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Objectives: To review the established and novel methods of plaque quantification employed in dental research, including a discussion of their merits and to present a new method of planimetrically measuring plaque using light induced fluorescence., Method: Quantitative light-fluorescence (QLF) images were acquired from the buccal surfaces of an individual who had refrained from oral hygiene both with and without traditional plaque disclosure. Digital photographs were also taken. Images were analysed using a novel method and a percentage plaque index produced., Results: Traditional plaque indices are problematic due to their integral nature and their failure to detect small, but potentially clinically relevant changes in plaque area. The use of a fluorescent technique demonstrated good reliability although there was no correlation between red fluorescent plaque and total disclosed plaque suggesting that the auto-fluorescing plaque is not a good measure of total plaque volume., Conclusions: The use of planimetric techniques can increase the power of plaque studies, potentially reducing the number of subjects and time required to separate therapies or products. Fluorescent methods of quantification have potential as they enable clear separation of the plaque covered and non-covered tooth surfaces.

Objectives: The objectives of this review were to give a comprehensive account of the methods used to determine dental plaque pH over the last 50 years, to review how these methods have been used in dental cariology research and to give an update as to how dental plaque pH studies might be developed in the future., Data: Published, referred papers and abstracts of conference proceedings in the literature were reviewed., Sources: A comprehensive search of the electronic databases PubMed and Medline, was undertaken. In addition, a hand search of the Index Denticus was done to identify relevant citations before 1966., Study Selection: Relevant published literature in peer-reviewed publications was reviewed. No additional inclusion criteria were applied., Conclusions: This comprehensive review gives an account of the background to, history of, relative merits and demerits of, applications of and future of dental plaque pH technologies.

Objective: To determine the effects of a detergent-free, whitening dentifrice using an in vivo plaque regrowth model with the novel application of QLF as a planimetric analysis tool., Method: A total of 20 subjects took part in a double blind, single-centre, crossover study in which slurry rinses were the only form of plaque control over a 5-day period. Following a washout and prophylaxis the subjects used 2 daily rinses in the absence of all other plaque control methods. Subjects returned to the clinic on the afternoon of day 5 when plaque was disclosed and assessed by the plaque index and area using both a photographic and novel fluorescent planimetric technique. A further 9-day washout was carried out and the rinse period repeated to ensure that each subject had used both experimental and comparator slurries., Results: Twenty subjects completed the trial. The test product showed a significant inhibition of plaque re-growth (16.9%) compared with a fluoride-matched comparator using the Turesky index (P < 0.0001), the photographic planimetric technique (17.5%) (P < 0.0001) and the novel QLF technique (18.4%) (P < 0.0001)., Conclusion: The results confirm that plaque inhibition capability of a detergent-free whitening dentifrice is at least as effective as a fluoride matched comparator. QLF is a promising tool for disclosed plaque quantification.

Objective: To determine the effects of a detergent-free dentifrice containing olive oil using in vitro microbiological tests and an in vivo plaque re-growth model., Method: A total of 20 subjects took part in a double blind, single-centre, crossover study in which slurry rinses were the only form of plaque control over a 5-day period. Following a washout and prophylaxis the subjects used 2x daily rinses in the absence of all other plaque control methods. Subjects returned to the clinic on the afternoon of day 5 when plaque was disclosed and assessed by plaque index and area. A further 9-day washout was carried out and the rinse period repeated to ensure that each subject had used both experimental and control slurries. In vitro microbiological experiments were conducted to examine the effects of olive oil alone and in the new dentifrice on bacterial growth and adhesion., Results: 20 subjects completed the trial. The olive oil product showed a significant inhibition of plaque re-growth (17.2%) compared with a fluoride-matched control using the Turesky index (p<0.0001) and also using the planimetric technique (22.0%) (p<0.0001). Significant decreases in bacterial growth and adhesion (measured as total viable count) were detected in the presence of olive oil and the new dentifrice., Conclusion: The results suggest that the experimental olive oil containing paste has potential value in the inhibition of plaque.

The aim of this study was to investigate the acid anions produced in plaque after chewing various cereal-based foods and fruits for one minute. Test foods were oranges, apples, bananas, Cornflakes, Branflakes, Weetabix, Alpen, white bread, wholemeal bread, rice and spaghetti, plus positive and negative controls of 10% sucrose and 10% sorbitol. 4 males and 3 females, aged 22-37 years, participated in the study .7 min from the start of chewing, 48-h plaque was collected from all accessible smooth surfaces, with no attempt to collect interproximal plaque, and centrifuged. Plaque fluid was withdrawn and analyzed by isotachophoresis for formate, succinate, lactate, acetate and propionate. At rest, acetate was the major anion present in plaque fluid, whereas following carbohydrate consumption, highest levels of lactate were detected followed by acetate. The amount of lactate only, detected in plaque fluid, was significantly correlated to the carbohydrate present in the food. It was concluded that important information regarding the acidogenicity of test foods is gained by studying the acid anion profile of plaque fluid.

The effect of sucrose rinses supplemented with L-LDH and NAD was investigated. When LDH or NAD were added to the sucrose rinse, the pH fall was less marked than with rinsing with sucrose alone. Significant differences (p < 0.05) in mean minimum pH and cH areas were observed when sucrose rinses were supplemented with LDH, NAD or with a combination of LDH and NAD, when compared with sucrose alone. Plaque fluid concentrations of lactate and acetate significantly decreased with all rinses when compared with sucrose alone. Phosphate levels also decreased, although not significantly, when NAD was the sole supplement. No significant decreases were observed in succinate, formate or propionate concentrations. The results confirm the beneficial effect of LDH in reducing the accumulation of lactate in plaque following a sucrose rinse. The inclusion of NAD in the rinse demonstrated the role of extracellular hydrogen acceptors in the reaction. Further work is required to identify endogenous hydrogen receptors and to clarify the fate of pyruvate resulting from the oxidation of lactate before the potential role of LDH as a cariostatic agent can be fully understood.

The protective role of saliva is demonstrated by the rampant caries seen in human subjects with marked salivary hypofunction, and in desalivated animals. In normal cases, however, the relationship between saliva flow and coronal or root caries experience is doubtful, and to examine the concept that stimulation of saliva might have protective effects against caries, one must look beyond a simple correlation between caries and flow rate. Protective properties of saliva which increase on stimulation include salivary clearance, buffering power, and degree of saturation with respect to tooth mineral. These benefits are maximized when saliva is stimulated after the consumption of fermentable carbohydrates, by reducing the fall in plaque pH leading to demineralization and by increasing the potential for remineralization. Plaque acid production is neutralized, and experimental lesions in enamel are remineralized, when gum is chewed to stimulate saliva after a carbohydrate intake. The pH-raising effects are more easily explained by the buffering action of the stimulated saliva than by clearance of carbohydrates. The remineralization action depends upon the presence of fluoride. These findings suggest that the protective actions of saliva can be mobilized by appropriate salivary stimulation, and that in addition to established procedures such as tooth cleaning and fluoride regimens, eating patterns which lead to saliva stimulation to increase the potential for saliva protection might be included in recommendations for caries prevention. Confirmation of this concept in clinical tests is required.

Previous studies have suggested that if either sugar-free or sucrose-containing chewing gum is chewed after acidogenic meals or snacks, the plaque pH response to the latter is reduced and the potential for remineralisation of experimental white spot caries lesions is enhanced. This study has compared the effects of both gums on plaque pH (measured by the sampling technique) in 10 subjects who chewed the gums after standard acidogenic challenges (sucrose rinse, cupcake, and fried chicken dinner). The results showed that while both gums can significantly reduce the acid response, sugar-free gum appears more effective.

The effect of fluoride and plaque, separately and together on the remineralization of early artificial caries lesions was studied in a group of volunteers in a double-blind randomized study design. Artificial caries lesions produced in slabs of human enamel were mounted buccally in silver bands cemented bilaterally to the mandibular first molar teeth. Volunteers wore the bands for two 2-week periods, rinsing daily with either a placebo or a 500 ppm fluoride mouthrinse. Plaque accumulation was encouraged by placing Dacron gauze over one of each pair of slabs. Sections were cut from the lesions and analyzed by microradiography followed by microdensitometry and image analysis. Microdensitometric data revealed that non-significant increases in mineralization had occurred in all of the test groups. However, the image analysis results indicated that significant remineralization had occurred in lesions after 2 weeks on the fluoride rinse regimen, with no significant remineralization in lesions exposed to the placebo. The presence or absence of plaque over the lesions had no effect on the amount of remineralization induced on the fluoride rinse regimen.

The effect of lactate dehydrogenase (LDH) on human plaque pH and acid anion concentrations in vivo was investigated. Rinsing with sucrose solutions supplemented with LDH (1 or 2 U/ml) gave rise to reduced pH fall, decreased cH area and an increase in the mean minimum pH when compared with rinsing with sucrose only (p less than 0.05). Mean acid anion estimations showed that plaque fluid concentrations of lactate, acetate and proprionate significantly decrease (p less than 0.05) following rinsing with sucrose containing LDH at both levels and decreases in succinate when 1 U/ml LDH was present, whereas formate concentrations were only significantly lower when 2 U/ml LDH was added to sucrose rinses. It is thought that the ability of LDH to produce an alteration in the acid end-products of plaque metabolism may provide some protection against dental caries.

Studies of the extracellular, free concentrations of substrates, growth factors, inhibitors, and end-products of metabolism to which the intact plaque microflora is exposed in situ can assist in the understanding of factors controlling plaque pathogenicity. Information is becoming increasingly available from analysis of fluid separated by centrifugation of plaques collected at various intervals after an intra-oral pulse of dietary or experimental substrate, or different procedures or treatments having cariostatic potential. Such analytical results give more information than those obtained by analysis of aqueous or other extracts, because they yield values of substrate concentration representing those occurring at the bacterial cell surface. The largest body of information concerns extracellular levels of acid end-products of sugar catabolism in relation to food quality or sequence, and of amino acids and other products of nitrogen metabolism, in relation to studies of the detailed metabolic events of the Stephan curve, and of the demineralizing effect of the plaque environment. Areas where little information is available and which merit further study include plaque clearance of salivary and other components with anti-caries activity (e.g., antibodies, enzymes, fluorides, cations, other antimicrobials, etc.), and substrate concentrations to determine gradients for diffusion into and out of plaque.

The primary aim of this study was to rank several reference foods (apple drink, caramel, chocolate, cookie, skimmed milk powder, snack cracker, and wheat flake) according to their plaque pH response as monitored in a panel of 12 volunteers by the plaque-sampling method for comparison with data previously reported with other methods used to assess cariogenicity potential. Secondary experiments (using subsets of the panel of subjects) were undertaken in an attempt to elucidate some of the reasons for the observed plaque pH changes. Oral carbohydrate retention was measured at a single time period after food use as total anthrone-positive carbohydrate material, and as specific acidogenic sugars by gas-liquid chromatography after gel-exclusion chromatography. The concentrations of acid anions in the plaque fluid after food consumption were measured by isotachophoresis eight min after food use. According to the plaque pH response, apple-flavored fruit drink and chocolate were the most acidogenic foods and skimmed milk powder the least acidogenic. There were significant correlations (p less than 0.05) between the plaque pH data and lactate-plus-acetate concentrations in plaque fluid, but the correlations between the pH data and any of the carbohydrate retention parameters were not significant.

Plaque pH in six adult subjects was monitored while they consumed a meal containing three foods. The marked fall in plaque pH caused by drinking sugared coffee was considerably reduced by eating the two non-sugary foods (egg and crisp-bread) before, during, or after the sugared coffee. This favorable action was negated by a pause between the sugary and non-sugary courses.

In studies of the pH response of dental plaque in situ to rinses with conventional sugars, glucose and maltose give similar falls in pH, and fructose only slightly smaller effects compared with sucrose. Lactose and galactose are less acidogenic, while the pH response to glucose syrups varies according to their composition. Of the sugar alcohols, sorbitol, mannitol, maltitol and lactitol are all slowly fermented to acid by oral bacterial, and xylitol is virtually non-fermentable. Acid formation in plaque by fermentable sugar alcohols can be too slow to overcome the buffering power of plaque and saliva and plaque pH can rise following exposure to these compounds. Lycasin, a synthetic sweetener containing sorbitol, maltitol and some higher sugar alcohols gives effects similar to its major constituents. Palatinit, L-sorbose and trichlorogalactosucrose are of low acidogenicity, but coupling sugar is more fermentable and can give rise to a substantial pH response, albeit less than sucrose. Non-nutritive intense sweeteners may affect plaque pH by their sialogogue effects.

In 2 separate studies, plaque was harvested (i) from 15 smokers and 15 non-smokers after a 48-h period without oral hygiene, and (ii) from 15 smokers and 12 non-smokers after a 48 h hygiene-free period following complete plaque removal by toothbrushing. No significant association emerged between wet weight of accumulated plaque and cigarette smoking, in either study. In the second study, the mean plaque calcium concentration was raised in smokers compared with non-smokers, and significantly raised in 7 heavy smokers relative to non-smokers. In both instances, calcium concentration was significantly raised relative to (total) phosphorus concentration, which may indicate that the additional calcium was not in mineral deposits. These results may reflect an association between calcium concentration of plaque and tobacco consumption.

The effects of chewing Parafilm and cheese following sucrose rinsing on human dental plaque pH and plaque fluid organic and amino acid concentrations were investigated. Immediate increases in plaque pH were observed following chewing with concomitant decreases in lactate and acetate concentrations and increases in the concentration of formate and many amino acids. Chewing with cheese when compared with Parafilm resulted in significantly higher (p less than 0.05) plaque fluid concentrations of most amino acids, although significant decreases (p less than 0.05) in phosphate, succinate, and acetate concentrations were observed. However, no significant difference in the levels of formate, lactate, and propionate were found between the two chewing treatments.

Co-60 gamma irradiation of the salivary glands of Macaca mulata monkeys fed a cariogenic diet led to the rapid onset of dental caries resembling that in irradiated human patients. Plaque fluid and saliva were sampled from irradiated monkeys, nonirradiated controls and a group of animals fed a noncariogenic diet in order to look for changes which might occur in inorganic composition related to the caries development and to dietary differences. Salivary calcium and phosphate levels were not markedly changed after irradiation: iodide levels were raised, while thiocyanate levels fell. In plaque fluid, calcium concentrations were not affected by irradiation, but were higher in animals fed a noncariogenic diet. Phosphate levels were higher with a cariogenic diet and further increased in irradiated animals. Magnesium levels were occasionally higher than those of calcium. Other differences in plaque fluid composition may be related to secondary effects of the concomitant gingival disease. The results do not point clearly at a specific change in the quality of the saliva produced by the residual gland tissue after irradiation which precipitates the rampant caries. It is more likely that the great reduction in the quantity of saliva with its protective constituents is responsible.

Irradiation of the major salivary glands of monkeys (Macaca mulatta) fed cariogenic diets leads to caries clinically indistinguishable from radiation caries in man. This study compares the organic composition of individual samples of plaque fluid and saliva from irradiated and control monkeys receiving the same cariogenic diet. Plaque and saliva were collected from fasting, tranquillised animals. Four irradiated animals were sampled repeatedly as were non-irradiated controls. Total protein, albumin, immunoglobulins A, G, and M, and the third component of complement (C'3) were quantitated in plaque fluid and whole saliva. Salivary amylase and peroxidase activities were also determined. Plaque fluid and saliva samples were also subjected to polyacrylamide gel electrophoresis. The total viable anaerobic count and numbers of Streptococcus mutans were determined in samples of plaque. The results suggest that the major effect of irradiation leading to increased numbers of S. mutans and caries susceptibility is in the amount, and not the composition, of the saliva produced by the residual gland tissue. The scanty flow of saliva may reduce the effectiveness of cleansing, buffering and lubrication mechanisms as well as resulting in a marked reduction in the total amount of specific and non-specific immune factors entering the mouth.

The relationship between these factors was studied in plaque and plaque fluid samples taken at intervals during the Stephan pH curve following a sucrose mouth rinse. Levels of lactate rose after the rinse, then fell during the pH recovery phase. Levels of acetate, propionate and phosphate fell after rinsing, then rose again. Amino acid concentrations also changed, with many showing a fall followed by a rise; others rising then falling; and some showing a more variable or complex pattern. In resting plaque fluid, only alanine, proline, glutamic acid, glycine and ammonia were present at concentrations above 1 mmol/l. Delta-aminovaleric acid was detected at levels below those that have been found in monkeys. Hydroxyproline and hydroxylysine were consistently detected, levels of arginine were generally low, and those of cystine consistently very low. The results may provide a basis for understanding the complex metabolic interrelations that occur in the course of the Stephan curve and which may reflect or produce the observed pH changes. They suggest that besides the amount of acid produced, the type of acid, buffering power and base production should be considered as determinants of plaque pH.

The effects on the metabolism of starch by plaques formed in human subjects during periods of dietary sucrose limitation or supplementation were studied. High sucrose (HS) plaques showed lower resting pH and pH minima, and higher concentrations of lactate ion after the starch mouth-rinse than low sucrose (LS) plaques. Plaque samples incubated with starch solutions in vitro showed no differences in final pH or acid-anion concentrations between HS and LS diets. Plaque-amylase activity was higher on HS than LS diets. Thus starch was more acidogenic when consumed as part of a diet already rich in sucrose, but it is unclear whether this was a specific or non-specific effect of starch on plaque metabolism.

Previous reports in the literature indicate that smoking is associated with increased plaque accumulation and/or mineralization. This might be due to an effect of tobacco smoke on properties of saliva. The present study examined the stability of components in fresh and incubated saliva in smokers and non-smokers. Saliva from smokers: collected immediately after smoking a cigarette, did not differ significantly from that of non-smokers in the turbidity developing after incubation (24 h, 37°C) which may reflect aggregation of salivary components by mechanisms analogous to those involved in plaque formation. This finding is in agreement with our previous result (Macgregor, Edgar & Greenwood 1985) which showed that the rate of plaque formation did not differ between smokers and non-smokers. Although an intimation of higher initial calcium concentrations, greater precipitation of calcium and development of more turbidity during incubation was observed in the salivas of smokers than those of non-smokers, the major reason for the greater plaque accumulation and calculus in smokers is probably inadequate oral hygiene. [ABSTRACT FROM AUTHOR]

Dental calculus contains a range of calcium phosphates, derived from plaque fluid and the saliva. Older deposits tend to contain more hydroxyapatite and ß tricalcium phosphate. Compounds which inhibit the transformation of calcium phosphate nuclei to hydroxyapatite and its subsequent crystal growth are known to be effective in restricting calculus deposition in vivo. Such compounds include the diphosphonates, polyphosphates, and zinc. Under physiological conditions, apatites tend to be calcium-deficient with matching HPO42- groups. If these groups are displaced, then crystal growth is inhibited. This can be demonstrated for heated apatite and FPO32- treatment. The latter, however, subsequently hydrolyzes with reformation of the HPO42- groups. Diphosphonates and polyphosphates adsorb upon nuclei, displacing surface phosphate in the process. The treated apatite nuclei will not grow until the adsorbed species are displaced or hydrolyzed. The cationic inhibitor, zinc, is acquired by the dental plaque and oral surfaces and is eluted in the saliva. Zinc is adsorbed upon apatite crystallites with displacement of calcium. When a surface coverage of approximately 3 μmol/m² is achieved, crystal growth is almost completely inhibited. Post-treatment with calcium desorbs the zinc. Inhibition of hydroxyapatite crystal growth and thence calculus formation thus depends upon displacement of surface ions from seed crystallites. The role of HPO42- groups in this and other physiological apatite processes is of particular interest. [ABSTRACT FROM AUTHOR]

The crucial role played by the actions of saliva in controlling the equilibrium between de- and remineralization in a cariogenic environment is demonstrated by the effects on caries incidence of salivary dysfunction and by the distribution of sites of caries predilection to those where salivary effects are restricted. However, of the several properties of saliva which may confer protective effects, it is not certain which are most important. A distinction can be made between static protective effects, which act continuously, and dynamic effects, which act during the time-course of the Stephan curve. Evidence implicates salivary buffering and sugar clearance as important dynamic effects of saliva to prevent demineralization; of these, the buffering of plaque acids may predominate. Enhanced remineralization of white spot lesions may also be regarded as dynamic protective effects of saliva. Fluoride in saliva (from dentifrices, ingesta, etc.) may promote remineralization and (especially fluoride in plaque) inhibit demineralization. The design of experiments using caries models must take into account the static and dynamic effects of saliva. Some models admit a full expression of these effects, while others may exclude them, restricting the range of investigations possible. The possibility is raised that protective effects of saliva and therapeutic agents may act cooperatively. [ABSTRACT FROM AUTHOR]

Information about several papers discussed at a symposium on dental plaque fluid sponsored by the Cariology Group and the Mineralized Tissue Group is presented. Topics of the symposium included the development of research on dental plaque fluid, analytical techniques used in examining fluid sites and modeling studies of the caries process.