminis­ diabetes, as increased transaminase activities have been
tration of different teas (green and black teas) of shown to cause liver damage (Hultcrantz et al., 1986).
C. aconitifolius showed a significant decrease in the serum Figure 7a–d shows the serum level of alanine aminotrans­
level of triglycerides, total cholesterol and LDL cholesterol, ferase, aspartate aminotransferase, alkaline phosphatase
compared to that of the diabetic control group. This and total protein of the animals after 28 days of treatment.
decrease could be explained by an inhibition of hepatic A significant decrease in the level of serum transaminases
synthesis of cholesterol by the phenolic compounds and alkaline phosphatase was observed in this work

160 600

140 c
500 c
Total cholesterol (mg/dL)
Triglycerides (mg/dL)

120 b
b 400
100 b b
a b b
80 300 a
60
200
40

20 100

0
0
T- T+ Met Met + GTC Met + BTC
T- T+ Met Met + GTC Met + BTC

90
c 70
80 d
bc
HDL-cholesterol (mg/dL)

70 bc 60
LDL-cholesterol (mg/dL)

60 b 50

50 a c
40 b
b
40
30 a
30
20
20
10 10

0 0
T- T+ Met Met + GTC Met + BTC T- T+ Met Met + GTC Met + BTC

Figure 6. Serum concentration of triglycerides, total cholesterol, HDL cholesterol and LDL cholesterol in animals after 28 days of treatment. T (-): Healthy
control; T (+): untreated diabetics; MET: Metformin; GTC: Green tea; BTC: Black tea. a, b, c and d: On the same graph, bands with different letters differ
significantly (p < .05).
 CYTA - JOURNAL OF FOOD 391

3500

3000 c 3000
bc c
bc 2500
2500 b
bc b b

AST (U/mL)
ALT (U/mL)

2000 2000
a a
1500 1500

1000 1000

500 500

0 0
T- T+ Met Met + GTC Met + BTC T- T+ Met Met + GTC Met + BTC
200
c 30
180
c
160 b 25 b
b
b b b

Total proteins (g/dL)

140
20
ALP (U/I)

120

100 a
15
80 a
60 10

40
5
20

0 0
T- T+ Met Met + GTC Met + BTC T(-) T(+) Met Met + GTC Met + BTC

Figure 7. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total protein after 28 days of
treatment. T (-): Healthy control; T (+): untreated diabetics; MET: Metformin; GTC: Green tea; BTC: Black tea. a, b and c: On the same graph, bands with different
letters differ significantly (p < .05).

compared to untreated diabetics, which could reflect that
the various “teas” of C. aconitifolius would correct the dis­
orders induced by diabetes in rats, this by regulating altera­
tions in liver enzymes and protecting cell membrane
permeability or avoiding liver cell rupture (Maria & Maira,
2020). These results are in agreement with Chukwu et al.
(2020) who reported that the aqueous and ethanolic
extracts of the leaves of C. aconitifolius could have dose-
dependent hepatoprotective potentials. Untreated animals
had significantly elevated serum ALT, AST and alkaline
phosphatase levels compared to healthy controls. An
increase in the activity of these enzymes reflects active
damage and inflammatory hepatocellular disorders
(Hultcrantz et al., 1986). In addition, an increase in serum
protein levels observed in this work would confirm that
bioactive compounds present in various “teas” of
C. aconitifolius improve diabetes-related disorders. Raza
and John (2007) reported that tea catechins can prevent
protein degradation by altering subcellular reactive oxygen
species production, glutathione metabolism and cyto­
chrome P450 2E1 activity.
3.5. Effect of treatments on serum urea and creatinine
Figure 8a,b shows serum urea and creatinine concentrations
of animals after 28 days of treatment. An increase in the
creatinine and urea levels in untreated diabetic animals
was observed. This could be linked to renal dysfunction in
untreated diabetics, which is due to the nephrotoxicity of
streptozotocin.
In this work, a significant decrease in the level of urea
and serum creatinine was observed in the animals having
received the “teas” of C. aconitifolius leaves. This could be
justified by the high content of bioactive compounds, in
particular phenolic compounds. The present results corro­
borate those of Oyagbemi & Odetola (2013) who showed
that C. aconitifolius has a nephroprotective effect. No sig­
nificant difference was observed between animals treated
with green tea and black tea from C. aconitifolius.
4. Conclusion
Simultaneous intake of green and black teas produced
from C. aconitifolius leaves with metformin treatment bet­
ter improved the health outcomes of type 2 diabetic rats
than the treatment with metformin alone. There was
a significant decrease in the fasting blood glucose, total
and LDL cholesterol and triglyceride levels of diabetic rats
after 28 days of treatment compared to animals treated
with metformin alone. Green and black tea infusions
decreased serum triglycerides, total and LDL cholesterol,
ALT, AST, creatinine and urea concentration in the serum
392 A.-M. MAHAMMAD ET AL.
450
c
400
350
300 b 4
Urea (mg/dL)
b
250
ab
200
150 a 2
100
50
0 0
T- T+ Met Met + GTC Met + BTC
Figure 8. Serum urea and creatinine concentrations in animals after 28 days of treatment. T (-): Healthy control; T (+): untreated diabetics; MET: Metformin; GTC:
Green tea; BTC: Black tea. a and b: On the same graph, bands with different letters differ significantly (p < .05).
of diabetic rats. Furthermore, their consumption also
increased the HDL cholesterol and the total protein con­
centrations. Consumption of black and green teas made
from C. aconitifolius leaves potentiates the antidiabetic
activity of metformin.
Disclosure statement
No potential conflict of interest was reported by the author(s).
Funding
This research did not receive any specific grant from funding agencies in
the public, commercial, or not-for-profit sectors.
Authors contribution
Woumbo Cerile Ypolyte conceived the work, supervised data analy­
sis and interpretation and read the paper. Mahammad Al-Mansour,
Tekou Florian Armel, Kentsop Michel Peguy and Djuine Vanessa
collected leaves samples, produced tea infusions, carried out experi­
mentations, analyzed and interpreted data and wrote the paper.
Kuate Dieudonné supervised the work and read the paper. All
authors have approved the final article.
References
Ajiboye, B., Ojo, O., Adeyonu, O., Imiere, O., Olayide, I., Fadaka, A., &
Oyinloye, B. (2016). Inhibitory effect on key enzymes relevant to
acute type-2 diabetes and antioxidative activity of ethanolic extract
of Artocarpus heterophyllus stem bark. Journal of Acute Disease, 5(5),
423–429. https://doi.org/10.1016/j.joad.2016.08.011
Asante, D. B., Effah-Yeboah, E., Barnes, P., Abban, H. A., Ameyaw, E. O.,
Boampong, J. N., Ofori, E. G., & Dadzie, J. B. (2016). Antidiabetic effect
of young and old ethanolic leaf extracts of Vernonia amygdalina:
A comparative study. Journal of Diabetes Research, 2016, 8252741.
https://doi.org/10.1155/2016/8252741
Bartels, H., Böhmer, M., & Heierli, C. (1972). Serum creatinine determina­
tion without protein precipitation. Clinica Chimica Acta, 37, 193–197.
https://doi.org/10.1016/0009-8981(72)90432-9
Chukwu, E., Osuocha, K., & Iwueke, A. (2020). Phytochemical profiling,
body weight effect and anti-hypercholesterolemia potentials of
Cnidoscolus aconitifolius leaf extracts in male albino rat. Journal of
Pharmacognosy and Phytotherapy, 12(2), 19–27. https://doi.org/10.
5897/JPP2016.0436
Dohou, N., Yamni, K., Tahrouch, S., Hassani, L. M. I., Badoc, A., & Gmira, N.
(2003). Screening of a phytochemical endemic ibero-Moroccan
Thymelaea lythroides. Bulletin de la Société de Pharmacie, 142, 61–78.
6
5 c
Creatinine (mg/dL)
b b b
3
a
1
T(-) T(+) Met Met + GTC Met + BTC
Fabri, R., Nogueira, M., Braga, F. G., Coimbra, E. S., & Scio, E. (2009).
Mitracarpus frigidus aerial parts exhibited potent antimicrobial, antil­
eishmanial, and antioxidant effects. Bioresource Technology, 100(1),
428–433. https://doi.org/10.1016/j.biortech.2008.05.053
Fossati, P., & Principe, L. (1982). Serum triacylglycerols determined color­
imetrically with an enzyme that produces hydrogen peroxide. Clinical
Chemistry, 28(10), 2077–2080. https://doi.org/10.1093/clinchem/28.
10.2077
Friedewald, W., Levy, R., & Frederickson, D. (1972). Estimation of the
concentration of low-density lipoprotein cholesterol in plasma, with­
out use of the preparative ultracentrifuge. Clinical Chemistry, 18(6),
499–502. https://doi.org/10.1093/clinchem/18.6.499
Garg, A., & Grundy, S. M. (1990). Management of dyslipidemia in NIDDM.
Diabetes Care, 13(2), 153–169. https://doi.org/10.2337/diacare.13.2.153
Gornall, A., Bardawill, C., & David, M. (1949). Determination of serum
proteins by means of the biuret reaction. The Journal of Biological
Chemistry, 177(2), 751–766. https://doi.org/10.1016/S0021-9258(18)
57021-6
Guevara-Cruz, M., Medina-Vera, I., Cu-Cañetas, T. E., Cordero-Chan, Y.,
Torres, N., Tovar, A. R., Márquez-Mota, C., Talamantes-Gómez, J. M.,
Pérez-Monter, C., Lugo, R., Gutiérrez-Solis, A. L., & Avila-Nava, A.
(2021). Chaya leaf decreased triglycerides and improved oxidative
stress in subjects with dyslipidemia. Frontiers in Nutrition, 8, 666243.
https://doi.org/10.3389/fnut.2021.666243
Huang, Y. C., Kao, J. T., & Tsai, K. S. (1997). Evaluation of two homo­
geneous methods for measuring high-density lipoprotein cholesterol.
Clinical Chemistry, 43(6 Pt 1), 1048–1055. https://doi.org/10.1093/clin
chem/43.6.1048
Hultcrantz, R., Glaumann, H., Lindberg, G., & Nilsson, L. H. (1986). Liver
investigation in 149 asymptomatic patients with moderately elevated
activities of serum aminotransferases. Scandinavian Journal of
Gastroenterology, 21(1), 109–113. https://doi.org/10.3109/
00365528609034632
International Diabetes Federation. (2021). The IDF diabetes atlas 10th
edition 2021. Retrieved February 9, 2022, form www.diabetesatlas.
org/fr/.
Iyke, W., Nwafor, A., Njoku, B., Nwoke, K., & Deebii, N. (2018).
Investigation of phytocomponents and hypoglycaemic effect of
hydro-methanolic leaf extract of Cnidoscolus aconitifolius (Spinach
Tree) in streptozotocin induced-diabetic Wistar rats. Journal of
Complementary and Alternative Medical Research, 5(3), 1–9. https://
doi.org/10.9734/JOCAMR/2018/26858
Katalinic, V., Milos, M., & Jukic, M. (2006). Screening of 70 medicinal plant
extracts for antioxidant capacity and total phenols. Food Chemistry,
94(4), 550–557. https://doi.org/10.1016/j.foodchem.2004.12.004
Kuate, D., Kengne, A., Biapa, C., Azantsa, B., & Muda, W. (2015).
Tetrapleura tetraptera spice attenuates high-carbohydrate, high-fat
diet-induced obese and type 2 diabetic rats with metabolic syn­
drome features. Lipids in Health and Diseases, 14(1), 50. https://doi.
org/10.1186/s12944-015-0051-0
Li, R., Douglas, T., Maiyoh, G., Adeli, K., & Theriault, A. (2006). Green tea
leaf extract improves lipid and glucose homeostasis in a fructose-fed

insulin-resistant hamster model. Journal of Ethnopharmacology, 104
(1–2), 24–31. https://doi.org/10.1016/j.jep.2005.08.045
Li, H., & Wang, X. (2009). Polyphenolic compounds and antioxidant
properties of selected China wines. Food Chemistry, 112(2), 454–460.
https://doi.org/10.1016/j.foodchem.2008.05.111
Maria, L., & Maira, R. (2020). Renal and hepatic disease: Cnidoscolus
aconitifolius as diet therapy proposal for prevention and treatment.
Journal of the American College of Nutrition. https://doi.org/10.1080/
07315724.2020.1810171
Marshall, E. (1913). A new method for the determination of urea in
blood. The Journal of Biological Chemistry, 15(3), 487. https://doi.org/
10.1016/S0021-9258(18)88512-X
Mensor, L., Menezez, F., Leitao, G., Reis, A., Dos Santos, T., Coube, C., &
Leitao, S. (2001). Screening of Brazilian plant extracts for antioxidant
activity by the use of DPPH free radical method. Phytotherapy
Research, 15(2), 127–130. https://doi.org/10.1002/ptr.687
Novelli, E., Diniz, Y., Galhardi, C., Ebaid, G., Rodrigues, H., Mani, F.,
Cicogna, A. C., Novelli Filho, J. L. V. B., & Fernandes, A. A. H. (2007).
Anthropometrical parameters and markers of obesity in rats.
Laboratory Animals, 41(1), 111–119. https://doi.org/10.1258/
002367707779399518
OECD. (2008). Guidelines for chemicals trials: Repeated dose oral toxicity
studies for 28 days in rodents. http://www.oecd-ilibrary.org.
Ojo, A., Ojo, O., Okesola, M., Ajiboye, B., & Oyinloye, B. (2019). Garcinia
kola extracts improve biochemical markers associated with erectile
function: Possible applications in clinical treatment? Acta Facultatis
Naissenis, 36(1), 15–26. https://doi.org/10.2478/afmnai-2019-0002
Ojo, O., Oloyede, O., Tugbobo, O., Olarewaju, O., & Ojo, A. (2014).
Antioxidant and inhibitory effect of scent leaf (Ocimum gratissimum)
on Fe2+ and sodium nitroprusside induced lipid peroxidation in rat
brain in vitro. Advances in Biological Research, 8(1), 8–17. https://doi.
org/10.5829/idosi.abr.2014.8.1.8145
Oluwatobi, T., Oluseyi, A., Regina, N., & Mopelola, A. (2020). Cnidoscolus
aconitifolius leaf extract exhibits comparable ameliorative potentials
with ascorbate in dimethylnitrosamine-induced bone marrow clasto­
genicity and hepatotoxicity. Clinical Nutrition Experimental, 29, 36–48.
https://doi.org/10.1016/j.yclnex.2019.11.003
Oyagbemi, A., & Odetola, A. (2013). Hepatoprotective and nephropro­
tective effects of Cnidoscolus aconitifolius in protein energy malnutri­
tion induced liver and kidney damage. Pharmacognosy Research, 5(4),
260. https://doi.org/10.4103/0974-8490.118817
Oyaizu, M. (1986). Studies on products of browning reaction.
Antioxidative activities of products of browning reaction prepared
from glucosamine. Japanese Journal of Nutrition, 44(6), 307–315.
https://doi.org/10.5264/eiyogakuzashi.44.307
Perez, Z., Macias, M., Hernandez, S., Siordia, A., & Jimenez, M. (2019).
Additional compounds and the therapeutic potential of Cnidoscolus
chayamansa (McVaugh) against hepatotoxicity induced by antituber­
cular drugs. Biomedicine and Pharmacotherapy, 117, 109140. https://
doi.org/10.1016/j.biopha.2019.109140
Potier, L. (2014). Role of the kallikrein-kinin system in diabetes and its
complications [Doctoral dissertation, Paris 6].
CYTA - JOURNAL OF FOOD 393
Raza, H., & John, A. (2007). In vitro protection of reactive oxygen
species-induced degradation of lipids, proteins and 2- deoxyribose
by tea catechins. Food and Chemical Toxicology, 45(10), 1814–1820.
https://doi.org/10.1016/j.fct.2007.03.017
Roy, D., Ferdioussi, N., Khatun, T., & Moral, M. (2016). Phytochemical
screening, nutritional profile and anti-diabetic effect of ethanolic
leaf extract of Cnidoscolus aconitifolius in streptozotocin induce
diabetic mice. International Journal of Basic & Clinical
Pharmacology, 5(5), 2244–2250. https://doi.org/10.18203/2319-
2003.ijbcp20163269
Temiz, M. (2021). Antioxidant and antihyperglycemic activities of
Scorzonera cinerea radical leaves in streptozotocin-induced diabetic
rats. Acta Pharmaceutica, 71(4), 603–617. https://doi.org/10.2478/
acph-2021-0045
Toolsee, N. A., Aruoma, O. I., Gunness, T. K., Kowlessur, S., Dambala, V.,
Murad, F., Googoolye, K., Daus, D., Indelicato, J., Rondeau, P.,
Bourdon, E., & Bahorun, T. (2013). Effectiveness of green tea in
a randomized human cohort: Relevance to diabetes and its
complications. BioMed Research International, 2013, 412379. https://
doi.org/10.1155/2013/412379
Trinder, P. (1969). Determination of glucose in blood using glucose oxidase
with alternative oxygen acceptor. Annals of Clinical Biochemistry, 6(1),
24–27. https://doi.org/10.1177/000456326900600108
Veljković, J. N., Pavlović, A., Mitic, S., Tošić, S., Stojanović, G., Kaličanin, B.
M., Stanković, D., Stojkovic, M., Mitić, M., & Brcanović, J. M. (2013).
Evaluation of individual phenolic compounds and antioxidant proper­
ties of black, green, herbal and fruit tea infusions consumed in Serbia:
Spectrophotometrical and electrochemical approaches. Journal of
Food and Nutrition Research, 52(1), 12–24.
World Health Organisation. (2022). Diabetes. Retrieved January 14, 2023,
from https://www.who.int/news-room/fact-sheets/detail/diabetes.
Woumbo, C., Kuate, D., Klang, M., & Womeni, H. (2021). Valorization of
Glycine max (Soybean) seed waste: Optimization of the
microwave-assisted extraction (MAE) and characterization of poly­
phenols from soybean meal using response surface methodology
(RSM). Journal of Chemistry, 2021, 12. https://doi.org/10.1155/2021/
4869909
Woumbo, C. Y., Kuate, D., Metue Tamo, D. G., & Womeni, H. M. (2022).
Antioxidant and antidiabetic activities of a polyphenol rich extract
obtained from Abelmoschus esculentus (okra) seeds using optimized
conditions in microwave-assisted extraction (MAE). Frontiers in
Nutrition, 9, 1030385. https://doi.org/10.3389/fnut.2022.1030385
Woumbo, C., Kuate, D., & Womeni, H. (2017). Cooking methods affect
phytochemical composition and anti-obesity potential of soybean
(Glycine max) seeds in Wistar rats. Heliyon, 3(8), e00382. https://doi.
org/10.1016/j.heliyon.2017.e00382
Zhao, C. N., Tang, G. Y., Cao, S. Y., Xu, X. Y., Gan, R. Y., Liu, Q., Mao, Q. Q.,
Shang, A., & Li, H. B. (2019). Phenolic profiles and antioxidant activ­
ities of 30 tea infusions from green, black, oolong, white, yellow and
dark teas. Antioxidants (Basel, Switzerland), 8(7), 215. https://doi.org/
10.3390/antiox8070215