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Effect of consuming different dairy products on calcium, phosphorus and pH levels of human dental plaque: a comparative study.


Milk and dairy products have a low cariogenic potential, but they are also claimed to be cariostatic [Johansson, 2002]. Studies to date have demonstrated that milk and milk products such as cheese and yogurt are beneficial to dental and oral health. They play a key role in preventing adverse dental issues such as dental caries and periodontitis [Al-Zahrani, 2006]. It has been shown that calcium, phosphorous and various bioactive peptides found in milk have important functions in the maintenance of dental health, through enamel protective and anti-caries effects [Merritt et al., 2006].

Dairy products particularly cheese have very low cariogenic potential and are therefore safe for teeth. According to available evidence chewing of hard cheese particularly cheddar cheese can prevent enamel demineralisation via at least two different mechanisms. Firstly by stimulating salivary flow which buffers dental plaque pH and secondly by increasing the calcium and phosphorus concentrations in dental plaque thus favouring remineralisation [Rugg-Gunn et al., 1975; Silva et al., 1986]. Enamel rehardening has been shown to occur with soft cheeses [Lewinstein et al., 1993] and even processed cheese can be cariostatic [Drummond et al., 2002] It was suggested that a substantial portion of the protective effect of cheese might be related to textural influences along with the casein or calcium phosphate content of cheese [Saroglu Sonmez and Aras, 2007].

There is a large volume of data that suggests milk and other milk products contain numerous bioactive compounds conducive to prevent dental caries [Merritt et al., 2006]. There is evidence that the addition of milk and milk products increases the calcium and phosphate content of food that in turn diminishes the extent of tooth demineralisation and enhances remineralisation [Kashket et al., 2002].

One of the objectives of dental research has ben to identify factors in foods that can protect the teeth against caries. Two major advantages of protective agents found in milk and/or milk products are that (1) it would be perceived as a natural product and (2) milk is recommended on account of its nutritional properties, chiefly as a source of calcium and protein, and accordingly its use does not raise any toxicological problems [Grenby et al., 2001].

The concentration of calcium and phosphate ions in plaque is important because these are the same elements within the hydoxyapatite crystals. If the plaque fluid adjacent to the tooth is supersaturated with calcium and phosphorus ions at a given pH, the enamel certainly cannot undergo demineralisation at its surface. Calcium is bound to proteins in the plaque. As the pH of plaque drops following acidogenesis, dissolution of enamel occurs and calcium and phosphate are liberated along with other bound ions in the plaque. In this way all the minerals in the plaque that are more soluble than hydroxyapatite serve to protect the apatite crystals by increasing the ionic saturation of the threatened area. Thus the present study is aimed to evaluate the effect of consuming different dairy products without any added sugar (sucrose) on the calcium, phosphorus and pH levels of human dental plaque.

Materials and methods

The present study was conducted to assess the calcium, phosphorus and pH levels of human dental plaque after consuming three different dairy products (milk, cheese and yogurt). The convenience sample of 68 voluntary students (34 with caries and 34 caries-free), aged 17-20 years were recruited from a dental college. Subjects who gave informed consent and those satisfying the inclusion criteria were included in the study. Subjects undergoing any orthodontic treatment, exhibiting dental fluorosis, antibiotic therapy or other chemotherapeutic procedure that could have an effect on plaque formation were excluded from the study.

The study subjects were classified as caries-free (CF) and caries groups (CG). From each group the study subjects were randomly divided into subgroups of cheese (n=8), milk (n=8), yogurt (n=8) and paraffin wax (control) (n=10). The ionic calcium, inorganic phosphorus and pH levels were assessed before and after consumption of dairy products. Ethical clearance for the study was obtained from the local institutional ethical committee before the start of study

Method of plaque collection and calcium, phosphorus and pH estimation at baseline. All the study subjects were asked to refrain from all oral hygiene measures for 48 hours and not to take any food before collection of plaque samples on the day of plaque collection in order to reduce the effect of food on calcium, phosphorus and pH levels of the plaque. Just before the baseline plaque estimations all the study subjects were made to rinse their mouth with plain water.

Pooled plaque samples were collected by removing all visible plaque from the 1st and 3rd quadrants of the mouth using sterile stainless steel Jacquette scalers and avoiding obvious food debris. Plaque was not collected from the mandibular incisors, as the plaque from this region, being close to the salivary glands, has a very high concentration of calcium and phosphorus and any small differences in the amount of plaque sampled from this area could have a marked influence on the results [Ashley, 1975].

A part of the harvested baseline pooled plaque sample was suspended in 0.4% EDTA (ethylene diamine tetra acetate) and thoroughly agitated for 5 minutes, so that all plaque particles could easily be dissolved. Plaque ionic calcium concentration was analysed using an electrolyte analyser (Spectrolab-K, Model no. 1600 DR-K, UV- Vis Metrolab S.A) and inorganic phosphorus by the method of Chen and Toribara [1956]. Plaque pH was determined using a standard portable digital pH meter with a glass microelectrode (model Eq- 612 Elicoelectronics, Mumbai, India).

Analysis of plaque after consuming dairy products. After the baseline estimation of plaque the subjects were allowed to chew and swish the different dairy products (cheese 5g, milk 15 ml and yogurt 5g) and paraffin wax (control) for 3 minutes followed by swishing their mouth with plain water so as to reduce the contamination of plaque with soft debris. The pooled plaque samples from the 2nd and 4th quadrants were analysed for ionic calcium, inorganic phosphate and pH levels after 10 mins using similar procedures. Similarly the pH of the control group was analysed for the pH changes before and after chewing paraffin wax. Statistical analyses were carried out using SPSS software package 15.0. Comparison between the caries-active and caries-free groups were performed using unpaired t-tests and between before and after eating dairy products using paired t-tests and the correlation coefficients were analysed between calcium, phosphorus and pH of dental plaque.


The result showed that the consumption of dairy products particularly cheese in caries-active subjects showed a significant rise in the mean plaque calcium and phosphorus levels followed by the yogurt group, while the milk group showed the least change and it was statistically non-significant. In the caries-free group, cheese and yogurt showed a statistically significant rise in the mean plaque calcium and phosphorus levels, among which the cheese group showed the highest rise followed by the yogurt and then the milk group. The rise in phosphorus levels for the milk group was non-significant. The control group showed the least and non-significant rise in the mean calcium levels in both caries-active and caries-free groups. While comparing the mean plaque calcium and phosphorus levels between caries-active and caries-free subjects, a statistically significant difference was observed after consuming cheese and milk, as the amount of rise in calcium and phosphorus concentration was greater in caries-free subjects as compared with caries-active subjects, while the yogurt and control groups showed no significant differences between the two groups (Table 1).

In both caries-active and caries-free subjects there was a strong correlation between plaque pH and calcium levels particularly in the cheese group (r = 0.763 **, 0.817 **) followed by the yogurt and milk group. Similarly, a strong correlation was observed between plaque pH and plaque phosphorus in caries-active and caries-free subjects, where the cheese group showed a strong correlation (r = 0.650 *, 0.585 **) followed by the yogurt and milk groups. The cheese and yogurt groups showed a statistically significant correlation with the calcium group. Only the cheese group showed a statistically significant correlation with the phosphorus group. The control group showed the least and non-significant correlation (Table 2).

Table 3 records that there was an increase in plaque pH after consumption of cheese. Caries-free subjects showed a greater increase. Milk and yogurt groups showed a decline in plaque pH, among which the yogurt group showed a greater decline in plaque pH when compared to the milk group. For the control group there was a slight increase in plaque pH in both caries-active and caries-free groups.


Food groups such as milk and milk products have low cariogenic potential and are most commonly recognised as exhibiting anti-caries activity [Aimutis, 2004]. The anti-cariogenic activity of milk and its products was attributed to the direct chemical effects of casein, phosphopeptides, calcium and phosphate [McDougall, 1977; Reynolds, 1998]. Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) has the ability to bind and stabilise calcium and phosphate in solution, as well as binding to both dental plaque and tooth enamel. The high concentration of calcium and phosphate ions in dental plaque have been extensively researched and proven to reduce the risk of enamel demineralisation and promote remineralisation of tooth enamel [Walsh, 2000]. It is customary in studies of this nature to ask the subjects to abstain from oral hygiene procedures for a set period of time prior to plaque collection. This may be preceded by a thorough scaling and polishing in order to standardise the baseline experimental condition. However, such standardisation may not accurately reflect the relationship in an older plaque which has been subjected to variable oral hygiene procedures. The investigation into the calcium and phosphorus concentrations of plaque which are normally present in subjects mouths may be of more practical significance. Such plaque will be referred to as "mature" and "pooled" even though its age will vary according to the subject's oral hygiene habits and dental treatment [Ashley, 1975].

The present study revealed that after the consumption of dairy products (cheese, milk and yogurt), the cheese group showed the highest rise in the mean plaque ionic calcium and inorganic phosphorus levels. The rise in phosphorus levels were less compared to calcium as it would be expected that some of the phosphate may be precipitated or form a colloidal suspension or may be bound to casein [Silva et al., 1987]. The results of this study are comparable to previous studies [Rugg-Gunn et al., 1975; Silva et al., 1986].

The greater mean plaque calcium levels observed after cheese consumption may be due to the calcium in cheese existing in a readily available form for diffusion into plaque. Another view is that casein from cheese or phosphopeptides from cheese form complexes which stabilise colloidal calcium phosphate and favours their diffusion into plaque [Moynihan et al., 1999]. There was also a significant correlation between plaque pH and plaque calcium, which suggests that increased calcium levels are associated with greater plaque pH levels.

The effect of cheese in raising plaque pH has also been observed by several other investigators [Rugg-Gunn et al., 1975; Silva et al., 1986]. Thus in summary cheese has two mechanisms of protection against enamel demineralisation: (1) by releasing calcium and phosphorus and therefore increasing their concentrations in the dental plaque, which inhibits demineralisation and favours remineralisation by a common-ion effect; and (2) by stimulating salivary flow and, consequently, buffering the dental plaque and exerting a sugar-clearing effect [Silva et al., 1986].

The yogurt group also showed a rise in mean plaque calcium and phosphorus levels slightly less than for the cheese group. The results of our study are similar to those of Ferrazzano et al. [2008]. Yogurt is very rich in calcium and phosphorus. Additionally, natural CPPs content in yogurt is higher than that in milk; due to proteolytic activity of microorganisms contained in the yogurt [Rasic and Kurmann, 1983].

Moreover, because of the lower pH of yogurt compared with that of milk, calcium is present in yogurt mostly in ionic form [Adolfsson et al., 2004]. In fact, fermentation has little effect on the mineral content of milk and therefore the total mineral content remains unaltered in the yogurt

It has been suggested that bacteria present in yogurt may have a positive effect on the ecology of dental plaque, reestablishing the microbial homeostasis broken down by the bacterial metabolism. Recent studies reported that yogurt and probiotic-containing milk and cheese consumption has been associated with a decrease in mutans streptococci in dental plaque and/or saliva [Nikawa et al., 2004; Caglar et al., 2005]. This hypothesis could represent a favourable outcome in terms of prevention of dental caries.

However, the milk group also showed a rise in mean plaque calcium and phosphorus levels, but the rise was less and statistically non-significant. The results of our study are comparable with that of Bowen and Pearson [1993]. In comparison with the cheese and yogurt groups, the milk group showed less of a rise in mean plaque calcium and phosphorus levels. This may be because, in milk, calcium phosphate is packaged in the inner portion of micelles and casein is the component of the inner shell of micelles [Walsh., 2000]. While comparing the milk group with the control group, the milk group showed a greater rise in mean plaque calcium and phosphorus levels. This is because of greater concentration of calcium, phosphorus and casein in milk as compared with the control (paraffin group). To increase the effectiveness of milk in remineralising enamel lesions it has to be fortified with 2.0-5.0 gram of CPP-ACP [Walker et al., 2006]. Milk also possesses many of the biological and physical properties desired for a saliva-substitute.

The present study also showed that the mean rise of ionic calcium and phosphorus concentration was higher in dental plaque of the caries-free group as compared with the plaque of the caries group, as the existence of a relationship between plaque calcium and phosphorus levels and caries is not entirely unexpected. Jenkins [1966], when discussing the importance of the saturation of saliva with calcium phosphate in preventing the dissolution of tooth surfaces, stressed that in caries it is the equilibrium between enamel and plaque, rather than between enamel and saliva that is important. Dawes and Jenkins [1962] associated the relative caries resistance of lower anterior teeth with the higher calcium and phosphate content of the plaque in this region. It has been suggested that critical pH would be lowest of all in this type of plaque as it seemed possible that these elements (calcium and phosphorous) would pass into solution with a fall in pH and maintain the aqueous phase in a saturated condition [Schroeder, 1963].

Our results are comparable with the study of Ashley [1975] in which a significant inverse relationship was reported between the plaque inorganic phosphorus level and caries experience. The reason for the increase in the mean concentration of inorganic phosphorus in the caries-free subjects is due to the presence of dicalcium phosphate and trimetaphosphate which increases enamel hardness. This may be due to the ability of these phosphates to remineralise or change the chemical composition of the enamel surface to a more insoluble apatite.

Although there was a rise in calcium and phosphorus concentration in dental plaque after yogurt consumption, there was no significant difference between caries-active and caries-free groups. This suggests that caries activity of an individual will not have any impact on calcium and phosphorus deposition in dental plaque by yogurt consumption. This may be due to the probiotic microorganisms present in yogurt which breaks down the metabolism of virulent microorganisms like Streptococcus mutans, and higher concentrations of calcium and phosphate would result in an increase in plaque pH [Adolfsson et al., 2004; Caglar et al., 2005].


Cheese and yogurt without added sugar (sucrose) are potentially non-cariogenic and to some extent cariostatic as they increases calcium and phosphorus and pH levels in dental plaque. Dairy products can be recommended as desserts after meals especially to school children, which would help to reduce the incidence of dental caries. Considering the public health perspective for developing countries like India, among all the three dairy products, yogurt should be preferred as it is easily available and its calcium and phosphorus content is higher, as well as containing enzymes (lactoferrin) and probiotic microorganisms that may hinder the metabolism of virulent microorganisms such as Streptococcus mutans, and re-establishes microbial homeostasis.


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T.L. Ravishankar *, V. Yadav *, P.S. Tangade *, A. Tirth *, T.R. Chaitra **

* Department of Public Health Dentistry, **Department of Pedodontics & Preventive Dentistry, Kothiwal Dental College and Research Centre, Mora Mustaqueem, Moradabad, India.

Postal address: Dr T.L. Ravishankar, Dept. of Public Health Dentistry, Kothiwal Dental College and Research Centre, Kanth Road, Moradabad-244001, Uttar Pradesh, India.

Table 1: Comparison of mean [+ or -] S.D. ionic calcium and phosphorus
(mmol/l) levels between caries-active and caries-free subjects after
consumption of dairy products.

Group         Calcium

              Caries-active        Caries-free

Cheese    B   4.73 [+ or -] 0.14   5.11 [+ or -] 0.12
          A   6.08 [+ or -] 0.72   8.23 [+ or -] 0.45 *
p value       0.002 *              0.000 *
Milk      B   4.70 [+ or -] 0.44   5.30 [+ or -] 0.32
          A   4.83 [+ or -] 0.41   5.67 [+ or -] 0.27 *
p value       0.656                0.017 *
Yogurt    B   4.74 [+ or -] 0.25   4.95 [+ or -] 0.19
          A   5.67 [+ or -] 0.28   5.84 [+ or -] 0.32 **
p value       0.000 *              0.000 *
Control   B   4.98 [+ or -] 0.02   5.19 [+ or -] 0.09
          A   5.03 [+ or -] 0.04   5.23 [+ or -] 0.25 **
p value       0.057 **             0.665 **

Group         Phosphorus

              Caries-active        Caries-free

Cheese    B   3.90 [+ or -] 0.18   4.62 [+ or -] 0.91
          A   4.78 [+ or -] 0.35   6.51 [+ or -] 0.91 *
p value       0.002(S)             0.262(S)
Milk      B   4.02 [+ or -] 0.54   4.21 [+ or -] 0.56
          A   4.12 [+ or -] 0.65   4.82 [+ or -] 0.61 *
p value       0.770 **             0.162 **
Yogurt    B   3.97 [+ or -] 0.37   4.12 [+ or -] 0.31
          A   4.72 [+ or -] 0.45   5.42 [+ or -] 0.42 **
p value       0.011 *              0.001 *
Control   B   4.11 [+ or -] 0.15   4.62 [+ or -] 0.33
          A   4.40 [+ or -] 0.38   4.98 [+ or -] 0.08 **
p value       0.46 **              0.58 **

* Significant ** Non-significant. Paired t-test to compare
caries-free and caries groups, unpaired t-test to compare
before and after consumption of dairy products

Table 2: Correlation between pH, calcium and phosphorus
after consumption of dairy products.

Group      Caries-active r value     Caries-free r value

           Calcium    Phosphorus    Calcium    Phosphorus

Cheese     0.763 *     0.650 *      0.817 *     0.585 *
Milk       0.391 **    0.459 **    -0.586 *    -0.462 **
Yoqurt    -0.643 *    -0.476 **    -0.594 *    -0.487 **
Control    0.125 **    0.144 **     0.373 **    0.205 **

* Significant ** Non-Significant

Table 3: Mean  [+ or -]  S.D. plaque pH of subjects
(caries and caries-free) at baseline and after 10 minutes
among cheese, milk, yogurt and control groups

Dairy Products        Baseline            10 Minutes

Cheese Group
Caries           6.10 [+ or -] 0.13   6.83 [+ or -] 0.12
Caries-free      6.96 [+ or -] 0.15   7.36 [+ or -] 0.13
Milk Group
Caries           6.14 [+ or -] 0.09   5.94 [+ or -] 0.09
Caries-free      7.04 [+ or -] 0.23   6.75 [+ or -] 0.17
Yoqurt Group
Caries           6.13 [+ or -] 0.18   5.59 [+ or -] 0.09
Caries-free      7.00 [+ or -] 0.09   5.63 [+ or -] 0.17
Control Group
Caries           6.10 [+ or -] 0.17   6.22 [+ or -] 0.19
Caries-free      6.90 [+ or -] 0.16   6.97 [+ or -] 0.15
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Author:Ravishankar, T.L.; Yadav, V.; Tangade, P.S.; Tirth, A.; Chaitra, T.R.
Publication:European Archives of Paediatric Dentistry
Article Type:Report
Geographic Code:9INDI
Date:Jun 1, 2012
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