Antibacterial effect of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs)

16 Pages·2015·1.3 MB·English

Basrah Journal of Science (B) Vol. 32(2), 166-181 ,2014
Antibacterial effect of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs)
Zainab R. Abdul-Hussein
Basra University / College of Science /Dept. of Biology
[email protected]
Abstract :-
Effect of five concentrations of five non-steroidal anti-inflammatory drugs (NSAID) included
(acetyl salicylic acid , ibuprofen , sodium diclofenate , mefenamic acid ,and piroxicam)
against five bacterial species (E. coli , Pseudomonas , Klebsiella , Staphylococcus , Bacillus)
via measuring the inhibition zone of drug . All drugs were effected on all bacterial species
except (mefenamic acid , piroxicam) which inhibited only Gramm positive bacteria .
Statistical analysis results showed that the most effected drug was acetyl salicylic acid and the
less effective one was mefenamic acid.

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Abdul-Hussein Z. R. Antibacterial effect of Non-Steroidal…
Introduction
The anti-inflammatory ,analgesic ,and anti- prostaglandin by inhibiting one or both of
pyretic properties of non-steroidal anti- COX isoenzymes .Eicosanoids two
inflammatory drugs (NSAIDs) are pathways ;the cyclooxygenase and
particularly useful in treating rheumatic and lipooxygenase pathway .COX is the key
other musculoskeletal disorders .Theses enzyme in the synthesis of prostaglandin
drugs have been introduced on the market .Two isoforms of the cox enzymes have
of the commercial potential for such been characterized COX1 and COX 2
compounds and testing to their utility in the which they are identical in structure but
treatment of pain and inflammations of have different substrate and inhibitor
varying origin (Nakka,et al.,2011). selectivity .(Rusu 2011).
The NSAIDs are among the most Anti-inflammatory non- steroidal agents
widely described drugs worldwide ,being could be divided in to four main groups 1-
the drug of the first choice in the treatment compounds capable of producing full
of degenerative inflammatory diseases. inhibition of both COX1&COX2with poor
Arachidonic acid (AA) is subsequently selectivity 2- compounds inhibits both
converted by lipooxygenases and COX1&COX2with preference towards
cyclooxygenases (COX) to eicosanoids . COX2, 3- compounds that strongly inhibit
Inhibition of cyclooxygenases and therefore COX2 and 4- compound that appeared to
prostaglandin production is common weak inhibitors for both COX
mechanism of action of the NSAIDs .In 1&COX2(Warner &Mitchel2004, Howard
addition AA is precursor for the production &Delafoutain 2004).
of eicosanoids ,known virulence factor
Different studies investigated the
,stimulating germ tube formation ,and
antibacterial and antifungal effect of the
inflammation during infection and can be
non- steroidal anti-inflammatory drugs .
incorporated in to the phospholipids .
NSAIDs possessed a moderate powerful
NSAIDs are inhibitors of the
effect against Gramm negative bacteria
cyclooxygenases (COX) isoenzymes .these
like Salmonella , E. coli ,Helicobacter
drugs block the synthesis of mammalian
pylori , Klebsiella ,and Enterobacter , as
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Basrah Journal of Science (B) Vol. 32(2), 166-181 ,2014
well as Gramm positive bacteria such as Extraction non -steroidal anti-
Staphylococcus ,Bacillus , Mycobacterium inflammatory drugs (NSAIDs):-
,and Listeria monocytogenes (Mazumdaret
Five type of non-steroidal anti-
al .,2009, Al-Janabi,2009,Al-Janabi,2010,
inflammatory drugs were used in this study
Bend et al .,1999 et al,2003).
including peroxicam ,ibuprofen , aspirin
Studies on NSAIDs also explored (salicylic acid ) , ponstan (mefenamic acid
their effect against fungal infection such as ),and voltarin(diclofenate sodium),all drugs
Candida, Cryptococcus ,Aspergillus, were extracted from tablets or capsule
Penicillium ,Trichoderma( Deva et al., according to their solubility.
2000,Novert et al,2003,Rusu et al .2009,Al-
Piroxycam extracted by dissolving
Bader,2009)
1gm powder in 15 ml chloroform with
The present study was designed to stirring for 1 hour ,filtered , and evaporated
investigate whether the pathogenic bacher at room temperature in dark.(Nakkaet al.
the pathogenic bacteria from different ,2011)
sources were susceptible to different type of
Ibuprofen was extracted by
non -steroidal anti-inflammatory drugs .
powdering 600 mg of tablets after removing
Materials and Methods:- the red coat ,homogenized , and dissolved
in 10 ml chloroform with stirring for 5
Bacterial Isolates:
minutes ,centrifuged ,excluded of
Different pathogenic bacteria ,i.e. E.
precipitate and left supernatant to dried at
coli , Klebsiella , Staphylococcus from
room temperature .(matcovic et al .,2005)
urinary tract infection and Pseudomonas
Salicylic acid was extracted by
from wound infection were used in present
powdering 600 mg of aspirin tablets ,and
study ,while Bacillus was the
dissolving in ethyl acetate with vigorous
onlyenvironmental isolate (all bacteria were
shaking for 2 minutes, filtered ,and dark
obtained from bacteriology laboratory
dried at room temperature.(Williamson
/dept. of biology /college of science.
,1989)

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Abdul-Hussein Z. R. Antibacterial effect of Non-Steroidal…
Mefenamic acid was extracted by for antibacterial effect determination(Al-
dissolving 10 mg of ponstan powder in Janabi ,2009).
glacial acetic acid at 70 oc. with stirring
All data were analyzed statistically
,filtered and dried atroom
by using ANOVA test .
temperature(Othman and Awades,2008) .
Results and Discussion:-
Diclofenate was extracted by
Large amounts of NSAIDs are consuming
dissolving 5mg of voltarin powder in 5ml
every day all over the world for treatment
methanol ,filtered ,and dried at room
of many inflammatory diseases. Obtaining
temperature .( Jawla,and Jain,2010)
data about antibacterial action of such drugs
Preparation of stock solution :To obtain
is still unclear due to variability of
the stock solution (100000µg/ml) of each
influencing factors.
drug,0.1gm from each extract was dissolved
Measurement of inhibition zone of
in 1 ml of DMSO (Dimethyl
non-steroidal anti-inflammatory drugs
sulfoxide).Then a serial dilution (10-
against bacterial spp. Revealed that aspirin
10000µg/ml)from the stock solution were
was the most effective drugs among the
prepared .
studied NSAIDs followed by ibuprofen and
Preparation of bacterial culture :-One
voltarin while each of ponstan and
colony from each bacterial stock culture
piroxicam gave the lowest effect
was inoculated in 4 ml nutrient broth and
.According to studied bacteria, results
incubated at 37oc for 24 hours .
showed that Bacillus was the most affected
Antibacterial assay :- 0.1 ml from each even at low concentration with large
bacterial broth was spreaded on nutrient inhibition zone . All of the 5 NSAIDs
agar plate by using sterile L-shape rod , affected the growth of Gramm positive
then well made with sterile cork-borer ,after bacteria at different concentrations
that 0.1 ml from each concentration of each .Susceptibility of Gramm negative bacteria
NSAIDs was added to well .All plates were towards NSAIDs were varied.
incubated at 37oc for 24 hours ,then

diameters of inhibition zone were measured

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Basrah Journal of Science (B) Vol. 32(2), 166-181 ,2014

Table (1) Diameter of inhibition of NSAIDs exposed bacteria
Bacteria E. coli Pseudom Klebsiel Staphyloco Bacillus
onas la ccus
NSAIDS

1*10^1 6 4
Sali 1*10^2 6 6 6
cyli
c 1*10^3 12 10 8 10
aci
d
1*10^4 12* 18 12 20 12
1*10^5 22 20 40 38 22
1*10^1 4 4
Ibu 1*10^2 10 6 6
pro
fen
1*10^3 12 10 6 4 8

1*10^4 16 12 10 12 12
1*10^5 20 14 12 18 16
1*10^1 6
Dic
lof 1*10^2 8 8 10
en
ate 1*10^3 10 14 14
sod
iu
1*10^4 12 12 22 18
m
1*10^5 12 12 34 30

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Abdul-Hussein Z. R. Antibacterial effect of Non-Steroidal…

1*10^1
1*10^2 6
Me 1*10^3 10
fen
am
1*10^4 14
ic
aci
1*10^5 12 16
d
1*10^1 8
Pir 1*10^2 10 8
oxi
ca
1*10^3 10 8
m
1*10^4 12 10
1*10^5 18 18
* Measurement of inhibition zone (mm)

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Basrah Journal of Science (B) Vol. 32(2), 166-181 ,2014

Aspirin gave the effect on all studied bacteria at high concentration and the diameter of
inhibition zone decreased with increasing of dilution .Table (1) figure(1)

1

1
[
[ T 4
2 T5 4 2 5
y
[ y [ [
p
T p3T T
3e
y e[y 3 y

p T p [ p
a

e BayaecilTlus 1 eStaphylococcus

p y [ q
a qea p 2 5T 4 a u
u e y[ [ o
q oaq 3 pT T q t
u t u a 1 ey y u e
o qeo p p o
t 1u t q Eea. coli e t f 1

e [ofe u e r [
2 T5tr o 4 qa a 4 o T5 2
[f yoe[ f t u f m y[ [
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yo [eyf o ut u o t e3y y
mp T prt m f oe o m h 1p p
e Klyeabohesi erlla t t Pseuedaoemonase
t pme t o ef e t
ah eqa h m r h d qa a
Figuree (1)Anu dtti ebact erial effect of aspirino f(salicylicf acid ) . meeasuremeontu o f diamete r of
inhibqit ion aohoq t zone mr of r eac h c oqconcentqration
1=1*u1d0^1,2= 1teuc*d10h^2,3=1*10^3,4=1*10^4,o5 =1*10^5o in mm. d u tu u
oo qeuo o e mt m o m eo o
tc umdt c h c e t t
eu ofoeeu d et t u n fe e
m trncm o h 172 h m t r
fe eoutfe c de e e of f
rn mm r n u o n o mr r
Abdul-Hussein Z. R. Antibacterial effect of Non-Steroidal…
Salicylate and related compounds such Pseudomonas aeruginosa (Al-Bakri, et al
aspirin are known to have a variety of effect .,2009).
on microorganisms (Price et al .,2000)
Another study showed that when
These effects include changes in membrane
aspirin administered to mice undergoing
potentials and virulence factors production,
treatment of tuberculosis infection
reduction in extracellular polysaccharide
(Mycobacterium tuberculosis) to determine
production (Price et al. 2000; Wang et al.
if these non-steroidal anti-inflammatory
2003)
drugs enhance pyrazinamide activity in
Aspirin possessed a broad spectrum vivo(Byrne et al .,2002).
antimicrobial activity against E. coli and
Diclofenate sodium revealed a significant effect against Gramm positive bacteria.
Pseudomonas not affected even at high concentration .Table (1) figure (2)

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Basrah Journal of Science (B) Vol. 32(2), 166-181 ,2014

1
1
[
[
2
T5
4 2 T5 4
3333 y 3333 y 3333 3333
p 3
3 3 p 3 3
e
3
e

1
a Staphylococ
Bacillus [ a

cus
T
q
5y q
u 4 p 3333 2 u

o 3333 3 e 3 o

t 3 1
t
e E. coli [
a e
T

f y
1 q f
r p
[ u r
o e
T o o 5 2
4 5 2
m 4
3333 y 3333 t 3333 m a 3333
3 p3 3 e 3 3
t 3
e t
Klebsihella Pseudoqmonas
f h

e u
a r e
o
Figure (2)Antiba cterial effect of voltarin o(diclofenate sodium ) . m easurement of
d t
q m d
diameter of inhibition zone of each concentration
o e
u o
1=1*10^1,2=1*10c^2,3=1*10^3,4=1*10^4,5=1*10^5 in mm.
o t c
u f
t h u
A time kill study of diclofenate come in part from its ability to inhibit the DNA
m r
e e m
synthesis of E. coeli and Listeria monocytogenes(Mazmudaret al .,2009) .Duttao et al .,(2007)
e
had demonstratedn that diclofenate had the ability to protect animal fromm lethality of
f d n
t
Salmonella . r o t
t
o c
o h
m u o
r e
m r
174
t e
t d
h n t
h o
t
Abdul-Hussein Z. R. Antibacterial effect of Non-Steroidal…
Ibuprofen has been inhibited the growth of both Gramm negative and Gramm positive
bacteria , as illustrated in table (1)and Figure (3)

1
1
[
[
2
T
T 5 2 5 4
4 y 22 y 22 22
22 2 3p 2 2
p
2 3 e 22
e
22 1 2

Staphylococ
2 [ a
Bacillus
a
T cus

2 5y 4 q
q
u 3 p 22 22 u
2
22 e 2 o
o
2 t
1t
a e
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2 5yf 4 Eq. coli 2 fT5 22
2
p 22 r 22 u ry 22

3 e 2 o 2 o 3op 2
22 m t m 22 e
2 Kleab siella e Pseu 2d omonas

t
t a
f h
qh
Figure (3)Antibacterial effect of ibuprofen . measurement of diameter of inhibition zone
r e
ue q
of each concentration 1=1*10^1,2=1*10^2,3=1*o10^3,4=1*10^4,5=1*10^5 in
o u
m d
td o
individual compounds ,which by turn
Ibuprofen belongs to propionic family of o
eo t
NSAIDs which consist of a phenol group in dtamage DNA and enzymces (Hensch et al.
c e
its structure . Thef uantibacterial effect of h.,2000; Boyd et al. ,2001) u.
e m
ibuprofen due to mrthe formation of free f
The less effectede compounds of
oe r
radicles and its hydrophobicity of the
d n
mn studied NSAIDs weore both of
o t
t 175 m
c
t
u o
ho t