On Stephen Phinney and an RQ of 0.62

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Now, oxidising long chain saturated fat gives you an RQ of 0.69. Lower than this needs a supplementary process of some sort. In the last post I had Table II from Stephen Phinney's 1980 paper. There are RQs below 0.69 all over the place and even the mean RQ of the 6 week fasting exercise test was 0.66,  with some individuals down at 0.62.

So how can we manipulate RQ values?

This is a graph taken from that nice paper on ketogenic diets for rats. The black line is the RQ of the chow fed rats. They are on 17% or so calories from fat, 64% of calories from starch and the rest is protein. Grey zones are night, white zones are daytime. Ratties are nocturnal, they eat their high carbohydrate diet at night. While they are eating they run their metabolism on glucose. This should give an RQ of 1.0 but we can see the RQ is greater than 1.0 during the times at which the rats are feeding:

We've seen this before during an OGTT in massively weight reduced people. Show them some glucose and they will immediately convert it to lipid and store it. After a mere 75g of glucose during an OGTT, these post obese ladies will develop an RQ over 1.0, see the top dashed line:

This is de novo lipogenesis, either routine in the rats on a 64% carbohydrate or pathological in the post obese ladies. Glucose arrives as an oxygen rich molecule. During the reorganisation to a very oxygen poor molecule oxygen is provided without it needing to be taken up through the lungs. Smaller oxygen flux per unit CO2 produced gives an RQ greater than 1.0.

So it's pretty easy to get a RQ above 1.0. How easy is it to get an RQ below 0.69?

As we all know, acetoacetate is unstable, spontaneously decarboxylating to acetone and CO2. On its own this isn't fast enough to be useful so we have acetoacetate decarboxylase to speed the process up. You find it in the liver and in the brain, mostly. The sorts of places where glucose might be useful.

Apart from being exhaled, what is the fate of acetone in the body? I can't imagine that we are deliberately forming the stuff enzymatically just to breathe it out... Well, here's a pathway I cribbed earlier, can't remember from which paper but one on basic acetone metabolism:

Soooo theoretically ketone bodies, via acetone and oxaloacetate , are glucose precursors. If you radio label acetone with (14)C, where does it end up?

"Radioactivity from (14)C acetone was not detected in plasma free fatty acids, acetoacetate, beta-hydroxybutyrate, or other anionic compounds, but was present in plasma glucose, lipids, and proteins".

Ketones to glucose. How much?

“On the basis of our specific activity data, we have calculated that 4-11% of plasma glucose production could theoretically be derived from acetone”.

The 11% was calculated for 21 day starved humans.

The most logical explanation for an RQ of 0.62 is that the person is performing a significant conversion of fat to glucose. This is completely plausible via acetoacetate, acetone and oxaloacetate. The exact steps are unimportant. What matters is that there will be an increased consumption of oxygen per unit CO2 produced. The RQ is just a ratio so increasing oxygen use will make it drop, possibly below that 0.69 of saturated fat oxidation.

Summary: We already know that total O2 consumption must and did drop on fat adaptation. We know from simple arithmetic that CO2 production drops even more that O2 usage when fat (vs glucose) is oxidised, to give us that normal RQ of 0.69.

If there is a further usage of O2 in the process of converting ketones derived from fat in to glucose, this would explain an RQ of 0.62.

Despite this "waste" of oxygen you still use less O2 per ATP from fat oxidation, even if doing some gluconeogenesis. We know this from the absolute VO2 measurements combined with the RQ values in Phinney's Table II and my back of envelope calculations.

I sit in awe of fat oxidation. We carry fat as long term energy storage for use in times of need. Under those conditions of privation this long term energy store allows very efficient ATP generation per unit oxygen, at the same time as reducing CO2 production, at the same time as generating a significant amount of glucose. Fatty acids and beta oxidation, with ketones thrown in, are just awesome.

I'm also hugely impressed by how far ahead of its time Stephen Phinney's paper was and how well it still stacks up against modern papers.

Peter

Endurance and oxygen flux during fatty acid oxidation

I was browsing Pubmed looking for other things when I came across this ancient publication by Stephen Phinney, from back in 1980. Those were the days. That was the Summer I contracted mycoplasmal arthritis as a final year vet student seeing practice in North Devon. Lots of NSAIDs then six weeks on tetracyclines once the serology came back positive. Yum. Anyway:

Capacity for Moderate Exercise in Obese Subjects after Adaptation to a Hypocaloric, Ketogenic Diet

Free full text, nice read. Replicable, if you have the equipment.

This is table II. For the first endurance test the treadmill was set at 3 mph with a personalised incline. The same test was then repeated after one and six weeks of a protein supplemented fast. For the six week treadmill run the settings were identical to the first sessions but the subjects wore backpacks containing weights equivalent to the bodyweight that they had lost during the 6 weeks of their fast. There was no exercise training in the interim.

The column to look at is the one in the middle labelled VO2 and expressed as ml/min. On a mixed diet the treadmill settings required a flux of oxygen of 1875ml/min to maintain this significant effort. After 6 weeks on a protein supplemented fast, wearing weights in a backpack to offset the bodyweight loss, with the same treadmill settings, subjects required 1497 ml/min of oxygen. That's a big drop.

Some of this is from the magic of ketones, BHB increasing the free energy of hydrolysis of ATP. A large chunk of this component should have been in place by the one week treadmill test as everyone was well in to ketosis at this time. There was some drop in VO2 needed by this time point, but nowhere like the drop by six weeks.

We can tell by the RQ values that at baseline (mean RQ 0.76) the subjects were oxidising a mixture of fat and carbohydrate (and some protein I guess). At one week there was more fat oxidation occurring (RQ 0.72). By six weeks we have the rather strange RQ of 0.66, lower than is produced even by the oxidation of pure saturated fat.

Oxidation of a fully saturated fat produces an RQ of 0.69. Hmmmmm...

I worked out that oxidising fat, on a simple mathematical basis, should give you a 5% improvement in ATP production per unit oxygen consumed compared with glucose. With Veech's increased Gibbs free energy of ATP hydrolysis while oxidising ketones we gain a combined 5% improvement in ATP supply alongside a decreased total ATP requirement. These are components of the decreased VO2 which didn't make it in to the discussion as they were either unknown (free energy of ATP hydrolysis) or hadn't been worked out (ATP per O2). Not a bad gain for six weeks of fasting. I can see, very clearly, that anyone with a chronic lung disease would be crazy to eat any way other than for a metabolism based on lipid oxidation with ketosis. It just makes sense.

I'll leave it now for this post and put up some ideas about the subnormal RQ figure as the next post.

Peter

Dave Asprey and Dr Veech

Dave Asprey has a very interesting extended discussion with Dr Veech on his Bulletproof website, mostly about ketones but also about the history of biochemistry and a number of other subjects. Dr Veech is very pro ketones while being surprisingly anti high fat diets, an interesting combination and clearly far from my own perspective. Much of the interview is simply fascinating in its own right but I'd just like to talk about the aspects with which I disagree. As one must.

Exclusive: Interview with Ketone Expert Dr. Richard Veech

The section of interest is around one hour five minutes in to the discussion. For those who would like some flavour of Dr Veech's uncomfortable stances on a fat based diet, cardiologists and cholesterol you can grind your teeth through the ten minutes leading up to that point. It's pretty obvious that Dr Veech has a lot of reading to do on cholesterol and cardiovascular health, should he so wish.

I had to correct a few relatively minor typos, nothing to do with what was said, just how it got written down, so I've put up the original text followed by my edited version. I've listened to this section three times now and I think my modified transcript is correct:



Original:

Dr. Veech: However, when you’re burning fat, you’re going through beta-oxidation. One reducing equivalent goes to NAD and one goes to pflavo protein. You’ve already lost 1/3 of your ATP in that step. Go back to your lennature Lehninger, beta-oxidation, you do one NADH, you do one NADH, you do one pflavo protein. You do that to keep from blowing the mitochondria up.

Tidied up:

Dr. Veech: However, when you’re burning fat, you’re going through beta-oxidation. One reducing equivalent goes to NAD and one goes to flavo protein. You’ve already lost 1/3 of your ATP in that step. Go back to your Lehninger, beta-oxidation, you do one NADH, you do one flavo protein. You do that to keep from blowing the mitochondria up.




I sold my copy of the biblical Lehninger at the end of my second year at vet school when we finished biochemistry as a subject in its own right. I have to say I really enjoyed the book. I also midnight question-spotted the urea cycle for my biochemistry written exam and it came up. Yeaha!

Sooooooo. Yes, beta oxidation (of saturated fats) does indeed yield one FADH2 which is embedded in a flavoprotein as well as generating an NADH. Electron transporting flavoprotein (ETF), which docks with ETF dehydrogenase, reduces the CoQ couple and omits pumping the four protons which could have been pumped had a second NADH been generated instead. My back-of-an-envelope calculations suggest an NADH could have been generated instead of the FADH2. This does indeed waste a four protons worth of the ATP which might have been generated.

And yes, the FADH2 is generated to stop damage to our mitochondria. So fats are bad?



With some slight discomfort I have to re-cite (yet again) the Protons thread. The whole point of generating inputs which reduce the CoQ couple is to drive reverse electron flow through complex I. Low levels to trigger insulin signalling, high levels to resist insulin signalling. H2O2 is the second messenger.

The waste of proton pumping by supplying FADH2 at the CoQ couple is offset by it being used to regulate the system. This applies to mitochondrial glycerol-3-phosphate dehydrogenase, driven by glycolysis, or ETFdh driven by beta oxidation of saturated fats. Both initially assist insulin signalling and, as substrate throughput increases, they facilitate resistance to the signal from insulin to accept more calories.

Another sooooooo. FADH2 is, I agree, wasteful but more importantly it also drives reverse electron flow through complex I. And undoubtedly you can have too much of a good thing. Waaaay back in the early Protons posts I spent a lot of time looking at FADH2:NADH ratios and decided that somewhere around palmitic or stearic acids there was a maximum healthy FADH2:NADH ratio, somewhere around 0.48. At that time Dr Speijer was kind enough to supply his paper looking at the development of the peroxisome in LECA, the Last Eukaryote Common Ancestor (not to be confused with LUCA, of the hydrothermal vents).

One major function of peroxisomes is to deal with very long chain fatty acids using a beta oxidation version which does not generate FADH2. Problem solved, no need to blow a gasket in our mitochondria. In fact the peroxisomes shorten VLC fatty acids to octanoate, much beloved of Dave Asprey and Dr Veech.

Minor aside: How might you explode your mitochondria if Dr Veech's concerns were correct re FFAs and his idea about FADH2 being used to reduce the supply of pumped protons from complex I? There is a technique to completely flood your mitochondria with NADH. It's called beta-hydroxybutyrate, particularly if supplied in large amounts from ketone esters. This enters the mitochondria using the monocarboxylate transporters, generates NADH as it converts to acetoacetate then goes on to generate three more NADHs from each of the two acetyl-CoA entering the TCA. These NADHs are potentially capable of pumping membrane popping numbers of protons through complex I, without concerning Dr Veech. And without actually doing any damage, that I can tell. End aside.

Next minor aside: Free fatty acids are not going to explode your mitochondria: Under ketogenic high fat eating there is a combination of elevated free fatty acids (happy happy), low insulin (so no loss of FFAs through insulin induced triglyceride formation), plenty of ATP in the mitochondria to allow uncoupling proteins to function (UCPs must have generous ATP on their mitochondrial matrix end to allow them to function) and those ad lib FFAs are necessary and available to actually carry the protons through the uncoupling proteins. FFAs are essential for uncoupling, no FFAs = no uncoupling, pax mtG3Pdh. FFAs = uncoupling = no exploding mitochondria. End second aside.

I'm left here with my view of a healthy metabolism as one based around beta oxidation of saturated fatty acids. Ketones as they happen, no stress. Nothing originating from the discussion has budged my entrenched position in the least, interesting though it has been to listen to.

Peter

When is a ketogenic diet not a ketogenic diet?

I've listened to a few youtube/podcasts videos recently. Seyfried, Veech and D'Agostino appear to besettled in to what looks like a "Ketone Ester" corner. There are many, many things which make me splutter a little in some of the things they say but I think it's hard to decry ketone bodies too badly. They clearly do things. It's quite possible that the main effect of flooding the TCA with acetyl-CoA is the inhibition of glycolysis. If it does nothing else, that seems worth doing.

The flip side is Ron Rosedale's view, shared by a few others. Towards the end of his presentation, largely about mTOR, he takes a position on ketones. I don't think he has anything against them per se, but what he really wants is a metabolic state based around beta oxidation of fatty acids. Just enough protein, minimal carbs, oxidise fats. If that throws off some ketones, so be it.

I clearly recall Wooo posting some time ago about the ketogenic diet for epilepsy and pointing out that what matters is compliance with the diet, not levels of a given ketone in blood or urine. If fatty acid oxidation dominates I'd be willing to bet the glial cells generate a ton of ketones which enter neurons without a dipstick in sight.

I think I might be in that camp. So I'm a little uncomfortable with medium chain triglycerides, octanoate alone, ketone salts and ketone esters. They clearly have benefits but they are not a route I would take currently.

I like ketones as a surrogate for fatty acid oxidation. Or should I say that I like beta oxidation, and ketones are a reasonable surrogate.

For the beta oxidation "camp" we have this paper:

Induction of ketosis in rats fed low-carbohydrate, high-fat diets depends on the relative abundance of dietary fat and protein.

It's a no nonsense sort of a paper. For the LC rats the diet was beef dripping. With added casein at 5.5%, 11.8% or 19.1% of calories and a few vitamins and minerals. The only carbs came from the vitamin/mineral mix. Anyone could get any rodent diet manufacturer to formulate it:

Rule one of a scientific paper: The methods must supply enough information to replicate the study.

Replication = validation. Without it your paper is worthless.

So rats on a LC diet are only in ketosis when over 90% of calories are supplied as beef dripping:

A BHB blood level of 28mg/dl is reasonable ketosis. That would be around 3.0mmol/l in new money. That's for the rats on 5.5% casein in their beef fat. All else was ns compared to the chow fed rats. There we have it. Decent ketosis in a rat is reliably achievable by feeding beef tallow. No MCTs, no ketone esters, no octanoate. Anyone could do this with their rodents. One niggle:

For the CICOtards: The LC animals were pair fed to the calories eaten by the chow fed rats, a feature of the paper I dislike a little. It's equal calories all round but the low carb rats probably ate less than their appetite would have dictated. This might have accentuated ketosis (but we'll never know...) and ad lib feeding might have blunted ketosis.

The weight gains themselves are fun:

Obviously the red squares are the ketogenic animals. There is a table of blood insulin, glucose, FGF-21 and FFAs but, well, we all know what the number have to be so there's not much need to go in to it in any detail. KDs work.

So if you want to know what a ketogenic diet does in a given medical condition, this is the one diet you have to use on a rat model. Maybe use butter instead of beef dripping (I'd prefer this for myself) but it looks like a gold standard to me... Ketones as a spin off of a whole body FFA based metabolism. The metabolic state is what interests me. Replicate at will.


Let's compare this with a ketone ester camp paper. This is the diet:

"...mice received KD-USF, a custom ketogenic diet designed by the authors and produced by Harlan Laboratories, fed ad libitum".

What's it made of? Dunno. Here are the macros:

What sort of fat? Dunno. Just fat. Maybe: Crisco? Fish oil? Canola? Coconut? Mmmm, butter? Replication anyone? If you can't replicate the study how can you tell whether the results were made up or real? Personally, I think the results are absolutely true. I have no doubt. Why? Because this is the level of ketones generated by the diet, where it says KD:

The ketogenic diet generated, estimated by zooming in on the above chart, something around 0.1-0.2mmol/l of BHB. No one would make up a figure that low, these are honest results. With added ketone esters we get up to almost exactly the level of ketones found in Bielohuby's truly ketogenic tallow fed rats, without the crippling expense of the ketone esters.

People shouldn't get me wrong. I have nothing against trying to use a ketogenic diet for management of cancer. I even think using ketone esters might be reasonable for folks who can't cope with cream, butter, eggs and a wide variety of meats and non starch veggies.

What I would prefer is for a diet described as ketogenic to actually generate some ketones. You cannot describe a diet generating 0.1mmol/l of BHB as ketogenic! Especially when close on 3.0mmol/l is easily achieved on a beef tallow based true ketogenic diet. It's not exactly surprising that adding 3.0mmol/l BHB derived from ketone esters should out performed a non-ketogenic "ketogenic" diet for cancer management! And the non ketogenic high fat diet did help a little, presumably by eliminating starch triggered insulin signalling...

I was a bit shocked that non of this was discussed in the discussion. Being driven by a love of ketone esters is no excuse for sloppy science. When you are on the winning side there is no need for this.

Peter

Would Franklin have taken a statin if they had been available in the 1800s?

The Franklin expedition of 129 men perished in the Arctic, primarily of scurvy and related illnesses, in its entirety. I've been reading Stefansson's retrospective account and speculation written in the 1950s. This paragraph stuck out as a nice warning, still applicable to the lipid hypothesis today, which we see crumbling before out eyes. How many have died prematurely of medical dogma?

"We concede that Franklin had the excuses which many others have used; that medical men told him nothing about meat being preventive of scurvy, and that they told him instead about lime juice and lemon juice as preventatives and curatives. But we return to our point that a man who makes exploration a profession, as Franklin had done, has no business to sacrifice his men to the dogma of current therapeutics when he can divide the entire literature of his own craft into two chains of events; the expeditions which had a good deal of fresh food [meat] and little or no scurvy; and those which had little or no fresh food and much scurvy".

Fat vs sugar. PUFA vs saturates. Statin vs giving the finger to Keys for the buried Minnesota data.

Peter

Glucosamine and aged mice

It would appear that glucosamine, the arthritis neutraceutical, promotes longevity.

D-Glucosamine supplementation extends life span of nematodes and of ageing mice

This paper is a mix of C. elegans and mouse work. Most of the findings are reasonably transferable but the core finding, that a metabolite of glucosamine inhibits glycolysis, was actually demonstrated in the worm section. Extended longevity was found both in C. elegans and ready-aged mice.

Glycolysis inhibition and life extension in C. elegans:

And for longevity in already aged mice:

It's worth noting that glucosamine usage in humans is observationally associated with extended lifespan.

To me the core finding is that the inhibition of glycolysis extends life. I like that. In the mice it was also found to cause insulin resistance, mild at the dose rate used in the paper. However I would expect resisting insulin to be associated with extending lifespan. I'm quite keen on extending my own healthspan. The longer this goes on, the happier I'll be. My plan is resist insulin.

Now. Is there any other simple way of inhibiting glycolysis which might rival extended glucosamine administration?

If we go to Veech's isolated, perfused, working rat hearts (again) we can see some interesting things:

I suppose the first thing we have to say is that the heart works quite well on the utterly non physiological substrate of isolated glucose.

Next is that insulin decreases glycolysis compared to this baseline. The increased glucose ingress to the cell is diverted to glycogen synthesis while insulin simultaneously increases the efficiency of the electron transport chain. So net flux through glycolysis drops from 5.6 to 3.8 micromol/min/ml. Ketones (a physiological mix of AcAc and BHB) drop glycolysis from the baseline 5.6  down to 1.7micromoles/min/ml, which I find quite impressive.

As always, Veech maintains a phobia about fat so we have no data about what the addition of fatty acids might do to glycolysis. Enough said that ketones suppress it quite well. Veech has a detailed discussion of the various redox couples which control this process. To a simpleton like myself we can just view the situation as observing when ketones flood the TCA with acetyl-CoA there is nowhere for glycolysis products to go to, so it stops. The effect of redox couples on the energy yield of ATP hydrolysis is a separate consideration.


Let's summarise: Ketone bodies markedly inhibit glycolysis. Note to push-biking self: They also promote glycogen formation.


Glucosamine drops glycolysis in C. elegans by 43% and gives a measurable longevity gain. Ketones in an isolated perfused working rat heart drop glycolysis by 66% and appear, in many papers, to have some favourable effects on many of the problems which come to us with age. I find that interesting.

Peter

Alcohol and fructose are the same to your liver

From Liz Miller via Facebook.

Alcoholic Liver Disease: Update on the Role of Dietary Fat

"The protective effects of dietary saturated fat (SF) and deleterious effects of dietary unsaturated fat (USF) on alcohol-induced liver pathology are well recognized and documented in experimental animal models of ALD. Moreover, it has been demonstrated in an epidemiological study of alcoholic cirrhosis that dietary intake of SF was associated with a lower mortality rates, whereas dietary intake of USF was associated with a higher mortality".

My italics.

Brief note to Saturophiles and Cholesterophiles:

I think we can say that we have won.

Peter

Dairy and diabetes

Mozaffarian has published this nice observational correlation between three markers of dairy consumption and subsequent onset of diabetes over the following 15 or so years.

Circulating Biomarkers of Dairy Fat and Risk of Incident Diabetes Mellitus Among US Men and Women in Two Large Prospective Cohorts

Any of the three markers is associated with a roughly halving of the incidence of diabetes. I sort of like this, living largely on bulk calories from dairy myself. I also like the size of the effect. If you consider how many people are in the process of developing diabetes in the USA alone, halving that incidence might make a significant contribution to reducing suffering.

If you eat a gram of C15:0 fat, what else comes bundled along with it? Well, in Swedish milk fat there is about a gram of C15:0 in 100g of total fat. That total fat is made up of 70% saturated fats, mostly palmitic acid, a decent dollop of stearic and myristic acids plus odds and sods of shorter chain saturates. These fats might be what reduces the diabetes risk. What makes me think that, other than my biases?

If you feed a normal Bl/6 mouse 40% of its calories as stearic acid, what happens to its blood glucose level? Taken from fig 2.3.1 part B. After 10 weeks the blood glucose of a normal mouse will be significantly lower than if it had been fed standard CIAB or 40% of an oleic acid/PUFA mix. That will be the red arrow:

The blood glucose lowering effect even occurs in db/db mice, a routine model used to vaguely represent T2 diabetes, green arrow. That's all fine and gives some sort of suggestion that it might be the saturated nature of dairy which is protective against diabetes. But why should this occur?

I'm drawn back to the perfused isolated pancreas and the insulin response to physiological levels of glucose in the presence of various fatty acids. This is the image I'm thinking of, from The Insulinotropic Potency of Fatty Acids Is Influenced Profoundly by Their Chain Length and Degree of Saturation:

Notice the marked but transient spike in insulin when glucose is raised from 3.0mmol/l to 12.5mmol/l, most obvious in the black squares representing stearic acid as the background FFA (palmitate is the black triangles). After the spike, which I think represents the first phase insulin response, there is a steady climb in insulin, equivalent to the second phase of insulin secretion. Obviously this is needed because it's an isolate pancreas prep, glucose is fixed at 12.5mmol/l in the perfusate. If the first phase insulin response does its job in real life the systemic circulation (and pancreas) would never see 12.5mmol/l of glucose. The surge of insulin would hit the liver and interact with the insulin receptor. Two things follow on from this. Most insulin would be metabolised following interaction with its receptor, so insulin would never flood the systemic circulation. Second effect is that insulin/insulin receptor activation would shut down hepatic glucose output while the Glut2 transporters continue to pretty well clear the portal vein of glucose.

So a first phase insulin response is designed to protect the systemic circulation from both hyperinsulinaemia and hyperglycaemia. That is its job.

If you want to obliterate the first phase insulin response what you need to do is to reduce reverse electron flow through complex I. Just take a peek at the open circles (oleic acid) and, even better, the closed circles (linoleic acid). For a healthy pancreas, from a healthy rat, you can eliminate the first phase insulin response to hyperglycamia just by choosing your background FFA.

Summary:

Arterycloggingsaturatedfat first phase insulin response 16ng/fraction

vs

Hearthealthypolyunsaturatedfat first phase insulin response 2ng/fraction.

Want to be fat? Use your second phase insulin response in the systemic circulation to pack fat and glucose in to adipocytes. But you must avoid the first phase response because this will keep glucose in the liver as harmless glycogen.

Try using sunflower oil or corn oil as advised by the Food Standards Agency. When you get so fat that your adipocytes start to spew unregulated FFAs, you can be a diabetic (congratulations!). You owe it all to your cardiologist. Or the FSA. Remember who to contact when you get your first diabetic amputation.

Peter

Oh, picked up this quote about dairy and heart disease from Tom Naughton's blog:

"Rather than suggesting that the saturated fats in dairy products are harmless, Aslibekyan and co-author Ana Baylin, an adjunct assistant professor of community health at Brown, hypothesize that other nutrients in dairy products are protective against heart disease, for all but perhaps the highest dairy consumption quintile in their study. The potentially beneficial nutrients include calcium, vitamin D, potassium, magnesium and conjugated linoleic acid (CLA)".

The authors are completely wrong in the interpretation of their own data. On every front. Saturated fats are NOT harmless. Shout it from the roof tops. THEY ARE PROTECTIVE.

"calcium, vitamin D, potassium, magnesium and conjugated linoleic acid (CLA)"?

Bollocks. Ask the mice on stearic acid.

Stearate, butter and leptin receptors: Speculation!

I suppose the first thing I have to say is that the Tatter Paper of the last blog entry is not science in the form that any scientist might recognise. Control your variables is rule one... I only posted on it because, by a complete and apparently unplanned accident, the deep fried chips were of similar macros to the boiled mashed potatoes (BMP) but differed in fat type, in a manner very exciting from a Protons point of view. Just to emphasise: This was a chance gem in a pile of wallanga*. Picking the gem out of the wallanga can get your fingers dirty, but it's worth it. Chance is occasionally very useful.

I also happened to notice that, again by chance, the BMP macros were going to pan out somewhere near 40% fat, using butter as the bulk calorie source. This too is quite exciting.

Here are the BMP's macros, just roughed out to round numbers:

The post before the Tatter Paper post was merely pointing out that control C57Bl/6 mice do NOT become obese on their high (40% of calories, chance, neat huh?) fat diet in Ms Reeves' PhD. Their weight might be a little heavier on an olive oil based diet with generous PUFA and a little lighter on a stearic acid based diet with minimal PUFA, but nothing dramatic and absolutely no obesity in sight.

From the 40% fat fed mice in the stearic acid PhD:

Not a perfect match but reasonable. Why is this interesting?

My (repetitive) idea is that a certain amount of input to the ETC via ETFdh drives reverse electron flow through complex I to limit adipocyte distention. Stearic acid plus chow starch does this early during a meal. Butter plus potato starch does this early during a meal. Canola oil plus potato starch doesn't.

We have no idea whether the butter with potato starch would carry on, long term, to a slim phenotype in people. The long term effect of stearate is downwards and of oleate/linoleate is upwards on bodyweight in mice, but the effect is small. How come?

High level signalling.

Anyone who has read Hyperlipid over the last few years will be well aware that I hate high level signalling. It usually takes a basic process, like the control of insulin signalling by the ratio of inputs at complex I, ETFdh and mtG3Pdh, and sticks a nice, glossy, superficial and somewhat opaque surface veneer over it. Then researchers can go off to find 25 or more genes which have some level of influence at some level of "higher-ness" of signalling above the core process. We then end up with a morass of over information with no one linking it all to the core process.

Such a high level signalling molecule is leptin. I have had relatively little interest in leptin over the years so may well be missing large chunks of information which are common knowledge to others on tinternet. The basic process seems to be that fat cells make leptin and the hypothalamus uses the information embedded in blood leptin levels to make a ton of decisions about energy homeostasis and energy use. Including appetite. Leptin secretion is related to adipocyte size but deeply under pinning adipocyte size is the ETC's control of insulin signalling, which sets cellular fat content. Leptin appears to provide some long term modulation of a series of repeated short term post prandial insulin events.

We can strip off the surface veneer of longer term leptin signalling from the core mitochondrial process by using db/db mice. The db/db mouse has non-functional leptin receptors. This means that the acute effect of mitochondrial signalling within adipocytes is not smoothed over or averaged out by the brain using leptin. The core process in fat cells takes over and can be seen via body weight and fat mass.

At peak energy flux stearic acid generates the maximum resistance to insulin's distending effect on adipocytes. Oleic/linoleic is far less able to generate insulin resistance to limit calorie ingress to each adipocyte.

The mice which are db/db homozygous become obese on chow (17% fat largely PUFA). On the 40% oleic/linoleic acid diet they become even more obese because they have plenty of dietary fat to store and a minimal ability to resist insulin's storage signalling. Stearic acid fed db/db mice also have a ton of fat available for storage. They don't lose any weight but they don't gain any weight either and they still up pretty damned close to normal mice fed normal chow. A little heavier, but not much:

The point of this post is emphasise that saturated fat, which I consider to drive physiological adipocyte insulin resistance, limits weight gain in leptin receptor KO mice. The fact it also cures their diabetes at the same time is another story.

Summary so far:

I consider that leptin smooths out the differences in fat storage produced by superoxide signalling originating from the ETC. I hate this, being a great fan of superoxide signalling. Stearate generated superoxide can largely offset the obesogenic effect of being a db/db mouse produced by the attendant lack of leptin signalling. It works with stearate at 40% of the diet, but not at 17% (elsewhere in the PhD).

The rest of this post is wild speculation originating from combining the Stearate PhD and the Tatter Paper:

Could this combo of spuds (or any other starch), butter (38% of calories, quite similar to 40% of calories from stearate) and meatballs work for people who are db/db in the same way as the stearate diet works in db/db mice?

Even more wild speculation, because the db/db genotype is fairly uncommon in humans:

Could the butter component of the "tatters" side-step failed leptin signalling (as in obesity) or relative/absolute hypoleptinaemia (as in the post-obese) in the same way that a ketogenic diet side steps the need for insulin signalling?

Further wild speculation:

Could putting a human on to a high saturated fat ketogenic diet sidestep most of the obesity problems currently prevalent in the world? By giving actual weight loss...

Make up your own mind...

Peter

A final comment on leptin. I'm unfamiliar with the massive complexity of leptin signalling. It seems to go on and on for ever. But just occasionally you come across little snippets of interest which suggest things about the function of leptin. There is a group who have developed an adipocyte specific leptin receptor knockout model. The only cells to lose their leptin receptors are the adipocytes. They still make leptin, they still release leptin, the liver still sees leptin, the hypothalamus still sees leptin. What happens?

"Despite a normal level of leptin receptors in the hypothalamus and normal food intake, mutant mice developed increased adiposity, decreased body temperature, hyperinsulinemia, hypertriglyceridemia, impaired glucose tolerance and insulin sensitivity, as well as elevated hepatic and skeletal muscle triglyceride levels".

The mice become obese and diabetic (on chow of course). Just by their adipocytes failing to perceive plasma leptin levels. And folks think the brain controls obesity! And of course you should be able to fix these adipocytes by supplying stearic acid as 40% of the diet.

Aside: The brain is clearly important in controlling all sorts of physiology. No one would deny this. Much as the computer of a modern car closely controls engine performance (my sister used to drive a Mitsubishi Lancer Evolution which turned out to develop 270bhp on a rolling road. She'd paid for the 315bhp version. Mitsubishi took out the computer, sent it to Japan for upgrade, refitted it and, hey presto, 315bhp. Never touched the engine), so too does the brain fine tune metabolism. But if metabolism is broken peripherally there's not much point looking in the brain. Trying to upgrade the performance of a Morris Minor by reprogramming its computer would be technically slightly difficult. When I used to tune Morris Minor engines computers still ran on punched cards. But the core process in the engines of a Lancer and a Moggy are the same. End aside.

So, did leptin arise to allow fat cells to monitor the fullness of other fat cells so as to maintain a reasonable level of fat stores? Then the brain started listening in? I don't know, but I find the idea interesting. And of course, the basic control of fat storage at that stage would then have been ETC derived superoxide. A little gets insulin signalling going. A lots shuts insulin signalling down. Insulin signalling, of course, is core. Even today.

Final final comment. This post makes me sound like Ray Peat. Something I find very embarrassing, to say the least.


About the asterisk:

*Wallang! Wallanga: You goin' in dat cave man? It's dark in there. We keep cows in there. We keep sheep in there. We keep pigs in there. Take care you don't step in no wallanga.

It needs a guitar, a folk club and a very long shaggy dog ballad to get you to this punch line.

Boiled mashed potatoes for miracle satiety?

The effects of potatoes and other carbohydrate side dishes consumed with meat on food intake, glycemia and satiety response in children.

With thanks to Mike Eades for the full text.

This is an interesting study. Given a meal of meatballs plus a choice of five different carbohydrate sources, a group of children ate a great deal less (in calories) of boiled mashed potatoes than of pasta, rice or either of two types of chips.

"The five treatment sessions consisted of ad libitum servings of (i) rice, (ii) pasta, (iii) boiled and mashed potato (BMP), (iv) baked French fries (BFF) and (v) fried French fries (FFF) with a fixed amount (100 g) of meatballs".

What did they find?

"... children consumed 30–40% less calories at meals with BMP (p less than 0.0001) compared with all other treatments, which were similar".

That's a LOT less calories! Potatoes seem to have some sort of magical satiety property. If you believe in magic. Table 1 gives an inkling of the problems with the study:

As you read through the cooking description you realise (red box) that the carbohydrates had very different amounts of added fat per unit carbohydrate and that some had butter (+/- added milk) while others had canola oil in varying doses. So when we look at Table 3 we have to realise that "CHO amount (g)" means an assorted mix of various fats and carbs:

We have to work back using Table 1 to find out what amounts of carbohydrate and fat were actually eaten and read the cooking details to find out what the fats were in each dish. Some arithmetic gives us this for what was actually eaten:

To my mind the trial here splits in to two. We have BMP, boiled mashed potatoes with 3g of carbohydrate per gram of butter, which is fairly well matched with FFF, chips deep fried in canola oil, with 2g of carbohydrate per gram of canola oil. Both are potatoes. Both provide a roughly similar ratio of calories/grams from glucose and fat. Both are relatively low carbohydrate per unit fat (compared to the other three meals, ie just in this study).

From the Protons point of view the relatively low carb BMP and FFF are supplying glucose from potatoes to drive complex I. However butter also supplies FADH2 at ETFdh, so generates a resistance within adipocytes (and elsewhere) to an excessive insulin facilitated calorie ingress during the period of maximal blood nutrient levels. When calories stop falling in to adipocytes, satiety kicks in. Using FADH2 this happens after eating 508 kcal. With FFF based on canola oil, ie potatoes steeped in 18 carbon omega 3 and 6 PUFA, the beta oxidation generates a much lower input at ETFdh (one less FADH2 per double bond) and so insulin sensitivity at peak nutrient uptake is maintained for longer, fat pours in to adipocytes for longer and almost twice as many calories are consumed (912 kcal) before satiety kicks in. I expect satiety to rise as blood nutrients rise. Not sequestering them in to adipocytes seems the best way to do this. More physiological insulin resistance. I'm guessing the brain does the actual sensing of both glucose and FFAs.

I like that. You can say what you like about the hypothalamus. I prefer to think about the adipocytes and their mitochondria as determining what gets done with food and hunger. There is some input from leptin of course, but that's another post.


The other three carbohydrate dishes are essentially lowish fat foods with between 7g and 10g of carbohydrate per gram of butter or canola oil.

In these lower fat preparations it takes three or four teaspoons of butter to generate satiety vs just under 6 teaspoons of canola oil, roughly twice as much fat is needed when carried with a similar amount of starch. A reasonable fit with a Protons point of view, though not as pleasing as the BMP vs FFF comparison.

How the study was developed is fascinating to think about.

What decisions were made at the planning stage? Obviously, someone had worked out, well before any grant application was submitted, that higher saturated fat with lower carb meals are by far the most satiating. Or maybe they are dumb and they were just lucky to get a result? Personally, I can't see how you engineer a study like this unless you are pretty clever and well informed, not at the mitochondrial level of course, but certainly at the butter level. Mashed potatoes, which already have something of a reputation as a miracle weight loss food, getting a helping hand... From a dollop of butter. It makes sense.

BTW this is Canada. I can't see how such a study would ever have gotten past any ethics review committee in the US of A. Imagine trying to feed BUTTER to American children. Immoral. Plus they might not eat up their carbs!

Peter

High fat fed mice on stearic acid

The concept of finding anything positive about palmitic acid is still tantamount to research suicide. However, stearic acid is a rather different matter. It's lipid "neutral" for those poor folks who still bow their heads and kneel before the altar of the lipid hypothesis. So you can publish good stuff about stearic acid with relative impunity.

Raymond sent me the PhD thesis of Valerie Reeves, Kentucky University.

Reeves, Valerie Lynn, "A DIET ENRICHED IN STEARIC ACID PROTECTS AGAINST THE PROGRESSION OF TYPE 2 DIABETES IN LEPTIN RECEPTOR DEFICIENT MICE (DB/DB)" (2012).Theses and Dissertations--Physiology. Paper 3.

Before we think about leptin receptor defective mice (another day), we can ask questions about the control groups. Such as:

What happens if you feed a fairly typical C57Bl/6 mouse 40% of its calories from fat, based on fully saturated stearic acid?

They stay significantly slimmer than they do on CIAB (chow) and probably slimmer than when fed on 40% oleic acid (olive oil w/o the PUFA).

(EDIT As Tucker pointed out in comments: You might be able to explain the relative weight gains in terms of omega 6 PUFA. Chow was about 13% of energy as PUFA, stearic acid diet about 5% PUFA and the oleic acid diet about 14% PUFA. The correlation of PUFA with fat gain isn’t perfect but it’s quite close… END EDIT)

Now this is clearly impossible, as anyone who has read anything about Bl/6 mice and fat will be very aware. So the poor girl did it again:

This time we have p values sprouting all over the graph like mould in a Winter bathroom. For mice, chow makes you fat. Olive oil makes you fat. Stearic acid doesn't. Impossible I know, but that's twice it has happened. For fat mass the p values never make pay dirt but the writing is on the wall for oleic acid and fat gain too:

The wild type control mice were so nice in this PhD thesis that I thought I'd just put up these few figures before we consider what might happen if (gasp) you put an obese, diabetic db-/- mouse on a highly saturated stearic acid based diet.

I think palmitic acid would do exactly the same as stearic acid did for these mice. But who would risk their career with a finding like that? The corollary is that when you see a C57Bl/6 mouse get fat on a high fat diet, you know there are lots of double bonds in that fat........

Peter

Not really much about swimming underwater (2)

Just a one liner after all the discussions about breath holding on a fat based diet:

Effects of Twenty Days of the Ketogenic Diet on Metabolic and Respiratory Parameters in Healthy Subjects.

The first person I came across doing this practically was a LC blogger back in my early days (probably 2002-ish) and I didn't realise why she was LC eating to manage her chronic lung disease, from the metabolic perspective. She was very focused on saturated fat, obviously (with hind sight!). I've not been through the above link's full text but you all know the depths of stupidity of most saturophobes. If this was corn oil and MCT based... Perhaps we could get a significant O2 consumption drop given some butter, dunno. Nice to see some medics taking this seriously!

Peter

With apologies to whoever put this up on Facebook, one of those saved links but no idea who it came from.

Life (11) Ferredoxin

Anyone who has read through the Life series will know that I have a great deal of time for reduced FeS moieties as the core energy source used during the transition between pre-biotic chemistry and something resembling life.

Bottom line: An electron from a reduced FeS moiety is able to reduce dissolved CO2 to a Ni bound CO- group using molecular hydrogen. If you supply a couple of geochemical CH3-SH molecules you can then generate acetyl-SH, precursor to acetyl-CoA, from this Ni-CO. After that it's down hill all the way to metabolism. This is where the watchmaker comes from who is going to make the watch which you might find in the jungle.

One of the earliest biological problems was to detach this reduced FeS from the inorganic cell wall and make it mobile. The solution is ferredoxin.

People have looked at the ferredoxin used by those bacteria which have done well for several billion years by developing the form of metabolism most closely allied to this very basic pre biotic chemistry. One of these ferredoxins was sequenced very early in the 1960's, while I was just a kid playing in the streets of Nottingham and Mike Russell (thanks Jack) had just gained his geology BSc from  London University.

Working with the ferredoxin sequence from Clostridium pasteurianum Eck and Dayhoff noticed some interesting things. There looks to have been a very early gene duplication, this allows the two sections of the protein to be compared to each other and this facilitated all sorts of speculation about its possible origin. A sort of molecular Rosetta Stone. Here is the sequence they started from in the nicely descriptive three letter code (it becomes more legible if you click on it):

To make comparisons easier to fit on a given line they then changed the three letter notation to the less descriptive single letter notation for amino acids during the rest of the discussion, which goes like this:

The legend to Fig 1 is quite self explanatory but, if anyone wants the full text to work though it line by line in more detail, I have the pdf. The end conclusion is that primordial ferredoxin could be derived from a simple repeating pattern of just 4 amino acids. These four:

From these four amino acids they suggest you can reverse engineer the process giving this as the process of generating ferredoxin:

Apart from a nice discussion about why a very early protein, given billions of years to evolve, remains so remarkably similar to its primordial sequence, they also have a think about what the ADSG polymer might have been doing before it was co-opted to pick up an FeS cluster. Possibly some sort of simple structural polymer. They also throw in the concept that the FeS might initially have been only chelated to cysteine, I would suggest as a solubilising agent. Again, cysteine is one of the most primordial of amino acids:

I found the paper and its proposals fascinating. They are talking about concepts which fit extremely well in to Mike Russell's ideas about hydrothermal vents at a time before there was any evidence that the vents existed. As far as I can tell it has no bearing on anything we might do today but I still like it. It says a great deal about where we might have come from.

Peter

Insulin glucagon and protein

Again from dissertante's query: How can chicken be found to raise blood glucose, acutely?

Many years ago, as a beginner at treating diabetic animals, I tried to balance insulin dose rate/timing against carbohydrate intake. Owners always asked if there was anything they could feed as treats etc. I used to suggest meat and fat as they shouldn't need insulin for processing.

This was a mistake. Dogs are, by the time we diagnose them, functionally type 1 diabetics. While fat is perfectly OK, protein certainly isn't.

Eating protein, for a type 1 diabetic, produces an immediate rise in blood glucose. This is nothing to do with gluconeogenic amino acids, the effect of which would expect to be delayed for several hours, if it occurs at all. While protein for an normal human being/animal is neutral on systemic blood glucose it never the less produces an immediate spike (by around 60 minutes) in blood insulin.

Dandona measured insulin and glucose, although not glucagon, after casein ingestion as we saw in the last post:

Eating 75g of casein protein more or less triples your blood insulin level but doesn't budge blood glucose down any more than cream does, which leaves insulin pretty well alone. Under normal conditions the casein induced spike in insulin is counterbalanced by a rise in glucagon. If the insulin rise does not occur (through beta cell failure) the glucagon will still rise and is unopposed, so hyperglycaemia is the net result, coming from a rise in hepatic glucose output.

This took me years to realise. Slow, I know but ah well.... It's now common knowledge and Dr Unger's glucagonocentric view of diabetic hyperglycaemia makes a great deal of sense.

So protein will provoke hyperglycaemia in the absence of an insulin response, via glucagon, in a type 1 diabetic. I would guess that the same would apply to an advanced type 2. It very recently occurred to me that an elevated blood glucose after protein intake might be a useful supplementary test for certain oddities in OGTTs.

I had an email a few weeks ago about OGTT results in long term, non diabetic low carb eaters. I don't know the exact details of duration of LC eating or the period of carb loading before the OGTT, but the end result after glucose ingestion was a sustained hyperglycaemia with profoundly depressed C-peptide levels.

The worry here is that long term LC might have led to endocrine pancreatic insufficiency. My initial thought was to wonder what the response to exogenous insulin might be, but this was probably the wrong line of thought.

What would be far more interesting would be to run an oral protein response test, looking at blood glucose, insulin, glucagon and C-peptide. Although, at a pinch, all you need is the blood glucose result. If a person has developed a significant loss of beta cells then the unopposed alpha cell glucagon response to this protein would produce hyperglycaemia. A normal insulin reaction in response to protein would produce normoglycaemia after said protein load.

We all know that after a month or two of LC eating that three days at 150g/d of carbs will restore a normal response to glucose. But the question is what time scale of carb loading is needed after several years of LC eating. The regulation of insulin secretion in response to glucose requires active glycolysis, regulated by glucokinase in the pancreas. Glucokinase gene expression is controlled by dietary glucose supply. If long term glucokinase down regulation takes longer than a few days of carbohydrate loading to reverse, this would produce intolerance to glucose but would have no effect on insulin secretion when driven by amino acids. It would be quite simple to differentiate between down regulation of the pancreatic glucose sensor from newly acquired type 1 diabetes during LC eating.

Summary: Elevated blood glucose after an oral protein load suggests genuine diabetes. Poor responsiveness to glucose after sustained LC eating simply reflects a mothballed glucose sensor, provided response to protein is normal.

Peter

Personalised nutrition: Eat fat

Personalized Nutrition by Prediction of Glycemic Responses

In the comments after the last post, dissertante asked about the above study. It's been around for a while and many folks have talked about it, Bill Lagakos being one of the more articulate. The study is enormous. The paper is quite long and, for various reasons, not exactly gripping reading for myself. So I may well have missed certain facts which are not immediately obvious. This is the summary of the study from the abstract:

My initial thought was to ask how the insulin response varied between people with a normoglycaemic response to junk food vs hyperglycaemic response. Typical junk foods considered in the study are the bananas vs the cookies in section G of Figure 2:

If normoglycaemia is bought at the cost of hyperinsulinaemia, it's not particularly attractive, to me anyway. Banana, cookie, who cares? The only way I can see that either of these is acceptable as food is if they are taken by the gut bacteria, converted to short chain fatty acids and so bypass the whole insulin/glucose signalling system. Many people seem to be happy to trust their health and glycaemic control to their gut bacteria. It takes all sorts I guess.

So, the implication is that we can use this massive level of investigation to make choices between carbs which spike glucose and carbs which don't. For us, on an individual basis, tailored nutrition. Without any idea of what these given sources of carbohydrate do to an individual's insulin levels. But, to be quite honest, it's junk vs junk anyway.

There is a snippet which shows a glimmer of interest in the use of fat to blunt the glycaemic response to carbohydrate by the group. This is what they say:

"The PDP [partial dependence plots, part of their model] of fat exhibits a beneficial effect for fat since our algorithm predicts, on average, lower PPGR [post prandial glucose response] as the meal’s ratio of fat to carbohydrates (Figure 4C) or total fat content (Figure S5A) increases, consistent with studies showing that adding fat to meals may reduce the PPGR (Cunningham and Read, 1989). However, here too, we found that the effect of fat varies across people".

Fat cannot reliably save us from carbohydrate induced hyperglycaemia. We still need personalised nutrition, even if we eat fat.

But what if we eat only fat? What would be the glycaemic response to 100ml of double cream, drunk on its own, for breakfast?

Dandona, on his way to drawing incorrect conclusions, gives us the glucose and insulin data for 100ml of double cream:

Drinking cream alone mildly reduces  insulin after a transient rise and point blank drops glucose throughout the study period. There may be minor individual variations in response but these are all contained within standard deviations which narrow with time after exposure... There is little scope for a pathological rise in glucose or insulin within those SDs.

So how much do we have to go begging, cap-in-hand, to our gut microbiota for a nice glucose AND insulin response to 100ml of cream? Not a lot. Ditto butter, lard, beef dripping...

The simple approach to personalised nutrition is to eat fat, cut out the middle man of our microbiota, limit glucose and reduce signalling through the insulin pathway while eating just enough protein to meet our needs. Anything else is going to need an awful lot of laboratory investigations to even get half the information we need to keep our blood glucose levels remotely normal while still using unknown amounts of insulin.

Personalised nutrition: Eat fat.

Peter

Oh, dissertante also mention that, for some people, chicken came through as a "bad" food in terms of post prandial glycaemia. That's another post I guess.

On drinking varnish

Dietary linoleic acid elevates the endocannabinoids 2-AG and anandamide and promotes weight gain in mice fed a low fat diet.

Raphi sent me this link early in the New Year. It’s nice. It demonstrates, at some level of complexity, that omega 6 PUFA at 8% of calories are obesogenic in mice, even if they are fed otherwise fat free CIAB. It’s all about endocannabinoid ligands and receptor activation. Potentially useful when folks get round to starting class actions against the cardiological community and any other health advisors warning against saturated fat. If you limit fat to 30% of calories and saturated fat to 10% you still have 20% PUFA/MUFA in your diet. That’s easily obesogenic. Your cardiologist made you fat. Sue now.

But all of this endocannabinoid stuff is what I call high level signalling. At the core mitochondrial level we know that omega 6 PUFA fail to limit insulin activity under situations where a saturated fat would shut down insulin mediated calorie ingress. In an adipocyte this means that, during oxidation of omega 6 PUFA, insulin continues to signal and fatty acids (and glucose) fall in to the adipocytes, stay there, and you get really hungry. Modified chemicals derived from this system of omega six fatty acids are overlaid on top of the core mitochondrial signalling. A modified derivative of arachidonic acid becomes an endocannabinoid ligand and makes you hungry and fat. The system takes something basic and develops an overlay of enormous complexity, this is what I call higher level signalling.

I hate higher level signalling. Give me the core process anyday.

On this front people may realise I have issues with omega 3 PUFA fats. From the ETC perspective they are worse than omega 6 PUFA and should be more obesogenic. But, in general they’re not. In fact there is a massive industry showing us how good they are for us. But there are suggestions that the core process which makes omega 6 PUFA obesogenic really do apply to the omega 3s. Bear in mind that we are only talking about linoleic and alpha linolenic acids here. Longer fatty acids go to peroxisomes for oxidation and have little influence on core mitochondrial processes, though they do perform a great deal of high level signalling. Here we go:

Sucrose counteracts the anti-inflammatory effect of fish oil in adipose tissue and increases obesity development in mice.

Notice the obesogenic effect of fish oil only shows when sucrose is present in the diet. Replacing sucrose with protein eliminates the effect. Fructose is an unstoppable source of cellular energy intake which needs insulin resistance to limit insulin signalling facilitated ingress of glucose. As insulin continues to act, fat cells sequester calories. Fish oil combined with sucrose is the worst, corn oil is intermediate and, without sucrose, none of the fats are obesogenic.

This makes me happy. I can see the core process at work, never mind what EPA and DHA say to g-protein coupled receptors.

There is another paper which shows a similar effect and I like it rather a lot because the cognitive dissonance, which shines through every word of the text, is rather entertaining. How can you get a life-sustaining source of funding if your data show that omega 3 PUFA are grossly obesogenic? They improve insulin signalling exactly as the ETC effects would predict. The cost of improved insulin responsiveness in adipocytes is obesity. Here we go again:

Adipose tissue inflammation induced by high-fat diet in obese diabetic mice is prevented by n-3 polyunsaturated fatty acids.

The values to look at begin with the weight gain. All we have to do is to subtract weight at the start of the study period from weight at the end (perhaps the authors don't do arithmetic?). Low fat group gained a gram, added saturated fat group gained 0.6 g, added omega 6 group lost* 2.4g and omega 3 group gained 10.4g.

Ten point four grams.

These are db/db mice which lack a functional leptin receptor. They are diabetic and I feel their chronic hyperglycaemia represents a similar drive to obesity as the fructose loading in the last study, ie an unregulated source of calories which drop in to adipocytes and which require insulin resistance to shut down whatever further caloric ingress it can practically do. Free fatty acids, a reasonable surrogate for the action of unmeasured insulin, are low so this suggests adipocyte sensitivity to insulin is high, hence the weight gain.

Weight gain in the alpha linolenic acid group was over 17 times that of the saturated fat group and 10 times that of the low fat group. Notice saturated fat protected (admittedly ns) against the weight gain seen on the low fat diet. The logic is obvious. What do the authors say? Well, I can find no mention in the discussion of this massive weight gain in the omega 3 group. Zilch. This is the quote from the only mention it gets, in the results section:

"Body weight at the end of the study was somewhat higher in db/db mice fed HF/3 compared with HF/S (Table 1)".

My emphasis.

There is no other mention of the hard fact that omega 3 fats are obesogenic. Also note that in relatively normal, non hyperglycaemic db/+ mice, the omega 3s are not obesogenic. Much the same as for non-fructose fed mice in the previous study.

Now look at the * I put in above. The omega 6 diabetic group LOST 2.4g. Ouch, at the core mitochondrial function level! How can this be? This needs no mention at all in the paper because p is greater than 0.05 (in the twisted stats used by the authors). But brownie points if you have noted the oddity about this particular group of mice.

Well done! Yes, in a group of 5 animals the standard deviation at the end of omega 6 feeding is 8.6. No other group had a standard deviation greater than 3 at any time. How do you get a standard deviation of 8.6? These are diabetic mice. Four gained weight, one became ill and this one lost a lot of weight. That's my guess, just trying to reverse engineer information out of the data supplied by a group of dissonant thinkers...

So, I went to an on-line standard deviation calculator and fed in various options where 4 mice gained some weight and one mouse lost a tonne of weight. Using a 2g gain for 4 possibly healthy mice and a 20g loss for the fifth poorly mouse we get four mice at 44g and one at 22g. This gives a mean weight at the end of the study of 39.5g to with an SD of just over 9. I think something like this is what happened. Would this group notice one skinny mouse in with four fat ones? Hahahahaha!

Summary: When PUFA are being oxidised in the mitochondria of adipocytes, those adipocytes are unable to resist the signal from insulin to distend with fat. The more double bonds in the PUFA has, the greater the effect. Linseed oil should be used for making varnish.

Peter

Paignton Zoo

So funny that both articles come from Paignton Zoo in Devon. Has anyone contacted the victims of Lynne Garton's Going Ape "Evo Diet"? To tell them to knock off the fruit and live on raw kale leaves? Good enough for monkeys....Luckily Garton's stupidity seems to have done no permanent damage to it's victims, beyond 12 days of flatulence in the "study"!

Going ape.

Monkeys banned from eating bananas at Devon zoo.

Thanks to Amber O'Hearn via Faceache for the second link.

Peter

Not really much about swimming underwater

*****MAJOR ERROR********

Down in the comments section Mateusz has very kindly found the error in my arithmetic for me. It makes the whole of this post completely incorrect and requires a great deal of working through posts based on this conclusion to correct my mistake and the implications this has for blood supply and oxygen consumption.

Fats require around about 5% more O2 per ATP cf glucose.

With apologies to everyone.

I know I said (first paragraph) that I would take this post down in embarrassment if I'd made an arithmetical error but, on balance, I think it should stay as a warning, to me as much as anyone else.

So I’m going to leave this post up unchanged, with this edit, as a warming to the immense power of confirmation bias. There’s a lot to do.

*****MAJOR ERROR********







Just before I hit post: I think the arithmetic and the logic here are sound on a ball-park basis but if anyone can point out any major flaws I stand to be corrected and will take the post down in embarrassment. But this is so simple in concept that I don't see why it's not standard fare... Here we go.

In the comments after a previous post it became pretty obvious that several LC eating folks noted a significant improvement in their ability to breath-hold while running their metabolism on fat rather than on glucose. Although this is rather counter intuitive based on the RQ (more oxygen is required per unit CO2 generated when you oxidise fat compared to glucose) what matters is the generation of ATP per unit oxygen or ATP per unit CO2 produced. I started with oxygen. Arithmetic goes like this:

Glucose oxidation is simple. Six carbons give 2ATP from glycolysis and a mix of NADH and FADH2 from the TCA:

6(CH2O) + 6O2 = 6CO2 + 6H2O      
RQ: CO2/O2 = 6/6 = 1.0
2 ATP + 10NADH + 2FADH2


A theoretical six carbon section of a chain of a fully saturated fatty acid gives this:

6(CH2) + 9O2 = 6CO2 + 3H2O        
RQ: CO2/O2 = 6/9 = 0.67
15NADH + 6FADH2


Three of the FADH2s are from acetyl CoA turning the TCA, the other three are from beta oxidation. For PUFA a theoretical alternating sequence of single and double bonds yields this:

6(CH1.5) + 8.25 O2 = 6CO2 + 4.5 H2O
RQ: CO2/O2 = 6/8.25 = 0.73
15NADH + 3FADH2


The first step of beta oxidation for PUFA yields no FADH2, so we just have the three from the TCA. Assuming the ETC works efficiently we pump these protons from our hydrogen supply:

NADH = 12H+
FADH2 = 8H+

And, very crudely, let’s assume at complex V, ATP synthase, we have 4H+ = 1 ATP (not true IRL!)

So we can calculate protons pumped, what this is worth in ATP and combine this with the O2 needed (from the chemical equations above) giving:


Glucose protons
10NADH = 120    2FADH2 = 16, total = 136 H+
ATP 34 + 2 = 36

ATP-gluc/O2 = 6.00

Saturated fat protons
15NADH = 180     6FADH2 = 48, total = 228 H+
ATP = 57

ATP-sat/O2 = 6.33

PUFA protons
15NADH = 180 3FADH2 = 24, total = 204 H+
ATP = 51

ATP-pufa/O2 = 6.12


Clearly fatty acids are better at generating ATP per unit O2 consumed. If a 70kg person, at rest, is consuming 200ml of oxygen per minute to produce a given amount of ATP while burning glucose they should be able to maintain that same amount of ATP on less oxygen.


But the difference seems pretty small. How small?

Through sins of education I tend to think of O2 consumption for an anaesthetised, mechanically ventilated patient. That person needs about 200ml/min of oxygen.

200ml O2 gives 6.00 x10^bw ATP if running on glucose (where 10^bw is a crude scalar to whole body ATP needs). On saturated fat:

200ml O2 gives 6.33 x 10^bw ATP

Or, more realistically:

190ml of O2 gives 6.00 x 10^bw ATP on fat, equivalent to 200ml O2 used on glucose. An oxygen sparing effect of 10ml/min is underwhelming on first consideration. It’s a 5% improvement. But this should be maintained at VO2 max. When oxygen delivery is the limiting factor in performance, running on fat gives you a 5% advantage.

This is simple arithmetic applied to the most basic of biochemistry processes.

Is butter a performance enhancing drug?

Yes, provided it displaces carbohydrate.

Should folks with ischaemic problems eat butter?

Yes, provided it displaces carbohydrate.

Does it taste good?

Yes, unqualified.

Of course, once you add in ketones, magic starts to happen to the energy yield of ATP hydrolysis. Ketones are not as arithmetically simple as fatty acids but we all know, from Veech and D'Agostino's work, that magical indeed they are.

Peter

Oh, I calculated CO2 per unit ATP produced too. On carbs ATP/CO2 = 6.00 as you would expect but on saturated fat the amount ATP produced per unit CO2 evolved is 9.5. CO2 build up makes you breathe, you make less per minute on fats. Breath holding is, arithmetically thinking, expected to be easier running on saturated fat. This is what we find.

*****EDIT*****

Hans pointed out in comments that the TCA provides a molecule of GTP which can convert to ATP from each acetyl-CoA. This gives two extra ATP's per glucose and three more ATP's per six carbons from saturated fat. I can't be *rsed to re do the math, but you get the picture.

*****END EDIT*****