Bul l et i n of Exper i ment al Bi ol ogy and Me di c i ne , No_ 3, 1 9 9 9 PHARMACOLOGYAND TOXI COLOGY 7 7 ~

Prolonged Bactericidal Effect of Sodium Hypochlorite

P. A. Galenko-Yaroshevskii, S. E. Gumenyuk,
S. G. Pavlenko, S. A. Babichev, V. Yu. Shevchuk,
S. I. Gumenyuk, and u G. Orlov
Translated f r om Byulleten' Eksperimental' noi Biologii i Meditsiny, Vol. 127, No. 3, pp. 305-307, March, 1999
Original article submitted February 3, 1998
Pol yvi nyl pyrrol i done prol ongs t he bactericidal effect of sodi um hypochl or i t e (NaOC1) by
25-30 t i mes. The ant i bact eri al act i vi t y of i mmobi l i zed NaOCI depends on t he pol ymer
concent r at i on i n t he solution. Excess of pol yvi nyl pyr r ol i done leads t o bl ockade of active
groups and reduces NaOCI activity.
Key Words: pol yvi nyl pyrrol i done; staphylococcus; Escheri chi a coli; sodi um hypochlorite
Hi gh i nci dence of nosocomi al i nfect i ons and mul t i -
drug r esi st ant strains of mi cr oor gani sms necessi t at e
search for new drugs and met hods for controlling puru-
l ent surgi cal infection. The met hod of i ndi rect electro-
chemi cal oxi dat i on wi t h sodi um hypochl ori t e (NaOC1)
is a per spect i ve t r eat ment . Thi s agent is hi ghl y ef-
fect i ve t owards purul ent surgical infection; it cancel s
antibiotic resi st ance of mi cr oor gani sms; the effects of
t he maj or i t y of antibacterial drugs are pot ent i at ed by
NaOCI; t he exo- and endot oxi ns of pat hogeni c mi cro-
or gani sms can be bound by sodi um hypochl ori t e [1,2,
4]. However , NaOC1 is an unst abl e compound, and its
t herapeut i c effect is l i mi t ed by several mi nut es, after
whi ch it rapidly degrades. This limits t he use of NaOC1
in l ocal t her apy of pur ul ent wounds.
We i nvest i gat ed t he possi bi l i t y of prol ongi ng t he
bact er i ci dal ef f ect of NaOC1 by i mmobi l i zat i on on
l ow- mol ecul ar - wei ght dext ran.
MATERI ALS AND METHODS
Sterile apyr ogeni c sol ut i ons of sodi um hypochl or i t e
wer e pr epar ed by el ect r ol ysi s of 0. 89% NaC1 in an
aut omat ed mode on an EDO- 4 devi ce (Regnatis, Mos-
cow). The concent rat i on of NaOC1 (600 and 1200 mg/
l i t er) was r egul at ed by changi ng t he exposur e and
Department of Pharmacology, Department of Surgery of Pediatric and
Dentistry Faculties, Department of Microbiology, Kuban State Medical
Academy, Krasnodar
current during electrolysis. The agent was mi xed (1:1)
wi t h sterile wat er sol ut i on of medi cal pol yvi nyl pyrro-
l i done (PVP) wi t h mol ecul ar wei ght of 12600__.2000
i n di fferent concent r at i ons at 18-20~ Under t hese
conditions compet i t i ve interactions of weakl y ori ent ed
anion groups l ed to fi xat i on of active oxygen on the
pol ymer [3].
Antibacterial act i vi t y of i mmobi l i zed NaOC1 was
i nvest i gat ed usi ng standard Escheri chi a coli J53 met
pr o- ZZ lac+ and Staphylococcus aureus 209 strains.
Cultures of standard strains were grown in oblique
meat - pept one agar ( MPA) for 16 h, and a bacterial
suspension (4x10 9 CFU in i sot oni c NaCI) was pre-
pared. The antibacterial effect of NaOCI depends on
t he initial concent rat i on of mi croorgani sms: the higher
t he concent rat i on of mi cr oor gani sms, t he hi gher t he
effect i ve concent rat i on of NaOC1. For si mul at i ng the
conditions in a purul ent wound. 5 ml NaOC1 wi t h and
wi t hout PVP was mi xed wi t h 0.2 ml fresh heparinized
human plasma.
The mi xt ure was i ncubat ed at 37~ Ior 24 h and
0. 5-ml aliquots t aken after certain intervals (15 min,
1. 6, 12. and 24 h) were i ncubat ed wi t h 0.1 ml bac-
terial suspension for 15 mi n at 37~ in a wat er bath.
The number of viable bact eri a was eval uat ed by the
met hod of serial dilutions after inoculation of the mix-
ture in dishes wi t h MPA (for S. aureus) or Endo me-
di um (for E. coli). Vari ous concent rat i ons of NaOCI
(600 and 1200 mg/liter) and PVP (0.5. 1, 2, 5, and
10%) were combi ned.
0007-4888/99/0003-0275522.00 9 Kluwer Academic/Plenum Publishers
276 Bulletin of Experimental Biology and Medicine, No 3 , 1999 PHARMACOLOGYAND TOXICOLOGY
o
5 H ~ 2 z,+
o , c n , r , , t o O o
' z 0.23 '
Fig. 1. CFU in a E. coil culture treated with NaOCI (600 rag/liter) and
pol yvi nyl pyr r ol i done.
PVP ~ @~ P VP
pvP PVP
Fig. 2. Bactericidal effect of NaOCI at polyvinyl pyrrolidone (PVP)
concentrations 1-2% (a) and >5% (b). BC: bacterial cell.
Bactericidal effect, %
100-
/
80-
/ H 1200 rag/liter
O---O 600 rag/liter
60
40-
20-
/
0 / 9 - ~=
2 4 6 8 10
Concentration, %
Fig. 3. Bactericidal effect of immobilized NaOCI for E. coil culture at
di f f er ent concent r at i ons of po~yvinyl pyrro~idone. Exposur e 60 min.
RESULTS
Whe n standard E. col i J53 met pr o- ZZ l ac+ strain was
i ncubat ed wi t h NaOC1 wi t hout PVP t he antibacterial
act i vi t y of t he sol ut i on di sappeared wi t hi n 1 h (Fig.
1). Addi t i on of PVP pr ol onged the ant i bact eri al ac-
t i vi t y of NaOC1 i n a dos e- dependent manner. In a
concent rat i on of 0. 5% PVP pr ol onged the agent ac-
t i vi t y t o 6 h. The maxi mum effect was obser ved at a
PVP concent rat i on of 1%, when the drug act i vi t y de-
creased onl y after 24 h. Furt her i ncrease in PVP con-
centration in the mi xt ure r educed its antibacterial ac-
tivity. Mi xt ures cont ai ni ng mor e t han 5% PVP pos -
sessed no antibacterial act i vi t y.
Similar results wer e obser ved wi t h S. aur e us cul-
tures, t hough the ant i bact eri al act i vi t y of i mmobi l i zed
NaOCI was l ower by an order of magni t ude. In the
control series, isotonic NaC1 did not suppress the viabi-
l i t y of bacteria. Cont r ol tests wi t h PVP in di fferent
concentrations wi t hout NaOCI s howed that the number
of CFU after exposur e of bacterial suspensi ons for 15
and 30 mi n di d not depend on the concent rat i on o1"
PVP and was vi rt ual l y t he same as i n i sot oni c NaCII
Hence, the bact eri ci dal effect of NaOCI depends
on t he concentration of PVP in the solution. The effect
of 1-2% PVP can be due t o bi ndi ng or bact eri a and
t oxi ns t o the carrier enabl i ng their cl ose cont act wi t h
the oxi dant (Fig. 2, a). The bact eri ci dal effect in this
case is due t o i mmedi at e effect of t ree NaOCI in the.
sol ut i on and pr ol onged effect of a rel at i vel y unst abl e
compl ex of PVP and hypochl or i t e di ssoci at i on pro- -
duct s ( hypoc hl or ous aci d HOC1 and h y p o c h l o r i t e .
i on OC1-). Supersat urat i on of the sol ut i on wi t h P VP
i ncreases cross-l i nki ng of act i ve pol ymer groups wi t h
NaOCI di ssoci at i on pr oduct s and yi el ds a tight matrix
wi t h bl ocked active groups. Theret ore, bacterial cells
cont act onl y wi t h surface act i ve groups in this com-
pl ex (Fig. 2, b), whi ch was conf i r med in experi ment s
wi t h di fferent concent rat i ons of NaOCI (Fig. 3).
Thus, pol ymeri c carrier considerably (25-30 times)
prol onged t he bact eri ci dal effect of NaOCI, whi l e an
excess of the pol ymer i c carrier in the sol ut i on bl ocked
act i ve groups and sharpl y decr eased antibacterial ac-
t i vi t y o f the compl ex. Pol ymer - i mmobi l i zed NaOC1 is
a pr omi si ng agent t or medi ci ne: pr ol ongat i on of its
effect for more t han 24 h makes i t a useful drug for
the t reat ment of i nt oxi cat i ons and in pyosept i c sur -
gery, part i cul arl y i n i nfect i ons caused by anaerobi c
microflora.
In t he control, NaOCI was r epl aced wi t h 0. 89%
NaC1. In order t o rul e out t he nonspeci f i c effect s of
PVP on bact eri al cel l s (t br exampl e, coaggl ut i nat i on),
t he number s o f CFU i n bact eri al suspensi on i n i so-
t oni c NaCI and PVP wer e compared.
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