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Original Article
EVALUATION OF ANTI-HYPERLIPIDEMIC ACTIVITY OF CAPSICUM FRUTESCENS EXTRACT
LICTO THOMAS
1
, JAGADISH VASUDEV KAMATH
2
1
Research Scholar, Bhagwant University, Ajmer, Rajasthan, India,
2
Department of Pharmacology, Shree Devi College of Pharmacy,
Mangalore, 574 142, Karnataka, India
Email: licto.thomas@gmail.com
Received: 04 Jan 2017, Revised and Accepted: 31 Apr 2017
ABSTRACT
Objective: The objective of the study was to evaluate the in-vivo anti-hyperlipidemic activity of Capsicum frutescens extracts.
Methods: The dried fruit powder were extracted with a three liquid phase extraction system. The acetone extract was isolated and the anti-
hyperlipidemic activity was evaluated.
Results: The anti-hyperlipidemic study was carried out by inducing hyperlipidemia in rats by means of triton. The serum collected was analyzed for
total cholesterol, triglyceride, low-density lipoprotein and high-density lipoprotein.
Conclusion: The result of the present study revealed that the acetone extract of the fruits of Capsicum frutescens possess anti-hyperlipidemic activity.
Keywords: Capsicum frutescens, Antihyperlipidemic activity, HDL-LDL, Triton
© 2017 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)
DOI: http://dx.doi.org/10.22159/ijcpr.2017v 9i4.21289
INTRODUCTION
Hyperlipidemia, also known as hyperlipoproteinemia or
dyslipidemia, is an abnormal elevation of lipid levels in the
bloodstream. These lipids include cholesterol, cholesterol
compounds, phospholipids and triglycerides, all carried in blood as
large molecules called lipoproteins. There are three types of
hyperlipidemia namely hyperlipoproteinemia (elevated levels of
lipoprotein in the blood), hypercholesterolemia (high cholesterol)
and hypertriglyceridemia (high triglycerides level in blood).
Hyperlipidemia affects lipid production, transportation in the
bloodstream and/or deposition in body cell. It is the first and foremost
factor that leads to diseases like atherosclerosis, coronary heart
disease, ischemic cerebrovascular disease, hypertension, obesity and
diabetes mellitus (Type-II) etc. [1, 2].
Although human beings have developed many allopathic drugs to
combat the hyperlipidemia and these drugs to a certain extent have
shown promising lipid-lowering activity. But these drugs have been
found to be associated with side effects. As a result of side effects
that are mostly associated with the allopathic system of medicine,
the world population today is turning back to a traditional system of
medicine as these medicines are free from side effects, easily
available with less cost. Thus, in the present scenario, more research
activities are required for the development of the drug from the
natural sources to meet the demand of world population.
Capsicum frutescens is a vegetable used daily, and the substance
capsaicin is responsible for its hot and spicy flavour, sought after in
gastronomy. Capsaicin and several related molecules are known by
the collective name capsaicinoids, and they are produced by all
plants of the genus Capsicum, with the exception of the bell pepper
(Capsicum annum), which produces no capsaicin.
The naturally occurring content of capsaicinoids in spices ranges
typically from 0.1 mg/g in chilli pepper to 2.5 mg/g in red pepper
and 60 mg/g in oleoresin red pepper [3]. Capsaicin and
dihydrocapsaicin are the major capsaicinoids produced; however,
others exist and are produced in smaller quantities.
Capsaicin is also known to inhibit Substance P, a neuropeptide that
is the key transmitter of pain to the brain thus providing relief to
pain. It has potent antibacterial properties that fight and prevent
chronic sinus infections, or sinusitis. Capsaicin is also a thermogenic
agent, which means it increases metabolic activity. This, in turn,
helps to burn calories and fat. Capsaicin may help to protect the
heart by reducing cholesterol, triglycerides and platelet aggregation.
Capsaicin is claimed to have high antioxidant values and HDL-
cholesterol-raising effect. Hence this study aims at exploring the
possible effects of Capsaicin on serum TC, TG, LDL and HDL levels in
experimental rats.
MATERIALS AND METHODS
Chemicals
Triton WR 1339 (Sigma-Aldrich), Cholesterol (SRL Mumbai),
Atorvastatin (Micro labs Pvt Ltd, B, lore), Anaesthetic ether-SD Fine-
chem Ltd., Mumbai, Chloroform-SD Fine Chem Ltd. Mumbai.
Formaline-SD Fine-chem Ltd., Mumbai. All chemicals and reagents
were of analytical grade. Diagnostic kits used for estimation of
cholesterol, triglycerides, HDL, LDL, VLDL were procured from
Robonik Diagnostic Ltd India.
Animals
Male Wistar rats aged between 8-10 w (250-300 g) were used for
the study. Animals were kept in the controlled condition in the
institutional animal house at an ambient temperature of 25-30 °C
and relative humidity of 55-60% and 12/12 h light/dark cycle and
were provided pellet diet al. ong with water ad libitum. The
experimental protocol was accepted by Institutional Animal Ethics
Committee.
Preparation of fruit extract
Mature Capsicum frutescens (bird’s eye chilli) were collected and
washed thoroughly with water and air dried in shade at room
temperature. It was ground into powder by a miniature high-speed
universal pulveriser. The three liquid phase extraction system of
acetone, K2HPO4 and n-hexane was prepared by weighing acetone
22% (w/w), K2HPO4 20% (w/w), n-hexane 10% (w/w) and water
(58% (w/w) and mixing. The powdered Capsicum frutescens was
added to this three liquid phase extraction system in a mass ratio of
1:20. The mixture was thoroughly vibrated for 10 min and then set-
tled at room temperature. After the separation of three phases, the
volume of the acetone was removed from the middle layer by using a
pipette and kept aside. This procedure was done again for three
International Journal of Current Pharmaceutical Research
ISSN- 0975-7066 Vol 9, Issue 4, 2017
Thomas et al.
Int J Curr Pharm Res, Vol 9, Issue 3, 165-168
166
times by adding fresh acetone into the same extract. The acetone
extracts were pooled and dried under vacuum. The percent yield of
capsaicin was 10 mg/g of Capsicum frutescens powder.
Electrospray mass spectrometry analysis
The electrospray mass spectrometric scans were monitored to check
for the presence of Capsaicinoids using AB Sciex 4000 mass
spectrometer with direct injection of sample dissolved in Methanol
in MS scan mode into the mass detector.
Dose preparation of standard and leaves extracts
Atorvastatin at a dose of 10 mg/Kg was prepared by suspending in
5% sucrose in water. The extract of leaves was dissolved in 5%
sucrose in water. Doses of 50 mg/Kg b.w. and 100 mg/Kg b.w. of the
extract were administered by oral route to the rats.
In vivo acute oral toxicity studies
In the present study, the acute oral toxicity of the extracts was
performed according to OECD Guideline 423 [4]. In this method, the
toxicity of the extracts was evaluated using the stepwise procedure,
each step using three Wistar rats. The rats were fasted prior to dosing
(food but not water should be withheld) for three to four hrs.
Following the period of fasting the animals were weighed and the
extract was administered orally at a dose of 1000 mg/Kg b.w. Animals
were observed individually after dosing at least once during the first
30 min; periodically during the first 24 h with special attention given
during the first 4 h and daily thereafter, for a total of 14 d.
Experimental design
A total of 30 male wistar rats were utilized and the animals were
randomly divided into 5 groups of 6 rats in each group:
Group I-Normal [standard diet+5% sucrose (p. o.)]
Group II–Diabetic control [standard diet+triton (i. p.)+5% sucrose
(p. o.)]
Group-III-Standard drug (10 mg/Kg b.w., p. o.)+triton+standard diet.
Group-IV-Diabetic control+Capsicum frutescens extract (50 mg/kg
body weight which is 5% of the dose used for acute toxicity
studies)+triton+standard diet
Group–V-Diabetic control+Capsicum frutescens extract (100 mg/kg
body weight which is 10% of the dose used for acute toxicity
studies)+triton+standard diet
Effect in Triton-induced hyperlipidemic rats
Animals were kept for fasting for 24 h and was injected with a
solution of Triton WR-1339 (dissolved in 0.9% NaCl) at a dose of
400 mg/kg body weight intraperitoneally. The plant extracts, at a
dose of 50 mg/kg and 100 mg/kg body weight were administered
orally through gastric intubation. The first dose was given
immediately after triton injection and a second dose 20 h later.
Blood samples were collected at 6, 24 and 48h after triton injection
and were used for the study of various biochemical parameters.
Blood was collected through retro-orbital plexus route on anesthesia
and centrifuged at 2000 rpm for 30 minutes [5-6].
Biochemical analysis
Serum samples were analyzed for
Total serum cholesterol (TC),
Triglyceride (TG)
High-density lipoprotein cholesterol (HDL-C)
Low-density lipoprotein (LDL-C)
LDL-C/HDL-C ratio was calculated as the ratio of plasma LDL-C to
HDL-C levels.
Data analysis
Data were statistically analyzed as mean+SEM and expressed as
non-significant P>0.05, just significant P<0.05, significant P<0.01
and highly significant P<0.001 by using ANOVA followed by
Dunnett’s t-test and unpaired t-test with Welch correction.
RESULTS
Electrospray mass spectrometry analysis
The presence of intense peaks at 306.3 and 308.3 indicated the presence
of the Capsaicinoids–Capsaicin and Dihydrocapsaicin (fig. 1).
Fig. 1: MS scan showing the peaks for capsaicin and dihydrocapsaicin
In vivo acute oral toxicity studies
Acute oral toxicity studies for the extracts of the leaves of Capsicum
frutescens revealed that both the extracts were non-toxic attested
dose levels and well tolerated by the experimental animals.
Effect on the lipid profile
The results of the lipid profile are tabulated in tables 1-3. The extract
showed a decrease in blood lipids in hyperlipidemic rats when
compared with the respective diabetic controls. The extract showed
Thomas et al.
Int J Curr Pharm Res, Vol 9, Issue 3, 165-168
167
a significant decrease in TC, TG and LDL along with an increase in
HDL level dose-dependently at the tested doses (50 mg/kg and 100
mg/kg) when administered in Triton-induced hyperlipidemic rats
(tables 1-3).
Table 1: Effect of capsaicinoids on total cholesterol and triglyceride levels in triton induced hyperlipidemic rats (Values are mean+SEM of
6 animals in each group)
Treatment
Cholesterol (mg/dl)
Triglycerides (mg/dl)
6h
24h
48h
6h
24h
48h
Normal
63.4+2.7
61.4+2.1
65.2+1.7
68.1+1.6
70.3+3.5
72.7+1.0
Diabetic control
106.4+2.1
a
164.4+5.3
a
119.0+2.1
a
130.2+1.6
a
148.7+8.6
a
105.2+2.9
a
Standard
91.8
+
2.2
***
94.0+4.0
***
108.6+2.4
*
104.0+3.0
***
81.1+2.6
***
78.0+1.9
***
CE(5
0 mg
/kg)
101.0
+
1.9
ns
142.8+6.6
*
113.8+2.5
ns
117.3+1.9
**
124.6+5.3
*
86.0+2.9
***
CE(10
0 mg
/kg)
96.2
+
2.7
*
120.2+5.7
***
111.8+2.4
ns
112.3+2.8
***
98.5+5.9
***
79.9+2.1
***
***
P<0.001,
**
P<0.01,
*
P<0.05,
ns
P>0.05 (Dunnett’s t-test)
a
P<0.001 (Unpaired t-test)
Table 2: Effect of capsaicinoids on LDL and HDL levels in triton induced hyperlipidemic rats (Values are mean+SEM of 6 animals in each
group)
Treatment
HDL (mg/dl)
LDL (mg/dl)
6h
24h
48h
6h
24h
48h
Normal
23.2+1.0
26.0+0.9
23.6+0.9
22.6+0.9
18.6+3.0
24.6+0
.6
Diabetic Control
16.2+1.0
a
14.1+1.6
a
14.7+1.1
a
43.6+2.4
a
84.2+6.5
a
31.7+2.3
a
Standard
27.2+1.8
***
34.7+3.8
***
29.3+2.2
***
31.3+2.6
***
10.6+1.0
***
17.4+1.0
***
CE(5
0 mg
/kg)
20.3+0.8
ns
25.3+1.7
*
22.0+0.9
*
33.6+2.2
*
26.2+2.9
***
23.8+1.5
**
CE(10
0 mg
/kg)
26.8+0.6
***
35.1+1.6
***
28.3+2.3
***
34.8+1.8
*
19.9+1.1
***
24.0+1.2
**
***
P<0.001,
**
P<0.01,
*
P<0.05,
ns
P>0.05 (Dunnett’s t-test)
a
P<0.001 (Unpaired t-test)
Table 3: Effect of capsaicinoids on TC/HDL ratio and LDL/HDL ratio in triton induced hyperlipidemic rats
Treatment
TC/HDL ratio
LDL/HDL ratio
6h
24h
48h
6h
24h
48h
Normal
2.7+0.1
2.4+0.1
2.8+0.1
1.0+0.1
0.7+0.1
1.0+0.0
Diabetic Control
6.6+0.3
12.3+1.4
8.3+0.6
2.7+0.3
6.4+1.0
2.2+0.3
Standard
3.4+0.2
2.8+0.3
3.8+0.3
1.2+0.1
0.3+0.1
0.6+0.1
CE(5
0 mg
/kg)
5.0+0.2
5.7+0.2
5.2+0.3
1.7+0.1
1.1+0.2
1.1+0.1
CE(10
0 mg
/kg)
3.6+0.2
3.5+0.3
4.1+0.4
1.3+0.1
0.6+0.1
0.9+0.1
DISCUSSION
The three liquid phase system consisting of n-hexane, acetone,
K2HPO4, and water was successfully used to extract capsanthin and
capsaicin. Capsanthin and capsaicin were partitioned mainly into the
n-hexane and acetone phases, respectively. They could be easily
obtained after removing the solvent for further purification. TLPE
was an effective method for the extraction and separation of
capsaicinoids [7].
Triton WR 1339, when administered in fasted rats, causes an
elevation in plasma lipid level. Initially there is a sharp increase in
the lipid level reaching a peak two to three times the control value
by 24h after the administration of Triton injection (Phase I–
synthetic phase), this increase in lipids will fall off within the next
24h i.e. 48h after the administration of Triton (Phase II–Excretion
phase) This increase in plasma lipids by Triton is thought to be due
to one of the following mechanisms; as due to increase in hepatic
synthesis of cholesterol or Triton physically alters very low density
lipoproteins (VLDL) rendering their removal from the blood. Drugs
interfering with cholesterol synthesis were shown to be active in
phase I, while drugs interfering with cholesterol excretion and
metabolism was active in Phase II [8-10].
Total cholesterol (TC) was determined by one step method of
Wybenga and Pillegi [7] based on the reaction between cholesterol
and cholesterol reagent (ferric oxide, ethyl acetate and sulfuric acid).
High-density lipoprotein (HDL) from blood was determined by a
two-step method i.e. initial separation of HDL from blood using a
precipitating agent and then the precipitated HDL was determined
by using calorimetric reaction with cholesterol reagent [11].
Triglyceride (TG) was determined calorimetrically by an enzymatic
reaction using glycerol-3-phosphate oxidase. Enzymatic splitting of
lipoprotein lipase along with reaction between 4-aminoantipyrin, 4-
chlorophenol and H
2
O
2
under the catalytic action of peroxidase
generates quinimine which is used as an internal indicator in this
calorimetric determination [12]. LDL was determined by using
Grindelwald's formula–LDL = TC-HDL-(TG/5) [13].
The preliminary screening revealed the presence of Capsaicin and
dihydrocapsaicin in the acetone extract. Capsaicinoids significantly
decreased the plasma triglycerides, total cholesterol, LDL
cholesterol and increased the high-density lipoprotein cholesterol.
The cholesterol-lowering action of capsaicinoids was attributed to
the inhibition of hepatic cholesterol synthesis, also by the
stimulation of the conversion of cholesterol to bile acids by up-
regulation of cholesterol 7α-hydroxylase expression and increasing
the excretions of bile acids in feces [14].
In conclusion, the present study shows that Capsaicinoids extracted
from Capsicum frutescens has a plasma LDL-C, cholesterol,
triglyceride lowering effect and has a plasma HDL-C raising effect.
We think that it is interesting and meaningful to report that dietary
intake of Capsaicinoids can improve plasma lipid levels.
AUTHOR CONTRIBUTION
L. T. designed all the experiments, carried out the main experimental
work and wrote the manuscript. J. V. K supported the idea of the
manuscript and provided a number of suggestions.
CONFLICT OF INTERESTS
Declare none
Thomas et al.
Int J Curr Pharm Res, Vol 9, Issue 3, 165-168
168
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How to cite this article
• Licto Thomas, Jagadish Vasudev Kamath. Evaluation of the
antihyperlipidemic activity of Capsicum frutescens extract. Int J
Curr Pharm Res 2017;9(4):165-168.
