Salivary Nitrite Research Articles

The effects of isoflavone supplementation plus combined exercise on salivary markers of oxidative stress in postmenopausal women.

This study tested the effect of isoflavone supplementation in addition to combined exercise training in salivary oxidative stress markers in non-obese postmenopausal women. Thirty-two postmenopausal women without hormone therapy were randomly assigned to exercise + placebo (n = 15) or exercise + isoflavone supplementation (n = 17) groups. They performed 30 sessions of combined exercises (aerobic plus resistance) over ten weeks and consumed 100 mg of isoflavone supplementation or placebo. Saliva samples were collected after an overnight fast. Superoxide dismutase, total antioxidant capacity, thiobarbituric acid reactive substances, catalase, total protein and nitrite were determined before and after ten weeks of the intervention. The ANOVA two-way analysis of variance was applied with α of 5%. Both groups increase (p<0.05) superoxide dismutase activity and decrease catalase levels. There was interaction (time × group) in both nitrite and thiobarbituric acid reactive substances results, with increase (p<0.05) in placebo group and decrease (p<0.05) in isoflavone group. No difference was found for total antioxidant capacity or total protein. The combination of isoflavone supplementation and exercise training can promote an antioxidant effect through reduction of lipid peroxidation and concentrations of salivary nitrite.

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 (NO3−)-nitrite (NO2−)-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 NO3−, which serves as a ‘substrate’ for this pathway, on numerous different health outcomes. One challenge has been to evaluate compliance with the NO3− 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 NO3−-rich beetroot juice (BJ) after rinsing their mouth with either water or commercially available antibacterial mouthwash. Salivary NO3− and NO2− 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 NO2−concentrations 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 NO3−/NO2−concentration. Sialin concentrations were measured at baseline and up to 5 h after BJ consumption.BJ elevated salivary NO3− and NO2− 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 NO2− (P < 0.001), while NO3−concentrations 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 NO2− concentrations measured by strips and Griess and chemiluminescence methods (rho = 0.83 and 0.77, respectively) and also when expressed as changes in salivary NO2− concentrations (rho = 0.80 and 0.79, respectively). Bland Altman analysis indicated a poor agreement for absolute NO2− concentrations between salivary strips and the chemiluminescence and Griess methods. A small significant negative correlation was found between changes in salivary sialin and salivary NO2ˉ concentrations (r = −0.20, P = 0.04). A non-significant positive correlation was observed between the change in salivary sialin and salivary NO3ˉ concentrations (r = 0.18, P = 0.06).This study suggests that commercially available salivary NO2− test strips provide a reasonable surrogate marker for monitoring changes in salivary NO2− concentrations in humans. However, the strips do not provide accurate estimates of absolute NO2− concentrations.

Dietary Nitrite Drives Disease Outcomes in Oral Polymicrobial Infections

Streptococcus mutans resides in the oral polymicrobial biofilm and is a major contributor to the development of dental caries. Interestingly, high salivary nitrite concentrations have been associated with a decreased prevalence of dental caries. Moreover, the combination of hydrogen peroxide–producing oral commensal streptococci and nitrite has been shown to mediate the generation of reactive nitrogen species, which have antimicrobial activity. The goal of this study was to examine whether nitrite affects S. mutans virulence during polymicrobial infections with the commensal Streptococcus parasanguinis. Here, we report that the combination of S. parasanguinis and nitrite inhibited S. mutans growth and biofilm formation in vitro. Glucan production, which is critical for S. mutans biofilm formation, was also inhibited in 2-species biofilms with S. parasanguinis containing nitrite as compared with biofilms that contained no nitrite. In the in vivo caries model, enamel and dentin carious lesions were significantly reduced in rats that were colonized with S. parasanguinis prior to infection with S. mutans and received nitrite in the drinking water, as compared with animals that had a single S. mutans infection or were co-colonized with both bacteria and received no nitrite. Last, we report that S. mutans LiaS, a sensor kinase of the LiaFSR 3-component system, mediates resistance to nitrosative stress. In summary, our data demonstrate that commensal streptococci and nitrite provide protection against S. mutans pathogenesis. Modulating nitrite concentrations in the oral cavity could be a useful strategy to combat the prevalence of dental caries.

Procyanidins in rice cooked with adzuki bean and their contribution to the reduction of nitrite to nitric oxide (•NO) in artificial gastric juice

In Japan, adzuki bean is cooked with rice. During the cooking, the colour of rice becomes pale red. It is postulated that the red pigment is produced from procyanidins and that the ingestion of red rice causes the production of nitric oxide (•NO) in the stomach by reacting with salivary nitrite. The increase in colour intensity accompanied the decrease in the amounts of procyanidins, suggesting the conversion of procyanidins into the red pigment during the cooking. In addition, the red pigment combined with rice strongly. The red-coloured rice produced •NO by reacting with nitrite in artificial gastric juice, and the amounts were dependent on the contents of procyanidins and the equivalents. It is suggested that although adzuki procyanidins were oxidised during cooking with rice, procyanidins and the equivalents bound to rice still have the ability to produce bioactive •NO in the stomach using nitrite in mixed whole saliva.

The oral nitrate-reducing capacity correlates with peak power output and peak oxygen uptake in healthy humans

Interest in inorganic nitrate and nitrite has grown substantially over the past decade as research has revealed the role of these anions in enhancing nitric oxide (NO) availability through an oral pathway. Nitrite synthesis in the mouth seems to be an important mechanism to feed the circulatory system with this anion. This is interesting since greater plasma nitrite concentration has been associated with better fitness levels in humans, but this question has not been investigated in relation to salivary nitrite concentration. Additionally, no previous study has investigated the oral nitrate-reducing capacity in regards to peak oxygen uptake (VO2peak) or peak power output (Wpeak) in humans. Thus, the main goal of this study was to investigate whether salivary nitrite and nitrate concentration and the oral nitrate-reducing capacity were associated with VO2peak and Wpeak in healthy humans.Fifty individuals (22 females and 28 males; 38.8 ± 14.3 years/old; BMI = 22.8 ± 3.9) performed a graded exercise test on a cycle ergometer to assess their VO2peak and Wpeak. Unstimulated salivary samples were taken before and 20 min after exercise to measure nitrate/nitrite, pH and lactate. The oral nitrate-reducing capacity was also assessed in 25 subjects before and after exercise.Oral nitrate-reducing capacity was positively associated with Wpeak (rs = 0.64; P = 0.001) and the VO2peak (rs = 0.54; P = 0.005). Similar correlations were found when these variables were analysed after exercise. In addition, a significant decrease in salivary pH (pre: 7.28 ± 0.361; post-exercise: 7.16 ± 0.33; P = 0.003) accompanied by an increase of salivary lactate (pre: 0.17 ± 0.14 mmol/L; post-exercise: 0.48 ± 0.38; P < 0.001) was found after exercise. However, these changes did not have any impact on salivary nitrate/nitrite concentration and the oral nitrate-reducing capacity after exercise.In conclusion, this is the first evidence showing a link between the oral nitrate-reducing capacity and markers of aerobic fitness levels in healthy humans.

The Acute Effects of Mat Pilates on Hemodynamic and Salivary Nitrite Responses After Exercise in Postmenopausal Women.

The aim of this study was to compare the hemodynamic and salivary responses after mat Pilates, aerobics, resistance exercises, and control. A total of 16 normotensive postmenopausal women performed: Pilates, 10 floor exercises; aerobics, 35 min on a treadmill (60-70% of heart rate reserve); resistance exercises, 60% of one-repetition maximum; and control, no physical exercise. Blood pressure and heart rate variability were evaluated at rest and 60min after the intervention. Saliva samples were collected at rest, immediately after, and 30 and 60min after exercise for analysis of nitrite concentration and total proteins. Systolic blood pressure, diastolic blood pressure, and mean blood pressure area under the curve were lower (p < .05) after both aerobic and resistance exercises sessions but not after the Pilates session when compared with the control session. Nitrite concentrations in saliva were higher 60min after the end of all exercise sessions. Heart rate variability was higher after the resistance exercise. Aerobic and resistance exercises were capable of decreasing arterial blood pressure after acute exercise.

Variability in nitrate-reducing oral bacteria and nitric oxide metabolites in biological fluids following dietary nitrate administration: An assessment of the critical difference

There is conflicting evidence on whether dietary nitrate supplementation can improve exercise performance. This may arise from the complex nature of nitric oxide (NO) metabolism which causes substantial inter-individual variability, within-person biological variation (CVB), and analytical imprecision (CVA) in experimental endpoints. However, no study has quantified the CVA and CVB of NO metabolites or the factors that influence their production. These data are important to calculate the critical difference (CD), defined as the smallest difference between sequential measurements required to signify a true change. The main aim of the study was to evaluate the CVB, CVA, and CD for markers of NO availability (nitrate and nitrite) in plasma and saliva before and after the ingestion of nitrate-rich beetroot juice (BR). We also assessed the CVB of nitrate-reducing bacteria from the dorsal surface of the tongue. It was hypothesised that there would be substantial CVB in markers of NO availability and the abundance of nitrate-reducing bacteria. Ten healthy male participants (age 25 ± 5 years) completed three identical trials at least 6 days apart. Blood and saliva were collected before and after (2, 2.5 and 3 h) ingestion of 140 ml of BR (∼12.4 mmol nitrate) and analysed for [nitrate] and [nitrite]. The tongue was scraped and the abundance of nitrate-reducing bacterial species were analysed using 16S rRNA next generation sequencing. There was substantial CVB for baseline concentrations of plasma (nitrate 11.9%, nitrite 9.0%) and salivary (nitrate 15.3%, nitrite 32.5%) NO markers. Following BR ingestion, the CVB for nitrate (plasma 3.8%, saliva 12.0%) and salivary nitrite (24.5%) were lower than baseline, but higher for plasma nitrite (18.6%). The CD thresholds that need to be exceeded to ensure a meaningful change from baseline are 25, 19, 37, and 87% for plasma nitrate, plasma nitrite, salivary nitrate, and salivary nitrite, respectively. The CVB for selected nitrate-reducing bacteria detected were: Prevotella melaninogenica (37%), Veillonella dispar (35%), Haemophilus parainfluenzae (79%), Neisseria subflava (70%), Veillonella parvula (43%), Rothia mucilaginosa (60%), and Rothia dentocariosa (132%). There is profound CVB in the abundance of nitrate-reducing bacteria on the tongue and the concentration of NO markers in human saliva and plasma. Where these parameters are of interest following experimental intervention, the CD values presented in this study will allow researchers to interpret the meaningfulness of the magnitude of the change from baseline.

Lipid and Protein Oxidation during in Vitro Gastrointestinal Digestion of Pork under Helicobacter pylori Gastritis Conditions.

Helicobacter pylori gastritis affects gastric pH and concentrations of ascorbic acid, hydrogen peroxide, hypochlorite, ammonia and urea, pepsin, and mucin. First, the separate effects of each of these altered factors on oxidation of pork were investigated during in vitro gastrointestinal digestion. Lipid and protein oxidation increased (range 23-48%) in duodenal digests of pork previously exposed to elevated (6.1) versus normal acidic stomach pH (2.3 to 3.5) conditions. Salivary nitrite reduced the formation of lipid and protein oxidation products (range 14-20%) under normal acidic but not elevated stomach pH conditions. Higher amounts of hydrogen peroxide and lower amounts of ascorbic acid decreased concentrations of lipid oxidation products in duodenal pork digests, whereas ammonia slightly stimulated protein oxidation during digestion. Second, two H. pylori gastritis-duodenal digestion models were installed using a set of altered compound concentrations at normal acidic or elevated stomach pH. The elevated pH-gastritis-duodenal digestion model increased pork protein oxidation compared with the normal pH-gastritis and the normal digestion model (14.3 ± 2.1 vs 8.2 ± 1.0 nmol DNPH/mg protein, P < 0.001). Compared with the other models, protein oxidation was also increased when nitrite-cured pork was exposed to the elevated pH-gastritis-duodenal digestion model (10.8 ± 1.4 vs 5.9 ± 0.8 nmol DNPH/mg protein, P < 0.001), but no significant effect of the model was observed when the pork was seasoned with herbs. Lipid oxidation was not or was marginally affected by the installed model.

Effects of Oral Supplementation With Nitrate-Rich Beetroot Juice in Patients With Pulmonary Arterial Hypertension—Results From BEET-PAH, an Exploratory Randomized, Double-Blind, Placebo-Controlled, Crossover Study

BackgroundThe nitrate–nitrite–nitric oxide (NO) pathway may represent a potential therapeutic target in patients with pulmonary arterial hypertension (PAH). We explored the effects of dietary nitrate supplementation, with the use of nitrate-rich beetroot juice (BRJ), in patients with PAH. Methods and ResultsWe prospectively studied 15 patients with PAH in an exploratory randomized, double-blind, placebo-controlled, crossover trial. The patients received nitrate-rich beetroot juice (∼16 mmol nitrate per day) and placebo in 2 treatment periods of 7 days each. The assessments included; exhaled NO and NO flow–independent parameters (alveolar NO and bronchial NO flux), plasma and salivary nitrate and nitrite, biomarkers and metabolites of the NO-system, N-terminal pro–B-type natriuretic peptide, echocardiography, ergospirometry, diffusing capacity of the lung for carbon monoxide, and the 6-minute walk test. Compared with placebo ingestion of BRJ resulted in increases in; fractional exhaled NO at all flow-rates, alveolar NO concentrations and bronchial NO flux, and plasma and salivary levels of nitrate and nitrite. Plasma ornithine levels decreased and indices of relative arginine availability increased after BRJ compared to placebo. A decrease in breathing frequency was observed during ergospirometry after BRJ. A tendency for an improvement in right ventricular function was observed after ingestion of BRJ. In addition a tendency for an increase in the peak power output to peak oxygen consumption ratio (W peak/VO2 peak) was observed, which became significant in patients reaching an increase of plasma nitrite >30% (responders). ConclusionsBRJ administered for 1 week increases pulmonary NO production and the relative arginine bioavailability in patients with PAH, compared with placebo. An increase in the W peak/VO2 peak ratio was observed after BRJ ingestion in plasma nitrite responders. These findings indicate that supplementation with inorganic nitrate increase NO synthase–independent NO production from the nitrate-nitrite-NO pathway.

Salivary cortisol and nitrite concentrations in school teachers: A longitudinal pilot study.

Occupational/mental stressors, which may be evaluated with measurements of salivary cortisol concentrations, affect the endothelial function and has implications on cardiovascular health. Nitric oxide (NO) is known to have an important role in cardiac function and may also be assessed in the saliva, but its participation in physiological responses to hypothalamic-pituitary-adrenal (HPA) axis stimulation is still not completely understood. The aim of the present study was to test the hypothesis that salivary NO (as nitrite) and cortisol concentrations in school teachers vary throughout the academic year. Saliva samples were collected from 13 teachers distributed across five secondary schools. The samples were collected at 6:30 a.m., 11:30 a.m., and 5:30 p.m. in the months of March, July, and December. Salivary cortisol levels were measured by enzyme immunoassay (EIA), and salivary NO concentration was determined by the quantification of nitrite. The area under the curve in relation to ground (AUC) was calculated to assess the cortisol and nitrite concentrations throughout the day. No significant changes were observed in cortisol or nitrite concentrations across the three periods of the school year, while cortisol and nitrite levels showed a low positive and significant correlation (r=0.3455, p=0.0336). The results suggest that changes in salivary cortisol levels are accompanied by changes in salivary nitrite levels. This is the first time that such association has been demonstrated. These results encourage further studies aimed to confirm the importance of salivary NO measurement in relation to occupational stress and cardiovascular health.

Music stimuli lead to increased levels of nitrite in unstimulated mixed saliva.

Concentration of salivary nitrate is approximately 10-fold to that of serum. Many circumstances such as acute stress could promote salivary nitrate secretion and nitrite formation. However, whether other conditions can also be used as regulators of salivary nitrate/nitrite has not yet been explored. The present study was designed to determine the influence of exposure to different music on the salivary flow rate and nitrate secretion and nitrite formation. Twenty-four undergraduate students (12 females and 12 males) were exposed to silence, rock music, classical music or white noise respectively on four consecutive mornings. The unstimulated salivary flow rate and stimulated salivary flow rate were measured. Salivary ionic (Na+, Ca2+ Cl-, and PO43-) content and nitrate/nitrite levels were detected. The unstimulated salivary flow rate was significantly increased after classical music exposure compared to that after silence. Salivary nitrite levels were significantly higher upon classical music and white noise stimulation than those under silence in females. However, males were more sensitive only to white noise with regard to the nitrite increase. In conclusion, this study demonstrated that classical music stimulation promotes salivary nitrite formation and an increase in saliva volume was observed. These observations may play an important role in regulating oral function.

Salivary nitrite production is elevated in individuals with a higher abundance of oral nitrate-reducing bacteria.

Nitric oxide (NO) can be generated endogenously via NO synthases or via the diet following the action of symbiotic nitrate-reducing bacteria in the oral cavity. Given the important role of NO in smooth muscle control there is an intriguing suggestion that cardiovascular homeostasis may be intertwined with the presence of these bacteria. Here, we measured the abundance of nitrate-reducing bacteria in the oral cavity of 25 healthy humans using 16S rRNA sequencing and observed, for 3.5 h, the physiological responses to dietary nitrate ingestion via measurement of blood pressure, and salivary and plasma NO metabolites. We identified 7 species of bacteria previously known to contribute to nitrate-reduction, the most prevalent of which were Prevotella melaninogenica and Veillonella dispar. Following dietary nitrate supplementation, blood pressure was reduced and salivary and plasma nitrate and nitrite increased substantially. We found that the abundance of nitrate-reducing bacteria was associated with the generation of salivary nitrite but not with any other measured variable. To examine the impact of bacterial abundance on pharmacokinetics we also categorised our participants into two groups; those with a higher abundance of nitrate reducing bacteria (> 50%), and those with a lower abundance (< 50%). Salivary nitrite production was lower in participants with lower abundance of bacteria and these individuals also exhibited slower salivary nitrite pharmacokinetics. We therefore show that the rate of nitrate to nitrite reduction in the oral cavity is associated with the abundance of nitrate-reducing bacteria. Nevertheless, higher abundance of these bacteria did not result in an exaggerated plasma nitrite response, the best known marker of NO bioavailability. These data from healthy young adults suggest that the abundance of oral nitrate-reducing bacteria does not influence the generation of NO through the diet, at least when the host has a functional minimum threshold of these microorganisms.

Uticaj dužine pušenja cigareta na funkciju endotelijalnih ćelija

Objective. Cigarette smoking represents an important factor for development of cardiovascular diseases. Both the amount and duration of cigarette consumption are in direct correlation to the increase in cardiovascular pathology. The aim of our research was to evaluate the influence of the cigarette smoking duration on the nitrate, nitrite, S-nitrosothiols and endothelin-1 levels in saliva of smokers. Methods. For the purpose of this study, a total number of 60 subjects were recruited. They were divided into 4 groups based on the duration of cigarette consumption - non smokers - control (n=15); group I (n=15) - consumption of cigarettes for 2 years; group II (n=15) - 4 years and group III (n=15) - 6 years. The blood pressure, weight and height of all subjects were measured. Body mass index was calculated. Determination of nitrate, nitrite, S-nitrosothiols and endothelin-1 was done in each salivary sample collected from 60 subjects. Results. Both systolic and diastolic blood pressure did not differ between the control group and group I. These parameters were significantly greater in group II and group III subjects compared to controls. Smokers of all groups and non-smokers had normal body mass index values. The levels of salivary nitrate, nitrite and S-nitrosothiols were significantly lower in group II and group III compared to control group. Salivary endothelin-1 concentrations were significantly higher in subjects of group II and group III than controls. Conclusion. In this study the substantial differences between smokers and non-smokers in levels of stable metabolites of NO and endothelin-1 are due to the causal effects of smoking.

Salivary nitrate-nitrite conversion capacity after nitrate ingestion and incidence of Veillonella spp. in elderly individuals.

Dietary nitrate has several beneficial effects, including blood pressure reduction and improved oxygen consumption efficiency, but in order to do so it must first be reduced to nitrite by oral bacteria. Veillonella spp., a strictly anaerobic group, are the most prevalent nitrate-reducing bacteria in the oral cavity. In response to some early studies that have hinted at inter- and intra-individual variation in salivary nitrate-nitrite conversion capacity, the purpose of the present study was to investigate the incidence of and variation in the Veillonella species V. atypica, V. dispar, and V. rogosae by direct PCR and to assess salivary nitrate-nitrite conversion capacity and its reproducibility after dietary nitrate consumption in 24 elderly individuals. V. atypica, V. dispar, and V. rogosae were detected in 10 (41.7%), 24 (100%), and 14 (58.3%) participants, respectively. The coefficients of correlation between the first and second experiments on increased nitrate/nitrite were 0.637 and 0.583, respectively, both of which were statistically significant (P < 0.01). In both experiments, 6 participants produced relatively low levels of nitrite (<0.5 mM Δ nitrite) while 7 produced relatively high levels (>1.0 mM Δ nitrite). The data suggested that V. dispar was the most prevalent species, being present even in individuals producing low levels of salivary nitrite.

Flavonoid-Rich Apple Improves Endothelial Function in Individuals at Risk for Cardiovascular Disease: A Randomized Controlled Clinical Trial.

The cardioprotective effects of apples are primarily attributed to flavonoids, found predominantly in the skin. This study aimed to determine if acute and/or chronic (4 weeks) ingestion of flavonoid-rich apples improves endothelial function, blood pressure (BP), and arterial stiffness in individuals at risk for cardiovascular diseases (CVD). In this randomized, controlled cross-over trial, acute and 4 week intake of apple with skin (high flavonoid apple, HFA) is compared to intake of apple flesh only (low flavonoid apple, LFA) in 30 participants. The primary outcome is endothelial function assessed using flow-mediated dilation (FMD) of the brachial artery, while main secondary outcomes are 24 h ambulatory BP and arterial stiffness. Other outcomes include fasting serum glucose and lipoprotein profile, plasma heme oxygenase-1 (Hmox-1), F2 -isoprostanes, flavonoid metabolites, and plasma and salivary nitrate (NO3- ) and nitrite (NO2- ) concentrations. Compared to LFA control, the HFA results in a significant increase in FMD acutely (0.8%, p<0.001) and after 4 weeks chronic intake (0.5%, p<0.001), and in plasma flavonoid metabolites (p<0.0001). Other outcomes are not altered significantly. A lower risk of CVD with higher apple consumption could be mediated by the beneficial effect of apple skin on endothelial function, both acutely and chronically.

Possible Reactions of Dietary Phenolic Compounds with Salivary Nitrite and Thiocyanate in the Stomach.

Foods are mixed with saliva in the oral cavity and swallowed. While staying in the stomach, saliva is contentiously provided to mix with the ingested foods. Because a salivary component of nitrite is protonated to produce active nitrous acid at acidic pH, the redox reactions of nitrous acid with phenolic compounds in foods become possible in the stomach. In the reactions, nitrous acid is reduced to nitric oxide (•NO), producing various products from phenolic compounds. In the products, stable hydroxybezoyl benzofuranone derivatives, which are produced from quercetin and its 7-O-glucoside, are included. Caffeic acid, chlorogenic acid, and rutin are oxidized to quinones and the quinones can react with thiocyanic acid derived from saliva, producing stable oxathiolone derivatives. 6,8-Dinitrosocatechis are produced from catechins by the redox reaction, and the dinitrocatechins are oxidized further by nitrous acid producing the quinones, which can make charge transfer complexes with the dinitrosocatechin and can react with thiocyanic acid producing the stable thiocyanate conjugates. In this way, various products can be produced by the reactions of salivary nitrite with dietary phenolic compounds, and reactive and toxic quinones formed by the reactions are postulated to be removed in the stomach by thiocyanic acid derived from saliva.

Effect of saliva collection methods and oral hygiene on salivary biomarkers

The aim of this study was to evaluate the influence of unstimulated and stimulated saliva collection methods, as well as tooth brushing, on the secretion rate of salivary total protein, nitrite, total antioxidant capacity and alpha-amylase. Saliva of 14 healthy individuals were collected with stimulation using Salivette®, Parafilm® and chewing gum and without stimulation from spit with and without fluid accumulation, before and after oral hygiene. Total protein, nitrite, total antioxidant capacity and alpha-amylase concentration (sAA) were evaluated. The collection of saliva stimulated with Parafilm® and chewing gum increased the salivary flow (1.5 ± 0.4 and 3.4 ± 0.7 mL/min, respectively) and the secretion rate of salivary total protein (1.0 ± 0.2 and 2.3 ± 0.5 mg/min, respectively). Also, chewing gum increases the salivary nitrite secretion (213 ± 58 nmol/min) and total antioxidant capacity (410 ± 47 nmol trolox eq/min). Interestingly, the unstimulated method without saliva accumulation prior to collection resulted in low sAA levels (23,531 ± 7979 pixel density). Furthermore, oral hygiene decreased salivary flow (1.3 ± 0.5 to 1.0 ± 0.4 mL/min), reduced the secretion rate of total protein (1.0 ± 0.5 to 0.6 ± 0.2 mg/min, p < .05) and increased sAA (13,159 ± 7114 to 20,075 ± 25,656 pixel density, p < .05). The type of stimulation can activate autonomous receptors responsible for the secretion and composition of saliva. Therefore, the evaluation of saliva collection methods and oral hygiene on salivary biomarkers is important for understanding and standardizing variations in salivary composition to strengthen the use of saliva as a diagnostic fluid.

Salivary Nitrite in Patients with Type 2 Diabetes Mellitus: Role of Diabetic Pharmacotherapy

Salivary Nitrite in Patients with Type 2 Diabetes Mellitus: Role of Diabetic Pharmacotherapy

Validation of two point-of-care tests against standard lab measures of NO in saliva and in serum

Nitric oxide (NO) is an endogenous signaling molecule, which plays important roles in cardiometabolic health. A significant source of NO is dietary nitrate (NO3), which is initially metabolized by oral bacteria into nitrite (NO2−) and is subsequently converted into NO once digested in the acidic gastric environment. Inexpensive non-invasive tests for measuring nitrite from saliva have been developed as a means for individuals to monitor their NO bioavailability. However, few studies exist in the literature validating and comparing these products with standard lab assays. The objective of this study was to validate two commonly used commercial strips: Nitric Oxide Test Strips (Berkeley Test) and Nitric Oxide Indicator Strips (Neogenesis) against standard lab measures for saliva and serum nitrite/nitrate. A stratified random sample of 20 non-smoking, overweight or obese participants between 40 to 65 years of age, were selected for this study from the baseline data of the San Juan Overweight Adults Longitudinal Study (SOALS). There was a significant correlation between the measures from the two nitrite-detecting-strips after controlling for the stratification variables (metabolic syndrome, and mouthwash use) (r = 0.75). Measurements from both strips correlated significantly with salivary nitrite levels (r = 0.76 for Berkeley strips; r = 0.59 for Neogenesis). Neither of the strips had a significant correlation with the levels of saliva nitrate, serum nitrite and serum nitrate. In conclusion, commercially available Berkeley and Neogenesis strips provide a reasonable surrogate for salivary, but not for systemic nitrite levels.

The effects of essential oil, povidone-iodine, and chlorhexidine mouthwash on salivary nitrate/nitrite and nitrate-reducing bacteria.

Dietary nitrate is reduced to nitrite and nitric oxide by microbial flora, and this activity is beneficial to vascular health. It has been reported that this bacterial process is inhibited by chlorhexidine mouthwash, although the effects of other products are largely unknown. This study examined the effects of several treatments on salivary nitrate/nitrite and nitrate-reducing bacteria. Twelve university staff and students performed mouth-washing with water (control), essential oil, 0.35% povidone-iodine, or 0.0025% chlorhexidine and then ate 100 g lettuce (110 mg nitrate content), followed by collection of saliva and tongue bacteria at the baseline, and 1, 5, and 10 h thereafter. The individual treatments were separated by an interval of one week. Salivary nitrate/nitrite was measured by the calorimetric method, and a representative nitrate-reducing bacterial species, Veillonella dispar, was detected and semi-quantified using a polymerase chain reaction (PCR) assay. Significant increases in salivary nitrate/nitrite were observed for all treatments (all P < 0.05). The PCR assay showed that water, essential oil, and povidone-iodine mouthwash had little effect, whereas V. dispar DNA bands were markedly inhibited after washing with chlorhexidine. These results suggest that essential oil and povidone-iodine mouthwash have little effect on oral nitrate-reducing activity. Salivary nitrite production was not reduced by chlorhexidine, but the fainter band of V. dispar DNA suggests that longer daily use might blunt this nitrate-reducing activity.