Evaluation of hypolipidemic effect of citrus lemon

Abstract

Herbs have been used for several cardio vascular diseases, such as congestive heart failure, hypertension, angina pectoris, atherosclerosis, arrhythmia, cerebral and venous insufficiency. Atherosclerosis is the primary cause of heart disease and stroke. Hence blood lipid levels are probably the major determinant of the development of cardiovascular disease. Citrus fruit and juices have long been considered a valuable part of a healthy and nutritious diet. It is well established that some of the nutrients in citrus promote health and provide protection against chronic disease. The present study has been specifically designed to investigate the hypolipidemic effects of citrus lemon juice in rabbits after high cholesterol diet for four weeks. The citrus lemon juice (1ml/kg/day) revealed a significant reduction in serum cholesterol, triglycerides; low density lipoprotein levels and resulted in an increase in high density lipoprotein. These results suggest that the hypocholesterolemic effects of citrus lemon juice may be due to its antioxidant effect.

Full text

Journal of Basic and Applied Sciences Vol. 6, No. 1, 39-43, 2010 ISSN: 1814-8085
EVALUATION OF HYPOLIPIDEMIC EFFECT OF CITRUS LEMON
Yasmin Khan, Rafeeq Alam Khan*, Syeda Afroz and Afshan Siddiq
Department of Pharmacology, Faculty of Pharmacy
University of Karachi, Karachi, Pakistan
ABSTRACT
Herbs have been used for several cardio vascular diseases, such as congestive heart failure, hypertension, angina
pectoris, atherosclerosis, arrhythmia, cerebral and venous insufficiency. Atherosclerosis is the primary cause of heart
disease and stroke. Hence blood lipid levels are probably the major determinant of the development of cardiovascular
disease. Citrus fruit and juices have long been considered a valuable part of a healthy and nutritious diet. It is well
established that some of the nutrients in citrus promote health and provide protection against chronic disease.
The present study has been specifically designed to investigate the hypolipidemic effects of citrus lemon juice in rabbits
after high cholesterol diet for four weeks. The citrus lemon juice (1ml/kg/day) revealed a significant reduction in serum
cholesterol, triglycerides; low density lipoprotein levels and resulted in an increase in high density lipoprotein. These
results suggest that the hypocholesterolemic effects of citrus lemon juice may be due to its antioxidant effect.
Keywords: Citrus lemon, Hypolipidemic effect.
INTRODUCTION
Cholesterol is an amphipathic sterol, hydrophilic at the
hydroxy end with a hydrophobic hydrocarbon side chain
and thus can be incorporated into membrane bilayers. As
well as being an essential structural component of cell
membrane, cholesterol determines membrane lipid
organization and structural integrity (Goldstein et al.,
2006; Ikonen, 2006). Hypercholesterolemia is
characterized by high levels of cholesterol in the blood .It
is not a disease but metabolic derangement that can
contribute to many disease, most notably cardiovascular
disease. It is closely related to the terms hyperlipidemia
and hyperlipoproteinemia .Elevated cholesterol in the
blood is due to abnormalities in the levels of lipoproteins,
the particles that carry cholesterol in the blood streams.
This may be related to diet, genetic factors (such as LDL
receptor mutations in familial hypercholesterolemia) and
the presence of other diseases such as diabetes and an
under active thyroid (Durrington etal, 2003).
Hypercholesterolemia is silent disorder that is usually
diagnosed after a heart attack or stroke. Cardiovascular
diseases have become a major clinical and public health
problem in south Asian countries specifically India,
Pakistan, Srilanka, Bangladesh and Nepal. This
population carries increased risk even if they migrate to
other countries (Iqbal et al., 2005).The national
cholesterol education program (NCEP) published a
guideline in 1993 for screening and treating
hyperlipidemia (Eric et al., 2002). The NCEP program
includes therapeutic life style changes such as reduced
intakes of saturated fats to < 7% of total calories and <
200 mg/day cholesterol. 25% to 35% daily fat intake
provides low saturated and trans fats to body. Therapeutic
options to reduce LDL also involve the use of plant
steroids, 2gm/day and increased intake of viscous fiber
(soluble fiber) about 10-25 gm/day. Similarly exercise
and life style modification is also beneficial. However
most of the allopathic drugs are toxic and generate free
radicals in the body (Beldeu, 2006). Thus present study
has been specifically designed to investigate the lipid
lowering effect of citrus lemon with minimal side effects.
MATERIALS AND METHODS
Selection of Animals
The study was carried out on 18 healthy white rabbits of
either sex weighing from 1000-1500 gm. Rabbits were
selected as experimental animals since biochemical
changes produced in rabbits are comparatively similar as
observed in humans (Irena et al., 1979). Moreover
sufficient amount of blood samples can easily be
collected, easy to handle and economical.
Animal Groups
Animals were divided into two groups control and treated
each containing 9 animals. Apparent health of these
animals was monitored during the conditioning period
under the laboratory environment for a week before
administration of drug specifically noticing loss of hair,
diarrhea, hematuria, skin ulceration and loss of activity.
They were housed individually in cages, under controlled
condition of temperature 23±2ºC. Diet and water was
provided ad libitum.
Dosing
Animals of both groups were given high cholesterol diet
(HCD) daily for a period of 30 days i.e. 0.125gm/kg
cholesterol (Merck) in 0.5% corn oil purchased from local
*Corresponding author: email: rkhan1959@gmail.com

J. basic appl. sci.
40
market. After 30 days animal of treated group were
administered fresh citrus lemon juice orally for 30 days in
the dose of 1ml/kg during the first phase of study, while
animals of control group were administered water through
same route equivalent to the volume of respective doses
according to their body weight. During the second phase
of study animals of treated group were further
administered fresh citrus lemon juice up to 45 days and
compared with control for the same period.
Sample Collection
Blood samples of 5 ml were collected thrice, first after 30
days of HCD then again after 30 and 45 days dosing of
citrus lemon juice from ear vein of the animals in gel tube
for evaluation of lipid profile.
Gross Toxicities
Overall gross toxicities such as skin ulceration, average
weight variation, hematuria, alopecia, loss of activity,
diarrhea, salivation, aggressive behavior and color of skin
in different groups of animals was observed during the
total period of experiment.
Animals of control group showed no gross toxicities, but
there was an overall increase in average weight of control
animals by 15 to 35 gm however animals at HCD
comparatively showed more increase in average weight of
the animals by 25 to 260 gm during the total period of
experiment except in second week, where there was
decrease in weight by 75 gm. Animals at HCD revealed
slight gross toxicities, such as yellow skin, alopecia
(especially at neck and foot), yellow urine with acidic pH
and loss of activity that may be due to severe diarrhea.
Animals at citrus lemon (1ml/kg) after HCD did not
reveal gross toxicities. However there was loss of weight
of 30 to100 gm during the total period of study. However
at the end of study there was 53 gm decrease in average
weight.
MORTALITY RATE
Table 1 reveals mortality rate of animals kept on HCD for
30 days, treated with lemon and control animals for 45
days. Deaths were observed in animals at HCD during 2nd
and 3rd weeks of dosing. However no death was observed
in animals of control and treated groups.
Table 1. Comparison of Overall Mortality Rate (%)
Animals Groups Mortality Rate
HCD 07/25(28.00)
Control 0/9(00.00)
Lemon 0/9(00.00)
No. of animals expired/total animals (percentage)
BIOCHEMICAL ANALYSIS
Serum samples collected at the end of dosing were
utilized for assessment of lipid profile using standard
reagent kits of Human Germany. All the values were
expressed as the mean and standard error to the mean.
Student test was performed and P values were observed.
Values of P<0.05 were considered as significant and
P<0.005 as highly significant. All statistical procedures
were performed according to the method of Alcaraz and
Jimenez (1989).
RESULTS
Table 2 reveals the comparative effects of citrus lemon on
cholesterol, triglycerides, LDL and HDL at the dose of
1ml/kg citrus lemon for 30 and 45 days respectively in
animals received high cholesterol diet for 30 days.
After 30 days animals showed highly significant decrease
in cholesterol 150.8 ± 19.1mg/dl in comparison to control
animals i.e. 345.3± 28 mg/dl. While after 45 days
decrease in cholesterol continued and a highly significant
decrease was observed i.e.16.9 ± 2.8 mg/dl in comparison
to the value of control animals i.e. 232.6±9.0 mg/dl.
Similarly a highly significant decrease in LDL
concentration was observed after 30 days i.e. 122.6± 13
mg/dl in comparison to control animals i.e. 273 ± 38
mg/dl. The decrease in the levels of cholesterol and LDL
persisted even after 45 days. Where as HDL concentration
was increased significantly after 30 days i.e. 7.27 ± 1.7
mg/dl than as compared to control i.e. 3.07 ± 0.26 mg/dl.
However increase in HDL level became insignificant after
45 days i.e. 2.15 ± 0.31 mg/dl as compared to control i.e.
2.1±0.32 mg/dl.
Decrease in triglycerides was insignificant after 30 days
as compared to control animals. However a highly
significant decrease was observed in triglyceride levels
after 45 days i.e. 26.69 ± 1.4 mg/dl in comparison to
control 40.3±3.5 mg/dl.
Table 3 reveals the %age reduction of cholesterol,
triglycerides, LDL and HDL at 1ml/kg dose of citrus
lemon for 30 and 45 days respectively in animals received
high cholesterol diet for 30 days.
DISCUSSION
Herbs have been used as medical treatments since the
beginning of civilization and some derivatives (e g,
aspirin, reserpine, and digitalis) have become mainstays of
human pharmacotherapy. Herbs have been used for
several cardiovascular diseases, such as congestive heart
failure, hypertension, angina pectoris, atherosclerosis,
arrhythmia and cerebral insufficiency. However, many
herbal remedies currently in use have not undergone

Khan et al. 41
careful pharmacological assessment, and some have the
potential to cause serious toxic effects and major drug-to-
drug interactions. The high prevalence of herbal use in
current medical practice throughout the world has
increased questions about the safety and efficacy of herbal
drugs for cardiac disease and clinicians must be informed
about the potential for benefit and harm. Thus continuous
research is necessary to elucidate the pharmacological
activities of the herbal remedies now being used to treat
cardiovascular diseases (Nick et al., 1998).
Vegetables, fruits and their constituents are potent
effectors of biological system in humans (Lampe,
1990).The cholesterol-lowering effects of vegetables,
fruits and their constituents have been examined in detail.
Daily doses of these food showing cholesterol-lowering
effects in human have been reported as follows; variety of
vegetables (750g /day) and fresh apples (350-400g/day),
guava fruit (0.5-1kg /day) , garlic (10-20g /day) and a low
saturated fat diet combined with vegetables, protein
(33g/day) and a high soluble fiber(18g/day) (Lampe,1990;
Jenkins, 1999).
Atherosclerosis is the primary cause of heart disease and
stroke. Hence, blood lipid content is probably the major
determinant of the development of cardiovascular disease.
Several authors have reported the hypolipidemic effects
of Citrus juice (Monforte, 1995; Gorinstein, 2004)
moreover, some studies suggest that a high dietary intake
of orange or grape juice might reduce hypercholestero-
lemia, and this was postulated to be largely due to the
principal Citrus flavanones, hesperidin from orange and
naringenin from grapefruit, as their glycosides, hesperitin
and naringin (Kurowska, 2000).
0
50
100
150
200
250
300
350
400
Cholesterol LDL HDLTriglycerides
serum mg/dl
0
50
100
150
200
250
300
Cholesterol LDL HDL Triglycerides
serum mg/dl
Fig. 1. Histogram showing comparative effects on lipid
profile in HCD and citrus lemon animals after 30 days. Fig. 2. Histogram showing comparative effects on lipid
profile in HCD and citrus lemon animals after 45 days.
Table 2. Comparative effects of citrus lemon on lipid profile after high cholesterol diet for 30 and 45 days
No. of Days Groups Cholesterol
Mg/dl LDL-CHOL
Mg/dl HDL-CHOL
Mg/dl Triglycerides
Mg/dl
Control 345.30±28.0 273.0±38.0 3.07±0.26 39.40±4.7 30 Lemon 150.80±19.1** 122.6±13.0** 7.27±1.7* 39.40±4.2
Control 232.60±9.0 148.1±19.0 2.10±0.32 40.30±3.5 45 Lemon 16.94±2.80** 76.8±6.10** 2.15±0.31 26.69±1.4**
n = 9, Average ± SEM, *p < 0.05 significant as compared to control, **p < 0.005 highly significant as compared to control
Table 3. Percentage reduction in lipid profile after 30 and 45 days administration of citrus lemon
Cholesterol LDL HDL Triglycerides
No. of days Groups %age
Control 51 52.2 21.4 36.1 30 Lemon 79 80.5 20.8 64
Control 68.5 73.8 40.4 41.2 45 Lemon 97.7 87.8 65 67.1
serum conc. m
g/
dl
serum conc. m
g/
dl

J. basic appl. sci.
42
The present study has been specifically designed to
investigate the hypolipidemic effects of Citrus lemon
juice in rabbits after high cholesterol diet for 4 weeks.
The Citrus lemon contains important natural components,
including citric acid, ascorbic acid, minerals and
flavonoids (Benavente et al, 1997) though health-related
properties of citrus lemon have been associated with their
content of vitamin C, but recently flavonoids have been
also shown to play a role in this respect. Some researchers
suggest that flavonoids have different biological
functions, including antioxidative, anti-inflammatory,
antiallergic, antiviral, antiproliferative, antimutagenic and
anticarcinogenic activities (Middleton & Kandaswami,
1986; Godeberg, 1994, Meyer, 1994 and Rice, 1997).
The citrus lemon juice (1ml/kg/day) revealed a significant
reduction in serum cholesterol, triglycerides, low-density
lipoprotein levels and resulted in an increase in high-
density lipoprotein(Fig 1 & 2). These results suggest that
the hypocholesterolemic effect of citrus lemon juice may
be due to its antioxidant effect since previous studies has
suggested that antioxidant (vitamin C) administration in
hypocholesterolemic rats improves endothelial function of
coronary and peripheral vessels (Henry et al., 1997). It is
widely accepted that elevations in cholesterol and LDL
plasma levels are major factors for coronary heart disease
There has been relationship between elevated LDL and
atherosclerosis. Since LDL in the blood gets deposited in
the blood vessel walls and becomes the major component
of atherosclerotic plaque. Studies suggest that
pathological process could be reversed by reducing the
serum LDL level (Ross, 1993). Since citrus lemon juice
has significantly reduced cholesterol and LDL levels
therefore it may be concluded that citrus lemon might
prevent atherosclerosis. Moreover there has been
significant increase in serum HDL levels which is
considered to be good cholesterol (Stein, 1999). The
increase in HDL has shown to slow down the
atherosclerosis process (Nofer, 2002). Therefore it can be
safely concluded that citrus lemon juice could be
considered as a valuable supplement to prevent coronary
diseases.
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