Chyawanprash (Chy) is an ayurvedic formulation commonly consumed in Indian households. Chy is a comprehensive herbal tonic, prepared from around 50 herbs employing anwala (
Learning and memory are two fundamental cognitive functions that confer us the ability to accumulate knowledge from our experiences [
Chyawanprash (Chy), a household remedy all over India, is popular for its nutritional value. In ayurveda, Chy is classified under the category of Rasayana, which aims at maintaining physique, vigor and vitality, while delaying the ageing process [
Chy is a complex mixture of more than 50 herbal ingredients. All the ingredients in Chy have been scientifically studied individually for their health benefits. The combination of these nutrients used in Chy in a specific quantity and manner of blending creates a powerful synergy for optimum health benefits. Chy is helpful in clearing the accumulated excreta by promoting digestion and excretion [
Chy manufactured by Dabur, India was used in the present study. The proportion of various constituents of Chy is given in Table
Proportion of constituents of Chy.
Common name | Plant name | Weight per 100 g of Chy |
---|---|---|
Dashmool | 3.98 g | |
Bala | 0.398 g | |
Mudgaparni | 0.398 g | |
Mashparni | 0.398 g | |
Karkatshringi | 0.398 g | |
Bhumiamlaki | 0.398 g | |
Jivanti | 0.398 g | |
Pushkarmool | 0.398 g | |
Agarkashta | 0.398 g | |
Haritaki | 0.398 g | |
Guruchi | 0.398 g | |
Katchur | 0.398 g | |
Mustak | 0.398 g | |
Punarnava | 0.398 g | |
Neelkamal | 0.398 g | |
Vasaka | 0.398 g | |
Kakanasa | 0.398 g | |
Yashtimadhu | 0.398 g | |
Varahikand | 0.796 g | |
Ashwagandha | 0.796 g | |
Satavari | 0.796 g | |
Vidarikand | 1.195 g | |
Anwala green | 90 g | |
Ghrit | Milk fat | 2.08 g |
Sesame oil | 1.20 g | |
Banslochan | 0.8 g | |
Akarkara | 0.126 g | |
Pippali | 1.12 g | |
Nagkesar | 0.116 g | |
Dalchini | 0.116 g | |
Tejpatra | 0.116 g | |
Lavang | 0.128 g | |
Elaichi | 0.588 g | |
Kesar | 0.021 g | |
Chandansaar | 0.0092 g | |
Abhrak Bhasam | Ayurvedic preparation | 0.188 g |
Muktasukti Pishti | Ayurvedic preparation | 0.063 g |
Silver foil | q.s. | |
Sugar | q.s. |
q.s.: quantity sufficient.
The drugs and chemicals employed in this study were obtained from following drug houses: Chy (Dabur, India), donepezil hydrochloride (Donep; Wokhardt Ltd., Baddi, India), alprazolam (Alp) (Ind-Swift, Baddi, India), Sco hydrobromide (Sigma Aldrich, USA), 1,1,3,3-tertramethoxy propane (Sigma Aldrich, USA), 5,5-dithiobis-2-nitrobenzoic acid (DTNB) and reduced glutathione (Sisco Research Laboratories Pvt. Ltd., Mumbai, India), Piracetam (Nootropil, UCB India Ltd, India), acetylcholine iodide, eserine salicylate, sodium dihydrogen phosphate, disodium hydrogen phosphate (Hi-Media, India). Sco HBr and Donep HCl were dissolved in distilled water. Alp was suspended in 1% carboxymetylcellulose sodium.
Swiss male mice (20–25 g, 3-4 months of age) were employed in this study, as consequent variation in estrogen levels in female mice may influence the cognitive behavior of the animals [
Exteroceptive behavioral models, such as (i) Morris water maze (MWM), (ii) Hebb-Williams maze (HWM) and (iii) elevated plus maze (EPM), and Interoceptive behavioral models, such as (i) Sco-induced amnesia (1.4 mg kg−1; i.p.) [
The procedure, technique and end point for testing memory were followed as per the parameters described earlier [
On fifth day (i.e., 19th day of Chy administration), the platform was removed. Mouse was placed in water maze and allowed to explore the maze for 120 s. Each mouse was subjected to four such trials and each trial was started from a different quadrant. Mean time spent in all the three quadrants, that is, Q1, Q2 and Q3 was recorded and the time spent in the target quadrant (TSTQ) in search of the missing platform provided as an index of retrieval. Care was taken not to disturb the relative location of water maze with respect to other objects in the laboratory.
HWM is an incentive-based exteroceptive behavioral model useful for measuring spatial working memory of rodents. The procedure, technique and end point for testing memory were followed as per the parameters described earlier in our studies [
EPM served as the exteroceptive behavioral model to evaluate short-term memory in mice. The procedure, technique and end point for testing memory was followed as per the parameters described earlier [
The effect of Chy on ambulation (spontaneous locomotor activity) was recorded using Medicraft photoactometer (INCO, Ambala, India) in different groups of mice.
Mice were divided in 38 groups comprising of six animals in each group. Chy was administered for 15 days. After completion of 15 days, behavioral studies were carried out using MWM (Groups I–XII), HWM (Groups XIII–XXIV) and EPM (Groups XXV–XXXVI). Chy administration was continued during maze studies, that is, Chy was administered for 16 days in case of EPM and HWM and for 19 days in case of MWM. The animals had free access to food and water before subjecting them to behavioral studies using mazes. Piracetam is an established memory enhancer [
Groups I, XIII, and XXV (control groups): Standard diet (without Chy) was administered to mice.
Groups II and III, XIV and XV and XXVI and XXVII (Chy groups): Chy in two concentrations (1 and 2% w/w in the diet) was administered to mice daily for 15 successive days.
Groups IV, XVI, and XXVIII (piracetam groups): Piracetam (400 mg kg−1; i.p.) was administered for 15 successive days to mice.
Groups V, XVII, and XXIX (Sco group): Sco (1.4 mg kg−1; i.p.) was administered daily from (days 1 to 4) 30 min before training on water maze.
Groups VI and VII, XVIII and XIX and XXX and XXXI (Chy + Sco groups): Chy in two concentrations (1 and 2% w/w in the diet) was administered to mice daily for 15 successive days. Sco (1.4 mg kg−1; i.p.) was injected after 90 min of Chy administration daily from days 15 to 18.
Groups VIII, XX, and XXXII (Piracetam + Sco group): Piracetam (400 mg kg−1; i.p.) was administered for 15 successive days to mice. Sco (1.4 mg kg−1; i.p.) was injected after 90 min of piracetam administration daily from days 15 to 18.
Groups IX, XXI and XXXIII (Alp group): Alp (0.5 mg kg−1; i.p.) was administered daily from (days 1 to 4) 30 min before training on water maze.
Groups X and XI, XXII and XXIII and XXXIV and XXXV (Chy + Alp groups): Chy in two concentrations (1 and 2% w/w in the diet) was administered to mice daily for 15 successive days. Alp (0.5 mg kg−1; i.p.) was injected after 90 min of Chy administration daily from days 15 to 18.
Groups XII, XXIV and XXXVI (Piracetam + Alp group): Piracetam (400 mg kg−1; i.p.) was administered for 15 successive days to mice. Alprazoalm (0.5 mg kg−1; i.p.) was injected after 90 min of piracetam administration daily from days 15 to 18.
After behavioral studies, animals were sacrificed by decapitation. The whole brain was removed, weighed and homogenized in an ice bath after adding 10 volumes of phosphate buffer (pH 7.4). The homogenate was centrifuged at 3000 rpm for 15 min and the resultant cloudy supernatant fluid was used for estimation of acetylcholinesterase (AChE), reduced glutathione (GSH) and thiobarbituric acid reactive substances (TBARS). Since Donep is a marketed preparation for the management of Alzheimer’s disease (AD), which acts through inhibition of acetyl-cholinesterase enzyme [
Group XXXVII (Donep group): Donep (0.1 mg kg−1; i.p.) was administered for 15 days daily to mice. Estimation of brain AChE, GSH and TBARS was carried out on 15th day after 90 min of Donep administration.
Group XXXVIII (Donep + Sco group): Donep (0.1 mg kg−1; i.p.) was administered for 15 days daily to mice. Sco (1.4 mg kg−1; i.p.) was injected on 15th day after 90 min of Donep administration. Estimation of brain AChE, GSH and TBARS was carried out on 15th day after 45 min of Sco administration.
The whole brain AChE activity was measured by the method of Ellman et al. with a slight modification [
The whole brain GSH level was measured by the method of Beutler et al. [
The whole brain TBARS level was measured by the method of Ohkawa et al. [
All the results were expressed as mean ± SEM. Data were analyzed using one way analysis of variance followed by
The administration of Chy has not affected the diet intake and weight of the animals in comparison to control group animals in the present study.
The administration of Sco (1.4 mg kg−1; i.p.) and Alp (0.5 mg kg−1; i.p.) before training trials induced amnesia in mice using MWM (Table
Effect of Chy on day 4 ELT and day 5 time spent in target quadrant (TSTQ) of mice using MWM.
Group | Treatment | Dose | Day 1 ELT (s) | Day 4 ELT (s) | Day 5 TSTQ (s) |
---|---|---|---|---|---|
I | Control (normal) | Standard diet | 69.7 ± 2.4 | 25.6 ± 1.8a | 73.4 ± 3.6 |
II | Chy | 1% w/w, in diet | 68.6 ± 1.6 | 24.2 ± 2.4 | 70.6 ± 4.2 |
III | Chy | 2% w/w, in diet | 65.6 ± 0.6 | 26.7 ± 2.9 | 72.8 ± 2.4 |
IV | Piracetam | 400 mg kg−1; i.p. | 67.9 ± 3.9 | 13.4 ± 3.1b | 89.9 ± 2.2e |
V | Sco | 1.4 mg kg−1; i.p. | 66.3 ± 3.5 | 56.2 ± 3.6b | 29.2 ± 4.9e |
VI | Chy + Sco | 1% w/w, in diet + 1.4 mg kg−1; i.p. | 64.3 ± 1.2 | 34.8 ± 2.1c | 52.1 ± 1.8f |
VII | Chy + Sco | 2% w/w, in diet + 1.4 mg kg−1; i.p. | 62.1 ± 4.3 | 25.4 ± 1.3c | 61.2 ± 4.6f |
VIII | Piracetam + Sco | 400 mg kg−1; i.p. + 1.4 mg kg−1; i.p. | 62.3 ± 2.9 | 14.4 ± 2.7c | 66.3 ± 2.8f |
IX | Alp | 0.5 mg kg−1; i.p. | 63.4 ± 2.9 | 59.9 ± 4.3b | 24.3 ± 1.2e |
X | Chy + Alp | 1% w/w, in diet + 0.5 mg kg−1; i.p. | 63.7 ± 2.7 | 32.1 ± 1.6d | 54.2 ± 3.2g |
XI | Chy + Alp | 2% w/w, in diet + 0.5 mg kg−1; i.p. | 60.1 ± 2.5 | 22.1 ± 4.1d | 64.6 ± 3.7g |
XII | Piracetam + Alp | 400 mg kg−1; i.p. + 0.5 mg kg−1; i.p. | 66.3 ± 1.9 | 17.3 ± 2.3d | 69.1 ± 3.6g |
Piracetam (400 mg kg−1; i.p.) is used as a standard drug. Values are mean ± SEM.
a
b
c
d
e
f
g
Effect of Chy administered for 15 successive days on time taken by mice to reach reward chamber (TRC) using HWM. Piracetam (Pira) (400 mg kg−1; i.p.) is used as a standard drug. Values are mean ± SEM; a
Effect of Chy administered for 15 successive days on TL of mice using EPM. Piracetam (Pira) (400 mg kg−1; i.p.) is used as a standard drug. Values are mean ± SEM; a
No statistically significant differences were observed in brain AChE activity of Chy-treated mice and control group mice. On the other hand, administration of Sco (1.4 mg kg−1, i.p.) significantly increased the brain AChE activity, which was reversed (
Effect of Chy administered for 15 successive days on brain AChE activity. Donep (0.1 mg kg−1; i.p.) is used as a standard drug. Values are mean ± SEM; a
No statistically significant differences were observed in TBARS levels (Figure
Effect of Chy administered for 15 successive days on brain TBARS. Donep (0.1 mg kg−1; i.p.) is used as a standard drug. Values are mean ± SEM; a
Effect of Chy administered for 15 successive days on brain GSH level. Donep (0.1 mg kg−1; i.p.) is used as a standard drug. Values are mean ± SEM; a
In the present study, Alp did not affect significantly the brain AChE activity, TBARS levels and GSH levels in any manner. Chy (1 and 2% w/w in the diet) when administered for 15 successive days did not show any significant change in locomotor activity of mice (scores: 193 ± 12 and 202 ± 14) as compared to control animals (score: 178 ± 09). The schematic diagram illustrating the beneficial effect of Chy on memory is depicted in Figure
Schematic diagram illustrating the beneficial effect of Chy on memory.
Dementia is a clinical syndrome characterized by the development of multiple cognitive defects that are severe enough to interfere with daily social and professional functioning [
In the present study, Sco and Alp produced amnesia in experimental animals as indicated by the increased day 4 ELT, reduced TSTQ (time spent to reach target quadrant) using MWM, increased TL using EPM and increased TRC (time taken to reach reward chamber) using HWM. Sco (centrally acting anti-muscarinic drug) and Alp (a benzodiazepine) have been extensively utilized to induce memory deficits in experimental animals [
Sco-induced amnesia is mediated through blockade of central muscarinic receptors [
An increased lipid peroxidation due to increased generation of free radicals and decreased scavenging of free radicals due to reduced antioxidant enzymes has been reported in AD brain [
It is noteworthy that phytoconstituents such as
The authors are deeply grateful to Dr D. D. S. Sandhu, Hon’ble Vice Chancellor, Guru Jambheshwar University of Science & Technology, Hisar and Dr Rajendar Singh, Chairman, Rajendra Institute of Technology & Sciences, Sirsa for support and institutional facilities.