Creating your own buffer for unbuffered NAD+

Sera

Member
Have any NAD+ that was unusable because it was unbuffered, and therefore like injecting lemon juice? Well with this guide you can turn that into a superior product with virtually zero discomfort.

I’m going to explain what I did and you can adapt the steps to your own set up as needed.

You will need:
1 30mL bottle bacteriostatic water.
sodium phosphate monobasic
sodium phosphate dibasic
pes sterile syringe filters 0.2micron
5-10ml Syringes
27g or similar needles
0.001 Milligram scale
Weigh boats ideally
Sterile 5-10ml vials
Lyophilised NAD+ (in my case it was 600mg.)

I recommend hydrated forms of these salts as it makes the masses more accurate as anhydrous versions can absorb varying amounts of moisture from the air, making the calculations less accurate.
Whichever form you get, you can use an AI tool to calculate the masses of each salt needed. You want to create an isotonic buffer (with no added osmotic agent such as NaCl) at 6.5pH with 30mL of water so just ask it for that and specify the exact forms you have.

I used anhydrous anyway (didn’t think of the moisture issue when I ordered it) so that was about 420mg of monobasic and 100mg of dibasic.

I weighed this with a milligram scale and a weigh boat. I then added the powder to the inside of a syringe barrel with a needle attached, and drew water from the bac water bottle into the syringe to dissolve the powder.

I then emptied the syringe into the bac water bottle and let it mix thoroughly.

This is the buffer ready, so use it to reconstitute a vial of nad+. In my case I needed 6ml to buffer 600mg of nad+ you may need more or less depending on how much nad+ you have per vial. Seems reasonable to assume 1ml per 100mg.
Use the 3ml vial and the syringe to hold the amount of buffer you need.

Prepare your filter by rinsing a ml or two of normal bac water through it before adding a needle to the end. Then swab your sterile vial with an alcohol swab, allow to dry, insert a 27-30g vent needle off centre in the stopper and then insert the filter needle on the opposite side of the vial also off centre.

Insert your syringe into the filter ensuring no air bubbles get trapped in the filter as these will greatly slow the pace of filtration. You can pull back on the plunger if any get stuck to draw them out and back into the syringe.

You’ll want to do this every time as the bac water buffer you made isn’t sterile. I think this should be fine given it contains benzyl alcohol and you will be filtering it.

However on reflection I might recommend to the reader to be on the safe side:
Filter the phosphate salts into the bac water bottle instead of just adding it straight. This would protect against any theoretical microbe growth that might happen in the bottle.

Anyway, after doing this a moment ago and injecting some to try it out, I can confirm it works with basically zero discomfort subcutaneously.
 
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Update: I got a little carried away here, it turns out the recipe didn’t really work. pH is still low and didn’t manage to increase to the target.
At first the pain really wasn’t noticeable so maybe it helped a little?

Back to the drawing board.
 
Update: I got a little carried away here, it turns out the recipe didn’t really work. pH is still low and didn’t manage to increase to the target.
At first the pain really wasn’t noticeable so maybe it helped a little?

Back to the drawing board.

PBS pH 6.5.
Would this not work?
 
Nah you gotta take that buffered mess (pH ~2.8) in an aqueous solution up to at least pH of 6 (with dilute NaOH drop wise) before adding phosphate buffer or your just spinning your wheels.

PBS pH 6.5 10x?
I've never used NAD+ so I'm not exactly sure, but we just want a buffer right?
 
Update:

250mg dibasic on it’s own brought the NAD+ to 6.5pH. Makes me wonder if 300 (half of the mass of the NAD+) would neutralise it and bring to pH 7.

The only problem with this technique is that it leads to a hypertonic solution which is a little uncomfortable to inject and you need to use it in the muscle to avoid lumps. A little bit like l-carnitine (another very hypertonic solution, except that l-carnitine from what I understand is usually a lot more hypertonic).

According to the patent, the next method I will try will likely result in an isotonic solution, suitable for use subq.
 
Simple NaOH works fine for PH adjustment. Aim for PH in 5.5 to 6 range. Buffering could be useful if you wanted to create a pharma/commercial grade product- for personal use it’s over kill.

NAD doesn’t last long in fridge, use with in 4 weeks or freeze for longer storage.
 

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