ABSTRACT

Ayurvedic formulation triphala was found to be effective in inhibiting γ-radiation induced damage in
microsomal lipids and plasmid pBR322 DNA. The fast reaction kinetic tools like pulse radiolysis and
stopped flow technique were used to asses its antioxidant activities and antioxidant equivalents. The
phytochemical analysis showed that triphala is rich in polyphenols (38± 3%) and tannins (35 ± 3%).
Based on these studies it is proposed that triphala is an effective antioxidant, which can act as a good
radio protector.



DETAILS

Introduction


In the recent past, there has been growing
interest in exploiting the biological activities of
different ayurvedic medicinal herbs, owing to
their natural origin, cost effectiveness and lesser
side effects ].

Triphala is one of the
ayurvedic medicinal herbal formulations
prescribed by most health care practitioners.

It isused as colon tonic, laxative, eye rejuvenator,
anti-inflammatory, anti-viral etc.

It is a composite
mixture of three medicinal herbs Amalaki
(Emblica officinalis), Haritaki (Terminalia
chebula) and Bibhitaki (Terminalia belerica).
Triphala is gentle for people of all ages, from
children to seniors and hence is recommended
for everybody [3].

Triphala has been tested as
an antioxidant and also as a radioprotector in
mice [4, 5]. In the present study, we tested the in
vitro antioxidant activity under γ-radiation
induced conditions.

In order to understand the
factors responsible for the antioxidant and radio
protection activity, free radical reactions and
phytochemical analysis of triphala were carried
out.






Experimental



Lipid peroxidation in microsomes and DNA
damage in pBR322 were carried out using 60Co
γ-source [2].

Nanosecond pulse radiolysis and stopped flow technique were used to study rates
of free radical reaction and to determine the
antioxidant equivalents. Phytochemical analysis
were carried out by using HPLC and absorption
spectrophotometry.




Results and Discussion


The aqueous extract of triphala (20 μg/ml)
inhibited γ-radiation induced lipid peroxidation in
rat liver microsomes at all the doses employed
(120 – 360 Gy) to the extent of 65 to 85%. By
using concentration profile studies (5-35 μg/ml)
at a fixed dose of 240 Gy, IC50 value of 10μg/ml
was determined.

Triphala (25-200 μg/ml) was also found to be effective (~35-75%) in inhibiting
γ-radiation induced (absorbed dose of 6 Gy)
strand breaks in plasmid pBR322 DNA



The above two studies suggest that triphala
exhibits antioxidant activity under γ-irradiation
conditions. Under these conditions, damage to
biomolecules is initiated by the free radicals
produced by the radiolysis of water. Hence it is
appropriate to study their free radical scavenging
ability.




Free radical reactions of Triphala




The radical scavenging experiments were
carried out by using fast reaction kinetic tools
like pulse radiolysis and stopped flow technique
and the reactivity of triphala towards different
radicals such as hydroxyl radicals, superoxide
radicals, DPPH and ABTS•− were determined.
Triphala was found to be an effective scavenger
of DPPH and superoxide radicals.


The reaction of ●OH with triphala, produced a
transient absorbing in the region 350 – 500 nm
with a major transient absorption peak at 350 nm
attributed to gallic acid type of radicals.



Phytochemical analysis


Phenolic acids, flavonoids and tannins are the
most commonly found polyphenolic compounds
in the plant extracts. In the present studies we
have estimated the total polyphenolic and tannin
content in triphala by using Folin- Ciocalteau
method and Folin-Denis method respectively.

It showed that triphala contains 38± 3%
polyphenols and 35 ± 3% tannins. The HPLC
analysis was carried out by using a C18 PCX
500 analytical column and mobile phase (0.05 M
HCl, 0.1 M KCl and varying the percentage of
acetonitrile from 2.5 – 32 %). The detector used
for HPLC analysis is a UV detector set at 260
nm.


It showed that triphala contains sufficient amount
of gallic acid (Figure 2), so that it can be used as
marker compound for in-vivo studies.





Conclusions



Triphala, a well known ayurvedic formulation,
exhibits antioxidant activity and radio protection
ability under in vitro conditions The polyphenolic
content in triphala confirm that the antioxidant
and radioprotecting ability of triphala arises from
the polyphenols, which reduce oxidative stress
by converting the reactive oxygen free radicals
to non-reactive products. The studies are of
great significance as the demand for herbal
products as antioxidants and radio protectors is
increasing constantly.


( SOURCE -- Founder’s Day Special Issue, 2005)

Note - The aim of this article is to increase awareness in Ayurvedic community about
wonderful uses of ayurvedic drugs , and to enhace use of evidence based medicine.