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`VOL 123 no.3 MARCH 1998
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`

`

`Analyst, March 1998, Vol. 123 (503-507)
`
`503
`
`Effects of copper and zinc ions on the germicidal
`properties of two popular pharmaceutical antiseptic
`agents cetylpyridinium chloride and povidone-iodinet
`
`Jacobus J. Zeelie* and Terrence J. McCarthy
`Unit f or Health Services, Port Elizabeth Technikon, Private Bag X6011 , Port Elizabeth, South
`Africa 6000
`
`The effects of copper and zinc ions on the rate of killing
`of Gram-negative bacterium Pseudom.onas aeruginosa,
`Gram-positive bacterium Staphylococcus aureus and
`fungal yeast Candida albicans by antiseptic agents
`cetylpyridinium chloride and povidone-iodine (Betadine)
`were investigated. In the 48 test cases copper and zinc
`ions clearly potentiated the antiseptic agents in 28 (58.3 % )
`cases and exhibited an improved (not clear potentiation)
`activity in 15 (31.3 % ) cases. In five (10.4 % ) cases there
`was no change in the antiseptics' antimicrobial activity. In
`general zinc potentiated the antiseptic agents more than
`copper. If an 'improved activity' was the only criterion
`for this study, then a more rapid antimicrobial effect was
`observed in 43 out of the 48 test cases, i.e., 90%.
`Keywords : Copper; zinc; cetylpyridinium chloride;
`povidone-iocline; antimicrobial; antiseptic
`
`Copper and zinc are two trace metals for which there are dail y
`recommended allowances (RDA). They are also well known for
`their antimic robi a l properti es . Copper is commonl y used as an
`antim icrobial agent in swimming pools and elsewhere whereas
`zinc is used phannaceuticall y in creams, ointments, eye drops,
`etc. to combat vari ous types of infection. In the past vari ous
`trace metals were used quite ex tensively as an tiseptic agents,
`but they have had to make way fo r more popular synthetic drugs
`such as antibiotics and other e laborate antimicrobial agents.
`These an tibi oti cs and an tiseptic agents are we ll known to cause
`allergies and a lso to allow resistance to develop in some micro(cid:173)
`organisms. They are also very ex pensive. To overcome
`microbial res istance doses are often increased to levels which
`simply aggravate (he problems already mentioned. Alter(cid:173)
`natively combinati ons of antimicrobial agents are not too
`infrequentl y emplo yed . The consequence is all too often a
`higher incidence of untoward effects. On the other hand copper
`and zinc combinations such as the sul fa tes are very well tested
`and allergies are quite unknown to members of the h_ealt_h
`profess ions. In this study an attempt wa made to e tabltsh if
`there ex ists antimicrobial interaction between these metal ions
`and two othe r pop ul ar antiseptic agents whi ch are used quite
`widely because of their w ide antimicrobial spectra and low
`incidence of side e ffects. T he antiseptic agents selected were the
`quatern ary ammonium compound cetylpyridinium chl oride and
`the orga nic iod ine compound pov idone-iodine. The effects of
`the meta l ions on these antiseptic agents were tested aga111st a
`typica l Gram-positive bacterium (Staphylococcus aureus),
`Gram-negative bacterium (Pseudomonas aeruginosa) and a
`fungal yeast (Candida albicans). All the microbial species were
`strains isolated from hospi talised patients with cond1tt on~ wh ich
`would not respond to the usual antibiotic therapy for 111f-ecttons
`caused by these micro-organisms.
`
`t Presented at The Sixth Nordic Symposium on Trace Elements in Human Health and
`Disease, Roskildc, Denmark, June 29- July 3, 1997.
`
`Experimental
`The concentration necessary for each of cetylpyridinium
`chloride and povidone-iodine to kill an inoculum of l X !06
`micro-organisms within 40 min, but not before 30 min, was
`determined. This value will be referred to as the minimum
`Increasing
`microbicidal concentrati on3014o or MMC30J40·
`amounts of the metal ions were then added to each of the
`antiseptic agents and the time to kill the same number of micro(cid:173)
`organisms noted. The micro-organisms were standardised
`spectrophotometricall y to contain 1 x 106 micro-organisms per
`0.0001 dm3 of the culture medium.
`
`MMC30140 determinations for the antiseptic agents
`(l ) Seri al dilutions were made in Normal Saline (the reacti on
`medium) for each of the antimicrobial test substances. These
`were sterilised at I l 5 °C for 30 min in an autoclave. (2) Each
`dilution was inoculated with I x I Q6 of the appropri ate micro(cid:173)
`organisms and kept at 37 °C in an incubator. (3) At JO min
`intervals and for 40 min subcultures were made into Tryptone
`Soya Broth whi ch contained 3% m/v Tween 80 as neu tralising
`agent (the recovery medium). A level of 3% m/v Tween 80 has
`been shown by several researchers 1- 5 to neutralise many
`antiseptic agents without inhibiting or ki lling the micro(cid:173)
`organisms themselves. In prelimi nary tests this concentr~tion
`was fo und to have no inhibitory effects on the te t m1cro(cid:173)
`oroanisms used in thi
`study. The subcu ltures were then
`in;ubated at 37 °C for 24 h. (4) After incubation the di lutions
`were visuall y checked fo r growth (optical density) .
`
`Minim.um lethal concentrations for the metal ions
`
`This test was necessary to avoid the erroneous assumption that
`a particul ar antiseptic-metal ion combinati on kil led a microbial
`populati on whereas the microbi cidal effects may have _been
`effected by the metal ions present and not necessan ly a
`combination of metal ions and antiseptic agents. The test was
`essentiall y the same as for the antiseptic agents, ~ut the total
`time period for subculturing wa 48 h. The assumption was that
`the minimum amount whi ch ki lled onl y after 48 h would not be
`ab le to kill within a time period of 40 min. T he ions were used
`as sulfate salts (copper sulfate and zinc sulfate). These salts
`were solu ble at the concentrations tested.
`
`Interactions
`( I) Solutions in Normal Sali ne containing the MMC30/40 fo r the
`antiseptic agents plus 5, I 0, 50 ~r 100 X I 0- 3 g dm3 of the meta l
`ions were prepared fo r each antisepti c agent, fo r each me~al salt
`and for each micro-organism. T hese solut1on_s ~ere stenltsed at
`l 15 °C fo r 30 min. The stock solutions conta1111ng the antisepti c
`agents and the metal salts were sterilise_d to e limmate the
`presence of possible resistant extraneous m1c~o-or~amsms and/
`or spores. (2) The dilutions containing the anttsept1c agents and
`
`

`

`504
`
`Analyst, March 1998, Vol. 123
`
`metal alts were inoculated with I x 106 of the appropriate
`micro-organism. (3) The inoculated dilutions were kept at 37 °C
`in an incubator. (4) At 10 min intervals 0.0001 dm3 amples
`were subcultured into the recovery medium and the latter
`reincubated at 37 °C for 24 h. (5) The subcultures were then
`visuall y examined for turbidity indicating microbial growth.
`The reason for choosino a reaction temperature of 37 °C ts
`because the selected anti~eptic agents are used on the skin and
`in body cavities where the average temperature comes close to
`37 °C.
`
`Results and Discussion
`All the micro-organi ms appeared to be sensitive to both
`Betadine (which contains I % m/v of povidone-iodine) and
`cetylpyridinium chloride (Table l and 2).
`The antimicrobial effects of the metals appeared to be
`particularly hi gh against Staphylococcus aureus. This Gram(cid:173)
`positive bacterium has been found in numerous inve ti gative
`studie
`to be very fragile and sensitive to most antimicrobial
`agents such as antiseptics, preservatives, disinfectants, ere.,
`currentl y on the market. The exception, however, is antibiotics.
`Strains resistant to several antibiotics are constantly emerging in
`hospitals where many medical staff members are healthy nasal
`carriers thereof. There are currently numerous reports on the
`resistance of Staphylococcus aureus to antibiotic agents. The
`apparent resi tance of the Gram-negative microbial species
`(Pseudomonas aeruginosa) is not unex pected. The resistance of
`this bacterium, especially in organic tissue , is very well
`documented. It is a bacterium which can survi ve on simple
`inorganic chemicals, being able to convert them to more
`elaborate organic requirements. Its ability to mutate is eq uall y
`well documented in the literature. The reason why this micro(cid:173)
`organism appear
`to be resistant to zinc ions, but not equall y
`resistant against copper ions, could be attributed to the fact that
`zinc preparations are over-the-counter pharmaceutical
`fo r
`which a prescripti on is not required. They are often applied to
`the skin or mucosa of the eye fo r purpo es of tissue regeneration
`
`and combatting infecti ons, whe reas copper-containing prepara(cid:173)
`tions are not that often used_. This constant exposure of
`Pseudomonas aeru?tnosa to z mc may be the cause of the
`development of re~1stance. Pseudomon_as aeruginosa is found
`on the human skm wh_ere 1t o_ften mfects wounds. Auto(cid:173)
`infec tt ons of the eye (pmk-eye 111fect1on) 1s also very com(cid:173)
`mon.
`The information in Table 3 is important to avoid mis(cid:173)
`interpretation of the obse rved killing effect by metal ion(cid:173)
`antiseptic agent combinati ons (Table 4 and 5). It should be
`clearly established tl1at the combination of agents killed the
`inoculum and not the meta l ions present as the latter also exert
`an antimicrobi al effect on the ir own.
`
`Copper
`5 X J0-3 g dm- 3: This level of copper ions would not be able
`to kill any of the micro-organisms within a 40 min period (see
`above for minimum letha l concentrations). The ki.lling effect of
`cetylpyridinium chloride against the yeast remained unchanged,
`but in the case of Betadine
`trong potentiation was evident.
`Cetylpyridinium chloride retained its original killing effects on
`Pseudomonas aeruginosa. Both anti septic agents were po(cid:173)
`tentiated against Staphylococcus aureus.
`10 X 10-3 g dm- 3: In all instance the micro-organisms were
`killed over the whole testing period of 40 min. Since this level
`of copper could not have killed Pseu.domonas aeruginosa and
`Candida albicans, a potentiated effect is obvious. The improved
`ki ll ing effect noted for Staphylococcus aureus could have been
`due to the antimicrobial properties of copper on its own against
`this micro-organism (minimum leth a l concentration being 6 X
`I 0-3 g dm-3) and potenti ation is therefore not clearly indicated.
`50-100 X J0-3 g dm -3 : T hese levels of copper, ions in
`combination with the antiseptic agents, did not allow growth to
`take pl ace at 5 min or longer. Once agai n, the improved effect
`is not nece sarily potentiation as the minim um lethal concentra(cid:173)
`ti ons for copper aga inst all the micro-organisms are lower than
`50 X 10-3 g dm-3 .
`
`Table_ 1 Minimum microbicidal concentrations30100 (MMC3or•o) at 37 °C for Betadine and cetylpyridin ium chloride agai nst three pathogenic micro·
`organisms•
`
`Antiseptic agent
`
`Betad ine
`(I% m/v povidone-iodine)
`
`Time/min
`
`Cetylpyridinium chloride
`
`Time/min
`
`IO
`
`20
`
`30
`
`40
`
`+
`+
`+
`+
`
`+
`+
`+
`+
`
`+
`+
`+
`+
`
`+
`+
`+
`
`+
`+
`+
`+
`
`+
`+
`+
`+
`
`+
`+
`
`+
`+
`+
`+
`
`+
`+
`+
`+
`
`•
`
`+
`
`+
`
`+
`
`10
`
`+
`+
`+
`+
`+
`
`+
`+
`+
`+
`
`20
`
`+
`+
`+
`+
`+
`
`+
`+
`+
`
`30
`
`+
`+
`+
`+
`+
`
`+
`+
`
`+
`+
`
`40
`
`(+)
`(+)
`(+)
`+
`
`+
`
`+
`
`Concentration
`tested (% m/v)
`3.600
`3.500
`3.488
`3.486
`3.484
`3.482
`5.038
`5.036
`5.034
`5.032
`5.030
`3.5
`3.4
`3.3
`3.2
`3. 1
`
`Pseudomonas aeruginosa
`
`Sraphylococcus a11re11s
`
`Candida albicans
`
`•
`
`+ =
`
`Concentrntion
`tested (% m/v)
`0.056
`0.055
`0.054
`0.053
`0.052
`0.051
`0.00040
`0.00038
`0.00036
`0.00034
`0.00032
`0.0025
`+
`+
`0.0020
`+
`+
`0.0018
`+
`+
`0.0016
`+
`+
`0.0015
`+
`+
`G
`0.0014
`rowth, - = no growth • = MMC
`·
`· ·
`-
`'
`30/40 - minimum concentrauon which kills after 40 min , but not after 30 min.
`
`

`

`Zinc
`5 x I o-3 g dm-3: This level of zinc ion on its own did not kill
`any of the micro-organisms in the absence of other antiseptic
`agents. Any improved effect noticed would therefore be due to
`potentiation. Cetylpyridinium chloride retained its original
`killing effects.against Candida albicans. The effect of cetylpyr(cid:173)
`idinium chlonde on Staphylococcus aureus also remained the
`same. Potentiation was strong in all other test cases.
`IO x J0- 3 g dm- 3: A potentiated effect was found for all the
`antiseptic agents against Candida albicans and Pseudomonas
`aeruginosa. Since thi s level of metal ion could have had a
`killing effect of its own against Staphylococcus aureus, the
`improved activity throughout does not necessarily indicate
`potentiation .
`
`Table 2 Summary of the minimum microbicidal concentrations30140
`(MMC30140) at 37 °C for Beradine and cetylpyridinium chloride against
`three pathogenic micro-organisms•
`
`Minimum contact killing concentration30140
`
`Antimicrobial agent
`
`S. aureus
`P. aernginosa
`C. albicans
`3.484
`3.200
`5.032
`Betadine
`0.00034
`0.055
`Cetylpyridinium chloride
`0.0015
`• P. aemginosa = Pseudomonas aeruginosa, S. aureus = Staphylo(cid:173)
`coccus aureus, C. albicans = Candida albicans; MMC30140 = Minimum
`concentration which ki ll s after 40 min , but not after 30 min.
`
`Table 3 Minimum lethal concentrations after 48 h at 37 °C for copper and
`zinc ions agai nst three pathogenic micro-organisms•
`
`Minimum lethal
`concentrat ions/
`X IQ- 3 g dm-3
`Metal ions
`
`Highest test
`concentration
`which did not kill/
`X !Q-3 g dm-3
`Metal ions
`
`Micro-organism
`Pseudomonas aerugi11osa
`19 17
`36
`Staphylococcus aureus
`9
`8
`Candida albicans
`39
`26
`• The above values are fo r the meta l ions Cu++ and Zn++ (as present in
`the salts CuS04·5 H20 and Z nS04·7H20).
`
`Cu++
`
`Zn++
`
`Cu++
`
`34
`6
`24
`
`Zn++
`
`1906
`8
`38
`
`Analyst, March 1998, Vol. 123
`
`505
`
`50 X I0-3 g dm-:: A better, improved killing effect was
`found throughout. This level of ion did kill Candida albicans
`and Staphylowccus aureus in previous studies, therefore any
`obs.erved killing effect better than
`that obtained for the
`antiseptic agents on their own, should rather be described as
`tmprov~d than. potentiated. Nevertheless, turbidity indicating
`mtcrobtal multtphcat10n was only present for 5 min in the case
`of cetylpyridinium chloride against Staphylococcus aureus. In
`all of.the remaining combinations potentiation was very strong
`as evidenced by the total absence of turbidity over the whole
`testing period.
`100 X L0- 3 g dm- 3: In all the cases growth was absent from
`5 mm onwards. This indicates the same very strong potentiating
`effect as when 50 X I 0-3 g dm-3 of the zinc ion was present.
`The growth observed for cetylpyridinium chloride against
`Staphylococcus aureus was completely absent.
`Several authors6--8 noticed that copper can carry certain
`substances which are extracellularly nontoxic into the microbial
`cell where they become intracellular toxins because of the fact
`that their interaction sites are inside and not on the cell. This
`effect was found to be more pronounced in Gram-positive
`micro-organisms as compared with Gram-negative bacteria. In
`this study, however, a better effect was noticed for the Gram(cid:173)
`negative bacterium and this mechanism is perhaps unlike ly.
`Several other mechanisms could possibly account for the
`increased ki.Uing effects found. Several
`in vesti gators7,9.to
`remarked that copper can deplete microbial cells of magnesium
`and the latter is essential for protective cell wall formation . The
`metal is also known to bind with phosphate groups. 11 A much
`better improvement of the overall killing effect was noticed for
`the Gram-negative bacterium as compared with the Gram(cid:173)
`positive species. One should also bear in mind the possibility
`that the oversupply of any one metal may not only deplete the
`microbial cell of other metals e sential for normal cell
`membrane structure and function as well as enzyme activity, but
`that it may also alter the cell 's need fo r other metal ions and in
`the process render the cell sensitive to damag ing substances
`such as antiseptic agents or even metal ions themselves.9, t2-t4 It
`may be possible that exces copper could affect the zinc
`requirement for dehydrogenase enzymes and the hyd rolys i of
`phosphates and peptide , all of which are found on the cell wall
`surface. Such an effect may render the cell more permeable to
`antiseptic agents and intracellular solutes. Lastl y, two research
`groupsts.16 proposed that polyvalent metal ions can cau e
`effecti ve charge neutralisation on microbial cells and this may
`lead to improved attachment and penetration of some antiseptic
`
`Table 4 Effects at 37 °C of copper and zinc ions on the minimum microbicidal concen1 ra1ions301•o (MM C301•0) at 37 °C of Betadine and cetylpyridinium
`chloride·
`
`Metal ion concentration/
`X IQ-3 g dm -3
`
`10
`
`20
`
`30
`
`40
`
`IO
`
`20
`
`30
`
`40
`
`10
`
`20
`
`30
`
`40
`
`10
`
`20
`
`30
`
`40
`
`Copper
`
`10
`
`Time/min
`
`5
`
`Z inc
`
`10
`
`+
`+
`
`( +)
`( +)
`
`( +)
`
`Antimicrobi al agents
`Betadine-
`P. aeruginosa (3.484)
`S. aureus (5.032)
`C. albicans (3 .200)
`Cetylpyridinium chloride-
`P. aeruginosa (0.055)
`(+)
`S. a11re11s (0.00034)
`+
`+
`+
`+
`( +)
`C. albicans (0.00 15)
`+
`+
`(+)
`+
`+
`(+)
`+
`+
`• + Growth; - = no growth; (+) = fai nt growth ; P. aem ginosa = Pse1tdomonas aeruginosa , S. a11re11s = Swphylococcus aureus, C. albicans =
`(appear in brackets) = minimum concentration which kills after 40 mm , but not after 30 mm; Note: Concentrnt ,ons of 50 and
`Candida albicans; MMc
`30140
`I 00 X I 0- 3 g dm-3 of the metal ions allowed no growth in all test cases.
`
`(+)
`
`(+)
`
`

`

`506
`
`Analyst, March 1998, Vo l. 123
`
`agents to the microbial cell. It is likely that more than one
`mechanism is involved and that the improved antimicrobial
`effects observed are the result of a combination of mechanisms.
`Zi nc ions exh ibited no antagonistic effect on the antimicrobial
`agents and an improved acti vity was observed in most of the
`combinations. Excellent potentiation was noted in several cases.
`The ki lling effects of zinc were mostl y the same as those
`observed fo r copper. In the case of copper, a better kilJi ng effect
`was noted fo r 42 out of 48 cases, whi le an improved kjlling
`effect was fo und fo r zinc in 45 out of 48 cases. No antagonistic
`effects were observed fo r these two metal ions. All micro(cid:173)
`organisms seemed to be affected adversely by the presence of
`copper and zinc ions. Zinc has been described as a carrier fo r
`oxine to obtai n an improved antifungal effect. 17,18 It was found
`in these studies that zinc improved the antifungal effects of the
`quaternary ammonium compounds, but not of Betadine. It is
`obvious that some sort of phys ical and chemical interaction
`between zinc and the antiseptic agent is of importance fo r this
`mechanism
`to apply. Z inc can displace metals such as
`magnesium1, 10.19 and copper9,12- 14 from microbial cells. The
`effect of magnesium depletion (as well as other metal ions) fro m
`a microbial ceU was discussed earlier. According to this
`mechanism, Gram-negati ve bacteria should be more sensitive
`than Gram-positive bacteria. This was fo und for the quaternary
`ammonium compounds against the bacteria (not the yeast), but
`not at all
`for benzyl alcohol and Betadine. The charge
`neutral isation effect described for copper may hold true for zinc
`
`as well. If zinc depleted the microbial cells or other essential
`me'.als, copper could be one of them. Copper is involved in the
`act1v1ty of some enzy mes such as ox1dases. 14 According t
`0
`Eagon and As bell , 19 zinc can in hibit some step of the ener
`transfer cycle which_ is involved in the transport of substra~~
`catalysed by magnesium . It can ind uce confo rmational changes
`111 the tertiary structu res of cell mem brane proteins involved in
`substrate transport.
`
`Conclusion
`All the types of antiseptic-metal ion interactions observed can
`be described as fo llows, taking into account that (I) the
`antiseptic agents should allow microbial growth for a period of
`up to 30 min, but not 40 mi n, and that (2) metal
`ion
`concentrations above the minimu m lethal levels as previously
`determined fo r a 48 h incubation period could have produced
`although unli kely over a 40 m in test period, an antimicrobial
`effect of their own.
`If the metal
`the minimum lethal
`is below
`level
`ion
`concentration and the possibility of an antimicrobial effect from
`the metal therefore unlikely, one can use the following as an
`- = excellent
`indication of the degree of interaction: -
`-
`-
`potentiation (EP); + - - - = good potentiation (GP); ++ - - =
`moderate potentiation (MP).
`the minimum lethal
`is above
`If the metal
`ion
`level
`concentration and the possibility of an anti-microbial effect
`
`Table 5 lnteracrion effects at 37 °C of copper and zinc ions on the min imum microbicidal concentrations30140 .(MMC30140) of Betadine and cetylpyridinium
`chloride*
`
`Metal ion concentration/X
`
`LQ-3 g dm- 3
`
`Copper
`
`Zinc
`
`5
`
`u
`MP
`EP
`
`10
`
`EP
`IA
`EP
`
`50
`
`EP
`IA
`IA
`
`100
`
`EP
`IA
`IA
`
`5
`
`EP
`EP
`EP
`
`Antimicrobial agents
`Beradine-
`P. aeruginosa (3.484)
`S. a11re11s (5.032)
`C. albicans (3.200)
`Cetylpyridinium chloride-
`EP
`EP
`EP
`EP
`EP
`EP
`EP
`U
`P. aeruginosa (0.055)
`GP
`EP
`GP
`U
`IA
`lA
`IA
`GP
`S. a11re11s (0.00034)
`EP
`EP
`U
`IA
`IA
`EP
`U
`C. albicans (0.00 15)
`MP
`= goo potent1at1on, EP = excellent potenuauon, IA = 1mprovemen1;
`• A = anta0 onism U = unchan °ed effect MP = moderate pote 1· 1· GP
`·
`·
`·
`· ·
`d
`.
`.
`"_
`•
`. " .
`•
`n 1a ion,
`P. ae, ugmosa - Pseuc:omonas ae, ug,nosa, S. aureus = Staphylococcus aureus, C. albicans = Candida albicans· MMC
`. appear in bracke1s = minimum
`3
`o14o
`concentration which ki lls after 40 mm, but 1101 after 30 min.
`'
`
`LO
`
`EP
`IA
`EP
`
`50
`
`EP
`IA
`IA
`
`100
`
`EP
`IA
`IA
`
`Ta ble 6 Summary of the effects of copper and zinc ions on the microbial ki ll ing rates of Betadine and cetylpyridinium chloride*
`
`Copper ions
`
`Zinc ions
`
`Polentiation
`Improved action
`Unchanged
`Potentiation
`Improved action
`Unchanged
`
`Betadine
`6
`5
`I
`7
`5
`0
`
`Cetylpyridinium chloride
`5
`5
`2
`10
`0
`2
`
`Copper and Zinc
`
`Potentiation
`13
`Improved action
`10
`Unchanged
`I
`• Summary: increased killi ng rate (potentiation + improved action):
`No effect:
`
`+
`+
`+
`23
`I
`
`· ·
`. Q
`f
`• (N
`uantmes o 5, I 0, 50 and I 00 x Io-, g dm -3
`ote.
`aemginosa, Sraphylococcus aureus and Candida a/bicans).
`
`Total
`28 (58.3%)
`LS (3 1.3%)
`5 ( 10.4%)
`43
`(89,6%)
`..2.
`(10.4%)
`Total: 48
`(1 00%)
`h ·
`f
`-
`0 eac
`,on were tested on the above anti septic agents' abilities to ki ll Pseudomonas
`
`15
`5
`4
`+
`+
`
`20
`4
`
`

`

`from the metal thus possible, although unlikely, over a 40 min
`testing period: -
`-
`-
`or + -
`-
`or ++ - would indicate an
`improved antimicrobial action (IA).
`If there is microbial growth for up to 30 min, but not at 40
`min: +++ - = unchanged effect (U).
`Growth throughout the 40 min testing period: ++++ =
`antagonism (A) .
`The various types of interaction can now be described as one
`of the fo llowing: E xcellent potentiation = EP, - - - -; Good
`potentiation = GP, + - - -; Moderate potentiation = MP, ++(cid:173)
`- ; Improved action = IA, - - - -/+ - - -/++ - - ; Unchanged
`effect = U, +++ - ; Antagonism = A, ++++. This scaling of
`responses is used in T able 5.
`Table 6 gives a summary of the antim icrobial activities of
`combinations of the vari o us antiseptic agents with copper and
`with zinc ions. For copper ions, in general, an excellent
`improvement in the killing activity of the antimicrobial agents
`was observed in most cases. No true antagonism was observed.
`It can be seen that a more rapid microbial killing rate ..yas
`observed in about 90% of test cases. True potentiation was seen
`in about 58% of cases and an improved action in about 31 % of
`cases.
`Antimicrobial agents such as povidone-iodine (e.g. , Beta(cid:173)
`dine, Podine) and cetylpyridinium chloride (e.g., Savlon,
`Cetavlon) are non-specific cytotoxic agents which may cause
`untoward effects like skin or mucosa! irritation, rashes and
`allergies, even at
`tested
`in-use concentrations
`in certain
`individ uals. Coppe r and zinc ions in the form of the metal salts
`tested are well known fo r their antimicrobial properties as
`pharmaceutical agents against (bacterial , fungal, viral , proto(cid:173)
`zoa!) infections of the skin, eyes, etc. If small quantities of these
`metal salts can be employed in pharmaceutical fo rmulations
`containing povidone-iodine and cetylpyridinium chloride,
`smalle