Why dietary nitrate is hard to beet – Andrew Webb

Why dietary nitrate is hard to beet - Andrew Webb



much in fact Society for inviting us to speak yeah it is a double act so attempt a few minutes each I think so it's in now is perhaps quite obvious that nitrates does things to the vascular chair but that's only become obvious in the last five to ten years it really wasn't obvious but it's quite difficult to see that so I'm going to try and take us back and show you why it is but it's obviously this on the title slide this is what obviously rocket which is high nitrate these or high nitrate containing foods spinach Popeye here's possibly right this is a fennel in the shape of a heart it's obviously good for your heart and the beat though this is beating in a beetroot and lots of vegetables there so this is that this is nitrites no2 – and this is nitrates only three – nitrite is used to cure meats and preserve them and nitrates is very high in green leafy leafy vegetables and beetroot so back in 1981 green hell you'll guess what the next names gonna live in a minute shade that actually we excrete a lot of nitrates in the year in but it's about four times more than the nitrates that you ingest so they had no idea where that was coming from at the time so after green leaf came along from the same group of Tannenbaum at all and he showed that it came from l-arginine so and it was around that time of course he'll arginine and Oh path was discovered spinach here which is also very high in dietary nitrates also contains a lot of elaj and it's put in the highest l-arginine containing vegetables soy protein also contains a lot for example so this was the l-arginine na na sono pathway or a large needs to substrate for much cocoa synthase and then this pathway therefore became established in the mid to late 80s and salvador McCadden I mean he's in there ninja medicine review in 1993 said that nitrites and nitrates are oxidized metabolites of nitric oxides essentially they were considered to be inert very useful for measuring nitric oxide activity but we thought to have no active to themselves obviously nitric oxides a highly active free radical so it's not moving on there was a concern however that nitrate nachos Amiens have been shown in the 50s 60s to be carcinogenic in rat models in rats because nitrites could form nitrous a means has a lot of concern about nitrite and also now vegetables of nitrate as this pathway was was starting to be demonstrated and so they were thought to be undesired residues in the food chain however the Mediterranean diet is high in fruit and vegetables particularly sort of salad vegetables and is associated with a lower incidence of cardiovascular disease and there's also a particular association with allowance of Commerce of disease and green leafy vegetables for example at Josh Shapiro shows and also the – diet is high infusion to vegetables has been estimated have a large content of their dietary nitrates and that may partly count for reducing blood pressure so just so you're clear nitrites and nitrates are inorganic as opposed to organic compounds and that have a hydrocarbon structure so here's a nitroglycerin or glycerin nitrates which is an organic nitrates now so it's different it's a volatile and explosive compounds if you drop it by accident it explodes quickly Alfred Nobel of course worked out how to stabilize it with silicone silicone paste to make dynamite he ended up getting angina later in life and treated himself with nitroglycerin which works partly by releasing much Cox idream established nitric oxide so the question was could inorganic nitrites also be a source of nitric oxides and it was known under very acidic conditions that you can acidify nitrites were no but was this relevant in physiology so in 1995 as Varsho din the language off isolated rat heart preparation and that if you perfuse it with nitrites under normal conditions then using EPR spectroscopy there's no generation of nitric oxide however if you then use 30 minutes of ischemia the rat hearts you get a very strong signal shown here which is a due to nitric oxide production and under these conditions to get a reducing condition with reduced oxygen and protons or hydrogen ions and so the pathway starts to come back this way as an alternative source of nitric oxide so the next question is is nitrite beneficial and in this paper 95 and looking at the recovery of the heart function this was the Rach pressure product and following in a scheme of reprovision injury it appeared that there nitrites was Dilla Terius it reduced the recovery of the heart function and interest in nitrites sort of waned off around that time I thought it's a cancer risk and it might be source of nitric oxide nitrous oxides is a pollutant but it does bad things in scheme reperfusion and yeah the field sort of went almost dead for a while I presented society of work I did with Ben Benjamin amrita al auliyah and 2002 showing controversially that nitrites in a similar scheme diffusion injury model appear to be very protective and here was the reduction from say line control versus nitrites at two concentrations so here 10 100 micro molar that was a lonely time at the time as a nature paper this is a society presentation 2002 but subsequent to this there have been no further studies showing a deleterious effect but there have been other studies by other groups grew from mark glad we in the town a year later in 2005 in the heart and the liver and other organs and a year-on-year they've been further studies and now this has gone into clinical trials and several trials in in Britain of nitrites and acute in acute myocardial infarction so the next question was doesn't like try it dilates blood vessels so we share another acidic or ischemic conditions the nitrite can get reduced back to air no but what under more under physiological conditions and so this paper came out from 2001 from the group of multi Karamazov foolish and it actually says in the title plasmon liked right so this one rather than nitrate reflects regional endothelial NOS synthase activity so this is the acute effect the increase enter activity you see an acute rising light right first but lacks intrinsic phase of duration activity so now it's only under ischemic perhaps hypoxic conditions in the same year on the same time this group from Yunnan Lumbergh and early mites burg in the Karolinska showed that under acidic conditions in the rat a litter so this is pH 6.6 of protons again you could detect more profound vasodilation of a alter with lower concentrations of nitrites so perhaps out situation however in 2003 the group of mark globulin showed published in Nature Medicine showing that so even with fairly large concentrations nitrite they're able to demonstrate an increase in foreign blood flow surveys dilation effects and they also showed about a seven millimeter reduction in the mean arterial pressure associated with it with a higher dose than the one shown there and so the next question is how is nitrite reduced to nitric oxide is it just acidic conditions or what else could do it and so the goofin mark gladwyn mainly pioneered this showing that deoxyhemoglobin so hemoglobin under the oxygenation conditions can reduce nitrates when their so this is going through the vasculature from oxygenating conditions in the larger arteries to the resistance vessels in skeletal muscle where nightwear oxygen gets removed by metabolism and you get reduction nitrite severe no and the balance point for maximum conversion nitrites when a is when hemoglobin is about 50 percent saturated with oxygen said the oxy hemoglobin is one way to convert nitrites or no and that that's the equation and then we showed that xanthine oxidase and heart tissue and this is rat heart tissue but also human heart tissue if you give allopurinol for example it's inhibited xanthine oxidase will reduce the amount of Mo produced from nitrites with later showed that xanthine oxidase also present on the red cells and so this is another site where night track can be reduced to I know and also surprisingly endothelial nitric oxide synthase itself could also act as an nitrite reductase to generate mo from nitrites and subsequently several other nitro trajectories have been demonstrated so that's a thinking about nitrites which is sort of where the field was focused or pulp field was fixed on at the time what what about nitrates could that generate might transfer an oak and going back to 1994 both been Benjamin and you and Lumbergh independently showed that darting nitrates saved whoops or giving letters for example would increase gastric NO NO levels and this Ben Benjamin showed that this had anti effective properties for example but it was subsequent paper by June nights with Ben Benjamin showed that whilst you get a big rise in gastric nitrates as you'd expect and plasma nitrates the gastric nitrite levels are very low so it was thought that you all you generate NO and not much gastric nitrite and so how does nitrates inhibits platelets all the question is does nitrates in favorite platelets and this was shown in 1999 we've quite profound inhibition of platelet function from nitrate so how does it do it so the next question and is it via snows s nitroso files and this is the chemistry if you're interested in the chemistry you can generate s no trace of vials from nitrates and nitrites and indeed they showed snow production in this in the stomach but couldn't demonstrate the presence of nitrogen files in the plasma in the circulation so it was slightly mysterious how nitrates I was having an aunty plates effect because in a publication just after that about canal hotel and they fed people lettuce for a few days over a reasonable sample size with 27 subjects but couldn't demonstrate any significant increase in plasma nitrite so if you took a lettuce or nitrates doctrine nitrate source it appeared that you were increasing of gastric NO not increasing gastric nitrite or plasma nitrites generating nitroso files in the stomach and they were possibly getting into circulation but that was couldn't be demonstrated and so this is the sort of situation you'd swallowed action nitrate this would be absorbed into the circulation and gets concentrated again in the saliva glands violence entry slime circulation and it would be reduced to nitrites by battery on the back of the tongue and then on to nitric oxide and other agents here and possibly some of this nitrous of files that reduce might get absorbed but didn't appear that nitrite was getting absorbed so we looked to see whether nitrate could increase plasma nitrites and this is how the study was portrayed in the Daily Mail but the subjects were fully clothed so it's not quite accurate and as as expected we got a very large rise in plasma nitrate and we also demonstrated a fairly acute rise in plasma nitrites although it took it wasn't as acute as the rise in nitrate it peaked at around two and a half to three hours and this is because of time to accumulate the night traits in the air provide the entry cycle circulation in the saliva glands and then secreted and then had to be converted to not try to move it to be absorbed so that's the explanation for the delay so here we were getting direct absorption of nitrites from the gut into the circulation so the next questions does nitrate therefore lower blood pressure but because it's generating nitrites and previously been shown that nitrites could lower the mean arterial blood pressure and yes indeed it did we had over a 10 millimeter reduction compared to control in the randomized crossover study in healthy volunteers and you can see here the peak drop in blood pressure compared to baseline was it around two and a half or so hours and that corresponded to the peak rise in plasma nitrite I'm saying you may have you seen this cartoon before but this you've swallowed the beetroot shown in purple here containing doctrine nitrate about two-thirds of nitrate is excreted in urine if fact Iria are here in the urinary tract to UTI that contain nitrate reductase enzymes they reduce the nitrates and nitrites that's a principle of the urine dipstick on the walls so the about 20 to 28 percent of nitrate load is concentrated in saliva glands and bacteria here which also have nitro productors enzymes has similar to a UTI reduce the nitrate to nitrite that's swallowed and then now we know that light right is absorbed and then in the circulation particularly deoxyhemoglobin for example the nitrite is reduced further to nitric oxide and we wanted to interrogate this a bit more and so this is just the first bit so we've got subjects to swallow nitrates of beetroot juice and then for the next three hours to spit out all the saliva and this was to stop them then absorbing nitrites and then having subsequent effects to show that it was nitrite that was having these effects on blood pressure for example so here we go splat up the saliva of three hours and there was no effect on the plasma nitrate concentration but it stopped completely the rise in with spitting all the saliva of three hours that rise we otherwise gotten crossover study of nitrites and also a similar thing with the blood pressure when they spat us all the saliva there's no drop in blood pressure but we could repeat drop in blood pressure with normal soiling of the saliva three hours saved just quickly because we'd shown previously that nitrites was protecting a skier effusion injury could nitrate as it's now a source of nitrite also be protective and indeed and a flow mediated dilation I our study of 20 minutes ischemia of 22 diffusion we show protection here with the purple bars this is beet treat to pre and post and this is just control water so all this the last question could nitrites inhibit platelets we've been shown in 2005 by nathan brian metal inmate recanvassing biology and if you gave the IP nitrite to rodents if anything there was an increase in place aggregation so well those know that enno could be an antiplatelet agent nitrite didn't seem to show that and everyone believed that at the time apart from us I thought surely at night tracks can reduce turnover and it'll be an to play lives so we looked at that in the spitting and swallowing study for example this is normal swallowing of saliva and we've got a significant inhibition post beat reduces his pre beet juice versus post we produce with normal stalling survival when the subject spat out of the saliva of three hours and then we tested plate aggregation again this effect was completely lost – this is collagen but we've got the same results – ADP and other agonists so the well established now na nitrite nitrate pathway I'm in 2008 the reverse of that the nitrate nitrite le path I was born so it's just surround the other way but that's only was that first is mentioned really in 2008 so only five years ago and this is some data we presented a couple of years ago to the society showing following nitrate ingestion of bigger eyes and nitrates and then the rising might try it and with Excel Dino we also get quite a large increase in exhaled in it as well so again nitrate nitrite any pathway and of course all this sort of stuff was node a long time go by the Chinese this is from the Dunhuang collection collection 2000 or so scrolls and is basically a description of using saltpeter potassium nitrate for patients who sound like they've got them acute coronary syndrome probably and they say this this will surely work at the bottom and this is of course while they were trying to create explosives as well and blow themselves up and look for an elixir of life at the same time the last point is all the other effects of nitrate nitrite the only pathway there's big concern about stomach cancer this has recently been quite extensive reviewed by Nathan brine and the states and a critical review the animal toxicology mixture of nitrite so is in the absence of administration of costume McKnight was aiming precursor there's no good evidence for carcinogenesis and the recent perspective that epidemiological studies also don't really demonstrating the SH in the in rodent models nitrogen I try to actually gastro protective against NSAID induced gastrula the–for example and as an alternative medicine a lot of patients with cancer think that their beetroot extract may be protective and there some in vitro and in vivo evidence for a variety of their cancer lines there are a lot of other effects which I haven't got time to go into but exercise was a big thing in the Olympics that's the secret of Team GB as Boris Johnson said was beetroot juice but a lot of other facts have been demonstrated subsequently and I think Vick will cover and some of these I'll skip over there's a very exciting area of research at the moment all all sorts of therapeutic avenues are possible for patients suggest the – diet and this is perhaps useful we published in a review last year Satnam I lived in myself that the nitrate veg table so if they interested in having a high nitrate diet rather than just buying beetroot juice which is one alternative and they can go for these high nitrates there things say for a UK portion that's got 80 grams so these high nitrate containing things green leafy vegetables and beetroot there's about two millimoles or I tell the patients nitrate units for an 80 gram a UK portion and then here in lower nitrate containing vegetables in water there is a acceptable daily intake but really this is probably a load of rubbish it was derived from 1962 studies in rats and dogs where they gave 500 milligrams per kilogram of nitrate with no observed adverse effects and therefore divided that by a hundredfold to get the human equivalent so first into man type dose if you like and that's where five milligrams kilogram of nitrates come sodium nitrate comes from so perhaps the ADI is actually the exceptional dog intake rather than the acceptable daily intake a browser might give through this but this is just a summary table of all the changes in the understanding of paradigm shifts that have occurred really all in the last decades with nitrites and nitrates game and finally if any of you are in doubt about how to take dietary nitrates if several people have tried wearing it so recommend bats necessary it doesn't work transdermally we don't think you nor Lady Gaga with that note or a hat they're all even math so you have to swallow it like that so you didn't have to be native to do it or spinach and Popeye also showed us how to expand it and finally to acknowledge AB lot of people who were involved in studies particularly a high return earlier I think as we can talk next and Nigel Benjamin Kennedy Cruikshank who's joined us recently who with him we're doing a beach reduce short term study with other things thank you

Add a Comment

Your email address will not be published. Required fields are marked *