Humidity, dew point and condensation: why cold vials attract moisture

The dew point is the temperature air must be cooled to, at the same pressure and moisture content, before it becomes saturated and water starts to condense (NOAA National Weather Service). That is why a vial taken out of a freezer mists over within seconds: the cold glass chills the air next to it below its dew point. The film of water comes from the air in the room, and if the vial is opened while it is still cold, some of it lands on the powder inside. This guide explains why it happens. It covers humidity and relative humidity, what the dew point is, a worked example with real numbers, and what that means for handling freeze-dried powders.

Terms are defined in the glossary.

What is humidity?

Humidity is the amount of water vapour in the air. Water evaporates from every damp surface, and the vapour mixes invisibly with the rest of the air. There are two common ways to express how much is there.

• Absolute humidity is the mass of water vapour in a given volume of air, usually in grams per cubic metre (g/m³).
• Relative humidity (RH) compares the water vapour present with the most the air could hold at that temperature, as a percentage. Air at 100% RH is saturated: it can't take up any more.

Relative humidity is the figure on weather forecasts and on most hygrometers, because it describes how close the air is to depositing water.

Warm air holds more water vapour

The most water vapour air can hold rises steeply with temperature. Worked out with the Magnus formula (Alduchov and Eskridge, 1996):

Air at 21 °C can hold almost four times as much water as air at 0 °C. So relative humidity depends on temperature as well as on how much water is present. Warm a sealed box of air and its relative humidity falls, though no water has left. Cool it and the relative humidity rises, until it reaches 100%.

What is the dew point?

The dew point is the temperature at which a particular parcel of air would become saturated if it were cooled without gaining or losing water. Cool air below its dew point and the excess vapour has to go somewhere: it condenses as liquid water on the coldest surfaces nearby. That is dew on grass at night, and mist on a cold glass.

The dew point is a better measure of how much water is in the air than relative humidity, because it doesn't change when the air warms or cools. A higher dew point means more moisture.

Dew point chart

Dew points at common room conditions, worked out with the Magnus formula below and given to the nearest degree:

Any surface colder than the dew point collects water from the air around it.

How to calculate dew point

The Magnus formula gives the dew point, T_d, from the air temperature T in °C and the relative humidity RH in percent (Alduchov and Eskridge, 1996; constants as given by Lawrence, 2005, via Wikipedia):

1. Work out γ = ln(RH / 100) + (b × T) / (c + T)
2. Then T_d = (c × γ) / (b − γ)

where b = 17.625 and c = 243.04 °C.

Worked example: a lab at 21 °C and 50% RH.

• ln(50 / 100) = −0.693
• (17.625 × 21) / (243.04 + 21) = 370.1 / 264.0 = 1.402
• γ = −0.693 + 1.402 = 0.709
• T_d = (243.04 × 0.709) / (17.625 − 0.709) = 172.3 / 16.92 ≈ 10.2 °C

So any surface below about 10 °C in that room will collect condensation.

Why condensation forms on a cold vial

Here is a worked example. A lab is at a typical room temperature of 21 °C and 50% relative humidity, so its dew point is about 10 °C.

• A vial from a fridge at about 4 °C is roughly 6 °C below the dew point. Water condenses on the glass as soon as it meets room air.
• A vial from a freezer at −20 °C is far below it. Where the glass is below 0 °C, the vapour freezes straight onto it as frost: the frost point is the dew point's name below freezing. As the vial warms, the frost melts into water.

The film keeps forming until the glass warms past the dew point. Then it starts to evaporate again.

In a humid room the problem is worse. At 21 °C and 70% RH the dew point is about 15 °C, so even a vial that has warmed part of the way can still be cold enough to collect water.

Why it matters for freeze-dried powders

The outside of the vial is not the risk. The risk is the powder inside:

• If a cold vial is opened, humid room air flows in and meets a cold, dry powder. Water condenses on the cake as well as on the glass.
• Freeze-dried peptides are hygroscopic: they take up water readily, even without condensation. A cold powder in humid air does both.
• Moisture leads to weighing errors, clumping and faster chemical breakdown over time. These are covered in hygroscopic powders.

How to avoid condensation

The advice in our other guides comes straight from the dew point:

1. Keep the vial sealed while it warms. A closed vial has only a tiny volume of air inside, so very little water can condense within it. Any condensation forms on the outside, where it does no harm.
2. Warm it inside a closed container, ideally a desiccator or a sealed box with a desiccant. The dry air inside has a low dew point, so the glass stays dry as it warms.
3. Give it time. Twenty to thirty minutes is the usual guide for a small vial. It needs longer from a freezer than from a fridge, and longer in a closed box than on an open bench.
4. Wipe the outside dry before opening, so drops can't run into the neck.
5. Open it briefly and reseal it promptly. See hygroscopic powders for weighing and resealing.

The same reasoning explains why vials are kept in a closed container inside a freezer or fridge. Every time the door opens, warm room air rushes in, and its moisture condenses or freezes on whatever is coldest.

Humidity in stability testing

Humidity matters to how long a material keeps, as well as when a vial is opened. The ICH stability guideline sets its standard storage conditions by both temperature and relative humidity. Long-term studies at room temperature use 25 °C / 60% RH, and accelerated studies use 40 °C / 75% RH (ICH Q1A(R2)). For a powder, the water it takes up can matter as much as the heat. See the Arrhenius equation and accelerated stability testing.

Measuring humidity

• Hygrometers measure relative humidity. Most modern ones use a small electronic sensor whose electrical properties change as it absorbs water. Cheap ones can be several percentage points out, so treat their readings as a guide.
• Temperature and humidity data loggers record both over time, which shows how the conditions change through the day or when a door is opened. See freezer temperatures for how loggers are used.
• Humidity indicator cards change colour at set humidity levels. They are cheap and useful inside a sealed box of vials. See desiccants and silica gel.

Typical indoor humidity

Indoor humidity varies with the season, the heating and the building. Heated rooms in winter are often fairly dry, and rooms in summer, or near sinks and steam, are damper. A dew point table like the one above shows why that matters: the damper the room, the warmer a vial must be before it can be opened without collecting water.

Related guides

For handling, weighing and resealing vials, see hygroscopic powders. For why a dry powder can travel without cold packs, and what to do when a parcel arrives, see why lyophilised peptides ship at room temperature. For storage conditions, see how to store freeze-dried peptides. Every lot has its own COA, published on the product page and on that lot's own page. See Quality.