September 17, 2014, Bethesda, Maryland
The National Heart, Lung, and Blood Institute convened this workshop to discuss the results of recent research on the effects of inorganic nitrate and nitrite on the cardiovascular system, possible long term effects of these compounds in the diet and drinking water, and future research needs including population-wide effects examined through epidemiological studies.
Research in the past decade has demonstrated that the body can use exogenous nitrite and nitrate to produce the important messenger molecule, nitric oxide. In small feeding trials, ingested nitrate is absorbed with great efficiency, after which some of it is reduced to nitrite by enterosalivary cycling. Nitrite is then available as a circulating storage pool for nitric oxide bioactivity. Subsequent reduction of nitrite to nitric oxide takes place in many tissues, particularly under conditions of hypoxia, resulting in vasodilation and platelet inhibition, and the enhancement of oxidative metabolic efficiency, among other functions.
It is possible that the cardiovascular protective effect of a diet rich in fruits and vegetables is in part due to its high nitrate content, contributed largely by leafy green vegetables and root vegetables. In an attempt to exploit the cardiovascular benefits of nitrate for the benefit of athletic performance, there has been a recent rapid proliferation of unregulated performance-enhancing supplements containing high amounts of nitrate. However, in contrast to a likely salutary role played by dietary nitrate and nitrite in cardiovascular health, there is a long history of research on a possible deleterious role of nitrate and nitrite in food and of nitrate in drinking water with the endogenous production of carcinogenic N-nitroso compounds.
To address the current state of our knowledge of the exposure and effects of dietary nitrate, we convened this working group of investigators in nitrate physiology; nitrate distribution in groundwater, soils, plants, and dietary components; cancer epidemiology; and cardiovascular disease epidemiology.
A review of recent discoveries surrounding the nitrate-nitrite-nitric oxide cycling in the body was presented, including the function of the enterosalivary cycle and the importance of nitrite reservoirs such as hemoglobin and myoglobin. Presenters also discussed the results of recent experiments on the effects of exogenous nitrate and nitrite on platelet function and coagulation, inflammation, cardiac function, blood pressure, and in the brain. The effect of exogenous nitrate and nitrite on nitric oxide availability to the mitochondria was shown to result in increased oxidative efficiency and result in heightened exercise performance without concomitant increase in oxygen uptake.
The availability and distribution of exogenous nitrate and nitrite in drinking water and diet was then addressed. Models of nitrate in U.S. groundwater were shown, and data were shared on the geographical distribution of excessive groundwater nitrate. Nitrate and nitrite levels in a typical U.S. diet were compared with those found in the traditional diets of other countries. Data were presented on the levels of nitrate and nitrite in human breast milk and infant formulas. It was noted that nitrate tolerance levels in drinking water have been set to limit the amount available to young infants in order to prevent acute methemoglobinemia, and participants engaged in considerable discussion regarding the degree to which much of the susceptibility of infants might be due to concurrent gastrointestinal infection.
The working group then considered ongoing large observational cohorts that may be useful in studying the long term effects of dietary nitrate exposures. Current ongoing U.S. population-based studies that capture detailed dietary information from large numbers of participants that represent the entire country are the National Health and Nutrition Examination Survey, sponsored by the Centers for Disease Control and Prevention, and the Reasons for Geographic and Racial Differences in Stroke cohort, sponsored by the National Institutes of Health. Representatives of both of these studies presented the current available data and plans for future data capture. However, limited data on drinking water are available from these studies, and the quantification of dietary nitrate ingestion is hampered by the often substantial variability in nitrate content within a single species of vegetable, and the lack of standardized dietary databases that list the nitrate content of dietary components.
Data were presented from several epidemiological studies of cancer and nitrate or nitrite exposure. Current studies have generally found no effect, or even some cancer protection, from nitrate in the vegetable content of the diet, perhaps due to the protection from N-nitrosation afforded by simultaneous ingestion of antioxidant components of vegetables. Drinking water nitrate is considered by cancer epidemiologists to be of more concern due to the lack of concurrent antioxidant consumption. Of new concern is the widespread and unregulated use of athletic supplements that may contain high levels of nitrate or nitrite without protective antioxidants.
-- Improve the standard dietary databases used in epidemiological research to include estimates of the nitrate content of commonly eaten vegetables, such that when dietary patterns are evaluated for cardiovascular benefit, the nitrate content is taken into account. Large cohort studies should incorporate measures of nitrate in urine and plasma whenever possible.
-- Add measures of drinking water nitrate content to observational cohort data collection, particularly those that include persons in rural areas drinking from private wells. Although these persons are a minority of the U.S. public, it is important to accurately ascertain whether they are at increased risk of cancer resulting from chronic high water nitrate ingestion without simultaneous antioxidant ingestion, and whether they experience any long term cardiovascular benefit from their high level of nitrate ingestion.
--Although current research in nitrate carcinogenicity does not support a role for nitrate consumed as vegetables in the diet, there is an urgent need to examine whether some of the popular high-nitrate athletic performance supplements may be increasing endogenous production of carcinogenic N-nitroso compounds, especially those supplements that do not incorporate antioxidants. Small feeding trials could be designed that include urine testing for N-nitroso compounds.
Amrita Ahluwalia, Ph.D
Barts and the London School of Medicine and Dentistry
Mark Gladwin, M.D.
University of Pittsburgh School of Medicine
Gary D. Coleman, Ph.D.
University of Maryland
Norman Hord, Ph.D., M.P.H., R.D.
Oregon State University
George Howard, Dr.P.H.
University of Alabama at Birmingham
Daniel Kim-Shapiro, Ph.D.
Wake Forest University
Martin Lajous, M.D., Ph.D.
Nacional de Salud Pública de Mexico
Filip Larsen, Ph.D.
David J. Lefer, Ph.D.
Louisiana State University Health Sciences Center
Leslie A. McClure, Ph.D.
University of Alabama at Birmingham
Bernard T. Nolan, Ph.D.
U.S. Geological Survey
Ryszard Pluta, M.D., Ph.D.
National Institute of Neurological Disorders and Stroke
Alan Schechter, M.D.
National Institute for Diabetes and Digestive and Kidney Diseases
Chia-Yih Wang, Ph.D.
National Center for Health Statistics
Mary H. Ward, Ph.D.
National Cancer Institute
Jane Harman, D.V.M., Ph.D.
Division of Cardiovascular Sciences