INTRODUCTION:

Whole Saliva is a mucoserous fluid which acts as safeguard for oral cavity^1,2. Saliva withits variable characteristic has a diagnostic role in different oral diseases^3,4,5,6 it is physical and chemical property has vital role in oral health as saliva pH buffer capacity and flow rate is important for caries control^7,8,9.

Unstimulated whole salivary flow rate is a basal flow rate while stimulated salivary flow rate represents the flow rate during stimuli such as during food intake.

Unstimulated salivary flow rate remains longer in the mouth and has important role for protection of oral tissue, while stimulated saliva remains for up to two hours so unstimulated salivary flow rate was considered in this study⁸.

Salivary pH is the most important features of saliva and directly correlated to buffering capacity ^[10].Normal salivary pH ranging 6.2 to 7.6, usually maintained near neutrality, between 6.7-7.3^11,12. Salivary buffer capacity is the ability of saliva to neutralize the pH to neutral pH after exposure to acidic or basic condition. Bicarbonate, phosphate and protein are the three buffers present in saliva; salivary buffer activity is one of the main functions of saliva^13,14,15,16.Saliva can buffer acids and maintain oral pH therefore protects oral cavity from acids that is produced by bacteria and is one of the main causes of dental demineralization so buffering capacity has positive role in mineralization^17,18.

As salivary flow rate is differs according to many variables such as age and gender and health status in addition buffer capacity directly can be affected by flow rate^19,20,21. Therefore in this study flow rate buffer capacity and pH studied in two age categories of normal adults in order to describe the changes of buffer capacity and flow rate according to age and have assumption on age and gender related changes on oral health.

MATERIAL AND METHODS:

Study design and participants:

This is a cross sectional study conducted on 250 adult healthy volunteers from which 120(48%) were male and 130(52%) were female. The study conducted on department of Biochemistry of Kabul University of Medical Sciences (KUMS) after approval by ethic committee of KUMS. All participants were informed about the research and signed informed consent form. All pregnant, smokers, those have systemic disease and who had radiography of neck and face in the last three months was excluded from study. All participants were categorized in two age groups 18-30 years old and 31-65 years. Normality test was considered before analyzing the data.

Salivarycollection:

Whole Unstimulated saliva was collected in sitting position between 8-10am. The participants were informed not to drink and eat at least two hours before saliva collection. Saliva was collected by drooling method in sterile tube up to 5ml. Saliva was immediately used for flow rate pH and buffer capacity determination.

Flow rate measurement:

Flow rate was calculated in milliliter /minute as total volume of saliva in milliliter was collected in a period of time (minutes).

pH and Buffer capacity measurement:

Digital pH meter (pHS-2F) used for measuring pH²². After measuring pH the Buffer capacity was measured by adding 1ml of saliva to 3ml of 0.005M hydrochloric acid in an open container and kept for 10minutes for CO₂ elimination then the pH was measured by pH meter. Buffer capacity was categorized to normal (pH=5-7) borderline (pH=4-5) and low buffer capacity (pH <4)²³.

Data analysis:

All data were analyzed in SPSS version 21 with CI=95% and α=0.05 concerned. After the determination of the distribution of data, Descriptive statistics were shown. All participants were divided in two age groups, 18-30 years (young adults) and 31-65 years old. The difference of variables in each age group with the concern of gender was investigated by Mann Whitney U test. Prevalence of low buffer capacity was investigated according to age and gender.

RESULTS”:

In this study 250 volunteers participated that 130(52%) of them were female and 120(48%) were male. Mean age was 30 years. Descriptive statistics of all data is shown in Table (1).

Table 1: Descriptive statistics of variables

		Age (years)	Flow rate (ml/min)	pH	Buffer capacity
N	Valid	250	250	250	250
N	Missing	0	0	0	0
Mean		29.8040	0.7607	6.4768	4.1349

Median		25.0000	0.6000	6.5000	4.0000

Std. Deviation		12.22109	0.48849	0.54757	.87664

Minimum		18.00	0.13	5.00	1.70

Maximum		65.00	3.00	8.00	6.50

As a result of Mann Whitney U test there is significant decrease (P=0.009) in flow rate and buffer capacity after age 30 but there is no significant differences (P=0.072) in pH of saliva among two age groups. Differences of variables according to gender in each group are shown in Table (2).

Table 2: Differences of variables in two age groups according to gender

Ranks
Age		Gender	N	Mean Rank	Sum of Ranks	Asymp.sig (two tailed)
18-30 years old	Flow rate ml/min	Female	95	75.77	7198.00	P=0.023^*
		Male	70	92.81	6497.00
		Total	165
	pH	Female	95	82.30	7818.50	P=0.825
		Male	70	83.95	5876.50
		Total	165
	Buffer capacity	Female	95	73.87	7018.00	P=0.004^*
		Male	70	95.39	6677.00
		Total	165
31-65 years old	Flow rate ml/min	Female	35	34.10	1193.50	P=0.005*
		Male	50	49.23	2461.50
		Total	85
	pH	Female	35	43.49	1522.00	P=0.878
		Male	50	42.66	2133.00
		Total	85
	Buffer capacity	Female	35	36.34	1272.00	P=0.036*
		Male	50	47.66	2383.00	P=0.036*
		Total	85

*p<0.05 and the differences is significant.

Among all participants 165(66%) were 18-30 years old while 85(34%) were between age 31-65 years old. result of Kolmogorov Simonov test (p=0.000) is shown that the data is not normally distributed.

Totally 88 participants (35.2%) have low buffer capacity (pH less than 4) and 64.8% has normal and borderline buffer capacity. It should be mention that 42.3% of females have low buffer capacity and 27.5% of males have low buffer capacity. Females have 1.5 times lower buffer capacity than males. Prevalence of lower buffer capacity in age 18-30 years is 0.85 times less than age group 31-65 years. Result of spearman test shows a weak positive correlation (r=0.13, p=0.036) between flow rate and buffer capacity in addition there is significant positive correlation (r=0.19, p=0.002) between buffer capacity and pH. But there is no significant correlation between pH and flow rate.

DISCUSSION:

Based on salivary flow rate, pH and buffering capacity studies risk of caries are more in females than male and oral hygiene is poor in males^20,24. In this study salivary flow rate is lower after age 30 years and decrease in flow rate is much more in females than males. Result of this study in regard to the flow rate of unstimulated saliva is the same as study of Rose-Anna Grace Pushpass et al²⁵ who reported low flow rate in older adult and research of Hiba Khaldoun Al-Tamimi and Nadia Aftan Al-Rawi ²⁶ that describes the lowering of flow rate with increasing age. Similarly, Ana Carolina Mosca et al ²⁷ observed a decrease of flow rate with aging which was higher in men than women. Dental caries is a multifactorial disease and low flow rate is one of the factors of dental caries²⁸as A. E. González-Aragón Pineda et al ²¹ reported that incidence of dental caries increases with low flow rate, therefore females are more prone for dental caries after age 30 years.

Salivary pH is one of the important characteristics of saliva that have important role on oral health. When salivary pH becomes lower than normal the acids in oral cavity breaks down the enamel, the longer this condition continues the more likely the development of caries ²⁹. Salivary buffer system is an important factor for salivary pH control and normal buffer capacity leads to good outcome of oral health ^{30, 31,32}. In this study salivary buffer capacity significantly decreased after age 30 and females have much lower buffer capacity than male that is similar to research conducted by Isamu Kado et al²⁴ who reported lower buffer effect in females than males. In addition.in this study positive weak correlation exists between salivary flow rate and buffer capacity which is similar to research reported by Katie P Wu et al ³³and study conducted by C. Fenoll Palomares et al³⁴. Besides these in study of Egija Zaura et al ³⁵and .V.T. Hemalatha³⁶ it was reported that salivary flow clears the acid and contributes to pH recovery .Study of Vijaya lakshmi Bolla et al ³⁷ reported that low salivary pH is a risk for higher incidence of caries and described a clearly association between salivary buffer capacity and dental caries. Therefore, females are more prone for dental caries due to low buffer capacity.It should be mentioned that in this study low sample size especially in age group 31-65 years old and unequally distribution of participants in each group are the limitations that should be considered in further researches.

CONCLUSION:

Finally it can be concluded that unstimulated salivary flow rate and buffer capacity is lower after age 30 and females have significant much lower flow rate and buffer capacity than males. So it can be assumed that females especially after age 30 are more prone to dental caries.

ACKNOWLEDGMENTS:

The authors are thankful from all volunteers for devoting their time in collecting samples.

CONFLICTS OF INTEREST:

The authors have no potential conflicts of interest to disclose.

REFERENCES:

1. L. Forcella, C. Filippi, T.Waltimo, A. Filippi. Measurement of unstimulated salivary flow rate in healthy children aged 6 to 15 years. Research and Science. 2018,128 (12):962-967.

2. Jyothi.S, Karthikeyan Murthykumar, Anupama Deepak, Dhanraj, Noorul Aneesa, Sowndarya Baskar et al. Drugs Inducing Xerostomia. Research J. Pharm. and Tech. 2016 May; 9(5). DOI: 10.5958/0974-360X.2016.00113.X.

3. Thirumalaisamy V, Gajendran P. Role of Salivary Interleukin 1 in Chronic Periodontitis: A Review. Research Journal of Pharmacy and Technology. 2018; 11(1): 390-2. http://dx.doi.org/10.5958/0974-360X.2018.00071.9

4. Indrawati R, Diyatri I, Rezkita F, Humaidy RS. Quantification molecular weight of human beta defensin-2 and human beta defensin-3 from saliva of caries patient.Research Journal of Pharmacy and Technology. 2020; 13(10): 4599-602. http://dx.doi.org/10.5958/0974-360X.2020.00810.0

5. Agarwal R, Lakshmi T. Salivary enzymes as biomarkers for periodontitis–an update. Research Journal of Pharmacy and Technology. 2014; 7(1): 98-100.

6. Nandakumar DG, Gheena S. Cortisol in Saliva of Uncontrolled Diabetes Patients and Adrenal Stress index. Research J. Pharm. and Tech. 2016; 9(10): 1788. http://dx.doi.org/10.5958/0974-360X.2016.00361.9

7. Ajmal Mir, Reyaz Farooq, Amir Purra and Fayaz Ahanger. Evaluation of impact of hypertension on salivary pH, salivary flow and buffering capacity: A retrospective analysis. International Journal of Advanced Research in Medicine. 2021; 3 (2): 411-414

8. P.L.F. Bonda, K. Brilli, F. Pattarino, A. Foglio-Bonda. Salivary flow rate and pH in patients with oral pathologies. European Review for Medical and Pharmacological Science. 2017; 21: 369-374.

9. Sareen A, Tandon S, Ramachandran A, Srimathi R. Saliva as A Diagnostic Tool for Detection of the Viruses: A Review. Research Journal of Pharmacy and Technology. 2018; 11(10): 4739-43. http://dx.doi.org/10.5958/0974-360X.2018.00863.6

10. A. Mohiti, F. Eslami, M.R Dehestani. Does Hypertension affect Saliva Properties?. J Dent Shiraz University Medical Science. 2020; 21(3): 190-194

11. A. A. Pratha, J. Prabakar. Comparing the effect of Carbonated and energy drinks on salivary pH- In Vivo Randomized Controlled Trial. Research J. Pharm. and Tech. 2019; 12(10). http://dx.doi.org/10.5958/0974-360X.2019.00809.6

12. Shetty SS, Wen LY, Dass W, Kaur G, Venugopal H, Hamzah MR, Leong SJ. Effects of green tea and black tea on salivary pH and flow rate in healthy individuals. Research Journal of Pharmacy and Technology. 2020; 13(2): 627-30. http://dx.doi.org/10.5958/0974-360X.2020.00119.5

13. F. Bechir, M. Pacurar, A. Tohati and S. Maria Bataga. Comparative Study of Salivary pH, Buffer Capacity, and Flow in Patients with and without Gastro esophageal Reflux Disease. Int. J. Environ. Res. Public Health. 2022; 19(1): 201.

14. Y.S.Wimardhani, Y.W.Kusuma, H. Sasanti, Anzany Tania DWI Putri Baringbin. G. P.Subita, A. S.Sarsito S.A Pradono, A. I. Soegyanto, F. Rahmayanti, I. Wardhany, A. T. DWI Putri Baringbing. Salivary profile of recovering drug users in Indonesia. Journal of International Dental and Medical Research. 2016; 9(1): 50-54.

15. F. S. AlHudaithi and D. Ali Alshammery. Screening of biochemical parameters in the orthodontic treatment with the fixed appliances: A follow-up study. Saudi Journal of Biological Sciences. 2021; 28(12): 6808–6814.

16. I. Taraboanta , Simona Stoleriu , Gianina Iovan, A. C. Ghiorghe, Irina Nica1e, Andra C. Taraboantan Gamen et al.Evaluation Of The Salivary Parameters In Facial Mask Wearers During Covid-19 Pandemic. Stoma Edu J. 2021; 8(4): 233-238.

17. A.Hosseini Yekani, A. Nadjarzadeh, M. Vossoughi, J. Zavar Reza, Ali Golkari. Relationship between Physicochemical Properties of Saliva and Dental Caries and Periodontal Status among Female Teachers Living in Central Iran. Journal of International Society of Preventive and Community Dentistry. 2018; 8(1): 48-55. DOI: 10.4103/jispcd.JISPCD_391_17

18. Majdy Idrees, Mohammad-Zakaria Nassani, Omar Kujan. Assessing the association between unstimulated whole salivary flow rate (UWSFR) and oral health status among healthy adult subjects: A cross-sectional study. Med Oral Patol Oral Cir Bucal. 2018; 23(4): 384-90.

19. A. A. Alkhateeb, Lloyd A. M. Richard B. Presland, Marilynn L. R. Donald L.Chi. Unstimulated Saliva-Related Caries Risk Factors in Individuals with Cystic Fibrosis: A Cross-Sectional Analysis of Unstimulated Salivary Flow, pH, and Buffering Capacity. Caries Res. 2017; 51: 1–6.

20. B. Dawlaty, S. Rahim. Study of unstimulated salivary flow rate, total protein, albumin concentration and estimated albumin globulin ratio changes in normal adults according to age and gender. Research J. Pharm. and Tech. 2022; 15(7). DOI: 10.52711/0974-360X.2022.00534

21. A. E. González-Aragón Pineda1, A. García Pérez1 and F. García-Godoy. Salivary parameters and oral health status.amongest adolescents in Mexico. BMC Oral Health. 2020; 20(1): 190

22. D. P. Antunes, R.M. M. Marinho, M.C. Valera Garakis, E. Bresciani. Buffer Capacity of Saliva as a Function of Time after Consumption of Sugary, Sugar-Free and Probiotic Chewing Gums. Brazilian Research in Pediatric Dentistry and Integrated Clinic. 2015; 5 (1): 153-161.

23. Rubbaey YA. Oral health status and caries experience among a group of asthmatic patients in Al-Najaf Al-Ashraf. Research Journal of Pharmacy and Technology. 2019; 12(9): 4099-101. http://dx.doi.org/10.5958/0974-360X.2019.00706.6

24. Isamu Kado, Ryo Kunimatsu, Yuki Yoshimi, Cynthia Concepcion Medina,Sakura Yamada1 and Kotaro Tanimoto. Surveillance of salivary properties of pre‑orthodontic patients in relation to age and sex. Scientific Reports. 2021; 11: 6555.

25. R.A. Grace Pushpass et al. Altered Salivary Flow, Protein Composition, and Rheology Following Taste and TRP Stimulation in Older Adults. Forniters in Physiology. 2019; 10: 652.Doi: 10.3389/fphys.2019.00652

26. H.Khaldoun Al-Tamimi and N. Aftan Al-Rawi.Age-Related Changes with Selected Salivary Physical Properties and Caries Experience among Healthy Adult Men. International Journal of Medical Research and Health Science. 2018; 7(9): 52-58.

27. Ana Carolina Mosca et al. How are macronutrient intake, BMI, ethnicity, age, and gender related to the composition of unstimulated saliva? A case study. Journal of Texture Studies. 2019; 50 (1): 53–61. DOI: 10.1111/jtxs.12362

28. A. Hadi Faha, Raed F. Al Huwaizi,Yamama A. A. Rubbaey and A. Falhuwaizi. Salivary pH, flow rate and Streptococcus mutants count in relation to oral health status among colored Eye Adolescent in Baghdad/Iraq. Biomedical and Pharmacology. 2018; 11(4).

29. Karishma S Halageri, Aruna C N, Padma K Bhat, Subodh Kumar. Comparison of salivary pH, flow rate and oral clearance rate between packaged fruit drink and fresh fruit juice in young adults: A comparative study. Journal of Advanced Medical and Dental Sciences Research. 2020; 8 (7).

30. M. R.Malekipour, M. Messripour and F. Shirani.Buffering Capacity of Saliva in Patients with Active Dental Caries. Asian Journal of Biochemistry. 2008; 3: 280-283.

31. F. A. Motamayel, P. Falsafi, M. T.Goodarzi, J.poorolaja. Comparsion of salivary pH, buffering capacity and alkaline phosphatase in smokers and healthy nonsmokers. Sultan Qaboos University Med J. 2016, 16(3): 317-321.

32. A.Almeslet, Alnamlah S, Alanzan L, Aldriwesh R, AlWehaiby S.Role of Salivary Biomarkers in Cystic Fibrosis: A Systematic Review. Biomed Res Int. 2022. DOI: 10.1155/2022/5818840. PMID: 35097122; PMCID: PMC8791744.

33. Katie P Wu , Jyh-Yuh Ke, Chia-Ying Chung, Chia-Ling Chen, Tsong-Long Hwang, Ming-Yen Chou, Alice M K Wong et al. Relationship between unstimulated salivary flow rate and saliva composition of healthy children in Taiwan.Chang Gung Med J. 2008; 31(3): 281-6.

34. C. Fenoll-Palomares, J.V. Muñoz Montagud, V Sanchiz, B Herreros, V. Hernández, M. Minguez, A Benages. Unstimulated salivary flow rate, pH and buffer capacity of saliva in healthy volunteers. Rev Esp Enferm Dig. 2004; 96(11) : 773-783.

35. Egija Zaura, Bernd W Brandt, Andrei Prodan, Maarten Joost Teixeira de Mattos,Sultan Imangaliyev, Jolanda Kool et al. On the ecosystem network of saliva in healthy young adults. The ISME Journal. 2017; 11(5): 1218–1231.

36. V.T. Hemalatha, Sivaranjini. P, Mani Sundar. N, .Julius. A, V. Aarthi Nisha, Sankar Narayanan R.A. Comparative Study Of Salivary Protein Concentration, Flow Rate, Buffer Capacity And pH Among Young And Elderly Subjects With Gingivitis And Periodontitis A Biochemical Study. Journal of Positive School Psychology. 2022; 6(3): 3756 –3766.

37. Vijaya Lakshmi Bolla, Surendra R. Munnangi, Manoj Kumar MG, U. Kumar Chowdary, Pradeep Koppulu and Lingam Amara Swapna. Correlation between the pH of saliva, plaque and buffering capacity of saliva. International Journal of Applied Dental Sciences. 2017; 3(4): 48-50.

Received on 17.09.2023 Modified on 02.11.2023

Research J. Pharm. and Tech 2024; 17(5):2213-2216.

DOI: 10.52711/0974-360X.2024.00348