Natural Stacks Acetylcholine Brain Food: Impressive HRV Booster?

The concept of heart rate variability (HRV) has gained much attention in the biohacking universe, and for legitimate reasons. Higher variability in the rhythm of the body’s blood pump has been associated with numerous health benefits. 

As such, identifying hacks to positively influence HRV is fertile ground. Heart rate and corresponding heart rate variability are heavily influenced by the primary connection between the brain and heart: the vagus nerve.

As a key component in the parasympathetic nervous system (PNS) - which is involved with relaxation - the vagus nerve utilizes chemical messengers called neurotransmitters to relay instructions between these critical organs. 

In other words, neurotransmitters are signaling substances in the nervous system.

The primary transmitter of the PNS is acetylcholine (ACh). Given this understanding, it has been hypothesized that oral supplementation of ACh modulators may impact HRV. 

This article explores the application of these concepts and reviews the first of a handful of ACh-supporting supplements: Acetyl-choline Brain Food by Natural Stacks. 

The question I'm trying to answer is whether Acetyl-choline Brain Food is capable of improving heart rate variability. If heart rate variability can be improved, then, arguably, overall relaxation and other health-related variables like sleep quality and cognitive performance may also improve.


Let’s consider my experience with this supplement:


Natural Stacks Acetyl-Choline Brain Food Summary


  • Scientific research implies modulating Ach pathways may impact HRV
  • Formula contains notable Ach precursors and modulators supported by science


  • No human trials showing efficacy on ACh levels or HRV status
  • Unable to measure direct impact on ACh levels 

My Conclusion:

Acetyl-choline Brain Food by Natural Stacks did not meaningfully impact heart rate variability or enhance mental clarity in me. This outcome warrants further exploration, however, and more of such tests are coming.


Can You Improve Heart Rate Variability with Acetylcholine Supplements?

Let's first take a step back:

This is the first part of a series I'm writing on whether acetylcholine supplements can improve heart rate variability. Other installments will test other acetylcholine supplements and consider whether they affect my heart rate variability.

The first question is how exactly heart rate variability is related to acetylcholine. Let's find out: 


So let's explore acetylcholine's relationship to heart rate variability:

Enhancing HRV is commonly held as a good thing in pursuit of health and longevity. Before we dive into how ACh supplements may influence HRV, I encourage readers to refresh themselves on the concepts of HRV. 

For that purpose, this website has done a deep dive into the topic previously: How to Easily Measure Heart Rate Variability.

During my experiment, HRV measurements were taken throughout the duration of the experiment through the Biostrap. 

The following link helps you understand what and how that Biostrap device measures HRV: Biostrap Review: An Essential Wearable for the Serious Biohacker?

(A second installment on the Biostrap is Biostrap EVO: An Excellent Evolution In Fitness Wearables.) 

Having covered HRV and how to measure it, let’s dig into the “why” of HRV.

Science shows that HRV is a marker for sympathetic (fight or flight)/parasympathetic (rest & digest) dominance in the nervous system (1). 

(Remember: the Sympathetic Nervous System can be abbreviated as "SNS", the parasympathetic counterpart as "PNS")

Academicians and clinicians suspect that the modern western lifestyle generally shifts the balance towards sympathetic dominance (2). Coupled with generally declining sleep quality and duration (highly recommend Matt Walker’s “Why We Sleep” book for more on sleep), the stage is set for focusing on HRV patterns and how to influence them. 

If we agree that HRV is a marker for SNS/PNS balance, and vetted science shows this is true, then logic suggests that enhancing PNS function/activity should correspond to detectable improvements in HRV. 

As noted in the aforementioned HRV review blog, excessive physical activity (high SNS activity) leads to decreased HRV measures whereas adequate rest (high PNS activity) can improve HRV outcomes.  

Stress reduction and consistent sleep habits are the best HRV improvement strategies presently known (3,4).

A question biohackers - like me - want to know is “can HRV be improved through supplementation?”  

Heart rate and HRV are primarily governed via the PNS through the vagus nerve (5). Science has elucidated that acetylcholine is the primary chemical messenger in this system (5). 

One critical assumption in the hypothesis does come into play here: does low and/or impeded ACh levels impact HRV? Let’s have a closer look at ACh biochemistry to gain some insight on this topic. 

ACh is synthesized in the nervous system from choline and acetyl-CoA. Acetyl-CoA is involved with breaking down carbohydrates, proteins, and fats, in your "mitochondria". Mitochondria are the energy-producing "factories" of your cells. However, Acetyl-CoA is also a building block of ACh!

Enzymes that facilitate the creation and deconstruction of ACh are present in high levels in and around the neurons that utilize ACh. Both the building blocks as well as the enzymes are focal points for supplements and drugs that seek to influence ACh function in the body.

But's let's zoom out and simplify my message:

The investigation reported in this and the following installments of this series lead to a research question. The question I'm trying to answer is: "do combinations of ACh building blocks and ACh enzyme modulators impact my HRV?"

In other words, "do supplements that likely affect acetylcholine levels influence my HRV?"

If you're an astute reader you may ask “why not just measure ACh levels?” Indeed, an investigation exploring relationships between central ACh levels and HRV measures would be instrumental to determine if aberrations in ACh neurotransmission impacts HRV. 

Alas, there is no clinically useful measure of central ACh levels to make this determination.  The acetylcholinesterase enzyme is responsible for degrading ACh following post-synaptic release. Its ubiquitous nature ensures that freely-circulating ACh is non-detectable in biological fluids.  

In plain English, after your nervous system has used acetylcholine it won't be detectable in your saliva or blood or other fluids. Hence, I'm measuring HRV as a proxy for acetylcholine function.

In the absence of a tool to measure ACh levels in the human body, the hypothesis of enhancing HRV through ACh supplementation relies upon the assumption that in some individuals, low HRV may be in part due to low endogenous ACh, and that supplementing with ACh building blocks and enzyme modulators may improve HRV through enhanced ACh/PNS communication. 

I do want to apologize for the exhaustive walkthrough of the logic behind this idea. But I feel this background information is necessary to understand the potential implications. 


The outcomes of my experiment could be a big deal, as many of the pharmaceuticals used to address neuropsychological disorders, such as depression, anxiety, and insomnia, work by similar mechanisms. 

Altering your neurotransmission (communication between nerve cells) to impart a clinical response is frequently the goal in medicine.

And given that you now have a basic understanding of acetylcholine, let’s move on to the supplement review:


Scientific Review Of Acetyl-Choline Brain Food Ingredients

Brain food contains 5 separate ingredients in total. Below, I've reviewed all of the ingredients in the Acetylcholine Brain Food supplement from Natural Stacks:


1. White Peony Extract (Paeonia lactiflora) (root) (Standardized to 161 mg paeoniflorin), 383 mg

The use of peony extract dates back to ancient Greece, where it had reported applications for snake bites.  Recent research has documented that paeoniflorin, one of many active ingredients in the extract, inhibits the ACh-degrading enzyme, acetylcholinesterase  (6). The premise is straightforward: slow down or block the enzyme that cleans ACh out of the synapse and there will be more ACh present to activate ACh receptors.

No published studies on the use of Peony extracts for the treatment of ACh deficiency disorders exist, so its supplement use is based on a hypothetical belief that oral ingestion will impart some effect on acetylcholinesterase enzymatic activity. 


2. Vitamin B1 (as thiamine HCL), 64 mg

Thiamine - or vitamin B1 - has been established as one of three critical enzymatic cofactors in the synthesis of Acetyl-CoA (7). 

Here again, the premise is simple: Ensure adequate supply of all the components necessary to construct the building blocks of the target molecule, in this case, the Acetyl-CoA portion of the Acetylcholine molecule.

Important to note here, one does not obtain Acetyl-CoA from the diet. The compound is also produced by cells in the body from already existing building blocks. 


3. Acetyl-L-Carnitine HCL, 319 mg

This ingredient may serve dual purposes in this supplement.  The formula is marketed as “daytime support for mental clarity”. Claims exist regarding the function of carnitine as a “mental booster”, the likes of which appear to be non-ACh mediated and therefore out of the scope of this investigation. 

Of relevance, per the manufacturer’s website, the “acetyl group” of the ingredient is naturally cleaved off the molecule in the body, which is then hypothetically available as a building block for the synthesis of ACh. I've not identified scientific studies that describe Acetyl-L-Carnitine’s oral ingestion as a primary acetyl group donor for ACh. 


4. Pantethine (Pantesin), 287 mg

Similar to thiamine/vitamin B1, this form of vitamin B5 is involved in the acetyl-CoA synthesis pathway by increasing cellular CoA levels (8) The “Pantesin” branded form of vitamin B5 in this supplement is claimed to be highly absorbable and bioactive.

For more info on this ingredient, see


5. Alpha GPC (L-glycerylphosphorylcholine), 192 mg

This ingredient is the industry standard “choline donator”. Per the Natural Stacks website, Alpha GPC reportedly has the greatest amount of scientific evidence supporting efficacy and adsorption. Rat studies have verified that oral ingestion of Alpha GPC in fact leads to increased central ACh levels (9).


My Final Verdict On The Ingredients:

The Natural Stacks Acetyl-Choline Brain Food formula checks a number of boxes regarding pathways to enhance ACh production and activity. Adequate dosing of the ingredients contained is hard to assess, but they appear to be in line with other formulas on the market with a similar purpose.  


Assessing Acetyl-Choline Brain Food impact on HRV

I’ve established that the formula has a combination of well-researched ingredients to positively impact ACh pathways and the ingredient doses recommended appear to be in line with industry peers.

Therefore, it's time to explore the effects the formula had on various quantitative and qualitative measures.  

Experimental design: I used one bottle or this Natural Stacks supplement (60 capsules, 3 capsules per serving) for 20 days. Prior to beginning the supplement, I tracked baseline HRV for one month using the Biostrap. I captured HRV measures throughout the experimental period as well as after discontinuation.


The graphic above shows baseline HRV data in the month prior to the experiment. Of note, my average HRV value during this time frame was 30 MS (Milli-Seconds).


The second graphic shows HRV data through the duration of the experimental period. The metrics show no change in baseline HRV levels. 

The 30 days following the experimental period also showed no change in HRV levels.  As such, it can be concluded that the use of this supplement at the suggested dose did not impart a measurable difference in my HRV.   

Surprise? Maybe! Let's look at the significance of that outcome now:


My Assessment Of Acetyl-Choline Brain Food

My n=1 experiment with this supplement did not produce a measurable response, nor did I notice any cognitive or mental clarity effects. Because the supplement is marketed as “Daytime Support for Mental Clarity”, I typically dosed the product between mid-morning and the noon-hour. 

Throughout the experimental period, no qualitative measures of mental performance were noted. 

Despite the lack of measurable outcomes, both quantitative and qualitative, I feel this is a good formula containing the right ingredients for those who want to support ACh neurotransmission.

The challenge, of course, is not having a tool available to assess for ACh deficiency, for whom this formula would be the right fit.        

Nevertheless, if you're one of these people who is eating a diet that's really deficient in acetylcholine precursors - such as eggs and liver - then Brain Food might be the right choice for you!

Of course, your experiments with this supplement can also end up differently than mine did. 


Finishing Thoughts: A Good Supplement That Didn't Work For Me

Natural Stacks has done a good job in assembling an ACh-supporting formula that may in fact deliver enhanced mental clarity for those in need of ACh support. 

As for my lack of response, it is possible that my system is not deficient in ACh and therefore any effect, whether mental clarity or the potential for HRV modulation, was not detectable.

It is also possible that the dose taken was not sufficient to meaningfully increase central ACh levels.

Furthermore, given the timing of the dose, it is possible that the ingredients were cleared from my system by the time HRV measurements commenced (Biostrap measures HRV throughout the night). Given these considerations, it is still possible that the supplement imparts HRV effects. 

Additional considerations: in the absence of the ability to measure ACh deficiency, assessing ACh supplementation on HRV would likely be best performed in individuals who experience positive responses in mental clarity while using this formula. The assumption being that if the supplement works for mental clarity, it may be due to ACh deficiency, and therefore, the potential to impact HRV via ACh is increased.

This investigation will continue, as there are more ACh-supporting supplements that warrant investigation as well as experimental designs to maximize the potential for impact. 

The bottom line is this: is Natural Stacks Brain Food a next-gen HRV booster? Not for me, but possibly for you!

And if you’ve read this far, I encourage you to consider measuring HRV while using ACh supplements and report back in the comment section. I’d love to hear if this idea moves the needle on HRV! 



This is a post by Joe Ailts. Joe has completed degrees in biotechnology (BS) and nutrition (MS) and is a science writer for Alexfergus. He has 14 years of experience in the clinical laboratory arena as well as in the dietary supplement industry.


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