Мочевая кислота в слюне как неинвазивный биомаркер у пациентов с артериальной гипертензией: перекрестное сравнительное исследование

Ахмед М.С., Альсарраф А.М.Ф., Швани М.А., Хамад Ш.А., Хавези Д., Ахмед О.Ш., Альраад А.Ш. Мочевая кислота в слюне как неинвазивный биомаркер у пациентов с артериальной гипертензией: перекрестное сравнительное исследование. Сибирский журнал клинической и экспериментальной медицины. 2026;41(3):141–147. https://doi. org/10.29001/2073-8552-2026-41-3-141-147

Hypertension (HTN) affects more than 1.28 billion adults worldwide and remains one of the leading causes of premature death from cardiovascular disease, stroke, and chronic kidney disease1. Despite advances in antihypertensive pharmacotherapy, a substantial proportion of patients remain undiagnosed or inadequately controlled, partly because standard blood pressure (BP) measurements require clinical infrastructure and are subject to the “white coat” effect [1]. In this context, the identification of reliable, non-invasive biological markers of hypertension and target organ damage is of considerable clinical importance.

Uric acid (UA) is the terminal product of purine catabolism in humans, generated from hypoxanthine and xanthine through the action of xanthine oxidase [2] In physiological conditions, UA functions as the predominant plasma antioxidant, accounting for approximately 70% of total plasma antioxidant activity. However, in conditions of excessive production or impaired renal clearance, UA assumes pro-inflammatory and pro-oxidant properties – promoting endothelial dysfunction, activating the renin-angiotensin-aldosterone system, and stimulating vascular smooth muscle proliferation – mechanisms directly relevant to the pathogenesis of hypertension [3, 4].

Saliva is an increasingly recognised diagnostic biofluid that reflects the systemic biochemical milieu while offering the advantage of non-invasive, painless, and simple collection [5]. Salivary UA is the principal salivary antioxidant, contributing 70–85% of total salivary antioxidant capacity (TSAC) [6]. A linear correlation between serum and salivary UA concentrations has been documented in several populations, suggesting that salivary UA may serve as a surrogate marker for systemic uric acid status [7].

Although prior reviews and cross-sectional studies have established associations between salivary UA and conditions such as gout, cancer, metabolic syndrome, and HIV infection, the literature on salivary UA in hypertension – particularly in adult populations – remains sparse [8]. Maciejczyk M., et al. [9] demonstrated in a paediatric hypertensive cohort that salivary UA in stimulated whole saliva (SWS) possessed an exceptionally high area under the receiver operating characteristic curve (AUC=0.98), with 94% sensitivity and 96% specificity for differentiating hypertensive from normotensive children. The Salurate study, a current prospective multicentre cohort, is evaluating weekly salivary UA self-monitoring as a predictor of hypertensive disorders of pregnancy [10]. However, an adult non-pregnant cohort has not been systematically studied.

The present study was therefore designed to: (i) compare salivary UA concentrations measured in both non-stimulated (NWS) and stimulated (SWS) whole saliva between adult hypertensive patients and normotensive controls; (ii) evaluate the correlation of salivary UA with blood pressure measurements and serum UA; and (iii) determine the diagnostic utility of salivary UA as a biomarker of hypertension using ROC analysis.

Material and Methods

This was a prospective, hospital-based case-control study conducted between April 2025 and March 2026. The study was approved by the Institutional Ethics Committee (Kurdistan Higher Council of Medical Specialties, No. 1784, date 12-3-2025) and conducted in accordance with the Declaration of Helsinki. Written informed consent was obtained from all participants prior to enrolment.

The sample size was calculated a priori using G*Power version 3.1 for a two-independent-group comparison (hypertensive vs. normotensive controls). The calculation was based on the expected difference in stimulated salivary uric acid levels derived from the pilot findings reported by Maciejczyk M. et al. [9], which demonstrated a large effect size between hypertensive and normotensive subjects. Assuming a two-tailed hypothesis, an effect size (Cohen’s d) of 0.80, an alpha error probability of 0.05, and a statistical power of 90% (β = 0.10), the minimum required sample size was estimated at 53 participants per group. To compensate for potential dropouts, incomplete saliva samples, or laboratory exclusions, the sample size was increased by approximately 10%, resulting in a final recruitment target of 60 participants in each group (total n = 120).

The hypertension group consisted of 60 adults (≥18 years) with established primary (essential) hypertension, defined according to the 2023 ESH/ESC guidelines as office systolic blood pressure (SBP) ≥ 140 mmHg and/or diastolic blood pressure (DBP) ≥ 90 mmHg on at least two occasions, or on stable antihypertensive monotherapy for ≥ 6 months.12 The control group comprised 60 age- and sex-matched normotensive volunteers recruited from hospital staff and community outreach, with no personal or family history of hypertension, with BP values below 130/85 mmHg on two independent readings.

Exclusion criteria for both groups included: (i) secondary hypertension (renovascular, endocrine, or drug-induced); (ii) known diabetes mellitus, chronic kidney disease (eGFR <60 mL/min/1.73 m²), or hepatic impairment; (iii) active oral infection, periodontal disease, xerostomia, or oral malignancy (as these conditions directly alter salivary composition); (iv) current use of medications known to affect UA levels (allopurinol, febuxostat, loop diuretics, aspirin > 325 mg/ day); (v) current or recent (within 4 weeks) use of antibiotics, vitamin supplements, or non-steroidal anti-inflammatory drugs; (vi) pregnancy; Smoking or tobacco chewing(vii) and (viii) inability to provide informed consent.

Office blood pressure was measured in the non-dominant arm using a validated oscillometric device (Omron M6 Comfort) following a minimum of 5 minutes of seated rest, with the arm and back supported. Three consecutive readings were taken at 2-minute intervals and the average of the second and third measurements was used for analysis. Ambulatory blood pressure monitoring (ABPM) was performed where clinically appropriate to confirm hypertension diagnosis using a Mobil-O-Graph 24-hour monitor.

All participants were instructed to fast for at least 8 hours, refrain from tooth brushing for 2 hours prior to sampling, and avoid caffeine, tobacco, and physical exertion for at least 30 minutes before collection. Saliva was collected between 8:00 and 10:00 a.m. to minimise circadian variation. Patients were instructed to abstain from antihypertensive medications on the morning of collection; however, for ethical reasons, patients on long-acting medications provided samples prior to their daily dose.

The collection protocol followed that described by Maciejczyk M., et al. [10] and Kamodyová N., et al. [12]. Briefly, participants rinsed their oral cavities twice with distilled water at room temperature. Saliva was collected in a sitting position with the head slightly inclined forward, minimising facial and lip movements. The first minute of secretion was discarded. Non-stimulated whole saliva (NWS) was collected by passive drooling for 15 minutes into a sterile, pre-cooled Falcon-type tube placed in ice. Stimulated whole saliva (SWS) was subsequently collected for 5 minutes by applying 10 µl of 2% (w/w) citric acid to the tip of the tongue every 30 seconds to induce salivation. The volume collected was measured immediately using a calibrated pipette. Salivary flow rates (mL/min) were calculated by dividing the volume collected by collection time. Hyposalivation was defined as NWS flow < 0.2 mL/min and SWS flow < 0.9 mL/min.

Immediately after collection, samples were centrifuged at 3,000 × g for 20 minutes at 4°C. To prevent oxidative degradation, 10 µl of 0.5 M butylated hydroxytoluene (BHT) per 1 mL saliva was added to the supernatant. Aliquots were stored at -80°C and analysed within 3 months of collection. Samples with visible blood contamination were excluded.

Uric acid concentrationin saliva and serum was determined spectrophotometrically using a commercial UASure II Uric Acid Test Kit (Apex Biotechnology Corp. Hsinchu, Taiwan). The assay is based on the formation of a coloured complex of uric acid with iron ions and 2,4,6-tripyridyl-s-triazine (TPTZ), measured at 490 nm. All measurements were performed in duplicate and absorbance was recorded with an Infinite M200 PRO Multimode Microplate Reader. Results were expressed as µg of uric acid per mg of total protein for salivary measurements, and as µmol/L for serum measurements.

Total protein content in saliva was determined using the bicinchoninic acid (BCA) method with a commercial kit (Thermo Scientific Pierce BCA Protein Assay), using bovine serum albumin (BSA) as the standard. Total antioxidant capacity (TAC) was determined by the spectrophotometric ABTS method, measuring changes in absorbance of the ABTS radical cation (ABTS•+) at 660 nm, with results calculated from a Trolox standard curve and expressed in µmol/mg protein [13].

Serum uric acid was measured using a standard enzymatic colorimetric method on a Cobas 8000 automated analyser (Roche Diagnostics). Standard haematological and biochemical panels (creatinine, fasting glucose, lipid profile, full blood count) were performed in the hospital laboratory on the same day of saliva collection.

All participants underwent a standardised dental examination by an experienced dentist to exclude those with poor oral hygiene or active periodontal disease. The Approximal Plaque Index (API), Gingival Index (GI), and Sulcus Bleeding Index (SBI) were recorded. Participants with API >20%, GI >0.5, or SBI >0.5 were excluded, as periodontal disease and dental caries are recognised confounders for salivary oxidative stress markers [14].

Data were tested for normality using the Shapiro-Wilk test. Between-group comparisons were made using Student's independent t-test for normally distributed variables. Categorical variables were compared using the chi-squared test. The Pearson correlation coefficient (r) was used to assess relationships between salivary UA and clinical parameters. To evaluate the ability of the identified predictors to distinguish hypertension, ROC analysis was performed, including calculation of the area under the curve (AUC), optimal cut-off values, sensitivity, and specificity using the Youden index.A p-value of <0.05 was considered statistically significant. All analyses were performed using SPSS v28.0 (IBM, USA) and GraphPad Prism 9 (GraphPad Software, CA, USA).

Results

A total of 120 participants were enrolled: 60 hypertensive patients (32 males, 28 females; mean age 48.3 ± 9.2 years) and 60 normotensive controls (30 males, 30 females; mean age 47.6 ± 10.1 years). The two groups were well matched for age and sex distribution ( p > 0.05 for both). Hypertensive patients had significantly higher systolic BP (146.3 ± 11.4 vs 118.5 ± 8.2 mmHg; p < 0.001), diastolic BP (91.2 ± 8.7 vs 76.4 ± 6.5 mmHg; p < 0.001), while serum creatinine, fasting glucose, and total cholesterol were compared to controls (Table 1). Forty-two of the 60 hypertensive patients (70%) were on antihypertensive therapy (ACE inhibitors, angiotensin receptor blockers, or calcium channel blockers). No participant exceeded one antihypertensive agent. Dental examination confirmed no significant differences in oral hygiene or gingival status between the groups.

Table 1 . Clinical characteristics of hypertensive patients and normotensive controls

Таблица 1 . Клиническая характеристика пациентов с артериальной гипертензией и нормотензивной контрольной группы

Variable	Hypertensive Group ( n = 60)	Control Group ( n = 60)	p
Age (years), mean±SD	48.3±9.2	47.6±10.1	0.692
Sex (Male/Female)	32/28	30/30	0.712
BMI (kg/m²), mean±SD	26.4±3.5	25.2±3.1	0.083
SBP (mmHg), mean±SD	146.3±11.4	118.5±8.2	<0.001
DBP (mmHg), mean±SD	91.2±8.7	76.4±6.5	<0.001
Duration of hypertension (years)	6.8±4.3	N/A
Antihypertensive medication n (%)	42 (70%)	0 (0%)
Serum creatinine (mg/dL)	0.85±0.18	0.82±0.15	0.342
Fasting glucose (mg/dL)	92.4±10.2	89.7±9.8	0.152
Total cholesterol (mg/dL)	178±32	165±28	0.062

Note: SBP – systolic blood pressure, DBP – diastolic blood pressure, BMI – body mass index, SD – standard deviation, N/A – not applicable.

Salivary UA concentrations in both NWS and SWS were significantly elevated in the hypertensive group compared to controls. In stimulated saliva, UA was 18.62 ± 3.27 µg/ mg protein in hypertensive patients vs. 6.14 ± 1.92 µg/ mg protein in controls ( p < 0.001). Similarly, NWS UA was 5.84 ± 1.43 vs. 2.21 ± 0.78 µg/mg protein ( p < 0.001). Serum UA was also significantly higher in the hypertensive group (401.7 ± 68.4 vs. 298.3 ± 52.1 µmol/L; p < 0.001). Total antioxidant capacity in both NWS and SWS was significantly higher in hypertensive patients, consistent with the compensatory antioxidant response to increased reactive oxygen species (ROS) production. Stimulated salivary flow was significantly lower in the hypertensive group (0.87 ± 0.11 vs. 1.76 ± 0.14 mL/min; p < 0.001), with 28 hypertensive patients (47%) meeting the criterion for hyposalivation in SWS. These results are summarised in Table 2.

In the hypertensive group, SWS UA correlated strongly and positively with SBP ( r = 0.79, p < 0.001), DBP ( r = 0.81, p < 0.001), and serum UA ( r = 0.84, p < 0.001). NWS UA

Table 2 . Salivary and serum uric acid and antioxidant parameters in hypertensive patients and controls

Таблица 2 . Содержание мочевой кислоты и антиоксидантные параметры в слюне и сыворотке крови у пациентов с артериальной гипертензией и контрольной группы

Parameter	Hypertensive Group ( n = 60)	Control Group ( n = 60)
Salivary UA – NWS (µg/mg protein)	5.84±1.43	2.21±0.78*
Salivary UA – SWS (µg/mg protein)	18.62±3.27	6.14±1.92*
Serum UA (µmol/L)	401.7±68.4	298.3±52.1*
TAC – NWS (µmol/mg protein)	1.84±0.41	0.91±0.28*
TAC – SWS (µmol/mg protein)	2.13±0.58	0.98±0.31*
Salivary flow – NWS (mL/min)	0.41±0.06	0.48±0.05*
Salivary flow – SWS (mL/min)	0.87±0.11*	1.76±0.14

Note: NWS – non-stimulated whole saliva, SWS – stimulated whole saliva, UA – uric acid, TAC – total antioxidant capacity. *p<0.001 vs. hypertensive group.

also correlated significantly with SBP ( r = 0.61, p < 0.001) and DBP ( r = 0.58, p < 0.001). Moderate correlations were identified between SWS UA and BMI ( r = 0.44, p = 0.003) and between SWS UA and serum creatinine ( r = 0.39, p = 0.008). In the control group, no significant correlations were observed between salivary UA and blood pressure. All statistically significant correlations are presented in Table 3.

Table 3 . Pearson correlations between salivary uric acid and clinical parameters in the hypertensive group

Таблица 3 . Корреляции Пирсона между содержанием мочевой кислоты в слюне и клиническими параметрами в группе пациентов с артериальной гипертензией

Pair of Variables	r	p-value
SWS UA and SBP	0.79	<0.001
SWS UA and DBP	0.81	<0.001
SWS UA and Serum UA	0.84	<0.001
NWS UA and SBP	0.61	<0.001
NWS UA and DBP	0.58	<0.001
SWS UA and BMI	0.44	0.003
SWS UA and serum creatinine	0.39	0.008

Note: SWS – stimulated whole saliva, NWS – non-stimulated whole saliva, UA – uric acid, SBP – systolic blood pressure, DBP – diastolic blood pressure, BMI – body mass index.

ROC analysis demonstrated that SWS UA had the highest diagnostic performance among the parameters tested, with an AUC of 0.97 (95% CI: 0.93–1.00), sensitivity of 92%, and specificity of 93% at an optimal cut-off value of > 9.8 µg/mg protein. NWS UA also performed well (AUC = 0.88, sensitivity 78%, specificity 81%). Serum UA, while significantly elevated, showed lower discriminatory performance (AUC = 0.79). These findings are presented in Table 4.

Discussion

To our knowledge, this is among the first prospective case-control studies to rigorously evaluate salivary UA as a non-invasive biomarker of essential hypertension in an adult population, incorporating both NWS and SWS measurements and a full redox antioxidant profile alongside a standardised saliva collection protocol. The central finding – that salivary UA, particularly in stimulated saliva, is markedly elevated in hypertensive adults and strongly correlated with blood pressure – builds upon and extends the findings of Maciejczyk, et al. [10] in hypertensive children, and is consistent with the emerging evidence implicating UA in hypertensive pathophysiology.

The significantly elevated salivary UA in the hypertensive group can be understood through the lens of xanthine oxidase (XO) biology. In hypertension, oxidative stress drives upregulation of XO activity, the enzyme responsible for the terminal steps of purine catabolism [3]. XO simultaneously generates UA and superoxide radicals (O2•-), thereby creating a dual biological burden: elevated UA on the one hand, and increased ROS on the other. This was reflected in our data by the significantly elevated total antioxidant capacity (TAC) in both NWS and SWS of hypertensive patients – a finding that appears counterintuitive but represents an adaptive response of the antioxidant barrier to the increased oxidant load, as also demonstrated by Maciejczyk M. et al [9] UA itself, as the predominant salivary antioxidant (contributing 70–85% of TSAC7), would be expected to increase proportionally with this adaptive upregulation.

The paradoxical pro-oxidant role of UA at high concentrations is now well recognised. When UA reacts with peroxynitrite – formed during the combination of superoxide and nitric oxide (NO) – it generates highly reactive amino carbonyl radicals capable of alkylating biomolecules, a process particularly prominent during oxidative/nitrosative stress conditions [15]. This mechanism explains why elevated UA, despite being the body's primary liquid-phase antioxidant, is also consistently associated with hypertension, cardiovascular disease, and metabolic syndrome [3, 4]. In our correlational analysis, the strong positive correlation between SWS UA and both SBP ( r = 0.79) and DBP ( r = 0.81) further supports this mechanistic framework, as it implies that the higher the blood pressure – and by extension the greater the systemic oxidative/nitrosative stress – the greater the salivary UA elevation.

The linear relationship between salivary and serum UA ( r = 0.84 in SWS) observed in our study is consistent with prior literature. Shibasaki K., et al. [16] demonstrated a statistically significant linear correlation between serum and salivary UA in 244 subjects across different UA status groups, and proposed salivary UA as a substitute for blood UA measurement. This was further corroborated by Zhao J.

Table 4 . ROC analysis of salivary and serum uric acid as biomarkers for hypertension

Таблица 4 . ROC-анализ слюнной и сывороточной мочевой кислоты в качестве биомаркеров артериальной гипертензии

Parameter	AUC	95% CI	Cut-off	Sensitivity (%)	Specificity (%)
SWS UA (µg/mg protein)	0.97	0.99-1.00	>9.8	92	93
NWS UA (µg/mg protein)	0.88	0.95-1.00	>3.6	78	81
Serum UA (µmol/L)	0.79	0.83-0.94	>352	72	74
TAC SWS (µmol/mg protein)	0.83	0.92-0.99	>1.52	79	80

Note: AUC – area under the ROC (Receiver Operating Characteristic) curve, NWS – non-stimulated whole saliva, SWS – stimulated whole saliva, UA – uric acid, TAC – total antioxidant capacity.

and Huang Y. [17] who showed that salivary UA tracks serum UA changes during urate-lowering therapy in patients with chronic gouty arthropathy, with a consistent ratio between compartments. Our findings support these observations and extend them to the hypertensive context in adults.

The ROC performance of SWS UA in our study (AUC = 0.97, sensitivity 92%, specificity 93%) is remarkably similar to the paediatric data reported by Maciejczyk M., et al. [9] (AUC = 0.98, sensitivity 94%, specificity 96%), suggesting that the discriminatory power of salivary UA for hypertension may be robust across the age spectrum. Importantly, SWS UA outperformed both NWS UA and serum UA in terms of diagnostic accuracy. This is likely because stimulated saliva, predominantly produced by the parotid glands, is secreted in higher volumes and is less susceptible to the dilutional and contamination effects that can affect NWS. Stimulation also promotes a more reproducible sample with higher protein concentrations, facilitating normalisation of UA values. Clinically, SWS UA emerges as the preferred candidate for a point-of-care saliva test.

The finding of reduced stimulated salivary flow in the hypertensive group is clinically relevant. Hyposalivation in hypertension may result from the reduced bioavailability of nitric oxide (NO), which plays a critical role in the parasympathetic regulation of salivary gland secretion [18]. Reduced NO bioavailability – a hallmark of hypertensive endothelial dysfunction – leads to impaired opening of aquaporin water channels in salivary secretory cells, resulting in decreased saliva production. This has direct clinical implications: patients with hypertension may be at increased risk of xerostomia, dental caries, and oral mucosal infections, reinforcing the importance of dedicated dental care in this population [9, 14].

From a translational perspective, our findings are highly relevant in the context of emerging salivary UA monitoring technologies. The Salurate system described by Chmielewska B., et al. [10] – a non-invasive, home-use colorimetric test with a smartphone application – represents a practical vehicle for delivering such a test in the community setting. While Salurate is currently being validated specifically for hypertensive disorders of pregnancy, the underlying platform could, in principle, be adapted for chronic hypertension monitoring in the general adult population. The simplicity of saliva collection, freedom from needle phobia and venepuncture risks, and suitability for home-based monitoring represent meaningful advantages over serum testing, particularly for elderly patients, those with needle phobia, or populations in resource-limited settings [8, 19].

There are several limitations to acknowledge. First, as a single-centre study, our sample may not fully represent the heterogeneity of hypertensive presentations. Second, the cross-sectional nature of the comparison limits causal inference; longitudinal data would be required to determine whether salivary UA rises before, concurrently with, or after the onset of elevated blood pressure. Third, although we excluded patients on allopurinol and other UA-modulating medications, it is possible that antihypertensive drugs – particularly ACE inhibitors and angiotensin receptor blockers – influence salivary composition indirectly via haemodynamic effects on salivary gland perfusion. Fourth, dietary purine intake was not controlled, which is a recognised confounder of UA levels. Future studies should incorporate dietary assessment tools and measure salivary UA serially over time to account for intra-individual variability. Fifth, we did not assess secondary oxidative stress markers such as malondialdehyde (MDA), advanced glycation end-products (AGE), or peroxynitrite, which would have provided a more complete characterisation of the salivary redox microenvironment.

Conclusion

This study demonstrates that salivary uric acid, measured in stimulated whole saliva, is significantly elevated in adults with essential hypertension compared to normotensive controls and exhibits strong positive correlations with systolic and diastolic blood pressure. The diagnostic performance of SWS UA (AUC = 0.97, sensitivity 92%, specificity 93%) suggests it may serve as a reliable, non-invasive new biomarker for identifying hypertension. The linear relationship between salivary and serum UA further validates saliva as a surrogate biological matrix for UA assessment. Reduced stimulated salivary flow in hypertensive patients highlights the need for heightened dental surveillance in this population. Larger multicentre prospective studies with longitudinal follow-up are warranted to validate these findings, establish reference intervals for salivary UA in hypertensive adults, and explore the clinical utility of home-based salivary UA selfmonitoring systems in chronic hypertension management.