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Effects of pH, Nitrite, and Ascorbic Acid on Nonenzymatic Nitric Oxide Generation and Bacterial Growth in Urine

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Effects of pH, Nitrite, and Ascorbic Acid on Nonenzymatic Nitric Oxide Generation and Bacterial Growth in Urine

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Abstract

Nitrite may be generated by bacteria in urine dur-ing urinary tract infections. Acidification of nitrite results in the formation of nitric oxide (NO) and other reactive nitrogen oxides, which are toxic to a variety of microorganisms. We have studied NO for-mation and bacterial growth in mildly acidified hu-man urine containing nitrite and the reducing agent vitamin C. Urine collected from healthy sub-jects was incubated in closed syringes at different pH values with varying amounts of nitrite and/or ascorbic acid added. NO generation was measured in headspace gas using a chemiluminescence tech-nique. A similar setup was also used to study the growth of three strains of bacteria in urine. Mildly acidified nitrite-containing urine generated large amounts of NO and this production was greatly po-tentiated by ascorbic acid. The growth of Esche-richia coli, Pseudomonas aeruginosa, and Staphylo-coccus saprophyticus was markedly reduced by the addition of nitrite to acidified urine. This inhibition was enhanced by ascorbic acid. In conclusion, we show that the growth of three common urinary pathogens is markedly inhibited in mildly acidified urine when nitrite is present. The bacteriostatic effect of acidified nitrite is likely related to the re-lease of NO and other toxic reactive nitrogen inter-mediates. These results may help to explain the well-known beneficial effects of urinary acidifica-tion with, e.g., vitamin C in treatment and preven-tion of urinary tract infection. © 2001 Elsevier Science Urinary acidification has long been used as an aid in the treatment of urinary tract infections (1, 2). Various agents have been used in attempts to lower urinary pH, these include, for example, ammonium chloride, methionine, gluconic acid, methenamine hippurate, and ascorbic acid. In 1959 McDonald and Murphy (3) suggested the use of ascorbic acid as an acidifying agent because of its high excretion in urine. It has been shown that ascorbic acid when given at the same time as methenamine mandelate keeps the urine pH below 5.6 (4). Urinary acidifica-tion with vitamin C has long been used as a house-hold remedy in the treatment of urinary tract infec-tions, although evidence to support its efficacy has been largely anecdotal. Despite numerous studies the mechanism of any antibacterial action of vita-min C still remains unclear (5). During the last two decades several observations have established that nitric oxide (NO) 2 , besides being a potent vasodilator and neurotramsmittor, also has antimicrobial properties (6, 7). The enzy-matic formation of NO from L-arginine and molecu-lar oxygen is catalyzed by three different isoenzymes 1 To whom correspondence should be addressed at Depart-ment of Surgery, Section

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... 8,9 The antimicrobial activities of reactive nitrogen species are exemplified by nitric oxide (NO). 10 The aim of the current study was to evaluate the antibacterial effects of human cathelicidin LL-37 in vitro used either alone or in combination with AN to find out if they have synergistic or antagonistic effects against standard strains of E.coli ATCC 25922 and S. aureus ATCC 25923 when used together. This idea is based on the knowledge that the two antibacterial agents can be present together at sites of infection. ...
... To determine the effect of acidified nitrite (AN) on the antimicrobial activity of LL-37, the experiments were repeated using the combination of solutions containing the two agents to give final concentrations of LL-37 ranging from 2-64μg/ml and fixed concentration of AN at (10μM NaNO2+5 mM AA) to achieve final pH 5 as reported by Carlsson et al. 10 ...
... At a low pH, nitrite is converted to a variety of nitrogen oxides that are toxic to bacteria. 10 The present study shows that LL-37 has antibacterial activity when used alone or in combination with the other test compound. This antibacterial activity was dose-dependent between the concentrations of 2 to 64µg/ ml. ...
... [11][12][13] Originally, nitrate was considered the endpoint product of endogenous NO metabolism; however, several studies have found that nitrate can be used as a potential donor of NO under specific circumstances. [25][26][27] The beneficial effects of nitrate have been demonstrated in a variety of animal and clinical trials. The nitrate-nitrite-NO pathway could modulate mitochondrial function and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase to alleviate cardiorenal disease and ischemia-reperfusion injury. ...
... These results suggest that nitrate may act through the nitrate-nitrite-NO pathway via XOR. [11][12][13][25][26][27] XOR catalyzes the metabolism of xanthine into uric acid and has been found to degrade nitrate to NO in biochemical experiments. 26 XOR is believed to be the primary source of nitrite-derived NO in living organisms. ...
Article
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Objective: Nitrate has been reported to protect cells via the nitrate-nitrite-nitric oxide (NO) pathway. Most studies tend to use nitrite to investigate the mechanisms of this pathway. However, the latest studies have confirmed that mammals can directly degrade nitrate via xanthine oxidoreductase (XOR). The hypothesis is that nitrate could play a protective role in inflammatory responses independent of bacterial nitrate reductases. Methods: Mouse RAW264.7 macrophages were pre-incubated with sodium nitrate (10, 100, and 500 µM) for 2 hours, and then treated with lipopolysaccharide (LPS) for 2 hours to induce inflammation. The Quantikine Immunoassay was used to measure interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) concentrations in the supernatant. The fluorescence intensity ratio of red/green from JC-1 was used to assay mitochondrial membrane potential. The fluorescence intensity of MitoSOX Red was used to indicate the generation of mitochondrial reactive oxygen species. Results: Nitrate partially reduced IL-6 and TNF-α secretion via reducing NF-κB signaling in LPS-induced macrophages. Nitrate also reduced the generation of mitochondrial reactive oxygen species by regulating mitochondrial function. These effects depended on XOR-derived NO but were independent of inducible nitric oxide synthase-derived NO. Conclusion: Nitrate regulates mitochondrial function via XOR-derived NO to partially inhibit LPS-induced inflammation.
... Variations in the levels of some urinary constituents such as glucose can influence microbial growth; glucosuria, for example, enhances the growth of E. coli in urine (Geerlings et al., 1999). Urine is naturally antiseptic due to its chemical makeup; nitrite in acidified urine inhibits the growth of gram-negative bacteria (Carlsson et al., 2001), and urinary proteins such as Tamm-Horsfall protein are antimicrobial (Raffi et al., 2005;Saemann et al., 2005). The low pH of urine combined with high concentrations of urea and hypertonicity inhibits the growth of most urogenital tract commensals (Chambers and Lever, 1996;Kucheria et al., 2005;Sobel, 1985). ...
... Comparisons with comprehensive studies of 'typical human urine' (Bouatra et al., 2013;Putnam, 1971) reveal that most of the concentrations of the constituents of the proposed Composite SHU medium (Ipe et al., 2016) are within normal ranges. However, a noteworthy constituent in Composite SHU medium, as proposed in a recent review (Ipe et al., 2016) and validated in the current study, which differs from earlier standard SHU formulations, as described in physiology and cell biology studies (Robertson and Scurr, 1986), is the necessary inclusion of 0.1% (v/v) casamino acids to support the growth of gram-negative bacteria, which have been shown to grow in artificial or normal human urine in multiple prior studies (Martino et al., 2003;Ong et al., 2009;Wenzler-Rottele et al., 2006;Roos et al., 2006a;Roos et al., 2006b;Russo et al., 1996;Vejborg et al., 2012;Carlsson et al., 2001;Storer et al., 2011;Deutch et al., 2006;Chambers and Lever, 1996) and additional supplementation with 0.2% (v/v) yeast extract to support the growth of gram-positive bacteria, which have been shown to grow in artificial or normal human urine in multiple prior studies (Ipe et al., 2015;Sakinc et al., 2009;Vebo et al., 2010;La Rosa et al., 2012). Dextrose and yeast nitrogen base supplementation has been used for fungal UTI studies such as for Candida sp. ...
... Oral intake of propolis and Hibiscus sabdariffa can decrease urinary pH [6]. Infected urine contains large amounts of nitrite, and acidification of nitrite results in the formation of nitric oxide (NO), which has antimicrobial properties [7,8]. ...
... Oral intake of propolis and Hibiscus sabdariffa can decrease urinary pH [6]. Infected urine contains large amounts of nitrite, and acidification of nitrite results in the formation of nitric oxide, which possesses antimicrobial properties [7,8]. Those mechanisms might explain the potential effectiveness of propolis and Hibiscus sabdariffa in the control of UTI. ...
Article
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Background: The objective of the study was to analyze the efficacy and safety of a medical device containing reticulated gelatin with hibiscus and propolis (RGHP) in the early treatment of patients with symptoms of urinary tract infection (UTI).
... It has been reported that cadaverine production increases when UPEC strains are grown under nitrosative stress, potentially conditioning them for enhanced colonization of the host 31,32 . This is of relevance since, during a UTI, there is a significant formation of nitric oxide and reactive nitrogen intermediates 33,34 . Nevertheless, the specific role of cadaverine in resistance to nitrosative stress is unclear. ...
... This contrasts with results in patients with BV, where cadaverine and tyramine are also elevated 8,9 . Using an in vitro model, cadaverine was found to be significantly increased, perhaps due to nitrosative stress 33,35 which is higher in a steady-state culture than in the human bladder, allowing for its accumulation. The www.nature.com/scientificreports ...
Article
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Many women suffer from urinary tract infections (UTIs). In addition to pain and increased urgency to urinate, malodour is a significant issue for these patients. The specific factors causing this malodour are unclear, and there are no targeted treatment options to counteract it effectively. We used a metabolomics approach to compare the chemical composition of metabolites in the urine of women with E. coli UTIs (n = 15) and those who are healthy (n = 10). The biogenic amines trimethylamine and putrescine, which cause malodour in other urogenital conditions, were significantly increased in UTI patients. Conversely, the precursor of trimethylamine, trimethylamine N-oxide, was lower. To further confirm the source of the malodorous compounds, in vitro experiments were conducted by incubating strains of uropathogenic E. coli in sterilized urine from healthy women. All tested strains accumulated trimethylamine and putrescine. Notably, cadaverine was also produced by E. coli strains in vitro; however, it was not significantly different between both groups. We confirmed that the malodorous amines TMA and putrescine are found in higher concentrations in the urine of patients with an E.coli-caused UTI.
... This latter observation is not surprising as it is established that DHA and MDHA scavenge NO [46,47]. The slowing of NO loss in hypoxia is probably due to reduction of nitrite (formed from NO oxidation during the experiment) by AA, which has been characterized in many systems [48,49,50] and is discussed in more detail below. ...
... doi:10.1371/journal.pone.0082611.g004 conditions inside hypoxic plant cells favor NO production by the direct reduction of nitrous acid by AA [21,48]. Our final experiment investigating the role of AA in NO metabolism was designed to test whether such conditions could yield measurable NO concentrations in vitro. ...
Article
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Ascorbic acid and hemoglobins have been linked to nitric oxide metabolism in plants. It has been hypothesized that ascorbic acid directly reduces plant hemoglobin in support of NO scavenging, producing nitrate and monodehydroascorbate. In this scenario, monodehydroascorbate reductase uses NADH to reduce monodehydroascorbate back to ascorbate to sustain the cycle. To test this hypothesis, rates of rice nonsymbiotic hemoglobin reduction by ascorbate were measured directly, in the presence and absence of purified rice monodehydroascorbate reductase and NADH. Solution NO scavenging was also measured methodically in the presence and absence of rice nonsymbiotic hemoglobin and monodehydroascorbate reductase, under hypoxic and normoxic conditions, in an effort to gauge the likelihood of these proteins affecting NO metabolism in plant tissues. Our results indicate that ascorbic acid slowly reduces rice nonsymbiotic hemoglobin at a rate identical to myoglobin reduction. The product of the reaction is monodehydroascorbate, which can be efficiently reduced back to ascorbate in the presence of monodehydroascorbate reductase and NADH. However, our NO scavenging results suggest that the direct reduction of plant hemoglobin by ascorbic acid is unlikely to serve as a significant factor in NO metabolism, even in the presence of monodehydroascorbate reductase. Finally, the possibility that the direct reaction of nitrite/nitrous acid and ascorbic acid produces NO was measured at various pH values mimicking hypoxic plant cells. Our results suggest that this reaction is a likely source of NO as the plant cell pH drops below 7, and as nitrite concentrations rise to mM levels during hypoxia.
... 22 As the solution pH remained between 7.3 to 7.6 upon the addition of 5 mM nitrite−ascorbic acid, we believe that a major proportion of nitric oxide was generated through the redox cycling pathway rather than through the acidification of nitrite pathway. 22,23 However, we cannot rule out the possible formation of HNO 2 in our system, which may contribute to the overall NO release, although the system was well-mixed during the addition of ascorbic acid. A dip in the nitric oxide level was also observed in the period between 100 and 250 s (Figure 2). ...
... It appears that nitrite and ascorbic acid in this case inhibit the cell proliferation of nitrifying bacteria. Such "antibiofilm" activity of the nitrite− ascorbic acid system is in agreement with the earlier reports, in particular on Gram-negative pathogens, 23,25,26 but no study had been done previously on environmentally relevant bacteria, such as nitrifying bacteria. The suppressed biofilm growth in our study appears to result from the quick burst of nitric oxide generation observed in the nitrite−ascorbic acid system ( Figure 2). ...
Article
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In this study, catalytic generation of nitric oxide by a copper(II) complex embedded within a poly(vinyl chloride) matrix in the presence of nitrite (source of nitric oxide) and ascorbic acid (reducing agent) was shown to effectively control the formation and dispersion of nitrifying bacteria biofilms. Amperometric measurements indicated increased and prolonged generation of nitric oxide with the addition of the copper complex when compared to that with nitrite and ascorbic acid alone. The effectiveness of the copper complex-nitrite-ascorbic acid system for biofilm control was quantified using protein analysis, which showed enhanced biofilm suppression when the copper complex was used in comparison to that with nitrite and ascorbic acid treatment alone. Confocal laser scanning microscopy (CLSM) and LIVE/DEAD staining revealed a reduction in cell surface coverage without a loss of viability with the copper complex and up to 5 mM of nitrite and ascorbic acid, suggesting that the nitric oxide generated from the system inhibits proliferation of the cells on surfaces. Induction of nitric oxide production by the copper complex system also triggered the dispersal of pre-established biofilms. However, the addition of a high concentration of nitrite and ascorbic acid to a pre-established biofilm induced bacterial membrane damage and strongly decreased the metabolic activity of planktonic and biofilm cells, as revealed by CLSM with LIVE/DEAD staining and intracellular adenosine triphosphate measurements, respectively. This study highlights the utility of the catalytic generation of nitric oxide for the long-term suppression and removal of nitrifying bacterial biofilms.
... Next, we tried to prepare and analyze DAF-FM-T containing the stable isotope nitrogen 15 N derived from NO. It was previously shown that nitrite is protonated and reduced to NO under acidic conditions (Carlsson et al. 2001). Therefore, 50 μm of DAF-FM was reacted with 10 mm 14 N-nitrite or 15 N-nitrite in phosphate buffer (pH 2.5) to prepare DAF-FM-T containing 14 N or 15 N, respectively. ...
Article
Nitric oxide (NO) is a ubiquitous signaling molecule synthesized from various nitrogen sources. An analytical method to identify a nitrogen source for NO generation was developed using liquid chromatography with tandem mass spectrometry in combination with stable isotope labeling. Our method successfully detected the 15N-labeled NO-containing compound generated from 15N-labeled substrate nitrite in vitro and in vivo.
... Nitrite is unique to the nitrogen oxides in its redox position between oxidative (NO2 radical) and reductive (NO radical) signalling and its relative stability in blood and tissue [Gladwin et al., 2005]. Once nitrite is formed, there are numerous pathways in the body for its further reduction to NO involving haemoglobin [Cosby et al., 2003], myoglobin [Rassaf et al., 2007], xanthine oxidoreductase [Godber et al., 2000], ascorbate [Carlsson et al., 2001], polyphenols [Gago et al., 2007] and protons [Benjamin et al., 1994] (Table 1). The generation of NO by these pathways is greatly enhanced during hypoxia and acidosis, thereby ensuring NO production in situations for which the oxygen-dependent NOS enzyme activities are compromised [Oestergaad et al., 2007]. ...
... Oral cavity commensal bacteria reduce nitrate to nitrite, which has a high oral bioavailability (>90 %) [61,63,64]. Nitrite present in the blood stream is reduced directly to NO, a reaction catalyzed by several molecules, particularly deoxygenated myoglobin [36, [71][72][73], and deoxygenated hemoglobin [74], but also xanthine oxidoreductase [75], mitochondrial respiratory chain enzymes [76], aldehyde oxidase [77], carbonic anhydrase [78], vitamin C [79], polyphenols [80,81], and even endothelial NO synthase [82,83]. Importantly, the conversion of nitrite to NO occurs much more rapidly in the context of hypoxia [84•] and acidosis [85•]. ...
Article
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The pathogenesis of exercise intolerance in patients with heart failure and preserved ejection fraction (HFpEF) is likely multifactorial. In addition to cardiac abnormalities (diastolic dysfunction, abnormal contractile reserve, chronotropic incompetence), several peripheral abnormalities are likely to be involved. These include abnormal pulsatile hemodynamics, abnormal arterial vasodilatory responses to exercise, and abnormal peripheral O2 delivery, extraction, and utilization. The nitrate-nitrite-NO pathway is emerging as a potential target to modify key physiologic abnormalities, including late systolic left ventricular (LV) load from arterial wave reflections (which has deleterious short- and long-term consequences for the LV), arterial vasodilatory reserve, muscle O2 delivery, and skeletal muscle mitochondrial function. In a recently completed randomized trial, the administration of a single dose of exogenous inorganic nitrate has been shown to exert various salutary arterial hemodynamic effects, ultimately leading to enhanced aerobic capacity in patients with HFpEF. These effects have the potential for both immediate improvements in exercise tolerance and for long-term “disease-modifying” effects. In this review, we provide an overview of key mechanistic contributors to exercise intolerance in HFpEF, and of the potential therapeutic role of drugs that target the nitrate-nitrite-NO pathway.
... A proportion of the swallowed nitrite-rich saliva is protonated in the acidic environment of the stomach forming nitrous acid (HNO 2 ) which in turn breaks down to form NO and other nitrogen oxide moieties [22,24]. This reaction is enhanced by increasing concentrations of nitrite, as well as by a low stomach pH and the presence of ascorbic acid [45] or polyphenols [46,47]. The gastric generation of NO has been shown to promote effects to support gastric mucosal integrity, including increased mucosal blood flow and mucus generation [48,49]. ...
Article
Nitric oxide (NO) is generated endogenously by NO synthases to regulate a number of physiological processes including cardiovascular and metabolic functions. A decrease in the production and bioavailability of NO is a hallmark of many major chronic diseases including hypertension, ischaemia-reperfusion injury, atherosclerosis and diabetes. This NO deficiency is mainly caused by dysfunctional NO synthases and increased scavenging of NO by the formation of reactive oxygen species. Inorganic nitrate and nitrite are emerging as substrates for in vivo NO synthase-independent formation of NO bioactivity. These anions are oxidation products of endogenous NO generation and are also present in the diet, with green leafy vegetables having a high nitrate content. The effects of nitrate and nitrite are diverse and include vasodilatation, improved endothelial function, enhanced mitochondrial efficiency and reduced generation of reactive oxygen species. Administration of nitrate or nitrite in animal models of cardiovascular disease shows promising results, and clinical trials are currently ongoing to investigate the therapeutic potential of nitrate and nitrite in hypertension, pulmonary hypertension, peripheral artery disease and myocardial infarction. In addition, the nutritional aspects of the nitrate-nitrite-NO pathway are interesting as diets suggested to protect against cardiovascular disease, such as the Mediterranean diet, are especially high in nitrate. Here, we discuss the potential therapeutic opportunities for nitrate and nitrite in prevention and treatment of cardiovascular and metabolic diseases.
... Since the 1920s, when the urinary nitrite test was first developed, the dipstick assay has been an important component of modern UTI diagnosis. The growth of bacteria in slightly acidic urine is inhibited if exogenous nitrite is added[3,4]. This inhibition was potentiated in the presence of the reducing agent ascorbic acid. ...
... An increase in NO generation would be expected to augment the antimicrobial of acidified nitrite. Ascorbic acid (vitamin C) has been shown to increase the formation of NO from nitrite 3,4,6 .The main problem with UTI that should be taken into consideration is related to the emergence of new strains of pathogens having multidrug resistance 2 . Nowadays physicians are treating UTI using high doses of antibiotic for long duration, which may impair renal function, particularly in old aged patients. ...
Article
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Background and Objectives: Urinary acidification induces strong antibacterial effect against different bacterial types involved in urinary tract infections. The aim of this study is to assess in vitro determination of the minimal inhibitory concentration of nitrite alone and acidified nitrite using addition of ascorbic acid in infected urine with different pH of the bacterial media. Methods: E.coli bacteria have been isolated from the urine of patients with urinary tract infection. Identification of E. coli was achieved using biochemical tests. Determination of bacteriostatic activity of acidified sodium nitrite was carried out using 96-wells microtiter plate. Acidification effect against UTI bacteria was assessed using sequential steps; feeding bacteria with NaNo3, then transferring them to acidified urine after incubation using ascorbic acid. Results: Ascorbic acid (40 mM) alone and sodium nitrite (200 mM) alone are considered to be a weak antibacterial agents on uropathgenic E. coli, while mixing 10 mM ascorbic acid with 625 µM sodium nitrite at pH 5 became a strong antibacterial agent against 14 isolates out of 32. Bacterial death can occur by sequential steps, first feeding bacteria with sodium nitrate and after incubation, transferring to the acidified urine at pH 5 and pH4.6 causing bacterial death. Conclusions: Strong antibacterial agent can be formed in acidified urine containing nitrite. This antibacterial agent is strongly pH and nitrite dependent and is increased by addition of ascorbic acid. The antibacterial strength is a nitrite-dose dependent in mildly acidified urine.
... Ascorbic acid (vitamin C) is often recommended as a supplement that can prevent recurrent UTIs by acidification of the urine [36]. In vitro studies suggest that it has a bacteriostatic effect in the urine, mediated by the reduction of urinary nitrites to reactive nitrogen oxides rather than by lowering urinary pH [55,56]. However, convincing clinical evidence to support this is lacking. ...
Article
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INTRODUCTION AND HYPOTHESIS: Recurrent urinary tract infections (UTIs) are more common in women and are frequently defined as ≥2 episodes in the last 6 months or ≥3 episodes in the last 12 months. In a primary care setting, 53 % of women above the age of 55 years and 36 % of younger women report a recurrence within 1 year. Thus, management and prevention of recurrent UTI is of utmost significance. This review aims to highlight the latest research in prevention strategies and suggest a management pathway. METHODS: A search was conducted on MEDLINE, Embase and the Cochrane Database of Systematic Reviews databases for the latest systematic reviews and high-quality randomized controlled trials. Special emphasis was placed on the remit "recurrent" and strongly adhered to. Furthermore, a Google search was conducted for current guidelines on the management of UTIs. RESULTS: Current prevention strategies include eliminating risk factors that increase the risk of acquiring recurrent UTI and continuous, post-coital and self-initiated antimicrobial prophylaxis. Other prospective preventative strategies, currently under trial, include use of vaccinations, D-mannose and lactobacillus (probiotics). CONCLUSION: Although risk factors should be identified and addressed accordingly, individualized antibiotic prophylaxis remains the most effective method of management. Non-antibiotic prevention strategies such as cranberry, vitamin C and methenamine salts lack strong evidence to be introduced as routine management options and as alternatives to antibiotics. Based on current evidence and guidelines, a management pathway is recommended. Emerging therapies require further evaluation before they can be recommended.
... This inhibition was enhanced by ascorbic acid. 10 Although is has been suggested that vitamin C has some effect on urinary acidification, reducing the urinary pH, the role of vitamin C as a urinary acidifier is still controversial. In this case, the diagnosis was suggested on clinical manifestations, urine pH, CT findings and cystoscopy. ...
Article
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Encrusted cystitis and pyeloureteritis are rare chronic infectious conditions characterized by mucosal inflammation and encrustations of the urinary tract. It is caused by fastidious growing urea splitting microorganisms, mainly Corynebacterium. Herein, we report an unusual case of an 80-year-old man with encrusted cystitis and pyeloureteritis who was previously treated with transcatheteral arterial chemoembolization for hepatocellular carcinoma. Abdomino-pelvic computerized tomography showed a bilateral hydronephrosis with calcifications of renal pelvis, ureter, and bladder. Cystoscopy showed calcified bladder mucosa with necrosis and bleeding. After transurethral removal of calcified plaques, the patient was treated with antibiotic and oral urine acidification. One-month follow-up cystoscopy showed that inflammation was improved and calcification was significantly reduced.
... Wound dressings that release NO levels typically at 500 ppm appeared to be efficient at killing biofilms of nosocomial pathogens Acinetobacter baumannii, MRSA, and P. aeruginosa when assessed in in vitro experiments [100]. The use of toxic NO has also been investigated for treatment of urinary tract infections, where the addition of 10 mM ascorbate and nitrite at 50 M to 5 mM, which under these conditions generate equimolar NO, cleared E. coli infections in artificial urine and in an urinary tract model [101,102]. Another study demonstrated the effectiveness of NO-charged catheters, typically releasing 2 to 60 M NO, in preventing E. coli infections [103]. ...
Article
Full-text available
Studies of the biofilm life cycle can identify novel targets and strategies for improving biofilm control measures. Of particular interest are dispersal events, where a subpopulation of cells is released from the biofilm community to search out and colonize new surfaces. Recently, the simple gas and ubiquitous biological signaling molecule nitric oxide (NO) was identified as a key mediator of biofilm dispersal conserved across microbial species. Here, we review the role and mechanisms of NO mediating dispersal in bacterial biofilms, and its potential for novel therapeutics. In contrast to previous attempts using high dose NO aimed at killing pathogens, the use of low, non-toxic NO signals (picomolar to nanomolar range) to disperse biofilms represents an innovative and highly favourable approach to improve infectious disease treatments. Further, several NO-based technologies have been developed that offer a versatile range of solutions to control biofilms, including: (i) NO-generating compounds with short or long half-lives and safe or inert residues, (ii) novel compounds for the targeted delivery of NO to infectious biofilms during systemic treatments, and (iii) novel NO-releasing materials and surface coatings for the prevention and dispersal of biofilms. Overall the use of low levels of NO exploiting its signaling properties to induce dispersal represents an unprecedented and promising strategy for the control of biofilms in clinical and industrial contexts.
... The host-defence mechanism of nitrite reduction has also been demonstrated to play a role in the urinary tract (52,53) and on the skin (54,55). ...
... It has been shown that nitrate absorbed from digestive tract can be reduced to nitrite by bacterial nitrate reductases in the oral cavity [7]. Nitrite can be further reduced to NO by several pathways including deoxyhemoglobin [8,9], deoxymyoglobin [10], xanthine oxidase [11,12], and non-enzymatic reduction in the presence of protons [13,14] or ascorbic acid [15]. Since nitrate and nitrite can be easily obtained from our diet, the potential NO-related bioactivities of those anions are getting more attention with regard to the cardiovascular benefits [16,17]. ...
Article
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Nitric oxide (NO), a small gas molecule, has long been known to be a potent inhibitor of platelet function but the physiological and pathological implications of platelet inhibition by NO have not been well clarified. We recently showed that the addition of nitrite to platelet-rich plasma in the presence of erythrocytes could inhibit platelet aggregation and this inhibitory effect of nitrite + erythrocytes was enhanced by deoxygenation of erythrocytes as measured by P-selectin expression and cGMP production. In order to study the nitrite effect on platelets at different oxygen levels, we used the flow cytometric assays to detect platelet membrane surface markers upon activation. The P-selectin and activated gpIIb/IIIa expression on platelet membranes in response to ADP, collagen and thrombin stimulation was measured at various hematocrit and oxygen levels. Nitrite (0.1 to 1.0 μM) significantly decreased the percentage of these surface markers on the platelet membrane at the hematocrit values above 23% and oxygen levels lower than 49 mmHg. The inhibitory effect of nitrite was augmented by increasing hematocrit values and decreasing oxygen saturation. C-PTIO (an NO scavenger) prevented the platelet inhibition by nitrite + erythrocytes whereas the inhibitors of NO synthase and xanthine oxidoreductase had no effect. These results support the proposal that circulating nitrite decreases platelet reactivity in the presence of partially deoxygenated erythrocytes through its reduction to NO, which may also explain certain differences between arterial and venous thrombosis and support directly the role of deoxyhemoglobin in this process. We believe that our flow cytometric assays offer a possibility to identify the individual molecular process involved in these effects.
... Ascorbic acid is taken up into target cells via specific transporters (106) . Ascorbic acid can also be reabsorbed by renal tube cells (107) . ...
Article
The biofortification of staple crops with vitamins is an attractive strategy to increase the nutritional quality of human food, particularly in areas where the population subsists on a cereal-based diet. Unlike other approaches, biofortification is sustainable and does not require anything more than a standard food-distribution infrastructure. The health-promoting effects of vitamins depend on overall intake and bioavailability, the latter influenced by food processing, absorption efficiency and the utilisation or retention of the vitamin in the body. The bioavailability of vitamins in nutritionally enriched foods should ideally be adjusted to achieve the dietary reference intake in a reasonable portion. Current vitamin biofortification programmes focus on the fat-soluble vitamins A and E, and the water-soluble vitamins C and B9 (folate), but the control of dosage and bioavailability has been largely overlooked. In the present review, we discuss the vitamin content of nutritionally enhanced foods developed by conventional breeding and genetic engineering, focusing on dosage and bioavailability. Although the biofortification of staple crops could potentially address micronutrient deficiency on a global scale, further research is required to develop effective strategies that match the bioavailability of vitamins to the requirements of the human diet.
... Since American cranberries (Vaccinium macrocarpon Ait.), and other Vaccinium berries (e.g., lingonberry, blueberry, European cranberry, etc.) or their products (juices, concentrates, and dietary supplements) contain proanthocyanidins, anthocyanins, ascorbic acid, fructose, and other compounds; although there have been mixed results in the literature on its effectiveness, regular consumption are thought to prevent urinary tract infections (UTIs) and offer other potential health benefits (Carlsson, Wiklund, Engrstrand, Weitzberg, & Lundberg, 2001;Foo, Lu, Howell, & Vorsa, 2000;Howell, 2002;Zafriri, Ofek, Adar, Pocinio, & Sharon, 1989). Problems with experimental differences regarding the ingested form (extract, juice, juice cocktail, concentrate, capsule, tablet, etc.), dosage, duration, and frequency, or the age and other demographics of subjects, or relevant infective organism monitored in the clinical trial have frequently been cited (Barbosa-Cesnik et al., 2011;Jepson & Craig, 2007McMurdo, Argo, Phillips, Daly, & Davey, 2009) and were well summarized by Raz, Chazan, and Dan (2004) and Jepson and Craig (2008). ...
Article
In this study, five common proanthocyanidin purification techniques were evaluated prior to phloroglucinolysis, followed by HPLC analysis. An optimized purification method was then used to identify and quantify the proanthocyanidins (extension and terminal units of epigallocatechin, catechin, epicatechin, A type trimer, and A type dimer) of commercially available cranberry products (juices, concentrates, tablets, and capsules; n = 17). Two size exclusion beads (Toyopearl 4 TSK HW-40C and Sephadex LH-20) were found suitable for proanthocyanidin purification, though proanthocyanidin extension and terminal unit composition was contingent upon the cleanup procedure utilized. These data illustrate that purification methods require consideration prior to conducting any cranberry proanthocyanidin analyses, and have to be accounted for when comparing values between studies. Total proanthocyanidin levels ranged from 11.7 (juice) to 436.4 (tablet) mg/100 mL or 100 g values obtained from Sephadex LH-20 purification, while total anthocyanin levels ranged from 0.54 (juice) to 98.00 (tablet) mg/100 mL or 100 g.
... When inorganic nitrite (NO 2 À ) is acidified, a variety of reactive RNIs are generated, including NO, a gas with antimicrobial properties [17,18]. In the presence of the reducing agent ascorbic acid, NO generation from nitrite is greatly enhanced [19]. Urinary pathogens such as Escherichia coli and Pseudomonas aeruginosa are killed if they are exposed to the combination of mildly acidified urine (pH 5 to 5.5) and nitrite, and the antibacterial effect is further enhanced by ascorbic acid [20,21]. ...
Article
Antibacterial nitrogen oxides including nitric oxide are formed from nitrite under acidic conditions. In a continuous-flow model of the urinary bladder we used the retention cuff of an all-silicone Foley catheter as a depot for export of nitrogen oxides. The cuff was filled with sodium nitrite (50mM) and an acidic buffer solution (pH 3.6) and the growth of nine common uropathogens in the surrounding artificial urine was measured along with biofilm formation on the catheter surface. In experiments with control catheters (NaCl) bacteria grew readily and biofilm developed within hours in five out of nine strains. In contrast, with test catheters bacterial counts were markedly reduced and biofilm formation by P. aeruginosa, K. pneumoniae and E. cloace was prevented while E. coli and S. aureus was unaffected. We conclude that antibacterial nitrogen oxides generated in the retention cuff of a urinary catheter diffuse into urine and prevent the growth of urinary pathogens and biofilm formation. Although promising, future studies will reveal if this novel approach can be clinically useful for the prevention of catheter-associated urinary tract infections.
... This effect was shown to be mediated by the reduction of urinary nitrites to reactive nitrogen oxides rather then by lowering urinary pH. 31,32 Methenamine Salts. Methenamine salts are hydrolyzed in the urine to form ammonia and formaldehyde. ...
Article
Recurrence after urinary tract infection (rUTI) is common in adult women. The majority of recurrences are believed to be reinfection from extraurinary sources such as the rectum or vagina. However, uropathogenic Escherichia coli are now known to invade urothelial cells and form quiescent intracellular bacterial reservoirs. Management of women with frequent symptomatic rUTI can be particularly vexing for both patients and their treating physicians. This review addresses available and promising management strategies for rUTI in healthy adult women.
... This confirms that NO 3 ( is the key 'active' ingredient responsible for the physiological changes observed following beetroot juice supplementation. It does not rule out, however, a synergistic role for other components of beetroot juice such as antioxidants and polyphenols which may facilitate the reduction of NO 3 ( to NO 2 ( and NO (Carlsson, Wiklund, Engstrand, Weitzberg, & Lundberg, 2001;Gago, Lundberg, Barbosa, & Laranjinha, 2007). Collectively, these results indicate that the reduced V O 2 following NO 3 ( supplementation is reproducible and can be observed across a range of different supplementation regimes and exercise modalities. ...
Article
Nitric oxide (NO) is a potent signalling molecule that influences an array of physiological responses. It was traditionally assumed that NO was derived exclusively via the nitric oxide synthase (NOS) family of enzymes. This complex reaction requires a five electron oxidation of L-arginine and is contingent on the presence of numerous essential substrates (including O2) and co-factors. Recently an additional, O2-independent, NO generating pathway has been identified, where nitrite (NO2 ) can undergo a simple one electron reduction to yield NO. NO2 is produced endogenously from the oxidation of NO and also from the reduction of dietary nitrate (NO3 ) by facultative bacteria residing on the tongue. Recent data show that dietary NO3 supplementation, which increases the circulating plasma [NO2 ], reduces the O2 cost of submaximal exercise in healthy humans. This finding is striking given that efficiency during moderate-intensity exercise has been considered to be immutable. There is evidence that the muscle ATP turnover at a fixed work rate is reduced and the mitochondrial P/O ratio is increased following NO3 supplementation, which offers important insights into the physiological bases for the reduced [Vdot] during exercise. NO3 supplementation has also been shown to improve exercise performance in both healthy and patient populations. Therefore, dietary NO3 supplementation may represent a practical and cost-effective method to improve exercise efficiency and exercise tolerance in humans. Given that a NO3 -rich diet may have numerous cardiovascular and other health benefits, dietary NO3 intake may have important implications for human lifelong health and performance.
... Flavonoids are proposed to modulate endothelial-dependent NO release, and nitrates impact on NO production from nitrite intermediates and it is possible that their combined consumption may result in additive or synergistic vascular responses. Furthermore formation of NO and other reactive nitrogen species in the stomach is enhanced by increasing nitrite concentrations, lower stomach pH and the presence of vitamin C or polyphenols (134)(135)(136) . Bondonno et al. (106) investigated the independent and additive effects of consumption of flavonoid-rich apples and nitrate-rich spinach. ...
Article
CVD remain the leading cause of death globally. Effective dietary strategies for their reduction are of high priority. Increasing evidence suggests that phytochemicals, particularly dietary flavonoids and nitrates, are key modulators of CVD risk reduction through impact on multiple risk factors. The aim of this review is to explore the evidence for the impact of flavonoid- and nitrate-rich foods and supplements on CVD risk, with specific reference to their importance as mediators of vascular health and platelet function. There is accumulating evidence to support benefits of dietary flavonoids on cardiovascular health. Dose-dependent recovery of endothelial function and lowering of blood pressure have been reported for the flavanol (-)-epicatechin, found in cocoa, apples and tea, through production and availability of endothelial nitric oxide (NO). Furthermore, flavonoids, including quercetin and its metabolites, reduce in vitro and ex vivo platelet function via inhibition of phosphorylation-dependent cellular signalling pathways, although further in vivo studies are required to substantiate these mechanistic effects. Hypotensive effects of dietary nitrates have been consistently reported in healthy subjects in acute and chronic settings, although there is less evidence for these effects in patient groups. Proposed mechanisms of actions include endothelial-independent NO availability, which is dependent on the entro-salivary circulation and microbial conversion of dietary nitrate to nitrite in the mouth. In conclusion, flavonoid- and nitrate-rich foods show promising effects on vascular function, yet further randomly controlled studies are required to confirm these findings and to determine effective doses.
... In a study with 110 pregnant women, a group taking ferrous sulfate 200 mg/day, folic acid 5 mg/day and ascorbic acid 100mg/day was compared with a group taking only ferrous sulfate and folic acid, and the finding showed that the frequency of UTI was lower in the group taking ascorbic acid for 3 months [32]. Currently, it is found in vitro experiments that vitamin C has bacteriostatic effect in urine, and the effect is thought to arise by vitamin C restoring urinary nitrites to active nitrogen oxides rather than by decreasing urinary pH [33,34]. ...
... Nitrite is further metabolized in blood and tissues into a variety of bioactive nitrogen oxides. This reduction is catalyzed through numerous pathways involving myoglobin/hemoglobin, ascorbate, xanthine oxireductase, and polyphenols [12,[15][16][17][18][19][20][21][22]. The production of NO from these pathways is enhanced by hypoxic and acidotic conditions [23][24][25]. ...
Article
Full-text available
Nitric oxide (NO) is a short-lived, ubiquitous signaling molecule that affects numerous critical functions in the body. There are markedly conflicting findings in the literature regarding the bimodal effects of NO in carcinogenesis and tumor progression, which has important consequences for treatment. Several preclinical and clinical studies have suggested that both pro- and antitumorigenic effects of NO depend on multiple aspects, including, but not limited to, tissue of generation, the level of production, the oxidative/reductive (redox) environment in which this radical is generated, the presence or absence of NO transduction elements, and the tumor microenvironment. Generally, there are four major categories of NO-based anticancer therapies: NO donors, phosphodiesterase inhibitors (PDE-i), soluble guanylyl cyclase (sGC) activators, and immunomodulators. Of these, NO donors are well studied, well characterized, and also the most promising. In this study, we review the current knowledge in this area, with an emphasis placed on the role of NO as an anticancer therapy and dysregulated molecular interactions during the evolution of cancer, highlighting the strategies that may aid in the targeting of cancer.
... conditions in the patient, or by clinical interventions that may disrupt drug efficacy (ascorbic acid treatment of urinary tract infections to lower urine pH) (Carlsson et al., 2001). (5) Many patients that develop multidrug-resistant infections have co-morbidities, immunosuppressive therapy and/or the presence of invasive medical devices that impact susceptibility to indicated pathogens (Paterson and Bonomo, 2005). ...
Article
Full-text available
Drug testing often excludes potent antibiotics for the treatment of bacterial infections, while frequently identifying antibiotics that are ineffective. However, drug testing under conditions that mimic natural infections succeeded in identifying effective antibiotics, even though these same antibiotics failed standard tests. This work suggests that standard drug-testing may be hindering patient treatment and slowing the process of discovery of new, effective, and safe antibiotics because it disqualifies effective compounds. These findings call for an overhaul of standardized drug testing which hasn't changed in > 50 years.
... Additionally, the inhibition was enhanced by vitamin C. These results help to explain the bacteriostatic effects of acidified nitrite because of the release of NO and other toxic reactive nitrogen intermediates and also the role of vitamin C in the treatment and prevention of UTI [63]. ...
Article
Full-text available
Background Urinary tract infection (UTI) is a common occurrence in females, during pregnancy, and in peri- and postmenopausal women. UTIs are associated with significant morbidity and mortality, and they affect the quality of life of the affected patients. Antibiotic therapy is an effective approach and reduces the duration of symptoms. Development of resistance, adverse effects of antibiotics, and other associated problems lead to establishing the research framework to find out the alternative approaches in controlling UTIs. Natural approaches have been extensively used for the management of various diseases to improve symptoms and also improve general health. Main body Different databases were employed to identify studies reporting on natural options including herbal medicines, vitamins, trace elementals, sugars, and probiotics without time limitations. Conclusion Herbal medicines can be effective at the first sign of the infection and also for short-term prophylaxis. Using vitamins, trace elementals, and/or sugars is an effective approach in preventing UTIs, and a combination of them with other antibacterial agents shows positive results. Probiotics have great potential for the threat of antibiotic over-usage and the prevalence of antibiotic-resistant microorganisms. This study may be of use in developing the efficient formulation of treatment of UTI.
... Upon swallowing, bio-converted nitrite enters the gastrointestinal tract and reaches peak plasma concentrations at nearly 2.5 h following a nitrate load [98]. Circulating nitrite is sequentially reduced to nitric oxide, a systemic process that can be mediated by a variety of endogenous nitrate reductases [103] including deoxyhemogoblin and ascorbic acid [103][104][105]. This final step in its bio-activation to nitric oxide is pH and oxygen pressure dependent, thus the reaction is greatly enhanced when oxygen levels are reduced, such as in ischemic tissue [106,107]. ...
Article
Full-text available
Objective: We aim to synthesize effects of repeated administration (≥3 days) of inorganic nitrate on blood pressure and arterial stiffness measures. Methods: We conducted a systematic review and meta-analysis of randomized controlled trials with at least 3 days treatment of inorganic nitrate on blood pressure and arterial stiffness in individuals with or without elevated cardiovascular disease risk. MEDLINE, EMBASE and the Cochrane Library were searched through 2 July 2019. Two independent reviewers extracted relevant study data. Data were pooled using the generic inverse variance method with random-effects model, and expressed as mean differences with 95% confidence intervals. Certainty in the evidence was assessed using GRADE. Results: Forty-seven trials were included (n = 1101). Administration of inorganic nitrate significantly lowered SBP [mean difference: -2.91 mmHg, 95% confidence interval (95% CI): -3.92 to -1.89, I = 76%], DBP (mean difference: -1.45 mmHg, 95% CI: -2.22 to -0.68, I = 78%], central SBP (mean difference: -1.56 mmHg, 95% CI: -2.62 to -0.50, I = 30%) and central DBP (mean difference: -1.99 mmHg, 95% CI: -2.37 to -1.60, I = 0%). There was no effect on 24-h blood pressure, augmentation index or pulse wave velocity. Certainty in the evidence was graded moderate for central blood pressure, pulse wave velocity and low for peripheral blood pressure, 24-h blood pressure and augmentation index. Conclusion: Repeated administration (≥3 days) of inorganic nitrate lower peripheral and central blood pressure. Results appear to be driven by beneficial effects in healthy and hypertensive individuals. More studies are required to increase certainty in the evidence.
... On the other hand, the pH meter is a medical device, which has been validated with patients (20,21), designed for urine pH self-monitoring, enabling patients to easily control urine pH on their own and its applicability may be extended to other urological pathologies where urinary pH plays an important role, such as acid-base imbalance diseases, urinary tract infections, cystitis, painful bladder syndrome or stent encrustation (22)(23)(24). ...
Preprint
Full-text available
Background: Encrustation of ureteral double J stents is a common complication that may affect its removal. The aim of the proposed study is to evaluate the efficacy and safety of a new oral composition to prevent double J stent encrustation in indwelling times up to 8 weeks. Methods: A double-blinded, multicenter, placebo-controlled trial was conducted with 105 patients with indwelling double J stents enrolled across 9 public hospitals in Spain. The patients were randomly assigned (1:1) into intervention (53 patients) or placebo (52 patients) groups for 3 to 8 weeks and both groups self-monitored daily their morning urine pH levels. The primary outcome of analysis was the degree of stent ends encrustation, defined by a 4-point score (0 – none; 3 – global encrustation) using macroscopic and electron microscopy analysis of crystals, after 3 to 8-w indwelling period. Score was exponentially transformed according to calcium levels. Secondary endpoints included urine pH decrease, stent removal, and incidence of adverse events. Results: The intervention group benefits from a lower global encrustation rate of stent ends than placebo group (1% vs 8.2%; p < 0.018). Mean encrustation score was 85.12 (274.5) in the placebo group and 18.91 (102.27) in the intervention group (p < 0.025). Considering the secondary end points, treated patients reported greater urine pH decreases (p = 0.002). No differences in the incidence of adverse events were identified between the groups. Conclusions: Our data suggest that the use of this new oral composition is beneficial in the context of ureteral double J indwelling by decreasing mean, as well as global encrustation. Trial registration: This trial was registered at www.clinicaltrials.gov under the name “Combined Use of a Medical Device and a Dietary Complement in Patient Urinary pH Control in Patients With an Implanted Double J Stent” with date 2nd November 2017, code NCT03343275, and URL: https://clinicaltrials.gov/ct2/show/record/NCT03343275?term=NCT03343275&draw=2&rank=1
... On the other hand, the pH meter is a medical device, which has been validated with patients(20,21), designed for urine pH self-monitoring, enabling patients to easily control urine pH on their own and its applicability may be extended to other urological pathologies where urinary pH plays an important role, such as acid-base imbalance diseases, urinary tract infections, cystitis, painful bladder syndrome or stent encrustation (22)(23)(24). ...
Preprint
Full-text available
Background: Encrustation of ureteral double J stents is a common complication that may affect its removal. The aim of the proposed study is to evaluate the efficacy and safety of a new oral composition to prevent double J stent encrustation in indwelling times up to 8 weeks. Methods: A double-blinded, multicenter, placebo-controlled trial was conducted with 105 patients with indwelling double J stents enrolled across 9 public hospitals in Spain. The patients were randomly assigned (1:1) into intervention (53 patients) or placebo (52 patients) groups for 3 to 8 weeks and both groups self-monitored daily their morning urine pH levels. The primary outcome of analysis was the degree of stent ends encrustation, defined by a 4-point score (0 – none; 3 – global encrustation) using macroscopic and electron microscopy analysis of crystals, after 3 to 8-w indwelling period. Score was exponentially transformed according to calcium levels. Secondary endpoints included urine pH decrease, stent removal, and incidence of adverse events. Results: The intervention group benefits from a lower global encrustation rate of stent ends than placebo group (1% vs 8.2%; p
... On the other hand, the pH meter is a medical device, which has been validated with patients [20,21], designed for urine pH self-monitoring, enabling patients to easily control urine pH on their own and its applicability may be extended to other urological pathologies where urinary pH plays an important role, such as acid-base imbalance diseases, urinary tract infections, cystitis, painful bladder syndrome or stent encrustation [22][23][24]. ...
Article
Full-text available
Background: Encrustation of ureteral double J stents is a common complication that may affect its removal. The aim of the proposed study is to evaluate the efficacy and safety of a new oral composition to prevent double J stent encrustation in indwelling times up to 8 weeks. Methods: A double-blinded, multicenter, placebo-controlled trial was conducted with 105 patients with indwelling double J stents enrolled across 9 public hospitals in Spain. The patients were randomly assigned (1:1) into intervention (53 patients) or placebo (52 patients) groups for 3 to 8 weeks and both groups self-monitored daily their morning urine pH levels. The primary outcome of analysis was the degree of stent ends encrustation, defined by a 4-point score (0 - none; 3 - global encrustation) using macroscopic and electron microscopy analysis of crystals, after 3 to 8-w indwelling period. Score was exponentially transformed according to calcium levels. Secondary endpoints included urine pH decrease, stent removal, and incidence of adverse events. Results: The intervention group benefits from a lower global encrustation rate of stent ends than placebo group (1% vs 8.2%; p < 0.018). Mean encrustation score was 85.12 (274.5) in the placebo group and 18.91 (102.27) in the intervention group (p < 0.025). Considering the secondary end points, treated patients reported greater urine pH decreases (p = 0.002). No differences in the incidence of adverse events were identified between the groups. Conclusions: Our data suggest that the use of this new oral composition is beneficial in the context of ureteral double J indwelling by decreasing mean, as well as global encrustation. Trial registration: This trial was registered at www.clinicaltrials.gov under the name "Combined Use of a Medical Device and a Dietary Complement in Patient Urinary pH Control in Patients With an Implanted Double J Stent" with date 2nd November 2017, code NCT03343275, and URL.
... On the other hand, the pH meter is a medical device, which has been validated with patients (20,21), designed for urine pH self-monitoring, enabling patients to easily control urine pH on their own and its applicability may be extended to other urological pathologies where urinary pH plays an important role, such as acid-base imbalance diseases, urinary tract infections, cystitis, painful bladder syndrome or stent encrustation (22)(23)(24). ...
Preprint
Full-text available
Background: Encrustation of ureteral double J stents is a common complication that may affect its removal. The aim of the proposed study is to evaluate the efficacy and safety of a new oral composition to prevent double J stent encrustation in indwelling times up to 8 weeks. Participants, design and intervention: A double-blinded, multicenter, placebo-controlled trial was conducted with 105 patients with indwelling double J stents enrolled across 9 public hospitals in Spain. The patients were randomly assigned (1:1) into intervention (53 patients) or placebo (52 patients) groups for 3 to 8 weeks and both groups self-monitored daily their morning urine pH levels. Outcome measurements and statistical analysis: The primary outcome of analysis was the degree of stent ends encrustation, defined by a 4-point score (0 – none; 3 – global encrustation) using macroscopic and electron microscopy analysis of crystals, after 3 to 8-w indwelling period. Score was exponentially transformed according to calcium levels. Secondary endpoints included urine pH decrease, stent removal, and incidence of adverse events. Results: The intervention group benefits from a lower global encrustation rate of stent ends than placebo group (1% vs 8.2%; p < 0.018). Mean encrustation score was 85.12 (274.5) in the intervention group and 18.91 (102.27) in the placebo group (p < 0.025). Considering the secondary end points, treated patients reported greater urine pH decreases (p = 0.002). No differences in the incidence of adverse events were identified between the groups. Conclusions: Our data suggest that the use of this new oral composition is beneficial in the context of ureteral double J indwelling by decreasing mean, as well as global encrustation. Trial registration: This trial was registered at www.clinicaltrials.gov under the name “Combined Use of a Medical Device and a Dietary Complement in Patient Urinary pH Control in Patients With an Implanted Double J Stent” with date 2nd November 2017, code NCT03343275, and URL: https://clinicaltrials.gov/ct2/show/record/NCT03343275?term=NCT03343275&draw=2&rank=1
... On the other hand, the pH meter is a medical device, which has been validated with patients (20,21), designed for urine pH self-monitoring, enabling patients to easily control urine pH on their own and its applicability may be extended to other urological pathologies where urinary pH plays an important role, such as acid-base imbalance diseases, urinary tract infections, cystitis, painful bladder syndrome or stent encrustation (22)(23)(24). ...
Preprint
Full-text available
Background: Encrustation of ureteral double J stents is a common complication that may affect its removal. The aim of the proposed study is to evaluate the efficacy and safety of a new oral composition to prevent double J stent encrustation in indwelling times up to 8 weeks. Participants, design and intervention: A double-blinded, multicenter, placebo-controlled trial was conducted with 105 patients with indwelling double J stents enrolled across 9 public hospitals in Spain. The patients were randomly assigned (1:1) into intervention (53 patients) or placebo (52 patients) groups for 3 to 8 weeks and both groups self-monitored daily their morning urine pH levels. Outcome measurements and statistical analysis: The primary outcome of analysis was the degree of stent ends encrustation, defined by a 4-point score (0 – none; 3 – global encrustation) using macroscopic and electron microscopy analysis of crystals, after 3 to 8-w indwelling period. Score was exponentially transformed according to calcium levels. Secondary endpoints included urine pH decrease, stent removal, and incidence of adverse events. Results: The intervention group benefits from a lower global encrustation rate of stent ends than placebo group (1% vs 8.2%; p
... On the other hand, the pH meter is a medical device designed for urine pH self-monitoring, enabling patients to easily control urine pH on their own and consequently, adjusting their treatment according to their physician's advice. This device has been validated with patients (20,21) and its applicability may be extended to other urological pathologies where urinary pH plays an important role, such as acid-base imbalance diseases, urinary tract infections, cystitis, painful bladder syndrome or stent encrustation (22)(23)(24). ...
Preprint
Full-text available
Background: Encrustation of ureteral double J stents is a common complication that may affect its removal. The aim of the proposed study is to evaluate the efficacy and safety of a new oral composition to prevent double J stent encrustation in indwelling times up to 8 weeks. Methods: A double-blinded, multicenter, placebo-controlled trial was conducted with 105 patients with indwelling double J stents enrolled across 9 public hospitals in Spain. The patients were randomly assigned (1:1) into intervention or placebo groups for 3 to 8 weeks and both groups self-monitored daily their morning urine pH levels. The primary outcome of analysis was the degree of stent ends encrustation, defined by a 4-point score (0 – none; 3 – global encrustation) using macroscopic and electron microscopy analysis of crystals, after 3 to 8-w indwelling period. Score was exponentially transformed according to calcium levels. Secondary endpoints included urine pH decrease, stent removal, and incidence of adverse events. Results: The intervention group benefits from a lower global encrustation rate of stent ends than placebo group (1% vs 8.2%; p < 0.018). Mean encrustation score was 85.12 (274.5) in the intervention group and 18.91 (102.27) in the placebo group (p < 0.025). Considering the secondary end points, treated patients reported greater urine pH decreases (p = 0.002). No differences in the incidence of adverse events were identified between the groups. Conclusions: Our data suggest that the use of this new oral composition is beneficial in the context of ureteral double J indwelling by decreasing mean, as well as global encrustation. Trial registration: This trial was registered at www.clinicaltrials.gov under the name “Combined Use of a Medical Device and a Dietary Complement in Patient Urinary pH Control in Patients With an Implanted Double J Stent” with date 2 nd November 2017, code NCT03343275, and URL: https://clinicaltrials.gov/ct2/show/record/NCT03343275?term=NCT03343275&draw=2&rank=1 Keywords: double J stent; encrustation; nutraceutical; L-methionine; phytin; pH.
... An increase in NO generation would be expected to augment the antimicrobial of acidified nitrite. Ascorbic acid (vitamin C) has been shown to increase the formation of NO from nitrite 3,4,6 .The main problem with UTI that should be taken into consideration is related to the emergence of new strains of pathogens having multidrug resistance 2 . Nowadays physicians are treating UTI using high doses of antibiotic for long duration, which may impair renal function, particularly in old aged patients. ...
... in this medium, as well as its effect on the production and stability of enzymes and the growth of microorganisms (Carlsson et al., 2001). ...
Thesis
Full-text available
Summary Proteus vulgaris it’s an opportunistic pathogen cause urinary tract infections and wound infections. Urease production, bacterial motility and fimbriae may favor the production of upper urinary tract infections (UTIs). Chondroitinase [EC 4.2.2.4] is an enzyme degrading hyaluronic acid, chondroitin sulfate, Dermatan sulfate or the like into a mixture of an unsaturated disaccharide and oligosaccharide. The enzyme is known to be produced by bacteria such as P. vulgaris. From September 2013 to December 2013, 125 from Al Kadhimiya Teaching Hospital, Baghdad Teaching Hospital and El Shaheed Ghazi al-Hariri Hospital. Samples were collected from patients suffering from UTI. These isolates were identified according to their morphological and biochemical tests, 48 isolates were found belong to genus Proteus with isolation percentage 38%. Proteus vulgaris percentage was 18.7% and Proteus mirables was 81.25 %. The ability of these isolates to produce chondroitinase enzyme was done by using solid media agar contain chondroitin sulfate descripted by Smith and Willett, (1968). P. vulgaris could not utilize chondroitin sulfate on solid medium because the enzyme production was intracellular. Chondroitinase production was induced on broth medium contain chondroitin sulfate descripted by Yamagata et al., (1986). All strain showed the ability to produce chondroitinase. P (8) strain showed the highest enzyme activity about 150 (unit/ ml), further detection was done by test extracted bacterial on solid medium contain chondroitinase. Optimum condition for production of chondroitinase was 37° C, pH media 8.0, incubation time 2 day. Extracted chondroitinase showed high ability to lose it activity after 24 hour even when kept at 4° C and -20° C. EDTA and glycerol addition to harvested cells (before ultra-sonication process) showed the ability to kept enzyme activity at 4° C and -20° C for weeks. The chondroitinase ABC of P. vulgaris P (8) strain was extracted and purified by precipitation with 60% saturation of ammonium sulfate, dialysis and filtrated with Sephadex 6B column. Optical density at 280 for fractions showed five peaks. The specific activities was reached 5000 (Unit/mg) at fourths peak, fraction (53-57) with 35.2% yield and 17.4 fold of purification. Purified chondroitinase has molecular weight 111254 Dalton. Optimum pH and temperature was 8.0, 37° C respectively. Chondroitinase lose it activity at alkaline and acidic pH more or lowest than (7-8) pH, also it loose its activity rapidly above 40° C. Chondroitinase activity was inhibited significantly with increasing antibiotics concentrations the remaining activity of the treated enzyme with 75µg/ml of ampicillin, amoxicillin, ciprofloxacin and cefotaxime was 58, 45, 63 and 61 % respectively while it showed in cephalexin increase in enzyme activity to 127% at 75 µg/ml. Effect of metal ions solution on chondroitinase showed increasing in enzyme activity to 120 % and 113 % when enzyme incubated with Mgcl2, Cacl2 respectively, while NH4cl , Kcl recode reduce in remain activity 95% , 86% respectively. Effect of chondroitinase on human cartilage showed the ability of enzyme to degraded the cartilage, turbidity was observed as a result of incubation the cartilage. Also optical density at 232nm was measured and showed increase gradually with a direct proportion relationship with incubation time for 14 day at 37°C Histopathology study of chondroitinase on animal tissue (mice) it showed ability of enzyme to reach different organ and knee cartilage when injection intraperitoneally, it cause necrosis and congestion with inflammation cell in heart, liver and kidney. While in knee cartilage it showed inflammation cell on synovial membrane and decalcification of bone. In direct injection with chondroitinase in knee cartilage it showed more change in tissue than intraperitoneally injection, necrosis, congestion, oedema and decalcification of bone that’s lead to formation abnormal bone.
... The antioxidant effect of this herb was proved in a study by Czyzowska et al. (2015) from the Netherlands. saprophyticus were assessed in a study conducted in Sweden, and concurrent use of nitrite and vitamin C was found to have a significant effect on inhibiting these UTI-causing bacteria (Carlsson, Wiklund, Engstrand, Weitzberg, & Lundberg, 2001). Castello, Girona, Gomez, Mur, and Garcia (1996) investigated the prophylactic effect of ascorbic acid 2 g daily on the prevention of UTI in people with spinal injury and showed that ascorbic acid had no significant effect on the prevention of UTI in these patients, which disagrees with the present study. ...
Article
Urinary tract infection (UTI) is an infection that can occur in any area of the urinary tract which is characterized by a positive urine culture (U/C). The risk of UTI following cesarean section (CS) increases due to procedures such as catheterization. In vitro studies have demonstrated the effect of Rosa canina fruit in preventing Escherichia coli growth. This study was conducted to determine the effect of R. canina fruit in preventing the incidence of UTI in women following CS. This triple-blind randomized clinical trial was conducted in 2016 on 400 women following CS with negative U/C in Alzahra and Taleghani educational hospitals in the city of Tabriz-Iran. Participants were assigned into two groups of 200 women using block randomization. Each group received a twice daily dose of 500 mg capsules containing R. canina or placebo from the second day after CS for 20 days. Women were assessed by U/C on the 7th–10th and 20th days following CS. UTI was significantly lower in the intervention group compared with the control in the follow-ups conducted on the 7th–10th days (odds ratio = 0.22; confidence interval 95% [0.07, 0.67]; p = .006) and 20th day (odds ratio = 0.32; confidence interval 95% [0.14, 0.75]; p = .008). But the incidence of cystitis in the two groups was not statistically significant (p > .05). R. canina fruit capsules were able to reduce the incidence of UTI after CS. Thus, it is likely that administration of this medication can promote maternal health following CS.
... It also works synergistically with MH, providing an acidic environment to aid conversion to formaldehyde (64). Finally, there is data to suggest that vitamin C can also have a bacteriostatic effect within urine, mediated by reduction of urinary nitrites to reactive nitrogen oxides (65,66). ...
Article
One in three women will experience a clinically significant urinary tract infection (UTI) by age twenty-four and almost half will have at least one in their lifetime. Recurrent UTIs (rUTIs) are defined as having greater than two infections in a 6-month period, or three infections over twelve months, with complete resolution for at least two weeks. These may be due to relapse from incomplete treatment (persistence) or re-infection (new source). It may be difficult to distinguish between the two, where the same organism is cultured. There are several risk factors for rUTIs including an impairment of the body's immune system and virulence factors. Reversible or treatable causes are sought and excluded in the patient's initial review. Patient's with rUTI are often complex and difficult to manage. The long-term management options in women are multimodal and should focus on prevention of relapse and recurrence. Behavioural factors include adequate hydration, care with sexual hygiene, reducing one's body mass index (BMI) and post-void residual (PVR) volume. There are several non-antimicrobial options for rUTIs which have become a multi-billion-dollar business. Unfortunately, there are numerous studies which fail to show any major benefit or having conflicting data. Vaccines are currently being explored as a prevention strategy, delivered through injection, intra-nasal sprays, or vaginal suppositories, which are made from combinations of heat killed uro-pathogenic strains. There are no widely available vaccines at present due to limited clinical success. It is well established that appropriate antibiotic therapy results in higher rates of symptom relief and bacterial eradication in women with uncomplicated cystitis. There are several options for antimicrobial use which have been shown to be highly effective in reducing the risk of rUTI in women. The pain and discomfort of the UTI must be balanced with the cost and risk of developing resistance when using antimicrobials. Continuous prophylaxis, pre- and postcoital voiding, and self-starting are the three commonly accepted options for prophylaxis. The choice between these will depend upon patient preference, cultures and previous pattern of infection. Intra-vesical instillation of hyaluronic acid and chondroitin sulphate have been used for glycosaminoglycan (GAG) layer replenishment for many indications, including interstitial cystitis, overactive bladder syndrome, radiation cystitis and prevention of rUTI. At present, intra-vesical therapies are reserved for only those with the most unresponsive rUTIs. The principles of treating rUTI are to break the cycle and to treat any reversible causes. With our ever-expanding research knowledge, there are now many useful products that may be used for the successful treatment of rUTI. A management plan including a combination of a non-antimicrobial and selective antimicrobial regime for a minimum of six months should be considered. It is a prudent clinician that clearly defines this management plan, with reassurance of a finite period of therapy.
... 25,26 However, AA has been suggested to prevent UTI in experimental studies by pH-dependent and pH-independent mechanisms. 27,28 We speculate that AA consumption in both groups could lead to a global reduction in UTI incidence. ...
Article
Introduction: Kidney transplant (KT) recipients are highly susceptible to urinary tract infections (UTIs). Few data are available regarding the effects of cranberry products in KT recipients. Subjects and methods: This was a randomized double-blind study comparing daily treatment with a cranberry capsule versus a placebo capsule for 6 months. The study protocol was conducted in accordance with the Helsinki Declaration and was approved by the local ethics committee. The study group comprised women undergoing KT for over 1 year, and the exclusion criteria included simultaneous antibiotic UTI prophylaxis. The primary endpoint was UTI occurrence, and the secondary endpoints were the time to first UTI, hospitalization, antibiotic resistance, graft function, immunosuppressive dose changes, and side effects. Results: 55 KT women at a single centre were included as follows: 25 were randomized to the cranberry group, and 30 were assigned to the placebo group. During the study period, 16.4% of the participants presented with a UTI. A diagnosis of UTI within the previous year and KT duration, but not cranberry use (p=0.95) were independent predictors of UTI. Discussion: Cranberry capsules appear to be safe in KT women; however, no clinical efficacy of UTI prophylaxis was demonstrated in this well-designed but underpowered study. Further multicentric and longer trial should be performed to determine cranberry clinical efficacy
... In parallel with the effects that we found with TEMPOL, reducing agents such as polyphenols caused nitrite to generate predominantly NO [14,34]. We found that treatment with TEMPOL was associated with greater increases in plasma nitrite and lower increases in plasma nitrate after 5 or 15 mg of sodium nitrite, possibly as a result of a more reductive gastric environment in the presence of TEMPOL. ...
Article
Orally administered nitrite exerts antihypertensive effects associated with increased gastric nitric oxide (NO) formation. While reducing agents facilitate NO formation from nitrite, no previous study has examined whether antioxidants with reducing properties improve the antihypertensive responses to orally administered nitrite. We hypothesized that TEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) could enhance the hypotensive effects of nitrite in hypertensive rats by exerting antioxidant effects (and enhancing NO bioavailability) and by promoting gastric nitrite-derived NO generation. The hypotensive effects of intravenous and oral sodium nitrite were assessed in unanesthetized freely-moving rats with L-NAME (Nω-nitro-L-arginine methyl ester; 100mg/kg; p.o.)-induced hypertension treated with TEMPOL (18mg/kg; p.o.) or vehicle. While TEMPOL exerted antioxidant effects in hypertensive rats, as revealed by lower plasma 8-isoprostane and vascular reactive oxygen species levels, this antioxidant did not affect the hypotensive responses to intravenous nitrite. Conversely, TEMPOL enhanced the dose-dependent hypotensive responses to orally administered nitrite, and this effect was associated with higher increases in plasma nitrite and lower increases in plasma nitrate concentrations. In vitro experiments using electrochemical and chemiluminescence NO detection under variable pH conditions showed that TEMPOL enhanced nitrite-derived NO formation, especially at low pH (2.0 to 4.0). TEMPOL signal evaluated by electron paramagnetic resonance decreased when nitrite was reduced to NO under acidic conditions. Consistent with these findings, increasing gastric pH with omeprazole (30mg/kg; p.o.) attenuated the hypotensive responses to nitrite and blunted the enhancement in plasma nitrite concentrations and hypotensive effects induced by TEMPOL. Nitrite-derived NO formation in vivo was confirmed by using the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (C-PTIO), which blunted the responses to oral nitrite. Our results showed that TEMPOL promotes nitrite reduction to NO in the stomach and enhanced plasma nitrite concentrations and the hypotensive effects of oral sodium nitrite through mechanisms critically dependent on gastric pH. Interestingly, the effects of TEMPOL on nitrite-mediated hypotension cannot be explained by increased NO formation in the stomach alone, but rather appear more directly related to increased plasma nitrite levels and reduced nitrate levels during TEMPOL treatment. This may relate to enhanced nitrite uptake or reduced nitrate formation from NO or nitrite.
Conference Paper
The relevance of pH assessment in clinical analysis, environmental and industrial control, has raised the demand for the development of portable, low cost and easy-to-use monitoring systems. This paper proposes a pH sensor printed on a paper support passivated with a solid-ink coating. The sensor exploits the pH sensitivity of a reduced graphene oxide functionalized with 3-(4-aminophenil)propionic acid. The sensor responded in the pH range [4], [10] and had a sensitivity of 46 mV/pH. Tests on human plasma and seawater proved this pH sensor to have similar performances than those of a commercial pH-meter with an uncertainty of 0.1 and 0.2 pH unit in plasma and seawater, respectively.
Chapter
Urinary catheters are one of the most commonly utilized medical devices worldwide. They are used in virtually every healthcare setting and contribute to improvements in patient care. While urinary catheters provide invaluable aid to patients, they are not without complications, the most notable being catheter-associated urinary tract infections (CAUTI). Once a urinary catheter is in place, pathogens may migrate to the bladder one of two ways, through the catheter lumen or extraluminally in the periurethral space. In vitro methods are useful tools for predicting clinical efficacy only if they accurately model the most important factors contributing to a clinical infection. Although in vitro methods can’t replace in vivo scenarios, outcomes may be improved as experiments approximate relevant criteria including the biofilm phenotype, time, media type, materials similarities and environment.
Thesis
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Inorganic nitrate (NO3-) and nitrite (NO2-) have generally been considered stable inactive end products of nitric oxide (NO) or unwanted residues in the food chain. While several recent studies surprisingly demonstrate that nitrite is reduced to bioactive NO in blood and tissues, the more stable anion nitrate is still considered to be inert. We investigated if inorganic nitrate could be bioactivated in vivo to generate nitrite systemically. After oral intake and absorption, nitrate is concentrated in saliva, where much is reduced to nitrite by oral bacterial nitrate reductases. We show here that systemic nitrite levels increase greatly after oral nitrate intake, demonstrating for the first time that nitrate is in fact a substrate for systemic generation of nitrite and eventually NO. We show that oral bacteria and the entero-salivary recirculation of nitrate play a major role in the in vivo bioactivation of nitrate. In addition to this major prokaryotic pathway, we discovered a mammalian functional nitrate reductase (eukaryotic pathway) that also regulates nitrite and NO homeostasis. Subsequent studies have confirmed robust physiological effects of dietary nitrate, all of which are compatible with generation of NO. These include a lowering of blood pressure and inhibition of oxygen consumption in humans, and protection against ischaemia reperfusion injury and reversal of metabolic syndrome in animals. This has made us speculate that the strong cardioprotective effects of a diet rich in vegetables, at least partly is explained by the high nitrate content in these foodstuffs. Differently from inorganic nitrate, organic nitrates such as glyceryl trinitrate (GTN) are generally recognized to act via NO donation, and these drugs have been used in the treatment of cardiovascular disorders for >100 years. Despite this long history, their metabolism is still a matter of debate. It is known that liver first pass metabolism can strongly affect their disposition and activity. Thus a careful investigation of the hepatic metabolism is crucial for compounds designed for oral administration. In the second part of this project the liver metabolism of a novel class of hybrid organic nitrates, the nitrooxybutyl-esters derivatives of anti-inflammatory or anti-oxidant compounds was investigated and compared with the prototypic organic nitrate GTN. These compounds are claimed to retain the properties of the parent compound with increased, NO-related, safety and tolerability. It was shown that nitrooxybutyl-ester derivatives are rapidly cleaved in vitro in liver fractions to their parent compounds and the organic nitrate moiety nitrooxybutyl alcohol (NOBA). As for GTN, NOBA is mainly denitrated by the glutathione S-transferase through a clearance based mechanism, i.e. direct metabolism to NOx (nitrite + nitrate) with no main acute bioactivation to NO. The NOx generated during first passage could therefore contribute to the “NO related” effect of organic nitrates when given orally. Moreover, since NOBA is only slowly denitrated in the liver in vitro it might have the potential to partly survive first passage metabolism and be bioactivated to NO in other tissues. A complete picture of the metabolic profile of this class of organic nitrates in different tissues will help to facilitate development of more powerful and selective drugs in different therapeutic areas. In conclusion the results of the present thesis laid the bases that reversed the status of inorganic nitrate from inert end product of NO metabolism to important reservoir of NO. It follows that also the nitrate and nitrite generated from organic nitrate metabolism might play an important role in the final biological effect of these molecules.
Article
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Article
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Recent studies of hydrogen sulfide (H2S) signaling implicate low molecular weight (LMW) thiol persulfides and other reactive sulfur species (RSS) as signaling effectors. Here, we show that a CstR protein from the human pathogen Enterococcus faecalis (E. faecalis), previously identified in Staphylococcus aureus (S. aureus), is an RSS-sensing repressor that transcriptionally regulates a cst-like operon in response to both exogenous sulfide stress and Angeli’s salt, a precursor of nitroxyl (HNO). E. faecalis CstR reacts with coenzyme A persulfide (CoASSH) to form interprotomer disulfide and trisulfide bridges between C32 and C61’ which negatively regulate DNA binding to a consensus CstR DNA operator. A ΔcstR strain exhibits deficiency in catheter colonization in a catheter-associated urinary tract infection (CAUTI) mouse model suggesting sulfide regulation and homeostasis is critical for pathogenicity. Cellular polysulfide metabolite profiling of sodium sulfide-stressed E. faecalis confirms an increase in both inorganic polysulfides and LMW thiols and persulfides sensed by CstR. The cst-like operon encodes two authentic thiosulfate sulfurtransferases and an enzyme we characterize here as an NADH and FAD-dependent coenzyme A (CoA) persulfide reductase (CoAPR) that harbors an N-terminal CoA disulfide reductase (CDR) domain and a C-terminal rhodanese homology domain (RHD). Both cysteines in the CDR (C42) and RHD (C508) domains are required for CoAPR activity and complementation of a sulfide-induced growth phenotype of a S. aureus strain lacking cstB, encoding a non-heme FeII persulfide dioxygenase. We propose that S. aureus CstB and E. faecalis CoAPR employ orthogonal chemistries to reduce CoASSH that accumulates under conditions of cellular sulfide toxicity and signaling.
Article
Background: Nitric oxide (NO), a potent signaling molecule, is known to inhibit platelet (PLT) function in vivo. We investigated how the levels of NO and its metabolites change during routine PLT storage. We also tested whether the material of PLT storage containers affects nitrite content since many plastic materials are known to contain and release nitrite. Study design and methods: For nitrite and nitrate measurement, leukoreduced apheresis PLTs and concurrent plasma (CP) were collected from healthy donors using a cell separator. Sixty-milliliter aliquots of PLT or CP were stored in CLX or PL120 Teflon containers at 20 to 24°C with agitation and daily samples were processed to yield PLT pellet and supernatant. In a separate experiment, PLTs were stored in PL120 Teflon to measure NO generation using electron paramagnetic resonance (EPR). Results: Nitrite level increased markedly in both PLT supernatant and CP stored in CLX containers at a rate of 58 and 31 nmol/L/day, respectively. However, there was a decrease in nitrite level in PLTs stored in PL120 Teflon containers. Nitrite was found to leach from CLX containers and this appears to compensate for nitrite consumption in these preparations. Nitrate level did not significantly change during storage. Conclusion: PLTs stored at 20 to 24°C maintain measurable levels of nitrite and nitrate. The nitrite decline in nonleachable Teflon containers in contrast to increases in CLX containers that leach nitrite suggests that it is consumed by PLTs, residual white blood cells, or red blood cells. These results suggest NO-related metabolic changes occur in PLT units during storage.
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Background: Omadacycline, an aminomethylcycline, was approved in 2018 for the treatment of acute bacterial skin and skin structure infections and community-acquired bacterial pneumonia. In a Phase Ib study, around 34% of the absorbed dose of omadacycline was shown to be excreted in urine-an important property for urinary tract infection (UTI) treatment. Therefore, omadacycline has been studied in two Phase II trials for the treatment of uncomplicated UTIs and acute pyelonephritis. The activity of omadacycline against UTI pathogens in human urine is important to understand in this context. Objectives: To study the in vitro activity of omadacycline against UTI pathogens in human urine supplemented with calcium and magnesium. Methods: Omadacycline activity was compared with that of levofloxacin against the urinary pathogens Escherichia coli, Klebsiella pneumoniae and Staphylococcus saprophyticus in standard medium, pooled normal human urine and neutral pH-adjusted pooled normal human urine spiked with calcium or magnesium at concentrations consistent with hypercalcaemia and hypermagnesaemia. Results: The activities of omadacycline and levofloxacin against these urinary pathogens were lower in urine relative to standard medium; addition of Mg2+ to broth and urine had a further negative impact on omadacycline activity, whereas the addition of Ca2+ had less of an impact. Levofloxacin activity was not substantially reduced in either broth or urine by the addition of divalent cations. Conclusions: The activity of omadacycline against UTI organisms was lower in urine relative to standard medium and was negatively impacted by magnesium. Omadacycline displayed slightly reduced activity when excess calcium was present, but, overall, the differences were ≤2-fold. These observations should be considered along with the pharmacokinetics of the agent for clinical context.
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Background Vitamin C may enhance nitric oxide (NO) production through stepwise reduction of dietary nitrate (NO3) to nitrite (NO2) to NO. The combined effect of vitamin C and NO3 supplementation is relatively unexplored in untreated hypercholesterolemia. Aims We aimed to examine whether co-administration of vitamin C and nitrate for 4-weeks would improve endothelial function (primary outcome), plasma NO metabolites, oxidative stress, and blood lipids (secondary outcomes). Methods Subjects 50–70 years of age with low density lipoprotein (LDL) > 130 mg/dL and RHI ≤2 were enrolled in this randomized double-blind crossover study. Subjects were assigned to two 4-week supplementation treatments starting with 70 ml of concentrated beetroot juice (CBJ) with 1000 mg of vitamin C (NC) or CBJ with matched placebo (N), then switched to alternate treatment following 2-week washout. The change in reactive hyperemia index (RHI), sum of plasma NO metabolites (NO2 + NO3 (NOx)), oxidized LDL (oxLDL), and serum lipids were assessed at baseline and at 4-weeks of each treatment period. Results Eighteen subjects (11M:7F) completed all study visits. No significant treatment differences were observed in RHI change (N: 0.21 ± 0.12; NC: 0.20 ± 0.17; p = 0.99). Secondary analysis revealed that a subgroup of NC subjects who started with a baseline RHI of <1.67 (threshold value for ED) had greater improvements in RHI compared to subjects with RHI >1.67 (1.23 ± 0.15 to 1.96 ± 0.19; n = 8 vs. 1.75 ± 0.11 to 1.43 ± 0.10; n = 8; p = 0.02). Compared to N, NC experienced a significant increase in plasma NOx (N: 94.2 ± 15.5 μmol/L; NC: 128.7 ± 29.1 μmol/L; p = 0.01). Although there was no significant difference in oxLDL change between treatments (N: −1.08 ± 9.8 U/L; NC: −6.07 ± 9.14 U/L; p = 0.19), NC elicited significant reductions in LDL (N: 2.2 ± 2; NC: −10.7 ± 23; p = 0.049), triglycerides (N: 14.6 ± 43; NC: −43.7 ± 45; p = 0.03), and no change in serum high density lipoprotein. Within treatment group comparisons showed that only NC reduced oxLDL significantly from baseline to 4 weeks (p = 0.01). Conclusions No between intervention differences were observed in RHI. RHI only improved in NC subjects with ED at intervention baseline. Four weeks of NC enriched the NO pool and promoted reduction of blood lipids and oxidative stress in subjects with hypercholesterolemia. These preliminary findings highlight a supplementation strategy that may reduce the progression of atherosclerotic disease and deserves further attention in studies using flow mediated dilation methods. Clinical trial registration www.clinicaltrials.gov (NCT04283630).
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Helicobacter pylori resists gastric acidity by modulating the proton-gated urea channel UreI, allowing for pHout-dependent regulation of urea access to intrabacterial urease. We employed pH- and Ca2+-sensitive fluorescent dyes and confocal microscopy to determine the location, rate, and magnitude of pH changes in an H. pylori-AGS cell coculture model, comparing wild-type bacteria with nonpolar ureI-deletion strains (ureI-ve). Addition of urea at pH 5.5 to the coculture resulted first in elevation of bacterial periplasmic pH, followed by an increase of medium pH and then pH in AGS cells. No change in periplasmic pH occurred in ureI-deletion mutants, which also induced a slower increase in the pH of the medium. Pretreatment of the mutant bacteria with the detergent C12E8 before adding urea resulted in rapid elevation of bacterial cytoplasmic pH and medium pH. UreI-dependent NH3 generation by intrabacterial urease buffers the bacterial periplasm, enabling acid resistance at the low urea concentrations found in gastric juice. Perfusion of AGS cells with urea-containing medium from coculture at pH 5.5 did not elevate pHin or [Ca2+]in, unless the conditioned medium was first neutralized to elevate the NH3/NH4+ ratio. Therefore, cellular effects of intrabacterial ammonia generation under acidic conditions are indirect and not through a type IV secretory complex. The pHin and [Ca2+]in elevation that causes the NH3/NH4+ ratio to increase after neutralization of infected gastric juice may contribute to the gastritis seen with H. pylori infection.
Article
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High values (800-6000 parts per billion) of nitric oxide (NO) in expelled air from the stomach were shown in humans by chemiluminescence technique. These NO values were more than 100 times higher than those found in orally exhaled air. Intragastric NO production is probably non-enzymatic, requiring an acidic environment, as NO in expelled air was reduced by 95% after pretreatment with the proton pump inhibitor omeprazole. Furthermore, large amounts of NO were formed in vitro from lettuce and saliva when placed in hydrogen chloride (pH < 2). In conclusion, large amounts of NO are formed intragastrically in humans and this source of NO may be of importance for the integrity of the gastric mucosa in health and disease. Measurements of NO in expelled air might be of value as a non-invasive method for estimation of gastric acidity.
Article
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The role of reactive nitrogen intermediates (RNI) such as nitric oxide (.NO) in host defense against pyogenic microorganisms is unclear, and the actual interactive effect of RNI and reactive oxidative intermediates (ROI) for microbial killing has not been determined. Since, in nature, ROI and RNI might be generated together within any local infection, we evaluated the separate and interactive effects of .NO and O2- on staphylococcal survival by using a simplified system devoid of eukaryotic cells. These studies showed that prolonged exposure of staphylococci to .NO does not result in early loss of viability but instead is associated with a dose-related delayed loss of viability. This effect is abrogated by the presence of hemoglobin, providing further evidence that the effect is RNI associated. Superoxide-mediated killing also is dose related, but in contrast to RNI-mediated killing, it is rapid and occurs within 2 h of exposure. We further show that the interaction of .NO and O(2)- results in decreased O(2)--mediated staphylococcal killing at early time points. .NO, however, appears to enhance or stabilize microbial killing over prolonged periods of incubation. This study did not produce evidence of early synergism of ROI and RNI, but it does suggest that .NO may contribute to host defense, especially when ROI-mediated killing is compromised.
Article
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It has been suggested that dietary nitrate, after concentration in the saliva and reduction to nitrite by tongue surface bacteria, is chemically reduced to nitric oxide (NO) in the acidic conditions of the stomach. This study aimed to quantify this in humans. Ten healthy fasting volunteers were studied twice, after oral administration of 2 mmol of potassium nitrate or potassium chloride. Plasma, salivary and gastric nitrate, salivary and gastric nitrite, and gastric headspace NO concentrations were measured over six hours. On the control day the parameters measured varied little from basal values. Gastric nitrate concentration was 105.3 (13) mumol/l (mean (SEM), plasma nitrate concentration was 17.9 (2.4) mumol/l, salivary nitrate concentration 92.6 (31.6) mumol/l, and nitrite concentration 53.9 (22.8) mumol/l. Gastric nitrite concentrations were minimal (< 1 mumol/l). Gastric headspace gas NO concentration was 16.4 (5.8) parts per million (ppm). After nitrate ingestion, gastric nitrate peaked at 20 minutes at 3430 (832) mumol/l, plasma nitrate at 134 (7.2) mumol/l, salivary nitrate at 1516.7 (280.5) mumol/l, and salivary nitrite at 761.5 (187.7) mumol/l after 20-40 minutes. Gastric nitrite concentrations tended to be low, variable, and any rise was non-sustained. Gastric NO concentrations rose considerably from 14.8 (3.1) ppm to 89.4 (28.6) ppm (p < 0.0001) after 60 minutes. All parameters remained increased significantly for the duration of the study. A very large and sustained increase in chemically derived gastric NO concentrations after an oral nitrate load was shown, which may be important both in host defence against swallowed pathogens and in gastric physiology.
Article
THE search for effective urinary antibacterial agents is never ending. Recent reports propose dl-methionine as an effective agent for urinary antisepsis. It has been said to lower urinary pH and promote excretion of bacteriostatic substances.1 , 2 About 40 per cent of the methionine given orally has been accounted for as increased urinary sulfate. Free methionine and aminobutyric acid are also excreted in the urine.3 Escherichia coli, streptococcus, pseudomonas, proteus and staphylococcus species of common urinary pathogens have been found to metabolize and require methionine.4 5 6 7 8 The d-isomer is not utilized for growth by these bacteria. Also, d-aminobutyric acid, a metabolite of . . .
In a discussion on the persistence with which attempts to dislodge the colon bacillus from the urinary tract are defeated, Barborka reminded me that the urine of patients who are receiving a diet high in fat and low in carbohydrate is considerably acidified. Shohl and Janney,1 a number of years ago, demonstrated that growth of colon bacilli was inhibited in urine of which the acidity was as great as pH 4.6 or 5.0, or alkalinity as much as pH 9.2 or 9.6.In October, I2 reported the results in two cases of bacilluria of Escherichia coli type, in which the ketogenic diet was given. Since that time, the treatment has been applied in several additional cases of persistent bacilluria, and the patients have been under observation long enough to allow the results to be evaluated.In the accompanying table is a summary. Several notable features emerged
Article
Thesis (Med. Sci. D.)--Columbia University. College of Physicians and Surgeons, 1941. Offprint: The journal of urology. Vol. 44, no. 5 (Nov. 1940). Includes bibliographical references.
Article
Ihe effect of ascorbic acid on urine pH was studied in 10 healthy male volunteers. Ascorbic acid tablets were given to each subject at two dosage levels and intervals (4 and 6 g/day divided into four and five doses). A crossover study design was used in which each of the four regimens was in simultaneous use in one of four subgroups. Each urine volume voided in a day was collected in a separate polyethylene bag, refrigerated and the pH measured at one time daily. After two days of ascorbic add therapy, the pH of each urine sample was determined during the remaining three, daJra of the regimen. A two-day "washout" period was followed between regimens. The four ascorbic acid treatments did lower the urine pH of all subjects combined, but the greatest effect was only 0.24 pH unit The 4-g/day (four doses) treatment resulted in a mean urine pH significantly different from either pretreatment or posttreatment controls; the 6-g/day (four doses) treatment was not different from either control period; and the other two treatments were significantly different from only the pretreatment control. It does not seem warranted to recommend ascorbic acid, in any dose or regimen, as a urinary acidifier.
Article
The recent popularization of self-prescribed large doses of vitamin C has increased the possibility for erroneous conclusions to be drawn from standard clinical methods used in urinary glucose monitoring, due to interference with these methods by the greatly elevated excretion of vitamin C. The coupled-enzym–echromogen strip tests showed erroneously negative glucose levels in urines of both a diabetic individual and a subject with a genetic low renal threshold for glucose when they were supplementing their normal diets with 1–2 g vitamin C per day. With this regimen, their urinary vitamin C levels reached 200 mg/dl (11.4 mmol/l). For normal urine with vitamin C added, false-positive tests for glucose were found using Benedict’s reagent when vitamin C was present at 250 mg/dl (14.3 mmol/l) or higher concentrations. In diabetic individuals consuming large quantities of vitamin C, this interference with standard coupled-enzyme–chromogen strip tests or Benedict’s test could present a significant problem in diagnosis and clinical management of the disease. A simple anion exchange method of treating the urine was used to correct the false results.
Article
Urinary nitrite, a precursor of carcinogenic nitrosamines, and nitrate were measured in 73 Japanese patients with urinary tract infections (UTI) and in two control groups. Nitrite was detected in 12% of patients with uncomplicated UTI (226 +/- 161 mumol/l) and in 42% of those with complicated UTI (375 +/- 297 mumol/l). None of the subjects in the control groups excreted detectable amounts of nitrite. The excretion of nitrite in four out of five nitrite excretors continued for at least 14-55 days. The concentrations of urinary nitrite were significantly correlated with those of nitrate- plus nitrite-nitrogen, which reflect dietary nitrate ingestion. The results suggest the importance of close monitoring of nitrite excretor group in UTI patients, to clarify the mechanism of the association between UTI and bladder cancer.
Article
Ascorbic acid and ascorbate ion (denoted together as ASC) inhibit nitrosation by competing for the nitrosating agents formed from nitrite (e.g. N2O3, NO+ and NOSCN). ASC is oxidized irreversibly by this reaction and the nitrite equivalents are reduced to nitric oxide (NO). In open, aerobic systems the effective stoichiometry of the reaction between ASC and nitrite is not fixed, but is determined by a competition between the physical removal of NO (and NO2) from the system and the oxidation of NO by dissolved O2. The oxidation of NO reconverts it to a nitrosating agent which may react again with the remaining ASC. To determine the conditions under which ASC is most effective as a nitrosation inhibitor, we examined the kinetics of the reactions between nitrite and ASC and between nitrite and proline. These reactions were studied in open shaker flasks as functions of pH, anion composition (SCN- and Cl-), temperature, and gas-liquid mass transfer rate. At lower mass transfer rates, at lower pH and/or in the presence of SCN- or Cl-, relatively more ASC was consumed by a given amount of nitrite. Increased temperature caused more or less ASC to be consumed by a given amount of nitrite, depending on the conditions. A mathematical model of the reactions and mass transfer steps was developed which describes each of these features. The model predicts the variable stoichiometry of the reaction between nitrite and ASC in open, aerobic systems, and clarifies the mechanisms by which ASC inhibits nitrosation.
Article
The acidifying effect of intravenous (IV) ascorbic acid was studied in seven healthy adult volunteers. After obtaining baseline urine and blood samples, 2-g IV doses of ascorbic acid were administered to each subject during a 20-minute period. Venous blood samples were obtained at times 0.5, 1, and 2 hours, and urine was collected at times 0.5, 1, 2, and 3 hours. Our results show that venous blood pH, plasma bicarbonate concentration, urine PCO2, and urine bicarbonate excretion did not change significantly during the study period. Urinary titratable acidity, ammonium excretion, and net hydrogen ion excretion decreased, and urinary pH actually showed a significant rise at two hours. We therefore conclude that IV ascorbic acid administered in recommended doses does not effectively acidify urine.
Article
The reactive nitrogen intermediate (RNI) nitric oxide (NO.) is formed from L-arginine by an NO. synthase and, following secondary reactions yielding additional toxic intermediates, nitrite (NO2-) and nitrate are formed. Nitrite, however, also has toxic properties. At acid pH, nitrous acid (HNO2) is bactericidal to Escherichia coli, in association with the loss of HNO2/NO2- and the uptake of oxygen, an effect which is increased by H2O2. Under conditions in which HNO2/NO2- +/- H2O2 were ineffective, the further addition of peroxidase (myeloperoxidase [MPO], eosinophil peroxidase, lactoperoxidase) or catalase resulted in bactericidal activity and the disappearance of HNO2/NO2-. Paradoxically, HNO2/NO2- also inhibited the bactericidal activity of MPO by the formation of a complex with MPO with a shift in the absorption spectrum, and by reaction with hypochlorous acid (HOCl) (the product of the chloride-supplemented MPO-H2O2 system), with loss of the bactericidal activity of HOCl and the disappearance of both HOCl and HNO2/NO2- from the reaction mixture. Thus, HNO2/NO2-, rather than being solely an end product of RNI formation, may influence antimicrobial activity either by acting alone, with H2O2, or with H2O2 and peroxidase as a source of toxic agents, or by inhibiting the peroxidase-mediated antimicrobial systems.
Article
The past decade has witnessed a veritableexplosionof interest in the simplemolecule nitric oxide (NO) as a vasodilator, neurotransmitter, and antimicrobial agent. NO and other reactive nitrogen intermediates exhibit cytostatic or cytocidal activity against a remarkable breadth of pathogenic microorganisms. Mammalian cells, including human cells, produce nitric oxide both constitutively and inducibly in response to inflammatory stimuli. This review will provide a brief overview of current knowledge regarding the antimicrobial activity of NO and the possible importance of this activity in infection, particularly with regard to intracellular pathogens.
Article
The bacteriostatic gas nitric oxide (NO) is formed when nitrite is acidified. Infected urine may contain considerable amounts of nitrite as a result of bacterial nitrate reductase activity, and detection of nitrite in urine is routinely used in the diagnosis of bacterial cystitis. We sought to determine whether NO was generated from acidified nitrite-containing urine. Furthermore, we also studied the growth of the urinary pathogen Escherichia coll in acidified nitrite-containing urine. Urine, collected from healthy control subjects or from patients with infected nitrite-containing urine, was acidified and incubated in a closed syringe with varying amounts of nitrite added. After 30 minutes, the headspace gas was removed and immediately injected into a chemiluminescence NO analyzer. In addition, NO was measured in urine collected from healthy control subjects after ingestion of vitamin C. Bacterial growth was measured continuously in control urine for 10 hours after incubation for 2 hours in acidic urine with varying concentrations of nitrite added. Large amounts of NO were released from infected nitrite-containing urine after mild acidification. NO was also released from acidified control urine if nitrite was added, and this release was greatly potentiated in the presence of vitamin C. Furthermore, the growth of E. coli was markedly reduced by the addition of nitrite to acidified urine. We propose that nitrite-producing bacteria induce their own death in acidic urine by supplying substrate for generation of bacteriostatic compounds such as NO. This mechanism might explain why urinary acidification and vitamin C may be effective in the treatment of bacteriuria.
Article
Death from chronic pyelonephritis of several girls with otherwise normal urinary tracts has focused our attention on the importance of controlling bacterial infections of the kidney. At the same time, difficulties encountered in the management of kidney infections were emphasized by the stormy courses of six girls with frequently recurring pyelonephritis. Although these children had anatomically normal urinary tracts and each infection could be eradicated by appropriate therapy utilizing information gained from urine cultures and sensitivity studies, reinfection invariably followed. Prior to referral, several therapeutic approaches had been tried without achieving permanent remission, including one month of hospitalization for treatment with massive doses of antibiotic combinations. Finally, a program was instituted for the purpose of preventing recurrences.1 After the infections had been cleared by appropriate antibacterial drugs, the patient was started on methenamine mandelate and, when indicated, a urinary acidifying agent. This paper describes this prophylactic program and its
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