|Year : 2021 | Volume
| Issue : 1 | Page : 74-78
|Assessment of salivary thiocyanate levels and pH in the saliva of smokers and nonsmokers with chronic periodontitis – A comparative study
C Naresh Kumar1, Subramaniam M Rao2, Ankur Jethlia3, Chandra Shekhar Linganna4, Manish Bhargava5, Devendra H Palve6
1 Department of Periodontology, G Pulla Reddy Dental College and Hospital, Kurnool, Andhra Pradesh, 9, India
2 Department of Periodontology, P.M. Nadagouda Memorial Dental College and Hospital, Bagalkot, Karnataka, India
3 Department of Maxillofacial Surgery and Diagnostic Sciences, Diagnostic Division, College of Dentistry, Jazan University, Saudi Arabia
4 Department of Dentistry, Sridevi Institute of Medical Science, Tumkur, Karnataka, India
5 Department of Oral Pathology, Manav Rachna Dental College, Faridabad, Haryana, India
6 Department of Oral Pathology and Microbiology, Swargiya Dadasaheb Kalmegh Smruti Dental College & Hospital, Nagpur, Maharashtra, India
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|Date of Submission||06-May-2019|
|Date of Decision||18-Feb-2020|
|Date of Acceptance||30-Nov-2020|
|Date of Web Publication||13-Jul-2021|
| Abstract|| |
Background: It has been speculated that the pathogenesis of diseases prompted by cigarette smoking includes oxidative damage by free radicals. Though, definitive evidence that smoking may cause the oxidative modification of target molecules in vivo is lacking. Therefore, the purpose of this study was to estimate and compare the levels of salivary Thiocyanate (SCN) and power of hydrogen (pH) in the saliva of smokers and nonsmokers with chronic periodontitis (ChP). Materials and Methods: A study population consisted of 60 male systemically healthy subjects in the age group of 20-65 years that was further divided into three groups: Group 1: 20 Healthy nonsmokers, who never smoked. Group 2: 20 nonsmokers with chronic periodontitis. Group 3: 20 smokers with chronic periodontitis. Unstimulated saliva was collected for at least 5 mins and clinical parameters; salivary pH and SCN thiocyanate levels were assessed using the spectrophotometric method. Statistical analysis was performed using SPSS Inc 21.0, Chicago, II, United States of America. Results: Data showed that the mean salivary SCN level, periodontal parameters were higher in smokers with chronic periodontitis as compared to nonsmokers with chronic periodontitis and healthy subjects (P < 0.05.) Post Hoc tests multiple comparisons Tukey Honest Significant Difference (HSD) among three groups were statistically significant (p < 0.05). Conclusion: Salivary thiocyanate levels remain increased by smoking in addition to the impact of periodontitis, and these results also indicated a significant change in the pH depending on the severity of the periodontal condition in smokers. Thus, the measurement of salivary thiocyanate may prove to be useful in the early detection of periodontal disease. The salivary pH shows significant changes and, consequently, relevant to the severity of the periodontal disease. Salivary pH may thus be used as a quick chairside diagnostic biomarker.
Keywords: Antioxidants, chronic periodontitis, cigarette smoking, pH, saliva
|How to cite this article:|
Kumar C N, Rao SM, Jethlia A, Linganna CS, Bhargava M, Palve DH. Assessment of salivary thiocyanate levels and pH in the saliva of smokers and nonsmokers with chronic periodontitis – A comparative study. Indian J Dent Res 2021;32:74-8
|How to cite this URL:|
Kumar C N, Rao SM, Jethlia A, Linganna CS, Bhargava M, Palve DH. Assessment of salivary thiocyanate levels and pH in the saliva of smokers and nonsmokers with chronic periodontitis – A comparative study. Indian J Dent Res [serial online] 2021 [cited 2023 Sep 24];32:74-8. Available from: https://www.ijdr.in/text.asp?2021/32/1/74/321372
| Introduction|| |
Periodontitis is an inflammatory disease that is affected by both periodontal pathogens and the host inflammatory responses. The imbalance between these two factors results in a loss of connective tissue attachment and alveolar bone breakdown. Smoking remains the most critical risk factor, with evidence suggesting that its periodontal effects occur regardless of what is smoked. Smoking is thought to exert its influence through affecting neutrophil function, causing shifts to a more pathogenic microflora, and causing sustained peripheral vasoconstriction.,
Cigarette smoking accounts for approximately half of periodontitis diagnosed in young adults. Smokers are almost three times more likely to show severe periodontal disease compared with nonsmokers. Current smokers were also 3.3 times more likely to attend a periodontal practice office compared to nonsmokers. The effect of smoking on periodontal tissues is cumulative and dose-dependent. Clinically, the smoking-associated periodontal disease presents with thick inflamed marginal gingiva and gingival recession. The buccal marginal gingiva of both upper and lower anterior teeth often shows the characteristic stain of smoker melanosis., The degree of alveolar bone destruction far exceeds the periodontal obliteration evident clinically. Thus, cigarette smoking is the single, modifiable environmental factor responsible for the excess prevalence of the periodontal disease in the population. The diagnosis of periodontal disease has been primarily based upon clinical and radiographic measures of periodontal tissue destruction. These parameters provide a quantity of past destruction and are of limited use in early diagnosis. Thiocyanate (SCN) ion is found inorganic and organic complexes and is a normal component of body fluids such as saliva, serum, tears and urine. Therefore saliva contains peroxidase enzymes and lysozyme among many other host innate defense systems. The saliva has three complete peroxidase components salivary peroxidase, peroxidase enzyme and myeloperoxidase, hydrogen peroxidase (H2O2), and an oxidizable substrate such as pseudohalide thiocyanate (SCN). Thiocyanate reacts with H2O2 to produce hypothiocyanite, which has antimicrobial properties. Also, thiocyanate plays a significant role in the peroxidase system by decreasing the toxicity of H2O2 produced by oral bacteria by reacting with it to produce less harmful hypothiocyanite., It has been stated that salivary thiocyanate concentrations in together stimulated and resting saliva to have a converse relation with gingival inflammation, and amount of plaque.
It is known that periodontal diseases in humans and other mammals are predominantly associated with Gram-negative anaerobic organisms and that before destructive periodontal diseases are initiated, these microorganisms must colonize tooth surfaces at and slightly below the marginal gingiva. Strong evidence exists to consider Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis Tannerella forsythia as etiologic agents. A study by Takahashi et al., on the effect of pH on the growth of microorganisms showed that P. gingivalis cultivates at a pH of 6.5-7.0, P. intermedia cultivates at a pH of 5.0-7.0 and F. nucleatum grow at a pH of 5.5-7.0. Accurate measurement of cigarette smoking is critical for understanding patterns of adult smoking behavior and essential for evaluating health education programs aimed at reducing or preventing the habit. Chemical measures such as salivary SCN show promising results in obtaining accurate, quantitative information on smoking habits.
Hence, to the simplest of our knowledge, there's no current information examining the salivary thiocyanate levels and hydrogen ion (pH) concentration in smokers and nonsmokers with chronic periodontitis. Hence, an attempt was made in this study to research the impact of smoking standing on the salivary level of SCN in smokers and nonsmokers with chronic periodontitis.
| Materials and Methods|| |
A total of 60 male patients aged 20-65 years of age were recruited from the outpatient Department of Periodontology, AECS Maaruti College of Dental Sciences and Research Centre, Bangalore, Karnataka, India. Sample size was calculated using G*power 22.214.171.124 considering a standard deviation of 1.9 mg% of salivary thiocyanate level using 80% power and 95% confidence level. None of the patients had received periodontal therapy or used antibiotics, anti-inflammatory agents, regular mouth wash, vitamin supplements, and any special dietary supplements in the last three months. Patients with systemic diseases like diabetes mellitus, cancer, coronary heart disease, hepatitis, cardiovascular disease, HIV infection and epilepsy were excluded from the study.
Group 1: 20 Healthy nonsmokers, who had never smoked, with no history of periodontal disease nor tooth loss caused by periodontitis and had no clinical signs of periodontitis (clinical attachment level [CAL] ≤1 mm, probing pocket depth [PPD] ≤3 mm, gingival index [GI] <1).
Group 2: 20 patients included for nonsmokers with chronic periodontitis group were diagnosed according to the classification of the 1999 American Academy of Periodontology workshop with the criteria of ≥20 residual teeth having ≥1 tooth with sites of pocket probing depth (PPD) ≥4 mm and clinical attachment level (CAL) ≥4 mm in all four quadrants.
Group 3: 20 smokers with chronic periodontitis who claimed to have smoked at least ten cigarettes per day for the past five years at the minimum.
Subjects were explained about the study and, based on their approval, were asked to read carefully and sign the consent form. The design of the study and procedures for obtaining informed consent was approved by the Ethical Committee and was performed in accordance with the Code of Ethics of the World Medical Association according to the Declaration of Helsinki of 1975 as revised in 2000.
Plaque index (PI), bleeding on probing dichotomous method (BOP was expressed as a percentage of all sites), probing depth (PD in mm), and clinical attachment level (CAL in mm) were measured at six sites and recorded on each tooth, except third molars. All clinical periodontal measurements were performed by the single examiner by using William's graduated periodontal probe (Hu-Friedy Mfg. Co., Rockwell St. Chicago).
The patients were scheduled to report between 9 am to 12 pm and were seated for 30 minutes before the sampling procedure and also instructed not to eat or drink two hours prior to saliva collection. The smokers were also prohibited from smoking one hour prior to saliva collection. The participants were instructed to rinse the mouth using distilled water. The unstimulated saliva was collected for at least 5 minutes in a sterile centrifuge tube (Tarsons products Pvt Ltd® Kolkata, India), and stored at -20° C until analyzed. Saliva samples were centrifuged at 3800 rpm for at least 10 min.
Analyses of thiocyanate activity
A volume of 0.5 ml of supernatant saliva obtained after centrifugation was mixed with 9.5 ml Fe(NO3)3 reagent; the following reaction occurs:
Fe+3 (aq) + SCN (aq) → FeSCN2+ (aq)
This FeSCN2+ complex exhibits deep red color, which can be conveniently measured with UV spectrophotometry at 447 nm wave lengths. The FeSCN2+ concentration of saliva solution was calculated through Lambert and Beers law.
Analyses of salivary pH
Dental salivary pH indicator strips [iGen pH test strips, China [Figure 1]] were used for the analysis of salivary pH 4.5-9.0.
Statistical analysis was performed using a software program SPSS Inc 21.0, Chicago, II, United States of America. Salivary thiocyanate, pH of saliva and clinical parameters were assessed using a one-way analysis of variance (ANOVA), followed by post hocTukey test when ANOVA suggested a significant difference among groups (P < 0.05).
| Results|| |
Data showed that the mean salivary thiocyanate levels 10.9 ± 1.2 mg% found in smokers with chronic periodontitis which were significantly higher (P < 0.005) as compared to nonsmokeres with chronic periodontitis 6.67 ± 0.85 mg % and healthy 3.09 ± 1.07 mg %. Significantly higher PPD 5.32 ± 0.17 mm, BOP 58.6 ± 0.6%, CAL 4.12 ± 0.14 mm, was shown in smokers with chronic periodontitis compared with other groups [Table 1]. It was found that pH of saliva in smokers with chronic periodontitis was acidic 6.10 ± 0.6 when compared with that of nonsmokers with chronic periodontits 6.30 ± 0.40, and pH of saliva was alkaline in healthy group 7.05 ± 0.56 (P < 0.05) [Table 1]. Post Hoc tests multiple comparisons Tukey Honest Significant Difference (HSD) among group 1 vs. group 2 and group 3; group 2 vs. group 1 and group 3; and group 3 vs. group 1 and group 2 were statistically significant (P < 0.05) [Table 2].
|Table 1: Salivary thiocyanate levels, pH and clinical parameters in Group 1, Group 2 and Group 3|
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|Table 2: Post Hoc tests multiple comparisons Tukey Honest Significant Difference (HSD)|
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| Discussion|| |
This study sets out to assess the association between smoking and periodontal disease through a life stage in the natural history of periodontitis about which little is known. The hypothesized increase in strength of important periodontal risk factors was observed, with smoking being particularly important. Intra-oral differences in periodontitis occurrence became more pronounced, with more BOP, PI, PD and CAL. In the present study, higher levels of salivary Thiocyanate and increased PI, BOP, PD and CAL with decreased salivary pH were observed in smokers with chronic periodontitis compared with nonsmokers with chronic periodontitis and healthy subjects. In the present study, female smokers were not evaluated because of their low prevalence in India.
Compelling reasons exist to use saliva as a diagnostic fluid. It meets the demands of being inexpensive, non-invasive, and easy to use diagnostic methods. As a clinical tool, saliva has many advantages over serum, including ease of collection, storing and shipping, and it can be obtained at low cost in sufficient quantities for analysis. For patients, the non-invasive collection techniques dramatically reduce anxiety and discomfort and simplify the procurement of repeated samples for monitoring over time. Saliva also is easier to handle for diagnostic procedures because it does not clot, thus lessening the manipulations required.,
According to Sanchez-Perez et al. saliva plays an important role in oral health monitoring, regulating and maintaining the integrity of oral mucosa. Saliva is necessary for the protection and lubrication of oral mucosal tissues, remineralization of teeth, digestion, taste sensation, stimulation, wash-out effect, pH balance and phonation. Human saliva contains a large number of solid (organic and inorganic) constituents such as proteins, sodium, potassium, SCN, immunoglobulins, etc. SCN levels in saliva are the most frequently used biochemical tests for establishing the incidence or prevalence of tobacco consumption among smokers.
In our study, the salivary SCN level was significantly increased in smokers with chronic periodontitis as compared to nonsmokers with chronic periodontitis. This is due to the fact that the main source of SCN is tobacco smoke, which is absorbed in the lungs and later metabolized to SCN (Benfari et al. Tenovuo). Borgers and Burckhard found that the high concentration of cyanide in tobacco smoke may persist in the oral cavity of smokers for some time, and the saliva thus contaminated may artifactually increase the cyanide level. Courant observed significantly higher concentrations of SCN in the saliva of smokers. This was consistent with our results and was also reported in various other studies (Tenovuo et al. Luepker et al. Lamberts et al.), which are also in agreement with our study.
According to Foss et al. SCN has a high specificity for heavy smokers, as compared to light smokers. Bliss et al. indicated that an SCN assay shows excellent sensitivity (over 90%) in chronic smokers who smoke more than 15 cigarettes per day, whereas in light (one to five cigarettes per day) and moderate smokers (five to ten cigarettes per day), the sensitivity of the SCN level was not found to be sigificant. This is consistent with the results of the present study as smokers included in our study were those who smoke at least ten cigarettes per day for more than five years. In our study, the pH of saliva was acidic in smokers with chronic periodontitis when compared with nonsmokers with chronic periodontitis and healthy. There are fewer studies on pH and smoking. A salivary concentration pH of 7.0 usually indicates a healthy dental and periodontal situation. At this pH, there is a lower-frequency of tooth decay combined and little or no calculus. Therefore, stable conditions ought to primarily be found during this environment. A saliva pH below 7.0 usually indicates acidemia (abnormal acidity of the blood). If a chronic condition exists, the mouth is a lot of at-risk of tooth decay, halitosis and periodontitis. Chronic acidemia can be a causative factor for a multitude of diseases affecting the whole body. A saliva pH above 7.0 usually indicates alkalinity. Excessive alkalinity can bring about the same anaerobic conditions as acidemia, but it is a much rarer condition. Plaque bacteria take calcium compounds in the environment and use the minerals to protect them from the high pH. The two key factors to plaque formation are; first, there must be oral bacteria to attack food particles and elevate the pH. Second, the pH must elevate above 7.6 to grow dental plaque crystals that cause periodontal disease. Thus, alkaline pH is essential for plaque growth suggesting the mildly alkaline pH of the saliva obtained from the subjects with generalised chronic gingivitis.
The limitations of the present study were it was conducted only among self-reported male smokers, and further work is to be carried out to determine the correlation of the enzyme in female smokers. Previous studies have found that there is a low prevalence of female smokers in India (≤4%). The smokers were purely categorised as those who had smoked more than ten cigarettes per day for more than five years and nonsmokers, based on self-reports. Former smokers were not included in the present study, and smokers were not categorized into light and heavy smokers and chronic periodontitis group were not classified further into mild, moderate and severe periodontitis.
Therefore, further studies incorporating a larger sample size and longitudinal studies, including female smokers, coupled with the estimation of serum cotinine assays are warranted. The preventive intervention has to consider, which includes oral health education, behavioral modification, tobacco cessation with nicotine replacement therapy, and appropriate periodontal treatment and follow up.
| Conclusion|| |
Salivary thiocyanate levels are increased by smoking in addition to the impact of periodontitis, and these results also indicated a significant change in the pH depending on the severity of the periodontal condition in smokers. Thus, the measurement of salivary thiocyanate may prove to be useful in the early detection of periodontal disease. The salivary pH shows significant changes and thus relevant to the severity of the periodontal disease. Salivary pH may thus be used as a quick chairside diagnostic biomarker.
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Conflicts of interest
There are no conflicts of interest.
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] [Full text]
Dr. Subramaniam M Rao
Department of Periodontology, P.M. Nadagouda Memorial Dental College and Hospital, Bagalkot, Karnataka
Source of Support: None, Conflict of Interest: None
[Table 1], [Table 2]
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