CAPE-rich Propolis for Autism?

CAPE (caffeic acid phenethyl ester) is a substance known to be an inhibitor of PAK1.  PAK1 has been shown at MIT to be implicated in various disorders including Fragile X and schizophrenia.  PAK1 inhibitors are also effective in research models of various cancers, including leukemia.

There are currently no approved PAK1 inhibitor drugs, although several are in development.

PAK1 is also implicated in Neurofibromatosis, and clinicians have researched various alternative PAK1 inhibiting substances.  The two most interesting ones that I have already written posts about are:-

·        Ivermectin, an old anti-parasite drug (also shown effective in leukemia)

·        BIO 30 propolis, rich in CAPE

Ivermectin is already used as an autism treatment by “alternative” doctors who think autism is caused by parasites.  We saw in a recent post that a study looking for parasites in people with autism (in the US) found none.  Ivermectin reportedly does improve autism, according to one reader of this blog and other anecdotal evidence.

I think Ivermectin is likely to be more potent than BIO30, but Ivermectin cannot be safely used continuously, without long breaks.

BIO-30 Trial

Having discussed the idea with one of the Japanese Neurofibromatosis clinicians, it seemed worthwhile to see the effect in our kind of autism.

As you may have seen in previous posts the science behind PAK1 is complex.  It has numerous, mainly bad, effects.  It is involved in dendritic spine morphology; this might be one area where ongoing “damage” is still being done.  So when asked what kind of change I expected/hoped to see, I said “cognitive improvement”.

According to recent research:-

CAPE alone has never been used clinically, due to its poor bioavailability/water-solubility; Bio 30 contains plenty of lipids which solubilize CAPE, and also includes several other anticancer ingredients that seem to act synergistically with CAPE.

Propolis is widely used as a natural remedy, but this was my first experience with it.  The first problem was how to take it; it sticks to everything.

My solution is to cut a small piece of toast and then apply 20 drops of propolis.  Since propolis has a strong flavor, I try to mask it with a layer of Nutella spread on top.

I gave this “honey medicine” at breakfast and in early afternoon.  

Trial Conclusion

There is a cognitive enhancing effect, noticeable not just to me.  The effect is visible almost straight away, but was more noticeable with a dose of 2 x 20 drops than with my original 1 x 20 drops.

At this dosage, it is not revolutionary, but it does indeed provide a real “nootropic”/cognitive enhancing effect.

Propolis for All?

At the dose I am using, I would think this “therapy” is only worthwhile in people whose autism is well-controlled already; meaning no stimming/stereotypy/OCD, allergies/GI problems all resolved, no aggression or anxiety;  these behaviours will mask any benefit.

I actually think this is the first thing I have come across that looks ideally suited for Asperger’s and other HFA.

I did look on line for people trying BIO30 for schizophrenia, all I found was someone else asking the same question:-

Apparently FRAX486 treats schizophrenia in mice due to PAK1 inhibition. Why does no one try Bio 30 Propolis for schizophrenia, as it is a PAK1 inhibitor as well?

Propolis does have numerous other ingredients, including many very interesting flavonoids.

As long as you are not one of the one percent of people with a bee allergy, propolis seems a very safe product.

If you live in Australia or New Zealand you can buy the CAPE-rich propolis locally.  As we learnt in previous posts, only two types of propolis were found to be PAK1 inhibitors, an expensive one from Brazil and the CAPE-rich BIO30 Propolis from New Zealand.

If anyone tries it, please let me know the result.  You only need one bottle and a few days to see if it has an effect.

What does Cancer Risk and Autism tell us?

Today’s post is a short one.

As you look deeper into how the body functions you come across many, only recently understood, pathways.  In reality these are still “works in progress”, but some will eventually lead to a better understanding of diseases like cancer, diabetes, Parkinson’s, Alzheimer’s and, eventually, many types of autism.

Within this blog we have seen how many common diseases share some underpinnings with autism.  As a result these diseases appear more commonly in people with autism, and so they get called comorbidities.

Some comorbidities get talked about quite a lot, things like epilepsy and MR/intellectual impairment.

For me the really interesting ones and the ones that might actual lead you to some therapeutic implication.  In this respect, allergies (food and airborne) have proved to be the most useful.

Not far behind are heart disease, diabetes and cancer.

In Paul Whiteley’s blog he recently highlighted a study showing how heart disease was increased in autism.  This has been noted before and I believe leads back to calcium channels, known to be dysfunctional in autism.  One particular channel is called Cav1.2 and it is widely expressed in the brain and the heart.  In earlier posts I have covered this channelopathy from the point of view of autism.  Not surprisingly, if you have Cav1.2 dysfunction in the brain, it might very well occur elsewhere.

There are little genetic errors called Single Nucleotide Polymorphisms, or SNPs.  In the CACNA1C gene there are 12,932 known SNPs.  Some of the most common ones are associated with autism, bipolar and schizophrenia.

You can look up this gene, or any other one, and see for yourself.

Genecards database

If you read the gene description above, the idea that heart disease is comorbid with autism is no surprise. 

The lower red arrow points at hypokalemic periodic paralysis.  This has appeared many times on this blog, along with Hypokalemic Sensory Overload.  I discovered long ago that there is a potassium ion channel dysfunction in autism; it appears to be behind the odd sensory overload experienced by many with autism and also in some people with ADHD.  What is interesting is that this dysfunction co-occurs with CACNA1C dysfunctions.

Cancer and Autism

The science behind cancer is complex and so as not to research it in vain, it is useful to know that there is solid evidence linking autism and cancer.

The following study of 8,438 people with autism, compared their incidence of cancer with the incidence in the general population

To understand the jargon first read this excerpt from a fact sheet on cancer statistics:

Fact Sheet: Explanation of Standardized Incidence Ratios

How an SIR Is Calculated

The expected number is calculated by multiplying each age-specific cancer incidence rate of the reference population by each age-specific population of the community in question and then adding up the results. If the observed number of cancer cases equals the expected number, the SIR is 1. If more cases are observed than expected, the SIR is greater than 1. If fewer cases are observed than expected, the SIR is less than 1.

Examples:

60 observed cases / 30 expected cases: the SIR is 60/30 = 2.0

Since 2.0 is 100% greater than 1.0, the SIR indicates an excess of 100%.

45 observed cases / 30 expected cases: the SIR is 45/30 = 1.5

Since 1.5 is 50% greater than 1.0, the SIR indicates an excess of 50%.

30 observed cases / 30 expected cases: the SIR is 30/30 = 1.0

A SIR of 1 would indicate no increase or decrease.

Here is the autism study:-

Risk of Cancer in Children, Adolescents, and Young Adults with Autistic Disorder

Objectives

To investigate whether individuals with autism have an increased risk for cancer relative to the general population.

Study design

We enrolled patients with autistic disorder from the Taiwan National Health Insurance database in years 1997-2011. A total of 8438 patients diagnosed with autism were retrieved from the Registry for Catastrophic Illness Patients database. The diagnosis of cancers was also based on the certificate of catastrophic illness, which requires histological confirmation. The risk of cancer among the autism cohort was determined with a standardized incidence ratio (SIR).

Results

During the observation period, cancer occurred in 20 individuals with autism, which was significantly higher than a total number of expected cancers with a SIR estimate of 1.94 (95% CI 1.18-2.99). The number of cancer in males was greater than the expected number with a SIR of 1.95 (1.11-3.16), but no excess risk was found for females with a SIR of 1.91 (0.52-4.88). Cancer developed more than expected in individuals age 15-19 years with the SIR of 3.58 (1.44-7.38), but did not differ in other age range groups. The number of cancers of genitourinary system was significantly in excess of the expected number (SIR 4.15; 95% CI 1.13-10.65), and increased risk was found in ovarian cancer with SIR of 9.21 (1.12-33.29).

Conclusions

Our study demonstrated that patients with autistic disorder have an increased risk of cancer.

So, overall, the risk of all cancers is about twice as high if you have autism.  

Certain cancers are particularly high risk and understanding why this is the case might lead to a better understanding of the “pathways” leading to some types of autism. Due to the rarity of some cancers, like ovarian, one might need to validate the result; note the (1.12-33.29) range for ovarian cancer.

Rather than worry about this risk, we should use these observations to understand and treat autism.

Just as we can counter the elevated risk of heart disease we can do the same for cancer.

Clearly the cancer pathways that will soon be appearing in this blog are relevant to autism.  But in the meantime anyone can reduce their cancer risk by ensuring a high level of antioxidants in their body.  People at higher risk are those with low levels of antioxidants, which include almost all older people and people of all ages with autism.

A vast wealth of information already exists showing the chemo-protective effect of antioxidants.  Cancer clearly generally results from multiple hits, and you may be unlucky to have a single gene that “ups” your risk.  By upping your antioxidant intake you can slash one risk, in this multiple step process.

It does not seem to matter which potent antioxidant you take, but you do need enough of it.  They are all slightly different and most likely a mix of several will yield the best result.

My current favourites are:-

·        NAC (N-acetyl cysteine)

·        ALA (Alpha lipoic acid) - Nrf2 activator

·        Sulforaphane – Nrf2 activator

·        Cocoa Flavanols

·        Lycopene (cooked tomato)

These should reduce both the risk of cancer risk and heart disease.

Other antioxidants mentioned in this blog include:-

·        L-Carnosine

·        Silibinin – Nrf2 activator

·        Selenium

One should be aware that avoiding cancer and treating an existing cancer are different tasks.  Once a cancer has developed, some antioxidants can interfere with the body’s own response mechanism.

My focus is preventative “medicine”.

We saw in an earlier post how children at risk of developing asthma could be identified by their atopic dermatitis.  By treating these children with a cheap mast cell stabilizer called Ketotifen, a trial showed how it was possible to avoid the onset of asthma.

I suspect that the same thing might be possible with epilepsy.  We saw in an earlier post that the first epileptic attack make a (epigenetic?) change, and thereafter there is a greatly increased risk of future seizures.

Other interesting preventative interventions, include statins to avoid Parkinson’s disease and Verapamil to avoid the onset of Type II diabetes.

I did explain all this to the European Medicines Agency some months ago, the idea of treating the comorbidities of autism BEFORE they occur.  Perhaps an idea before its time?

Statins for Cancer and Autism? Another case for PTEN?

When I first started this blog and my investigation into the biology of autism, I did shy away from the more complex areas like genetics.  I assumed that this would be best left to the “experts” and be beyond the powers of those without fancy laboratory tools.

My literature review took me early on to oxidative stress and then neuroinflammation.  I deduced that in the case of neuroinflammation, it might be possible to control inflammatory cytokines using statins.  I also noted the use of statins in TBI (Traumatic Brain Injury). I thought it would be harmless to do a quick trial, not really expecting anything to happen; but it did, and from the very first dose.

The literature is full of references to lipid dysfunction in autism and one large sub-group in autism is known to have high cholesterol.  Cholesterol and inflammation are now known to go hand in hand.  When inflammation is present, the body can react by laying down a protective layer of cholesterol.  The problem is that too much cholesterol is not good for you either.  The real culprit is not the cholesterol, it is the inflammation.

If you are in the high cholesterol autism group, a cholesterol lowering drug that is also anti-inflammatory may be “just what the doctor ordered”.

Be warned that another subgroup in autism has very low cholesterol.  In a study at the Kennedy Krieger Institute, 19% of children had extremely low cholesterol, meaning lower than 99% of typical children.

Autism: New Study Discovers Statistically Significant Link Between Abnormally Low Cholesterol Levels And Autism Spectrum Disorders

There is a rare condition, leading to autism called Smith-Lemli Opitz syndrome (SLOS).  SLOS is caused by a mutation in an enzyme involved in cholesterol synthesis; the resulting biochemical characteristics may be predictable. Most patients have lowered plasma cholesterol levels.

Since cholesterol testing is cheap and widely available, you can easily determine which group you are in.

This post is for the high cholesterol cohort.

Note well how meaningless a figure for the "average cholesterol level" in autism would be. In the autism literature they frequently take the mean average for all data, thus missing the point. 

Why Statins for Autism?

My initial logic was that since inflammatory markers are often elevated in autism and that oxidative stress and inflammation are self-reinforcing, it would be logical to find an effective anti-inflammatory agent.  Steroids might fit the bill, but they cause plenty of side effects in long term use; their short term use in autism can be remarkably effective.  So I looked further, and having screened the literature, ended up convincing myself of the potential of statins.  Read all about cytokine storms in the old posts, if you are interested.

I choose  Atorvastatin (also known as Lipitor or Sortis), since it freely crosses the blood brain barrier (BBB) and is safely used my tens of millions of people around the world.

It worked.

Explaining Statin Therapy to others.

The most important thing is to have a therapy that works;  but then you have to explain it to others.

I was recently explaining it again to a doctor relative, who was asking how I could be sure it works.  I explained that every time I stop using it, within a day behaviour changes in the same predictable way.  It is as if people with autism have an inhibitory barrier; there are things they can do, want to do, but something is blocking them from doing them.

Examples are numerous.  Speech being one.  Plenty of kids with autism are non-verbal, everything is physically functional, yet they do not talk, even when they want to communicate.

At the age Monty, now aged 11 with ASD, tried the statin he was relatively verbal.  The immediate change in him was that he suddenly started to play the piano, by himself.  Odd it may sound.

In his earlier years he would often get “stuck”.  He would be upstairs and unable to come downstairs, somebody had to go up and get him.

When I now stop the statin, he will again get “stuck”.  He will stand in the kitchen and want to leave and just say “go that way”, but not move.  You have to take his hand, so that he can “go that way”.

A Better Explanation?

Now I have another explanation of why statins may be effective in one large sub-group of autism.

Statins up-regulate a known key dysfunctional autism gene, and protein, called PTEN.  I mentioned PTEN in a previous post, since one chemical released by eating broccoli also up-regulates PTEN.

Science has already shown that things that down-regulate PTEN (like seizures) make autism worse.

The full science behind PTEN will come in a later post.

Statins and Cancer

Regular readers will recall that PTEN is also a tumor suppressor gene and is therefore a target for cancer research.

Thinking the way I do, I know that statins increase PTEN and that this should slow cancer growth.  Hundreds of millions of people take statins and many millions get cancer, so what about people on statins getting cancer?

A quick check on google and there we have studies showing that people on statins get less cancer and that in common cancers like that of the prostate, the outcome is better when statins are taken.

Now this is not a cancer blog, but you do not have to dig very deep to uncover a wealth of supporting evidence.

Statins and Prostate Cancer: Novel, Encouraging Study

Postoperative statin use and risk of biochemical recurrence following radical prostatectomy: results from the Shared Equal Access Regional Cancer Hospital (SEARCH) database

Conclusion

In this retrospective cohort of men undergoing RP, post-RP statin use was significantly associated with reduced risk of BCR. Whether the association between post-RP statin use and BCR differs by race requires further study. Given these findings, coupled with other studies suggesting that statins may reduce risk of advanced prostate cancer, randomized controlled trials are warranted to formally test the hypothesis that statins slow prostate cancer progression.

Statins and prostate cancer (Harvard Medical School)

Statins could reduce the risk of breast cancer

Statins could reducethe risk of pancreatic cancer

Statin use and risk of liver cancer: an update meta-analysis

 Conclusions This meta-analysis suggests that statin is associated with a significant risk reduction of liver cancer when taken daily for cardiovascular event prevention. However, this preventive effect might be overestimated due to the exposure period, the indication and contraindication of statins and other confounders. Statins might be considered as an adjuvant in the treatment of liver cancer.

Statins and PTEN

I am no cancer expert, but I can read the literature and the evidence is pretty compelling to me.  It is not enough, however, for doctors to prescribe statins to avoid cancer.  They are so busy prescribing statins to over 50s for other reasons, it does not really matter.

We came across PPAR previously.  PPAR gamma is a pathway to treat type 2 diabetes and the old type 2 diabetes drug Pioglitazone has shown promise in an autism study.

PPAR alpha, beta and gamma in Autism, Heart Disease and Diabetes

 Effect of pioglitazone treatment on behavioral symptoms in autistic children

At that time I was more interested in PPAR-alpha, due to its role in mast cell stabilization.

It is via PPAR-gamma, that statins up regulate PTEN.

You do not want to overdo it, because at very high doses too much PPAR gamma protein will be produced and you risk causing type 2 diabetes.

Low doses of statins are trouble free for most people, but high doses are associated with increased risk of diabetes and all kinds of aches and pains.

The statin effect in autism does not increase with higher doses, only a small dose is required.

Regulation of the PTEN promoter by statins and SREBP.

Abstract

Germline mutations in the tumor-suppressor gene PTEN predispose to heritable breast cancer. The transcription factor peroxisome proliferator-activated receptor-gamma (PPARgamma) has also been implicated as a tumor suppressor pertinent to a range of neoplasias, including breast cancer. We previously demonstrated that lovastatin may signal through PPARgamma and directly upregulate PTEN expression at the transcriptional level. In our current study, we show that simvastatin, pravastatin and fluvastatin can induce PTEN expression in a dose-dependent manner. This resulted from an increase in PTEN mRNA indicating transcriptional upregulation. In addition, we observed, for the first time, that upregulation of sterol response element-binding protein (SREBP), known to induce PPARgamma expression, can increase PTEN expression. Using reporter assays, we observed that both the statins and SREBP could specifically induce PPARgamma-mediated transcription. However, the statins do not appear to signal through SREBP. Furthermore, our results indicate that SREBP utilizes PPARgamma's transcriptional activity to induce PTEN transcription, whereas the statins signal through PPARgamma's protein activity to upregulate PTEN expression. Overall, our observations suggest that statins signal through another transcription factor, in a PPARgamma-dependent manner, which in turn induces PTEN transcription. We, therefore, studied the full-length PTEN promoter through serial deletion reporter assays and electromobility shift assays and identified a region between -854 and -791 that binds an as-yet-unidentified transcription factor, through which the statins induce PTEN expression. Since PTEN is constitutively active, our data indicate it may be worthwhile to examine statin and SREBP stimulation as mechanisms to increase PTEN expression for therapeutic and preventative strategies in cancer, diabetes mellitus and cardiovascular disease

PTEN dysfunction in Cancer and Autism

I will cover this point in more detail in the post on PTEN, but note that the PTEN gene dysfunctions found in 10% of people with autism are generally different to the ones found in cancer.  We also have the difference between whether the PTEN gene is mutated or there is PTEN loss.

There should be two identical copies of the PTEN gene. When one copy is mutated, the protein it produces was found to inhibit the protein produced by the good copy. In other cases, one copy of the PTEN gene is OK, but the other got deleted.   This turned out to be better than having one mutant version.

Different mutations in PTEN are linked to different outcomes.  The known autism mutations are called H118P, H93R and H123Q.  If you have a C124S mutation you would be at risk of something called thyroid follicular carcinoma and not autism.

It is all very complicated and I have to say some conclusions in the research are contradictory.

But it is reported that about 10% of people with autism have an identifiable PTEN mutation.  I am more interested in whether PTEN is an interesting protein in the other 90%.

We saw in the fragile X research that even though this affects only 1% of cases with autism, some experimental therapies for fragile X worked on people with autism, but without fragile X.  At the time I thought that very odd.

My assumption is that PTEN is interesting for more than the 10%.

Conclusion

So there are now 2 plausible reasons why statin therapy may be effective in people with classic autism and elevated cholesterol:-

·        Reduction in inflammatory cytokines 

·        Up-regulation of PTEN

Maybe it is both.

It may be that in people with autism and low cholesterol, and so not suited to statins, they may also have low levels of PTEN.

We saw in a recent post that when you eat fresh broccoli in addition to Sulforaphane, you also produce Indole-3-carbinol (I3C).   I3C also up-regulates  PTEN.

Using Peter logic, if statins have an immediate effect then quite likely so would I3C.

Whatever Next?

Well, for those few of you who have discovered the “magical” beneficial effects of mast cell stabilizers, like Verapamil and Cromolyn Sodium, on both autistic behaviours and severe allergies, here is a preview of what is coming next:-

PPAR-gamma modulates allergic inflammation through up-regulation of PTEN

Recent studies have indicated that PPAR-gamma plays an important role in anti-inflammatory responses and that PPAR-gamma signaling is associated with regulation of PTEN expression. It is known that up-regulation of PTEN expression reduces asthmatic pathogenesis.

These findings suggest that PPAR-gamma uses PTEN to modulate asthmatic responses The signaling mechanism by which stimulation of PPAR-gamma with the agonists regulates PTEN expression as well as Akt phosphorylation remains to be lucidated. However, our results agree with the observation that the anti-inflammatory action of PPAR-gamma agonists is mediated via up-regulation of PTEN.

In other words, increasing PTEN minimizes allergies.  Perhaps, via feedback loops, increasing allergies reduces PTEN?

Seizures also reduce PTEN.

Reduced PTEN leads to increased autistic behaviours.

Not surprisingly we will come back, yet again, to mast cells.

For us, it really does seem that PTEN is a key piece in the puzzle;  but a puzzle with a solution.

How to make Sulforaphane (Broccoli) at home

I hope he took his Sulforaphane

This month thousands of runners braved thick smog at the Beijing marathon, with some even donning masks as air pollution soared to 16 times the maximum recommended level.

Johns Hopkins have been trialing their Sulforaphane in China as a therapy to counter the health effects of air pollution.

It was proposed that the potent anti-oxidant and chemoprotective protective properties of Sulforaphane would be a cheap way to protect the health of people living in highly polluted environments.

Broccoli sprout beverage enhances detoxification of air pollutants in clinical trial

 or the actual study:-

Rapid and Sustainable Detoxication of Airborne Pollutants by Broccoli Sprout Beverage: Results of a Randomized Clinical Trial in China

Abstract

Broccoli sprouts are a convenient and rich source of the glucosinolate, glucoraphanin, which can generate the chemopreventive agent, sulforaphane, an inducer of glutathione S-transferases (GST) and other cytoprotective enzymes. A broccoli sprout–derived beverage providing daily doses of 600 mmol glucoraphanin and 40 mmol sulforaphane was evaluated for magnitude and duration of pharmacodynamics action in a 12-week randomized clinical trial. Two hundred and ninety-one study participants were recruited from the rural He-He Township, Qidong, in the Yangtze River delta region of China, an area characterized by exposures to substantial levels of airborne pollutants. Exposure to air pollution has been associated with lung cancer and cardiopulmonary diseases. Urinary excretion of the mercapturic acids of the pollutants, benzene, acrolein, and crotonaldehyde, were measured before and during the intervention using liquid chromatography tandem mass spectrometry. Rapid and sustained, statistically

significant (P _ 0.01) increases in the levels of excretion of the glutathione-derived conjugates of benzene (61%), acrolein (23%), but not crotonaldehyde, were found in those receiving broccoli sprout beverage compared with placebo. Excretion of the benzene-derived mercapturic acid was higher in participants who were GSTT1-positive than in the null genotype, irrespective of study arm assignment.

Measures of sulforaphane metabolites in urine indicated that bioavailability did not decline over the 12-week daily dosing period. Thus, intervention with broccoli sprouts enhances the detoxication of some airborne pollutants and may provide a frugal means to attenuate their associated long-term health risks.

Now this blog is not about pollution, but you might be interested to know that such pollution not only increases cancer risk (plus respiratory diseases, of course) but also increases the incidence of autism.

  

Study Suggests Link Between Air Pollution Exposure And Autism

How to make Sulforaphane at Home

Hopefully you can now see the potential benefits of Sulforaphane.  As I said in the earlier post, twenty years has passed since Johns Hopkins discovered Sulforaphane and there have been numerous studies and experiments done.  What follows is just my synthesis and conclusions of that work.

1.     Eating Broccoli

Broccoli does contain glucosinolate and the required enzyme myrosinase.  If you eat copious amount of raw broccoli or very lightly cooked (steaming for 2 minutes) you will produce Sulforaphane in your body.  The amount required would be literally pounds/kilos each and every day, to come close the therapeutic doses.

Frozen broccoli has no active myrosinase and over-cooked broccoli has no myrosinase.

Clever tricks developed to get round this include:-

·        Eating a small piece of raw broccoli (to provide  myrosinase) with your cooked broccoli

 ·        Adding a tiny amount of daikon radish to frozen broccoli.  This is really a great idea, since only 0.25% Daikon is needed, you get 99.75% broccoli and will never even notice or taste the daikon.  The idea is that this should be done by the food processor when they freeze the broccoli, you would not do anything at home.

  

Modifying the processing and handling of frozen broccoli for increased sulforaphane formation.

Abstract

Frozen broccoli can provide a cheaper product, with a longer shelf life and less preparation time than fresh broccoli. We previously showed that several commercially available frozen broccoli products do not retain the ability to generate the cancer-preventative agent sulforaphane. We hypothesized that this was because the necessary hydrolyzing enzyme myrosinase was destroyed during blanching, as part of the processing that frozen broccoli undergoes. This study was carried out to determine a way to overcome loss of hydrolyzing activity. Industrial blanching usually aims to inactivate peroxidase, although lipoxygenase plays a greater role in product degradation during frozen storage of broccoli. Blanching at 86 °C or higher inactivated peroxidase, lipoxygenase, and myrosinase. Blanching at 76 °C inactivated 92% of lipoxygenase activity, whereas there was only an 18% loss in myrosinase-dependent sulforaphane formation. We considered that thawing frozen broccoli might disrupt membrane integrity, allowing myrosinase and glucoraphanin to come into contact. Thawing frozen broccoli for 9 h did not support sulforaphane formation unless an exogenous source of myrosinase was added. Thermal stability studies showed that broccoli root, as a source of myrosinase, was not more heat stable than broccoli floret. Daikon radish root supported some sulforaphane formation even when heated at 125 °C for 10 min, a time and temperature comparable to or greater than microwave cooking. Daikon radish (0.25%) added to frozen broccoli that was then allowed to thaw supported sulforaphane formation without any visual alteration to that of untreated broccoli.

2.     Eating Broccoli Sprouts

It was shown that broccoli seeds and broccoli sprouts (5 day old broccoli) contain highly concentrated amounts of glucosinolate and the required enzyme myrosinase.  It is reported to be about 20 times higher in sprouts than full grown broccoli.

Broccoli sprouts are eaten uncooked and so no myrosinase is lost in food preparation.

Following all the Johns Hopkins research and commercialization, in many parts of the world you can readily buy fresh broccoli sprouts, many sold by companies licensed by the company run by the son of the original researcher at John Hopkins.

It was reported that that original lead researcher tries to regularly eat 4oz (120g) a week of broccoli sprouts, which is not so much.

However if you want to achieve the therapeutic doses in the clinical trials this will not be enough.

Trials used between 50 and 150 micromoles of Sulforaphane.

Rather unhelpfully they do not equate this to a measure accessible to lay people. 

If you recall your high school chemistry just go to Wikipedia and look up Sulforaphane:

Molecular formula	C6H11NOS2
Molar mass	177.29 g/mol

To convert to grams you just multiply by 177.29.

So the trials used dosages between 8.8 mg and 26.6 mg of sulforaphane.

Most of these trials are in adults and most people reading this blog are interested in treating children, so let’s work with the figure of 8mg of sulforaphane.

Bioavailability and inter-conversion of sulforaphane and erucin in humansubjects consuming broccoli sprouts or broccoli supplement in a cross-over study design

Abstract

Broccoli consumption may reduce the risk of various cancers and many broccoli supplements are now available. The bioavailability and excretion of the mercapturic acid pathway metabolites isothiocyanates after human consumption of broccoli supplements has not been tested. Two important isothiocyanates from broccoli are sulforaphane and erucin. We employed a cross-over study design in which 12 subjects consumed 40 grams of fresh broccoli sprouts followed by a 1 month washout period and then the same 12 subjects consumed 6 pills of a broccoli supplement. As negative controls for isothiocyanate consumption four additional subjects consumed alfalfa sprouts during the first phase and placebo pills during the second. Blood and urine samples were collected for 48 hours during each phase and analyzed for sulforaphane and erucin metabolites using LC-MS/MS. The bioavailability of sulforaphane and erucin is dramatically lower when subjects consume broccoli supplements compared to fresh broccoli sprouts. The peaks in plasma concentrations and urinary excretion were also delayed when subjects consumed the broccoli supplement. GSTP1 polymorphisms did not affect the metabolism or excretion of sulforaphane or erucin. Sulforaphane and erucin are able to interconvert in vivo and this interconversion is consistent within each subject but variable between subjects. This study confirms that consumption of broccoli supplements devoid of myrosinase activity does not produce equivalent plasma concentrations of the bioactive isothiocyanate metabolites compared to broccoli sprouts. This has implications for people who consume the recommended serving size (1 pill) of a broccoli supplement and believe they are getting equivalent doses of isothiocyanates.

Following consumption of 40 g of alfalfa sprouts or 6 placebo pills, no SFN or ERN metabolites were detected in plasma or urine from the four subjects in the control group (Figure 1). In contrast subjects who consumed 40 g of broccoli sprouts (150 and 71 μmoles glucoraphanin and glucoerucin, respectively) or 6 supplement pills (121 and 40 μmoles glucoraphanin and glucoerucin, respectively) had considerable amounts of SFN and ERN metabolites in both plasma and urine.

In 12 hours about 145 micromols of SFN and ERN were excreted in urine.  From the chart it looks likes SFN:ERN is about 2:1.  So assume about 95 micromols of SFN (sulforaphane).

In the following study using frozen sulforaphane made at Johns Hopkins about 85 micromols were excreted in 12 hours 

Bioavailability of sulforaphane from two broccoli sprout beverages: Results of a short term, cross-overclinical trial in Qidong, China

In the Johns Hopkins trial above the dosage was 800 μmol of glucoraphanin in GRR (the blue lines above) and 150 μmol of sulforaphane in SFR beverages (green lines above). The drugs were mixed with mango juice and water.

We compare the green line with the earlier study and see that 40 g of sprouts is a similar dose to 150 μmol of Johns Hopkins sulforaphane.

Now I did ask Johns Hopkins how many grams of broccoli sprouts yields 50 μmol of Johns Hopkins sulforaphane.  They did reply and said that the level varies among sprouts and so it is impossible to say.

We have seen in this blog, to date, that while nothing is 100% certain in autism or autism therapies, once you have exceeded a certain level of probability, it is worth giving things a try.  If you wait for 100% certainty, you will never move.

So while you will never know exactly how much sulforaphane is in your sprouts, it does look like a fair estimate is 3.8 μmol /g.

So if you want 50 μmol, then you would need to eat about 13g of sprouts a day.

To achieve the adult dose of 150  μmol you would need to eat 40g of sprouts a day.

As a double check compare this to what the original lead researcher is reporting to be taking, for preventative therapy.  He takes 4 oz. a week.  This is 113.4g or 16g a day.

This dose appears not to have harmed him, and he is now 91 years old!

Paul Talalay

Paul Talalay was born in Germany , but emigrated to England with his family in 1933, shortly after the Nazi Party came to power. He was educated at Bedford School and, in 1940, he travelled to the United States to enter Massachusetts Institute of Technology where he majored in biology

Talalay's career has been devoted to cancer research and the achievement of early protection against cell damage. A pioneer in the field of chemoprotective research strategies, Talalay and his colleagues devised simple cell culture methods for detecting phytochemicals which appear to boost enzymes that detoxify carcinogens in the body. This work led to the isolation of sulforaphane, found in broccoli, as a potent inducer of detoxifying phase two enzymes. These findings, published in 1992,  attracted worldwide attention as a major breakthrough in understanding the potential link between cruciferous vegetable consumption and reduced cancer risk.

Since I have no signs of any other Germans appearing on my Dean’s List and there are already plenty of Americans, he goes down as a German.  Nikola Tesla had the same problem, with four countries claiming him as their own (USA, Austria, Croatia and Serbia).

 

3.     Mixing Daikin Powder with Broccoli Powder

Many people do not like eating broccoli.  I do suggest you try eating it raw; it really is not bad at all, and much better than the soggy, over cooked, variety.

For those preferring powders and pills, the third method involves mixing freeze dried Daikin Radish with freeze dried broccoli.

It turns out that while the myrosinase in broccoli is not heat or cold stable, the daikon radish root is a good source of heat stable myrosinase.  This radish is commonly used in Japan and is available cheaply in freeze dried form.

This is the powder that was proposed to be added to Frozen broccoli, so that it would be a source of sulforaphane.

Broccoli powder is produced in large volumes for the supplement industry, which package it in capsules and sell it on to you.

Why nobody thought of including active myrosinase from daikon radish is beyond me.  It is not expensive.

There appears to be one broccoli supplement that does actually do what it says on the label and produce some  sulforaphane.  Perhaps it includes some Daikin powder ?  It was tested in the US.

That supplement is made in Australia.  It is not cheap.

It is claimed that:-

A 1-gram serve of EnduraCell powder is equivalent to 12 grams of fresh sprouts (with their sulforaphane inhibitors deactivated) and contains 30mg of Glucoraphanin that yields 12 mg Sulforaphane.

Research has shown that generally broccoli supplements do not perform, perhaps this one is different?

4.     Combining Broccoli Sprouts with Broccoli Powder

Since broccoli sprouts, like daikin radish, contains copious amounts of myrosinase, you could also combine fresh broccoli sprouts with broccoli powder.  This has actually been studied in research projects and does work.

Sulforaphane absorption and excretion following ingestion of a semi-purified broccoli powder rich in glucoraphanin and broccoli sprouts in healthy men.

Abstract

Sulforaphane (SF) is a chemopreventive isothiocyanate (ITC) derived from the myrosinase-catalyzed hydrolysis of glucoraphanin, a thioglucoside present in broccoli. Broccoli supplements often contain glucoraphanin but lack myrosinase, putting in question their ability to provide dietary SF. This study compared the relative absorption of SF from air-dried broccoli sprouts rich in myrosinase and a glucoraphanin-rich broccoli powder lacking myrosinase, individually and in combination. Subjects (n=4) each consumed 4 meals consisting of dry cereal and yogurt with 2 g sprouts, 2 g powder, both, or neither. Blood and urine were analyzed for SF metabolites. The 24 h urinary SF recovery was 74%, 49%, and 19% of the dose ingested from broccoli sprouts, combination, and broccoli powder meals, respectively. Urinary and plasma ITC appearance was delayed from the broccoli powder compared to the sprouts and combination. A liver function panel indicated no toxicity from any treatment at 24 h. These data indicate a delayed appearance in plasma and urine of SF from the broccoli powder relative to SF from myrosinase-rich sprouts. Combining broccoli sprouts with the broccoli powder enhanced SF absorption from broccoli powder, offering the potential for development of foods that modify the health impact of broccoli products.

Conclusion

More good news is that when you make sulforaphane in the above fashion, you also make some other interesting substances; one of these is Indole-3-carbinol (I3C).  I3C itself has some extremely interesting properties for both cancer and autism.  I3C up-regulates a protein called PTEN, encoded by the PTEN gene.  PTEN is dysfunctional in autism and, in general terms, may need to be up-regulated.  Indole-3-carbinol is one of the few, safe, known, ways to up-regulate PTEN.  PTEN is also a tumor suppressor gene and so in people with some cancers, up-regulating PTEN will slow cancer progression.

Anyway, it really does look like broccoli may be good for cancer and autism.

Bon Appetit!