Download Nitric oxide synthase activation is a unique mechanism of garlic action

136s Biochemical Society Transactions ( 1 995) 23
Nitric oxide synthase activation is a unique mechanism of garlic
action.
INDRAJIT DAS, NUSRAT S. KHAN AND 'SUREN R.
SOORANNA
in a dose dependent manner (Fig. 2). NOS activities are suppressed
at higher concentrations of garlic, although NO production must
have increased, as reflected by increased nitrite concentrations.
This is probably due to NO acting as a feedback modulator of its
own synthesis [7].
4cdemic Departments of Psychiatry and 'Obstetrics &
Gynaecology. Charing Cross & Westminster Hospital, London W6
8RP.
Garlic (Alliwn sarivum L . ) is thought to decrease blood
pressure, inhibit platelet aggregation, reduce blood fat and
cholesterol as well as possessing antibacterial and antifungal
properties [I]. The main active agents in garlic extracts are allicin
and ajoene, but several other chemicals have been isolated and
studied [2,3]. Nitric oxide (NO) is linked to hypertension [4] and
the relevant properties of garlic prompted us to investigate whether
the above mentioned effects of garlic extracts are due to the
activation of nitric oxide synthase (NOS), the enzyme that produces
NO.
Garlic extracts were prepared by 1 % alcoholic extraction of
fresh cloves of garlic or from Kwai garlic tablets (gm/rnL). This
was diluted and the effects upon calcium-dependent NOS activities
determined in 2 different in v i m systems. Human platelets were
isolated and prepared for assaying NOS activity in the presence and
absence of garlic extracts at different dilutions. Garlic extracts were
also added to human placental villous tissue and incubated at 37°C
for 45min. Tissues were then separated by centrifugation and
supernatants removed for determination of nitrite concentrations by
chemiluminescence [ S ] . The incubated tissues were rapidly frozen
and homogenates prepared for determination of NOS activities.
Assays were performed in platelets and homogenates by following
the conversion of radiolabelled L-arginine to L-citrulline in the
presence and absence of the specific inhibitor, N'-monomethyl-Larginine [6].
NOS activity increased in a dose dependent manner in
platelet preparations after incubation with garlic extract (Fig. I).
The incubation of placental tissues with garlic extracts also showed
an increase in NOS activity and the metabolite, nitrite,
**
600
4-9
.-
4-9
0
a
400
v)
0
z
be
200
0
A
Control
1/500
r
c
400
200
300
=aE
--
z
-
c
0
-I
4
200
ln
0
2
n
100
H
100
-4
3,
a
0
n
0
Control
112000
11500
11100
Dilution of Garlic E x t r a c t
Fig. 2. The effect of rarlic extract on NOS activitv in ulacental
villous tissues. 100% NOS activity = 1.31 f 2.2 nmoles of
citrulline formed per min per g of protein and 100% nitrite = 2.96
f 0.79 nmoles of nitrite formed per min per g of protein.
Placental villi (1-2min') were incubated in 4mL in Ham's medium
containing various dilutions of garlic extract. The results are the
means f SEM of 5 experiments. * represents P < 0.05 when
compared to controls by the paired Student's f test.
A variety of sulphur containing compounds are formed depending
on the way in which garlic is treated prior to extraction. Both
water and alcoholic extracts possesses some activity of the above
properties claimed. Production of NO by activation of calcium
dependent NOS is probably the most novel mechanism by which
garlic can exert some of its therapeutic properties claimed so far.
Recent studies have shown that both calcium-dependent and
calcium-independent NOS activities are decreased in placental
tissues of pregnancies complicated by pre-eclampsia and growth
retardation [8]. The use of garlic to alleviate these and other
vascular-related diseases warrants further investigation.
We (N.S.K) acknowledge financial support from Horton Hospital
League of Friends.
r
.->
1
0
1/100
1/50
1/10
Dilution of Garlic Extract
Fig. 1, The effect of rarlic extract on NOS activity i n a cell-free
platelet ureuaration. 100% NOS activity = 2.01 k 0.4 nmoles of
citrulline formed per m i n per g of protein. The results are the
means k SEM of 3 - 6 experiments. *, ** and *** represent P <
0.05, 0.02 and 0.005 respectively when compared to controls by
the Student's f test.
1. McMahon, F.G. & Vargas, R. (1993) Phurmcrc.orhcrcipy,
13,406-407
2. Srivastava, K.C. & Tyagi, O.D. (1993) Pro.stcifi/onc/in.s
Leukorricves Essenr. Furry Acids. 49, 587-595
3. Lawson, L . D . , Ransom, D.K. & Hughes, B.G. (1992) 7liromh.
RES. 65, 141-156
4. Moncada, S . , Palmer, R.M.J. & Higgs, E.A. (1991) Phumr.
Rev. 43, 109-142
5 . Ignarro, L.J., Fukuto, J.M., Griscavage, J.M., Rogers, N.E. &
Byrns, R.E. (1993) Proc. Natl. Acad. Sri. USA. 90, 8103-8107
6 . Salter, M., Knowles, R.G. & Moncada, S . (1991) FEBS. Leu.
291, 145-149
7. Rogers, E.R. & Ignarro, L.J. (1992) Bioc'hem. Biophys. Rrs.
Commun. 189, 242-249
8. Morris, N.H., Ramsay, B., Steer, P.J., Pearson, J & Sooranna,
S.R. (1994) Er. J . Ohstef. Gynuecol. Submitted.