Scand. ]. Dent. Res. 1975: 83: 18-25 (Key words: dental plaque; gingivitis)
Effect of Listerine® on dental plaque and gingivitis JAN FORNELL, YVONNE SUNDIN AND JAN LINDHE Department of Periodontology, Faculty of Odontology, University of Gothenburg, Gothenburg, Sweden /VBSTRACT — The present study was performed in 10 adults in order to evaluate the effect of an antiseptic mouthrinse (Listeririe®) on the rate of dental plaque formation and gingivitis development during a 2-week period when all efforts towards active mechanical oral hygiene were withdrawn. The study was performed as a crossover study and was carried out during four consecutive 2-week periods. During the first and third periods (preparatory periods) the participants were subjected to repeated professional tooth cleanings in order to establish plaque- and gingiviti.s-free dentitions. During the second and fourth periods (test and control periods) the participants were not allowed to brush their teeth but rinsed their mouths three times a day with Listerine or a placebo mouthwash. Plaque Index, Gingival Index, gingival fluid flow, and crevicular leukocytes were assessed on d 0, 2, 4, 7, and 14. On d 7 and 14, dental plaque was removed from the right and left jaws respectively and the wet weights determined. The chemotactic activity elaborated by the plaques was studied in Boyden chambers. During the Listerine test period, significantly lower Plaque and Gingival Index values were scored and lower amounts of plaque could be sampled in comparison to the control period. (Received for publication 11 June, accepted 16 October 1974)
Observations made in clinical trials and animal experiments have demonstrated that the microbial plaque plays a central role in the etiology of gingivitis, periodontitis, and caries (SOCRANSKY 1970, LOE, FEHR & ScHioTT 1970, HAMP 1973). Attempts have therefore been made to develop techniques for facilitating and improving the effect of toothbrushing (for review see BAY 1972, BERGENHOLTZ 1972). The maintenance of adequate standards of oral hygiene for prolonged periods by mechanical tooth cleaning, however, is laborious (LINDHE & KOCH
utilize drugs incorporated in mouthwashes and dentifrices, foi" example, to minimize the rate of plaque formation. The aim of the present investigation in young adults was to appraise the effect of an antiseptic mouthrinse (Listerine®, Warner-Lambert Pharmaceutical Company, U.S.A.) on the rate of (1) dental plaque formation and (2) development of gingivitis during 2 weeks' abstinence from active mechanical cleaning of the teeth.
1967, KOGH & LINDHE 1970). Consequently, efforts have also been made to
Ten adults, aged 19 to 30, participated in the clinical trial. At an initial examination the oral
Material and methods
EFFEGT OF LISTERINE® ON PLAQUE
19
Table 1 Results of clinical examination on day 0 of the two periods of abstinence from oral hygiene. No significant difference between the two groups regarding Pll, GI, amount of gingival fluid and crevicular leukocytes, x = mean; s.e. = standard error of the mean Gingival fluid X s.e.
Grevicular leukocytes X s.e.
0.08 ± 0.01 0.11 ± 0.02
0.27 ± 0.06 0.16 ± 0.04
153 ± 33 145 ± 27
N.S.
N.S.
N.S.
Day
Pll
GI
0 (L) 0 (P)
0.09 ± 0.02 0.18 ± 0.03 N.S.
hygiene status of the participants was assessed according to the criteria of the Plaque Index system (Pll; SILNESS & LOE 1964). The gingival conditions were analyzed according to the Gingival Index system (GI; LOE & SILNESS 1963, and measurements of gingival fluid and crevicular leukocytes (LOE & HOLM-PEDERSEN 1965, ATTSTROM & EGELBERG 1971). This initial examination was followed by an initial preparatory 2-week period during which the teeth were first scaled and polished until staining with basic fuchsin showed no plaque. Each individual was also instructed in adequate oral hygiene habits, including use of the Bass method (B.\ss 1948) of toothbrushing and interdental cleaning with floss or toothpicks. Once every third day during this preparatory period the participants were subjected to professional dental cleaning as described by LINDHE & AXELSSON (1973). At the end of this first preparatory period, i.e. on day 0 of the proper trial, the Plaque and Gingival Indices were assessed anew and gingival exudate and crevicular leukocytes were sampled. The clinical measurements were made in the following tooth areas: 16, 15, 14, 11, 21, 24, 25, 26, 36, 35, 34, 31, 46, 4.5, 44, 41.* Gingival and Plaque Indices were scored on all tooth surfaces, but gingival exudate and crevicular leukocytes were sampled only on the buccal part of the gingiva. The individual mean values were calculated and are given in Table 1. The participants were then told to abstain
* The "two-digit" system. In: World Health Organization: Oral health surveys. Basic methods. WHO, Geneva, p. 18.
from all mechanical tooth cleaning procedures during a 2-week period. They were instructed to rinse their mouths three times a day (in the morning, at noon, and in the evening) with 20 ml of either a Listerine mouthwash (test persons 1—5) or a placebo solution (test persons 6-10). The participants were given individual instruction in the use of the mouthwash, i.e. to rinse the mouth vigorously for 1 min each time. Listerine® antiseptic mouthrinse is a. 2b % aqueous alcoholic solution of the following essential oils: eucalyptol, thymol, mentliol and methyl salicylate. It is acidified to pH 4.2 with benzoic and boric acid. Measurements of the Plaque and Gingival Index and sampling of the gingival exudate and crevicular leukocytes were repeated after 2, 4, 7, and 14 d of the experiment. The following teeth and gingival areas were examined at the various intervals after day 0: day 2: 16, 15, 14, 11; day 4: 36, 35, 34, 31; day 7: 46, 45, 44, 41; day 14: 26, 25, 24, 21. The mean Pll, GI, gingival exudate and crevicular leukocyte scores were determined and the differences between these and those noted on day 0 were calculated. On day 7 the plaque formed on all surfaces of the right maxillary and mandibular teeth was sampled. The teeth were first dried by a blast of air and kept dry by cotton rolls placed in the vestibule. The supragingival plaque was then sampled from the tooth surfaces with the use of a sterile carver. The wet weight of each plaque sample was determined. On day 14 plaque was sampled in the same way from the left maxillary and mandibular teeth. After the end of the first main test period the participants were allowed to clean their teeth with a toothbrush, toothpicks and dentifrice. They were also subjected to professional
FORNELL, SUNDIN AND LINDHE
20
Tabble 2 Plaque formation and gi7igivitis development during the two test periods, x = mean; s.e. = stajidard error of the mean Day
Plaque Index Listerine Placebo Gingival Index Listerine Placebo Gingival fluid Listerine Placebo Crevicular leukocytes Listerine Placebo
2 X
4 s.e.
X
7 s.e.
X
14 s.e.
X
s.e.
0.27 ±0.05 0.8210.13
0.53±0.11 1.36±0.12
0.82±0.12 1.66±0.12
0.93±0.13 1.87 ±0.07
0.08 ±0.02 0.13±0.02
O.12±O.O3 0.33±0.08
0.25±0.07 0.50±0.06
0.71±0.10 1.29±0.06
0.04 ±0.03 0.12 ±0.05
0.18±0.07 0.09 ±0.07
0.34±0.18 0.41 ±0.15
0.22±0.07 0.34±0.08
90 ±23 112±20
n2±19 68±12
72±19 95 ±16
158±32 1G7±33
tooth cleaning in the same way as during the initial preparatory period. After 2 weeks of active oral hygiene, a second test period was started during which the participants rinsed their mouths with Listerine mouthwash (test persons 6-10) or with the placebo solution (test persons 1-5). The oral hygiene and the conditions of the gingivae were studied in the same way as during the first test period. Plaque was sampled after 7 and 14 d of abstaining from oral hygiene. The chemotactic activity for rabbit neutrophil leukocytes of the plaque formed during the two test periods was determined using an in vitro system described by BOYDEN (1962) and in accordance with a technique presented by LiNDHE & HELLDEN (1972). The wet weight of the individual plaque samples (from day 7 or from day 14) was determined, after which the "Listerine" and "placebo" plaques were pooled separately. Each pool was suspended in 1 ml 0.15 M NaCl, homogenized in a Whirlimixer® for at least 2 min, centrifuged at 12,000 g for 30 min at -|-4°C. The supernatant was filtered through a 0.45 i-im Millipore® filter (Millipore Filter Corp., New Bedford, Mass., U.S.A.). The protein content of the filtrate was determined according to LOWRY, ROSEBROUGH, FARR & RANDALL (1951) with tyrosine as a standard. The protein content of the plaque filtrates was then adjusted by dilution in saline in such a way that the four plaque pools contained equal amounts of protein.
Ten tests were performed with each of the four plaque filtrate pools. In addition, five tests were carried out with Gey's medium, and five tests with casein placed in the lower compartment of the Boyden chamber (for details see TYNELIUS-BRATTHALL & LINDHE 1974).
The significance of differences between pair-' ed observations was tested by the Student's t test (CHILTON 1967). Alterations in gingival fluid flow and number of crevicular leukocytes during the no oral hygiene periods were studied by analysis of variance.
Results At the beginning of the two test periods, i.e. on day 0 of the Listerine period and on day 0 of the placebo period, there was no significant difference in Plaque Index, Gingival Index, gingival fluid or crevicuiar leukocytes between the two groups (Table 1). On abstinence from active oral hygiene during the placebo period large amounts of plaque developed. The Plaque Index scores increased during the first 2 d from, on the average, 0.18 to 0.82 (increase of 0.64) and then to 1.36 (day 4), 1.66 (day 7) and 1.87 (day 14). The development
EFFECT OF LISTERINE® ON PLAQUE
21
Table 3 Pll and GI scores from day 0 representing the buccal/oral and approximal tooth s_urfaces during the two periods of abstinence from active oral hygiene, x = mean; s.e. = standard error of the mean Day
Plaque Index Buccal/oral Listerine Placebo Approximal Listerine Placebo Gingival Index Buccal/oral Listerine Placebo Approximal Listerine Placebo
2 X
X
14
7
4 s.e.
s.e.
X
s.e.
X
s.e.
0.15±0.05 0.44±0.12
0.30 ±0.09 1.12±0.12
0.60±0.12 1.46±0.12
0.53 ±0.08 1.51 ±0.09
0.40 ±0.06 1.05±0.19
0.74±0.19 1.59±0.16
0.96±0.16 1.92±0.13
1.33±0.18 2.30±0.11
0.02 ±0.01 0.02±0.01
0.07 ±0.03 0.23 ±0.08
0.14±0.05 0.36 ±0.06
0.58±0.11 1.06±0.07
0.15±0.04 0.25 ±0.04
O.17±O.O5
0.37±0.10
0.44 ±0.15 0.63 ±0.08
0.84±0.11 1.47±0.11
of plaque during the Listerine period was substantially retarded (Table 2). At the end of the no oral hygiene period the Plaque Index score of the Listerine group was only 0.93 compared to 1.87 for the placebo. Table 3 gives the increase in the Pll scores from day 0 in the various tooth areas. The mean values show that the Listerine mouthrinse had its strongest effect on the buccal and the lingual tooth surfaces, where after a 14-d abstinence from toothbrushing the calculated increase of the Plaque Index score was 0.52. The corresponding increase for the placebo group was 1.48. Plaque development was also markedly retarded on the approximal surfaces. Thus after 14 d of no mechanical tooth cleaning in the placebo group the increase of the Plaque Index score for the approximal surfaces was 1.98, which should be compared with 1.16 for the Listerine period. It should also be observed that at each examination the Plaque Index scores during the placebo oeriod were significantly (P
Effect of Listerine® on dental plaque and gingivitis JAN FORNELL, YVONNE SUNDIN AND JAN LINDHE Department of Periodontology, Faculty of Odontology, University of Gothenburg, Gothenburg, Sweden /VBSTRACT — The present study was performed in 10 adults in order to evaluate the effect of an antiseptic mouthrinse (Listeririe®) on the rate of dental plaque formation and gingivitis development during a 2-week period when all efforts towards active mechanical oral hygiene were withdrawn. The study was performed as a crossover study and was carried out during four consecutive 2-week periods. During the first and third periods (preparatory periods) the participants were subjected to repeated professional tooth cleanings in order to establish plaque- and gingiviti.s-free dentitions. During the second and fourth periods (test and control periods) the participants were not allowed to brush their teeth but rinsed their mouths three times a day with Listerine or a placebo mouthwash. Plaque Index, Gingival Index, gingival fluid flow, and crevicular leukocytes were assessed on d 0, 2, 4, 7, and 14. On d 7 and 14, dental plaque was removed from the right and left jaws respectively and the wet weights determined. The chemotactic activity elaborated by the plaques was studied in Boyden chambers. During the Listerine test period, significantly lower Plaque and Gingival Index values were scored and lower amounts of plaque could be sampled in comparison to the control period. (Received for publication 11 June, accepted 16 October 1974)
Observations made in clinical trials and animal experiments have demonstrated that the microbial plaque plays a central role in the etiology of gingivitis, periodontitis, and caries (SOCRANSKY 1970, LOE, FEHR & ScHioTT 1970, HAMP 1973). Attempts have therefore been made to develop techniques for facilitating and improving the effect of toothbrushing (for review see BAY 1972, BERGENHOLTZ 1972). The maintenance of adequate standards of oral hygiene for prolonged periods by mechanical tooth cleaning, however, is laborious (LINDHE & KOCH
utilize drugs incorporated in mouthwashes and dentifrices, foi" example, to minimize the rate of plaque formation. The aim of the present investigation in young adults was to appraise the effect of an antiseptic mouthrinse (Listerine®, Warner-Lambert Pharmaceutical Company, U.S.A.) on the rate of (1) dental plaque formation and (2) development of gingivitis during 2 weeks' abstinence from active mechanical cleaning of the teeth.
1967, KOGH & LINDHE 1970). Consequently, efforts have also been made to
Ten adults, aged 19 to 30, participated in the clinical trial. At an initial examination the oral
Material and methods
EFFEGT OF LISTERINE® ON PLAQUE
19
Table 1 Results of clinical examination on day 0 of the two periods of abstinence from oral hygiene. No significant difference between the two groups regarding Pll, GI, amount of gingival fluid and crevicular leukocytes, x = mean; s.e. = standard error of the mean Gingival fluid X s.e.
Grevicular leukocytes X s.e.
0.08 ± 0.01 0.11 ± 0.02
0.27 ± 0.06 0.16 ± 0.04
153 ± 33 145 ± 27
N.S.
N.S.
N.S.
Day
Pll
GI
0 (L) 0 (P)
0.09 ± 0.02 0.18 ± 0.03 N.S.
hygiene status of the participants was assessed according to the criteria of the Plaque Index system (Pll; SILNESS & LOE 1964). The gingival conditions were analyzed according to the Gingival Index system (GI; LOE & SILNESS 1963, and measurements of gingival fluid and crevicular leukocytes (LOE & HOLM-PEDERSEN 1965, ATTSTROM & EGELBERG 1971). This initial examination was followed by an initial preparatory 2-week period during which the teeth were first scaled and polished until staining with basic fuchsin showed no plaque. Each individual was also instructed in adequate oral hygiene habits, including use of the Bass method (B.\ss 1948) of toothbrushing and interdental cleaning with floss or toothpicks. Once every third day during this preparatory period the participants were subjected to professional dental cleaning as described by LINDHE & AXELSSON (1973). At the end of this first preparatory period, i.e. on day 0 of the proper trial, the Plaque and Gingival Indices were assessed anew and gingival exudate and crevicular leukocytes were sampled. The clinical measurements were made in the following tooth areas: 16, 15, 14, 11, 21, 24, 25, 26, 36, 35, 34, 31, 46, 4.5, 44, 41.* Gingival and Plaque Indices were scored on all tooth surfaces, but gingival exudate and crevicular leukocytes were sampled only on the buccal part of the gingiva. The individual mean values were calculated and are given in Table 1. The participants were then told to abstain
* The "two-digit" system. In: World Health Organization: Oral health surveys. Basic methods. WHO, Geneva, p. 18.
from all mechanical tooth cleaning procedures during a 2-week period. They were instructed to rinse their mouths three times a day (in the morning, at noon, and in the evening) with 20 ml of either a Listerine mouthwash (test persons 1—5) or a placebo solution (test persons 6-10). The participants were given individual instruction in the use of the mouthwash, i.e. to rinse the mouth vigorously for 1 min each time. Listerine® antiseptic mouthrinse is a. 2b % aqueous alcoholic solution of the following essential oils: eucalyptol, thymol, mentliol and methyl salicylate. It is acidified to pH 4.2 with benzoic and boric acid. Measurements of the Plaque and Gingival Index and sampling of the gingival exudate and crevicular leukocytes were repeated after 2, 4, 7, and 14 d of the experiment. The following teeth and gingival areas were examined at the various intervals after day 0: day 2: 16, 15, 14, 11; day 4: 36, 35, 34, 31; day 7: 46, 45, 44, 41; day 14: 26, 25, 24, 21. The mean Pll, GI, gingival exudate and crevicular leukocyte scores were determined and the differences between these and those noted on day 0 were calculated. On day 7 the plaque formed on all surfaces of the right maxillary and mandibular teeth was sampled. The teeth were first dried by a blast of air and kept dry by cotton rolls placed in the vestibule. The supragingival plaque was then sampled from the tooth surfaces with the use of a sterile carver. The wet weight of each plaque sample was determined. On day 14 plaque was sampled in the same way from the left maxillary and mandibular teeth. After the end of the first main test period the participants were allowed to clean their teeth with a toothbrush, toothpicks and dentifrice. They were also subjected to professional
FORNELL, SUNDIN AND LINDHE
20
Tabble 2 Plaque formation and gi7igivitis development during the two test periods, x = mean; s.e. = stajidard error of the mean Day
Plaque Index Listerine Placebo Gingival Index Listerine Placebo Gingival fluid Listerine Placebo Crevicular leukocytes Listerine Placebo
2 X
4 s.e.
X
7 s.e.
X
14 s.e.
X
s.e.
0.27 ±0.05 0.8210.13
0.53±0.11 1.36±0.12
0.82±0.12 1.66±0.12
0.93±0.13 1.87 ±0.07
0.08 ±0.02 0.13±0.02
O.12±O.O3 0.33±0.08
0.25±0.07 0.50±0.06
0.71±0.10 1.29±0.06
0.04 ±0.03 0.12 ±0.05
0.18±0.07 0.09 ±0.07
0.34±0.18 0.41 ±0.15
0.22±0.07 0.34±0.08
90 ±23 112±20
n2±19 68±12
72±19 95 ±16
158±32 1G7±33
tooth cleaning in the same way as during the initial preparatory period. After 2 weeks of active oral hygiene, a second test period was started during which the participants rinsed their mouths with Listerine mouthwash (test persons 6-10) or with the placebo solution (test persons 1-5). The oral hygiene and the conditions of the gingivae were studied in the same way as during the first test period. Plaque was sampled after 7 and 14 d of abstaining from oral hygiene. The chemotactic activity for rabbit neutrophil leukocytes of the plaque formed during the two test periods was determined using an in vitro system described by BOYDEN (1962) and in accordance with a technique presented by LiNDHE & HELLDEN (1972). The wet weight of the individual plaque samples (from day 7 or from day 14) was determined, after which the "Listerine" and "placebo" plaques were pooled separately. Each pool was suspended in 1 ml 0.15 M NaCl, homogenized in a Whirlimixer® for at least 2 min, centrifuged at 12,000 g for 30 min at -|-4°C. The supernatant was filtered through a 0.45 i-im Millipore® filter (Millipore Filter Corp., New Bedford, Mass., U.S.A.). The protein content of the filtrate was determined according to LOWRY, ROSEBROUGH, FARR & RANDALL (1951) with tyrosine as a standard. The protein content of the plaque filtrates was then adjusted by dilution in saline in such a way that the four plaque pools contained equal amounts of protein.
Ten tests were performed with each of the four plaque filtrate pools. In addition, five tests were carried out with Gey's medium, and five tests with casein placed in the lower compartment of the Boyden chamber (for details see TYNELIUS-BRATTHALL & LINDHE 1974).
The significance of differences between pair-' ed observations was tested by the Student's t test (CHILTON 1967). Alterations in gingival fluid flow and number of crevicular leukocytes during the no oral hygiene periods were studied by analysis of variance.
Results At the beginning of the two test periods, i.e. on day 0 of the Listerine period and on day 0 of the placebo period, there was no significant difference in Plaque Index, Gingival Index, gingival fluid or crevicuiar leukocytes between the two groups (Table 1). On abstinence from active oral hygiene during the placebo period large amounts of plaque developed. The Plaque Index scores increased during the first 2 d from, on the average, 0.18 to 0.82 (increase of 0.64) and then to 1.36 (day 4), 1.66 (day 7) and 1.87 (day 14). The development
EFFECT OF LISTERINE® ON PLAQUE
21
Table 3 Pll and GI scores from day 0 representing the buccal/oral and approximal tooth s_urfaces during the two periods of abstinence from active oral hygiene, x = mean; s.e. = standard error of the mean Day
Plaque Index Buccal/oral Listerine Placebo Approximal Listerine Placebo Gingival Index Buccal/oral Listerine Placebo Approximal Listerine Placebo
2 X
X
14
7
4 s.e.
s.e.
X
s.e.
X
s.e.
0.15±0.05 0.44±0.12
0.30 ±0.09 1.12±0.12
0.60±0.12 1.46±0.12
0.53 ±0.08 1.51 ±0.09
0.40 ±0.06 1.05±0.19
0.74±0.19 1.59±0.16
0.96±0.16 1.92±0.13
1.33±0.18 2.30±0.11
0.02 ±0.01 0.02±0.01
0.07 ±0.03 0.23 ±0.08
0.14±0.05 0.36 ±0.06
0.58±0.11 1.06±0.07
0.15±0.04 0.25 ±0.04
O.17±O.O5
0.37±0.10
0.44 ±0.15 0.63 ±0.08
0.84±0.11 1.47±0.11
of plaque during the Listerine period was substantially retarded (Table 2). At the end of the no oral hygiene period the Plaque Index score of the Listerine group was only 0.93 compared to 1.87 for the placebo. Table 3 gives the increase in the Pll scores from day 0 in the various tooth areas. The mean values show that the Listerine mouthrinse had its strongest effect on the buccal and the lingual tooth surfaces, where after a 14-d abstinence from toothbrushing the calculated increase of the Plaque Index score was 0.52. The corresponding increase for the placebo group was 1.48. Plaque development was also markedly retarded on the approximal surfaces. Thus after 14 d of no mechanical tooth cleaning in the placebo group the increase of the Plaque Index score for the approximal surfaces was 1.98, which should be compared with 1.16 for the Listerine period. It should also be observed that at each examination the Plaque Index scores during the placebo oeriod were significantly (P
