Validity and reliability of test strips for the measurement of salivary nitrite concentration with and without the use of mouthwash in healthy adults.

The nitrate (NO)-nitrite (NO)-nitric oxide (NO) pathway has received considerable interest in recent years as a potential target for nutritional interventions designed to increase NO production, and elicit therapeutic effects in humans. In particular, studies have evaluated the effects of supplemental dietary NO, which serves as a 'substrate' for this pathway, on numerous different health outcomes. One challenge has been to evaluate compliance with the NO interventions. A recent advance in this field has been the development of a non-invasive, simple and rapid method to measure nitrite concentrations in saliva using small test salivary strips. In the present study, ten healthy adults were recruited to a randomised, crossover study and received an acute dose of NO-rich beetroot juice (BJ) after rinsing their mouth with either water or commercially available antibacterial mouthwash. Salivary NO and NO concentrations were measured at baseline and up to 5 h after BJ consumption using the gold-standard chemiluminescence and a colorimetric Griess assay. In addition, two salivary test strips (Berkeley Test strips, CA, USA) were used to measure NOconcentrations at the same time points. Five observers read the strips and inter- and intra-observer reliability was measured. The Bland-Altman method was used to provide a visual representation of the agreement between the methods used to evaluate salivary NO/NOconcentration. Sialin concentrations were measured at baseline and up to 5 h after BJ consumption. BJ elevated salivary NO and NO concentrations when the mouth was rinsed with water (both P < 0.01), as assessed via both chemiluminescence and Griess methods. Rinsing the mouth with antibacterial mouthwash attenuated markedly the increase in NO (P < 0.001), while NOconcentrations were unaffected (P > 0.05). The Intra-Class Coefficients of Correlation (ICC) showed a high inter- and intra-observer reliability (r > 0.8). A significant positive correlation was found between absolute salivary NO concentrations measured by strips and Griess and chemiluminescence methods (rho = 0.83 and 0.77, respectively) and also when expressed as changes in salivary NO concentrations (rho = 0.80 and 0.79, respectively). Bland Altman analysis indicated a poor agreement for absolute NO concentrations between salivary strips and the chemiluminescence and Griess methods. A small significant negative correlation was found between changes in salivary sialin and salivary NO- concentrations (r = -0.20, P = 0.04). A non-significant positive correlation was observed between the change in salivary sialin and salivary NO- concentrations (r = 0.18, P = 0.06). This study suggests that commercially available salivary NO test strips provide a reasonable surrogate marker for monitoring changes in salivary NO concentrations in humans. However, the strips do not provide accurate estimates of absolute NO concentrations.