Bubble Point Testing
Updated July 17, 2023
PLEASE do this step to ensure your preparation is sterile. It requires no additional tools.
At this point you should have one or more 100mL vials with filtered product. Before we go through the hassle of putting everything in vials, let’s make sure the filters held up during filtration!
We rely primarily on the syringe filter to ensure the sterility of the end product. While people will speak at length about other ways to sterilize DIY brews, filters are the only scientifically validated method of sterilization and depyrogenation of your preparation that are available for you in a DIY lab.
For this reason, we urge you to conduct a bubble point test to ensure that your syringe filter maintained integrity throughout your filtration process. Bubble point testing does not require any additional tools beyond what you already have on hand, and it takes only a minute to conduct plus some light math that we are going to help you with. This is a test we do on the syringe filter(s) we used to prepare our vials. If you did a 2nd pass filtration for sterility testing do not conduct a bubble point test on any 2nd pass filters.
- Find the bubble point of your syringe filters,
A. It should be listed on the package or on the product’s webpage. You can also use a new filter soaked in MCT oil and this methodology to calculate it. - Find the current air pressure at your location using today’s weather, convert from inHg to PSI,
B. - Remove the syringe filter from the syringe you were filling with, and then fill that syringe (or a new one) with air to an exact measurement,
C, e.g. 10.0mL. Reattach the syringe filter. Attach a large gauge needle to the filter. - Submerge the needle in a cup of tap water. Slowly exert pressure onto the plunger, attempting to push air through the filter. When the needle starts emitting a steady stream of air bubbles record the volume marked on the syringe,
D. - You should have the below values:
A - Rated bubble point pressure B - Current air pressure C - Starting volume D - Actual bubble point volume
Go to the Boyle’s Law Calculator and input your values. Initial pressure is B. Initial volume is C. Final volume is D. Make sure you’re using the correct units of measurement.
The calculator will give you a value for final pressure, call this E - actual bubble point
How does the rated bubble point of your filter, A, compare to the actual bubble point you calculated? If the actual bubble point is lower than the rated bubble point, you need to ask if your filter failed during filtration.
For example, if our filter is rated to 86psi, and our actual bubble point was 77psi, we’re going to consider the filter functional. It withstood a lot of pressure, and we’re working with only estimates here. Laboratory bubble point tests use more sophisticated methods of measurement.
On the other hand, if our filter is rated to 86psi, and our actual bubble point was 40psi, then this is a sign that there was a major failure with the filter during use. It is likely that a significant amount of medicine did not properly pass through the membrane.
If your filter failed, it might be worth running an experiment on a new filter from the same batch to determine where it’s bubble point is (once the filter is saturated in oil). It’s (unlikely but) possible the batch was incorrectly labeled
If we determine that our filter failed during use, we will redo all of our work. Working in the still air box or laminar flow hood we will remove all the mixture from all the vials with a syringe and needle. We will use a new syringe and new syringe filter and re-filter everything into NEW sterilized and depyrogenated vials.
HOWEVER, some filters will fail due to chemical degradation. There is always a chance that if this happened that chemicals from the filter leeched into your medicine. It’s unclear if another round of filtration would remove that. For this reason we recommend only PTFE filters or nylon filters if PTFE is unavailable. These are the only filter membranes that have a studied compatability with Benzyl Benzoate.