Patent Document:

broadly , the present invention contemplates the use of an activating inhibitor in a stabilized chlorine dioxide mixture to make possible the lowering of the ph of the chlorine dioxide mixture at the time it is used in the mouth . clinical observations by the inventor have led to the discovery that an activating inhibitor phosphate such as disodium hydrogen phosphate , sodium dihydrogen phosphate , or , preferably , trisodium phosphate , will allow the stabilized chlorine dioxide to remain stable and effective at a lower ph than has been thought possible . in addition , the phosphate is a detergent which , when used in place of the detergent sodium laurel sulfate , provides a nonsudsing toothpaste . nonsudsing toothpastes are preferred for use with electric toothbrushes because electric toothbrushes tend to spray sudsing toothpastes about the room during use . further , people tend to quit brushing when a mouth full of suds accumulates in the mouth ; a nonsudsing toothpaste does not cause this psychological event . the first step in the formation of plaque on a clean tooth surface is the formation of acquired pellicle . studies by others have shown the following to be part of the acquired pellicle formative process . glycoproteins of salivary and other mucous gland origin are attached to the hydroxyapatite crystals . ( roukima , p . a . and nieuw amerongen a . v ., sulphated glycoproteins in human saliva ; saliva and dental caries ( sp . supp . microbiology abst .) 1979 , pp . 76 , embery , g ., the role of anionic glycoconjugates , particularly sulphated glycoproteins in relation to the oral cavity , saliva and dental caries . ( supp . microbiol abstr . ), information retrieval 1978 , pp . 105 - 111 ). sulphated glycoproteins have a strong affinity to the calcium anion ( ibid ., pp 105 - 108 ). most major salivary secreted glycoproteins may be bound to certain ester sulphates ( ibid ). these sulphated glycoproteins have been related to bacterial agglutination or clumping ( ibid ., pp 108 ). clinical observations by the inventor have led to the discovery that the process of acquired pellicle can be inhibited by the use by humans of stabilized chlorine dioxide . through such observations it has been learned that the chlorine dioxide reacts with the sulphated glycoproteins to inhibit pellicle formation . this process results primarily from , but is not limited to , oxidation of the sulphide bonds . since acquired pellicle is the first step in plaque formation , this initial inhibition alters the sequence of events to follow . the second step , bacterial adhesion , and subsequent steps are consequently retarded . no disulphate enzymes capable of cleaning the sulphate moieties of glycoproteins are known . bacterial agglutinigation includes the conversion of sucrose to glucans and fructrans by enzymes known as glycosyltransferases . these enzymes are of bacterial origin . the plaque mass becomes a complex extra cellular ( of microorganisms ) matrix containing sulphated glucosamineglycans , proteglycans , glycoproteins , sugar , proteins and lipids , which aid in the process of bacterial agglutination ( schluger , s ., yuodelis r . and page r ., periodontal disease , chapter 6 , pp . 135 to 166 , 1977 , lea and febiger , phila , pa . newbrun e ., polysaccharide synthesis in plaque ; microbiol aspects of dental caries , vol iii ( supp . microbiology abstr . ), 1976 , pp 649 - 664 .) these compounds include the presence of sulphur and become unstable in the presence of high oxygen compounds . the oxygen splits the sulphide bonds to form sulphates or so 2 . clinical observations by the inventor have led to the conclusion that all of these biochemical compounds are attacked to a greater or lesser extent by stabilized chlorine dioxide . since these compounds may be used as nutrients for bacteria , the reduction of the compounds will inhibit bacterial growth . more specifically , the stabilized chlorine dioxide oxidizes carbohydrates , chondroitin sulphates , glucosaminglycans , glycoproteins , proteins and lipids . since these compounds arise as bacterial byproducts and debris from dead and dying cells , are of salivary origin and are the mechanism of agglutination of the plaque mass , their degradation / oxidation retards plaque growth . the initial bacterial residents of the plaque mass are aerobic , oxygen using organisms . the saliva bathing the plaque provides the source of oxygen . as the plaque thickens and the oxygen using bacteria increases in numbers , the deeper layers have a reduced oxygen content . the greater the aerobic population of plaque matrix , the lower the oxygen level in the saliva . this permits the deeper layers of the plaque matrix to develop an anaerobic population of bacteria ( globerman , d . y ., and kleinberg , i ., intro - oral po 2 and it &# 39 ; s relation to bacterial accumulation on the oral tissues : saliva and dental caries . ( sp . supp . microbiology abstr .) 1976 pp . 275 - 292 ). clinical observations by the inventor lead to the discovery that the use of stabilized chlorine dioxide with an activating inhibitor in an oral health preparation such as a mouth wash or toothpaste will raise the level of oxygen in the saliva . the raised level of oxygen within the plaque matrix will inhibit anaerobic bacterial growth . as periodontitis is caused by anaerobic bacteria , the potential for the development of periodontitis is reduced by stabilized chlorine dioxide . the inhibition of acquired pellicle formation , the prevention of bacterial agglutinization and the oxidation of the plaque mass through the use of chlorine dioxide with an activating inhibitor phosphate in a mouth wash or toothpaste are independent of the germicidal capacity of such mouth wash or toothpaste . furthermore , these factors in combination with the bacteriocidal capacity of chlorine dioxide in the mouth wash or toothpaste renders the mouth wash or toothpaste an effective pellicle and plaque inhibitor . the permeability of sublingual mucous tissue within the mouth is increased substantially by exposure to hydrogen sulfide ( h 2 s ) and methyl mercaptan ( ch 3 sh ). ( gaffer and rizzo papers referenced in &# 34 ; effect of hydrogen sulfide and methyl mercaptan on the permeability of oral mucosa &# 34 ;, j . dent res . 63 ( 7 ), jul . 1984 , pages 994 - 997 ). accordingly , the toxic bacterial products attendant plaque which produce these compounds have a related effect on tissue permeability . since chlorine dioxide breaks the disulphide bonds of both these compounds , the use of chlorine dioxide with an activating inhibitor phosphate in a toothpaste would reduce the penetration potential of pathogenic materials . evidence exists that endotoxin and lipopolysaccharide from gram negative bacteria are the worst of the products to penetrate the tissues . application of endotoxin to gingiva has caused gingival inflammation . ( ibid ). chlorine dioxide used in treatment of plaque acts upon attendant gram negative bacteria . thereby , the inventor has learned through experimentation and observation that chlorine dioxide with an activating inhibitor phosphate in a mouth wash or toothpaste can be a preventative product leading to oral health . the us and efficacy of this composition is described below . the stability of chlorine dioxide at ph 6 . 8 in the presence of phosphate . 1 . purogene ( 2 % clo 2 ), lot # 8907 . 41 , 1 gallon , manufactured by bio - cide , international , p . o . box 2700 , norman , okla . 73070 . a 10 % solution of monobasic sodium phosphate was prepared in distilled water . ten ml was placed into each of four beakers . one of each of the four beakers received 1 , 2 . 5 , 5 , and 10 ml of chlorine dioxide concentrate ( 2 % clo 2 ), respectively . all solutions were diluted to 90 ml with distilled water , adjusted to ph 6 . 8 with 1n naoh and 1n hcl , diluted to 100 ml and placed in screw cap bottles . solutions containing dibasic and tribasic sodium phosphate and a distilled water blank control were prepared in a similar manner . chlorine dioxide content and ph was determined for each solution on days 0 , 7 , 14 , 21 and 28 in accordance with standard methods for the examination of water and wastewater , 17th edition , 1989 . as shown in table 1 , the content of chlorine dioxide was stable in all sodium phosphate solutions and distilled water control over the 28 day test period . the ph of all samples ranged from 6 . 1 to 7 . 6 . the stability of chlorine dioxide in various toothpastes at ph 5 . 0 . 5 . 5 , and 6 . 0 . stabilized chlorine dioxide in alkaline solutions is present as sodium chlorite . acidification of sodium chlorite results in the liberation of chlorine dioxide gas which is very reactive against organic material . 1 . tartar control crest toothpaste , original flavor , lot # 9342tk , manufactured by procter and gamble , cincinnati , ohio , 45202 . 2 . aquafresh for kids fluoride toothpaste bubblemint flavor , lot m292b , manufactured by beecham products , usa , pittsburgh , pa ., 15230 . 3 . generic toothpaste without a detergent ( unlabeled ), supplied by perry ratcliff , dds , 7125 east lincoln drive , scottsdale , ariz . 85253 . 4 . trisodium phosphate , na 3 po 4 . 12h 2 o . fisher scientific , fair lawn , n . j . table 1__________________________________________________________________________results showing the stability of chlorine dioxide solution at ph 6 . 8 indistilled water and 1 % sodium phosphate , monobasic , dibasic , andtribasic day theroetical 0 7 14 21 28solution % clo . sub . 2 ph % clo . sub . 2 ph % clo . sub . 2 ph % clo . sub . 2 ph % clo . sub . 2 ph % clo . sub . 2__________________________________________________________________________distilled water 0 . 02 6 . 8 0 . 02 6 . 9 0 . 02 6 . 9 0 . 02 6 . 5 0 . 02 6 . 5 0 . 02 0 . 05 6 . 8 0 . 05 6 . 9 0 . 05 6 . 9 0 . 05 7 . 1 0 . 05 6 . 9 0 . 05 0 . 1 6 . 8 0 . 1 6 . 9 0 . 1 7 . 0 0 . 1 7 . 7 0 . 1 7 . 6 0 . 1 0 . 2 6 . 8 0 . 2 6 . 9 0 . 2 6 . 9 0 . 2 7 . 2 0 . 2 7 . 2 0 . 21 % na . sub . 2 hpo . sub . 4 0 . 02 6 . 8 0 . 02 6 . 1 0 . 02 6 . 7 0 . 02 6 . 7 0 . 02 6 . 8 0 . 02 ( disodium 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05hydrogen 0 . 1 6 . 8 0 . 1 6 . 9 0 . 1 6 . 9 0 . 1 6 . 8 0 . 1 6 . 8 0 . 1phosphate ) 0 . 2 6 . 8 0 . 2 6 . 9 0 . 2 6 . 9 0 . 2 6 . 9 0 . 2 6 . 8 0 . 21 % nah . sub . 2 po . sub . 4 0 . 02 6 . 8 0 . 02 6 . 7 0 . 02 6 . 8 0 . 02 6 . 7 0 . 02 6 . 8 0 . 02 ( sodium 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05 6 . 8 0 . 05 6 . 9 0 . 05dihydrogen 0 . 1 6 . 8 0 . 1 6 . 8 0 . 1 6 . 8 0 . 1 6 . 9 0 . 1 6 . 9 0 . 1phosphate ) 0 . 2 6 . 8 0 . 2 6 . 8 0 . 2 6 . 8 0 . 2 6 . 9 0 . 2 6 . 9 0 . 21 % na . sub . 3 po . sub . 4 0 . 02 6 . 8 0 . 02 6 . 8 0 . 02 6 . 4 0 . 02 6 . 9 0 . 02 7 . 0 0 . 02 ( trisodium 0 . 05 6 . 8 0 . 05 7 . 0 0 . 05 7 . 1 0 . 05 6 . 9 0 . 05 7 . 0 0 . 05phosphate ) 0 . 1 6 . 8 0 . 1 7 . 5 0 . 1 7 . 5 0 . 1 7 . 0 0 . 1 6 . 9 0 . 1 0 . 2 6 . 8 0 . 2 7 . 0 0 . 2 7 . 1 0 . 2 6 . 9 0 . 2 6 . 9 0 . 2__________________________________________________________________________ 5 . purogene , ( 2 % chlorine dioxide ), lot # 8907 : 41 , manufactured by bio - cide international , inc ., p . o . box 2700 , normal , okla . 73070 . each toothpaste was divided into three 50 g portions and placed in plastic jars with lids . one portion of each toothpaste was adjusted to ph 5 . 0 , 5 . 5 , and 6 . 0 , respectively , with 1n naoh and 1n hcl . subsequently 5 ml 2 % chlorine dioxide was added to each portion and mixed thoroughly , yielding a chlorine dioxide concentration of approximately 0 . 15 %. toothpaste was dispensed into four jars , 50 g per jar . jars were divided into groups of two each . jars in the first group each received 1 . 2 g na 3 po 4 . 12h 2 o ( 1 % na 3 po 4 ) while jars in second group each received 2 . 3 g na 3 po 4 . 12h 2 o ( 2 % na 3 po 4 ). one jar from each group was adjusted to ph 5 . 0 and 6 . 0 , respectively . subsequently 5 ml 2 % chlorine dioxide was added to each portion and mixed thoroughly , yielding a chlorine dioxide concentration of approximately 0 . 15 %. the ph and chlorine dioxide content of all jars was determined . jars were stored in the dark at @ 20 degrees c . and reassayed for ph and chlorine dioxide content on days 7 , 14 , 21 and 28 . as shown in table 2 , the ph values of crest and generic toothpaste with 1 % and 2 % trisodium phosphate were stable during the twenty - eight day test period . in sharp contrast , the ph values of aquafresh toothpaste increased from 0 . 4 to 1 . 0 units during the 28 day test period . table 3 shows the percentage of chlorine dioxide remaining in the various toothpastes on days 0 , 7 , 14 , 21 , and 28 . chlorine dioxide in crest toothpaste gradually decreased during the 28 day test period . on day 28 , 6 % of the original chlorine dioxide was remaining in crest toothpaste at ph 5 . 0 . in sharp contrast , 54 % of the chlorine dioxide was present in crest toothpaste at ph 6 . 0 , nine fold greater than at ph 5 . 0 chlorine dioxide content in aqua fresh toothpaste remained stable for the first 21 days ( 78 - 100 % of clo 2 remaining ) at all ph levels . a sharp drop in chlorine dioxide concentration occurred on day 28 ; the reason for the decrease is not clear . chlorine dioxide content in generic toothpastes containing 1 % and 2 % trisodium phosphate at ph 5 . 0 gradually decreased during the 28 day test period . on the 28th day , 5 to 27 % of the chlorine dioxide remained , respectively . in sharp contrast , generic toothpaste containing 1 % and 2 % trisodium phosphate at ph 6 . 0 remained relatively stable during the 28 day test period ( 80 to 100 % chlorine dioxide remaining ). this is three to 20 fold greater retention of chlorine dioxide than at ph 5 . 0 . of all the toothpastes tested , generic toothpaste with either 1 % or 2 % trisodium phosphate at ph 6 . 0 appears to be the most compatible with chlorine dioxide . table 2______________________________________results showing ph values of toothpastesduring the twenty - eight day test period daytoothpaste 0 7 14 21 28______________________________________crest 5 . 0 5 . 0 5 . 0 5 . 0 5 . 0 5 . 5 5 . 5 5 . 5 5 . 5 5 . 5 6 . 0 6 . 0 6 . 0 6 . 0 6 . 0aqua fresh 5 . 0 5 . 6 5 . 7 5 . 9 6 . 0 5 . 5 5 . 7 5 . 7 5 . 8 5 . 9 6 . 0 6 . 2 6 . 3 6 . 4 6 . 6generic w / 1 % 5 . 0 5 . 0 5 . 0 5 . 0 5 . 1na . sub . 3 po . sub . 4 6 . 0 6 . 0 6 . 0 6 . 0 6 . 1generic w / 2 % 5 . 0 5 . 1 5 . 0 5 . 1 5 . 2na . sub . 3 po . sub . 4 6 . 0 6 . 0 6 . 0 6 . 0 6 . 1______________________________________ table 3______________________________________results showing the stabilityof chlorine dioxide in toothpastesat various ph levels duringtwenty - eight day test period percent chlorine dioxide remaining on daystoothpaste ph * 0 7 14 21 28______________________________________crest 5 . 0 100 56 38 19 6 5 . 5 100 60 47 40 27 6 . 0 100 85 77 62 54aqua fresh 5 . 0 100 86 86 93 43 5 . 5 100 89 89 78 22 6 . 0 100 100 100 100 17generic w / 1 % 5 . 0 100 55 25 10 5na . sub . 3 po . sub . 4 6 . 0 100 100 100 93 80generic w / 2 % 5 . 0 100 73 87 33 27na . sub . 3 po . sub . 4 6 . 0 100 82 82 88 82______________________________________ * ph day 0 . 1 . purogene ( 2 % chlorine dioxide ), lot # 8907 : 41 , manufactured by bio - cide international , inc ., p . o . box 2700 , norman , okla . 73070 . 2 . test organisms : streptococcus mutans ( atcc # 27152 ), streptococcus sanguis ( atcc # 10556 ), and candida albicans ( atcc # 18804 ) 10 . mcfarland nephelometer tube no . 1 . density of this tube is equivalent to a bacterial suspension of 3 × 10 8 organisms per ml . 13 . sodium dihydrogen phosphate , nah 2 po 4 . 7h 2 o . ( fisher scientific , fair lawn , n . j . 14 . trisodium phosphate , na 3 po 4 . 12h 2 o . albright & amp ; wilson , p . o . box 80 , oldbury , narley , west midlands , b694ln , england . 15 . sodium monofluorophosphate , na 2 fpo 3 , ref no . ob 12837 , manufactured by albright and wilson , p . o . box 80 , oldbury , narley , west midlands , b694ln , england . dpd reagent and phosphate buffer reagent were prepared in accord with standard methods for the examination of water and wastewater , 17th edition , p . 9 - 54 ( 1989 ). a ten percent sodium dihydrogen phosphate solution was prepared in distilled water . ten ml was placed into each of five beakers . one of each of the five beakers received 0 , 1 , 2 . 5 , 5 , and 10 ml of chlorine dioxide concentrate ( 2 % clo 2 ), respectively . all solutions were diluted to 90 ml with distilled water , adjusted to ph 6 . 0 with 1n naoh and 1n hcl , diluted to 100 ml and placed in screw cap bottles . solutions containing 0 ppm chlorine dioxide were filter sterilized prior to use . solutions containing trisodium phosphate and sodium monofluorophosphate were prepared in a similar manner . suspensions of each organism were prepared in butterfield &# 39 ; s buffer from 48 hour agar cultures and turbidity adjusted to a mcfarland tube # 1 . subsequently 0 . 1 ml of this suspension was diluted in 50 ml of saline . the diluted microorganism suspensions were now ready for use . one ml of test suspension was aliquoted into each of five sterile 16 × 125 mm screw cap tubes . each of the five tubes received 4 ml of a solution containing either 0 , 200 , 500 , 1000 , or 2000 ppm chlorine dioxide in 1 % sodium dihydrogen phosphate . each tube was shaken for ten seconds and immediately inactivated with 0 . 25 ml 15 % sodium thiosulfate . solutions containing 1 % trisodium phosphate and 1 % sodium monofluorophosphate were handled in a similar manner . one ml of test suspension was dispensed into two sterile 16 × 125 mm screw cap tubes . each tube received 4 ml 2000 ppm chlorine dioxide in 1 % sodium dihydrogen phosphate . the first tube received 0 . 25 ml sodium thiosulfate , while the second tube received none . subsequently each tube was tested for residual chlorine dioxide by adding 0 . 3 ml phosphate buffer reagent and 0 . 3 ml dpd reagent to each tube . neutralized tubes were colorless , while nonneutralized tubes were pink . solutions of trisodium phosphate and sodium monofluorophosphate containing 2 , 000 ppm chlorine dioxide were handled in a similar manner . one ml test suspension of each organism was treated with 4 ml butterfield &# 39 ; s buffer and 0 . 25 ml 10 % sodium thiosulfate as a negative control . sterility tests on all reagents were run parallel to experiments by plate counted method . the plate counted method and sterility tests were conducted in accord with standard methods for the examination of water and wastewater , 17th edition , p . 9 - 54 ( 1989 ). as shown in tables 4 , 5 and 6 , 99 - 100 % of the organisms were killed when challenged with 2 , 000 ppm ( 0 . 2 %) chlorine dioxide in either 1 % sodium dihydrogen phosphate or trisodium phosphate . in sharp contrast , only fifty percent of s . mutans were killed when challenged with 2 , 000 ppm chlorine dioxide in sodium monofluorophosphate . s . sanguis and c . albicans were 99 - 100 % killed when challenged with 1 , 000 ppm ( 0 . 1 %) chlorine dioxide in either 1 % sodium dihydrogen phosphate or trisodium phosphate . in sharp contrast , s . mutans was killed 24 and 76 %, respectively , when similarly challenged . 1 , 000 ppm chlorine dioxide in 1 % sodium monofluorophosphate yielded a 12 % kill when used against s . mutans . chlorine dioxide concentrations of 200 ( 0 . 02 %) and 500 ppm ( 0 . 05 %) in the presence of phosphates demonstrated marginal bacteriocidal activity against s . mutans , s . sanguis , and c . albicans ( 6 - 66 % kill ). table 4______________________________________results showing the bacteriocidalactivity of chlorine dioxide inphosphate solutions at ph 6 . 0 againststreptococcus mutans . phosphate solutionclo . sub . 2 negative ( ppm ) control * nah . sub . 2 hpo . sub . 4 na . sub . 3 po . sub . 4 na . sub . 2 fpo . sub . 3______________________________________ 0 17 , 000 ** 16 , 000 ( 6 )*** 18 , 000 ( 0 ) 14 , 000 ( 18 ) 200 nd 15 , 000 ( 12 ) 16 , 000 ( 6 ) 14 , 000 ( 18 ) 500 nd 15 , 000 ( 12 ) 15 , 000 ( 12 ) 14 , 000 ( 18 ) 1000 nd 13 , 000 ( 24 ) 4 , 100 ( 76 ) 15 , 000 ( 12 ) 2000 nd 2 ( 99 ) 120 ( 99 ) 8 , 500 ( 50 ) ______________________________________ * butterfield &# 39 ; s buffer ** organisms / ml *** percent kill nd = not done table 5______________________________________results showing the bacteriocidalactivity of chlorine dioxide inphosphate solutions at ph 6 . 0 againststreptococcus sanguisphosphate solutionclo . sub . 2 negative ( ppm ) control * 1 % nah . sub . 2 hpo . sub . 4 1 % na . sub . 3 po . sub . 4______________________________________ 0 29 , 000 ** 22 , 000 ( 24 )*** 25 , 000 ( 14 ) 200 nd 22 , 000 ( 24 ) 19 , 000 ( 34 ) 500 nd 16 , 000 ( 45 ) 11 , 000 ( 62 ) 1000 nd 360 ( 99 ) 0 ( 100 ) 2000 nd 0 ( 100 ) 0 ( 100 ) ______________________________________ * butterfield &# 39 ; s buffer ** organisms / ml *** percent kill nd = not done table 6______________________________________results showing the bacteriocidalactivity of chlorine dioxide inphosphate solutions at ph 6 . 0 againstcandida albicansphosphate solutionclo . sub . 2 negative ( ppm ) control * 1 % nah . sub . 2 hpo . sub . 4 1 % na . sub . 3 po . sub . 4______________________________________ 0 95 , 000 ** 64 , 000 ( 33 )*** 55 , 000 ( 42 ) 200 nd 58 , 000 ( 39 ) 64 , 000 ( 33 ) 500 nd 47 , 000 ( 51 ) 32 , 000 ( 66 ) 1000 nd 250 ( 99 ) 0 ( 100 ) 2000 nd 17 ( 99 ) 5 ( 99 ) ______________________________________ * butterfield &# 39 ; s buffer ** organisms / ml *** percent kill nd = not done the stability of chlorine dioxide at ph 6 . 8 in the presence of 0 . 02 % phosphate . the following is an example of how to test the stability of chlorine dioxide at ph 6 . 8 in the presence of 0 . 02 % phosphate . 1 . purogene ( 2 % clo 2 ), lot # 8907 . 41 , 1 gallon , manufactured by bio - cide , international , p . o . box 2700 , norman , okla . 73070 . a 0 . 2 % solution of monobasic sodium phosphate is prepared in distilled water . ten ml is placed into each of four beakers . one of each of the four beakers receives 1 , 2 . 5 , 5 , and 10 ml of chlorine dioxide concentrate ( 2 % clo 2 ), respectively . all solutions are diluted to 90 ml with distilled water , adjusted to ph 6 . 8 with 1n naoh and 1n hcl , diluted to 100 ml and placed in screw cap bottles . solutions containing dibasic and tribasic sodium phosphate and a distilled water blank control are prepared in a similar manner . chlorine dioxide content and ph is determined for each solution on days 0 , 7 , 14 , 21 and 28 in accordance with standard methods for the examination of water and wastewater , 17th edition , 1989 , in order to determine the stability of chlorine dioxide over time . the stability of chlorine dioxide in various toothpastes having a 0 . 02 % and 0 . 05 % na 3 po 4 at ph 5 . 0 . 5 . 5 , and 6 . 0 . the following is an example of how to test the stability of chlorine dioxide in various toothpastes having either 0 . 02 % or 0 . 05 % na 3 po 4 . stabilized chlorine dioxide in alkaline solutions is present as sodium chlorite . acidification of sodium chlorite results in the liberation of chlorine dioxide gas which is very reactive against organic material . 1 . tartar control crest toothpaste , original flavor , lot # 9342tk , manufactured by procter and gamble , cincinnati , ohio , 45202 . 2 . aquafresh for kids fluoride toothpaste bubblemint flavor , lot m292b , manufactured by beecham products , usa , pittsburgh , pa ., 15230 . 3 . generic toothpaste without a detergent ( unlabeled ), supplied by perry ratcliff , dds , 7125 east lincoln drive , scottsdale , ariz . 85253 . 4 . trisodium phosphate , na 3 po 4 . 12h 2 o . fisher scientific , fair lawn , n . j . 5 . purogene , ( 2 % chlorine dioxide ), lot # 8907 : 41 , manufactured by bio - cide international , inc ., p . o . box 2700 , normal , okla . 73070 . each toothpaste is divided into three 50 g portions and placed in plastic jars with lids . one portion of each toothpaste is adjusted to ph 5 . 0 , 5 . 5 , and 6 . 0 , respectively , with 1n naoh and 1n hcl . subsequently 5 ml 2 % chlorine dioxide is added to each portion and mixed thoroughly to yield a chlorine dioxide concentration of approximately 0 . 15 %. toothpaste is dispensed into four jars , 50 g per jar . the jars are divided into groups of two each . jars in the first group each receive 0 . 24 na 3 po 4 . 12h 2 o ( 0 . 02 na 3 po 4 ) while jars in second group each receive 0 . 6 g na 3 po 4 . 12h 2 o ( 0 . 05 % na 3 po 4 ). one jar from each group is adjusted to ph 5 . 0 and 6 . 0 , respectively . subsequently a sufficient amount of 2 % chlorine dioxide is added to each portion and mixed thoroughly to yield a chlorine dioxide concentration of approximately 0 . 15 %. the ph and chlorine dioxide content of all jars is determined . the jars are stored in the dark at @ 20 degrees c . and reassayed for ph and chlorine dioxide content on days 7 , 14 , 21 and 28 , in order to determine the stability of the various toothpastes . the following is an example of how to test the antimicrobial effectiveness of chlorine dioxide in 0 . 02 % phosphate solution . 1 . purogene ( 2 % chlorine dioxide ), lot # 8907 : 41 , manufactured by bio - cide international , inc ., p . o . box 2700 , norman , okla . 73070 . 2 . test organisms : streptococcus mutans ( atcc # 27152 ), streptococcus sanguis ( atcc # 10556 ), and candida albicans ( atcc # 18804 ) 10 . mcfarland nephelometer tube no . 1 . density of this tube is equivalent to a bacterial suspension of 3 × 10 8 organisms per ml . 13 . sodium dihydrogen phosphate , nah 2 po 4 . 7h 2 o . ( fisher scientific , fair lawn , n . j . 14 . trisodium phosphate , na 3 po 4 . 12h 2 o . albright & amp ; wilson , p . o . box 80 , oldbury , narley , west midlands , b694ln , england . 15 . sodium monofluorophosphate , na 2 fpo 3 , ref no . ob 12837 , manufactured by albright and wilson , p . o . box 80 , oldbury , narley , west midlands , b694ln , england . dpd reagent and phosphate buffer reagent is prepared in accord with standard methods for the examination of water and wastewater , 17th edition , p . 9 - 54 ( 1989 ). a 0 . 2 percent sodium dihydrogen phosphate solution is prepared in distilled water . ten ml is placed into each of five beakers . one of each of the five beakers receives 0 , 1 , 2 . 5 , 5 , and 10 ml of chlorine dioxide concentrate ( 2 % clo 2 ), respectively . all solutions are diluted to 90 ml with distilled water , adjusted to ph 6 . 0 with 1n naoh and 1n hcl , diluted to 100 ml and placed in screw cap bottles . solutions containing 0 ppm chlorine dioxide are filter sterilized prior to use . solutions containing trisodium phosphate and sodium monofluorophosphate are prepared in a similar manner . suspensions of each organism are prepared in butterfield &# 39 ; s buffer from 48 hour agar cultures and turbidity adjusted to a mcfarland tube # 1 . subsequently 0 . 1 ml of this suspension is diluted in 50 ml of saline . the diluted microorganism suspensions are then ready for use . one ml of test suspension is aliquoted into each of five sterile 16 × 125 mm screw cap tubes . each of the five tubes receives 4 ml of a solution containing either 0 , 200 , 500 , 1000 , or 2000 ppm chlorine dioxide in 1 % sodium dihydrogen phosphate . each tube is shaken for ten seconds and immediately inactivated with 0 . 25 ml 15 % sodium thiosulfate . solutions containing 1 % trisodium phosphate and 1 % sodium monofluorophosphate are handled in a similar manner . one ml of test suspension is dispensed into two sterile 16 × 125 mm screw cap tubes . each tube receives 4 ml 2000 ppm chlorine dioxide in 0 . 02 % sodium dihydrogen phosphate . the first tube receives 0 . 25 ml sodium thiosulfate , while the second tube receives none . subsequently each tube is tested for residual chlorine dioxide by adding 0 . 3 ml phosphate buffer reagent and 0 . 3 ml dpd reagent to each tube . neutralized tubes are colorless , while nonneutralized tubes are pink . solutions of trisodium phosphate and sodium monofluorophosphate containing 2 , 000 ppm chlorine dioxide are handled in a similar manner . one ml test suspension of each organism is treated with 4 ml butterfield &# 39 ; s buffer and 0 . 25 ml 10 % sodium thiosulfate as a negative control . sterility tests on all reagents are run parallel to experiments by plate counted method . the plate counted method and sterility tests are conducted in accord with standard methods for the examination of water and wastewater , 17th edition , p . 9 - 54 ( 1989 ), in order to determine the antimicrobial effectiveness of chlorine dioxide in 0 . 02 % phosphate solution . it will be seen from the foregoing that the mixture of stabilized chlorine dioxide and phosphates as part of a mouth wash or toothpaste has an improved shelf life , and is effective as a bactericide superior to other compositions used today . both stabilized chlorine dioxide , and phosphates , have been used for many years in other areas and extensive study in animals and in man have demonstrated the compounds , low toxicity and safety . it will be obvious that various changes , alterations and modifications to the method and process described herein may be made , to the extent that such changes , alterations and modifications do not depart from the spirit and scope of the appended claims therein intended to be encompassed herein .

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