Effect of standardized aqueous extract of Withania somnifera on tests of cognitive and psychomotor performance in healthy human participants

Abstract

Withania somnifera is an herbal medicine that has been known to possess memory-enhancing properties. The current study involved an assessment of cognitive and psychomotor effects of Withania somnifera extract in healthy human participants.
In this prospective, double-blind, multi-dose, placebo-controlled, crossover study, 20 healthy male participants were randomized to receive 250 mg two capsules twice daily of an encapsulated dried aqueous extract of roots and leaves of Withania somnifera or a matching placebo for a period of 14 days. Cognitive and psychomotor performance was assessed pre-dose (day 1) and at 3 hrs post-dose on day 15 using a battery of computerized psychometric tests. After a washout period of 14 days, the subjects crossed-over to receive the other treatment for a further period of 14 days as per prior randomization schedule. Same battery of test procedures were performed to assess cognitive and psychomotor performance.
Significant improvements were observed in reaction times with simple reaction, choice discrimination, digit symbol substitution, digit vigilance, and card sorting tests with Withania somnifera extract compared to placebo. However, no effect can be seen with the finger tapping test.
These results suggest that Withania somnifera extract can improve cognitive and psychomotor performance and may, therefore, be a valuable adjunct in the treatment of diseases associated with cognitive impairment.

Full text

12 Pharmacognosy Research | January-March 2014 | Vol 6 | Issue 1
Address for correspondence:
Prof. Usha Rani Pingali, Department of Clinical Pharmacology
and Therapeutics, Nizam’s Institute of Medical Sciences,
Panjagutta, Hyderabad, India. E-mail: ushapingali@yahoo.com
Effect of standardized aqueous extract of Withania
somnifera
on tests of cognitive and psychomotor
performance in healthy human participants
Usharani Pingali, Raveendranadh Pilli, Nishat Fatima
Department of Clinical Pharmacology and Therapeutics, Nizam’s Institute of Medical Sciences, Panjagutta, Hyderabad, India
Submitted: 20-04-2013 Revised: 20-06-2013 Published: 12-12-2013
INTRODUCTION
Evaluation of effects of drugs on psychomotor performance
during the new drug development process is an important
aspect, because of their potential to contribute to accidents,
whether on the road, at work place, or at home. Effects of
drugs on the central nervous system (CNS) can be assessed
in terms of effects on behavior, vigilance, attention,
cognition, neurophysiological activity of the brain and
on the neuroendocrine functions. Psychometric tests are
used in clinical pharmacological studies for quantitative
documentation of CNS effects of drugs during the early
phases of drug development.[1]
Withania somnifera,
also known as ashwagandha, is an
important herb in Ayurvedic and indigenous medical
systems. The plant is known as
“
Medhya Rasayan
”
[2-4] or mind
rejuvenate, used in enhancing memory and overall brain
functioning. The active principles of
Withania somnifera
sitoindosides VII-X and Withaferin A (glycowithanolides)
have shown an antioxidant effect in the brain, which may
be responsible for its diverse pharmacological properties.
Withania somnifera
has been used to promote physical
and mental health, to provide defense against disease
and adverse environmental factors, and to arrest the
aging process. Several studies[5-9] indicated that
Withania
somnifera
possesses antioxidant, anti-tumor, anti-stress,
anti-infl ammatory, immunomodulatory, hematopoietic,
anxiolytic, anti-depressive, rejuvenating properties and
was found to play a signifi cant role in the prevention of
different CNS disorders, especially in the conditions of
stress and neurodegenerative diseases, which includes
Parkinson’s and Alzheimer’s disorders, tardive dyskinesia,
cerebral ischemia, and also in the management of the drug
addiction. It has a cognition promoting effect and was
found to be useful in children with memory defi cit and old
Background:
Withania somnifera is an herbal medicine that has been known to possess
memory-enhancing properties. The current study involved an assessment of cognitive
and psychomotor effects of Withania somnifera extract in healthy human participants.
Materials and Methods: In this prospective, double-blind, multi-dose, placebo-controlled,
crossover study, 20 healthy male participants were randomized to receive 250 mg two
capsules twice daily of an encapsulated dried aqueous extract of roots and leaves of Withania
somnifera or a matching placebo for a period of 14 days. Cognitive and psychomotor
performance was assessed pre-dose (day 1) and at 3 hrs post-dose on day 15 using a
battery of computerized psychometric tests. After a washout period of 14 days, the subjects
crossed-over to receive the other treatment for a further period of 14 days as per prior
randomization schedule. Same battery of test procedures were performed to assess cognitive
and psychomotor performance. Results: Signifi cant improvements were observed in reaction
times with simple reaction, choice discrimination, digit symbol substitution, digit vigilance, and
card sorting tests with Withania somnifera extract compared to placebo. However, no effect
can be seen with the fi nger tapping test. Conclusion: These results suggest that
Withania
somnifera extract can improve cognitive and psychomotor performance and may, therefore,
be a valuable adjunct in the treatment of diseases associated with cognitive impairment.
Key words: Central nervous system, psychometric tests, Withania somnifera
extract
ORIGINAL ARTICLE
PHCOG RES.
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DOI: 10.4103/0974-8490.122912
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ABSTRACT

Pharmacognosy Research | January-March 2014 | Vol 6 | Issue 1 13
Pingali,
et al.
: Withania somnifera and psychometric tests
age people with memory loss. In a study, Schliebs
et al
.[10]
reported that
Withania somnifera
improves the cognitive
capabilities of the brain by increasing the capacity of
muscarinic receptors. The present study was thus planned
to evaluate the effect of
Withania somnifera
extract on
cognitive and psychomotor performance in healthy human
participants.
MATERIALS AND METHODS
Study medication
Each capsule contains either an inert placebo or 250 mg
of highly standardized dried aqueous extract of the
roots and leaves of
Withania somnifera
[11] (SENSORIL®,
Natreon Inc, USA). The
Withania somnifera
extract was
derived from withaferin A and corresponding withanolide
glycoside-predominant, genetically uniform chemotype,
cultivated in the central and northern provinces of India.
Withania somnifera
root and leaf material was processed
using a water-based extraction protocol and assessed using
high performance thin layer chromatography analysis of
fractions against standard references (CAMAG Linomat
V applicator, CAMAG TLC Scanner, and WinCats
software version 1.3.4; CAMAG, Sonnenmattstr. Muttenz,
Switzerland). Each
Withania somnifera
extract capsule
contains not less than 10% withanolide glycosides, not
more than 0.5% of withaferin-A and not less than 32%
of oligosaccharides. The placebo capsules were identical in
appearance and contain 49.7% (w/w) of microcrystalline
cellulose, 49.5% (w/w) of lactose, and 0.69% (w/w) of
magnesium stearate.
Study methodology
Twenty-six healthy male participants, aged 20-35 yrs, were
enrolled into the study. The study was approved by the
Institutional Ethics Committee and registered in the clinical
trial registry of India (CTRI/2013/04/003537) and was
carried out in accordance with the Declaration of Helsinki.
Experimental procedures were explained in detail to the study
participants, and written informed consent was obtained prior
to testing. A detailed medical history (including smoking,
alcohol habits), physical examination, hematological, hepatic
and renal parameters, electrocardiogram (ECG), and chest
X-ray were done in all the participants. Subjects were
excluded if there was any evidence of physical illness, drug
abuse or aberrant laboratory fi ndings during screening. All
the subjects abstained from nicotine, caffeine, and alcohol
intake for at least 24 hrs, prior to and during the test day.
The participants were trained in study procedures on at least
two occasions prior to the study to make them familiar with
the testing device.
This was a prospective, double-blind, multi-dose,
placebo-controlled, crossover study, with participants
randomized to receive either
Withania somnifera
extract
or placebo capsules in Run I. On the study day, after
an overnight fast, participants were asked to relax and
sit comfortably for half an hour before initiation of
test procedures in the clinical research unit. All the test
measurements were recorded at baseline between 8:00 and
9:00 AM in an adequately lit, quiet experimental room with
an ambient temperature of 22° ± 2 °C to avoid distraction
and increase the comfort level of the subject. Subjects then
received the study medication as per prior randomization
schedule, with instructions for them to take two capsules
twice-a-day (i.e., in the morning and in the evening) with
240 ml of water daily for 14 days. All the test measurements
were recorded 3 hrs after the morning dose administration
on Day-15. A washout period of 14 days was given between
the treatments. Subjects were then crossed over to receive
the second treatment i.e., 2 capsules twice daily for 14 days
for Run II. All the test procedures were repeated pre- and
post-treatment as described for Run I. Participants were
asked to report any side-effects with either treatment.
If there was any adverse event, same was noted down
in the case record form. Compliance was evaluated by
questioning the subject and counting the leftover capsules.
The drugs assigned for each subject was prepared by a
staff member otherwise not involved in the present study
using a randomization program (Statistica: Stat Soft, Inc,
USA). After the study, the assigned drugs were unblinded
for statistical analysis. Figure 1, provides an overview of
the requirement, allocation, and follow-up of participants.
Assessments
The following tests were administered
Tests of psychomotor performance
Cognitive and Psychomotor performance was assessed
using an indigenously developed automated psychometric
test battery, whose sensitivity and validity has been
previously established in the department.[12] The tests used
in the study are sensitive and can detect drug-induced
changes in psychomotor performance in healthy subjects.
The entire session of tests presented took approximately
20 minutes, and in each session, tests were administered
in the following order. Each test was done thrice, and an
average of three readings was taken.
Finger tapping test
The duration of the test is 10 sec, during which the
subject has to continuously tap on the “Enter Button” on
the response box in quick succession. The results were
presented in terms of average reaction time in milliseconds
as well as total number of clicks attempted. The test
provides information on motor system performance.
Simple reaction test
The duration of the test is 60 sec. In this test, on the click

14 Pharmacognosy Research | January-March 2014 | Vol 6 | Issue 1
Pingali,
et al.
: Withania somnifera and psychometric tests
of a start button, the test time will begin and a picture of
a boy will appear on the center of the screen for 20 times.
The subject has to press the ‘BOY’ symbol button on
response box as quickly as possible every time the ‘BOY’
picture appears on the monitor. Each time, the BOY
picture will remain on the screen for 1 sec, and there will
be a time gap of 1.5-2.5 sec between the appearances of
subsequent BOY pictures. Results are presented in terms
of average reaction time, correct attempts, and wrong
attempts. This test assesses attention and sensory-motor
performance of brain.
Choice discrimination test
The duration of the test is 60 sec. In this test, on the click
of a start button, the test time will begin and a picture of
a boy or girl will appear randomly for 10 times each on the
center of the screen. The subject has to press the ‘BOY’
symbol button with right index fi nger and ‘GIRL’ symbol
button with left index fi nger on the response box as quickly
as possible corresponding to the picture. Each time, the
picture will remain on the screen for 1 sec, and there will
be a time gap of 1.5-2.5 sec between the appearances of
the subsequent pictures. Results are presented in terms
of average reaction time, correct attempts, and wrong
attempts. This test assesses the attention and sensory-motor
performance of the brain and estimates the psychomotor
response speed.
Digit symbol substitution test
In this test, the upper panel of the screen will display 1-9
Figure 1: Requirement, allocation, and follow-up of participants

Pharmacognosy Research | January-March 2014 | Vol 6 | Issue 1 15
Pingali,
et al.
: Withania somnifera and psychometric tests
digits with their corresponding target symbol placed over
each digit. The subject has to carefully concentrate and
remember the corresponding digit for these symbols. On
click of start button, the symbols that are shown in the
panel will appear randomly one after other on the center of
the screen. The subject has to press the corresponding digit
as quickly as possible on the response box when the target
symbol appears. The total duration of the test is 90 sec.
Results are presented in terms of average reaction time,
total attempts, correct attempts, and wrong attempts. This
test assesses attention, response speed, central integration,
and visuo-motor coordination.
Digit vigilance task
A target digit was randomly selected and constantly
displayed to the right of the monitor screen. On click of
start button, a series of digits will appear one at a time on
the center of the screen at the rate of 105 digits/min. The
subject has to press the “Enter Button” on the response
box as quickly as possible every time the digit in the series
matched the target digit. The task lasts for 60 sec, and
there will be 45 stimulus-target matches. The results are
presented in terms of average reaction time (millisec),
total attempts, correct attempts, and wrong attempts. The
test assesses alertness and vigilance while placing minimal
demands on two other components of attention: Selectivity
and capacity.
Card sorting test
In this test, the participant was asked to sort a set of
52 cards based on the different colors and shape using a
digital pen. Results are presented in terms of the average
time taken to complete the sorting and the number of
correct and wrong cards. This test assesses sensory, motor,
central integrative and executive functions.
Safety assessments
Physical examination, systemic examination, vital signs,
hematological and laboratory tests like hemoglobin,
complete blood picture, renal function tests, liver function
tests, lipid profile, and complete urine examination
including ECG were performed at screening visit, baseline,
and end of treatment during each cross-over period.
Statistical analysis
The statistical analysis was carried using Graphpad
prism software, Version 4 (Graphpad software Inc.
Sandiego, California, USA). All the data were presented
as Mean ± SD. Within subject and pair-wise comparisons
between the two treatments (Withania somnifera extract
Vs Placebo) were tested for statistical signifi cance using
one way ANOVA and paired
t
-test. Statistical signifi cance
was at
P
< 0.05.
Sample size calculation
Assuming a 15% difference between two treatments,
sample size of 20 subjects was calculated based on
two-sided differences with the type I error α being 0.05 and
type II error β at 80%. Taking into account study design
and total number of times subject visiting the study center
and expected dropout of 30%, a total of 26 subjects were
enrolled into the study.
RESULTS
Total 26 subjects were screened, out of which four
subjects were excluded because of abnormal laboratory
investigations and two subjects did not report to
randomization visit.
The mean age of the 20 subjects was 24.90 ± 4.18 yrs,
with a mean BMI of 22.38 ± 1.15 kg/m2. There were no
signifi cant differences in baseline characteristics between
the two groups.
The results of the outcome measures are depicted in
Table 1. As seen from the table,
Withania somnifera
extract
signifi cantly decreased reaction time (RT) in SRT, CDT,
DSST, DVT, and CST compared to that of baseline and
placebo. However, no signifi cant effect can be seen with
FTT.
Compared to placebo, the mean percentage change in
reaction time for the CDT, DSST, and DVT decreased
signifi cantly. However, no signifi cant change can be seen
with FTT, SRT, and CST [Figures 2 and 3].
All subjects completed both treatments, and no participant
discontinued the study. Compliance to study medication
was good, and all the subjects in the
Withania somnifera
extract group and in the placebo group took more than
90% of the study drugs. Safety laboratory parameters were
within normal range, and none of the treatments affected
the safety lab parameters.
DISCUSSION
The present study demonstrated the favorable effects of
Withania somnifera
extract on cognitive and psychomotor
performance in healthy participants. Psychometric tests like
FTT, CDT, DSST, DVT, and CST were performed using
computerized psychometric test system, which have gained
popularity in recent times in clinical studies of psychiatric
disorders and to evaluate the effects of psychotropic
drugs.[13] Psychometric tests are used as biological markers
in early phase 1 trials to establish preliminary objective
pharmacodynamic data.[1] In the present study, the

16 Pharmacognosy Research | January-March 2014 | Vol 6 | Issue 1
Pingali,
et al.
: Withania somnifera and psychometric tests
psychomotor performance tests used are well-established
with sensitivity to the depressant and stimulant effects for
a wide range of psychotropic drugs.
Withania somnifera
is a
known brain tonic in Ayurveda
postulated to act in the human body by modulating the
neuro-endocrino-immune systems and have been found
to be a rich source of antioxidants.
In the present study, we have observed that
Withania
somnifera
extract decreased the reaction time signifi cantly
in SRT, CDT, DSST, DVT, and CST following 14 days
treatment (1000 mg/day) compared to placebo, indicating its
positive effect on psychomotor function. Similarly, there was
signifi cant reduction in mean percent reaction time by 6%
in SRT, 7% in DSST, and 3% in DVT compared to placebo.
The decrease in the reaction time could be interpreted
as a central effect, since
Withania somnifera
helps to
maintain homeostasis and psychomotor abilities in
times of stress. Clinical trials have shown that
Withania
somnifera
can alleviate a reactive type of depression
without causing sedation. Instead, it optimizes mental
and psychomotor performance by easing the mental
stress bundle.[8] Our results are in par with the above
data where we have not witnessed any sedative effect in
the subjects during treatment with
Withania somnifera
extract. The mechanism by which
Withania somnifera
exerted benefi cial effects is presently not clear; however,
animal studies have shown that
Withania somnifera
is
capable of improving memory and enhancing cognitive
function by modulation of cholinergic neurotransmission.
As per the literature, leaf and fruit extracts of
Withania
somnifera
consisting of mixtures of sitoindosides VII–X
and withaferin-A produces an increase in the cortical
muscarinic acetylcholine capacity, which may partly
explain cognition-enhancing effect demonstrated in
animal and human models. Schliebs
et al.
[10] studied the
effects of sitoindosides VII-X and withaferin isolated
from aqueous methanol extracts from the roots of
Withania somnifera
on brain cholinergic, glutamatergic,
Table 1: Effects of placebo and Withania somnifera extract on psychomotor performance tests
Placebo Withania somnifera extract
Pre-treatment
Tests TC Avg RT CC WC TC Avg RT CC WC
FTT 52.60±4.52 192±14.22 - - 54.05±4.52 186.7±16.23 - -
SRT 20±0 313±45.46 20±0 0±0 20±0 306±38.84 20±0 0±0
CDT 19.90±0.30 476±33.61 19.50±0.68 0.40±0.68 19.95±0.22 471±34.40 19.49±0.88 0.36±0.80
DSST 55±7.83 1690±278 54±8.67 1.65±1.56 55.13±7.61 1695±262 53±9.09 1.37±0.90
DVT 44.30±3.58 1371±79.49 37±3.16 7.15±2.0 43.35±1.94 1360±57.02 35.65±2.85 8.60±2.87
CST 52±0 98±17.11 51.20±0.89 0.80±0.89 52±0 95.90±19.13 51.25±0.85 0.90±0.91
Post-treatment
FTT 51.95±5.37 194±14.31 - - 55.05±4.95 183±17.31 - -
SRT 20±0 316±26.35 20±0 0±0 20±1 287±43.34**## 20±0 0±0.89
CDT 19.90±0.30 477±29.60 19.55±0.88 0.35±0.81 19.90±0.44 455±50.52*#19.55±0.60 0.60±0.68
DSST 55±6.60 1739±312 53.55±12.12 1.88±1.19 59±08.13 1557±233***#57±8.49 2±1.48
DVT 43.50±3.03 1398±82.18 37.30±2.94 6.10±1.74 46±3.50 1317±87.98**## 38±6.54 8±4.80
CST 52±0 97.65±19.93 51.55±0.60 0.45±0.60 52±0 84.30±12.66*#51.50±0.76 0.80±0.83
Data represented as Mean±SD *
P
<0.05,**
P
<0.01, ***
P
<0.001 compared to pre-treatment of Withania somnifera extract #
P
<0.05, ##
P
<0.01 compared to post-treatment
of placebo. TC : Total clicks; CC : Correct clicks; WC : Wrong clicks; Avg. RT : Average reaction time (millisec. for FTT, SRT, CDT, DSST, DVT and sec. for CST)
Figure 2: Mean % change in RT in FTT, SRT, and CDT before and
after intake of Placebo and
Withania somnifera
extract #
P
< 0.05
compared to placebo
Figure 3: Mean % change in RT in DSST, DVT, and CST before and
after intake of Placebo and
Withania somnifera
extract #
P < 0.05,
##
P < 0.01 compared to placebo

Pharmacognosy Research | January-March 2014 | Vol 6 | Issue 1 17
Pingali,
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: Withania somnifera and psychometric tests
and GABAergic receptors in male Wistar rats. The
activity of AchE levels is enhanced in lateral septum
and globus pallidus, whereas it is decreased in vertical
diagonal band. Similarly, the activity of M1 cholinergic
receptor binding is enhanced in lateral septum and
frontal cortex, whereas M2 receptor binding sites
increased in frontal and parietal cortex regions. The
data suggests that the compounds - sitoindosides VII-X
and withaferin preferentially affect events in the cortical
cholinergic-signal transduction cascade. The drug-induced
increase in cortical muscarinic acetylcholine receptor
capacity might partly explain the cognition-enhancing
and memory-improving effects of
Withania somnifera
extracts in animals and in humans.
Similarly, Dhuley
et al.
[14] tested
Withania somnifera
,
comprising sitoindosides VII–X and Withaferin-A, to prove
their effect on improving short-term memory in mice. The
test was done using electric shock therapy, treatment with
scopolamine (an anti-cholinergic agent that produces
defi cits in learning and memory retention) as well as testing
mice that had never been on the electric grid before. The
mice treated with
Withania somnifera
(200 mg/kg of body
weight) produced a signifi cant decrease in the amount
of errors while trying to reach the shock-free zone after
1 hr of training. The effect of
Withania somnifera
on the
scopolamine-treated mice reversed the defi cits in retrieval
and acquisition. This study showed an effectiveness of
Withania somnifera
in improving short-term memory in
both naive and amnesiac mice.
Though there is not much data on the effects of
Withania
somnifera
on psychomotor performance in healthy
participants, research by Karnick CR[15] in a randomized,
double-blind study reported that
Withania somnifera
signifi cantly improved integrated sensorimotor function,
auditory reaction time, and mental arithmetic ability
compared to Panax ginseng and placebo in 30 healthy
subjects. The results of the present study demonstrate
that the natural products like
Withania somnifera
is a
promising realm that awaits neuropharmacologist to
exploit.
CONCLUSION
The measurement of cognitive and psychomotor
performance is highly important in evaluating and
predicting treatment outcomes in healthy subjects as well as
in patients. In the present study,
Withania somnifera
extract
has shown to improve the cognitive and psychomotor
performance in healthy human participants. However,
multicentric long-term clinical studies in patients are
required to confi rm its therapeutic effi cacy in disease
states associated with impaired cognition and psychomotor
function.
ACKNOWLEDGMENTS
The authors are grateful to the Natreon INC. USA for providing
study medications (SENSORIL® and placebo capsules) and
Dr. I. Sravanthi, Ayurvedic Physician, for her expert advice.
We extend our thanks to Indian Council of Medical Research,
New Delhi, India for providing the extramural grant for
development of instrument for evaluating cognitive and
psychomotor performance under Advanced ICMR Clinical
Pharmacodynamic Center for Evaluation of Pharmacodynamic
Effects of Drugs.
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Cite this article as: Pingali U, Pilli R, Fatima N. Effect of standardized
aqueous extract of
Withania somnifera
on tests of cognitive and psychomotor
performance in healthy human participants. Phcog Res 2014;6:12-18
Source of Support: Natreon INC. USA for providing study medications
(SENSORIL® and placebo capsules), Indian Council of Medical Research,
New Delhi, India for providing the extramural grant for development of
instrument, Confl ict of Interest: None declared.