Steam Sterilization Cycles, Part 2: Liquids

Following our first post about Steam Sterilization Cycles—where we addressed Gravity vs. Vacuum Cycles—this second article discusses the Liquids Cycle.

Just about every lab must sterilize some type of liquid solution, such as lysogeny broth (otherwise known as LB Broth), media, agar, buffer, saline, and water.  And as such, sterilization of a liquid in a vented container requires a special type and the proper application of a cycle known as the Liquids Cycle.

The Liquids Cycle

Liquids rely on the Liquids Cycle to avoid a phenomenon known as “boil-over.” Boil-over is simply a liquid boiling so violently that it spills over the top of its container.  Boil-over will occur if the pressure in your autoclave chamber is released too quickly during the exhaust phase of the cycle.  Significant liquid volume can be lost to boil-over, and this can result in unwanted spills on the bottom of your chamber and subsequent cleanup.

To help prevent boil-over during the exhaust phase, the chamber pressure must be released slowly.  This process is controlled by the sterilizer’s control system.  Controlling the exhaust rate allows the liquid load to cool off as the surrounding chamber pressure is decreased.

Be sure your autoclave is programmed and equipped with the right software and hardware to handle a slow exhaust Liquids Cycle.  Quite often, a Jacket Blowdown Option will be supplied to assist with minimizing boil-over.  During a Liquids Cycle, this option automatically exhausts the steam from the jacket (which surrounds the chamber) and thus allows heat to be drawn out of the chamber and the liquid load at a faster rate. The exhaust rate for a Liquids Cycle is different from a standard Gravity or Vacuum Cycle, where the chamber pressure is released quickly.

NOTE: It is important to recognize that larger liquid loads will take longer to both heat up and cool down. See the chart below:

The autoclave operator should keep this in mind, especially for large beakers or carboys.  For instance, a 30-minute Liquids Cycle for a 500mL flask won’t necessarily achieve the same results if sterilizing a 5 L flask.  It is a best practice to A) validate your liquid loads with hermetically sealed biological indicators and B) minimize container volumes so that cycle times remain manageable.

Now you know what causes boil-over and how to avoid it when running a Liquids Cycle in your autoclave. However, while the Liquids Cycle prevents boil-over, it may also allow a small percentage of the liquid load to evaporate. So, when sterilizing very small volumes, it becomes imperative to minimize or eliminate evaporation.

To mitigate evaporation, more advanced sterilization cycle applications may be necessary. Please stay tuned for our next article in our Steam Sterilization Cycles series, where we will discuss the Air-Overpressure Cycle.

To learn more about the Liquids Cycle, check out our video here:

What causes a liquid to boil over?

Well, let’s start by thinking about boiling a pot of water. At sea level, water boils at 212˚F (100˚C). In Denver, CO, however, where the elevation is 5,280 ft. above sea level, water boils at 203˚F (95˚C). But why is there a difference in boiling temperature? The answer is in the elevation, and the amount of atmospheric pressure pushing *down* on the water. At sea level, the atmospheric pressure is only 14.6 psia (equivalent to 0 psig or 0 gauge pressure), while Denver’s atmospheric pressure is only 12.1 psia. This relationship between pressure and temperature is what determines the boiling point of water; not just its temperature.

Now, let’s think about the conditions inside an autoclave. During sterilization, a liquid load is heated to a temperature of 250˚F (121˚C). This temperature can only be achieved if the liquid load is subjected to steam pressure. During the sterilization phase, an autoclave chamber typically operates at a pressure of 15 psia above atmospheric pressure (roughly 29.6 psia). This additional pressure is just enough to keep the liquid load from boiling. Upon completion of the sterilization cycle, the chamber must exhaust the steam, which means the temperature and the pressure in the chamber will decrease. To prevent boil-over, the chamber pressure must decrease slowly to allow the temperature of the load to remain below the boiling point. If the pressure is exhausted all at once, the temperature of the load will be above its boiling point, resulting in instant and violent boiling.

While you wait for our next article in this series, we encourage you to ask us any questions you may have. Contact us today.