Concentration Dependent Effect of ()-Epicatechin
on Na
+
/K
+
-ATPase and Ca
2+
-ATPase Inhibition
Induced by Free Radicals in Hypertensive Patients:
Comparison with L-ascorbic Acid
Navneet Kumar,
1
* Ruchi Kant,
1
Pawan Kumar Maurya
2
and Syed Ibrahim Rizvi
3
1
Department of Biomedical Sciences, Jimma University, Jimma, P.B.-378, Ethiopia
2
Amity Institute of Biotechnology, Amity University Uttar Pradesh, Noida, 201303, India
3
Department of Biochemistry, University of Allahabad, U.P., India
Although the antioxidant properties of flavonoids are well documented, it is still unclear whether these effects are
dependent on radical scavenging or iron chelating activities. Oxidative stress, a state of excessive reactive oxygen
species (ROS) activity, is associated with vascular disease conditions such as hypertension. Both the anti- and pro-
oxidant effects of tea catechins have been implicated in the alterations of cellular functions that determine their
chemoprotective and therapeutic potentials in health and diseases. The present study examined the concentration
dependent (10
7
to 10
4
M) effects of ()-epicatechin and L-ascorbic acid on Na
+
/K
+
-ATPase and Ca
2+
-ATPase
activity in hypertensive patients and normal subjects. L-ascorbic acid has been used as a positive control to compare
the effect of ()-epicatechin. A significant (p < 0.0001) decrease in the activities of Na
+
/K
+
-ATPase and Ca
2+
-
ATPase was observed in hypertensive patients compared with normal subjects. We report that ()-epicatechin
shows a significant (p < 0.001) dose-dependent protective effect against oxidative stress induced by tert-butyl
hydroperoxide (t-BHP), which is manisfested as a decrease in the activity of erythrocyte Na
+
/K
+
-ATPase and
Ca
2+
-ATPase, in hypertensive patients as well as normal subjects. The effect of L-ascorbic acid was also significant
(p < 0.001) and was comparable with that of ()-epicatechin. Copyright © 2012 John Wiley & Sons, Ltd.
Keywords: epicatechin; erythrocyte; hypertension; oxidative stress.
INTRODUCTION
The naturally occurring polyphenols, especially those
belonging to the class of flavonoids, have recently
been receiving increasing attention owing to their strong
antioxidant activity (Pandey and Rizvi, 2009). Tea
(Camelia sinensis) is the most popular beverage world-
wide. Tea contains polyphenolic compounds collectively
known as catechins belonging to the flavonoid family.
Catechins are known to possess antioxidant, anticancer,
cardio-protective (Tijburg et al., 1997), hypoglycaemic
(Rizvi et al., 2005) and antiageing (Maurya and Rizvi,
2009; Maurya and Prakash, 2011) effects. L-ascorbic acid
is one of the most important antioxidants present in
plasma, it can donate a hydrogen atom and form a rela-
tively stable ascorbyl free radical (Weber et al., 1996).
Recently we have reported the protective role of
()-epicatechin in hypertensive patients (Kumar et al.,
2010) and also as an antiageing compound (Maurya and
Rizvi, 2009; Maurya and Prakash, 2011). The present
study was carried out to evaluate the concentration
dependent effect of ()-epicatechin on erythrocyte mem-
brane bound Na
+
/K
+
-ATPase and Ca
2+
-ATPase activities
in hypertensive patients as well as normal subjects after
induction of oxidative stress by tert-butyl hydroperoxide
(t-BHP). L-ascorbic acid, a well-known antioxidant
compound has been used as a positive control to compare
the effect of ()-epicatechin.
The membrane bound Na
+
/K
+
-ATPase (EC 3.6.1.3) is the
enzymatic basis of univalent cation transport (Sweadner
and Goldin, 1980). It is widely believed that impairment
in Na
+
/K
+
-ATPase activity may play a major role at the
cellular level in the pathophysiology of many diseases
including hypertension (Jaitovich and Bertorello, 2010).
It is also important in regulating various signal functions
of Ca
2+
(Carafoli, 1987; Dally et al., 2007).
MATERIAL AND METHODS
Selection of subjects. Venous blood from hypertensive
patients was taken after informed consent. The mean
age of hypertensive patients was 53.3 5.4 yr, mean
systolic pressure was 150.6 6.7 mm Hg and diastolic
pressure was 109.9 4.8 mm Hg. The BMI of the
subjects ranged between 25.5 kg/m
2
and 29.5 kg/m
2
.
Care was taken to select those subjects who were not
on chemotherapy. The patient was not suffering from
any other disease. All the experiments were carried
out within the 2–3 h of collection of sample of both
control and hypertensive patients. The protocol of study
was in conformity with the guidelines of the Institutional
Ethical Committee.
Determination of Na
+
/K
+
-ATPase activity. Na
+
/K
+
-ATPase
activity was assayed as described earlier (Suhail and Rizvi,
1987). The final assay mixture contained 0.4–0.9 mg
* Correspondence to: N. Kumar, Department of Biomedical Sciences,
College of Medical Sciences and Public Health, Post Box-378, Jimma
University, Jimma, Ethiopia.
E-mail: navneetbiochem@yahoo.co.in; navneet.kumar@ju.edu.et
PHYTOTHERAPY RESEARCH
Phytother. Res. (2012)
Published online in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/ptr.4624
Copyright © 2012 John Wiley & Sons, Ltd.
Received 20 July 2011
Revised 05 January 2012
Accepted 16 January 2012